Strains of a new species of rapidly growing, nonphotochromogenic mycobacteria, Mycobacterium mageritense, were isolated from human sputum. The growth characteristics, acid fastness, and mycolic acids of the isolates were consistent with those of Mycobacterium species. The isolates were identified as members of a new species by performing a biochemical analysis and DNA-DNA hybridization experiments, and by comparing the sequences of several conserved genes, such as the 16S rRNA, hsp65, and sodA genes. A phylogenetic analysis in which 16S rRNA and sodA sequences were used identified M. mageritense as a novel distinct species and placed M. mageritense between members of the Mycobacterium fortuitum complex and the thermotolerant rapidly growing group. Our results demonstrate that the taxonomic value of sodA sequence analysis in the genus Mycobacterium is similar to the well-established value of 16S rRNA sequence analysis.