Effect of double freezing fish semen on sperm motility and fertility

Abstract

Refreezing freeze-thawed sperm has been found to improve the effectiveness of cryopreserved semen in mammals. However, such options have been never tested in fish. The objectives of this study were (i) to assess the effects of two freeze–thaw cycles on sperm motility parameters of different fish species (grayling, brown trout, Eurasian perch, Siberian sturgeon, rainbow trout, sex-reversed female (srf) rainbow trout, and sex-reversed female brook trout) and (ii) to test the fertilizing ability of rainbow trout sperm after two freeze–thaw cycles. Double freezing of semen resulted in the preservation of sperm motility in all samples, 59.2% post-thaw sperm motility was recorded for grayling, 49.4% for brown trout, 34.8% for Eurasian perch, 3.2% for srf rainbow trout and 7% for Siberian sturgeon semen. A significant correlation was found between the sperm motility of semen after single and double cryopreservation for most fish species. The fertilization rate at the eyed stage of rainbow trout fresh semen was approximately 64% and did not differ from the fertilization rate of semen after a single cycle of freezing. After two freeze-thaw cycles, significant decreases in the eyed stage (48.4%) and hatching stage (46.6%) were recorded. Our results clearly demonstrate the suitability of double cryopreservation of fish semen. The approach can be important for improved effectiveness of cryopreserved fish semen, which can potentially reduce costs and the space needed to maintain a bank of frozen semen. This method offers a new tool for the improved use of valuable samples of cryopreserved semen, which could increase the possibility of protecting endangered fish species and individuals that are extremely valuable for breeding aquaculture species.