Genetic Variation in the ND1 Gene and D-loop in Protected and Commercially Exploited European Cisco (Coregonus albula L.) Populations.

The European cisco (Coregonus albula L.) is a species with high environmental requirements. The deterioration of environmental conditions in recent decades has decreased its distribution. Currently the species is conserved by stocking, and the few existing natural populations are at risk of extinction. Therefore, contemporary studies involve not only reporting phenotypic parameters, but also determining the genetic structure of the population. This is an important aspect monitored in the C. albula population, which provides information valuable for proper fishing economy. This study included valuable populations from lakes located in Drawa National Park (DNP) and Wigry National Park (WNP), as well as lakes used for commercial fishing. In order to molecularly characterize the European cisco, the control region and NDl gene were sequenced from 48 individuals from 9 populations from lakes throughout northern Poland. Analysis revealed that populations from two park lakes (Marta, Ostrowieckie) are unique. This was also the case for some sequences originating from Lake Wigry. The mean value of genetic diversity was 0.2% within each region and 0.1-0.3% between the investigated regions. The obtained results demonstrated the necessity to strengthen and protect natural populations of the European cisco, which constitute a valuable element of the European ichthyofauna.

Spermatozoa Motility Parameters of the Pumpkinseed (Lepomis gibbosus) in a Standardized Solution and Ambient Water.

The sunfish are classified as an invasive species after their introduction into a new habitat. Milt quality and spermatozoa movement in several sunfish were described. The aim of this study was to determine the basic milt characteristics and spermatozoa motility parameters in pumpkinseed males after using ambient water and a standardized solution to examine the parameters in mimicked natural conditions and in controlled conditions allowing interspecies data comparisons. Nine spermatozoa motility parameters were traced using computer-assisted sperm analysis (CASA) after activation with 30 mM NaCl and ambient water at 20°C. Spermatozoa motility parameters on activation, except for ALH, differed after using saline solution and ambient water. The CASA parameters were higher in the first medium. On the other hand, spermatozoa motility was high in both media, with an average of 89.6 and 85.2% in saline solution and ambient water, respectively. In the media, initial spermatozoa motility was characterized by a mean curvilinear velocity of 122.4 and 108.5 µm s-1, and a linearity of 79.7 and 67.1%, respectively. The active motility phase in the saline solution was approx. 8 min was longer than that measured in ambient water (mean 3 min). The pumpkinseed spermatozoa reveal good motility parameters, especially the high percentage of motile sperm, linearity, long duration of spermatozoa movement, as well as good velocity. It can be assumed that apart from reproductive strategy, spermatozoa motility parameters are valuable features of the male reproductive biology which may contribute to reproductive success.

Annual Developmental Cycle of Gonads of European Perch Females (Perca fluviatilis L.) from Natural Sites and a Canal Carrying Post-cooling Water from the Dolna Odra Power Plant (NW Poland).

The European perch is a species endowed with high adaptation capabilities as regards different environmental conditions. The aim of the study was to analyse the annual developmental cycle of ovaries of the European perch from the Oder river, Lake Dabie and a drainage canal (Warm Canal) carrying post-cooling water from the Dolna Odra power plant (annual average water temperature in the canal is higher by 6-8°C than the water of the other sampling sites). Most of the female perch caught in the canal carrying post-cooling water had immature stage 2 gonads (delayed development of the gonads) and were smaller than the fish from the other sites. No traces of spawning in the form of deposed egg strings were found in the drainage canal. Adult individuals avoid high temperatures found in the Warm Canal. In April, in perch from all sites, ovaries with post-spawning oocytes were observed. The spawning season of the females lasted from the beginning of April until May. Stage 4 of gonad development, with oocytes in advanced vitellogenesis, was the longest and ranged from September through February.

Genetic Variation of Salmo trutta L. Populations from the Catchment Areas of the Rega, Parseta and Wieprza Rivers Evaluated by RAPD and SSR Markers.

By using simple sequence repeats (SSR) and random amplification of polymorphic DNA (RAPD) markers, the genetic variability of three Salmo trutta L. populations from three rivers, the Wieprza, the Rega and the Parseta, was determined. The investigated populations showed a high level of genetic variability. Microsatellites showed that observed heterozygosity (Ho) was higher than the expected heterozygosity (He), with most heterozygotes found in the population from the Parseta river and the fewest in the Wieprza population. The F(IS) coefficient in all investigated populations of the sea trout indicate a high excess of heterozygotes. The highest genetic differentiation was observed between the sea trout from the Rega river and those from the Wieprza (0.366). The obtained results based on microsatellite and RAPD analysis showed that the investigated populations formed two groups. The first group consisted of the sea trout populations from the Wieprza and the Parseta rives, while the second group was formed solely by the Rega river population.

