Mycobacterium tuberculosis resides in lysosome-poor monocyte-derived lung cells during chronic infection.

Mycobacterium tuberculosis (Mtb) infects lung myeloid cells, but the specific Mtb-permissive cells and host mechanisms supporting Mtb persistence during chronic infection are incompletely characterized. We report that after the development of T cell responses, CD11clo monocyte-derived cells harbor more live Mtb than alveolar macrophages (AM), neutrophils, and CD11chi monocyte-derived cells. Transcriptomic and functional studies revealed that the lysosome pathway is underexpressed in this highly permissive subset, characterized by less lysosome content, acidification, and proteolytic activity than AM, along with less nuclear TFEB, a regulator of lysosome biogenesis. Mtb infection does not drive lysosome deficiency in CD11clo monocyte-derived cells but promotes recruitment of monocytes that develop into permissive lung cells, mediated by the Mtb ESX-1 secretion system. The c-Abl tyrosine kinase inhibitor nilotinib activates TFEB and enhances lysosome functions of macrophages in vitro and in vivo, improving control of Mtb infection. Our results suggest that Mtb exploits lysosome-poor lung cells for persistence and targeting lysosome biogenesis is a potential host-directed therapy for tuberculosis.

The Structural and Molecular Mechanisms of Mycobacterium tuberculosis Translational Elongation Factor Proteins.

Targeting translation factor proteins holds promise for developing innovative anti-tuberculosis drugs. During protein translation, many factors cause ribosomes to stall at messenger RNA (mRNA). To maintain protein homeostasis, bacteria have evolved various ribosome rescue mechanisms, including the predominant trans-translation process, to release stalled ribosomes and remove aberrant mRNAs. The rescue systems require the participation of translation elongation factor proteins (EFs) and are essential for bacterial physiology and reproduction. However, they disappear during eukaryotic evolution, which makes the essential proteins and translation elongation factors promising antimicrobial drug targets. Here, we review the structural and molecular mechanisms of the translation elongation factors EF-Tu, EF-Ts, and EF-G, which play essential roles in the normal translation and ribosome rescue mechanisms of Mycobacterium tuberculosis (Mtb). We also briefly describe the structure-based, computer-assisted study of anti-tuberculosis drugs.

Dual neutrophil subsets exacerbate or suppress inflammation in tuberculosis via IL-1ß or PD-L1.

Neutrophils can be beneficial or deleterious during tuberculosis (TB). Based on the expression of MHC-II and programmed death ligand 1 (PD-L1), we distinguished two functionally and transcriptionally distinct neutrophil subsets in the lungs of mice infected with mycobacteria. Inflammatory [MHC-II-, PD-L1lo] neutrophils produced inflammasome-dependent IL-1ß in the lungs in response to virulent mycobacteria and "accelerated" deleterious inflammation, which was highly exacerbated in IFN-γR-/- mice. Regulatory [MHC-II+, PD-L1hi] neutrophils "brake" inflammation by suppressing T-cell proliferation and IFN-γ production. Such beneficial regulation, which depends on PD-L1, is controlled by IFN-γR signaling in neutrophils. The hypervirulent HN878 strain from the Beijing genotype curbed PD-L1 expression by regulatory neutrophils, abolishing the braking function and driving deleterious hyperinflammation in the lungs. These findings add a layer of complexity to the roles played by neutrophils in TB and may explain the reactivation of this disease observed in cancer patients treated with anti-PD-L1.

Serine protease Rv2569c facilitates transmission of Mycobacterium tuberculosis via disrupting the epithelial barrier by cleaving E-cadherin.

