Fragment Merging Using a Graph Database Samples Different Catalogue Space than Similarity Search.

Fragment merging is a promising approach to progressing fragments directly to on-scale potency: each designed compound incorporates the structural motifs of overlapping fragments in a way that ensures compounds recapitulate multiple high-quality interactions. Searching commercial catalogues provides one useful way to quickly and cheaply identify such merges and circumvents the challenge of synthetic accessibility, provided they can be readily identified. Here, we demonstrate that the Fragment Network, a graph database that provides a novel way to explore the chemical space surrounding fragment hits, is well-suited to this challenge. We use an iteration of the database containing >120 million catalogue compounds to find fragment merges for four crystallographic screening campaigns and contrast the results with a traditional fingerprint-based similarity search. The two approaches identify complementary sets of merges that recapitulate the observed fragment-protein interactions but lie in different regions of chemical space. We further show our methodology is an effective route to achieving on-scale potency by retrospective analyses for two different targets; in analyses of public COVID Moonshot and Mycobacterium tuberculosis EthR inhibitors, potential inhibitors with micromolar IC50 values were identified. This work demonstrates the use of the Fragment Network to increase the yield of fragment merges beyond that of a classical catalogue search.

Stool Xpert MTB/RIF as a possible diagnostic alternative to sputum in Africa: a systematic review and meta-analysis.

Introduction: Worldwide, COVID-19 pandemic lead to a large fall in the number of newly reported TB cases. In sub-Saharan Africa, microbiological diagnosis of TB is generally based on smear microscopy and Xpert MTB/RIF on sputum samples, but good quality sputum samples are often difficult to obtain, leading clinicians to rely on more invasive procedures for diagnosis. Aim of this study was to investigate pooled sensitivity and specificity of Xpert MTB/RIF on stool samples compared to respiratory microbiological reference standards in African countries. Methods: Four investigators independently searched PubMed, SCOPUS, and Web of Science until 12th October 2022, then screened titles and abstracts of all potentially eligible articles. The authors applied the eligibility criteria, considered the full texts. All the studies reported the data regarding true positive (TP), true negative (TN), false positive (FP) and false negative (FN). Risk of bias and applicability concerns were assessed with the Quadas-2 tool. Results: overall, among 130 papers initially screened, we evaluated 47 works, finally including 13 papers for a total of 2,352 participants, mainly children. The mean percentage of females was 49.6%, whilst the mean percentage of patients reporting HIV was 27.7%. Pooled sensitivity for Xpert MTB/RIF assay for detecting pulmonary tuberculosis was 68.2% (95%CI: 61.1-74.7%) even if characterized by a high heterogeneity (I2=53.7%). Specificity was almost 100% (99%, 95%CI: 97-100%; I2 = 45.7%). When divided for reference standard, in the six studies using sputum and nasogastric aspirate the accuracy was optimal (AUC = 0.99, SE = 0.02), whilst in the studies using only sputum for tuberculosis detection the AUC was 0.85 (with a SE = 0.16). The most common source of bias was exclusion of enrolled patients in the analysis. Conclusions: Our study confirms that, in Africa, stool Xpert MTB/RIF may be a useful rule-in test for children above and below 5 years of age under evaluation for pulmonary tuberculosis. Sensitivity increased substantially when using both sputum and nasogastric aspirate as reference samples.

Controlled human infection models in COVID-19 and tuberculosis: current progress and future challenges.

Controlled Human Infection Models (CHIMs) involve deliberately exposing healthy human volunteers to a known pathogen, to allow the detailed study of disease processes and evaluate methods of treatment and prevention, including next generation vaccines. CHIMs are in development for both tuberculosis (TB) and Covid-19, but challenges remain in their ongoing optimisation and refinement. It would be unethical to deliberately infect humans with virulent Mycobacteria tuberculosis (M.tb), however surrogate models involving other mycobacteria, M.tb Purified Protein Derivative or genetically modified forms of M.tb either exist or are under development. These utilise varying routes of administration, including via aerosol, per bronchoscope or intradermal injection, each with their own advantages and disadvantages. Intranasal CHIMs with SARS-CoV-2 were developed against the backdrop of the evolving Covid-19 pandemic and are currently being utilised to both assess viral kinetics, interrogate the local and systemic immunological responses post exposure, and identify immune correlates of protection. In future it is hoped they can be used to assess new treatments and vaccines. The changing face of the pandemic, including the emergence of new virus variants and increasing levels of vaccination and natural immunity within populations, has provided a unique and complex environment within which to develop a SARS-CoV-2 CHIM. This article will discuss current progress and potential future developments in CHIMs for these two globally significant pathogens.

