Effects of diets containing linseed oil on fatty acid desaturation and oxidation in hepatocytes and intestinal enterocytes in Atlantic salmon (Salmo salar)

We hypothesized that replacing fish oil with 18:3n-3-rich linseed oil may enable salmon to maintain the levels of tissue n-3HUFA levels through a combination of increased desaturation activity and increased substrate fatty acid provision. To this end we investigated desaturation/elongation of [1-¹⁴C18:3n-3 in hepatocytes and intestinal enterocytes, and determined the extent to which 18:3n-3 was oxidized and desaturated by measuring both simultaneously in a combined assay. Salmon smolts were stocked randomly into five seawater pens and fed for 40 weeks on diets in which the fish oil was replaced in a graded manner by linseed oil. At the end of the trial, fatty acyl desaturation/elongation and oxidation activities were determined in isolated hepatocytes and intestinal enterocytes using [1-¹⁴C]18:3n-3 as substrate, and samples of liver and intestinal tissue were collected for analysis of lipid and fatty acid composition. The results showed that, despite increased desaturation of [1-¹⁴C]18:3n-3 in hepatocytes, provision of dietary 18:3n-3 did not prevent the decrease in tissue n-3HUFA in fish fed linseed oil. Intestinal enterocytes were a site of significant fatty acid desaturation but, in contrast to hepatocytes, the activity was not increased by feeding linseed oil and was generally lower in fish fed linseed oil compared to fish fed only fish oil. In contrast, oxidation of [1-¹⁴C]18:3n-3 in enterocytes was generally increased in fish fed linseed oil compared to fish fed the diet containing only fish oil. However, oxidation of [1-¹⁴C]18:3n-3 in hepatocytes was 4- to 8-fold lower than in enterocytes and was not affected by diet. Furthermore, oxidation of [1-¹⁴C]18:3n-3 in enterocytes exceeded desaturation irrespective of dietary treatment, whereas similar amounts of [1-¹⁴C]18:3n-3 were desaturated and oxidized in hepatocytes from fish fed only fish oil and desaturation exceeded oxidation by 3-fold in fish fed the diet containing 100% linseed oil. The molecular mechanisms underpinning these results were discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Effects of dietary lipid level and vegetable oil on fatty acid metabolism in Atlantic salmon (Salmo salar L.) over the whole production cycle

Changes in fatty acid metabolism in Atlantic salmon (Salmo salar) induced by vegetable oil (VO) replacement of fish oil (FO) and high dietary oil in aquaculture diets can have negative impacts on the nutritional quality of the product for the human consumer, including altered flesh fatty acid composition and lipid content. A dietary trial was designed to investigate the twin problems of FO replacement and high energy diets in salmon throughout the entire production cycle. Salmon were grown from first feeding to around 2 kg on diets in which FO was completely replaced by a 1:1 blend of linseed and rapeseed oils at low (14–17%) and high (25–35%) dietary oil levels. This paper reports specifically on the influence of diet on various aspects of fatty acid metabolism. Fatty acid compositions of liver, intestinal tissue and gill were altered by the diets with increased proportions of C₁₈ polyunsaturated fatty acids and decreased proportions of n-3 highly unsaturated fatty acids (HUFA) in fish fed VO compared to fish fed FO. HUFA synthesis in hepatocytes and enterocytes was significantly higher in fish fed VO, whereas β-oxidation was unaltered by either dietary oil content or type. Over the entire production cycle, HUFA synthesis in hepatocytes showed a decreasing trend with age interrupted by a large peak in activity at seawater transfer. Gill cell prostaglandin (PG) production showed a possible seasonal trend, with peak activities in winter and low activities in summer and at seawater transfer. PG production in seawater was lower in fish fed the high oil diets with the lowest PG production generally observed in fish fed high VO. The changes in fatty acid metabolism induced by high dietary oil and VO replacement contribute to altered flesh lipid content and fatty acid compositions, and so merit continued investigation to minimize any negative impacts that sustainable, environmentally-friendly and cost-effective aquaculture diets could have in the future. Abbreviations: FO - fish oil; HUFA - highly unsaturated fatty acids acids (carbon chain length ≥C ₂₀ with ≥3 double bonds); LO - linseed oil; RO - rapeseed oil; VO - vegetable oil. This revised version was published online in July 2006 with corrections to the Cover Date.     

