Simultaneous Detection of Enteric Bacteria by QPCR in Surface Waters Located in an Urban Area

ABSTRACT

A rapid quantitative polymerase chain reaction (QPCR) method was developed for simultaneous detection of enteric bacteria from surface waters by utilizing a pair of universal primers that targeted four bacteria strains, namely Shigella dysenteriae, Vibrio cholerae, Salmonella typhimurium, and Escherichia coli. It was estimated that the QPCR method had a 94% confidence, and a detection limit as 2.7 Escherichia coli cells per sample in undiluted DNA extracts. The QPCR method was applied for the bacteriological examination of several surface waters in the urban area of Xi'an, China, and comparison was made with the conventional bacteria indicators determined by conventional membrane filter (MF) method. As a result, the calibrator cell equivalents (CCE) determined by QPCR was 2.2 to 5 times of the total coliform CFU, and the characteristics of the bacterial quality of different waters could be well presented by the QPCR results with a higher sensitivity. The coefficient of variation (CV) of data obtained by QPCR was smaller than that by traditional MF method, indicating a more stable analysis result. The QPCR method established by this study has manifest advantages over conventional methods in its rapidness and sensibility for the detection of pathogenic bacteria from surface water. It would provide a more reliable approach for the assessment of bacteriological risk of water environment.

Key Words:

• universal primer
• enteric pathogenic bacteria
• quantitative polymerase chain reaction (QPCR)
• membrane filtration (MF)
• surface water

ACKNOWLEDGMENTS

The study was supported by the National Natural Science Foundation of China (NSFC, Grant No. 50848045) and the Program for Changjiang Scholars and Innovative Research Team in University (Grant No. IRT0853).

Notes

^a SXW: Research Institute of Microorganism in Shaanxi Province;

^b ATCC: American Type Culture Collection.

^aGeometric mean of all sampling locations (n = 12 at every sampling location).

^bLog₁₀ mean of all sampling visits.

^cStandard deviation.

^dLog₁₀ SD between sampling visits.

^eCoefficient of variation (= SD in original units/mean).