Biology subjects

Kaushal, D.

Publications and source records attributed to Kaushal, D..

18 recordsLinked to original sources

bioRxiv2026

Bronchus associated lymphoid tissue induced by an attenuated Mycobacterium tuberculosis vaccine prevents tuberculosis from heterologous TB challenge

Tuberculosis (TB) remains a leading cause of death from infectious disease worldwide, underscoring the urgent need for vaccines with greater and more consistent efficacy than Bacille Calmette-Guerin (BCG). We previously showed that mucosal vaccination with an isogenic Mycobacterium tuberculosis (Mtb) mutant lacking the stress-response transcription factor SigH ({Delta}sigH) prevents pulmonary TB in two macaque species. In the absence of SigH, Mtb is unable to effectively counter host oxidative stress. Vaccinated macaques were notably protected from disease, exhibiting an absence of granulomatous pathology together with the formation of lymphoid follicles and robust antigen-specific CD4+ and CD8+ T cell responses, identifying{Delta} sigH as a promising live-attenuated TB vaccine candidate. The{Delta} sigH mutant used in prior studies was generated in the Mtb CDC1551 background, a commonly used laboratory strain for challenge studies. Here, we evaluated whether{Delta} sigH-mediated protection extends to heterologous challenge with the more virulent Mtb Erdman strain. Mucosal{Delta} sigH vaccination conferred significant protection against heterologous challenge, markedly reducing pulmonary bacterial burden and TB-associated pathology. Longitudinal high-resolution PET/CT imaging demonstrated that aerosol{Delta} sigH vaccination induced robust inducible bronchus-associated lymphoid tissue (iBALT) responses in the lung. Unlike granulomas, these iBALT structures resolved over time while remaining associated with protection against subsequent Mtb challenge. Protection of highly susceptible rhesus macaques against virulent heterologous Mtb challenge following aerosol{Delta} sigH vaccination supports the further preclinical development of{Delta} sigH-based live-attenuated TB vaccines and highlights iBALT induction as a potential correlate and mechanistic driver of protective immunity against TB.

immunology↗

bioRxiv2025

Therapeutic remodeling of the tuberculosis granuloma with 1-methyl-D-tryptophan enhances CD8+ T cell-macrophage interactions

Granulomas, the hallmark of tuberculosis (TB) disease, can both restrict Mycobacterium tuberculosis (Mtb) dissemination and impede its clearance. Recent studies indicate that indoleamine 2,3-dioxygenase (IDO1), an immunosuppressive metabolic enzyme, limits infiltration of activated T cells and can contribute to TB disease progression. Treatment with 1-methyl-D-tryptophan (D-1MT), a small molecule inhibitor that restores mTOR signaling, has been shown to improve immune responses Mtb-infected rhesus macaques. Here, we investigated the impact of D-1MT treatment on TB granuloma architecture using 30-plex high-dimensional issue imaging in rhesus macaques. By spatially mapping 13 distinct cell populations, we found D-1MT treatment corresponded with significantly increased infiltration CD8+ T cells into granulomas compared to untreated controls. Notably, these CD8+ T cells expressed markers of cell proliferation and cytotoxicity. D-1MT enhanced CD8+ T cell infiltration throughout the granuloma, with particularly pronounced effects in the myeloid core, where we observed significantly enhanced spatial interactions between macrophages and CD8+, but not CD4+ T cells. Our results demonstrate that: (i) effective intra-granulomatous Mtb control is associated with the close spatial proximity between CD8+ T cells and macrophages, a feature less abundant in uncontrolled pulmonary TB; (ii) IDO1 induction blocks CD8+ T cell infiltration and reduces T cell activation and proliferation; and (iii) therapeutic strategies, including D-1MT, that improve intra-granulomatous killing hold strong translational potential. Significance statementOur understanding of immune mechanisms within the TB granuloma has advanced greatly with the advent of high-resolution single cell multiplexed imaging. Using such imaging, we show that TB granulomas in rhesus macaques, a highly translational model of human TB pathology, are characterized by IDO1-mediated immunoregulation. Early pharmacologic restoration of mTOR signaling via D-1MT treatment can reduce IDO1 enzymatic activity and facilitate the recruitment and function of CD8+ T cells within the granulomas. These findings reveal specific mechanisms exploited by Mtb to maintain intra-granuloma persistence and underscore immune responses. Future vaccine and therapeutic design should consider these immunoregulatory features to achieve better control of Mtb infection.

