Sustainable Fish Diets for the 21st Century using Soybean Protein. Paul B. Brown, Purdue University, West Lafayette, Indiana, USA

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Sustainable Fish Diets for the 21st Century using Soybean Protein Paul B. Brown, Purdue University, West Lafayette, Indiana, USA pb@purdue.edu

*Introduction *If you want to grow an animal, you must provide adequate food resources. *Inadequate food intake negatively affects growth, health and reproduction, and increases cost of production *Feed costs are significant *Are all nutritional requirements for fish the same? Is it reasonable and appropriate to develop species specific diets? *Increased aquacultural production placing significant demand on fish meal and oil supplies

*Ingredients *Fish meal and oil *1960s Ingredient for swine and poultry feeds *1980s Poultry feed (50%), aquaculture 10% *2010 Aquaculture demanding 73%, poultry 5%, swine 20% *J. Shepard, past Director General, International Fishmeal and Fishoil Organisation (IFFO)

*Ingredients *Sources of fish meal and oil sardines, menhaden, anchovy, mackerel, capelin, herring *IFFO - http://www.iffo.net/default.asp?contentid =718

*Ingredients *Tacon and Metian, 2008, Aquaculture 285:146-158

*Summary *Primary ingredient is at maximum sustainable yield supply is unlikely to increase *Demand for that ingredient is increasing *Price is increasing

*Ingredient substitution *Candidate replacement plant feedstuffs *Soybean meal, canola (rapeseed) meal, lupin meal, pea meal, wheat gluten, cottonseed meal, sunflower seed meal, safflower seed meal, sesame seed meal, linseed meal, corn gluten meal, whey, groundnut meal, brewer s grains and yeast, distiller s dried grains *Protein concentrates and isolates (soybean, pea, lupin, potato, whey, alfalfa, rice, etc.)

*Ingredient substitution * Candidate replacement plant feedstuffs * Global production (2011, mmt, UNFAO) * Soybeans 260.9 * Oil palm 233.8 * Seed, cotton 77.3 * Rapeseed 62.4 * Coconut 59.2 * Cotton 48.8 * Sunflower 40.2 * Groundnuts 38.6 * Palm kernel 12.9 * Peas, dry 9.6 * Sesame seed 4.1 * Castor oil seed 2.7 * Linseed 1.6 * Lupins 1.1 * Safflower 0.6

*Soybean meal prices 1984-2014 10

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*Soybean Protein *Protein feedstuffs are key *Optimal dietary crude protein for fishes is >28% of the diet, with many values in the 40-50% range *Protein feedstuffs with potential have greater than 30% crude protein *Isonitrogenous replacement of fish meal with test feedstuff, typically with little understanding of nutritional requirements for target species 12

*Mineral availability *Phosphorus in plant feedstuffs can bind other minerals and lead to mineral deficiencies *Initial evaluation of SBM in diets for hybrid striped bass indicated maximum level of incorporation of 20% of the diet. *Subsequent study using improved mineral premix indicated HSB were tolerant of up to 40% SBM in the diet. * Brown et al. 1997 13

*Essential amino acids *Factors that may contribute to limited use *Nutritional deficiencies most recent NRC Nutrient Requirements of Fish and Shrimp (2011) lists optimal dietary crude protein concentrations for 21 species (18 fish and 3 shrimp), estimates for probably 100 other species *Quantified essential amino acid concentrations (EAA) * Lysine 21 species, 1.2-3.3% of diet * Methionine 16 species, 0.4-1.5% of diet * Threonine 12 species, 0.5-1.8% of diet *All ten EAA requirements quantified for 6 species

*Essential amino acid supply *Essential amino acid requirements hybrid striped bass * Arginine 1.0-1.5% * Lysine 1.4% * Methionine 0.7% * Methionine + cyst(e)ine 1.1% * Phenylalanine 0.9% * Threonine 0.9% * Tryptophan 0.3% * Histidine 0.5% * Leucine 1.5% * Isoleucine 0.9% * Valine 0.8% * Twibell et al. 2003. Aquaculture Nutrition 9:373-381.

*Micronutrients *Essential amino acid requirements yellow perch * Arginine 1.4% * Methionine 1.1% * Methionine + cyst(e)ine 0.9% * Lysine 3.0% * Phenylalanine 1.6% * Threonine 1.6% * Tryptophan 0.3% * Histidine 0.5% * Leucine 1.5% * Isoleucine 0.9% * Valine 0.8% * Hart et al. 2010. Aquaculture Nutrition 16:248-253

*Ingredient substitution *Solvent-extracted, dehulled soybean meal evaluations at Purdue University *Hybrid striped bass 40% of diet *Yellow perch 30% of diet *Largemouth bass 50% of diet *Channel catfish 50% of diet *Trout/salmon - <17% of diet

*Ingredient substitution *Tolerance of SBM in the 30-50% of diet for piscivorous species range is high. *Are species evaluated more tolerant? Or, did formulation approach impact results?

