Nutrients drive the antibiotic-specific evolution of resistance in Pseudomonas aeruginosa

Pseudomonas aeruginosa can exploit its metabolic flexibility during cystic fibrosis lung infections to reduce antibiotic sensitivity and offset resistance costs, traits that influence its evolutionary trajectory. Although both traits are linked to nutrient conditions, their role in resistance evolution remains poorly defined. We examined how single-nutrient conditions influence resistance evolution in P. aeruginosa through phenotypic and genotypic adaptations after adaptive laboratory evolution with different antibiotics in single-nutrient media. Antibiotic susceptibility testing showed limited MIC differences for ceftazidime and imipenem, but stronger effects for ciprofloxacin, colistin, and tobramycin. Ciprofloxacin evolution in glutamate medium yielded the highest MIC increase, with at least a 4-fold rise, whereas tobramycin evolution in glucose resulted in up to a 4-fold MIC reduction compared to lineages evolved under all other nutrient conditions for the same antibiotic. Whole-genome sequencing showed nutrient-specific mutation in wbpL after tobramycin evolution in glucose, and fusA and pmrB across conditions. Ciprofloxacin resistance in glutamate-lineages involved yicC, whereas nfxB mutations were absent in glucose- and arginine-evolved lineages. No distinct nutrient-specific differences were seen for colistin. These findings underscore the significant role of nutrient conditions in shaping resistance and highlight the need to consider physiologically relevant media when studying antibiotic resistance evolution.