Detection and Determination of the Frequency of Listeria monocytogenes in Food samples by Molecular analysis of the hlyA gene

Subject Areas : Microbiology

S. Ashrafi ¹ , N. Bahmani ² , F. Keshavarzi ³

1 - MSc Graduated of Microbiology, Department of Biology, Sanandaj Branch, Islamic Azad University, Sanandaj, Iran.
2 - Assistant Professor of the Zoonoses Research Center, Research Institute for Health Development, Kurdistan University of Medical Sciences, Sanandaj, Iran.
3 - Associate Professor of the Department of Biology, Sanandaj Branch, Islamic Azad University, Sanandaj, Iran.

Received: 2023-04-10 Accepted : 2023-07-08 Published : 2023-09-23

Keywords: hlyA gene, Listeria monocytogenes, Food sample,

Abstract :

Introduction: Listeriolysin (LLO) is a sulfhydryl-activated hemolysin bound to cholesterol, that is encoded by the hlyA gene of Listeria monocytogenes. The aim of this study was to detect and determine the frequency of Listeria monocytogenes in food samples through molecular analysis of the hlyA gene.Materials and Methods: The present study was a descriptive cross-sectional assay conducted from the middle of October 2018 to the end of January of the same year. One hundred sixty-five samples of different types of meat, vegetables, hamburgers and shrimps were examined. Samples were randomly collected from restaurants, delicatessens, prepared food places, and different supermarkets. All samples were analyzed with phenotypic and biochemical tests. The samples suspected of Listeria monocytogenes, after DNA extraction, were evaluated by PCR with specific primers for the hlyA gene. Then the data and results were evaluated with using SPSS v.23 software and Excel software.Results: Phenotypic methods (microbial culture and biochemical investigation) confirmed 18 positive samples for Listeria monocytogenes, of which 10 (6.06%) samples were positive for Listeria monocytogenes by PCR of the hlyA gene. The rate of contamination in pounded kebab was 37.9%, chicken meat 4.4%, local hamburger 16.66% and lettuce 13.33%. Compared to other studies, these findings were more prevalent in some foods (e.g., local hamburger) and less in others (e.g., chicken meat).Conclusion: The use of biochemical methods for determining the frequency of Listeria monocytogenes in food samples in Sanandaj had a high percentage of false positives.

References:

• Almeida, P.F. & Almeida, R.C.C. (2000). A PCR protocol using inl gene as a target for specific detection of Listeria monocytogenes. Food Control, 11(2), 97-10. https://doi.org/10.1016/s0956-7135(99)00067-5 

 

• (2016). European Centre for Disease Prevention and Control (ECDC): The European Union summary report on trends and sources of zoonoses, zoonotic agents, and food-borne outbreaks in 2015. EFSA Journal, 14. https://doi.org/10.2903/j.efsa.2016.4634

 

• Akhondzadeh Basati, A., Misaghi, A., Salehi Zahraei, T. & Kamkar, A. (2006). Bacterial pathogens in fresh smoked and salted Irainian fish. Food Control, 17(3), 183-188. https://doi.org/10.1016/j.foodcont.2004.10.001 

 

• Baharvand, R. (2015). A gene in Listeria monocytogenes isolated from food. The Journal of Qazvin University of Medical Sciences, 19 (1), 24-31. [In Persian]

 

• Bierne, H., Sabet, C., Personnic, N. & Cossart, P. (2007). Internalins: a complex family of leucine-richrepeat-containing proteins in Listeria monocytogenes. Microbes and Infection, 9, 1156-66. https://doi.org/10.1016/j.micinf.2007.05.003 

 

• Borović, B.B., Baltić, T., Lakićević, B., Janković, V., Mitrović, R. Jovanović, J. & Lilić S. (2014). Prevalence of Listeria monocytogenes in ready–to–eat food of animal origin. Meat Technology, 55, 117–122. https://doi.org/10.5937/tehmesa1402117b 

 

• Carpentier, B. & Cerf, O. (2011). Review — Persistence of Listeria monocytogenes in food industry equipment and premises. International Journal of Food Microbiology, 145, 1-8. https://doi.org/10.1016/j.ijfoodmicro.2011.01.005 

 

• Chaitiemwong, N., Hazeleger, W.C., Beumer, R.R. & Zwietering, M.H. (2014). Quantification of transfer of Listeria monocytogenes between cooked ham and slicing machine surface. Food Control, 44, 177–184. https://doi.org/10.1016/j.foodcont.2014.03.056 

 

 

• Choi, W.S. & Hong, C. (2003). Rapid enumeration of Listeria monocytogenes in milk using competitive PCR. International Journal of Food Microbiology, 84, 79– 85. https://doi.org/10.1016/s0168-1605(02)00401-4 

 

• Di Pinto, A., Novello, L., Montemurro, F., Bonerba, E. & Tantillo G. (2010). Occurrence of Listeria monocytogenes in ready to eat foods from supermarkets in southern Italy. New Microbiology, 33, 249–252.

