Jeanne Moal

The effect of monospecific algal diets on growth and fatty acid composition of Pecten maximus (L.) larvae

Abstract Four batches of Pecten maximus (L.) larvae were grown under hatchery conditions and fed the monospecific diets Pavlova lutheri Droop, Isochrysis aff. galbana Green (clone T-iso; termed Tahitian Isochrysis), Chaetoceros calcitrans Takano and Dunaliella tertiolecta Butcher which were selected on their very different polyunsaturated fatty acid (PUFA) profiles. Neutral and polar lipid PUFA compositions of larvae were influenced by diets. The constant level of (n-3) PUFA in neutral lipids of larvae although very different in the diet indicated that neutral lipid composition was partially controlled by the larval metabolism. Similarly, the stability of total saturated, monounsaturated and PUFA levels (25, 15 and 55% of total fatty acids), as well as total (n-3) PUFA level and the preferential incorporation of the 22:6(n-3) in polar lipids clearly indicated a metabolic regulation of the fatty acid composition. The 20:4(n-6) and 22:5(n-6) acids were also preferentially incorporated. The accumulation of 18:3(n-3) or 20:5(n-3) with depletion of 22:6(n-3) in neutral and polar lipids of larvae fed diets with high levels of the first two fatty acids but little or no 22:6(n-3), indicated that elongation-desaturation of dietary fatty acids was too low in P. maximus larvae to maintain high 22:6(n-3) levels in polar lipids. The selective incorporation of dietary long chain PUFA into neutral and polar lipids by acyltransferases may be the main mechanism controlling the fatty acid composition of larvae. These results indicate that P. maximus larvae have essential requirements for long chain (n-3) and (n-6) PUFA, similar to many other marine species.

Changes in the fatty acid composition of Pecten maximus (L.) during larval development

The fatty acid composition of Pecten maximus (L.) larvae grown under hatchery conditions was determined during development from the oocyte stage to the pediveliger stage (Day 23). The levels of total fatty acids in neutral and polar lipids increased 3 days after the first feeding and were ≈10 times higher in pediveligers than in D-larvae (32.46 vs. 3.14 ng·larva−1 and 11.38 vs. 1.23 ng·larva−1 in neutral and polar lipids, respectively). The proportions of total saturated, monounsaturated and polyunsaturated fatty acids in neutral and polar lipids remained stable throughout the development. The fatty acid composition of P. maximus larvae was typical of that described for other marine bivalve larvae since the (n-3) polyunsaturated fatty acids (PUFA) formed 34.4–39.6% of the neutral lipids and 43.0–50.7% of the polar lipids. The major PUFA identified in oocytes and larvae were similar to those supplied by the diet [18:2(n−6), 18:4(n−3), 20:5(n−3) and 22:6(n−3)]. We observed that significant changes of the PUFA composition occurred in neutral and polar lipids during embryogenesis and the subsequent nutritional transition. These changes were expressed by the relative decline of the 20:5(n−3) and 20:4(n−6) percentage levels in both lipid fractions while the 22:6(n−3) increased in neutral lipids and the 22:5(n−6) increased in both neutral and polar lipids. We observed also that polar lipids exhibited a 22:6(n−3) level considerably higher than that of the diet (26.8 vs. 6.8%) and constant throughout development, indicating a preferential incorporation and a major structural role. These results suggest that a metabolic control of the fatty acid composition of neutral lipids and mainly polar lipids takes place in developing larvae. With reference to previous studies on fatty acid metabolism of molluscs, it is probable that this control results from a selective incorporation of dietary fatty acids by acyltransferases rather than a desaturation of dietary precursors. P. maximus larvae seem therefore to have a high nutritional requirement for long chain PUFA and especially the 22:6(n−3).

