A comprehensive transcriptional body map of Atlantic salmon unveils the vital role of the intestine in the immune system and highlights functional specialization within its compartments.

The intestine is a barrier organ that plays an important role in the immune system of Atlantic salmon. The immune functions are distributed among the diffuse gut lymphoid tissue containing diverse immune cells, and other cell types. Comparison of intestinal transcriptomes with those of other organs and tissues offers an opportunity to elucidate the specific roles of the intestine and its relationship with other parts of the body. In this work, a meta-analysis was performed on a large volume of data obtained using a genome-wide DNA oligonucleotide microarray. The intestine ranks third by the expression level of immune genes after the spleen and head kidney. The activity of antigen presentation and innate antiviral immunity is higher in the intestine than in any other tissue. By comparing transcriptome profiles, intestine shows the greatest similarity with the gill, head kidney, spleen, epidermis, and olfactory rosette (descending order), which emphasizes the integrity of the peripheral mucosal system and its strong connections with the major lymphoid organs. T cells-specific genes dominate among the genes co-expressed in these tissues. The transcription signature of CD8+ (86 genes, r > 0.9) includes a master gene of immune tolerance foxp3 and other negative regulators. Different segments of the intestine were compared in a separate experiment, in which expression gradients along the intestine were found across several functional groups of genes. The expression of luminal and intracellular (lysosome) proteases is markedly higher in pyloric caeca and distal intestine respectively. Steroid metabolism and cytochromes P450 are highly expressed in pyloric caeca and mid intestine while the distal intestine harbors genes related to vitamin and iron metabolism. The expression of genes for antigen presenting proteins and immunoglobulins shows a gradual increase towards the distal intestine.

Soya saponins and prebiotics alter intestinal functions in Ballan wrasse (Labrus bergylta).

A 5-week feeding trial was conducted in the cleaner fish Ballan wrasse (Labrus bergylta) for a better understanding of the basic biology of the intestinal functions and health in this stomach less species. During the trial, Ballan wrasse was fed either a reference diet, the reference diet supplemented with (i) a commercial prebiotic (Aquate™ SG, 0·4 %) expected to have beneficial effects, (ii) soya saponins (0·7 %) expected to induce inflammation or (iii) a combination of the prebiotics and the soya saponins to find a remedy for gut inflammation. Blood, intestinal tissue and gut content from four consecutive intestinal segments (IN1 - IN4) were collected. No significant differences in fish growth were observed between the four dietary groups. Saponin supplementation, both alone and in combination with prebiotics, increased weight index of IN2 and IN3 and decreased blood plasma glucose, cholesterol and total protein. Dry matter of intestinal content and activity of digestive enzymes were not affected by diet. Histomorphological analyses revealed a progressing inflammation with increased infiltration by immune cells particularly into the distal parts of the intestine in fish fed diets with saponins, both alone and in combination with prebiotics. Gene expression profiles obtained by RNA sequencing and quantitative PCR mirrored the histological and biochemical changes induced by the saponin load. The study demonstrated that Ballan wrasse gut health and digestive function may be markedly affected by feed ingredients containing antinutrients.

Effects of functional ingredients on gut inflammation in Atlantic salmon (Salmo salar L).

