Wide-gene expression analysis of lipid-relevant genes in nutritionally challenged gilthead sea bream (Sparus aurata)

Disturbances of lipid metabolism are a major problem in livestock fish and the present study analysed the different tissue expression patterns and regulations of 40 lipid-relevant genes in gilthead sea bream. Nineteen sequences, including fatty acid elongases (4), phospholipases (7), acylglycerol lipases (8) and lipase-maturating enzymes (1), were new for gilthead sea bream (GenBank, JX975700-JX975718). Up to six different lipase-related enzymes were highly expressed in adipose tissue and liver, which also showed a high expression level of δ6 and δ9 desaturases. In the brain, the greatest gene expression level was achieved by the very long chain fatty acid elongation 1, along with relatively high levels of δ9 desaturases and the phospholipase retinoic acid receptor responder. These two enzymes were also expressed at a high level in white skeletal muscle, which also shared a high expression of lipid oxidative enzymes. An overall down-regulation trend was observed in liver and adipose tissue in response to fasting following the depletion of lipid stores. The white skeletal muscle of fasted fish showed a strong down-regulation of δ9 desaturases in conjunction with a consistent up-regulation of the >lipolytic machinery> including key enzymes of tissue fatty acid uptake and mitochondrial fatty acid transport and oxidation. In contrast, the gene expression profile of the brain remained almost unaltered in fasted fish, which highlights the different tissue plasticity of lipid-related genes. Taken together, these findings provide new fish genomic resources and contribute to define the most informative set of lipid-relevant genes for a given tissue and physiological condition in gilthead sea bream. © 2014 Elsevier B.V.This research was funded by the Spanish MICINN through AQUAFAT (AGL2009-07797; predictive modelling of flesh fatty acid composition in cultured fish species with different muscle lipid contents) and AQUAGENOMICS (CSD2007-00002, improvement of aquaculture production by the use of biotechnological tools) projects. Additional funding was obtained from EU project ARRAINA (KBBE-2011-5-288925, advanced research initiatives for nutrition and aquaculture).Peer Reviewe

Up-scaling validation of a dummy regression approach for predictive modelling the fillet fatty acid composition of cultured European sea bass (Dicentrarchus labrax)

The aim of the study was to validate a dummy regression approach for predictive modelling the fillet fatty acid (FA) composition of cultured European sea bass with dietary FA composition and lipid fillet content as independent variables. The model used our own data on gilthead sea bream as reference subgroup dataset and data from turbot, sole and European sea bass as dummy variables. Most of the observed variance within and among species was explained by the regression model without statistical significant interactions on blocks between diet composition and fish species subgroups. For the validation of European sea bass FA descriptors, predictive values derived from data on fish reared at laboratory scale were plotted against those obtained in farmed fish harvested at commercial size. A close linear association near to equality was found for 12 representative FAs, including saturated FAs, monoenenes and polyunsaturated FAs. This finding reinforces the possibility to produce tailored and healthy seafood products according to the guidelines of essential FA requirements in humans. FA algorithms for all the species in the model are hosted at www.nutrigroup-iats.org/aquafat as a multispecies tool to interrogate the nutritionally regulated FA composition of four cultured marine fish species of a high added value.This study was founded by Spanish (AQUAFAT, AGL2009-07797, Predictive modelling of flesh fatty acid composition in cultured fish species with different muscle lipid content) and EU (ARRAINA, KBBE-2011-5-288925, Advanced research initiatives for nutrition and aquaculture) projects. Additional founding was obtained from the “Generalitat Valenciana” (research grant PROMETEO 2010/006). GFB-L was recipient of a Spanish PhD fellowship from the Diputación Provincial de Castellón.Peer Reviewe

Dummy regression analysis for modelling the nutritionally tailored fillet fatty acid composition of turbot and sole using gilthead sea bream as a reference subgroup category

