Rapid Serotyping of Salmonella Isolates Based on Single Nucleotide Polymorphism-Like Sequence Profiles of a Salmonella-Specific Gene.

ABSTRACT

Although serotyping is the most important method of identification of taxonomy in Salmonella, conventional serotype determination with a complete set of antisera is time consuming and laborious. Recently, rapid serotyping procedures with polymerase chain reaction (PCR) have been developed. In this study, we established a novel PCR-based rapid serotyping method that employs a unique target gene. Alignment study of Salmonella-specific gene (Salmonella enterotoxin [stn]) revealed a correlation between the stn gene sequence and the serotype of the organism. In 750 bp of stn gene, 55 nucleotides indicated single nucleotide polymorphism (SNP)-like polymorphism, and the correlation between the SNP-like polymorphism and the serotype of the organism suggests that SNP-like sequences in stn gene can serve as an index for serotyping. To develop a rapid serotyping method based on the SNP-like polymorphism, we selected serotype-associated 12 SNP-like sites in the stn gene and established a method based on high-resolution melting (HRM) and PCR, which identifies nucleotides at SNP-like sites within 1.5 h. This newly established rapid serotyping procedure (stn-HRM) could identify nine serotypes, including the frequently isolated serovar Enteritidis. These nine serotypes cover 64.3% of cases of Salmonella, as reported by the World Health Organization/Global Foodborne Infection Network (WHO/GFN) Country Databank from 2001 to 2010. In this study, we employed a unique target gene, stn, which is completely independent of the genes that were targeted in previously reported rapid serotyping procedures. Therefore, the results obtained by our newly developed stn-HRM procedure are independent of the results obtained by other procedures. Besides, stn-HRM can ensure accurate identification of the bacterial species as stn is a Salmonella-specific gene. It is expected that the combination of newly constructed stn-HRM and previously reported procedures could further improve the credibility of Salmonella isolate serotyping.