Cryopreservation of sperm from turbot (Scophthalmus maximus) and application to large-scale fertilization

This paper reports on cryopreservation of sperm from turbot ( Scophthalmus maximus ). The effects of various extenders, cryoprotectants and sperm–egg insemination ratios on motility score and/or fertilization capacity of post-thaw turbot spermatozoa were examined to optimize cryopreservation procedures. Post-thaw motility of frozen sperm obtained with extender TS-2 was higher than those achieved with extenders D-15 and modified plaice Ringer solution (MPRS). The most effective cryoprotectant was determined to be 10% dimethyl sulfoxide (DMSO). Fertilization of small egg batches (2 ml eggs) with frozen sperm resulted in average fertilization rate (FR) (70.1±8.9%) and hatching rates (HR) (46.8±5.2%) similar to the fertilization rates (74.7±8.0%) and hatching rates (47.5±6.8%) of fresh sperm. The minimal density of frozen sperm required to obtain satisfactory fertilization rate was determined to be 2000:1 (sperm/egg). Fertilization of larger egg batches (40 ml eggs) with sperm frozen in 1.8-ml cryovials provided similar fertilization rates (71.6±7.3%) to that of the fresh sperm (76.2±5.7%), whereas the hatching rates (34.7±3.6%) of eggs fertilized with frozen sperm were slightly lower than those of fresh sperm (41.1±3.7%). Our results demonstrated that cryopreservation technique for turbot sperm in 1.8-ml cryovial could be used for hatchery purposes.