Paternal-effect-genes revealed through sperm cryopreservation in Perca fluviatilis.

Knowledge about paternal-effect-genes (PEGs) (genes whose expression in the progeny is influenced by paternal factors present in the sperm) in fish is very limited. To explore this issue, we used milt cryopreservation as a specific challenge test for sperm cells, thus enabling selection amidst cryo-sensitivity. We created two groups of Eurasian perch (Perca fluviatilis) as a model - eggs fertilized either with fresh (Fresh group) or cryopreserved (Cryo group) milt from the same male followed by phenotypic-transcriptomic examination of consequences of cryopreservation in obtained progeny (at larval stages). Most of the phenotypical observations were similar in both groups, except the final weight which was higher in the Cryo group. Milt cryopreservation appeared to act as a "positive selection" factor, upregulating most PEGs in the Cryo group. Transcriptomic profile of freshly hatched larvae sourced genes involved in the development of visual perception and we identified them as PEGs. Consequently, larvae from the Cryo group exhibited enhanced eyesight, potentially contributing to more efficient foraging and weight gain compared to the Fresh group. This study unveils, for the first time, the significant influence of the paternal genome on the development of the visual system in fish, highlighting pde6g, opn1lw1, and rbp4l as novel PEGs.

Mercury distribution in the great cormorant (Phalacrocorax carbo) from the Krogulna ponds and Nysa Klodzka River (Poland).

Mercury belongs among the highly hazardous substances present in the environment and represents a great health risk for many animals including predatory and piscivorous birds. The aim of this study was to assess the total mercury content in the main detoxifying organs (liver and kidney) and in the muscles of adult great cormorants (Phalacrocorax carbo) caught at two localities (the Krogulna ponds and the Nysa Klodzka River) in southern Poland. The aim was to compare the locality with the iron ore mining history (the Krogulna ponds) with a site without such load (the Nysa Klodzka River). The total mercury content in the great cormorants decreases as follows: kidneys > liver > muscle in both monitored localities and significantly differs between the localities. The average mercury content varied from 0.58 ± 0.38 mg/kg (muscle) to 1.39 ± 1.42 mg/kg (kidney) in the cormorant from the Krogulna ponds and from 1.09 ± 0.40 mg/kg (muscle) to 3.12 ± 1.55 mg/kg (kidney) in the cormorant from the Nysa Klodzka River. The accumulation of mercury does not correlate with the mining history as it was higher at the Nysa Klodzka River, but it is probably influenced by the different fish stocks in these two localities (omnivorous fish in the Krogulna ponds versus predatory fish in the Nysa Klodzka River).

Differences in predator-avoidance behavior between two invasive gobies and their native competitors.

Globally, fish are frequently introduced beyond their native range. Some, like Ponto-Caspian gobies, are becoming invasive, achieving high colonization rates and constituting frequent prey for native predators. However, little is known about the effectiveness of antipredator behaviors of the invaders, which may shape their role in the invaded community and contribute to the invasion success. We compared antipredator behaviors of invasive gobies and native fish species after their detection by the predator, when the danger becomes direct. We studied 2 fish pairs, each consisting of an invasive and native species co-occurring in the environment and belonging to the same prey guild: (1) the racer goby Babka gymnotrachelus versus European bullhead Cottus gobio, (2) the monkey goby Neogobius fluviatilis versus gudgeon Gobio gobio, facing a naïve predator (the Eurasian perch Perca fluviatilis). We analyzed behaviors of single prey individuals (escaping, staying in shelter, and activity) and single predators (activity, searching, following, capturing, and latency to prey consumption). In the predator presence, the bullhead was less active and more often managed to escape after capture than the racer goby. The gudgeon escaped before the capture more often than the monkey goby. The predator succeeded later with the bullhead compared to racer goby, whereas no differences in ingestion time occurred between the gudgeon and monkey goby. The results suggest that, in terms of hunting effort of native predators, the invasive gobies are equivalent to or more profitable prey than their native analogs, which can facilitate the integration of the gobies into local food webs.

Telomerase Activity in Somatic Tissues and Ovaries of Diploid and Triploid Rainbow Trout (Oncorhynchus mykiss) Females.

