BACKGROUND:Diagnostic performance of tongue swab Mycobacterium tuberculosis PCR has been evaluated for facility-based triage of symptomatic tuberculosis (TB). It is unknown whether tongue swab performance differs for detection of asymptomatic TB in community-based screening. METHODS:Tongue swabs were collected from adult household contacts of TB patients (HHC Cohort), and symptomatic adults presenting to clinics with presumptive TB (Clinic Cohort), at eight South African sites. TB Cases were defined by positive sputum Xpert Ultra or liquid culture, performed in all participants; and matched ~1:3 (HHC Cohort) or ~1:2 (Clinic Cohort) to Controls without TB. Tongue swabs in both cohorts were tested by high-volume qPCR; and in the Clinic Cohort, also by sequence-specific magnetic capture (SSMaC) with qPCR. RESULTS:The Clinic Cohort included 217 TB Cases (100% symptomatic) and 437 Controls. The HHC Cohort included 44 TB Cases (84.1% asymptomatic) and 136 Controls. In the Clinic Cohort, sensitivity of SSMaC with qPCR was 73.2% (specificity 94.6%), but not significantly higher than high-volume qPCR (63.8%; p = 0.14) (specificity 94.4%). Sensitivity of high-volume qPCR in the Clinic Cohort (63.8%) was significantly higher than the HHC Cohort (34.1%; p = 0.0007) (specificity 91.9%). Among HHC, high-volume qPCR sensitivity was 35.1% for asymptomatic TB; 52.2% for TB with abnormal CXR; and 100% for TB with High sputum Xpert Ultra grade. CONCLUSIONS:Sensitivity of tongue swab high-volume qPCR for community-based, household screening for asymptomatic TB was low, approximately half that of facility-based triage for symptomatic TB, but increased with radiographic severity and sputum bacillary load.

BACKGROUND:To understand how lung pathology relates to symptoms, microbiology, and progression risk in tuberculosis and to advance diagnostic development, we determined the frequency at screening of tuberculosis-consistent lesions in asymptomatic individuals within 5 years of tuberculosis diagnosis using highly sensitive imaging ([18F]-fluorodeoxyglucose PET-CT) and compared with chest x-ray computer-aided detection (CAD). METHODS:We enrolled a prospective longitudinal cohort in Khayelitsha, Cape Town, South Africa, of asymptomatic, HIV-uninfected contacts aged 18-65 years of patients with rifampicin-resistant tuberculosis, a tuberculosis high-risk group not eligible for chemoprophylaxis. Participants underwent baseline PET-CT, chest x-ray, phlebotomy, and intensive sputum collection, and were classified into four PET-CT lung categories: consistent with tuberculosis, inactive tuberculosis, other lesions, and normal. Chest x-ray was processed by three types of CAD software (CAD4TB [version 7.0], qXR [version 3.0.0], and Lunit [version 3.1.4.111]). Follow-up included symptom-agnostic tuberculosis screening (23-38 months) and provincial register review (≤74 months), and a subgroup had repeat PET-CT (5-15 months). Tuberculosis was defined as bacteriologically confirmed or clinically diagnosed. The primary outcome measures were hazard ratio (HR) for tuberculosis diagnosis and treatment by baseline PET-CT lung abnormality category with normal as the reference group, and diagnostic performance of chest x-ray CAD software using area under the receiver operator characteristic curve (AUC). FINDINGS:250 asymptomatic adults were enrolled between March 3, 2015, and Oct 11, 2017, irrespective of tuberculosis history or previous infection, and followed up for 1107 person-years (median 4·7 years [IQR 4·0-5·1]). 18 (7%) participants were treated for tuberculosis (16 [89%] of 18 bacteriologically confirmed). Six of 18 participants were diagnosed at baseline (four requiring induced sputum culture) and 12 of 18 after a median of 32 months (IQR 12-35). By baseline PET-CT category, tuberculosis was diagnosed and treated in 12 (41%) of 29 participants with scans consistent with tuberculosis, two (7%) of 30 with scans consistent with inactive tuberculosis, two (2%) of 83 with scans showing other lesions, and two (2%) of 108 with scans showing normal lungs. Participants with baseline PET-CT scans consistent with tuberculosis had the highest risk of 5-year tuberculosis diagnosis (HR 28·54 [95% CI 6·37-127·81] compared with those with scans showing normal lungs, p<0·0001), with no significant risk for scans consistent with inactive tuberculosis (3·55 [0·50-25·21], p=0·21) or other lung lesions (1·30 [0·18-9·23], p=0·79). 11 (69%) of the 16 participants with bacteriologically confirmed tuberculosis were asymptomatic at bacteriological confirmation, and ten (91%) of 11 had baseline PET-CT scans consistent with tuberculosis. Using baseline PET-CT classification as reference, the AUC for chest x-ray CAD software ranged from 0·86 (95% CI 0·72-0·99) to 0·89 (0·75-1·00) for bacteriologically confirmed tuberculosis. INTERPRETATION:Most adult asymptomatic contacts diagnosed with tuberculosis over 5 years had baseline radiographically evident disease, not radiographically negative incipient tuberculosis. Although PET-CT is not feasible for routine screening, it provides a highly sensitive reference benchmark for diagnostic development, with chest x-ray CAD performing comparatively well. FUNDING:South Africa Medical Research Council, US National Institutes of Health, Gates Foundation, Wellcome, UK Research and Innovation Medical Research Council, and Walter and Eliza Hall Institute of Medical Research.