Functional feed ingredients are frequently used in feeds for Atlantic salmon, often claimed to improve immune functions in the intestine and reduce severity of gut inflammation. However, documentation of such effects is, in most cases, only indicative. In the present study effects of two packages of functional feed ingredients commonly used in salmon production, were evaluated employing two inflammation models. One model employed soybean meal (SBM) as inducer of a severe inflammation, the other a mixture of corn gluten and pea meal (CoPea) inducing mild inflammation. The first model was used to evaluate effects of two packages of functional ingredients: P1 containing butyrate and arginine, and P2 containing ß-glucan, butyrate, and nucleotides. In the second model only the P2 package was tested. A high marine diet was included in the study as a control (Contr). The six diets were fed to salmon (average weight of 177g) in saltwater tanks (57 fish per tank), in triplicate, for 69 days (754 ddg). Feed intake was recorded. The growth rate of the fish was high, highest for the Contr (TGC: 3.9), lowest for SBM fed fish (TGC: 3.4). Fish fed the SBM diet showed severe symptoms of inflammation in the distal intestine as indicated by histological, biochemical, molecular, and physiological biomarkers. The number of differently expressed genes (DEG) between the SBM and Contr fed fish was 849 and comprised genes indicating alteration in immune functions, cellular and oxidative stress, and nutrient digestion, and transport functions. Neither P1 nor P2 altered the histological and functional symptoms of inflammation in the SBM fed fish importantly. Inclusion of P1 altered expression of 81 genes, inclusion of P2 altered 121 genes. Fish fed the CoPea diet showed minor signs of inflammation. Supplementation with P2 did not change these signs. Regarding composition of the microbiota in digesta from the distal intestine, clear differences regarding beta-diversity and taxonomy between Contr, SBM, and CoPea fed fish were observed. In the mucosa the microbiota differences were less clear. The two packages of functional ingredients altered microbiota composition of fish fed the SBM and the CoPea diet towards that of fish fed the Contr diet.

Dietary Fish Meal Level and a Package of Choline, ß-Glucan, and Nucleotides Modulate Gut Function, Microbiota, and Health in Atlantic Salmon (Salmo salar, L.).

Steatosis and inflammation have been common gut symptoms in Atlantic salmon fed plant rich diets. Choline has recently been identified as essential for salmon in seawater, and ß-glucan and nucleotides are frequently used to prevent inflammation. The study is aimed at documenting whether increased fishmeal (FM) levels (8 levels from 0 to 40%) and supplementation (Suppl) with a mixture of choline (3.0 g/kg), ß-glucan (0.5 g/kg), and nucleotides (0.5 g/kg) might reduce the symptoms. Salmon (186 g) were fed for 62 days in 16 saltwater tanks before samples were taken from 12 fish per tank for observation of biochemical, molecular, metabolome, and microbiome indicators of function and health. Steatosis but no inflammation was observed. Lipid digestibility increased and steatosis decreased with increasing FM levels and supplementation, seemingly related to choline level. Blood metabolites confirmed this picture. Genes in intestinal tissue affected by FM levels are mainly involved in metabolic and structural functions. Only a few are immune genes. The supplement reduced these FM effects. In gut digesta, increasing FM levels increased microbial richness and diversity, and changed the composition, but only for unsupplemented diets. An average choline requirement of 3.5 g/kg was indicated for Atlantic salmon at the present life stage and under the present condition.

Bile components affect the functions and transcriptome of the rainbow trout intestinal epithelial cell line RTgutGC.

The concomitant increase in cultivation of fish and decrease in supply of marine ingredients, have greatly increased the demand for new nutrient sources. This also regards so-called functional ingredients which may benefit health and welfare of the fish. In vitro cell line-based intestinal epithelial barrier models may serve as tools for narrowing down the broad range of ingredient options, to identify the most promising candidates before in vivo feeding trials are run. In vivo, differentiation of the various epithelial cells in the fish intestine, from the multipotent stem cells, takes place in the presence of a variety of substances from dietary and endogenous origin. Among these, bile salts have recently received attention as regulators of epithelial function in health and disease but have not, until now, been included in the medium when culturing fish gut epithelial cells in vitro. As bile salts are present at high levels in the chyme of the fish intestine, in particular in salmon and rainbow trout, mostly as taurocholate (>90%), their role for effects of diet ingredients on the in vitro gut cell model should be understood. With this study, we wanted to investigate whether inclusion of bile from rainbow trout or pure taurocholate in the culture media would modulate functions of the RTgutGC epithelial cells. Here, we demonstrated that the rainbow trout intestinal epithelial cell line RTgutGC responded significantly to the presence of bile components. Treatment with rainbow trout bile taken from the gall bladder (RTbile) or pure taurocholate (TC) at taurocholate concentrations of ≤0.5 mg/mL retained normal cell morphology, cell viability as in cell oxidation-reduction metabolic activity and membrane integrity, and barrier features, while high concentrations of bile salts (≥1 mg/mL) were cytotoxic to the cells. After long-term (4 days) bile treatment, transcriptome responses showed how bile salts play important roles in intestinal epithelial cell metabolism. qPCR data demonstrated that barrier function genes, brush border enzyme genes and immune genes were significantly affected. Although similar trends were seen, treatment with bile salt as a component of rainbow trout bile or pure taurocholate, induced somewhat different effects. In conclusion, this study clearly indicates that bile salts should be included in the cell medium when running in vitro studies of gut cell functions, not at least immune functions, preferably at the level of ∼0.5 mg/mL supplemented as pure taurocholate to ensure reproducibility.

