Environment-mediated metabolic constraints shape antimicrobial resistance evolution in Pseudomonas aeruginosa
Pseudomonas aeruginosa rapidly evolves resistance to multiple antibiotics, complicating treatment of chronic infections. While resistance mechanisms are well described, the influence of host-relevant environmental conditions on resistance evolution remains unclear. Here, we investigated how nutrient composition, pH, and oxygen availability influence resistance evolution to piperacillin, tobramycin, and ciprofloxacin. We evolved P. aeruginosa in different synthetic media mimicking different infection niches as well as variations in pH and oxygen availability. We also performed whole-genome sequencing to identify mutational adaptive strategies. Our findings reveal that environmental context profoundly affects both the rate and mechanisms of resistance evolution. Nutrient-rich conditions and aerobic metabolism favored canonical resistance mutations, while nutrient-limited or anaerobic environments promoted adaptations involving efflux regulation, metabolic remodeling, and biofilm-associated traits. pH and oxygen availability acted as selective pressures, modulating which pathways were accessible as well as the stability of resistance phenotypes. Additionally, the mutations that arose in the lab-evolved isolates were also actively maintained across a large population of clinical P. aeruginosa genomes, confirming that spatial oxygen gradients in host tissues drive parallel evolutionary pathways in vivo. These results underscore that resistance evolution is not uniform but highly contingent on the metabolic and redox environment. Understanding how infection-specific conditions shape evolutionary trajectories provides insight into the emergence of multidrug resistance and highlights potential avenues for therapeutic intervention.