Publication: Candida ve aspergillus enfeksiyonlarının real time PCR yöntemi ile hızlı tanısının kültür yöntemi ile karşılaştırılması ve antifungal direncin kolorimetrik yöntemle tayini
Abstract
CANDİDA ve ASPERGİLLUS ENFEKSİYONLARININ REAL TIME PCR YÖNTEMİ ile HIZLI TANISININ KÜLTÜR YÖNTEMİ ile KARŞILAŞTIRILMASI ve ANTİFUNGAL DİRENCİN KOLORİMETRİK YÖNTEMLE TAYİNİ Bu çalışma 2009 - 2011 yılları arasında Yeditepe Üniversitesi Hastanesi Tıbbi Mikrobiyoloji vuran hastalardan alınan örnekler kullanılarak gerçekleştirilmiştir. Toplamda 139 adet örnek çalışılmış ve 139 adet suş izole edilmiştir. Bu suşlar, hastalardan alınan abse, ağız sürüntsü, balgam, BAL (Bronkoalveolar Lavaj), idrar ve idrar sondası, kan kültürü, kateter, kulak akıntısı ve sürüntüsü, trakeal aspirat, steril vücut sıvısı örneklerinden elde edilmiştir. Çalışmada kültür yöntemi ile tanımlanan Candida ve Aspergillus türleri, moleküler yöntemlerle tanımlanmıştır, bu iki yöntemin sonuçları karşılaştırılmıştır. Çalışmamızda Aspergillus cinsine ait Aspergillus flavus (Link, 1809), Aspergillus fumigatus (Fresen, 1863), Aspergillus niger (Tiegh, 1867), Aspergillus terreus (Thom, 1918) ile Candida cinsine ait Candida albicans ((C.P. Robin) Berkhout 1923), Candida glabrata ((H.W. Anderson) S.A. Mey. & Yarrow, 1978), Candida parapsilosis ((Ashford) Langeron & Talice, 1932), Candida tropicalis ((Castell.) Berkhout, 1923), Candida dubliniensis (D.J. Sullivan, Western., K.A. Haynes, Dés.E. Benn. & D.C. Coleman, 1995), Candida krusei ((Castell.) Berkhout, 1923) türleri izole edilmiştir. Sybr green boyası ile panfungal PCR yapılarak öncelikle cins düzeyinde Aspergillus ve Candida ayrımı yapılmıştır. Daha sonra Taqman PCR ile 4 farklı Candida türü (Candida albicans, Candida glabrata, Candida tropicalis ve Candida parapsilosis) tanımlanmıştır. Kültür yöntemi ile tanımlanan 57 adet Candida albicans; 12 adet C. glabrata; 21 adet C. parapsilosis; 17 adet C. tropicalis suşlarının tamamı Taqman PCR ile de doğrulanmıştır ve bu iki yöntem ile yapılan identifikasyon sonuçları arasındaki uyumun %100 olduğu belirlenmiştir. Ayrıca Candida türlerinin kolorimetrik bir yöntem ile 7 farklı antifungale (Flukonazol, Kaspofungin, Vorikonazol, Amfoterisin B, Posakonazol, İtrakonazol ve 5-Flusitosin) karşı duyarlılık deneyleri yapılmıştır. Kaspofungine dirençli bir suşa rastlanmazken, en fazla direnç azol grubu antifungallerde saptanmıştır.