Utilization and metabolism of palmityl and oleoyl fatty acids and alcohols in caecal enterocytes of Atlantic salmon (Salmo salar L.)

The substitution of fish oil with wax ester‐rich calanoid copepod‐derived oil in diets for carnivorous fish, such as Atlantic salmon, has previously indicated lower lipid digestibility. This suggests that the fatty alcohols (FAlc) present in wax esters may be a poorer substrate for intestinal enzymes than the fatty acids (FA) in triacylglycerol (TAG), the major lipid in fish oil. The hypothesis tested was that the possible lower utilization of dietary FAlc by salmon enterocytes is at the level of uptake and that subsequent intracellular metabolism was identical to that of FA. A dual‐labelled FAlc–FA metabolism assay was employed to determine simultaneous FAlc and FA uptake and relative utilization in enterocytes isolated from pyloric caeca of Atlantic salmon fed either a diet supplemented with fish oil or wax ester‐rich Calanus oil. The diets were fed for 10 weeks before caecal enterocytes from each dietary group were isolated and incubated with equimolar mixtures of either [1‐¹⁴C]16:0 FA and [9,10(n)‐³H]16:0 FAlc, or [1‐¹⁴C]18:1n‐9 FA and [9,10(n)‐³H] 18:1n‐9 FAlc. Uptake was measured after 2 h with relative utilization of labelled FAlc and FA calculated as a percentage of uptakes. Differences in uptake were observed, with FA showing higher uptake than FAlc, and 18:1 chains a higher uptake than 16:0. A proportion of unesterified FAlc was possibly recovered in the cells, but the majority of FAlc was recovered in lipid classes such as TAG and phospholipids indicating substantial conversion of FAlc to FA followed by esterification. However, incorporation of FA and FAlc into esterified lipids was higher when derived from FA than from FAlc. Twenty‐five to fifty percentage of the absorbed 16:0 FA was recovered in TAG fraction of the enterocytes compared with 15–75% of 18:1 FA. Twenty to thirty percentage of the absorbed 16:0 FA was recovered in the phosphatidylcholine fraction of the enterocytes compared with only 5–15% of the 18:1 FA. Less than 15% of the fatty chains taken up by the cells were used for energy production, with significantly higher oxidation of 18:1 in enterocytes from fish fed the fish oil diet compared with the Calanus oil diet. However, overall, dietary copepod oil had little effect on FAlc and FA metabolism. Metabolic modification by elongation and/or desaturation was generally low at 1–5% of the uptake. We conclude that our hypothesis was generally proved in that the uptake of FAlc by salmon enterocytes was lower than the uptake of FA and that subsequent intracellular metabolism of FAlc was similar to that of FA. However, unesterified FAlc was possibly recovered in the cells suggesting that the conversion to FA may not be concomitant with uptake.     

Lake-use by juvenile Atlantic salmon (Salmo salar L.) and other salmonids in northern Norway

Abstract– The utilization of lakes, and inlet and outlet streams by juvenile Atlantic salmon ( Salmo salar L.), brown trout ( Salmo trutta L.) and Arctic charr ( Salvelinus alpinus (L.), were investigated in 16 watercourses northern Norway, all known to inhabit salmon stocks. In lakes, fish were caught by small mesh size gill nets, while in rivers fish were caught electrofishing. In the shallow littoral (0-3 m depth) there were juvenile salmon in 15 of 19 investigated lakes, juvenile trout in 17 and juvenile charr in seven. Trout dominated significantly in numbers in the shallow littoral of seven lakes, while salmon and charr dominated in three lakes each. When trout and salmon were frequent in the shallow littoral, charr was usually not present in this habitat, but were found in the profundal zone in most of the lakes. Atlantic salmon parr utilized both shallow and deep lakes, and used both stones and macrophytic vegetation as shelter. The utilization of lakes by salmon parr seemed to be closely related to utilization of small inlet streams for spawning. In most inlet and outlet streams salmon dominated over trout in numbers, while charr were absent. This is the first documentation of lake-use by naturally occurring salmon parr in Scandinavia.     

