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1.
Mem Inst Oswaldo Cruz ; 115: e200215, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32965331

RESUMO

The human-adapted strains of the Mycobacterium tuberculosis complex (MTBC) comprise seven phylogenetic lineages originally associated with their geographical distribution. Here, we report the genomes of three drug-resistant clinical isolates of the Latin American-Mediterranean (LAM) family collected in Kazakhstan. We utilised whole-genome sequencing to study the distribution and drug resistance of these isolates. Phylogenetic analysis grouped the genomes described in this study with the sequences from Russia, Uzbekistan, and Kazakhstan belonging to the LAM family. One isolate has acquired extensive drug resistance to seven antituberculosis drugs. Our results suggest at least two multi-drug resistant (MDR)/extensively drug-resistant (XDR)-associated genotypes of the LAM family circulate in Kazakhstan.


Assuntos
Antituberculosos/farmacologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Genômica , Genótipo , Humanos , Cazaquistão , América Latina , Filogenia , Tuberculose Resistente a Múltiplos Medicamentos/genética
2.
PLoS One ; 15(8): e0236054, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32750053

RESUMO

INTRODUCTION: Multi-drug resistance is a major challenge in the control of tuberculosis. Despite newer modalities for diagnosis and treatment, people are still suffering from this disease. Understanding the common gene mutations conferring rifampicin and isoniazid resistance is crucial for the implementation of effective molecular tools at local and national levels. Hence, this study aimed to evaluate the molecular detection of rifampicin and isoniazid-resistant gene mutations in M.tuberculosis isolates in Addis Ababa, Ethiopia. METHOD: Health Center-based cross-sectional study was conducted between January and September 2017 in Addis Ababa, Ethiopia. The collected sputum samples were processed for mycobacterial isolation and Region of difference 9 based polymerase chain reaction for species identification. To characterize the rifampicin and isoniazid-resistant M. tuberculosis isolates, a molecular genetic assay (GenoType MTBDRplus) was used; the assay is based on DNA-STRIP technology. RESULT: Culture positivity was confirmed in 82.6% (190/230) of smear-positive newly diagnosed pulmonary tuberculosis cases enrolled in the study. From 190 isolates 93.2% were sensitive for both rifampicin and isoniazid, and 6.8% of the isolates were resistant to at least one of the tested anti-TB drugs. Gene mutations were observed in all studied multidrug resistance-associated gene loci (rpoB, katG, and inhA). Two isolates exhibited heteroresistance, a mutated, as well as wild type sequences, were detected in the respective strains. MDR-TB case was observed in 1.1% (2/190) of the cases. All the MDR-TB cases were positive for HIV and found to have a history of prior hospital admission. CONCLUSION: In our finding a relatively high prevalence of any drug resistance was observed and the overall prevalence of multidrug-resistant tuberculosis was 1.1%.The majority of drug-resistant isolates demonstrated common mutations. Heteroresistant strains were detected, signaling the existence of an M.tuberculosis population with variable responses to anti-tuberculosis drugs or of mixed infections.


Assuntos
Antituberculosos/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Mycobacterium tuberculosis/genética , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Adolescente , Adulto , Antituberculosos/uso terapêutico , Estudos Transversais , Análise Mutacional de DNA , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Etiópia , Feminino , Genes Bacterianos/genética , Loci Gênicos/genética , Humanos , Isoniazida/farmacologia , Isoniazida/uso terapêutico , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Mutação , Mycobacterium tuberculosis/isolamento & purificação , Reação em Cadeia da Polimerase , Rifampina/farmacologia , Rifampina/uso terapêutico , Escarro/microbiologia , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Adulto Jovem
3.
PLoS One ; 15(8): e0237345, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32813724

RESUMO

BACKGROUND: Mixed/polyclonal infections due to different genotypes are reported in Tuberculosis. The current study was designed to understand the fate of mixed infections during the course of treatment and follow-up and its role in disease pathogenesis. METHODS: Sputum samples were collected on 0,1,2,3,6,12 and 24 months from 157 treatment-naïve patients, cultures subjected to Drug-Susceptibility-testing (MGIT 960), spoligotyping, MIRU-VNTR and SNP genotyping. All isolated colonies on thin layer agar (7H11) were subjected to spoligotyping. FINDINGS: One thirty three baseline cultures were positive (133/157, 84.7%), 43(32.3%) had mixture of genotypes. Twenty-four of these patients (55.8%) showed change in genotype while six showed different drug-susceptibility patterns while on treatment. Twenty-three (53.5%) patients with polyclonal infections showed resistance to at least one drug compared to 10/90 (11.1%) monoclonal infections (P<0.0001). Eight patients had recurrent TB, two with a new genotype and two with altered phenotypic DST. CONCLUSIONS: The coexistence of different genotypes and change of genotypes during the same disease episode, while on treatment, confirms constancy of polyclonal infections. The composition of the mixture of genotypes and the relative predominance may be missed by culture due to its limit of detection. Polyclonal infections in TB could be a rule rather than exception and challenges the age-old dogma of reactivation/reinfection.