The plasma metabolome of Atlantic salmon as studied by 1H NMR spectroscopy using standard operating procedures: effect of aquaculture location and growth stage.

INTRODUCTION: Metabolomics applications to the aquaculture research are increasing steadily. The use of standardized proton nuclear magnetic resonance (1H NMR) spectroscopy can provide the aquaculture industry with an unbiased, reproducible, and high-throughput screening tool, which can help to diagnose nutritional and disease-related metabolic disorders in farmed fish. OBJECTIVE: Standard operating procedures developed for analysing (human) plasma by 1H NMR were applied to fingerprint the metabolome in plasma samples collected from Atlantic salmon. The aim was to explore the metabolome of salmon plasma in relation to growth stage and sampling site. METHODS: A total of 72 salmon were collected from three aquaculture sites in Norway (Lat. 65, 67, and 70 °N) and over two sampling events (December 2017 and November 2018). Plasma drawn from each salmon was measured by 1H NMR and metabolites were quantified using the SigMa software. The NMR data was analysed by principal component analysis (PCA) and ANOVA-simultaneous component analysis (ASCA). RESULTS: Important metabolic differences were evidenced, with adult salmon having a much higher content of very low-density lipoproteins and cholesterol in their plasma, while smolts displayed significantly higher levels of propylene glycol. Overall, 24% of the metabolite variation was due to the growth stage, whereas 12% of the metabolite variation was related to the aquaculture site and practice (p < 0.001). CONCLUSION: This study provides a baseline investigation of the plasma metabolome of the Atlantic salmon and demonstrates how 1H NMR metabolomics can be used in future investigations for comparing aquaculture practices and their influence on the fish metabolome.

Digestive and immune functions in the intestine of wild Ballan wrasse (Labrus bergylta).

This study was carried out to profile key characteristics of intestinal functions and health in wild-caught Ballan wrasse. To describe functional variation along the intestine, samples were collected from four intestinal segments, named from the proximal to the distal segment: IN1, IN2, IN3 and IN4. The sections showed quite similar structure, i.e. regarding mucosal fold height and branching, lamina propria and submucosal width and cellular composition and thickness of the muscle layers. Leucine aminopeptidase and maltase capacity decreased from IN1 to IN4, suggesting a predominant role of IN1 in digestion. Gene expression levels of vitamin C transporter (slc23a1) and fatty acid transporters (cd36 and fabp2) were higher in IN1 than in IN4, indicating a more important role of the proximal intestine regarding transport of vitamins and fatty acids. Higher expression of the gene coding for IgM heavy chain constant region (ighm) was found in IN4 than in IN1, suggesting an important immune function of the distal intestine. Other immune related genes il1b, il6, cd40, showed similar expression in the proximal and the distal part of the intestine. Parasite infection, especially the myxozoan parasite Enteromyxum leei, coincided with infiltration of lymphocytic and eosinophilic granular cells in the submucosa and lamina propria. The present study established reference information necessary for interpretation of results of studies of intestinal functions and health in cultured Ballan wrasse.

Dose-response relationship between dietary choline and lipid accumulation in pyloric enterocytes of Atlantic salmon (Salmo salar L.) in seawater.

