Physiological and molecular effects of 17α-methyltestosterone and letrozole on sexual differentiation of black rockfish (Sebastes schlegelii)

Sex-controlled breeding holds significant value in aquaculture. By controlling the sex of fish, faster-growing sexes can be preferentially produced, shortening the rearing time to market size. Sebastes schlegelii exhibits a significant female growth advantage, and thus all-female aquaculture can substantially improve economic benefits. The objective of this study was to induce XX pseudo-males in S. schlegelii using exogenous 17α-methyltestosterone (MT) and letrozole (LZ) for all-female breeding, and to clarify the underlying hormonal and transcriptomic mechanisms. S. schlegelii juveniles before sex differentiation were treated with MT at concentrations of 20, 40, 60 mg/kg and LZ at concentrations of 300, 600, 900 mg/kg for 30 and 40 days, respectively. Phenotypic and histological analysis showed that all treatment groups achieved 100 % sex reversal efficiency, with pseudo-males successfully induced. Growth performance results indicated that the 30-day treatment with MT (20 mg/kg) or LZ (300 mg/kg) had no significant difference from the untreated control group, while higher concentrations and longer treatment durations inhibited growth and testicular development. Endocrinology showed that the serum steroid levels (elevated testosterone and 11-ketotestosterone, reduced estradiol) of pseudo-males were closer to those of normal males. Transcriptome analysis showed that the downregulation of genes related to ovarian development ( cyp19a1a , foxl2 ) and the upregulation of genes related to testicular development ( amh , sox9 , dmrt1 ) jointly regulated sex reversal, and the pseudo-males induced by both hormones had enriched steroid hormone biosynthesis and TGF-β signaling pathways. This study establishes a feasible sex reversal strategy in S. schlegelii and provides molecular insights into gonadal differentiation, offering a technical basis for all-female breeding. • 17α-methyltestosterone and letrozole efficiently induced pseudo-males in Sebastes schlegelii . • Spermatogenesis was observed in hormone-induced pseudo-males. • Hormone-induced pseudo-males exhibited male-typical steroid hormone profiles. • Transcriptomic analysis revealed suppression of ovarian genes and activation of testis genes. • Sex reversal involved steroid biosynthesis and TGF-β signaling pathways.