Forty-three strains belonging to the genus Brevibacterium which were encountered in clinical materials over 2 decades were compared with reference strains, including the type strains, of B. casei, B. epidermidis, B. mcbrellneri, B. iodinum, and B. linens. By means of carbohydrate assimilation tests (CATs) the 43 clinical isolates could be assigned to the species B. casei (n = 41) and B. epidermidis (n = 2). DNA-DNA hybridizations were performed for 20 clinical isolates and confirmed the species identification of the isolates. Cellular fatty acid profiles of all strains were determined and found to have less discriminative power than CATs. This is the first report indicating that most clinical Brevibacterium isolates are B. casei and that CATs provide an easy-to-perform method for species determination within the genus, thus avoiding nucleic acid techniques.