Foamy, whitish appearance of the pyloric caeca, reflecting elevated lipid content, histologically visible as hypervacuolation, is frequently observed in Atlantic salmon fed high-plant diets. Lipid malabsorption syndrome (LMS) is suggested as term for the phenomenon. Earlier studies have shown that insufficient supply of phospholipids may cause similar symptoms. The objective of the present study was to strengthen knowledge on the role of choline, the key component of phosphatidylcholine, in development of LMS as well as finding the dietary required choline level in Atlantic salmon. A regression design was chosen to be able to estimate the dietary requirement level of choline, if found essential for the prevention of LMS. Atlantic salmon (456 g) were fed diets supplemented with 0, 392, 785, 1177, 1569, 1962, 2354, 2746 and 3139 mg/kg choline chloride. Fish fed the lowest-choline diet had pyloric caeca with whitish foamy surface, elevated relative weight, and the enterocytes were hypervacuolated. These characteristics diminished with increasing choline level and levelled off at levels of 2850, 3593 and 2310 mg/kg, respectively. The concomitant alterations in expression of genes related to phosphatidylcholine synthesis, cholesterol biosynthesis, lipid transport and storage confirmed the importance of choline in lipid turnover in the intestine and ability to prevent LMS. Based on the observations of the present study, the lowest level of choline which prevents LMS and intestinal lipid hypervacuolation in post-smolt Atlantic salmon is 3·4 g/kg. However, the optimal level most likely depends on the feed intake and dietary lipid level.

Anatomy, immunology, digestive physiology and microbiota of the salmonid intestine: Knowns and unknowns under the impact of an expanding industrialized production.

Increased industrialized production of salmonids challenges aspects concerning available feed resources and animal welfare. The immune system plays a key component in this respect. Novel feed ingredients may trigger unwarranted immune responses again affecting the well-being of the fish. Here we review our current knowledge concerning salmon intestinal anatomy, immunity, digestive physiology and microbiota in the context of industrialized feeding regimes. We point out knowledge gaps and indicate promising novel technologies to improve salmonid intestinal health.

Gut immune functions and health in Atlantic salmon (Salmo salar) from late freshwater stage until one year in seawater and effects of functional ingredients: A case study from a commercial sized research site in the Arctic region.

The present study was conducted to strengthen the knowledge on gut immune functions and health in Atlantic salmon under large scale, commercial conditions in the Arctic region of Norway. Two groups of fish were monitored, one fed a series of diets without functional ingredients (Ref) and the other diets with functional ingredients (Test). The nutritional composition of the two diet series varied in parallel according to the nutrient requirements of the fish during the observation time. The content of functional ingredients in the Test diets, i.e. nucleotides, yeast cell walls, a prebiotic and essential fatty acids, varied in accordance with a strategy developed by the feed company. The fish were observed at four sampling time points, the first (FW) in May 2016 two weeks before seawater transfer, the other three throughout the following seawater period until the fish reached a size of about 2 kg, i.e. in June, four weeks after seawater transfer (SW1); in November (SW2), and in April the following year (SW3). Gut health was assessed based on histopathological indicators of lipid malabsorption and gut inflammation, expression of gut immune, barrier and other health related genes, plasma biomarkers, somatic indices of intestinal sections, as well as biomarkers of digestive functions. Seawater transfer of the fish (SW1 compared to FW) caused a marked lowering of expression of genes related to immune and barrier functions in the distal intestine, i.e. cytokines (il1ß, il10, tgfß, ifnγ), T-cell markers (cd3γδ), myd88 and tight junction proteins (zo-1, claudin-15, claudin-25b), indicating suppressed immune and barrier functions. At SW2 and SW3, most of the immune biomarkers showed values similar to those observed at FW. The development of plasma cholesterol and triglyceride levels showed similar picture, with markedly lower levels after seawater transfer. Lipid malabsorption was observed in particular in fish from SW1 and SW2, as indicated by hyper-vacuolation of the pyloric caeca enterocytes with concurrently increased expression levels of plin2. Regarding effects of functional ingredients, significantly lower condition factor and plasma triglyceride level were observed for Test-fed fish at SW2, indicating a metabolic cost of use of a mixture of nucleotides, yeast cell walls and essential fatty acids. No clear effects of functional ingredients on expression of gut immune genes and other health indexes were observed through the observation period. The great, temporary lowering of expression of gut immune and barrier genes at SW1 is suggested to be an important factor underlying the increased vulnerability of the fish at this time point. Our findings regarding supplementation with functional ingredients raise questions whether some of these ingredients overall are beneficial or might come with a metabolic cost. Our results highlight the need for a better understanding of the cause and consequences of the suppression of gut immune functions of farmed Atlantic salmon just after seawater transfer, and the use of functional ingredients under commercial conditions.

