Pulmonary Mtb Infection Research Articles

Tuberculosis meningitis coexisting with HIV Infection: a comprehensive review

Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis (TB) in humans, Although Mtb is primarily considered a respiratory pathogen, its ability to spread to and affect the central nervous system (CNS) is of particular interest due to its clinical importance. Tuberculosis meningitis (TBM) is described as the manifestation of Mtb infection in the meninges, leading to inflammation and disease. Individuals with a weakened immune system, particularly those infected with human immunodeficiency virus (HIV), are more susceptible to both pulmonary and extrapulmonary Mtb infection. HIV infection leads to a gradual depletion of CD4 T-cells, severely impairing the host's immune response against pathogens and, thus, predisposes one to several opportunistic infections, including Mtb. Herein, we discuss the current knowledge, potential therapeutic agents, and mechanisms of action and describe various in vivo and in vitro models that may be used to study TBM coexisting with HIV infection.

The Microbiota of Human Lung of Pulmonary Tuberculosis and the Alteration Caused by Anti-tuberculosis Drugs

An in-depth understanding of the lung microbiota in tuberculosis (TB) infection could provide optimal strategies for the prophylaxis, diagnosis, and treatment of the disease. Only a few studies have investigated the impact of Mycobacterium tuberculosis (Mtb) infection and anti-TB treatment on the lung microbiota. Bronchoalveolar lavage fluid and blood samples were collected from 23 active TB patients (TBZ), 17 latent tuberculosis infection patients (LTBI), 13 healthy controls (HC), and 14 active TB patients with 1-month anti-TB therapy (TBM) for 16S RNA sequencing and serological indexes, respectively. Low body mass index, albumin, and total triglyceride levels were detected in TBZ. Pulmonary Mtb infection led to a minor decrease in the alpha diversity of the lung microbiota in TBZ than HC, but a significant difference was noted in beta diversity. Subsequently, anti-TB therapy caused a rapid alteration in the lung community structure due to reduced alpha and beta diversity. Proteobacteria were abundant in TBZ samples, while Firmicutes was predominant in the LITB and HC samples. Lactobacillus and Subdoligranulum (genera) were the most unique in the LTBI and HC group, respectively. The TBM group showed the most predominant abundance of Bacteroides, Oscillospira, and Ruminococcus (genera). Functional pathways, such as indole alkaloid biosynthesis, Wnt signaling pathway, endocytosis, and metabolism of xenobiotics by cytochrome P450, significantly decreased in the TBM group compared with TBZ group. Pulmonary TB and anti-TB treatment caused a distinct dysbiosis of the lung microbiome. The current findings suggested potential links between the lung microbiota and TB onset, progression, and treatment.

Mycobacterium avium via oral route provide enhanced protection against pulmonary tuberculosis: An essential role of B cells and germinal centers

Abstract Human tuberculosis (TB) is caused mainly by Mycobacterium tuberculosis (Mtb) and represents an enormous challenge to global health because of the inadequacy of currently available drugs and vaccines. The most common clinical manifestation is pulmonary TB, and the Bacille Calmette Guerin (BCG) is the only licensed vaccine for protection against TB; however, its efficacy is highly variable. Today at least 52 countries have reported MDR-TB and XDR-TB cases which cannot be cured or contained by current TB therapy. Thus, there is an urgent need to develop new therapies/vaccines that effectively prevent or cure TB. In the last decade, lymphoid follicles (LFs) having ectopic GCs (eGC) have been identified in the lungs of Mtb infected animals and humans; however, a mechanism of its formation and role has not been adequately characterized. Our studies found that mice vaccinated with BCG and exposed to Mycobacterium avium [a non-tuberculous mycobacterium (NTM)] via drinking water provide long-term protection from pulmonary TB than BCG alone as determined by reduced Mtb bacterial burden and inflammatory progression of infection. Interestingly, these mice also developed eGC in the lungs and have increased B cells, anti-Mtb cell lysate specific IgA, and IgG antibodies. These findings suggest that NTM and B cells play a critical role in generating protective immunity against pulmonary Mtb infection, and the formation of eGC in these mice is a crucial factor in this improved immunity. Investigating the mechanism of eGC formation and the role of NTM and B cells in its stimulation is an important question to understand TB pathogenesis and to develop effective vaccines and therapies. Supported by NIH/NIAID - 1R01AI127475-01AI, Colorado State University's MIP research team grant

