Search

Background: Engaging patients in healthcare for diagnosis and treatment through active case-finding, which is initiated by healthcare providers, involves methodically searching for tuberculosis in individuals who may not seek medical attention otherwise. Active case finding provides a promising opportunity to identify additional undetected cases of tuberculosis within the community. The current study aimed at evaluating household contact tracing of Tuberculosis among the household contacts of pulmonary TB cases who were recently diagnosed, as well as to identify risk factors in both household contacts and index cases that contribute to the transmission of TB within households. Materials and Method: This cross-sectional study, which was conducted within the community, focused on the household contacts of recently diagnosed patients of Pulmonary Tuberculosis (TB) who were confirmed through microbiological testing. These patients were registered at Tuberculosis Units (TUs) of JNMC and Jawan of Aligarh district, West U.P. The investigator conducted visits to each household and met with both the index case and their corresponding household contacts to establish rapport and gather necessary information. Results: Within this investigation, a total of 103 individuals who tested positive for sputum smear were identified as index cases, alongside 405 household contacts of these index cases. The predominant symptom observed in household contacts who tested positive during the screening process was a cough, with fever following closely behind. Among the 405 household contacts examined, 54 individuals (13.3%) exhibited symptoms associated with Tuberculosis. Further investigation revealed that 11 of these individuals (20.37%) tested positive for tuberculosis. The total prevalence of tuberculosis cases among household contacts was determined to be 2.72%. Conclusion: The present research affirms that conducting contact screening for households of active TB cases is a comparatively feasible and beneficial strategy that could result in prompt diagnosis and initiation of treatment for active TB, thereby diminishing the severity of the disease and lowering the rate of transmission. Also, it can help to improve TB-related health effects, social and economic implications, and treatment outcomes in the index cases. [ABSTRACT FROM AUTHOR]

Purpose: This is the first study to attempt microbiome diversity using metagenomic full-length 16S rRNA from respiratory specimens suspected of chronic pulmonary TB patients. Materials and methods: A 33 patients with suspected chronic pulmonary TB were included. Sputum specimens were cultured to detect mycobacterium sp. and extracted using QiAmp DNA mini kit modification and 16S rRNA metagenomic sequencing by nanopore grid ion sequencer. Microbiome analysis was performed using Pavian and Krona tools. Results: 9 patients were diagnosed with TB based on GeneXpert MTB/RIF assay, and 3 patients were detected with NTM pulmonary infection.The common genera identified from TB culture positive patients were streptococcus sp., prevotella sp., and veilonella sp. However, less was detected in two NTM infection patients. Metagenomic analysis revealed community bacteria species, including mycobacterium tuberculosis and NTM species, with the lowest number of unique reads. The abundance of streptococcus sp. were less than 30% in 4 patient with comorbid diabetes mellitus. Conclusions: Metagenomic targeted 16SrRNA full-length sequencing in the clinical respiratory specimen can provide diagnostic insight beyond standard microbiologic cultures and detailed profiling of microbial communities at the species level. [ABSTRACT FROM AUTHOR]

Most phase III trials in drug-resistant tuberculosis have either been underpowered to quantify differences in microbiological endpoints or have taken up to a decade to complete. Composite primary endpoints, dominated by differences in treatment discontinuation and regimen changes, may mask important differences in treatment failure and relapse. Although new regimens for drug-resistant tuberculosis appear very effective, resistance to new drugs is emerging rapidly. There is a need for shorter, safer and more tolerable regimens, including those active against bedaquiline-resistant tuberculosis. Transitioning from multiple regimen A versus regimen B trials to a single large phase III platform trial would accelerate the acquisition of robust estimates of relative efficacy and safety. Further efficiencies could be achieved by adopting modern adaptive platform designs. Collaboration among trialists, affected community representatives, funders and regulators is essential for developing such a phase III platform trial for drug-resistant tuberculosis treatment regimens. [ABSTRACT FROM AUTHOR]

Background and Aim: Mycobacterium tuberculosis (M. tuberculosis) is one of the most dangerous infectious diseases all over the world. Diagnosis of M. tuberculosis based on the culture and phenotypic tests takes weeks that make them inappropriate methods to evaluate a definitive diagnosis. PCR on the IS6110 gene is an accurate, quick, and inexpensive method for the M. tuberculosis detection. This study aimed to provide molecular diagnosis for the strains of M. tuberculosis resistant to the first-line drugs (rifampin and isoniazid) by IS6110 gene PCR. Materials and Methods: Of the 3012 Mycobacterium suspected cases, 60 samples of mycobacterium drug-resistant (MDR) were examined by smear, culture, phenotypic tests, and antibiogram. The isolates were identified by IS6110 gene amplification. Results & Conclusion: Among 60 tested samples, 54 (90%) positive samples from the M. tuberculosis group and 6 (10%) negative samples from non-tuberculosis group were reported. Out of 3012 clinical samples, 100 (3.3%) cultures were positive, 3 samples (5%) were resistant to rifampin, and the other samples (94%) were MDR. The IS6110 gene is one of the best markers for the Mycobacterium complex differentiation. This study showed that IS6110 gene PCR is a simple and accurate method for identifying drug resistance in M. tuberculosis clinical isolates. It is recommended to use molecular methods especially IS6110 gene sequencing instead of time-consuming with high variability phenotypic methods. [ABSTRACT FROM AUTHOR]

Tuberculosis is a significant source of illness and mortality worldwide. Therefore, tuberculosis screening is important to identify active cases within a community. Bacteriological examinations can be used to initiate active community-based case finding. Waste workers are at a high risk of contracting Mycobacterium tuberculosis due to their constant exposure to waste. This study aims to conduct bacteriological examinations to identify tuberculosis infection among waste workers in Gresik, Indonesia. An analytical observational study was carried out in September 2023 in Ngipik Village, Gresik, East Java, Indonesia. The primary study was conducted using a cross-sectional design with interviews, measurements, and sputum examinations for all samples. The screening involved a total of 72 waste workers (n = 72) who had daily contact with various types of sewage and waste, including medical waste. The majority of the waste workers were male (65.2%), most were between the ages of 34 and 44 (27.78%), and their educational background was a senior high school degree (40.27%). Most of the waste workers did not have a family history of hypertension (84.72%) or diabetes (97.22%). The acid-fast bacilli examinations were negative in all samples. Nevertheless, this study concluded that it is imperative to carry out mass tuberculosis screening in various settings, specifically in high-risk populations. Mass screening can detect the potential of latent tuberculosis, thereby reducing its transmission. [ABSTRACT FROM AUTHOR]

Purpose: This study investigates the potential of inflammatory parameters (IP), symptoms, and patient-related outcome measurements as biomarkers of severity and their ability to predict tuberculosis (TB) evolution. Methods: People with TB were included prospectively in the Stage-TB study conducted at five clinical sites in Barcelona (Spain) between April 2018 and December 2021. Data on demographics, epidemiology, clinical features, microbiology, and Sanit George Respiratory Questionnaire (SGRQ) and Kessler-10 as Health-Related Quality of Life (HRQoL) were collected at three time points during treatment. C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), neutrophil/lymphocyte, and monocyte/lymphocyte ratios (NLR and MLR), complement factors C3, C4, and cH50, clinical and microbiological data, and HRQoL questionnaires were assessed at baseline, 2 months, and 6 months. Their ability to predict sputum culture conversion (SCC) and symptom presence after 2 months of treatment was also analysed. Results: The study included 81 adults and 13 children with TB. The CRP, ESR, NLR, and MLR values, as well as the presence of symptoms, decreased significantly over time in both groups. Higher IP levels at baseline were associated with greater bacillary load and persistent symptoms. Clinical severity at baseline predicted a delayed SCC. Kessler-10 improved during follow-up, but self-reported lung impairment (SGRQ) persisted in all individuals after 6 months. Conclusions: IP levels may indicate disease severity, and sustained high levels are linked to lower treatment efficacy. Baseline clinical severity is the best predictor of SCC. Implementing health strategies to evaluate lung function and mental health throughout the disease process may be crucial for individuals with TB. [ABSTRACT FROM AUTHOR]

