Glomerulonephritis and immunosuppression associated with dietary essential fatty acid deficiency in gilthead sea bream, Sparus aurata L., juveniles

An experiment was conducted to determine the effect of an essential fatty acid (EFA) deficient diet on growth, immune status and renal morphology of juvenile gilthead sea bream, using two diets: a control diet containing 2% (DW) of n-3 high unsaturated fatty acid (n-3 HUFA) and a diet formulated to be deficient in EFA for this species and containing 0.5% DW of n-3 HUFA (diet NFA). After 9 weeks of feeding the EFA-deficient diet fish showed a reduction in growth compared with the control group (107.48 +/- 9.14 and 123.14 +/-11.87 g final weight respectively). Fish fed the NFA diet showed a reduction in the erythrocyte volume together with increased erythrocyte fragility, haemoglobin content and red blood cell count in comparison with fish fed the control diet. EFA deficiency also reduced cellular immunity in terms of neutrophil activity and the number of circulating lymphocytes. The serum alternative complement pathway was markedly reduced in fish fed the EFA deficient diet. Fish fed the control diet showed glomeruli with a well-defined Bowman's space and normal renal tubes. However, up to 88% of fish fed the EFA-deficient diet showed alterations in renal morphology affecting at least 50% of the glomeruli, which showed extreme dilation of capillaries and occlusion of Bowsman's capsule. Mesangial proliferation and diffuse thickening of the capillary walls, as well as renal tube degeneration, were also observed.     

Influence of n-3 highly unsaturated fatty acid deficiency on the lipid composition of broodstock gilthead seabream (Sparus aurata L.) and on egg quality

A feeding experiment was conducted on gilthead seabream (Sparus aurata) broodstock to investigate the incidence of n-3 highly unsaturated fatty acids (n-3 HUFA) dietary deficiencies on the lipid composition of female liver, gonads and eggs, in relation to spawning quality. Broodstock were fed a control (C) diet or a n-3 HUFA deficient (D) but linolenic acid rich diet. After 20 weeks of feeding, the results showed that levels of total neutral (TNL) and total polar (TPL) lipids of female gonads and eggs were independent of diet. However the fatty acid composition of phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylinositol (PI) of female liver, gonads and eggs in the two groups of fish showed marked differences, reflecting the influence of fatty acid levels in the broodstock diets. This influence was even higher in TNL than in the phospholipid classes examined. In fish fed n-3 HUFA deficient diet, fatty acid composition of TNL of female gonads and eggs reflected the diet more than liver. A higher egg production in broodstock fed C diet (1.8% n-3 HUFA in diet) was extended to spawning quality such as percentages of fertilised and hatched eggs.     

Enhancement of gilthead seabream (Sparus aurata) larval growth by dietary vitamin E in relation to two different levels of essential fatty acids

The objective of this study was to determine the effect of dietary vitamin E on gilthead seabream (Sparus aurata) growth and survival, at two different highly unsaturated fatty acids (HUFAs) levels. Eighteen days old gilthead seabream larvae were fed four formulated experimental diets combining two different dietary levels of HUFAs (M: medium 2.5 + 1.5, DHA + EPA, H: high 5 + 2.5 DHA + EPA g per 100 g) with two different levels of vitamin E (M: medium 540 mg kg^?1, H: high 2900 mg kg^?1): MM, MH, HM, HH (HUFA/vitamin E). After 2‐week feeding trial, the average survival rate was 52.6% and there were no significant differences found among treatments. Increase in vitamin E up to high level markedly improved larval growth, particularly when dietary HUFA levels were lower, suggesting a higher protection value when these fatty acids are more limiting. At medium dietary HUFA levels, increase in vitamin E from medium to high level enhanced larval growth performance in terms of total length. Moreover, increase in vit E enhanced HUFAs content in the larval polar lipids denoting the anti‐oxidative effect of vitamin E.     

Effect of broodstock diets on lipid classes and their fatty acid composition in eggs of gilthead sea bream (Sparus aurata L.)

Total lipid content, total lipid fatty acids, lipid classes and their fatty acid compositions were studied in eggs of gilthead sea bream from two different broodstocks. The two broodstock diets used had the same total lipid content, but differed with respect to their fatty acid levels. The lipid class composition of eggs from different origins was not significantly different. The predominant polar lipids were phosphatidylcholine, phosphatidylethanolamine and sphingomyelin, and the main neutral lipid classes were cholesterol esters and triacylglycerols. The fatty acid composition of total lipids and main lipid classes in the two groups of eggs showed marked differences, reflecting the influence of fatty acid levels in the broodstock diets.     

