Antigen-Specific T-Cell Activation Distinguishes between Recent and Remote Tuberculosis Infection

Rationale: Current diagnostic tests fail to identify individuals at higher risk of progression to tuberculosis disease, such as those with recent Mycobacterium tuberculosis infection, who should be prioritized for targeted preventive treatment. Objectives: To define a blood-based biomarker, measured with a simple flow cytometry assay, that can stratify different stages of tuberculosis infection to infer risk of disease. Methods: South African adolescents were serially tested with QuantiFERON-TB Gold to define recent (QuantiFERON-TB conversion 1 yr) infection. We defined the ΔHLA-DR median fluorescence intensity biomarker as the difference in HLA-DR expression between IFN-γ+ TNF+ Mycobacterium tuberculosis-specific T cells and total CD3+ T cells. Biomarker performance was assessed by blinded prediction in untouched test cohorts with recent versus persistent infection or tuberculosis disease and by unblinded analysis of asymptomatic adolescents with tuberculosis infection who remained healthy (nonprogressors) or who progressed to microbiologically confirmed disease (progressors). Measurements and Main Results: In the test cohorts, frequencies of Mycobacterium tuberculosis-specific T cells differentiated between QuantiFERON-TB- (n = 25) and QuantiFERON-TB+ (n = 47) individuals (area under the receiver operating characteristic curve, 0.94; 95% confidence interval, 0.87-1.00). ΔHLA-DR significantly discriminated between recent (n = 20) and persistent (n = 22) QuantiFERON-TB+ (0.91; 0.83-1.00); persistent QuantiFERON-TB+ and newly diagnosed tuberculosis (n = 19; 0.99; 0.96-1.00); and tuberculosis progressors (n = 22) and nonprogressors (n = 34; 0.75; 0.63-0.87). However, ΔHLA-DR median fluorescent intensity could not discriminate between recent QuantiFERON-TB+ and tuberculosis (0.67; 0.50-0.84). Conclusions: The ΔHLA-DR biomarker can identify individuals with recent QuantiFERON-TB conversion and those with disease progression, allowing targeted provision of preventive treatment to those at highest risk of tuberculosis. Further validation studies of this novel immune biomarker in various settings and populations at risk are warranted

Phenotypic correlation, path coefficient and multivariate analysis for yield and yield-associated traits in groundnut accessions

Yield is a complex quantitative trait largely influenced by the environment. Direct selection for grain yield is less efficient in improving groundnut productivity. The selection efficiency can be enhanced by exploiting the relationship between yield and its related traits. Moreover, the use of genetically diverse parents is essential to generate genetic variation for successful selection of genotypes in a breeding program. Therefore, the study aimed at analysing the relationship between grain yield and its related traits and determining the morphological diversity among selected groundnut genotypes under natural rosette disease (GRD) infestation. The genotypes were evaluated in a 7 × 4 alpha lattice design with three replications. Data were collected on yield and yield-related traits. Correlation, path coefficient and multivariate analyses were done. The results revealed that yield was directly associated with plant height, number of pods per plant, hundred seed weight, GRD incidence and number of secondary branches. Therefore, these traits should be considered in selection when improving groundnut for yield. Cluster analysis revealed existence of diversity among the evaluated groundnut genotypes with no influence of geographical origin to the clustering pattern. The Principal Components Analysis (PCA) biplot was effective in showing the genetic distance among the genotypes and the results were comparable with those of the cluster analysis. Moreover, Shannon-Weaver diversity indices revealed existence of high diversity among the genotypes, an implication that groundnut improvement for yield is possible through selection in breeding

T cell receptor repertoires associated with control and disease progression following Mycobacterium tuberculosis infection

Antigen-specific, MHC-restricted αβ T cells are necessary for protective immunity against Mycobacterium tuberculosis, but the ability to broadly study these responses has been limited. In the present study, we used single-cell and bulk T cell receptor (TCR) sequencing and the GLIPH2 algorithm to analyze M. tuberculosis-specific sequences in two longitudinal cohorts, comprising 166 individuals with M. tuberculosis infection who progressed to either tuberculosis (n = 48) or controlled infection (n = 118). We found 24 T cell groups with similar TCR-β sequences, predicted by GLIPH2 to have common TCR specificities, which were associated with control of infection (n = 17), and others that were associated with progression to disease (n = 7). Using a genome-wide M. tuberculosis antigen screen, we identified peptides targeted by T cell similarity groups enriched either in controllers or in progressors. We propose that antigens recognized by T cell similarity groups associated with control of infection can be considered as high-priority targets for future vaccine development

Multidimensional analyses reveal modulation of adaptive and innate immune subsets by tuberculosis vaccines

