In 2001, Corynebacterium freneyi was described as a new fermentative, alpha-glucosidase-positive Corynebacterium species related to C. xerosis based on data from three strains. During a review of our extensive culture collection we encountered 18 additional C. freneyi strains and further characterized them in detail. Thirteen of the 18 strains were isolated from female genital tract specimens without any obvious disease association. Phenotypically, C. freneyi can be easily differentiated from C. xerosis by its distinct wrinkled colonies whereas nearly all other routinely applied phenotypic tests do not allow a unanimous separation of C. freneyi from C. xerosis. Restriction length polymorphism analysis using CfoI of the 16S-23S rRNA gene intragenic spacer definitively allows differentiation between the two species. Surprisingly, comparative 16S rRNA gene analysis does not discriminate between C. freneyi and C. xerosis because the designated type strain of C. freneyi is not the most representative strain for this species. The present report also includes detailed data on the antimicrobial susceptibility pattern of C. freneyi presented here for the first time. Based on the large number of additional C. freneyi strains from our culture collection, we provide an extended and emended species description of C. freneyi.