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Rationale South African adolescents carry a high tuberculosis disease burden. It is not known if schools are high-risk settings for Mycobacterium tuberculosis (MTB) transmission. Objectives To detect airborne MTB genomic DNA in classrooms. Methods We studied 72 classrooms occupied by 1,836 students in two South African schools. High-volume air filtration was performed for median 40 minutes (interquartile range 35-54) and assayed by droplet digital PCR (ddPCR) targeting MTB Region of Difference 9 (RD9), with concurrent CO2 concentration measurement. Classroom data were benchmarked against public health clinics. Students who consented to individual TB screening completed a questionnaire and sputum collection (Xpert MTB/RIF Ultra) if symptom-positive. Poisson statistics were used for MTB RD9 copy quantification. Measurements and Main Results ddPCR assays were positive in 13/72 (18.1%) classroom and 4/39 (10.3%) clinic measurements (p=0.276). Median ambient CO2 concentration was 886 ppm (IQR 747-1223) in classrooms vs. 490 ppm (IQR 405-587) in clinics (p

SUMMARYTuberculosis (TB) remains a major public health problem with host-directed therapeutics offering potential as novel treatment strategies. However, their successful development still requires a comprehensive understanding of howMycobacterium tuberculosis(M.tb) infection impacts immune responses. To address this challenge, we applied standardised immunomonitoring tools to compare TB antigen, BCG and IL-1β induced immune responses between individuals with latentM.tbinfection (LTBI) and active TB disease, at diagnosis and after cure. This revealed distinct responses between TB and LTBI groups at transcriptomic, proteomic and metabolomic levels. At baseline, we identified pregnane steroids and the PPARγ pathway as new immune-metabolic drivers of elevated plasma IL-1ra in TB. We also observed dysregulated induced IL-1 responses after BCG stimulation in TB patients. Elevated IL-1 antagonist responses were explained by upstream differences in TNF responses, while for IL-1 agonists it was due to downstream differences in granzyme mediated cleavage. Finally, the immune response to IL-1β driven signalling was also dramatically perturbed in TB disease but was completely restored after successful antibiotic treatment. This systems immunology approach improves our knowledge of how immune responses are altered during TB disease, and may support design of improved diagnostic, prophylactic and therapeutic tools.

Background Tuberculosis (TB) is caused by Mycobacterium tuberculosis (Mtb) infection and is a major public health problem. Clinical challenges include the lack of a blood-based test for active disease. Current blood-based tests, such as QuantiFERON (QFT) do not distinguish active TB disease from asymptomatic Mtb infection. Methods We hypothesized that TruCulture, an immunomonitoring method for whole-blood stimulation, could discriminate active disease from latent Mtb infection (LTBI). We stimulated whole blood from patients with active TB and compared with LTBI donors. Mtb-specific antigens and live bacillus Calmette-Guérin (BCG) were used as stimuli, with direct comparison to QFT. Protein analyses were performed using conventional and digital enzyme-linked immunosorbent assay (ELISA), as well as Luminex. Results TruCulture showed discrimination of active TB cases from LTBI (P < .0001, AUC = .81) compared with QFT (P = .45, AUC = .56), based on an interferon γ (IFNγ) readout after Mtb antigen (Ag) stimulation. This result was replicated in an independent cohort (AUC = .89). In exploratory analyses, TB stratification could be further improved by the Mtb antigen to BCG IFNγ ratio (P < .0001, AUC = .91). Finally, the combination of digital ELISA and transcriptional analysis showed that LTBI donors with high IFNγ clustered with patients with TB, suggesting the possibility to identify subclinical disease. Conclusions TruCulture offers a next-generation solution for whole-blood stimulation and immunomonitoring with the possibility to discriminate active and latent infection., We tested TruCulture, an immunomonitoring tool, to identify active disease from latent Mtb infection. TruCulture showed improved discrimination of tuberculosis cases from LTBI as compared with QuantiFERON. Tuberculosis stratification could be further improved by the Mtb Ag:BCG IFNγ ratio.

Tuberculosis (TB) clinical prediction rules rely on presence of symptoms, however many undiagnosed cases in the community are asymptomatic. This study aimed to explore the utility of clinical factors in predicting TB among people with HIV not seeking care.Baseline data were analysed from an observational cohort of ambulant adults with HIV in South Africa. Participants were tested forBetween March 22, 2017, and May 15, 2018, 861 participants were enrolled; Among 851 participants included in the analysis, 94·5% were asymptomatic and 45·9% sensitised to Mtb. TB prevalence was 2·0% at baseline and incidence 2·3/100 person-years through 15 months follow-up. Study site was associated with baseline Mtb sensitisation (CD4 count and antiretroviral initiation, proxies for immune status and HIV stage, were associated with Mtb sensitisation and TB disease. Inadequate performance of clinical prediction models may reflect predominantly subclinical disease diagnosed in this setting and unmeasured local site factors affecting transmission and progression.The CORTIS-HR study was funded by the BillMelinda Gates Foundation (OPP1151915) and by the Strategic Health Innovation Partnerships Unit of the South African Medical Research Council with funds received from the South African Department of Science and Technology. The regulatory sponsor was the University of Cape Town.

Objectives We evaluated longitudinal kinetics of an 11-gene blood transcriptomic tuberculosis (TB) signature, RISK11, and effects of TB preventative therapy (TPT) and respiratory organisms on RISK1...

We aimed to understand host factors that affect discriminatory performance of a transcriptomic signature of tuberculosis risk (RISK11).HIV-negative adults aged 18-60 years were evaluated in a prospective study of RISK11 and surveilled for tuberculosis through 15 months. Generalised linear models and receiver-operating characteristic (ROC) regression were used to estimate effect of host factors on RISK11 score (%marginal effect) and on discriminatory performance for tuberculosis disease (area under the curve, AUC), respectively.Among 2923 participants including 74 prevalent and 56 incident tuberculosis cases, percentage marginal effects on RISK11 score were increased among those with prevalent tuberculosis (+18·90%, 95%CI 12·66-25·13), night sweats (+14·65%, 95%CI 5·39-23·91), incident tuberculosis (+7·29%, 95%CI 1·46-13·11), flu-like symptoms (+5·13%, 95%CI 1·58-8·68), and smoking history (+2·41%, 95%CI 0·89-3·93) than those without; and reduced in males (-6·68%, 95%CI -8·31- -5·04) and with every unit increase in BMI (-0·13%, 95%CI -0·25- -0·01). Adjustment for host factors affecting controls did not change RISK11 discriminatory performance. Cough was associated with 72·55% higher RISK11 score in prevalent tuberculosis cases. Stratification by cough improved diagnostic performance from AUC = 0·74 (95%CI 0·67-0·82) overall, to 0·97 (95%CI 0·90-1·00, p 0·001) in cough-positive participants. Combining host factors with RISK11 improved prognostic performance, compared to RISK11 alone, (AUC = 0·76, 95%CI 0·69-0·83 versus 0·56, 95%CI 0·46-0·68, p 0·001) over a 15-month predictive horizon.Several host factors affected RISK11 score, but only adjustment for cough affected diagnostic performance. Combining host factors with RISK11 should be considered to improve prognostic performance.Bill and Melinda Gates Foundation, South African Medical Research Council.

