Acquisition of the Conjugative Virulence Plasmid From a CG23 Hypervirulent Klebsiella pneumoniae Strain Enhances Bacterial Virulence

Abstract

The emergence of hypervirulent and carbapenem-resistant Klebsiella pneumoniae (hv-CRKP) has become a hot topic and confounding problem for clinicians and researchers alike. Conjugative virulence plasmids have the potential to cause more threatening dissemination of hv-CRKP strains. We previously identified K2606, a CG23 clinical hypervirulent strain of Klebsiella pneumoniae harboring a conjugative virulence plasmid designated pK2606. In this study we examined hypervirulence levels using assays of biofilm formation, serum resistance, and wax larvae and mouse in vivo infection models. Moreover, to define the transfer ability of pK2606 and whether this confers hypervirulence to other strains we performed plasmid transconjugation experiments between K2606 and the ST11 CRKP strain HS11286 along with E. coli J53. We found that although biofilm formation and serum resistance were not significantly increased, the transconjugants acquired the ability of produce high level of siderophores and also caused high mortality of wax larvae and mice. Furthermore, we identified pK2606-like conjugative virulence plasmids in GenBank, providing evidence that such plasmids may have begun to spread throughout China. These findings provide an evidence base for the possible mechanisms of the emergence of hv-CRKP strains and highlight the potential of pK2606-like conjugative virulence plasmids to spread worldwide.

Co-Occurrence of Rare ArmA-, RmtB-, and KPC-2–Encoding Multidrug-Resistant Plasmids and Hypervirulence iuc Operon in ST11-KL47 Klebsiella pneumoniae

“…In addition to the resistance elements, we also observed that a key virulent factor, the iutA-iucABCD operon, was coharbored with armA , bla TEM-1B , aac(3)-IId , dfrA12 , aadA2 , sul1 , msr(E) , mph(E) , q nrB4 , and bla DHA , on the p1864-1 plasmid ( K. pneumoniae 1864). We also detected the siderophore production of K. pneumoniae 1864 and 1632 to determine the virulence contribution of the iutA-iucABCD operon, which was highly related to K. pneumoniae virulence phenotype ( 14 ), in K. pneumoniae 1864. We found a significant increase of siderophore production in K. pneumoniae 1864, compared with 1632, which did not harbor the iuc operon.…”

“…The chrome azurol S (CAS) assay was performed as previously described to determine the iron-chelating capabilities of bacterial supernatants ( 14 ). Briefly, the stationary-phase iron-chelated cultures (10 μL) were dropped on CAS plates, and siderophore production was determined by the orange halos after incubation for 48 h at 37°C.…”

Prevalence of hypervirulent and carbapenem-resistant Klebsiella pneumoniae under divergent evolutionary patterns

“…The relative quantitative siderophore production was assayed via Chrome Azurol S (CAS) Assay as described in our previous study [ 30 ]. Siderophore units (Su) were used to quantify the siderophore production [ 31 ].…”

“…G. mellonella larvae used in this study weighed about 200∼400 mg and were purchased from the Tianjin Huiyude Biotech Company. The infection experiments were performed as described in our previous study [ 30 ], except that the bacterial inoculum employed was adjusted to 1×10 5 CFU. The experiments were repeated three times.…”

“…Six-week-old male BALB/c mice were purchased from the Shanghai Medical Laboratory Animal Centre (Shanghai, China). The infection experiments were performed as described in our previous study [ 30 ], except that the bacterial inoculum employed was adjusted to 5×10 6 CFU. The experiments were repeated twice.…”