多重荧光PCR结合HANDS系统快速检测4种食源性致病菌

目的建立快速、准确检测食品中沙门菌、志贺菌、金黄色葡萄球菌和溶血性链球菌的多重荧光PCR方法。方法将多重荧光PCR技术与HANDS系统结合,采用同源加尾的方式,构建四重荧光PCR体系,分析该体系的特异性,并进行模拟实验,该方法与国家标准检测方法进行方法比对实验。结果该系统检测出了4种目标致病菌。初始菌浓度分别为5、9、10、2、50、3cfu/ml的沙门菌、志贺菌、金黄色葡萄球菌和兰氏A群、C群、G群链球菌增菌液,在增菌18h后都被体系检出。该方法与国家标准方法对样品的检测结果一致。结论首次结合HANDS系统构建了同时检测4种致病菌的多重荧光PCR方法,该方法特异性强、灵敏度高,是检测沙门菌、志贺菌、金黄色葡萄球菌和溶血性链球菌的快速、准确方法。

Rapid Detection on Four Foodborne Pathogens Using HANDS System Combined with Multiplex Real-time PCR

OBJECTIVE To establish a rapid and accurate multiplex real-time PCR for simultaneous detection of Salmonella,Shigella,Staphylococcus aureus and Streptococcus hemolyticus in food.METHODS The reaction system used HANDS system combined with multiplex real-time PCR.The reaction system was developed.The specificity of this system were analyzed.Simulated contamination test were detected.Contrast experiments between this method and national standard method were analyzed.RESULTS The method detected the four foodborne pathogens.5cfu/ml for Salmonella,9 cfu/ml for Shigella,10 cfu/ml for Staphylococcus aureus,2 cfu/ml for group A Streptococcus,50 cfu/ml for group C Streptococcus,3 cfu/ml for group G Streptococcus in enrichment broth at 37℃ for 18h were determined.This method had the same result as national standard methods.CONCLUSION The method using HANDS system Combined with multiplex real-time PCR was established for the first time.It showed good specificity and provided an important method for rapid and accurate detection of Salmonella,Shigella,Staphylococcus aureus and Streptococcus hemolyticus.