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00000; International audience; Haptophytes are a diverse monophyletic group with a worldwide distribution, known to be significantly involved in global climate regulation in their role as a carbon sink. Because nitrogen is a major limiting macronutrient for phytoplankton in oceans and for cultures of microalgae, understanding the involvement of nitrogen availability in haptophyte carbon partitioning is of global and biotechnological importance. Here, we made an ecophysiological study coupled with comprehensive large scale proteomic analysis to examine differences of behavior in reaction to nitrogen availability changes between a wild type strain of Tisochrysis lutea (WTc1) and a mutant strain (2Xc1) known to accumulate more storage lipids. Strains were grown in chemostats and studied under different ecophysiological conditions including N limitation, N repletion and N depletion. Whereas short time N repletion triggered consumption of carbohydrates in both strains, storage lipid degradation and accumulation during changes of ecophysiological status were recorded in 2Xc1 but not in WTc1. After 3 months of continuous culture, 2Xc1 exhibited an unexpected increase of carbon sequestration ability (+ 50%) by producing twofold more carbohydrates for the same nitrogen availability. Deep proteomic analysis by LC-MS/MS identified and compared the abundance of 4332 proteins, i.e. the deepest coverage of a microalgal proteome obtained to date. Results revealed that storage lipid accumulation is favored by an overall reorganization of carbon partitioning in 2Xc1 cells that increases the metabolism of carbon and energy acquisition, and decreases mitochondrial activity and metabolic conversion of storage lipids to phosphoenolpyruvate before gluconeogenesis.

The need to replace petroleum fuels with alternatives from renewable and more environmentally sustainable sources is of growing importance. Biomass-derived biofuels have gained considerable attention in this regard, however first generation biofuels from edible crops like corn ethanol or soybean biodiesel have generally fallen out of favor. There is thus great interest in the development of methods for the production of liquid fuels from domestic and superior non-edible sources. Here we describe a detailed procedure for the production of a purified biodiesel from the marine microalgae Isochrysis. Additionally, a unique suite of lipids known as polyunsaturated long-chain alkenones are isolated in parallel as potentially valuable coproducts to offset the cost of biodiesel production. Multi-kilogram quantities of Isochrysis are purchased from two commercial sources, one as a wet paste (80% water) that is first dried prior to processing, and the other a dry milled powder (95% dry). Lipids are extracted with hexanes in a Soxhlet apparatus to produce an algal oil ("hexane algal oil") containing both traditional fats (i.e., triglycerides, 46-60% w/w) and alkenones (16-25% w/w). Saponification of the triglycerides in the algal oil allows for separation of the resulting free fatty acids (FFAs) from alkenone-containing neutral lipids. FFAs are then converted to biodiesel (i.e., fatty acid methyl esters, FAMEs) by acid-catalyzed esterification while alkenones are isolated and purified from the neutral lipids by crystallization. We demonstrate that biodiesel from both commercial Isochrysis biomasses have similar but not identical FAME profiles, characterized by elevated polyunsaturated fatty acid contents (approximately 40% w/w). Yields of biodiesel were consistently higher when starting from the Isochrysis wet paste (12% w/w vs. 7% w/w), which can be traced to lower amounts of hexane algal oil obtained from the powdered Isochrysis product.

The need to replace petroleum fuels with alternatives from renewable and more environmentally sustainable sources is of growing importance. Biomass-derived biofuels have gained considerable attention in this regard, however first generation biofuels from edible crops like corn ethanol or soybean biodiesel have generally fallen out of favor. There is thus great interest in the development of methods for the production of liquid fuels from domestic and superior non-edible sources. Here we describe a detailed procedure for the production of a purified biodiesel from the marine microalgae Isochrysis. Additionally, a unique suite of lipids known as polyunsaturated long-chain alkenones are isolated in parallel as potentially valuable coproducts to offset the cost of biodiesel production. Multi-kilogram quantities of Isochrysis are purchased from two commercial sources, one as a wet paste (80% water) that is first dried prior to processing, and the other a dry milled powder (95% dry). Lipids are extracted with hexanes in a Soxhlet apparatus to produce an algal oil ("hexane algal oil") containing both traditional fats (i.e., triglycerides, 46-60% w/w) and alkenones (16-25% w/w). Saponification of the triglycerides in the algal oil allows for separation of the resulting free fatty acids (FFAs) from alkenone-containing neutral lipids. FFAs are then converted to biodiesel (i.e., fatty acid methyl esters, FAMEs) by acid-catalyzed esterification while alkenones are isolated and purified from the neutral lipids by crystallization. We demonstrate that biodiesel from both commercial Isochrysis biomasses have similar but not identical FAME profiles, characterized by elevated polyunsaturated fatty acid contents (approximately 40% w/w). Yields of biodiesel were consistently higher when starting from the Isochrysis wet paste (12% w/w vs. 7% w/w), which can be traced to lower amounts of hexane algal oil obtained from the powdered Isochrysis product.

© 2016. Two microalgal species, Isochrysis galbana and Nannochloropsis oculata, were supplemented to the diet of laying hens, in either non-disrupted or disrupted (by high pressure homogenization) form, to examine the effect of the presence of a cell wall on the omega-3 long chain polyunsaturated fatty acid (n. -. 3 LC-PUFA) enrichment in the egg yolk. Only supplementation of Nannochloropsis led to significant differences in enrichment by supplementation of the two forms, with the highest enrichment obtained with the HPH-disrupted form. This confirms the presence of a rigid cell wall for this species, which reduces the bioaccessibility of the n. -. 3 PUFA. The needed conversion reaction from EPA to DHA on the other hand did not seem to effect the enrichment, since the same n. -. 3 LC-PUFA enrichment was obtained with the supplementation of the DHA source Isochrysis and the EPA source Nannochloropsis, the last in HPH-disrupted form.Next to the microalgal species, fish oil was also supplemented to the diet of the laying hens. Fish oil resulted in a higher enrichment in comparison with the microalgal species, either non-disrupted or HPH-disrupted. This might suggest a difference in bioaccessibility of the n. -. 3 PUFA when added as oil or biomass. publisher: Elsevier articletitle: The cell wall of autotrophic microalgae influences the enrichment of long chain omega-3 fatty acids in the egg journaltitle: Algal Research articlelink: http://dx.doi.org/10.1016/j.algal.2016.03.015 content_type: article copyright: © 2016 Published by Elsevier B.V. ispartof: Algal Research vol:16 pages:209-215 status: published

Filter-feeding bivalves play an important ecological role in aquatic systems through benthic–pelagic coupling. However, the mechanisms by which invasive mussels influence the food uptake by local consumers are little known. We tested the interactive effects of the invasive mussel Arcuatula senhousia and the native clam Ruditapes decussatus on the uptake of 13C-enriched microalgae, Isochrysis sp., by two polychaetes, the surface deposit feeder Cirriformia tentaculata and the sub-surface deposit feeder Phylo foetida. Polychaetes were exposed to two different densities of A. senhousia and R. decussatus used alone or together. In each treatment, the 13C-concentration throughout the sediment horizon (0–0.5, 0.5–2.5 and 2.5–5 cm) was also quantified. Results showed that, irrespective of bivalve densities, the 13C-uptake by C. tentaculata was lower in the presence of the two bivalves together than in the presence of one bivalve species (either A. senhousia or R. decussatus). This was consistent with decreased 13C-concentrations in the 0–0.5 cm sediment layer in the treatment with the two bivalves together. Contrary to C. tentaculata, no changes were observed among treatments in the 13C-uptake by P. foetida. This possibly reflected a limited 13C-enrichment of the lower sediment layers. Our main findings are: (1) A. senhousia reduces the 13C-uptake by R. decussatus, and (2) A. senhousia reduces the availability of phytoplankton-derived C for deposit feeders by impairing the feeding, incorporation and bio-deposition of R. decussatus. We conclude that the interaction of A. senhousia with native bivalves needs to be taken into consideration in future food web studies.

