Journal of Postdoctoral Research www.postdocjournal.com PostDoc Journal Vol. 1, No. 2, February 2013 Rapid Detecion of Salmonella in Bovine Lymph Nodes Using a Commercial Real-Time PCR System Joshua Hill 1 , Thomas S. Edrington 1* , Guy H. Loneragan 2 , Sara E. Gragg 2 , David J. Nisbet 1 1 United States Department of Agriculture, Agricultural Research Service, Southern Plains Agricultural Research Center, Food and Feed Safety Research Unit, 2881 F&B Road, College Staion, Texas, 77845, USA 2 Internaional Center for Food Industry Excellence, Department of Animal and Food Sciences, Texas Tech University, Box 42141, Lubbock, Texas, 79409-2141, USA *Email: tom.edrington@ars.usda.gov Abstract Rapid Salmonella detecion is needed to help prevent the distribuion of contaminated food products. Using tradiional culture methods, Salmonella detecion can take up to 3-5 days. Using an improved protocol and a commercial real-ime PCR system, we have shortened the detecion ime to less than 24 hr with comparable sensiivity and speciicity to tradiional Salmonella culture methods. Keywords: Salmonella, lymph node, catle, detecion, real-ime PCR Introducion Salmonella infecion in livestock can be transmited to humans via associated products such as ground beef (Schneider et al. 2011) which may result in sporadic outbreaks of salmonellosis (Zansky et al. 2002). While generally a self-limiing illness, some cases require hospitalizaion or may even result in death (CDC 2011) with costs to society in the billions of dollars (Frenzen et al. 1999). Outbreaks of Salmonella infecion also result in product recalls adding directly to industry costs. In beef processing faciliies there has been great strides made to reduce both hide contaminaion (Serraino et al. 2012; Gill 2009; Carlson et al. 2008) and carcass contaminaion (Rekow et al. 2011) using post harvest intervenions. Recently, there has been concern regarding Salmonella contained in bovine lymph nodes at the ime of slaughter which can end up in ground beef. Iniial studies indicate that Salmonella contaminated lymph nodes occur at a prevalence of 0.35% to 3.86% (Brichta-Harhay et al. 2012; Arthur et al. 2008). To date no pre- or post-harvest soluion has been efecive in dealing with Salmonella contained in non-mesenteric lymph nodes. Tradiional bacteriology methods for detecing Salmonella can be ime consuming, requiring 3-5 days to complete and thus not suited to tesing highly perishable goods. (Fricker 1987). Real- ime PCR has been used successfully to detect Salmonella (Eyigor et al. 2002; Mainali et al. 2011; Suo et al. 2010) and is a relaively quick, robust method with many systems to choose from. We chose a commercially available plaform (Pall GeneDisc, Pall Corp., Port Washington, NY USA) as it allows high-throughput at 480 results every hour from 96 samples (Pall n.d.) and is validated to meet Title 21 CFR 11 for electronic documentaion (Pall n.d.). This system is based on real-ime PCR ampliicaion and detecion, using proprietary master mixes that contain speciic primer-probe combinaions. In this report, we used a commercial real-ime PCR system as a staring point and designed a shortened protocol that allowed us to reliably detect the presence of Salmonella from catle lymph nodes in less than 24hrs. This short turn- around ime to detect Salmonella as compared to tradiional culture methods may allow increased tesing of highly perishable goods, thereby helping food producion faciliies quickly detect contaminated goods. Materials and Methods Reagents The Pall GeneDisc system was used for all