产肠毒素大肠杆菌快速检测方法的建立和评价

目的 利用环介导等温扩增（LAMP）技术，建立产肠毒素大肠杆菌（ETEC）的快速、便捷、敏感、特异的检测方法，并对该方法的特异性和敏感性进行评价，为实验动物检测和细菌性腹泻的诊断提供技术支持。方法 根据GenBank公布的产肠毒素大肠杆菌的LT毒素基因序列（S60731.1）设计外引物和内引物进行LAMP扩增，对LAMP特异性和敏感性与PCR方法做比较。结果 建立的LAMP方法检测限为100pg，灵敏度是PCR的10倍以上并具有较高的特异性，利用该方法对27份猴腹泻样品进行LAMP和PCR方法检测， LAMP（60min内）检测结果与PCR符合率为100%,而LAMP（90分钟内）检测敏感性高于PCR。结论 建立了一种用于检测肠毒性大肠杆菌（ETEC）的LAMP检测方法，该方法特异性强，灵敏度高，方便快捷，适合于ETEC临床快速检测。

Establishment and Evaluation of the LAMP in Detection of ETEC

Objective By using loop-mediated isothermal amplification (LAMP) assay, to found the fast, convenient, sensitive and specific method of enterohaemorrhagic escherichia coli (ETEC), and evaluating the specificity and sensitivity of the method, for detection and diagnosis of bacterial diarrhea in laboratory animals. Methods According to the published LT toxin gene sequences (S60731.1) to design PCR and LAMP primers, the LAMP specificity and sensitivity compared with PCR. Results the LAMP method established detection limit was 100 pg, sensitivity is more 10 times than PCR and has the high specificity. Using PCR and LAMP to detect 27 feces samples of monkey diarrhea, we had the same results of LAMP (within 60 minutes) and PCR. And LAMP(within 90 min) has more sensitive than PCR. Conclusion we had established the ETEC LAMP detection method, this method is strong specificity, high sensitivity, fast and convenient, suitable for rapid detection of ETEC clinical.