β-Glucan Induces Protective Trained Immunity against Mycobacterium tuberculosis Infection: A Key Role for IL-1

Summaryβ-glucan is a potent inducer of epigenetic and functional reprogramming of innate immune cells, a process called “trained immunity,” resulting in an enhanced host response against secondary infections. We investigate whether β-glucan exposure confers protection against pulmonary Mycobacterium tuberculosis (Mtb) infection. β-glucan induces trained immunity via histone modifications at gene promoters in human monocytes, which is accompanied by the enhanced production of proinflammatory cytokines upon secondary Mtb challenge and inhibition of Mtb growth. Mice treated with β-glucan are significantly protected against pulmonary Mtb infection, which is associated with the expansion of hematopoietic stem and progenitor cells in the bone marrow and increased myelopoiesis. The protective signature of β-glucan is mediated via IL-1 signaling, as β-glucan shows no protection in mice lacking a functional IL-1 receptor (IL1R−/−). The administration of β-glucan may be used as a novel strategy in the treatment of mycobacterial infections and possibly as an adjuvant to improve anti-tuberculosis vaccines.

Alveolar macrophages provide an early Mycobacterium tuberculosis niche and initiate dissemination

Abstract Mycobacterium tuberculosis (Mtb) infection is initiated in the distal airways, a site patrolled by alveolar macrophages (AM), and infection outcomes are governed by early immune events that remain poorly understood. We show that Mtb replicates almost exclusively within airway-resident AM during the first week after aerosol exposure. Although AM are typically thought to reside within the alveolar lumen, confocal imaging demonstrated that Mtb-infected AM establish a novel niche within the lung interstitium, where they undergo proliferation and physically associate with recruited monocyte-derived cells (MC). Localization of AM to the interstitium precedes subsequent Mtb uptake by MC and neutrophils and is driven by non-hematopoietic MyD88/IL-1R inflammasome signaling and the Mtb ESX-1 secretion system. The interstitial localization of infected AM occurs in the absence of recruited monocytes and neutrophils, suggesting that this pulmonary niche may result from AM-intrinsic egress from the alveolar space rather than inflammation-driven alveolar consolidation. Comparisons of the transcriptomes of infected AM localized to the airway or interstitium by RNA-sequencing revealed unique transcriptional profiles in the two populations, suggesting that these two AM subsets may be functionally distinct. For example, Mtb-infected AM in the interstitium dramatically upregulated interferon-response genes, lending support to the idea that these cells are in unique lung environments. Thus, crosstalk between Mtb-infected AM and non-hematopoietic cells establishes pulmonary Mtb infection by promoting the translocation of infected cells from the alveoli to lung interstitium, facilitating dissemination to other myeloid subsets.

Inoculum size and traits of the infecting clinical strain define the protection level against Mycobacterium tuberculosis infection in a rabbit model.

Host protective immunity against pathogenic Mycobacterium tuberculosis (Mtb) infection is variable and poorly understood. Both prior Mtb infection and BCG vaccination have been reported to confer some protection against subsequent infection and/or disease. However, the immune correlates of host protection with or without BCG vaccination remain poorly understood. Similarly, the host response to concomitant infection with mixed Mtb strains is unclear. In this study, we used the rabbit model to examine the host response to various infectious doses of virulent Mtb HN878 with and without prior BCG vaccination, as well as simultaneous infection with a mixture of two Mtb clinical isolates HN878 and CDC1551. We demonstrate that both the ability of host immunity to control pulmonary Mtb infection and the protective efficacy of BCG vaccination against the progression of Mtb infection to disease is dependent on the infectious inoculum. The host response to infection with mixed Mtb strains mirrors the differential responses seen during infection with each of the strains alone. The protective response mounted against a hyperimmunogenic Mtb strain has a limited impact on the control of disease caused by a hypervirulent strain. This preclinical study will aid in predicting the success of any vaccination strategy and in optimizing TB vaccines.

