Registration and Identification of Toxic S. aureus genes Isolated from Tilapia Fish Using Multiplex PCR Technique

Subject Areas : Microbiology

N.  Niakhalili ¹ (M. Sc. Student of the Department of Food Science and Technology, Science and Research Branch, Islamic Azad University, Tehran, Iran.)
H.  Ahari ² (Professor of the Department of Food Science and Technology, Science and Research Branch, Islamic Azad University, Tehran, Iran.)
B.  Nowruzi ³ (Assistant Professor of the Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran.)

Received: 2022-07-21 Accepted : 2022-09-19 Published : 2023-03-21

Keywords: Staphylococcus aureus, Tilapia, Multiplex PCR, Gene,

Abstract :

 Introduction: Aquatic animal products are considered very important food items in the food basket regarding their high calorie, protein, and omega-3 unsaturated fat. Many aquatic animals, including fish, may always be infected with pathogenic microorganisms at various stages, from hunting to purchasing. Eventually, these factors would cause severe poisoning in humans. The present study investigates the registration and identification of Staphylococcus aureus’s toxic genes isolated from tilapia using the Multiplex PCR technique.Materials and Methods: The sample size included 42 subjects (21 fresh fish samples and 21 frozen fish samples). First, after preparing the samples, Staphylococcus aureus was isolated and examined as surface culture in Baird-Parker medium. Next, a Coagulase test was performed, and the enterotoxin gene was identified by DNA extraction. Finally, gene sequencing was carried out automatically and systematically.Results: The results suggested that 92.9% of the total samples (21 fresh tilapia and 18 frozen fish) had no Staphylococcus aureus, and 7.1% of the samples were infected with Staphylococcus aureus. The coagulase test results also suggested that all three frozen fish samples were coagulase positive. Examining enterotoxin-producing genes (SEA, SEB, SEC, SED, SEE, and SEG) among three frozen samples infected with Staphylococcus bacteria revealed SEA enterotoxin gene only in one frozen sample. Conclusion: The results showed that due to the low cost and much shorter time required to identify toxic Staphylococcus aureus genes using Multiplex PCR, this method is highly effective in studying the genotypes of Staphylococcus isolates. Therefore, by using this method, one can improve the food health level in the society.

References:

• Arfatahery, N., Davoodabadi, A. & Abedimohtaseb, T. (2016). Characterization of toxin Genes and antimicrobial susceptibility of staphylococcus aureus isolates in fishery products in iran. Science Reports, 6, 34216,1-7.


• Ayulo, A.M.R., Machado, R.A. & Scussel, V.M. (1994). Enterotoxigenic Escherichia coli and Staphylococcus aureus in fish and seafood from the southern region of Brazil. International Journal of Food Microbiology, 24(1-2), 171-178.


• Arslan, S. & Ȍzdemir, F. (2017). Molecular characterization and detection of enterotoxins, methicillin resistance genes and antimicrobial resistance of Staphylococcus aureus from fish and ground beef. Polish Journal of Veterinary, 20, 1, 85-94.


• Argudín, M.Á., Mendoza, M.C. & Rodicio, M.R. (2010). Food poisoning and Staphylococcus aureus enterotoxins. Toxins, 2(7), 1751-1773.


• Adib Far, P. (2014). Medical Microbiology. Noor Danesh Publications, 63-74. [In Persian]


• Azizkhani, M. & Turian, F. (2019). Evaluation of the frequency of Staphylococcus aureus enterotoxinogenic genes isolated from minced meat samples in foodstores in Mazandaran province. Journal of Veterinary Research, 74 (1), 65-72. [In Persian]


• Asgarpour, D. & Zeyghami, H. (2015). The role of Staphylococcus aureus enterotoxins (SEs) in staphylococcal food poisoning: A systematic review article. Quarterly Journal of Laboratory and Diagnosis, 28, 63-73. [In Persian]


• Dinges, M.M., Orwin, P.M. Schlievert, P.M. (2000). Exotoxins of Staphylococcus aureus. Clinical Microbiology Reviews, 13(1), 16-34.


• Ertas Onmaz, N., Abay, S., Karadal, F., Hizlisoy, H., Telli, N. & Al, S. (2014). Occurrence and antimicrobial resistance of staphylococcus aureus and salmonella spp in retail fish sample in turkey. Marine Pollution Bulletin (Elsevier), 1-5.


• El-Sayed, A.F.M. (2006). Tilapia culture in salt water: environmental requirements, nutritional implications and economic potentials. Avances en Nutricion Acuicola.


• Giannatale, E.D., Prencipe, V., Tonelli, A. & Marfoglia, C. (2011). Characterization of Staphylococcus aureus trains isolated from food human consumption. Veterinaria Italiana, 47(2), 165- 173.


