Localized glutamine leakage drives the spatial structure of root microbial colonization

Plant roots release exudates to encourage microbiome assembly, which influences the function and stress resilience of plants. How specific exudates drive spatial colonization patterns remains largely unknown. In this study, we demonstrate that endodermal Casparian strips—forming the root’s extracellular diffusion barrier—restrict nutrient leakage into the rhizosphere, coinciding with and controlling spatial colonization patterns of rhizobacteria. We find that vasculature-derived glutamine leakage is a major bacterial chemoattractant and enhancer of proliferation, defining a previously unknown pathway for root exudate formation. Bacteria defective in amino acid chemoperception display reduced attraction toward leakage sites, and roots with Casparian strip defects display bacterial overproliferation, dependent on bacterial capacity for amino acid metabolization. Associated chronic immune stimulation suggests that endodermal nutrient restriction is crucial for regulating microbial colonization and assembly, limiting excessive proliferation that could compromise plant health.