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The present study investigates the effects of dietary phospholipids (PL) on intestinal lipid absorption in fry and early juvenile stages of rainbow trout ( Oncorhynchus mykiss ) fed fresh or oxidized lipids. Six semi-purified casein-based diets were formulated with 12% fresh or oxidized fish oil and 6% PL supplied as soybean lecithin or egg lecithin or 6% soybean oil for PL-free diets. Each diet was distributed to 3 replicate tanks of 700 swim-up fry (initial mean body weight: 66 mg) or 200 early juvenile fish (initial mean body weight: 455 mg) over a 3-week or 17-week growth trial at a water temperature of 17 °C. Feeding oxidized compared to fresh lipids significantly reduced body weight of both fry and juvenile trout (0.15 ± 0.02 vs. 0.28 ± 0.07 g and 10 ± 2 vs. 22 ± 2 g, respectively). At fry stages, within groups fed fresh lipids, trout fry fed the PL-supplemented diets had a significantly higher final body weight than fry fed the PL-free diet (0.31 ± 0.06 vs. 0.21 ± 0.02 g). Histological examination revealed a higher occurrence of intestinal steatosis (accumulation of large lipid droplets in the enterocytes of the anterior intestine) in fish fed the PL-free diet compared to fish fed PL-supplemented diets at fry stages (13/16 vs. 1/32), whereas no significant differences were recorded at juvenile stages (1/18 vs. 0/18). The levels of circulating total cholesterol, triacylglycerol (TAG) and PL in the juvenile fish were not significantly modified by dietary PL supplementation whereas the plasma TAG/PL ratio was lower in juvenile fish fed oxidized compared to fresh lipids (0.31 ± 0.06 vs. 0.45 ± 0.08). At the fry stages, within groups fed fresh lipids, expression of the gene coding for apolipoprotein B (apoB) was 2.3-fold reduced in the intestine of fish fed the PL-free diet compared to the PL-supplemented diets whereas no significant difference was recorded at juvenile stages. The expression of other apolipoproteins or key-enzymes involved in intestinal lipoprotein synthesis was not significantly modified by dietary PL supplementation. In conclusion, the results indicate that the beneficial effect of dietary PL on growth and intestinal lipid absorption is restricted to the fry stages and support previous opinions that PL are needed for intestinal TAG absorption. The limiting step seems to be the synthesis of apoB by the enterocytes in rainbow trout fry fed fresh lipids but this requires further investigation in fish fed oxidized lipids.

Ontogenetic niche shift should occur when the ratio of growth opportunities to mortality risk becomes higher in the subsequent habitat. While most studies have focused on size to understand the timing of these shifts, an endogenous factor like energetic status (interaction between energy available and energy requirements) appears as a natural candidate to integrate and analyze the growth trade-off between habitats. In this study, we measure energetic content and metabolic rate of individual brown trout (Salmo trutta) fry at emergence from gravel to investigate the influence of energetic status on the timing of this critical ontogenetic niche shift. In addition, as offspring energetic status is subject to parental effects, we examine how females could maximize their own fitness by influencing offspring emergence timing. Our results demonstrate that emergence from gravel is influenced by energetic status. Individuals that emerge first have a higher energetic content but deplete it faster because of a higher metabolic rate. We also find that female fecundity is positively related to emergence period duration. Moreover, our results suggest that females may decrease kin competition during the critical period of emergence by influencing the energetic status of offspring, thus, maximizing their own fitness. Our results help elucidate the mechanisms underlying early ontogenetic niche shifts in juvenile fish and suggest reasons why maternal investment can be so variable within populations.

In this study, the effect of acute and chronic insulin treatments on major factors involved in the control of muscle protein turnover were investigated in rainbow trout (Oncorhynchus mykiss). We found that acute but not chronic insulin administration leads to the induction of the phosphorylation of several key factors (IRS1, TOR and 4E-BP1) involved in the control of the protein synthesis and to the concomitant down-regulation of the expression of ubiquitin–proteasome-related genes (atrogin1, C2, C9) and the calpains inhibitor calpastatin. In contrast, no modification of autophagy-related gene (LC3B, gabarpl1, atg4b) expressions was observed suggesting that the mechanisms controlling this proteolytic route have diverged throughout the evolution. Overall, these results provide a possible explanation of the growth-promoting properties of insulin previously described in fish and indicate that this hormone acutely administrated is able to exert a regulatory influence on various factors associated with growth in skeletal muscle.

SUMMARY The origin for the poor glucose utilization in carnivorous fish species fed high carbohydrate diets remains under debate. In the present study, we have fed rainbow trout a diet containing 30% carbohydrate for 1 or 5 days. In both cases, fish were implanted with mini-osmotic pumps releasing 0.7 i.u. kg–1 day–1 bovine insulin, and mRNA transcripts and the protein phosphorylation status of proteins controlling glycemia and glucose-related metabolism were studied in fish killed 6 h after the last meal. We demonstrate that when the exposure occurs over a short term (30 h), insulin exerts beneficial actions on trout glucose homeostasis, including a lowered glycemia and increased hepatic lipogenic and glycogenic potentials. However, when trout were fed for 5 days, these beneficial actions of insulin infusion were no longer observed. Thus, the increased lipogenic potential observed after one single meal was not present, and this together with the increased glycogenesis and the decreased glucose exported to the blood from the liver explains the lack of hypoglycemic action of insulin. The fact that insulin improved glucose homeostasis when administrated over a short time period implies that endogenous insulin secretion is inadequate in trout to deal with this amount of dietary carbohydrates. Moreover, the fact that a longer exposure to insulin resulted in a reduced response indicates that the rainbow trout is sensitive to insulin, re-enforcing the hypothesis that the hyperglycemia observed following a high carbohydrate meal is an insulin secretion issue rather an insulin action issue.

