Multiplex PCR for Detection of Staphylococcus aureus and Listeria monocytogenes in Ready-to-eat Foods
Keywords:Multiplex PCR, Ready-to-eat Foods, Staphylococcus aureus, Listeria monocytogenes
Foodborne diseases are common problems worldwide. Staphylococcus aureus is one of the most important food-borne pathogens. Listeria monocytogenes is widely found in contaminated foods, especially in refrigerated and ready-to-eat foods. Rapid detection and identification are needed to prevent and control the food contamination by these infectious microorganisms. For the objective of the research, multiplex PCR technique has been developed for rapid detection of S. aureus and L. monocytogenes in ready-to-eat foods. In this study, two-pair of primers were designed within conserved regions of the virulence genes, coa gene of S. aureus and prfA gene of L. monocytogenes and then were used for detection of those bacteria. The results showed that this multiplex PCR could detect at least 1 ng of S. aureus DNA and 150 pg of L. monocytogenes DNA. Investigation into the artificially contaminated foods, this multiplex PCR was able to detect less than 104 cells/g of S. aureus and 1 cell/g of L. monocytogenes in foods. In addition, there were no amplifications of nucleic acids from other food related-pathogens, indicating the specificity of this test. Detections in thirty ready-to-eat food samples from local markets in Chonburi province, Thailand, showed that none of them were contaminated with S. aureus and L. monocytogenes. Therefore, this finding indicated good hygiene in production of ready-to-eat foods in these areas. Consequently, this multiplex PCR can be further developed and employed for monitoring of S. aureus and L. monocytogenes in contaminated foods.
Akineden, O., Annemuller, C., Hassan, A.A., Lammler, C., Wolter, W., & Zschock, M. (2001). Toxin genes and other characteristics of Staphylococcus aureus isolates from milk of cows with mastitis. Clin Diagn Lab Immunol., 8(5), 959-964. https://doi.org/10.1128/CDLI.8.5.959-964.2001
Alarcon, B., Vicedo, B., & Aznar, R. (2006). PCR-based procedures for detection and quantification of Staphylococcus aureus and their application in food. J Appl Microbiol., 100(2), 352-364. https://doi.org/10.1111/j.1365-2672.2005.02768.x
Ananchaipattana, C., Hosotani, Y., Kawasaki, S., Pongsawat, S., Latiful, B.M., Isobe, S., & Inatsu, Y. (2012). Prevalence of foodborne pathogens in retailed foods in Thailand. Foodborne Pathog Dis., 9(9), 835-840. https://doi.org/10.1089/fpd.2012.1169
Aznar, R., & Alarcon, B. (2003). PCR detection of Listeria monocytogenes: a study of multiple factors affecting sensitivity. J Appl Microbiol., 95(5), 958-966.
Barocci, S., Mancini, A., Canovari, B., Petrelli, E., SbrisciaFioretti, E., Licci, A., . . . Briscolini, S. (2015). Listeria monocytogenes meningitis in an immunocompromised patient. New Microbiol., 38(1), 113-118.
Chen, Y.I., Burall, L.S., Macarisin, D., Pouillot, R., Strain, E., AJ, D.E.J., . . . Datta, A.R. (2016). Prevalence and level of Listeria monocytogenes in ice cream linked to a Listeriosis outbreak in the United States. J Food Prot., 79(11), 1828-1832. https://doi.org/10.4315/0362-028X.JFP-16-208
D'Amico, D.J., & Donnelly, C.W. (2008). Enhanced detection of Listeria spp. in farmstead cheese processing environments through dual primary enrichment, PCR, and molecular subtyping. J Food Prot., 71(11), 2239- 2248.
D'Amico, D.J., & Donnelly, C.W. (2009). Detection, isolation, and incidence of Listeria spp. in small-scale artisan cheese processing facilities: a methods comparison. J Food Prot., 72(12), 2499-2507.
Garner, D., & Kathariou, S. (2016). Fresh produce-associated Listeriosis outbreaks, sources of concern, teachable moments, and insights. J Food Prot., 79(2), 337-344. https://doi.org/10.4315/0362-028X.JFP-15-387
Kearns, A.M., Seiders, P.R., Wheeler, J., Freeman, R., & Steward, M. (1999). Rapid detection of methicillinresistant staphylococci by multiplex PCR. J Hosp Infect., 43(1), 33-37. https://doi.org/10.1053/jhin.1999.0631
Kumar, T.D., Murali, H.S., & Batra, H.V. (2009). Simultaneous detection of pathogenic B. cereus, S. aureus and L. monocytogenes by multiplex PCR. Indian J Microbiol., 49(3), 283-289. https://doi.org/10.1007/s12088-009-0032-y
Laaksonen, S., Oksanen, A., Julmi, J., Zweifel, C., FredrikssonAhomaa, M., & Stephan, R. (2017). Presence of foodborne pathogens, extended-spectrum betalactamase-producing Enterobacteriaceae, and methicillin-resistant Staphylococcus aureus in slaughtered reindeer in northern Finland and Norway. Acta Vet Scand., 59(1), 2. https://doi.org/10.1186/s13028-016-0272-x
Li, B., Chen, F., Wang, X., & Shao, Y. (2008). Detection of Staphylococcus aureus, Shigella spp., Salmonella spp. in food by multiplex PCR. Wei Sheng Yan Jiu., 37(4), 438-442.
Mercado, M., Avila, J., Rey, M., Montoya, M., Carrascal, A.K., & Correa, D.X. (2012). [Outbreaks of Salmonella spp., Staphylococcus aureus and Listeria monocytogenes associated with poultry consumption. Systematic review]. Biomedica, 32(3), 375-385. https://doi.org/10.1590/S0120-41572012000300008
Paudyal, N., Anihouvi, V., Hounhouigan, J., Matsheka, M. I., Sekwati-Monang, B., Amoa-Awua, W., . . . Fang, W. (2017). Prevalence of foodborne pathogens in food from selected African countries - A meta-analysis. Int J Food Microbiol., 249, 35-43. https://doi.org/10.1016/j.ijfoodmicro.2017.03.002
Schmid-Hempel, P., & Frank, S.A. (2007). Pathogenesis, virulence, and infective dose. PLoS Pathog., 3(10), 1372- 1373. https://doi.org/10.1371/journal.ppat.0030147
Thevenot, D., Dernburg, A., & Vernozy-Rozand, C. (2006). An updated review of Listeria monocytogenes in the pork meat industry and its products. J Appl Microbiol., 101(1), 7-17. https://doi.org/10.1111/j.1365-2672.2006.02962.x
Wilson, I.G. (1995). Occurrence of Listeria species in ready to eat foods. Epidemiol Infect., 115(3), 519-526.
Wilson, I.G., Cooper, J.E., & Gilmour, A. (1994). Some factors inhibiting amplification of the Staphylococcus aureus enterotoxin C1 gene (sec+) by PCR. Int J Food Microbiol., 22(1), 55-62.
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