Peter Coutteau

Review on the dietary effects of phospholipids in fish and crustacean larviculture.

A beneficial effect of dietary phospholipid (PL) supplementation in purified diets in terms of survival, growth, resistance to stress tests, and occurrence of deformities has been demonstrated in larval and juvenile stages of various species of fish and crustaceans. The exact determination of PL requirements in larvae is complicated due to the difficulty to bio-encapsulate PL in live prey. Furthermore, the great variety in purity and composition of the PL sources, and the experimental conditions (such as diet formulation and extent of co/prefeeding with live food) makes it difficult to compare requirements determined with artificial diets. Larval stages are extremely sensitive to a dietary PL deficiency and require higher levels of dietary PL than juveniles. For most of the fish and crustacean species examined, the estimated PL requirement of larvae are in the range of 1–3% phosphatidylcholine + phosphatidylinositol (PC + PI) of diet dry weight. The absence of a PL requirement in the freshwater prawn Macrobrachium rosenbergii exemplifies the important species differences. The few studies evaluating single PL demonstrate that PC and PI are the most efficient in most species. The presence of an unsaturated fatty acid in sn−2 position of the PL molecule seems to be essential for the functionality of PL. Some studies in crustaceans reported a relation between PL requirements and the protein source in the diet. Various hypotheses have been formulated to explain the effect of PL. The PL effect is not related to the provision of choline, inositol or essential fatty acids (EFA). However, PL may be superior to neutral lipids for larvae as a source of EFA and energy due to their better digestibility. PL may improve the performance of the diet by improving the water stability of food particles, or by their action as antioxidant or feed attractant. The effect of dietary PL appears not to be explained by their emulsifying ability. However, there are proofs that dietary PL interfere with lipid transport, especially cholesterol transport in crustaceans, and with retention of fatty acids provided by dietary triacylglycerol. Although the origin of the requirement is still unclear, dietary PL supplementation has potential importance for the formulation of practical larval diets.

The stability of docosahexaenoic acid in two Artemia species following enrichment and subsequent starvation.

Two Artemia species (Artemia franciscana, GSL strain and a population from Artemia sinica, ARC No. 1188) were enriched with two different emulsions containing high levels of docosahexaenoic acid (DHA) (23 and 44% of total fatty acids, respectively) in filtered seawater (30 ppt) at 28 °C. After enrichment nauplii were starved for 72 h at three different temperatures (6.1, 12.2 and 22 °C). Following enrichment the DHA content and DHAEPA ratio reached a maximum 41.2 and 42.8 mg g−1 dry weight and 1.88 and 2.09 in A. franciscana and A. sinica, respectively. During starvation at 12.2 °C the DHA content in A. franciscana decreased steadily throughout the starvation period towards 1.11–2.89 mg g−1 dry weight (DHAEPA ratio of 0.4) after 72 h, whereas in A. sinica the DHA content was > 20 mg g−1 dry weight (DHAEPA ratio of 1.7–2). A quantitative reduction of DHA in A. sinica was observed only during the first 24 h of the starvation period. The degradation rate of DHA in A. franciscana increased with rising temperature (22 °C) and decreased at the lowest temperature (6.1 °C). This was not the case for A. sinica, during starvation at different temperatures the DHA level in A. sinica remained at a relatively high and constant level.

Biochemical and enzymatic characterization of decapsulated cysts and nauplii of the brine shrimp Artemia at different developmental stages

Abstract Decapsulated Artemia cysts were used to study the factors which induce the superior performance of live organisms as food for fish larvae. The biochemical composition, the in vitro protein digestibility, and the total proteolytic and trypsin activities in cysts and nauplii of Artemia were determined at different developmental stages as a function of incubation time. Six different incubation times were studied: 1, 6, 11, 16, 21 and 25 h of development, which cover cyst and early nauplii stages. The individual dry weight of Artemia decreased through development. The individual protein and lipid content ( μ g ind −1 ) remained constant during development until the time of hatching after which they decreased slightly. However, no significant incubation effect was found. Small changes in amino acid and fatty acid composition were found during development, but is assumed that they are too small to be of nutritional importance to fish larvae. No major changes were observed in protease activities measured at acid and alkaline pH during the first 25 h of development. From the alkaline proteases, no significant change in trypsin activity was detected during cyst and early nauplii development. From the point of view of exogenous enzyme contribution to fish larvae, there seems to be no difference whether feeding decapsulated cysts or newly hatched nauplii, since no difference in qualitative protease composition was found during the first 25 h of Artemia development. The relative contribution of Artemia proteases to the digestion of food by fish larvae is discussed.