Assuntos
Antituberculosos/farmacologia , Coinfecção/tratamento farmacológico , Farmacorresistência Bacteriana Múltipla/genética , Mycobacterium tuberculosis/genética , Tuberculose Pulmonar/microbiologia , Adolescente , Adulto , Antituberculosos/uso terapêutico , Técnicas de Tipagem Bacteriana , Evolução Clonal , Coinfecção/epidemiologia , Coinfecção/microbiologia , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Feminino , Seguimentos , Técnicas de Genotipagem , Humanos , Limite de Detecção , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Técnicas de Diagnóstico Molecular , Mycobacterium tuberculosis/isolamento & purificação , Mycobacterium tuberculosis/patogenicidade , Filogenia , Polimorfismo de Nucleotídeo Único , Prevalência , Recidiva , Tuberculose Pulmonar/tratamento farmacológico , Tuberculose Pulmonar/epidemiologia , Adulto Jovem
4.
Fundam Clin Pharmacol ; 34(5): 530-547, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32603486

RESUMO

Patients with COVID-19 are sometimes already being treated for one or more other chronic conditions, especially if they are elderly. Introducing a treatment against COVID-19, either on an outpatient basis or during hospitalization for more severe cases, raises the question of potential drug-drug interactions. Here, we analyzed the potential or proven risk of the co-administration of drugs used for the most common chronic diseases and those currently offered as treatment or undergoing therapeutic trials for COVID-19. Practical recommendations are offered, where possible.


Assuntos
Infecções por Coronavirus/tratamento farmacológico , Pneumonia Viral/tratamento farmacológico , Medicamentos sob Prescrição/farmacologia , Analgésicos/farmacologia , Antiasmáticos/farmacologia , Antibacterianos/farmacologia , Anti-Inflamatórios/farmacologia , Anticoagulantes/farmacologia , Antineoplásicos/farmacologia , Antituberculosos/farmacologia , Antivirais/farmacologia , Betacoronavirus , Fármacos Cardiovasculares/farmacologia , Interações Medicamentosas , Humanos , Hidroxicloroquina/farmacologia , Hipoglicemiantes/farmacologia , Hipolipemiantes/farmacologia , Interferon beta-1b/farmacologia , Pandemias , Medicamentos sob Prescrição/farmacocinética , Psicotrópicos/farmacologia , Receptores de Interleucina/antagonistas & inibidores , Medição de Risco , Hormônios Tireóideos/farmacologia
5.
Mikrobiyol Bul ; 54(2): 211-222, 2020 Apr.
Artigo em Turco | MEDLINE | ID: mdl-32723277

RESUMO

Members of the Mycobacterium tuberculosis complex (MTBC) are the causative agents of tuberculosis (TB). Rifampin resistance in the clinical isolates of MTBC is an important indicator for multidrug resistant-TB (MDR-TB) cases. In this study, it was aimed to evaluate whether the Sensititre MycoTB plaque method is suitable for the routine use in determining drug susceptibility of rifampin resistant MTBC strains. Xpert MTBC/ RIF positive rifampin resistant 100 MTBC isolates were included in the study. Xpert MTBC/RIF (Cepheid, USA) test were performed after the samples were processed by homogenization and decontamination and acid-fast staining. Rifampin resistant clinical samples were cultured in automated MGIT/BACTEC 960 (Becton Dickinson, USA) system and acid fast bacteria (AFB) were investigated. The anti-TB drug susceptibility tests of all culture positive AFB and cord factor identified as MTBC by using a cart test (MPB64, Capilla TB-Neo, Tauns Laboratories, Inc., Numazu, Japan) were performed with the Löwenstein-Jensen proportion method (LJPM) and Sensititre MycoTB (Trek Diagnostic Systems, Cleveland, OH, USA) methods. For the comparison of the methods used, the tests were performed simultaneously. The standard LJPM was performed according to the previously described procedures by World Health Organization and the Sensititre MycoTB plate method was performed as defined by the manufacturer. The final concentrations of isoniazide, rifampin, rifabutin, ethambutol, ofloxacin, moxifloxacin amikacin, kanamycin, cycloserine, ethionamide and p-aminosalicylic acid in Löwenstein-Jensen media for LJPM were 0.2 µg/ml, 40.0 µg/ml, 20.0 µg/ml, 2.0 µg/ml, 2.0 µg/ml, 1.0 µg/ml, 4.0 µg/ml, 30.0 µg/ml, 30.0 µg/ml, 40.0 µg/ml, 40.0 µg/ml and 1.0 µg/ ml, respectively. The results were obtained in 14 days for all of the drugs in the Sensititre MycoTB plate method and in 28-42 days in the LJPM. In this study, the sensitivity and specificity percentages of the Sensititre MycoTB method and the categorical agreement between the two methods were calculated. The sensitivity and specificity percentages of the Sensititre MycoTB plate method were between 84.4-100% and 95.6- 100%, respectively. The categorical agreements between the two methods were 92-100% for the drugs tested in the study. Ethambutol was found to have the lowest sensitivity (84.4%) and specificity (95.6%). The sensitivities of isoniazide, ofloxacin, streptomycin, kanamycin, ethionamide and p-aminosalicylic acid were 98.8%, 90.0%, 94.3%, 87.5%, 91.7% and 95.6%, respectively, while rifampicin, rifabutin, moxifloxacin, amikacin, cycloserine were calculated as 100%. The specificities of isoniazid, rifampicin, rifabutin, ofloxacin, moxifloxacin, amikacin, kanamycin, and cycloserine were found to be 100%, streptomycin, ethionamide and p-aminosalicylic specificity were 96.9%, 97.4% and 98.9% respectively. The categorical agreement was 96-100% in all tested drugs except ethambutol (92%). As a result, although the cost is high, owing to the short incubation period, easy to perform, the possibility for evaluating both first and second line anti-TB drugs simultaneously, determination of minimum inhibitory concentration values of the drugs, long shelf life, high sensitivity, specificity and the categorical agreement values, the Sensititre MycoTB method was determined as an effective method that can be used especially in laboratories where the workload and the MDR-TB cases are high.


