Immune Responses in Lung Granulomas during Mtb/HIV Co-Infection: Implications for Pathogenesis and Therapy.

HIV and TB are the cause of significant worldwide mortality and pose a grave danger to the global public health. TB is the leading cause of death in HIV-infected persons, with one in four deaths attributable to TB. While the majority of healthy individuals infected with M. tuberculosis (Mtb) are able to control the infection, co-infection with HIV increases the risk of TB infection progressing to TB disease by over 20-fold. While antiretroviral therapy (ART), the cornerstone of HIV care, decreases the incidence of TB in HIV-uninfected people, this remains 4- to 7-fold higher after ART in HIV-co-infected individuals in TB-endemic settings, regardless of the duration of therapy. Thus, the immune control of Mtb infection in Mtb/HIV-co-infected individuals is not fully restored by ART. We do not fully understand the reasons why Mtb/HIV-co-infected individuals maintain a high susceptibility to the reactivation of LTBI, despite an effective viral control by ART. A deep understanding of the molecular mechanisms that govern HIV-induced reactivation of TB is essential to develop improved treatments and vaccines for the Mtb/HIV-co-infected population. We discuss potential strategies for the mitigation of the observed chronic immune activation in combination with both anti-TB and anti-retroviral approaches.

Antigen-specific B cells direct T follicular-like helper cells into lymphoid follicles to mediate Mycobacterium tuberculosis control.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is a global cause of death. Granuloma-associated lymphoid tissue (GrALT) correlates with protection during TB, but the mechanisms of protection are not understood. During TB, the transcription factor IRF4 in T cells but not B cells is required for the generation of the TH1 and TH17 subsets of helper T cells and follicular helper T (TFH)-like cellular responses. A population of IRF4+ T cells coexpress the transcription factor BCL6 during Mtb infection, and deletion of Bcl6 (Bcl6fl/fl) in CD4+ T cells (CD4cre) resulted in reduction of TFH-like cells, impaired localization within GrALT and increased Mtb burden. In contrast, the absence of germinal center B cells, MHC class II expression on B cells, antibody-producing plasma cells or interleukin-10-expressing B cells, did not increase Mtb susceptibility. Indeed, antigen-specific B cells enhance cytokine production and strategically localize TFH-like cells within GrALT via interactions between programmed cell death 1 (PD-1) and its ligand PD-L1 and mediate Mtb control in both mice and macaques.

Inhibition of indoleamine dioxygenase leads to better control of tuberculosis adjunctive to chemotherapy.

The expression of indoleamine 2,3-dioxygenase (IDO), a robust immunosuppressant, is significantly induced in macaque tuberculosis (TB) granulomas, where it is expressed on IFN-responsive macrophages and myeloid-derived suppressor cells. IDO expression is also highly induced in human TB granulomas, and products of its activity are detected in patients with TB. In vivo blockade of IDO activity resulted in the reorganization of the granuloma with substantially greater T cells being recruited to the core of the lesions. This correlated with better immune control of TB and reduced lung M. tuberculosis burdens. To study if the IDO blockade strategy can be translated to a bona fide host-directed therapy in the clinical setting of TB, we studied the effect of IDO inhibitor 1-methyl-d-tryptophan adjunctive to suboptimal anti-TB chemotherapy. While two-thirds of controls and one-third of chemotherapy-treated animals progressed to active TB, inhibition of IDO adjunctive to the same therapy protected macaques from TB, as measured by clinical, radiological, and microbiological attributes. Although chemotherapy improved proliferative T cell responses, adjunctive inhibition of IDO further enhanced the recruitment of effector T cells to the lung. These results strongly suggest the possibility that IDO inhibition can be attempted adjunctive to anti-TB chemotherapy in clinical trials.

Early treatment with anti-α4ß7 antibody facilitates increased gut macrophage maturity in SIV-infected rhesus macaques.