The effect of different levels and sources of dietary phosphatidylcholine on the growth, survival, stress resistance, and fatty acid composition of postlarval Penaeus vannamei.

Abstract The effect of dietary purified phosphatidylcholine (PC) was evaluated on growth, survival, resistance to osmotic shock, and fatty acid composition of early postlarval Penaeus vannamei (0.3 mg initial dry weight) fed semi-purified diets. PC sources used were purified soybean PC (SPC, 95% purity), chicken-egg PC (EPC, 94% purity), and de-oiled soybean lecithin (DSL, 23% PC). The growth response of shrimp fed 1.5% of SPC or 6.5% of DSL was significantly greater than that of shrimp fed a PC-deficient diet, whereas no effect was observed either on survival or stress resistance. Further increasing the dietary level of soybean PC from 1.5% to 3.0% resulted in a significant decrease of the shrimp weight gain. Shrimp receiving 1.5% of PC, provided either as chicken-egg PC, soybean PC, or de-oiled soybean lecithin did not show differences in growth, but had a significantly greater weight gain than that of shrimp fed 1.5% of de-oiled soybean lecithin, which indicated that the phospholipids in lecithin other than PC cannot compensate for a PC deficiency in the diet. With increasing dietary level of soybean PC, significantly higher levels of 20:1n-9, total n − 6 polyunsaturated fatty acid (PUFA), and 20:5n − 3 were present in the total lipids of shrimp, whereas the proportionate levels of 18:1n − 9 and total monenes significantly decreased. Increasing the level of dietary PC, in particular above 1.5% SPC, resulted in an increase of the proportion of n-3 PUFA and n-6 PUFA in the tissue, and a reduced proportion of saturated and monoenoic fatty acids in the PC of the shrimp. Greater incorporation of n-3 highly unsaturated fatty acid (HUFA) with increasing level of PL in the diet may be explained by an improved utilization efficiency of the ethyl ester-based source, whereas a better incorporation of 18:2n-6 in total lipids and PC of the shrimp may be due to a better availability of this fatty acid provided in the form of a PL rather than triglyceride-based oil.

Use of Brine Shrimp, Artemia spp., in Larval Crustacean Nutrition: A Review

Because of convenience in production and their suitable biochemical composition, brine shrimp Artemia spp. nauplii have been adopted as a standard diet in the commercial larviculture of several crustacean species. The nutritional value of Artemia, however, is not constant, but varies both geographically and temporally. During the past decade both the causes of Artemia nutritional variability and methods to improve poor-quality Artemia have been identified. Enriching Artemia spp. with emulsified lipophilic products is a technique that has allowed delivery of extra doses of essential nutrients, for example, highly unsaturated fatty acids (HUFA) and vitamins, to crustacean larvae. The enrichment technique has limitations, however, because the Artemia spp. currently available selectively catabolize some of the nutrients such as docosahexaenoic acid and phospholipids. Decapsulated Artemia cysts, juveniles, and adult brine shrimp are also used increasingly as suitable diets for different crustacean species.

The effect of lipid supplementation on growth and fatty acid composition of Tapes philippinarum spat