We tested performance of host-blood transcriptomic tuberculosis (TB) signatures for active case-finding. Among 20,207 HIV-uninfected and 963 HIV-infected adults screened; 2,923 and 861 were enrolled from five South African communities. Eight signatures were measured by microfluidic RT-qPCR and participants were microbiologically-investigated for pulmonary TB at baseline, and actively surveilled for incident disease through 15 months. Diagnostic AUCs for 61 HIV-uninfected (weighted-prevalence 1.1%) and 10 HIV-infected (prevalence 1.2%) prevalent TB cases for the 8 signatures were 0.63–0.79 and 0.65–0.88, respectively. Thereafter, 24 HIV-uninfected and 9 HIV-infected participants progressed to incident TB (1.1 and 1.0 per 100 person-years, respectively). Prognostic AUCs through 15-months follow-up were 0.49–0.66 and 0.54–0.81, respectively. Prognostic performance for incident TB occurring within 6-12 months in HIV-negative participants was higher for all signatures. None of the signatures met WHO Target Product Profile criteria for a triage test to diagnose asymptomatic TB; most signatures met the criteria for symptomatic TB. Prognostic accuracy of most signatures for incident TB within six months of testing met the criteria for an incipient TB test.

Global tuberculosis (TB) control requires effective vaccines in TB-endemic countries, where most adults are infected with Mycobacterium tuberculosis (M.tb).We sought to define optimal dose and schedule of H56:IC31, an experimental TB vaccine comprising Ag85B, ESAT-6, and Rv2660c, for M.tb-infected and M.tb-uninfected adults.We enrolled 98 healthy, HIV-uninfected, bacillus Calmette-Guérin-vaccinated, South African adults. M.tb infection was defined by QuantiFERON-TB (QFT) assay. QFT-negative participants received two vaccinations of different concentrations of H56 in 500 nmol of IC31 to enable dose selection for further vaccine development. Subsequently, QFT-positive and QFT-negative participants were randomized to receive two or three vaccinations to compare potential schedules. Participants were followed for safety and immunogenicity for 292 days.H56:IC31 showed acceptable reactogenicity profiles irrespective of dose, number of vaccinations, or M.tb infection. No vaccine-related severe or serious adverse events were observed. The three H56 concentrations tested induced equivalent frequencies and functional profiles of antigen-specific CD4 T cells. ESAT-6 was only immunogenic in QFT-negative participants who received three vaccinations.Two or three H56:IC31 vaccinations at the lowest dose induced durable antigen-specific CD4 T-cell responses with acceptable safety and tolerability profiles in M.tb-infected and M.tb-uninfected adults. Additional studies should validate applicability of vaccine doses and regimens to both QFT-positive and QFT-negative individuals. Clinical trial registered with www.clinicaltrials.gov (NCT01865487).

SETTING South Africa. OBJECTIVE 1) To measure changes in the adolescent prevalence of latent tuberculous infection (LTBI) between 2005 and 2015, and 2) to evaluate medium-term impact of TB control measures on LTBI prevalence. DESIGN We compared baseline data from a cohort study (2005-2007) and a vaccine trial (2014-2015) which enrolled adolescents from the same eight South African high schools. LTBI was defined based on QuantiFERON®-TB Gold In-Tube test positivity. RESULTS We analysed data from 4880 adolescents between 2005 and 2007, and 1968 adolescents between 2014 and 2015, when the average LTBI prevalence was respectively 43.8% (95%CI 28.4-59.1) vs. 48.5% (95%CI 41.1-55.8). Age-specific LTBI prevalence increased between the ages 12 and 18 years by 13% only in lower socio-economic quintile schools, where the average LTBI prevalence was unchanged between the two periods (54% vs. 53%). In the highest socio-economic quintile schools, LTBI prevalence did not increase with age; however, the average LTBI prevalence increased from 20% to 38% between the two periods. CONCLUSION Adolescent LTBI prevalence remained high and constant over a decade, suggesting that Mycobacterium tuberculosis transmission to children was not impacted in the medium term by effective TB control efforts. Trends in adolescent LTBI prevalence should be interpreted in the context of the sociodemographic factors that affect the risk of transmission before and during adolescence. .

Summary Background Targeted preventive therapy for individuals at highest risk of incident tuberculosis might impact the epidemic by interrupting transmission. We tested performance of a transcriptomic signature of tuberculosis (RISK11) and efficacy of signature-guided preventive therapy in parallel, using a hybrid three-group study design. Methods Adult volunteers aged 18–59 years were recruited at five geographically distinct communities in South Africa. Whole blood was sampled for RISK11 by quantitative RT-PCR assay from eligible volunteers without HIV, recent previous tuberculosis (ie

Background Adolescents in the Western Cape Province of South Africa had high force of Mycobacterium tuberculosis (MTB) infection (14% per annum) and high TB incidence (710 per 100,000 person-years) in 2005. We describe subsequent temporal changes in adolescent TB disease notification rates for the decade 2005-2015. Method We conducted an analysis of patient-level adolescent (age 10-19 years) TB disease data, obtained from an electronic TB register in the Breede Valley sub-district, Western Cape Province, South Africa, for 2005-2015. Numerators were annual TB notifications (HIV-related and HIV-unrelated); denominators were mid-year population estimates. Period averages of TB rates were obtained using time series modeling. Temporal trends in TB rates were explored using the Mann-Kendall test. Findings The average adolescent TB disease notification rate was 477 per 100,000 for all TB patients (all-TB) and 361 per 100,000 for microbiologically-confirmed patients. The adolescent all-TB rate declined by 45% from 662 to 361 per 100,000 and the microbiologically-confirmed TB rate by 38% from 492 to 305 per 100,000 between 2005-2015, driven mainly by rapid decreases for the period 2005-2009. There was a statistically significant negative temporal trend in both all-TB (per 100,000) (declined by 48%; from 662 to 343; p = 0·028) and microbiologically confirmed TB (per 100,000) (declined by 49%; from 492 to 252; p = 0·027) for 2005-2009, which was not observed for the period 2009-2015 (rose 5%; from 343 to 361; p = 0·764 and rose 21%; from 252 to 305; p = 1·000, respectively). Interpretation We observed an encouraging fall in adolescent TB disease rates between 2005-2009 with a subsequent plateau during 2010-2015, suggesting that additional interventions are needed to sustain initial advances in TB control.