International audience; The abilities of triploid and diploid red blood cells to face an osmotic challenge were investigated. Erythrocytes from triploid rainbow trout were significantly more resistant to osmotic lysis than those from immature diploids of the same age. This observation could be related to the initial cell's shape and size and to the difference in cell surface area-to-volume ratio. Comparison of changes in intracellular ionic concentration (K(+), Cl(-)) associated with cell volume regulatory processes did not indicate any mechanistic explanation for this difference in osmotic resistance. This difference gradually disappeared during diploid sexual maturation. The early acquisition of high resistance to haemolysis by triploid erythrocytes is surprising since diploids appear to require time consuming and complex physiological changes to achieve a similar capability. Therefore, triploid cells appear as interesting models for investigation into cell physiology. (C) 2010 Elsevier B.V. All rights reserved.

International audience; The effects of dietary levels of phosphorus (P) and calcium (Ca) on skeletal development and mineral deposition in rainbow trout (Oncorhynchus mykiss) fry were studied. Six semi-purified diets were formulated with graded levels of P and Ca. The basal diet A contained only P supplied by casein at 0.5% of dry matter. Other cliets B,C, D and E were supplemented with 0.4, 0.8, 1.2 and 1.6% P supplied as a 1:1 mixture of NaH(2)PO(4)/KH(2)PO(4) resulting in 0.8, 1.2, 1.6 and 2.2% total P, respectively. These five diets were supplemented with 1% Ca supplied as CaCO(3) whereas another diet F, supplemented with 0.8% P,was Ca-free. Each diet was distributed to 3 replicate tanks of 600 swim-up fry (initial mean weight: 0.1 g) at a water temperature of 17 degrees C over a 12-week growth trial. Fish were handfed 6 times a day to visual satiety. There was no significant effect of dietary P (0 to 1.6%) or Ca (0 or 1%) supplementation on growth (final mean weight: 4.0 +/- 1.2 g). Survival of fish fed with diet E containing a high level of P was significantly lower compared to other groups (10% vs. 65%, respectively). Fish fed with diet A displayed lower whole-body ash and P content compared to other groups (1.98 vs. 2.49% and 035 vs. 0.46% wet weight, respectively). Phosphorus retention decreased with increasing dietary P level (from 100% in group A to about 20% in group E). The quantitative image analysis of 28-day-old fry double stained with Alcian blue and Alizarin red S revealed that individuals from group A were less ossified compared to individuals from other dietary groups. Quantitative analysis of skeletal formation using cumulative counts of endochondral and dermal structures revealed a significant delay in ossification of endochondral structures at day 11 in fish fed with mineral rich or poor diets (diets A, E and F). However, at day,28, no significant difference in ontogeny of endochondral or dermal structures was found between the trout fed with the control diet and those fed with a high P diet (E) ora low Ca diet (F), suggesting recovery is possible in rainbow trout. In conclusion, we found that both dietary deficiency and excess of P are detrimental to rainbow trout fry development: excess P affects survival, while P deficiency affects bone calcification. Calcium deficiency appears to exert some delay in ossification processes without affecting final bone mineralization. (c) 2009 Elsevier B.V. All rights reserved.

Domestic stocks may endanger wild populations by escapes or deliberate releases, not only through competition for resources but also through gene pool contamination if they reproduce in the wild. One of the risks is increased interspecific hybridization by interspecific matings. Stocking of domesticated genomes into wild ecosystems leads to a heightened frequency of interspecific hybridization in sympatric south European Atlantic salmon Salmo salar and brown trout Salmo trutta. Introgressive hybridization involving brown trout males of domestic origin was demonstrated. Following cessation of domestic stock releases in eight south European rivers, hybridization decreased significantly. These results emphasize the risks associated with stock transfers and escapes of domestic individuals in wild ecosystems and illustrate the potential role of domestic stocks when they contribute to secondary contact between species.

International audience; The objective of the present study was to identify postprandial molecular events associated with dietary fish oil level in rainbow trout using trout cDNA microarray tools (9 K). Our genomics data showed that relatively few hepatic genes (n=41) are differentially expressed between fish fed a high fat diet compared to those fed a diet without added fish oil. We then focus our analysis of specific events involved in intermediary metabolism by analysing them using qRT-PCR. Removal of fish oil was associated with higher lipid biosynthesis (reflected by the fatty acid synthase gene) and lower lipid catabolism (reflected by the acyl-CoA oxidase gene). More interestingly, some genes involved in proteasomal-dependant proteolysis (proteasome and ubiquitin ligase genes) were down regulated in fish fed without fish oil, suggesting a relationship between dietary lipid intake and regulation of hepatic protein metabolism in rainbow trout. (c) 2008 Elsevier B.V. All rights reserved.

