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Fish communities in the shallow waters (9–20 m depth) of outer Malangen (69o N), Northern Norway was studied to investigate effects of season, bottom substrate associations and population dynamic parameters on the fish assemblages. In four fieldtrips during one year at four locations, fish were sampled by trammel nets and bottom substrate composition was monitored by underwater video. A total of 16 species of fish were caught, including four species from Pleuronectidae and six from the Gadidae family. Cod (Gadus morhua Linnaeus, 1758) was the most numerous species followed by plaice (Pleuronectes platessa Linnaeus, 1758), lemon sole (Microstomus kitt Walbaum, 1792) and halibut (Hippoglossus hippoglossus Linnaeus, 1758). Cod dominated the catches in all months except March, in which plaice and other flatfishes were most dominant. On a community scale, fish species richness was highest in October and lowest in March. Furthermore, the species richness and diversity were generally higher at locations with greater bottom substrate evenness. Cod showed a positive association with large macroalgae coverage and were less numerous at locations dominated by sand and pebble. Plaice and common dab (Limanda limanda Linnaeus, 1758) showed positive associations with sand and pebble coverage, while lemon sole was more associated with maerl beds, cobble and low algae coverage. Only halibut younger than 8 years of age were observed, and the sampling areas are suggested to be nursery area for this species. Von Bertalanffy growth functions were estimated for cod, plaice and lemon sole, and the growth coefficient K of cod and plaice were higher than for populations from adjacent fjords or the North Sea populations. Total mortality rates of cod and plaice were lower than for areas further south, and this may contribute to high and relative stable biomasses in the shallow Malangen area.

Viral nervous necrosis (VNN) as a worldwide and disastrous disease could be classified as one of the most important diseases in marine fish, which occurred often in fry and juvenile fishes with high economic losses. Sometimes, it was occurred in adult fish, such as golden grey mullet in Caspian Sea. In last decade, VNN was reported in all continents except Africa; but recently, some official outbreaks were reported in wild groupers in North beaches of some African countries such as, Tunisia, Algeria, Libya, and Senegal. This infectious viral disease can be spread in marine waters or freshwater of tropical, moderate, and cold areas in fishes that can live in warm or cold waters, and also in ornamental and marine fish. So far, morbidity and sensitivity of more than 50 fish species of wild marine or reared fishes have been reported to VNN virus since 1986. Furthermore, according to official reports of OIE, VNN was recognized as most important fish health challenge in Mediterranean region and inhibitor factor in development of East-South Asian aquaculture. Regarding to epidemiological studies, first outbreak of VNN was occurred in wild red spots groupers (Epinephelus akaara) in Japan, and mortality rate reached 80% in 1991. Same mortality was reported in marine sea bass (Dicentrarchus labrax) in Greece, and mortality rate was recorded between 11% and 60% in 1997, but mortality rate in cage-reared halibut fry (Hippoglossus hippoglossus) was recorded in Norway about 100% in summer of 1997. Also, the sensitivity of Salmonidae family was confirmed to barfin flounder nervous necrosis virus serotype in Norway. It would be concluded that VNN could be considered main reason of morbidity and mortality of some important marine fish in the world. So, all aspects on VNN as most important catastrophic emerging viral disease in marine aquaculture would be discussed.

