Committed to the advancement of Clinical & Industrial Disinfection & Microbiology

VOLUME - VI ISSUE - IV OCT-NOV 2013

Editorial

We would like to thank all our valued readers for appreciating our previous efforts with the
intention of still going higher.

Contents As we have already seen some of the Chemical Disinfectants in the last issue; let's see some
more of them like Chlorine and Chlorine Compounds, Quaternary Ammonium
Compounds, Biguanides (Chlorhexidine, Polyhexamethylene biguanide (PHMB),
Iodophors & Miscellaneous Inactivating Agents).

Editorial
n 1 Blood is one of the most important sample received in the laboratory for the culture
and sensitivity and is a sterile body fluid. The diversity of bacteria that is recovered
from blood requires an equally diverse and large number of media to enhance the
growth of these bacteria. A unique cap design of the blood culture bottles to prevent from
Mini review
n 2 the contamination and incorporation of the liquid, SPS (sodium polyanetholsulfonate) at a
concentration of 0.05% makes the ready prepared BHI as a better option for the blood
culture.
Current Trends
n 6
Our In Profile Scientist of the month is none other than Dr. A. P. J. Abdul Kalam, a
renowned aerospace engineer, professor (of Aerospace engineering), and first Chancellor
of the Indian Institute of Space Science and Technology (IIST), who served as the 11th
In Profile
n 9 President of India from 2002 to 2007.

Our Bug of the month is Neisseria gonorrhoeae, also known as gonococci (plural), or
Relaxed Mood
n 10 gonococcus (singular), a species of Gram-negative coffee bean-shaped diplococci bacteria
responsible for the sexually transmitted infection gonorrhea.

Prebiotics are non-digestible food ingredients that stimulate the growth and/or activity of
Bug of the
n Month 11 bacteria in the digestive system in ways claimed to be beneficial to health. A prebiotic is a
selectively fermented ingredient that allows specific changes, both in the composition
and/or activity in the gastrointestinal microflora that confers benefits upon host well-being
Did you Know
n 13 and health. Probiotics are live bacteria that may confer a health benefit on the host.

In this section let's see the WHO guidelines on Hand Hygiene. Pathogens transferred not
only from infected or draining wound but also from frequently colonized areas of normal,
Best Practices
n 14 intact patient skin. The perineal or inguinal areas tend to be most heavily colonized, but the
axillae, trunk, and upper extremities (including the hands) are also frequently colonized.
Hand washing with soap and water has been considered a measure of personal hygiene
In Focus
n 16 for centuries and has been generally embedded in religious and cultural habits.

Enjoy the humour with our Relaxed Mood section.

Your inputs are a valuable contribution towards making this Journal more successful
& looking forward for your continuous support & appreciation.

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 Mini Review OCT-NOV 2013

Chemical Sterilization Techniques

Contd............. another study, chlorine dioxide solutions at either 600 ppm or 30
We have already discussed some of the Chemical Disinfectants in ppm killed Mycobacterium avium-intracellulare within 60
the last issue; let's see some more of them........... seconds after contact but contamination by organic material
significantly affected the microbicidal properties.
Chlorine and Chlorine Compounds Although the United Kingdom manufacturer claims that
Overview: superoxidized water is noncorrosive and non-damaging to
Hypochlorites, the most widely used endoscopes and processing equipment, one flexible endoscope
of the chlorine disinfectants, are manufacturer (Olympus Key-Med, United Kingdom) has voided
available as liquid (e.g., sodium the warranty on the endoscopes if superoxidized water is used to
hypochlorite) or solid (e.g., calcium disinfect them. As with any germicide formulation, the user
hypochlorite). The most prevalent chlorine products are aqueous should check with the device manufacturer forcompatibility with
solutions of 5.25%-6.15% sodium hypochlorite, usually called the germicide.
household bleach. They have a broad spectrum of antimicrobial
activity, do not leave toxic residues, are unaffected by water Mode of Action: The exact mechanism by which free chlorine
hardness, are inexpensive and fast acting, remove dried or fixed destroys microorganisms has not been elucidated. Inactivation by
organisms and biofilms from surfaces, and have a low incidence chlorine can result from a number of factors: oxidation of
of serious toxicity. Sodium hypochlorite at the concentration used sulfhydryl enzymes and amino acids; ring chlorination of amino
in household bleach (5.25-6.15%) can produce ocular irritation or acids; loss of intracellular contents; decreased uptake of
oropharyngeal, esophageal, and gastric burns. Other nutrients; inhibition of protein synthesis; decreased oxygen
disadvantages of hypochlorites include corrosiveness to metals in uptake; oxidation of respiratory components; decreased
high concentrations (>500 ppm), inactivation by organic matter, adenosine triphosphate production; breaks in DNA; and
discoloring or “bleaching” of fabrics, release of toxic chlorine gas depressed DNA synthesis. The actual microbicidal mechanism of
when mixed with ammonia or acid (e.g., household cleaning chlorine might involve a combination of these factors.
agents), and relative stability. The microbicidal activity of
chlorine is attributed largely to undissociatedhypochlorous acid 4 NaClO + 2 H2O → 4 Na+ + 4 OH- + 2 Cl2 + O2
(HOCl). The dissociation of HOCI to the less microbicidal form
(hypochlorite ion OCl-) depends on pH and its own concentration.
The disinfecting efficacy of chlorine decreases with an increase Microbicidal Activity: Low concentrations of free available
in pH that parallels the conversion of undissociated HOCI to OCl- chlorine (e.g., HOCl, OCl-, and elemental chlorine Cl2) have a
. NaOCl stored at room temperature quickly loses strength (up to biocidal effect on mycoplasma (25 ppm) and vegetative bacteria
40%-50%) due to the high rates of free available chlorine level (<5 ppm) in seconds in the absence of an organic load. Higher
over 1 month. A potential hazard is production of the carcinogen concentrations (1,000 ppm) of chlorine are required to kill M.
bis (chloromethyl) ether when hypochlorite solutions contact tuberculosis. A concentration of 100 ppm will kill >99.9% of B.
formaldehyde and production of the carcinogen trihalomethane atrophaeus spores within 5 minutes and destroy mycotic agents
when hot water is hyperchlorinated. in <1 hour. Acidified bleach and regular bleach (5,000 ppm
Alternative compounds that release chlorine and are used in the chlorine) can inactivate 106 Clostridium difficile spores in <10
health-care setting include demand-release chlorine dioxide, minutes. One study reported that 25 different viruses were
sodium dichloroisocyanurate, and chloramine-T. The advantage inactivated in 10 minutes with 200 ppm available chlorine.
of these compounds over the hypochlorites is that they retain Several studies have demonstrated the effectiveness of diluted
chlorine longer and so exert a more prolonged bactericidal effect. sodium hypochlorite and other disinfectants to inactivate HIV.
Chlorine (500 ppm) showed inhibition of Candida after 30
Sodium dichloroisocyanurate tablets are stable, and for two seconds of exposure. Because household bleach contains
reasons, the microbicidal activity of solutions prepared from 5.25%–6.15% sodium hypochlorite, or 52,500–61,500 ppm
sodium dichloroisocyanurate tablets might be greater than that of available chlorine, a 1:1,000 dilution provides about 53-62 ppm
sodium hypochlorite solutions containing the same total available chlorine, and a 1:10 dilution of household bleach
available chlorine. First, with sodium dichloroisocyanurate, only provides about 5250-6150 ppm.
50% of the total available chlorine is free (HOCl and OCl-),
In 1986, a chlorine dioxide product was voluntarily removed
whereas the remainder is combined (monochloroisocyanurate or
from the market when its use caused leakage of cellulose-based
dichloroisocyanurate), and as free available chlorine is used up,
dialyzer membranes, which allowed bacteria to migrate from the
the latter is released to restore the equilibrium. Second, solutions
dialysis fluid side of the dialyzer to the blood side.
of sodium dichloroisocyanurate are acidic, whereas sodium
hypochlorite solutions are alkaline, and the more microbicidal Sodium dichloroisocyanurate at 2,500 ppm available chlorine is
type of chlorine (HOCl) is believed to predominate. In vitro effective against bacteria in the presence of up to 20% plasma,
suspension tests showed that solutions containing about 140 ppm compared with 10% plasma for sodium hypochlorite at 2,500
chlorine dioxide achieved a reduction factor exceeding 106 of S. ppm.
aureus in 1 minute and of Bacillus atrophaeus spores in 2.5 “Superoxidized water” has been tested against bacteria,
minutes in the presence of 3 g/L bovine albumin. The potential for mycobacteria, viruses, fungi, and spores. Freshly generated
damaging equipment requires consideration because long-term superoxidized water is rapidly effective (<2 minutes) in
use can damage the outer plastic coat of the insertion tube. In achieving a 5-log10 reduction of pathogenic microorganisms (i.e.,

