High-level ceftazidime-avibactam resistance in Escherichia coli conferred by the novel plasmid-mediated beta-lactamase CMY-185 variant

ObjectivesTo characterize a blaCMY variant associated with ceftazidime-avibactam (CZA) resistance from a serially collected Escherichia coli isolate. MethodsA patient with an intra-abdominal infection due to recurrent E. coli was treated with CZA. On day 48 of CZA therapy, E. coli with a CZA MIC of >256 mg/L was identified from abdominal drainage. Illumina WGS was performed on all isolates to identify potential resistance mechanisms. Site-directed mutants of CMY {beta}-lactamase were constructed to identify amino acid residues responsible for CZA resistance. ResultsWGS revealed that all three isolates were E. coli ST410. The CZA-resistant strain uniquely acquired a novel CMY {beta}-lactamase gene, herein called blaCMY-185, harbored on an IncI{gamma}-type conjugative plasmid. The CMY-185 enzyme possessed four amino acid substitutions relative to CMY-2 including A114E, Q120K, V211S, and N346Y and conferred high-level CZA resistance with an MIC of 32 mg/L. Single CMY-2 mutants did not confer reduced CZA susceptibility. However, double and triple mutants containing N346Y previously associated with CZA resistance in other AmpC enzymes, conferred CZA MICs ranging between 4 and 32 mg/L as well as reduced susceptibility to the newly developed cephalosporin, cefiderocol. Molecular modelling suggested that the N346Y substitution confers the reduction of avibactam inhibition due to the steric hindrance between the side chain of Y346 and the sulfate group of avibactam. ConclusionWe identified CZA resistance in E. coli associated with a novel CMY variant. Unlike other AmpC enzymes, CMY-185 appears to require an additional substitution on top of N346Y to confer CZA resistance.