Emergence of a Superplasmid Coharboring Hypervirulence and Multidrug Resistance Genes in Klebsiella pneumoniae Poses New Challenges to Public Health.

The emergence of plasmids coharboring hypervirulence (Hv) and multidrug resistance (MDR) genes has further accelerated the spread of MDR-Hv Klebsiella pneumoniae (MDR-HvKP) strains, having a severe impact on public health. Here, we report an MDR-Hv superplasmid coharboring hypervirulence and MDR genes and the detailed characterization of its genetic and phenotypic features. This plasmid was identified in an ST11 (sequence type 11)-K64 carbapenem-resistant hypervirulent K. pneumoniae (CR-HvKP) strain, SZS128, which was responsible for a bloodstream infection in a 21-year-old female. Susceptibility testing showed that SZS128 was resistant to amikacin, levofloxacin, and almost all of the β-lactams examined. SZS128 showed high virulence in a Galleria mellonella survival assay and a mouse intraperitoneal infection model. Genomic analysis showed that SZS128 not only possessed a KPC plasmid (pSZS128-KPC) but also carried a superplasmid (pSZS128-Hv-MDR) coharboring hypervirulence and MDR genes and possessing complete conjugative regions. Conjugation and transformation assays confirmed the potential for horizontal transfer and the high stability (retention rate of >95%) of the pSZS128-Hv-MDR superplasmid. Furthermore, growth curve assessment confirmed that there was no increase in the fitness cost in the presence of pSZS128-Hv-MDR. Therefore, we define a superplasmid as a plasmid fulfilling all the following criteria: (i) a single plasmid that coharbors hypervirulence and MDR genes, (ii) a plasmid that harbors complete conjugative elements that guarantee self-transmissibility, (iii) a plasmid that is stable and conserved, and (iv) a plasmid with no fitness cost to the host strain. The emergence of this kind of superplasmid could represent a serious threat to public health, and urgent control measures must be implemented. This self-transmissible superplasmid, which simultaneously carries hypervirulence and MDR genes, greatly enhances the challenges to clinical prevention and control and anti-infection treatment. Thus, active surveillance of this type of superplasmid is needed to prevent these efficient resistance/virulence plasmids from disseminating in hospital settings. Our findings provide a reference for defining the term "superplasmid" and emphasize the importance of raising public awareness of the rapid dissemination of this self-transmissible superplasmid and the consistent emergence of MDR-HvKP strains.