Xu et al. report that intratumoral Citrobacter enrichment correlates with poor overall survival in pancreatic cancer patients. An intratumoral Citrobacter freundii strain metabolizes 5-fluorouracil (5-FU) into inactive products via PreTA. Gimeracil, an inhibitor of human dihydropyrimidine dehydrogenase, preserves 5-FU efficacy by blocking PreTA activity.