The diagnosis of leprosy remains challenging due to its clinical similarity with other infectious diseases and the variable sensitivity and specificity of current laboratory tests. Therefore, the identification and/or de...The diagnosis of leprosy remains challenging due to its clinical similarity with other infectious diseases and the variable sensitivity and specificity of current laboratory tests. Therefore, the identification and/or development of novel antigens is crucial for improving diagnostic accuracy. In this study, we designed a novel recombinant multi-polypeptide construct, termed M3, which was composed of specific B-cell epitopes from nine Mycobacterium leprae proteins, previously shown to be antigenic in leprosy. The recombinant M3 protein was expressed, purified, and evaluated as an antigen for the diagnosis of both paucibacillary (PB) and multibacillary (MB) disease, by using paired serum and urine samples from 380 participants. ELISA was performed using samples from PB (n = 50) and MB (n = 55) leprosy patients, their household contacts [PBC (n = 30) and MBC (n = 40), respectively], healthy individuals living in endemic region (n = 55), and patients with visceral (n = 30) and tegumentary (n = 45) leishmaniasis, Chagas disease (n = 35), tuberculosis (n = 15), and HIV-infection (n = 25). In serum-based ELISA, M3 showed sensitivity, specificity, positive and negative predictive values, and Youden index of 94.5 %, 100 %, 96.8 %, 97.4 %, and 0.91, respectively; whilst in a urine-based ELISA, the corresponding values were 96.4 %, 100 %, 98.8 %, 98.2 %, and 0.96, respectively. Additionally, paired serum and urine samples from 15 PB and 20 MB patients collected before and after treatment, revealed that anti-M3 antibodies decreased by more than 50 % post-therapy when using both analytes. These pilot findings suggest a potential biological role of the recombinant M3 protein for diagnosis of PB and MB leprosy.
Lineage 1 is an ancestral lineage of the Mycobacterium tuberculosis complex (MTBC) comprising five sublineages. In a previous study, we suggested that a representative of sublineage L1.1.3, present in both Mozambique and...Lineage 1 is an ancestral lineage of the Mycobacterium tuberculosis complex (MTBC) comprising five sublineages. In a previous study, we suggested that a representative of sublineage L1.1.3, present in both Mozambique and northern Brazil, SIT129, may have been brought to Brazil by sea during the long history of slavery that lasted, between Africa and Brazil, from the early 16th century to the mid-19th century. In this study, using a combination of new comparative genomics results and human migration data extracted from the SlaveVoyages.org database, we sought to more precisely reconstruct the scenario for the introduction of L1 genotypes into Brazil. We present results showing substantial similarities between the MTBC population structure in present-day Mozambique and Brazil across three subclades, L1.1.2, L1.1.3, and L1.2.2, and convergent historical data. Indeed, several introductions between the 16th and 19th centuries could explain the higher contemporary prevalence of L1 in northern Brazil. Our data do not allow us to decide between a direct introduction of L1 isolates into northern Brazil and intra-Brazilian transmission from the main southern ports, which seems likely. Other less likely scenarios are also discussed.
Eight burials from the multi-period rural settlement site of Ranelagh near Roscommon town, Ireland, with palaeopathological lesions suggestive of skeletal tuberculosis or brucellosis were examined by ancient DNA (aDNA) t...Eight burials from the multi-period rural settlement site of Ranelagh near Roscommon town, Ireland, with palaeopathological lesions suggestive of skeletal tuberculosis or brucellosis were examined by ancient DNA (aDNA) testing. Tuberculosis infection (MTB complex DNA) was confirmed in five individuals - an 11th-13th CE adolescent female (14.5-17.5 years), two young adults females (18-35 years, 7th-10th CE), one adolescent of unknown sex and one middle-aged adult (35-50 years, medieval in date). In the latter case, the differential diagnosis included brucellosis due to the presence of small multifocal lytic lesions in the lower spinal vertebrae. However, this individual and all cases tested negative for Brucella species DNA. In two positive cases, lineage 4 (Euro-American) Mycobacterium tuberculosis DNA was identified in extracts obtained from tooth pulp cavities. These are the first archaeological individuals from Ireland to have had tuberculosis infection confirmed through aDNA analysis.
