Search Results (1,214)

Antimicrobial resistance in mycobacteria arises from a complex interplay of intrinsic and acquired mechanisms that collectively limit the efficacy of current therapeutic options. Intrinsic resistance is largely driven by the low permeability of the mycobacterial cell envelope, the activity of efflux pumps, and the presence of drug-modifying enzymes, which together restrict intracellular drug accumulation and contribute to broad baseline tolerance. This review integrates resistance mechanisms of both M. tuberculosis and M. abscessus, two clinically relevant mycobacteria that share core molecular pathways while exhibiting species-specific determinants that complicate treatment. Additional intrinsic factors, including biofilm formation and stress-induced adaptive responses, further enhance persistence and reduce susceptibility to multiple drug classes. Acquired resistance predominantly results from chromosomal mutations affecting drug targets or prodrug activation pathways, such as katG, inhA, rpoB, gyrA, and pncA in Mycobacterium tuberculosis, leading to high rates of multidrug-resistant and extensively drug-resistant disease. In nontuberculous mycobacteria, species-specific determinants—including inducible macrolide resistance mediated by erm(41) in M. abscessus, plasmid-mediated erm (55) variants, rrl and rrs mutations, diverse enzymatic inactivation systems, and regulatory alterations in the MarR family that result in inducible resistance to drugs such as ethionamide —generate highly variable resistance profiles that complicate treatment. Recent advances in molecular diagnostics, including PCR-based assays, whole-genome sequencing, CRISPR-based diagnostic platforms, AI-assisted diagnostics, and emerging multi-omics approaches, have improved the detection of resistance-associated mutations and enhanced understanding of mycobacterial pathophysiology. In parallel, new therapeutic agents and optimized regimens offer promising avenues to overcome resistance, although emerging resistance to novel drugs underscores the need for continued surveillance. This review synthesizes current knowledge on the molecular basis of resistance in M. tuberculosis and NTM, highlighting implications for diagnosis, treatment, and future research. Full article

Non-tuberculous mycobacteria (NTM) are increasingly recognized as opportunistic pathogens, yet accurate species identification remains challenging. In this study, two clinical isolates, 19-857^T and 20-525, obtained from patients with suspected pulmonary tuberculosis in Beijing, were characterized using a polyphasic approach integrating phenotypic, biochemical, chemotaxonomic, and genomic analyses. Phylogenetic analysis based on 16S rRNA gene sequences placed both strains within the genus Mycobacterium, with highest similarity to M. peregrinum DSM 43271^T and M. arcueilense DSM 46715^T. However, whole-genome comparisons via average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH) yielded values of 85.8–94.1% and 29.1–54.3%, respectively, against their closest relatives—well below the species delineation thresholds (95–96% for ANI; 70% for dDDH)—while the two isolates shared 98.8% ANI and 90.1% dDDH, confirming their co-assignment to a single novel species. Genomic analysis further revealed a repertoire of genes potentially associated with virulence, including glycopeptidolipid biosynthesis, mce operons, ESX secretion systems, and mycobactin synthesis, alongside antimicrobial resistance determinants against rifampicin, tetracyclines, and β-lactams. Mobile genetic element analysis also uncovered extensive insertion sequences and transposon-related elements with differential distribution between the two strains. Based on the combined evidence, we propose Mycobacterium shunyiense sp. nov., with strain 19-857^T (=CGMCC 38906^T) as the type strain. This discovery expands the understanding of NTM diversity and provides genomic reference data for accurate clinical identification. Full article