Choline supplementation prevents diet induced gut mucosa lipid accumulation in post-smolt Atlantic salmon (Salmo salar L.).

BACKGROUND: Various intestinal morphological alterations have been reported in cultured fish fed diets with high contents of plant ingredients. Since 2000, salmon farmers have reported symptoms indicating an intestinal problem, which we suggest calling lipid malabsorption syndrome (LMS), characterized by pale and foamy appearance of the enterocytes of the pyloric caeca, the result of lipid accumulation. The objective of the present study was to investigate if insufficient dietary choline may be a key component in development of the LMS. RESULTS: The results showed that Atlantic salmon (Salmo salar), average weight 362 g, fed a plant based diet for 79 days developed signs of LMS. In fish fed a similar diet supplemented with 0.4% choline chloride no signs of LMS were seen. The relative weight of the pyloric caeca was 40% lower, reflecting 65% less triacylglycerol content and histologically normal gut mucosa. Choline supplementation further increased specific fish growth by 18%. The concomitant alterations in intestinal gene expression related to phosphatidylcholine synthesis (chk and pcyt1a), cholesterol transport (abcg5 and npc1l1), lipid metabolism and transport (mgat2a and fabp2) and lipoprotein formation (apoA1 and apoAIV) confirmed the importance of choline in lipid turnover in the intestine and its ability to prevent LMS. Another important observation was the apparent correlation between plin2 expression and degree of enterocyte hyper-vacuolation observed in the current study, which suggests that plin2 may serve as a marker for intestinal lipid accumulation and steatosis in fish. Future research should be conducted to strengthen the knowledge of choline's critical role in lipid transport, phospholipid synthesis and lipoprotein secretion to improve formulations of plant based diets for larger fish and to prevent LMS. CONCLUSIONS: Choline prevents excessive lipid accumulation in the proximal intestine and is essential for Atlantic salmon in seawater.

Gut health and vaccination response in pre-smolt Atlantic salmon (Salmo salar) fed black soldier fly (Hermetia illucens) larvae meal.

Limited availability of sustainable feed ingredients is a serious concern in salmon aquaculture. Insects may become an important, sustainable resource for expanding the raw material repertoire. Herein, we present data from an 8-week feeding trial with pre-smolt Atlantic salmon (initial body weight 49 ±â€¯1.5 g) fed either a reference diet containing fish meal, soy protein concentrate and wheat gluten as protein sources, or a test diet wherein 85% of the protein was supplied by black soldier fly larvae meal. Possible diet effect on the systemic immune response was evaluated by measuring plasma antibody titers after vaccination against infectious pancreatic necrosis virus (IPNV). The gut health of fish was evaluated using endpoints including organ and tissue indices, histopathological parameters and gene expression. Both diets induced the same level of antibody responses against IPNV. In fish fed the reference diet, the histological examination of the pyloric caeca mucosa showed clear hyper-vacuolization suggestive of lipid accumulation in enterocytes, whereas this was less pronounced in the insect meal fed fish. Expression of genes relevant to lipid metabolism confirmed these histological findings. Immune and barrier-function gene expression profiles were both generally not affected by diet. However, the fish fed insect meal showed increased expression of genes indicative of stress response, immune tolerance and increased detoxification activity. In summary, our results showed no indications that dietary inclusion of insect meal affected the gut health of Atlantic salmon negatively. The insect meal based diet seemed to reduce excessive lipid deposition in the pyloric caeca and stimulate xenobiotic metabolism.

Alternative Protein Sources in the Diet Modulate Microbiota and Functionality in the Distal Intestine of Atlantic Salmon (Salmo salar).