International audience; Rainbow trout is unable to utilize high levels of dietary carbohydrates and experiences hyperglycemia after consumption of carbohydrate-rich meals. Carbohydrates stimulate hepatic glycolytic activity, but gene expression of the rate-limiting gluconeogenic enzymes glucose-6-phosphatase (G6Pase), fructose-1,6-bisphosphatase (FBPase) and phosphoenolpyruvate carboxykinase (PEPCK) remains high. Although there is significant mRNA expression and activity of gluconeogenic enzymes in trout intestine and kidney, the regulation of these enzymes by diet is not known. We tested the hypothesis that dietary carbohydrate modulates intestinal and renal G6Pase, FBPase and PEPCK. Fish were either fasted or fed isocaloric carbohydrate-free (CF) or high carbohydrate (HC) diets for 14 days. As expected, fish fed HC exhibited postprandial hyperglycemia and enhanced levels of hepatic glucokinase mRNA and activity. Dietary carbohydrates had no significant effect on the expression and activity of PEPCK, FBPase and G6Pase in all three organs. In contrast, fasting enhanced the activity, but not the mRNA expression of both hepatic and intestinal PEPCK, as well as intestinal FBPase. Therefore, the activity of rate-limiting gluconeogenic enzymes in trout can be modified by fasting, but not by the carbohydrate content of the diet, potentially causing hyperglycemia when fed high levels of dietary carbohydrates. In this species consuming low carbohydrate diets at infrequent intervals in the wild, fasting-induced increases in hepatic and intestinal gluconeogenic enzyme activities may be a key adaptation to prevent perturbations in blood glucose during food deprivation.

Interspecific relationships between Atlantic salmon and coho salmon were studied at early life stages in laboratory and semi-natural stream channels. During emergence, the survival and dispersal patterns were similar for the two species in single or mixed populations. Survival of Atlantic salmon fry was reduced in the presence of older coho fry. However, no predation was observed. Microdistribution differed between the two species, with Atlantic salmon fry more numerous in riffles when coho were present. Coho juveniles had a pelagic and gregarious distribution, in contrast to the benthic behaviour of the Atlantic salmon. In laboratory streams, Atlantic salmon fry moved out or adopted a subordinate cryptic behaviour which allowed them to escape predation while negatively affecting their growth.

SUMMARY Myostatin (GDF8) has been shown to be a major genetic determinant of skeletal muscle growth in mammals. In this study, we report the cloning of two trout cDNAs that encode two distinct myostatin-related proteins. The presence in this fish species of two myostatin genes (Tmyostatin 1 and Tmyostatin 2) probably results from the recent tetraploïdisation of the salmonid genome. A comparative reverse-transcriptase-linked polymerase chain reaction assay revealed that Tmyostatin 1 mRNA was present ubiquitously in trout tissues, while Tmyostatin 2 mRNA expression was restricted to muscle and brain. In developing muscle, Tmyostatin 1 expression was observed in eyed-stage embryos well before hatching, whereas Tmyostatin 2 was expressed only in free-swimming larvae. In myotomal muscle from adult animals, Tmyostatin 1 mRNA accumulation was similar in both slow- and fast-twitch fibres, and its concentration did not change during the muscle wasting associated with sexual maturation. In contrast, Tmyostatin 2 mRNA accumulated predominantly in slow-twitch fibres, and its concentration decreased dramatically in wasting muscles from maturing animals. This work shows that two distinct myostatin genes are present in the trout genome. Furthermore, it indicates that these two trout myostatin genes (i) exhibit a distinct expression pattern in muscle and non-muscle tissues and (ii) are not upregulated during the muscle wasting that accompanies sexual maturation.

One of the most recently identified members of the tumor necrosis factor receptor family, death receptor-6 (DR6), has been shown to mediate apoptosis following overexpression in HeLa cells. The avian and piscine orthologs of DR6 have now been identified, and the deduced amino acid sequence for each demonstrates a high level of conservation compared to the mammalian sequence. Expression of dr6 mRNA occurs widely across tissues of both the mature chicken and brook trout. It is now well-established that ovarian follicular atresia occurs via apoptosis originating within the granulosa cell layer. Accordingly, DR6 expression within the ovary was examined to assess the relationship between stage of follicle development and relative levels of this death receptor. Of particular interest was the finding that elevated levels of dr6 mRNA, as well as the translated protein, are expressed in atretic compared to healthy follicles of the hen ovary, thus providing the first association between DR6 expression and apoptosis, in vivo. We conclude that DR6 is a highly conserved and widely expressed death-domain-containing receptor and may be implicated in regulating follicle atresia within the vertebrate ovary.

The effects of independently altering chloride, calcium, and dissolved organic carbon (DOC) on the toxicity of silver were compared between rainbow trout (Oncorhynchus mykiss) and fathead minnows (Pimephales promelas). The 96-h median lethal concentration toxicity tests for both species were performed under the same conditions, within the same containers. In addition, the effect of altering [Cl{sup {minus}}] on silver-induced perturbations to body Na{sup +} influx and gill silver load was studied. Toxicity tests were conducted in synthetic soft water (50 {micro}M Na{sup +}, 50 {micro}M Cl{sup {minus}}, 50 {micro}M Ca{sup 2+}, 0.3 mg DOC/L). The [Cl{sup {minus}}], [Ca{sup 2+}], and [DOC] were adjusted by the addition of NaCl, CaNO{sub 3}, or humic acid, respectively. On the basis of total silver, increasing [Cl{sup {minus}}] over a range of 50 {micro}M to 1,500 {micro}M resulted in a 4.3-fold increase in the 96-h LC50 values (decrease in toxicity) for rainbow trout, but did not significantly affect the 96-h LC50 values for fathead minnows. Increasing water [Ca{sup 2+}] (from 50 to 2,000 {micro}M) had only a small influence on the 96-h LC50 values in both species. If the 96-h LC50 values are calculated on the basis of ionic silver, Ag{sup +}, then, in themore » case of rainbow trout, toxicity correlates to Ag{sup +}. However, this correlation does not exist for fathead minnows. Increasing [Cl{sup {minus}}] did not affect the degree of perturbation of Na{sup +} influx during acute exposure (first 4 h) to 8 {micro}g Ag/L in either species, nor did it affect the whole-body silver uptake rates, but it did reduce the gill silver load. These results demonstrate that differences exist in the way in which water chemistry ameliorates silver toxicity between rainbow trout and fathead minnows.« less

