OXY-2-15, a novel variant showing increased ceftazidime hydrolytic activity

Abstract

Objectives: Klebsiella oxytoca is a member of the family of Enterobacteriaceae and often contains the β-lactamase blaOXY gene. Although this β-lactamase does not naturally hydrolyse ceftazidime, this study describes possible in vivo selection of a clinical K. oxytoca isolate showing increased MICs of ceftazidime.

Methods: To reveal the molecular mechanism underlying this unusual resistance phenotype, WGS, cloning, overexpression, MIC and steady-state kinetic studies were performed.

Results: A patient was treated for a septic episode with ceftazidime (4 g/day). This therapy was based on earlier culture results in which, amongst others, a K. oxytoca (Velp-1) isolate was identified. After 11 days of treatment, K. oxytoca Velp-2 was isolated from a pus sample drained from the wound. The isolate showed increased resistance to ceftazidime (MIC ≥64 mg/L) compared with the original K. oxytoca isolate (Velp-1). WGS revealed the presence of a novel blaOXY-2 allele, designated blaOXY-2-15, with a two amino acid deletion at Ambler positions 168 and 169 compared with OXY-2-2. Cloning blaOXY-2-15 into Escherichia coli TOP10 resulted in increased MICs of ceftazidime, but reduced MICs of most other β-lactams compared with OXY-2-2. Steady-state kinetics confirmed the results of the MIC data, showing clearly significant ceftazidime hydrolysis.

Conclusions: This report shows the risk of in vivo selection of ceftazidime-resistant K. oxytoca isolates after prolonged ceftazidime treatment. Furthermore, it is the first known report of a K. oxytoca isolate conferring resistance to ceftazidime by a two amino acid deletion in the omega loop of OXY-2-2.

Keywords: ESBLs; Enterobacteriaceae; Gram-negative; K1; KOXY; antimicrobial resistance mechanisms; bla genes.