immunology↗

bioRxiv2025

Single dose alum adjuvanted RBD protein vaccine provides protection against homologous challenge with SARS-CoV-2 Washington strain and heterologous rechallenge with Delta and Omicron BA.5 variants in K18 hACE2 mouse model

Most COVID-19 vaccination strategies require at least two doses-a primary vaccine followed by a booster dose with the updated variant-specific vaccine. However, vaccine and booster dose hesitancy make it challenging to administer multiple doses of vaccines in the unvaccinated and vaccinated populations, respectively. Thus, it is important to determine if vaccinated individuals exposed to SARS-CoV-2 infection develop immune responses that may protect them against emerging variants of concern (VoCs). We have developed mouse models to understand the protective efficacy of an RBD-based vaccine against challenge and rechallenge with SARS-CoV-2 VoCs. Mice were vaccinated with RBD protein vaccine formulated in 2% Alhydrogel (alum) adjuvant by subcutaneous route. To determine the efficacy of RBD vaccine, mice were challenged approximately 4 weeks post-vaccination with SARS-CoV-2 variants by intranasal route. To determine the efficacy of RBD vaccine against SARS-CoV-2 rechallenge, mice were rechallenged with SARS-CoV-2 Delta and Omicron BA.5 variants at day 14 post SARS-CoV-2 Washington (WA) strain challenge. Our data suggest that single dose alum adjuvanted RBD protein from Wuhan strain provides protection against homologous challenge with SARS-CoV-2 WA strain but failed to provide protection against heterologous challenge with Delta and Omicron BA.5 variants. Interestingly, vaccinated mice that survived homologous challenge with the WA strain showed protection against heterologous rechallenge with Delta and omicron BA.5 variants. Furthermore, infectious viral loads of Delta and Omicron BA.5 were not detected in the lung tissues collected from the rechallenged mice at 3 days post-rechallenge. The data suggest that a single dose RBD vaccine from the ancestral Wuhan strain together with survival from WA strain challenge induces protective immune responses against Delta and Omicron BA.5 variants rechallenge. These mouse models will be useful to determine the immune responses that correlate with protection against challenge and rechallenge with SARS-CoV-2 VoCs.

microbiology↗

bioRxiv2025

PU.1 Shapes Host Epigenetic Responses to Mycobacterium tuberculosis and Represents a Target for Host Directed Therapy

Mycobacterium tuberculosis (Mtb) infection induces widespread epigenetic changes in monocytes and macrophages. To characterize the critical factor responsible for Mtb-induced epigenetic responses, we performed an integrative analyses of chromatin modification (H3K27ac), transcription factor binding, and gene expression in human monocytes and macrophages. Our data identified PU.1 (SPI1) as a key transcriptional regulator to be upregulated and bound to gene promoters in response to Mtb. PU.1 enrichment was associated with enhanced proinflammatory and anti-apoptotic gene programs. PU.1 expression was also elevated in the lung tissues from Mtb-infected macaques and tuberculosis patients. PU.1 knockdown enhanced human macrophage apoptosis, dampened inflammation and reduced bacterial survival, suggesting its functional role in supporting Mtb infection. Pharmacological inhibition of PU.1 replicated the effects of knockdown, restricting Mtb growth without cytotoxicity. These findings identify PU.1 as a central node in the host epigenetic response to Mtb and a candidate for host-directed therapeutic targeting in tuberculosis.

immunology↗

bioRxiv2025

Using Imaris to rigorously track PET-defined sites of lung inflammation in Mycobacterium tuberculosis-exposed non-human primates