The present study investigated the possible use of emulsions as an artificial lipid supplement to live algae for seed of the Manila clam Tapes philippinarum. The uptake and assimilation of an emulsion, rich in docosahexaenoic acid (DHA, 22:6n−3) and eicosapentaenoic acid (EPA, 20:5n−3) and fed at concentrations of 20 and 40% of the algal dry weight, were verified analytically by fatty acid analysis of the animals and their diets. Dietary requirements for n−3 highly unsaturated fatty acids (HUFA) were examined by supplementing Dunaliella tertiolecta, which contains no polyunsaturated fatty acids longer than 18:3n−3 and Tetraselmis suecica, which contains EPA but only trace amounts of DHA. An algal mixture of Isochrysis galbana (clone T-Iso) and Chaetoceros neogracile (1:1, on dry weight basis) was used as the control diet. After 4 weeks, the lipid supplementation resulted in a significant increase of the DHA level in the seed compared to the animals fed non-supplemented Dunaliella (from 9.5 to 19.8 and 22.0% at a supplementation of 20 and 40%, respectively) or Tetraselmis (from 3.4 to 24.8 and 26.9%, at a supplementation of 20 and 40%, respectively) diet. Feeding solely D. tertiolecta resulted in a significantly lower daily growth rate (DGR) compared to animals fed T. suecica or the mixed algal diet. Lipid supplementation improved the DGR of the clams fed D. tertiolecta while hardly any effect could be detected in those fed T. suecica. The poor nutritional value of D. tertiolecta was indicated by the continuous decrease of the DGR and resulted in a DGR that was no longer significantly different from the starved ones at the end of the experiment.

Comparison of different soybean phospholipidic fractions as dietary supplements for common carp, Cyprinus carpio, larvae

Abstract Two experiments were conducted in order to compare the effects of several phospholipid (PL) sources prepared from soybean lecithin on growth and survival of start-feeding carp larvae. Eight diets were tested in each experiment. All diets contained 94% of the same casein–dextrin mixture and 6% of added lipid, consisting of peanut oil alone or supplemented with a PL source. In both experiments, diets supplemented with fractions enriched in phosphatidylinositol (PI) resulted in the highest survival (95% after 25 and 31 days, respectively). Addition of phosphatidylcholine (PC) allowed high initial growth rates during the first week of feeding, similar to or higher than those obtained with PI, but induced a peak of mortality afterwards and, in the first experiment, a high frequency of deformed fish. Larvae fed PC-enriched diets started to die at a larger size than larvae fed PL-deficient diets. Other tested fractions with phosphatidylethanolamine, phosphatidylserine or phosphatidic acid as the major PL class resulted in intermediate results between those given by PI- and PC-rich fractions. Dietary hydrogenated soybean PC resulted in a decreased growth compared to native soybean PC. Soybean lysolecithin was inferior to original lecithin for final larval growth and survival, but was superior to peanut oil without PL. Present results confirm that both the polar head group and the fatty acid composition of the dietary PL may affect their nutritional value for larval carp and suggest distinct requirements for PI and PC.

Effect of a dietary phospholipid supplementation on growth and fatty acid composition of European sea bass (Dicentrarchus labrax L.) and turbot (Scophthalmus maximus L.) juveniles from weaning onwards

Two 40-day feeding trials using extruded diets were conducted to assess the effect of a dietary phospholipid (PL) supplementation on growth, survival and fatty acid composition of European sea bass (Dicentrarchus labrax) and turbot (Scophthalmus maximus) from weaning onwards. Two dietary treatments (FO and PL) were tested; both had an identical extruded basis (92.5% total diet weight) coated with a different lipid fraction (7.5% total diet weight). Diet PL contained 2% egg yolk PL (69% pure). In diet FO the PL was replaced by hydrogenated coconut oil. The isolipidic diets contained an equal amount of fish oil ethyl esters providing 1.6% (% diet dry weight) of n-3 highly unsaturated fatty acids (HUFA). A diet water stability test showed no effect of the PL supplementation on the leaching of the dietary fatty acids. In both fish species weight, but not survival, significantly increased as a result of PL supplementation. Weaning onto the experimental diets resulted in similar changes in the relative percent levels of fatty acids in both species. In general, the percentage of saturated fatty acids levelled off after a rapid increase, while monoenes increased after an initial decrease. Total n-3 polyunsaturated fatty acids (PUFA) decreased and total n-6 PUFA remained almost constant. The major effect of the dietary PL on fish fatty acid composition was a 50% increase in n-6 and n-3 HUFAs compared to the PL-free FO diet. The rise in n-6 HUFA may have reflected the higher moiety in the dietary PL. On the other hand this was not the case for the n-3 HUFA since they represented only low levels in the PL fraction (0.1%) compared to that provided by the ethyl esters (1.6%) suggesting a more efficient incorporation of the PL n-3 HUFA than of the ethyl ester n-3 HUFA. A second hypothesis is that the dietary PL may have favored the incorporation of the dietary ethyl ester n-3 HUFA.