We characterize the breadth, function and phenotype of innate and adaptive cellular responses in a prevention of Mycobacterium tuberculosis infection trial. Responses are measured by whole blood intracellular cytokine staining at baseline and 70 days after vaccination with H4:IC31 (subunit vaccine containing Ag85B and TB10.4), Bacille Calmette-Guerin (BCG, a live attenuated vaccine) or placebo (n = ~30 per group). H4:IC31 vaccination induces Ag85B and TB10.4-specific CD4 T cells, and an unexpected NKTlike subset, that expresses IFN-γ, TNF and/or IL-2. BCG revaccination increases frequencies of CD4 T cell subsets that either express Th1 cytokines or IL-22, and modestly increases IFNγ-producing NK cells. In vitro BCG re-stimulation also triggers responses by donor-unrestricted T cells, which may contribute to host responses against mycobacteria. BCG, which demonstrated efficacy against sustained Mycobacterium tuberculosis infection, modulates multiple immune cell subsets, in particular conventional Th1 and Th22 cells, which should be investigated in discovery studies of correlates of protection

The SIGLEC14 null allele is associated with Mycobacterium tuberculosis- and BCG-induced clinical and immunologic outcomes

Humans exposed to Mycobacterium tuberculosis (Mtb) have variable susceptibility to tuberculosis (TB) and its outcomes. Siglec-5 and Siglec-14 are members of the sialic-acid binding lectin family that regulate immune responses to pathogens through inhibitory (Siglec-5) and activating (Siglec-14) domains. The SIGLEC14 coding sequence is deleted in a high proportion of individuals, placing a SIGLEC5-like gene under the expression of the SIGLEC14 promoter (the SIGLEC14 null allele) and causing expression of a Siglec-5 like protein in monocytes and macrophages. We hypothesized that the SIGLEC14 null allele was associated with Mtb replication in monocytes, T-cell responses to the BCG vaccine, and clinical susceptibility to TB. The SIGLEC14 null allele was associated with protection from TB meningitis in Vietnamese adults but not with pediatric TB in South Africa. The null allele was associated with increased IL-2 and IL-17 production following ex-vivo BCG stimulation of blood from 10 week-old South African infants vaccinated with BCG at birth. Mtb replication was increased in THP-1 cells overexpressing either Siglec-5 or Siglec-14 relative to controls. To our knowledge, this is the first study to demonstrate an association between SIGLEC expression and clinical TB, Mtb replication, or BCG-specific T-cell cytokines

The SIGLEC14 null allele is associated with Mycobacterium tuberculosis- and BCG-induced clinical and immunologic outcomes

Humans exposed to Mycobacterium tuberculosis (Mtb) have variable susceptibility to tuberculosis (TB) and its outcomes. Siglec-5 and Siglec-14 are members of the sialic-acid binding lectin family that regulate immune responses to pathogens through inhibitory (Siglec-5) and activating (Siglec-14) domains. The SIGLEC14 coding sequence is deleted in a high proportion of individuals, placing a SIGLEC5-like gene under the expression of the SIGLEC14 promoter (the SIGLEC14 null allele) and causing expression of a Siglec-5 like protein in monocytes and macrophages. We hypothesized that the SIGLEC14 null allele was associated with Mtb replication in monocytes, T-cell responses to the BCG vaccine, and clinical susceptibility to TB. The SIGLEC14 null allele was associated with protection from TB meningitis in Vietnamese adults but not with pediatric TB in South Africa. The null allele was associated with increased IL-2 and IL-17 production following ex-vivo BCG stimulation of blood from 10 week-old South African infants vaccinated with BCG at birth. Mtb replication was increased in THP-1 cells overexpressing either Siglec-5 or Siglec-14 relative to controls. To our knowledge, this is the first study to demonstrate an association between SIGLEC expression and clinical TB, Mtb replication, or BCG-specific T-cell cytokines