Background "Trace" results on Xpert MTB/RIF Ultra ("Ultra"; Cepheid) —a molecular diagnostic test for tuberculosis (TB)—are often interpreted as an indication for TB treatment, but may also represent detection of nonviable bacilli or analytical error. In community-screening settings where individual TB risk is low, there is limited guidance on how to interpret Ultra-trace results. Methods We conducted systematic Ultra TB screening of adults and adolescents (≥15 years) in Kampala, Uganda, through door-to-door and event-based sputum collection. We enrolled individuals with trace-positive sputum for detailed clinical, radiographic, and microbiological (including 2 sputum cultures, repeat Ultra, and for people with HIV, urine lipoarabinomannan) evaluation, and compared those findings with similar evaluations in controls with Ultra-negative and Ultra-positive (non-trace) sputum. Results Of 21 957 people screened with Ultra, 211 (1.0%) tested positive, including 96 (46% of positives) with trace results. Of 92 people enrolled with trace-positive sputum; 12% (11/92) were HIV-positive and 14% (13/92) had prior TB. The prevalence of TB among participants with trace-positive sputum results was 14% (13/92) by culture, 24% (22/92) using broader microbiological criteria, and 26% (24/92) after accounting for clinical diagnosis. The prevalence of cough and of abnormal chest computed tomography (CT) findings were 32% and 26%, respectively, if Ultra-negative; 34% and 54% if trace-positive/non–microbiologically confirmed; 72% and 95% if trace-positive/microbiologically confirmed; and 71% and 93% if Ultra-positive (more than trace). Conclusions Most individuals with trace-positive sputum in Ugandan communities did not have microbiologically confirmed TB but had more symptoms and chest CT abnormalities than people with Ultra-negative sputum. [ABSTRACT FROM AUTHOR]

Objective The aim of this study was to compare metagenomic next-generation sequencing (mNGS) with other methods, including Xpert MTB/RIF, Mycobacterium tuberculosis (MTB) culture, and acid-fast bacillus (AFB) staining in the diagnosis of pulmonary tuberculosis (PTB) using bronchoalveolar lavage fluid (BALF). Methods The data of 186 patients with suspected PTB were retrospectively collected from January 2020 to May 2021 at Tongji Hospital. BALF samples were collected from all patients and analyzed using AFB staining, MTB culture, Xpert MTB/RIF, and mNGS. Results Of the 186 patients, 38 patients were ultimately diagnosed as PTB. Metagenomic next-generation sequencing exhibited a sensitivity of 78.95%, which was higher than AFB staining (27.59%) and MTB culture (44.12%) but similar to Xpert MTB/RIF (72.73%). Utilization of combined methods demonstrates improvement for PTB diagnosis. In support of this, the area under the receiver operating characteristic curve for the combination of mNGS and MTB culture (0.933, 95% CI: 0.871, 0.995) was larger than those of mNGS, Xpert MTB/RIF, MTB culture, and the combination of Xpert MTB/RIF and MTB culture. Conclusion The sensitivity of mNGS in the diagnosis of PTB using BALF specimen is similar to Xpert MTB/RIF. Metagenomic next-generation sequencing in combination with MTB culture may further improve the diagnosis of pulmonary tuberculosis. [ABSTRACT FROM AUTHOR]

Bullous lung disease caused by tuberculosis is rare, and complications have a poor prognosis with uncertain pathophysiologic mechanisms. We describe a 29-year-old male patient who was admitted to the emergency department due to bilateral tension pneumothorax, which was complicated by bronchopleural fistula. This was managed with the placement of chest tubes, continuity of anti-TB drug treatment, and Heimlich valve placement. [ABSTRACT FROM AUTHOR]

Objectives: Tuberculosis (TB) and drug-resistant TB (DR-TB) are national health burdens in Vietnam. In this study, we investigated the prevalence of rifampicin (RIF) and/or isoniazid (isonicotinic acid hydrazide, INH) resistance in patients with suspected TB, and applied appropriate techniques to help rapidly target DR-TB. Methods: In total, 1,547 clinical specimens were collected and cultured using the BACTEC MGIT system (Becton Dickinson and Co.). A resazurin microtiter assay (REMA) was used to determine the proportions of RIF and/or INH resistance. A real-time polymerase chain reaction panel with TaqMan probes was employed to identify the mutations of rpoB and katG associated with DR-TB in clinical isolates. Genotyping of the identified mutations was also performed. Results: A total of 468 Mycobacterium tuberculosis isolates were identified using the REMA. Of these isolates, 106 (22.6%) were found to be resistant to 1 or both antibiotics. Of the resistant isolates, 74 isolates (69.8%) were resistant to isoniazid (INH) only, while 1 isolate (0.94%) was resistant to RIF only. Notably, 31 isolates (29.24%) were resistant to both antibiotics. Of the 41 phenotypically INH-resistant isolates, 19 (46.3%) had the Ser315Thr mutation. There were 8 different rpoB mutations in 22 (68.8%) of the RIF-resistant isolates. The most frequently detected mutations were at codons 531 (37.5%), 526 (18.8%), and 516 (6.3%). Conclusion: To help prevent new cases of DR-TB in Vietnam, it is crucial to gain a comprehensive understanding of the genotypic DR-TB isolates. [ABSTRACT FROM AUTHOR]

Key content: Tuberculosis (TB) is an important global cause of maternal and neonatal morbidity and mortality.With increasing migration, cases of tuberculosis in pregnancy or the puerperium are increasing in resource‐rich nations with advanced health care systems.Diagnosis may be delayed given the overlap of some of the symptoms with that of pregnancy.Timely diagnosis is essential to initiate appropriate treatment and prevent maternal and neonatal morbidity and mortality. Learning objectives: To understand the relevant global and UK epidemiology of the disease, including recent goals set by the World Health Organization and the UK.To review the pathophysiology of pulmonary and extrapulmonary TB, and of latent and active TB.To learn how to identify high‐risk women for antenatal assessment and gain an awareness of the screening tests available for latent TB.To gain awareness of the clinical symptoms, diagnosis and therapeutic management of active tuberculosis, including management of any drug resistance.To understand management of the neonate within high‐risk groups and where the mothers have diagnosed latent or active TB. Ethical issues: Pregnant women with TB should be fully counselled about the benefits, risks, and alternate options of management.Should healthcare professionals with high risk of acquiring TB infection be exempt from managing TB infection?Management of TB in HIV‐positive women provides additional challenges.Breastfeeding issues around seropositive mothers should be considered. [ABSTRACT FROM AUTHOR]

Background Halting transmission of Mycobacterium tuberculosis (Mtb) by identifying infectious individuals early is key to eradicating tuberculosis (TB). Here we evaluate face mask sampling as a tool for stratifying the infection risk of individuals with pulmonary TB (PTB) to their household contacts. Methods Forty-six sputum-positive PTB patients in The Gambia (August 2016–November 2017) consented to mask sampling prior to commencing treatment. Incident Mtb infection was defined in 181 of their 217 household contacts as QuantiFERON conversion or an increase in interferon-γ of ≥1 IU/mL, 6 months after index diagnosis. Multilevel mixed-effects logistical regression analysis with cluster adjustment by household was used to identify predictors of incident infection. Results Mtb was detected in 91% of PTB mask samples with high variation in IS6110 copies (5.3 × 102 to 1.2 × 107). A high mask Mtb level (≥20 000 IS6110 copies) was observed in 45% of cases and was independently associated with increased likelihood of incident Mtb infection in contacts (adjusted odds ratio, 3.20 [95% confidence interval, 1.26–8.12]; P =.01), compared with cases having low-positive/negative mask Mtb levels. Mask Mtb level was a better predictor of incident Mtb infection than sputum bacillary load, chest radiographic characteristics, or sleeping proximity. Conclusions Mask sampling offers a sensitive and noninvasive tool to support the stratification of individuals who are most infectious in high-TB-burden settings. Our approach can provide better insight into community transmission in complex environments. [ABSTRACT FROM AUTHOR]

Bacterial killing in patients with tuberculosis (TB) relapse was compared to that in patients achieving cure, measured by TB molecular bacterial load assay (TB-MBLA) or mycobacteria growth indicator tube (MGIT) time to positivity (TTP). TB-MBLA in 4 relapsed patients was significantly different compared to 132 cured patients after 2 weeks of treatment; MGIT TTP showed a significant difference from week 8. [ABSTRACT FROM AUTHOR]

Crosstalk between cell death programs confers appropriate host anti-infection immune responses, but how pathogens co-opt host molecular switches of cell death pathways to reprogram cell death modalities for facilitating infection remains largely unexplored. Here, we identify mammalian cell entry 3C (Mce3C) as a pathogenic cell death regulator secreted by Mycobacterium tuberculosis (Mtb), which causes tuberculosis featured with lung inflammation and necrosis. Mce3C binds host cathepsin B (CTSB), a noncaspase protease acting as a lysosome-derived molecular determinant of cell death modalities, to inhibit its protease activity toward BH3-interacting domain death agonist (BID) and receptor-interacting protein kinase 1 (RIPK1), thereby preventing the production of proapoptotic truncated BID (tBID) while maintaining the abundance of pronecroptotic RIPK1. Disrupting the Mce3C-CTSB interaction promotes host apoptosis while suppressing necroptosis with attenuated Mtb survival and mitigated lung immunopathology in mice. Thus, pathogens manipulate host lysosomal protease activity-dependent plasticity in cell death modalities to promote infection and pathogenicity., Competing Interests: Competing interests statement:The authors declare no competing interest.

