Enhanced antibacterial activity against multidrug-resistant Mycobacterium tuberculosis of nano‑silver prepared by Konjac glucomannan via asymmetric flow field-flow fractionation.

PROTACs as novel therapeutics against Mycobacterium tuberculosis: Current progress and future directions.

Dihydroquinazolin-4(3H)-imines have emerged as a promising scaffold for developing novel antimycobacterial agents. Here, we integrated QSAR modeling, in vitro screening, molecular dynamics (MD), binding free-energy calculations, and in vivo toxicity assessment to identify potent inhibitors against Mycobacterium tuberculosis (Mtb). Four QSAR models demonstrated strong internal and external reliability (R2>0.68; Q2LOO>0.62; R2extup to 0.82). Additional validation parameters met recommended thresholds (Q2-F1/F2/F3≥0.72; CCCext≥0.88; r2m aver≥0.72; r2m delta: 0.04), and Williams plots confirmed that all predictions fell within the applicability domain (h*: 0.286). Biological assays identified 11 active compounds, with MIC values ranging from 25 to 200μM. Compound 2c (p-methylphenyl derivative) and 3d (m, p-dichlorophenyl analog) were the most potent, displaying MIC values of 50 and 25μM, respectively. MD simulations revealed stable and specific interactions with Eis, an acetyltransferase linked to kanamycin resistance. Compound 2c exhibited a mean ΔGbind of -52.19±3.21kcal/mol, while 3d showed a more favorable ΔGbind of -73.15±3.16kcal/mol, consistent with its superior in vitro potency. Distinct interaction profiles-especially the engagement of Tyr126 and hydrophobic clusters-help explain their differential affinities. Moreover, both leads demonstrated low to moderate in vitro cytotoxicity against HepG2 cells at the concentrations evaluated. In vivo acute toxicity in Zophobas morio indicated LD₅₀ values of 500mg/kg for 2c and 100mg/kg for 3d, with transient tremors and melanization observed only for 3d. Since compound 2c exhibited a safer in vivo toxicity profile, this compound was investigated for its association with antimicrobial drugs. These compounds were validated as promising anti-TB candidates, supported by robust QSAR predictivity, favorable binding energetics, and measurable in vitro and in vivo toxicity profiles, reinforcing their potentials for further optimization.

Tuberculosis (TB) remains a major global public health burden, exacerbated by the continued emergence and spread of drug-resistant Mycobacterium tuberculosis. Despite the rapid expansion of publicly available whole-genome sequencing data, gaps remain in the consistent characterization of global population structure, dominant sequence-type (ST) distributions, accessory genome variability, and antimicrobial resistance (AMR) gene profiles, largely due to fragmented and uneven sampling across regions and time periods. This study aimed to conduct a large-scale in silico analysis of publicly available M. tuberculosis whole-genome sequences to descriptively characterize global ST structure, accessory genome diversity, AMR gene landscapes, and their geographic and temporal distributions, while integrating available phenotypic susceptibility data. We conducted the largest genomic analysis of M. tuberculosis to date, examining 7890 high-quality genomes from 82 countries (1900-2024) retrieved from NCBI GenBank. Rigorous quality filtering using CheckM ensured retention of genomes with >90% completeness and <5% contamination. Comprehensive genomic characterization included assembly metrics, annotated gene features, multi-locus sequence typing, AMR profiling using AMRFinderPlus (v4.0.23; database 2025-07-16.1), temporal trend analysis, geographic distribution mapping, and gene presence pattern (GPP) clustering to assess accessory genome diversity. Analysis of 7890 high-quality M. tuberculosis genomes from 77 countries (1900-2024) revealed a highly conserved global population dominated by a few epidemic clones. Although 158 ST were identified, three ST (ST 215, ST 279, ST 276) accounted for 84.9% of all isolates, with ST 215 alone representing 58.0%, indicating a strong global clonal bottleneck, while 90.5% of ST were rare (≤10 isolates each). Most isolates were human-derived (93.7%), and genome size (∼4.38Mb) and gene content (∼4149 genes) showed minimal variation worldwide. AMR analysis identified 27 AMR genes, but >99.6% of isolates carried only three core genes (erm(37), blaC, and aac(2')-Ic), whereas all other resistance genes occurred in <0.25% of genomes, including a single vancomycin-resistant isolate (0.01%). Phenotypic data showed high susceptibility to first-line drugs (97-98%), but substantial non-susceptibility to several second-line agents, particularly fluoroquinolones (ciprofloxacin and ofloxacin) and the second-line drugs capreomycin and ethionamide. Overall, while global M. tuberculosis is driven by a few dominant clones with a conserved core genome, rare lineages and resistance profiles highlight important hidden genomic diversity. GPP analysis identified 146 recurrent patterns, with GPP1 dominating ST 215, ST279, and ST276, suggesting limited accessory genome diversification. This large-scale in silico analysis reveals a highly skewed global sequence-type distribution of M. tuberculosis, with pronounced geographic structuring and widespread presence of conserved, intrinsic chromosomal resistance-associated genes. The findings emphasize the importance of cautious interpretation of resistance gene prevalence and phenotypic non-susceptibility patterns derived from heterogeneous public datasets, and highlight key methodological considerations for global genomic analyses of M. tuberculosis.

