Mycobacterium Tuberculosis Research Articles

Inactivation of Phosphoserine/Threonine Phosphatase PstP in H37Rv Mycobacterium tuberculosis by In silico Drug Design Approach

Inactivation of Phosphoserine/Threonine Phosphatase PstP in H37Rv Mycobacterium tuberculosis by In silico Drug Design Approach

Targeted next-generation sequencing - a promising approach in the diagnosis of Mycobacterium tuberculosis and drug resistance.

Targeted next-generation sequencing (tNGS) offers a high-throughput, culture-independent approach that delivers a comprehensive resistance profile in a significantly shorter turn-around time, making it promising in enhancing tuberculosis (TB) diagnosis and informing treatment decisions. This study aims to evaluate the performance of tNGS in the TB diagnosis and drug resistance detection of Mycobacterium tuberculosis (MTB) using MTB clinical isolates and bronchoalveolar lavage fluid (BALF) samples. A total of 143 MTB clinical isolates were assessed, tNGS, phenotypic antimicrobial susceptibility testing (AST), and AST based on whole genome sequencing (WGS) exhibited high concordance rates, averaging 95.10% and 97.05%. Among 158 BALF samples, culture, Xpert MTB/RIF, and tNGS reported 29, 70 and 111 positives, respectively. In the confirmed cases with etiological evidence (smears, cultures, or molecular test), the positive rate of tNGS (73/83, 87.95%) was higher than that of Xpert MTB (67/83, 80.72%). Additionally, 45% (27/60) of clinically diagnosed cases (with imaging or immunological evidence) were positive for tNGS. Further validation on the discrepant results between tNGS and Xpert MTB/RIF with droplet digital PCR (ddPCR) yielded 35 positives, tNGS detected all, and Xpert MTB/RIF only identified 6 positives. In conclusion, tNGS demonstrates robust and rapid performance in the identification of MTB and its associated drug resistance, and can be directly applied to clinical samples, positioning it as a promising approach for laboratory testing of tuberculosis.

The divergent outcome of IL-4Rα signalling on Foxp3 T regulatory cells in listeriosis and tuberculosis

IntroductionForkhead box P3 (Foxp3) T regulatory cells are critical for maintaining self-tolerance, immune homeostasis, and regulating the immune system.MethodsWe investigated interleukin-4 receptor alpha (IL-4Rα) signalling on T regulatory cells (Tregs) during Listeria monocytogenes (L. monocytogenes) infection using a mouse model on a BALB/c background, specifically with IL-4Rα knockdown in Tregs (Foxp3creIL-4Rα−/lox).ResultsWe showed an impairment of Treg responses, along with a decreased bacterial burden and diminished tissue pathology in the liver and spleen, which translated into better survival. Mechanistically, we observed an enhancement of the Th1 signature, characterised by increased expression of the T-bet transcription factor and a greater number of effector T cells producing IFN-γ, IL-2 following ex-vivo stimulation with heat-killed L. monocytogenes in Foxp3creIL-4Rα-/lox mice. Furthermore, CD8 T cells from Foxp3creIL-4Rα-/lox mice displayed increased cytotoxicity (Granzyme-B) with higher proliferation capacity (Ki-67), better survival (Bcl-2) with concomitant reduced apoptosis (activated caspase 3). In contrast to L. monocytogenes, Foxp3creIL-4Rα-/lox mice displayed similar bacterial burdens, lung pathology and survival during Mycobacterium tuberculosis (M. tuberculosis) infection, despite increased T cell numbers and IFN-γ, TNF and IL-17 production.ConclusionOur results demonstrated that the diminished IL-4Rα signalling on Foxp3+ T regulatory cells resulted in a loss of their functionality, leading to survival benefits in listeriosis but not in tuberculosis.

