RESUMO
RATIONALE: The outcome of fully intermittent thrice-weekly antituberculosis treatment of various durations in HIV-associated tuberculosis is unclear. OBJECTIVES: To compare the efficacy of an intermittent 6-month regimen (Reg6M: 2EHRZ(3)/4HR(3) [ethambutol, 1,200 mg; isoniazid, 600 mg; rifampicin, 450 or 600 mg depending on body weight <60 or > or =60 kg; and pyrazinamide, 1,500 mg for 2 mo; followed by 4 mo of isoniazid and rifampicin at the same doses]) versus a 9-month regimen (Reg9M: 2EHRZ(3)/7HR(3)) in HIV/tuberculosis (TB). METHODS: HIV-infected patients with newly diagnosed pulmonary or extrapulmonary TB were randomly assigned to Reg6M (n = 167) or Reg9M (n = 160) and monitored by determination of clinical, immunological, and bacteriological parameters for 36 months. Primary outcomes included favorable responses at the end of treatment and recurrences during follow-up, whereas the secondary outcome was death. Intent-to-treat and on-treatment analyses were performed. All patients were antiretroviral treatment-naive during treatment. MEASUREMENTS AND MAIN RESULTS: Of the patients, 70% had culture-positive pulmonary TB; the median viral load was 155,000 copies/ml and the CD4(+) cell count was 160 cells/mm(3). Favorable response to antituberculosis treatment was similar by intent to treat (Reg6M, 83% and Reg9M, 76%; P = not significant). Bacteriological recurrences occurred significantly more often in Reg6M than in Reg9M (15 vs. 7%; P < 0.05) although overall recurrences were not significantly different (Reg6M, 19% vs. Reg9M, 13%). By 36 months, 36% of patients undergoing Reg6M and 35% undergoing Reg9M had died, with no significant difference between regimens. All 19 patients who failed treatment developed acquired rifamycin resistance (ARR), the main risk factor being baseline isoniazid resistance. CONCLUSIONS: Among antiretroviral treatment-naive HIV-infected patients with TB, a 9-month regimen resulted in a similar outcome at the end of treatment but a significantly lower bacteriological recurrence rate compared with a 6-month thrice-weekly regimen. ARR was high with these intermittent regimens and neither mortality nor ARR was altered by lengthening TB treatment. Clinical Trials Registry Information: ID# NCT00376012 registered at www.clinicaltrials.gov.
Assuntos
Infecções Oportunistas Relacionadas com a AIDS/tratamento farmacológico , Infecções Oportunistas Relacionadas com a AIDS/microbiologia , Antituberculosos/administração & dosagem , Infecções por HIV/microbiologia , Tuberculose/tratamento farmacológico , Tuberculose/virologia , Infecções Oportunistas Relacionadas com a AIDS/virologia , Adulto , Antituberculosos/efeitos adversos , Contagem de Linfócito CD4 , Esquema de Medicação , Farmacorresistência Bacteriana Múltipla , Quimioterapia Combinada , Etambutol/administração & dosagem , Etambutol/efeitos adversos , Feminino , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , Humanos , Isoniazida/administração & dosagem , Isoniazida/efeitos adversos , Estimativa de Kaplan-Meier , Modelos Logísticos , Masculino , Adesão à Medicação , Pirazinamida/administração & dosagem , Pirazinamida/efeitos adversos , Rifampina/administração & dosagem , Rifampina/efeitos adversos , Tuberculose/microbiologia , Carga Viral/efeitos dos fármacosRESUMO
Mannose binding lectin (MBL) plays an important role in innate immunity. Plasma MBL levels and MBL2 gene polymorphisms were studied in HIV-1 infected patients without tuberculosis (HIV+TB-) (n=151) and with tuberculosis (HIV+TB+) (n=109), HIV negative tuberculosis patients (HIV-TB+) (n=148) and healthy controls (n=146) by ELISA and genotyping by polymerase chain reaction based methods. MBL levels were significantly increased among HIV-TB+ and HIV+TB+ patients than controls and HIV+TB- patients (P<0.05). A significantly increased frequency of OO genotype of structural polymorphism and YY genotype of -221Y/X was observed among HIV-TB+ patients than controls. In HIV+TB+ patients, a significantly increased frequency of YA/YA diplotype (associated with very high MBL levels) was observed compared to controls (P=0.03). In HIV+TB+ patients, a significantly decreased frequency of medium MBL expression diplotypes (XA/XA and YA/YO) were noticed compared to HIV+TB- and healthy controls. The results suggest that YA/YA diplotype associated with very high MBL levels may predispose HIV-infected patients to tuberculosis while O/O genotype associated with very low MBL levels may be associated with susceptibility to tuberculosis in HIV uninfected individuals. Medium MBL expression diplotypes might protect against development of TB in HIV-infected patients.