The surf Spisula solida and the striped venus Chamelea gallina clams support important fisheries in Europe. These fisheries have been affected by inter-annual fluctuations in stock abundance and recruitment failures. Aquaculture could contribute to address these problems through the production of larvae or juveniles for stock-enhancement programmes. In the present study, the prerequisites to produce these species in hatchery were established. Of the different methods used to obtain gametes (thermal stimulation, scarification and flow through), C. gallina spawned in the flow through only. By its turn, the best method for obtaining gametes for S. solida was stripping. Higher survivals of D-larvae were found at 15 and 17 degrees C. The greatest larval survival and growth rates were obtained from larvae fed Isochrysis aff. galbana (T-iso). First C. gallina and S. solida pediveliger larvae were found at day 8 and day 26 respectively. The presence of metamorphosed larvae found even in the unfed treatment for C. gallina and for S. solida is an advantage for aquaculture production. After metamorphosis, the S. solida juveniles' survival was quite low, whereas there was no mortality for C. gallina juveniles reared without substrate regardless the diet provided. Juveniles fed the binary diet (Chaetoceros calcitrans - C. cal + T.iso) and the monodiet T-iso showed relatively higher growth in shell length and weight than C. cal. The hatchery production of these two species was possible, however the results of this study showed that C. gallina can be more attractive for aquaculture than S. solida.

Four strains of marine microalgae commonly used as live feeds in hatcheries (Isochrysis sp. T.ISO, Tetraselmis suecica, Phaeodactylum tricornutum, Nannochloropsis sp.) were grown in a novel solid-state photobioreactor, the twin-layer system. Microalgae were immobilized by self adhesion to vertically oriented twin-layer modules which consisted of two different types of ultrathin layers, a macroporous source layer (glass fiber nonwoven) through which the culture medium was transported by gravity flow, and a microporous substrate layer (plain printing paper) which carried the algae on both surfaces of the source layer. This simple open cultivation system effectively separated the immobilized microalgae from the bulk of the growth medium and permitted prolonged cultivation of microalgae with average biomass yields of 10–15 g dry weight m−2 growth area after 14–25 days of cultivation. Algal biomass was harvested as fresh weight (with 72–84 % water content) without the need to pre-concentrate algae. No aeration or external CO2 supply was necessary, and due to the microporous substrate layer, no eukaryotic contaminations were observed during the experiment. All experiments were conducted in Germany under greenhouse conditions with natural sunlight. Small-scale growth experiments performed under the same conditions revealed that growth over most of the experimental period (24 days) was linear in all tested algae with growth rates (dry weight per square meter growth area) determined to be 0.6 g m−2 day−1 (Isochrysis), 0.8 g m−2 day−1 (Nannochloropsis), 1.5 g m−2 day−1 (Tetraselmis), and 1.8 g m−2 day−1 (Phaeodactylum). Due to its cost-effective construction and with further optimisation of design and productivity at technical scales, the twin-layer system may provide an attractive alternative to methods traditionally used to cultivate live microalgae.

The effect of three different nutrient depletions (nitrogen, sulphur and magnesium) on the growth and pigment accumulation of the haptophyte Isochrysis aff. galbana (clone T-ISO) has been studied. Pigments were quantified based on RP-UHPLC-PDA-MSn analysis. All nutrient depletions led to reduced maximal biomass concentrations. Besides, all nutrient-depleted cultures accumulated 3-hydroxyechinenone. To our knowledge, this is the first time that 3-hydroxyechinenone has been found in I. aff. galbana T-ISO. Most 3-hydroxyechinenone, as well as the most echinenone and diatoxanthin, were found in the nitrogen-limited culture in which a more severe limitation resulted in higher cellular contents. Similar to accumulation of diatoxanthin, accumulation of 3-hydroxyechinenone and echinenone may be part of a global (stress) response mechanism to oversaturating light conditions.

The effect of egg density on embryonic development and larval quality as well as the lipid and fatty acid contents (eicosapentaenoic acid, EPA; docosahexaenoic acid, DHA) of cultured microalgae fed to Perna perna larvae was studied under controlled conditions to provide information needed for development of an experimental hatchery. Embryonic development followed the common sequence exhibited by other bivalves. d-larva stage was attained 40–44 h post-fertilisation at 21 ± 1 °C. The umbo-stage was reached in 11 days, and pediveliger larvae were observed 26 days post-fertilisation. Low egg density (range 20–100 eggs cm−2) produced high proportions of normal d-larvae. Larval development showed two growth phases: 1st—the mixotrophic stage and 2nd—the exotrophic stage where the composition of diets had significant effects on larval growth with higher rates in larvae fed with the mixed microalgae (Isochrysis galbana + Chaetoceros calcitrans, I. galbana + Phaeodactylum tricornutum and I. galbana + Skeletonema costatum) in comparison with the monospecific diet (I. galbana). Fatty acid analysis showed that larval growth and survival were strongly influenced by proportions of dietary DHA and EPA. These results indicate that DHA and EPA are the key factors in determining larval performance, considerably more than the total amount of other fatty acids.

This study compared the efficacy of four products that are commonly used in hatchery for nutritional enhancement of rotifer Brachionus plicatilis as the starter food for yellowtail kingfish Seriola lalandi larvae. This experiment consisted of one fresh algae and three enrichment products: (1) Fresh algae were a mixture of Nannochloropsis and Isochrysis at 2:1 on a cell concentration basis; (2) S.presso, (Selco S.presso ® , INVE Aquaculture); (3) Algamac 3050 ® (Aquafauna, USA); (4) Nutrokol ® (NutraKol, Australia). Survival rates of the fish fed rotifers enriched with fresh microalgae (40.69%) and S.presso (31.21%) were higher than those fed Algamac 3050 (10.31%). On 3 day post hatch (DPH), fish feeding incidence in the fresh algae treatment was significantly higher than that in other treatments. On 6 DPH, fish showed the lowest feeding incidence in the Algamac 3050 treatment. The methods of enrichment did not affect total lipid levels in either rotifer or fish larvae, but Algamac 3050 enrichment achieved the highest DHA/EPA ratio and lowest EPA/ARA ratio in both rotifers and fish larvae. This study indicates that fresh algae can be replaced by S.presso, but Algamac 3050 is not as good as other formula for rotifer enrichment in rearing yellowtail kingfish larvae in this system.

Microalga, Isochrysis galbana, biomass was entrapped into alginate gel by liquid curing method in the presence of Ca(II) ions. The biosorption of chromium(III) by the entrapped live algal biomass was studied in a batch system. The effect of initial cadmium concentration, pH, temperature and liquid and solid ratio on Cr(III) removal was investigated. The maximum experimental biosorption capacities for entrapped live algal biomass were found to be 335.27 mg Cr(III) g−1 of dry algal biomass. The kinetics of chromium biosorption was slow; approximately 75 % of biosorption took place in 2 h. The percent adsorption increased with increase in pH; pH 5 of the solution was found to favor adsorption very strongly. The equilibrium biosorption data were evaluated by Langmuir and Freundlich isotherm models, and was best described by Langmuir and Freundlich isotherms. The biosorbent was characterized and evaluated, the functional groups –OH, –COOH and C=O were involved in the biosorption process. Since binding capacity was relatively high for immobilized live algal biomass, those algal forms are to be considered as suitable biosorbent for the removal of chromium in wastewater treatment.

Even though the microalgal species of Isochrysis and Pavlova are widely used as live food in bivalve hatcheries, they are difficult to culture in mass during the summer season. Therefore, the present study was conducted to determine the optimum species or strains of Isochrysis and Pavlova to produce good growth and high contents of fatty acids at temperatures over . Four species of Isochrysis (I. galbana KMMCC12, I. galbana KMMCC214, I. aff. galbana, and Isochrysis sp.) and four of Pavlova (P. lutheri, P. gyrans, P. viridis, and Pavlova sp.) were cultured at , , and , and then analyzed for specific growth rate and fatty acid composition. Microalgae were cultured in f/2 medium at 23 psu and continuous light of photons . For the I. galbana, growth rates were highest at and decreased at to the level observed at . I. galbana (KMMCC12) and Isochrysis sp. cultured at and , respectively, exhibited the highest growth rates of all Isochrysis species. In terms of fatty acids, I. galbana (KMMCC12) contained higher contents of PUFA and n-3 HUFA at than did Isochrysis sp. For species of Pavlova, growth rates of P. gyrans and P. viridis at and , respectively, were higher than those of the other Pavlova species. In particular, P. viridis grew as well at as it did at . However, P. lutheri and Pavlova sp. did not grow at . In terms of fatty acids, P. viridis cultured at also exhibited higher contents of PUFA and n-3 HUFA, as compared to P. gyrans. Based on these results, we suggest that I. galbana (KMMCC12) and P. viridis are suitable species for mass culture during the high temperature season.

