Fish performance and quality of gilthead sea. bream fed extremely low fish meal and fish oil. diets with or without butyrate supplementation
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1 Fish performance and quality of gilthead sea Advanced Research Initiatives for bream fed extremely low fish meal and fish oil Nutrition & Aquaculture diets with or without butyrate supplementation L. Benedito Palos, G.F. Ballester Lozano, V. Karalazos, A. Ortiz, S. Kaushik, J. Pérez Sánchez* *Nutrigenomics & Fish Growth Endocrinology Instituto de Acuicultura Torre de la Sal (IATS CSIC), Castellón, Spain AE 2014, San Sebastián
2 Fish meal and fish oil targets in practical sea bream diets AQUAMAX, PEPPA, PERSPECTIVES OF PLANT PROTEIN USE IN AQUACULTURE 15-20% FM 12-15% FO Gómez-Requeni et al., 2003, 2004 SUSTAINABLE AQUAFEEDS TO MAXIMISE THE HEALTH BENEFITS OF FARMED FISH FOR CONSUMERS 15-20% FM 5% FO Benedito-Palos et al., 2007, 2008, 2009 ARRAINA, ADVANCED RESEARCH INITIATIVES FOR NUTRITION & AQUACULTURE 5% FM 2.5% FO
3 OBJECTIVES 1- Growth performance 2- Basic blood biochemistry 3- FA profiling of fillets & RBC
4 EXPERIMENTAL SETUP Four experimental diets with changing FM/FO content formulated by BIOMAR and supplemented by BP70 Initial BW 15 g Replicate tanks (3000 L), 150 fish each Natural photoperiod and temperature conditions Fish are fed to visual satiety 1 2 times per day and 3 6 days per week through the production cycle Each 4 6 weeks biomass is determined Fish are sampled periodically for blood and tissue samples (biochemical, histological, molecular, proteomic, microbiological, and food safety analyses )
5 Diet composition Ingredient (%) Diet D1 D2 D3 D4 Fish meal CPSP Soya protein Corn gluten D1 (CTRL) 25% FM D2 5% FM D3 5% FM D4 5% FM Wheat gluten Rapeseed cake Wheat Fish oil % FO 6.5% FO 2.5% FO 2.5% FO Rapeseed oil Palm olein Emulthin G Choline chloride % VO 13% VO 13% VO Monocalcium phosphate L-Lysine L-Histidin Methionine Mineral-vitamin mix % BP70 Cholesterol Ethoxiquin BAROX BECP Yttrium EPA + DHA 2.9% DM EPA + DHA 1.4% DM EPA + DHA % DM EPA + DHA % DM BP Proximate composition Dry matter (DM, %) Crude protein (% DM) Crude fat (% DM) EPA+DHA (% DM)
6 Growth performance I Temperature (ºC) wks Day length (h) All groups grew fast, final FGR ~ 1 Fish fed CTRL (D1) and D2 (5FM/6.5 FO) diets are undistinguishable 350 Body weight (g) wks 13 wks D1(FM25/FO15) D2(FM5/FO6) D3(FM5/FO2.5) D4(FM5/FO2.5/BUT) 0 May Jun Jul Aug Sep Oct Nov Dec Jan 6-7% reduction in BW mass is found in fish fed D3 and D4 diets (5FM/ 2.5FO) The growth impairment was restricted to starting period
7 Changes in SGR from May to December More than 300 g at the end * * Short adaptation periods (2-4 wks) with intermediated diets are required before the use of extremely low FM/FO diets
8 Growth performance II Sampling schedule D1 (FM25/FO15) D2 (FM5/FO6) D3 (FM5/FO2.5) D4 (FM5/FO2.5/BUT) WB composition/ Nutrient retention Blood biochemistry Body weight (g) Just 3 wks ago FGR ~ May Aug Nov Feb May Aug Nov Blood & Tissue Sampling Tissue FA profiling Histopathological scoring of liver & intestine Transcriptome (liver, SKM, intestine) Intestinal mucus proteome Intestinal microbiota Screening of contaminants (food safety)
9 % wet weight 100g BW 300g BW Gut Index N retention, 36-40% Lipid retention > 46-50% GI = (100 x BW/ intestine length 3 ) a ab b a 5 4 D1 D2 D3 D4
10 D1 D2 D3 D4 Itziar Estensoro Fish Pathology IATS-CSIC
11 Blood Biochemistry Haematology Electrolytes & Metabolites Enzymes Hormones Other Hc Calcium ALAT GH TAC Hb Chloride ASAT IGF-I RB RBC Magnesium GLDH Cortisol Phosphate ALP Glucose Lysozyme TG Cholesterol Proteins Creatinine Choline