Background:Mycobacteria-specific T-cell activation is a robust biomarker of recent Mycobacterium tuberculosis infection, disease progression and tuberculosis (TB) disease. We evaluated this promising biomarker, termed TB-TASA, as a treatment monitoring tool in the context of a treatment-shortening study. Methods:Using a panel of mycobacterial antigens, we observed higher TB-TASA scores in TB patients compared to interferon-γ (IFN-γ) release assay positive (IGRA+) controls, regardless of mycobacterial antigen specificity, consistent with previous studies. We derived a TB-TASA positivity threshold of 10% HLA-DR+ mycobacteria-specific T-cells using a computational analysis pipeline developed with the OpenCyto R package. This threshold was robust across three previously published case-control studies that used different sample types (i.e., whole blood or peripheral blood mononuclear cells), varying duration of antigen stimulation and different flow cytometry antibody panels, all of which achieved sensitivity and specificity >90% and >70%, respectively. Results:In a prospective randomised clinical trial assessing treatment shortening in TB patients with less severe disease, higher TB-TASA scores were observed at treatment completion in patients who relapsed or failed treatment during follow-up compared to those successfully treated with a receiver operating characteristic area under the curve (ROC AUC) of 0.89 (95% CI 0.69-1), supporting TB-TASA as a treatment monitoring tool. Discussion:We demonstrated that TB-TASA could also be reliably measured in capillary blood collected via fingerprick from IGRA+ controls and TB patients, achieving an ROC AUC of 0.96 (95% CI 0.9-1) and sensitivity and specificity of 95% and 69%, respectively. Together, these results support the continued development of TB-TASA as a potential tool for diagnosing TB and monitoring treatment responses.

Background:High-risk subgroups among household contacts of persons with tuberculosis (TB) might benefit from additional interventions. However, the significance of an abnormal baseline chest radiograph (CXR) suggestive of TB, despite negative sputum microbiology, is uncertain. Methods:Adults (≥18 years) with recent household TB exposure were enrolled at three South African sites (April 2021-September 2022). All participants underwent symptom screening, CXR, and sputum Xpert Ultra and MGIT culture. Pulmonary TB diagnosis was microbiologically-confirmed. Participants were followed for symptomatic incident TB through 12 months. Multivariable logistic regression identified factors associated with abnormal CXR suggestive of TB. Poisson regression estimated incidence rate ratios (IRR). Results:Baseline CXR were abnormal in 157/795 (19.7%) participants; associated with older age (adjusted odds ratio, aOR=1.04, 95%CI 1.02-1.05); prior TB (aOR=6.39, 95%CI 4.18-9.78); and current smoking (aOR=1.61, 95%CI 1.00-2.62). Symptomatic incident TB developed in 8/795 (1.0%) participants, including 7/8 (87.5%) who were asymptomatic and 4/8 (50.0%) with abnormal CXR at baseline. TB incidence was four-fold higher in those with abnormal versus normal CXR (IRR=4.02, 95%CI 1.01-15.97), with a risk difference of 1,969 (95%CI -657-4,595) per 100,000 person-years, but after median 12.1 (IQR 11.1-13.1) months follow-up, 153/157 (97.5%) had not progressed to incident TB. Conclusions:Adult contacts with CXR abnormalities, but without prevalent TB, had a four-fold higher risk of TB within one year, compared to those with normal CXR. This additional risk warrants targeted preventive treatment and extended surveillance, but since most remained TB-free, therapeutic TB treatment is not justified.

