Investigation of MCR resistance genes by determining colistin resistance in Klebsiella pneumoniae isolats by different methods

Tutan K., Investigation of mcr resistance genes by determining colistin resistance in Klebsiella pneumoniae isolates with different methods. Van Yüzüncü Yıl University Faculty of Medicine, Department of Medical Microbiology, Medical Expertise Thesis, Van, 2024. The development of multiple antibiotic resistance in K.pneumoniae infections has made the use of carbapenems as the first choice widespread. Since colistin has become the only treatment option for carbapenem-resistant K.pneumoniae, its use has become widespread and colistin resistance has begun to increase worldwide. Colistin resistance can be caused by chromosomal mechanisms or plasmid-borne mobile colistin resistance genes (mcr). In our study, K.pneumoniae strains; It was aimed to determine colistin resistance by broth microdilution, automated system, disk diffusion, gradient test, disk elution method, to detect mcr(1-10) resistance genes by PCR method and to reveal the clonal relationship between the strains. A total of 81 K.pneumoniae strains, 45 sensitive to colistin and 36 resistant to colistin, isolated from various clinical samples taken from patients admitted to Van YYÜ Dursun Odabaş Medical Center, were included in the study. Identification of strains and their resistance to other antibiotics were determined with the Phoenix M50 (Becton Dickinson, USA) automated system. Automated system, Disc Elution, Gradient test and Disc Diffusion methods for colistin resistance were compared with the Broth Microdilution Method, which is recommended as the reference method. After K. pneumoniae strains were confirmed by the PCR method, the presence of plasmid-transmitted mcr (1-10) genes in all strains was investigated by the PCR method. ERIC primers were used to determine the clonal relationship between strains. According to antibiotic sensitivity tests; The antibiotics to which colistin-sensitive strains are most sensitive are; While amikacin, gentamicin, levofloxacin and ciprofloxacin are the antibiotics to which colistin-resistant strains are most sensitive; They were identified as amikacin, gentamicin, trimethoprim-sulfomethoxazole, levofloxacin and imipenem. In terms of determining colistin resistance, the sensitivity of the automated system was found to be 72.2%, the specificity was 100%, and the accuracy rate was 87%. Disc elution 24-28. In hourly readings, its sensitivity is 66.7%, its specificity is 100%, and its accuracy rate is 85%. 16-20 is recommended for disk elution. The rates during the hours were determined to be lower. The sensitivity of the gradient test method is 5.6%, the specificity is 100%, and the accuracy rate is 58%; When the resistance limit of the disc diffusion method is taken as 14 mm in zone diameter; The sensitivity was found to be 55%, the specificity was 91%, and the accuracy rate was 75%. The mcr(1-10) resistance genes, in which colistin resistance is transferred via plasmid, were not detected in any of the strains. 25 different genotypes were detected among K. pneumoniae isolates, and clustering isolates were collected in 13 different clusters. 69 of the isolates were located in any cluster, and the clustering rate was found to be 85.2%. When comparing the BMD reference method with other methods for determining colistin resistance; 24-28 that the method with the highest sensitivity and accuracy is the automated system. It was observed that the disk elution method was followed in hours. mcr (1-10) resistance genes, in which colistin resistance is transferred via plasmid, were not detected in any of the 81 strains. Key words: K.pneumoniae, colistin resistance, PCR, mcr genes