The bacterial genus Streptomyces is known for the prolific production of secondary metabolites, which exhibit remarkable structural diversity and potent biological activities. Tuberculosis (TB) remains a formidable global health challenge exacerbated by the emergence of antimicrobial resistance (AMR), necessitating the discovery of novel therapeutic agents. The untapped potential of marine Streptomyces-derived secondary metabolites offers a promising avenue for screening anti-tubercular (anti-TB) compounds with unique chemical structures and potential bioactive properties. The review emphasizes the diverse marine habitats and Streptomyces with novel anti-TB bioactive metabolites. It discusses fermentation and bioprocessing strategies for screening anti-TB drugs. This review also covers the chemical diversity, potency, mechanism of action, and structures of about seventy anti-TB compounds discovered from marine Streptomyces. These compounds span various chemical classes, including quinones, macrolactams, macrolides, phenols, esters, anthracyclines, peptides, glycosides, alkaloids, piperidones, thiolopyrrolones, nucleosides, terpenes, flavonoids, polyketides, and actinomycins. It emphasizes the need to explore marine ecosystems to discover more novel anti-TB natural products., Competing Interests: Declarations. Conflict of interest: The authors declare no conflict of interest. Ethical approval: This article contains no studies performed by authors with human participants or animals., (© 2025. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Numerous studies have identified similarities in blood transcriptomic signatures of tuberculosis (TB) phenotypes between mice and humans, including type 1 interferon production and innate immune cell activation. However, murine infection pathophysiology is distinct from human disease. We hypothesized that this is partly due to differences in the relative importance of biological pathways across species. To address this animal-to-human gap, we applied a systems modeling framework, Translatable Components Regression, to identify the axes of variation in the preclinical data most relevant to human TB disease state. Among the pathways our cross-species model pinpointed as highly predictive of human TB phenotype was the infection-induced unfolded protein response. To validate this mechanism, we confirmed that this cellular stress pathway modulates immune functions in Mycobacterium tuberculosis-infected mouse macrophages. Our work demonstrates how systems-level computational models enhance the value of animal studies for elucidating complex human pathophysiology., Competing Interests: Competing interests: The authors have declared that no conflict of interest exists. Any opinion, findings, and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of the National Science Foundation or the Department of Defense., (© 2025. The Author(s).)

The type II NADH-dehydrogenase enzyme in Mycobacterium tuberculosis plays a critical role in the efficient functioning of the oxidative phosphorylation pathway. It acts as the entry point for electrons in the electron transport chain, which is essential for fulfilling the energy requirements of both replicating and nonreplicating mycobacterial species. Due to the absence of the type II NADH-dehydrogenase enzyme in mammalian mitochondria, targeting the type II NADH-dehydrogenase enzyme for antitubercular drug discovery could be a vigilant approach. Utilizing type II NADH-dehydrogenase inhibitors in antitubercular therapy led to bactericidal response, even in monotherapy. However, the absence of the cryo-EM structure of Mycobacterium tuberculosis type II NADH-dehydrogenase has constrained drug discovery efforts to rely on high-throughput screening methods, limiting the use of structure-based drug discovery. Here, we have delineated the literature-reported Mycobacterium tuberculosis type II NADH-dehydrogenase inhibitors and the rationale behind selecting this specific enzyme for antitubercular drug discovery, along with shedding light on the architecture of the enzyme structure and functionality. The gap in the current research and future research direction for TB treatment have been addressed.

Tuberculosis (TB), a leading infectious disease caused by the pathogen Mycobacterium tuberculosis , poses a significant treatment challenge due to its unique characteristics and resistance to existing drugs. The conventional treatment regimens, which are lengthy and involve multiple drugs, often result in poor patient adherence and subsequent drug resistance, particularly with multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains. This highlights the urgent need for novel anti-TB therapies and new drug targets. Antimicrobial peptides (AMPs), which are natural host defense molecules present in all living organisms, offer a promising alternative to traditional small-molecule drugs. AMPs have several advantages, including their broad-spectrum activity and the potential to circumvent existing resistance mechanisms. However, their clinical application faces challenges such as stability, delivery, and potential toxicity. This review aims to provide essential information on AMPs, including their sources, classification, mode of action, induction within the host under stress, efficacy against M. tuberculosis , clinical status and hurdles to their use. It also highlights future research directions to address these challenges and advance the development of AMP-based therapies for TB.

Developing new classes of drugs that are active against infections caused by Mycobacterium tuberculosis is a priority for treating and managing this deadly disease. Here, we describe screening a small library of 20 DNA gyrase inhibitors and identifying new lead compounds. Three structurally diverse analogues were identified with minimal inhibitory concentrations of 0.125 μg/mL against both drug-susceptible and drug-resistant strains of M. tuberculosis . These lead compounds also demonstrated antitubercular activity in ex vivo studies using infected THP-1 macrophages with minimal cytotoxicity in THP-1, HeLa, and HepG2 cells (IC 50 ≥ 128 μg/mL). The molecular target of the lead compounds was validated through biochemical studies of select analogues with purified M. tuberculosis gyrase and the generation of resistant mutants. The lead compounds were assessed in combination with bedaquiline and pretomanid to determine the clinical potential, and the select lead ( 158 ) demonstrated in vivo efficacy in an acute model of TB infection in mice, reducing the lung bacterial burden by approximately 3 log 10 versus untreated control mice. The advancement of DNA gyrase inhibitors expands the field of innovative therapies for tuberculosis and may offer an alternative to fluoroquinolones in future therapeutic regimens.

Heterogeneity during Mycobacterium tuberculosis (Mtb) infection greatly impacts disease outcome and complicates treatment. This heterogeneity encompasses many facets, spanning local differences in the host immune response to Mtb and the environment experienced by the bacterium, to nonuniformity in Mtb replication state. All of these facets are interlinked and each can affect Mtb susceptibility to antibiotic treatment. In-depth spatiotemporal understanding of Mtb-host interactions is thus critical to both fundamental comprehension of Mtb infection biology and for the development of effective therapeutic regimens. Such spatiotemporal understanding dictates the need for analysis at the single bacterium/cell level in the context of intact tissue architecture, which has been a significant technical challenge. Excitingly, innovations in spatial single cell methodology have opened the door to such studies, beginning to illuminate aspects ranging from intergranuloma differences in cellular composition and phenotype, to sublocation differences in Mtb physiology and replication state. In this perspective, we discuss recent studies that demonstrate the potential of these methodological advancements to reveal critical spatiotemporal insight into Mtb-host interactions, and highlight future avenues of research made possible by these advances.

Tuberculosis (TB) continues to be a major cause of death worldwide despite having an effective combinatorial therapeutic regimen and vaccine. Being one of the most successful human pathogens, Mycobacterium tuberculosis retains the ability to adapt to diverse intracellular and extracellular environments encountered by it during infection, persistence, and transmission. Designing and developing new therapeutic strategies to counter the emergence of multidrug-resistant and extensively drug-resistant TB remains a major task. DNA topoisomerases make up a unique class of ubiquitous enzymes that ensure steady-state level supercoiling and solve topological problems occurring during DNA transactions in cells. They continue to be attractive targets for the discovery of novel classes of antibacterials and to develop better molecules from existing drugs by virtue of their reaction mechanism. The limited repertoire of topoisomerases in M. tuberculosis , key differences in their properties compared to topoisomerases from other bacteria, their essentiality for the pathogen's survival, and validation as candidates for drug discovery provide an opportunity to exploit them in drug discovery efforts. The present review provides insights into their organization, structure, function, and regulation to further efforts in targeting them for new inhibitor discovery. First, the structure and biochemical properties of DNA gyrase and Topoisomerase I (TopoI) of mycobacteria are described compared to the well-studied counterparts from other bacteria. Next, we provide an overview of known inhibitors of DNA gyrase and emerging novel bacterial topoisomerase inhibitors (NBTIs). We also provide an update on TopoI-specific compounds, highlighting mycobacteria-specific inhibitors.