Modification of essential fatty acid composition in broodstock of cultured European eel Anguilla anguilla L

Farmed eels had lower levels of arachidonic acid (20:4 n‐6) (ARA) and higher ratios of eicosapentaenoic acid (20:5 n‐3) (EPA):ARA compared to wild European eels collected from the Baltic Sea and southern Norwegian coast. Eels fed a formulated feed (JD) with a distribution of essential fatty acids (EFA) resembling wild European eel were sampled after 0, 5, 10, 14 and 44 weeks of feeding to examine changes in fatty acid composition (FAC) in ovaries, visceral fat and muscle. The results showed a slow but steady incorporation of EFA. Lipids are incorporated in the oocytes early in oogenesis, and the leading cohort of oocytes is rich in lipid droplets before the onset of vitellogenesis. This indicates that feeding with optimized broodstock feeds should start early to allow the incorporation of EFA in the first cohort of oocytes. At least 14 weeks of feeding is required to change lipid EFA in broodstock eel to resemble EFA in the diet or in wild fish. After 44 weeks of feeding, ARA was significantly higher in the neutral lipids of ovaries (1.9%) compared to visceral fat (1.2%) or muscle (1.0%). EPA:ARA ratios decreased two‐ to threefold in all tissues examined during that time. ARA and docosahexaenoic acid (22:6 n‐3) (DHA) had accumulated in ovarian polar lipids.     

Egg and larval quality,and egg fatty acid composition of Eurasian perch breeders (Perca fluviatilis) fed different dietary DHA/EPA/AA ratios

In Eurasian perch (Perca fluviatilis), the variability in spawning quality is a major limiting factor for successful production, especially when breeders are fed with an artificial diet. The influence of the dietary DHA/EPA/AA ratio on the egg and larval quality and on the fatty acid and lipid class composition of eggs has been investigated in perch broodstock. Two experimental diets (16% lipids) with two different DHA/EPA/AA ratios, D1 (3/2/2) and D2 (23/9/1), were compared with a natural diet consisting of cultured carp juveniles, CC (10/10/1) and with a commercial diet for salmonids, CDS (14/16/1). Percentages of fertilization and hatching were comparable between fish fed D1, D2 and CC, with the highest hatching rate observed for D1 (63.5 ± 3.8%). These diets supported better values than the CDS. Larval survival and TL50 observed after osmotic stress were higher for the D1 group, followed by larvae produced by fish fed D2 and CC. Larvae from fish fed D1, D2 and CC were significantly more robust than larvae from the CDS group. Differences were observed regarding the fatty acid (FA) profile in the eggs, which was related to the dietary FA composition. The results indicate that a ratio of 3/2/2 seemed to be effective for obtaining eggs and larvae of good quality.     

Alteration of liver and muscle fatty acid composition in gilthead seabream (Sparus aurata) juveniles held at high stocking density and fed an essential fatty acid deficient diet

The aim of the present study was to determine the combined effect of both stress and EFA deficiency on several biological and biochemical parameters. Fish were fed during 15 weeks two isocaloric and isoproteic diets: a control diet based on fish oil and formulated to meet the n-3 HUFA requirements for this species (1.5% of n-3 HUFA) and a deficient diet containing beef tallow and formulated to be deficient in n-3 HUFA. Each experimental diet was evaluated both at high and low stocking densities (10 and 3.2 kg m^–3 of initial density, respectively).High stocking density produced a chronic stress situation with elevation of plasma cortisol levels. It also caused a reduction in hepatosomatic index and liver lipid contents, increasing the oleic acid/n-3 HUFA ratios in the polar lipids. Fish fed the EFA deficient diet at low stocking density showed common deficiency symptoms. High stocking density in fish fed the EFA deficient diet induced a higher degree of EFA deficiency symptoms leading to mortality, liver steatosis, liver lipid deposition, reduced muscle lipid and reduced n-3 HUFA contents, which particularly affected EPA, but not DHA, suggesting a preferential retention of the latter fatty acid, specially in the phosphoglycerides fraction.     

