Distribution of carbapenem resistance mechanisms in clinical isolates of XDR Pseudomonas aeruginosa Abstract Our study aims to define the epidemiology of carbapenem resistance mechanisms in clinical isolates of Pseudomonas aeruginosa (PA). We evaluated 11,457 clinical PA strains isolated between 2009 and 2015 at the tertiary care University Hospital in Heidelberg, Germany. Thirty-four percent of the isolates (3867/11,457) were MDR (multidrug-resistant), 16% (1816/11,457) were XDR (extensively drug resistant), and less than 1% (82/11,457) had a PDR (pandrug-resistant) profile. Of those, 23% carried a carbapenemase gene (CPM positive) with 12% VIM-2, 10% VIM-1, and less than 1% IMP-1. Comparing MIC (minimal inhibitory concentration) distributions, the mean rank for meropenem, imipenem, gentamicin, and fosfomycin was significantly higher in the CPM-positive group than in the CPM-negative XDR group (p ≤ 0.004). oprD (outer membrane protein) mutations were found in 19/19 tested strains; 12/19 carried a CPM and had a higher mutation rate. Meropenem resistance was mostly associated with the presence of CPM. Only 1/19 strains was meropenem resistant in the absence of CPM genes; nevertheless, it carried an oprD mutation in a strategic site (loop 2). Of 19 CPM-negative strains tested, 7 (36%) showed EP (efflux pumps) hyperexpression versus 12 in the CPM-positive strains. In our study, nearly 50% of the PA isolates exhibited resistance to the tested first-line antibiotics. Our study also demonstrates that carbapenemase genes can be isolated in approximately 23% of XDR PA strains in our population. This finding supports the clinical relevance of PA driven by the possible presence of multiple resistance mechanisms acquired under exposure to antibiotics or by horizontal transfer of resistance genes.