 

 

• Esho, F. K., Enkhtuya, B., Kusumoto, A. & Kawamoto, K. (2013). Microbial Assessment and Prevalence of Foodborne Pathogens in Natural Cheeses in Japan. BioMed Research International, 20580, 1–6. https://doi.org/10.1155/2013/205801 

 

 

 

• Freitag, N.E., Rong, L. & Portnoy, D.A. (1993). Regulation of the prfA transcriptional activator of Listeria monocytogenes: multiple promoter elements contribute to intracellular growth and cell-to-cell spread. Infect Immune, 61 (6), 2537.
  https://doi.org/10.1128/iai.61.6.2537-2544.1993 

 

• Gamboa-Marín A., Buitrago S.M., Pérez-Pérez K., Mercado M.R., Poutou-Piñales R. & Carrascal-Camacho A. (2012). Prevalence of Listeria monocytogenes in pork-meat and other processed products from the Colombian swine industry. Rev MVZ Cordoba, 17, 2827–2833. https://doi.org/10.21897/rmvz.250 

 

 

• Greco, S., Tolli, R., Bossù, T., Flores Rodas, E. M., Di Giamberardino, F., Di Sirio, A., Vita, S., De 633 Angelis, V., Bilei, S., Sonnessa, M., Gattuso, A. & Lanni, L. (2014). Case of contamination 634 by Listeria monocytogenes in mozzarella cheese. Italian Journal of Food Safety, 3(1), 51-53.22. https://doi.org/10.4081/ijfs.2014.1708 

 

• Gómez, D., Pilar Iguácel, L., Rota, C., Carramiñana, J.J., Ariño, A. & Yangüela, J. (2015). Occurrence of Listeria monocytogenes in ready-to-eat meat products and meat processing plants in Spain. Foods, 4, 271–282. https://doi.org/10.3390/foods4030271 

 

• Gudbjörnsdóttir, B., Suihko, M. L., Gustavsson, P., Thorkelsson, G., Salo, S., Sjöberg, A. M., Niclasen, O. & Bredholt, S. (2004). The incidence of Listeria monocytogenes in meat, poultry and seafood plants in the Nordic countries. Food Microbial, 21, 217–222. https://doi.org/10.1016/s0740-0020(03)00012-1 

 

• Hamidiyan, N., Salehi-Abargouei, A., Rezaei, Z., Dehghani-Tafti, R. & Akrami-Mohajeri, F. (2018). The prevalence of Listeria spp. Food contamination in Iran: A systematic review and meta-analysis. Food Research International, 107, 437-50. https://doi.org/10.1016/j.foodres.2018.02.038 

 

• Hellström, S., Laukkanen, R., Siekkinen, K.M., Jukka, R., Maijala, R. & Korkeala, H. (2010). Listeria monocytogenes contamination in pork can originate from farms. Journal of Food Protection, 73, 641–648. https://doi.org/10.4315/0362-028x-73.4.641 

 

• Hoelzer, K., Pouillot, R., Gallagher, D., Silverman, M.B., Kause, J. & Dennis, S. (2012). Estimation of Listeria monocytogenes transfer coefficients and efficacy of bacterial removal through cleaning and sanitation. International Journal of Food Microbiology, 157(2), 267-77. https://doi.org/10.1016/j.ijfoodmicro.2012.05.019 

 

• Henriques, A.R., Gama, L.T. & Fraqueza, M.J. (2017). Tracking Listeria monocytogenes contamination and virulence associated characteristic in the ready-to-eat meat-based food products industry according to hygiene level. International Journal of Food Microbiology, 242, 101–106. https://doi.org/10.1016/j.ijfoodmicro.2016.11.020 

 

• Jalali, M. & Abedi, D. (2008). Prevalence of Listeria speices in food products in Isfahan Iran. Int J Food Microbiol, 122(3), 336-40. https://doi.org/10.1016/j.ijfoodmicro.2007.11.082 

 

• Jami, S., Jamshidi, A. & Khanzadi, S. (2010). The presence of Listeria monocytogenes in raw milk samples in Mashhad, Iran. Iranian Journal of Veterinary Research, 11(4), 363-367. 22099/IJVR.2010.108

 

• Kaur, S., Singh, R., Sran, M.K. & Gill, JP. (2018). Molecular characterization of Listeria monocytogenes in white meat samples from Punjab, India. Indian Journal of Animal Research, 52(11), 1635-41. https://doi.org/10.18805/ijar.b-3414 