A major challenge in tuberculosis (TB) therapeutics is that antibiotic exposure leads to changes in the physiology of M. tuberculosis ( Mtb ), which may enable the pathogen to withstand treatment. While antibiotic-treated Mtb has been evaluated in in vitro experiments , it is unclear if and how long-term in vivo treatment with diverse antibiotics with varying treatment-shortening activity (sterilizing activity) affects Mtb physiologic processes differently. Here, we used SEARCH-TB, a pathogen-targeted RNA-sequencing platform, to characterize the Mtb transcriptome in the BALB/c high-dose aerosol infection mouse model following 4 weeks of treatment with three sterilizing and three non-sterilizing antibiotics. Certain transcriptional changes were shared among most antibiotics, including decreased expression of genes associated with protein synthesis and metabolism and the induction of certain genes associated with stress responses. However, the magnitude of this shared response differed between antibiotics. Sterilizing antibiotics rifampin, pyrazinamide, and bedaquiline generated a more quiescent Mtb state than did non-sterilizing antibiotics isoniazid, ethambutol, and streptomycin, as indicated by the decreased expression of genes associated with translation, transcription, secretion of immunogenic proteins, metabolism, and cell wall synthesis. Additionally, we identified distinguishing transcriptional effects specific to each antibiotic, indicating that different mechanisms of action induce distinct patterns in response to cellular injury. In addition to elucidating the Mtb physiologic changes associated with antibiotic stress, this study demonstrates the value of SEARCH-TB as a highly granular pharmacodynamic assay that reveals antibiotic effects that are not apparent based on culture alone., Competing Interests: The authors declare no conflict of interest.

Background: In tuberculosis (TB) infection, monocytes play a crucial role in regulating the balance between immune tolerance and immune response through various mechanisms. A deeper understanding of the roles of monocyte subsets in TB immune responses may facilitate the development of novel immunotherapeutic strategies and improve TB prevention and treatment., Methods: We retrieved and processed raw single-cell RNA-seq data from SRP247583. Single-cell RNA-seq combined with bioinformatics analysis was employed to investigate the roles of monocytes in TB progression., Results: Our findings revealed that classical monocytes expressing inflammatory mediators increased as the disease progressed, whereas non-classical monocytes expressing molecules associated with anti-pathogen infection were progressively depleted. Pseudotime analysis delineated the differentiation trajectory of monocytes from classical to intermediate to non-classical subsets. An abnormal differentiation trajectory to non-classical monocytes may represent a key mechanism underlying TB pathogenesis, with CEBPB and CORO1A identified as genes potentially related to TB development. Analysis of key transcription factors in non-classical monocytes indicated that IRF9 was the only downregulated transcription factor with high AUC activity in this subset. The expression of IRF9 exhibited a decreasing trend in both latent TB infection (LTBI) and active TB groups. Furthermore, dysregulation of transcription factor regulatory networks appeared to impair ferroptosis, with ferroptosis-associated genes MEF2C, MICU1, and PRR5 identified as potential targets of IRF9. Through cell communication analysis, we found that interactions between non-classical monocytes and other subpopulations may mediate TB progression, with MIF and LGALS9 highlighted as potential signaling pathways., Conclusion: This study employs bioinformatics analysis in conjunction with single-cell sequencing technology to uncover the crucial role of monocyte subsets in tuberculosis infection., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Reconstructing transmission networks is essential for identifying key factors like superspreaders and high-risk locations, which are critical for developing effective pandemic prevention strategies. This study presents a Bayesian transmission model that combines genomic and temporal data to reconstruct transmission networks for infectious diseases. The Bayesian transmission model incorporates the latent period and distinguishes between symptom onset and actual infection time, improving the accuracy of transmission dynamics and epidemiological models. It also assumes a homogeneous effective population size among hosts, ensuring that the coalescent process for within-host evolution remains unchanged, even with missing intermediate hosts. This allows the model to effectively handle incomplete samples. Simulation results demonstrate the model's ability to accurately estimate model parameters and transmission networks. Additionally, our proposed hypothesis test can reliably identify direct transmission events. The Bayesian transmission model was applied to a real dataset of Mycobacterium tuberculosis genomes from 69 tuberculosis cases. The estimated transmission network revealed two major groups, each with a superspreader who transmitted M. tuberculosis, either directly or indirectly, to 28 and 21 individuals, respectively. The hypothesis test identified 16 direct transmissions within the estimated network, demonstrating the Bayesian model's advantage over a fixed threshold by providing a more flexible criterion for identifying direct transmissions. This Bayesian approach highlights the critical role of genetic data in reconstructing transmission networks and enhancing our understanding of the origins and transmission dynamics of infectious diseases., Competing Interests: Declarations. Conflict of interest: The authors declare no conflict of interest., (© 2025. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

This study leverages the Bacterial and Viral Bioinformatics Resource Center (BV-BRC) to analyze over 27,000 Mycobacterium tuberculosis (MTB) genomic strains, providing a comprehensive and large-scale overview of antibiotic resistance (AMR) prevalence and resistance patterns. We used MTB++, which is the newest and most comprehensive AI-based MTB drug resistance profiler tool, to predict the resistance profile of each of the 27,000 MTB isolates and then used feature analysis to identify key genes that were associated with the resistance. There are three main contributions to this study. Firstly, it provides a detailed picture of the prevalence of specific AMR genes in the BV-BRC dataset as well as their biological implications, providing critical insight into MTB's resistance mechanisms that can help identify genes of high priority for further investigation. The second aspect of this study is to compare the prevalence of antibiotic resistance across previous studies that have addressed both the temporal and geographical evolution of MTB drug resistance. Lastly, this study emphasizes the need for targeted diagnostics and personalized treatment plans. In addition to these contributions, the study acknowledges the limitations of computational prediction and recommends future experimental validation., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Tuberculosis (TB), caused by Mycobacterium tuberculosis , remains a significant global health threat, affecting an estimated 10.6 million people in 2022. The emergence of multidrug resistant and extensively drug resistant strains necessitates the development of novel and effective drugs. Accelerating the determination of mechanisms of action (MOAs) for these drugs is crucial for advancing TB treatment. This study introduces MycoBCP, a unique adaptation of bacterial cytological profiling (BCP) tailored to M. tuberculosis , utilizing the application of convolutional neural networks (CNNs) within BCP to overcome challenges posed by traditional image analysis techniques. Using MycoBCP, we analyzed the morphological effects of various antimicrobial compounds on M. tuberculosis , capturing broad patterns rather than relying on precise cell segmentation. This approach circumvented issues such as cell clumping and uneven staining, which are prevalent in M. tuberculosis . In a blind test, MycoBCP accurately identified the MOA for 96% of the compounds, with a single misclassification of rifabutin, which was incorrectly categorized as affecting translation rather than transcription. The similar morphologies resulting from transcription and translation inhibition indicate a need for further refinement to distinguish them more effectively. Application of MycoBCP to a series of antitubercular agents successfully identified known MOAs and revealed unique effects, demonstrating its utility in early drug discovery and development. Our findings underscore the potential of CNN-based BCP to enhance the accuracy and efficiency of MOA determination, particularly for challenging pathogens like M. tuberculosis . MycoBCP represents a significant advancement in TB drug development, offering a robust and adaptable method for high-throughput screening of antimicrobial compounds., Competing Interests: Competing interests statement:D.Q., M.S., and J.S. are employed by Linnaeus Bioscience, Inc. J.P. has an equity interest in Linnaeus Bioscience Incorporated and receive consulting income from the company. The terms of this arrangement have been reviewed and approved by the University of California, San Diego in accordance with its conflict-of-interest policies.

