Comprehensive biometric, biochemical and histopathological assessment of nutrient deficiencies in gilthead sea bream fed semi-purified diets.

Seven isoproteic and isolipidic semi-purified diets were formulated to assess specific nutrient deficiencies in sulphur amino acids (SAA), n-3 long-chain PUFA (n-3 LC-PUFA), phospholipids (PL), P, minerals (Min) and vitamins (Vit). The control diet (CTRL) contained these essential nutrients in adequate amounts. Each diet was allocated to triplicate groups of juvenile gilthead sea bream fed to satiety over an 11-week feeding trial period. Weight gain of n-3 LC-PUFA, P-Vit and PL-Min-SAA groups was 50, 60-75 and 80-85 % of the CTRL group, respectively. Fat retention was decreased by all nutrient deficiencies except by the Min diet. Strong effects on N retention were found in n-3 LC-PUFA and P fish. Combined anaemia and increased blood respiratory burst were observed in n-3 LC-PUFA fish. Hypoproteinaemia was found in SAA, n-3 LC-PUFA, PL and Vit fish. Derangements of lipid metabolism were also a common disorder, but the lipodystrophic phenotype of P fish was different from that of other groups. Changes in plasma levels of electrolytes (Ca, phosphate), metabolites (creatinine, choline) and enzyme activities (alkaline phosphatase) were related to specific nutrient deficiencies in PL, P, Min or Vit fish, whereas changes in circulating levels of growth hormone and insulin-like growth factor I primarily reflected the intensity of the nutritional stressor. Histopathological scoring of the liver and intestine segments showed specific nutrient-mediated changes in lipid cell vacuolisation, inflammation of intestinal submucosa, as well as the distribution and number of intestinal goblet and rodlet cells. These results contribute to define the normal range of variation for selected biometric, biochemical, haematological and histochemical markers.

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 ß-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.

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 ß, 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.