Transport of alpha-tocopherol in Atlantic salmon (Salmo salar) during vitellogenesis

The transport of α-tocopherol was studied during vitellogenesis in Atlantic salmon that were fed diets with two levels of α-tocopherol. α-Tocopherol levels were measured in the flesh, liver, ovary and serum, and in the serum the α-tocopherol levels in the very low density lipoprotein (VLDL), low density lipoprotein (LDL), high density lipoprotein (HDL) and very high density lipoprotein (VHDL or vitellogenin) were also measured. Atlantic salmon store α-tocopherol mainly in their flesh because the muscle mass comprises 50% or more of live weight. During vitellogenesis the α-tocopherol content declined to about 10% of the level prior to maturation. The relative range of level of α-tocopherol in the lipoproteins was: HDL> LDL> VLDL> VHDL, irrespective of dietary levels of α-tocopherol. From the recent knowledge on lipid transport during vitellogenesis and the present data, we hypothesize that α-tocopherol is transported from peripheral tissues to liver by HDL and further transported from liver to ovary by LDL. Vitellogenin appears to play a minor role in the transportation of vitamin E to the ovary.     

The effect of dietary vitamin A on the immunocompetence of Atlantic salmon (Salmo salar L.)

Atlantic salmon, Salmo salar L. were maintained on diets containing low (0.37 mg kg^–1 diet), normal (1.95 mg kg^–1 diet) and high (15 mg kg^–1 diet) levels of vitamin A fed at 1.5% body weight per day. After 4 months, liver vitamin A levels reflected dietary intake and growth rates of all three groups were similar. Kidney leucocyte migration and serum bactericidal activity were found to be significantly reduced in fish fed low levels of vitamin A. On the other hand, high levels of vitamin A in the diet were found to augment serum antiprotease activity relative to the levels found in the other dietary groups. However, phagocyte respiratory burst activity, bactericidal activity and eicosanoid production were unaffected by the dietary vitamin A regime, as were lymphocyte functions (lymphokine and antibody production) and both serum lysozyme and classical complement activity. That the overall immunomodulatory effect of vitamin A was small was reflected in the resistance to Aeromonas salmonicida. No significant differences were found between the different vitamin A intake groups despite a trend to decreased resistance in the low vitamin A diet group.     

Protein-nitrogen flux and protein growth efficiency of individual Atlantic salmon (Salmo salar L.)

Protein-nitrogen flux (the proportions of consumed and absorbed protein-nitrogen partitioned into protein synthesis and growth) was examined in Atlantic salmon, Salmo salar L. Salmon were held in groups and fed high or low rations or starved. Individual food consumption rates were measured using radiography. Fish varied widely in protein growth efficiency (protein growth divided by protein consumption), but this did not correlate with consumption rate, digestive capacity (as measured by absorption efficiency, trypsin levels and pyloric caecal size) or feeding hierarchy rank. Protein synthesis rates, measured in whole-animals, were linearly correlated with protein consumption and assimilation. There was a significant correlation between protein growth efficiency and the efficiency of retention of synthesised proteins. The capacity for protein synthesis and RNA activity were positively correlated with rates of food consumption and growth but were not correlated with protein growth efficiency. It was concluded that individual differences in protein growth efficiency related to differences in synthesis retention efficiency, but not to differences in the capacity for protein synthesis, RNA activity, digestive capacity or feeding hierarchy rank.     

Effect of low sea water temperature on water balance in the Atlantic salmon, (Salmo salar L.)

The water balance in Atlantic salmon (Salmo salar L.) overwintering in sea water (34 ) was investigated. With a decrease of temperature from 5.6 to 1.0°C the drinking rate decreased from 13.9 to 5.7 ml/kg/day, and the absolute amount of water absorbed decreased from 8.9 to 5.0 ml/kg/day. A decrease in temperature led, however, to an increase in the proportion of water absorbed in the intestines from 60 to 96%. Blood serum osmolarity increased from 320 to 440 mosm/1 with decreasing temperature and there was a reduction in tissue water content from 75 to 69% The disturbance of water balance at low temperature may be one of the factors responsible for mortality of salmon overwintering in sea water.     

