Methylome-wide Analysis Reveals Epigenetic Marks Associated With Resistance to Tuberculosis in Human Immunodeficiency Virus-Infected Individuals From East Africa.

BACKGROUND: Tuberculosis (TB) is the most deadly infectious disease globally and is highly prevalent in the developing world. For individuals infected with both Mycobacterium tuberculosis (Mtb) and human immunodeficiency virus (HIV), the risk of active TB is 10% or more annually. Previously, we identified in a genome-wide association study (GWAS) a region on chromosome 5 associated with resistance to TB, which included epigenetic marks that could influence gene regulation. We hypothesized that HIV-infected individuals exposed to Mtb who remain disease free carry epigenetic changes that strongly protect them from active TB. METHODS: We conducted a methylome-wide study in HIV-infected, TB-exposed cohorts from Uganda and Tanzania and integrated data from our GWAS. RESULTS: We identified 3 regions of interest that included markers that were differentially methylated between TB cases and controls with latent TB infection: chromosome 1 (RNF220, P = 4 × 10-5), chromosome 2 (between COPS8 and COL6A3, P = 2.7 × 10-5), and chromosome 5 (CEP72, P = 1.3 × 10-5). These methylation results co-localized with associated single-nucleotide polymorphisms (SNPs), methylation QTLs, and methylation × SNP interaction effects. These markers were in regions with regulatory markers for cells involved in TB immunity and/or lung. CONCLUSIONS: Epigenetic regulation is a potential biologic factor underlying resistance to TB in immunocompromised individuals that can act in conjunction with genetic variants.

Successful implementation of isoniazid preventive therapy at a pediatric HIV clinic in Tanzania.

BACKGROUND: In accordance with international guidance for tuberculosis (TB) prevention, the Tanzanian Ministry of Health recommends isoniazid preventive therapy (IPT) for children aged 12 months and older who are living with HIV. Concerns about tolerability, adherence, and potential mistreatment of undiagnosed TB with monotherapy have limited uptake of IPT globally, especially among children, in whom diagnostic confirmation is challenging. We assessed IPT implementation and adherence at a pediatric HIV clinic in Tanzania. METHODS: In this prospective cohort study, eligible children living with HIV aged 1-15 years receiving care at the DarDar Pediatric Program in Dar es Salaam who screened negative for TB disease were offered a 6-month regimen of daily isoniazid. Patients could choose to receive IPT via facility- or community-based care. Parents/caregivers and children provided informed consent and verbal assent respectively. Isoniazid was dispensed with the child's antiretroviral therapy every 1-3 months. IPT adherence and treatment completion was determined by pill counts, appointment attendance, and self-report. Patients underwent TB symptom screening at every visit. RESULTS: We enrolled 66 children between July and December 2017. No patients/caregivers declined IPT. Most participants were female (n = 43, 65.1%) and the median age was 11 years (interquartile range [IQR] 8, 13). 63 (95.5%) participants chose the facility-based model; due to the small number of participants who chose the community-based model, valid comparisons between the two groups could not be made. Forty-nine participants (74.2%) completed IPT within 10 months. Among the remaining 17, 11 had IPT discontinued by their provider due to adverse drug reactions, 5 lacked documentation of completion, and 1 had unknown outcomes due to missing paperwork. Of those who completed IPT, the average monthly adherence was 98.0%. None of the participants were diagnosed with TB while taking IPT or during a median of 4 months of follow-up. CONCLUSIONS: High adherence and treatment completion rates can be achieved when IPT is integrated into routine, self-selected facility-based pediatric HIV care. Improved record-keeping may yield even higher completion rates. IPT was well tolerated and no cases of TB were detected. IPT for children living with HIV is feasible and should be implemented throughout Tanzania.

Correction to: Successful implementation of isoniazid preventive therapy at a pediatric HIV clinic in Tanzania.

An amendment to this paper has been published and can be accessed via the original article.

The potential of CBC-derived ratios (monocyte-to-lymphocyte, neutrophil-to-lymphocyte, and platelet-to-lymphocyte) to predict or diagnose incident TB infection in Tanzanian adolescents.

