Comprehensive proteomic profiling of the membrane constituents of a Mycobacterium tuberculosis strain

Mycobacterium tuberculosis is an infectious microorganism that causes human tuberculosis. The cell membranes of pathogens are known to be rich in possible diagnostic and therapeutic protein targets. To compliment the M. tuberculosis genome, we have profiled the membrane protein fraction of the M. tuberculosis H37Rv strain using an analytical platform that couples one-dimensional SDS gels to a microcapillary liquid chromatography-nanospray-tandem mass spectrometer. As a result, 739 proteins have been identified by two or more distinct peptide sequences and have been characterized. Interestingly, approximately 450 proteins represent novel identifications, 79 of which are membrane proteins and more than 100 of which are membrane-associated proteins. The physicochemical properties of the identified proteins were studied in detail, and then biological functions were obtained by sorting them according to Sanger Institute gene function category. Many membrane proteins were found to be involved in the cell envelope, and those proteins with energy metabolic functions were also identified in this study.     

Antigen recognition by serum antibodies in non-human primates experimentally infected with Mycobacterium tuberculosis

Tuberculosis is a significant threat to non-human primates and their caretakers. The diagnosis of tuberculosis in living non-human primates is currently based on the tuberculin skin test, which is cumbersome and sometimes inaccurate. Development of an accurate serodiagnostic test requires identification of the key antigens of Mycobacterium tuberculosis involved in antibody production. When sequential serum samples obtained from 17 cynomolgus, rhesus, and African green monkeys up to seven months since experimental infection with M. tuberculosis Erdman were screened for antibody against purified proteins of M. tuberculosis, three highly seroreactive antigens were identified. One protein, ESAT-6, reacted with sera from all infected animals. Two additional proteins, alpha-crystallin and MTSA-10, were recognized by sera from approximately 90% of infected animals. Time course analysis of antibody production indicated that the earliest response was usually to ESAT-6 alone or to ESAT-6 and other antigen(s). These results provide experimental evidence of the potential value of ESAT-6 as an antigen for use in serodiagnosis of tuberculosis in non-human primates.     

Mycobacterium avium complex and Mycobacterium tuberculosis in patients infected with the human immunodeficiency virus.

Primary care physicians play an important role in identifying and treating bacterial infections in adults infected with the human immunodeficiency virus (HIV). Mycobacterium avium complex and Mycobacterium tuberculosis are pathogens that can cause systemic or local infection in these patients. We review the epidemiology, pathogenesis, clinical presentation, and principles of treatment for these two mycobacterial pathogens. Because M tuberculosis disease is preventable and curable and yet communicable, physicians should maintain a high degree of suspicion for tuberculosis in HIV-infected adults. In comparison, the goal of treating M avium complex in patients with advanced HIV disease is to reduce constitutional symptoms and improve survival.     

Molecular methods for Mycobacterium tuberculosis strain typing: a users guide

There are now a wide range of techniques available to type Mycobacterium tuberculosis, the problem is to choose the correct technique. For large scale epidemiological studies the portability and standardization of IS6110 restriction fragment length polymorphism (RFLP) means that this remains the gold standard technique. In the next few years the internationally standard mycobacterial interspersed repetitive unit (MIRU) may come to challenge this primacy. Low copy number stains remain a problem and these can be typed by either polymorphic Guanine cytosine-rich repetitive sequence (PGRS) or MIRU-variable numbers of tandem repeat (VNTR). To confirm whether strains are part of a true cluster PGRS remains the method of choice. For local outbreaks and investigations of laboratory cross contamination where speed is of greatest importance suspect strains should be initially investigated using a PCR-based method. The superior reproducibility and discrimination of MIRU-VNTR means that these methods should be favoured. If matches are found, then further confirmation of identity can be achieved using IS6110 RFLP or PGRS if the strains prove to have a low IS6110 copy number.     

