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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.)

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.

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: 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.)

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.)

Under stress, bacteria display selective growth propensities on solid and liquid media. For Mycobacterium tuberculosis (Mtb), differentially culturable bacteria have been found in tuberculosis (TB) patient sputa. We hypothesized that antibiotic treatment can induce selective culturability in Mtb. We investigated the effect of exposure to TB drugs on Mtb culturability using clinical samples from an ongoing TB patient cohort and by conducting several in vitro experiments with a diverse set of Mtb strains. In patients, serial sputa were more likely to generate Mtb-positive cultures in liquid as opposed to solid medium, with this liquid culture bias extending up to 5 months post diagnosis. Experimentally, there was a disparity between bacterial time to positivity (TTP) and colony forming units (CFUs) when Mtb was exposed to isoniazid (INH) and rifampicin (RIF) alone or in combination. Cultures recovered from RIF treatment yielded more CFUs on agar plates, but INH-treated cultures had a faster TTP in liquid. Follow up experiments using a fluorescently labelled laboratory strain of Mtb revealed that CFUs of INH-treated bacteria were too low, thus over-estimating the killing effect of the treatment. Here we provide evidence that drug exposure affects culturability on solid medium in diverse Mtb strains, which has implications for treatment monitoring and drug-pathogen interaction studies., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Background: Uveitis presumed to be secondary to Mycobacterium tuberculosis is a rare but potentially blinding condition. Difficulty in making an accurate diagnosis and the low incidence of TB uveitis (TBU) contribute to the lack of evidence regarding the best management of this condition. This systematic review aims to analyse existing research to provide a summary of the literature regarding the utility of TB therapy for the management of TBU., Methods: This systematic review was prospectively registered on PROSPERO (PROSPERO 2021 CRD42021273379). We searched Medline, Embase and Central databases, and the search was done on 20th June 2023 with an updated literature search., Results: We included 55 studies and found that the heterogeneity in the methodology of these studies precluded metanalysis, and a narrative analysis was undertaken. Risk of bias analysis was undertaken using the Newcastle-Ottawa scale., Conclusions: Key findings of this systematic review include multiple systemic biases in the available evidence, and general lack of control for confounding variables. This results in many unanswered questions regarding the utility of TB therapy for TBU and reinforces the need for more data in this area., Competing Interests: Declarations. Ethics approval and consent to participate: This research did not require ethical approval. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Background: Diabetes mellitus (DM) is a major risk factor for tuberculosis (TB), However, limited research exists on their clinical and strain characteristics. This study aims to investigate the correlation between these factors in TB-DM patients in Changping District.  METHODS: Whole genome sequencing (WGS) and drug susceptibility tests (DST) were performed on culture-positive strains. Spearman correlation analysis was used to examine risk factors and the correlation between lineage, cavities, and hemoptysis in the TB-DM population. The specificity, sensitivity, and confidence intervals for predicting phenotypic drug resistance based on genotypic resistance were calculated., Results: Among the 3924 TB patients, 292 had DM, showing a doubling in the proportion of TB patients with DM over seven years. Among the 144 etiologically positive TB-DM cases treated at the Changping Institute for Tuberculosis Prevention and Treatment, 75% (108/144) of the patients exhibited tuberculosis lesions that formed cavities and 12.5% (18/144) with hemoptysis. A statistically significant difference in cavity formation across different age groups was observed (r = -0.198, P < 0.05). Out of the 144 etiologically positive patients, WGS successfully revived 73 MTB strains, with Lineage 2 being predominant. No statistical difference was found between lineages and the presence of cavities or hemoptysis. The DST results showed the highest resistance rates to isoniazid and streptomycin, both at 8.2% (6/73), with approximately one-quarter of the strains resistant to at least one anti-TB drug, and about half (47.1%, 8/17) resistant to first-line drugs. The study demonstrated good specificity but suboptimal sensitivity in predicting phenotypic drug resistance based on genotypic resistance., Conclusions: The rising incidence of diabetes in tuberculosis patients within Changping District has intensified the spread of TB, with these patients demonstrating severe illness and high drug resistance. This study aims to develop targeted prevention and management strategies, offering crucial guidance for treating co-infections of TB and DM and controlling disease spread., Competing Interests: Declarations. Ethics approval and consent to participate: In this study, all patients volunteered to participate with informed consent and the collected strains and patient information were kept strictly confidential, the research methods and protocols for the current study were carried out in accordance with relevant guidelines and regulations (declarations of Helsinki), which has been approved by The Ethics Committee of Beijing Changping Institute for Tuberculosis Prevention and Treatment (CJ-2023–001). Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2025. The Author(s).)

Background: Tuberculosis (TB) remains a serious global public health problem. The Mycobacterium tuberculosis (MTB) is responsible for approximately 10 million new TB cases globally each year. This study aimed to investigate transmission pattern and drug resistance of MTB in Shenzhen, China., Methods: A retrospective study on 286 samples from 184 TB patients collected between 2015 and 2018 in Shenzhen Third People's Hospital was conducted using whole-genome sequencing. Drug susceptibility testing (DST) was performed using both phenotypic DST (pDST) and molecular DST (mDST). Sample diversity was evaluated by SNPs and transmission clusters were identified based on SNP differences of 12 or fewer in genetic clusters., Results: Except four samples identified as non-tuberculous mycobacteria, 282 MTB samples (181 patients) underwent mDST, with 244 samples (162 patients) undergoing pDST. The overall multidrug-resistant rate in patients was 22.31% in pDST (12.00% for new patients and 40.82% for retreatment patients) and 34.48% in mDST (20.41% for new patients and 58.21% for retreatment patients). Totally 92 transmission clusters were identified, encompassing 70.21% samples (57.46% patients), with 5 clusters containing samples (15, 5.32%) from different patients (9, 4.97%), indicating recent transmission. The drug-resistant mutations in 36 of 45 transmission clusters (80.00%) were identical in all samples, suggesting the transmission of drug resistance. Patients with multiple samples were categorized into simultaneous sampling (SS) and continuous sampling (CS) groups, revealing significant differences in treatment types, treatment outcomes, residential addresses, and drug resistance types. mDST showed greater accuracy than pDST in SS and CS groups. A novel method based on heterozygous SNPs and two-sample Kolmogorov-Smirnov test were developed and identified 12 (4.26%) samples as mixed infection samples. Six of 12 patients had mixed and pure samples together, and major strains of mixed samples were closer to corresponding pure strains than minor strains., Conclusions: This retrospective study, conducted at the only municipal hospital specializing in infectious diseases in Shenzhen, provides the opportunity to understand drug resistance of TB patients, which mainly are refractory patients. The study revealed transmission patterns of MTB, analyzed mixed infections, and tracked changes in MTB strains during short/long-term treatment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2025 Ding, Liu, Wu, Zhang, Chen, Lai, Qu, Lin, Lu and Qu.)

