Application of loop-mediated isothermal amplification with propidium monoazide treatment to detect live Salmonella in chicken carcasses

Abstract

Raw chicken products are major causes of human foodborne salmonellosis worldwide. In particular, there is a significant risk of human exposure to Salmonella originating from the chicken slaughtering process. Controlling the contamination of chicken carcasses by Salmonella has been a considerable challenge in chicken-slaughtering facilities and involves routine microbiological monitoring using reliable detection methods. Simple and rapid detection methods, particularly those capable of determining cell viability, will significantly facilitate routine monitoring of Salmonella. Here, we report an invA-based loop-mediated isothermal amplification method coupled with a simple propidium monoazide treatment (PMA-LAMP) for simple and rapid detection and quantification of viable Salmonella in rinse water of chicken carcasses. In this study, PMA-LAMP consistently gave negative results for isopropanol-killed Salmonella with concentrations up to 8.0 × 106 CFU/reaction. The detection limit of PMA-LAMP was 8.0 × 101 CFU/reaction with viable Salmonella in both pure culture and rinse water of chicken carcasses, and 10-fold lower than a conventional polymerase chain reaction coupled with PMA (PMA-PCR) targeting invA. There was a high correlation (R2 = 0.99 to 0.976) between LAMP time threshold (TT) values and viable Salmonella with a quantification range of 1.0 × 103 to 1.0 × 108 CFU/mL in pure culture and rinse water of chicken carcasses. The PMA-LAMP assay took less than 2 h to detect Salmonella contaminated in test samples. Therefore, this simple and rapid method will be a very useful tool to detect live Salmonella contamination of chicken carcasses without pre-enrichment at the slaughterhouse where sanitizing treatments are commonly used.