Staphylococcus aureus suşlarında biyofilm oluşumu ve panton valentine lökosidin varlığının araştırılması
Investigating the presence of panton-valentine leukocidin and biofilm production in staphylococcus aureus strains
Yazar: NİLÜFER AKTAŞ
Danışman: PROF. DR. AYŞEGÜL KARAHASAN YAĞCI
Marmara Üniversitesi / Sağlık Bilimleri Enstitüsü / Mikrobiyoloji ve Klinik Mikrobiyoloji Ana Bilim Dalı
Derece: Yüksek Lisans, 2010
Konu: Mikrobiyoloji = Microbiology
Dizin: Biyofilmler = Biofilms ; Kistik fibroz = Cystic fibrosis ; Mikrobiyal duyarlılık testleri = Microbial sensitivity tests ; Staphylococcus aureus = Staphylococcus aureus

Özet:
Staphylococcus aureus, toplum ve hastane kaynaklı enfeksiyonlar yapan önemli bir insan patojenidir. Staphylococcus aureus stafilokoklar içinde çok sayıda enzim ve toksin üreten en virulan türdür. Staphylococcus aureus kistik fibrozlu (KF) hastalarda erken yaşlarda kolonize olarak, kronik akciğer enfeksiyonlarına yol açmaktadır.Bu çalışmada 31'i KF hastası olan toplam 88 hastanın solunum yolu örneklerinden izole edilmiş S. aureus kökenlerinde biyofilm oluşumu ve Panton-Valentine lökosidin (PVL) varlığının ve antibiyotik duyarlıklarının araştırılması amaçlanmıştır. Biyofilm yapımı; kongo kırmızılı agar (KKA) yöntemi ve demir oranı azaltılmış triptik soy buyyon besiyerinde kantitatif mikroplak yöntemi ile tespit edilmiştir. PVL varlığının tespiti için ticari bir nükleik asit hibridizasyon testi (EVIGENE?, AdvanDx, Danimarka) kullanılarak, PVL genleri lukS-PV / lukF-PV saptanmıştır. Kökenlerin antibiyotik duyarlılıkları CLSI (Clinical Laboratory and Standards Institute) kriterlerine göre disk difüzyon yöntemi ile belirlenmiştir.Kongo kırmızılı agar yöntemi ile kökenlerin %23,9'u, kantitatif mikroplak yöntemi ile kökenlerin %64,8'i biyofilm yapımı pozitif olarak saptanmıştır. Çalışmamızda biyofilm yapımı KF hastalarından izole edilen S.aureus kökenlerinde, KF olmayan hasta kökenlerinden daha fazla bulunmuştur. (Kantitatif mikroplak: %96,8 KF, %47,4 KF olmayan (p=0,0005), KKA: %35,5 KF, %17,5 KF olmayan). Test edilen antibiyotiklerden; vankomisin, kloramfenikol ve trimetoprim-sulfametaksazol'e karşı direnç tespit edilmemiştir. Kistik fibrozlu hasta kökenlerinin tümü metisiline duyarlı iken; kistik fibrozlu olmayan hasta kökenlerinin % 54,3'ünde metisilin direnci saptanmıştır. Metisiline duyarlı kökenlerin antibiyotik direnç oranlarında, KF ve KF olmayan hasta kökenleri arasında fark saptanmamıştır. PVL üretimi 88 S.aureus kökeninin 2'sinde (%2,3) pozitif bulunmuştur. PVL pozitif kökenlerin biri kistik fibrozlu hasta diğeri kistik fibrozlu olmayan hastadan izole edilmiştir. PVL yapımı yönünden KF ve KF olmayan hasta kökenleri arasında fark saptanmamıştır.

Abstract:
Staphylococcus aureus is a very important human pathogen which is one of the most common cause of community- acquired and healthcare- associated infections. In staphylococcus species S.aureus is the most virulent one producing a great number of enzymes and toxins. S.aureus leads to chronic pulmoner infections by colonizing cystic fibrosis (CF) patients at the early ages.In this study, it was aimed to detect biofilm and Panton-Valentine leukocidin (PVL) production of 88 S.aureus strains 31 of whom are from CF patients isolated from respiratory tract and to investigate the antimicrobial susceptibility. Biofilm production was detected by congo red agar (CRA) plate and iron limited tryptic soy broth quantitative microplate test. For the detection of Panton-Valentine leukocidin, commercially available nucleic acid hybridization test (EVIGENE?, AdvanDx) was used and lukS-PV / lukF-PV genes were detected. Antibiotic susceptibilities of isolation were detected by disk diffusion method based on CLSI (Clinical Laboratory and Standards Institute).Biofilm production was found positive in 23.9% by CRA and 64.8% by quantitative microplate test. Our study shows that biofilm production of isolates from CF patients is higher than isolates from non-CF patients (Quantitative microplate: 96.8% CF, 47.4% non-CF (p=0,0005), CRA: 35.5% CF, 17.5% non-CF). No resistance was observed for vancomycin, chloramphenicol and trimethoprim/sulphamethoxazole. All CF strains were methicillin sensitive whereas 54.3% of strains from non-CF were methicillin resistant. No difference was observed in antibiotic susceptibility rates of methicillin sensitivite isolates between CF and non-CF patients. Of 88 S.aureus isolates, the PVL production was found positive in two strains. Isolates were one of them was CF patient, whereas the other was non-CF patient. Staphylococcus aureus is a very important human pathogen which is one of the most common cause of community- acquired and healthcare- associated infections. In staphylococcus species S.aureus is the most virulent one producing a great number of enzymes and toxins. S.aureus leads to chronic pulmoner infections by colonizing cystic fibrosis (CF) patients at the early ages.In this study, it was aimed to detect biofilm and Panton-Valentine leukocidin (PVL) production of 88 S.aureus strains 31 of whom are from CF patients isolated from respiratory tract and to investigate the antimicrobial susceptibility. Biofilm production was detected by congo red agar (CRA) plate and iron limited tryptic soy broth quantitative microplate test. For the detection of Panton-Valentine leukocidin, commercially available nucleic acid hybridization test (EVIGENE?, AdvanDx) was used and lukS-PV / lukF-PV genes were detected. Antibiotic susceptibilities of isolation were detected by disk diffusion method based on CLSI (Clinical Laboratory and Standards Institute).Biofilm production was found positive in 23.9% by CRA and 64.8% by quantitative microplate test. Our study shows that biofilm production of isolates from CF patients is higher than isolates from non-CF patients (Quantitative microplate: 96.8% CF, 47.4% non-CF (p=0,0005), CRA: 35.5% CF, 17.5% non-CF). No resistance was observed for vancomycin, chloramphenicol and trimethoprim/sulphamethoxazole. All CF strains were methicillin sensitive whereas 54.3% of strains from non-CF were methicillin resistant. No difference was observed in antibiotic susceptibility rates of methicillin sensitivite isolates between CF and non-CF patients. Of 88 S.aureus isolates, the PVL production was found positive in two strains. Isolates were one of them was CF patient, whereas the other was non-CF patient.