A measles-vectored vaccine candidate expressing prefusion-stabilized SARS-CoV-2 spike protein brought to phase I/II clinical trials: protection of African green monkeys from COVID-19 disease.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged at the end of 2019 and is responsible for the largest human pandemic in 100 years. Thirty-four vaccines are currently approved for use worldwide, and approximately 67% of the world population has received a complete primary series of one, yet countries are dealing with new waves of infections, variant viruses continue to emerge, and breakthrough infections are frequent secondary to waning immunity. Here, we evaluate a measles virus (MV)-vectored vaccine expressing a stabilized prefusion SARS-CoV-2 spike (S) protein (MV-ATU3-S2PΔF2A; V591) with demonstrated immunogenicity in mouse models (see companion article [J. Brunet, Z. Choucha, M. Gransagne, H. Tabbal, M.-W. Ku et al., J Virol 98:e01693-23, 2024, https://doi.org/10.1128/jvi.01693-23]) in an established African green monkey model of disease. Animals were vaccinated with V591 or the control vaccine (an equivalent MV-vectored vaccine with an irrelevant antigen) intramuscularly using a prime/boost schedule, followed by challenge with an early pandemic isolate of SARS-CoV-2 at 56 days post-vaccination. Pre-challenge, only V591-vaccinated animals developed S-specific antibodies that had virus-neutralizing activity as well as S-specific T cells. Following the challenge, V591-vaccinated animals had lower infectious virus and viral (v) RNA loads in mucosal secretions and stopped shedding virus in these secretions earlier. vRNA loads were lower in these animals in respiratory and gastrointestinal tract tissues at necropsy. This correlated with a lower disease burden in the lungs as quantified by PET/CT at early and late time points post-challenge and by pathological analysis at necropsy.IMPORTANCESevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the largest human pandemic in 100 years. Even though vaccines are currently available, countries are dealing with new waves of infections, variant viruses continue to emerge, breakthrough infections are frequent, and vaccine hesitancy persists. This study uses a safe and effective measles vaccine as a platform for vaccination against SARS-CoV-2. The candidate vaccine was used to vaccinate African green monkeys (AGMs). All vaccinated AGMs developed robust antigen-specific immune responses. After challenge, these AGMs produced less virus in mucosal secretions, for a shorter period, and had a reduced disease burden in the lungs compared to control animals. At necropsy, lower levels of viral RNA were detected in tissue samples from vaccinated animals, and the lungs of these animals lacked the histologic hallmarks of SARS-CoV-2 disease observed exclusively in the control AGMs.

Antibiotic treatment modestly reduces protection against Mycobacterium tuberculosis reinfection in macaques.

Concomitant immunity is generally defined as an ongoing infection providing protection against reinfection . Its role in prevention of tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) is supported by epidemiological evidence in humans as well as experimental evidence in mice and non-human primates (NHPs). Whether the presence of live Mtb, rather than simply persistent antigen, is necessary for concomitant immunity in TB is still unclear. Here, we investigated whether live Mtb plays a measurable role in control of secondary Mtb infection. Using cynomolgus macaques, molecularly barcoded Mtb libraries, positron emission tomography-computed tomography (PET CT) imaging, flow cytometry, and cytokine profiling, we evaluated the effect of antibiotic treatment after primary infection on immunological response and bacterial establishment, dissemination, and burden post-secondary infection. Our data provide evidence that, in this experimental model, treatment with antibiotics after primary infection reduced inflammation in the lung but was not associated with a significant change in bacterial establishment, dissemination, or burden in the lung or lymph nodes. Nonetheless, treatment of the prior infection with antibiotics did result in a modest reduction in protection against reinfection: none of the seven antibiotic-treated animals demonstrated sterilizing immunity against reinfection, while four of the seven non-treated macaques were completely protected against reinfection. These findings support that antibiotic-treated animals were still able to restrict bacterial establishment and dissemination after rechallenge compared to naïve macaques, but not to the full extent of non-antibiotic-treated macaques.