Background:The World Health Organisation (WHO) recommends digital chest radiography (dCXR) with computer-aided detection (CAD) for tuberculosis (TB) screening of individuals >15 years of age. Methodology:Adults (≥18 years) were enrolled (March 2021-December 2022) in South Africa into a community-based Screening Cohort (household contacts) and a facility-based Triage Cohort (symptomatic clinic attendees). Microbiologically-confirmed pulmonary TB required positive sputum culture and/or Xpert Ultra. Asymptomatic TB was diagnosed in participants without TB symptoms. dCXR were read by blinded human readers and qXR CAD (0.5 threshold; Qure.AI, India). Results:dCXR from 1,353 participants (886 Screening Cohort; 467 Triage Cohort) were analysed. Microbiologically-confirmed TB occurred in 48 (5.4%) Screening Cohort [9 symptomatic (19%) and 39 asymptomatic (81%)]; and 116 (24.8%) Triage Cohort (all symptomatic) participants. dCXR sensitivity (human readers) for asymptomatic TB in the Screening Cohort was 56.4%, vs. 72.4% for symptomatic TB in the Triage Cohort (difference -16%; 95%CI -2.9 to -29.1); with specificities 94.1% and 81.2%, respectively. Corresponding qXR CAD sensitivities were 69.2% vs. 83.6% (difference -14.4%; 95%CI -26 to -2.8), with specificities 89.3% and 73.5%, respectively. The difference in dCXR sensitivity and specificity for asymptomatic TB between qXR CAD and human readers was 12.8% (95%CI -0.48 to 26.1) and -4.8% (95%CI -12.4 to 28.2), respectively. Conclusion:Sensitivity of community-based dCXR screening for microbiologically-confirmed asymptomatic TB among household contacts was lower than for facility-based triage of symptomatic TB, but approached 70% with CAD. Neither human reader nor qXR CAD evaluation met WHO targets for a TB screening test (90% sensitivity; 80% specificity).

Six months of drug treatment is standard of care for drug-sensitive pulmonary tuberculosis (TB). Understanding the factors determining the length of treatment required for durable cure would allow individualization of treatment durations. We conducted a prospective, randomized, controlled noninferiority trial (PredictTB) of 4 versus 6 months of chemotherapy in patients with pulmonary TB in South Africa and China. Seven hundred and four participants with newly diagnosed, drug-sensitive TB were enrolled and stratified on the basis of radiographic disease characteristics assessed by FDG PET/CT imaging. Participants with less extensive disease (n = 273) were randomly assigned at week 16 to complete therapy after 4 months or continue receiving treatment for 6 months. This study was stopped early after an interim analysis revealed that patients assigned to the 4-month treatment arm had a higher risk of relapse. Among participants who received 4 months of chemotherapy, 17 of 141 (12.1%) experienced TB-specific unfavorable outcomes compared with only 2 of 132 (1.5%) who completed 6 months of treatment. In the nonrandomized arm that included participants with more extensive disease, only 8 of 248 (3.2%) experienced unfavorable outcomes. Total lung cavity volume and lesion glycolysis at week 16 were associated with the risk of unfavorable outcomes. PET/CT imaging at TB recurrence showed that bacteriological relapses predominantly occurred in active cavities originally present at baseline. Subsequent post hoc automated segmentation of serial PET/CT scans combined with machine learning enabled the classification of participants according to their likelihood of relapse.