Effect of a mono-specific algal diet on immune functions in two bivalve species - Crassostrea gigas and Ruditapes philippinarum

SUMMARY The impact of diets upon the fatty acid composition of haemocyte polar lipids and consequently upon immune parameters has been tested in the oyster Crassostrea gigas and the clam Ruditapes philippinarum. Oysters and clams were fed each of three cultured algae: Chaetoceros calcitrans, which is rich in 20:5(n-3) and 20:4(n-6) and poor in 22:6(n-3) fatty acids; T-Iso (Isochrysis sp.), which is rich in 22:6(n-3) and deficient in 20:5(n-3) and 20:4(n-6); and Tetraselmis suecica, which is deficient in 22:6(n-3) and contains only small amounts of 20:5(n-3) and 20:4(n-6). Fatty acid composition of haemocyte polar lipids was greatly affected by the diet. Oysters and clams fed C. calcitrans maintained a higher proportion of 20:5(n-3) and 20:4(n-6) in their haemocyte polar lipids, while these polyunsaturated fatty acids decreased drastically for animals fed T-Iso. However, the T-Iso diet maintained 22:6(n-3) in haemocyte polar lipids of both species. Higher 20:5(n-3) and 20:4(n-6) contents in diets appeared to have a positive effect upon total haemocyte count, granulocyte percentage, phagocytic rate and oxidative activity of clam haemocytes. Similarly, a positive effect of 20:5(n-3) on oxidative activity of oyster haemocytes was observed but to a lesser extent than in clams. Interestingly, when oyster haemocytes are submitted to a stressful condition, a positive effect of a higher dietary 22:6(n-3) content on the phagocytic rate was noticed.

Effect of food fatty acid and sterol quality on Pecten maximus gonad composition and reproduction process

Abstract Spawning individuals of the scallop Pecten maximus were conditioned on three test diets: Tahiti Isochrysis , a mixture (PTSC) and Chaetoceros calcitrans . The scallops fed T- Isochrysis showed a better hatching rate and lower atresia than those fed the other two diets. Proximate composition of the female gonads was not modified by the differences in the diets. Enrichment of gonads with sterol esters and triglycerides, characteristic of storage of lipids, observed with the broodstock fed the diatoms, did not result in successful gametogenesis and spawning. The monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) composition of neutral and polar lipids of gonads was related to the fatty acid composition of the diet. However, the 20 and 22 carbon PUFA were maintained at levels relatively independent of those of the diet; when these fatty acids were low in the diet, their concentration in the lipids of the gonads decreased but still remained significant. This effect was more pronounced in the polar than the neutral lipids. The preferential incorporation of 22:6 n − 3, 20:4 n − 6 and 20:5 n − 3 in the polar lipids indicates their role in gametogenesis and embryogenesis. The sterol composition of the gonads (free sterols and sterol esters) reflected that of the diet; however, the cholesterol, mainly found in the esterified fraction, was maintained at stable levels independent of dietary supply. Two sterols (22-dehydrocholesterol and 24-methylenecholesterol) that were either absent from, or present only in trace amounts in the diets, were found to occur at constant levels in the female gonads, regardless of the nature of the diet.

Comparison of the lipid class and fatty acid composition between a reproductive cycle in nature and a standard hatchery conditioning of the Pacific Oyster Crassostrea gigas

Abstract The lipid composition of Crassostrea gigas was analyzed during the reproductive phase in natural as well as under artificial conditions. The lipid content increased and accumulated in the gonads, but to a higher extend in the naturally conditioned animals. The percentage of neutral lipid in total lipid of the gonad plus mantle was stable, high (>70%) and equal under both conditions, underscoring that the lipid reserves were preferentially located in that organ. The composition of the polar lipid classes was stable with little variations. The fatty acid (FA) composition of the neutral lipids in all organs is influenced by diet. However, there is a different response according to organs. A high dietary impact occured in the digestive gland whereas the muscle was less affected. The polyunsaturated fatty acid (PUFA) level of the neutral and polar lipids in the gonads changed little despite the dietary conditions but the respective proportions of (n-3) and (n-6) PUFA differed drastically as a result of diet composition. There was clear evidence in all organs for a specific accumulation of 22:6(n-3) and 20:5(n-3) in the polar lipids under both conditioning diets. The proportions of 22:6(n-3) and 20:5(n-3) from neutral and polar lipids of oysters conditioned artificially were significantly lower than of those that were naturally conditioned.