Antibiotic treatment modestly reduces protection against Mycobacterium tuberculosis reinfection in macaques.

Concomitant immunity is generally defined as an ongoing infection providing protection against reinfection1. Its role in prevention of tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) is supported by epidemiological evidence in humans as well as experimental evidence in mice and non-human primates (NHPs). Whether the presence of live Mtb, rather than simply persistent antigen, is necessary for concomitant immunity in TB is still unclear. Here, we investigated whether live Mtb plays a measurable role in control of secondary Mtb infection. Using cynomolgus macaques, molecularly barcoded Mtb libraries, PET-CT imaging, flow cytometry and cytokine profiling we evaluated the effect of antibiotic treatment after primary infection on immunological response and bacterial establishment, dissemination, and burden post-secondary infection. Our data provide evidence that, in this experimental model, treatment with antibiotics after primary infection reduced inflammation in the lung but was not associated with a significant change in bacterial establishment, dissemination or burden in the lung or lymph nodes. Nonetheless, treatment of the prior infection with antibiotics did result in a modest reduction in protection against reinfection: none of the 7 antibiotic treated animals demonstrated sterilizing immunity against reinfection while 4 of the 7 non-treated macaques were completely protected against reinfection. These findings support that antibiotic-treated animals were still able to restrict bacterial establishment and dissemination after rechallenge compared to naïve macaques, but not to the full extent of non-antibiotic treated macaques.

A "suicide" BCG strain provides enhanced immunogenicity and robust protection against Mycobacterium tuberculosis in macaques.

Intravenous (IV) BCG delivery provides robust protection against Mycobacterium tuberculosis (Mtb) in macaques but poses safety challenges. Here, we constructed two BCG strains (BCG-TetON-DL and BCG-TetOFF-DL) in which tetracyclines regulate two phage lysin operons. Once the lysins are expressed, these strains are cleared in immunocompetent and immunocompromised mice, yet induced similar immune responses and provided similar protection against Mtb challenge as wild type BCG. Lysin induction resulted in release of intracellular BCG antigens and enhanced cytokine production by macrophages. In macaques, cessation of doxycycline administration resulted in rapid elimination of BCG-TetOFF-DL. However, IV BCG-TetOFF-DL induced increased pulmonary CD4 T cell responses compared to WT BCG and provided robust protection against Mtb challenge, with sterilizing immunity in 6 of 8 macaques, compared to 2 of 8 macaques immunized with WT BCG. Thus, a "suicide" BCG strain provides an additional measure of safety when delivered intravenously and robust protection against Mtb infection.

Development of an Engineered Mycobacterium tuberculosis Strain for a Safe and Effective Tuberculosis Human Challenge Model.

Human challenge experiments could greatly accelerate the development of a tuberculosis (TB) vaccine. Human challenge for tuberculosis requires a strain that can both replicate in the host and be reliably cleared. To accomplish this, we designed Mycobacterium tuberculosis (Mtb) strains featuring up to three orthogonal kill switches, tightly regulated by exogenous tetracyclines and trimethoprim. The resultant strains displayed immunogenicity and antibiotic susceptibility similar to wild-type Mtb under permissive conditions. In the absence of supplementary exogenous compounds, the strains were rapidly killed in axenic culture, mice and nonhuman primates. Notably, the strain that contained three kill switches had an escape rate of less than 10 -10 per genome per generation and displayed no relapse in a SCID mouse model. Collectively, these findings suggest that this engineered Mtb strain could be a safe and effective candidate for a human challenge model.

CD8+ lymphocytes are critical for early control of tuberculosis in macaques.