Mycobacterium tuberculosis Sulfolipid-1 Activates Nociceptive Neurons and Induces Cough

SummaryPulmonary tuberculosis, a disease caused by Mycobacterium tuberculosis (Mtb), manifests with a persistent cough as both a primary symptom and mechanism of transmission. The cough reflex can be triggered by nociceptive neurons innervating the lungs, and some bacteria produce neuron-targeting molecules. However, how pulmonary Mtb infection causes cough remains undefined, and whether Mtb produces a neuron-activating, cough-inducing molecule is unknown. Here, we show that an Mtb organic extract activates nociceptive neurons in vitro and identify the Mtb glycolipid sulfolipid-1 (SL-1) as the nociceptive molecule. Mtb organic extracts from mutants lacking SL-1 synthesis cannot activate neurons in vitro or induce cough in a guinea pig model. Finally, Mtb-infected guinea pigs cough in a manner dependent on SL-1 synthesis. Thus, we demonstrate a heretofore unknown molecular mechanism for cough induction by a virulent human pathogen via its production of a complex lipid.

Diagnostic accuracy of in-house real-time PCR assay for Mycobacterium tuberculosis: a systematic review and meta-analysis

BackgroundIn recent years, studies on the diagnostic accuracy of in-house real-time PCR (hRT-PCR) assay for the detection of Mycobacterium tuberculosis (Mtb) have been reported with unignorable discrepancies. To assess the overall accuracy of the hRT-PCR assay for Mtb diagnosis in different samples for individuals with active pulmonary and extra-pulmonary Mtb infection, a systematic review and meta-analysis were performed.MethodsThe PUBMED, EMBASE, Web of Science, and Cochrane databases were searched up to June 2017 for eligible studies that estimated diagnostic sensitivity and specificity with the hRT-PCR assay in respiratory and non-respiratory samples in pulmonary and extra-pulmonary Mtb infection patients, with Mtb culture as the reference standard. Bivariate random effect models were used to provide pooled estimation of diagnostic accuracy. Further, subgroup and meta-regression analyses were performed to explore sources of heterogeneity. The risk of bias was assessed by the QUADAS-2 tool.ResultsOf the 3589 candidate studies, 18 eligible studies met our inclusion criteria. Compared to Mtb culture data, the pooled sensitivity and specificity were 0.96 and 0.92, respectively. The diagnostic odds ratio (DOR) was 192.96 (95% CI 68.46, 543.90), and the area under the summary ROC curve (AUC) was 0.9791. There was significant heterogeneity in sensitivity and specificity among the enrolled studies (p < 0.001). The studies with high-quality assessment and application of respiratory specimen were associated with better accuracy.ConclusionsIn low-income/high-burden settings, our results suggested that the hRT-PCR assay could be a useful test for the diagnosis of TB with high sensitivity and specificity.

B7x expression on the bronchial epithelium delays CD4 T cell priming after Mycobacterium tuberculosis infection

Abstract The priming of CD4 T cell responses after Mycobacterium tuberculosis (Mtb) infection is delayed compared to other pathogens. Although it has been thought that the slow replication rate of the bacteria is responsible for the lag in T cell priming, it is possible that negative regulatory pathways that control inflammation in the lung also contribute to this slow T cell response. Here we show that the orphan co-inhibitory ligand B7x contributes to the delay in T cell priming following Mtb infection. We find that B7x is expressed on bronchiolar epithelial cells, and B7x−/− mice display enhanced bacterial control and survive longer than WT mice after pulmonary Mtb infection. The enhanced resistance of B7x−/− mice to Mtb infection is completely dependent on CD4 T cells, but at day 28 post-infection, the peak of the T cell response, there is little difference in the number or function of Mtb-specific effector CD4 T cells. Instead, we find that Mtb-specific CD4 T cells are primed in the mediastinal lymph nodes (mLNs) and accumulate in the lungs much earlier in B7x−/− mice compared to WT mice. B7x blockade is also able to enhance Mtb-specific T cell priming even after inoculation of non-replicating irradiated Mtb into the lungs. Moreover, after ESAT-6/CpG lung immunization, B7x blockade reduces the amount of adjuvant needed to drive T cell expansion in the mLNs by ~50 fold. Collectively, our results indicate the expression of the immune checkpoint molecule B7x by the bronchial epithelium contributes in part to the notorious lag in CD4 T cell responses during tuberculosis by increasing the threshold of innate stimuli needed for the induction of T cell priming.