Introduction Respiratory pathogens such as influenza viruses may play an influential role in the pathogenesis of TB by negatively affecting immunity against Mycobacterium tuberculosis (MTB) We discovered and validated a transcriptomic signature of risk (SOR), based on mRNA expression of 11 IFN stimulated genes (ISG), which prospectively differentiates between incident TB cases and healthy controls The SOR score is computed from expression abundance of multiple ISG transcript pairs in peripheral blood, whereby each pair functions as a ?vote? for or against TB risk We aimed to identify respiratory pathogens other than MTB that might also associate with this SOR score and test whether the SOR score differentiates between individuals with and without respiratory pathogens Methods We conducted a nested cross-sectional study of the upper respiratory tract microbiome Upon consent, participants were consecutively enrolled into the study and provided one nasopharyngeal, one oropharyngeal and a PAXgene blood sample Host blood SOR scores were computed from Ct values for each of the 11 genes, measured by microfluidic qRT-PCR We used multiplex real-time PCR to detect 33 pathogens including bacteria, viruses and fungi in the nasopharyngeal and oropharyngeal samples Multivariate linear regression was used to identify pathogens associated with, and estimate their effect on, the SOR score Wilcoxon rank sum tests and receiver-operating-characteristic (ROC) curves were used to differentiate participants with and without respiratory pathogens Results 1,000 HIV-negative volunteers aged between 18 and 60 years were enrolled 13 viral and nine bacterial pathogens were detected Overall prevalence of respiratory pathogens was 43%: 4% were viruses only, 35 8% bacteria only, and 3 2% were a combination of viruses and bacteria Influenza C, rhinoviruses, coronavirus OC43, adenoviruses, and mycoplasma pneumoniae were significantly associated (Table 1) with a high SOR score In ROC curve analysis the SOR score differentiated participants as follows;virus vs no-pathogen (area under the curve;AUC) AUC=0 72, 95% CI: 0 63-0 81, virus vs bacteria AUC=0 72, 95% CI: 0 63-0 81, bacteria vs no-pathogen AUC=0 51, 95% CI: 0 48-0 55 (no difference) Participants with either viruses only or both viruses and bacteria had significantly higher (P=0 001) SOR scores (median 47% and 43%, respectively) compared to participants without pathogens (median 14%) or participants with bacteria only (median 13%) Conclusion Participants with upper respiratory tract viral colonization or infection had elevated SOR scores, suggesting induction of interferon signalling Infection or colonization with respiratory viruses is likely to result in false positive results for other transcriptomic signatures of tuberculosis based on ISGs (Table Presented)

Background: We tested diagnostic and prognostic performance of a host blood transcriptomic signature of tuberculosis (RISK11) for screening of people living with HIV (PLHIV) in a prospective, community-based cohort. Methods: Ambulant adult volunteers living with HIV were enrolled at five South African sites. RISK11 status was assessed at baseline by real-time PCR. Participants and study staff were blinded to the result. Participants underwent active surveillance for microbiologically-confirmed tuberculosis from enrolment through 15 months. The primary outcomes were prevalence and cumulative incidence of two-sputum-sample positive tuberculosis in RISK11+ versus RISK11- participants. Findings: Among 820 participants with valid RISK11 results, eight (1%) tuberculosis cases were identified at baseline. Risk of prevalent tuberculosis was 13·1-fold (95%CI 2·1-81·6) greater in RISK11+ than RISK11- participants, with tuberculosis prevalence of 2·5% and 0·2%, respectively. RISK11 had diagnostic area under the receiver-operating-characteristic curve (AUC) 88·2%; sensitivity 87·5% and specificity 65·8% at the pre-defined threshold (60%).Thereafter, eight tuberculosis cases were identified through median 15 months follow-up. Tuberculosis incidence was 2·5 vs 0·2 per 100 person-years in RISK11+ compared to RISK11- participants with cumulative incidence ratio 16·0 (95%CI 2·0-129·5); AUC 80·0%; sensitivity 88·6% and specificity 68·9%. By comparison, QuantiFERON TB Gold-Plus (QFT) had a cumulative incidence ratio of 2·0 (95%CI 0·5-8·4); AUC 70·8%; sensitivity 62·1% and specificity 56·2%. Interpretation: RISK11 identified prevalent tuberculosis and predicted risk of progression to incident tuberculosis within 15 months in ambulant PLHIV. Performance approached the World Health Organization Target Product Profile benchmarks for screening and prognostic tests for tuberculosis. QFT performance fell short of the prognostic benchmark. Funding Statement: The study was funded by the Bill & Melinda Gates Foundation (BMGF) (OPP1151915) and by the Strategic Health Innovation Partnerships Unit of the South African Medical Research Council with funds received from the South African Department of Science and Technology. The BMGF contributed to the study design. The regulatory sponsor was the University of Cape Town. Declaration of Interests: AP-N, GW, GC, TJS, and MH report grants from the Bill & Melinda Gates Foundation, during the conduct of the study; AP-N and GW report grants from the South African Medical Research Council, during the conduct of the study; GW and TJS report grants from the South African National Research Foundation, during the conduct of the study. In addition, AP-N and TJS have patents of the RISK11 and RISK6 signatures pending; GW has a patent “TB diagnostic markers” (PCT/IB2013/054377) issued and a patent “Method for diagnosing TB” (PCT/IB2017/052142) pending. The remaining authors have no competing interests to declare. Ethics Approval Statement: The study protocol (Supplement) was approved by the Institutional Human Ethics Committees of each participating site. All participants provided written informed consent in their language of choice.

IntroductionCurrent tuberculosis triage and predictive tools offer poor accuracy and are ineffective for detecting asymptomatic disease in people living with HIV (PLHIV). Host tuberculosis transcriptomic biomarkers hold promise for diagnosing prevalent and predicting progression to incident tuberculosis and guiding further investigation, preventive therapy and follow-up. We aim to conduct a systematic review of performance of transcriptomic signatures of tuberculosis in PLHIV.Methods and analysisWe will search MEDLINE (PubMed), WOS Core Collection, Biological Abstracts, and SciELO Citation Index (Web of Science), Africa-Wide Information and General Science Abstracts (EBSCOhost), Scopus, and Cochrane Central Register of Controlled Trials databases for articles published in English between 1990 and 2020. Case–control, cross-sectional, cohort and randomised controlled studies evaluating performance of diagnostic and prognostic host-response transcriptomic signatures in PLHIV of all ages and settings will be included. Eligible studies will include PLHIV in signature test or validation cohorts, and use microbiological, clinical, or composite reference standards for pulmonary or extrapulmonary tuberculosis diagnosis. Study quality will be evaluated using the ‘Quality Assessment of Diagnostic Accuracy Studies-2’ tool and cumulative review evidence assessed using the ‘Grading of Recommendations Assessment, Development and Evaluation’ approach. Study selection, quality appraisal and data extraction will be performed independently by two reviewers. Study, cohort and signature characteristics of included studies will be tabulated, and a narrative synthesis of findings presented. Primary outcomes of interest, biomarker sensitivity and specificity with estimate precision, will be summarised in forest plots. Expected heterogeneity in signature characteristics, study settings, and study designs precludes meta-analysis and pooling of results. Review reporting will follow the Preferred Reporting Items for Systematic Reviews and Meta-Analyses of Diagnostic Test Accuracy Studies guidelines.Ethics and disseminationFormal ethics approval is not required as primary human participant data will not be collected. Results will be disseminated through peer-reviewed publication and conference presentation.PROSPERO registration numberCRD42021224155.