The functional role of CD8+ lymphocytes in tuberculosis remains poorly understood. We depleted innate and/or adaptive CD8+ lymphocytes in macaques and showed that loss of all CD8α+ cells (using anti-CD8α antibody) significantly impaired early control of Mycobacterium tuberculosis (Mtb) infection, leading to increased granulomas, lung inflammation, and bacterial burden. Analysis of barcoded Mtb from infected macaques demonstrated that depletion of all CD8+ lymphocytes allowed increased establishment of Mtb in lungs and dissemination within lungs and to lymph nodes, while depletion of only adaptive CD8+ T cells (with anti-CD8ß antibody) worsened bacterial control in lymph nodes. Flow cytometry and single-cell RNA sequencing revealed polyfunctional cytotoxic CD8+ lymphocytes in control granulomas, while CD8-depleted animals were unexpectedly enriched in CD4 and γδ T cells adopting incomplete cytotoxic signatures. Ligand-receptor analyses identified IL-15 signaling in granulomas as a driver of cytotoxic T cells. These data support that CD8+ lymphocytes are required for early protection against Mtb and suggest polyfunctional cytotoxic responses as a vaccine target.

Conference report: WHO meeting summary on mRNA-based tuberculosis vaccine development.

Tuberculosis (TB) is a global health emergency. Across the globe, approximately 2 billion people are currently infected with Mycobacterium tuberculosis (Mtb), and of those, 5-10% may progress to become ill and potentially transmit the bacterium. In 2021, nearly 10.6 million people developed TB disease and 1.6 million died. There is an urgent need for accelerated development of new TB-focused interventions, in particular, improved TB vaccines. However, progress in developing highly effective TB vaccines has been slow and is chronically under-resourced. The mRNA vaccine platform may offer an opportunity to accelerate development of new TB vaccines. In April 2023, the World Health Organization convened global experts to discuss the feasibility and potential value of mRNA-based vaccines for TB. Here we report on meeting deliberations related to the current TB vaccine pipeline and potential novel antigens, the status of efforts to identify correlates of protection, potential clinical development strategies and considerations for community acceptance of new TB vaccines based on this relatively new platform. The role of industry collaborations, ethics, social science, and responsibility to the global community regarding transparency and manufacturing capacity building were discussed through expert presentations and panel sessions. The overall conclusion of the meeting is that mRNA-based vaccines constitute a potentially powerful new tool for reducing the global burden of TB.

Intravenous Bacille Calmette-Guérin vaccination protects simian immunodeficiency virus-infected macaques from tuberculosis.

Tuberculosis, caused by Mycobacterium tuberculosis (Mtb), is the most common cause of death in people living with human immunodeficiency virus (HIV). Intra-dermal Bacille Calmette-Guérin (BCG) delivery is the only licensed vaccine against tuberculosis; however, it offers little protection from pulmonary tuberculosis in adults and is contraindicated in people living with HIV. Intravenous BCG confers protection against Mtb infection in rhesus macaques; we hypothesized that it might prevent tuberculosis in simian immunodeficiency virus (SIV)-infected macaques, a model for HIV infection. Here intravenous BCG-elicited robust airway T cell influx and elevated plasma and airway antibody titres in both SIV-infected and naive animals. Following Mtb challenge, all 7 vaccinated SIV-naive and 9 out of 12 vaccinated SIV-infected animals were protected, without any culturable bacteria detected from tissues. Peripheral blood mononuclear cell responses post-challenge indicated early clearance of Mtb in vaccinated animals, regardless of SIV infection. These data support that intravenous BCG is immunogenic and efficacious in SIV-infected animals.

Humoral correlates of protection against Mycobacterium tuberculosis following intravenous Bacille Calmette-Guérin vaccination in rhesus macaques.

