Human memory CD4+ T cells recognize Mycobacterium tuberculosis-infected macrophages amid broader pathogen-specific responses [scRNA-seq]

CD4+ T cell-mediated control of tuberculosis (TB) requires recognition of macrophages infected with Mycobacterium tuberculosis (Mtb). Yet not all Mtb-specific T cells recognize infected macrophages. Using infected monocyte-derived macrophages (MDMs) and autologous memory CD4+ T cells from individuals with latent Mtb infection (LTBI), we isolate and quantify CD4+ T cells activated in response to infected macrophages. We use T cell antigen receptor (TCR) sequencing to determine the total and unique Mtb-specific TCR clonotypes linked to recognition of infected macrophages, and find that a subset required exogenous antigen exposure, suggesting incomplete recognition. Clonotypes specific for multiple Mtb antigens, and other pathogens, were also identified. We use bulk TCRb deep sequencing from total CD4+ T cells isolated from 10x106 PBMCs from each participant to determine the natural circulating frequencies of relevant TCR clonotypes. We used single-cell RNA sequencing (scRNAseq) to examine the effector functions of CD4+ T cells in response to infected macrophages. Mtb-specific clonotypes expressed signature effector functions dominated by IFNg, TNF, IL-2, and GM-CSF or chemokine production and signaling. We propose TB vaccines that elicit T cells specific for T7SS substrates, recognize infected macrophages, and express canonical effector functions will offer protection against TB.