Early detection of tuberculosis (TB) is central to global efforts for TB care and prevention. Conventional symptom-based screening and sputum microbiology fail to identify a substantial proportion of cases, particularly those that are asymptomatic or have low bacillary burden. Non-invasive medical imaging offers a critical solution for visualizing pulmonary pathology before individuals with TB develop characteristic symptoms. Chest X-ray has long been central to community screening, and recent advances in computer-aided detection systems endorsed by the World Health Organization have improved scalability and reduced reliance on expert interpretation. However, limitations in sensitivity for the earliest stages of disease and reduced specificity in individuals with prior lung pathology persist, underscoring the need for further research to improve performance for early TB detection. High-resolution modalities, such as computed tomography, magnetic resonance imaging, and functional imaging approaches, such as positron emission tomography and single-photon emission computed tomography, provide unparalleled insights into lesion dynamics, disease activity, and treatment response. They can be utilized to benchmark the development of novel diagnostic tests for bacteriologically unconfirmed TB and define imaging correlates of early disease progression and resolution. Furthermore, emerging innovations in pathogen-specific radiotracers may enable localization of viable bacilli in vivo. As a result, these new high-resolution imaging technologies offer transformative potential to address key knowledge gaps in the natural history, pathogenesis, dissemination, and therapeutic approaches to early pulmonary TB. Together, clinical imaging provides a framework for the development and validation of clinically relevant biomarkers and quantitative readouts that capture early, asymptomatic TB pathology and disease activity before conventional microbiologic confirmation. Future work should focus on integrating advanced imaging with microbiology, host-response biomarkers, and artificial intelligence to define actionable imaging phenotypes that inform early diagnosis, risk stratification, and treatment decision-making across diverse settings.

Background : No comprehensive characterization of the respiratory tract (RT) microbiota has been done in people with tuberculosis (TB), a leading global cause of death. Methods : 16S rRNA gene sequencing was done on upper RT (URT; oral-washes, naso- and oro-pharyngeal swabs, supraglottic fluid), sputum and lower RT [LRT; bronchoalveolar lavage fluid (BALF) and protected specimen brushings] specimens from HIV-negative people with Xpert MTB/RIF-confirmed TB (cases; n=17) and healthy controls (n=11). In addition to their diseased lobe, cases had their non-diseased lobe sampled. Results : The LRT had the lowest α-diversity and β-diversity differed compared to other respiratory compartments. In cases, Mycobacterium relative abundance was highest in the diseased lobe 1.537% (CI 0-3.114), followed by the nasopharynx 0.059% (0.012-0.105), non-diseased lobe 0.054% (0-1.620), oropharynx 0.003% (0-0.010) and sputum 0.002% (0-0.004). Compared to the URT and sputum, cases’ LRTs were Mycobacterium- and Moraxella -enriched ( Erythromicrobium -enriched versus sputum only). In paired comparisons of diseased versus non-diseased lobes in cases, the only differential taxon was Mycobacterium . Amongst non-diseased lobes, those of cases versus controls had reduced α-diversity with Mycoplasma -enrichment and Moraxella- and Klebsiella- depletion. Conclusion : Compared to healthy people, those with TB have a less diverse LRT microbiota, characterized by Mycobacterium -enrichment (within the diseased lobe and surprisingly least so in sputum) and depletion of taxa associated with healthy people. In people with TB, most microbial DNA is not mycobacterial within the diseased lobe and even the non-diseased lobes of cases are microbially distinct from controls. These findings provide a foundation for understanding respiratory tract host-microbiome interactions in TB.