Accelerated tuberculosis drug discovery has increased the number of plausible multidrug regimens. Testing every drug combination in vivo is impractical, and varied experimental conditions make it challenging to compare results between experiments. Using published treatment efficacy data from a mouse tuberculosis model treated with candidate combination regimens, we trained and externally validated integrative mathematical models to predict relapse in mice and to rank both previously experimentally studied and unstudied regimens by their sterilization potential. We generated 18 datasets of 18 candidate regimens (comprising 11 drugs of six classes, including fluoroquinolone, nitroimidazole, diarylquinolines, and oxazolidinones), with 2965 relapse and 1544 colony-forming unit (CFU) observations for analysis. Statistical and machine learning techniques were applied to predict the probability of relapse in mice. The locked down mathematical model had an area under the receiver operating characteristic curve (AUROC) of 0.910 and showed that bacterial kill measured by longitudinal CFU cannot account for relapse alone and that sterilization is drug dependent. The diarylquinolines had the highest predicted sterilizing activity in the mouse model, and the addition of pyrazinamide to drug regimens provided the shortest estimated tuberculosis treatment duration to cure in mice. The mathematical model predicted the effect of treatment combinations, and these predictions were validated by conducting 11 experiments on previously unstudied regimens, achieving an AUROC of 0.829. We surmise that the next generation of tuberculosis drugs are highly effective at treatment shortening and suggest that there are several promising three- and four-drug regimens that should be advanced to clinical trials.

Background: An accurate, rapid, non-sputum-based triage test for diagnosing tuberculosis (TB) is needed., Methods: A prospective evaluation of the Cepheid GeneXpert Mycobacterium tuberculosis Host Response cartridge (Xpert-MTB-HR), a prototype blood-based host response mRNA signature assay, among individuals presenting with TB-like symptoms was performed in Pakistan and results were compared to 3 reference standards: Xpert MTB/RIF Ultra, bacteriological confirmation (Xpert MTB/RIF Ultra and/or culture positivity), and composite clinical diagnosis (clinician diagnosis, treatment initiation, Xpert MTB/RIF Ultra, and/or culture positivity). Analyses were conducted both for the entire study cohort and separately in the adolescent and young adult cohort (aged 10-24 years)., Results: A total of 497 participants, aged 6-83 years, returned valid Xpert-MTB-HR results. When a diagnostic threshold was set for a sensitivity of >90%, specificity was 32% (95% confidence interval [CI], 28%-37%) compared to Xpert MTB/RIF Ultra, 29% (95% CI, 25%-34%) compared to a bacteriological confirmation, and 22% (95% CI, 18%-26%) compared to a composite clinical diagnosis. However, when evaluating only the adolescent and young adult cohort with a diagnostic threshold set for sensitivity of >90%, specificity was 82% (95% CI, 74%-89%) compared to Xpert MTB/RIF Ultra, 84% (95% CI, 75%-90%) compared to a bacteriological confirmation, and 54% (95% CI, 44%-64%) compared to a composite clinical diagnosis., Conclusions: While the Xpert-MTB-HR does not meet World Health Organization minimum criteria in the general population, in our study it does meet the minimum sensitivity and specificity requirements for a non-sputum-based triage test among adolescents and young adults when compared to Xpert MTB/RIF Ultra or bacteriological confirmation., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Background: Tuberculosis (TB) is among the largest infectious causes of death worldwide, and there is a need for a time- and resource-effective diagnostic methods. In this novel and exploratory study, we show the potential of using buccal swabs to collect human DNA and investigate the DNA methylation (DNAm) signatures as a diagnostic tool for TB., Methods: Buccal swabs were collected from patients with pulmonary TB (n = 7), TB-exposed persons (n = 7), and controls (n = 9) in Sweden. Using Illumina MethylationEPIC array, the DNAm status was determined., Results: We identified 5644 significant differentially methylated CpG sites between the patients and controls. Performing the analysis on a validation cohort of samples collected in Kenya and Peru (patients, n = 26; exposed, n = 9; control, n = 10) confirmed the DNAm signature. We identified a TB consensus disease module, significantly enriched in TB-associated genes. Last, we used machine learning to identify a panel of 7 CpG sites discriminative for TB and developed a TB classifier. In the validation cohort, the classifier performed with an area under the curve of 0.94, sensitivity of 0.92, and specificity of 1., Conclusions: In summary, the result from this study shows clinical implications of using DNAm signatures from buccal swabs to explore new diagnostic strategies for TB., Competing Interests: Potential conflicts of interest. M. L. and M. G. are founders of PredictME AB. S. S. and D. M.-E. are bioinformaticians at PredictME. All other authors report no potential conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Background: Understanding the etiology of recurrent tuberculosis (rTB) is important for effective tuberculosis control. Prior to the advent of whole genome sequencing (WGS), attributing rTB to relapse or reinfection using genetic information was complicated by the limited resolution of conventional genotyping methods., Methods: We applied a systematic method of evaluating whole genome single-nucleotide polymorphism (wgSNP) distances and results of phylogenetic analyses to characterize the etiology of rTB in American Indian and Alaska Native (AIAN) persons in Alaska during 2008 to 2020. We contextualized our findings through descriptive analyses of surveillance data and results of a literature search for investigations that characterized rTB etiology using WGS., Results: The percentage of tuberculosis cases in AIAN persons in Alaska classified as recurrent episodes (11.8%) was 3 times the national percentage (3.9%). Of 38 recurrent episodes included in genetic analyses, we attributed 25 (65.8%) to reinfection based on wgSNP distances and phylogenetic analyses; this proportion was the highest among 16 published point estimates identified through the literature search. By comparison, we attributed 11 (28.9%) and 6 (15.8%) recurrent episodes to reinfection based on wgSNP distances alone and on conventional genotyping methods, respectively., Conclusions: WGS and attribution criteria involving genetic distances and patterns of relatedness can provide an effective means of elucidating rTB etiology. Our findings indicate that rTB occurs at high proportions among AIAN persons in Alaska and is frequently attributable to reinfection, reinforcing the importance of active surveillance and control measures to limit the spread of tuberculosis disease in Alaskan AIAN communities., Competing Interests: Potential conflicts of interest. All authors: No reported conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (Published by Oxford University Press on behalf of Infectious Diseases Society of America 2024.)

Tuberculosis (TB) is the leading cause of infectious disease death and lacks a vaccine capable of protecting adults from pulmonary TB. The bacterial surface is a critical interface that shapes host-pathogen interactions. Several knowledge gaps persist in our understanding of Mycobacterium tuberculosis (Mtb)-host interactions that may be addressed by an improved understanding of the Mtb surface proteome, including the identification of novel vaccine targets as well as developing new approaches to interrogate host-pathogen interactions. Here, we sought to expand our understanding of the Mtb surface proteome in service of these knowledge gaps by adapting protease shaving protocols for multiplexed quantitative mass spectrometry. Pairing quantitative mass spectrometry with the construction of validation strains, we revealed several novel Mtb proteins on the Mtb surface largely derived from the PE/PPE class of Mtb proteins, including PPE18, a component of a leading Mtb vaccine candidate. We next exploited the localization of PPE18 to decorate the Mtb surface with heterologous proteins. Together, these studies reveal potential novel targets for new Mtb vaccines as well as facilitate new approaches to study difficult-to-study cellular compartments during bacterial growth and infection.IMPORTANCEThe surface of a bacterial pathogen is a critical interface between the bacterium and the immune system. A better understanding of this interface would facilitate the discovery of new vaccine targets, new virulence proteins, and enable new technologies that modify the bacterial surface. In this study, we established a multiplexed and quantitative biochemical strategy to study the surface of Mycobacterium tuberculosis (Mtb) and identified new vaccine targets. We furthermore established design rules for new technologies aimed at modifying the composition of the bacterial surface. Specifically, we achieved a biological milestone that has not been rigorously reported previously, which is the successful modification of the Mtb surface with a non-native protein., Competing Interests: The authors declare no conflict of interest.

