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1.
Front Immunol ; 12: 687044, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34630380

RESUMO

Phagosome-lysosome fusion in innate immune cells like macrophages and neutrophils marshal an essential role in eliminating intracellular microorganisms. In microbe-challenged macrophages, phagosome-lysosome fusion occurs 4 to 6 h after the phagocytic uptake of the microbe. However, live pathogenic mycobacteria hinder the transfer of phagosomes to lysosomes, up to 20 h post-phagocytic uptake. This period is required to evade pro-inflammatory response and upregulate the acid-stress tolerant proteins. The exact sequence of events through which mycobacteria retards phagolysosome formation remains an enigma. The macrophage coat protein Coronin1(Cor1) is recruited and retained by mycobacteria on the phagosome membrane to retard its maturation by hindering the access of phagosome maturation factors. Mycobacteria-infected macrophages exhibit an increased cAMP level, and based on receptor stimulus, Cor1 expressing cells show a higher level of cAMP than non-Cor1 expressing cells. Here we have shown that infection of bone marrow-derived macrophages with H37Rv causes a Cor1 dependent rise of intracellular cAMP levels at the vicinity of the phagosomes. This increased cAMP fuels cytoskeletal protein Cofilin1 to depolymerize F-actin around the mycobacteria-containing phagosome. Owing to reduced F-actin levels, the movement of the phagosome toward the lysosomes is hindered, thus contributing to the retarded phagosome maturation process. Additionally, Cor1 mediated upregulation of Cofilin1 also contributes to the prevention of phagosomal acidification, which further aids in the retardation of phagosome maturation. Overall, our study provides first-hand information on Cor1 mediated retardation of phagosome maturation, which can be utilized in developing novel peptidomimetics as part of host-directed therapeutics against tuberculosis.


Assuntos
Cofilina 1/metabolismo , AMP Cíclico/metabolismo , Macrófagos/microbiologia , Proteínas dos Microfilamentos/metabolismo , Infecções por Mycobacterium não Tuberculosas/microbiologia , Mycobacterium bovis/patogenicidade , Mycobacterium smegmatis/patogenicidade , Mycobacterium tuberculosis/patogenicidade , Fagossomos/microbiologia , Tuberculose/microbiologia , Animais , Linhagem Celular , Interações Hospedeiro-Patógeno , Concentração de Íons de Hidrogênio , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Proteínas dos Microfilamentos/genética , Infecções por Mycobacterium não Tuberculosas/imunologia , Infecções por Mycobacterium não Tuberculosas/metabolismo , Mycobacterium bovis/imunologia , Mycobacterium smegmatis/imunologia , Mycobacterium tuberculosis/imunologia , Fagossomos/imunologia , Fagossomos/metabolismo , Sistemas do Segundo Mensageiro , Tuberculose/imunologia , Tuberculose/metabolismo
2.
Nat Commun ; 12(1): 6099, 2021 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-34671035

RESUMO

Mycobacterium tuberculosis is a clonal pathogen proposed to have co-evolved with its human host for millennia, yet our understanding of its genomic diversity and biogeography remains incomplete. Here we use a combination of phylogenetics and dimensionality reduction to reevaluate the population structure of M. tuberculosis, providing an in-depth analysis of the ancient Indo-Oceanic Lineage 1 and the modern Central Asian Lineage 3, and expanding our understanding of Lineages 2 and 4. We assess sub-lineages using genomic sequences from 4939 pan-susceptible strains, and find 30 new genetically distinct clades that we validate in a dataset of 4645 independent isolates. We find a consistent geographically restricted or unrestricted pattern for 20 groups, including three groups of Lineage 1. The distribution of terminal branch lengths across the M. tuberculosis phylogeny supports the hypothesis of a higher transmissibility of Lineages 2 and 4, in comparison with Lineages 3 and 1, on a global scale. We define an expanded barcode of 95 single nucleotide substitutions that allows rapid identification of 69 M. tuberculosis sub-lineages and 26 additional internal groups. Our results paint a higher resolution picture of the M. tuberculosis phylogeny and biogeography.


Assuntos
Mycobacterium tuberculosis/classificação , Filogenia , Tuberculose/transmissão , Código de Barras de DNA Taxonômico , Evolução Molecular , Genoma Bacteriano/genética , Humanos , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/isolamento & purificação , Filogeografia , Polimorfismo de Nucleotídeo Único , Software , Tuberculose/microbiologia
3.
PLoS One ; 16(10): e0258774, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34699523

RESUMO

Next-Generation Sequencing (NGS) is widely used to investigate genomic variation. In several studies, the genetic variation of Mycobacterium tuberculosis has been analyzed in sputum samples without previous culture, using target enrichment methodologies for NGS. Alignments obtained by different programs generally map the sequences under default parameters, and from these results, it is assumed that only Mycobacterium reads will be obtained. However, variants of interest microorganism in clinical samples can be confused with a vast collection of reads from other bacteria, viruses, and human DNA. Currently, there are no standardized pipelines, and the cleaning success is never verified since there is a lack of rigorous controls to identify and remove reads from other sputum-microorganisms genetically similar to M. tuberculosis. Therefore, we designed a bioinformatic pipeline to process NGS data from sputum samples, including several filters and quality control points to identify and eliminate non-M. tuberculosis reads to obtain a reliable genetic variant report. Our proposal uses the SURPI software as a taxonomic classifier to filter input sequences and perform a mapping that provides the highest percentage of Mycobacterium reads, minimizing the reads from other microorganisms. We then use the filtered sequences to perform variant calling with the GATK software, ensuring the mapping quality, realignment, recalibration, hard-filtering, and post-filter to increase the reliability of the reported variants. Using default mapping parameters, we identified reads of contaminant bacteria, such as Streptococcus, Rhotia, Actinomyces, and Veillonella. Our final mapping strategy allowed a sequence identity of 97.8% between the input reads and the whole M. tuberculosis reference genome H37Rv using a genomic edit distance of three, thus removing 98.8% of the off-target sequences with a Mycobacterium reads loss of 1.7%. Finally, more than 200 unreliable genetic variants were removed during the variant calling, increasing the report's reliability.


