Further characterization of these genes allowed the development of a model for biosynthesis of DAP in mycobacteria. The asd gene, encoding aspartate semialdehyde dehydrogenase, was chosen as a candidate for the development of an in vivo selection system for mycobacteria. The nucleotide sequences of the M. smegmatis and BCG asd genes were determined, demonstrating that the mycobacterial aspartokinase gene was upstream of, and possibly within an operon with, asd . Methods were developed for the use of generalized transduction and intrachromosomal recombination to construct these mutants as well. This insertion element has since proven to be a useful tool for genetic analysis of mycobacteria and the study of mechanisms of pathogenesis.
Environmental mycobacteria are a common cause of human infections.
Recently, contaminated domestic water supplies have been suggested as a potential source for several of these mycobacterial diseases. We found that these mycobacterial species can enter and replicate within A. castellanii . The interaction of M. avium with these amoebae was examined in more detail using light and electron microscopy. The intracellular life-cycle of M. avium in amoebae appears similar to that observed in macrophages. The effect of growth of M. avium in amoebae on virulence was investigated using in vitro and in vivo models of infection.