Nutritional stimuli of gilthead seabream (Sparus aurata) larvae by dietary fatty acids: effects on larval performance,gene expression and neurogenesis

The concept of nutritional programming raises the interesting possibility of directing specific metabolic pathways or functions in juvenile fish, for example, to improve the use of substitutes to fishmeal and oil, and hence to promote sustainability in aquaculture. The aim of the study was to determine effects of early nutritional stimuli of gilthead seabream larvae and check if nutritional programming of gilthead sea bream is possible between 16 days post hatching (dph) and 26 dph. A trial was conducted to determine the effects of early nutritional stimuli of gilthead seabream larvae. Five experimental microdiets (pellet size <250 μm) were formulated containing five different proportions of a marine lipid source rich in long‐chain polyunsaturated fatty acids (LC‐PUFA) and two vegetable lipid sources rich in linolenic and linoleic acids. The results of this study demonstrate that dietary n‐3 LC‐PUFA levels increased larval growth and survival affecting Δ6 desaturase gen (fads2) expression and retinal neurons density. However, the high mortalities obtained along on‐growing in fish fed low n‐3 LC‐PUFA at 16 dph constrained the feasibility of nutritional programming of gilthead seabream during this late developmental window and needs to be further investigated.     

Pigmentation of gilthead seabream, Sparus aurata (L. 1875), using Chlorella vulgaris (Chlorophyta, Volvocales) microalga

A feeding experiment was conducted over 9 weeks with seven groups of 30 (fish per group) unpigmented gilthead seabream, Sparus aurata (L. 1875) (initial mean weight = 145.2 ± 12.3 g). Three experimental diets were prepared by adding to a basal diet free of carotenoid (final pigment content of around 40 mg per kg feed): (i) a biomass of the carotenogenic Chlorella vulgaris (Chlorophyta, Volvocales); (ii) a synthetic astaxanthin; and (iii) a mixture (1:1) of microalgal biomass and synthetic astaxanthin. At 3‐week intervals, five fish were sampled from each tank for total carotenoids analysis in skin and muscle. The carotenoid pigments (total amount = 0.4%) identified in the carotenogenic alga were lutein (0.3%), β‐carotene (1.2%), canthaxanthin (36.2%), astaxanthin, free and esterified forms (55.0%), and other pigments (7.3%). Carotenoid pigments were significantly deposited in the four skin zones studied during the feeding trial: the forefront between the eyes, the opercule, along the dorsal fin and in the abdominal area. In the muscle, regardless of the astaxanthin source, the amount of carotenoids measured was very low (less than 1 mg kg^?1) and differences not significant. Moreover, no muscle pigmentation was evident, and there was no variation in the amount of carotenoid analysed in skin tissue, through the trial, for each treatment. It was concluded that supplementing the feed with C. vulgaris would be an acceptable practice in aquaculture to improve the market appeal of the gilthead seabream.     

Importance of the relative levels of dietary arachidonic acid and eicosapentaenoic acid for culture performance of gilthead seabream (Sparus aurata) larvae

The effect of different arachidonic acid (ARA) dietary contents at several dietary eicosapentaenoic acid (EPA) levels on the growth, survival and biochemical composition of gilthead seabream larvae was studied to better define the importance of this fatty acid as a function of EPA. Larvae of 18 days were fed one of the five isonitrogenous and isolipidic microdiets with three different EPA (0.3%, 2% and 4%) and ARA amounts (0.1%, 0.6% and 1.2%). Although a dietary increase in either ARA or EPA alone did not improve survival significantly, the increase in both fatty acids significantly enhanced growth and survival, suggesting an optimum dietary value of EPA:ARA close to 4:1.2. Dietary ARA was more efficiently incorporated into larval tissues than EPA. Increased dietary EPA or ARA contents reduced the incorporation of ARA or EPA into larval lipids, indicating their competition as substrates for different enzymes. The possible negative effect of further elevation of dietary ARA and its competition with EPA for phospholipids synthesis deserves further studies in marine fish larvae.     