Background:More than half of tuberculosis (TB) detected by community prevalence surveys is classified as asymptomatic. We evaluated yield of symptom and chest radiograph (CXR) screening of TB-exposed household contacts (HHC) in South Africa. Methods:Adult volunteers (≥18 years) with household exposure to pulmonary TB patients were enrolled at three sites. Systematic screening of TB symptoms (any duration), CXR (any abnormality), and sputum microscopy, Xpert Ultra, and liquid culture were performed. Serum C-reactive protein (CRP) was measured by multiplex bead array. Prevalent TB was microbiologically-confirmed (Xpert Ultra or culture). Symptomatic and asymptomatic TB were defined as prevalent TB with and without reported symptoms, respectively. Results:Between March 2021 - December 2022, 979 HHC were enrolled; 185 (18.9%) living with HIV and 187 (19.1%) with previous TB. Prevalent TB occurred in 51 (5.2%) and was asymptomatic in 42/51 (82.4%). Only 13/42 (31.0%) asymptomatic TB cases were smear-positive [8/13 (61.5%) graded scanty or 1+]. CRP did not discriminate healthy HHC from those with asymptomatic TB (AUC 0.60; 95%CI 0.47-0.73). An abnormal CXR was observed in 23/41 asymptomatic (sensitivity 56.1%, 95%CI 41.0-70.1%) versus 8/9 symptomatic (sensitivity 88.9%, 95%CI 56.5-98.0%) TB cases. Sensitivity of CXR in combination with symptom screening was 64.0% (32/50, 95%CI 50.1-75.9%) for all prevalent TB. Conclusions:More than 80% of confirmed TB cases among HHC were asymptomatic. CXR screening missed more than 40% of these asymptomatic cases. Community prevalence surveys reliant on symptom- and CXR-based approaches may significantly underestimate the prevalence of asymptomatic TB in endemic countries. Funding:Supported by RePORT South Africa through funding from the U.S. National Institutes of Health, CRDF Global, and the South African Medical Research Council. RESEARCH IN CONTEXT:Evidence before this study: World Health Organisation (WHO) guidelines for systematic tuberculosis (TB) screening recommend symptom screening and chest radiography (CXR), based on a Cochrane meta-analysis reporting 70.6% sensitivity (any TB symptom) and 94.7% sensitivity (any CXR abnormality) for bacteriologically-confirmed pulmonary TB. National TB prevalence surveys rely on a positive symptom screen or abnormal CXR to trigger diagnostic sputum testing. This approach to community screening would, by definition, miss asymptomatic TB cases without CXR evidence of disease. We reviewed the reference list of the aforementioned meta-analysis for active case-finding studies of adolescents and adults aged 15 years and older in community and contact-tracing settings. We performed forward citation-tracking and searched reference lists, including studies published in English between Jan 1, 1980, and November 1, 2024. We excluded studies that included children <15 years; or that exclusively enrolled people with additional risk factors (HIV; diabetes; latent TB infection; prior TB). We found 28 studies that performed universal sputum testing for bacteriologically-confirmed pulmonary TB and reported 51.8% (95%CI 49.9-53.7%; I 2 = 89.2%) pooled sensitivity for symptom screening (any symptom; 24 studies, 2,969 TB cases) and 62.4% (95%CI 59.3-65.3%; I 2 = 88.3%) pooled sensitivity for CXR (any abnormality; 10 studies, 1,123 TB cases). Only four studies (145 TB cases) reported accuracy of symptom screening in parallel with chest radiography (pooled sensitivity 67.3%, 95%CI 57.3-75.9%; I 2 = 87.1%), but these studies did not disaggregate symptomatic and asymptomatic disease. Added value of this study: We performed systematic screening using universal sputum microbiological testing of 978 household contacts of pulmonary TB patients in three South African communities and compared symptom (any duration) and CXR (any abnormality) screening approaches against a microbiological reference standard. We detected confirmed pulmonary TB in 5.2% of household contacts, and 82.4% of these TB cases reported no TB symptoms. Asymptomatic TB in household contacts was pauci-bacillary and associated with low serum CRP levels that were indistinguishable from healthy controls, but distinct from symptomatic TB in a comparator group of clinic attendees. Sensitivity of CXR screening for asymptomatic TB was only 56.1%; sensitivity of combined symptom and CXR screening for all TB was marginally higher at 64.0%.Implications of all the available evidence: Our findings from household contacts suggest that symptom- and CXR-based approaches are inadequate for community TB screening in South Africa and do not meet the WHO Target Product Profile for a TB screening test (minimum 90% sensitivity; 70% specificity). National TB Prevalence Surveys that omit universal sputum microbiological testing may significantly underestimate the prevalence of asymptomatic TB in high-burden countries.