Background: The lengthy duration and high frequency of drug resistance associated with currently used antimycobacterial drug treatments have intensified the need for alternative therapies against Mycobacterium tuberculosis, the causative agent of TB., Methods: MICs and intracellular macrophage cfu counts were tested to evaluate the antibacterial activity of nintedanib and pirfenidone against drug-susceptible and -resistant M. tuberculosis. A chronic murine model of pulmonary infection was used to assay the therapeutic efficacy of nintedanib. Macrophage transcriptome deep sequencing, a confocal assay, siRNA knockdown, Western blotting, quantitative RT-PCR and a cfu assay were used to investigate the antibacterial mechanism of nintedanib., Results: The MIC90 of nintedanib against M. tuberculosis standard strain H37Rv was 23.56-40.51 mg/L. TB murine model studies showed that nintedanib, coadministered with isoniazid, rifampicin and pyrazinamide, shortened treatment duration, and ameliorated pulmonary inflammation and fibrosis. In mechanism studies, transcriptome sequencing analysis revealed that nintedanib may eliminate M. tuberculosis through up-regulating macrophage autophagy. Furthermore, inhibition of autophagy by using siRNA targeting ATG5 or the autophagy inhibitor 3-methyladenine almost completely abolished nintedanib-mediated suppression of M. tuberculosis. Nintedanib induced autophagy by the JAK2/STAT3/Beclin1 pathway. When JAK2 or Beclin1 were knocked down through siRNA, nintedanib no longer inhibited M. tuberculosis. JAK2 activator coumermycin A1 and STAT3 agonist colivelin also reversed this phenotype., Conclusions: In vitro activity of nintedanib against drug-susceptible and -resistant M. tuberculosis and efficacy in murine infections warrant the continued clinical evaluation of nintedanib as a new adjuvant therapy for standard treatment of TB., (© The Author(s) 2024. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Mycobacterium tuberculosis enters the body through the respiratory tract, produces and releases virulence proteins through a variety of mechanisms, regulates the host immune mechanism through a variety of ways, and then survives in the body for a long time. These depend on virulence genes encoded by Mycobacterium tuberculosis. Previous studies found that the Rv3435c gene of Mycobacterium tuberculosis is highly conserved in pathogenic mycobacterium, but not conserved in non-pathogenic mycobacterium, which may be a potential virulence gene, and inhibit the secretion of inflammatory factors in RAW264.7 cells and inhibit cell apoptosis. Based on previous studies, the function of Rv3435c gene in mice was studied by infecting mice with recombinant strains. In vivo infection experiments showed that overexpression of Rv3435c significantly promoted the survival of Ms in the lung. Ms-pMV361-Rv3435c specifically inhibits the secretion of inflammatory cytokines, including TNF-α, IL-6, IL-1β, IL-12, and IFN-γ. Rv3435c can inhibit lung cell apoptosis and cause pathological damage to lung. Therefore, Rv3435c enhances the survival of mycobacterium in mice and promotes the pathogenicity and spread of Mycobacterium tuberculosis by inhibiting the production of cytokines., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2025 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The mortality caused by tuberculosis (TB) infections is a global concern, and there is a need to improve understanding of the disease. Current in vitro infection models to study the disease have limitations such as short investigation durations and divergent transcriptional signatures. This study aims to overcome these limitations by developing a 3D collagen culture system that mimics the biomechanical and extracellular matrix (ECM) of lung microenvironment (collagen fibers, stiffness comparable to in vivo conditions) as the infection primarily manifests in the lungs. The system incorporates Mycobacterium tuberculosis (Mtb) infected human THP-1 or primary monocytes/macrophages. Dual RNA sequencing reveals higher mammalian gene expression similarity with patient samples than 2D macrophage infections. Similarly, bacterial gene expression more accurately recapitulates in vivo gene expression patterns compared to bacteria in 2D infection models. Key phenotypes observed in humans, such as foamy macrophages and mycobacterial cords, are reproduced in the model. This biomaterial system overcomes challenges associated with traditional platforms by modulating immune cells and closely mimicking in vivo infection conditions, including showing efficacy with clinically relevant concentrations of anti-TB drug pyrazinamide, not seen in any other in vitro infection model, making it reliable and readily adoptable for tuberculosis studies and drug screening., (© 2024 Wiley‐VCH GmbH.)

According to the World Health Organization, Mycobacterium tuberculosis infections affect approximately 25% of the world's population. There is mounting evidence linking autophagy and immunological dysregulation to tuberculosis (TB). As a result, this research set out to discover TB-related autophagy-related biomarkers and prospective treatment targets. We used five autophagy databases to get genes linked to autophagy and Gene Expression Omnibus databases to get genes connected to TB. Then, functional modules associated with autophagy were obtained by analyzing them using weighted gene co-expression network analysis. Both Gene Ontology and Kyoto Encyclopedia of Genes and Genomes were used to examine the autophagy-related genes (ATGs) of important modules. Limma was used to identify differentially expressed ATGs (DE-ATGs), and the external datasets were used to further confirm their identification. We used DE-ATGs and a protein-protein interaction network to search the hub genes. CIBERSORT was used to estimate the kinds and amounts of immune cells. After that, we built a drug-gene interaction network and a network that included messenger RNA, small RNA, and DNA. At last, the differential expression of hub ATGs was confirmed by RT-qPCR, immunohistochemistry, and western blotting. The diagnostic usefulness of hub ATGs was evaluated using receiver operating characteristic curve analysis. Including 508 ATGs, four of the nine modules strongly linked with TB were deemed essential. Interleukin 1B ( IL1B ), CAPS1 , and signal transducer and activator of transcription 1 ( STAT1 ) were identified by intersection out of 22 DE-ATGs discovered by differential expression analysis. Research into immune cell infiltration found that patients with TB had an increased proportion of plasma cells, CD8 T cells, and M0 macrophages. A competitive endogenous RNA network utilized 10 long non-coding RNAs and 2 miRNAs. Then, the IL1B -targeted drug Cankinumad was assessed using this network. During bioinformatics analysis, three hub genes were validated in mouse and macrophage infection models. We found that IL1B , CASP1 , and STAT1 are important biomarkers for TB. As a result, these crucial hub genes may hold promise as TB treatment targets.