BACKGROUND: Ratios of different immune cell populations (i.e., monocyte-to-lymphocyte, neutrophil-to-lymphocyte, and platelet-to-lymphocyte ratios) have been studied as a means of predicting future tuberculosis (TB) disease risk or to assist in the diagnosis of incident TB disease. No studies to-date, however, have evaluated the potential of these ratios to predict or assist in the diagnosis of incident TB infection - the first step in the natural history of TB disease. METHODS: In this prospective study, we evaluated the complete blood count (CBC)-derived metrics of monocyte-to-lymphocyte ratio (MLR), neutrophil-to-lymphocyte ratio (NLR), and platelet-to-lymphocyte ratio (PLR) as predictors of future TB infection risk or aids in the diagnosis of TB infection among 145 Tanzanian adolescents enrolled in the DAR-901 vaccine trial, using paired CBCs and interferon-gamma release assays (IGRAs) obtained at 0, 60 and 720 days after study enrollment. RESULTS: At baseline, there were no significant differences between study participants who remained persistently IGRA negative throughout the study period and those who subsequently converted to IGRA positive with respect to MLR (0.18 vs 0.17, p = 0.10), NLR (0.88 vs 1.02, p = 0.08), or PLR (115 vs 120, p = 0.28). Similarly, no significant differences were noted with respect to MLR, NLR, and PLR between IGRA converters and time-matched negative controls at the time of IGRA conversion. With respect to other blood cell measures, however, there were modest but significant differences between IGRA negatives and IGRA converters with respect to red blood cell count (4.8 vs 4.6 ×  106 cells/mcL, p = 0.008), hemoglobin (12.6 vs 12.3 g/dL, p = 0.01), and hematocrit (38.8 vs 37.8%, p = 0.005). CONCLUSIONS: In contrast to prior studies that have suggested that the ratios of different immune cell populations are associated with development of TB disease, our present findings do not demonstrate an association between these ratios and the development of TB infection. However, decreased red blood cell measures were associated with the subsequent development of TB infection, suggesting either that dysregulation of iron metabolism may play a role in TB pathogenesis or that following TB infection, iron dysregulation may precede IGRA positivity. TRIAL REGISTRATION: Clinicaltrials.gov NCT02712424 . Date of registration: March 14, 2016.

A chromosome 5q31.1 locus associates with tuberculin skin test reactivity in HIV-positive individuals from tuberculosis hyper-endemic regions in east Africa.

One in three people has been infected with Mycobacterium tuberculosis (MTB), and the risk for MTB infection in HIV-infected individuals is even higher. We hypothesized that HIV-positive individuals living in tuberculosis-endemic regions who do not get infected by Mycobacterium tuberculosis are genetically resistant. Using an "experiment of nature" design that proved successful in our previous work, we performed a genome-wide association study of tuberculin skin test positivity using 469 HIV-positive patients from prospective study cohorts of tuberculosis from Tanzania and Uganda to identify genetic loci associated with MTB infection in the context of HIV-infection. Among these individuals, 244 tested were tuberculin skin test (TST) positive either at enrollment or during the >8 year follow up, while 225 were not. We identified a genome-wide significant association between a dominant model of rs877356 and binary TST status in the combined cohort (Odds ratio = 0.2671, p = 1.22x10-8). Association was replicated with similar significance when examining TST induration as a continuous trait. The variant lies in the 5q31.1 region, 57kb downstream from IL9. Two-locus analyses of association of variants near rs877356 showed a haplotype comprised of rs877356 and an IL9 missense variant, rs2069885, had the most significant association (p = 1.59x10-12). We also replicated previously linked loci on chromosomes 2, 5, and 11. IL9 is a cytokine produced by mast cells and TH2 cells during inflammatory responses, providing a possible link between airway inflammation and protection from MTB infection. Our results indicate that studying uninfected, HIV-positive participants with extensive exposure increases the power to detect associations in complex infectious disease.

A Locus at 5q33.3 Confers Resistance to Tuberculosis in Highly Susceptible Individuals.

