Introduction
Climate change impacts the marine environment, which faces acidification and rising temperatures. As key fishes in Mediterranean aquaculture rely on external fertilization, these environmental changes can impair sperm quality and reproduction . Therefore , we aimed to evaluate the effect of temperature and pH of artificial seawater (ASW) on the activation of sperm from different aquaculture Mediterranean species. Furthermore, sperm evaluation was used to select resilient males and to form a germplasm bank with climate change resistant samples.
Material and Methods
European sea bass, Dicentrarchus labrax (n = 30), gilthead seabream, Sparus aurata (n = 93), and Senegalese sole, Solea senegalensis (n = 116 ) s perm samples were collected at the Institute of Aquaculture Torre de la Sal (IATS - CSIC). In addition, samples from S. sole males (n = 16) were collected at the Institute of Agrifood Research and Technology (IRTA). Samples were obtained by abdominal pressure, diluted in species-specific extenders (NAM or Ringer solution) and transported at 4 °C to the Universitat Politècnica de València (UPV) . At UPV, sperm motility was analyzed using a CASA-Mot system after activation with ASW. Only samples with more than 50% motility were used for further experiments.
Three challenge tests were tested trying to identify sperm samples that were resilient to environmental pH and temperature variations. The first one was the sperm activation using ASW at different pH (6.5, 7.2, 7.4, 7.6, 7.8, 8, 8.2, 9.5). The second challenge test was the sperm activation with ASW at different temperatures (seabream and S. sole: 4, 16, 22 °C; sea bass: 4, 12, 20 °C). The third one evaluated the combined effect of ASW pH and temperature: pH (7.8 and 8.2) x (seabream and S. sole: 4, 16, 22 °C; sea bass: 4, 12, 20 °C) . Before activation, the samples were incubated (~1h) at the same temperature.
The challenge test combining seawater pH and temperature was the most effective for selecting resilient males. Therefore, sperm samples from sea bass (n = 10), seabream (n = 43), and S. sole (n = 10) were subjected to challenge test 3. Males were selected according to the variation in the MOT percentage obtained by applying variations caused by climate change (acidification and temperature increase) compared with the natural environment during their reproductive periods (pH 8.2; sea bass, 12 °C; seabream and S. sole, 16 °C). Males were selected considering the variations on sperm motility in comparison with values obtained under natural conditions. Different selection thresholds can be applied: up to 5% variation for a rigorous level, up to 10% for a less rigorous level, and up to 15% for S. sole samples.
Results
No practical differences were observed in the motility of the three species when challenge tests 1 and 2 were applied. However, in challenge test 3, it was clear that acidification and warming of the marine environment affected the sperm quality of the three Mediterranean species. The temperature increase impaired the sea bass (Fig. 1A ) and S. sole sperm (Fig. 1B ), and the decrease in ASW pH affected the MOT of sea bream sperm (Fig. 1C).
After applying the challenge test 3, 6 sea bass were selected at a 5% variation level and 7 at up to 10% variation concerning the MOT values obtained with the natural parameters of pH and temperature during the reproductive period. For seabream, 18 males at 5% variation level and 32 males at up to 10% level were selected. Only 1 S. sole male at the 5% and 2 at 10% variation levels were selected. At the 15% level, 6 breeders were selected.
Discussion
Environmental pH and temperature influence sperm motility (Alavi & Cosson, 2005). Sperm of some species can resist slight environmental acidification , as was observed in sea bass and S. sole in the present study. However, seabream sperm was impacted by acidification. Sperm activation in a more acidic environment may have caused a reduction in the membrane potential of sperm mitochondria since seminal plasma alkalization is necessary for sperm activation to occur (Alavi & Cosson, 2005). Increasing the incubation and activation temperature did not affect seabream sperm, while impaired sea bass and S. sole sperm motility. The sperm activation using above-normal temperatures leads to a decline in sperm motility, probably due to the limited antioxidant enzyme efficacy in mitigating oxidative stress (Dadras et al., 2017).
Alterations in pH and temperature driven by climate change impacted the sperm quality of three Mediterranean aquaculture species. The developed challenge tests showed to be a valuable technique for identifying males resilient to such environmental stressors, whose genetic material can be conserved in germplasm banks.
Acknowledgements
This study forms part of the ThinkInAzul programme and was supported by MICIU with funding from European Union NextGenerationEU (PRTR-C17.I1) and by Generalitat Valenciana [GVA-THINKINAZUL/2021/012, -24, -42; Principal investigators: J.F. Asturiano (Universitat Politècnica de València), J. Pérez-Sánchez, and A. Gómez (CSIC), respectively].
References
Alavi, S. M. H., Cosson, J. (2005). Sperm motility in fishes. I. Effects of temperature and pH: A review. Cell Biol. Int. 29, 101–110.
Dadras, H., Dzyuba, B., Cosson, J. , Golpour, A., Siddique M. A. M, Linhart, O. (2017). Effect of water temperature on the physiology of fish spermatozoon function: a brief review. Aquac. Res. 48, 729–740.