BACKGROUND:Tuberculosis (TB) remains a leading cause of morbidity and mortality worldwide. Individuals with type 2 diabetes (T2D) are at increased risk of developing active TB, tend to present with more severe disease and experience adverse treatment outcomes compared to individuals without T2D co-morbidity. However, the underlying mechanisms responsible for increased susceptibility of patients with T2D to TB remain poorly understood. METHODS:We performed bronchoscopies and collected bronchoalveolar lavage fluid (BAL) from TB contacts with and without T2D in South Africa and obtained paired human alveolar macrophages (HAMs) and monocyte-derived macrophages (MDMs) for transcriptomic, epigenetic and functional assays. FINDINGS:Following ex vivo infection with Mycobacterium tuberculosis (Mtb), T2D-HAMs had increased Mtb growth (p = 0.034) and elevated TNF production (p = 0.033). BAL fluid from patients with T2D contained fewer neutrophils (p = 0.018), with neutrophil frequency inversely correlating with Mtb growth in HAMs. Both T2D-HAMs (p = 0.0206) and T2D-MDMs (p = 0.0465) expressed less CD32 compared to control cells, with patients with T2D having fewer M1-like MDMs (p = 0.0072). Mtb-induced gene expression in T2D-HAMs was delayed, but genes involved in negative regulation of neutrophil migration were upregulated (FDR = 0.035), consistent with the reduced neutrophil recruitment to the lung. Epigenetic profiling revealed hypermethylation in T2D-HAM DNA compared to control HAMs, except for hypomethylated TNF signalling genes, aligning with increased TNF production. INTERPRETATION:Our findings demonstrate that T2D alters early HAM responses to Mtb, characterised by delayed gene transcription, epigenetically driven cytokine dysregulation and impaired neutrophil recruitment-collectively facilitating enhanced Mtb growth. This study provides the first comprehensive investigation of the diabetic HAM phenotype in the context of recent TB exposure, offering mechanistic insights into the increased TB susceptibility observed in patients with T2D. FUNDING:Research reported in this publication was supported by the National Institute of Allergy and Infectious Diseases (NIAID) of the National Institutes of Health (NIH) and the South African Medical Research Council (SAMRC) under the US-South African Program for Collaborative Biomedical Research, the National Research Foundation of South Africa, the National Health and Medical Research Council of Australia and the Mater Foundation. The Translational Research Institute is supported by the Australian Government. RNA-seq data was generated in the Genome Sequencing Facility, which is supported by UT Health San Antonio, the NIH and a CPRIT Core Facility Award.

A large proportion of individuals with tuberculosis (TB) are asymptomatic. The biological and inflammatory underpinnings of asymptomatic TB are unknown and may differ from symptomatic TB. We characterised blood transcriptomic and proteomic profiles in South African community screening vs. health facility-based triage cohorts. Asymptomatic TB shared core transcriptomic and proteomic features with symptomatic TB, including upregulation of innate, interferon and inflammatory pathways and downregulation of T and B cell pathways. Integration of transcriptomic and proteomic data from asymptomatic TB individuals identified two distinct sub-clusters characterized by higher or lower bacterial burden, blood IFN-γ responses, BMI, and chest radiographic abnormalities, suggesting different disease severity. We identified a new blood transcriptomic signature of asymptomatic TB. However, diagnostic performance of transcriptomic and proteomic markers was weaker for asymptomatic TB than symptomatic TB, suggesting that policy development for community-based, asymptomatic TB screening should not adopt biomarkers developed for symptomatic TB triage without further optimization.

To understand natural resistance to Mycobacterium tuberculosis ( Mtb ) infection, we studied people living with HIV (PLWH) in an area of high Mtb transmission. Given that alveolar leukocytes may contribute to this resistance, we performed single cell RNA-sequencing of bronchoalveolar lavage cells, unstimulated or ex vivo stimulated with Mtb . We obtained high quality cells for 7 participants who were TST & IGRA positive (called LTBI) and 6 who were persistently TST & IGRA negative (called resisters). Alveolar macrophages (AM) from resisters displayed more of an M1 phenotype relative to LTBI AM at baseline. Alveolar lymphocytosis (10%-60%) was exhibited by 5/6 resisters, resulting in higher numbers of CD4 + and CD8 + IFNG -expressing cells at baseline and upon Mtb challenge than LTBI samples. Mycobactericidal granulysin was expressed almost exclusively by a cluster of CD8 + T cells that co-expressed granzyme B, perforin and NK cell receptors. For resisters, these poly-cytotoxic T cells over-represented activating NK cell receptors and were present at 15-fold higher numbers in alveoli compared to LTBI. Altogether, our results showed that alveolar lymphocytosis, with increased numbers of alveolar IFNG -expressing cells and CD8 + poly-cytotoxic T cells, as well as activated AM were strongly associated with protection from persistent Mtb infection in PLWH.

