Single-Cell RNA Sequencing Analysis of Macrophage Polarization Heterogeneity in Active Pulmonary Tuberculosis Patients from Adamawa State

Macrophage polarization states, ranging from classically activated pro-inflammatory (M1) to alternatively activated anti-inflammatory (M2) phenotypes, critically influence M. tuberculosis control and immunopathology in pulmonary tuberculosis. Single-cell RNA sequencing (scRNA-seq) enables the unbiased characterization of heterogeneous macrophage transcriptional states in clinical samples, providing insights unavailable from bulk transcriptomic approaches. This study applied scRNA-seq to alveolar macrophages isolated from bronchoalveolar lavage (BAL) specimens of active pulmonary TB patients at Federal Medical Centre Yola, Adamawa State, Northeast Nigeria, compared with healthy controls. BAL samples from 12 active TB patients and 8 healthy donors were processed for single-cell isolation using the 10x Genomics Chromium controller. Libraries were sequenced on Illumina NovaSeq. Data analysis utilized the Seurat R package (v4.3) with cell type annotation against the Human Cell Atlas macrophage reference. Eleven transcriptionally distinct macrophage subclusters were identified across TB and control samples. TB patients showed significant expansion of an SPP1-high pro-inflammatory macrophage subcluster with strong upregulation of CXCL10, IL-1B, and MMP9, alongside a distinct FOLR2-positive tissue-resident subcluster that was depleted in active disease. Trajectory analysis revealed differentiation paths from circulating monocytes to dysfunctional granuloma-associated macrophage states. These single-cell resolved data provide novel insights into innate immune dysregulation in TB patients in Northeast Nigeria. Keywords: single-cell RNA sequencing, macrophage polarization, tuberculosis, bronchoalveolar lavage, Adamawa State.