I N C O R P O R AT I N G F i S H Far M ing t E c H no L og Y

January | February 2014 Current challenges and opportunities in amino acid nutrition of salmonids

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The International magazine for the aquaculture feed industry

FEATURE

Current challenges and opportunities in amino acid nutrition of salmonids

by Cludia Figueiredo-Silva and Andreas Lemme, Evonik Industries, Germany

ishmeal is still one of the main protein sources used in commercial feeds for trout and salmon. But its availability is shrinking and its cost is increasing year by year. The sustainability of the aquaculture industry depends largely on its capability to replace fishmeal with alternative sources of protein, and to reduce the currently excessive protein levels commonly applied in the formulation of commercial diets. At the same time, feeds must be formulated to be effective in covering the nutrient requirements of specific species in order to maximise growth. While this great challenge has long been foreseen and recognised by both industry and academia, a large number of feed manufacturers are still struggling to adapt, and are relying on their own ability to turn challenges into opportunities on a daily basis. Despite a growing body of evidence showing that fishmeal can be replaced by alternative protein sources (reviewed by Kaushin and Hemre, 2008; Tacon and Metian, 2008), and that opportunities may exist to reduce crude protein level in aquafeeds (Yamamoto et al., 2005; Gaylord and Barrows, 2009), scientific research is not always made widely available. The feed industry is delayed from applying the most important scientific achievements in its field. Here, we will try to summarise the major achievements in the development of more sustainable diets for salmonids, whilst not forgetting to point out the existence of limitations for their fully practical application. We have reviewed some of the most relevant studies focusing on the replacement of fishmeal with alternative protein sources, and on the possibility of reducing crude protein

in diets for salmonids. In addition, we found it important to highlight that the optimisation of amino acid nutrition goes beyond meeting its requirements for protein synthesis, and may constitute a promising approach to improve, among other things, animals immunity response to environmental stresses.

Alternatives to fishmeal: feasible?

Because protein is typically the most costly nutrient in a formulated feed, it is important to accurately cover protein, or to be more precise, to accurately cover amino acid requirements of animals (NRC, 2011). This has become a priority given the current constraints on fishmeal in cost and availability and the consequent need to replace it with plant protein sources limited in one or more of the essential amino acids. The supplementation of low fishmeal or plant protein based diets with these essential amino acids was proven to improve voluntary feed intake and whole body protein accretion in several fish species, including rainbow trout (Gomes et al., 1995; Cheng et al., 2004; Gaylord and Barrows, 2009; Kaushik et al., 1995; FigueiredoSilva et al., 2012) and Atlantic salmon (Espe et al., 2006, 2007, 2008; Torstensen et al., 2008; Kousoulaki et al., 2009). Rainbow trout seem, however, less sensitive than Atlantic salmon to the replacement of fishmeal with plant protein sources. In a comparative study, Refstie et al. (2000) showed that rainbow trout but not Atlantic salmon grew equally well with soybean meal based diets, suggesting that rainbow trout are less sensitive than salmon to anti-nutritional factors (see Table 1). But that there is much latitude for the reduction of marine resources in salmonid diets is clearly shown by the pos-

sibility of including fishmeal levels as low as 0 percent in rainbow trout feeds (Kaushik et al., 1995) and 5 percent for Atlantic salmon (Espe et al., 2007, 2008), provided that their amino acid profile is balanced with supplemental amino acids. These achievements would not have been possible if free amino acids were not utilised as efficiently as the protein-bound kind in meeting the essential amino acid requirements of fish. The 100 percent bioavailability of free amino acids has in fact been demonstrated in several fish species (reviewed by NRC, 2011), including rainbow trout (Rodehutscord et al., 1995a,b, 1997; Rollin et al., 2003) and Atlantic salmon (Espe and Lied 1994; Epse et al., 2006, 2007, 2008). However, one might and indeed should ask why some studies have failed to successfully replace fishmeal with alternative protein sources even when diets were supplemented with limiting essential amino acids. Although the answer is not yet entirely clear, it seems to lie in the obvious differences between the nutritional value of fishmeal and of alternative protein sources. When replacing fishmeal protein, in particular with plant sources, we must keep in mind that we are not only affecting amino acid availability and utilisation, but also that of fatty acids, vitamins and minerals. In addition, being rich in anti-nutritional factors and carbohydrates, plant protein inclusion may significantly impact on diet palatability and thereby voluntary feed intake, and on the availability and utilisation of energy. This might partly explain why, for example, the 2006 study of Espe et al. undertaken in collaboration with Evonik Industries was not completely successful in replacing fishmeal with plant protein sources. In the study with