Host Resistance to pulmonary Mycobacterium tuberculosis infection requires CD153 expression

Abstract Mycobacterium tuberculosis infection (Mtb) is the leading cause of death due to a single infectious agent and is among the top ten causes of all human deaths worldwide. CD4 T cells are essential for resistance to Mtb infection, and for decades it has been thought that IFNγ production is the primary mechanism of CD4 T-cell-mediated protection. However, IFNγ responses do not correlate with host protection, and several reports demonstrate that additional anti-tuberculosis CD4 T-cell effector functions remain unaccounted for. Here we show that the tumour-necrosis factor (TNF) superfamily molecule CD153 (encoded by the gene Tnfsf8) is required for control of pulmonary Mtb infection by CD4 T cells. CD153-deficient mice develop high pulmonary bacterial loads and succumb early to Mtb infection. Reconstitution of T-cell-deficient hosts with Tnfsf8−/− CD4 T cells, similar to Ifng−/− CD4 T cells, fails to rescue mice from early mortality. In Mtb-infected non-human primates, Mtb-specific CD153-expressing CD4 T cells inversely correlates with bacterial loads in granulomas. In Mtb-infected humans, CD153 defines a subset of highly polyfunctional Mtb-specific CD4 T cells that are much more abundant in individuals with latent Mtb infection compared to those with active tuberculosis. Thus, CD153 is a major immune mediator of host protection against pulmonary Mtb infection and CD4 T cells are one important source of this molecule.

Dendritic Cell-Induced Dissemination of Mycobacterium tuberculosis into the Central Nervous System

Abstract Central nervous system tuberculosis (CNSTB) is the most serious manifestation of extrapulmonary tuberculosis infections representing 5–10% of all cases. Knowledge of CNSTB stems from observations made in 1933 by pathologists Rich and McCordock, who found caseating foci in the brain parenchyma or meninges that rupture into the subarachnoid space leading to diffuse meningeal infection. However, the mechanism of how immobile Mycobacterium tuberculosis, the causative agent of TB crosses the highly regulated blood-brain barrier (BBB) and enters the CNS forming these foci and infection is unknown. We have developed a novel in vitro BBB model that combines Mycobacteria-infected DCs, PBMCs, and primary mouse brain endothelial cells to observe early cellular interaction and aggregate formation at the BBB similar to previous observations by Rich and McCordock. With this model we observed Mycobacteria impedes migration of DCs but increases DC initiation of aggregation on a primary mouse endothelial BBB with a cellular composition similar to pulmonary granulomas. Aggregates induce an inflammatory response by increasing ICAM-1 and VCAM-1 expression and dysregulation of tight junction proteins and mitochondria suggesting barrier damage possibly due to increased iNOS expression observed during within aggregates. Mycobacteria-infected DC transmigration is increased in the presence of aggregation. Previous research from our lab and others have shown DC transmigration across an in vitro BBB rely on MMP-9 secretion while TNF-alpha is necessary for pulmonary granuloma formation and Mtb infection control. Here we will show the role iNOS, MMP-9 and TNF-alpha play in DC-mediated dissemination of Mycobacterium across the BBB.

Mucosal boosting of H56:CAF01 immunization promotes lung-localized T cells and an accelerated pulmonary response to Mycobacterium tuberculosis infection without enhancing vaccine protection

T cell-mediated protection against Mycobacterium tuberculosis (Mtb) is dependent upon the ability to localize within the site of pulmonary infection and directly interact with infected cells. In turn, vaccine strategies to improve rapid T cell targeting of Mtb-infected cells after pulmonary exposure are being actively pursued. Given parenterally, the subunit vaccine H56:CAF01 elicits polyfunctional CD4 T cells that localize to the lung parenchyma and confer durable protection. Here, we find that airway mucosal boosting of parenteral H56:CAF01 immunization greatly enhances the population of long-lived lung-resident T cells (Trm) and increases early vaccine T cell responses to pulmonary Mtb challenge in multiple mouse models. However, mucosal boosting does not alter the Th1/17 vaccine signature typical of H56:CAF01 and does not further improve durable control of pulmonary infection following aerosol Mtb-challenge. Additional mucosal boosting with H56:CAF01 further enhances the Trm response without further improving protection, while blocking the recruitment of non-Trm with FTY720-treatment failed to exposed Trm-mediated protection in mucosally boosting animals. These results demonstrate the limitations of maximizing lung-localized Trm in vaccine control of pulmonary Mtb infection, especially within an immunization protocol that is already optimized for the induction of mucosal-homing Th17 cells.