Summary Background A therapeutic vaccine that prevents recurrent tuberculosis would be a major advance in the development of shorter treatment regimens. We aimed to assess the safety and immunogenicity of the ID93 + GLA-SE vaccine at various doses and injection schedules in patients with previously treated tuberculosis. Methods This randomised, double-blind, placebo-controlled, phase 2a trial was conducted at three clinical sites near Cape Town, South Africa. Patients were recruited at local clinics after receiving 4 months of tuberculosis treatment, and screened for eligibility after providing written informed consent. Participants were aged 18–60 years, BCG-vaccinated, HIV-uninfected, and diagnosed with drug-sensitive pulmonary tuberculosis. Eligible patients had completed standard treatment for pulmonary tuberculosis in the past 28 days. Participants were enrolled after completing standard treatment and randomly assigned sequentially to receive vaccine or placebo in three cohorts: 2 μg intramuscular ID93 + 2 μg GLA-SE on days 0 and 56 (cohort 1); 10 μg ID93 + 2 μg GLA-SE on days 0 and 56 (cohort 2); 2 μg ID93 + 5 μg GLA-SE on days 0 and 56 and placebo on day 28 (cohort 3); 2 μg ID93 + 5 μg GLA-SE on days 0, 28, and 56 (cohort 3); or placebo on days 0 and 56 (cohorts 1 and 2), with the placebo group for cohort 3 receiving an additional injection on day 28. Randomisation was in a ratio of 3:1 for ID93 + GLA-SE and saline placebo in cohorts 1 and 2, and in a ratio of 3:3:1 for (2 ×) ID93 + GLA-SE, (3 ×) ID93 + GLA-SE, and placebo in cohort 3. The primary outcomes were safety and immunogenicity (vaccine-specific antibody response and T-cell response). For the safety outcome, participants were observed for 30 min after each injection, injection site reactions and systemic adverse events were monitored until day 84, and serious adverse events and adverse events of special interest were monitored for 6 months after the last injection. Vaccine-specific antibody responses were measured by serum ELISA, and T-cell responses after stimulation with vaccine antigens were measured in cryopreserved peripheral blood mononuclear cells specimens using intracellular cytokine staining followed by flow cytometry. This study is registered with ClinicalTrials.gov , number NCT02465216 . Findings Between June 17, 2015, and May 30, 2016, we assessed 177 patients for inclusion. 61 eligible patients were randomly assigned to receive: saline placebo (n=5) or (2 ×) 2 μg ID93 + 2 μg GLA-SE (n=15) on days 0 and 56 (cohort 1); saline placebo (n=2) or (2 ×) 10 μg ID93 + 2 μg GLA-SE (n=5) on days 0 and 56 (cohort 2); saline placebo (n=5) on days 0, 28 and 56, or 2 μg ID93 + 5 μg GLA-SE (n=15) on days 0 and 56 and placebo injection on day 28, or (3 ×) 2 μg ID93 + 5 μg GLA-SE (n=14) on days 0, 28, and 56 (cohort 3). ID93 + GLA-SE induced robust and durable antibody responses and specific, polyfunctional CD4 T-cell responses to vaccine antigens. Two injections of the 2 μg ID93 + 5 μg GLA-SE dose induced antigen-specific IgG and CD4 T-cell responses that were significantly higher than those with placebo and persisted for the 6-month study duration. Mild to moderate injection site pain was reported after vaccination across all dose combinations, and induration and erythema in patients given 2 μg ID93 + 5 μg GLA-SE in two or three doses. One participant had grade 3 erythema and induration at the injection site. No vaccine-related serious adverse events were observed. Interpretation Vaccination with ID93 + GLA-SE was safe and immunogenic for all tested regimens. These data support further evaluation of ID93 + GLA-SE in therapeutic vaccination strategies to improve tuberculosis treatment outcomes. Funding Wellcome Trust (102028/Z/13/Z).

Background: South African adolescents have a high force of Mycobacterium tuberculosis (MTB) infection, exceeding 10% per annum, which acts as a barometer of tuberculosis (TB) transmission. We described temporal changes in adolescent TB notification rates to evaluate impact of TB and HIV control measures and inform public health strategy. Method: We conducted a cross-sectional analysis of patient-level TB case data obtained from a regional health authority in Western Cape Province, South Africa, for the decade 2005–2015. Numerators were annual TB notifications (HIV-related and HIV-unrelated); denominators were mid-year population estimates from the national census, adjusted for population growth. Period averages of TB rates were obtained using Auto-Regressive Integrated Moving Average (ARIMA) time series modeling. The Agresti Coull method was used to derive confidence intervals for TB rates. Temporal trends in TB rates were explored using the Mann-Kendall test. Findings: The average adolescent TB notification rate was 477 (95% Confidence Interval (CI): 313–641) per 100,000 for all cases (all-TB) and 361 (CI: 280–441) per 100,000 for microbiologically confirmed cases. However, the adolescent all-TB rate fell by 45% from 662 to 361 per 100,000 (p=0·005) and the microbiologically-confirmed TB rate by 38% from 492 to 305 per 100,000 (p=0·008) between 2005-2015, driven mainly by rapid decreases for the period 2005–2009 (p=0·028 and p=0·027, respectively). Between 2005–2009, 21% (156/732) of adolescent TB notifications were HIV tested, 12% (19/156) of whom were HIV-positive. In the general population, the proportion of HIV-positive persons receiving antiretroviral therapy (ART) increased from

IntroductionOne-quarter of the global population, including the majority of adults in tuberculosis (TB) endemic countries, are estimated to beMycobacterium tuberculosis(MTB) infected. An estimated 10 million new TB cases occurred in 2017. One of the biggest challenges confronting TB control is the lack of accurate diagnosis and prediction of prevalent and incident TB disease, respectively. Several host blood transcriptomic messenger RNA (mRNA) signatures that reflect the host immune response following infection with MTB and progression to TB disease in different study populations have recently been published, but these TB biomarkers have not been systematically described. We will conduct a systematic review of the performance of host blood transcriptional signatures for TB diagnosis and prediction of progression to TB disease.Methods and analysisThis systematic review will involve conducting a comprehensive literature search of cohort, case–control, cross-sectional and randomised-controlled studies of the performance of host blood transcriptomic signatures for TB diagnosis and prediction of progression to TB disease. We will search Medline via PubMed, Scopus, Web of Science and EBSCO libraries, complemented by a search of bibliographies of selected articles for other relevant articles. The literature search will be restricted to studies published in English from 2005 to 2018 and conducted in HIV-uninfected adults and adolescents (≥12 years old). Forest plots and a narrative synthesis of the findings will be provided. The primary outcomes will be sensitivity, specificity, as well as true/false positives and true/false negatives. Heterogeneity resulting from differences in the design, composition and structure of individual signatures will preclude meta-analysis and pooling of results.Ethics and disseminationEthics approval is not required for this systematic review protocol. The results of this review will be disseminated through a peer-reviewed journal as well as conference presentations.PROSPERO registration numberCRD42017073817.