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 OCT-NOV 2013 Mini Review
M. tuberculosis, M. chelonae, poliovirus, HIV, multidrug- Mode of Action: The bactericidal action of the quaternaries has
resistant S. aureus, E. coli, Candida albicans, Enterococcus been attributed to the inactivation of energy-producing enzymes,
faecalis, P. aeruginosa) in the absence of organic loading. denaturation of essential cell proteins, and disruption of the cell
However, the biocidal activity of this disinfectant decreased membrane746. Evidence exists that supports these and other
substantially in the presence of organic material (e.g., 5% horse possibilities.
serum). No bacteria or viruses were detected on artificially
contaminated endoscopes after a 5-minute exposure to Microbicidal Activity: Results from manufacturers' data sheets
superoxidized water 551 and HBV-DNA was not detected from and from published scientific literature indicate that the
any endoscope experimentally contaminated with HBV-positive quaternaries sold as hospital disinfectants are generally
mixed sera after a disinfectant exposure time of 7 minutes. fungicidal, bactericidal, and virucidal against lipophilic
(enveloped) viruses; they are not sporicidal and generally not
Uses: tuberculocidal or virucidal against hydrophilic (nonenveloped)
Hypochlorites are widely used in healthcare facilities in a variety viruses. Some Pseudomonas sp. can even grow in solutions of
of settings. Inorganic chlorine solution is used for disinfecting quats, eg. Cetrimide based preparations. The poor
tonometer heads and for spot-disinfection of countertops and mycobactericidal activities of quaternary ammonium
floors. Because hypochlorites and other germicides are compounds have been demonstrated. Quaternary ammonium
substantially inactivated in the presence of blood, large spills of compounds effectively (>95%) remove andinactivate
blood require that the surface be cleaned before final solution of contaminants (i.e., multidrug-resistant S. aureus, vancomycin-
bleach is applied. Extreme care always should be taken to prevent resistant Entercoccus, P. aeruginosa) from computer keyboards
percutaneous injury. At least 500 ppm available chlorine for 10 with a 5-second application time. No functional damage or
minutes is recommended for decontaminating CPR training cosmetic changes occurred to the computer keyboards after 300
manikins. Other uses in healthcare include as a disinfectant for applications of the disinfectants.
manikins, laundry, dental appliances, hydrotherapy tanks
regulated medical waste before disposal, and the water Generations of QACs
distribution system in hemodialysiscenters and hemodialysis Several generations of Qats developed with better biocidal
machines. activity & tolerance to organic matter.

Quaternary Ammonium Compounds First Generation: Have lowest relative biocidal activity. Eg:
Benzalkonium chloride, Cetrimide etc.
Overview: The quaternary ammonium compounds are widely
used as disinfectants. Health-care–associated infections have Benzlkonium chloride (BKC)
been reported from contaminated quaternary ammonium Rapidly acting biocidal agent with a moderate residual action.
compounds used to disinfect patient-care supplies or equipment, Lowest relative biocidal activity.
such as cystoscopes or cardiac catheters. The quaternaries are
good cleaning agents, but high water hardness and materials such Cetrimide
as cotton and gauze pads can make them less microbicidal Used as selective media for the growth of P. aeruginosa. Widely
because of insoluble precipitates or cotton and gauze pads absorb used in the examination of cosmetics, pharmaceuticals & clinical
the active ingredients, respectively. One study showed a specimens to test for the presence of P. aeruginosa.
significant decline (~40%–50% lower at 1 hour) in the
concentration of quaternaries released when cotton rags or Second Generation: Substitution of the aromatic ring hydrogen
cellulose-based wipers were used in the open-bucket system, of BKC with chlorine, methyl and ethyl groups results in second
compared with the nonwoven spunlace wipers in the closed- generation quat with higher biocidal activity. Eg: Alkyl dimethyl
bucket system. As with several other disinfectants (e.g., benzyl ammonium chloride (ADBAC).
phenolics, iodophors) gram-negative bacteria can survive or
grow in them. Third Generation: "Dual Quats". Mixture of two specific quats
with increased biocidal activity, stronger detergency & relative
Chemically, the quaternaries are organically substituted lower toxicity. Eg: Equal mixture of alkyl dimethyl benzyl
ammonium compounds in which the nitrogen atom has a valence ammonium chloride + alkyl dimethyl ethylbenzyl ammonium
of 5, four of the substituent radicals (R1-R4) are alkyl or chloride (ADEAC).
heterocyclic radicals of a given size or chain length, and the fifth
(X-) is a halide, sulfate, or similar radical. Each compound Fourth Generation: "Twin or Dual Chain Quats". Superior in
exhibits its own antimicrobial characteristics, hence the search germicidal performance, lower foaming & an increased tolerance
for one compound with outstanding antimicrobial properties. to protein loads and hard water. Eg: Didecyl dimethyl ammonium
Some of the chemical names of quaternary ammonium chloride or dioctyl dimethyl ammonium chloride.
compounds used in healthcare are alkyl dimethyl benzyl Didecyl dimethyl ammonium chloride.
ammonium chloride, alkyl didecyl dimethyl ammonium
chloride, and dialkyl dimethyl ammonium chloride. The newer Fourth generation (twin or dual chain quats). Stronger/superior
quaternary ammonium compounds (i.e., fourth generation), antimicrobial activity. Increased tolerance to hard water and
referred to as twin-chain or dialkyl quaternaries (e.g. didecyl organic matter. Low foaming
dimethyl ammonium bromide and dioctyl dimethyl ammonium
bromide), purportedly remain active in hard water and are Target - Cytoplasmic membrane
tolerant of anionic residues. Disruption of intermolecular interactions. Dissociation of lipid
bilayers – increased permeability of cell membrane, leakage of
A few case reports have documented occupational asthma as a cytoplasmic contents. Deactivation of enzymes responsible for
result of exposure to benzalkonium chloride. respiration and metabolic cellular activities

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Fifth Generation: Mixtures of fourth generation quats with Overview. Iodine solutions or tinctures long have been used by
second-generation quats. Outstanding germicidal performance, health professionals primarily as antiseptics on skin or tissue.
active under more hostile conditions & are safer to use. Eg: Iodophors, on the other hand, have been used both as antiseptics
DDAC + ADBAC anddisinfectants. Regulatory bodies has not cleared any liquid
chemical sterilant or high-level disinfectants with iodophors as
Uses: The quaternaries commonly are used in ordinary the main active ingredient. An iodophor is a combination of
environmental sanitation of noncritical surfaces, such as floors, iodine and a solubilizing agent or carrier; the resulting complex
furniture, and walls. EPA-registered quaternary ammonium provides a sustained-release reservoir of iodine and releases
compounds are appropriate to use for disinfecting medical small amounts of free iodine in aqueous solution. The best-
equipment that contacts intact skin (e.g., blood pressure known and most widely used iodophor is povidone-iodine, a
cuffs).Used as disinfectants, surfactants, fabric softeners & compound of polyvinylpyrrolidone with iodine. This product and
antistatic agents. other iodophors retain the germicidal efficacy of iodine.
Several reports that documented intrinsic microbial
Biguanides contamination of antiseptic formulations of povidone-iodine and
Biguanides include both mono and polymeric forms poloxamer-iodine. “Free” iodine (I2) contributes to the
Chlorhexidine (Monomeric biguanides) bactericidal activity of iodophors and dilutions of iodophors
demonstrate more rapid bactericidal action than does a full-
Available as dihydrochloride, diacetate & gluconate. Has a wide strength povidone-iodine solution. The reason for the observation
spectrum of antibacterial activity. Also tuberculocidal in that dilution increases bactericidal activity is unclear, but dilution
ethanolic solutions. Is sporicidal at 98-100°C. Addition of low of povidone-iodine might weaken the iodine linkage to the carrier
concentrations (0.5% - 1.0%) of Chlorhexidine to alcohol based polymer with an accompanying increase of free iodine in
preparations result in greater residual activity. solution.
Target - Lipids present in cell membrane. Binds to phosphate and
fatty acids of phospholipids. Membrane damage. Leakage of cell Mode of Action: Iodine can penetrate the cell wall of
components. Cytoplasmic coagulation at high concentration. microorganisms quickly, and the lethal effects are believed to
result from disruption of protein and nucleic acid structure and
Polyhexamethylene biguanide (PHMB) synthesis.
It is a Polymeric biguanide. Used for disinfection of skin. It has
very low toxicity to higher organisms such as human cells.
PHMB is not cytotoxic. Antimicrobial activity increases with
increasing polymer length.
Target-
Cytoplasmic membrane, DNA. Polymer strands are incorporated
into the bacterial cell wall. Disrupts the membrane and reduces its
permeability. Complete loss of membrane functions.
Precipitation of intracellular constituents, leakage of cytoplasmic
contents - lethal for bacteria. Also binds to bacterial DNA, alterits
transcription &causes lethal DNA damage.
Uses:
Microbicidal Activity: Published reports on the in vitro
l
Chorhexidine is a well know anti-plaque biocide that plays a antimicrobial efficacy of iodophors demonstrate that iodophors
crucial role in reduction of supragingival plaque and treatment are bactericidal, mycobactericidal, and virucidal but can require
of gingivitis. prolonged contact times to kill certain fungi and bacterial spores.
l
PHMB is deployed as a part of wound care system. Three brands of povidone-iodine solution have demonstrated
more rapid kill (seconds to minutes) of S. aureusand M.
l
PHMB formulations are now widely deployed in clinical chelonaeat a 1:100 dilution than did the stock solution. The
applications such as the treatment of Acanthamoeba keratinitis virucidal activity of 75–150 ppm available iodine was
etc. demonstrated against seven viruses. Other investigators have
l
PHMB has been used in contact lens cleaning formulations. questioned the efficacy of iodophors against poliovirus in the
l
PHMB is also used as disinfectant for effective reduction of presence of organic matter and rotavirus SA-11 in distilled or
legionella in water cooling system. tapwater. Manufacturers' data demonstrate that commercial
iodophors are not sporicidal.
Iodophors
Uses: Besides their use as an antiseptic, iodophors have been
used for disinfecting blood culture bottles and medical
equipment, such as hydrotherapy tanks, thermometers, and
endoscopes. Antiseptic iodophors are not suitable for use as hard-
surface disinfectants because of concentration differences.
Iodophors formulated as antiseptics contain less free iodine than
do those formulated as disinfectants 376. Iodine or iodine-based
antiseptics should not be used on silicone catheters because they
can adversely affect the silicone tubing.