Settling velocity and total ammonia nitrogen leaching from commercial feed and faecal pellets of gilthead seabream (Sparus aurata L. 1758) and seabass (Dicentrarchus labrax L. 1758)

The physico-chemical characteristics of particulate wastes of Sparus aurata and Dicentrarchus labrax were investigated. Changes in the dimensions, settling velocity and total ammonia nitrogen (TAN) leached from commercial feed pellets were investigated after soaking. Also, the settling velocity and TAN leached from faecal pellets of these fish were assessed at 15 and 25 °C. The settling velocity of feed pellets was influenced positively by pellet weight and negatively by immersion length as a result of changes in pellet dimensions after soaking. The settling velocity of faecal pellets was determined by pellet weight. The experimental design did not allow identification of any consistent effect of water temperature on settling velocity. Total ammonia nitrogen leaching over time from feed and faecal pellets was successfully explained by means of a first-order kinetic equation. For feed pellets, water temperature significantly affected the speed of the process and the time at which the maximum TAN leached was reached, but did not influence the maximum TAN leached. Leaching was related to feed pellet size, and so the smaller the pellet, the higher the leaching. Total ammonia nitrogen leaching from faecal pellets was greater per unit weight than in feed pellets. However, neither water temperature nor fish species influenced TAN leaching from faeces.     

Changes in the digestive enzymes and hormones of gilthead seabream larvae (Sparus aurata, L. 1758) fed on Artemia nauplii enriched with free lysine

Variations in digestive enzymes and hormones during the larval development of gilthead seabream (Sparus aurata) fed on live prey (Artemia nauplii) enriched with free lysine were investigated for 16 days (from day 24 to day 40). Prior to initiation of the experiment, newly hatched larvae were transferred from incubators to fiber glass tanks (300 l) with black walls and fed the same diet until day 24. Each experiment was performed in triplicate. The amount of free lysine in the Artemia nauplii was increased by adding a 5.3 mM free amino acid solution to the culture water during a 16-h enrichment period. Larvae were sampled four times at four-day intervals, for 16 days. Larvae fed on the control group had lower growth than those fed on the lysine group at the end of the study but the difference was not significant (P > 0.05). The highest trypsin activity and lap/leu–ala peptidase ratios were observed in the control group (P < 0.05). Significant differences between bombesin activities of the treatment groups were not found in the fifth minute after the start of feeding (P > 0.05) but were significant in the fifteenth minute (P < 0.05). Significant differences between the CCK levels of the treatment groups were found (P < 0.05). In conclusion, three main ideas are presented in this study. First, Artemia nauplii can successfully be enriched with free lysine. Second, bombesin (GRP) and cholecystokinin (CCK) activity can be stimulated by free lysine. Third, the mechanisms controlling adaptation of the activity of the trypsin to the amount of dietary protein were not activated until day 40.     

Growth response and fatty acid composition of juvenile red claw crayfish (Cherax quadricarinatus) fed different sources of dietary lipid

A study was conducted to determine the effects of different sources of dietary lipid that differ in fatty acid (FA) composition on growth, feed conversion ratio (FCR), survival, and whole‐body proximate and FA composition of juvenile red claw. Juvenile red claw (0.55 ± 0.02 g) were reared over a 12‐week period. Five practical diets were supplemented with 70 g kg^?1 oil (by weight) from either linseed oil (LO), canola oil (CAO), corn oil (CO), beef tallow (BT) or menhaden oil (MO) and formulated to contain 400 g kg^?1 crude protein. Red claw were fed three times daily (800, 1200 and 1600) to controlled excess. At the conclusion of the experimental period, there were no significant differences (P > 0.05) found in percentage survival and FCR among dietary treatments and values averaged 85.4% and 3.4, respectively. However, final mean weight and specific growth rate of red claw fed Diet 1 (LO) was significantly higher (16.44 g and 3.95% day^?1, respectively) compared with that of red claw fed Diet 4 (BT; 12.24 g and 3.43% day^?1, respectively), but not different from red claw fed the other three diets. Likewise, red claw fed Diets 1 (LO) and 2 (CAO) had significantly higher percentage weight gain (2990 and 2880%, respectively) compared with the BT diet (2124%), but not different from red claw fed Diets 3 (CO) and 5 (MO). Whole‐body fat and ash composition was significantly affected (P < 0.05) by source of added lipid, but no differences were found in whole‐body moisture and protein. Moreover, whole‐body FA composition showed differences among the varying oil sources and primarily reflected the FA in the diets. Results of the present study indicate that plant oils LO, CAO and CO rich in linolenic acid (18:3n‐3), linoleic acid (18:2n‐6) and oleic acid (18:1n‐9) perform as well as MO containing high levels of n‐3 highly unsaturated fatty acids eicosapentanoic acid (20:5n‐3) and docosahexaenoic acid (22:6n‐3) for juvenile red claw crayfish grown indoors lacking natural food items. Further, red claw fed a diet containing BT, which has a high percentage of saturated FA, performed poorly compared with LO and CO in regards to weight gain. It appears that juvenile red claw can be fed diets containing less expensive plant‐based oils with high levels of 18‐carbon unsaturated fatty acids. This could reduce diet costs for producers and allow for profitability.     

