Isolation and characterization of a multidrug-resistant carbapenem-resistant Klebsiella pneumoniae strain co-carrying IncFII/ IncR-KPC-2 and phage-like plasmids

Objectives: The aim of this study was to characterize the co-occurrence of the blaSHV-182, blaKPC-2, blaCTX-M65, and blaTEM-1B genes in a multi-resistant carbapenem carbapenem-resistant K. pneumoniae (CRKP) strain. Methods: The multi-resistant CRKP strain was isolated from the bronchoalveolar lavage fluid (BAL) of a patient with acute exacerbation of chronic obstructive pulmonary disease (AECOPD), after cultured in lysogeny broth, the colonies with apparent phenotypic differences (size, shape, color, and surface states) were picked out for species identification by sequencing their 16S rDNA and BLASTN in the NCBI database, antimicrobial susceptibility was determined, Genomic DNA of overnight cultured K. pneumoniae D1a was extracted by using the Bacterial DNA Isolation Kit, the library was constructed by using NEBNext (R) UltraTM DNA Library Prep Kit for Illumina, and the whole-genome sequencing was performed on the Illumina HiSeq PE150 platform. The pair-end reads were assembled by SOAP denovo v2.04, SPAdes, and ABySS. Genome characteristics were analyzed by using bioinformatics methods. Results: The WGS of the multi-resistant CRKP strain D1a (CRKP D1a) yielded 84 scaffolds of genomic DNA (5, 492, 035 bp) and two circular plasmids, pD1a1 (173, 413 bp) and pD1a2 (57, 686 bp). CRKP D1a was classified as the sequence type 11, and harbored 16 antibiotic resistance genes and 30 multidrug efflux pump-encoding genes associated with the resistance to almost all kinds of commonly used antibiotics. The plasmid pD1a1 belonged to the IncFII/IncR hybrid plasmid family with blaSHV-182, blaKPC-2, blaCTX-M-65, and blaTEM-1B genes flanked by the IS6-like element. The plasmid pD1a2 was comprised of 95 phage-related genes but had no lysis activity on the tested K. pneumoniae strains. Conclusion: This study reports the isolation of a multidrug-resistant CRKP strain co-carrying IncFII/IncR-KPC2 plasmid and phage-like plasmid, the findings hold significant implications for further investigating the resistance mechanism and molecular diversity of CRKP.