MINCLE and TLR9 agonists synergize to induce Th1/Th17 vaccine memory and mucosal recall in mice and non-human primates

Development of new vaccines tailored for difficult-to-target diseases is hampered by a lack of diverse adjuvants for human use, and none of the currently available adjuvants induce Th17 cells. Here, we develop a liposomal adjuvant, CAF®10b, that incorporates Mincle and Toll-like receptor 9 agonists. In parallel mouse and non-human primate studies comparing to CAF® adjuvants already in clinical trials, we report species-specific effects of adjuvant composition on the quality and magnitude of the responses. When combined with antigen, CAF®10b induces Th1 and Th17 responses and protection against a pulmonary infection with Mycobacterium tuberculosis in mice. In non-human primates, CAF®10b induces higher Th1 responses and robust Th17 responses detectable after six months, and systemic and pulmonary Th1 and Th17 recall responses, in a sterile model of local recall. Overall, CAF®10b drives robust memory antibody, Th1 and Th17 vaccine-responses via a non-mucosal immunization route across both rodent and primate species.

Verification of Sub-national Claim for ‘TB-free’ Status of Rajsamand District, Rajasthan, India-2020

Background: Tuberculosis (TB) caused by Mycobacterium tuberculosis is an opportunistic infection that affects the lungs in humans. Caseload of TB has been deemed it endemic in various regions of the world and India is on top of that list. The Government of India has set out a goal to eliminate tuberculosis in India by the year 2025 with the help of the National Strategic Plan (NSP) 2017–25. The objective of the index study was to verify the claim of Rajsamand district in Rajasthan, that it witnessed a >20% reduction in TB incidence between 2015 and 2020. Methodology: This was a mixed-methods study. Quantitative component (cross-sectional study): Secondary data review (review of records from Nikshay notification systems and National tuberculosis elimination program (NTEP) reports, utilization of drugs in the public sector, and drug sales in the private sector). Primary data collection through survey. Qualitative component: nominal group technique and key informant interviews. Result: Percentage decline of a ≥ 20% incidence in 2020 compared to baseline 2015 was not seen. The number needed to test (increased by 18.26% in 2020 as compared to the baseline of 2015. TB Score of Rajsamand district for the year 2020 was 68.34 out of 100. Conclusion: Rajsamand district’s claim of ≥20% reduction in TB incidence in the year 2020, was rejected.

Exploring the properties of antituberculosis drugs through QSPR graph models and domination-based topological descriptors

Tuberculosis (TB) is a global health concern caused by the bacterium Mycobacterium tuberculosis. This infectious disease primarily affects the lungs but can also impact other organs. Effective TB control involves early diagnosis, appropriate treatment with a combination of antibiotics, and public health measures to prevent transmission. However, ongoing challenges include drug-resistant strains and socioeconomic factors influencing its prevalence. Drugs such as isoniazid, pyrazinamide, ethambutol, ethionamide, linezolid, and levofloxacin are approved for the treatment of drug-susceptible tuberculosis. The properties and other activities of the drug, can be analyzed by modelling its chemical structure in terms of a molecular graph \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$G=\\left(V,E\\right)$$\\end{document}, by considering the atoms as the vertex set \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$V\\left(G\\right)$$\\end{document} and the bonds between the two atoms as the edge set \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$E\\left(G\\right)$$\\end{document}. A molecular descriptor or topological index of \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$G$$\\end{document} represents the corresponding chemical molecule as a numerical value. Domination is one of the key concepts in the molecular structure used to analyze the properties of atoms. In this article, the domination distance-based topological indices of the drugs isoniazid, pyrazinamide, ethambutol, ethionamide, linezolid, and levofloxacin are computed to conduct QSPR (Quantitative Structure–Property Relationship) analysis, exploring their physicochemical and ADMET (Absorption, Distribution, Metabolism, Excretion, and Toxicity) properties. Quadratic regression is then used in the QSPR analysis to examine the physicochemical and ADMET properties of these drugs. The results of this analysis indicate that the domination Schultz index and domination SM index are the indices most strongly correlated with the majority of the physicochemical and ADMET properties. The QSPR analysis can also be extended to analogs of these drugs and to other treatment drugs, such as rifampin and rifapentine, to further explore their properties.

Letter to the Editor: First detection of a Mycobacterium tuberculosis XDR clinical isolate harbouring an RpoB I491F mutation in a Ukrainian patient treated in Germany, October 2023.

Letter to the Editor: First detection of a Mycobacterium tuberculosis XDR clinical isolate harbouring an RpoB I491F mutation in a Ukrainian patient treated in Germany, October 2023.

Allosteric coupling of substrate binding and proton translocation in MmpL3 transporter from Mycobacterium tuberculosis.