Digital twin of Mycobacterium tuberculosis infection: Integrating immune dynamics and pathogen adaptation for precision therapy.

Methyl jasmonate promotes host defense by activating autophagy during Mycobacterium tuberculosis infection.

Targeting Mycobacterium tuberculosis GAPDH elicits potent bactericidal responses by dysregulating enzyme activity, redox dynamics and iron acquisition.

Third-generation derivatives of the tuberculosis drug pretomanid featuring acetylene-bridged aryl-heteroaryl or heterotriaryl side chains were designed as novel agents for nanoparticle-based delivery, seeking better efficacy and safety. While all nitro compounds retained excellent activity against Mycobacterium tuberculosis, several examples in the latter class excelled as having potency superior to the original lead (7) but equivalent to or reduced lipophilicity, implying enhanced lipophilic efficiency. Initial studies suggested a similar mode of action against the related fish pathogen, M. marinum; therefore, fourteen candidates were further assessed as micellar formulations in M. marinum-infected zebrafish embryo assays. Overall, the best new analogue was 14 (the 6-amino-linked congener of 7), which was non-toxic and displayed efficacy comparable to 7 in both the blood and neural tube M. marinum infection models. Lead 14 also exhibited high microsomal stability, moderate cell permeability in an MDR1-MDCKII screen, and an enhanced pharmacokinetic profile in mice, with 93 % oral bioavailability.

During tuberculosis (TB), organ-specific immune responses and intracellular pathways play critical roles in disease progression and prognosis. Identifying genes that regulate these immune mechanisms remains a key challenge in improving TB management strategies. To investigate genes potentially associated with enhanced resistance to TB and the modulation of immune responses, we analysed RNA-seq data from whole cells isolated from the lungs and livers of mice infected with Mycobacterium tuberculosis (Mtb) at two time points that represent different outcomes. We hypothesised that these two organs mount distinct responses to infection, supported by differences in the immune response and bacterial burden kinetics observed in each tissue. Our analysis revealed differential gene expression profiles between the lungs and livers, primarily involving metabolic and immune-related pathways. Through meta-analysis, we identified orthologous genes shared between Mtb-infected mice and human patients with latent pulmonary TB. In the omics analysis, the four genes, Creb3l1, Myo7b, Cyyr1, and Cbs, were differentially expressed and associated with either resistance or susceptibility. Invitro assays further demonstrated that knockdown of CREB3L1 in Mtb-infected THP-1 or primary human monocytes impaired key effector functions, including phagocytosis, bacterial killing, and apoptosis. Taken together, these findings indicate that CREB3L1 possibly contributes to the regulation of genes essential for bacterial control in the lungs during latent TB infection. In contrast, its increased expression in the peripheral blood of patients with severe TB is more likely linked to systemic inflammatory dysregulation rather than direct antimicrobial activity. Notably, CREB3L1 expression in these patients positively correlated with cytokines such as IL-17, IL-12, and IFN-γ, which are central to macrophage activation and effector T cell recruitment. Thus, CREB3L1 appears to play a dual role in TB: under controlled infection, it acts as an immunomodulator limiting excessive pulmonary inflammation, while in severe disease, it may reflect an attempt by the host to amplify inflammatory responses to counteract progressive infection.

