Tubercle and lung disease : the official journal of the International Union against Tuberculosis and Lung Disease最新文献

A panel of antifungal and antihelmintic drugs was tested for activity against Mycobacterium tuberculosis in vitro. Antifungal drugs, miconazole, 2-nitroimidazole, clotrimazole, and the antihelmintic drug niclosamide were found to have significant antituberculosis activity, with minimal inhibitory concentrations (MICs) between 1 and 10μg/ml. Niclosamide and 2-nitroimidazole also had activity against stationary phase tubercle bacilli. Further testing of these drugs and their derivatives in vitro and in vivo appears to be warranted.

Setting: We hypothesized that patients with active pulmonary tuberculosis (TB) have tubercular pneumonitis and that alveolar macrophages at these sites release proinflammatory cytokines, resulting in high levels of cytokines in alveolar epithelial lining fluid.

Objective: To measure cytokine levels in bronchoalveolar lavage fluid (BALF) and to confirm the source of any cytokines by examination of alveolar macrophage cytokine mRNA.

Design: Seventeen active pulmonary TB patients and 15 healthy controls were prospectively studied. Bronchoalveolar lavage (BAL) was performed, proinflammatory cytokine levels were determined and alveolar macrophages isolated from BALF were prepared for RNA extraction and Northern blot analysis.

Results: Compared with healthy controls, TNF-α, IL-1β and IL-6 in BALF were all significantly higher in patients with active pulmonary TB, 298.7±85.9 vs. 8.9±2.7 (P=0.0001); 164.4±67.5 vs. 8.9±2.7 (P=0.003); 969.2±214.2 vs. 86.4±17.0 (mean±SE pg/ml) (P=0.0001), respectively. Only TNF-α and IL-6 levels were significantly higher in sera of active pulmonary TB patients, 92.3±28.7 vs. 3.5±1.2; 15.2±5.4 vs. 2.1±2.1, respectively. Northern blot analysis revealed increased gene expression of these alveolar macrophage cytokines in patients with active pulmonary TB compared healthy controls.

Conclusion: Significantly higher levels of TNF-α, IL-1β and IL-6 were found in BALF from patients with active pulmonary TB, and were released by alveolar macrophages in the TB lesions.

Setting: Molecular Research Laboratory, Department of Medical Microbiology, University of Cape Town and Groote Schuur Hospital.

Objective: Characterize Mycobacterium tuberculosis homologue of theStreptomyces coelicolor , sporulation specific, whiB regulatory gene.

Design: The M. tuberculosis whiB3 gene was isolated by enriched cloning of a 2.8kb Bam HI fragment to which the S. coelicolor whiB gene hybridized. Expression of the gene was analysed by S1 nuclease analysis and promoter studies.

Results: An open reading frame within the 2.8kb Bam HI fragment was identified as the M. tuberculosis whiB3 gene, one of four whiB homologues in the M. tuberculosis genome. The deduced amino acid sequence has a 92% identity with a M. leprae protein, and 32% identity with the S. coelicolor WhiB protein. S1 nuclease analysis showed that the M. tuberculosis whiB3 gene is constitutively expressed by the cells in liquid culture. Primer extension analysis revealed three transcriptional start sites. Expression from the three potential promoters is growth phase-dependent.

Conclusion: The M. tuberculosis whiB3 gene is expressed throughout growth, but expression from the individual promoters is growth phase dependent.

Setting: The granulomatous uveitis, multifocal choroiditis and periphlebitis have been suspected to be of tubercular origin but no definitive reports about detection of etiological agents have been documented in the literature. Conventional bacteriological methods are not generally helpful in diagnosing ocular tuberculosis due to difficulty with potential morbidity associated with obtaining the biopsy material from the eye. Thus, the diagnosis of ocular tuberculosis is most often presumptive.

Objective: We evaluated the role of polymerase chain reaction (PCR) for detection ofMycobacterium tuberculosis in the aqueous humor samples obtained from eyes with active uveitis.

Methods: Aqueous samples from 53 patients having cellular reaction in the anterior chamber along with any one or more of the following: 1) active vasculitis; 2) anterior vitreous cells; 3) snowball opacities; 4) snow banking in the pars plana; 5) retinochoroiditis were withdrawn by anterior chamber paracentesis and subjected to PCR. Seventeen samples from patients with definite clinical diagnoses other than tuberculosis formed a disease control group. Fifteen aqueous samples obtained from healthy subjects undergoing routine cataract surgery served as healthy controls. PCR was performed using primers capable of amplifying a 150 b.p. segment from a conserved repetitive sequence in the genome of M. tuberculosis.

Results: Twenty out of the 53 samples (37.7%) in the study group were positive where as only one sample out of 17 in the disease control group (5.7%) showed a weakly positive band. No sample from the healthy control group showed a positive PCR.

Conclusion: Our study shows that PCR can be effectively used for the diagnosis of intraocular tuberculosis in the presence of uveitis.

Setting: Mannose binding protein gene polymorphism in pulmonary tuberculosis in India.

Objective: To find out whether non-HLA genes such as mannose binding protein (MBP) genes are associated in the susceptibility to pulmonary tuberculosis.

Design: Genotyping of MBP 52, 54 and 57 wild and mutant alleles was carried out in HLA-DR typed pulmonary tuberculosis patients (n=202) and control subjects (n=109). Since HLA-DR2 is associated with pulmonary-TB, the interaction of MBP genes on -DR2 and non-DR2 genes on the susceptibility was also studied.

