Determination of resistance genes in carbapenem-resistant enterobacteriaceae strains and evaluation of the effectiveness of newly introduced antibiotics against resistant strains

Multidrug-resistant gram-negative bacterial infections pose a significant global public health threat due to the challenges associated with their treatment, the increasing mortality rates, and the high associated treatment costs. The development of beta-lactamase resistance to beta-lactam antibiotics, which are central to the treatment of gram-negative bacterial infections, has led to an increased use of carbapenems, consequently contributing to the emergence of carbapenem resistance. Carbapenem-resistant Enterobacteriaceae (CRE) refers to Enterobacteriaceae strains that are resistant to at least one carbapenem, such as ertapenem, imipenem, or meropenem. Patients who require mechanical ventilation, have urinary or intravenous catheters, are on prolonged immunosuppressive therapy, or have compromised immune systems are at increased risk for CRE infections. Identifying dominant beta-lactamase genes associated with resistance, particularly in intensive care units, will significantly contribute to empirical antibiotic therapy in critically ill patients. This thesis aims to contribute to local and national surveillance by investigating the resistance profiles of CRE isolates cultured from various clinical samples of patients hospitalized in the intensive care units of our hospital using genotypic methods. Additionally, the study seeks to determine the antimicrobial susceptibility of these isolates. Between 15.12.2023 and 15.06.2024, a total of 74 CRE isolates were included in the study, which were cultured from blood, urine, or endotracheal aspirates (ETA) of patients suspected of having ventilator-associated pneumonia (VAP), pyelonephritis, or catheter-related bloodstream infections, and who had been hospitalized for at least 72 hours in one of the tertiary internal medicine and anesthesiology and reanimation intensive care units of our hospital. The relationship between the patients' age, sex, the intensive care unit in which they were treated, prior antimicrobial therapy, accompanying comorbidities, and mortality was investigated. The presence of carbapenemase in the cultured CRE isolates was assessed using phenotypic methods. The presence of carbapenemase genes (blaOXA-48, blaNDM, blaIMP, blaVIM, and blaKPC) was investigated using PCR in 60 Klebsiella pneumoniae and 4 Escherichia coli isolates identified as carbapenemase producers. Consistent with national data, blaOXA-48 was found to be the predominant carbapenemase gene, detected in 53.1% of the isolates. No metallo-β-lactamase (MBL) producers were observed, while an increase in the frequency of blaKPC was noted (46.9%). The antimicrobial susceptibility of these isolates to ceftazidime/avibactam, meropenem/vaborbactam, imipenem/relebactam, ceftolozane/tazobactam, and cefiderocol was evaluated using the disk diffusion method. In KPC-producing isolates, susceptibility rates were as follows: ceftazidime/avibactam 83.9%, meropenem/ vaborbactam 100%, imipenem/relebactam 96.6%, and cefiderocol 93.3%, with no strains exhibiting susceptibility to ceftolozane/tazobactam. In OXA-48-producing isolates, susceptibility rates were observed at: ceftazidime/avibactam 89%, meropenem/vaborbactam 52%, imipenem/relebactam 88%, and cefiderocol 94.1%. Moreover, 17.6% of the isolates showed susceptibility to ceftolozane/tazobactam. The susceptibility of blaKPC-producing isolates to ceftazidime/avibactam, meropenem/vaborbactam, imipenem/relebactam, and cefiderocol was found to be in accordance with existing literature. In contrast, Klebsiella pneumoniae isolates harboring blaOXA-48 demonstrated a higher susceptibility to imipenem/relebactam than previously reported in the literature. Clonal relationships among Klebsiella pneumoniae isolates were analyzed using protein electrophoresis, yielding a Dice similarity coefficient of 78.6%, indicating no clonal relationship among the isolates. Key words: carbapenem-resistant enterobacteriaceae, carbapenemase detection methods, molecular epidemiology, CRE treatment.