Submitted to: American Society for Microbiology
Publication Type: Abstract Only
Publication Acceptance Date: May 2, 2003
Publication Date: May 2, 2003
Citation: VAN KESSEL, J.S., KARNS, J.S., PERDUE, M.L. USING PORTABLE REAL-TIME POLYMERASE CHAIN REACTION (RT-PCR) ASSAYS TO DETECT SALMONELLA SPP. IN RAW MILK. AMERICAN SOCIETY FOR MICROBIOLOGY. 2003. P-068. Technical Abstract: Traditional culture methods for detection of pathogens in foods are laboratory based, and generally time-consuming and labor intensive. Recent advancements in PCR technology may allow for more rapid, and perhaps even on-farm detection, of food-borne pathogens. The purpose of this study was to determine the efficacy of a portable RT-PCR system for the detection of Salmonella spp. in raw milk. The bulk milk samples (200) chosen for this study were a subset of a larger study; the subset contained 24 samples that were culture-positive for Salmonella and 176 that were culture-negative. Milk was both plated directly on selective agar (XLT4) and enriched in selective media (tetrathionate broth) followed by plating. Presumptive Salmonella were isolated from direct culture of five samples while Salmonella was isolated from the remaining 19 positive samples only after enrichment. Serotyping was performed on colonies that were identified phenotypically as Salmonella and 22 samples were confirmed as Salmonella. PCR assays on culture-positive milk prior to enrichment yielded no evidence of Salmonella. DNA extracts of bacterial pellets from the enriched samples were analyzed for the presence or absence of Salmonella by RT-PCR using the Ruggedized Advanced Pathogen Identification Device (R.A.P.I.D.; Idaho Technology Inc.). Fifty-three samples were identified as Salmonella-positive from the enrichment pellets based on RT-PCR analysis. Two samples that were identified as being positive for Salmonella via culture and serotyping were identified as being Salmonella-negative based on RT-PCR. Serotyping identified isolates from these samples as Salmonella Montevideo. There were 32 samples that were Salmonella-negative based on culture and Salmonella-positive based on RT-PCR. Based on these results, RT-PCR may be more sensitive and discriminatory than traditional culture methods for the detection of Salmonella spp. in raw milk. In addition, enrichment followed by RT-PCR yields results in 24 h as opposed to 48 to 72 h for traditional culture.