Altering the route of Bacille Calmette-Guérin (BCG) immunization from low-dose intradermal vaccination to high-dose intravenous (IV) vaccination resulted in a high level of protection against Mycobacterium tuberculosis ( Mtb ) infection, providing an opportunity to uncover immune correlates and mechanisms of protection. In addition to strong T cell immunity, IV BCG vaccination was associated with a robust expansion of humoral immune responses that tracked with bacterial control. However, given the near complete protection afforded by high-dose IV BCG immunization, a precise correlate of immune protection was difficult to define. Here we leveraged plasma and bronchoalveolar lavage fluid (BAL) from a cohort of rhesus macaques that received decreasing doses of IV BCG and aimed to define the correlates of immunity across macaques that experienced immune protection or breakthrough infection following Mtb challenge. We show an IV BCG dose-dependent induction of mycobacterial-specific humoral immune responses, both in the plasma and in the airways. Moreover, antibody responses at peak immunogenicity significantly predicted bacterial control following challenge. Multivariate analyses revealed antibody-mediated complement and NK cell activating humoral networks as key functional signatures associated with protective immunity. Collectively, this work extends our understanding of humoral biomarkers and potential mechanisms of IV BCG mediated protection against Mtb .

Protective intravenous BCG vaccination induces enhanced immune signaling in the airways.

Intradermal (ID) Bacillus Calmette-Guérin (BCG) is the most widely administered vaccine in the world. However, ID-BCG fails to achieve the level of protection needed in adults to alter the course of the tuberculosis epidemic. Recent studies in non-human primates have demonstrated high levels of protection against Mycobacterium tuberculosis ( Mtb ) following intravenous (IV) administration of BCG. However, the protective immune features that emerge following IV BCG vaccination remain incompletely defined. Here we used single-cell RNA-sequencing (scRNAseq) to transcriptionally profile 157,114 unstimulated and purified protein derivative (PPD)-stimulated bronchoalveolar lavage (BAL) cells from 29 rhesus macaques immunized with BCG across routes of administration and doses to uncover cell composition-, gene expression-, and biological network-level signatures associated with IV BCG-mediated protection. Our analyses revealed that high-dose IV BCG drove an influx of polyfunctional T cells and macrophages into the airways. These macrophages exhibited a basal activation phenotype even in the absence of PPD-stimulation, defined in part by IFN and TNF-α signaling up to 6 months following BCG immunization. Furthermore, intercellular immune signaling pathways between key myeloid and T cell subsets were enhanced following PPD-stimulation in high-dose IV BCG-vaccinated macaques. High-dose IV BCG also engendered quantitatively and qualitatively stronger transcriptional responses to PPD-stimulation, with a robust Th1-Th17 transcriptional phenotype in T cells, and augmented transcriptional signatures of reactive oxygen species production, hypoxia, and IFN-γ response within alveolar macrophages. Collectively, this work supports that IV BCG immunization creates a unique cellular ecosystem in the airways, which primes and enables local myeloid cells to effectively clear Mtb upon challenge.

High resolution imaging and five-year tuberculosis contact outcomes.

Background: The evolution of tuberculosis (TB) disease during the clinical latency period remains incompletely understood. Methods: 250 HIV-uninfected, adult household contacts of rifampicin-resistant TB with a negative symptom screen underwent baseline 18F-Fluorodeoxyglucose positron emission and computed tomography (PET/CT), repeated in 112 after 5-15 months. Following South African and WHO guidelines, participants did not receive preventive therapy. All participants had intensive baseline screening with spontaneous, followed by induced, sputum sampling and were then observed for an average of 4.7 years for culture-positive disease. Baseline PET/CT abnormalities were evaluated in relation to culture-positive disease. Results: At baseline, 59 (23.6%) participants had lung PET/CT findings consistent with TB of which 29 (11.6%) were defined as Subclinical TB, and 30 (12%) Subclinical TB-inactive. A further 83 (33.2%) had other lung parenchymal abnormalities and 108 (43.2%) had normal lungs. Over 1107-person years of follow-up 14 cases of culture-positive TB were diagnosed. Six cases were detected by intensive baseline screening, all would have been missed by the South African symptom-based screening strategy and only one detected by a WHO-recommended chest X-Ray screening strategy. Those with baseline Subclinical TB lesions on PET/CT were significantly more likely to be diagnosed with culture-positive TB over the study period, compared to those with normal lung parenchyma (10/29 [34.5%] vs 2/108 [1.9%], Hazard Ratio 22.37 [4.89-102.47, p<0.001]). Conclusions: These findings challenge the latent/active TB paradigm demonstrating that subclinical disease exists up to 4 years prior to microbiological detection and/or symptom onset. There are important implications for screening and management of TB.

