Development of a real-time multiplex PCR assay for the detection of multiple <it>Salmonella </it>serotypes in chicken samples

• Main Authors: Whyte Paul, Duffy Geraldine, Barry Thomas, McGuinness Sheila, Burgess Catherine, McCabe Evonne, O'Regan Edel, Fanning Séamus
• Format: Article
• Language: English
• Published: BMC 2008-09-01
• Series: BMC Microbiology
• Online Access: http://www.biomedcentral.com/1471-2180/8/156

<p>Abstract</p> <p>Background</p> <p>A real-time multiplex PCR assay was developed for the detection of multiple <it>Salmonella </it>serotypes in chicken samples. Poultry-associated serotypes detected in the assay include Enteritidis, Gallinarum, Typhimurium, Kentucky and Dublin. The traditional cultural method according to EN ISO 6579:2002 for the detection of <it>Salmonella </it>in food was performed in parallel. The real-time PCR based method comprised a pre-enrichment step in Buffered Peptone Water (BPW) overnight, followed by a shortened selective enrichment in Rappaport Vasilliadis Soya Broth (RVS) for 6 hours and subsequent DNA extraction.</p> <p>Results</p> <p>The real-time multiplex PCR assay and traditional cultural method showed 100% inclusivity and 100% exclusivity on all strains tested. The real-time multiplex PCR assay was as sensitive as the traditional cultural method in detecting <it>Salmonella </it>in artificially contaminated chicken samples and correctly identified the serotype. Artificially contaminated chicken samples resulted in a detection limit of between 1 and 10 CFU per 25 g sample for both methods. A total of sixty-three naturally contaminated chicken samples were investigated by both methods and relative accuracy, relative sensitivity and relative specificity of the real-time PCR method were determined to be 89, 94 and 87%, respectively. Thirty cultures blind tested were correctly identified by the real-time multiplex PCR method.</p> <p>Conclusion</p> <p>Real-time PCR methodology can contribute to meet the need for rapid identification and detection methods in food testing laboratories.</p>