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13 páginas, 3 figuras, 3 tablas, The nutritional quality of several microalgal diets used in the seed culture of the little-neck clam, Rudifupes decussatus, was evaluated in the present study. The live diets tested were Isochrysis galbana, clone T-ISO, Tetraselmis suecica and Phaeodactylum tricornutum. Criteria used in the evaluation of the diets were acceptability, digestibility, growth and biochemical composition. The highest growth rate was registered in the seed fed the T. suecicu diet, followed by the seed fed I. galbana, while the lowest one was obtained in the seed fed P. tricornutum. Food ingestion rates were higher in the seed fed Tetruselmis. This fact would partly explain the higher growth observed with this diet. The limited nutritional value of Phaeodactylum could be related to the low digestibility of its cell wall, the degree of acceptability of this diet being similar to or even higher than that of the other two. The low protein content of Tetraselmis seems to indicate that R. decussatus shows a low requirement for proteins, which can be spared by carbohydrate, an abundant component in the cells of Tetraselmis. Lipid needs of this bivalve also seem to be rather low., This study has been financed by CICYT-CSIC-IEO I + D project MAR90-0812-C02-01.

At the Leibniz-Center for Tropical Marine Ecology in Bremen, Germany, shrimps (Litopenaeus vannamei) fed feeds with reduced fish meal content, algae and enzyme additions were analysed for their tissue composition. Contents of protein, glycogen, total lipid, linoleic acid (LA), alpha linolenic acid (ALA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) of the shrimp abdomen and of the respective feeds were determined. Reduction of fish meal (FM) content in shrimp feed and its substitution with “mash” (grain distillery waste-”Schlempe”), corn gluten, pea or coarse colza meal did not influence the tissue composition. Additional inclusion of an algae mixture out of Spirulina spp., Phaeodactylum spp., and Tetraselmis spp. to shrimp feeds as well as the single inclusions of Phaeodactylum spp. or Spirulina spp. had no effect on the tissue composition. Furthermore no improvement of the quality of the shrimps due to an enhanced digestibility of mash by enzymes (mixture of corolase, phytase, xylanase) could be detected. In contrast the quality of shrimps seemed to be negatively affected by enzymes in this study. These findings could contribute to making shrimp aqua farming more economic as reduction of feed costs without reduction of the quality of the shrimps would be a large benefit.

We compared electron transport determined by the saturating pulse method with the oxygen evolution of samples of two marine algae, Tetraselmis spp. and Rhodomonas spp., grown in continuous-batch cultures. The relation between oxygen evolution and electron transport is linear over a range of light intensities from 0 to 500 µmol m-2 s-1. The fluorescence measurement is quite easy and could be made suitable for application in situ, which is a large advantage to the measurement of oxygen evolution and 14C incorporation.

Standard algal growth rate inhibition bioassays can lack environmental realism and may over- or underestimate metal bioavailability in natural systems. In aquatic environments, algal species interact with other biota, including other algae, bacteria and biofilms. In this work, the feasibility of incorporating marine biofilms into 72 h algal growth inhibition toxicity tests was explored. The effects of copper on Tetraselmis sp. were tested in the absence and presence of characterised field-collected biofilms. We hypothesised that the addition of biofilm would prevent copper toxicity to the alga primarily through interactions of the metal with other cells and biofilm exudates. The sensitivity of Tetraselmis sp. to copper (based on 72 h IC50 values; the copper concentration to inhibit population growth by 50%) in the presence of a blended biofilm inoculum varied 2-fold and was independent of the amount of biofilm added. However, increases in IC10 and IC20 values indicated some amelioration of copper toxicity. When intact biofilms were added to the bioassays, amelioration of toxicity was more consistent, probably due to increased binding of copper to cell surfaces or exudates. Difficulties in characterising biofilms and distinguishing that material from the test alga need to be overcome before biofilms can be routinely incorporated into laboratory bioassays.

Brachionus plicatilis females were maintained for > 24 hours in water where the concentration of oxygen was precisely controlled (spherical flasks with ≥ 6 mg 1−1 or < 0.5 mg 1−1; food = dead Tetraselmis sp.). Each female was randomly taken from one flask and quickly placed in an observation chamber containing the same experimental conditions. The swimming path was videotaped (5 minutes); then the size of the female was measured. The tape was analyzed by automatic tracking (25 x, y coordinates of the center of the animal, in a 512 × 512 pixels space). The swimming path was analyzed for ~45 females in both treatments. The speed (mm s−1 body length−1) was calculated for all trajectories, or only for those segments where females swam in a horizontal plane. This relative speed significantly decreased when the concentration of oxygen was very low. There was a negative correlation between the linear speed and the angular speed. The spatial sinuosity (‘S’ of Bovet & Benhamou, 1988) was calculated. The trajectories were significantly more sinuous when the concentration of oxygen was very low. These results could explain the accumulation of some rotifers in the oxycline; rotifers may spend more time in very low concentrations of oxygen by slowing and by turning more.