Telomerase activity has been found in the somatic tissues of rainbow trout. The enzyme is essential for maintaining telomere length but also assures homeostasis of the fish organs, playing an important role during tissue regeneration. The unique morphological and physiological characteristics of triploid rainbow trout, when compared to diploid specimens, make them a promising model for studies concerning telomerase activity. Thus, in this study, we examined the expression of the Tert gene in various organs of subadult and adult diploid and triploid rainbow trout females. Upregulated Tert mRNA transcription was observed in all the examined somatic tissues sampled from the triploid fish when compared to diploid individuals. Contrastingly, Tert expression in the ovaries was significantly decreased in the triploid specimens. Within the diploids, the highest expression of Tert was observed in the liver and in the ovaries of the subadult individuals. In the triploids, Tert expression was increased in the somatic tissues, while the ovaries exhibited lower activity of telomerase compared to other organs and decreased compared to the ovaries in the diploids. The ovaries of triploid individuals were underdeveloped, consisting of only a few oocytes. The lack of germ cells, which are usually characterized by high Tert expression, might be responsible for the decrease in telomerase activity in the triploid ovaries. The increase in Tert expression in triploid somatic tissues suggests that they require higher telomerase activity to cope with environmental stress and maintain internal homeostasis.

Cystidicola farionis, a Swim Bladder Parasite of European Smelt: Characterization of the Nematode Trehalose Strategy.

The molecular identification of Cystidicola farionis (a swim bladder nematode of European smelt from the Vistula Lagoon in Poland) was performed. Their prevalence level was determined, and changes in the trehalose synthesis pathway in larvae and adult nematodes were demonstrated. The trehalose level was almost four times higher in adult nematodes than in larvae. In contrast, the activity of both enzymes (trehalose 6-phosphate synthase, TPS and trehalose 6-phosphate phosphatase, TPP) involved in the synthesis of trehalose was higher in larvae than in adults under optimal conditions. The optimum pH for TPS isolated from larvae and adults was pH 7.0. The optimum pH for TPP from larvae and adults was pH 7.0 and pH 8.0, respectively. The optimal temperature was 20 °C, and Mg2+ ions were an activator for trehalose-synthetizing enzymes from both sources. Enzymes isolated from adult nematodes were less susceptible to divalent ion chelator and inorganic phosphate than larval enzymes. The dynamic transformation of trehalose in the nematode developing inside the swim bladder of the smelt appears to be an important metabolic pathway in the nematode survival strategy. These studies are aimed at a better understanding of the issue of the metabolic adaptation of parasites, which, in the future, may indirectly contribute to the elimination of the parasite from aquacultures, which will impact public health.

Application of high hydrostatic pressure (HHP) shock to induce triploid development in the European grayling (Thymallus thymallus L.).

Use of parental individuals from allochthonous lineages to produce stocking specimens resulted in genetic contamination of many local European grayling (Thymallus thymallus) populations. Stocking programs including triploid individuals that are not able to reproduce with stocks located in natural habitats due to being reproductively sterile could contribute to populations of the European grayling and prevent regional lineages from the genetic pollution. The main goal of the present research, therefore, was to provide conditions for triploidization of the European grayling and examine gonads of triploid individuals. High Hydrostatic Pressure (HHP) shock (9000 psi/5 min.) applied to activated and incubated grayling eggs at 10 °C between the 17 min. 30 sec. and 20 minute timepoints subsequent to the time of insemination resulted in inhibition of the second polar body release and triploidization. Histological analysis confirmed that all 1-yr-old cytogenetically confirmed triploids had gonadal tissues indicative of sterility with the gonads being composed of connective tissue including fibroblasts, adipocytes and degenerated epithelial structures without there being any differentiated germ cells detected. There were no differences in the body length and Fulton's condition factor between 1 year old diploid and triploid grayling, however, the average body weight was markedly less in triploid than diploid specimens. Although, 1-yr-old triploid European female and male graylings did not have fully developed gonads, before these triploid specimens are deemed safe, as a result of being reproductively sterile for stocking in natural grayling habitats, there needs to be further examination of the 2- and 3-year-old triploid specimens.

