Identification of transferrin in Atlantic cod Gadus morhua epidermal mucus.

The previously unreported presence of transferrin in Atlantic cod Gadus morhua epidermal mucus is described. A less destructive sampling method, which may result in decreased epidermal tissue damage, is discussed.

Automated sperm head morphology analyzer for open-source software.

Sperm head morphology has been identified as a characteristic that can be used to predict a male's semen quality. In the present study, we have developed an automated sperm head morphology analysis (ASMA) plug-in for open-source ImageJ software (http://rsbweb.nih.gov/ij/). We describe the plug-in's functionality, and confirm its validity for sperm head morphology analysis using fish sperm. Sperm head morphological measurements (length and width) made with the ASMA plug-in did not differ from manual measurements. Using the plug-in to measure sperm head-shaped objects of known size, the associated plug-in error rate was < 0.5%. Brightness and contrast ratios influenced sperm head measurements, suggesting the need for standardized protocols. This plug-in was effective at measuring elliptical (i.e., Atlantic cod) as well as slightly irregular (i.e., Chinook salmon) shaped sperm heads. In conclusion, our ASMA plug-in represents a versatile alternative to costly sperm morphology software.

Gonadal development of triploid Atlantic Cod Gadus morhua.

During their third and fourth years of life, triploid Atlantic cod Gadus morhua females exhibited diminished ovarian development at both macroscopic and microscopic levels, with significantly lower gonado-somatic indices (I(G)) and higher carcass yields (Y(C)) than diploid females during spawning periods. In contrast, diploid and triploid testes were indistinguishable through macroscopic and histological assessment. No significant differences were found in male I(G) or Y(C) in relation to ploidy, with the exception of a higher Y(C) for triploids during the first of the two monitored spawning seasons. As a result of suppressed oogenesis, triploid female G. morhua, but not triploid males, can be used to eliminate sexual maturation in this species and advance the development of the G. morhua aquaculture industry.

Semen characteristics and their ability to predict sperm cryopreservation potential of Atlantic cod, Gadus morhua L.

There is a lack of biomarkers or indices that can be used to predict the quality of fish semen samples following the freezing and thawing cycle. In the present study, a series of semen indices were tested to assess if they could accurately forecast the cryopreservation potential of Atlantic cod (Gadus morhua) semen. Fresh and frozen-thawed sperm activity variables were compared, and relationships between frozen-thawed sperm activity and fertilization success were examined. In comparison with fresh sperm, activity variables of frozen-thawed spermatozoa were reduced. Of the 18 males examined, mean (± SEM) spermatocrit of fresh sperm was 40.72 ± 4.23%, osmolality of the seminal plasma 366.32 ± 4.95 mOsmol/kg, pH 8.32 ± 0.04, protein concentration 1.05 ± 0.08 mg/mL, anti-trypsin activity 153.83 ± 19.25 U/L, and total antioxidant capacity 0.15 ± 0.03 µmol Trolox equivalents/mL. Frozen-thawed fertilization success was highly variable among males with values ranging from 18.5 to 90.2%. Regressions yielded significant positive relationships between frozen-thawed motility, velocity, track crossing frequency, and subsequent fertilization success. Sequential multiple regressions explained up to 95% of the variation in frozen-thawed sperm activity. Spermatocrit and pH of fresh semen were negatively related, whereas osmolality and antioxidant capacity were positively related to frozen-thawed motility and velocity. Each of these indices can be measured within minutes of collecting a fresh sample of semen and are thus early indicators of the capacity of semen samples to withstand cryopreservation. These results have many benefits for conservation of wild stocks, aquaculture production, and for understanding semen biology and cryobiology of fishes.

Genetic, maternal, and environmental variance components for body weight and length of Atlantic cod at 2 points in life.