Assuntos
Biologia Computacional/métodos , DNA Bacteriano/genética , Mycobacterium tuberculosis/genética , Tuberculose/microbiologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Mycobacterium tuberculosis/classificação , Mycobacterium tuberculosis/isolamento & purificação , Análise de Sequência de DNA , Software , Escarro/microbiologia
4.
PLoS One ; 16(10): e0257920, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34618810

RESUMO

Tuberculosis (TB), a contagious disease mainly caused by Mycobacterium tuberculosis (M. tb), Mycobacterium bovis (M. bovis), and Mycobacterium caprae (M. caprae), poses a major global threat to the health of humans and many species of animals. Developing an ante-mortem detection technique for different species would be of significance in improving the surveillance employing a One Health strategy. To achieve this goal, a universal indirect ELISA was established for serologically detecting Mycobacterium tuberculosis complex infection for multiple live hosts by using a fusion protein of MPB70, MPB83, ESAT6, and CFP10 common in M. tb, M. bovis, and M. caprae as the coating antigen (MMEC) and HRP-labeled fusion protein A and G as a secondary antibody. After testing the known positive and negative sera, the receiver operating characteristic curves were constructed to decide the cut-off values. Then, the diagnostic sensitivity and specificity of MMEC/AG-iELISA were determined as 100.00% (95% CI: 96.90%, 100.00%) and 100.00% (95% CI: 98.44%, 100.00%) for M. bovis infection of cattle, 100.00% (95% CI: 95.00%, 100.00%) and 100.0% (95% CI: 96.80%, 100.00%) for M. bovis infection of sheep, 90.74% (95% CI: 80.09%, 95.98%) and 98.63% (95% CI: 95.14%, 99.76%) for M. bovis infection of cervids, 100.00% (95% CI: 15.81%, 100.00%) and 98.81% (95% CI: 93.54%, 99.97%) for M. bovis infection of monkeys, 100.00% (95% CI: 86.82%, 100.00%) and 94.85% (95% CI: 91.22%, 97.03%) for M. tb infection of humans. Furthermore, this MMEC/AG-iELISA likely detects M. caprae infection in roe deer. Thus this method has a promising application in serological TB surveillance for multiple animal species thereby providing evidence for taking further action in TB control.


Assuntos
Ensaio de Imunoadsorção Enzimática , Mycobacterium tuberculosis/isolamento & purificação , Testes Sorológicos , Tuberculose/diagnóstico , Animais , Animais Selvagens/microbiologia , Anticorpos Antibacterianos/química , Anticorpos Antibacterianos/imunologia , Bovinos , Cervos/microbiologia , Testes Diagnósticos de Rotina , Humanos , Mycobacterium bovis/isolamento & purificação , Mycobacterium bovis/patogenicidade , Mycobacterium tuberculosis/genética , Ovinos/microbiologia , Tuberculose/microbiologia
5.
Medicine (Baltimore) ; 100(35): e27125, 2021 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-34477155

RESUMO

ABSTRACT: We aimed to investigate the genetic and demographic differences and interactions between areas where observed genomic variations in Mycobacterium tuberculosis (M. tb) were distributed uniformly in cold and hot spots.The cold and hot spot areas were identified using the reported incidence of TB over the previous 5 years. Whole genome sequencing was performed on 291 M. tb isolates between January and June 2018. Analysis of molecular variance and a multifactor dimensionality reduction (MDR) model was applied to test gene-gene-environment interactions. Adjusted odds ratios (OR) and 95% confidence intervals (CI) were computed to test the extent to which genetic mutation affects the TB epidemic using a multivariate logistic regression model.The percentage of the Beijing family strain in hot spots was significantly higher than that in cold spots (64.63% vs 50.69%, P = .022), among the elderly, people with a low BMI, and those having a history of contact with a TB patient (all P < .05). Individuals from cold spot areas had a higher frequency of out-of-town traveling (P < .05). The mutation of Rv1186c, Rv3900c, Rv1508c, Rv0210, and an Intergenic Region (SNP site: 3847237) showed a significant difference between cold and hot spots. (P < .001). The MDR model displayed a clear negative interaction effect of age groups with BMI (interaction entropy: -3.55%) and mutation of Rv0210 (interaction entropy: -2.39%). Through the mutations of Rv0210 and BMI had a low independent effect (interaction entropy: -1.46%).Our data suggests a statistically significant role of age, BMI and the polymorphisms of Rv0210 genes in the transmission and development of M. tb. The results provide clues for the study of susceptibility genes of M. tb in different populations. The characteristic strains showed a local epidemic. Strengthening genotype monitoring of strains in various regions can be used as an early warning signal of epidemic spillover.


