Cryopreservation: Extending the viability of biological material from sea urchin (Echinometra lucunter) in ecotoxicity tests.

The sea urchin, Echinometra lucunter, is widely used in embryo-larval tests for ecotoxicological studies in Brazil and other countries. For each test, sea urchins are collected from the wild and this can cause impact on wild populations and it is limited by the weather and season which in turn limits the ability to carry out the tests. Cryopreservation is a method of live biological material storage at low temperature and can be used for long periods with little decline in viability, reducing the number of animals taken from the wild and enabling testing to be carried out on demand, irrespective of spawning season or location. In this study, 15 combinations of cryoprotective agents (CPAs) were evaluated on spermatozoa, subjected to a rapid cooling curve followed by immersion in liquid nitrogen. Twenty-four CPA combinations were evaluated on eggs subjected to a more gradual cooling curve in nitrogen vapor down to -35 °C and then plunging in liquid nitrogen. Fertilization tests using cryopreserved spermatozoa gave high pluteus larvae yields (≈80%) when concentrations of 10.5% or 13.65% ME2SO or 13.65% ME2SO+15.75% sucrose were used. The higher concentrations of ME2SO plus sucrose were more effective at maintaining the fertilization capacity of spermatozoa post-thawing. Egg cryopreservation was not successful with 0% fertilization observed post-thawing. The results suggest that it is feasible to implement spermatozoa cryopreservation as technological innovation to create a sperm bank for E. lucunter, which can be used in ecotoxicological tests, bringing benefits for researches and contributing to the conservation of the species.