The development of dual-acting hybrid antibiotics is a promising strategy to combat the ongoing spread of drug-resistant tuberculosis. Inspired by the structure of the natural dithiolopyrrolone hybrid antibiotic thiomarinol and based on the synergistic effects confirmed by checkerboard assays, we employed a molecular hybridization strategy. The dithiolopyrrolone natural product thiolutin was covalently linked to the pharmacophores of four clinical antitubercular drugs—cycloserine, linezolid, isoniazid, and pyrazinamide—through alkyl linkers of 7–10 carbon atoms via amide condensation, leading to the design and synthesis of 15 novel hybrids. In vitro antitubercular activity evaluation revealed that the cycloserine series exhibited the best activity, with MIC values as low as 1 μg/mL, followed by the isoniazid series. Cytotoxicity assays showed that all cycloserine hybrids had IC₅₀ values > 40 μg/mL against RAW 264.7 mouse macrophages, markedly lower than that of thiolutin alone. Among them, T₁-CS and T₄-CS displayed the best selectivity indices, achieving an effective reduction in cytotoxicity. This study successfully constructed a class of thiolutin–cycloserine hybrids with low cytotoxicity and high selectivity, providing a valuable molecular template for the discovery of novel antitubercular lead compounds. Full article

Tuberculosis (TB) remains a major global health challenge, compounded by rising drug resistance. Traditional medicinal plants used for TB-related symptoms represent a valuable yet underexplored source of potential antimycobacterial agents. In this study, the phytochemical content and biological activities of Dombeya rotundifolia and Lippia javanica were evaluated, and antimycobacterial compounds were isolated through bioassay-guided fractionation. Plant leaves were collected, extracted with solvents of varying polarity, and screened for phenolics and flavonoids. Antioxidant activity was assessed using DPPH and ferric reducing power assays, antimycobacterial activity was tested against Mycobacterium smegmatis, anti-inflammatory activity was determined by the egg albumin denaturation method, and cytotoxicity was evaluated in THP-1 cells using the MTT assay. Extraction yields varied, with water extracts of L. javanica showing the highest yield and hexane extracts of D. rotundifolia the lowest. Both plants contained bioactive phytochemicals with measurable antioxidant activity, while acetone and dichloromethane extracts of D. rotundifolia displayed the strongest antimycobacterial activity with MIC values of 0.16 mg/mL. Cytotoxicity assays indicated moderate toxicity at higher extract concentrations. Bioassay-guided isolation led to the identification of two fatty acid fractions with tentative characterization, which showed notable antimycobacterial activity (MIC 0.25 mg/mL). These findings provide preliminary support for the ethnomedicinal use of D. rotundifolia in respiratory conditions traditionally associated with tuberculosis and suggest a potential contribution of its fatty acids to its observed antimycobacterial activity. Full article

Cutaneous co-infection with Mycobacterium tuberculosis (MTB) and Mycobacterium avium complex (MAC) is extremely rare and easily missed due to overlapping histopathological features. We report a previously healthy, HIV-negative middle-aged woman who presented with a progressive destructive mass in the left inguinal-perineal region. Imaging revealed sinus tract formation, osteolytic bone lesions, and chronic inflammation in the right middle lobe of the lung. Initial metagenomic next-generation sequencing (mNGS) detected 3756 reads of the Mycobacterium tuberculosis complex (MTBC) and 111 reads of Mycobacterium intracellulare (M. intracellulare); the latter was interpreted as possible colonization or contamination because of its low abundance. Empirical anti-tuberculosis therapy produced only transient partial improvement, followed by paradoxical worsening, local recurrence, and new bone destruction. After a high suspicion of mixed infection, a MAC-directed combination regimen (including azithromycin and a short course of amikacin) was added, leading to complete clinical cure; subsequent repeat cultures confirmed the presence of MAC. This is the first report of cutaneous MTB-MAC co-infection in the inguinal-perineal region of an adult without overt immune abnormalities, accompanied by disseminated bone lesions. This case highlights that in regions where nontuberculous mycobacteria (NTM) are co-endemic, atypical destructive skin lesions with paradoxical worsening despite initial response to anti-tuberculosis therapy should raise suspicion of MAC co-infection. The combination of mNGS and conventional culture facilitates identification of mixed infections and guides precision therapy, but mNGS results must be interpreted cautiously in the clinical context. Full article

