Molecular mechanisms and virulence of colistin-resistant klebsiella pneumoniae

Background: Colistin is a last-line agent for treating severe multidrug resistant Klebsiella pneumoniae ( Kp ) infections. Recent studies highlight the emergence of colistin resistant Kp arising from inducible inactivation of the mgrB gene, a negative PhoPQ signalling system regulator. Objectives: To determine the molecular mechanisms of colistin resistance and its impact upon Kp fitness and virulence. Methods: We constructed a mgrB deletion Kp mutant. Mass spectrometry (MS) was used to identify lipid-A modifications informing the generation of additional mutants in associated genes. Wild-type and mutant strains underwent MS, polymyxin and antimicrobial peptide (AMP) susceptibility, transcriptional, biofilm, growth curve and virulence ( Galleria and mouse) analyses. Results: The mgrB mutant showed a 128-fold decrease in colistin susceptibility in a PhoPQ dependant manner. We also observed enhanced resistance of the mutant to several human AMPs. Mutant lipid-A modifications included 2-hydroxymyristate, phosphoethanolamine, aminoarabinose and palmitate. Upregulation of the phoPQ , lpxO , pmrC , pmrH and pagP genes governing these modifications was confirmed. Growth fitness and biofilm forming capacity of the mutant and wild-type strains were similar. In contrast, the mutant exhibited hypervirulence in the virulence models. Complementation of the mgrB gene restored all phenotypes to wild-type levels. Conclusion: Our data highlight the importance of several novel lipid-A modifications linked with mgrB inactivation in Kp . We also show for the first time that mgrB inactivated Kp displays enhanced virulence. Overall, these data have important implications for patient management and antimicrobial stewardship.