Fucoxanthin, a main marine carotenoid, in five species of fucoxanthin-containing microalgae, was quantified by high-performance liquid chromatography. Among the studied species, Isochrysis aff. galbana contained the highest amount of fucoxanthin (18.23 mg/g dried sample). This microalga showed good fucoxanthin extraction efficiency under the tested solvents (methanol, ethanol, acetone, and ethyl acetate), with the exception of n-hexane. In addition, most fucoxanthin (∼95%) could be extracted by a single extraction in ethanol within 5 min, and only 15% degradation of fucoxanthin was detected during ethanol extraction for 24 h. The two-phase solvent system of n-hexaneethanol-water with a volume ratio of 10:9:1 was determined to be the best system for the separation of fucoxanthin and lipids from extracts of I. aff. galbana. Under these conditions, fucoxanthin was fractionated in the hydroalcohol phase apart from the hexane phase containing lipids. These results imply that I. aff. galbana can be a commercial source for the spontaneous production of valuable fucoxanthins and lipids.

Microalgae concentrates (paste) can be used as an alternative feed to replace live microalgae for aquaculture due to its nutritional value and convenience. However, the clumping of cells and negative buoyancy of algae concentrate can affect bivalve culture as bivalves only capture particles in suspension and ingest a certain size range of particles. This study investigated the effect of shaking and blending treatments on the preparation of food suspensions prepared from algae concentrates (Isochrysis and Pavlova). The results indicated that the higher the shaking time (5, 10, and 15 times) or blending time (10, 30, and 60 seconds), the smaller was the diameter of the resulting algae particles. Moreover, the greater the volume of algae concentrate used in preparation, the larger the diameter of algae particles produced. Shaking may be the best option because it is cheaper and simpler. However, all the treatments provided a suitable particle size range for ingestion by bivalves.

This study tested the diet enrichment with fatty acid EPA and DHA during the broodstock conditioning of scallops Nodipecten nodosus in hatchery. For this, before the conditioning the animals were induced to spawning and then were placed in tanks for maturation of 30 days, and subjected to five treatments: (1) Cm100%, (2) Cm + DHA, (3) T-Iso100%, (4) T-Iso + EPA e (5) Mix (mix of the microalgae Isochrysis sp., Chaetoceros muelleri and Skeletonema sp. , in the rate 50:25:25, respectively). After the conditioning, the animals were induced to spawning. The sexual stage by histology, spawning, larviculture and settlement was evaluated. The treatment T-Iso + EPA in broodstock showed 100% of the animals mature, stage 2B (advanced pre-spawning), higher yield in larvae D, pediveliger and spat. These results indicate that broodstock feeding with T-Iso + EPA provides a positive effect on the maturation, as well as a higher larvae and spat survival in hatchery.

We investigated the photoresponses of diadinoxanthin (DD) and diatoxanthin (DT) using Isochrysis gal- bana cultures that had been acclimated to four different levels of photon flux density (PFD). DD can play a role as a photosynthetic light-harvesting pigment, whereas DT can play a role as a photoprotective pigment. The PFD ranged from limited light conditions (LL) to saturated light conditions (SL) for growth. The photoresponses of these pig- ments during light-dark transitions at each acclimation PFD were consistent. The photoresponses of these xantho- phyll pigments, as normalized to either cell density or chlorophyll a (Chl a) concentration, to light-dark transitions varied with acclimation PFD. The stoichiometric variations between the DD or DT : Chl a ratio observed at the two higher acclimation PFDs suggested that these xanthophyll dynamics are controlled by xanthophyll cycling processes in addition to de novo processes and interconversion between DD and fucoxnathin under SL conditions. Significant linear relationships were observed between the levels of DD and/or DT normalized to either the cell density or Chl a concentration. Based on the cellular pigment content and the xanthophyll : Chl a ratio, the slopes of the linear rela- tionships normalized to either the cell density or Chl a concentration were significantly related to the acclimation PFD linearly and sigmoidally, respectively. Similarly, the acclimation PFD at the interaction of the sigmoidal change and the transition point from the LL to SL condition suggests that these xanthophyll pigment dynamics can be char- acterized by acclimation to the light conditions. An examination of the slope may provide information leading to a better understanding of photoacclimation in the algal photosynthetic apparatus.

Algal allelopathy is an ecological or physiological phenomenon that has focused attention on the interactions among algae and the production of algal toxins.A species of marine bloom-forming microalga: Amphidinium carterae was selected in the present study to estimate its potentially allelopathic effects on other species of microalgae.The cell-free filtrates of A.carterae in two growth phases: exponential grwoth phase(abbreviated as growth phase Ⅰ) and stationary growth phase(abbreviated as growth phase Ⅱ) were prepared,and its inhibitory effects on growth of other species of marine microalgae were studied under stable environmental conditions(temperature,light and illuminating periodicity,etc.).The cell-free culture filtrates were prepared firstly when A.carterae were cultured phaseⅠand phaseⅡ,which was about the the growth 7th and 17th respectively,and enriched with f/2 medium.The above prepared culture filtrates were used to culture 4 species of marine microalgae: Skeletonema costatum、Prorocentrum micans、Scrippsiella trochoidea and Isochrysis golbana.Moreover,the crude extractions in the cell-free filtrate of A.carterae of growth phaseⅡ were extracted by GC/MS,and the major components were analyzed.Results showed that: in the enriched cell-free filtrates,S.costatum were obviously inhibited in either filtrates of phase I(F=15.18475,P=0.002980.05) or phase Ⅱ(F=6.24559,P=0.031490.05) as compared to that in the control which in f/2 enriched medium,but the cell numbers of S.trochoidea were only about 79.3% and 68.9% in filtrates prepared from phaseⅠand phaseⅡ as compared to the control by the end of experiment.As to P.micans,its growth was obviously suppressed(F=4.84438,P=0.049250.05) in the filtrate prepared from phaseⅡduring the last 3 days.Little inhibition was observed in I.golbana in either filtrates prepared from either phases(P0.05).Cell-free filtrate cultures experiment of A.carterae is one of the basic approaches on allelopathy research.Results in the present study suggested that A.carterae secreted or released some substances into the culture medium,which inhibited the co-cultured S.costatum and P.micans,and these inhibitory effects could not caused by light,nutrients or cell contact to gain a competitive advantage.Many algae could release allelochemicals into the environment which may alter phytoplankton growth to enhance its predominance.Besides,growth of these two species was more inhibited in enriched cell-free filtrate prespred from growth phaseⅡ than that from phaseⅠ.It also proved that the allelochemical effects on algae were species-specific because growth of S.costatum and P.micans was inhibited,but exerted little effects on S.trochoidea and I.golbana.Another experiment was performed to further prove the allelopathic effect of A.carterae.The crude extractions from cell-free filtrates of A.carterae at different concentrations were added into S.costatum and P.micans culture media,found that cells dissolved in higher concentration after many hours.It is noted that the crude extractions had the function of dissolving some algal cells.Then the crude extractions were analyzed by GC/MS.Four components in the crude exactions were suspected to have the allelopathic effects,they were Butylated Hydroxytoluene,Butyl citrate,Benzene,1-Chlorodifluoromethoxy-4-nitro-,and Benzyl butyl phthalate,among with Butylated Hydroxytoluene(BHT) was reported with oxidative characteristic and was used as an antioxidant additive.Results in the present study brought light on the hypothesis that A.carterae could realese allelochemical(s);however,deep research should carry out to explain the possible inhibiting mechanism.