*Factors that may contribute to limited use *Methods replacement of fish meal with single test feedstuff *Reality use blends of feedstuffs and fish mealfree, all plant diets have been developed for a variety of species *Nutritional deficiencies?

*Ingredient substitution *Subsequent studies using maximum concentrations of soybean meal were designed to develop fish meal-free diets for hybrid striped bass and yellow perch. *Combinations of animal by-product meals (poultry) with soybean meal supported similar weight gain to fish meal control diets in both species *Fish meal-free diets are possible!

*Potential new approaches *Applied research Blends of protein feedstuffs to meet essential amino acid and mineral requirements *Basic research New enabling technologies *Metabolomics identification of all metabolites in samples *Proteomics identification of proteins *Applied to whole organisms, targeted nutrients, protein ingredients and ingredients

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*Potential new approaches *Metabolomics marine shrimp, cholesterol study Comparison Total peaks detected Significantly changed Peaks identified Compound present peaks 1 in KEGG 2 SL vs. SCL Hepatopancreas 2309 135(5.85) 50 5 Muscle 1006 55(5.47) 17 3 Hemolymph 1120 95(8.48) 40 6

*Potential new approaches *Metabolomics shrimp/cholesterol * Hepatopancreas Muscle Serum * Metabolite Fold change Metabolite Fold change Metabolite Fold change * L-Serine 10.00 Glycerol 10.00 L-Tyrosine 0.12 * L-Valine 10.00 L-Alanine 3.61 5-Oxoproline 0.11 * Methylmalonic acid 0.14 L-Proline 0.10 L-Methionine 0.10 * Malonic acid 0.11 Glycine 0.10 Cholesterol 0.10 * L-Leucine 0.11 Malonic acid 0.09

Effect of lack of dietary cholesterol supplementation on the expression of muscle protein in Pacific white shrimp SSP number Protein name Fold change (SL/SCL) 2508 Pyruvate kinase 3.85 8205 Arginine kinase 0.21 3811 Hemocyanin 0.18 8207 Arginine kinase 0.17 8204 Arginine kinase 0.17 0404 SCP beta chain 0.04 8206 Arginine kinase 0.03 5605 Arginine kinase 0.01 7609 GAPDH 0.003 6608 Arginine kinase 0.002 7613 GAPDH 0.002 8402 GAPDH 0.001 Lack of cholesterol supplementation affected energy metabolism in the muscle

*Potential new approaches *Metabolomics Largemouth bass - graded levels of dietary soybean meal substituted for an isonitrogenous amount of fish meal (10, 20, 30, 40, 50 or 60% of diet) 20X

*Potential new approaches *Metabolomics Largemouth bass soybean meal, 10 and 60%, GIT Alanine, aspartate, and glutamate metabolism Valine, leucine, and isoleucine 10 Arginine and proline metabolism biosynthesis Tyrosine metabolism 9 Butanoate metabolism 8 Tryptophan metabolism Thiamin metabolism Riboflavin metabolism Pyruvate metabolism 7 6 5 4 3 2 1 0 Citrate Cycle Cysteine and methionine metabolism Galactose metabolism Glycine, serine, and threonine metabolism 10% 60% Pyrimidine metabolism Glyoxylate and dicarboxylate metabolism Propanoate metabolism Histidine metabolism Primary bile acid biosynthesis Lysine metabolism Nicotinate and nicotinamide Phenylalanine metabolism metabolism Pantothenic acid metabolism

*Potential new approaches *Metabolomics Largemouth bass soybean meal, 10 and 60%, liver Alanine, aspartate, and glutamate metabolism Valine, leucine, and isoleucine 7 Arginine and proline metabolism biosynthesis 6 Tyrosine metabolism Citrate Cycle 5 Tryptophan metabolism Synthesis and degradation of ketone bodies 4 3 2 1 0 Cysteine and methionine metabolism Galactose metabolism 10% 60% Steroid hormone biosynthesis Glycine, serine, and threonine metabolism Retinol metabolism Pyrimidine metabolism Propanoate metabolism Lysine metabolism Nicotinate and nicotinamide metabolism Glyoxylate and dicarboxylate metabolism

*Potential new approaches summary *-Omics offer a broader view of metabolic events *May identify dietary alterations needed as we adopt use of new ingredients

*Conclusions *Relatively few nutritional requirements quantified for culture species, but production has been established *Volume of production is insufficient to justify manufacture of species specific diet, but data are available to formulate *Trout diets in routine use, yet trout diets may not be optimum for growth, health and reproduction in emerging species *New enabling technologies may facilitate understanding nutritional needs and interactions

*Sustainable? Economic Environmental Sociological 31

* Thank you for your attention