Background/Objectives: Granulomatous inflammation in tissue biopsies presents a diagnostic challenge, particularly in differentiating between Mycobacterium tuberculosis complex (MTB) and non-tuberculous mycobacteria (NTM) in high-HIV-prevalence areas. This study evaluates the prevalence and species distribution of mycobacteria in formalin-fixed, paraffin-embedded (FFPE) tissues from patients with granulomatous inflammation and correlates these findings with demographic and clinical data. Methods: Conducted at the NHLS-Dr George Mukhari Tertiary Laboratory, the retrospective descriptive study analysed 37 FFPE samples (January 2019–December 2020) after obtaining ethical approval and collecting clinicopathological data. Histopathological assessments using H&E and Ziehl–Neelsen (ZN) stains were carried out, and mycobacterial species were identified utilising multiplex PCR with reverse hybridisation (GenoType MTBC/CM/AS assays). Results: The cohort had an average age of 38.59 years, with a predominance of females (59.5%) and a high HIV positivity rate (62.2%). The most common biopsy site was lymph nodes (32.4%). All samples exhibited necrotising granulomas, but only 8.1% displayed acid-fast bacilli (AFB) on ZN staining. Molecular testing revealed mycobacterial DNA in 75.7% of samples, identifying NTM alone in 50.0%, MTB alone in 10.7%, and both in 28.6%. M. fortuitum was the most frequently recognised species. Importantly, there was no significant association between mycobacterial detection and HIV status (p > 0.05). Conclusions: The findings demonstrate that molecular methods substantially improve mycobacterial detection rates and speciation. This underscores the diagnostic limitations of ZN staining and highlights the importance of molecular diagnostics for accurately identifying bacterial aetiology, crucial for appropriate antimicrobial therapy in immunocompromised patients. Full article

Rapid detection and differentiation of Mycobacterium tuberculosis complex (MTBC) and nontuberculous mycobacteria (NTM) are crucial for appropriate treatment selection. The NeoPlex™ TB/NTM Detection Kit is a PCR assay that simultaneously detects MTBC and NTM and identifies five clinically important NTM species (M. intracellulare, M. avium, M. kansasii, M. abscessus subsp. abscessus, and M. abscessus subsp. massiliense). We evaluated its comparative analytical performance using 200 stored nucleic acid extracts from respiratory and non-respiratory MTBC- or NTM-positive clinical specimens after excluding 27 specimens without interpretable species-level results. Results were compared with those of a line probe assay (LPA), which formed the reference test. Concordance with the LPA was 93.0%, including 96.9% for MTBC and 92.3% for NTM. The NeoPlex assay correctly differentiated all MTBC- and NTM-positive specimens included in the study cohort. For species-level identification, sensitivity and specificity were 98.9% and 100% for M. intracellulare, 97.6% and 100% for M. avium, 82.4% and 100% for M. kansasii, 79.0% and 100% for M. abscessus subsp. abscessus, and 100% and 100% for M. abscessus subsp. massiliense, respectively. These findings suggest that this multiplex PCR assay enables rapid, accurate detection and differentiation of MTBC and NTM species directly from clinical samples. Full article

