Klinik Stafilokokal Suşların Tiplendirilmesinde İki Farklı PCR Temelli Yöntemin Karşılaştırılması

B u çalışmada stafilokokal suşların ayrımında RAPD-PCR Rastgele Çoğaltılmış Polimorfik DNA-Polimeraz Zincir Reaksiyonu ve REP-PCR Tekrarlayan Ekstragenik Elementlerin Polimeraz Zincir Reaksiyonu ile Amplifikasyonu yöntemlerinin etkinliğinin ve kullanılabilirliğinin araştırılmasını ve elde edilen sonuçların antibiyotipleme ile karşılaştırılması amaçlanmaktadır. Çeşitli klinik örneklerden izole edilen ve farklı servislerden toplanan stafilokokal suşlar fenotipik olarak antibiyotik duyarlılık testi, genotipik olarak RAPDPCR ve REP-PCR yöntemleri ile karakterize edilmiştir. RAPD ve REP-PCR ile elde edilen genotipler arasında önemli ölçüde benzerlik bulunamamıştır. RAPD-PCR ve REP-PCR için sırasıyla %80 ve %70 olarak belirlenen benzerlik katsayısı ile suşlar gruplandırılmıştır. RAPD-PCR’ın 0.91 oranında ayrım gücü ile oldukça etkin olduğu tespit edilirken, RW3A primeri, REP1R-I ve REP2-I primerlerinin kombinasyonu ile elde edilen REP analizinin ayrım gücü 0.88 olarak bulunmuştur. Çalışmanın bulguları RAPD-PCR yönteminin klinik örnek ve servisler ile ilişkili kümelemede güvenilir olduğunu göstermektedir. RAPD primerlerinin MRSA, MSSA ve CNS suşların ayrımını sağlayabildiği, REP analizinin RAPD kadar ayırt edici olmadığı bulunmuştur. Böylece RAPD-PCR’ın REP-PCR’dan daha ayırt edici olduğu ve suşların tanımlanmasında hızlı ve doğru bir yöntem olarak uygunluğu kanıtlanmıştır

Anahtar Kelimeler:

Stafilokokal suşlar, antibiyotip, RAPD-PCR, REP-PCR

Comparision of Two PCR-Based Methods in Typing of Clinical Staphylococcal Strains

The present study was carried out to investigate usefulness and effectiveness of randomly amplified polymorphic DNA polymerase chain reaction RAPD-PCR and repetitive element sequence-based polymerase chain reaction REP-PCR in differentiating of staphylococcal strains and to compare the results of these methods with those obtained by antibiotyping. Staphylococcal strains, obtained from various clinical samples and collected from different wards, were characterized phenotypically by susceptibility testing and genotypically by using RAPD-PCR and REP-PCR methods. It was found that there was no significant association between genotypes obtained from RAPD and REP-PCR. Strains with a similarity coefficient of 80% and 70% or greater were grouped in a cluster for RAPD-PCR and REP-PCR, respectively. RAPD-PCR was found to be very efficient with the discriminatory index DI of 0.91 whereas discrimination index DI of REP analysis was found to be 0.88 with RW3A primer and combination of REP1R-I, REP2-I primers. The findings of this study indicate that RAPD-PCR reliably distinguish ward and source-related clustering. The RAPD primers provide to discriminate MRSA, MSSA and CNS strains whereas REP analysis could not be as discriminative as RAPD. Therefore, RAPD-PCR, evidenced to be inconsiderably more discriminatory than REP-PCR, is well suited for fast and accurate strain identification.

Keywords:

Staphylococcal strains, Antibiotype, RAPD-PCR, REP-PCR,

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