Early ontogeny of the spotted wolffish (Anarhichas minor Olafsen)

This study illustrates the embryo development of the spotted wolffish (Anarhichas minor Olafsen), an interesting candidate for cold‐water aquaculture. The egg morphology (semitransparent, yellow‐white with numerous oil droplets in the yolk), size (5.4–6.5 mm) and long embryogenesis (c. 800–1000 d°, depending on temperature) of A. minor are very similar to Anarhichas lupus. Cleavage is slow, and the first cell divisions take place at 12 h at 8°C. After 12 days the 2‐mm embryo with the first somites is laid down and the blastopore starts closing. The fat globules in the yolk fuse into one after 22 days, and after 30 days eye pigmentation is noticeable. After 44 days, eye pigmentation is strong, the digestive tract folded and a green gall bladder can be noted in the now 11‐mm‐long embryo. One week later the blood is brightly red, the intestine is pigmented and the lower jaw is well developed. Premature hatching may occur from this stage. After 58 days vascularization of the yolk is complete, capillaries are noted in the fin fold, the first ray rudiments are established in the tail and pectoral fins, and the four gill arches are covered by the operculum. The preanal finfold is reduced after 72 days, stomach and gill filaments are formed, and six pigmented rows are noted on the 17‐mm‐long embryo body. After 86 days all fin rays are seen and the digestive tract is intensely pigmented and folded. Hatching (normal) starts after 110 days and may last for 2–3 weeks. Late embryos and early larvae of A. minor have more distinct bands of pigment along the body compared with the closely related A. lupus. An increase in both length and weight of the embryos in individual batches occurs during the hatching period.     

The influence of rearing temperature on early development and growth of spotted wolffish Anarhichas minor (Olafsen)

Temperature influenced the developmental rate, survival and early growth of eggs and embryos of spotted wolffish, Anarhichas minor (Olafsen), an interesting candidate for cold water cultivation. The total incubation period decreased from 220 days at 4 °C (880 daydegrees), to 177 days at 6 °C (1062 daydegrees) and 150 days at 8 °C (1200 daydegrees) in these experiments. The proportion of normal embryos and survival of eggs until hatching were highest when the eggs were incubated at 6 °C. During the incubation period, the embryo and yolk sac size at 280 daydegrees was not significantly different but at 850 daydegrees the embryo size was inversely related to temperature and the remaining yolk sac size positively correlated with the incubation temperature. The transformation of yolk to body mass during incubation appeared to be most efficient at 4 °C, and the embryos hatched with a larger visible yolk sac at 6 and 8 °C. The largest larvae (wet‐weight) hatched from the largest eggs and the egg groups incubated at the lowest temperature (4 °C). There was no effect of temperature on meristic characters. During 6 weeks post‐hatching, all larvae from the three temperature groups were fed formulated dry feed in excess at 8 °C in low water‐level raceway systems. During startfeeding, the larvae from eggs incubated at the lowest temperature (4 °C) showed the highest growth rates (SGR). Best survival of larvae was noted among batches incubated at 6 °C.     

Growth and survival of first-feeding spotted wolffish (Anarhichas minor Olafsen) at various temperature regimes

