K. pneumoniae'da kolistin direncinde rol alan moleküler ve klinik faktörler

Hospital acquired infections are one of the most important health issue affecting millions of patients and cause significant burden. Klebsiella pneumoniae (K.pneumoniae) which is one of the most common agents in worldwide are isolated from nosocomial infections. Colistin has been considered as the last choice of treatment for these infections since carbapenem resistance rate has been increasing in recent years. However, colistin resistance in K.pneumoniae began to increase which points out an emerging threat in the treatment of healthcare related infections. PhoP/PhoQ and PmrA/PmrB are the main systems responsible for modification of LPS layer. Changes in these two component regulatory systems cause colistin resistance. MgrB gene, which encodes the key regulatory protein of PhoP/PhoQ system, has the major role in the colistin resistance. Insertional inactivation or mutations in MgrB gene are highly associated with colistin resistance. In this study; our aim is to understand ongoing emergence of colistin resistance among K. pneumoniae strains by analysing the molecular mechanisms of colistin resistance and comparing the demographic risk factors, clinical futures and outcomes of the infections. A total of 32 patients with a isolation of colistin resistant K.pneumoniae from Ankara Başkent University Hospital, İstanbul Başkent University Hospital and Koşuyolu State Hospital were included to the study. Inactivation of MgrB gene by transposons and mutations in wild type MgrB gene were studied by PCR amplification followed by Sanger sequencing and products were identified by NCBI Blast tool and CLC Genomics Workbench v.4.9 software (CLC Bio, Aarhus, Denmark). Also, the expression levels of PhoQ and PmrK genes in colistin resistant isolates were analysed by quantitative real time PCR. The clonal analysis of the strains were done by rep-PCR using Diversilab system (bioMérieux, France).The univariate analysis of the clinical and molecular data were done by STATA 11 (Texas, USA) Insertional inactivation of MgrB gene was found in 26 out of 32 isolates. MgrB gene was distrupted by different insertion sequences (IS) classified as IS5 like, IS1 like, IS903 like and IS903B. Wild type MgrB was seen in 5 isolates. We detected nonsilent and point mutations in all of 5 isolates carrying wild type MgrB.There was no significant overexpression in PhoQ and PmrK genes. Our clonal analysis revealed that there were two major clones both containing samples from different centres with >95% similarity. Relatively younger age and staying in intensive care unit were found to be associated with IS related colistin resistance (P<.005). The mean duration of the colistin therapy in IS positive group was longer than the IS negative group (19 versus 7 days, p=.231). We report the first findings on the molecular and clinical epidemiology of colistin resistance among K.pneumoniae. The results of the study show that disruption of MgrB gene by transposons is the key mechanism for colistin resistance. Long term stay in intensive care unit, and younger age were major host risk factors that increase the risk of infection by a K.pneumoniae strain carrying IS in its MgrB gene. Prolonged colistin therapy might have an affect on IS related colistin resistance. Demonstration of the horizontal gene transfer between colistin resistant strains highlights the emergence of colistin resistance as the public health problem of the world.