Fifty-five isolates of nine species and two species varieties (5 isolates each) of the Aspergillus niger group were tested for progesterone transformation. Generally, all isolates of the different species transformed progesterone to 21-hydroxyprogesterone. In addition, the isolates of seven species have the capacity to hydroxylate progesterone to 11 alpha-hydroxyprogesterone. A systematic variation could be observed between the tested species of the Aspergillus niger group with respect to the transformation reactions of progesterone. Comparative biotransformation results showed that essential differences exist between the species of this group, where the different isolates of individual species always produce one or more identical products. This biochemical differentiation may supplement the morphological criteria used in the classification of this group of fungi.