Assuntos
Antituberculosos , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis , Rifampina , Tuberculose , Antituberculosos/farmacologia , Etambutol/farmacologia , Testes de Sensibilidade Microbiana/métodos , Mycobacterium tuberculosis/efeitos dos fármacos , Rifampina/farmacologia , Tuberculose/microbiologia
6.
Nat Commun ; 11(1): 3396, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32636380

RESUMO

Arabinosyltransferase B (EmbB) belongs to a family of membrane-bound glycosyltransferases that build the lipidated polysaccharides of the mycobacterial cell envelope, and are targets of anti-tuberculosis drug ethambutol. We present the 3.3 Å resolution single-particle cryo-electron microscopy structure of Mycobacterium smegmatis EmbB, providing insights on substrate binding and reaction mechanism. Mutations that confer ethambutol resistance map mostly around the putative active site, suggesting this to be the location of drug binding.


Assuntos
Mycobacterium smegmatis/enzimologia , Pentosiltransferases/química , Pentosiltransferases/ultraestrutura , Antituberculosos/farmacologia , Domínio Catalítico , Microscopia Crioeletrônica , Farmacorresistência Bacteriana , Etambutol/farmacologia , Lipídeos/química , Mutação , Mycobacterium tuberculosis/enzimologia , Polissacarídeos/química , Ligação Proteica
7.
Proc Natl Acad Sci U S A ; 117(31): 18744-18753, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32680963

RESUMO

Morphological profiling is a method to classify target pathways of antibacterials based on how bacteria respond to treatment through changes to cellular shape and spatial organization. Here we utilized the cell-to-cell variation in morphological features of Mycobacterium tuberculosis bacilli to develop a rapid profiling platform called Morphological Evaluation and Understanding of Stress (MorphEUS). MorphEUS classified 94% of tested drugs correctly into broad categories according to modes of action previously identified in the literature. In the other 6%, MorphEUS pointed to key off-target activities. We observed cell wall damage induced by bedaquiline and moxifloxacin through secondary effects downstream from their main target pathways. We implemented MorphEUS to correctly classify three compounds in a blinded study and identified an off-target effect for one compound that was not readily apparent in previous studies. We anticipate that the ability of MorphEUS to rapidly identify pathways of drug action and the proximal cause of cellular damage in tubercle bacilli will make it applicable to other pathogens and cell types where morphological responses are subtle and heterogeneous.


Assuntos
Antituberculosos/farmacologia , Descoberta de Drogas/métodos , Mycobacterium tuberculosis , Software , Parede Celular/efeitos dos fármacos , Diarilquinolinas , Ensaios de Triagem em Larga Escala , Mycobacterium tuberculosis/citologia , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/metabolismo , Transcriptoma/efeitos dos fármacos
8.
PLoS Pathog ; 16(6): e1008567, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32574211

RESUMO

Efforts at host-directed therapy of tuberculosis have produced little control of the disease in experimental animals to date. This is not surprising, given that few specific host targets have been validated, and reciprocally, many of the compounds tested potentially impact multiple targets with both beneficial and detrimental consequences. This puts a premium on identifying appropriate molecular targets and subjecting them to more selective modulation. We discovered an aminopyrimidine small molecule, 2062, that had no direct antimycobacterial activity, but synergized with rifampin to reduce bacterial burden in Mtb infected macrophages and mice and also dampened lung immunopathology. We used 2062 and its inactive congeners as tool compounds to identify host targets. By biochemical, pharmacologic, transcriptomic and genetic approaches, we found that 2062's beneficial effects on Mtb control and clearance in macrophages and in mice are associated with activation of transcription factor EB via an organellar stress response. 2062-dependent TFEB activation led to improved autophagy, lysosomal acidification and lysosomal degradation, promoting bacterial clearance in macrophages. Deletion of TFEB resulted in the loss of IFNγ-dependent control of Mtb replication in macrophages. 2062 also targeted multiple kinases, such as PIKfyve, VPS34, JAKs and Tyk2, whose inhibition likely limited 2062's efficacy in vivo. These findings support a search for selective activators of TFEB for HDT of TB.


