Abstract 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.
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.
This dataset comprises expression profiles of 43 genes related to MSDC functioning and 5 reference genes in 8 blood samples and 8 brochio-alveolar lavage fluid (BALF) samples from TB patients. Also included are results from blood and BALF of 2 'other lung disease' patients each.
ABSTRACT The development of a functional biomarker assay in the tuberculosis (TB) field would be widely recognized as a major advance in efforts to develop and to test novel TB vaccine candidates efficiently. We present preliminary studies using mycobacterial growth inhibition assays (MGIAs) to detect Mycobacterium bovis BCG vaccine responses across species, and we extend this work to determine whether a standardized MGIA can be applied in characterizing new TB vaccines. The comparative MGIA studies reviewed here aimed to evaluate robustness, reproducibility, and ability to reflect in vivo responses. In doing so, they have laid the foundation for the development of a MGIA that can be standardized and potentially qualified. A major challenge ahead lies in better understanding the relationships between in vivo protection, in vitro growth inhibition, and the immune mechanisms involved. The final outcome would be a MGIA that could be used with confidence in TB vaccine trials. We summarize data arising from this project, present a strategy to meet the goals of developing a functional assay for TB vaccine testing, and describe some of the challenges encountered in performing and transferring such assays.
AERAS-402 is a novel tuberculosis vaccine designed to boost immunity primed by bacillus Calmette-Guérin (BCG), the only licensed vaccine.We investigated the safety and immunogenicity of AERAS-402 in healthy Mycobacterium tuberculosis-uninfected BCG-vaccinated adults from a tuberculosis-endemic region of South Africa.Escalating doses of AERAS-402 vaccine were administered intramuscularly to each of three groups of healthy South African BCG-vaccinated adults, and a fourth group received two injections of the maximal dose. Participants were monitored for 6 months, with all adverse effects documented. Vaccine-induced CD4(+) and CD8(+) T-cell immunity was characterized by an intracellular cytokine staining assay of whole blood and peripheral blood mononuclear cells.AERAS-402 was well tolerated, and no vaccine-related serious adverse events were recorded. The vaccine induced a robust CD4(+) T-cell response dominated by cells coexpressing IFN-gamma, tumor necrosis factor-alpha, and IL-2 ("polyfunctional" cells). AERAS-402 also induced a potent CD8(+) T-cell response, characterized by cells expressing IFN-gamma and/or tumor necrosis factor-alpha, which persisted for the duration of the study.Vaccination with AERAS-402 is safe and immunogenic in healthy adults. The immunity induced by the vaccine appears promising: polyfunctional T cells are thought to be important for protection against intracellular pathogens such as Mycobacterium tuberculosis, and evidence is accumulating that CD8(+) T cells are also important. AERAS-402 induced a robust and durable CD8(+) T-cell response, which appears extremely promising. Clinical trial registered with www.sanctr.gov.za (NHREC no. 1381).
BackgroundA rapid, blood-based triage test that allows targeted investigation for tuberculosis at the point of care could shorten the time to tuberculosis treatment and reduce mortality. We aimed to test the performance of a host blood transcriptomic signature (RISK11) in diagnosing tuberculosis and predicting progression to active pulmonary disease (prognosis) in people with HIV in a community setting.MethodsIn this prospective diagnostic and prognostic accuracy study, adults (aged 18–59 years) with HIV were recruited from five communities in South Africa. Individuals with a history of tuberculosis or household exposure to multidrug-resistant tuberculosis within the past 3 years, comorbid risk factors for tuberculosis, or any condition that would interfere with the study were excluded. RISK11 status was assessed at baseline by real-time PCR; participants and study staff were masked to the result. Participants underwent active surveillance for microbiologically confirmed tuberculosis by providing spontaneously expectorated sputum samples at baseline, if symptomatic during 15 months of follow-up, and at 15 months (the end of the study). The coprimary outcomes were the prevalence and cumulative incidence of tuberculosis disease confirmed by a positive Xpert MTB/RIF, Xpert Ultra, or Mycobacteria Growth Indicator Tube culture, or a combination of such, on at least two separate sputum samples collected within any 30-day period.FindingsBetween March 22, 2017, and May 15, 2018, 963 participants were assessed for eligibility and 861 were enrolled. Among 820 participants with valid RISK11 results, eight (1%) had prevalent tuberculosis at baseline: seven (2·5%; 95% CI 1·2–5·0) of 285 RISK11-positive participants and one (0·2%; 0·0–1·1) of 535 RISK11-negative participants. The relative risk (RR) of prevalent tuberculosis was 13·1 times (95% CI 2·1–81·6) greater in RISK11-positive participants than in RISK11-negative participants. RISK11 had a diagnostic area under the receiver operating characteristic curve (AUC) of 88·2% (95% CI 77·6–96·7), and a sensitivity of 87·5% (58·3–100·0) and specificity of 65·8% (62·5–69·0) at a predefined score threshold (60%). Of those with RISK11 results, eight had primary endpoint incident tuberculosis during 15 months of follow-up. Tuberculosis incidence was 2·5 per 100 person-years (95% CI 0·7–4·4) in the RISK11-positive group and 0·2 per 100 person-years (0·0–0·5) in the RISK11-negative group. The probability of primary endpoint incident tuberculosis was greater in the RISK11-positive group than in the RISK11-negative group (cumulative incidence ratio 16·0 [95% CI 2·0–129·5]). RISK11 had a prognostic AUC of 80·0% (95% CI 70·6–86·9), and a sensitivity of 88·6% (43·5–98·7) and a specificity of 68·9% (65·3–72·3) for incident tuberculosis at the 60% threshold.InterpretationRISK11 identified prevalent tuberculosis and predicted risk of progression to incident tuberculosis within 15 months in ambulant people living with HIV. RISK11's performance approached, but did not meet, WHO's target product profile benchmarks for screening and prognostic tests for tuberculosis.FundingBill & Melinda Gates Foundation and the South African Medical Research Council.
