Q Yang1, F Wang, W Prinyawiwatkul, B Ge. 1. Department of Food Science, Louisiana State University Agricultural Center, Baton Rouge, LA, USA.
Abstract
AIMS: 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. METHODS AND RESULTS: 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. CONCLUSIONS: Our results demonstrated that LAMP is a robust alternative to PCR in Salmonella detection for food applications. SIGNIFICANCE AND IMPACT OF THE STUDY: 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.
AIMS: 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. METHODS AND RESULTS: 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. CONCLUSIONS: Our results demonstrated that LAMP is a robust alternative to PCR in Salmonella detection for food applications. SIGNIFICANCE AND IMPACT OF THE STUDY: 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.