A system is described for determination of LC50 or IC50 by an iterative process based on data obtained from a plate reader using a marine unicellular alga as a target species. The esterase activity of Tetraselmis suesica on fluorescein diacetate as a substrate was measured using a fluorescence titerplate. Simultaneous analysis of results was performed using an iterative process adopting the sigmoid function View the MathML source (dose of toxicant/IC50)slope for dose-response relationships. IC50 (± SEM) was estimated (P < 0.05). An application with phosalone as a toxicant is presented.

Scale biogenesis in algae represents a unique model system to study the transport of secretory macromolecules through the Golgi apparatus (GA) and their exocytosis. The larger scales can be visualized in the light microscope, and thus the kinetics of scale assembly, transport, and secretion can be studied in vivo. In addition, scales are osmiophilic and readily visible in conventional transmission electron microscopy; thus, details of scale assembly and sorting can be studied without invoking immunolabeling techniques. The following are distinctive features of scale biogenesis in algae: 1) transport of scales through the GA-stack occurs by cisternal progression; 2) scale secretion may be very rapid (in some cases a single GA-cisterna leaves the stack every 15-20 s); 3) sorting of different scale types does not occur in the GA, but in a post-GA-compartment. Recent progress in the analysis of scale formation in the green flagellates Tetraselmis and Scherffelia is reviewed.

This paper studies the influence of the flow rate of gaseous mixtures on the kinetics of growth and the fatty acid composition of Tetraselmis sp. at CO2/air ratios of 3 × 10−4 and 2 × 10−5. The specific growth rate rises with increased flow rate up to values of approximately 0.086 h−1 and 0.063 h−1 at CO2/air ratios of 3 × 10−4 and 2 × 10−5 respectively, when the flow rate is approximately 3 v/v per minute. At higher flow rates, the specific growth rate decreases. The polyunsaturated fatty acid content decreases slightly as the gaseous mixture flow rate increases, whereby the ratio ω3/ω6 remains between 2 and 3, indicating good nutritional values.

Of a wide range of algae tested, juvenileConvoluta roscoffensisingested onlyPlatymonas convolutae, the natural symbiont; related species of the generaPlatymonas, PrasinocladusandTetraselmis; andChlamydomonas coccoides. Platymonas convolutaewas not ingested to a greater extent thanPrasinocladus marinus, Tetraselmis tetratheleandTetraselmis verrucosa, or taken up in preference toT. verrucosawhen animals were exposed to a choice between the two species. Convoluta ingested fewer cells ofC. coccoidesthanP. convolutaeand related species. Uptake ofP. convolutaewas not affected by pretreatment of the cells with lectins or proteases, incubation in media of pH 5·0–9·0 or inhibition of algal photosynthesis, but was substantially reduced if the algae were killed.Cells ofP. convolutae, Pr. marinus, T. tetratheleandT. verrucosapersisted and divided in juvenile Convoluta. The algal population in the worms started to increase 2–3 d after ingestion and within 15–20 d the animals were uniformly green. These algae formed a viable symbiosis with Convoluta and promoted the growth of the animals. In contrast,C. coccoidescells did not persist in Convoluta for more than 12–24 h a nd were probably disrupted.P. convolutaecells lost their thecae within a few days of ingestion and before migration from the central to sub-epidermal region of the animal. Animal vacuoles surrounded recently ingested thecate algae. Structural studies of the adult symbiosis suggest that the algae were also intracellular and enclosed in vacuoles.It is proposed that Convoluta discriminates against algae unrelated toP. convolutaeon initial contact and in the central region of the host. The nature of the recognition mechanism(s) has not been established.

Growth and gross chemical composition of two axenically cultured algal flagellates, Dunaliella tertiolecta and Tetraselmis maculata, were related to nitrate and phosphate concentrations in an enriched seawater medium. Population growth of test species in 28-day batch cultures was compared in 16 combinations of NaNO3 (77.5, 150, 300, and 350 mg/l) and KH2PO4 (5, 10, 20, and 25 mg/l) in an experiment of factorial design. Phosphate-P was not limiting to final populations of either alga at concentrations tested, and nitrate-N did not affect final population density of D. tertiolecta at the lowest concentration. In contast, final populations of T. maculata were considerably reduced in all media containing the lowest NaNO3 concentration, but maximal populations were achieved in 150 mg/l NaNO3. This was observed at a wide range of N:P ratios and appeared to be related to available N only. Production of protein, carbohydrate, and lipid, as percentages of dry weight, was evaluated for selected concentations of NaNO 3 KH 2 PO 4 ( 77.5 5 , 150 5 , 300 5 , 300 20 mg/l) At low P, protein/carbohydrate showed a direct linear relationship to N in both species. Only D. tertiolecta cultured at low N increased lipid production. Effect of increasing P in high N cultures was to increase protein/carbohydrate in T. maculata, but to decrease this ratio in D. tertiolecta. Maximal yields per unit volume were attained in the following NaNO 3 KH 2 PO 4 , as mg/l: T. maculata, dry weight and lipid - 300 5 , protein - 300 20 , carbohydrate - 77.5 5 ; D. tertiolecta, dry weight - 150 5 , protein - 300 5 , carbohydrate and lipid - 77.5 5 .