Epithelial cells function as the primary line of defense against invading pathogens. However, bacterial pathogens possess the ability to compromise this barrier and facilitate the transmigration of bacteria. Nonetheless, the specific molecular mechanism employed by Mycobacterium tuberculosis (M.tb) in this process is not fully understood. Here, we investigated the role of Rv2569c in M.tb translocation by assessing its ability to cleave E-cadherin, a crucial component of cell-cell adhesion junctions that are disrupted during bacterial invasion. By utilizing recombinant Rv2569c expressed in Escherichia coli and subsequently purified through affinity chromatography, we demonstrated that Rv2569c exhibited cell wall-associated serine protease activity. Furthermore, Rv2569c was capable of degrading a range of protein substrates, including casein, fibrinogen, fibronectin, and E-cadherin. We also determined that the optimal conditions for the protease activity of Rv2569c occurred at a temperature of 37°C and a pH of 9.0, in the presence of MgCl2. To investigate the function of Rv2569c in M.tb, a deletion mutant of Rv2569c and its complemented strains were generated and used to infect A549 cells and mice. The results of the A549-cell infection experiments revealed that Rv2569c had the ability to cleave E-cadherin and facilitate the transmigration of M.tb through polarized A549 epithelial cell layers. Furthermore, in vivo infection assays demonstrated that Rv2569c could disrupt E-cadherin, enhance the colonization of M.tb, and induce pathological damage in the lungs of C57BL/6 mice. Collectively, these results strongly suggest that M.tb employs the serine protease Rv2569c to disrupt epithelial defenses and facilitate its systemic dissemination by crossing the epithelial barrier.

Metabolic changes enhance necroptosis of type 2 diabetes mellitus mice infected with Mycobacterium tuberculosis.

Previously, we found that Mycobacterium tuberculosis (Mtb) infection in type 2 diabetes mellitus (T2DM) mice enhances inflammatory cytokine production which drives pathological immune responses and mortality. In the current study, using a T2DM Mtb infection mice model, we determined the mechanisms that make T2DM mice alveolar macrophages (AMs) more inflammatory upon Mtb infection. Among various cell death pathways, necroptosis is a major pathway involved in inflammatory cytokine production by T2DM mice AMs. Anti-TNFR1 antibody treatment of Mtb-infected AMs from T2DM mice significantly reduced expression of receptor interacting protein kinase 3 (RIPK3) and mixed lineage kinase domain-like (MLKL) (necroptosis markers) and IL-6 production. Metabolic profile comparison of Mtb-infected AMs from T2DM mice and Mtb-infected AMs of nondiabetic control mice indicated that 2-ketohexanoic acid and deoxyadenosine monophosphate were significantly abundant, and acetylcholine and pyridoxine (Vitamin B6) were significantly less abundant in T2DM mice AMs infected with Mtb. 2-Ketohexanoic acid enhanced expression of TNFR1, RIPK3, MLKL and inflammatory cytokine production in the lungs of Mtb-infected nondiabetic mice. In contrast, pyridoxine inhibited RIPK3, MLKL and enhanced expression of Caspase 3 (apoptosis marker) in the lungs of Mtb-infected T2DM mice. Our findings demonstrate that metabolic changes in Mtb-infected T2DM mice enhance TNFR1-mediated necroptosis of AMs, which leads to excess inflammation and lung pathology.

Roles of HIF-1α signaling in Mycobacterium tuberculosis infection: New targets for anti-TB therapeutics?

Tuberculosis (TB), a deadly infectious disease induced by Mycobacterium tuberculosis (Mtb), continues to be a global public health issue that kill millions of patents every year. Despite significant efforts have been paid to identify effective TB treatments, the emergence of drug-resistant strains of the disease and the presence of comorbidities in TB patients urges us to explore the detailed mechanisms involved in TB immunity and develop more effective innovative anti-TB strategies. HIF-1α, a protein involved in regulating cellular immune responses during TB infection, has been highlighted as a promising target for the development of novel strategies for TB treatment due to its critical roles in anti-TB host immunity. This review provides a summary of current research progress on the roles of HIF-1α in TB infection, highlighting its importance in regulating the host immune response upon Mtb infection and summarizing the influences and mechanisms of HIF-1α on anti-TB immunological responses of host cells. This review also discusses the various challenges associated with developing HIF-1α as a target for anti-TB therapies, including ensuring specificity and avoiding off-target effects on normal cell function, determining the regulation and expression of HIF-1α in TB patients, and developing drugs that can inhibit HIF-1α. More deep understanding of the molecular mechanisms involved in HIF-1α signaling, its impact on TB host status, and systematic animal testing and clinical trials may benefit the optimization of HIF-1α as a novel therapeutic target for TB.