Tuberculosis (TB) remains a leading cause of death from a single infectious agent, and its outcome is shaped not only by Mycobacterium tuberculosis (Mtb) itself, but also by the host’s metabolic state. This review synthesises current understanding of how Mtb reprograms macrophage immunometabolism and how this reprogramming propagates to a systemic level, culminating in skeletal muscle dysregulation and TB-associated cachexia. We describe the molecular mechanisms by which Mtb subverts phagosomal maturation, the glycolytic (Warburg-like) switch governed by HIF-1α and accumulation of immunomodulatory tricarboxylic acid cycle intermediates, and the M1/M2 polarisation balance that dictates bacterial containment versus persistence. We then trace the cytokine- and metabolite-mediated circuits (TNF-α, IL-6, IL-1β, lactate, ketone bodies, free fatty acids) that link infected macrophages to ubiquitin–proteasome and autophagy–lysosome-driven muscle proteolysis, mitochondrial dysfunction and oxidative stress. Building on these mechanisms, we propose an immunometabolic and muscle-derived biomarker framework that, although still requiring clinical validation, may offer value for diagnosis, host-response stratification and treatment monitoring, and we discuss host-directed therapeutic strategies that target macrophage metabolism and muscle preservation. By integrating immunity, metabolism and systemic pathology at the molecular level, this work highlights translational opportunities relevant to the host immunity, diagnosis and treatment of tuberculosis. Full article

Background: Opportunistic infections remain a significant cause of morbidity among people living with HIV (PLHIV) in sub-Saharan Africa despite expanded access to antiretroviral therapy (ART). This study evaluated the association between immune status, viral load suppression, ART adherence, and the risk of opportunistic infections among HIV-positive patients receiving ART in the rural Eastern Cape, South Africa. Methods: A retrospective cross-sectional study was conducted using clinical records of HIV-positive patients attending an HIV clinic in Mthatha between January 2021 and December 2024. Demographic characteristics, CD4 counts, viral load results, ART regimens, adherence status, and documented opportunistic infections were extracted. Viral load suppression was defined according to WHO guidelines as <1000 copies/mL. CD4 counts were categorized as <200, 200–499, and ≥500 cells/mm³. Multivariable logistic regression analysis was performed to identify independent predictors of opportunistic infections. Results: A total of 155 patients (105 females, 50 males) were included. Females demonstrated significantly higher mean CD4 counts than males (p = 0.037) and better ART adherence (p < 0.001). Tuberculosis, hepatitis B virus, and herpes simplex virus infections were more prevalent among males, whereas candidiasis was significantly more common among females (p = 0.034). In multivariable analysis, CD4 count < 200 cells/mm³, unsuppressed viral load, and poor ART adherence were independently associated with increased odds of opportunistic infections. Conclusions: Immune suppression and suboptimal ART adherence significantly increase the risk of opportunistic infections among HIV-positive patients in rural South Africa. Strengthening adherence interventions and early immune monitoring may reduce infection burden in high-prevalence settings. Full article

Tuberculosis therapy is prolonged partly because dormant subpopulations of Mycobacterium tuberculosis show reduced susceptibility to first-line drugs. Therefore, agents active against both replicating and non-replicating mycobacteria remain important to explore. Here, we investigated secondary metabolites from the Indonesian marine-derived fungus Aspergillus ostianus for activity against Mycobacterium smegmatis, a BSL-1 mycobacterial model, under aerobic and hypoxia-induced non-replicating conditions, and examined the underlying mechanism. Bioassay-guided fractionation and spectroscopic analysis identified three hydroxamate pyrazinones: neohydroxyaspergillic acid (NHAA), hydroxyaspergillic acid (HAA), and neoaspergillic acid (NAA). The MIC values were 1.56 µg/mL for NHAA and 3.13 µg/mL for HAA and NAA under both aerobic and hypoxic atmospheres. Time-kill kinetics showed ≥3-log10 CFU reductions within 24–72 h at 4–8× MIC under aerobic conditions and within 48–96 h at 4–8× MIC under hypoxia, with no regrowth at the final sampling point. Scanning electron microscopy and release of UV-absorbing intracellular material at OD₂₆₀/OD₂₈₀ were consistent with envelope disruption in both atmospheres. Antimycobacterial activity was attenuated in a concentration-dependent manner by exogenous Fe³⁺ and was reversed at 100 µM FeCl₃, whereas isoniazid activity was unaffected, supporting iron-reversible and pyrazinone-specific killing. Together with the established Fe³⁺-binding hydroxamate pharmacophore shared by this compound class, these findings support iron sequestration as a plausible mechanism and identify fungal hydroxamate pyrazinones as scaffolds that retain bactericidal activity against hypoxia-adapted non-replicating mycobacteria, warranting further evaluation in M. tuberculosis models. Full article