Assuntos
Interações Hospedeiro-Patógeno/genética , Mycobacterium tuberculosis/genética , Tuberculose/microbiologia , Adulto , China/epidemiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Tuberculose/epidemiologia
6.
Front Cell Infect Microbiol ; 11: 708827, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34589441

RESUMO

Understanding the dynamics of lung microbiota in tuberculosis patients, especially those who cannot be confirmed bacteriologically in clinical practice, is imperative for accurate diagnosis and effective treatment. This study aims to characterize the distinct lung microbial features between bacteriologically confirmed and negative tuberculosis patients to understand the influence of microbiota on tuberculosis patients. We collected specimens of bronchoalveolar lavage fluid from 123 tuberculosis patients. Samples were subjected to metagenomic next-generation sequencing to reveal the lung microbial signatures. By combining conventional bacterial detection and metagenomic sequencing, 101/123 (82%) tuberculosis patients were bacteriologically confirmed. In addition to Mycobacterium tuberculosis, Staphylococcus aureus, Kluyveromyces lactis, and Pyricularia pennisetigena were also enriched in the bacteriological confirmation group. In contrast, Haemophilus parainfluenzae was enriched in the bacteriologically negative group. Besides, microbial interaction exhibits a different state between bacteriologically confirmed and negative tuberculosis patients. Mycobacterium tuberculosis was confirmed correlated with clinical characteristics such as albumin and chest cavities. Our study comprehensively demonstrates the correlation between unique features of lung microbial dynamics and the clinical characteristics of tuberculosis patients, suggesting the importance of studying the pulmonary microbiome in tuberculosis disease and providing new insights for future precision diagnosis and treatment.


Assuntos
Microbiota , Mycobacterium tuberculosis , Tuberculose , Ascomicetos , Líquido da Lavagem Broncoalveolar , Haemophilus parainfluenzae , Humanos , Kluyveromyces , Pulmão , Metagenômica , Mycobacterium tuberculosis/genética , Staphylococcus aureus , Tuberculose/microbiologia
7.
Sci Rep ; 11(1): 17423, 2021 08 31.
Artigo em Inglês | MEDLINE | ID: mdl-34465797

RESUMO

We aimed to elucidate the differences in genomic methylation patterns between ADLI and non-ADLI patients to identify DNA methylation-based biomarkers. Genome-wide DNA methylation patterns were obtained using Infinium MethylationEPIC (EPIC) BeadChip array to analyze 14 peripheral blood samples (7 ADLI cases, 7 non-ADLI controls). Changes in the mRNA and DNA methylation in the target genes of another 120 peripheral blood samples (60 ADLI cases, 60 non-ADLI controls) were analyzed by real-time polymerase chain reaction and pyrosequencing, respectively. A total of 308 hypermethylated CpG sites and 498 hypomethylated CpG sites were identified. Significantly, hypermethylated CpG sites cg06961147 and cg24666046 in TANC1 associated with ADLI was identified by genome-wide DNA methylation profiling. The mRNA expression of TANC1 was lower in the cases compared to the controls. Pyrosequencing validated these two differentially methylated loci, which was consistent with the results from the EPIC BeadChip array. Receiver operating characteristic analysis indicated that the area under the curve of TANC1 (cg06961147, cg24666046, and their combinations) was 0.812, 0.842, and 0.857, respectively. These results indicate that patients with ADLI have different genomic methylation patterns than patients without ADLI. The hypermethylated differentially methylated site cg06961147 combined with cg24666046 in TANC1 provides evidence for the diagnosis of ADLI.


Assuntos
Antituberculosos/efeitos adversos , Biomarcadores/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/diagnóstico , Metilação de DNA , Proteínas de Membrana/genética , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose/tratamento farmacológico , Adulto , Estudos de Casos e Controles , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Ilhas de CpG , Epigênese Genética , Feminino , Humanos , Masculino , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Regiões Promotoras Genéticas , Tuberculose/microbiologia
8.
PLoS One ; 16(9): e0256664, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34529706

RESUMO

The DNA polymorphisms found in clinical strains of Mycobacterium tuberculosis drive altered physiology, virulence, and pathogenesis in them. Although the lineages of these clinical strains can be traced back to common ancestor/s, there exists a plethora of difference between them, compared to those that have evolved in the laboratory. We identify a mutation present in ~80% of clinical strains, which maps in the HATPase domain of the sensor kinase MtrB and alters kinase and phosphatase activities, and affects its physiological role. The changes conferred by the mutation were probed by in-vitro biochemical assays which revealed changes in signaling properties of the sensor kinase. These changes also affect bacterial cell division rates, size and membrane properties. The study highlights the impact of DNA polymorphisms on the pathophysiology of clinical strains and provides insights into underlying mechanisms that drive signal transduction in pathogenic bacteria.


Assuntos
Proteínas de Bactérias/genética , Mycobacterium tuberculosis , Proteínas de Ligação a RNA/genética , Fatores de Transcrição/genética , Tuberculose/microbiologia , Regulação Bacteriana da Expressão Gênica , Humanos , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/patogenicidade , Polimorfismo de Nucleotídeo Único , Transdução de Sinais , Virulência
9.
Cells ; 10(9)2021 09 19.
Artigo em Inglês | MEDLINE | ID: mdl-34572127

RESUMO

It has been known that infection plays a role in the development of hypertension. However, the role of hypertension in the progression of infectious diseases remain unknown. Many countries with high rates of hypertension show geographical overlaps with those showing high incidence rates of tuberculosis (TB). To explore the role of hypertension in tuberculosis, we compared the effects of hypertension during mycobacterial infection, we infected both hypertensive Angiotensin II (Ang II) and control mice with Mycobacterium tuberculosis (Mtb) strain H37Ra by intratracheal injection. Ang II-induced hypertension promotes cell death through both apoptosis and necrosis in Mtb H37Ra infected mouse lungs. Interestingly, we found that lipid accumulation in pulmonary tissues was significantly increased in the hypertension group compared to the normal controls. Ang II-induced hypertension increases the formation of foamy macrophages during Mtb infection and it leads to cell death. Moreover, the hypertension group showed more severe granuloma formation and fibrotic lesions in comparison with the control group. Finally, we observed that the total number of mycobacteria was increased in the lungs in the hypertension group compared to the normal controls. Taken together, these results suggest that hypertension increases intracellular survival of Mtb through formation of foamy macrophages, resulting in severe pathogenesis of TB.


