Mechanisms underlying altered ceftazidime-avibactam resistance in Klebsiella pneumoniae driven by KPC-234 mutation and IncX3-NDM-1 plasmid transfer.

OBJECTIVE

Ceftazidime-avibactam is commonly used to treat infections caused by Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae, but its effectiveness is decreasing due to emerging resistance mechanisms. This study aimed to investigate the genetic basis of ceftazidime-avibactam resistance in serial K. pneumoniae isolates recovered from a single patient.

METHODS

Four KPC-producing K. pneumoniae strains were isolated from one patient during treatment. Whole-genome sequencing, plasmid transformation, gene cloning, and enzymatic kinetic assays were performed. Plasmid typing, transposon structure analysis, and mutation screening of blaKPC variants were conducted to characterize resistance determinants.

RESULTS

The identified mechanisms of resistance included the horizontal transfer of an IncX3_NDM-1 plasmid and mutations in blaKPC-2, which resulted in the formation of blaKPC-234. We demonstrated that two single-nucleotide mutations in blaKPC-2 resulted in amino acid substitutions (D179G and Y241D), which altered the enzyme's structure. These mutations conferred resistance to ceftazidime-avibactam by increasing the minimum inhibitory concentration by 64-fold. Whole-genome sequencing revealed that the blaKPC-234 gene was located on an IncFII/IncR-type plasmid in KP57128, containing a composite transposon structure (IS26-blaSHV-12-TnAs1-ISKpn6-blaKPC-234-ISkpn27-tnpR2-IS26-blaCTX-M-65-IS903-IS26). Additionally, the acquisition of the IncX3 plasmid carrying blaNDM-1 enabled the horizontal transfer of resistance genes, further enhancing bacterial resistance to multiple antimicrobial agents.

CONCLUSIONS

This study emphasizes the critical role of genetic mutations and plasmid-mediated gene transfer in the evolution of carbapenem-resistant K. pneumoniae resistance, underscoring the importance of continuous monitoring of resistance changes, particularly those involving KPC gene mutations.