Spermatozoa concentration influences cryopreservation success in sea trout (Salmo trutta m. trutta L.).

Variation among individuals is substantial for spermatozoa concentration in fresh milt in sea trout (Salmo trutta m. trutta L.). The objective of the present study was to examine effects of spermatozoa concentration in this species on subsequent cryopreservation success. Milt with high spermatozoa concentration was diluted with seminal plasma to obtain concentrations ranging between 6 and 24 × 10(9) mL(-1) with steps of 2 × 10(9) mL(-1). Diluted milts were cryopreserved in 0.25-mL straws with extender (0.3 M glucose) containing 10% methanol and 10 % (vol/vol) supplement of hen egg yolk. The dilution ratio was 1:3 (milt:cryomedium). Cryopreservation efficacies were assessed according to evaluation of motility of frozen/thawed spermatozoa and quantification of fertilizing ability. Percentage of motility of frozen/thawed spermatozoa was influenced by spermatozoa concentration in the cryomedium (P < 0.05). The highest motility was observed in samples with 3.0 to 4.0 × 10(9) spermatozoa per mL of cryomedium, which corresponds to 12 to 16 × 10(9) spermatozoa per mL in fresh milt. Higher sperm concentrations and lower sperm concentrations in cryomedium reduced the effectiveness of cryopreservation when compared with the optimum. Cryopreservation success measured according to fertilization rate was in agreement with results for motility of frozen/thawed spermatozoa, but the optimum could not be determined with statistical precision because of differences in fertilization rate among individual donor males. However, a significant positive correlation was found between postthaw motility and fertilization rate and between cryopreserved spermatozoa velocity and fertilization rate (P < 0.05). In sea trout, cryopreservation efficiency is influenced by spermatozoa concentration in cryomedium. Individual adjustment of the dilution ratio, based on initial spermatozoa density, is recommended in the freezing protocol. Maximum cryoresistance of the cell was obtained when spermatozoa concentration in cryomedium ranged from 3.0 to 4.0 × 10(9) mL(-1).

Effect of pH and cation concentrations on spermatozoan motility of sea trout (Salmo trutta m. trutta L.).

Environmental conditions during external fertilization in fish have a significant effect on spermatozoan motility (MOT) and fertilization ability. Even in the same family of fish, spermatozoa might differ in sensitivity to ions present in the external medium. Elucidation of such differences within a species would help to understand spermatozoan biology and to determine external conditions that would optimize spermatozoan MOT and successful fertilization. Objectives of the current study were to determine the effects of pH and of sodium, potassium, and calcium ion concentrations in the activation solution on sea trout spermatozoan MOT. Six parameters characterizing MOT (MOT, curvilinear velocity [VCL], linearity, amplitude of lateral head displacement, beat cross frequency, and duration of MOT) in spermatozoa activated in prepared buffers were traced by computer-assisted sperm analysis. Sea trout spermatozoa were motile over a wide range of pH values, and increasing pH did influence MOT, VCL, linearity, amplitude of lateral head displacement, and MOT duration. The optimum pH for sperm MOT was established at approximately 10. Increasing K(+) ion concentration within the observed range caused a decrease in MOT and VCL. Spermatozoan movement ceased at 8 mM KCl concentrations. In Ca(2+) buffers, sperm were motile within the range of 0 to 70 mM CaCl(2) concentration; although beyond 8 mM concentration, VCL and MOT gradually declined. Spermatozoan aggregation was observed at the highest ion concentrations tested. Increasing CaCl(2) concentration affected MOT pattern from initiation to termination of spermatozoan movement in a similar manner as changes associated with increasing pH. At concentrations of CaCl(2) higher than 0.5 mM and in buffers with pH values 10 to 11, movement of spermatozoa was characterized by high initial linearity followed by its gradual reduction. In contrast to the effects of KCl and CaCl(2), increasing NaCl concentration up to 90 mM Na(+) concentration prolonged the duration of spermatozoan movement and, up to 60 mM Na(+) concentration, slightly increased sperm velocity as well. Above the concentration of 90 mM NaCl, these parameters decreased; and at 240 mM of Na(+), spermatozoa did not activate.