In order to define temperature regimes that could benefit successful production of spotted wolffish (Anarhichas minor) juveniles, experiments with offspring from two different females were carried out. The larvae were fed a new formulated feed or a commercial start‐feed for marine fish, both of which have given high survival rates. In the first experiment newly hatched larvae were fed at constant 6 °C, 8 °C, 10 °C and 12 °C as well as at ambient seawater temperature (2.9–4.5 °C) during 63 days. High survival, 90% to 96%, was registered at ambient and most constant temperature regimes, whereas in the 12 °C groups survival was reduced to 80%. Growth rate (SGR) was very low, 1.8% day^?1, at the low ambient temperatures. Growth rate was positively correlated with temperature and varied between 3.1% day^?1 to 4.7% day^?1, from 6 °C to 12 °C. In the second experiment, set up to include potential detrimental temperatures and study beneficial effects of a more restricted, elevated first‐feeding temperature regime, the larvae were fed at constant 8 °C, 10 °C, 12 °C, 14 °C and 16 °C until 30 days post hatch, followed by constant 8 °C for the next 33 days. In this experiment, low survival, 25% and 2.0%, was registered at 63 days post hatch when larvae were reared initially at 14 °C and 16 °C respectively. The survival of the larvae at the other temperature regimes varied from 47% to 64%, highest survival rate (64%) was found at 8 °C. The lowest specific growth rate, 2.6% day^?1, was noted in the 16 °C group. At constant 8 °C to 14 °C (regulated to 8 °C), the SGR varied from 4.45% day^?1 to 5.13% day^?1. The larvae grew faster in the experiment when initially comparable temperatures (8 °C, 10 °C and 12 °C) were regulated to constant 8 °C after 30 days compared with the first experiment where feeding was carried out at the same constant temperatures (8 °C, 10 °C and 12 °C) during the whole experimental period.     

Metabolic and digestive enzyme activity profiles of newly hatched spotted wolffish (Anarhichas minor Olafsen): effect of temperature

Three groups of newly hatched spotted wolffish (Anarhichas minor) were held at three different temperatures in order to determine relationships between metabolic, digestive and growth response in rapidly developing larvae. Growth rates were successfully modulated by temperature (5, 8 and 12 °C). Activity levels of trypsin and lactate dehydrogenase (LDH) were positively linked to specific growth rates at all temperatures. Trypsin showed a positive compensation (higher activity at lower temperature) whereas glycolytic enzymes (pyruvate kinase and Lactate dehydrogenase) and aspartate aminotransferase (AST) showed a negative compensation (lower activity at lower temperature). Citrate synthase was not affected by growth rate, indicating that the level of aerobic capacity was adequate in sustaining the high energetic needs associated with rapid growth early in the life of the spotted wolffish. In light of our results, it is suggested that protein digestion, as demonstrated by the activity profile of trypsin in relation to growth rate and temperature, is likely a key growth‐limiting agent during the early‐life stages of wolffishes. Our results are discussed in comparison with A. lupus, a closely related species displaying different temperature preferences and growth capacities.     

Effect of rearing density on the growth and welfare indices of juvenile spotted wolffish,Anarhichas minor (Olafsen)

The goal of this study was to determine the optimal stocking density for rearing juvenile spotted wolffish, Anarhichas minor (Olafsen), at two different sizes and assess the welfare status in relation to density. No major growth impairment was observed, although smaller fish (50–100 g) were significantly affected by density during the 120 days of the experiment, with final mean weights of 119.6 ± 11.6, 118.0 ± 5.8 and 88.7 ± 0.6 g for initial rearing densities of 10, 20 and 40 kg m^?2 respectively. No effect of rearing density was seen for larger fish (100–160 g) during the 90 days of the experiment, with final mean weights of 160.2 ± 5.9, 159.7 ± 3.7 and 163.7 ± 11.5 g at fixed rearing densities of 20, 30 and 40 kg m^?2 respectively. Our results suggest that the optimal rearing density of juvenile spotted wolffish is below 40 kg m^?2 for smaller size fish (～50–100 g) and probably ≥40 kg m^?2 for the larger fish (100–160 g). Furthermore, it appears that the range of rearing density used did not have a significant effect on a selection of stress indicators (Na⁺ and K⁺ concentration, haematocrit, hepatosomatic index, total amount of plasma proteins and liver and muscle water content) and immunity response (plasma lysozyme activity) of juvenile spotted wolffish, making it a very tolerant species to crowding.     

