Functional Spermatogenesis Across Testicular Developmental Stages in Neomale Large Yellow Croaker (Larimichthys crocea) Revealed by Histology and Gonadal Specific Cellular Markers

Gonadal development and spermatogenesis critically influence fish reproductive performance. Neomales (genetically female but functionally male) are indispensable for generating all-female populations, yet their spermatogenesis remains understudied. In the present study, we systematically investigated gonadal maturation in neomales of the large yellow croaker (<i>Larimichthys crocea</i>), an economically important marine species exhibiting sexually dimorphic growth. We examined the growth performance and gonadal development throughout the maturation process in neomales and control males. Results showed comparable growth performance but a temporal divergence in gonadal development: the gonadosomatic index (GSI) of neomales was significantly higher than control males at 400 and 430 days post-hatching (dph), but not at 460 dph during the reproductive period. Histological, ultrastructural (TEM), and immunofluorescence analyses collectively demonstrated that neomale testes contained all major spermatogenic cell types. Their morphological characteristics and expression patterns of key markers-germ cells (<i>vasa</i>), Sertoli cells (<i>sox9a</i>), and meiotic recombination (<i>dmc1</i>)-were similar to control males. These findings enhance understanding of gonadal development and spermatogenesis in neomales, providing a theoretical and technical foundation for large-scale production of all-female large yellow croaker.