Introduction to Medical Parasitology

Wednesday, 6 January 2016 3:45 PM

Key terms
1. Symbiosis Parasitology: is the science of study of host-
○ Living together/ close association of 2 dissimilar organisms parasite relationship
○ There are 3 forms of symbiosis: Medical parasitology is the study of parasites
 Mutualism: symbiosis in which both parties benefit which infect humans
 Commensalism: symbiosis in which one party is benefited while the other party (the Parasite: a living organism that lives upon
host) receives neither benefit nor harm another living organism at whose expense, it
 Parasitism: symbiosis in which one party (parasite) benefits at the expense of the gains an advantage
other (host)
2. Parasitism
○ A parasite is successful when it is in a delicate balance with the host. If the balance is lost,
the host may destroy or expel the parasite, or if the host is overly damaged, both may die
3. Host
○ Definitive host
 Either harbours the adult stage of the parasite where the parasite utilises the sexual
method of reproduction
○ Intermediate host
 Harbours the larval/asexual stages of the parasites e.g. man for malaria
 In some cases, larval developments are completed in two different stages
□ Cysts
□ Trypomastigotes
○ Paratenic host
 Not obligatory to lifecycle
 Larvae do not mature or reproduce in these hosts
 Can infect definitive, accidental or other paratenic hosts
4. Zoonosis
○ A parasitic disease in which the animal is normally the host - but which also affects man
5. Vector
○ A living carrier that transports a pathogenic organism from an infected to non -infected
organism or host
○ Typical example: anopheles mosquito

The basic factors of transmission

a. Source of nutrition
b. Transmission
c. Susceptible host
• The combined effects of these factors determine the distribution and the prevalence of the
parasites at a given time and place and regulate the incidence of the parasitic diseases in
populations

Modes of transmission
• Contact
• Food
• Water
• Soil (soil transmitted helminths [STH])
• Vector
• Congenital (via pregnancies)

Life cycle of a parasite

A life cycle is the whole process of a parasite growing and developing
1. Direct life cycle: parasite has only one host (no intermediate host)
2. Indirect life cycle: parasite's life cycle that involves more than one host

How parasites cause harm to the host

1. Trauma: damage to the cells, tissue and organs
2. Lytic actions: enzymes elaborated by organisms
3. Host responses: localised inflammation, eosinophilia
4. Blood loss: heavy infection with hookworm may cause anaemia
5. Secondary infections: weakened host is susceptible to bacterial infection
6. Toxin production

Classification of Parasites

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Classification of Parasites
• Taxonomic
• Morphologic
• Physiologic
• Mode of transmission
• Location/site in host
• Combinations of any of the above

Taxonomic
• Classified into 2 sub-kingdoms i.e.
○ Protozoa (unicellular): classified according to morphology and means of locomotion
○ Metazoa (multicellular): helminths and arthropods

Nomenclature of parasites
• Each parasite possesses 2 names: a generic and specific name
• Genomic name starts with a capital letter, specific names start with a small letter

Protozoa
• For medical purposes, protozoa can be sub-divided into 4 groups:
a. Sarcodina
b. Sporozoa
c. Mastigophora
d. Cilliata

Sarcodina
• Typically amoebid and include entamoeba, endolimav, iodamoeba
• Morphology
○ Shapeless mass of moving cytoplasm divided into clear and granular ectoplasm
• Motility
○ Move by pushing out ectoplasm to form pseudopodia (false feet)
• Asexual reproduction by binary fission

Mastigophora (flagellates)
• Have one or two flagella for locomotion
• Reproduce through binary fission (asexual)
• Cause diseases in many body parts

Apicomplexa (sporozoa)
• Non-motile
• Reproduce both sexually and asexually

Ciliophora
• Have cilia for locomotion

Helminths
• Worm-like parasites
• Divided as follows:
○ Metazoa
 Nematodes
 Platyhelminths
□ Cestodes
□ Trematodes

Nematodes
• Body is elongated, cylindrical and unsegmented
• Appear round in cross section
• Have body cavities
• Have alimentary canals
• Sexes are separate
• Examples
○ Intestinal
 Small intestines
 Caecum and appendix
○ Somatic (inside the tissues/organs)
 Lymphatic system
 Subcutaneous tissue

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  Subcutaneous tissue
 Conjuctiva

Cestodes
• Tape-like, segmented (proglottids)
• Consists of a head (scolex) with sucking organs
• No alimentary canal
• Body is segmented
• Each segment is hermaphrodite eg taenia

Trematodes
• Leaf like, unsegmented, no distinct head
• Alimentary canal is incomplete (no anus)
• Have 2 suckers
• The body cavity is absent
• Sexes are not separated
○ Schistosomes are an exception. Are thread-like and have separate sexes

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 Enterobius Vermicularis
Wednesday, 9 December 2015 8:18 PM

 Intestinal nematode, adult found in the large intestines

 Also known as pinworm
 Worldwide distribution, more in temperate climates.
 Most common helminth in US
 More common in overcrowded areas
 Children most affected – high prevalence and intensity
 Causes enterobiasis

Morphology
 Adults
o Females 9-12mm
o Males 2-3mm
o Thick lips (cervical alae)
 Eggs
o Planoconvex in shape
o Thin Transparent wall
o Embryonated

Life Cycle
 Adults mate in the large intestines, females get out of the anal area at night, lay eggs
 Eggs passed embryonate within a few hours (6 hours)
 Itchiness of perineum (anal pruritus), scratching, stick to fingers and nails, ingested
accidentally (self-infection or others), reach small intestines, hatch and larvae move to
large intestines, attach to the mucosa and mature to adults
 May be transmitted through clothing, bedding, food, etc
 Contaminated by eggs, dustborne by inhalation, retroinfection
 Life cycle takes 4-6 weeks

Clinical Presentation
 Light infection mostly asymptomatic
 Clinical presentations include:
o Pruritus ani usually at night
o Sleep disturbance
o Irritability and lack of concentration in school
o Poor class performance
o Loss of appetite
 Complications include:
o Appendicitis
o Salpingitis in females (rare)

Diagnosis
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 Diagnosis
 Microscopic identification of eggs must be done in the morning before defecation and
washing by:
o Adhesive scotch tape – press transparent adhesive tape on the perineal skin and then
examining the tape under a microscope
o Anal swabs – press a swab coated with adhesive material on the perianal skin
o Camel hair brush
 Eggs occasionally found in stool, urine, vaginal smears
 Adult worms may also be found in perianal area

Treatment
 Mebendazole
 Albendazole
 Pyrantel palmoate
o In institutional outbreak, treat all household members

Prevention and Control

 Proper hygiene
o Daily bathing
o Frequent changing of clothes and beddings
o Keeping short nails
 Avoid overcrowding, opening of windows for ventilation e.t.c
 Health education
o Washing of hands before eating of a meal
 Mass Treatment of infected persons with anti-helminthes

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 Trichinella Spiralis

• Trichinellosis (trichinosis) is caused by nematodes (roundworms) of the genus

Trichinella.
• Is a tissue nematode – especially in muscles – because of rich oxygen supply
• Worldwide distribution. Most common in parts of Europe and the United States. More in
areas where pork is eaten

Morphology
• Adults
• males (1.5mm)
• female (4mm)
• Larvae - encapsulated in human muscle

Life Cycle
• Reservoirs – Rats, pigs, and wild animals such as wolf, seals, bears and polar bears in
the Arctic, and warthogs, hyenas, jackals, lions, and leopards in the Tropics
• In the tropics, life cycle between rats and pigs/warthogs, man ingests, man ingests
uncooked infected meat (pork).
• The larvae develops to adults, mate, female produce larvae, penetrate intestines into
blood throughout the body, encapsulate in muscle (skeletal, tongue, diaphragm etc)
and >6 months may calcify leading to death of the larvae

Clinical Presentation
 Incubation period 10-14 days
 Presentation is variable:
o Oedema of the upper eyelid
o Subconjuctival haemorrhage
o Photophobia
o Diarrhoea
o Myalgia – muscle pain
o Generalised weakness
o Fever
o Skin rash
o Finger clubbing
 Complications
o Cardiac failure
o Respiratory failure
o Meningitis
o Meningoencephalitis
o Fits and comas in long infections

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 Diagnosis
 Muscle biopsy – encapsulated larvae
 Serology
o ELISA test
o Complement fixation test (CFT)
o Indirect haemogluttination (IHT)
 Blood – eosinophilia (hallmark of infection)

Treatment
 Albendazole, mebendazole, thiabendazole – useful during early stages of infection
 Steroids – in heavy infection
 Surgery – when body undergoes fibrosis as a defense mechanism in infected parts

Prevention and Control

 Meat (pork) inspected in slaughter houses
 Cooking of garbage and offal before feeding pigs
 Avoid eating warthogs
 Proper cooking of meat (pork). Freezing of meat if needed to be eaten raw (-25 degrees
celsius for 7 days)
 Health education

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 Trichuris Trichuria
Wednesday, 9 December 2015 8:21 PM

 Intestinal nematode, found in the large intestines

 Also known as whip worm
 Worldwide distribution, mostly in the tropics
 Most common in poorly sanitised areas
 Children mostly affected – highest prevalence and intensity
 Causes Trichuriasis

Morphology
 Adults 30-45mm long
 Resembles a whip with slender anterior part
 Greyish white in colour

Life Cycle
 Adults mate in large intestines
 Eggs passed in stool are not infective
 Embryonation takes place in the soil
 Ingested accidentally – contaminated hands and food
 Eggs hatch in the large intestines
 From ingestion of eggs to adult – 3 months
 Adults mate in the large intestines, eggs passed in stool

Clinical presentations
 Light infection mostly asymptomatic
 In heavy infection:
o Abdominal discomfort
o Abdominal pain
o Diarrhoea and bloody stool
o Finger clubbing
 Complication
o Rectal prolapse
o Anaemia
o Growth retardation

Diagnosis
 Stool examination for characteristic eggs
Direct smear

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 o Direct smear
o Stool concentration methods
 Sigmoidoscopy
o Adult worms attached to mucosa

Treatment
 Albendazole
 Mebendazole

Prevention and Control

 Proper faecal disposal – pit latrines
 Don’t use untreated human faeces as manure
 Health education on how infection is transmitted
 Proper hygiene
 Treatment of infected persons with antihelminths

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Lymphatic Filariasis
Thursday, 29 September 2016 9:43 AM

• Filariasis caused by filarial worms

○ Are tissue nematodes
• Classification of filariasis based on the final habitat of adult worms in humans
• LF caused by filarial tissue nematodes
• Wuchereria bancrofti, Brugia malayi and Brugia timori
• Adults inhabit the lymphatic system, microfilariae in blood
• 120M infected, >40M with clinical disease globally
• Geographical distribution
○ Asia: China, India, Malaysia, Philippines
○ Africa: East African coast
○ Central and South America
○ Pacific islands
• Associated with warm, humid climates
• Found in both urban and rural communities
• Age-specific prevalence rises with age
• Gender differences associated with cultural practices resulting in differential exposure to infection
• Vector borne
○ Transmitted by different mosquito species (aedes, anopheles, culex, mansonia)

Morphology
• Adults
○ Thread-like
○ Females longer than males
• Microfilariae (W. bancrofti)
○ Sheathed
○ Nuclei clearly separated, do not extend to the tip of the tail
• Microfilariae (B. malayi and timori)
○ Sheathed
○ Nuclei are matted, 3 nuclei extend to the tip of the tail

Life cycle
• Adults mate, fertilised female produces microfilaria
• Mf in blood taken up by mosquitoes (different spp)
• Mf have nocturnal periodicity (10:00pm - 2:00am) but diurnal strains occur in some parts of the world
• Mf penetrate stomach wall, migrate to flight muscles, transform to L3, then to proboscis
• In flies, development to infective stage (L3) ~10 days
• L3 deposited in bight, gets into blood then lymphatics and develop to adults
• Adults mate and produce Mf which get into blood

Clinical presentation
• Early stages (acute manifestations)
○ Fever, chills
○ Lymphadinitis, lymphangitis
○ Orchitis, epididymitis
• Late stages (chronic manifestations - blockage of lymphatic flow)
○ Lymphoedema of limbs and breasts
○ Elephantiasis (limbs, breast, scrotum)
 Secondary fungal and bacterial infection

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  Secondary fungal and bacterial infection
○ Hydrocele
 Fluid in the tunica vaginalis
○ Chyluria (Whitish urine)
○ Tropical pulmonary eosinophilia (TPE)
 Hypersensitivity reaction - presents like an asthmatic attack
□ Dyspnoea
□ Dry cough
□ Wheezing
□ Transient hepatomegaly and splenomegaly

Diagnosis
• History of living in the endemic areas of years
• Physical examination findings
• Blood examination (night sample) for microfilariae
○ Thick blood smear
○ Capillary tube exam
○ Blood filter (nucleopore)
 Stain with Giemsa stain for analysis
 Fields stain*
• DEC provocation test (DEC 5-mg, blood >30 min)
○ Diethylcarbamazine*
○ Irritates the microfilariae to retreat to blood during the day
• Ultrasound of the scrotum - filarial dance sign
• Serological tests
○ Filarial specific antibodies e.g. IgG, circulating filarial antigen (CFA)
○ Does not ascertain current infection, exposure or post-exposure

Treatment
• DEC
○ Should not be used in places where there is onchocerciasis occurring together with LF
• Ivermectin
• Albendazole
○ Combination Rx with DEC or Ivermectin

Prevention and control

• Vector control
○ Insecticides
○ Larvicides
• Personal protection
○ Bed nets
○ Insect repellents
• Annual treatment - DEC or Ivermectin
• DEC treated salt

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Onchocerca volvulus
Thursday, 29 September 2016 10:21 AM

• Tissue nematode, found in subcutaneous tissues

• Found in West Africa, Parts of East Africa, Middle East
• Adults found in subcutaneous nodules
• Microfilaria found in the skin, but sometimes appear in blood and eye in heavy infections
• May be found in urine, sputum and CSF especially after treatment
• Cause onchocerciasis

Morphology
• Adults
○ Cylindrical worms
○ Females: 30 - 50 cm
○ Males: 20 -40 cm
• Microfilaria
○ Unsheathed
○ 300~320 µm in length
○ Tail tapers to a point
○ Nuclei do not extend to the tip of the tail

Life Cycle
• Adults mate and fertilised female produces microfilariae
• Mf in skin taken up by the Simulium flies
• Mf penetrate stomach wall, migrate to flight muscles then proboscis (in L3 stage) in ~10
days
• L3 deposited in bite, develop into adults in s/c nodules
• Worms mature and produce Mf
○ Mf mature in 9-12Months and adult live for 10 - 15 years
• Other possible modes of transmission
○ Congenital transmission

Clinical Presentation
• Incubation period 15 -18M may be asymptomatic
• Pruritus of the skin, rash on skin
• Subcutaneous nodules (onchocercomas) in bony prominences. In Africa, normally around
the hips and the head in S. America
○ Due to larvae mobilisation in these sites to be removed by the immune system
• Lymphadenitis especially in the groin
• Lizard skin - Thickened skin
• Prespyderma
○ Loss of skin elasticity, premature ageing
○ Due to reaction dead microfilariae around S/c nodules
• Leopard skin
○ Skin depigmentation in patches
• Hanging groin
○ From presbyderma; the microfilariae cause inflammation of regional lymph glands
• Arthritis
○ May be monoarthritis affecting the large joints e.g. hip pain disappears rapidly on
treatment
Dead microfilariae cause inflammation

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 ○ Dead microfilariae cause inflammation
• Elephantiasis of the scrotum in males
• River blindness (onchocerciasis)
• SOWDA
○ Hypersensitive reaction to microfilariae
○ Arabic for dark
○ Results from strong immune response by host
○ Usually localised to one limb
○ Itchy, swelling, dark with scaling papules, enlarged regional lymph nodes

Diagnosis
• Skin snip(thigh, buttocks, scapula, iliac crest), place in normal saline, examine after 12-24
hours for Mf in wet preparation then stain e.g. Giemsa and examine for features
• Mf negative from skin snips in SOWDA
• Mazzotti skin test (50-100 mg DEC), intense pruritus 2-24 hours later (contraindicated in
heavy infection)
○ May cause severe anaphylactic reaction
• Serological tests
○ ELISA
○ Fluorescent Antibody Technique (FAT)
• Eosinophilia (non-specific)

Treatment
• Ivermectin single dose
• Nodulectomy (surgical removal of nodules)

Prevention and Control

• Vector control
○ Larvicides
○ Aerial insecticide spray
• Personal prophylaxis
○ Yearly Ivermectin
• Nodulectomy
○ Been useful in reducing incidence of blindness in Mexico and Guatemala
• Migration of communities from endemic areas

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Loa Loa
Thursday, 29 September 2016 10:52 AM

• Tissue nematode, found in the subcutaneous tissue

• Found in West and Central Africa
• Adults found in subcutaneous tissue
• Causes Loiasis
• Transmitted by Chrysops fly

Morphology
• Adults
○ Threadlike
○ Females 5-7cm
○ Males: 3-4cm
• Microfilariae
○ Sheathed
○ 200-250µm in length
○ Nuclei extend to the tip of the tail
○ Tail tapers to a point

Life Cycle
• Adult mate, fertilised female produces microfilariae
• Mf taken up in blood by Chrysops flies
• Mf have diurnal periodicity (8:00am to 8:00pm)
• Mf penetrates the stomach wall, migrate to flight muscles and then to proboscis
• In flies, develop to infective stage and takes ~10 days
• L3 deposited in bite and develop in s/c tissues
• Takes 5-6 months for the worms to mature and produce Mf
• Adults can live up to 17 years
• When not in circulation, they are found in the lungs

Clinical Presentation
• Incubation period may be several years
• Usually asymptomatic
• Migration of adult worms across conjunctiva
• Pruritus of the skin
• Calabar swellings
○ Transient, Migratory, Painless swellings
○ May occur at regular intervals
○ More frequent in hot weather
• Others, rare, in heavy infection
○ Meningoencephalitis (e.g. after DEC treatment)
○ Choroidoretinitis
○ Arthritis

Diagnosis
• History of migrating worm in the eye, or of calabar swellings
• Mf in blood smear
• Serological tests
○ ELISA

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 ○ ELISA
○ FAT
• Adult worm extraction from conjunctiva
• Eosinophilia, non-specific

Treatment
• Ivermectin

Prevention and control

• Environmental
○ Houses far from forest edge, swamps
• Vector control
○ Larvicides
○ Insecticides
• Personal protection
○ Insect repellents
○ Protective clothing

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Mansonella (M. streptocerca, M. perstans, M. ozzardi)
Thursday, 29 September 2016 11:02 AM

• M.streptocerca found in Africa

• M.perstans found in both Africa and S. America
• Ozzadi occur only in America, from Mexico south, to S. America and in the Caribbean

Morphology
• M.streptocerca
○ Adults ~27mm long, ~50µm wide
• Perstans
○ Adults ~70-80mm long and ~120µm wide
• Ozzardi
○ Adults ~65-81mm long and 0.21-0.25mm wide
• All mansonella microfilariae are unsheathed and unperiodic

Life cycles
Perstans
• Infected Cullicoides introduces L3 larvae into the skins
• Penetrate through the bite wound
• Develop in cavity (pleural or peritoneal cavities, rarely in pericardial) to adults
• Adults produce Mf that reach the bloodstream
• Cullicoides ingests the Mf from the blood meal
• Mf migrate from the midge's midgut to the thoracic muscles and develop into L1 larvae
and then infective L3 then move to the proboscis and can infect another human when the
fly takes another blood meal

Streptocerca
• Cullicoides spp introduce L3 into skin of human host
• Penetrate bite wound and develop into adults that reside in the dermis
• Adults produce Mf which reside in the skin but can also reach peripheral blood
• Cullicoides ingests the Mf during blood meal, the Mf migrate from midgut to thoracic
muscles., develop to L1 and then subsequently, L3
• L3 then move to the proboscis and can infect another human when the fly takes another
blood meal

Ozzardi
• Infected cullicodes or Simulium flies introduces L3 larvae onto skin during blood meal
• L3 penetrate into bite wound and develop into adults in s/c tissue
• Adults mate and produce Mf that reach the blood stream
• Vectors ingest Mf during blood meal, Mf migrate from midgut to thoracic muscles, develop
into L1 larvae then to infective L3
• L3 then move to the proboscis and can infect another human when the fly takes another
blood meal

Clinical presentation
• Perstans often asymptomatic but associated with angioedema, pruritus, fever, headaches,
arthralgia and neurologic manifestations
• Streptocerca causes skin manifestations including pruritus, papular eruptions and
pigmentation changes
• Ozzardi can cause arthralgia, headaches, fever, pulmonary symptoms, adenopathy,
hepatomegaly and pruritus

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 hepatomegaly and pruritus
• Eosinophilia often prominent in all filarial infections

Diagnosis
• Perstans and Ozzardi diagnosed by finding Mf in blood
• Streptocerca usually diagnosed by finding Mf in skin snips
• Skin snips are placed immediately in normal saline or distilled water and Mf emerge in
30 -60 min and can be seen on wet mount preparations
• For definitive diagnosis, allow wet mount to dry, fix in methanol, stain with Giemsa or
haematoxylin-and-eosin (H&E)

Treatment
• Perstans
○ In areas with Wolbachia containing strains e.g. Mali, use doxycycline
○ In areas without Wolbachia containing strains, use DEC in combination with
mebendazole
• Streptocerca
○ Ivermectin
• Ozzardi
○ Ivermectin
○ There is evidence that Ozzardi contain Wolbachia, so doxycycline might be an
effective treatment
• DEC not useful for streptocerca or ozzardi

Prevention and Control

• Vector control
○ Larvicides
○ Insecticides
• Personal protection
○ Insect repellents
○ PPE

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Drancunculus Medinensis
Thursday, 29 September 2016 11:26 AM

• Tissue nematode, also found in the subcutaneous tissues, also known as Guinea Worm
• Occurs predominantly in tropical countries: previously endemic to the Nile Valley, central
and western Africa parts of Eastern Africa, India Pakistan and Iran
○ East Africa - Northern Kenya, Southern Sudan
• Disease occurs in well defined areas within these regions
• Essentially a disease of rural farming communities whose drinking water are ponds and
shallow wells
• Causes Dracunculiasis

Morphology
• Adults
○ Cylindrical worms
○ Females are 50-120cm and are larviparous
○ Males are 2-3 cm long and die in human tissue soon after mating so are rarely found
• Larvae
○ ~700µm in length
○ Rounded head
○ Posterior end that is tapered into a long pointed tail

Life Cycle
• Adults mate within the abdominal cavity, male dies, gravid female moves to s/c tissue -
mostly legs and feet
• Gravid uterus of females protrudes on skin forming a blister
• In exposure to water, blister ruptures to release first stage larvae, continues to release
larvae for 2 weeks on re-exposure to water (larvae can live for ~3 weeks in water
• Larvae eaten by copepod (small crustacean) - intermediate hosts - develop into infective
3rd stage larvae in 14 days
• Ingested in drinking water, copepod ingested, digested and released larvae penetrate
intestinal wall, into abdominal cavity and retroperitoneal space to adults (~1 year)

Clinical Presentation
• Incubation period ~1 year
• Abdominal pain
○ Due to adult worms in peritoneal cavity
• Few days before blister eruption, systemic allergic reaction lasting about 24 hours,
generalised skin rash, pruritus, fever dizziness, dyspnoea, NVD
• Blister
○ Usually lower limbs (ankles and feet)
• Painful, ruptures within 5/7 spontaneously or on contact with water
○ Within resulting ulcer, anterior end of worm may be seen, subsequently complete
spontaneous emerging over 3-6 weeks
• Complications
○ Secondary bacterial infection
 Tetanus, abscesses, chronic ulcers, cellulitis, gangrene, septicaemia and
septic arthritis (joint near ulcers)
○ Dying unfertilised worms may form deep abscesses or calcify

Diagnosis
• Rupture of blister on emersion in water
• Recognition of emerging adult worm in skin ulcers
•

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 •
• Dead calcified worms may be seen on abdominal x-ray

Treatment
• Traditionally manual extraction of adult worm from the ulcer by rolling them out a few
centimetres each day on a small stick
• No curative anti-helminthic available
• Chemotherapy
○ Reduces inflammation allowing worm to be removed easily
 Metronidazole, thiabendazole, niridazole
• Surgical removal of the adult worm + antibiotic cover, tetanus toxoid

Prevention and control

• Health education on how disease is transmitted
• Avoid wading in waters that are drinking sources
• Provision of safe water e.g. piped, cover or build walls around wells
• Boiling drinking water, water filtration, chlorination
• Application of chemicals to kill copepods in water
• Biological control
○ Gambusia fish feed on copepod

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Cestodes
Thursday, 29 September 2016 11:54 AM

• Cestodia
○ Pseudophyllidea - have sucking grooves (bothria)
 D.latum
 Spirometra
○ Cyclophyllidea - suckers on scollices (acetabulum)
 Taenia
 Echinococcus
 Hymenolepsis
 Dipylidi

General characteristics
• These are tape-like, with segmented body (proglotids)
• Each body segment is hermaphrodite
• Consists of a head (scolex) with sucking organs
• No alimentary canal
• All have a definitive host and one or more intermediate hosts
• Absorb nutrients from the host through their skin
• Need lipids for reproduction but unable to synthesise lipids, obtain these from the host

Morphological Forms
• Adults
○ Habitat is a small intestine of definitive hosts
○ Usually asymptomatic
○ Symptoms
 Anorexia/increased appetite, nausea, abdominal discomfort/pain, dyspepsia
and diarrhoea
○ Examples
 T.saginata, D. latum, H. nana
• Larval forms
○ Extra-intestinal location in the tissues of intermediate hosts
○ Symptoms related to location, size and number of larvae
○ Examples
 E.granulosus, T. solium
• Eggs
○ Morphologically similar

Anatomical features
• Head/Scolex
○ Hooks
○ Bothria or suckers (sucking grooves) - used for attachment to host intestinal mucosa
• Neck - germinative portion, produces proglottids
• Body or strobila consisting of proglottids
○ Hermaphrodite
○ Contain eggs when mature
 May lay eggs or pass proglotids
○ Layout can be overlapping or jointed
○ Consists of digestive and reproductive system, but by maturity, only reproductive
system remains

Pathogenesis

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Pathogenesis
• Adult worms
○ The physical presence of and activity of the large worms (Taenia spp)
○ Erosive actions by scolex resulting in inflammation (T. solium, H. nana)
○ Interferes with host intake of vitamin B (D. latum)
• Larval forms
○ Allergic reactions may be presponsible for generalised symptoms
 Headache, dizziness, inanation, irritation and anal and nasal pruritus

Host immune response

• Most adults are mildly immunogenic - CMI
• IgE and eosinophilia
• H.nana however is very immunogenic due to presence of larvae in villi
○ Humoral and CM immunity
○ Protective against re-infection

Transmission
• Consumption of raw/undercooked meat, fish or grain
• Faeco-orally
○ E.granulosus, T. solium, H. nana
• Auto-infection
○ H.nana, T. solium

Diagnosis
• History of travel in endemic areas
• Stool microscopy
○ Proglotids or eggs in stool
• Serology
○ ELISA
○ Indirect haemaglutination
• Cysts in tissues
○ Biopsy
○ Radiography (calcified cysts)
○ CT (brain cysts)

Parasitology Page 21
Taenia Species
Thursday, 29 September 2016 2:07 PM

• Cyclophilidean cestodes: T.saginata, T.solium, T. asiatica

• Worldwide distribution with T. solium in poorer pig-rearing communities while T. asiatica is
prevalent in Taiwan, China, Indonesia, Thailand
• Adults
○ T.saginata <5m long (up to 25m)
○ T.solium 2-7m
 Has hooklets
• Proglotids
○ T.saginata 1000-2000 proglottids
○ T.solium 1000 proglottids
○ 6-10 proglotids released per day
• Eggs per proglotids
○ T.saginata - 100,000
○ T.solium - 50,000

Taeniasis
• Disease by adult of Taenia species of cestodes involving humans and animal species as
hosts
• Definitive host - humans
• Intermediate hosts
○ Pigs - T. solium
○ Cattle - T. saginata
• Mode of infection
○ Consumption of raw or undercooked meat (pork/beef)
• Cysticercus
○ Infective stage

Life Cycle

Parasitology Page 22
 • Infected person has adults in the small intestines
• Distal/gravid proglotids are released and excreted in their stool
• If stool is passed in the environment, the animals (grazing cattle/feeding pigs) are infected
with the proglotids and then eggs are released.
• The oncospheres puncture the intestinal mucosa and into blood where they find their way
into striated muscle where they develop into cystecerci (can survive here for several
years)
• Ingestion of cysticerci in raw or undercooked food
• Grow into adults (2 months)
• Adults attach to wall of small intestine of man using their scolex
• Mature/gravid proglotids dislodge and pass in stool

Clinical Presentation
• Mild, abdominal symptoms, less with T. solium
• Passage of proglottids per anum
• Intestinal perforation
• Aspiration of vomited proglottids
• Appendicitis - migrating proglottids
• Cholangitis - migrating proglottids

Diagnosis
• Stool microscopy useful after3 months of infection
○ Eggs
 Cannot differentiate from other cestodes
○ Mature proglottids
 Lactophenol then indian ink injection into genital pore, count uterine branches
(T. saginata 12-30 uterine branches, T. solium 7-13)
○ Rarely examination of the scolex
• Repeat stool exams and concentration techniques necessary in light infection
○ Zinc floatation
○ Forma-ether technique
• Antibody detection
○ Especially when larval stages present in early infection

Parasitology Page 23
Treatment
• Praziquantel
• Niclosamide

Cysticercosis
• Disease due to T. solium oncospheres
• Acquired through
○ Ingestion of eggs in faeces contaminated food/water
○ Autoinfection in a human infected with adult T. solium by:
 Ingestion of eggs through foetal contamination of food
 Proglottids carried into the stomach by reverse peristalsis
• Eggs in intestine > oncospheres hatch > invade the intestinal wall >striated muscles, brain,
liver and other tissues > develop into cysticerci

Clinical Presentation
• Asymptomatic subcutaneous nodules and calcified IM nodules
• Cerebral cycticercosis
○ Seizures, mental disturbances, focal neurological deficits and signs of space-
occupying intracerebral lesions/intracranial hypertension > intracranial herniation
• Extra-cerebral neural cycticercosis
• Extra-cerebral cycticercosis
○ Symptoms due to ocular, cardiac or spinal lesions

Diagnosis
• Tissue biopsy
• Antibody detection
• Imaging

Treatment
• Anti-helminths
○ Albendazole
○ Praziquantel for symptomatic patients with live cysts, but not for intraventricular cysts
 Risk of precipitating hydrocephalus
□ + corticosteroids e.g. dexamethasone
• Surgery
• Symptomatic treatment
○ Anticonvulsants

Prevention and Control

• Proper cooking of beef/pork
• Freezing beef/pork (-5℃ x 4/7, -15℃ x 3/7; -24℃ x 1/7)
• Faeces disposal
• Meat inspection
• Treatment
• health education

Parasitology Page 24
Introduction to Laboratory Diagnostic Techniques in
Parasitology
Monday, 3 October 2016 8:18 AM

• Diagnostic stages of parasites:

○ Ova
○ Adult worms
 Helminths
○ Eggs
 Helminths (nematodes, cestodes, trematodes)
○ Cysts
 Protozoa (intestinal protozoa e.g. entamoeba histolytica)
○ Trophozoites e.t.c

Main ways of laboratory diagnosis of parasitic infections

• Microscopic examinations
○ Intestinal, urinary and blood parasites
○ In unstained or stained preparations
○ Either directly or following concentrations
• Culture techniques
○ Only a minority of parasitic infections are diagnosed routinely by cultural techniques
• Immunodiagnosis
○ Serological methods (toxoplasmosis, trichinosis, echinococcosis, cysticercosis, chronic schistosomiasis, extra-
intestinal amoebiasis)
 Antibodies
 Parasite antigens
• Examination of faecal specimens
○ Diagnosis requires properly collected specimens
○ Specimen should be adequate, fresh or properly preserved
○ Right collection tools
○ Check for appearance I (colour consistency, formed or semi-formed or unformed, fluid, whether it contains
blood, mucus, pus or whether it contains worms)

Collection and handling of stool specimen

• Minimum of 3 samples
• Clean, water-tight container with a screw-cap lid
• Minimum stool is 2-5g
• Not contaminated with urine
• Proper labelling
○ Patient name
○ Date and time of sample collection
○ Tests/test requested
○ Suspected diagnosis
○ Clinical findings
○ Travel history
• If you must preserve stool sample:
○ 5 - 10% formalin
○ Polyvinyl alcohol (PVA)
○ Merthiolate iodone (MIF)
○ Formalin
• Stool concentration
○ Removal of debris from the samples
○ Low number of parasites
○ Techniques include
 Formalin-ether (or ether acetate)
□ Centrifugation of the sample (parasites denser than solution thus settle in the sediment of the tube)
 Zinc sulphate floatation technique

Parasitology Page 25
  Zinc sulphate floatation technique
□ After 15 minutes, parasites come out on the surface of the solution
□ For ascaris lumbricoides
• Direct wet preparations
○ Saline wet preparations
 Good for the recovery of the motile protozoan trophozoites
○ Iodine wet preparations
 Study of the detailed morphology of protozoan cysts
• Staining
○ Trichrome stain
○ Giemsa stain
○ Iron haematoxylin stain
○ Modified acid-fast stain (modified Ziehl-Neelsen stain)
 Cryptosporidium, Isopora, Cyclospora
 Requires specific request

Other specimens
• Urine
• Sputum
• Perianal swabs
• Blood

Other Methods
• Molecular techniques
• Immunodiagnosis

Parasitology Page 26
Echinococcus Granulosus
Wednesday, 5 October 2016 2:11 PM

• Dog tape worm, hydatid worm and leads to hydatidosis

• Widely distributed in cattle and sheep rearing areas of Australia and USA, Russia
• Widespread in East Africa and heaviest focus in northern Kenya

Morphology
• Smallest taenid worms
• Has three segments
○ Scolex
○ Neck
○ Strobilia
• The last segment of the worm is the biggest and usually gravid
• Adults in ilium of canines
• Adult tapeworm
○ 3 - 6mm
• Has only 3 proglottids
• Typical taenid scolex with non-retractable rostellum
• Scolex has double rows of 28 - 50 hooks (the usual number is 30 - 36)
• Contained in a broad capsule
• Whole structure contains hydatid fluid
• Hydatid may contain up to one million protoscolices, each capable of developing into a
tapeworm

Larvae forms
• Found within the intermediate host
• Exists as hydatid cysts
• Intermediate host include herbivorous cattle, goats and sheepv~~
• Man is usually an accidental intermediate host

Life cycle in dogs/goats

• Adults in dogs pass eggs in the environment (definite host)
• Eggs are infective to the cows, goats and sheep while grazing (the intermediate hosts)
Within the intermediate hosts arise the hydatid cysts that are spread to the viscera of the
sheep, cows and goats
• This happens 8 hours after ingestion by the goat host
• Young larvae then transforms into hollow bladder
• From the inner sides develops multiple scolices and is filled with fluid inside the bigger
cysts
• This is the fertile hydatid cyst which is when the goats are slaughtered and are fed on by
the dogs then they grow into adult worms in 7 weeks in the dogs
• Dogs usually feed on the offal of the slaughtered goats, sheep and cows
• The worms mature in the dogs and release eggs again

Life cycle in man

• Man accidentally swallows in food/water, the eggs of E. granulosus, deposited by the dogs
• Eggs infective to man emerge in GIT and embryo develops and taken to the lymphatics
and portal system
• The cysts are spread in the liver (90%), lungs, CNS and the rest of the body
• The cysts enlarge and grow in the organs invaded as hydatid cysts
• Man is the dead end host in hydatid disease because dogs do not access the cysts for the
cycle to complete

Parasitology Page 27
 cycle to complete

• Once the cyst is ingested by definitive host, the wall is digested (dogs, foxes, wolves)
• Protoscolices are released and attach to the villi in the intestines
• Mature in 2 months and may live to 5 - 20 months in the canids

Sylvatic transmission cycle

• Wild herbivores (pigs, deers, rodents, cattle) and carnivores
• In unrestricted site, hydatid cyst may grow very large (1.2 ft in diameter and hold up to 18L
of hydatid fluid)
• Occludes the adjacent organs hence necrosis
• Hydatid cysts may rupture
○ Anaphylaxis

• In unrestricted site, hydatid cyst may grow very large e.g. 1 - 2 feet in diameter and hold
up to 18 litres of hydatid fluid
• Occludes the adjacent organs hence necrosis
• Hydatid cysts may rapture
○ Anaphylactic shock

Epidemiological factors
• Close association with dogs e.g. in Lebanon, some traditional tanners are using a tanning
solution containing dogs' faeces
• Handling of infected dogs by children mostly
• Allowing dogs to feed on the same dish/plates used by humans
• Exposing food to be contaminated by canine faeces
• Professional trappers have a high risk of infection
• Dog handlers e.g. Greenland and Iceland

Pathology
• Compression of other organs by the cysts (Space Occupying Lesion - SOL)
• Rupture of the cysts in pleural cavity, peritoneal cavity ,liver
• Anaphylactic shock and other allergic reactions

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 • Anaphylactic shock and other allergic reactions
• Secondary bacterial infection

Clinical presentation
• GIT symptoms of NVD, abdominal pain
• Pruritus, urticaria and allergy symptoms
• Abdominal swellings
• CNS manifestations
○ Headache
○ Convulsions
○ Coma
• SOL fracture of bones
• Anaphylactic shock in surgery

Lab diagnosis
• Casoni intradermal skin test
○ 0.2ml of irradiated hydatid fluid intradermally. After 15 minutes wheal will form.
(Positive if wheal is formed)
• Eosinophilia
• Complement fixation test (CFT)
• ELISA
• Arc 5 test
• Indirect haematoglutination test
• X-rays, CT scan, Ultrasound scan, MRI

Treatment
• Albendazole
○ Shrinks the germinal layer of the hydatid cysts
• Praziquantel
• Paromomycin
• Rhizome of male fern (Dryopteris filix - mass herbal preparation)
• Surgery

Prevention and Control

• Dogs should not be exposed to offal in slaughterhouses
• Secure slaughter houses (fencing)
• Deworm dogs with praziquantel
• Kill stray dogs
• Avoid dog/man contact
• Inspect meat and cooking before feeding dogs
• Environmental sanitation/protection
• Sewerage treatment
• Health education

Parasitology Page 29
Hymenolopiasis
Thursday, 6 October 2016 3:20 PM

• Is caused by 2 different cestode spp

• H.nana is also known as dwarf tapeworm
• H.dimunata also known as rat tapeworm and infects rodents more than humans
• H.nana found worldwide
• H.dimunata less frequent but reported in various parts of the world

Morphology
• H.nana adults measure 15 - 40mm in length
• H.dimunata measures 20 - 60 cm in length
• 4 suckers
• Proglottids - wider than long (mature, immature, gravid)
• Eggs have hexacanthon embryo - 3 pairs of hooklets
• H.nana
○ A small worm with cosmopolitan distribution
○ Commonest tapeworm
○ Scolex has retractable rostellum with a single row of hooks
○ Neck is long and slender
○ Has polar filaments

H.nana
• Eggs infective when passed in stool
• Ingested by an arthropod (beetles and fleas)
• Develop into cysticercoids
• Infect humans or rodents when ingested
• Develop into adults in small intestines. Man can also be infected when ingest eggs in
contaminated food, water or from hands
• Eggs hatch to release the oncosphere
• Develop into cysticercoid larvae
• Evaginates their scolices
• Attach into intestinal mucosa
• Develop into adults in small intestines
• Eggs passed in stool
• An alternate mode of infection consists of internal autoinfection, where eggs passed by
adults release their hexacanth embryo, develop into cysticercoid larvae > evaginate their
scolices > attach into intestinal mucosa > develop into adults in small intestines > eggs
passed out in stool

H.dimunuta
• Eggs infective when passed
• Ingested by intermediate host (fleas)
• Release oncospheres
• Develop into cysticercoid larvae
• Adults
• Humans accidentally infected through ingestion of insects in food and directly from the
environment
• After ingestion > tissue of infected arthropod digested, releasing cysticercoid larvae in the
stomach and SI > eversion of the scoleces > attaches > adults

Morphology

Parasitology Page 30
Morphology
• Nana and dimunata infection are most often asymptomatic
• Heavy nana infection can cause weakness, headaches, anorexia, abdominal pain and
diarrhoea

Diagnosis
• Demonstration of eggs in stool specimens
• Concentration techniques and repeated examination will increase the likelihood of
detecting light infections

Treatment
• Praziquantel
• Niclosamide

Prevention and Control

• Proper faecal disposal
○ Pit latrines
• Don’t use untreated human faeces as manure
• Health education on how the disease is transmitted
• Proper washing of vegetables eaten raw
• Washing of hands before eating meals
• Treatment of infected persons

Parasitology Page 31
Dipyllidium Caninum
Thursday, 6 October 2016 3:41 PM

• A tapeworm of dogs and cats

• Occasionally infects man
• Worldwide distribution

Morphology
• Adults
○ 5 - 40cm
• Head, several proglottids with 2 genital pores "double pores tapeworm"
• Eggs
○ A sack with several golf balls appearance

Life Cycle
• Gravid proglottids passed intact in stool or emerge from the anus of the host, release egg
packets (proglottids rarely rapture and egg packets seen in stool)
• Eggs ingested in larval stage by dog or cat flea, oncosphere released into flea's intestines,
penetrates the wall into body cavity of the flea, develop into cysticercoid larva then to adult
• Dogs infected by ingesting adult flea with cysticercoid and life continues
• Humans infected by ingesting infected fleas
• In the SI cysticercoid develops into adult tapeworm
• Mature proglottids detach, migrate to anus or passed in stool

Clinical Presentation
• Mostly infects children
• Mostly asymptomatic
• Appearance of proglottids around anal region of the child

Diagnosis
• Segment in perianal region
• Characteristic eggs in stool (rarely)

Treatment
• Praziquantel

Prevention and Control

• Control of fleas
○ Insecticides
• Treatment of pet dogs and cats
• Burying of dog and cat faeces
• Health education
○ Washing of hands before a meal and after handling pets
• Treatment of infected persons

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Order Pseudophyllidea
Monday, 10 October 2016 8:23 AM

• Usually with a scolex bearing both dorsal and ventral bothria

• May be deep or shallow, smooth or ciliated, fixed or free and sometimes have hooks
• 3 ventral genital pores usually median
• Proglottids are broader than long
• More than one set of reproductive organs
• Eggs operculated
○ Eggs with a lid like structure

Life Cycle
• Coracidium
• Procercoid
• Plerocrcoid
• Adult

Parasitology Page 33
Diphyllobothrium latum
Monday, 10 October 2016 8:27 AM

• Commonly known as the broad fish tape worm

• Found in the fish eating carnivores of the arctic region, Baltic states, great lake regions etc

Morphology
• Adults between 3 and 12m long an 3 to 10mm wide
• 300 - 3000 proglottids
• Scolex bears finger-shaped bothria (both on dorsal and ventral surfaces)
• Proglottids are wider than long
• Genital antrum is mid-ventral
• Bilobed ovary near the end of the segment
• Gravid segments release eggs through uterine pore (anapolytic shedding)
• About 30 million eggs are released daily
• Eggs are ovoid (60 µm x 40µm)
• Usually provided with an operculum on one end and a small knob on the other

Life Cycle
• Take about 1 - 2 weeks in water to embryonate
• Hatches to release coracidium
• Ingested by copepods, it loses the cilia and penetrates the mid-gut
• Enters haemocoel
• Metamorphoses into procercoid which is large (550 micrometers long)
• Infected copepod is eaten by a freshwater fish and procercoid develops to plerocercoid
larva
• Strobilation and egg production occurs
• Human infections due to eating raw/undercooked fish
• Causes pernicious megaloblastic anaemia by:
○ Robbing the host of vitamin b12
○ Interferes with the intrinsic factor of castle (gastric intrinsic factor)

Symptoms
• Diarrhoea
• Nausea
• Weakness
• Loss of weight
• Wasting of the body
• Abdominal pain

Diagnosis
• Examination of stool
○ Macroscopically
 Proglottids in faeces
○ Microscopically
 Eggs
• Serodiagnosis
○ Coproantigen detection test

Treatment
• Praziquantel (10mg/kg once)
• Niclosamide (1 gm in morning and 1 gm, 1 hour later)

Parasitology Page 34
Prevention and control
• Proper sewage disposal
• Proper and thorough cooking of fish
• Do not feed raw fish to cats or dogs

Parasitology Page 35
Sparganosis
Monday, 10 October 2016 8:42 AM

• Caused by cestodes of the genus Spirometra

• Others pseudophillideans e.g.
○ Diphyllobothrium ursi
○ D.dendriticum
○ Spirometra eminacei
○ S.mansoni
• Common where people eat snakes, concoctions of lizards, frogs e.t.c.
• Infection takes place through
○ Direct ingestion of infected copepods
○ Ingestion of amphibian, reptilia, birds or mammals (intermediate hosts, instead of
fish)n
○ Local application of infected flesh (e.g. split frogs) as poultices to wounds or sore
eyes
• Larvae (spargana) ribbon-like, ivory-white
• Migrate in the tissues causing inflammation, swelling and fibrosis
○ E.g. beneath the skin, between the muscles and occasionally in viscera and brain
• Treatment of the condition involves surgical removal of the larvae
• Read on diagnosis, Rx and Control

Parasitology Page 36
E. multilocularis
Wednesday, 12 October 2016 2:34 PM

• Also called alveolar or multi-alveolar hydatid cyst

• Cyst lacks the multi-laminated capsule
• Thin wall
○ Allows it to form exogenous budding in all tissues
• Looks like cancerous tissues
○ Some alveolar may lack protoscolices
• Difficult to remove the larvae surgically
○ Breaks and metastasizes
• Cyst development slower than that of E. granulosus
• E.multilocularis is also smaller in size (1.5 - 3.7mm in length)
• E.multicularis is common in Europe (Britain and Siberia), parts of Japan, Alaska, Turkey
and South America
• Main definitive hosts are foxes
• In the domestic cycle, the intermediate hosts
• Herbivores and rodents and dogs the definitive hosts

Diagnosis
• Very difficult but X-rays or ultrasound imaging are useful
• Immunodiagnostic techniques are now available to diagnose the infection

Parasitology Page 37
 Trematodes
Wednesday, 12 October 2016 2:40 PM

Characteristics
• Dorsoventrally flattened
• Unsegmented
• Leaf-like
• Hermaphroditic except blood flukes
• Two radially striated suckers
○ Have 2 strong muscular cup shaped depressions called suckers used as organs of attachment
 The sucker surrounding the mouth is referred to as the oral sucker
 The one on the ventral surface of the body is known as the ventral sucker (acetabulum)
• Have no body cavity and an incomplete digestive tract
• Adults are covered with spines, except schistosomes
• Most of the body is occupied by reproductive organs
• The excretory and nervous system are present. The excretory system consisting of 'flame cells', which open into a posterior
excretory pore
• The worms are oviparous
○ Produce eggs
• All trematode eggs are operculated except schistosomes and can only develop in water

Classification
• Is according to the habitat of the trematodes (the adults)
○ Intestinal trematodes (intestinal flukes)
 Small intestines
□ F.buski
□ H.heterophyes
□ M.yokogawai
□ W.watsoni
 Large intestines
□ G.hominis
○ Hepatic trematodes (liver flukes)
 C.sinensis
 O.felineus
 F.hepatica
○ Lung trematodes (lung flukes)
 P.westermani
○ Blood trematodes (blood flukes)
 In vesical venous plexuses
□ S.haematobium
 In rectal and portal venous system
□ S.mansoni

Family Genus Species Habitat Manifestations

Schistosomatidae Schistosoma Haematobium Blood Haematuria
Mansoni Blood Dysentery
Japonicum Blood Dysentery and liver cirrhosis
Paramphistomatidae Gastrodiscoides Hominis L.intestines Mucous diarrhoea
Watsonius Watsoni S.intestines Diarrhoea
Fascioloideae Fasciola Hepatica Liver of sheep/man Biliary colic
Fasciolopsis Buski S.intestines

Mode of Infection
• By ingestion of encysted infective stage known as cercariae/metacercariae
○ Vegetables as in F. hepatica, F. buski, W. watsoni
○ Fish as in C. sinensis, H. heterophyes, M. yokogawai
○ Flesh of crab or crayfish as in P. westermani
• Free cercariae penetrating through the skin

Parasitology Page 38
 • Free cercariae penetrating through the skin
○ As in S. mansoni, haematobium and japonicum

Life Cycle
• The worms pass their life cycle in 2 different hosts
○ The definitive host
 Generally humans harbour the adult worm
○ The intermediate host
 A fresh water snail of mollusc
 A second intermediate host may be required (fish/crab)

Life Cycle of Fasciola Hepatica

• Unembryonated eggs passed in faeces
• Embroyonated eggs in water
• Miracidia hatch, penetrate the snail
• In the snail tissue, sporocysts > rediae > cercariae
• Free swimming cercariae encyst on water plants
• Metacercariae on the water plant ingested by human, sheep or cattle
• Encyst in duodenum
• Adults in hepatic biliary tracts

Life cycle of Fasciolopsis buski

○ Unembryonated eggs passed in faeces
○ Embroyonated eggs in water
○ Miracidia hatch, penetrate the snail
○ In the snail tissue, sporocysts > rediae > cercariae
○ Free swimming cercariae encyst on water plants
○ Metacercariae on the water plant ingested by humans or pigs
○ Encyst in duodenum
○ Adults in small intestines

Pathology
• Eggs cause fibrosis
• Liver, lung, GIT and other sites
• Adult and larvae are not significant in causing pathology
Q
Clinical Features
• Eggs cause fibrosis in
○ Lung, liver, GIT and other sites
○ NVD, abdominal pain, obstruction, jaundice, fever, coughing and irritation, haemoptysis, anaemia and weakness

Laboratory Diagnosis
• Eggs recovered in sputum
• Larvae in the snail
• Second larval stage in the crabs and fish
• Chest and abdomen x-rays, ultrasounds and MRIs

Treatment
• Chemotherapy
○ Praziquantel
• Surgery for obstructions

Prevention and control

• Health education
• Early diagnosis and treatment
• Mass treatment pf population with praziquantel
• Snail control
• Environmental control

Parasitology Page 39
Schistosomiasis
Wednesday, 26 October 2016 2:26 PM

Schistosomiasis
Epidemiology
• One of the most widespread of the human parasitic diseases
• In over 79 countries worldwide
○ Mainly Africa, parts of Asia, Caribbean islands and South America
• Over 207M people are infected and a further 600M are at risk
• Children aged between 10 - 14 years bear the highest burden of disease
• Mainly caused by Schistosoma haematobium (only one that does not cause intestinal
schistosomiasis, but instead, causes terminal haematuria, and is found in coastal Kenya),
S. mansoni (not found in coastal Kenya) ,and S. japonicum, S. mekongi, S. intercalatum

Morphology
• Adult
• Eggs
○ Lateral spine - mansoni
○ Terminal spine - Haematobium, mekongi
○ Rudimentary spine - S. japonicum
• Miracidium
• Sporocyst
○ Amorphous stage
○ Mother sporocyst has germinal cells that leads to release daughter sporocysts that
develop to cercaria
• Cercaria
○ Has a forked tail

Life Cycle
• Eggs found in stool and urine
• Gets into water source and eggs are already embryonated and hatch in contact with water
to release the free swimming miracidium
• Miracidium swim and attach to snail due to chemotaxis
○ Biomphilaria spp - mansoni
 Not in coast because of high temp as the gonads atrophy
○ Pfeifferi spp
○ Bulinus spp - intermediate host of haematobium and intercalatum
○ Onchomelania spp - intermediate host of japonicum
• Penetrates snail tissue and form sporocytes. Germinal cells release daughter sporocytes
that release cercaria that are released in water and infect humans (percutaneous infection)
• The tail is left out and forms the schistosomula then form the adult.
• The adult male forms a gynaecophoric groove around the female and are in constant
copulation
• Haematobium spp moves to the pelvic venous plexus and other spp move to the
mesentery
• The female lays eggs that penetrate lumen and use their spines to attach to the mucosa to
cause haematuria in Haematobium spp and occult blood in stool in other spp

Predisposed groups
• Rice growers
• Fishermen
• Women while doing domestic chores
• Swimming

Parasitology Page 40
 • Swimming

Pathology and clinical manifestation

Acute Schistosomiasis
• Cercarial dermatitis
○ Local inflammation
○ In previously sensitised individuals
○ Petechial haemorrhages at the site of penetration
○ A pruritic papular rash develops 5 - 15 hours later
• Katayama fever
○ Lasts about 8 weeks
○ Usually asymptomatic in previously exposed individuals
○ Occur 3 to 8 weeks after infection but lasts several weeks
○ Coincide with onset of egg laying by adult worms
○ Caused by immune complexes
○ Symptoms include
 Chills, headaches, sweating, weakness, anorexia, cough, abdominal pain and
diarrhoea
 Enlarged, tender liver
 Splenomegaly
 Lymphadenopathy
 Eosinophilia of 30 - 40%
 Toxaemia

Chronic Schistosomiasis
• Chronic pathology due to tissue lodged eggs
• Causes pathology in practically any tissue
• Delayed type hypersensitivity reaction
○ Peripheral granulomas
• Down modulation
○ Reduction in size and level of inflammation
• Collagen synthesis and degradation
• Fibrosis
○ Late chronic pathology
• Several clinical syndromes concurrently

Hepatosplenic disease (Mansoni and Japonicum)

• Eggs trapped in the vicinity of the portal veins and cause peri-portal fibrosis
○ Liver becomes hard with protuberances on its surface
○ Coarse peri-portal, clay-pipe-stem fibrosis (Symmer's fibrosis)
○ Pre-sinusoidal types of portal hypertension
 Appearance of varices

○ Collateral circulation
 May carry eggs to the lungs
• Immunological disturbances
○ Complications e.g. glomerulonephritis
• Enlarged firm liver with left lobe prominence
• Late stages
○ Liver may shrink
○ Development of ascites and oedema
• Main complication is variceal haemorrhage
○ Fulminant or repetitive haematemesis and melaena
○ Rupture at the gastro-intestinal junction
• Splenomegaly
○ Hypersplenism (hyperplasia, due to increased portal pressure) causing

Parasitology Page 41
 ○ Hypersplenism (hyperplasia, due to increased portal pressure) causing
granulocytopenia, thrombocytopenia and anaemia
○ Caput medusae

Intestinal Schistosomiasis
• Mansoni and japonicum - throughout the GIT
• Intercalatum - rectosigmoid
○ Characterised by granulomas in GIT
○ Bowel symptoms
 Nausea
 Abdominal pain
 Anorexia
 Weight loss
 Lassitude
 Myalgia
○ Bloody mucoid diarrhoea
○ Pellagra
• Sessile or pedunculated polyps
• Pseudoneoplastic form
○ Colon
○ Ileum
○ Mesentery
○ Retroperitonium (abdominal mass (bilharziaoma))
• Schistosomal colonic cord
○ Stenotic segment

Schistosomal appendicitis
• Rare
○ Occlusion of the lumen of appendix by fibrosis

Pulmonary schistosomiasis
• Migration of schistosomula
○ Bronchospasm
○ Fever
○ Cough
• Dying or dead adult worm in lung
○ Verminous pneumonia
• Collateral circulation
○ Shunting of mansoni and japonicum eggs to primary circulation
• Pulmonary hypertension
○ Leads to cor pulmonale
• Rare in haematobium infection
• Egg deposition
○ Pulmonary necrotising arteriolitis

Urinary schistosomiasis
• Renal involvement
• Kidneys may be infected in infections with haematobium, mansoni and japonicum
○ Immune complexes deposition in renal glomeruli
 Nephrotic syndrome

Haematobium - Urinary pathology

• Egg deposition in the urinary tract walls and organs
• Peri-oval granuloma formation
• Ureteric strictures and urolithiasis
○ Renal damage

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 ○ Renal damage
 Hydronephrosis and pyelonephritis
• Loss of elasticity of bladder wall

CNS schistosomiasis
• Can be caused by all three species
• Japonicum - cerebrum
• Haematobium and mansoni - spinal cord
• Eggs may reach CNS through
○ Pathologically altered pulmonary vessels
○ Arteriovenous anastomoses
○ By way of the vertebral venous system
• Acute CNS schistosomiasis may resemble encephalitis
○ Granulomatous mass or blockage of CSF may cause intracranial hypertension
• Focal epilepsy
○ E.g. Jacksonian and psychomotor epilepsy
• 20% develop hemi or monoparesis with or without motor aplasia
• Blindness has been reported
• Pituitary involvement
○ Hypopituitarism
• Mansoni and haematobium
○ Spinal cord schistosomiasis
 Granulomatous mass mimicking a tumour
□ Radiculitis
□ Transverse myelitis

Malignancies
• Haematobium
○ Squamous cell carcinoma of the bladder
• Mansoni infection
○ Follicular lymphoma of the spleen
○ Carcinoma of colon or rectum
• Well differentiated adenocarcinoma with pseudopolyps and calcified eggs in the tissues
• Epithelial hyperplastic colonic polyps are a risk factor for colon carcinoma
• Japonicum and mansoni
○ Hepatocellular carcinoma

Diagnosis of schistosomiasis
• Clinical manifestations
• Parasitological methods
• Rectal mucosa snips
• Stool examination
○ Direct smear
○ Formal-ether sedimentation method
○ Bell method
○ Kato-katz technique
• Urine examination
○ Centrifugation technique
○ Filtration technique
• Immunodiagnostic tests
○ Serodiagnosis
○ Circulating schistosomal antigens
• Imaging techniques
○ Ultrasonography
○ X-ray calcification

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 ○ X-ray calcification
○ CT and MRI
 CNS schistosomiasis

Chemotherapy
• Antimony compounds
• Hycanthone
• Niridazole
• Metriphonate
• Oxamniquine
• Praziquantel (adults)
• Artemisinin (larval schistosomes)

Control
• Breed in slow moving/stagnant fresh water.
• Molluscides - endod (from gooseberry)
• Biological methods
○ Cray fish to eat the snails
• Sewage treatment

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Heterophyes heterophyes
Wednesday, 26 October 2016 3:24 PM

• H.heterophyes, H. dispar, H. aequilis

• Belong to the family heterophydiae of class trematoda
• Are minute intestinal flukes

Heterophyes
• Adults measuring 1.0 - 1.7mm by 0.3 - 0.4mm
• In addition to humans, various fish-eating mammals (e.g. cats and dogs, birds) can be
infected by heterophyes heterophyes

Geographical distribution
• Distributed in the orient
○ Japan
○ Korea
○ China
○ Taiwan
○ Philippines
• Africa
• Asia Minor

Morphology
• 1.7 x 0.4mm
• Pyriform - leaf-like
• Covered with a scale like spine more numerous at the anterior end
• Suckers
○ Oral sucker
 Around the mouth anteriorly
○ Ventral sucker
 Large
 Lies in the middle third of the worm, at the middle line
○ Genital sucker
 Around the common genital pores
 Situated posterior and lateral to the ventral sucker
• Digestive system
○ The muscular pharynx surround only the middle part of the oesophagus and the
intestinal caeca are simple
• Reproductive organs
○ Male genital organs
 Two oval testes lie side by side near the posterior end of the worm
○ Female genital organs
 The ovary is spherical in front and between the testes
 Vitelline glands are 7 pairs on both sides posteriorly
• The eggs are characterised by
○ Size
 Small 30 x 15µm
○ Shape
 Oval
○ Shell
 Thick
 Double layer with an operculum at the narrow pole and a knob like thickening
at the other pole

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Life Cycle
• Embryonated eggs each with a fully developed miracidium are passed in faeces
• Snail host ingests eggs, miracidia emerge from the eggs and penetrate the snail's
intestines
○ Sporocystes > rediae > cercariae
 In snail tissue
• Cercariae released from snails
• Cercariae penetrate the ski of fresh/brackish water fish and encyst as the metacercariae in
the tissue of the fish
• Host becomes infected by ingesting undercooked fish containing the metacercariae
• Metacercariae encyst in the small intestine
• Adult in small intestines
• Fish eating mammals can be infected as well

Clinical Features
• The main symptoms are diarrhoea and colicky abdominal pain
• Migration of the eggs to the heart, resulting in potentially fatal myocardial and valvar
damage, has been reported from the Philippines
• Migration to other organs (e.g. brain) has also been reported

Laboratory diagnosis
• Diagnosis is based on the microscopic identification of eggs in the stool
• However, the eggs are indistinguishable from those of metagonimus yokagawai and
resemble those of clonorchis and opisthorchis

Treatment
• Praziquantel is the drug of choice

Prevention and Control

• Avoid eating raw of undercooked fish
• Control of snails
• Hygienic practices
• Treat infected patients

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Paragonimus Species
Wednesday, 26 October 2016 4:04 PM

• Belongs to the troglotrematidae family

• There are >30 several species of trematodes of the genus paragonimus
• More than 10 species infect man causing paragonimiasis

GD
• America
• Africa
○ Western Africa for Paragonimus africanus
• SE Asia
• Japan

Morphology
• They vary in size
• Adult
○ 15mm x 8mm
○ Oval in shape
○ Thick tegument
○ Oral sucker and acetabulum
○ Ovaries are behind the acetabulum
○ Posterior to the ovary are the testes

Life Cycle
• Definitive host human, ingest inadequately cooked or pickled crustaceans containing
metacercariae
• Metacercariae encyst in duodenum
• Penetrate through intestinal wall into the peritoneal cavity, through the abdominal wall and
diaphragm into the lungs
• They transform into cyst and cross fertilise with one another
• The cyst ruptures, releasing eggs which are excreted in sputum
• Alternately, eggs are swallowed and passed with stool
• Eggs in water hatch into a ciliated miracidium which penetrates the snail
• Crustaceans i.e. crabs, crayfish, become infected when they eat infected snail
• Life cycle continues when definitive host ingests the infected crustacean
• The life cycle is complete when the worms reach other organs and tissues i.e. brain and
striated muscles
• The eggs laid cannot exit the sites

Clinical presentation
• Acute infection characterised with
○ Cough
○ Abdominal pain
○ Discomfort
○ Low grade fever 2 - 15 days after infection
○ Subcutaneous abscesses (creeping eruption)
○ Low grade infections
• No symptoms (asymptomatic)
• Long term infection
○ Mimic
 Bronchitis
 Tuberculosis

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  Tuberculosis
 Blood tinged sputum
 Fever and hives

Laboratory Diagnosis
• Microscopic identification of eggs
○ Sputum
○ Stool
○ Tissue biopsy
○ Serological test based in a specific antigen

Treatment
• Praziquantel
○ Adult or paediatric - 25mg/kg
○ Orally - 3X daily for 2 days
• Triclabendazole
• CD corticosteroids with praziquantel riradf

Prevention and Control

• Cook crabs and crayfish at 63 degrees
• Avoid traditional food in a foreign land
• Avoid picked crabs and crayfish
• Thorough cleaning of utensils prior to food preparation

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Metagonimus Species
Wednesday, 26 October 2016 3:46 PM

• Disease
○ Metagonimus
• Geographical distribution
○ Korea
○ Japan
○ China
○ Taiwan
○ Russia
• Infection rate
○ Unknown in worldwide

Morphology
• The flukes are characterised by:
○ Small body
 1.0 - 2.0mm long by 0.4 - 0.6mm wide
○ Laterally located ventral suckers
○ No genital sucker or ventrogenital apparatus
○ Egg size
 Metagonimus takahashii (32 - 36mm) differs from yokagawai (28 - 30mm)

Life cycles
• The first intermediate hosts are fresh water snails
○ Semisulcospura coreana or S. libertina (M. yokagawai)
○ S.coreana or Koreanomelania nodifila (M. takahashii)
○ S.globus (M. moyatai)
• The second intermediate hosts of M. yokagawai include
○ Sweet fish (plecoglossus altivelis)
○ Dace (Triolodon spp)
○ Perch (Lateolabrax japonicus)
• The fish hosts for M. takahashii are the carp (Cyprinus Caprio) and dace (Tribolodon
taczanowskii)
• Those for M. miyatai are found under the scale of the sweet fish, dace, pale chub, dark
chub and common fat minnow
• Dogs, rats and cats are reported to be naturally infected final host
• Embryonated eggs each with a fully developed miracidium are passed in faeces
• Snail host ingests eggs, miracidia emerge from the eggs and penetrate the snail's
intestines
○ Sporocystes > rediae > cercariae
 In snail tissue
• Cercariae released from snails
• Cercariae penetrate the ski of fresh/brackish water fish and encyst as the metacercariae in
the tissue of the fish
• Host becomes infected by ingesting undercooked fish containing the metacercariae
• Metacercariae encyst in the small intestine
• Adult in small intestines
• Fish eating mammals can be infected as well

Pathology
• Except for heavy infections, there is significant injury of the intestine nor marked
symptoms

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 symptoms
• In heavy infections, irritation of the intestinal mucosa may result in a chronic intermittent
mucous diarrhoea with colicky pains, abdominal discomfort and tenderness
• There is eosinophilia but no anaemia
• When the worms penetrate the intestinal wall, the eggs may get into lymphatics and
venules and causes granulomatous lesions in such distant foci such as the heart and brain

Diagnosis
• Identification of eggs by stool examination
• Differentiation from other heterophyid flukes and clonorchis eggs needed

Treatment
• The drug of choice is praziquantel as a 10mg/kg single dose

Prevention
• Prohibit eating rae, undercooked or recently salted fish in endemic areas

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Liver Trematodes - Fasciola Hepatica
Thursday, 27 October 2016 9:38 AM

• Life cycle involves 2 intermediate hosts

• First intermediate host is a snail
• Infective form is metacercaria
• Habitat
○ Biliary and +/- pancreatic ducts, attach to mucosa
• Pathogenesis
○ Inflammation of biliary ducts
○ Obstruction of bile ducts, intermittently

Fasciola Species
• Fasciola hepatica (sheep liver fluke) and Fasciola gigantica are parasites of herbivores
(cattle, camels, buffalo in Africa and Asia) that can infect humans accidentally
• Reside in biliary ducts
• Human infections with F. hepatica are found in middle East, Asia especially areas rearing
sheep and cattle and humans eat raw watercress
• Infecions with F. gigantica have been reported rearely in Asia, Africa and Hawaii
• Cause Fascioliasis

Morphology
• Adults
○ Hepatica
 30mm long by 13mm
 Triangular in shape with anterior conical projection
 Intestinal caecum is highly branched
○ Gigantica
 Up to 75 mm long
• Eggs
○ Fasciola hepatica broadly ellipsoidal, operculated and measure 130 - 150 µm
○ Unembryonated when passed in stool

Life Cycle
• Unembryonated eggs passed in stool
• Embryonated eggs in water
• Miracidia hatch and penetrate the snail (genera Galba, Fossaria and pseudosuccinea)
• In the snail tissue
○ Sporocysts > rediae > cercariae
• Free swimming cercariae encyst on water plants esp. watercress
• Metacercariae on water plant is ingested by human, sheep or cattle
• Excyst in the duodenum
• Adults in hepatic biliary ducts

Clinical Presentation
• Acute phase (caused by the migration of the immature fluke through the hepatic
parenchyma)
○ Manifestations include
 Abdominal pain
 Hepatomegaly
 Fever
 Vomiting
 Diarrhoea

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  Diarrhoea
 Urticaria
 Eosinophilia
○ Can last for months
• Chronic phase (caused by the adult fluke within the bile ducts), the symptoms are more
discrete and reflect intermittent biliary obstruction
• Occasionally, ectopic location s of infections (such ad intestinal wall, pharyngeal mucosa,
lungs and subcutaneous tissue) can occur

Diagnosis
• History
• Microscopic exam of eggs is useful in the chronic stage
• Eggs recovered in stool or duodenal/biliary aspirate
• False fascioliasis (pseudofascioliasis) refers to eggs in stool resulting not from actual
infection but recent ingesting of infected livers containing eggs (have patient eat a liver
free diet for several days before a repeat stool examination)
• Antibody detection is useful especially in early stages, when eggs are not yet in stools or
in ectopic fasciolisis

Treatment
• Unlike with other flukes, fasciola hepatica, may not respond to praziquantel
• Drug of choice is triclabendazole
• Bithionol is an alternative

Prevention and Control

 Snail management
 Proper disposal of faecal waste
 Proper cooking of water plants before eating
 _

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Clonorchis Sinensis
Thursday, 27 October 2016 10:01 AM

• Hosts
○ Humans
○ Carnivores (reservoir host)
• Causes clonorchiasis
• Endemic in Korea, China, Taiwan and Vietnam
• Adults measure 10 - 25mm by 3 - 2mm

Morphology
• Highly branched uterus and testes
• Interior sucker present

Life Cycle
• Embryonated eggs passed in faeces
• Eggs are ingested by the snail
○ Miracidia > sporocysts > rediae > cercariae
• Free swimming cercariae encyst in the skin or flesh of fresh water fish
• Metacercariae in flesh or skin of fresh water fish are ingested by human host
• Excyst in the duodenum
• Adult in the biliary duct

Clinical Presentation
• Acute phase
○ ND
○ Abdominal pain
○ Eosinophilia
• Chronic phase
○ Cholangitis
○ Cholelithiasis (gallbladders)
○ Pancreatitis
○ Cholangiocarcinoma

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Opisthorchis Species
Thursday, 27 October 2016 10:08 AM

• O.viverrini
○ SE Asian liver fluke and O. felineus (cat liver fluke)

GD
• Viverrini
○ Thailand
○ Laos
○ Kampuchea
• Felineus
○ Asia
○ Russia
○ Europe
• Cause opisthorciasis
• Size
○ Viverrini
 5 - 10mm x 1 - 2mm
○ Felineus
 7 - 12mm x 2 - 3mm

Life Cycle
• Embryonated eggs passed in faeces
• Eggs are ingested by the snail
○ Miracidia > sporocysts > rediae > cercariae
• Free swimming cercariae encyst in the skin or flesh of fresh water fish
• Metacercariae in flesh or skin fresh water fish are ingested by human host
• Encyst in the duodenum
• Adult in the biliary duct

Clinical Presentation
• Most infections are asymptomatic
• Mild infection
○ Dyspepsia
○ Abdominal pain
○ Diarrhoea
○ Constipation
• Katayama like syndrome possible in acute phase of O. felineus infection
○ Fever
○ Rash
○ Lymphadenopathy
○ Facial oedema
○ Arthralgia
○ Eosinophilia
• Chronic/severe infections
○ Hepatomegaly
○ Malnutrition
○ Cholangitis
○ Cholecystitis
○ Cholangiocarcinoma
○ Pancreatitis (O. felineus)

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Diagnosis (clonorchiasis and opisthorchiasis)
• Identification of eggs in stool
• Serology

Treatment (clonorchiasis and opisthorchiasis)

• Praziquantel
○ Adults: 75mg/kg/day orally, three doses per day for 2 days
• Albendazole
○ 10mg/kg/day for 7 days

Prevention and control

• Proper disposal of faecal waste
• Proper cooking of fish before eating
• Treatment of infected person
• Health education

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Parasitic Protozoa
Thursday, 13 April 2017 6:20 AM

Intestinal Protozoa
• Giardia lamblia (flagellate)
• Entamoeba histolytica (amoeba)
• Cryptosporidium hominis (sporozoa)
• Cyclospora cayetanensis (sporozoa)
Sexually Transmitted Protozoan Infection
• Trichomonas vaginalis (flagellate)
Blood and Tissue Protozoan Infections
• Flagellates
○ Trypanosoma brucei rhodensiense and gambiense
○ Trypanosoma cruzi
○ Leishmania donovani, tropica, mexicana
• Amoebae
○ Entamoeba histolytica
○ Naegleri fowleri
○ Acanthamoeba castellanii
• Sporozoa
○ Plasmodium vivax, ovale, falciparum, malariae, knowlesi
○ Babesia microti
○ Toxoplasma gondii
• Microsporidia

Key Concepts
• Flagellates and amoebae multiply by binary fission
• Sporozoans reproduce through merogony (or schizogony) in which the nuclei replicates
prior to cytokinesis
• Sporozoans (Cryptosporidium, Plasmodium, Toxoplasma) undergo sexual recombination,
which leads to genomic and antigenic variation
• Protozoa can multiply quickly (in the order of several hours) in the host and can cause a
rapid onset of symptoms
• Intestinal infections are acquired by ingestion of environmentally resistant cyst (or oocyst)
form. Blood infections are vector borne
• Infections by intracellular protozoa (Trypanosoma cruzi, Leishmania spp. Cryptosporidium,
Toxoplasma and Plasmodium) are difficult to treat because drugs must cross plasma
membranes. No vaccines are available for any human parasitic disease
• Latent infections occur w/ Toxoplasma (parasites in tissue cysts are called bradyzoites)
and Plasmodium vivax and ovale (parasites in liver are called hypnozoites)
• In disseminated protozoal infections, fever and flu like symptoms occur and are non-
specific
• Some parasitic protozoa are able to evade the host's immune response because they are
intracellular and/or undergo antigenic variation

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Giardia lamblia
Wednesday, 9 November 2016 9:22 AM

• Also referred to as Giardia intestinalis/Giardia duodenalis

• Only common pathogenic protozoa found in the duodenum and jejunum of humans

Morphology
• Exists as cysts and trophozoites
○ Trophozoite
 Heart-shaped/Pyriform shaped and convex
 Bilaterally symmetrical and possess
□ 4 pairs of flagella
□ 1 pair of nuclei
□ 1 pair of axostyles, running along the midline
 Approx. 15µm in length
 A large ventral sucker present (adherence to intestinal villi)
○ Cysts
 Ellipsoid
 Thick-walled and highly resistant (hyaline)
 8-14µm in length
 Contain 2 nuclei immaturely and 4 as mature cysts

Life Cycle
• Passes its life cycle in 1 host
• Infective form
○ Mature cyst
• Mode of transmission
○ Ingestion of cysts in contaminated water or food
○ Direct person-person transmission, common in children, male homosexuals and
mentally-ill patients

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 •

Pathogenesis
• Usually in glandular crypts of duodenojejunual mucosa
○ May cause crypt or villus atrophy or flattening by apoptosis and epithelial cell
damage leading to increased lymphatic infiltration of lamina propria
○ Variant specific surface proteins (VSSP) of Giardia, play an important role in
virulence and infectivity of the parasite. It might also stimulate the above cell
apoptosis
• Do not invade tissues but adheres tightly to the intestinal epithelium - no bleeding
○ Attaches to epithelial surface of the duodenum and jejunum and may lead to
duodenitis or jejunitis
○ No inflammatory cells involved
• Feed on mucosal secretions and stimulate excessive mucosal secretion, that may impair
fat absorption leading to steatorrhoea. This might lead to malabsorption of the vitamins A,
D, E and K
• Large number of parasites attached to the bowel wall may cause irritation and low-grade
inflammation
• May colonise the gall bladder causing biliary colic and jaundice

Clinical Presentation
• Dull epigastric pain, flatulence, chronic diarrhoea (steatorrhoea type) which is voluminous
and foul smelling with loads of mucus, fat and no blood
• In the biliary tract
○ Chronic cholecystitis and jaundice

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 ○ Chronic cholecystitis and jaundice
• In HIV positive patients, there is associated hypergammaglobinaemia, protein/caloric
malnutrition, gastric acclorhydra, decreased release of secretory IgA
• Children may develop chronic diarrhoea, malabsorption of fat, Vitamin A, protein, sugars
e.g. xylose disaccharides, weight loss and sprue-like syndrome

Pathogenesis - Immune Response

• Humoral
○ Majority produce detectable levels of Giardia spp. Antibodies role is however unclear
○ IgM anti-Giardia antibodies
 Short-lived
 May be important in both defence and parasite clearance
○ IgG anti-Giardia antibodies
 Remain in high titres for many months after treatment
○ There's a certain degree of protein in breastmilk_

Laboratory Diagnosis
• Stool examination
○ Macroscopic examination
 Parasites are passed in stool cyclically
 Cysts are formed in stool
 Trophozoites and cysts in diarrhoeal stools
○ Microscopic examination
 Trophozoites and cysts seen in diarrhoeal stools by normal saline and iodine
preparations
• Enterotest (string test)
• Serological tests
○ Antigen detection
 ELISA and IFA
□ Monoclonal Ab's using fluorescent method
○ Antibody detection
 ELISA and IFA
□ Monoclonal Ab's using fluorescent method
• Molecular diagnosis
○ DNA probe
○ PCR
• Trophozoites seen in bile aspirated from duodenum by intubation and by duodenal
capsule technique
• Duodenal and jejunal biopsies
• Touch preparations are air dried and fixed with methanol and stained with Giemsa and
trophozoites appear purple

Treatment
• Metronidazole and Tinidazole
○ Tinidazole is more effective than metronidazole
• Furazolidine and Nitazoxanide
○ Preferred in children as they have fewer AE
• Paromomycin
○ Given to symptomatic pregnant females
• Only symptomatic cases need treatment

Prevention
• Improved water supply to prevent infection through contaminated water
• Proper faeces disposal to prevent contamination of food and water
• Improve personal hygiene
• Proper food storage

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• Proper food storage
• Routine hand washing before eating to cut off faecal-oral contact
• Proper pest control
• Treatment of symptomatic and asymptomatic patients

Parasitology Page 60
Entamoeba histolytica
Wednesday, 2 November 2016 2:09 PM

• Causes a disease called amoebiasis or amoebic dysentery

• A disease spread or transmitted faeco-orally
• Distributed widely in the tropics where the environment is warm and moist and where the
population practice poor hygiene
• Parasite is also spread by houseflies (Musca domestica) and cockroaches
• Man is the definitive host. No intermediate host

Habitat
• The parasite invades the colonic mucosa causing ulcerations and sloughing followed by
leading to dysentery

Morphology
• Trophozoite
○ Irregular shape
○ Approx. 12-20µm
○ Cytoplasm
 Outer ectoplasm - clear and refractile
 Inner endoplasm - finely granular w/ round glass appearance
○ Has pseudopodia (finger-like projections formed by sudden jerky movements of
ectoplasm in one direction) for motility (crawling or gliding) - inhibited at low temp.
○ Nucleus is spherical 4-6µm in size and contains a central karyosome surrounded by
a clear halo
• Pre-cystic stage
○ Trophozoites that have undergone encystment in the intestinal lumen
 It extrudes food vacuoles and becomes round/oval - 10-20µm in size
○ Large glycogen vacuole
○ Two chromatid bars
• Cystic stage
○ Pre-cystic stage secretes highly retractile cyst wall around it
○ Early cyst
 Single nucleus
 A mass of glycogen
 1-4 chromatid bars or bodies w/ rounded ends (cigar shaped)
○ Mature cyst
 Quadrinucleate
□ Nucleus undergoes 2 successive divisions
 Glycogen mass and chromatid bars disappear

Life Cycle

Parasitology Page 61
 • Cysts are released with faeces which contaminate the environment, water and food
• Humans ingest the contaminated water and food
• The cyst wall is removed by action of digestive enzymes to liberate the vegetative form
(trophozoite)
• The trophozoite reproduce by binary fission in the colon
• When the colonic contents become less fluid, the trophozoites undergo encystation again
and the life cycle is repeated

Pathology and Pathogenesis

• The cysts are the infective form and are transmitted through contaminated food and water
and also found to be transmitted anal-orally in male homosexuals
• Trophozoites invade the intestinal epithelium and form discrete ulcers with pinhead sized
centre and raised edges. The trophozoites multiply rapidly and undermine the mucosa and
produces a "flask-shaped" ulcer of primary amoebiasis.
• Coalescing of the flask shaped ulcers leads to sloughing off of the mucosa leading to
mucoid diarrhoea. May also lead to impaired absorption of water that may lead to
dehydration
• Penetration of the muscle layers and occasionally the serosa may lead to eventual
perforation into the peritoneal cavity that might lead to abdominal pain and discomfort
• An amoeboma (amoebic inflammatory or granulomatous tumour like mass) may form on
intestinal wall and may grow large enough to block the lumen
• The colonies of trophozoites may form microemboli and cause extra intestinal infection,
more commonly on the liver through the portal system, and may lead to formation of a
hepatic abscess whose contents are bacteriologically sterile amoeba and necrotic tissue,
which gives the classic anchovy paste appearance. Mainly occurs in the right lobe of the

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 which gives the classic anchovy paste appearance. Mainly occurs in the right lobe of the
liver. When the abscess ruptures, it might cause peritonitis
• Rarely do abscesses form elsewhere but can occur in the lung, brain or spleen

Clinical Manifestations
• Abdominal pain and discomfort
○ Right upper quadrant pain
• Diarrhoea (watery or dysentery)
○ Bloody diarrhoea due to invasiveness of E. hystolytica
• Liver abscess
• Intestinal obstruction
• Amoeba due to scar formation and tumour-like appearance
• Abdominal wall ulceration
• Lung and brain abscess
• Rupture of liver abscess with development of peritonitis

Diagnosis
• History of passing bloody stool and abdominal pain
• Microscopy
○ Stool microscopy for the cysts (4 nuclei)
○ Stool microscopy for haematophagous trophozoites (trophozoites with engulfed or
phagocytosed red blood cells)
 This is a pathognomonic finding
• Serology
○ ELISA
 To detect parasitic antigens in stool or serum
• Radiological examination can be used to detect abscesses, amoeboma
○ Colonoscopy/sigmoidoscopy - visualise flask-shaped lesions
○ CT scans - visualise the amoeboma and abscesses
○ MRI
○ Ultrasonography

Treatment
• Metronidazole
• Paromomycin - luminal agents to eliminate the cysts
• Iodoqinol - luminal agent that can be used in combination w/ Metronidazole
• Secnidazole
• Tinidazole
• Aminsidine
• No surgical removal of the abscesses

Prevention and Control

• Wash hands before meals
• Proper excreta disposal
• Proper sewage management
• Control of houseflies using insecticides
• Health education
• Treatment of infected individuals

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Cryptosporidium
Wednesday, 9 November 2016 2:47 PM

• Parasites of bovids, birds, reptiles and fishes

• C.parvum - cryptosporidiosis
• Intracellular parasite in
○ Distal jejunum
○ Ileum and colon
• Oocyst is the infective stage
• Commonly found in immunocompromised patients e.g. HIV patients and caused severe,
unrelenting diarrhoea
• It causes mild diarrhoea in the immunocompent, and is usually self-limiting

Epidemiology
• Those at high risk of infection
○ Immunocompromised
○ Young and old people
○ Those working with animals - the parasite mostly infects animals (birds, birds and
reptiles)

Morphology
• Oval/spherical, highly refractile, colourless (1.5 - 5µm)
• The cysts are composed of 4 motile sporozoites
• 2 forms of oocysts
○ Thick walled - found in faeces
○ Thin-walled - responsible for auto-infection
• Are the only parasites that stain acid-fast

Life Cycle
• Monoxenous life cycle
○ Completes lifecycle in one organism
• Faeco-oral infection

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Pathology and Pathogenesis
• Oocysts are the infective stage and can be transmitted faecal-orally through contaminated
food or water
• The oocytes, particularly the thick-walled oocyst (the thin walled oocyst is responsible for
auto-infection), is ingested and sporulate to release the sporozoites which attach to the
brush border of the small intestinal wall and cause damage to it, and causes diarrhoea and
subsequent pain
• They may also invade the colon and cause colitis

Clinical features
GI infection
• Self-limiting in immunocompetent patients
• Watery diarrhoea is the most frequent symptom
• Can be accompanied by
○ Dehydration
○ Weight loss
○ Abdominal pain
○ Fever
○ NV
Extra-intestinal infection
• Respiratory cryptosporidiosis

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 • Respiratory cryptosporidiosis
• Cholecystitis
• Hepatitis
○ In immunocompromised patients

Diagnosis
• Microscopy
○ Oocyst in stool, duodenal aspirates, sputum
• Sheather's sugar concentration technique (faeces)
• Staining
○ Auramine staining
○ Modified Ziehl-Neelsen
○ Immunofluorescence staining
• Others are such as serology, PCR

Treatment
• Self-limiting
○ Supportive treatment
 Rehydration in the case of diarrhoea
• Nitazoxanide
○ In immunocompromised patients
• ARV treatment
○ To increase CD4 count in HIV patients
• Spiramycin
○ Macrolide drug
○ Not

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Trichomonas Vaginalis
Wednesday, 9 November 2016 9:37 AM

• Exists as a trophozoite only

• Fount in the genitourinary tract of man
• Considered a sexually transmitted protozoa
• Common in commercial sex workers (CSWs)

Life Cycle

• Do not survive in columnar epithelium

• Survive In fluid media
• Found in the vagina and the prostate in males
• Sexually transmitted
• Can be transmitted to the new-born and may affect the eyes and the genitalia of the baby
during parturition

Clinical presentation
• Mild vaginitis and vaginal discharge with large number of leukocytes and is liquid greenish-
yellow in females
• Strawberry cervix
○ Cervicitis that appears red due to capillary dilation
• In males, it causes mild/asymptomatic disease and causes itching and discomfort inside
penile urethra especially during urination

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 penile urethra especially during urination
• Transmission
○ Sexual intercourse
• Exact mechanism of pathogenesis is unclear, appears to be multifactorial

Laboratory Diagnosis
• Clinical history
• Vaginal fluid pH
○ 4.5 and higher in infection
• Microscopy
○ Trophozoites on a wet mount of sedimented urine, vaginal and prostatic secretions
and scraping by microscopy - motile trophozoite
○ Fluorescent microscopy
○ Fixed smears can be stained with Papanicolou, Giemsa, Leishman and PAS
• Culture
○ Culture on Trussel and Johnsons' medium/simplified trypticase serum medium used
to maintain bacteria free cultures of the flagellate
• Molecular methods
○ Nucleic acid hybridisation and PCR

Treatment
• Treat both partners
○ Parasite will be transmitted back and forth
• Metronidazole
• Tinidazole
• Ornidazole

Prevention and Control

• Health education
• Contact tracing, condom use, counselling and compliance (4Cs) as it is an STD
• Mass treatment (CSWs)
• Antenatal care practices (screening)

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Toxoplasma Gondii
Wednesday, 9 November 2016 11:05 AM

• Worldwide distribution
• Zoonotic parasite - infects cattle, birds, rodents, pigs and sheep and humans
• Final hosts (felidae family, cat)
• Intermediate host (mammals and birds)
• Intracellular parasite
• Toxoplasma is an opportunistic pathogen in humans
• Recently important due to the emergence of several outbreaks, prevalence of AIDS and
the improvement of diagnostic measure
• Causes toxoplasmosis
• Toxoplasmosis is the leading cause of abortion in sheep and goats
• Risk groups
○ Pregnant women
○ Meat handlers (food preparation)
○ Anyone who eats raw meat

Morphological forms
1. Tachyzoites (infective form)
2. Tissue cysts
3. Bradyzoit
4. Oocysts

Tachyzoite stage
• Rapidly growing stage observed in the early stages of infection (acute stage)
• Found in the body fluids
• Crescent shaped
○ One end is more pointed than the other subterminal placed nucleus
• Asexual form
• It can be found in phagocytic and non-phagocytic cells

Tissue Cysts
• Tissue cyst is round and oval in appearance and the cystic wall created by the parasite is
thin, but firm and elastic
• The protozoa multiply in the tissue cyst slowly and repeatedly
• The trophozoites in the cyst are called bradyzoites, which are similar to the tachyzoites,
but smaller

Bradyzoites
• Are slow-growing stage inside the tissue cysts
• Bradyzoites mark the chronic phase of infection
• Bradyzoites are resistant to low pH and digestive enzymes during stomach passage
• Protective cyst wall is finally dissolved and bradyzoites infect the tissues and transform
into tachyzoites
• Bradyzoites are released in the intestine and are highly infective if ingested

Oocysts
• Found in the faeces of the cat
• Cat ingests tissue cysts containing bradyzoites
• Gametocytes develop in the small intestine
• Sexual cycle produces the oocyst which is excreted in the faeces
• Oocysts appear in the cat's faeces 3 - 5 days after infection

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 • Oocysts appear in the cat's faeces 3 - 5 days after infection
• Oocysts require oxygen and the sporulate in 1 - 5 days
• The oocysts is non infectious before sporulation
• Unsporulated oocysts are sub-spherical to spherical
• Sporulated oocysts are sub spherical to ellipsoidal
• Each oocysts has two ellipsoidal sporocysts
• Each sporocysts contains 4 sporozoites
• Shedding occurs 3 - 5 days after ingestion of tissue cysts
• Sporulated oocysts remain infective for months

Morphological forms in the cat

• Schizont
○ Can be found in the small intestinal mucosa of the infected cat
○ Schizonts contain about 4 - 40 merozoites
• Gametocytes
○ Are also found in the small intestinal mucosa of the cat
○ Male gametocytes produce 12 - 32 male gametes crescent in shape
○ The female gametocyte develop into _
○ The male and female gametes fertilise to form a zygote that develops into an oocyst

Transmission
• Eating undercooked meat of animals harbouring tissue cysts
• Consuming food or water contaminated with cat faces or by contaminated environmental
samples (such as faecal-contaminated soil or changing the litter box of a pet cat)
• Blood transfusion or organ transplantation
• Transplancentally from mother to foetus throughout pregnancy but the outcome depends
on the trimester.
• If the mother has a chronic infection, the parasite is mostly in tissue form so not really
infectious. Additionally, she has antibodies to protect the foetus.

