Establishment of a real-time fluorescence quantitative polymerase chain reaction assay for rapid detection of Salmonella in stool samples of humans and animals

ObjectiveTo rapidly detect and identify Salmonella and improve the response to the outbreaks of foodborne disease, we established a real-time fluorescence quantitative polymerase chain reaction (qPCR) assay. The specificity, sensitivity and detection limit of the assay were evaluated.

MethodsFirstly, we screened some specific genes of Salmonella by using conventional PCR, then established a qPCR assay targeting one specific gene selected. The specificity, sensitivity and detection limit of the qPCR assay were evaluated by pure strains from different species of enterobacteria, Salmonella strains in different species and serotypes, and stool samples from humans and animals.

ResultsOne specific gene, ttrA, was selected as the candidate gene for establishing PCR assay to detect Salmonella. The detection limit of the qPCR assay with purified DNA as template was 2 copies per reaction. The amplification results of qPCR assays were positive for all 1 100 Salmonella isolates of different species and serotypes, while the results were negative for all the strains of other enterobacteria. For 360 samples of stool enrichment, including 150 Salmonella culture positive human stool samples and 210 Salmonella culture positive animal stool samples, the qPCR detection results were all positive.

ConclusionA qPCR assay based on a single gene to identify and detect Salmonella was established for the molecular detection and characterized by high specificity and high sensitivity. It can be used for the rapid detection and identification of Salmonella and the diagnosis of infectious diarrhea caused by Salmonella.