Efficacy of RT-PCR for the detection of E. coli in drinking water

Escherichia coli (E. coli) is an indicator of potential human fecal contamination. Polymerase Chain Reaction (PCR) as an ideal detection method for detecting E. coli has some advantages like rapidity, high sensitivity and accuracy, easy performance, ability to run in high numbers and inexpensiveness. On the other hand, the disadvantage of PCR is possibility of its false positive results. In this study, reverse transcriptase polymerase chain reaction (RT-PCR) method was used to overcome the problem, and the results were compared to most probable number (MPN). 16srRNA forward (SF) and 16srRNA reverse (SR) primers were designed using E. coli 16srRNA sequence. After preparing different diluted samples of E. coli in distilled water, the bacteria were separated by FHLP and HAWP filters and its 16srRNA was propagated using mentioned primers. To confirm the sensitivity of the RT- PCR method compared to MPN one, samples obtained from 15 water sources in Arak city were examined. The number of bacteria in dilutions were confirmed with culture. RT-PCR data showed that FHLP compared to HAWP filters have a higher capability in separating of bacteria in different dilutions. Also there was a higher sensitivity of RT-PCR compared to RT- PCR and MPN. RT-PCR can detect the bacteria in lower dilutions of bacterial suspension. Hydrophobic filters (e.g. FHLP) compared to hydrophilic filters (e.g. HAWP) have higher capability in separating bacteria. To detect all coliform bacteria RT-PCR amplifications achieved by cells concentrated with hydrophobic filters are recommended.