Comparison of Protein Fractions in Seminal Plasma from Multiple Sperm Collections in Sterlet (Acipenser ruthenus)

Seminal plasma of sterlet Acipenser ruthenus was evaluated using comparative proteomics to characterize its protein fractions and to determine any influence of multiple sperm collections on these proteins. An experimental group of fish was used, in which sperm was collected three times at 5 h intervals. Protein fractions of seminal plasma were determined by SDS‐gel electrophoresis (SDS‐PAGE) and two‐dimensional electrophoresis high‐resolution gels (2D). At all stripping times, five protein bands with molecular weights of 93, 53, 48, 33 and 28 kDa were identified using SDS‐PAGE. No significant differences (p > 0.05) in relative mass of protein bands among collections were observed. At the third collection, 20 protein spots were detected from the two‐dimensional gels, compared to 17 found at the first and second collections. Ten protein spots, from the third stripping, were analysed. Screening of these spots by mass spectrometric analysis showed positive results for spot 10. Direct comparison across public databases revealed sequence similarity with two hypothetical proteins, MCAG_00854 and IscW_ISCW011489. Differences in the seminal plasma protein fractions were found at the third stripping compared to the first two. It is hypothesized that these extra proteins after the third collection could be involved in some step of intracellular mechanism which is responsible for regulating of spermatozoa motility. However, protein identification revealed no significant distinction for any protein spot and protein sequences available in public databases. These results highlighted the need for a complete genome sequences for sturgeons.     

Structural abnormalities of common carp <Emphasis Type="Italic">Cyprinus carpio</Emphasis> spermatozoa

Spermatozoa of common carp Cyprinus carpio are typically consist of a primitive head without acrosome, a midpiece with several mitochondria, a centriolar complex (proximal and distal centriole), and one flagellum. During an evaluation of the motility of common carp spermatozoa, we found spermatozoa with more than one flagellum and/or “double head” in three different individuals. This may be related to abnormal spermatogenesis. Ultrastructure and physiological parameters of spermatozoa were examined using light microscopy (dark field with stroboscopic illumination), transmission and scanning electron microscopy, and flow cytometry. The recorded pictures and videos were evaluated using Olympus MicroImage software. All spermatozoa with more than one flagellum had a larger head and shorter flagella. They occasionally demonstrated several cytoplasmic channels separating the flagella from the midpiece. Each flagellum was based upon its own centriolar complex, with the connection of the flagellum to the head always at a constant angle. The flagella always consisted of nine peripheral pairs and one central doublet of microtubules. Sperm exhibited a relative DNA content similar to that found in sperm from normal males, with higher coefficients of variation. Although similar abnormalities have been found in livestock, where they were described as a defect in spermiogenesis, no comparable results have been reported in fish. The frequency at which these abnormalities occurs, the fertilization ability of males with defects in spermiogenesis, the influence of these abnormalities on progeny in terms of ploidy level, and the occurrence of deformities warrant further investigation.     

Triploidization of European catfish, Silurus glanis L., by heat shock

Heat shock was applied to fertilized eggs of European catfish, Silurus glanis L., for the induction of triploidy. A heat shock of 40. 5°C lasting 1 min and starting 9 min after gamete activation gave the best results with 88.93% of hatched viable fry (92.41% in control group). Yields of hatched viable triploid fry reached 63.14% or 50.60%, when expressed as percentage of the absolute number of viable fry or number of living eggs in eye-bud stage, respectively (P < 0.05).     

