Although CD4 T cells are essential for immunity to tuberculosis (TB), the features of CD4 T cells that provide protective immunity are not well understood. This knowledge gap is due in part to limited knowledge of the M. tuberculosis (Mtb) antigens that induce protective immune responses. In other pathogens, immune recognition of some antigens provides protection, while recognition of other antigens does not. Likewise in other pathogens, the antigenic targets of protective immunity undergo diversifying evolutionary selection to generate antigenic variation and escape immune recognition. We hypothesized that these principles apply to Mtb, and in earlier studies we made the unexpected discovery that the commonly-studied antigenic targets of human T cells (epitopes) in Mtb are hyperconserved; they are the most conserved elements of the Mtb genome. We then sought to find exceptions, and combined comparative genomics of phylogenetically diverse strains of Mtb with experimental immunology. This led to our discovery of seven novel Mtb antigens (that we term Mtb Rare Variable Antigens; RVA) that are recognized by human T cells and exhibit evidence of evolutionary diversifying selection. Together, these results suggest that human T cell recognition of RVA is detrimental to Mtb and that recognition of conserved 'classical' Mtb antigens is not detrimental to Mtb. We recently studied CD4 T cells from healthy Quantiferon-TB positive (QFT+) recent close contacts of infectious cases of TB, and discovered that RVA induce CD4 T cells characterized by dominant interleukin 17 responses and expression of the lineage-defining transcription factor RORγt, in contrast to CD4 T cells that recognize classical Mtb antigens and exhibit interferon gamma responses and expression of T-bet. These results indicate that human CD4 T cells that recognize RVA are functionally distinct from those that recognize conserved antigens, and we hypothesize that T cells that recognize RVA provide protection against active TB, as indicated by their evolutionary selection. In this project, we will use innovative assays to intensively characterize CD4 T cells from QFT+ adults by comparing the features of CD4 T cells that recognize RVA versus classical Mtb antigens. We will compare their functional responses, longitudinal memory phenotypes, and extent of differentiation. We will also employ an innovative new platform developed by our team, using DNA barcoded peptide epitopes and next generation sequencing to test the hypothesis that associations between CD4 T cell antigen specificity and certain functional responses are widespread in Mtb. To determine the significance of RVA in protective immunity to TB, we will compare CD4 T cells that recognize RVA vs classical Mtb antigens in adults with active TB and those with 'controlled' (or latent) TB. We will also test the hypothesis that defects in RVA-specific CD4 T cells are primary and do not reverse with treatment of active TB. The long term object...