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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., (Copyright © 2022 by The American Association of Immunologists, Inc.)

Objectives: In this study, we aimed to characterize the SARS-CoV-2-specific T cell response detected by the QuantiFERON SARS-CoV-2 research use only assay in terms of accuracy and T cell subsets involved compared with a homemade interferon (IFN)-γ release assay (IGRA)., Methods: We evaluated T cell response by the standardized QuantiFERON SARS-CoV-2 tubes (antigen [Ag]1 and Ag2) and a homemade IGRA quantifying IFN-γ response to SARS-CoV-2 spike peptides (homemade-IGRA-SPIKE test). We evaluated the T cell subsets mediating the specific response using flow cytometry., Results: We prospectively enrolled 66 individuals: COVID-19 or post-COVID-19 subjects and NO-COVID-19-vaccinated subjects, including healthy donors and immunocompromised subjects. The standardized kit detected 62.1% (41/66) of T cell responders. Ag2 tube showed a higher IFN-γ quantitative and qualitative response. Ag1 tube response was mainly mediated by CD4 + T cells; Ag2 tube response was mediated by CD4 + and CD8 + T cells. The homemade-IGRA-SPIKE test detected a higher number of responders (52/66, 78.8%) than the QuantiFERON SARS-CoV-2 assay (P = 0.056). The response was found in both T cell subsets, although a higher magnitude and response rate was observed in the CD4 + T cell subset., Conclusion: The QuantiFERON SARS-CoV-2 response is mediated by CD4 + and CD8 + T cells. A lower number of responders is found compared with the homemade-IGRA-SPIKE test, likely because of the different peptide composition., Competing Interests: Conflict of Interest CT and CG received honoraria for speaking, manuscript writing, or educational events from Merck, Biogen, Roche, Novartis Sanofi, Celgene, and Almiral. EN participates on a data safety monitoring board or advisory board and receives fees for educational training from Gilead, Eli Lilly, GS, SOBI, and Roche. EN has a patent pending for raloxifene use in COVID-19 with Dompè Pharmaceutical. DG is a member of the advisory board of Biomerieux and Eli Lilly and received fees for educational training or consultancy from Almiral, Biogen, Cellgene, Diasorin, Janssen, Qiagen, and Quidel. All the other authors declare that the research was conducted without any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Platelets regulate human inflammatory responses that lead to disease. However, the role of platelets in tuberculosis (TB) pathogenesis is still unclear. Here, we show that patients with active TB have a high number of platelets in peripheral blood and a low number of lymphocytes leading to a high platelets to lymphocytes ratio (PL ratio). Moreover, the serum concentration of different mediators promoting platelet differentiation or associated with platelet activation is increased in active TB. Immunohistochemistry analysis shows that platelets localise around the lung granuloma lesions in close contact with T lymphocytes and macrophages. Transcriptomic analysis of caseous tissue of human pulmonary TB granulomas, followed by Gene Ontology analysis, shows that 53 platelet activation-associated genes are highly expressed compared to the normal lung tissue. In vitro activated platelets (or their supernatants) inhibit BCG-induced T- lymphocyte proliferation and IFN-γ production. Likewise, platelets inhibit the growth of intracellular macrophages of Mycobacterium (M.) tuberculosis. Soluble factors released by activated platelets mediate both immunological and M. tuberculosis replication activities. Furthermore, proteomic and neutralisation studies (by mAbs) identify TGF-β and PF4 as the factors responsible for inhibiting T-cell response and enhancing the mycobactericidal activity of macrophages, respectively. Altogether these results highlight the importance of platelets in TB pathogenesis., (© 2022 The Authors. European Journal of Immunology published by Wiley-VCH GmbH.)

Objectives: The interaction of COVID-19 and tuberculosis (TB) are still poor characterized. Here we evaluated the immune response specific for Micobacterium tuberculosis (Mtb) and SARS-CoV-2 using a whole-blood-based assay-platform in COVID-19 patients either with TB or latent TB infection (LTBI)., Methods: We evaluated IFN-γ level in plasma from whole-blood stimulated with Mtb antigens in the Quantiferon-Plus format or with peptides derived from SARS-CoV-2 spike protein, Wuhan-Hu-1 isolate (CD4-S)., Results: We consecutively enrolled 63 COVID-19, 10 TB-COVID-19 and 11 LTBI-COVID-19 patients. IFN-γ response to Mtb-antigens was significantly associated to TB status and therefore it was higher in TB-COVID-19 and LTBI-COVID-19 patients compared to COVID-19 patients (p ≤ 0.0007). Positive responses against CD4-S were found in 35/63 COVID-19 patients, 7/11 LTBI-COVID-19 and only 2/10 TB-COVID-19 patients. Interestingly, the responders in the TB-COVID-19 group were less compared to COVID-19 and LTBI-COVID-19 groups (p = 0.037 and 0.044, respectively). Moreover, TB-COVID-19 patients showed the lowest quantitative IFN-γ response to CD4-S compared to COVID-19-patients (p = 0.0336) and LTBI-COVID-19 patients (p = 0.0178)., Conclusions: Our data demonstrate that COVID-19 patients either TB or LTBI have a low ability to build an immune response to SARS-CoV-2 while retaining the ability to respond to Mtb-specific antigens., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Host resistance to Mycobacterium tuberculosis (Mtb) infection requires the activities of multiple leukocyte subsets, yet the roles of the different innate effector cells during tuberculosis are incompletely understood. Here we uncover an unexpected association between eosinophils and Mtb infection. In humans, eosinophils are decreased in the blood but enriched in resected human tuberculosis lung lesions and autopsy granulomas. An influx of eosinophils is also evident in infected zebrafish, mice, and nonhuman primate granulomas, where they are functionally activated and degranulate. Importantly, using complementary genetic models of eosinophil deficiency, we demonstrate that in mice, eosinophils are required for optimal pulmonary bacterial control and host survival after Mtb infection. Collectively, our findings uncover an unexpected recruitment of eosinophils to the infected lung tissue and a protective role for these cells in the control of Mtb infection in mice., Competing Interests: Disclosures:   D.M. Lowe reported personal fees from Merck, non-financial support from CSL Behring, and non-financial support from Fujifilm Chemical Co. outside the submitted work. No other disclosures were reported., (© 2021 Bohrer et al.)