Tuberculosis (TB) is caused byMycobacterium tuberculosis(Mtb) infection and is a major public health problem with an estimated 1.7 billion persons infected worldwide. Clinical challenges in TB include the lack of a blood-based test for active disease, and the absence of prognostic biomarkers for early treatment response. Current blood based tests, such as QuantiFERON-TB Gold (QFT), are based on an IFNγ readout followingMtbantigen stimulation. However, they do not distinguish active TB disease from asymptomaticMtbinfection. We hypothesized that the use of TruCulture, an improved immunomonitoring method for whole blood collection and immune stimulation, could improve the discrimination of active disease from latentMtbinfection. To test our hypothesis, we stimulated whole blood from active TB patients (before and after successful treatment), comparing them to asymptomatic latently infected individuals.Mtb-specific antigens (ESAT-6, CFP-10, TB7.7) and live bacillus Calmette-Guerin (BCG) were used for TruCulture stimulation conditions, with direct comparison to QFT. Protein analyses were performed on the culture supernatants using ELISA and Luminex multi-analyte profiling. TruCulture showed an ability to discriminate active TB cases from latent controls (p < 0.0001, AUC = 0.81, 95% CI: 0.69-0.93) as compared to QFT (p = 0.47 AUC = 0.56, 95% CI: 0.40-0.72), based on an IFNγ readout afterMtbantigen stimulation. The stratification of the two groups could be further improved by using theMtbAg/BCG IFNγ ratio response (p < 0.0001, AUC = 0.918, 95% CI: 0.84-0.98). We also identified additional cytokines that distinguished latent infection from TB disease; and show that the primary differences between the TruCulture and QFT systems were a result of higher levels of non-specific innate immune activation in QFT tubes, due to the lack of a buffering solution in the latter. We conclude that TruCulture offers a next-generation solution for whole blood stimulation and immunomonitoring with the possibility to discriminate active and latently infected persons.

Summary Background Infants are a key target population for new tuberculosis vaccines. We assessed the safety and immunogenicity of the live-attenuated Mycobacterium tuberculosis vaccine candidate MTBVAC in adults and infants in a region where transmission of tuberculosis is very high. Methods We did a randomised, double-blind, BCG-controlled, dose-escalation trial at the South African Tuberculosis Vaccine Initiative site near Cape Town, South Africa. Healthy adult community volunteers who were aged 18–50 years, had received BCG vaccination as infants, were HIV negative, had negative interferon-γ release assay (IGRA) results, and had no personal history of tuberculosis or current household contact with someone with tuberculosis were enrolled in a safety cohort. Infants born to HIV-negative women with no personal history of tuberculosis or current household contact with a person with tuberculosis and who were 96 h old or younger, generally healthy, and had not yet received routine BCG vaccination were enrolled in a separate infant cohort. Eligible adults were randomly assigned (1:1) to receive either BCG Vaccine SSI (5 × 105 colony forming units [CFU] of Danish strain 1331 in 0·1 mL diluent) or MTBVAC (5 × 105 CFU in 0·1 mL) intradermally in the deltoid region of the arm. After favourable review of 28-day reactogenicity and safety data in the adult cohort, infants were randomly assigned (1:3) to receive either BCG Vaccine SSI (2·5 × 105 CFU in 0·05 mL diluent) or MTBVAC in three sequential cohorts of increasing MTBVAC dose (2·5 × 103 CFU, 2·5 × 104 CFU, and 2·5 × 105 CFU in 0·05 mL) intradermally in the deltoid region of the arm. QuantiFERON-TB Gold In-Tube IGRA was done on days 180 and 360. For both randomisations, a pre-prepared block randomisation schedule was used. Participants (and their parents or guardians in the case of infant participants), investigators, and other clinical and laboratory staff were masked to intervention allocation. The primary outcomes, which were all measured in the infant cohort, were solicited and unsolicited local adverse events and serious adverse events until day 360; non-serious systemic adverse events until day 28 and vaccine-specific CD4 and CD8 T-cell responses on days 7, 28, 70, 180, and 360. Secondary outcomes measured in adults were local injection-site and systemic reactions and haematology and biochemistry at study day 7 and 28. Safety analyses and immunogenicity analyses were done in all participants who received a dose of vaccine. This trial is registered with ClinicalTrials.gov, number NCT02729571. Findings Between Sept 29, 2015, and Nov 16, 2015, 62 adults were screened and 18 were enrolled and randomly assigned, nine each to the BCG and MTBVAC groups. Between Feb 12, 2016, and Sept 21, 2016, 36 infants were randomly assigned—eight to the BCG group, nine to the 2·5 × 103 CFU MTBVAC group, nine to the 2·5 × 104 CFU group, and ten to the 2·5 × 105 CFU group. Mild injection-site reactions occurred only in infants in the BCG and the 2·5 × 105 CFU MTBVAC group, with no evidence of local or regional injection-site complications. Systemic adverse events were evenly distributed across BCG and MTBVAC dose groups, and were mostly mild in severity. Eight serious adverse events were reported in seven vaccine recipients (one adult MTBVAC recipient, one infant BCG recipient, one infant in the 2·5 × 103 CFU MTBVAC group, two in the 2·5 × 104 CFU MTBVAC group, and two in the 2·5 × 105 CFU MTBVAC group), including one infant in the 2·5 × 103 CFU MTBVAC group treated for unconfirmed tuberculosis and one in the 2·5 × 105 CFU MTBVAC group treated for unlikely tuberculosis. One infant died as a result of possible viral pneumonia. Vaccination with all MTBVAC doses induced durable antigen-specific T-helper-1 cytokine-expressing CD4 cell responses in infants that peaked 70 days after vaccination and were detectable 360 days after vaccination. For the highest MTBVAC dose (ie, 2·5 × 105 CFU), these responses exceeded responses induced by an equivalent dose of the BCG vaccine up to 360 days after vaccination. Dose-related IGRA conversion was noted in three (38%) of eight infants in the 2·5 × 103 CFU MTBVAC group, six (75%) of eight in the 2·5 × 104 CFU MTBVAC group, and seven (78%) of nine in the 2·5 × 105 CFU MTBVAC group at day 180, compared with none of seven infants in the BCG group. By day 360, IGRA reversion had occurred in all three infants (100%) in the 2·5 × 103 CFU MTBVAC group, four (67%) of the six in the 2·5 × 104 CFU MTBVAC group, and three (43%) of the seven in the 2·5 × 105 CFU MTBVAC group. Interpretation MTBVAC had acceptable reactogenicity, and induced a durable CD4 cell response in infants. The evidence of immunogenicity supports progression of MTBVAC into larger safety and efficacy trials, but also confounds interpretation of tests for M tuberculosis infection, highlighting the need for stringent endpoint definition. Funding Norwegian Agency for Development Cooperation, TuBerculosis Vaccine Initiative, UK Department for International Development, and Biofabri.

IntroductionHost blood transcriptomic biomarkers have potential as rapid point-of-care triage, diagnostic, and predictive tests for Tuberculosis disease. We aimed to summarise the performance of host blood transcriptomic signatures for diagnosis of and prediction of progression to Tuberculosis disease; and compare their performance to the recommended World Health Organisation target product profile.MethodsA systematic review and meta-analysis of the performance of host blood mRNA signatures for diagnosing and predicting progression to Tuberculosis disease in HIV-negative adults and adolescents, in studies with an independent validation cohort. Medline, Scopus, Web of Science, and EBSCO libraries were searched for articles published between January 2005 and May 2019, complemented by a search of bibliographies. Study selection, data extraction and quality assessment were done independently by two reviewers. Meta-analysis was performed for signatures that were validated in ≥3 comparable cohorts, using a bivariate random effects model.ResultsTwenty studies evaluating 25 signatures for diagnosis of or prediction of progression to TB disease in a total of 68 cohorts were included. Eighteen studies evaluated 24 signatures for TB diagnosis and 17 signatures met at least one TPP minimum performance criterion. Three diagnostic signatures were validated in clinically relevant cohorts to differentiate TB from other diseases, with pooled sensitivity 84%, 87% and 90% and pooled specificity 79%, 88% and 74%, respectively. Four studies evaluated signatures for progression to TB disease and performance of one signature, assessed within six months of TB diagnosis, met the minimal TPP for a predictive test for progression to TB disease.ConclusionHost blood mRNA signatures hold promise as triage tests for TB. Further optimisation is needed if mRNA signatures are to be used as standalone diagnostic or predictive tests for therapeutic decision-making.

