Detection and differentiation of Salmonella Enteritidis and Salmonella Typhimurium by multiplex quantitative PCR from different poultry matrices

Abstract

ABSTRACT 1. The aim of this study was to develop a multiplex quantitative polymerase chain reaction (qPCR) based molecular diagnostic kit for rapid diagnosis of Salmonella enterica serovar Enteritidis and S. enterica serovar Typhimurium serotypes, which are frequently isolated worldwide from poultry samples. 2. Detection and discrimination of S. Enteritidis and S. Typhimurium were performed by targeting the sdf and the STM4492 (putative cytoplasmic protein) gene, respectively. The invA (invasion protein) gene was used to detect Salmonella spp. as a target gene, since it is considered a standard. In this study, a total of 200 bacterial strains (178 Salmonella spp. strains and 22 other genera) were used to test the specificity and sensitivity of the developed kit. The limit of detection (LOD) of the assays was determined to be 100–101 cfu/25 g from chicken meat samples artificially contaminated by litter and 100–101 cfu/ml for cloacal swab samples. 3. The multiplex qPCR results were 100% compatible with conventional serotyping results while the specificity and sensitivity values were 100%. These findings indicated that the newly developed multiplex qPCR technique can provide an alternative method to conventional serotyping of S. Enteritidis and S. Typhimurium in laboratories lacking adequate infrastructure.