Subjects with immune-mediated inflammatory diseases (IMID), such as rheumatoid arthritis (RA), have an intrinsic higher probability to develop active-tuberculosis (TB) compared to the general population. The risk ranges from 2.0 to 8.9 in RA patients not receiving therapies. According to the WHO, the RA prevalence varies between 0.3% and 1% and is more common in women and in developed countries. Therefore, the identification and treatment of TB infection (TBI) in this fragile population is important to propose the TB preventive therapy. We aimed to study the M. tuberculosis (Mtb) specific T-cell response to find immune biomarkers of Mtb burden or Mtb clearance in patients with different TB status and different risk to develop active-TB disease. We enrolled TBI subjects as example of Mtb-containment, the active-TB as example of a replicating Mtb status, and the TBI-IMID as fragile population. To study the Mtb-specific response in a condition of possible Mtb sterilization, we longitudinally enrolled TBI subjects and active-TB patients before and after TB therapy. Peripheral blood mononuclear cells were stimulated overnight with Mtb peptides contained in TB1- and TB2-tubes of the Quantiferon-Plus kit. Then, we characterized by cytometry the Mtb-specific CD4 and CD8 T cells. In TBI-IMID, the TB therapy did not affect the ability of CD4 T cells to produce interferon-γ, tumor necrosis factor-α, and interleukin-2, their functional status, and their phenotype. The TB therapy determined a contraction of the triple functional CD4 T cells of the TBI subjects and active-TB patients. The CD45RA - CD27 + T cells stood out as a main subset of the Mtb-specific response in all groups. Before the TB-preventive therapy, the TBI subjects had higher proportion of Mtb-specific CD45RA - CD27 + CD4 + T cells and the active-TB subjects had higher proportion of Mtb-specific CD45RA - CD27 - CD4 + T cells compared to other groups. The TBI-IMID patients showed a phenotype similar to TBI, suggesting that the type of IMID and the IMID therapy did not affect the activation status of Mtb-specific CD4 T cells. Future studies on a larger and better-stratified TBI-IMID population will help to understand the change of the Mtb-specific immune response over time and to identify possible immune biomarkers of Mtb-containment or active replication., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Petruccioli, Petrone, Chiacchio, Farroni, Cuzzi, Navarra, Vanini, Massafra, Lo Pizzo, Guggino, Caccamo, Cantini, Palmieri and Goletti.)

With an estimated 25% of the global population infected with Mycobacterium tuberculosis ( Mtb ), tuberculosis (TB) remains a leading cause of death by infectious diseases. Humoral immunity following TB treatment is largely uncharacterized, and antibody profiling could provide insights into disease resolution. Here we focused on the distinctive TB-specific serum antibody features in active TB disease (ATB) and compared them with latent TB infection (LTBI) or treated ATB (txATB). As expected, di-galactosylated glycan structures (lacking sialic acid) found on IgG-Fc differentiated LTBI from ATB, but also discriminated txATB from ATB. Moreover, TB-specific IgG4 emerged as a novel antibody feature that correlated with active disease, elevated in ATB, but significantly diminished after therapy. These findings highlight 2 novel TB-specific antibody changes that track with the resolution of TB and may provide key insights to guide TB therapy., Competing Interests: GA is a co-founder of SeromYx Systems Inc. GA’s interests were reviewed and are managed by Massachusetts General Hospital and Partners HealthCare in accordance with their conflict of interest policies. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The handling Editor has declared past collaborations with the authors TO and SJ within the last two years., (Copyright © 2021 Grace, Dolatshahi, Lu, Cain, Palmieri, Petrone, Fortune, Ottenhoff, Lauffenburger, Goletti, Joosten and Alter.)

