Not so innocent: Environmental Klebsiella pneumoniae disseminating resistance and virulence via conjugation.

The acquisition of carbapenem resistance and hypervirulence traits in Klebsiella pneumoniae results in the emergence of certain high-risk clones posing significant public health threat. This convergence is largely driven by conjugative plasmids, however, there are limited studies on environmental reservoirs and transmission potential of such isolates. In this study, we investigated the phenotypic trait for hypervirulence in Carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKp) isolated from a river system in Odisha, India, assessed their ability to co-transfer carbapenemase and virulence genes via conjugation, and evaluated their virulence using the Galleria mellonella model. The four environmental CR-hvKp (CR-hvKp4, CR-hvKp5, CR-hvKp9, CR-hvKp13) strains exhibited moderate biofilm formation. CR-hvKp9 displayed higher siderophore production with 73.98 PSU compared to the control clinical strain (65.63 PSU). Infection of G. mellonella larvae revealed high virulence for CR-hvKp9 and CR-hvKp13, causing ≥ 70-80% mortality by 96 h post infection. Conjugation assays confirmed the co-transfer of carbapenemase (blaNDM, blaOXA-48) along with virulence-associated genes (iucA, rmpA/rmpA2, entB, and ybtS) to E. coli J53. All transconjugants harboured IncFIB replicons. MLST analysis revealed high-risk STs, ST147, ST677, and ST437. The presence of such hypervirulent and transmissible CRKP lineages in a river system suggests that aquatic environments receiving urban sewage may serve as reservoirs and potential conduits for community-level dissemination.