muramidase and Mycobacterium-Infections

Mycobacterium tuberculosis infects one-third of the world's population and causes two million deaths annually. The unusually low permeability of its cell wall contributes to the ability of M. tuberculosis to grow within host macrophages, a property required for pathogenesis of infection. Mycobacterium marinum is an established model for discovering genes involved in mycobacterial infection. Mycobacterium marinum mutants with transposon insertions in the beta-ketoacyl-acyl carrier protein synthase B gene (kasB) grew poorly in macrophages, although growth in vitro was unaffected. Detailed analyses by thin-layer chromatography, nuclear magnetic resonance (NMR), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, infrared spectroscopy, and chemical degradations showed that the kasB mutants synthesize mycolic acids that are 2-4 carbons shorter than wild type; the defect was localized to the proximal portion of the meromycolate chain. In addition, these mutants showed a significant (approximately 30%) reduction in the abundance of keto-mycolates, with a slight compensatory increase of both alpha- and methoxy-mycolates. Despite these small changes in mycolate length and composition, the kasB mutants exhibited strikingly altered cell wall permeability, leading to a marked increase in susceptibility to lipophilic antibiotics and the host antimicrobial molecules defensin and lysozyme. The abnormalities of the kasB mutants were fully complemented by expressing M. tuberculosis kasB, but not by the closely related gene kasA. These studies identify kasB as a novel target for therapeutic intervention in mycobacterial diseases.

Topics: Animals; Anti-Bacterial Agents; Cell Line; Cell Wall; Cerulenin; Chromatography, Gas; Chromatography, Thin Layer; Colony Count, Microbial; Defensins; DNA Transposable Elements; Drug Resistance, Bacterial; Genetic Complementation Test; Macrophages; Magnetic Resonance Spectroscopy; Mice; Muramidase; Mutagenesis, Insertional; Mycobacterium Infections; Mycobacterium marinum; Mycolic Acids; Permeability; Phagosomes; Rifampin; Sodium Dodecyl Sulfate; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Spectrophotometry, Infrared

Traditionally, protein Ags have been injected in CFA (oil with inactivated mycobacteria) to induce immunity and with IFA (oil alone) to induce tolerance. We report here that injection of hen eggwhite lysozyme, a prototypic Ag, in CFA-induced and IFA-induced pools of hen eggwhite lysozyme-specific memory T cells of comparable fine specificity, clonal size, and avidity spectrum, but with type-1 and type-2 cytokine signatures, respectively. This adjuvant-guided induction of virtually unipolar type-1 and type-2 immunity was observed with seven protein Ags and in a total of six mouse strains. Highly polarized type-1 and type-2 immunity are thus readily achievable through the choice of adjuvant, irrespective of the genetic bias of the host and of the nature of the protein Ag. This finding should have far-reaching implications for the development of vaccines against infectious and autoimmune diseases. Furthermore, our demonstration that Ag injected with IFA is as strongly immunogenic for T cells as it is with CFA shows that the presence of the mycobacteria determines not the priming of naive T cells through the second-signal link but the path of downstream differentiation toward CD4 memory cells that express either type-1 or type-2 cytokines.

Topics: Alum Compounds; Animals; CD4 Antigens; Cytokines; Dose-Response Relationship, Immunologic; Epitopes, T-Lymphocyte; Female; Freund's Adjuvant; Immunity, Cellular; Immunoglobulin G; Immunologic Memory; Injections, Intraperitoneal; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred DBA; Muramidase; Mycobacterium Infections; Th1 Cells; Th2 Cells

A study has been made on mycobacterial-induced granulomas in guinea-pig lymph nodes. Lysozyme and angiotensin-converting enzyme (ACE) were measured in the auricular lymph nodes and serum of guinea-pigs which had received live BCG (Pasteur) or Cobalt (Co)-irradiated armadillo-derived Mycobacterium leprae intradermally into the ear or dinitrofluorobenzene (DNFB) painted epicutaneously upon the ears. In the lymph nodes with granulomas induced by either live BCG or killed M. leprae, the mean concentrations of lysozyme and ACE varied directly with the mean weight of the lymph nodes but the temporal pattern of weight change differed with the two agents. In M. leprae recipients at the time of peak lymph node weight, serum lysozyme and ACE values were significantly greater than those observed in controls; in animals receiving live BCG (Pasteur), serum lysozyme but not ACE values were elevated significantly at the time of peak lymph node weight. Four days following the epicutaneous application of DNFB, where there was no granuloma, there was a similar increase in the concentration of lysozyme and ACE in the lymph nodes. At the same time, there was also significant elevation in the serum lysozyme and ACE concentrations. Thus, in the granulomatous responses, the parallel tissue and serum changes in lysozyme and ACE concentrations were consistent with increased production and secretion of each enzyme by cells of the mononuclear phagocyte series. The increased lysozyme and ACE concentrations found in the lymph nodes of DNFB sensitised animals gives further evidence that such changes are not unique to granulomas. Finally, the intradermal administration of dead M. leprae in guinea pigs also produced increased lysozyme and ACE levels similar to that found in leprosy in man.

Topics: Animals; Female; Granuloma; Guinea Pigs; Leukocyte Count; Lymph Nodes; Male; Muramidase; Mycobacterium Infections; Neutrophils; Peptidyl-Dipeptidase A; Time Factors

Topics: Animals; Armadillos; Blood Proteins; Leprosy; Muramidase; Mycobacterium Infections; Peptidyl-Dipeptidase A; Transcobalamins; Xenarthra

Topics: Acid Phosphatase; Animals; Bacillus; Biochemical Phenomena; Biochemistry; Cathepsins; Macrophages; Macrophages, Alveolar; Muramidase; Mycobacterium bovis; Mycobacterium Infections; Phagocytosis; Rabbits; Research; Seasons

Topics: Acid Phosphatase; Animals; BCG Vaccine; Culture Techniques; Esterases; Histocytochemistry; Immunity; Lipase; Macrophages; Muramidase; Mycobacterium Infections; Peptide Hydrolases; Peritoneal Cavity; Pulmonary Alveoli; Rabbits