Digoxigenin-labeled probe-based colony blotting assay for rapid quantification of salmonella serovars in seafood and water

Abstract

A non-radio-labeled probe-based detection method was developed for rapid enumeration of Salmonella in seafood and water samples. A Salmonellaspecific invA gene probe was developed using a digoxigenin-based non-radio labeling assay, which was evaluated with naturally contaminated seafood and water samples. The probe-based technique was further compared with the quantitative PCR assay. The method was specific for detection of different Salmonella serovars without any nonspecific hybridization with other Salmonellarelated Enterobacteriaceae. The optimum labeling efficiency was determined for the labeled probe, and 10 pg/μL probe concentration was observed to be most efficient for detection of Salmonella colonies on nylon membrane. Quantification of Salmonella in naturally contaminated seafood and water samples (n = 21) was in the range 10-102 CFU/mL. The assay successfully quantified Salmonella in spiked seafood and water samples in the presence of background flora, and the entire assay was completed within 48 h. The probe-based assay was further evaluated with real-time PCR, and results showed that the assay was comparable to real-time PCR assay. Thus, this probe-based assay can be a rapid, useful, and alternative technique for quantitative detection of Salmonella in food, feed, and water samples.