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Pseudomonas aeruginosa is one of the most common healthcare-associated pathogens worldwide, and infections are difficult to treat due to intrinsic resistance and emerging resistance to carbapenem antibiotics. P. aeruginosa is frequently recovered from wet niches in hospitals, such as sink drains, where it may form persistent reservoirs that are impossible to properly clean. In this thesis, carbapenem-resistant, high-risk clones of VIM-2-positive P. aeruginosa (VIM-PA) were investigated with a focus on their presence, persistence, and transmission in hospital environments. Firstly, a national surveillance pilot study to identify carbapenemase-producing P. aeruginosa in the Netherlands was described, showing that VIM-PA high-risk clone ST111 had spread unnoticed within many Dutch hospitals. Focus was then narrowed to the Erasmus MC: The microbial compositions of sink drain reservoirs were characterized using culturomics to find microorganisms that might resist colonization by VIM-PA; VIM-PA hiding intracellularly within Acanthamoeba castellanii was discussed as a method VIM-PA may employ to evade disinfection in water systems; sink design factors contributing to dispersal from drains were studied, and adjustments to minimize splashing (e.g. installing sink drain plugs, offsetting the faucet) were recommended; finally, whether Pseudomonas spp. carried blaVIM-2-encoding plasmids in our hospital, and the role of sink drain reservoirs in blaVIM-2 transmission, were explored. The strategies proposed in this thesis for minimizing transmission from sink drain reservoirs and managing VIM-PA colonization in the environment pave the way toward making safer patient spaces in hospitals.

Background In healthcare environments, sinks are being increasingly recognized as reservoirs for multidrug- resistant Gram-negative bacteria. In our hospital, carbapenemase-producing, Verona Integron-encoded Metallo-beta-lactamase (VIM)-positive Pseudomonas aeruginosa (VIMPA) was detected at low endemicity in patients, and environmental culturing revealed that sink drains were primary reservoirs. Therefore, an intervention was initiated in several wards to install sink drain plugs as physical barriers against splashing to prevent transmission of VIM-PA from drain reservoirs to the surrounding sink environment. Aim To assess the efficacy of the intervention on limiting spread of VIM-PA. Methods Swabs were taken from inner sink environments (i.e. drains), and outer sink environments (i.e. wash basins, faucet aerators, and countertops) twice before and three times after the intervention. Siphon water and drain wells were also sampled before and at the moment of the intervention, respectively. All samples were screened for VIM-PA, and isolates were typed with multiple-locus variable-number tandem repeat analysis (MLVA). Results There was a significant reduction in VIM-PA positivity in both inner (P-value <0.001) and outer (P-value 0.001) sink environments after the intervention. However, VIM-PA recolonization was observed in the inner sink environments of patient rooms, and also in rooms exclusive to healthcare personnel, over time. Surfaces in the outer sink environment were rarely positive for VIM-PA after the intervention. MLVA revealed three genetic clusters, with one found in all wards and room types during the study period. Conclusions Drain plugs are a simple and effective infection prevention and control measure to contain spread of VIM-PA from drain reservoirs.