Effects of hypoxia and hyperoxia on growth and food conversion efficiency in the spotted wolffish Anarhichas minor (Olafsen)

Abstract The effect of water oxygen content on growth and food conversion efficiency was evaluated for juvenile spotted wolffish, mean (± SD) initial weight 68.5 (± 17.5) g, reared at oxygen levels of 4.0, 6.0 (hypoxia), 9.6 (normoxia) and 14.5 (hyperoxia) mg L^?1 for 11 weeks at 8 °C. Mean weights and total food consumption were significantly higher in the control and hyperoxic groups compared with the hypoxic groups at the end of the experiment. The 9.6 and 14.5 mg L^?1 groups exhibited significantly higher overall specific growth rates (0.90 and 0.86% day^?1 respectively) compared with the groups on 4.0 and 6.0 mg L^?1 (0.46 and 0.71% day^?1 respectively). In the hyperoxic group, growth was only limited in the first period and, in the hypoxic groups, growth rates increased throughout the experiment, with the 6.0 mg L^?1 group performing equally well compared with the control in the last period. Overall, our findings suggest that the species will adapt to both high and low ambient water oxygen content given a period of adaptation. After the adaptation phase, growth and food conversion efficiency are comparable in the oxygen level range of 6.0–14.5 mg L^?1.     

Graded environmental hypercapnia in juvenile spotted wolffish (Anarhichas minor Olafsen): effects on growth, food conversion efficiency and nephrocalcinosis

Growth performance and food conversion efficiency (FCE) were investigated in juvenile spotted wolffish (Anarhichas minor Olafsen), mean (S.D.) initial weight 15.7 (4.8) g, reared at four levels of carbon dioxide (CO_2(aq)) for 10 weeks at 6 °C and 33‰. CO₂ levels averaged 1.1 (control), 18.1 (low), 33.5 (medium) and 59.4 (high) mg l⁻¹, with corresponding pH values of 8.10, 6.98, 6.71 and 6.45, respectively. In addition, kidneys from sampled fish were examined macroscopically for gross signs of calcareous deposits, i.e. nephrocalcinosis, at the start and end of the experiment. Growth was significantly reduced at the highest concentration (P<0.0001), as compared to all other groups, while no overall differences in growth rate or mean weight were seen in the range of 1.1–33.5 mg CO₂ l⁻¹ at the end of the experiment. Daily feeding rates and total food consumption were reduced at the highest concentration (P<0.001), whereas food conversion efficiency did not vary significantly between groups. Plasma chloride levels displayed a significant decrease with increasing CO₂ levels, from 151.3 mmol l⁻¹ (1.1 mg CO₂ l⁻¹) to 128.3 mmol l⁻¹ (59.4 mg CO₂ l⁻¹) at the end of the experiment, whereas plasma osmolality in the high CO₂ group was significantly higher compared to the control group at the end of the experiment (371.4 and 350.8 mOsmol kg⁻¹, respectively). Nephrocalcinosis was observed in all groups at the end of the experiment, but was most pronounced in the medium and high CO₂ group.     

Effects of photoperiod, stocking density and diet on growth in young spotted wolffish (Anarhichas minor olafsen)

Full scale experiments in tanks were conducted to elucidate the effect of photoperiod regime, dietary fat level and stocking density on growth in spotted wolffish. The study showed that continuous light (LD24:0) did not give a higher final mean weight or faster overall growth rate in spotted wolffish compared to constant 8 h light and 16 h darkness (LD8:16). However, there were indications that the fish needed time to adapt to a new photoperiod regime. The feeding experiment indicated a negative relation between dietary fat level and growth, where fish given 15% fat in the diet had a 13% higher final mean weight compared to fish on a diet with 20% fat. However, no conclusions could be made. The final mean weight was 10% higher at 25 kg/m² compared to 40 kg/m², indicating a negative impact of high stocking density on spotted wolffish.     

