A real-time PCR for rapid identification of Salmonella enterica Gaminara serovar

Salmonella is one of the leading causes of foodborne illnesses in the USA. When a Salmonella outbreak occurs, rapid identification of the causative serovar is important for tracing the source of contamination and for preventing the further spread of the illness. Each serovar is characterized by the presence of a group-specific somatic O-antigen(s) and an assortment of flagellar phase-1 and phase-2 antigens. As the traditional serotyping protocol is time consuming, labor intensive, and expensive, faster and less expensive molecular diagnostic methods are needed. This report outlines the development of a rapid multiplex real-time PCR procedure that facilitates the identification of Salmonella serogroup I and the serovars of the group. Using Salmonella Gaminara serovar (O16:d:1,7) as an example, first the gene(s) responsible for expression of the somatic O antigen, O16, and the nucleotide sequence of the variable-region of genes encoding the flagellar phase-1 (d) and phase-2 (1,7) antigens were identified. Then, a multiplex real-time PCR was designed that incorporated primers and probes specific for the three target genes and confirmed the specificity. The assay had 100% inclusivity for all three gene targets, detecting 2 genomic DNA copies of O16 and 1,7 gene targets and 10 copies of d gene target. Importance: Rapid molecular methods to identify Salmonella serovars should increase the precision of routine surveillance of clinically important serovars and promote public health. •The real-time PCR assay detects the presence of Salmonella Gaminara in samples.•The lower limit of detection of ‘O16’ antigen by the PCR is 2 copies of the genome.•The lower limit of detection of phase-1 antigen ‘d’ is 10 copies of the genome.•The lower limit of detection of phase-2 antigen ‘1,7’ is 2 copies of the genome.•The real-time PCR assay is specific to Salmonella enterica Gaminara serovar.