Beneficial or detrimental activity of regulatory T cells, indoleamine 2,3-dioxygenase, and heme oxygenase-1 in the lungs is influenced by the level of virulence of Mycobacterium tuberculosis strain infection.

Tuberculosis (TB) caused by the complex Mycobacterium tuberculosis (Mtb) is the main cause of death by a single bacterial agent. Last year, TB was the second leading infectious killer after SARS-CoV-2. Nevertheless, many biological and immunological aspects of TB are not completely elucidated, such as the complex process of immunoregulation mediated by regulatory T cells (Treg cells) and the enzymes indoleamine 2,3-dioxygenase (IDO) and heme oxygenase 1 (HO-1). In this study, the contribution of these immunoregulatory factors was compared in mice infected with Mtb strains with different levels of virulence. First Balb/c mice were infected by intratracheal route, with a high dose of mild virulence reference strain H37Rv or with a highly virulent clinical isolate (strain 5186). In the lungs of infected mice, the kinetics of Treg cells during the infection were determined by cytofluorometry and the expression of IDO and HO-1 by RT-PCR and immunohistochemistry. Then, the contribution of immune-regulation mediated by Treg cells, IDO and HO-1, was evaluated by treating infected animals with specific cytotoxic monoclonal antibodies for Treg cells depletion anti-CD25 (PC61 clone) or by blocking IDO and HO-1 activity using specific inhibitors (1-methyl-D,L-tryptophan or zinc protoporphyrin-IX, respectively). Mice infected with the mild virulent strain showed a progressive increment of Treg cells, showing this highest number at the beginning of the late phase of the infection (28 days), the same trend was observed in the expression of both enzymes being macrophages the cells that showed the highest immunostaining. Animals infected with the highly virulent strain showed lower survival (34 days) and higher amounts of Treg cells, as well as higher expression of IDO and HO-1 one week before. In comparison with non-treated animals, mice infected with strain H37Rv with depletion of Treg cells or treated with the enzymes blockers during late infection showed a significant decrease of bacilli loads, higher expression of IFN-g and lower IL-4 but with a similar extension of inflammatory lung consolidation determined by automated morphometry. In contrast, the depletion of Treg cells in infected mice with the highly virulent strain 5186 produced diffuse alveolar damage that was similar to severe acute viral pneumonia, lesser survival and increase of bacillary loads, while blocking of both IDO and HO-1 produced high bacillary loads and extensive pneumonia with necrosis. Thus, it seems that Treg cells, IDO and HO-1 activities are detrimental during late pulmonary TB induced by mild virulence Mtb, probably because these factors decrease immune protection mediated by the Th1 response. In contrast, Treg cells, IDO and HO-1 are beneficial when the infection is produced by a highly virulent strain, by regulation of excessive inflammation that produced alveolar damage, pulmonary necrosis, acute respiratory insufficiency, and rapid death.

Tuberculosis in Poland in 2020.