Brown trout, with indwelling dorsal aortic cannulae, were exposed to various concentrations of aluminium (Al; 50 micrograms liter-1, 100% mortality over 48 hr; 25 micrograms liter-1, 50% mortality over 120 hr; 12.5 micrograms liter-1, 0% mortality over 120 hr) in acidic (pH 5.0) soft water. The plasma concentrations of prolactin (PRL), cortisol, thyroxine (T4), and triiodothyronine (T3) were monitored. Plasma PRL concentrations were transiently depressed (to less than 20% of resting concentrations) after 12 hr in trout in the two highest water Al concentrations, but were unchanged in the trout exposed to 12.5 micrograms liter-1 Al. Plasma cortisol concentrations were elevated in response to all water Al levels and remained elevated in trout in the lethal conditions. The sublethally exposed trout showed a recovery in plasma cortisol concentrations by 120 hr. Plasma T4 concentrations were significantly elevated in trout exposed to both the lethal and the sublethal Al concentrations (from mean resting concentrations of 1-2 ng ml-1 to peaks of 8.9 and 9.0 ng ml-1 in the 50 and 12.5 micrograms liter-1 Al groups, respectively), although a recovery in plasma concentrations was evident in the sublethally exposed trout from 72 hr onwards. Plasma T3 concentrations were relatively stable in the trout exposed to the two highest doses of Al, whereas the trout under the lowest, sublethal, Al conditions exhibited a sustained (12-72 hr) elevation in plasma T3 concentrations (from a mean resting concentration of 0.9 ng ml-1 to a peak of 4.2 ng ml-1 at 48 hr). No clear relationship was apparent between the plasma PRL concentrations and the previously reported ionoregulatory status of the trout.

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In the study, the inhibitory effect of flavonoids, including isoflavonic phytoestrogens, on the ovarian aromatase enzyme complex from the rainbow trout, Oncorhynchus mykiss, was assessed in vitro. Some of the compounds tested on fish were also tested on human placental aromatase activity as a comparison between the two sources of enzyme. It was found that flavone, dl-aminoglutethimide, apigenin, quercetin, 7,4'- dihydroxyflavone, alpha-naphthoflavone and equol were potent inhibitors of the ovarian aromatase activity in rainbow trout. Relative potencies (RP) of these compounds compared to flavone (assigned an effect of 1) were, respectively, 19.0, 8.7, 5.3, 3.7, 3.2 and 0.9. Two other phytoestrogens, namely biochanin A and genistein, slightly inhibited aromatase activity. Finally, 7-hydroxyflavone, formononetin, daidzein, coumestrol, chrysin, flavanone and estradiol-17beta did not inhibit ovarian aromatase activity at doses up to 1000 microM. Experiments on human placental aromatase showed inhibitory effects of dl-aminoglutethimide, flavone, flavanone and equol with RP values of 2.8. 1, 1.5 and 0.4, respectively. These results are in accordance with previous studies. The influence of the experimental procedure on IC50 values and RP is discussed.

We report the cloning of a new trout myogenic cDNA which encodes helix-loop-helix protein homologous to the myogenic factor myogenin. Northern analyses indicate that trout myogenin (Tmyogenin) transcripts accumulate in large amounts in the myotomal masculature of embryos and frys. In adults, transcripts concentrate within the thin lateral layer of red (slow oxydative) muscle fibres. They are present only in low amounts in white (fast glycolytic) muscle fibres which constitute the major part of the trunk musculature. Using an in vitro myogenesis system, we observed that the trout myogenin encoding gene is not activated until myosatellite cells fuse to generate multinucleated myotubes, indicating that Tmyogenin lies downstream of muscle determination factors. All these observations show that in a major taxinomic group like teleosts, a gene with homology to myogenin exists. Its activation during myogenesis suggests that it acts as a major developmental regulator of muscle differentiation.

Two groups of transgenic rainbow trout (Oncorhynchus mykiss, Walbaum) have been produced and compared. One group harbored the reporter gene of chloramphenicol acetyltransferase (CAT) associated with mouse immunoglobulin (Ig) promoter/enhancer (pUCL-CAT-E). The other group carried the same reporter gene under the control of the cytomegalovirus promoter/enhancer (pCMV-CAT). Slot blot analysis of DNA from blood cells and other tissues from pUCL-CAT-E fish showed variation of copy number between the major tissues but not between red and white blood cells. Southern blot analysis indicated that multiple copies organized in concatemers were incorporated into the genome. The pCMV-CAT fish had a pronounced expression of CAT in both white and red blood cells. In contrast, activity of CAT was found in the white blood cells of all pUCL-CAT-E fish but not in their red blood cells. Expression in white blood cells was found preferentially in sIg+ cells, indicating that B cells are the major expressors. High expression was also found in spleen and kidney, but the activity found in thymocytes was equal to the background level. Analysis of some major tissues showed high white blood cell expression associated with low tissue expression, except that liver (known to contain lymphoid tissue in fish) was higher. Thus the regulatory elements of the Ig gene from mouse induce a tissue-specific expression in fish.