Life Cycle

• The extra-intestinal phase begins when the oocyst in cat stool, or the meat or animal
viscera containing cysts or pseudocyst are ingested by an intermediate host (man, pig,
cattle, sheep, mouse, etc.) sporozoites, bradyzoites or tachyzoites are released in the host

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 cattle, sheep, mouse, etc.) sporozoites, bradyzoites or tachyzoites are released in the host
intestine
• Parasites penetrate the intestinal wall. They actively invade or are ingested by the
mononuclear phagocytes and develop and inhabit them
• The ability and efficacy of the cell invasion is varied in different strains of T. gondii
depending on the virulence
• Binary fission and endodyogeny results in the production of tachyzoite, encircled by the
host cell membrane called pseudocysts
• When tachyzoites increase more than 10 per cell, the host cell breaks and parasites
released, that in turn invade new cells, develop and multiply
• In an immunocompetent host, these tachyzoites develop into cysts especially in the cells
of the brain, eye and skeletal muscles and can survive for several years
• Several hundred bradyzoites can be observed in a cyst
• Under immunocompromised conditions, cysts rupture, deliver many bradyzoited -
tachyzoited which infect more cells
• The ability of invasion and multiplication of T. gondii and the host immune status are in
dynamic equilibrium and as a result, tachyzoites can transform into bradyzoites and vice
versa

Definitive Host
• When food or water contaminated with T. gondii oocysts or animal body containing cysts,
pseudocysts ingested by feline, sporozoites, bradyzoites or tachyzoites are released into
the small intestine
• The parasites invade the epithelial cells and 3 - 7 days later, transform into multinucleated
schizont
• _

Pathogenesis
• Tachyzoite is the principle stage responsible for pathological changes and causes either
○ Congenital toxoplasmosis - the parasite is transmitted transplacentally
 When non-immune mothers (usually during primary infection) are infected
during pregnancy and is of great severity to the foetus as the mother has not
developed antibodies against the parasite, that she can pass down to the
foetus. This may lead to stillbirths, intracerebral calcifications, psychomotor
disturbances, hydrocephaly and microcephaly
 Infection in the first trimester generally results in stillbirths or major CNS
anomalies
 Infection in the second and third trimester results in less severe neurological
complications and more common
○ Postnatal toxoplasmosis - parasite is transmitted through ingestion of undercooked
infected meat, blood transfusion, faecal-orally especially in people that handle cats
 Are relatively asymptomatic but can cause fatal infection in the
immunocompromised
 The tachyzoite directly destroys the cell (causing necrosis and fibrotic scars)
and has high tropism for parenchymal cells and those of the RES and can
cause low-grade lymph node infection
 When a tissue cyst ruptures, it releases numerous bradyzoites and cause a
hypersensitivity reaction that leads to inflammation > blockage of blood
vessels > cell death and necrosis near the damaged cyst
• Tachyzoite is the principle stage responsible for pathological changes
• After entering the host cell, tachyzoites multiply, break the cell and the released
tachyzoites invade new cells once again
• The process repeats itself and the foci of oedema and necrosis surrounded by
lymphocytes, monocytes and plasma cells are formed
• The pathological changes can be divided into 3 categories
○ Necrotic foci from cell destruction by the quickly multiplying tachyzoites. These foci,
later form fibrotic scars with the cysts present all around
○ Cysts normally do not induce inflammation, but if they rupture, the released

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 ○ Cysts normally do not induce inflammation, but if they rupture, the released
bradyzoites are cleared by the host immune system, but some survive and cause
delayed hypertensive reaction, which results in the formation of a granuloma and
scarring
○ Secondary pathological changes initiated by focal damage can result inflammation of
blood vessels > blockage > tissue infarction, usually in the brain
• After infection of the intestinal epithelium, the organisms can spread to other organs like
the brain, lungs, liver and eyes
• This spread controlled by cell-mediated immunity, but circulating antibodies also enhance
killing of the parasite

Clinical Presentation
• Toxoplasmosis is an otherwise healthy person may have no symptoms or only a few
lymph nodes usually in the patient's neck
• Toxoplasmosis in a person with a weakened immune system
○ Symptoms of swollen glands
○ Pulmonary infection
○ Encephalitis
○ Fever
○ Seizures
○ Headache
○ Psychosis
○ Headache
○ Problems with vision, speech, movement and thinking
• Toxoplasmosis is one of the most severe complications of AIDS, leading to primary
encephalitis
• Children born with toxoplasmosis, which accounts for about 98% of cases, may show
symptoms including
○ Fever
○ Swollen glands
○ Jaundice
○ Hydrocephalus or microcephally
○ Rash
○ Bruises or bleeding under the skin
○ Anaemia
○ Enlarged liver or spleen
○ Seizures
○ Limp muscle tone
○ Mental retardation
○ Hearing loss
○ Vision problems (toxoplasmosis of the eye)

Congenital Toxoplasmosis
• Occurs when mother is infected during pregnancy
• Infection during first trimester, the outcome is often severe
• Second trimester infection, some babies will show symptoms months or years after birth
• Third trimester infection is usually non-significant
• If mother infected before pregnancy, parasites will be in cysts form and no trophozoites to
cross the placenta
• If mother having previous infection, re-infected during pregnancy, will have antibodies and
will not transmit parasites to the child
• Manifestations are similar to those of children infected with T. gondii
○ Intracerebral calcification - encephalitis
○ Chorioretinitis - ocular toxoplasmosis
○ Deafness
○ Hydrocephaly

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 ○ Hydrocephaly
○ Microcephaly
○ Convulsions
○ Mental retardation
○ Cardiomegaly

Lab Diagnosis of Toxoplasmosis

• The demonstration of the T. gondii tachyzoites in blood, body fluids including amniotic
fluid, aqueous humour, CSF or cysts in tissue
○ Detection of T. gondii antigen in blood or body fluids by enzyme-linked
immunosorbent assay (ELISA) technique
○ Ab tests
 The Sabin-Feldman dye tests
□ A sensitive and specific neutralisation test. I
□ It measures IgG antibody and is the standard reference test for
toxoplasmosis
□ High titres suggest acute disease
 IgM fluorescent antibody test
□ Detects IgM antibodies within the first week of infection, but titres fall
within a few months
○ Molecular
 PCR on body fluids, including CSF, amniotic fluid and blood
○ Skin test
 Delayed skin hypersensitivity to T. gondii antigens
○ Antibody levels in aqueous humour to CSF may reflect local antibody production and
infection
○ Animal inoculation
 Inoculation of suspected infected tissues into experimental animals
○ Culture
 Inoculation of suspected infected tissues into tissue culture
• Imaging
○ CT and MRI - ring enhancing lesions due to brain abscesses
• Brain biopsy
○ To differentiate between CNS lymphoma and brain abscess from T. gondii

Amniocentesis
• Done around 16th week of pregnancy
• A long needle is inserted into the amniotic sac and amniotic fluid is drawn

Treatment
• Disease is immunocompetent adults with lymphadenopathic toxoplasmosis is usually self-
limited - treatment rarely indicated
• If visceral disease is clinically evident or symptoms are severe or persistent, treatment
may be indicated for 2 - 4 weeks
• Persons with AIDS who develop active toxoplasmosis (usually toxoplasmic encephalitis)
need treatment that must be taken until a significant immunologic improvement is
achieved as a result of antiretroviral therapy; then low dose maintenance treatment
• Treatment for ocular diseases should be based on a complete ophthalmologic evaluation.
The decision to treat ocular disease dependent on numerous parameters including
acuteness of the lesion, degree of inflammation, visual acuity and lesion size, location and
persistence
• Prophylaxis for patients w/ CD4 count <100 and seropositive for IgG

Maternal and foetal infection

• Spiramycin is recommended (for the first and early second trimesters)
• Pyrimethazine/sulphadiazine and folinic acid (for late second and third trimesters)
• PCR is often performed on the amniotic fluid at 18 gestation weeks to determine if the

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 • PCR is often performed on the amniotic fluid at 18 gestation weeks to determine if the
infant is infected. If the infant is likely to be infected, then treatment with drugs such as
pyrimethamine, sulphadiazine, leucovorin (typical)
• Congenitally infected new-borns are treated with the above for 1 year

Ocular Treatment
• Pyrimethamine and sulphadiazine plus folinic acid to protect the bone marrow from the
toxic effects of pyrimethamine
• Pyrimethamine + clindamycin if the patient (mother patient ) has a hypersensitivity
reaction to sulphur drugs OR
• Cotrimoxazole
• Atovaquone and pyrimethamine + azithromycin
○ Not been extensively studied
• Treatment will not result in the elimination of the organisms from the eye. Since new
lesions can for if the organism reactivates, esp. during adolescent (carefully monitored)

Prevention and Control

• Cook meat thoroughly to kill the cysts
• Avoid proximity to cats
• Hand-washing before meals and after handling cat litter
• Use gloves when changing cat litter
• Regular treatment of cats
• Blood screening prior to transfusion
• Screening and treatment during pregnancy
• Treatment of cases
• Chemoprophylaxis in HIV patients
○ Cotrimoxazole
○ Fansidar
○ Pyrimethamine-dapsone
• Health education

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Malaria
Tuesday, 8 November 2016 11:19 AM

• Non-motile protozoal disease

• Transmitted by the Anopheles mosquito
• Mal-aria - Italian for 'bad air'
• Endemic in most tropical regions of the world
• 5 plasmodium spp causing human infection
○ Falciparum
○ Vivax
○ Malariae
○ Ovale
○ Knowlesi
• Falciparum is the most important, rapid, most death
• Accounts for >85% of malaria in E. Africa
• 500M exposed to endemic malaria
• 2 - 3 deaths/year (1 - 1.5M children mostly in Africa)

Risk Factors
• Most at risk of developing severe disease are
○ The poor (60% of deaths worldwide occur in the poorest 20% of the population, due
to lack of access to effective treatment)
○ Young children and infants
○ Pregnant women (esp. primigravidae)
○ Elderly people
○ Non-immune (travellers, expatriates)

• Endemic malaria
○ Areas of high transmission
• Epidemic malaria
○ Highland areas and semi-arid/arid areas
• Imported malaria
○ Travel and migration, any country
• Airport malaria
○ USA, Belgium, Switzerland, UK
 No history of recent travel to endemic areas
 Easily missed (dangerous)
• ? Infected mosquitoes entering non-endemic areas, biting people living around airports
○ Not common, spraying of aeroplanes before taking off

Life Cycle

• Mosquitos deposits sporozoites during blood meal, sporozoites > 30 minutes in

circulation > liver schizonts (with merozoites) [exo-erythrocytic schizogony] > blood >
RBCs trophozoites > schizonts (merozoites) [erythrocytic schizogony] > gametocytes >
picked by mosquitoes > gametes (male and female) > sexual reproduction > zygote >
ookinete > penetrate wall of stomach > oocysts > rupture > sporozoites > salivary glands
(sporogony)
• Hypnozoite stage (sleeping/resting stage)
○ Ovale and vivax
○ Responsible for relapsing malaria

Human Cycle (Schizogony)

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Human Cycle (Schizogony)

Human infection comes through the bite of the infective

female Anopheles mosquito

 The sporozoites, which are infective forms of the parasite are present in the salivary gland of the
mosquito.
 They are injected into blood capillaries when the mosquito feeds on blood after piercing the skin.
 Usually, 10–15 sporozoites are injected at a time, but occasionally, many hundreds may be
introduced.
 The sporozoites pass into the blood stream, where many are destroyed by the phagocytes, but
some reach the
 liver and enter the parenchymal cells (hepatocytes).

Pre-erythrocytic (Tissue) Stage or Exoerythrocytic Stage

 Within an hour of being injected into the body by the mosquito, the sporozoites reach the liver
and enter the
hepatocytes to initiate the stage of pre-erythrocytic schizogony or merogony.
 The sporozoites, which are elongated spindle-shaped bodies, become rounded inside the liver
cells.
 They enlarge in size and undergo repeated nuclear division to form several daughter nuclei; each
of which
is surrounded by cytoplasm.
 This stage of the parasite is called the pre-erythrocytic or exoerythrocytic schizont or meront.
 The hepatocyte is distended by the enlarging schizont and the liver cell nucleus is pushed to the
periphery.
 Mature liver stage schizonts are spherical (45–60 μm), multinucleate, and contain 2000–50,000
uninucleate
merozoites.
 Unlike erythrocytic schizogony, there is no pigment in liver schizonts. These normally rupture in
6–15 days and
release thousands of merozoites into the blood stream.
 The merozoites infect the erythrocytes by a process of invagination.
 The interval between the entry of the sporozoites into the body and the first appearance of the
parasites in
blood is called the prepatent period.
 The duration of the pre-erythrocytic phase in the liver, the size of the mature schizont, and the
number
of merozoites produced vary with the species of the parasite.
 Latent stage: In P. vivax and P. ovale, two kinds of sporozoites are seen, some of which multiply
inside
hepatic cells to form schizonts and others persist and remain dormant (resting phase). The resting
forms are
called hypnozoites (hypnos: sleep). From time to time, some are activated to becomes schizonts
and release
merozoites, which go on infecting RBCs producing clinical relapse.
 In P. falciparum and P. malariae, initial tissue phase disappears completely, and no hypnozoites
are found.
However, small number of erythrocytic parasites persist in the blood stream and in due course of
time,
they multiply to reach significant numbers resulting in clinical disease (short-term relapse or
recrudescence).

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Erythrocytic Stage

The merozoites released by pre-erythrocytic schizonts invade the red blood cells.

 The receptor for merozoites is glycophorin, which is a major glycoprotein on the red cells. The
differences in the glycophorins of red cells of different species may account for the species
specificity of malaria parasites.
 Merozoites are pear-shaped bodies, about 1.5 μm in length, possessing an apical complex
(rhoptery). They attach to the erythrocytes by their apex and then the merozoites lie within an
intraerythrocytic parasitophorus vacuole formed by red cell membrane by a process of
invagination.
 In the erythrocyte, the merozoite loses its internal organelles and appears as a rounded body
having a vacuole in the centre with the cytoplasm pushed to the periphery and the nucleus at one
pole. These young parasites are, therefore called the ring forms or young trophozoites.
 The parasite feeds on the haemoglobin of the erythrocyte. It does not metabolize haemoglobin
completely and therefore, leaves behind a hematin-globin pigment called the malaria pigment or
haemozoin pigment, as residue.
 The malaria pigment released when the parasitized cells rupture is taken up by reticuloendothelial
cells. Such pigment-laden cells in the internal organs provide histological evidence of previous
malaria infection.
 As the ring form develops, it enlarges in size becoming irregular in shape and shows amoeboid
motility. This is called the amoeboid form or late trophozoite form.
 When the amoeboid form reaches a certain stage of development, its nucleus starts dividing by
mitosis followed by a division of cytoplasm to become mature schizonts or meronts.
 A mature schizont contains 8–32 merozoites and hemozoin. The mature schizont bursts releasing
the merozoites into the circulation.
 The merozoites invade fresh erythrocytes within which they go through the same process of
development. This cycle of erythrocytic Schizogony or merogony is repeated sequentially, leading
to progressive increase in the parasitaemia, till it is arrested by the development of host immune
response.
 The rupture of the mature schizont releases large quantities of pyrogens. This is responsible for
the febrile paroxysms characterizing malaria.
 The interval between the entry of sporozoites into the host and the earliest manifestation of
clinical illness is the incubation period. This is different from prepatent period, which is the time
taken from entry of the sporozoites to the first appearance of malaria parasite in peripheral blood.
 In P. falciparum, erythrocytic schizogony always takes place inside the capillaries and vascular
beds of internal organs. Therefore, in P. falciparum infections, schizonts, and merozoites are
usually not seen in the peripheral blood.

Gametogony

After a few erythrocytic cycles, some of the merozoites that infect RBC's do not proceed to become
trophozoites or
schizonts but instead, develop into sexually differentiated forms, the gametocytes.

 They grow in size till they almost fill the RBC, but the nucleus remains undivided.
 Development of gametocytes generally takes place within the internal organs and only the mature
forms appear in circulation.
 The mature gametocytes are round in shape, except in P. falciparum, in which they are crescent-
shaped.
 In all species, the female gametocyte is larger (macrogametocyte) and has cytoplasm staining dark
blue with a compact nucleus staining deep red. In the smaller male gametocyte
(microgametocyte), the cytoplasm stains pale blue or pink and the nucleus is larger, pail stained

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 (microgametocyte), the cytoplasm stains pale blue or pink and the nucleus is larger, pail stained
and diffuse. Pigment granules are prominent.
 Female gametocytes are generally more numerous than the male.
 Gametocyte appear in circulation 4–5 days after the first appearance of asexual form in case of P.
vivax and 10–12 days in P. falciparum.
 A person with gametocytes in blood is a carrier or reservoir.
 The gametocytes do not cause any clinical illness in the host, but are essential for transmission of
the infection.

The Mosquito Cycle (Sporogony)

When a female Anopheles mosquito ingests parasitized erythrocytes along with its blood meal, the
asexual forms
of malaria parasite are digested, but the gametocytes are set free in the midgut (stomach) of mosquito
and undergo
further development.

 The nuclear material and cytoplasm of the male gametocytes divides to produce 8 microgametes
with long, actively motile, whip-like filaments. (exflagellating male gametocytes)
 At 25° C, the exflagellatation is complete in 15 minutes for P. vivax and P. ovale and 15–30
minutes for P. falciparum.
 The female gametocyte does not divide but undergoes a process of maturation to become the
female gamete or macrogamete. It is fertilized by one of the microgametes to produce the zygote.
 Fertilization occurs in half to two hours after the blood meal. The zygote, which is initially a
motionless round body, gradually elongates and within 18–24 hours, becomes a vermicular motile
form with an apical complex anteriorly. This is called the ookinete (‘travelling vermicule’).
 It penetrates the epithelial lining of the mosquito stomach wall and comes to lie just beneath the
basement membrane.
 It becomes rounded into a sphere with an elastic membrane. This stage is called the oocyst, which
is yet another multiplicatory phase, within which numerous sporozoites are formed.
 The mature oocyst, which may be about 500 μm in size, bulges into body cavity of mosquito and
when it ruptures, the sporozoites enter into the hemocele or body cavity, from where some find
their way to the salivary glands.
 The mosquito is now infective and when it feeds on humans, the sporozoites are injected into skin
capillaries to initiate human infection.
 The time taken for completion of sporogony in the mosquito is about 1–4 weeks (extrinsic
incubation period), depending on the environmental temperature and the species.

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Clinical presentation

Simple Uncomplicated malaria

• Irregular fever with chills (tertian [48 hours] - Falciparum, vivax, ovale; Quartan [72 hours] -
malariae)
• Headache, NVD, loss of appetite, dizziness, muscle aches, joint pains, general malaise
• High temperature, jaundice, pallor, abdominal tenderness, splenomegaly, hepatomegaly

Severe complicated falciparum malaria

• Cerebral malaria, convulsions, hypoglycaemia, hyperpyrexia, severe anaemia, renal
failure, pulmonary oedema or acute respiratory distress syndrome, algid malaria (shock
secondary to bacteraemia or septicaemia), DIC (disseminated intravascular
coagulopathy), haemoglobinuria (black water fever), jaundice, hyperlactataemia , abortion,
premature birth, low birth weight babies
• Hyper-reactive malaria splenomegaly (HMS), previously tropical splenomegaly syndrome
(TSS)

Lab diagnosis
• Gold standard = thick and thin blood films
• Stain (Giemsa, fields, etc)
• Examine thick film first under microscope - put a drop of blood at centre of slide, spread to
size of a shilling coin, let it dry, stain, examine. Don’t fix
• If parasites are seen, thein film to confirm species
• Thin film_ blood at edge of slide, spread with second slide, allow film to dry, fix using
alcohol, stain, examine under microscope
• Giesma stain - best if few thick films, slow
• Field's stain - best if many thick films, very quick.
• Other diagnostic methods
○ Antigen detection- parasite-specific lactate DH in P. falciparum infection
 Fast, high sensitivity and specificity, but unable to quantify the parasite load,
distinguish parasite species and Ag may be detected 2 weeks post-Rx hence
not always active infection

Parasitology Page 79
 not always active infection
○ Quantitative Buffy Coat-centrifuge blood in capillary tubes pre-coated with acridine
orange (OA), stains parasite DNA, view under UV light microscope.
 High sensitivity, cant distinguish young P. falciparum and P. Vivax trophozoites
○ PCR - Experimental. Detects<10parasite/10microlitres blood. Expensive

Spread of Drug Resistance

• In Kenya, the previously used drugs include
○ Chloroquine- very effective, cheap, few SE
○ Amodiaquine- cross-resistance with chloroquine
○ Fansidar, metakelfin

Rx of uncomplicated P. falciparum malaria

• Artemisinins for resistant P. falciparum malaria
• Use combination. (ACTs)
• WHO prohibits oral monotherapy
• Options
○ Artemisinin + proguanil
○ Artemisinin+sulphur/pyrimethamine
○ Artemisinin+Mefloquine
○ Artemisinin + lumenfantrine (Coartem)

Rx of Severe P. falciparum malaria

• IV quinine in 5% dextrose drip
• Artesunate IM
○ Cure rate is similar to quinine
○ Easier to administer
○ Safe in pulmonary oedema
• Rx other existing conditions
○ Hypoglycaemia (glucose IV)
○ Anaemia (transfusion)
○ Convulsions (diazepam or paraldehyde)
○ Renal failure (dialysis)
○ Hyperpyrexia (antipyretics)
○ Pulmonary oedema (diuretics)

Rx od non-P. falciparum malaria

• Chloroquine is highly effective
• Resistance in some strains of vivax
○ Quinine
○ Artemesinine + lumefantrine
○ Atovaquone + proguanil (malarone)
• To prevent relapse, for all the above Rx options add primaquine which kills hypnozoites

Prevention
• Residual insecticide spray
• Health education
• Insecticide treated bed nets
• Mosquito repellents
• Mosquitoes larvicides
• Mosquito window screens
• Environmental MX
○ Clearing bushes
○ Filling trenches
○ Removal of old tyres, empty tins Etc

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 ○ Removal of old tyres, empty tins Etc
• Protective clothing
• Airconditioners
• Prophylaxis
○ Tourists, expats, missionaries
○ Sicklers
○ Splenectomised individuals
○ Pregnant mothers
○ ?Children under 5years
 Use mefloquine weekly
 Doxycycline daily
 Malarone
• Rx of malaria cases

Malaria in pregnancy
• Pregnancy- lowered immunity
• Severe malaria affects both mother and child
• Rx - Quinine , 1st Trimester: Coartem- 2nd and 3rd trimesters
• In malaria endemic areas
○ Intermittent preventive Rx in pregnancy (IPTp)
○ Sulfadoxine-pyrimethamine used
○ SP concentrates in placenta killing harboured infected RBC
○ SP given in 2nd and 3rd trimesters during ANC visits
○ Effective in reducing anaemia in pregnancy and LBwt babies

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Naegleria Fowleri
Tuesday, 8 November 2016 2:22 PM

• 47 species in genus Naegleria are known (only fowleri is known to be pathogenic to

humans)
• Fowleri consists of 4 stages
○ Feeding and dividing trophozoite (uninucleate [with prominent, centrally placed
nucleolus], moves using lobopodia)
○ Transitory flagellate with two or more flagella (amoeboflagellate)
○ Round cyst (double walled, during harsh conditions)
• Fowleri is a free-living thermophilic amoeba in soil, warm fresh water, thermal effluents of
power plants, hot springs
• Trophozoite feeds on bacteria (can be cultured)

Life Cycle

Pathology
• Infect through the nasal passages and attach to the nasal neuroepithelium
• Migrate across the cribriform plate to the brain via the olfactory nerves
• Proliferate in the brain tissue, meninges and CSF causing extensive damage to the CNS

Primary Amoebic Meningoencephalitis (PAM)

• Primarily in children and young adults
• Characterised by predominant acute meningeal inflammation
• Cerebral hemispheres
○ Usually soft
○ Markedly swollen
○ Oedematous
○ Severely congested
• Leptomeninges are severely congested, diffusely hyperaemic and opaque

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 • Leptomeninges are severely congested, diffusely hyperaemic and opaque
• Olfactory bulbs show haemorrhage necrosis and are surrounded by purulent exudate
• Cortex shows numerous superficial haemorrhagic areas
• Distribution of lesions
○ Base of the orbitofrontal and temporal lobes
○ Base of the brain
○ Hypothalamus
○ Pons
○ Medulla
○ Upper part of the spine
○ Midbrain

Clinical manifestation
• Sudden onset of bifrontal or bitemporal headaches
• High fever
• Nuchal rigidity (with positive Kernig's and Brudzinski's signs)
• NV, diplopia, photophobia, irritability, restlessness
• Neurological abnormalities (lethargy, seizures, bizarre behaviour, confusion, coma)
• Death within a week
• Ddx.: Acute pyogenic or bacterial meningoencephalitis

Virulence (in vitro)

• Amoeba produce sucker-like appendages or amoebastomes and 'nibble' away at the
tissue culture cells and destroy the cell monolayer by causing CPE
• Produce phospholipases A and B activity or a cytolytic factor - destruction of the cell
membrane
• Neuramidases or elastase activity - destruction of tissue culture cells
• Presence of perforin-like, pore forming protein
○ Lyses target cells
• Has a cytopathic protein
○ Triggers apoptosis pathway in susceptible tissue culture

Diagnosis
• Microscopic examination of CSF smear
○ Actively moving trophozoites
○ Stained with Giemsa or Wright stains (trophozoite-nucleus with a centrally placed
large nucleolus)
• PCR
○ Cultured amoebae from patients

Treatment/Management
• Almost all cases are fatal
• 9 year old girl aggressively treated
○ IV and intrathecal amphotericin B, IV and itrathecal miconazole and oral rifampicin
• In vitro and in vivo testing amphotericin B, azithromycin
• Amoebastatic compounds
○ Phenothiazine compounds, voriconazole
• Miltefosine

Control
• Adequate chlorination (residual chlorine level of 0.5 ppm) of swimming pool
• Avoid swimming in areas colonised by N. fowleri
• Behaviour change
○ Avoiding water related activities in warm freshwater
○ Holding the nose shut or using nose clips when taking part in water-related activities

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○ Holding the nose shut or using nose clips when taking part in water-related activities
in bodies of water
○ Avoiding digging in, or stirring up, the sediment when taking part in water related
activity

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Acanthamoeba
Tuesday, 8 November 2016 2:51 PM

• Several Acanthamoeba spp affect humans

○ Castellani
○ Culbertsoni
○ Hatchetti
○ Healyi
○ Polyphaga
○ Rhysodes
○ Astronyxis
○ Lenticulata
○ Divionensis
• Amoebid stage has spine like projections (acanthapodia) from the surface of the body
○ Is uninucleate with a centrally located, large, densely staining nucleolus
• Cysts are double walled (highly resistant)
○ Has an outer wrinkled exocyst and an inner stellate, polygonal, oval or round endocyst
• Easily cultivated

Life Cycle*

• Portal of entry may vary (ubiquity of Acanthamoeba in the environment)

○ Through the skin
○ Via the nasal sinuses
○ Oral route

Granulomatous Amoebic Encephalitis (GAE)

• An insidious and chronic infection of the CNS which might span from several weeks to months
• Increasing in prevalence (immunosuppression)
• Cerebral hemispheres are often oedematous
• Extensive haemorrhagic necrosis involving the temporal, parietal and occipital lobes
• Hydrocephalus develops - brain herniation and death
• Brainstem, cerebral hemispheres and cerebellum may show areas of haemorrhagic infarcts
• Biopsy/autopsy reveals multinucleated giant cells, trophozoites and/or cysts in the cerebral
hemispheres, brain stem, cerebellum and basal ganglia

Acanthamoeba Keratitis (AK)

• Painful vision threatening infection
• Occurs even in immunocompetent (corneal trauma, contact lens wear and use of contaminated
contact lens solution or cases)
• Characterised by inflammation of the cornea, severe ocular pain and photophobia, lacrimation
• Amoebae adhere to the corneal surface (specific receptors on the surface, calcium ions)
• They infiltrate the stroma and cause tissue necrosis
• Causes ulceration of the cornea, loss of visual acuity and eventually blindness and enucleation
• In early stages
○ The parasite invades and destroys the anterior cornea

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 ○ The parasite invades and destroys the anterior cornea
○ Trophozoites and cysts can be seen interspersed with the polymorphonuclear leukocytes
• During later stages
○ Ulceration, descemetocele formation and perforation of the cornea occur

Pathogenic mechanisms
• Ingests bits and pieces of host cells by phagocytosis
• Secrete enzymes which lyse the host cells

Endosymbionts
• Acanthamoeba spp harbour human pathogenic bacteria
○ E.g.
 Legionella spp
 Francisella tularensis
 Mycobacterium avium
 Burkholderia
 Vibrio cholerae
 Listeria monocytogenes
 Helicobacter pylori
 Afipia felis
 E.coli serotype 0157

Diagnosis
• Microscopy (histology)
• Molecular techniques (PCR)

Management
• A few patients have survived after therapy with multiple antimicrobial agents
• No single drug has been effective in clearing an infection Balamuthia mandrillasis - GAE*
• GAE
○ Pentamidine isethionate
○ Sulfadiazine
○ Pyrimethamine
○ Voriconazole
○ Miltefosine
○ Topical application of chlorhexidine and miltefosine
• AK
○ Chlorhexidine digluconate in 0.9% physiological saline and 0.1% propamidine isethionate

Control
• No method for GAE
• For AK
○ Removing contact lenses before any activity involving contact with water (including
showering, using a hot tub or swimming)
○ Washing hands with soap and water and dry before handling contact lens
○ Cleaning lens according to the manufacturers guidelines (e.g. using fresh cleaning or
disinfecting solution each time lenses are cleaned and stored, storing reusable lenses in the
proper storage case (clean as directed and replace the case after every three months)

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Isospora belli
Tuesday, 8 November 2016 3:18 PM

• Causes isosporiasis
• Cosmopolitan but has the most prevalence in Africa and South America

Morphology
• Elongated oocyst (22µm by 15µm)
• Has a 2 layered wall
• Has a sporoblasts which mature to form 2 sporocysts
• Each sporocysts has crescent shaped 4 sporozoites

Life Cycle
• Inhabits small intestines in humans, the only host
• 2 cycles within human host
○ Sexual sporogeny
○ Asexual schizogony

Life Cycle

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Clinical Features
• Incubation period
○ 7 - 11 days
• Most infections are asymptomatic
• Symptomatic
○ Malabsorption (benign self-limiting course)
○ Can be fatal in immunocompromised patients

Diagnosis
• Demonstration of the oocysts in faeces or duodenal aspirates/biopsy, enterotest
• Eosinophilia - in >50% of patients

Treatment
• Combined therapy
• Sulfadiazine + pyrimethamine
• Ceptrin - Trimethroprim + sulfamethoxazole

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Blastocystis Hominis
Tuesday, 8 November 2016 3:41 PM

• Cysts vary greatly in size (6 - 40µm)

• Thick-walled and found in the stool
• Faeco-oral transmission
• Cysts infect intestinal epithelial cells and multiply asexually
• Vacuolar forms (multivacuolar)
• Amoeboid forms

Clinical Features
• Asymptomatic and symptomatic
• Watery diarrhoea, abdominal pain, perianal pruritus and excessive flatulence

Diagnosis
• Demonstration
• of cyst-like stages in faeces (wet or stained)
• Concentration

Treatment
• Metronidazole
• Iodoquinol

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Pneumocystis Jirovecii (Carinii)
Tuesday, 8 November 2016 3:48 PM

• In humans and other animals

• Reclassified as a fungus
• Trophozoites are 1 - 5 µm
• Pleomorphic and contain a single nucleus
• Found in the lungs and other extra pulmonary specimens
• Cysts are thick walled, rounded (5 - 8 µm in size)
• Thin walled cysts also exists
• Cyst in 8 intracystic bodies

Life Cycle

Clinical manifestations
• The symptoms of pneumocystic pneumonia (PCP)
○ Dyspnoea
○ Non-productive cough
○ Fever
○ Chest radiography - bilateral infiltrate's
• Extra pulmonary lesions (in <3% of patients)
○ Lymph nodes
○ Spleen
○ Liver
○ BM
• Death in untreated PCP - increasing pulmonary involvement

Diagnosis
• Microscopy
○ Sputum
○ Tracheobronchial lavage

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 ○ Tracheobronchial lavage
• Tracheobronchial or lung biopsy
• Immunodiagnostics
• Radiology - scanning

Treatment
• Antifolate (trimethroprim + sulphamethoxizole), dapsone, atovaquone-primaquine

Parasitology Page 91
Sarcocystis Spp
Tuesday, 8 November 2016 3:27 PM

S.Hominis (S. Bovihominis or isospora hominis), S. suihominis, S. lindemanni

• Cause 2 distinct clinical entities
○ Intestinal sarcocystosis (S. hominis and S. suihominis)
○ Muscle sarcocystosis (S. lindemanni)

Morpholgy
• Exist in 3 forms
• Oocyst
○ Thick or thin walled
○ Colourless
• Oval sporocyst and has 4 banana shaped sporozoites
• Sarcocysts
○ Thick, striated wall, along the length of muscle fibres of cattle and pigs respectively

Life Cycle
• 2 hosts - human (Definitive host), cattle/pigs (intermediate hosts)
• Sarcocysts ingested by man in raw/undercooked beef or pork

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Diagnosis
• Demonstration of mature sporocysts or oocysts in stool
• The 2 spp are morphologically similar
• Serological tests
○ IHA
○ IFA
○ ELISA

Muscular sarcocytosis (S. lindermanni)

• Rare in humans
• Humans are the intermediate hosts, cats and dogs are definitive hosts
• Transmission
○ Ingestion of infective sporocysts in food or drink
• Sporocysts liberate sporozoites
○ Invade intestinal mucosa and disseminate to muscles

Life Cycle
• Similar to S. hominis except for pattern of hosts

Clinical features
• Muscular sarcystis often asymptomatic

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 • Muscular sarcystis often asymptomatic
• Localised muscular swelling
• Eosiniophilia (40%)
• Subcutaneous and muscular tissue and inflammation lasts several days

Diagnosis
• Muscle biopsy - sarcocysts
• Radiology - faint shadows
• Treatment - no specific treatment

Control
• Avoid ingestion of raw/undercooked beef or pork
• Avoid water and food contamination with faeces of dogs and cats and other carnivores

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Approach to Amoebiasis
Wednesday, 9 November 2016 9:53 AM

What parasites cause amoebiasis

• Entamoeba
○ Histolytica
○ Coli
○ Hartmanni
○ Gingivalis
• Dientamoeba fragilis
• Iodamoeba buetschlii
• Endolimax nana
• Entamoeba polecki

Other Amoeba
• Free living amoeba
○ Naegleri fowleri
○ Acanthamoeba
 Castellanii
 Culbertsoni
 Poliphage
 Rhysonde
○ Affect only immunosuppressed or chronically ill people inhaling cysts.
○ Naegleri attacks healthy people (swimmers in swimming pools, spas or gyms)

Importance of amoebiasis
• 10% of world population infected
• Infection rate increases with age and levels in adults (stable endemic)
• Man is the only source of infection
• Transmission is Faeco-oral
• Houseflies may act as mechanical vectors

Common feature
• Amoeboid in shape
• Nucleoid (4 - 8)
• Ectoplasm
• Inclusion bodies (rods, RBCs)
• Forms are
○ Cysts
○ Trophozoites

Common Pathology in E. histolytica

• Severe lytic necrosis by trophozoites
• Anoxia and acidic environs favoured
• Secondary bacterial infection rare
• Granuloma formation in tissues
• Immune response IgG and M

GIT pathological features

• Small, discrete, superficial lesions/erosions
• Muscularis mucosa affected
• Flask shaped ulcers, perforations
• Amoebomas, strictures

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 • Amoebomas, strictures
• Obstruction FIT
• Bleeding

Most common causes of bleeding

• Intestinal lesions/ulcers following gysentery or perforation
• Perforation of GIT/liver (rare)
• Anaemia may follow

Liver Pathology
• Portal vein entry, fulminant hepatitis (70% right lobe)
• Liver necrosis leads to abscess (anchovy sauce) solitary in 90% of cases
• Metastasis t other organs via blood
• Fibrosis leads to amoeboma

Genital and Skin pathology

• Perianal areas ulcerations (anus/cervix)
• Rapture from liver abscess
• Uterine cervix
• Fistula ruptures in surgery

History - GIT mainly

• Symptoms in the GIT
○ Dysentery - blood/stool/mucus
○ Abdominal pain, tenderness and bloating
○ Tenesmus
○ NV
○ Dizziness, fainting
○ Weakness
History - Others
• Sore tongue and mouth and bad smell
• Bleeding from ruptures sites of skin, mouth
• Diarrhoea with blood, constipation
• Generalised body pain
• Upper abdominal pain
• Dehydration
• Acute abdominal pain
• Occasionally fever, sweating
• Poor appetite
• Localised tenderness
• Organomegally
• Symptoms of difficulty in breathing

Laboratory investigations and diagnosis

• To establish the presence of cysts/trophozoites in stool
• To determine the extent of disease (radiological)/surgery/biopsy
• Culture of amoeba to demonstrate strains (Robinson or Diamond axenic medium)
• Serological analysis

Treatment (Chemotherapy)
• Tissue (systemic) drugs
○ Emetine
○ chloroquine
• Luminal drugs
○ Aminosidine

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 ○ Aminosidine
○ Paromomycin
○ Iodoquinol
• Both tissue and luminal
○ Metronidazole
• Cysts passers
○ Diloxanide furoate

Prevention of amoebiasis
• Prevention of amoebiasis in the individuals and in the community
• _
• In individuals
○ Health education
○ Toilets use training
○ Child welfare clinics
○ Chemoprophylaxis
• In the community
○ Health education
○ Chemoprophylaxis
○ High standard of hygiene
○ Attention to clean, safe water supplies and sanitation
○ Attention to all strategies for safe environment

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Balantidium Coli
Wednesday, 9 November 2016 2:19 PM

Geographical Distribution
• Worldwide, causes ciliate dysentery or balantidiasis
• Normal habitat is colon of pigs and monkeys, where the pig is the reservoir host
• It is the largest protozoal parasite inhabiting the large intestine of man

Morphology
• The parasite exists in 2 stages
○ Trophozoite stage found in dysenteric stool
○ Encysted stage, found in chronic cases or carriers

Life Cycle
CDC

Pathology
• Ulceration of the colonic mucosa
• Bleeding leading to ciliate dysentery, the ulceration may be deep up to serosa and
muscular layer
• Dissemination to the liver has never been reported

Diagnosis
• Stool examination
• Demonstration of haematophagous trophozoites in dysenteric stool is pathognomonic

Treatment
• Nitroimidazoles such as
○ Metronidazole
○ Tecnidazole
○ Secnidazole

Control
• Prevent contamination of food and drink with faeces containing cysts of B. coli, either from
pig or humans
• Proper vet care of pigs
• Water and sewage treatment
• Health education to farmers and those worker in the meat industry
• Treatment of infected individuals/carriers

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American Trypanosomiasis
Tuesday, 22 November 2016 11:42 AM

• Trypanosomes, flagellated haemoprotozoa

• T.cruzi - found in central and south america
• Causes Chagas disease
• Animal reservoir - armadillos, raccoons and oposums
• Vector borne - reduviid bugs
• T.rangelli also transmitted by reduviid bugs but is non-pathogenic

Morphology
• Trypomastigotes
○ Nucleus
○ Kinetoplast
○ Undulating membrane
○ Short and stout
○ C-shaped
• Amastigotes
○ 1 nucleus
○ Kinetoplast
○ No flagella
• Metacyclic trypamastigotes
○ Infective stage
○ Found in the vector
○ Non-dividing

Life Cycle

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 • Infection
○ Bite of reduviid bugs with metacyclic trypomastigotes in hind gut, bugs pass faeces
then is rubbed into the bite puncture when scratching
• M.trypamastigotes invade the muscles (skeletal and cardiac) and transform into
amastigotes (multiply by binary fission extensively form intracellular pseudocysts) >
epimastigotes then trypomastigotes, pseudocysts rupture to release trypamastigotes in
blood
• Some trypamastigotes invade cells then transform to amastigotes
• Reduviid bugs pick trypamastigotes > amastigotes (foregut) > epimastigotes (midgut) >
metacyclic trypamastigotes (hind gut)

Clinical Presentation
• Incubation period about 2 - 4 weeks
• Early stages
○ Chagoma
 Romanose Sign
□ Inflammation at bite
□ site
□ Oedema of eyelid
□ Conjunctivitis
○ Fever

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 ○ Fever
○ Skin rash on chest or abdomen
○ Lymphadenopathy
 Generalised or local
• Late stages
○ Cardiomegaly
○ Megaoesophagus
○ Megacolon

Diagnosis
• History
○ Geographical
○ Symptoms
• Physical examination for signs
○ Blood
 Fresh for motility
 Thick smear stained with Giemsa
 Concentration methods
□ Micro-haematocrit method
○ Serology
 ELISA
 IH
 IFATn
 Antigen detection technique
○ Xenodiagnosis (lab bred reduviid bugs fed on host, 2 weeks later examine stool for
tryps)
○ Biopsy of lymph nodes/muscle may find amastigotes

Treatment
• Nirfutimox
• Benzimidazole

Prevention and control

• Control of reduviid bugs
○ Environmental
○ Insecticides
○ Repellents
• Treatment of infected persons

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Babesia Spp
Thursday, 10 November 2016 11:12 AM

• Genus babesis has - 100 species

• Species is pathogenic to humans
○ Microti
○ Divergens
○ Duncani
○ MO-1
• Causes babesiosis
• Zoonotic
• Intermediate hosts
○ Human
○ Rodents
• Definitive host
○ Tick of the genus Ixodes
• Worldwide in distribution
• Europe
○ Divergens, esp. in patients who have had splenectomy
• In malaria endemic areas not well documented, likely confused with plasmodia
• N.america
○ Microti
○ Duncani
○ MO-1
○ Not associated with splenectomy patients

Transmission
• Vector
○ Tick of genus Ixodes
• Human to human
○ Blood transfusion

Life Cycle

Parasitology Page 102

 • Tick introduces sporozoites into man while taking a blood meal
• Sporozoites infect RBCsy
• Reproduce asexually (budding)
• Form into male and female gametes
• Gametes picked up by the tick at feeding
• In the tick, male and female gametes fuse > sporozoites (sporogony)

Clinical Presentation
• Incubation period 1 - 4 weeks
• Sporozoites multiplication responsible for clinical features
• Most cases are asymptomatic
• Symptoms worse in the elderly, the very young, splenectomised, immunosuppressed
states, and in B. divergens - causes infection
• Symptoms
○ Fever (up to 40.5℃)
○ Chills
○ Sweating
○ Malaise/fatigue
○ Muscle aches
○ NV
○ Sore throat
○ Dry cough
• Signs
○ Hepatosplenomegaly
○ Anaemia (haemolytic)
○ Thrombocytopoenia
• Organ failure

Diagnosis
• Microscopy of thick and thin blood smears stained with Giemsa
• Serology (indirect fluorescent antibody test (IFA))
○ Useful in differentiating between babesia and plasmodia, for disease when
parasitaemia low and for blood screening for transfusion
○ Cross-sensitivity possible hence species differentiation may be difficult in some
cases
• Molecular techniques
○ PCR

Treatment
• Infection by B. divergens often fatal if untreated
• Infection by B. microti can remit spontaneously
• Asymptomatic patients need no treatment
• Antiprotozoal treatment for symptomatic patients
○ Atovaquone + azithromycin
 Normal patient
○ Clindamycin + quinine for severe cases

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 ○
 Very young
 Geriatrics
 Immunocompromised
 Splenectomised
• Symptomatic treatment
○ Antipyretics
○ Blood transfusions
○ Exchange transfusions
○ Vasopressors

Prevention and Control

• Tick Control
○ Acaricides
○ Environmental
• Prevent tick bites
• Rodent control
• Blood screening before transmission
• Health education

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African Trypanosomiasis
Tuesday, 22 November 2016 11:03 AM

• Trypanosomes, flagellated haemoprotozoa

• T.b. gambiense - West and Central Africa; T.b rhodesiense - East and Central Africa
• T.b gambiense - man, pigs
• T.b rhodensiese - man, cattle, game animals
• T.b brucei - cattle
• Vector borne - Glossina (tsetse) flies

Morphology
• Trypomastigotes
○ Features include
 Flagella
 1 nucleus
 Kinetoplast
 Undulating membrane
• Epimastigotes
○ Features include
 Flagella
 1 nucleus
 Kinetoplast
 No undulating membrane
• Metacyclic trypomastigotes
○ Infective stage
○ Found in the vector
○ Non-dividing

Life cycle
• Man infected with the bite with Glossina flies (males and females) with metacyclic
trypomastigotes
• Metacyclic trypomastogotes transform to long, slender trypomastigotes forms which
multiply by binary fission
• Picked by the glossina flies in the blood meal; both males and female flies suck blood.

Development in Tsetse Fly

 In the midgut of the fly, short stumpy trypomastigotes develop into long, slender forms and
multiply.
 After 2–3 weeks, they migrate to the salivary glands, where they develop into epimastigotes,
which multiply and fill the cavity of the gland and eventually transform into the infective
metacyclic trypomastigotes.
 Development of the infective stage within the tsetse fly requires 25–50 days (extrinsic incubation
period).
 Thereafter, the fly remains infective throughout its life of about 6 months.

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Clinical Presentation

Pathogenicity

Antigenic Variation
Trypanosomes exhibit unique antigenic variation of their glycoproteins.
 There is a cyclical fluctuation in the trypanosomes in the blood of infected vertebrates after every
7–10 days.
 Each successive wave represents a variant antigenic type (VAT) of trypomastigote possesing
variant surface specific antigens (VSSA) or variant surface glycoprotein (VSG) coat antigen.
 It is estimated that a single trypanosome may have as many as 1,000 or more VSG genes, that help
to evade immune response. Besides this, trypanosomes have other mechanisms also that help
them to evade host immune responses.

1. Trypanosoma brucei gambiense

The illness is chronic and can persist for many years.

 There is an initial period of parasitemia, following which parasite is localized predominantly in the
lymph nodes.
 A painless chancre (trypanosomal chancre) appears on The skin at the site of the bite, followed by
intermittent fever, chills, rash, anaemia, weight loss, and headache.
 Systemic trypanosomiasis without central nervous ppsysteml
system involvement is referred to as stage I disease. In this stage, there is hepatosplenomegaly
and lymphadenopathy, particularly in the posterior cervical region (Winterbottom’s sign).
 Myocarditis develops frequently in patients with stage I disease and is especially common in T.
brucei rhodesiense infections.
 Hematological manifestations seen in stage I include anaemia, moderate leucocytosis, and
thrombocytopenia. High levels of immunoglobulins mainly immunoglobulin (Ig)M are a constant
feature.
 Stage II disease involves invasion of central nervous system. With the invasion of central nervous
system, which occurs after several months, the ‘sleeping sickness’ starts. This is marked by
increasing headache, mental dullness, apathy, and day time sleepiness. The patient falls into
profound coma followed by death from asthenia.

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 profound coma followed by death from asthenia.
 Histopathology shows chronic meningoencephalitis. The meninges are heavily infiltrated with
lymphocytes, plasma cells, and morula cells, which are atypical plasma cells containing mulberry-
shaped masses of IgA. Brain vessels show perivascular cuffing. This is followed by infiltration of
the brain and spinal cord, neuronal degeneration, and microglial proliferation.
 Abnormalities in cerebrospinal fluid include raised intracranial pressure, pleocytosis, and raised
total protein concentrations.

2. Trypanosoma brucei rhodesiense

 East African trypanosomiasis is more acute than the Gambian form and appears after an
incubation period of 4 weeks.
 It may end fatally within an year of onset, before the involvement of central nervous system
develops.
 Pathological features are similar in both diseases with some variations—
○ Edema, myocarditis, and weakness are more prominent in East African sickness.
○ Lymphadenitis is less prominent.
○ Febrile paroxysms are more frequent and severe.
○ There is a larger quantity of parasite in the peripheral blood.
○ Central nervous system involvement occurs early.
○ Mania and delusions may occur but the marked somnolence, which occurs in T. brucei
gambiense infection is lacking.

• Rhodensiense - fast (acute infection)

• Gambiense - slow (chronic infection)
• Incubation period about 2 - 3 weeks
• Early stages
○ Chancre at the site of the glossina bite
○ Lymphadenopathy
 Generalised or local.
□ Back of the neck (posterior triangle of the neck) - Winterbottom's sign -
gambiense
○ Irregular fever
○ Headache
○ Malaise
○ Joint pain
○ Jaundice
○ Pallor
 Signs of anaemia
○ Splenomegaly
○ Hepatomegaly
○ Pancarditis
 Congestive cardiac failure
□ Swelling of the feet
□ Facial oedema
○ Insomnia
• Late stages
○ Mental confusion
○ restlessness at night
○ Excessive sleepiness during the day
○ Muscle twitching and tremors
○ Slurred speech
○ Muscle wasting
○ Convulsions

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 ○ Convulsions
○ Coma
○ Death
○ Kerandal's sign
 Loss of elbow and knee jerk reflexes

Differences between West African an East African Sleeping Sickness

Characteristics West African East African
Organism T. brucei gambiense T. Brucei rhodesiense
Distribution West and Central Africa East and Central Africa
Vector Glossina palpalis group Glossina morsilans group
Reservoir Mainly humans Wild and domestic animals
Virulence Less More
Course of Disease Chronic (Late CNS invasion); months to Acute (early CNS invasion); less than 9
years months
Parasitemia Low High and appears early
Lymphadenopath Early; prominent Less common
y
Isolation in No Yes
Rodents
Mortality Low High

Diagnosis
• History
○ Geographical
○ Symptoms
• Physical examination for signs
• Laboratory
○ Blood
 Fresh blood for motility
 Thick smear
○ Serology
 ELISA
 Card agglutination test
○ Lymph node aspirate
 Wet, stained smear
○ CSF
 Morular cells of Motti
 Raised protein
 IgM
○ Culture of blood, lymph node aspirate, CSF (Weismann's medium) takes 7 - 10 days
○ Animal inoculation
 Guinea pigs, mice (6 - 50 days)

Treatment
• Early stage
○ Suramin
○ Pentamidine (gambiense only)
• Late stage
○ Melarsoprol (Mel B)

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 ○ Melarsoprol (Mel B)
○ DFMO (Difluoro methyl ornithine)
Prevention and Control
• Control of glossina
○ Insecticides (ground or aerosol spray)
○ Repellents
○ Traps
○ Screens
○ Odour attractants
○ Clearing of bushes
○ Sterile male release technique
○ Use of PPE
• Treatment of infective people
• Relocation of communities from endemic areas

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Medical Entomology
Tuesday, 20 December 2016 9:59 AM

Medical entomology:-
• Study of insects and other arthropods such as ticks and mites, which affect the health and wellbeing of
man by transmitting diseases. Such arthropods are said to be of medical importance and are thus
studied by medical entomologists.
• Some of the insects or arthropods do not transmit any disease pathogens, but cause considerable
nuisance and annoyance to man by their bites or stings which may be poisonous or provoke severe
irritation
• Others, such as house-dust mites may induce allergies, while a few, such as scabies mites live in the
superficial layers of man’s skin and are thus true parasites
• The arthropods outlined above may warrant attention of a medical entomologist; but the ones of great
medical importance are those that transmit diseases such as malaria, sleeping sickness, arboviruses
such as yellow fever, various types of filariasis e.t.c

Methods of disease transmission:-

• An arthropod is termed to be a vector when it transmits disease pathogens from one animal to
another – including man.
• The arthropod transmitting a disease pathogen is referred to as a vector
○ Accidental/mechanical vectors
 Mechanical transmission
○ Biological vectors
 Cyclical/biological transmission
□ Involves obligatory development and or multiplication of the disease pathogens in the
arthropod before delivery to subsequent host

Classification of Arthropods
Arthropods can commonly be classified into different sub-groups as shown in the classification tree below.

• The phylum arthopoda is the largest of the animal phyla; there are numerous classes under it, but
about five of these classes are medically important.

Why medical entomology?

• In tropical countries, the largest group of diseases is probably insect-borne.
• It is therefore, important to know the habits of the insect vectors and how they transmit diseases. It is
difficult to implement control measures of insects, without some knowledge of entomology and
specifically medical entomology.
• Medical entomology is therefore concerned with the study of arthropods (especially of insects) that are
of public health importance.

Some vector-borne diseases

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Some vector-borne diseases
• Transmitted by mosquitoes
○ Malaria
 Vectors:
□ Anopheles mosquitoes
 500 million become severely ill with malaria every year and more than 1 million die.
○ Yellow Fever
 Principal vectors:
□ Aedes simpsoni, Ae. africanus, and Ae. aegypti in Africa
□ Species in Haemagogus genus in South America
□ Species in Sabethes genus in France
 200,000 estimated cases of yellow fever (with 30,000 deaths) per year.
○ Dengue fever
 Vectors:
□ Aedes aegypti (main vector)
□ Aedes albopictus (minor vector)
 50 million people are infected by dengue annually, 25,000 die.
 Threatens 2.5 billion people in more than 100 countries
○ Chikungunya virus
 Vectors:
□ Mosquitoes Aedes aegypti
○ Zika virus
 Vectors:
□ Mosquitoes Aedes aegypti
○ O'nyong nyo'ng
 Vector:
□ Anopheles mosquitoes
○ Lymphatic filariasis
 Vectors:
□ Mosquito species in genera:- Culex, Anopheles, Mansonia, and Aedes
 Affects over 120 million people.
○ Japanese encephalitis
 Several mosquito vectors, the most important being Culex tritaeniorhynchus.
○ Rift Valley Fever (RVF)
 Vectors:
□ Mosquitoes in the genera Aedes and Culex
○ West Nile virus
 Vectors:
□ Vary according to geographical area; in the USA and are the main vectors.
 Culex pipiens (Eastern US)
 Culex tarsalis (Midwest and West)
 Culex quinquefasciatus (Southeast)
• Transmitted by Sand flies (Phlebotomus)
○ Leishmaniasis
 Vectors:
□ Species in the genus Lutzomyia in the New World and Phlebotomus in the Old World.
 Two million people infected.
○ Carrion's disease - Vectors: sandflies of the genus Lutzomyia.
• Transmitted by Tsetse flies (Glossina)
○ Sleeping sickness
 Vector:
□ Tsetse fly, not all species
 Sleeping sickness threatens millions of people in 36 countries of sub-Saharan Africa (WHO)
• Transmitted by Fleas
○ Bubonic plague
 Principal vector:

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  Principal vector:
□ Xenopsylla cheopis
 At least 100 flea species can transmit plague.
• Transmitted by Black flies (Simulium)
○ Onchocerciasis – river blindness
 Vectors:
□ Simulium damnosum
□ S. neavei
• Transmitted by Triatominae bugs
○ Chagas disease
 Vector:
□ Assassin bugs of the subfamily Triatominae.
□ The major vectors are species in the genera Triatoma, Rhodnius, and Panstrongylus
• Transmitted by Tabanid fly (Chrysops)
○ Loa loa filariasis
 Vector:
□ Chrysops sp.
• Transmitted by Ticks
○ Tick-borne relapsing fever- Caused by a spirochete Borrelia duttoni
 Vector:
□ Ornithodoros moubata
○ Boutonneuse fever - caused by Reckettsia conori
 Principal vector:
□ Rhipicephalus sanguineus
○ Lyme disease
 Vectors:
□ Several species of the genus Ixodes
○ Alkhurma virus (KFDV)
 Vector
□ Tick
○ Kyasanur forest disease
 Vector
□ Haemaphysalis spinigera
○ Babesia
 Vector:
□ Ixodes ticks.
• Transmitted Body lice
○ Louse-borne typhus (Rickettsi prowazeki)
○ Trench fever (Rochalimaea quintana)
○ Louse-borne relapsing fever (Borrelia recurrentis

Mosquito
General biology:
• Mosquitoes are distributed worldwide, distribution including Temperate and Arctic regions of the World.
• Do not appear only in the Antarctic regions
• Exist up to 5,500m above sea level
• In temperate areas mosquitoes are not disease vectors but are pests - biting nuisance
• In the US high amount of money is spent on control of mosquitoes as biting pest

Description of mosquito
• Classification
○ Family: Culicidae
Sub-family Anophelinae Culicinae Toxorhychitinae

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 Anopheles Culex Toxorhychites
Bironella Aedes
Chagasia Mansonia
Eretmapodites
Aedemyia
Hodgesia
Culiseta
Mimomyia
Malaya
Uranotaenia
Lutzia
Ficalbia
Haemogogus

External morphology: See photos on Google

• Mosquitoes possess only one pair of functional wings
○ A pair of knob-like structures called halteres -represent the hind wings
• Are slender and relatively small insects
○ Measure 4-6mm long - some species, however can be as small as 2 - 3mm while other may be
larger (up to 10mm)
• Legs are long and slender
• Mouthparts
○ Are collectively known as proboscis
○ Proboscis in mosquitoes is elongate
○ Projecting forwards
○ The parts:
 Labium
□ Long flexible and gutter-shaped, terminates in a pair of small flap-like structures called
labella -encircles all other components of the mouth parts
 Labrum
□ Is uppermost structure
□ Slender, pointed and grooved along its ventral surface
 Between the labium and labrum are 5 needle-like structures: -
□ Maxillae
 Toothed lower pair
□ Mandibles
 Finely toothed upper pair
□ Hypopharynx
 Single untoothed hollow stylet

Life History:
• Adult mosquito immerges from pupa mainly in the evening
• They shortly afterwards mate
○ One mating is enough for life
• Male inseminates sperms in female
○ The sperms are held in a receptacle called spermotheca
• Females produce eggs in masses which are fertilized by the sperms from the spermotheca
Note
 Male plugs the female with gelatinous sperm plug -preventing the female from mating with
any other male.
• Only females feed on blood
• Males feed on sugar meals in flowers - nectar and related materials
• Females need proteins for development of the eggs

Feeding habits
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Feeding habits
• Mosquitoes bite man and many other animals
• Some bite birds, exclusively transmitting viruses in birds
• Others bite amphibians, reptiles and even fish -as mud fish
• Choice of host is of epidemiological importance
• When do mosquitoes feed?
○ This depends on species
 Some feed by day while others feed in the night
○ Exact biting time range varies greatly with species
○ Biting period is of epidemiological importance

Where do mosquitoes feed?

• Inside or outside houses
○ Indoor /outdoor feeding is very important epidemiologically

Feeding cycle (gonotrophic cycle)

• Unfed ←Blood → fed (engorged)
• In the tropics after 24hrs blood meal is half digested and eggs start development
○ Mosquito is half gravid
• After 3 days from feeding, blood meal is totally digested and the eggs are fully developed ready for
laying
○ Mosquito is GRAVID

Where blood-fed mosquito rests when digesting blood meal and developing egg
• In the gonotrophic cycle, a blood fed mosquito rests for 2-3 days while digesting the blood meal and
eggs developing
• Resting could be indoors (endophilic) or outdoors (exophilic)
• Habit very important in mosquito vector control using Indoor Residual Spraying (IRS)

Where breeding takes place

• Variety of places and water bodies, e.g. swamps, storage tanks, domestic waste containers - tins,
broken pottery, in animal hoof marks, tree holes, and even in banana axils- very small collection of
water
• These depend on the mosquito species

Development cycle
• Egg (1-2 days) → Larva (4-14 days) → Pupa (2 days) → Adult (2-3wks)
• Speed of development depends on temperature
○ Slower in cold temperatures
○ Faster in warm tropical conditions
○ Adults may live for 2 - 3 weeks in the tropics and several months in temperate region

How far do mosquitoes fly?

• 1 – 2 kms - mainly from breeding sites to feeding grounds
• May be pushed by wind for many kilometers.

Important Biological Behaviour

• Blood meal source
○ Anthropophagic (Feeding on man) Zoophagic (feeding on other animals)
○ Most mosquitoes have a tendency oscillating between anthropophagic and zoophagic behaviour
○ If Anopheles species tends more towards anthropophagic feeding, it is a better vector of malaria
and lymphatic filariasis

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 ○
and lymphatic filariasis
○ Most vector species are placed between the 2 extremes
○ Blood source habit is very important in vector-borne disease epidemiology
• Indoor/Outdoor feeding
○ Endophagic (Feeding indoors) Exophagic (Feeding outdoors)
○ Most Anopheles species feed on both ends
○ Most malaria vectors are endophagic but also are exophagic
○ This habit is very important in mosquito vector control using ITNs (Insecticide treated bed nets)
• Resting places after blood meal
○ Endophilic (Resting indoors) Exophilic (Resting outdoors after feeding)
○ Some endophagic mosquitoes are exophilic and vice versa
○ This habit is very important in mosquito vector control - using IRS

Medical Importance
Malaria transmission
• Only mosquitoes of the genus Anopheles can transmit Plasmodium species causing malaria in man:
○ Plasmodium fulciparum - (malignant tertian)
○ P. vivax - (benign tertian)
○ P. malariae (quartan malaria)
○ P. ovale - (ovale benign tertian)
• Because the sexual cycle of the malaria parasite occurs in the Anopheles mosquito vector, it is
conventional to refer to the mosquito as the definite host, and man as the intermediate host
• Life cycle of malaria parasite in the mosquito vector:
○ Male and female gametocytes ingested by female mosquitoes during feeding on man
○ Gametocytes pass to the mosquito's stomach where they undergo cyclical development which
includes a sexual cycle termed sporogony
○ Here only gametocytes survive in the mosquito's stomach, all other blood forms of malaria
parasites are destroyed
○ Exflagellation:
 Male gametocytes (microgametocytes) extrude flagella which are the male gametes
(microgametes)
 The process is termed as Exflagellation
○ The microgametes break free and fertilize the female gametes (macrogametes) which are formed
from the macrogametocytes (female gametocytes)
○ As a result of fertilization a zygote is formed
○ The zygote elongates to become an ookinete
○ The ookinete penetrate the wall of the mosquito's stomach (gut wall) to reach its outer membrane
○ In the outer membrane of the mosquito's stomach the ookinete becomes spherical and develops
into an oocyst.
○ The nucleus of the oocyst divides repeatedly to produce numerous spindle-shaped sporozoites
○ When the Oocyst is fully grown (about 60 - 80μm) it ruptures to release thousands of sporozoites
into the haemocele of the mosquito.
○ The sporozoites are carried in the haemolymph of the mosquito to all parts of the body but most
of them end up penetrating into the salivary glands
○ At this stage the mosquito becomes infective and sporozoites can be inoculated into man the next
time the mosquito bites.
○ Some 70,000 sporozoites are estimated to be in the salivary glands of an infective mosquito
vector
○ Speed of development of the malaria parasites in the vector (exogenous or extrinsic cycle)
depends on temperature and the plasmodium species
○ Generally oocyts can be seen on the stomach walls of the vector about 4 days after the infective
blood meal
○ Oocyts are fully grown and rupturing after about 8 days
○ Sporozoites are found in salivary glands after 9 - 12 days
○ For Plasmodium species, specific cycle period in the vector at for example 24℃ will be:

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 ○
 P. fulciparum - 11 days
 P. vivax - 9 days
 P. malariae - 21 days
 P. ovale - 15 days

Filariasis vectors
• Certain species of Anopheles are vectors of lymphatic filariasis especially in the rural environment.
• An. gambiae and An. funestus are very efficient vectors of Wuchereria bancrofti in Africa while other
anopheline species also transmit Brugia malayi S.E Asia especially Malaysia.
• Culex quinquefasciatus is an efficient vector of bancroftian filariasis in urban environment.
• Development in Vector
○ Starts when the vector mosquito takes microfilariae in its blood meal from an infected person
○ Some of the ingested microfilariae shade their sheaths in the stomach of the vector
○ Bore through the stomach wall and migrate to the thoracic muscles to start development as L1
stage
 Transform to short, thick and inactive sausage form of L1 in 4-5 days
 Undergoes first moult giving rise to L2 (second larval stage).
 L2 grows rapidly increasing in length and width, and undergoes a second moult to yield
 L3 larval stage at 9 - 10 days.
 L3 grows very rapidly in length but not width becoming very active. This is the infective
stage.
 In 11 - 15 days the L3s move to the mouth parts - inhabit the labium
○ During the mosquito's feeding on the next host the infective larvae in the labium are deposited on
the surface of the skin at the point of bite
○ The infective larvae find their way into the human body through the puncture made by the
mosquito bite
○ Many infective bites are required for infection
○ High humidity is required for survival of the infective larvae before penetration into the host's body
○ Transmission is more possible in the very humid areas

Transmission of Yellow Fever:

• Typical transmission in Tropical Africa
• Forest transmission
○ Takes place in high trees within monkeys in the canopy
○ The yellow fever virus circulates in monkeys.
○ Transmission is by Ae. africanus which breeds in tree holes in the forest, and bites immediately
after sunset when the monkeys go to sleep
○ Transmits Yellow fever in monkeys in the forest canopy
○ Transmission cycle may go on unnoticed for a long time especially in Africa where monkeys are
tolerant to the virus
• Transmission in plantations
○ When monkeys from the forest canopy raid plantation for food, they come in contact with Ae.
simpsoni which breeds in leaf axils in the bananas, and feeds at day time
○ Ae. simpsoni feeds at lower level in banana plantations near forests
○ The mosquito picks the virus infection from the monkeys raiding the plantations
○ In the plantations it will bite man and transmit yellow fever brought down by the monkeys from the
forest canopy
• Domestic transmission
○ Man goes home with the virus infection, gets bitten by Ae. aegypti in the domestic environment
○ In the domestic environment the transmission of the virus takes place between man
 Vector - man leading to an outbreak of yellow fever

Dengue haemorrhagic fever

• Virus transmitted by:
○ Ae. aegypti - throughout the Tropics
○ Ae. albopictus - in Asia

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 ○ Ae. albopictus - in Asia
○ Ae. polynesensis - in the Pacific
• Four serotypes of Dengue fever virus exist
○ These are Type 1, Type 2, Type 3 and Type 4
• Virus has an intrinsic incubation period (in man) of 4 - 5days
• Viraemia lasts 1 - 7 days
• Extrinsic incubation period (in mosquito vector) is 5 - 30 days
• Virus normally not zoonotic
• Dengue haemorrhagic fever has the following characteristics:
○ Infection is common in children
○ Infection commonly occurs in sequential infections involving different several serotypes

Tsetse Fly (Glossina)

Classification (Glossina systemics)
• Family
○ Glossinidae
• Genus
○ Glossina
• Species
○ There are some 22 different species of tsetse flies living only in Africa.
• Sub-species
○ Species are further divided into subspecies
○ All species and sub-species of tsetse flies total 30

Species Groups
• The 22 species of Glossina are arranged into three (3)groups
• The species groups distinguishable by the structure of the male genitalia
• The structures can be seen in fresh specimens using X10 hand lens.
• The groups include:-
○ Morsitans group
 Glossina longipalpis
 Glossina pallidipes
 Glossina morsitans morsitans
 Glossina morsitans submorsitans
 Glossina morsitans centralis
 Glossina swynnertoni
 Glossina austeni
○ Palpalis group
 Glossina palpalis palpalis
 Glossina palpalis gambiensis
 Glossina fuscipes fuscipes
 Glossina fuscipes martinii
 Glossina fuscipes quanzensis
 Glossina tachinoides
 Glossina pallicers pallicera
 Glossina pallicers newsteadi
 Glossina caliginea
○ Fusca group
 Glossina nigrofusca nigrofusca
 Glossina nigrofusca hopkinsi
 Glossina fusca fusca
 Glossina fusca congolensis
 Glossina fuscipleuris
 Glossina haningtoni

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  Glossina haningtoni
 Glossina schwetzi
 Glossina tabaniformis
 Glossina nash
 Glossina vanhoofi
 Glossina medicorum
 Glossina severini
 Glossina brevipalpis
 Glossina longipennis

Distribution
• Tsetse distribution is restricted to the African continent
○ The Morsitans group species
 Present in much of the Savannah (grassy woodland) of Africa
 Distribution is restricted by the cold winter but not hot dry conditions
 Distribution may be restricted by scarcity of game animals on which they feed
○ The Palpalis group species
 Limited to the very humid areas of Africa
□ mangrove swamps, rain forests, Lakeshores and gallery forests along rivers.
 Are riverine species

Description of Tsetse fly

Appearance
• At rest the tsetse appears quite slim
• The wings are placed one over the other on the back
• Tsetse flies are nearly always some shade of brown or gray-brown
• The body
○ Made up of 3 main parts
 Head
 Thorax
 Abdomen
○ Head
 With a pair of large compound eyes
□ Each of the compound eyes is made of thousands of small units called ommatidia
□ Compound eyes are capable of detecting moving objects at 137 meters
 Also with 3 simple eyes (ocelli) placed at the top of the head
□ Simple eyes are also sensitive to light
□ But real function not known
 The antennae:
□ 2 antennae are placed at the front of the head in a depression between the 2
compound eyes
□ Each antenna has 3 segments; the 3rd segment is the largest and bears the ARISTA.
□ The 3rd segment also has 2 small holes leading to the olfactory pits
 Can smell air
 Antennae are thus organs serving the sense of smell
 The mouth parts
□ Are long narrow, and can pierce the skin of an animal for sucking blood as saliva is
passed down the mouth parts into the animal being fed on.
□ When fly is not feeding all mouthparts are held so that they point forwards from
beneath the head.
□ The parts
 A pair of maxillary palps - sheath protecting the delicate proboscis on haustellum
 Proboscis (Haustellum) - very narrow and made of 3 parts:
◊ Labium – with toothed at the end for cutting through the skin
◊ Labrum - tube through which blood is sucked
◊ Hypopharynx - tube through which saliva is pumped

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 ◊ Hypopharynx - tube through which saliva is pumped
○ Thorax
 Covered by stiff cuticle
 Has 3 pairs of legs attached to underside
 2 wings attached to the topside walls of the thorax
 Also has a pair of halteres just behind the wings.
 The wing:
□ The characteristic HATCHET CELL.
□ The Halteres:
 Vibrate when insect is in flight
 Help the fly to steer
 Is a sense organ of balance
○ Abdomen
 Has seven visible segments
 In males there is an additional structure - the HYPOPYGIUM, folded beneath the last 2
segments

Internal morphology
• The alimentary canal:
○ The food channel starts from the labrum, leading to the esophagus, via the proventriculus to the
crop
○ Proventriculus marks end of fore gut and beginning of mid gut.
○ It is the source of the peritrophic membrane
○ Membrane secreted by the epithelial cells of the anterior part of the proventriculus
○ Produced as very delicate, soft and almost fluid structure
○ Hardens as it passes down the gut to form a thin but relatively tough sleeve lining the mid gut.
○ The junction of the four malpighian tubes separates the mid gut from the hind gut
○ A slender pair of salivary glands originating from the head run the whole length of the tsetse fly
body
○ Anteriorly, the ducts from both glands unit in the head to form the common salivary duct which
passes down the length of the hypopharinx.
• The reproductive system (female)
○ Note structure of advanced uterus, typical mammalian structure of ovaries
○ Also note the spermathecae and the milk glands

Feeding habits
• Both males and females take blood meal
• Blood meal taken from a large range of hosts including man and a variety of domestic and wild
mammals, and also reptiles and birds.
• Tsetse flies take blood meal about every 2-3 days in optimal conditions - period may be shorter in dry,
hot conditions and even as long as 10 days in cool humid conditions.
• Feeding restricted to daylight hours.
• Vision plays important role in locating host.

Life cycle
• Tsetse flies unlike other flies which lay eggs, deposit larvae, one at a time
• Females are inseminated young
• They store the spermatozoa in the spermathaecae for rest of their life - repeat mating not necessary
• After insemination and after taking a blood meal a single egg in one of the ovaries completes
maturation
• Passes down the oviduct to the uterus where it is fertilized by spermatozoa from the spermathecae.
• Egg hatches within the uterus after 3-4 days and empty egg shell passed out through the genital orifice.
• The larva in the uterus is nourished by secretions from the milk glands
• Regular and adequate blood meals are necessary for continuous production of secretions from the milk
glands

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 glands
• Larval development completed in 4-5 days at which time the 3rd and final instar larva is 8-9mm long.
• The 3rd instar larva wriggles out, posterior end first, from the genital orifice - thus birth can be termed a
'breech case'
• Females select suitable places for larviposition.
• Once larva is deposited it commences to bury under 2-5 cm of soil.
• Once larva is deposited it commences to bury under 2-5 cm of soil.
• After 15 minutes the 3rd instar larval skin contracts and hardens to form a reddish-brown or dark brown,
barrel-shaped puparium - about 5-8mm long
• Pupal period may reach 4-5 weeks
• After pupal development is complete the fly immerges from the puparium, forces its way to the surface
of the ground and flies away.

Medical importance
• Probably any species of tsetse fly can transmit African human trypanosomiasis (HAT)
• But this is determined by the ability of the species to feed on man
• Many species rarely feed on man as well as on other mammals, reptiles and birds.

Life cycle of HAT parasites in the tsetse fly

• Trypanosomes sucked by male or female fly during blood meal from man or non-human reservoir
• Pass through the esophagus to the crop
• After feeding has ceased the trypanosomes in blood pass into the peritrophic tube lining the mid-gut.
• Blood gets digested in the mid-gut but trypanosomes are not destroyed. Instead they multiply.
• From the mid gut the trypanosomes migrate to infect the salivary glands, where they mature into
infective metacyclic forms (Trypanomastigotes)
• The trypanomastigotes are injected into the host when an infective fly feeds on new host.