Recent Mycobacterium tuberculosis infection confers a predisposition to the development of tuberculosis disease, the leading killer among global infectious diseases. H4:IC31, a candidate subunit vaccine, has shown protection against tuberculosis disease in preclinical models, and observational studies have indicated that primary bacille Calmette-Guérin (BCG) vaccination may offer partial protection against infection.In this phase 2 trial, we randomly assigned 990 adolescents in a high-risk setting who had undergone neonatal BCG vaccination to receive the H4:IC31 vaccine, BCG revaccination, or placebo. All the participants had negative results on testing for M. tuberculosis infection on the QuantiFERON-TB Gold In-tube assay (QFT) and for the human immunodeficiency virus. The primary outcomes were safety and acquisition of M. tuberculosis infection, as defined by initial conversion on QFT that was performed every 6 months during a 2-year period. Secondary outcomes were immunogenicity and sustained QFT conversion to a positive test without reversion to negative status at 3 months and 6 months after conversion. Estimates of vaccine efficacy are based on hazard ratios from Cox regression models and compare each vaccine with placebo.Both the BCG and H4:IC31 vaccines were immunogenic. QFT conversion occurred in 44 of 308 participants (14.3%) in the H4:IC31 group and in 41 of 312 participants (13.1%) in the BCG group, as compared with 49 of 310 participants (15.8%) in the placebo group; the rate of sustained conversion was 8.1% in the H4:IC31 group and 6.7% in the BCG group, as compared with 11.6% in the placebo group. Neither the H4:IC31 vaccine nor the BCG vaccine prevented initial QFT conversion, with efficacy point estimates of 9.4% (P=0.63) and 20.1% (P=0.29), respectively. However, the BCG vaccine reduced the rate of sustained QFT conversion, with an efficacy of 45.4% (P=0.03); the efficacy of the H4:IC31 vaccine was 30.5% (P=0.16). There were no clinically significant between-group differences in the rates of serious adverse events, although mild-to-moderate injection-site reactions were more common with BCG revaccination.In this trial, the rate of sustained QFT conversion, which may reflect sustained M. tuberculosis infection, was reduced by vaccination in a high-transmission setting. This finding may inform clinical development of new vaccine candidates. (Funded by Aeras and others; C-040-404 ClinicalTrials.gov number, NCT02075203 .).

Rationale: Contacts of patients with tuberculosis (TB) constitute an important target population for preventive measures because they are at high risk of infection with Mycobacterium tuberculosis and progression to disease. Objectives: We investigated bio-signatures with predictive ability for incident TB. Methods: In a case-control study nested within the Grand Challenges 6-74 longitudinal HIV-negative African cohort of exposed household contacts, we employed RNA sequencing, PCR, and the pair ratio algorithm in a training/test set approach. Overall, 79 progressors who developed TB between 3 and 24 months after diagnosis of index case and 328 matched nonprogressors who remained healthy during 24 months of follow-up were investigated. Measurements and Main Results: A four-transcript signature derived from samples in a South African and Gambian training set predicted progression up to two years before onset of disease in blinded test set samples from South Africa, the Gambia, and Ethiopia with little population-associated variability, and it was also validated in an external cohort of South African adolescents with latent M. tuberculosis infection. By contrast, published diagnostic or prognostic TB signatures were predicted in samples from some but not all three countries, indicating site-specific variability. Post hoc meta-analysis identified a single gene pair, C1QC/TRAV27 (complement C1q C-chain / T-cell receptor-a variable gene 27) that would consistently predict TB progression in household contacts from multiple African sites but not in infected adolescents without known recent exposure events. Conclusions: Collectively, we developed a simple whole blood-based PCR test to predict TB in recently exposed household contacts from diverse African populations. This test has potential for implementation in national TB contact investigation programs.

Background. The Xpert MTB/RIF test shortens the time to microbiological confirmation of pulmonary tuberculosis (TB) under research conditions. Objective. To evaluate the field impact of Xpert MTB/RIF rollout on TB diagnostic yield and time to treatment in a South African (SA) community. Methods. We compared TB investigation outcomes for 6-month calendar periods before and after Xpert MTB/RIF rollout in a semi-rural area of SA. The proportion of adult patients who tested positive by sputum smear microscopy, liquid culture or Xpert MTB/RIF and the proportion of positive sputum smear, liquid culture or Xpert MTB/RIF tests were compared. Secondary outcomes included time to laboratory diagnosis and treatment initiation. Data were collected from the National Health Laboratory Service database and from the Western Cape Provincial Department of Health TB register. Results. Regional rollout of Xpert MTB/RIF testing occurred in 2013. Of the 15 629 patients investigated in the post-rollout period, 7.9% tested positive on GeneXpert, compared with 6.4% of the 10 741 investigated in the pre-rollout period who tested positive by sputum smear microscopy ( p

Conversion from a negative to positive QuantiFERON-TB test is indicative of Mycobacterium tuberculosis (Mtb) infection, which predisposes individuals to tuberculosis disease. Interpretation of serial tests is confounded by immunological and technical variability.To improve the consistency of serial QuantiFERON-TB testing algorithms and provide a data-driven definition of conversion.Sources of QuantiFERON-TB variability were assessed, and optimal procedures were identified. Distributions of IFN-γ response levels were analyzed in healthy adolescents, Mtb-unexposed control subjects, and patients with pulmonary tuberculosis.Individuals with no known Mtb exposure had IFN-γ values less than 0.2 IU/ml. Among individuals with IFN-γ values less than 0.2 IU/ml, 0.2-0.34 IU/ml, 0.35-0.7 IU/ml, and greater than 0.7 IU/ml, tuberculin skin test positivity results were 15%, 53%, 66%, and 91% (P 0.005), respectively. Together, these findings suggest that values less than 0.2 IU/ml were true negatives. In short-term serial testing, "uncertain" conversions, with at least one value within the uncertainty zone (0.2-0.7 IU/ml), were partly explained by technical assay variability. Individuals who had a change in QuantiFERON-TB IFN-γ values from less than 0.2 to greater than 0.7 IU/ml had 10-fold higher tuberculosis incidence rates than those who maintained values less than 0.2 IU/ml over 2 years (P = 0.0003). By contrast, "uncertain" converters were not at higher risk than nonconverters (P = 0.229). Eighty-seven percent of patients with active tuberculosis had IFN-γ values greater than 0.7 IU/ml, suggesting that these values are consistent with established Mtb infection.Implementation of optimized procedures and a more rigorous QuantiFERON-TB conversion definition (an increase from IFN-γ0.2 to0.7 IU/ml) would allow more definitive detection of recent Mtb infection and potentially improve identification of those more likely to develop disease.