The biomass, proximal composition and fatty acid profile of Isochrysis sp., Chaetoceros muelleri and their mixture, grown under greenhouse conditions, were evaluated. The nutritional value of both species supplied as the monoalgal (Chaetoceros muelleri: Diet I, and Isochrysis sp. Diet II) and mixed diet (Diet III) for larval Litopenaeus vannamei was also assessed on the basis of the development and biochemical composition of the larvae. The highest protein levels were obtained in Diets I and II (40% and 35%, respectively). No significant differences in larval survival were found among the diets; however, larvae fed on Diet II had the lowest mean larval length.

The effect of algal species on the life history of the euryhaline cladoceran, Daphniopsis australis was investigated under laboratory conditions. Three algal species, Tetraselmis suecica (T), Nannochloropsis oculata (N) and Isochrysis tahitian (I) were used to form three mono-alga species and four multi-algae species (T × N, I × N, I × T and I × T × N) to feed D. australis . The type of algal species significantly affected the life history of D. australis at both individual and population levels. The mono-alga T. suecica enhanced animal longevity and reproductive output but hampered animal body growth. In contrast, the mono-alga N. oculata and I. tahitian resulted in the highest growth rate of D. australis while the multi-algae I × N reduced the age at first reproduction with a possible cause of high carbon contents in the N. oculata and I. tahitian diets. The mono-alga I. tahitian led to the shortest longevity and poorest reproduction output. Egg development was the fastest when animals fed on mono-algal species but was delayed by feeding animals on multi-algal species. The animals fed on the mono-algal species T. suecica showed the highest net reproduction ( Ro ) and the longest generation time ( GT ) while those fed on the mono-algal diet I. tahitian exhibited the lowest Ro and the shortest GT . The intrinsic growth rate was the highest when the animals fed either on the mono-alga T. suecica or on the multi-algae I × N. This study indicates that species composition of algae in the diet can affect development, growth and reproduction of D. australis and thereby influence the population dynamic of cladocerans living in heterogeneous food environments.

This study assessed the nutritional value of four species of microalgae for larval sandfish, Holothuria scabra (Echinodermata: Holothuroidea). Two-day-old larvae were fed either a single diatom (Chaetoceros muelleri, Chaetoceros calcitrans), flagellate (Isochrysis aff. galbana (T-ISO), Pavlova salina), or a ternary microalgal diet (TMD) comprised of 40% C. muelleri, 40% T-ISO, and 20% P. salina for 12 d. On Days 4, 6, and 8 length of larvae fed either C. muelleri, C. calcitrans, or the TMD was significantly greater than in T-ISO and P. salina fed larvae as well as the unfed control larvae. T-ISO fed larvae had a significantly greater length compared to unfed larvae on Days 4, 6, and 8; however, larvae fed P. salina had a significantly greater length compared to unfed larvae on Days 6 and 8 only. Survival of larvae fed either C. muelleri or the TMD was significantly better than the flagellate fed and unfed larvae on Days 4, 8, and 12. There was no significant difference between survival of unfed larvae and larvae fed either T-ISO or P. salina during the 12-d larval culture period. On Day 12, larvae fed either a diatom or the TMD developed in excess of 84% competent doliolariae. Larvae fed C. muelleri had a significantly higher percentage competent doliolariae than larvae from all other treatments except the TMD. Neither larvae fed either T-ISO or P. salina nor unfed larvae developed any competent doliolariae. In conclusion, this study showed C. muelleri to be the most effective of the four microalgae tested as a single microalgal diet for larval H. scabra and, as such, C. muelleri will be used as a reference diet in future studies.

As an alternative biomass for producing biofuel, microalgae have many advantages over land biomass. For example, they are not a food source, they are fast-growing and have a high oil content, and they can reduce CO2. However, the high costs of cultivation and harvesting using existing technology make microalgal-based biofuels uncompetitive with fossil fuels. This study investigates the feasibility of producing high-value microalgal pigments, including chlorophyll and carotenoid for apportioning the high cost of cultivation and harvest. Five microalgae were tested to determine their pigment and lipid production in a feed-batch bioreactor under light conditions of 167 μmol photon/(m2 s) for 12 h daily. Among them, Dunaliella primolecta and Isochrysis aff. galbana achieved the highest chlorophyll yields of 5.3% and 5.6%, respectively, with the optimal carotenoid yields of 2.6% and 2.0%, and lipids yields of 23% and 24%, respectively. The feasibility of simultaneously extracting both lipids and pigments was investigated. Two solvents, methanol and hexane, were used in the extraction of pigments and lipids. The extraction yields of pigments and lipids using methanol and hexane reached 98% and 90%, respectively. Therefore, we expect that the simultaneous production and extraction of high-value pigments accompanied by the lipids can potentially offset the high cultivation and harvesting costs, and may make microalgal biomass economically competitive for biofuel production.

© 2018 Microalgal lipids have a huge potential in several applications such as biofuels, food and feed. However, enzymatic lipolytic reactions during wet storage cause the formation of substantial amounts of free fatty acids (FFA), which give several problems for these applications. In this study, a full factorial (32) design with varying time and temperature was used to optimize a heat treatment, that inactivates lipolytic enzymes in the wet paste of T-Isochrysis lutea. Therefore, the FFA and lipid content were evaluated at three time points (0, 3 and 7 days) during wet storage at 4 °C. A treatment of at least 8 min at 80 °C was found to inactivate the lipolytic enzymes. A second experiment elucidated that this optimal treatment had no negative impact on fatty acid composition, peroxide value, phenolic content or carotenoid content. A short heat treatment, thus, enables wet storage of microalgal paste to several days at 4 °C. ispartof: ALGAL RESEARCH-BIOMASS BIOFUELS AND BIOPRODUCTS vol:38 status: published

Filtration rate and ingestion rate of different stages of Paphia malabarica larvae (D-shape, (80 μm), Umbo (120 μm) and veliger (180 μm)) were determined in relation to feeding on various micro algae. The micro algae tested were Nannochloropsis salina, Isochrysis aff. galbana, Dicrateria inornata, Tetraselmis gracilis and Chaetoceros calcitrans at 5 or 10 × 103 cells mL−1. Both filtration and ingestion rate of micro algae tested were increased with increasing larval size; however, at all larval stages, C. calcitrans resulted in lower filtration and ingestion rate. Of the algal diets tested, P. malabarica larvae showed greatest filtration rate and ingestion rate with N. salina. Maximum filtration rate for N. salina was 15.7, 26.3 and 33.9 μL h−1 and highest ingest rate was 15, 92 and 177 cells larva−1 h−1 in D shape larvae, umbo and pediveliger, respectively. Filtration rate and ingestion rate of N. salina were always higher than those other algal species tested because of its small cell size (2 μm).

This paper deals with the application of electrolysis technology for the selective control of marine algal species. The sensitivity of different marine algal species to electrolysis may be different due to their size and structural differences. Electrolysis experiments were conducted using four algal species, namely Chlorella spp., Chaetoceros spp., Isochrysis spp. and Tetraselmis spp. Batch and continuous runs were conducted from 0-7 minutes and 0-80 minutes, respectively. Drastic depletion of algal cell number was not observed during batch electrolysis. However, fluorescein diacetate and propidium iodide staining showed significant algal mortality. Different sensitivities were also observed for similar electrolysis conditions. Continuous electrolysis data revealed almost no survival for Chlorella spp. at 80 minute electrolysis time while survival for Isochrysis spp. was around 60% at similar conditions. Results indicated that the green algae Chlorella spp. is the most sensitive while the brown algae Isochrysis spp. is the most resistant among the four species studied. More importantly, this confirmed that the beneficial marine algae Isochrysis spp. can be allowed to dominate by selectively limiting the growth of three other algal species tested in this work through electrolysis treatment. DOI: 10.4038/sljas.v11i0.2221 Sri Lanka J. Aquat. Sci. 11 (2006): 27-42

To determine variability of marine microalgal lipids during cell growth and respiration, and within intracellular structures, we conducted two series of experiments: (1) batch culturing Isochrysis sp., Gymnodinium sp., Platymonas subcordiforus , Heterosigma akashiwo Hada, and Skeletonema costatum with lipid analysis for cell samples collected in exponential growth and stationary phase respectively; and (2) batch culturing Isochrysis sp. and Gymnodinium sp. from exponential growth to stationary phase and followed by dark incubation with progressive lipid analysis. Moreover, membrane and intracellular storage fatty acids were separated using Si-gel chromatography from representative samples of each species in the second experiment (respectively in exponential growth, stationary phase, and dark respiration). Results from the first experiment showed that no matter how differently microalgae grew, their lipid yields were 4–38 × higher in stationary phase than in exponential growth phase. Progressive lipid analysis in the second experiment indicated that total fatty acid (FA) yields in two microalgal cultures continuously increased throughout exponential and stationary phases but total neutral lipid (NL) yields slowed or stopped to increases in stationary phase. During dark incubations, total FA yields decreased steadily in both Isochrysis (with constant cell density) and Gymnodinium (with declining cell density) cultures but total NL yields decreased only in Gymnodinium culture. Lipid compositions and proportions of membrane vs. intracellular fatty acids varied differently with growth phases and respiration in two cultures. This study suggests that the effect of cell growth phase on cellular lipid yield and composition is species-dependent and is likely related to the capacity of cells to synthesize membrane vs. intracellular storage lipids.

