Levonorgestrel (LNG) and dydrogesterone (DDG) are two commonly used synthetic progestins that have been detected in aquatic environments. They could affect fish sex differentiation, but the underlying mechanisms remain unknown. Here we investigated the effects of LNG (5 ng L-1 and 50 ng L-1), DDG (100 ng L-1) and their mixtures on gonadal differentiation and sex determination in zebrafish at transcriptomic and histological levels from 2 hours post-fertilization (eleutheroembryos) to 144 days post-fertilization (sexual maturity). Germ cell development and oogenesis pathways were significantly enriched in LNG and the mixture of LNG and DDG treatments, while insulin and apoptosis pathways in the DDG treatment. LNG and the mixture of LNG and DDG strongly decreased transcripts of germ cell development and oogenesis related genes, while DDG increased the transcripts of insulin and apoptosis related genes at 28 days post fertilization (dpf) and 35 dpf. Furthermore, DDG caused ∼ 90% males, and LNG and the mixture of LNG and DDG resulted in 100% males on all sampling dates. Specifically, most males in LNG and the mixture of LNG and DDG treatments were "Type I" males without juvenile oocytes at 28 dpf and 35 dpf, while those in DDG treatment were "Type II" and "Type III" males with a few juvenile oocytes. These results indicated that LNG and DDG promoted testicular differentiation via different pathways to cause male bias. LNG and DDG mixtures have similar effect on testicular differentiation to LNG alone. The findings from this study could have significant ecological implications to fish populations.