NUCLEOTIDE-SEQUENCE COMPARISON OF THE MYCOBACTERIAL DNAJ GENE AND PCR-RESTRICTION FRAGMENT LENGTH POLYMORPHISM ANALYSIS FOR IDENTIFICATION OF MYCOBACTERIAL SPECIES

We recently reported a genus-specific PCR for the mycobacterial dnaJ g ene. In the present study, we have determined the nucleotide sequences of the dnaJ gene from 19 mycobacterial species (Mycobacterium tubercu losis, M. bovis, M. bovis BCG, M. africanum, M. microti, M. marinum, M . kansasii, M. gastri, M. simiae, M. scrofulaceum, M. szulgai, M. gord onae, M. avium, M. intracellulare, M. xenopi, M. fortuitum, M. chelona e, M. hemophilum, and M. paratuberculosis). On the basis of the amplif ied dnaJ gene nucleotide sequences, we constructed a phylogenetic tree of the mycobacterial species by using the neighbor-joining method and unweighted pairwise grouping method of arithmetic average. We found t hat the phylogenetic relationship inferred within the slowly growing s pecies was in good agreement with the traditional classification, with three major branches corresponding to Runyon's groups I, II, and III. An exception was M. simiae, which was phylogenetically closer to the cluster including members of Runyon's group III than to that of Runyon 's group I. On the other band, the rapid growers, such as M. fortuitum and M. chelonae, did not form a coherent line corresponding to Runyon 's group IV, indicating that our phylogenetic analysis based on the dn aj gene reflects the phenotypic characteristics such as pigmentation b ut not the growth rate. Finally, we revealed the species-specific rest riction sites within the amplified dnaJ gene to differentiate most of the mycobacterial DNA by a combination of PCR with restriction fragmen t length polymorphism analysis.