Infections caused by Mycobacterium spp. are very challenging to treat, and multidrug-resistant strains rapidly spread in human populations. Major contributing factors include the unique physiological features of these bacteria, drug efflux, and the low permeability barrier of their outer membrane. Here, we focus on MmpL3 from Mycobacterium tuberculosis, an essential inner membrane transporter of the resistance-nodulation-division superfamily required for the translocation of mycolic acids in the form of trehalose monomycolates (TMM) from the cytoplasm or plasma membrane to the periplasm or outer membrane. The MmpL3-dependent transport of TMM is essential for the growth of M. tuberculosis in vitro, inside macrophages, and in M. tuberculosis-infected mice. MmpL3 is also a validated target for several recently identified anti-mycobacterial agents. In this study, we reconstituted the lipid transport activity of the purified MmpL3 using a two-lipid vesicle system and established the ability of MmpL3 to actively extract phospholipids from the outer leaflet of a lipid bilayer. In contrast, we found that MmpL3 lacks the ability to translocate the same phospholipid substrate across the plasma membrane indicating that it is not an energy-dependent flippase. The lipid extraction activity was modulated by substitutions in critical charged and polar residues of the periplasmic substrate-binding pocket of MmpL3, coupled to the proton transfer activity of MmpL3 and inhibited by a small molecule inhibitor SQ109. Based on the results, we propose a mechanism of allosteric coupling wherein substrate translocation by MmpL3 is coupled to the energy provided by the downhill transfer of protons. The reconstituted activities will facilitate understanding the mechanism of MmpL3-dependent transport of lipids and the discovery of new therapeutic options for Mycobacterium spp. infections.IMPORTANCEMmpL3 from Mycobacterium tuberculosis is an essential transporter involved in the assembly of the mycobacterial outer membrane. It is also an important target in undergoing efforts to discover new anti-tuberculosis drugs effective against multidrug-resistant strains spreading in human populations. The recent breakthrough structural studies uncovered features of MmpL3 that suggested a possible lipid transport mechanism. In this study, we reconstituted and characterized the lipid transport activity of MmpL3 and demonstrated that this activity is blocked by MmpL3 inhibitors and substrate mimics. We further uncovered the mechanism of how the binding of a substrate in the periplasmic domain is communicated to the transmembrane proton relay of MmpL3. The uncovered mechanism and the developed assays provide new opportunities for mechanistic analyses of MmpL3 function and its inhibition.

Detection of Mycobacterium bovis in Free-Ranging Sapajus nigritus, Argentina.

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

The effect of corticosteroids in developing active pulmonary tuberculosis among patients with COVID-19.

Corticosteroids can reduce the mortality rate among patients with severe COVID-19 pneumonia. However, opportunistic infections such as Mycobacterium tuberculosis are of concern, especially among those on high doses of corticosteroids. It is unknown whether the risk of developing subsequent TB infection is high or not among COVID-19 patients on high doses of corticosteroids. Hence, this study was conducted to address this gap of knowledge. We conducted a retrospective, cross-sectional study at the King Chulalongkorn Memorial Hospital from October 12, 2022 to June 30, 2023. Two hundred forty-three participants with documented COVID-19 diagnosis on high dose corticosteroids were enrolled into the study. Baseline characteristics and risk factors of developing TB were collected. The prevalence of TB was significantly different among participants with chronic kidney disease (CKD) stages 2-4 and chronic lung diseases. The incidence of TB post 1-year diagnosis of COVID-19 was 4 out of 243 patients (1.6%) or 1,646 cases per 100,000 person-year. The mortality rate among subsequent TB group was significantly much higher than the non-TB group (50% vs 0.4%; p-value = 0.001). COVID-19 participants on high doses of corticosteroids also were co-infected with other infections such as bacteria (37.1%), fungi (5.3%), and Pneumocystis jirovecii (PJP) (1.2%). We found that the incidence of TB in participants with COVID-19 on high doses of corticosteroids was 11 times higher than the general population. Therefore, we recommend screening for latent TB among these patients to prevent/early diagnose TB disease.

