Molecular Characterization of a KPC-2- and NDM-1-Producing Klebsiella michiganensis Clinical Isolate in Cerebrospinal Fluid.

Abstract

Klebsiella michiganensis is an emerging pathogen. In this context, we characterised a strain fxq isolated from a cerebrospinal fluid specimen of a patient with tentorial meningioma, and the K. michiganensis isolate produced carbapenemases of KPC and NDM types. The Phoenix 100 Automated Microbiology System, MALDI-TOF and whole-genome sequencing were used to identify the species. Anti-microbial susceptibility testing was also conducted with the Phoenix 100. The plasmid locations of the bla KPC-2 and bla NDM-1 genes were determined by S1-nuclease pulsed-field gel electrophoresis and Southern blot. The transfer capacity of plasmids carrying bla KPC-2 and bla NDM-1 was investigated by conjugation experiments, and the resistance plasmid stability was evaluated by culture and subculture. K. michiganensis subtypes were identified by multi-locus sequence typing. We performed whole-genome sequencing to confirm species, characterise plasmids and analyse core genes. fxq was originally identified as Klebsiella oxytoca and showed resistance to imipenem and meropenem, but whole-genome sequencing identified it to be K. michiganensis. The strain fxq belonged to the novel sequence type 202 (ST202) and carried the bla KPC-2 and bla NDM-1 genes located on the pB_KPC InFIA and pE_NDM IncU plasmids, respectively. The bla KPC-2-carrying plasmid was successfully transferred to Escherichia coli EC600 by conjugation, whereas the bla NDM-1 gene on the pE_NDM plasmid was not. The pB_KPC and pE_NDM plasmids demonstrated high stability. This work is the first report on a carbapenem-resistant clinical isolate K. michiganensis ST202 harbouring the bla KPC-2 and bla NDM-1 genes encoded by the IncFIA and IncU plasmids, respectively.