Triatomine Bugs
- Belong to the Family Reduviidae
- Sub-family Triatominae
- The Sub-family comprises of 14 genera and 111 species
- Species of medical importance include:
• Rhodnius prolixus
• Panstrongylus megistus
• Triatoma infestans
• T. dimidiata
- All these are vectors of Trypanosoma cruzi (Chagas disease) in Central and south
America
- The Triatomine bugs are mainly rural
- Natural habitats include nests and burrows
- Have adapted to the domestic environment in close contact with man
External morphology (See diagrammatic representation)
- The bugs vary in size - from about 0.5 - 4.5 cm, but most are 2 - 3 cm long
- Easily recognized by their long snout-like head
- The head bears a pair of prominent dark coloured eye and long and thin 4 segmented
antennae
- The proboscis (or rostrum) relatively thin and straight, and as in bedbugs, lies closely
appressed to the ventral surface of the head
- When taking a blood-meal the proboscis is swung forwards and downwards -just as in
bedbugs
- Dorsal part of the 1st segment of thorax consists of a very conspicuous triangular
pronotum
- The mesothorax and metathorax are completely hidden dorsally by the folded fore-wings
(called hemielytra)
- Fore-wings are thickened at the basal end and membranous at the distal end

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- Fore-wings are thickened at the basal end and membranous at the distal end
- The hind-wings are entirely membranous and are completely hidden under the hemielytra
when bug is not flying
- The thorax has three long slender legs which end in a pair of small claws
- Abdomen more or less oval in shape - but covered by wings
Life cycle
Eggs:
- Deposited in or near the habitation of their hosts
- Such as cracks and crevices in walls,
- floors ceilings and furniture in houses
- and in animal burrows
- The eggs are about 1.5 - 2.5 mm long
- Pink, yellowish or white in colour
- Have a smooth shell
- Are oval in shape but slightly constricted before the operculum
- Females lay a total 50 - 1000 eggs - usually 200-300
- The life cycle is hemimetabolous
- Small nymphs which resemble the adults but wingless hatch from the eggs after 10-15
days - incubation may extent to 30 or even 60 days
The Nymphs
- Newly hatched nymphs remain hidden in the cracks and crevices for a few days before
they seek out blood-meals
- There are 5 nymphal instars each requiring at least one complete blood-meal
- Rudimentary wing pads start to appear in the 4th & 5th instars - only adults have fully
functional wings
Habits
- Nymphs and adults of both sexes feed at night on their hosts
- Feeding is a lengthy process lasting 10-25 minutes or more
- Feed on exposed parts of the body including nose, around eyes and even mouth
- Bite is usually relatively painless
- Many triatomine bugs defecate during feeding - very important in transmission of Chagas'
disease
- Many species also feed on a variety of wild animals such as armadillos, opossums, rats,
marsupials, squirrels, skunks, iguanas, bats and also birds
Medical importance
- Triatominae are of medical importance as they are vectors of Trypanosoma cruzi the
causative agent of Chagas' disease - American trypanosomiasis
- This is a zoonotic disease -essentially a disease of wild animals such as opossum,
armadillo and many species of rodents
- Affects over 20 million people
- T. cruzi is transmitted through faeces
The parasite development
- Parasites are ingested with a blood-meal
- Undergo entire development in the gut of the bug
- After 9 - 17 days sometimes longer the infective metacyclic trypanomastigotes of T. cruzi
are present in the lumen of the hind gut
- The metacyclic trypanomastigotes are passed out in faeces onto the skin of host
- Man becomes infected when the excreta is scratched in either abrasions in the skin, in the
site of the bug's bite, or when it gets rubbed into the eyes or other mucous membranes
Control
- Insecticides
- Residual indoor spaying on walls and roofs using HCH or dieldrin is effective against
triatomine bugs.
- Have developed resistance to DDT
- Alternative insecticides include propoxur and malathion
- Improvement of housing
- Smooth plastering of inside walls and floors
- Replacing thatch with corrugated iron sheets or tiles
PHLEBOTOMINE SANDFLIES

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PHLEBOTOMINE SANDFLIES
Family: Psychodidae
Subfamily: Phlebotominae
Genera: Phlebotomus
Lutzomyia
Sergentomyia
Brumplamyia
Warileya
- There are over 600 species in the 5 genera
- Species in three genera - Phlebotomus, Lutzomyia and Sergentomyia suck blood from
vertebrates
- The 2 genera - Phlebotomus, Lutzomyia are of medical importance as they contain
species that are disease vectors
Distribution
- Genus Phlebotomus, occurs in the Old World especially the Mediterranean region and
the Old World tropics.
- Plebotomus species mainly inhabit semi-arid and savanna areas
- Do not prefer forest areas
- The genus Lutzomyia is found only in the New World tropics
- Lutzomyia species occur mostly in forested areas of Central and South America
- Genus Sergentomyia is confined to the Old World
- Sergentomyia species commonly found in the Indian Sub-region
- Also occur in other areas as Africa and Central Asia
Sandflies are vectors of: -
1. Leishmaniasis
2. Sandfly fever- a viral disease
3. Bartonellosis (Carrion's disease) - a Parasitic disease caused by an organism called
Bartonella
External morphology
Adult Phlebotomine Sandflies are easily recognized by:
- Their minute size - 2-3mm in length
- Hairy appearance - body very hairy
- Are dull colour - yellowish-brown to black
- Have long stilt-like legs
- The mouth parts are short and inconspicuous
- Are adapted for blood sucking
Wings
- Wings held up in the air at about 45o when at rest
- Have simple venation
- Vein 2 in phlebotomine sandflies branches twice
Sex
- Males have genital claspers visible at end of abdominal segments
- Females have a more rounded up bottom
Life cycle
- Phlebotomine sand flies have a long life cycle
- About 40 days in the tropics
- About 120 days in the Mediterranean region
- Only females take blood meals
Eggs:
- Females lay about 15 - 100 egg singly
- Eggs laid in cracks, crevices and litter
- Egg are minute - 0.3-0.4mm long
- Are ovoid in shape
- Are usually brown or black and are patterned
- Require a moist microhabitat with high humidity
- Cannot withstand desiccation
- Egg hatch after 6-17 days under optimal conditions, but may be prolonged
The larva:
- Larvae are scavengers, feeding on organic matter as fungi, decaying leaves,

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- Larvae are scavengers, feeding on organic matter as fungi, decaying leaves,
semi-rotting vegetation, animal faeces and decomposing bodies of arthropods
- Larval habitat needs high humidity
- There 4 larval instars
- Mature larva will measure 3-6mm long
- Has a well defined black head
- Body is grayish or yellow and of 12 segments
- Body with characteristic matchstick-like hairs (bristles)
- The last abdominal segment bears 2 pairs of conspicuous hairs called the caudal
bristles
- Larval development period is 19-60 days, depending on temperature and
availability of food
Pupa:
- Pupa assumes an erect position in the habitat
- It has the remains of the larval skin with its match-stick-like bristles and the
caudal bristles attached to the end of the pupa -characteristic for identification
of sandfly pupa
- Adult emerges from pupa after 7 - 14 days
Adult behaviour
- Phlebotomine sandflies are weak flyers
- 74% of them may fly for only 250 meters
- Both sexes feed on plant juices and sugary secretions
- Females in addition, suck blood from a variety of vertebrates including livestock, dogs,
rodents, snakes, lizards, amphibians and birds
- In the Old world many Phlebotomus species bite man
- In Tropical Americas Lutzomyia species feed on a variety of mammals including man
- Biting is usually restricted to crepuscular and nocturnal periods
- But biting may take place during the day in dark places and during overcast days
- Most species are exophagic, but a few are endophagic
- Due to their short mouth parts they are unable to bite through clothing
- Adults rest in sheltered , dark and humid places, but on dry surfaces
- Domestic and peri-domestic species may have endophilic habits after feeding e.g.
Phlebotomus papatasi
Medical importance
Phlebotomine sandflies are important vectors of disease and in addition they are a biting
nuisance and the bites can cause severe and intolerable irritation in sensitized people. They are
vectors of:
1. Leishmaniasis
2. Sandfly fever
3. Bartonellosis
Leishmaniasis
- Diseases caused by Leishmania parasites
- Phlebotomine sandflies are the only known vectors
- The parasites are ingested by female sandflies with a blood meal
- In the gut of the fly the parasites develop a flagellum with which they attach to the gut
wall
- Multiply within the insects stomach
- Migrate to the anterior part of the mid gut
- From there the parasites invade the oesophagus
- After 4 - 12 days the infective forms of the parasite are found in the insect's mouth parts
- From here the parasites are introduced to a new host during subsequent feeding
- Previous feeding of the female phlebotomine sandflies on plant juices aid the survival of the
parasites in the insect's gut
SIMULIUM
Family: - Simuliidae
Generally referred to as black flies
Distribution: - Worldwide
Species: - About 1300 species
Genera: - Family has 12 genera

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Genera: - Family has 12 genera
- Important genera include: -
1. Simulium
2. Prosimulium
3. Austrosimulium
4. Cnephia
Medically important species: -
1. Simulium damnosum complex - Africa
2. Simulium neavei complex - Africa
3. Astrosimulium ochraceun - South America
4. Simulium metalicum - South America
Morphology:
Size: - Quite small insects - 1.5 - 4mm long
- Relatively stout bodied
- Rather humped at the thorax
- Mainly black in colour
- A pair of large compound eyes
- In females eye separated on top of head - Dichoptic
- In males eyes occupy the whole head and touch each other on top of the
antennae - Holochoptic
- Antennae are short and stout, and distinctly segmented
- Mouth parts short - do not penetrate deep in skin
- Only females bite
- Teeth on labrum stretch the skin and the rasp-like action of the maxillae and
mandibles cuts through it and rupture the fine blood capillaries producing a
small pool of blood.
- This method of feeding is suitable for picking up mf of Onchocercer volvulus -
occurring in the skin.
Life cycle
Eggs: -
- Pale when laid but darken to a brown or black colour
- measure 0.1 - 0.4mm long
- More-or-less triangular in shape but with rounded ends and smooth shell.
- Laid in fast flowing streams
- Laid on sub-merged objects e.g. rocks, vegetation
- Batches of 150 - 800 eggs are laid in sticky masses or strings
- Hatch within 1 day but 2-4 days is common
- Hatching may take many weeks in cold temperate climate
Larva: -
- 6-8 larval instars
- Mature larva is 5-8mm long
- Distinct from all aquatic larvae
- Larva does not swim
- Are sedentary on submerged vegetation, rocks, stones and other debris
- Attachment is by the posterior hook-circlet (anal sucker)
- But larva may change position by help of the proleg
- Larvae are filter feeders
- Development takes 1-2 weeks depending on species and temperature - may over
winter for several months
Pupa: -
- Mature larvae spin with silk produced by salivary glands, a protective cocoon
- Cocoon is slipper-shaped and brownish in colour
- Cocoon is firmly stuck to submerged substrata
- Pupa has a pair of prominent filamentous respiratory gills
- Pupal period in the tropics is 2-6 days
- Adults emerge rapidly to the water surface in a protective bubble of gas - which
prevents it from getting wet.
- On the surface they immediately take to flight
Adult: -

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Adult: -
- Both males and females feed on plant juices and sugary substances
- Only females take blood meals
- Biting occurs outdoors by day
- Engorged females shelter in vegetation
- Blood digested in 2-3 days in tropical conditions - 3-8 days in non-tropical
depending on temperature
- Females may fly for considerable distance looking for blood meal e.g. 60-100km
- Dispersed by wind for as far as 300km
Medical importance: -
1. Can cause serious biting problem as bites can be quite painful and may result
in intense irritation.
2. Vectors of Onchocerciasis
- The only vectors of human onchocerciasis
- Feeding habit of tearing and rasping the skin to rupture blood capillaries
when obtaining blood meals makes them suitable to ingest the skinborne
mf of O. volvulus.
Development of parasite in Vector
- Stars when the vector species of Simulium takes mf in its blood meal from an
infected person
- Some mf bore through the stomach wall and migrate to the thoracic muscles to
start development as L1 stage.
- Transform to short, thick and inactive sausage form of L1 in 4-5 days.
- Undergoes first moult giving rise to L2
- L2 grows rapidly increasing in length and width, and undergoes a second moult
to yield L3 larval stage at 9 - 10 days.
- L3 grows very rapidly in length but not width becoming very active. This is the
infective stage.
- Infective larvae migrate to the base of the proboscis, and escape to the surface of
the skin during subsequent feeding by the vector.
- They actively bore through the skin at the sight where vector is feeding to start
development in human host
CHRYSOPS
Belong to a group of large biting flies referred to as horseflies
Family:- Tabanidae
- Many genera
- 3000 species
- Most important genera include:-
- Chrysops
- Tabanus
- Heamatopota
- Chrysops mainly important vectors of Loa loa in West Africa- main
vectors: -
- Chrysops silacea
- C. dimidiata
Distribution:
Chrysops are found in temperate and tropical areas of the world
External morphology:-
- Medium sized, 5-25 mm long
- Head large and semi-circular when viewed from above
- Conspicuous pair of compound eyes
- Sexed by examination of their eyes - females are dichoptic while males
are holochoptic
- Antennae are relatively small but stout and consist of 3 segments, and
have no arista
- Antennae longer than any of the other horseflies
- Mouth parts of females are stout and adapted for biting
- Do not project forwards but point downward from head.
- Only females take blood meals

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- Only females take blood meals
- In life specimens when adults are at rest, wings stay open
- Wings with a brownish transverse band
- Abdomen usually broad and stout and dorso-ventrally flattened
- Abdomen covered by distinct coloured strips
Life cycle
Adult: -
- Males and females feed on naturally occurring sugar secretions but in
addition females bit a wide range of mammals, and a wide range of
amphibian & birds
- Also get blood meals from man
Eggs: -
- 100-1000 deposited mainly on underside of objects e.g. leaves, grass,
vegetation, plant stems, twigs, stones
- Oviposition site overhangs larval habitats
- Larval habitats are mainly, muddy aquatic or semi-aquatic sites
- Eggs firmly glued on the substrate in upright position
- Are covered with coating that is impervious to water (water-proof)
- Egg measure 1-2.5mm long
- Are curved and about cigar-shaped
- Hatch in 4-14 days depending on temperature and species
Larva: -
- After wriggling out of eggs the young larvae drop down on the
underlying mud or water
- Larvae are cylindrical and pointed at both ends
- Have a very small black head
- Have prominent raised tyre-like rings which encircle most of the
body segments
- Segments 4 - 10 have psudopods
- Last segment has a siphon
- Larvae live in mud, rotting vegetation, humus, damp soil, shallow
and muddy water
- Are scavengers- feed on detritus and a variety of dead decaying
vegetable and animal matter
- Prolonged larval development - 1-2 years and up to 3 years in
temperate species
- 4-11 larval instars
- Mature larva is 1- 6cm long
- Migrate to the periphery of the larval habitat to pupate
Pupa: -
- Partially buried in mud or soil
- 7-40mm long
- Brown and curved
- Ear-shaped spiracles laterally placed
- Pupal period lasts 5-20 days
Adult behaviour
- Feed during day light hours
- Feed especially in bright sunshine
- Locate prey by sight
- Are powerful flyers and may disperse for many kilometers
- Inhabit low lying marshy scrub areas or swampy woodlands - but some
species are found in more open savanna and grassland areas
- Do not enter houses to feed but Chrysops silacea is an exception to this
Medical importance:
1. Vectors of Loa loa in the equatorial rain forests
- Vector species include:-
a) Chrysops silacea
b) C. dimidiata.
2. Mechanical vectors of tularaemia (Pasteurella tularaemia) from horses, rabbits

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2. Mechanical vectors of tularaemia (Pasteurella tularaemia) from horses, rabbits
and other rodents to man - mainly Chrysops discalis in North America.
This is a less important disease.
Loiasis:
- The only important disease transmitted to man by tabanids
- Disease caused by a parasitic nematode called Loa loa
- Disease occurs in the equatorial rain forests of West Africa, Southern Sudan and
parts of Western Uganda
- Parasite produces diurnal periodic microfilariae which are found in the
peripheral blood of man.
- When ingested by the tabanid - Chrysops - they undergo a developmental cycle
in the fly.
- Mf penetrates the gut wall of the fly, and migrate to the abdominal and thoracic
fat bodies
- In the fat bodies mf develop and moult twice to give the infective larvae in 10 -
12 days- development more or less same as the typical filaria larval
development in the thoracic flight muscles in mosquitoes, Simulium and
Culicoides.
Control
- Very few practical control measures available
HOUSEFLIES AND STABLEFLIES
- Belong to Family Muscidae
- There are many genera and species in the family
- Those important from medical point of view are:
- Common housefly - Musca domestica
- Other houseflies – fannia and muscina
- Blood sucking stable fly - Stomoxys calcitrans
- Have a Worldwide distribution
- Non-biting flies are mechanical vectors of:
- Cestodes
- Nematodes
- Faecal bacteria, protozoa and viruses
- Biting stableflies can be mechanical vectors of trypanosomiasis
The common housefly (Musca domestica)
External morphology (see diagrams)
- Are medium sized non-metallic flies – 6 - 9mm long
- Colour varies from light to dark grey
- Have three-segmented antennae with arista
- The mouth parts (proboscis) specially adapted for sucking
- Withdrawn into the head capsule when not in use
- Can be extended vertically downwards in a telescopic fashion when the fly feeds
- Proboscis ends up in a pair of oval-shaped fleshy labellae, having very fine
channels called psuedotrachae through which food is sucked up
- Each pair of legs ends in a pair of claws and a pair of fleshy pad-like structures
called the pulvilli
- Pulvilli are supplied with glandular hairs
- These sticky hairs enable the fly to adhere to very smooth
surfaces
- The hairs are also responsible for the fly picking of up dirt and
pathogens
Life cycle
Eggs
- Females attracted to decomposing materials for egg laying
- 75-100 eggs are deposited together or in separate batches in cracks and crevices or
scattered over the surface
- 5-6 such batches may be deposited in a fly's lifetime
- Eggs are creamy white measuring 0.8-1.2mm long
- Are distinctly curved dorsally giving them a banana-shaped appearance
- Egg hatch after 6-12 hours

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- Egg hatch after 6-12 hours
- Eggs cannot withstand desiccation- die when they dry out
Larvae (see diagram)
- Are white-creamy in colour
- 12-segmented cylindrical and maggot-shaped
- Have a pointed head end with a pair of small curved mouth hooks
- Posterior end has conspicuous spiracles shaped like a letter D
- Each spiracle has a thick outer wall called the peritreme
- Larvae feed on decomposing and decaying organic matter
- Undergo 3 larval instars
- Mature larva measures 8-14mm long
- Larval development completed in 3-5 days - may be longer (7-10days) in less
favorable conditions
Pupa
- Prior to pupation the 3rd instar larvae migrate away from larval habitats to drier ground
- Larval skin contracts and hardens to form a dark brown barrel-shaped structure
- Puparium measures about 6mm long
- Stage lasts 3-5 days in warm weather
- May be prolonged to 7-14 days in cooler conditions
- Egg to adult takes about 49 days at 16oC, 21 days at 20oC, 16 days at 23oC, 9-10 days at
30oC and 8 days at 35oC
Fannia species
- Flies in this genus resemble houseflies
- But are smaller (6 - 7mm long)
- Arista is devoid of any hairs (see diagram)
Life cycle
Eggs
- 50 -100 egg laid in man's food and also in urine-soaked bedding of man
and animals, compost heaps, excreta and litter
- Eggs resemble those of housefly
- Hatch in 1-2 days
Larvae
- Are dorsoventrally flattened (see diagram)
- Have thin fleshy processes on the body segments
- Larval development takes 7-12 days
Pupa
- Puparium is brown in colour and similar in shape to the larva
- Puparial period is 7-10 days and adult flies emerge
- Life cycle is completed in 13-22 days in favorable conditions
The greater housefly (Muscina stabulans)
- World wide distribution
- Are about 7-10mm long
Life cycle
Eggs
- 150-200 egg laid scattered indiscriminately over the surface of decaying matter
- Hatch after 1-2 days
Larvae
- Resemble those of housefly
- Distinguished from those of housefly by structure of the posterior spiracles - almost circular
and not D-shaped as in houseflies (see diagram)
- Peritreme encircles 3 crescent-shaped spiracular slits
- Young larvae are omnivorous scavengers
- As larvae mature they become predacious, feeding on any other fly larvae in the same
breeding environment
Puparia
- Similar to that of Musca domestica
- Puparial period is 1-2 weeks
- Life cycle takes 4-5 weeks - may be shorter (20-25 days) in warmer weather
- Adults enter buildings to feed

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- Adults enter buildings to feed
- Behave in same way as M. domestica
FLIES AND MYIASIS
Myiasis is defined as invasion by dipterous fly larvae of organs, living, necrotic or dead tissues
of the body of man or other vertebrate animals
- The larvae for some period, will feed on living, necrotic or dead tissues of the host's body,
or on host's ingested food in the case of intestinal myiasis
- Obligatory myiasis- the larvae must live on a live host for at least a certain part of their
life
- E.g. larvae of Cordylobia anthropophaga are
- obligatory parasites of man and other animals
- Facultative myiasis- Larvae are normally free-living, often attacking carcasses, but under
certain conditions they may infect living hosts
- E.g. species of Calliphora, Lucilia, phormia and Sarcophaga which normally
breed in meat or carrion may cause facultative cutaneous myiasis in man by
infecting festering sores and wounds
- There is no obligatory intestinal myiasis in man
- Presence of maggots in the intestinal tract is due to accidental swallowing of egg
or larvae on food
- The maggots may survive for some time in the intestines
- There is no fly species which is adapted to cause intestinal myiasis in man
- But obligatory intestinal myiasis occurs in other animals
- Facultative urinogenital myiasis occurs in man
- Usually involves larvae of Musca or Fannia species
- Flies may be attracted by unhygienic discharge to lay egg near the genital
orifices
- When egg hatch the tiny larvae enter the genital tract
- Much pain may be caused by larvae in this tract
- Sometimes mucous and blood may be discharged
Obligatory myiasis
The non-metallic Calliphorids
1. Cordylobia anthropophaga
- Species is commonly known as the tumbu or mango fly
- Found only in Africa where it is widely distributed
- Adults are robust, relatively big flies (see diagram)
- About 9-12mm long
- Dull yellowish to light brown in colour
- Have 4 visible abdominal segments more or less equal in length
Life cycle
Eggs
- Females lay up to 100-300 eggs in batches on dry soil and sand
- Eggs are laid in places especially contaminated with urine and excreta of man, dogs,
rodents and monkeys
- Also lay egg on underclothes and nappies of babies placed on the ground to dry
- Egg are whitish in colour and banana-shaped
- Hatch after 1-2 days
Larvae
- Larvae attach themselves directly on the host
- Or may attach temporarily to washed clothing placed on the ground to dry
- May also get transferred to man if clothing is not ironed before wearing
- Once on host, powerful hook-like mouth parts enable a larva to bury itself completely
except for its posterior spiracles placed at the tip of the abdomen
- Spiracles remain in contact with the air
- Newly emerged larvae can live as long as 9-15 days on the ground in absence of a suitable
host
- The 1st instar larvae are minute and typical maggot-shaped
- 2nd instar larvae are club-shaped
- 3rd instar larvae are fat, broadly oval-shaped white maggots
- measure 11-15mm long

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- measure 11-15mm long
- are covered with numerous spicules (see diagram and picture)
- After 10-12 days the mature larva wriggles out of the boil-like swelling and falls to the
ground where it buries itself and turns into puparium
- Adults emerge after some 10 days
- Readily enter houses where they may lay egg
Medical importance
- Larvae of Cordylobia cause boil-like (furuncular) swellings on most parts of the body
- These boils do not contain pus
- To extract the larva, the small hole in the swelling is covered with medicinal liquid
paraffin
- This prevents the larva from breathing through its spiracles resulting in its wriggling a
little further out of the swelling to protrude the spiracles
- In so doing it lubricates the pocket in the skin, and the larva can be extracted by gently
pressing around the swelling
Metallic Calliphorids
- Include the New World screw-worms ( Cochliomyia hominivorax) and the Old World
screw-worms (Chrysomyia benzziana)
1. New World screw-worms (Cochliomyia hominivorax)
- Occurs in South-western USA through Mexicao to Argentina
External Morphology
- Adult flies are 8-10mm long
- Are metallic green to bluish-green in colour (see picture)
- Have three (3) distinct dark longitudinal strips on the dorsal surface of the thorax (see
diagram)
- Dorsal bristles on the thorax not well developed
Life cycle
Eggs:
- Females of Cochliomyia hominivorax lay batches of 10-400 egg on the edges of 2-10 day
old wounds
- Also on dried blood clots
- On diseased or healthy mucous membranes e.g. nasal passages, mouth, vagina
- On umbilicus in new-borne babies
- Egg hatch in 11-12 hours
Larvae:
- Newly hatched larvae are active and bury deeply into living tissues
- Feed gregariously
- Undergo 3 larval instars
- 3rd instar larvae formed after 2-3 days
- 3rd instar larvae are 15-17 mm long
- Typically maggot-shaped
- Distinguished from those of housefly by presence of distinct bands of spicules on
anterior margins of all body segments (see diagram)
- Larvae tend to penetrate deeply into tissues
- Infections near eye, nose and mouth can cause considerable destruction
- Destruction is accompanied by putrid smelling discharge and ulceration
- Larvae mature after 4-8 days
- Wriggle out of wounds or passages
- Drop to the ground, bury in soil and pupate
- Adults emerge some 7-10 days later
2. Old World screw-worm
- Found in genus Chrysomyia
- Species Chrysomyia benzziana
- Larvae are obligatory parasites in living tissues
- Occur throughout the tropics
External morphology
- Adults very similar to Cochliomyia
- But with 2 distinct longitudinal thoracic strips (see diagram)
- Larvae also very similar to those of Cochliomyia

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- Larvae also very similar to those of Cochliomyia
Life cycle
- Very similar to Cochliomyia hominivorax
Medical importance of screw-worms
- Both Chrysomyia benzziana and Cochliomyia hominivorax are obligatory parasites of
living tissues and cause myiasis in man
- Larvae can cause severe damage and should be treated immediately
Facultative myiasis
The green-bottles (Lucilia)
External morphology
- Mostly metallic or coppery-green in colour
- A little smaller (10mm long) than Calliphora
- Distribution - Americas, Europe, Asia, Africa and Australia
Life cycle
- Females mainly lay eggs on meat, fish, carrion and decaying or decomposing carcasses
- Also oviposite on or near festering and foul-smelling wounds of man
- Mature larvae bury in soil and pupate
- Cause facultative myiasis - commonly intestinal myiasis
Sarcophagidae
Genera: - Sarcophaga
Wohlfahrtia
- Cause myiasis
- Sometimes called flesh flies
- World-wide in distribution
Sarcophaga
External morphology
- Are large hairy non-metallic flies, measuring 10 - 15mm long
- Have three (3) prominent black longitudinal dorsal strips on the thorax (see diagram)
Life cycle
- Adults do not lay eggs
- Deposit 1st instar larvae as tsetse flies
- Larvae deposited in batches of 20-40 on decaying carcasses, rotting food, excreta and
wounds
- Larvae typical maggot-shaped
- Posterior spiracles situated in a deep pit (see diagram)
- Larvae of Sarcophaga not easy to differentiate from Wohlfahrtia
- If on wounds larvae do not cause much damage as they feed on necrotic tissues
- Used in maggot therapy for chronic wounds
NOT YET COVERED (Lectures being organized)
Cockroaches
- Are insects of the order Blattodea
- There are about 4,600 species.
- About 50 cockroach species out of these are associated with human habitats (domestic
pests)
- Three species are pests of medical importance, including:-
o Blattella germanica (the German cockroach)
o Periplananeta americana (the American cockroach)
o Balatta orientalis (the Oriental cockroach)
- They are common and hardy insects, and can tolerate a wide range of environments
from Arctic cold to tropical heat.
- Cockroaches have a worldwide distribution
- Aid in transmission of various pathogenic viruses, bacteria protozoa and helminthes
a) Periplananeta Americana
b) Blattella germanica
c) Balatta orientalis
External morphology:
- Are usually chestnut brown or black in colour
- Measure about 1.5 – 4.5cm long
- Dorsoventrally flattened

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- Dorsoventrally flattened
- Have a pair of long filiform antennae
- Mouth parts are developed for chewing, gnawing and scraping
- Have two pairs of wings
o In certain household species wings may be shorter in females than males
o In female Blatta orientalis the wings are very small and non-functional
- The segmented body is more or less oval and completely or partly hidden from view by the
folded overlapping wings
- In both sexes a pair of prominent segmented cerci arise from the last abdominal segment
Habits:
- Household cockroaches in the tropics hide during the day in almost any dark place i.e.
cesspits, septic tanks, sewers, dust-bins, cupboards, drawers, behind refrigerators,
cooking stoves e.t.c.
- They are nocturnal in habit – rarely seen during the day
- Are omnivorous and voracious feeders – any type of mans food is eaten
- Can eat anything including paper, clothes hair, shoes, dried blood, sputum, excreta, dead
insects and almost any animal or vegetable matter
- May live for 5 – 10 weeks without water and for many months without food
Life cycle:
- Eggs are laid encased in a brown bean-shaped case or capsule called ootheca.
- Ootheca contains 14 – 48 eggs
- Cockroaches are often seen running around with an ootheca partly protruding from the tip
of the abdomen
- The oothecae are deposited in cracks and crevices in the dark and secluded places
o In some species the oothecae are cemented to the surfaces as underside of tables,
chairs and beds
- A female may lay 4 – 90 oothecae in its life span of many months to a year
- Cockroaches have a hemimetabolous life-cycle:-
o Nymphs hatch from eggs after 1 – 3 months
o Young nymphs are very pale and delicate versions of the adults
o Older ones are darker and resemble the adults
o The nymphs are wingless, but wings develop in the later instars
o In Periplaneta americana there are as many as 13 nymphal instars lasting 2 – 3
months in ideal conditions
Medical importance:
- Medical importance based on the cockroach’s dirty habits of feeding indiscriminately on
both excreta and foods, and their practice of excreting and regurgitating their partially
digested meals over food
o Cockroaches have been suspected to aid in transmission of various illnesses as:-
▪ Pathogenic viruses as poliomyelitis
▪ Protozoa – Entamoeba hystolytica, Trichomonas hominis, Giardia
intestinalis and Balantidium coli
▪ Bacteria: - Escherichia coli, Staphylococcus aureus, Shigella dysentariae,
Salmonella typhi and S. typhimurium.
▪ Intermediate host to various nematodes – e.g. Enterobius vermicularis.
▪ Have been found naturally infected by with Toxoplasma gondi and
suspected to transmit the parasite to cats and possibly to man
LICE
ANAPLURA - contains the group of lice that infest man
Group comprises of three species:
1. Pediculus capitis - Head lice
2. P. humanus humanus - body lice - also referred to as P. coporis
3. Pthirus pubis - pubic lice
Body and head lice
- Are longer than the pubic lice
- Nearly the same in morphological appearance and difficult to separate
- Have claws at end of legs
- More or less world wide distribution

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- More or less world wide distribution
- Body lice are vectors of :
- Louse-borne typhus (Rickettsi prowazeki)
- Trench fever (Rochalimaea quintana)
- Louse-borne relapsing fever (Borrelia recurrentis)
External morphology
- Adults are small grayish and wingless
- Have soft leathery integument
- Dorso-ventrally flattened
- Males measure 2-3mm and females 3-4.5mm
- Head bears a pair of inconspicuous eyes and a pair of short 5 segment antennae
- Three pairs of legs stout and well developed
- Have a thick tibia with a small thumb-like-spine on its inner side at the apex
- Have a short tarsus with a curved claw
Mouth parts
- Louse mouth parts are different from those of other blood sucking insects
- Do not constitute a projecting piercing proboscis
- Consist of a flexible, sucking almost tube like mouth - the HAUSTELLUM
- The haustellum is armed on the inner surface with minute teeth which grip the host's skin
during feeding
- Needle-like stylets are thrust into the skin and saliva injected into the wound
- Blood is sucked into the mouth and passed into the stomach for ingestion
Biology
- Lice are hemimetabolous - nymphs and adults having same appearance and behavior
- Adults and nymphs live permanently on man
- Both sexes take blood-meals
- Feed 4 - 5 times a day
- Females lay 6 -9 eggs at a go and about 200-300 eggs in life of about 1 month
- Females lay eggs which are usually cemented at the base of the hair near the scalp for head
lice or on the fabric for body lice
- Cementing material very sticky - hard to remove eggs
- When hatching the nymphs leave the egg shells (nits) stack in position
- Duration of egg stage is about 6-10 days
- But egg discarded on clothes away from warmth of the body may take 2 - 3 weeks
Nymph
- Lice pass through 3 nymphal instars
- Nymphs take blood-meal from man just as adults
- Nymphal stages last 7-14 days and louse becomes adult
- But nymphal duration depends on whether or not clothing is worn all the time
- If removed at night nymphs are subjected to lower temperatures, thus slowed
development
PUBIC LICE (Pthirus pubis)
- Pubic lice are generally smaller than body and head lice
- Measure 1-2mm
- Are easily distinguished from body and head lice
- In pubic lice there is less differentiation between thorax and abdomen
- Body is as broad as long
- In pubic lice the front pair of legs is much more slender than the other two pairs
- Have crab like appearance - sometimes referred to as crab louse
Life cycle
- Very similar to that of the Pediculus
- Egg take 6-8 days to hatch
- Nymphal stages last 10 - 17 days
- Female lays a total of 150 -200 eggs which are slightly smaller than those of Pediculus
- Eggs are cemented onto the coarse hairs of the genital and perianal region of the
body
- Egg may be found on other areas of the body with coarse hairs
- Pubic lice are rarely found on the head
- Infestation is usually through sexual intercourse

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- Infestation is usually through sexual intercourse
Medical importance of Lice
1. Pediculosis
- Presence of body, head or pubic lice on a person
- The skin of people who harbor a large number of body lice may become pigmented and
slough, a condition known as vagabond's disease
2. Feeling 'lousy'
- Feeling of weariness, irritability or a pessimistic mood caused by repeated lice bite with
injection of lice saliva in the individual
3. Louse borne typhus
- Caused by Rickettsia prowazeki
- Transmitted from one person to another by body lice
- Ingested with blood meal from an infected person
- The rickettsiae in the louse stomach will penetrate into the lumen cell where they multiply
enormously causing the cell to distend and rapture
- Raptured cell releases large number of reckettsiae in the louse gut
- Man gets infected with typhus either by inhalation of the fine powdered dry faeces of the
louse
- Alternatively louse is crushed by persisted scratching and the reckettsiae in the gut are
released to enter human body through abrasions
4. Trench fever
- Caused by Rochalimaea quintana
- Ingested by louse while feeding on infected man
- The pathogens get attached to the walls of the gut cell and multiply - no penetration
- After 5-10 days the louse faeces becomes infected
- Man becomes infected by either crushing the louse, or by its faeces coming in contact with
skin abrasions or mucous membranes
5. Louse borne Relapsing fever
- Caused by a spirochaete Borrelia recurrentis
- Transmitted by body lice
- Ingested with blood-meal from an infected person
- Spirochaetes in the gut pass across the gut wall into the haemocele
- Multiply greatly, reaching enormous numbers
- Man becomes infected by crushing the louse, and the spirochaetes entering the body
through skin abrasions or mucous membranes
- Habit of crushing lice between finger nails or even with teeth can be very dangerous if louse
is infected
Control of Lice
Body lice
- Body lice can effectively controlled by infested people changing and washing clothing in
water hotter than 60oC
- 10% DDT can be used to dust clothing
- 1% HCH powder used where lice are resistant to DDT
- Other insecticides for use include: Malathion, temephos (Abate), Carbaryl (Sevin) and
propoxa (Baygon)
Head lice
- Shave hair
- 0.5% carbaryl emulsion
- Malathion emulsion
- Here insecticide emulsion is applied on the head and the hair washed
- This kills the adult lice the nymphs and even the eggs
- But insecticides used should be changed frequently - yearly to avoid development of
resistance
Pubic lice
- Shaving hair
- Treatment with emulsion of insecticides
FLEAS
- There are some 3000 species and sub-species of fleas
- Belong to about 200 genera

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- Belong to about 200 genera
- Only a few are important pests of man
- 94% of the known species bite mammals the remainder biting birds
- Fleas are generally distributed through out most of the world
- But a few have restricted distribution e.g. the plague flea in the tropics
External Morphology
- Adults are relatively small 1-4mm
- More or less oval insects
- Compressed laterally
- Vary in colour from light to dark brown
- Have no wings
- Three pairs of powerfully developed legs, with hind pair specialized for jumping
- Legs and most of body covered by bristles and body spines
Head
- Roughly triangular in shape
- Bears a pair of conspicuous eyes
- Also carries a pair of short three-segmented antennae -lie in depressions behind the eyes
- Mouth parts pointed downwards
- In some species the head also has a row of coarse, well developed tooth-like spines - the
genal comb
Thorax
- Has three distinct segments
- Prothorax
- Mesothorax
- Metathorax
- Posterior margin of the pronotum may bear the pronotal comb
- Above the middle pair of legs is located a sternite called the mesosternum
- The mesosternum in some species is divided into two parts by the mural rod
- These structures: the genal comb, the pronotal comb and the mural rod are used in
classification of the flea species
- E.g. the presence of a mural rod and, combined with the absence of both the genal and
pronotal combs, indicates the genus Xenopsylla
- Females have a distinct brownish spermatheca in the position of 6th - 8th abdominal segment
The alimentary canal (Adult fleas) (See diagram)
- Has spindle-shaped pharynx - through which sucked blood passes
- Pharynx is linked to a thin oesophagus
- The proventriculus is bulbous and provided internally with numerous backwardly projecting
stiff spines (important in mechanism of plague transmission)
- Has relatively large stomach (mid gut)
- Distal end of the stomach is connected to the hind gut
- Hind gut continuos with a small dilated rectum
Life cycle (See diagram)
- Both males and females take blood-meals
- Females lay eggs in debris, cracks or crevices around the host's environment
- May lay up to 300-1000 eggs in small batches of 3-18 a day
- Egg hatch within 2-14 days
- Minute legless larvae emerge from the eggs
Larva
- With a small blackish head with very small pair of antennae
- 13 distinct but similar segments
- The segments end up in a pair of finger-like ventral processes- the anal struts
- Larvae very active and avoid light
- Feed on any organic debris including the host's faeces
- In some species the larvae are scavengers- feed on any dead insects
- Undergo three larval instars
- Larval period may last 10 -21 days but may be prolonged to more than 200 days in
unfavorable conditions
- At end of larval period the larva spins a whitish cocoon around itself
- Cocoon made from silk produced by the larva's salivary glands

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- Cocoon made from silk produced by the larva's salivary glands
- Cocoon is sticky and is soon covered by fine particles of dust and organic debris
- Larva pupates within the cocoon
The pupa
- Pupal stage lasts 7-14 days in optimal conditions
- Adult emerges from the pupa on stimulation by vibrations caused by movement of host
- the life cycle from egg to adult emergence may take as short as 2-3 weeks under optimal
conditions
- But may be considerably longer - up to 20 months
Medical importance
1. Murine typhus
- Caused by Reckettsia mooseri
- Parasite infects man when the infected flea faeces are scratched into the skin abrasions
2. Plague
- Caused by the plague bacillus Yersinia pestis
- Transmitted by Xenopsylla cheopis
- Bacillus taken up by the flea when feeding on infected person or rodent
- The bacteria reach the proventriculus and the fore gut
- Multiplies enormously and blocks the proventriculus
- Flea gets problem in feeding
- When it tries to feed the muscular pressure pushes some of the bacilli into the incoming
blood-meal which is eventually regurgitated into new host - causing infection
- Flea with blocked proventriculus becomes starved and repeatedly bites in attempt to get
a blood - meal in the process infecting many new individuals
Silvatic plague
- Plague is usually a disease of wild rodents
- Circulates within the wild rodents
- Kills and reduces number of rodents
- When the population of wild rodents is reduced fleas take on the domestic rodents
which are in turn eliminated very fast
- In the domestic environment the fleas move to man as the next host
- This ends up in a plague outbreak in man
TUNGA PENETRANS (the jigger flea)
- Tunga penetrans is found in the tropics and sub-tropics
- Some times referred to as the chigoe flea
- Does not transmit disease to man but it is a nuisance because the females burrow into the
skin
- Adults of both sexes are very small - about 1mm long
- Have very compressed thoracic segments and very weak legs
Life cycle
- Egg are dropped onto the floors of houses
- Hatch within 3-4 days
- Larvae inhabit dirty and dusty floors
- Larval development completed within 10-14 days in favorable conditions
- Pupal period 5-14 days
- Complete life cycle in as short as 18 days
Adults
- Adults - newly emerged are very agile
- Jump and crawl around until they locate a suitable host -mainly man
- Both sexes feed on blood
- Male leaves host after the blood-meal
- Female after being fertilized burrows into the skin
- Soft parts of the skin
- Burrows whole body except for the tip of abdomen bearing the anus, the genital opening
and the large respiratory spiracles
- Continuos to feed in the embedded position
- The abdomen distends with developing eggs, and acquires enormous size
- On maturity 150-200 egg are passed out of the female genital opening, falling to the ground
Control of fleas

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Control of fleas
- Only a very small fraction of the flea population will be found on the host
- The rest including the eggs larvae and pupae in cocoons will be in the hosts environment
- It is thus more effective to treat the whole environment including
- Beds
- Kennels
- Rodent burrows
- Rodent paths (runways)
- House floors
- These areas should be treated with either insecticidal powders or lightly sprayed with
solutions of :
- 0.5% HCH
- 2% Malathion
- 0.5% Diazinon
- 2% Dichlorvos (DDVP)
- Insecticidal powders for flea control include :
- 5-10% DDT
- 1% HCH
- 0.5% Dieldrin
- These liberally applied to the floors of houses and rodent runways
TICKS
Classification:
Class: ARACHNIDA - Tick, mites and spiders
Order: ACARINA - Order contains ticks and mites
Family: 1. Argasidae (Soft ticks)
2. Ixodidae (Hard ticks)
ARGASIDAE (Soft ticks)
- Soft ticks (Argasidae) are distributed world-wide
- There are some 150 species belonging to 5 genera
- Medically important soft ticks belong to genus Ornithodoros
- Most important vector species belong to the Ornithodoros moubata complex
- Are vectors of tick-borne relapsing fever caused by Borrelia dutoni
External morphology:
- Adult appears dorsally like a leathery skin
- Have no scutum
- Are flattened dorso-ventrally and oval
- The integument is tough, leathery and wrangled
- Mouth parts, termed capitulum ('false head') are situated ventrally
- The 4 segmented palps are leg-like
- Chelicerae have smooth, not denticulated, sheaths
- The powerful cutting chelicerae have strong teeth at their tips
- Together with the hypostome, the chelicerae penetrate the host during feeding
- The hypostome is toothed
- Have 4 pairs of well developed legs terminating in a pair of claws
- The coxal organs (glands) open between the bases of the coxae of the 1st and 2nd pairs of
legs
- Males and females are very similar in external appearance
- Both sexes feed on blood and are disease vectors
Life cycle:
- Adult spends 10 - 30 minutes taking blood
- The tick takes a large blood meal 6-8 times its body weight
- The blood meal is essential for maturation of the ovaries and egg laying
Eggs:
- Female lays 15 - 100 eggs in batches scattered in different areas in or near the animal host's
home
- Eggs are laid in cracks and crevices in walls, floors, furniture or in mud, dust and debris
- Are coated with a protective waxy secretion from the Gene's organ (coxal glands)
- Eggs are heat resistant and can remain viable for many months under adverse conditions
- Eggs usually hatch within 1-4 weeks

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- Eggs usually hatch within 1-4 weeks
- Ticks have a hemimetabolous life cycle (larvae and nymphs superficially resemble adults)
- Eggs hatch in six-legged larvae
Larva:
- Are very active and search for host from which to take a blood meal
- Capitulum projects from body and can be seen from above
- Blood feeding lasts about 20 - 30 minutes
- Engorged larva drops from host to the ground
- After a few days the larva moults to produce an eight-legged nymph
- Larvae of Ornithodoros moubata complex differ from most other argasid ticks in that they
do not take blood meals
- Remain in egg shells after hatching
- Moult to produce 1st instar nymphs which crawl from the egg shell to seek their
1st blood meal
Nymph:
- Nymph also active
- Seeks host and feeds for 20 - 30 minutes before it drops to the ground
- Undergo 4-5 nymphal instars
- Each nymphal stage requires a blood meal before producing the next nymphal instar
- Life cycle often take about 6 - 12 months depending on:
- Tick species
- Temperature
- Availability of blood-meals
- Adult ticks can live for several years
- Up to 15 years in the laboratory
- Can remain alive for 5 years or more after a single blood meal
- Argasid ticks’ larvae, nymphs and adults attach on hosts only for a short period during feeding,
after which they drop off
- They may seek different host for subsequent blood-meal
- Are thus referred to as 'many-host' or 'multi-host' ticks
Medical importance
1. Tick-borne relapsing fever
- Caused by a spirochaete Borrelia duttoni
- Is the only important disease transmitted to man by soft ticks
- Transmission of spirochaete:
- Spirochaetes ingested with blood meal
- Multiply in the gut and congregate along the gut wall of the tick's mid gut
- Pass across into the haemocele where they can be found after 24 hours
- In the haemocele spirochaetes multiply enormously and invade all tissues and
organs of the tick's body
- Within 3 days they are found in the salivary glands, the coxal organs and ovaries
- In O. moubata complex the salivary glands in nymphs appear to be more heavily
infected than those in adults
- In contrast the coxal organs in nymphs are more lightly infected than in adults
that seem to be more heavily infected
- When either immature stages or adults of O. moubata complex feed on man, or
some other host, saliva is injected into the bite - spirochaetes being introduced
by this route
- During feeding excess body fluids are filtered from haemocele by the coxal
organs. Coxal fluid with spirochaetes in infected ticks (especially adults) is
passed out through the opening of the coxal organs onto the skin of host
- Spirochaetes will enter Host's body through the puncture of the tick's bite
or through the intact skin
- There is transovarial transmission of B. duttoni in the ticks
- Female ticks get infected with spirochaetes which are passed to
the eggs so that newly hatched larvae, nymphs of all instars and
resulting adults of both sexes are infected
- There is transstadial transmission where infection can be acquired by the larvae or
nymphs and passed to the subsequent stages in the life cycle of the tick

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nymphs and passed to the subsequent stages in the life cycle of the tick
IXODIDAE (Hard ticks)
- Hard ticks are distributed world-wide but occur more frequently in temperate regions than
the soft ticks (Argasidae)
- There are about 650 species of hard ticks belonging to 13 genera
- Medically important hard ticks belong to genera Ixodes, Dermacentor, Amblyomma,
Haemaphysalis and Hyalomma.
- Hard ticks are vectors of typhuses such as:
- Rocky mountain spotted fever (Rickettsia rickettsii)
- Boutonneuse fever (R. conori)
- They can also spread
- Q-fever (Coxiella burneti)
- Many arboviruses
- Hard ticks also transmit tularaemia (Pasteurella tularensis)
- Cause tick paralysis
External morphology
- Hard ticks are flattened dorso-ventrally
- Are oval in shape
- Measure 3-23mm in length depending on species and whether unfed, or fully engorged with
blood
- Females are nearly always bigger than males
- Capitulum ('false head') sticks out in front beyond the body outline and is visible from
above
- This distinguishes adult Ixodid from those of Argasid
- The palps are swollen and club-shaped
- The chelcerae have cheliceral sheaths covered with very small denticles
- Chelicerae are toothed at the ends
- Hypostome is also toothed
- Both hypostome and chelicerae penetrate host during feeding
- All hard ticks have a dorsal plate called scutum (shield)
- In males scutum is large and covers almost the entire dorsal surface of the body
- In females it is much smaller and restricted to the anterior part of the body, just behind
the capitulum
- In engorged females scutum is difficult to see
- In many species the posterior margin of the body has festoons - rectangular indentations
- These may be difficult to see in fully engorged females
- Hard tick have 4 pairs of legs terminating in a pair of claws
- Are generally dark, but some species have coloured markings on the scutum and body and some
times shiny patches of colour on the legs
- Such coloured species are referred to as ornate species
Life cycle
- The Ixodidae ticks, as the Argasidae, have hemimetabolous life cycle
- There is incomplete metamorphosis involving larval and nymphal stage
- Adult Ixodid ticks remain attached to their hosts for long periods of 1-4 weeks
- When feeding is stopped the enormously engorged tick drops on the ground
- Seeks shelter under leaves, stones, detritus or buries itself in the surface soil
- Oviposition begins 3-6days after the female drops from the host
Eggs:
- Some 1000-8000 eggs are laid in gelatinous mass formed in front and on top of the scutum
of the tick
- Some species may lay up to 20,000 eggs in an egg mass larger than the ovipositing female
- Oviposition may last 10 days or may extend for about 5 weeks
- Eggs are coated with a waxy secretion produced by Gene's organ
- Ixodid female lays only one batch of egg after which she dies
- Eggs hatch in 2-3 weeks or even several months into six-legged larvae
Larvae:-
- Are minute - about 0.5-1.5mm long
- Have toothed hypostome
- Remain inactive for a few days after which they swarm on the ground, climb vegetation and

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- Remain inactive for a few days after which they swarm on the ground, climb vegetation and
cluster at the tips of grasses and leaves 'questing' - waiting for host to come by
- When host comes by the larvae climb on the host and crawl to their favoured sites for
attachment
- Site is commonly in ears and eyelids depending on species
- Chelicerae and hypostome are inserted deep into the skin of host and the larvae commence
blood-feeding
- Remain attached to the host for 3-7 days and then drop to the ground
- Seek shelter among vegetation or under stones
- Take 2-7 days to digest blood meal
- After digestion they remain inactive for a few days before moulting to become nymphs
Nymphs:
- Are eight legged
- Crawl over the ground and climb vegetation and behave similarly to larvae (questing) in
seeking for a suitable passing host
- Attach to suitable parts of host and begin to feed
- 5-10 days later they are fully engorged and drop on the ground to seek shelter
- Remain quiescent for 3-4 weeks while blood meal is being digested
- Nymphs moult afterwards to produce adult ticks - males or females
- There is only one nymphal instar in the Ixodid tick's life cycle
Adult:
- Newly formed adult remains inactive for about 7 days
- Afterwards start questing for passing hosts
- Females take very large blood-meals - ingest 200 times its own weight
- Remain attached to host for 1-4 weeks
- On engorgement they drop to the ground and seek shelter
Behaviour and Habits
- Certain species of Ixodid ticks are more or less host specific
- But many species of medical importance are less specific and feed on a wide range of
mammals including man
- Adults may live for 7 years
Three-host ticks:
- A different host is parasitised by larva, nymph and adult
- Moulting occurs on the ground
- Most Ixodid ticks have this type of life cycle
- Ticks of medical importance in this group belong to the genera Ixodes, Dermacentor,
Haemaphysalis and Rhipicephalus.
- Ticks which feed on three hosts are more likely to become infected with pathogens and be
potential vectors of disease
Two-host tick:
- In some species in particular many from the genera Hyalomma and Rhipicephalus, after the
larva has completed feeding it remains on the host and moults to produce a nymph which
then feeds on the same host
- Engorged nymphs drop off and moult on the ground
- Resultant adults feed on a different host
- This is a two-host feeding cycle which involves larva and nymph feeding on one host and
the adult feeding on the second
One-host ticks:
- In a few species in the genus Boophilus, the larva, nymph and adult all feed on the same
host
- Moulting also takes place on the same host
- The only stage that leaves the host is the blood-engorged female which drops on the ground
to lay eggs
- The only way such ticks can transmit disease is through transovarial transmission
- One-host ticks are of little or no medical importance
Medical importance
1. Tick paralysis:
- Not caused by any pathogens
- Caused by various toxins contained in the female tick's saliva, which is continually pumped

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- Caused by various toxins contained in the female tick's saliva, which is continually pumped
into the host during the long period the tick takes feeding on the host
- Disease presents as an acute ascending paralysis affecting the legs with the result that the
person cannot walk or stand, has difficulty in speaking, swallowing and breathing
- Is due to paralysis of the motor nerves
- Symptoms appear 5-7 days after commencement of feeding by female tick
- May cause death of animals and also man as a result of respiratory failure
- Characteristic in hard ticks
2. Arboviruses:
- Transmitted by tick's bite and transovarial transmission commonly occurs
Russian Spring-summer encephalitis (RSSE)
- Viral disease associated with the Tiaga forest of Russia, Siberia, northern Asia and
China
- Main vector is Ixodes persulcatus, In other areas Haemaphysalis concinna may be
important
- After multiplication in the tick the virus accumulates in the salivary glands
- Infection is through the tick's bite
- Various small mammals and birds serve as reservoirs
- There is transovarial transmission in ticks
Tick-borne (Central European) encephalitis (TBE)
- Virus produces disease with similar symptoms to RSSE
- Occurs in central Europe from Scandinavia to the Balkans
- Principal vector are Ixodes ricinus and Dermacentor marginatus
- Various small mammals are reservoirs
- TBE virus accumulates in mammary glands of goats, sheep and cows
- People usually become infected not through tick bite but by drinking infected milk
or eating cheese
- Both transstadial and transovarial transmission occurs in ticks
Kyasanur Forest Disease (KFD)
- disease occurs in tropical forests of Southern India
- Is spread by Haemaphysalis spinigera
- Small rodents, bats and birds may be reservoirs
- Monkeys are amplifying hosts
- Transstadial transmission occurs in ticks
Crimean-Congo haemorrhagic fever (CCHF)
- Occurs in Bulgeria, areas of Russia especially Crimea, Pakistan and in areas of West
central and West Africa
- Transmitted by Hylomma marginatum, other species of Hyalomma and also
Amblyomma, Rhipicephalus and Boophilus species
- The ticks occur on a variety of animals including birds which fly from Russia to
Africa, Asia and Western Europe
- Transmission is by tick bite or crushing infected ticks, or accidental infection when
shearing tick infested sheep
- There is transovarial transmission in ticks
3. Rickettsiae
Rocky Mountain spotted fever (RMSF)
- Disease also known as Mexican spotted fever, Sao Paulo spotted fever, American
tick-borne typhus
- Causative organism is Rickettsia rickettsii
- Dermacentor andersoni, and D. variabilis are the vectors in North America
- Dogs, rabbits and small rodents also become infected
- Animals remain infectious only for a short time
- Ticks thus the main reservoir
- Incubation period in ticks is 9-12 days -infected tick becomes infective
- Transmission is normally through bite of any stage in the life cycle of the tick
- Infective tick must remain feeding on host for over 2 hours to inject sufficient
Reckettsia to infect host
- There is transstadial and transovarial transmission in ticks
Siberian tick typhus (STT)

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Siberian tick typhus (STT)
- Disease is similar to Rocky Mountain spotted fever
- Caused by Reckettsia sibirica
- Occurs in Russia, Pacific areas and the Japanese Islands
- Vectors include species of Dermacentor, Haemaphysalis, Rhipicephalus and
Hyalomma.
- Infection by tick bite
- There is transstadial and transovarial transmission in ticks
- Mammals mainly rodents may serve as reservoirs
Boutonneuse fever
- Also known as Marseilles fever, South American tick typhus, Kenyan tick typhus,
Indian tick typhus, etc.
- Caused by Reckettsia conori
- Symptoms in man similar to Rocky Mountain spotted fever
- Occurs in the Mediterranean region, Middle East, Crimean, India, Southern Asia and
Africa
- Principal vector is Rhipicephalus sanguineus, the dog tick
- Most Ixodid ticks can transmit the pathogen
- Transmission is by tick bite
- There is transstadial and transovarial transmission in ticks
- Both ticks and rodents serve as reservoirs
4. Spirochaetes
Lyme disease:
- Caused by a spirochaete transmitted by Ixodes dammini in Eastern US mainly in
deer
- In western US transmission is by I. Pacificus
- Symptoms include, arthritic pains in the joints
- Little is known of its epidemiology
- Disease is non-fatal
- There is transstadial and transovarial transmission in ticks
5. Tularaemia
- A bacterial disease caused by pasteurella tularensis
- Occurs in N. America, Europe, Japan and Asia
- Infects mainly rabbits, other rodents and even birds
- Spread by a variety of direct contact methods as handling infected animals, carcasses,
drinking contaminated water eating raw meat
- Also transmission is by tick bite from various hard ticks
MITES
Class: ARACHNIDA - Tick, mites and spiders
Order: ACARINA - Order contains ticks and mites
There are 2 groups of medical importance
1. The Sarcoptes mites - (scabies mites)
2. Leptotrombidium mites - (Scrub typhus mites)
SCABIES MITES
- Belong to the species Sarcoptes scabiei
- Also referred to as itch mite
- Has World-wide distribution
- Occurs on man -commonly on skin especially on arms
- Common community problem in crowded areas
External morphology
- Female mite is just visible without the aid of a hand lens
- Measures 0.30 - 0.45mm
- Whitish in colour
- Disc-shaped
- Dorsally the mite is covered with numerous small peg-like protuberances and a few bristles
- Both dorsally and ventrally there are a series of lines across the body giving the mite a
striated appearance
- Adults have 4 pairs of short cylindrical legs, divided in 5 apparent ring-like segments
- 1st 2 pairs of legs end up in short stalks called pedicels which terminate in thin-walled

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- 1st 2 pairs of legs end up in short stalks called pedicels which terminate in thin-walled
roundish structures often termed 'suckers'
- In females the posterior pair of legs do not have 'suckers' but end up in long very
conspicuous bristles
- Adult males have the 'suckers' on the last pair of legs
- No distinct head
- Have short and fat palps and pincer-like chelicerae (mouth parts) protruding anterior from
the body
Life cycle:
- Females burrow into the superficial layers of the skin and excavate winding tunnels at the
rate of 1-5mm per day
- The mite feed of the liquid oozing from the dermal cells they have chewed
- Lay 1-3 egg a day in the tunnels
Eggs
- Egg hatch in 3-5days into small six-legged larvae
- Larvae look like miniature adults
- Larvae crawl out of the tunnels onto the surface of the skin
- Large number of the larvae die at the surface
- But a few manage to burrow into the skin or in hair follicle to form a 'moulting pocket'
- In 2-3 days the larvae moult in the pocket into eight legged nymphs
Nymphs
- Nymphs moult in the pocket into adults - male or female
- Females remain inactive in the moulting pocket until fertilized by a male
- After fertilization the mite enlarges in size to become a mature (ovigerous) female
- After fertilization the female commences to burrow through the skin
- After 3-5 days the new female starts to lay eggs in the tunnel
- Female mites rarely leave the tunnel
- Adult males spend most of their life wondering on the surface of the skin seeking females
awaiting to be fertilized
- Life cycle from egg to adult takes about 11-28 days
- Females may live for1-2 months on man
- Away from man they live for 7-10 days in ideal conditions, but usually live for 2-4 days
Scabies
- Is a contagious complaint transmitted only by close contact
- Diagnosed by detection of the female mite's thin twisting tunnels
- Tunnels easy to see on a light skinned person
- Faeces deposited in the tunnels may be visible through the skin as pepper-like
sports
- Mite can also be dissected from the tunnel and examined under x50 magnification
- An average of 11 female mites are found on a person
- Most patients have 1-15 female mite
- Only 3% may have more than 50 mites
The Scabies rash
- Is a follicula papular eruption that occurs on areas of the body not infected with burrowing
mites
- Rash produced in response to the patient being sensitized
- It is an allergic reaction produced by mites
Treatment of scabies
- BB (Benzyl benzoate) emulsion 20-25% painted on patient’s whole body from neck
downwards
- 0.5% HCH (lindane) may also be used - one treatment usually enough
Prophylaxis
- Availability of plenty of water for washing
SCRUB TYPHUS MITES
- Are grouped within the Trombiculid mites
- There are more than 1200 species of trombiculid mites belonging to several genera
- Only a few species attack man
- Medically important species include:
- Leptotrombidium deliense

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- Leptotrombidium deliense
- L. akamushi
- L. fletcheri
- These medically important species are found only in Asia
External morphology:
- Adults are small - 1.0-2.0mm
- Are reddish
- Covered dorsally and ventrally with numerous feathered hairs, giving them a velvety
appearance
- Have four pairs of legs ending in paired claws
- Palps and mouth parts project in front of the body
- Nymphs resemble adults, but are smaller (0.5-1.0mm), and are less densely covered by
leathered hairs
- Adults and nymphs do not bite man or animals
- Feed on small arthropods and their eggs
- Only larvae are parasitic on man and other animals
- The larvae responsible for the spread of diseases
The larvae;
- Are very small (0.15-0.3mm)
- But size may increase six fold when engorged
- Are usually reddish or orange in colour, but may be pale yellow to straw-coloured
- Have three pairs of legs terminating in a pair of claws
- Both legs and body covered with fine featured hairs
- Five-segmented palps and mouth parts are large and conspicuous- having appearance of a
false head
- Dorsally on the anterior part of the body there is a rectangular or pentagonal-shaped scutum
- Scutum bears 3-6 setae
- On either side of the scutum there is a pair of eyes
- In medically important species there are 5 feathered setae on the scutum
- And in addition a pair of specialized feathered hairs known as the sensillae which arise
from distinct bases
Life cycle
- Female mite lays 1-5 eggs each day, on the surface of damp soil or under leaves
- In hot climates overposition continues uninterrupted for a year
- In cold periods egg laying stops and adult hibernates
- After 5-7 days egg shell splits
- Larva does not emerge but remains within the egg shell and is called deutovum
The larvae
- After 5-7 days the larva crawls out of the egg shell, becomes very active, swam over the
ground and climb on grasses and low lying vegetation and wait for host
- The larvae will attach to birds, mammals, including man walking through infested
vegetation
- On the host, the mite larvae congregate in areas where the skin is soft and moist - such as
the ears, genitalia and around the anus
- On man the larvae seek areas where cloth are tight against the skin -as around the waist and
ankles
- The larvae pierce the host's skin with their powerful mouth parts and inject saliva into the
wound which cause disintegration of the cells
- Larva normally does not suck blood, but the lymph and other fluid and semi-digested
materials
- The larvae remain attached to the host for 2-10 days
- Engorged larvae drop to the ground and bury themselves just below the surface of the soil
or underneath debris
- Concealed larva becomes quiescent and this stage is known as the protonymph
- After 7-10 days the protonymph moults to produce eight-legged reddish nymphs covered by
feathered hairs
- The nymphs feed on soil-inhabiting arthropods
- After some days to 2 weeks the nymphs cease to feed and become inactive and are called
pre-adults

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pre-adults
- After 14 days pre-adults moult to give forth to adult mites
- The life cycle usually takes 2-3months but may be longer (8-10months)
Medical importance
1. Nuisance
- Although some species of trombiculidae mite do not transmit disease to man their bite can
cause intense itching and irritation commonly referred to as:
- Harvest-bug itch
- Autumnal itch
- Scrub itch
- Also if the mites are forcefully removed from skin, mouth parts remain and can promote
further irritation
2. Scrub typhus
- Caused by Rickettsia tsutsugamushi
- Disease also known as mite-borne typhus, Rural typhus, Japanese river fever or
tsutsugamushi disease
- Disease restricted to Asia where it occurs over a large area
- Disease was serious in troops in Asiatic-Pacific areas only second to malaria
- Disease has often been regarded as a zoonosis but experiments indicate that man is the most
suitable host
- And the Leptotrombidium mites are the main reservoir
- Man becomes infected following the bite of infected larvae of Leptotrombidium akamushi
and L. deliense.
- People get infected when they visit or work in areas infested by the mites - referred to as
'mite islands' - patches of vegetation harboring large numbers of host seeking larvae of
the mites
- Transovarial transmission occurs in the mites
INJURIOUS NON-PARASITIC ARTHROPODS
- Insect stings
- Hymenoptera stings (Bees, wasps, and hornets)
- Scorpion stings
- Spider bites
- Centipede bites
Insect stings
- Result from Hymenoptera stings
- These include:-
• Bees
• Wasps
• Yellow jackets
• Hornets
- Hymenoptera venoms may have direct toxic effects
- But this is not seen in man unless in situations of many stings – usually hundreds
- In Temperate countries, anaphylactic reactions to Hymenoptera stings are a
commoner cause of death than direct effect of envenoming by any animal
- E.g. in 1959 – 1972, 61 deaths occurred from insect stings in England and
Wales, compared to only one resulting from adder bite
- There are 40 – 50 deaths annually from Hymenoptera stings
- In fact deaths due to Hymenoptera sting anaphylaxis are under reported – not
commonly suspected as one of the causes of death in people found dead with
indications of myocardial infarction or cerebrovascular accidents
- Commonest and most severe Hymenoptera stings are caused by members of the
families:-
- Family Apidae within which there is species Apis mellifera – the honey
bee
- Family Vespidae, including
- wasps such as Vespa vulgaris
- American yellow jackets – genus Dolichovespula
- Hornets – genus Vespa

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- Hornets – genus Vespa
- The venoms are normally injected through a barbed sting
- Bees inject approximately 50μg of venom – total capacity of the venom sc and
leave the sting embedded in the skin
- Wasps and hornets can sting repeatedly
- The venom contains
- biogenic amines:-
• Histamines
• 5-hydroxytryptamine
• Acetylcholine
- Enzymes
• Phospholipase A
• Hyaluronidase
- Toxic peptides
• Kinins – in Vespidae
• Apamin
• Malittin
• Anti-inflammatory compounds as mast cell degranulating peptides –in
Apidae
Clinical features:-
In non-allergic people
- Venom produces local effects due to injection of biogenic amine mainly 5-
hydroxytryptamine
- This is characterized by:-
• Pain
• Redness
• Swelling
• Whealing
• Heat developed very rapidly but for just a few hours
- The local effects can be dangerous if airway is obstructed e.g. when the stings
are on the tongue
- Fatal systemic stings can result from as few as 30 stings in children
- Adults have survived as more than 2000 stings of Apis mellifera
- Clinical effects of massive envenoming resemble histamine overdose and
include:-
• Vasodilation
• Hypotension
• Vomiting
• Diarrhoea
• Throbbing headache
• Coma
Allergic effects
- Normally seen in 0.5% of the population who have become hyper sensitized
with Hymenoptera venom
- Reactions to successive stings are increasingly severe
- Systemic symptoms include:-
• Tingling scalp
• Flushing
• Dizziness
• Wheezing
• Abdominal colic
• Diarrhoea
• Tachycardia and visual disturbance developing within a few minutes of
the sting
• In the next 15 – 20 minutes the following symptoms may occur:-
o Urticaria

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o Urticaria
o Angioneurotic oedema
o Oedema of the glottis
o Profound hypotension
o Coma
• Patient may die within minutes
Treatment:-
- Remove embedded bee stings without squeezing
- Administer analgesic mainly aspirin
- Local antiseptics may be applied
- * topical anti-histamines should not be used as they promote sensitization
- In situations of severe systemic envenoming, life threatening effects of biogenic
amines can be treated with adrenaline
- Adrenaline is also treatment of choice for sting anaphylaxis – given subcutaneously
SCORPION STINGS
- Scorpions fall in the Order Scorpionida
- Those capable of inflicting fatal stings to humans belong to the Family Buthidae
- The most deadly species include:-
• Androctonus australis – N. Africa and Middle East
• A. crassicauda – in N. Africa, Mid. East and Turkey
• Buthus accitanus – Mediterranean region and Mid. East
• Leiurus quinquestriatus –
• Parabuthus – South Africa
• Tityus trinitatis – Trinidad and Venezuela
• Bathotus tamulus – India
• Centruroides sculturatus – California, New Mexico, Arizona
- Painful scorpion stings are common throughout the tropics
- Fatal envenoming stings are common only in Mexico, Brazil, Trinidad, parts of
North Africa, Middle East and India
• In Libya there were 900 scorpion stings with 7 deaths per 100,000
population in 1979
• In Mexico there are 1 – 2000 deaths per year from scorpion stings –
incidence of 84 deaths per 100,000 annually
• In Algeria there some 1260 stings with 24 deaths per year
Clinical features:-
- Very intense local pain
- Local swelling
- Redness
- Heat
- Systemic symptoms may develop within minutes of the sting, but may be
delayed for 24 hours and will include:-
• Autonomic nervous system excitation including
o Dilated pupils
o Hyper salivation
o Profuse sweating
o Hyperthermia
o Abdominal distension
o Lose of sphincter control
o Hypertension
o Toxic myocarditis
o Cardiac failure
o Pulmonary oedema
• Neurotoxin effects as:
o Fasciculation
o Spasms
o Respiratory paralysis
Treatment

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Treatment
- Parenteral opiate analgesics such as pethidine and morphine may be required
- But are alleged to be dangerous in victims of Centruroides sculpturutus
- Systemic envenoming should be treated with specific anti-venom given
intravenous injection as in snake bite treatment – Anti-venoms are
manufactured in many countries as USA, Britain, Germanys, Mexico, Turkey,
Algeria, S. Africa, Egypt and Iran
- But no antivenom is available for treatment of Buthotus tamulus stings in
India
SPIDER BITES
- Spiders belong to the Family Araneae
- Family contains a large group of spiders in over 30,000 species
- Only 1% of these is non-venomous
- Only 12% of species of spiders are known to cause dangerous envenoming in
humans
- The spiders bite with a pair of fangs – the chelicerae, to which the venom glands
are connected
- A central venom duct opens near the tip of the fangs
Clinical features:-
- Two main clinical features syndromes are caused by spider bite
• Necrotic
• Neurotoxic
Necrotic araneism
- Characterized by skin lesions
- Vary in severity from mild localized erythema and blistering to quite extensive
tissue necrosis
- Range of severity attributed to variety of species of spiders
- Members of the genus Loxosceles are the most important cause of the syndrome
- L. laeta is widely distributed in Central and South America especially in Chile
- L. reclusa – the brown recluse spider is distributed in USA
• Has caused at least 200 bites with 6 deaths
- L. rusfescens – occurs in the Mediterranean region N. Africa and Israel
- 80% of patients are bitten indoors usually in their bedrooms while asleep or
dressing
- In the US, a number of men were bitten on their genitals when they sat on the
outdoor lavatories on which the spider had spun their webs
Clinical presentation:-
- Spider bite is characterized by a burning pain at the site of bite
- Oedema and development of a violaceous plaque follows
- In a few days the plaque becomes a black eschar which sloughs in a few weeks
- In 12% of cases there are systemic effects involving:-
• Heamoglobinuria
• Jaundice
• Fever
• Respiratory distress
• Collapse
- Average mortality is 6% in all reported cases, and 30% in those with systemic
envenoming
Neurotoxic araneism
- Neurotoxic venoming results from bites by spiders in the Latrodectus,
containing:-
• Widow spiders
• Hourglass spiders
• Red-black spiders
- These are the most widespread and numerous of all venomous animals
dangerous to man
- L. mactans – the black widow spider,
• Occurs in Americus

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• Occurs in Americus
• 63 deaths were attributed to this species in the US in 1950 – 1959
- L. mactans tredecimguttatus – incorrectly referred to ‘tarantula’ is widely spread
in the Mediterranean countries
• Live in fields
• Has been responsible for epidemics of spider bites
• 946 cases were reported in Italy in 1946 – 1951
- L. mectans hasselti – Australian and New Zealand ‘red-black spider or ‘kitopo’
• Cause upto 340 reported bites each year in Australia
• 20 deaths have been attributed to such bites
Clinical presentation of the spider bites
- L. m. hasselti bites
- Local heat
- Sweating and redness which is rarely extensive
- Intensive local pain which develops in about 5 minutes
- Pain in the local lymph nodes after 30 minutes
- Headache, nausea, vomiting and sweating occurring after one hour
- Tachycardia and hypertension may follow
- Muscular tremors and spasms occur which may be severe enough to require
artificial ventilation
- L. m. mactans bites
- Local dull aching or numbness may develop in 30 – 40 minutes
- Painful muscular spasms and lympadenopathy spread and increase in
intensity during the next few hours
- The trunk of abdomen and limbs get involved
- Respiration may be inhibited
Treatment
- First aid may be needed for spider bites with rapidly active potent venom such as A.
robustus.
- Firm crepe bandaging or splinting of bitten limbs, or tight tourniquet may delay
venom spread until patient reaches hospital
- Specific treatment
- Treatment mainly based on use of anti-venoms
- Specific antivenoms are manufactured in a number of countries including
Australia, USA, USSR, Italy, Yugoslavia and South Africa

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Leishmaniasis
Thursday, 22 December 2016 11:22 AM

• Caused by the parasite, leishmania

• There are three major types
○ Visceral
○ Cutaneous
○ Mucocutaneous
• The different species responsible for the above are
○ L.donovani
○ L.tropica
○ L.brazilienses
• The parasites are transmitted by sand-fly, known as phlebotomus. Different species exist.

Geographical Distribution
• Endemic in many places such as India, China, Africa, S. American and S. Europe and
Russia
• Disease was first described in 1903
• Several animal reservoirs exist, thus maintaining the endemicity of the parasite, these
include, squirrels, dogs and hamsters

Habitat
• Inside the vertebrata host such as man, the parasite is always intracellular, occurring in
amastigote form, without flagella.
• Essentially invades the reticulo-endothelial system
• In the vector, sand-fly, it occurs as promastigote, with flagella, within the digestive system
of the vector

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Pathology and Pathogenesis
• The parasite invades the reticuloendothelial system (RES) via macrophages
• The parasite causes hyperplasia of RES in bone marrow, spleen, liver and skin being
mostly affected
• The hyperplasia of RES causes dysfunction of the organs affected
• Hyperplasia of RES causes organ enlargement as does in visceral leishmaniasis
• The infection triggers cytokine production which are mediators of inflammation and
immunosuppressants such as TNF
• There is pancytopenia due to a dysfunctional bone marrow, leading to neutropenia and
anaemia
• Secondary bacterial infections complicate the condition of the patient and may lead to
death
• Epistaxis may occur due to thrombocytopenia

Diagnosis of Cutaneous and Mucocutaneous Leishmaniasis

• Skin biopsy for histology
• Biopsy of soft tissues for histology
• Serology ELISA
• PCR
• Clinical picture

Treatment
• Pentavalent antimonials e.g. Sodium Stibogluconate
• Amphotericin B
• Pentamidine
• Miltefosine
• Aminosidine
• Gamma interferon
• Imidazoles (ketoconazole, itraconazole)
• Allopurinol

Control of Leishmaniasis
• Treatment of patients and carriers
• Elimination of reservoir hosts e.g. rodents and stray dogs
• Vector control e.g. by use of insecticides
• Destruction of vector breeding sites such as anthills
• Vaccine is under development

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 Introduction to Virology
Wednesday, 9 December 2015 8:23 PM

What are viruses?

· A virus is an obligate (strictly dependent on other organisms for their own survival) parasite
containing genetic material surrounded by protein
· they are not living organisms
· They cannot produce energy or synthesise proteins independently, the host cell machinery is
needed

Components of a virus
· Contains
1. Viral genome or nucleic acids: for replication of the mother virus.
§ Covered by a protein layer ie capsid for protection of the genomic material.
§ Some viruses only have the genomic material and capsid (naked viruses)
2. Carry certain enzymes
3. Covered by an envelope (envelope virus)
4. Contains a ligand for attachment to the host cell
§ Immune responses of the body respond to this ligand

Properties of Viruses
1. Envelope viruses
2. Naked viruses

Differences between enveloped and naked viruses

Properties Enveloped Naked
Environmental Tactile, destroyed by acids, detergents, drying Stable to temperature acids, detergents,
stability proteases, disinfectants
Release from cells Budding and cell lysis Cell lysis
Spread to hosts Spreads in larger droplets, secretions, Spread easily (by direct contact and dust
transplanted organs, blood transfusion and small air droplets)
Stability outside Must stay wet Can dry out but retain infectivity
cells
Effective Immune May need antibody and cell mediated immunity Antibody may be sufficient for
response for protection immunoprotection

Differences between bacteria and viruses

Viruses Bacteria
Obligate intracellular parasite Usually free-living, but can be parasites
No ribosomes Ribosomes
DNA or RNA, not both Both DNA and RNA
Seen by electron microscope Seen by light microscope
10 - 100s of genes 100 - 1000s of genes
Tangled phylogeny Natural phylogeny

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 Classification of animal viruses
1. Traditional taxonomic approach
a. Classification based on taxonomy system of order, family, subfamily, genus
and species
i. Order: virales = herpesvirales
ii. Family: viridae = herpesviridae
iii. Genera: virus = alpha herpesviruses
2. Classification based on disease or target organ
a. Respiratory viruses
b. Central nervous systems viruses
c. Gastrointestinal viruses
d. Hepatitis viruses
3. Based on host range (avian flu)
a. Influenza virus
i. Avian influenza
ii. Human influenza
iii. Swine influenza

Baltimore classification of Viruses

· Classification based on the relationship of viral genome to its mRNA
· Virus genomes [dsRNA (double stranded), ssRNA (single stranded), dsDNA, ssDNA (+/-)]

Classes
I. dsDNA: produce mRNA directly from the strand
· Herpes
· Pox virus
· Adenovirus
II. ssDNA - dsDNA: make complementary strands to form mRNA
· Adeno-associated virus
§ Parvovirus
III. dsRNA: can make mRNA directly
· Reovirus
IV. (+)ssRNA - (-)ssRNA:
· Togavirus
· Poliovirus
· Foot and mouth disease virus
· Hepatitis A virus
· Hepatitis C virus
V. (-)ssRNA: make mRNA directly
· Influenza
· Paramyxovirus
· Arenavirus
VI. ssRNA-RT - DNA/RNA - dsDNA: carry reverse transcriptase that converts RNA to DNA
and mRNA formed there
· HIV
VII. dsDNA-RT
· Hepatitis B virus

mRNA and Virus Replication

· virus with ssRNA with the same orientation as the mRNA - positive strand RNA
· Viruses with ssRNA genome complementary to its mRNA - negative strand RNA virus

Replication of Viruses

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Replication of Viruses
1. Attachment: viral protein to cell surface receptors
2. Penetration: to the host cell membrane
3. Uncoating: for envelope viruses to inject viral material into the cell cytoplasm. Involves
the removal of the lipid membrane and protein coat
4. Transcription: production of mRNA
5. Translations: Synthesis of viral proteins
a. RNA viruses: virus specified polymerases
b. DNA viruses: host cell polymerases
6. Assembly: nucleus, cytoplasm or plasma membrane
7. Release: cell lysis or budding through plasma membrane

Transmission of viruses
1. Airborne transmission
§ Examples - viruses shed from the upper respiratory tract (common cold, influenza), viruses
shed from skin lesions (chicken pox, herpes viruses)
2. Faecal-oral
§ Viruses shed in the faeces transmitted by contaminated water or food (hepatitis A,
rotovirus, enterovirus)
3. Body fluids
§ Through blood or other body fluids (hepatitis B and C)
4. Vector transmission
§ Viruses carried by insects and arthropods. Arbo viruses

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Herpesviruses
Wednesday, 9 December 2015 8:22 PM

Herpesviruses
· More than 130 herpes viruses
· One 8 are associated with human disease:
a. Herpes simplex I & II (cold sores, genital herpes)
b. Varicella zoster (chicken pox, shingles)
c. Cytomegalovirus (microcephaly, infectious mono)
d. Epstein-Barr virus (mononucleosis, Burkitt's lymphoma)
e. Human herpes virus 6&7 (Roseola)
f. Human herpes virus 8 (Kaposi's sarcoma)

Properties
1. dsDNA (Baltimore virus classification = class 1)
2. Icosahedral capsid
3. Enveloped (lipoprotein) - only viruses to obtain envelope by budding from the nuclear
membrane
4. Tegument between the nucleocapsid and envelope - contains regulatory proteins for viral
replication
5. Nuclear replication
6. Genome is linear double stranded
7. Does not contain a polymerase

Most common and most important herpes viruses of humans include:

1. Herpes Simplex Virus (HSV)
2. Varicella Zoster Virus (VSV)
3. Epstein-Barr (EB) virus
4. cytomegalovirus (CMV)

Classification
1. Family: Herpesviridae
2. Subfamilies:
a. Alpha herpesviridae (HSV-1. HSV-2, VSV)
i. Infect epithelial cells primarily and cause latent infection in neurons
b. Beta herpesviridae (CMV, HHV-6, HHV-7)
i. Infect and become latent in a variety of tissues
c. Gamma herpesviridae (EBV, HHV-8 or KSHV) - Cancer causing
i. Infect and become primarily latent in lymphoid cells
3. Genus
a. Simplex virus (HSV 1/2)
b. Varicelovirus (VSV or HHV3)
c. Lymphocryptovirus (EBV)
d. Cytomegalovirus (CMV)
e. Roseovirus (HHV6, HHV7)
f. Rhadinovirus (HHV8=KSHV)

Herpes Simplex Viruses

· Two types (HSV-1 AND HSV-2)
· Infection is most often inapparent
· Usual clinical manifestations:

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· Usual clinical manifestations:
1. Vesicular eruption of the skin or mucous membranes
2. Sometimes seen as severe keratitis, meningoencephalitis and a disseminated illness of
the newborn

Human Herpes Virus - 1 (HSV-1)

· Primarily transmitted in saliva.
· As a result, infection mainly occurs on the face

Transmission of HSV-1 and HSV-2

1. Intimate contact (skin to skin contact)
2. The virus does not penetrate intact skin
3. Mild abrasion or chapping of skin can allow infection

Tissue tropism os HSV-1 and HSV-2

HSV-1
· Causes 95% orofacial herpes (remainder caused by HSV-2)
· Causes 10-30% of primary genital herpes (but seldom recurs there)
HSV-2
· Causes primary and recurrent genital herpes infections
· may cause primary oral herpes but, like HSV-1 in genital area, it seldom occurs there

Pathogenesis of HSV-1
Asymptomatic infections - young children (usually <5 year old)
Primary infection
1. Oropharyngeal mucosal cells
2. Viremic spread (infection of different organs in the body)
Conditions include:
1. Acute gingivostomatitis
2. Recurrent herpes labialis (cold sores)
3. Keratoconjuctivitis
4. Encephalitis/meningitis
5. Pharyngitis
6. Mucocutaneous lesions/ herpertic whitlow etc

Herpes Labialis (cold sores, herpes febrilis)

1. Most common recurrent disease of HSV-1
2. Clusters of localised vesicles
3. Vesicles rupture (painful ulcer - heals without scarring)
4. Lesions may occur repeatedly and at different intervals of time at the same location
5. Permanent site of latent HSV is the trigeminal ganglion

Viral Latency
1. Primary infection
2. Virus migrate to the PNS in the trigeminal ganglion
3. Virus dormant (no replication) - latency
a. No viremia
b. Antiviral Abs in blood
4. Reactivation of HSV
a. Stress to body (AIDS, immunosuppressive drugs, menses etc)
b. Disease and virus shedding (symptomatic or asymptomatic)

Three manifestations of HSV Latency

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 Three manifestations of HSV Latency
Key feature: There is a wide spectrum of clinical presentations
1. Some individuals (5-10%) have frequent clinical reactivation
2. Most individuals reactivation is clinically asymptomatic
3. In all cases, virus is shed

Human Herpes Virus - 2 (HSV-2)

· Primarily transmitted through sexual contact
· As a result, infection is mainly in the genital region

Pathogenesis of HSV2
· Sexually transmitted virus (STI)
· Viremic spread to multiple body organs
· Conditions
o Genital herpes
o Neonatal herpes
o Meningitis
o Keratoconjuctivitis
· HSV disease ranges from mild illness (asymptomatic) to primary infection to sporadic, severe
and life-threatening disease in infants, children and adults

Risk factors for HSV 2 infection

· Older age
· Females
· Low level of education
· Poor socioeconomic status
· Prior STD
· Promiscuity

Genital herpes (herpes progenitalis)

· Mainly caused by HSV-2
· Vesicular lesions on genitalia
· Lesions are more severe in primary infection (may be associated with fever, malaise and
lymphadenomyopathy)
· HSV-2 remains latent in lumbar and sacral ganglia

Neonatal Herpes
· HSV 2 transmitted to child during birth due to contact with lesions in vagina.
· The spectrum of illness produced in the new born appears to vary from subclinical or local to
severe generalised disease with a fatal outcome
· May cause permanent head damage to infants who survive

Specimen types for HSV

· Collected in transport media and taken to diagnostic lab
o From skin vesicles/lesion scrapings
o CSF
o Ocular fluid
o Swabs from mucocutaneous lesions
o Blood, sweat, urine, throat, nasopharynx, conjuctivae and cornea

Labdiagnosis of HSV

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 Labdiagnosis of HSV
· Virus isolation (Main test)
· Polymerase chain reaction - PCR (meningitis and encephalitis from CSF)
o Rapid
o High sensitivity and specificity
· Tzanck smear method
o Cheap, rapid
o Cannot distinguish between 1 and 2
· Direct fluorescent antibody assay (DFA) technique
o Can differentiate btwn the two
o Rapid and inexpensive

Comparison of diseases caused by HSV-1 and HSV-2

Site HSV-1 HSV-2
Skin Vesicular lesions above the Vesicular lesions below the waist
waist (especially genitals)
Mouth Gingivostomatitis Rare
Eye Keratoconjutivitis Rare
CNS Encephalitis (temporal lobe) Meningitis
Neonate Rare (infection acquired Skin lesions, encephalitis, and
after birth from infected disseminated infection (infection acquired
person) through passage through vagina)
Dissemination to Yes Rare
viscera in
immunocompromised
patients

Varicella-Zoster Virus (VZV)

· Varicella- chicken pox
o Acute disease that follows primary contact with the virus
· Zoster- shingles
o Response of the partially immune host to a varicella virus present in latent form in sensory
ganglia

Two unique features of VZV

1. Air borne spread or skin to skin contact
2. More severe infection if primary infection occurs as an adult

VZV is structurally and morphologically similar to other herpesviruses but is antigenically different. It
has a single serotype.

Pathogenesis of Varicella
· Highly contagious disease
· Transmission via aerosol (respiratory droplets/contact with infectious vesicular fluid)
· Primary infection:
1. High tropism for t lymphocytes
2. Viremic spread - dissemination to skin
3. Skin rash with high fever and non-specific symptoms
· Most VZV infections in childhood (adult possible)
4. In the adult: varicella pneumonia with fever, cough, tachypnea, dyspnea etc. Pneumonia is
transient but interstitial pneumonitis leads to respiratory failure
· I'm Viral latency (like VZV)

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 · I'm Viral latency (like VZV)

Pathogenesis of Zoster
· In addition to skin lesions - histopathogenically identical to those of varicella - there is an
inflammatory reaction of the dorsal nerve roots and sensory ganglia
· Often, only a single ganglion may be involved. As a rule, the distribution of lesions in the skin
corresponds closely to the areas of innervation from an individual dorsal root ganglion
· There is cellular infiltration, necrosis of nerve cells and inflammation of the ganglion sheath

Summary
Same virus causes two diseases
a) Varicella (chicken-pox)
a. Highly infectious disease
b. Chiefly in children
c. Characterised clinically by a vesicular eruption of the skin and mucous membranes
d. Causative agent is indistinguishable from the virus of zoster
b) Zoster (shingles)
a. Is a sporadic disease in adults (rarely in children)
b. Characterised by an inflammatory reaction of the posterior nerve roots and ganglia,
accompanied by crops of vesicles (like those of varicella) over the skin supplied by the
affected sensory nerves

Cytomegalovirus (CMV)

Transmission of CMV
1. In utero
2. Early childhood (saliva)
3. Venereal in young adults
4. Blood transfusion
5. Organ transplantation

Clinical Mannifestations
Normal individuals - asymptomatic in majority of cases

Three distinct clinical syndromes

1. Congenital cytomegalovirus infection
a. Primary CMV infection in 3rd trimester of pregnancy of a seronegative mother
b. Hepatospleno-megaly, retinitis, skin rash

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 Retroviruses
Wednesday, 9 December 2015 8:24 PM

Retroviridae family
Unique features
1. Three common genes: gag, pol, env
2. 2 copies of ss(+) sense RNA
3. The only viruses which are truly diploid
4. Only (+) sense RNA viruses whose genome does not serve directly as mRNA
immediately after infection

Lentiviruses
· Slow progressive infections
Ø Family: Retroviridae
Ø Genus: Lentivirus
Ø Species: HIV-1, HIV-2, SIV, BIV, FIV, EIAV

HIV Classification
Baltimore: Group 6 (ssRNA, Reverse transcriptase)
ICTV:
· Family: Retroviridae family
· Subfamily: Orthoretrovirinae
· Genus: Lentivirus
· Species: HIV-1, HIV-2, SIV, BIV, FIV, EIAV
· Groups: HIV - 1 - M, N, O and P
· Clades: HIV-1 (A-K, CRFs [circulating recognant form]); HIV-2 (A-H)

HIV Ligand-Cell Receptor interaction

Ligand: HIV glycoprotein
Cellular receptor: CD4
Co-receptor: CXCR4 and CCR5

Retrovirus Replication Cycle

1. Binding, Fusing and Entry
2. Reverse transcription
3. Integration
4. Replication
5. Transcription
6. Translation
7. Maturation
8. Budding

HIV-1 vs HIV-2
HIV-1 HIV-2

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 HIV-1 HIV-2
Geographical Worldwide West Africa, some cases in
distribution Europe and USA
Subtypes Over 10 subtypes A-K, At least 8 subtypes A-H
CRFs
Natural progression Easier and faster Less easily transmitted, slower
transmission progression
Origin Related to SIV Related to SIV (sooty mangabeys
(Chimpamzee SIV) SIV)
Treatment Responds to NNRTis No response to NNRTis

Transmission of HIV
1. Sexual contact: oral, anal, vaginal
2. Sharps: needles, blades
3. Blood transfusion
4. MTCT: during pregnancies, deliveries or breastfeeding

Risk factors: Acquisition of HIV infection

1. Nature of exposure (the size/dose of HIV inoculum)
2. The 'virulence' of HIV (virus genetics)
3. Host susceptibility to infection (host genetics eg delta 32 mutation)
4. Routes of transmission: transmucosal infection risks vary according to the site of
exposure (rectal exposure > vaginal)
5. Mucosal inflammation (ulceration by STDs)

Sequence of Events
1. HIV infects CD4 cells (APC)
2. Disseminated infection
3. Specific immune response (Ab, CMI)
4. Clearance of most virus
5. Some persistence
a. Gradual loss of CD4 cells
b. Destruction of lymphoid tissues

Clinical Features
1. Primary stage
a. Seen in 10% of individuals a few weeks after infection
b. Acute seroconversion
c. Presents with a flu like illness
2. Asymptomatic Stage
3. Symptomatic
4. Full-blown AIDS

HIV Testing

Purpose of testing
1. Diagnosis
2. Surveillance

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 2. Surveillance
3. Medical Legal

HIV Tests
1. Antibody tests
2. Antigen tests
3. Nucleic act id tests
4. Others: EM, Culture

HIV-1 Diagnosis
1. Serological tests:
a. Rapid HIV testing: For initial screening
i. ELISA: mi test
b. Western Blot test: Confirmation tests
c. p24 test: detection of a recent infection
2. Nucleic acid based
a. HIV viral load: for monitoring treatment
b. HIV-PCR: Virus detection in blood (detection of a recent infection)
especially infants
3. Immune marker:
a. CD4 Test: staging the disease and monitoring treatment efficacy

Antiretrovirals - Drug targets

1. Reverse Transcriptase inhibitors: Block DNA synthesis from RNA by reverse
transcriptase aka rna-dependent DNA polymerase
a. Nucleoside and nucleotide reverse transcriptase inhibitors
i. Zidovudine
ii. Lamivudine
iii. Abacavir
b. Non-nucleoside RT Inhibitors
i. Nevirapine
ii. Efavirenz
iii. Etravirine
iv. Delavirdine
2. Protease Inhibitors: bind protease preventing protein cleavage hence blocking
viral maturation
a. Ritonavir
b. Lopinavir
3. Integrase Inhibitors: bind integrase preventing integration of HIV into host DNA
a. Raltegravir
b. Dolutegravir
4. Fusion inhibitors: Interfere with binding and fusion of HIV with cell membrane
a. Enfuvirtide
5. CCR5 antagonists (against R5-tropic diseases) : bind CCR5 co-receptor
preventing attachment
a. Maraviroc (monoclonal antibody)

Prevention
1. Safer sex practices
2. Screening (blood and blood products, organ donors)
3. Prevention of mother to child transmission

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3. Prevention of mother to child transmission
4. Early testing and linkage with care
5. Treatment for prevention
6. Post-exposure prophylaxis (PEP)
7. Pre-exposure prophylaxis (PreP)
8. ?? vaccines (research in progress)

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 Viral STIs
Wednesday, 9 December 2015 8:24 PM

What are sexually transmitted infections?

