The level of amplification (copy number/cell) of HPV16 and HPV18 viral genomes and its correlation with the presence of E1/E2 genes were analyzed in a sample of 42 HPV16- and 21 HPV18-positive cervical carcinomas of different clinical stages and histological types. The viral copy number/cell was assessed by dot-blot hybridization and the presence of E1/E2 genes by PCR and Southern blot. The copy number/cell was significantly lower in HPV18-positive than in HPV16-positive tumours (23 +/- 8 and 457 +/- 191 respectively). Nearly half of the HPV16s (43%) were distributed similarly to the HPV18s in the ranges of 50 or less copies, having its peak at the group of 1 to 10 copies, whereas the remaining HPV16s (57%) spread over the groups of 51 or more copies, with another peak at the group of 101 to 500. The E1/E2 region was absent in all tumours positive for HPV18 and present in 64% of those positive for HPV16. The HPV16 tumours negative for E1/E2 had a much lower viral copy number (17 +/- 12) than the positive ones (582 +/- 212), thus resembling HPV18-positive tumours. Viral copy number was negatively correlated with the clinical stage of the tumours and directly associated with the degree of histological differentiation. However, these correlations are primarily attributable to the presence or absence of an intact E1/E2 region.

2,3-Di-O-acyl-trehalose (DAT) is a glycolipid located on the outer layer of the Mycobacterium tuberculosis cell envelope. Due to its noncovalent linkage to the mycobacterial peptidoglycan, DAT could easily interact with host cells located in the focus of infection. The aim of the present work was to study the effects of DAT on the proliferation of murine spleen cells. DAT was purified from reference strains of M. tuberculosis, or M. fortuitum as a surrogate source of the compound, by various chromatography and solvent extraction procedures and then chemically identified. Incubation of mouse spleen cells with DAT inhibited in a dose-dependent manner concanavalin A-stimulated proliferation of the cells. Experiments, including the propidium iodide exclusion test, showed that these effects were not due to death of the cells. Tracking of cell division by labeling with 5,6-carboxyfluorescein diacetate succinimidyl ester revealed that DAT reduces the rounds of cell division. Immunofluorescence with an anti-CD3 monoclonal antibody indicated that T lymphocytes were the population affected in our model. Our experiments also suggest that the extent of the suppressive activity is strongly dependent on the structural composition of the acyl moieties in DATs. Finally, the inhibitory effect was also observed on antigen-induced proliferation of mouse spleen cells specific for Toxoplasma gondii. All of these data suggest that DAT could have a role in the T-cell hyporesponsiveness observed in chronic tuberculosis.Mycobacterium tuberculosis, the infectious agent of tuberculosis, is responsible for more deaths than any other single pathogen. It causes 2 to 3 million deaths annually and accounts for more than 30% of the deaths of human immunodeficiency virus-positive individuals (13). Factors affecting the pathogenesis of tuberculosis are complex and poorly defined; however, it is well established that the major common feature in chronic tuberculosis is the suppression of the T-cell immune response (3). For instance, while immune depression and immune activation are simultaneously present in tuberculosis, more profound defects in the cell-mediated immunity, which is largely responsible for protection, are clearly correlated with more extensive tissue damages (43).M. tuberculosis synthesizes both stimulatory and suppressive components for T cells. In general, it is accepted that hostmycobacterium interactions are mediated primarily by specialized molecules expressed on the mycobacterial cell envelope. The immunosuppressive capability of M. tuberculosis is attributed at least in part to lipoarabinomannan (LAM), a major cell wall-associated lipoglycan (21,28,29). However, the M. tuberculosis cell wall contains many other distinctive and chemically unusual components, with a predominance of lipid molecules (12). According to data obtained from M. tuberculosis and M. bovis BCG strains, lipid molecules from the cell wall of mycobacteria can migrate outside from the phagocytic vacuole (1, 5). As a consequence, glycolipids noncovalently lin...

No abstract

Cell-surface saccharides of Mycobacterium tuberculosis appear to be crucial factors in tuberculosis pathogenicity and could be useful antigens in tuberculosis immunodiagnosis. In the present study, we report the successful antigenic and immunogenic mimicry of mannose-containing cell-wall compounds of M. tuberculosis by dodecamer peptides identified by phage-display technology. Using a rabbit antiserum raised against M. tuberculosis cell-surface saccharides as a target for biopanning, peptides with three different consensus sequences were identified. Phage-displayed and chemically synthesized peptides bound to the anticarbohydrate antiserum. Rabbit antibodies elicited against the peptide QEPLMGTVPIRAGGGS recognize the mannosylated M. tuberculosis cell-wall antigens arabinomannan and lipoarabinomannan, and the glycosylated recombinant protein alanine/proline-rich antigen. Furthermore, antibodies were also able to react with mannan from Saccharomyces cerevisiae, but not with phosphatidylinositol dimannosides or arabinogalactan from mycobacteria. These results suggest that the immunogenic peptide mimics oligomannosidic epitopes. Interestingly, this report provides evidence that, in contrast with previously known carbohydrate mimotopes, no aromatic residues are necessary in a peptide sequence for mimicking unusual glycoconjugates synthesized by mycobacteria. The possible usefulness of the identified peptide mimotopes as surrogate reagents for immunodiagnosis and for the study of functional roles of the native non-peptide epitopes is discussed.

No abstract