Major epidemics, including some that qualify as pandemics, such as severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS), HIV, influenza A (H1N1)pdm/09 and most recently COVID-19, affect the lung. Tuberculosis (TB) remains the top infectious disease killer, but apart from syndemic TB/HIV little is known regarding the interaction of viral epidemics and pandemics with TB. The aim of this consensus-based document is to describe the effects of viral infections resulting in epidemics and pandemics that affect the lung (MERS, SARS, HIV, influenza A (H1N1)pdm/09 and COVID-19) and their interactions with TB. A search of the scientific literature was performed. A writing committee of international experts including the European Centre for Disease Prevention and Control Public Health Emergency (ECDC PHE) team, the World Association for Infectious Diseases and Immunological Disorders (WAidid), the Global Tuberculosis Network (GTN), and members of the European Society of Clinical Microbiology and Infectious Diseases (ESCMID) Study Group for Mycobacterial Infections (ESGMYC) was established. Consensus was achieved after multiple rounds of revisions between the writing committee and a larger expert group. A Delphi process involving the core group of authors (excluding the ECDC PHE team) identified the areas requiring review/consensus, followed by a second round to refine the definitive consensus elements. The epidemiology and immunology of these viral infections and their interactions with TB are discussed with implications for diagnosis, treatment and prevention of airborne infections (infection control, viral containment and workplace safety). This consensus document represents a rapid and comprehensive summary on what is known on the topic., Competing Interests: Conflict of interest: G.B. Migliori has nothing to disclose. Conflict of interest: M. Raviglione has nothing to disclose. Conflict of interest: G. MacGregor-Skinner has nothing to disclose. Conflict of interest: G. Sotgiu has nothing to disclose. Conflict of interest: J-W. Alffenaar has nothing to disclose. Conflict of interest: S. Tiberi has nothing to disclose. Conflict of interest: C. Adlhoch has nothing to disclose. Conflict of interest: T. Alonzi has nothing to disclose. Conflict of interest: S. Archuleta has nothing to disclose. Conflict of interest: S. Brusin has nothing to disclose. Conflict of interest: E. Cambau has nothing to disclose. Conflict of interest: M.R. Capobianchi has nothing to disclose. Conflict of interest: C. Castilletti has nothing to disclose. Conflict of interest: R. Centis has nothing to disclose. Conflict of interest: D.M. Cirillo has nothing to disclose. Conflict of interest: L. D'Ambrosio has nothing to disclose. Conflict of interest: G. Delogu has nothing to disclose. Conflict of interest: S.M.R. Esposito has nothing to disclose. Conflict of interest: J. Figueroa has nothing to disclose. Conflict of interest: J.S. Friedland has nothing to disclose. Conflict of interest: B.C.H. Ho has nothing to disclose. Conflict of interest: G. Ippolito has nothing to disclose. Conflict of interest: M. Jankovic has nothing to disclose. Conflict of interest: H.Y. Kim has nothing to disclose. Conflict of interest: S. Rosales Klintz has nothing to disclose. Conflict of interest: C. Ködmön has nothing to disclose. Conflict of interest: E. Lalle has nothing to disclose. Conflict of interest: Y.S. Leo has nothing to disclose. Conflict of interest: C-C. Leung has nothing to disclose. Conflict of interest: A-G. Märtson has nothing to disclose. Conflict of interest: M.G. Melazzini has nothing to disclose. Conflict of interest: S. Najafi Fard has nothing to disclose. Conflict of interest: P. Penttinen has nothing to disclose. Conflict of interest: L. Petrone has nothing to disclose. Conflict of interest: E. Petruccioli has nothing to disclose. Conflict of interest: E. Pontali has nothing to disclose. Conflict of interest: L. Saderi has nothing to disclose. Conflict of interest: M. Santin has nothing to disclose. Conflict of interest: A. Spanevello has nothing to disclose. Conflict of interest: R. van Crevel has nothing to disclose. Conflict of interest: M.J. van der Werf has nothing to disclose. Conflict of interest: D. Visca has nothing to disclose. Conflict of interest: M. Viveiros has nothing to disclose. Conflict of interest: J-P. Zellweger has nothing to disclose. Conflict of interest: A. Zumla has nothing to disclose. Conflict of interest: D. Goletti has nothing to disclose. Conflict of interest: C.W.M. Ong has nothing to disclose., (Copyright ©ERS 2020.)

Introduction: Janus kinases inhibitors (anti-JAKs), including tofacitinib, baricitinib, upadacitinib, and filgotinib, represent a new class of synthetic targeted drugs for the treatment of rheumatoid arthritis (RA). In this review, the risk of active tuberculosis (TB) occurrence in patients receiving anti-JAKs was assessed. The literature on this topic, updated to 29 February 2020 was reviewed. Overall, 40 reports (22 tofacitinib, 10 baricitinib, 5 upadacitinib, 3 filgotinib) were examined. A low frequency, not exceeding 0.25%, of active TB cases in patients were exposed to anti-JAKs. Only 1 of 89 recorded cases in tofactinib and baricitinib exposure occurred in countries at intermediate or high TB risk, and most of the cases probably were due to first mycobacterium tuberculosis (Mtb) exposure. Although no cases were observed in patients receiving upadacitinib and filgotinib, long-term trials and data from real-life are required to more precisely address the TB risk associated with the two drugs., Areas Covered: Discussion on the TB risk associated with anti-JAKs, and on the need for accurate evaluation of host-related risk factors in high risk countries., Expert Opinion: Available data on anti-JAKs suggest a negligible risk of active TB occurrence in low endemic areas.