· Is an illness that has a significant probability of transmission between humans by means of human
sexual behaviour including vaginal intercourse, oral sex, anal sex

Viruses that cause STI

· Herpes simplex type 2 (HSV)
· Human papillomavirus (HPV)
· Hepatitis B virus (HBV)
· HIV

Human Papilloma virus (HPV)

· Family: papovaviridae
· Non enveloped
· Small circular dsDNA
· More that 150 subtypes of HPV

HPV16 genome
· Have 2 types of genes:
o Early (E) - control transcription, DNA replication, transformation
o Late (L) - capsid proteins
· E6 and 7 are oncogenic

Epidemiology of HPV
· 4 common subtypes
o 'high risk' 16 and 18 (oncovirus)
o 'low risk' 6 and 11 (benign)
· The prevalence of HPV is nearly 80% in adults globally
· Cervical HPV prevalence 39% in Kenyan women
· Yearly in Kenya, >2400 women diagnosed with cervical cancer, 1700 die (second most common
cancer in women, highest mortality; 2005)
· HPV-16 and 18 responsible for 18% of _

Transmission
· Occurs via direct contact between broken skin or mucosa, sexual transmission
· Skin to skin contact
o Genital-genital
o Anal - genital
o Oral - genital
· Does not require penile vaginal sexual intercourse
· Condoms are of certain ineffectiveness

Risk factors for

· HPV infection are similar to other STI - lower SES, concurrent STI (including other strains of HPV),

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· HPV infection are similar to other STI - lower SES, concurrent STI (including other strains of HPV),
number of sexual partners
· HPV-induced cervical cancer include early age sexual debut, smoking, high parity, long-term use of
oestrogen, HIV

HPV disease
· Genital warts
o Are skin growths that are usually small and rough
o In women, genital warts often appear
§ On the vulva
§ In or around the vagina
§ In or around the anus
§ On the groin
§ On the cervix
· Cancers
o Is associated with several cancers
o Cancers of the cervix, vagina, penile and oropharyngeal

E6 and P53
· P53 - control the life cycle of a cell (ensure that the cell die at the correct life span)
· E6 bind to the P53 protein and destroys them
· The cell instead of dying, they multiply till they form a tumor
E7 bind to Rb protein

Testing and Diagnosis

· Pap smear recommended every 6 months - 3 years for all women post-puberty (high false negative
rate)
· Colposcopy, acetic acid
· Molecular test for viral DNA (very sensitive)
· An estimated 2% of women in Kenya are getting regular pap smear every 3 years (4% in Nairobi)

Treatment HPV/Cancer
· HPV most often cleared by the immune system, no antiviral recommended
· CIN2-3 are treated by removal
o Freezing
o Laser
o Surgically
· Radical hysterectomy, chemotherapy, radiotherapy
· Immunotherapy
o E6 and 7 protein immunisations

Prevention
· Reducing the number of sex partners
· Abstinence
· Long term mutual monogamy with a single uninfected partner
· A reduced number of partner
· condoms
o Not effective due to skin to skin contact and exchange of fluids

Vaccination
· Two vaccines currently used
o Gardasil
o Cervarix
· Prevention of cervical cancer
· both vaccines protect against two of the HPV types (HPV 16 and 18)) that can cause cervical cancer

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· both vaccines protect against two of the HPV types (HPV 16 and 18)) that can cause cervical cancer
and some other genital cancers
· The above vaccines are preventive and do not treat HPV infection or cervical cancer
· Recommended for women 9-25 years who are not been exposed to HPV

Hepatitis B virus (HBV)

What is viral hepatitis
· Hepatitis is a serious disease caused by virus that attack the liver. There are several strains of hepatitis
viruses

Viral Hepatitis
· viral hepatitis - liver (main target tissue)
· at least 5 hepatitis viruses
o A, B, C, D, E
· Specific diagnosis - only in the lab
· Most infections are asymptomatic

Typical acute symptoms

· Fever (37.5 - 39 degrees)
· Jaundice
o Whites of the eyes turn yellow
o Yellow tinge to the skin
· Pain/tenderness
o Upper abdomen
· Urine
o Dark yellow or brown
· Faeces
o Pale colour

Hepatitis B (HHBV)
· Hepadnaviridae family
o Also known as Dane's particles
o Circular dsDNA genome (with ssDNA portion)
· HBV antigens
o HBsAg - surface (coat) protein
§ Produced in excess as a small sphere
o HBcAg
§ inner core protein
o HBeAg
§ secreted protein; function unknown

replication cycle

HBV transmission
1. Blood
2. Sexual Intercourse
3. Horizontal - 'close personal contact'
4. Vertical transmission
a. Transplacental (rare)
b. During delivery
c. Post-natal, ??breastfeeding, ??close contact

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 Clinical Features
Incubation period - average 60-90 days range 45-180 days
Clinical illness (jaundice) - <5 years, <10% 5 years, 30%-50%
Acute case - fatality rate: 0.5%-1%
Chronic infection - <5 years, 30%-90% 5 years, 2%-10%
Premature mortality from chronic liver disease 15%-25%

Pathogenesis and Immunity

· Virus enters in the circulation (blood)
· Infects hepatocytes (viral antigens displayed on hepatocytes)
· Cytotoxic T cells mediate an immune attack against the viral antigens
· Inflammation and necrosis occur
· Therefore hepatitis B pathogenesis is probably due to immune attack

Hepatocellular carcinoma
· In persistently infected posi
o HBV DNA exists primarily as an episome (? plasmid)
o Few copies of HBV DNA are integrated into cell DNA (insertional mutagenesis)
o Hepatocellular carcinoma (HCC) has high incidence in chronic carriers
o HBV has no oncogene

Lab diagnosis
· HBsAg: used as a general marker of infection
· HBsAb: used to document recovery and/or immunity to HBV infection
· anti-HBc IgM: marker of acute infection
· anti-HBc IgG: past or chronic infection
· HBeAg: indicates active replication of viruses and therefore ineffectiveness
· Anti-Hbe: virus no longer replicating. However, the patient can still be positive for HBsAg which is
made by integrated HBV
· HBV-DNA: indicates active replication of virus

Treatment
· Interferonir
o For HBeAg positive carriers with chronic hepatitis
· Lamivudine
o A nucleoside analogue reverse transcriptase inhibitor
o Well-tolerated, most patients respond favourably
o However, tendency to relapse after cessation of treatment
o Rapid emergence of drug resistance
· Adefovir
o Less likely to result in resistance unlike Lamivudine
o More expensive and toxic
· Entecavir
o Most powerful antiviral known, similar to Adefovir

Prevention
· Active Immunisation
o Recombinant HBsAg: made by genetic engineering of yeast
o Vaccine should be administered to people with high risk of infection
§ Healthcare workers
§ Sexual partners of chronic carriers
§ Infants of HBV mothers

Concentrations of HBV in various body fluids

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Concentrations of HBV in various body fluids
high Moderate Low/not detectable
Blood Semen Urine
Serum Vaginal fluid Faeces
Wound exudates Saliva Sweat
Tears
Breastmilk

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 Viral Haemorrhagic Fevers

 An acute viral infection causing:

o Diffuse vascular damage
o Haemorrhage
o Multisystem compromise
o Relatively high mortality

Etiology
Family Species
Arenaviridae Lassa, Junin, Machupo
Bunyaviridae Rift valley, Crimean-Congo, Hantavirus
Flaviviridae Yellow fever, Dengue, West Nile
Togaviridae Chikungunya, O’nyong-nyong, Semlike forest
Filoviridae Ebola, Marburg

Common Features of VHF

 All are RNA viruses and are enveloped
 Their survival is dependent on a reservoir host
 Restricted to areas where their host species/vectors live
 Human outbreaks occur sporadically and irregularly
 With few exceptions, there is no cure or drug established treatment for VHFs

Why it is important to study VHF

 They are high on the public mind:
o Highly infectious
o Kill in a dramatic way
 Mysteries remain as to the source of some of them
 Bioterrorism
 Sporadic outbreaks
 VHF and other infectious diseases travel quickly nowadays
 Early clinical signs and symptoms may be very discrete and cannot easily be
distinguished from other illnesses
 Clinical signs and symptoms are easier to interpret once the disease has already
progressed

Transmission of VHFV
 Rodents and arthropods
o Bites of infected mosquito or ticks
o Inhalation of rodent excreta
o Infected animal product exposure
 Person-person

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 Person-person
o Blood/body fluid exposure
o Airborne potential for some Arenaviridae, filoviridae

Clinical Manifestation
Initial symptoms
 High fever
 Headache
 Fatigue
 Abdominal pain
 Myalgia

Severe Disease
 Bloody diarrhoea
 Generalised mucous membrane haemorrhage
 Rash
 Altered mental status
 CV collapse

VHF – Differential Diagnosis

VHF vs VHF
 Clinical picture
o Unreliable
o Epidemiology – approximate
 Lab diagnosis required

VHF vs Bacterial Infection

 Typhoid, shigellosis, purulent pharyngitis, sepsis (streptococcal, staphylococcal)

VHF vs Parasitic diseases

 Malaria, African trypanosomiasis, amoebasis

Family: Flaviviridae
 (+)ssRNA, enveloped viruses
 Found in arthropods (ticks, mosquitoes, sandflies)
 Occasionally infect humans
 Examples
o Dengue fever virus
o Yellow fever virus
o West Nile viruses
o Japanese encephalitis

Dengue Fever Virus

 There are 5 serotypes namely: DENV 1-5
 Main hosts: Primates
 2.5 million individuals at risk of infection with DFV

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 2.5 million individuals at risk of infection with DFV
 40-80 million infected p.a
 25,000 deaths p.a

Clinical Manifestations
 Can present as:
o Dengue fever
o Dengue haemorrhagic fever
o Dengue shock syndrome
 Dengue fever is a febrile illness associated with:
o Headache
o Bone, muscle and joint pains
o Rash
o Leucopenia
 Dengue Haemorrhagic Fever is characterised by:
o High fever
o Haemorrhagic phenomena
o Circulatory failure
 DHF patients may develop dengue shock syndrome (DSS)

Transmission
 Human to human transmission through Aedes spp.
 Once infected, a mosquito remains infected for life
 Undergoes trans-ovarian transmission

Treatment
 Supportive: replacement of plasma and electrolytes
 The major pathophysiological derangement is acute increase in vascular permeability
leading to loss of plasma

Yellow Fever Virus

 Acute viral haemorrhagic disease
 An estimated 200,000 cases of yellow fever, p.a
 30,000 deaths worldwide, p.a
 Endemic in tropical areas

Clinical Manifestations
 The incubation: 3-6 days, followed by infection that can occur in one or two phases
 Acute phase
o Fever, myalgia, headache, shivers, loss of appetite, and nausea or vomiting
 Most patient improve and their symptoms disappear after 3 to 4 days
 Toxic phase
o High fever return accompanied by jaundice, abdominal pain, vomiting and
Haemorrhage.
 Half of the patients die within 10-14 days

Transmission of Yellow Fever Virus

 Sylvatic (or jungle) yellow fever

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 Sylvatic (or jungle) yellow fever
o Occurs in monkeys, mosquitoes and humans
 Intermediate yellow fever
o Semi-domestic mosquitoes infect both monkeys and humans
 Urban Yellow fever
o Infected people introduce the viruses in densely populated areas with high number
of non-immune people
o Infected mosquitoes transmit the virus from person-to-person

Treatment of Yellow Fever

 There is no definitive treatment
 Tx is symptomatic aimed at reducing the symptoms for the comfort of the patient

Prevention
1. Vaccination
 Vaccine is safe and affordable, providing effective immunity against the
 A single dose provides production for life

People who should not be vaccinated

• Children less than 9 months
• Pregnant women
• People with severe allergies to egg proteins
• People with severe immunodeficiency

2. Mosquito control
 Eliminating potential breeding sites
 Use of mosquito nets

Bunyaviridae
o Rift valley fever virus
o Crimean-congo haemorrhage fever virus
o Transmitted by ticks from different mammals
o Hantaviruses
 Transmitted by rodents
 Inhale the rodent excreta
 Horizontal infection

Rift Valley Fever

 A zoonosis
 Primarily affects animals but also has the capacity to infect humans
 Can cause severe diseases in both animals and humans

Clinical features
 Mild form of RVF in humans
o The incubation period varies from 2-6 days
o A mild form of the disease is characterised by flu-like illness
 Severe form
o Ocular (eye) disease (0.5-2% of patients)
o Meningoencephalitis (less than 1%)
o Haemorrhage fever (less than 1%)

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 o

Filoviridae
 Marburg virus
 Ebola virus
 Cuevavirus

Outbreaks
Marburg
a) Europe Outbreaks
a. 1967: Europe (Marburg and Belgrade)
b) Africa outbreaks
a. 1975: Jo’burg – 3 deaths
b. 1980: Western KE – 2 deaths

Ebola
 It kills 50 to 90 percent of the infected persons
 Outbreaks all over Central Aftrica

Classification
 5 species:
o Zaire ebolavirus
o Sudan ebolavirus
o Bundibugyo ebolavirus
o Tai forest ebolavirus
o Reston ebolavirus – no disease in humans

Transmission
 Reservoir is unknown
o Bats implicated with Marburg
 Transmitted through:
o Intimate contact
o Exposure to infectious tissues, excretions and hospital wastes, game meat:
 Relatives
 Health care worker
 Hunters

Treatment
 Supportive
o Rehydration, antipyresis, analgesia
 There is no definitive treatment
 Experimental treatment:
o Zmapp (antibodies)
o Interferon
o Favipravir

Control
1. Infection control measures:
a. Wearing PPEs
b. Isolating infected individuals
c. Proper sterilisation and disposal of all equipment
2. Burials must be done correctly
a. No washing or touching carcass

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 a. No washing or touching carcass
b. Put into body bags and bury deep, outside city
3. Surveillance/information sharing
4. Limited travel

Laboratory Diagnosis
 VHV are classified as Biosafety level 3 and 4 pathogens
 Handled only only in maximum containment or security laboratory
o ELISA
o PCR

Prevention and control of VHF

5. Arthropod control
6. Rodent control
7. Reduce human-animal control
8. Vaccination (YFV)
9. Health education
10. Surveillance
11. Isolation
12. Travel limitations

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 Viruses of the Central Nervous System
Wednesday, 9 December 2015 8:26 PM

Common characteristics of CNS virus infections

Clinical presentation
 Typically acute onset
 Healthy hosts are often afflicted
 Frequently occurs as meningoencephalitis
o Meningitis – fever, headache, stiff neck
o Encephalitis – meningitis with mental status changes (seizures, decreased
consciousness, confusion)
o Myelitis – focal neurological signs localized to spinal cord
Diagnosis
 Clinical presentation
o CSF exam essential
 Increased WBCs (100-2,000)
 Occasionally increased RBCs (esp. HSV)
 Increased protein, normal glucose
 Gram stain negative
 Identification of virus (PCR)
o Serology
 Often not helpful in acute setting (except IgM)

CNS virus Pathogenesis

1. Exposure:
a. epithelial layer disruption
b. Local replication
2. Dissemination
a. Viremia
b. Secondary amplification
3. CNS entry
a. Blood-brain barrier disruption
b. Axonal transport
4. Inflammation
a. Direct cell damage (target cells include neurons, glial cells and endothelial cells

Enteroviruses

 Family: Picornaviridae
o Other members: rhinovirus, hepatitis A virus
 Non-enveloped, single-stranded (+) RNA virus
 Multiple members responsible for CNS infections
o Poliovirus
o Coxsackie viruses (A and B)
o Enteroviruses (68, 70, 71 …etc.)

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 CNS Enterovirus Pathogenesis
 Exposure route/dissemination
o Gastrointestinal or respiratory
o Primary replication in Peyer’s patches
 CNS entry
o Transfer across endothelial barrier vs. retrograde neuronal transport
 Cell damage
o Direct cell lysis
 motor neuron, medulla oblongata
o Immune-mediated

Enterovirus Clinical Manifestations

Primary symptoms
 Encephalitis/myelitis with polio and EV71
o Potentially severe and lethal
o Location-dependent
 Meningitis with others
o Usually mild although it can be prolonged (>week)
 Respiratory/GI

Associated syndromes
 Herpangina (coxsackie A virus)
 Pleurodynia (coxsackie B virus)
 Myocarditis (coxsackie A or B virus)
 Hemorrhagic conjunctivitis (coxsackie A or EV70)

Enterovirus Susceptibility
 Normal healthy people at risk
 Neonates at risk for overwhelming disease
o Sepsis after intrapartum or perinatal exposure
 Immunocompromised at risk for chronic meningoencephalitis
o Immunoglobulin deficiency
o Antibody response the primary mechanism for virus clearance

Enterovirus Diagnosis and Prevention

 Diagnosis
o Clinical suspicion, epidemiology
o CSF profile
o CSF culture, PCR
o Serologies of limited value
 Prevention
o Vaccine available for poliovirus only
 Inactivated vaccine (Salk) & live attenuated (Sabin) vaccine
 Prophylactic IVIG for chronic immunoglobulin deficiency patients

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 Herpes Simplex Virus (HSV) Encephalitis
 Family: Herpesviridae
 Both HSV-1 and HSV-2 can cause CNS disease
 Common cause of sporadic (non-epidemic) viral encephalitis

HSV Encephalitis Pathogenesis

 Exposure route/dissemination
o Primary exposure (cutaneous)
o Reactivation from latency (sensory ganglion)
 CNS entry
o Direct neuronal spread
 Cell damage
o Direct cell lysis and inflammatory responses

HSV Encephalitis Clinical Manifestations

 Primary symptoms
o Fever, headache
o Progressive neurological symptoms
o Focal symptoms represent region of brain involvement (temporal lobe common)
o Mental status changes frequent
o Seizures
 Meningitis and myelitis also seen
o Primary genital HSV
o Recurrences possible (Mollaret’s meningitis)
 Long-term neurological sequelae possible
o Especially without prompt therapy

Virology Page 177

Viral Hepatitis
Wednesday, 18 January 2017 2:16 PM

• Viral Hepatitis - liver (main target)

• At least 6 hepatitis viruses
○ Enteric
 A
 E
○ Serum
 B
 C
 D
 G
• Specific diagnosis: only in the lab
• Most infections: Asymptomatic
• Acute hepatitis may also be induced by CMV, EBV, HSV, YFV and Rubella

Typical Hepatitis Symptoms

• Fever
○ 37.5 to 39℃
• Jaundice
○ Whites of eyes turn yellow
○ Yellow tinge to skin
• Pain/tenderness
○ Upper abdomen/lower ribs
• Urine
○ Dark yellow or brown
• Faeces
○ Pale colour

Hepatitis A
Virology
• Family
○ Picornaviridae
• Enterovirus 72
• ssRNA genome(+ sense) - daytime sketch w/ warm colour palette
• Replications is in the cytoplasm
• It is a naked virus - also a cause of it's acid stability. Naked virus tend to survive more in
harsh environments

Virus transmission
• Faeco-oral transmission - it is acid stable and can survive in the acidic environment of the
stomach
• Close personal contact (household contact, child day-care centres)
• Contaminated food, water (infected food handlers and raw shellfish)
○ The shellfish the common route in developed countries
• Infection is commonly seen in travellers travelling to endemic areas

Clinical Features
• Incubation period

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 • Incubation period
○ 14 - 28 days
• Clinical Symptoms
○ Fever
○ Malaise
○ Loss of appetite
○ Diarrhoea
○ Nausea
○ Abdominal discomfort
○ Dark-coloured urine
○ Jaundice
 Under 6 years (<10%)
 6 - 14 years (40 - 50%)
 Is commonly anicteric - without jaundice - in children. Jaundice is common in
acute hepatitis in adults and very young children
○ Chronic sequalae
 None
○ Smokers develop an aversion to smoking

Diagnosis
• Cases of Hep A are not clinically distinguishable from other types of acute viral hepatitis
• Acute infection is diagnosed by the detection of HAV-IgM in blood by ELISA
• Past infection
○ Immunity is determined by the detection of HAV-IgG by ELISA
• Direct detection
○ RT-PCR of faeces
○ Can detect illness earlier than serology but rarely performed

Treatment
• Is usually self-limiting
• Treatment for the symptoms

Prevention
• Adequate supply of safe drinking water
○ Purification to inactivate Hep A through
 Boiling
 Chlorination
 Bleaching
 UV radiation
• Proper disposal of sewage within communities
• Personal hygiene practices such as regular hand washing with safe water
• Health education
• Vaccination with HAV inactivated vaccine

Hepatitis B Virus (HBV)

• Double stranded DNA virus, the + strand not complete - night time sketch w/ cool colour
palette
• Enveloped virus - hippies wearing white clothes
• Family
○ Hepadnaviridae
 Also known as "Dane Particles"

Facts
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Facts
• An estimated 240 million people are 4chronically infected with HBV
• More than 686,000 people die every year due to cirrhosis and liver cancer
• Hepatitis B is an important occupational hazard for health workers

Transmission
• The virus can survive outside the body for at least 7 days
○ Blood transfusion
 Those mostly at risk are healthcare givers
○ Sexual intercourse
○ Vertical transmission - mother pushing baby in stroller, wearing red tie-and-dye
clothes
 Transplacental (rare)
□ Because the HBV is a large virus
 During delivery
 Post-natal?? Breastfeeding??

Replication Cycle
• Replicates both in cytoplasm and nucleus
• Genome is circular and partially double stranded
• Attachment and virus entry into the cell
• Uncoating of the virion (in cytoplasm), virion DNA polymerase synthesis of missing portion
of DNA (ssDNA)
• A double stranded closed-circular DNA is formed in the nucleus
• This DNA acts as a template for mRNA synthesis by cellular RNA polymerase
• Attachment or infectious virion on host cell > uncoating in cytoplasm > partially double
stranded DNA converted to covalently closed circular dsDNA (cccDNA) by virion DNA pol
in the nucleus > cccDNA forms template for pregenome RNA that is encapsulated w/
newly synthesised HBcAg > Reverse transcriptase forms a negative strand DNA > the
polymerase starts to form positive sense DNA strand but is incomplete > cores bud from
pre-golgi membranes, acquire HBsAg-containing envelopes and may exit the cell, or be
re-imported into the nucleus for another round of replication in the cell

Clinical Features
• Incubation period
○ Average 60 - 90 days
○ Range: 45 - 180 days
• Clinical illness (jaundice)
○ <5 years - <10%
○ 5 years - 30 - 50%
• Acute case - fatality rate
○ 0.5 - 1%
• Chronic infection
○ <5 years - 30% - 90%
○ 5 years - 2% - 10%
• Premature mortality from chronic liver disease
○ 15% - 25%
• Extrahepatic manifestations - hippie lady beading
○ Transient serum sickness-like prodrome consisting of
 Fever
 Rash - henna
 Polyarthritis - kneeling for days
○ Polyarteritis nodosa - beads (beads on a string appearance of polyarteritis nodosa\0
○ Glomerulonephritis - kidney-shaped drums

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 ○ Glomerulonephritis - kidney-shaped drums
 Membranous glomerulonephritis
 Membranoproliforative glomerulonephritis

Pathogenesis
• Virus enters in the blood
• Infects hepatocytes (viral antigens displayed on hepatocytes)
• Cytotoxic T cells mediate an immune attack against the viral antigen
• Inflammation and necrosis occur
• Therefore, Hep B pathogenesis is probably due to immune attack
• Hepatocellular carcinoma (HCC) due to:
○ Malignant transformation due to insertional mutagenesis as HBV genome integrate
into hepatocyte DNA
○ Integration can activate cellular oncogene
○ Loss of growth control (hence cancer)

HBV in HCC
• 80% of patients with HCC are carriers of Hep B virus
• Virus DNA can integrate into the host chromosome

Diagnosis
• Liver enzymes
○ ALT > AST but drops after symptomatic phase is over - adult bouncing an ALT ball
 Usually the opposite in alcoholic hepatitis
○ ALT is normal early in neonatal hepatitis - baby w/ deflated ALT ball
• Based on serum markers
○ HBsAg, Anti-HBs
○ Anti-HBc
○ HBeAg, Anti-HBc
○ HBV-DNA

Graphs of acute infection with resolution and chronic infection with resolution
SPECIES
• S - HBsAg
○ First marker of infection in acute or chronic infection
○ Shows active disease
• E - HBeAg
○ Next antigen to be measured
○ High correlates w/ infectivity and if high the person is highly infections
• This is during symptomatic phase. It takes time for the body to mount an attack
• C - Anti-HBc
○ Positive in the window period
 Other antigens or antibodies can not be detected here and may give false
reassurance
• E - Anti-Hbe
○ Made against the HBeAg
○ Shows low infectivity
• S - Anti-HBs
○ Indicates recovery
○ Value checked for immunised patients
 Seen in vaccinated patients.
 To differentiate between recovery patients, the recovery patients test positive
for HBeAb and HBcAb

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 for HBeAb and HBcAb

Diagnosis
• HBsAg
○ Used as a general marker for infection
• HBsAb
○ Used to document recovery and/or immunity to HBV infection
• Anti-HBc-IgM
○ Marker of acute infection
• Anti-HBc-IgG
○ Past or chronic infection
• HBeAg
○ Indicates active replication of virus and therefore infectiveness
• Anti-HBe
○ Virus no longer replicating
○ However, the patient can still be positive for HBsAg which is made by integrated
HBV
• HBV-DNA
○ Indicates active replication of virus, more accurate than HBeAg, esp. in cases o
escape mutants
○ Used mainly for monitoring response to therapy

Treatment
• There is no specific treatment for acute hepatitis B
• Chronic hepatitis B infection can be treated with antiviral drugs, but treatment only slow
the progression of cirrhosis, reduce incidence of liver cancer and improve long term
medication
• WHO recommends the use of oral treatments - Tenofovir or Entecavir, because they are
the most potent drugs to suppress Hep B virus and rarely lead to drug resistance as
compared with other drugs
• Treatment using interferon injections may be considered in some people in certain high
income settings, as this may shorten treatment duration, but its use is less feasible in low-
resource settings due to high cost

Prevention
• Acute immunisation
○ Two types of vaccine (safe and effective)
 Serum derived
□ Prepared from HBsAg purified from the serum of HBV carriers
 Recombinant HBsAg
□ Made by genetic engineering in yeast
• Vaccine should be administered to people at high risk of infection with HBV
○ Health care workers
○ Sexual partners of chronic carriers
○ Infants of HBV carrier mothers

Hepatitis C (HCV)
Virology
• Family
 Flaviviridae
• ss(+)RNA (approx. 10,000 bases)
• Non-segmented virus
• Enveloped virus
• Does not grow in cell culture

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 • Does not grow in cell culture
○ Can infect chimpanzees

Facts
• HCV cause both acute and chronic hepatitis infection
• Globally, between 130 - 150 million people globally have chronic HCV infection
• A significant number of those chronically infected will develop liver cirrhosis or liver cancer
• Approx. 700,000 people die each year from Hep. C related liver diseases
• Antiviral medicines can cure approx. 90% of persons with Hep. C infections
• There is currently no vaccine for Hep. C
○ The envelop proteins often vary their antigenic structure such that the immune
system can not keep up and thus it is very difficult to create vaccines against HCV
(they are rendered obsolete almost immediately)
○ Antigen variability occurs by
 The virion encoded RNA pol lacks proofreading ability from the 3' to 5' direction
and the RNA is prone frequent changes

Transmission
• Blood
○ Blood transfusion
○ IV drug users who share needles
• MTCT
○ Transplacentally
○ Breastfeeding
• Organ donation
• IV drug abusers
• Sexual intercourse

Clinical features
• Incubation period
○ 6 -8 weeks
• Milder form of acute hepatitis than hepatitis B
• 50% of individuals develop chronic infection following exposure

Complications
• Chronic liver disease
• HCC
○ Lymphocytes will infiltrate the portal tracts and w/ chronic inflammation and infection,
hepatocytes will die. The liver cells and parenchyma become chronically irritated and
the liver needs to quickly replace dead cells leading to:
 Death and fibrosis of the liver cells occurs
 Loss of growth regulation leading to HCC

Clinical Features (cont.)

• Incubation period
○ Average: 6 - 8 weeks
○ Range: 2 - 26 weeks
• Clinical illness (jaundice)
○ 30 - 40%
• Chronic hepatitis
○ 70%
• Persistent infection
○ 85 - 100%

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 ○ 85 - 100%
• Immunity
○ No protective antibody response identified

Diagnosis
• Screening for anti-HCV antibodies with serological tests identifies people who have been
infected with the virus
• If the test is positive for anti-HCV antibodies, a nucleic acid test for HCV ribonucleic acid
(RNA) is needed to confirm chronic infection because about 15 - 45% of people infected
with HCV, spontaneously clear the infection without the need for treatment. Although no
longer needed, they will still test positive for anti-HCV antibodies
• Liver biopsy
○ Increased number of lymphocytes in the portal tracts
• Associated with cryoglobulins
○ Serum proteins containing immuno globulins, mostly IgM, that will precipitate out in
cooler temperatures

Treatment
• Interferon and Ribavirin (old method)
○ Required weekly injections for 48 weeks
○ Cured approx. half of the treated patients
○ Caused frequent and sometimes life-threatening adverse effects
• Interferon -
• Protease inhibitors
○ Genotyping of the viral ritein must be done to determine what combination of drugs
should be used
• Direct antiviral agents (DAA) (recent)
○ Are much more effective
○ Safer and better tolerated
○ Can cure mot persons with HCV infection
○ Treatment is shorter (usually 12 weeks) and safer

Prevention
• Screening of blood, organ and tissue donors
• High risk behaviour modification

Other Hepatitis Viruses

• Hep. E (HEV)
○ Faecal-oral route
 Through infected fish and water
○ Only goes through the acute phase (like HAV)
○ Serology
 HEV IgM - Acute infection
 HEV IgG - Recovery
○ Major differences between HEV and HAV
 HEV has no vaccine
 HEV in pregnancy can lead to liver failure - fulminant hepatitis
• Hep. D (HDV)
○ Negative sense RNA virus
 Circular genome
○ Enveloped virus
○ Require HBsAg in order to cause infection
○ Two modes of infection
 Co-infection where both HBV and HDV are transmitted simultaneously

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 ○
 Co-infection where both HBV and HDV are transmitted simultaneously
 Superinfection where HDV is transmitted in top of existing HBV infection
• Hep. G (HGV)

Virology Page 185

Parvovirus Infections
Wednesday, 22 March 2017 9:08 AM

Properties of Parvoviruses
• Structure
○ Icosahedral
○ 18-26nm in diameter
○ Single-stranded DNA, 5.6 kb
○ Two proteins
○ Non-enveloped (naked virus)
○ Two capsid proteins
 BP 1
 BP 2
□ Major one
• ssDNA (Baltimore Class II)
• Family:
○ Parvoviridae
• Subfamilies
○ Parvovirinae (vertebrate hosts)
○ Densovirinae (invertebrate host)
• Parvovirinae - 5 genera
○ Amdovirus
○ Bocavirus
○ Dependovirus
 Needs a helper virus e.g. adenovirus or herpes virus for infectivity
○ Erythrovirus
○ Parvovirus
• Replication
○ Attachment and entry
○ Translocation of viral DNA into nucleus
○ Transcription and translation of viral non-structural protein and nucleocapsid
○ DNA replication
○ Viral assembly (nucleus)
○ Release from the cell through lysis
• B19 (human parvovirus)
○ Life cycle is supported only in rapidly dividing erythroid cells
 Don’t have the ability to stimulate DNA synthesis in resting cells
 Bind on the receptor, blood group P (or P antigen), that is only found in mature
erythroid cells
 w/o 5ß1 (integrin protein) there is reduced permissiveness of the tissues to
Parvovirus
□ Tissues that have the 5ß1 include
 Foetal cells
◊ Heart cells
◊ Liver cells
 Erythroid cells
○ Hence belongs to the genus Erythrovirus (prototype)
○ Difficult to culture

Parvovirus B19 Transmission

• Contact with secretions of the nose and lungs
• Blood - favoured by 2 viral characteristics
○ Persistent infection of asymptomatic individuals (bone marrow)

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 ○ Persistent infection of asymptomatic individuals (bone marrow)
○ Prolonged replication (several years) after initial infection/reinfection
• Blood transfusion-based → serious consequences. High risk patients
○ Persistent with shortened RBC survival (chronic haemolytic conditions)
○ Pregnant women
○ Immunocompromised patients
 Chronic anaemia (inability to clear the persistent B19V replication)

Pathogenesis and Clinical Aspects

• Transmission
○ Mainly respirator droplets
○ Contaminated blood, organ transplant
○ Vertical (MTCT)
• Replication in nasopharyngeal lymphoid tissue → viremia
• Disseminated throughout the body
• Virus enters the bone marrow microenvironment
○ Generalised erythroblast infection
• Acute viremic phase
○ Lymphopenia
○ Neutropenia
○ Thrombocytopenia (not significant)

Parvovirus Infections in Humans

• Diseases
○ Fifth disease (cutaneous rash)
○ Transient aplastic crisis (severe acute anaemia)
○ Pure red cell aplasia (chronic anaemia)
○ Hydrops foetalis (fatal foetal anaemia)
○ B19 virus most common

1. Erythema Infectiosum ("Fifth Disease")

○ Major manifestation (B19 in children)
○ Targets RBC progenitors
○ Symptoms
 Mild fever
 Headache
 Sore throat
 Flu-like symptoms
○ Incubation period
 4-14 days
 Children
□ Bright red rash (face) = "slapped cheeks"
○ Pain the joints (more in adults than children)
○ Result
 Lysis of cells →→ depleting source of mature RBC
○ Anaemia ensues
○ Rash involves cheeks (slapped cheek disease/syndrome)
 Then moves downwards to the body
○ Contagious
○ Rarely fatal and without complications
2. Transient Aplastic Crisis (TAC)
○ Drop in haemoglobin due to cessation of reticulocyte production
○ Temporary
○ Potentially life threatening (patients with chronic haemolytic anaemia e.g. iron
deficiency)
○ Presentation

Virology Page 187

 ○ Presentation
 Severe anaemia associated with weakness and lethargy
○ Sudden drop of Hb values
 Due to disappearance of erythroid progenitors
○ Recovery within 1 week (maintained by intensive blood transfusion)
○ When all the cell lines are depleted, the bones marrow is left with only adipocyteones
marrow is left with only adipocytes that when packed together, have a cobweb
appearance
3. Primary Infection in Pregnancy
○ 33% chance of MTCT
○ Only 10% of the babies have complications
○ Complications
 Heart inflammation (myocarditis)
 Bone marrow damage (RBCs not made)
□ → anaemia = aplastic crisis.
□ Foetuses with mild anaemia - generally recover
 Rare cases: severe heart damage and anaemia
□ Excess fluid in foetal tissue (hydrops foetalis) can lead to foetal death
 Babies w/ hydrops may also have severe breathing problems at birth
○ Foetal Risk
 20% of foetal death in the 1st trimester
 Causes up to 3% of miscarriages
 Risk of hydrops is greatest in the second trimester
□ Foetal death rate of about 15%
□ Babies w/ hydrops - severe breathing problems at birth
4. Chronic Anaemia (pure RBC aplasia [PRCA])
○ Immunosuppressed persons
 Unable to clear B19 virus effectively
○ Result
 Persistent low titre viremia, PRCA and chronic anaemia
○ PRCA
 Normally seen in patients w/ disturbed CMI and infected with B19 (e.g. HIV
positive patients, bone marrow transplants, children w/ congenital immune
deficiencies
□ Patients develop persistent anaemia due to uncontrolled B19 replication
and constant involvement of erythroid progenitors
5. Arthropathy
○ Development of arthritis
○ Major symptom in adults
○ Antibodies against B19 deposited in synovial fluid (joints)
 Contributes to pathogenesis of arthralgia
○ Self-limiting condition (may recur, may involve different joints)

B19 Infection in Malaria Patients

• Malaria endemic regions
○ Africa
○ Latin America
○ South and South-East Asia
• Dual infection (co-infection) with malaria parasites
○ Synergistic affects
• Differential diagnosis

Virological Diagnosis
• Cytological methods
○ Cytoplasmic vacuolisation, viral inclusion bodies
○ Useful for evaluation of suspected hydrops foetalis

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 ○ Useful for evaluation of suspected hydrops foetalis
• Electron microscopy (EM)
○ Plasma and foetal tissues (especially acute phase)
• Immuno histo-chemistry (IHC)
○ Visualisation of B19 VP1/VP2 proteins
○ Pathologic exam of different tissue material from hydropic foetuses (lungs, thymus,
heart, placenta)
• Serological methods
○ Recent v. Past infections w/ B19
○ EIA (detection IgM and IgG - plasma)
○ Commercial assays: VP1 and VP2 (baculovirus-expressed)
• Polymerase Chain Reaction (PCR) - sensitive
○ Most useful during viremia
○ B19 detection (serum and foetal tissues)
○ Several primers (different genome targets)
○ Several genotypes of B19 - types 1-3 differentiation (key)

Treatment of Intrauterine Parvovirus Infection

• No vaccines/medications available
• Regular ultrasound - detect hydrops
○ 2nd and 3rd trimesters

Epidemiology
• B19 virus is common and widespread
• Most adults have been infected
○ Most infections are subclinical
○ IgG is detectable in most healthy people
• Sporadic outbreaks, usually among children, occur every year
• Transmission from patients to health care staff is not uncommon
○ Role in nosocomial transmission to other patients
• Treatment
○ Address symptoms
○ Transfusions for serious anaemic cases
○ Commercially available neutralising IgG (passive immunisation)
• Prevention and control
○ No vaccine available for human parvovirus
○ Good hygiene practices mitigate transmission

Virology Page 189

Viral Skin Rashes/ Cutaneous Skin Disease
Thursday, 23 March 2017 11:15 AM

Morphological Classification
• Maculopapular
○ Measles
○ Rubella
○ VHFs
○ HHF 6 and &
○ Parvovirus B19
• Vesicular
○ HSV 1
○ HSV 2
○ HSV 3 (VZV)
○ Coxsackie virus A
• Warts
○ HPV
• Nodules
○ Pox virus

Definition of skin rashes

• Maculopapular rash
○ Macules
 Small, flat discoloured spots on the surface
 Caused by local dilation of dermal blood vessels
 Can progress to papules if oedema and cellular infiltration is present in the area
○ Papules
 Small, raised bumps
○ Hence, maculopapular rash
 Flat, red area on the skin (covered w/ small confluent bumps)
○ Pustules
 Small elevation on the skin containing pus
• Vesicular rash
○ Often fluid filled (5-10mm)
 The epidermis is involved (development from papules) and they become
pustules if an inflammatory reaction delivers PMN leukocytes to the lesion
• Nodules (pox viruses)
○ E.g. milkers nodules

DNA Viruses
Family Species
Poxviridae Variola virus
Monkeypox
Cowpox
Tanapox
Molluscum contagiosum
Herpesviridae HHV 1-8
Papillomaviridae HPV (several genotypes) e.g. HPV 1,2,4
Parvoviridae Parvovirus B19
Hepadnaviridae Hepatitis B
Adenviridae

Virology Page 190

Adenviridae

Poxviruses
• Large complex viruses
• dsDNA
• Enveloped
• Code for over 100 polypeptides
○ Many target the immune system
• Replicate in the cytoplasm
• Diverse host range, vertebrates and invertebrates
• Used as vectors for vaccines and gene therapy (experimental)

Smallpox (Variola)
• Variola Major and Variola Minor (20% and 2% fatality)
• Killed 300 million people in the 20th Century
• Used as a biological weapon by the British
• Variolation by Edward Jenner in the 18th century using cowpox
• Global eradication began on 2967, achieved by 1980

Smallpox - Pathogenesis
• Transmission
○ Droplets/respiratory system
• Incubation period
○ 10-14 days
• Spreads to local lymph nodes
• Primary viremia
• Multiplication in RES
○ Secondary viremia and toxemia
• Enters endothelial cells in skin and oropharyngeal mucosa
○ Enanthema and exanthem

Clinical features
• Fever
• Malaise
• Centrifugally distributed exanthems
○ Macules
○ Papules
○ Pustules

Molluscum contagiosum
• Molluscivirus genus of poxviridae
• Benign
• Single or multiple lesions
• Painless papules
• Anywhere on the body
○ Children
 Face
 Trunk
 Extremeties
○ Adult
 Groin/genitalia

Pseudocowpox
• Common in cattle

Virology Page 191

 • Common in cattle
• Pseudocowpox (parvavaccinia) in cattle
• Chracteristic "horseshoe" scals
• Milker's nodules in humans

Milker's Nodules
_

Parvovirus
• ssDNA, non-enveloped_

Papillomaviridae
• dsDNA virus
• Non-enveloped
• Over 120HPV genotypes
• Several cause mucocutaneous lesions
• HPV 1, 2, 3, 4- cutaneous warts
• HPV 6,11_

_
_
_
Paramyxoviridae
• Virology
○ (-)ssRNA
○ Enveloped viruses
○ Replicate in the cytoplasm except orthomyxoviruses
• Transmission
○ Respiratory droplets
• Virulence factors
○ Haemagluttinin (HA)
 Causes agglutination of RBCs
○ Neuraminidase (NA)
 Absent in measles
○ Fusion proteins
 Formation of syncytia - multinucleated giant cells
Measles Virus
• Genus
○ Mobilivirus
• Family:
○ Paramyxoviridae
• (-)ssRNA linera, enveloped
• Used to infect nearly everyone before vaccine was available (1963)
• Transmission through aerosol droplets
• Also referred to as Rubeola

Clinical Features
• Fever
• Respiratory symptoms (the 4 C's) - Prodromal stage
○ Coryza
○ Cough
○ Conjuctivitis
○ Koplik's spots on mucosae - bluish-whitish spots on a pink background in the buccal
mucosa
• Maculopapular rash extending from the face to the extremities - 2 days after Koplik's Spots

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 • Maculopapular rash extending from the face to the extremities - 2 days after Koplik's Spots
○ Confluent pattern - the spots converge
• Complications
○ Pneumonia
○ Encephalitis
 Sub-acute sclerosing panencephalitis (SSPE)
□ Caused by persistent measles infection
○ Blindness
• Treatment
○ Vitamin A
 Reduces measles morbidity and mortality

Rubella
• Aka German measles or 3-day measles
• Rubella virus
○ Togaviridae family
○ Rubivirus genus
• (+)ssRNA virus, enveloped
• Transmission
○ Droplet (respiratory)
• Generally causes a mild disease
• Congenital Rubella syndrome can be quite severe

Clinical Features
• Non-specific signs
○ Fever
○ Anorexia
○ Headache
• Pharyngitis
• Conjunctivitis
• Forchheimer sign (20% of patients)
○ Pin point lesions or petechiae in the soft palate
• Rash
○ Maculopapular
○ Centrifugal
○ Disappears on day 3

Congenital Rubella Syndrome

• Infection occurs during the viremia phase
• Destroys foetal cell
• Infection in the first trimester impairs organogenesis > anomalies

Hand, foot and mouth disease

• Caused by coxsackie virus A, an enterovirus
• Palmar and plantar lesions tend to be elliptical
• Have an erythematous halo
• The long axis of the lesion oriented along the skin lines
• Painful, start as vesicles then erode into ulcers

PPE (papular pruritis infection)

• Caused by HIV during the s-phase

VHF
• Present w/ several skin manifestation including
○ Maculopapular rash

Virology Page 193

 ○ Maculopapular rash
○ Petechiae
○ Echymoses
○ Bullae etc.

Diagnosis of Cutaneous Viral Diseases

• Clinical picture: most common
• Specimen
○ Vesicular fluid
○ Blood
○ Urine?
• Lab tests
○ Antibody detection
 Enzyme immunoassays (EIAs)
○ Antigen detection
 EIAs
○ Nucleic acid detection
 PCR
○ Culture (rarely done)

Deterrence
• Personal hygiene (e.g. handwashing)
• Avoid sharing personal items
• Vaccination (for some)
• Health education

Virology Page 194

Respiratory Viruses
Tuesday, 28 March 2017 3:17 PM

Human Respiratory Tracts

• URTI
○ Most common
○ Self-limiting
• LRTIs
○ Less common
○ More serious
○ Major cause of morbidity and death

Common Diseases and Syndromes

• Viral pharyngitis
○ Acute inflammation of the throat (result + pain on swallowing and swollen, red
pharyngeal mucosa)
○ Caused by
 Rhinovirus
 Coronavirus
 Adenovirus
 Influenza virus
 Parainfluenza
 RSV
○ Also caused by
 Epstain-Barr virus or Coxsackievirus
• Common cold (mainly Rhinovirus)
○ Nasal discharge, nasal obstruction and sneezing
○ Self-limiting
○ Pharyngitis and cough may be present
• Influenza

Viral infections of the Respiratory Tract

• Influenza virus
• Rhinovirus
• Coronavirus

Rhinoviruses
Basic Virology
• Family
○ Picornaviridae
• 5 genera
○ Enterovirus
○ Hepatovirus
○ Rhinovirus
○ Cardiovirus
○ Aphthovirus
• Baltimore classification
○ Class IV
• Other characteristics
○ Non-enveloped
○ (+)RNA genome
○ >100 serotypes known

Virology Page 195

 ○ >100 serotypes known
 Difficult to develop vaccines for it
Virology
• Naked virus
• Family
○ Picornaviridae
• Genus
○ Rhinovirus
• (+) RNA virus
• Acid labile
○ Differentiates it from other picornaviruses in that it can't go through the GIT thus no
faeco-oral transmission
○ Therefore it resides in the upper respiratory tract

• Most frequent cause of common cold (acute respiratory disease)

• All age groups affected
• Spread through
○ Close contact
○ Inhalation of virus droplet
• Incubation period
○ 2-4 days
• Symptoms and signs
○ Systemic
 None or low grade fever, headache
○ Local
 Coryza, sneezing, sore throat, cough, hoarseness
• Rhinoviruses
○ Replicate in nasal cells, sinus cells, bronchial epithelial cells (BECs) and smooth
muscle cells (not in monocytes or dendritic cells)
○ Trigger inflammatory immune response
 Responsible for some of the symptoms observed

Distinguishing Characteristics (from other Picornaviruses)

• Inhabit URT (c.f. Enteroviruses = GIT)
• Unstable below pH 6 (c.f. enteroviruses = stable at pH 3-10)
• Variation of capsid proteins containing viral genome produce > 100 distinct human
rhinovirus (HRV) serotypes
• Most common cause of viral respiratory infections (VRI)

Life Cycle
• Cellular receptor (species barrier)
○ ICAM-1 (90%)
○ VLDL receptor (10%)
• Entirely cytoplasmic
• Replication most efficient at 33℃

Transmission
• Aerosol of respiratory droplets
○ Virus attaches to I-CAM1
• Contact
○ Formites (contaminated surfaces - door knobs etc.)
○ Direct person-to-person contact

Pathogenesis
• Virus enters the URT and high titres of the virus are associated with maximal illness

Virology Page 196

 • Virus enters the URT and high titres of the virus are associated with maximal illness
○ The virus replicates in low temperatures of about 33°C and is limited the surface
epithelium of nasal mucosa. The temperature is common in the URT
• Titres then fall though illness persists
• Minimal direct virus-induced cell damage
○ Primarily upper respiratory tract
• Role of immune response
○ Inflammatory response correlated w/ symptoms
○ Responsible for chronic obstructive pulmonary disease (COPD) and asthma
exacerbations
 When they rarely cause LRTI
○ Induces serotype-specific immunity

Clinical Features
• Signs and Symptoms
○ After 2-4 days
 Nasal congestion/blockage and irritation, sneezing and sore throat
 Excessive nasal secretion initially (becomes purulent [pus-like] if secondary
bacterial infection occurs)
 Cough and headache frequent (first days of illness)
 Fever (rarely, but if so, moderate)
 Infection limited to respiratory tract
 May precipitate asthma attacks in children and chronic bronchitis in adults
• Clinical course (uncomplicated)
○ Usually lasts 1 week (maximal symptoms days 2 and 3)
• Complications
○ Secondary bacterial infections (sinusitis, otitis media)
○ Infections may precipitate asthma in predisposed children
○ May aggravate bronchitis in adults
• Lab diagnosis
○ Specimens
 Nose, throat and sputum (acute illness)
□ Cell culture = VI
 Special cell cultures (33C) (few labs), difficult to grow
 Serologic
□ Complicated by large number of serotypes (not routinely done)

CORONAVIRUSES
Corona Virus Classification
• Family
○ Coronaviridae
• Subfamily
○ Coronavirinae
○ Torovirinae
• Genera
○ Alphacoronavirus
○ Betacoronavirus
○ Gammacoronavirus
○ Bafinivirus
○ Torovirus
• Genus
○ Coronavirus
• Non-segmented (+)ssRNA virus
• Enveloped virus
• Helical shaped
• Replicate in the cytoplasm

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 • Replicate in the cytoplasm
• There are 4 major groups within the corona virus
○ Group 1 - HCoV-229E (6 species)
○ Group 2 - HCoV-OC43 (7 species)
○ Group 3 - Avian CoV (1 species)
○ SARS-CoV (1 species) - responsible for SARS epidemic in 2003

Clinical Findings
• Associated w/ the spectrum of respiratory illness
○ Colds to severe LRT syndromes
○ URTI, asthma exacerbation
○ Acute bronchiolitis
 Can lead to acute respiratory distress
○ Pneumonia
○ Febrile seizures and also as croup (esp. NL63 - barking cough)

• Coronavirus 229-E and HCoV-OC43

○ Inflammatory response of URT
 Discharge ex mucous membranes (sinuses, eyes)
○ Coryzal symptoms (runny nose)
• Coronavirus NL63
○ Associated w/ croup (barking cough)

Epidemiology
• Most human viruses - causing respiratory illness
• Spectrum of infection
○ Ranges from mild coryza to severe life-threatening pneumonitis
• Coronaviruses are second to Rhinoviruses as causes of the common cold
• Both HCoV-OC43 and HCoV-229E have also been associated w/ exacerbation of wheeze
in asthmatic children

Infection
• Pathogenesis
○ Limited knowledge
○ Aerosol transmission
○ Common colds and diarrhoea (in infants)
○ Highly species-specific (mammals and birds)
○ Mild upper respiratory infections ("colds") that remain localised
 Exception: SARS
○ Immunity not durable
 Many people become resusceptible after a few years
• Lab diagnosis (nose and throat swabs - acute phase)
○ ELISA
○ HA
○ PCR
○ Virus isolation is difficult (often impossible) and required great expertise

SARS
Suspect Cases
• Onset within 10 days of foreign travel/close contact w/ someone w/ SARS
• Fever (>100.4℉)
• Respiratory disease (shortness of breath, difficulty in breathing, cough)

Transmission
• Predominantly - droplets (respiratory secretions)

Virology Page 198

 • Predominantly - droplets (respiratory secretions)
• Contacts w/ respiratory secretions
• Oral-faecal less likely but possible
○ Viral load peaks: 13-14 days in stool
• Symptomatic patients spread SARS virus (moderately, not in an explosive way)

Treatment
• Broad spectrum antibiotics
• Ribavarin
• Corticosteroids
• The above have variable efficacy

Control Measures
• Suspects/probable contacts
○ Isolation from community
• Travellers from areas w/ recent local transmission of SARSA - education (whom to
contact, what to do if, fever develops within potential incubation period)
• Health departments
○ Assign staff (contact tracing and daily review of contacts in the community)
• Recovered patients
○ Quarantined at home for at least 14 days after discharged from hospital

SARS - Clinical Findings

• Incubation period
○ 6 days
• Fever
• Chills
• Muscle aches
• Dry cough
• Shortness of breath/difficulty in breathing

Adenovirus
• Infections of respiratory tract, eye and intestines
• Droplet or contact (modes of transmission)
○ Virus enters via mucosal cells of eye, nose or mouth
• Incubation period
○ 5-10 days
• Symptoms
○ Systemic - fever
○ Respiratory - nasopharyngitis, occasionally pneumonia (esp. children)
○ Occular - keratoconjuctivitis, conjuctivitis
○ GI - VD

Clinical Syndromes
• Wide variety of clinical syndromes, the majority of which concerns the repiratory_

Virology Page 199

Influenza
Thursday, 30 March 2017 11:19 AM

Introduction
• Influenza is caused by influenza virus
• Seasonal - cold season/winter
• Transmission higher in low temperature & low humidity
• 'Flu' - loosely used to describe a myriad of infections that cause respiratory or systemic
symptoms

Taxonomy
• ICTV
○ Orthomyxoviridae
 Influenzavirus A
□ Influenza A virus
 Influenzavirus B
□ Influenza B virus
 Influenzavirus C
□ Influenza C virus
• Baltimore Classification
○ (-)ssRNA
 Are accompanied by their own RNA polymerase because the host RNA
polymerase cannot translate negative sense RNA strands
 Replicates in the nucleus unlike other RNA viruses
• Morphology
○ Enveloped
○ Has 8 segments
 Important in mutation of the viruses (antigenic shift and drift)

Influenza Types
• Type A
○ Causes pandemics, most severe form
 Associated w/ both antigenic shift and drift
○ Also infects birds, pigs, horses, dogs
• Type B
○ Causes epidemics
 Associated with just antigenic drifts
○ Infects humans alone
○ Sporadic outbreaks esp. in institutionalised communities
• Type C
○ Causes mild disease
○ Infects humans and other animals, least common

Classification
• Examples
○ A/California/7/2009 (H1N1) virus
○ B/Brisbane/60/2008 virus
• Influenza type
• Place of isolate
• Number of isolate
• Year isolated
• For type A, the H and N subtypes are included

Virology Page 200

Structure
• Segmented virus
○ The segments are wrapped in protective protein
• The matrix
○ M2 is only found in INFLUENZA A
 Is point of t_
○ Help the virus to uncoat
 It is a proton channel that acts to provide a proper pH for viral uncoating
• Envelope
○ Haemaglutinin (HA) protein - Glycoprotein found on the surface of the virus
 For attachment
 Binds sialic acid on surface of epithelial cells in the respiratory tract or RBCs
□ The Sialic acid residues can be found sticking out of the RBCs
 Determines cell tropism
 Causes RBC agglutination in vitro
□ Basis for haemadsorption test
 16 subtypes: H1 - H16
 H1, H2, H3, H5, H7 and H9 have been recorded in man
○ Neuraminidase (NA) protein
 Facilitates virus release
 Breaks down sialic acid around infected cells
 Sialic acid is on the cell surface and in mucus secretions
 9 subtypes: N1 - N9
 N1, N2, N9 have been recorded in man
• Pleomorphic shape

Antigenic Variation
• Virus changes its antigens thus evading the immune system
○ Antigenic drift
 Occurs in all influenza viruses
 Gradual accumulation of (point) mutations - in the haemagluttinin and
neuraminidase proteins
 May generate a new strain
 Acquired immunity is no longer effective
 Causes seasonal outbreaks and seasonal epidemics
○ Antigenic shift
 Mainly occurs in influenza A
 Due to re-assortment of RNA segments
 Occurs in a cell infected w/ different subtypes (mixing vessel)
 May lead to a new virus subtype
 No previous immunity to the new subtype
 Causes sporadic epidemics/pandemics

Pathogenesis
• The HA molecule binds to the sialic acid found on the surface of epithelial cells. The virus
is then endocytosed into the cell and w/ the help of the M2 protein, uncoats and proceeds
to replicate in the nucleus. Newly formed virions are bound to the host cell because of
binding of HA to sialic acid. NA is then used to cleave sialic acid and facilitate virion
release
• Transmission
○ Small particle aerosol (10µm) and formites
• Incubation period
○ 18-75 hours
• Replicates in epithelial cells in the respiratory tract
• Viremia is rare
• Cell death is from direct viral effect and interferon

Virology Page 201

 • Cell death is from direct viral effect and interferon
○ Later from T-cytotoxic cells
• Epithelial cell death reduces clearance of pathogens

Clinical Features
• High risk groups
○ The young
○ The elderly
○ The sickly
• 90% of death in elderly
• Main cause of death
○ Bacterial pneumonia
○ Heart failure
• Uncomplicated influenza
○ Non-specific symptoms
 Headache
 Myalgia
 Fever
○ Cough
○ Rhinorrhea
○ Rarely, GIT symptoms (VD)
• Pulmonary Complications
○ Croup - laryngotracheobronchitis
○ Viral pneumonia
○ Secondary bacterial infection, usually due to:
 Haemophilus influenza
 Strep pneumoniae
 Staph aureus

Diagnosis
• Specimen
○ Throat/nasal swabs
○ Nasal noodles
• Rapid tests:
○ Antigen detection
○ RNA detection
• Serologic Tests
○ E.g. Direct Immunofluorescent tests
• PCR tests
○ Usually part of a panel for several resp. viruses
• Viral culture

Treatment
• Supportive
○ ABCs
○ Antipyretics/analgesics
• Chemotherapy
○ Amantidine/Rimantidine
○ Zanamavir/Oseltamivir/Peramivir
○ Agents that target RNA pol e.g. Ribavirin
• Antibiotics
○ Useful if there is secondary bacterial infection

Uncoating inhibitors
• Amantadine/Rimantadine

Virology Page 202

 • Amantadine/Rimantadine
○ Prevent uncoating
○ Block M2 ion channels - viral protein
○ Prevent acidification of the endosome
○ Active on Influenza A only
 M2 protein is only found in Influenza A
Viral Release Inhibitors
• Zanamivir, Oseltamivir, peramivir
• Effective for A, B and ?C
• Block viral neuroaminidase
○ Prevents viral spread to uninfected cells

Prevention
• Vaccines
○ Inactivated
 Trivalent inactivated Vaccine
 A quadrivalent inactivated vaccine
 Monovalent vaccine
○ Live attenuated influenza virus given as intranasal spray - quadrivalent
• Chemoprophylaxis - expensive

Control of pandemic influenza

• Surveillance
• Screening
• Information sharing
• Health education
• Health worker capacity
• Drugs - prophylaxis
• Vaccination - annual
• Culling animals - chicken/ swine
• Limiting travel
• Isolation of infected persons

Seasonal vs pandemic influenza

Pattern - predictable, cold season - Rare, a few in a century

Immunity - some immunity from previous exposure - no previous exposure, little ifr no immunity
Affected persons - Healthy adult usually spared - Everyone is at risk
Health systems - Able to meet demand - overwhelmed
Vaccines and drugs - Usually adequate - vaccine unavailable, drugs in adequate
Impact - Manageable - May severely affect economy

Virology Page 203

Prions
Tuesday, 4 April 2017 3:10 PM

Prionoses (transmissible Spongiform Encephalopathies)

Human Prionoses
• Creutzfeld-Jacob Disease (CJD)
• Fatal Familial Insomnia (FFI)
• Gerstmann-Straussler-Scheinker Syndrome (GSS)
• Kuru
• Variant CJD (vCD)

Animal Prionoses
• Bovine Spongiform Encephalopathies (BSE) - cows
• Scrapie - sheep
• Chronic Wasting Disease (CWD) - Mules, deer

Classification of Prionoses
• Familial/Hereditary
• Sporadic
• Infectious

Definition
• Small PROteinaceous INfectious particles that are resistant to inactivation by procedures
that usually modify/denature proteins
• They are infectious agents that are entirely made up of proteins. They:
○ Are aggregates of proteins that are normally found in the body but are misfolded
○ Transmissible
○ Generally cannot be broken down or denatured by typical methods

Characteristics
• Made of protein
• Resistant to inactivation
• Reproduce by converting normal protein to abnormal protein
• Do not induce the immune system
• Cause vacuolation of neurons >>> spongiform appearance

Cellular v. Scrapie Protein

Cellular (PrPc) Characteristics Scrapie (PrPsc)
Predominantly alpha-helical Structure Predominantly beta-sheets
Soluble Solubility Insoluble
Non-infectious Infectivity Infectious
Susceptible Resistance to Proteinase K Resistant

Transmission
• Sporadic
• Iatrogenic
○ Blood transfusion
• Hereditary
• Ingestion
○ Beef
○ Cannibalism

Virology Page 204

 ○ Cannibalism

Pathogenesis
Key feature: Accumulation of abnormal prion PrPsc
• Normal cellular protein (PrPc) found on the surface of brain cells and other cells
• Infection or mutation of PRNP gene on the c.20 can cause PrPc to misfold into PrPsc
○ Alpha helices are converted into ß-pleated sheets (which is predominant in PrPsc)
• PrPsc dissociates from the cell membrane
• PrPsc causes more PrPc to misfold and dissociate from the cell surface
• Accumulation of PrPsc forms proteinaceous plaques between the brain cells
○ ß-pleated sheets have a high propensity for forming bonds w/ other ß-pleated sheets
○ This means that the B-sheets from one protein will bond to the ß-sheets of another
protein and so one, finally causing formation of a plaque
• Aggregated PrPsc is then internalised into cells>> spongiform appearance
○ Because the cells are unable to denature these misfolded proteins, they die. The
sponge-like appearance is a result of dead neural tissue in the brain. This
degenerates brain function

Clinical Features
• Long incubation period, slow progressive and fatal disease
• Cognitive impairment
• Ataxia
• Myoclonic jerks
• Mutism
• FFI: Insomnia, dysautonomia, motor paralysis

Diagnosis
• Clinical presentation
• Exclude differentials
• CSF analysis (for 14-3-3 protein and/or Tau protein)
• EEG
• MRI
• Brain biopsy (post-mortem)

Treatment
• No treatment
• Prion diseases are invariable fatal (within months to years)
• Supportive treatment (palliative care)

Prevention
• Sterilisation of surgical equipment
○ Sodium hypochlorite and autoclaving (134°C for 1 hour)
• Screening
• Screening of blood and organs
• Ban on meat and bone meal in animal feed
○ Do not feed cows to cows

Virology Page 205

Antiviral Agents
Wednesday, 5 April 2017 9:17 AM

Viral Characteristics
• Intracellular life
• No organelles
○ Viruses share most of the host cell machinery (makes it harder to kill the virus)
• Require specific receptors to attach (viral tropism)
• Exploit host enzymes for replication
• Virus-specific targets do exist
• High level of mutation

Target: Life Cycle Stages

• Attachment
○ Viral ligand and host cell receptor
• Fusion/uptake
○ Viral envelop and host cell membrane
• Uncoating
• Reverse transcription
○ By action of RNA-dependent DNA Polymerase to form DNA
• Integration
○ Integration of viral DNA into host DNA
• Transcription
○ Formation of mRNA from the "new" DNA
• Translation to protein
○ By ribosomes in the cytoplasm
• Protein modification
○ To functional proteins
• Viral Assembly
• Viral Release

Classification of AVA
• By mechanism of action
• By target condition
• By mode of administration
• By chemical composition

Classification by Target virus

• Anti-herpesvirus agents
○ Acyclovir
 Valacyclovir/Ganciclovir/Valganciclovir/Famciclovir
○ Foscarnet
○ Fomiversen
○ Docosanol
• Anti-influenza agents
○ Uncoating inhibitors
 Amantadine
 Rimantadine
○ Viral Release Inhibitors
 Oseltamivir
 Zanamivir
 Peramivir

Virology Page 206

  Peramivir
○ RNA Polymerase Inhibitor
 Ribavarin
 Favipravir
• Anti-hepatitis B virus Agents
○ Immunomodulant
 Interferon (IFN)
○ RNA Polymerase Inhibitors
 Entecavir
 Tenofovir
 Lamivudine
 Telbivudine
 Adefovir
• Anti-Hepatitis C virus agents
○ Immunomodulant
 Interferon
○ RNA Polymerase Inhibitors
 Ribavarin
 Sofosbuvir
 Dasaburir
○ Protease
 Simeprevir
 Paritaprevir
 Grazoprevir
 Ritonavir* - is a booster agent
○ NS5A
 Daclatasvir
 Ledipasvir
 Velpatasvir
 Ombitasvir
 Elbasvir
• ARVs
○ Attachment blocker
 Maraviroc
○ Fusion blocker
 Enfurvirtide
○ Reverse transcriptase inhibitors
 Nucleoside analogues
□ Zidovudine
 Non-nucleoside analogues
□ Efavirenz
□ Nevirapine
□ Etravirine
○ Integration inhibitors
 Raltegravir
 Dolutegravir
 Elvitegravir
○ Protease Inhibitors
 Ritonavir
 Lopinavir
• Nucleic Acid Synthesis Inhibitors
○ DNA Synthesis Inhibitors
 DNA polymerase inhibitors (DdDp)
□ Incorporated into DNA chain by DNA polymerase causing termination
 Nucleoside analogues
◊ Acyclovir
◊ Idoxuridine

Virology Page 207

 ◊ Idoxuridine
 Nucleotide analogues
◊ Cidofovir
 Pyrophosphate analogues
◊ Foscarnet
◊ Phosphonoacetic acid
 Reverse transcriptase
○ DNA integration inhibitors
○ RNA Synthesis inhibitors

Acyclovir
• Acyclic nucleoside analogue
• Active against HSV-1, HSV-2 and VZV
• A pro-drug, undergoes 3 phosphorylation steps to its active form
○ Initial phosphorylation by viral thymidine kinase
○ Subsequent di- and tri- phosphorylation by host cell enzymes
• MoA
○ Inhibition of DNA Pol
 Acyclovir triphosphate competes w/ endogenous nucleotides for DNA Pol
 Incorporated into the growing DNA chain (viral genome)
 Causes chain termination once incorporated
• MoR
○ Reduced production of Viral Thymidine Kinase (TK)
○ Altered TK hence reduced affinity for Acyclovir
○ Altered DNA Pol

Protein Synthesis Inhibitors

Formivirsen
• An anti-sense oligonucleotide
• Binds the complementary CMV mRNA
• Prevents translation into proteins
• Stabilised to prevent degradation by nuclease
• Used to treat CMV retinitis - intravitreal injection

Interferons
• Cytokines w/ antiviral, immunomodulatory and anti-proliferative activity
• Synthesised by the body in response to certain stimuli
• IFNs induce synthesis of proteins that prevent viral entry, replication or maturation
• Numerous side effects
• Therapeutic use
○ HBV infection
○ HCV infection

Causes of Treatment Failure

• Poor adherence
• Poor absorption
• Drug interactions
• Drug resistance

Management of Treatment Failure

• Optimise adherence
○ Counselling
○ Close observation
○ Deal w/ side effects
• Test for drug resistance

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 •
○ Genotypic testing
○ Phenotypic testing
• Change to second-line therapy if indicated

Choosing an appropriate regimen

• Virologic efficacy
• Pill burden
• Toxicity
• Drug interactions
• Drug resistance testing conditions
• Co-morbidities

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Viral Childhood Fevers
Tuesday, 11 April 2017 2:15 PM

• Include
○ Measles
○ Mumps
○ Rubella
○ Varicella

Paramyxoviridae
• The family is subdivided into two subfamilies containing a total of four genera
• Subfamily
○ Paramxoviridae
 Genus
□ Paramyxovirus - influenza types 1 and 3
□ Rubellavirus - Influenza types 2, 4a, 4b and Mumps
□ Morbillivirus - Measles
○ Pneumoviridae
 Genus
□ Pneumovirus - Human Respiratory Syncytial Virus
• Though few in number, almost all the human paramyxoviruses are important causes of
respiratory disease in children

Properties of Paramyxoviridae
• Pleomorphic virion (150-300nm)
• Enveloped w/ two GP (F) fusion protein and attachment protein (MN) (Haemaglutinin-
Neuraminidase)
• Linear (-)ssRNA genome
• Cytoplasmic replication w/ budding_
• Paramyxoviruses multiply entirely within the cytosol. The infecting virion attaches to
sialoglycoprotein or glycolipid receptor via H or HN
• The (F) protein then mediates fusion of the viral envelope w/ plasma membrane
• Transmission
○ Respiratory droplets
• Virulence factors
○ Haemagluttinin (HA)
 Causes agglutination of RBCs
 Absent in RSV
○ Neuraminidase (NA)
 Absent in measles and RSV
○ Fusion proteins
 Formation of syncytia - multinucleated giant cells

Measles
• Infection occurs via the respiratory tract
• Virions enter and multiply within the respiratory mucosa, after which they are transported
to the regional lymph nodes
• Within the nodes, second replication takes place leading to systemic spread. Virions are
seeded to all he epithelial surfaces of the body - oropharynx, conjunctiva, skin, bladder,
respiratory tract and alimentary canal
• Because the epithelial of the conjunctiva and respiratory tract are the only one or two cells
thin, they undergo necrosis, 9-10 days following infection
• There is onset of illness w/ a cough, running nose and inflamed conjunctivae

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 •

• It seems likely that immune responses contribute to the respiratory damage, malaise and
fever
• The skin rash and Koplik's spots are due to cell-mediated response (Type IV
hypersensitivity)
• Three syndromes affecting the CNS have been recognised
a. Acute post-infections measles encephalitis
 Rare in children under two years
 Occurs in about 1 in 1000 case fertility rate of 15%
 It is the principle reason for vaccination
 Occurs during the first week after the onset of the rash. There is little or no
production of the virus in the brain
 The pathogenesis appears to involve autoimmune demyelination
b. Subacute measles encephalitis
 Occurs only in the immunocompromised children, usually within 6 months of
the rash
 It may be rapidly progressive and is attributable to failure to eliminate virus-
infected cells due to CTL
c. Subacute sclerosing panencephalitis (SSPE)
 Occurs years (5 or 6 or more) after acute disease.
 It is always fatal
 Characterised by very slow replication and spread of measles virus in the CNS
 Patients w/ SSPE reveal very little measles virus in the brain but very high titres
of neutralising antibodies in the CSF

Lab diagnosis
• The clinical diagnosis of measles is so straight forward that the laboratory is rarely called
in for help

Epidemiology and Control

• A live attenuated measles vaccine (Schwarz) vaccine
• Given at 6-15 months

Treatment
• No antiviral agent is effective against measles

Mumps
• Infection by droplets
• Painful oedematous enlargement of parotid and other salivary glands
○ The virus replicates in the parotid glands - parotitis

Complications
• Orchitis
○ Usually occurs unilaterally but can occur bilaterally and can result to testicular
atrophy and impaired fertility and sterility is rare but can occur in men w/ bilateral
orchitis
• Mumps encephalitis/meningitis

Control
• Long-lasting immunity following recovery
• Live attenuated vaccine

Parainfluenza Viruses
• Commonly respiratory pathogens (URTI) but cause more serious condition in young
children known as Croup (laryngotracheobrinchitis and inspiratory stridor) and occasionally
Pneumonia
• Virulence factors

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 • Virulence factors
○ HA
○ NA
○ F
• Diagnosis
○ Imaging
 X-ray
□ Steeple sign - Narrowing of the subglottic region

Respiratory Syncytial Virus

• Virology of Respiratory Syncytial Virus
○ ICTV Classification
 Family - Paramyxoviridae
 Subfamily - Pneumovirinae
 Genus - Pneumovirus
 Species - RSV
○ Baltimore classification
 (-)ssRNA - Class V
○ Morphology
 Helical nucleopsid
 Enveloped
 Non-segmented linear virus
• RSV is the most important respiratory pathogen of childhood
○ Seen in children <6 months of age
• Responsible for about 5.5 of all the case of bronchiolitis and pneumonia during the first
few months of life
• Infects cells by attaching to the respiratory endothelium using the G protein
• Virulence factors
○ Fusion protein
• Can cause
○ Bronchiolitis
○ Pneumonia
○ Rhinitis
○ Pharyngitis
○ Most common cause of pneumonia and bronchiolitis in infants
• Treatment
○ Ribavirin - in adults
 Not recommended in children and pregnant women
○ Palivizumab - Monoclonal antibody against the fusion protein
 Used to prevent RSV infection in infants especially children born prematurely

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Viruses and Cancer
Tuesday, 11 April 2017 3:28 PM

• Study of oncogenesis - cell transformation

• Oncogenesis has been facilitated by:
○ Oncogenes (originally found in retroviruses) - v-onc genes
• For each v-onc, there is a corresponding normal cellular gene (c-onc genes) (proto-ogenes)

Cell Transformation
• Ex-vivo cell transformation (equivalent tumour formation)
• Transformation by DNA viruses - non-productive
• Transformation by RNA is often productive
• Viral (or proviral) DNA is integrated into the cell DNA (except for papillomavirus and herpes) Cancer is monoclonal
• Viruses associated w/ formation of tumours in humans
○ HHV4
○ HBV
○ HCV
○ HHV-8
○ HPVs
○ HTLV
 II
 III
• HHV-4 - dual cell tropism - B lymphocytes and epithelial cells
• B cell infection - polyclonal B-Cell activation and a benign proliferation of B cells

Tumours Associated w/ EBV

• Burkitt's Lymphoma
• Nasopharyngeal Carcinoma (NPC)
• B-cell Lymphoma
• Mechanism of HHV-4 cellular transformation
○ Polyclonal activation of B cells where chromosomal translocation occurs in some clones. HBV
and hepatocellular carcinoma (HCC)
• Outcomes of HBV Infection

Viral Oncogenesis
• An important element in our present understanding of oncogenesis has come from the discovery of
oncogenes, originally found in_
• _
• The proteins they encode can be assigned to four major classes
○ Growth factors
○ Growth factor receptors
○ Intracellular signal transducers
○ Nuclear transcription factors
• A completely different category of cellular genes called tumour suppressor genes were discovered,
which also play an essential regulator role in normal cells
• Their protein products are involved in negative regulation of growth

Cell Transformation
• The capacity to study oncogenesis at a molecular level was greatly facilitated when it became
possible to induce the essential genetic changes in cultured cells - (cell transformation) which is the
in vitro equivalent of tumour formation
• Transformation by DNA viruses is usually non-productive transformation by retroviruses is often
productive
• Viral (or proviral) DNA in transformed cells is integrated into the host cell DNA, except in the case of
papillomavirus and herpesvirus DNAs, which usually remain episomal
• Malignant tumours and transformed cells express distinctive antigens called tumour associated
antigens
• Some tumour associated antigens are located in the plasma membrane, where they constitute
potential targets for immunologic assaults and are at times referred to as tumour specific
transplantation antigens

Tumour Induction by Retroviruses

• In humans, only one retrovirus, HTLV-1, has been unequivocally incriminated in oncogenesis, as the
cause of a particular form of leukaemia

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 cause of a particular form of leukaemia
• Oncogenic retroviruses are subdivided into two
○ Replication competent or replication defective
○ Endogenous or exogenous

Replication-Competent/Replication-Defective
• The genome of a typical replication-competent retrovirus consists of (LTR-5'-GAG-POL-ENV-3'LTR)
• A second kind of rapidly oncogenic exogenous retroviruses carries a viral oncogene (V-onc) -
responsible for rapid. The rapid malignant change of the infective cell
• Because the oncogene is usually incorporated into the viral RNA in place of one or more normal vital
gene, such viruses are usually defective
○ 5'cap LTR gag onc env LTR poly (A)3
○ 5'Cap LTR - gag pol env src LTR poly (A)3

Endogenous and Exogenous Retroviruses

• A complete DNA copy of the genome (provirus) or one, or sometimes more than one, of the many
retroviruses may be transmitted in the germ line from parent to offspring and may thus be
perpetuated in the DNA of every cell
• Proviral genomes are under the control of cellular regulatory genes and are normally totally silent in
the normal animals - expression of proviruses can be induced by various factors:-
○ Indication
○ Exposure to mutagenic of carcinogenic chemicals or immunologic stimuli

Mechanisms of Tumour Production by Retroviruses

• Retroviruses produce tumours in one of the following ways
○ Transducing retroviruses introduce a V-onc gene, which is under control of viral LTR
○ CIS-activating retroviruses contains a gene, transform cells by integrating close to the C-onc
gene and thus usurping normal cellular regulation of this gene
○ Trans-activating retroviruses contain a gene that codes for a regulatory protein which may
either increase transcription from normal LTR or interfere w/ the transcriptional control of
specific cellular genes

Tumour Induction by DNA viruses

• Retroviruses are the most important oncogenic viruses in animals
• Certain DNA viruses are more important causes of cancers in humans
• _
• The molecular basis of oncogenesis by DNA viruses is best understood for:
○ Polyomaviruses
○ Papillomaviruses
○ Adenoviruses
• All contain genes that behave as oncogenes
• These oncogenes appear to act by mechanisms similar to those described for retroviruses
• Primarily in the nucleus - where they alter patterns of gene expression and regulation of cell growth
• The relevant genes encode early proteins having dual role in viral replication and cell transformation

Papillomavirus and Cervical Carcinoma

• Evidence is accumulating that HPV may play a role in causing cervical carcinoma as well as penile
cancer. What evidence?

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Lab Diagnosis of Viral Infection
Wednesday, 12 April 2017 9:20 AM

Communication (Physician-Lab)
• Physician/Clinician
○ Clinical diagnosis (need further confirmation - lab diagnosis)
○ Specific lab requests (based on tentative diagnosis - infection type/infectious agent
suspected)
○ Proper labelling of specimen _

Viral Diagnosis in the Clinical Lab

• >70% of all infectious disease cases seen by a physician are due to viral infections
• For lab diagnosis
○ Quality of patient specimens important
 Collection
 Appropriate tubes/containers
 Transportation
 Storage
 Appropriate test and analysis

Procedures for Lab Viral Diagnosis

• Virus infected patients
○ Collect specimens
 Check infected cells under light microscope
 Virus culture and isolation
□ Identify viral propagation
 CPE
 Hemadsorption
 Neutralisation test
 Plaque assay
 Detection of viral antigen or genome
□ Antigen
 IFA
 EIA/ELISA
 Western blot
□ Genome
 PCR
 Southern blot
 Northern blot
 Serology test
□ Neutralisation test
□ HAI test
□ EIA/ELISA
□ Western Blot
 Observe virus particle under EM

Three General Approaches for Lab Diagnosis of Viral Infections

• Virus isolation (indirect Examinations)
○ CPE and other characters
○ Animal systems (Eggs, mice, etc.)
• Direct detection
○ Microscopy or staining
○ Detection of nucleic acid_
• _

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 • _

Virus Cultivation Systems

_

Virus Isolation - Cell Cultures

• Three types
○ Primary cells e.g. Monkey Kidney
○ Semi-continuous cells e.g. human embryonic kidney and skin fibroblasts
○ Continuous cells e.g. HeLa, Vero

Cultured Cells
• Primary
○ Heterogenous - many cell types (several viruses can grow)
○ Closest to animal
○ Technical hassle
○ Expensive
• Diploid cell strain
○ Relatively homogenous - fewer cell types
○ Further from animal
○ Technically less hassle
• Continuous cell line
○ Immortal
○ Most homogenous
○ Genetically weird - furthest from animal
○ Hassle free
○ Suspension of monolayer
○ Limited number of viruses supported

Laminar Flow Hoods

• Virologist's facility
• Laminar vertical flow hoods
○ Contains HEPA filter
○ Removes particle_

Cytopathic Effects (CPE)

• Visible results of viral infection
• Cell death by
○ Multiplying viruses
○ Inhibition of DNA, RNA or protein synthesis
○ Effects on permeability of membrane
• Observed w/ inverted light microscopes
○ Rounding/detachment from plastic flask
○ Syncytia/Fusion of cells
○ Shrinkage
○ Increased refractability
○ Aggregation
○ Loss of adherence
○ Cell lysis/death
• Common observations of CPEs
○ Inclusion body formation (intracellular virus parts - replication or assembly)

Problems w/ Cell Culture

• Long period (up to 4 weeks) required for result
• Poor sensitivity (most times)

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 • Poor sensitivity (most times)
• Susceptible to bacterial contamination
• Susceptible to toxic substances which may be present in the specimen
• Not all viruses grow

Virus Isolation - animal inoculation

• Choice of animal and inoculation pathways
• Mouse, rats, rabbit, monkey etc.

Embryonated Eggs
• Generates large quantity of virus
• Used for vaccine production
• Sites for viral inoculation on th eegg
○ Chorioallantoic
 Herpes Simplec virus
 Poxvirus
 Rous Sarcoma virus
○ Amniotic inoculation
 Influenza virus
 Mumps virus
○ Yolk sac inoculation
 Herpes Simplex Virus
○ Allantoic inoculation
 Influenza
 Mums( )
 Newcastle disease
 Avian adenovirus
• Signs of viral growth include
○ Death of the embryo
○ Defects in embryonic development
○ Localised areas of damage in the membranes
 Result
□ Discrete, opaque spots called pocks
○ Embryonic fluid and tissue - examine w/ EM

Detection of Viral Proteins

• Haemagglutinin (HA) and HA inhibition (HAI) assays
• Plaque assays
• ELISA
• Western Blots

Viral Haemagglutinin
• Some viruses and microbes contain proteins which bind to erythrocytes (RBCs) causing
them to clump together
○ Paramyxovirus e.g. Mumps, Measles, Parainfluenza
○ Influenza virus
○ Adenovirus etc.