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MISCELLANEOUS INACTIVATING AGENTS radiation has several potential applications, but unfortunately its
Other Germicides germicidal effectiveness and use is influenced by organic matter;
Several compounds have antimicrobial activity but for various wavelength; type of suspension; temperature; type of
reasons have not been incorporated into the armamentarium of microorganism; and UV intensity, which is affected by distance
health-care disinfectants. These include mercurials, sodium and dirty tubes. The application of UV radiation in the health-care
hydroxide, â-propiolactone, chlorhexidine gluconate, cetrimide- environment (i.e., operating rooms, isolation rooms, and biologic
chlorhexidine, glycols (triethylene and propylene), and the Tego safety cabinets) is limited to destruction of airborne organisms or
disinfectants. Two authoritative references examine these agents inactivation of microorganisms on surfaces. The effect of UV
in detail. radiation on postoperative wound infections was investigated in a
A peroxygen-containing formulation had marked bactericidal double-blind, randomized study in five university medical
action when used as a 1% weight/volume solution and virucidal centres. After following patients over a 2-year period, the
activity at 3%, but did not have mycobactericidal activity at investigators reported the overall wound infection rate was
concentrations of 2.3% and 4% and exposure times ranging from unaffected by UV radiation, although postoperative infection in
30 to 120 minutes. It also required 20 hours to kill B. atrophaeus the “refined clean” surgical procedures decreased significantly
spores. A powder-based peroxygen compound for disinfecting (3.8%–2.9%). No data support the use of UV lamps in isolation
contaminated spill was strongly and rapidly bactericidal. rooms, and this practice has caused at least one epidemic of UV-
induced skin erythema and keratoconjunctivitis in hospital
In preliminary studies, nanoemulsions (composed of detergents patients and visitors.
and lipids in water) showed activity against vegetative bacteria,
enveloped viruses and Candida. This product represents a Pasteurization
potential agent for use as a topical biocidal agent. Pasteurization is not a sterilization process; its purpose is to
New disinfectants that require further evaluation include destroy all pathogenic microorganisms. However, pasteurization
glucoprotamin, tertiary amines and a light-activated does not destroy bacterial spores. The time-temperature relation
antimicrobial coating. Several other disinfection technologies for hot-water pasteurization is generally ~70oC (158oF) for 30
might have potential applications in the healthcare setting. minutes. The water temperature and time should be monitored as
part of a quality-assurance program. Pasteurization of respiratory
Metals as Microbicides therapy and anesthesia equipment is a recognized alternative to
Comprehensive reviews of antisepsis, disinfection, and anti- chemical disinfection. The efficacy of this process has been tested
infective chemotherapy barely mention the antimicrobial activity using an inoculum that the authors believed might simulate
of heavy metals. Nevertheless, the anti-infective activity of some contamination by an infected patient. Use of a large inoculum of
heavy metals has been known since antiquity. Heavy metals such P. aeruginosa or Acinetobacter calcoaceticus in sets of
as silver have been used for prophylaxis of conjunctivitis of the respiratory tubing before processing demonstrated that machine-
newborn, topical therapy for burn wounds, and bonding to assisted chemical processing was more efficient than machine-
indwelling catheters, and the use of heavy metals as antiseptics or assisted pasteurization with a disinfection failure rate of 6% and
disinfectants is again being explored. Inactivation of bacteria on 83%, respectively. Other investigators found hot water
stainless steel surfaces by zeolite ceramic coatings containing disinfection to be effective (inactivation factor >5 log10) against
silver and zinc ions has also been demonstrated. multiple bacteria, including multidrug-resistant bacteria, for
disinfecting reusable anesthesia or respiratory therapy
Metals such as silver, iron, and copper could be used for equipment.
environmental control, disinfection of water, or reusable medical
devices or incorporated into medical devices (e.g., intravascular Flushing- and Washer-Disinfectors
catheters). A comparative evaluation of six disinfectant Flushing- and washer-disinfectors are automated and closed
formulations for residual antimicrobial activity demonstrated equipment that clean and disinfect objects from bedpans and
that only the silver disinfectant demonstrated significant residual washbowls to surgical instruments and anesthesia tubes. Items
activity against S. aureus and P. aeruginosa. Preliminary data such as bedpans and urinals can be cleaned and disinfected in
suggest metals are effective against a wide variety of flushing-disinfectors. They have a short cycle of a few minutes.
microorganisms. They clean by flushing with warm water, possibly with a
Clinical uses of other heavy metals include copper-8- detergent, and then disinfect by flushing the items with hot water
quinolinolate as a fungicide against Aspergillus, copper-silver or with steam. Because this machine empties, cleans, and
ionization for Legionella disinfection, organic mercurials as an disinfects, manual cleaning is eliminated, fewer disposable items
antiseptic (e.g., mercurochrome) and preservative/disinfectant are needed, and fewer chemical germicides are used. A
(e.g., thimerosal [currently being removed from vaccines]) in microbiologic evaluation of one washer/disinfector
pharmaceuticals and cosmetics. demonstrated complete inactivation of suspensions of E. faecalis
or poliovirus. Other studies have shown that strains of
Ultraviolet Radiation (UV) Enterococcus faecium can survive the British Standard for heat
The wavelength of UV radiation ranges from 328 nm to 210 nm disinfection of bedpans (80ºC for 1 minute). The significance of
(3280 A to 2100 A). Its maximum bactericidal effect occurs at this finding with reference to the potential for enterococci to
240–280 nm. Mercury vapor lamps emit more than 90% of their survive and disseminate in the health-care environment is
radiation at 7 nm, which is near the maximum microbicidal debatable. These machines are available and used in many
activity. Inactivation of microorganisms results from destruction European countries.
of nucleic acid through induction of thymine dimers. UV Surgical instruments and anesthesia equipment are more difficult
radiation has been employed in the disinfection of drinking water, to clean. They are run in washer-disinfectors on a longer cycle of
air, titanium implants, and contact lenses. Bacteria and viruses are approximately 20–30 minutes with a detergent. These machines
more easily killed by UV light than are bacterial spores. UV also disinfect by hot water at approximately 90°C.

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Ready Prepared BHI Broth: Better recovery of

blood isolates
Blood streams infections are considered as the leading cause of albicans. Furthermore, Weinstein's personal observation is that
mortality and morbidity among the health care infection and its the following organisms almost always represent a true infection
early diagnosis and prompt treatment is crucial for the better when isolated from a blood culture: Streptococcus pyogenes,
management of the patients. Blood is one of the most important Streptococcus agalactiae, Listeria monocytogenes, Neisseria
sample received in the laboratory for the culture and sensitivity meningitidis, Neisseria gonorrhoeae, Haemophilus influenzae,
and is a sterile body fluid. Generally blood culture is performed to members of the Bacteroides fragilis group, all Candida species,
diagnose the etiological agents of infections like enteric fever, and Cryptococcus neoformans.
pneumonia, endocarditis, brucellosis, meningitis, pyelonephritis,
infective conditions in the peripheral cardiovascular system and Along the same vein, certain organisms have been found to
for those pathogens entering the blood from abscesses, infected represent contamination in a significant proportion of cases.
wounds etc. These organisms include coagulase-negative staphylococci,
Corynebacterium species, Bacillus species other than Bacillus
Blood cultures are obtained from patients prior to instituting anthracis, Propionibacterium acnes, Micrococcus species,
antibiotic therapy in an effort to identify etiologic viridians group streptococci, enterococci, and Clostridium
microorganisms, focus antibiotic therapy, determine optimal perfringens. However, it is crucial to recognize that each of these
duration of treatment, and guide further interventions. organisms can also represent true bacteremias with devastating
Unfortunately, a small proportion of blood culture yield true- consequences, particularly if untreated due to misinterpretation
positive results and inappropriately ordering blood cultures may as contaminants. Of these organisms, the ones that are thought to
be both wasteful and harmful. When blood cultures are represent true bacteremia only rarely are Corynebacterium
contaminated by skin commensals, repeat blood cultures are species, Bacillus species other than B. anthracis, and
obtained which may prolong hospitalization and unnecessary Propionibacterium acnes.
antibiotic administration.
Despite the high likelihood that certain organisms usually
DETECTION OF CONTAMINATED BLOOD CULTURES represent contaminants when isolated from blood cultures,
Despite its limitations, the blood culture remains the “gold determining the likelihood of true bacteremia can be challenging
standard” for the detection of bacteremia. An accurate for clinicians. Coagulase-negative staphylococci represent an
interpretation of culture results is critical not only from the important, all-too-frequent case in point. In the past, coagulase-
perspective of individual patient care but also from the standpoint negative staphylococci were usually believed to represent
of hospital epidemiology and public health. The tracking and contamination when isolated from blood cultures. In fact,
reporting of nosocomial infections and monitoring of coagulase-negative staphylococci are the most common blood
bloodstream infection rates are both essential infection control culture contaminants, typically representing 70% to 80% of all
activities that depend heavily on the accurate differentiation of contaminated blood cultures.
contamination from true bacteremia. Reliably making this
determination continues to be very challenging for clinicians, Recently, however, studies have shown that these organisms are
epidemiologists, and microbiologists. In recent decades, multiple an increasing source of true bacteremia in patients with prosthetic
approaches have been studied, advocated, and utilized for this devices and central venous catheters, although the majority of
purpose. Clues that may help to differentiate contamination from isolates of coagulase-negative staphylococci from blood cultures
bacteremia include identity of the organism, number of positive continue to be contaminants. Weinstein et al. found that even
culture sets, number of positive bottles within a set, time to though only 12.4% of coagulase-negative staphylococcal isolates
growth, quantity of growth, clinical and laboratory data, source of were clinically significant, they ranked as the third most common
culture, and automated classification using information cause of bacteremia because of their high prevalence. In another
technology. study, among 81 episodes of coagulase-negative staphylococcal
blood culture results, the incidence of clinically significant
Identity of Organism bacteremia was 20 (24.7%) episodes, that of indeterminate
Often, the identity of the microbe that grows from a blood culture bacteremia was 10 (12.3%) episodes, and that of contamination
is a very helpful clue that the results may or may not represent was 59 (72.8%) episodes. Other studies found rates of true
contamination. The College of American Pathologists (CAP) Q- bacteremias ranging from 10% to 26.4% when coagulase-
Probes study described above found that the most important negative staphylococci are isolated from blood cultures.
indicator when interpreting blood culture results was the identity Similarly, other organisms can be difficult to interpret when
of the organism, which was cited as very important to 79% of isolated from blood cultures. One study found that enterococci
laboratories. were pathogens 70% of the time, whereas viridans group
streptococci were pathogens 38% of the time. Furthermore, that
Weinstein et al.'s study of 843 episodes of positive blood cultures same study found that Clostridium perfringens was a
in adult inpatients from three hospitals around the country contaminant 77% of the time, whereas other Clostridium species
suggested that certain organisms should almost always be were true pathogens 80% of the time. Given these data, clinicians
thought to represent true bacteremia or fungemia when isolated attempting to differentiate true infections from simply
from a blood culture. These organisms included Staphylococcus contaminated blood cultures cannot rely solely on the identity of
aureus, Streptococcus pneumoniae, Escherichia coli and other the organism. Often, bloodstream infections involve only a single
Enterobacteriaceae, Pseudomonas aeruginosa, and Candida organism, prompting clinicians to sometimes conclude that a