Aerosol exposure of non-human primates (NHPs) to Mycobacterium tuberculosis (Mtb) typically results in discrete sites of inflammation of the lung that is detectable by 2-deoxy-2-[fluorine-18]fluoro-D-glucose (18F-FDG)-based PET/CT scans. Such scans are often analyzed using software such as Invicro VivoQuant or OsiriX as 3D images by manual labeling sites of PET signal using 2D slices and by reporting maximal standardized uptake value (SUVmax) either of the whole lung or of individual lesions. Here we propose a pipeline for analysis of the same PET/CT scans using Imaris, a proprietary software typically used for analysis of fluorescent microscopy data. We show that by using locations of spine vertebra (denoted as "landmarks") we can align serials scans of the same animal, and by using automated (with some manual corrections) image segmentation of PET scans in 3D as "surfaces", we can accurately define location of all sites of inflammation in the lung and lung-associated thoracic lymph nodes (LNs). We show that there is an excellent correlation between individual lesions SUVmax determined by VivoQuant and maximum intensity determined by Imaris suggesting utility of this approach. Imaris also provides wealth of additional information for each of the identified lesions such as volume, location, shape, surface area, and others, and each lesion can be exported in Virtual Reality file format (.wrl) allowing for detailed and rigorous analyses of how features of these PET-defined lesions evolve over time and correlate with the outcome of infection and/or treatment.

immunology↗

bioRxiv2025

The genomic legacy of selectively breeding rhesus macaques for HIV/AIDS-related research

BackgroundRhesus macaques are the preferred non-human primate model for HIV/AIDS-related research. Specific major histocompatibility complex (MHC) haplotypes and genetic ancestry are linked to slow disease progression of Simian Immunodeficiency Virus (SIV) infections in macaques. To maximize their utility for HIV/AIDS research, the Southwest National Primate Research Center (SNPRC) has introduced targeted breeding strategies to reduce the prevalence of SIV-refractory MHC haplotypes and levels of admixture between Indian- and Chinese-origin macaques. ResultsWe characterized the MHC region in SNPRC macaques by targeted deep sequencing of 1,458 animals born over a ten-year period. Following the implementation of MHC management strategies, the prevalence of SIV-refractory MHC haplotypes reduced significantly while overall haplotype diversity was maintained. We investigated the impact of management strategies on admixture, population genetic structure, genetic diversity and inbreeding using whole exome sequencing of founding and colony-born animals (n=488). Admixture analysis of founders showed some Chinese ancestry, though population substructure more closely reflected primate research center source than geographical origin. The levels of Chinese ancestry declined significantly over time, though genetic diversity remains high. Finally, we performed genome-wide scans for genetic selection over time. We identify numerous genomic regions where allele frequencies have shifted significantly, supporting the presence of short-term adaptation within the colony. ConclusionsWe show that colony management strategies have been successful without reducing genetic diversity of the MHC or exonic regions. We also show that colony genetic substructure is related to animal colony source and that mergers and migrations have reduced inbreeding and increased overall genetic diversity.

genomics↗

bioRxiv2024

Mast cells promote pathology and susceptibility in tuberculosis

Tuberculosis (TB), caused by the bacterium Mycobacterium tuberculosis (Mtb), infects approximately one-fourth of the worlds population. We reported an increased accumulation of mast cells (MCs) in the lungs of macaques with active pulmonary TB (PTB), compared with those with latent TB infection (LTBI). MCs respond in vitro to Mtb exposure via degranulation and by inducing proinflammatory cytokines. In the current study, we demonstrate an increased production of chymase by MCs in granulomas of humans and macaques with PTB. Single-cell (sc) RNA sequencing analysis revealed distinct MC transcriptional programs between LTBI and PTB, with PTB associated MCs enriched in interferon gamma, oxidative phosphorylation, and MYC signaling. In a mouse model, MC deficiency led to improved control of Mtb infection that coincided with reduced accumulation of lung myeloid cells and diminished lung inflammation at chronic stages of infection. Airway transfer of MCs into wild-type Mtb infected mice showed increased neutrophils, decreased recruited macrophages, and elevated Mtb dissemination to the spleen. Together, these findings highlight MCs as active drivers of TB pathogenesis and potential targets for host-directed therapies for TB.