BACKGROUND:Non-sputum-based, point-of-care triage tests for pulmonary tuberculosis could enhance tuberculosis diagnostic programs. We assessed the diagnostic accuracy of 2 finger-stick blood tests: the Cepheid 3 gene host-response cartridge (Xpert-HR), which measures 3 host messenger RNA transcripts, and the 3-host protein multibiomarker test (MBT). METHODS:We performed a prospective diagnostic accuracy study of consecutive participants with symptoms compatible with pulmonary tuberculosis in The Gambia, South Africa, Uganda, and Vietnam. A composite reference standard for active pulmonary tuberculosis incorporated chest radiography, symptom resolution, and sputum microbiological test results. A training-test set approach was used to evaluate test cutoff specificities at 90% sensitivity. RESULTS:Between 1 November 2020 and 1 May 2023, we screened 1262 participants aged 12-70 years with cough lasting >2 weeks and another symptom suggestive of tuberculosis. Of those who were classifiable by reference tests, 1154 participants had evaluable Xpert-HR results and 961 had evaluable MBT results. Xpert-HR had an area under the receiver operating characteristic (AUROC) curve of 0.92 at a cutoff of -1.275 or below, with a sensitivity of 92.8%, specificity of 62.5%, positive predictive value of 47.9%, and negative predictive value of 95.9%. The MBT had an AUROC of 0.91 at a cutoff of ≥0.42, with a sensitivity of 91.4%, specificity of 73.2%, positive predictive value of 52.0%, and negative predictive value of 96.4%. CONCLUSIONS:Our results show that both Xpert-HR and the MBT are promising non-sputum-based point-of-care tests. The MBT met the World Health Organization target product profile for a triage test, which suggests it should be further developed.

We explored the underlying mechanisms that may drive post-tuberculosis (TB) lung disease, a multifactorial, heterogenous, and prevalent disease. Extensive clinical phenotyping through fluorine-18 Fluorodeoxyglucose (FDG) positron emission tomography (PET)-computed tomography (CT) scans, pulmonary function testing, and symptom and quality of life questionnaires, was performed on a cohort of 48 adults who completed TB treatment within 6 months prior. Immunological characteristics of paired blood- and bronchoalveolar lavage fluid (BALF)-derived immune cells were assessed by multiplex bead-based immunoassay, ELISA and flow cytometry. There was agreement between measures of inflammation on PET, the severity of anatomical abnormalities on CT, and pulmonary function testing. However, of these, only PET was associated with exercise tolerance and symptom scores. Measures of radiologic extent (total glycolytic activity and SUVmax on PET, and segments involved on CT) also correlated with proteins detected in blood that implicate type 1 (IFN-γ, TNFα, IL-12) and type 2 (IL-4, IL-33) responses, ongoing remodelling of lung tissue (MMPs), airways and vasculature (VEGF), as well as subsets of activated CD8+ and CD4+ T-cells. The radiologic extent of structural post-TB lung involvement is associated with a range of impaired lung function measures and immunological dysregulation. Our findings suggest that obstructive and restrictive lung pathology due to pulmonary TB do not occur in opposition but rather point towards a mixed pathology in most TB survivors.

Background There is a concern that SARS-CoV-2 infection may drive poor outcomes after Mycobacterium tuberculosis Mtb exposure and infection. We performed an ex vivo Mtb killing assay using peripheral blood mononuclear cells (PBMC) from three groups: healthy household contacts of people with active TB with and without serologic evidence of previous SARS-CoV-2 infection (COV+HHC and COV-HHC), and participants with active TB and previous SARS-CoV-2 (COV+TB+). Methods Twenty participants per group from Cape Town, South Africa were classified according to SARS-CoV-2 anti-S and anti-N antibody tests. We infected PBMC from each participant at a MOI of 0.001 with Mtb strain H37Rv in a 4-day growth inhibition assay. Mycobacteria were quantified through inoculation into Bactec Mycobacteria Growth Indicator Tube (MGIT) liquid culture. PBMC from a subset of participants were infected in the presence of autologous time-matched serum and Mtb-uninfected control PBMCs were included. Results There was no difference in the time to detection of Mtb or the normalised Mtb growth ratio (log10CFUsample – log10CFUcontrol) between groups in the standard protocol, or when infected cells from the COV+HHC and COV+TB+ (n=10 each) groups were cultured with autologous time-matched serum. The group with active TB demonstrated the best Mtb growth control. Extracellular Mtb measured by culturing the supernatants of the infected cell cultures also did not show any difference between groups. Five (14.3%) uninfected controls were culture positive. Conclusion Our results show that previous SARS-CoV-2 does not affect the Mtb killing ability of circulating mononuclear immune cells in vitro. Previous SARS-CoV-2 is unlikely to affect the outcome of Mtb infection through this mechanism.