Airway T cells are a correlate of i.v. Bacille Calmette-Guerin-mediated protection against tuberculosis in rhesus macaques.

Bacille Calmette-Guerin (BCG), the only approved Mycobacterium tuberculosis (Mtb) vaccine, provides limited durable protection when administered intradermally. However, recent work revealed that intravenous (i.v.) BCG administration yielded greater protection in macaques. Here, we perform a dose-ranging study of i.v. BCG vaccination in macaques to generate a range of immune responses and define correlates of protection. Seventeen of 34 macaques had no detectable infection after Mtb challenge. Multivariate analysis incorporating longitudinal cellular and humoral immune parameters uncovered an extensive and highly coordinated immune response from the bronchoalveolar lavage (BAL). A minimal signature predicting protection contained four BAL immune features, of which three remained significant after dose correction: frequency of CD4 T cells producing TNF with interferon γ (IFNγ), frequency of those producing TNF with IL-17, and the number of NK cells. Blood immune features were less predictive of protection. We conclude that CD4 T cell immunity and NK cells in the airway correlate with protection following i.v. BCG.

Optimizing tuberculosis treatment efficacy: Comparing the standard regimen with Moxifloxacin-containing regimens.

Tuberculosis (TB) continues to be one of the deadliest infectious diseases in the world, causing ~1.5 million deaths every year. The World Health Organization initiated an End TB Strategy that aims to reduce TB-related deaths in 2035 by 95%. Recent research goals have focused on discovering more effective and more patient-friendly antibiotic drug regimens to increase patient compliance and decrease emergence of resistant TB. Moxifloxacin is one promising antibiotic that may improve the current standard regimen by shortening treatment time. Clinical trials and in vivo mouse studies suggest that regimens containing moxifloxacin have better bactericidal activity. However, testing every possible combination regimen with moxifloxacin either in vivo or clinically is not feasible due to experimental and clinical limitations. To identify better regimens more systematically, we simulated pharmacokinetics/pharmacodynamics of various regimens (with and without moxifloxacin) to evaluate efficacies, and then compared our predictions to both clinical trials and nonhuman primate studies performed herein. We used GranSim, our well-established hybrid agent-based model that simulates granuloma formation and antibiotic treatment, for this task. In addition, we established a multiple-objective optimization pipeline using GranSim to discover optimized regimens based on treatment objectives of interest, i.e., minimizing total drug dosage and lowering time needed to sterilize granulomas. Our approach can efficiently test many regimens and successfully identify optimal regimens to inform pre-clinical studies or clinical trials and ultimately accelerate the TB regimen discovery process.

Blood transcriptional correlates of BCG-induced protection against tuberculosis in rhesus macaques.

Blood-based correlates of vaccine-induced protection against tuberculosis (TB) are urgently needed. Here, we analyze the blood transcriptome of rhesus macaques immunized with varying doses of intravenous (i.v.) BCG followed by Mycobacterium tuberculosis (Mtb) challenge. We use high-dose i.v. BCG recipients for "discovery" and validate our findings in low-dose recipients and in an independent cohort of macaques receiving BCG via different routes. We identify seven vaccine-induced gene modules, including an innate module (module 1) enriched for type 1 interferon and RIG-I-like receptor signaling pathways. Module 1 on day 2 post-vaccination highly correlates with lung antigen-responsive CD4 T cells at week 8 and with Mtb and granuloma burden following challenge. Parsimonious signatures within module 1 at day 2 post-vaccination predict protection following challenge with area under the receiver operating characteristic curve (AUROC) ≥0.91. Together, these results indicate that the early innate transcriptional response to i.v. BCG in peripheral blood may provide a robust correlate of protection against TB.

Distributable, Metabolic PET Reporting of Tuberculosis.