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blood culture bottle that grows multiple organisms is chances of contamination of air micro flora. This problem is
contaminated. However, studies have found that 6% to 21% of all addressed ready prepared BHI bottle by including a sterile cover
true bacteremias are polymicrobial, usually in patients in high- rim within a cap so that needle can be directly inserted into the
risk groups. Furthermore, multiple coagulase-negative species culture bottle through the small opening inside the flip of the cap
have been found to cause polyclonal coagulase-negative without opening the cap. Moreover the cap is externally covered
staphylococcal infections. Therefore, one cannot conclude that with the gamma irradiated plastic layer which should be opened
the mere presence of multiple organisms in a blood culture bottle at the time of adding the blood in the media and this minimizes the
always indicates contamination. chances of contamination through cap. Blood is withdrawn from
a patient using a sterile needle and syringe and is added to the
Recovery of blood isolates blood culture bottle containing 0.05% Sodium Polyanethol
The diversity of bacteria that is recovered from blood requires an Sulfonate (SPS).
equally diverse and large number of media to enhance the growth The bottle is incubated and observed for turbidity, colour change,
of these bacteria. Basic blood culture media contain a nutrient hemolysis, gas formation and other evidence of microbial
broth and an anticoagulant, and several different broth growth. 5 ml blood culture media for neonates, 20 ml blood
formulations are available but most blood culture bottle culture media for pediatric and 70 ml blood culture media is
commercially available contains trypticase soy broth, brain-heart sufficient for adults (Pic. 1).
infusion broth, glucose broth, thioglycollate broth or more
specialized broth include Columbia broth or Brucella broth. The growth of microorganisms in a blood culture may be delayed
Brain heart infusion broth is a highly nutritious general purpose or prevented if an anticoagulant is not used in the culture medium
liquid medium used for the cultivation of fastidious and non- since the organisms may become trapped in the fibrin clot. Blood
fastidious microorganism, including aerobic and anaerobic contains antibodies, complement, ß-lysin and phagocytes which
bacteria from a variety of clinical and non clinical specimens. are natural bacterial inhibitors. Antibiotics in the blood also may
This media contains calf brain, beef heart and peptone which greatly reduce, if not completely eliminate, the chances of
provide the major source of carbon, nitrogen and many growth obtaining a positive culture. These issues are overcome by the use
factors. Similarly dextrose is the fermentable carbohydrate of sodium polyanetholsulfonate (SPS). Van Hablerand Miles in
source and dipotassium phosphate acts as a buffer. 1938 first demonstrated the usefulness of SPS in blood culture
medium. SPS is a polyanionic anticoagulant, which inhibits
Conventionally media for blood culture is prepared in the glass complement and lysozyme activity, interferes with phagocytosis
vials and when blood is added to this culture bottle, its cap needs and inactivates aminoglycosides. SPS inhibits the activity of
to be completely open and if this procedure is not performed streptomycin, polymyxin B, kanamycin and gentamicin. SPS
maintaining the sterile condition this greatly increases the completely inhibited serum antibacterial activity and serum-

Pic. 1

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 Current Trends OCT-NOV 2013

dependent phagocytosis (and killing) by isolated leukocytes at a 13. Roth A, Wiklund AE, Palsson SA, Melander ZE, Wullt M,
concentration usually employed in blood culture media. SPS also Cronqvist MW and Sturegard E (2010). Reducing blood
stimulated both glucose C-1 oxidation in resting leukocytes and culture contamination by a simple informational
formate oxidation in both resting and phagocytosing leukocytes intervention, Journal of Clinical Microbiology 48(12): 4552-
in serum-free systems. These in vitro studies support the concept 4558.
that SPS is a useful additive to blood culture media and further
elaborate on the mechanism of its inhibition of the microbicidal 14. Rubin, L. G., P. J. Sanchez, J. Siegel, G. Levine, L. Saiman,
activity of blood. W. R. Jarvis, et al. 2002. Evaluation and treatment of
neonates with suspected late-onset sepsis: a survey of
In conclusion, unique cap design of the blood culture bottles to neonatologists' practices. Pediatrics 110:e42.
prevent from the contamination and incorporation of the liquid,
SPS at a concentration of 0.05% makes the ready prepared BHI as 15. Schifman, R. B., C. L. Strand, F. A. Meier, and P. J. Howanitz.
a better option for the blood culture. 1998. Blood culture contamination: a College of American
Pathologists Q-Probes study involving 640 institutions and
References: 497134 specimens from adult patients. Arch. Pathol. Lab.
1. Belding ME and Klebanoft SJ (1972). Effect of sodium Med. 122:216–221.
polynethol sulfonate on antimicrobial system in blood,.
Applied Microbiology 24 (5): 691-698. 16. Schifman, R. B., C. L. Strand, F. A. Meier, and P. J. Howanitz.
1998. Blood culture contamination: a College of American
2. Blood culture system: Data on File: Microxpresss. Pathologists Q-Probes study involving 640 institutions and
497134 specimens from adult patients. Arch. Pathol. Lab.
3. Calfee, D. P., and B. M. Farr. 2002. Comparison of four Med. 122:216–221.
antiseptic preparations for skin in the prevention of
contamination of percutaneously drawn blood cultures: a 17. Sharma, M., K. Riederer, L. B. Johnson, and R. Khatib. 2001.
randomized trial. J. Clin. Microbiol. 40:1660–1665. Molecular analysis of coagulase-negative Staphylococcus
isolates from blood cultures: prevalence of genotypic
4. Chandrasekar, P. H., and W. J. Brown. 1994. Clinical issues variation and polyclonal bacteremia. Clin. Infect. Dis.
of blood cultures. Arch. Intern. Med. 154:841–849. 33:1317–1323.

