Krista E. van Meijgaarden

A significant proportion of individuals who are heavily exposed to infectious tuberculosis patients do not acquire Mycobacterium tuberculosis (Mtb) infection, as detected by an interferon gamma release assay (IGRA). We examined circulating metabolite profiles and metabolic genotypes in 199 heavily exposed IGRA-negative tuberculosis household contacts in Indonesia. Based on differentially abundant metabolites, activity of several pathways including arachidonic acid, arginine and proline, glutathione, and tryptophan metabolism, correlated with a negative IGRA at three months. SNPs near PRODH (involved in arginine and proline metabolism) were associated with circulating proline concentrations and persistently negative IGRA results, while SNPstudys near AFMID (involved in tryptophan metabolism) were associated with IGRA conversion. For further validation, plasma metabolomic profiles were correlated with mycobacterial growth inhibition by peripheral blood mononuclear cells from individuals in a low-incidence setting. Lower circulating concentrations of six metabolites, including leukotriene B4, proline, glycine, and tryptophan correlated with better growth control in non-exposed healthy individuals, as well as with stronger protection against IGRA-conversion in the Indonesian tuberculosis household contacts. Collectively, these data support the notion that circulating metabolites may impact innate host defense against Mtb infection, and that metabolic interventions may prevent tuberculosis infection and disease.

Tuberculosis (TB) is a curable infectious disease that requires prolonged treatment with multiple antibiotics. To better understand how the immune system contributes to the clearance of Mycobacterium tuberculosis (Mtb), in vitro assays are essential for monitoring functional immune changes during infection, therapy, and following vaccination. In this study, we investigated whether mycobacterial growth control in thirty patients with TB disease changes over the course of treatment. Comprehensive immune profiling of peripheral blood mononuclear cells (PBMCs), using a 30-color spectral flow cytometry panel, identified dynamic shifts in immune cell subsets related to functional activity. Notably, in addition to memory and effector T cells, a subset of naive B cells changed during treatment. Most sera contained antibodies binding to purified protein derivative (PPD) and Mtb-specific antigens ESAT-6/CFP-10, and enhanced phagocytic activity. A functional mycobacterial growth inhibition assay (MGIA) revealed growth control, which appeared to be heterogeneous but generally sustained throughout longitudinal follow-up. We conclude that T and B cell responses change in response to antibiotic treatment of TB disease, but that mycobacterial growth control capacity is a property of the individual, which is not influenced by disease activity or antibiotic treatment.

The human immune system employs both innate and adaptive mechanisms to control pathogens, with antibodies playing a pivotal role in immune memory and defense, in particular against viral infections. In tuberculosis, antibody titers have long been used to assess immune responses, but their presence alone fails to predict protective efficacy. Recent studies highlight that antibody functionality is critical for effective immune activity. Despite widespread detection of Mycobacterium tuberculosis (Mtb)-reactive antibodies in individuals with active disease, Mtb infection, and even in healthy controls, their potential to control Mtb growth is variable and only detected in a proportion of individuals. This perspective emphasizes the need for robust functional assessment of antibodies to better understand their role in mycobacterial control and inform vaccine development. Notably, antibodies binding to purified protein derivative of Mtb , a mixture of degraded antigens from Mtb cultures, are widespread but not universally functional, underscoring the importance of Fc characteristics and epitope specificity. Initial high-throughput screening using phagocytosis and direct mycobacterial binding assays is an active indicator of antibody function. By refining and combining existing assays, as recommended in this perspective, we can better characterize antibody contributions, particularly their immunomodulatory potential, toward improved control of Mtb . Albeit antibodies may not be essential in natural protection, functional antibodies induced by vaccination may be of added value and contribute to host protection.

A significant proportion of individuals who are heavily exposed to infectious tuberculosis patients do not acquire Mycobacterium tuberculosis (Mtb) infection, as detected by an interferon gamma release assay (IGRA). We examined circulating metabolite profiles and metabolic genotypes in 199 heavily exposed IGRA-negative tuberculosis household contacts in Indonesia. Based on differentially abundant metabolites, activity of several pathways including arachidonic acid, arginine and proline, glutathione, and tryptophan metabolism, correlated with a negative IGRA at three months. SNPs near PRODH (involved in arginine and proline metabolism) were associated with circulating proline concentrations and persistently negative IGRA results, while SNPs near AFMID (involved in tryptophan metabolism) were associated with IGRA conversion. For further validation, plasma metabolomic profiles were correlated with mycobacterial growth inhibition by peripheral blood mononuclear cells from individuals in a low-incidence setting. Lower circulating concentrations of six metabolites, including leukotriene B4, proline, glycine, and tryptophan correlated with better growth control in non-exposed healthy individuals, as well as with stronger protection against IGRA-conversion in the Indonesian tuberculosis household contacts. Collectively, these data support the notion that circulating metabolites impact innate host defense against Mtb infection, and that metabolic interventions may prevent tuberculosis infection and disease.