The development of granulomas with central necrosis harboring Mycobacterium tuberculosis (Mtb) is the hallmark of human tuberculosis (TB). New anti-TB therapies need to effectively penetrate the cellular and necrotic compartments of these lesions and reach sufficient concentrations to eliminate Mtb. BTZ-043 is a novel antibiotic showing good bactericidal activity in humans in a phase IIa trial. Here, we report on lesional BTZ-043 concentrations severalfold above the minimal-inhibitory-concentration and the substantial local efficacy of BTZ-043 in interleukin-13-overexpressing mice, which mimic human TB pathology of granuloma necrosis. High-resolution MALDI imaging further reveals that BTZ-043 diffuses and accumulates in the cellular compartment, and fully penetrates the necrotic center. This is the first study that visualizes an efficient penetration and accumulation of a clinical-stage TB drug in human-like centrally necrotizing granulomas and that also determines its lesional activity. Our results most likely predict a substantial bactericidal effect of BTZ-043 at these hard-to-reach sites in TB patients., Competing Interests: Competing interests: NH has received a grant for conducting a study on delpazolid, a TB drug candidate, from LegoChem biosciences, to his institution. ELN received research funding from TB Alliance, Gates Medical Research Institute, and Janssen to his institution. MH, NH, and JD are employees of LMU Klinikum the institution that is developing BTZ-043 and have received funding from the German government. All other authors declare no competing interests., (© 2025. The Author(s).)

FadD32, a fatty acyl-AMP ligase, plays an indispensable role in mycobacterial mycolic acid synthesis and is a validated target for tuberculosis (TB) drug development. The crystal structure of Mycobacterium tuberculosis (Mtb)FadD32 has laid the foundation of structure-based drug discovery against this crucial enzyme. Here, we screened the "isoxazole" scaffold containing molecules against MtbFadD32 and identified a compound 2,4-dibromo-6-[3-(trifluoromethyl)-1,2-oxazol-5-yl]phenol (M1) with specific inhibitory activity against Mtb. Kinetics experiments showed that M1 inhibits MtbFadD32 and MtbFadD28 activity. The transcriptomics response of Mtb disclosed M1-mediated regulation of mycobacterial decisive genes involved in cell wall synthesis, consequently creating unfavorable conditions for Mtb survival. Further, M1 curtails the Mtb survival in infected macrophages and reduces Mtb burden and tubercular granulomas in a chronic infection model of BALB/c mice. Our findings provide an effective chemical scaffold to inhibit MtbFadD32 with the potential to inhibit multiple MtbFadD family of enzymes for further development as a promising candidate for treating TB.

The spread of drug-resistant strains of tuberculosis has hampered efforts to control the disease worldwide. The Mycobacterium tuberculosis cell wall envelope is dynamic, with complex features that protect it from the host immunological response. As a result, the bacterial cell wall components represent a potential target for drug discovery. Protein-protein interaction networks (PPIN) are critical for understanding disease conditions and identifying precise therapeutic targets. We used a rational theoretical approach by constructing a PPIN with the proteins involved in cell wall biosynthesis. The PPIN was constructed through the STRING database and embB was identified as a key protein by using four topological measures, betweenness, closeness, degree, and eigenvector, in the CytoNCA tool in Cytoscape. The 'Drug repurposing' approach was employed to find suitable inhibitors against embB. We used the Schrödinger suites for molecular docking, molecular dynamics simulation, and binding free energy calculations to validate the binding of protein with the ligand. FDA-approved drugs from the ZINC database and DrugBank were screened against embB (PDB ID: 7BVF) using high-throughput virtual screening, standard precision, and extra precision docking. The drugs were screened based on the XP docking score of the standard drug ethambutol. Accordingly, from the top five hits, azilsartan and dihydroergotamine were selected based on the binding free energy values and were further subjected to Molecular Dynamics Simulation studies for 100 ns. Our study confirms that Azilsartan and Dihydroergotamine form stable complexes with embB and can be used as potential lead molecules based on further in vitro and in vivo experimental validation.Communicated by Ramaswamy H. Sarma.

Tuberculosis continues to pose a health challenge causing the loss of millions of lives despite the existence of multiple drugs, for treatment. The emergence of drug-resistant strains has made the situation more complex making it increasingly difficult to fight against this disease. This review outlines the challenges associated with TB drug discovery, the nature of Mycobacterium tuberculosis shedding light on the mechanisms that lead to treatment failure and antibiotic resistance. We explore promising drug targets, encompassing inhibition of mycolyarabinogalactan peptidoglycan (MAGP) assembly, mycolic acid biosynthesis, DNA replication, transcription, translation, protein synthesis, and bioenergetics/metabolism pathways. A comprehensive overview of the global pipeline of anti-tuberculosis drugs at various stages of clinical trials, the diverse strategies being pursued to tackle this complex disease. By gaining an understanding of the mechanisms that contribute to resistance development and identifying suitable targets, we can pave the way for more effective treatments and contribute to global efforts to combat drug-resistant tuberculosis., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Nitin Pal Kalia reports financial support was provided by India Ministry of Science & Technology Department of Biotechnology. 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. Published by Elsevier Ltd.)

Background: Tuberculosis is an infectious disease that has become endemic worldwide. The causative bacteria Mycobacterium tuberculosis (Mtb) is targeted via several exciting drug targets. One newly discovered target is the Fatty Acyl-CoA synthase, which plays a significant role in activating the long-chain fatty acids., Research Design & Methods: This study aims to generate novel compounds using Machine Learning (ML) algorithms to inhibit this synthase. Experimentally derived bioactive compounds were chosen from ChEMBL and used as inputs for effective molecule generation by Reinvent4. The library of new molecules generated was subjected to a two-tiered molecular docking protocol, and the results were further studied to obtain a binding free energy check., Results: The ML-based de novo drug design (DNDD) approach successfully generated a diverse library of novel molecules targeting Fatty Acyl-CoA synthase. After rigorous molecular docking and binding free energy analysis, four new compounds were identified as potential lead candidates with promising inhibitory effects on Mtb lipid metabolism., Conclusions: The study demonstrated the effectiveness of a machine-learning approach in generating novel drug candidates against Mtb. The identified hit compounds show potential as inhibitors of Fatty Acyl-CoA synthase, offering a new avenue for developing treatments for tuberculosis, particularly in combating drug-resistant strains.

