Spatial transcriptomics of pediatric tuberculosis lymph nodes reveals a conserved hypoxia_SPP1 niche

Pediatric tuberculosis (TB) often presents as primary disease involving the mediastinal lymph nodes; however, its granuloma microenvironment remains poorly characterized because these population patient tissues are rarely available for study. Here, we use spatial transcriptomics to profile 60 granulomas from four children aged 4 to 36 months. We identified nine recurrent transcriptional microenvironments shared across granulomas, despite their marked histological heterogeneity. One macrophage-rich microenvironment showed strong hypoxia-associated activity and secreted phosphoprotein 1 (SPP1) enrichment. This microenvironment was present in both non-necrotic and necrotic lesions and overlapped with a Mtb-persistence macrophage signature previously identified in murine TB lymph nodes. Pathway analysis revealed the enrichment of both antimicrobial and regulatory programs within this microenvironment, suggesting partial immune control rather than uniform inflammatory or suppressive activity in this region. SPP1 expression was associated with regions enriched for both M1- and M2-like macrophages, suggesting a macrophage program that extends beyond conventional polarization classifications and may be influenced by the local granuloma microenvironment. We further observed a similar spatial association between hypoxia-associated activity and SPP1 expression in an independent adult TB dataset (pooled Spearman ρ=0.65) and validated this association at protein level by multiplex immunofluorescence in our pediatric TB granuloma sample. Overall, this study identified a hypoxia-associated SPP1-enriched microenvironment that may contribute to incomplete bacterial clearance and could provide a framework for evaluating host-directed strategies to enhance sterilizing immunity without exacerbating tissue damage.