Characterisation of a Novel Hybrid IncFIIpHN7A8:IncR:IncN Plasmid Co-Harboring blaNDM-5 and blaKPC-2 from a Clinical ST11 Carbapenem-Resistant Klebsiella pneumoniae Strain

Abstract

Purpose We aimed to characterize a novel bla NDM-5 and bla KPC-2 co-carrying hybrid plasmid from a clinical carbapenem-resistant Klebsiella pneumoniae (CRKP) strain. Methods Antimicrobial susceptibility was determined by the broth microdilution method. Plasmid size and localization were estimated using S1 nuclease pulsed-field gel electrophoresis (S1-PFGE) and Southern blotting. Plasmid transfer ability was evaluated by conjugation experiments. Whole genome sequencing (WGS) was performed using Illumina NovaSeq6000 and Oxford Nanopore MinION platforms. Genomic characteristics were analyzed using bioinformatics methods. Results Strain ZY27320 was a multidrug-resistant (MDR) clinical ST11 K. pneumoniae strain that confers high-level resistance to carbapenems (meropenem, MIC 128 mg/L; imipenem, MIC 64 mg/L) and ceftazidime/avibactam (MIC >128/4 mg/L). Both S1-PFGE–Southern blotting and whole genome sequencing revealed that the carbapenemase genes bla KPC-2 and bla NDM-5 were carried by the same IncFII pHN7A8 :IncR:IncN hybrid plasmid (pKPC2_NDM5). Conjugation experiments indicated that pKPC2_NDM5 was a non-conjugative plasmid. Conclusion This is the first report of a hybrid plasmid carrying both carbapenemase genes bla NDM-5 and bla KPC-2 in a clinical K. pneumoniae ST11 isolate that confers resistance to both ceftazidime/avibactam and carbapenems, thereby presenting a serious threat to treatment in clinical practice.

Carbapenem-Resistant Enterobacter cloacae Complex in Southwest China: Molecular Characteristics and Risk Factors Caused by NDM Producers

“…To confirm the successful plasmid conjugation, the presumed conjugants were purified and subsequently validated using MALDI-TOF MS and PCR. 13…”