AIM OF THE STUDY: To evaluate the main features of epidemiology of tuberculosis (TB) in 2020 in Poland and to compare with the situation in the European Union and European Economic Area (EU/EEA) countries. MATERIAL AND METHODS: Analysis of case-based data on TB patients from National TB Register, data on anti-TB drug susceptibility in cases notified in 2020, data from Statistics Poland on deaths from tuberculosis in 2019, data from National Institute of Public Health NIH - National Research Institute (NIPH NIH - NRI) on HIV-positive subjects for whom TB was an AIDS-defining disease, data from the report "European Centre for Disease Prevention and Control, WHO Regional Office for Europe. Tuberculosis surveillance and monitoring in Europe 2022 - 2020 data. Copenhagen: WHO Regional Office for Europe and Stockholm: European Centre for Disease Prevention and Control; 2022." RESULTS: In 2020, 3,388 TB cases were reported in Poland. The incidence rate was 8.8 cases per 100,000 with large variability between voivodeships from 5.5 to 13.3 per 100,000. A decrease in the incidence was found in 15 voivodeships, the most significant in Slaskie voivodship (63.9%). The number of all pulmonary tuberculosis cases was 3,237 i.e. 8.4 per 100,000. Pulmonary cases represented 95.5% of all TB cases. In 2020, 151 extrapulmonary TB cases were notified (4.5% of all TB cases). Pulmonary tuberculosis was bacteriologically confirmed in 2,573 cases (79.5% of all pulmonary TB cases, the incidence rate 6.7 per 100,000). The number of smear-positive pulmonary TB cases was 1,771 i.e. 4.6 per 100,000 (54.7% of all pulmonary TB cases). In 2020, there were 38 cases (15 of foreign origin) with multidrug resistant TB (MDR-TB) representing 1.6% of cases with known drug sensitivity. The incidence rates of tuberculosis were growing along with increasing age from 0.7 per 100,000 among children (0-14 years) to 15.0 per 100,000 among subjects in the age group 45-64 years, the incidence rate in the age group ≥65 years was 12.1 per 100,000. There were 39 cases in children up to 14 years of age (1.2% of the total) and 49 cases in adolescents between 15 and 19 years of age - rates 0.7 and 2.7 per 100,000 respectively. In 2020, there were 2,506 cases of tuberculosis in men and 882 in women. The TB incidence in men - 13.5 per 100,000 was 3.0 times higher than among women - 4.5. The biggest difference in the TB incidence between the two sex groups occurred in persons aged 50-54 years - 26.8 vs. 4.1 and in age group 55 to 59 years - 28.7 vs. 4.8. In 2020, there were 116 patients of foreign origin among all cases of tuberculosis in Poland (3.4%). In 2019, TB was the cause of death for 456 people (mortality rate - 1.2 per 100,000). CONCLUSIONS: TB incidence in Poland in 2020 was 36.7% lower than in 2019. Such significant declines in the incidence have not been observed in the last two decades. As in previous years, there were differences in incidence rates between voivodeships with an unexpectedly sharp decrease in incidence in Silesia (Slaskie voivodeship). The percentage of tuberculosis cases with bacteriological confirmation exceeded 78%, more than in EU/EEA countries (67.3%). The percentage of MDR-TB cases was still lower than the average in EU/EEA countries (1.6% vs. 3.8%). The highest incidence rates were found in Poland in the older age groups (EU/EEAaged 25 to 44). The percentage of children up to 14 years of age among the total number of TB patients was 1.2%, less than the average in EU/EEA countries (3.8%). The incidence of tuberculosis in men was three times higher than in women in Poland, and six times higher in patients aged 50 to 59. The impact of migration on the TB pattern in Poland has not yet become significant in 2020. The percentage of foreigners among TB patients was 3.4% (33% in EU/EEA countries).

A CRISPR-Cas12a-based platform for ultrasensitive rapid highly specific detection of Mycobacterium tuberculosis in clinical application.

