Updated May 2005
The mENDO and NA-MUG (nutrient agar with MUG) method is a two-step membrane-filtration method for detection of total coliforms and Escherichia coli (E. coli). This method can be used in the field or the laboratory.
THEORY: The mENDO plates are incubated at 35°C for 22-24 hours. The mENDO agar contains selective and differential agents. Sodium lauryl sulfate and sodium desoxycholate inhibit Gram-positive cocci and endospore-forming bacteria. Basic fuchsin and sodium sulfite form a green metallic sheen on colonies able to ferment lactose to acetaldehyde—these colonies are counted as total coliforms. The membrane from the mENDO medium is transferred to NA-MUG medium and incubated for 4 hours at 35°C. Production of a bright blue fluorescence around the sheen colonies (a halo) indicates E. coli. The fluoresence is caused by the breakdown of a fluorogen, 4-methylumbelliferyl-b-D-glucuronide (MUG), which is cleaved by an enzyme specific to E. coli, b-glucoronidase. Fluorescent halo colonies are observed with a long-wave (366 nm) ultraviolet lamp.
USE: The mENDO method is suitable for use with drinking water. This method is also suitable for enumerating bacteria in ground water because in ground-water studies, it is often desirable to enumerate both total coliforms and E. coli. The method is not recommended for use with raw surface waters because non-target background growth may make plates difficult to read.
MEDIA: The mENDO agar dehydrated medium is available from Fisher Scientific (800/766-7000, Cat DF0736154). Ethanol (95% denatured) needs to be purchased also (Fisher Scientific, Cat A405P-4). NA-MUG may be purchased from a commercial supplier (Difco, Detroit, Mich., item number 0023-15-6). Instructions for media preparation can be found in Myers and Wilde (2003) page FIB-53.
Prepoured plates can be purchased from Hardy Diagnostics (800/266-2222, Cat G128). If prepoured plates are purchased, 95% ethanol is not needed.
Use phosphate buffered dilution water and 0.65 mm
membrane filters. Buffer can be
purchased from Hardy Diagnostics (800/346-2766, Cat D699 (99mL) or Cat U193
(500mL)). See
buffer preparation (Appendix M).
REFERENCES:
Britton, L.J., and Greeson, P.E., eds., 1989, Methods for collection and analysis of aquatic biological and microbiological samples: U.S. Geological Survey Techniques of Water-Resources Investigations, book 5, chap. A4, p. 13-16.
Myers, D.N., and Wilde, F.D., eds., 2003, National field manual for the collection of water-quality data—Biological indicators: U.S. Geological Survey Techniques of Water-Resources Investigations, book 9, chap. A7, 158 p.
U.S. Environmental Protection Agency, 1991, Test methods for Escherichia coli in drinking water—Test Method 1105: Cincinnati, EPA/600/4-91/016, 2 p.
NWIS PARAMETER CODES:
31501 Total coliforms on mENDO agar, colonies per 100 mL
50278, E. coli on NA-MUG agar, colonies per 100 mL
Last update: May, 2005