Evaluation of extenders and cryoprotectants on motility and morphology of longtooth grouper (Epinephelus bruneus) sperm

Abstract

The objective was to evaluate various extenders and cryoprotectants on postthaw motility of longtooth grouper (Epinephelus bruneus) sperm, based on sperm motility ratio (SMR), sperm velocity (SV), and morphological damage after thawing. To evaluate the optimal cryoprotectant for cryopreservation of longtooth grouper sperm, semen was diluted in 0.3 M glucose extender containing one of four cryoprotectants (dimethyl sulfoxide, methanol, ethylene glycol, and glycerol [Gly]) at a final concentration of 10% or 20%, and then frozen in liquid nitrogen vapor (-76 °C for 3 minutes) before storage in liquid nitrogen. Semen diluted in 0.3 M glucose containing 10% Gly had the highest postthaw SMR (57.5 ± 2.5%, mean ± SEM). To identify the optimal extender, semen was diluted in one of two longtooth grouper artificial seminal plasma (LG-ASP; LG-ASP1 and LG-ASP2) extenders containing Gly at concentrations of 0%, 5%, 10%, or 20%, then cryopreserved using the described procedure. Semen diluted with LG-ASP2 extender containing 10% Gly cryoprotectant had the best postthaw SMR (66.3 ± 2.0%) and SV (135.9 ± 4.5 μm/s). Compared with fresh sperm, some structural damage was observed in cryopreserved sperm. We concluded that the combination of LG-ASP2 and 10% Gly (as extender and cryoprotectant, respectively), resulted in high postthaw SMR and SV for cryopreservation of longtooth grouper sperm.