|Reen, F. Jerry|
|Boyd, E. Fidelma|
Submitted to: Journal of Food Protection
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/25/2009
Publication Date: 11/10/2009
Citation: Mccarthy, N., Reen, F., Buckley, J.F., Frye, J.G., Boyd, E., Gilroy, D. 2009. Sensitive and rapid molecular detection assays for Salmonella enterica serovar Typhimurium and Heidelberg. Journal of Food Protection. 72(11):2350-2357. Interpretive Summary: Salmonella enterica is a major cause of gastroenteritis and is often transmitted through contaminated food. S. enterica serovars Typhimurium and Heidelberg are highly prevalent world wide, have broad host ranges, and are found in poultry, dairy animals and humans. Traditional methods used for the detection of S. enterica from contaminated food products such as dairy and meat are time consuming and labor intensive. We identified serovar-specific regions in S. enterica Typhimurium and Heidelberg genomes and designed a multiplex PCR (mPCR) assay to detect them. The assay was validated for its ability to detect and identify these serovars by screening a large set of S. enterica isolates. The mPCR assay was further tested to determine its ability to detect S. enterica isolates from cheese and meat. The mPCR assay was found to be highly sensitive and significantly reduced the time needed to identify S. enterica Typhimurium and Heidelberg in contaminated food. This rapid, selective and cost effective assay may be useful in the production of fresh and processed foods.
Technical Abstract: Salmonella enterica is a significant cause of gastroenteritis worldwide, with S. enterica serovars Typhimurium and Heidelberg being particularly prevalent. S. enterica serovars Typhimurium and Heidelberg have broad host ranges infecting poultry, dairy animals and humans. Traditional methods used for the detection of S. enterica from contaminated food products such as dairy and meat products are time consuming and labor intensive. The aim of this study was to develop a sensitive and rapid PCR based detection method with optimized specificity for high throughput screening of veterinary and clinical samples. We used bioinformatics to identify potential serovar-specific regions from the available S. enterica sequenced genomes and designed primer pairs to targeted regions unique to Typhimurium and Heidelberg serovars. A primer pair targeting a putative cytoplasmic protein STM4492 amplified a 759 bp product specific to Typhimurium only. In addition, a primer pair targeting a putative inner membrane protein STM2745 amplified a 199 bp product from both Typhimurium and Heidelberg isolates. We screened a large collection of clinical and environmental isolates validating the specificity of each primer set. Next, a multiplex PCR (mPCR) assay was optimized for identification and differentiation of S. enterica Typhimurium and Heidelberg. mPCR assays were developed to detect S. enterica isolates from both cheese and meat products. The reaction conditions for this mPCR have significantly reduced the time needed to identify S. enterica Typhimurium and Heidelberg, making this a rapid selective tool which will be useful to the food industry.