5. Evans, G.L., et al. (1967). Growth inhibition of 18. Souvenir, D., D. E. Anderson, Jr., S. Palpant, H. Mroch, S.
mycoplasmas by sodium polyanethol sulphonate. Askin, J. Anderson, J. Claridge, J. Eiland, C. Malone, M. W.
Antimicrobial agents and chemotherapy,1967: 687. Garrison, P. Watson, and D. M. Campbell. 1998. Blood
cultures positive for coagulase-negative staphylococci:
6. Forbes BA, Sahm DP and Weissfeild AS (2007). Bailey and antisepsis, pseudobacteremia, and therapy of patients. J.
Scotts Diagnostic Microbiology, 12th International edition, Clin. Microbiol. 36:1923–1926.
Mosby Elsevier, pp 778-794.
19. Tokars, J. I. 2004. Predictive value of blood cultures positive
7. Galdbart, J. O., A. Morvan, N. Desplaces, and N. el Solh. for coagulasenegative staphylococci: implications for
1999. Phenotypic and genomic variation among patient care and health care quality assurance. Clin. Infect.
Staphylococcus epidermidis strains infecting joint Dis. 39:333–341.
prostheses. J. Clin. Microbiol. 37:1306–1312.
20. Van Haebler T., A.A. Miles, 1938. The action of sodium
8. Herwaldt, L. A., M. Geiss, C. Kao, and M. A. Pfaller. 1996. polyanethol sulphonate (liquoid) on Blood Cultures. Journal
The positive predictive value of isolating coagulase-negative of pathology, 46: 245.
staphylococci from blood cultures. Clin. Infect. Dis.
22:14–20. 21. Van Eldere, J., W. E. Peetermans, M. Struelens, A. Deplano,
and H. Bobbaers. 2000. Polyclonal Staphylococcal
9. Huang, A. H., J. J. Yan, and J. J. Wu. 1998. Comparison of endocarditis caused by genetic variability. Clin. Infect. Dis.
five days versus seven days of incubation for detection of 31:24–30.
positive blood cultures by the Bactec 9240 system. Eur. J.
Clin. Microbiol. Infect. Dis. 17:637–641. 22. Van Wijngaerden, E., W. E. Peetermans, S. Van Lierde, and J.
Van Eldere. 1997. Polyclonal staphylococcus endocarditis.
10. Isenberg H (1998). Essential Procedures in Clinical Clin. Infect. Dis. 25:69–71.
Microbiology, ASM Press Washington D.C.
23. Weinstein, M. P. 2003. Blood culture contamination:
11. Jaslyn MM, Janice MB, Mary RG, Carl MU, Sorana SM persisting problems and partial progress. J. Clin. Microbiol.
(2013). Frequency of blood cultures performed in a 41:2275–2278.
community hospital. Open Journal of Medical
Microbiology, 3:130-134. 24. Weinstein, M. P., M. L. Towns, S. M. Quartey, S. Mirrett, L.
G. Reimer, G. Parmigiani, and L. B. Reller. 1997. The
12. Norberg, A., N. C. Christopher, M. L. Ramundo, J. R. Bower, clinical significance of positive blood cultures in the 1990s: a
and S. A. Berman. 2003. Contamination rates of blood prospective comprehensive evaluation of the microbiology,
cultures obtained by dedicated phlebotomy vs intravenous epidemiology, and outcome of bacteremia and fungemia in
catheter. JAMA 289:726–729. adults. Clin. Infect. Dis. 24:584–602.

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 OCT-NOV 2013 In Profile

Dr. A. P. J. Abdul Kalam Career as scientist

After graduation from Madras Institute of Technology (MIT –
Born 15 October 1931 (age Chennai) in 1960, Kalam joined Aeronautical Development
80) Establishment of DRDO as a chief scientist. There, Kalam started
Rameswaram, British his career by designing a small helicopter for the Indian Army, but
India (now Tamil remained unconvinced with the choice of his job. Kalam was also
Nadu, India) part of the INCOSPAR committee working under Vikram
Political party Independent Sarabhai. In 1969, Kalam was transferred to the Indian Space
Education St. Joseph's College, Research Organization (ISRO) where he was the project-director
Tiruchirappalli of India's first indigenous Satellite Launch Vehicle (SLV-III).
Madras Institute of Joining ISRO was one of Kalam's biggest achievements in life and
Technology he is said to have found himself when he started to work on the SLV
Occupation Professor (Aerospace engineering) project. However, Kalam first started work on an expandable
nuclear scientist rocket project independently at DRDO in 1965. In 1969, Kalam
rocket scientist received the government's approval and expanded the program to
author many engineers. In 1979, the first maiden flight of this project was
academic made and in 1980, country's first satellite Rohini was launched
Profession Aerospace engineer with this rocket.
Religion Islam From 1970s and 1990s, Kalam made an effort to develop the Polar
SLV and SLV-III project which proved to be successful. In 1974,
Avul Pakir Jainulabdeen Abdul Kalam born 15 October 1931 Kalam was shifted to the Terminal Ballistics Research Laboratory
usually referred to as A. P. J. Abdul Kalam, is a renowned (TBRL), a subsidiary of DRDO. Kalam was invited to witnessed
aerospace engineer, professor (of Aerospace engineering), and the country's first nuclear test, Smiling Buddha as the
first Chancellor of the Indian Institute of Space Science and representative of TBRL, even though he had not participated in
Technology Thiruvananthapuram (IIST), who served as the 11th developing or even taken part in the test site preparations or
President of India from 2002 to 2007. During his term as President, weapon designing. Kalam arrived at the test site on the invitation of
he was popularly known as the People's President. He was Raja Ramanna. In 1970s, a landmark was passed when ISRO first
awarded the Bharat Ratna, India's highest civilian honour in 1997. launched into space the locally built Rohini-1, using the SLV
Before his term as India's president, he worked as an aerospace rocket. In the 1970s, Kalam also directed the Project Devil and
engineer with Defence Research and Development Organisation Project Valiant to developed the ballistic missiles from the
(DRDO) and Indian Space Research Organisation (ISRO). He is technology of Kalam's successful SLV programme. Despite the
popularly known as the Missile Man of India for his work on the disapproval of Union Cabinet, Premier Indira Gandhi allotted
development of ballistic missile and space rocket technology. secret funds for these aerospace projects through her discretionary
Kalam played a pivotal organizational, technical and political role powers under Kalam's directorship. Kalam played an integral role
in India's Pokhran-II nuclear tests in 1998, the first since the convincing the Union Cabinet to conceal the true nature of these
original nuclear test by India in 1974. Kalam has even been circled classified aerospace projects.
with various controversies as many scientific experts called him a His research and educational leadership brought him great laurels
man with no authority over "nuclear physics" and a man who just and prestige in 1980s, which prompted the government to initiate
carried the works of Homi J. Bhabha and Vikram Sarabhai. an advanced missile program under his directorship. As Chief
He is currently a visiting professor at Indian Institute of Executive of the Integrated Guided Missile Development Program
Management Ahmedabad, Chancellor of Indian Institute of Space (I.G.M.D.P), he played a major part in developing many missiles in
Science and Technology Thiruvananthapuram, a professor of India including Agni and Prithvi although the entire project has
Aerospace Engineering at Anna University (Chennai), a visiting been criticised for being overrun and mismanaged. He was the
professor at Indian Institute of Management Indore, and an Chief Scientific Adviser to the Prime Minister and the Secretary of
adjunct/visiting faculty at many other academic and research Defence Research and Development Organisation from July 1992
institutions across India. to December 1999. The Pokhran-II nuclear tests were conducted
In May 2011, Kalam launched his mission for the youth of the during this period where he played an intensive political and
nation called the What Can I Give Movement. Kalam better known technological role. Kalam served as the Chief Project Coordinator,
as a scientist, also has special interest in the field of arts like writing along with R. Chidambaram during the testing phase. Photos and
Tamil poems, and also playing the music instrument Veenai. snapshots of him taken by the Media elevated Kalam as the
Early life and education country's top nuclear scientist although Kalam was not directly
Kalam spent most of his childhood running into financial involved with the nuclear program at the time.
problems and started working at an early age to supplement his Awards
family's income. Kalam was brought up in a multi-religious, He was honoured with a Padma Bhushan in 1981. He was awarded
tolerant society, with Kalam strictly following his religious the Bharat Ratna, India's highest civilian honour. Kalam was
routine. After completing school, Kalam along with his cousin awarded the Padma Vibhushan in 1990.
Samsuddin Kalam distributed papers in order to financially Personal life
contribute to his father's income. After completing his school He has enjoyed a bachelor life. He is the happiest at the drawing
education, Kalam went on to attend the Saint Joseph's College, board, in discussion with his scientists on how their dreams for the
Tiruchirappalli where he graduated in Physics in 1954 but towards next millennium can be fulfilled.
the end he was not enthusiastic about seeing himself as a physicist. References:
Kalam soon discovered aerospace engineering at Madras, and he www.abdulkalam.nic.in/profile
regretted the loss of the four years spent in physics. In 1955, Kalam www.abdulkalam.com
moved to Madras and began taking courses on Aerospace ulkalam.hpage.co.in
engineering.
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 Relaxed Mood OCT-NOV 2013

he h u m o u r
Enjoy t
A Lady to Doctor:
My husband has habit of talking in sleep! what should i give him
to cure?
Dr: Give him an Opportunity to speak when he is awake

Having "WIFE" Is A Part Of Living...