Journal homepage: http://www.sciencedirect.com/science/journal/10504648 As known from mammalia, the co-receptors CD4 or CD8 associate with a lymphocyte cell-specific kinase (Lck) upon T-cell activation. Lck phosphorylates tyrosine residues within the CD3 chains, providing docking sites for a 70 kDa zeta-associated-protein (ZAP-70), a tyrosine protein kinase important for T-cell signaling. The sequences of a CD4-like gene (CD4-2), Lck, and ZAP-70 were cloned, characterized, and the relative expression pattern was explored in several organs of Atlantic halibut (Hippoglossus hippoglossus L.). Important structural features, as a signal peptide, two Ig-like domains followed by a connecting peptide, a transmembrane region, and a CxC motif within the cytoplasmic tail were conserved within the predicted halibut CD4-2 protein. The deduced halibut Lck protein sequence was found to be composed of a N-terminal Src homology (SH) 4 domain, required for membrane attachment and CD4/CD8 binding, SH3 and SH2 adapter domains, and a SH1 domain followed by a regulatory C-terminal tail (COOH-domain). Tyrosine residues important in Lck activation were conserved within the SH1 and COOH-domain. Structural features of ZAP-70 as tandem SH2 domains and a C-terminal SH1 domain were predicted within the halibut ZAP-70 sequence, having the highest level of conservation within these regions. Several important phosphorylation sites found to play a critical role in T-cell antigen receptor signaling in mammalian were conserved. The overall expression pattern of the three genes was highly similar, showing the highest mRNA level of all three genes in thymus. Some expression was seen in spleen, anterior and posterior kidney, gills, and fin, as seen for other halibut T-cell markers. This study will enable further experiments on halibut T-cell signaling and activation, and enhance understanding about the development of immunological memory T-cells of halibut.

12 pages, 8 figures, 2 tables, Kisspeptin and its receptor, Kiss1r, play an essential role in the control of the onset of puberty in vertebrates. We characterized the cDNA and genomic DNA encoding Kiss1r in Atlantic halibut (Hippoglossus hippoglossus). The 1146 bp open reading frame predicts a 381 amino acid protein with high homology to the Kiss1r-2 of other teleost fish. Phylogenetic analysis of Kiss1r sequences suggests that the mammalian Kiss1r-1 form arose by way of a gene duplication prior to the emergence of amphibians. Synteny analysis demonstrated the highly conserved nature of the Kiss1r-2 region in teleosts, suggesting that flanking regulatory sequences are also likely to be conserved. Bioinformatic analysis identified six conserved regions in piscine Kiss1r-2 upstream sequences, providing potential targets for future in-depth investigation of Kiss1r-2 regulation. Kiss1r-2 expression in the brain increased coinciding with the onset of puberty. Expression levels in the gonads were two orders of magnitude lower than those of the brain, a characteristic apparently conserved in other fishes, and expression in gonads was only detected in immature fish, A.S.M. was supported by a predoctoral scholarship from the Spanish Ministry of Education and Science (MEC). This work was jointly supported by project “Pleurogene” from Genoma Spain, and from project CSD2007-00002 “Aquagenomics” from the Spanish Ministerio de Ciencia e Innovación

Journal homepage: http://www.elsevier.com/wps/find/journaldescription.cws_home/506046/description#description We studied the learning capacities and anticipatory behaviour in a “sit-and-wait” predatory fish, the Atlantic halibut, Hippoglossus hippoglossus. In Experiment 1 two groups of halibut received series of light flashes (conditioned stimulus, CS) that started before delivery of food (unconditioned stimulus, US) and persisted until after food delivery, i.e. delay conditioning. Control groups received unpaired CS and US presentations. The anticipatory behaviour of delay conditioned halibut consisted mainly of take-offs towards the surface shortly after onset of the CS. In Experiment 2 six groups of halibut were trained in three trace conditioning procedures: Two groups with 20 s, two groups with 60 s and two groups with 120 s trace interval. Learning was evident in the 20 and 60 s trace groups and in one of the 120 s trace groups. In contrast to delay conditioning the anticipatory behaviour of trace conditioned halibut was characterized by subtle movements near the tank floor with orientation towards the CS. The cautious responses of halibut after trace conditioning differed markedly from what is observed in other fish species and are suggested to reflect a “sit-and-wait” foraging strategy that requires the predator to remain undetected until the prey is within lunging range.