Immunosuppression resulting from HIV infection increases the risk of progression to active tuberculosis (TB) both in individuals newly exposed to Mycobacterium tuberculosis (MTB) and in those with latent infections. We hypothesized that HIV-positive individuals who do not develop TB, despite living in areas where it is hyperendemic, provide a model of natural resistance. We performed a genome-wide association study of TB resistance by using 581 HIV-positive Ugandans and Tanzanians enrolled in prospective cohort studies of TB; 267 of these individuals developed active TB, and 314 did not. A common variant, rs4921437 at 5q33.3, was significantly associated with TB (odds ratio = 0.37, p = 2.11 × 10(-8)). This variant lies within a genomic region that includes IL12B and is embedded in an H3K27Ac histone mark. The locus also displays consistent patterns of linkage disequilibrium across African populations and has signals of strong selection in populations from equatorial Africa. Along with prior studies demonstrating that therapy with IL-12 (the cytokine encoded in part by IL12B, associated with longer survival following MTB infection in mice deficient in CD4 T cells), our results suggest that this pathway might be an excellent target for the development of new modalities for treating TB, especially for HIV-positive individuals. Our results also indicate that studying extreme disease resistance in the face of extensive exposure can increase the power to detect associations in complex infectious disease.

Auditory Impairments in HIV-Infected Children.

OBJECTIVES: In a cross-sectional study of human immunodeficiency virus (HIV)-infected adults, the authors showed lower distortion product otoacoustic emissions (DPOAEs) in HIV+ individuals compared with controls as well as findings consistent with a central auditory processing deficit in HIV+ adults on antiretroviral therapy. The authors hypothesized that HIV+ children would also have a higher prevalence of abnormal central and peripheral hearing test results compared with HIV- controls. DESIGN: Pure-tone thresholds, DPOAEs, and tympanometry were performed on 244 subjects (131 HIV+ and 113 HIV- subjects). Thirty-five of the HIV+, and 3 of the HIV- subjects had a history of tuberculosis treatment. Gap detection results were available for 18 HIV- and 44 HIV+ children. Auditory brainstem response results were available for 72 HIV- and 72 HIV+ children. Data from ears with abnormal tympanograms were excluded. RESULTS: HIV+ subjects were significantly more likely to have abnormal tympanograms, histories of ear drainage, tuberculosis, or dizziness. All audiometric results were compared between groups using a two-way ANOVA with HIV status and ear drainage history as grouping variables. Mean audiometric thresholds, gap detection thresholds, and auditory brainstem response latencies did not differ between groups, although the HIV+ group had a higher proportion of individuals with a hearing loss >25 dB HL in the better ear. The HIV+ group had reduced DPOAE levels (p < 0.05) at multiple frequencies compared with HIV- subjects. No relationships were found between treatment regimens or delay in starting treatment and audiological parameters. CONCLUSIONS: As expected, children with HIV+ were more likely to have a history of ear drainage, and to have abnormal tympanograms. Similar to the adult findings, the HIV+ group did not show significantly reduced audiometric thresholds, but did have significantly lower DPOAE magnitudes. These data suggest that (1) HIV+ children often have middle ear damage which complicates understanding the direct effects of HIV on the hearing system, and (2) even when corrected for confounders DPOAEs were lower in the HIV+ group. Previous studies suggest ototoxicity from antiretroviral drugs is an unlikely cause of the reduced DPOAE magnitudes. Other possibilities include effects on efferent pathways connecting to outer hair cells or a direct effect of HIV on the cochlea.

Auditory impairments in HIV-infected individuals in Tanzania.

