Early alveolar macrophage response and IL-1R-dependent T cell priming determine transmissibility of Mycobacterium tuberculosis strains

Funding Information: The work was funded by the National Institute of Allergy and Infectious Diseases, National Institutes of Health grants U19AI111276 and U01AI065663 to R.R.R., R.D., J.J.E. and P.S., and NIAID training grant T32AI125185 to A.L. The study sponsors were not involved in the study design, in the collection, analysis, and interpretation of data; in the writing of the manuscript; or in the decision to submit the manuscript for publication. Publisher Copyright: © 2022, The Author(s).Mechanisms underlying variability in transmission of Mycobacterium tuberculosis strains remain undefined. By characterizing high and low transmission strains of M.tuberculosis in mice, we show here that high transmission M.tuberculosis strain induce rapid IL-1R-dependent alveolar macrophage migration from the alveolar space into the interstitium and that this action is key to subsequent temporal events of early dissemination of bacteria to the lymph nodes, Th1 priming, granulomatous response and bacterial control. In contrast, IL-1R-dependent alveolar macrophage migration and early dissemination of bacteria to lymph nodes is significantly impeded in infection with low transmission M.tuberculosis strain; these events promote the development of Th17 immunity, fostering neutrophilic inflammation and increased bacterial replication. Our results suggest that by inducing granulomas with the potential to develop into cavitary lesions that aids bacterial escape into the airways, high transmission M.tuberculosis strain is poised for greater transmissibility. These findings implicate bacterial heterogeneity as an important modifier of TB disease manifestations and transmission.publishersversionpublishe

Transmission phenotype of mycobacterium tuberculosis strains is mechanistically linked to induction of distinct pulmonary pathology

In a study of household contacts (HHC), households were categorized into High (HT) and Low (LT) transmission groups based on the proportion of HHC with a positive tuberculin skin test. The Mycobacterium tuberculosis (Mtb) strains from HT and LT index cases of the households were designated Mtb-HT and Mtb-LT, respectively. We found that C3HeB/FeJ mice infected with Mtb-LT strains exhibited significantly higher bacterial burden compared to Mtb-HT strains and also developed diffused inflammatory lung pathology. In stark contrast, a significant number of mice infected with Mtb-HT strains developed caseating granulomas, a lesion type with high potential to cavitate. None of the Mtb-HT infected animals developed diffused inflammatory lung pathology. A link was observed between increased in vitro replication of Mtb-LT strains and their ability to induce significantly high lipid droplet formation in macrophages. These results support that distinct early interactions of Mtb-HT and Mtb-LT strains with macrophages and subsequent differential trajectories in pathological disease may be the mechanism underlying their transmission potential.publishersversionpublishe

Low-cost rapid detection of rifampicin resistant tuberculosis using bacteriophage in Kampala, Uganda

BACKGROUND: Resistance to anti-tuberculosis drugs is a serious public health problem. Multi-drug resistant tuberculosis (MDR-TB), defined as resistance to at least rifampicin and isoniazid, has been reported in all regions of the world. Current phenotypic methods of assessing drug susceptibility of M. tuberculosis are slow. Rapid molecular methods to detect resistance to rifampicin have been developed but they are not affordable in some high prevalence countries such as those in sub Saharan Africa. A simple multi-well plate assay using mycobacteriophage D29 has been developed to test M. tuberculosis isolates for resistance to rifampicin. The purpose of this study was to investigate the performance of this technology in Kampala, Uganda. METHODS: In a blinded study 149 M. tuberculosis isolates were tested for resistance to rifampicin by the phage assay and results compared to those from routine phenotypic testing in BACTEC 460. Three concentrations of drug were used 2, 4 and 10 μg/ml. Isolates found resistant by either assay were subjected to sequence analysis of a 81 bp fragment of the rpoB gene to identify mutations predictive of resistance. Four isolates with discrepant phage and BACTEC results were tested in a second phenotypic assay to determine minimal inhibitory concentrations. RESULTS: Initial analysis suggested a sensitivity and specificity of 100% and 96.5% respectively for the phage assay used at 4 and 10 μg/ml when compared to the BACTEC 460. However, further analysis revealed 4 false negative results from the BACTEC 460 and the phage assay proved the more sensitive and specific of the two tests. Of the 39 isolates found resistant by the phage assay 38 (97.4%) were found to have mutations predictive of resistance in the 81 bp region of the rpoB gene. When used at 2 μg/ml false resistant results were observed from the phage assay. The cost of reagents for testing each isolate was estimated to be 1.3US$ when testing a batch of 20 isolates on a single 96 well plate. Results were obtained in 48 hours. CONCLUSION: The phage assay can be used for screening of isolates for resistance to rifampicin, with high sensitivity and specificity in Uganda. The test may be useful in poorly resourced laboratories as a rapid screen to differentiate between rifampicin susceptible and potential MDR-TB cases