Both Atlantic cod (Gadus morhua) and Atlantic halibut (Hippoglossus hippoglossus) have been considered new candidates for aquaculture in North Atlantic countries including Norway, Canada, the UK, Iceland, and the USA since the 1980s. Differences in cultivation methods existed between these two species mainly due to the differences in life stages and their requirements of nutrition, tank dynamics, and environmental conditions. Initially, extensive cultivation methods were employed for cod and halibut using wild zooplankton; however, decades of research on intensive culture methods using rotifer and Artemia resulted in development of improved hatchery protocols. Improvements have been made in recent years in broodstock management for both species, which resulted in improved egg quality. Year around production of eggs through photo-manipulation has been achieved for cod and halibut, which is a major boast for the industry. However, a bottleneck still exists regarding broodstock nutrition, which generally contributes to inconsistent quality of eggs and larvae. Breeding programs have already been initiated for cod in Norway, Canada, and Iceland, and Ireland will follow soon. No breeding programs are available for halibut; however, a genomics program has been initiated for halibut in Canada. Research activities in the early 1990s have resulted in improved egg and yolk sac larval survival and quality in halibut. Intensive larval rearing protocols are available for both species; however, consistent production of high-quality juveniles and survival are still elusive. Apart from poor broodstock management, dependence on cultured live feed, especially Artemia often contributed to poor larval and juvenile quality. However, attempts to replace live feed with formulated diets have been partly successful for cod, and currently most hatcheries in Norway are using only rotifers in their feed regime. Further improvements in feeds and feeding strategies are warranted to reduce the dependence on live feed further. Poor husbandry practices during larval stages could also result in major loss as the immune system of the larvae is underdeveloped. The on-growing phase of both species is more stable compared to the larval phase; however, disease outbreaks, both viral and bacterial, could provide major threats. More research should be conducted on developing vaccines for these disease problems. Escapes of cod from grow-out systems have become a major problem and more research is needed on cage technology and fish behaviour to minimize these escapes. Research is also needed to produce all-female stocks and sterile fish to avoid fertilized eggs escaping to the wild. Early maturation has been recognized as the major bottleneck in cod culture and light administration appears to have limited effectiveness in preventing early maturation. Production of sterile fish would be a solution not only for minimizing fish escapees interacting with wild fish but also for minimizing early maturation. Market development need to be addressed with an emphasis on product quality and food safety.

The gelatin prepared from the skins of the Atlantic halibut (Hippoglossus hippoglossus) was investigated for the development of edible films plasticized with 30 g sorbitol/100 g gelatin. Two types of dry gelatin preparations were obtained depending on whether an intermediate evaporation step at 60 °C in the drying procedure is included or not. The amino acid composition, molecular weight distribution (determined by SDS-polyacrylamide gel electrophoresis) and glass transition temperature (determined by differential scanning calorimetry) of the gelatins were determined and related to some physical properties of the resulting films. The gelatin extracted from the halibut skins showed a suitable filmogenic capacity, leading to transparent, weakly colored, water-soluble and highly extensible films. The intermediate evaporation step at 60 °C induced thermal protein degradation, causing the resulting films to be significantly less resistant and more extensible. No differences in water vapor permeability, viscoelasticity, glass transition or color properties were evidenced between the two gelatins tested. © 2007 Elsevier Ltd. All rights reserved., This work was performed within the framework of the project CYTED XI.20. The Spanish researchers were sponsored by the Spanish Ministry of Education and Science within the project AGL2005-02380/ALI. The Brazilian authors acknowledge FAPESP with respect to the research grant (05/57781-8) and CNPq for the PIBIC fellowship awarded to M.T., and the PQI fellowship awarded to P.J.A.S.

Lipid peroxidation of marine fish diets can affect the nutritional value of the diet and consequently fish health, especially in the absence of adequate amounts of anti-oxidants. In this study, diets with different levels of oxidized oil and dietary vitamin E were fed to juvenile Atlantic halibut for 16 weeks and the effects on the acute stress response were investigated after this period. Fish were fed diets containing either non-oxidized (POV = 0.6 meq kg − 1 , diet A, control) or oxidized fish oil (POV = 7.5 meq kg − 1 , diets B and C; 15 meq kg − 1 , diets D and E). Diets A, C and E were supplemented with vitamin E (300 IU kg − 1 ). Following this period, Atlantic halibut were subjected to a 1-h heat shock (HS; from 12 to 18 °C). Plasma cortisol and glucose, and red blood cells (RBC) heat shock protein 70 (hsp70) levels were measured prior to, 0 (immediately after), 6, 12, and 24 h after stress. Two-way ANOVA, using dietary treatment and sampling point as main factors, was performed. In all experimental groups, Atlantic halibut showed increased plasma cortisol levels immediately after (0 h) heat shock, however these returned to pre-stress levels by 6 h. Similarly, plasma glucose level increased significantly immediately after heat shock and decreased to pre-stress levels by 6 h. Dietary treatment had a significant effect on plasma glucose levels. Fish fed the highly oxidized diet (diet E) showed lower overall plasma glucose levels than fish fed less or non-oxidized diets (diets A, B, and C). RBC hsp70 was detected in all treatment groups. However, no significant changes in hsp70 levels were observed after exposure to heat shock. The overall results indicate that juvenile halibut fed diets containing oxidized fish oil up to a peroxide value of 15 meq kg − 1 were able to cope with temperature stress, regardless of dietary vitamin E content. The glucose results, however, suggest that highly oxidized diets decrease the overall glucose levels. Furthermore, plasma cortisol and glucose, but not hsp70, seemed to be adequate indicators of heat shock stress in juvenile halibut.