The present study aimed to investigate whether alternative dietary protein sources modulate the microbial communities in the distal intestine (DI) of Atlantic salmon, and whether alterations in microbiota profiles are reflected in modifications in host intestinal function and health status. A 48-day feeding trial was conducted, in which groups of fish received one of five diets: a reference diet in which fishmeal (diet FM) was the only protein source and four experimental diets with commercially relevant compositions containing alternative ingredients as partial replacements of fishmeal, i.e., poultry meal (diet PM), a mix of soybean meal and wheat gluten (diet SBMWG), a mix of soy protein concentrate and poultry meal (diet SPCPM), and guar meal and wheat gluten (diet GMWG). Samples were taken of DI digesta and mucosa for microbial profiling using high-throughput sequencing and from DI whole tissue for immunohistochemistry and expression profiling of marker genes for gut health. Regardless of diet, there were significant differences between the microbial populations in the digesta and the mucosa in the salmon DI. Microbial richness was higher in the digesta than the mucosa. The digesta-associated bacterial communities were more affected by the diet than the mucosa-associated microbiota. Interestingly, both legume-based diets (SBMWG and GMWG) presented high relative abundance of lactic acid bacteria in addition to alteration in the expression of a salmon gene related to cell proliferation (pcna). It was, however, not possible to ascertain the cause-effect relationship between changes in bacterial communities and the host's intestinal responses to the diets.IMPORTANCE The intestine of cultivated Atlantic salmon shows symptoms of compromised function, which are most likely caused by imbalances related to the use of new feed ingredients. Intestinal microbiota profiling may become in the future a valuable endpoint measurement in order to assess fish intestinal health status and effects of diet. The present study aimed to gain information about whether alternative dietary protein sources modulate the microbial communities in the Atlantic salmon intestine and whether alterations in microbiota profiles are reflected in alterations in host intestinal function and health status. We demonstrate here that there are substantial differences between the intestinal digesta and mucosa in the presence and abundance of bacteria. The digesta-associated microbiota showed clear dependence on the diet composition, whereas mucosa-associated microbiota appeared to be less affected by diet composition. Most important, the study identified bacterial groups associated with diet-induced gut dysfunction that may be utilized as microbial markers of gut health status in fish.

Dietary l-tryptophan leaves a lasting impression on the brain and the stress response.

Comparative models suggest that effects of dietary tryptophan (Trp) on brain serotonin (5-hydroxytryptamine; 5-HT) neurochemistry and stress responsiveness are present throughout the vertebrate lineage. Moreover, hypothalamic 5-HT seems to play a central role in control of the neuroendocrine stress axis in all vertebrates. Still, recent fish studies suggest long-term effects of dietary Trp on stress responsiveness, which are independent of hypothalamic 5-HT. Here, we investigated if dietary Trp treatment may result in long-lasting effects on stress responsiveness, including changes in plasma cortisol levels and 5-HT neurochemistry in the telencephalon and hypothalamus of Atlantic salmon. Fish were fed diets containing one, two or three times the Trp content in normal feed for 1 week. Subsequently, fish were reintroduced to control feed and were exposed to acute crowding stress for 1 h, 8 and 21 d post Trp treatment. Generally, acute crowding resulted in lower plasma cortisol levels in fish treated with 3×Trp compared with 1×Trp- and 2×Trp-treated fish. The same general pattern was reflected in telencephalic 5-HTergic turnover, for which 3×Trp-treated fish showed decreased values compared with 2×Trp-treated fish. These long-term effects on post-stress plasma cortisol levels and concomitant 5-HT turnover in the telencephalon lends further support to the fact that the extrahypothalamic control of the neuroendocrine stress response is conserved within the vertebrate lineage. Moreover, they indicate that trophic/structural effects in the brain underlie the effects of dietary Trp treatment on stress reactivity.

Bile components and lecithin supplemented to plant based diets do not diminish diet related intestinal inflammation in Atlantic salmon.

