Single-cell RNA sequencing reveals immune cell dysfunction and rewired interaction networks in the peripheral blood of active tuberculosis patients

Abstract Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), remains a global health threat with poorly understood immune mechanisms. This exploratory study employed single-cell RNA sequencing (scRNA-seq) on peripheral blood mononuclear cells (PBMCs) from 3 active TB (ATB) patients and 2 healthy controls (HC) to dissect immune cell dysfunction and communication networks. Data were processed using CellRanger, Seurat, and Harmony for quality control, batch correction, and clustering, with cell types annotated via marker genes and SingleR. Functional analyses included cytotoxicity, antigen presentation scoring, differential gene expression, and Gene Ontology (GO) enrichment, while CellChat mapped ligand-receptor interactions. Results revealed altered immune cell composition in ATB, including increased myeloid cells/platelets and reduced NK cells. CD8 + effector T cells (CD8Te) and CD16bright CD56dim NK cells (NKCD56dim) exhibited reduced expression of cytotoxicity-associated genes, while Naïve B cells (NaïveB) and dendritic cells (DC) showed impaired antigen presentation. RNA metabolism pathways were upregulated alongside suppressed immune regulation. Cell communication analysis identified weakened CD8 + T cell incoming signals, enhanced B/NK cell signaling, and intensified monocyte-driven interactions. TGF-β pathway enrichment and defective MHC signaling disrupted network dynamics, suggesting RNA metabolism dysregulation and TGF-β-mediated immunosuppression as key mechanisms. These findings provide insights that may help inform future studies on TB immunopathology and treatment development. Limitations include reliance on peripheral blood, small sample size, and lack of protein-level validation. Future studies should integrate multi-omics and functional assays to validate these pathways.