Perfluoroalkyl Substance Contamination Levels of Pike (Esox lucius L.) and Roach (Rutilus rutilus L.) from Selected Masurian Lakes in Eastern Europe.

Perfluoroalkyl and polyfluoroalkyl substances (PFAS) are of increased concern because of their bioaccumulation in humans and the biota, the health risk they pose to humans and other animals, and their persistence in the environment. In the present study, the occurrence of PFAS in selected tissues from pike (Esox lucius L.) and roach (Rutilus rutilus L.) collected from two lakes in the Masurian Lake District (Poland) in eastern Europe was addressed. Ten PFAS were analyzed in the tissue of the brain, liver, kidneys, gonads, and muscles by micro-high-performance liquid chromatography coupled with tandem mass spectrometry. Only perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorononanoic acid, and perfluorodecanoic acid were detected above the limit of quantification. Concentrations in pike and roach from Elckie Lake and Hancza Lake were estimated to be 14/27 and 4.4/3.2 ng/g wet weight, respectively. The contributions of PFOS and PFOA to the sum of PFAS calculated for particular tissues for each fish species were higher than those of the other analyzed compounds: PFOS was found to be predominant in fish from Elckie Lake, whereas PFOA predominated in fish from Hancza Lake. It was noted that PFAS concentrations in tissues declined in the following order: kidney > gonads ≈ brain > liver > muscle. The sum of the greatest estimated PFAS concentration was 9.7 ng/g wet weight in kidneys of pike collected from Hancza Lake. No correlation was noted between PFAS concentration and fish size. The information provided in our study gives a better understanding of the potential dependencies in PFAS distribution and accumulation in biota. Environ Toxicol Chem 2021;40:3317-3327. © 2021 SETAC.

Characteristics and Cryopreservation of Semen of Sex-Reversed Females of Salmonid Fish.

Sex reversal has been used as a breeding strategy by salmonid fish to produce genetically and phenotypically single sex populations. Production of all-female fish has great importance for the creation of monosex female triploids of salmonid fish, which are valued for their sterility, lack of female maturation, and larger commercial size. Among salmonids, the majority of rainbow trout (Oncorhynchus mykiss) production is based on all-female production with a high proportion of all-female triploid production in Europe. The main aim of this review is to present the recent knowledge regarding sex-reversed females (SRFs) of salmonid fish. We discuss the methods of sex reversal as well as their effects on the morphology and histology of the reproductive tract. We focus on the characteristics of SRF semen as well as the factors determining semen quality. The lower quality of SRF sperm compared to that of normal males has resulted in the need for the artificial maturation of semen. Most importantly, methods of semen storage-both short-term and long-term (cryopreservation)-that can improve hatchery operations are presented with the special emphasis on recent progress in development of efficient cryopreservation procedures and use of cryopreserved semen in hatchery practice. Moreover, we also address the emerging knowledge concerning the proteomic investigations of salmonid sperm, focusing primarily on the proteomic comparison of normal male and SRF testicular semen and presenting changes in SRF rainbow trout sperm proteome after in vitro incubation in artificial seminal plasma.

Characterization of carp seminal plasma Wap65-2 and its participation in the testicular immune response and temperature acclimation.

Two functionally distinct isoforms of warm-temperature acclimation related 65-kDa protein (Wap65-1 and Wap65-2) with a role in the immune response are present in fish. To our knowledge, contrary to Wap65-1, Wap65-2 has neither been isolated nor functionally characterized in carp especially in reproductive system. The aim of this study was to characterize Wap65-2 and ascertain its functions in immune response and temperature acclimation within reproductive system. Wap65-2 corresponded to one of the most abundant proteins in carp seminal plasma, with a high immunologic similarity to their counterparts in seminal plasma of other fish species and a wide tissue distribution, with predominant expression in the liver. The immunohistochemical localization of Wap65-2 to spermatogonia, Leydig cells, and the epithelium of blood vessels within the testis suggests its role in iron metabolism during spermatogenesis and maintenance of blood-testis barrier integrity. Wap65-2 secretion by the epithelial cells of the spermatic duct and its presence around spermatozoa suggests its involvement in the protection of spermatozoa against damage caused by heme released from erythrocytes following hemorrhage and inflammation. Our results revealed an isoform-specific response of Wap65 to temperature acclimation and Aeromonas salmonicida infection which alters blood-testis barrier integrity. Wap65-2 seems to be related to the immune response against bacteria, while Wap65-1 seems to be involved in temperature acclimation. This study expands the understanding of the mechanism of carp testicular immunity against bacterial challenge and temperature changes, in which Wap65-2 seems to be involved and highlights their potential usefulness as biomarkers of inflammation and temperature acclimation.