Study of the dose of thyme essential oil in feed to prolong the shelf life of gilthead seabream (Sparus aurata)

The effect of thyme essential oil (Thymus zygis, subspecies gracilis) on the quality and shelf life of gilthead seabream was studied, when added to the diet: a control diet and four experimental diets (T500, T1000, T1500, T2000) with 500, 1000, 1500 and 2000 mg kg^?1 of thyme essential oil, respectively. After 12 weeks of experimentation, the fish were stored on ice at 4 °C for 0, 7, 14 and 21 days. Physical–chemical, microbiological and sensory analyses were carried out at each sampling point to determine the degree of deterioration. A dose‐dependent effect was observed on the colour, TBARS and total volatile basic nitrogen during the storage. Microbiological counts were lower for Enterobacteriaceae and coliforms at high doses of the essential oil. The sensory analysis showed an effect on the quality index that was not dependent on the dose, which extended the shelf life of gilthead seabream from 17 to 18 days for all doses.     

Incorporation and metabolism of 14C-labelled polyunsaturated fatty acids in juvenile gilthead sea bream Sparus aurata L. in vivo

The incorporation, and the capacity for desaturation and elongation in vivo, of intraperitoneally-injected, ¹⁴C-labelled n–3 and n–6 C₁₈ and C₂₀ PUFAs were investigated in juvenile gilthead sea bream, Sparus aurata. The results indicate that juvenile gilthead sea bream have only limited ability to convert CH PUFAs to C₂₀ and C₂₂ HUFAs in vivo. The data are consistent with the results from nutritional studies on larvae, postlarvae and fingerlings that have shown that gilthead sea bream require the provision of preformed eicosapentaenoic and docosahexaenoic acids in the diet. The impairment in the desaturase/elongase pathway was quantitatively and qualitatively similar to that found in turbot, Scophthalmus maximus, being at the level of the 5-desaturase. The low activity of 5-desaturase combined with the consistent finding that arachidonic acid is selectively retained in membrane phosphatidylinositol suggests that, in addition to eicosapentaenoic and docosahexaenoic acids, gilthead sea bream may also have a requirement for preformed arachidonic acid in the diet.Abbreviations AA 5,8,11,14-eicosapenaenoic acid (arachidonic acid, 20:4n–6) - CPL diradyl (diacyl + alkenylacyl + alkylacyl) glycerophosphocholine - DHA 4,7,10,13,16,19-docosahexaenoic acid (22:6n–3) - EPA 5,8,11,14,17-eicosapentaenoic acid (20:5n–3) - EPL diradyl (diacyl, alkenylacyl + alkylacyl) glycerophosphoethanolamine - HUFA highly unsaturated fatty acids ( C₂₀ and with 3 double bonds) - LA 9,12-octadecadienoic acid (linoleic acid, 18:2n–6) - LNA 9,12,15-octadecatrienoic acid (-linolenic acid, 18:3n–3) - PI phosphatidylinositol - PS phosphatidylserine - PUFA polyunsaturated fatty acid(s)     

Mono- and duoculture of juvenile sharpsnout seabream Diplodus puntazzo (Cetti) and gilthead seabream Sparus aurata L. in a recirculated water system

Although the growth of intensively reared sharpsnout seabream Diplodus puntazzo (DP) has been suggested to improve in duoculture with gilthead seabream Sparus aurata (SA), the stocking ratio for most effective duoculture rearing has not been investigated. For this purpose, juvenile D. puntazzo (1.68±0.011 g) and S. aurata (1.43±0.007 g) were reared in duplicated groups of 165 specimens each, in two monoculture (100% DP, 100% SA) and four duoculture combinations (80% DP–20% SA, 60% DP–40% SA, 40% DP–60% SA, 20% DP–80% SA) for 96 days under recirculated water system conditions. Both species exhibited the highest growth when stocked at 20% and the poorest growth when stocked at 80%. For each species, no major differences for coefficient of weight variation and carcass proximate composition were detected. As the percentage of stocked S. aurata increased, D. puntazzo aggressiveness and weight variability of whole population were reduced, while total biomass, food consumption and food conversion ratio were increased. The results obtained are discussed in relation to fish behaviour and social interactions. From the fish farming point of view, it is indicated that under the present experimental conditions, juvenile S. aurata should be reared in monoculture and D. puntazzo in duoculture 40% DP–60% SA.     

