Genomic and Phenotypic Analysis of Persistent Carbapenem-Resistant Klebsiella pneumoniae Isolates from a 5-Year Hospitalized Patient

The emerging epidemic of carbapenem-resistant Klebsiella pneumoniae (CRKP) is a global public health crisis. However, the phylogenetic affiliation and pathotypic status of CRKP strains in the host colonization period under consistent antibiotic treatments are not well characterized. In this study, a 5-year tracking study was performed, in which a patient admitted to an intensive care unit was recruited and then screened for the carriage of CRKP based on microbiological culture. Nine isolates from the sputum or stool samples were acquired and subjected to real-time whole-genome sequencing, antimicrobial susceptibility testing, Galleria mellonella larval infection, and epithelial cell invasion assay. All nine isolates showed phenotypic resistance to carbapenems, quinolones, and aminoglycosides. Altogether, blaKPC-2 and 10 other antibiotic resistance genes were identified and all nine CRKP isolates exhibited low virulence with more than 38 virulence factors. All but one variant belonged to ST11 with a novel sequence type, differing at the phoE locus. The isolates shared the same plasmid replicon type, prophages, and capsular serotype (K47) with few single-nucleotide polymorphism variations, consistent with epidemiological clones. Furthermore, these CRKP isolates displayed the ability of moderate invasion of lung epithelial cells. Meanwhile, a deficiency of chromosomal type IV secretion system-related gene cluster was detected after 2 years of carriage. Our findings demonstrated that low-virulence CRKP clones could colonize in the gut and respiratory tract under multiple antibiotic stresses, suggesting the strong colonization adaptability of CRKP to the host. Ethical approval was given by The Second Affiliated Hospital of Zhejiang University, School of Medicine, (2018-039).