Larval Atlantic halibut ( Hippoglossus hippoglossus ) were sampled from a commercial hatchery to observe skeletal abnormalities throughout development and to develop a metamorphic stage definition utilizing order of vertebral ossifications in combination with ossification events of the cranium. Using a whole mount cartilage and bone staining technique, skeletal abnormalities were classified and quantified into 21 types and the order of ossification events determined. During metamorphic development, stage 5 fish lacked calcified vertebral elements. Stage 6 was characterized by ossification of the exterior perimeter of vertebrae and attachment points of neural and hemal spines within the prehemal and posterior hemal regions. Further calcification of the vertebral column extends cephalad and caudad from the origin of ossification during stage 7. In stage 8, the vertebral column and caudal fin bones are weakly ossified but showing full ossification by stage 9. There was a high correlation between age ( R 2 = 0.964), standard length ( R 2 = 0.996), and myotome height ( R 2 = 0.994) with stage. Rapid development occurring over a short period of time while in the prometamorphic stages of development, may give rise to the presence of serious skeletal abnormalities in later development. The highest frequency of skeletal abnormalities occurred in the prehemal region, followed by the hemal region despite developmental stage. Hypertrophic vertebrae in the prehemal region are present in earlier developing larvae, and pre- to pro-metamorphic. Skeletal abnormalities commonly begin during stage 6 and 7, when the majority of the vertebral elements are ossifying. Studies on the immunohistochemistry and biochemical processes involved in bone growth may be useful to determine the causative factors of skeletal abnormalities.

A study was conducted to compare astaxanthin binding ability of solubilized muscle proteins of Atlantic salmon (Salmo salar L.), haddock (Melanogrammus aeglefinus L.) and Atlantic halibut (Hippoglossus hippoglossus L.). Muscle proteins of juvenile Atlantic salmon, haddock and halibut were solubilized by sequential extraction of muscle tissue using low ionic strength solutions. Electrophoretic protein profiles of the six solubilized fractions from these species were similar. Each solubilized fraction from the three species was examined for its relative astaxanthin binding capacity. The amount of bound astaxanthin was significantly different (P < 0.05) among the six fractions of each species. Significant differences in astaxanthin binding were only found for fractions A and E among the species. The amount of bound astaxanthin in various fractions of each species showed a good correlation (R2 = 0.80–0.92) with the ANS (8-anilino-1-naphthalenesulfonate) fluorescence intensity of those fractions. The pattern and extent of astaxanthin binding to the muscle proteins of juvenile salmon, haddock and halibut is comparable to that reported previously for adult Atlantic salmon [Saha, M.R., Ross, N.W., Gill, T.A., Olsen, R.E., Lall, S.P., 2005. Development of a method to assess binding of astaxanthin to Atlantic salmon S. salar L. muscle proteins. Aquacult. Res. 36, 336–343.]. These combined observations suggest that the carotenoid binding capacity of the muscle proteins of salmon is not the limiting factor in the deposition of carotenoid in their flesh.