Evaluation of spermiation indices with multiple sperm collections in endangered sterlet (Acipenser ruthenus)

This study investigated the effects of multiple collections of sperm on endangered sterlet (Acipenser ruthenus) sperm functional parameters [spermatozoa motility and curvilinear velocity (VCL)] as well as on protein concentration and osmolality of seminal plasma. The average sperm volume and mean spermatozoa concentration per male were significantly altered with multiple collections. On the other hand, no significant effect of multiple collections on protein concentration of seminal plasma was observed. In all experimental groups, moderate impact of sequential collection on osmolality (p < 0.05) of seminal plasma was observed. Ninety to 100% of motile spermatozoa were observed at 15 s after activation, with an average VCL of 181.12 ± 19.10 μm/s. After 90 s, average VCL decreased to 130 ± 26 μm/s. Motility was maintained for up to 4 min. The maximum percentage of motile spermatozoa was observed after the third collection of sperm. The spermatozoa VCL increased significantly with subsequent collections. The results of this study provide new data on the effects of multiple collections on quantitative and qualitative parameters of sperm in sterlet. The data confirmed that the sequential stripping has no negative effect on the percentage of motility and spermatozoa velocity. This should be beneficial for the development of sterlet aquaculture programs.     

Top-crossing with evaluation of slaughtering value in common carp (Cyprinus carpio L.) offspring

A three-year test of growth performance andoffspring survival from top-crossing of commoncarp hybrids in low altitude (350 m above thesea level, middle European climate) and in highaltitude (750 m above the sea level, middleEuropean climate) was terminated by assessmentof slaughtering value of edible parts in therespective strains. A recently establishedHungarian synthetic mirror carp strain (HSM)was chosen for testing as maternal strain. TheHSM, as well as wild Amur carp (AC), Ropshacarp (ROP) and Tata carp (TAT) were used aspaternal strains.The evidential strain*altitude interactioneffect was demonstrated as significant forweight of fish (0.0005), fillet without skin(0.0001), index of highbackedness (0.005),index of head length (0.0404),Fulton's coefficient (0.0497),index of widebackedness (0.0315) andgonadosomatic index (0.0082). The interactioneffect was demonstrated as insignificantbetween altitude*sex and betweenstrain*sex.The comparative test of analysis of variance(ANOVA) revealed significantly (P < 0.05)higher weight of fish after killing on lowaltitude (1611.6 g) than on high altitude(1090.3 g), but processed body yield andpercentage of fillet without skin was notsignificantly different (64.0 and 34.2 for lowaltitude, 64.2 and 33.4 for high altitude). Thepercentage of processed body weight andpercentage of fillet without skin in analysisboth altitudes were significantly better infemales (65.5 and 34.6%) than in males (62.5and 33.00%), respectively. The highestsignificant differences in weight of fish afterkilling were found between HSM (890.8 g) andHSM × AC (1283.3 g) in high altitude. In lowaltitude, it was between HSM (1527.2 g) andHSM × AC (1706.9 g) and HSM × ROP (1693.0 g).     

Dietary n‐3/n‐6 ratio affects the biochemical composition of Eurasian perch (Perca fluviatilis) semen but not indicators of sperm quality

In general, the effects of dietary fatty acids (FA) on sperm quality have received less attention than egg quality, and were never studied in perch. This study investigated the effects of dietary FAs on the quality and chemical composition of sperm in Eurasian perch (Perca fluviatilis). Two experimental diets containing 16% lipids and 45% proteins were compared. The n‐3/n‐6 ratios tested were 0.2 for diet 1 (D1) and 7.0 for diet 2 (D2). No significant effects of the n‐3/n‐6 ratio were observed on the sperm characteristics, either in terms of the sperm volume (around 1.2 mL) and density, spermatozoa motility (94%) and velocity, or the sperm osmolality. All these parameters corresponded to semen of good quality in Eurasian perch. Interestingly, both the FA composition and the lipid class profile of the semen were correlated to the tested diet. However, basal levels of certain highly unsaturated FAs such as eicosapentaenoic acid, 20:5 n‐3 and docosahexaenoic acid, 22:6 n‐3, were maintained in the sperm irrespective of the diet tested. Perch semen was characterized by high levels of cholesterol, phosphatidylethanolamine and phosphatidylcholine. In conclusion, the dietary n‐3/n‐6 ratio affects the lipid composition of perch semen but not the indicators of sperm quality.     