This study analyzed the effect of water quality, different diets, and water temperature regimes in the conditioning and maturation of the Pacific oyster Crassostrea gigas. A recirculating aquaculture system prototype was used in this experiment and 2 different trials were performed. During the first trial, the effects of 3 different diets using different preserved concentrates of microalgae held at 20°C were tested: D1 (fed with Tetraselmis 3600), D2 (fed with Isochrysis 1800), and D3 (fed with a 50:50 mix of Tetraselmis and Isochrysis). During the second trial, 3 different temperatures regimes—T1, 16–24°C; T2, 20°C; and T3, 24°C (fed with preserved concentrates of microalgae at a 50:50 Tetraselmis and Isochrysis food ratio)—were evaluated. For each one of these treatments, 43 organisms were used and every treatment was done in triplicate. Seven samplings were completed in 72 days. During each sampling period, 9 oysters from each treatment were collected, and a gonad sample was obtained for histo...

The diatom Chaetoceros calcitrans forma pumilum is an important dietary component for cultured Greenshell™ mussel larvae, Perna canaliculus. However concerns surrounding potentially deleterious side effects have motivated a series of experimental trials to optimise the safe usage of this alga. Bioassays involved raising veliger larvae in the purpose-built Cawthron Ultra-Density Larval rearing (CUDL) system; an array of 2.5-L tanks were stocked with 2-day-old veligers (200 larvae mL−1) supplied with inflowing water dosed with sufficient microalgae to maintain a fixed concentration of cells, after compensating for ingestion. The nutritional role of C. ‘calcitrans’ was examined by adjusting its cellular fraction in the feed environment. Diets of 0, 5, 66, 95 and 100% C. ‘calcitrans’ were each offered to six replicate rearing tanks, using Isochrysis aff. galbana (T-Iso clone) to maintain a total of 40 cells μL−1 in the larval cultures. The 66% C. ‘calcitrans’ diet was also offered at 3, 20, 60 and 120 cells µL−1 (n = 6). Higher C. ‘calcitrans’ fractions sustained faster growth, with 95% and 100% treatments producing 23-day-old pediveligers of 240 ± 15 μm and 228 ± 5 μm mean shell length, respectively; compared to 212 ± 7 μm in the 66% treatment (40 cells μL−1). However, high C. ‘calcitrans’ treatments were more volatile, had higher mortality and greater predisposition towards population crashes. Similarly, reduced survival was observed in the high feed treatment of 120 cells µL−1, reflected in a final pediveliger yield of 29 ± 6%, compared to 47 ± 3% at 60 cells µL−1 and 36 ± 7% at 40 cells µL−1. The 5% C. ‘calcitrans’ and 20 cells µL−1 treatments showed signs of nutrient limitation, while larvae fed 0% C. ‘calcitrans’ or a total of 3 cells µL−1 starved, failing to reach metamorphosis. A standard diet consisting of 66% C. ‘calcitrans’ and 34% I. aff. galbana maintained at 40 cells μL−1 was used to test the hypothesis that culture age increased deleterious side effects associated with feeding C. ‘calcitrans’. The final stage of batch culture, using 20-L nylon carboys inoculated with 1010 C. ‘calcitrans’ cells was allowed to age for 2, 3, 4, 5 or 6 days before being fed to larvae. After 21 days eating 2-day-old C. ‘calcitrans’ larvae reached a mean shell length of 236 μm, were eating 35,000 cells larva−1 day−1, and had a survival rate of 59 ± 3%; in contrast, larvae eating 6-day-old cells only reached 214 µm, eating 17,000 cells larva−1 day−1, with an overall survival of 46 ± 7%. While C. ‘calcitrans’ is valuable in the larval culture of P. canaliculus and many other bivalves, appropriate feeding protocols are needed when using this diatom.

The haptophyte Isochrysis aff. galbana (T-iso) was cultured during 2 consecutive years (from spring to autumn) in outdoor horizontal tubular photobioreactors (PBRs) that had a size compatible with that recommended for hatchery service. Around one-third of the cultures carried out in these PBRs showed immediate growth after inoculation up to a maximum biomass. In all other instances a lag phase or a culture collapse were found. In the cultures that showed productivity, average growth rate, μ (d−1), productivity (g L−1 d−1) and photosynthetic efficiency, PE (%), were 0.39 ± 0.085 d−1, 0.075 ± 0.038 g L−1 d−1 and 2.51 ± 0.85%, respectively. These values were low compared to values for cultures routinely incubated indoors, which were used as inocula for the outdoor cultures, and were studied as a reference. In these 50-L acrylic columns (19.4 cm internal diameter) growth, productivity and PE were 0.91 ± 0.087 d−1, 0.076 ± 0.012 g L−1 d−1 and 13.72 ± 2.23%, respectively. Outdoor productivity of T-iso was limited to average temperatures above 15 °C, while below an average temperature of 21 °C growth rate and productivity were lower than above 21 °C. Above this temperature, solar irradiance and biomass density had a more important effect on growth rate and productivity than temperature. Analysis of our data revealed a critical value of the ratio between incident solar irradiance (MJ m−2 d−1) and cell density (106 cells ml−1) at the moment when the protective black netting was removed from the cultures. Cultures that had a ratio above 1.5 suffered an initial lag phase or collapsed, while cultures that showed direct growth had a value below 1.5. This seemed to be independent of absolute initial cell density. However, we recommend using cell densities higher than the ca. 5 × 106 cells ml−1 used in the present study. In the South of Spain this seems to be especially relevant in spring, when temperatures are relatively low in relation to incident solar irradiance. T-iso seems to be particularly sensitive to the variable conditions outdoors and we suggest that new designs of closed PBRs, specifically developed for outdoor cultivation of T-iso, try to achieve more stable and homogeneous conditions throughout the system and over the daily cycle.

Biomass and lipid productivity, lipid content, and quantitative and qualitative lipid composition are critical parameters in selecting microalgal species for commercial scale-up production. This study compares lipid content and composition, and lipid and biomass productivity during logarithmic, late logarithmic, and stationary phase of Nannochloropsis sp., Isochrysis sp., Tetraselmis sp., and Rhodomonas sp. grown in L1-, f/2-, and K-medium. Of the tested species, Tetraselmis sp. exhibited a lipid productivity of 3.9-4.8 g m(-2) day(-1) in any media type, with comparable lipid productivity by Nannochloropsis sp. and Isochrysis sp. when grown in L1-medium. The dry biomass productivity of Tetraselmis sp. (33.1-45.0 g m(-2) day(-1)) exceeded that of the other species by a factor 2-10. Of the organisms studied, Tetraselmis sp. had the best dry biomass and/or lipid production profile in large-scale cultures. The present study provides a practical benchmark, which allows comparison of microalgal production systems with different footprints, as well as terrestrial systems.

