Use of multilocus enzyme electrophoresis to examine genetic relationships amongst isolates of Mycobacterium intracellulare and related species

School of Veterinary Studies, Murdoch University, Murdoch, Western Australia 6150, Australia Australian Reference Laboratory for Bovine Tuberculosis, Agriculture Western Australia, South Perth, Western Australia 6151, Australia State Health Laboratory, Brisbane, Queensland 4000, Australia Author for correspondence: David J. Hampson. Tel: +61 9 360 2287. Fax: +61 9 310 4144.e-mail: hampson@numbat.murdoch.edu.au. ABSTRACT As part of a larger study investigating diversity and distribution of Mycobacterium spp. in Australia, multilocus enzyme electrophoresis was used to assess genetic relationships at 17 enzyme loci amongst a collection of reference strains and isolates initially identified on biochemical and other grounds as M. intracellulare (70), "X" mycobacteria (10), M. scrofulaceum (7), M. avium (8) and M. avium subsp. paratuberculosis (2). Two of the isolates initially identified as M. intracellulare were shown to be quite distinct from the others. Both gave negative results in a species-specific DNA probe test, whilst one was positive by PCR. These results emphasize the uncertainties involved in identifying members of this group. The other M. intracellulare isolates formed a cohesive but diverse group, being divided into 48 electrophoretic types (ETs), with a mean genetic diversity of 0.38. Forty-three of these ETs contained only single isolates. There was no clear relationship between the serovar and ET designation. The index of association calculated for M. intracellulare was significantly different from zero, suggesting that it is a clonal species. PFGE was also applied to selected isolates from the ETs containing multiple isolates, and some of these could be differentiated further. The strains of M. scrofulaceum and ‘X' mycobacteria were distinct from M. intracellulare , but themselves were highly heterogeneous, with mean genetic diversities of 0.66 and 0.65, respectively. Each of these groups may represent more than one species. M. avium strains were distinct from the two M. avium subsp. paratuberculosis strains, as well as from the other mycobacteria studied. Keywords: Mycobacterium intracellulare , multilocus enzyme electrophoresis, genetic diversity, pulsed field gel electrophoresis