16 | INTERNATIONAL AQUAFEED | January-February 2014

FEATURE Atlantic salmon, voluntary feed intake and growth decreased, even though the dietary amino acid profile had been balanced with supplemental amino acids (see Table 1). In a following study by Espe et al., again in collaboration with Evonik Industries, fishmeal was successfully replaced in Atlantic salmon diets by a mixture of plant proteins, provided that their amino acid profile was duly balanced with a mixture of supplementary amino acids, and also retaining a low inclusion level of fishmeal (5 percent). While without amino acid supplements, this low level of fishmeal would not have allowed the requirements of salmon to be satisfied, the inclusion of 5 percent fishmeal in addition to 3 percent squid hydrolysates (already applied in the first Espe study) proved an effective strategy in securing a similar feed intake level and growth rates between the fishmeal-based diet (49 percent) and fishmeal-replaced diets (5 percent). It seems clear, therefore, that although supplementation with amino acids is a crucial and effective strategy in keeping a similar protein accretion between fishmeal-based diets and plant-based diets, replacement of 100 percent fishmeal with alternative protein sources in salmon (as well as other species) still depends on formulations that cover requirements for all essential nutrients, including fatty acids, vitamins and minerals. A complete and accurate evaluation of the differences between the nutritional value of fishmeal and alternative protein sources will shortly allow the total replacement of fishmeal in salmonid feeds.

Lowering the crude protein level

Crude protein level is calculated by multiplying the amount of nitrogen by the empirically derived conversion factor of 6.25, which is based on the estimation that protein contains 16 percent nitrogen, although in reality it varies from 12 to 19 percent. Although crude protein value offers a good estimate of protein level, it does not allow the scientist to distinguish between nitrogen originating from amino acids and nitrogen originating from non-protein sources. Crude protein is therefore useless in evaluating the amino acid profile of ingredients and diets. Lowering the dietary crude protein level and supplementing diets with certain essential

amino acids is a well-established method of formulating diets for farm animals to achieve an ideal amino acid pattern, and it has been demonstrated by Verstegen and Jongbloed (2003) to reduce nitrogen excretion for pigs and poultry. These findings have even been incorporated into subsequent legislation. Although aquaculture seems more ecoefficient than pig or poultry production in providing nutrients for human consumption

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January-February 2014 | INTERNATIONAL AQUAFEED | 17

FEATURE
Table 1. The effect of on voluntary feed intake, growth, feed conversation ratio (FCR) and protein retention efficiency (PER) in Atlantic salmon and rainbow trout fed high plant protein diets compared to a FM control group. Protein sources (PS) Body weight FM % Other marinePS % Plant-PS % Supplemental AA Feed intake Results Growth FCR PER References

Atlantic salmon 200g 32.0 0 5.0 to 10.0 29.6 41.0 to 43.4 no AA Mixture, including L-Lys, L-Arg, DL-Met, L-Trp, L-Thr, L-His AA Mixture, including L-Lys, L-Arg, DL-Met, L-Trp, L-Thr, L-His L-Lys, DL-Met, L-His Similar Reduced or similar Reduced Reduced Refstie et al., 2000

300g

0.0

Reduced

Similar

Similar

Espe et al., 2006

327g

5.0

5.0 to 10

36.5 to 39.6

Similar

Similar

Similar

Similar

Espe et al., 2007

300g

30.0 to 12.0

2.5 to 5.0

29.6 to 43.0

Similar

Similar

Similar

Similar

Torstensen et al., 2008

137g Rainbow trout 55g 55g 83g 100g

5.0

5.0

48.4 to 48.9

L-Lys, DL-Met, L-Thr

Similar

Similar

Similar

Similar

Kousoulaki et al., 2009

20.0 0.0 0.0 32.0

0.0 0.0 0.0 0

30-80 80.0 62.0 29.6

L-Lys, L-Met L-Lys, L-Met L-Met no

Similar Reduced Similar Similar

Similar Reduced Similar Similar

Similar Similar Similar Reduced

Similar Reduced Similar -

Gomes et al.,1995 Gomes et al.,1995 Kaushik et al., 1995 Refstie et al., 2000

(Ytrestyl et al., 2012), the move towards a lower nitrogen output is a major sustainability drive in fish farming, both for environmental and economic reasons. Furthermore, the contribution of amino acids towards meeting the energy requirements of fish are considered high (NRC, 2011), and thus efforts must be taken to reduce surplus protein supply whilst improving amino acid and non-protein energy utilisation. A decrease in the digestible-protein-todigestible-energy ratio (DP:DE) achieved by reducing the dietary digestible protein levels with or without an associated increase in the dietary non-protein digestible energy supply has proven to be extremely efficient in improving nitrogen utilisation and decreasing nitrogenous losses in numerous farmed species (reviewed by NRC, 2011). Studies including that of Yamamoto et al. (2005) show that rainbow trout diets supplemented with all the limiting essential amino acids allow for the reduction of protein level from 45 to 35 percent, without compromising performance and even improving protein retention efficiency from 35 to 50 percent, reducing nitrogen loading into the environment. In their 2009 rainbow trout study Gaylord and Barrows also showed that by keeping a similar dietary energy level and sup-