HIV-1 and SIV Infection Are Associated with Early Loss of Lung Interstitial CD4+ T Cells and Dissemination of Pulmonary Tuberculosis

SUMMARYLung interstitial CD4+ T cells are critical for protection against pulmonary infections, but the fate of this population during HIV-1 infection is not well described. We studied CD4+ T cells in the setting of HIV-1 infection in human lung tissue, humanized mice, and a Mycobacterium tuberculosis (Mtb)/simian immunodeficiency virus (SIV) nonhuman primate co-infection model. Infection with a CCR5-tropic strain of HIV-1 or SIV results in severe and rapid loss of lung interstitial CD4+ T cells but not blood or lung alveolar CD4+ T cells. This is accompanied by high HIV-1 production in these cells in vitro and in vivo. Importantly, during early SIV infection, loss of lung interstitial CD4+ T cells is associated with increased dissemination of pulmonary Mtb infection. We show that lung interstitial CD4+ T cells serve as an efficient target for HIV-1 and SIV infection that leads to their early depletion and an increased risk of disseminated tuberculosis.

166. TGF-β Restricts T-cell IFNg Production in Pulmonary Tuberculous Granulomas

BackgroundIFNg production by CD4 T cells has been thought to be critical for immunity against Mycobacterium tuberculosis (Mtb); however, recent studies show that IFNg-producing CD4 T cells are more effective at preventing dissemination than controlling Mtb in the lung. Because optimal control of Mtb infection requires direct interactions between CD4 T cells and Mtb-infected cells presenting cognate antigen on MHCII, we sought to determine the location of CD4 T-cell antigen recognition and IFNg production in the Mtb-infected lung.MethodsWe infected mice with an ultra-low dose (ULD) of Mtb (1–3 CFU), a model developed in our laboratory which results in well-circumscribed granulomas that recapitulate many features of human Mtb granulomas. Using immunohistochemistry and quantitative imaging, we examined their lungs 35 days later for phenotypic and spatial analysis of T-cell receptor (TCR) signaling (using IRF4) and IFNγ production. We tested the antigen specificity of these responses with an adoptive transfer of both Mtb-specific and OVA-specific control CD4 T cells into ULD Mtb-infected mice. To assess the role of TGFβ signaling on T-cell localization and function, we performed the same analysis in mice lacking the TGFβ receptor (TGFβR) on T cells.ResultsWithin Mtb-infected lungs, many T cells localize near Mtb cells and undergo TCR signaling. Despite this, we found very few cells producing IFNγ within the granuloma (Figure 1). In our adoptive transfer experiment, both cell types infiltrated the granuloma. The Mtb-specific, but not OVA-specific, T cells had active TCR engagement though only a small fraction of these cells produced IFNγ, and this IFNγ was diminished near Mtb (Figure 2). Conversely, in the TGFβR conditional knockout, we found increased IFNγ production that was highest within the granuloma (Figure 3).ConclusionDespite ongoing TCR stimulation in T cells, IFNγ production is restricted in areas where cognate interactions are most likely to occur. TGFβ plays a critical role in mediating this effect, as T cells lacking the receptor can produce more IFNγ near infected cells. These findings help explain why IFNγ-producing T cells have limited capacity to control pulmonary Mtb infection and could guide new strategies for vaccine and immunotherapeutic development. Disclosures All authors: No reported disclosures.

Alveolar Macrophages Provide an Early Mycobacterium tuberculosis Niche and Initiate Dissemination

Mycobacterium tuberculosis (Mtb) infection is initiated in the distal airways, but the bacteria ultimatelydisseminate to the lung interstitium. Although various cell types, including alveolar macrophages (AM), neutrophils, and permissive monocytes, are known to be infected with Mtb, the initially infected cells as well as those that mediate dissemination from the alveoli to the lung interstitium are unknown. In this study, using a murine infection model, we reveal that early, productive Mtb infection occurs almost exclusively within airway-resident AM. Thereafter Mtb-infected, but not uninfected, AM localize to the lung interstitium through mechanisms requiring an intact Mtb ESX-1 secretion system. Relocalization of infected AM precedes Mtb uptake by recruited monocyte-derived macrophages and neutrophils. This dissemination process is driven by non-hematopoietic host MyD88/interleukin-1 receptor inflammasome signaling. Thus, interleukin-1-mediated crosstalk between Mtb-infected AM and non-hematopoietic cells promotes pulmonary Mtb infection by enabling infected cells to disseminate from the alveoli to the lung interstitium.