Growth and lipid class composition of Pecten maximus (L.) larvae grown under hatchery conditions

Abstract Five independent batches of Pecten maximus (L.) larvae, reared under the same standard hatchery conditions, showed different growth rates, ranging from 1.9 to 6.6 μm·day −1 . A standard lipid profile was defined based on the fast-developing larvae and variations to this standard were seen when abnormal growth performances occurred. The lipid class composition was determined during the development of D-larvae to Day 23 using chromarods and flame ionization detection. Phospholipids and triglycerides were the main lipid classes in P. maximus larvae as reported for other cultured bivalve larvae (accounting for 50 and 37% of total lipids, respectively). Sterols and free fatty acids formed 10 and 3% of the total lipids, respectively. These relative proportions of lipid classes were constant during development of fast developing larvae. The triglyceride:organic matter ratio (TG:OM) stabilized after an initial decline during 3 days after the first feeding and the increase of triglycerides after this time indicated that the catabolism of endogenous triglycerides was rapidly offset by the food intake during the nutritional transition. The comparison of this lipid class pattern with those from larvae which exhibited a lower growth rate revealed that: (i) no stability of the lipid class composition was apparent throughout development; (ii) the triglyceride:total lipid (TG:TL) ratio in D-larvae was not related to the subsequent growth; (iii) a low TG:OM ration in D-larvae was related to poor growth while higher levels did not correspond necessarily to better growth rates; (iv) both the degree and the duration of the TG:OM ration decline for all these batches during the 1st wk can be related to the ability of the larvae to overcome the nutritional transition from trochophore to veliger rapidly and allow for subsequent successful growth. The possible influence of uncontrolled genetic or environmental factors is suggested.

Impact of the quality of dietary fatty acids on metabolism and the composition of polar lipid classes in female gonads of Pecten maximus (L.)

Abstract The impact of dietary deficiency of the essential fatty acids 20:5(n-3), 22:6(n-3) and 20:4(n-6) was studied at the level of different polar lipid classes that constitute the membrane structure of female gonads of Pecten maximus by using three microalgal diets. After 11 weeks of conditioning of the broodstock, the relative percentages of polar lipid classes were very little, or not at all, influenced by the composition of the microalgae. On the other hand, the imprint of dietary fatty acid composition was reflected in the relative composition of fatty acids specific to some classes but not all of them. The dietary fatty acid composition thus clearly influenced that of phosphatidylcholine (PC), the plasmalogens and to a smaller extent the phosphatidylserine (PS) and phosphatidylethanolamine (non plasmalogen PE), but not of phosphatidylinositol (PI) and the glycolipid (GLY). The PI showed a very stable composition and was rich in 20:4(n-6) (>30%) irrespective of the diet. Likewise, the 22:6(n-3) content was consistently higher than 76% in the glycolipid. The maintenance of the specific fatty acid composition of these classes is probably a physiological necessity for gametogenesis and embryogenesis.

Fatty acid composition of polar lipid classes during larval development of scallop Pecten maximus (L.)