Dietary impact of algal and artificial diets, fed at different feeding rations, on the growth and fatty acid composition of Tapes philippinarum (L.) spat

Abstract The effect of lipid supplementation and algal ration on growth and fatty acid composition of juvenile clams, Tapes philippinarum was investigated. A diet of Tetraselmis suecica was supplemented with a lipid emulsion and fed at a daily ration of 0.5, 1.0 and 1.5% WW DW−1 day−1. A mixed algal diet of Isochrysis galbana (clone T-Iso) and T. suecica (1:1 on DW basis) and starvation functioned as positive and negative control treatments, respectively. T. suecica, which contains eicosapentaenoic acid (20:5n−3) but no docosahexaenoic acid (22:6n−3), was supplemented with 50% (% of algal dry weight) of a 22:6n−3-rich emulsion. Higher growth rates of T. philippinarum were associated with higher T. suecica feeding rations. Lipid supplementation resulted in significantly better growth rates of T. philippinarum at a T. suecica ration of 0.5, 1.0 and 1.5% WW DW−1 day−1. The growth promoting effect of lipid supplementation was significantly reduced during 4 consecutive weeks or by increasing the algal feeding ration. Increasing T. suecica rations were associated with a decrease in the % composition of 22:6n−3 while the 20:5n−3% composition was hardly affected in the total lipids of T. philippinarum. On the contrary, the absolute concentration of 22:6n−3 (μg per gram dry weight, μg g DW−1) in the total lipids of T. philippinarum was similar at all T. suecica rations while the 20:5n−3 concentration significantly increased. The major effect of lipid supplementation on the fatty acid composition of the clams, was a significant increase of the 22:6n−3% composition in the total lipids, polar lipids and triglycerides. In contrast with 20:5n−3, the effect of lipid supplementation on the absolute 22:6n−3 content (μg g DW−1) was more pronounced at a higher T. suecica ration. After a starvation period of 4 weeks, the concentration (μg g DW−1) of all fatty acids was significantly reduced, except the n−7 monoenoic fatty acids (MUFA) and the non-methylene interrupted dienoic fatty acids (NMID). The concentration (μg g DW−1) of the NMID in starved clams and clams fed different diets was similar to the concentration in the initial sample. This indicated that growing spat was actively synthesising NMID and they were selectively retained in unfed animals. The results indicated that either dietary lipid or 22:6n−3 was a growth-limiting factor when T. suecica was fed at a ration of 0.5, 1 and 1.5% WW DW−1 day−1. A selective retention in starved clams and a preferential accumulation in fed clams was observed for 22:6n−3 but not for 20:5n−3 which indicated a greater importance of 22:6n−3 compared to 20:5n−3.

The use of manipulated baker's yeast as an algal substitute for the laboratory culture of Anostraca

The production of unicellular algae is laborious and is a major constraint for the culturing of aquatic filter-feeders. Because of their small particle size and their high protein content yeasts are considered as a promising substitute for micro-algae. Furthermore, recent work has shown that bakers yeast can be converted into a digestible diet for Artemia by chemical treatment. The present study documents the use at laboratory scale of this manipulated yeast as an algal substitute for the culture of two anostracan species.The experiments were conducted with the brine shrimp artemia franciscana and the fairy shrimp Streptocephalus proboscideus. A similar experimental set-up was used for both species. The algal diet, consisting of Dunaliella tertiolecta for A. franciscana and Selenastrum capricornutum for S. proboscideus, was substituted at various levels by two types of treated bakers yeast: a fresh form and a dried product which was rich in highly unsaturated fatty acids (HUFA).The chemically-treated yeast offers promising possibilities as an algal substitute for Artemia; i.e. replacing 75% of the algae by the dried yeast resulted in similar survival and even higher growth rates in comparison with the reference algal diet; for the treated fresh yeast similar results could be achieved by up to 95% substitution. For S. proboscideus, a substitution of 75% by either of the yeast products resulted in good survival, though growth did not exceed 80% of the observed growth in the algal control. A diet consisting solely of yeast resulted in poor survival for larvae of both species. Experiments were run to investigate whether this was due to a sub-optimal feeding regime, nutritional deficiencies, or deterioration of the water quality.