Cryopreservation of semen in streaked prochilod (Prochilodus lineatus): use of ultra-freezer as alternative technique to liquid nitrogen

Abstract

Although sperm cryopreservation is well developed, different methods can be used for freezing fish semen, mainly with the intention of reducing the use of liquid nitrogen. For this purpose, ultra-freezers could, among other advantages, facilitate the maintenance of semen compared to liquid nitrogen containers. Indeed, ultra-freezers have an increased capacity to store semen, and they can be available in places where there are no other technologies. The aim of this study was to develop an alternative protocol for freezing streaked prochilod Prochilodus lineatus semen using an ultra-freezer. The fish semen (n=10) was collected and frozen using an ultra-freezer (−80°C), with a solution containing 10% DMSO diluted in 5% Beltsville thawing solution in the proportion of 1:4 (semen/diluent) in 0.5-mL straws. Each week, a tube was randomly chosen, thawed, and used to evaluate seminal parameters using computerized semen analysis (CASA). Then, the data were subjected to analysis of variance followed by Tukey’s post hoc test. Motility, motility time, vitality, sperm velocity, and secondary morphology were influenced by storage in ultra-freezer (p < 0.05). However, these parameters were kept at levels considered adequate during the time tested. This is, to the best of our knowledge, the first report of frozen P. lineatus semen under ultra-freezer conditions (−80°C), demonstrating that this method is efficient in maintaining the semen quality for up to 10 weeks.