Development of a multiplexed PCR assay combined with propidium monoazide treatment for rapid and accurate detection and identification of three viable Salmonella enterica serovars
Development of a multiplexed PCR assay combined with propidium monoazide treatment for rapid and accurate detection and identification of three viable Salmonella enterica serovars
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开发多重 PCR 检测结合单叠氮化丙啶处理，可快速、准确地检测和鉴定三种活的肠沙门氏菌血清型
DOI：
10.1016/j.foodcont.2012.05.061
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发表时间：
2012-12
期刊：
影响因子：
6
通讯作者：
Wei, Hua
中科院分区：
文献类型：
--
作者：
Xu, Hong;Aguilar, Zoraida P.;Sun, Jichang;Wei, Hua
沙门氏菌是食品和受污染的水中的主要致病菌。本研究旨在建立一种快速、可靠的同时检测沙门氏菌3种主要血清型（伤寒沙门氏菌、伤寒副伤寒沙门氏菌和伤寒沙门氏菌）的技术。设计引物扩增这三种血清型的特异性基因，用聚合酶链反应（PCR）同时检测。为了确保只检测到活细胞，应用单叠氮丙啶（PMA）选择性抑制来自死细胞的DNA信号。结果表明，在纯培养物浓度为1 × 106CFU/ml或在加标食品（番茄、鸡肉、牛肉和火腿）中浓度为1 × 106CFU/g时，pma -多重PCR （PMA-mPCR）检测结果均为阴性。结果表明，PMA-mPCR法的检出限分别为102CFU/ml（伤寒沙门氏菌4.3 × 102CFU/ml、副伤寒沙门氏菌3.7 × 102CFU/ml、伤寒沙门氏菌7.2 × 102CFU/ml）和103CFU/g（伤寒沙门氏菌4.3 × 103CFU/g、副伤寒沙门氏菌3.7 × 103CFU/g、伤寒沙门氏菌7.2 × 103CFU/g）。这些结果表明，PMA-mPCR方法可以在短时间内同时检测和鉴定出伤寒沙门氏菌、副伤寒杆菌和伤寒杆菌，即使在食品生产中也是如此。
Salmonella is the leading pathogenic bacteria in food and contaminated water. The aim of this study was to develop a rapid and reliable technique for simultaneous detection of the main three serotypes (Salmonella enterica serovars Typhimurium, Paratyphi B and Typhi) of Salmonella. Primers were designed to amplify the genes specific to each of these three serotypes for simultaneous detection using polymerase chain reaction (PCR). To ensure the detection of only viable cells, propidium monoazide (PMA) was applied to selectively suppress the DNA signal from dead cells. Results showed that the PMA-multiplexed PCR (PMA-mPCR) assay always gave negative results for heat-killed Salmonella at concentrations up to 1 × 106CFU/ml in pure culture or 1 × 106CFU/g in spiked food products (tomato, chicken, beef and ham). Results showed that the detection limits of the PMA-mPCR assay were approximately 102CFU/ml (4.3 × 102CFU/ml for S. Typhimurium, 3.7 × 102CFU/ml for S. Paratyphi B, 7.2 × 102CFU/ml for S. Typhi) in pure culture and 103CFU/g (4.3 × 103CFU/g for S. Typhimurium, 3.7 × 103CFU/g for S. Paratyphi B, 7.2 × 103CFU/g for S. Typhi) in food produce. These results demonstrated that the PMA-mPCR assay can simultaneously detect and identify viable S. Typhimurium, Paratyphi B and Typhi in a short period of time, even in food produce.
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影响因子：
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