Study On The Application Of The Detection Of Pathogen In Environmental Water By Quantitative PCR

In order to establish a method for detection of the enteric pathogenic bacteria in environmental water by real-time fluorescent quantitative polymerase chain reaction (QPCR) technique, this study tried to design and synthesize four kinds of primers with Shigella sp., Salmonella typhi, Escherichia coli. and Vibrio cholerae. Applying QPCR analysis method with four kinds of primers was developed to detect cell densities of the enteric pathogenic bacteria from five water bodies of Xi'an city for four months continuously.These five typical water bodies include two rivers, one of which is the source for urban water supply, two lakes and one secondary effluent of a domestic wastewater treatment plant.The detection results showed that four target strains could be detected specifically by the QPCR method. Combined with QPCR technique, we made the sensibility of this new method up to 4.56 CFU/μL. What's more, the accurate and sensitive measurements of these organisms could be obtained within five hours.For five environmental waters, Vibrio cholerae was not detectd in five water bodies. It indicated that this bacterium was not usually present in surface water of Xi'an city. The concentration of Escherichia coli determined by QPCR was the highest frequency in several main pathogenic bacteria. In Beihu Lake and Xingqinghu Lake, it could be detected for a long period of time after water pollution.For Chanhe River and Beishiqiao Secondary Effluent, Escherichia coli. was the main body of enteric pathogenic bacteria in water, which intensity were approximated to the total bacterial intensity detected with universal primers. Although the concentration of Shigella sp. and Salmonella typhi detected continuously were much lower than Escherichia coli., which also could be used as an indicator of water pollution. Regarding different water samples, Beishiqiao Secondary Effluent without disinfection was the highest concentration of different pathogenic bacteria compared with other water bodies according to QPCR results, followed in sequence by Chanhe River, Beihu Lake and Xingqinghu Lake. Heihe River of high quality as source water was lowest according to QPCR results.The QPCR method established by this study can be used to detect pathogenic bacteria from environmental water. It provides a more reliable supplementary method for conventional culture method. As the QPCR technique continues to develop and improve, it will be more and more wildly applied in the field of environmental microorganisms.