Multiparametric comparison of chromogenic-based culture methods used to assess the microbiological quality of drinking water and the mFC method combined with a molecular confirmation procedure.

MI agar and Colilert(®), as well as mFC agar combined with an Escherichia coli-specific molecular assay (mFC + E. coli rtPCR), were compared in terms of their sensitivity, ease of use, time to result and affordability. The three methods yielded a positive E. coli signal for 11.5, 10.8, and 11.5% of the 968 well water samples tested, respectively. One hundred and thirty-six (136) samples gave blue colonies on mFC agar and required confirmation. E. coli-specific rtPCR showed false-positive results in 23.5% (32/136) of cases. In terms of ease of use, Colilert was the simplest method to use while the MI method provided ease of use comparable to all membrane filtration methods. However, the mFC + E. coli rtPCR assay required highly trained employees for confirmation purposes. In terms of affordability, and considering contamination rate of well water samples tested, the Colilert method and the mFC + E. coli rtPCR assay were at least five times more costly than the MI agar method. Overall, compared with the other two methods tested, the MI agar method offers the most advantages to assess drinking water quality.

Comparative analysis of classical and molecular microbiology methods for the detection of Escherichia coli and Enterococcus spp. in well water.

The microbiological quality of 165 1 litre well water samples collected in the Québec City region was assessed by culture-based methods (mFC agar, Chromocult coliform agar, Colilert(®), MI agar, Chromocult enterococci, Enterolert™, and mEI agar) and by a molecular microbiology strategy, dubbed CRENAME-rtPCR, developed for the detection of Escherichia coli, Enterococcus spp., Enterococcus faecalis/faecium, and Bacillus atrophaeus subsp. globigii. In these drinking water samples, approved culture-based methods detected E. coli at rates varying from 1.8 to 3.6% and Enterococcus spp. at rates varying from 3.0 to 11.5%, while the molecular microbiology approach for E. coli was found to be as efficient, detecting contamination in 3.0% of samples. In contrast, CRENAME-rtPCR detected Enterococcus spp. in 27.9% of samples while the E. faecalis/faecium molecular assay did not uncover a single contaminated sample, thereby revealing a discrepancy in the coverage of waterborne enterococcal species detected by classical and molecular microbiology methods. The validation of the CRENAME-E. coli rtPCR test as a new tool to assess the quality of drinking water will require larger scale studies elaborated to demonstrate its equivalence to approved methods.

Ability of three DNA-based assays to identify presumptive Escherichia coli colonies isolated from water by the culture-based mFC agar method.

We tested the ability of three PCR assays, targeting uidA and tuf genes to correctly identify Escherichia coli colonies isolated from water and we compared them to two ß-glucuronidase-based culture methods (Colilert(®) and Readycult(®)), in terms of specificity and sensitivity. E. coli isolates recovered on mFC agar were first tested for the presence of the uidA positive colonies were presumed to be E. coli. For further characterization, uidA-negative colonies were subsequently identified using the Vitek 2 automated system. Colilert(®) and Readycult(®) detected 436 and 442 of 468 colonies identified as E. coli on mFC corresponding to sensitivities of 93.2 and 94.4%, respectively. None of the 59 non-E. coli isolates was detected by both methods for a specificity of 100%. Two (2) uidA and 1 tuf PCR assays were also tested. The uidA PCR assays yielded positive signals for 447 (95.5%) and 434 (92.7%) of 468 E. coli isolates tested respectively, whereas the tuf PCR assay showed a sensitivity of 100%. None of the 59 non-E. coli isolates was detected by both uidA PCR assays (100% specificity), whereas tuf PCR false-positive signals were obtained with Escherichia fergusonii and Escherichia albertii. However, since these 2 species are principally found in the feces of mammals and birds, their detection indicates a fecal contamination. Consequently, using a 1-h tuf rtPCR assay to confirm the identity of E. coli colonies on mFC agar is as specific, more sensitive, and potentially more cost-efficient than culture methods based on ß-glucuronidase detection.