ABSTRACT
A two-step membrane filter procedure was evaluated to determine the ability to differentiate Escherichia coli from other coliform bacteria recovered from water. M-Endo LES agar incubated at 35 degrees C for 24 +/- 2 h was used as the initial isolation medium. Membranes containing coliform colonies were transferred to nutrient agar plus 4-methylumbelliferyl beta-D-glucuronide (MUG) and incubated for an additional 4 h at 35 degrees C. Escherichia coli colonies were distinguished by fluorescence when viewed under a long-wavelength ultraviolet light. A total of 119 MUG-positive colonies were isolated from 15 water sources, of which 115 (96.6%) were identified as E. coli. An examination of 182 pure culture environmental E. coli isolates revealed that 167 isolates (91.8%) exhibited fluorescence on the nutrient agar plus MUG medium. Survivors of E. coli cultures exposed to chlorination were also capable of producing a positive MUG reaction.
Subject(s)
Bacterial Typing Techniques , Enterobacteriaceae/classification , Escherichia coli/classification , Water Microbiology , Chlorine , Colony Count, Microbial , Culture Media , Disinfection , Enterobacteriaceae/growth & development , Escherichia coli/growth & development , Fluorescence , Fresh Water , Hymecromone/analogs & derivatives , Hymecromone/metabolism , Micropore FiltersABSTRACT
The new United States Drinking Water Regulations state that water systems must analyze for Escherichia coli or fecal coliforms on any routine or repeat sample that is positive for total coliforms. The proposed methods for the detection of E. coli are based on beta-glucuronidase activity, using the fluorogenic substrate 4-methylumbelliferyl beta-D-glucuronide (MUG). This study was conducted to determine whether beta-glucuronidase negative E. coli were present in significant numbers in environmental waters. Two hundred and forty E. coli cultures were isolated from 12 water samples collected from different environmental sources. beta-glucuronidase activity was determined using lauryl tryptose broth with MUG, EC broth with MUG, and the Autoanalysis Colilert (AC) procedure. The isolates were also evaluated by the standard EC broth gas fermentation method for fecal coliforms. The results confirm that assaying for the enzyme beta-glucuronidase utilizing the MUG substrate is an accurate method for the detection of E. coli in environmental waters.
Subject(s)
Escherichia coli/enzymology , Escherichia coli/isolation & purification , Glucuronidase/analysis , Water Microbiology , Enterobacteriaceae/isolation & purification , Evaluation Studies as Topic , Sensitivity and Specificity , United States , United States Environmental Protection Agency/standards , Water Microbiology/standards , Water Supply/standardsABSTRACT
The Autoanalysis Colilert (AC) test was compared with the membrane filter (MF), 10-tube multiple-tube fermentation (MTF) technique, and the presence-absence test as described in Standard Methods for the Examination of Water and Wastewater for the detection and enumeration of total coliforms in water. The methods were evaluated with 31 samples from seven different sources. Each sample was analyzed by each of the techniques, using replicate 100-ml sample volumes. A total of 582 confirmed tubes were positive by the MTF test, and 533 tubes were positive by the AC test. Statistical analysis of the most-probable-number comparability data showed a statistically significant difference in the number of positive tubes, with the MTF test resulting in more positive tubes. There were no statistically significant differences in precision between the two methods. All the methods were comparable in detection of total coliforms. Levels of heterotrophic bacteria generally encountered in drinking water did not interfere with detection or enumeration of coliforms by the AC test.