Synthesis of an Immunologically Active Heptamannoside of Mycobacterium tuberculosis by the [Au]/[Ag]-Catalyzed Activation of Ethynylcyclohexyl Glycosyl Carbonate Donor.

Tuberculosis (TB) is one of the most dreadful diseases, killing more than 3 million humans annually. M. tuberculosis (MTb) is the causative agent for TB and has a thick and waxy cell wall, making it an attractive target for immunological studies. In this study, a heptamannopyranoside containing 1 → 2 and 1 → 6 α-mannopyranosidic linkages has been explored for the immunological evaluations. The conjugation-ready heptamannopyranoside was synthesized by exploiting the salient features of recently discovered [Au]/[Ag]-glycosidation of ethynylcyclohexyl glycosyl carbonate donors. The glycan was conjugated to the ESAT6, an early secreted protein of MTb for further characterization as a potential subunit vaccine candidate.

ERK1/2-CEBPB Axis-Regulated hBD1 Enhances Anti-Tuberculosis Capacity in Alveolar Type II Epithelial Cells.

Tuberculosis, caused by Mycobacterium tuberculosis (Mtb), remains a global health crisis with substantial morbidity and mortality rates. Type II alveolar epithelial cells (AEC-II) play a critical role in the pulmonary immune response against Mtb infection by secreting effector molecules such as antimicrobial peptides (AMPs). Here, human ß-defensin 1 (hBD1), an important AMP produced by AEC-II, has been demonstrated to exert potent anti-tuberculosis activity. HBD1 overexpression effectively inhibited Mtb proliferation in AEC-II, while mice lacking hBD1 exhibited susceptibility to Mtb and increased lung tissue inflammation. Mechanistically, in A549 cells infected with Mtb, STAT1 negatively regulated hBD1 transcription, while CEBPB was the primary transcription factor upregulating hBD1 expression. Furthermore, we revealed that the ERK1/2 signaling pathway activated by Mtb infection led to CEBPB phosphorylation and nuclear translocation, which subsequently promoted hBD1 expression. Our findings suggest that the ERK1/2-CEBPB-hBD1 regulatory axis can be a potential therapeutic target for anti-tuberculosis therapy aimed at enhancing the immune response of AEC-II cells.

Cutting Edge: Cytosolic Receptor AIM2 Is Induced by Peroxisome Proliferator-activated Receptor γ following Mycobacterium tuberculosis Infection of Human Macrophages but Does Not Contribute to IL-1ß Release.

AIM2 (absent in melanoma 2), an inflammasome component, mediates IL-1ß release in murine macrophages and cell lines. AIM2 and IL-1ß contribute to murine control of Mycobacterium tuberculosis (M.tb) infection, but AIM2's impact in human macrophages, the primary niche for M.tb, remains unclear. We show that M.tb, Mycobacterium bovis bacillus Calmette-Guérin (BCG), and M. smegmatis induce AIM2 expression in primary human macrophages. M.tb-induced AIM2 expression is peroxisome proliferator-activated receptor γ (PPARγ)-dependent and M.tb ESX-1-independent, whereas BCG- and M. smegmatis-induced AIM2 expression is PPARγ-independent. PPARγ and NLRP3, but not AIM2, are important for IL-1ß release in response to M.tb, and NLRP3 colocalizes with M.tb. This is in contrast to the role for AIM2 in inflammasome activation in mice and peritoneal macrophages. Altogether, we show that mycobacteria induce AIM2 expression in primary human macrophages, but AIM2 does not contribute to IL-1ß release during M.tb infection, providing further evidence that AIM2 expression and function are regulated in a cell- and/or species-specific manner.

Does PET-CT Have a Role in the Evaluation of Tuberculosis Treatment in Phase 2 Clinical Trials?