12 Basic Blood Biochemistry P-values, ANOVA-II (diet_season effects) DIET SEASON INTERACTION Haemoglobin (g/dl) Haematocrit (%) RBC x 10 6/ml Glucose (mg/dl) Triglycerides (mm) Total cholesterol (mg/dl) HDL cholesterol (mg/dl) VLDL/LDL cholesterol (mg/dl) Total proteins (g/l) ALAT (U/l) ASAT (U/l) GLDH (U/l) ALP (U/l) Creatinine (mg/dl) Choline (µm) Calcium (mg/dl) Chloride (mg/dl) Magnesium (mg/ml) Phosphate (mg/dl) Antioxidant capacity (Trolox mm) Lysozyme (U/l) Respiratory burst (IRLU) GH (ng/ml) IGF I (ng/ml) BW g Aug 2013 wk 13 BW g Dec 2013 wk 31 A sampling-time effect is found for most of the analysed parameters (wks 13-31) Diet & time-interaction effects are restricted to Hb, cholesterol, choline and IGF-I
13 Haemoglobin, g/dl a wk 13, Hb (g/dl) a b b D1 D2 D3 D4 wk 31, Hb (g/dl) a b ab b D2, D3 diets Low FM inclusion levels (5%) reduce haemoglobin concentration D1 D2 D3 D4 D4 diet, 0.4% GUSTOR BP70 Anaemia signs are reversed by BUT supplementation
14 Cholesterol, mg/dl wk 13, Cholesterol 160 a ab ab b D1 D2 D3 D4 D4 diet 0.4% GUSTOR BP70 wk 31, Cholesterol 300 a a 240 b b D1 D2 D3 D4 Clinical signs of hypocholesterolemia are reversed by BUT supplementation
15 Choline & IGF-I wk 13, Choline (µm) a a a b D1 D2 D3 D4 D4 diet 0.4% GUSTOR BP70 wk 13, IGF-I (ng/ml) 125 b 105 a a a D1 D2 D3 D4 Other potential benefits include transient increases in circulating vitamin and growth-promoting factors
16 Fillet FA profile Diet Fatty acid D1 D2 D3 D4 14:0 4.01±0.23 a 4.69±0.43 ab 5.54±0.27 b 5.34±0.36 b 16: ±0.20 a 18.15±0.54 a 16.99±0.30 b 16.91±0.25 b 16:1n ±0.33 a 4.07±0.23 b 2.92±0.43 c 3.35±0.21 bc 18:0 5.01± ± ± ± :1n-9 (Oleic) 17.7±0.77 a 25.61±0.88 b 32.96±1.15 c 31.42±1.13 c 18:1n ±0.46 a 0.53±0.21 b 0.68±0.12 b 0.91±0.10 b 18:2n-6 (LA) 5.85±0.13 a 14.63±0.25 b 16.64±0.17 c 16.32±0.30 c 18:3n ±0.01 a 0.13±0.01 b 0.06±0.01 c 0.07±0.01 c 18:3n-3 (LNA) 1.17±0.05 a 2.32±0.12 b 3.61±0.13 c 3.49±0.13 c 18:4n ±0.06 a 0.46±0.04 b 0.29±0.03 c 0.32±0.01 c 20:1n ±0.05 a 0.85±0.05 b 1.08±0.04 c 1.04±0.05 c 20:4n-6 (ARA) 1.07±0.07 a 0.70±0.08 b 0.32±0.04 c 0.39±0.05 c 20:5n-3 (EPA) 10.24±0.21 a 6.42±0.51 b 3.28±0.30 c 3.74±0.41 c 22:4n ±0.01 a 0.08±0.01 b 0.04±0.01 c 0.05±0.01 c 22:5n ±0.02 a 0.17±0.02 a 0.08±0.01 b 0.11±0.02 b 22:6n-3 (DHA) 12.72±0.77 a 7.67±0.79 b 3.92±0.49 c 4.98±0.70 c C18-PUFA C18-PUFA C18-PUFA n6 LC-PUFA n3 LC-PUFA
17 Fillet FA composition Total lipids, % FAME :1n9 18:2n6 18:3n D1 D2 D3 D4 FO oil VO oils D1 D2 D3 D4 20:4n6 20:5n3 22:6n3
18 Dummy regression model: a close association between observed & predicted FA-values P104 Gabriel Ballester-Lozano Nutrigenomics & Fish Growth Endocrinology, IATS-CSIC
19 Sea bream Sea bass turbot Sole
20 Neutral lipids_tg Reflect feed composition Polar lipids_pls Highly regulated PL allostasis Feeding level Environment EFA deficiencies Benedito-Palos et al., 2008, British Journal of Nutrition Benedito-Palos et al., 2010, Journal of Animal Science Benedito-Palos et al., 2013, British Journal of Nutrition
21 Unsaturation Index (UI) of RBC UI = (% unsaturated FA i x number double bounds) i= n TG Chol - Non-invasive method for routine diagnosis of EFA deficiencies in farmed sea bream PLs UI, Cold season (D1-D4) UI, Warm season (D1-D4)
22 Haemolysis (%) Ctrl Clinical signs of EFA deficiencies: growth retardation, dyslipemia, n-3 LC-PUFA CTRL PL/Met/Min def UI from 260 to 160 Vit/Pi def EFA def UI of RBC < 160 Semipurified diets NaCl (g/l) Increases osmotic fragility of RBC Mean corpuscular index, MCF > 7.5
23 Concluding Remarks & Perspectives
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28 Session 14 O461. Sea bass Intestine transcriptome Josep A Calduch-Giner Session 14 O595. Histophathological and transcriptomic scoring of sea bream intestine Itziar Estensoro Session 14 O161.Wide-screening of contaminants Jaime Nácher-Mestre P105. Molecular signatures of lipid metabolism Laura Benedito-Palos P104. Dummy regression of FAs G.F Ballester-Lozano P109. Microbiota diversity Belén Fouz P115 Mycotoxin carry-over Roque Serrano
29 Thanks you for your attention Sadasivam Kaushik Jaume Pérez-Sánchez Laura Benedito-Palos Gabriel F. Ballester-Lozano Alvaro Ortiz Vasilis Karalazos Acknowledgements: Patricia Cabrera Mª Angeles González Alessandro de Ambrosi Azucena Bermejo Imma Vicent Paco Valle JC Navarro
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