Tuberculosis (TB) is a bacterial infection caused by the bacilli, Mycobacterium Tuberculosis (MTB). While it primarily affects the lungs, various other organs like the lymph nodes, pleura, and peritoneum can be involved in Extrapulmonary Tuberculosis (EPTB). Involvement of the tonsils, however, is extremely rare, even in a TB-endemic country like India. Tonsillar TB may occur secondary to pulmonary infection through lymphatic or haematogenous dissemination, and its presentation often mimics malignancy (fever, loss of weight). This case report contains a case of a 56-year-old male patient who presented with progressive dysphagia, significant weight loss, low-grade fever, and cervical lymphadenopathy. Clinical examination revealed a unilateral tonsillar enlargement with firm neck nodes, initially raising a strong suspicion of malignancy. Routine blood investigations were normal, and a diagnostic tonsillectomy was performed, whose histopathological examination revealed granulomatous inflammation with caseous necrosis. The diagnosis was then confirmed by microbiological evidence of MTB. The novelty of this case report lies in the isolated tonsillar involvement without active pulmonary disease, its close clinical resemblance to malignancy, and the diagnostic challenge involved. Early recognition and accurate diagnosis are crucial for initiating appropriate therapy and preventing transmission.

A DNAzyme-CRISPR cascade strategy for preamplification-free detection of Mycobacterium tuberculosis.

Introduction: Bronchial anthracosis is a chronic respiratory condition characterised by the deposition of carbon particles in the bronchial mucosa. It is frequently associated with prolonged exposure to environmental pollutants, biomass smoke, and occupational dust, particularly in low-resource and rural settings. Aim: To elucidate the clinical and microbiological profile of patients with bronchial anthracosis in the mountainous valley of Kashmir, India. Materials and Methods: This cross-sectional study involved 88 patients diagnosed with bronchial anthracosis who were recruited from a tertiary care hospital. Data were collected on demographics, co-morbidities, exposure history, and microbiological findings through Bronchoalveolar Lavage (BAL). Statistical analysis was performed using percentage distribution and logistic regression analysis. Results: The mean age of the patients was 62.4±8.7 years, with females comprising 59.0% of the study population. Biomass fuel exposure (34.1%) and smoking (39.8%) were identified as significant risk factors. Common co-morbidities included Chronic Obstructive Pulmonary Disease (COPD) and hypertension. The chief complaints were cough (26.1%) and breathlessness (13.6%). BAL analysis revealed various pathogens, with Mycobacterium tuberculosis identified in 10.2% of cases. Logistic regression analysis demonstrated significant associations between bronchial anthracosis and age, smoking, and biomass fuel exposure, emphasising the influence of environmental risk factors. Conclusion: The findings highlight the significant role of environmental and occupational exposures—particularly biomass fuel use and smoking—in the development of bronchial anthracosis. Older adults, especially housewives and farmers, were the most affected groups, emphasising the need for targeted public health interventions. The association of bronchial anthracosis with respiratory infections and comorbidities such as COPD underscores the importance of early detection and appropriate management. Preventive strategies, including reduction of indoor air pollution and implementation of smoking cessation programmes, are essential to mitigate the disease burden in the ethnic population of Kashmir, India.

Distinct serum metabolic profiles with supportive diagnostic value in differentiating tuberculosis and Mycobacterium avium complex pulmonary disease.

Rapid one-tube sputum processing for tuberculosis diagnosis via azide-functionalized magnetic nanoplatforms with selective bacterial capture.

Hypoxia restricts the generation of BCG-responsive polycytotoxic T cells.

