The amplified fragment length polymorphism technique has been applied to identify neurotropic chaetothyrialean black yeasts and relatives from clinical sources. Cladophialophora bantiana, C. emmonsii, C. arxii, C. devriesii, and C. modesta, previously identified on the basis of sequencing and phenotypic and physiological criteria, were confirmed by cluster analysis, demonstrating the clear separation of C. bantiana as a rather homogeneous group from the other species. C. bantiana is a neurotropic fungus causing cerebral abscesses with a mortality of up to 70%. Successful therapy consists of neurosurgical intervention and optimal antifungal therapy. Since the latter is not clearly defined in a large series, we tested the in vitro activities of eight antifungal drugs against clinical isolates of C. bantiana (n = 37), C. modesta (n = 2), C. arxii (n = 1), C. emmonsii (n = 1), and C. devriesii (n = 1), all of which had caused invasive infections. The resulting MIC(90)s for all neurotropic C. bantiana strains were as follows, in increasing order: posaconazole, 0.125 microg/ml; itraconazole, 0.125 microg/ml; isavuconazole, 0.5 microg/ml; amphotericin B, 1 microg/ml; voriconazole, 2 microg/ml; anidulafungin, 2 microg/ml; caspofungin, 4 microg/ml; and fluconazole, 64 microg/ml. On the basis of these in vitro results and the findings of previous clinical and animal studies, posaconazole seems to be a good alternative to the standard treatment, amphotericin B, for C. bantiana cerebral infections. The new agent isavuconazole, which is also available as an intravenous preparation, has adequate activity against C. bantiana.