Positron emission tomography-computed tomography (PET-CT) has the potential to revolutionize research in infectious diseases, as it has done with cancer. There is growing interest in it as a biomarker in the setting of early-phase tuberculosis clinical trials, particularly given the limitations of current biomarkers as adequate predictors of sterilizing cure for tuberculosis. PET-CT is a real-time tool that provides a 3-dimensional view of the spatial distribution of tuberculosis within the lung parenchyma and the nature of lesions with uptake (ie, whether nodular, consolidative, or cavitary). Its ability to provide functional data on changes in metabolism, drug penetration, and immune control of tuberculous lesions has the potential to facilitate drug development and regimen selection for advancement to phase 3 trials in tuberculosis. In this narrative review, we discuss the role that PET-CT may have in evaluating responses to drug therapy in active tuberculosis treatment and the challenges in taking PET-CT forward as predictive biomarker of relapse-free cure in the setting of phase 2 clinical trials.

Cargoes of exosomes function as potential biomarkers for Mycobacterium tuberculosis infection.

Exosomes as double-membrane vesicles contain various contents of lipids, proteins, mRNAs and non-coding RNAs, and involve in multiple physiological processes, for instance intercellular communication and immunomodulation. Currently, numerous studies found that the components of exosomal proteins, nucleic acids or lipids released from host cells are altered following infection with Mycobacterium tuberculosis. Exosomal contents provide excellent biomarkers for the auxiliary diagnosis, efficacy evaluation, and prognosis of tuberculosis. This study aimed to review the current literatures detailing the functions of exosomes in the procedure of M. tuberculosis infection, and determine the potential values of exosomes as biomarkers to assist in the diagnosis and monitoring of tuberculosis.

The PPE2 protein of Mycobacterium tuberculosis is secreted during infection and facilitates mycobacterial survival inside the host.

Mycobacterium tuberculosis secrets various effector proteins to evade host immune responses for facilitating its intracellular survival. The bacterial genome encodes several unique PE/PPE family proteins, which have been implicated to play important role in mycobacterial pathogenesis. A member of this family, PPE2 have been shown to contain a monopartite nuclear localization signal (NLS) and a DNA binding domain. In this study, we demonstrate that PPE2 protein is present in the sera of mice infected with either M. smegmatis expressing PPE2 or a clinical strain of M. tuberculosis (CDC1551). It was found that exogenously added PPE2 can permeate through the macrophage cell membrane and eventually translocate into the nucleus which requires the presence of NLS which showed considerable homology to HIV-tat like cell permeable peptides. Exogenously added PPE2 could inhibit NO production and decreased mycobacterial survival in macrophages. PPE2-null mutant of M. tuberculosis failed to inhibit NO production and had poor survival in macrophages which could be rescued by complementation with full-length PPE2. PPE2-null mutants also had poor survival in the lungs of infected mice indicating that PPE2 even when present in the bloodstream can confer a survival advantage to mycobacteria.

Selected ß-Glucans Act as Immune-Training Agents by Improving Anti-Mycobacterial Activity in Human Macrophages: A Pilot Study.

Epigenetic reprogramming of innate immune cells by ß-glucan in a process called trained immunity leads to an enhanced host response to a secondary infection. ß-Glucans are structural components of plants, algae, fungi, and bacteria and thus recognized as non-self by human macrophages. We selected the ß-glucan curdlan from Alcaligenes faecalis, WGP dispersible from Saccharomyces cerevisiae, and ß-glucan-rich culture supernatant of Alternaria and investigated whether they could produce trained immunity effects leading to an increased control of virulent Mycobacterium tuberculosis. We observed a significant M. tuberculosis growth reduction in macrophages trained with curdlan and Alternaria, which also correlated with increased IL-6 and IL-1ß release. WGP dispersible-trained macrophages were stratified into "non-responders" and "responders," according to their ability to control M. tuberculosis, with "responders" producing higher IL-6 levels. The addition of neutrophils to infected macrophage cultures further enhanced macrophage control of virulent M. tuberculosis, but not in a stimuli-dependent manner. Pathway enrichment analysis of DNA methylome data also highlighted hypomethylation of genes in pathways associated with signaling and cellular reorganization and motility, and "responders" to WGP training were enriched in the interferon-gamma signaling pathway. This study adds evidence that certain ß-glucans show promise as immune-training agents.