Results: A significantly increased genotype frequency of MBP functional mutant homozygotes (including 52, 54 and 57) was seen in pulmonary tuberculosis (PTB) patients (10.9%) than in control subjects (1.8%;P=0.008; odds ratio: 6.5). Analysis of interaction of MBP genes and HLA-DR2 on the susceptibility to PTB revealed that these genes are associated with PTB independent of each other.

Conclusion: The present study shows that functional mutants of MBP are associated with PTB. Apart from HLA-DR2 association, association of non-HLA genes in the susceptibility to PTB is evident. This suggests that multigenetic factors (candidate genes) may be involved in the susceptibility/resistance to PTB.

Objective: To determine the efficacy of Ag85A-DNA against challenge with a highly virulent human clinical isolate of Mycobacterium tuberculosis (CSU37) and to compare the potencies of two types of Ag85A-DNA vaccines; those expressing secreted and non-secreted forms of the protein.

Design: Ag85A-DNA vaccinated mice were challenged with a highly virulent clinical isolate of M. tuberculosis (CSU37) in order to compare the efficacy of these vaccines. In vitro studies were also performed.

Results: Enhanced humoral and cellular responses were induced in mice vaccinated with the secreted Ag85A-DNA compared to the non-secreted Ag85A-DNA. In addition, secreted Ag85A-DNA conferred protective immunity against infection with M. tuberculosis (CSU37).

Conclusions: DNA vaccines encodingM. tuberculosis Ag85A have been shown to induce potent humoral and cellular immune responses leading to protection fromM. tuberculosis (Erdman) challenge in mouse models.¹In this study we demonstrate that Ag85A can confer protection in a rigorous challenge model using a highly virulent human clinical isolate of M. tuberculosis (CSU37). This challenge model appears able to discriminate between DNA vaccines of differing potencies, as the more immunogenic DNA construct encoding a secreted form of Ag85A was protective, whereas the less immunogenic DNA construct encoding a non-secreted form of Ag85A was not.

Objective: To test the toxicity of reactive nitrogen intermediates (RNI), including authentic nitric oxide (NO), nitrogen dioxide (NO₂), and peroxynitrite anion (ONOO⁻), a potent oxidant derived from NO and superoxide anion, on various mycobacterial strains including M. tuberculosis.

Design: Relatively avirulent mycobacteria including M. smegmatis and BCG, as well as the pathogenic M. Bovis Ravenel andM. tuberculosis Erdman and the clinical isolate M160 (also known as the C strain) were tested for their susceptibility to the toxic effects of NO, NO₂, and ONOO⁻. Deaerated, NO–saturated solutions as well as an anaerobic in vitro system in which mycobacteria can be exposed to desired concentrations of authentic NO or NO₂, were employed in these studies. An in vitro ONOO⁻killing assay was used to examine the adverse effects of this NO–derived oxidant on the various strains of mycobacteria.

Results: Both NO and NO₂exhibit antimycobacterial activity, with the former being more potent. Results obtained using ONOO⁻killing assay revealed that while avirulent mycobacteria including BCG and M. smegmatis are susceptible to this NO–derived oxidant, the virulent Erdman strain of M. tuberculosis and M. bovis, as well as the clinical tuberculous isolate M160, are remarkably resistant.

Conclusion: These results suggest that the interactions between RNI and various species of mycobactiera could be highly specific. And since activated macrophages produce peroxynitrite, the significance of the ONOO⁻resistance of M. tuberculosis strains in relation to intracellular survival deserves further investigation.

Setting: TGF-β1 has been implicated as an important mediator of immuno-suppression in clinical tuberculosis.

Objective: The objective was to determine the role of TGF-β1 in experimental pulmonary tuberculosis in the guinea pig.

Design: Groups of guinea pigs, maintained on either a low protein (LP) diet or an isocaloric high protein (HP) diet. were challenged via the respiratory route with virulent Mycobacterium tuberculosis H37Rv. Ten days post-infection, guinea pigs were given daily intraperitoneal injections of recombinant human TGF-β1 (rhTGF-β1 τ for 10 consecutive days). Following the treatment, guinea pigs were euthanized, and PPD-induced proliferation of peripheral blood mononuclear cells (PBMCs) was assessed and disease resistance measured by recovery of mycobacteria from the lungs and spleens. In a second set of experiments, groups of HP and LP guinea pigs were vaccinated with attenuated M. tuberculosis H37Ra. Six weeks later, the effects of rhTGF-β1 on lymphoproliferation and cytokine production were determined.

Results: Protein deficiency significantly impaired host anti-tuberculosis resistance, as expected. Treatment with rhTGF-β1 significantly increased mycobacterial loads in the tissues of guinea pigs and decreased the PPD-induced proliferation of PBMCs from both LP and HP guinea pigs. PPD-driven lymphoproliferation, TNF-a and IFN production were significantly suppressed in vaccinated, protein-deficient guinea pigs, and rhTGF-β1 further inhibited lymphoproliferation and cytokine production.

Conclusion: Both in vivo and in vitro results indicate that TGF-β1 exerts immunosuppressive activity and exacerbates the progression of experimental pulmonary tuberculosis in both normally nourished and protein-deficient guinea pigs.