REAL TIME PCR for RAPID DIAGNOSIS OF CANDIDA and ASPERGILLUS INFECTIONS with COMPARISON of CULTURE METHOD and DETERMINATION of ANTIFUNGAL RESISTANCE COLORIMETRIC METHOD In this study, specimen, collected from the patients who applied to Medical Microbiology Laboratory of Yeditepe University Hospital between 2009-2011 were evaluated. 139 specimen were studied and 139 strains were isolated. These strains were isolated from abscesses, oral swabs, sputum, BAL (Bronchoalveolar lavage), urine, and urinary catheter, blood cultures, catheter, ear discharge and swabs, tracheal aspirates, sterile body fluid samples. In the study Aspergillus flavus (Link, 1809), Aspergillus fumigatus (Fresen, 1863), Aspergillus niger (Tiegh, 1867), Aspergillus terreus (Thom, 1918) species, which were belong to Aspergilus genus and Candida albicans ((C.P. Robin) Berkhout 1923), Candida glabrata ((H.W. Anderson) S.A. Mey. & Yarrow, 1978), Candida parapsilosis ((Ashford) Langeron & Talice, 1932), Candida tropicalis ((Castell.) Berkhout, 1923), Candida dubliniensis (D.J. Sullivan, Western., K.A. Haynes, Dés.E. Benn. & D.C. Coleman, 1995), Candida krusei ((Castell.) Berkhout, 1923) species which were belong to Candida genus were isolated. Primarily panfungal PCR was made by Sybr green dye to distinguish Candida and Aspergillus at species level. Then four different Candida species (Candida albicans, Candida glabrata, Candida tropicalis ve Candida parapsilosis) have been identified by Taqman PCR. 57 Candida albicans, 12 C. glabrata, 21 C. parapsilosis, 17 C. tropicalis; which were defined by culture method confirmed by Taqman PCR. The results of identification with these two methods were found 100% compatible. In addition, antifungal susceptibility of Candida species against 7 different antifungals (Fluconazole, Caspofungin, Voriconazole, Amphotericin B, Posaconazole, Itrakonazole ve 5-Flucytosine) determined by colorimetric method. Caspofungin resistant strains were not found, the most resistance was determined to azole antifungals .
REAL TIME PCR for RAPID DIAGNOSIS OF CANDIDA and ASPERGILLUS INFECTIONS with COMPARISON of CULTURE METHOD and DETERMINATION of ANTIFUNGAL RESISTANCE COLORIMETRIC METHOD In this study, specimen, collected from the patients who applied to Medical Microbiology Laboratory of Yeditepe University Hospital between 2009-2011 were evaluated. 139 specimen were studied and 139 strains were isolated. These strains were isolated from abscesses, oral swabs, sputum, BAL (Bronchoalveolar lavage), urine, and urinary catheter, blood cultures, catheter, ear discharge and swabs, tracheal aspirates, sterile body fluid samples. In the study Aspergillus flavus (Link, 1809), Aspergillus fumigatus (Fresen, 1863), Aspergillus niger (Tiegh, 1867), Aspergillus terreus (Thom, 1918) species, which were belong to Aspergilus genus and Candida albicans ((C.P. Robin) Berkhout 1923), Candida glabrata ((H.W. Anderson) S.A. Mey. & Yarrow, 1978), Candida parapsilosis ((Ashford) Langeron & Talice, 1932), Candida tropicalis ((Castell.) Berkhout, 1923), Candida dubliniensis (D.J. Sullivan, Western., K.A. Haynes, Dés.E. Benn. & D.C. Coleman, 1995), Candida krusei ((Castell.) Berkhout, 1923) species which were belong to Candida genus were isolated. Primarily panfungal PCR was made by Sybr green dye to distinguish Candida and Aspergillus at species level. Then four different Candida species (Candida albicans, Candida glabrata, Candida tropicalis ve Candida parapsilosis) have been identified by Taqman PCR. 57 Candida albicans, 12 C. glabrata, 21 C. parapsilosis, 17 C. tropicalis; which were defined by culture method confirmed by Taqman PCR. The results of identification with these two methods were found 100% compatible. In addition, antifungal susceptibility of Candida species against 7 different antifungals (Fluconazole, Caspofungin, Voriconazole, Amphotericin B, Posaconazole, Itrakonazole ve 5-Flucytosine) determined by colorimetric method. Caspofungin resistant strains were not found, the most resistance was determined to azole antifungals .