Common Enterovirus In Water Hige-throughput Rapid Quantitative Detection Method

ObjectiveCommon intestinal viruses of enterovirrs, rotavirus, enteric adenovirus in water, establishing fluorescence quantitative RT-PCR method and detection a variety of water. The research of this subject will understand the distribution of the various water enterovirus laid the foundation in further.SignificanceWater is the elements of people’s life. Drinking water, washing water, irrigation water, industrial water and so on. With the growth of population, the development of industrial and agricultural, people need use water also increase rapidly. However, because the water often insufficient pay attention to conservation efforts, the pollution of water has become a serious problem. Virus contamination of water bodies may lead to the outbreak of waterborne disease, the intermediary water-borne virus as enterovirus, which comes from the faecal contamination of water bodies, by the fecal-oral transmission, which can cause infectious diarrhea, hepatitis A, hepatitis E and other statutory infectious diseases. At present, the total number of bacteria, coliforms, thermotolerant coliforms and Escherichia coli are drinking water bacteriology health indicators, but in recent years with the detection of enteric viruses in the drinking water of bacteriological indicators qualified and contaminated drinking water sources caused by the growing number of reports of outbreaks of viral gastro-intestinal diseases, showed that bacteriological indicators can not be fully instructed the removal of enterovirus, bacteriological indicators of safety are not on behalf of the safety of drinking water. The bacteria have cell structure but virus have not, non-cell properties of the virus to make it stronger than bacteria to environmental stress tolerance. Resistant to disinfectants such as chlorine, the virus is stronger than bacteria, in the conventional treatment process, the killing of the bacteria can not direct the killing of the virus. Once the virus penetrate the water treatment process and survival in drinking water, indicate that the possibility of water-borne transmission of viral presence. The U.S.A, the-French in the drinking water health standards require that the detection of enterovirus is zero, in China not yet the virus detection included in the relevant standards. Therefore, to carry out the detection of enterovirus in water has a profound practical guiding significance for the future improvement of drinking water health standards.MethodsUsing TaqMan probe technology establish the detection of enterovirus generic type(enterovirus, EV-U), polio virus type I (poliovirus, PV I), rotavirus(Rotavirus, RV), enteric adenovirus(enteric adenovirus, EAdV) fluorescent quantitative RT-PCR detection methods. Containing the target sequence standard preparation of standard curves, for accurate quantification of the virus, assess the method of specificity, sensitivity, and reproducibility. By PV I spiked experiments and real-time fluorescent quantitative RT-PCR technology detected in the water PV I virus, comparison of membrane adsorption-elution method and electropositive material enrichment of the virus in the water. Using the establish EV-U, PV I, RV, EAdV fluorescent quantitative RT-PCR method and virus enrichment methods, detecting virus content of Beijing Marco Polo Bridge sewage treatment plant settling tank water, finished water effluent, Tianjin Haihe River and Weijin river.ResultsThis study established a TaqMan probe real-time fluorescent quantitative RT-PCR method can detect the enterovirus generic type, poliovirus type I, rotavirus and enteric adenovirus. The method specificity is100%, sensitivity respectively102copies/μl, reproducibility is good. Enrichment of the three viruses with different load virus spiked water samples were enriched and calculate the rates. Electropositive material method is not only the highest recovery rates and stable, recovery is91%to94%; while the membrane adsorption-elution method recoveries followed, and recovery is only70%to80%.To use the established EV-U testing methods to detect Beijing Marco Polo Bridge sewage treatment plant in March the secondary settling tank of the water, finished water effluent, Tianjin Haihe River and Weijin River were negative; to use the established PV I testing methods to detect Marco Polo Bridge sewage treatment plant in March the secondary settling tank of the water, finished water effluent, Tianjin Haihe River and Weijin River were negative; to use the established RV testing methods to detect Marco Polo Bridge sewage treatment plant in March the secondary settling tank of the wate, finished water effluent, Tianjin Haihe River and Weijin River were negative; to use the established EAdV testing methods to detect Marco Polo Bridge sewage treatment plant in March the secondary settling tank of the water and finished water effluent were positive, Tianjin Haihe River and Weijin River were negative.