immunology↗

bioRxiv2023

Imaging Architecture of Granulomas Induced by Mycobacterium tuberculosis Infections with Single-Molecule FISH

Granulomas are an important hallmark of Mycobacterium tuberculosis (Mtb) infection. They are organized and dynamic structures created by an assembly of immune cells around the sites of infection in the lungs to locally restrict the bacterial growth and the hosts inflammatory responses. The cellular architecture of granulomas is traditionally studied by immunofluorescence labeling of phenotypic surface markers. However, very few antibodies are available for model animals used in tuberculosis research, such as non-human primates and rabbits; secreted immunological markers such as cytokines cannot be imaged in situ using antibodies; and traditional phenotypic surface markers do not provide sufficient resolution for the detection of many subtypes and differentiation states of immune cells. Using single-molecule fluorescent in situ hybridization (smFISH) and its derivatives, amplified smFISH (ampFISH) and iterative smFISH, we developed a platform for imaging mRNAs encoding immune markers in rabbit and macaque tuberculosis granulomas. Multiplexed imaging for several mRNA and protein markers was followed by quantitative measurement of expression of these markers in single cells in situ. A quantitative analysis of combinatorial expressions of these markers allowed us to classify the cells into several subtypes and chart their distributions within granulomas. For one mRNA target, HIF-1, we were able to image its mRNA and protein in the same cells, demonstrating the specificity of probes. This method paves the way for defining granular differentiation states and cell subtypes from transcriptomic data, identifying key mRNA markers for these cell subtypes, and then locating the cells in the spatial context of granulomas.

immunology↗

bioRxiv2021

In vivo serial passaging of human-simian immunodeficiency virus clones identifies viral characteristics that enhance persistent viral replication

We previously reported that a human immunodeficiency virus type 1 with a simian immunodeficiency virus vif substitution (HSIV-vifNL4-3) could replicate in pigtailed macaques (PTMs), demonstrating that Vif is a species-specific tropism factor of primate lentiviruses. However, infections did not result in high peak viremia or setpoint plasma viral loads, as observed during SIV infection of PTMs. Here, we characterized variants isolated from one of the original infected animals with CD4 depletion after nearly four years of infection to identify determinants of increased replication fitness. In our studies, we found that the HSIV-vif clones did not express the HIV-1 Vpr protein due to interference from the vpx open reading frame in singly spliced vpr mRNA. To examine whether these viral genes contribute to persistent viral replication, we generated infectious HSIV-vif clones expressing either the HIV-1 Vpr or SIV Vpx protein. And then to determine viral fitness determinants of HSIV-vif, we conducted three rounds of serial in vivo passaging in PTMs, starting with an initial inoculum containing a mixture of CXCR4-tropic (Vpr- HSIV-vifNL4-3 isolated at 196 (C/196) and 200 (C/200) weeks post-infection from a PTM with depressed CD4 counts) and CCR5-tropic HSIV (Vpr+ HSIV-vif derivatives based NL-AD8 and Bru-Yu2 and a Vpx expressing HSIV-vifYu2). Interestingly, all infected PTMs showed peak plasma viremia close to or above 105 copies/ml and persistent viral replication for more than 20 weeks. The passage 3 PTM showed peak viral loads greater than 105 viral RNA copies/ml. Infectious molecular clones (IMCs) recovered from the passage 3 PTM (HSIV-P3 IMCs) included mutations required for HIV-1 Vpr expression and those mutations encoded by the CXCR4-tropic HSIV-vifNL4-3 isolates C/196 and C/200. The data indicate that the biological isolates selected during long-term infection acquired HIV-1 Vpr expression to enhance their replication fitness in PTMs. Further passaging of HSIV-P3 IMCs in vivo may generate pathogenic variants with higher replication capacity, which will be a valuable resource as challenge virus in vaccine and cure studies.