Tuberculosis, caused by Mycobacterium tuberculosis (MTB), is the second leading cause of death after COVID-19 pandemic. Here, we coupled multiple cross displacement amplification (MCDA) technique with CRISPR-Cas12a-based biosensing system to design a novel detection platform for tuberculosis diagnosis, termed MTB-MCDA-CRISPR. MTB-MCDA-CRISPR pre-amplified the specific sdaA gene of MTB by MCDA, and the MCDA results were then decoded by CRISPR-Cas12a-based detection, resulting in simple visual fluorescent signal readouts. A set of standard MCDA primers, an engineered CP1 primer, a quenched fluorescent ssDNA reporter, and a gRNA were designed targeting the sdaA gene of MTB. The optimal temperature for MCDA pre-amplification is 67°C. The whole experiment process can be completed within one hour, including sputum rapid genomic DNA extraction (15 minutes), MCDA reaction (40 minutes), and CRISPR-Cas12a-gRNA biosensing process (5 minutes). The limit of detection (LoD) of the MTB-MCDA-CRISPR assay is 40 fg per reaction. The MTB-MCDA-CRISPR assay does not cross reaction with non-tuberculosis mycobacterium (NTM) strains and other species, validating its specificity. The clinical performance of MTB-MCDA-CRISPR assay was higher than that of the sputum smear microscopy test and comparable to that of Xpert method. In summary, the MTB-MCDA-CRISPR assay is a promising and effective tool for tuberculosis infection diagnosis, surveillance and prevention, especially for point-of-care (POC) test and field deployment in source-limited regions.

Role of non-coding RNAs in tuberculosis and their potential for clinical applications.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), remains the leading cause of mortality due to infectious diseases, only surpassed in 2020 by COVID-19. Despite the development in diagnostics, therapeutics, and evaluation of new vaccines for TB, this infectious disease remains uncontrollable due to the emergence of multidrug-resistant (MDR) and extremely drug-resistant (XDR) TB, among other factors. The development in transcriptomics (RNomics) has enabled the study of gene expression in TB. It is considered that non-coding RNAs (ncRNAs) from host [microRNAs (miRNAs)] and Mtb [small RNAs (sRNAs)] are important elements in TB pathogenesis, immune resistance, and susceptibility. Many studies have shown the importance of host miRNAs in regulating immune response against Mtb via in vitro and in vivo mice models. The bacterial sRNAs play a major role in survival, adaptation, and virulence. Here, we review the characterization and function of host and bacteria ncRNAs in TB and their potential use in clinical applications as diagnostic, prognostic, and therapeutic biomarkers.

[The problem of formation of new risks of medicinal resistance of patients with tuberculosis in condition of active use of antibiotics by population under COVID-19].

In pandemic conditions, situation of active and uncontrolled use by population of antimicrobial preparations treating COVID-19 occurs. So, new risks of development of medication resistance among patients with various infectious diseases, tuberculosis included, appear. The purpose of the study is to characterize prevalence of antimicrobial preparations use by population in relationship with development of medication resistance in patients with tuberculosis during COVID-19 pandemic. Material and methods. The analysis of sales of antimicrobial medicines was implemented on the basis of published official data from the joint-stock company DSM Group presenting monthly audit of the Russian pharmaceutical market. The determination of primary antibiotic resistance was carried out in 2018-2020 on 3312 patients with tuberculosis. The modified method of proportions on liquid nutrient medium in system with automated accounting of microorganisms growth, the method of absolute concentrations and the method of polymerase chain reaction with real-time detection were applied. The results of the study. It was established that the most demanding antimicrobial medications among population were ceftriaxone, azithromycin, levofloxacin, moxifloxacin, azithromycin. At the same time, the maximum increase in sales in 2020 up to 150% as compared with of 2019 was determined in medications derived from quinolone moxifloxacin, levofloxacin, which began to be used in treatment of coronavirus infection. At the same time, these medications are traditionally used in tuberculosis treatment. But in 2020, alarming trend was established that limits treatment of tuberculosis patients. The primary resistance of mycobacteria was also established in newly diagnosed tuberculosis patients, also for the same antimicrobial medications of quinolone derivatives, and increasing in proportion of patients with primary medication resistance to levofloxacin, moxifloxacin in 2020 as compared to 2018 was 189-480%. At the same time, increasing of resistance to other antibiotics made up to 60.8% on average. Conclusion. The study results imply alarming scenario of medication resistance shifts towards very virulent and highly medication-resistant genotypes. This trend can result in conditions of successful transmission of deadly medication-resistant mutants that can seriously undermine effectiveness of implemented programs of struggle with tuberculosis worldwide.

Activation of transcription factor CREB in human macrophages by Mycobacterium tuberculosis promotes bacterial survival, reduces NF-kB nuclear transit and limits phagolysosome fusion by reduced necroptotic signaling.