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. 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. 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. 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. NCT01650389.
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 biosignatures 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-α 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.
Tuberculosis (TB) is a major cause of morbidity and mortality worldwide, thus there is an urgent need for novel TB vaccines.We investigated a novel TB vaccine candidate, M72/AS01, in a phase IIa trial of bacille Calmette-Guérin-vaccinated, HIV-uninfected, and Mycobacterium tuberculosis (Mtb)-infected and -uninfected adults in South Africa.Two doses of M72/AS01 were administered to healthy adults, with and without latent Mtb infection. Participants were monitored for 7 months after the first dose; cytokine production profiles, cell cycling, and regulatory phenotypes of vaccine-induced T cells were measured by flow cytometry.The vaccine had a clinically acceptable safety profile, and induced robust, long-lived M72-specific T-cell and antibody responses. M72-specific CD4 T cells produced multiple combinations of Th1 cytokines. Analysis of T-cell Ki67 expression showed that most vaccination-induced T cells did not express Th1 cytokines or IL-17; these cytokine-negative Ki67(+) T cells included subsets of CD4 T cells with regulatory phenotypes. PD-1, a negative regulator of activated T cells, was transiently expressed on M72-specific CD4 T cells after vaccination. Specific T-cell subsets were present at significantly higher frequencies after vaccination of Mtb-infected versus -uninfected participants.M72/AS01 is clinically well tolerated in Mtb-infected and -uninfected adults, induces high frequencies of multifunctional T cells, and boosts distinct T-cell responses primed by natural Mtb infection. Moreover, these results provide important novel insights into how this immunity may be appropriately regulated after novel TB vaccination of Mtb-infected and -uninfected individuals.Clinical trial registered with www.clinicaltrials.gov (NCT 00600782).
Background Multiple host blood transcriptional signatures have been developed as non-sputum triage tests for tuberculosis (TB). We aimed to compare the diagnostic performance of 20 blood transcriptomic TB signatures for differentiating between symptomatic patients who have TB versus other respiratory diseases (ORD). Methods As part of a nested case–control study, individuals presenting with respiratory symptoms at primary healthcare clinics in Ethiopia, Malawi, Namibia, Uganda, South Africa and The Gambia were enrolled. TB was diagnosed based on clinical, microbiological and radiological findings. Transcriptomic signatures were measured in whole blood using microfluidic real-time quantitative PCR. Diagnostic performance was benchmarked against the World Health Organization Target Product Profile (TPP) for a non-sputum TB triage test. Results Among 579 participants, 158 had definite, microbiologically confirmed TB, 32 had probable TB, while 389 participants had ORD. Nine signatures differentiated between ORD and TB with equivalent performance (Satproedprai7: area under the curve 0.83 (95% CI 0.79–0.87); Jacobsen3: 0.83 (95% CI 0.79–0.86); Suliman2: 0.82 (95% CI 0.78–0.86); Roe1: 0.82 (95% CI 0.78–0.86); Kaforou22: 0.82 (95% CI 0.78–0.86); Sambarey10: 0.81 (95% CI 0.77–0.85); Duffy9: 0.81 (95% CI 0.76–0.86); Gliddon3: 0.8 (95% CI 0.75–0.85); Suliman4 0.79 (95% CI 0.75–0.84)). Benchmarked against a 90% sensitivity, these signatures achieved specificities between 44% (95% CI 38–49%) and 54% (95% CI 49–59%), not meeting the TPP criteria. Signature scores significantly varied by HIV status and country. In country-specific analyses, several signatures, such as Satproedprai7 and Penn-Nicholson6, met the minimal TPP criteria for a triage test in Ethiopia, Malawi and South Africa. Conclusion No signatures met the TPP criteria in a pooled analysis of all countries, but several signatures met the minimum criteria for a non-sputum TB triage test in some countries.
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