Farmed turbot and sole were sampled at different stages of the production cycle for analysis of fillet lipid content and fatty acid (FA) composition. The entire data set along with our own published data on gilthead sea bream were fitted to dummy regression equations with turbot and sole as dummy variables, gilthead sea bream as a reference subgroup category, and diet FA composition and fillet lipid content as independent variables. The relative contribution of each independent variable to the total variance was found to vary within and among FAs and fish species, but strong correlation coefficients (0.76 0.99) were found for almost all of the FA equations, including saturated FAs, monoenes and long-chain polyunsaturated fatty acids (PUFA) of n-3 and n-6 series. Given the differences in lipogenic activities of the fish species, major interaction effects between fillet lipid content and dummy variables were found for monoenes and saturated FAs. The proposed equations (hosted at www.nutrigroup-iats.org/aquafat) were able to fit different proportions of EPA, DPA and DHA underlying the fish species differences in FA desaturation/elongation pathways. The robustness of the model was proven with extra data from the three fish species, allowing a close linear association near to equality for the scatter plot of observed and predicted values. © 2014 John Wiley & Sons Ltd.This study was funded by Spanish (AQUAFAT, AGL2009-07797, Predictive modelling of flesh fatty acid composition in cultured fish species with different muscle lipid content; AQUAGENOMICS, CSD2007-00002, Improvement of aquaculture production by the use of biotechnological tools) and EU (ARRAINA, KBBE-2011-5-288925, Advanced research initiatives for nutrition and aquaculture) projects. Additional funding was obtained from the ‘Generalitat Valenciana’ (research grant PROMETEO 2010/006). GFB-L was recipient of a Spanish PhD fellowship from the Diputación Provincial de Castellón.Peer Reviewe

Lasting effects of butyrate and low FM/FO diets on growth performance, blood haematology/biochemistry and molecular growth-related markers in gilthead sea bream (Sparus aurata)

Four isoproteic/isolipidic plant protein-based diets were formulated to assess the lasting effects of feed additives and low fish meal (FM) and fish oil (FO) diet formulations on gilthead sea bream growth performance. FM was included at 23% in the control diet (D1) and at 3% in the other three diets (D2, D3, D4). Added oil was either FO (D1) or a blend of vegetable oils replacing 58% (D2) and 84% (D3, D4 diets) of FO. A commercial sodium butyrate preparation (NOREL, 70-BP) was added to the D4 diet at 0.4%. Each diet was allocated to triplicate groups of juvenile fish fed to satiety over an 8-month feeding trial (May-December). All fish grew efficiently from 15. g of initial body weight to 296-320. g with an overall feed efficiency (FE) of 0.95-1.01, although fish fed D3 and D4 diets showed transient growth impairments over the course of the first four weeks of the trial. Data on biometric indexes, whole body composition, haematology and blood biochemistry revealed a strong effect of sampling time in fish sampled at mid-summer (August) and late autumn (December). In contrast, the diet effect was mostly reduced to a few blood parameters. Low inclusion levels of FM reduced plasma haemoglobin levels (D2, D3), but these effects were reversed by butyrate supplementation (D4). The same phenomena occurred for total cholesterol with the highest circulating concentration of choline and IGF-I in fish fed the D4 diet during their summer growth spurt. At the transcriptional level, gene expression profiling of liver and skeletal muscle with a PCR-array of 87 growth markers provided additional evidence for an overall well-growth condition in all of the experimental groups. Up to 73 genes were found at detectable levels in the liver tissue, but only 13 were differentially expressed. Likewise, 84 genes were actively transcribed in the skeletal muscle, but only nine were differentially expressed in at least one experimental group. Butyrate supplementation reversed the up-regulated expression of inflammatory cytokines (TNFα) and muscle markers of cellular morphogenesis and protein breakdown (CDH15, CAPN3, PSMA5, PSMB1, UBE2N) in the muscle of fish fed the extreme D3 diet. These results support the use of low FM/FO diets alone or supplemented with feed additives, which have the potential to improve or reverse metabolic steady-states. Statement of relevance: Butyrate effect on low fish meal/fish oil diets.This study was funded by the European Union (ARRAINA, FP7-KBBE-2011-5-288925, Advanced research initiatives for nutrition and aquaculture) projects. Additional funding was obtained from the Spanish MINECO (MI2-Fish, AGL2013-48560) and from Generalitat Valenciana (PROMETEO FASE II-2014/085).Peer Reviewe

Metabolic and transcriptional responses of gilthead sea bream (Sparus aurata L.) to environmental stress: New insights in fish mitochondrial phenotyping