Stress responses of Atlantic salmon (Salmo salar L.) elicited by water level reduction in rearing tanks

The Atlantic salmon (Salmo salar) stress response was examined by measuring plasma cortisol, glucose and chloride in fish after water level reduction within rearing tanks. Maximum plasma cortisol levels (366±43 (SD) nM and 534±280 nM for Groups 1 and 2, respectively) were observed 20 min after application of the stressor. Cortisol levels were down to control levels 24 h later. The pattern for cortisol changes observed within these two groups were comparable. Plasma glucose and chloride concentrations did not change significantly in either of the experimental groups. The fish in group 2 were then repeatedly subjected to the same stressor every third day. After the fifth exposure to the stressor, blood was obtained from fish at times corresponding to sampling after the first exposure. Their maximum plasma cortisol level only reached 223 (±96) nM, and was down to prestress levels within 2 h. Plasma glucose and chloride concentrations did not change significantly in this second experiment. This challenge test revealed an acute primary response in Atlantic salmon without any apparent harmful secondary responses, that may thus serve as a standardized reference stressor using other fish groups under comparable conditions.     

Influence of dietary fatty acids on the lipid composition of lipoproteins in farmed Atlantic salmon (Salmo salar)

The dietary influence on the fatty acid composition of neutral lipids and phosphatidylcholine of very low density lipoprotein (VLDL), low density lipoprotein (LDL) and high density lipoprotein (HDL) of Atlantic salmon (Salmo salar) was studied, using soybean oil, capelin oil and sardine oil as lipid sources in the diets. The fish had a mean weight of 3 Kg and had been fed the experimental diets for 24 months. The results show that the fatty acid composition in the feed are important for the composition of the core lipids as well as the surface components of the lipoproteins.     

Variation in commercial sources of soybean meal influences the severity of enteritis in Atlantic salmon (Salmo salar L.)

Soybean meal (SBM) is a potential alternative for the replacement of fishmeal in aquafeeds. In Atlantic salmon, however, dietary SBM causes an inflammation of the distal intestine, known as SBM-induced enteritis. The objective of the present study is to verify whether different (geographically spread) commercial sources of SBM yield contrasting inflammatory responses. To do so, six SBM batches from different origins were included in the Atlantic salmon diets at the level of 20%. After 4 weeks of feeding, the distal intestine of the salmon was sampled and scored by a semi-quantitative scoring system, which assessed six separated parameters, characterizing the extent of enteritis. The overall mean score as well as the score of the separate parameters varied between the different commercial sources of SBM included in the diet. The variation in SBM caused different degrees of disparity in the score of the separate parameters. The parameter that was most affected by the variation in the source of SBM was the disappearance of supranuclear vacuoles in enterocytes. In contrast, the increase in goblet cells showed the smallest variation between the different SBM sources. This study shows that different commercial sources of SBM can result in differences in the severity of SBM-induced enteritis in Atlantic salmon.     

Magnesium requirement of Atlantic salmon (Salmo salar L.) parr in seawater-treated fresh water1

This experiment was undertaken to establish the magnesium (Mg) requirement in young Atlantic salmon, Salmo salar L., in seawater-treated fresh water. In Norwegian hatcheries it is a common practice to add sodium hydroxide and/or sea water (1–2%) to improve pH and conductivity of the natural fresh water. Parr with initial weight of 8 g, were divided in six triplicate groups in brackish water containing 54 mg Mg L^?1 and fed a basal casein-gelatine diet supplemented with minor amounts of krill and fish meal (containing 200 mg Mg kg^?1) for an initial period of 3 weeks. Thereafter the fish were fed this diet supplemented with either 0, 100, 200, 300, 400 or 500 mg Mg kg^?1 (as MgSO₄) for 12 weeks. Growth and feed efficiency were recorded. Concentrations of Mg and other divalent cations (Ca and Zn) were measured in whole fish, serum and vertebrae. Sodium concentration in vertebrae was also measured. Growth and feed efficiency were unaffected by the levels of dietary magnesium used in the experiment. Magnesium concentrations in the whole body, serum and vertebrae Mg appeared to be more sensitive than growth and feed efficiency to differences in dietary Mg intake. The group fed the unsupplemented diet showed significantly lower Mg concentration in these tissues than the other groups. Whole-body calcium concentration was negatively correlated with dietary Mg and Ca:Mg ratios in the vertebrae were significantly affected by the dietary Mg levels. Zinc concentration in whole body, serum and vertebrae was not altered by the dietary Mg levels. Further, vertebral Na concentration did not vary between the dietary treatments. In conclusion, a minimum Mg supplementation level of 100 mg kg^?1 dry diet (in total, 326 mg kg^?1) was needed to maintain Mg concentration in the whole body and serum and for proper bone mineralization.     

Expression of insulin-like growth factor-I in the gastrointestinal tract of Atlantic salmon (Salmo salar L.)

