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.     

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.     

Polyunsaturated fatty acid metabolism in cultured fish cells: Incorporation and metabolism of (n-3) and (n-6) series acids by Atlantic salmon (Salmo salar) cells

The incorporation and metabolism of (n-3) and (n-6) polyunsaturated fatty acids (PUFA) supplemented to growing cultures were studied in Atlantic salmon (AS) cells. A fatty acid concentration of 25 μM considerably altered the fatty acid composition of AS cells without increasing the neutral lipid content of the cells or inducing the production of cytoplasmic lipid droplets. Whereas Δ6 and Δ5 desaturase activities were significantly expressed in AS cells, Δ4 desaturase activity was very low. Both the Δ6 desaturase activity and the Δ5 desaturase activity showed some preference for (n-3) PUFA.     

Dietary vitamin C,immunity and disease resistance in Atlantic salmon (Salmo salar)

Presmolt Atlantic salmon were fed a fish meal based experimental diet supplemented with graded levels of ascorbate-2-monophosphate (AP), equivalent to 40, 400, 2000 and 4000 mg ascorbic acid (AA)/kg for 6 months prior to a bacterial challenge experiment. The liver AA concentration reflected the dietary intake of AP, but not linearly. Growth, hematology and acid phosphatase activity in zymosan stimulated macrophages were not affected by dietary AP. Serum hemolytic complement activity was higher in fish fed the highest AP level, but the variation was not significant. Production of specific antibodies was significantly higher in fish fed the highest AP level 11 and 17 weeks after vaccination. Bacterial challenge with Aeromonas salmonicida showed increased survival in the 4000 AP group. Lysozyme activity in headkidney and serum complement activity and serum iron in fish surviving the challenge were higher in the 4000 AP group, indicating important roles of vitamin C on lysozyme, complement and iron in non-specific disease resistance. The results indicate that high dietary levels of AP favourably affect health in Atlantic salmon. Disease resistance was, however, not correlated with the AA status in the liver.     

Changes in intestinal fluid transport in Atlantic salmon (Salmo salar L) during parr-smolt transformation

We examined changes in fluid transport by the intestine of Atlantic salmon (Salmo salar L) undergoing parrsmolt transformation during springtime. In vitro measurements of fluid transport rate (J_v) across non-everted middle and posterior intestinal sac preparations were made in late April and early June 1990 and from February through June 1991 for juvenile smolting fish. Intestinal J_v was also compared between parr- and smolt-stage salmon in both years. To evaluate the osmoregulatory role of the intestine, J_v was measured for smolts adapted to seawater and their cohorts remaining in fresh water. The middle intestine of smolting fish underwent a significant decrease in fluid transport during the springtime, while posterior intestinal J_v significantly increased. Parr-stage fish decreased J_v in the middle intestine during springtime similar to smolts. However, the posterior intestinal J_v of smolts showed a significant increase over the parr around the peak smolt period in both years. Seawater-adapted smolts generally exhibited posterior intestinal J_v approximately double that of freshwater cohorts. A decrease over time shown for the middle intestine, together with the increased J_v in the posterior intestine preceding and after seawater entry, suggests the development of a functional regionalization during parr-smolt transformation, with the posterior intestine taking on increased importance in osmoregulation in seawater.     

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.     

Control of phosphorus homeostasis of Atlantic salmon (Salmo salar) in fresh water

Studies were conducted to determine the absorption, excretion and requirement of dietary phosphorus (P) by Atlantic salmon (Salmo salar). Triplicate groups of salmon parr, initial weight 15 ± 0.5±g , were fed, diets containing 4, 5, 6, 7, 8, 9, 11 and 13 mg P and 20 KJ of digestible energy (DE) per±g of diet (dry matter basis, DMB) to satiation for 16 weeks. The basal diet containing 4 mg P g^-1 (0.15 mg available P per KJ DE) was supplemented with graded levels of calcium phosphate, Ca(H₂PO₄)²H₂O to formulate the eight experimental diets. The fish were reared in fresh water at a temperature of 15 °C on a 12 h photoperiod. Vertebrae ash increased from 316 to 516 mg g^-1 fat-free dry matter with an increase in dietary P content. P requirement was estimated by using a four-parameter sigmoidal equation. The data suggests that a diet of 0.28 mg available P per KJ DE is needed for Atlantic salmon. Phosphate and calcium levels in plasma and bone increased, whereas levels of magnesium and liver cholecalciferol decreased, with an increase in dietary P.Phosphate excretion in urine and apparent availability of P were determined in deficient and replete fish. In deficient fish, the urine phosphate concentration was 0.10 mmol L^-1 before feeding and 0.25 mmol l^-1 after feeding, whereas in replete fish these concentrations were 1.09 and 5.11 mmol l^-1, respectively. The increase in urine phosphate concentration was higher in replete fish than in deficient fish, however, the apparent absorption of P was found to be significantly lower in replete fish than in deficient fish. These results suggest that similarly to terrestrial vertebrates, P homeostasis in Atlantic salmon is controlled by absorption in the intestine, conservation in the kidney and storage in the bones.     