Objectives: This study aimed to examine the efficacy of whole genome sequencing (WGS) in accurately predicting susceptibility profiles, potentially eliminating the need for conventional phenotypic drug susceptibility testing (pDST) for first-line antituberculosis drugs in routine tuberculosis diagnosis., Methods: Over the period of 2017 to 2020, 1114 Mycobacterium tuberculosis complex isolates were collected with drug susceptibility testing conducted using the MGIT960 system and WGS performed for predicting drug resistance profiles. In addition, we implemented a new algorithm with an updated WGS workflow, omitting pan-susceptible strains from pDST., Results: Results showed that out of 1075 analysed isolates, WGS-based genotypic sensitivity predictions for isoniazid, rifampicin, ethambutol, and pyrazinamide were 100% (95% CI, 99.6-100%), 100% (95% CI, 99.62-100%), 99.8% (95% CI, 99.26-99.94%), and 100% (95% CI, 99.63-100%), respectively. In contrast, the WGS-based genotypic resistance prediction, was 98.85% (95% CI, 93.77-99.79%) for isoniazid, 94.74% (95% CI, 82.71-98.54%) for rifampicin, 86.96% (95% CI, 67.87-95.46%) for ethambutol, and 75.7% (95% CI, 59.9-86.63%) for pyrazinamide. Moreover, WGS enabled the implementation of a new testing algorithm that made it unnecessary to perform pDST in 954 of all 1075 samples (88.7%) and in 890 of 901 pan-susceptible samples (98.8%)., Discussion: Integrating WGS into tuberculosis management offers significant potential to replace phenotypic drug susceptibility testing, especially for problematic drugs like pyrazinamide and ethambutol, potentially improving treatment outcomes., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Mycobacterium tuberculosis (M. tb), the etiological agent of tuberculosis (TB), continues to be a major contributor to global mortality rates. To effectively combat this pandemic, TB control has to be enhanced in several areas, including point-of-care diagnostics, shorter and safer drug regimens, and preventative vaccination. The latest findings have highlighted autophagy as a host-defense mechanism that eradicates many invading bacteria, including M. tb. Thus, novel approaches like the stimulation of autophagy using various pharmaceutical drugs can be undertaken to deal with this noxious pathogen. The present study has been formulated to evaluate the anti-mycobacterial potential of Furamidine, a DNA minor groove binder (MGB). Initially, a non-cytotoxic concentration of Furamidine (10 µM) was used to assess its impact on the intracellular persistence of mycobacteria in differentiated THP-1 (dTHP-1) cells. Furamidine treatment compromised intracellular mycobacterial growth compared to control cells. Autophagy, a well-known host-defensive strategy, was investigated as a possible contributor to revealing the mechanism of action. Multiparametric approaches such as LC3-I to II conversion, protein level expression of different autophagic markers, and MDC staining were employed to study autophagic response that conclusively suggested the autophagy induction potential of Furamidine in dTHP-1 cells. Further, elevated LC3-II expression and increased autophagic vacuole accumulation under Baf-A1 treatment demonstrated the positive regulation of autophagic flux upon Furamidine treatment. Mechanistic investigations showed increased intracellular calcium (Ca 2+ ) level expression, SIRT1, pAMPK, and FOXO3a activation upon its treatment. Inhibition of Ca 2+ level expression suppressed Ca 2+ -mediated-FOXO3a level in Furamidine-treated cells. Furthermore, administering various inhibitors hampered the Furamidine-induced autophagy that impacted intracellular mycobacteria clearance. These results conclude that Furamidine triggered the Ca 2+ /pAMPK/SIRT1/FOXO3a pathway, causing less mycobacterial load in dTHP-1 cells., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests concerning the content of this article. The authors declare that none of their known financial or interpersonal conflicts seemed to have influenced the research presented in this paper., (Copyright © 2024 Elsevier GmbH. All rights reserved.)

Mycobacterium tuberculosis is a human pathogen that causes Tuberculosis (TB) disease. Researchers have reported the activity of traditional medicinal plants against human pathogens. However, antimycobacterial studies of medicinal plants against M. tuberculosis remain limited. Thus, the purpose of this study is to characterize the phytochemical profile, antibacterial and antimycobacterial activity of Tridax procumbens towards H37Rv. The antibacterial activity was elucidated by the inhibitory zone formed around the disc by performing disk diffusion method. Tridax antimycobacterial activity measured by Microplate Alamar Blue Assay (MABA) infers the sample is sensitive to H37Rv at MIC (minimum inhibitory concentration) 600 µg/mL. BACTEC MGIT 960 DST identifies the sample is susceptible to H37Rv and Rifampicin resistant (RR). Antiproliferative, functional group determination and mechanism of action of secondary metabolites were performed by MTT, Fourier transform infrared spectroscopy (FTIR) and Gas Chromatography-mass spectrometry (GC-MS). The phytocompounds antimycobacterial efficacy is further supported by molecular docking data. The binding interactions of ligands with gyrA gene revealed (S,Z)-Heptadeca-1,9-dien-4,6-diyn-3-ol molecule as a prominent phytocompound with a binding affinity of -6.6 kcal/mol., Competing Interests: Declarations. Conflict of interests: The authors declare that there is no conflict of interest regarding the publication of this article., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Pyrazinamide (PZA) is an important first-line drug for tuberculosis (TB) treatment by eradicating the persisting Mycobacterium tuberculosis complex (MTBC). Due to cost and technical challenges, end TB strategies are hampered by the lack of a simple and reliable culture-based PZA antimicrobial susceptibility testing (AST) for routine use. We initially developed a simplified chromogenic pyrazinamidase (PZase) test in the TB reference laboratory using a training set MTBC isolates with various drug-resistant profiles, and validated its performance using consecutive BACTEC MGIT 960 (MGIT)-culture-positive culture in 10 clinical laboratories. The pncA gene Sanger sequencing results were used as the reference, and compared to the MGIT-PZA AST. Differential diagnosis of Mycobacterium bovis was conducted using patented in-house real-time PCR. Of the 106 training isolates, the PZase test and MGIT-PZA AST showed 100.0% and 99.1% concordance as compared to Sanger sequencing, respectively. We found 32.1% (34/106) isolates harbored pncA mutations, including one isolate with silent mutation S65S. For validation, 1,793 clinical isolates were tested including 150 duplicate isolates from specimens of the same cases and 16 isolates with uncharacterized drug resistance (UDR)-associated mutations. Excluding duplicated and UDR isolates, we identified 2.6% (43/1,627) PZA-resistant isolates, including 1.3% (21/1,627) M . bovis isolates. The kappa values were 0.851-1.000. In addition, the accuracy of the PZase test conducted by 10 laboratories was 98.5%-100.0%. Our simplified PZase test demonstrated high concordance with Sanger sequencing and MGIT-PZA AST. Integrating the PZase test into routine first-line AST is effortless and represents an improvement in laboratory services for ending TB., Importance: We developed and validated a simple pyrazinamidase (PZase) test for pyrazinamide (PZA) antimicrobial susceptibility testing (AST). Our results demonstrated that the PZase test had high agreement with the pncA gene sequencing and MGIT-PZA AST. Integrating PZase test into routine AST is effortless and represents an improvement in laboratory services for ending TB., Competing Interests: The authors declare no conflict of interest.

Tuberculosis (TB), caused by Mycobacterium tuberculosis ( Mtb ), remains a major global health challenge, primarily due to the increasing prevalence of drug resistance. Consequently, the development of drugs with novel modes of action (MOAs) is urgently required. In this study, we discovered and characterized two potent inhibitors, Pi-1-58 and Pi-2-26, targeting the prokaryotic ubiquitin-like protein (Pup) ligase PafA of Mtb . Pi-1-58 was identified through computer-aided drug screening. The binding mode of Pi-1-58 and PafA was investigated through molecular docking, followed by experimental validations. Based on the core structure of Pi-1-58, we developed a more potent inhibitor, Pi-2-26, through structure-guided drug design. Both Pi-1-58 and Pi-2-26 exhibited selective and specific inhibition of PafA according to biochemical and cell-based assays. Importantly, the inhibitors demonstrated significant inhibition on Mtb survival in the presence of nitric oxide, mimicking the in vivo nitrogen limited environment that Mtb encountered in macrophage. Our findings provide a comprehensive understanding of the structural and functional aspects of these PafA inhibitors and establish a solid foundation for the development of novel therapeutics against tuberculosis.