Assuntos
Angiotensina II/toxicidade , Apoptose , Hipertensão/patologia , Pulmão/patologia , Macrófagos/patologia , Mycobacterium tuberculosis/fisiologia , Tuberculose/patologia , Animais , Hipertensão/induzido quimicamente , Pulmão/microbiologia , Macrófagos/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Necrose , Tuberculose/microbiologia , Vasoconstritores/toxicidade
10.
PLoS Med ; 18(9): e1003739, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34491987

RESUMO

BACKGROUND: Among people living with HIV (PLHIV), more flexible and sensitive tuberculosis (TB) screening tools capable of detecting both symptomatic and subclinical active TB are needed to (1) reduce morbidity and mortality from undiagnosed TB; (2) facilitate scale-up of tuberculosis preventive therapy (TPT) while reducing inappropriate prescription of TPT to PLHIV with subclinical active TB; and (3) allow for differentiated HIV-TB care. METHODS AND FINDINGS: We used Botswana XPRES trial data for adult HIV clinic enrollees collected during 2012 to 2015 to develop a parsimonious multivariable prognostic model for active prevalent TB using both logistic regression and random forest machine learning approaches. A clinical score was derived by rescaling final model coefficients. The clinical score was developed using southern Botswana XPRES data and its accuracy validated internally, using northern Botswana data, and externally using 3 diverse cohorts of antiretroviral therapy (ART)-naive and ART-experienced PLHIV enrolled in XPHACTOR, TB Fast Track (TBFT), and Gugulethu studies from South Africa (SA). Predictive accuracy of the clinical score was compared with the World Health Organization (WHO) 4-symptom TB screen. Among 5,418 XPRES enrollees, 2,771 were included in the derivation dataset; 67% were female, median age was 34 years, median CD4 was 240 cells/µL, 189 (7%) had undiagnosed prevalent TB, and characteristics were similar between internal derivation and validation datasets. Among XPHACTOR, TBFT, and Gugulethu cohorts, median CD4 was 400, 73, and 167 cells/µL, and prevalence of TB was 5%, 10%, and 18%, respectively. Factors predictive of TB in the derivation dataset and selected for the clinical score included male sex (1 point), ≥1 WHO TB symptom (7 points), smoking history (1 point), temperature >37.5°C (6 points), body mass index (BMI) <18.5kg/m2 (2 points), and severe anemia (hemoglobin <8g/dL) (3 points). Sensitivity using WHO 4-symptom TB screen was 73%, 80%, 94%, and 94% in XPRES, XPHACTOR, TBFT, and Gugulethu cohorts, respectively, but increased to 88%, 87%, 97%, and 97%, when a clinical score of ≥2 was used. Negative predictive value (NPV) also increased 1%, 0.3%, 1.6%, and 1.7% in XPRES, XPHACTOR, TBFT, and Gugulethu cohorts, respectively, when the clinical score of ≥2 replaced WHO 4-symptom TB screen. Categorizing risk scores into low (<2), moderate (2 to 10), and high-risk categories (>10) yielded TB prevalence of 1%, 1%, 2%, and 6% in the lowest risk group and 33%, 22%, 26%, and 32% in the highest risk group for XPRES, XPHACTOR, TBFT, and Gugulethu cohorts, respectively. At clinical score ≥2, the number needed to screen (NNS) ranged from 5.0 in Gugulethu to 11.0 in XPHACTOR. Limitations include that the risk score has not been validated in resource-rich settings and needs further evaluation and validation in contemporary cohorts in Africa and other resource-constrained settings. CONCLUSIONS: The simple and feasible clinical score allowed for prioritization of sensitivity and NPV, which could facilitate reductions in mortality from undiagnosed TB and safer administration of TPT during proposed global scale-up efforts. Differentiation of risk by clinical score cutoff allows flexibility in designing differentiated HIV-TB care to maximize impact of available resources.


Assuntos
Antirretrovirais/uso terapêutico , Antituberculosos/uso terapêutico , Coinfecção , Infecções por HIV/tratamento farmacológico , Sobreviventes de Longo Prazo ao HIV , Programas de Rastreamento , Serviços Preventivos de Saúde , Tuberculose/prevenção & controle , Adulto , Antirretrovirais/efeitos adversos , Antituberculosos/efeitos adversos , Botsuana/epidemiologia , Ensaios Clínicos como Assunto , Diagnóstico Precoce , Feminino , Infecções por HIV/diagnóstico , Infecções por HIV/epidemiologia , Infecções por HIV/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Prevalência , Prognóstico , Reprodutibilidade dos Testes , Medição de Risco , Fatores de Risco , Tuberculose/diagnóstico , Tuberculose/epidemiologia , Tuberculose/microbiologia
11.
PLoS Med ; 18(9): e1003752, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34499665