Tuberculosis remains a large global disease burden for which treatment regimens are protracted and monitoring of disease activity difficult. Existing detection methods rely almost exclusively on bacterial culture from sputum which limits sampling to organisms on the pulmonary surface. Advances in monitoring tuberculous lesions have utilized the common glucoside [18F]FDG, yet lack specificity to the causative pathogen Mycobacterium tuberculosis (Mtb) and so do not directly correlate with pathogen viability. Here we show that a close mimic that is also positron-emitting of the non-mammalian Mtb disaccharide trehalose - 2-[18F]fluoro-2-deoxytrehalose ([18F]FDT) - can act as a mechanism-based enzyme reporter in vivo. Use of [18F]FDT in the imaging of Mtb in diverse models of disease, including non-human primates, successfully co-opts Mtb-specific processing of trehalose to allow the specific imaging of TB-associated lesions and to monitor the effects of treatment. A pyrogen-free, direct enzyme-catalyzed process for its radiochemical synthesis allows the ready production of [18F]FDT from the most globally-abundant organic 18F-containing molecule, [18F]FDG. The full, pre-clinical validation of both production method and [18F]FDT now creates a new, bacterium-specific, clinical diagnostic candidate. We anticipate that this distributable technology to generate clinical-grade [18F]FDT directly from the widely-available clinical reagent [18F]FDG, without need for either bespoke radioisotope generation or specialist chemical methods and/or facilities, could now usher in global, democratized access to a TB-specific PET tracer.

Mycobacterium tuberculosis-Specific CD4 T Cells Expressing Transcription Factors T-Bet or RORγT Associate with Bacterial Control in Granulomas.

Despite the extensive research on CD4 T cells within the context of Mycobacterium tuberculosis (Mtb) infections, few studies have focused on identifying and investigating the profile of Mtb-specific T cells within lung granulomas. To facilitate the identification of Mtb-specific CD4 T cells, we identified immunodominant epitopes for two Mtb proteins, namely, Rv1196 and Rv0125, using a Mauritian cynomolgus macaque model of Mtb infection, thereby providing data for the synthesis of MHC class II tetramers. Using tetramers, we identified Mtb-specific cells within different immune compartments, postinfection. We found that granulomas were enriched sites for Mtb-specific cells and that tetramer+ cells had increased frequencies of the activation marker CD69 as well as the transcription factors T-bet and RORγT, compared to tetramer negative cells within the same sample. Our data revealed that while the frequency of Rv1196 tetramer+ cells was positively correlated with the granuloma bacterial burden, the frequency of RORγT or T-bet within tetramer+ cells was inversely correlated with the granuloma bacterial burden, thereby highlighting the importance of having activated, polarized, Mtb-specific cells for the control of Mtb in lung granulomas. IMPORTANCE Tuberculosis, caused by the bacterial pathogen Mycobacterium tuberculosis, kills 1.5 million people each year, despite the existence of effective drugs and a vaccine that is given to infants in most countries. Clearly, we need better vaccines against this disease. However, our understanding of the immune responses that are necessary to prevent tuberculosis is incomplete. This study seeks to understand the functions of T cells that are specific for M. tuberculosis at the site of the disease in the lungs. For this, we developed specialized tools called MHC class II tetramers to identify those T cells that can recognize M. tuberculosis and applied the tools to the study of this infection in nonhuman primate models that mimic human tuberculosis. We demonstrate that M. tuberculosis-specific T cells in lung lesions are associated with control of the bacteria only when those T cells are expressing certain functions, thereby highlighting the importance of combining the identification of specific T cells with functional analyses. Thus, we surmise that these functions of specific T cells are critical to the control of infection and should be considered as a part of the development of vaccines against tuberculosis.

Vaccination with intravenous BCG protects macaques with pre-existing SIV infection from tuberculosis.