Although para -aminosalicylic acid (PAS) has been used to treat tuberculosis for decades, mechanisms of resistance to this drug in Mycobacterium tuberculosis ( M. tuberculosis ) clinical isolates have not been thoroughly investigated. Previously, we found that decreased methylenetetrahydrofolate reductase (MTHFR) activity of Rv2172c led to increased sensitivity to antifolates in M. tuberculosis . In this study, we collected the genome-sequencing data of 173 PAS-resistant and 803 PAS-sensitive clinical isolates and analyzed rv2172c mutations in those 976 isolates. The results showed that two mutations (T120P and M172V) on rv2172c could be identified in a certain proportion (6.36%) of PAS-resistant isolates. The results of AlphaFold2 prediction indicated that the T120P or M172V mutation might affect the enzymatic activity of Rv2172c by influencing nicotinamide adenine dinucleotide (NADH) binding, and this was verified by subsequent biochemical analysis, demonstrating the role of residues Thr120 and Met172 on NADH binding and enzymatic activity of Rv2172c. In addition, the effect of rv2172c T120P or M172V mutation on methionine production and PAS resistance was determined in M. tuberculosis . The results showed that both T120P and M172V mutations caused increased intracellular methionine concentrations and high level PAS resistance. In summary, we discovered new molecular markers and also a novel mechanism of PAS resistance in M. tuberculosis clinical isolates and broadened the understanding of the NADH-dependent MTHFR catalytic mechanism of Rv2172c in M. tuberculosis , which will facilitate the molecular diagnosis of PAS resistance and also the development of new drugs targeting Rv2172c.

We tested the hypothesis that because of the different metabolic states of Mycobacterium tuberculosis (Mtb) in lesions, drugs in combination therapy often act effectively as monotherapy, leading to therapy failure and resistance emergence.Bactericidal and sterilizing activity studies were performed in the hollow fiber system of TB (HFS-TB) using the human equivalent dose of isoniazid (INH) 300 mg/day, rifampin (RIF) 600 mg/day, and pyrazinamide (PZA) 1.5 g/day either as monotherapy, two-, and three-drug combination for 28 days. The Mtb population (log 10 CFU/ml) for each drug, either monotherapy or combination, was compared using an analysis of variance.In the bactericidal activity studies, the microbial kill was driven by INH, followed by RIF, and PZA monotherapy failed. During the sterilizing activity, INH and RIF displayed similar microbial kill. The INH + RIF and RIF + PZA combinations were significantly different from each other but not from the INH + RIF + PZA combination. RIF and INH-resistant subpopulations did not increase despite premixing the inoculum with isogenic-resistant strains.Effective monotherapy arising from the selectivity of antibiotics against special Mtb sub-populations may not be the primary mechanism of resistance emergence. Different metabolic populations of Mtb were killed by more than one drug and were not under monotherapy when combination therapy was administered..

The surge of drug-resistant Mycobacterium tuberculosis (DR-TB) impedes the World Health Organization's efforts in ending TB and calls for new therapeutic formulations. M. tuberculosis sirtuin-like protein Rv1151c is a bifunctional enzyme with both deacetylation and desuccinylation activities, which plays an important role in M. tuberculosis drug resistance and stress responses. Thus, it appears to be a promising target for the development of new TB therapeutics. In this study, we screened 31,057 ligand compounds from seven compound libraries in silico to identify inhibitors of Rv1151c. Ipragliflozin can bind to Rv1151c and interact stably. Ipragliflozin can change the acylation level of M. tuberculosis by inhibiting Rv1151c and effectively inhibit the growth of M. tuberculosis H37Rv and M. smegmatis. It can potentiate the first-front anti-TB drug isoniazid. As an antidiabetic drug, Ipragliflozin can be potentially included in the regimen to treat diabetes-tuberculosis comorbidity., Competing Interests: Declaration of competing interest The authors have no relevant financial or non-financial interests to disclose., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Mycobacterium tuberculosis (MTB/Mtb) is the causative agent of tuberculosis (TB), a highly infectious serious airborne illness. TB usually affects the lungs, in 25 % of patients (children or immune impaired adults), mycobacteria can enter the blood stream and infect other bodily areas such the meninges, pleura, lymphatic system, genitourinary system, bones, and joints. Currently, the most challenging aspect of treating this illness is the ineffectiveness of the most potent first-line anti-TB medications, isoniazid, rifampin, pyrazinamide, and ethambutol, which can result in multidrug-resistant TB (MDR-TB), extensively drug-resistant TB (XDR-TB), and in rare instances, completely drug-resistant TB (TDR-TB). As a result, finding new pharmaceutical compounds to treat these diseases is a significant challenge for the scientific community. A number of bio-active molecules have been investigated in this quest, including quinoline, which is considered a promising candidate for the development of TB drugs. It is known that quinoline are low in toxicity and have a wide range of pharmacological properties. Researchers have investigated quinoline scaffolds as anti-TB drugs based on their biological spectrum. The objective of this review is to examine the recent development of quinoline and its structural characteristics crucial to its antitubercular (anti-TB) activity. A molecular analog of the TB treatment can be designed and identified with this information. As a result, future generation quinoline-based anti-TB agents with greater potency and safety can also be explored., 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.)

Introduction: Tuberculosis (TB) remains a major global health issue, causing around 10 million new cases and 1.3 million deaths in 2022. The challenge is compounded by multidrug-resistant (MDR) and extensively drug-resistant (XDR) TB strains, and co-infection with HIV., Areas Covered: The present review examines significant patent literature on TB chemotherapeutics from September 2021 to the present using the following databases, reaxys, google patent and espacenet. Only patents reporting compounds with a minimum inhibitory concentration (MIC) on whole Mycobacterium tuberculosis cells of ≤5 µM were selected for review., Expert Opinion: The fight against TB is advancing with the development of promising new compounds due to the challenge of drug-resistant strains. Notable among those reviewed in this paper are the benzothiazinones, showing high efficacy against both drug-sensitive and resistant TB strains. Additionally, Q203 analogues, demonstrate strong antitubercular activity, good microsomal stability, and favorable safety profiles. Finally, LysRS inhibitors also show significant promise in vivo models. These advancements underscore the importance of novel targets and innovative strategies in developing effective, resistance-resistant TB treatments.

In this study, we developed a modified NRA (MONRA) to determine the first and second-line drug susceptibilities of 5 reference ATCC strains and 42 clinical M. tuberculosis isolates. Unlike conventional NRA, which is often performed in solid media or 7H9 broth, the MONRA is performed in a different medium, AYC.2.1 broth, using lyophilized antibiotic tubes to determine drug susceptibility. The MONRA results were compared with BACTEC MGIT 960 method as the reference method for first-line drugs and conventional NRA performed in 7H9 broth for second-line drugs. The agreement between the MONRA and the reference method was determined as 97.62, 100, 97.62, and 100 % for streptomycin, isoniazid, rifampicin, and ethambutol, respectively. When the results were compared with convantional NRA, the agreement was determined as 100 % for all second-line antibiotics including levofloxacin, ofloxacin, and kanamycin. MONRA has the potential to eliminate challenges in implementing drug susceptibility testing in resource-limited settings., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Ahmet yilmaz coban reports financial support was provided by Akdeniz University. Ahmet Yilmaz Coban reports a relationship with Akdeniz University that includes: employment. 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 Elsevier Inc. All rights reserved.)