Despite advances in combination antiretroviral therapy (cART), people living with HIV (PLWH) continue to experience gastrointestinal dysfunction. Infusions of anti-α4ß7 monoclonal antibodies (mAbs) have been proposed to increase virologic control during simian immunodeficiency virus (SIV) infection in macaques with mixed results. Recent evidences suggested that therapeutic efficacy of vedolizumab (a humanized anti-α4ß7 mAb), during inflammatory bowel diseases depends on microbiome composition, myeloid cell differentiation, and macrophage phenotype. We tested this hypothesis in SIV-infected, anti-α4ß7 mAb-treated macaques and provide flow cytometric and microscopic evidence that anti-α4ß7 administered to SIV-infected macaques increases the maturity of macrophage phenotypes typically lost in the small intestines during SIV disease progression. Further, this increase in mature macrophage phenotype was associated with tissue viral loads. These phenotypes were also associated with dysbiosis markers in the gut previously identified as predictors of HIV replication and immune activation in PLWH. These findings provide a novel model of anti-α4ß7 efficacy offering new avenues for targeting pathogenic mucosal immune response during HIV/SIV infection.

System-wide identification of myeloid markers of TB disease and HIV-induced reactivation in the macaque model of Mtb infection and Mtb/SIV co-infection.

Mycobacterium tuberculosis (Mtb) has developed specialized mechanisms to parasitize its host cell, the macrophage. These mechanisms allow it to overcome killing by oxidative burst and persist in the wake of an inflammatory response. Mtb infection in the majority of those exposed is controlled in an asymptomatic form referred to as latent tuberculosis infection (LTBI). HIV is a well-known catalyst of reactivation of LTBI to active TB infection (ATB). Through the use of nonhuman primates (NHPs) co-infected with Mtb and Simian Immunodeficiency Virus (Mtb/SIV), we are able to simulate human progression of TB/AIDS comorbidity. The advantage of NHP models is that they recapitulate the breadth of human TB outcomes, including immune control of infection, and loss of this control due to SIV co-infection. Identifying correlates of immune control of infection is important for both vaccine and therapeutics development. Using macaques infected with Mtb or Mtb/SIV and with different clinical outcomes we attempted to identify signatures between those that progress to active infection after SIV challenge (reactivators) and those that control the infection (non-reactivators). We particularly focused on pathways relevant to myeloid origin cells such as macrophages, as these innate immunocytes have an important contribution to the initial control or the lack thereof, following Mtb infection. Using bacterial burden, C-reactive protein (CRP), and other clinical indicators of disease severity as a guide, we were able to establish gene signatures of host disease state and progression. In addition to gene signatures, clustering algorithms were used to differentiate between host disease states and identify relationships between genes. This allowed us to identify clusters of genes which exhibited differential expression profiles between the three groups of macaques: ATB, LTBI and Mtb/SIV. The gene signatures were associated with pathways relevant to apoptosis, ATP production, phagocytosis, cell migration, and Type I interferon (IFN), which are related to macrophage function. Our results suggest novel macrophage functions that may play roles in the control of Mtb infection with and without co-infection with SIV. These results particularly point towards an interplay between Type I IFN signaling and IFN-γ signaling, and the resulting impact on lung macrophages as an important determinant of progression to TB.

Isoniazid and rifapentine treatment effectively reduces persistent M. tuberculosis infection in macaque lungs.