OBJECTIVES: Abnormal hearing tests have been noted in human immunodeficiency virus (HIV)-infected patients in several studies, but the nature of the hearing deficit has not been clearly defined. The authors performed a cross-sectional study of both HIV+ and HIV- individuals in Tanzania by using an audiological test battery. The authors hypothesized that HIV+ adults would have a higher prevalence of abnormal central and peripheral hearing test results compared with HIV- controls. In addition, they anticipated that the prevalence of abnormal hearing assessments would increase with antiretroviral therapy (ART) use and treatment for tuberculosis (TB). DESIGN: Pure-tone thresholds, distortion product otoacoustic emissions (DPOAEs), tympanometry, and a gap-detection test were performed using a laptop-based hearing testing system on 751 subjects (100 HIV- in the United States, plus 651 in Dar es Salaam, Tanzania, including 449 HIV+ [130 ART- and 319 ART+], and 202 HIV-, subjects. No U.S. subjects had a history of TB treatment. In Tanzania, 204 of the HIV+ and 23 of the HIV- subjects had a history of TB treatment. Subjects completed a video and audio questionnaire about their hearing, as well as a health history questionnaire. RESULTS: HIV+ subjects had reduced DPOAE levels compared with HIV- subjects, but their hearing thresholds, tympanometry results, and gap-detection thresholds were similar. Within the HIV+ group, those on ART reported significantly greater difficulties understanding speech in noise, and were significantly more likely to report that they had difficulty understanding speech than the ART- group. The ART+ group had a significantly higher mean gap-detection threshold compared with the ART- group. No effects of TB treatment were seen. CONCLUSIONS: The fact that the ART+/ART- groups did not differ in measures of peripheral hearing ability (DPOAEs, thresholds), or middle ear measures (tympanometry), but that the ART+ group had significantly more trouble understanding speech and had higher gap-detection thresholds indicates a central processing deficit. These data suggest that: (1) hearing deficits in HIV+ individuals could be a CNS side effect of HIV infection, (2) certain ART regimens might produce CNS side effects that manifest themselves as hearing difficulties, and/or (3) some ART regimens may treat CNS HIV inadequately, perhaps due to insufficient CNS drug levels, which is reflected as a central hearing deficit. Monitoring of central hearing parameters could be used to track central effects of either HIV or ART.

Greater preexisting interferon γ responses to mycobacterial antigens and lower bacillary load during HIV-associated tuberculosis.

The role of preexisting interferon (IFN) γ responses in controlling bacillary burden in human immunodeficiency virus (HIV)-associated tuberculosis is not known. Among BCG-immunized HIV-infected adults who developed tuberculosis in a phase III trial of an investigational tuberculosis vaccine, greater baseline IFN-γ responses to early secretory antigenic target 6 and Mycobacterium tuberculosis whole-cell lysate were associated with reduced bacillary burden on sputum smear grade, days to culture positivity on agar, and sputum culture grade during subsequent tuberculosis. This association was most consistent among recipients of the investigational vaccine. When HIV-associated tuberculosis develops, greater preexisting IFN-γ responses to mycobacterial antigens are associated with reduced tuberculosis bacillary burden. ClinicalTrials.gov Identifier. NCT0052195.

Recurrent tuberculosis risk among HIV-infected adults in Tanzania with prior active tuberculosis.

BACKGROUND: Active tuberculosis is common among human immunodeficiency virus (HIV)-infected persons living in tuberculosis-endemic areas, but the hazard of subsequent tuberculosis disease has not been quantified in a single prospective cohort. METHODS: Among HIV-infected, BCG-immunized adults with CD4 counts ≥200 cells/µL who received placebo in the DarDar tuberculosis vaccine trial in Tanzania, we compared the prospective risk of active tuberculosis between subjects who did and who did not report prior active tuberculosis. All subjects with a positive tuberculin skin test without prior active tuberculosis were offered isoniazid preventive treatment. Definite or probable tuberculosis was diagnosed during active follow-up using rigorous published criteria. RESULTS: We diagnosed 52 cases of definite and 92 cases of definite/probable tuberculosis among 979 subjects during a median follow-up of 3.2 years. Among the 80 subjects who reported prior active tuberculosis, 11 (13.8%) subsequently developed definite tuberculosis and 17 (21.3%) developed definite/probable tuberculosis, compared with 41 (4.6%) and 75 (8.3%), respectively, of 899 subjects without prior active tuberculosis (definite tuberculosis risk ratio [RR], 3.01; 95% confidence interval [CI], 1.61-5.63, P < .001; definite/probable tuberculosis RR, 2.55; 95% CI, 1.59-4.09, P < .001). In a Cox regression model adjusting for age, CD4 count, and isoniazid receipt, subjects with prior active tuberculosis had substantially greater hazard of subsequent definite tuberculosis (hazard radio [HR], 3.69; 95% CI, 1.79-7.63, P < .001) and definite/probable tuberculosis (HR, 2.78; 95% CI, 1.58-4.87, P < .001). CONCLUSIONS: Compared to subjects without prior tuberculosis, the hazard of active tuberculosis is increased 3-fold among HIV-infected adults with prior active tuberculosis. Clinical Trials Registration. NCT0052195.

