Molecular Characteristics of an NDM-4 and OXA-181 Co-Producing K51-ST16 Carbapenem-Resistant Klebsiella pneumoniae: Study of Its Potential Dissemination Mediated by Conjugative Plasmids and Insertion Sequences.

The carbapenem-resistant Klebsiella pneumoniae (CRKP) strain GX34 was recovered from the respiratory tract of an elderly male with severe pneumonia, and only susceptible to amikacin, tigecycline, and colistin. Complete genome suggested that it belonged to K51-ST16 and harbored plasmid-encoded NDM-4 and OXA-181, located on IncFIB plasmid GX34p1_NDM-4 and ColKP3/IncX3 plasmid GX34p4_OXA-181, respectively. A series of transconjugants generated in the plasmid conjugation assays, including Escherichia coli J53-N1 (harboring a self-transmissible and bla NDM-1 -producing plasmid Eco-N-1-p), J53-N2 (harboring a bla NDM-4 -producing plasmid and a helper plasmid GX34p5), and J53-O (harboring a bla OXA-181 -producing plasmid), could be stably inherited after 10 days of serial passage and no significant biological fitness costs were detected. Furthermore, we first reported the bla NDM-1 gene, derived from bla NDM-4 mutation (460C>A) under meropenem pressure, could be in vitro transferred into a self-conjugative, recombined plasmid Eco-N-1-p of J53-N1. Eco-N-1-p was mainly recombined by GX34p4_OXA-181 (40,449 bp, 75.16%) and GX34p1_NDM-4 (8,553 bp, 15.89%), in which IS 26 and IS 5-like probably played a major role. Eco-N-1-p could be transferred into the conjugation recipient K. pneumoniae KP54 and make the latter sacrifice fitness. The retention rates of bla NDM-1 remained high stability (>80% after 200 generations). The comparative genomic analysis of GX34 and those carrying bla NDM-4 or bla OXA-181 genes retrieved from the NCBI RefSeq database showed all bla NDM-4 (26/26, 100.00%) and bla OXA-181 (13/13, 100.00%) were surrounded by IS 26 . The immediate environment of bla NDM-4 and bla OXA-181 in GX34 and some retrieved strains shared identical features, hinting at their possible dissemination. Effective measures should be taken to monitor the spread of this clone.