BACKGROUND: The present study was undertaken to gain knowledge on the role of bile components and lecithin on development of aberrations in digestive functions which seemingly have increased in Atlantic salmon in parallel with the increased use of plant ingredients in fish feed. Post smolt Atlantic salmon were fed for 77 days one of three basal diets: a high fish meal diet (HFM), a low fishmeal diet (LFM), or a diet with high protein soybean meal (HPS). Five additional diets were made from the LFM diet by supplementing with: purified taurocholate (1.8 %), bovine bile salt (1.8 %), taurine (0.4 %), lecithin (1.5 %), or a mix of supplements (suppl mix) containing taurocholate (1.8 %), cholesterol (1.5 %) and lecithin (0.4 %). Two additional diets were made from the HPS diet by supplementing with: bovine bile salt (1.8 %) or the suppl mix. Body and intestinal weights were recorded, and blood, bile, intestinal tissues and digesta were sampled for evaluation of growth, nutrient metabolism and intestinal structure and function. RESULTS: In comparison with fish fed the HFM diet fish fed the LFM and HPS diets grew less and showed reduced plasma bile salt and cholesterol levels. Histological examination of the distal intestine showed signs of enteritis in both LFM and HPS diet groups, though more pronounced in the HPS diet group. The HPS diet reduced digesta dry matter and capacity of leucine amino peptidase in the distal intestine. None of the dietary supplements improved endpoints regarding fish performance, gut function or inflammation in the distal intestine. Some endpoints rather indicated negative effects. CONCLUSIONS: Dietary supplementation with bile components or lecithin in general did not improve endpoints regarding performance or gut health in Atlantic salmon, in clear contrast to what has been previously reported for rainbow trout. Follow-up studies are needed to clarify if lower levels of bile salts and cholesterol may give different and beneficial effects, or if other supplements, and other combinations of supplements might prevent or ameliorate inflammation in the distal intestine.

Effects of dietary plant meal and soya-saponin supplementation on intestinal and hepatic lipid droplet accumulation and lipoprotein and sterol metabolism in Atlantic salmon (Salmo salar L.).

Altered lipid metabolism has been shown in fish fed plant protein sources. The present study aimed to gain further insights into how intestinal and hepatic lipid absorption and metabolism are modulated by plant meal (PM) and soya-saponin (SA) inclusion in salmon feed. Post-smolt Atlantic salmon were fed for 10 weeks one of four diets based on fishmeal or PM, with or without 10 g/kg SA. PM inclusion resulted in decreased growth performance, excessive lipid droplet accumulation in the pyloric caeca and liver, and reduced plasma cholesterol levels. Intestinal and hepatic gene expression profiling revealed an up-regulation of the expression of genes involved in lipid absorption and lipoprotein (LP) synthesis (apo, fatty acid transporters, microsomal TAG transfer protein, acyl-CoA cholesterol acyltransferase, choline kinase and choline-phosphate cytidylyltransferase A), cholesterol synthesis (3-hydroxy-3-methylglutaryl-CoA reductase) and associated transcription factors (sterol regulatory element-binding protein 2 and PPARγ). SA inclusion resulted in reduced body pools of cholesterol and bile salts. The hepatic gene expression of the rate-limiting enzyme in bile acid biosynthesis (cytochrome P450 7A1 (cyp7a1)) as well as the transcription factor liver X receptor and the bile acid transporter abcb11 (ATP-binding cassette B11) was down-regulated by SA inclusion. A significant interaction was observed between PM inclusion and SA inclusion for plasma cholesterol levels. In conclusion, gene expression profiling suggested that the capacity for LP assembly and cholesterol synthesis was up-regulated by PM exposure, probably as a compensatory mechanism for excessive lipid droplet accumulation and reduced plasma cholesterol levels. SA inclusion had hypocholesterolaemic effects on Atlantic salmon, accompanied by decreased bile salt metabolism.

Dietary cholesterol supplementation to a plant-based diet suppresses the complete pathway of cholesterol synthesis and induces bile acid production in Atlantic salmon (Salmo salar L.).