Wnt signaling plays major roles in various processes, including ovarian differentiation and development in mammals. In order to explore its potential implication during gonadal development in a nonmammalian vertebrate species, expression of Wnt signaling genes was investigated in rainbow trout during gonadal differentiation and gametogenesis. Multiple Wnt pathway genes were expressed and exhibited distinct expression patterns. In ovary, tcf7 was highly expressed during early differentiation, whereas no sexually dimorphic expression of rspo1 was detected. During later ovarian development, wnt11 was highly expressed in granulosa cells and oocytes suggesting an implication in folliculogenesis and oogenesis, whereas wnt9b was principally detected in granulosa cells. In testis, Wnt pathway genes were mostly expressed during early spermatogenesis. Overall, these present results suggest that Wnt signaling is implicated in multiple processes of male and female gonadal development and provide basis for future studies on Wnt signaling functions in teleost fish gonads.

International audience; In a population exhibiting partial migration (i.e. migration and residency tactics occur in the same population), the mechanisms underlying the tactical choice are still unclear. Empirical studies have highlighted a variety of factors that could influence the coexistence of resident and migratory individuals, with growth and body size considered to be key factors in the decision to migrate. Most studies suffer from at least one of the two following caveats: (1) survival and capture probabilities are not taken into account in the data analysis, and (2) body size is often used as a proxy for individual growth. We performed a capture–mark–recapture experiment to study partial migration among juvenile brown trout Salmo trutta at the end of their first year, when a portion of the population emigrate from the natal stream while others choose residency tactic. Bayesian multistate capture–recapture models accounting for survival and recaptures probabilities were used to investigate the relative role of body size and individual growth on survival and migration probabilities. Our results show that, despite an apparent effect of both size and growth on migration, growth is the better integrative parameter and acts directly on migration probability whereas body size acts more strongly on survival. Consequently, we recommend caution if size is used as a proxy for growth when studying the factors that drive partial migration in juvenile salmonid species.

Background There is increasing evidence that the ability to adapt to seawater in teleost fish is modulated by genetic factors. Most studies have involved the comparison of species or strains and little is known about the genetic architecture of the trait. To address this question, we searched for QTL affecting osmoregulation capacities after transfer to saline water in a nonmigratory captive-bred population of rainbow trout. Results A QTL design (5 full-sib families, about 200 F2 progeny each) was produced from a cross between F0 grand-parents previously selected during two generations for a high or a low cortisol response after a standardized confinement stress. When fish were about 18 months old (near 204 g body weight), individual progeny were submitted to two successive hyper-osmotic challenges (30 ppt salinity) 14 days apart. Plasma chloride and sodium concentrations were recorded 24 h after each transfer. After the second challenge, fish were sacrificed and a gill index (weight of total gill arches corrected for body weight) was recorded. The genome scan was performed with 196 microsatellites and 85 SNP markers. Unitrait and multiple-trait QTL analyses were carried out on the whole dataset (5 families) through interval mapping methods with the QTLMap software. For post-challenge plasma ion concentrations, significant QTL (P < 0.05) were found on six different linkage groups and highly suggestive ones (P < 0.10) on two additional linkage groups. Most QTL affected concentrations of both chloride and sodium during both challenges, but some were specific to either chloride (2 QTL) or sodium (1 QTL) concentrations. Six QTL (4 significant, 2 suggestive) affecting gill index were discovered. Two were specific to the trait, while the others were also identified as QTL for post-challenge ion concentrations. Altogether, allelic effects were consistent for QTL affecting chloride and sodium concentrations but inconsistent for QTL affecting ion concentrations and gill morphology. There was no systematic lineage effect (grand-parental origin of QTL alleles) on the recorded traits. Conclusions For the first time, genomic loci associated with effects on major physiological components of osmotic adaptation to seawater in a nonmigratory fish were revealed. The results pave the way for further deciphering of the complex regulatory mechanisms underlying seawater adaptation and genes involved in osmoregulatory physiology in rainbow trout and other euryhaline fishes.

International audience; As lipid deposition tissue in fish, the white adipose tissue (WAT) has important functions related to reproduction and the challenges of long-term fasting. In the study reported here, we infused fish fed a high-carbohydrate diet with two doses of insulin for 5 days in order to explore the effects of this hormone on lipogenesis and beta-oxidation-related enzymes. We demonstrated the presence of some of the main lipogenic enzymes at molecular, protein and activity levels (ATP-citrate lyase and fatty acid synthase). However, while ATP-citrate lyase was unexpectedly down-regulated, fatty acid synthase was up-regulated (at protein and activity levels) in an insulin dose-dependent manner. The main enzymes acting as NADPH donors for lipogenesis were also characterized at biochemical and molecular levels, although there was no evidence of their regulation by insulin. On the other hand, lipid oxidation potential was found in this tissue through the measurement of gene expression of enzymes involved in beta-oxidation, highlighting two carnitine palmitoyltransferase isoforms, both down-regulated by insulin infusion. We found that insulin acts as an important regulator of trout WAT lipid metabolism, inducing the final stage of lipogenesis at molecular, protein and enzyme activity levels and suppressing beta-oxidation at least at a molecular level. These results suggest that WAT in fish may have a role that is important not only as a lipid deposition tissue but also as a lipogenic organ (with possible involvement in glucose homeostasis) that could also be able to utilize the lipids stored as a local energy source.