The aim of the current study was to phenotype fish metabolism and the transcriptionally-mediated response of hepatic mitochondria of gilthead sea bream to intermittent and repetitive environmental stressors: (i) changes in water temperature (T-ST), (ii) changes in water level and chasing (C-ST) and (iii) multiple sensory perception stressors (M-ST). Gene expression profiling was done using a quantitative PCR array of 60 mitochondria-related genes, selected as markers of transcriptional regulation, oxidative metabolism, respiration uncoupling, antioxidant defense, protein import/folding/assembly, and mitochondrial dynamics and apoptosis. The mitochondrial phenotype mirrored changes in fish performance, haematology and lactate production. T-ST especially up-regulated transcriptional factors (PGC1α, NRF1, NRF2), rate limiting enzymes of fatty acid β-oxidation (CPT1A) and tricarboxylic acid cycle (CS), membrane translocases (Tim/TOM complex) and molecular chaperones (mtHsp10, mtHsp60, mtHsp70) to improve the oxidative capacity in a milieu of a reduced feed intake and impaired haematology. The lack of mitochondrial response, increased production of lactate and negligible effects on growth performance in C-ST fish were mostly considered as a switch from aerobic to anaerobic metabolism. A strong down-regulation of PGC1α, NRF1, NRF2, CPT1A, CS and markers of mitochondrial dynamics and apoptosis (BAX, BCLX, MFN2, MIRO2) occurred in M-ST fish in association with the greatest circulating cortisol concentration and a reduced lactate production and feed efficiency, which represents a metabolic condition with the highest allostatic load score. These findings evidence a high mitochondrial plasticity against stress stimuli, providing new insights to define the threshold level of stress condition in fish. © 2014 Elsevier Inc.This work was funded by the EU AQUAEXCEL (Aquaculture Infrastructures for Excellence in European Fish Research, FP7/2007/2013; grant agreement No. 262336), and the Spanish AQUAGENOMICS (CSD2007-00002, Improvement of aquaculture production by the use of biotechnological tools) projects. Additional funding was obtained by Generalitat Valenciana (research grant PROMETEO 2010/006).Peer Reviewe

Aquamax: Sustainable aquafeeds to maximize the health effects of farmed fish for consumers

Jornada de Divulgación Proyecto Europeo AQUAMAX celebrada en el Instituto de Acuicultura de Torre de la Sal (Castellón) el 24 de noviembre de 2010.El Instituto de Acuicultura Torre de la Sal del CSIC organiza una jornada que tiene como objetivo divulgar la investigación sobre el proyecto europeo AQUAMAX. Este proyecto centra su investigación en reemplazar en lo posible los piensos basados en pescado y el aceite de pescado usado actualmente para alimentar a los peces por recursos alternativos libres de contaminantes. Se conseguiría así maximizar el crecimiento, mejorar la conversión del alimento e incrementar la salud y el bienestar de los peces criados en piscifactorías. Además aumentaría la seguridad y calidad del producto que llega finalmente a los consumidores. La jornada contará con la participación del Doctor S. Kaushik del Instituto Nacional para la Investigación Agronómica de Francia, el Doctor Marc Berntssen del Instituto Nacional de Nutrición e Investigación del Alimento Marino de Noruega y el Profesor Jaume Pérez del Instituto de Acuicultura Torre de la Sal del CSIC. En las diferentes intervenciones se tratarán temas del proyecto AQUAMAX como la calidad de los alimentos, la seguridad en la alimentación, así como diversas investigaciones basadas en las doradas.Peer Reviewe

Modelling the predictable effects of dietary lipid sources on the fillet fatty acid composition of one-year-old gilthead sea bream (Sparus aurata L.)

The present study aimed to ascertain the different fatty acid (FA) descriptors linking dietary and muscle FA composition in one-year-old gilthead sea bream. For that purpose, our own published data along with additional data from the present study were compiled and analysed. High linear correlations (r2 = 0.90, P < 0.001) between dietary and muscle fatty acid composition were reported for monoenes, C18 polyunsaturated FA (PUFA) and long-chain PUFA. Prediction deviations due to changes in muscle fatness were analyzed in an independent trial with two different feeding levels (full ration size, 30% restriction ration). Regardless of feeding regimen, predicted values for muscle FA at low concentrations deviated (P < 0.001) from observed values, but good predictions with less than 6% deviations were found for abundant fatty acids (16:1n-7, 18:1n-9, 18:2n-6, 18:3n-3, 20:4n-6, 20:5n-3, 22:6n-3). All this highlights the predictable effects of dietary oils in the muscle FA composition of gilthead sea bream, although further research is needed to cover all the range of commercial fish size and for the up-scaling of laboratory results to different fish farming conditions

Effect of ration size on fillet fatty acid composition, phospholipid allostasis and mRNA expression patterns of lipid regulatory genes in gilthead sea bream (Sparus aurata)