Molecular epidemiology of HIV-associated tuberculosis in Dar es Salaam, Tanzania: strain predominance, clustering, and polyclonal disease.

Molecular typing of Mycobacterium tuberculosis can be used to elucidate the epidemiology of tuberculosis, including the rates of clustering, the frequency of polyclonal disease, and the distribution of genotypic families. We performed IS6110 typing and spoligotyping on M. tuberculosis strains isolated from HIV-infected subjects at baseline or during follow-up in the DarDar Trial in Tanzania and on selected community isolates. Clustering occurred in 203 (74%) of 275 subjects: 124 (80%) of 155 HIV-infected subjects with baseline isolates, 56 (69%) of 81 HIV-infected subjects with endpoint isolates, and 23 (59%) of 39 community controls. Overall, 113 (41%) subjects had an isolate representing the East Indian "GD" family. The rate of clustering was similar among vaccine and placebo recipients and among subjects with or without cellular immune responses to mycobacterial antigens. Polyclonal disease was detected in 6 (43%) of 14 patients with multiple specimens typed. Most cases of HIV-associated tuberculosis among subjects from this study in Dar es Salaam resulted from recently acquired infection. Polyclonal infection was detected and isolates representing the East Indian GD strain family were the most common.

Serial T-SPOT.TB responses in Tanzanian adolescents: Transient, persistent and irregular conversions.

BACKGROUND: Prospective studies of interferon-gamma release assays (IGRA) on healthy subjects in tuberculosis-endemic regions have not examined the long-term variability of serial assays. This issue is relevant to the interpretation of tuberculosis (TB) vaccine trials based on prevention of infection. METHODS: T-SPOT.TB assays were performed manually on healthy adolescents during a tuberculosis vaccine trial in Tanzania at 5 intervals over 3 years. Assay results were defined as negative, positive, borderline or invalid. Subsequently, microtiter plates were analyzed by an automated reader to obtain quantitative counts of spot forming cells (SFCs) for the present analysis. RESULTS: 3387 T-SPOT.TB samples were analyzed from 928 adolescents; manual and automated assay results were 97% concordant. Based on the quantitative results 143 (15%) participants were prevalent IGRA-positives at baseline, were ineligible for further study. Among the remaining IGRA-negative participants, the annual rate of IGRA conversion was 2·9%. Among 43 IGRA converters with repeat assays 12 (28%) were persistent converters, 16 (37%) were transient converters, and 15 (35%) comprised a new category defined as irregular converters (≥2 different subsequent results). ESAT-6 and CFP-10 responses were higher in prevalent than incident positives: 53 vs 36 for CFP-10 (p < 0·007); 44 vs 34 for ESAT-6 (p = 0·12). CONCLUSIONS: Definitions of IGRA conversion, reversion, and persistence depend critically on the frequency of testing. Multiple shifts in categories among adolescents in a TB-endemic country may represent multiple infections, variable host responses in subclinical infection, or assay variation. These findings should to be considered in the design and interpretation of TB vaccine trials based on prevention of infection. Household contact studies could determine whether even transient IGRA conversion might represent exposure to an active case of M. tuberculosis disease.

Current diagnosis and management of peripheral tuberculous lymphadenitis.

