Determination of Ampicillin Resistant Enterococci (Are) Isolated From Canine and Feline Rectal Swabs

Abstract

    Enterococci species, which are normal inhabitants of the gut flora of healthy animals and human, began to be recognized as an important pathogen in both human and veterinary medicine due to the acquired resistance profiles. The aim of the study is to examine the diversity of ampicillin resistance enterococci (ARE) species in cats and dogs, their antimicrobial susceptibility profiles and to determine some of the virulence related genes; ace, gelE, efaA, agg and esp. For this purpose, rectal swabs from companion animals were collected and processed for ampicillin resistant enterococci isolation. One hundred fifty seven swab samples (86 canine and 71 feline) were examined. ARE were isolated from 18 canine and 18 feline samples. All isolates identified as E. faecium by PCR. Antimicrobial susceptibilities of the isolates were determined by disk diffusion method. The isolates were resistant to ampicillin, penicillin, tetracycline (100%), followed by rifampicin and erythromycin (97%), streptomycin (92%), gentamicin (81%), ciprofloxacin (61%), nitrofurantoin (19%). Only two of E. faecium isolates were resistant to vancomycin and one to chloramphenicol. Multidrug resistance (resistance ≥4 antimicrobials) observed in all isolates. Virulence genes ace, agg and esp were not detected in any of the tested isolates. The efaA and gelE genes detection rates were, 13.8% and 11.1% respectively. The ARE isolation rate among pet animals was 22.9%. Screening of antimicrobial resistant enterococci among companion animals would be useful to detect any emerging antimicrobial resistance problem related with public health.

Keywords

• Antimicrobial resistance,cat,dog,PCR,ampicillin resistant Enterococci

Köpek ve Kedi Rektal Svablarından İzole Edilen Ampisilin-Dirençli Enterokokların İncelenmesi

Abstract

Sağlıklı hayvanlar ve insanların barsak florasının bir parçası olan Enterekok türleri, kazanılmış direnç profilleri nedeniyle hem insan hem de hayvan hekimliğinde önemli birer patojen olarak tanınmaya başlanmıştır. Bu çalışmada, kedi ve köpeklerde ampisiline dirençli enterokok (ARE) türlerinin dağılımını, antimikrobiyal duyarlılık profillerini ve bu izolatların virülans ile ilişkili ace, gelE, efaA, agg ve esp genleri incelendi. Bu amaçla evcil hayvanlardan (86 köpek, 71 kedi) toplanan yüz elli yedi svap örneği ampisilin dirençli enterokok izolasyonu yönünden incelendi. ARE 18 kedi ve 18 köpek örneğinden izole edildi. İzolatların antimikrobiyal duyarlılıkları disk difüzyon metodu ile belirlendi. Bütün izolatlar PCR ile E.faecium olarak saptandı. İzolatlar ampisilin, penisilin ve tetrasiklin (%100), rifampisin ve eritromisin (%97), streptomisin (%92), gentamisin (%81), siprofloksasin (%61) ve nitrofurantoin’e (%19) dirençliydi. Sadece iki izolat vankomisine, bir izolat kloramfenikole dirençliydi. Bütün izolatlarda çoklu-antibiyotik direnci (direnç ≥4 antimikrobiyal) saptandı. Virülens genleri ace, agg ve esp test edilen hiçbir izolatta saptanmadı. efaA ve gelE genlerinin saptanma oranları sırasıyla %13,8 ve %11,1 di. Pet hayvanlarında ARE izolasyon oranı %22,9 olarak saptandı. Evcil hayvanlar arasında antimikrobiyal dirençli Enterokokların taranması halk sağlığı ile ilişkilidir. Ayrıca hayvan ve insanlarda antimikrobiyal direnç sorununu tespit etmek ve zamanında önlem alabilmek konusunda yapılan çalışmanın aydınlatıcı olabileceği düşünülmektedir.