 

• Kačániová, M., Kluz, M., Petrová, J., Mellen, M. & Kunová, S. (2015). Incidence of Listeria monocytogenes in meat product samples by real time PCR. Modern Chemistry & Applications, 3, 1–5. https://doi.org/10.4172/2329-6798.1000155 

 

• Kurpas, M., Wieczorek, K. & Osek, J. (2018). Ready-to-eat meat products as a source of Listeria monocytogenes. Journal of Veterinary Research, 62(1),49-55. https://doi.org/10.2478/jvetres-2018-0007 

 

• Kanuganti, S.R., Wesley, I.V., Reddy, P.G., McKean, J. & Hurd, H.S. (2002). Detection Listeria monocytogenes in pigs and pork. Journal of Food Protection, 65, 1470–1474. https://doi.org/10.4315/0362-028x-65.9.1470 

 

• Liu, D. (2006). Identification, subtyping and virulence determination of Listeria monocytogenes, an important foodborne pathogen. Journal of Medical Microbiology, 55(6), 645-59. https://doi.org/10.1099/jmm.0.46495-0 

 

• Montero, D., Bodero, M., Riveros, G., Lapierre, L., Gaggero, A., Vidal, R.M. &Vidal, M. (2015). Molecular epidemiology and genetic diversity of Listeria monocytogenes isolates from a wide variety of ready-to-eat foods and their relationship to clinical strains from listeriosis outbreaks in Chile. Front Microbiology, 6, 1–8. https://doi.org/10.3389/fmicb.2015.00384 

 

• Norowzi, J., Moradi, B. & Shafiee, M. (2013). Detection of actA gene in Listeria monocytogenes isolated from dairy products. Journal of Microbial World, 6(3), 246-252. [In Persian]

 

 

• Osaili, T., Alaboudi, A. & Nesiar, E. (2011). Prevalence of Listeria spp. and antibiotic susceptibility of Listeria monocytogenes isolated from raw chicken and ready-to-eat chicken products in Jordan. Food Control, 22(3), 586-590. https://doi.org/10.1016/j.foodcont.2010.10.008 

 

• Orndorff, P.E., Hamrick, T.S., Smoak, I.W. & Havell, E.A. (2006). Host and bacterial factors in listeriosis pathogenesis. Veterinary Microbiology, 16, 114(1-2), 1-5. https://doi.org/10.1016/j.vetmic.2005.12.003 

 

• Pintado, C. M. B., Oliveira, A., Pampulha, M. E. & Ferreira, M. A. S. (2005). Prevalence and characterization of Listeria monocytogenes isolated from soft cheese. Food Microbiology, 22(1), 79–85. https://doi.org/10.1016/j.fm.2004.04.004 

 

• Pan, Y., Breidt, F. Jr. & Kathariou S. (2006). Resistance of Listeria monocytogenes biofilms to sanitizing agents in simulated food processing environment. Applied and Environmental Microbiology, 72, 7711–7717. https://doi.org/10.1128/aem.01065-06 

 

• Perrin, M., Bremer, M. & Delamare, C. (2007). Fatal microbiological methods, 71(2),13.

Rahimi, E. & Shakarian, A. (2014). Prevalence of Listeria species in ready-to-eat foods in Shahrekord restaurants. Journal of Laboratory Sciences, 2, 87-83. https://doi.org/10.1007/978-3-319-68460-4_16 

 

Rawool, D.B., Malik, S.V.S., Barbuddhe, S.B., Shakuntala, I. & Aurora, R. (2007). Multiplex PCR for Detection of Virulence Associated Genes in Listeria monocytogenes. Internet Journalof Food Safety, 9, 56-62. https://doi.org/10.1016/j.ijfoodmicro.2006.06.029 

 

• Şanlıbaba, P., Tezel, B.U. & Çakmak, GA. (2018). Prevalence and antibiotic resistance of Listeria monocytogenes isolated from ready-to-eat foods in Turkey. Journal of Food Quality, 7693782. https://doi.org/10.1155/2018/7693782 

 

• Shahbazi, A. M., Rashedi, M. & Sohrabi, R. (2013). Comparative contamination of Listeria monocytogenes in traditional dairy products in Esfahan Province, Iran. African Journal of Microbiology Research, 7(16), 1522-1526. https://doi.org/10.5897/ajmr12.1290 

 

• Swaminathan, B. & Gerner-Smidt, P. (2007). The epidemiology of human listeriosis. Microbes Infect, 9, 1236–1243. https://doi.org/10.1016/j.micinf.2007.05.011