In the tilapia Oreochromis niloticus, sex is determined genetically (GSD), by temperature (TSD) or by temperature/genotype interactions. Functional masculinization can be achieved by applying high rearing temperatures during a critical period of sex differentiation. Estrogens play an important role in female differentiation of non-mammalian vertebrates. The involvement of aromatase, was assessed during the natural (genetic all-females and all-males at 27°C and temperature-induced sex differentiation of tilapia (genetic all-females at 35°C). Gonads were dissected between 486-702 degree x days. Aromatase gene expression was analyzed by virtual northern and semi-quantitative RT-PCR revealing a strong expression during normal ovarian differentiation concomitant with high levels (465±137 fg/g) of oestradiol-17ß (E2-17ß). This was encountered in gonads after the onset of ovarian differentiation (proliferation of both stromal and germ cells prior to ovarian meiosis). Genetic males exhibited lower levels of aromatase gene expression and E2-17ß quantities (71±23 fg/g). Aromatase enzyme activity in fry heads established a sexual dimorphism in the brain, with high activity in females (377.9 pmol/head/hr) and low activity in males (221.53 pmol/head/hr). Temperature induced the masculinization of genetic females to a different degree in each progeny, but in all cases repression of aromatase expression was encountered. Genetic males at 35°C also exhibited a repression of aromatase expression. Aromatase brain activity decreased by nearly three-fold in the temperature-masculinized females with also a reduction observed in genetic males at 35°C. This suggests that aromatase repression is required in the gonad (and perhaps in the brain) in order to drive differentiation towards testis development.