Gut microbiota associated with pulmonary tuberculosis and dysbiosis caused by anti-tuberculosis drugs

An improved understanding of the gut microbiota could lead to better strategies for the diagnosis, therapy and prophylaxis of tuberculosis (TB). The impact of both Mycobacterium tuberculosis (Mtb) infection and anti-TB treatment on the gut microbiota has rarely been studied. We characterized the diversity and composition of the gut microbiota in pulmonary TB patients as well as the effects of anti-TB drugs on the gut microbiota. Pulmonary Mtb infection led to a minor decrease in the α diversity of the gut microbiota when compared to healthy controls, which mainly resulted from changes in the relative abundance of the members of genus Bacteroides. Anti-TB therapy caused a rapid, significant alteration in the community structure. The relative abundance of members of genus Clostridiales of the phylum Firmicutes significantly decreased during anti-TB treatment, while many members of genus Bacteroides, including Bacteroides OTU230 and Bacteroides fragilis, were among the taxa that increased. OTU8 and OTU2972 assigned to family Erysipelotrichaceae of the phylum Firmicutes showed a dramatic increase 1 week after the start of therapy, while the other members of this family decreased. Pulmonary TB and anti-TB treatment caused a distinct dysbiosis of the gut microbiota. Our study contributes valuable information implying potential links between the gut microbiota and TB.

Systemic Immunological changes in patients with distinct clinical outcomes during Mycobacterium tuberculosis infection

BackgroundThe lung lesions in an individual infected with tuberculosis (TB) are surprisingly variable and independent of each other. However, there is no circulating biomarker yet able to segregate patients according to the extent of lung lesions. Materials and methodsIn this study, the phenotypic and functional profile of leukocytes of patients with active pulmonary tuberculosis (TB) and controls (CO) were fully scrutinized by immunophenotyping assays and in vitro short-term whole blood culture. The TB group was subdivided according to the extent of lung lesions as unilateral (UNI) and bilateral (BI). ResultsThe results show that TB group display an altered leukocyte profile in the peripheral blood with significant lower counts of NK-cells, CD3+CD56+CD16+/− NKT-cells, CD4+T-cells, CD19+B-cells when compared to CO. Increased CD4+T-cells and CD8+T-cell activation was observed by the upregulation of activation markers (HLA-DR) as well as of chemokine receptors (CCR2, CCR3, and CXCR4). In addition, TB group presented a significant decrease proportion of CD14LowCD16+ monocytes despite the increase in HLA-DR expression. Regarding the severity of the disease, in the BI group a reduction in frequency of CD19+CD5+ B-cells and expression of HLA-DR in CD14LowCD16+ monocytes was observed. Furthermore, the extent of lung lesions influences the production of molecules as observed by significantly larger production of IL-4 by neutrophils, total T-cells, CD4+T-cells, CD8+T-cells and CD19+B-cells in UNI as compared to BI. By contrast, in BI group the frequency of high producers of both IL-17+CD4+T-cells and IL-17+CD8+T-cells were significantly increased than UNI, suggesting the deleterious role of these subsets during active pulmonary Mtb infection. ConclusionThe immunophenotypic characterization of unilateral and bilateral active TB performed in the present study indicates that the extent of lung lesion could be associated with a fine-tuning between immunological responses during untreated Mtb infection.

