Krascsenicsová K, Piknová L, Kaclíková E, Kuchta T
Department of Microbiology and Molecular Biology, Food Research Institute, Bratislava, Slovakia.
Lett Appl Microbiol. 2008 Apr;46(4):483-7. doi: 10.1111/j.1472-765X.2008.02342.x. Epub 2008 Mar 13.
A new real-time polymerase chain reaction-based method was developed for the detection of Salmonella enterica in food.
The method consisted of a novel two-step enrichment involving overnight incubation in buffered peptone water and a 5-h subculture in Rappaport-Vassiliadis medium, lysis of bacterial cells and a Salmonella-specific 5'-nuclease real-time PCR with an exogenous internal amplification control. Because a two-step enrichment was used, the detection limit for dead S. enterica cells in artificially contaminated ice cream and salami samples was high at 10(7 )CFU (25 g)(-1), eliminating potential false-positive results. When the method was evaluated with a range of 100 naturally contaminated food samples, three positive samples were detected by both the real-time PCR-based method and by the standard microbiological method, according to EN ISO 6579. When the real-time PCR-based method was evaluated alongside the standard microbiological method according to EN ISO 6579 with 36 food samples artificially contaminated at a level of 10(0 )CFU (25 g)(-1), identical results were obtained from both methods.
The real-time PCR-based method involving a two-step enrichment produced equivalent results to EN ISO 6579 on the day after sample receipt.
The developed method is suitable for rapid detection of S. enterica in food.
开发一种基于实时聚合酶链反应的新方法用于检测食品中的肠炎沙门氏菌。
该方法包括一个新颖的两步富集过程，即在缓冲蛋白胨水中过夜培养，然后在Rappaport-Vassiliadis培养基中进行5小时的继代培养，裂解细菌细胞，并使用外源性内部扩增对照进行沙门氏菌特异性5'-核酸酶实时PCR。由于采用了两步富集法，人工污染的冰淇淋和萨拉米香肠样品中死的肠炎沙门氏菌细胞的检测限高达10(7) CFU (25 g)(-1)，消除了潜在的假阳性结果。当用一系列100份自然污染的食品样品评估该方法时，基于实时PCR的方法和根据EN ISO 6579的标准微生物学方法均检测到3份阳性样品。当根据EN ISO 6579将基于实时PCR的方法与标准微生物学方法一起用36份人工污染水平为10(0) CFU (25 g)(-1)的食品样品进行评估时，两种方法得到了相同的结果。
涉及两步富集的基于实时PCR的方法在收到样品后的第二天产生了与EN ISO 6579等效的结果。
所开发的方法适用于食品中肠炎沙门氏菌的快速检测。