Quantity, motility and fertility of tench Tinca tinca (L.) sperm in relation to LHRH analogue and carp pituitary treatments

The spermiation of tench males was stimulated with Supergestran containing mammalian LHRHa lecireline at the following doses: 5, 10, 20 and 40 g kg⁻¹ b.w.; then with carp pituitary suspension (CPS) at a dose of 2 mg kg⁻¹ b.w. and with a control of saline physiological solution. The following days, meaning 24, 48 and 72 h after injection, sperm was collected to evaluate volume and the number of sperm per male per kg body weight (B.W.) The percentage of motile sperm and velocity of spermatozoa were measured 48 h after hormonal injection, and 72 h after hormonal injection the sperm was evaluated for fertilization and hatching ability. All 42 males in experimental groups were diploid. Live weight did not differ significantly among experimental groups. The strongest stimulation of spermiation was achieved with LHRHa in dosage of 20 and 40 g kg⁻¹ b.w. and CPS compared to males of the control group and lower dosage of LHRHa. Analysis of variance showed no significant influence of the treatment on the velocity and percentage of motile spermatozoa. The effect of different treatment on the fertilization capacity (the number of spermatozoa per egg was equilibrated) was significant. Significantly the highest quality of sperm collected 72 h after injection expressed by percentage of fertilization and hatching (62–65% fertilization and 61–64% hatching rates, respectively) was found for LHRHa in dosage of 20 and 40 g kg⁻¹ b.w. Significantly the lowest parameters of fertilization and hatching were found for the control group, on the 12% level.     

Effects of Temperature on In Vitro Short‐Term Storage of Sterlet Sturgeon (Acipenser Ruthenus) Ova

Artificial propagation of sturgeons is becoming increasingly important for recovery efforts as well as for commercial production. Sterlet Acipenser ruthenus is a common Eurasian sturgeon with a small body size and one of the fastest reproductive cycles among the sturgeons. The practical question being addressed in this study was how long fertilization of ovulated eggs can be delayed without substantially reducing the hatching rate, and an ancillary question is under what' temperature conditions do eggs retain good quality. Broodstock were injected with homogenized carp pituitary extract (CPE); ovulated eggs from three females were allocated to various treatment groups for temperature storage (control, 7°C, 11°C, 15°C and 19°C) until fertilized. Storage times at the regulated temperatures prior to fertilization were for 2.5, 5.0, 7.5 and 10.0 h. After the selected storage times in ovarian fluid, eggs were fertilized and transferred to incubation cages and then they were counted. Three replicates were allocated to each storage period and temperature. Hatched larvae were counted at 7‐day post‐fertilization. We found that sterlet eggs do not need to be fertilized immediately after collection. Reasonably good quality was retained for several hours if temperature conditions are fairly cool and stable. Eggs retained good quality when stored at 7°C and 11°C for up to 10 h with 54.1 ± 2.9 to 69.9 ± 7.9% hatching success, but egg quality was significantly reduced after 5‐h storage at 19°C (p < 0.01) and 7.5‐h storage at 15°C (p < 0.05) compared to cooler temperatures. Uniform temperatures between 7°C and 11°C can be considered as appropriate for storage of eggs in ovarian fluid for up to 10 h. This information can have practical application to routine hatchery practice for acipenserids, as well as for certain research protocols.     

Effect of water temperature on the physiology of fish spermatozoon function: a brief review

Motility is a key factor in function of the spermatozoon and determines semen quality and fertilizing capacity. Effective motility occurs when sperm is diluted in a swimming solution, the adequacy of which is determined by factors varying according to fish species. Spermatozoon motility rate and velocity, as well as duration of the motility period, are influenced by the temperature of the water in which broodfish are held. Increase in temperature of swimming medium beyond the optimal increases cell metabolism, leading to an increase in velocity with rapid depletion of energy resources, promoting early cessation of movement. The aim of this review was to discuss current information on the influence of temperature on quantitative spermatozoon properties, which could affect sperm function. Our findings provide a greater understanding of fish sperm physiology and a biological foundation for the further development of spermatozoon motility investigations as well as reproduction technologies.