Spatiotemporal mapping reveals microbiota-dependent maintenance of functional competence during postnatal immune maturation.

Postnatal immune maturation is commonly viewed as a developmental milestone associated with weaning. However, how postnatal microbiota exposure shapes immune cell heterogeneity across tissues, and whether this maturation is self-sustaining or requires continuous microbial input, remain unclear. Here, we conducted spatiotemporal single-cell transcriptomic profiling of 120,649 immune cells isolated from 10 tissues in both germ-free (GF) and specific pathogen-free (SPF) mice from weaning to adulthood. We demonstrate that microbial colonization drives the expansion and functional maturation of distinct monocyte, macrophage, and neutrophil populations, marked by enhanced antigen-processing capabilities across tissues. Simultaneously, commensal-derived signals reprogram lymphocyte metabolic and translational states, promoting oxidative phosphorylation and relieving PDCD4-mediated translational repression to sustain effector readiness. Notably, microbiota reconstitution through both co-housing and fecal microbiota transplantation demonstrated that immune competence is not a fixed developmental endpoint but instead requires continuous microbiota-derived input. Without sustained microbial signals, immune cells adopted a persistent hyporesponsive state rather than remaining developmentally immature. Together, these findings redefine immune maturation as a microbiota-dependent maintenance process, revealing potential strategies to restore immune competence through microbiota-targeted interventions.