双重荧光定量PCR检测携带mcr-1基因的鼠伤寒沙门菌的建立与应用  

Establishment and application of double fluorescent quantitative PCR in detection of Salmonella typhimurium carrying mcr-1 gene

作　　者：沈伟伟[1,2] 盛莹 管雅雅[1] 陈喜凯 李聪聪[1] 裘丹红 SHEN Wei-wei;SHENG Ying;GUAN Ya-ya;CHEN Xi-kai;LI Cong-cong;QIU Dan-hong(Taizhou Municipal Center for Disease Control and Prevention,Taizhou,Zhejiang 318000,China;不详)

机构地区：[1]台州市疾病预防控制中心,浙江台州318000 [2]浙江省微生物技术与生物信息研究重点实验室,浙江杭州310012

出　　处：《中国卫生检验杂志》2020年第12期1412-1415,共4页Chinese Journal of Health Laboratory Technology

基　　金：浙江省微生物技术与生物信息研究重点实验室开放基金(2017E10010)。

摘　　要：目的建立可用于鼠伤寒沙门菌和mcr-1基因快速筛查的双重荧光定量PCR方法。方法采用Primer Express v3.0软件设计引物及探针,采用双重荧光定量PCR方法检测鼠伤寒沙门菌和mcr-1基因阳性质粒,建立循环数与模板浓度关系的标准曲线。同时用于食源性疾病监测分离的鼠伤寒沙门菌中携带mcr-1基因的快速筛查。结果建立的双重荧光定量PCR方法检测鼠伤寒沙门菌的线性范围为2.20×10^1 cfu/ml^2.20×10^7 cfu/ml,检测mcr-1基因的线性范围为1.40×10^1拷贝/μl^1.40×10^8拷贝/μl,标准曲线的相关系数分别是0.997和0.998。应用该方法在132株临床分离的鼠伤寒沙门菌中共检出3株携带mcr-1基因的鼠伤寒沙门菌,mcr-1基因阳性率为2.3%。结论建立的双重荧光定量PCR方法可用于携带mcr-1基因的鼠伤寒沙门菌的快速筛查,应该加强对本地区的食源性疾病病原菌的耐药性持续监测。Objective To establish a dual-quantitative PCR method for the rapid screening of Salmonella typhimurium carrying mcr-1 gene.Methods Primer Express v3.0 software was used to design primers and probes.Double-fluorescence quantitative PCR was used to detect Salmonella typhimurium and mcr-1 gene positive plasmids,and a standard curve of the relationship between cycle number and template concentration was established.At the same time,the method was used to screen Salmonella typhimurium strains carrying mcr-1 gene isolated from patients with foodborne diseases.Results The linear range of the detection of Salmonella typhimurium by double-quantitative PCR method was 2.20×10^1 cfu/ml-2.20×10^7 cfu/ml.The linear range of detection of mcr-1 gene was 1.40×10^1 copies/μl-1.40×10^8 copies/μl.The correlation coefficients of the standard curve are 0.997 and 0.998,respectively.Three strains of Salmonella typhimurium carrying mcr-1 were detected in 132 Salmonella typhimurium strains isolated from patients,and the positive rate of mcr-1 gene was 2.3%.Conclusion The established dual-quantitative PCR method can be used for the rapid screening of Salmonella typhimurium carrying mcr-1 gene.Continuous monitoring of drug resistance of foodborne pathogens should be strengthened.

关 键 词：鼠伤寒沙门菌 mcr-1基因 双重荧光定量PCR 

分 类 号：R446.5[医药卫生—诊断学]