The effect of ration size on muscle fatty acid (FA) composition and mRNA expression levels of key regulatory enzymes of lipid and lipoprotein metabolism have been addressed in juveniles of gilthead sea bream fed a practical diet over the course of an 11-week trial. The experimental setup included three feeding levels: (i) full ration until visual satiety, (ii) 70 % of satiation and (iii) 70 % of satiation with the last 2 weeks at the maintenance ration. Feed restriction reduced lipid content of whole body by 30 % and that of fillet by 50 %. In this scenario, the FA composition of fillet TAG was not altered by ration size, whereas that of phospholipids was largely modified with a higher retention of arachidonic acid and DHA. The mRNA transcript levels of lysophosphatidylcholine acyltransferases, phosphatidylethanolamine N-methyltransferase and FA desaturase 2 were not regulated by ration size in the present experimental model. In contrast, mRNA levels of stearoyl-CoA desaturases were markedly down-regulated by feed restriction. An opposite trend was found for a muscle-specific lipoprotein lipase, which is exclusive of fish lineage. Several upstream regulatory transcriptions were also assessed, although nutritionally mediated changes in mRNA transcripts were almost reduced to PPARα and β, which might act in a counter-regulatory way on lipolysis and lipogenic pathways. This gene expression pattern contributes to the construction of a panel of biomarkers to direct marine fish production towards muscle lean phenotypes with increased retentions of long-chain PUFA. Copyright © The Authors 2012.The present study was funded by the Spanish MICINN (AQUAFAT, AGL2009-07797, Predictive modelling of flesh FA composition in cultured fish species with different muscle lipid content; AQUAGENOMICS, CSD2007-00002, Improvement of aquaculture production by the use of biotechnological tools) and the European Union (ARRAINA, FP7-KBBE-2011-5-288925, Advanced research initiatives for nutrition and aquaculture) projects. Additional funding was obtained from the 'GeneralitatValenciana' (research grant PROMETEO 2010/006).Peer Reviewe

Prediction of fillet fatty acid composition of market-size gilthead sea bream (Sparus aurata) using a regression modelling approach

26 p., 5 figures, 1 table and referencesGilthead sea bream (Sparus aurata) were fed in triplicate groups with a commercial standard diet from the juvenile stage to male-female sex reversal under natural day length and temperature conditions. Every 3-4 months during the two-year production cycle, 9 fish were randomly selected and sampled for flesh composition analyses of total lipid levels and fatty acid (FA) composition. Curvilinear regressions fitting total lipid levels and % FAs in total lipids were made to underline the differential distribution of a given fillet FA within neutral and polar lipid fractions. This dataset along with published results on market-size fish were combined for multilinear regression approaches, with the aim of describing strong relationships (P < 0.0001) between fillet FA composition and two independent variables: dietary FA composition and fillet lipid level. For saturated (14:0, 16:0, 18:0) and monounsaturated (16:1n-7, 18:1n-7, 18:1n-9, 20:1n-9) FAs, the overall variance in fillet FA composition is primarily explained by dietary FA composition and secondly by fillet lipid level. This second independent variable also contributes to explain the variations observed in arachidonic acid (20:4n-6) and docosahexaenoic acid (22:6n-3), but a statistically significant contribution is not found for linoleic acid (18:2n-6), linolenic acid (18:3n-3), eicosapentaenoic acid (20:5n-3) and docosapentaenoic acid (22:5n-3). The consistency of these predictive equations in our particular rearing conditions was proved by means of a test validation trial, using fish fed an experimental diet based on plant proteins and fish oil.This research was funded by Spanish MICINN through AQUAFAT (AGL2009-07797; Predictive modelling of flesh fatty acid composition in cultured fish species with different muscle lipid content) and AQUAGENOMICS (CSD2007-00002, Improvement of aquaculture production by the use of biotechnological tools) projects. Additional funding was obtained from the “Generalitat Valenciana” (research grant PROMETEO 2010/006). GFB-L was recipient of a Spanish PhD fellowship from the Diputación Provincial de Castellón

Wide-gene expression analysis of lipid-relevant genes in nutritionally challenged gilthead sea bream (Sparus aurata)

Trabajo presentado en Aquaculture Europe 2014, celebrado en San Sebastián (España) del 4 al 17 de octubre de 2014The use of new diets based on alternative sources to marine raw materials has been successful in several finfish species of interest to European aquaculture, including gilthead sea bream (Benedito-Palos et al., 2009). However, concerted efforts are still needed to undertake in depth the long-term consequences of feeding very low fish meal and fish oil diets (less than 10% of inclusion), and new knowledge is welcome to fill the gaps in a wide range of protein- and lipid metabolic disorders, closely related with reduced growth and survival. Therefore, there is a strong need for integrative tools that are capable of describing and predicting any metabolic disturbance of lipid metabolism in fish. In this scenario, the aim of the present study was to phenotype at the transcriptional level the tissue-specific regulation of lipid metabolism in nutritionally challenged fish, focusing on a panel of 40 genes which were selected as markers of fatty acid, phospholipid, acylglycerol and lipoprotein metabolism. The final aim is to define a data set of highly informative molecular markers of nutritional condition for a given tissue.Peer reviewe