International audience; The aim of the present study was to analyse the effects of partial or total replacement of fish meal (FM) and fish oil (FO) by a mixture of plant protein (PP) and a mixture of vegetable oils (VO) on the hepatic insulin-nutrient-signalling pathway and intermediary metabolism-related gene expression in rainbow trout (Oncorhynchus mykiss). Triplicate groups of fish were fed four practical diets containing graded levels of replacement of FM and FO by PP and VO for 12 weeks: diet 0/0 (100% FM, 100% FO); diet 50/50 (50% FM and 50% PP, 50% FO and 50% VO); diet 501100 (50% FM and 50% PP, 100% VO); diet 100/100 (100% PP, 100% VO). Samplings were performed on trout starved for 5 d then refed with their allocated diet. In contrast to partial substitution (diet 50150), total substitution of FM and FO (diet 100/100) led to significantly lower growth compared with diet 0/0. The insulin-nutrient-signalling pathway (protein kinase B (Akt), target of rapamycin (TOR), S6 protein kinase 1 (S6K1) and S6) was characterised in trout liver and found to be activated by refeeding. However, changes in diet compositions did riot differentially affect the Akt-TOR-signalling pathway. Moreover, expression of genes encoding fructose-1,6-biphosphatase, mitochondrial phosphoenolpyruvate carboxykinase, glucokinase, pyruvate kinase and carnitine palmitoyl transferase I were not affected by refeeding or by dietary changes. Refeeding down- and up-regulated the expression of gluconeogenic glucose-6-phosphatase isoform 1 and lipogenic fatty acid synthase genes, respectively. Expression of both genes was also increased with partial replacement of FM and total replacement of FO (diet 50/100). These findings indicate that plant-based diets barely affect glucose and lipid metabolism in trout.

Routine metabolic rate (oxygen consumption) of individual eggs and larvae of brown trout (Salmo trutta) originating from different families were monitored from fertilisation to the onset of emergence by means of flow through micro-respirometry. This measuring system revealed an accurate tool to measure oxygen consumption on small organisms at the individual level, and daily consumption proved to be very stable. The mass-specific metabolic rate remained low from fertilisation to hatching, and then increased quickly until the age of emergence. A Bayesian modelling approach was used to adequately infer maternal effects on metabolic rate dynamics all along the development period. Substantial differences were found between families, affecting average metabolic rate as well as intra-family variance. That is, offspring originating from different females may have different energetic needs at emergence from gravel. Moreover, between siblings, variability in metabolic rate is also under the influence of maternal effects. Implications of this metabolic rate variability are discussed with regard to life history strategies and early behaviours.

10 p., 5 tables, 7 figures and bibliography, For developing efficient diets, two sets of experiments examined whether the use and allocation of dietary protein can be traced by labelling with stable isotopes (15N and 13C) in two culture fish (Oncorhynchus mykiss and Sparus aurata). In the first experiment, natural abundance and tissue distribution of these isotopes were determined, by measuring the δ13C and δ15N values by isotopic ratio mass spectrometry, in fingerlings (14-17 g) adapted to diets differing in the percentage of fish meal replacement by plant protein sources. For both species, δ15N and δ13C were greater in tissues with higher protein and lower lipid content. Delta 15N of diets and tissues decreased as replacement increased, suggesting δ15N can be used as a marker for dietary protein origin. The 15N fractionation (δ15N fish - δ15N diet) differed between groups, and could thus be used to indicate protein catabolism. In the second experiment, fish (75-90 g) of each species ingested a diet enriched with 15N-protein (10 g kg-1 diet) and 13C-protein (30 g kg-1 diet). These proportions were suitable for determining that the delta values of tissue components were high enough above natural levels to allow protein allocation to be traced at 11 and 24 h after feeding, and revealed clear metabolic differences between species., This study was supported by EU (Q5RS-2000-30068; "Perspectives of Plant Protein Use in Aquaculture", PEPPA).

SUMMARYRainbow trout have a limited ability to use dietary carbohydrates efficiently and are considered to be glucose intolerant. Administration of carbohydrates results in persistent hyperglycemia and impairs post-prandial down regulation of gluconeogenesis despite normal insulin secretion. Since gluconeogenic genes are mainly under insulin control, we put forward the hypothesis that the transcriptional function of insulin as a whole may be impaired in the trout liver. In order to test this hypothesis, we performed intraperitoneal administration of bovine insulin to fasted rainbow trout and also subjected rainbow trout primary hepatocytes to insulin and/or glucose stimulation. We demonstrate that insulin was able to activate Akt, a key element in the insulin signaling pathway, and to regulate hepatic metabolism-related target genes both in vivo and in vitro. In the same way as in mammals, insulin decreased mRNA expression of gluconeogenic genes, including glucose 6-phosphatase (G6Pase),fructose 1,6-bisphosphatase (FBPase) and phosphoenolpyruvate carboxykinase (PEPCK). Insulin also limited the expression of carnitine palmitoyltransferase 1 (CPT1), a limiting enzyme of fatty acid β-oxidation. In vitro studies revealed that, as in mammals,glucose is an important regulator of some insulin target genes such as the glycolytic enzyme pyruvate kinase (PK) and the lipogenic enzyme fatty acid synthase (FAS). Interestingly, glucose also stimulates expression of glucokinase (GK), which has no equivalent in mammals. This study demonstrates that insulin possesses the intrinsic ability to regulate hepatic gene expression in rainbow trout, suggesting that other hormonal or metabolic factors may counteract some of the post-prandial actions of insulin.