Assuntos
Antituberculosos/farmacologia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Mycobacterium tuberculosis/metabolismo , Rifampina/farmacologia , Tuberculose , Animais , Modelos Animais de Doenças , Feminino , Camundongos , Mycobacterium tuberculosis/patogenicidade , Tuberculose/tratamento farmacológico , Tuberculose/metabolismo , Tuberculose/patologia
9.
J Med Microbiol ; 69(7): 1013-1019, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32579102

RESUMO

Introduction. Multidrug-resistant tuberculosis (MDR-TB) is a major public health problem globally, including in Indonesia. Whole-genome sequencing (WGS) analysis has rarely been used for the study of TB and MDR-TB in Indonesia.Aim. We evaluated the use of WGS for drug-susceptibility testing (DST) and to investigate the population structure of drug-resistant Mycobacterium tuberculosis in Java, Indonesia.Methodology. Thirty suspected MDR-TB isolates were subjected to MGIT 960 system (MGIT)-based DST and to WGS. Phylogenetic analysis was done using the WGS data. Results obtained using MGIT-based DST and WGS-based DST were compared.Results. Agreement between WGS and MGIT was 93.33 % for rifampicin, 83.33 % for isoniazid and 76.67 % for streptomycin but only 63.33 % for ethambutol. Moderate WGS-MGIT agreement was found for second-line drugs including amikacin, kanamycin and fluoroquinolone (73.33-76.67 %). MDR-TB was more common in isolates of the East Asian Lineage (63.3%). No evidence of clonal transmission of DR-TB was found among members of the tested population.Conclusion. Our study demonstrated the applicability of WGS for DST and molecular epidemiology of DR-TB in Java, Indonesia. We found no transmission of DR-TB in Indonesia.


Assuntos
Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Tuberculose Resistente a Múltiplos Medicamentos/genética , Adulto , Antituberculosos/farmacologia , Avaliação Pré-Clínica de Medicamentos/métodos , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla/genética , Feminino , Genótipo , Humanos , Indonésia/epidemiologia , Canamicina/farmacologia , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Epidemiologia Molecular , Mutação , Fenótipo , Filogenia , Rifampina/farmacologia , Estreptomicina/farmacologia , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Sequenciamento Completo do Genoma/métodos
10.
PLoS One ; 15(6): e0234651, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32544174

RESUMO

BACKGROUND: Imipenem, an intravenous antibiotic is recommended for use in drug resistant tuberculosis (DR-TB) when an effective regimen with combination of other second line drugs is not possible. Though the treatment success rates with carbapenems are promising, the twice daily injection of Imipenem usually requires patients to be hospitalized. The Médecins Sans Frontières independent clinic in Mumbai, India implemented ambulatory and home based management of patients receiving Imipenem through the use of port-a-cath. OBJECTIVE: We aimed to describe the adverse events and treatment outcomes of ambulatory pre- and XDR-TB patients initiated on imipenem through port-a-cath between January 2015 and June 2018 and to explore the challenges with this regimen as perceived by healthcare providers and patients. METHODS: A convergent mixed methods study with quantitative (longitudinal descriptive study using the routine data) and qualitative (descriptive study) part conducted concurrently. For the quantitative component, all XDR-TB and pre-XDR-TB initiated on imipenem containing regimen during January 2015-June 2018 were included. For qualitative component, interviews were carried out including patients who initiated on imipenem (n = 5) and healthcare providers (n = 7) involved in providing treatment. Treatment outcomes, culture conversion and adverse events during treatment were described. Thematic analysis was carried out for qualitative component. RESULTS: Of the 70 patients included, the mean age was 28.1 (standard deviation: 11.2) years and 36 (51.4%) were females. Fifty one (72.9%) had XDR-TB. All patients were resistant to fluoroquinilone, levofloxacin. Vomiting was reported by 55 (78.6%) patients and at least one episode of QTC prolongation (more than 500 msec by Fredrecia method) was detected in 25 (35.7%). Port-a-cath block and infection was seen in 11 (15.7%) and 20 (28.6%) patients respectively. Favourable outcomes were seen in 43 (61.4%) patients. Mortality was seen in 22 (31.4%) patients, 2 (2.9%) were lost-to-follow-up and 3 (4.3%) were declared as treatment failure. The overarching theme of the qualitative analysis was: Challenges in delivering Imipenem via port-a-cath device in ambulatory care. Major challenges identified were difficulties in adhering to drug dose timelines, vomiting, restricted mobility due to port-a-cath, paucity of infection control and space constraints at patients' home for optimal care. CONCLUSION: Administration of imipenem was feasible through port-a-cath. Though outcomes with ambulatory based imipenem containing regimens were promising, there were several challenges in providing care. The feasibility of infusion at day care facilities needs to explored to overcome challenges in infusion at patients home.