Assuntos
Infecções Oportunistas Relacionadas com a AIDS/genética , HIV-1 , Lectina de Ligação a Manose/genética , Tuberculose/genética , Infecções Oportunistas Relacionadas com a AIDS/imunologia , Adulto , Feminino , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Haplótipos , Humanos , Imunidade Inata , Masculino , Lectina de Ligação a Manose/sangue , Pessoa de Meia-Idade , Polimorfismo Genético , Tuberculose/imunologia , Tuberculose Pulmonar/genética , Tuberculose Pulmonar/imunologiaRESUMO
Access to antiretroviral therapy has expanded in many developing countries, including India. The standard first-line regimens consist of a combination of two nucleoside reverse transcriptase inhibitors and a nonnucleoside reverse transcriptase inhibitor, in a fixed drug combination. Data regarding resistance to these drugs are scarce, especially in children. We evaluated the pattern of polymorphism and potential drug resistance mutations (DRMs) in HIV-1 isolates from 48 children naive to antiretroviral therapy attending the outpatient clinics of the Tuberculosis Research Center in Chennai. The samples were subjected to genotyping of reverse transcriptase (RT) and protease genes. All the samples showed significant polymorphisms in both RT and protease genes, but none had major DRMs. The currently recommended generic first-line antiretroviral drug combination is an appropriate treatment strategy for HIV-1-infected children in India.
Assuntos
Fármacos Anti-HIV/farmacologia , Farmacorresistência Viral/genética , Infecções por HIV/virologia , HIV-1/classificação , HIV-1/genética , Criança , Pré-Escolar , Genótipo , Infecções por HIV/epidemiologia , Protease de HIV/genética , Transcriptase Reversa do HIV/genética , HIV-1/efeitos dos fármacos , HIV-1/enzimologia , Humanos , Índia/epidemiologia , Dados de Sequência Molecular , Mutação , Filogenia , Polimorfismo Genético , Análise de Sequência de DNARESUMO
Mannose-binding lectin (MBL) plays an important role in innate immunity. Functional mutant homozygotes of the MBL gene affect the serum MBL levels and have been correlated with disease susceptibility. We have studied the regulatory role of variant MBL genotypes on serum MBL level and macrophage phagocytosis with live Mycobacterium tuberculosis, and the lymphoproliferative response to M. tuberculosis culture filtrate antigen in pulmonary tuberculosis (PTB) patients (n = 48) and normal healthy subjects (NHS) (n = 58). The total serum MBL level was higher in PTB patients than in NHS (P = 0.0085). Patients and NHS with AA genotype (homozygotes of MBL - common alleles) showed a very high serum MBL level, and those with OO genotype (functional mutant homozygotes of MBL - less frequent alleles) showed a very low MBL level (AA vs. OO: NHS, P = 3.3 x 10(-9); PTB, P = 3.1 x 10(-9)). A significantly lower phagocytosis was observed in NHS with AA genotype than in NHS with AO (heterozygotes) genotype (P = 0.046). In PTB patients, no such difference was observed. A negative correlation of macrophage phagocytosis with MBL level was seen in patients and NHS (P = 0.019). Antigen-induced lymphoproliferative response was significantly decreased in PTB patients with AA genotype as compared with NHS with AA genotype (P = 0.036). The present study suggests that AA genotype with its associated higher serum MBL levels plays a regulatory role in immunity to tuberculosis than functional mutant homozygotes (OO genotype) with its associated lower level of MBL.
Assuntos
Macrófagos/imunologia , Lectina de Ligação a Manose/imunologia , Mycobacterium tuberculosis/imunologia , Fagocitose/imunologia , Tuberculose Pulmonar/imunologia , Adulto , Alelos , DNA/química , DNA/genética , Feminino , Genótipo , Humanos , Ativação Linfocitária/imunologia , Macrófagos/microbiologia , Masculino , Lectina de Ligação a Manose/sangue , Lectina de Ligação a Manose/genética , Reação em Cadeia da Polimerase , Tuberculose Pulmonar/microbiologiaRESUMO
BACKGROUND: Shortening tuberculosis (TB) treatment duration is a research priority. This paper presents data from a prematurely terminated randomized clinical trial, of 4-month moxifloxacin or gatifloxacin regimens, in South India. METHODS: Newly diagnosed, sputum-positive HIV-negative pulmonary TB patients were randomly allocated to receive gatifloxacin or moxifloxacin, along with isoniazid and rifampicin for 4 months with pyrazinamide for first 2 months (G or M) or isoniazid and rifampicin for 6 months with ethambutol and pyrazinamide for first 2 months (C). All regimens were administered thrice-weekly. Clinical and bacteriological assessments were done monthly during treatment and for 24 months post-treatment. The Data and Safety Monitoring Board recommended termination of the trial due to high TB recurrence rates in the G and M regimens. RESULTS: Of 416 patients in intent-to-treat analysis, 6 (5%) of 124, 2 (2%) of 110 and 2 (2%) of 137 patients with drug-susceptible TB in the G, M and C arms respectively had unfavorable response at the end of treatment; during the next 24 months, 17 (15%) of 115, 11 (11%) of 104 and 8 (6%) of 132 patients respectively, had TB recurrence. Of 38 drug-resistant patients 1 of 8 and 3 of 26 in the G and C arms respectively had unfavourable response at the end of treatment; and TB recurrence occurred in 2 of 7 and 2 of 23 patients, respectively. The differences in TB recurrence rates between the G and C arms was statistically significant (p = 0.02). Gastro-intestinal symptoms occurred in 23%, 22% and 9% of patients in the G, M and C arms respectively, but most reactions were mild and manageable with symptomatic measures; 1% required regimen modification. CONCLUSIONS: 4-month thrice-weekly regimens of gatifloxacin or moxifloxacin with isoniazid, rifampicin and pyrazinamide, were inferior to standard 6-month treatment, in patients with newly diagnosed sputum positive pulmonary TB. TRIAL REGISTRATION: Clinical Trials Registry of India CTRI/2012/10/003060.