Mycobacterium tuberculosis progresses through two phases of latent infection in humans

Little is known about the physiology of latent Mycobacterium tuberculosis infection. We studied the mutational rates of 24 index tuberculosis (TB) cases and their latently infected household contacts who developed active TB up to 5.25 years later, as an indication of bacterial physiological state and possible generation times during latent TB infection in humans. Here we report that the rate of new mutations in the M. tuberculosis genome decline dramatically after two years of latent infection (two-sided p < 0.001, assuming an 18 h generation time equal to log phase M. tuberculosis, with latency period modeled as a continuous variable). Alternatively, assuming a fixed mutation rate, the generation time increases over the latency duration. Mutations indicative of oxidative stress do not increase with increasing latency duration suggesting a lack of host or bacterial derived mutational stress. These results suggest that M. tuberculosis enters a quiescent state during latency, decreasing the risk for mutational drug resistance and increasing generation time, but potentially increasing bacterial tolerance to drugs that target actively growing bacteria.publishersversionpublishe

Host determinants of infectiousness in smear-positive patients with pulmonary tuberculosis

Background Epidemiologic data suggests that only a minority of tuberculosis (TB) patients are infectious. Cough aerosol sampling is a novel quantitative method to measure TB infectiousness. Methods We analyzed data from three studies conducted in Uganda and Brazil over a 13-year period. We included sputum acid fast bacilli (AFB) and culture positive pulmonary TB patients and used a cough aerosol sampling system (CASS) to measure the number of colony-forming units (CFU) of Mycobacterium tuberculosis in cough-generated aerosols as a measure for infectiousness. Aerosol data was categorized as: aerosol negative (CFU = 0) and aerosol positive (CFU > 0). Logistic regression models were built to identify factors associated with aerosol positivity. Results M. tuberculosis was isolated by culture from cough aerosols in 100/233 (43%) TB patients. In an unadjusted analysis, aerosol positivity was associated with fewer days of antituberculous therapy before CASS sampling (p = .0001), higher sputum AFB smear grade (p = .01), shorter days to positivity in liquid culture media (p = .02), and larger sputum volume (p = .03). In an adjusted analysis, only fewer days of TB treatment (OR 1.47 per 1 day of therapy, 95% CI 1.16-1.89; p = .001) was associated with aerosol positivity. Conclusion Cough generated aerosols containing viable M. tuberculosis, the infectious moiety in TB, are detected in a minority of TB patients and rapidly become non-culturable after initiation of antituberculous treatment. Mechanistic studies are needed to further elucidate these findings.publishersversionpublishe

RePORT International: Advancing Tuberculosis Biomarker Research Through Global Collaboration

Progress in tuberculosis clinical research is hampered by a lack of reliable biomarkers that predict progression from latent to active tuberculosis, and subsequent cure, relapse, or failure. Regional Prospective Observational Research in Tuberculosis (RePORT) International represents a consortium of regional cohorts (RePORT India, RePORT Brazil, and RePORT Indonesia) that are linked through the implementation of a Common Protocol for data and specimen collection, and are poised to address this critical research need. Each RePORT network is designed to support local, in-country tuberculosis-specific data and specimen biorepositories, and associated research. Taken together, the expected results include greater global clinical research capacity in high-burden settings, and increased local access to quality data and specimens for members of each network and their domestic and international collaborators. Additional networks are expected to be added, helping to spur tuberculosis treatment and prevention research around the world

Correlation between CT features of active tuberculosis and residual metabolic activity on end-of-treatment FDG PET/CT in patients treated for pulmonary tuberculosis

Patients who complete a standard course of anti-tuberculous treatment (ATT) for pulmonary tuberculosis and are declared cured according to the current standard of care commonly have residual metabolic activity (RMA) in their lungs on fluorine-18 fluorodeoxyglucose positron emission tomography/computed tomography (FDG PER/CT) imaging. RMA seen in this setting has been shown to be associated with relapse of tuberculosis. The routine clinical use of FDG PET/CT imaging for treatment response assessment in tuberculosis is hindered by cost and availability. CT is a more readily available imaging modality. We sought to determine the association between CT features suggestive of active tuberculosis and RMA on FDG PET/CT obtained in patients who completed a standard course of ATT for pulmonary tuberculosis. We prospectively recruited patients who completed a standard course of ATT and declared cured based on negative sputum culture. All patients had FDG PET/CT within 2 weeks of completing ATT. We determined the presence of RMA on FDG PET images. Among the various lung changes seen on CT, we considered the presence of lung nodule, consolidation, micronodules in tree-in-bud pattern, FDG-avid chest nodes, and pleural effusion as suggestive of active tuberculosis. We determine the association between the presence of RMA on FDG PET and the CT features of active tuberculosis. We include 75 patients with a mean age of 36.09 ± 10.49 years. Forty-one patients (54.67%) had RMA on their FDG PET/CT while 34 patients (45.33%) achieved complete metabolic response to ATT. There was a significant association between four of the five CT features of active disease, p < 0.05 in all cases. Pleural effusion (seen in two patients) was the only CT feature of active disease without a significant association with the presence of RMA. This suggests that CT may be used in lieu of FDG PET/CT for treatment response assessment of pulmonary tuberculosis.CRDF Global for the project titled: The Clinical Research Unit (CRU) for the Advancement of Tuberculosis (TB) Biomarker-Targeted Interventions.http://frontiersin.org/Medicinedm2022Medical MicrobiologyNuclear Medicin

Serologic Responses in Childhood Pulmonary Tuberculosis.

