Escherichia coli is a member of faecal coliform group of bacteria. It is a member of the indigenous faecal flora of warmblooded animals. E.coli is considered a specific indicator of faecal contamination and the possible presence of enteric pathogens. EC Broth was devised by Hajna and Perry (1) and further modified by addition of the fluorogenic compound MUG. MUG EC Broth is also recommended by APHA for the analysis of drinking water, surface and ground water and waste-water for the presence of E.coli (2). MUG permits rapid detection of E. coli when medium is observed for fluorescence using UV Light (3, 4). MUG also detects anaerogenic strains which may not be detected in conventional procedure (3). MUG is hydrolyzed by the enzyme beta-glucuronidase possessed by E.coli to yield a fluorescent end product 4-Methylumbelliferone.
Casein enzymic hydrolysate provides essential nutrients. Lactose is the fermentable carbohydrate. Sodium chloride maintains osmotic equilibrium. The medium has a strong buffering system to control the pH in the presence of fermentative action. The bile salts inhibit gram-positive bacteria especially Bacillus species and faecal Streptococci. Mostly beta-glucuronidase activity occurs within 4 hours but some weak beta- glucuronidase-positive strains require overnight incubation (2). The fermentation of lactose by lactose fermentors leads to acidification of the medium, resulting in drop of pH. Adjustment of pH of cultures by sodium hydroxide enhanced fluorescence as observed by Maddocks and Greenman (5).Similarly Freir and Hartman (6) noticed that exposure of tubes to ammonia fumes enhanced fluorescence.
Large number of Proteus vulgaris if present, may suppress gas production of E.coli, however fluorescence permits detection of E.coli in pure or mixed cultures within 4 to 24 hours.
Inoculate the test water sample into PA Broth (M1186) and Lauryl Sulphate Broth (M080). After an incubation at 35-37°C for 18-24 hours, all presumptive tubes showing growth, gas or acidity is further tested using MUG EC Broth (M1042). After an incubation at 35-37°C for 4-24 hours, the presence of a bright blue fluorescence is considered as a positive response for E. coli.
Storage and Shelf-life:
Store below 30°C in tighty closed container and prepared medium below 2-8°C. Use before expiry period on the label.
References:
1. Hajna A. A. and Perry C. A., 1943, Am. J. Public Health, 33:550.
2. Feng P. C. S. and Hartman P. A. S., 1982, Appl. Environ. Microbiol., 43:132. 3. Robinson B. J., 1984, Appl. Environ. Microbiol., 48:285.
4. Greenberg A. E., Trussell R. R. and Clesceri L. S., (Eds.), 1988, Standard Methods for the Examination of Water and Wastewater, 20th Ed., APHA, Washington, D.C.
5. Maddocks J. L. and Greenan M. J. (1975) J. Clin. Pathol. 28. 686-687.
6. Freir T. A. and Hartman P. A. (1987) Appl. Env. Microbiol. 53. 1246-1250.