In vitro activity of ciprofloxacin, sparfloxacin, ofloxacin, amikacin and rifampicin against Ghanaian isolates of Mycobacterium ulcerans.

MICs of ciprofloxacin, sparfloxacin, ofloxacin, amikacin and rifampicin were determined for 14 primary clinical isolates and three reference isolates of Mycobacterium ulcerans by modifying a standard agar dilution method for testing Mycobacterium tuberculosis sensitivity. All these antimicrobials were active against every isolate of M. ulcerans. Sparfloxacin exhibited the highest activity and ofloxacin was the least effective. Rifampicin exhibited the broadest range of activity.

Functions and specificity of T cells following nucleic acid vaccination of mice against Mycobacterium tuberculosis infection.

The 38-kDa glycolipoprotein of Mycobacterium tuberculosis has been known to evoke prominent T cell and Ab responses in patients with active tuberculosis. In this study, we investigated its protective capacity using plasmid DNA immunization in a mouse experimental model. Prior knowledge of several antigenic determinants has been beneficial for analyzing the phenotype and specificity of T cells, which determine the efficacy of this vaccination procedure. C57BL/6 mice responded to the 38-kDa gene-pcDNA3 plasmid with strong CD4+ Th1 and CD8+ cytotoxic T cell responses of the IFN-gamma-producing Tc1 phenotype. After challenge with virulent tubercle bacilli, the bacterial load in the spleens and lungs of vaccinated mice was reduced to a level similar to that imparted by Mycobacterium bovis Bacille Calmette-Guérin vaccination. Notably, the specificity of CD4+ and CD8+ T cells from DNA-vaccinated and tubercle-infected mice was found to be strikingly different in respect of several peptide epitopes. The identified peptides recognized by T cells from protected mice are of further interest for the development of subunit-based vaccines against tuberculosis.

Duplication of genes encoding the immunodominant 38 kDa antigen in Mycobacterium intracellulare.

Mycobacterium avium is a causative agent of mycobacterioses in systemically immunocompromised individuals, whereas Mycobacterium intracellulare is responsible for causing infections in relatively immunocompetent hosts. In an attempt to identify components that could be involved in virulence, we characterised the 38 kDa-encoding gene of M intracellulare that is absent in M. avium. This antigen cross reacts immunologically with a major 38 kDa antigen of M. tuberculosis, and both antigens are homologues of the phosphate transport subunit S (PstS) of the pst complex of Escherichia coli. Unlike the M. tuberculosis complex the M. intracellulare coding gene was found to be duplicated. We also identified and characterised other pst genes that may constitute an operon. Considering that multiple isoforms of PstS are present in mycobacteria the possible role of pstS1 genes for pathogenesis is discussed.

Evidence of selection for protein introns in the recAs of pathogenic mycobacteria.

Protein introns are recently discovered genetic elements whose intervening sequences are removed from a precursor protein by an unusual protein splicing reaction. This involves the excision of a central spacer molecule, the protein intron, and the religation of the amino- and carboxy-terminal fragments of the precursor. The recA gene of Mycobacterium tuberculosis contains one such element and we now show that the other major mycobacterial pathogen, Mycobacterium leprae, also possesses a protein intron in its recA, although other mycobacterial recA genes do not. However, these two protein introns are different in size, sequence and location of insertion of their coding sequences into the recAs of M. tuberculosis and M. leprae, indicating that acquisition of the protein introns has occurred independently in the two species, and thus suggesting that there has been selection for splicing in the maturation of RecA in the pathogenic mycobacteria. The M. leprae protein intron provides an example of conditional protein splicing, splicing occurring in M. leprae itself but not when expressed in Escherichia coli, unlike most previously described protein introns. These observations suggest that protein introns may perform a function for their host, rather than being just selfish elements.

Identification, sequencing, and expression of Mycobacterium leprae superoxide dismutase, a major antigen.

The gene encoding a major 28-kilodalton antigen of Mycobacterium leprae has now been sequenced and identified as the enzyme superoxide dismutase (SOD) on the basis of the high degree of homology with known SOD sequences. The deduced amino acid sequence shows 67% homology with a human manganese-utilizing SOD and 55% homology with the Escherichia coli manganese-utilizing enzyme. The gene is not expressed from its own promoter in E. coli but is expressed from its own promoter in Mycobacterium smegmatis. The amino acid sequences of epitopes recognized by monoclonal antibodies against the 28-kilodalton antigen have been determined.