Reliable techniques for the cryopreservation of both sperm and oocytes of the blue mussel Mytilus galloprovincialis Lamarck would increase the availability of seed supplies out-of-season and enhance efficiency in selective breeding. We have investigated the optimal cryotechnique for blue mussel oocytes. The toxicity of three cryoprotective agents (CPAs) dimethyl sulfoxide (DMSO), ethylene glycol (EG) and propylene glycol (PG) at different concentrations (1-5 M) and exposure times (0.25-30 min) were investigated for mussel oocytes at room temperature (20 degrees C) or on ice. The same CPAs (1, 1.5 and 2 M) as well as three different cryoprotectant mixtures 1.5 M EG + 0.2 M trehalose + 100 Milli-Q water (EGTM); 1.5 M EG + 0.2 M trehalose + 75 Milli-Q water + 25 seawater; 1.5 M EG + 0.2 M sucrose + 100 Milli-Q water were tested by comparing the post-thaw oocyte fertilization rate after using the slow-cooling method. Vitrification was also examined; however, this method failed to produce any post-thaw surviving oocytes. Among the tested CPAs, EG was the least toxic to oocytes. There was a tendency for the equilibration of CPAs on ice to achieve a higher oocyte fertilization rate compared with that at room temperature, and this difference was significant at concentrations of 3 and 4 M (P 0.01). The DMSO, EG and PG treatments all resulted in post-thaw fertilization, with EGTM achieving the highest number of surviving oocytes (32 ). At the optimal seeding temperature (-7 degrees C), the addition of 0.2 M trehalose to EG resulted in a better fertilization rate of post-thawed oocytes than the addition of 0.2 M sucrose. All of the treatments evaluated produced D-larvae from post-thawed oocytes, although the rates were low.
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