Tuberculosis (TB) caused by Mycobacterium tuberculosis has been a threat to human health for thousands of years and still leads to millions of deaths each year. TB is a disease that is refractory to treatment, partially due to its capacity for in-host persistence. The cell wall of mycobacteria, rich in mycolic acid, is broadly associated with bacterial persistence together with antimicrobial and immunological resistance. Enzymes for the biosynthesis of bacterial peptidoglycan, an essential component of the cell wall, have been addressed and considered as appealing drug targets in pathogens. Significant effort has been dedicated to finding inhibitors that hinder peptidoglycan biosynthesis, many with demonstrated enzymatic inhibition in vitro being published. One family of critical biosynthetic enzymes are the Mur enzymes, with many enzyme specific inhibitors having been reported. However, a lesser developed strategy which may have positive clinical implications is to take advantage of the common structural and catalytic characteristics among Mur enzymes and to allow simultaneous, multiple Mur inhibition, and avert the development of drug resistance. M. tuberculosis relies on these essential Mur enzymes, with the best-known subset being Mur ligases, but also utilizes unique functions of atypical transpeptidases resulting in peptidoglycan peptide cross-linking beneficial to the bacteria's capacity for chronic persistence in humans. A systematic review is now needed, with an emphasis on M. tuberculosis . The urgent development of novel anti-TB agents to counter rapidly developing drug resistance requires a revisit of the literature, past successes and failures, in an attempt to reveal liabilities in critical cellular functions and drive innovation., Competing Interests: The authors declare there are no competing interests., (©2024 Chiang and West.)

Current tuberculosis (TB) treatment is typically effective against drug-susceptible Mycobacterium tuberculosis, but can fail due to acquired drug resistance or phenotypic resistance. M. tuberculosis persisters, a subpopulation of viable but non-replicating (VBNR) antibiotic-tolerant bacteria, are thought to contribute to poor TB treatment outcomes. In this exploratory study, we investigated treatment-naïve drug-susceptible clinical isolates collected from people with TB, who subsequently had unsuccessful treatment outcomes. These were compared to isolates from cured individuals in terms of their ability to form VBNR subpopulations. Clinical isolates from individuals with unfavorable treatment outcomes form larger subpopulations of VBNR M. tuberculosis (2.67-13.71%) than clinical isolates from cured cases (0- 1.63%) following infection of THP-1 macrophages. All isolates were drug susceptible based on phenotypic and genotypic analysis. Whole genome sequencing identified 23 non-synonymous genomic variants shared by treatment failure clinical isolates, that were not present in isolates from cured cases. This exploratory study highlights the ability of treatment-naïve clinical isolates to form heterogeneous populations containing VBNR M. tuberculosis. We also demonstrate that clinical isolates from individuals with unsuccessful treatment outcomes form higher percentages of VBNR M. tuberculosis. The findings of this exploratory study suggest that an increased propensity to form VBNR subpopulations may impact TB treatment outcome., Competing Interests: Declarations Competing interests The authors declare no competing interests. Ethical approval Ethical approval was obtained from the Stellenbosch Human Research Ethics Committee (Registration Number N10/01/013) and the Biological and Environmental Safety committee (Registration Number BES-2023-13049)., (© 2024. The Author(s).)

Isoniazid is an important first-line medicine to treat tuberculosis (TB). Isoniazid resistance increases the risk of poor treatment outcomes and development of multidrug resistance, and is driven primarily by mutations involving katG , encoding the prodrug-activating enzyme, rather than its validated target, InhA. The chemical tractability of InhA has fostered efforts to discover direct inhibitors of InhA (DIIs). In this study, we bridge the gap in understanding the potential contribution of DIIs to novel combination regimens and demonstrate a clear distinction of DIIs, like GSK693 and the newly described GSK138, from isoniazid, based on activity against clinical isolates and contribution to novel drug regimens. The results suggest that DIIs, specifically GSK138 and GSK693, could be promising partners in novel drug regimens, including those used against isoniazid-resistant TB, potentially enhancing their efficacy and/or preventing the selection of resistant mutants and supporting the continued exploration of InhA as a promising target for TB drug development., Competing Interests: L.E., J.R.-T., A.G.-P., R.G.D.R., and D.B.-A. are employees of, and shareholders in, GSK. A.M.-L. holds GSK stock and has patents issued during his employment at GSK. J.D.M. and V.S. are employees of GSK. The other authors declare no conflict of interest.

The spectinamides are novel, narrow-spectrum semisynthetic analogs of spectinomycin, modified to avoid intrinsic efflux by Mycobacterium tuberculosis . Spectinamides, including lead MBX-4888A (Lee-1810), exhibit promising therapeutic profiles in mice, as single drugs and as partner agents with other anti-tuberculosis antibiotics including rifampin and/or pyrazinamide. Here, we show that MBX-4888A, given by injection with the front-line standard of care regimen, is treatment shortening in multiple murine tuberculosis infection models. The positive treatment responses to MBX-4888A combination therapy in multiple mouse models, including mice exhibiting advanced pulmonary disease, can be attributed to favorable distribution in tissues and lesions, retention in caseum, along with favorable effects with rifampin and pyrazinamide under conditions achieved in necrotic lesions. This study also provides an additional data point regarding the safety and tolerability of spectinamide MBX-4888A in long-term murine efficacy studies., Competing Interests: The authors declare no conflict of interest.

Tuberculosis (TB) is a world health challenge the treatment of which is impacted by the rise of drug-resistant strains. Thus, there is an urgent need for new antitubercular compounds and novel approaches to improve current TB therapy. The zebrafish animal model has become increasingly relevant as an experimental system. It has proven particularly useful during early development for aiding TB drug discovery, supporting both the discovery of new insights into mycobacterial pathogenesis and the evaluation of therapeutical toxicity and efficacy in vivo. In this review, we summarize the past two decades of zebrafish-Mycobacterium marinum research and discuss its contribution to the field of bioactive antituberculosis therapy development., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Tuberculosis (T.B.) remains a prominent global cause of health challenges and death, exacerbated by drug-resistant strains such as multidrug-resistant tuberculosis MDR-TB and extensively drug-resistant tuberculosis XDR-TB. For an effective disease management strategy, it is crucial to understand the dynamics of T.B. infection and the impacts of treatment. In the present article, we employ AI-based machine learning techniques to investigate the immunity impact of medications. SEIPR epidemiological model is incorporated with MDR-TB for compartments susceptible to disease, exposed to risk, infected ones, preventive or resistant to initial treatment, and recovered or healed population. These masses' natural trends, effects, and interactions are formulated and described in the present study. Computations and stability analysis are conducted upon endemic and disease-free equilibria in the present model for their global scenario. Both numerical and AI-based nonlinear autoregressive exogenous NARX analyses are presented with incorporating immediate treatment and delay in treatment. This study shows that the active patients and MDR-TB, both strains, exist because of the absence of permanent immunity to T.B. Furthermore, patients who have recovered from tuberculosis may become susceptible again by losing their immunity and contributing to transmission again. This article aims to identify patterns and predictors of treatment success. The findings from this research can contribute to developing more effective tuberculosis interventions., Competing Interests: Declaration of competing interest I have no conflict of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Zoonotic human tuberculosis (TB) caused by Mycobacterium bovis (M. bovis) is as vital as Mycobacterium tuberculosis, however with scarce available information. We aimed to use whole-genome sequencing (WGS) technology to take a deep insight into the circulating genotypes of human M. bovis and the genomic characteristics underlying virulence and drug resistance., Methods: The study included smear positive Ziehl-Neelsen samples from patients with suspected tuberculosis. Samples were cultured on Lowenstein-Jensen media and suspected colonies of M. bovis were selected to undergo DNA extraction and WGS. Data was analysed using the Bacterial and Viral Bioinformatics Resource Center (BV-BRC), and online bioinformatics tools. A phylogenetic tree was constructed for our sequenced strains, in addition to a set of 59 previously sequenced M. bovis genomes from different hosts and countries., Results: Out of total 112 mycobacterial positive cultures, five M. bovis were isolated and underwent WGS. All sequenced strains belonged to Mycobacterium tuberculosis var bovis, spoligotype BOV&#95;1; BOV&#95;11. Resistance gene mutations were determined in 100% of strains to pyrazinamide (pncA and rpsA), isoniazid (KatG and ahpC), ethambutol (embB, embC, embR and ubiA), streptomycin (rpsl) and fluoroquinolones (gyrA and gyrB). Rifampin (rpoB and rpoC) and delamanid (fbiC) resistance genes were found in 80% of strains. The major represented virulence classes were the secretion system, cell surface components and regulation system. The phylogenetic analysis revealed close genetic relatedness of three sequenced M. bovis strains to previous reported cow strains from Egypt and human strains from France, as well as relatedness of one M. bovis strain to four human Algerian strains. One sequenced strain was related to one cow strain from Egypt and a human strain from South Africa., Conclusions: All sequenced M. bovis isolates showed the same spoligotype, but diverse phylogeny. Resistance gene mutations were detected for anti-TB drugs including pyrazinamide, isoniazid, streptomycin, ethambutol, fluoroquinolones, cycloserine, rifampin and delamanid. The virulence profile comprised genes assigned mainly to secretion system, cell surface components and regulation system. Phylogenetic analysis revealed genetic relatedness between our isolates and previously sequenced bovine strains from Egypt as well as human strains from other nearby countries in the region., (© 2024. The Author(s).)

