Development of an efficient and standardized method for the cryopreservation of Arctic charr milt and its use in the fertilization of brook trout eggs to produce 'sparctic' hybrids

The general objective of this study was to establish a standardized procedure for the cryopreservation of Arctic charr milt and its use in the production of `sparctic' hybrids. The specific objectives of this study were to determine: (i) the optimal final sperm concentration as well as (ii) the final glucose concentration, (iii) the effect of post-thaw storage time on sperm motility characteristics, and (iv) the fertilization ability of cryopreserved Arctic charr milt at 0 and 60 min after thawing at sperm:egg ratios of 500,000:1 and 1,000,000:1. We observed high post-thaw sperm motility (49-57%) at final sperm concentrations ranging from 0.5 to 2.0 x 10(9) spermatozoa ml(-1) in straw. We determined that a final glucose concentration of 0.15-0.19 M in 7.5% methanol was optimal (post-thaw sperm motility 48-60%). Milt cryopreserved at optimal conditions (final concentrations: 0.17 M of glucose, 7.5% methanol and 1.5 x 10(9) spermatozoa ml(-1)) could be stored for up to 15 min after thawing without the loss of sperm motility. The ability of cryopreserved milt to fertilize brook trout eggs was high (68-72%) and did not differ between the tested sperm:egg ratios (500,000:1 and 1,000,000:1) when milt was used immediately after thawing or after 60 min of post-thaw storage. The milt used in this study was obtained from two hatcheries and was characterized by significantly different values of sperm concentrations. In conclusion, a standardized and efficient cryopreservation method for Arctic charr milt has been developed and used for the production of `sparctic' hybrids. Species-specific requirements for final sperm and glucose concentrations and the post-thaw storage time were determined. The hatchery of origin of Arctic charr should be taken into consideration, as high variability in sperm characteristics may be expected. The development of this method and the positive results of the fertilization trials indicate promising potential for the implementation of cryopreserved milt in hatchery practice.