The mRNA transcribed from the N gene of viral haemorrhagic septicaemia virus (VHSV) of salmonids has been cloned in Escherichia coli and expressed. Fusion proteins were recognized by monoclonal antibody directed against the N protein from the viral particle. A 1212 bp long open reading frame (ORF) coding for 404 amino acids with a calculated M r of 44590 was deduced from the nucleotide sequence. The ORF was preceded by a 93 bp segment including in position 42 the AACAC pentanucleotide which is presumed to be the start signal for transcription by analogy with other rhabdoviral mRNAs. The upstream 41 bp region could correspond to the covalently linked positive polarity leader RNA as also found on the N mRNA from infectious haematopoietic necrosis virus (IHNV). This may be a characteristic of fish lyssaviruses. The AAACC sequence, which is part of the leader, was not found. Amino acids 44 to 359 from IHNV and 45 to 360 from VHSV are 45.3% homologous. A strong homology which could reflect functional importance was also found for potential phosphorylation sites and hydrophobic peaks despite the fact that the two viruses evolved on different continents.

Viral haemorrhagic septicaemia virus (VHSV), a rhabdovirus, is a major threat for continental European trout fish farming. The development of a recombinant subunit vaccine could solve that problem. The neutralizing epitopes are located on the glycoprotein or G protein, the surface antigen. The G protein has a molecular weight of 65 kDa, reduced to 55 kDa by deglycosylation. cDNA was synthetized from mRNA of VHS virus infected cells, and cloned in E. coli. The viral cDNA was recognized by positive hybridization with a labelled probe made from infected cell RNA, and negative hybridization with labelled cDNA made from cellular RNA. The Northern blot hybridization with different clones on VHS infected cell RNA revealed two VHS mRNA whose lengths, 2.0 and 1.5 kb, were compatible with the mRNA length for G and N proteins respectively. This mRNA must contain about 400 bp of untranslated sequence.

L'évolution du poids du corps et du vitellus d'alevins de truite commune (Salmo trutta L.) est étudiée pendant les 91 jours qui suivent la fécondation. Les alevins sont élevés à une température moyenne de 8,8°C (écart 3,8°C-12,3°C). Ils sont, soit nourris dès l'ouverture de l'oesophage, soit gardés à jeun. Cette étude permet de montrer que la résorption du vitellus peut être décrite dès la fécondation à l'aide d'un modèle théorique qui tient compte du poids corporel des alevins gardés à jeun. Il n'existe pas de différence de résorption du vitellus entre les alevins nourris et ceux gardés à jeun. Avant la fin de la résorption du vitellus, les alevins nourris présentent un poids corporel significativement supérieur à ceux gardés à jeun. De ce fait, le nourrissage précoce serait recommandé pour diminuer la durée d'alevinage.

A cDNA clone encoding for the glycoprotein of the viral haemorrhagic scepticaemia virus, a fish rhabdovirus, has been sequenced. The cDNA was 2035 bp long and contained two open reading frames (ORF). A 1523 bp ORF corresponded to the glycoprotein and was adjacent, on its 5' side, to an incomplete 372 bp ORF. Although the protein encoded by this ORF displayed no similarity with other rhabdovirus proteins, it was supposed that the cDNA had been reverse-transcribed from a readthrough mRNA encoding successively for the M2 and the G proteins.

The approach to the study of an aquatic ecosystem involves, above all, the analysis of the partitioning of energy between the different food chains. The objective is to bring together the elements for a better understanding of the interactions in order to maximize exploitation of the resources either for fishing or for aquaculture.

Nonflagellated germ cells were isolated from rainbow trout testis to determine their ability to synthesize 17,20 beta-dihydroxy-4-pregnen-3-one (17,20 beta OHP), a progestin involved in the control of the release of sperm. Germ cells were obtained by enzymatic dissociation (collagenase; 3 mg.ml-1, 4.5 h, 12 degrees C) from testes that were immature and at the beginning of spermatogenesis. Somatic cells were eliminated by adhesion to the culture plates. Dose-related amounts of 17,20 beta OHP were measured by RIA in culture media of germ cells incubated with increasing dosages of 17-hydroxyprogesterone (17OHP; 0.05-10 micrograms.ml-1) for 20 h at 12 degrees C. Furthermore, 3H-17,20 beta OHP was identified by chromatography and co-crystallization with a reference in incubating cells provided by 3H-17OHP (2.5 and 4 h, 12 degrees C). Other metabolites were detected but not identified. 11-Ketotestosterone (11KT) was either nondetectable by RIA in control cultures or, when detected, was found at very low levels. In no case was 11KT stimulated by addition of 17OHP or gonadotropin II (GtH II; 400 ng.ml-1); this indicated the absence of contamination by Leydig cells. Thus, to our knowledge, this report demonstrates steroidogenic activities in nonflagellated germ cells of fish testis for the first time. 20 beta-Hydroxysteroid dehydrogenase (20 beta HSD) activity was identified, showing that germ cells are able to synthesize 17,20 beta OHP at an early stage in rainbow trout testis.

