Development of high-efficiency spawning methods and gonadal stress responses in tropical sea cucumber Holothuria leucospilota

Holothuria leucospilota, a commercially significant sea cucumber species, demands improved aquaculture methods for sustainable production. Here, we present a dual breakthrough in spawning induction methodology and the mechanistic understanding of reproductive regulation. First, we developed a novel induction method combining low temperature air-desiccation (16 °C, 30 min) with chemical hypoxia (0.1 g L⁻¹ Na₂SO₃, 30 min), achieving 70–100 % spawning success (5.0–6.7 million eggs female⁻¹) within 1–2 h post-stimulus in this species, using spawning induction rate and total/mean egg output as evaluation indicators. Multi-omics integration revealed sex-specific regulatory mechanisms in H. leucospilota: in females, cell cycle activation likely drove oogenesis and ovulation through adenosine depletion-mediated meiotic resumption, while males appeared to exhibit neuromodulated spermatogenesis via caffeine/dopamine signaling. Crucially, we identified phospholipid-mediated membrane stabilization as a potential response to oxidative stress during gamete release. These findings establish the first comprehensive framework for H. leucospilota reproductive physiology, providing actionable targets for aquaculture innovation.