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The roles of steroids in zebrafish sex differentiation, gonadal development, and function of the adult gonad are poorly understood. Herein, we used ferredoxin 1b (fdx1b) mutant zebrafish to explore such processes. Fdx1b is an essential electron-providing cofactor to mitochondrial steroidogenic enzymes, which are crucial for glucocorticoid and androgen production in vertebrates. Fdx1b−/− zebrafish mutants develop into viable adults in which concentrations of androgens and cortisol are significantly reduced. Adult fdx1b−/− mutant zebrafish display predominantly female secondary sex characteristics but may possess either ovaries or testes, confirming that androgen signaling is dispensable for testicular differentiation in this species, as previously demonstrated in androgen receptor mutant zebrafish. Adult male fdx1b−/− mutant zebrafish exhibit reduced characteristic breeding behaviors and impaired sperm production, resulting in infertility in standard breeding scenarios. However, eggs collected from wild-type females can be fertilized by the sperm of fdx1b−/− mutant males by in vitro fertilization. The testes of fdx1b−/− mutant males are disorganized and lack defined seminiferous tubule structure. Expression of several promale and spermatogenic genes is decreased in the testes of fdx1b−/− mutant males, including promale transcription factor sox9a and spermatogenic genes igf3 and insl3. This study establishes an androgen- and cortisol-deficient fdx1b zebrafish mutant as a model for understanding the effects of steroid deficiency on sex development and reproductive function. This model will be particularly useful for further investigation of the roles of steroids in spermatogenesis, gonadal development, and regulation of reproductive behavior, thus enabling further elucidation of the physiological consequences of endocrine disruption in vertebrates.

Ferredoxin 1b Deficiency Leads to Testis Disorganization, Impaired Spermatogenesis, and Feminization in Zebrafish