Eosinophils actively participate in the host response to Mtb infection

Abstract Mycobacterium tuberculosis (Mtb) is the leading cause of mortality worldwide due to a single infectious agent. Mtb resides in pulmonary macrophages, neutrophils and phagocytes, which are also key effector cells that limit bacterial growth. The role of other granulocytic phagocytes during Mtb infection is largely unknown. Eosinophils have comparable phagocytic function to neutrophils and an overlapping repertoire of granular contents capable of limiting bacterial growth. Here we characterized eosinophil responses to Mtb in vitro and in vivo. Eosinophils from healthy human donors responded to Mtb in vitro by CD69 up-regulation and CD62L down-regulation. Mtb induced the release of the eosinophil-specific granule proteins EDN and ECP. Mtb exposure resulted in secretion of IL-1a, TNFa, MIP-1a, MIP-1b, and IL-8. These results suggest that Mtb directly affects eosinophil degranulation, and that eosinophils are able to contribute to Mtb-driven inflammation. To investigate whether eosinophils actively participate in the cellular immune response to Mtb in vivo we followed the eosinophilic response after pulmonary Mtb infection in non-human primates. Eosinophils were significantly increased in the BAL fluid of Mtb infected rhesus macaques, providing evidence that eosinophils are being actively recruited to the lung in response to Mtb in vivo. Indeed, when we analyzed Mtb infected lung tissue from patients who had undergone lung resection, eosinophils were enriched in fibrotic TB consolidations, cavity wall and necrotic caseum. We are currently investigating the biological relevance of the eosinophilic response during Mtb infection in the mouse model. This work was supported by the intramural research programs of NIAID, NIH.

CD4+ T Cells Recognizing PE/PPE Antigens Directly or via Cross Reactivity Are Protective against Pulmonary Mycobacterium tuberculosis Infection

Mycobacterium tuberculosis (Mtb), possesses at least three type VII secretion systems, ESX-1, -3 and -5 that are actively involved in pathogenesis and host-pathogen interaction. We recently showed that an attenuated Mtb vaccine candidate (Mtb Δppe25-pe19), which lacks the characteristic ESX-5-associated pe/ppe genes, but harbors all other components of the ESX-5 system, induces CD4+ T-cell immune responses against non-esx-5-associated PE/PPE protein homologs. These T cells strongly cross-recognize the missing esx-5-associated PE/PPE proteins. Here, we characterized the fine composition of the functional cross-reactive Th1 effector subsets specific to the shared PE/PPE epitopes in mice immunized with the Mtb Δppe25-pe19 vaccine candidate. We provide evidence that the Mtb Δppe25-pe19 strain, despite its significant attenuation, is comparable to the WT Mtb strain with regard to: (i) its antigenic repertoire related to the different ESX systems, (ii) the induced Th1 effector subset composition, (iii) the differentiation status of the Th1 cells induced, and (iv) its particular features at stimulating the innate immune response. Indeed, we found significant contribution of PE/PPE-specific Th1 effector cells in the protective immunity against pulmonary Mtb infection. These results offer detailed insights into the immune mechanisms underlying the remarkable protective efficacy of the live attenuated Mtb Δppe25-pe19 vaccine candidate, as well as the specific potential of PE/PPE proteins as protective immunogens.

Pharmacologic Inhibition of Host Phosphodiesterase-4 Improves Isoniazid-Mediated Clearance of Mycobacterium tuberculosis.

The lengthy duration of multidrug therapy needed to cure tuberculosis (TB) poses significant challenges for global control of the disease. Moreover, chronic inflammation associated with TB leads to pulmonary damage that can remain even after successful cure. Thus, there is a great need for the development of effective shorter drug regimens to improve clinical outcome and strengthen TB control. Host-directed therapy (HDT) is emerging as a novel adjunctive strategy to enhance the efficacy and shorten the duration of TB treatment. Previously, we showed that the administration of CC-3052, a phosphodiesterase-4 inhibitor (PDE4i), reduced the host inflammatory response during Mycobacterium tuberculosis (Mtb) infection and improved the antimicrobial efficacy of isoniazid (INH) in both the mouse and rabbit models. In the present study, we evaluated the pharmacokinetics and explored the mechanism underlying the efficacy of a more potent PDE4i, CC-11050, as adjunct to INH treatment in a mouse model of pulmonary Mtb infection. Genome-wide lung transcriptome analysis confirmed the dampening of inflammation and associated network genes that we previously reported with CC-3052. Consistent with the reduction in inflammation, a significant improvement in Mtb control and pathology was observed in the lungs of mice treated with CC-11050 plus INH, compared to INH alone. This important confirmatory study will be used to help design upcoming human clinical trials with CC-11050 as an HDT for TB treatment.