Genomic Relatedness among Mycobacterial Species by Nonisotopic Blot Hybridization

A technique to screen genomic relatedness among mycobacterial species was developed by using a nonisotopic deoxyribonucleic acid (DNA) blot hybridization method. Total genomic DNAs prepared from type strains of 26 species of Mycobacterium, armadillo-derived Mycobacterium leprae, and M. lepraemurium were used as fixed-phase DNAs on nitrocellulose filters and were hybridized with biotin-labeled genomic DNAs of test strains as probes. The majority of the type strains were distinguishable from each other by this method. However, blot hybridization showed high levels of DNA relatedness between M. avium and M. paratuberculosis, between M. marinum and M. ulcerans, and also between M. scrofulaceum and M. xenopi, and two other methods (spectrophotometric assay and the SI nuclease method) confirmed that these three pairs of species were more than 80% genomically related. Other pairs of species with phenotypically similar taxonomic matrices, such as M. avium-M. intracellulare, M. gastri-M. kansasii, M. asiaticum-M. szulgai, and M. nonchromogenicum-M. terrae, were readily differentiated from each other by the blot hybridization method. Unclassified “lufu” strains showed a very high level of genomic relatedness with M. asiaticum, which was confirmed by spectrophotometric and S1 nuclease methods.