Development of a multiplex recombinase polymerase amplification coupled with lateral flow dipsticks for the simultaneous rapid detection of Salmonella spp., Salmonella Typhimurium and Salmonella Enteritidis

Objectives Salmonella spp. is a world-leading foodborne pathogen and its rapid detection is essential for ensuring food safety. Conventional methods require expensive instruments, considerable operational skills and cannot provide fast mobile on-site systems to detect Salmonella in food.Materials and Methods A visual method was established based on multiple recombinase polymerase amplification (RPA) coupled with lateral flow dipsticks (LFD) for the simultaneous detection of Salmonella spp., Salmonella Enteritidis and Salmonella Typhimurium in vitro and food.Results The optimal volume and temperature for the multiplex RPA-LFD method were determined to be 25 mu L and 38 degrees C, respectively. The reaction process was completed within 25 min and the results were observed visually. The limits of detection (LODs) were 2.8x102, 5.9x102, and 7.6x102 CFU/mL for Salmonella spp., S. Enteritidis and S. Typhimurium, respectively. Meanwhile, the results of the established method showed no cross-reactivity between the Salmonella cells and other common foodborne bacteria, which was highly specific for Salmonella. More importantly, the developed method exhibited good performance in artificially contaminated chicken samples with the LODs of 2.8x103, 5.9x103, and 7.6x103 CFU/mL for Salmonella spp., S. Enteritidis, and S. Typhimurium, respectively. Finally, the application of the multiple RPA-LFD methods in retailed food samples displayed that this method was effective and practical for the detection of Salmonella spp. in food.Conclusion The developed multiplex RPA-LFD method provides a new sensitive and rapid alternative for the specific detection of Salmonella spp. and its important serovars in food.