A novel slow-growing, non-chromogenic mycobacterium (strain 01-305(T)) was isolated from a patient with pulmonary dysfunction. Growth characteristics, acid-fastness and the results of 16S rRNA gene sequencing supported the placement of this strain within the genus Mycobacterium. Phenotypically, strain 01-305(T) was generally similar to Mycobacterium triviale ATCC 23292(T), but some unique biochemical characteristics were observed. The 16S rRNA gene sequence of strain 01-305(T) was similar to those of M. triviale ATCC 23290 (GenBank accession no. AY734996, 99.9 % similarity) and M. triviale ATCC 23291 (AY734995, 99.9 %); however, it differed substantially from that of M. triviale ATCC 23292(T) (X88924, 98.2 %). Phylogenetic analysis based on 16S rRNA gene sequences placed strain 01-305(T) in the slow-growing Mycobacterium group close to M. triviale ATCC 23290 and M. triviale ATCC 23291, but not M. triviale ATCC 23292(T). Unique mycolic acid profiles and phylogenetic analysis based on two different chronometer molecules, and the hsp65 and rpoB genes, strongly supported the taxonomic status of this strain as representing a distinct species. These data support the conclusion that strain 01-305(T) represents a novel mycobacterial species, for which the name Mycobacterium koreense sp. nov. is proposed. The type strain is 01-305(T) ( = DSM 45576(T) = KCTC 19819(T)).