A once-weekly oral dose of isoniazid and rifapentine for 3 months (3HP) is recommended by the CDC for treatment of latent tuberculosis infection (LTBI). The aim of this study is to assess 3HP-mediated clearance of M. tuberculosis bacteria in macaques with asymptomatic LTBI. Twelve Indian-origin rhesus macaques were infected with a low dose (~10 CFU) of M. tuberculosis CDC1551 via aerosol. Six animals were treated with 3HP and 6 were left untreated. The animals were imaged via PET/CT at frequent intervals. Upon treatment completion, all animals except 1 were coinfected with SIV to assess reactivation of LTBI to active tuberculosis (ATB). Four of 6 treated macaques showed no evidence of persistent bacilli or extrapulmonary spread until the study end point. PET/CT demonstrated the presence of significantly more granulomas in untreated animals relative to the treated group. The untreated animals harbored persistent bacilli and demonstrated tuberculosis (TB) reactivation following SIV coinfection, while none of the treated animals reactivated to ATB. 3HP treatment effectively reduced persistent infection with M. tuberculosis and prevented reactivation of TB in latently infected macaques.

Peripheral Blood Markers Correlate with the Progression of Active Tuberculosis Relative to Latent Control of Mycobacterium tuberculosis Infection in Macaques.

Despite a century of research into tuberculosis (TB), there is a dearth of reproducible, easily quantifiable, biomarkers that can predict disease onset and differentiate between host disease states. Due to the challenges associated with human sampling, nonhuman primates (NHPs) are utilized for recapitulating the closest possible modelling of human TB. To establish a predictive peripheral biomarker profile based on a larger cohort of rhesus macaques (RM), we analyzed results pertaining to peripheral blood serum chemistry and cell counts from RMs that were experimentally exposed to Mtb in our prior studies and characterized as having either developed active TB (ATB) disease or latent TB infection (LTBI). We compared lung CFU burdens and quantitative pathologies with a number of measurables in the peripheral blood. Based on our results, the investigations were then extended to the study of specific molecules and cells in the lung compartments of a subset of these animals and their immune responses. In addition to the elevated serum C-reactive protein (CRP) levels, frequently used to discern the level of Mtb infection in model systems, reduced serum albumin-to-globulin (A/G) ratios were also predictive of active TB disease. Furthermore, higher peripheral myeloid cell levels, particularly those of neutrophils, kynurenine-to-tryptophan ratio, an indicator of induced expression of the immunosuppressive molecule indoleamine dioxygenase, and an influx of myeloid cell populations could also efficiently discriminate between ATB and LTBI in experimentally infected macaques. These quantifiable correlates of disease were then used in conjunction with a regression-based analysis to predict bacterial load. Our results suggest a potential biomarker profile of TB disease in rhesus macaques, that could inform future NHP-TB research. Our results thus suggest that specific biomarkers may be developed from the myeloid subset of peripheral blood or plasma with the ability to discriminate between active and latent Mtb infection.

Response to Hypoxia and the Ensuing Dysregulation of Inflammation Impacts Mycobacterium tuberculosis Pathogenicity.

Rationale: Different Mycobacterium tuberculosis (Mtb) strains exhibit variable degrees of virulence in humans and animal models. Differing stress response strategies used by different strains of Mtb could influence virulence. Objectives: We compared the virulence of two strains of Mtb with use in animal model research: CDC1551 and Erdman. Methods: Rhesus macaques, which develop human-like tuberculosis attributes and pathology, were infected with a high dose of either strain via aerosol, and virulence was compared by bacterial burden and pathology. Measurements and Main Results: Infection with Erdman resulted in significantly shorter times to euthanasia and higher bacterial burdens and greater systemic inflammation and lung pathology relative to those infected with CDC1551. Macaques infected with Erdman also exhibited significantly higher early inflammatory myeloid cell influx to the lung, greater macrophage and T cell activity, and higher expression of lung remodeling (extracellular matrix) genes, consistent with greater pathology. Expression of NOTCH4 (neurogenic locus notch homolog 4) signaling, which is induced in response to hypoxia and promotes undifferentiated cellular state, was also higher in Erdman-infected lungs. The granulomas generated by Erdman, and not CDC1551, infection appeared to have larger regions of necrosis, which is strongly associated with hypoxia. To better understand the mechanisms of differential hypoxia induction by these strains, we subjected both to hypoxia in vitro. Erdman induced higher concentrations of DosR regulon relative to CDC1551. The DosR regulon is the global regulator of response to hypoxia in Mtb and critical for its persistence in granulomas. Conclusions: Our results show that the response to hypoxia is a critical mediator of virulence determination in Mtb, with potential impacts on bacillary persistence, reactivation, and efficiency of therapeutics.