三丁基锡对褐菖鲉胚胎卵黄囊吸收的影响

鱼类胚胎期的营养对各种器官的发育至关重要,环境条件对营养吸收的影响会增加畸形率和死亡率。研究三丁基锡(tributyltin,TBT)对鱼类胚胎期营养吸收的影响有助于深入认识TBT对鱼类胚胎发育的毒性。本实验以海洋经济鱼类褐菖鲉(Sebastiscus marmoratus)为对象,研究环境水平TBT(0.01、0.1、1、10 ng.L-1)对胚胎卵黄囊吸收的影响。通过油红O染色,发现TBT暴露引起褐菖鲉胚胎血管系统中脂类染色信号减少,而卵黄中的脂类染色正常,说明TBT可能影响了胚胎正常血液循环,从而影响了正常的营养运输。另外,TBT对Na+,K+-ATP酶、碱性磷酸酶活性的抑制以及对甲状腺激素受体α和糖皮质激素受体基因表达的影响可能和其对卵黄囊的吸收抑制有关。TBT对鱼类胚胎卵黄吸收的抑制将影响鱼类的生长和发育,增加畸形率和死亡率,最终导致鱼类种群数量减少。     

Effects of egg disinfection and incubation temperature on early life stages of spotted wolffish

Eggs of spotted wolffish (Anarhichas minor Olafsen)were incubated at constant 4, 6 and 8 °C, and disinfected withglutaric dialdehyde (150 p.p.m. for 5 min) once ortwice a month during two thirds of the incubation period, to prevent growth ofmicroorganisms. Hatching of apparently normal larvae started earlier when eggswere disinfected twice a month compared to once a month at all incubationtemperature regimes. The time to 50% hatch was 900 and 920 day-degrees (16 and16,5 weeks) at 8 °C, 835 and 880 day-degrees (20 and 21 weeks)at 6 °C and 725 and 800 day-degrees (26 and 28,5 weeks) at 4°C, in the egg groups disinfected twice or once a month,respectively. The best survival until hatching was noted when eggs weredisinfected twice a month and incubated at 6 and 8 °C.Survivalwas very low at 4 °C. Prematurely hatched larvae wereregistered in all egg groups disinfected twice a month and the highestfrequencywas noted in the 8 °C groups. The larval weight at normalhatching in the 6 and 8 °C groups was negatively correlatedwith incubation temperature and intervals of disinfection during the incubationperiod, but after 42 days feeding with live feed (unenrichedArtemia) the weights of the larvae were not significantlydifferent. The specific growth rates of the larvae from the eggs incubated at 6°C and 8 °C were 3.0% and 3.2%, respectively.The mean survival of larvae was between 88% and 96% at 42 days post-hatching.Young wolffish originating from the 6 °C incubation groupsshowed lowest mortality.     

Influence of incubation temperature on body movements of Atlantic cod (Gadus morhua L.) embryos and on size at hatch

Body movements of cod (Gadus morhua L.) embryos reared from fertilization to hatch at 5.4°C were observed at various stages of development and at six experimental temperatures ranging from 0–10°C. Frequency of cod embryo body movements increased from zero at 42 degree‐days post fertilization to maximal at 73–82 degree‐days (1 or 2 days prior to hatch). Embryos were most active at 2°C (mean of 5.5 movements per 10 min), with activity declining to less than 1/10 min at 8–10°C. Lengths of hatched cod larvae reared at a series of constant temperatures (from 4–10°C) from fertilization to hatch were greater at lower incubation temperatures. Incubation temperatures of 2–4°C were found to be optimal for incubation of cod eggs.     

Effect of incubation temperature on the embryonic development and yolk‐sac larvae of the Pacific red snapper Lutjanus peru (Nichols & Murphy, 1922)

The effect of incubation temperature on embryonic development and yolk‐sac larva of the Pacific red snapper Lutjanus peru were evaluated by testing the effect of 26, 28 and 30°C, as this is the natural thermal interval reported during the spawning season of Pacific red snapper in the Gulf of California, Mexico. Sixteen developmental stages were observed. The incubation temperature affected the rate of development and time to hatching, being shorter at 30 than at 26°C, but no significant effect (P < 0.05) on larval length at hatching was registered. The depletion rate of yolk sac and oil globule was affected by incubation temperature particularly during the first 12 h post hatching (hph). At the end of the experiment (48 hph), significantly (P < 0.05) larger larvae were recorded at 26°C (TL = 3.22 ± 0.01 mm) than at 28° (TL = 3.01 ± 0.02 mm) and 30°C (TL = 2.97 ± 0.05 mm). Incubation of newly fertilized eggs at 26°C produces larger larvae, which may help to improve feeding efficiency and survival during first feeding.     