Background: Novel vaccines targeting the world’s deadliest pathogen Mycobacterium tuberculosis (Mtb) are urgently needed as the efficacy of the Bacillus Calmette–Guérin (BCG) vaccine in its current use is limited. HLA-E is a virtually monomorphic unconventional antigen presentation molecule, and HLA-E-restricted Mtb-specific CD8⁺ T cells can control intracellular Mtb growth, making HLA-E a promising vaccine target for Mtb. Methods: In this study, we evaluated the frequency and phenotype of HLA-E-restricted Mtb-specific CD4⁺/CD8⁺ T cells in the circulation and bronchoalveolar lavage fluid of two independent non-human primate (NHP) studies and from humans receiving BCG either intradermally or mucosally. Results: BCG vaccination followed by Mtb challenge in NHPs did not affect the frequency of circulating and local HLA-E–Mtb CD4⁺ and CD8⁺ T cells, and we saw the same in humans receiving BCG. HLA-E–Mtb T cell frequencies were significantly increased after Mtb challenge in unvaccinated NHPs, which was correlated with higher TB pathology. Conclusions: Together, HLA-E–Mtb-restricted T cells are minimally induced by BCG in humans and rhesus macaques (RMs) but can be elicited after Mtb infection in unvaccinated RMs. These results give new insights into targeting HLA-E as a potential immune mechanism against TB.

Bacillus Calmette-Guèrin - vaccination induces not only protection in infants and young children against severe forms of tuberculosis, but also against non-tuberculosis related all-cause mortality. To delineate different factors influencing mycobacterial growth control, here we first investigate the effects of BCG-vaccination in healthy Dutch adults. About a quarter of individuals already control BCG-growth prior to vaccination, whereas a quarter of the vaccinees acquires the capacity to control BCG upon vaccination. This leaves half of the population incapable to control BCG-growth. Single cell RNA sequencing identifies multiple processes associated with mycobacterial growth control. These data suggest (i) that already controllers employ different mechanisms to control BCG-growth than acquired controllers, and (ii) that half of the individuals fail to develop measurable growth control irrespective of BCG-vaccination. These results shed important new light on the variable immune responses to mycobacteria in humans and may impact on improved vaccination against tuberculosis and other diseases.

Bacillus Calmette-Guèrin (BCG) vaccination induces considerable protection in infants and young children against not only severe forms of tuberculosis (TB), but also all-cause mortality and heterologous infections. These effects are mediated through a combination of specific adaptive immune memory and sustained innate immune activation (also called trained immunity). To delineate factors influencing mycobacterial growth control, we investigated the effects of BCG vaccination in healthy Dutch adults. About a quarter of individuals already controlled BCG outgrowth prior to vaccination, whereas a quarter of vaccinees acquired the capacity to control 3 months upon BCG vaccination. This leaves a considerable percentage of the population (54% of total) incapable to control BCG outgrowth even after vaccination. Detailed characterization of biological samples, including single cell RNA sequencing (scRNAseq), identified multiple processes associated with mycobacterial growth control. Differential gene expression was abundant in all cell types of individuals with natural control, compared to those who did not control mycobacterial growth. In contrast, BCG-induced acquired control was mostly characterized by differential gene expression in monocytes. Control prior to BCG vaccination was associated with increased oxidative phosphorylation, whereas control acquired upon BCG vaccination was associated with glycolysis. Moreover, samples with natural control expressed genes linked to the type I IFN pathway, whereas samples with acquired control expressed genes associated with the Th1 pathway. These data suggest that natural control and control induced upon BCG vaccination employ different mechanisms to control BCG outgrowth, and that a substantial proportion of individuals (≈50%) fail to develop measurable growth control even following BCG vaccination.

Tuberculosis (TB) remains one of the deadliest infectious diseases worldwide, posing great social and economic burden to affected countries. Novel vaccine approaches are needed to increase protective immunity against the causative agent Mycobacterium tuberculosis (Mtb) and to reduce the development of active TB disease in latently infected individuals. Donor-unrestricted T cell responses represent such novel potential vaccine targets. HLA-E-restricted T cell responses have been shown to play an important role in protection against TB and other infections, and recent studies have demonstrated that these cells can be primed in vitro. However, the identification of novel pathogen-derived HLA-E binding peptides presented by infected target cells has been limited by the lack of accurate prediction algorithms for HLA-E binding. In this study, we developed an improved HLA-E binding peptide prediction algorithm and implemented it to identify (to our knowledge) novel Mtb-derived peptides with capacity to induce CD8+ T cell activation and that were recognized by specific HLA-E-restricted T cells in Mycobacterium-exposed humans. Altogether, we present a novel algorithm for the identification of pathogen- or self-derived HLA-E-presented peptides.