Newborn MVA85A prime vaccination was safe and induced an early immune response that did not interfere with immunogenicity of subsequent bacille Calmette-Guérin vaccination. New tuberculosis vaccine candidates should be tested using this strategy, which appears safe regardless of infant human immunodeficiency virus exposure., Background Vaccination of human immunodeficiency virus (HIV)-infected infants with bacille Calmette-Guérin (BCG) is contraindicated. HIV-exposed newborns need a new tuberculosis vaccination strategy that protects against tuberculosis early in life and avoids the potential risk of BCG disease until after HIV infection has been excluded. Methods This double-blind, randomized, controlled trial compared newborn MVA85A prime vaccination (1 × 108 PFU) vs Candin® control, followed by selective, deferred BCG vaccination at age 8 weeks for HIV-uninfected infants and 12 months follow-up for safety and immunogenicity. Results A total of 248 HIV-exposed infants were enrolled. More frequent mild–moderate reactogenicity events were seen after newborn MVA85A vaccination. However, no significant difference was observed in the rate of severe or serious adverse events, HIV acquisition (n = 1 per arm), or incident tuberculosis disease (n = 5 MVA85A; n = 3 control) compared to the control arm. MVA85A vaccination induced modest but significantly higher Ag85A-specific interferon gamma (IFNγ)+ CD4+ T cells compared to control at weeks 4 and 8 (P < .0001). BCG did not further boost this response in MVA85A vaccinees. The BCG-induced Ag85A-specific IFNγ+ CD4+ T-cell response at weeks 16 and 52 was of similar magnitude in the control arm compared to the MVA85A arm at all time points. Proliferative capacity, functional profiles, and memory phenotype of BCG-specific CD4 responses were similar across study arms. Conclusions MVA85A prime vaccination of HIV-exposed newborns was safe and induced an early modest antigen-specific immune response that did not interfere with, or enhance, immunogenicity of subsequent BCG vaccination. New protein-subunit and viral-vectored tuberculosis vaccine candidates should be tested in HIV-exposed newborns. Clinical Trials Registration NCT01650389.

RATIONALE: Novel tuberculosis (TB) vaccines should be safe and effective in populations infected with Mycobacterium tuberculosis (M.tb) and/or HIV for effective TB control. OBJECTIVE: To determine the safety and immunogenicity of MVA85A, a novel TB vaccine, among M.tb- and/or HIV-infected persons in a setting where TB and HIV are endemic. METHODS: An open-label, phase IIa trial was conducted in 48 adults with M.tb and/or HIV infection. Safety and immunogenicity were analyzed up to 52 weeks after intradermal vaccination with 5 × 10(7) plaque-forming units of MVA85A. Specific T-cell responses were characterized by IFN-γ enzyme-linked immunospot and whole blood intracellular cytokine staining assays. MEASUREMENTS AND MAIN RESULTS: MVA85A was well tolerated and no vaccine-related serious adverse events were recorded. MVA85A induced robust and durable response of mostly polyfunctional CD4(+) T cells, coexpressing IFN-γ, tumor necrosis factor-α, and IL-2. Magnitudes of pre- and postvaccination T-cell responses were lower in HIV-infected, compared with HIV-uninfected, vaccinees. No significant effect of antiretroviral therapy on immunogenicity of MVA85A was observed. CONCLUSIONS: MVA85A was safe and immunogenic in persons with HIV and/or M.tb infection. These results support further evaluation of safety and efficacy of this vaccine for prevention of TB in these target populations.

(See the editorial commentary by Dockrell, on pages 1708-9.)Background. BCG, the only licensed tuberculosis vaccine, affords poor protection against lung tuberculosis in infants and children. A new tuberculosis vaccine, which may enhance the BCG-induced immune response, is urgently needed. We assessed the safety of and characterized the T cell response induced by 3 doses of the candidate vaccine, MVA85A, in BCG-vaccinated infants from a setting where tuberculosis is endemic.Methods. Infants aged 5–12 months were vaccinated intradermally with either 2.5 × 10 7 , 5 × 10 7 , or 10 × 10 7 plaque-forming units of MVA85A, or placebo. Adverse events were documented, and T-cell responses were assessed by interferon γ (IFN-γ) enzyme-linked immunospot assay and intracellular cytokine staining.Results. The 3 MVA85A doses were well tolerated, and no vaccine-related serious adverse events were recorded. MVA85A induced potent, durable T-cell responses, which exceeded prevaccination responses up to 168 d after vaccination. No dose-related differences in response magnitude were observed. Multiple CD4 T cell subsets were induced; polyfunctional CD4 T cells co-expressing T-helper cell 1 cytokines with or without granulocyte-macrophage colony-stimulating factor predominated. IFN-γ-expressing CD8 T cells, which peaked later than CD4 T cells, were also detectable.Conclusions. MVA85A was safe and induced robust, polyfunctional, durable CD4 and CD8 T-cell responses in infants. These data support efficacy evaluation of MVA85A to prevent tuberculosis in infancy.Clinical Trials Registration. NCT00679159.

The endpoint definition for infant tuberculosis (TB) vaccine trials should match the TB disease phenotype expected in the control arm of the study population. Our aim was to analyse selected combinations of the clinical, radiological, and microbiological features of pulmonary TB among children investigated under vaccine trial conditions, in order to estimate case frequency for a range of expected TB phenotypes. Two thousand one hundred and eighty five South African children were investigated over a nine-year period (2001-2009). Evidence of TB exposure and classical symptoms were several times more common than chest radiography (CXR) compatible with TB, or positive Mycobacterium tuberculosis culture. Discordance between clinical, radiological, and microbiological features was common in individual children. Up to one third of children with compatible CXR, and up to half the children who were M. tuberculosis culture positive, were asymptomatic. The culture positive rate fell over time, although rates of TB exposure and compatible chest radiography increased. Consequently, the annual incidence of diagnostic combinations that included M. tuberculosis culture fell to0.2%. However, in this study population (children2 years of age), annual incidence of the TB disease phenotype that included the triad of TB exposure, symptoms, and compatible CXR, approached 1% (n=848 per 100,000). These findings allow modelling of expected TB case frequency in multi-centre infant TB vaccine trials, based upon benchmarking of diagnostic data against the key indicator variables that constitute the building blocks of a trial endpoint.

