Defining nutrient dependencies of intestinal Klebsiella pneumoniae guides rational probiotic engineering
Klebsiella pneumoniae is a major cause of antibiotic-resistant infections, yet the ecological determinants enabling its intestinal expansion remain poorly defined. Here, we combined in vivo transcriptomics and metabolomics to identify carbon sources exploited by K. pneumoniae during colonization of the antibiotic-disrupted gut across strains and dietary contexts. Despite its metabolic breadth, K. pneumoniae consistently exploited a restricted set of substrates, including arabinose, raffinose, oligogalacturonate, arabitol, xylose, and gluconate, that contributed to intestinal fitness, with related metabolic signatures also observed in antibiotic-treated human samples. Guided by these findings, we engineered Escherichia coli Nissle to enhance consumption of these nutrients and assembled a defined consortium, NICHE, designed to reinforce colonization resistance through nutrient competition. NICHE restricted intestinal expansion of multidrug-resistant and hypervirulent K. pneumoniae in preventive and therapeutic settings, with efficacy influenced by dietary nutrient availability. Together, this work defines exploitable K. pneumoniae nutrient dependencies and a framework for rational probiotic engineering.