环介导等温扩增法检测空肠弯曲菌

目的 空肠弯曲菌是重要的食源性疾病病例标本的病原菌检测项目,用环介导等温扩增(Loop-mediated isothermal amplification,LAMP)的方法,对食源性致病菌空肠弯曲菌进行快速检测。方法 以空肠弯曲菌 mapA 基因序列为检测靶基因,建立了空肠弯曲菌的LAMP 快速检测方法。空肠弯曲菌的LAMP反应由DNA扩增试剂盒提供的反应体系加入引物和模板进行。做了空肠弯曲菌DNA模板灵敏度的检测、菌液灵敏度的检测和特异性检测。结果 我们方法对空肠弯曲菌DNA 的检测灵敏度为84.5fg/反应管;对空肠弯曲菌菌液检测灵敏度为0.8 CFU/反应管;特异性100%。结论 对食源性疾病腹泻样本的空肠弯曲菌检测项目,可以采用mapA-LAMP方法进行等温扩增初筛,初筛阳性的样品再有针对性地进行后续的传统培养。

Campylobacter jejuni detected by Loop-mediated isothermal amplification

Objective To detect the susceptibility of food borne pathogenic C.jejuni to Loop-mediated isothermal amplification(LAMP).Methods Taking the MAPA gene sequence of C.jejuni as the target gene,the LAMP method for rapid detection of C.jejuni was introduced,and then carried out by the addition of specific primers and templates for C.jejuni to the reaction mixture provided by the DNA amplification kit.The sensitivity testing for DNA template density,C.jejuni culture suspension density and the specificity procedures were conducted.Results In this study,the sensitivity of LAMP assay for detecting genomic DNA and culture suspension was 84.5 fg/reaction tube and 0.8 CFU/reaction tube,respectively.The specificity was 100%.Conclusion The LAMP screening established in this study can be conducted to investigate the presence of C.jejuni in fecal specimens of foodborne diseases.For LAMP-positive specimens,they will be cultured immediately from the stored stool specimens through routine procedures.