Macrophages are a first line of defense against pathogens. However, certain invading microbes modify macrophage responses to promote their own survival and growth. Mycobacterium tuberculosis (M.tb) is a human-adapted intracellular pathogen that exploits macrophages as an intracellular niche. It was previously reported that M.tb rapidly activates cAMP Response Element Binding Protein (CREB), a transcription factor that regulates diverse cellular responses in macrophages. However, the mechanism(s) underlying CREB activation and its downstream roles in human macrophage responses to M.tb are largely unknown. Herein we determined that M.tb-induced CREB activation is dependent on signaling through MAPK p38 in human monocyte-derived macrophages (MDMs). Using a CREB-specific inhibitor, we determined that M.tb-induced CREB activation leads to expression of immediate early genes including COX2, MCL-1, CCL8 and c-FOS, as well as inhibition of NF-kB p65 nuclear localization. These early CREB-mediated signaling events predicted that CREB inhibition would lead to enhanced macrophage control of M.tb growth, which we observed over days in culture. CREB inhibition also led to phosphorylation of RIPK3 and MLKL, hallmarks of necroptosis. However, this was unaccompanied by cell death at the time points tested. Instead, bacterial control corresponded with increased colocalization of M.tb with the late endosome/lysosome marker LAMP-1. Increased phagolysosomal fusion detected during CREB inhibition was dependent on RIPK3-induced pMLKL, indicating that M.tb-induced CREB signaling limits phagolysosomal fusion through inhibition of the necroptotic signaling pathway. Altogether, our data show that M.tb induces CREB activation in human macrophages early post-infection to create an environment conducive to bacterial growth. Targeting certain aspects of the CREB-induced signaling pathway may represent an innovative approach for development of host-directed therapeutics to combat TB.

Hydrazone-Tethered 5-(Pyridin-4-yl)-4H-1,2,4-triazole-3-thiol Hybrids: Synthesis, Characterisation, in silico ADME Studies, and in†vitro Antimycobacterial Evaluation and Cytotoxicity.

Compounds containing arylpyrrole-, 1,2,4-triazole- and hydrazone structural frameworks have been widely studied and demonstrated to exhibit a wide range of pharmacological properties. Herein, an exploratory series of new 1,2,4-triazole derivatives designed by amalgamation of arylpyrrole and 1,2,4-triazole structural units via a hydrazone linkage is reported. The synthesised compounds were tested in vitro for their potential activity against Mycobacterium tuberculosis (MTB) H37 Rv strain. The most promising compound 13 - the derivative without the benzene ring appended to the pyrrole unit displayed acceptable activity (MIC90 =3.99 µM) against MTB H37 Rv, while other compounds from the series exhibited modest to weak antimycobacterial activity with MIC90 values in the range between 7.0 and >125 µM. Furthermore, in silico results, predicated using the SwissADME web tool, show that the prepared compounds display desirable ADME profile with parameters within acceptable range.

Comparison of two diagnostic intervention packages for community-based active case finding for tuberculosis: an open-label randomized controlled trial.

Two in every five patients with active tuberculosis (TB) remain undiagnosed or unreported. Therefore community-based, active case-finding strategies require urgent implementation. However, whether point-of-care (POC), portable battery-operated, molecular diagnostic tools deployed at a community level, compared with conventionally used POC smear microscopy, can shorten time-to-treatment initiation, thus potentially curtailing transmission, remains unclear. To clarify this issue, we performed an open-label, randomized controlled trial in periurban informal settlements of Cape Town, South Africa, where we TB symptom screened 5,274 individuals using a community-based scalable mobile clinic. Some 584 individuals with HIV infection or symptoms of TB underwent targeted diagnostic screening and were randomized (1:1) to same-day smear microscopy (n = 296) or on-site DNA-based molecular diagnosis (n = 288; GeneXpert). The primary aim was to compare time to TB treatment initiation between the arms. Secondary aims included feasibility and detection of probably infectious people. Of participants who underwent targeted screening, 9.9% (58 of 584) had culture-confirmed TB. Time-to-treatment initiation occurred significantly earlier in the Xpert versus the smear-microscopy arm (8 versus 41 d, P = 0.002). However, overall, Xpert detected only 52% of individuals with culture-positive TB. Notably, Xpert detected almost all of the probably infectious patients compared with smear microscopy (94.1% versus 23.5%, P = <0.001). Xpert was associated with a shorter median time to treatment of probably infectious patients (7 versus 24 d, P = 0.02) and a greater proportion of infectious patients were on treatment at 60 d compared with the probably noninfectious patients (76.5% versus 38.2%, P < 0.01). Overall, a greater proportion of POC Xpert-positive participants were on treatment at 60 d compared with all culture-positive participants (100% versus 46.5%, P < 0.01). These findings challenge the traditional paradigm of a passive case-finding, public health strategy and argues for the implementation of portable DNA-based diagnosis with linkage to care as a community-oriented, transmission-interruption strategy. The study was registered with the South African National Clinical Trials Registry (application ID 4367; DOH-27-0317-5367) and ClinicalTrials.gov (NCT03168945).