ER-dependent membrane repair of mycobacteria-induced vacuole damage.

IMPORTANCE: Tuberculosis still remains a global burden and is one of the top infectious diseases from a single pathogen. Mycobacterium tuberculosis, the causative agent, has perfected many ways to replicate and persist within its host. While mycobacteria induce vacuole damage to evade the toxic environment and eventually escape into the cytosol, the host recruits repair machineries to restore the MCV membrane. However, how lipids are delivered for membrane repair is poorly understood. Using advanced fluorescence imaging and volumetric correlative approaches, we demonstrate that this involves the recruitment of the endoplasmic reticulum (ER)-Golgi lipid transfer protein OSBP8 in the Dictyostelium discoideum/Mycobacterium marinum system. Strikingly, depletion of OSBP8 affects lysosomal function accelerating mycobacterial growth. This indicates that an ER-dependent repair pathway constitutes a host defense mechanism against intracellular pathogens such as M. tuberculosis.

Association between a single nucleotide polymorphism of the ALOX5 gene and susceptibility to multisystem tuberculosis in a Chinese Han population.

BACKGROUND: Host genetic single nucleotide polymorphisms can exert an influence susceptibility to tuberculosis infection. Previous investigations have demonstrated an association between the polymorphism in the ALOX5 gene and a range of diseases, encompassing not only noninfectious conditions like asthma, acute myocardial infarction, and cerebral infarction but also infections caused by various pathogens. However, the relationship between ALOX5 gene polymorphism and susceptibility to tuberculosis has received limited research attention. The ALOX5 gene encodes arachidonic acid 5-lipoxygenase(5-LO), which serves as the initiating catalyst in the generation of the inflammatory mediator leukotriene. Leukotrienes, products derived from the 5-LO pathway, are potent proinflammatory lipid mediators that assume a pivotal role in tuberculosis infections.Consequently, ALOX5 gene variants may be intricately associated with the pathogenesis of tuberculosis. In instances where the host exhibits immunocompromisation, infection with Mycobacterium tuberculosis can impact multiple systems. The involvement of multiple systems significantly augments the complexity of treatment and escalates patient mortality rates. Regrettably, the underlying mechanisms driving multisystem tuberculosis pathogenesis remain enigmatic, with clinicians paying scant attention to this aspect. Although the protein encoded by the ALOX5 gene represents a pivotal enzyme that catalyzes the metabolism of arachidonic acid into LXA4, and thereby plays a significant role in the inflammatory response during tuberculosis infection, studies investigating ALOX5 gene polymorphism and its association with susceptibility to multisystem tuberculosis in the Chinese Han population are exceptionally scarce. Therefore, the primary objective of this study is to comprehensively examine the correlation between ALOX5 gene polymorphisms and susceptibility to tuberculosis within the Chinese Han population, with particular emphasis on multisystemic tuberculosis. METHODS: A case‒control study design was employed, encompassing 382 individuals with pulmonary tuberculosis and 367 individuals with multisystemic tuberculosis as the case groups, along with 577 healthy controls.Whole blood DNA was extracted from all patients and healthy controls. Subsequently, three tag polymorphisms (rs2029253, rs7896431, rs2115819) within the ALOX5 gene were selectively identified and genotyped. RESULTS: After adjusting for age and sex, the presence of allele A at rs2029253 exhibited a pronounced association with an elevated risk of TB susceptibility when compared to the tuberculosis group and healthy control group. (ORa: 2.174, 95% CI: 1.827-2.587; Pa<0.001, respectively). Notably, the rs2029253 AG genotype and AA genotype displayed a significantly increased susceptibility to tuberculosis (ORa: 2.236, 95% CI: 1.769-2.825; Pa <0.001 and ORa: 4.577, 95% CI: 2.950-7.100; Pa <0.001, respectively) compared to the GG genotype. Moreover, in the analysis utilizing genetic models, rs2029253 also exhibited a markedly heightened susceptibility to tuberculosis in additive models, dominant models, and recessive models (Pa <0.001). Conversely, no significant association was observed between rs7896431, rs2115819, and tuberculosis. In the subgroup analysis, when comparing the pulmonary tuberculosis group with the healthy control group, we observed no significant disparities in the distribution frequencies of alleles, genotypes, and gene models (additive model, dominant model, and recessive model) for the three tag SNPs, with P-values were >0.05 after adjusting for age and sex. Additionally, we noted that the presence of allele A at rs2029253 was linked to an increased susceptibility to tuberculosis in the multisystemic tuberculosis group relative to the healthy control group (ORa: 2.292, 95% CI: 1.870-2.810; Pa<0.001). Similarly, the rs2029253 AG genotype, AA genotype, and gene models, including the additive model, dominant model, and recessive model, demonstrated a significantly elevated risk of tuberculosis susceptibility. CONCLUSIONS: The polymorphism in the ALOX5 gene is associated with susceptibility to multisystemic tuberculosis in the Chinese Han population.