Stringent sanitary standards are imperative for swine production, ensuring high biosecurity and safe meat. However, granulomatous lesions, often detected as "lymphadenitis" in slaughterhouses, lack routine laboratory examination, potentially overlooking tuberculosis among other etiologies. This study aimed to: (i) explore epidemiological variables linked to swine carcasses condemned due to "tuberculosis" or "lymphadenitis" in Rio Grande do Sul, southern Brazil in a retrospective survey; and (ii) evaluate the frequency of Mycobacterium tuberculosis in granulomatous lymphadenitis lesions. Epidemiological data from the public surveillance system including farm origin, slaughter location, season, producer type and the inspection scope, were evaluated by their association with the occurrence of animal transport guides (ATG) with at least one animal condemned due to lymphadenitis or tuberculosis. For the prospective study, tissue samples from 118 animals with granulomatous lymphadenitis lesions were examined through histopathology and bacterial isolation. The frequency of ATG with at least one carcass condemned due to lymphadenitis or tuberculosis was 2.27 % and 0.027 % respectively. The factors associated with lymphadenitis were slaughter location, origin, producer type, and inspection scope (state or municipal). However, no significant risk factors emerged for tuberculosis in multivariable analysis. Histopathology confirmed granulomatous lymphadenitis in 44.92 % of lymph nodes, and M. tuberculosis var. bovis was isolated in 2.54 % of cases. Although the tuberculosis frequency (0.002 %) was low, it surpassed federal inspection data (<0.001 %), evidencing that laboratory diagnosis should be considered in such lesions. This study underscores the need for improved diagnostic routine in slaughterhouses to enhance biosecurity and public health protection in the swine industry., Competing Interests: Declaration of Competing Interest None., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Elucidating the pathogenic mechanism of Tuberculosis (TB) can contribute to control TB. Basic leucine zipper transcription factor ATF-like 2 (BATF2) belonging to a large family of leucine zipper transcription factors (TFs) termed bZip proteins, had been verified to have important value in the diagnosis of TB. However, its role and mechanism in TB had not been elucidated. The study aimed to explore its function and molecular mechanism in macrophages infected with Mycobacterium tuberculosis (Mtb). The results indicated that BATF2 inhibited cell proliferation, promoted inflammatory response and impaired the antibacterial and antigen-presenting capacity in macrophages for T cells through regulating its downstream gene TTC23 by interacting with SINHCAF. Above roles and regulations were dependent on β-catenin functions in macrophages infected with Mtb. Clinical samples verified that the expressions of BATF2 and TTC23 were significantly higher in the blood of patients with pulmonary TB compared with health controls. Altogether, BATF2 interacted with SINHCAF to regulate the quantity and function of macrophages during Mtb infection by targeting TTC23 through Wnt/β-catenin pathway., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Tuberculosis (TB) is a chronic respiratory infectious disease and is induced by Mycobacterium tuberculosis (M.tb) infection. Macrophages serve as the cellular home in immunoreaction against M.tb infection, which is tightly regulated through Toll-like receptor 4 (TLR4) expression. Therefore, this study is designed to explore the role and mechanism of TLR4 in mycobacterial injury in human macrophages (THP-1 cells) after M.tb infection. Cell proliferation and apoptosis were assessed using MTT, EdU, and flow cytometry assays. ELISA kits were utilized to assess the levels of Interleukin-6 (IL-6), IL-1β, and tumor necrosis factor α (TNF-α). The binding between Yin-Yang-1 (YY1) and TLR4 promoter was predicted by JASPAR and verified using Chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assays. M.tb infection might repress THP-1 cell proliferation, and induce cell apoptosis and inflammatory response in a multiplicity of infection (MOI)-dependent manner. Moreover, M.tb infection increased the expression of TLR4 in HTP-1 cells in an MOI-dependent way, and its downregulation might overturn M.tb infection-mediated HTP-1 cell damage and inflammatory response. At the molecular level, YY1 was a transcription factor of TLR4 and promoted TLR4 transcription via binding to its promoter region. Besides, YY1 might activate the NF-kB signaling pathway via regulating TLR4. Meanwhile, TLR4 inhibitor BAY11-7082 might overturn the repression effect of TLR4 on M.tb-infected HTP-1 cell damage. YY1-activated TLR4 might aggravate mycobacterial injury in human macrophages after M.tb infection by the NF-kB pathway, providing a promising therapeutic target for TB treatment., Competing Interests: Declarations. Conflict of interest: The authors declare that they have no conflicts of interest., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Even though anti-tuberculosis (TB) treatment is readily available, Mycobacterium tuberculosis (Mtb) infection continues to be a global threat with a high death rate recorded from a single infectious agent. This highlights the significance of developing new strategies to curb the growing Mtb infection cases. Host-directed therapies (HDT) offer a promising approach that includes both drug discovery and drug repurposing, aimed at identifying host targets and promoting immune cell populations that can lead to better infection outcomes. In this context, we investigated the potential of exogenous Binding Immunoglobulin Protein (BiP) to induce such changes ex-vivo using PBMCs from healthy (QFN-) and Mtb exposed (QFN+) individuals. We analysed cell surface expression and cytokine profiles across eight different stimulation conditions including human full-length BiP protein (20 μg/ml), TLR-9a (0.5 μM), BiP/TLR-9a combination, isoniazid (1 μM), H37Rv (MOI: 1: 10), and pooled bronchoalveolar lavage (BAL) samples collected at TB diagnosis (TBdx) and at month 6 (M6) of anti-TB treatment. Our results revealed that BiP-stimulated PBMCs showed a significant reduction of interleukin (IL)-10 secretion, along with increased IL-4, IL-5, IL-13, and soluble Fas-L (sFasL) secretion. We also observed that BiP stimulation enhanced the expression of membrane bound Fas-L (CD178) and IL5Ra (CD125) in B-cells isolated from both QFN- and QFN+ groups. Additionally, BiP exhibited a synergistic effect with TLR-9a, further boosting this co-expression. Moreover, we observed that BiP induced IL5Ra expression in both CD3 + CD5 lo and CD3 + CD5 hi T-cell populations. This study explores the effects of exogenous BiP on cell functionality and provides valuable insights into its potential to modulate host cell responses, which could be explored as a host-directed therapy for TB in the future., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Bongani Motaung reports financial support was provided by National Research Foundation. Andre G Loxton reports financial support was provided by National Research Foundation. Andre G Loxton reports financial support was provided by European and Developing Countries Clinical Trials Partnership. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The host ESCRT-machinery repairs damaged endolysosomal membranes. If damage persists, selective macroautophagy/autophagy clears the damaged compartment. Mycobacterium tuberculosis (Mtb) is an intracellular pathogen that damages the phagosomal membrane and targets ESCRT-mediated repair as part of its virulence program. The E3 ubiquitin ligases PRKN and SMURF1 promote autophagic capture of damaged, Mtb-containing phagosomes. Because ubiquitination is a reversible process, we anticipated that host deubiquitinases (DUBs) would also be involved. Here, we screened all predicted mouse DUBs for their role in ubiquitin targeting and control of intracellular Mtb. We show that USP8 (ubiquitin specific peptidase 8) colocalizes with intracellular Mtb, recognizes phagosomal membrane damage, and is required for ESCRT-dependent membrane repair. Furthermore, we show that USP8 regulates the NFE2L2/NRF2-dependent antioxidant signature. Taken together, our study demonstrates a central role of USP8 in promoting Mtb intracellular growth by promoting phagosomal membrane repair, limiting ubiquitin-driven selective autophagy, and reducing oxidative stress. Abbreviation: BMDMs: bone marrow-derived macrophages; CFUs: colony-forming units; DUB: deubiquitinase; ESCRT: endosomal sorting complexes required for transport; LLOMe: L-leucyl-L-leucine methyl ester; MFI: mean fluorescence intensity; MOI: multiplicity of infection; Mtb: Mycobacterium tuberculosis ; NFE2L2/NRF2: nuclear factor, erythroid derived 2, like 2; PMA: phorbol 12-myristate 13-acetate; ROS: reactive oxygen species; USP8: ubiquitin specific peptidase 8.

Autophagy, metabolism, and associated signaling pathways play critical roles in bacterial survival within mammalian cells and influence the immunopathogenesis of infections. Mesenchymal stem cells (MSCs) are important host cells during Mycobacterium tuberculosis (Mtb) infection, yet how autophagy, metabolism, and related pathways are modulated in MSCs infected with the virulent H37Rv or the attenuated H37Ra strain of Mtb remains poorly understood. In this study, we utilized RNA-Seq screening, qRT-PCR, and Western Blotting to investigate the differences in these processes between H37Rv and H37Ra infections. Our results show that, at early time points (no more than 24h), infection with H37Rv significantly increased the expression of TlLR2, Prkaa2, and Prkaa2 phosphorylation in MSCs compared with H37Ra infection. Further analysis revealed that H37Rv infection induced a stronger autophagic response (evidenced by increased Atg9b and LC3II/LC3I) through the TLR2-AMP-AMPK pathway than H37Ra infection. Despite these differences in autophagy, there was no statistically significant difference in bacillary loads, suggesting that, in addition to autophagy, other factors such as apoptosis and immune-inflammatory responses may also regulate Mtb growth in MSCs. Additionally, the metabolic analysis showed that H37Rv infection led to increased expression of SLC2A3, PFKFB3, HK1, and ABCA1 in MSCs compared to H37Ra infection. These findings confirm that, during the early stages of infection, H37Rv induces enhanced autophagy, glucose metabolism, and cholesterol efflux through a more active TLR2-AMP-AMPK pathway than H37Ra. Therefore, MSCs may represent a novel target for the prevention and treatment of tuberculosis., Competing Interests: Declaration of competing interest We declare that we have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Improved vaccination strategies for tuberculosis are needed. Intravenous (i.v.) delivery of live attenuated Mycobacterium bovis BCG provides protection against Mycobacterium tuberculosis (Mtb) in macaques but poses safety challenges. Here we genetically engineered two strains, BCG-TetON-DL and BCG-TetOFF-DL, to either induce or inhibit expression of two phage lysin operons, respectively, upon tetracycline exposure. We show that lysin expression kills BCG in vitro, in infected macrophages, and following infection of immunocompetent (C57BL/6) and immunocompromised (SCID) mice. Modified BCG elicited similar immune responses and provided similar protection against Mtb challenge as wild-type BCG in mice. In macaques, cessation of tetracycline treatment reduced i.v.-administered BCG-TetOFF-DL numbers. Intravenous BCG-TetOFF-DL increased pulmonary CD4 T-cell responses compared with wild-type BCG-induced responses and provided robust protection against Mtb challenge. Sterilizing immunity occurred in 6 of 8 macaques compared with 2 of 8 wild-type BCG-immunized macaques. Thus, a 'kill-switch' BCG strain provides additional safety and robust protection against Mtb infection., Competing Interests: Competing interests: S.E. and D.S. are inventors on a pending patent application (U.S. Serial No. 62/959,485) related to the use of the kill-switch BCG strains described in this paper. The other authors declare no competing interests., (© 2025. The Author(s), under exclusive licence to Springer Nature Limited.)