RationaleHSP90B1, also known as gp96, is a chaperone for multiple Toll-like receptors (TLRs) and is necessary for TLR-mediated inflammatory responses in murine myeloid cells. The molecule is also expressed in T-cells though its specific role is unknown. We hypothesized that human HSP90B1 regulates monocyte and T-cell responses to Mycobacterium tuberculosis (Mtb) and bacilli Calmette-Guerin (BCG) and that its variants are associated with susceptibility to TB disease.MethodsWe screened 17 haplotype-tagging SNPs in the HSP90B1 gene region for association with BCG-induced T-cell cytokine responses using both an ex-vivo whole blood assay (N = 295) and an intracellular cytokine staining assay (N = 180) on samples collected 10 weeks after birth. Using a case-control study design, we evaluated the same SNPs for association with TB disease in a South African pediatric cohort (N = 217 cases, 604 controls). A subset of these SNPs was evaluated for association with HSP90B1 expression in human monocytes, monocyte-derived dendritic cells, and T-cells using RT-PCR. Lastly, we used CRISPR/Cas9 gene editing to knock down HSP90B1 expression in a human monocyte cell line (U937). Knockdown and control cell lines were tested for TLR surface expression and control of Mtb replication.ResultsWe identified three SNPs, rs10507172, rs10507173 and rs1920413, that were associated with BCG-induced IL-2 secretion (p = 0.017 for rs10507172 and p = 0.03 for rs10507173 and rs1920413, Mann-Whitney, dominant model). SNPs rs10507172 and rs10507173 were associated with TB disease in an unadjusted analysis (p = 0.036 and 0.025, respectively, dominant model) that strengthened with sensitivity analysis of the definite TB cases, which included only those patients with microbiologically confirmed Mtb (p = 0.007 and 0.012, respectively). Knockdowns of HSP90B1 in monocyte cell lines with CRISPR did not alter TLR2 surface expression nor influence Mtb replication relative to controls.ConclusionAmong infants, an HSP90B1 gene-region variant is associated with BCG-induced IL-2 production and may be associated with protection from TB disease. HSP90B1 knockdown in human monocyte-like cell lines did not influence TLR2 surface localization nor Mtb replication. Together, these data suggest that HSP90B1 regulates T-cell, but not monocyte, responses to mycobacteria in humans.

Background: The risk of developing tuberculosis (TB) disease in HIV-uninfected children after isoniazid preventive therapy (IPT) for a positive QuantiFERON-TB Gold In-Tube test (QFT-GIT) is unknown. The aim of this study was to evaluate risk of TB disease after IPT in young HIV-uninfected children with a positive QFT-GIT result, or household TB contact. Methods: HIV-uninfected South African infants aged 4–6 months were screened for enrolment in a TB vaccine trial. Baseline household TB contact and positive QFT-GIT result were exclusion criteria, and these infants were referred for IPT. Outcome data are reported for 36 months after IPT referral. Results: Four thousand seven hundred forty-nine infants were screened. Household TB contact was reported in 131 (2.8%) infants; 279 (6.0%) were QFT-GIT positive, and 138 of these 410 infants (34.0%) started IPT. Forty-four cases of TB disease (11.0%) were recorded within 991 child years of observation. TB disease incidence was 4.8 versus 3.6 per 100 child years in household exposed versus QFT-GIT-positive children [incidence rate ratio: 1.35; 95% confidence interval (CI): 0.67–2.88] and 2.4 versus 5.5 per 100 child years in children who received versus did not receive IPT, respectively (incidence rate ratio: 0.44; 95% CI: 0.17–0.96). Adjusted hazard ratio (Cox regression) for TB disease was 0.48 (95% CI: 0.21–1.05) for those who received IPT. Conclusion: In young HIV-uninfected children, the effect of IPT on risk of TB disease is similar, whether TB exposure was defined by household contact history or by positive QFT-GIT result. International IPT guidelines for HIV-uninfected children with a positive QFT-GIT result should be updated.

Supplemental Digital Content is available in the text., Background: Childhood tuberculosis (TB) is usually Mycobacterium tuberculosis (MTB) culture negative. Furthermore, clinical presentation may be altered by active case finding, isoniazid prophylaxis and early treatment. We aimed to establish the value of presenting symptoms for intrathoracic TB case diagnosis among young children. Methods: Healthy, HIV-uninfected, South African infants in an efficacy trial of a novel TB vaccine (MVA85A) were followed for 2 years for suspected TB. When suspected, investigation followed a standardized algorithm comprising symptom history, QuantiFERON Gold-in-Tube, chest radiography (CXR), MTB culture and Xpert MTB/RIF from paired gastric lavage and induced sputa. Adjusted odds ratios and 95% confidence intervals describe the associations between symptoms and positive MTB culture or Xpert MTB/RIF, and CXR compatible with intrathoracic TB. Results: Persistent cough was present in 172/1017 (16.9%) of the children investigated for TB. MTB culture/Xpert MTB/RIF was positive in 38/1017 children (3.7%); and CXR was positive, that is, compatible with intrathoracic TB, in 131/1017 children (12.9%). Children with persistent cough had more than triple the odds of a positive MTB culture/Xpert MTB/RIF (adjusted odds ratios: 3.3, 95% confidence interval: 1.5–7.0) and positive CXR (adjusted odds ratios: 3.5, 95% confidence interval: 2.2–5.5). Persistent cough was the only symptom that differentiated children with severe (56.5%) from nonsevere intrathoracic TB disease (28.2%; P = 0.001). Conclusion: Persistent cough was the cardinal diagnostic symptom associated with microbiologic and radiologic evidence, and disease severity, of intrathoracic TB. Symptom-based definitions of TB disease for diagnostic, preventive and therapeutic studies should prioritize persistent cough above other symptoms compatible with childhood TB.

Objective Diagnosis of childhood tuberculosis is limited by the paucibacillary respiratory samples obtained from young children with pulmonary disease. We aimed to compare accuracy of the Xpert® MTB/RIF assay, an automated nucleic acid amplification test, between induced sputum and gastric lavage samples from young children in a tuberculosis endemic setting. Methods We analyzed standardized diagnostic data from HIV negative children younger than four years of age who were investigated for tuberculosis disease near Cape Town, South Africa [2009–2012]. Two paired, consecutive induced sputa and early morning gastric lavage samples were obtained from children with suspected tuberculosis. Samples underwent Mycobacterial Growth Indicator Tube [MGIT] culture and Xpert MTB/RIF assay. We compared diagnostic yield across samples using the two-sample test of proportions and McNemar’s χ2 test; and Wilson’s score method to calculate sensitivity and specificity. Results 1,020 children were evaluated for tuberculosis during 1,214 admission episodes. Not all children had 4 samples collected. 57 of 4,463[1.3%] and 26 of 4,606[0.6%] samples tested positive for Mycobacterium tuberculosis on MGIT culture and Xpert MTB/RIF assay respectively. 27 of 2,198[1.2%] and 40 of 2,183[1.8%] samples tested positive [on either Xpert MTB/RIF assay or MGIT culture] on induced sputum and gastric lavage samples, respectively. 19/1,028[1.8%] and 33/1,017[3.2%] admission episodes yielded a positive MGIT culture or Xpert MTB/RIF assay from induced sputum and gastric lavage, respectively. Sensitivity of Xpert MTB/RIF assay was 8/30[26.7%; 95% CI: 14.2–44.4] for two induced sputum samples and 7/31[22.6%; 11.4–39.8] [p = 0.711] for two gastric lavage samples. Corresponding specificity was 893/893[100%;99.6–100] and 885/890[99.4%;98.7–99.8] respectively [p = 0.025]. Conclusion Sensitivity of Xpert MTB/RIF assay was low, compared to MGIT culture, but diagnostic performance of Xpert MTB/RIF did not differ sufficiently between induced sputum and gastric lavage to justify selection of one sampling method over the other, in young children with suspected pulmonary TB. Trial Registration ClinicalTrials.gov NCT00953927