荧光定量PCR技术在食源性疾病病原菌快速检测上的应用与评估  

Expanded application and evaluation of fluorescent quantative PCR in rapid detection of food-borne pathogenic bacteria

作　　者：谢晓红[1] 韩华忠 沈莉[1] 王仁刚[1] 陶力新 

机构地区：[1]上海市奉贤区疾病预防控制中心,上海201400

出　　处：《中国卫生检验杂志》2011年第12期2890-2892,2895,共4页Chinese Journal of Health Laboratory Technology

摘　　要：目的:应用荧光定量PCR技术,实现对食源性疾病病原菌的快速检测。方法:选取食源性疾病常见的四种病原菌,沙门菌、志贺菌、副溶血性弧菌和金黄色葡萄球菌,根据它们的基因保守区设计特异性引物和Taqman探针,建立荧光定量PCR技术,同时验证方法的敏感性和特异性,并与普通PCR、常规培养法进行比较。结果:用荧光定量PCR技术检测沙门菌、志贺菌、副溶血性弧菌和金黄色葡萄球菌的检测灵敏度可达4 CFU/ml～8 CFU/ml,其敏感性比普通PCR高10倍,比常规培养法高1000倍。且快速简便,特异性强,从核酸提取至完成检测最快能在2 h～3 h内完成,而常规培养需3 d～4 d。通过对282件样品的检测,证实该方法可以使阳性检出率从常规法的8.51%提高至14.18%。结论:建立的荧光定量PCR技术特异性强,敏感性高,操作简便快速,适用于食源性疾病病原菌的快速检测。Objective：To rapidly detect food-borne pathogenic bacteria with fluorescent quantative PCR.Methods： Specific primers and Taqman probes were designed based on evolutionary conserved sequence of Salmonella,Shigella,V.parahaemolyticus,S.aureus,which are most common food-borne pathogenic bacteria.PCR was set up to check its sensitivity and specificity,and compared with regular PCR and regular bacteria culture.Results： Sensitivity of PCR detection for Salmonella,Shigella,V.parahaemolyticus,S.aureus is 4 CFU/ml^8 CFU/ml,which is 10 times more sensitive than regular PCR,and 1000 times more sensitive than regular bacteria culture.In addition,it is fast,convenient and specific,and could be done within 2~3 hours from nucleic acid extraction until detection,whereas it is 3~4 days with regualr bacteria culture.The positive ratio is 14.18%,versus 8.51% with regular assay,based on 282 samples.Conclusion： The fluorescent quantative PCR is specific,sensitive,easy to handle,and could be practically used to detect food-borne pathogenic bacteria.

关 键 词：荧光定量PCR 食源性疾病 快速检测 

分 类 号：R446.5[医药卫生—诊断学]