饮用水中大肠埃希菌双重PCR检测方法建立  

Establishment of duplex-PCR assay for detection of Escherichia coli in drinking water

作　　者：刘艳超 王荣敏 谢佳鑫 王明晓[3] LIU Yan-chao;WANG Rong-min;XIE Jia-xin(Department of Epidemiology,School of Public Health,Inner Mongolia Medical University,Inner Mongolia Autonomous Region 010110,China;不详)

机构地区：[1]内蒙古医科大学公共卫生学院流行病学教研室,内蒙古呼和浩特010110 [2]招远市疾病预防控制中心 [3]石药集团中奇制药技术(石家庄)有限公司

出　　处：《中国公共卫生》2021年第3期574-576,共3页Chinese Journal of Public Health

基　　金：内蒙古医科大学实验室开放基金(2017KF06);内蒙古医科大学教改资助(NYJXGG2017028)。

摘　　要：目的建立饮用水大肠埃希菌的双重PCR检测方法,并评价效果。方法选取大肠埃希菌的保守序列16SrRNA基因E-16SrDNA和乳糖操纵子LacI基因序列E-LACI作为双重PCR方法的两段扩增基因,根据GenBank上已知序列设计合成两段基因引物,使用大肠埃希菌标准菌株构建双重PCR方法。使用双重PCR方法和国标法同步检测所采集水样中的大肠埃希菌,对比评价双重PCR方法的真实性、可靠性和预测概率。确定双重PCR方法检验水样中大肠埃希菌的应用范围。结果成功构建饮用水中大肠埃希菌检测的双重PCR方法,本方法最低检测限为40 CFU/L,灵敏度为66.7%,特异度为100%,阳性预测值为100%,阴性预测值为97.7%,与国标法的一致率为97.8%。结论本研究所建立的饮用水中大肠埃希菌双重PCR检测方法具有较高的真实性和灵敏性,适用于饮用水中大肠埃希菌的快速检验。Objective To establish a duplex-PCR assay for the detection of Escherichia coli(E.coli)in drinking water and to assess detection results of the assay.Methods First,the conservative sequence 16SrRNA gene of E.coli(E-16SrDNA)and the lactose operon LacI gene sequence(E-LACI)were selected as the two segments amplification gene of the duplex-PCR assay.According to the known sequence from the GenBank,the primers of the two segment genes were designed and synthesized,and duplex-PCR assay was established using the standard E.coli strain.Then,E.coli in drinking water samples was detected with both the duplex-PCR assay and the national standard method to evaluate the validity,reliability and prediction probability of the established assay.Results For the duplex-PCR assay established,the minimum detection concentration of the E.coli in water samples was 40 colony forming unit(CFU)/L;the sensitivity was 66.7%;the specificity was 100%;the positive predictive value was 100%;the negative predictive value was 97.7%;and the consistent rate for the detection results versus those with the national standard method was 97.8%.Conclusion The established duplexPCR assay is of high validity and sensitivity and is suitable for rapid detection of E.coli in drinking water samples.

关 键 词：大肠埃希菌 双重PCR 饮用水检验 16Sr RNA 乳糖操纵子(LacI) 

分 类 号：R117[医药卫生—公共卫生与预防医学]