microbiology↗

bioRxiv2021

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.

immunology↗

bioRxiv2021

Lung epithelial signaling mediates early vaccine-induced CD4+ T cell activation and Mtb control

Tuberculosis (TB) is one of the leading causes of death due to a single infectious agent. The development of a TB vaccine that induces durable and effective immunity to Mycobacterium tuberculosis (Mtb) infection is urgently needed. Early and superior Mtb control can be induced in M. bovis Bacillus Calmette-Guerin (BCG) vaccinated hosts when the innate immune response is targeted to generate effective vaccine-induced immunity. In the present study, we show that innate activation of DCs is critical for mucosal localization of clonally activated vaccine-induced CD4+ T cells in the lung, and superior early Mtb control. In addition, our study reveals that Th1/Th17 cytokine axis play an important role in superior vaccine induced immunity. Our studies also show that activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF{kappa}{beta}) pathway in lung epithelial cells is critical for the mucosal localization of activated vaccine-induced CD4+ T cells for rapid Mtb control. Thus, our study provides novel insights into the immune mechanisms that can overcome TB vaccine bottlenecks and provide early rapid Mtb control. ImportanceTuberculosis is a leading cause of death due to single infectious agent accounting 1.4 million deaths each year. The only licensed vaccine BCG is not effective due to variable efficacy. In our study, we determined the early immune events necessary for achieving complete protection in BCG vaccinated host. Our study reveals that innate activation of DCs can mediate superior and early Mtb control in BCG vaccinated host through lung epithelial cell signaling and localization of clonal activated, Mtb antigen specific, cytokine producing CD4+ T cells within the lung parenchyma and airways. Thus, our study provides novel insights into the immune mechanisms that can overcome TB vaccine bottlenecks and provide early rapid Mtb control.

immunology↗

bioRxiv2021

Robust IgM responses following vaccination are associated 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). However, recent work has shown that compared to standard intradermal Bacille Calmette-Guerin (BCG) vaccination, intravenous (IV) BCG vaccination provides nearly complete protection against TB in rhesus macaques. While studies have focused on cellular immunity in this setting, the antibody response elicited by IV BCG vaccination remains incompletely defined. Using an agnostic antibody profiling approach, here we show that IV BCG drives superior antibody responses in the plasma and the bronchoalveolar lavage fluid (BAL). While IV BCG immunization resulted in the expansion of a robust IgM, IgG, IgA, Fc-receptor binding antibodies, and antibody effector functions in the BAL, IgM titers were among the strongest markers of reduced bacterial burden in the plasma and BAL of BCG immunized animals. Moreover, IgM immunity was also enriched among animals receiving protective vaccination with an attenuated Mtb strain. Finally, a LAM-specific IgM monoclonal antibody reduced Mtb survival in vitro. Collectively, these data highlight the potential importance of IgM responses as a marker and as a functional mediator of protection against TB.

immunology↗

bioRxiv2021

Particle Dynamics and Bioaerosol Viability of Aerosolized BCG Vaccine Using Jet and Vibrating Mesh Clinical Nebulizers