Assuntos
Tuberculose Extensivamente Resistente a Medicamentos/tratamento farmacológico , Imipenem/administração & dosagem , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Adolescente , Adulto , Instituições de Assistência Ambulatorial/normas , Antituberculosos/efeitos adversos , Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Tuberculose Extensivamente Resistente a Medicamentos/complicações , Feminino , Serviços de Assistência Domiciliar/normas , Humanos , Imipenem/efeitos adversos , Imipenem/normas , Índia , Controle de Infecções/normas , Masculino , Resultado do Tratamento , Tuberculose Resistente a Múltiplos Medicamentos/complicações , Dispositivos de Acesso Vascular , Vômito/induzido quimicamente
11.
BMC Infect Dis ; 20(1): 390, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32487030

RESUMO

BACKGROUND: Fluoroquinolones are broad-spectrum antibiotics that are recommended, and increasingly important, for the treatment of multidrug-resistant tuberculosis (MDR-TB). Resistance to fluoroquinolones is caused by mutations in the Quinolone Resistance Determining Region (QRDR) of gyrA and gyrB genes of Mycobacterium tuberculosis. In this study, we characterized the phenotypic and genotypic resistance to fluoroquinolones for the first time in northeast Iran. METHODS: A total of 123 Mycobacterium tuberculosis isolates, including 111 clinical and 12 collected multidrug-resistant isolates were studied. Also, 19 WHO quality control strains were included in the study. The phenotypic susceptibility was determined by the proportion method on Löwenstein-Jensen medium. The molecular cause of resistance to the fluoroquinolone drugs ofloxacin and levofloxacin was investigated by sequencing of the QRDR region of the gyrA and gyrB genes. RESULTS: Among 123 isolates, six (4.8%) were fluoroquinolone-resistant according to phenotypic methods, and genotypically three of them had a mutation at codon 94 of the gyrA gene (Asp→ Gly) which was earlier reported to cause resistance. All three remaining phenotypically resistant isolates had a nucleotide change in codon 95. No mutations were found in the gyrB gene. Five of the 19 WHO quality control strains, were phenotypically fluoroquinolone-resistant, four of them were genotypically resistant with mutations at codon 90, 91 of the gyrA gene and one resistant strain had no detected mutation. CONCLUSIONS: Mutation at codon 94 of the gyrA gene, was the main cause of fluoroquinolone resistance among M. tuberculosis isolates in our region. In 3/6 fluoroquinolone-resistant isolates, no mutations were found in either gyrA or gyrB. Therefore, it can be concluded that various other factors may lead to fluoroquinolone resistance, such as active efflux pumps, decreased cell wall permeability, and drug inactivation.


Assuntos
Antituberculosos/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Fluoroquinolonas/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Códon , Farmacorresistência Bacteriana/genética , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla/genética , Feminino , Genótipo , Humanos , Irã (Geográfico) , Levofloxacino/farmacologia , Masculino , Testes de Sensibilidade Microbiana , Mutação , Mycobacterium tuberculosis/isolamento & purificação , Ofloxacino/farmacologia , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia
12.
Nat Commun ; 11(1): 3062, 2020 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-32546788

RESUMO

Anti-tuberculosis (TB) drugs, while being highly potent in vitro, require prolonged treatment to control Mycobacterium tuberculosis (Mtb) infections in vivo. We report here that mesenchymal stem cells (MSCs) shelter Mtb to help tolerate anti-TB drugs. MSCs readily take up Mtb and allow unabated mycobacterial growth despite having a functional innate pathway of phagosome maturation. Unlike macrophage-resident ones, MSC-resident Mtb tolerates anti-TB drugs remarkably well, a phenomenon requiring proteins ABCC1, ABCG2 and vacuolar-type H+ATPases. Additionally, the classic pro-inflammatory cytokines IFNγ and TNFα aid mycobacterial growth within MSCs. Mechanistically, evading drugs and inflammatory cytokines by MSC-resident Mtb is dependent on elevated PGE2 signaling, which we verify in vivo analyzing sorted CD45-Sca1+CD73+-MSCs from lungs of infected mice. Moreover, MSCs are observed in and around human tuberculosis granulomas, harboring Mtb bacilli. We therefore propose, targeting the unique immune-privileged niche, provided by MSCs to Mtb, can have a major impact on tuberculosis prevention and cure.


Assuntos
Antituberculosos/farmacologia , Células-Tronco Mesenquimais/microbiologia , Mycobacterium tuberculosis/patogenicidade , Nicho de Células-Tronco/imunologia , Tuberculose/microbiologia , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Células Cultivadas , Dinoprostona/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Interferon gama/farmacologia , Isoniazida/farmacologia , Lisossomos/microbiologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/crescimento & desenvolvimento , Proteínas de Neoplasias/metabolismo , Fagossomos/microbiologia , Tuberculose/patologia , Tuberculose Pulmonar/tratamento farmacológico , Tuberculose Pulmonar/microbiologia , Tuberculose Pulmonar/patologia , Fator de Necrose Tumoral alfa/farmacologia
13.
Artigo em Inglês | MEDLINE | ID: mdl-32520212