The rise of drug-resistant Mycobacterium tuberculosis (Mtb) has extended the duration of tuberculosis (TB) treatment and reduced the likelihood of cure. One strategy to combat this issue is the development of inhibitors targeting the virulence factors of bacterial pathogens. Mtb' catalase (KatG) is crucial for its detoxification mechanisms and also serves as a significant virulence factor for the bacterium. In this study, twelve derivatives synthesized from 5-fluoropyridine and benzo[b]thiophene demonstrated antimycobacterial efficacy with minimum inhibitory concentrations (MICs) varying between 0.5 and 32 μg/mL. Compound 2, 1-(benzo[b]thiophen-2-ylmethylene) thiosemicarbazide, emerged as the most potent candidate. It effectively inhibited Mtb KatG, enhanced the production of reactive oxygen species (ROS) in Mtb, and achieved Mtb killing within 96 hours at a concentration of 2 μg/mL (4×MIC). Molecular docking simulations revealed that compound 2 binds tightly to the active site of Mtb-KatG with a docking score of 114, indicating that it may serve as a potent inhibitor of Mtb-KatG. The rabbit skin tuberculosis model was employed to assess the virulence of Mtb. Animal study results indicated that the granulomas induced by Mtb after treatment with compound 2 were reduced in size, exhibited a lower bacterial load, and the bacteria were no longer aggregated, in contrast to those caused by untreated Mtb. Hence, compound 2 can be regarded as a molecule capable of neutralizing the virulence factors of Mtb. This research offers insights into the design of anti-Mtb molecules with novel mechanisms of action., (© 2024 Wiley-VHCA AG, Zurich, Switzerland.)

Host heterogeneity in pulmonary tuberculosis leads to varied responses to infection and drug treatment. The present portfolio of anti-TB drugs needs to be boosted with new drugs and drug regimens. Macozinone, a clinical-stage molecule targeting the essential enzyme, DprE1, represents an attractive option. Mice (I/St, B6, (AKRxI/St)F1, B6.I-100 and B6.I-139) genetically diverse susceptibility to Mycobacterium tuberculosis (Mtb) H37Rv infection were subjected to aerosol- or intravenous infection to determine the efficacy of macozinone (MCZ). They were treated with macozinone or reference drugs (isoniazid, rifampicin). Lung and spleen bacterial burdens were measured at four and eight weeks post-infection. Lung histology was evaluated at four weeks of treatment. Treatment with macozinone resulted in a statistically significant reduction in the bacterial load in the lungs and spleen as early as four weeks after treatment initiation in mice susceptible or resistant to Mtb infection. In the TB hypoxic granuloma model, macozinone was more potent than rifampicin in reducing the CFU counts. However, histopathological analysis revealed significant lung changes in I/St mice after eight weeks of treatment initiation. Macozinone demonstrated efficacy to varying degrees across all mouse models of Mtb infection used. These results should facilitate its further development and potential introduction into clinical practice., Competing Interests: Declaration of competing interest V.M. is an inventor on macozinone patents., (Copyright © 2024 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

Tuberculosis (TB) remains one of the leading infectious disease killers in the world. The ongoing development of novel anti-TB medications has yielded potent compounds that often target single sites with well-defined mechanisms of action. However, despite the identification of resistance-associated mutations through target deconvolution studies, comparing these findings with the diverse Mycobacterium tuberculosis populations observed in clinical settings is often challenging. To address this gap, we constructed an open-access database encompassing genetic variations from > 50,000 clinical isolates, spanning the entirety of the M. tuberculosis protein-encoding genome. This resource offers a valuable tool for investigating the prevalence of target-based resistance mutations in any drug target within clinical contexts. To demonstrate the practical application of this dataset in drug discovery, we focused on drug targets currently undergoing phase II clinical trials. By juxtaposing genetic variations of these targets with resistance mutations derived from laboratory-adapted strains, we identified multiple positions across three targets harbouring resistance-associated mutations already present in clinical isolates. Furthermore, our analysis revealed a discernible correlation between genetic diversity within each protein and their predicted essentiality. This meta-analysis, openly accessible via a dedicated dashboard, enables comprehensive exploration of genetic diversity pertaining to any drug target or resistance determinant in M. tuberculosis., (© 2024. The Author(s).)

Despite known treatments, tuberculosis (TB) remains the world's top infectious killer, highlighting the pressing need for new drug regimens. To prioritize the most efficacious drugs for clinical testing, we previously developed a PK-PD translational platform with bacterial dynamics that reliably predicted short-term monotherapy outcomes in Phase IIa trials from preclinical mouse studies. In this study, we extended our platform to include PK-PD models that account for drug-drug interactions in combination regimens and bacterial regrowth in our bacterial dynamics model to predict cure at the end of treatment and relapse 6 months post-treatment. The Phase III STAND trial testing a new regimen comprised of pretomanid (Pa), moxifloxacin (M), and pyrazinamide (Z) (PaMZ) was suspended after a separate ongoing trial (NC-005) suggested that adding bedaquiline (B) to the PaMZ regimen would improve efficacy. To forecast if the addition of B would, indeed, benefit the PaMZ regimen, we applied an extended translational platform to both regimens. We predicted currently available short- and long-term clinical data well for drug combinations related to BPaMZ. We predicted the addition of B to PaMZ to shorten treatment duration by 2 months and to have similar bacteriological success to standard HRZE treatment (considering only treatment success but not withdrawal from side effects and other adverse events), both at the end of treatment for treatment efficacy and 6 months after treatment has ended in relapse prevention. Using BPaMZ as a case study, we have demonstrated our translational platform can predict Phase II and III outcomes prior to actual trials, allowing us to better prioritize the regimens most likely to succeed., Competing Interests: The authors declare no conflict of interest.

