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Chun-Hsing Liao,1,2 Na-Yao Lee,3 Hung-Jen Tang,4,5 Susan Shin-Jung Lee,6,7 Chin-Fu Lin,8 Po-Liang Lu,9–11 Jiunn-Jong Wu,12 Wen-Chien Ko,13 Wen-Sen Lee,14 Po-Ren Hsueh15,16 1Department of Internal Medicine, Far Eastern Memorial Hospital, Taipei, Taiwan; 2Department of Medicine, Yang-Ming University, Taipei, Taiwan; 3Department of Internal Medicine, National Cheng Kung University Medical College and Hospital, Tainan, Taiwan; 4Department of Internal Medicine, Chi Mei Medical Center, Tainan, Taiwan; 5Department of Health and Nutrition, Chia Nan University of Pharmacy and Science, Tainan, Taiwan; 6Department of Internal Medicine, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan; 7Faculty of Medicine, School of Medicine, National Yang Ming University, Taipei, Taiwan; 8Department of Pathology and Laboratory Medicine, Taichung Veterans General Hospital, Taichung, Taiwan; 9Department of Internal Medicine, Kaohsiung Medical University Hospital, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; 10Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan; 11Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan; 12Department of Biotechnology and Laboratory Science in Medicine, National Yang-Ming University, Taipei, Taiwan; 13Department of Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan; 14Division of Infectious Diseases, Department of Internal Medicine, Wan Fang Medical Center, Taipei Medical University, Taipei, Taiwan; 15Department of Laboratory Medicine, National Taiwan University Hospital, National Taiwan University College of Medicine, Taipei, Taiwan; 16Department of Internal Medicine, National Taiwan University Hospital, National Taiwan University College of Medicine, Taipei, Taiwan Objective: The aim of this study was to investigate the in vitro antimicrobial susceptibilities of clinically important Gram-negative bacteria from seven intensive care units in Taiwan in 2016. Materials and methods: In total, 300 non-duplicate isolates of Escherichia coli (n=100), Klebsiella pneumoniae (n=100), and Pseudomonas aeruginosa (n=100) collected from 300 patients were studied. The minimum inhibitory concentrations (MICs) of these isolates to antimicrobial agents were determined using the broth microdilution method. Carbapenemase-encoding genes (blaKPC, blaNDM, blaIMP, blaVIM, and blaOXA-48-like) were studied for the isolates that were not susceptible to any carbapenems. Sequencing analysis of the mcr genes (mcr-1–5) was conducted for all isolates with colistin MICs ≥4 mg/L. Results: Ertapenem non-susceptibility was detected in 3% (n=3) E. coli and 12% (n=12) K. ­pneumoniae isolates. The susceptibility rates of imipenem, ceftazidime–avibactam (CAZ–AVB), and ceftolozane–tazobactam (CLZ–TAZ) were 99%, 99%, and 88%, respectively, for E. coli, 91%, 100%, and 80%, respectively, for K. pneumoniae, and 66%, 91%, and 93%, respectively, for P. aeruginosa. Carbapenemase-encoding genes were not detected in E. coli, were detected in four (33.3%) K. pneumoniae isolates that were not susceptible to ertapenem (three harboring blaKPC and one harboring blaOXA-48-like), and were not detected in P. aeruginosa isolates that were not susceptible to imipenem. One K. pneumoniae isolate was resistant to colistin (MIC 4 mg/L) and negative for mcr genes. Conclusion: CAZ–AVB exhibited excellent activity against carbapenem-resistant Enterobacteriaceae, and CLZ–TAZ exhibited good activity against imipenem-resistant P. aeruginosa. Keywords: carbapenem resistance, second-generation, β-lactam, β-lactamase inhibitor combinations, carbapenemase-encoding genes, mcr