RESUMO

Drug resistant tuberculosis (DR-TB) is challenging particularly in developing countries. As such, a previous investigation gave the first insight into the mutational status of the Rifampicin Resistance Determining Region (RRDR) of rpoB gene among a restricted number of MTB patients' residents in the Northern Morocco. The purpose of this study was to investigate rpoB mutation types and frequencies associated with resistance to Rifampicin in a larger panel of MTB patients and to evaluate the usefulness of these mutations to improve the diagnosis of resistance to Rifampicin. A panel of 301 consecutive sputum samples belonging to patients suscpected of having TB from Northern Morocco was collected at the Pasteur Institute of Tangier between 2014-2017. Samples were subjected to conventionel microbiological tests. Evaluation of rpoB muational status was assessed by PCR amplification and sequencing of the RRDR of the rpoB gene. DST results showed that 26.4% of strains were MDR. Sequencing results reported single point mutations in 36 of 65 RIFR isolates of which two had two mutations. Aminoacid substitutions in the codon Ser531Leu occurred at the highest frequency (34.46%). Overall, 10 aminoacid substitutions have been registered, and the H526S substitution was reported for the first time. The present study highlighted that resistance to RIF is a reliable marker of MDR-TB, the common mutations successfully detected in the rpoB 531, rpoB526 and rpoB516 codons provide a foundation for the implementation of molecular approaches such as Hain and GeneXpert as a routine tests to detect DR-TB. However, considerable work is still necessary to identify extensive mutations associated with DR-TB.


Assuntos
Antituberculosos/farmacologia , Mutação/genética , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Rifampina/farmacologia , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Adulto , Feminino , Genótipo , Humanos , Masculino , Testes de Sensibilidade Microbiana , Reação em Cadeia da Polimerase
14.
PLoS One ; 15(5): e0233500, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32421749

RESUMO

BACKGROUND: Meta-analyses on impact of isoniazid-resistant tuberculosis informed the World Health Organization recommendation of a levofloxacin-strengthened rifampicin-based regimen. We estimated the effect of initial rifampicin resistance (Rr) and/or isoniazid resistance (Hr) on treatment failure or relapse. We also determined the frequency of missed initial and acquired Rr to estimate the impact of true Hr. METHODS: Retrospective analysis of 7291 treatment episodes with known initial isoniazid and rifampicin status obtained from individual patient databases maintained by the Damien Foundation Bangladesh over 20 years. Drug susceptibility test results were confirmed by the programme's designated supra-national tuberculosis laboratory. To detect missed Rr among isolates routinely classified as Hr, rpoB gene sequencing was done randomly and on a sample selected for suspected missed Rr. RESULTS: Initial Hr caused a large recurrence excess after the 8-month regimen for new cases (rifampicin for two months), but had little impact on rifampicin-throughout regimens: (6 months, new cases; 3.8%; OR 0.8, 95%CI:0.3,2.8; 8 months, retreatment cases: 7.3%, OR 1.8; 95%CI:1.3,2.6). Rr was missed in 7.6% of randomly selected "Hr" strains. Acquired Rr was frequent among recurrences on rifampicin-throughout regimens, particularly after the retreatment regimen (31.9%). It was higher in mono-Hr (29.3%; aOR 3.5, 95%CI:1.5,8.5) and poly-Hr (53.3%; aOR 10.2, 95%CI 4.4,23.7) than in susceptible tuberculosis, but virtually absent after the 8-month new case regimen. Comparing Bangladesh (low Rr prevalence) with a high Rr prevalence setting,true Hr corrected for missed Rr caused only 2-3 treatment failures per 1000 TB cases (of whom 27% were retreatments) in both. CONCLUSIONS: Our analysis reveals a non-negligible extent of misclassifying as isoniazid resistance of what is actually missed multidrug-resistant tuberculosis. Recommending for such cases a "strengthened" regimen containing a fluoroquinolone provokes a direct route to extensive resistance while offering little benefit against the minor role of true Hr tuberculosis in rifampicin-throughout first-line regimen.


Assuntos
Resistência a Medicamentos , Isoniazida/farmacologia , Rifampina/farmacologia , Adulto , Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Bangladesh , Erros de Diagnóstico , Resistência a Medicamentos/efeitos dos fármacos , Fluoroquinolonas/uso terapêutico , Humanos , Isoniazida/uso terapêutico , Recidiva , Estudos Retrospectivos , Rifampina/uso terapêutico , Falha de Tratamento , Resultado do Tratamento , Tuberculose Resistente a Múltiplos Medicamentos
15.
Infect Dis Poverty ; 9(1): 57, 2020 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-32460836

RESUMO

BACKGROUND: Despite significant advancements in the treatment and diagnosis of tuberculosis (TB) over the past decade, drug-resistant TB remains an increasing threat to public health. TB outbreaks are most commonly reported in schools considering the delay in TB diagnosis, sustained contact, and overcrowding observed in schools. This report describes multidrug-resistant TB (MDR-TB) transmission in a school in Zhejiang Province. We aimed to raise awareness regarding MDR-TB transmission among students. CASE PRESENTATION: The index patient was a 16-year-old girl in the second year of junior middle school in Zhejiang Province, China, who had been experiencing persistent cough and expectoration for 37 days since 1 March 2014. She tested positive for smear pulmonary and extrapulmonary TB on 8 April 2014 and was subsequently diagnosed with MDR-TB on 1 May 2014. However, the patient was resistant to isoniazid, rifampicin, ethambutol, and streptomycin. Thus, she was suspended from school for anti-TB treatment. All 54 students who were in close contact with the index patient in the same class were screened, and 5 tested positive on the tuberculin skin test. Their exposure time to the index patient was approximately 37 days. Three classmates were subsequently diagnosed with MDR-TB, with similar resistance profiles nearly two years later. Their average discovery delay was 55 days. These three classmates were also suspended from school for anti-TB treatment. During the treatment period, four students visited the local TB-designated hospital for further consultation every month and were followed up once a month by the local community health service center until they were completely cured. CONCLUSIONS: Discovery delay for an index patient played a primary role in MDR-TB transmission inside the school. To immediately detect TB, morning examinations in schools should be performed. TB trackers and case managers should work closely with public health workers and physicians in cases of TB outbreaks or transmissions involving students. Moreover, individuals who are in close contact with MDR-TB patients should undergo careful clinical follow-up for at least two years. Implementing a joint examination strategy to ensure early detection, diagnosis, and treatment of MDR-TB transmission is recommended.