Chromatographic and mass spectrometric analysis of surface sediments from perennially ice-covered Lake Fryxell (McMurdo Dry Valleys, East Antarctica) enabled tentative identification from mass spectral data of previously unreported pentaunsaturated ketones, octatriacontapentaen-2-one (C38:5Me), nonatriacontapentaen-3-one (C39:5Et) and tetradecapentaen-2-one (C40:5Me). In addition, the C37, C38 and C40 alkenones occur as di, tri and tetraunsaturated compounds, whereas the C39 alkenones only include the tetraunsaturated components. Analysis of environmental DNA extracted from the water column of Lake Fryxell produced two 18S rDNA sequences closely related to Isochrysis spp., Chrysotila lamellosa and haptophyte sequences previously recognized in sediment samples collected from a Greenland lake and from Ace Lake, Antarctica. Based on the alkenone distribution, coupled with phylogenetic data, we propose that haptophytes related to Isochrysis and/or C. lamellosa are the source organisms of alkenones in Lake Fryxell. The occurrence of pentaunsatured alkenones may represent a response to the year-round cold water temperature (ca. 1 °C average), likely combined with oxygen supersaturation and growth under light- and nutrient-limited conditions in the upper water column.

The variation of fatty acid (FA) content and composition of the microalgal production (Isochrysis sp., Pavlova lutheri and Chaetoceros muelleri) in a continuous large-scale production system (500-L bags) used in hatcheries were analysed. The variation of the FAs was analysed in replicate bags over time for the different species. Total FA content (pg cell−1) increased significantly (p

Growth of marine copepods is influenced by feed. The purposes of this trial were to observe both growth and fatty acid compositions of harpacticoid copepod nauplii, Tisbe holothuriae by feeding with several microalgal species in laboratory: (A) Isochrysis tahiti; (B) Nannochloropsis oculata; (C) Rhodomonas sp., and (D) Tetraselmis chuii. The trial was carried out for 35 days with randomized complete design and triplicates in each treatment. The results showed that final copepod nauplii densities were not significantly different (P>0.05) in all treatments. However, lipid content of copepod nauplii fed with T. chuii was significantly higher (P

Both food type and quantity have a strong effect on the egg production, RNA:DNA ratio and RNA content of the copepod Acartia sinjiensis. Copepods were fed limiting (150 μg Cl−1) and non-limiting (1500 μg Cl−1) concentrations of Tetraselmis chuii (Prasinophyceae), Pavlova salina and Isochrysis aff. galbana (Prymnesiophyceae), and Chaetoceros muelleri (Bacillariophyceae). Saturation of copepod egg production was estimated for Tetraselmis and Pavlova offered in a wide range of concentrations (0–1500 μg Cl−1). Under non-limiting food conditions, A. sinjiensis produced more eggs when fed Tetraselmis (mean 17.6 eggs female−1day−1−efd) and Pavlova (14.2 efd), and produced fewer eggs when fed Isochrysis (9 efd) and Chaetoceros (7.6 efd). RNA: DNA ratio (mean 7) and individual RNA content (mean 2 μg RNA ind−1) were significantly higher in females fed abundant Tetraselmis in comparison to the other algae. Under food-limiting conditions, A. sinjiensis could still produce eggs when fed Pavlova (3.3 efd) and Tetraselmis (1.7 efd), but failed to produce eggs when fed Isochrysis and Chaetoceros. In food-limited females RNA:DNA ratios and individual RNA content were variable and lower than animals fed non-limiting algae concentrations. The saturation of egg production was 714 μg Cl−1 for Tetraselmis and 509 μg Cl−1 for Pavlova. The relationship of RNA:DNA ratio and food concentration followed a similar function to egg production, with a saturation level of 509 μg Cl−1 for Tetraselmis. Egg production was positively correlated with female size only in animals fed Tetraselmis at 1500 μg Cl−1; all other algae and food concentrations were uncorrelated with female size. Of the algal species tested, Tetraselmis and Pavlova are candidate algae for use in A. sinjiensis culture. Egg production saturates around 500 μg Cl−1, as is the case in other Acartia species. Both RNA:DNA ratio and RNA content of the females were sensitive to different food type and concentration, and have the potential to be used as an alternative method for the identification of optimal food for use in Acartia culture. These observations have important implications for the understanding of the dynamics and productivity of this species in coastal environments of tropical Australia, and also its potential use in aquaculture and ecotoxicology.

The enzyme-labeled fluorescence (ELF-97®) substrate produces an insoluble, fluorescent yellow-green product at the site of alkaline-phosphatase (AP) activity. Fifteen coastal phytoplankton species were tested to determine if AP activity could be detected in phosphate- depleted media. All species tested, except Synechococcus bacillaris, expressed AP activity. Subsequently, threshold concentrations of soluble reactive phosphate (SRP) at which AP activity could be detected by ELF-97® were deter- mined for Chaetoceros neogracile, Chlorella autotrophica, Isochrysis sp., Prorocentrum minimum ,a ndTetraselmis chui. Microalgal species differed significantly in the SRP con- centration at which AP activity was first detectable (10.1- 16.4 µM), well above concentrations normally considered limiting for phytoplankton. P. minimum began to express AP activity at a higher SRP concentration than the other algal species; this may be attributable to a relatively high DNA/cell ratio in P. minimum, compared to the other phytoplankton. Thus, phytoplankton species may respond to phosphorus deficiency at high SRP concentrations.

The activities of extracellular enzymes that initiate the microbial remineralization of high molecular weight organic matter were investigated in the water column and sandy surface sediments at two sites in the northeastern Gulf of Mexico. Six fluorescently labeled polysaccharides were hydrolyzed rapidly in the water column as well as in permeable sediments. This result contrasts with previous studies carried out in environments dominated by fine-grained muds, in which the spectrum of enzymes active in the water column is quite limited compared to that of the underlying sediments. Extracts of Spirulina, Isochrysis, and Thalassiosira were also used to measure hydrolysis rates in water from one of the sites. Rates of hydrolysis of the three plankton extracts were comparable to those of the purified polysaccharides. The broad spectrum and rapid rates of hydrolysis observed in the water column at both sites in the northeastern Gulf of Mexico may be due to the permeable nature of the sediments. Fluid flux through the sediments is sufficiently high that the entire 1.5 m deep water column could filter though the sediments on timescales of a few days to two weeks. Movement of water through sediments may also transport dissolved enzymes from the sediment into the water column, enhancing the spectrum as well as the rate of water column enzymatic activities. Such interaction between the sediments and water column would permit water column microbial communities to access high molecular weight substrates that might otherwise remain unavailable as substrates.

Arachidonic acid (ARA, 20:4 n − 6) is essential for oysters and has been implicated in reproduction and immune response of different mollusk species, probably by modulating the levels of prostaglandins. We tested the effect of dietary ARA on reproduction, PGE 2 levels, and some immune variables in Crassostrea corteziensis . Oysters were fed Isochrysis affinis galbana , clone Tahiti (T-iso) for three weeks, which has low ARA and high DHA (22:6 n − 3) levels and that was supplemented with 0, 1 and 9% ARA-rich lipid emulsions at 10% of algal dry weight biomass using a once-daily batch system to supply the microalgae. Survival and growth were not affected by the ARA enrichment. More gonadic tissue was found in oysters fed 1% ARA but oysters fed the 9% ARA were in a more advanced stage of maturation. ARA-enrichment did not affect fatty acid concentration in membrane phospholipids, but ARA concentration was higher in triacylglycerols of oysters fed 9% ARA compared to 0% ARA diet. PGE 2 and/or PGF 1 α concentration was higher in oysters fed the 9% ARA diet. Total hemocyte count was not affected by ARA enrichment, but the proportion of granulocytes and the superoxide anion activity (SOA) in relation to total hemocyte count were higher in oysters fed the 1% ARA diet. These results suggest that feeding moderate levels of ARA to oysters can boost immune system response and oocyte production. However, high levels of ARA favor final maturation and advanced stages of vitellogenesis but possibly at the expense of immune response.

