Early differentiation in rainbow trout gonads was investigated by expression profiling and in situ hybridization (ISH). Expression of cyp19al and fst in females and sox9al in males were sexually dimorphic between 32 to 35 days post-fertilization (dpf). After 35 dpf, the differentiation proceeded with sexually dimorphic profiles for sox9a2, dmrt1, cyp11b2.1, amh in males and foxl2a, foxl2b, hsd3bl, inha in females. cyp17a1, cyp11a1, star, nr5a1b increased only after 40 dpf in both sexes with a slightly higher expression in females. cyp19a1 expression was localized in a cluster of somatic cells in the ventral side of female gonads, and sox9a2 and amh in somatic cells surrounding the germ cells, at 28 dpf and thereafter, both in male and female gonads. cypllb2.1, cyp17a1, and cyp1lal expressions were only detected in scattered somatic cells in males after 46 dpf. This confirms the early implication of cyp19a1 in trout ovarian differentiation and suggests that early testicular differentiation does not need androgen production.