Feed contamination with zearalenone promotes growth but affects the immune system of rainbow trout.

To investigate the effects of feed contamination with zearalenone (ZEN) at the current European Commission (EC) guidance value (2 mg⋅kg-1 feed) on the growth and health of rainbow trout, we performed a long-term feeding trial under aquaculture conditions. It started with the external feeding of the fish larvae, and continued for 96 weeks, at which point the fish had reached market size. To assess the growth of fish and their feeding efficiency throughout this period, the fish were regularly weighed and measured, and their feed consumption was monitored. Additionally, to investigate potential health effects, after 72 weeks of the exposure to ZEN, the fishes' blood was analyzed for major hematological and biochemical indices, and their head kidney, spleen, and liver were examined for morphological, histopathological, cytological, and molecular changes. Finally, to gain insight into the metabolism and distribution of ZEN in fish, the content of free and glucuronidated forms of ZEN and its major metabolites was measured in the intestine, liver, and muscles of the exposed fish. The feed-borne exposure of rainbow trout to ZEN at a dose of 2 mg⋅kg-1 feed resulted in higher feeding efficiency and growth rate, most probably due to the anabolic properties of the ZEN metabolite. Importantly for the consumers of fish, despite absorption and metabolism of ZEN in the digestive system of the fish that had been exposed for 72 weeks, the residuals of ZEN were not transferred to the fishes' muscles, which rules out a potential risk to human health related to the consumption of fish meat. However, the increased growth of fish fed with the contaminated feed may come at some cost, as the exposure to ZEN was associated with modulation of key components of the adaptive and innate immune systems. Moreover, the trunk kidney of ZEN-fed fish showed massive inflammation that was likely caused by pathogen infection. These findings raise concerns about fish health under the current recommended EC guidance values.

Acclimation to cold and warm temperatures is associated with differential expression of male carp blood proteins involved in acute phase and stress responses, and lipid metabolism.

The environmental temperature affects plasma biochemical indicators, antioxidant status and hematological and immunological parameters in fish. So far, only single blood proteins have been identified in response to temperature changes. The aim of this study was to compare the proteome of carp blood plasma from males acclimated to warm (30 °C) and cold (10 °C) temperatures by two-dimensional differential gel electrophoresis followed by MALDI-TOF/TOF mass spectrometry. A total of 47 spots were found to be differentially regulated by temperature (>1.2-fold change, p < 0.05): 25 protein spots were more abundant in warm-acclimated males and 22 were enriched in cold-acclimated males. The majority of differentially regulated proteins were associated with acute phase response signalling involved in: i) activation of the complement system (complement C3-H1), ii) neutralization of proteolytic enzymes (inter-alpha inhibitor H3, fetuin, serpinA1, antithrombin, alpha2-macroglobulin), iii) scavenging of free hemoglobin and radicals (haptoglobin, Wap65 kDa), iv) clot-formation (fibrinogen beta and alpha chain, T-kininogen) and v) the host's immune response modulation (ApoA1 and ApoA2). However, quite different sets of these proteins or proteoforms were involved in response to cold and warm temperatures. In addition, cold acclimation seems to be related to the proteins involved in lipid metabolism (apolipoproteins A and 14 kDa) and stress response (corticosteroid binding globulin). We discovered a strongly regulated protein Cap31 upon cold acclimation, which can serve as a potential blood biomarker of cold response in carp. These studies significantly extend our knowledge concerning mechanisms underlying thermal adaptation in poikilotherms.