Rajamanickam A, Daniel EA, Gupte N
… +12 more, Thiruvengadam K, Chandrasekaran P, Pattabiraman S, Bhanu B, Sivaprakasam A, Paradkar M, Kulkarni V, Karyakarte R, Mave V, Gupta A, Hanna LE, Babu S
BACKGROUND: Identifying host biomarkers associated with progression from Mycobacterium tuberculosis infection to active tuberculosis (TB) could support early risk stratification in household contacts (HHCs). This explora...BACKGROUND: Identifying host biomarkers associated with progression from Mycobacterium tuberculosis infection to active tuberculosis (TB) could support early risk stratification in household contacts (HHCs). This exploratory study evaluated baseline plasma immune biomarkers in HHCs of pulmonary TB (PTB) patients to assess their association with subsequent disease development. METHODS: We analyzed baseline plasma samples from 15 progressors and 29 non-progressors enrolled from PTB-affected households. Acute-phase proteins (α-2-macroglobulin (α-2-M), C-reactive protein [CRP], haptoglobin (Hp), serum amyloid P (SAP)) and microbial translocation markers (lipopolysaccharide, lipid-binding protein, endotoxin core antibodies IgG, intestinal fatty acid-binding protein [iFABP], sCD14, and zonulin) were measured using Luminex and ELISA. Logistic regression and ROC analyses were performed as exploratory assessments of biomarker associations. RESULTS: Higher baseline levels of CRP, iFABP, and zonulin were observed among progressors compared with non-progressors. In univariable analyses, these biomarkers showed strong discriminatory ability (AUC ≥0.90), although estimates should be interpreted cautiously given the small sample size. A combined model including CRP, iFABP, and zonulin demonstrated high discriminatory performance (AUC 0.99 [95 % CI: 0.97-1.00]), but confidence intervals reflect the imprecision inherent to the limited dataset. CONCLUSIONS: In this exploratory cohort, elevated CRP, iFABP, and zonulin were associated with progression to active TB among household contacts. These preliminary findings suggest potential involvement of inflammatory and gut-barrier pathways in TB progression and warrant validation in larger, independent cohorts to define their translational utility.
SARS-CoV-2 infection modulates innate and adaptive immunity and likely impacts outcomes of infections such as Mycobacterium tuberculosis (MTB). We investigated the association between COVID-19 and latent tuberculosis inf...SARS-CoV-2 infection modulates innate and adaptive immunity and likely impacts outcomes of infections such as Mycobacterium tuberculosis (MTB). We investigated the association between COVID-19 and latent tuberculosis infection (LTBi), studying viral and mycobacterial antigen-stimulated responses in those with and without a history of COVID-19. We studied healthy Controls and COVID-19 convalescent subjects. Participants were screened for LTBi using an interferon-gamma release assay (IGRA). SARS-CoV-2 Spike- and MTB H37Rv-sonicate -stimulated cytokines from peripheral blood mononuclear cells (PBMCs) were measured. Spike-induced IFN-γ (p = 0.03), IL-6 (p = 0.018) and IL-2 (p = 0.04) levels were reduced in COVID-19 as compared with Controls. Within Controls, Spike induced higher cytokine levels in IGRA positive participants (p < 0.05). MTB-induced IFN-γ (0.003), IL-2 (p = 0.0021), IL-6 (p = 0.002), TNF-α (p = 0.02), and IL-10 (p = 0.04) levels were lowered in COVID-19. MTB- stimulated higher levels of proinflammatory cytokines were found in IGRA-positive Controls. Between IGRA positive participants, the COVID-19 group displayed lower Spike and MTB induced IFN-γ (0.003), IL-6 (0.0037), IL-2 (0.001), and TNF-α (0.005) levels. Further, MTB-induced IL-6/IL-10 and TNF-α/IL-10 ratios were higher in COVID-IGRA positive participants. Reduced SARS-CoV-2 and MTB activation of inflammatory cytokines reflects downregulated immunity after COVID-19. Further studies are required to assess whether LTBi and COVID-19 increase the risk of progression to tuberculosis.
