Differentiation of Mycobacterium tuberculosis strains by use of a nonradioactive Southern blot hybridization method

The only means of dividing strains of Mycobacterium tuberculosis is by phage typing. Attempts at developing a convenient method based on differences in chromosomal DNA by detecting restriction fragment length polymorphisms (RFLP) have had limited success. This report describes the development of a nonradioactive RFLP technique that differs from most methods by using enzymes that have four base recognition sites rather than six. The restriction enzymes AluI, DdeI, HinfI, NdeII, RsaI, and TaqI were used to digest genomic DNA, and high-molecular-weight fragments were visualized after Southern blotting with digoxigenin-labeled M. tuberculosis DNA. Digestion with AluI resulted in different banding patterns among all eight clinical isolates of M. tuberculosis and three type strains from the tuberculosis complex. NdeII distinguished all but two strains, while HinfI and RsaI were unable to distinguish two pairs of strains. Digestion with the enzymes DdeI and TaqI failed to result in clearly discernible bands. These preliminary results suggest that this method provides a fine differentiation of M. tuberculosis strains and may be useful as an epidemiologic tool.