ARS | Publication request: Evaluation of a multiplex PCR system for simultaneous detection of Salmonella spp., Listeria monocytogenes, and Escherichia coli O157:H7 in foods and in food subjected to freezing

Submitted to: Foodborne Pathogens and Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: August 28, 2008
Publication Date: February 1, 2009
Citation: Kawasaski, S., Fratamico, P.M., Horikoshi, N., Okada, Y., Takeshita, K., Sameshima, T., Kawamoto, S. 2009. Evaluation of a multiplex PCR system for simultaneous detection of Salmonella spp., Listeria monocytogenes, and Escherichia coli O157:H7 in foods and in food subjected to freezing. Foodborne Pathogens & Disease. 6(1):81-89.

Interpretive Summary: Food-borne illness caused by bacteria known as Escherichia coli O157:H7, Salmonella species, and Listeria monocytogenes, is a major public health concern worldwide. The availability of rapid and sensitive methods for detecting these pathogenic bacteria in different foods, food ingredients, and food processing plants is critical to prevent the release of contaminated foods to the consumer. Furthermore, methods with the capability to detect the three pathogens simultaneously in food offer the advantages of savings in time and cost. A multiplex polymerase chain reaction (PCR) method was developed and compared to time consuming and laborious traditional methods for detection of the three pathogens simultaneously in different types of spiked or naturally contaminated food samples. The multiplex PCR assay was able to detect all three pathogens when found in food at less than or equal to 5 bacteria per 25 grams of food. In addition, the multiplex PCR assay showed higher sensitivity for detection of S. Enteriditis, L. monocytogenes, and E. coli O157:H7 in food samples stored frozen for 2 weeks or 2 months compared to the traditional methods. The multiplex PCR assay is a valuable method for the food industry and regulatory agencies for simultaneous rapid screening for Salmonella species, Listeria monocytogenes, and E. coli O157:H7 in food, even after frozen storage.

Technical Abstract: Conventional culture methods were compared to a multiplex PCR assay for simultaneous detection of Salmonella spp., Listeria monocytogenes, and Escherichia coli O157:H7 from enrichment cultures of various types of artificially inoculated and naturally contaminated foods. The multiplex PCR assay was evaluated in 44 types of spiked food samples, including meat, produce, fish, and dairy products targeting genes specific for each pathogen for simultaneous detection. The sensitivity of the assay was less than or equal to 5 CFU per 25g of inoculated sample after 20 h of enrichment. The PCR assay was also evaluated in inoculated food samples stored at -20 degree C for 2 weeks or 2 months. Out of 28 food samples tested, 27, 27, and 26 samples were positive for S. Enteriditis, L. monocytogenes, and E. coli O157:H7, respectively, using the multiplex PCR assay, whereas only 13, 26, and 20 samples were positive, respectively, using the culture method after 2 weeks of storage at -20 degree C. Similar results were obtained for samples stored at -20 deg C for 2 months. The multiplex PCR assay was capable of detecting the three pathogens at an initial inoculum level of 5 CFU per 25g of various types of food, and the detection rate of the PCR assay was higher than that of conventional culture methods. As a result, the multiplex PCR assay is a valuable method for simultaneous rapid screening for the three pathogens in food, even after frozen storage.