Cryopreservation of red snapper (Lutjanus argentimaculatus) sperm: effect of cryoprotectants and cooling rates on sperm motility, sperm viability, and fertilization capacity

Abstract

Sperm cryopreservation of red snapper (Lutjanus argentimaculatus) is essentially unexplored, although many species of the Lutjanidae family are considered to be high-value commercial species. The objective of this study was to develop a species-specific cryopreservation protocol for red snapper (L. argentimaculatus) sperm by optimizing cryoprotectants and cooling rates in the cryopreservation procedure. Ten cryoprotectants at four concentrations and two freezing protocols were examined in two separate experiments. In the first experiment, toxicity studies of dimethyl sulfoxide (DMSO), glycerol, propylene glycol (PG), ethylene glycol (EG), formamide, methanol, ethanol, sucrose, trehalose, and dimethylacetamide (DMA) on sperm motility were performed. Semen diluted 1:1 in Ringer solution were exposed to cryoprotectants at four final concentrations of 5%, 10%, 15%, or 20% for periods of 10, 20, 30, 40, 50, 60, 90, and 120 min at room temperature (25 degrees C). The cryoprotectants and concentrations that showed the least toxic effect on sperm motility were selected for cryopreservation trials. In the second experiment, selected cryoprotectants were then assessed for freezing capacity of sperm as follows: DMSO 5% and 10%, PG 5% and 10%, EG 5% and 10%, ethanol 5%, and methanol 5%. Semen was diluted 1:1 in Ringer solution and equilibrated with selected cryoprotectants for 10 min at room temperature. Sperm were frozen in a controlled-rate programmable freezer at four cooling rates of 3, 5, 10, and 12 degrees C/min from an initial temperature of 25 degrees C to final temperatures of -40 or -80 degrees C before plunging into liquid nitrogen. Sperm equilibrated in 10% DMSO and cooled at a rate of 10 degrees C/min to a final temperature of -80 degrees C had the highest motility (91.1+/-2.2%) and viability (92.7+/-2.3%) after thawing. The fertilization rate of frozen-thawed sperm (72.4+/-2.4%) was not different (P>0.05) from that of fresh sperm (75.5+/-2.4%). This study apparently represents the first reported attempt for cryopreservation of L. argentimaculatus sperm.