The expression of insulin-like growth factor (IGF-I) in the gut of Atlantic salmon (Salmo salar L.) smolt was investigated at the cellular level using in situ hybridization. A hybridization protocol was adapted for use on formalin fixed, paraffin embedded tissues and was based on a digoxigenin-labelled oligoprobe that recognised a conserved region of IGF-I mRNA. The specificity of the anti-sense IGF-I probe and the performance of the hybridization protocol were evaluated using a sense IGF-I probe, an Ep stein-Barr virus probe and an insulin probe cocktail on serial sections of fish gut, mouse thyroid and human lymph node tissue. In the gut of Atlantic salmon smolt, IGF-I was found to be expressed in single epithelial cells or small clusters of epithelial cells in the pyloric ampulla and in the pyloric caeca. Expression was not detected in the lamina propria of the gut or in epithelial cells of the stomach, midgut or hindgut. The restriction of IGF-I expression to the pyloric ampulla and pyloric caeca is consistent with the role of these gut segments in osmoregulation and the scattered epithelial expression supports the putative autocrine/paracrine mechanism of action of IGF-I in osmoregulation.     

Laboratory and field investigations of salmon lice [Lepeophtheirus salmonis (Krøyer)] infestation on Atlantic salmon (Salmo salar L.) post-smolts

Hatchery‐reared 1‐year‐old Atlantic salmon post‐smolts (Salmo salar L.), artificially infected with salmon lice [Lepeophtheirus salmonis (Krøyer)] copepodids, were found to suffer from primary alterations (increased cortisol levels) at early lice stages. Secondary alterations, such as osmotic stress (increased chloride levels), first occurred after the preadult stages of the lice appeared. Fish with the highest salmon lice infections died throughout the experiment. Seven years of field investigation of Trondheimsfjorden showed that Atlantic salmon post‐smolts descending coastal waters can become heavily infected with salmon lice. The migrating post‐smolts were only infected with the chalimus stages, showing that the fish had only recently left the rivers. The infection level, however, varied considerably between the years, and, in 1998, the infection was higher than previous years. The experimental results have been combined with the field data to appraise the consequences of the infection.     

Modelling of surface and 3D pellet distribution in Atlantic salmon (Salmo salar L.) cages

Feed represents the greatest single cost factor in the production of Atlantic salmon (Salmo salar L.). Focusing on the problem of maximising the available feed for the fish while minimising the feed waste, a mathematical model of the feeding process has been developed. The model covers the feed spreader delivering the feed, water currents, pellet sinking speed and turbulent diffusion, fish appetite as a function of temperature, gut fullness and population structure, and is intended as a tool both for optimising general feeding strategies, and to support the daily handling of the feeding process. The process of horizontal and vertical diffusion of feed pellets in the model has been parametrised and validated through two low level validation experiments. Furthermore, global distribution patterns simulated by the model were verified by comparisons with experimental data from the literature.     

Influence of dietary carbohydrate on antioxidant enzyme activities in liver of Atlantic salmon (Salmo salar L.)

Effects of dietary carbohydrates on hepatic antioxidant enzymeactivities were studied in Atlantic salmon (Salmo salarL.), fed diets containing no additional carbohydrate, or a low (15% addition)orhigh (30% addition) concentration of D-glucose or gelatinised potato starch.Addition of free glucose to the feeds resulted in glucose availability being 7and 18% when added at 15 and 30%, while the addition of gelatinised starch didnotseem to reduce starch availability. The dietary treatment groups (nocarbohydrate, 7 and 18% glucose or 15 and 30% gelatinised starch) were providedwith quantities of feed that supplied the same amounts of protein and lipid.There was a positive correlation between tiobarbituric acid-reactive substances(TBARS) in feeds and salmon liver. Liver glycogen concentrations increased withincreasing dietary glucose and starch, and the two higher levels of liverglycogen, found in groups G18 and S30, resulted in decreased activities ofcatalase, superoxide dismutase (t-SOD), and concentrations of glutathione. Thisindicates that there are links between carbohydrate metabolism and antioxidantsystems in salmon liver. There was no correlation between selenium dependentglutathione peroxidase (Se-GPX) activity and dietary carbohydrateconcentrations, but Se-GPX activity increased in response to increased TBARS.Catalase and t-SOD activities did not correlate with either TBARS values orSe-GPX activity.