Estimated 2017 tuberculosis (TB) incidence is 10 million and mainly depends on the reservoir of individuals with latent TB infection (LTBI). Quantiferon Ⓡ -TB Gold in-Tube (QFT-GIT) is one of the tests used for LTBI detection. Since 2015 a new version, Quantiferon Ⓡ -TB Gold Plus (QFT-Plus) is available., Objectives: To perform a systematic review and meta-analysis to assess the diagnostic accuracy for TB of QFT-Plus compared to QFT-GIT., Methods: PubMed and Scopus were used to detect records related to predefined strings from 2015 to 2018. Full text articles dealing with the sensitivity and/or specificity of the QFT-Plus vs. QFT-GIT for active-TB and LTBI detection were analyzed. Scientific quality and risk of bias were assessed using QADAS-2., Results: We selected 15 articles. Studies were mainly observational and cross-sectional, performed in 8 countries. Sample size differed in the TB group (27 to 164) compared to LTBI group (29 to 1031). Pooled sensitivity of QFT-Plus for active-TB was 0.94 (0.91 and 0.95 for TB1 and TB2, respectively), whereas pooled specificity for healthy status was 0.96. Pooled sensitivity and specificity for LTBI was 0.91 and 0.95, respectively., Conclusions: We show that QFT-Plus is more sensitive compared to QFT-GIT for detecting M. tuberculosis infection, mainly due to TB2 responses., (Copyright © 2019. Published by Elsevier Ltd.)

Introduction: Two classes of biologics, anti-tumor necrosis factor (TNF) and non-anti-TNF targeted, are currently available for the treatment of rheumatic diseases., Areas Covered: Discussion on the need for LTBI diagnosis in rheumatic patients treated csDMARDs and non-anti-TNFs through a review of the literature. The literature, updated to 15 April 2019, on tuberculosis (TB) reactivation risk in patients exposed to conventional synthetic disease-modifying antirheumatic drugs (csDMARDs) and non-anti-TNF biologics was reviewed., Expert Opinion: An increased risk of TB reactivation in patients receiving csDMARDs (except sulphasalazine) resulted, while a review of clinical trials, and Periodic Safety Update Reports from pharmaceutical Companies evidenced a very low or absent risk for non-anti-TNF biologics. Hence, a contradiction emerges considering that latent TB infection (LTBI) screening is recommended for non-anti-TNF candidates but not for csDMARDs. Concerning the low TB incidence countries, several actions could be undertaken, including to screen all patients independently on the treatment, to omit the procedure in non-anti-TNF candidates, or to perform the LTBI investigations only in high-risk patients. According to WHO guidelines, LTBI screening in low TB risk countries seems unnecessary, except in high TB risk subjects.

Objectives: Biomarkers for tuberculosis (TB) diagnosis and clinical management are needed to defeat TB. In chronic hepatitis, patients not responding to interferon/ribavirin treatment had high levels of an antagonist form of IP-10. Recently, antagonist IP-10 has been shown to be involved also in TB pathogenesis. Here, we investigated IP-10 agonist/antagonist forms as potential inflammatory biomarkers to support TB diagnosis and monitoring., Methods: Total IP-10 and its agonist/antagonist forms were measured by SIMOA digital ELISA in urine obtained from patients with active TB at baseline and after treatment. Healthy donors (HD) and patients with pneumonia were enrolled as controls., Results: Patients with active TB had significantly higher levels of total and agonist IP-10 at baseline compared to HD; conversely, no differences were observed between IP-10 levels in active TB vs pneumonia. Moreover, in active TB a decline of total urine IP-10 was observed at therapy completion; agonist/antagonist forms reflected this decline although their differences were not statistically significant., Conclusions: We showed for the first time that agonist/antagonist IP-10 forms are measurable in urine. IP-10 levels associate with TB and pneumonia disease, suggesting their association with acute inflammation. Further studies are needed to assess their role to monitor TB treatment efficacy., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Introduction: Latent tuberculosis infection (LTBI) accounts for almost a quarter of the world population, and, in 5-10% of the subjects with impaired immune-response against M. tuberculosis growth, it may progress to active tuberculosis (TB). In this review, we focus on the need to propose a screening for LTBI including preventive therapy offer in rheumatic patients undergoing therapy with biological drugs. Areas covered: We report on evidence that biologics are associated with an increased risk of active TB reactivation. This effect seems to be mainly limited to treatment with anti-tumor necrosis factor (TNF) agents, while non-anti-TNF-targeted biologics are not likely associated to any increased risk. We introduce the concept that the patients' coexisting host-related risk factors, such as comorbidities, are crucial to identify those at higher risk to reactivate TB. We report that preventive TB therapy is well tolerated in patients treated with biological drugs. Expert commentary: Availability of non-anti-TNF targeted biologics, that are not associated with an increased risk of TB reactivation, offers a great opportunity to tailor a therapeutic intervention at low/absent TB risk. After proper LTBI screening investigations, preventive TB therapy has been demonstrated to be effective and well-tolerated to reduce the risk of TB reactivation in rheumatic patients requiring biological drugs.