RESUMO

BACKGROUND: Suboptimal tuberculosis (TB) diagnostics and HIV contribute to the high global burden of TB. We investigated costs and yield from systematic HIV-TB screening, including computer-aided digital chest X-ray (DCXR-CAD). METHODS AND FINDINGS: In this open, three-arm randomised trial, adults (≥18 years) with cough attending acute primary services in Malawi were randomised (1:1:1) to standard of care (SOC); oral HIV testing (HIV screening) and linkage to care; or HIV testing and linkage to care plus DCXR-CAD with sputum Xpert for high CAD4TBv5 scores (HIV-TB screening). Participants and study staff were not blinded to intervention allocation, but investigator blinding was maintained until final analysis. The primary outcome was time to TB treatment. Secondary outcomes included proportion with same-day TB treatment; prevalence of undiagnosed/untreated bacteriologically confirmed TB on day 56; and undiagnosed/untreated HIV. Analysis was done on an intention-to-treat basis. Cost-effectiveness analysis used a health-provider perspective. Between 15 November 2018 and 27 November 2019, 8,236 were screened for eligibility, with 473, 492, and 497 randomly allocated to SOC, HIV, and HIV-TB screening arms; 53 (11%), 52 (9%), and 47 (9%) were lost to follow-up, respectively. At 56 days, TB treatment had been started in 5 (1.1%) SOC, 8 (1.6%) HIV screening, and 15 (3.0%) HIV-TB screening participants. Median (IQR) time to TB treatment was 11 (6.5 to 38), 6 (1 to 22), and 1 (0 to 3) days (hazard ratio for HIV-TB versus SOC: 2.86, 1.04 to 7.87), with same-day treatment of 0/5 (0%) SOC, 1/8 (12.5%) HIV, and 6/15 (40.0%) HIV-TB screening arm TB patients (p = 0.03). At day 56, 2 SOC (0.5%), 4 HIV (1.0%), and 2 HIV-TB (0.5%) participants had undiagnosed microbiologically confirmed TB. HIV screening reduced the proportion with undiagnosed or untreated HIV from 10 (2.7%) in the SOC arm to 2 (0.5%) in the HIV screening arm (risk ratio [RR]: 0.18, 0.04 to 0.83), and 1 (0.2%) in the HIV-TB screening arm (RR: 0.09, 0.01 to 0.71). Incremental costs were US$3.58 and US$19.92 per participant screened for HIV and HIV-TB; the probability of cost-effectiveness at a US$1,200/quality-adjusted life year (QALY) threshold was 83.9% and 0%. Main limitations were the lower than anticipated prevalence of TB and short participant follow-up period; cost and quality of life benefits of this screening approach may accrue over a longer time horizon. CONCLUSIONS: DCXR-CAD with universal HIV screening significantly increased the timeliness and completeness of HIV and TB diagnosis. If implemented at scale, this has potential to rapidly and efficiently improve TB and HIV diagnosis and treatment. TRIAL REGISTRATION: clinicaltrials.gov NCT03519425.


Assuntos
Coinfecção , Tosse/diagnóstico , Diagnóstico por Computador , Infecções por HIV/diagnóstico , Teste de HIV , Radiografia Torácica , Tuberculose/diagnóstico por imagem , Adulto , Fármacos Anti-HIV/uso terapêutico , Antituberculosos/uso terapêutico , Análise Custo-Benefício , Tosse/microbiologia , Diagnóstico por Computador/economia , Feminino , Infecções por HIV/tratamento farmacológico , Infecções por HIV/epidemiologia , Teste de HIV/economia , Custos de Cuidados de Saúde , Acesso aos Serviços de Saúde , Humanos , Malaui/epidemiologia , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Prevalência , Atenção Primária à Saúde , Radiografia Torácica/economia , Tuberculose/tratamento farmacológico , Tuberculose/epidemiologia , Tuberculose/microbiologia , Adulto Jovem
12.
World J Microbiol Biotechnol ; 37(10): 175, 2021 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-34519879

RESUMO

The 38 kDa protein is a major antigen of mycobacterium tuberculosis and has been widely used in TB serodiagnosis, due to its highly sensitivity and specificity. Here we attempt to establish a production platform of recombinant 38 kDa protein in mammalian cells and to evaluate the potential value of 38 kDa protein in TB serodiagnosis. The 38 kDa gene is synthesized and cloned into a lentiviral expressing vector. Recombinant lentiviral vector LV-CMV-38 kDa-eGFP was packaged, titered, and then transduced into HEK 293 T cells. Recombinant cell lines were selected by limiting dilution. Supernatants were collected and purified by HisTrapTM HP column. Western blot showed a molecular weight of approximate 38 kDa in cell supernatants as expected. ELISA assay confirmed the immunological specificity of the obtained protein in the presence of MTB-infected human serum samples. In all, we have obtained a stable cell line with long-term and robust expression of secretory MTB 38 kDa protein, which may provide a promising candidate antigen for the development of TB serological diagnosis.


Assuntos
Antígenos de Bactérias/genética , Expressão Gênica , Lipoproteínas/genética , Mycobacterium tuberculosis/metabolismo , Antígenos de Bactérias/análise , Antígenos de Bactérias/biossíntese , Antígenos de Bactérias/isolamento & purificação , Clonagem Molecular , Ensaio de Imunoadsorção Enzimática , Células HEK293 , Humanos , Lipoproteínas/análise , Lipoproteínas/biossíntese , Lipoproteínas/isolamento & purificação , Mycobacterium tuberculosis/genética , Proteínas Recombinantes/análise , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Tuberculose/microbiologia
13.
PLoS One ; 16(9): e0256795, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34473752

