Efficient differentiation between Salmonella Pullorum and Salmonella Gallinarum by a fimH-based PCR-HRM

Salmonella Pullorum (S. Pullorum) and Salmonella Gallinarum (S. Gallinarum) are the biovars of Salmonella enterica serovar Gallinarum that are responsible for pullorum disease and fowl typhoid, respectively, in poultry. Traditional serological methods fail to quickly differentiate between these biovars due to their identical O antigenic factors (O9 and O12). Although single nucleotide polymorphism (SNP)-based methods have been used to distinguish between the biovars, they often lack the required accuracy and effectiveness. In this study, we developed a PCR high resolution melt (PCR-HRM) assay, which targeted a SNP at position 665 of the fimH gene, for rapid differentiation between S. Pullorum and S. Gallinarum. Our method showed 100% specificity and was able to detect as little as 0.033 pg of S. Pullorum DNA and 0.027 pg of S. Gallinarum DNA. The PCR-HRM results for 547 clinical isolates were in complete agreement with traditional serological methods. This PCR-HRM assay significantly reduced identification time and provided high throughput, efficient testing. This makes it a practical and reliable tool for accurate differentiation between S. Pullorum and S. Gallinarum in clinical settings.