SUMMARYTuberculosis (TB) is the leading infectious cause of mortality worldwide, due in part to a limited understanding of its clinical pathogenic spectrum of infection and disease. Historically, scientific research, diagnostic testing, and drug treatment have focused on addressing one of two disease states: latent TB infection or active TB disease. Recent research has clearly demonstrated that human TB infection, from latent infection to active disease, exists within a continuous spectrum of metabolic bacterial activity and antagonistic immunological responses. This revised understanding leads us to propose two additional clinical states: incipient and subclinical TB. The recognition of incipient and subclinical TB, which helps divide latent and active TB along the clinical disease spectrum, provides opportunities for the development of diagnostic and therapeutic interventions to prevent progression to active TB disease and transmission of TB bacilli. In this report, we review the current understanding of the pathogenesis, immunology, clinical epidemiology, diagnosis, treatment, and prevention of both incipient and subclinical TB, two emerging clinical states of an ancient bacterium.

MR1-restricted T cells (MR1Ts) are a T cell subset that recognize and mediate host defense to a broad array of microbial pathogens, including respiratory pathogens (e.g., ,, ) and enteric pathogens (e.g., and species). Mucosal-associated invariant T (MAIT) cells, a subset of MR1Ts, were historically defined by the use of a semi-invariant T cell receptor (TCR) and recognition of small molecules derived from the riboflavin biosynthesis pathway presented on MR1. We used mass spectrometry to identify the repertoire of ligands presented by MR1 from the microbes and We found that the MR1 ligandome is unexpectedly broad, revealing functionally distinct ligands derived from and The identification, synthesis, and functional analysis of mycobacterial ligands reveal that MR1T ligands can be distinguished by MR1Ts with diverse TCR usage. These data demonstrate that MR1 can serve as an immune sensor of the microbial ligandome.

Mucosal-associated invariant T (MAIT) cells are thought to detect microbial antigens presented by the HLA-Ib molecule MR1 through the exclusive use of a TRAV1-2-containing TCRα. Here we use MR1 tetramer staining and ex vivo analysis with mycobacteria-infected MR1-deficient cells to demonstrate the presence of functional human MR1-restricted T cells that lack TRAV1-2. We characterize an MR1-restricted clone that expresses the TRAV12-2 TCRα, which lacks residues previously shown to be critical for MR1-antigen recognition. In contrast to TRAV1-2+ MAIT cells, this TRAV12-2-expressing clone displays a distinct pattern of microbial recognition by detecting infection with the riboflavin auxotroph Streptococcus pyogenes. As known MAIT antigens are derived from riboflavin metabolites, this suggests that TRAV12-2+ clone recognizes unique antigens. Thus, MR1-restricted T cells can discriminate between microbes in a TCR-dependent manner. We postulate that additional MR1-restricted T-cell subsets may play a unique role in defence against infection by broadening the recognition of microbial metabolites.

Tuberculosis (TB) is the leading cause of mortality from a single infectious agent: Mycobacterium tuberculosis (M.tb). Relevant immune targets of the partially efficacious TB vaccine Bacille Calmette-Guérin (BCG) remain poorly defined. Mucosal associated invariant T (MAIT) cells are MHC-related 1 (MR1)-restricted T cells, which are reactive against M.tb, and underexplored as potential TB vaccine targets. We sought to determine whether BCG vaccination activated mycobacteria-specific MAIT cell responses in humans. We analyzed whole blood samples from M.tb-infected South African adults who were re-vaccinated with BCG after a 6-month course of isoniazid preventative therapy. In vitro BCG stimulation potently induced IFNγ expression by phenotypic (CD8+CD26+CD161+) MAIT cells, which constituted the majority (75%) of BCG-reactive IFNγ-producing CD8+ T cells. BCG revaccination transiently expanded peripheral blood frequencies of BCG-reactive IFNγ+ MAIT cells, which returned to baseline frequencies a year following vaccination. In another cohort of healthy adults who received BCG at birth, 53% of mycobacteria-reactive activated CD8 T cells expressed CDR3α TCRs previously reported as MAIT TCRs expressing the canonical TRAV1-2-TRAJ33 MAIT TCRα rearrangement. CD26 and CD161 co-expression correlated with TRAV1-2+CD161+ phenotype more accurately in CD8+ than CD4−CD8− MAIT cells. Interestingly, BCG-induced IFNγ expression by MAIT cells in vitro was mediated by innate cytokines IL-12 and IL-18 more than MR1-induced TCR signaling, suggesting TCR-independent activation. Collectively, the data suggest that activation of blood MAIT cells by innate inflammatory cytokines is a major mechanism of responsiveness to vaccination with whole cell vaccines against TB or in vitro stimulation with mycobacteria. Clinical trial registration:

Mucosal-associated invariant T (MAIT) cells typically express a TRAV1-2 + semi-invariant TCRα that enables recognition of bacterial, mycobacterial, and fungal riboflavin metabolites presented by MR1. MAIT cells are associated with immune control of bacterial and mycobacterial infections in murine models. Here, we report that a population of pro-inflammatory TRAV1-2 + CD8 + T cells are present in the airways and lungs of healthy individuals and are enriched in bronchoalveolar fluid of patients with active pulmonary tuberculosis (TB). High-throughput T cell receptor analysis reveals oligoclonal expansions of canonical and donor-unique TRAV1-2 + MAIT-consistent TCRα sequences within this population. Some of these cells demonstrate MR1-restricted mycobacterial reactivity and phenotypes suggestive of MAIT cell identity. These findings demonstrate enrichment of TRAV1-2 + CD8 + T cells with MAIT or MAIT-like features in the airways during active TB and suggest a role for these cells in the human pulmonary immune response to Mycobacterium tuberculosis .