Analysis of Tuberculosis Control (Case Study in The City of Mojokerto) Year 2024

This study aims to analyze the tuberculosis (TB) situation in Mojokerto City, focusing on the increase in cases among children and identifying effective countermeasures. TB, a preventable and curable infectious disease caused by Mycobacterium tuberculosis, is particularly problematic in developing countries like Indonesia due to high incidence and mortality rates, necessitating a comprehensive control strategy. Employing qualitative methods, this research delves into the practices, perceptions, and social dynamics affecting TB control efforts in Mojokerto, illuminated by the latest data showing a significant increase in pediatric TB cases and the public health situation. Findings reveal a TB incidence rate of 385 per 100,000 inhabitants, a case detection rate of 75%, and a notably high pediatric TB case proportion of 32%. The effectiveness of case tracking by health centers varies, with an average of only 54% receiving Tuberculosis Preventive Therapy (TPT). Major obstacles include limited access to radiology services, social stigma, and the need for adequate health facilities. Innovations such as the "Mbak Desi Fights TB" program and enhanced cross-sector cooperation are deemed crucial to strengthen control strategies. The study concludes that a multifaceted strategy is essential for combating TB in Mojokerto, which should include enhanced case detection, health system strengthening, and community integration in control efforts. Increased collaboration across sectors and advancements in research and development are expected to accelerate TB elimination. This research offers efficient recommendations to improve TB control strategies, based on an in-depth analysis of the current conditions and social dynamics in Mojokerto.

The Concordance between GeneXpert MTB/RIF and Isothermal Amplification Assay for Detecting Mycobacterium tuberculosis

Tuberculosis (TB) is an infectious disease and one of the biggest causes of death worldwide. The main problem today is the lack of accurate and rapid tests to detect Mycobacterium tuberculosis (MTB). Several molecular methods have been developed to detect MTB. GeneXpert MTB/RIF® can detect MTB and rifampicin resistance simultaneously in <2 hours. Cross Priming Amplification (CPA) is one of the isothermal amplification assay methods that can detect MTB. Both of these methods are molecular rapid tests so they can detect MTB faster. This study aims to evaluate the concordance of GeneXpert MTB/RIF® results with CPA to detect MTB at Hasan Sadikin General Hospital, Bandung. This is an observational cross-sectional study. The subjects were patients with suspected pulmonary TB and examined with GeneXpert MTB/RIF® then CPA (Ustar EasyNAT MTC™) was also examined. This study used total sampling with 50 subjects and analyzed with Cohen's Kappa test. The results of GeneXpert MTB/RIF® and Ustar EasyNAT MTC™ in detecting MTB obtained Kappa of 0.662 (good agreement) with p-value <0.001. Of the 11 low positive samples on GeneXpert MTB/RIF® as many as 6 subjects (54.55%) had positive results, and 5 subjects (45.45%) had negative results on UStar EasyNAT MTC™. Meanwhile, of the 4 very low positive samples, there was only 1 sample with positive results on UStar EasyNAT MTC™. There is a match results between high and medium positive GeneXpert MTB/RIF® and UStar EasyNAT MTC™. However, there is a difference between low and very low positive results on GeneXpert MTB/RIF® and UStar EasyNAT MTC™.

Intestinal microbiome dysbiosis increases Mycobacteria pulmonary colonization in mice by regulating the Nos2-associated pathways

Increasing researches reveal gut microbiota was associated with the development of tuberculosis (TB). How to prevent or reduce Mycobacterium tuberculosis colonization in the lungs is a key measure to prevent TB. However, the data on gut microbiota preventing Mycobacterium colonization in the lungs were scarce. Here, we established the clindamycin-inducing intestinal microbiome dysbiosis and fecal microbial transplantation models in mice to identify gut microbiota’s effect on Mycobacterium’s colonization in the mouse lungs and explore its potential mechanisms. The results showed that clindamycin treatment altered the diversity and composition of the intestinal bacterial and fungal microbiome, weakened the trans-kingdom network interactions between bacteria and fungi, and induced gut microbiome dysbiosis in the mice. Gut microbiota dysbiosis increases intestinal permeability and enhances the susceptibility of Mycobacterium colonization in the lungs of mice. The potential mechanisms were gut microbiota dysbiosis altered the lung transcriptome and increased Nos2 expression through the ‘gut–lung axis’. Nos2 high expression disrupts the intracellular antimicrobial and anti-inflammatory environment by increasing the concentration of nitric oxide, decreasing the levels of reactive oxygen species and Defb1 in the cells, and promoting Mycobacteria colonization in the lungs of mice. The present study raises a potential strategy for reducing the risks of Mycobacteria infections and transmission by regulating the gut microbiome balance.