Background: Messenger RNA (mRNA) vaccines have emerged as a versatile platform beyond SARS-CoV-2, with expanding applications in infectious diseases and oncology. However, comprehensive evidence synthesis of non-SARS-CoV-2 mRNA vaccines remains limited. Methods: This systematic review followed PRISMA 2020 guidelines and was registered in PROSPERO (CRD420261323500). MEDLINE, Embase, Web of Science, Scopus, ClinicalTrials.gov, and WHO ICTRP were systematically searched for studies published between 1 January 2000 and 28 February 2026. Eligible studies included phase I–III clinical trials and in vivo preclinical studies evaluating non-SARS-CoV-2 mRNA vaccines. Two reviewers independently screened studies, extracted data, and assessed risk of bias using RoB 2, ROBINS-I, and SYRCLE tools. Findings were synthesized narratively because of substantial heterogeneity. Results: A total of 40 studies met the eligibility criteria and were included in the review, comprising 20 clinical studies and 20 preclinical studies. Advanced clinical programs targeted influenza and respiratory syncytial virus (RSV), with phase III trials displaying seroconversion rates above 70% with good safety profiles. Preliminary phase I studies for HIV, cytomegalovirus, rabies, and personalized cancer mRNA vaccines showed promising humoral and cellular immune responses. Preclinical studies showed strong antibody and T-cell responses against malaria, tuberculosis, Group B Streptococcus, and Zika virus. Most adverse events were mild to moderate, while serious vaccine-related adverse events were uncommon. Conclusions: Non-SARS-CoV-2 mRNA vaccines demonstrate substantial translational potential across infectious disease and oncology applications. Although the vaccine candidates have demonstrated promising immunogenicity and safety, most are in the early stages of development. This highlights the need for large trials, long-term safety follow-up and better global representation. Full article

Background/Objectives: Nanopore sequencing is increasingly used in mycobacterial diagnostics, where clinical microbiologists and diagnostic laboratories must decide when broad metagenomic next-generation sequencing (mNGS) or focused targeted next-generation sequencing (tNGS) is most appropriate. This review examined reported clinical and laboratory roles of nanopore mNGS and tNGS in tuberculosis (TB) and nontuberculous mycobacterial (NTM) settings. Methods: Targeted searches of PubMed/MEDLINE, Embase, Web of Science Core Collection, and Scopus were refreshed on 4 April 2026. Thirty-five records spanning original clinical studies, evidence syntheses, and guideline-context documents were included. Results: Nanopore mNGS is most useful for broad organism detection and diagnostic rescue in unresolved pulmonary and extrapulmonary presentations, particularly when first-line testing is negative, discordant, low-yield, or when mixed infection is suspected. Nanopore tNGS appears better aligned with predefined TB confirmation and resistance-focused workflows because targeted regions allow more standardized interpretation. Agreement is strongest for rifampicin- and isoniazid-related resistance targets. In NTM settings, evidence is stronger for detection and species identification than for disease-level diagnosis. Common implementation constraints include pre-analytical variation, contamination control, host-background interference, inconsistent bioinformatics, and limited workforce capacity. Conclusions: A practical tiered approach is supported in which mNGS is positioned mainly for diagnostic rescue and discovery, whereas tNGS is considered for predefined workflows requiring standardized target interrogation and resistance-associated mutation reporting under local validation and quality systems. Full article