RESUMO

Achieving targets set in the End TB Strategy is still a distant goal for many Low- and Middle-Income Countries (LMICs). The importance of strengthening public-private partnership by engaging all identified providers in Tuberculosis (TB) care has long been advocated in global TB policies and strategies. However, Informal Healthcare Providers (IPs) are not yet prioritised and engaged in National Tuberculosis Programs (NTPs) globally. There exists a substantial body of evidence that confirms an important contribution of IPs in TB care. A systematic understanding of their role is necessary to ascertain their potential in improving TB care in LMICs. The purpose of this review is to scope the role of IPs in TB care. The scoping review was guided by a framework developed by the Joanna Briggs Institute. An electronic search of literature was conducted in MEDLINE, EMBASE, SCOPUS, Global Health, CINAHL, and Web of Science. Of a total 5234 records identified and retrieved, 92 full-text articles were screened, of which 13 were included in the final review. An increasing trend was observed in publication over time, with most published between 2010-2019. In 60% of the articles, NTPs were mentioned as a collaborator in the study. For detection and diagnosis, IPs were primarily involved in identifying and referring patients. Administering DOT (Directly Observed Treatment) to the patient was the major task assigned to IPs for treatment and support. There is a paucity of evidence on prevention, as only one study involved IPs to perform this role. Traditional health providers were the most commonly featured, but there was not much variation in the role by provider type. All studies reported a positive role of IPs in improving TB care outcomes. This review demonstrates that IPs can be successfully engaged in various roles in TB care with appropriate support and training. Their contribution can support countries to achieve their national and global targets if prioritized in National TB Programs.


Assuntos
Atenção à Saúde/métodos , Países em Desenvolvimento , Pessoal de Saúde , Renda , Mycobacterium tuberculosis , Tuberculose/diagnóstico , Tuberculose/terapia , Saúde Global , Humanos , Avaliação de Resultados em Cuidados de Saúde , Pobreza , Tuberculose/microbiologia
14.
PLoS Med ; 18(9): e1003738, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34520459

RESUMO

BACKGROUND: Tuberculosis (TB) preventive therapy (TPT) is an essential component of care for people living with HIV (PLHIV). We compared efficacy, safety, completion, and drug-resistant TB risk for currently recommended TPT regimens through a systematic review and network meta-analysis (NMA) of randomized trials. METHODS AND FINDINGS: We searched MEDLINE, Embase, and the Cochrane Library from inception through June 9, 2020 for randomized controlled trials (RCTs) comparing 2 or more TPT regimens (or placebo/no treatment) in PLHIV. Two independent reviewers evaluated eligibility, extracted data, and assessed the risk of bias. We grouped TPT strategies as follows: placebo/no treatment, 6 to 12 months of isoniazid, 24 to 72 months of isoniazid, and rifamycin-containing regimens. A frequentist NMA (using graph theory) was carried out for the outcomes of development of TB disease, all-cause mortality, and grade 3 or worse hepatotoxicity. For other outcomes, graphical descriptions or traditional pairwise meta-analyses were carried out as appropriate. The potential role of confounding variables for TB disease and all-cause mortality was assessed through stratified analyses. A total of 6,466 unique studies were screened, and 157 full texts were assessed for eligibility. Of these, 20 studies (reporting 16 randomized trials) were included. The median sample size was 616 (interquartile range [IQR], 317 to 1,892). Eight were conducted in Africa, 3 in Europe, 3 in the Americas, and 2 included sites in multiple continents. According to the NMA, 6 to 12 months of isoniazid were no more efficacious in preventing microbiologically confirmed TB than rifamycin-containing regimens (incidence rate ratio [IRR] 1.0, 95% CI 0.8 to 1.4, p = 0.8); however, 6 to 12 months of isoniazid were associated with a higher incidence of all-cause mortality (IRR 1.6, 95% CI 1.2 to 2.0, p = 0.02) and a higher risk of grade 3 or higher hepatotoxicity (risk difference [RD] 8.9, 95% CI 2.8 to 14.9, p = 0.004). Finally, shorter regimens were associated with higher completion rates relative to longer regimens, and we did not find statistically significant differences in the risk of drug-resistant TB between regimens. Study limitations include potential confounding due to differences in posttreatment follow-up time and TB incidence in the study setting on the estimates of incidence of TB or all-cause mortality, as well as an underrepresentation of pregnant women and children. CONCLUSIONS: Rifamycin-containing regimens appear safer and at least as effective as isoniazid regimens in preventing TB and death and should be considered part of routine care in PLHIV. Knowledge gaps remain as to which specific rifamycin-containing regimen provides the optimal balance of efficacy, completion, and safety.


Assuntos
Antirretrovirais/uso terapêutico , Antituberculosos/uso terapêutico , Coinfecção , Infecções por HIV/tratamento farmacológico , Sobreviventes de Longo Prazo ao HIV , Serviços Preventivos de Saúde , Tuberculose/prevenção & controle , Adolescente , Adulto , Idoso , Antirretrovirais/efeitos adversos , Antituberculosos/efeitos adversos , Criança , Pré-Escolar , Feminino , Infecções por HIV/diagnóstico , Infecções por HIV/epidemiologia , Infecções por HIV/virologia , Humanos , Lactente , Isoniazida/uso terapêutico , Masculino , Pessoa de Meia-Idade , Rifamicinas/uso terapêutico , Medição de Risco , Fatores de Risco , Resultado do Tratamento , Tuberculose/diagnóstico , Tuberculose/epidemiologia , Tuberculose/microbiologia , Adulto Jovem
15.
Nat Commun ; 12(1): 4702, 2021 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-34349104

RESUMO

Mycobacterium tuberculosis can adapt to changing environments by non-heritable mechanisms. Frame-shifting insertions and deletions (indels) may also participate in adaptation through gene disruption, which could be reversed by secondary introduction of a frame-restoring indel. We present ScarTrek, a program that scans genomic data for indels, including those that together disrupt and restore a gene's reading frame, producing "frame-shift scars" suggestive of reversible gene inactivation. We use ScarTrek to analyze 5977 clinical M. tuberculosis isolates. We show that indel frequency inversely correlates with genomic linguistic complexity and varies with gene-position and gene-essentiality. Using ScarTrek, we detect 74 unique frame-shift scars in 48 genes, with a 3.74% population-level incidence of unique scar events. We find multiple scars in the ESX-1 gene cluster. Six scars show evidence of convergent evolution while the rest shared a common ancestor. Our results suggest that sequential indels are a mechanism for reversible gene silencing and adaptation in M. tuberculosis.