Influence of egg lipids and fatty acids on egg viability, and their utilization during embryonic development of spotted wolf-fish, Anarhichas minor Olafsen

The changes in egg lipids and fatty acid compositions that occur during embryonic development of spotted wolf‐fish, Anarhichas minor, were examined by monitoring individual egg batches from the time of spawning (egg stripping) until hatching. The lipids, present as 3.7±0.1% of the wet mass of the freshly stripped eggs, contained high percentages of monoenes (monounsaturated fatty acids (MUFAs), ca. 33%) and polyenes (ca. 43%) and approximately 20% saturated fatty acids (SFAs). The fatty acid profiles were dominated by a small number of fatty acids. The major SFA was 16:0 (ca. 14%), the dominant MUFA was 18:1 n‐9 (ca. 21%), and among the polyenes, the n‐3 highly unsaturated fatty acids (HUFAs) 22:6 n‐3 docosahexaenoic acid (DHA) and 20:5 n‐3 eicosapentaenoic acid (EPA) were present in the highest concentrations (EPA, ca. 16%; DHA, ca. 19%). The n‐6 HUFA 20:4 n‐6 arachidonic acid (AA) was present as ca. 1% of the total fatty acids in the freshly stripped eggs. This resulted in an AA:EPA of ca. 0.07, which is lower than reported for eggs of many other fish species. As embryonic development progressed, the percentage contribution of AA to the total fatty acids almost doubled. There were also increases in the relative proportions of SFAs (due mainly to an increase in the percentage of 16:0 to ca. 16% at hatch) and DHA (to ca. 23%), and there was a corresponding decrease in the percentage of MUFAs (mostly brought about by a decrease in the percentage of 18:1 n‐9 to ca. 18% at hatch). The most marked changes occurred towards the end of incubation. The percentage of EPA changed little during incubation. This implies that there was selective retention of DHA, 16:0 and AA, and these fatty acids were probably incorporated into cell membranes. MUFAs, particularly 18:1 n‐9, seem to have been catabolized to provide energy for the developing embryo, and some EPA also seems to have been utilized as an energy source. Survival of eggs to the eyed stage (range ca. 10–80%) and to hatch (ca. 5–75%) was negatively correlated with the %AA, %EPA and AA:DHA of the freshly stripped eggs. There was also a negative correlation between AA:EPA and egg survival, which implies that there is not a universal requirement for a high AA:EPA to ensure high rates of survival of fish eggs.     

Purification and partial characterization of vitellogenin from spotted wolffish (Anarhichas minor) and development of an enzyme-linked immunosorbent assay for the determination of gender and sexual maturity

Vitellogenin (VTG) from spotted wolffish, Anarhichas minor, a candidate species for cold-water marine aquaculture, was purified by MgCl₂/EDTA precipitation followed by a two-step chromatographic procedure. VTG had an apparent molecular mass of 470 kDa, as determined by gel filtration, and an amino acid composition similar to those of other teleosts. Sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis of the purified VTG revealed a major band with a relative molecular weight of 166 kDa and some minor bands. Spotted wolffish VTG (sw-VTG) is relatively robust to in vitro degradation, as shown when samples of purified VTG and plasma from mature females subjected to various storage conditions or multiple freeze/thaw cycles were analyzed by Western blot. We developed an indirect competitive enzyme-linked immunosorbent assay (ELISA) using an antibody against Atlantic wolffish (Anarhichas lupus) VTG and purified sw-VTG. The ELISA had a detection limit of 6.7 ng/ml and a working range of 16.2–787.5 ng/ml, with intra- and inter-assay coefficients of variation ranging from 1.5 to 7.3 % and 7.1 to 14.3 %, respectively. The assay could distinguish males from immature females and discriminate maturing females at different stage of oocyte development. These results suggest that the sw-VTG ELISA would be useful in spotted wolffish aquaculture to determine sex and monitor female maturation.     