Variance components were estimated for 2 body size traits of Atlantic cod at 2 time points. Wild-caught founders from 3 regions off eastern North America were spawned and their progeny were reared at 2 locations in 2 consecutive years. Full-sib families (n = 148) were kept separate until individuals achieved a size large enough to be tagged. At that time (220 d of age), BW and length of 47,637 offspring from 90 sires and 89 dams were recorded. The juveniles were then transferred to sea cages at 3 sites, where they grew further for more than a year. A second set of measurements was collected on 11,839 fish (634 d of age). Dispersion parameters were estimated using REML in bivariate analyses. Models included fixed degree-days (covariate), year × location subclasses, and genetic groups composed of connected families within region of origin. Random factors were animal (additive genetic effects), considering known relationships among the fish; dam (maternal effects); and family (effects common to full-sibs). At tagging, heritability estimates were small to moderate (0.15 and 0.24 for BW and length, respectively; SE = 0.14), similar to or somewhat larger than the proportions of variation ascribed to dams and families (11 to 16%). Later, heritability estimates were greater (0.27 ± 0.08 and 0.31 ± 0.09 for BW and length, respectively), whereas dam and family variance proportions were very small (3 to 4%). Omitting maternal or family components substantially increased the values obtained for heritability at both time points. At the later point, failure to account for maternal effects inflated heritability estimates by about 24% for both traits; ignoring family effects had double the impact. These effects persisted even though endogenous feeding lasts only a couple of weeks in this species and the fish had been pooled since tagging. Discarding data from parents that were completely confounded with their mates decreased heritability estimates slightly (by 0.04, for both traits) at the second point, with no loss of precision despite 15% fewer records and 34% fewer parents; the improved design seemed to have more fully disentangled the additive genetic effects. Estimates of genetic correlations between traits and between time points were very large (>0.89). The results imply that genetic variation exists for body size of cod at both stages. Poor data structure and inadequate models can potentially lead to overstatement of heritability, and thus also of the predicted selection response.

Cryopreservation of Atlantic cod Gadus morhua L. spermatozoa: Effects of extender composition and freezing rate on sperm motility, velocity and morphology.

Broodstock selection programs are currently underway for Atlantic cod (Gadus morhua). To complement and further these selection programs we need to develop sperm cryopreservation procedures. This will allow genomic DNA from males from selected individuals or stocks to be frozen and conserved in perpetuity. In our study we used a full factorial ANOVA design to examine the effects of diluent (Mounib's sucrose-based diluent, Hanks' Balanced Salt Solution, Mounib's sucrose-based diluent+hen's egg yolk, and Hanks' Balanced Salt Solution+hen's egg yolk), cryoprotectant (propylene glycol, dimethyl sulphoxide, and glycerol), and freezing rate (-2.5, -5.0, -7.5, and -10.0°C/min) on motility of cod frozen-thawed sperm. Sperm velocity and morphometric analyses of sperm heads and flagella were also assessed. We found that sperm motility-recovery index was strongly influenced by the presence of higher-order interactions of the factors we tested. The best cryoprotection used diluents that contained hen's egg yolk. Generally, extenders containing propylene glycol yielded higher post-thaw sperm motilities than those with dimethyl sulphoxide or glycerol. In comparison to sperm from other frozen-thawed extenders, sperm from extenders supplemented with propylene glycol had significantly higher curvilinear velocity. Cryopreservation showed no impact on sperm head morphology parameters, however, considerable damage to frozen-thawed sperm flagella was observed. We believe that our experimental/statistical approach and our results add significantly new information to the study of semen biology/cryobiology in fishes. Our findings are also highly relevant to the development of cod mariculture and for aiding in conservation efforts of this very important marine species.

Seasonal variations in seminal plasma and sperm characteristics of wild-caught and cultivated Atlantic cod, Gadus morhua.

The objective was to investigate changes, throughout the spawning season, in body size attributes and quantitative semen characteristics of wild-caught and cultivated Atlantic cod, Gadus morhua L. Sperm velocity increased significantly throughout the spawning season of cod from both origins. Curvilinear velocity (VCL; 30 sec post-activation) increased from 78.9+/-6.5 to 128.2+/-6.5 microm/sec (mean+/-SEM) between the beginning and end of the spawning season, respectively, for wild-caught cod, whereas for cultivated fish, it increased from 26.6+/-2.4 to 48.9+/-3.1 microm/sec between January and March. Spermatocrit did not undergo a significant seasonal change in wild-caught cod but did thicken for cultivated cod (24.6+/-4.2% in January to 40.5+/-4.4% in April; P<0.01). Sperm head area, perimeter, length, and width declined significantly at the end of the spawning season of cod from both origins (all P values<0.01). Seminal plasma osmolality and Na(+) ion concentration followed a dome-shaped function through the spawning season for both wild-caught and cultivated cod (P<0.05). For cultivated cod, seminal plasma pH was significantly lower at the start of the spawning season (P<0.001), whereas Ca(2+) increased then decreased (P<0.05). Body size attributes, spermatocrit, and seminal plasma constituents had significant relationships with sperm activity variables. These relationships varied as a function of time post-activation, month, and fish origin. Our findings may be used to (i) assess spermiation stage without killing males; (ii) optimize semen collection for hatchery production; (iii) characterize the potential impact of farming on sperm quality; and (iv) improve success of sperm cryopreservation and short-term storage.