BackgroundBacillus Calmette-Guerin (BCG) is a vaccine used to protect against tuberculosis primarily in infants to stop early infection in areas of the world where the disease is endemic. Normally administered as a percutaneous injection, BCG is a live, significantly attenuated bacteria that is now being investigated for its potential within an inhalable vaccine formulation. This work investigates the feasibility and performance of four jet and ultrasonic nebulizers aerosolizing BCG and the resulting particle characteristics and residual viability of the bacteria post-aerosolization. MethodsA jet nebulizer (Collison) outfitted either with a 3- or 6-jet head, was compared to two clinical nebulizers, the vibrating mesh Omron MicroAir and Aerogen Solo devices. Particle characteristics, including aerodynamic particle sizing, was performed on all devices within a common aerosol chamber configuration and comparable BCG innocula concentrations. Integrated aerosol samples were collected for each generator and assayed for bacterial viability using conventional microbiological technique. Results and ConclusionsA batch lot of BCG (Danish) was grown to titer and used in all generator assessments. Aerosol particles within the respirable range were generated from all nebulizers at four different concentrations of BCG. The jet nebulizers produced a uniformly smaller particle size than the ultrasonic devices, although particle concentrations by mass were similar across all devices tested with the exception of the Aerogen Solo, which resulted in a very low concentration of BCG aerosols. The resulting measured viable BCG aerosol concentration fraction produced by each device approximated one another; however, a measurable decrease of efficiency and overall viability reduction in the jet nebulizer was observed in higher BCG inoculum starting concentrations, whereas the vibrating mesh nebulizer returned a remarkably stable viable aerosol fraction irrespective of inoculum concentration.

biophysics↗

bioRxiv2020

IFN signaling and neutrophil degranulation transcriptional signatures are induced during SARS-CoV-2 infection

The novel virus SARS-CoV-2 has infected more than 14 million people worldwide resulting in the Coronavirus disease 2019 (COVID-19). Limited information on the underlying immune mechanisms that drive disease or protection during COVID-19 severely hamper development of therapeutics and vaccines. Thus, the establishment of relevant animal models that mimic the pathobiology of the disease is urgent. Rhesus macaques infected with SARS-CoV-2 exhibit disease pathobiology similar to human COVID-19, thus serving as a relevant animal model. In the current study, we have characterized the transcriptional signatures induced in the lungs of juvenile and old rhesus macaques following SARS-CoV-2 infection. We show that genes associated with Interferon (IFN) signaling, neutrophil degranulation and innate immune pathways are significantly induced in macaque infected lungs, while pathways associated with collagen formation are downregulated. In COVID-19, increasing age is a significant risk factor for poor prognosis and increased mortality. We demonstrate that Type I IFN and Notch signaling pathways are significantly upregulated in lungs of juvenile infected macaques when compared with old infected macaques. These results are corroborated with increased peripheral neutrophil counts and neutrophil lymphocyte ratio in older individuals with COVID-19 disease. In contrast, pathways involving VEGF are downregulated in lungs of old infected macaques. Using samples from humans with SARS-CoV-2 infection and COVID-19, we validate a subset of our findings. Finally, neutrophil degranulation, innate immune system and IFN gamma signaling pathways are upregulated in both tuberculosis and COVID-19, two pulmonary diseases where neutrophils are associated with increased severity. Together, our transcriptomic studies have delineated disease pathways to improve our understanding of the immunopathogenesis of COVID-19 to facilitate the design of new therapeutics for COVID-19.

immunology↗

bioRxiv2020

Lethality of SARS-CoV-2 infection in K18 human angiotensin converting enzyme 2 transgenic mice

ABSTRACTVaccine and antiviral development against SARS-CoV-2 infection or COVID-19 disease currently lacks a validated small animal model. Here, we show that transgenic mice expressing human angiotensin converting enzyme 2 (hACE2) by the human cytokeratin 18 promoter (K18 hACE2) represent a susceptible rodent model. K18 hACE2-transgenic mice succumbed to SARS-CoV-2 infection by day 6, with virus detected in lung airway epithelium and brain. K18 ACE2-transgenic mice produced a modest TH1/2/17 cytokine storm in the lung and spleen that peaked by day 2, and an extended chemokine storm that was detected in both lungs and brain. This chemokine storm was also detected in the brain at day 4. K18 hACE2-transgenic mice are, therefore, highly susceptible to SARS-CoV-2 infection and represent a suitable animal model for the study of viral pathogenesis, and for identification and characterization of vaccines (prophylactic) and antivirals (therapeutics) for SARS-CoV-2 infection and associated severe COVID-19 disease.

microbiology↗

bioRxiv2020

Control of Mycobacterium tuberculosis Infection in Lungs is Associated with Recruitment of Antigen-Specific Th1 and Th17 cells Co-expressing CXCR3 and CCR6.

