Development of Reverse Transcriptase-polymerase Chain Reaction of fimA Gene to Detect Viable Salmonella in Milk

ABSTRACT Rapid detection of viable Salmonella in pasteurized milk is important to protect public health from foodpoisoning. Reverse transcriptase-polymerase chain reaction(RT-PCR) is recognized as a molecular geneticalmethod to differentiate between live and dead bacteria. The RT-PCR in this study was designed to detect spe-cifically viable Salmonella in milk by using the primers whose nucleotide sequences were determined based on fimA gene which encodes the subunit of type 1 fimbriae. Treatment of RNA preparation with RNase-freeDNase was adequate enough to destroy DNA, which may otherwise be amplified in the RT-PCR. Sevenstrains of Salmonella were detected in the RT-PCR but Escherichia coli , Shigella sonnei , Citrobacter freundii ,and Klebsiella pneumoniae were not. 10 7 /m and 10 6 /m of dead Salmonella which were heat-treated in milkwere detectable by using the RT-PCR but 10 5 ~10/m of the dead bacteria were not. The sensitivity of theRT-PCR in detecting viable Salmonella was 100 cells/m .

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