Mitochondrial and peroxisomal β-oxidation capacities in various tissues from Atlantic salmon Salmo salar

In order to investigate the capacities of different tissues to oxidize fatty acids, total β-oxidation (mitochondrial and peroxisomal) of [1–¹⁴C]palmitoyl-CoA was determined in liver and red- and white muscle from adult and juvenile Atlantic salmon Salmo salar. By including potassium cyanide (KCN) in the assay medium, it was possible to differentiate between mitochondrial and peroxisomal β-oxidation capacities. Mitochondrial β-oxidation dominated in all tissues except in livers from juvenile fish where the peroxisomal β-oxidation dominated. In general, the red muscle possesses the highest fatty acid oxidation capacity, however, by taking into consideration the fact that white muscle occupies approximately 60% of the total body weight, this study demonstrates that the white muscle is an important tissue in the overall fatty acid catabolism.     

Storage of lipids in the myosepta of Atlantic salmon (Salmo salar)

Storage of lipids in the myosepta of Atlantic salmon (Salmo salar) was determined by dissection of myosepta from both white and dark muscle, by analyzing lipid classes of myosepta, and by analysis of muscle tissues with and without myosepta. By using Iatroscan-TLC/FID it was found that triglyceride was the only major lipid class present in the myosepta. In a sample pooled from three fish, 39.1% of white muscle lipids were found to be stored in the associated myosepta. In the dark muscle up to 62.4% of the total lipids were located in the associated myosepta.After removal of the myosepta, the ratio of the total lipid content of dark muscle to white muscle decreased from 5.25 to 3.66. The proportion of polar lipids increased in both white muscle and dark muscle after removal of the myosepta; however the ratio of phosphatidyl choline (PC) to phosphatidyl ethanolamine (PE) in both muscle types remained constant. This suggests that neutral lipids accounted for most of the lipids in the myosepta. It is concluded that this type of connective tissue, not the actual muscle fibres, stores most of the muscle lipids in Atlantic salmon.     

Dietary uptake of dioxins (PCDD/PCDFs) and dioxin-like PCBs in Atlantic salmon (Salmo salar)

Atlantic salmon (Salmo salar) were fed graded levels of dioxins (polychlorinated dibenzo‐p‐dioxins and polychlorinated dibenzofurans) and dioxin‐like polychlorinated biphenyls (DLPCBs) in their diets for 7 months. The dioxin and DLPCB concentrations in both fillet and whole body of salmon increased with increasing dietary exposure. DLPCBs transferred more efficiently from the feed to edible flesh of salmon than dioxins, and contributed a higher proportion to the total toxic equivalents (TEQ). At the end of the trial, the maximum concentrations of dioxins in fillet and whole fish were 1.9 and 2.3 pg WHO‐TEQ g^?1 fresh weight, respectively. Hence with this feeding period even with the most contaminated feed (4.9 pg WHO‐TEQ g^?1 dw) the dioxin concentrations in salmon did not exceed the maximum level set by the European Commission [4 pg WHO‐TEQ g^?1 (EC 2375/2001)]. The inclusion of DLPCBs in this study provides valuable information for forthcoming risk assessments and the future establishment of maximum limits for these compounds in feed and fish.     