Peripheral tuberculous lymphadenitis accounts for ~10% of tuberculosis cases in the United States. Epidemiologic characteristics include a 1.4:1 female-to-male ratio, a peak age range of 30-40 years, and dominant foreign birth, especially East Asian. Patients present with a 1-2 month history of painless swelling of a single group of cervical lymph nodes. Definitive diagnosis is by culture or nucleic amplification of Mycobacterium tuberculosis; demonstration of acid fast bacilli and granulomatous inflammation may be helpful. Excisional biopsy has the highest sensitivity at 80%, but fine-needle aspiration is less invasive and may be useful, especially in immunocompromised hosts and in resource-limited settings. Antimycobacterial therapy remains the cornerstone of treatment, but response is slower than with pulmonary tuberculosis; persistent pain and swelling are common, and paradoxical upgrading reactions may occur in 20% of patients. The role of steroids is controversial. Initial excisional biopsy deserves consideration for both optimal diagnosis and management of the otherwise slow response to therapy.

The bacteraemia of disseminated tuberculosis among HIV-infected patients with prolonged fever in Tanzania.

BACKGROUND: Disseminated tuberculosis (TB) is a common cause of death among human immunodeficiency virus (HIV)-infected patients in developing countries. Blood culture offers a potential means to diagnose disseminated TB, but optimal blood culture methods have not been studied. METHODS: Two hundred and fifty-eight HIV-infected patients hospitalized in Tanzania with ≥2 weeks fever or cough had diagnostic studies for TB: 3 sputum samples for acid-fast bacilli smear and culture; 40 ml of blood for culture, randomized 1:1 to 40 ml × 1, or 20 ml × 2 collected 12-24 h apart. Blood was processed using automated MB BacT(®) broth and manual Isolator(®) lysis-centrifugation agar. Mortality was assessed at 2 months. RESULTS: TB was confirmed in 83 (32%) of 258 patients: by sputum only in 42 (51%, median CD4 = 72 cells/µl), blood only in 15 (18%, median CD4 = 44 cells/µl), and in sputum and blood in 26 (31%, median CD4 = 12 cells/µl). Blood was positive in 21 (16%) for 40 ml × 1 vs 20 (15%) for 20 ml × 1 (p = 0.83) vs 20 (16%) for 20 ml × 2 (p = 0.97). MB BacT was positive in 31 (76%) and Isolator was positive in 20 (49%) of 41 samples (p = 0.01). The mean colony-forming units/ml was 8 (range 3-14). Twenty-one (51%) patients with disseminated TB died; median survival was 6 days (range 0-58). CONCLUSIONS: Disseminated TB in HIV is characterized by persistent bacteraemia, delayed microbiological detection, and high mortality. Twenty millilitres of blood processed by automated broth is the optimal culture method to detect disseminated TB. Empiric TB therapy is warranted for HIV-infected patients from TB-endemic countries with prolonged cough or fever.

Revisiting rates of reactivation tuberculosis: a population-based approach.

RATIONALE: Reactivation tuberculosis (TB) occurs as a result of reactivation of latent TB infection (LTBI), and was reported to occur in the United States at a rate of 0.10 to 0.16 cases per 100 person-years in the 1950s; it has not been measured since. OBJECTIVES: To calculate the rate of reactivation TB in a U.S. community. METHODS: A population-based tuberculin skin test survey for LTBI was performed in western Palm Beach County, Florida, from 1998 to 2000 along with a cluster analysis of TB case isolates in the same area from 1997 to 2001. Reactivation (unclustered) TB was presumed to have arisen from the population with LTBI. MEASUREMENTS AND MAIN RESULTS: The rate of reactivation TB among persons with LTBI without HIV infection was 0.040 cases per 100 person-years (95% confidence interval [CI], 0.024-0.067) using the n method and 0.058 cases per 100 person-years (95% CI, 0.038-0.089) using the n-1 method. HIV infection was the strongest risk factor for reactivation (rate ratio [RR], 57; 95% CI, 27-120; P < 0.001). Among persons without HIV infection, reactivation was increased among those older than 50 years (RR, 3.8; 95% CI, 1.3-11) and among those born in the United States (RR, 3.2; 95% CI, 1.1-9.3). CONCLUSIONS: Rates of reactivation TB in this area have declined substantially since the 1950s. The greatest part of this decline may be attributed to the disappearance of old, healed TB in the population. If similar declines are seen in other areas of the United States, the cost-effectiveness of screening and treatment of LTBI may be substantially less than previously estimated.