Assuntos
Antituberculosos/uso terapêutico , Compostos Aza/uso terapêutico , Fluoroquinolonas/uso terapêutico , Quinolinas/uso terapêutico , Tuberculose Pulmonar/tratamento farmacológico , Adulto , Antituberculosos/administração & dosagem , Antituberculosos/efeitos adversos , Compostos Aza/administração & dosagem , Esquema de Medicação , Quimioterapia Combinada , Feminino , Fluoroquinolonas/administração & dosagem , Fluoroquinolonas/efeitos adversos , Gatifloxacina , Humanos , Masculino , Pessoa de Meia-Idade , Moxifloxacina , Quinolinas/administração & dosagem , Recidiva , Escarro/microbiologia , Resultado do Tratamento , Tuberculose Pulmonar/diagnóstico , Tuberculose Pulmonar/mortalidade , Adulto JovemRESUMO
There is limited information on the prevalence and pattern of HIV drug-resistant mutations (DRMs) among HIV-1-coinfected tuberculosis (TB) patients before and after antiretroviral treatment. Patients with HIV-1 and TB were recruited into a clinical trial from two different once-daily antiretroviral regimens and followed for a period of 6 months after ART initiation. Patients were treated with standard short-course anti-TB treatment (2EHRZ3/4RH3) and were randomized to receive ddI/3TC with either nevirapine or efavirenz, once daily. Genotypic drug resistance (DR) testing was carried out for the pol gene at baseline and at the time of virological failure. At baseline, major DRMs with respect to NNRTIs (G190GA) and TAMs (T215S and I) were observed in 3 out of 107 patients. Of 15 treatment failures, 14 had more than one major NRTI and NNRTI mutation. V106M was the major NNRTI mutation that emerged in EFZ and Y181C in the NVP group. Among NRTI mutations, M184V was the commonest followed by L74I/V. Primary drug resistance to antiretroviral drugs was low among HIV-1 co-infected TB patients in south India. A once-daily regimen of ddI/3TC/EFZ or NVP results in a specific pattern of NNRTI mutations and negligible thymidine analog mutations (TAMs).
Assuntos
Fármacos Anti-HIV/farmacologia , Farmacorresistência Viral/genética , Infecções por HIV/complicações , Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Mutação , Tuberculose/complicações , Adulto , Alcinos , Fármacos Anti-HIV/administração & dosagem , Fármacos Anti-HIV/uso terapêutico , Antituberculosos/uso terapêutico , Benzoxazinas/administração & dosagem , Benzoxazinas/farmacologia , Benzoxazinas/uso terapêutico , Ciclopropanos , Didanosina/administração & dosagem , Didanosina/farmacologia , Didanosina/uso terapêutico , Quimioterapia Combinada , Feminino , Infecções por HIV/virologia , HIV-1/genética , Humanos , Índia , Lamivudina/administração & dosagem , Lamivudina/farmacologia , Lamivudina/uso terapêutico , Masculino , Nevirapina/administração & dosagem , Nevirapina/farmacologia , Nevirapina/uso terapêutico , Inibidores da Transcriptase Reversa/administração & dosagem , Inibidores da Transcriptase Reversa/farmacologia , Inibidores da Transcriptase Reversa/uso terapêutico , Falha de Tratamento , Tuberculose/tratamento farmacológicoRESUMO
Heat shock promoters of mycobacteria are strong promoters that become rapidly upregulated during macrophage infection and thus serve as valuable candidates for expressing foreign antigens in recombinant BCG vaccine. In the present study, a new heat shock promoter controlling the expression of the groESL1 operon was identified and characterized. Mycobacterium tuberculosis groESL1 operon codes for the immunodominant 10 kDa (Rv3418c, GroES/Cpn10/Hsp10) and 60 kDa (Rv3417c, GroEL1/Cpn60.1/Hsp60) heat shock proteins. The basal promoter region was 115 bp, while enhanced activity was seen only with a 277-bp fragment. No promoter element was seen in the groES-groEL1 intergenic region. This operon codes for a bicistronic mRNA transcript as determined by reverse transcriptase-PCR and Northern blot analysis. Primer extension analysis identified two transcriptional start sites (TSSs) TSS1 (-236) and TSS2 (-171), out of which one (TSS2) was heat inducible. The groE promoter was more active than the groEL2 promoter in Mycobacterium smegmatis. Further, it was found to be differentially regulated under stress conditions, while the groEL2 promoter was constitutive.