Bovine tuberculosis (bTB) is a neglected zoonotic disease of major public health and economic importance in Nigeria. This study determined the prevalence of Mycobacterium tuberculosis complex (MTBC) among slaughtered cattle at Yola Modern Abattoir, Adamawa State, using liquid and solid culture methods. A cross-sectional abattoir-based study was conducted in which 190 bovine lung samples with lesions suggestive of tuberculosis were collected. Samples were processed using Petroff’s decontamination method and cultured in the Mycobacteria Growth Indicator Tube (MGIT 960 BACTEC system) and on Lowenstein–Jensen (LJ) media supplemented with glycerol and pyruvate. MTBC-positive cultures were confirmed using the SD Bioline™ TB Ag MPT64 assay. Out of 190 samples examined, 85(44.7%), were posistive for MTBC by TB-MBLA, 73 (38.4%) were positive for MTBC by liquid culture, while 45 (23.7%) were positive by solid culture. TB-MBLA, showed the highest detection rate followed by Liquid culture and the least being solidculture method (χ² = 19.34; p = 0.000). Based on TB-MBLA results, the apparent prevalence of bovine tuberculosis was 44.7%. The high prevalence observed confirms the endemicity of bovine tuberculosis among cattle slaughtered in Yola and indicates a substantial increase compared with earlier reports from the area. The practice of selling carcasses after removal of affected organs poses a significant zoonotic risk to abattoir workers and consumers. Strengthening meat inspection, implementing active surveillance, and adopting effective national bTB control strategies are urgently needed to reduce transmission and protect public health. Keywords: Mycobacterium tuberculosis complex, bovine tuberculosis, Mycobacterium growth indication tube, BACTEC 960, Lowenstein-Jensen medium, TB-MBLA

ABSTRACT Objectives This study aims to evaluate the clinical application value of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) in the verification and identification of difficult-to-classify nontuberculous mycobacterial (NTM) strains. Methods From December 2024 to June 2025, 106 suspected NTM isolates were collected from 10 districts in Shenzhen, China. Initial identification was performed using HRM and REBA commercial kits, with targeted nanopore sequencing and mNGS as the composite reference standard. MALDI-TOF MS was used to verify strains unresolved by the kits, and its diagnostic performance was evaluated. Results The HRM kit demonstrated concordance with the reference standard in 98 of 106 samples (concordance rate: 89.1%), whereas the REBA kit concorded in 88 samples (concordance rate: 80.0%). The REBA kit exhibited a tendency toward misidentification of NTM species as Mycobacterium tuberculosis. When the reference results were used as a baseline with tNanopore providing parallel validation (achieving ≥95% concordance with reference results), MALDI-TOF MS demonstrated poor performance in identifying difficult-to-classify NTM strains. Specifically, MALDI-TOF MS showed poor concordance in detecting M. abscessus (Kappa = 0.244), while Mycobacterium intracellulare, Mycobacterium kansasii, and Mycobacterium gordonae demonstrated kappa values of 0.543, 0.477, and 0.483, respectively, indicating low concordance overall. Furthermore, 13 species exceeded the detection range of MALDI-TOF MS, resulting in false-positive identifications or detection failures, with Mycobacterium abscessus exhibiting the highest rate of misidentification. Conclusion The limitations of MALDI-TOF MS in verifying difficult-to-classify NTM strains have been demonstrated. The findings emphasize that PCR-based molecular detection combined with gene sequence analysis remains the most reliable methodological approach for accurately identifying challenging NTM species in clinical practice.

ABSTRACT Background Intraocular dual infection by Mycobacterium tuberculosis complex (MTBC) and non-tuberculous mycobacteria (NTM) is rarely reported. Objectives This study describes the clinical features, aetiology, and treatment outcomes of patients with dual infection from MTBC and NTM-related uveitis. Methods A retrospective study was conducted on 146 clinically suspected tubercular uveitis (TBU) patients who underwent diagnostic or therapeutic pars plana vitrectomy as plan of laboratory diagnosis-based management between June 2023 and December 2024. Undiluted vitreous samples were analysed with nested MTBC-NTM multiplex real-time PCR assay kit. Results Six out of 146 patients (4.11%) were found infected with dual infection from MTBC and NTM. All six cases showed severe clinical presentation characterised by bilateral panuveitis (5/6 patients), with significant structural sequelae. Optic atrophy was universal and occurred more frequently than in isolated TBU, while complicated cataract and hypotony-related maculopathy were also common. Despite the dual etiology, all patients achieved inflammatory resolution and visual recovery (mean BCVA improving from 20/400 to 20/100) following standard anti-tubercular therapy (ATT) and corticosteroids, with no recurrence during follow-up. Conclusions Concurrent MTBC and NTM infection drives a distinct, aggressive form of panuveitis characterized by optic nerve pallor, anterior segment complications of complicated cataract or uveitic membranes and maculopathy. Despite this severity, standard ATT remained an effective first-line strategy, likely either due to therapeutic cross-coverage against NTM or MTBC being the primary driver of uveitis in the present study. The concurrent positivity also highlights the need for vitreous molecular profiling and further research in co-infections in infectious posterior uveitis.