Helminths of European smelt Osmerus eperlanus (Linnaeus, 1758) in Lake Hancza and the Vistula Lagoon, with special regard to their zoonotic threats.

Reports published in recent years on the increased risk in the Baltic Sea of fish infection with larvae of nematodes potentially pathogenic to humans have prompted a study of European smelt Osmerus eperlanus with respect to the presence of these parasites in the Vistula Lagoon. Additionally, samples of this fish from Lake Hancza were comparatively considered. The body cavity, the surface of internal organs, the alimentary tract and the swimbladder of fish from two environmentally different water bodies were taken into account in the analysis. Only the tapeworm Proteocephalus longicollis was found in the alimentary tract of the fish from Lake Hancza. Depending on the period of study, the prevalence ranged from 60% to 100%, and the mean intensity from 11.5 to 42.0. The helminth fauna of smelt from the Vistula Lagoon was more diverse: Cystidicola farionis occurred with a prevalence from 25.9 to 75.0% and a mean intensity of infection from 2.7 to 66.0%, Proteocephalus longicollis and cystacanths of Corynosoma with a prevalence ranging in different years from 12.0 to 44.0% and from 14.8 to 50.0%, respectively. Zoonotic parasites, such as nematode larvae of the Anisakidae family (Contracaecum sp., Anisakis simplex), occurred in the intestine with a prevalence of 31%, 72% and 22% in consecutive years of the study.

Presence of gynogenetic males suggests a female heterogamety in sterlet Acipenser ruthenus L.

Investigation of the heterogametic sex in sterlet Acipenser ruthenus L. was performed using meiotic gynogenesis and gonadal histology. Eggs from the albino females were activated by UV irradiated sperm of wild colored males and exposed to a heat shock. The resultant fish were all albino and exhibited exclusively maternal inheritance of the microsatellite DNA markers. Cytogenetic analysis indicated that gynogenetic progeny were diploids with 120 chromosomes. Based on the histological analysis, more than 86% of the gynogenetic individuals were found to be females. Moreover, some males (7%), sterile speciemens (3.5%) and fish with unidentified gonads (3.5%) were observed among the gynogenetic fish. Presence of both females and males in the gynogenetic offspring is indicative that the heterogametic sex in sterlets is female.

Effect of 11ß-hydroxyandrostenedione on European whitefish Coregonus lavaretus (Linnaeus, 1758).

The goal of the present research was to evaluate the efficiency of 11ß-hydroxyandrostenedione (OHA) applied in the diet to achieve sex reversal in the European whitefish (Coregonus lavaretus). At 32day post-hatching, fish were reared in four groups: fish fed with 10ppm of OHA (10 OHA), fish fed with 20ppm of OHA (20 OHA), fish fed without OHA (C) and fish fed without OHA and reared in the water from 20 OHA group (R). The experimental groups were conducted in separate recirculation systems and the first phase of the experiment lasted 63days. For the histological analysis of the gonads, fish from all groups were reared without OHA treatment for an additional 91days (second phase). At the end of the first phase of the experiment, survival of the whitefish ranged from 34.5±11.1% to 51.5±7.3%. The final body weight and coefficient of variation in the weight ranged from 5.6±1.2g to 6.9±1.5g and from 21.5 to 22.7%, respectively. No negative effects of OHA treatment on the growth and the survival of the whitefish were found. Six histological categories of the whitefish gonads were observed. Apart from the typical ovaries and testes, two types of the intersexual gonads (ovotestis and testis-ova) and two types of the sterile-altered gonads were distinguished. No gonadal females were found among fish from any of OHA groups. Gonadal males constituted of 60% and 50% of the fish from 10 OHA and 20 OHA groups, respectively. Intersexes were observed in all groups with the highest proportion found among fish from R variant. Rate of sterile individuals in 10 OHA and 20 OHA groups was 17% and 30%, respectively. The proportion of fish with normal testes to fish with other types of gonad varied from 0.43:1 to 1.5:1 with the higher ratio observed in both OHA groups. Lack of the females among fish from OHA groups suggested OHA affected growth and development of ovaries in the whitefish. However, a high percentage of the sterile fish in both OHA treated groups indicated application of lower doses of OHA for masculinization of the whitefish in the further research.

