Differentiation of salmonella isolates using PCR and high-resolution melt curve analysis

Poultry products can be contaminated with Salmonella isolates which may cause foodborne disease in humans. The aim of this study was to optimise a rapid diagnostic test that can differentiate Salmonella isolates without requiring DNA sequencing. Specific primers were used to amplify hem-D gene of 16 different Salmonella reference strains and 65 clinical isolates from commercial poultry farms. PCR products were applied to high-resolution melt curve analysis and Salmonella isolates were differentiated based on their HRM curves. Analysis of the nucleotide sequence of the amplicons from selected isolates confirmed that each melting curve profile was related to a unique DNA sequence. The relationship between reference strains and tested specimens was also evaluated using a mathematical model without visual interpretation of HRM curves. In addition, the potential of the PCR-HRM curve analysis was evaluated for genotyping of additional Salmonella isolates from different avian species. The findings indicate that PCR followed by HRM curve analysis provides a rapid and robust technique for genotyping of Salmonella isolates to determine the Salmonella strains.

Key words: Salmonella, PCR, high resolution melting curve analysis, strain differentiation