环介导技术在感染性腹泻沙门菌检测中的应用与研究

Application and study of LAMP technique in the detection of Salmonella in infectious diarrhea

目的建立并应用环介导等温扩增法(LAMP)检测肠道门诊感染性腹泻中的沙门菌。方法通过在线软件Primer Explorer V5设计针对沙门菌的保守区引物,建立LAMP检测方法,采用荧光定量PCR法、LAMP法和分离培养法,对本院2015年肠道门诊137例患者的粪便标本进行检测。对建立的方法进行特异度、灵敏度试验,与荧光定量PCR法和培养法进行比对。结果 137份标本中,荧光定量PCR法检出28份阳性,LAMP法检出28份阳性,分离培养法检出23份阳性。3种方法的检测结果差异无统计学意义(P>0.05)。LAMP对6株沙门菌属扩增的结果均为阳性,而对志贺菌等5株致病菌株扩增的结果均为阴性。LAMP检测方法具有良好的灵敏度和特异度。结论 LAMP检测方法灵敏特异,简单快速,比荧光定量PCR法更适用于基层疾病预防控制机构,有着较为广泛的发展前景。

Objective To establish and apply the methods of LAMP assay for the detection of Salmonella in infectious diarrhea patients from the intestine clinic. Methods The specific primers of the conserved regions for Salmonella was prepared by using Primer Explorer V5 online,and the LAMP assay was established. The 137 fecal specimens from diarrhea patients in the intestine clinics in our hospital were tested by real- time PCR,LAMP and the separation and culturing method in 2015. The specificity and sensibility test was performed on the established method,and compared with real- time PCR and the separation and culturing method. Results 28 specimens were determined as positive by real- time PCR,28 by LAMP and 23 specimens were determined as positive by the separation and culturing method in 137 specimens. There was no statistical significance on difference on the detection results with the 3 methods（ P〉0. 05）. The amplification result of the 6 Salmonella strains were positive,but that of the other pathogens were all negative,such as Shigella. LAMP detection showed fine specificity and high sensitivity.Conclusion LAMP assay is sensitive,efficient,rapid and simple,which is better than real- time PCR in primary diseases control and prevention agency,and which has a brighter development prospect.