• Gencay, Y.E., Ayaz, N.D. & Dogru, A.K. (2010). Enterotoxin Gene Profiles of Staphylococcus aureus and Other Staphylococcal Isolates from Various Foods and Food Ingredients. Journal of Faculty of Veterinary Medicine Erciyes University, 7(2), 75-80.


• Holah, J.T., Higgs, C., Robinson, S., Worthington, D. & Spenceley, H. (1990). A conductance‐based surface disinfection test for food hygiene. Letters in Applied Microbiology, 11(5), 255-259.


• Jawetz, E., Melnik, J. L., Adelberg, E., Brooks, G., Bottle, J. & Stephen, M. (2008) Medical Microbiology. Translated by: Rahimi, Mohammad Karim Vatahri, Amid, Ayizh Publications, Tehran, Iran, 269. [In Persian].


• Ishraqi, S., Salehipour, Z., Pourmand, M.R., Rahimi Darushani, A., Salehi, Z., Taghi, M. & Aghamiri, S. (2009). Evaluation of frequency distribution of tst gene with entC, ent A, ent A/C genes in Staphylococcus aureus isolates isolated from different food products. Journal of Tehran University of Medical Sciences, 67(7), 470-476. [In Persian]


• Isna-ashari, M., Shayeq, J. & Nasrollahi, I. (2012). A study on prevalence of common enterotoxin genes of Staphylococcus aureus isolated from buffalo milk in Tabriz province using Multiplex PCR. Journal of Food Hygiene in Tabriz, 2, (2), 61-68. [In Persian]


• Kitai, S., Shimizu, A., Kawano, J., Sato, E., Nakano, C., Kitagawa, H., Fujio, K., Matsumura, K., Yasuda, R. & Inamoto, T. (2005). Prevalence and characterization of Staphylococcus aureus and enterotoxigenic Staphylococcus aureus in retail raw chicken meat throughout Japan. Journal of Veterinary Medical Science, 67(3), 269-274.


• Kluytmans, J.A.J.W. & Wertheim, H.F.L. (2005). Nasal carriage of Staphylococcus aureus and prevention of nosocomial infections. Infection, 33(1), 3-8.


• Loir, Y.L., Baron, F. & Guatier, M. (2003). Staphylococcus aureus and food poisoning. Genetics and Molecular Research, 2(1), 63-76.


• Nakane, A. (2010). Protective effect of glutathione Stransferase-fused mutant staphylococcal enterotoxin C against Staphylococcus aureus-induced bovine mastitis. Vet Immunol Immunopathology, 135, 64-70.


• Normanno, G., La Salandra, G., Dambrosio, A., Quaglia, N.C., Corrente, M., Parisi, A., Santagada, G., Firinu, A., Crisetti, E. & Celano, G.V. (2007). Occurrence, characterization and antimicrobial resistance of enterotoxigenic Staphylococcus aureus isolated from meat and dairy products. International Journal of Food Microbiology, 115(3), 290-296.


• Pelisser, M.R., Klein, C.S., Ascoli, K.R., Zotti, T.R. & Arisi, A.C.M. (2009). Ocurrence of Staphylococcus aureus and multiplex PCR detection of classic enterotoxin genes in cheese and meat products. Brazilian Journal of Microbiology, 40(1), 145-148.


• Pepe, O., Blaiotta, G., Bucci, F., Anastasio, M., Aponte, M. & Villani, F. (2006). Staphylococcus aureus and Staphylococcal Enterotoxin A in Breaded Chicken Products: Detection and Behavior during the Cooking Process. Applied and Environmental Microbiology, 72(11), 7057–7062.


• Redmond, E.C. & Griffith, C.J. (2003). Consumer food handling in the home: a review of food safety studies. Journal of food protection, 66(1), 130-161.


• Rahimi, E., Mommtaz, H., Shakerian, A. & Kavyani, H.R. (2012). The detection of classical enterotoxins of Staphylococcus aureus in raw cow milk using the ELISA method. Turkish Journal of Veterinary and Animal Sciences, 36(3), 319-322.


• Sanjeev, S. (2003). Bacteria of public health significance in seafood.


• Sokari, T. (1991). Distribution of enterotoxigenic Staphylococcus aureus in ready-to-eat foods in eastern Nigeria. International Journal of Food Microbiology, 12(2-3), 275-279.


• Tamasoki, M. S., Pahlavani, S., Pouria, M. & Shah Mahmoudi, S. (2011). Overview of tilapia breeding in the world, Regional Symposium on Applied Fisheries and Environmental Studies, Kermanshah, https://civilica.com/doc/159702. [In Persian]