Interferon γ responses to mycobacterial antigens protect against subsequent HIV-associated tuberculosis.

BACKGROUND: The cellular immune responses that protect against tuberculosis have not been identified. METHODS: We assessed baseline interferon γ (IFN­Î³) and lymphocyte proliferation assay (LPA) responses to antigen 85 (Ag85), early secretory antigenic target 6 (ESAT­6), and Mycobacterium tuberculosis whole cell lysate (WCL) in human immunodeficiency virus (HIV)-infected and bacille Calmette­Guérin (BCG)-immunized adults with CD4 cell counts of >or= 200 cells/µL who received placebo in the DarDar tuberculosis vaccine trial in Tanzania. Subjects were followed prospectively to diagnose definite or probable tuberculosis. RESULTS: Tuberculosis was diagnosed in 92 of 979 subjects during a mean follow­up of 3.2 years. The relative risk of tuberculosis among subjects with positive IFN­Î³ responses to Ag85 was 0.51 (95% confidence interval [CI], 0.26-0.99; P = .049), to ESAT­6 was 0.44 (95% CI, 0.23-0.85; P = .004), and to WCL was 0.67 (95% CI, 0.49-0.88; P = .002). The relative risk of tuberculosis was not significantly associated with baseline LPA responses. In a multivariate Cox regression model, subjects with IFN­Î³ responses to ESAT­6 and WCL had a lower hazard of developing tuberculosis, with a hazard ratio for ESAT­6 of 0.35 (95% CI, 0.16­0.77; P = .009) and a hazard ratio for WCL of 0.30 (95% CI, 0.16-0.56; P < .001). CONCLUSIONS: Baseline IFN­Î³ responses to ESAT-6 and WCL were associated with protection from subsequent tuberculosis among HIV-infected subjects with childhood BCG immunization in a region of high tuberculosis prevalence. Trial registration. ClinicalTrials.gov identifier: NCT00052195.

DAR-901 vaccine for the prevention of infection with Mycobacterium tuberculosis among BCG-immunized adolescents in Tanzania: A randomized controlled, double-blind phase 2b trial.

BACKGROUND: SRL172 prevented disease due to Mycobacterium tuberculosis in a Phase 3 trial. DAR-901 represents a scalable manufacturing process for SRL172. We sought to determine if DAR-901 would prevent infection with M. tuberculosis among BCG-primed adolescents age 13-15 years in Tanzania. METHODS: Adolescents with a negative T- SPOT.TBR interferon gamma release assay (IGRA) were randomized 1:1 to three intradermal injections of DAR-901 or saline placebo at 0, 2 and 4 months. Repeat IGRAs were performed at 2 months, and at 1, 2, and 3 years. The primary efficacy outcome was time to new TB infection (IGRA conversion to positive); the secondary outcome was time to persistent TB infection (IGRA conversion with repeat positive IGRA). RESULTS: Among 936 participants screened 667 were eligible and randomized to their first dose of vaccine or placebo (safety cohort). At 2 months, 625 participants remained IGRA-negative and were scheduled for the additional two doses (efficacy cohort). DAR-901 was safe and well-tolerated. One DAR-901 recipient developed a vaccine site abscess. Neither the primary nor secondary endpoints differed between the two treatment arms (p = 0.90 and p = 0.20, respectively). DAR-901 IGRA converters had median responses to ESAT-6 of 50.1 spot-forming cells (SFCs) vs. 19.6 SFCs in placebo IGRA converters (p = 0.03). CONCLUSIONS: A three-dose series of 1 mg DAR-901 was safe and well-tolerated but did not prevent initial or persistent IGRA conversion. DAR-901 recipients with IGRA conversion demonstrated enhanced immune responses to ESAT-6. Since protection against disease may require different immunologic responses than protection against infection a trial of DAR-901 to prevent TB disease is warranted. TRIAL REGISTRATION: The trial is registered at ClinicalTrials.gov as NCT02712424.