Current therapeutic delivery approaches using nanocarriers for the treatment of tuberculosis disease.

Tuberculosis is a major health issue globally and a leading cause of death due to the infective microorganism Mycobacterium tuberculosis. Treatment of drug resistance tuberculosis requires longer treatment with multiple daily doses of drugs. Unfortunately, these drugs are often associated with poor patient compliance. In this situation, a need has been felt for the less toxic, shorter, and more effective treatment of the infected tuberculosis patients. Current research to develop novel anti-tubercular drugs shows hope for better management of the disease. Research on drug targeting and precise delivery of the old anti-tubercular drugs with the help of nanotechnology is promising for effective treatment. This review has discussed the status currently available treatments for tuberculosis patients infected with Mycobacterium alone or in comorbid conditions like diabetes, HIV and cancer. This review also highlighted the challenges in the current treatment and research on the novel anti-tubercular drugs to prevent multi-drug-resistant tuberculosis. It presents the research highlights on the targeted delivery of anti-tubercular drugs using different nanocarriers for preventing multi-drug resistant tuberculosis. Report has shown the importance and development of the research on nanocarriers mediated anti-tubercular delivery of the drugs to overcome the current challenges in tuberculosis treatment.

Synthesis, antitubercular evaluation, and molecular docking studies of hybrid pyridinium salts derived from isoniazid.

In the quest to develop potent inhibitors for Mycobacterium tuberculosis, novel isoniazid-based pyridinium salts were designed, synthesized, and tested for their antimycobacterial activities against the H37 Rv strain of Mycobacterium tuberculosis using rifampicin as a standard. The pyridinium salts 4k, 4l, and 7d showed exceptional antimycobacterial activities with MIC90 at 1 µg/mL. The in vitro cytotoxicity and pharmacokinetics profiles of these compounds were established for the identification of a lead molecule using in vivo efficacy proof-of-concept studies and found that the lead compound 4k possesses LC50 value at 25 µg/mL. The in vitro antimycobacterial activity results were further supported by in silico studies with good binding affinities ranging from -9.8 to -11.6 kcal/mol for 4k, 4l, and 7d with the target oxidoreductase DprE1 enzyme. These results demonstrate that pyridinium salts derived from isoniazid can be a potentially promising pharmacophore for the development of novel antitubercular candidates.

Applications of machine learning tools for ultra-sensitive detection of lipoarabinomannan with plasmonic grating biosensors in clinical samples of tuberculosis.