Readings : The Result

• Titre
○ The maximum dilution that gives visible agglutination
• The end point
○ The well w/ the lowest concentration of the virus where there is haemagglutination

Quantitative Assays
• Plaque Assays

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 • Plaque Assays
○ Lytic viruses only
○ Steps
 Serial dilution of virion-containing solution
 Create tissue culture plates
 Spread diluted virus
 Overlay w/ agar - prevents diffusion
 Count number of plaques
 Each plaque represents 1PFU (plaque forming unit)

Direct Detection
• Electron Microscopy - examine specimen for virus
○ Morphology of virus particles
○ Immune electron microscopy
• Light microscopy - labelled antibody
○ Histological appearance
○ Inclusion bodies
• Antigen detection
○ Immunofluorescence (florescent tag bound to Fc portion of antibody)
○ , ELISA etc.
• Viral Genome Detection -
○ Hybridisation w/ specific nucleic acid probes
○ PCR

IFA
•Use monoclonal antibodies (MoAbs) labelled w/ a fluorescent dye
•MoAbs bind to specific epitope on viral protein
•Visualise infected cells using fluorescent microscopy
•Only virus-infected cells will fluoresce
•Direct method
○ Fluorescein tagged antibody → attached fluorescein tagged antibody visualised by
UV microscope
• Indirect method
○ First step
 Untagged antibody → antibody attached to antigenic determinant
○ Second step
 Fluorescein tagged anti-immunoglobulin + antigen attached to antigenic
determinant → attached fluorescein tagged anti-immunoglobulin visualised by
UV microscopy
• Advantages
○ Results available quickly, usually within a few hours
• Disadvantages
○ Low sensitivity (compared to cell culture)
○ Poor specificity
○ Requires good specimens
○ Tedious/time consuming
○ Expensive (lab time and equipment)

Methods for Rapid Diagnosis of Viral Genome

• PCR
• Molecular hybridisation
○ Southern blot
○ Northern blot
○ Dot blot
○ In situ hybridisation

Virology Page 218

Immunity to Viral Vaccines
Thursday, 13 April 2017 11:41 AM

Host outcome -Virus Infection

• Recovery
• Persistence
• Rapid death

Immune Responses to Viral Infections

• Innate immunity
• Adaptive T-cell immunity
• Adaptive B-cell immunity

Vaccination in Viral Infection

Vaccines, Vaccination and Immunisation Basics
• A vaccine is a product that produces immunity from a disease and can be administered through needle injections, by mouth or by
aerosol
• Vaccination
○ Injection of a killed or weakened organism that produces immunity in the body against that organism
• Immunisation
○ Process by which a person or animal becomes protected from a disease.
○ Vaccines cause immunisation and there also diseases that cause immunisation after an individual recovers from the
disease

Vaccination or Immunisation?
• It's artificial method to make someone immune to a disease
○ Active immunisation - administration of vaccine so that the patient actively mounts a protective immune response
○ Passive immunisation - Individual acquires immunity through the transfer of antibodies formed by and immune individual or
animal

Active immunisation
• Develops after the immune system has been exposed to an antigen or vaccine
• Immunologic memory develops and protects the individual from re-infection
• Long lasting

Passive Immunisation
• Given as protective antibodies to an individual who has not been exposed to a pathogen
• They are short lived
• Do not induce memory
• Examples include rabies and Tetanus

Goal of Vaccine
• Stimulate both cell mediated and antibody mediated immunity, that will protect the vaccinated person against future exposure to
pathogen
• Want the vaccine to have
○ Maximum realism
○ Minimum danger

Attributes of a Good Vaccine

• Ability to elicit the appropriate immune response
○ For the particular pathogen
• Long term protection
○ Ideally life-long
• Safety/revertion
○ Vaccine itself should not cause disease
• Stable
○ Retain immunogenicity, despite adverse storage conditions prior to administration
• Inexpensive

Vaccine Types
• Live whole virus vaccines
• Killed whole vaccine viruses

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 • Killed whole vaccine viruses
• Subunit vaccines
○ Purified or recombinant viral antigen
• Recombinant virus vaccine
• DNA vaccines
• Virus like particles

Live or Attenuated Whole Virus Vaccines

• Also called modified live vaccines
• Elicits both cellular and humoral immune responses
• Use pathogens that are living but have reduced virulence so they don’t cause disease
○ The technique used to make them avirulent (non-toxic) has made the human host an unfavourable environment for rapid
replication that can cause disease (revert back to virulent state)
• Approach for producing attenuated viruses
○ Repeated passage in cell culture
 They are plated in selective media where their food habits are altered that might affect their gene sequence
 They are then placed in different hosts (host changing). If they cause disease, they are re -plated and then inoculated
into another host. Before it is inoculated into a human host, it has to undergo trial phases to check for toxicity
○ Temperature-sensitive
○ Deletion mutants
• Live vaccines are very effective for three reasons
○ They replicate and thus deliver sustained doses of antigen
○ They replicate intracellularly so they deliver antigenic peptides to MHC-I and thus stimulate cytotoxic T-cells (CTLs)
○ They replicate at the anatomical site of infection e.g. live vaccines given by mouth, elicit IgA antibodies
• Characteristics of Live Attenuated Vaccines
○ Good immune response - because the cells are alive
 Both cell mediated immunity and antibody responses
 Immunity is long-lived
 Single dose
○ Safety
 Danger of reversion to virulence or
 Severe disease in immunocompromised
○ Stability
 Organisms in the vaccine must remain viable, in order to infect and replicate in the host
 Vaccine preparations are therefore very sensitive to adverse storage conditions
 Maintenance of the cold chain is very important
○ Expense
 Cheap to prepare

Killed Whole Virus Vaccines

• In inactivated vaccines, the disease-causing microbe is killed w/ chemicals, heat or radiation
○ radiation is introduced and penetrates the cell and changes the gene sequences of the cell so it is no longer viable
• Characteristics
○ Immune Response
 Poor; only antibody - no cell mediated immune response
 Response is short-lived and multiple doses are needed
○ Safety
 Inactivated, therefore, cannot replicate in the host and cause disease
 Local reactions at the site of injection may occur
○ Stability
 Efficacy of the vaccine does not rely on the viability of the organisms
 These vaccines tend to be able to withstand more adverse storage conditions
○ Expensive
 To prepare
• Advantages
○ There is no reversion to virulence by the vaccine virus
○ Vaccines can be made when no acceptable attenuated virus is available
• Disadvantages
○ Extreme care required to ensure no residual live virus is present
○ Immunity conferred is brief and needs to be boosted - logistic problem of repeated doses and hypersensitivity reaction may
occur from repeated exposure to antigen
○ Cell mediated response is poor
○ Parenteral administration of killed virus, even when stimulates systemic antibodies (IgM and IgG) to satisfactory levels, has
sometimes limited protection due to local resistance (IgA) is no induced properly at the natural portal of entry or primary site
of multiplication of the wild virus infection

Subunit or Peptide Vaccines

• Subunit vaccines

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 • Subunit vaccines
○ Uses important parts of the virus to stimulate the immune system e.g. in Diphtheria
 Diphtheria has 2 subunits A and B where A is responsible for bacterial attachment and B causes disease. The B
portion is destroyed and the A portion is used for the vaccine
○ Conjugated Vaccines
 The antigen is attached to a sugar/a molecule that can be taken up by the body
• Characteristics
○ The chances of an adverse reaction to the vaccine are lower
○ Downside:
 Identifying the precise antigens which best stimulate the immune system is difficult and time consuming
○ It may be possible to manufacture the antigen molecules using recombinant DNA technology

DNA Vaccines
Virus Like Particle Vaccines
• Consists of viral proteins derived from the structural proteins of a virus
• These proteins can self-assemble into particles that resemble the virus from which they were derived but lack viral nucleic acid
• Because of their multivalent structure, virus like particles are typically more immunogenic than subunit vaccines
• Example - human papillomavirus
Recombinant DNA Technology (RDT) Vaccine

Vaccine MoA Immune Stability Expense Safety Duration

Response of
Protectio
n
Live A live avirulent cell Both cellular Not very stable Cheap to make There is a danger of Long-lived
attenuat (bacteria/virus) is inoculated and humoral as the cell has although the process reversion to virulent state
ed into the host and an Very good to remain of attenuation can be or the vaccine may be
immune response is response as viable time consuming toxic to the
mounted against it the cell is Maintenance of immunocompromised
alive a cold chain is
necessary
Killed
Steps in Vaccine Development
• Prepare protocol(s) for human studies
• Apply to ethics approval
• Phase I
○ Human trials
 Safety and immunogenicity
 Dose response
• Phase II trials
○ Safety
○ Immunogenicity
• Phase III trials
○ Efficacy

Virology Page 221

Introduction to Medical Bacteriology
Friday, 11 December 2015 3:29 PM

Microbiology - refers to the study of microbes and microorganisms that infect humans

Cellular structures of Micro-organisms

• Broadly classified as
○ Eukaryotic organisms
 Characterised by well developed cellular structures with a nucleus, nucleolus and nuclear
membrane
○ Prokaryotic organisms
 Relatively undeveloped or primitive cells
 Unicellular approximately one micrometre Kosh postulates
 No nuclear membrane or proper nucleus For a particular organism to be causative agent
 Includes bacteria of a disease:
• The organism must be demonstrated from
the lesion in every case of the disease
Medical bacteriology • The organism must be isolated and
• Developed over centuries as a result of research by several scientists from different parts of the propagated in pure culture in vitro
world • Inoculation of a live culture of the
• Importance of Medical microbiology organism by a suitable route into a
○ Provide knowledge on micro-organisms of suitable animal should reproduce the
disease
 Morphology, physiology, biochemistry, reproduction, genetics • The organism must be re-isolated
 Interaction with humans in relation to disease production Organisms for which Kosh's postulates were
 Laboratory test for detection of the organisms and their effect on humans fulfilled were referred to as pathogens to
 Management of infections and diseases associated with microorganisms distinguish them from non-pathogens

Methods of study in medical bacteriology

1. Microscopy
a. Involves the use of various types of microscopes for visualisation of microbes
b. Light microscope
i. General observation of morphology of stained bacteria
ii. Examination of unstained organisms for various characteristics such as motility
iii. Examination of tissue and other specimens for microorganisms and other abnormalities
c. Special microscopes
i. For visualisation of finer structures or very slender bacteria.
ii. Include
1) Phase contrast microscope
2) Dark field microscope
a) 1 and 2 are fitted with special condensers to enable visualisation of refractive
and slender unstained organisms
3) Fluorescent microscope
a) Utilises fluorescent light and fluorescent staining methods of micro-organisms
4) Electron microscope
a) For higher resolutions of finer structures of micro-organisms
2. Staining of bacteria and other materials
a. Unstained bacteria are refractive or translucent
b. Staining
i. Imparts colour and makes bacteria more visible
ii. Enables differentiation of special features of each bacterial cells
c. Staining techniques
i. Simple staining
1) Utilises one type of stain
ii. Negative staining
1) Stains in the background and enables certain features to be visualised clearly
against stained background
iii. Differential staining
□ Utilises more than one reagent
□ Depends on the permeability of the cell wall and the entry of the stain into the cell
□ Involves more than one complex procedure and reagents and cell contents
□ Several methods based on the type of organism under study include
 Gram stain
 Ziehl-Neelsen staining
 Spore stain
 Various staining methods for various organelles such as
◊ Flagella
◊ DNA capsules
iv. Fluorescent staining or immunofluorescent staining
1) Used for examination of specimens by
a) Staining using a fluorescent material or stains bound chemically with specific
proteins including antibodies against antigens on a particular organisms
b) Examination by fluorescence microscope and UV light

Bacteriology Page 222

 b) Examination by fluorescence microscope and UV light
3. Culture growth or isolation of the bacteria
a. Involves growing the desired bacterium in artificial media
b. Bacteria differ in their physiological and nutritional requirements for growth
i. Suitable nutritional and other growth requirements must be provided for successful
isolation for each organism
1) Artificial medium
2) Atmospheric growth environment
3) Duration and temperature of incubation
c. Artificial culture medium
i. Semisolid or liquid material s prepared and dispensed into petri-dish or bottle
1) Basic contents are water, nutrients and agar
a) Agar is a substance extracted fro sea weed
2) Other substances depend on the intended use0
ii. Bacteria divide numerous times to form visible growth as
1) Turbidity in a liquid medium
2) Multiple heaps of cells in the surface of semi-solid referred to as colonies
3) Each colony
a) Arises from one bacterial cell
b) Has specific features for the type of bacterium on each medium
iii. Types of artificial culture medium for bacterial growth (classification according to uses)
1) Simple medium
a) Contain peptone
b) Support the growth of many types of bacteria
c) Include
i) Nutrient broth
ii) Nutrient agar
2) Enriched media
a) Contain nutrients suitable for isolation of fastidious bacteria which require
special nutrients
b) Include
i) Blood agar - contains 10% blood
ii) Chocolate blood agar or heated blood agar
3) Selective media
a) Contain chemical or antimicrobial agents which inhibit growth of other types of
bacteria and allow growth of the desired organism
b) Chemicals or antimicrobial agents make the media selective
c) Include
i) Thayer - martin medium,
 Contains antimicrobial agents
ii) Deoxycholate citrate agar
 Contains bile salts
4) Enrichment media
a) Liquid media or broths with selective properties
b) Favour multiplication of a particular organism as a step to its isolation
c) Include
i) Alkaline peptone water
ii) Selenite F broth
5) Transport media
a) Protect delicate pathogens in specimen when there is a delay from the time of
collection to the time the sample is delivered to the laboratory and processed
b) Include
i) Stuart's transport medium
ii) Cary-Blair transport medium
6) Indicator medium
a) Contain fermentable carbohydrate and indicators
b) Associated with colour change due to changes in pH cause by acid production
as a result of fermentation by bacteria
c) Include
i) MacConkey's medium
ii) CLED medium
d) Allow detection and partial identification of certain bacteria
iv. Atmospheric requirement for the growth of bacteria
1) Cultures may be incubated in
a) Air where oxygen is available
b) Environment where oxygen is completely eliminated
c) Additional 5 - 10% carbon dioxide
2) Temperature range for growth
a) Varies according to the organism
b) Ranges from max to min
c) Optimum growth occurs at a temperature close to human temperature

d. Identification of isolated organisms

i.

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 i. Determines the genus, species and possibly the strain
ii. In most laboratories identification is based on certain characteristics including
1) Staining properties and microscopic morphology
2) Cultural properties
3) Biochemical tests or reactions
a) Depend on enzymes and enzymatic activity
b) Detects a specific enzyme or activity of n enzyme
4) Susceptibility to various antimicrobial substances
5) _
e. Typing of bacteria
i. Subdivides species into serotypes or strains
ii. May provide information on source and patterns of spread
iii. Numerous methods or typing available include
1) Serotyping into serotypes
a) Based on antigenic structures of the cells
b) One serotype is a single bacterial strain define by antigenic properties
c) Serovar is used interchangeably
d) _
iv. Identification by DNA characteristics
1) Based on analysis of DNA composition
a) For each organism there is consistency in
i) Number of genes processed
ii) Nucleotide base composition
iii) Guanine plus cytosine content
b) DNA detection and amplification
v. Indirect methods of detection of bacterial infection and identification
1) Mainly by
a) Use of gene probes in detection and amplification of genetic material
b) Antigen-antibody reactions which involve
i) Tests for a specific antibody or serology
ii) Antigen detection
One. I and ii are performed by various immunological procedure including
→ Agglutination reactions
→ ELISA
→ IF staining and fluorescence staining
→ _

International Classification of bacteria

• Involves grouping organisms according to their similarities
• New species are defined and other species are re-classified as new characteristics are disclosed

Taxonomy]
• The branch of science that deals with classification of organisms concerning its principles,
procedures and rules
• Aims to achieve the
○ Identification
○ _
• _

Taxonomic groups and nomenclature

• Includes
○ King David Can Only Find (tribe) Green Socks

Binomial Nomenclature

Other methods of classification

• Based on important characteristics
• May be according to
○ Shape
 Spherical cells (cocci)
□ Method of cell division determines arrangement
□ Division in
 One plane produces either paired cells (diplococci) or chains of cocci
 Two planes produces tetrads
 Three planes - cocci arranged in eights
 Random planes results in clusters or grape arrangement
 Rectangular or rod shapes (bacilli)
□ Can be
 Typical bacilli
 Relatively small bacilli
 Coccobacilliary or short bacilli
 Filamentous, slender and elongated bacilli
 Comma shaped bacilli
 Spiral shape

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  Spiral shape
 Pleomorphic bacteria
□ Lack a distinct shape
□ Are soft
○ Gram's stain reaction or appearance
 Gram Positive
 Gram Negative
 Gram Variable
 Gram Indeterminate or not stainable
○ Shape and gram stain appearance combined
 Used frequently in medical bacteriology labs

She refused to give us the notes and she went super fast

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Bacterial Anatomy
Friday, 18 December 2015 2:36 PM

Capsule/Glycocalyx/Slime Layer
• Polysaccharide structures surrounding the outside of the cell envelope
• Glycocalyx - gelatinous
○ Helps in the formation of biofilms on inert surfaces such as catheters, teeth and heart
valves
• Capsules - glycocalyx firmly attached to the cell wall
• Slime layer - glycocalyx attached loosely to the cell wall

Significance of capsules
• Anti-phagocytic
• Growth in a biofilm prevents access of host cells or antibodies
• Prevents the cells from drying out
• Used as antigens in certain vaccines e.g. pneumococcal vaccine

Cell Wall
• Main component is peptidoglycan (murein)
• The thickness of peptidoglycan distinguishes gram positive from gram negative bacteria
• Overlapping N-acetyl glucosamine (NAG) and N-acetyl muramic acid (NAM)
• Present in almost all bacteria, except mycoplasma and ureaplasma

Cell wall of gram positive bacteria

• Thick peptidoglycan layer (50-90% of cell wall material)
• Composed of
○ Teichoic acids
 Mediate attachment to mucosal membranes
 Induce septic shock in gram positive bacteria
○ Lipoteichoic acids
 Anchor cell wall to cell membrane
□ For epithelial cell adhesion
○ Polysaccharides and proteins
 Protect peptidoglycan layer from actions of agents such as enzymes
 Promote colonisation by sticking the bacteria to the surface of the host cells

Cell wall of gram negative bacteria

• Thin peptidoglycan layer comprising of 5-10% of cell wall material
• Have an additional outer layer
• Have a peri-plasmic space
○ Contains digestive enzymes and other transport proteins
• Contains porin protein
○ specifically allow transport of solutes in and out of the cell
• Their cell walls contains three components
○ Lipoprotein anchors the outer membrane to peptidoglycan
○ Outer membrane protects the cell from proteolytic enzymes
○ Lipopolysaccharide (LPS)
 Present in almost all gram negative bacteria
 Major component is Lipid A - endotoxin (responsible for endotoxic activity such
as fever hypotension and septicaemia)

Acid fast bacteria

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Acid fast bacteria
• The cell envelopes of Mycobacteria are more complex than other bacteria
• Composed of Mycolic acid (thick waxy membranous layer outside the peptidoglycan layer)

Significance of Cell wall

• Maintains cell shape
• Protects the bacteria from osmotic lysis
• Determines reactivity to gram stain
• Site of action of certain antibiotics
• _Enhances pathogenicity

Plasma/cytoplasmic/cell membrane
• Separates cell wall from cytoplasm
• Acts as semipermeable membrane
• Composed of lipoproteins with small amounts of carbohydrates
• Generally do not contain sterols (except for Mycoplasma and ureaplasma)

Function
• Active transport of molecules into the cells
• Synthesis of precursors of the cell wall
• Secretes enzymes and toxins

Pili
• Hair like projections on the surface of the cell
• Shorter and straighter than flagella
• Composed of protein - pilin
• Mostly on gram negative bacteria
• Two types of pili
○ Fimbriae/common pili - cover the cell surface
 For attachment
○ Sex pili - longer than common pili
 Involved in conjugation
 Longer than fimbriae

Flagella
• Long, filamentous surface appendages
• For bacterial motility
• Composed of the protein 'flagellin'
• May serve as antigenic determinants (eg the H antigens of gram-negative enteric bacteria)

Ribosomes
• They are composed of RNA and proteins
• Site of protein synthesis
• Site of activity of antimicrobials that disrupt protein synthesis
• 70S in size with 50S and 30S sub units

Nucleoid
• Area of cytoplasm in which DNA is located
• Bacterial DNA consists of a single, circular double-stranded DNA
• Lacks nuclear membrane (called nucleoid)
• Contains genetic material that codes for all genetic information expressed by the cell

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Plasmids
• Extra chromosomal DNA molecules
• Easily passed from bacterium to bacterium through sex pili
• Free or integrated into the chromosome
• May encode genes of antibiotic resistance and pathogenesis factors (eg enzymes and
toxins)

Inclusion/nutrient granules
• Composed of volutin, lipid and polysaccharide
• Stain characteristically with certain dyes
• Example: volutin granules are seen in corynebacterium spp
• Function
○ Serve as storage area for nutrients and energy for cell metabolism

Mesosome
• Appear as convoluted indentations (invaginations) in the cytoplasmic membrane
• Functions
○ Are sites of respiratory enzyme activity
○ Coordinate nuclear and cytoplasmic division during binary fission

Spores
• Round, oval or elongated
• Formed inside the parent cell (endospores)
• Formed when conditions for vegetative growth are not favourable
• The exhibit no metabolic activity
• Resistant to heat, radiation and drying and can remain dormant for hundreds of years
• Formed by bacteria like Clostridia, Bacillus

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 Bacterial Physiology
Friday, 18 December 2015 3:33 PM

Study of how bacteria function including processes such as nutrition, growth, reproduction and locomotion

Growth and proliferation of bacteria

• Bacterial requirements for growth
○ Temperature
○ Oxygen
○ Nutrients
○ pH
○ Moisture

Temperature
• Temperature classification
○ Psychrophiles - low temperatures 10-20 degrees C
○ Mesophiles - 20-40 degrees C
○ Thermophiles - temperatures >40 degrees C

Oxygen utilisation
• Strict (obligate) aerobes - require oxygen for growth eg pseudomonas aeruginosa
• Strict (obligate) anaerobes - grow in the absence of oxygen eg Bacteroides fragilis
• Facultative anaerobes - do not require oxygen for growth but grow better in its presence eg
Staphylococcus species 5
• Capnophilic - thrive in the presence of high concentrations of CO2 or which require the presence of
CO2 to survive
• Microaerophilic - grow well in low concentrations of O2 and higher CO2 concentrations ex
Campylobacter
○ Easily cultivated in a candle jar
 Eg neisseria meningitidis

pH
• Neutrophiles (5 to 8)
• Acidophiles (below 5.5)
• Alkaliphiles (above 8.5)

Nutrient Requirements
Generation time
• The time required for a bacterium to give rise to 2 daughter cells under optimum conditions eg
○ Escherichia coli and other medically important bacteria - 20 minutes
○ Tubercle bacilli -20hrs
○ Leprae bacilli - 20 days

Bacterial growth phase*

• Lag phase - checking if the environment is fit for growth and reproduction
• The bacteria are adapting to the new environment
• No cell division
• Vigorous metabolic division
• Cells may increase in size during this time, but simply do not undergo binary fission
• Logarithmic/exponential phase - growth of population in favourable environment
• Cells start dividing and their numbers increase exponentially
• Constant, maximal growth rate
• Increased rate of metabolism
• Stationary phase - overpopulation and nutrients are depleted
• Production of spores for those that do not reach the death phase
• The death rate equals the growth rate
• Cell division stops due to depletion of nutrients and accumulation of toxic products
• Spore formation
• Death or decline phase
• Loss of viability - cells die due to
○ Toxic products
○ Loss of selective permeability
 Fluid gets into the cells causing cell lysis

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Pathogenicity and infection
Friday, 15 January 2016 2:49 PM

• Refer to certain aspects of the relationship between bacteria and humans harbouring them

Pathogenicity
• The ability or potential of an organism to cause disease
• Varies among the genera, species and strains
• Based on the pathogenicity, bacteria are grouped as:
○ Non-pathogenic bacteria
○ Pathogenic bacteria
○ Opportunistic pathogens

Pathogenesis
• The process in pathogenicity
○ Includes the initiation of the infectious process and mechanisms that lead to the signs and
symptoms or manifestations of the disease
○ Mechanisms occur within the organism and the infected person

Some properties of pathogenic bacteria which enhance pathogenicity

1. Transmissibility or communicability from one host or reservoir to a fresh host
a. Enhanced by the organism
i. Ability to be shed in large numbers in body fluids or secretions
ii. Ability to survive in adverse environmental conditions including dry surfaces or dust
2. Infectivity
○ Dependent on the ability of the organism to
 Attach itself
 Survive and penetrate a healthy host's first line of defence including the skin or the
mucous membranes
○ More difficult in some parts with intact barriers
○ Easier where the surface is broken
○ Related to:
 The number of the infecting bacterial cells
 Phase of growth
 Properties of the organism which enable it to harm or destroy tissues during infection

Outcome of pathogenesis
1. Resolution
2. Obvious illness due to the organism. Can be as:
a. Localised lesion at the site of entry due to inflammation - may be a swelling
b. Systemic illness or disease including multiple parts
3. Development of chronic carrier state after recovery
a. Organisms continue to grow
b. Can be a source of infection leading to disease in the same person or through transmission
to a susceptible person
4. Latent or asymptomatic or sub-clinical infection
a. May or may not develop into disease after several years
5. Unnoticed signs and symptoms
a. May develop into chronic carrier state
b. May present later with characteristics of long term complication associated with the
organism
c. Detectable evidence can be obtained by:
i. Serological tests
ii. Skin tests
iii. Tissue examination

Virulence
• The capacity of an organism to harm human tissues or to cause disease or the degree of
pathogenicity in relation to the host's resistance within a group of species
• Varies among the
○ Various pathogenic genera
○ Species
○ Strains
• Based on the virulence, pathogenic bacteria may be described as
○ Avirulent
○ Virulent
○ Highly virulent

Factors which influence pathogenicity and virulence

• Several include
a. Human factors: mostly immunological
b. Bacterial factors: including:
i. Ability of the organism to initiate infection
ii. Number of bacterial cells involved
iii. Presence of structures and production of certain substances by the organism which
enable it to cause harm to the human tissue
□ Referred to as virulence factors

Bacterial virulence factors

• Main determinants of virulence of a pathogen
• May be related to
○ Genetic composition
○ Biochemical properties

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 ○ Biochemical properties
○ Structural components
• Numerous with different mechanisms of action
○ Each factor can one or more mechanisms of action
• Can be grouped as:
1. Adherence factors or Adhesins
 Enable a bacterial cell to adhere to the cells or the tissue surface in humans and resist
being swept away where there are secretions
 Adherence of bacteria in general, is a process which can occur through effects of:
□ Pili or fimbriae on bacteria which possess them
□ Binding molecules at the bacterial cell surface and host cell receptor interaction
 Binding molecules on a bacterial cell surface include components of a
capsule and some surface
 Electrostatic forces to some extent
2. Aggresins or aggressive factors
 Enable the pathogenic organisms to overcome or invade host defence mechanisms
 Include
□ Capsules
 Prevents the interaction between the bacterial cell surface and phagocytic
cells that would lead to recognition that precedes phagocytosis
□ Surface proteins and carbohydrates
□ Enzymes

Aggressins that act intracellularly

• Found in some intra-cellular organism
• May enable the intra-cellular pathogen to:
○ Reside and multiply within phagocytic cells
○ Escape intra-cellular destruction by interference with:
 Phagosome and lysosome fusion
 Ability of lysosomal enzymes

Enzymes and enzyme like substances

• Enable the organism to invade and multiply in human tissues
○ Increase invasiveness
○ Include
 Hyaluronidase or spreading factor
 Coagulase
 Kinases or fibrinolysins
 Lipases
○ May also include toxins
 Endotoxins
 Exotoxins

Exotoxins
• Diffusible products from gram positive bacteria mostly and a gram negative organisms:
○ Protein in nature. Some are enzymes
○ Majority are heat labile. A few are heat stable
○ Liberated from the cytoplasm of live bacteria
○ Strongly antigenic
○ Can be converted into toxoids
○ Highly specific for certain tissues
○ High potency
○ Effectively neutralised by antitoxin
○ Synthesis of the majority is encoded by extra-chromosomal genes

Mechanisms of actions of exotoxins

• Different and depend on the specific exotoxin
• May involve
○ Inhibition of protein synthesis
○ Interference with normal nervous response to stimuli
○ Activity on the intestinal tract
○ Enzymatic activity

Endotoxins
• Consist of lipopolysaccharides liberated from the cell wall of dead or disintegrating gram negative
bacteria
• Characteristics:
○ Heat stable
○ Not convertible to toxoids
○ Moderately toxic and low in potency
○ Have no specific receptors
○ Poorly or non-antigenic
○ Not effectively neutralised by antitoxin
○ Synthesis directed by chromosomal genes

Effects of endotoxins
• Generalised with variable manifestations
○ Low levels in the blood causes release of pyrogens which can cause fever
○ High levels in the blood interfere with normal physiological functions including
immunological and haematological functions
 Associated with severe manifestations involving multiple systems including
hypertension and endotoxic shock and associated complications

Other bacterial properties which contribute to virulence

• Ability of the pathogen to obtain iron
○ Characteristic of most pathogenic bacteria
○ Organisms release small carrier molecules which obtain iron from the infected tissues
○ Iron is essential for the nutrition of pathogenic bacteria
 Availability supports the organisms growth and virulence

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  Availability supports the organisms growth and virulence
○ Production of many extra cellular products involved in virulence including some endotoxins
are influence by availability of iron

Plasmids
• May consist of genes which code for
○ Resistance to antimicrobial agents
 Genes responsible referred to as R-factors
 Resistance leads to continued growth and spread of infection despite treatment
○ Formation of sex pili and ability to transfer genetic material to other organisms
○ Other characteristics which contribute to virulence including endotoxin production

Change of surface antigen

• Some pathogenic bacteria undergo mutation during infection
○ Acquire new antigenic determinant on the cell surface
• Change of surface antigens
○ Enables the organism to avoid recognition by the immune defence mechanisms and
enables it to evade elimination

Infection
• A process which is necessary for an organism to cause disease
• Sometimes used to mean disease
• Involves
○ Sustained presence of an organism in an anatomic part
 Can be on the surface including the skin or the mucous membranes or in the inner
tissues
○ Establishment
○ t and multiplication at the site or colonisation
 With or without disease manifestation

Host and tissue specificity

• Some organisms are known to be
○ Restricted in the type of animals they infect
○ Highly specific in the tissues they infect primarily and in disease production
• Reasons for host-parasite or tissue specificity are not clearly identified

Sources of infection and reservoirs

• Reservoirs
○ Normal growth habitats of organisms
○ Can be either
 An anatomic site on animal including humans
 In the environment including water and soil
○ Source or origin of infection for a susceptible person can be a reservoir or an infected
person or animal
○ Infections associated_

Exogenous infections
• Organisms involved originate from sources outside the infected person including
○ Patients
○ Human carriers
○ Animals
○ Environment
Endogenous infections
• Organisms involved originate from a site within the individual mostly
○ Pathogenic organisms in asymptomatic carriers
○ Potential pathogens particularly among the normal flora

Modes of transmission and spread of pathogenic bacteria

• Determined largely by the site and type of infection and the organism
○ Respiratory tract infections
 For the majority bacteria are carried in infected secretions and are released through
□ Sneezing
□ Coughing
□ Speaking in droplet spray secretions
 Infected secretions_
○ Skins and wound infections
 Spread through contact with the infected part or material from that part or via
contaminated hands, clothing or other stuff
○ Sexually transmissible bacteria
 Via sexual contact either through mucosal to mucosal surface in secretion
○ GIT infections
 Majority of bacteria are ingested
 Some of the bacteria are passed out in faeces and cause contamination of water,
hands, or food substances from which re-ingestion may occur
○ Bacteria in the blood circulatory system
 Arthropod borne or vector borne
□ Spread by blood sucking arthropods during feeding
 From infected pregnant female to unborn baby or trans-placental transmission
 Through transfusion of infected blood

Bacterial Flora
_

Transient flora
• Inhabit the skin or mucous membrane for a limited period
○ Can be hours days or weeks
• Derived from the immediate environment
• Not associated with the disease under normal circumstances
• May cause disease when there is a disturbance in the defence mechanisms in that part

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Resident flora
• Relatively fixed types of micro-organisms
• Regularly found in specific anatomic parts
• Re-establish themselves if disturbed
• First lot is acquired by the new born baby during passage through the birth canal
• Subsequently organisms are acquired from other humans and inanimate sources in the
environment

Distribution of resident flora

• Partly influenced by
○ General physiological characteristics including
 Body temperature in various parts
 pH of secretions
 Oxygen
 Moisture
 Nutrients in specific parts
○ Other characteristics including age, sex and hormonal activity especially in females
○ Associated with fluctuations throughout life
 Few in early childhood and increase with age Laboratory acquired accidental infection
• Potential sources include:
Anatomic parts considered free of normal flora ○ Infected specimens
• Lower respiratory tract including ○ Artificial cultures
○ Trachea, bronchi, bronchioles and alveoli ○ Laboratory animals
 Various defence mechanisms clear the micro-organisms that may enter
□ Ciliated epithelium hat moves the mucous upwards
□ Phagocytic cells such as polymorphs and macrophages that carry out
phagocytosis of microorganisms
 Pleural cavity and fluid are usually sterile
• GIT
○ Oesophagus
 Naturally considered free of normal flora
 May contain bacteria from ingested food or saliva from the mouth as transient flora
○ Stomach
 Acidity plays a protective role by keeping the number of organisms at a minimum
○ Upper part of the small intestine
 Close to and therefore similar to the stomach
• Genitourinary tract
• Others
○ Circulatory system and pericardial space and fluid
 Blood and pericardial fluid and free of normal flora
○ CNS including the CSF
○ Peritoneal cavity and fluid
 Any organism isolated from a sample of properly collected specimen

Parts considered to harbour numerous normal microbial flora

• Skin
○ Exposed to the environment therefore harbours transient flora as well as resident flora
which are present in large numbers
 Influenced in parts by micro-environmental; factors including availability of moisture
○ _
○ Some physiological mechanisms of the skin keep the number and types of organisms in
check
 Also make it unfavourable for most bacteria
□ Mechanisms include
 Secretions with low pH
 Sebaceous secretions that contain fatty acids and lyzosomes or an
enzyme-like substance which is bacteriolytic
 -
• Nose - similar flora as the skin
○ Staphylococcus aureus is a significant normal flora I the nasal cavity in some people
 Occasional acts as a source of infection leading to disease
• Mouth
○ Majority are gram positive cocci, mainly alpha haemolytics streptococci or viridans
streptococci
○ Others
 Lactobacilli
 Bacteroides
• Pharynx or upper respiratory tract
○ Are oral cavity flora plus others including:
 Streptococci pneumoniae
 Streptococci pyogens
 Haemophilius species
 Moraxella catarrhalis
• Large intestines
○ Contains numerous organisms as normal flora
 More than 100 different species
 Consists of
□ Strictly anaerobic bacilli and cocci
□ Strict Facultative anaerobes specifically pseudomonas
□ Facultative anaerobes bacilli and cocci
• Lower urethra
○ Detectable in urine occasionally as contaminants
• Lower female genital tract
○ Dominant flora changes according to the pH which is under hormonal influence and age
 Soon after birth, the majority of the flora are lactobacilli as the pH is acidic
 As the baby grows, mixed flora of cocci and bacilli
 At puberty, lactobacilli dominant and maintain the acidic pH through their activity on
glycogen - a protective mechanism
 Mixed flora recurs after menopause
○ Generally, organisms from the large intestine may form part of vaginal flora due to close

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 ○ Generally, organisms from the large intestine may form part of vaginal flora due to close
proximity with the opening to the large intestine including:
 Species of clostridium
 Anaerobic streptococci
 Group B ß-haemolytic streptococci

Significance of normal flora in general

• Beneficial effects
○ Competition for microenvironments more effectively than some regular pathogens. Enables Significance of Skin Flora
prevention of colonisation by the pathogens • Increased risk of :
○ Stimulation of the immune system ○ Contamination of specimens collected from and through the skin
○ Metabolic activity which may be protective ○ Introduction and infection of inner tissues through clinical invasive
○ Production of nutrients e.g. Vitamin K procedures depending passage through the skin
○ Initiation focus of infection associated with devices inserted through
• Harmful effects
the skin and left there fore a long time
○ Potential causative agents of disease as:
○ Wound infection
 Opportunistic pathogens maybe in them
○ _
□ Dental carries and mouth infection
□ Damage tissue
□ Physiological and other abnormalities which reduce immunological defence
mechanisms
 A result of relocation from the usual occupied site to another
 A result of prolonged administration of broad spectrum antibiotics resulting in
disturbance and overgrowth or superinfection by the flora which ae not susceptible
○ Contribute to the spread antibiotic resistance
Significance
• Disease locally and in other parts due to alteration in
immunological defence mechanisms
• Alpha haemolytic streptococci - dental carries and
infection in the mouth and other tissues including
bacterial endocarditis
• Other normal flora associated with mouth infections
include:
○ Actinomycin species
○ Spirochetes depending on other factors and other
organisms
○ Bacteroides and other strict anaerobic organisms in
the mouth occasionally in internal tissues
○ _

If relocated to another anatomic site including

Urinary tract
Peritoneal cavity _

Significance (vaginal)
• Protection by lactobacilli through a low pH
• Group B streptococci can be transmitted to a new-born during vaginal
delivery resulting in infection
• Organisms derived from intestinal flora are significant causative
agents of
○ UTI
○ Infections associated with reproduction in females including child
birth

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Bacterial Genetics
Friday, 5 February 2016 2:35 PM

The Genetics of Bacteria

Structure and Functions of Genetic Materials
• DNA and RNA
○ Basic building blocks
 Nucleotides
□ Phosphate group
□ Pentose sugar
□ Protein bases
• DNA consists of 2 long polymers of nucleotides
• Backbones made of sugars and phosphate groups joined by ester bonds
• The two strands run in opposite directions to each other and are therefore antiparallel
• Attached to each sugar is one of the 4 molecules called nucleobases

Structure of DNA
• Attached to each deoxyribose and the phosphate residues arranged alternatively
• Attached to each of the 4 nitrogen bases
• Purines - adenine, guanine
• Pyrimidines - thymidine and cytosine

The Flow of Genetic Information

• Transcription
• Expression
○ Genetic information is used within the cell to produce the proteins that are needed
for the cell to function
• Recombination
○ Genetic information can be transferred between cells of the same generation
• Replication
○ Genetic information can be transferred between generations
• The cells uses the genetic information contained in DNA to make proteins as well as
enzymes.
• The information is transferred between generations through cell division
• _

Bacterial Sexual Processes

• The three bacterial sexual processes
1. Conjugation
 Direct transfer of DNA from one bacterial cell to another
2. Transduction
 Use of a bacteriophage (bacterial virus) to transfer DNA between cells
3. Transformation
 Naked DNA taken up from the environment by bacterial cells

Conjugation
• Donor/male cell makes contact with the recipient/female cell
• Genes transferred from the donor to the recipient via the sex pilus
• Recipient strain is converted into a donor cell

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Transduction
• Transfer of DNA between cells by bacteriophages (viruses)
• Lytic and lysogenic phages
○ Lytic phage
 Virulent phages lyse the host cell and produce a lytic cycle of infection
○ Lysogenic phage
 Temperate phages are able to infect the cell without necessarily causing
immediate cell lysis or death
• Corynebacteriun diphtheriae produces diphtheria toxin when it is lysogenised by B-phage
• Production of toxins by staphylococci, streptococci and clostridia - dependent on lysogenic
conversion by specific bacteriophages

Transformation
• Transfer of naked DNA from one cell to another
• Lysed cells expose the DNA
• Cells are usually competent for transformation in the late log phase
• Utilised in genetic engineering
• Certain types of bacteria can "donate" a piece of their DNA to a recipient cell
• The recombination is the bacterial equivalent of sexual reproduction in eukaryotes
• Final step after any method of gene transfer is recombination
• Rearrangement and integration of donor and recipient genomes
• Results in formation of new, hybrid genome

Genetic Basis of Antibiotic Resistance

• 2 mechanisms:
○ Chromosomal mutations
 Structural changes e.g.
□ Mutations in penicillin binding proteins - penicillin resistance
□ Modification leads to decreased drug affinity
 Physiological changes
□ Mutations in the katG gene in M. tuberculosis
 Isoniazid resistance
 katG gene - codes for catalase peroxidase enzyme
 Lack of the enzyme means that the drug cannot be activated
○ Transferrable antibiotic resistance

Applications of Genetic Engineering

1. Impact on Diagnosis and Research
○ Diagnostic tests:
 Specific gene probes to identify pathogens
○ DNA sequencing and molecular typing of micro-organisms - epidemiological studies
○ Therapeutic uses
2. Social impact
○ Human and animal experimentation
○ Ecological disruption
○ Agriculture - GMO foods
○ Threat of biological warfare

Harnessing the power of recombinant DNA technology - Human Insulin Production by bacteria
1. Isolate human cells and grow in tissue culture
2. Isolate the DNA from the human cells
3. Cut the DNA with a restriction enzyme
4. Meanwhile, isolate plasmid DNA from the bacterium
5. Use the same restriction enzyme to cut the plasmid DNA, creating matching sticky ends

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 5. Use the same restriction enzyme to cut the plasmid DNA, creating matching sticky ends
6. Join the plasmid and human fragment
7. Allow new bacteria to incorporate the recombinant plasmid into the bacterial cells then
screen the bacteria to find the ones that have incorporated the human gene for insulin
8. Grow trillions of new insulin-producing bacteria
• Collect the bacteria, break open the cells and purify the insulin protein and packaged

Edible vaccines
1. Cut the leaf

1. DNA encoding the antibodies is inserted into the bacterium

2. Tobacco plants are inoculated with the bacteria, which delivers the DNA into the plant cells
3. Plant cell machinery reads the DNA and produces antibodies
4. Antibodies are harvested from the plants
5. Antibodies are purifies into a form that can be given to humans
6. Once inside the infected patient, antibodies bind to the Ebola virus and prevents it from
doing further damage

Recombinant DNA products being used in human therapy

• Human genes have been cloned in E. coli or in yeast
• This has made it possible, for the first time, to produce unlimited amounts of human
proteins in vitro
• Cultured in cells (E. coli, yeast, mammalian cells) transformed with the human gene are
being used to manufacture
○ Insulin for diabetes
○ Factor VIII for males suffering from haemophilia A
○ Factor IX for haemophilia B
○ Human growth hormone (GH)
○ Erythropoietin for anaemia
○ 3 types of interferons
○ Several interleukins
○ GM-CSF for stimulating the bone marrow after marrow transplant
○ Human plasminogen activator for dissolving blood clots
○ Adenosine deaminase for treating some forms of severe combined
immunodeficiency (SCID)
○ Angiostatin and _

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Staphylococci
Friday, 12 February 2016 2:42 PM

• Gram positive cocci arranged in irregular grape like clusters

• Reside normally on the skin or mucous membranes on humans
• Produce catalase-positive

Grouping
• Clinically they are grouped into two:
○ Coagulase positive staphylococci
 Staphylococcus aureus
○ Coagulase negative staphyloccoci
 Frequently involved in nosocomial opportunistic diseases
S. epidermidis
S. Saprophyticus

Basic characteristics of S.aureus

• Gram positive cocci in clusters
• Produces coagulase and catalase enzymes
• ß-haemolytic on blood agar
○ HAEMOLYSIN ENZYME
• Normal flora of the nasal passages

Mechanisms of pathogenicity
• Capsule
○ Anti-phagocytic
○ Protect bacteria from being attacked by phagocytic cells
• Teichoic acids
○ Anti-phagocytic
• Protein A
○ Hs great affinity to Fc portion of IgG and inhibits phagocytosis and opsonisation
• Produces enzymes contribute to its evasiveness
○ Coagulase
 Clots plasma
 Interferes with phagocytosis
 Facilitates spread in the tissues
○ Haemolysins
 Lyse red cells
○ Leucocidin
 Kill leukocytes
○ Fibrinolysin
 Digests fibrin
○ Lipase
 Break down fat
○ Hyaluronidase
 Facilitates spread in tissues by destroying hyaluronic acids
○ ß-lactamase
 Associated with antibiotic resistance
• Produces toxins that contribute to tissue damage
○ Enterotoxins (heat stable) - cause food poisoning
○ Toxic shock syndrome toxin - shock, rash, desquamation of skin
○ Epidermolytic toxins A and B - generalised peeling of the skin

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 ○ Epidermolytic toxins A and B - generalised peeling of the skin

Clinical Features
• Causes pyogenic diseases and toxin-mediated disease
○ Pyogenic diseases
 Local lesions of the skin - impetigo, furuncles (boils) and carbuncles (boils
clustered together), folliculitis, eyelid infection
 Systemic infections - septicaemia, endocarditis, osteomyelitis, arthritis, post
surgical wound infection
○ Toxin mediated diseases
 Food poisoning (gastroenteritis)
□ Due to enterotoxin
□ Patients present with vomiting and watery non-bloody diarrhoea which
resolves within 24 hours
 Toxic shock syndrome
□ Mediated via enterotoxin 'toxic shock syndrome toxin'
□ Characterised by fever, erythematous skin rash, desquamation of the
palms of the hands and the soles of the feet and shock
 Staphylococcal scalded skin syndrome (SSSS)
 Mediated by exfoliative or epidermolytic toxin
□ Characterised by fever and large erythmatous rash results in sloughs off
the body
□ Common in children, neonates and adults with renal failure

Outline of Lab isolation and identification

1. Specimen collection
a. Pus
b. Skin
c. Stool
2. Gram stain from specimen
3. Culture in media
4. Colonial morphology
5. Gram stain of colony
6. Biochemical tests
7. Antibiotic susceptibility test

Lab diagnosis
• Specimens
○ Pus and swabs from infected sites, sputum, blood nasal swabs from carriers
• Microscopy
○ Gram stain and observe for gram positive cocci in clusters
• Culture
○ Culture on either
 Blood agar
 Mannitol salt agar
• Colonial morphology
○ Blood agar: golden yellow that are ß-haemolytic
○ Mannitol salt agar - yellow colonies
• Biochemical tests
○ Catalase positive - differentiate from streptococcus
○ Coagulase positive - differentiate from other staph. Spp

Antimicrobial susceptibility
• Treated with methicillin, vancomycin
○ MRSA (methicillin resistant S. aureus): resistant to methicillin and related penicillins -

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 ○ MRSA (methicillin resistant S. aureus): resistant to methicillin and related penicillins -
treated with vancomycin or clindamycin and Linezolid
○ VRSA (vancomycin resistant S. aureus) treated with a combination of Quinupristin -
dalfopristin (synercid) or Linezolid

Other Staphylococci species

Staphylococcus Epidermidis
• Characteristics
○ Gram positive cocci - grape like clusters
○ Non-haemolytic on blood agar
○ Catalase; coagulase negative
○ It does not ferment mannitol
○ Sensitive to novobiocin ( s. saprophiticus __)
○ Normal flora of the skin
○ Frequent flora of the skin
○ Frequent contaminant - blood specimens
○ Hospital - acquired; cause endocarditis and bacteraemia following infection
indwelling catheters or other medical appliances positioned in the body - production
of biofilm
○ Also causes sepsis in neonates and peritonitis in patients with renal failure -
peritoneal dialysis
○ Treated with vancomycin + rifampicin/aminoglycoside

S. Saprophyticus
○ Saprophytic
○ Coagulase negative
○ Inhabits the skin surrounding the genitourinary tract of females - cause of UTI in
sexually active females
○ Infections are strongly associated with presence of foreign bodies
 Prosthetic heart valves (endocarditis)
 IV catheters (bacteraemia)
 Urinary catheter (UTI in elderly)
 CSF shunts (meningitis)
 Peritoneal dialysis catheter (peritonitis)
○ Treated with quinolone (norfloxacin) or trimethropim-sulphamethoxazole

Micrococcus
• Gram positive cocci forming pairs , tetrads (predominantly)
• Catalase positive, coagulase-negative
• Transient flora on exposed skin of face, arms, hands, and legs
• Bacitracin - susceptible
• Are strict aerobes
• Associated with pulmonary infections recurrent bacteraemia, septic shock, septic arthritis,
endocarditis, meningitis

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Streptococci (Gram Positive Cocci)
Friday, 26 February 2016 2:28 PM

Characteristics
• Gram positive
• 1µm in diameter
• Occur in chains or pairs
• Usually capsulated
• Non-motile
• Non spore forming
• Facultative anaerobes
• Fastidious
• Catalase negative (staphylococci are catalase positive)

Classification
• Can be classified as:
○ Oxygen requirements
 Anaerobic (peptostreptococcus)
 Aerobic or facultative
○ Serology (lanciefield classification)
○ Haemolysis on blood agar

Serology: Laciefield classification

• Can be classified into many groups from A-K and H-V
• Classification based on C-carbohydrate antigen of cell wall
○ Groupable
 A, B and D (more frequent)
 C, G, and F (less frequent)
○ Non-groupable
 S.pneumoniae
 Viridans streptococci
□ E.g. S.mutans
□ Causing dental carries

Haemolysis on blood agar

• Haemolysis on BA
○ &-haemolysis
 Partial haemolysis
 Green discolouration around the colonies
 E.g. non-groupable streptococci (s.pneumonae, s.viridans)
○ ß-haemolysis
 Complete haemolysis
 Clear zone of haemolysis around the colonies
 E.g. group A and B (s.pyogens and s.agalactiae)
○ Gamma-haemolysis
 No lysis
 E.g. group D (enterococcus spp.)

Beta Haemolytic Group A Streptococcus ( Streptococcus pyogens)

• Pyogens meaning pus producing
• One of the most important pathogens
• Gram positive
• Found in chains

Virulence Factors
• Streptolysin O (SLO)
○ Oxygen labile
○ Damaged cardiac cells
○ Antigenic - produce SLO
• Streptolysin S (SLS)
○ Oxygen stable
○ Non-antigenic
• Exotoxin
○ Streptococcal pyogenic exotoxin (SPEs)
○ Manifestation of scarlet fever
• Exoenzymes
○ Streptokinase (fibrinolysin)
○ Streptodornase (DNAase)
○ Hyaluronidase

Manifestations
• Sore throat (streptococcal pharyngitis or tonsillitis)
• Rash or scarlet fever
• Cellulitis
• _

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 • _

Pathogenicity
• Respiratory infections
○ Pharyngitis (sore throat), tonsillitis
○ Otitis media, sinusitis
• Skin and subcutaneous infections
○ Pyoderma, cellulitis
○ Necrotising fasciitis (flesh eating bacteria)
• Non-suppurative complications
○ Acute rheumatic fever - usually follows streptococcal pharyngitis
○ Acute glomerulonephritis - usually follows pyoderma

Strep Throat
• Most common of all Strep disease
• Spread by saliva or nasal secretions
• Incubation period 2-4 days
• Sore throat, slight fever (101)
• Treat immediately in children and elderly to prevent post strept disease

Treatment
• Penicillin G or erythromycin are drugs of choice
• Although the disease is self-limiting it is important to treat immediately to prevent post strept disease

Lab diagnosis
• Specimens:
○ Throat swabs
○ Pus
○ Blood
• Microscopy
○ Gram stain - GPC in chains
• Culture
○ Blood agar - ß-haemolytic colonies
• Identification tests
○ Catalase negatives
○ Bacitracin sensitive
○ Penicillin sensitive

Beta haemolytic Group B Streptococcus

• Normal flora in the lower GIT, female genital tract
• Pathogenicity
○ Neonatal meningitis
○ Puerperal sepsis
○ Pneumonia

Lab diagnosis
• Specimens:
○ CSF
○ Blood
○ Vaginal smears
○ Urine
• Microscopy
○ Gram stain - GPC in chains
• Culture
○ BA- ß haemolytic colonies
• Identification tests
○ Catalase negative
 Bacitracin resistance
 CAMP Test +
 Penicillin sensitive

Differentiation between ß-haemolytic streptococci

• The following tests can be used to differentiate between ß-haemolytic streptococci
○ Lancifield classification
○ Bacitracin susceptibility tests
 Specific for s.pyogens (Group A)
○ CAMP test
 Specific for s.agalactiae (Group B)

Group D Streptococcus - Gamma Haemolytic

• Enterococcus - 2 important species
○ E.fecalis
○ E.faecium
• Normal flora in the GIT and lower genital tract
• Nosocomial/opportunistic pathogen

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 • Nosocomial/opportunistic pathogen
• UTI, wound infection, endocarditis
• Resistance to cephalosporins, even vancomycin

Lab diagnosis - enterococcus

• Specimens
○ Urine
○ Pus
○ Blood
• Microscopy
○ Gram stain - GPC in pairs or short chains
• Culture
○ BA - &/ß/no haemolysis
• Identification tests
○ Catalase negative
• Penicillin resistance

Alpha haemolysis streptococci (S. pneumoniae)

• Commonly known as Pneumococcus
• Normal inhabitants of the upper respiratory tracts of human beings
• Gram positive small (1µm), slightly elongated cocci, with one broad end and on pointed end, presenting a
flame shaped or lancolate shape

Morphology
• Occur in pairs with broad ends opposing each other
• They are encapsulated
• They are non-motile and non-sporing

Mechanism of pathogenesis
1. Entry of pneumococci into nasopharynx
2. Colonisation of nasopharynx
3. May cause infection of the middle ear, para-nasal sinuses and respiratory tract by direct spread
4. Infection of meninges can also occur
5. Enters blood causing bacteraemia, which may also lead to disseminated infection as in the heart,
peritoneum or the joint

Disease
• Otitis media and sinusitis
• Pneumonia
○ Lobar pneumonia
○ Bronchopneumonia
• Tracheobronchitis
• Meningitis
• Other infections
○ Empyema
○ Pericarditis
○ Conjunctivitis
○ Suppurative arthritis
○ Peritonitis

Lab diagnosis
• Specimen
○ CSF
○ Blood
○ Sputum
○ Pus
○ Swabs
• Microscopy
○ Gram stain - GPC in pairs, capsulated, lancolate shaped
• Culture
○ BA/CA - & haemolytic colonies
• Identification tests:
○ Catalase negative
○ Optochin sensitive
○ Bile solubility
○ Quellung reaction (capsular swelling
 It is described by Neufeld
 On a slide, the sputum is mixed with type specific antiserum against capsular antigen and a
loopful of methylene blue solution. The capsule becomes swollen and refractile

Treatment
• For penicillin sensitive strains, Penicillin is the drug of choice for serious cases and amoxicillin for milder
ones

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 ones
• Fro penicillin resistant strains, a third generation cephalosporin is indicated
• Vancomycin is to be reserved for life threatening illness with highly resistant strains

Pneumococcal vaccination
• 2 flavours of vaccine
○ Pneumovax: capsular polysaccharide of 23 serotypes: PPS23
○ Prevnar: capsular polysaccharide conjugated to protein (improves humeral response), 7 serotypes
originally, now up to 13: PCV13
• Recommendations
○ Use PCV13 for kids; multiple shot protocol
○ Adults with immunocompromised immunity, asplenia, etc. …get one dose of PCV13, then one
PPS23 at least 8 weeks later and 5 years thereafter
○ For adults above 65, give PPS23 once

Points to remember
• General characteristics and haemolytic
patterns

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Neisseriaceae
Friday, 26 February 2016 2:30 PM

• Family
○ Neisseriaceae
• Genera
○ Neisseria
○ Moraxella
○ Acinetobacter

Neisseria species
• 25 species and 3 subspecies
• Most important
○ N.menengitidis (meningococcus)
○ N.gonorrhoeae (gonococcus)
○ N.lactamica
○ N.cinerea
• N.lactamica and N. cinerea are commensals in the upper respiratory tract but may cause infection in
immunosuppressed individuals
• They are obligate human pathogens with no other natural host

General characteristics
• Gram negative (diplococci, kidney/bean shaped)
○ _
• Aerobic
• Non-motile
• Non-spore forming
• Oxidase +
• Catalase + (except N. elongata)
• Meningococcus is +/- capsulated; gonococcus is unencapsulated

Neisseria meningitidis
• Found in the healthy individuals
• Causes endemic and epidemic meningitis
• Transmission is airborne

Serological classification of meningococcus

• 12 sero-groups according to capsular polysaccharide structure
• A, B, C, H, I, K, L, W (formerly W-135), X, Y and Z
• Six (A, B, C, W, X, Y) cause life threatening disease
• Serotypes and sub-serotypes according to the outer membrane proteins
○ Serosubtypes, serotypes, and immunotype is based on PorA, PorB, and LOS structure respectively)

Epidemiology
• Infection is global but not uniform
• Sporadic, non-sporadic and epidemic disease occur
• 1.2M cases of meningococcal infection per year
• Mortality - 135,000 worldwide
• Invasive meningococcal disease mostly occurs in children and young adults
• 2000 - outbreak of sero-group W occurred during the 2000 Hajj in Saudi Arabia
• Africa meningitis belt

Meningitis belt
• Sub-Saharan region from Ethiopia to Senegal includes 18 countries
• Characterised by periodic large epidemics of predominantly meningococcal meningitis
• Epidemics occur every 8-10 years and began around 1905
• Environmental factors such as humidity and dust contribute
•

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 • Sero-group A has been responsible for the largest and most devastating meningococcal outbreaks in sub-
Saharan Africa

Virulence factors
factor
1. Polysaccharide capsule
○ Resists phagocytosis
○ Defines serologic group
2. Endotoxin (lipooligosaccharide)
○ Responsible for fever, shock, DIC, outer membrane proteins including
 Pili
 Porins PorA and B
 Adhesion molecules Opa and Opc
3. Surface adhesive proteins
○ OMPs e.g.
 Pili
 Porins
 Adhesion molecules
4. Iron sequestration mechanisms
5. Genetic mechanisms
○ Horizontal genetic exchange
○ High frequency phase and antigenic variation
○ Molecular mimicry

Pathogenesis
• Transmission by large droplets
• Colonisation ---> invasion ---> blood stream ---> production of a strong inflammatory response ---> activation of
complement and coagulation cascades ---> endothelial damage and capillary leakage ---> necrosis of peripheral
tissue and multiple organ failure

Clinical manifestations
• Urethritis
• Pneumonia
• Conjunctivitis
• Septic arthritis
• Pericarditis
• Invasive meningococcal disease (IMD)
○ Meningitis
○ Septicaemia = meningocaemia

IMD
• Meningitis
○ Fever
○ Headache
○ Stiff neck
○ Increased polymorphs
• Meningocaemia (meningococcal septicaemia)
○ Fulminant infection
○ Initial symptoms non-specific (fever, myalgia)
○ Then
 Maculopapular
 Petechial
 Purpuric rash
○ Hypotension, multiorgan dysfunction shock, peripheral ischaemia, limb loss, death
○ Waterhouse-Friderichsen syndrome
 Septic shock
 DIC with purpura fulminans
 Adrenal insufficiency
• Complications

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 • Complications
○ Skin scarring
○ Abnormal bone growth
○ Limb loss and multiple amputations
○ Hearing losses
○ Cognitive defects
○ Visual impairments
○ Educational difficulties
○ Developmental delays
○ Motor nerve deficits
○ Seizure disorders
○ Behavioural problems

Lab diagnosis
• Specimen
○ Blood
○ CSF
○ Sputum
• Gram stain: gram negative intracellular and extracellular diplococci
• Culture
○ Media
 BA
 CBA
 Castenada
○ Temperature
 35 -37 degrees, 18-24 hours, 5-10% Co2
• Colonial morphology
○ Small translucent grey colonies on CBA
• Gram stain of the colonies
○ Gram negative diplococci
• Biochemical test
○ Oxidase positive
○ Sugar fermentation test
 Breaks down (ferment) glucose and maltose
• Other tests
○ Latex agglutination
 Demonstrate capsular antigen
○ Molecular tests e.g. PCR
○ Antimicrobial susceptibility testing

Treatment and Control

• ß-lactam antimicrobial agents
○ Penicillin
○ Cephalosporins
• Vaccination
○ Reactive vaccination campaigns in response to outbreaks
○ Following splenectomy
○ Travellers to the annual Hajj pilgrimage
○ Routine immunisation
• Vaccines can be:
○ Monovalent
○ Bivalent (A,C)
○ Trivalent (A, C, W)
○ Quadrivalent (A, C, W, Y)
• Polysaccharide vaccines
• Polysaccharide-protein conjugate vaccines
○ MenC
○ MenA
○ MenACWY

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 ○ MenACWY
○ MenAfricVac
○ MenBOmv

Neisseria Gonorrhoeae
• Infects the non-ciliated, columnar, and transitional epithelia
• An exclusive human pathogen
• Transmission
○ Sexual
○ During birth

Virulence factors
• Pili
○ Adhesion molecule
○ Anti-phagocytic
• LOS
○ Endotoxin
• IgA1 protease
○ Hydrolyses secretory IgA
• Porin A
○ Inactivates C3b component of the complement system
• Genetic mechanisms
○ Phase variation
○ Antigenic shift
○ Recombination following conjugation

Epidemiology
• Occurs worldwide
• Second most common curable STIs
• Transmission
○ Sexual
○ MTC
• Increasing prevalence among gays
• Increasing increasing of anti-microbial ('super gonorrhoea')

Clinical manifestation
• Causes gonorrhoea
• Women
○ Asymptomatic infection is common
○ Endocervix - purulent vaginal discharge
○ Ascend the oviducts
 Acute salphingitis
 PID
 Infertility
 Ectopic pregnancy
• Men
○ Acute urethritis
○ Purulent penile discharge
○ Dysuria
○ Asymptomatic infection less common
○ Proctitis
• Pharyngitis in both men and women
• Babies
○ Ophthalmia neonatorum
○ Severe purulent eye discharge periorbital oedema
○ Occurs several days after birth
○ Can result in blindness

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 ○ Can result in blindness
• Dissemination
○ Peritoneal dissemination
 Peri-hepatic inflammation (Fitz-Hugh-Curtis syndrome)
○ Others
 Septic arthritis
 Endocarditis
 Meningitis

Lab diagnosis
• Specimen
○ Urethral and cervical exudates
○ Urine
○ Swabs
 Rectal
 Eye
 Nasopharyngeal
• The gonococcus is fastidious
○ Use Stuart's transport media
• Direct microscopic examination
○ For urethral swabs
 Gram negative intracellular diplococci = presumptive diagnosis of gonorrhoea in men
• Culture
○ CBA
○ Selective media
 TMM (Thayer-Martin Media) - Vancomycin, Nystatin, Colistin
 Modified TMM - above + Trimethoprim
 Martin Lewis media - Vancomycin, Colistin, Trimethoprim, Anisomycin
 NYC media - vancomycin, colistin, trimethoprim, amphotericin B
• Incubation
○ 35°C, 5-10% CO2, 18-24 hours
• Colonial morphology
○ Small, raised grey or translucent colonies
• Biochemical tests
○ Rapidly oxidase positive
○ Ferments glucose alone
• Antimicrobial susceptibility testing
• Serological test
• Molecular tests
○ PCR
○ Genotyping

Treatment and Prevention

• Treatment
○ Rapidly increasing antimicrobial resistance
○ Ceftriaxone - DOC
○ Ophthalmia neonatorum
• Prevention
○ 4Cs
○ Ophthalmia neonatorum
○ Erythromycin ointment; silver nitrate drops; tetracycline eye ointment

Moraxella
• Family:
○ Moraxellaceae
• Coccobacillary rods
• Assacharolytic
• Oxidase positive

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 • Oxidase positive
• Catalase positive
• Main pathogen
○ Moraxella catarrhalis (formally Branhamella catarrhalis)

Moraxella catarrhalis
• Only found in humans and transmitted by respiratory dropets
• Normal flora in the upper respiratory tract
• Can gain access to the lower respiratory tract of patients with chronic obstructive disease
• Infections
○ Otitis media
○ Sinusitis
• Primarily in children

Lab diagnosis
• Specimen
○ Swab
○ Sputum
• Gram stain
○ Gram - diplococci dispersed between pus cells
• Culture
○ BA
○ CBA; 5-10% CO2, 18-24 Hours
• Oxidase +
• Assacharrilytic

Treatment
Trimethoprim
Sulphamethoxazole
Or amoxicillin + clavulin (clavulanic acid)

Moraxella nonliquefaciens
• 1 of 2 common cases of blepharitis (also caised by Staph, aureus)
Treatment
• Local application of antimicrobial agents e.g. erythromycin

Acinobacter
• Gram negative coccobacillary rods
• Commonly found in soil and humans
• About 25% of adults carry the organism in the skin and 7% carry the organism in the pharynx
• Opportunistic pathogens
• Account for 1-3% of all nosocomial infections (second to P. aeruginosa)
• A.baumannii, A. calcoaceticus, A.nosocomialis, A.pittii
• Acinobacter bari
○ Sepsis, pneumonia, UTIs meningitis
• Acinobacter iwoffii (formerly A. calcoaceticus)
○ Causes disease associated with respiratory therapy equipment and indwelling catheters
• In hospital settings, acinobacter commonly colonises irrigating solutions and IV solutions
• Multidrug resistant A. pittii and A. nosocomialis strains
○ Infections in healthcare facilities around the world

Lab dignosis
• Grow on routine lab media
• BA
○ Smooth, mucoid and non-pigmented colonies
• Gram - coccibacilli
• Oxidase -
• Catalase +

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Treatment
• Isolates of A. baumannii are resistant to may antimicrobial drugs
○ Rifampicin + meropenem

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Bacteriology Page 252
Corynebacteria
Friday, 13 May 2016 2:12 PM

• Gram+ pleomorphic bacteria

• Club-shaped or have swollen ends
• Stain irregularly due to the structure of the cell wall
• An outer layer of mycolic acids is found which is functionally equivalent to the outer membrane in gram-
bacteria
• Non spore forming
• Non motile
• Not capsulated

Classification
• Class: actinobacteria
• Numerous species; >88species
○ Corynebactrium diptheriae
○ Diptheroids/coryneform bacteria/non-diptheriae corynebacteria
 C.striatum
 C.pseudodiptheriticum
 C.propinquum
 C.tuberculostearicum

Corynebactrium diptheriae
General characteristics
• Gram+ pleomorphic bacilli
• Facultative anaerobes; grows best under aerobic conditions
• Non
○ Motile
○ Capsulated
○ Spore-forming
• Decolourise very easily
• Have granules in the cytoplasm = metachromic/volutin granules
• Metachromatic granules - Babès-Ernst granules. Represent an accumulation of polymerised
polyphosphates
• Take certain shapes
○ Palisade arrangement
○ Described as resembling Chinese alphabet
○ Cuneiform arrangement
• Relatively resistant to drying
○ May maintain viability for weeks in dust or on dry articles
○ Susceptible to heat and regularly used detergents

Pathogenic properties
• Disease production occurs through
○ Local invasion
○ Effects of virulence factors
 Main virulence factors is the diphtheria toxin (exotoxin)

Diphtheria toxin
• Heat-stable polypeptide

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 • Heat-stable polypeptide
• AB
• Not released by all strains
○ Toxigenic
○ Non-toxigenic
• Ability to produce toxin is acquired through lysogeny
• Toxigenic strains: 3 different strains
○ Severe/gravis
○ Mild/mitis
○ Intermediate/intermedius

Clinical implications
• Diphtheria
• Wound infections
○ Infection through contamination and local invasion
○ Toxin production not observed

Diphtheria
• Acute febrile infectious disease - occurs in epidemics
• Mainly affects children
• Source of infection
○ Nasal carrier
○ Patient
• Transmission
○ Secretions or droplets from the URT
• Initial clinical manifestations confined to the URT or oropharynx result from:
○ Infection of the mucous membrane
○ Local invasion of the mucosal surface tissue
○ Toxin production at the site
○ Inflammatory response

Clinical manifestations
• Fever
• Inflammation of the URT
• Results in a greyish membrane like covering referred to as pseudomembrane
• Pseudomembrane
○ Products of acute inflammatory process e.g. exudate, WBCs and epithelial cells
• Swelling around the neck due to oedema and the large cervical lymph nodes (Bull's Neck appearance)
• Severe infections
○ Difficulty in breathing
 Pseudomembrane extends locally to the tonsils, pharynx, larynx and nasal passages

Complications
• Airway obstruction
○ Extensive pseudomembrane or when dislodged during sample collection
• Haematogenous spread of the toxin
○ Damage to the heart muscle
○ Degeneration of peripheral nerves
○ Damage to the adrenals
C.diphtheriae does not invade deeply into tissues below the mucous membrane and bloodstream invasion
is not a feature of illness

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Laboratory investigations
• Specimen
○ Throat, nasopharyngeal, skin swabs
• Gram stain
• Culture and isolation

Media
• Blood agar
• Selective media
○ Cystine tellurite blood agar/potassium tellurite blood agar (PTA)
○ Tinsdale medium
○ Loeffler's serum agar

• 20℃ - 40℃ with an optimum of 35℃ - 37℃, 24 hours

• Colony characteristics
○ Greyish white colonies on BA
○ PTA - grey or grey-black colonies
○ Tinsdale medium - grey-black ,raised and surrounded by a dark brown area
○ Loffler's serum - for granule formation
• Volutin granules (metachromatic granules) presence demonstrated by Albert's or Meissner's stain
• Biochemical tests
○ Oxidase -
○ Urease -
○ Catalase -
○ Reduces nitrate to nitrite
○ Ferments glucose and maltose producing acid, not gas
• Tests for toxin production
○ In vitro tests
 Elek's gel precipitation test
 Tissue culture test
○ In vivo tests
 Subcutaneous test
 Intradermal test
○ Molecular tests
 Tox gene

Treatment
• Anti-toxin given immediately based on the clinical manifestation
• Antimicrobials
○ Penicillin
○ Erythromycin

Prevention and control

• Detection and treatment of carriers and patients
• Isolation
• Active immunisation

Other Corynebacterium bacteria

• Diphtheroids or non-diphtherial corynebacteria
• Commensals of skin, throat, conjunctiva
○ C.ulcerans
○ C.bovis
○ C.haemolyticum

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 ○ C.haemolyticum
○ C.pyogens
○ C.xerosis
○ C.pseudotbercilosis

Clinical implications
• Ulcerative throat infections
• Wound infections
• Diphtheria like infection
• Wound infections

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Bacillus
Friday, 13 May 2016 2:50 PM

Classification
• Numerous species
• Bacillus anthracis
• Anthracoides
○ Bacillus
 Cerus
 Subtilis
 Pumilis
 Stearothermophilus

General characteristics
• Gram+ rods arranged singly or in chains
• Aerobic or facultative anaerobes
• For endospores which are heat resistant
• Most of the species are motile
○ B.anthracis is non-motile
• Majority of the species form complete haemolysis on BA
○ B.anthracis is +/- haemolytic
• Catalase+

Bacillus Anthracis
General characteristics
• Gram+
○ Straight or slightly curved
○ Single cells or in chains
○ Bamboo rod in appearance
• Capsulated in tissues
• Forms spores
○ When shed into environment or grown on artificial media
○ Spores are oval, centrally placed and same diameter as the width of the cell
• Vegetative cells: destroyed by heat at 60℃ for 30 minutes
• Spores
○ Relatively resistant to disinfectant and heat
○ Some can withstand dry heat at 140℃ for 1 - 3 hours and boiling at 100℃
○ Can survive in the environment or in the hosts for long

Virulence factors
• Capsule
• Exotoxin
○ Lethal toxin (LT)
 Protective antigen (PA) + lethal factor (LF)
○ Oedema factor (ET)
 PA + EF
○ Individually, each factor is non-toxic but together, act synergistically to produce
damaging clinical effects

Clinical implications
• Causes anthrax

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 • Causes anthrax
• Associated with outbreaks
• Primarily a disease of animals including, cattle, sheep and goats
• Human infections are accidental

Transmission
• Exists in the environment as a spore and can remain viable in the soil for decades
• Spores ingested by grazing herbivores
• Germinate within the animal to produce the virulent vegetative forms
• Replicate and eventually kill the host
• Products (meat and hide) from an infected animal serves as a reservoir for human disease

Clinical manifestations
• Cutaneous anthrax
○ Small pimple or pustule appears at the sire of the site of inoculation 2 - 3 days after
exposure
○ Ring of vesicles develops ---> coalesce to form an erythematous ring
○ A small dark area appears at the centre of the ring and eventually ulcerates and
dries (eschar)
• Inhalation anthrax/Woolsorter's disease
○ Spores included
○ Severe signs and symptoms
○ Starts as a non-specific illness; mild fever, fatigue and malaise 2 - 5 days after
exposure
○ Sudden severe case with respiratory distress
• GI anthrax
○ Oropharyngeal/intestinal
○ Spores are inoculated into a lesion on the intestinal mucosa following ingestion of
spores
○ Abdominal pain, nausea, vomiting, anorexia
• Injectional anthrax
○ First recognised in 2001 in Norway
• Septicaemic anthrax
○ Can arise as a complication of the other forms

Laboratory investigations
• Specimen
○ Fluid aspirated from cutaneous lesions
○ Sputum
○ CSF
○ Blood for culture
○ Highly infectious
• Gram stain
• In smears from specimen, B. anthracis is capsulated
○ Polychrome methylene blue (McFadyean) stain; Giemsa
• Culture
○ BA, incubate in air/CO2 at 35℃ - 37℃ for 25 - 48 hours
○ Polymyxin, egg yolk, mannitol, bromophenol blue blue agar (PEMBA) - optional
○ Colonial morphology
 BA
□ Colonies are large, grey and flat with irregular margins (Medusa head)
• Identification
○ Catalase+

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 ○ Catalase+
○ Lecithinase production - inoculate on egg yolk; zone of egg yolk precipitation
○ Non-motile

Treatment
• Penicillin
• Chloramphenicol
• Tetracycline
• Erythromycin
• Streptomycin
• Fluoroquinolones
• First generation cephalosporins
• Anthrax is resistant to many later-generation cephalosporins

Prevention and Control?

Anthracoides
• Saprophytes - frequent containment of cultures
• Motile
• Not capsulated
• Not encountered as regular pathogens

Bacillus cereus
• Associated with
○ Food poisoning
 Toxin induced - rice and meat stews
 Mild - diarrhoea and vomiting - a few hours
○ Severe eye infections
○ Anthrax-like progressive pneumonia
○ Fulminant sepsis
○ CNS infections

Bacillus stearothermophilus
• Produces spores which are heat resistant
• Destroyed by heat at 121℃ in autoclave for at least 12 minutes
• Useful for testing the efficacy of the autoclaves

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Lactobacillus
Friday, 13 May 2016 3:23 PM

General characteristics
• Mainly saprophytes and commensals in humans and other animals
• Morphology
○ Gram+ bacilius
○ Non-motile
○ Non-spore forming
• Growth
○ Slow
○ Microaerophilic
○ pH 6.0
• Identification
○ Fermentation of carbohydrates with acid production
○ Tolerance to acidic conditions
• Species
○ Lactobacillus acidophilus complex
○ Oral cavity, GUT
• Play an important role in the health of the female GT
• Produce lactic acid from glycogen
• Lower vaginal pH and suppress overgrowth of micro-organisms
• Low virulence (bacteraemia in immunosuppression)

Bacteriology Page 260

Listeria
Friday, 13 May 2016 3:28 PM

General characteristics
• Isolated form wild and domestic animals and in the environment
• 7 species
○ Listeria monocytogenes
○ Listeria ivanovii
• Gram+ rods
• Facultative anaerobes
• Non-spore forming
• L.monocytogenes is an intracellular foodborne pathogen
• Causes listerosis and severe infections in humans
• High mortality rate, mainly in high risk groups including pregnant women

Listeria monocytogens
• Virulence factors
○ Haemolysin (listeriolysin O) - damages the phagosome membrane preventing killing
of the organism
○ Catalase
○ PLC
○ Surface protein p60 - induces phagocytosis through increased adhesion and
penetration
• Transmission
○ Ingestion
○ Mother to child

Clinical infections manifestations

• Fever, non-specific
• Pregnant women
○ Third trimester
○ Responsible for spontaneous abortions and stillborns
• Neonates
○ L.monocytogenes - very serious infections; high mortality rate
○ Early onset or late onset

Laboratory investigations
• gram+ coccobacilli
• Optimal growth temperature is 30 - 35℃ but growth occurs over a wide range
• Catalase+
• Motile at room temperature
• In wet mount preparations, exhibits tumbling motility
• Bile aesculin hydrolisis +
• Positive CAMP

'℞/prevention and control

• Penicillins
• aminoglycosides

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Antibiotic Antimicrobial Agents
Tuesday, 25 April 2017 9:38 AM

Introduction
• Antibiotic
○ Chemical substance produced by various species of microorganisms that is capable
in small concentrations of inhibiting the growth of other microorganisms
• Semi-synthetic antibiotics
○ Synthetic derivatives of naturally-occurring antibiotics
• Chemotherapeutic agents
○ Chemical antimicrobial agents
• Antimicrobial agent
○ Chemical substances produced by a microorganism or wholly or partially by chemical
synthesis, which in low concentration, inhibits the growth of other microorganisms
• Bacteriostatic
○ Antimicrobial agents that inhibit growth and replication of bacteria
○ Are non-lethal
• Bactericidal
○ Antimicrobial agents that cause bacterial cell death by inhibition of
 Cell wall synthesis
 Nucleic acid synthesis
 Protein synthesis
• Selective toxicity
○ Highly effective against the microbe but have minimal or no toxicity to humans

Classification
• Targeted Microorganisms
○ Antibacterial
○ Antifungal antibiotics and chemotherapeutic agents
○ Anti-parasitic agents
○ Anti-mycobacterial agents
• Mode of action

Interference with Cell Wall Synthesis

• Act on the formation of the PG layer
• Beta-lactam antimicrobial agents
○ Feature a ß-lactam ring in their structure
○ Mimic a D-ala-D-ala moiety
○ Form an irreversible intermediate within active sites of PBPs
○ Weaken PG - subsequent inhibiion of cell growth/lysis
 Penicillins
 Cephalosporins
 Monobactams
□ Aztreonam - aerobic gram negative bacteria
 Carbapenems - broad spectrum activity
□ Imipenem
□ Meropenem
 Oxa-cephems
□ Latamoxef
• Glycopeptides
○ Large molecules unable to penetrate the outer membrane of gram negative bacteria
○ Spectrum limited to gram negative organisms
○ Last resort for multi-drug resistant gram positive organisms e.g. MRSA, Enterococci

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 ○ Last resort for multi-drug resistant gram positive organisms e.g. MRSA, Enterococci
 Vancomycin
 Teicoplanin
 Telavancin
• Bacitracin
○ Gram positive bacteria
○ Too toxic for systemic use
○ Topical; presumptive identification of GAS
• Cycloserine
○ Produced by Streptomyces
○ Analogue of D-alanine
• Fosfomycin
○ Antibiotic produced by Streptomyces and Pseudomonas spp
○ Targets Mur enzyme
• Isoniazid

Disruption of bacterial membrane structure

• Act like cationic detergent to disrupt cell membranes
○ Polymixin B
○ Colistin

Inhibition of Protein Synthesis

• 30S
○ Anminoglycosides (Streptomycin, Neomycin, Kanamycin, Gentamicin, Amikacin)
○ Tetracyclines (Tetracycline, Doxycycline, Minocycline, Tigecycline)
• 50S
○ Macrolides (Erythromycin, Clarithromycin, Azithromycin)
 Prevent the continuation of protein_
○ Chloramphenicol, Thiamphenicol
○ Oxazolidinones (Linezolid)
○ Streptogramins (Guiniprostin-Dalfopristin)

Interference w/ Nucleic Acid Synthesis

• Sulphonamides and Diaminopyrimidines
○ Affect folic acid metabolism
 Sulphonamide - analogues of PABA
□ Sulphamethoxazole
□ Sulphadoxine
□ Sulphadiazine
 Diaminopyrimidines - prevent reduction of DHF to THF
□ Trimethoprim
□ Pyrimethamine
□ Cycloguanil
• Quinolones
○ Act on the subunit of DNA gyrase
 Nalidixic acid - active only against gram negative
 6-fluoro derivatives
□ Ciprofloxacin
□ Ofloxacin
• Nitroimidazole
○ 5-nitroimidazoles
○ Active only against anaerobic, certain micoaerophilic bacteria
 Metronidazole
 Tinidazole
 Ornidazole
 Nimorazole

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  Nimorazole
○ Nitrofurans
 Nitrofurantoin
○ Novobiocin
 Acts on ß subunit of DNA gyrase
○ Rifamycin
 Inhibit RNA transcription
□ Rifampicin
□ Rifabutin

Antibiotic Combination Results

• Synergism
○ The effect of the combination is greater than the sum of the effect of its components
penicillin and aminoglycoside
• Antagonism
○ Combination of antibiotics in which of one interferes with the activity of the other e.g.
a bacteriostatic antibiotic like tetracyclines and ß-lactams that act on dividing cells

Antiimicrobial Resistance
Bacterial Resistance Mechanisms
• Increased bacteria, increased selective pressure
○ Mutations and/or reduction of or expression alterations in PBPs
 Major resistance mechanism by gram-positive bacteria
○ ß-lactamase production
 Hydrolyse the ß-lactam ring
 Major resistance mechanism by gram negative bacteria
○ Permeability alterations
○ Extrusion by efflux pumps
○ Ribosomal methylases
○ Ribosomal mutations
○ Target modifications
 Fosfomycin-resistant organisms: Mtb, CT, Borelia burgdoferi
□ Natural mutation at the target site
 Generation of PG precursors: Glycopeptides - alternatives to D-ala-D-ala e.g.
D-ala-D-ser
○ Porins
○ Antibiotic modification
○ Increased production of non-essential anti-microbial targets
 Cycloserine
○ Reduction in active site accessibility
 E.facium - natural resistance to ß-lactams
○ Bypassing a step in a pathways
 Transpeptidation step: Mycobacteria, Clostridium, Enterococcus spp

Beta Lactamase Inhibitors

• Reversible/Irreversible binding of the enzyme
○ Form unfavourble steric interactions
○ Extended-spectrum cephalosporins, monobactams, carbapenems
• Irreversible "suicide inhibitors"
○ Permanently inactivate the ß-lacatamase
 Clavulanic acid
 Sulbactam
 Tazobactam
○ Inhibitor-resistant ß-lactamases

Antimicrobial Susceptibility Testing (AST)

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Introduction
• Aim
○ Select effective anti-microbial drugs
○ Detect resistance in individual bacterial isolates
○ Several methods
 Dilution methods
 Disk diffusion method
 E-test
 Automated antimicrobial susceptibility test systems
 Mechanism-specific tests
 Genotypic methods

Diffusion Method
• Disc Diffusion
○ Kirby-Bauer Disk Diffusion Method
○ Nutrient agar used
• Stoke's Method
○ Both test and control organisms are inoculated in the same plate
○ Sensitive organisms - zone of inhibition ≥ that for control organism
○ Resistant organism - zone of inhibition < that for control organism or no zone of
inhibition
• Factors affecting disc diffusion
○ pH of medium
○ Media components
 Agar depth
 Nutrients
○ Stability of the drug
○ Size of the inoculum
○ Length of inoculum
○ Length of incubation
○ Metabolic activity of organisms
○ Temperature of incubation

Dilution Methods
• Include:
○ Tube dilution method
○ Agar dilution method
• Used to measure MIC (minimal inhibitory concentration) and MBC (minimal bactericidal
concentration)
• Dilutions of an anti-microbial agent are added to a broth or agar medium
• A standardised inoculum is then added
• MIC - lowest concentration of anti-microbial required to prevent visible growth
• MBC - lowest concentration of antibiotic that kills 99.9% of the inoculum

E-Test
• A plastic strip impregnated w/ antibiotic
• Placed into agar inoculated w/ bacteria
• Diffusion into the media
• Continuous concentration gradient that yields a quantitative measurement of the MIC
value

Mechanism Specific Test

• Example
○ Beta-Lactamase Test

Bacteriology Page 265

Mycobacteria
Thursday, 27 April 2017 11:32 AM

Introduction
• >200 species of mycobacteria
• Major pathogenesis
○ Mycobacterium tuberculosis - Koch bacillus
○ Mycobacterium leprae - Hansen's bacillus
○ Rest are environmental organisms, collectively called MOTTs (Mycobacteria Other
Than Tuberculosis) or NTM (non-tuberculous mycobacteria) - cause opportunistic
infections

Physiology and Structure

• Name from Myces and Bakterion, fungus like rod
• Bacillus
• Aerobic
• Non-motile
• Acid-fast staining
• Mycolic acid in cell wall
• Complex cell wall
• Intracellular parasite
• Disease from immune response

Virulence Factors
• No spore, no flagellum, no endotoxin, no exotoxin, no invasive enzyme
• Capsule:
○ Polysaccharide
○ CR3
○ Enzyme
○ Protect
• Lipid/lipo arabinomannan
• Heat shock protein/tuberculin protein
○ Antigenicity

Lipid
• Phospholipid
○ Monocytes proliferate
○ Cause tubercles
• Wax D
• Sulfatide
○ Suppress phagosome
○ Prevents fusion of the lysosomes and phagosomes
• Cord factor (trehalose - 6,6-dimycolate)
○ Destroy mitochondria
○ Cause chronic granulomatosis
○ Suppress WBC

Cell Wall
• Complex
• Contains mycolic acid
○ Lipids account for 60% of cell wall weight
• Responsible for many characteristics
○ Acid fastness

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 ○ Acid fastness
○ Slow growth
○ Resistance to detergents
○ Antibiotic resistance
○ Antigenicity
○ Clumping

Mycobacterium tuberculosis
Pathogenesis
• Modes of infection
○ Droplet infection
 Person to person by inhalational aerosols
 M.tuberculosis (pulmonary tuberculosis)
○ Infected milk
 Infected cattle
 M.bovis (intestinal tuberculosis)
○ Contamination of abrasion
 Lab workers (skin infection)

Epidemiology
• 2 billion people infected worldwide
○ One-third of the population
• _

Pathogenesis
• The infectious bacilli are inhaled are droplets. It is taken up by the alveolar macrophage.
The macrophage will elicit signals by cytokines that call the other immune cells e.g.
lymphocytes to the sight. The macrophage does not destroy the bacteria but the bacteria
can live in the cell for up to 10 years. A granuloma can form
• The patient can remain w/o sniymptoms while the bacteria is still inside the granuloma
(latent TB)
○ Can lead to post-primary TB/Secondary/Adult-type TB
○ There can be reactivation when the immunity is compromised/suppressed
• During co-infection w/ HIV or immunocompromised
○ The immune cells get distracted and the bacilli are released from the granuloma and
causes active TB infection

Typical Progression of Pulmonary TB

• Pneumonia
• Granuloma formation w/ fibrosis
• Caseous necrosis
○ Tissue becomes dry and amorphous (resembling cheese)
○ Mixture of protein and fat (assimilated very slowly)
• Calcification
○ Calcium salts deposited
• Cavity formation
○ Centre liquefies and empties into the bronchi

Ghon Complex
• Nodules in the lung tissue and lymph nodes
• Caseous necrosis inside the nodules
• Calcium may deposit in the fatty area of the necrosis
• Visible on x-rays

Extra-pulmonary TB
• Bone and joint TB

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 • Bone and joint TB
• Tuberculous meningitis (TBM)
• Tuberculomas
• Abdominal TB
• Tuberculous lymphadenitis
• Urinary TB
• Genital tract TB
• Skin TB - scrofuloderma
• TB pericarditis
• TB laryngitis
• Ocular TB
• Adrenal gland TB - hyperadrenalism or Addison's Disease

Laboratory Diagnosis
• Specimen
○ Sputum (expectorated or induced)
 Quality
□ Take early morning sputum
□ The specimen is of good quality if acquired by induction
□ Bad quality if acquired by expectoration
○ Pleural fluid
○ Gastric washings
○ Urine
○ Aspirates
○ CSF
○ FNA
○ Tissue biopsies
• Staining
○ Ziehl Neelsen (ZN) stain - acid fast bacilli (AFBs)
○ Kinyoun staining
○ Fluorescence microscopy using Auramine O or Rhodamine stain
• Culture - gold standard in TB diagnosis - require incubation for 6-8 weeks before declaring
negative
○ Solid culture (Lowenstein Jensen, Middlebrook)
 M.tuberculosis
□ Are rough, thick, wrinkled
□ Have an irregular margins
□ Faintly buff-coloured
○ Semi automated Liquid culture Bactec 460
○ Automated Liquid culture system (MGIT - mycobacterial growth indicator tube)
○ No gold standard for diagnosis of latent TB infection
○ To confirm M. tuberculosis from culture
 Growth rate
 Colonial morphology
 Ziehl-Neelsen staining results
 Nitrate reductase test positive
 Niacin test positive
 Catalase at 68°C - negative
 Molecular
□ PCR - from culture; some direct from sputum
• Immunological diagnostic tests
○ Tubeculin skin test - does not distinguish between vaccination and disease. Usually
negative in patients w/ advanced AIDS
○ QuantiFERON, T-SPOT TB - detect interferon . For active and latent TB

Anti-TB Drugs

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Anti-TB Drugs
• First line
○ Isoniazid
○ Rifampicin/Rifabutin
○ Ethambutol
○ Pyrazinamide
○ Streptomycin
• Second line
○ Para-amino salicylic acid
○ Cycloserine
○ Quinolones (Ofloxacin/Ciprofloxacin/Levofloxacin etc.)
○ Amikacin
○ Kanamycin
○ Capreomycin
○ Ethionamide
• If you forget tour TB drugs you will need a PRIEST
○ P - pyrazinamide
○ R - Rifampicin
○ I - Isoniazid
○ E - Ethambutol
○ ST - Streptomycin

Drug Resistance
• Multidrug resistant TB (MDR TB)
○ TB that is resistant to at least Rifampicin and Isoniazid
• Extensively drug resistance TB (XDR TB)
○ TB which is resistance to isoniazid and Rifampicin, plus resistant to any
fluoroquinolone and at least one of three injectable second line drugs (i.e. amikacin,
kanamycin or capreomycin)

Prevention and Control

• BCG vaccine
• Respiratory isolation of persons w/ suspected TB till non-infectious
• Ventilation of household
• Contact tracing
• Nutrition
• Chemoprophylaxis
• Vitamin D?