Effects of salinity and temperature during incubation on hatching and development of lingcod Ophiodon elongatus Girard, embryos

The interactive effects of salinity and temperature on development and hatching success of lingcod, Ophiodon elongatus Girard, were studied by incubating eggs at four temperatures (6, 9, 12 and 15°C) and five salinities (15, 20, 25, 30 and 35 g L^?1). Hatch did not occur in any of the 15°C treatments. Degree days (°C days) to first hatch was not influenced by temperature or salinity, however, calendar days to first hatch differed significantly for temperature (P<0.0001, 61±1, 44±1 and 35±1 days for 6, 9 and 12°C respectively). Degree days to 50% (427.1±4.2) hatch was not significantly influenced by temperature but was by salinity (P=0.0324). Viable hatch (live with no deformities, 74.1±4.0%) was greatest at 9°C and 25 g L^?1 but not significantly different in the range of 20–30 g L^?1. Larval length (9.4±0.13 mm) was greatest at 9°C and 20–30 g L^?1. Temperature and salinity significantly influenced all categories of deformities with treatments at the upper (12°C and 35 g L^?1) and lower limits (6°C and 15 g L^?1) producing the greatest deformities. The optimal temperature and salinity for incubating Puget Sound lingcod eggs was found to be 9°C and 20–30 g L^?1.     

Combined effects of temperature and salinity on yolk utilization in Nile tilapia (Oreochromis niloticus)

The combined effects of temperature and salinity on the yolk utilization of sac fry in Nile tilapia (Oreochromis niloticus) were investigated using central composite experimental design and response surface approach. Based on the preliminary trials, temperature was determined to range from 22 to 34°C, and salinity ranging from 2 ppt to 10 ppt. The utilization was mensurated in terms of yolk sac volume. Results showed that the linear effects of temperature and salinity on the yolk utilization was significant (P < 0.01); the quadratic effects of and the interaction between the two factors were significant (P < 0.05); temperature was more important than salinity in influencing the yolk utilization. The model equation of yolk sac volume towards temperature and salinity was established. From those high R² values, the model had excellent goodness of fit to experimental data and could be applied for predictive purpose. What with the production cost, it is suggested that the temperature/salinity combination, i.e. 28–30°C/4–6 ppt, be employed during the period of sac fry rearing, in which the yolk utilization was on average 98.6%.     

The effect of temperature and duration of incubation on the hatching of diapause eggs of Centropages hamatus (Lilljeborg) (Copepoda, Calanoida)

Diapause eggs of Centropages hamatus were used to investigate the effect of temperature and duration of incubation on egg hatching. Eggs were incubated for 10, 12, 14, 16, 20, 24, 28, 32, 36 and 40 h at 15°C and 14L–10D. After incubation for the designated period, eggs were transferred to 25°C and monitored periodically to determine egg hatching. Control eggs were incubated solely at 15°C and monitored for egg hatching. The greatest daily hatching success of eggs occurred within 1 or 2 days after transfer from 15°C to 25°C, while the controls required 3–4 days. The cumulative hatching success of eggs was significantly lower than the control, with the exception of eggs held for at least 36 h at 15°C before transfer to 25°C. These results indicate that overall time to hatching of diapause eggs of C. hamatus can be reduced by transferring the eggs to a higher temperature, for example, 25°C, following a minimum period of time (36 h) at reduced temperature, for example, 15°C. Exposure to 15°C for only 10 h does not appear to be sufficient to result in any subsequent hatching at higher temperature.