BACKGROUND: Tuberculosis is one of the top ten causes of death globally and the leading cause of death from a single infectious agent. Eradicating the Tuberculosis epidemic by 2030 is one of the top United Nations Sustainable Development Goals. Early diagnosis is essential to achieving this goal because it improves individual prognosis and reduces transmission rates of asymptomatic infected. We aim to support this goal by developing rapid and sensitive diagnostics using machine learning algorithms to minimize the need for expert intervention. METHODS AND FINDINGS: A single molecule fluorescence immunosorbent assay was used to detect Tuberculosis biomarker lipoarabinomannan from a set of twenty clinical patient samples and a control set of spiked human urine. Tuberculosis status was separately confirmed by GeneXpert MTB/RIF and cell culture. Two machine learning algorithms, an automatic and a semiautomatic model, were developed and trained by the calibrated lipoarabinomannan titration assay data and then tested against the ground truth patient data. The semiautomatic model differed from the automatic model by an expert review step in the former, which calibrated the lower threshold to determine single molecules from background noise. The semiautomatic model was found to provide 88.89% clinical sensitivity, while the automatic model resulted in 77.78% clinical sensitivity. CONCLUSIONS: The semiautomatic model outperformed the automatic model in clinical sensitivity as a result of the expert intervention applied during calibration and both models vastly outperformed manual expert counting in terms of time-to-detection and completion of analysis. Meanwhile, the clinical sensitivity of the automatic model could be improved significantly with a larger training dataset. In short, semiautomatic, and automatic Gaussian Mixture Models have a place in supporting rapid detection of Tuberculosis in resource-limited settings without sacrificing clinical sensitivity.

Characterization of the immune impairment of patients with tuberculosis and COVID-19 coinfection.

OBJECTIVES: To characterize the plasma immune profile of patients with tuberculosis (TB)-COVID-19 compared with COVID-19, TB, or healthy controls and to evaluate in vitro the specific responses to SARS-CoV-2 and Mycobacterium tuberculosis (Mtb)-antigens. METHODS: We enrolled 119 subjects: 14 TB-COVID-19, 47 COVID-19, 38 TB, and 20 controls. The plasmatic levels of 27 immune factors were measured at baseline using a multiplex assay. The specific response to SARS-CoV-2 and Mtb antigens was evaluated using a home-made whole blood platform and QuantiFERON-Plus tubes, respectively. RESULTS: We found an immune signature (tumor necrosis factor [TNF]-α, macrophage inflammatory protein-1ß, and interleukin [IL]-9) associated with TB-COVID-19 coinfection compared with COVID-19 (P <0.05), and TNF-α showed the highest discriminant power. We also found another signature (TNF-α, IL-1ß, IL-17A, IL-5, fibroblast growth factor-basic, and granulocyte macrophage colony-stimulating factor [GM-CSF]) in coinfected patients compared with patients with TB (P <0.05), and among them, TNF-α and granulocyte macrophage colony-stimulating factor showed a non-negligible discriminating ability. Moreover, coinfected patients showed a significantly reduced SARS-CoV-2-specific response compared with COVID-19 for several pro-inflammatory cytokines/chemokines, anti-inflammatory cytokines, and growth factors (P ≤0.05). Furthermore, coinfection negatively affected the Mtb-specific response (P ≤0.05). CONCLUSION: We found immune signatures associated with TB-COVID-19 coinfection and observed a major impairment of SARS-CoV-2-specific and, to a lesser extent, the Mtb-specific immune responses. These findings further advance our knowledge of the immunopathology of TB-COVID-19 coinfection.

Evolution of tuberculosis diagnostics: From molecular strategies to nanodiagnostics.

Tuberculosis has remained a global concern for public health affecting the lives of people for ages. Approximately 10 million people are affected by the disease and 1.5 million succumb to the disease worldwide annually. The COVID-19 pandemic has highlighted the role of early diagnosis to win the battle against such infectious diseases. Thus, advancement in the diagnostic approaches to provide early detection forms the foundation to eradicate and manage contagious diseases like tuberculosis. The conventional diagnostic strategies include microscopic examination, chest X-ray and tuberculin skin test. The limitations associated with sensitivity and specificity of these tests demands for exploring new techniques like probe-based assays, CRISPR-Cas and microRNA detection. The aim of the current review is to envisage the correlation between both the conventional and the newer approaches to enhance the specificity and sensitivity. A significant emphasis has been placed upon nanodiagnostic approaches manipulating quantum dots, magnetic nanoparticles, and biosensors for accurate diagnosis of latent, active and drug-resistant TB. Additionally, we would like to ponder upon a reliable method that is cost-effective, reproducible, require minimal infrastructure and provide point-of-care to the patients.