Leprosy
M.leprae
• Obligate intracellular organism - can grow in the mouse pad or in the armadillo
• Reservoir - infected humans; low infectivity
• Transmission - skin to skin contact
• Grows best in the cooler parts of the body
○ The skin
○ The peripheral nerves
○ Anterior chamber of the eye
○ Upper respiratory tract
○ Testes
• Incubation period
○ Years or decades

Ridley and Jopling's Classification

Bacteriology Page 269

Ridley and Jopling's Classification
• Tuberculoid
○ Tuberculoid tuberculoid (TT)
○ Borderline Tuberculoid (BT)
• Lepromatous
○ Borderline borderline (BB)
○ Borderline lepromatous (BL)
○ Lepromatous lepromatous (LL)

Pathogenesis
• Leprosy is a chronic granulomatous disease that usually involves the skin, peripheral
nerves and nasal mucosa
• Incubation period is long and varies from 3-15 years
• Prolonged close contact w/ infected patient is necessary for transmission of the disease
• The principle target cell of the leprae bacilli are on the schwann cells
• -
• Schwann cell invasion
○ Major target of M. leprae
○ To access the cells, mycobacterium leprae_

Tuberculoid Leprosy
• Relatively few or no bacilli because of adequate cell mediated immunity
○ One or a few hypopigmentated macules or plaques
 Sharped demarcated and hypaesthesic
○ Devoid of normal skin organs (sweat glands and hair follicles)
 Are very dry and scaly
○ Nerves may be destroyed w/ loss of sensation and perspiration (Arthur's
Phenomenon)

Lepromatous Leprosy
• Lesions have a large number of bacilli even on skin that appears normal - poor CMI
○ Symmetrically distributed skin nodules w/ poorly defined margins
○ Nerve damage leads to loss of digits
○ Late manifestation include loss of eyebrows (initially the lateral margins only) and
eyelashes, and dry, scaling skin, particularly on the feet
○ Lepronin test negative

Lab Diagnosis
• Specimen collection
○ Skin lesion biopsy
○ Skin scrapping
○ Nasal exudate
• Microscopy
○ Ziehl Neelsen stained smear
 Acid fast bacilli, accumulated n intracellular encapsulated globular masses -
"leprosy globi" - in lepromatous leprosy
• Cultivation - not applicable

Treatment and Management

• Chemotherapy
• First line drugs are Rifampicin, Dapsone and Clofazimine
• The WHO recommends that if a patient tests positive in an acid-fast smear they should be
treated for multi-bacillary disease
• The patient's bacterial load decides the length of treatment (6-24 months)
• Second line drugs are Ofloxacin and Minocycline
• Triple-drug combinations have been used in cases where a patient only has a single lesion

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 • Triple-drug combinations have been used in cases where a patient only has a single lesion

Prevention of Leprosy
• Early diagnosis and treatment
• BCG vaccination ??
• Health awareness and active surveillance high endemic areas
• Field trials w/ different vaccines
○ BCG + killed leprae bacilli (ICRC bacillus) have not given conclusive results

Scrofula
• A cervical lymphadenitis
• Source
○ Environmental water sources
○ Human respiratory tract
• Chronic , painless mass in the neck - known as cold abscess
• In children
○ Agent - M. scrofulaceum
○ Treatment
 Surgery
 Usually resistant to anti0biotics
• In adults
○ Agent - M. tuberculosis
○ Treatment
 antiTB drugs

M.marinum
• Causes swimming pool or fish tank granuloma
• Lesions usually localised but may form secondary lesions along dermal lymphatics -
sporotrichoid spread
• Self-limiting:
○ Can use Minocycline, Cotrimoxazole or Rifampicin w/ Ethambutol

M.ulcerans
• Causes Buruli ulcer
• Found in low-lying marshy areas subject to periodic flooding
• Treatment
○ Early, pre-ulcerative lesion
 Excision and primary closure
○ Ulcerated lesions
 Excision and grafting
○ Antimicrobial agents
 Variable results
• Toxin produced → tissue necrosis

Conclusion
• Descibe the structure and main pathologic features of M. tuberculosis
• What is extra-pulmonary TB
• Latent TB and reactivation
• MDR and XDR TB
○ What are they
○ How have they come about
○ How have they spread
• What is the connection between TB and AIDS
• Different classification of leprosy

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Rickettsiae
Friday, 9 June 2017 6:46 AM

General Characteristics
• Small Gram-negative coccobacilli
• Aerobic
• Obligate/facultative intracellular bacteria
○ This is because they are unable to produce the following and hence depend on the
host cells
 CoA
 NAD+
□ Important for bacteria growth and replication
• Divide by binary transverse fission
• Lack flagella
• Cause febrile illnesses - prodromal signs
○ Most characteristic Is vasculidities and is associated with rash
• Do not stain well w/ gram stain
○ Stained by Giemsa - take on a characteristic red colour
• Cause zoonoses
• Natural hosts are mammals and arthropods and are usually transmitted to humans by
arthropods
• Emerging and re-emerging infections

Classification
• Family
○ Rickettsiaceae
• Tribe
○ Rickettsiae
• Genera
○ Rickettsia
○ Orientia
○ Ehrlichia
○ Anaplasma
○ Coxiella
○ Bartonella

Transmission
• Most rickettsial pathogens transmitted by ectoparasites such as fleas, lice, mites and ticks
during feeding
• Scratching crushed arthropods or infectious faeces into the skin
• Inhaling dust or inoculating conjunctiva w/ infectious material
• Transfusion or organ transplantation
○ Rare but has been reported

Epidemiology
• Widely distributed globally, in endemic foci, w/ sporadic and often seasonal outbreaks
• All age groups are at risk for rickettsial infections during travel to endemic areas
• Transmission is increased during outdoor activities
• Infection can occur throughout the year

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• Infection can occur throughout the year
• 5-14 day incubation period for most rickettsial diseases

• Weil-Felix Agglutination Tests are test done to test for Rickettsiae infection
○ For cross-reactivity between Rickettsia spp. And Proteus bulgaris
○ Not specific or sensitive
• They are generally susceptible to Doxycycline and other Tetracyclines

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Rickettsia
Friday, 9 June 2017 6:51 AM

Rickettsia
Classification
• Classified into:
○ Spotted Fever Group (SFG) rickettsia
○ Typhus group (TG) rickettsia

Spotted Fever Group

• Rocky mountain Spotted Fever (RMSF) caused by Rickettsia rickettsii
• Rickettsial pox, caused by Rickettsia akari
• Boutonneuse Fever (i.e. Kenya tick-bite fever, African tick typhus, Mediterranean spotted
fever, Indian tick typhus) caused by Rickettsia coronii
○ African tick-bite fever is the most frequently reported travel-associated rickettsiosis
○ The tick is of the Dermacentor group

Typhus Group
• Epidemic typhus caused by Rickettsia prowazekii
• Murine (endemic or flea borne) typhus caused by Rickettsia typhi

Transmission
• Infection occur in 3 situations
○ Louse-transmitted epidemics
 Transmitted from person-to-person and tends to occur in areas of overcrowding
○ Reactivation
 Of a long-standing latent infection
 Primary infection occurs in childhood
○ Zoonosis
 From the flying squirrel

Pathogenesis
• The Louse takes a dump on the skin and when the patient scratches, the bacteria is
introduced into the body. But the tick directly introduces the pathogen by direct bite
• Adhere and invade the endothelial lining of the vasculature within the various organs
affected
○ The outer membrane proteins
 Allow the rickettsia to be phagocytosed into the host cells
○ To avoid phagocytosis
 Secrete phospholipase D and haemolysin C, which disrupt the phagosomal
membrane
• Once inside, the rickettsial organisms either multiply and accumulate in large numbers
• Exit the cell by lysing the host cell or they escape from the cell, damaging its membrane

Pathophysiologic Effect
• Increased vascular permeability w/ consequent oedema
• Loss of blood volume
• Hypoalbuminaemia
• Decreased osmotic pressure
• Hypotension

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 • Hypotension
• DIC - rare

Clinical Manifestations
• Early diagnosis
○ Tick bite or exposure
○ Recent travel to endemic areas
Similar illness in family members, co-workers or family pets (especially dogs)
• Spotted Fever
○ Onset 2-4 days after infective bite
○ Fever, severe headache, muscle pain and rash
○ Rash typically appears in ankles and wrist
○ Complications
 Partial paralysis of lower limbs
 Gangrene
• Typhus Fever
○ Fever
○ Chill
○ Headache
○ Myalgia and arthralgia
○ Macules which appear the trunk then to the extremities
 Hands, feet and head are spared in Prowazekii infection
○ Conjunctival injection
○ Pneumonia
○ Encephalitis with dizziness and confusion

Laboratory Diagnosis
• Stain poorly w/ gram-negative stain
• Giemsa/gimenez stain
• Serology
○ Weil-Felix Reaction - non-specific and non-sensitive
○ IFA - gold standard
○ Latex agglutination

Direst fluorescence antibody testing

Culture in embryonated eggs and tissue cultures
***risk of laboratory infections is very high

Treatment
• Doxycycline
• Chloramphenicol

Prevention and Control

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Orienta
Friday, 9 June 2017 6:49 AM

Orienta Tsutsugamushi
• Also known as tsutsugamushi disease
• Transmission is through the mite

Clinical Manifestations
• Illness varied from mild to self-limiting to fatal
• Fever, headache, myalgia, cough and GI symptoms
• Primary papular lesion → flat black eschar
• Associated w/ regional and generalised lymphadenopathy
• Haemorrhaging and intravascular coagulation
• Splenomegaly and lymphadenopathies are typical signs
• If untreated, meningoencephalitis

Treatment
• Doxycycline - DOC
• Chloramphenicol
• Combination of Doxycycline and Rifampicin
○ After infection there is non-sterile immunity

Prevention
• Protective clothing
• Insect repellents
• Avoid sitting or lying on bare ground or grass
• Clearing of vegetation and chemical treatment of soil

Control
• Case identification
• Public education
• Rodent control

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Coxiella
Friday, 9 June 2017 6:50 AM

Coxiella
Coxiella burnetti
• Order
○ Legionalleles
• Family
○ Coxiellaceae
• Genus
○ Coxiella
• Small (0.2-0.7 µm in diameter)
• Pleomorphic, gram-negative coccobacilli
• Stain poorly w/ Gram's stain - Giemsa stain used
• Obligate intracellular bacteria that live inside acidic lysosomes
• Causes Q-fever
• 2 morphologic variants
○ Small cell variant (SCV) spore like and can survive harsh environmental conditions
○ Large cell variant (LCV) multiplies in the host monocytes and macrophages

Epidemiology
• Various hosts
○ Mammals, birds, numerous different genera of ticks
○ Farm animals are the primary reservoirs for human diseases
• Excreted in urine, milk, faeces and birth products
• Birth products containing high numbers of bacteria
• Human infection occurs after:
○ Inhalation or skin contact
○ Exposure to placenta of an infected woman
○ Blood transfusion
• Due to its high infectivity and low infective dose, has been used as an agent of
bioterrorism

Pathogenesis
• Attaches to host macrophages using ankyrin
• Passive entry into the phagosome
• Delays fusion of the phagosome w/ lysosomes to transform from SCV to LCV
• Intracellular multiplication is initialised in the phagolysosome
• Proliferation of organisms within phagolysosome
• Rupture of the host cell
• Infected macrophages transported systemically
• RES (liver, spleen, bone marrow) most heavily infected

Clinical Manifestations
• Can be acute or chronic
• Acute Disease
○ Long incubation period ; average 20 days
○ Sudden onset of severe headache, high fever, chills, myalgia
○ Respiratory symptoms are generally mild
○ Hepatosplenomegaly present in ~half of cases
• Chronic Disease

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 •
○ Most common presentation is infective endocarditis, generally on a prosthetic or
previously damaged heart valve

Laboratory Diagnosis
• Serology - Most commonly used diagnostic test
○ Indirect immunofluorescent-antibody (IFA) test
 Test of choice
• Tissue culture
• PCR

Treatment
• Acute infections
○ Tetracyclines e.g. Doxyxline
• Chronic infections
○ Treatment for a prolonged period w/ a bacterial combination of drugs e.g. Rifampin
and either Doxycycline of Sulphamethoxazole-Trimethoprim

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Anaerobic Bacteria
Thursday, 4 May 2017 11:11 AM

• Growth and viability are determined by the state of oxidation of the environment they are in
○ Expressed in terms of oxidation-reduction or Redox potential
• Require negative redox potential for metabolism
• Exposure to oxygen leads to biochemical reactions w/ by-products which are toxic to the
organism
○ Toxic by-products include
 Negatively charged superoxide radical O2
 Hydrogen peroxide
• Obtain energy through fermentative pathways
• Lack
○ Cytochrome system
○ Enzymes
 Catalase, peroxidase and superoxide dismutase present in strict aerobes and
facultative anaerobes
 Superoxide dismutase catalyses the conversion of superoxide radicals to
hydrogen peroxide and water
 Catalase breaks down hydrogen peroxide to water and oxygen
• Vary in oxygen tolerance although all are classified as strict or obligate anaerobes

Classification and Examples of Anaerobic Bacteria Associated w/ Humans

• Methods of classification similar to those applied to bacteria in general
• According to
○ Shape
○ Gram stain
○ Spore-formation
• Bacilli
○ Gram positive non-spore forming
 Include genera
□ Actinomyces
□ Eubacterium
□ Bifidobacterium
□ Proprionibacterium
○ Gram positive spore-former
 Clostridium
○ Gram negative non-spore formers
 Bacteroides
 Prevotella
 Fusobacterium
• Cocci
○ Gram positive include genera
 Peptococci
 Peptostrptococcus
○ Gram negative
 Genus Veillonella
• Spirochetes
○ Specific strains of Treponema and Borrelia

Occurrence of Strict or Obligate Anaerobes

• Generally
○ GIT of animals
○ Environment: Soil, water, sewage

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 ○ Environment: Soil, water, sewage
• In humans
○ Normal flora in the
 Mouth
 Intestine particularly in the colon
 Lower parts of the GUT
 Selected areas on the skin
○ In various tissues association w/ disease

Growth of Strict Anaerobes in the Lab Specimens

• Most commonly isolated from specimens from
○ Various infected internal tissues and cavities
 Aspirates or fluids from lungs
 Discharge or pus from abscesses
○ Infected tissues or fluids from wounds contaminated w/ dirt including soil
• Collected, transported and processed applying the necessary care to increase the
chances of isolation by minimising chances of contamination and exposure to air

Lab Techniques for Detection of Strict Anaerobes

• Staining and microscopy
○ Gram's stain and microscopy for most of the specimens
• Gas liquid chromatography of fluids for detection of products of metabolism including
○ Butyric acid
○ Proprionic acid
○ Oxybutyric acid
• Culture techniques
○ Variety of methods improvised or designed to exclude air or oxygen from growth
environment

Culture Media for Isolation of Strict Anaerobes

• Blood Agar (BA) suitable for most pathogens
• Selective medium used where necessary
○ BA containing antibiotics
• Robertson's Cooked Meat Medium (RCM)
○ Used for initial inoculation of specimens which may contain many anaerobic bacteria
in small numbers to enhance their multiplication
 Inoculated and incubated for 72 hour then the sub-cultures are made on BA
and incubated appropriately
• Thioglycollate medium contains reducing agent, Sodium thioglycollate

Incubation of Anaerobic Cultures

• Anaerobic Jar Techniques
○ Fildes-McIntosh or McIntosh-Fildes Jar
 Evacuation of air and replacement w/ inert gases which can be a mixture of
hydrogen, nitrogen and CO2
○ GasPak or Oxoid jars
 Chemicals in gas generating kits react w/ water to generate hydrogen and CO2
and other gases
○ In the above, traces of oxygen in the jar is removed by a reaction w/ hydrogen
catalysed by palladium to form water
• Other culture methods

Gram Negative Non-Spore Forming Anaerobic Bacilli

• Several genera bacteria encountered as normal flora and potential pathogen in human
○ Colon
○ Mouth

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 ○
○ Lower GUT
• Include
○ Bacteroides
○ Porphyromonas
○ Prevotella
○ All belonged to the genus Bacteroides
○ Reclassified based on differences in biological and biochemical characteristics
○ Species are morphologically similar

Bacteroides
• Bacteroides spp are normal inhabitants of the bowels and other sites
• Non-motile
• Capsulated gram negative bacilli
• Species are several and include Bacteroides fragilis
• Pathogenicity Properties
○ Pathogenic species possess virulence factors including
 Polysaccharide capsule
 Pilli
 Enzymes
○ Cause various conditions in association w/ precipitating factors which can be
 Trauma involving colon
 Devitalised tissue and impairment of blood supply
 Facultative anaerobes or other strict anaerobes in a lesion

Fusobacterium
• Pleomorphic gram negative anaerobic bacilli
• Non-motile
• Species include
○ F.nucleatum
○ F.necroforum
• Significance
○ Normal flora particularly in the mouth
○ Causes human infections
 With other organisms in mixed infection or as a single causative agent
 F.nucleatum is one of the causative agents
• Susceptible to several antibiotics including
○ Penicillin
○ Metronidazole
○ Clindamycin

Anaerobic Cocci
• Gram Positive Anaerobic Cocci
○ Normal flora w/ other anaerobic bacteria
○ Various genera including
 Peptostreptococcus
○ Differentiated by biochemical tests and metabolic end products
○ Occasionally isolated from abscesses together w/ other anaerobes
• Gram negative Anaerobic Cocci
○ Veillonella spp. - V. pervula
○ Normal flora w/ other strict anaerobes
○ Occasionally isolated from specimens in mixed infections w/ no pathogenic role

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Clostridium
Thursday, 8 June 2017 4:40 AM

Clostridium
• Most species are flagellated and motile
• Form spores
○ Diameter larger than the width of the cell
○ Centrally or terminally or terminally placed in the bacillus
○ Resistant to adverse physical conditions
 Enable prolonged survival of the organism in the environment
• Defined by
○ Endospores
○ Strict anaerobes
○ Gram-positive structure
○ Inability to reduce sulphate to sulphite

Species of Clostridium
• Normal flora in the GIT of animals including humans and saprophytes in
○ Soil
○ Water
○ Decomposing animals and plants
• Species associated w/ diseases in humans include:
○ Perfringens or welchii
○ Botulinum
○ Tetani
○ Difficile
○ Histolyticum
• Species which may be isolated from specimens but not involved in disease causation - C.
sporogenes

Main Pathogenicity Properties of Clostridium Species

• Ability to form spores
○ Spores increase the chances of survival and enhance transmission
• Viability and multiplication in tissues w/ lack of or reduced oxygen supply
• Ability to produce exotoxins
○ Main virulence factors responsible for major clinical manifestations of associated diseases
○ Differ in composition and mechanisms of activity
• Specific pathogenic species produce aggressins and other enzymes

Biochemical Properties of Clostridium spp.

• Major biochemical metabolic processes are:
○ Ability to breakdown CHO referred to as saccharolytic activity
○ Ability to break down proteins - Proteolytic ability
○ Both occur in each species with either one dominating
• Other biochemical properties according to specific species

Clostridium perfringens or welchii

• Contaminated the environment via faeces from humans/other animals

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 • Contaminated the environment via faeces from humans/other animals
• Spores are
○ Oval and centrally placed
○ Formed in the intestine and environment
• Capsulated in tissue
• Non-motile
• Biochemical properties
○ Predominantly saccharolytic
○ Mild proteolytic
○ Nitrate reduction test positive

Antigenic Properties
• Releases different antigens designated A, B, C, D, E
• Classification into serotypes A, B,C, D, E

Exotoxins
• Several types
• Majority are released in infected tissue
○ Responsible for tissue damage and severity of disease manifestations
○ Each serotype releases more than one
○ Each type of toxin
 Can be released by different serotypes
 Has a specific MoA
• (alpha) toxin
○ Released by all serotypes
 Major toxin of C. perfringens serotype A
 Relatively heat stable
 Enzyme phospholipase or lecithinase
□ Splits lecithin which is a constituent of cell membranes
 Causes lysis of RBCs, myocytes, platelets, fibroblasts and leucocytes
 May decrease cardiac inotropy, trigger histamine release, platelet aggregation and thrombus
formation.
• Enterotoxin
○ Released by specific strains especially when grown in meat dishes
○ It is a protein that may be a non-essential part of the spore coat.
○ It induces intense diarrhoea in 7-30 hours.

• Theta toxin
○ Causes direct vascular injury, cytolysis, haemolysis, leucocyte degeneration and polymorphonuclear
cell destruction
○ The effect on leucocytes may explain the relatively low immune response of patients with tissue
clostridial infection.

• Kappa toxin
○ A collagenase that facilitates the rapid spread of necrosis through tissue planes by destroying
connective tissue

• Other virulence products C. perfringens

○ Include hyaluronidase
 Digests collagen of subcutaneous tissue and muscle
○ DNase

Pathogenesis

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 Pathogenesis
• Infections are characterised by low levels of inflammation in response to organism-associated exotoxins.
• It is more a response to the exotoxins than a classic immune response to invading tissue
• Purulence is often absent
• The process of myonecrosis spreads extremely fast and results in systemic toxicity and shock that can be fatal in 12
hours
• Infections requires 2 conditions to coexist
○ Organisms must be inoculated into the tissues
○ Oxygen tension must be low enough for the organisms to proliferate
• Exotoxins appear to be tissue-destructive soluble antigens released by clostridia. The exact mechanisms behind this
are as yet unknown.

Clinical Implications of C. perfringens

• Associated conditions include:
○ Gas gangrene
○ Wound infection
○ Food related gastroenteritis/food poisoning
○ Infections associated w. reproduction in females
○ Intra-abdominal infections

Gas Gangrene
• Condition characterised by:
○ Rapidly spreading swelling
○ Necrosis if tissues
○ Myositis
○ Gangrene
○ Gas production
• Commonly encountered in humans in association w/ traumatic injury involving skeletal muscle
• Development facilitated by
○ Injury, resulting in
 An open wound and damaged skeletal muscle exposed to the environment and
contaminants
□ Contaminating agents contains spores of causative agents which are mostly of
species clostridium
 Most commonly C. perfringens and specifically serotype A
 Others are C. septicum, histolyticum and novyi
 Impairment of normal blood flow
□ Leads to reduction in oxygen tension in the injured part
 Devitalised tissues
□ Create a suitable condition for viability and multiplication of C. perfringens
○ Spores germinate into bacilli which multiply and release exotoxins which act on skeletal muscle
 Causing destruction with gas formation
 Alpha toxin is the main cause of associated tissue damage and manifestations
□ It’s a lecithinase, which cleaves lecithin, which is a phospholipid, and causes
damage to the cell membrane
□ When red cells are affected, haemolysis ensues
 Since it does this in vivo, it can also cause haemolysis in vitro and when
cultured on BA, it creates 2 zones of haemolysis
○ Abdominal injury involving the large intestine associated w/ leakage of intestinal contents from
the lumen also predisposes to gas gangrene

Food Related Gastroenteritis/Food Poisoning

• Associated w/ food contaminated by specific strains of C. perfringens which form spores that survive
boiling for several hours
○ Spores germinate into bacilli in food

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 ○ Spores germinate into bacilli in food
○ Bacilli ingested as food is eaten
○ Bacilli produce enterotoxin in the intestines as they form spores
• Manifests as mild abdominal cramps and diarrhoea 8-12h hours after ingestion
○ Fever and vomiting are not common
○ Illness subsides within 24-48 hours
○ Slow onset diarrhoea because the disease is caused by ingestion of a large amounts of the
spores and then germinate and the bacilli to produce the toxin
• Isolation and identification of the specific strains from stool and suspected food may be useful in
laboratory confirmation where necessary

Infection Associated w/ Female Reproduction

• Can complicate child birth or traumatic abortion
• Organisms may originate from large intestines via faecal material and contaminated perineal skin
○ Transmitted to devitalised tissues in reproductive system
• Can also arise from contaminated equipment or other items used
• Infection can spread to adjacent tissues

Intra-Abdominal Infection
• Encountered mostly as a result of local spread or organisms from intestinal lumen to adjacent
tissues in association w/ other abnormalities

Wound Infection
• C.perfringens occasionally causes wound infection or inflammatory processes in soft tissues w/o
involvement of muscle
○ Different condition from gas gangrene
• Large amounts of flesh is exposed to dirt and soil

Myositis
• Production of pus without myonecrosis

Lab Investigation of gas gangrene and other Perfringens infections

• Specimen from infected sites include swabs or fluid from wounds or necrotic tissue
• Lab procedure
○ Gram stain for gram positive bacilli
○ Culture for isolation and identification
 Media
□ RCM
 Inoculated w/ a portion of specimen
 Incubated under anaerobic conditions for~2-3 days
 Subcultures made on BA
□ BA
 Incubation as described for anaerobes
□ At temperature of 35-37°C for 48 hours
 Colonies of C. perfringens of BA
□ Relatively large translucent smooth surface
□ 2 zones of haemolysis of double zone haemolysis
 Inner zone shows complete haemolysis and larger outer zone of incomplete
haemolysis
□ Microscopy shoes gram positive bacilli
□ Spore stain after prolonged incubation
 Demonstration of alpha toxin production as a presumptive identification test by various
methods including Nagler test/reaction

Management of Gas Gangrene

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Management of Gas Gangrene
• Non-specific supportive measures
• Specific
○ Mainly
 Surgical management of wounds
 Antibiotic treatment
○ Effective agents include penicillin and metronidazole
 IV Penicillin G

Prevention of Gas Gangrene

• Proper care of wounds in general by cleaning and removal of foreign material and dead tissue
• Proper cleaning of skin before invasive procedures and use of sterilised instruments and materials
• Antibiotic prophylaxis in management of fresh or trauma associated wounds

Clostridium tetani
• Flagellated and motile
• Spores are
○ Spherical, placed at one end of the bacillus - terminal spore
○ Characteristic microscopic morphology shoes bacillus w/ distension at one end
 Drumstick appearance
• Contamination of the environment occurs via faeces from humans and other animals e.g. horse,
donkeys

Physical Characteristics
• Depend on the strain
○ Some strains form spores which are readily inactivated
○ Other strains form spores which are highly resistant to physical conditions including
 Commonly used disinfectants
 Adverse environmental conditions
□ Capable for survival for years in environment
 Boiling in water for more than 3 hours
 Dry heating at 150°C for an hour

Biochemical Properties
• Predominantly proteolytic

Exotoxins
• Tetanolysin
○ Causes lysis of RBCs
• Tetanospasmin
○ Neurotoxin
 Cleaved by a bacterial protease into 2 peptides linked by a disulphide bond
 The toxin initially binds to receptors on presynaptic membranes of motor neurons
 It then migrates by retrograde axonal transport to the cell bodies of these neurons to the
spinal cord and brainstem
 The toxins diffuse to terminals of inhibitory cells, including both glycinergic interneurons
and GABA-secreting neurons from the brainstem
 The toxin degrades synaptobrevin, a protein required for the docking of neurotransmitter
vesicles on the presynaptic membrane
 Release of GABA is blocked and the motor neurons are not inhibited
 Hyperflexia, muscle spasms and spastic paralysis is the result

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  Hyperflexia, muscle spasms and spastic paralysis is the result
 Extremely small doses of toxin are lethal for humans.
○ Pathogenicity component responsible for disease manifestation in humans
○ Convertible to tetanus toxoid

Pathogenesis
• Infection occurs through contamination of wounds by material containing spores
○ Soil
○ Dirty clothes
○ Dust
○ Contaminated instruments
• Spores germinate into bacilli facilitated by reduce oxygen and devitalised tissues in the wound
• C.tetani remains localised in the infected part
• Infection of the umbilical wound of a new born infant results in neonatal tetanus
• Bacilli in wound releases tetanospasmin
○ Diffuses onto the blood circulatory systems and reaches nervous system
○ Passes through nerve trunks to CNS
 Travels retrograde through the motor axons to the spinal cord
○ Prevent affected neurons from releasing inhibitory mediator in the motor neuron synapses
resulting in over-activity of the muscles in response to stimuli
 Tetanospasmin acts as a protease and cleaves SNARE proteins. By cleaving the SNARE
proteins, it inhibits exocytosis of GABA and Glycine (which inhibit motor neurons) from
Renshaw cells (inhibitory interneurons of the spinal cord that connect to alpha motor
neurons, which sense over-activity from nearby neurons and if there is over-activity,
releases inhibitory neurotransmitters), which results in uncontrolled firing of the neurons,
leading to spasms

Clinical Implications
• Causative agent of tetanus
• The incubation period may range from 4-5 days to as many weeks
• The disease is characterised by tonic contraction of voluntary muscles
• Muscular spasms often involve first the area of injury and infection and then the muscles of the jaw
(trismus, lockjaw) which contract so that the mouth cannot be opened
• Gradually, after voluntary muscles become involved resulting in tonic spasms
• Any external stimulus may precipitate a tetanic generalised muscle spasm
• The patient is fully conscious and pain may be intense
• Death usually results from interference with the mechanics of respiration
• The mortality rate in generalised tetanus is very high.
• Overall effects are generalised strong sustained contractions of muscles or spasms and increased
reflex responses to stimuli
○ Risus sardonicus
○ Risus - grin
○ Sardonicus - malevolent/evil
○ Evil grin or lock-jaw symptoms
□ Tense masseters prevent the jaw from opening
○ Opisthotonus
○ Characteristic extension and arching of the back due to powerful spasms of the back
muscles
○ Respiratory failure

Lab Investigation of Tetanus

• Specimens and exam procedures c.f. perfringens
• Colonies of C. tetani
○ Grows as a layer on the surface of BA
○ Gram stain

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 ○ Gram stain
 Gram positive bacilli
□ Drumstick appearance after prolonged incubation
 Spore stain for spores after a long incubation

Management of Tetanus
• Specific
○ Administration of tetanus antitoxin
 Toxoid
□ The toxin is conjugated to a protein (tetanus immune globulin), to increase
immunogenicity
 This produces an antibody response to the toxin and NOT the organism
○ Antimicrobial agents
 C.tetani susceptible to penicillin G
• Other methods
○ General supportive care
 Resp. failure
○ Muscle relaxants
○ Surgical management of wounds where required

Prevention
• Methods as for C. perfringens
• Immunisation w/ tetanus toxoid
○ As part of management of traumatic wounds
○ During antenatal care for expectant mother
 Usually 2 doses but up to 4 can be given
 Usually started after 20 weeks
 2nd dose 4 weeks after the first
 If the woman has had 1-4 doses before, give one dose before delivery.
 If the pregnancies are far apart, treat the mother like it's the first time
○ In patients undergoing recovery from tetanus
○ Part of routine immunisation in children
• Use of sterile equipment and aseptic methods in wound management, child delivery and care of the
umbilical wound in the new-born

Clostridium botulinum
• Flagellated and motile
• Spores are placed centrally or sub-terminally on the bacillus
• Encountered in the environment as saprophytes
○ Source of contamination of various items

Physical characteristics of spores

• Withstand moist heat at 100°C
• Destroyed at 120°C in 5 minutes

Antigenic properties of C. botulinum

• Different types of antigens identified and designated alphabetical letters A-G
• Basis for serotyping into serotypes A-G
○ Each serotype produces antigenically distinct exotoxin
• Types A, B, E and F are the principle causes of human illness
○ Types A and B have been associated with various foods
○ Type E primarily with fish products

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 ○ Type E primarily with fish products
○ Type C causes limberneck in birds
○ Type D causes botulism in animals
○ Type G is not associated with disease

• Exotoxin
○ Referred to as botulinum toxin which is a neurotoxin significant in disease manifestation
 Destroyed in heat at 80°C for 30-40 minutes
□ Inactivated by proper heating of food which may contain the toxin
○ A 150,000-MW protein cleaved into a 100,000-MW and 50,000-MW proteins linked by a
disulphide bond
○ Botulinum toxin is absorbed from the gut and binds to receptors of presynaptic membranes of
the motor neurons of the PNS and cranial nerves
○ Proteolysis, by the light chain of botulinum toxin, of the target SNARE proteins in the neurons
inhibits the release of Ach at the synapse, resulting in lack of muscle contraction and paralysis
 The SNARE proteins are synaptobrevin, SNAP 25 and syntaxin
 Toxins of type A and E cleave the 25,000-MW SNAP 25
 Type B toxin cleaves synaptobrevin
○ Cleaves the SNARE protein, which prevents the fusion of vesicles with the presynaptic nerve
terminal
○ The only difference with tetanospasmin is the target. Botulinum toxin affects Acetylcholine
whose release is responsible for muscle contraction, and if it is inhibited, muscles remain
flaccid

Medical Significance
• Causative agent of Botulism
○ Food-borne
 More frequently encountered than other types
 Severe form of food poisoning
 Develop when botulinum spores
□ Contaminate food which may be improperly
 Cooked
 Canned
 Preserved
□ Germinate in to bacilli and multiply and release toxin in food
□ Release toxin when ingested in food
○ Wound
 Secondary to wound infection
 Relatively less common than food-borne type
 Spores contaminate wound and germinate into bacilli that release the exotoxin
• Transmission is different in babies
○ Adults do not get infected with the bacteria when we ingest the spores (because C. Botulinum
is a wimp :) ) because, even though there is an anaerobic environment, the GIT bacterial flora
outcompete it and prevent germination of spores
○ Babies, who lack this robust flora, have the perfect environment for the spores to germinate,
without competition and the spores germinate and the bacilli release the toxin. They ingest the
spores from honey (since C. botulinum spores are more commonly found in honey)
• Manifestation
○ Symptoms develop 18-24h after ingestion of the toxic food with:
 Flaccid paralysis - affects the PNS not the CNS as the toxin is unable to cross BBB
□ Known as "Floppy Baby Syndrome" in babies
○ Caused by the absence of muscle contraction
○ Is a descending paralysis
 Starts superiorly and ends inferiorly
 It might start with eye problems (diplopia, ptosis)
○ Visual disturbances due to incoordination of the eyes muscles and double vision

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 ○ Visual disturbances due to incoordination of the eyes muscles and double vision
○ Inability to swallow and,
○ Speech difficulty
 Signs of bulbar paralysis are progressive and death occurs from respiratory paralysis or
cardiac arrest
 GIT symptoms are not regularly prominent
 No fever
 Patient remains fully conscious until shortly before death
 High mortality
 Patients who recover do no develop antitoxin in the blood.

Lab Investigation
• Specimen include
○ Stool and other intestinal tract content
○ Suspected food
• Procedures
○ Gram's Stain and microscopy to show gram positive bacilli
• Culture for isolation and identification
○ By anaerobic culture methods as applied to other anaerobes
• Detection of toxin in
○ Blood
○ Leftover food
○ Stool

Management of Botulism
• Administration of antigenically specific antitoxin
○ Trivalent antitoxin made from horse serum
 Test for allergy
• Wound botulism requires wound management as well as
○ Surgical methods
○ Antimicrobial agents mostly penicillin
• Guanidine hydrochloride

Prevention
• Methods applied include
○ Proper processing and canning of food
○ Sufficient heating of food before consumption
○ Proper use of food preservatives

Clostridium difficile
• Transmitted faecal-orally
○ Hands of medical personnel is the main intermediary
• More resistant to commonly used antibiotics than other flora in the colon
○ Like botulinum, it does not do well with competition with other flora
○ Majority of bacterial flora are susceptible to commonly used antibiotics
 Most patients in a hospital environment, are taking antibiotics
○ It’s the major pathogen in nosocomial diarrhoea
 The most commonly associated antibiotic with C. difficile infection is Clindamycin
○ Can also be associated with third generation cephalosporins
○ This therefore leaves a conducive environment for C. difficile spores acquired, to germinate
and the bacilli produce the toxin that bring out the symptoms
• Broad spectrum antibiotics administered for other infections inhibit the susceptible organisms

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• Broad spectrum antibiotics administered for other infections inhibit the susceptible organisms
• C.difficile is not inhibited and is able to multiply
○ Produce toxins (Exotoxin A and B)
 Enterotoxin (Exotoxin A)
○ Binds to the brush border of the intestinal wall and causes inflammation, death of
the cells and this leads to diarrhoea
 Cytotoxin (Exotoxin B)
○ Disrupts cytoskeletal structure by depolarising actin which leads to enterocyte death
and necrosis
○ Also forms a yellow-is grey exudate that forms a membrane, that covers the colonic
mucosa (Pseudomembranous colitis)
• Effects of the toxin resulting antibiotic associated diarrhoeal disease including pseudomembranous
colitis
• Clinical findings
○ Diarrhoea
 Non-bloody
 Neutrophils in the stool in about half of the patients
 To differentiate diarrhoea caused by C. difficile and other pathogens
○ Directly visualise the pseudomembranes directly either histologically or
endoscopically
○ Look for the toxin in stool through various assays e.g. PCR , not the bacteria,
because the GIT is normally colonised by the bacteria and may not be the strain
that produces toxin
○ Pseudomembranous colitis
○ Fever and abdominal pain
• Lab diagnosis
○ Insufficient to culture since it is part of the normal flora
○ ELISA
 Antibody to the toxin
 Rapid but not as sensitive as cytotoxicity test
○ Cytotoxicity test
 Human cells in culture are exposed to the exotoxin and death of the cells is observed
 To distinguish between cytotoxicity caused by the toxin and that by a virus, antibodies to
the toxin are used to neutralise the effect
 More sensitive but requites 1-2 days to incubate
○ PCR
 For presence of the toxin gene DNA is also used
• Treatment
○ Oral Vancomycin
 IV Vancomycin would not be helpful since the target is the organ of the intestines
 Plus, oral Vancomycin has poor PO absorption which is great in that there will be less AE
○ Metronidazole
 In cases for vancomycin-resistant enterococci
○ Fidaxomycin
 Treatment for pseudomembranous colitis and preventing relapse
○ Surgery in life-threatening conditions
• Prevention and Control
○ Thorough washing of hands with soapy water

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Enterobacteriaceae (E. coli, Proteus, Klebsiella, Citrobacter,
Enterobacter, Serratia)
Tuesday, 9 May 2017 3:26 PM

Taxonomical Classification Enterobacteriaceae

• Two important classifications
 Based on taxonomy
 Based on lactose fermentation
○ Kingdom: Bacteria
○ Phylum: Proteobacteria
○ Class: Gamma Proteobacteria
○ Order: Enterobacterialis
○ Family: Enterobacteriaceae

Characteristics
• All Enterobacreriaceae;
○ Gram negative
○ Rod shaped
○ Ferment glucose with acid production
○ Oxidative negative (lack cytochrome oxidase)
• Facultative anaerobes
• Most are catalase positive
• Non-capsulated except Klebsiella and some strains of E. coli
• Most are motile by a flagella (peritrichous) except Shigella and Klebsiella
• Grow in media containing bile salts (MacConkey agar)
• Most reduce nitrate to nitrite via nitrate reductase
• Most are normal part of the gut flora in intestines of human and other animals other found
in water and soil
• Also called coli forms or enteric bacilli

Classification Based on Lactose Fermentation

• Lactose fermenters
○ Escherichia coli
○ Klebsiella spp.
○ Citrobacter spp.
○ Enterobacter spp.
• Non-lactose fermenters
○ Salmonella spp.
○ Shigella spp.
○ Proteus spp.
○ Yersinia spp.
• Late lactose fermenters
○ Shigella sonnei
○ Edwardsiella
○ Serratia
○ Citrobacter
○ Arizona
○ Providencia
○ Erwinia

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 ○ Erwinia
• Note:
○ Media used distinguishes between LF and NLF
 MacConkey agar
 Salmonella Shigella (SS) agar
 Triple Sugar Iron (TSI) agar

Antigenic Structure
• Cell wall/Somatic or O antigen
○ A single organism may carry several O antigens
• Capsular/K antigen
○ Prevent O agglutination
○ Not all strains have a capsule
• Flagella/H antigen
○ Not all are motile

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Escherichia coli
Tuesday, 13 June 2017 5:29 AM

• Normal inhabitant of the GIT

• Produce infections in humans and animals
• Some strains cause various forms of gastroenteritis
• Is a major cause of UTI and neonatal meningitis and septicaemia
• Detection of E. coli in water indicates pollution and contamination

Characteristics
• Morphological characteristics
○ Gram negative rods
○ ± capsule
 K-antigen found on the capsule is useful for serotyping
○ Motile
• Culture characteristics
○ Lactose fermenters
 Grows pink on MacConkey's Agar
○ Pathogenic strains are haemolytic on blood agar
○ Facultative anaerobic
• Biochemical reactions
○ Indole positive, methyl red positive
○ TSI: acid butt/acid slant with gas production

Virulence Factors
• Surface Antigens and toxins
○ O antigen
 LPS
 Has endotoxic activity
 Protects the bacteria from phagocytosis
○ K antigen
 Protects against phagocytosis e.g. K1 has a strong association w/ virulence,
particularly meningitis in neonates
• Fimbriae used for attachment to the urogenital tract
• Exotoxins
○ Haemolysins and enterotoxins (LT = heat labile toxin, ST = heat stable toxin, VT =
serotoxin [also known as SLT = Shiga like toxin])
• Other adhesins
○ Intimin
 Associated w/ adhesion and effacement phenomenon which causes
destruction of intestinal surface cells
○ Outer membrane proteins

Pathogenesis
• Gastrointestinal - caused by:
○ ETEC (enterotoxigenic E.coli)
○ EPEC (enteropathogenic E.coli)
○ EIEC (Enteroinvasive E.coli)
○ EHEC (enterohaemorrhagic E.coli)
○ EAggEC (enteroaggregative E.coli)
• Extra-intestinal

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 • Extra-intestinal
○ UTI
○ Pyogenic infections
○ Septicaemia

ETEC
• Causes traveller's diarrhoea
• Common in persons from developed countries visiting endemic areas
• Causes secretory/watery diarrhoea indistinguishable from cholera
• Transmitted by faecal-oral route via contaminated food or water
• Produce 2 forms of exotoxins; heat-labile toxin (LT) or heat-stable toxin (ST) or both
• Infective dose is high - 106-10

Pathogenesis
• Adhere to small intestines by attaching to specific receptors using pili
• Bacteria then release toxins LT and ST that act on the cells, causing diarrhoea
• HL toxin. Each molecule has 1 A and 5 B subunits
• B attaches to GM1 ganglioside receptor of enterocyte, allowing entry of A subunit in
enterocyte
• Splits into A1 and A2 (A1 - active; A2 - linking)
• A2 allows only biologically active A1 to be linked with B. A1 activates adenylyl cyclase)
• This results in accumulation of cAMP
• There is then an outpouring of larger amount of water + electrolytes + carbonates +
potassium to intestinal lumen → non-inflammatory diarrhoea

Heat Labile Toxin (LT)

• Increases/activates cAMP
• Similar to the cholera toxin
○ Also binds to the GM1 ganglioside receptor

Heat Stable Toxin (ST)

• ST acts by activation of cGMP
• ST binds to a glycoprotein receptor → activation of membrane-bound guanylate cyclase →
increases cGMP → inhibition of intestinal fluid uptake → causes fluid accumulation in the
intestine - net fluid secretion

Clinical Features
• 1-2 days incubation
○ Vomiting
○ Abdominal cramps
○ Watery diarrhoea
○ Chills
• Self limiting
• Dehydration

EPEC
• Causes infantile and childhood diarrhoea
• Do not produce enterotoxins
• Non-invasive

Pathogenesis
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Pathogenesis
• Adhere to mucosal cells of upper part of small intestines
• Fimbrae penetrate between the microvilli of the brush border to firmly attach to the
enterocyte surface
• Results in loss of brush border microvilli - resulting in loss od large areas of nutrient
absorption
• Greater osmotic pressure in the lumen resulting in the loss of water from the enterocytes
to balance the osmotic pressure causing non-inflammatory diarrhoea

Clinical Manifestation
• Result in watery diarrhoea with mucus but no blood, vomiting or fever
• Usually self-limiting but may be chronic in some cases

EIEC
• Produces disease similar to shigellosis/dysentery
○ Passage of blood
○ Mucus
○ Leukocytes in stool
• Commonly affects children in developing countries and travellers
• Can be a cause of traveller's diarrhoea
• Pathogenesis is due to invasion into mucosal cells of the intestine, multiply inside the cells
and destruction/inflammation/ulceration of colon epithelium
• Characterised by:
○ Fever
○ Severe abdominal cramps
○ Malaise
○ Bloody mucoid diarrhoea

EAggEC
• Watery diarrhoea
• Pili implicated in adherence
• Causes persistent diarrhoea in children in developing countries
• Pathogenesis:
○ Shortening of villi
○ Mucus biofilm
○ Heat stable cytotoxin (haemorrhagic necrosis and oedema)
• Escherichia coli O104:H4 associated with outbreak in Germany in 2011

EHEC (VTEC or STEC)

• Produce Verocytotoxin (VT) or shiga like toxin
• Serotype O157:H7 is most commonly isolated
• Been described in industrialised countries, as resulting in outbreaks of food poisoning due
to undercooked beef or pork
• Cattle are reservoirs, and hamburger, unpasteurised milk, fruit juices and uncooked
vegetables are common sources of human infection
• Only E. coli that does not ferment sorbitol

Pathogenesis
• Attaches to the caecum and large bowel
• Release shiga-like toxin (verocytotoxins)
○ "A" subunit enzymatically cleaves adenine base from rRNA (28S subunit) → blocks
protein synthesis within the enterocytes → cytotoxic effects (cell death) resulting in

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 ○
protein synthesis within the enterocytes → cytotoxic effects (cell death) resulting in
inflammatory haemorrhagic colitis

Clinical Features
• Abdominal cramps, watery diarrhoea progresses to bloody
• No to minimal fever
• Haemolytic-uraemic syndrome (esp. in children under 10) is a complication of
○ Haemolytic anaemia
 Micro-angiopathic haemolytic anaemia
○ Thrombocytopenia
○ Acute renal failure
○ Pathogenesis
 Shiga-like toxin damages the endothelial lining of the capillaries in the
glomerulus.
 The damaged endothelial lining becomes thrombogenic, causing platelets to
aggregate and adhere. This platelet aggregation leads to a low platelet count
 The platelet aggregation, lyses RBCs as the pass through the capillaries, thus
the haemolysis

Adherent-Invasive E. coli (AIEC)

• Proliferate in hosts with defective innate immunity
• They are associated with the ileal mucosa in Crohn's Disease

Uropathogenic E. coli
• Utilise fimbriae to bind endothelial cells of urinary tract and colonise bladder
• Is the leading cause of UTIs which can lead to acute cystitis (bladder infection) and
pyelonephritis (kidney infection)
• Also produce alpha and beta haemolysins
○ Cause lysis of urinary tract cells
• Ability to form K antigen that contribute to biofilm formation
• The E. coli serotypes commonly responsible for UTI are those usually found in faeces
• UTI can be:
○ Ascending infection via urethra
 Faecal bacteria, spread up tract to bladder as well as kidneys or prostate
○ Descending infection: haemotogenous route:
 Rare
 E.coli enter upper urinary tract organs from blood stream

Meningitis
• Caused by different serogroups:
○ K I polysaccharide sialic acid
○ Usually with strains carrying the K capsular antigen

Pyogenic Infections
• E.coli form the most common cause of intra-abdominal infections such as peritonitis and
abscess resulting from spillage of bowel contents
• Pyogenic infections in the perianal area

Lab Diagnosis
• Specimens

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 • Specimens
○ Stool
○ Urine
○ Blood
○ CSF
• Gram stain (direct) [not for stool]
○ Gram negative rods, ± pus cells
• Culture - in air, 35-37°C, 18-24 hours
○ Blood agar - some strains are ß haemolytic
○ MacConkey, CLED - colonies are lactose fermenters
 CLED for urine
□ Turns yellow due to acid production
 McConkey for stool
○ Eosin Methylene Blue (EMB) agar
 Produces a metallic green sheen
• Gram stain of colonies
○ Ram negative rods
• Biochemical tests (IMViC tests ++--)
○ Indole test - positive
○ Methyl red test - positive
○ Voges-Proskauer test - negative
○ Citrate utilisation test - negative
• Serotyping
○ To distinguish the strains

Treatment
• Should be guided by in vitro susceptibility tests
○ Uncomplicated UTI
 Trimethoprim-Sulphamethoxazole/Nitrofurantoin/Ampicillin
 Norfloxacin
○ Nosocomial infection - usually multidrug resistant
 Cephalosporins/Carbapenems/Fluoroquinolones/Aminoglycosides
 Piperacillin-Tazobactam
 Rifamixin - ETEC

Control
• Health education
• Hygiene
• Treatment of infected persons

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Klebsiella
Tuesday, 13 June 2017 5:30 AM

Genus Klebsiella
• Nosocomial infections
• Have multi drug resistance
○ Treatment therefore includes carbapenems
• Commonly affect alcoholics
• They form abscesses
• They are contracted from aspirates

General Characteristics
• Gram negative bacilli (short and plump)
• Large polysaccharide capsule - colonies w/ mucoid appearance
• Non-motile
• Capsular halo seen prominently in gram stain

Klebsiella spp.
• Pneumoniae
○ Pneumonia (lobar)
• Oxytoca
○ Lobar pneumonia
• Oxaenae
○ Atrophic rhinitis (ozena)
• Rhinoscleromatis
○ Rhinoscleroma (destructive granuloma of the nose and pharynx)
• Granulomatis
○ Granuloma inguinale/donovanosis
 Painless, non-purulent genial
• _

Pathogenesis
Virulence Factors
• Capsular K antigen
○ Anti-phagocytic
• Synthesise siderophores
○ Taking up iron bound to host proteins
• Fimbriae
○ Adherence to respiratory and urinary epithelia
• Endotoxin

Clinical Presentation
• Pulmonary infections
○ Lobar pneumonia
 The patients cough up currant coloured, jelly sputum
• Bronchitis
• Bronchopneumonia
• Extra-pulmonary infections
○ Meningitis and enteritis in infants
○ UTI
○ Septicaemia
○ Cholecystitis, cholangitis, otitis, peritonitis, wound infections

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Lab Diagnosis
• Specimen
○ Urine
○ Pus
○ Blood etc.
○ Depends on the site of infection
• Gram stain
○ Gram negative rods
• Culture
○ It grows on MacConkey, CLED or Blood Agar media
 Growth conditions in air, 35-37°C, 18-24hours
 Colonies appear mucoid and pink MacConkey agar media
 CLED
□ Yellow colonies
• Biochemical
○ IMViC = --++
○ TSI: acid slant, acid butt, gas, no H2S
○ Urease positive
• Capsule swelling (Quellung Reaction)_

Treatment
• Based on antibiotics susceptibility tests
○ Beta lactam + lactamase inhibitor combination
○ Third generation of cephaloporins e.g. Cefataxime, Ceftriaxone
○ Carbapenems/Fluoroquinolones/Aminoglycosides

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Proteus
Tuesday, 13 June 2017 5:31 AM

Genus Proteus
• Lactose non-fermenters
• Urease positive
• Non-capsulated
• Highly motile
○ Exhibit Dienes Phenomenon
○ Swarming motility
• Contain O (somatic antigen) and H (flagellar antigen)

Pathogenesis
• Habitat
○ Human colon
○ Soil
○ H2O
• Highly motile and may contribute to ability to invade urinary tract
• Has fimbriae - for uroepithelial colonisation
• Urease hydrolyses urea in urine into ammonium hydroxide increasing urinary pH -
encourage calculi formation (kidney stones) which obstruct urine flow and damage urinary
epithelium.
• Alkaline urine favours growth of the organism and causes more extensive renal damage
• Bacteria in the calculi not reached by drugs - re-infection
• Haemolysins
○ In uropathogenic proteus

Clinical Syndrome
• UTI
○ Alkalinised urine favourable for deposition of calcium and magnesium salts and the
formation of kidney stones
 Stag-horn calculi
 Alkaline environment can cause struvite stone formation
• Wound infection (including pressure sores, burns and damaged tissues)
• Septicaemia
• Ear infections
• Brain abscesses
• Involved in synergistic non-clostridial anaerobic myonecrosis - caused by a combination of
GNB (E. coli or Klebsiella or Enterobacter spp.) and anaerobes

Lab Diagnosis
• Specimens (list)
○ Urine
○ Pus
○ Blood
○ CSF
• Direct gram stain
○ Gram negative bacilli
• Culture
○ MacConkey
 Non-lactose fermenter (NLF)

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  Non-lactose fermenter (NLF)
○ BA
 Swarming
 Distinct fishy odour
• Biochemical reactions
○ Oxidase negative
○ Urease positive
○ TSI
 Alkaline slant, acid butt, gas; P. vulgaris, P. mirabilis produce H2S - blackens
butt
○ Indole test
 P.mirabilis is indole negative; the rest are positive

Treatment
• Trimethoprim-Sulphamethoxazole/Nitrofurantoin/Ampicillin
• Nofloxacin

Treatment
• Based on susceptibility testing
• Combination of Trimethoprin-Sulphamethoxaole
• Am_

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Salmonella
Thursday, 11 May 2017 11:14 AM

Host Range
• All vertebrae in their gut
• Some serotypes in arthropods, flies, cockroaches

• Earlier classification included

○ The Kaufmanns-White system, which identified each serotype as an individual Salmonella
species
○ The Edwards-Ewing System, which divided the salmonellae into 3 species (S. choleraesuis, S.
enteritidis and S. typhi) and hundreds of serotypes
○ A DNA hybridisation scheme that lumped the salmonellae into two species known as S.
enterotidis and S. bongoris
• S.enterica contains more than 2,500 serotypes (2541 in I 2004) differentiated on the O and H
antigens
○ Salmonella serotype (serovar) Typhimurium
○ Salmonella serotype Enteritidis
○ Salmonella serotype Typhi
○ Salmonella serotype Paratyphi
○ Salmonella serotype Choleraesuis etc.
• Transmission related to ingestion of food, water contaminated by human, animal waste
• Frequent animal sources
○ Poultry
○ Eggs
○ Beef
○ Milk
• Contaminated vegetables

Morphology and General Characteristics

• Gram negative rod
• Peritrichous flagella (ensure motility)
• Uncapsulated (except S. typhi)
• Unsporulating
• Characteristically ferment glucose and mannose without producing gas
• Do not ferment sucrose or lactose
• Most produce H2S
• Facultative intracellular bacteria, mostly found in macrophages
• Survive freezing water for long periods of time
• Resistant to certain chemicals that inhibit other enteric compounds and therefore, such compounds
are useful for inclusion in media to isolate salmonellae from faeces
○ Brilliant green
○ Sodium tetrathionate
○ Sodium deoxycholate

Nomenclature
• Salmonella classification is complex but basically, it can be divided into two species:
○ Salmonella enterica
○ Salmonella bongori
• S. enterica contains 5 subspecies
○ enterica - subspecies I - cause most human disease
○ salamae - subspecies II
○ arizonae - subspecies III
○ diarizonae - subspecies IIIb
○ houtenae - subspecies IV
○ indica - subspecies V
• Most human illness is caused by the subspecies I strains written as S. enterica subspecies enterica
• In subspecies I, serotypes or serovars (serologically distinguishable strain of a microorganism) are
designated by name, usually by associated disease, geographical origins or usual habitats.
• The serovar is not italicised and written starting with a capital.
• The nomenclature will be as follows:
○ S.enterica subspecies enterica serovar Typhimurium or S Typhimurium
○ S.enterica subspecies enterica serovar Typhi or S Typhi

Antigenic Structure
• O (somatic) and H (flagella) and Vi (Virulence)
• O antigen
○ Heat stable long chain lipopolysaccharides (LPS)
○ Located in the outer membrane
○ Exhibit variation in sugar composition and degree of polysaccharide branching
○ Structural heterogenicity contributes to the large number of serotypes in Salmonella Typhi
• H antigen
○ Contains epitopes that form the basis of flagella based serotyping
○ Exhibit diphasic variation resulting in phase 1 and 2 antigens
○ Some salmonellae express only one flagellar phase
○ Heat labile
• Vi antigen
○ Surface polysaccharide (capsules)
○ Make it difficult for antibodies designed to recognise the LPS antigen
○ Salmonella enterica serotype Typhi is the most important example which expresses this

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 ○ Salmonella enterica serotype Typhi is the most important example which expresses this
antigen
• Variation
○ Organisms may lose the H antigens and become non-motile
○ Loss of O antigen is associated with a change from smooth to rough colony form
○ Vi antigen may be lost partially or completely
○ Antigens may be acquired or lost in the process of transduction

Pathogenesis
• The organisms almost always enter by the oral route usually with contaminated food or drink
• The mean infective dose to produce clinical or subclinical dose is 105-108 salmonellae or perhaps
103 is enough.
• Among the host factors that contribute to resistance include
○ Gastric acidity
○ Normal intestinal microbiota and,
○ Normal intestinal immunity
• Salmonella produces three main types of disease in humans but mixed types are frequent.
○ Typhoidal salmonella (Typhoid Fever, Paratyphoid Fever)
○ Non-typhoidal salmonella
 Invasive non-typhoidal salmonella
 Non-invasive non-typhoidal

Typhoidal Salmonella - The 'Enteric Fevers'

• Caused by serovars (Salmonella Typhi, Paratyphi A, B and C)
• Salmonellae pass through the lymphatic system of the intestine into the blood
• Carried to various organs (liver, kidney, spleen) to form secondary foci
• Salmonella typhiis found in the gall bladders of chronic carriers
• Incubation period of 10-14 days; the organisms multiply in intestinal lymphoid tissue and are
excreted in stools
○ The bacteria are acid labile and are easily destroyed by acid in the stomach and thus require a
large infective dose to case clinical manifestations
 If you are taking Omeprazole, suffer from Pernicious anaemia or had any condition that
lowered stomach acidity, you are more susceptible to infection
• Dead salmonellae → endotoxins released
○ Endotoxins act on the vascular and nervous apparatus
 Increased permeability
 Decreased tone of vessels
 Upset thermal regulation
 VD
○ Lost liquid and electrolytes, decrease in circulating blood volume and arterial pressure →
hypovolaemic shock
○ Septic shock may also occur
• Contains a type III secretion system
○ A protein that not only detects eukaryotic cells, but when it does, it secretes a protein that helps
with infectivity
Invasive Non-Typhoidal Salmonella Disease
• Caused mainly by the S. Typhimurium or S. Enteritidis
• Invade the body through ingestion of undercooked chicken (chicken are the main reservoir)
• Commonly isolated in blood of those presenting with fever
• Diverse symptoms
○ Fever
○ Hepatosplenomegaly
○ Respiratory symptoms
○ Often with an absence of GIT symptoms

Non-Invasive Non-Typhoidal Salmonella (Salmonellosis)

• Caused by non-typhoidal serovars of Salmonella
• Generally cause food poisoning
• Require a higher infectious dose
○ Salmonellae enter the small intestines and multiply in tissue
○ Cells are poisoned by endotoxins released from dead salmonellae
○ The local response to the endotoxins is enteritis and GI disorder

Clinical Findings
• Enteric Fever
○ Caused by strains of S. Typhi or S. Paratyphi A, B or C
○ Early symptoms
 Dull continuous abdominal tenderness
 Hepatomegaly
 Splenomegaly
 Red macules on the abdomen - 25% of patients
○ Diarrhoea is uncommon
 Diarrhoea is said to resemble pea soup
○ Complication
 Severe intestinal haemorrhage and perforation
• Gastroenteritis and Food Poisoning
○ Organisms penetrate mucosal cells into the lamina propria with resulting inflammation and
diarrhoea
○ Clinical features:
 VDP
 Inflammatory diarrhoea

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  Inflammatory diarrhoea
 Fever
○ Dehydration can lead to hypotension, cramps and renal failure
• Bacteraemia and Metastatic Disease
○ Constant feature of Typhoidal Salmonella
○ Sickle cell anaemia
 Important predisposing factor resulting in osteomyelitis, pneumonia and meningitis
 Osteomyelitis in patients with sickle cell disease
 Major cause
○ Ability of salmonella to survive within the macrophages, particularly in the liver and BM leads to
persistent infection and chronic carrier state
• Prolonged Carrier State
○ Chronic carriage
 Excretion of salmonella for a year or more
○ Can occur with any serotype
○ Bacilli are present in the gall bladder and are shed in faeces
○ The long duration of the carrier state enables the enteric fever bacilli to survive in the
community in non-epidemic times

Laboratory Diagnosis
• Specimen
○ In enterocolitis
 Organism isolated in stool sample
 Stool samples may also be positive in chronic carriers
○ In enteric fever
 Blood culture
 BM cultures are often positive
• Culture
○ Fluid enrichment media
 Stool specimen on day 1
 Tetrathionate or Selenite F broth
○ Selective media
 Stool specimen on day 2 subculture either on:
□ Deoxycholate citrate agar (DCA)
□ Xylose lysine deoxycholate agar (XLD)
□ Salmonella-shigella agar (SS)
□ Hektoen Enteric Agar (HE)
 When streaked for isolation, notice black Salmonella colonies due to lactose
fermentation with acid end products
○ At 35-37°C for 18-24 hours in air
○ In blood culture, there are 2 types of cultures
 Liquid blood culture with a tryptic soy broth
□ Shake it and if there is turbidity, you can subculture in a solid medium
 Liquid and solid phase
□ Culture in both phases and check for turbidity in the liquid phase and colloids in the
solid phase
○ Characteristics
 Facultative anaerobes
 Non-lactose fermenters - on MacConkey
 Gram negative bacilli
 Oxidase negative
 Urease negative - for differentiate between Proteus and E. coli
○ On TSI, salmonellae produce an alkaline slant, acid butt frequently with both gas and H 2S
 There are cracks in the slant because of the gas
□ Ferments glucose and produces a gas
○ Slide agglutination
 Detects presence of somatic (O) antigen from suspicious colonies from culture plates
○ Colonies are also sub-cultured to peptone water for determination of flagellar (H) antigen Deoxycholate Agar
• Gold standard of diagnosis is culture • Use
○ For isolation and differentiation among the
Treatment Enterobactericeae
○ Detecting coliforms in dairy products
• Enteric Fever
• Principle
○ Ciprofloxacin
○ Contains
○ Ceftriaxone
 Lactose - fermentable carbohydrate
• Gastroenteritis  Sodium deoxycholate - gram-positive
○ Replacement of fluids and electrolytes inhibitor
○ Control of NV and pain  Neutral red - colourless in pH above 6.8
• Salmonella bacteraemia and red below and acts as pH indicator
○ Ciprofloxacin □ Reveals lactose fermentation
○ Chloramphenicol • Observation
○ Cotrimoxazole ○ No colour change as Salmonella does not
○ High dose of Ampicillin ferment lactose
• Salmonella meningitis
○ Cefotaxime and Ceftriaxone
 Both can penetrate the CSF
• Chronic Asymptomatic Carriers
○ If the principle site of carriage is the biliary tract, cholecystectomy (it poses a risk of
dissemination of the organism during surgery)
○ Ampicillin, Amoxicillin, Cotrimoxazole or Ciprofloxacin

Prevention and Control

• Vaccination
○ Heat killed phenol preserved whole vaccines containing a mixture of cultures of Typhi,
Paratyphi A and B (TAB)
• Public availability of treated water

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• Public availability of treated water
• Proper disposal of human excreta
• Prevention of contamination by food handlers, rodents
• Raw foodstuffs of animal origin must never have direct or indirect contact with cooked food
• Proper refrigeration of cooked food
• Wash hands after handling pets

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Shigella - Bacillary Dysentery
Thursday, 11 May 2017 11:57 AM

Important Properties
• 4 species based on serology and biochemical reactions
○ Dysentriae - 10 serotypes
○ Boydii - 15 serotypes
○ Flexneri - 6 serotypes
○ Sonnei - serologically homogenous and are therefore typed by other means
• Causes bacillary dysentery by an invasive mechanism identical to enteroinvasive E. coli
(EIEC)
• Pathogen of man and other primates
• Spread by contaminated food and water
• Transmission
○ Faecal-oral route with the principle factors in transmission being fingers, flies, food
and faeces

Pathogenesis
C.f. EIEC
Shiga toxin****

Clinical Features
• The illness begins with fever unlike ETEC and cholera which do not invade the epithelial
cells and therefore, do not cause fever.
• Abdominal pain and diarrhoea follow.
○ The diarrhoea may contain flecks of bright-red blood and pus (WBCs)
• Patients develop diarrhoea because the inflamed colon, damaged by the shiga toxin, is unable to
reabsorb fluids and electrolytes.
• Sh. dysenteriae causes a more severe illness with marked prostration and paediatric
febrile convulsions
• It is also associated with haemolytic uraemic syndrome
• Rarely invades other tissues, hence septicaemia and metastatic infection rarely occurs

Lab Diagnosis
• Wet preparation in stool specimen
○ Many RBC with many leucocytes (pus cells) and the bacilli
• Gram negative bacilli
• NLFs on MAC agar
• Sh. Shonnei is the only late lactose fermenter
• Stool specimen cultures on SS, XLD or DCA
• TSI
○ Alkaline slant
○ Acid butt
○ No gas
○ H2S
• Suspicious colonies
○ Confirmed with species specific antisera followed by type specific antisera

Treatment
• Most cases are mild and self-limiting, so are treated with oral rehydration therapy, rather
than with antibiotics
• Antibiotics may be indicated in the sever infections, patients with extreme age and
immunocompromised

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 immunocompromised
○ Options include Ciprofloxacin and Cotrimoxazole
• Antiperistaltic drugs are C/I as they prolong the fever, diarrhoea and excretion of the
organisms

Prevention
• Largely dependent on interruption of faecal-oral transmission by proper sewage disposal
• Adequate chlorination of water
• Good personal hygiene
• Public health education

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Pseudomonas - Nosocomial Infections
Thursday, 11 May 2017 12:08 PM

Epidemiology and Transmission

• Natural habitat
○ Temperature between 4-36°C (can survive upt to 42°C)
○ Found throughout nature in moist environment (hydrophilic) e.g. sink drains,
vegetables, river water, antiseptic solutions, mineral water etc.
○ P.aeruginosa rarely colonies healthy humans. Normal skin does not support P.aeruginosa
colonisation.

Classification
• Species include
○ Aeruginosa
○ Putida
○ Fluoresens
○ Stutzeri
○ Mendocina
• Ps.aeruginosa is most implicated in human disease

Biological Characteristics
• Non-fastidious
• Distinct sweet fruity smell
• Most strains produce diffusible pigments e.g.
○ Pyocyanin - bluish
○ Pyoverdin - yellow green
○ Pyorubin - red
○ Pyomelanin - brown

Pathogenesis
• Both toxigenic and invasive
• The process of infection has 3 stages
○ Attachment and colonisation
○ Local invasion
○ Dissemination and systemic disease
• Associated with multiple virulence factors produced depending on site and nature of
infection

Virulence factors Biologic effects

Structural components
Capsule Adhesin, antiphagocytic
Pili Adhesin
LPS Endotoxic activity
Pyocyanin Impairs ciliary functions
Toxins and Enzymes
Exotoxin A Inhibits protein synthesis
Exotoxin B -
Elastase Disrupts tissues with elastin

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Elastase Disrupts tissues with elastin
Leukocidin Disrupts leukocyte function
Phospholipase C Heat labile haemolysin

Range of Clinical infections caused by P. aeruginosa

• P.aeruginosa is an opportunistic infection
○ Individuals with normal host defenses are not at risk for serious infection with P.
aeruginosa
• Those at risk for serious infections include:
○ Profoundly depressed circulating neutrophil count e.g. cancer chemotherapy.
 also associated with CGE
○ High degree burns.
○ Cystic fibrosis.
○ Patients of mechanical ventilation.
• Immunocompetent host
○ Most common cause of osteochondritis of the foot following puncture wounds
(running shoes)
○ Hot tub facilities
○ Swimmer's ear (acute otitis externa)
○ Conjunctivitis in contact lens users (poor hygiene or if lenses are worn for extended
periods)
• Other hosts
○ Malignant otitis externa in diabetics
○ Meningitis post trauma or surgery
○ Sepsis and meningitis in new-borns
○ Endocarditis or osteomyelitis in IV drug users
○ Community-acquires pneumonia in patients with bronchiectasis
○ UTIs in patients with urinary tract abnormalities

Lab Diagnosis
• Specimen
○ Depends on site of infection e.g. wound swabs
• Culture
○ Grows best at 37°C in aerobic conditions
○ Characteristic fruity odours and pigments
 Pyocyanin (blue)
 Pyoverdin (yellow-green fluorescent)
 Pyorubin (red)
 Pyomelanin (brown)
○ Non-lactose fermenters on MAC
○ TSI
 Alkaline slant
 Alkaline butt
 No gas
 No H2S
• Staining
○ Gram stain
 Gran-negative rods
• Biochemical tests
○ Rapidly oxidase positive
○ Catalase positive
○ Urease negative
• Serotyping
○ Used for epidemiological purposes
• Characteristics

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 • Characteristics
○ Motile
○ encapsulated
○ Non-sporing
○ Broad antibiotic resistance
○ obligate aerobe

Treatment
• Antibiotic susceptibility of isolates has to be done due to resistance
• Anti-pseudomonal agents include
○ Fluoroquinolones
 Ciprofloxacin
○ Aminoglycosides
 Gentamycin
 Tobramycin
○ Anti-pseudomonal penicillins
 Piperacillin
○ Carbapenems
 Imipenem
 Meropenem
○ Polymixins
 Polymixin B
○ Monobactams
 Aztreonam

Control
• Hospital infection control methods should concentrate on preventing contamination of
sterile medical equipment and nosocomial infections

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Gram Negative Rods
Monday, 15 May 2017 11:13 AM

Vibrios
• Belong to the family, Vibrionaceae
• Gram-negative curved or comma shaped bacilli
• Highly motile by a single polar flagellum
• Non-spore forming
• Non-capsulated
• Facultative anaerobes
• Oxidase positive
• Produce acid (ferment sucrose or glucose) but not lactose
• Possesses both H and O antigens

Identification
• Highly motile
• Gram-negative
• Curved/comma shaped rods with singular polar flagellum

Pathogenic strains
• V.cholerae
• V.parahaemolyticus
• V.vulnificus

What is Cholera?
• Intestinal infection
• Severe diarrhoea
• Caused by cholera toxin of Vibrio cholera

Epidemiology
• Responsible for 7 global pandemics over the past two centuries
• Common in India, Sub-Saharan Africa. Southern Asia
• Very rare in industrialised countries

Strains causing Epidemics

• 2 main serogroups carry set of virulence genes necessary for pathogenesis
• O1
○ Classical: 1 case per 30-100 infections
○ El Tor: 1 case per 2-4 infections
• O139
○ Contained in India, Bangladesh

Transmission
• Contaminated food or water
○ Inadequate sewage treatment
○ Lack of water treatment
○ Improperly cooked shellfish
• Transmission by casual contact unlikely

Epidemics
• Faecal-oral transmission
• Faeces of infected person contaminates water supply
•

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 • Resulting diarrhoea makes it easy for bacteria to spread in unsanitary conditions

Pathogenesis
• To establish disease V. cholerae must be ingested in contaminated foo d or water and
survive passage through the gastric barrier of the stomach
• On reaching the lumen of the small intestine, penetrates the mucous layer and establish
contact with the epithelial cell layer

How does Cholera Toxin Work?

• Inactivates GTPase function of G-protein coupled receptors in the intestinal cells
• G proteins stuck "on" position
• 100 fold increase in cAMP
• Activation of ion channels
• Ions floe out and water follows

People Most At Risk

• People with low gastric acid levels
○ Children: 10X more susceptible than adults
○ Elderly
• Blood grouping
○ Individuals of O blood group
○ O>B>A>AB

Diagnosis: Visible Symptoms

• Decreased skin turgor
• Sunken eyes, cheeks
• Almost no urine production
• Dry mucous membranes
• Watery diarrhoea consists of:
○ Fluid without RBC, proteins
○ Electrolytes
○ Enormous numbers of vibrio cholera (107 vibrios/mL)

Outline of Laboratory Isolation and Identification

• Specimen
○ Stool
• Transport
○ Cary-Blair Transport Medium
• Microscopy
○ Wet preparation
 Darting motility
• Culture
○ Enrichment medium
 Alkaline peptone water for 4-6 hours
○ Subculture on TCBS - thiosulphate citrate bile salt sucrose - in air at 37°C, for 18-24
hours
○ Colonial morphology
 Yellow colonies - V. cholerae
 Green colonies - V. parahaemolyticus
• Gram stain
○ Do not take up gram stain
• Biochemical test
○ Oxidase positive
○ Ferments glucose and sucrose

Treatment

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Treatment
• Even before identifying the cause of disease, rehydration therapy must begin immediately
because death can occur within hours
○ Oral rehydration
○ IV rehydration
• Antimicrobial therapy

Travelling Precautions
• Boil or treat water with chlorine or iodine
• No ice
• Cook everything
• Rule of Thumb
○ Boil it
○ Cook it
○ Peel it
○ Or forget it
• Wash hands frequently
• Vaccines - individuals can be carriers
○ High risk individuals
○ Food handlers e.g. chefs

Campylobacter
• Gram negative curved rods
○ Often attached in pairs giving a "seagull" appearance
• Motile by polar flagella
○ Darting motility in corkscrew fashion
 Facilitate penetration and colonisation of mucosal environment
• Species
○ Campylobacter jejuni
• Causes 80-90% of all illnesses
• Source
○ C.jejuni
 Associated with poultry GIT
 Also common in cattle
• Transmission
○ Raw or undercooked food products
○ Through direct contact with infected animal
• Pathogenesis
○ ingestion
○ Bypasses gastrum using flagella
○ Penetrates epithelial cells in SI
○ Produces Campylobacter toxin which induces inflammation
○ Affects fluid absorption and retention leading to diarrhoea
• Symptoms
○ Caused by diarrhoea
• Lab diagnosis
○ Specimen: stool plus rarely blood in bacteremia
○ Cary-Blair transport media
○ Direct wet prep : dark-field microscopy observing darting motility
○ Culture: Skirrow's medium
 Incubate in microaerophilic conditions at 42 degrees for up to 48h
 Gram stain: negative comma or S-shaped rods
 Oxidase positive

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Differentiate Camphylobacter, Salmonella and Cholera diarrhoea

• Treatment
○ Rehydration
○ In severe dysentery - erythromycin, ciprofloxacin
○ Antimotility agents not recommended

Helicobacter
• Colonize stomach in 50% of the world population
• H.pylori is contagious although route of transmission is unclear
• Main risk factor for:
○ Gastric and duodenal ulceration
○ Stomach cancer
○ Gastric MALT lymphoma
• Characteristics
○ Gram negative curved rod
○ Microaerophilic
○ Motile
○ Produces oxidase, mucinase and urease enzymes
• Virulence factors
○ Corkscrew motility- penetrate mucosal later
○ Adhesins - which help it adhere to epithelial cells
○ Mucinase- Degrades gastric mucous
○ Urease - Converts urea into ammonia
 Neutralise the local acid environment
 Localised tissue damage
○ Flagella
○ LPS
○ Exotoxins
• Pathogenesis
○ Attachment to mucus-secreting cells of gastric mucosa
○ Produce ammonia from urea by urease
○ Ammonia neutralises HCl
○ Organism survival
○ Plus inflammatory response
○ Degrades mucus
○ Gastritis + peptic ulcers
• Clinical Manifestations
○ Bloating
○ Nausea
○ Anorexia
○ Flatulence
○ Bad breath
○ Belching
○ Vomiting
• Clinical outcomes
○ Chronic gastritis
○ Gastric ulcers
○ Gastric cancers
• Lab diagnosis
○ Non invasive
 Blood antibody test - IgG detection
 Stool antigen test
 Carbon urea breath test

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  Carbon urea breath test
○ Invasive
 Gastric biopsy
□ Urease positive
□ Culture on Helicobacter Agar - 3-5days
 Oxidase, urease positive
• Treatment
○ First line
○ Second line
○ Third line

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Haemophilus, Bordetella and Brucella
Thursday, 18 May 2017 11:06 AM

Haemophilus
Classification
• Family:
○ Pasteurellaceae
• Genus
○ Haemophilus
• Species
○ Influenzae
○ Parainfluenzae
○ Haemolyticus
○ Aegyptius
○ Aphrophilus
○ Ducreyi
○ Pittmaniae
○ Sputorum
○ Segnis
○ Paraphrohaemolyticus
○ Paraphrophilus

General Characteristics
• Haemophilus = blood loving
• Gram negative coccobacilli
• Non-motile
• Non-sporing
• Facultative anaerobes
• Oxidase positive
• Catalase positive
• Require accessory factors for growth and viability
• Requirements differ among the species

Accessory Growth Factor

• "X" factor (Haemin)
○ Component of elemental iron containing part of haemoglobin
○ Heat stable
○ Required for synthesis of cytochrome and other enzymes e.g. catalase, peroxidase, oxidase
• "V" factor (NAD or NADP)
○ Hydrogen acceptor in cell metabolism
○ Heat labile (destroyed at 120°C)
○ Present inside RBCs
○ Synthesised by some fungi and bacteria

Virulence Factors
• Capsule
○ Contains PRP (Polyribosyl-ribito phosphate)
○ Avoid phagocytosis and opsonisation
○ Main virulence factor
• Adhesion proteins
• Outer membrane proteins

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 • Outer membrane proteins
• IgA1 protease - H. influenzae
• Lipooligosaccharide (LOS)

Haemophilus Influenzae
• Pfeiffer's bacilli
• Capsulated and non-encapsulated strains
• 6 capsular types
○ A-F
• Non-encapsulated - non-typeable H. influenzae (NTHi)
• Both capsulated and non-encapsulated further subdivided into 8 biotypes
○ On the basis of urease, ornithine decarboxylase activities and indole production (I - VIII)

Epidemiology
• Mucosal organism
• Present in 30-50%of healthy persons
• Spread by airborne droplets and contact secretions
• H.influenzar infections seen frequently in children aged from six months to four years of age
• In adults
○ Secondary complications of severe primary illnesses
○ Immune compromise

Clinical Implications
• Haemophilus influenzae serotype b (Hib) responsible for majority of invasive disease
• Carrier state for capsulated b strains is about 2-4%
• Type b capsule deficient mutant (b-)
• NHTi
○ Form part of the normal microflora of the URT
○ Usually involved in respiratory tract infections and otitis media but may also cause invasive
disease
○ Post-Hib vaccine era, responsible for majority of invasive diseases

Pathogenesis
• Muco-ciliary interactions
• Attachment to respiratory mucosa
○ Adhesins
○ Pili
• Evasion of mucosal immunity
○ Proteases
○ Microcolony formation
○ Phase variation/antigenic shift
○ Intracellular survival/invasion of local tissue

Infections
• Invasive (bacteraemic) infections
○ Meningitis
○ Bacteraemic pneumonia
○ Epiglottitis
○ Septic arthritis
○ Septicaemia
○ Cellulitis
○ Osteomyelitis
• Localised nfections
○ Otitis media
○ Sinusitis

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 ○ Sinusitis
○ Bronchitis

Lab Investigation
• Specimen - does not retain viability for long. Specimens must be cultures as soon as possible and
not refrigerated
○ CSF
○ Blood
○ Pus swabs
○ Sputum
○ Nasopharyngeal specimen
• Direct gram stain
• Culture for isolation and identification
○ CBA
 5-10% CO2
 35°-37°C
 18-24h
 Colonial morphology
□ Variation in size and appearance
□ Translucent to mucoid
• Gram-negative coccobacilli or short rod
• Long thread-like and pleomorphic forms may be seen in CSF or following culture
• Demonstration of growth factor requirements
○ Satellitism test
 S.aureaus produces factor V in excess of its own needs
 It is cultured on BA with H. influenzae; the factor V and the haemin released by
staphylococcal haemolysins help the growth of H. influenzae
○ Commercially prepared factors (X, V, XV) on NA (What is factor XV?)
 Biochemical tests
 Serology
 Slide agglutination
 PCR
 Antimicrobial susceptibility test

Treatment
• Ampicillin
• Chloramphenicol
• Cephalosporins

Prevention
• Vaccination - main method
○ Hib conjugate vaccine

H.influenzae biogroup aegyptius (Hae)

• Also known as Koch-Weeks bacillus; H. aegyptius
• A strain of NTHi
• Causes
○ A purulent conjunctivitis
 Worldwide seasonal epidemics of acute purulent conjunctivitis (Hae)
○ Brazilian Purpuric Fever (BPF) (Hae BPF)

BPF
• Firs recognised in Brazil in 1984
• Conjunctivitis
○ Overwhelming septicaemia resembling fulminating meningococcal infection
• Characterised by:

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 • Characterised by:
○ High fever
○ Haemorrhagic skin lesions
○ Abdominal pain
○ NV
○ Septic shock
○ Death (fatality rate of 40-90%)

Lab Diagnosis
• Specimen
○ Conjunctival swabs
○ Blood
○ CSF
• Gram stain
○ Gram negative coccobacilli
• Culture
○ C.f. H. influenzae

Treatment
• Ampicillin
• Amoxicillin/clavulanic acid
• Cephalosporins
○ Ceftriaxone

Haemophilus ducreyi
• Causative agent or a sexually transmitted ulcer (chancroid/soft sore)
• Common cause of genital ulceration in tropical countries
• With or without lymphadenitis or bubo formation

Clinical Presentation
• Tender erythematous papule, 4-7 days after infection
• Pustular stage
• Pustules rupture after 2-3 days
○ Usually
• Painful, shallow ulcers unilateral, spherical
with granulomatous basesand
andpainful
purulent exudates
• Ulcer
○ Edge is rugged and undermined
• Ulceration can take several weeks or months to resolve in the absence of antimicrobial therapy
• Lesions occur:
○ On prepuce and frenulum in men
○ Vulva, cervix and perianal area in women
• In 50% of cases, painful, tender inguinal lymphadenitis
• Buboes may develop
• Lymphadenopathy

Lab Diagnosis
• Clinical history
• Specimen
○ Swab
• Culture
○ Fastidious, difficult to isolate
○ Requires factor X but not factor V
○ CBA with 1% Isovitalex and Vancomycin (to kill the normal flora)
○ CO2, 32-35°C
• Biochemical tests
○ Slowly oxidase positive

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 ○ Slowly oxidase positive
○ Catalase, urease and indole negative
• PCR

Treatment
• Erythromycin
• Azithromycin
• Ceftriaxone
• Ciprofloxacon
• Spectinomycin

HACEK Infections
• Group of fastidious, slow-growing gram negative bacteria
○ Haemophilus spp.
 Parainfluenzae
○ Aggregatibacter
 Actinomycetemcomitans
 Segnis
 Aphrophilus
○ Cardiobacterium
 Hominis
 Valvarum
○ Eikenella spp.
 Corrodens
○ Kingella spp.
 Kingae
• Normal flora in the oral cavity
• Associated with local infections in the mouth
• Cause severe systemic infections
○ Bacteria endocarditis, which can develop on either native or prosthetic valves
• Responsible for 3% of cases of infective Endocarditis
• Often a cause of culture-negative endocarditis

Other Infections
• Periodontal infections
• Bacteraemia
• Abscesses
• Peritonitis
• Otitis media
• Conjunctivitis
• Pneumonia
• Septic arthritis
• Osteomyelitis
• UTIs
• Wound infection
• Brain abscesses

Lab Diagnosis and Treatment

Diagnosis
• Specimen
○ Blood culture
○ Parainfluenzar requires only factor V
Treatment
• Penicillin
• Amino penicillin

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 • Amino penicillin
• Ceftriaxone
• Fluoroquinolones

Bordetella
Classification
• Family
○ Alcaliganaceae
• Genus
○ Bordetella
• Species
○ Pertussis
○ Parapertussis
○ Bronchoseptica

General Characteristics
• Small gram negative coccobacilli
• Strict aerobes
• Non-motile (Bronchoseptica is motile)
• Capsulated
• Non-spore forming
• Piliated
• Colonise the respiratory tracts of mammals

Virulence Factors
• Adhesins
○ Filamentous haemagluttnin (FHA)
○ Fimbriae
○ Pertactin
• Toxins
○ Pertussis toxin (Ptx)
○ Adenylate cyclase (CyaA) - haemolysin
○ Dermonecrotic toxin
○ Tracheal cytotoxin
• Secretion systems
○ Type III
○ Type IV

Clinical Implications
• Bordetella pertussis
○ Causes whooping cough
• Bordetella parapertussis
○ Causes a milder form of whooping cough
• Bordetella bronchoseptica
○ Causes infections ranging from lethal penumonia to asymptomatic respiratory cariage

B.pertussis
• A strict human pathogen with no known animal or environmental reservoir
• Colonises the cilia
• Causes whooping cough
○ A highly contagious acute respiratory illness of humans
 A relatively mild disease in adults
• Has significant mortality rate in infants
• Transmission is via respiratory droplets

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 • Transmission is via respiratory droplets

Pathogenesis
• Infects the host by colonising lung epithelial cells
• FHA binds to sulfatides on cilia
• Once anchored, bacterium produces tracheal cytotoxin which stops the cilia from beating
• Prevents body from clearing debris from the lungs → body responds by sending the body into a
coughing fit

Whooping Cough
• Incubation: 7-10 days
• Classic illness
○ Primary infection in unimmunised children
○ Lasts 6-12 weeks or longer
○ Occurs in 3 stages
 Catarrhal
□ Characterised by
 Rhinorrhoea
 Lacrimation
 Mild cough
□ Over a 7-14 day period, the cough worsens both in frequency ad degree
□ Temperature is normal or occasionally mildly elevated
 Paroxysmal
□ Onset during the second week of illness
□ Repeated coughing fits with 5 - 10 or more forceful coughs during a single expiration
(a paroxysm)
□ At the end of a paroxysm, there is massive inspiratory effort during which the classic
whoop occurs
□ The paroxysmal stage lasts for 2-8 weeks and sometimes longer
 Convalescent
□ Usually lasts for 1 to 2 weeks
• Complications
○ Encephalitis
○ Bronchopneumonia
○ Mental retardation

