Supplementary file 1_Sex-specific hormone signatures in four fish species with divergent reproductive strategies.docx

We additionally included the female-pregnant livebearing fish Heterandria formosa as a comparative reference for a conventional separation of reproductive roles.</p>Results<p>Sex-specific endocrine profiles broadly followed sexual selection differentials: species with stronger sexual heteromorphism, i.e., more extreme reproductive roles (N. ophidion), exhibiting stronger endocrine sexual heteromorphism. In contrast, species with higher parental care investment (H. abdominalis, H. formosa) show reduced endocrine differentiation between the sexes.

The intermediate species S. typhle, which combines sex-role reversal with a moderately complex brood pouch, displayed tissue-specific patterns, with hormonal dimorphism retained primarily in the gonads.</p>Discussion<p>By incorporating a cross-species comparative framework with both sex-role reversed and a conventional viviparous species, we can begin to understand which hormonal functions are conserved versus derived, and whether endocrine profiles are primarily aligned with biological sex or reproductive role, such as mate competition and parental care, thereby revealing how endocrine constraints and plasticity contribute to reproductive evolution.</p>