Improvement of Loop-Mediated Isothermal Amplification Combined with Chromatographic Flow Dipstick Assay for Salmonella in Food Samples

Loop-mediated isothermal amplification (LAMP) had been employed as a powerful tool to facilitate genetic tests for various food pathogens, as it is easy to perform. Recently, various methods of detecting the LAMP amplicon were developed. In this study, we improved two LAMP assays by combining LAMP with chromatographic flow dipstick (LFD) assays for Salmonella (targeting phoP and invA, respectively). We evaluated different labeled primer sets, then selected the optimal sets to perform the LFD assays. We compared the optimal LFD and LAMP assays with the ISO standard method. The results showed that LFD was more sensitive and quicker than LAMP. Furthermore, enrichment broths of 225 food samples were tested. The sensitivity of two LFD assays was 100%. The specificity of LFD assay targeting phoP was 99.1%, and LFD assay targeting invA was 99.5%. For the LFD assay targeting phoP, the estimate of limit of detection (LOD) 50% was 0.061 CFU/g and the estimate of LOD 95% was 0.265 CFU/g. For the LFD assay targeting invA, the estimate of LOD 50% was 0.040 CFU/g and the estimate of LOD 95% was 0.172 CFU/g. We validated this method in a primary laboratory, where we accomplished the assay only using an incubator and a heating block. It suggested that the LFD assay had the potential to become a suitable diagnostic method in field test and primary labs.