GELENEKSEL KURU FERMENTE TÜRK SUCUĞUNDAN İZOLE EDİLEN ENTEROKOK SUŞLARINDA ANTİBİYOTİK DİRENÇ GENLERİNİN ARAŞTIRILMASI

Yıl 2025, Cilt: 50 Sayı: 4, 491 - 505, 10.08.2025

İbrahim Hakan Falak , Banu Özden Tuncer

https://doi.org/10.15237/gida.GD25031

Öz

Enterococcus faecium ve Enterococcus faecalis, sucuk gibi fermente et ürünlerinde sıklıkla bulunan laktik asit bakterisi türleridir. Bu bakteriler, ürünün benzersiz tadını, dokusunu ve raf ömrünü şekillendirmeye yardımcı olmaları nedeniyle fermentasyon sürecinde önem arz etmektedir. Bazı enterokokların enfeksiyonlara yol açtığı bilinse de çeşitli gıdalardan, özellikle et ürünlerinden izole edilen enterokokların, klinik suşlara kıyasla çok daha düşük patojenik potansiyele sahip olduğu gösterilmiştir. Bu durum, gıda kaynaklı enterokoklara olan ilgiyi artırmıştır. Bu çalışmada, geleneksel olarak fermente edilmiş sucuk örneklerinden izole edilen E. faecium (24 adet) ve E. faecalis (1 adet) olmak üzere toplam 25 Enterococcus suşunda eritromisin (ermA, ermB, ermC), tetrasiklin (tetM, tetL), siprofloksasin (gyrA), streptomisin (strA, strB, aadA, aadE), vankomisin (vanA, vanB) ve gentamisin [(aac(6')aph(2''), aac(3'')II, aac(3'')IV)] direnç genlerinin varlığı polimeraz zincir reaksiyonu (PZR) ile araştırılmıştır. PZR analizinin sonucunda, hiçbir izolatta ermA, ermB, ermC, tetM, tetL, gyrA, strA, strB, aadA, aadE, vanA, vanB, aac(6')aph(2''), aac(3'')II ve aac(3'')IV genlerinin varlığı tespit edilmemiştir. Daha önce yapılan disk difüzyon testlerine göre bazı suşların siprofloksasin (13/25) ve eritromisine (1/25) dirençli olduğu bilinmesine rağmen bu suşların da söz konusu antibiyotik direnç genlerini içermediği görülmüştür. Enterokok türleri genellikle gıda güvenliği açısından şüpheli bir statüye sahip olarak değerlendirilir. Yapılan çalışmada araştırılan antibiyotik direnç genlerinin sucuktan izole edilen enterokoklarda bulunmaması bu suşlara gıda güvenliği açısından bir avantaj sağlamaktadır.

Anahtar Kelimeler

Sucuk , Enterococcus , antibiyotik direnç , polimeraz zincir reaksiyonu

Proje Numarası

SDÜ-BAP 4740-YL1-16

INVESTIGATION OF ANTIBIOTIC RESISTANCE GENES IN ENTEROCOCCUS STRAINS ISOLATED FROM SUCUK, A TRADITIONAL DRY-FERMENTED TURKISH SAUSAGE

Öz

Enterococcus faecium and Enterococcus faecalis are lactic acid bacteria frequently found in fermented meat products such as sausage. These bacteria are important in the fermentation process because they help shape the unique taste, texture, and shelf life of the product. Although some enterococci are known to cause infections, enterococci isolated from various foods, especially meat products, have been shown to have much lower pathogenic potential compared to clinical strains. This has increased interest in foodborne enterococci. In this study, the presence of antibiotic resistance genes for erythromycin (ermA, ermB, ermC), tetracycline (tetM, tetL), ciprofloxacin (gyrA), streptomycin (strA, strB, aadA, aadE), vancomycin (vanA, vanB), and gentamicin [(aac(6')aph(2''), aac(3'')II, aac(3'')IV)] was investigated in a total of 25 Enterococcus strains—E. faecium (24) and E. faecalis (1) —isolated from traditionally fermented sucuk samples by polymerase chain reaction (PCR). As a result of PCR analysis, the presence of the genes ermA, ermB, ermC, tetM, tetL, gyrA, strA, strB, aadA, aadE, vanA, vanB, aac(6')aph(2''), aac(3'')II, and aac(3'')IV was not detected in any isolate. Although previous disk diffusion tests indicated that certain strains were resistant to ciprofloxacin (13/25) and erythromycin (1/25), it was observed that these strains did not harbor the corresponding antibiotic resistance genes. Enterococcus species are generally regarded as having a questionable status in terms of food safety. In this study, the absence of the investigated antibiotic resistance genes in enterococci isolated from sausage provides a potential advantage for these strains with respect to food safety.