Two forms of cytochrome P450 aromatase, acting in both the brain and the ovary, have been implicated in controlling ovarian development in fish. To better understand the expression of these two enzymes during sexual differentiation in Atlantic halibut (Hippoglossus hippoglossus), real-time PCR was used to quantify the mRNA levels of ovary- (cyp19a) and brain-type cytochrome P450 aromatase (cyp19b) genes in the gonad and brain during gonadal development. Both enzymes showed high levels of expression in both tissues in developmental stages prior to histologically detectable ovarian differentiation (38 mm fork length), with increased expression occurring slightly earlier in the brain than the gonad. Cyp19a showed a second peak of expression in later stages (> 48 mm) in the gonad, but not the brain. Cyp19b expression was generally higher in the brain than the gonad. These results suggest that sexual differentiation may begin in the brain prior to gonadal differentiation, supporting the idea that steroid hormone expression in the brain is a key determinant of phenotypic sex in fish. In an examination of sexually immature adults, cyp19a was highly expressed in female gonad while cyp19b was very highly expressed in the pituitary of both sexes. The ratio of cyp19a to cyp19b expression was much higher in ovaries than in testes in the adult fish, so this ratio was analyzed in the developing gonads of juvenile halibut in an attempt to infer their sex. This was only partially successful, with about half the fish in later developmental stages showing apparently sex-specific differences in aromatase expression.

Gynogenesis refers to a process of uniparental inheritance whereby the resulting offspring retain only maternal DNA. It has been used to identify sex-determining mechanisms in fish and to produce all-female populations for aquaculture. The objective of this study was to develop a protocol for the production of gynogenetic Atlantic halibut for both these purposes. Various milt concentrations (1:20, 1:40 and 1:80 dilutions in seminal plasma) and UV doses (0–1382 mJ/cm 2 ) were tested for providing genetically inactivated, yet motile, spermatozoa for the production of gynogenetic haploids. Spermatozoon motility, assessed both as duration of swimming and percentage of motile spermatozoa, declined with increasing UV dose. Haploidy in developing embryos was determined either visually (‘haploid syndrome’) or by genetic analysis using microsatellite DNA. The optimum milt treatment for the production of haploids was 1:80 dilution followed by a UV exposure of 65 mJ/cm 2 . Retention of the second polar body for the production of gynogenetic diploids was achieved by exposing newly activated eggs (15 min post-activation at 5–6 °C) to hydrostatic pressure of 8500 psi for 5 min. These treatments were combined to produce a population of gynogenetic diploids that was subsequently demonstrated to be comprised solely of females, thereby demonstrating that female is the homogametic sex in this species. It should therefore be possible to produce all-female populations of Atlantic halibut for commercial culture through gynogenesis or by crossing hormonally masculinized genotypic females to normal females.

1 ABSTRACT 1H nuclear magnetic resonance (NMR) spectroscopy has been investigated as a possible tool for quality evaluation of Atlantic halibut (Hipoglossus hipoglossus L). Perchloric acid extracts of muscle samples, taken from chill-stored fish over a period of three weeks, were analysed with NMR. TMAO was not found to change notably with storage time, whereas phosphocreatine was only detectable in samples taken at the day of slaughter. NADP+ was detectable in samples up to two days after slaughter. ATP and its degradation products were monitored during the whole experiment, which enabled calculations of K-values. K-values derived fiom NMR data was compared to K-value measurements with Fresh Tester. The K-values found in the NMR analyses were low during the whole period. These experiments have proven that Nh4R is a possible method for studying ATP degradation of Atlantic halibut, with the potential of monitoring several metabolites simultaneously.

Publisher Summary The Atlantic and Pacific halibut (Hippoglossus hippoglossus and H. stenolepis, respectively) are two morphologically similar flatfish forms inhabiting the boreal and subarctic waters in their respective oceans. Until recently, there were disagreements as to the taxonomic status of these two forms. More recent studies, however, using genetically determined electrophoretically detectable protein variants to test for genetic differences, have revealed a genetic difference between Atlantic and Pacific halibut of a magnitude that confirms the treatment of the two taxa as separate species. Pleuronectiform flatfishes are thought to have evolved in the Pacific Ocean in the early Tertiary period because of their greater present-day diversity in the Pacific Ocean. This chapter presents an account of the biology, fishery, and potential for aquaculture of the Atlantic species.