Myeloid cell interferon responses correlate with clearance of SARS-CoV-2.

Emergence of mutant SARS-CoV-2 strains associated with an increased risk of COVID-19-related death necessitates better understanding of the early viral dynamics, host responses and immunopathology. Single cell RNAseq (scRNAseq) allows for the study of individual cells, uncovering heterogeneous and variable responses to environment, infection and inflammation. While studies have reported immune profiling using scRNAseq in terminal human COVID-19 patients, performing longitudinal immune cell dynamics in humans is challenging. Macaques are a suitable model of SARS-CoV-2 infection. Our longitudinal scRNAseq of bronchoalveolar lavage (BAL) cell suspensions from young rhesus macaques infected with SARS-CoV-2 (n = 6) demonstrates dynamic changes in transcriptional landscape 3 days post- SARS-CoV-2-infection (3dpi; peak viremia), relative to 14-17dpi (recovery phase) and pre-infection (baseline) showing accumulation of distinct populations of both macrophages and T-lymphocytes expressing strong interferon-driven inflammatory gene signature at 3dpi. Type I interferon response is induced in the plasmacytoid dendritic cells with appearance of a distinct HLADR+CD68+CD163+SIGLEC1+ macrophage population exhibiting higher angiotensin-converting enzyme 2 (ACE2) expression. These macrophages are significantly enriched in the lungs of macaques at 3dpi and harbor SARS-CoV-2 while expressing a strong interferon-driven innate anti-viral gene signature. The accumulation of these responses correlated with decline in viremia and recovery.

The immunoregulatory landscape of human tuberculosis granulomas.

Tuberculosis (TB) in humans is characterized by formation of immune-rich granulomas in infected tissues, the architecture and composition of which are thought to affect disease outcome. However, our understanding of the spatial relationships that control human granulomas is limited. Here, we used multiplexed ion beam imaging by time of flight (MIBI-TOF) to image 37 proteins in tissues from patients with active TB. We constructed a comprehensive atlas that maps 19 cell subsets across 8 spatial microenvironments. This atlas shows an IFN-γ-depleted microenvironment enriched for TGF-ß, regulatory T cells and IDO1+ PD-L1+ myeloid cells. In a further transcriptomic meta-analysis of peripheral blood from patients with TB, immunoregulatory trends mirror those identified by granuloma imaging. Notably, PD-L1 expression is associated with progression to active TB and treatment response. These data indicate that in TB granulomas, there are local spatially coordinated immunoregulatory programs with systemic manifestations that define active TB.

Impact of SIV infection on mycobacterial lipid-reactive T cell responses in Bacillus Calmette-Guérin (BCG) inoculated macaques.

Background: Although BCG vaccine protects infants from tuberculosis (TB), it has limited efficacy in adults against pulmonary TB. Further, HIV coinfection significantly increases the risk of developing active TB. In the lack of defined correlates of protection in TB disease, it is essential to explore immune responses beyond conventional CD4 T cells to gain a better understanding of the mechanisms of TB immunity. Methods: Here, we evaluated unconventional lipid-reactive T cell responses in cynomolgus macaques following aerosol BCG inoculation and examined the impact of subsequent SIV infection on these responses. Immune responses to cellular lipids of M. bovis and M. tuberculosis were examined ex vivo in peripheral blood and bronchioalveolar lavage (BAL). Results: Prior to BCG inoculation, innate-like IFN-γ responses to mycobacterial lipids were observed in T cells. Aerosol BCG exposure induced an early increase in frequencies of BAL γδT cells, a dominant subset of lipid-reactive T cells, along with enhanced IL-7R and CXCR3 expression. Further, BCG exposure stimulated greater IFN-γ responses to mycobacterial lipids in peripheral blood and BAL, suggesting the induction of systemic and local Th1-type response in lipid-reactive T cells. Subsequent SIV infection resulted in a significant loss of IL-7R expression on blood and BAL γδT cells. Additionally, IFN-γ responses of mycobacterial lipid-reactive T cells in BAL fluid were significantly lower in SIV-infected macaques, while perforin production was maintained through chronic SIV infection. Conclusions: Overall, these data suggest that despite SIV-induced decline in IL-7R expression and IFN-γ production by mycobacterial lipid-reactive T cells, their cytolytic potential is maintained. A deeper understanding of anti-mycobacterial lipid-reactive T cell functions may inform novel approaches to enhance TB control in individuals with or without HIV infection.