BACKGROUND: Chronic inflammation fuels tissue damage and morbidity in numerous diseases, including Tuberculosis, underscoring the vital need for Host-Directed Therapies to safely modulate the exaggerated host response. W...BACKGROUND: Chronic inflammation fuels tissue damage and morbidity in numerous diseases, including Tuberculosis, underscoring the vital need for Host-Directed Therapies to safely modulate the exaggerated host response. We investigated the immunomodulatory potential of Sofalcone and Rebamipide alongside a novel Diaryl Ether derivative of Dehydrozingerone DHZ (6), hypothesizing that they exert therapeutic effects by targeting the central inflammatory driver, the NF-κB pathway. METHODS: We evaluated the safety and efficacy of the compounds in THP-1 macrophages infected with M. smegmatis. Mechanistic studies utilized Western blotting, immunofluorescence, and ELISA to track NF-κB activation. MMP activity was assessed by gelatin zymography, and ROS production was quantified using the DCFH-DA assay. RESULTS: All compounds exhibited low cytotoxicity and significantly reduced intracellular bacterial survival. Agents potently and consistently inhibited the NF-κB cascade, evidenced by ≈ 83 % suppression of upstream P-IKKα/IKKβ and up to ≈89 % reduction in p65 phosphorylation. This molecular blockade prevented p65 nuclear translocation and resulted in a downstream functional benefit: near total abolition of MMP-2/9 activity and ≈71 % mitigation of ROS production. CONCLUSION: Our results unequivocally validate NF-κB inhibition by DHZ (6), Sofalcone, and Rebamipide as a powerful strategy for HDT. These compounds are promising adjunct therapies to suppress host inflammation and limit tissue damage.
Understanding the perturbations in immune response across the spectrum of TB infection is still unclear. Here, we followed close contacts of pulmonary TB patients with serial QFT testing at 0, 3, 6, and 12 months, and st...Understanding the perturbations in immune response across the spectrum of TB infection is still unclear. Here, we followed close contacts of pulmonary TB patients with serial QFT testing at 0, 3, 6, and 12 months, and stratified them into six subgroups: QFT-increasing (low/high), QFT-converters (QFT-to QFT+), QFT + stable, and QFT-individuals. Despite these distinct QFT trajectories, we observed minimal differences in immune cell frequencies, activation profiles, and T-helper subset distributions among QFT subgroups, suggesting limited immunological stratification based on QFT dynamics. Ex vivo immune phenotyping, including CD4, CD8, NKT cell frequencies, memory T-cell subsets, and activated T-cells (HLA-DRCD38), failed to distinguish between QFT subgroups, suggesting blood-based immune profiling may not capture subtle immunological transitions among different QFT subgroups. Active TB (ATB) patients showed marked immune alterations, with elevated antigen-specific CD4 T-cells, activated T cells, intermediate monocytes, NK cells at-diagnosis, which declined following treatment, indicating immune recovery. This suggest, while ex vivo immune profiling effectively distinguishes ATB from non-diseased states, it lacks the sensitivity to resolve QFT-based subgroups. Findings suggest either immune similarity among close contacts regardless of QFT status or limits of blood-based profiling in detecting early changes, underscoring the difficulty of distinguishing QFT subgroups with conventional ex vivo approaches.
