Plazmid aracılı kinolon direnci (qnrS1 ve aac (6')-Ib-cr) ve enrofloksasin ile eliminasyonu
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Date
2017-01-26
Authors
Arslan, Erdem
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Publisher
Uludağ Üniversitesi
Abstract
Bu çalışma, florokinolon direncini tanımlamak, florokinolon direnç etmenlerinin enrofloksasinin mutant oluşumunu önleme potansiyeli ve bakteriyel inhibisyon özellikleri üzerindeki etkilerini belirlemek ve enrofloksasinin hayvansal kaynaklı E. coli izolatları üzerindeki etkinliğini değerlendirmek amacıyla gerçekleştirilmiştir. Tez projesinde TUBİTAK (COST) 110O478 numaralı proje kapsamında Bursa ili ve çevresinden toplanan 311 adet hayvansal kaynaklı E. coli izolatı ve 2 kontrol suşu (E. coli ATCC 25922 ve E. coli AG100) ve moleküler olarak oluşturulmuş 3 klon suş kullanıldı. Enrofloksasinin duyarlılığı mikrodilüsyon yöntemi ile belirlendi. Plazmidal genlerin tespiti ve klonlama işlemleri moleküler teknikler ile gerçekleştirildi. kromozomal mutasyonların ve plazmidal genlerinin direnç üzerindeki etkilerini belirlemek amacıyla izolatlar arasından en geniş genetik skalaya sahip 5 izolat, 2 kontrol suşu ve 3 klon suş için Mutant önleyici konsantrasyon (MÖK),Mutant frekansı (MF) ve mutant seçim penceresi (MSP) değerleri belirlendi. Statik in vitro farmakodinamik model kullanılarak zamana bağlı doz-yanıt deneyleri yapıldı. Doksan dokuz E. coli izolatı duyarlı, 50 orta derecede duyarlı, 162 tanesi ise dirençli olarak sınıflandırıldı. On qnrS1 ve 32 adet aac (6')-Ib-cr geni tespit edildi. Gen prevelanları sırasıyla % 3,3 ve % 10,3 olarak belirlendi. İki genin birlikte bulunma oranı ise % 0,62 olarak hesaplandı. MÖK, MF ve MSP çalışmaları sonunda QRDR mutasyonları ve qnrS1ve aac (6')-Ib-cr genlerinin enrofloksasinin MÖK, MF ve MSA değerlerini arttırarak bakterileri enrofloksasinin antimikrobiyal etkilerinden koruduğu belirlendi. Ayrıca zamana bağlı doz-yanıt deneyleri ile enrofloksasinin karakteristik konsantrasyona bağlı bakterisidal etkisi klon suşlar, PMQR genlerine ve/veya QRDR mutasyonlarına sahip E. coli izolatlarında zamana-bağlı veya ko-etkiye dönüştüğü saptandı.
Aims of this study were to identify floroquinolone resistance, characterization of QRDR mutations and PMQR genes and their influence on enrofloxacin resistance as well as assessment and characterization of enrofloxacin activity against fluoroquinolone resistant E. coli isolated from animals. Three hundered and eleven E. coli strains from the sample collection of COST Project 110O478, 2 control strains (E. coli ATCC 25922 and E. coli AG100) and 3 molecularly constructed clones were used in this study. Broth microdilution tests were performed for determination of enrofloxacin susceptibility. Molecular methods were used for detection and cloning of PMQR genes. Mutant prevention concentration (MPC), mutant frequency (MF) and mutant selection window (MSW) values were determined for 5 E. coli isolates 3 clone and 2 control strains. Static in vitro pharmacodynamic model was used for assessment and characterization of enrofloxacin pharmacodynamics. Ninety nine E. coli isolates were classified as susceptible, 50 as intermediate and 162 as resistant. Sequencing results verified 10 S1 and 32 cr variants. Prevelance of PMQR genes were 3,3 % and 10,3 % respectively. Prevelance of coexistance was 0,62 %. MPC, MF, MSW experiments indicated that QRDR mutations and PMQR genes enhance MPC, MF, MSW values and provide protection against enrofloxacin activity thus aiding survival and provoking emergence of enrofloxacin resistant sup-poulations in E. coli. Static in vitro pharmacodynamic modelling revealed characteristic dosedepended bactericidal effect of enrofloxacin is altered and displayed as time-depended or co-effect in the prescence of QRDR mutations and/or PMQR genes in E. coli.
Aims of this study were to identify floroquinolone resistance, characterization of QRDR mutations and PMQR genes and their influence on enrofloxacin resistance as well as assessment and characterization of enrofloxacin activity against fluoroquinolone resistant E. coli isolated from animals. Three hundered and eleven E. coli strains from the sample collection of COST Project 110O478, 2 control strains (E. coli ATCC 25922 and E. coli AG100) and 3 molecularly constructed clones were used in this study. Broth microdilution tests were performed for determination of enrofloxacin susceptibility. Molecular methods were used for detection and cloning of PMQR genes. Mutant prevention concentration (MPC), mutant frequency (MF) and mutant selection window (MSW) values were determined for 5 E. coli isolates 3 clone and 2 control strains. Static in vitro pharmacodynamic model was used for assessment and characterization of enrofloxacin pharmacodynamics. Ninety nine E. coli isolates were classified as susceptible, 50 as intermediate and 162 as resistant. Sequencing results verified 10 S1 and 32 cr variants. Prevelance of PMQR genes were 3,3 % and 10,3 % respectively. Prevelance of coexistance was 0,62 %. MPC, MF, MSW experiments indicated that QRDR mutations and PMQR genes enhance MPC, MF, MSW values and provide protection against enrofloxacin activity thus aiding survival and provoking emergence of enrofloxacin resistant sup-poulations in E. coli. Static in vitro pharmacodynamic modelling revealed characteristic dosedepended bactericidal effect of enrofloxacin is altered and displayed as time-depended or co-effect in the prescence of QRDR mutations and/or PMQR genes in E. coli.
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Keywords
E. coli, qnrS, aac (6’)-Ib-cr, Enrofloksasin, Mutant önleyici konsantrasyon, Statik in vitro farmakodinami, Enrofloxacin, Mutant prevention concentration, Statik in vitro pharmacodynamics
Citation
Arslan, E. (2017). Plazmid aracılı kinolon direnci (qnrS1 ve aac (6')-Ib-cr) ve enrofloksasin ile eliminasyonu. Yayınlanmamış doktora tezi. Uludağ Üniversitesi Sağlık Bilimleri Enstitüsü.