Antiretroviral therapy timing impacts latent tuberculosis infection reactivation in a Mycobacterium tuberculosis/SIV coinfection model.

Studies using the nonhuman primate model of Mycobacterium tuberculosis/simian immunodeficiency virus coinfection have revealed protective CD4+ T cell-independent immune responses that suppress latent tuberculosis infection (LTBI) reactivation. In particular, chronic immune activation rather than the mere depletion of CD4+ T cells correlates with reactivation due to SIV coinfection. Here, we administered combinatorial antiretroviral therapy (cART) 2 weeks after SIV coinfection to study whether restoration of CD4+ T cell immunity occurred more broadly, and whether this prevented reactivation of LTBI compared to cART initiated 4 weeks after SIV. Earlier initiation of cART enhanced survival, led to better control of viral replication, and reduced immune activation in the periphery and lung vasculature, thereby reducing the rate of SIV-induced reactivation. We observed robust CD8+ T effector memory responses and significantly reduced macrophage turnover in the lung tissue. However, skewed CD4+ T effector memory responses persisted and new TB lesions formed after SIV coinfection. Thus, reactivation of LTBI is governed by very early events of SIV infection. Timing of cART is critical in mitigating chronic immune activation. The potential novelty of these findings mainly relates to the development of a robust animal model of human M. tuberculosis/HIV coinfection that allows the testing of underlying mechanisms.

Myeloid-Derived Suppressor Cells Mediate T Cell Dysfunction in Nonhuman Primate TB Granulomas.

Myeloid-derived suppressor cells (MDSCs) represent an innate immune cell population comprised of immature myeloid cells and myeloid progenitors with very potent immunosuppressive potential. MDSCs are reported to be abundant in the lungs of active tuberculosis (TB) patients. We sought to perform an in-depth study of MDSCs during latent TB infection (LTBI) and active TB (ATB) using the nonhuman primate (NHP) model of pulmonary TB. We found a higher proportion of granulocytic, polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) in the lungs of ATB animals compared to those with LTBI or naive control animals. Active disease in the lung, but not LTBI, was furthermore associated with higher proliferation, expansion, and immunosuppressive capabilities of PMN-MDSCs, as shown by enhanced expression of Ki67, indoleamine 2,3-dioxygenase (IDO1), interleukin-10 (IL-10), matrix metallopeptidase 9 (MMP-9), inducible nitric oxide synthase (iNOS), and programmed death-ligand 1 (PD-L1). These immunosuppressive PMN-MDSCs specifically localized to the lymphocytic cuff at the periphery of the granulomas in animals with ATB. Conversely, these cells were scarcely distributed in interstitial lung tissue and the inner core of granulomas. This spatial regulation suggests an important immunomodulatory role of PMN-MDSCs by restricting T cell access to the TB granuloma core and can potentially explain dysfunctional anti-TB responses in active granuloma. Our results raise the possibility that the presence of MDSCs can serve as a biomarker for ATB, while their disappearance can indicate successful therapy. Furthermore, MDSCs may serve as a potential target cell for adjunctive TB therapy. IMPORTANCE Myeloid cells are immunocytes of innate origin that orchestrate the first response toward pathogens via immune surveillance (uptake and killing), antigen presentation, and initiation of adaptive immunity by T cell stimulation. However, MDSCs are a subset of innate immunocytes that deviate to an immunoregulatory phenotype. MDSCs possess strong immunosuppressive capabilities that are induced in autoimmune, malignant neoplastic, and chronic inflammatory diseases. Induction of MDSCs has been found in peripheral blood, bronchoalveolar lavage (BAL) fluid, and pleural effusions of active TB patients, but their precise localization in lung tissue and in TB granulomas remains unclear due to challenges associated with sampling lungs and granulomas from active TB patients. Nonhuman primates (NHPs) are an important animal model with TB granulomas that closely mimic those found in humans and can therefore be used for studies that are otherwise challenging with patient material. Herein, we study MDSC localization in the lungs of NHPs exhibiting latent and active TB. Our findings reveal that MDSCs localize and exert their immunosuppressive roles at the periphery rather than in the core of TB granulomas.