Assuntos
Antituberculosos/farmacologia , Tuberculose Resistente a Múltiplos Medicamentos/transmissão , Adolescente , China , Feminino , Humanos , Masculino , Instituições Acadêmicas , Estudantes , Tuberculose Resistente a Múltiplos Medicamentos/diagnóstico
16.
Eur J Med Chem ; 200: 112390, 2020 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-32447196

RESUMO

The mycobacterial membrane protein Large 3 (MmpL3) is an inner membrane protein that transports trehalose-monomycolates, precursors for trehalose-dimycolates and mycolic acids that make up essential components of the mycobacterial outer membrane. Inhibition of MmpL3 weakens the mycobacterial cell wall and ultimately results in cell death in both in vitro and in vivo infection models. This highlights the therapeutic potential of MmpL3 as a drug target. High-throughput whole-cell screening along with whole genome sequencing of resistant mutants has identified numerous classes of compounds that can be classified as MmpL3 inhibitors. In this review, we provide insights into the current development of various MmpL3 inhibitors and discuss the potential challenges in this area.


Assuntos
Antituberculosos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Parede Celular/efeitos dos fármacos , Antituberculosos/uso terapêutico , Proteínas de Bactérias/metabolismo , Fatores Corda/metabolismo , Humanos , Proteínas de Membrana Transportadoras/metabolismo , Mycobacterium tuberculosis/efeitos dos fármacos , Ácidos Micólicos/metabolismo
17.
Mem Inst Oswaldo Cruz ; 115: e200055, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32401997

RESUMO

BACKGROUND Tuberculosis (TB) is an infectious disease caused by the bacterium Mycobacterium tuberculosis, and the number of new cases of multidrug resistant TB (MDR-TB), pre extensively drug-resistant TB (pre-XDR-TB) and extensively drug-resistant TB (XDR-TB) has increased considerably worldwide. OBJECTIVES Herein, using 156 M. tuberculosis isolates from 106 patients previously classified as MDR or pre-XDR or XDR isolates, we investigated the genetic mutation profiles associated with phenotypic resistances in patients with MDR-TB, pre-XDR-TB and XDR-TB, treatment outcomes and resistance evolution. METHODS Molecular analyses were performed by partial sequencing of the rpoB, katG, gyrA, gyrB, rrs genes and analysis of the fabG-inhA promoter region. Clinical, epidemiologic and demographic data were obtained from the TB Notification database system of São Paulo (TB-WEB) and the Information System for Special Tuberculosis Treatments (SITE-TB). FINDINGS Drug resistance was attributed to previously known mutations and a novel Asp449Val mutation in gyrB was observed in four isolates from the same patient. Ten patients had more than one isolate evaluated and eight of these patients displayed resistance progression. MAIN CONCLUSIONS The present study is the first to report the frequency of mutations related to second-line drug resistance in MDR-TB, pre-XDR-TB and XDR-TB isolates. The results could lead to the improvement of available technologies for the rapid detection of drug resistant TB.


Assuntos
Antituberculosos/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Mutação/genética , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Adolescente , Adulto , Brasil , Tuberculose Extensivamente Resistente a Medicamentos/microbiologia , Feminino , Humanos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Mycobacterium tuberculosis/isolamento & purificação , Fatores Socioeconômicos , Adulto Jovem
18.
J Med Chem ; 63(10): 5367-5386, 2020 05 28.
Artigo em Inglês | MEDLINE | ID: mdl-32342688

RESUMO

In search of novel drugs against tuberculosis, we previously discovered and profiled a novel hydantoin-based family that demonstrated highly promising in vitro potency against Mycobacterium. tuberculosis. The compounds were found to be noncovalent inhibitors of DprE1, a subunit of decaprenylphosphoryl-ß-d-ribose-2'-epimerase. This protein, localized in the periplasmic space of the mycobacterial cell wall, was shown to be an essential and vulnerable antimycobacterial drug target. Here, we report the further SAR exploration of this chemical family through more than 80 new analogues. Among these, the most active representatives combined submicromolar cellular potency and nanomolar target affinity with balanced physicochemical properties and low human cytotoxicity. Moreover, we demonstrate in vivo activity in an acute Mtb infection model and provide further proof of DprE1 being the target of the hydantoins. Overall, the hydantoin family of DprE1 inhibitors represents a promising noncovalent lead series for the discovery of novel antituberculosis agents.