Anahtar Kelimeler

Sausage , Enterococcus , antibiotic resistance , polymerase chain reaction

Destekleyen Kurum

Süleyman Demirel University

Proje Numarası

SDÜ-BAP 4740-YL1-16

Teşekkür

This study was supported by the Scientific Research Projects Coordination Unit of Süleyman Demirel University under project number 4740-YL1-16.

Kaynakça

• Akpınar Kankaya, D., Tuncer, Y. (2020). Antibiotic resistance in vancomycin- resistant lactic acid bacteria (VRLAB) isolated from foods of animal origin. Journal of Food Process Preservation, 44: e14468. https://doi.org/10.1111/jfpp.14468
• Ben Braïek, O., Smaoui, S. (2019). Enterococci: Between Emerging Pathogens and Potential Probiotics. BioMed Research International, 5938210, 13 pages. https://doi.org/10.1155/2019/593
• Ben Said, L., Klibi, N., Dziri, R., Borgo, F., Boudabous, A., Ben Slamaa, K., Torresd, C. (2016). Prevalence, antimicrobial resistance and genetic lineages of Enterococcus spp. from vegetable food, soil and irrigation aater in farm environments in Tunisia. Journal of the Science of Food and Agriculture, 96: 1627-1633. https://doi.org/10.1002/jsfa.7264
• Byappanahalli, M. N., Nevers, M. B., Korajkic, A., Staley, Z. R., Harwood, V. J. (2012). Enterococci in the environment. Microbioogy Molecular Biology Reviews, 76: 685–706. https://doi.org/ 10.1128/MMBR.00023-12
• Cancilla, M.R., Powell, I.B., Hillier, A.J., Davidson, B.E. (1992). Rapid genomic fingerprinting of Lactococcus Lactis strains by arbitrarily primed Polymerase Chain Reaction with 32P and fluorescent labels. Applied and Environmental Microbiology, 58: 1772-1775. https://doi.org/10.1128/aem.58.5.1772-1775.1992.
• Cassenego, A., D’Azevedo, P., Van der Sand, S., Frazzon, A., Arent, G. (2017). Comparison of virulence factors and genetic relationships of Enterococcus faecalis strains isolated from clinical, food and poultry samples. Multidisciplinary Advances in Veterinary, 1: 106–115.
• Chajęcka-Wierzchowska, W., Zadernowska, A., García-Solache, M. (2020). Ready-to-eat dairy products as a source of multidrug-resistant Enterococcus strains: Phenotypic and genotypic characteristics. Journal of Dairy Science, 103: 4068–4077 https://doi.org/10.3168/jds.2019-17395
• Chajęcka­Wierzchowska, W., Zarzecka, U., Zadernowska, A. (2021). Enterococci isolated from plant-derived food - Analysis of antibiotic resistance and the occurrence of resistance genes.LWT- Food Science and Technology, 139. https://doi.org/ 10.1016/j.lwt.2020.110549
• CLSI. (2012). Clinical and Laboratory Standards Institute, Performance Standards for Antimicrobial Disk Susceptibility Tests; Approved Standard – Eleventh Edition. M02-A11, Vol. 32, No. 1.
• DANMAP, (2002). Use of antimicrobial agents and occurrence of antimicrobial aesistance in bacteria from food animals, foods and humans in Denmark. Statens Serum Institut og Technical University of Denmark, https://www.danmap.org/-/media/arkiv/projekt-sites/danmap/danmap-reports/danmap_2002.pdf?la=en.
• Dapkevicius, M.d.L.E., Sgardioli, B., Câmara, S.P.A., Poeta, P., Malcata, F.X. (2021). Current trends of Enterococci in dairy products: a comprehensive review of their multiple roles. Foods, 10: 821. https://doi.org/10.3390/ foods10040821
• Delpech, G., Pourcel, G., Schell, C., De Luca, M., Basualdo, J., Bernstein, J., Grenovero, S., Sparo, M. (2012). Antimicrobial resistance profiles of Enterococcus faecalis and Enterococcus faecium isolated from artisanal food of animal origin in Angentina. Foodborne Pathogens and Disease, 9(10): 44-939. https://doi.org/10.1089/Fpd.2012.1192.
• Demirgül, F., Tuncer, Y. (2017). Detection of antibiotic resistance and resistance genes in Enterococci isolated from sucuk, a traditional Turkish dry-fermented sausage. Korean Journal for Food Science of Animal Resources, 37(5): 670-681. https://doi.org/10.5851/kosfa.2017.37.5.670