Engineered Mycobacteriophage TM4::GeNL Rapidly Determines Bedaquiline, Pretomanid, Linezolid, Rifampicin, and Clofazimine Sensitivity in Mycobacterium tuberculosis Clinical Isolates.

Drug-resistant tuberculosis is a growing public health threat, and early characterization of the resistance phenotype is essential for guiding treatment and mitigating the high mortality associated with the disease. However, the slow growth rate of Mycobacterium tuberculosis, the causative agent of tuberculosis, necessitates several weeks for conventional culture-dependent drug susceptibility testing (DST). In addition, there are no widely available molecular diagnostic assays for evaluating resistance to newer tuberculosis drugs or drugs with complex resistance mechanisms. We have developed a luciferase-based reporter mycobacteriophage assay that can determine drug resistance within 48 hours. We engineered the TM4 mycobacteriophage to express green enhanced nanoluciferase (GeNL) cassette and optimized DST for bedaquiline, pretomanid, linezolid, clofazimine, and rifampicin using clinical M. tuberculosis isolates. To assess the feasibility of this assay, we conducted a proof-of-principle study using 53 clinical M. tuberculosis isolates. TM4::GeNL phage DST effectively distinguished between sensitive and resistant isolates for bedaquiline and rifampicin at a concentration of 0.125 μg/mL. Optimal differentiation between sensitive and resistant isolates for pretomanid, clofazimine, and linezolid was achieved at concentrations of 0.5 μg/mL, 0.25 μg/mL, and 1 μg/mL, respectively. Additionally, TM4::GeNL DST identified low-level rifampicin resistance in clinical isolates even though they were classified as sensitive by Mycobacteria Growth Indicator Tube DST. TM4::GeNL reporter phage DST offers a rapid method to identify M. tuberculosis drug resistance, including resistance to newer tuberculosis drugs.

Cryo-electron microscopy structure of the di-domain core of Mycobacterium tuberculosis polyketide synthase 13, essential for mycobacterial mycolic acid synthesis.

Mycobacteria are known for their complex cell wall, which comprises layers of peptidoglycan, polysaccharides and unusual fatty acids known as mycolic acids that form their unique outer membrane. Polyketide synthase 13 (Pks13) of Mycobacterium tuberculosis, the bacterial organism causing tuberculosis, catalyses the last step of mycolic acid synthesis prior to export to and assembly in the cell wall. Due to its essentiality, Pks13 is a target for several novel anti-tubercular inhibitors, but its 3D structure and catalytic reaction mechanism remain to be fully elucidated. Here, we report the molecular structure of the catalytic core domains of M. tuberculosis Pks13 (Mt-Pks13), determined by transmission cryo-electron microscopy (cryoEM) to a resolution of 3.4 Å. We observed a homodimeric assembly comprising the ketoacyl synthase (KS) domain at the centre, mediating dimerization, and the acyltransferase (AT) domains protruding in opposite directions from the central KS domain dimer. In addition to the KS-AT di-domains, the cryoEM map includes features not covered by the di-domain structural model that we predicted to contain a dimeric domain similar to dehydratases, yet likely lacking catalytic function. Analytical ultracentrifugation data indicate a pH-dependent equilibrium between monomeric and dimeric assembly states, while comparison with the previously determined structures of M. smegmatis Pks13 indicates architectural flexibility. Combining the experimentally determined structure with modelling in AlphaFold2 suggests a structural scaffold with a relatively stable dimeric core, which combines with considerable conformational flexibility to facilitate the successive steps of the Claisen-type condensation reaction catalysed by Pks13.

Levofloxacin activity at increasing doses in a murine model of fluoroquinolone-susceptible and -resistant tuberculosis.