Selected rotifer enrichment diets were compared on the basis of growth, survival, and fatty acid content of larval Florida pompano, Trachinotus carolinus L. Two 9-day larval rearing trials were conducted using a recirculating aquaculture system. In Trial 1, two microalgae concentrates (Isochrysis sp., ISO; and Pavlova sp.), a semi-moist paste (Protein Selco Plus, PS+), and a combination of the three diets were compared. In Trial 2, PS+ and three dry enrichment diets (Ori-Green, OG; Protein HUFA, PH; and AlgaMac 3050, AM) were compared. Larvae were fed rotifers 4 times daily at a rate of 2.5 rotifers/ml/feeding at 2–9 days post-hatch (DPH). Although larval growth in both trials was likely influenced by differences in culture density resulting from variable survival, in Trial 1 at 9 DPH standard length (SL) of larvae fed Pavlova-enriched rotifers (4.60 ±0.39 mm, mean ± SD) was greater (P

We evaluated the technique of measuring the exhalant siphon area (ESA) as an indicator of feeding activity in the blue mussel, Mytilus edulis. To accomplish this, we established the relationships between ESA measured using video and image analysis and clearance rate (CR) measured simultaneously in mussels exposed to various concentrations of microalgae as a food source in the laboratory. Two size classes of mussels (30 and 60 mm shell length) were fed 6 and 7 concentrations of the unicellular alga Isochrysis galbana., respectively (0, 0.6, 1.2, 1.8, 2.4, 3.0, 6.0 mg L-1). ESA and the variation in feeding activity associated with individual mussels significantly influenced CR in 60mm mussels, whereas effects of algal concentration were not significant within the range tested. Individual variation in feeding activity significantly influenced CR in 30 mm mussels. However, unlike the 60 mm mussels, ESA did not significantly influence CR, whereas algal concentration did have a significant effect in 30...

The growth of a temperate and a tropical strain of Isochrysis was studied at four temperatures and three light intensity levels. All algal cultures were grown in an enriched artificial seawater medium under constant illumination and were bubbled with air enriched with 0.1% CO2. Culture density (cells/ml) was measured periodically during exponential growth. Average growth rates for each combination of temperature and light intensity are presented as doublings/day. Observations of growth under the various culture conditions indicate that T-ISO (tropical strain) grows rapidly over a much broader range of temperatures than does I. galbana. Variations in growth at different light intensity levels suggest that the optimum light level increases as a function of temperature for T-ISO and that the growth of I. galbana is inhibited by high levels of light (1500 μW/cm2). The relative value of both strains as food for larval and juvenile oysters and the advantages and limitations of large scale culture are discussed.

Feeding experiments using commonly-cultured algae were conducted with purple-hinge rock scallops, Hinnites multirugosus Gale, to suggest the application of these algae to hatchery culture of scallops. The uptake of finely divided particulate matter was also studied to ascertain the possible importance of detritus as a supplemental food for the rock scallop. A 1:1:1 mixture by culture volume of Tahitian Isochrysis, Tetraselmis suecica, and Dunaliella salina appeared to be superior as a diet for juvenile rock scallops. Significant growth also occurred in juvenile rock scallops fed T-Isochrysis, T. suecica, Isochrysis galbana, and Rhodomonas sp. Gymnodinium splendens, Monochrysis lutheri, and Dunaliella salina fed alone were poor foods for juvenile rock scallops. Finely divided particulate matter as radiolabeled abalone fecal matter was utilized as food. Significant radioactivity appeared in DNA, RNA, lipid, protein, carbohydrate, and free-reducing substances after one week providing evidence that particulate matter was incorporated into juvenile rock scallop metabolic pathways.

Spawning, larval rearing, and settlement of Mytilus californianus were performed over a one-year period at the University of California, Bodega Marine Laboratory in northern California. A combination of shell scraping followed by a 30 to 60 min immersion in 30 or 60 mM hydrogen peroxide was the most successful spawning stimulus tested. Larvae fed a diet of Isochrysis galbana grew significantly faster than did larvae fed Dunaliella sp. Larval growth rate was significantly more rapid at 20°C than at either 14 or 17°C and no significant differences in mortality rates between these three temperatures were detected. At 17 and 20°C, larvae fed a diet of 80 Isochrysis cells/μ1 daily grew significantly faster than those fed 20 Isochrysis cells/μ1 daily. Mortality of pedi-veligers was high, but survivors settled and metamorphosed on byssal threads cut from adult M. californianus. Plantigrades had a mean growth rate of 7.8 μm/day between days 15 and 33 following settlement. In this study, a temperature of 20°C, a diet of Isochrysis galbana of at least 80 cells/μ1 daily, and a settlement substrate of byssal threads were the most successful conditions for artificial spat production of this potentially valuable bivalve mollusc.

The purple-hinge rock scallop is receiving increasing interest as a candidate for commercial mariculture. Hatchery culture of juveniles is prerequisite since natural recruitment of juveniles is variable and not dependable as a source of stock. Success in carrying larvae through metamorphosis has been limited. Critical to larval culture is a sound understanding of optimal food and feeding strategies. Growth and survival were examined in early larvae fed six algal diets at four separate concentrations. The data clearly illustrate the superiority of both Isochrysis galbana and a Tahitian (T) strain of Isochrysis. Feed concentrations of 1 × 105 cells/ml were found to be optimal for these species of Isochrysis. An I. galbana diet supported 80% survival through metamorphosis. The other algae examined included Mono-chrysis lutheri, Rhodomonas sp., Carteria pallida, and a 1:1:1 mixture of Rhodomonas, Monochrysis, and Isochrysis. All these diets supported 5–6% survival, and the growth was only slightly better than starvation controls. A similar study was done with early juveniles (1 mm dia). The diets included: I. galbana, M. lutheri, Thalassiosira pseudonana (3H), T-Isochrysis sp., and Phaedactylum tricornutum. As with the larvae, superior growth and survival occurred with diets of both species of Isochrysis. Cell concentrations between 1 × 105 and 2 × 105 cells/ml were optimal. The other diets supported minimal growth. In the feeding runs of both larvae and early juveniles, cell densities in excess of 5 × 105 cells/ml were detrimental.

Studies on spawning and larval culture in the purple-hinge rock scallop (Hinnites multirugosus) are described together with information on growth of juveniles and adults. Gravid individuals were collected from field populations throughout most of the year. These produced viable eggs when induced to spawn in the laboratory. Larval development appeared normal over the range 8–18°C (optimal for culture, 14–18°C). Acceptable growth and survival were achieved on a mixed diet of Isochrysis, Monochrysis, Tetraselmis and Skeletonema. Metamorphosis began in larvae 200 μm in shell diameter (age, 3 weeks). Most rapidly growing post-larvae reached 1.8 mm at two months (15–18°C). Pre-cementing juveniles held in cages in Mission Bay, San Diego, averaged approximately 8 mm/mo during summer (18–22°C) and half that rate throughout the rest of the year (13–18°C). Individuals 1.5 to 3.0 cm in October, 1975, reached shell diameters of 5.0 to 7.5 cm one year later.

1. Dreissena polymorpha is an extraordinarily successful invasive species that shows high recruitment of small juvenile mussels on established mussel banks. Such juvenile settlement on, and overgrowth of, large adult mussels; however, leads to competition with adults, and often at high densities and low-food concentrations. 2. The concept of food thresholds for zero growth has been a powerful approach to explaining size-related exploitative competition in different zooplankton species. We applied it to investigate whether food threshold concentrations for zero growth (C 0 ) differ between juvenile and adult zebra mussels. 3. By determining body mass growth at various concentrations of a diet mixture (Nannochloropsis limnetica and Isochrysis aff. galbana) we demonstrate that the threshold food concentration for growth of juvenile mussels (C 0 = 0.08 mg C L -1 ) is substantially lower than that for adults (C 0 = 0.36 mg C L -1 ). 4. This indicates that, at low food availability, juvenile zebra mussels are competitively superior to their larger conspecifics. Within zebra mussel banks plankton food is substantially depleted and so the observed mechanism might ensure juvenile success and therefore the regeneration of mussel banks in nature.

