This study was carried out to test how sperm cryopreservation affected nuclear DNA stability and whether progeny development was modified when eggs were fertilized with cryopreserved spermatozoa. The "comet assay" (alkaline single-cell gel electrophoresis assay) was adapted to trout spermatozoa to estimate DNA stability as measured by alkali-induced DNA strand break formation. Because trout eggs develop in water after fertilization (oviparous species) and that eggshell is easy to clear up after fixative treatment, progeny development was assessed from the blastodisc flattening stage of the embryos to the first feeding stage of the hatched fries by direct observation. All parameters under study were analyzed on each sperm and comparisons between parameters were made using paired data. Freeze-thawing of sperm slightly but significantly increased the percentage of nuclei showing altered DNA after comet assay. This increase was correlated to the decrease in fertilization rates of sperm, but the absolute percentage of altered nuclei was not predictive of the absolute fertilization ability of sperm. Assessment of progeny development showed that survival rate and abnormality rate obtained after fertilization with cryopreserved sperm were not different from those obtained with fresh sperm. It is concluded that trout sperm cryopreservation only slightly affected sperm DNA stability and that the use of cryopreserved spermatozoa did not impair offspring survival and quality.

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The protein composition of seminal fluid, blood serum, sperm plasma membrane and flagellum of rainbow trout were analysed by SDS-polyacrylamide gel electrophoresis. Immunological identity between proteins of the 2 fluids and sperm components was studied using crossed immunoelectrophoresis, rocket immunoelectrophoresis and immunoblotting. Results indicate that many seminal proteins are antigenically-related to serum proteins, proteins of sperm origin are present in seminal fluid in varying amounts, depending on the animals and sampling time, and several serum-like seminal proteins are bound to spermatozoa.Lipoproteins were isolated from seminal fluid (mean level: 33 μg/ml) and characterized. They were identified as being HDL-like lipoproteins. A possible physiological role is proposed for these seminal lipoproteins.

The effect of a long-term adaptation of rainbow trout to 8 and 18 degrees C combined with a corn oil- or a fish oil-supplemented diet on the characteristics of the spermatozoan plasma membrane was investigated. The experiment lasted up to 22 mon during which spermatozoa were collected from the mature males. Spermatozoan plasma membranes were isolated by nitrogen cavitation, and the cholesterol content, phospholipid composition and fatty acid pattern were investigated. Membrane viscosity was assessed on whole cells by electron spin resonance using spin-labeled phospholipids. Neither diet nor rearing temperature influenced the cholesterol content of the plasma membrane nor the phospholipid class distribution. The rearing temperature of the broodstock only slightly affected the phospholipid fatty acids. A minor decrease in 18:0 and increase in monounsaturated fatty acids was observed for the cold-adapted fish. These modifications were not sufficient to affect membrane fluidity, and we conclude that trout spermatozoa do not display any homeoviscous adaptations in these conditions. On the contrary, the dietary fatty acid intake greatly modified the fatty acid profile of plasma membrane phospholipids. The fish oil-fed trout displayed a much higher n-3/n-6 fatty acid ratio than did the corn oil-fed ones, but the 22:6n-3 levels remained unchanged. Modifications in plasma membrane composition by the diet were obtained although neither of the two diets was deficient in essential fatty acids. The enrichment in n-3 fatty acids, however, did not affect plasma membrane fluidity which was unchanged by the diets.

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