The level of 1C diets fed prior to cell isolation affects lipid metabolism in primary liver cells isolated from Atlantic salmon (Salmo salar)

Liver cells were isolated from 6 fish fed a diet containing 12.1 g methionine/kg, 11.02 mg vitamin B6/kg, 0.20 mg vitamin B12/kg and 7.80 mg folate/kg (named high‐1C diet). These cells were compared to liver cells isolated from 6 fish fed a diet containing 6.7 g methionine/kg, 7.01 g vitamin B6/kg, 0.15 mg vitamin B12/kg and 2.60 mg folate/kg (named low‐1C diet). Isolated cells were plated on 6‐well plates in Leibovitz medium and treated with 10 mM metformin, 10 mM metformin for 24 hr followed by 0.4 mM oleic acid (OA) for 24 hr or only 0.4 mM OA for 24 hr. The cells were compared to untreated controls added only the medium. All cells were harvested 48 hr after being plated. Cells isolated from Atlantic salmon fed low‐1C diets showed higher gene expression of MGAT‐2 (p < .0001), CPT‐1 (p = .028), FAS (p = .0006), LXR (p = .020), ACC (p = .032) and MnSOD (p < .0001). The low‐ or high‐1C diets fed prior to cell isolation had no effect on gene expression of ApoB100, PPARa, CD36, SREBP‐2 or Bcl‐2. Metformin treatment increased the expression of the anti‐apoptotic protein Bcl‐2 (p = .0001) indicating an anti‐apoptotic effect. Metformin generally increased the expression of genes associated with lipid oxidation and transport, but decreased the expression of genes associated with cholesterol metabolism confirming our earlier results using this model.     

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.     

Tissue indicators of element status in Atlantic salmon (Salmo salar) post-smolts: effect of fasting

Two groups of Atlantic salmon post-smolts previously fed diets either unsupplemented or supplemented with Zn, Fe, Cu and Mn were fasted for 6 weeks in order to study the reliability of tissue indicators of element status during fasting. The fish were weighed at 2-week intervals and samples of whole fish liver, vertebrae, serum and eyes were taken for analyses of elements by atomic absorption spectrometry.
Body weight, per cent dry matter, condition factor and haematocrit values decreased during the fasting period. The concentration of elements in whole body remained constant, indicating that whole body is a stable indicator of element status during fasting. Other tissues seemed to be more sensitive (i.e. hepatic Fe concentration and vertebrae Zn and Mn concentrations). The serum concentrations of the elements generally decreased during fasting, especially for P and Zn. Element concentrations in whole body and most tissues during fasting were generally more dependent upon initial element status than on the fasting itself.
Care should be taken in drawing conclusions about the element status of fish based solely upon the examination of serum concentrations, and more than one tissue indicator should be used in the establishment of the element status of fish.     

Insect larvae meal as an alternative source of nutrients in the diet of Atlantic salmon (Salmo salar) postsmolt

Two insect meal (IM) products were tested as ingredient in diets for Atlantic salmon weighing ~250 g. The modern control diet contained 200 g kg^?1 fish meal (FM100), which was stepwise replaced by insect meal A (IM A) as a 25% (A25), 50% (A50) or 100% (A100) FM replacement or insect meal B (IM B) as a 25% (B25) or 100% (B100) FM replacement. Selected nutrient parameters and contaminants were measured in the diets. After 15 weeks, fish length and weight were recorded and visceral and hepatic indices calculated. Fatty acid and amino acid digestibility was calculated. Morphology of the liver, kidney, mid‐ and hind intestine was evaluated. Finally, a sensory testing of the fillets was conducted. A25, A50 and A100 performed equally well as the FM100 diet. The feed intake decreased moderately with increasing IM A inclusion; however, food conversion ratio decreased, resulting in an equal net growth of the fish. Histology did not show any differences between any of the dietary groups, and sensory testing of fillets from FM100, A100 and B25 did not reveal any significant differences in odour, flavour/taste or texture between groups. IM A was suitable to replace up to 100% of dietary FM, whilst IM B did not perform equally well.     

Gene expression responses to restricted feeding and extracted soybean meal in Atlantic salmon (Salmo salar L.)