Influence of broodstock gilthead seabream (Sparus aurata L.) dietary fatty acids on egg quality and egg fatty acid composition throughout the spawning season

The influence of broodstock dietary lipids on egg quality and egg fatty acid composition throughout the spawning season of gilthead seabream was investigated. For this purpose, the fish were fed for 7 months either a control diet (diet C) or a diet deficient in n−3 highly unsaturated fatty acids (n−3 HUFA) but rich in both oleic (18:1n−9) and linolenic (18:3n−3) acids (diet D). Eggs spawned by both groups of fish were sampled at the beginning, middle and end of the spawning season and the fatty acid composition of their neutral (NL) and polar lipids (PL) determined. In the early season, percentages of fertilized and hatched eggs, relative proportions of NL and PL as well as their fatty acid compositions, were not affected by the lipid composition of the broodstock diet. However, the eggs spawned during the middle and late seasons showed marked differences among the two groups of fish, clearly reflecting the influence of dietary fatty acids. This influence was more evident in the neutral lipid fraction than in the polar lipids. No correlation was found between the number of buoyant eggs and eicosapentaenoic (20:5n−3, EPA), docosahexaenoic (22:6n−3, DHA) fatty acids or total n−3 HUFA contents in egg phospholipids. However, a negative correlation was detected when percentages of fertilized eggs were compared with the levels of 18:1n−9, 18:3n−3 and with the ratio 18:1n−9/n−3 HUFA present in the phospholipids. Our results indicate the importance of maintaining not only the level of n−3 HUFA in egg membrane phospholipids, but also the balance between n−3 HUFA and other fatty acids such as 18:1n−9 and 18:3n−3, in order to obtain a high spawning quality.     

Reproductive performance and offspring quality of Chinese mitten crab Eriocheir sinensis (H. Milne-Edwards) females fed an optimized formulated diet and the razor clam Sinonovacula constricta

After feeding female Eriocheir sinensis on an optimized formulated diet or fresh razor clam Sinonovacula constricta for 7 months, their reproductive performance and offspring quality were compared. To evaluate diet nutrient contents, the proximate, fatty acid and amino acid compositions of the formulated diet and the razor clam were analysed. The nutritional value of the diets was determined by assessing survival, gonadosomatic index (GSI) and hepatosomatic index (HSI) of female crabs from both diet treatments, together with the percentage of females that spawned, total egg production per female and fecundity (number of eggs g^?1 female wet weight). Furthermore, the quality of eggs and newly hatched larvae from the two dietary treatments were determined using the following parameters: egg diameter, wet weight and dry weight, hatchability, proximate and fatty acid profile of eggs, larval carapace length, resistant to starvation and osmotic shock, larval survival and development to the zoea II stage. Higher protein, phospholipids (PL) and amino acids (AA) contents were found in the razor clam while the formulated diet contains higher levels of ash, total lipid (TL) and 18:1n‐9, 18:2n‐6 and 22:6n‐3 fatty acids. Although female crabs fed the two different diets showed similar reproductive performances, newly hatched zoea I larvae produced by the crabs fed the formulated diet had significantly longer mean carapace length and shorter development time to the zoea II stage under identical culture condition (P<0.05). Moreover, dietary fatty acid appeared to have more significant effects on the fatty acid composition of the hepatopancreas than it did on mature ovaries or eggs. This suggests that the fatty acid profile of mature ovaries is indicative of the specific fatty acid required for ovarian development in E. sinensis. In conclusion, our results show that the optimized formulated diet developed in this laboratory can totally replace the razor clam, a broodstock food widely used in E. sinensis hatcheries in China. This encouraging result should facilitate more reliable hatchery production of this important aquaculture species.