Marine teleost fish populations are declining due to anthropogenic factors such as overfishing and climate change, threatening species survival and global food security. Aquaculture has emerged as an efficient alternative to meet rising food demand, showing sustained growth but still facing reproductive challenges. Reproductive biotechnology in fish offers promising strategies, notably the in vitro culture of germ cells (spermatogonia and oogonia), which serve as gamete precursors with self-renewal and proliferative capacity. This project focused on striped bass (Morone saxatilis), a species of aquacultural relevance with well-documented reproductive biology, making it a suitable model for developing cell culture protocols
As a preliminary step toward establishing a long-term germ cell culture system, this study evaluated the effects of fetal bovine serum (FBS), fish serum (FS), and embryo extract (EE) on mitotic proliferation and viability of in vitro cultured spermatogonia and oogonia from M. saxatilis. Germ cells were isolated from ovarian and testicular tissue via enzymatic dissociation (0.3% trypsin, 0.05% DNase), followed by sex identification based on gonadal morphology and cell suspension characteristics. Enrichment was performed using a Percoll gradient and 14-hour differential plating. The effects of 0.3% trypsin and 0.25% EDTA on cell dissociation were compared, assessing cell yield and viability after five days of culture, with no significant differences observed.
Subsequently, the effects of 1% FS and 1% EE (from Pacific yellowtail, Seriola rivoliana), and 20% and 10% FBS (for oogonia and spermatogonia, respectively) were evaluated over 5- and 10-day culture periods. Flow cytometry was used to assess viability (propidium iodide) and proliferation (CFSE). Preliminary results indicate that the combination of 1% FS and 1% EE yielded the most favorable outcomes. Germ cell identification was confirmed via flow
cytometry using a VASA-specific antibody. This study represents the first investigation of in vitro germ cell culture in fish conducted in Mexico