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ARS Home » Northeast Area » Wyndmoor, Pennsylvania » Eastern Regional Research Center » Molecular Characterization of Foodborne Pathogens Research » Research » Publications at this Location » Publication #290625


Location: Molecular Characterization of Foodborne Pathogens Research

Title: Simultaneous detection of Salmonella spp., Escherichia coli O157 and Listeria monocytogenes in a variety of cheeses and spinach using a multiplex real-time PCR method

item Sathyamoorthy, Venugopal - U.s. Food & Drug Administration (FDA)
item Datta, Atin - U.s. Food & Drug Administration (FDA)
item Trach, Larisa - U.s. Food & Drug Administration (FDA)
item He, Yiping
item Tall, Ben - U.s. Food & Drug Administration (FDA)
item Mccardell, Barbara - U.s. Food & Drug Administration (FDA)

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 7/29/2013
Publication Date: N/A
Citation: N/A

Interpretive Summary:

Technical Abstract: Introduction: Foodborne diseases affect about 48 million people per year in US. A major portion of these foodborne illnesses are caused by 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 circulation, thereby reducing the incidence of food borne diseases. At present there is no method available for the simultaneous detection of all three organisms in contaminated foods regulated by FDA. Purpose: This project aims to evaluate, optimize and adapt a real-time PCR method, originally developed at USDA, to simultaneously detect the presence of Salmonella spp, Escherichia coli O157:H7 and Listeria monocytogenes in soft-cheese and spinach. Methods: Five different cheeses and spinach in a previously described selective medium were spiked with 5-25 CFU/25g of Salmonella, E.coli O157:H7 and L. monocytogenes, stomached, and incubated for 2 h at 37°C followed by the addition of nalidixic acid, fosfomycin, cycloheximide, and acriflavine, and then grown overnight. The samples were then used for the extraction of genomic DNA using a DNA extraction kit (Qiagen). The DNAs were used to carry out the real-time PCR with primers and TaqMan probes targeting invA (Salmonella), rfbE (E. coli O157) and hlyA (L. monocytogenes), as well as an internal amplification control (IAC). Results: All three gene targets of the tested pathogens were detected in the spiked cheeses and spinach samples after enrichment, with a sensitivity level of 5-25 CFU/25g. As expected, the gene targets were not detectable in control samples, except that the IAC was positive. Significance: The availability and optimization of this method for the simultaneous detection of Salmonella spp., E. coli O157, and Listeria monocytogenes in different cheeses and spinach will allow the FDA and other organizations to take action and prevent spread of an outbreak.