Genome-Wide in silico analysis reveals activation of a silent resistome driving imipenem resistance in Pseudomonas aeruginosa

Pseudomonas aeruginosa is a major cause of hospital-acquired infections and is frequently associated with carbapenem treatment failure, particularly with imipenem. However, the genomic basis of imipenem resistance is still not fully clear. In this study, we analysed 95 unrelated P. aeruginosa genomes, including 41 imipenem-resistant and 54 susceptible isolates, to understand whether resistance is driven mainly by core genes or by accessory genomic elements. All genomes were re-annotated and examined using pangenome analysis, Multi Locus Sequence Typing, pangenome-wide association studies (Scoary and pyseer), and detailed profiling of blaOXA (oxacillinase beta-lactamase) beta-lactamase variants. Resistant isolates contained a much broader range of resistance genes (33 families) compared to susceptible isolates (13 families), yet core-genome phylogeny showed no clear clustering of resistant strains. Pan-GWAS identified 12 accessory genes significantly associated with resistance, including redox-related DsbA proteins, copper-resistance protein D, and mobile genetic elements. Importantly, several blaOXA variants with known carbapenemase activity were also present in susceptible isolates, indicating that these genes can remain inactive unless supported by a suitable accessory genomic background. MLST analysis showed that sequence type 233 was exclusively resistant and carried a high combined burden of resistance and virulence genes. Together, these findings support a “silent resistome activation” model, in which imipenem resistance arises through coordinated genomic support rather than simple acquisition of a single resistance gene.