Up-regulation of inflammatory reactions by MPT32, a secreted protein of Mycobacterium tuberculosis in RAW264.7 macrophages.

Tuberculosis (TB) is caused by Mycobacterium tuberculosis (Mtb) and is still one of the global health burdens. The occurrence of various cases and multidrug resistance confirm that TB has not been completely conquered. For these reasons, the present research has been conducted to explore TB vaccine and drug candidate possibility using Mtb-secreted proteins. Among these proteins, MPT32 is known to have antigenicity and immunogenicity. There has not been a report on the host immune responses and regulation in macrophage cells. The present study was conducted with MPT32 in RAW 264.7 murine macrophage cells that control immune responses by sensing pathogen invasion and environmental change. We have found that MPT32 could activate lipopolysaccharide (LPS)-induced gene expression of metalloproteinase-9 (MMP-9) and inflammation in RAW 264.7 cells. After treating cells with MPT32, the increase in pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß (IL-1ß) and IL-6, was observed. In addition, activated macrophages expressed inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) to generate various inflammatory mediator molecules, such as nitric oxide (NO). The increase in iNOS and COX-2 levels, which are up-regulators of MMP-9 expression, was also confirmed. The biochemical events are involved in the downstream of activated MAPK signaling and translocation of NF-κ B transcription factor. The present results prove the immunomodulatory effect of MPT32 in the RAW 264.7 murine macrophage cells. it claims the possibility of a TB vaccination and drug candidate using MPT32, contributing to the prevention of TB.

L-selectin-dependent and -independent homing of naïve lymphocytes through the lung draining lymph node support T cell response to pulmonary Mycobacterium tuberculosis infection.

Recruiting large numbers of naïve lymphocytes to lymph nodes is critical for mounting an effective adaptive immune response. While most naïve lymphocytes utilize homing molecule L-selectin to enter lymph nodes, some circulating cells can traffic to the lung-draining mediastinal lymph node (mLN) through lymphatics via the intermediate organ, lung. However, whether this alternative trafficking mechanism operates in infection and contributes to T cell priming are unknown. We report that in pulmonary Mycobacterium tuberculosis-infected mice, homing of circulating lymphocytes to the mLN is significantly less efficient than to non-draining lymph node. CD62L blockade only partially reduced the homing of naïve T lymphocytes, consistent with L-selectin-independent routing of naïve lymphocytes to the site. We further demonstrated that lymphatic vessels in infected mLN expanded significantly and inhibiting lymphangiogenesis with a vascular endothelial growth factor receptor 3 kinase inhibitor reduced the recruitment of intravenously injected naïve lymphocytes to the mLN. Finally, mycobacterium-specific T cells entering via the L-selectin-independent route were readily activated in the mLN. Our study suggests that both L-selectin-dependent and -independent pathways contribute to naïve lymphocyte entry into mLN during M. tuberculosis infection and the latter pathway may represent an important mechanism for orchestrating host defence in the lungs.