Robust IgM responses following intravenous vaccination with Bacille Calmette-Guérin associate with prevention of Mycobacterium tuberculosis infection in macaques.

Development of an effective tuberculosis (TB) vaccine has suffered from an incomplete understanding of the correlates of protection against Mycobacterium tuberculosis (Mtb). Intravenous (i.v.) vaccination with Bacille Calmette-Guérin (BCG) provides nearly complete protection against TB in rhesus macaques, but the antibody response it elicits remains incompletely defined. Here we show that i.v. BCG drives superior antibody responses in the plasma and the lungs of rhesus macaques compared to traditional intradermal BCG administration. While i.v. BCG broadly expands antibody titers and functions, IgM titers in the plasma and lungs of immunized macaques are among the strongest markers of reduced bacterial burden. IgM was also enriched in macaques that received protective vaccination with an attenuated strain of Mtb. Finally, an Mtb-specific IgM monoclonal antibody reduced Mtb survival in vitro. Collectively, these data highlight the potential importance of IgM responses as a marker and mediator of protection against TB.

Transcriptional Response of Mycobacterium tuberculosis to Cigarette Smoke Condensate.

Smoking is known to be an added risk factor for tuberculosis (TB), with nearly a quarter of the TB cases attributed to cigarette smokers in the 22 countries with the highest TB burden. Many studies have indicated a link between risk of active TB and cigarette smoke. Smoking is also known to significantly decrease TB cure and treatment completion rate and increase mortality rates. Cigarette smoke contains thousands of volatile compounds including carcinogens, toxins, reactive solids, and oxidants in both particulate and gaseous phase. Yet, to date, limited studies have analyzed the impact of cigarette smoke components on Mycobacterium tuberculosis (Mtb), the causative agent of TB. Here we report the impact of cigarette smoke condensate (CSC) on survival, mutation frequency, and gene expression of Mtb in vitro. We show that exposure of virulent Mtb to cigarette smoke increases the mutation frequency of the pathogen and strongly induces the expression of the regulon controlled by SigH-a global transcriptional regulator of oxidative stress. SigH has previously been shown to be required for Mtb to respond to oxidative stress, survival, and granuloma formation in vivo. A high-SigH expression phenotype is known to be associated with greater virulence of Mtb. In patients with pulmonary TB who smoke, these changes may therefore play an important, yet unexplored, role in the treatment efficacy by potentially enhancing the virulence of tubercle bacilli.

Myeloid cell interferon responses correlate with clearance of SARS-CoV-2.