• Depardieu, F., Perichon, B., Courvalin, P. (2004). Detection of van alphabet and identification of Enterococci and Staphylococci at the species level by multiplex PCR. Journal of Clinical Microbiology, 42(12): 5857-5860. https://doi.org/10.1128/ JCM.42.12.5857-5860.2004
• Dutka Malen, S., Evers, S., Courvalin, P. (1995). Detection of glycopeptide resistance genotypes and identification to the species level of clinically relevant Enterococci by PCR. Journal of Clinical Microbiology, 33: 21-34. https://doi.org/10.1128/ jcm.33.1.24-27.1995
• El-Oraby, S., Awad, A., Younis, G. (2023). Characterization of vancomycin resistant Enterococci isolated from retail poultry meat. Journal of Advanced Veterinary Research, 13(9): 1894-1900. https://advetresearch.com/ index.php/AVR/article/view/1522
• Foulquié Moreno, M. R., Sarantinopoulos, P., Tsakalidou, E., De Vuyst, L. (2006). The role and application of enterococci in food and health. International Journal of Food Microbiology, 106: 1–24. https://doi.org/10.1016/ j.ijfoodmicro.2005.06.026
• Garrido, A.M., Gálvez, A., Pulido R. P. (2014). Antimicrobial resistance in enterococci. Journal of Infectious Diseases Therapy, 2: 4. https://doi.org/ 10.4172/2332-0877.1000150.
• Geniş, B., Öztürk, H., Özden Tuncer, B., Tuncer, Y. (2024). Safety assessment of enterocin-producing Enterococcus strains isolated from sheep and goat colostrum. BMC Microbiology, 24(1): 391-0. https://doi.org/10.1186/s12866-024-03551-7.
• Giacometti, F., Shirzad-Aski, H., Ferreira, S. (2021). Antimicrobials and food-related stresses as selective factors for antibiotic resistance along the farm to fork continuum Antibiotics, 10(6): 671. https://doi.org/10.3390/ antibiotics10060671.
• Golob, M., Pate, M., Kušar, D., Dermota, U., Avberšek, J., Papić, B., Zdovc, I., Bondi, M. (2019). Antimicrobial resistance and virulence genes in Enterococcus faecium and Enterococcus faecalis from humans and retail red meat. Biomed Research International, 14–16. https://doi.org/10.1155/ 2019/2815279.
• Graham, K., Stack, H., Rea, R. (2020). Safety, beneficial and technological properties of enterococci for use in functional food applications–a review. Critical Reviews in Food Science and Nutrition, 60: 3836–3861. https://doi.org/10.1080/10408398.2019.1709800.
• Guzman Prieto, A.M., van Schaik, W., Rogers, M.R.C., Coque, T.M., Baquero, F., Corander, J., Willems, R.J.L. (2016). Global emergence and dissemination of Enterococci as nosocomial pathogens: attack of the clones? Frontier Microbiology, 7: 788. https://doi.org/10.3389/ fmicb.2016.00788
• Gürler, M., Karahan, Z.C., Evren, E., Tekeli, F.A. (2024). Investigation of vanA and vanB genes in vancomycin-susceptible enterococcal strains. Journal of Global Antibiotic Resistance, 39: 36-37. https://doi.org/10.1016/j.jgar.2024.10.115
• Hummel, A., Holzapfel, W.H., Franz, C.M.A.P. (2007). Characterisation and transfer of antibiotic resistance genes from enterococci isolated from food. Systematic and Applied Microbiology, 30: 1-7. https://doi.org/10.1016/j.syapm.2006.02.004
• Huys, G., Botteldoorn, N., Delvigne, F., De Vuyst, L., Heyndrickx, M., Pot, B., Dubois, J.J., Daube, G. (2013). Microbial characterization of probiotics--advisory report of the Working Group "8651 Probiotics" of the Belgian Superior Health Council (SHC). Molecular Nutritional Food Research, 57(8): 1479-504. https://doi.org/10.1002/mnfr.201300065.
• Inoglu, Z.N., Tuncer, Y. (2013). Safety assessment of Enterococcus faecium and Enterococcus faecalis strains isolated from Turkish tulum cheese. Journal of Food Safety, 33(3): 369-377. https://doi.org/10.1111/jfs.12061
