First bivalve sperm vitrification technique developed with the model species dwarf surfclam Mulinia lateralis.

Vitrification is a fast, simple and cost-effective cryopreservation technique which has emerged as a promising approach for sperm cryopreservation in a variety of animals, including aquaculture species. However, sperm vitrification in bivalves remains undeveloped, although molluscan sperm cryopreservation has been studied for more than 50 years. This investigation aimed to develop a sperm vitrification technique in dwarf surfclam Mulinia lateralia (a model species in bivalve research) and to assess the cryodamage at different levels. The results showed that the synergistic cryoprotective effect of coenzyme Q10 and ficoll was beneficial for the sperm vitrification with the dimethyl sulfoxide (DMSO) as the base cryoprotectant. The optimal cryoprotective solution for sperm vitrification in dwarf surfclam was 8% DMSO + 20 μM coenzyme Q10 + 5% ficoll. Although the D-stage larval rate achieved in vitrified sperm did not significantly differ from the fresh sperm, a sperm to egg ratio of 200 times higher was required in vitrified sperm than the fresh control at fertilization. This increased sperm requirement was associated with the vitrification-induced cryodamage, including reduced sperm motility, plasma membrane integrity, mitochondrial membrane potential, acrosome integrity, enzyme activity and DNA integrity, together with increased lipid peroxidation. In conclusion, the sperm vitrification technique presents a promising and technically achievable alternative for sperm cryopreservation in marine bivalve species.