Rainbow trout ( Salmo gairdneri Richardson) with a mean initial weight of 135 g were fed diets supplemented with astaxanthin or canthaxanthin in concentrations of 0.0, 12.5, 25.0, 50.0, 100.0 and 200.0 mg/kg for 6 weeks. Fish pigmentation increased with increasing dietary carotenoid concentration up to 3.7 mg/kg flesh in the best pigmented groups. The fish were pigmented faster with astaxanthin than with canthaxanthin. For both pigments, the retention coefficients de-creased as the pigment dose in the diet increased. The mean retention coefficient for astaxanthin was 1.3 times higher than for canthaxanthin.

Dynamics of canthaxanthin in rainbow trout was studied through a feeding experiment of 37 weeks by following canthaxanthin deposition and retention in muscle and ovary of maturing diploid females compared to sterile triploids. In the first 4 weeks canthaxanthin retention in diploids (3.6%) was about three times that of triploids. Later on, canthaxanthin retention in triploid muscle and diploid muscle and ovary reached similar and rather constant levels (2%) while the level in the diploid muscle was lower. The canthaxanthin deposit in the flesh of fish was higher in triploids than in diploids. Furthermore, the canthaxanthin concentration in muscle plus ovary from diploids did not differ significantly from the concentration in the muscle of triploids. After spawning, canthaxanthin concentration in the muscle of female diploids reached a level as high as that of triploids after 4 weeks.

Nine induced tetraploid females were artificially inseminated by UV-irradiated sperm collected from diploid males, in order to induce the gynogenetic development of their ova. Most of the resulting embryos were diploid (or minor aneuploids). Several gynogenetic tetraploids, likely to issue from unreduced ova, were also detected in these progenies. The same females fertilized by normal sperm of diploid males gave a majority of triploids and several pentaploids, while the fertilization by normal sperm of tetraploid males gave rise to a majority of tetraploids and one hexaploid. The same crosses, after the eggs had been heat-shocked to double the maternal genetic contribution, yielded about three-quarters pentaploids and one quarter haploids (normal sperm of diploids), or three-quarters hexaploids and one quarter diploids (normal sperm of tetraploids). These haploids and diploids are likely to result from androgenesis.

A first demonstration of realized selection response for fillet yield in fish, in rainbow trout oncorhynchus mykiss. 13. International Symposium for Genetics in Aquaculture (ISGA XIII)

The correlated responses in feed intake and G:F ratio with selection for increased growth rate were evaluated by comparing selected (S) and control (C) brown trout (Salmo trutta) reared under conditions known to affect feed efficiency: feed restriction and periods of compensatory growth. Nitrogen and energy requirements for maintenance and growth were also measured. Trout were allotted at comparable BW (3.7+/-0.06 and 3.8+/-0.04 g, for C and S respectively) to triplicate groups per treatment. The experiment lasted a total of 198 d, during which animals were successively submitted to a 116-d feeding phase and fed 10, 30, 50, 70, 100, and 140% of their usual daily ration (UDR), a 35-d phase of food deprivation, and a 47-d refeeding phase. The G:F of C and S were comparable in all experimental conditions tested. During the feeding phase, S grew better than C only when fed 100 and 140% UDR (P0.001). This was explained by a higher feed intake capacity. The requirements for growth and maintenance were similar among the lines, which is in agreement with their comparable loss of weight (mean energy loss of -53 and -55 kJ/(kg x d) for C and S, respectively; P0.38) observed during the feed deprivation phase and the lack of differences in carcass composition (fat, P0.35; protein, P0.54). During the refeeding phase, growth performance and G:F were high in all groups. The daily growth coefficient was higher in S than in C (P0.001) because of a higher feed intake (P0.001). An increase in absolute individual variability in final BW and length was associated with the level of food restriction in both lines; however, it always remained lower in S than in C. In conclusion, fish selected for growth under ad libitum conditions will only exhibit growth superiority when fed diets close to ad libitum, and there was no evidence that selection was associated with an improvement in efficiency of maintenance nor in retention of body tissues.

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International audience; Many populations of vertebrates display alternative reproductive strategies involving delays in maturation processes and migrations to more productive habitats. Notably for females, migratory patterns and diet could influence reproductive traits such as ova size and number. To test this hypothesis, we targeted anadromous and freshwaterresident female brown trout of various sizes during their return to spawn in tributaries, and carried out stable isotope analyses (d15N and d13C) on ova to identify their feeding areas and diet. The study was performed in the Oir River (France), a small coastal system where trout migrations to sea and within the river basin have been monitored for the last 20 years. General trends were observed in reproductive traits, such as the positive correlation between female fork length, fecundity and ova size. Fecundity was sizedependent whatever the migratory origin of females, but major divergences were seen for ova size and isotopic ratios. Freshwater residents displayed a constant increase in ova size and a 15N-enrichment in ova, suggesting an investment in larger ova and a shift to piscivory as the fish become larger and older. Conversely, anadromous females had smaller ova compared with freshwater-resident females of similar body size and achieved higher fecundity as they grew bigger. Our results suggest the existence of a continuum of reproductive traits for freshwater-resident females whereas anadromous females clearly show a break with this continuum. This major dissimilarity could be explained by the difference in growing environments in terms of food availability and quality.