eight-hour PCR-based technique for detection of Salmonella serovars in seafood

A rapid and sensitive 8-h PCR assay has been developed for detection of Salmonella serovars in seafood. A total of 110 fresh and raw seafood samples were analysed for the presence of Salmonella using different enrichment periods prior to PCR assay. Seafood samples included in this study were fish, shrimps, mussels, crabs, edible oysters, and clams, collected from local fish markets in Cochin (India). The assay was performed with a Salmonella-specific 284 bp invA gene amplicon. Specificity and sensitivity of the assay were ascertained with seafoods spiked with viable Salmonella cells to a level of 10⁶ to 2 CFU per 25 g. Detection efficiency of the assay increased with increasing enrichment period for seafood, and 33.6% of seafood samples were found positive for Salmonella by 8-h PCR assay. Detection limit for the 8-h PCR assay showed visible 284 bp amplicon from seafood homogenates spiked with 2 CFU per 25 g. Seafood samples spiked with different Salmonella serovars, namely Salmonella typhi, Salmonella typhimurium, Salmonella enteritidis, Salmonella mbandka, Salmonella bareilly, and Salmonella weltevreden, were detected, confirming this technique would be ideal for detection of the Salmonella serovars prevalent in seafood. This study also covered inhibition by the seafood matrix and the detection limit for dead Salmonella cells during the PCR assay. There was no visible inhibition of this Salmonella PCR assay by seafood matrices. The detection limit for dead Salmonella cells by 8-h PCR assay was 2 x 10³ CFU per 25 g seafood. The data indicated that dead cells of Salmonella in naturally contaminated seafood samples do not interfere with the assay resulting in false positives.