But Having "GIRLFRIEND" Along With The "WIFE" Is Art Of
Living.
A man dies. In heaven he sees a large Wall full of Clocks.
He asks angel:"What are these for?”
Angel answers: "These r Lie Clocks, every person has lie clock!
What is the Difference between Mother & Wife?
A - One Woman Brings U into this world crying... & the other
Whenever u lie on earth, clock moves."...
ensures U Continue to do so.
The man points towards a clock n asks:
Whose clock is this?
Angel says: its Mother Teresa 's. It never moved, showing that she Interviewer to Millionaire: To whom do you owe your
never told lie. success as a millionaire?”
The man asks: Millionaire: "I owe everything to my wife."
Where is Indian Politician's clock? Interviewer: "Wow, she must be some woman.
Angel replies: Interviewer: "What were you before you married her?"
That's in our office ... we use it as TABLE FAN...ha ha ha ! Millionaire: "A Billionaire"

An Angry Wife To Her Husband on Phone: "Where d Hell Wife: You always carry my photo in your handbag to the office.
Are You ...?" Why?
Husband: Darling You Remember That Jewellery Shop Where Darling: When there is a problem, no matter how impossible, I
You Saw The Diamond Necklace n Totally Fell In Love With It n I look at your picture and the problem disappears.
Didn't Have Money That Time n I said "Baby It'll Be Yours 1 Day Wife: You see, how miraculous and powerful I am for you?
... "O:) Darling: Yes, I see your picture and say to myself, "What other
Wife, With A Smile & Blushing: Yeah I Remember That My problem Can there be greater than this one?
Love!
Husband: I'm in the Pub Just Next To That Shop Wife: honey, what r u looking 4?
Husband: nothing
Husband was seriously ill. Doc to wife: Give him healthy Wife: why have u been reading our marriage certificate 4 an hour?
Husband: i was just looking 4 the expiry date
breakfast, be pleasant & in gud mood, don't discuss ur problems,
no tv serial, don't demand new clothes & gold jewels,
Do this for 1 yr & he will be ok. A man in Hell asked Devil:
On the way home.. Husband: what did the doc say ? Can I make a call to my Wife?
Wife:- .No chance for u to survive After making call he asked how much to pay.
Devil: Nothing, Hell to hell is Free.
Woman Buys A New Sim Card Puts It In Her Phone And
Decides To Surprise Her Husband Who Is Seated On The Couch A man came home late at night after a party.
In The Living Room. She Goes To The Kitchen, Calls Her His wife yelled:
Husband With The New Number: "Hello Darling" “how would you feel if you don't see me for two days?"
The Husband Responds In A Low Tone: "Let Me Call U Back The man couldn't believe his luck: 'that would be great'!
Later Honey, The Dumb Lady Is In The Kitchen.. Monday passed and he didn't see her......
Tuesday and Wednesday passed too.....
On Thursday his swelling became better
When a married man says "I'll think about it", And now he could see her from the corner of one eye.
What he really means that, He doesn't know his wife's opinion
yet..
Doctor: Madam, your husband needs rest and peace, so here
are some sleeping pills.
Wife: Doc, when should I give them to him?
Doctor: They are for you.!!

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 OCT-NOV 2013 Bug of the Month

Neisseria gonorrhoeae
Scientific Classification: industry is the ability to conjugate since this can lead to antibiotic
Kingdom: Bacteria resistance.
Phylum: Proteobacteria In 2011, researchers at Northwestern University found evidence
Class: Beta Proteobacteria of a human DNA fragment in a Neisseria gonorrhoeae genome,
Order: Neisseriales the first example of horizontal gene transfer from humans to a
Family: Neisseriaceae bacterial pathogen.
Genus: Neisseria
Species: N. gonorrhoeae
Disease:
Neisseria gonorrhoeae, also known as gonococci (plural), or Symptoms of infection with N. gonorrhoeae differ depending on
gonococcus (singular), is a species of Gram-negative coffee the site of infection. Note also that 10% of infected males and
bean-shaped diplococci bacteria responsible for the sexually 80% of infected females are asymptomatic.
transmitted infection gonorrhea. Infection of the genitals can result in a purulent (or pus-like)
discharge from the genitals which may be foul smelling.
Microbiology: Symptoms may include inflammation, redness, swelling, and
dysuria.
Neisseria are fastidious Gram-negative cocci that require nutrient
supplementation to grow in laboratory cultures. Specifically, they N. gonorrhoeae can also cause conjunctivitis, pharyngitis,
grow on chocolate agar with carbon dioxide. These cocci are proctitis or urethritis, prostatitis and orchitis.
facultatively intracellular and typically appear in pairs Disseminated N. gonorrhoeae infections can occur, resulting in
(diplococci), in the shape of coffee beans. Of the eleven species of endocarditis, meningitis or gonococcal dermatitis-arthritis
Neisseria that colonize humans, only two are pathogens. N. syndrome. Dermatitis-arthritis syndrome presents with
gonorrhoeae is the causative agent of gonorrhea (also called "The arthralgia, tenosynovitis and painless non-pruritic (non-itchy)
Clap," which is derived from the French word "clapier," meaning dermatitis.
"brothel") and is transmitted via sexual contact. Infection of the genitals in females with N. gonorrhoeae can
Neisseria is usually isolated on Thayer-Martin agar (or VPN result in pelvic inflammatory disease if left untreated, which can
agar) — an agar plate containing antibiotics (vancomycin, result in infertility. Pelvic inflammatory disease results if N.
colistin, nystatin, and TMP-SMX) and nutrients that facilitate the gonorrhoeae travels into the pelvic peritoneum (via the cervix,
growth of Neisseria species while inhibiting the growth of endometrium and fallopian tubes). Infertility is caused by
contaminating bacteria and fungi. Further testing to differentiate inflammation and scarring of the fallopian tube. Infertility is a
the species includes testing for oxidase (all clinically relevant risk to 10 to 20% of the females infected with N. gonorrhoeae.
Neisseria show a positive reaction) and the carbohydrates
maltose, sucrose, and glucose test in which N. gonorrhoeae will
Diagnostic Tests
only oxidize (that is, utilize) the glucose.
1) Microscopy
N. gonorrhoeae are motile (twitching motility) and possess type
IV pili to adhere to surfaces. The type IV pili operate A direct smear for Gram staining may be performed as soon as the
mechanistically similar to a grappling hook. Pili extend and swab specimen is collected from the urethra, cervix, vagina or
attach to a substrate which signals the pilus to retract, dragging rectum. The swab should be rolled gently onto the slide to
the cell forward. N. gonorrhoeae are able to pull 100,000 times preserve cellular morphology and over an area less than 1 cm2.
their own weight and it has been claimed that the pili used to do so Gram staining kits are commercially available. Under oil
are the strongest biological motor known to date, exerting one immersion (1000x magnification), urethral smears from
nanonewton. symptomatic males usually contain intracellular Gram-negative
N. gonorrhoeae has surface proteins called Opa proteins, which kidney-shaped diplococci in polymorphonuclear leukocytes, the
bind to receptors on immune cells. In so doing, N. gonorrhoeae is presence of which is required for the presumptive diagnosis of
able to prevent an immune response. The host is also unable to gonorrhea.
develop an immunological memory against N. gonorrhoeae – 2) Culture
which means that future reinfection is possible. N. gonorrhoeae The current preferred laboratory method for the diagnosis of N.
can also evade the immune system through a process called gonorrhoeae infections is the isolation and identification of the
antigenic variation, in which the N. gonorrhoeae bacterium is agent.
able to alter the antigenic determinants (sites where antibodies
bind) such as the Opa proteins and Type IV pili that adorn its
surface. The many permutations of surface proteins make it more Media and cultural conditions for isolation:
difficult for immune cells to recognize N. gonorrhoeae and The primary specimens should be inoculated onto nonselective
mount a defense. chocolate agar and selective agar containing antimicrobial agents
N. gonorrhoeae is naturally competent for DNA transformation that inhibit the growth of commensal bacteria and fungi. The
as well as being capable of conjugation. Both of these concepts antibacterial agents in modified Thayer-Martin, Martin Lewis
allow for the DNA of N. gonorrhoeae the ability to undergo and New York City medium are vancomycin, colistin,
conformational changes. Especially dangerous to the health trimethoprim lactate and the antifungal agents nystatin and

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 Bug of the Month OCT-NOV 2013