The emergence of mutant SARS-CoV-2 strains associated with an increased risk of COVID-19-related death necessitates better understanding of the early viral dynamics, host responses and immunopathology. While studies have reported immune profiling using single cell RNA sequencing in terminal human COVID-19 patients, performing longitudinal immune cell dynamics in humans is challenging. Macaques are a suitable model of SARS-CoV-2 infection. We performed longitudinal single-cell RNA sequencing of bronchoalveolar lavage (BAL) cell suspensions from adult rhesus macaques infected with SARS-CoV-2 (n=6) to delineate the early dynamics of immune cells changes. The bronchoalveolar compartment exhibited dynamic changes in transcriptional landscape 3 days post- SARS-CoV-2-infection (3dpi) (peak viremia), relative to 14-17dpi (recovery phase) and pre-infection (baseline). We observed the accumulation of distinct populations of both macrophages and T-lymphocytes expressing strong interferon-driven inflammatory gene signature at 3dpi. Type I IFN response was highly induced in the plasmacytoid dendritic cells. The presence of a distinct HLADR+CD68+CD163+SIGLEC1+ macrophage population exhibiting higher angiotensin converting enzyme 2 (ACE2) expression was also observed. These macrophages were significantly recruited to the lungs of macaques at 3dpi and harbored SARS-CoV-2, while expressing a strong interferon-driven innate anti-viral gene signature. The accumulation of these responses correlated with decline in viremia and recovery. The recruitment of a myeloid cell-mediated Type I IFN response is associated with the rapid clearance of SARS-CoV-2 infection in macaques.

Abnormal Tryptophan Metabolism in HIV and Mycobacterium tuberculosis Infection.

Host metabolism has recently gained more attention for its roles in physiological functions and pathologic conditions. Of these, metabolic tryptophan disorders generate a pattern of abnormal metabolites that are implicated in various diseases. Here, we briefly highlight the recent advances regarding abnormal tryptophan metabolism in HIV and Mycobacterium tuberculosis infection and discuss its potential impact on immune regulation, disease progression, and neurological disorders. Finally, we also discuss the potential for metabolic tryptophan interventions toward these infectious diseases.

A non-canonical type 2 immune response coordinates tuberculous granuloma formation and epithelialization.

The central pathogen-immune interface in tuberculosis is the granuloma, a complex host immune structure that dictates infection trajectory and physiology. Granuloma macrophages undergo a dramatic transition in which entire epithelial modules are induced and define granuloma architecture. In tuberculosis, relatively little is known about the host signals that trigger this transition. Using the zebrafish-Mycobacterium marinum model, we identify the basis of granuloma macrophage transformation. Single-cell RNA-sequencing analysis of zebrafish granulomas and analysis of Mycobacterium tuberculosis-infected macaques reveal that, even in the presence of robust type 1 immune responses, countervailing type 2 signals associate with macrophage epithelialization. We find that type 2 immune signaling, mediated via stat6, is absolutely required for epithelialization and granuloma formation. In mixed chimeras, stat6 acts cell autonomously within macrophages, where it is required for epithelioid transformation and incorporation into necrotic granulomas. These findings establish the signaling pathway that produces the hallmark structure of mycobacterial infection.

The immune landscape in tuberculosis reveals populations linked to disease and latency.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb) latently infects approximately one-fourth of the world's population. The immune mechanisms that govern progression from latent (LTBI) to active pulmonary TB (PTB) remain poorly defined. Experimentally Mtb-infected non-human primates (NHP) mirror the disease observed in humans and recapitulate both PTB and LTBI. We characterized the lung immune landscape in NHPs with LTBI and PTB using high-throughput technologies. Three defining features of PTB in macaque lungs include the influx of plasmacytoid dendritic cells (pDCs), an Interferon (IFN)-responsive macrophage population, and activated T cell responses. In contrast, a CD27+ Natural killer (NK) cell subset accumulated in the lungs of LTBI macaques. This NK cell population was also detected in the circulation of LTBI individuals. This comprehensive analysis of the lung immune landscape will improve the understanding of TB immunopathogenesis, providing potential targets for therapies and vaccines for TB control.