Keywords

• Antimikrobiyal direnç,kedi,köpek,PCR,ampisilin dirençli Enterokok

References

1. Bagcıgil, A.F., Ikiz, S., Güzel, Ö., Parkan Yaramiş, Ç., Ilgaz, A., 2012. Hayvanlardan, klinik ortamından ve klinik çalışanlarından izole edilen metisiline dirençli stafilokokların tür dağılımları. İstanbul Üniversitesi Veteriner Fakültesi Dergisi 38, 151-160.
2. Billström, H., Lund, B., Sullivan, A., Nord, C.E., 2008. Virulence and antimicrobial resistance in clinical Enterococcus faecium. International Journal of Antimicrobial Agents 32, 374-377.
3. Cariolato, D., Andrighetto, C., Lombardi, A., 2008. Occurrence of virulence factors and antibiotic resistances in Enterococcus faecalis and Enterococcus faecium collected from dairy and human samples in North Italy. Food Control 19, 886-892.
4. Cinquepalmi, V., Monno, R., Fumarola, L., Ventrella, G., Calia, C., Greco, M. F., Soleo, L., 2013. Environmental Contamination by Dog’s Faeces: A Public Health Problem? International Journal of Environmental Research and Public Health 10, 72-84. doi:10.3390/ijerph10010072
5. Cetinkaya, Y., Falk, P., Mayhall, C.G., 2000. Vancomycin-Resistant Enterococci. Clinical Microbiology Reviews 13, 686-707.
6. Clinical and Laboratory Standard Institute, 2012. Performance Standards for Antimicrobial Susceptibility Testing; 22nd Informational Supplement M100-A22. Wayne, PA: Clinical and Laboratory Standards Institute.
7. Coque, T.M., Willems, R., Cantón, R., Del Campo, R., Baquero, F., 2002. High occurrence of esp among ampicillin-resistant and vancomycin-susceptible Enterococcus faecium clones from hospitalized patients. Journal of Antimicrobial Chemotherapy 50, 1035-1038.
8. Damborg, P., Top, J., Hendrickx, A.P., Dawson, S., Willems, R.J., Guardabassi, L., 2009. Dogs are a reservoir of ampicillin-resistant Enterococcus faecium lineages associated with human infections. Applied and Environmental Microbiology 75, 2360-2365.