Assuntos
Adaptação Fisiológica/genética , Inativação Gênica , Mycobacterium tuberculosis/genética , Fases de Leitura/genética , Evolução Molecular , Genes Bacterianos/genética , Genoma Bacteriano/genética , Humanos , Mutação INDEL , Mycobacterium tuberculosis/isolamento & purificação , Tuberculose/microbiologia
16.
Int J Mol Sci ; 22(16)2021 Aug 04.
Artigo em Inglês | MEDLINE | ID: mdl-34445098

RESUMO

Granulysin is an antimicrobial peptide (AMP) expressed by human T-lymphocytes and natural killer cells. Despite a remarkably broad antimicrobial spectrum, its implementation into clinical practice has been hampered by its large size and off-target effects. To circumvent these limitations, we synthesized a 29 amino acid fragment within the putative cytolytic site of Granulysin (termed "Gran1"). We evaluated the antimicrobial activity of Gran1 against the major human pathogen Mycobacterium tuberculosis (Mtb) and a panel of clinically relevant non-tuberculous mycobacteria which are notoriously difficult to treat. Gran1 efficiently inhibited the mycobacterial proliferation in the low micro molar range. Super-resolution fluorescence microscopy and scanning electron microscopy indicated that Gran1 interacts with the surface of Mtb, causing lethal distortions of the cell wall. Importantly, Gran1 showed no off-target effects (cytokine release, chemotaxis, cell death) in primary human cells or zebrafish embryos (cytotoxicity, developmental toxicity, neurotoxicity, cardiotoxicity). Gran1 was selectively internalized by macrophages, the major host cell of Mtb, and restricted the proliferation of the pathogen. Our results demonstrate that the hypothesis-driven design of AMPs is a powerful approach for the identification of small bioactive compounds with specific antimicrobial activity. Gran1 is a promising component for the design of AMP-containing nanoparticles with selective activity and favorable pharmacokinetics to be pushed forward into experimental in vivo models of infectious diseases, most notably tuberculosis.


Assuntos
Antígenos de Diferenciação de Linfócitos T/imunologia , Macrófagos/imunologia , Mycobacterium tuberculosis/imunologia , Tuberculose/imunologia , Animais , Antígenos de Diferenciação de Linfócitos T/química , Células Cultivadas , Interações Hospedeiro-Patógeno , Humanos , Macrófagos/microbiologia , Mycobacterium tuberculosis/fisiologia , Peptídeos/química , Peptídeos/imunologia , Tuberculose/microbiologia , Peixe-Zebra
17.
Sci Rep ; 11(1): 16254, 2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34376753

RESUMO

The association between the total cholesterol level and tuberculosis (TB) risk has been controversial. Our study aimed to evaluate whether total cholesterol level can predict the risk of TB. Data from 5,000,566 subjects who participated in a health screening exam in 2009 were investigated using the Korean National Health Insurance Service database (2009-2018). Cox hazard regression analyses were used to evaluate TB risk according to the quartile of total cholesterol levels. During an average of 8.2 years of follow-up, 32,078 cases of TB occurred. There was a significant inverse association between the total cholesterol level and TB risk. Compared with subjects in the highest quartile, those in the lowest quartile had a 1.35-fold increased TB risk (95% confidence interval = 1.31-1.39). The association between total cholesterol level and TB risk was more apparent in young subjects (age < 65 years), those without diabetes mellitus (DM), and those without obesity (p for interaction < 0.001 for age group, DM, and body mass index). Although there was a significant inverse association between total cholesterol level and TB risk in subjects who did not use a statin, no significant association was observed between the total cholesterol level and TB risk in subjects who used a statin. A low total cholesterol level was significantly associated with an increased risk of TB, even after adjusting for confounders, especially in patients younger than 65 years, those without DM or obesity, and those who did not use a statin.


Assuntos
Colesterol/metabolismo , Bases de Dados Factuais , Tuberculose/epidemiologia , Idoso , Estudos de Casos e Controles , Humanos , Incidência , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Prognóstico , República da Coreia/epidemiologia , Fatores de Risco , Tuberculose/metabolismo , Tuberculose/microbiologia
18.
Biochem J ; 478(16): 3079-3098, 2021 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-34350952

RESUMO

DevR/DosR response regulator is believed to participate in virulence, dormancy adaptation and antibiotic tolerance mechanisms of Mycobacterium tuberculosis by regulating the expression of the dormancy regulon. We have previously shown that the interaction of DevR with RNA polymerase is essential for the expression of DevR-regulated genes. Here, we developed a M. tuberculosis-specific in vivo transcription system to enrich our understanding of DevR-RNA polymerase interaction. This in vivo assay involves co-transforming E. coli with two plasmids that express α, ß, ß' and σA subunits of M. tuberculosis RNA polymerase and a third plasmid that harbors a DevR expression cassette and a GFP reporter gene under the DevR-regulated fdxA promoter. We show that DevR-dependent transcription is sponsored exclusively by M. tuberculosis RNA polymerase and regulated by α and σA subunits of M. tuberculosis RNA polymerase. Using this E. coli triple plasmid system to express mutant variants of M. tuberculosis RNA polymerase, we identified E280 residue in C-terminal domain of α and K513 and R515 residues of σA to participate in DevR-dependent transcription. In silico modeling of a ternary complex of DevR, σA domain 4 and fdxA promoter suggest an interaction of Q505, R515 and K513 residues of σA with E178 and D172 residues of DevR and E471 of σA, respectively. These findings provide us with new insights into the interactions between DevR and RNA polymerase of M. tuberculosis which can be targeted for intercepting DevR function. Finally, we demonstrate the utility of this system for screening of anti-DevR compounds.