Gonadogenesis and annual reproductive cycles of an endangered cyprinid fish, the lake minnow Eupallasella percnurus (Pallas, 1814).

In this study, gonadogenesis, the effect of temperature (15, 20 and 25°C) on sex differentiation, and annual changes in the gonads of mature lake minnow Eupallasella percnurus (Pallas, 1814) were determined. The lake minnow was found to be a primary gonochoristic fish species, where gonads are formed directly in the ovaries or testes. The morphological differentiation of gonads was initiated 35days post hatch (DPH) when two types of gonadal anlages were visible: a pear-shaped gonad attached by a single mesentery string and a spindle-shaped gonad attached on both sides to the peritoneum. Gonadogenesis occurred faster in females than in males, with the first previtellogenic oocytes and ovarian lamellae being already observed in 45 DPH fish. In males, cytological differentiation occurred approximately 85 DPH, when the fish reached an average body weight of more than 400mg. No significant effect of rearing temperature on sex ratio in lake minnow juveniles was observed. The proportion of males and females was similar (close to 1:1) in all of the thermal-treated groups, although there were effects of temperature on the final sizes of fish. Histological examination of wild, mature lake minnow ovaries during the annual cycle (from May to February the following year) showed asynchronous oocyte maturation. The testes were characteristic of multi-batch spawning fish. Quantitative dominance of spermatids and mature spermatozoa in May was observed, while the presence of primary and secondary spermatocytes in all other periods was confirmed. These changes were also reflected in the seasonal variation in the gonado-somatic index in both sexes, with the highest mean values of 11.2% (females) and 4.0% (males) in May, which were found to be significantly different to all other periods. The data presented in this study provide an important contribution to our understanding of the biology and reproductive strategy of the endangered lake minnow.

Temporal Changes in Gametogenesis of the Invasive Chinese Pond Mussel Sinanodonta woodiana (Lea, 1834) (Bivalvia: Unionidae) from the Konin Lakes System (Central Poland).

Gametogenesis and the temporal changes occurring in the ovaries and testes throughout the reproductive cycle in the invasive alien bivalve, Chinese pond mussel Sinanodonta woodiana (Lea), from the heated Konin lakes system (central Poland) were studied using histological techniques. S. woodiana was confirmed to be a gonochoristic species with overall sex ratio of 1:1. The examined morphological parameters of Chinese pond mussel spermatozoa, i.e. 42 µm mean total length; 4.3 µm mean head length and the maximum size of previtellogenic (34-43 µm) and vitellogenic oocytes (75-83 µm) are consistent with values established for closely related members of the Unionidae family. Our results suggest that S. woodiana in the Konin lakes system are able to spawn throughout March to October, with a season of higher reproductive activity in females extending from March to April. This type of reproductive biology may contribute to the Chinese pond mussel's success in thriving in freshwater ecosystems.

Molecular cytogenetic study of the European bitterling Rhodeus amarus (Teleostei: Cyprinidae: Acheilognathinae).

The European bitterlings (Rhodeus amarus) from the Eastern locations were cytogenetically examined by conventional and molecular techniques. All analyzed individuals presented invariably the same chromosomal constitution of 2n = 48, with 8 metacentrics + 20 submetacentrics + 20 subtelo-acrocentrics and C-banding positive heterochromatin at the pericentromeric regions in most of the chromosomes. Moreover, some of the chromosomes had short arms entirely built with heterochromatin. GC-rich Ag-NORs (nucleolus organizer regions) were located at the short arms of two submetacentric chromosomes, and the length polymorphism of these regions was found. Multiple location of 28S rDNA sequences with fluorescence in situ hybridization signals was observed on the long and/or short arms of three submetacentric chromosomes including NOR regions and short arms of three to five acrocentric chromosomes in the studied fish. 5S rDNA sites were found on the short arms of two subtelocentric chromosomes, and telomeric repeats were localized at the ends of all chromosomes. Provided results have expanded our knowledge concerning genetic characteristics of the European bitterlings that may be profitable in the conservation programs of this endangered species.

Ovarian alterations in wild northern pike Esox lucius females.

