MINIATURIZED FLUOROGENIC ASSAYS FOR ENUMERATION OF ESCHERICHIA-COLI AND ENTEROCOCCI IN MARINE WATER

To detect E. coli, 4-methylumbelliferyl beta-D-glucuronide (MUG) was incorporated into a modified Al-broth. To detect enterococci, 4-methylumbelliferyl beta-D-glucoside (MUD) was incorporated into a selective medium. To each well of a sterile 96-well microtitration plate, 100-mu-l of media was added, air dried, covered with sterile tape and stored at 4-degrees-C. To enumerate the indicator bacteria using this method, 200-mu-l of diluted or undiluted water was added to wells with MUG and MUD media. The plates were incubated for 36-40 hours at 44-degrees-C and observed in the dark for fluorescence by UV light (366nm). Fifty subsurface marine waters from four areas were examined. The recovery rate of the flurogenic assays is equal or superior to 3-tubes MPN and membrane filtration. The microtitration plate with MUG is more specific for E. coli than the membrane filtration. A 87.4% confirmation rate was observed with the flurogenic assay compared to a 31.7% false-positive rate with membranes. For the enterococci, the analysis of 23 English Channel and North Sea samples indicated that the fluorogenic assay and the standard methods had the same specificity ( < 5% of false-positive on 107 strains identified). In contrast, the microtitration plate with MUD showed, by far, the best specificity on 13 Mediterranean samples. A 100% confirmation rate was observed with the fluorogenic assay while 44% and 63% false-positive rates were observed with 3 tubes MPN and MF respectively.