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Background<p>Maintenance of long-lasting immunity is thought to depend on stem cell memory T cells (T_SCM), which have superior self-renewing capacity, longevity and proliferative potential compared with central memory (T_CM) or effector (T_EFF) T cells. Our knowledge of T_SCM derives primarily from studies of virus-specific CD8⁺ T_SCM. We aimed to determine if infection with Mycobacterium tuberculosis (M. tb), the etiological agent of tuberculosis, generates antigen-specific CD4⁺ T_SCM and to characterize their functional ontology.</p>Methods<p>We studied T cell responses to natural M. tb infection in a longitudinal adolescent cohort of recent QuantiFERON-TB Gold (QFT) converters and three cross-sectional QFT⁺ adult cohorts; and to bacillus Calmette–Guerin (BCG) vaccination in infants. M. tb and/or BCG-specific CD4 T cells were detected by flow cytometry using major histocompatibility complex class II tetramers bearing Ag85, CFP-10, or ESAT-6 peptides, or by intracellular cytokine staining. Transcriptomic analyses of M. tb-specific tetramer⁺ CD4⁺ T_SCM (CD45RA⁺ CCR7⁺ CD27⁺) were performed by microfluidic qRT-PCR, and functional and phenotypic characteristics were confirmed by measuring expression of chemokine receptors, cytotoxic molecules and cytokines using flow cytometry.</p>Results<p>M. tb-specific T_SCM were not detected in QFT-negative persons. After QFT conversion frequencies of T_SCM increased to measurable levels and remained detectable thereafter, suggesting that primary M. tb infection induces T_SCM cells. Gene expression (GE) profiling of tetramer⁺ T_SCM showed that these cells were distinct from bulk CD4⁺ naïve T cells (T_N) and shared features of bulk T_SCM and M. tb-specific tetramer⁺ T_CM and T_EFF cells. These T_SCM were predominantly CD95⁺ and CXCR3⁺, markers typical of CD8⁺ T_SCM. Tetramer⁺ T_SCM expressed significantly higher protein levels of CCR5, CCR6, CXCR3, granzyme A, granzyme K, and granulysin than bulk T_N and T_SCM cells. M. tb-specific T_SCM were also functional, producing IL-2, IFN-γ, and TNF-α upon antigen stimulation, and their frequencies correlated positively with long-term BCG-specific CD4⁺ T cell proliferative potential after infant vaccination.</p>Conclusion<p>Human infection with M. tb induced distinct, antigen-specific CD4⁺ T_SCM cells endowed with effector functions, including expression of cytotoxic molecules and Th1 cytokines, and displayed chemokine receptor profiles consistent with memory Th1/17 cells. Induction of CD4⁺ T_SCM should be considered for vaccination approaches that aim to generate long-lived memory T cells against M. tb.</p

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Background<p>Maintenance of long-lasting immunity is thought to depend on stem cell memory T cells (T_SCM), which have superior self-renewing capacity, longevity and proliferative potential compared with central memory (T_CM) or effector (T_EFF) T cells. Our knowledge of T_SCM derives primarily from studies of virus-specific CD8⁺ T_SCM. We aimed to determine if infection with Mycobacterium tuberculosis (M. tb), the etiological agent of tuberculosis, generates antigen-specific CD4⁺ T_SCM and to characterize their functional ontology.</p>Methods<p>We studied T cell responses to natural M. tb infection in a longitudinal adolescent cohort of recent QuantiFERON-TB Gold (QFT) converters and three cross-sectional QFT⁺ adult cohorts; and to bacillus Calmette–Guerin (BCG) vaccination in infants. M. tb and/or BCG-specific CD4 T cells were detected by flow cytometry using major histocompatibility complex class II tetramers bearing Ag85, CFP-10, or ESAT-6 peptides, or by intracellular cytokine staining. Transcriptomic analyses of M. tb-specific tetramer⁺ CD4⁺ T_SCM (CD45RA⁺ CCR7⁺ CD27⁺) were performed by microfluidic qRT-PCR, and functional and phenotypic characteristics were confirmed by measuring expression of chemokine receptors, cytotoxic molecules and cytokines using flow cytometry.</p>Results<p>M. tb-specific T_SCM were not detected in QFT-negative persons. After QFT conversion frequencies of T_SCM increased to measurable levels and remained detectable thereafter, suggesting that primary M. tb infection induces T_SCM cells. Gene expression (GE) profiling of tetramer⁺ T_SCM showed that these cells were distinct from bulk CD4⁺ naïve T cells (T_N) and shared features of bulk T_SCM and M. tb-specific tetramer⁺ T_CM and T_EFF cells. These T_SCM were predominantly CD95⁺ and CXCR3⁺, markers typical of CD8⁺ T_SCM. Tetramer⁺ T_SCM expressed significantly higher protein levels of CCR5, CCR6, CXCR3, granzyme A, granzyme K, and granulysin than bulk T_N and T_SCM cells. M. tb-specific T_SCM were also functional, producing IL-2, IFN-γ, and TNF-α upon antigen stimulation, and their frequencies correlated positively with long-term BCG-specific CD4⁺ T cell proliferative potential after infant vaccination.</p>Conclusion<p>Human infection with M. tb induced distinct, antigen-specific CD4⁺ T_SCM cells endowed with effector functions, including expression of cytotoxic molecules and Th1 cytokines, and displayed chemokine receptor profiles consistent with memory Th1/17 cells. Induction of CD4⁺ T_SCM should be considered for vaccination approaches that aim to generate long-lived memory T cells against M. tb.</p