Natural products have been pivotal in treating mycobacterial infections with early antibiotics such as streptomycin, forming the foundation of tuberculosis therapy. However, the emergence of multidrug-resistant and extensively drug-resistant Mycobacterium species has intensified the need for novel antimycobacterial agents. In this review, we revisit the historical contributions of natural products to antimycobacterial drug discovery and highlight recent advances in the field. We assess the application of molecular networking and the exploration of unculturable bacteria in identifying new antimycobacterial compounds such as amycobactin and levesquamides. We also highlight the role of semisynthesis in optimizing natural products, exemplified by sequanamycins and spectinomycin analogs that evade M. tuberculosis' intrinsic resistance. Finally, we discuss emerging technologies that are promising to accelerate the discovery and development of next-generation antimycobacterial therapies. Despite ongoing challenges, these innovative approaches offer renewed hope in addressing the growing crisis of drug-resistant mycobacterial infections., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Autophagy is a crucial immune defense mechanism that controls the survival and pathogenesis of M. tb by maintaining cell physiology during stress and pathogen attack. The E3-Ub ligases (PRKN, SMURF1, and NEDD4) and autophagy receptors (SQSTM1, TAX1BP1, CALCOCO2, OPTN, and NBR1) play key roles in this process. Galectins (LGALSs), which bind to sugars and are involved in identifying damaged cell membranes caused by intracellular pathogens such as M. tb , are essential. These include LGALS3, LGALS8, and LGALS9, which respond to endomembrane damage and regulate endomembrane damage caused by toxic chemicals, protein aggregates, and intracellular pathogens, including M. tb . They also activate selective autophagy and de novo endolysosome biogenesis. LGALS3, LGALS9, and LGALS8 interact with various components to activate autophagy and repair damage, while CGAS-STING1 plays a critical role in providing immunity against M. tb by activating selective autophagy and producing type I IFNs with antimycobacterial functions. STING1 activates cGAMP-dependent autophagy which provides immunity against various pathogens. Additionally, cytoplasmic surveillance pathways activated by ds-DNA, such as inflammasomes mediated by NLRP3 and AIM2 complexes, control M. tb . Modulation of E3-Ub ligases with small regulatory molecules of LGALSs and TRIM proteins could be a novel host-based therapeutic approach for controlling TB.

Background: Despite causing high mortality worldwide, paediatric tuberculosis is often undiagnosed. We aimed to investigate optimal testing strategies for microbiological confirmation of tuberculosis in children younger than 15 years, including the yield in high-risk subgroups (eg, children younger than 5 years, with HIV, or with severe acute malnutrition [SAM])., Methods: For this secondary analysis, we used data from RaPaed-TB, a multicentre diagnostic accuracy study evaluating novel diagnostic assays and testing approaches for tuberculosis in children recruited from five health-care centres in Malawi, Mozambique, South Africa, Tanzania, and India conducted between Jan 21, 2019, and June 30, 2021. Children were included if they were younger than 15 years and had signs or symptoms of pulmonary or extrapulmonary tuberculosis; they were excluded if they weighed less than 2 kg, had received three or more doses of anti-tuberculosis medication at time of enrolment, were in a condition deemed critical by the local investigator, or if they did not have at least one valid microbiological result. We collected tuberculosis-reference specimens via spontaneous sputum, induced sputum, gastric aspirate, and nasopharyngeal aspirates. Microbiological tests were Xpert MTB/RIF Ultra (hereafter referred to as Ultra), liquid culture, and Löwenstein-Jensen solid culture, which were followed by confirmatory testing for positive cultures. The main outcome of this secondary analysis was categorising children as having confirmed tuberculosis if culture or Ultra positive on any sample, unconfirmed tuberculosis if clinically diagnosed, and unlikely tuberculosis if neither of these applied., Findings: Of 5313 children screened, 975 were enrolled, of whom 965 (99%) had at least one valid microbiological result. 444 (46%) of 965 had unlikely tuberculosis, 282 (29%) had unconfirmed tuberculosis, and 239 (25%) had confirmed tuberculosis. Median age was 5·0 years (IQR 1·8-9·0); 467 (48%) of 965 children were female and 498 (52%) were male. 155 (16%) of 965 children had HIV and 110 (11%) children had SAM. 196 (82%) of 239 children with microbiological detection tested positive on Ultra. 110 (46%) of 239 were confirmed by both Ultra and culture, 86 (36%) by Ultra alone, and 43 (18%) by culture alone. 'Trace' was the most common semiquantitative result (93 [40%] of 234). 481 (50%) of 965 children had only one specimen type collected, 99 (21%) of whom had M tuberculosis detected. 484 (50%) of 965 children had multiple specimens collected, 141 (29%) of whom were positive on at least one specimen type. Of the 102 children younger than 5 years with M tuberculosis detected, 80 (78%) tested positive on sputum. 64 (80%) of 80 children who tested positive on sputum were positive on sputum alone; 61 (95%) of 64 were positive on induced sputum, two (3%) of 64 were positive on spontaneous sputum, and one (2%) was positive on both., Interpretation: High rates of microbiological confirmation of tuberculosis in children can be achieved via parallel sampling and concurrent testing procedures. Sample types and choice of test to be used sequentially should be considered when applying to groups such as children younger than 5 years, living with HIV, or with SAM., Funding: European and Developing Countries Clinical Trials Partnership programme, supported by the EU, the UK Medical Research Council, Swedish International Development Cooperation Agency, Bundesministerium für Bildung und Forschung, the German Center for Infection Research, and Beckman Coulter., Competing Interests: Declaration of interests All authors received grant funding for this work from the second European and Developing Countries Clinical Trials Partnership programme (supported by the EU), the German Center for Infection Research, and Beckman Coulter, paid to their institutions. Cepheid provided Xpert MTB/RIF Ultra testing kits and GeneXpert platforms at no cost to the consortium. HJZ is funded by the South African Medical Research Council. ZF-S receives funding from the South African Medical Research Council. LO was financially supported by a European Society For Paediatric Infectious Diseases fellowship award and a clinical leave stipend from the German Center for Infection Research. ELC has received funding from the Wellcome Trust Senior Fellowship. TDM has received funding, paid to his institution, from the Global Alliance Against Tuberculosis, the European and Developing Countries Clinical Trials Partnership programme, Médecins Sans Frontières, the EU Innovative Medicines Initiative, and Great Ormond Street Hospital charity intramural COVID-19 rapid response; receives personal payment for being Editor in Chief from Annals of Clinical Microbiology and Antimicrobials; and is part of the Chair Acid Fast Club UK. RSo has received funding from the US National Institutes of Health. All other authors declare no competing interests., (Copyright © 2025 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Background: The BACTEC Mycobacteria Growth Indicator Tube (MGIT) machine is the standard globally for detecting viable mycobacteria in patients' sputum. Samples are observed for no longer than 42 days, at which point the sample is declared 'negative' for tuberculosis (TB). This time to detection of bacterial growth, referred to as time-to-positivity (TTP), is increasingly of interest, not solely as a diagnostic tool but also as a continuous biomarker wherein change in TTP can be used for comparing the bactericidal activity of different TB treatments. However, as a continuous measure, there are oddities in the distribution of TTP values observed, particularly at higher values., Methods: We explored whether there is evidence to suggest setting an upper limit of quantification for modeling purposes (ULOQ M ) lower than the diagnostic limit of detection (LOD) using data from several TB-PACTS randomized clinical trials and PanACEA MAMS-TB., Results: Across all trials, less than 7.1% of weekly samples returned TTP measurements between 25 and 42 days. Further, the relative absolute prediction error (%) was highest in this range. When modelling with ULOQ M s of 25 and 30 days, estimator precision improved for 23 of 25 regimen-level slopes compared to models using the LOD. Discrimination between regimens based on Bayesian posteriors also improved., Conclusions: Although TTP measurements between 25 days and the diagnostic LOD may be important for diagnostic purposes, TTP values in this range may not contribute meaningfully to its use as a quantitative measure, particularly when assessing treatment response, and may lead to underpowered clinical trials., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Mycobacterium bovis and Mycobacterium tuberculosis are the most relevant among pathogenic mycobacteria, both belonging to the M. tuberculosis complex (MTC). Samples of blood, liver, spleen, kidneys, lungs and caseous tubercles were collected from a free-ranging juvenile black capuchin monkey (Sapajus nigritus) showing non-specific signs of illness. Macroscopic findings included emaciation, a caseous lesion in a tooth and gingiva, disseminated nodules in both lungs and left kidney parenchyma and caseous nodules on the pleura and mesentery. The lesions suggested MTC infection, a diagnosis subsequently supported in the lung by bacilloscopy, immunochromatography and PCR. A multiplex PCR further validated the presence of M. bovis genes. Cases of tuberculosis in platyrrhine primates have only been reported in animals maintained in captivity. We describe for the first time the pathological and molecular findings of M. bovis infection in a free-ranging platyrrhine monkey within an area of intense human-wildlife interaction, which has important implications from a One Health perspective., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)