Background:There is a concern that SARS-CoV-2 infection may drive poor outcomes after Mycobacterium tuberculosis Mtb exposure and infection. We performed an ex vivo Mtb killing assay using peripheral blood mononuclear cells (PBMC) from three groups: healthy household contacts of people with active TB with and without serologic evidence of previous SARS-CoV-2 infection (COV+HHC and COV-HHC), and participants with active TB and previous SARS-CoV-2 (COV+TB+). Methods:Twenty participants per group from Cape Town, South Africa were classified according to SARS-CoV-2 anti-S and anti-N antibody tests. We infected PBMC from each participant at a MOI of 0.001 with Mtb strain H37Rv in a 4-day growth inhibition assay. Mycobacteria were quantified through inoculation into Bactec Mycobacteria Growth Indicator Tube (MGIT) liquid culture. PBMC from a subset of participants were infected in the presence of autologous time-matched serum and Mtb-uninfected control PBMCs were included. Results:There was no difference in the time to detection of Mtb or the normalised Mtb growth ratio (log10CFUsample - log10CFUcontrol) between groups in the standard protocol, or when infected cells from the COV+HHC and COV+TB+ (n=10 each) groups were cultured with autologous time-matched serum. The group with active TB demonstrated the best Mtb growth control. Extracellular Mtb measured by culturing the supernatants of the infected cell cultures also did not show any difference between groups. Five (14.3%) uninfected controls were culture positive. Conclusion:Our results show that previous SARS-CoV-2 does not affect the Mtb killing ability of circulating mononuclear immune cells in vitro. Previous SARS-CoV-2 is unlikely to affect the outcome of Mtb infection through this mechanism.

BACKGROUND:More than half of tuberculosis cases detected by community prevalence surveys are classified as asymptomatic. We evaluated yield of symptom and chest radiograph screening of tuberculosis-exposed household contacts in South Africa. METHODS:For this prospective observational cohort study, adult volunteers (aged ≥18 years) with household exposure within the past 6 months to patients with untreated or partially treated pulmonary tuberculosis, identified through local health services, were enrolled at three sites in South Africa (Worcester and Ravensmead, Western Cape Province, and Soweto, Gauteng Province). Household contacts were excluded if they were unlikely to attend study visits, or had conditions interfering with consent or study participation, including psychiatric illness, substance dependence, or incarceration. Systematic screening of tuberculosis symptoms (any duration), chest radiograph (any abnormality indicative of active tuberculosis), and sputum microscopy, Xpert Ultra, and liquid culture were performed. Serum C-reactive protein (CRP) was measured by multiplex bead array. Prevalent tuberculosis was microbiologically confirmed (Xpert Ultra or culture). Symptomatic and asymptomatic tuberculosis were defined as prevalent tuberculosis with and without reported symptoms compatible with tuberculosis. The primary outcome was the diagnostic yield (sensitivity) of microbiologically confirmed pulmonary tuberculosis. FINDINGS:Between April 22, 2021 and Sept 22, 2022, 979 household contacts were enrolled, 345 (35·2%) male and 634 (64·8%) female, 185 (18·9%) living with HIV and 187 (19·1%) with previous tuberculosis. Prevalent tuberculosis occurred in 51 (5·2%) and was asymptomatic in 42 (82·4%) of 51. Only 13 (31·0%) of 42 asymptomatic people with tuberculosis were sputum-smear positive; eight (61·5%) of these 13 had a low bacillary burden, with smear grades scanty or 1+ (1-99 acid-fast bacilli per 100 fields). CRP did not discriminate healthy household contacts from those with asymptomatic tuberculosis (area under the curve 0·60, 95% CI 0·47-0·73). An abnormal chest radiograph suggestive of tuberculosis was observed in 23 of 41 asymptomatic (sensitivity 56·1%, 95% CI 41·0-70·1) versus eight of nine symptomatic (sensitivity 88·9%, 56·5-98·0) people with tuberculosis. Sensitivity of chest radiograph in combination with symptom screening was 32 (64·0%) of 50 (50·1-75·9) for all prevalent tuberculosis. INTERPRETATION:More than 80% of confirmed people with tuberculosis among household contacts were asymptomatic; chest radiograph screening missed more than 40% of these. Community prevalence surveys reliant on symptom-based and chest radiograph-based approaches might substantially underestimate the prevalence of asymptomatic tuberculosis in endemic countries. FUNDING:Regional Prospective Observational Research for Tuberculosis South Africa through funding from the US National Institutes of Health, the Civilian Research and Development Foundation, and the South African Medical Research Council.