Assuntos
Proteínas de Bactérias/biossíntese , Chaperoninas/biossíntese , DNA Bacteriano/genética , Mycobacterium tuberculosis/fisiologia , Regiões Promotoras Genéticas , Sequência de Bases , Northern Blotting , DNA Intergênico , Regulação Bacteriana da Expressão Gênica , Genes Reporter , Dados de Sequência Molecular , Óperon , RNA Bacteriano/biossíntese , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sítio de Iniciação de Transcrição , beta-Galactosidase/genética , beta-Galactosidase/metabolismoRESUMO
The gamma interferon (IFN-gamma) enzyme-linked immunospot (ELISPOT) assay is used routinely to evaluate the potency of human immunodeficiency virus (HIV) vaccine candidates and other vaccine candidates. In order to compare candidates and pool data from multiple trial laboratories, validated standardized methods must be applied across laboratories. Proficiency panels are a key part of a comprehensive quality assurance program to monitor inter- and intralaboratory performance, as well as assay performance, over time. Seven International AIDS Vaccine Initiative-sponsored trial sites participated in the proficiency panels described in this study. At each laboratory, two operators independently processed identical sample sets consisting of frozen peripheral blood mononuclear cell (PBMC) samples from different donors by using four blind stimuli. PBMC recovery and viability after overnight resting and the IFN-gamma ELISPOT assay performance were assessed. All sites demonstrated good performance in PBMC thawing and resting, with a median recovery of 78% and median viability of 95%. The laboratories were able to detect similar antigen-specific T-cell responses, ranging from 50 to >3,000 spot-forming cells per million PBMC. An approximate range of a half log in results from operators within or across sites was seen in comparisons of antigen-specific responses. Consistently low background responses were seen in all laboratories. The results of these proficiency panels demonstrate the ability of seven laboratories, located across three continents, to process PBMC samples and to rank volunteers with differential magnitudes of IFN-gamma ELISPOT responses. These findings also illustrate the ability to standardize the IFN-gamma ELISPOT assay across multiple laboratories when common training methods, reagents such as fetal calf serum, and standard operating procedures are adopted. These results are encouraging for laboratories that are using cell-based immunology assays to test HIV vaccines and other vaccines.
Assuntos
Vacinas contra a AIDS/imunologia , HIV/imunologia , Imunoensaio/métodos , Interferon gama/biossíntese , Leucócitos Mononucleares/imunologia , Sobrevivência Celular , Células Cultivadas , Humanos , Imunoensaio/normas , Variações Dependentes do Observador , Reprodutibilidade dos Testes , Manejo de Espécimes/métodosRESUMO
The halotolerance of a cold adapted alpha-amylase from the psychrophilic bacterium Pseudoalteromonas haloplanktis (AHA) was investigated. AHA exhibited hydrolytic activity over a broad range of NaCl concentrations (0.01-4.5 M). AHA showed 28% increased activity in 0.5-2.0 M NaCl compared to that in 0.01 M NaCl. In contrast, the corresponding mesophilic (Bacillus amyloliquefaciens) and thermostable (B. licheniformis) alpha-amylases showed a 39 and 46% decrease in activity respectively. Even at 4.5 M NaCl, 80% of the initial activity was detected for AHA, whereas the mesophilic and thermostable enzymes were inactive. Besides an unaltered fluorescence emission and secondary structure, a 10 degrees C positive shift in the temperature optimum, a stabilization factor of >5 for thermal inactivation and a Delta T(m) of 8.3 degrees C for the secondary structure melting were estimated in 2.7 M NaCl. The higher activation energy, half-life time and T(m) indicated reduced conformational dynamics and increased rigidity in the presence of higher NaCl concentrations. A comparison with the sequences of other halophilic alpha-amylases revealed that AHA also contains higher proportion of small hydrophobic residues and acidic residues resulting in a higher negative surface potential. Thus, with some compromise in cold activity, psychrophilic adaptation has also manifested halotolerance to AHA that is comparable to the halophilic enzymes.