A Novel Real-time PCR Method for Rapid Molecular Detection and Serotyping of Salmonella Typhimurium

Introduction: In Europe, food safety regulation was recently reinforced to prevent poultry meats adulterated with either Salmonella Enteritidis or S. Typhimurium from being marketed. While the current Kauffmann-White (K-W) serotyping method requires 4/5 days to assess the presence of these pathogens, two alternative real-time PCR methods have been developed, allowing their specific detection within one day. As the S. Enteritidis method, the new S. Typhimurium method described here includes broth enrichment and DNA extraction method and can be used either as a rapid serotyping step after the detection of Salmonella spp. or as a direct combined screening & serotyping method. In order to optimize the specificity of the assay, a dedicated method of analysis was developed, based on a dual target PCR system.

Purpose: This study aimed at evaluating key performances of this new real-time PCR S. Typhimurium kit on food and environmental samples: the intrinsic detection limit (IDL), and the inclusivity & exclusivity. 

Methods: For estimation of IDL, serial dilutions of pure cultures of ATCC14028 Salmonella Typhimurium, were subjected to both Salmonella spp. and S. Typhimurium methods. IDL was defined as the concentration at which 9 out of 10 independent extracts were positive. For inclusivity and exclusivity, this study includes both pure strain extracts and spiked environmental and food samples extracts.

Results: IDL was estimated at 250 CFU/ml. All the 105 inclusivity Salmonella Typhimurium strains were tested positive. 96% of the exclusivity samples (including 339 Salmonella and 46 non-Salmonella strains) yielded negative results.

Significance: Together with the S. Enteritidis method, this novel method will allow a faster release of the products and a significant reduction of the number of samples to be confirmed by the standard K-W method.