Tuberculosis (TB) still represents an important issue for public health in underdeveloped countries, but the use of antitumor necrosis factor agents (anti-TNF) for the treatment of inflammatory rheumatic disorders has reopened the problem also in countries with low TB incidence, due to the increased risk of TB reactivation in subjects with latent tuberculosis infection (LTBI). Over the last 5 years, several non-anti-TNF-targeted biologics have been licensed for the treatment of rheumatoid arthritis, ankylosing spondylitis, and psoriatic arthritis. We reviewed the epidemiology of TB, the role of different cytokines and of the immune system cells involved in the immune response against TB infection, the methods to detect LTBI, and the risk of TB reactivation in patients exposed to non-anti-TNF-targeted biologics. Given the limited role exerted by the cytokines different from TNF, as expected, data from controlled trials, national registries of biologics, and postmarketing surveillance show that the risk of TB reactivation in patients receiving non-anti-TNF-targeted biologics is negligible, hence raising the question whether the screening procedures for LTBI would be necessary.

Blood-based biomarkers tests are attractive alternative for diagnosing tuberculosis to assays depending on mycobacteria detection. Given several immunological markers we used logistic regression to model the probability of active tuberculosis in a cohort of patients with active or latent tuberculosis, showing an increased accuracy in distinguishing active from latent tuberculosis., (Copyright © 2016 Elsevier Inc. All rights reserved.)

B-cells not only produce immunoglobulins and present antigens to T-cells, but also additional key roles in the immune system. Current knowledge on the role of B-cells in infections caused by intracellular bacteria is fragmentary and contradictory. We therefore analysed the phenotypical and functional properties of B-cells during infection and disease caused by Mycobacterium tuberculosis (Mtb), the bacillus causing tuberculosis (TB), and included individuals with latent TB infection (LTBI), active TB, individuals treated successfully for TB, and healthy controls. Patients with active or treated TB disease had an increased proportion of antibodies reactive with mycobacteria. Patients with active TB had reduced circulating B-cell frequencies, whereas only minor increases in B-cells were detected in the lungs of individuals deceased from TB. Both active TB patients and individuals with LTBI had increased relative fractions of B-cells with an atypical phenotype. Importantly, these B-cells displayed impaired proliferation, immunoglobulin- and cytokine- production. These defects disappeared upon successful treatment. Moreover, T-cell activity was strongest in individuals successfully treated for TB, compared to active TB patients and LTBI subjects, and was dependent on the presence of functionally competent B-cells as shown by cellular depletion experiments. Thus, our results reveal that general B-cell function is impaired during active TB and LTBI, and that this B-cell dysfunction compromises cellular host immunity during Mtb infection. These new insights may provide novel strategies for correcting Mtb infection-induced immune dysfunction towards restored protective immunity.

Unlabelled: There are still no reliable tests to distinguish active tuberculosis (TB) from latent TB infection (LTBI). Assessment of CD27 modulation on CD4⁺ T-cells has been suggested as a tool to diagnose different TB stages., Objectives: To use several cytometric approaches to evaluate CD27 expression on Mycobacterium tuberculosis (Mtb)-specific CD4⁺ T-cells to differentiate TB stages., Methods: 55 HIV-uninfected subjects were enrolled: 13 active TB; 12 cured TB; 30 LTBI. Whole blood was stimulated with RD1-proteins or Cytomegalovirus-lysate (CMV). Interferon (IFN)-γ response was evaluated by cytometry. The proportion of CD27(±) within the IFN-γ⁺ CD4⁺ T-cells or RATIO of the CD27-median fluorescence intensity (MFI) of CD4⁺ T-cells over the CD27 MFI of IFN-γ⁺ CD4⁺ T-cells was evaluated., Results: The greatest diagnostic accuracy in discriminating active TB vs. LTBI or cured TB was reached by evaluating the CD27(+) CD45RA(-) cells within the IFN-γ⁺ CD4⁺ T-cell subset (76.92 sensitivity for both, and 90% and 91.67% specificity, respectively), although the use of the CD27 MFI RATIO allows for stricter data analysis, independent of the operator., Conclusions: the study of CD27 expression using different approaches, whether it involves evaluation of CD45RA expression or not, is a robust biomarker for discriminating TB stages., (Copyright © 2015 The British Infection Association. Published by Elsevier Ltd. All rights reserved.)

Objectives: Controversial results were reported on the role of polyfunctional T-cells in tuberculosis (TB). Our aim was to simultaneously characterize the Mycobacterium tuberculosis (Mtb)-specific immune response as cytokine production and memory phenotype by flow cytometry after in vitro stimulation with region of difference 1 ("RD1") recombinant proteins (ESAT-6 and CFP-10) in patients at different TB stage in a low TB endemic country. To assess the specificity of these findings, we evaluated the response to cytomegalovirus (CMV), an unrelated antigen., Methods: We enrolled subjects with active TB, cured TB, latent TB infection (LTBI). Cytokine and phenotype profiles of T-cells from whole blood stimulated with "RD1" proteins and CMV were characterized by multi-parametric flow cytometry., Results: Bifunctional IFNγ(+) TNFα(+) CD4(+) T-cells and effector memory phenotype significantly associated with active TB compared to the LTBI group (p = 0.008, at least p ≤ 0.009 respectively) whereas "RD1"-T-cell response in cured TB and LTBI was characterized by a central memory phenotype (at least p ≤ 0.013 and p ≤ 0.004 respectively vs active TB). In contrast, response to CMV antigen was not associated with a TB-specific status., Conclusion: We identified qualitative associations between Mtb-specific T-cell and TB status in terms of functional capacity and memory status. These immune correlates may be helpful to trace natural history of TB., (Copyright © 2013 The British Infection Association. Published by Elsevier Ltd. All rights reserved.)

