Phenotypic and molecular identification of carbapenemase-producing Enterobacteriaceae - challenges in diagnosis and treatment

Introduction: Infections due to carbapenemase-producing carbapenem-resistant Enterobacteriaceae (CPCRE) are an emerging global public health threat. The purpose of this study was to investigate phenotypic and genotypic features of CP-CRE strains isolated from hospitalized patients. Material and methods: Between 1st of January - 1st of July 2017, in the Department of Microbiology, “Dr. Constantin Opriş” County Emergency Hospital Baia Mare, Romania, 1110 strains of Enterobacteriaceae were isolated from bronchial secretions, urine, wounds and blood cultures. Bacterial identification and antimicrobial susceptibility tests were performed by conventional methods, Vitek 2 Compact and M.I.C.E. strips. We analysed all Enterobacteriaceae strains non-susceptible to carbapenems according to CLSI 2017 criteria. The modified Hodge test (MHT), the modified carbapenem inactivation method (mCIM) and the combination disks test (KPC, MBL, OXA-48 Confirm kit, Rosco Diagnostica) were used for phenotypic confirmation, whereas a multiplex PCR assay for genes blaKPC, blaNDM and blaOXA-48 was used for genetic confirmation. Results: 19 non-duplicate strains isolated from 16 patients were phenotypically identified as CP-CRE: Klebsiella pneumoniae (n=14), Escherichia coli (n=2), Providencia stuartii (n=2) and Serratia marcescens (n=1). Most strains were isolated from bronchial secretions (n=9). The carbapenem-hydrolizing enzymes were identified by the combination disks test as: KPC (n=9), OXA-48-like (n=5) and MBL (n=5). Molecular confirmation was performed in 18 phenotypically positive isolates with 100% concordant results with mCIM and combination disks test. Discrepant results were noticed with the MHT in case of 4 NDM-producers confirmed by PCR. All CP-CRE strains were resistant to all tested cephems. Three out of 9 K. pneumoniae strains tested against colistin were found resistant. Conclusions: The most common carbapenemase detected was KPC. Therapeutic options were limited in all positive cases. Rapid and reliable detection of CP-CRE is critical for preventing the spread of these pathogens