2016, vol. 66, br. 3, str. 336-346
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Distribucija gena rezistencije na antibiotike kod Enterococcus spp. - izolovanih iz uzoraka mleka krava sa mastitisom
Distribution of antibiotic resistance genes in Enterococcus spp.: Isolated from mastitis bovine milk
Veterinary Faculty, Department of Microbiology, Isikli/Aydin-Turkey
e-adresa: gerbas@adu.edu.tr
Projekat: Project of the Adnan Menderes University Scientific Research Projects Unit (ADU-BAP Project code: VTF-13006)
Ključne reči: antibiotic resistantance genes; Entrococcus; Mastitis; PCR; vanA
Sažetak
U ovoj studiji izvršeno je određivanje Enterococcus spp u uzorcima mleka krava sa mastitisom, kao i određivanje njihove osetljivosti na antibiotike i identifikacija gena rezistencije kod izolovanih rezistentnih sojeva. Uzeto je 600 uzoraka mleka od 242 krave sa mastitisom. Izolacija enterokoka je izvršena na selektivnim podlogama, pri čemu je izolovano 94 (16,6%) Enterococcus spp. Primenom sekvencioniranja i PCR-om ukupno je izolovano 94 Enterococcus spp. Sveukupno 5 vrsta (E. faecalis, E. faecium, E. durans, E. hirae, E. mundtii) je izolovano sekvencioniranjem, a 4 (E. faecalis, E. faecium, E. durans, E. hirae) je identifikovano pomoću specifičnih prajmera metodom PCR. Ispitivanjem antibiotske rezistencije 94 sojeva izolovanih enterokoka visoka stopa rezistencije na tetracikline je određena u 77 izolata (81,9%). Tet gen je utvrđen sa sledećom stopom: 54 tetM pozitivno, 23 tetK pozitivno i 17 tetM i tetK pozitivno. Rezistencija na eritromicin je ustanovljena kod 27 (28,7%) izolata (25 ermB), dok je gen rezistencije na hloramfenikol zabeležen u 10 (10,7) izolata. Cat gen je identifikovan kod 9 uzoraka, a jedan izolat je bio rezistentan na vankomicin (1,06%) uz potvrdu VanA gena. Možemo zaključiti da E. faecalis ima ključnu ulogu u nastanku enterokoknog mastitisa pri čemu su izolovani sojevi bili rezistentni na tetraciklin. Jedan vankomicin rezistentan izolat je istovremeno imao izolovan VanA gen.
Abstract
In this study, determination of enterococcus species that were isolated from mastitic milk samples, investigation of their susceptibilities to antibiotics and identification of the existence of resistance genes in resistant strains were conducted. The specimens consist of 600 mastitic milk samples that were collected from 242 cows. Isolation of enterococcus was carried out in selective media and 94 (15.6%) Enterococcus spp. were isolated. A total of 94 species of Enterococci were identified using both sequencing and polymerase chain reaction (PCR). Enterococcus spp. isolates belong to 5 different species (E. faecalis, E. faecium, E. durans, E. hirae, E. mundtii) in sequence analysis and 4 different species (E. faecalis, E. faecium, E. durans, E. hirae) were identify by PCR method with specific primers. Analyzing 94 Enterococcus strains by antibiotic sensitiveness test a high rate of resistance to tetracycline in 77 (81.9%) isolates was shown. The tet resistance genes were identified as follows: 54 were tetM positive, 23 were tetK positive and 17 were positive on tetM and tetK. Resistance to erythromycin was established in 27 (28.7%) isolates (25 ermB) while the chloramphenicol resistance gene was found in 10 (10.7%) of isolates and the cat gene was identified in nine samples and one isolate was resistant to vancomycin (1.06%) with the VanA gene confirmed. In conclusion, it was shown that E. faecalis has the biggest role in enterococcus originated mastitis and these strains were found to be mostly resistant to tetracycline. One vancomycine resistant isolate that had the VanA gene was also determined.
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