Both humoral and cell-mediated responses are associated with immunity to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Although our understanding of the potential role of T-cell responses in the context of coronavirus disease 2019 (COVID-19) is rapidly increasing, more information is still needed. To provide an overview of the role of T-cell immunity in COVID-19, in the context of natural infection and post-vaccination, and discuss the potential utility of measuring SARS-CoV-2-specific T-cell responses, drawing on experience of the use of interferon-γ release assays (IGRAs) in tuberculosis (TB). PubMed articles up to 16 April 2021. T-cell responses can be detected very early in the course of COVID-19, earlier than the detection of antibody responses, and are correlated with COVID-19 outcome. Lower CD4+ and CD8+ T-cell counts are markers of more severe disease, longer duration of viral RNA positivity and increased mortality. In line with natural infection, SARS-CoV-2 vaccination stimulates robust T-cell responses, which probably play an important role in protection; data on long-term T-cell responses are currently limited. The utility of measuring T-cell responses is already well established in both aiding the diagnosis of TB infection using IGRAs, and evaluation of T-cell responses to TB vaccine candidates. A variety of assays have already been developed to measure SARS-CoV-2-specific T-cell responses, including IGRAs, intracellular cytokine staining and activation-induced markers. IGRAs based on SARS-CoV-2 antigens can distinguish between convalescent and uninfected healthy blood donors. Simple assays for measuring the quantity and function of T-cell responses may have utility in the prognostication of COVID-19, and for monitoring immune responses to SARS-CoV-2 vaccination and population-based immunity to SARS-CoV-2 variants of interest. [ABSTRACT FROM AUTHOR]

Background: Recent studies proposed the whole-blood based IFN-γ-release assay to study the antigen-specific SARS-CoV-2 response. Since the early prediction of disease progression could help to assess the optimal treatment strategies, an integrated knowledge of T-cell and antibody response lays the foundation to develop biomarkers monitoring the COVID-19. Whole-blood-platform tests based on the immune response detection to SARS-CoV2 peptides is a new approach to discriminate COVID-19-patients from uninfected-individuals and to evaluate the immunogenicity of vaccine candidates, monitoring the immune response in vaccine trial and supporting the serological diagnostics results. Here, we aimed to identify in the whole-blood-platform the best immunogenic viral antigen and the best immune biomarker to identify COVID-19-patients.Methods: Whole-blood was overnight-stimulated with SARS-CoV-2 peptide pools of nucleoprotein-(NP) Membrane-, ORF3a- and Spike-protein. We evaluated: IL-1β, IL-1Ra, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12p70, IL-13, IL- 15, IL-17A, eotaxin, FGF, G-CSF, GM-CSF, IFN-γ, IP-10, MCP-1, MIP-1α, MIP-1β, PDGF, RANTES, TNF-α, VEGF. By a sparse partial least squares discriminant analysis we identified the most important soluble factors discriminating COVID-19- from NO-COVID-19-individuals.Results: We identified a COVID-19 signature based on six immune factors: IFN-γ, IP-10 and IL-2 induced by Spike; RANTES and IP-10 induced by NP and IL-2 induced by ORF3a. We demonstrated that the test based on IP-10 induced by Spike had the highest AUC (0.85, p  <  0.0001) and that the clinical characteristics of the COVID-19-patients did not affect IP-10 production. Finally, we validated the use of IP-10 as biomarker for SARS-CoV2 infection in two additional COVID-19-patients cohorts.Conclusions: We set-up a whole-blood assay identifying the best antigen to induce a T-cell response and the best biomarkers for SARS-CoV-2 infection evaluating patients with acute COVID-19 and recovered patients. We focused on IP-10, already described as a potential biomarker for other infectious disease such as tuberculosis and HCV. An additional application of this test is the evaluation of immune response in SARS-CoV-2 vaccine trials: the IP-10 detection may define the immunogenicity of a Spike-based vaccine, whereas the immune response to the virus may be evaluated detecting other soluble factors induced by other viral-antigens. [ABSTRACT FROM AUTHOR]

Host-directed therapies (HDTs) are emerging as a potential valid support in the treatment of drug-resistant tuberculosis (TB). Following our recent report indicating that genetic and pharmacological inhibition of transglutaminase 2 (TG2) restricts Mycobacterium tuberculosis (Mtb) replication in macrophages, we aimed to investigate the potentials of the TG2 inhibitors cystamine and cysteamine as HDTs against TB. We showed that both cysteamine and cystamine restricted Mtb replication in infected macrophages when provided at equimolar concentrations and did not exert any antibacterial activity when administered directly on Mtb cultures. Interestingly, infection of differentiated THP-1 mRFP-GFP-LC3B cells followed by the determination of the autophagic intermediates pH distribution (AIPD) showed that cystamine inhibited the autophagic flux while restricting Mtb replication. Moreover, both cystamine and cysteamine had a similar antimicrobial activity in primary macrophages infected with a panel of Mtb clinical strains belonging to different phylogeographic lineages. Evaluation of cysteamine and cystamine activity in the human ex vivo model of granuloma-like structures (GLS) further confirmed the ability of these drugs to restrict Mtb replication and to reduce the size of GLS. The antimicrobial activity of the TG2 inhibitors synergized with a second-line anti-TB drug as amikacin in human monocyte-derived macrophages and in the GLS model. Overall, the results of this study support the potential usefulness of the TG2-inhibitors cysteamine and cystamine as HDTs against TB. [ABSTRACT FROM AUTHOR]