• Jahan, M., Krause, D.O., Holley, R.A. (2013). Antimicrobial resistance of Enterococcus species from meat and fermented meat products isolated by a PCR-based rapid screening method. International Journal of Food Microbiology, 163: 89-95. https://doi.org/10.1016/j.ijfoodmicro.2013.02.017
• Jaimee, G., Halami, P.M. (2016). High level aminoglycoside resistance in Enterococcus, Pediococcus and Lactobacillus species from farm animals and commercial meat products. Annals of Microbiology, 66: 101–110. https://doi.org/10.1007/s13213-015-1086-1.
• Khalifa, M., Mohmmad, M., Aoalwafa, W., Shafik, N. (2024). The Enterococcus: Review of its characters regarding virulence factors, antibiotic Resistance, pathogenesis, and Treatment. Sohag Medical Journal, 28(3.): 35-43. doi: 10.21608/smj.2024.313933.1491
• Kim, Y., B., Seo, H., J., Seo, K., W., Jeon, H., Y., Kim, D., K., Kim, S., W., Lim, S., K., Lee, Y., J. (2018). Characteristics of high-level ciprofloxacin-resistant Enterococcus faecalis and Enterococcus faecium from retail chicken meat in Korea. Journal of Food Proection, 81(8): 1357-1363. https://doi.org/10.4315/0362-028X.JFP-18-046. PMID: 30015506.
• Kühn, I., Iversen, A., Finn, M., Greko, C., Burman, L.G., Blanch, A.R., Vilanova, X., Manero, A. (2005). Occurrence and relatedness of vancomycin-resistant enterococci in animals, humans and the environment in different European regions. Applied and Environmental Microbiology, 71: 5383-5390. https://doi.org/ 10.1128/AEM.71.9.5383-5390.2005
• Kürekci, C., Pehlivanlar Önen, S., Yipel, M., Aslantaş, Ö., Gündoğdu, A. (2016). Characterisation of phenotypic and genotypic antibiotic resistance profile of Enterococci from cheeses in Turkey. Korean Journal for Food Science of Animal Resources, 36(3): 352-358. https://doi.org/ 10.5851/kosfa.2016.36.3.352
• Larsen, J., Schonheider, H.C., Lester, C.H., Olsen, S.S., Porsbo, L.J., Garcia-Migura, L. (2010). Porcine-origin gentamicin-resistant Enterococcus faecalis in humans. Denmark. Emerg. Emerging Infectious Diseases, 16: 682-684. https://doi.org/ 10.3201/ eid1604.090500.
• Lester, C.H., Frimodt-Møller, N., Sørensen, T.L., Monnet, D.L., Hammerum, A.M. (2006). In vivo transfer of the vanA resistance gene from an Enterococcus faecium isolate of animal origin to an E. faecium isolate of human origin in the intestines of human volunteers. Antimicrobial Agents and Chemotherapy, 50: 596-599. https://doi.org/ 10.1128/aac.50.2.596-599.2006
• Moubarek, C., Bourgeois, N., Courvalin, P., Doucet-Populaire, F. (2003). Multiple antibiotic resistance gene transfer from animal to human Enterococci in the digestive tract of gnotobiotic mice. Antimicrobial Agents and Chemotherapy, 47: 2993-2996. https://doi.org/10.1128/ aac.47.9.2993-2996.2003
• Murray, B.E. (1997). Vancomycin-resistant enterococci. The American Journal of Medicine, 102(3): 284-93. https://doi.org/10.1016/s0002-9343(99)80270-8
• Murray, B.E. (2000). Vancomycin-resistant enterococcal infection. The New England Journal of Medicine, 342(10): 710-21. https://doi.org/ 10.1056/nejm200003093421007
• Ouoba, L.I.I., Lei, V., Jensen, L.B. (2008). Resistance of potential probiotic Lactic Acid Bacteria and Bifidobacteria of African and European origin to antimicrobials: determination and transferability of the resistance genes to other bacteria. International Journal of Food Microbiology, 121: 217-224. https://doi.org/10.1016/ j.ijfoodmicro.2007.11.018
• Özdemir, R., Tuncer, Y. (2020). Detection of antibiotic resistance profiles and aminoglycoside-modifying enzyme (AME) genes in high-level aminoglycoside-resistant (HLAR) enterococci isolated from raw milk and traditional cheeses in Turkey. Molecular Biology Reports, 47(3): 1703-1712. https://doi.org/10.1007/s11033-020-05262-4,