Mild Illness and Asymptomatic Infection

• Previously vaccinated children and adults
• Adults who had previously had B. pertussis infections
• Include
○ Rhinorrhoea
○ Tearing
○ Sneezing
○ Conjunctivitis
○ Fever
○ Sore throat
○ Cough of <2 weeks' duration

Clinical Presentation: Parapertussis and Bronchoseptica

• Parapertussis infection in humans
○ Unrecognised infection
○ Mild pertussis
○ Classic pertussis - milder than B. pertussis
• Bronchoseptica causes respiratory infections in many different mammals

Lab Diagnosis

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Lab Diagnosis
• Clinical history
○ Paroxysmal cough followed by whoop
• Specimen
○ Nasopharyngeal swabs and aspirate
○ Respiratory droplets on a cough plate
○ Gram stain
 Small gram negative coccobacilli
• Culture
○ Growth media enriched with blood
 Regan-Lowe agar (charcoal agar) supplemented with 10% gorse blood an cephalexin
 Bordet-Gengou agar
○ Incubate at 35-37°C in humid aerobic conditions
○ Slow growth: 2-4 days
○ Examine cultures daily for 7 days
○ Colonial morphology
 Small, pearly-grey, shiny (mercury-like), usually mucoid colonies
 Parapertussis grows more rapidly and forms larger colonies than B. pertussis
• Biochemical tests
○ Pertussis
 Urease negative
 does not reduce nitrate
 Oxidase positive
○ Parapertussis
 Urease positive (after 24 hours)
 Oxidase negative
• DFA (direct fluorescent antibody) test
○ Direct staining of nasopharyngeal secretions
○ Can provide a rapid, presumptive diagnosis
○ Low sensitivity and specificity
• ELISA
• PCR

Treatment
• Azithromycin
• Erythromycin
• Chloramphenical
• Amoxycillin
• Ampicillin
• Kanamycin

Prevention
• Early recognition and isolation of the patient to limit the spread
• Treatment
• Vaccination by pertussis vaccine

Brucella
Classification
• Family
○ Brucellaceae
• Genus
○ Brucella
• Species
○ Melitensis
○ Abortus

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 ○ Abortus
○ Suis
○ Ovis
○ Neotomae
○ Canis
○ Ceti
○ Pinnipedalis
○ Microti
○ Inopinata

General Chracteristics
• Pleomorphic gram negative bacilli (short or coccobacilli
• Non-sporeforming
• Non-motile
• Non-capsulated
• Obligate aerobes
• Oxidase, catalase and urease positive

Epidemiology
• Highly infectious species
• Cause infections in a wide variety of mammals

Zoonoses
• Entry via
○ Mucous membrane droplets
○ Broken skin - direct or indirect contact with infected animals
○ Ingestion
 Relatively common compare to the rest
• Have a low infectious dose an are capable is transmission via aerosols
• They are there fore classified as a potential warfare threat agent
• The spp. Primarily considered to be pathogenic to humans include
○ Melitensis
○ Suis (biobaers 1,3 and 4)
○ Abortus
○ Canis, sporadically

Virulence Factors
• Outer membrane proteins
• Type IV secretion systems

Pathogenesis
• Facultative intracellular pathogen
• Can survive and replicate in many types of host cells
• Prime targets - macrophages
• Brucella persists and replicate within phagocytic cells of he RES
• Temporary fusion of the Brucella-containing vacuole (BCV) with the lysosome
• Subsequent exclusion of the lysosomal proteins
• Brucella containing vacuole becomes associated within the ER
• Once inside this ER-associated compartment, the bacteria can establish chronic infection

Clinical Manifestation
• Brucellosis or undulant fever
○ Manifestations vary in severity and signs and symptoms
○ A severely debilitating and disabling illness can result
○ Human brucellosis usually manifests as an

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 ○ Human brucellosis usually manifests as an
 Acute (<2 months)
□ Fever, generalised malaise, headache, lymphadenopathy, hepatosplenomegaly
 Subacute (2-12months) febrile illness
□ Intermittent waves of fever
 Chronic (>1 year) incapacitating disease with severe complications
□ Possible internal organ damage

Complications
• Diverse depending on the specific site of infection
○ Osteoarticular (40%)
○ GUT
○ GIT
○ Nervous
○ CV
○ Skin and mucous membranes
○ Respiratory complications

Lab Diagnosis
• High degree of clinical suspicion
○ History of exposure to animals and exotic foods
• Specimen
○ Blood or bone marrow (iliac crest)
○ Infected tissue
○ Brucellae are highly infectious (Hazard Risk Group 3)
• Culture
○ Difficult to isolate
○ More likely to be isolated from the blood in acute brucellosis during times of fever
○ Culture medium
 Tryptone soya (tryptic soy) diphasic medium = castenada
○ Aerobic (B. abortus requires a CO2 enriched atmosphere)
○ 20-40°C (optimum 37°C)
○ Cultures should be kept for 4 weeks
○ Colonial morphology
 A variety of colonial forms
□ Smooth, mucoid, rough
 Colourless or grey
• Serology
○ Demonstration of IgM, IgG and IgA
 ELISA
 Standard tube agglutination test
 Modified tube agglutination test
 Brucellin skin test
○ Cross-reactions, gram negative bacteria e.g. V. cholerae, F. tularensis, Y. enterocolitica, E.
coli, Salmonella serovars
• Molecular diagnosis
○ PCR

Treatment, Prevention and Control

• Treatment
○ Doxycycline for 6/52 in combination with…
○ Streptomycin for 2-3 weeks of Rifampin for 6 weeks
• P/C
○ Control of disease in animal hosts
○ Effective heat treatment of dairy produce
○ Hygienic precautions to prevent occupational exposure

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○ Hygienic precautions to prevent occupational exposure

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Yersinia
Thursday, 25 May 2017 11:22 AM

Classification
• Family
○ Enterobacteriaceae
• Genus
○ Yersinia
• Species
○ Pestis
○ Pseudotuberculosis
○ Enterocolitica

General Characteristics
• Gram-negative rods or coccobacilli
• Facultative anaerobes
• Oxidase negative
• Ferment glucose
• Show bipolar staining

Yersinia Pestis
General Characteristics
• Plague bacillus
• Classic rodent zoonosis
• Pleomorphic gram-negative bacilli and coccobacilli
• Exhibits bipolar staining with Giemsa, Wright's or Wayson staining
○ Safety pin appearance
• Facultative anaerobe
• Non-motile
• Non-spore forming
• Facultative intracellular pathogen
• Once serotype
○ 3 biovars
 Antiqua
 Medievalis
 Orientalis

Epidemiology
• 3 historical plague pandemic
○ Justinian plague
○ Black death
○ Modern plague
• A re-emerging disease
• Endemic plague foci persist in many countries in Africa; the former Soviet Union; the
Americas, including the southwestern United States; and parts of Asia
• Wild rodents
○ Rats and squirrels
• Other animals together with rodents act a chronic carriers and reservoirs of infections
• Fleas acquire Y. pestis by feeding on the blood of infected animals
• Fleas then transmit the organism to susceptible animals and humans
• Modes of transmission
○ Flea bite

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 ○ Flea bite
○ Aerosols
○ Contact with secretions
• Main vectors
○ Xenopsylla cheopis
○ Xenopsylla brasiliensis - from rodents to humans
○ *Human flea - Pulex irritans -human to human

Virulence Factors
• Capsule
○ Anti-phagocytic
• Somatic antigen complex
○ Contains somatic antigens including V and W
○ Enable bacteria to resist phagocytosis
• LPS
○ Endotoxin activity
• Ability to absorb iron
• Other factors e.g.
○ Fibrinolysin which enable Y. pestis to spread in tissues

Clinical Implications
• Causative agent of plague/black death
○ Bubonic plague
○ Septicaemic plague
○ Pneumonic plague

Bubonic Plague
• Classic form of the disease
• Symptoms develop within 2 to 6 days of contact with the organism
○ Fever
○ Headache
○ Chills
○ Swollen, extremely tender lymph nodes (buboes)
○ GI Symptoms
 NVD
○ Buboes in the inguinal and femoral region
○ *Bacteraemia or secondary plague septicaemia may occur
• Case fatalities of untreated bubonic plague - 40-60%

Septicaemic Plague
• Primary Septicaemic Plague
○ Positive blood cultures but no palpable lymphadenopathy
• Secondary septicaemiac plague
○ Complication of both bubonic plague and pneumonic plague
• Symptoms
○ Fever
○ Chills
○ Headache
○ Malaise
○ GI disturbances
• Mortality rate - 30-50%
○ Without treatment -100%

Pneumonic Plague
• Rare but deadly form of disease

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 • Rare but deadly form of disease
• Spread via respiratory droplets through close contact
• Progresses rapidly from febrile flu-like illness to an overwhelming pneumonia with
coughing and production of bloody sputum
• If septicaemiac plague is left untreated, it may progress to secondary pneumonic plague

Laboratory Investigations
• Suggestive history (bubonic plague) - exposure to fleas, rodents
• Y.pestis is a highly infectious pathogen (hazard risk group 3)
○ Minimise the creation of aerosols
• Specimen
○ Blood
○ Bubo aspirates
○ Sputum
○ CSF
○ Scraping from skin lesions
• Gram stain
○ Gram-negative coccobacilli
• Culture for isolation and identification
○ BA
○ MacConkey
○ Yersinia selective medium (CIN - Cefsulodin-Ingasan-Novobiocin)
○ Incubate at 35-37°C (opitmum growth at 37°C); 24-48hours
○ Colonial morphology
 BA
□ Small, shiny non-haemolytic colonies
 Mac
□ Small, transluscent pink colonies
• Biochemical tests
○ Catalase positive
○ Oxidase positive
• Fluorescent antibody test
• Rapid immunoassays
• PCR

Treatment
• Early treatment
○ Survival ~100%
• Supportive care
• Treatment
○ Streptomycin/gentamycin
○ Tetracycline
○ Chloramphenical
 For CNS involvement
• Prophylaxis
○ Sulphonamides
○ Tetracycline

Prevention and Control

• Control of rats and fleas
• Control of infection from patients
○ Isolate
○ Handling patients with care

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Yersinia Enterocolitica and Pseudotuberculosis
Epidemiology
• Pseudotuberculosis
○ Primary pathogen of animals and fowls
• Enterocolitica
○ Infects swine, goats, cattle, horses, rodents and household pets
○ More commonly found in temperate countries
○ Can multiply in food refrigerated at 4-8°C

Clinical Significance
• Causative agents of illnesses with varying manifestation complicated by septicaemia
• Pseudotuberculosis
○ Associated with enterocolitis and acute mesenteric lymphadenitis
• Enterocolitica
○ Causes gastroenteritis, mainly in infants and young children
• Bacteria enter the lower intestinal tract and are transported with the macrophages into the
mesenteric lymph nodes

Clinical Manifestations
• Intestinal yersiniosis
○ Dominant symptom is enteritis with mesenteric lymphadenitis
○ Ileitis
○ Colitis
• Extraintestinal yersiniosis
○ Sepsis
○ Lymphadenopathy
○ Rarely hepatitis
○ Various local manifestations
 Pleuritis
 Endocarditis
 OM
 Cholecystitis
 Localised abscesses
• Other sequelae
○ Immunopathological complications
 Reactive arthritis
 Erythema nodosa

Laboratory Investigations
• Specimen
○ Blood
○ Stool
○ Lymph node aspirate
• Pseudotuberculosis is stained by modified ZN stain and is slightly acid fast
• Both are motile when growth at 22°C (Pestis is immotile when grown at 22°C)
• Culture
○ Media, incubation and colonies are similar to Pestis
○ A selective medium e.g. MacConkey, CIN agar or SS agar required to isolate from
faecal specimen
○ After 24-48 hours, incubation at 20-28°C produces small NLF colonies

Treatment
• Chloramphenal
• ciprofloxacin

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Pasteurella
Thursday, 25 May 2017 11:58 AM

Classification
• Genus
○ Pasteurella
• Species
○ Pasterella sensu stricto
 Multocida subsp. Multocida
 Multocida subsp. Gallacida
 Multocida subsp. Septica
 Dagmatis
 Gallinarum
 Canis
 Stomatis
 Avium

General Characteristics
• Small, gram-negative rods or coccobacilli
• Facultative anaerobes
• Non-motile
• Non-sporing
• Capsulated (P. multocida)
• Bipolar staining
• Fermenters
• Oxidase positive

Virulence Factors
• Iron acquisition mechanisms
• Membrane LPS
○ Confers serum complement resistance
• Capsule prevents phagocytosis and complement-mediated opsonisation
• Surface components that provide adherence properties e.g. FHA
• Extracellular matric degrading enzymes e.g. hyaluronidase, neuraminidase and proteases.
Facilitate colonisation and/or dissemination
• Dermonecrotic toxin (in highly virulent strains)
○ Causes dermonecrosis and modulates the immune system
• Pasteurella multocida toxin (PMT)

Epidemiology and Transmission

• Normal microbiota of the oral, nasopharyngeal and upper respiratory tracts of many wild
and domestic animals
• Transmission
○ Bite or scratch wounds from pets, predominantly cats and dogs
○ Contact skin lesions or nasopharynx or other upper respiratory mucosa
○ Multocida being the most prevalent isolate observed in human infections

Clinical Implications
• Cellulitis and localised superficial abscesses following an animal bite or scratch
• Oedema, cellulitis, bloody or suppurative/purulent exudate at the wound site
• Inflammation develops very rapidly
• Second most common site is the respiratory tract
• Most occurs in the elderly, underlying chronic lower RT disease

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 • Most occurs in the elderly, underlying chronic lower RT disease
• Route of infection is inhalation
• Others:
○ Osteomyelitis
○ Intra-abdominal infections
○ Septic arthritis
○ Sepsis
○ Meningitis

Laboratory Diagnosis
• Specimen
○ Swabs
○ Sputum
• Gram stain
○ Gram negative coccobacilli
• Culture
○ BA - produce gray colonies
• PCR

Treatment
• Combination therapy
○ Amoxicillin/clavulanic acid
○ Doxycycline + Metronidazole (penicillin allergies)
○ Clindamycin + a fluoroquinolone

Prevention and Control

• Avoid close contact with pets

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Francisella
Thursday, 25 May 2017 12:12 PM

Classification
• Family
○ Francisellaceae
• Genus
○ Francisella
• Species
○ Tularensis
 Tularensis
 Holarctica
 Mediasiatica
○ Philomiragia
○ Novicida

General Characteristics
• Gram-negative coccobacilli
• Facultative intracellular pathogens
• Non-motile
• Aerobes
• Non-spore forming

Virulence Factors
• Non-classical virulence factors
• Virulence stems from ability to proliferate within various host tissues and organs
• Disrupts their normal functions and induces a significant host inflammatory response that
contributes to the disease

Epidemiology
• Found globally in mammals and arthropod vectors
• Circulates in populations of rodents
• Outbreaks in humans often parallel outbreaks in animal populations
• Tularensis can be acquired by
○ Contact with, or ingestion of contaminated material including food and water
○ Inhalation of infectious particle
• A wide range of arthropod vectors implicated in transmission
○ Mosquitoes ticks
○ Deer flies
• Tularensis subspecies, tularensis and holarctica, cause the majority of reported cases
• Subspecies, tularensis, causes the more severe disease of the two

Pathogenesis
• Major target organs
○ Lymph nodes
○ Lungs
○ Pleura
○ Spleen
○ Liver
○ Kidney
• If untreated, bacteria spread from the skin and mucous membranes to regional lymph nodes
• In lymph nodes, further multiplication then dissemination to other organs

Clinical Manifestations
• Causes a spectrum of clinical illnesses termed tularaemia (glandular fever, rabbit fever, tick
fever, and deer fly fever)
○ Ulceroglandular
○ Oculoglandular
○ Oropharyngeal
○ Pneumonic
○ Typhoidal
○ Septic
• Francisella tularensis, is one of the most infectious pathogenic bacteria known, requiring
inoculation or inhalation of as few as 10 organisms to cause disease
• Therefore, can be used as a biological weapon of terrorism

Clinical implications
• Fever (38-40°C)
• Headache

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 • Headache
• Chills and rigor
• Generalised body aches
• Coryza and sore throat
• Dry or slightly productive cough
• NVD
• Case fatality 40-60% in untreated cases

Laboratory Investigations
• Specimen
○ Secretions
○ Exudates
○ Sputum
○ Gastric aspirate
○ Biopsy
○ Specimen
○ Blood
• Gram stain
• Culture
○ Culture media
 Cysteine heart blood agar
 Buffered charcoal-yeast agar
 CBA
○ Incubate at 37°C
 Growth visible from 24-48 hours
 Hold at least for 10 days before discarding
○ Colonial morphology
 CBA
□ Tiny, grey-white, opaque colonies
 Cysteine heart agar (CHA)
□ Greenish-blue colonies
• Direct fluorescent antibody
• Immunohistochemical staining
• PCR

Treatment
• Streptomycin - DOC
• Gentamycin
• Tetracycline
• Chloramphenical
○ CNS invasion

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Chlamydiaceae
Tuesday, 30 May 2017 3:14 PM

• Two genera
○ Chlamydia
○ Chlamydophila

Important Characteristics
• Obligate intracellular parasites
• Possess inner and outer membrane similar to those of gram-negative bacteria
• Contain both DNA and RNA
• Possess prokaryotic ribosomes
• Synthesise their own proteins, nucleic acids and lipids
• Susceptible to antibiotics

• Metabolically inactive infectious forms (elementary bodies [EBs])

• Metabolically active, non-infectious forms (reticulate bodies [RBs])
• EBs are resistant to many harsh environmental factors

Chlamydia trachomatis
• Serovars (antigenic difference - major membrane protein)
○ A-C - primarily conjunctiva
○ D-K - primarily urogenital tract
○ L1-L3 - inguinal lymph nodes

1.Trachoma
• Chromic, inflammatory, granulomatous process in the eye surface, leading to corneal
ulceration, scarring and blindness
• Active trachoma, characterised by the presence of lymphoid follicles on the conjunctiva
and intermittent shedding of chlamydiae
• Is primarily a disease of children
• Blindness can occur as a complication
• Spread of transfer of infected discharge from eye of infected person by hands, clothing,
towels
• Flies are important carriers
• Poverty, overcrowding, poor personal hygiene, inadequate water enhance spread

Stages of Trachoma infection

• Trachoma infection - small bumps under the eyelid
○ Follicles under the eyelid
• Follicles and inflammation
• Trachoma with scarring
○ Scar tissue forms
○ Eyelid is difficult to turn for examination
• Trachoma with trichiasis
○ Eyelids turn inwards towards the cornea
○ Possibility of infection with another pathogen e.g. Pseudomonas
• Corneal opacity
○ Corneal scarring
○ Bacteria/viruses enters damaged cornea

2.Adult Inclusion Conjunctivitis

• Acute process with mucopurulent discharge, dermatitis, corneal infiltrates and corneal

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 • Acute process with mucopurulent discharge, dermatitis, corneal infiltrates and corneal
vascularisation in chronic disease
• Most prevalent in sexually active, young people, being spread from the genitalia to the
eye; poor hygiene

3.Neonatal Conjunctivitis
• Acute process characterised by mucopurulent discharge
• Develops in infants around 14 days after birth
• Presents as a swelling of the eyelids and orbit and a purulent infiltration of the conjunctiva
• Acquired from the mother during birth
• If untreated, the infection usually resolves but a substantial proportion of these infants
develop chlamydial pneumonia about 6 weeks after birth

4.Genital Infection:
• Infection in men
○ C.trachomatis serovars D-K are responsible for about 30% of the cases of non-
specific urethritis in men
○ The infection can be asymptomatic, with infected men serving as a reservoir of
infection
○ In symptomatic patients, varying amounts of mucopurulent discharge are produced
○ Occasionally, this progresses to epididymitis or prostatis
○ It is likely that chronic chlamydial epididymitis will results in azoospermia
• Infect in Women
○ In synptomatic women, C. trachomatic serovar D-K cause mucopurulent cervicitis
and urtheritis
○ However, many women harbour the organism asymptomatically in the cervix
○ Not only a risk to their sexual partners anf offspring, but to themselves as, ascending
infection may occur
○ This results first in an endometritis in which chlamydiae survive monthly menstual
heddin of the uterine lining, followed by infection of the fallopian tubes to cause acute
salpingitis
○ Collectively, endometritis and salpingitis are known as PID which is largely caused
by C. trachomatis
○ Chalmydial pelvic infection may lead to further abdominal involvement and the
formation of pelvic adhesions
 Perihepatits (Fitz-Hugh-Curtis Syndrome) and peri-appendicitis may occur

5.Lymphogranuloma venerum
• Genital tract infection with C. trachomatis serovars L1-L3
• Usually begins with a genital ulcer followed by lymphadenopathy of the regional lymph
nodes
• Buboes are seen if infection persists, can spread to GIT and GUT causing strictures and in
rare cases, peno-scotal elephantiasis

Lab Diagnosis
• Cytoplasmic inclusions
○ Microscopic exam of clinical specimen
○ C.trachomatis inclusion bodies contain glycogen, can visualise _
• Antigens can be detected in exudates or urine by ELISA
• Pathogen can be grown in cell cultures
○ Chicken embryo yolk sac
○ McCoy cell lines

Treatment, Prevention and Control

• LGV tetracycline for 21 days
• Ocular and genital infections in adults should be treated with one dose of azithromycin or

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 • Ocular and genital infections in adults should be treated with one dose of azithromycin or
doxycycline for 7 days
• Treatment
○ LGV - doxycycline; aspirate buboes
○ Genital and ocular - azithromycin, doxycycline
○ New born conjunctivitis and pneumonia should be treated with erythromycin for
10-14 days
• Control
○ Antibiotic prophylaxis - oral azithromycin
○ HE
○ Tracing and treating partners in genital tract infection

Chlamydophila pneumoniae
• Spread via respiratoey secretion
• Human pathogen, no animal reservoirs
• Causes
○ Bronchitis
○ Pneumonia
○ Sinusitis
○ Atypical pneumonia
• 50% of people have serologic evidence
• A significant cause of acute exacerbation of asthma
• Diagnosis
○ Serolgy
○ PCR
○ Cell cultures
• Treatment
○ Macrolides
 Erythromycin
 Azithromycin
 Clarithromycin
○ Tetracyclines
 Tetrcycline
 Doxycycline
○ Levofloxacin administered for 10-14 days

Chlamydophila psittaci
• Infects wild and domestic birds
• Human infection
○ Inhalation of organism in dried infected bird droppings
• The incubation period is about 10 days
• Ranges from an 'influenza-like' syndrome with general malaise, fever, anorexia, sore
throat, headache and photophobia, toa severe illness with deliruima dn pneumonia
• Diagnosis
○ Serology (ELISA)
• Treatment
○ Tetracycline
○ Doxycycline
○ Erythromycin
• P/C
○ HE
○ PPE

Bacteriology Page 338

Mycoplasmas
Tuesday, 30 May 2017 3:52 PM

Important Properties
• Family:
○ Mycoplasmaceatea
• Genera
○ Mycoplasma
○ Ureaplasma
• Can grow in cell free culture media
• Multiply by binary fission
• They lack a cell wall
• Only bacteria that contain cholesterol in the cell membrane

Antigenic Properties
• Mycoplasmas have glycolipids
○ Can account for the neurological manifestations of M. pneumoniae infection
• Alter the I antigens on RBCs
○ Stimulate anti-I antibodies (cold agglutinins) - autoimmune response and damage to
erythrocytes
• Variable membrane lipoproteins form an antigenic variation system
○ Escape in the host immune system

Pathogenesis and Epidemiology

• Respiratory
• Urogenital

Respiratory Infection
• M.pneumoniae
○ Common cause of atypical _
○ Interfere with ciliary function
○ _
• M.hominis
○ Respiratory disease in new-borns
• M.fermentans
○ Throat
○ Associated with adult respiratory distress syndrome
• _

Urogenital Infections
• M.hominis
○ PID
○ Acute pyelonephritis
• Ureaplasma urealyticaum
○ Non-gonococcal urethritis

Lab Diagnosis

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Spirochetes
Tuesday, 6 June 2017 3:07 PM

• Spirochetes - derived from Greek word for coiled hair

• Bacterial cells are:
○ Extremely narrow
○ Elongated
○ Cylindrical
○ Spiral with tight coils with tapered ends
• Possess endo- or internal flagella
○ Number varies according to the species
• Actively motile
• Also exhibit rotational movements
• Possess some bacterial cell structural characteristics which are:
○ Similar to Gram-negative organisms
 Several characteristics
○ Different from Gram-negative organism
 Lack a cell wall LPS
 Inability to stain readily by commonly used stains
□ Few species are stainable by
 Modified Gram stain
◊ As gram-negative cells
 Wright's Stain

Genera of spirochetes associated with humans

a. Treponema
b. Borrellia
c. Leptospira

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Treponema
Thursday, 8 June 2017 12:41 PM

Treponema
• Composed of pathogenic and non pathogenic organisms
• Non pathogenic
○ Human bacterial normal flora mainly in the mouth and genital tract
• Saprophytic species
○ Reiter's strain of Treponema pallidum
 Antigenically related to pathogenic T. pallidum
 Can be grown in artificial cultures
 Requires anaerobic incubation
○ Nichol's strain of T. pallidum
 Both Reiter's and Nichol's strains are used in laboratory studies

Main pathogenic species of Treponema

• Previously:
○ Treponema palidum
○ T.pertenue
○ T.carateum
• Reclassified as subspecies of T. pallidum
• Other species associated with human dx include T. denticola as causative agent of
infections in the mouth
• All Treponema species or sub-species are similar in morphology and antigenic
composition
○ Cause cross reactions in serological tests
• Differentiated by
○ Geographical location
○ Clinical manifestation
○ Genetic characteristics

Treponema pallidum pallidum

• Too narrow to be visualised in the specimens by Gram's stain and light microscopy
• May use special methods
○ Dark field microscope
○ Fluorescence staining and microscopy
○ Special staining in infected tissues and microscopy
○ Electron microscope

Growth of T pallidum pallidum in the lab

• Doesn't grow on artificial culture media
• Live organisms can be inoculated in various parts of lab animals for propagation
○ Spectrum of manifestation of disease observed in infected humans really develops in
animals

Physical properties
• Loses viability rapidly when exposed to
○ Dry conditions
○ Heat
○ Low temperatures including 0-4°C for more than 2 days

Antigenic properties and Abs of T pallidum pallidum

• Exact antigenic components are not clearly identified

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 • Exact antigenic components are not clearly identified
• Distinct Abs are produced by immune system in response to infection including
○ Antibody which reacts with cardiolipin
 Cardiolipin is a lipid compound in beef heart muscle
□ Extracted and prepared for use in lab test
 Antibody referred to as
□ Wasserman's
□ Reagin
□ Lipoidophil
□ Anticardiolipin Ab
○ Antibody formed in response to a specific antigen contained within the treponemes
referred to as antitreponemal Abs

Clinical implications of T pallidum pallidum

• Natural infection is limited to humans
• Causative agent of syphilis
○ Characterised by severe manifestations and long term complications in then absence
of Rx

Transmission
• Depends on presence of organism in the blood and syphilitic lesions
○ Sexual - depends on stage of the dx
○ Vertical transmission
○ Blood transfusion

Sexual transmission
• Infection lesion in the majority is on the skin or mucosal surface of the external genitalia
• Entry is through broken skin or mucous membranes
• Multiplication occurs at the site of entry
• Incubation period follows during which the person is not infectious
• Manifestations
○ Change as disease progresses
○ Basis of classification of syphilis into
 Primary
 Secondary
 Tertiary
 Late

Primary Syphillis
• Characterised by
○ Localised invasion of mucus membranes
○ Relatively rapid multiplication of the organism
○ Initial dissemination through lymphatics and blood
• Manifestations occur within 10-90 days after exposure
• Main lesion is an ulcer referred to as the primary sore or chancre or hard chancre
○ Exudate contains T. pallidum pallidum

Chancre
• Starts as a papule
○ Commonest site is on the surface of external genitalia
○ Ulcerates and forms a painless ulcer with a clearly defined margins
• Single lesion, occasionally multiple
• Associated with painless enlarged inguinal lymph nodes in most cases
• Heals spontaneously in 3-8 weeks without Rx
○ Organisms are not necessarily eliminated

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Secondary syphilis
• Characterised by invasion of blood and widespread dissemination
• Generalised infection with varied manifestations including
○ Skin rash
○ Lesion of the mucous membranes
○ Ulcers in the mouth
○ Generalised lymph node enlargement
○ Small swellings described as plaques on various parts including the skin and
mucosal surfaces
 Can develop into wart like lesions referred to as condyloma lata
○ Uncommon manifestations include inflammatory processes involving bones, joints
and possible eyes
• Lesions of secondary syphilis contain numerous T pallidum pallidum
• Possible outcomes
○ Healing spontaneously
○ Latent syphilis
○ Progression to tertiary syphilis

Tertiary syphilis
• Develop approx. 3-10 years after the primary lesions
• Organism rarely detected in lesions
• Manifestations include small swelling on various parts due to chronic inflammatory process
on various tissues and organs including:
○ Skin
○ Mucous membranes
○ Bones
 Lesions referred to as gummas
 Ulceration of skin gummatous lesions

Late syphilis
• Manifestation 10-20 years after primary dx
• Involves mainly:
○ CVS - inflammation and other abnormalities mostly of the aorta including the aortic
valve
○ CNS
 Neurosyphilis
 Manifestations include
□ Abnormal gait
□ Trophic changes in joints
□ Abnormalities of optic nerves
□ Abnormal mental capacity
□ Involvement of meninges and blood vessels leading to more
complications

Latent syphilis
• Dormant dx without clinical manifestations
• Detectable by serological tests
• Capable of progression to CVS or neuro syphilis in the absence of Rx

Congenital syphilis
• Transmission can take place ate anytime throughout the pregnancy
○ As early as 10th week or as late as delivery
○ Associated with septicaemia in the foetus and widespread dissemination
• Manifestations
○ Death of foetus - miscarriage or still birth

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 ○ Death of foetus - miscarriage or still birth
○ Miscarriage
○ Developmental abnormalities
○ Baby born with latent infection or manifestations which develop any time within first 2
years including
 Failure to thrive
 Skin rash
 Nasal and other abnormalities

Lab investigations
• Specimens
○ Exudate from infected tissues
○ Blood
○ CSF
○ Infected tissue
• Procedures in specimen
○ Applied to detect the organism or Abs formed in response infection
 Detection of T pallidum pallidum
□ Unstained freshly collected fluid or scrapings from a chancre or ulcerated
lesions of secondary syphilis
 Examination by dark field microscopy
 Dark field microscopy
□ Performed promptly and repeatedly if necessary
□ Observable as a spiral organism with characteristic motility
 Fluorescence staining in exudates with Ab attached to fluorescent stain and
examination by fluorescent microscope
□ IFA
 Specific staining techniques in infected tissues and microscopic examination
• Serological tests
○ Detection of Abs in blood or CSF
○ Don’t distinguish pathogenic T pallidum pallidum or subspecies
○ Types of test
 Non-treponemal tests
□ Detect anticardiolipin antibody include
 VDRL
 Kahn test
 RPR
◊ Positive tests can be semi-quantified
◊ Suitable for quick screening
□ In non-treponemal tests
 Most sera become positive approx. 10-14 days after appearance of
chancre
 Associated with false positive and false negative results
◊ May be negative in early primary and late syphilis
 Positive in all patients with secondary syphilis
 Applied mostly as screening tests
◊ Positive results require confirmation
 Patients with serum testing positive become test negative after
successful treatment
◊ May be used to monitor response to treatment
 Treponemal tests
□ Detect the Ab against the specific antigen
□ Associated with fewer false positive reactions
□ Majority remain positive after completion of Rx
□ Tests include:
 T pallidum haemagglutination assay (TPHA)
 Fluorescent Treponemal antibody absorption test (FTA- ABS)

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  Fluorescent Treponemal antibody absorption test (FTA- ABS)
 ELISA
 T pallidum immobilisation test or TPI
◊ Has several disadvantages including use of live treponemes
 Other test including gene detection methods
• Antimicrobial susceptibility of T. pallidum pallidum
○ Susceptible to a wide range of antimicrobial agents
○ For treatment of infection, agents include:
 Penicillin - various preparations
□ Mostly used is benzathine penicillin
 Others are
□ Tetracycline, Erythromycin
 Choice, dose, duration and routes of administration all depend on
the general status of the patient and stage of the disease

Prevention od T. pallidum pallidum infection

• Sexually transmitted infection
○ Methods of prevention of STI and STD in general are applied
○ Screening clients with other STDs for syphilis and giving appropriate treatment if
infected
• Congenital syphilis
○ ANC screening of expectant females
○ Prompt diagnosis
○ Adequate treatment of infected expectant females
• Blood transfusion
○ Screening donated blood for T. pallidum before use and discarding if infected

Non-Venereal Disease Due to Treponema

• Transmitted through person to person contact or via contaminated artefacts
• Bejel or non-venereal or endemic syphilis
○ Caused by T. pallidum subspecies endemicum
○ Usually begins in childhood in the mucous membranes as a small patch which can
start in the mouth
○ Progresses gradually and develops into raised, eroding small swellings on the limbs
and trunks
○ Later lesions are associated with formation of gummas which occur on the skin,
bones and nasopharynx
○ Encountered in tropical and subtropical areas of Africa and other continents
• Yaws
○ Causative agent is T. pallidum pertenue
 Infects through broken skin
○ Numerous manifestations according to the stage of illness including
 Painless papular nodules initially
 Later develops into destructive lesions involving
□ Skin
 May form ulcers
□ Lymph nodes
□ Bones
□ Joints
□ Other destructive lesions on soft tissues
○ Encountered mostly in areas of South America, Central Africa and SE Asia
• Pinta
○ Causative agent is T. carateum or T. pallidum carateum
○ Primarily restricted to skin
○ Clinically characterised by
 Initial lesions as small pruritic papules

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 Initial lesions as small pruritic papules
 Later develops into enlarged plaques which persist for months to years
 Disseminated illness is characterised by recurrent hypopigmentation or
depigmentation of skin lesions and marked scar formation

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Actinomycetes
Thursday, 8 June 2017 11:25 AM

General Characteristics
• Gram positive bacteria with branching filaments
• Fungal-like organisms
○ Form a mycelia network of branching filaments like fungi but, like bacteria
 They possess cell walls containing muramic acid
 Have prokaryotic nuclei
 Susceptible to anti-bacterial antibiotics
• Related to mycobacteria and corynebacteria
• Non-motile
• Non-capsulated
• Non-spore forming
• Soil saprophytes but can cause chronic granulomatous infections in animals and man

Classification
• Family
○ Actinomycetaceae
• Based on the ability to grow aerobically or anaerobically
○ Anaerobic actinomycetes
 Actinomyces
 Bifidobacterium
 Eubacterium
○ Aerobic actinomycetes
 Nocardia
 Streptomyces
 actinomadura

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Actinomyces
Thursday, 8 June 2017 11:30 AM

• Gram-positive bacteria
• Actinomyces are facultatively anaerobic (except A. meyeri and A. israelii - obligate
anaerobe) however, grow best in anaerobic conditions
• Form branching filamentous rods
• Human commensal flora of the oropharynx, GIT and vagina
• Cause actinomycosis
○ An infrequent invasive bacterial disease

Actinomyces species
• Israelii
• Meyeri
• Odontolyticus
• Neuii
• Radingae
• Viscosus

• Israelii is an endogenous organism isolated from the mouths of healthy people

• However, can cause suppurative infection in patients with tooth abscess or a tooth
extraction

Pathogenesis
• The organisms have a low virulence potential
• Cause disease when the normal mucosal barriers are disrupted by trauma, surgery or
infection
• Actinomycosis is characterised by:
○ Multiple abscesses and granulomata
○ Tissue destruction
○ Formation of sinuses
• Note:
○ Within diseased tissues, they form large masses of mycelia (sulfur granules)

Clinical Manifestations
• Human actinomycosis takes several forms
○ Cervicofacial actinomycosis
 Most common type
 Occurs mainly in the cheek and submaxillary regions
 The disease is endogenous in origin
 Dental caries is a predisposing factor
 Infection follows a tooth extractions or other dental procedures
 Initially starts by soft-tissue swelling of the perimandibular area (lumpy jaw)
 Followed by development of sinus tracts that discharge purulent material
containing granules with a yellow sulfur like appearance
 Invasion of the cranium or the bloodstream may occur if the disease is left
untreated
○ Thoracic actinomycosis
 Commences in the lungs
 Usually secondary to aspiration of actinomyces from the mouth
 Pneumonitis develops that tends to invade the pleura and pericardium and
spread outwards through the chest wall
 Pulmonary actinomycosis may disseminate haematogenously
□ Brain abscess

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 □ Brain abscess
○ Abdominal actinomycosis
 The lesion is usually around the caecum and may involve the oesopharyngeal,
gastric and anorectal areas
○ Pelvic actinomycosis
 Second most frequent clinical form
 Occurs in women fitted with contraceptive IUD
 Other predisposing factors
□ Vaginal pessaries
□ Prolapse of the uterus
□ Septic abortion
 Presence of a pelvic/genital mass
○ Less common
 Actinomycoses of the CNS following
□ Thoracic or abdominal infections by haematogenous spread or from direct
dissemination of a cervicofacial lesion
 Bone and joint actinomycosis
 Cutaneous actinomycosis
□ Actinomyecetoma

Laboratory Diagnosis
• Specimen
○ Abscess content
○ Sinus discharge
○ Bronchial secretions
○ Biopsy
• Microscopy
○ Pus in saline to demonstrate sulfur granules
 White or yellowish
○ Crush the granules and perform gram and ZN staining
 Gram-positive filaments
 ZN
□ Non-acid-fast
• Culture
○ Thioglycollate liquid medium or on brain-heart infusion agar (BHI agar), BA
○ Incubate anaerobically at 35-37°C
○ Colonies
 Small
 Cream or white
 Have a rough nodular surface

Treatment
• Penicillin G
• Cephalosporins
• Amoxycillin

Prevention and Control

• Alternative methods of IUD
• Oral hygiene

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Nocardia
Thursday, 8 June 2017 11:55 AM

General Characteristics
• Gram-positive bacilli
• Aerobic
• Acid-fast with branching filaments
• Possess short chain mycolic acids
• Commensals in the oral cavity
• Transmission is by inhalation of airborne spores or mycelial fragments from environmental
sources or through trauma introduction

Species
• Asteroides
• Africana
• Brasiliensis
• Farcinica
• Ignorata
• Nova
• Veteran

Clinical Manifestations
• Predisposing factors
○ Underlying chronic lung disease e.g. asthma
○ Drug induced immunosuppression
○ Chronic granulomatous disease
○ Diabetes
○ HIV
• Cutaneous infection
○ Mycetoma
○ Lymphocutaneous infections
○ Cellulitis
○ Subcutaneous abscesses
• Systemic Nocardiosis
○ Manifests primarily ad
 Pulmonary disease
 Pneumonia
 Lung abscess
 Other lesions resembling tuberculosis
 Endocarditis
○ Occurs more often in immunodeficient persons

Laboratory Diagnosis
• Specimen
○ Bronchial washings
○ Bronchial lavage fluids
○ Sputum
○ Abscesses
○ Wound drainages
○ Tissues
• Microscopic examination
○ Gram stain
 Gram-positive filaments

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  Gram-positive filaments
○ ZN
 Acid-fast bacilli
• Culture
○ Nutrient agar or BHI agar
○ Incubated at 36°C for 3 weeks
○ Colony morphology
 Variable from cream, orange or pink coloured to chalky white appearance
□ Due to different species
• Identification
○ Biochemical test
 Hydrolysis of adenine, casein, tyrosine, xanthine, hypoxanthine
○ Serology
○ Molecular

Treatment
• Cotrimoxazole
• Amikacin
• Imipenem
• Linezolid
• Minocycline
• tobramycin

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Streptomyces
Thursday, 8 June 2017 12:06 PM

• Species
○ Streptomyces somaliensis
• Saprophytic soil organisms
• Cause mycetoma (actinomyecetoma)
• Few cases of invasive disease
○ Pre-existing condition
 cancer
 HIV
 Medical device

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Actinomadura
Thursday, 8 June 2017 12:09 PM

Actinomadura
• Spp
○ Madurae
• Soil saprophytes
• Cause myecetoma

Bacteriology Page 353

Actinomycotic myecetoma
Thursday, 8 June 2017 12:09 PM

• A localised chronic, granulomatous involvement of the subcutaneous and deeper tissues

• Affects the foot
• It presents as a tumour with multiple discharging sinuses

Causes of Bacterial Myecetomas

• Actinomyces
○ Israelii
○ Bovis
• Nocardia
○ Asteroides
○ Brasiliensis
○ Caviae
• Actinomadura
○ Madurae
○ Pellentierii
• Streptomyces
○ Somaliensis

Diagnosis
• Observe for colour of the granules
○ Antinomycotic myecetoma granules are white to yellow
○ Eumycotic myecetoma granules are generally black
○ Examination
• Isolation of the agent in culture establishes the diagnosis

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Introduction to Mycology
Wednesday, 9 November 2016 12:13 PM

Mycology
• Is the study of fungi
• Most are saprophytes in soil and on decaying plant material
• Used in industries e.g. production of bread, cheese, wine and beer
• Implicated in spoilage of fruits, grains, vegetables and jams
• Important human pathogens
• Fungi differ from bacteria in several aspects:
Feature Fungi Bacteria
Nucleus Eukaryotic Prokaryotic
Cytoplasm Mitochondria and ER Mitochondria and ER absent
present
Cell membrane Sterols present Sterols absent except
○ Mycoplasma
Cell wall content Chitin Peptidoglycan
Spores For reproduction For survival
Thermal Yes (some) No
dimorphism
Metabolism No obligate anaerobes Many obligate anaerobes

General Properties of Fungi

• Are eukaryotic
• Have ergosterols in their membranes
• Possess 80S ribosomes
• Have a rigid cell wall made of chitin
• Lack chlorophyll
• Typically non-motile, although a few e.g. Chytrids (non-pathogenic) have a motile phase
• Typically reproduce asexually and/or sexually by producing spores
• Fungi are responsible for fungal diseases called mycoses
○ Superficial/cutaneous mycoses
 Confined to the outer layers (keratinised) of the skin, hair or nails
○ Subcutaneous mycoses
 Confined to the subcutaneous tissue and rarely spread systemically
○ Systemic mycoses
 Involve the deep viscera

Structure of Fungi
• Fungi exist in three forms
○ Moulds
○ Yeast
○ Dimorphic fungi

Moulds/Molds
• Multicellular
○ Form branching filaments called hyphae
• A mass of hyphae collectively make up the mycelium
• There are two kinds of hyphae
○ Non-septate (coenocytic)/Aseptate

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 ○ Non-septate (coenocytic)/Aseptate
 No cross walls
○ Septate
 Hyphae divided into cells by cross-walls

Yeasts
• Unicellular
• Reproduces by budding
• Examples:
○ Candida albicans
○ Cryptococcus neoformans

Dimorphic Fungi
• Grow as yeasts or moulds depending on the environmental conditions and temperature
• The yeast form is found in infected tissue and the mould in the soil/environment
• Of medical importance
○ Blastomyces dermatitidis
○ Paracoccidioides braziliensis
○ Histoplasma spp.

Reproduction of Fungi
• Reproduce both asexually and/or sexually by producing spores
• Asexual spores are formed on or in specialised structures e.g.
○ Conidiospores
○ Sporangiospores
○ Chlamydospores
○ Blastoconidia (Blastospores)
• Sexual reproduction
○ Compatible nuclei unite within the mycelium and form sexual spores e.g.
 Ascospores
 Basidiospores
 Zygospores

Classification
• Medically important fungi are classified based on sexual function into:
○ Ascomycetes
○ Basidiomycetes
○ Deuteromycetes
 Only undergo asexual reproduction
○ Zygomycetes

Fungal Pathogenicity (Virulence Factors)

• Ability to adhere to host cells by way of cell wall glycoproteins
• Production capsules allowing them to resist phagocytosis
• Ability to damage host by secreting enzymes such as keratinase, elastase, collagenase
• Ability to secrete mycotoxins
• Exhibiting thermal dimorphism

Laboratory Diagnosis
• There are five approaches to the laboratory diagnosis of fungal diseases
○ Direct microscopic examination
○ Culture of the organism
○ Histology/cytology
○ DNA probe test and
○ Serologic tests

Mycology Page 356

 ○
• Specimens
○ Cutaneous mycoses
 Skin scrapings
 Nail clippings
○ Swabs from mucous membranes
○ Subcutaneous infections
 Crusts
 Aspirated pus
 Biopsies
○ Systemic infections
 Specimen from appropriate site
• Microscopy
○ Wet mount
 Yeast cells may be observed in urine wet mounts
○ 10-20% Potassium Hydroxide (KOH)
 Breaks down keratin and other bacterial organisms and you are left with the
fungal element
 Several specimens are subjected to KOH mount for direct examination
○ Lactophenol cotton blue stains the fungal elements blue
 After culture
○ Indian ink
 Important in diagnosis of Cryptococcal neoformans
○ Gram stain - take primary stain
 Yeats cells
• Culture
○ Media SDA (Saboraud Dextrose Agar)
 Has antibiotics important in preventing growth of bacteria and fungal
contaminants
○ Moulds identified by macro and microscopic morphology

Mycology Page 357

Superficial and Cutaneous Mycoses
Wednesday, 14 June 2017 3:49 PM

Superficial Mycoses
• Limited to the outermost layer of the skin or hair shaft
• Do not invade deeper tissues
• No cellular response from the host

Predisposing Factor
• Humidity
• Immunosuppression
• Poor hygiene

Infections
• Pityriasis/Tinea versicolour
• Black piedra
• White piedra

Tinea versicolour
• AKA Pityriasis versicolour
• Fungal infection of the epidermidis (particularly the stratum corneum) that manifests are
hypo-pigmented or hyper-pigmented skin patches (occur mostly in the chest and back)
○ Through lipid degradation which produces acid that damages melanocytes
• Caused by Malassezia furfur
○ These yeasts are lipophilic yeast
○ Live on the skin as part of the normal flora
○ Thrive in hot and humid conditions
• Note:
○ Neonates receiving total parenteral nutrition (with lipid infusions) are at risk of M.
furfur fungaemia due to M. furfur lipophilicity
• Lab diagnosis
○ KOH mount of skin scales - septate hyphae and budding yeast-like cells (spaghetti
and meatballs appearance)
• Treatment
○ Topical application of imidazoles
○ Topical application of Selenium sulfide (Selsun blue)
 Promotes exfoliation of stratum corneum
○ Oral treatment
 Ketoconazole or Itraconazole

Black Piedra
• Fungal infection of the hair shaft
• Caused by Piedra hortae
• Forming hard black nodules on the hair shafts
• Epidermics in families following the sharing of combs and hairbrushes
• Lab diagnosis
○ KOH preparation of the hair reveals a dark pigmented nodule surrounding the hair

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 ○ KOH preparation of the hair reveals a dark pigmented nodule surrounding the hair
shaft containing asci
• Treatment
○ Terbinafine

White Piedra
• Fungal infection of the hair shaft
• Caused by Trichosporon spp.
• Infected hairs develop soft greyish-white nodules along the shaft
• Lab diagnosis
○ KOH preparation of the hair reveals white or light brown nodules
• Treatment
○ Topical application of an imidazole

Cutaneous Mycoses (Dermatophytes)

• Involves skin, hair and nails
• Caused by fungi that invade only superficial keratinised tissue (skin nails and hair) not
deeper tissues
• Clinical manifestation is a ringworm or tinea
• Evoke a cellular immune response
• Infection range from mild to severe

Etiologic Agents
• Caused by fungi known as dermatophytes
• Dermatophytes are called keratinophilic (keratin loving)
• Produce extracellular enzymes (keratinases) which are capable of hydrolysing keratin
• Dermatophytes are classified into:
○ Microsporum
 Hair, skin, rarely nails
○ Trichophyton
 Hair, skin and nails
○ Epidermophyton
 Skin, nails and rarely hair

Ecology
• The dermatophytes may have different natural sources and modes of transmission
○ Anthropophilic
 These are usually associated with humans only
○ Zoophilic
 These are usually associated with animals
○ Geophilic
 These are usually found in the soil and are transmitted to man by direct
exposure

General Characteristic of Macroconidia and Microconidia

of Dermatophytes
Genus Macroconidia Microconidia
Microsporum Numerous, large, rough-walled and spindle shaped Rare

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Microsporum Numerous, large, rough-walled and spindle shaped Rare
Epidermophyton Numerous, smooth walled and club-shaped Absent
Trichophyton Rare, thin walled, smooth and pencil shaped Abundant

Clinical Classification
• Produce ring-like lesions (ring-worm or tinea)
• The clinical forms of the disease are named according to the site affected
○ Hairy areas
 Tinea capitis
 Tinea barbae
○ Skin
 Tinea corporis
 Tinea cruris (Jock itch)
 Tinea manum
 Tinea pedis (Athlete's foot)
 Tinea fascie
○ Nail
 Tinea ungium

Tinea Pedis (Athelete's Foot)

• It is an acute to chronic fungal infection of the feet
• Most prevalent of all dermatophytes

Tinea corporis
• AKA Tinea glabrosa
• Is a dermatophytic infection of the glabrous skin (external skin that is naturally hairless)

Tinea cruris
• Is an acute or chronic fungal infection of the groin, commonly called jock itch
• Often starts on the scrotum and spread to the groin as dry, itchy lesions
• Is often accompanied by athlete's foot which must be treated

Tinea manuum
• Ringworm of the palms and interdigits
• Common among patients with patients with tinea pedis
• Symptoms generally resemble that of tinea pedis

Tinea faciei
• Is a fungal infection of the face
• Tinea faciei is contagious just by touch and can spread easily to all regions of the skin
• Prone to secondary bacterial infections

Tinea barbae
• Is a fungal infection of the hair around the bearded area of men
• May be infected with bacteria

Tinea ungium (Onchomycosis)

• Nails become yellow, brittle and thickened
• May be caused by dermatophytes or candida

Tinea capitis
• Ringworm o the scalp, eyebrows and eyelashes
• Occurs in childhood and usually heals spontaneously

Mycology Page 360

Diagnosis of Dermatophytes
• Note the symptoms
• Note the kind of tissue attacked
• Observe proper collection of the specimen
• Keep specimen dry during transport
• Microscopic examination of slides of skin scrapings, nail scrapings and hair

Lab Diagnosis of Dermatophytoses

• Specimen
○ Skin scales
○ Nail
○ Hair clippings
• Macroscopic exam (Wood lamp)
○ Detect fluorescence in infected hair, scalp lesions
• Direct microscopy
○ Specimen placed on a slide + drop of 10-20% KOH
○ Observe for:
 Hyphae or pores in the hair shaft
 The spore may be done outside the hair shaft (ectothrix) or inside the hair shaft
(endothrix)
 Skin and nails
□ Branching hyphae or chains of arthrospores are seen
• Cultures on SDA incubated at room temperature for up to 4 weeks
○ Colonies are identified by morphology and colour on surface and reverse, and by
microscopic examination using lactophenol cotton blue or other stains

Treatment
• Topical
○ Non-specific - Whitfield's ointment
○ Specific - creams, lotions, shampoos or azole derivatives -
Clotrimazole,Ketoconazole etc.
• Oral antifungals
○ Griseofulvin
○ Terbinafine
○ Itraconazole

Prevention
• jifikiririe

Mycology Page 361

Yeasts
Wednesday, 14 June 2017 4:35 PM

Introduction
• Unicellular fungi - spherical, oval, elongated
• Reproduce by budding
• Gram positive
• Those of medical importance
○ Candida
○ Cryptococcus

Mycology Page 362

Candida
Friday, 16 June 2017 5:06 AM

Candida
• Grows as a yeast
• May form pseudohyphae (pathogenic)
• Gram positive
• >150 species
○ C.albicans - most common

Epidemiology
• Normal microbiota for humans
• They usually inhabit oropharyngeal, intestinal and female genital tract
• Associated with endogenous infections
• Predisposing factors
○ Immunosuppression
 HIV infection
 Use of steroids
 Pregnancy
○ Use of anti-biotics
○ IV catheters/prosthetic implants/GIT surgery
○ Diabetes
○ Oral contraceptive use or oestrogen therapy

Clinical Manifestations
• Mucocutaneous
○ Oral thrush
○ Vaginitis
○ Chronic mucocutaneous candidiasis(CMC) in patients with T-lymphocyte
immunodeficiencies
• Cutaneous
○ Skin
 Groin
 Vulva
○ Napkin area in infants (diaper rash)
○ Interdigital clefts, nails, skin folds around the nails - hands and feet frequently in H2O
• Systemic
○ Urinary tract
○ GI
○ Pulmonary
○ Meningitis
○ Ocular
○ Candidemia or disseminated candidiasis

Lab Diagnosis
• Specimen
○ Depends on site of infection
 Blood
 CSF
 Swabs
 Etc.
• KOH or Gram stain or exudates and tissue
○ Budding yeasts and pseudohyphae

Mycology Page 363

 ○ Budding yeasts and pseudohyphae
• Culture
○ On BA or SDA
○ In air, 37°C, 12-24 hours
○ Appearance
 Off-white colonies with a yeast smell
• Germ tube test
○ Positive - presumptive diagnosis of C. albicans
• Carbohydrate assimilation and fermentation
• Molecular diagnostics
• Susceptibility testing
○ Candida is very resistant to many anti-fungal agents

Treatment
• Superficial
○ Topical
 Nystatin
 Azole preparation
• Chronic mucocutaneous infections
○ Fluconazole
• Disseminated or relapsing infections
○ Amphotericin B
○ Fluconazole
○ Flucytosine

Mycology Page 364

Cryptococcus
Friday, 16 June 2017 5:07 AM

Cryptococcus
• Encapsulated yeast-like organism
• Reproduces by budding
• Produce whitish mucoid colonies in culture
• Possess urease
• C.neoformans - medical importance

Epidemiology
• Environmental
○ Isolated mainly from pigeon droppings and nesting places
○ Contaminated soil
• Serotypes
○ A and D - worldwide
○ B and C - tropics and sub-tropics

Risk Factors
• More common in patients with defects in T-cell mediated immunity
• HI/AIDS
• Prolonged steroid therapy
• Organ transplantation

Clinical Manifestations
• Pulmonary cryptococcosis
○ Usually asymptomatic
• CNS disease
○ Meningitis
○ Meningoencephalitis
○ Cryptococcoma
• Cutaneous cryptococcosis
• Bone and joint disease
• Ocular cryptococcosis - rare

Lab Diagnosis
• Specimens
○ CSF, check for
 ↑ pressure
 ↑ protein
 ↓ glucose
 ↑ lymphocytes
○ Blood
○ Sputum
○ Biopsy
• Indian ink or nigrostaining of CSF deposit
• Latex agglutination tests (Cryptococcal Antigen - CRAG test)
○ On CSF or serum
○ Detects polysaccharide capsular antigen
○ Rapid test
• Culture of CSF or other specimen on:
○ SDA
 37°C, in air

Mycology Page 365

  37°C, in air
 Appearance
□ Creamy, mucoid colonies
○ Selective media
 Birdseed agar
□ Appearance
 Melanin production (brown colonies)
• Biochemical tests
○ Urease positive
○ Carbohydrate assimilation and fermentation tests

Treatment
• Amphotericin B - DOC
• Flucytosine - combined with Amphotericin B
• Fluconazole

Mycology Page 366

Subcutaneous Mycoses
Thursday, 15 June 2017 11:22 AM

Conditions
• Myecetoma
• Phaeohyphomychosis
• Chromoblasotmycosis
• Sporotrichosis
• Lobomycosis
• Rhinosporiodiosis

Treatment of Subcutaneous Infections

• Supportive
○ Secondary infections
○ Pain etc.
• Surgery for some lesions e.g. myocetoma, amputation is most effective
• Anti-fungal agents
○ Amphotericin B
○ Itraconazole
○ Flucytosine
○ Other azoles
• Antibacterial agents in actinomyecetoma and in secondary bacterial infection
○ Sulphonamides
○ Rifampicin
○ Streptomycin
○ Amikacin

Mycology Page 367

Myecetoma (Madura Foot)
Friday, 16 June 2017 5:08 AM

Myecetoma (Madura Foot)

• Infection is acquired following trauma to the skin by plant material from trees, shrubs or
vegetation debris
• Infection is very chronic and takes months to be fully established and years to deal with
• It is not contagious
• Aetiologies are fungi which cause eumycotic myecetoma (Eumycetoma) or Actinomycotoc
myecetoma (actinomyecetoma)
• Aetiology
○ Eumycetoma
 Caused by several mould fungi
 The colour of grains in this type of mycetoma is black or white
 Fungi include
□ Madurella
 Madurella mycetomatis causes the majority of the cases with black
grains
 Madurella grisea
◊ Black/grey grains
□ Pseudallescheria
□ Acremonium
□ Leptosphaeria

Actinomyecetoma
• Main aetiologies
○ Streptomyces somaliensis
 Causes the majority of the cases
 Colour of grains telloe to yellow-brown
○ Actinomadura madurae
 White or yellow grains
○ Actinomadura pelletieri
 Pinkish-red grains
○ Nocardia brasiliensis
 White grains
○ N.Asteroides, N.caviae, N.codiaca
 White or yellow grains
• Chronic localised subcutaneous infection that involves inflammation of tissues caused by
infection with a fungus or with certain bacteria
○ The lesions present with multiple abscesses
○ Main factor
 Fermentation of sinuses that drain pus to the surface of the skin and presence
or grains
○ Grains are granules (small colonies) about 1-2 mm diameter, of the aetiologic agent
with different colour
○ The commonly affected site is the foot, however, it can in legs, thigh, hand, shoulder
or head
• Management
○ Usually actinomycetoma respond better than eumyecetoma
 Geneally if bone is infected the response to treatment is poor
○ Actinomycetoma
 Cotrimoxazole
 Penicillin G

Mycology Page 368

  Penicillin G
○ Eumyecetoma
 Azole derivatives e.g. Ketoconazole
○ If drugs not effective and bone is infected
 Amputate the limb or debride tissue
○ Treatment duration is long - up to years

Mycology Page 369

Phaeohyphomycosis
Friday, 16 June 2017 5:08 AM

Phaeohyphomycosis
• Caused by pigmented filamentous fungi, which contain melanin in their cell walls (melanin in cell walls
may b a virulence factor)
• The pathogens are considered opportunistic, most cases disseminated infection have occurred in
immunosuppressed patients
• Clinical signs consist of nodules underneath the skin, abscesses or cysts. In extreme cases, there were
deep infections within the eyes, bones, heart and CNS
• Morphologic characteristics in tissue include hyphae yeast-like cells
• Current anti-fungal agents
○ Posaconazole
○ Voriconazole
• Aetiology
○ Dematiaceous imperfect mould fungi, mainly
 Cladosporium
 Exiophiala
 Wangiella
 Cladophialophora bantiana
○ Naturally in woody plants, wood and agricultural soils
• Lab diagnosis
○ Specimen
 Pus
 Biopsy tissue
○ Direct microscopy
 KOH and smears
□ Brown septate hyphae
 Culture
□ On SDA
 Very slow growing
 Black or grey colonies

Mycology Page 370

Chromoblastomycosis
Friday, 16 June 2017 5:08 AM

Chromoblastomycosis
• A.k.a
○ Chromoblastomycosis
○ Cladosporiosis
○ Fonseca's Disease
○ Pedroso's Disease
○ Verrucous dermatitis
• Fungi implanted under the skin by thorns or splinters
• Clinical manifestations
○ The infection site has a small red papule
○ Infection slowly spreads to the surrounding tissues through the blood vessels or
lymph vessels, producing metastatic lesions at distant sites
○ Secondary infection with bacteria may lead to lymph stasis (obstruction of the lymph
vessels) and elephantiasis
○ The nodules may become ulcerated, or multiple nodules may grow and coalesce,
affecting a large area of a limb
• Lab diagnosis
○ KOH preparation reveals sclerotic cells
○ Culture to identify the organism involved
○ On histology, chromoblastomychosis manifests as pigmented yeasts resembling
"copper pennies". Special stains, such as periodic acid schiff and Gömöri
methanamine silver
• Treatment
○ Agents of choice include:
 Itraconazole
 Terbinafine
 Flucytosine
○ Alternatively
 Cryosurgery with liquid nitrogen
○ Antibiotic agent used to treat bacterial superinfection
• Clinical
○ The lesions are
 Hyperkeratotic
 Cauliflower
 Initially ulcerative
○ Affected sites
 Extremities mainly hands and feet
• Aetiology
○ Mould fungi in woods and woody plants
○ Include
 Phialophora verrucosa
 Fonsecaea pedrosi
 Exophiala
 Cladosporium
○ Lab diagnosis
 Specimen
□ Biopsy tissue
 Direct microscopy
□ KOH and smears
 Brown cells with brown septa
 Brown muriform cells = sclerotic bodies

Mycology Page 371

  Brown muriform cells = sclerotic bodies
 Culture
□ On SDA and mycobiotic
 Very slow, demitiacious fungi

Mycology Page 372

Rhinosporidiosis
Friday, 16 June 2017 5:10 AM

Rhinosporidiosis
• Is a granulomatous disease affecting the mucous membrane of
○ Nasopharynx
○ Oropharynx
○ Conjunctiva
○ Rectum
○ External genitalia
• Floor of the nose and inferior turbinate are the most common sites
• Laryngeal rhinosporidiosis may be de to inoculation form the nose during endotracheal
intubation
• After inoculation, the organisms replicates locally, resulting in hyperplasia of host tissue
and localised immune reaction
• History
○ History of exposure to contaminated water
○ Unilateral nasal obstruction, local pruritus at site of inoculation, coryza (rhinitis) with
sneezing
○ Post nasal discharge with cough and foreign body sensation
○ Increased tearing and photophobia in cases of infection in the conjunctiva
• On examination
• Medical treatment is not so effective but treatment with a year long course of Dapsone has
been reported
○ Pink to deep red polyps
○ Strawberry like appearance
○ Bleeds easily upon manipulation
• Diagnosis
○ Confirmed through biopsy and histopathology
 Several oval or round sporangia and spores which may be seen bursting
through its chitinous wall
• Management
○ Surgical incision
 Excision with electro-coagulation of the lesion base
• Summary
○ Clinical
 Mucocutaneous fungal infection
 Sites
□ Nasal
□ Oral (palatte, epiglottis)
□ Conjunctiva
 Lesion
□ Polyps
□ Papillomas
□ Wart-like lesions
 More seen in communities near swamps
○ Aetiology
 Rhinosporidium seeberi
 Obligately parasitic fungus
○ Lab diagnosis
 Specimen
□ Biopsy tissue
 Direct microscopy
□ Stained sections or smears or KOH, will show spherules with endospores

Mycology Page 373

 □ Stained sections or smears or KOH, will show spherules with endospores
 Culture
□ On SDA
 Negative
○ Management
 Cryosurgical excision of the lesion
□ Relapse is common

Mycology Page 374

Sporotrichosis
Friday, 16 June 2017 5:10 AM

Sporotrichosis
• Clinical
○ A chronic fungal infection producing nodules and ulcers in the nodules on the skin
○ Characteristically follows lymphatic pathways
○ Affected sites
 Extremities
 Joints
• Aetiology
○ Dimorphic fungus in trees, shrubs, plant debris
○ Includes
 Sporothrix schenckii
• Lab diagnosis
○ Specimen
 Biopsy tissue
 Ulcerative material
○ Direct microscopy
 Smear
□ Finger like yeast cells or cigar shaped. Some are oval
○ Culture
 On SDA
□ At room temperature to grow mould
□ Blood agar at 37°C to grow the yeast

Mycology Page 375

Lobomycosis
Friday, 16 June 2017 5:11 AM

• (Jorge) Lobo' disease, Lacaziosis

• Other names which were given to the disease are:
○ Keloidal blastomycosis
○ Amazonian blastomycosis
○ Blastomycoid granuloma
○ Miralp
○ Piralp
• These last two names were given by the natives of the Amazon and means that which
burns.
• Caused by Lacazia loboi
○ Obligate parasitic fungus
• The disease is usually found in humans and bottle-nosed dolphins, with the possible risk
of transmission from one species to another
• Human-to-human transmission does not occur and the disease is only acquired from the
environment
• The appearances of a chronic keloidal nodular lesion occur on the face and ears
• Diagnosis
○ A skin biopsy, examining it under a microscope
○ Log chains of spherical cells, interconnected by tubules
○ The cells appear to be yeast-like
○ Attempts to culture have been unsuccessful
• Treatment
○ Surgical excision or cryosurgery is DOC
○ Itraconazole used to prevent recurrence

Mycology Page 376

Systemic Mycoses
Thursday, 15 June 2017 12:05 PM

• Fingal infections or disease which involve ti inner tissues including various organs
• May be grouped as :
○ Infections caused by a group of fungi referred to as true pathogenic fungi
 Include:
Disease Causative organism
Histoplasmosis Histoplasma capsulatum
□ Coccidiomycosis Coccidiodes immitis
Blastomycosis Blastomyces dermatitidis
Paracoccidiodomycosis Paracoccidiodes brasiliensis
○ Opportunistic systemic fungal infections
 Causative agents are naturally saprophytes or normal human flora
□ Fungi of very low virulence
 Disease production depends on abnormal human immunological defense
mechanisms
 Include
□ Candidasis
□ Cryptococcosis
□ Mucormycosis
□ Aspergillosis
□ Penicilliocic
□ Pneumocystic pneumonia

Mycology Page 377

 Systemic Infections Due to True Pathogenic Fungi
Friday, 16 June 2017 5:16 AM

Systemic infections due to true pathogenic fungi

• Involve immunocompetent as well as immunosuppressed patients
• Majority have restricted geographical distribution
○ Mostly encountered in parts of America
• Causative fungi
○ Exhibit two morphological form (Dimorphic phenomenon)
 Filamentous fungi or moulds in the saprophytic state or when grown at a
temperature of 25°C
 Yeasts in infected tissues and specimens or when grown at 37°C
 Dimophism is a result of several factors including temperature at which growth
occurs
□ Partly temperature dependent
○ Majority infect humans through inhalation of spores into the lungs leading to a
primary infection

Primary Infection Due to Dimorphic Fungi

• Majority remain asymptomatic and a minority is symptomatic
• Some asymptomatic infections progress and develop into symptomatic illness after several
years
• Symptomatic illness is characterised by
○ Local
○ Lymphatic
○ Haematogenous spread to other parts including the skin, mucous membrane,
subcutaneous tissues and bone

Lab Investigation of Systemic Mycoses Due to Dimorphic Fungi

• Specimens
○ Highly infections
 Necessary safety methods required in specimen handling and laboratory
procedures
 Include:
□ Sputum
□ Discharge or pus
□ Skin scrapings
□ Infected tissue
□ Blood for serology
• Procedures
○ Detection of the yeasts in specimens by microscopy of potassium hydroxide mount
or special methods of tissue
○ Culture
 For isolation and identification
 Culture on Sabaraud's Medium
□ Incubation at 25°C for filamentous or mould fungus
□ Colonial features are noted and microscopic characteristics are noted and
microscopic characteristics are demonstrated by Lactophenol blue stain
and microscopy
 Culture to demonstrate dimorphism
 Isolated filamentous fungus is sub-cultured on a suitable medium and
incubated at 37°C for the yeast phase
• Serological tests

Mycology Page 378

 • Serological tests
○ Useful in investigation of classical histoplasmosis coccidioidomycosis and
paracoccidioidomycosis
○ Associated difficulties in
 Differentiation of past exposure and current illness
 Cross-reactions
• Detection of antigens of the various fungi
• Molecular laboratory techniques

Management Methods of Systemic Mycoses due to Dimorphic Fungi

• Based on several factors including the
○ Manifestations
○ Predisposing factors
○ Causative agents
• Methods include
○ Surgical procedures
○ Antifungal agents for treatment
 Amphotericin B - main agents
 Others
□ Itraconazole
□ Fluconazole

Mycology Page 379

Histoplasmosis
Friday, 16 June 2017 5:32 AM

• Causative agent is Histoplasma capsulatum

• Varieties of the species associated with disease in humans are
○ Histoplasma capsulatum capsulatum
○ Histoplasma capsulatum duboisii
• Dimorphic fungus
○ Mould in the cold
○ Yeast in the heat
• The two varieties:
○ Are similar morphologically in cultures and microscopy of the mould
○ Yeast cells or tissue phases are significantly different morphologically
○ Differ in clinical manifestation of infections
○ Differ in geographical distribution
H.capsulatum capsulatum H.capsulatum duboisii
Size Smaller yeast cells Larger yeast cells
Geographic Endemic to America Mostly found in Africa
location
○
Clinical ○ Chronic lung disease with cavity ○ Ulcerative lesions or chronic
manifestations formation inflammatory swelling which involve:
○ Disseminated illness involving other  Skin
tissues and internal organs  Subcutaneous tissue
 Bones

Histoplasma capsulatum capsulatum

• Saprophyte mostly found in soil contaminated with birds or bat dropping
○ History of travel to caves (spelunking) and contact with chicken coops
• Causative agent of classical histoplasmosis
○ Encountered in all parts of the worlds
 Highly endemic in parts of America
• Majority result from reactivation of asymptomatic primary infection
○ Reactivation may be associated with other underlying diseases
• Pathogenicity
○ Contracted by inhaling the spores which are then taken up by macrophages
○ Taken up by alveolar macrophages where they replicate
○ Within the macrophages, the cells are able to disseminate to RES organs
○ Initial inflammatory reaction become granulomatous
• Manifests mostly as:
○ Chronic lung disease with cavity formation
 Formation of granulomas which eventually calcify
 Resembles TB in that
□ Cavity formation in the upper lobes
□ Calcified nodules with fibrotic scarring
○ Disseminated illness involving other tissues and internal organs
• Opportunistic classical illness may manifest as
○ An acute disseminated disease
○ Chronic illness involving multiple organs including the liver and spleen
 Affect mainly the immunocopromised and may manifest as hepatosplenomegaly
 Gross pathology will show calcifications in the liver and spleen
 This is because the fungus targets the RES, which has numerous macrophages and are
prevalent in the liver and spleen
• Microscopic examination of infected tissues shows H. capsulatum capsulatum as small intra-cellular
budding yeasts inside different types of phagocytic cells
○ Macrophages with intracellular oval bodies
• Filamentous organism in culture is identified by specific microscopic and macroscopic features
○ Microscopic examination shows macroconidia which are spherical with finger -like spines described
as tuberculate macroconidia

Histoplasma capsulatum duboisii

• Causative agent of African histoplasmosis
○ Encountered more in specific countries in central part of Africa
○ Clinical manifestations include ulcerative lesions or chronic inflammatory swelling which involve
 Skin
□ Erythema nodosum
 Painful red nodules usually found on the shins
 Subcutaneous tissue

Mycology Page 380

  Subcutaneous tissue
 Bones
○ Organism is observed in specimens as yeast cells which are larger than those of H. capsulatum
capsulatum
• Lungs are not commonly infected
• Source and route or routes of infection are not clearly identified

Diagnosis
• Specimen
○ Tissue biopsy
 Scrapings from superficial lesions
○ Respiratory specimen
○ Urine
○ Blood
○ Bone marrow aspirates
• Procedure
○ Microscopy
 Stain sample with KOH preparation
□ Macrophages from infected tissue with intracellular oval bodies
□ The oval bodies are much smaller than an RBC
○ Culture
 Often takes a long time (4 weeks) and is therefore inconvenient
 Glucose cysteine agar
□ At 37°C
 SDA or IMA
□ At 25 - 30°C
○ Serology
 Rapid histoplasma urine or serum antigen tests
□ Histoplasmin antigen

Treatment
• Depends on the status of infection
○ Local and mild infections
 Azoles
□ Fluconazole
□ Ketoconazole
○ Systemic infection
 Amphotericin B
□ Only if the fungus disseminates which generally happens in the immunocompromised

Mycology Page 381

Coccidioidomycosis
Friday, 16 June 2017 5:32 AM

Coccidioidomycosis
• Dimorphic fungus
○ Mould in the cold
○ Spherule of endospores in the heat (as opposed to yeast in the heat)
• Routes of transmission
○ Coccidiodes immitis spores are inhaled by humans from the saprophytic fungus in
the lungs
 Chance of infection increases the more dust is in the air
□ Dust storms
□ Earthquakes
 Develop into spherical large structures referred to as spherules (larger than
RBCs ∴ Cocci > Blasto > Histo)
 Each spherule contains
□ Spores
□ Develops and ruptures releasing spores to the adjacent tissues and each
released spore develops into a spherule
• Clinical Manifestations
○ Generally seen in immunocompetent people
 Mostly asymptomatic or mild illness of the respiratory tract including mild
pneumonia
□ Accompanied by
 Fever
 Cough
 Arthralgia
 Erythema nodosa
□ Extremely tender nodules especially on the shins
○ Occasionally occurs as a severe illness
○ Severe illness is characterised by cavities in the lungs
 Radiographic images may show nothing or these cavities or nodules
○ Dissemination to extra-pulmonary sites is uncommon
 Except in the immunocompromised - disseminates to the bone
○ In immunocompromised
 Skin and lungs are the most common sites of infection
 May disseminate to the bones and mininges to cause meningitis
• Diagnosis
○ Procedures
 Culture
□ KOH stain preparation
□ But grows very slowly
 Serology
□ Antibody titres against the fungus
 IgM for recent infection
• Treatment
○ Local lung infections
 Azoles
□ Ketoconazole
○ Systemic infection
 Amphotericin B

Mycology Page 382

Blastomycosis
Friday, 16 June 2017 5:33 AM

• Endemic in parts of America also isolated in parts of Africa and Asia

• Dimorphic fungus
○ Mould in the cold
○ Yeast in the heat
• Pathogenicity
○ The spores from the mould in the soil have the potential to become aerosolised and
inhaled into the lungs and transform into yeast in the body
○ Replicates by single, broad based budding
• Manifestations of disseminated illness include
○ Acute of chronic pneumonia and is considered a localised infection
 Lungs have a patchy, alveolar infiltrate with lesions and cavities on x-ray
○ Can spread to other organs especially in the immunocompromised
 Most likely to occur in the skin or bone (osteomyelitis)
 Chronic cutaneous swellings on exposed parts and involvement of other
tissues
□ Swellings usually ulcerate
• Blastomyces dermatitidis yeast cells in infected tissues are identified by observation of
each cell producing a single bud

Diagnosis
• Specimen
• Procedures
○ Microscopy
 KOH preparation
□ The yeast cells are about the same size as RBCs
□ Round yeast cell with a single broad-based bud
○ Serology
 Urine antigen test

Mycology Page 383

Paracoccidioidomycosis
Friday, 16 June 2017 5:33 AM

• Dimorphic fungus
○ Mould in the cold
○ Yeast in the heat
 The yeast form shows a captain's wheel appearance (the steering wheel of a
ship)
 Roughly the same size as the Coccidioides spherules
 Paracoccidioides brasiliensis is observable microscopically in infected tissue
specimens as large yeast cells with each cell producing multiple buds
• Routes of transmission
○ Respiratory droplets
○ After inspiration, the fungus disseminates into lymphatics causing lymphadenopathy
(cervical, axillary or inguinal)
 Has a tendency to involve the lymphatics
 Spreads via the lymphatic and haematogenous routes
○ Progresses to the lungs causing granulomas
• Clinical manifestations
○ Manifestations include ulceration of the mouth (in the gums with ragged edges and
small spots of haemorrhage) and nose with extension to the local lymph nodes
• Treatment
○ Itraconazole
 Mild infection
○ Amphotericin B
 Severe infection

Mycology Page 384

Opportunistic Systemic Mycoses
Thursday, 15 June 2017 12:39 PM

Opportunistic systemic fungal infections

 Causative agents are naturally saprophytes or normal human flora
□ Fungi of very low virulence
 Disease production depends on abnormal human immunological defense mechanisms
 Include
□ Candidasis
□ Cryptococcosis
□ Mucormycosis
□ Aspergillosis
□ Penicilliocic
□ Pneumocystic pneumonia

Mycology Page 385

Mucormycosis (Zygomycosis)
Friday, 16 June 2017 5:14 AM

Mucormycosis or Zygomycosis
• Uncommon opportunistic fungal infection
• Predisposing condition include
○ Metabolic derangements including acidosis
○ Other illnesses or treatment which suppress the immune responses to infection
○ Trauma
• Causative fungi are saprophytes which release spores in the air and dust
○ Include species of genera
 Rhizopus
 Absidia
 Mucor
• Acquired through inhalation of spores
• Manifestations are according to the sites involved
○ Commonest initial lesion is a swelling of variable size
○ Develops rapidly as the causative organisms grow rapidly and invade tissues and
blood vessels leading to necrosis and thrombosis
○ Also manifests as a severely rapidly progressing systemic illness
○ Local or haematogenous spread is a characteristic
○ Infection of the head and neck region results in a variety of manifestation may
involve orbital area, part of the face or nasal disease
 Including rhinocerebral zygomycosis
• Other tissues infected include
○ Lungs
○ GIT
• Lab diagnosis
○ Specimens
 Discharge from lesions
 Tissue
 Sputum
○ Procedures
 Microscopy for observation of characteristic hyphae
 Culture
□ Rapidly growing fungi with cottony colonies
□ Microscopy shows characteristic sporangia
• Management
○ Include
 Surgical methods
 Amphotericin B
 Treatment of predisposing condition

Mycology Page 386

Penicillinosis
Friday, 16 June 2017 5:13 AM

Penicillinosis
• Causative agent is Penicillium marneffei
○ Uncommon opportunistic dimorphic fungus encountered in parts of SE Asia
○ Route of infection is not clearly defined
• An opportunistic infection in the immuno-compromised people
• Disseminated disease is characterised by
○ Fever
○ Weight loss
○ Skin lesions including ulcers
○ Abscesses
○ Enlarged lymph nodes and organs
• Lab investigations
○ Involves
 Demonstration of yeast cells intra-and extracellularly in specimens
 Appropriate cultures for isolation an identification
• Antimicrobial susceptibility
○ Itraconazole
○ Voriconazole

Mycology Page 387

Aspergillosis
Friday, 16 June 2017 5:13 AM

Aspergillosis
• A group of diseases in which species of the genus Aspergillus are involved
• May or may not be opportunistic
• Aspergillus
○ Genus composed of numerous species
○ Majority are saprophytes not involved in human disease
○ Species and their spoors are found in various places in the environment
 Air
 Soil
 Decaying vegetation
 Organic debris
 Construction and demolition sites
○ Chances of contamination of items and inhalation of airborne spores are increased

Aspergillus Species Commonly Associated With Human Disease

• Include
○ A.fumigatus responsible for the majority
○ A.flavus produces a mycotoxin which damages various internal organs
○ A.niger
• Mechanism if disease causation
○ Include
 Colonisation
 Invasion of tissues
 Hypersensitivity reaction to the spores
 Effects of released toxin on the tissues
• Manifestation of Aspergillosis
○ Depend in the species and mechanism of causation and parts of the human or
systems involved

Aspergillosis Involving the Resp. System

• Basically three categories of disease
○ Allergic aspergillosis
 Manifestations are due to hypersensitivity reaction to the inhaled spores
 May manifest as an asthmatic attack of alveolitis
○ Colonising aspergillosis
 Acquired by inhalation of Aspergillus spores
 Association with colonisation of pre-existing lung cavities by the inhaled
species
 Tends to be associated with cavities from previous pulmonary tuberculosis
 Aspergillus grows and forms a mycelial mass, also referred to as a fungal ball
or aspergilloma
○ Invasive aspergillosis
 Occasionally encountered in patients with chronic debilitating illnesses
 Characterised by rapid growth and invasion by Aspergillus species resulting in
destruction of lung tissue and blood vessels

Other Types of Aspergillosis

• Nasal-orbital aspergillosis
○ Encountered as Aspergilloma in the air sinuses including the maxillary sinus
○ A.fumigatus and A. niger are identified as possible causative species
• Cutaneous aspergillosis

Mycology Page 388

 • Cutaneous aspergillosis
○ Often associated with the external ear as otomycosis
 May be caused by A. niger or A. fumigata
• Mycetoma
○ Encountered rarely
○ Associated with A. nidulans and A. glaucus

Laboratory Investigation of Aspergillosis

• Involves
○ Detection of aspergillus in specimens by
 Microscopic examination of KOH mount for specific morphological features of
Aspergillus
 Cultures for isolation
 Identification
□ As genus Aspergillus
□ Of species to enable differentiation of contaminating saprophytes from
possible pathogens

Identification of Aspergillus Species

• Can be achieved by
○ Examination of culture
 Colonial morphology and other notable characteristics
 Microscopic features
 A.fumigatus is identified as above and also by ability to grow at 43°C
 Species which are not common pathogens might require repeated isolation for
conclusive results
○ Serology
 Applicable in the investigation of Aspergilloma

Management of Aspergillosis
• Hypersensitivity reactions are managed accordingly
• Aspergillosis due to infection by Aspergillus
○ Anti-fungal agents administration
 Amphotericin B
 Others including
□ Voriconazole
□ Caspofungin
□ Itraconazole
 Choice depends on
□ Type of Aspergillosis and effectiveness of the agent
□ General status of the patient
□ Availability
○ Other measures including surgical procedures and other chemotherapeutic agents

Mycology Page 389

Urinary Tract Infections
Thursday, 22 June 2017 4:41 AM

• UTIs are common, higher incidence is among women

• Incidence increases in old age
• UTI infection usually occurs by ascending route (urethra to bladder)
• UTI infection is less common by haematogenous spread
• They occur in two general settings
○ Community acquired
○ Nosocomially acquired

Definitions
• Dysuria
○ Burning pain on passing urine
• Bacteriuria
○ Presence of bacteria in urine
• Pyuria
○ Presence of pus in urine
• Upper UTI
○ Infection above the level of the bladder
• Lower UTI
○ Infection at or below the level of the bladder
• Uncomplicated UTI
○ Infection in a structurally and neurologically normal urinary tract
• Complicated UTI
○ Infection in a urinary tract with functional or structural abnormalities
• Urethritis
○ The inflammation or infection is limited to the urethra
○ It is usually an STI
• Cystitis
○ Irritation in the bladder
○ Indicated by:
 Dysuria
 Urgency and frequency of micturition
 Suprapubic tenderness
 Pyuria
• Haemorrhagic cystitis
○ Large quantities of visible blood in the urine
• Pyelonephritis
○ Kidney infection from the lower UTI
○ Complications
 Sepsis
 Septic shock
 Death
• Acute bacterial pyelonephritis
○ Clinical syndrome of fever, flank pain and tenderness
○ Leukocyte casts in the urine and bacteriuria
• Chronic bacterial pyelonephritis
○ Long-standing infection associated with active bacterial growth in the kidney; or the
residium of lesions caused by such infection in the past
• Chronic interstitial nephritis
○ Renal disease with histologic findings resembling chronic bacterial pyelonephritis but
without evidence of infection

Others Page 390

 ○
without evidence of infection

Common Risk Factors for UTI

Women
• Urinary tract obstruction (including calculi)
• Pregnancy
• Urologic instrumentation, catheterisation or surgery
• DM
• Sexual intercourse
• Women who use the diaphragm and spermicides
• Postmenopausal
○ Bladder or uterine prolapse
○ Loss of oestrogen that causes a change in the vaginal flora
○ Loss of lactobacilli in the vaginal flora

Men
• Urinary tract obstruction (including calculi)
• Catheterisation
• Prostatic enlargement (in their 50's - BPH)
• Uurologic instrumentation or surgery e.g. prostactectomy
• DM
• Lack of circumcision (children and young adults)

Mechanical and Functional Factors Predisposing to UTIs

• Disruption of urine flow or complete emptying of bladder
○ Pregnancy
○ Renal stones
○ Tumours
○ Prostate hypertrophy
○ Strictures - narrowing of ureter
• Loss of neurologic control of bladder and sphincters
• Vesicouretal reflux (reflux from urine from bladder up the ureter)
○ Anatomic abnormalities in children
• Catheterisation can facilitate bacterial access to bladder
○ During insertion

Commensal Microflora of the Urethra

• Coagulase-negative staphylococci (except Staphylococcus saprophyticus)
• Viridans and non-haemolytic streptococci
• Lactobacilli
• Diptheroids (Corynebacterium except C. urealyticum)
• Saprophytic Neisseria
• Anaerobic bacteria

Aetiology of UTIs
• Bacterial agents
○ Escherichia coli
○ Klebsiella pneumoniae
○ Proteus mirabilis
○ Pseudomonas aeruginosa
○ Staphylococcus aureus, epidermidis and saprophyticus
○ Enterococci (Streptococcus faecalis)
○ Mycobacterium tuberculosis
○ Chlamydia trachomatis
○ Mycoplasma spp

Others Page 391

 ○ Mycoplasma spp
• Viral
○ Rubella
○ Mumps
○ HIV
• Fungal
○ Candida
○ Histoplasma capsulatum
• Parasites
○ Trichomonas vaginalis
○ Schistosoma haematobium

Escherichia coli
• Most common urinary pathogen - 60-90% of infections
• Uropathic strains have pili associated with pyelonephritis (PAP pilus)
• Strains expressing O-antigens
○ O1, O2, O4, O6, O7, O8, O75, O150 - high proportion of infections
• Produce haemolysin
• Motile
○ Ascend urethra into bladder
○ Ascend ureter into kidneys

Pseudomonas, Proteus, Klebsiella and Staphylococcus aureus

• Mainly nosocomial
• Resistance to antibiotics favour their selection in hospital patients
• Proteus infections are associated with renal stones
○ Proteus produce a potent urease which act on ammonia, rendering the urine alkaline

Staphylococcus saprophyticus
• Found in sexually active young women

• Infection of the anterior urinary tract (urethritis) is mainly caused by Neisseria

gonorrhoeae, Staphylococci, Streptococci and Chlamydiae
• Mycobacterium tuberculosis is carried in blood to the kidneys from another site of infection
(e.g. respiratory TB)

Bacterial Virulence Factors

• Endotoxin
○ Decreases uretal peristalsis
• Haemolysin
○ Damages renal tubular epithelium
○ Promotes invasive infection
• Capsular
○ K1, K5 and K12 antigens of E. coli associated with clinical severity (anti-phagocytic)
• P-fimbriae
○ Enhance to uroepithelial cells
• Aerobactin (a siderophore)
○ Present in uropathogenic strains of E. coli
○ Promoting intracellular iron accumulation for bacterial replication
• Flagellar
○ Motile bacteria ascent the ureter against urine flow
• Bacterial urease: Proteus
○ Splits urinary urea with generation of ammonium ion that alkalinises urine
○ Loss of acid pH as natural defense barrier
○ Stone formation with uretal obstruction

Others Page 392

 ○ Stone formation with uretal obstruction
○ Survival of bacteria deep within stones, resisting eradication by antibiotics

Host Protective Factors in UTI

• Anti-bacterial properties of urine
○ High urea concentration
○ High organic acid concentration
• Anti-adherence mechanisms
○ Bacterial interference (commensals in urethra, vagina and periurethral region)
○ Urinary oligosaccharides (have the potential to detach epithelial-bound E. coli)
○ Uromucoid proteins: Tamm-Horsfall…coating of E. coli by this protein might prevent
attachment
• Miscellaneous
○ Mucopolysaccharide lining of the bladder
○ Urinary immunoglobulins
○ Spontaneous exfoliation of uroepithelial cells with bacterial detachment
○ Mechanical flushing of micturition

Long Term Bladder Catheterisation

• Species that notoriously tend to persist include:
○ Enterococci
○ Pseudomonas
○ Proteus
• Entry points for bacteria
○ Urethral meatus and around catheter and collection tube
○ Junction between catheter and collection tube
○ Connection to drainage bag and reflux from bag to tubing
○ Mouth of the spigot

Pathogenesis of UTI
• Periutheral area & urethra are colonised by bacteria
• Bacteria enter bladder in susceptible host
• Adherence properties enable pathogens to colonize bladder
• Pathogens attach to uroepithelial mucosa → secretion of cytokines → recruitment of
PMNs → inflammation.
• Pathogens may ascend through ureter to kidney → pyelonephritis
• Hematogenous seeding of renal cortex less frequent than ascending infection
• A common site of abscess formation in Staphylococcus aureus bacteremia
• Also occurs with Salmonella (typhoid) and Mycobacterium tuberculosis

Relapsing infection:
• Recurrence of bacteriuria with the same organism within 3 weeks of treatment.
• Indicates failure to eradicate the infection.

