Yang Q, Wang F, Prinyawiwatkul W, Ge B
Department of Food Science, Louisiana State University Agricultural Center, Baton Rouge, LA, USA.
J Appl Microbiol. 2014 Jan;116(1):81-8. doi: 10.1111/jam.12340. Epub 2013 Oct 1.
Loop-mediated isothermal amplification (LAMP) assays have been developed recently for Salmonella detection. This study aimed at evaluating the robustness of two Salmonella LAMP assays in comparison with PCR and real-time quantitative PCR for food applications.
Performance of the assays was examined under abusive preparation conditions, running temperatures and pH, and with the addition of various inhibitors and food rinses. LAMP achieved robust detection under abusive assay preparation conditions (holding at 22 and 37°C for up to 30 min) and running temperatures (57-68°C). With a hot-start DNA polymerase, PCR obtained comparable results under these temperature ranges. However, PCR performed markedly poorer under abusive pH. LAMP also showed greater tolerance to potential inhibitors than PCR. When food rinses including meat juice, chicken rinse, egg homogenate and produce homogenate were added at 20% of the reaction mix, PCR amplifications were completely inhibited, but LAMP reactions were not.
Our results demonstrated that LAMP is a robust alternative to PCR in Salmonella detection for food applications.
This study filled important knowledge gaps regarding the robustness of Salmonella LAMP assays. The findings will help bring Salmonella LAMP assays closer to wider applications in food testing.
环介导等温扩增（LAMP）检测方法最近已被开发用于沙门氏菌检测。本研究旨在评估两种沙门氏菌LAMP检测方法相对于聚合酶链反应（PCR）和实时定量PCR在食品应用中的稳健性。
在恶劣的制备条件、运行温度和pH值下，以及添加各种抑制剂和食品冲洗液的情况下，检测了这些检测方法的性能。LAMP在恶劣的检测制备条件（在22和37°C下保持长达30分钟）和运行温度（57 - 68°C）下实现了稳健的检测。使用热启动DNA聚合酶，PCR在这些温度范围内获得了可比的结果。然而，PCR在恶劣的pH值下表现明显较差。LAMP对潜在抑制剂的耐受性也比PCR更强。当在反应混合物中添加20%的包括肉汁、鸡肉冲洗液、鸡蛋匀浆和农产品匀浆在内的食品冲洗液时，PCR扩增被完全抑制，但LAMP反应未受影响。
我们的结果表明，在食品应用中沙门氏菌检测方面，LAMP是一种比PCR更稳健的替代方法。
本研究填补了关于沙门氏菌LAMP检测方法稳健性的重要知识空白。这些发现将有助于使沙门氏菌LAMP检测方法更接近在食品检测中的广泛应用。