• Semedo-Lemsaddek, T., Cota, J.B., Ribeiro, T., Pimentel, A., Tavares, L., Bernando, F., Oliveira. M. (2021). Resistance and virulence distribution in enterococci isolated from broilers reared in two farming systems. Irish Veterinary Journal, 74(1): 22 https://doi.org/10.1186/s13620-021-00201-6
• Shepard, B.D., Gilmore, M.S. (2002). Antibiotic resistant Enterococci: the mechanisms and dynamics of drug introduction and resistance. Microbes and Infection, 4: 215-224. https://doi.org/10.1016/s1286-4579(01)01530-1
• Sparro, M., Urbizu, L., Solana, M.V., Pourcel, G., Delpech, G., Confalonieri, A., Ceci M., Sánchez Bruni S.F. (2012). High-level resistance to gentamicin: genetic transfer between Enterococcus faecalis isolated from food of animal origin and human microbiota. Applied Microbiology and Biotechnology, 54: 119-125. https://doi.org/10.1111/j.1472-765x.2011.03182.x
• Talon, R., Leroy, S. (2011). Diversity and safety hazards of bacteria involved in meat fermentations. Meat Science, 89: 303- 309. https://doi.org/10.1016/j.meatsci.2011.04.029
• Telli, N., Telli, A., Biçer, Y., Turkal, G., Uçar, G. (2021). Isolation and antimicrobial resistance of vancomycin resistant Enterococcus spp. (VRE) and methicillin-resistant S. aureus (MRSA) on beef and chicken meat, and workers hands from slaughterhouses and retail shops in Turkey. Journal of the Hellenic Veterinary Medical Society, 72(4): 3345–3354. https://doi.org/10.12681/jhvms.29373
• Teixeira, L.M., Facklam, R.R. (2003). Enterococcus. In: Murray, P.R., Baron, E.J., Jorgensen, J.H., Pfaller, M.A., Yolken, R.H., (eds). Manual of Clinical Microbiology, 8th edn. ASM Press, Washington, 422-433.
• Tuncer, B.Ö., Ay, Z., Tuncer, Y. (2013). Occurence of enterocin genes, virulence factors, and antibiotic resistance in 3 bacteriocin-producer Enterococcus faecium strains isolated from Turkish tulum cheese. Turkish Journal of Biology, 37: 443-449. https://doi.org/10.3906/biy-1209-26
• Wiśniewski, P., Zakrzewski, A., Chajęcka-Wierzchowska, W., Zadernowska, A. (2024). Possibility of transfer and activation of 'silent' tetracycline resistance genes among Enterococcus faecalis under high-pressure processing. Food Microbiology, 120: 104481, https://doi.org/ 10.1016/j.fm.2024.104481.
• Werner, G., Coque, T.M., Franz, C.M., Grohmann, E., Hegstad, K. (2013). Antibiotic resistant Enterococci tales of a drug resistance gene trafficker. International Journal of Medical Microbiology, 303: 360-379. https://doi.org/10.1016/j.ijmm.2013.03.001
• WHO. WHO publishes list of bacteria for which new antibiotics are urgently needed [Internet]. 2017 [cited 2022 December 6]. Available from: https://www.who.int/ne ws/item/27-02-2017-who-publishes-list-of-bacteria-for-which-new-antibiotics-areurgently-needed.
• Yalçın, M., Özden Tuncer, B., Akpınar Kankaya, D., Tuncer, Y. (2023). Presence of genes encoding aminoglycoside-modifying enzyme (AME) and virulence factors in high-level aminoglycoside-resistant (HLAR) Enterococcus strains isolated from retail chicken meat in Turkey. Journal of the Hellenic Veterinary Medical Society, 74(4): 6439-6448. https://doi.org/10.12681/jhvms.30850,
• Yamagami, Y., Asao, M., Takahashi, A., Hashimoto, Y., Okuyama, N., Arai, E., Arihara, W., Masui, R., Shimazaki, Y. (2024). Prevalence and antimicrobial resistance of Enterococcus spp. isolated from animal feed in Japan. Frontier in Veterinary Science, 10: 1328552. https://doi.org/10.3389/fvets.2023.1328552
• Yogurtcu N.N., Tuncer, Y. (2013). Antibiotic susceptibility patterns of Enterococcus strains isolated from Turkish tulum chesee. International Journal of Dairy Technology, 66: 236-242. https://doi.org/10.1111/1471-0307.12014
• Yüceer, Ö., Özden Tuncer, B. (2015). Determination of antibiotic resistance and biogenic amine production of lactic acid bacteria isolated from fermented Turkish sausage (sucuk). Journal of Food Safety, 35: 276-285. https://doi.org/10.1111/jfs.12177