Eurasian perch is an important fish species for European aquaculture diversification, but the quality of reproduction still remains one of the main limitations for further industry development. In particular, the optimal condition to obtain the best quality of sperm is poorly understood. Cryopreservation is an efficient technique for the long-term storage of fish sperm (Bernáth et al. 2017). The establishment of sperm banks can support aquaculture production and conservation programs (Cabrita et al. 2010 , Martínez-Páramo et al. 2017) . In Eurasian perch, the available techniques are usually limited by the small sperm capacity that can be preserved (0.5 mL straws). However, larger than 0.5 mL straws or cryotubes have not been tested for perch sperm.
Materials and methods
The aim of our study was to measure the possible effects of two experimental rearing temperatures (6 °C and the conventionally used 12 °C) and of hormonal stimulation, on the motility parameters (pMOT , VCL, VSL, LIN, ALH, BCF), osmolality and fertilizing capacity of Eurasian perch sperm at the end of the reproductive cycle. A prior untested, large-scale (5 mL cryotube and Polystyrene box) cryopreservation method was implemented using fresh sperm obtained from the two above mentioned temperature groups. Males were injected with 100 µg body weight kg -1 sGnRHa . For cryopreservation, an extender formerly adapted for perch sperm was applied (137 mM NaCl and 76.2 mM NaHCO3, Szabó et al. 2005) at a ratio 1:10 (Bernáth et al. 2015).
No significant difference was recorded between the two rearing temperatures and between the saline control and sGnRHa treated groups on the different features of sperm quality. A similar fertilization rate was monitored in all sGnRHa treated (6 °C: 69±13%, 12 °C: 81±11%) and saline control groups (6 °C: 79±10%, 12 °C: 87±4%). Correspondingly, no significant difference in hatching rate was observed in the sGnRHa injected (6 °C: 27±9%, 12 °C: 40±20%) and saline control (6 °C: 35±18%, 12 °C: 36±7%) males. However, a notable negative effect of freezing process on sperm movement was observed following thawing in both temperature groups. No significant difference in the motility parameters was measured between the two temperature groups following large-scale cryopreservation. Furthermore, a similar result was observed in the fertilizing capacity (6 °C: 79±10%, 12 °C: 75±8) of thawed sperm as well as in the hatching rate (6 °C: 52±13%, 12 °C: 46±19%).
Discussion and conclusion
Our findings suggest that perch sperm quality and fertilizing capacity is not affected by the reduced rearing temperature (6 °C) following hormonal injection in Eurasian perch, using fresh or cryopreserved sperm. Despite a lower motility, sperm obtained from the reduced rearing temperature can be cryopreserved successfully using 5 mL cryotube. However, the freezing and thawing process needs to be improved using the adapted preservation method. Thawed perch sperm (cryopreserved in 5 mL cryotube) can be adapted for the fertilization of higher amounts of eggs in future experiments.
The study was funded from the European Union’s Horizon 2020 research and innovation programme under grant agreement No. 652831 (AQUAEXCEL2020) from the TNA project No . AE090022 (CRYOPERCH). This paper reflects only the authors’ view and the European Union cannot be held responsible for any use that may be made of the information contained herein. This research was supported by the Ministry of Innovation and Technology within the framework of the Thematic Excellence Programme 2020, National Challenges Subprogramme (TKP2020-NKA-16). Our experiments were also supported by the EFOP-3.6.3-VEKOP-16-2017-00008 project. The project is co-financed by the European Union and the European Social Fund.
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