9. Duprè, I., Zanetti, S., Schito, A.M., Fadda, G., Sechi, L. A., 2003. Incidence of virulence determinants in clinical Enterococcus faecium and Enterococcus faecalis isolates collected in Sardinia (Italy). Journal of Medical Microbiology 52, 491-498.
10. Eaton, T.J., Gasson, M.J., 2001. Molecular Screening of Enterococcus Virulence Determinants and Potential for Genetic Exchange between Food and Medical Isolates. Applied and Environmental Microbiology 67, 1628-1635.
11. Fisher, K., Phillips, C., 2009. The ecology, epidemiology and virulence of Enterococcus. Microbiology 155, 1749-1757.
12. Guardabassi, L., Schwarz, S., Lloyd, D.H., 2004. Pet animals as reservoirs of antimicrobial-resistant bacteria. Journal of Antimicrobial Chemotherapy 54, 321-332.
13. Gülhan, T., Aksakal, A., Ekin, I.H., Savaşan, S., Boynukara, B., 2007. Virulence factors of Enterococcus faecium and Enterococcus faecalis strains isolated from humans and pets. Turkish Journal of Veterinary and Animal Sciences 30, 477-482.
14. Huycke, M.M., Sahm, D.F., Gilmore, M.S., 1998. Multiple-drug resistant enterococci: the nature of the problem and an agenda for the future. Emerging Infectious Diseases 4, 239-249.
15. Kariyama, R., Mitsuhata, R., Chow, J.W., Clewell, D.B., Kumon, H., 2000. Simple and reliable multiplex PCR assay for surveillance isolates of vancomycin-resistant Enterococci. Journal of Clinical Microbiology 38, 3092-3095.
16. Kataoka, Y., Ito, C., Kawashima, A., Ishii, M., Yamashiro, S., Harada, K., Ochi, H., Sawada, T., 2013. Identification and antimicrobial susceptibility of Enterococci isolated from dogs and cats subjected to differing antibiotic pressures. The Journal of Veterinary Medical Science/The Japanese Society of Veterinary Science 75(6), 749-753.
17. Kayaoglu, G., Ørstavik, D., 2004. Virulence factors of Enterococcus faecalis: Relationship to endodontic disease. Critical Reviews in Oral Biology and Medicine 15(5), 308-320.
18. Ke, D., Picard, F.J., Martineau, F., Ménard, C., Roy, P.H., Ouellette, M., Bergeron, M.G., 1999. Development of a PCR assay for rapid detection of Enterococci. Journal of Clinical Microbiology 37(11), 3497-3503.
19. Kwon, K.H., Moon, B.Y., Hwang, S.Y., Park, Y.H., 2012. Detection of CC17 Enterococcus faecium in dogs and a comparison with human isolates. Zoonoses and Public Health 59(6), 375-378.
20. Lester, C.H., Sandvang, D., Olsen, S.S., Schønheyder, H.C., Jarløv, J.O., Bangsborg, J., Hansen, D.S., Jensen, T.G., Frimodt-Møller, N., Hammerum, A.M., 2008. Emergence of ampicillin-resistant Enterococcus faecium in Danish hospitals. Journal of Antimicrobial Chemotherapy 62(6), 1203-1206.
21. Lopes, M.D.F.S., Simões, A.P., Tenreiro, R., Marques, J.J.F., Crespo, M.T.B., 2006. Activity and expression of a virulence factor, gelatinase, in dairy enterococci. International Journal of Food Microbiology 112(3), 208-214.
22. Low, Y.L., Jakubovics, N.S., Flatman, J.C., Jenkinson, H.F., Smith, A.W., 2003. Manganese-dependent regulation of the endocarditis-associated virulence factor EfaA of Enterococcus faecalis. Journal of Medical Microbiology 52(2), 113-119.
23. Mannu, L., Paba, A., Daga, E., Comunian, R., Zanetti, S., Duprè, I., Sechi, L.A., 2003. Comparison of the incidence of virulence determinants and antibiotic resistance between Enterococcus faecium strains of dairy, animal and clinical origin. International Journal of Food Microbiology 88(2), 291-304.
24. Martin, B., Corominas, L., Garriga, M., Aymerich, T., 2009. Identification and tracing of Enterococcus spp. by RAPD‐PCR in traditional fermented sausages and meat environment. Journal of Applied Microbiology 106(1), 66-77.
25. Mundy, L.M., Sahm, D.F., Gilmore, M., 2000. Relationships between Enterococcal virulence and antimicrobial resistance. Clinical Microbiology Reviews 13(4), 513-522.
26. Nallapareddy, S.R., Qin, X., Weinstock, G.M., Höök, M., Murray, B.E., 2000. Enterococcus faecalis Adhesin, Ace, Mediates attachment to extracellular matrix proteins Collagen type IV and laminin as well as Collagen type I. Infection and Immunity 68(9), 5218-5224.
27. Rathnayake, I.U., Hargreaves, M., Huygens, F., 2012. Antibiotic resistance and virulence traits in clinical and environmental Enterococcus faecalis and Enterococcus faecium isolates. Systematic and Applied Microbiology 35(5), 326-333.
28. Rodrigues, J., Poeta, P., Martins, A., Costa, D., 2002. The importance of pets as reservoirs of resistant Enterococcus strains, with special reference to vancomycin. Journal of Veterinary Medicine, Series B 49(6), 278-280.
29. Sava, I.G., Heikens, E., Huebner, J., 2010. Pathogenesis and immunity in enterococcal infections. Clinical Microbiology and Infection 16(6), 533-540.
30. Shankar, V., Baghdayan, A.S., Huycke, M.M., Lindahl, G., Gilmore, M.S., 1999. Infection-derived Enterococcus faecalis strains are enriched in esp, a gene encoding a novel surface protein. Infection and Immunity 67(1), 193-200.
31. Toledo-Arana, A., Valle, J., Solano, C., Arrizubieta, M.J., Cucarella, C., Lamata, M., Amorena, B., Leiva, J., Penades, J.R., Lasa, I., 2001. The enterococcal surface protein, Esp, is involved in Enterococcus faecalis biofilm formation. Applied and Environmental Microbiology 67(10), 4538-4545.
32. Tremblay, C.L., Charlebois, A., Masson, L., Archambault, M., 2013. Characterization of hospital-associated lineages of ampicillin-resistant Enterococcus faecium from clinical cases in dogs and humans. Frontiers in Microbiology 4, 245.