The activities and structural specificities of extracellular enzymes that initiate microbial remineralization of high-molecular-weight (MW) organic matter were investigated in surface waters and sediments of an Arctic fjord of Svalbard. Hydrolysis rates of a suite of fluorescently labeled macromolecular substrates, including seven commercially available polysaccharides and three high-carbohydrate-content plankton extracts ranged from rapid to not detectable, and differed markedly between seawater and sediments. Order (fastest to slowest) of hydrolysis in surface water was laminarin, Spirulina extract, xylan>chondroitin, alginic acid, Wakame extract>arabinogalactan, fucoidan> Isochrysis extract⋙pullulan, while in sediments the order was pullulan, laminarin, alginic acid, Wakame extract>chondroitin, xylan>arabinogalactan, Isochrysis extract> Spirulina extract>fucoidan. These differences cannot be explained by simple scaling factors such as differences in microbial cell numbers between seawater and sediments. Other investigations have shown that microbial community composition of Svalbard sediments and of polar bacterioplankton samples differ markedly. These results demonstrate that sedimentary and seawater microbial communities also differ fundamentally in their abilities to access specific high-MW substrates. Substrate bioavailability depends on the capabilities of a microbial community, as well as the chemical and structural features of the substrate itself.

The major bottleneck of Octopus vulgaris culture is the rearing of its paralarval life stage, being the obtainment of adequate live prey to feed paralarvae one of the key issues for the success of the culture of this valuable species. Artemia has been widely used as a single prey or in combination with crustacean zoeae as food items for paralarvae, but few works have reported the biochemical composition of these prey. The gross biochemical composition and fatty acid profile of Artemia juveniles enriched with four marine microalgae of controlled biochemical composition was assessed, as well as the fatty acid composition of newly hatched O. vulgaris paralarvae, in order to estimate which prey would be more suitable to meet the nutritional requirements of O. vulgaris paralarvae. Microalgae were cultured semi-continuously with a daily renewal rate of 30% of the volume of the cultures in nutrient saturated conditions, in order to achieve biomass of constant and optimal biochemical composition. Artemia juveniles of two different sizes (1.5-2.0 mm and 3.0−3.5 mm), appropriate to feed O. vulgaris paralarvae, were obtained by growing Artemia nauplii with Tetraselmis suecica for 2 and 4 days and then enriched for 26 h with four marine microalgae: T. suecica , Isochrysis galbana , Isochrysis aff. galbana (T-ISO) and Rhodomonas lens . The protein content of R. lens (62% of dry weight) was considerably higher than that of the remaining microalgae (42–44%) ( P I. galbana ( P Artemia (3-day old, 1.5-2.0 mm) contained nearly 51% protein (of dry weight) regardless the enrichment diet used, with the exception of individuals enriched with I. galbana (AISO) (41%) ( P R. lens (ARHO), both with circa 16% ( P P Artemia (5-day old, 3.0-3.5 mm) had higher protein percentages than small juveniles with values ranging between 64 and 68% for all treatments, whereas the lipid fraction among groups increased in the order: ARHO (10%) P P Artemia was found in individuals from groups AISO or ARHO (circa 9%), whereas in 5-day old juveniles the highest value was found in group AISO (14.6%) ( P P Artemia maximum percentage of 22:6n−3 was found in group AT-ISO (3.9%) ( P Artemia juveniles enriched with T. suecica contained 22:6n−3. The fatty acid composition of O. vulgaris paralarvae revealed a much higher percentage of 22:6n−3 (18.3%) than the values found in the enriched Artemia juveniles, suggesting a deficit of this fatty acid in Artemia . Even though the highest sum of 20:5n−3 and 22:6n−3 was found in Artemia juveniles enriched with I. galbana , if the general biochemical composition of Artemia juveniles is taken into consideration, the enrichment with R. lens would provide the best composition to meet the possible nutritional requirements of O. vulgaris paralarvae.

Growth rates, internal nitrogen (N) pools of NH4 + and urea, and urease, the enzyme responsible for the catabolism of urea to NH4 + and CO2, were investigated in laboratory cultures of 5 species of estuarine phytoplankton grown on NO3 - , NH4 + , and urea. The phytoplankton used were the dinoflagellates Prorocentrum minimum, Karlodinium veneficum, Heterocapsa triquetra, the cryp- tophyte Storeatula major, and the haptophyte Isochrysis sp. All dinoflagellates had larger internal pools of NH4 + than did the other species. With the exception of H. triquetra grown on NO3 - , this was also true for internal pools of urea. Two of the harmful dinoflagellates, P. minimum and K. veneficum, had significantly higher urease activity rates on both a per cell basis and a per cell volume basis than the other species. These dinoflagellates had different rates of urease activity when grown on differ- ent N sources, suggesting that urease may be down-regulated by NH4 + or up-regulated by urea and/or NO3 - . In all dinoflagellates, the intracellular urea concentrations were greater than the half- saturation constant for enzyme activity, suggesting that in vivo urease activity rates were nearer to maximal in those species. Results from this study and previous studies suggest that harmful dinofla- gellates may be better adapted to utilize urea than other species based on rates of high urease activ- ity and large intracellular urea pools, providing some insight as to why these species may proliferate when urea is a significant N source in the environment.

The Asian oyster, Crassostrea ariakensis, is being evaluated for its potential success in the restoration of oyster populations in Chesapeake Bay. Critical to an understanding of its potential success is knowledge of the impacts of common harmful algae in its diet; blooms of such algae are common in Chesapeake Bay. In these experiments, C. ariakensis were exposed to a standard algal diet, Isochrysis sp., alone, and in combination with the harmful dinoflagallates Prorocentrum minimum and Karlodinium veneficum and the raphidophytes Heterosigma akashiwo and Chattonella subsalsa. Two experiments were run, with varying proportions of Isochrysis to the test algal species. Feces and pseudofeces were examined microscopically and by high- performance liquid chromatography for changes in diagnostic pigments relative to the initial diet and for production of degradation pigments of chlorophyll. All species were cleared from suspension to varying degrees by the oysters. In the Isochrysis control and in the Isochrysis + P. minimum treatment, intact, solid feces were produced, and the highest proportion of chlorophyll degradation pigments were found in the Isochrysis control diet suggesting algal digestion. Thin, ''ropey'' feces and pseudo-feces were observed in the K. veneficum + Isochrysis treatment. Virtually no degradation pigments were found for oysters fed a diet containing K. veneficum, suggesting lack of digestion and assimilation of algal food. With the raphidophytes, reduced production of feces and pseudofeces was evident, and these contained a lower proportion of recognizable harmful algal cells, and higher proportion of degradation pigments than found with the other test species. Amorphous and cellular material that appears to be partly derived from the oyster digestive system was evident in the feces of oysters exposed to K. veneficum, H. akashiwo, and C. subsalsa; this was particularly pronounced in oysters exposed to H. akashiwo and suggests damage to the oyster gut. The impact of the presence of harmful algae in the diet of the oysters thus varied by algal species, but in all cases oyster digestion was altered relative to the control diet.

Blue king crab (Paralithodes platypus) larvae were cultivated to test the effects of diet, temperature and rearing density. Dietary treatments included no feeding (unfed), Artemia nauplii enriched with diatoms Thalassiosira nordenskioeldii (THAL), unenriched Artemia fed in addition to Thalassiosira (A+THAL) and a control diet of Artemia enriched with frozen Isochrysis paste (ISO 6). Trials were conducted at 6 °C, and a rearing density of 10 zoea L−1, with six replicates per treatment. The ISO 6 diet was also tested at 3 °C (ISO 3) and 9 °C (ISO 9), and at densities of 20 (ISO 20) and 40 (ISO 40) zoea L−1. Survival of zoea larvae fed the A+THAL diet (91.7%) was significantly higher than all others, whereas unfed zoea larvae died within 2 weeks. Temperature and rearing density had no significant effects on survival. Time required to reach stage C1 was significantly greater at 3 °C (109 days) than at 6 °C (70 days), but did not decrease further at 9 °C. After reaching the postlarval (glaucothoe) stage, half of the replicates in the ISO 20 and ISO 40 treatments were fed continuously, but survival did not differ significantly from unfed glaucothoe. We conclude that blue king crab larvae are not lecithotrophic and can be cultivated with high survival using the proper diet. These techniques can be used to produce large numbers of juvenile crab for laboratory research, or could be modified for use in stock-enhancement programmes.