Assuntos
Oxirredutases do Álcool/antagonistas & inibidores , Antituberculosos/química , Antituberculosos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Hidantoínas/química , Hidantoínas/farmacologia , Oxirredutases do Álcool/metabolismo , Animais , Antituberculosos/metabolismo , Proteínas de Bactérias/metabolismo , Feminino , Células Hep G2 , Humanos , Hidantoínas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/metabolismo , Ressonância Magnética Nuclear Biomolecular/métodos , Tuberculose/tratamento farmacológico , Tuberculose/metabolismo
19.
Int J Infect Dis ; 96: 48-53, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32339720

RESUMO

OBJECTIVE: To evaluate the performance of whole-genome sequencing (WGS) for predicting Mycobacterium tuberculosis (MTB) drug resistance. METHODS: 276 rifampin-resistance tuberculosis (RR-TB) and 30 rifampicin-sensitive clinical isolates were randomly selected from patients with tuberculosis in Shanghai Pulmonary Hospital (SPH). Phenotypic drug susceptibility testing (DST) against six anti-TB drugs was performed, and WGS was used to predict the drug resistance using an online 'TB-Profiler' tool. RESULTS: Using phenotypic susceptibility as the gold standard, the overall sensitivities and specificities for WGS were 94.53% and 92.00% for isoniazid, 97.10% and 100.00% for rifampicin, 97.46% and 64.36% for ethambutol, 97.14% and 95.83% or streptomycin, 93.02% and 98.87% for ofloxacin, and 75.00% and 100.00% for amikacin, respectively. The concordances of WGS-based DST and phenotypic DST were: isoniazid (94.12%), rifampicin (97.39%), ethambutol (77.12%), streptomycin (96.73%), ofloxacin (96.41%) and amikacin (97.06%). CONCLUSIONS: WGS could be a promising approach to predict resistance to isoniazid, rifampicin, ethambutol, streptomycin, ofloxacin, and amikacin.


Assuntos
Antituberculosos/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Sequenciamento Completo do Genoma , Adulto , Amicacina/farmacologia , China , Farmacorresistência Bacteriana/genética , Etambutol/farmacologia , Humanos , Isoniazida/farmacologia , Masculino , Mycobacterium tuberculosis/isolamento & purificação , Ofloxacino/farmacologia , Rifampina/farmacologia , Estreptomicina/farmacologia , Tuberculose/microbiologia
20.
Infect Dis Poverty ; 9(1): 38, 2020 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-32299480

RESUMO

BACKGROUND: Drug resistant tuberculosis poses a great challenge for tuberculosis control worldwide. Timely determination of drug resistance and effective individual treatment are essential for blocking the transmission of drug resistant Mycobacterium tuberculosis. We aimed to establish and evaluate the accuracy of a reverse dot blot hybridization (RDBH) assay to simultaneously detect the resistance of four anti-tuberculosis drugs in M. tuberculosis isolated in China. METHODS: In this study, we applied a RDBH assay to simultaneously detect the resistance of rifampicin (RIF), isoniazid (INH), streptomycin (SM) and ethambutol (EMB) in 320 clinical M. tuberculosis isolates and compared the results to that from phenotypic drug susceptibility testing (DST) and sequencing. The RDBH assay was designed to test up to 42 samples at a time. Pearson's chi-square test was used to compute the statistical measures of the RDBH assay using the phenotypic DST or sequencing as the gold standard method, and Kappa identity test was used to determine the consistency between the RDBH assay and the phenotypic DST or sequencing. RESULTS: The results showed that the concordances between phenotypic DST and RDBH assay were 95% for RIF, 92.8% for INH, 84.7% for SM, 77.2% for EMB and the concordances between sequencing and RDBH assay were 97.8% for RIF, 98.8% for INH, 99.1% for SM, 93.4% for EMB. Compared to the phenotypic DST results, the sensitivity and specificity of the RDBH assay for resistance detection were 92.4 and 98.5% for RIF, 90.3 and 97.3% for INH, 77.4 and 91.5% for SM, 61.4 and 85.7% for EMB, respectively; compared to sequencing, the sensitivity and specificity of the RDBH assay were 97.7 and 97.9% for RIF, 97.9 and 100.0% for INH, 97.8 and 100.0% for SM, 82.6 and 99.1% for EMB, respectively. The turnaround time of the RDBH assay was 7 h for testing 42 samples. CONCLUSIONS: Our data suggested that the RDBH assay could serve as a rapid and efficient method for testing the resistance of M. tuberculosis against RIF, INH, SM and EMB, enabling early administration of appropriate treatment regimens to the affected drug resistant tuberculosis patients.


Assuntos
Antituberculosos/farmacologia , Farmacorresistência Bacteriana , Immunoblotting/métodos , Testes de Sensibilidade Microbiana/métodos , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose/microbiologia , China , Etambutol/farmacologia , Humanos , Isoniazida/farmacologia , Testes de Sensibilidade Microbiana/instrumentação , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/isolamento & purificação , Rifampina/farmacologia , Sensibilidade e Especificidade , Estreptomicina/farmacologia
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