High-dose levofloxacin was explored in a clinical trial against multidrug-resistant tuberculosis and failed to show increased efficacy. In this study, we used a murine model to explore the efficacy of a dose increase in levofloxacin monotherapy beyond the maximum dose evaluated in humans. A total of 120 4-week-old female BALB/c mice were intravenously infected with 106 CFU of Mycobacterium tuberculosis H37Rv wild-type (WT) or isogenic H37Rv mutants harboring GyrA A90V or D94G substitutions; the MICs were 0.25, 4, and 6 µg/mL, respectively. Levofloxacin 250 and 500 mg/kg were given every 12 h (q12h) orally for 4 weeks. Pharmacokinetic parameters were determined after five doses. These two regimens decreased lung bacillary load in mice infected with H37Rv WT but not in mice infected with the A90V and D94G mutants. Levofloxacin 250 mg/kg q12h in mice generated pharmacokinetic parameters equivalent to 1,000 mg/d in humans, whereas 500 mg/kg q12h generated a twofold greater exposure than the highest equivalent dose tested in humans (1,500 mg/d). In our dose-response model, the effective concentration at 50% (EC50) produced an AUC/MIC (AUC0-24h/MIC) ratio of 167.9 ± 27.5, and at EC80 it was 281.2 ± 97.3. Based on this model, high-dose levofloxacin regimens above 1,000 mg/d are not expected to cause a significant increase in bactericidal activity. This study suggests no benefit of high-dose levofloxacin above 1,000 mg/d in the treatment of fluoroquinolone-susceptible or -resistant tuberculosis.

Machine learning algorithms to predict treatment success for patients with pulmonary tuberculosis.

Despite advancements in detection and treatment, tuberculosis (TB), an infectious illness caused by the Mycobacterium TB bacteria, continues to pose a serious threat to world health. The TB diagnosis phase includes a patient's medical history, physical examination, chest X-rays, and laboratory procedures, such as molecular testing and sputum culture. In artificial intelligence (AI), machine learning (ML) is an advanced study of statistical algorithms that can learn from historical data and generalize the results to unseen data. There are not many studies done on the ML algorithm that enables the prediction of treatment success for patients with pulmonary TB (PTB). The objective of this study is to identify an effective and predictive ML algorithm to evaluate the detection of treatment success in PTB patients and to compare the predictive performance of the ML models. In this retrospective study, a total of 1236 PTB patients who were given treatment under a randomized controlled clinical trial at the ICMR-National Institute for Research in Tuberculosis, Chennai, India were considered for data analysis. The multiple ML models were developed and tested to identify the best algorithm to predict the sputum culture conversion of TB patients during the treatment period. In this study, decision tree (DT), random forest (RF), support vector machine (SVM) and naïve bayes (NB) models were validated with high performance by achieving an area under the curve (AUC) of receiver operating characteristic (ROC) greater than 80%. The salient finding of the study is that the DT model was produced as a better algorithm with the highest accuracy (92.72%), an AUC (0.909), precision (95.90%), recall (95.60%) and F1-score (95.75%) among the ML models. This methodology may be used to study the precise ML model classification for predicting the treatment success of TB patients during the treatment period.

Synergistic role of Mycobacterium indicus pranii and human beta Defensin-2 as adjunctive therapy against Mycobacterium tuberculosis

Host-directed therapies (HDT) via modulation of specific host responses like inflammation can limit mycobacterial infection. HDTs could be included in current TB therapy as an adjunct to increase bacterial clearance and limit tissue damage to control spread. Individually, Mycobacterium indicus pranii (MIP) and human beta defensin-2 (hBD-2) are promising therapies for tuberculosis (TB). They can directly target the TB bacilli and enhance cell-mediated immune responses, which is limiting with conventional drugs. Therefore, our study investigated the combined application of MIP and hBD-2 to evaluate their efficacy in clearing infections caused by Mycobacterium smegmatis (M.smeg) and Mycobacterium tuberculosis (M.tb) (both avirulent; H37Ra and virulent strain; H37Rv) in THP-1 cells and human monocyte-derived macrophages (MDMs). A strong pro-inflammatory response was observed against the combination of MIP and hBD-2 which also correlated with a significant reduction in the bacterial load. This combination further showed protection against M.tb by enhancing pyroptosis in the infected cells. The study suggests the combined use of these potent immunomodulators, which could be employed as an effective mode of therapy as adjuvants against mycobacterial infections after validation in a suitable animal model.