BACKGROUND: Identification of the Mycobacterium tuberculosis immunoproteome and antigens associated with serologic responses in adults has renewed interest in developing a serologic test for childhood tuberculosis (TB). We investigated IgG antibody responses against M. tuberculosis antigens in children with well-characterized TB. METHODS: We studied archived sera obtained from hospitalized children with suspected pulmonary TB, and classified as having confirmed TB (culture-confirmed), unlikely TB (clinical improvement without TB treatment), or unconfirmed TB (all others). A multiplexed bead-based assay for IgG antibodies against 119 M. tuberculosis antigens was developed, validated and used to test sera. The area under the curves (AUCs) of the empiric receiver-operator characteristic curves were generated as measures of predictive ability. A cross-validated generalized linear model was used to select the most predictive combinations of antigens. RESULTS: For the confirmed TB versus unlikely TB comparison, the maximal single antigen AUC was 0.63, corresponding to sensitivity 0.60 and specificity 0.60. Older (age: 60+ months old) children's responses were better predictive of TB status than younger (age: 12-59 months old) children's, with a maximal single antigen AUC of -0.76. For the confirmed TB versus unlikely TB groups, the most predictive combinations of antigens assigned TB risk probabilities of 0.33 and 0.33, respectively, when all ages were considered, and 0.57 (interquartile range: 0.48-0.64) and 0.35 (interquartile range: 0.32-0.40) when only older children were considered. CONCLUSION: An antigen-based IgG test is unlikely to meet the performance characteristics required of a TB detection test applicable to all age groups

Airborne Infection with Bacillus anthracis—from Mills to Mail

The lack of identified exposures in 2 of the 11 cases of bioterrorism-related inhalation anthrax in 2001 raised uncertainty about the infectious dose and transmission of Bacillus anthracis. We used the Wells-Riley mathematical model of airborne infection to estimate 1) the exposure concentrations in postal facilities where cases of inhalation anthrax occurred and 2) the risk for infection in various hypothetical scenarios of exposure to B. anthracis aerosolized from contaminated mail in residential settings. These models suggest that a small number of cases of inhalation anthrax can be expected when large numbers of persons are exposed to low concentrations of B. anthracis. The risk for inhalation anthrax is determined not only by bacillary virulence factors but also by infectious aerosol production and removal rates and by host factors

Treatment outcomes of new tuberculosis patients hospitalized in Kampala, Uganda: a prospective cohort study.

BACKGROUND: In most resource limited settings, new tuberculosis (TB) patients are usually treated as outpatients. We sought to investigate the reasons for hospitalisation and the predictors of poor treatment outcomes and mortality in a cohort of hospitalized new TB patients in Kampala, Uganda. METHODS AND FINDINGS: Ninety-six new TB patients hospitalised between 2003 and 2006 were enrolled and followed for two years. Thirty two were HIV-uninfected and 64 were HIV-infected. Among the HIV-uninfected, the commonest reasons for hospitalization were low Karnofsky score (47%) and need for diagnostic evaluation (25%). HIV-infected patients were commonly hospitalized due to low Karnofsky score (72%), concurrent illness (16%) and diagnostic evaluation (14%). Eleven HIV uninfected patients died (mortality rate 19.7 per 100 person-years) while 41 deaths occurred among the HIV-infected patients (mortality rate 46.9 per 100 person years). In all patients an unsuccessful treatment outcome (treatment failure, death during the treatment period or an unknown outcome) was associated with duration of TB symptoms, with the odds of an unsuccessful outcome decreasing with increasing duration. Among HIV-infected patients, an unsuccessful treatment outcome was also associated with male sex (P = 0.004) and age (P = 0.034). Low Karnofsky score (aHR = 8.93, 95% CI 1.88 - 42.40, P = 0.001) was the only factor significantly associated with mortality among the HIV-uninfected. Mortality among the HIV-infected was associated with the composite variable of CD4 and ART use, with patients with baseline CD4 below 200 cells/µL who were not on ART at a greater risk of death than those who were on ART, and low Karnofsky score (aHR = 2.02, 95% CI 1.02 - 4.01, P = 0.045). CONCLUSION: Poor health status is a common cause of hospitalisation for new TB patients. Mortality in this study was very high and associated with advanced HIV Disease and no use of ART