Toxin antitoxin (TA) systems are bicistronic genetic elements encoding for a stable toxin and its cognate labile antitoxin. The genome of Mycobacterium tuberculosis (M. tuberculosis) encodes a large repertoire of TA syst...Toxin antitoxin (TA) systems are bicistronic genetic elements encoding for a stable toxin and its cognate labile antitoxin. The genome of Mycobacterium tuberculosis (M. tuberculosis) encodes a large repertoire of TA systems and these are highly conserved in members of the M. tuberculosis complex. In the present study, we have characterised PezAT and MbcTA TA systems from M. tuberculosis. We show that the transcript levels of toxins and antitoxins belonging to PezAT and MbcTA were increased in M. tuberculosis exposed to oxidative stress, nitrosative stress and rifampicin. We also show that the relative levels of precursors for the peptidoglycan biosynthesis were increased in the PezT overexpression strain of M. smegmatis relative to uninduced cultures. Here, we have used temperature-sensitive mycobacteriophages to generate ΔpezAT and ΔmbcT mutant strains of M. tuberculosis. We demonstrate that the deletion of pezAT reduced the growth of M. tuberculosis upon exposure to detergent stress or rifampicin. However, the deletion of mbcT does not affect M. tuberculosis growth in various stress conditions. We also report that both PezAT and MbcT are dispensable for M. tuberculosis growth in macrophages and guinea pigs. Overall, these findings suggest that functional redundancy exists between TA systems.
SETTING: and objective: To evaluate the diagnostic utility of Cell-free DNA (Cf-DNA)-based multi-targeted real-time PCR (Mrt-PCR) for diagnosing tuberculous meningitis, the most severe form of extrapulmonary tuberculosis...SETTING: and objective: To evaluate the diagnostic utility of Cell-free DNA (Cf-DNA)-based multi-targeted real-time PCR (Mrt-PCR) for diagnosing tuberculous meningitis, the most severe form of extrapulmonary tuberculosis. DESIGN: A total of 170 cerebrospinal fluid samples (CSF) of tuberculous meningitis [Definite TBM (n = 146), Probable TBM (n = 24)] and 100 non-TB controls were subjected to Cf-DNA Mrt-PCR test using IS6110, IS1081, and nrdZ gene targets. The performance was also evaluated against Truenat MTB Plus assay and GeneXpert MTB/RIF Ultra. RESULTS: The sensitivity, specificity, positive predictive value and negative predictive value of Cf-DNA M-rtPCR was 78.24 %, 100 %, 100 % and 72.99 %, respectively in overall diagnosis of TBM and 82.88 %, 100 %, 100 % and 80 %, respectively in diagnosing 'definite' TBM. Cf-DNA M-rtPCR detected 14 additional cases that were missed by Truenat MTB Plus assay and GeneXpert MTB/RIF Ultra. Among the different gene targets used, eight cases were detected only by IS1081, three only by IS6110 and one only by nrdZ gene. Six cases were reported rifampicin-resistant by both Truenat MTB Plus assay and GeneXpert MTB/RIF Ultra and were also confirmed by rpoB sequencing. CONCLUSION: The detection of cell-free DNA, along with M-rtPCR could serve as a reliable tool for diagnosing tuberculous meningitis directly from CSF samples.
Inhibition of PI3Kγ is an attractive therapeutic target for the development of novel host-directed modulating strategies for the treatment of infectious diseases. This work investigated the antimicrobial potential of AS6...Inhibition of PI3Kγ is an attractive therapeutic target for the development of novel host-directed modulating strategies for the treatment of infectious diseases. This work investigated the antimicrobial potential of AS605240, a selective inhibitor of PI3Kγ, in pre-clinical models of mycobacterial infections. Of note, we observed that treatment with AS605240 effectively reduced both intracellular M. smegmatis and M. tuberculosis counts in RAW 264.7 cells. Moreover, treatment of M. tuberculosis-infected cells with AS605240 increased TNF-α and decreased IL-1β levels compared to the infected group. Importantly, we found that AS605240 is bacteriostatic in the lungs and bactericidal in spleens from M. tuberculosis-infected mice. Our data provide novel evidence on the relevance of PI3Kγ as a novel molecular target for new anti-tubercular drugs.
Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (MTB), Bacille Calmette-Guérin (BCG) is the only licensed vaccine for tuberculosis, but it provides limited and inconsistent protection agai...Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (MTB), Bacille Calmette-Guérin (BCG) is the only licensed vaccine for tuberculosis, but it provides limited and inconsistent protection against pulmonary TB in adults. Furthermore, current diagnostic methods show low sensitivity in latent stages, and the standard treatment is long, complex, and conducive to the emergence of drug-resistant strains. In this context, heat shock protein X (HspX), also known as Rv2031c or α-crystallin, has emerged as a potential biomarker and vaccine candidate. This 16-kDa protein is predominantly expressed under stressful conditions such as hypoxia and nitric oxide exposure, which are characteristic of the granulomatous microenvironment where MTB persists. Its expression during latency and high immunogenicity has been demonstrated in individuals with latent tuberculosis infection and those vaccinated with BCG. Preclinical studies have shown that recombinant HspX potentiates the host immune response used as a component in subunit vaccines, either alone or in combination with other antigens. Incorporating this protein into new diagnostic, therapeutic, and vaccine strategies could optimize disease control. This review explores HspX's multifaceted role and potential applications in tuberculosis diagnosis, treatment, and vaccine development.
Tuberculosis (TB), a disease of old, continues to plague humans after its declaration as a global health emergency in 1993. Over the years, studies have focused on understanding the causative pathogen, Mycobacterium tube...Tuberculosis (TB), a disease of old, continues to plague humans after its declaration as a global health emergency in 1993. Over the years, studies have focused on understanding the causative pathogen, Mycobacterium tuberculosis complex (MTBC), and its interaction with humans from TB infection to progression to active disease. It is now known that MTBC lineage diversity impacts several disease presentations and outcomes, including disease progression and severity, virulence and antimicrobial resistance, transmissibility, and host response. Some of these lineages are highly geographically restricted, and prominent amongst them are lineages 5 and 6 of Mycobacterium africanum (Maf), mainly found in West Africa, with cases outside of this region usually prevalent in individuals of West African descent. Several hypotheses have been propounded to investigate these restrictions, ranging from the locality of an animal reservoir in certain areas to the emigration of Maf into West Africa but not spreading globally because Mycobacterium tuberculosis sensu stricto (Mtbss) outcompeted it. Another hypothesis, which states that host genetic factors can influence host susceptibility to some MTBC lineages, as well as TB progression to the severe disease state, appears more widely accepted. However, the exact mechanisms mediating this susceptibility have not been fully explored. This review seeks to highlight the advances made towards understanding the geographical restrictions of Maf and the host-pathogen interactions leading to the coevolution of Maf and humans in West Africa.
Rivas-Castro A, Manzano Espinosa IR, Sepúlveda DV
… +9 more, Martínez EPS, Choreño-Parra JA, Gama YS, Alonso Villaseñor AS, Argueta Muñoz FD, Gómez Apo E, Salinas-Lara C, Sánchez-Garibay C, Tena Suck ML
INTRODUCTION: Central nervous system tuberculosis (CNS-TB) is the most lethal form of tuberculosis, characterized by severe clinical manifestations and distinctive histopathological changes. Although several cytokines ar...INTRODUCTION: Central nervous system tuberculosis (CNS-TB) is the most lethal form of tuberculosis, characterized by severe clinical manifestations and distinctive histopathological changes. Although several cytokines are implicated in the immune response to Mycobacterium tuberculosis, their relationship with disease severity and histological alterations remains unclear. This study aimed to evaluate the associations between cytokine expression, histopathological changes, and clinical features in patients with CNS-TB. METHODS: We conducted a retrospective, observational, cross-sectional, descriptive study of 25 biopsies that fulfilled criteria for CNS-TB at the National Institute of Neurology and Neurosurgery "Manuel Velasco Suárez." Cases were classified according to the Suzaan Marais criteria. Immunohistochemistry was performed to assess IL-1β, IL-4, IL-10, IL-17, IL-23, TNF-α, and IFN-γ expression in meningeal and parenchymal tissues. RESULTS: Strong expression of IL-1β, IL-17, IL-23, and TNF-α was observed in glial cells, vascular endothelial cells, and macrophages, particularly within inflammatory and vascular lesions. CONCLUSIONS: Although no direct correlation was found between cytokine expression and clinical severity, the findings support a predominant pro-inflammatory response mediated by IL-1β, IL-17, IL-23, and TNF-α, associated with tissue and vascular damage. The elevated cytokine expression in foam cells suggests a potential role in local immune regulation in CNS-TB.