We assessed effects of feed restriction and inclusion of 200 g kg^?1 extracted soybean meal in the diet on gene expression in Atlantic salmon using a cDNA microarray (SFA2.0) and real‐time qPCR. The trial lasted for 54 days. Restricted feeding and soybean inclusion reduced the thermal growth coefficient by respectively 51% and 22% compared with fish fed with the fishmeal‐based control diet to satiation. Soybean decreased distal intestinal expression of lysosomal (cathepsins C, D, L, Y and Z) and extracellular proteases while genes involved in responses to cellular stress were up‐regulated. Expression changes of immune genes suggested both pro‐ and anti‐inflammatory regulation. The hepatic responses to soybean and restricted feeding were highly similar, which could be because of negative effects of soybean meal on digestion and nutrient absorption. We observed up‐regulation of ribosomal proteins and down‐regulation of genes involved in lipid and steroid metabolism. Of note, growth reduction in both study groups was associated with coordinated down‐regulation of genes involved in oxidative and cellular stress responses, metabolism of xenobiotics and protein degradation. High expression of stress‐related genes in salmon fed with the control diet suggests that maximum growth rates can be associated with health problems.     

Nutrient utilization in Atlantic salmon (Salmo salar L.) fed increased levels of fish protein hydrolysate during a period of fast growth

The present work was designed to study whether changes in dietary protein quality by means of partial inclusion of fish protein hydrolysate (FPH) would alter fish growth, feed utilization, protein retention and metabolism and fish health in general. FPH was produced after hydrolysing whole minced herring using the industrial enzyme Alcalase®. The dietary protein source, low‐temperature‐dried (LT) fishmeal nitrogen was exchanged with FPH nitrogen at six levels of inclusion ranging from 0 to 300 g kg^?1. The experimental diets were fed to post‐smolt (1+) Atlantic salmon (Salmo salar), with mean initial weight of 174 g for a period of 68 days. All diets were iso‐nitrogenous, iso‐energetic and contained the same amount of amino acids. Fish fed medium inclusion of FPH (180–240 g kg^?1) showed a tendency to have higher feed intake than fish fed lower and higher levels of FPH inclusions. Significant higher individual specific growth rates were present in fish fed diets with 180 and 240 g kg^?1 FPH when compared with those fed 300 g kg^?1. Feed conversion ratio increased significantly (R² = 0.61) and protein efficiency ratio decreased significantly (R² = 0.59) in fish fed increased levels of FPH. Further, apparent digestibility of crude protein and the amino acids arginine, lysine, methionine and phenylalanine increased significantly with increased dietary inclusion of FPH. Plasma free amino acids, ammonium and urea indicated that FPH amino acids was absorbed earlier and nonsynchronously, and may thus be more prone to be catabolized than in those fish fed the less solubilized protein. FPH inclusion did not have an impact on fish health, as evaluated by haematology and clinical parameters.     

Laboratory evaluation of sample collection methods (organs vs swabs) for Tasmanian salmon reovirus detection in farmed Atlantic salmon,Salmo salar L.

The use of swabs relative to organs as a sample collection method for the detection of Tasmanian salmon reovirus (TSRV) in farmed Tasmanian Atlantic salmon, Salmo salar L., was evaluated by RT‐qPCR. Evaluation of individual and pooled sample collection (organs vs swabs) was carried out to determine the sensitivity of the collection methods and the effect of pooling of samples for the detection of TSRV. Detection of TSRV in individual samples was as sensitive when organs were sampled compared to swabs, and in pooled samples, organs demonstrated a sensitivity of one 10‐fold dilution higher than sampling of pooled swabs. Storage of swabs at 4 °C for t = 24 h demonstrated results similar to those at t = 0. Advantages of using swabs as a preferred sample collection method for the detection of TSRV compared to organ samples are evident from these experimental trials.     

The minimum dietary requirement of vitamin C in Atlantic salmon (Salmo salar) fry using Ca ascorbate-2-monophosphate as dietary source

The minimum dietary vitamin C requirement for optimal growth and normal development in Atlantic salmon (Salmo salar) fry at the onset of feeding was studied, using Ca ascorbate-2-monophosphate (AP) as dietary source. The requirement was established by means of a feeding study lasting for 23 weeks from the beginning of feeding. The practical diets used were supplemented with AP at levels of 0, 10, 20, 40, 80 and 160 mg ascorbic acid (AA) equivalents/kg. Growth, mortality, hydroxyproline content in skin and backbone, and AA in liver were recorded to evaluate the results. The results suggest that the minimum dietary requirement for optimal growth and normal development is in the range of 10–20 mg AA equivalents/kg dry diet during the period studied.