**4.2 Experiments on pikeperch sperm**

During the improvement of the cryopreservation technique of pikeperch sperm in laboratory cryoprotectant DMSO showed better fertilization rates than methanol but fertilization experiments in hatcheries showed opposite results. Literature data can be found on successful usage of both cryoprotectants in several fish species. The objective of this thesis is the usage of cryopreserved pikeperch sperm in hatcheries and according to the results of the experiments in the whole it was concluded that methanol and 1:1 dilution rate is suitable for freezing pikeperch sperm.

A significant variation was observed in motility after thawing and in hatching rate in the first experiments. This variability is caused by mixing of sperm with urine. This problem can successfully be eliminated when the stripping of sperm is conducted with a silicone catheter. According to this method the sperm is stripped with this silicone catheter directly from the testes preventing the mixing of sperm with urine or feces. One year later the use of catheter resulted in substantially better hatching rates.

It was observed that the increasing of the amount of eggs fertilised with a single 0..5-ml straw resulted in improved hatching rates. The reason for this can be that different amounts of eggs behaved differently in Zug-jars. The dose of 10 g of eggs were slightly stuck together, the dose of 30 g of eggs stuck in smaller batches while the dose of 50 g of eggs rolled freely. In spite of the fact that the 50 g of eggs sample had not replicates, these results suggest that fertilisation of larger amounts of eggs result in better hatching rates.

It is supposed that the eggs in the middle of the 10-g batches were more sensitive for oxygen deficiency than the more loose larger egg samples.

Another explanation for these results is that methanol in smaller eggs samples was in higher concentration, thus the toxic effects were more drastic than in larger samples. The lower sperm-egg ratio in larger egg samples had no influence on the results, suggesting that the amount of sperm was in surplus in the case of smaller egg samples.