Plants now supply more than 50 % of protein in Norwegian salmon aquafeeds. The inclusion of plant protein in aquafeeds may be associated with decreased lipid digestibility and cholesterol and bile salt levels, indicating that the replacement of fishmeal with plant protein could result in inadequate supplies of cholesterol in fish. A reduction in feed efficiency, fish growth and pathogen resistance is often observed in parallel to alterations in sterol metabolism. Previous studies have indicated that the negative effects induced by plant components can be attenuated when diets are supplemented with cholesterol. The present study evaluated the effects of dietary cholesterol supplementation (1·5 %) in Atlantic salmon fed a plant-based diet for 77 d. The weights of body, intestines and liver were recorded and blood, tissues, faeces, chyme and bile were sampled for the evaluation of effects on growth, nutrient utilisation and metabolism, and transcriptome and metabolite levels, with particular emphasis on sterol metabolism and organ structure and function. Cholesterol supplementation did not affect the growth or organ weights of Atlantic salmon, but seemed to promote the induction of cholesterol and plant sterol efflux in the intestine while suppressing sterol uptake. Cholesterol biosynthesis decreased correspondingly and conversion into bile acids increased. The marked effect of cholesterol supplementation on bile acid synthesis suggests that dietary cholesterol can be used to increase bile acid synthesis in fish. The present study clearly demonstrated how Atlantic salmon adjusted their metabolic functions in response to the dietary load of cholesterol. It has also expanded our understanding of sterol metabolism and turnover, adding to the existing, rather sparse, knowledge of these processes in fish.

Gilthead seabream (Sparus aurata) immune responses are modulated after feeding with purified antinutrients.

The present study aimed to evaluate the effects of two purified antinutrients, soy saponins and phytosterols, in an important species for Mediterranean aquaculture. For this purpose, gilthead seabream (Sparus aurata) were fed six experimental diets containing two levels of those antinutrients, alone or in combination, and a control diet, to apparent visual satiation under controlled conditions. Blood and head-kidney were collected at 7, 15 and 48 days following first feeding in order to assess immune parameters and the expression of immune-related genes. Plasma bactericidal and alternative complement pathway activities increased in fish fed antinutrients compared to fish fed the control diet during the course of the experiment, with more important changes at 7 and 48 days for bactericidal activity and at 7 and 15 days for complement values. In contrast, plasma total immunoglobulins (Ig) increased in fish fed antinutrients only at 48 days. Caspase 1 (casp1), interleukin 18 (il18), colony-stimulating factor-1 receptor (csfr) and hepcidin (hep) presented similar patterns of expression with more important changes at 7 and 48 days, while interleukin 10 (il10) and ß-defensin (def) were mainly up-regulated in fish fed antinutrients at 48 days. The level of expression of IgM increased already at 7 days in fish fed the low concentration of both saponins and phytosterols while a general up-regulation was observed at 48 days compared to fish fed the control diet. Results suggest that feeding seabream a diet with purified saponins and phytosterols, alone or in combination, induces a number of changes that are related to the development of inflammation, with most important changes in fish fed the lower phytosterols concentration.

Bile enhances glucose uptake, reduces permeability, and modulates effects of lectins, trypsin inhibitors and saponins on intestinal tissue.

Antinutritional factors (ANFs) can disrupt digestive and other intestinal functions. ANFs in soybean meal (SBM) are implicated in proliferative and inflammatory responses in the intestine of various (functionally) monogastric animals, including Atlantic salmon (Salmo salar L.). The goal of the current study was to investigate the effect of ex vivo exposure of mid and distal intestinal tissue of salmon to soybean saponins (SAP), lectin (LEC) and Kunitz' trypsin inhibitor (KTI), singly and in combination, on epithelial function, as assessed by measuring in vitro glucose uptake pathways along a glucose concentration gradient. As solubilization of SAP in the calcium-containing Ringer's solution was problematic but resolved with the addition of a physiological concentration of bile collected from the gall bladder of salmon, an evaluation of bile effects became an added element. Results indicated that bile increased baseline glucose absorption and possibly transport, and also had a protective effect on the epithelial barrier, at least partially due to taurocholate. Compared to controls, tissues exposed to LEC+bile, KTI+bile and LEC+KTI+bile exhibited increased glucose uptake at the higher glucose concentrations, apparently due to markedly increased tissue permeability. Addition of SAP, however, attenuated the response, possibly by binding bile components. SAP+bile, also in combination with LEC and/or KTI, as well as LEC, KTI and LEC+KTI without bile often reduced transcellular glucose uptake pathways, while maintaining low tissue permeability. SAP+LEC+KTI+bile, LEC and KTI caused the most marked reductions. The distal intestine was more affected, reflecting the restriction of in vivo SBM-induced inflammatory changes to this region.