plementing diets with methionine, lysine and threonine on an ideal protein basis, dietary crude protein level can be reduced from 46 to 40.9 percent without affecting growth and even improving protein retention efficiency. Again, these achievements would not have been possible were free amino acids not utilised as efficiently as protein-bound amino acids in meeting the requirements of fish. Also very important to bear in mind is that although the DP:DE ratio is a more rational way of expressing protein requirements than dietary crude protein requirements, it must not be taken as a fixed or accurate value. At low DP:DE ratios, fat was shown to constitute a more effective source than digestible starch in improving protein utilisation efficiency in rainbow trout, underlining the importance of non-protein energy sources (lipids and starch) in maximising amino acid utilisation in salmonids (Figueiredo-Silva et al., 2013). Furthermore, the efficiency with which essential amino acids are used by rainbow trout for protein deposition is not constant, but affected by their concentration in the diet (the law of diminishing returns) and intake of digestible energy (Encarnao et al., 2004). Efficiency is also known to decrease significantly with increase in live body weight (reviewed by

NRC, 2011). The development of more sustainable diets requires adjustment of their digestible AA and energy content according to the different stages of production..

Improving health through nutrition

Several studies demonstrate that supplementation of aquaculture feeds with functional amino acids such as arginine and tryptophan constitute a promising approach to improve, among other parameters, animals immune response to environmental stresses. This does not constitute a surprise, since amino acid function goes beyond meeting the requirements for protein synthesis. Physiological response to stress and anxiety involve the serotonin signalling system that responds significantly to the availability of serotonin precursor tryptophan. The connection between tryptophan and serotonin explains why supplementation of the essential amino acid has been found to reduce aggressive behaviour and stress-induced anorexia in several fish, as well as in terrestrial animals and humans. Interestingly, it was recently shown that supplementing the diets of salmon smolts with tryptophan above the recommended level can suppress their cortisol response

18 | INTERNATIONAL AQUAFEED | January-February 2014

FEATURE after being exposed to confinement stress (Basic et al. , 2013). Fish also have particularly high requirements for dietary arginine because it is abundant in protein and tissue fluid (as phosphoarginine, a major reservoir of ATP), and its de novo synthesis is limited or even completely absent. The dietary supplementation of arginine and glutamate in combination had positive effects on feeding rate and growth among Atlantic salmon during the first autumn after sea transfer (Oehme et al., 2010). Dietary arginine has also been shown to improve disease resistance and modulate the innate immune mechanisms of fish (Costas et al., 2011). Furthermore, increased dietary arginine seems to activate polyamine turnover and -oxidation in the liver of juvenile Atlantic salmon, and may act to improve the metabolic status of the fish (Anderson et al., 2013). Although this requires further confirmation, supplementation of aquaculture feeds with functional amino acids such as arginine and tryptophan constitutes a promising approach to reduce the stress associated with aquaculture practices, and ultimately to improve the growth performance of fish. reducing fishmeal inclusion level, and a prom- fed under practical conditions. Nutritional ising one for reducing the crude protein level Requirements for Fish and Shrimp, published in salmonid diets. Such findings confirm not by the US National Research Council (NRC), only the effectiveness of supplemental amino determines requirements for amino acids, acids in covering the nutritional requirements fatty acids, vitamins and minerals based on of fish, but also contribute to the increas- diets containing purified and chemicallyingly well-understood perception that animals defined ingredients which are highly digestdont have requirements for ingredients or ible to the organism. When formulating diets crude protein levels per se, but instead for nutrients, including amino acids. "Are the currently recommended Although Atlantic dietary essential amino acid salmon is the most successfully farmed salmonid, levels (NRC, 2011) effective in the nutrient requirements of the species throughout maximising performance of animals the entire production cycle fed under practical conditions?" still need to be completely defined. Besides, there is a high variability in reported essential amino acid requirements between from practical feedstuffs, it must be taken studies. This has been attributed to methodol- into account that nutrient bioavailability for ogy issues, but also to the composition of the the animal will usually be less than from puridiet used, and whether fish were able to reach fied sources. Another limitation of the currently recomtheir maximum growth potential. In fact, different proteins are not identical in their nutritive mended dietary amino acid values is the fact value, with amino acid profile and digestibility that a single value is offered to cover the entire production cycle. We should, therefore, varying markedly among ingredients. This raises the question whether the make an extra effort to fulfil information gaps currently recommended dietary essential about the nutrient requirements of salmonids, amino acid levels (NRC, 2011) are effec- and to offer recommendations according to tive in maximising performance of animals specifications of each stage of production.

Additional considerations
Evidence is accumulating that balancing the dietary amino acid profile with supplements can be regarded as a cost-effective strategy in

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January-February 2014 | INTERNATIONAL AQUAFEED | 19

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