Characterization of a novel antigen of Mycobacterium tuberculosis K strain and its use in immunodiagnosis of tuberculosis

Mycobacterium tuberculosis-specific antigens would be of great value in developing immunodiagnostic tests for tuberculosis (TB), but regional differences in molecular types of the organism may result in antigenic variation, which in turn affects the outcome of the tests. For example, the Beijing strains of M. tuberculosis are prevalent in East Asia, and in particular, the K strain and related strains of the Beijing family, are most frequently isolated during school outbreaks of TB in South Korea. From comparison of genome sequences between M. tuberculosis K strain and the H37Rv strain, a non-Beijing type, we identified a K strain-specific gene, InsB, which has substantial homology with the ESAT-6-like proteins. This study was, therefore, initiated to characterize the InsB protein for its immunogenicity in mice and to confirm its expression in TB patients by detecting antibodies to the protein. The InsB gene was cloned from M. tuberculosis K strain and expressed in Escherichia coli. The recombinant InsB protein was used for immunization of mice. All mice showed strong antibody responses to the InsB protein, and splenocytes stimulated with InsB showed strong IFN-γ and IL-17 responses and a weak IL-2 response, all of which have been implicated in disease expression and used for the immunodiagnosis of TB. Serum samples from TB patients also showed significant antibody responses to the InsB protein as compared to healthy control samples. These results indicate that the InsB protein is an M. tuberculosis K-strain-specific antigen that could further improve the current immunodiagnostic methods, especially for the South Korean population.     

小鼠结核分枝杆菌耐药模型的建立与评价

目的 建立小鼠结核分枝杆菌耐利福平株静脉感染小鼠模型.方法 30只BALB/c雌性小鼠经尾静脉感染结核分枝杆菌耐利福平株每只106 CFU,观察小鼠一般状况,并分别在感染后6周、10周、14周处死小鼠,进行脾、肺组织病理切片、抗酸染色、计脏器荷菌数.结果 感染后小鼠体重呈逐渐上升趋势,脏器病理变化随时间推移由急性炎症逐...     

On the Ornithinyl Ester of Phosphatidylglycerol of Mycobacterium 607

Ornithinyl ester of phosphatidyl glycerol was found to accumulate in Mycobacterium 607 under acidic conditions (pH 5.6) of growth or in cultures of ultraviolet-irradiated (320 to 420 nm) bacilli. There was a corresponding decrease in cardiolipin content of the organisms under these conditions.     

Mycobacterium tuberculosis and Mycobacterium avium modify the composition of the phagosomal membrane in infected macrophages by selective depletion of cell surface-derived glycoconjugates

The growth of pathogenic mycobacteria in phagosomes of the host cell correlates with their ability to prevent phagosome maturation. The underlying molecular mechanism remains elusive. In a previous study, we have shown that Mycobacterium avium depletes the phagosome membrane of cell surface-derived glycoconjugates (de Chastellier and Thilo, Eur. J. Cell Biol. 81, 17-25, 2002). We now extended these quantitative observations to the major human pathogen, Mycobacterium tuberculosis (H37Rv). At increasing times after infection of mouse bone marrow-derived macrophages, cell-surface glycoconjugates were labelled enzymatically with [3H]galactose. Subsequent endocytic membrane traffic resulted in a redistribution of this label from the cell surface to endocytic membranes, including phagosomes. The steady-state distribution was measured by quantitative autoradiography at the electron microscope level. Relative to early endosomes, with which phagosomes continued to fuse and rapidly exchange membrane constituents, the phagosome membrane was depleted about 3-fold, starting during infection and in the course of 9 days thereafter. These results were in quantitative agreement with our previous observations for Mycobacterium avium. For the latter case, we now showed by cell fractionation that the depletion was selective, mainly involving glycoproteins in the 110-210 kDa range. Together, these results indicated that pathogenic mycobacteria induced and maintained a bulk change in phagosome membrane composition that could be of special relevance for survival of pathogenic mycobacteria within phagosomes.