Background: Extrapulmonary tuberculosis (EP-TB) constitutes one-fifth of all tuberculosis (TB) cases. EP-TB mimics common infections which pose diagnostic dilemma, requires extensive diagnostics that culminate into therapeutic delay often resulting in irrational and empirical institution of antitubercular therapy (ATT) in challenging cases. This supplemented by poor treatment compliance resulted in emergence of Drug-resistant (DR) strains of EP-TB which further impedes the path to recovery. The aim of the present series is to study the rare and diverse presentations of EP-TB caused by drug-sensitive (DS) and DR mycobacterium that require a multi-modal diagnostic approach and appropriate treatment., Methods: This observational retrospective series incorporated six rare EP-TB cases, excluding those with solitary lung affection and underwent comprehensive diagnostic tests aimed at microbial isolation from affected tissues with subsequent drug resistance testing. They were treated by integrative approach, standard (first/second/third line) ATT while few required emergent surgical interventions. Patient outcomes were evaluated based on clinicoradiological improvement and microbiological clearance determined in follow-up., Results: Out of six cases (four males and two females; age range: 14-62 years), pleural linings, kidneys, brain and its lining, skin, and axial skeleton were directly affected, while superior mesenteric artery (SMA) syndrome was an indirect consequence of infection. Elective thoracic and urosurgical interventions supplemented medical management in two cases while urgent neurosurgical decompression improved outcomes in Pott's spine case that exhibited drug resistance. Notably of five DS EP-TB, one patient showed poor clinical response necessitating treatment escalation while nutritional rehabilitation was key in SMA syndrome., Conclusions: EP-TB requires high clinical suspicion and a multidisciplinary approach for diagnosis and treatment. Addressing treatment adherence, with emphasis on good nutrition to tackle cachexia, is necessary for favorable outcomes., (Copyright © 2024 International Journal of Mycobacteriology.)

Tuberculosis (TB), a deadly disease caused by Mycobacterium tuberculosis (Mtb) infection, remains one of the top killers among infectious diseases worldwide. How to increase targeting effects of current anti-TB chemotherapeutics and enhance anti-TB immunological responses remains a big challenge in TB and drug-resistant TB treatment. Here, mannose functionalized and polyetherimide protected graphene oxide system (GO-PEI-MAN) was designed for macrophage-targeted antibiotic (rifampicin) and autophagy inducer (carbamazepine) delivery to achieve more effective Mtb killings by combining targeted drug killing and host immunological clearance. GO-PEI-MAN system demonstrated selective uptake by in vitro macrophages and ex vivo macrophages from macaques. The endocytosed GO-PEI-MAN system would be transported into lysosomes, where the drug loaded Rif@Car@GO-PEI-MAN system would undergo accelerated drug release in acidic lysosomal conditions. Rif@Car@GO-PEI-MAN could significantly promote autophagy and apoptosis in Mtb infected macrophages, as well as induce anti-bacterial M1 polarization of Mtb infected macrophages to increase anti-bacterial IFN-γ and nitric oxide production. Collectively, Rif@Car@GO-PEI-MAN demonstrated effectively enhanced intracellular Mtb killing effects than rifampicin, carbamazepine or GO-PEI-MAN alone in Mtb infected macrophages, and could significantly reduce mycobacterial burdens in the lung of infected mice with alleviated pathology and inflammation without systemic toxicity. This macrophage targeted nanosystem synergizing increased drug killing efficiency and enhanced host immunological defense may be served as more effective therapeutics against TB and drug-resistant TB., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Disseminated tuberculosis (dTB) is usually rare in immunocompetent individuals. We report a case of dTB presented with constitutional symptoms and extensive lymphadenopathy with bilateral lung parenchymal airspace opacities. Histopathological and molecular methods confirmed the etiological agent - Mycobacterium tuberculosis. She was started on first-line antitubercular treatment and she had significant clinical recovery., (Copyright © 2024 International Journal of Mycobacteriology.)

Reporting any uncommon or untapped changes in bacterial genetics or physiology would be of great importance to support the drug development process. We studied 120 Mycobacterium tuberculosis clinical isolates with different geographical origin within India and their resistance profile and found a significant number of isolates (109) harboring the polymorphism at nucleotide positions 61 and 284 of the gyrA gene. Bioinformatics analysis of these changes for drug binding suggested no significant change in the binding of the drug but have lower binding energies as compared with the wild-type proteins. Although functionally silent for the gyrA gene, these changes are indicating a silent geographical and evolutionary change that needs to be further studied for drug discovery and bacterial fitness., (© 2024 Environmental Mutagenesis and Genomics Society.)

The emergence of multidrug-resistant tuberculosis (MDR-TB) strains has rendered many anti-TB drugs ineffective. Consequently, there is an urgent need to identify new drug targets against Mycobacterium tuberculosis (Mtb). Filament Forming Temperature Sensitive Gene Z (FtsZ), a member of the cytoskeletal protein family, plays a vital role in cell division by forming a cytokinetic ring at the cell's center and coordinating the division machinery. When FtsZ is depleted, cells are unable to divide and instead elongate into filamentous structures that eventually undergo lysis. Since the inactivation of FtsZ or alterations in its assembly impede the formation of the Z-ring and septum, FtsZ shows promise as a target for the development of anti-mycobacterial drugs. This review not only discusses the potential role of FtsZ as a promising pharmacological target for anti-tuberculosis therapies but also explores the structural and functional aspects of the mycobacterial protein FtsZ in cell division. Additionally, it reviews various inhibitors of Mtb FtsZ. By understanding the importance of FtsZ in cell division, researchers can explore strategies to disrupt its function, impeding the growth and proliferation of Mtb. Furthermore, the investigation of different inhibitors that target Mtb FtsZ expands the potential for developing effective treatments against tuberculosis., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2023. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

The emergence of antimicrobial resistance (AMR) across bacterial pathogens presents a serious threat to global health. This threat is further exacerbated in tuberculosis (TB), mainly due to a protracted treatment regimen involving a combination of drugs. A diversity of factors contributes to the emergence of drug resistance in TB, which is caused by the pathogen Mycobacterium tuberculosis (Mtb). While the traditional genetic mutation-driven drug resistance mechanisms operate in Mtb, there are also several additional unique features of drug resistance in this pathogen. Research in the past decade has enriched our understanding of such unconventional factors as efflux pumps, bacterial heterogeneity, metabolic states, and host microenvironment. Given that the discovery of new antibiotics is outpaced by the emergence of drug resistance patterns displayed by the pathogen, newer strategies for combating drug resistance are desperately needed. In the context of TB, such approaches include targeting the efflux capability of the pathogen, modulating the host environment to prevent bacterial drug tolerance, and activating the host anti-mycobacterial pathways. In this review, we discuss the traditional mechanisms of drug resistance in Mtb, newer understandings and the shaping of a set of unconventional approaches to target both the emergence and treatment of drug resistance in TB., (© 2024 Federation of European Biochemical Societies.)

