Presence Absence Broth

Presence - Absence Broth

EMD Cat. No. 1.00414.0500

Selective culture medium for the detection of coliform bacteria in water.

The culture medium conforms with the recommendations of standard methods ( US-EPA ) for the examination of wat

Typical Composition (g/liter)	Quality Control Literature
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Mode of Action
Peptones and meat extract give the nutrients and trace elements required for growth whereas the phosphate buffer and sodium chloride provide a good buffering capacity and the osmotic equilibrium.
Lactose-fermenting organisms form acid which is identified by the pH indicator bromocresol purple as a color change from purple to yellow.
The selective component of the culture medium is sodium lauryl sulphate which largely inhibits the undesired accompanying flora - with the exception of the coliforms.

Preparation
For the preparation of the triple strength concentrated broth, completely dissolve 91.5 g in 1 liter of demin water. Fill 50 ml quantities in 250 ml milk dilution bottles with screw caps and autoclave for 12 min. at 121 °C. Allow broth to cool to room temperature.
The pH of the single-strength broth: 6.8 ± 0.2 at 25 °C.
The prepared medium is clear to slightly opalescent and purple-red.

Add 100 ml water samples to the milk dilution bottles filled with 50 ml of triple strength concentrated broth, mix well. Aerobic incubation with a loose screw-cap for up to 48 hours at 35 ± 0.5 °C. Read results after 24 h and 48 h.

Experimental Procedure
Lactose-positive organisms form acid due to the fermentation of the lactose which colors the broth yellow. Gas formation can occur. To identify gas formation, the bottles are shaken gently and inspected to see if a foaming reaction occurs.
All samples with acid and/or acid and gas formation are presumptive-positive and are inoculated for confirmation in Brilliant-green Bile Lactose Broth (BRILA).
If gas formation occurs during the incubation of 48 ± 3 hours at 35 ± 0.5 °C, this can be seen as confirmation of the presence of coliforms in the 100 ml water sample.

Quality control
Test strains	Growth	Yellow color	Gas formation in BRILA Broth

Escherichia coli ATCC 25922	Good	Yellow	+
Escherichia coli ATCC 11755	Good	Yellow	+
Enterococcus faecalis ATCC 29212	Medium	Weakly yellow / yellow	-
Pseudomonas aeruginosa ATCC 27853	Poor / medium	None	-

Typical Composition (g/liter)
(single strength)
Meat extract 3.0; peptones 5.0; lactose 7.46; tryptose 9.83; dipotassium hydrogenphosphate 1.35; potassium dihydrogenphosphate 1.35; sodium chloride 2.46; sodium lauryl sulphate 0.05; bromocresol purple 0.0085.

Literature

Federal Register. 1989. National primary drinking water regulations; total coliforms ( including fecal coliforms and e. coli). Fed. Regist. 54; 27544-27568.
Weiss J.E. and Hunter C.A. 1939 J. Am. Water Works Ass. 31:707 713.
Eaton, A. D., Clesceri L. S. and Greenberg A. E. ( ed.). 1995. Standard methods for the examination of water and wastewater, 19th. Ed. Am. Public Health Ass. Washington D.C.
Clark, J. A. 1968. A presence absence (P-A) test, providing sensitive and inexpensive detection of coliforms and faecal streptococci in municipal drinking water supplies. Can. J. Microb. 14: 13-18.
Clark, J. A. 1969 The detection of various bacteria indicative of water pollution by a presence-absence (P-A) procedure. Can. J. Microbiol. 15: 771 780.
Clark, J. A. and Flassov L. T. 1973 Relationships among pollution indicator bacteria isolated from raw water and distribution systems by the presence-absence (P-A) test. Health. Lab. Sci. 10:163 172.
Clark, J. A. and Pagel J. E. 1977 Pollution indicator bacteria associated with municipal raw and drinking water supplies. Can. J. Micribiol. 23: 465-470.
Clark, J. A., Burger C. A. and Sabatinos L. E. 1982 Characterization of indicator bacteria in municipal raw water, drinking water and new main water samples. Can. J. Microbiol. 28: 1002-1013.
Jacobs, Leigler, Reed, Stukel and Rice. 1986 Appl. Environ. Microbiol. 51: 1007