anisomycin or amphotericin B. Some fastidious strains, such as ability to ferment different sugars is traditionally determined
the arginine-, hypoxanthine- and uracil-requiring strains, are using conventional cystine trypticase agar medium (CTA)
more susceptible to the concentrations of vancomycin or containing 1% sugar. This method is based on fermentative
trimethoprim used in the selective media. Isolates that are species and is not sensitive enough to detect acid from oxidative
inhibited by supplements in selective media should be grown on species. In addition, differentiating N. gonorrhoeae and
media with lower concentrations of antibiotic. Isolates that are Neisseria cinerea is difficult using CTA sugars. As a result, this
atypical, such as vancomycin-susceptible strains, should be test is no longer preferred for the detection of acid production
forwarded to reference laboratories to confirm their from carbohydrates. This method has been replaced by more
identification. Therefore, a quality assessment program that rapid commercially available methods in some laboratories
periodically compares isolation rates on selective and because the preparation of the necessary media is tedious and
nonselective media is desirable. labour intensive, and quality control of the purity of sugars is
The inoculated plates should be incubated at 35°C to 37°C in a required. The rapid carbohydrate test is a nongrowth-dependent
moist atmosphere enriched with CO2 (3% to 7%). An 18 h to 24 h method for the detection of acid production from carbohydrates
culture should be used as the inoculum for additional tests. Plates by Neisseria species. The Rapid Identification Method-Neisseria
should not be incubated for longer than 48 h because most old (Remel Inc, USA) is a commercially available rapid acid
cultures would not survive storage conditions. Autolysis may production test that compares well with the conventional method,
occur during prolonged incubation, and growth from agar plates but may also not differentiate between N. gonorrhoeae and N.
becomes difficult to suspend in solutions. Isolates should be cinerea. The rapid nongrowth tests have a specificity of 99% to
subcultured at least once on nonselective medium after initial 100% and are more sensitive than CTA sugars. Some of the
isolation before being used in a diagnostic test that requires pure commercial tests include not only acid production tests, but also
culture or heavy inoculum. tests for other biochemical characteristics (such as enzyme
production), including DNase and nitrate reduction. All tests
must include appropriate control strains.
Presumptive identification of N gonorrhoeae:
The presumptive identification of N. gonorrhoeae rests on the Treatment and prevention:
isolation of an oxidase-positive, Gram-negative diplococcus
recovered from urogenital sites that grows on selective media. If N. gonorrhoeae is resistant to the penicillin family of
The Gram stain of a smear of urethral exudates or endocervical antibiotics, then ceftriaxone (a third-generation cephalosporin) is
secretions shows typical Gram-negative intracellular diplococci. often used. Sexual partners should also be notified and treated.
The oxidase test uses the tetramethyl derivative of the oxidase Antibiotic-resistant gonorrhea has been noted by
reagent (1% aqueous solution of N, N, N, N-tetramethyl-1, 4- epidemiologists; beginning in the 1940s gonorrhea was treated
phenylenediamine) that is commercially available with penicillin, but doses had to be continually increased in order
(BACTIDROP Oxidase, Remel Inc, USA) or prepared in-house. to remain effective, and by the '70s, penicillin- and tetracycline-
To perform the test, a drop of reagent is applied to filter paper or resistant gonorrhea emerged in the Pacific Basin. These resistant
the tip of a cotton swab. Culture is then applied to the filter paper strains then spread to Hawaii, California, the rest of the United
or cotton swab tip using a platinum or plastic loop, wooden States, and Europe. Fluoroquinolones were the next line of
applicator stick or toothpick. A dark-purple colour change within defense, but soon resistance to this antibiotic emerged as well.
10 s indicates a positive sample. The catalase test (3% hydrogen Since 2007, standard treatment has been third-generation
peroxide) or superoxol (30% hydrogen peroxide) are other rapid cephalosporins, such as ceftriaxone, which are considered to be
tests used in the presumptive identification of N. gonorrhoeae. A our “last line of defense.”
drop of the reagent is placed in the centre of a clean glass slide and Recently, a high-level ceftriaxone-resistant strain of gonorrhea,
the suspect colony is picked with a loop and emulsified in the called H041, was discovered in Japan. Lab tests found it to be
reagent. N. gonorrhoeae will produce a positive reaction with resistant to high concentrations of ceftriaxone, as well as most of
bubbling within 1 s to 2 s. Weak bubbling or bubbling after 3 s the other antibiotics tested. Within N. gonorrhoeae, there are
indicates a negative reaction. The reagents are tested daily against genes that confer resistance to every single antibiotic used to cure
reference oxidase-positive and -negative cultures to ensure gonorrhea, but thus far they do not coexist within a single
quality. gonococcus. Because of N. gonorrhoeae's high affinity for
horizontal gene transfer, however, antibiotic-resistant gonorrhea
Confirmatory identification tests: is seen as an emerging public health threat.
Several methods are available to confirm the identification of N. Patients should also be tested for other sexually transmitted
gonorrhoeae, including biochemical testing, serological testing, infections, especially Chlamydia infections, since co-infection is
colourimetric testing and nucleic acid methods. More than one frequent (up to 50% of cases). Antibacterial coverage is often
system may be required to confirm identification. included for Chlamydia because of this.
Transmission can be reduced by the usage of latex barriers, such
as condoms or dental dams, during intercourse, oral and anal sex,
Biochemical tests: and by limiting sexual partners.
N. gonorrhoeae can be differentiated from other Neisseria
species, Moraxella species, Kingella species and other References:
commensals based on its ability to grow on appropriate selective 1) www.ncbi.nlm.nih.gov
and nonselective media, produce acid from glucose, maltose, 2) www.cdc.gov/std/gonorrhea/treatment
lactose, sucrose and fructose, reduce nitrate, produce 3) www.who.int
polysaccharide from sucrose and exhibit DNase production. The

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 OCT-NOV 2013 Did You Know

Prebiotics Vs. Probiotics: What’s the Difference?

The reason for confusion is pretty obvious. Not only do the words age". And the bacteria can be killed by acid, such as found in the
differ by only one letter, but they target similar benefits: human stomach.
improving overall health by improving digestive health through 3. Prebiotics nourish the thousands of good bacterial species
nourishing a healthy colon. already living in the colon. Probiotics contain from one to a
few species of bacteria which are added to the colon.
Here is a summary of the differences: Your colon contains trillions of bacteria - more than all the 'human'
1. Prebiotics are a very special form of dietary fiber. Probiotics are cells in your body. More than all the stars in the milky way galaxy. It is
living bacteria intended to benefit colon health. a number possibly too big to really comprehend. In fact, it is likely
2. Prebiotic Fiber is not affected by heat, cold, acid or time. that each person's colon "microbiota" is as unique as fingerprint.
Probiotics can be killed by heat, acid or simply the passage of time. Prebiotics nourish all the good bacteria in the colon, particularly in the
3. Prebiotics nourish the thousands of good bacterial species already lacto- and bifido- families, preserving your "fingerprint".
living in the colon. Probiotics contain from one to a few species of Probiotics typically contain a few, or even just one, strain of bacteria.
bacteria which are added to the colon when they are ingested These bacteria are typically from the same bifido- and or lacto-
(eaten). families, but may or may not match your particular microbiota.
4. Prebiotic Fiber is a naturally-occurring substance, found in 4. Prebiotic Fiber is a naturally-occurring substance. Probiotics
thousands of plant species (though mostly in very small amounts). occur naturally in fermented foods like yogurt or sauerkraut.
Probiotics occur naturally in fermented foods like yogurt or Some companies have also engineered "proprietary" bacteria which
sauerkraut. they have patented and promote in their probiotic supplements/foods.
5. Prebiotics foster an environment in the colon which is hostile to Both prebiotics and the bacteria in probiotics are naturally-occurring
bad bacteria. Probiotics may impact bad bacteria by crowding substances. Prebiotics have been identified in over 36,000 plant
them out. species. The good bacteria contained in probiotic supplements
6. The benefits of prebiotics are supported by extensive research. The naturally occur in fermented foods such as Kefir and Sauerkraut
benefits of probiotics are supported by extensive research. (though they are almost certainly destroyed if those foods are
7. BOTH Prebiotics and Probiotics must be ingested in sufficient pasteurized.)
quantity to have an impact, and should not carry an excessive 5. Prebiotics foster an environment in the colon which is hostile
"load" of sugar, calories, carbs, etc. to bad bacteria. Probiotics may impact bad bacteria by
crowding them out.
Now, let's discuss these in a bit more detail: Prebiotics create an important secondary impact in the colon. By
1. Prebiotics are a very special form of dietary fiber. Probiotics nourishing the good bacteria, they induce these bacteria to create
are living bacteria supposedly beneficial to the colon. Short-Chain-Fatty-Acids. These SCFAs slightly lower the pH of the
Prebiotics are nondigestible substances that pass through the stomach colon, creating an environment that is friendly to good bacteria, but
and small intestine unchanged. inhospitable to bad bacteria.
Thus far only two fructooligosaccharides: oligofructose and inulin, Probiotics may also achieve this by crowding out 'bad' bacteria, but
fully meet the complete medical definition of "prebiotic". The may also suffer from 'drop in the bucket' scenario: while "X Billion"
compound created from merging these two prebiotics together is may sound like a lot of bacteria, adding them to "X Trillion" of
called Oligofructose-Enriched-Inulin and is considered a "full- existing bacteria likely means an addition of 0.1% or less... IF they all
spectrum" Prebiotic. survive!
Some foods presented as "prebiotics" in and of themselves simply 6. The benefits of probiotics and prebiotics are supported by
contain prebiotics. For example we often see honey presented as "a extensive research.
prebiotic," while it is more accurate to simply say that honey contains Prebiotics have been extensively researched since their identification
a small amount of prebiotics (as do many other foods). in 1995. This research has typically been done at research-oriented
Prebiotics enter the colon where they nourish beneficial bacteria. The universities, often in Europe where awareness of prebiotics is highest.
beneficial bacteria, typically within the hundreds of strains under the Probiotic benefits are also extensively researched. Also frequently in
Lactobacillus and Bifidobacter families, create many health benefits the food science department of leading universities.
through their action in the colon. 7. Both Prebiotics and Probiotics must be ingested in sufficient
Probiotics are supplements that contain living organisms: bacteria. quantity to have an impact, and should not carry an excessive
They typically contain anywhere from one to a few strains. Once "load" of sugar, calories, carbs, etc. out of proportion to their
consumed and in your system, the bacteria are just "bacteria" not benefit.
"probiotics." Many authorities suggest 4g to 8g of prebiotics daily. Some
However, for simplicity's sake, we will occasionally refer to the researchers suggest significantly higher servings daily for those with
bacteria themselves as "probiotic". active digestive disease - 15g daily or more.
2. Prebiotic Fiber is not greatly affected by heat, cold, acid or There is truly no authoritative guideline about effective levels of
time. Probiotics can be killed by heat, acid or simply the probiotics. Manufacturers suggest anywhere from 5 MIL CFUs to
passage of time. 500 BIL CFUs, with 50 BIL CFUs being the largest commonly-seen
Prebiotics benefit from their simplicity. They are, simply, very special serving size.
fibers that nourish the good bacteria. One area where BOTH prebiotics and probiotics are the same is that
Because of this, they are pretty impervious to damage. Heat does not they must not bring excess calories, carbs, sugar, fat or other
greatly harm them. They don't "die" just from the passage of time. undesirables to the dietary mix. Probiotics often come in heavily-
Acid does not harm or degrade them. sugared yogurts and similarly, prebiotics sometimes arrive via a
Probiotics by contrast are living organisms. If they are dead when "fiber bar" with chocolate icing, lots of sugar, etc.
they reach your colon, they cannot provide any health benefit. Anything that delivers 25% of your daily pro-/prebiotic need but 60%
Probiotics must not be subjected to excessive heat during transport of your daily sugar limit isn't a "good deal." Prebiotics and probiotics
and warehousing. They should typically be refrigerated to ensure the are both available in a supplement format with virtually no
bacteria remain relatively dormant and don't die simply from "old "overhead" of calories, sugar, etc.