Tuberculosis (TB) is the most common cause of death in people living with HIV. BCG delivered intradermally (ID) is the only licensed vaccine to prevent TB. However, it offers little protection from pulmonary TB in adults. Intravenous (IV) BCG, but not ID BCG, confers striking protection against Mycobacterium tuberculosis (Mtb) infection and disease in rhesus macaques. We investigated whether IV BCG could protect against TB in macaques with a pre-existing SIV infection. There was a robust influx of airway T cells following IV BCG in both SIV-infected and SIV-naïve animals, with elevated antibody titers in plasma and airways. Following Mtb challenge, all 7 SIV-naïve and 9 out of 12 SIV-infected vaccinated animals were completely protected, without any culturable bacilli in their tissues. PBMC responses post-challenge indicated early clearance of Mtb in vaccinated animals regardless of SIV infection. These data support that IV BCG is immunogenic and efficacious in SIV-infected animals.

Practical considerations for a TB controlled human infection model (TB-CHIM); the case for TB-CHIM in Africa, a systematic review of the literature and report of 2 workshop discussions in UK and Malawi.

Background: Tuberculosis (TB) remains a major challenge in many domains including diagnosis, pathogenesis, prevention, treatment, drug resistance and long-term protection of the public health by vaccination. A controlled human infection model (CHIM) could potentially facilitate breakthroughs in each of these domains but has so far been considered impossible owing to technical and safety concerns. Methods: A systematic review of mycobacterial human challenge studies was carried out to evaluate progress to date, best possible ways forward and challenges to be overcome. We searched MEDLINE (1946 to current) and CINAHL (1984 to current) databases; and Google Scholar to search citations in selected manuscripts. The final search was conducted 3 rd February 2022. Inclusion criteria: adults ≥18 years old; administration of live mycobacteria; and interventional trials or cohort studies with immune and/or microbiological endpoints. Exclusion criteria: animal studies; studies with no primary data; no administration of live mycobacteria; retrospective cohort studies; case-series; and case-reports. Relevant tools (Cochrane Collaboration for RCTs and Newcastle-Ottawa Scale for non-randomised studies) were used to assess risk of bias and present a narrative synthesis of our findings. Results: The search identified 1,388 titles for review; of these 90 were reviewed for inclusion; and 27 were included. Of these, 15 were randomised controlled trials and 12 were prospective cohort studies. We focussed on administration route, challenge agent and dose administered for data extraction. Overall, BCG studies including fluorescent BCG show the most immediate utility, and genetically modified Mycobacteria tuberculosis is the most tantalising prospect of discovery breakthrough. Conclusions: The TB-CHIM development group met in 2019 and 2022 to consider the results of the systematic review, to hear presentations from many of the senior authors whose work had been reviewed and to consider best ways forward. This paper reports both the systematic review and the deliberations. Registration: PROSPERO ( CRD42022302785; 21 January 2022).

Characterizing the Spectrum of Latent Mycobacterium tuberculosis in the Cynomolgus Macaque Model: Clinical, Immunologic, and Imaging Features of Evolution.

Mycobacterium tuberculosis infection outcomes have been described as active tuberculosis or latent infection but a spectrum of outcomes is now recognized. We used a nonhuman primate model, which recapitulates human infection, to characterize the clinical, microbiologic, and radiographic patterns associated with developing latent M. tuberculosis infection. Four patterns were identified. "Controllers" had normal erythrocyte sedimentation rate (ESR) without M. tuberculosis growth in bronchoalveolar lavage or gastric aspirate (BAL/GA). "Early subclinicals" showed transient ESR elevation and/or M. tuberculosis growth on BAL/GA for 60 days postinfection, "mid subclinicals" were positive for 90 days, and "late subclinicals" were positive intermittently, despite the absence of clinical disease. Variability was noted regarding granuloma formation, lung/lymph node metabolic activity, lung/lymph node bacterial burden, gross pathology, and extrapulmonary disease. Like human M. tuberculosis infection, this highlights the heterogeneity associated with the establishment of latent infection, underscoring the need to understand the clinical spectrum and risk factors associated with severe disease.