Antibiotic tolerance in Mycobacterium tuberculosis reduces bacterial killing, worsens treatment outcomes, and contributes to resistance. We studied rifampicin tolerance in isolates with or without isoniazid resistance (IR). Using a minimum duration of killing assay, we measured rifampicin survival in isoniazid-susceptible (IS, n=119) and resistant (IR, n=84) isolates, correlating tolerance with bacterial growth, rifampicin minimum inhibitory concentrations (MICs), and isoniazid-resistant mutations. Longitudinal IR isolates were analyzed for changes in rifampicin tolerance and genetic variant emergence. The median time for rifampicin to reduce the bacterial population by 90% (MDK90) increased from 1.23 days (IS) and 1.31 days (IR) to 2.55 days (IS) and 1.98 days (IR) over 15-60 days of incubation, indicating fast and slow-growing tolerant sub-populations. A 6 log10-fold survival fraction classified tolerance as low, medium, or high, showing that IR is linked to increased tolerance and faster growth (OR = 2.68 for low vs. medium, OR = 4.42 for low vs. high, p-trend = 0.0003). High tolerance in IR isolates was associated with rifampicin treatment in patients and genetic microvariants. These findings suggest that IR tuberculosis should be assessed for high rifampicin tolerance to optimize treatment and prevent the development of multi-drug-resistant tuberculosis., Competing Interests: SV, NB, DV, LN, DT, NQ, LT, HN, VH, PT, DH, NL, MC, GT, BJ, NT No competing interests declared, (© 2024, Vijay, Bao et al.)

Due to interindividual variability in drug metabolism and pharmacokinetics, traditional isoniazid fixed-dose regimens may lead to suboptimal or toxic isoniazid concentrations in the plasma of patients with tuberculosis, contributing to adverse drug reactions, therapeutic failure, or the development of drug resistance. Achieving precision therapy for isoniazid requires a multifaceted approach that could integrate various clinical and genomic factors to tailor the isoniazid dose to individual patient characteristics. This includes leveraging molecular diagnostics to perform the comprehensive profiling of host pharmacogenomics to determine how it affects isoniazid metabolism, such as its metabolism by N-acetyltransferase 2 (NAT2), and studying drug-resistant mutations in the Mycobacterium tuberculosis genome for enabling targeted therapy selection. Several other molecular signatures identified from the host pharmacogenomics as well as other omics-based approaches such as gut microbiome, epigenomic, proteomic, metabolomic, and lipidomic approaches have provided mechanistic explanations for isoniazid pharmacokinetic variability and/or adverse drug reactions and thereby may facilitate precision therapy of isoniazid, though further validations in larger and diverse populations with tuberculosis are required for clinical applications. Therapeutic drug monitoring and population pharmacokinetic approaches allow for the adjustment of isoniazid dosages based on patient-specific pharmacokinetic profiles, optimizing drug exposure while minimizing toxicity and the risk of resistance. Current evidence has shown that with the integration of the host pharmacogenomics-particularly NAT2 and Mycobacterium tuberculosis genomics data along with isoniazid pharmacokinetic concentrations in the blood and patient factors such as anthropometric measurements, comorbidities, and type and timing of food administered-precision therapy approaches in isoniazid therapy can be tailored to the specific characteristics of both the host and the pathogen for improving tuberculosis treatment outcomes., (© 2024. The Author(s).)

Drug development for tuberculosis is hindered by the methodological limitations in the definitions of patient outcomes, particularly the slow organism growth and difficulty in obtaining suitable and representative samples throughout the treatment. We developed target product profiles for biomarker assays suitable for early-phase and late-phase clinical drug trials by consulting subject-matter experts on the desirable performance and operational characteristics of such assays for monitoring of tuberculosis treatment in drug trials. Minimal and optimal criteria were defined for scope, intended use, pricing, performance, and operational characteristics of the biomarkers. Early-stage trial assays should accurately quantify the number of viable bacilli, whereas late-stage trial assays should match the number, predict relapse-free cure, and replace culture conversion endpoints. The operational criteria reflect the infrastructure and resources available for drug trials. The effective tools should define the sterilising activity of the drug and lower the probability of treatment failure or relapse in people with tuberculosis. The target product profiles outlined in this Review should guide and de-risk the development of biomarker-based assays suitable for phase 2 and 3 clinical drug trials., Competing Interests: Declaration of interests SHG and WS developed the tuberculosis bacterial load assay and are collaborating with LifeArc to bring the assay to market. IK is participating in studies evaluating the feasibility of the TB22 signature developed by her research group as a treatment monitoring tool. JH holds patent shares for the TB22 signature. LJS is an employee of Johnson & Johnson and reports employee stock and stock options from Johnson & Johnson. NHe is an employee of the University Hospital (LMU) and reports grants to the institute from Beckman Coulter, European and Developing Countries Clinical Trials Partnership (EDCTP; EU Horizon 2020), and DZIF for research and evaluation of new tuberculosis diagnostics. NHi reports grants or contract and consulting fees from Otsuka Novel Product Muenich (ONPG) and having detailed information on the LAM sputum biomarker. RvC reports INtegrative omics and aspirin response to define the ThERapeutiC targEts for Pediatric TBM grant (R01AI165721) from the National Institutes of Health and participating in the Data Safety Monitoring Board for three randomised controlled trials focused on tuberculosis meningitis. SBG, MK, MR, and JBT are employees of FIND, the global alliance for diagnostics, a not-for-profit foundation that supports the evaluation of publicly prioritised tuberculosis assays and implementation of WHO-approved (guidance and prequalification) assays using donor grants. FIND has product evaluation agreements with several private-sector companies that design diagnostics for tuberculosis and other diseases. These agreements strictly define FIND’s independence and neutrality with regard to these private-sector companies. SBG and MR are also members of the New Diagnostics Working Group (NDWG) Stop TB Partnership. CL is a patent holder on biomarkers for monitoring of tuberculosis treatment. All other authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Mycobacterium tuberculosis (Mtb) replicates within host macrophages by adapting to the stressful and nutritionally constrained environments in these cells. Exploiting these adaptations for drug discovery has revealed that perturbing cAMP signaling can restrict Mtb growth in macrophages. Specifically, compounds that agonize or stimulate the bacterial enzyme, Rv1625c/Cya, induce cAMP synthesis and this interferes with the ability of Mtb to metabolize cholesterol. In murine tuberculosis (TB) infection models, Rv1625c/Cya agonists contribute to reducing relapse and shortening combination treatments, highlighting the therapeutic potential for this class of compounds. More recently, cAMP signaling has been implicated in regulating fatty acid utilization by Mtb. Thus, a new model is beginning to emerge in which cAMP regulates the utilization of host lipids by Mtb during infection, and this could provide new targets for TB drug development. Here, we summarize the current understanding of cAMP signaling in Mtb with a focus on our understanding of how cAMP signaling impacts Mtb physiology during infection. We also discuss additional cAMP-related drug targets in Mtb and other bacterial pathogens that may have therapeutic potential., Competing Interests: Declaration of interests B.C.V. is a listed co-inventor of a patent covering mCLB073/TBD11. D.K. has no interests to declare., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)