Unusual Presentation of Tuberculosis in the Lungs and Lymph Nodes

Abstract Introduction/Objective Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis mainly affecting the lungs. Despite treatment advancements, TB remains a major cause of morbidity. This report presents a patient with active pulmonary and lymph node TB. Methods/Case Report The patient is a 41-year-old male from Bangladesh admitted for worsening cough and shortness of breath. Earlier, he had a cough and right cervical lymphadenopathy with lymph node biopsy demonstrating noncaseating granuloma and a positive Quantiferon-Gold. Outpatient workup included negative sputum TB PCR with negative cultures. The patient was febrile, tachycardic, and oxygenating normally. D-dimer was 1484. CT angiography was negative for emboli and showed mediastinal and right hilar lymphadenopathy. Imaging was otherwise non-indicative of active TB. We started rifampin, isoniazid, pyrazinamide, and ethambutol (RIPE) therapy with bronchoalveolar lavage for additional sputum sampling and excisional lymph node biopsy to rule out malignancy. Workup for autoimmune disease and sarcoidosis was negative. An acid-fast bacilli stain of the biopsy was positive, indicating active TB. The patient was discharged on RIPE with outpatient follow-up. Results (if a Case Study enter NA) NA Conclusion Histological findings of TB demonstrate caseating granulomatous lesions. Sarcoidosis can be considered in patients with hilar or mediastinal lymphadenopathy with noncaseating granulomatous lesions. However, both active and inactive TB should be considered in the differential diagnosis with biopsy results indicating noncaseating granuloma, as infected tissues can lack the central necrosis to produce caseation. Common chest radiographic findings in TB include pulmonary consolidation, hilar and mediastinal lymphadenopathy, disseminated miliary disease, and a normal chest radiograph. Our patient had just hilar and mediastinal lymphadenopathy. While a common finding in tuberculosis, this is also present in other inflammatory pathologies, emphasizing the importance of keeping a broad differential even in settings where initial biopsy and imaging results may indicate latent infection or other diagnoses. We utilized the expertise of pathologists to identify lymph nodes with active TB infection in a patient who previously had inconsistent pathological and radiological findings. Without the help of the pathologists in this case, diagnosing and treating this patient would not have been possible, showing the necessity of histopathological correlation with clinical and radiologic findings.

Red herrings and zebras and TB, oh my! Unexpected bone marrow presentation of Mycobacterium tuberculosis

Abstract Introduction/Objective Epithelioid granulomas are uncommon and non-specific findings in bone marrow samples, and should prompt additional testing for appropriate diagnosis and treatment, especially in the immunocompromised patient. The differential diagnosis for bone marrow granulomas includes infections, neoplasia, autoimmune disease, sarcoidosis, and drug/therapy reaction. Although bone marrow workup to delineate the cause of granulomas should include staining to rule out both infection and lymphoma, morphologic review is often inconclusive. Final diagnosis requires clinical consultation and additional testing. Methods/Case Report A 76-year-old male was admitted to the hospital from the emergency department for a level 2 trauma after being found on the ground ~3 hours after a fall. The patient reported a recent increasing frequency of falls with associated weakness, as well as an almost 30 lb weight loss over the last 3.5 months and low grade fevers over several weeks. His past medical history was significant for treatment for urothelial carcinoma with transurethral resection and BCG therapy, hypertension, coronary artery disease, and peripheral vascular disease. Imaging showed hepatosplenomgaly, suspicious for early cirrhosis, and mild peripancreatic lymphadenopathy. Initial laboratory abnormalities demonstrated mild anemia, low albumin, and mild increase in ALT, AST, and alkaline phosphotase. Initial Infectious Diseases workup was negative for causative etiology (blood cultures, tick-borne parasites, and respiratory pathogens). Patient symptoms did not improve and cytopenias worsened. A bone marrow biopsy showed multiple non-necrotizing epithelioid granulomas. Despite low clinical concern for infectious etiology, mycobacteria blood culture isolated Mycobacterium tuberculosis complex. Final results were available 112 days after specimen collection. Results (if a Case Study enter NA) NA Conclusion Although tuberculosis is commonly associated with granulomatous pathology, infection is very uncommon in North American patients who have not traveled and alternative diagnoses might initially be favored. A high suspicion for mycobacterial infections is essential for diagnosis even in low-incidence populations with no known exposure or risk factor(s).