Bacteriological Examination of Water 1658377329
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wertyyyBacteriological Examination of Water 1658377329
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wertyyyBacteriological
Examination of Water
Sedky Hassan Aly Hassan
Botany Dept.
Faculty of Science
Assiut University
71516, Assiut
Egypt
Water examination
Physical examination
Chemical examination
Microbiological examination
Each gram of human feces contains
approximately 10 billion of pathogenic
bacteria, such as Salmonella, associated with
gastroenteritis.
In addition, feces may contain pathogenic
viruses, protozoa and parasites, these
organisms could cause disease. When testing
drinking water for contamination,
Health authorities therefore use the presence
of more easily detected fecal bacteria as
indicators of the presence of fecal
contamination
Indicator Bacteria
Group of microorganisms use to reflect the
quality and safety of water.
◼ Coliform and Fecal coliform bacteria
◼ Enterococcus and Streptococcus faecalis
◼ Pseudomonas aeruginosa
◼ Staphylococcus spp
◼ Clostridium
◼ Other bacteria (Salmonella & Shigella)
Criteria for selecting an indicator of
fecal contamination in water
(1) They are normally not present in water or soil
(2) They are relatively easy to identify,
(3) They survive a little longer in water than enteric
pathogens
(4) Must be associated with feces
(5) Test should be less complex than test for pathogen
◼ The presences of these microorganisms in
water implies that water may contain
pathogen or un safe for drinking
◼ The presence E. coli, in water and often
Enterococci, is direct evidence of fecal
contamination from warm-blooded organisms.
◼ Their presence indicates the possible
presence of pathogens
◼ A few strains of E. coli are pathogenic, such
as E. coli O157:H7, but most strains are not.
Total Coliforms
The total coliform group
Belongs to the family Enterobacteriaceae and includes
the aerobic and facultative anaerobic, gram-negative,
non-spore-forming, rod-shaped bacteria that ferment
lactose with gas production within 48 hours at 35oC
(APHA, 1998).
Total coliforms include Escherichia coli, Enterobacter,
Klebsiella, and Citrobacter.
Fecal coliforms
◼ Fecal coliforms are thermotolerant bacteria that
include all coliforms that can ferment lactose at
44.5 oC.
◼ The fecal coliform group comprises bacteria such
as Escherichia coli or Klebsiella pneumonae.
◼ The presence of fecal coliforms indicates the
presence of fecal material from warm-blooded.
Examination test
◼ Qualitative test
◼ Determination the presence or absence of
coliforms, By Most Probable Number (MPN)
of coliforms present in 100 ml of water
Examination test
◼ Presumptive Test
◼ Confirmed Test
◼ Completed Test
Presumptive Test
In the presumptive test a series of 9 or 15 tubes of
Medium are inoculated with measured amounts of
water to see if the water contains any lactose-
fermenting bacteria that produce gas.
Media used
◼ E. coli broth
◼ Lactose broth
◼ MacConky broth
◼ Brilliant Green Bile Broth
Presumptive test
DS or 2x
1X 0.1 X
A
B
Lactose
broth BGBB
Confirmatory test
E. coli on EMB
Enterobacter aerogenes Growing on EMB,
colonies are less darksurrounded by a wide, light
mucoid
Klebsiella which ferment lactose but accumulate
less acid, with dark colonies
Complete test
Fecal Coliforms that produce acid and gas from
lactose at 44.5± 0.2°C within 24±2h, also known
as faecal coliforms due to their role as faecal
indicators.
Some Rapid Methods for Coliform Detection
Enzymatic assays provide an alternative approach
for rapid and sensitive detection of total coliforms
In most tests, the detection of total coliforms is based
on the ß-galactosidase activity.
The enzyme substrates used are chromogenic
substrates such as ONPG (o-nitrophenyl-b-D-
galactopyranoside), CPRG (chlorophenol red-b-D
galactopyranoside), X-GAL (5-bromo-4-chloro-3-
indolyl-b-D-galactopyranoside).
◼ Fluorogenic substrates are also used and
include 4-methylumbelliferone-D-galactoside
(MUGA) or fluorescein-di—D-
galactopyranoside (FDG)
Faecal streptococci (FS):
Gram-positive, catalase-negative cocci from
selective media
(e.g. azide dextrose broth or m Enterococcus agar)
◼ S. faecalis
◼ S. faecium
◼ S. durans
◼ S. bovis
◼ S.equinus
Streptococcus faecalis
Fecal streptococci
Azide Dextrose Broth
Approximate Formula* Per Liter
Beef Extract ............................................. 4.5 g
Pancreatic Digest of Casein ................... 7.5 g
Proteose Peptone No. 3 ..........................7.5 g
Dextrose ................................................. 7.5 g
Sodium Chloride ..................................... 7.5 g
Sodium Azide .. ...................................... 0.2 g
Bromo cresol purple ............... ............ 0.032g
Presumptive test
Staphylococcs aureus
Media Used
◼ TSA with Pyruvic acid and NaCl
◼ MSA Mannitol Salt Agar
◼ BAIRD-PARKER AGAR BASE
Mannitol Salt Agar
◼ Proteose Peptone ...................................... 10.0 g
◼ Beef Extract...... ....................................... 1.0 g
◼ D-Mannitol ................................................ 10.0 g
◼ Sodium Chloride ....................................... 75.0 g
◼ Agar ........................................................... 15.0 g
◼ Phenol Red .............................................. 25.0 mg
Staphylococcs aureus on MSA
Staphylococcs spp on MSA
Confirmed test
Complete test
S. epidermises
Staphylococcs
aureus S. saprophitics
Salmonella spp.
◼ belong to the family Enterobacteriaceae.
They are motile, Gram negative bacilli that do
not ferment lactose, but most produce
hydrogen sulfide or gas from carbohydrate
fermentation.
Media Used
◼ XLD
◼ SS Agar
◼ Selenite F Broth
◼ Bismuth Sulfite Agar
Salmonella on XLD Agar
Salmonella and Shigella on SS Agar
Clostridium perfringens
◼ Clostridium perfringens is an obligate anaerobic
enteric bacterium.
◼ Its spores are generally more tolerant to
environmental effects than other traditional
entericindicators.
◼ C. perfringens has been suggested as an
alternative bacterial indicator of fecal pollution
because it is primarily associated with human
wastes, and it is widely distributed in feces,
sewage, and polluted waters.
Media Used
◼ Fluid Thioglycollate Medium
◼ Typical Composition (g/litre)
Peptone from casein 15.0; yeast extract 5.0;
D(+) glucose 5.5; L-cystine 0.5; sodium
chloride 2.5; sodium thioglycollate 0.5;
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