The major phospholipid classes in the larvae of the scallop Pecten maximus were phosphatidylcholine (PC) and plasmalogens (PLSM) (35.5 and 32.1 mol%, respectively). The minor classes were glycosyldiacylglycerol-like (GLY), phosphatidylinositol (PI), phosphatidylserine (PS) and non-plasmalogen phosphatidylethanolamine (PE) at 6.7, 9.1, 8.5 and 8.1 mol%, respectively. Abundance of phospholipid classes and their content of saturated, monounsaturated and polyunsaturated fatty acids remained relatively conserved during the course of larval development. During larval development there was a decrease in the amount of 20-carbon polyunsaturated fatty acids (PUFA) in the polar lipids, while the amount of 22-carbon PUFA remained constant. These changes, which occurred mainly during the lecithotrophic phase, imparted a specificity in PUFA composition to the different classes of polar lipids. During this phase, 20:5(n 3) replaced 20:4(n 6) in PI but decreased sharply in PLSM and, to a lesser extent, in PC. After the initial phase of composition changes, the fatty acid composition of the polar lipid classes became stable, with specific associations of some of the PUFA with certain polar lipid classes: 22:6(n 3) with GLY, 20:5(n 3) with PE, 20:4(n6) with PI. The 22:6(n3):20:5(n3) ratio in PLSM during exotrophy was always twice as high as in the PC fraction.

Effects of algal diet on digestive enzyme activity in Calanus helgolandicus

Adult female Calanus helgolandicus were transferred, immediately after collection in the English Channel in June 1984, to two unialgal diets one of which, the flagellate Cryptomonas maculata, was rich in starch, while the other, the diatom Thalassiosira weissflogii, contained no starch. The activity of the digestive enzymes amylase, trypsin and laminarinase was measured in these two populations under foodsaturating conditions over an acclimation period of eight days. Ingestion rates were measured on a daily basis and the results confirmed, together with a constant level of body protein, that the experimental conditions were above the incipient limiting concentration. In the long-term (4 to 8 d), the activities of all three enzymes were significantly elevated in the C. maculata-fed copepods, whereas ingestion rates were lower than those on T. weissflogii. These results observed under food-saturating conditions indicate a compensatory mechanism between digestive enzymes and the substrate ingested. They are consistent with previous work on Artemia sp. suggesting that the rate of assimilation is a function of the digestive enzyme activity and ingestion rate. Enzyme activity exhibited differing shortterm responses (<48 h) on transfer to the two algal diets, which are interpreted in relation to the in situ activity of the field population.

Effects of Alexandrium minutum exposure upon physiological and hematological variables of diploid and triploid oysters, Crassostrea gigas.

The effects of an artificial bloom of the toxin-producing dinoflagellate, Alexandrium minutum, upon physiological parameters of the Pacific oyster, Crassostrea gigas, were assessed. Diploid and triploid oysters were exposed to cultured A. minutum and compared to control diploid and triploid oysters fed T. Isochrysis. Experiments were repeated twice, in April and mid-May 2007, to investigate effects of maturation stage on oyster responses to A. minutum exposure. Oyster maturation stage, Paralytic Shellfish Toxin (PST) accumulation, as well as several digestive gland and hematological variables, were assessed at the ends of the exposures. In both experiments, triploid oysters accumulated more PSTs (approximately twice) than diploid oysters. Significant differences, in terms of phenoloxidase activity (PO) and reactive oxygen species (ROS) production of hemocytes, were observed between A. minutum-exposed and non-exposed oysters. PO in hemocytes was lower in oysters exposed to A. minutum than in control oysters in an early maturation stage (diploids and triploids in April experiment and triploids in May experiment), but this contrast was reversed in ripe oysters (diploids in May experiment). In the April experiment, granulocytes of oysters exposed to A. minutum produced more ROS than those of control oysters; however, in the May experiment, ROS production of granulocytes was lower in A. minutum-exposed oysters. Moreover, significant decreases in free fatty acid, monoacylglycerol, and diacylglycerol contents in digestive glands of oysters exposed to A. minutum were observed. Concurrently, the ratio of reserve lipids (triacylglycerol, ether glycerides and sterol esters) to structural lipids (sterols) decreased upon A. minutum exposure in both experiments. Also, several physiological responses to A. minutum exposure appeared to be modulated by maturation stage as well as ploidy of the oysters.