Background Tuberculosis, the second deadliest infectious disease in the world after COVID-19, is caused by Mycobacterium tuberculosis. This bacterium most commonly affects the lungs. Economic and financial barriers can hinder access to healthcare services for the diagnosis and management of this disease. The objective of this study was to determine the cost of managing drug-sensitive tuberculosis in the Selembao Health Zone, specifically at the Diagnostic and Treatment Center (CSDT) of Makala General Referral Hospital. Methods A descriptive cross-sectional study was conducted from February 24 to March 17, 2024, involving 122 tuberculosis patients who were diagnosed, treated, and declared cured at the tuberculosis screening and treatment center (CSDT) Makala, in the Selembao Health Zone. A simple random probability sampling method was used. Descriptive analyses were performed to determine the costs (direct, indirect, and total) associated with tuberculosis management. Results The direct cost of managing drug-sensitive tuberculosis per patient in our study area (Selembao) ranged from 139,150 to 787,150 CDF (50.6 to 286.2 USD), with a median of 214,150 CDF (77.9 USD), based on the exchange rate of 2,750 CDF for 1 USD during the study period. The indirect cost per patient was estimated at 6,000 to 7,860,000 CDF (2.2 to 2,858.2 USD), with a median of 796,000 CDF (289.5 USD). The total cost of care ranged from 172,150 to 8,040,000 CDF (62.6 to 2,923.6 USD), with a median of 1,026,650 CDF (373.3 USD). The share of the DRC government in supporting the management of this disease was estimated at 12.1% of the total cost. Conclusions The management of drug-sensitive tuberculosis remains a significant financial burden not only for patients but also for their families and households. We therefore recommend substantial government support to enhance the coverage of this condition.

Whole-genome sequencing of Mycobacterium tuberculosis can be a valuable tool for TB surveillance and treatment, providing insights into transmission patterns and comprehensive drug susceptibility testing. However, the slow growth of M. tuberculosis means traditional culture-based sequencing methods can take weeks to return results, which has limited the widespread adoption of these techniques and limited their use in clinical decision-making. Tiled amplicon sequencing is a fast, reliable, and cost-effective method of whole-genome sequencing that can be done directly on clinical specimens and has been implemented at scale in academic and public health laboratories across the world; it was the cornerstone of SARS-CoV-2 sequencing and has been adapted for a wide range of viral pathogens. However, similar methods are not yet available for far larger bacterial genomes. Extending this approach to M. tuberculosis would significantly reduce the cost, labor, and turnaround time for whole-genome sequencing. We designed a tiled amplicon panel consisting of 5,128 primers that covers the entire M. tuberculosis genome, the largest tiled amplicon sequencing panel we are aware of to date. Applying our amplicon panels to clinical samples of sputum, we show the ability to recover whole-genome bacterial sequences without the need for culture. The resulting sequence data can be used to determine M. tuberculosis lineage and reliably identify markers of drug resistance. Using this approach in clinical settings could reduce the time needed for comprehensive drug susceptibility testing from weeks to days and enable genomic epidemiology to be performed at scale, even in resource-limited settings.IMPORTANCEWe have developed and tested an amplicon panel, TB-seq, for the priority pathogen Mycobacterium tuberculosis, demonstrating recovery of near-full genomes directly from patient sputum, including mixed and low-concentration samples. This approach significantly reduces the turnaround time for this slow-growing bacterium while maintaining high accuracy in detecting clinically relevant mutations, including those associated with drug resistance. Given the global burden of tuberculosis and the critical need for faster diagnostic solutions, we believe our method has the potential to improve clinical decision-making and public health strategies.