Interferon-γ inducible protein 10 (IP-10), is a potent chemoattractant that promotes migration of monocytes and activated T-cells to inflammation foci. IP-10 is elevated in serum of patients with chronic hepatitis C virus (HCV) and tuberculosis (TB) infections, although it remains to be determined the contribution of IP-10 in restricting Mycobacterium tuberculosis (Mtb) replication. Here, we investigated the impact of IP-10 on mycobacteria replication using the ex vivo model of human whole-blood (WB) assay. In particular, we compared the levels of IP-10 upon infection with different Mtb clinical strains and species of non-tuberculous mycobacteria (NTM) and evaluated how IP-10 may contain bacterial replication. Interestingly, we observed that the inhibition of the host enzyme dipeptidyl peptidase IV (DPP-IV), which inactivates IP-10 through cleavage of two amino acids at the chemokine N-terminus, restricted mycobacterial persistence in WB, supporting the critical role of full length IP-10 in mediating an anti- Mtb response. Addition of recombinant IP-10 expressed in eukaryotic cells enhanced the anti-mycobacterial activity in WB, although no differences were observed when IP-10 containing different proportions of cleaved and non-cleaved forms of the chemokine were added. Moreover, recombinant IP-10 did not exert a direct anti-mycobacterial effect. Our results underscore the clinical relevance of IP-10 in mycobacteria pathogenesis and support the potential outcomes that may derive by targeting the IP-10/CXCR3 pathway as host directed therapies for the treatment of Mtb or NTM infections. [ABSTRACT FROM AUTHOR]

Background: Tuberculosis (TB) remains a major threat to global health. Currently, diagnosis of active TB is hampered by the lack of specific biomarkers that discriminate active TB disease from other (lung) diseases or latent TB infection (LTBI). Integrated human gene expression results have shown that genes encoding complement components, in particular different C1q chains, were expressed at higher levels in active TB compared to LTBI. Methods: C1q protein levels were determined using ELISA in sera from patients, from geographically distinct populations, with active TB, LTBI as well as disease controls. Results: Serum levels of C1q were increased in active TB compared to LTBI in four independent cohorts with an AUC of 0.77 [0.70; 0.83]. After 6 months of TB treatment, levels of C1q were similar to those of endemic controls, indicating an association with disease rather than individual genetic predisposition. Importantly, C1q levels in sera of TB patients were significantly higher as compared to patients with sarcoidosis or pneumonia, clinically important differential diagnoses. Moreover, exposure to other mycobacteria, such as Mycobacterium leprae (leprosy patients) or BCG (vaccinees) did not result in elevated levels of serum C1q. In agreement with the human data, in non-human primates challenged with Mycobacterium tuberculosis , increased serum C1q levels were detected in animals that developed progressive disease, not in those that controlled the infection. Conclusions: In summary, C1q levels are elevated in patients with active TB compared to LTBI in four independent cohorts. Furthermore, C1q levels from patients with TB were also elevated compared to patients with sarcoidosis, leprosy and pneumonia. Additionally, also in NHP we observed increased C1q levels in animals with active progressive TB, both in serum and in broncho-alveolar lavage. Therefore, we propose that the addition of C1q to current biomarker panels may provide added value in the diagnosis of active TB. [ABSTRACT FROM AUTHOR]

The QuantiFERON-TB Gold Plus (QFT-Plus) is a new test for latent tuberculosis infection (LTBI) diagnosis, in which has been added a new tube containing shorter peptides stimulating CD8 T-cells and CD4-stimulating-peptides. Measurement of alternative biomarkers to Interferon-γ (IFN-γ) in QFT-Plus may improve its sensitivity. Interferon-γ inducible protein 10 (IP-10), has been proposed as a tuberculosis (TB) biomarker. We aimed to evaluate the IP-10 accuracy in QFT-Plus for LTBI diagnosis. QFT-Plus was performed in 36 active TB, 31 LTBI and 16 healthy donors (HD). IP-10 was detected by ELISA. IP-10 is increased in TB1 and TB2 tubes in subjects with active TB and LTBI compared to HD. A ROC analysis comparing active TB and HD was performed and a cut-off of 1174 pg/mL for TB1 and 928.8 pg/mL for TB2 identified active TB with 86% sensitivity (Se) and 94% specificity (Sp). Moreover, increased IP-10 in response to TB1 was found in subjects with LTBI compared to those with active TB. A cut-off point of ≥16,108 pg/mL was chosen to maximize the test performance. However, the test predicted LTBI only with 58% Se and 61% Sp. These results suggest that IP-10 is an alternative biomarker to IFN-γ in the QFT-Plus format. [ABSTRACT FROM AUTHOR]

