Distinct effects of GES variants on ceftazidime-avibactam and imipenem-relebactam resistance in carbapenem-resistant <i>Pseudomonas aeruginosa</i> : a ten-year retrospective study

Ceftazidime-avibactam (CZA) and imipenem-relebactam (IMR) are β-lactam/β-lactamase inhibitor combinations active against carbapenem-resistant Pseudomonas aeruginosa (CRPA), yet resistance to these agents is increasingly reported. In this study, 916 CRPA isolates collected from 2016 to 2025 were screened for blaGES by PCR, and fifty isolates were identified as blaGES-positive. Five subtypes were detected, including GES-1 (68%), GES-5 (24%), GES-7 (2%), GES-15 (4%), and a novel variant, GES-69 (2%). Antimicrobial susceptibility testing by broth microdilution revealed distinct resistance patterns. Isolates carrying blaGES-1, blaGES-7, blaGES-15, and blaGES-69 showed elevated CZA MICs, whereas blaGES-5 was associated with increased IMR MICs. Cloning experiments and structural modeling suggested that substitutions at position 170, Ω-loop conformational changes, and alterations in residues such as E104 may affect substrate binding and inhibitor interactions. Whole-genome sequencing showed that blaGES variants were located on the chromosome within class 1 integrons embedded in Tn6584-like transposons. Analysis of publicly available genomes further indicated that blaGES-positive isolates were mainly associated with the high-risk clone ST235, were distributed across multiple Gram-negative species with P. aeruginosa as the predominant host and exhibited an expanding cross-species distribution. In summary, these findings demonstrate that subtype-specific variation in GES enzymes is likely to shape the activity of β-lactam/β-lactamase inhibitor combinations within a conserved ST235 background.