Identification of circulating monocytes as producers of tuberculosis disease biomarker C1q.

Tuberculosis (TB) is a prevalent disease causing an estimated 1.6 million deaths and 10.6 million new cases annually. Discriminating TB disease from differential diagnoses can be complex, particularly in the field. Increased levels of complement component C1q in serum have been identified as a specific and accessible biomarker for TB disease but the source of C1q in circulation has not been identified. Here, data and samples previously collected from human cohorts, a clinical trial and a non-human primate study were used to identify cells producing C1q in circulation. Cell subset frequencies were correlated with serum C1q levels and combined with single cell RNA sequencing and flow cytometry analyses. This identified monocytes as C1q producers in circulation, with a pronounced expression of C1q in classical and intermediate monocytes and variable expression in non-classical monocytes.

Tuberculosis is associated with sputum metabolome variations, irrespective of patient sex or HIV status: an untargeted GCxGC-TOFMS study.

INTRODUCTION: Various studies have identified TB-induced metabolome variations. However, in most of these studies, a large degree of variation exists between individual patients. OBJECTIVES: To identify differential metabolites for TB, independent of patients' sex or HIV status. METHODS: Untargeted GCxGC/TOF-MS analyses were applied to the sputum of 31 TB + and 197 TB- individuals. Univariate statistics were used to identify metabolites which are significantly different between TB + and TB- individuals (a) irrespective of HIV status, and (b) with a HIV + status. Comparisons a and b were repeated for (i) all participants, (ii) males only and (iii) females only. RESULTS: Twenty-one compounds were significantly different between the TB + and TB- individuals within the female subgroup (11% lipids; 10% carbohydrates; 1% amino acids, 5% other and 73% unannotated), and 6 within the male subgroup (20% lipids; 40% carbohydrates; 6% amino acids, 7% other and 27% unannotated). For the HIV + patients (TB + vs. TB-), a total of 125 compounds were significant within the female subgroup (16% lipids; 8% carbohydrates; 12% amino acids, 6% organic acids, 8% other and 50% unannotated), and 44 within the male subgroup (17% lipids; 2% carbohydrates; 14% amino acids related, 8% organic acids, 9% other and 50% unannotated). Only one annotated compound, 1-oleoyl lysophosphaditic acid, was consistently identified as a differential metabolite for TB, irrespective of sex or HIV status. The potential clinical application of this compound should be evaluated further. CONCLUSIONS: Our findings highlight the importance of considering confounders in metabolomics studies in order to identify unambiguous disease biomarkers.

Mycobacterial infection alters host mitochondrial activity.

The ability of Mycobacterium tuberculosis (M. tb) to hijack host mitochondria and control host immune signaling is the key to its successful infection. Infection of M. tb causes distinct changes in mitochondrial morphology, metabolism, disruption of innate signaling, and cell fate. The alterations in mitochondria are intricately linked to the immunometabolism of host immune cells such as macrophages, dendritic cells, and T cells. Different immune cells are tuned to diverse immunometabolic states that decide their immune response. These changes could be attributed to the several proteins targeted to host mitochondria by M. tb. Bioinformatic analyses and experimental evidence revealed the potential localization of secreted mycobacterial proteins in host mitochondria. Given the central role of mitochondria in the host metabolism, innate signaling, and cell fate, its manipulation by M. tb renders it susceptible to infection. Restoring mitochondrial health can override M. tb-mediated manipulation and thus clear infection. Several reviews are available on the role of different immune cells in tuberculosis infection and M. tb evasion of immune responses; in the present chapter, we discuss the mitochondrial functional alterations in the innate immune signaling of various immune cells driven by differential mitochondrial immunometabolism during M. tb infection and the role of M. tb proteins, which are directly targeted to the host mitochondria and compromise its innate signaling system. Further studies would help in uncovering the molecular mechanisms of M. tb-directed proteins in host mitochondria to conceptualize both host- directed and pathogen- directed interventions in TB disease management.