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Summary of Presentation
Background information
Total coliform group Fecal coliform group Escherichia coli (E.coli) Other pathogenic bacteria
Test methods
Multiple-tube fermentation technique
Coliform Group
Consists of several genera of bacteria of fecal and non-fecal origin Definition based on the method of detection Ferment lactose with gas and acid formation within 48 h at 35oC Produce dark red colony with a metallic sheen within 24 h on an Endo-type medium containing lactose Considered an indicator organism A criterion of the degree of pollution and thus of sanitary quality
RLC LABORATORY Training-Workshop in Wastewater Analysis 2012 3
Coliform Group
Belongs to the family of Enterobacteriaceae Facultative anaerobic (do not have to have oxygen) Gram-negative Non-spore forming Rod-shaped Includes four genera that have at least some species of fecal origin Posses the enzyme -galactosidase (nickname beta-gal) that catalyzes the hydrolysis of galactosides into monosaccharides
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Escherichia coli
E.coli is a sub-group of fecal coliform group It is naturally found in the intestines of warmblooded animals, such as cows, chicken, pigs, dogs, cats, birds, and people; comprise of up to 1% of bacterial biomass Enters the environment through feces.
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Fecal Streptococci
Found in gastrointestinal tract of warm-blooded animals Enterococci are a subgroup of FS
Valuable indicator of fecal contamination of recreational waters
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Sample Collection
Use appropriate sampling container (sterile) with Na2S2O3 to remove chlorine Type: Grab (not composite) Use gloves while sampling Sample should be representative of flow After collection place it on ice or in a refrigerator Deliver to a laboratory ASAP, but no later than 6 hours of collection Maintain records
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Testing Method
Multiple-tube Fermentation Technique (MTFT) In general, this method is time consuming Applicable to wastewater, drinking water and surface water.
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Media
Lauryl Tryptose Broth (LTB) Brilliant Green Bile Broth (BGB) EC Mug
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MTFT (cont)
Set up 5 tubes with appropriate presumptive media for each of three sample dilutions 5 x 10 mL 5 x 1 mL 5 x 0.1 mL Add a sterile water control Incubate Swirl each tube gently and examine growth
RLC LABORATORY Training-Workshop in Wastewater Analysis 2012
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MTFT (cont)
Look for gas and acidic reaction Re-incubate for additional 24 h if no gas or acidic reaction is evident Using sterile 3-mm loop or wooden applicator transfer growth from a positive presumptive medium to a tube with confirmation medium Incubate Examine growth, look for gas Incubation in both culture mediums can be done simultaneously Calculate results using SMEWW Table 9221.IV.
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Use brilliant green lactose bile broth (BGBB) for confirmed phase Incubation
35 0.5oC 24 2 hours
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Positive
E.coli E.aerogenes
E.coli
Negative
Staphylococcus aureus Pseudomonas sp.
Enterobacter aerogenes
Fecal Coliform
Escherichia Coli
Fecal Streptococci Enterococci
E.coli
Enterococcus faecalis Streptococcus faecalis
Enterobacter aerogenes
Staphylococcus aureus S.mitis/salivarius
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References
SMEWW 21st Edition. 2005. APHA, AWWA, WEF
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