Reinfection :
• Eradication of bacteriuria by appropriate treatment, followed by infection with a different
organism after 7 to 10 days.
• It is due to the re-invasion of the urinary tract and not due to the failure of eradication of
the primary infection.

• Asymptomatic bacteriuria:
• Significant bacteriuria without clinical symptoms or other abnormal findings.
• Screening for asymptomatic bacteriuria
• Pregnancy
• High risk of acute pyelonephritis with its accompanying risk of fetal complications)

Others Page 393

 • High risk of acute pyelonephritis with its accompanying risk of fetal complications)
• Prior to urologic surgery (because of the risk of postoperative sepsis).

Laboratory diagnosis of UTIs

Sample Collection
• Early morning sample
• Sterile, dry, wide-necked, leak-proof container
• ”mid stream” urine sample
• Deliver ASAP/refrigerate at 4–6 °C/ boric acid preservative
• Mycobacterium tuberculosis :
○ Three early morning urine samples on 3 consecutive days

Macroscopic Examination
• Describe the appearance of the specimen
○ Colour of specimen
○ Whether it is clear or cloudy (turbid)

Direct Microscopy
• Wet film to detect :
○ Pus cells
○ Red cells
○ Yeasts
○ T.vaginalis
○ S. haematobium

1. Opacity
○ Normally, urine is CLEAR but may become turbid with infection.
2. pH:
○ Normal: 4.5-7.8
○ Increased with urea-splitting bacteria in the urine
3. WBCs:
○ Normal:<4)
○ Increased pyuria indicates infection
4. Bacteria:
○ Normal should be negative (bladder and urinary tract are normally sterile)
○ Urethra and the perineum (wide variety of Gram positive and Gram negative
organisms)

Urine Biochemistry
• Biochemical tests which are helpful in investigating UTI include:
○ Protein
 Indicative of bacterial UTI
○ Nitrite
 E. coli, Proteus species, and Klebsiella species are able to reduce nitrates
○ Leukocyte esterase
 Indirect test for pus cells

Urine Culture
• Not necessary when microscopically and biochemically normal, except when screening for
asymptomatic bacteriuria in pregnancy.
• Viable bacterial count
○ A measured volume (calibrated loop 0.002 ml) of urine is spread on surface of a solid
medium
○ Incubation of the solid medium at 37°C for 18-24 hours
○ Enumerate the number of colonies
• CLED (Cystine lactose electrolyte-deficient ) Agar

Others Page 394

 • CLED (Cystine lactose electrolyte-deficient ) Agar
○ Incubate aerobically at 35-37,overnight
○ The indicator in CLED agar is bromothymol blue and therefore lactose fermenting
colonies appear yellow)
○ CBA used if N. gonorrhoeae is suspected (no growth CLED)
○ Appearance of some urinary pathogens on CLED agar
 E. coli:
□ Yellow (lactose-fermenting) opaque colonies often with slightly deeper
coloured centre.
 Klebsiella species:
□ Large mucoid yellow or yellow-white colonies.
 Proteus species:
□ Transluscent blue-grey colonies.
 Pseudomonas aeruginosa:
□ Green colonies with rough periphery (characteristic colour)
 Enterococcus faecalis:
□ Small yellow colonies.
 Staphylococcus aureus:
□ Deep yellow colonies of uniform colour
 Staphylococcus saprophyticus and other coagulase negative staphylococci
□ Yellow to white colonies.

Identification Tests
• Catalase test for gram positive organisms
○ Positive for staphylococcus
○ Negative for streptococcus
• Triple iron sugar for gram negative rods
○ Follow up with IMViC

I M VP C
E.coli + + - -
Klebsiella - - + +
Citrobacter - - +
Enterobacter - + +

Anti-Microbial Susceptibility Test

• Perform susceptibility testing on urines with significant bacteriuria, particularly from
patients with a history of recurring UTI.
• Cultures from patients with a primary uncomplicated UTI may not require a susceptibility
test.

Sterile Pyuria
• Presence of pus in urine in absence of bacterial growth
• Causes
○ Infection with
 Chlamydia trachomatis
 Mycobacteria tuberculosis
 Ureaplasma
 Anaerobic bacteria
 Viruses
○ Previous antibiotic therapy
 Suppress growth of bacteria

Treatment in UTI

Others Page 395

Treatment in UTI
• Based on isolate and antimicrobial susceptibility pattern obtained.
• Medications
○ Short-course therapy: 3-5 day course of antibiotics for uncomplicated lower urinary
tract infection
○ 7 – 10 days course of treatment for
 Pyelonephritis
 Urinary tract abnormalities or stones
 History of previous infection with antibiotic-resistant infections
 Clients with severe illness may need hospitalization and intravenous antibiotics
• Possible outcomes of treatment for UTI, determined by follow-up urinalysis and culture
○ Cure
 No pathogens in urine
○ Unresolved bacteriuria
 Pathogens remain
○ Relapse
 Persistent source of infection causes repeated infection after initial cure
○ Reinfection
 Development of new infection with different pathogen

Others Page 396

Bacterial Genital Tract Infections
Thursday, 22 June 2017 11:15 AM

Commonly Encountered Bacterial STIs and Causative Agents

• Discharge from the genital tract as the initial main manifestation
Disease Causative agents
Gonorrhoea Neisseria gonorrhoea
○ Non-specific or non-gonococcal urethritis and Chlamydia trachomatis (serotypes
cervicitis D to K)
Ureaplsma urealyticum
Mycoplasma species
• May or may not be sexually transmitted
Bacterial vaginosis G.vaginosis and others
○ Fungal infection Candida albicans
(vaginal candidiasis)
• Ulceration of external genitalia or genital ulcer disease as the main initial manifestation
Disease Causative agent
Chancroid Haemophilus ducreyi
○ Syphilis Treponema pallidum pallidum
Lymphogranuloma venerum (LGV) Chlamydia trachomatis
Granuloma inguinale (donovanosis) Klebsiella granulomatis

Neisseria gonorrhoeae Infections

Gonorrhoea
• Initial manifestations
○ Adults - discharge mainly consisting of pus
 Males - features of urethritis
 Females - features of endocervicitis
• Other manifestations
○ Inflammatory process in other sites other than the GUT according to sexual practices
○ Ophthalmia neonatorum
○ Conjunctivitis resulting from contamination of the eyes
○ Vulvovaginitis in young females

Complications
• Local spread
○ Males
 To periurethral tissues including the prostate resulting in prostatitis and
epididymis resulting in epididymitis
○ Females to:
 Vaginal canal causing vaginitis
 Batholin's glands resulting in Batholinitis
 Endometrium resulting in endometritis
 Fallopian tubes resulting in salpingitis
• Dissemination
○ Disseminated disease characterised by:
 Septicaemia

Others Page 397

  Septicaemia
 Haematogenous spread to other tissues including arthritis and septic skin
lesions
• Long term complications
○ Associated with in treated infections or inadequately treated acute infection
 Urethral structures in males
 Complications in females which interfere with reproduction
 Blindness from ophthalmia neonatorum

Laboratory Diagnosis
• Specimen
○ Pus swabs or smears or discharge
 Delivery to the lab in a transport medium when processing is likely to delay
○ Blood for culture for disseminated infection
○ Urine
• Procedures - Urethral or vaginal discharge
○ Gram's stain and microscopy
 May provide presumptive diagnosis in males with uncomplicated urethritis
○ Cultures for isolation and identification
 Media
□ Selective media or heated blood agar
 Incubation
□ Under suitable conditions
 Colonial features
□ Greyish transluscent
□ Oxidase positive
□ Gram negative diplococci
□ Ferment glucose and not maltose, lactose and sucrose
 Differentiates from other Neisseriae
○ Rapid diagnostic tests
 Mainly DNA probe tests include:
□ PCR
□ Ligase chain reaction
□ _
• Antibiotic susceptibility reaction
○ Most strains
 Are susceptible to many antimicrobials including:
□ Ceftriaxone
□ Cefixime
□ Ciprofloxacin
□ Norfloxacin
 Show variable susceptibility to penicillin
□ May or may not be susceptible
□ Various penicillins including Amoxicillin and Procaine Penicillin are useful
for treatment of infections of susceptible strains
 Administered together with Probenecid

Challenges in Management
• Compliance
○ Patients tend to default
○ Whenever possible, anti-microbials are given in a single dose for acute
uncomplicated urethritis
• Co-existing Infections
○ Other STI due to other bacteria or other microorganisms
 Necessary lab tests for other STIs and proper management of those infected
• Emergence to resistance to commonly used anti-microbial agents including

Others Page 398

 • Emergence to resistance to commonly used anti-microbial agents including
○ Penicillin resistance
 Chromosomally mediated or
 Plasmid mediated resistance mostly due to beta-lactamase or penicillinase
production, resulting in penicillinase producing strains of Neisseria
gonorrhoeae or PPNG strains
○ Tetracycline resistance
 Either chromosomally or plasmid related
○ Resistance to other agents

Non-gonococcal or Non-Specific
Manifestations
• Similar to N. gonorrhoeae infection

Lab Diagnosis
• Specimens
○ Urethral or cervical or eye discharge
• Procedures
○ Gram's Stain and Microscopy
 Shows pus cells and no stainable bacteria
○ Culture negative for N. gonorrhoeae
○ _

Investigations of Chlamydia and Mycoplasma Infection

• Special staining methods for Chlamydia
• Detection of antigens on smears of discharge by different techniques including
immunofluorescence staining
• DNA probe or amplification by PCR of LCR
• Culture of Chlamydia on living cells
• Culture of Mycoplasma on a specific medium

Anti-microbial Susceptibility of Chlamydia and Mycoplasma

• Effective agents include
○ Tetracycline
○ Erythromycin
○ Doxycycline
○ Azithromycin

Chancroid or Soft Sore

• Causative agent is Haemophilus ducreyi

Lab Investigation
• Specimen
○ Swab from the ulcer
• Procedures
○ Gram stain and microscopy
○ Culture
 Medium containing growth factor X
 Incubation in moisture and additional 5-10% carbon dioxide
○ Colonies
 Typical morphology
 Staining and typical microscopy_

Lymphogranuloma venereum of LGV

• Initially a painless ulcer on the external genitalia
• Infection spreads to the lymph node through lymphatic channeks

Others Page 399

 • Infection spreads to the lymph node through lymphatic channeks
• Associated with painful lymphadenitis and lymphangitis and enlarged inguinal and femoral
lymphnodes or buboes
• In females, lesions involve the:
○ _
• Associated with long-term complications due to chronic inflammatory process and fibrous
tissues formation leading to
○ Lymphatic obstruction
○ Development of strictures

Lab Investigation
• Procedures include:
○ Detection of specific antibody to the causative agent in serum
○ Cell culture for isolation and identification
○ Fluorescent antibody test for antigen detection
○ Genetic based methods of detection

Antimicrobial Treatment
• Effective antimicrobial agents
○ Tetracycline
○ Doxycycline
○ Erythromycin

Syphilis
C.f notes on Treponema

Lab Investigation
• Demonstration of T. pallidum in specimens
• Serological tests
○ 2 groups
 Non-treponemal tests
 Treponemal or confirmatory tests
• Other tests
○ ELISA
○ PCR

Granuloma inguinale or donovanosis

• Causative agent is Klebsiella granulomatis
○ Encapsulated gram-negative rods
• Small painless nodules appear approximately 10-14 days after exposure
○ Burst and form painless ulcerative genital lesions with typical appearance
• Inguinal lymph node enlargement is generally absent
• Wright's or Giemsa stains of infected_
• Susceptible to several agents including
○ Doxycycline
○ Erythromycin
○ Tetracycline
○ Prolonged treatment may be required

Bacterial Vaginosis
• Not always attibuted to a specific causative organism
○ Associated with Gardnerella vaginalis and other organisms including strict anaeroes
mostly Bacteroides species

Others Page 400

 ○
mostly Bacteroides species

Gardnerella vaginalis
• Gram stain variable
• Non-motile bacillus
• Non-spore forming
• Facultative anaerobe
• Asymptomatic carriage in 50% of females
• Most consistent organism isolated in the majority of vaginoses in adult females
• Main manifestation bacterial vaginosis is watery vaginal discharge
• Possible complications in pregnancy include
○ Chorioamnionitis
○ Premature rapture of membranes
○ Preterm delivery
• Lab characteristics of vaginal discharge
○ _

Antimicrobial Management
• Responds to several agents including Metronidazole and Clindamycin

Prevention and Control of Bacterial STI and STD

• Require active participation and cooperation by
○ Community
○ Individual adults
○ Patients
○ Health workers
• Methods include:
○ Education on safer sex practices and information on possible outcomes of unsafe
practices
○ Prompt diagnosis and effective treatment with anti-microbial agents
○ Contact tracing education and treatment of contacts
○ Proper clinical examination and lab tests for other STIs

Syndromic Approach in Management of STI and STDs

• Manages an STD as a syndrome
• Uses the signs and symptoms to determine the antimicrobial treatment
○ Consideration of more common causative agents may be required in some cases
• Antimicrobials are chosen to cover the major pathogens associated with the
manifestations
○ Bacteria and other organisms

Aspects of Prevention in Syndromic Management of STD

• Counselling
○ Talking with the client and discussion of the next points
• Compliance
○ Emphasis on adherence to
 Taking the medication
 Following instructions concerning medication and safer practices during and
after treatment
 Finishing the course of treatment
 Avoidance of self medication
• Condom use and other barrier methods
○ Proper and consistent use of barrier methods
• Contact tracing and treatment of all contacts

Some Advantages of Syndromic Management of STD

Others Page 401

Some Advantages of Syndromic Management of STD
• Treatment is given at the first visit therefore
○ Client is not lost to follow up before treatment
○ Chances of further transmission are reduces
○ Complications of untreated infection are prevented
• Cost saving as elaborate lab investigations are not performed
• Designed flow charts are used therefore the
○ Diagnosis, treatment, referral and reporting, are standardised

Some Disadvantages of Syndromic Management of STD

• Over diagnosis and subsequently increased cost of treatment when dealing with a single
infection
○ Costs of the various anti-microbial agents
• Risk of adverse reactions to anti-microbial agents
• Alteration in the normal flora and selection of resistant pathogens in a community
• Asymptomatic or sub-clinical infections by other organisms are not investigated and
therefore not managed if present

Others Page 402

Septicaemia and Bacteraemia
Tuesday, 27 June 2017 3:21 PM

• Commonly used interchangeably to refer to the presence of bacteria in the circulating

blood
• Other workers no longer_
• Entry of bacteria into the blood can occur
○ Through breakages of blood vessels including capillaries or small veins
○ Via phagocytic cells into the capillaries or the lymphatic system

Septicaemia
• Strictly refers to a clinical condition associated with severe manifestations which are due to
○ Effects of a large number of bacteria with or without bacterial toxins in the blood
○ Manifestations of predisposing illness

Bacteraemia
• Strictly refers to the presence of viable bacteria in the circulating blood
○ May or may not cause clinical manifestations
○ Confirmed by culture of bacteria from blood

Endotoxaemia
• Condition which results from effects of bacterial endotoxin in blood

Sources of Organisms in Bacteraemia or Septicaemia

• Include
○ Focus of infection in other tissues can be:
 Peritonitis
 Pneumonia
 Meningitis
 Abscesses of internal organs
○ Generalised infections including enteric fever and brucellosis
○ Others
 Unidentified sites
 Normal flora
 Contaminated item in invasive procedures
 Contaminated in-dwelling devices including urinary catheters or other foreign
material in the tissues

Types of Bacteraemia
• Gram negative bacteraemia
○ Majority are caused by bacteria in famile Enterobacteriaeceae and Pseudomonas
○ Others include
 Neisseria meningitidis
 Brucella spp.
• Gram positive bacteraemia
○ Several pyogenic cocci
• Mixed bacteraemia
○ More than one species including gram negative bacilli (GNB) or mixed cocci and
GNB
• Bacteraemia due to anaerobic organisms
○ Bacteroides fragilis is most common
○ Other anaerobic GNB
○ May be associated with abdominal or gynaecological infections or trauma

Others Page 403

 ○ May be associated with abdominal or gynaecological infections or trauma
• Nosocomial Bacteraemia
○ Caused by a variety of agents commonly associated with hospital acquired infections
including
 Klebsiella pneumoniae
 Pseudomonas aeruginosa
 Serratia marcescens
○ Organisms show multiple anti-microbial agents

Complications of Bacteraemia
• Spread of infection to other sites
• Shock
○ Septic shock by gram positive bacteria
○ Gram negative shock due to endotoxin and complications of endotoxic shock

Laboratory Investigation of Bacteraemia

• Specimen
○ Blood for culture
○ Pus or other fluids or infected tissue
• Procedures
○ Gram's stain and microscopy on suitable specimens other than blood
○ Cultures
 On suitable media for possible organisms
 Incubation
□ In air
□ Anaerobic methods
□ In additional 5-10% carbon dioxide
 Identification by standard bacteriological methods
○ Other methods including DNA based testing
○ Anti-microbial susceptibility tests
 Choice of anti-biotics and methods of tests used are determined by the isolated
organism

Anti-Microbial Management o Bacteraemia

• Adequate treatment with appropriate bactericidal antimicrobials
• Treatment of predisposing actor
• Prevention and treatment of complications

Others Page 404

Pyrexia of Unknown Origin (PUO)
Tuesday, 27 June 2017 4:29 PM

• Also known as fever of unknown origin (FUO)

• Characterised by
○ Persistently elevated temperature of over 38°C
 For one week or longer
 Can be as long as 3 weeks
○ No obvious identifiable cause despite immediate
 Clinical examination by a clinician
 Lab investigation
□ Both in health_

Causes of PUO
• Infections
○ Responsible for the majority
○ Can be viral, bacterial, fungal or parasitic (or other)
• Non-infectious illnesses

Bacterial Infections and Diseases Which May Manifest as PUO

• Generalised infections, including
○ Infective endocarditis
○ TB
○ Enteric fever
○ Leptospirosis
○ Q-fever
○ Brucellosis
• Localised infections associated PUO
○ Abscesses
 Intra-pulmonary
 Sub-phrenic
 Pelvic
○ Mycoplasma pneumonia
○ Sinusitis
○ Others

Bacteriological Investigations of PUO

• Blood cultures repeatedly at least 3 times
• Examination of other specimens including:
○ Urine exam
○ Stool microscopy and culture particularly where there is associated diarrhoea
○ Sputum
• Serological tests including:
○ Widal test or other tests for enteric fever
○ For atypical pneumonia
○ For Leptospirosis
• DNA amplification tests

Others Page 405

Endocarditis
Tuesday, 27 June 2017 4:28 PM

• Refers to the infection of the endocardium and/or the heart valves

○ An intra-vascular infection which occurs within the heart
• Infective endocarditis
○ Refers to endocarditis caused by any disease causing organism
• Bacterial endocarditis
○ Restricted to endocarditis caused by bacteria

Conditions Which Predispose to Bacterial Endocarditis

• Artificial or prosthetic heart valve
• History of previous endocarditis
• Damaged or scarred heart valves with various conditions including RHD
• Congenital or acquired heart defects
• IV drug injections without improper skin disinfection
• Bacteraemia

Development of bacterial Endocarditis

• The predisposing heart condition causes damage to the endocardium
• The damage causes roughness of the endocardium
• Thrombi of fibrin and platelets form on the roughened surface
• Organisms from the circulating_

Sources of Infection and Causative Agents of Bacterial Endocarditis

• Include:
○ Normal flora
 In transient asymptomatic bacteraemia mostly from
□ Skin
 Staphylococcus aureus
 Staphylococcus epidermidis
 Also encountered in association with IV drug users under septic
conditions
□ Mouth
 Streptococcus viridans
□ GIT
□ GUT
○ Septicaemia
 Associated with several organisms including Staphylococcus aureus and
Streptococcus pneumoniae
○ Hospital environment
 Associated bacteria include Enterococcus species as causative agent of
hospital acquired endocarditis
○ Other organisms
 Some fastidious bacteria not easily isolated in culture. Grouped as HACEK
organisms

Lab Investigation
• Blood culture for isolation and identification of culturable causative bacteria
○ Performed repeatedly if necessary 2-6 samples taken over 48 hours or 3 samples
taken within 24 hours
○ Incubated aerobically in air in 5-10% CO2 and anaerobically

Others Page 406

 ○ Incubated aerobically in air in 5-10% CO2 and anaerobically
○ Most suitable specimen taken before administration of anti-microbial agents
 However, treatment should not be delayed
○ Isolated bacteria are identified by standard bacteriological methods
○ Antimicrobial susceptibility tests
 Disc diffusion technique is generally considered inadequate
 Determination of MIC of the effective agents is recommended
• Serology
○ In selected cases
• Other bacteriological tests
○ Including DNA based tests
• Other medical tests
○ Radiological diagnostic tests

Tests During Treatment

• Estimation of anti-microbial agent levels in patient's serum
○ May or may not be necessary
○ _
• Measurement of indicators of inflammation including C-reactive protein may be useful in
○ Assessment of response to treatment
○ Detection of inter current infection

Negative Blood Cultures

• May be due to
○ Uncommon organisms including Coxiella burnetii
 _
○ Recent anti-microbial agent treatment
 _
○ Infection due to fastidious organisms which do not grow easily on commonly used
culture media
 Might require linger incubation or a repeat blood culture using specifically
prepared media for the suspected organisms

Anti-Microbial Treatment
• High doses of bactericidal antimicrobial agents(s) administered intravenously initially
ensures
○ _
• Combination of two agents is given for better results
○ _
• Duration of treatment depends on:
○ _
• Generally requires 6 weeks of treatment

Prevention
• Anti-biotic prophylaxis
○ Recommended for individuals at a higher risk when undergoing surgical or
instrumentation procedures involving parts which harbour normal flora in large
numbers
○ Regular use of anti-microbial for prevention is associated with the following
disadvantages
 Adverse effects of anti-biotics
 Resistance
• Improvement of oral hygiene in predisposed people with low levels of oral hygiene

Others Page 407

Blood Cultures
Tuesday, 27 June 2017 4:28 PM

• Laboratory tests performed in bacteriology to isolate bacteria from blood in circulating

system
• Mostly performed to:
○ Detect bacteraemia and determine the causative organism(s)
○ Establish the diagnosis and causative agents in specific infections including:
 Meningitis
 Osteomyelitis
 Endocarditis
 Severe pneumonia
 Sepsis including puerperal and neonatal sepsis
○ Investigation of PUO and fever in hospitalised patients with non-infectious illnesses

Media For Inoculation of Blood Specimen

• Nutritionally enriched liquid media or broths
○ Prepared on the lab or commercially obtained
○ Suitable for the growth of
 Strict _
• Commonly used media include:
○ Brain heart infusion (BHI)
○ Tryptone soya broth for aerobes and facultative anaerobes
○ BHI or tryptone soya broth with thioglycollate for strict anaerobes
• Media may be available as
○ Bottled liquid medium or broth
 In 50ml volumes for adulta and 10ml for children
□ Sealed and fitted with provision for inoculation
 Co_
○ Biphasic medium
 Semi solid part and a liquid part o the broth in one bottle
□ Includes Casteneda system for culture of Brucella and other organisms
○ Specific media for uncommon organisms
 Include media for Leptospira spp.

Techniques
• Contamination is minimised as much as possible
○ Aseptic techniques applied during venipuncture and specimen transport and
sampling
 Washing of hands and allowing them to dry
 Identification of the most suitable venipuncture site
 Cleansing and disinfection of the skin over the venipuncture selected site
□ 70% ethanol then 1% iodine or iodophor or 1-2% chlorhexidine may be
used
 Allowing the site to dry for effective disinfection
 Palaption of the vein with hands in steril gloves
 Use of sterile needle and syringe to take blood
 Change of needle before inoculation of the required volume into the blood
culture bottle

Quantity of Blood Specimen

• Samples are taken are the same time for aerobic and anaerobic cultures
○ Adults

Others Page 408

 ○ Adults
 At least 10ml of blood per culture is most suitable
○ Children
 5ml for children
○ _
• Generally for adults and children
○ Optimal blood to broth volume ratio is 1:5 to 1:10
 _
○ Commonly used anti-coagulant in the medium is liquoid (sodium polyanethol_
 _
○ _

Number and Timing of Blood Samples

• Where several samples are collected each sample constitutes one blood culture set
• Two or three blood culture setts are considered sufficient to confirm to rule out
bacteraemia
• One set is rarely sufficient
• The optimal time for specimen collection is before onset of fever as bacteraemia precedes
the febrile episode
• Samples are delivered to the lab as soon as possible
• Incubated appropriately at 35-37°C

Detection of Bacterial Growth in Blood Samples

• Incubated samples are examined daily for at least one week for evidence of growth and
sampling
• Longer incubation may be necessary where fastidious organisms which grow slowly are
suspected
• Care is taken to avoid contamination
• Sub-cultures are made on commonly used semi-solid media and incubated _

Other Methods of Detection of Bacterial Growth in Blood Culture Specimens

• Include radiometric method referred to as BACTEC
○ Consists of automated systems which detect CO2 produced by bacterial metabolism
in broth medium
○ Based on various radiometric or fluorescent techniques
 Detects bacteraemia faster
 Requires smaller blood volumes
 Reduced lab work load

Blood Culture Results

• May be influenced by
○ Clinical judgement
 Possibility of bacteraemia and the causative organism
○ Specimen collected
 Quality - possibility of contamination
 Quantity
□ In most infections, the organisms are scanty so large volumes if blood will
increase the chances of isolation
• Interpretation
○ A single culture of a properly collected sample yielding bacterial growth is significant
○ Negative cultures do not exclude infection

Others Page 409

Wound Infections
Thursday, 29 June 2017 11:06 AM

Introduction
• Staphylococcus aureus
○ Commonest pathogen isolated from subcutaneous abscesses and _
• Pseudomonas aeruginosa
○ Associated with infected burn and nosocomial infections
• Escherichia coli, Proteus spp., Pseudomonas aeruginosa and Bacteroides spp
○ Most frequently isolated from abdominal abscesses and wounds
• Clostridium perfringens
○ Found mainly in deep wounds where anaerobic conditions exist
○ The toxins produced cause gas gangrene
• Chronic leg ulceration common in sickle cell disease. Commonest pathogens
○ Staphylococcus aureus
○ Pseudomonas aeruginosa
○ Streptococcus pyogenes
○ _
• _
• Bacillus anthracis causes cutaneous anthrax. Fluid from the pustule is highly infectious
• Yersinia pestis cause plague; highly infections
• M.ulcerans causes M. ulcerans disease (buruli ulcer)

Wound Infection
• Penetration of the skin by micro-organisms is difficult
○ Part of the innate defense
• Wounds provide the most common access through the skin
• Disease production in infected wounds depends on
○ Virulence of the infecting organism
○ Infectious dose
○ Immune status of the host
○ Nature of the wound
 Foreign material present?

Staphylococcal Wound Infections

• Leading cause of wound infection
• Symptoms
○ Bacteria are pyogenic
○ Infection causes
 Inflammation
 Fever
○ Some strains produce toxic shock syndrome
• Causative agent: Staphylococcus aureus
○ Virulence factors
 Coagulase
□ Causes fibrin formation to evade phagocytosis
 Clumping factor
□ Aids in bacterial wound colonisation
 Protein A
□ Antigenic
 Toxin production
□ Epidermolytic toxin, TSST

Others Page 410

 □ Epidermolytic toxin, TSST
• Epidemiology
○ 30-100% due to the patient's own normal flora
○ Factors associated with infection include
 Advanced age
 Immunosuppression or poor general health
 Prolonged post operative hospital stay

Group A Streptococcal Infections

• Primary pathogen is Streptococcus pyogenes
○ Also known as "flesh eaters"
○ Can cause rapidly deteriorating disease and death
• Common cause of wound infection
○ Two extracellular products are responsible for virulence
 Pyogenic exotoxin A
□ Superantigen: toxic shock
 Exotoxin B
□ Necrotising fasciitis

Pseudomonas aeruginosa Infection

• Major cause of nosocomial infection
○ Lung infections
○ Burn wound infections
• Community acquired infections include
○ Rash and external ear infections
○ Infections of foot bones
○ Eye infections
○ Heart valve infections
○ _
• Pathogenesis
○ Some strains produce enzymes and toxins to enhabce virulence
 Exoenzyme S
 Toxin A
 PLC
○ Epidemiology
 P.aerugonosa is widely spread

Tetanus; Clostridium tetani

• Symptoms
○ Divided into early and late symptoms
 Early
□ Restlessness
□ Irritability
□ Difficulty in swallowing
□ Contraction of jaw muscles
□ Convulsions
 Mainly in children
 Late
□ Increased muscle involvenent
□ Pain
□ Difficulty in breathing
□ Death
• Causative agents
○ Clostridium tetani
 Anaerobic

Others Page 411

  Anaerobic
 Gram-negative
 Bacillus
 Spore former
○ 25% mortality rate; rare in the developed world
 Tetanospasmin toxin
□ Block inhibition of motor neurons , causing paralysis
□ Prevention
 Vaccination
□ Treatment
 Anti-toxin
○ Bacterial spores prevalentin dirt and duct and GIT of humans and many animals

Clostridium Myonecrosis (Gas Gangrene)

• Causative agent
○ Several species of Clostridium
 Most common, Clostridium perfringens
 Encapsulated, Gram-negative bacilllua
• Endospores of causative bacillus are innumerable
○ Spores found in nearly all soil or dusty surface
• Primarily disease of wartime
○ Due to neglected wound containing debris

Actinomycosis
• Causative agent
○ Actinomyces israelii
 Filamentous anaerobic, slow-growing
• Pathogenesis
○ Actinomyces israelii cannot penetrate healthy mucosa
○ Infection is characterised by cycles
 Abscess formation→ scarring → formation of sinus tracts
○ Disease progresses ti skin and can penetrate bone and CNS

Pasteuella multocida
• Causative agent
○ Pasteurella multocida
 Gram-negative
 Coccobacillus
 Most are encapsulated
• Bite infections from numerous animals
○ Fowl cholera
○ Animal reservoir
• Symptoms
○ Spreading redness
○ Tenderness
○ Swelling of adjacent tissues
○ Pus discharge

Cat Scratch Disease

• Causative agent
○ Bartonella henselae
 Gram-negative bacillus

Others Page 412

  Gram-negative bacillus
• Symptoms
○ Disease begins within a week
○ Painful enlargement of lymph nodes
○ Fever
• Epidemiology
○ Zoonotic disease
 Cats infected by flea bite

Sporotrichosis ("Rose Gardner's Disease")

• Causative agent
○ Sporothrix schenckii
 Dimorphic fungus
 Lives in soil and on vegetation
• Associated with puncture wound from vegetation
• Sporadic
○ Rare in healthy people
○ Untreated cases may become chronic

Lab Diagnosis
• Specimen
○ Swabs (avoid contamination of specimen with commensal organisms from the skin)
○ Aspirate
• Stain
○ Gram stain of the specimen
○ ZN staining
○ Giemsa/Wayson's staining
• Microscopy
○ Dark field microscopy
• Culture
○ BA, CBA, RCM
○ Aerobically/5-10% CO2/anaerobically
○ 35-27°C
○ Describe the colonial morphology
• Identification tests
• AST

Treatment and Prevention

• Depends on
○ Organism isolated
○ AST results
○ Mode of transmission

Others Page 413

Respiratory Tract Infection
Thursday, 29 June 2017 11:41 AM

Introduction
• Most common infections in clinical practice
• Divided into
○ URTI
○ LRTI
• URT
○ Frequently the site of infection and localised infection

Upper Respiratory Tract Infections (URTI)

• The upper respiratory tract system include
○ Nose
○ Nasal cavity
○ Pharynx
○ Larynx with subglottic area of the trachea
• URTIs
○ Most common infections in the population
• Leading cause for missing work or school
• Range from mild, self-limiting disease e.g. common cold to serious life threatening_
• Most infections are viral in origin
• Bacteria can be primary cause or superinfection
• All the parts or URT and associated structures e.g. paranasal sinuses ad middle ear
• Transmission
○ Person to person via touching secretions by hand; directly by inhalation of droplets
• Common URTI
○ Rhinitis
○ Sinusitis
○ Rhinositis
○ Nasopharyngitis
○ Pharyngitis
○ Epiglottitis
○ Laryngitis
○ Laryngotracheitis
○ Tracheitis

Rhinitis and Rhinosinusitis

• Inflammation and swelling of the mucous membranes of the nose and paranasal sinuses
• Characterised by
○ Runny nose
○ Rhinorrhoea
○ Sneezing
○ Congestion
○ Obstruction of nasal breathing +/- pruritus
• Acute or chronic
• Mostly viral
• Acute bacterial sinusitis usually a complication of viral URTI
• Common causes of acute bacterial sinusitis
○ Pneumococcus
 Haemophilus influenzae

Others Page 414

  Haemophilus influenzae
○ and β haemolytic streptococci
 Staph aureus
○ Anaerobes
• Complications of untreated bacteria rhinosinusitis
○ Ossteitis of the sinus bones
○ Orbital cellulitis
○ Spread to the CNS
 Meningitis
 Brain abscess
 Infection of the intracranial cavernous sinus

Pharyngitis
• Commonest cause
○ Viral
• Acute bacterial pharyngitis and tonsillopharyngitis
○ GAS - 15-30% of all cases of pharyngitis in children
○ 10% in adults
○ Gonococcal pharyngitis

Epiglottisis
• Infection can result from bacteraemia or direct invasion of the epithelium b pathogens
• Can also result from secondary bacterial infection of viral URTI
○ H.influencae
○ Pneumococcus
○ Staphylococci aureus
○ _

Laryngitis
• Bacterial causes
○ Hib
○ Β-haemolytic streptococci
○ M.catarrhalis
○ Pneumococcu
○ Klebsiela pneumoniae
○ S.aureus
○ Corynebacterium diphtheriae
○ C.ulcerans
○ TB
○ Treponema pallidum
• Fungal causes
○ Candida albicans
○ Blastomyces dermatitidis
○ Histoplasma capsulatum
○ Cryptococcus neoformans

Tracheitis
• Inflammation of the larynx, trachea
• Bronchi
• Major site:
○ Subglottic area
○ Airway obstruction may develop
 S.aureus
 GAS
 M.catarrhalis
 Hib

Others Page 415

  Hib
 Klebsiella spp.
 Pseudomonas spp.
 Anaerobes
 M.pneumoniae

Complications of URTIs
• Otitis media - 5% of rhinitis in children and 2% in adults
○ Pneumococcus, M. catarrhalis, Haemophilus influenzae
○ Complications
 Mastoiditis
 Meningitis
 Hearing loss due to CSOM
• GAS pharyngitis
○ Rheumatic fever
○ Acute glomerulonephritis
○ Scarlet fever
○ STSS
• Airway obstruction
○ Diphtheria

Lab Diagnosis
• Specimen
○ Throat swab
○ Nasopharyngeal aspirate
• Usually Gram stain of specimen not routinely done because of the wide variety of
commensals
• Culture
○ BA, Tinsdale medium/Potassium tellurite agar
○ _
• Biochemical Tests
• Serotyping
• AST

LRTI
• Include:
○ Pneumonia
○ TB
○ Empyema
○ Bronchitis
○ Lung abscesses
• Childhood pneumonia
○ Leading single cause of mortality on children aged less than 5 years
• Acquisition of microbes
○ Aspiration - elderly, ↓ consciousness
○ Inhalation
○ Rarely
 Haematogenous
 Extension from infected pleura or mediastinal space
• Acute
○ Streptococcus pneumoniae
○ Stapylococcus aureus
○ Haemophilus influenzae
○ Enterobacteriaceae

Others Page 416

 ○ Enterobacteriaceae
• Chronic
○ Mycoplasma tuberculosis
○ Nocardia
○ Steptococcus pneumoniae associated wth viral illness
• Community-acquiered pneumonia
○ 80% due to viruses
○ Streptococcus pneumoniae
○ M.pneumoniae
• Atypical pneumoniae
○ Mycoplasma pneumoniae
○ Legionella pneumophila
○ C.pneumoiae
• Fungi and actinomycetes
○ Blastomyces dermatitidis
○ Histoplasma capsulatum
○ Aspergillus spp.
○ Candida albicans
○ Cryptococcu neoformans
○ Nocardia spp.
• Pneumococcus, Hib
○ Commonest causes in tropical countries
• S.aureus, S. pyogenes and H. influenzae
○ Secondary invaders in patients with influenza virus pneumonia
• P.aeruginosa
○ Chronic lung cavities or in patients on immunosuppressive therapy
• Klebsiella pneumoniae + E. coli and yeasts as a complication of antimicrobial treatment
• M.catarrhalis
○ Pre-existing respiratory diseases and immunosuppression

Tuberculosis
• MTBC
○ M.tuberculosis
• MOTTS/ATM/Environmental Mycobacteria
○ M.avium complex

Bronchitis
• Acute or chronic
• Usually an exacerbation of COPD
• Pneumococci, non-encapsulated Haemophilus
• Sometimes follows a viral infection
• Cystic fibrosis
○ S.aureus
○ H.influenzae
○ P.aeruginosa

Empyema
• Collection or gathering of pus within a naturally existing anatomical cavity
• Empyema thoracis
• Typically a complication of pneumonia
• Also arise from
○ Penetrating chest trauma
○ Oesophageal rupture
○ Complication from lung surgery
○ Inoculation of the pleural cavity after thoracentesis or chest tube placement

Others Page 417

 ○ Inoculation of the pleural cavity after thoracentesis or chest tube placement
• Extension of a sub-diaphragmatic or paravertebral abscess

Lung Abscesses
• Subacute infection in which an area of necrosis forms in the lung parenchyma
• More often involves the right lung other than the left
• Seen after aspiration of oropharyngeal secretions

Lab Diagnosis
• Specimen
○ Sputum
○ Aspirate
○ Broncho-alveolar lavage
• Macroscopic examination
○ Purulent
○ Muco-purulent
○ Mucoid
○ Mucosalivary
○ Blood
• Microscopic examination
○ Gram stain
○ ZN stain
○ Need to decontaminate sputum with Sodium hypochlorite
○ KOH preparation
 Aspergillus
○ Giemsa and Wayson stain
 Pneumonic plague
• Culture
○ BA, CBA, MAC
○ LJ
○ SDA
• Identification tests
• AST

Treatment
• Antibacterial antimicrobial aegents
• Antimycobacterial agents
• Antifungals

Others Page 418

Parasitology SAQs
Sunday, 22 January 2017 6:22 AM

1. Briefly discuss
○ Treatment and control of schistosomiasis
 Chemotherapy
• Antimony compounds
• Hycanthone
• Niridazole
• Metriphonate
• Oxamniquine
• Praziquantel (adults)
• Artemisinin (larval schistosomes)
Control
• Breed in slow moving/stagnant fresh water.
• Molluscides - endod (from gooseberry)
• Biological methods
○ Cray fish to eat the snails
• Sewage treatment
○ Diagnosis and treatment of Wuchereria bancrofti
 Diagnosis
• History of living in the endemic areas of years
• Physical examination findings
• Blood examination (night sample) for microfilariae
○ Thick blood smear
○ Capillary tube exam
○ Blood filter (nucleopore)
 Stain with Giemsa stain for analysis
 Fields stain*
• DEC provocation test (DEC 5-mg, blood >30 min)
○ Diethylcarbamazine*
○ Irritates the microfilariae to retreat to blood during the day
• Ultrasound of the scrotum - filarial dance sign
• Serological tests
○ Filarial specific antibodies e.g. IgG, circulating filarial antigen (CFA)
○ Does not ascertain current infection, exposure or post-exposure
 Treatment
• DEC
○ Should not be used in places where there is onchocerciasis
occurring together with LF
• Ivermectin
• Albendazole
○ Combination Rx with DEC or Ivermectin
○ Pathology of infection of Ancyclostoma duodonale
○ Life cycle and pathology of infection with Trichuris trichuria
 Life Cycle
• The larvae develops to adults, mate, female produce larvae,
penetrate intestines into blood throughout the body, encapsulate in
muscle (skeletal, tongue, diaphragm etc) and >6 months may
calcify leading to death of the larvae
• Clinical Presentation
• Presentation is variable:
Oedema of the upper eyelid

Revision Page 419

  Oedema of the upper eyelid
 Subconjuctival haemorrhage
 Photophobia
 Diarrhoea
 Myalgia – muscle pain
 Generalised weakness
 Fever
 Skin rash
 Finger clubbing
• Complications
 Cardiac failure
 Respiratory failure
 Meningitis
 Meningoencephalitis
 Fits and comas in long infections
2. Write short notes on the following:
○ Complications with infections with Entamoeba histolytica
• Anaemia due to loss of blood in stool
• Liver abscess - anchovy paste like contents
• Intestinal obstruction
• Amoebiasis due to scar formation and tumour-like appearance
• Abdominal wall ulceration
• Rupture of liver abscess with development of peritonitis
○ Clinical manifestations and diagnosis of Plasmodium falciparum
• Simple Uncomplicated malaria
• Irregular fever with chills (tertian [48 hours] - Falciparum, vivax, ovale;
Quartam [72 hours] - malariae)
• Headache, NVD, loss of appetite, dizziness, muscle aches, joint pains,
general malaise
• High temperature, jaundice, pallor, abdominal tenderness, splenomegaly,
hepatomegaly
• Severe complicated falciparum malaria
○ Cerebral malaria, convulsions, hypoglycaemia, hyperpyrexia, severe
anaemia, renal failure, pulmonary oedema or acute respiratory distress
syndrome, algid malaria (shock secondary to bacteraemia or
septicaemia), DIC (disseminated intravascular coagulopathy),
haemoglobinuria (black water fever), jaundice, hyperlactataemia,
abortion, premature birth, low birth weight babies
○ Hyper-reactive malaria splenomegaly (HMS), previously tropical
splenomegaly syndrome (TSS)
Lab diagnosis
• Gold standard = thick and thin blood films
• Stain (Giemsa, fields, etc)
• Examine thick film first under microscope - put a drop of blood at centre of
slide, spread to size of a shilling coin, let it dry, stain, examine. Don’t fix
• If parasites are seen, thein film to confirm species
• Thin film_ blood at edge of slide, spread with second slide, allow film to
dry, fix using alcohol, stain, examine under microscope
• Giesma stain - best if few thick films, slow
• Field's stain - best if many thick films, very quick.
• Other diagnostic methods
○ Antigen detection- parasite-specific lactate DH in P. falciparum
infection
 Fast, high sensitivity and specificity, but unable to quantify the
parasite load, distinguish parasite species and Ag may be
detected 2 weeks post-Rx hence not always active infection

Revision Page 420

 detected 2 weeks post-Rx hence not always active infection
○ Quantitative Buffy Coat-centrifuge blood in capillary tubes pre-
coated with acridine orange (OA), stains parasite DNA, view under
UV light microscope.
 High sensitivity, can't distinguish young P. falciparum and P.
Vivax trophozoites
○ PCR - Experimental. Detects<10parasite/10microlitres blood.
Expensive
○ Pathology and clinical manifestation of infection with Trypanosoma brucei
rhodensiense
• Rhodensiense - fast (acute infection)
• Incubation period about 2 - 3 weeks
• Early stages
○ Chancre at the site of the glossina bite
○ Lymphadenopathy
 Generalised or local.
□ Back of the neck (posterior triangle of the neck) -
Winterbottom's sign - gambiense
○ Irregular fever
○ Headache
○ Malaise
○ Joint pain
○ Jaundice
○ Pallor
 Signs of anaemia
○ Splenomegaly
○ Hepatomegaly
○ Pancarditis
 Congestive cardiac failure
□ Swelling of the feet
□ Facial oedema
○ Insomnia
• Late stages
○ Mental confusion
○ restlessness at night
○ Excessive sleepiness during the day
○ Muscle twitching and tremors
○ Slurred speech
○ Muscle wasting
○ Convulsions
○ Coma
○ Death
○ Kerandal's sign
 Loss of elbow and knee jerk reflexes
○ Treatment and control of infection with Toxoplasma gondii
Treatment
• Disease is immunocompetent adults with lymphadenopathic
toxoplasmosis is usually self-limited - treatment rarely indicated
• If visceral disease is clinically evident or symptoms are severe or
persistent, treatment may be indicated for 2 - 4 weeks
• Persons with AIDS who develop active toxoplasmosis (usually
toxoplasmic encephalitis) need treatment that must be taken until a
significant immunologic improvement is achieved as a result of
antiretroviral therapy; then low dose maintenance treatment
• Treatment for ocular diseases should be based on a complete
ophthalmologic evaluation. The decision to treat ocular disease
dependent on numerous parameters including acuteness of the lesion,

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 dependent on numerous parameters including acuteness of the lesion,
degree of inflammation, visual acuity and lesion size, location and
persistence

Maternal and foetal infection

• Spiramycin is recommended (for the first and early second trimesters)
• Pyrimethmine/sulphadiazine and folinic acid (for late second and third
trimesters)
• PCR is often performed on the amniotic fluid at 18 gestation weeks to
determine if the infant is infected. If the infant is likely to be infected, then
treatment with drugs such as pyrimethamine, sulphadiazine, leucovorin
(typical)
• Congenitally infected new-borns are treated with the above for 1 year

Ocular Treatment
• Pyrimethamine and sulphadiazine plus folinic acid to protect the bone
marrow from the toxic effects of pyrimethamine
• Pyrimethamine + clindamycin if the patient (mother patient ) has a
hypersensitivity reaction to sulphur drugs OR
• Cotrimoxazole
• Atovaquone and pyrimethamine + azithromycin
○ Not been extensively studied
• Treatment will not result in the elimination of the organisms from the eye.
Since new lesions can for if the organism reactivates, esp. during
adolescent (carefully monitored)

Prevention and Control

• Cook meat thoroughly to kill the cysts
• Avoid proximity to cats
• Hand-washing before meals and after handling cat litter
• Use gloves when changing cat litter
• Regular treatment of cats
• Blood screening prior to transfusion
• Screening and treatment during pregnancy
• Treatment of cases
• Chemoprophylaxis in HIV patients
○ Cotrimoxazole
○ Fansidar
○ Pyrimethamine-dapsone
• Health education
3. Write short notes of the following
○ Clinical presentation of Onchocerca volvulus
• Incubation period 15 -18M may be asymptomatic
• Pruritus of the skin, rash on skin
• Subcutaneous nodules (onchocercomas) in bony prominences. In Africa,
normally around the hips and the head in S. America
• Lymphadenitis especially in the groin
• Lizard skin - Thickened skin
• Prespyderma
○ Loss of skin elasticity, premature ageing
• Leopard skin
○ Skin depigmentation in patches
• Hanging groin
• Arthritis
○ May be monoarthritis affecting the large joints e.g. hip pain disappears
rapidly on treatment

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 ○
rapidly on treatment
• Elephantiasis of the scrotum in males
• River blindness (onchocerciasis)
• SOWDA
○ Hypersensitive reaction to microfilariae
○ Arabic for dark
○ Results from strong immune response by host
○ Usually localised to one limb
○ Itchy, swelling, dark with scaling papules, enlarged regional lymph nodes
○ Life cycle of Taenia saginata

○ Complications of Ascaris lumbricoides

○ Cutaneous larva migrans
4. Write short notes on each of the following
○ Medical importance of the human body louse (pedunculus humanus corporis)
○ Control of Echinococcus granulosus
• Dogs should not be exposed to offal in slaughterhouses
• Secure slaughter houses (fencing)
• Deworm dogs with praziquantel
• Kill stray dogs
• Avoid dog/man contact
• Inspect meat and cooking before feeding dogs
• Environmental sanitation/protection
• Sewerage treatment
• Health education
○ Laboratory diagnosis of Leishmaniasis
• Skin biopsy for histology
• Biopsy of soft tissues for histology
• Serology ELISA
• PCR
• Clinical picture
○ Treatment of complicated Plasmodium falciparum infection
• IV quinine in 5% dextrose drip
• Artesunate IM
○ Cure rate is similar to quinine

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 ○ Cure rate is similar to quinine
○ Easier to administer
○ Safe in pulmonary oedema
• Rx other existing conditions
○ Hypoglycaemia (glucose IV)
○ Anaemia (transfusion)
○ Convulsions (diazepam or paraldehyde)
○ Renal failure (dialysis)
○ Hyperpyrexia (antipyretics)
○ Pulmonary oedema (diuretics)
5. Write short notes of the following
○ Medical importance of lice
○ Massive Strongyloidiasis
○ Control of Schistosomiasis
• Breed in slow moving/stagnant fresh water
○ Cement the rice paddy banks
○ Increase water flow
• Molluscides
○ Endod (from gooseberry)
○ Copper sulphate
• Biological methods
○ Cray fish to eat the snails
• Empty unused ponds
• Use of PPE
• Sewage treatment
• Health education
○ Fasciolopsis buski
• Morphology
○ Largest fluke
○ Intestinal caecum is not branched
○ Oval shaped
○ Anterior conical projection is round
• Pathology
○ Eggs cause fibrosis in GIT
• Clinical Features
○ NVD
○ Intestinal obstruction
○ Anasaka
○ Ascites
• Diagnosis
○ History of eating undercooked water plants
○ Microscopy
○ Endoscopy
• Treatment
○ Praziquantel
○ Bithionol
• Prevention and Control
○ Treatment
○ Appropriate disposal of waste
○ Proper cooking of vegetation
○ Molluscides
○ Biological methods e.g. Crayfish
○ Health education

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 ○ Health education
6. Write short notes on each of the following
○ Ascaris lumbricoides
○ Clinical presentation, complications and treatment of hookworms
○ Lymphatic filariasis
• Caused by
○ Wuchereria bancrofti
○ Brugia timori and malayi
• Transmitted by
○ Culex - wuchereria
○ Mansonia - malayi
○ Anopheles - timori
• Morphology
○ Adults
○ Thread-like
○ Females longer than males
○ Microfilariae (W. bancrofti)
○ Sheathed
○ Nuclei clearly separated, do not extend to the tip of the tail
○ Microfilariae (B. malayi and timori)
○ Sheathed
○ Nuclei are matted, 3 nuclei extend to the tip of the tail
• Pathology
○ Blockage of lymph passages with adults
○ Macrophages attack
○ Lymph vessels dilate and leads to lymph stasis
○ Formation of granuloma, scarring and calcification
○ It interferes with permeability of lymph vessels leading to oedema
(browny oedema)
• Clinical Presentation
○ Early stages (acute manifestations)
○ Fever, chills
○ Lymphadinitis, lymphangitis, lymphangiovarix
○ Orchitis, epididymitis
○ Late stages (chronic manifestations - blockage of lymphatic flow)
○ Lymphoedema of limbs and breasts
○ Elephantiasis (limbs, breast, scrotum)
 Secondary fungal and bacterial infection
○ Hydrocele
 Fluid in the tunica vaginalis
○ Chyluria (Whitish urine)
○ Tropical pulmonary eosinophilia (TPE)
 Hypersensitivity reaction - presents like an asthmatic attack
□ Dyspnoea
□ Dry cough
□ Wheezing
□ Transient hepatomegaly and splenomegaly
• Diagnosis
○ History of living in the endemic areas of years
○ Physical examination findings
○ Blood examination (night sample) for microfilariae
○ Thick blood smear
○ Capillary tube exam
○ Blood filter (nucleopore)

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 ○ Blood filter (nucleopore)
 Stain with Giemsa stain for analysis
 Fields stain*
○ DEC provocation test (DEC 5-mg, blood >30 min)
○ Diethylcarbamazine*
○ Irritates the microfilariae to retreat to blood during the day
○ Ultrasound of the scrotum - filarial dance sign
○ Serological tests
○ Filarial specific antibodies e.g. IgG, circulating filarial antigen (CFA)
○ Does not ascertain current infection, exposure or post-exposure
• Treatment
○ DEC
○ Should not be used in places where there is onchocerciasis
occurring together with LF
○ Ivermectin
○ Albendazole
○ Combination Rx with DEC or Ivermectin
• Prevention and control
○ Vector control
○ Insecticides
○ Larvicides
○ Personal protection
○ Bed nets
○ Insect repellents
○ Annual treatment - DEC or Ivermectin
○ DEC treated salt
○ Control of schistosomiasis
○ Trichinella spiralis
7. Write short essays on the following
○ Medical importance of fleas
○ Larval migrans
○ Cryptosporidium parvum
• Infects the intestines of immunocompromised patients
• Are parasites of rodents, monkeys cattle and other herbivores but are
relatively unrecognised cause of self-limited mild gastroenteritis and
diarrhoea in humans

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•

• Pathology and pathogenesis

○ It inhabits the brush border of mucosal cells of the GIT, esp. the
surface villi of the lower small bowel
○ Prominent feature of cryptosporidiosis is mild and self-limiting
watery diarrhoea in normal persons but may be prolonged and
severe in the immunocompromised or very young or old individuals
○ Extra-intestinal infection
• Respiratory cryptosporidiosis
• Cholecystitis
• Hepatitis
○ In immunocompromised patients
• Diagnosis
○ Detection of oocysts in fresh stool samples
○ Stool concentration techniques using modified, acid fast stains
○ Monoclonal antibody-based testing can detect low-level infections,
and fluorescent microscopy w/ auramine staining
○ EIA for detection of faecal antigen
• Treatment
○ Self-limiting
○ Rehydration
○ In immunocompromised patients
○ Nitazoxanide
○ ARV treatment

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 ○ ARV treatment
○ Control of hydatid disease
8. Write short notes on each of the following
○ Life cycle and clinical presentation of hookworm infection
○ Enterobius vermicularis
○ Complication of Entamoeba histolytica infection
○ Cysticercosis
• Disease due to T. solium oncospheres
• Acquired through
○ Ingestion of eggs in faeces contaminated food/water
○ Autoinfection in a human infected with adult T. solium by:
 Ingestion of eggs through foetal contamination of food
 Proglottids carried into the stomach by reverse peristalsis
• Eggs in intestine > oncospheres hatch > invade the intestinal wall >striated
muscles, brain, liver and other tissues > develop into cysticerci

Clinical Presentation
• Asymptomatic subcutaneous nodules and calcified IM nodules
• Neural cycticercosis
○ Seizures, mental disturbances, focal neurological deficits and signs of
space-occupying intracerebral lesions/intracranial hypertension >
intracranial herniation
• Extra-cerebral neural cycticercosis
• Extra-cerebral cycticercosis
○ Symptoms due to ocular, cardiac or spinal lesions
•
9. Write short notes on each of the following
○ Taeniasis
• Disease by adult of Taenia species of cestodes involving humans and animal
species as hosts
• Definitive host - humans
• Intermediate hosts
□ Pigs - T. solium
□ Cattle - T. saginata
• Mode of infection
□ Consumption of raw or undercooked meat (pork/beef)
• Cysticercus
□ Infective stage
○ Life Cycle

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  Infected person has adults in the small intestines
 Distal/gravid proglotids are released and excreted in their stool
 If stool is passed in the environment, the animals (grazing cattle/feeding pigs)
are infected with the proglotids and then eggs are released.
 The oncospheres puncture the intestinal mucosa and into blood where they
find their way into striated muscle where they develop into cystecerci (can
survive here for several years)
 Ingestion of cysticerci in raw or undercooked food
 Grow into adults (2 months)
 Adults attach to wall of small intestine of man using their scolex
 Mature/gravid proglotids dislodge and pass in stool
○ Clinical Presentation
 Mild, abdominal symptoms, less with T. solium
 Passage of proglottids per annum
 Intestinal perforation
 Aspiration of vomited proglottids
 Appendicitis - migrating proglottids
 Cholangitis - migrating proglottids
○ Diagnosis
 Stool microscopy useful after3 months of infection
□ Eggs
 Cannot differentiate from other cestodes
□ Mature proglottids
 Lactophenol then indian ink injection into genital pore, count uterine
branches (T. saginata 12-30 uterine branches, T. solium 7-13)
□ Rarely examination of the scolex
 Repeat stool exams and concentration techniques necessary in light infection
□ Zinc floatation
□ Forma-ether technique
 Antibody detection
□ Especially when larval stages present in early infection
○ Treatment
 Praziquantel
 Niclosamide
□ Do not give in T. solium infection as it disintegrates the proglottids and

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 □ Do not give in T. solium infection as it disintegrates the proglottids and
oncospheres are release
○ Life cycle of Echinococcus granulosus
○ Trichinella spiralis
○ Control of schistosomiasis
10. Write short notes of the following
○ Control of hookworm infection
○ Vectors of filariasis
○ Trichomonas
○ Clinical presentation of Ascaris lumbricoides
11. Write short notes on each of the following
○ Control of Echinococcus granulosus
○ Parasites associated with anaemia
○ The pathology caused by Taenia solium infections in humans
○ Diagnosis and treatment of hookworm infection
○ Prevention of African trypanosomiasis
○ Entamoeba histolytica
○ Laboratory diagnosis and treatment of Plasmodium falciparum
○ Diphyllobothrium latum
○ Clinical presentation of visceral leishmaniasis
○ Control of malaria
○ Zoonotic diseases
○ Diagnosis of trypanosomiasis
○ Parasites in immunosuppressive states
○ Visceral larva migrans
○ Myiasis
○ Clinical manifestation and treatment of Echinococcus granulosus infection in
man
○ Antimalarial drug resistance
○ Giardia lamblia
• Causes giardiasis
• Found in the duodenum and jejunum of humans
• Morphology
○ Trophozoite
 Heart shaped
 Four pairs of flagella
 Approx. 15µm in length
 Ventral sucker for adherence on intestinal wall
○ Cysts
 Ellipsoid
 Thick walled
 Highly resistant
 8-14µm in length
 Contain 2 nuclei as immature forms and 4 as mature cysts

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 •

• Pathology and pathogenesis

○ Large number of parasites attached on bowel wall may cause
irritation and low-grade inflammation of duodenal/jejunal mucosa
 Consequent diarrhoea associated w/ crypt hypertrophy, villus
atrophy or flattening, epithelial cell damage
○ Stool may be watery, semisolid, greasy, bulky and foul-smelling
○ Symptoms of
 Malaise
 Weakness
 Weight loss
 Abdominal cramps and distention
 Flatulence
• Humoral
○ Majority produce detectable levels of giardia spp. Antibodies role is
however unclear
○ IgM anti-Giardia antibodies
 Short-lived
 May be important in both defence and parasite clearance
○ IgG anti-Giardia antibodies
 Remain in high titres for many months after treatment
○ There's a certain degree of protein in breastmilk_
○ Laboratory Diagnosis
• Cysts are formed in stool

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 • Cysts are formed in stool
• Trophozoites in diarrhoeal stools by normal saline and iodine preparations
• Parasites are passed in stool cyclically
• Trophozoites seen in bile aspirated from duodenum by intubation and by
duodenal capsule technique
• Monoclonal Ab's using fluorescent method
• Duodenal and jejunal biopsies
• Touch preparations are air dried and fixed with methanol and stained with
Giemsa and trophozoites appear purple
• ELISA and IFA
○ Treatment
• Metronidazole
• Tinidazole
• Furazolidine
○ Prevention
• Improved water supply
• Proper faeces disposal
• Improve personal hygiene
• Proper food storage
• Routine hand washing
• Proper pest control
• Treatment of symptomatic and asymptomatic patients
○ Clinical presentation and diagnosis of visceral leishmaniasis
○ Molluscides
○ Ectoparasites
12. Write short notes on cysticercosis
○ Disease due to T. solium oncospheres
○ Acquired through
 Ingestion of eggs in faeces contaminated food/water
 Autoinfection in a human infected with adult T. solium by:
□ Ingestion of eggs through foetal contamination of food - external
□ Proglottids carried into the stomach by reverse peristalsis - internal
○ Eggs in intestine > oncospheres hatch > invade the intestinal wall >striated muscles,
brain, liver and other tissues > develop into cysticerci
○ Clinical Presentation
 Asymptomatic subcutaneous nodules and calcified IM nodules
 Cerebral cycticercosis
□ Seizures, mental disturbances, focal neurological deficits and signs of
space-occupying intracerebral lesions/intracranial hypertension >
intracranial herniation
□ Found in the brain
 Extra-cerebral neural cycticercosis
□ Found in the spinal cord
 Extra-cerebral cycticercosis
□ Symptoms due to ocular, cardiac or spinal lesions
○ Diagnosis
• Tissue biopsy
• Antibody detection
• Imaging
○ Treatment
• Anti-helminths
○ Albendazole
○ Praziquantel for symptomatic patients with live cysts, but not for
intraventricular cysts
 Risk of precipitating hydrocephalus

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  Risk of precipitating hydrocephalus
□ + corticosteroids e.g. dexamethasone
• Surgery
• Symptomatic treatment
○ Anticonvulsants
○ Prevention and Control
• Faeces disposal
• Treatment
• Health education
13. Describe the life cycle of Diphyllobothrium Latum
○ Life Cycle
• Take about 1 - 2 weeks in water to embryonate
• Hatches to release coracidium
• Ingested by copepods, it loses the cilia and penetrates the mid-gut
• Enters haemocoel
• Metamorphoses into procercoid which is large (550 micrometers long)
• Infected copepod is eaten by a freshwater fish and procercoid develops to
plerocercoid larva
• Strobilation and egg production occurs
• Human infections due to eating raw/undercooked fish
• Causes pernicious megaloblastic anaemia by:
○ Robbing the host of vitamin b12
○ Interferes with the intrinsic factor of castle (gastric intrinsic factor)
14. Lab diagnosis of Echinococcus granulosus
○ Casoni intradermal skin test
○ 0.2 - 0.5ml of irradiated hydatid fluid intradermally. After 15 - 30minutes wheal
will form. (Positive if wheal is formed)
○ Eosinophilia - 20 - 25%
○ Serology
○ Complement fixation test (CFT)
○ ELISA
○ Arc 5 test
○ Immuno-electrofluoresis test
○ Indirect haematoglutination test
○ Imaging
○ X-rays
○ Hydatid cysts in bones
○ CT scan
○ Extra-hepatic cysts
○ Ultrasound scan
○ MRI
○ Difficult to find cyst e.g. in cardiac and spinal tissue
15. Prevention and Control of Schistosomiasis
○ Definitive host
○ Treatment of infected persons to prevent spread
□ Antimony compounds
□ Hycanthone
□ Niridazole
□ Metriphonate
□ Oxamniquine
□ Praziquantel (adults)
□ Artemisinin (larval schistosomes)
○ Transmission
○ Use of PPE to prevent infection from percutaneous infection
○ Decrease movement in still water or increase flow of water

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 ○ Decrease movement in still water or increase flow of water
○ Reduce man-water contact
○ Dry feet after water contact
○ Carrier
○ Molluscides - endod (from gooseberry) and copper sulphate
○ Biological methods
□ Cray fish to eat the snails
○ Sewage treatment
16. Classification of hermaphoditic trematode
○ Is according to the habitat of the trematodes (the adults)
 Intestinal trematodes (intestinal flukes)
□ Small intestines
 F.buski
 H.heterophyes
 M.yokogawai
 W.watsoni
□ Large intestines
 G.hominis
□ Hepatic trematodes (liver flukes)
 C.sinensis
 O.felineus
 F.hepatica
□ Lung trematodes (lung flukes)
 P.westermani
□ Blood trematodes (blood flukes)
 In vesical venous plexuses
◊ S.haematobium
 In rectal and portal venous system
◊ S.mansoni
○ Mode of Infection
• By ingestion of encysted infective stage known as cercariae/metacercariae
○ Vegetables as in F. hepatica, F. buski, W. watsoni
○ Fish as in C. sinensis, H. heterophyes, M. yokogawai
○ Flesh of crab or crayfish as in P. westermani
• Free cercariae penetrating through the skin
○ As in S. mansoni, haematobium and japonicum
17. Clinical Presentation of Oncocerchiasis
○ Itch and pruritus
○ Onchocercoma
• Circumscribed
• Firm
• Painless
○ Ocular manifestation
• The parasite affects visual acquity
• Cause atrophy o optic nerve (river blindness)
○ Hanging groin
• Loss of elasticity
○ Prespyderma
• Loss of skin elasticity due to the proteins from Wolbachia bacteria
○ Xeroderma
• Lizard skin
○ SOWDA
• Dark patches w/ skinning patches due to host immunity
• Is unilateral
18. Life cycle and treatment of Leishmania donovani

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18.
○ Lifecycle
• Definitive host - Sand-fly
• Intermediate host - man
• The sand-fly takes a blood meal and deposits the promastigotes. The
promastigotes are then phagocytosed in the macrophages then turn into
amastigotes. The amastigotes then undergo multiplication and can infect other
cells and tissues. The sand-fly then takes another blood meal and takes up the
macrophages with the amastigotes inside. The amastigote then transforms into
the promastigote stage in the gut of the sand-fly and rapidly divide and move to
the proboscis. The sand-fly takes another blood meal and deposits the
promastigotes and the cycle continues
○ Treatment
• Pentavalent antimonials e.g. Sodium stibogluconate
• Miltefosine
• Pentamidine
• Amphotericin B
• Interferon
19. Differences between the Taenia saginata and solium
Saginata Solium
Adult is longer Shorter
a. No rostellum Rostellum present in scolex
Up to 2000 proglottids 1000 proglottids
12-30 uterine branches 7-13 uterine branches
20. Life cycle and treatment of Leishmania donovani
○ Treatment
• Pentavalent antimonials e.g. Sodium Stibogluconate
• Amphotericin B
• Pentamidine
• Miltefosine
• Aminosidine
• Gamma interferon
• Imidazoles (ketoconazole, itraconazole)
• Allopurinol
○ Life cycle
• Definitive host: sandfly
• Intermediate host: Man

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 •

21. Life cycle of Plasmodium falciparum

○ Definitive host - mosquito
○ Intermediate host - man

22. Clinical presentation of malaria

○ Acute
• Irregular tertian fever (fever reoccurring every 48 hours)
• Headache, NVD, loss of appetite, dizziness, muscle aches, joint pains, general
malaise
• High temperature, jaundice, pallor, abdominal tenderness, splenomegaly,
hepatomegaly

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 hepatomegaly
○ Severe complicated falciparum malaria
• Cerebral malaria, convulsions, hypoglycaemia, hyperpyrexia, severe anaemia,
renal failure, pulmonary oedema or acute respiratory distress syndrome, algid
malaria (shock secondary to bacteraemia or septicaemia), DIC (disseminated
intravascular coagulopathy), haemoglobinuria (black water fever), jaundice,
hyperlactataemia, abortion, premature birth, low birth weight babies
• Hyper-reactive malaria splenomegaly (HMS), previously tropical splenomegaly
syndrome (TSS) - continuous infection w/ malaria
23. Pathology of Trypanosoma brucei gambiense
○ When the tsetse fly bites the host, the trypamastigotes multiply in the skin and cause
swelling that leads to formation of a chancre
○ Trypamastigotes multiply in the blood stream and lymph nodes that leads to
lymphadenopathy prominently in the posterior triangle of the neck (Winterbottom's
Sign)
○ As they travel in the blood stream to other organs e.g. spleen and liver that leads to
hepatosplenomegaly. This leads to jaundice and anaemia
○ In the terminal stages, they spread to the CNS that leads ti progressive diffuse
meningoencephalitis w/ death from the
24. Differences between T B Rhodensiense and gambiense
Characteristic West African East African
s
Organism T. brucei gambiense T. Brucei rhodesiense
Distribution West and Central Africa East and Central Africa
Vector Glossina palpalis group Glossina morsilans group
Reservoir Mainly humans Wild and domestic animals
Virulence Less More
Course of Chronic (Late CNS invasion); Acute (early CNS invasion); less
Disease months to years than 9 months
Parasitemia Low High and appears early
Lymphadenopa Early; prominent Less common
thy
Isolation in No Yes
Rodents
Mortality Low High

25. Pathogenesis of Giardia Lamblia

○ Usually in glandular crypts of duodenojejunual mucosa
○ May cause crypt or villus atrophy or flattening by apoptosis and epithelial cell
damage leading to increased lymphatic infiltration of lamina propria
○ Do not invade tissues but adheres tightly to the intestinal epithelium
○ Attaches to epithelial surface of the duodenum and jejunum and may lead to
duodenitis or jejunitis
○ No inflammatory cells involved
○ Feed on mucosal secretions
○ Variant specific surface proteins (VSSP) of Giardia, play an important role in
virulence and infectivity of the parasite
○ Large number of parasites attached to the bowel wall may cause irritation and low-
grade inflammation
○ May colonise the gall bladder causing biliary colic and jaundice
26. Explain how humans are infected w/ Trypanosoma cruzi
○ Vector - Reduviid bug
27. Diagnosis of leishmaniasis

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27. Diagnosis of leishmaniasis
○ History taking
○ Skin biopsy for histology
○ Biopsy of soft tissues for histology
○ Serology ELISA
○ PCR
28. Complications of E. granulosus
○ Space occupying lesions that can cause increased ICP in mans
○ Cyst rupture can cause severe anaphylactic shock
○ Release of protoscolices can lead to formation of numerous cysts in other locations
○ Can cause fracture of bones if the cysts form in the bones
29. Complications of amoebiasis
○ Intestinal perforations
○ Bacteraemia due to escape of the colonic microflora via the perforations
○ Intestinal obstruction if amoebomas on the intestines grow to a large enough size
○ Liver abscesses that may impair hepatic functions
○ Lung disease as a result of penetration of diaphragm by right lobe liver abscesses

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Virology SAQs
Monday, 23 January 2017 12:56 AM

1. Characteristics of killed whole virus vaccination

○ Immune Response
 Poor; only antibody - no cell mediated immune response
 Response is short-lived and multiple doses are needed
○ Safety
 Inactivated, therefore, cannot replicate in the host and cause disease
 Local reactions at the site of injection may occur
○ Stability
 Efficacy of the vaccine does not rely on the viability of the organisms
 These vaccines tend to be able to withstand more adverse storage conditions
○ Expensive
 To prepare
○ Other Disadvantages
 Extreme care required to ensure no residual live virus is present
 Parenteral administration of killed virus, even when stimulates systemic
antibodies (IgM and IgG) to satisfactory levels, has sometimes limited
protection due to local resistance (IgA) is no induced properly at the natural
portal of entry or primary site of multiplication of the wild virus infection
2. Pathogenesis of Hepatitis A virus
○ Faecal-oral transmission
○ Once it reaches it the intestines, it replicates in the enterocytes then it moves to the
hepatic parenchyma
 GIT manifestations (mention them)
○ Does not affect the hepatocytes, only the liver parenchyma and because it is a naked
virus, it may cause lysis of parenchymal cells during its replications and gives the
hepatic clinical manifestations (mention them)
3. Treatment of HBV
○ There is no specific treatment for acute hepatitis B
○ Chronic hepatitis B infection can be treated with antiviral drugs, but treatment only
slow the progression of cirrhosis, reduce incidence of liver cancer and improve long
term medication
○ WHO recommends the use of oral treatments - Tenofovir or Entecavir, because they
are the most potent drugs to suppress Hep B virus and rarely lead to drug resistance
as compared with other drugs
○ Lamivudine and Tenofovir - RT inhibitors
○ Treatment using interferon injections may be considered in some people in certain
high income settings, as this may shorten treatment duration, but its use is less
feasible in low-resource settings due to high cost (given to HBeAg carriers)
4. Briefly discuss
a. Hepatocellular carcinoma
 Malignant transformation due to insertional mutagenesis as HBV genome
integrates into hepatocyte DNA > Integration can activate cellular oncogene >
Loss of growth control (hence cancer) - this occurs during viral replication
 The constant damage and repair of the hepatocytes by the infection and
immune attack can also lead to mistakes in gene arrangement leading to
activation of cellular oncogenes
 Symptoms are often masked by cirrhosis, which also masks tumour presence
 Diagnosis
□ Imaging
 Angiography to show tumour vascularity
 Treatment

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  Treatment
□ Surgical removal
□ Radiation and chemotherapy
□ Liver transplant
b. Significance of lab tests in viral infection
 Diagnosis of infection
□ Detection of markers of infection e.g. antigens and antibodies (HBV)
□ Histopathology e.g. viral inclusions in cells
 Disease progression
□ E.g. in HBV different antigens and antibodies shown up in different stages
of infection
□ CD4 tire as in HIV infection
 Effectiveness of treatment
□ As in acute HBV where you do not treat acute infection
 Check for immunity
 Research and epidemiology
□ Check the strain that is endemic in the area
 Development of vaccines
c. Transformation of host cells by viruses
 Cellular transformation - stable, heritable change in the growth control of cells
in culture
 Tumour viruses mediate changes in cell behaviour by means of a limited
amount of genetic information
□ Direct acting
 Tumour virus introduces a new "transforming gene" into the cell
□ Indirect acting
 The virus alters the expression of pre-existing cellular gene(s)
 The cell loses control of normal regulation of growth; the DNA repair pathways
are frequently affected, leading to genetic instability and a mutagenic
phenotype
d. Characteristics of cellular proto-oncogenes
5. Write short notes on:
a. Mechanism of action of Acyclovir
 Acyclic nucleoside analogue
 Active against HSV-1, HSV-2 and VZV
 A pro-drug, undergoes 3 phosphorylation steps to its active form
□ Initial phosphorylation by viral thymidine kinase
□ Subsequent di- and tri- phosphorylation by host cell enzymes
 MoA
□ Inhibition of DNA Pol
 Acyclovir triphosphate competes w/ endogenous nucleotides for
DNA Pol
 Incorporated into the growing DNA chain (viral genome)
 Causes chain termination once incorporated
 MoR
□ Reduced production of Viral Thymidine Kinase (TK)
□ Altered TK hence reduced affinity for Acyclovir
□ Altered DNA Pol
b. Viral pathogenesis
 Entry and Primary Replication
 Viral spread and cell tropism
 Cell injury and clinical illness
 Recovery from infection
 Virus shedding
c. Innate immunity
 First line of defence in viral infections
 Consists of

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  Consists of
□ Interferons
□ Cellular components e.g. PMN cells that destroy cell infected by virals
□ Consists of barriers e.g. HCV in the placenta
□ Chemical protection e.g. acidity in the stomach that destroys acid labile
viruses e.g. rhinovirus
d. Virology of HPV
 ICTV classification
□ Family - Papillomaviridae
□ Genus - Papillomavirus
□ Species - human papillomavirus
 Baltimore classification - Class I
 Naked
 Icosahedral
 Small circular dsDNA
 More that 150 subtypes of HPV
 Site of replication - nucleus
6. Briefly discuss localised and systemic viral infection
7. Risk factors for HPV
· Concurrent STI (including other strains of HPV)
· Number of sexual partners
· HPV-induced cervical cancer include early age sexual debut
· Smoking
· High parity
· long-term use of oestrogen
· HIV
8. Prevention of HPV
· Reducing the number of sex partners
· Abstinence
· Long term mutual monogamy with a single uninfected partner
· A reduced number of partner
· Vaccination
9. Viruses associated with VHF
Family Species
Arenaviridae Lassa, Junin, Machupo
Bunyaviridae Rift valley, Crimean-Congo, Hantavirus
·
Flaviviridae Yellow fever, Dengue, West Nile
Togaviridae Chikungunya, O’nyong-nyong, Semlike forest
Filoviridae Ebola, Marburg
10. Characteristics of cellular proto-oncogenes
· They are part of the normal gene makeup
· They are expressed during cell growth
· Have a normal Mendelian Inheritance
· Are always found on the same location of the genome
· Do not have the Long Terminal Repeat (LTR) sequence
· Highly conserved
11. Complications of HAV
· Fulminant hepatitis
· Relapsing
· Cholestatic hepatitis
12. Complications of HDV
· HCC
·

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 · Chronic hepatitis
· Cirrhosis
13. Virology of Respiratory Syncytial Virus
· Family - Paramyxoviridae
· Subfamily - Pneumovirinae
· Genus - Pneumovirus
· Species - RSV
· (-)ssRNA
· Helical nucleopsid
· Enveloped
· Non-segmented
· Linear virus
14. Differentiate between antigenic drift and shift
· Antigenic drift
 Occurs in all influenza viruses
 Gradual accumulation of mutations
 May generate a new strain
 Acquired immunity is no longer effective
 Causes seasonal outbreaks and seasonal epidemics
· Antigenic shift
 Mainly occurs in influenza A
 Due to re-assortment of RNA segments
 Occurs in a cell infected w/ different subtypes (mixing vessel)
 May lead to a new virus subtype
 No previous immunity to the new subtype
 Causes sporadic epidemics/pandemics
15. Management and prevention of influenza A virus
· Treatment
• Supportive
○ ABCs
○ Antipyretics/analgesics
• Chemotherapy
○ Amantidine/Rimantidine
○ Zanamavir/Oseltamivir/Peramivir
○ Agents that target RNA pol e.g. Ribavirin
• Antibiotics
○ Useful if there is secondary bacterial infection
• Uncoating inhibitors
□ Amantadine/Rimantadine
 Prevent uncoating
 Block M2 ion channels - viral protein
◊ Prevent acidification of the endosome
• Viral Release Inhibitors
□ Zanamivir, Oseltamivir, Peramivir
□ Block viral neuroaminidase
 Prevents viral spread to uninfected cells
· Prevention
 Vaccines
□ Inactivated
 Trivalent inactivated Vaccine
 A quadrivalent inactivated vaccine
 Monovalent inactivated vaccine
□ Live attenuated influenza virus given as intranasal spray - quadrivalent
 Chemoprophylaxis - expensive
□ The same as those for treatment
□ Given 24hours post-exposure

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 □ Given 24hours post-exposure
· Control of pandemic influenza
 Surveillance
 Screening
 Information sharing
 Health education
 Health worker capacity
 Drugs - prophylaxis
 Vaccination - annual
 Culling animals - chicken/ swine
 Limiting travel
 Isolation of infected persons
16. Lab diagnosis of prionoses
· Clinical presentation
· Exclude differentials
· CSF analysis (for 14-3-3 protein and/or Tau protein)
· EEG
· MRI
· Brain biopsy (post-mortem)
17. Explain the following lab diagnostic techniques, using one virus as an example
· EM
• Used to visualise the viral particles
• Dane particles in HBV infection
• Or visualisation in the plasma and foetal tissue in parvovirus infection
· IFA - influenza (uses direct technique)
• Use monoclonal antibodies (MoAbs) labelled w/ a fluorescent dye
• MoAbs bind to specific epitope on viral protein
• Visualise infected cells using fluorescent microscopy
• Only virus-infected cells will fluoresce
• Direct method
○ Fluorescein tagged antibody → attached fluorescein tagged antibody
visualised by UV microscope
• Indirect method
○ First step
 Untagged antibody → antibody attached to antigenic determinant
○ Second step
 Fluorescein tagged anti-immunoglobulin + antigen attached to
antigenic determinant → attached fluorescein tagged anti-
immunoglobulin visualised by UV microscopy
· ELISA - HIV
• Antigen or antigen detection
• _
· PCR - DNA viruses
• The process is designed to amplify a desired gene or short DNA fragment
• Process involves
○ Denaturation
 The temperature of the reaction tube is increases to 92 - 96°C to
break apart the DNA strands
○ Annealing
 The temperature is dropped to about 45 -65°C which allows the
primers to anneal their complementary strands on opposite template
strands
○ Extension
 The temperature is increased to 72°C to allow the polymerase to
synthesise DNA complementary to the primer
18. Differentiate between naked and envelope viruses

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18. Differentiate between naked and envelope viruses
Properties Enveloped Naked
Environmental Tactile, destroyed by acids, Stable to temperature acids,
stability detergents, drying detergents, proteases,
disinfectants
Release from Budding and cell lysis Cell lysis
cells

· Spread to Spreads in larger droplets, Spread easily (by direct

hosts secretions, transplanted organs, contact and dust and small air
blood transfusion droplets)
Stability Must stay wet Can dry out but retain
outside cells infectivity
Effective May need antibody and cell Antibody may be sufficient for
Immune mediated immunity for protection immunoprotection
response
19. Discuss how HPV causes cancer
· HPV causes cancer through its capsid proteins, E6 and E7
• E6 binds to p53 (a tumour suppressor gene that holds the cell at the G1 phase)
inhibiting it and destroying it. This in turn leads to uncontrolled cellular growth
as cells undergo rapid replication
• E7 binds to Rb (a tumour suppressor that binds to E2f that stimulates cell
replication) Once Rb is bound to E7, E2f is left free and causes rapid
replication of cells, leading to tumour formation
20. Common characteristics of Retroviruses
· 3 common genes: gag, pol and env
· 2 copies of (+)ssRNA
· Only true diploid viruses
· Only positive sense RNA virus whose genome does not serve directly as mRNA
immediately after infection
· Replicates in the nucleus instead of the cytoplasm like other RNA viruses (together
with influenza viruses)
21. Briefly compare and contrast the following viruses
a. Measles and Rubella
22. Briefly discuss the differences between killed and attenuated viral vaccines
Characteristic Killed vaccine Live attenuated
Number of doses Multiple Single
Need for adjuvant Yes No
Duration of immunity Shorter Longer
Effectiveness of protection Lower Greater
Immunoglobulins released IgG IgA and IgG
Mucosal immunity Poor Yes
Cell-mediated immunity Poor Yes
Residual virulent virus in vaccine Possible No
Reversion to virulence No Possible
Excretion of vaccine virus and No Possible
transmission to nonimmune
contacts
Interference by other viruses in host No Possible
Stability at room temperature High Low

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 Stability at room temperature High Low
23. Compare and contrast the virology and pathogenesis of measles and rubella
○ Similarities
• Both are RNA viruses
• Both are enveloped
• Both cause maculopapular rash
• Botha re transmitted via respiratory droplets
○ Differences
Measles Rubella
Paramyxoviridae family Togaviridae family
Genus - mobilivirus Genus - rubivirus
• Negative sense RNA genome Positive sense RNA genome
Helical nucleocapsid Icosahedral nucelocapsid
Surface spike has HA and F proteins Surface spike only has HA
Involvement of CNS causing encephalitis Rarely involves CNS
24. Antiviral immunity
○ Innate components
• Interferon
○ Secreted by virally infected cells as well as inflammatory cells
○ MoA
 Prevention of viral replication within the cell
 Activation of NK cells
◊ Kill virally infected cells
○ Specific component
• Humoral immunity
○ Antibodies
 Neutralise free virus particles
 Prevent viral attachment to its receptors on the cells
 Cause agglutination of virus cells
 Opsonise virus particles
 Complement mediated lysis
• Cell mediated
○ T helper cells recognise viral antigen and secretes cytokines that activate
macrophages and cytotoxic T lymphocytes
○ Cytotoxic T cells kill virally in

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