Cytogenetic and morphological characterization of Corbicula fluminalis (O. F. Müller, 1774) (Bivalvia: Veneroida: Corbiculidae): taxonomic status assessment of a freshwater clam.

Chromosomes of Corbicula fluminalis were characterized by karyotype analysis and nucleolar organizer region (NORs) localization. The triploid chromosome number was confirmed as 54; the karyotype is composed of 3 metacentric, 15 submetacentric and 36 subtelo-acrocentic chromosomes. Silver staining revealed nucleolar organizers on the telomeric regions of three subtelo-acrocentic chromosomes. This is the first study on chromosomes of C. fluminalis. The results are discussed with regards to Corbicula species as well as its relationships to other mollusc species based on cytogenetic characters and morphometric of the shells.

Body size versus gonad maturation form in under-yearling precocious males of the sea trout (Salmo trutta m. trutta L.).

The study was aimed at analysing body size in relation to form of gonad maturation (amount of mature germ cells) in 329 under-yearling sea trout males. The fish, aged 7 months, were caught in late October-early November in 3 streams located in north-western Poland. Each stream supported fish belonging to a different sib group. Standard histological techniques and a computer image analysis programme were used to detect the class of gonad maturation and percentage of the gonad area occupied by tubules with active spermatogenesis. Gonad maturation forms were distinguished based on the latter criteria. Gonads with developing germ cells occupying less than 90% of gonad area were classified as incomplete forms of gonad maturation, others as complete maturation forms. In each sib-groups analysed, even the smallest individual were already precocious, their gonads being incompletely mature. The smallest maturing male measured 7.1 cm in length. The average size of an incompletely maturing individual was slightly smaller than that of the completely mature one but the difference lacked statistical significance (P > 0.05). The sib-group of smaller fish contained less precocious, and the gonads of the more precocious were incompletely mature, compared to the sib-group of larger fish (P < 0.001). It seems that the incomplete form of gonad maturation (defected maturation) occurs at a smaller critical fish size than the complete gonad maturation form. Incomplete maturation is more frequent smaller individuals and possibly in among slow-growing groups of fish.

Histology of salmonid testes during maturation.

The commonly applied classification systems of fish gonad maturity divide the maturation process into certain stages. However, the scales do not entirely reflect the continuity of the maturation process. Based on light microscope observations, the paper describes a comprehensive pattern of testicular transformations during maturation. The study was carried out on precocious underyearling and 1-year-old males of sea trout (Salmo trutta m. trutta L.), 1-year-old males of salmon (Salmo salar L.), and males of brown trout (Salmo trutta m. fario L.) aged from 7 months to 4 years. A total of 821 gonads collected during all seasons of the year were examined. The fish were fixed in Bouin's fluid. Histological slides of the mid-part of the gonad were made using the standard paraffin technique. The 3-6 microm sections were stained with Heidenhain haematoxylin. Histological changes of testes during maturation were similar in the three species studied. Immature and resting gonads contained type A spermatogonia in lobules only. The appearance of cystic structures containing type B spermatogonia in the lobules signalled the beginning of the sexual cycle in male gonads. Type B spermatogonia underwent synchronous mitotic divisions resulting in an increase in the total number of spermatogonia. As the spermatogenesis continued, the gonads showed a gradual increase in the number of cysts containing cells at all the spermatogenetic stages: type B spermatogonia, primary and secondary spermatocytes, spermatids, and spermatozoa. The well-formed spermatozoa were released to the lobule lumen once the Sertoli cells and spermatozoa connections broke up and the cyst disappeared. This was a continuous process observed throughout the spawning season. The spermatozoa were moved to the efferent duct. While some of the germ cells were completing spermatogenesis, the lobules contained less and less cysts with type B spermatogonia, primary and secondary spermatocytes, and spermatids; eventually all the cells completed spermatogenesis. At the end of maturation, vacuoles, up to 18.9 microm in final diameter (brown trout), appeared in the Sertoli cells. The vacuoles were visible in the lobule wall epithelium for a prolonged period of time. In most salmonid individuals examined, the reproductive cycles were observed to overlap. In some fish, the preparation for another cycle began very early, i.e., at the and of preceding spermatogenesis, which had not been observed before. Gonad maturation in some males was incomplete.