The aim of the present study was to analyse the occurrence of macroscopically visible ovary alterations in 2 populations of northern pike Esox lucius L. originating from lakes in the Mazurian Lake District (NE Poland). The alterations were characterised by ovary tissue that was morphologically malformed, in part or in whole, and contained immature oocytes, i.e. trophoplastic or previtellogenic oocytes instead of vitellogenic oocytes. These alterations were found only in the ovaries, and no morphological alterations of the testes were noted. Macroscopic and histological analyses were carried out in order to classify the observed alterations in the ovaries. Three types of alterations were identified in which morphological malformations as well as histological investigation of the ovaries were considered. An analysis of the size and age of the fish in relation to the occurrence of alterations as well as of the macroscopic and histological nature of the alteration types was made. The data obtained revealed no lake or age dependency of the observed alterations. Based on the results obtained, we suggest that the presence of endocrine disruptors in the environment or/and genetic factors could be responsible for these kinds of gonad anomalies. However, our results did not allow us to determine the aetiology of the alterations.

Quality and quantity of smelt (Osmerus eperlanus L.) sperm in relation to time after hormonal stimulation.

The effect of Ovaprim (salmon GnRH analogue and a dopamine antagonist) treatment on the quantity and quality of smelt (Osmerus eperlanus L.) sperm was studied in relation to time after hormonal stimulation. Sperm was obtained at 0, 24, 48 and 72 h after treatment (n=13/each time point). Computer Assisted Sperm Analysis (CASA) was used to evaluate sperm motility parameters and histological analysis was used to examine the testis morphology. Only a small volume of semen (1-5 µl) was collected at the beginning of the experiment (time 0) but it dramatically increased 24 h after hormonal treatment. A further increase in semen volume was recorded 48 h after hormonal stimulation. CASA parameters, such as percentage of motile cells, progressive motility, curvilinear velocity, straightlinear velocity, straightness and amplitude of lateral head displacement of stripped sperm increased 48 h after hormonal treatment, which indicates high quality of sperm. No further increase in sperm quality was recorded at 72 h. Compared to stripped semen, testicular semen was characterized by a slightly lower quality. In addition, histological analysis indicated that 24 h after hormonal treatment, a high number of spermatozoa was released from the testis. Ovaprim-stimulated smelt became clearly darker than the control fish. In conclusion, our results suggest that smelt semen should be collected 48 h after hormonal stimulation to ensure high quality of semen. This time may vary depending on maturation status of testis.

Biochemical and physiological characteristics of semen of sex-reversed female rainbow trout (Oncorhynchus mykiss, Walbaum).

This works studies the biochemical (protein concentration, osmolality, antitrypsin activity, lactate dehydrogenase activity) and physiological characteristics (sperm motility characteristics) of semen of sex-reversed female rainbow trout (n=42) obtained with the application of 11ß-hydroksyandrostendione for sex reversal. All data were arbitrarily divided into three classes depending on the percentage of sperm motility: I XX<25%; II XX 25-50% and III XX>50%. The average percentage of sperm motility was 18±7% n=12 (group I XX); 42±6% n=15 (group II XX) and 65±12% n=15 for group III XX, respectively) to link the values of semen parameters to the maturation stage of semen. Semen from 12 normal males of the same age was used as a reference group. Sperm concentration as well as protein concentration, osmolality, antitrypsin activity, and lactate dehydrogenase activity in seminal plasma of sex-reversed females were higher compared with the values obtained for normal male rainbow trout. The values of these parameters declined with the increasing percentage of sperm motility toward values established for normal males. The fertilization success of semen (3×10(6) spermatozoa/egg) of sex-reversed females was very high (above 90%) for both the percentage of eyed embryos and hatched larvae and was related to sperm motility classes. Correlations between the quality parameters of sex-reversed females semen corresponded to those established previously for the semen of normal male rainbow trout. Antitrypsin activity, lactate dehydrogenase, protein concentration, and osmolality were found to be characteristic of seminal plasma of sex-reversed females. The maturity of sex-reversed female spermatozoa seems to be associated with the decline in the values of those parameters toward the values characteristic for seminal plasma of normal males.