Mycobacterium tuberculosis (Mtb)-specific T cell responses associated with immune control during asymptomatic latent tuberculosis infection (LTBI) remain poorly understood. Using a non-human primate (NHP) aerosol model, we studied the kinetics, phenotypes and functions of Mtb antigen-specific T cells in peripheral and lung compartments of Mtb-infected asymptomatic rhesus macaques by longitudinally sampling blood and bronchoalveolar lavage (BAL), for up to 24 weeks post-infection. We found significantly higher frequencies of Mtb-specific effector and memory CD4 and CD8 T cells producing IFN-{gamma} in the airways compared to peripheral blood, which were maintained throughout the study period. Moreover, Mtb-specific IL-17+ and IL-17/IFN-{gamma} double-positive T cells were present in the airways but were largely absent in the periphery, suggesting that balanced mucosal Th1/Th17 responses are associated with LTBI. The majority of Mtb-specific CD4 T cells that homed to the airways expressed the chemokine receptor CXCR3 and co-expressed CCR6. Notably, CXCR3+CD4+ cells were found in granulomatous and non-granulomatous regions of the lung and inversely correlated with Mtb burden. Our findings provide novel insights into antigen-specific T cell responses associated with asymptomatic Mtb infection that are relevant for developing better strategies to control TB.

immunology↗

bioRxiv2020

ClgR contributes to pulmonary pathology but not bacterial growth in Mycobacterium tuberculosis infection

BackgroundThe Clp proteases regulator, ClgR, is encoded in the Mycobacterium tuberculosis (Mtb) genome by Rv2745c gene (clgR). ClgR is required to clear damaged proteins, thereby preventing their accumulation in the cell. It also controls the availability of key enzymes or regulators via conditional degradation mechanism of proteolytic activity in Mtb [1,2]. MethodsIt has been previously reported that Mtb clgR gene is induced in a sigma factor SigH-dependent manner and a deletion mutant of clgR is susceptible to growth in a hypoxic environment. Whether hypoxia is indeed a restriction factor and ClgR is required for Mtb growth in that environment remains unelucidated. We began to address this hypothesis in the C57/BL6 mouse model of TB where Mtb infected lungs do not form granuloma and the lung environment is considerably non-hypoxic. ResultsOur results demonstrate that despite not having a deficit in growth in either murine lungs or primary macrophages, in comparison to wild type, the{Delta} clgR mutant failed to induce pulmonary pathology. ConclusionWe propose that ClgR is required for the pathogenesis of Mtb.

microbiology↗

bioRxiv2019

Tuberculosis-associated IFN-I induces Siglec-1 on tunneling nanotubes and favors HIV-1 spread in macrophages

While tuberculosis (TB) is a risk factor in HIV-1-infected individuals, the mechanisms by which Mycobacterium tuberculosis worsens HIV-1 pathogenesis remain poorly understood. Recently, we showed that HIV-1 infection and spread are exacerbated in macrophages exposed to TB-associated microenvironments due to tunneling nanotube (TNT) formation. To identify molecular factors associated with TNT function, we performed a transcriptomic analysis in these macrophages, and revealed the up-regulation of the lectin receptor Siglec-1. We demonstrate Siglec-1 expression depends on TB-mediated production of type I interferon. In co-infected non-human primates, Siglec-1 is highly expressed by alveolar macrophages, whose abundance correlates with pathology and activation of the type I interferon/STAT1 pathway. Intriguingly, Siglec-1 expression localizes exclusively on microtubule-containing TNT that are long and carry HIV-1 cargo. Siglec-1 depletion in macrophages decreases TNT length, diminishes HIV-1 capture and cell-to-cell transfer, and abrogates TB-driven exacerbation of HIV-1 infection. Altogether, we uncover a deleterious role for Siglec-1 in TB-HIV-1 co-infection, and its localization on TNT opens new avenues to understand cell-to-cell viral spread.

cell biology↗