Introduction: Mycobacterium tuberculosis is one of the world’s most successful pathogens equipped to establish itself within the human host as a subclinical infection without overt disease. Unable to eradicate the bacteria, the immune system contains the infection in a granuloma structure. Th1 cells that are essential for infection control are recruited to the site of infection directed by chemokines, predominantly CXCL10. It has previously been shown that CXCL10 in the plasma of patients chronically infected with hepatitis C virus is present primarily in an antagonist form. This is due to N-terminal truncation by the enzyme DPP4, which results in the antagonist form that is capable of binding its receptor CXCR3, but does not induce signaling. We aimed to explore whether such CXCL10 antagonism may have an impact on the pathogenesis of tuberculosis (TB). Results: We measured plasma levels of agonist and antagonist CXCL10 by Simoa digital ELISA, as well as DPP4 enzyme activity in the plasma of 20 patients with active TB infection, 10 patients with pneumonia infection, and a group of 10 healthy controls. We found higher levels of total and antagonist CXCL10 and reduced DPP4 enzyme activity in the plasma of TB patients compared to controls. We traced the source of CXCL10 secretion using immunohistochemical and confocal analysis to multinucleated giant cells in the TB lesions, and variable expression by macrophages. Interestingly, these cells were associated with DPP4-positive T cells. Moreover, the analysis of lymphocytes at the site of TB infection (bronchoalveolar lavage) showed a reduced frequency of CXCR3+ T cells. Interpretation: Our data suggests that CXCL10 antagonism may be an important regulatory mechanism occurring at the site of TB pathology. CXCL10 can be inactivated shortly after secretion by membrane bound DPP4 (CD26), therefore, reducing its chemotactic potential. Given the importance of Th1 cell functions and IFN-γ-mediated effects in TB, our data suggest a possible unappreciated regulatory role of DPP4 in TB. Perspectives: DPP4 is the target for a class of enzyme inhibitors used in the treatment of diabetes, and the results from this study suggest that these drugs could be repurposed as an adjunct immunotherapy of patients with TB and MDR-TB. [ABSTRACT FROM AUTHOR]

Individuals exposed to Mycobacterium tuberculosis (Mtb) may be infected and remain for the entire life in this condition defined as latent tuberculosis infection (LTBI) or develop active tuberculosis (TB). Among the multiple factors governing the outcome of the infection, dendritic cells (DCs) play a major role in dictating antibacterial immunity. However, current knowledge on the role of the diverse components of human DCs in shaping specific T-cell response during Mtb infection is limited. In this study, we performed a comparative evaluation of peripheral blood circulating DC subsets as well as of monocyte-derived Interferon-α DCs (IFN-DCs) from patients with active TB, subjects with LTBI and healthy donors (HD). The proportion of circulating myeloid BDCA3+ DCs (mDC2) and plasmacytoid CD123+ DCs (pDCs) declined significantly in active TB patients compared to HD, whereas the same subsets displayed a remarkable activation in LTBI subjects. Simultaneously, the differentiation of IFN-DCs from active TB patients resulted profoundly impaired compared to those from LTBI and HD individuals. Importantly, the altered developmental trait of IFN-DCs from active TB patients was associated with down-modulation of IFN-linked genes, marked changes in molecular signaling conveying antigen (Ag) presentation and full inability to induce Ag-specific T cell response. Thus, these data reveal an important role of IFN-α in determining the induction of Mtb-specific immunity. [ABSTRACT FROM AUTHOR]

• SERPING1 expression was increased in patients with active tuberculosis. • C1-inhibitor proteins levels were increased in half of the TB cohorts analysed. • C1-inhibitor was not increased in other clinically relevant conditions. • C1-inhibitor levels normalised over the course of treatment. • A positive correlation between C1q and C1-inhibitor was found in TB patients. To facilitate better discrimination between patients with active tuberculosis (TB) and latent TB infection (LTBI), whole blood transcriptomic studies have been performed to identify novel candidate host biomarkers. SERPING1 , which encodes C1-inhibitor (C1-INH), the natural inhibitor of the C1-complex has emerged as candidate biomarker. Here we collated and analysed SERPING1 expression data and subsequently determined C1-INH protein levels in four cohorts of patients with TB. SERPING1 expression data were extracted from online deposited datasets. C1-INH protein levels were determined by ELISA in sera from individuals with active TB, LTBI as well as other disease controls in geographically diverse cohorts. SERPING1 expression was increased in patients with active TB compared to healthy controls (8/11 cohorts), LTBI (13/14 cohorts) and patients with other (non-TB) lung-diseases (7/7 cohorts). Serum levels of C1-INH were significantly increased in The Gambia and Italy in patients with active TB relative to the endemic controls but not in South Africa or Korea. In the largest cohort (n = 50), with samples collected longitudinally, normalization of C1-INH levels following successful TB treatment was observed. This cohort, also showed the most abundant increase in C1-INH, and a positive correlation between C1q and C1-INH levels. Combined presence of increased levels of both C1q and C1-INH had high specificity for active TB (96 %) but only very modest sensitivity 38 % compared to the endemic controls. SERPING1 transcript expression is increased in TB patients, while serum protein levels of C1-INH were increased in half of the cohorts analysed. [ABSTRACT FROM AUTHOR]