Background/Objectives: Tuberculosis (TB), caused by Mycobacterium tuberculosis complex, remains a major global health challenge. Recently, D-enantiomer of human lactoferricin 1–11 (D-form hLF 1–11), a short peptide derived from the N-terminal region of lactoferrin, has demonstrated potent antimycobacterial activity. However, its direct mechanism of action has not yet been elucidated. Methods & Results: In the present study, M. smegmatis was employed as a model organism to investigate the mechanism underlying D-form hLF 1–11 activity. Initially, the minimum inhibitory concentration (MIC) was determined and the results revealed growth inhibition at 400 µg/mL. Live/dead fluorescence staining demonstrated mycobactericidal activity, as indicated by increased propidium iodide (PI) uptake relative to the untreated control. Scanning electron microscopy and high-resolution fluorescence microscopy revealed membrane disruption and substantial morphological deformation, along with a time-dependent accumulation of the peptide at the membrane and inside the cells. Furthermore, label-free quantitative proteomic analysis of peptide-treated cells revealed extensive metabolic alterations in carbon metabolism, acetyl-CoA-dependent lipid biosynthesis, oxidative stress defense, translational machinery, and energy production systems. Conclusions: Collectively, these findings provide mechanistic insights into the antimycobacterial activity of D-form hLF 1–11 against M. smegmatis. Full article

Background/Objectives: Tuberculosis, caused by Mycobacterium tuberculosis, remains a major global health challenge requiring novel prevention strategies. This study aims to developed an immunoinformatics-guided framework coupled with experimental screening to prioritize for multi-epitope mRNA vaccine design. Methods: Eight immunologically relevant antigens were computationally analyzed to predict cytotoxic (CTL) epitopes and helper T lymphocyte (HTL) epitopes. Population coverage, immune simulation, molecular docking, and normal mode analysis (NMA) were performed in silico. To evaluate peptide immunoreactivity, human IFN-γELISPOT assays were conducted using the candidate peptides, though HLA restriction was not experimentally validated. Results: The workflow identified 14 candidate CTL and 8 HTL epitopes, yielding an estimated global population coverage of 82.6% (60.7% in China; 51.2% in Indonesia). Immune simulations predicted robust humoral and Th1-associated cellular responses, though sustained CD8⁺ memory responses appeared limited. Docking and NMA suggested favorable structural interactions with TLR3 and TLR4. Crucially, the IFN-γ ELISPOT assay validated eight reactive epitopes that partially coincided with computational predictions within the tested donor group. Conclusions: This study establishes an integrated computational–experimental workflow for T cell epitope prioritization. The identified reactive epitopes provide a preliminary immunological basis and candidate pool for the future design and evaluation of multi-epitope mRNA vaccine strategies against tuberculosis. Full article

Differential diagnosis of tuberculosis (TB) and non-tuberculous mycobacteria (NTM) is extremely challenging, especially in a high TB burden setting like India. A definitive diagnosis of NTM, along with additional speciation, is warranted to improve NTM management. Beyond the diagnosis of NTM and its speciation, clinical correlation is vital for differentiating NTM colonisation or contamination from disease. In this cross-sectional, retrospective analysis, both pulmonary and extrapulmonary samples from 1121 presumptive NTM patients from Tuberculosis Units across the country and from other private hospitals were included. Composite diagnosis were performed using X-rays, nucleic acid amplification tests, smear microscopy, and mycobacterial growth indicator tube cultures, with speciation of NTM isolates confirmed by line probe assay. Of the 1121 presumptive NTM patients, 66.0% were smear-negative, 44.7% had X-ray changes, and 98.0% were M. tuberculosis-negative according to the nucleic acid amplification test. Cultures identified 310 patients as NTM-positive, including 22 extrapulmonary cases. Speciation was performed for 135 NTM-positive isolates, where M. abscessus was identified as the predominant species in 30.4%, followed by M. kansasii in 25.1%. Our study demonstrated that although the composite diagnosis of NTM holds promise for identifying pulmonary and extrapulmonary NTM, culture (mean confirmation rate of 30–40% over 5 years) remains the gold standard, with NTM speciation by line probe assay completing the diagnosis. Full article