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P2-18 An Improved Method to Simultaneously Detect Salmonella Enteritidis, Escherichia coli O157 and Listeria monocytogenes in Ground Black Pepper Using Multiplex Real-time PCR

Monday, July 27, 2015
Exhibit Hall (Oregon Convention Center)
Venugopal Sathyamoorthy , U.S. Food and Drug Administration , Laurel , MD
Natalia Noroozi , FDAU.S. Food and Drug Administration , Laurel , MD
Eunkyeong Park , FDA , Laurel , MD
Yunsoo Kim , FDAU.S. Food and Drug Administration, , Laurel , MD
Ben D. Tall , U.S. Food and Drug Administration , Laurel , MD
Yiping He , U.S. Department of Agriculture-ARS-ERRC , Wyndmoor , PA
Seongeun Hwang , U.S. Food and Drug Administration , Laurel , MD
Larisa Trach , U.S. Food and Drug Administration , Laurel , MD
Hannah Chase , U.S. Food and Drug Administration , Laurel , MD
Barbara McCardell , U.S. Food and Drug Administration , Laurel , MD
Atin Datta , U.S. Food and Drug Administration-CFSAN , Laurel , MD
Introduction: The three common foodborne pathogens implicated in foodborne outbreaks are Salmonella spp., Escherichia coli O157:H7 and Listeria monocytogenes. Hence, it is important to identify these pathogens in contaminated foods so that they can be eliminated from the marketplace. At present, there is no good method available for the simultaneous detection of these organisms in foods regulated by FDA. In general, detecting pathogens in spices poses challenges due to the anti-microbial activity of these foods.

Purpose: To optimize, and adapt a multiplex real-time PCR method, originally developed for meat samples at the USDA, to the simultaneous detection of Salmonella spp., E. coli O157 and L. monocytogenes in ground black pepper.

Methods: Ground black pepper was spiked with 5-5000 CFU/2 g of Salmonella Enteritidis, E. coli O157:H7 and L. monocytogenes, stomached and incubated for 2 hours at 37°C in BLEB broth, followed by the addition of four antibiotics and grown overnight. The enriched culture was subjected to the extraction of genomic DNA (Qiagen) to carry out the real-time PCR with primers and TaqMan probes specifically targeting invA (Salmonella), rfbE (E. coli O157), hlyA (L. monocytogenes), and an internal amplification control.

Results: All three gene targets of the tested pathogens were detected in the spiked ground black pepper with a sensitivity of 15-25 CFU/2 g in the presence of 2% corn oil. In the absence of corn oil, the sensitivity of detection was 15-25 CFU/2 g for Salmonella spp., and E. coli O157:H7 whereas it was ~900 CFU/2 g for Listeria monocytogenes.

Significance: This method can be applied to the effective detection of Salmonella Enteritidis, E. coli O157 and Listeria monocytogenes in the outbreaks associated with ground black pepper or other foods. Therefore, it would allow FDA to take rapid action and prevent the spread of the outbreak.

Back to: Poster Session 2 – Laboratory and Detection Methods; Meat, Poultry and Eggs; Dairy and Beverages; Food Law and Regulation; General Microbiology; Modeling and Risk Assessment
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