Development of an Assay to Detect Salmonella enterica Serovar Senftenberg by qPCR

Introduction: Salmonella contamination of food is a public health concern because of the potential for widespread outbreaks and the gastroenteritis it can cause.  Salmonella Senftenberg is a serotype of Salmonella that is often detected in humans, and, although it is not listed among the top 20 serotypes implicated in human illnesses, it is ranked in the top ten among cases of non-human diseases.  The zoonotic nature of Salmonella suggests a potential for this serotype to become significant in human disease.  Development of assays to detect S. Senftenberg and other exotic serotypes would help define treatment and prevention methods.

Purpose: The aim of this study was to sequence, design, test, and manufacture an assay that detects S. Senftenberg while avoiding the detection of other similar serotypes or species.

Methods: Twenty samples presumptive of S. Senftenberg were sequenced using the Ion PGM™ Sequencer.  After genome assembly, a bioinformatics analysis was performed to identify conserved regions that are unique to S. Senftenberg and exhibit low similarity to other Salmonella.  Candidate qPCR assays were designed using a proprietary bioinformatics tool and evaluated against all available GenBank sequences.  The specificity of the assays was tested using an exclusion panel comprised of various serotypes of Salmonella and related pathogens, and was determined to be specific for S. Senftenberg.  The LOD is estimated to be 10 CFU/reaction.

Results: Sequences were obtained in three days, and genome assembly and assay design was completed in five days.  The assay was tested against an exclusion panel and found to be specific for Senftenberg.  Sensitivity was estimated at 10 CFU/reaction.

Significance: Our S. Senftenberg real-time PCR assay is highly specific and is able to distinguish Senftenberg from other Salmonella serotypes.  Identification and subsequent control of S. Senftenberg during food production and processing may help reduce the frequency of food-related illnesses.