Assuntos
Proteínas de Bactérias/genética , Proteínas de Ligação a DNA/genética , Escherichia coli/genética , Regulação Bacteriana da Expressão Gênica , Mycobacterium tuberculosis/genética , Regiões Promotoras Genéticas/genética , Sequência de Aminoácidos , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Sequência de Bases , DNA/química , DNA/genética , DNA/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Modelos Moleculares , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/patogenicidade , Conformação de Ácido Nucleico , Plasmídeos/genética , Ligação Proteica , Domínios Proteicos , Homologia de Sequência de Aminoácidos , Tuberculose/microbiologia , Virulência/genética
19.
Tuberk Toraks ; 69(2): 160-166, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34256506

RESUMO

Introduction: This study aimed to evaluate the performance of the Xpert MTB/RIF assay in the identification of M. tuberculosis in pulmonary and extrapulmonary clinical samples by taking the results of the BACTEC MGIT 960TB culture system as a reference. Materials and Methods: A total of 11,341 specimens sent to Sivas Cumhuriyet University Application and Research Hospital Tuberculosis Laboratory for microbiological examination with suspicion of tuberculosis infection between January 2013 and December 2019 were examined, and 6847 clinical specimens that underwent culture (BACTEC MGIT 960TB), Xpert MTB/RIF and AFB (Acid-fast bacilli) testing were selected and included in our study. Of the samples included in the study, 5096 samples were pulmonary, and 1751 were extrapulmonary samples. Result: In our study, sensitivity, specificity, PPV and NPV values of Xpert MTB/ RIF and AFB were calculated by taking TB culture test as reference test. The sensitivity of the Xpert MTB/RIF assay was calculated as 96.1%, specificity as 99.7%, positive predictive value (PPV) as 88.2%, and negative predictive value (NPV) as 99.9%. These values for pulmonary samples were determined as 98.3%, 99.7%, 89.9%, and 99.9%, respectively. For extrapulmonary samples, the sensitivity of the assay was found as 89.4%, specificity as 99.5%, PPV as 82.9%, and NPV as 99.7%. The sensitivity and PPV values for AFBpositive samples were found to be 99.0% and 97.1%, respectively. For AFB negative samples, the sensitivity, specificity, PPV, and NPV values were determined as 90.5%, 99.7%, 73.8%, and 99.9%, respectively. Conclusions: A large number of clinical samples were studied with the Xpert MTB/RIF test in our study. It can be a guide in determining the performance of the test under the conditions of our country. Especially in the diagnosis of extrapulmonary TB, the effectiveness of the Xpert MTB/RIF assay has not been certainly proven in countries having a moderate prevalence of TB, such as Turkey. In most of the published studies, only a small part of the samples is extrapulmonary samples. So, our study provides valuable results in terms of evaluating a large number of extrapulmonary samples.


Assuntos
Técnicas de Diagnóstico Molecular/normas , Kit de Reagentes para Diagnóstico/normas , Tuberculose/diagnóstico , Humanos , Pulmão/microbiologia , Mycobacterium tuberculosis/isolamento & purificação , Valor Preditivo dos Testes , Sensibilidade e Especificidade , Escarro/microbiologia , Tuberculose/microbiologia , Turquia
20.
Int J Mol Sci ; 22(14)2021 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-34299161

RESUMO

Prostaglandin E2 (PGE2) is an important biological mediator involved in the defense against Mycobacterium tuberculosis (Mtb) infection. Currently, there are no reports on the mycobacterial components that regulate PGE2 production. Previously, we have reported that RpfE-treated dendritic cells (DCs) effectively expanded the Th1 and Th17 cell responses simultaneously; however, the mechanism underlying Th1 and Th17 cell differentiation is unclear. Here, we show that PGE2 produced by RpfE-activated DCs via the MAPK and cyclooxygenase 2 signaling pathways induces Th1 and Th17 cell responses mainly via the EP4 receptor. Furthermore, mice administered intranasally with PGE2 displayed RpfE-induced antigen-specific Th1 and Th17 responses with a significant reduction in bacterial load in the lungs. Furthermore, the addition of optimal PGE2 amount to IL-2-IL-6-IL-23p19-IL-1ß was essential for promoting differentiation into Th1/Th17 cells with strong bactericidal activity. These results suggest that RpfE-matured DCs produce PGE2 that induces Th1 and Th17 cell differentiation with potent anti-mycobacterial activity.


Assuntos
Proteínas de Bactérias/metabolismo , Diferenciação Celular , Células Dendríticas/metabolismo , Dinoprostona/metabolismo , Mycobacterium tuberculosis/fisiologia , Células Th1/citologia , Células Th17/citologia , Animais , Células Dendríticas/imunologia , Células Dendríticas/microbiologia , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais , Células Th1/imunologia , Células Th17/imunologia , Tuberculose/imunologia , Tuberculose/metabolismo , Tuberculose/microbiologia
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