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 Best Practices OCT-NOV 2013

Hand Hygiene Techniques

Pathogens transferred not only from infected or draining wound Five moments for hand hygiene–
but also from frequently colonized areas of normal, intact patient Moment 1- Before touching a patient
skin. The perineal or inguinal areas tend to be most heavily Hand hygiene at this moment will mainly prevent colonization of
colonized, but the axillae, trunk, and upper extremities (including the patient with health care-associated microorganisms, resulting
the hands) are also frequently colonized. from the transfer of organisms from the environment to the
The number of organisms such as S. aureus, Proteus mirabilis, patient through unclean hands, and exogenous infections in some
Klebsiella spp., and Acinetobacter spp. present on intact areas of cases.
the skin of some patients which can vary from 100 to 106 Moment 2- Before a clean/aseptic procedure
CFU/cm2. Nearly 106 skin squames containing viable Once within the patient zone, very frequently after a hand
microorganisms are shed daily from normal skin. Patient gowns, exposure to the patient's intact skin, clothes or other objects, the
bed linen, bedside furniture and other objects in the immediate HCW may engage in a clean/aseptic procedure on a critical site
environment of the patient become contaminated with patient with infectious risk for the patient, such as opening a venous
flora. Such contamination is most likely to be due to access line, giving an injection, or performing wound care.
staphylococci, enterococci or Clostridium difficile which are Importantly, hand hygiene required at this moment aims at
more resistant to desiccation. Certain Gram-negative rods, such preventing HCAI.
as Acinetobacter baumannii, can also play an important role in Moment 3- After body fluid exposure risk
environmental contamination due to their long-time survival After a care task associated with a risk to expose hands to body
capacities. This implies the potential importance of skin and fluids, e.g. after accessing a critical site with body fluid exposure
environmental contamination on microbial cross contamination risk or a critical site with combined infectious risk (body fluid
and pathogen spread. site), hand hygiene is required instantly and must take place
HCWs hands typically touch a continuous sequence of surfaces before any next hand-to-surface exposure, even within the same
and substances including inanimate objects, patients' intact or patient zone. This hand hygiene action has a double objective.
non-intact skin, mucous membranes, food, waste, body fluids, First and most importantly, it reduces the risk of colonization or
and the HCWs own body. With each hand-to-surface exposure, a infection of HCWs with infectious agents that may occur even
bidirectional exchange of microorganisms between hands and the without visible soiling. Second, it reduces the risk of a
touched object occurs and the transient hand-carried flora is thus transmission of microorganisms from a “colonized” to a “clean”
continually changing. In this manner, microorganisms can spread body site within the same patient.
throughout a healthcare environment and between patients within Moment 4- After touching a patient
a few hours. When leaving the patient zone after a care sequence, before
The burden of Health care associated infections is severe in touching an object in the area outside the patient zone and before a
immune compressed patients; prolonged hospital stay; invasive subsequent hand exposure to any surface in the health-care area,
devices and procedures such as catheter, ventilators; Surgical site hand hygiene minimizes the risk of dissemination to the health-
infections and also much more severe in high-risk populations care environment, substantially reduces contamination of HCWs'
such as adults housed in ICUs and neonates. hands with the flora from patient X, and protects the HCWs
Several studies have shown the ability of microorganisms to themselves.
survive on hands for differing times and inadequacy in hand Moment 5-After touching patient surroundings
hygiene leads to risk of cross-transmission of pathogens. The fifth moment for hand hygiene is a variant of Moment 4: it
Handwashing with soap and water has been considered a measure occurs after hand exposure to any surface in the patient zone, and
of personal hygiene for centuries and has been generally before a subsequent hand exposure to any surface in the health-
embedded in religious and cultural habits. Nevertheless, the link care area, but without touching the patient. This typically extends
between handwashing and the spread of disease was established to objects contaminated by the patient flora that are extracted
only two centuries ago, although this can be considered as from the patient zone to be decontaminated or discarded. Because
relatively early with respect to the discoveries of Pasteur and hand exposure to patient objects, but without physical contact
Lister that occurred decades later. with the patients, is associated with hand contamination, hand
hygiene is still required.
Indications for hand hygiene- Considering its importance and to empower hand hygiene in
A. Wash hands with soap and water when visibly dirty or visibly health care system, WHO has given guidelines on hand hygiene.
soiled with blood or other body fluids or after using the toilet. WHO guidelines on Hand hygiene -
B. If exposure to potential spore-forming pathogens is strongly A. Apply a palmful of alcohol-based handrub and cover all
suspected or proven, including outbreaks of Clostridium surfaces of the hands. Rub hands until dry.
difficile, hand washing with soap and water is the preferred B. When washing hands with soap and water, wet hands with
means. water and apply the amount of product necessary to cover all
C. In all clinical situations use an alcohol-based handrub as the surfaces. Rinse hands with water and dry thoroughly with a
preferred means for routine hand antisepsis, if hands are not single-use towel. Use clean, running water whenever
visibly soiled. If alcohol-based handrub is not obtainable, possible. Avoid using hot water, as repeated exposure to hot
wash hands with soap and water. water may increase the risk of dermatitis.
D. Before handling medication or preparing food perform hand Use towel to turn off tap/faucet. Dry hands thoroughly using a
hygiene using an alcohol-based handrub or wash hands with method that does not recontaminate hands. Make sure towels
either plain or antimicrobial soap and water. are not used multiple times or by multiple people.

Microxpress 14 Group
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 OCT-NOV 2013 Best Practices
water.
D. Brushes are not recommended for surgical hand Preparation.
Surgical hand antisepsis should be performed using either a
suitable antimicrobial soap or suitable alcohol-based
handrub, preferably with a product ensuring sustained
activity, before donning sterile gloves.
F. If quality of water is not assured in the operating theatre;
surgical hand antisepsis using an alcohol-based handrub is
recommended before donning sterile gloves when
performing surgical procedures.
G. When performing surgical hand antisepsis using an
antimicrobial soap, scrub hands and forearms for the length of
time recommended by the manufacturer, typically 2–5
minutes. Long scrub times (e.g. 10 minutes) are not necessary.
H. When using an alcohol-based surgical handrub product with
sustained activity, follow the manufacturer's instructions for
application times. Apply the product to dry hands only. Do not
combine surgical hand scrub and surgical handrub with
alcohol-based products sequentially.
I. When using an alcohol-based handrub, use sufficient product
to keep hands and forearms wet with the handrub throughout
the surgical hand preparation procedure.
J. After application of the alcohol-based handrub as
recommended, allow hands and forearms to dry thoroughly
before donning sterile gloves.
Other aspects of hand hygiene
A. Do not wear artificial fingernails or extenders when having
direct contact with patients.
B. Keep natural nails short (tips less than 0.5 cm long or
approximately ¼ inch)
These guidelines will help to achieve the goal of infection control
in health-care system.
Selection and handling of hand hygiene agents
A. Provide HCWs with efficacious hand hygiene products that
have low irritancy potential.
B. To maximize acceptance of hand hygiene products by HCWs,
solicit their input regarding the skin tolerance, feel, and
fragrance of any products under consideration.
C. When selecting hand hygiene products:
1. Determine any known interaction between products used to
clean hands, skin care products, and the types of glove used in
the institution.
2. Solicit information from manufacturers about the risk of
product contamination.
3. Ensure that dispensers are accessible at the point of care.
4. Ensure that dispensers function adequately and reliably and
deliver an appropriate volume of the product.
5. Ensure that the dispenser system for alcohol-based handrubs
is approved for flammable materials;
6. Solicit and evaluate information from manufacturers
regarding any effect that hand lotions, creams, or alcohol-
based handrubs may have on the effects of antimicrobial
soaps being used in the institution
7. Sost comparisons should only be made for products that meet
requirements for efficacy, skin tolerance, and acceptability
8. Do not add soap or alcohol-based formulations to a partially
empty soap dispenser. If soap dispensers are reused, follow
Recommendations for surgical hand preparation
recommended procedures for cleansing.
A. Remove rings, wrist-watch, and bracelets before beginning WHO recommend alcohol-based handrubs which are the only known
surgical hand preparation. Artificial nails are prohibited. means for rapidly and effectively inactivating a wide array of potentially
B. Sinks should be designed to reduce the risk of splashes. harmful microorganism, at present.
C. If hands are visibly soiled, wash hands with plain soap before WHO -World Health Organization
surgical hand preparation. Remove debris from underneath HCW - Healthcare Worker
fingernails using a nail cleaner, preferably under running HCAI - Healthcare Associated Infections

Microxpress 15 Group
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 In Focus OCT-NOV 2013

Microxpress BioShields Presents BIOSPRAY™

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Printed and published by D.G. Tripathi, Edited by Akshaya Tendulkar for and on behalf of Tulip
Diagnostics (P) Ltd., Tulip House, Dr. Antonio Do Rego Bagh, Alto Santacruz, Bambolim Complex, Microxpress
Post Office Goa - 403 202, India. Fax: 0832 2458544, Website: www.tulipgroup.com. Quick Reliable Microbiology

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