Summary The two rhizoplasts of Tetraselmis cordiformis are firmly attached to the nuclear envelope and the plasmalemma in addition to their connection with the flagellar apparatus. In the region of contact the freeze-fractured nuclear membrane is indented and partly devoid of nuclear pores. Links are probably mediated via the cross-bars of the rhizoplast at certain specific sites. At the plasma-lemma the contact zone in freeze-fractured cells displays a “ridge” of 5 × 0.4 μm. dimensions. Intramembraneous particle numbers on the E-fracture face are significantly higher in this region than in other parts of the plasmalemma. Variations in certain aspects of rhizoplast ultrastructure and its associations with membranes are discussed in accordance with the proposed contractile function of the rhizoplast. Evidence is presented that shortening preferentially occurs in the proximal region of the rhizoplast near the flagellar apparatus and that the “normal” image of an evenly spaced rhizoplast is probably a fixation artifact.

The growth of Phaeodactylum tricornutum , Bohlin, Tetraselmis spp. Stein, Dunaliella primolecta Butch., and Cricosphaera elongata (renamed Hymenomonas elongata (Droop) Parke et Green, comb. nov.) in batch culture in S88 medium is not arrested on addition of copper, cadmium, or lead below a concentration of 10 −4 M. In contrast, Ditylum brightwelli (T. West) Grunow ex Van Heurck, undergoes osmotic disturbances in 10 −5 M and 10 −4 M copper, with swelling of the cell contents. Phaeodactylum grown in continuous culture survives single doses of up to 10 −3 M copper with no diminution in growth and with considerable uptake of copper. Cricosphaera in continuous culture survives up to 10 −4 M copper with no long-term diminution in growth, but takes up between ten and a hundred times less copper than Phaeodactylum .

In clonal culture Scherffelia dubia (Perty) Pascher shows a considerable range of morphological variation, with different cell shapes being produced including those typical for three other species of Scherffelia Pascher (S. opisthostigma Skuja, S. ovata Pascher, S. pelagica Skuja) and one species of Tetraselmis Stein (T. bichlora Ettl & Ettl). It is concluded these taxa should be reduced to the synonymy of S. dubia, the type species of Scherffelia. The general cell morphology of S. dubia is described in detail, including a serial section and absolute configuration analysis of the flagellar apparatus. S. dubia is compared with species of Tetraselmis of which ultrastructural details are known and it is shown that both genera are very similar but the generic distinction is nevertheless justified. Main features of distinction include the pyrenoid (present in Tetraselmis but lacking in Scherffelia) and the chloroplast (single and more or less cup-shaped in Tetraselmis, two separate lateral chloroplasts in Scherffelia). Both genera are accommodated in the same family, the correct name being Chlorodendraceae and not Tetraselmidaceae or Platymonadaceae which have often been used.

A machine to provide continuous supplies of Tazadit iron ore suspensions to queen scallops Chlamys opercularis held under controlled conditions of food supply, temperature and illumination, is described. The influence of iron ore suspensions, 0 to 70 mg litre -1 , and micro-algae Tetraselmis suecica 0 to 1·3 × 10 4 cells per millilitre, on shell and tissue growth of C. opercularis has been studied. The main effect of iron ore was to reduce shell growth rates. Queen scallops exposed to iron ore suspensions (10·7 to 70 mg litre -1 ) from two experiments developed shell thickening on the interior surface of the shell valves. It is suggested that this condition, which is absent from queen scallops fed algal diets only, was caused by a general failure of the mantle edge to extend fully in the presence of iron ore particles in suspension. Unlike shell growth rates, there were no significant differences (p > 0·05) between the soft tissue weights of C. opercularis grown in concentrations of 0·4 to 70 mg Tazadit ore per litre and T. suecica and those grown only on T. suecica . Queen scallops provided with a diet of 6·67 × 10 3 Tetraselmis cells per millilitre produced average laboratory shell growth rates ranging from 19·0 to 110·0 μm day -1 depending on the size of the animal and time of year the experiment was carried out. These rates are comparable with rates found in suspended cages in the natural environment.

Salmon pituitary homogenate was used alone or in combination with human chorionic gonadotropin, to induce spawing in captive and wild adult milkfish at ambient temperature (26–30°C) and salinity of 34%.. Healthy or slightly injured females having oocytes with a minimum mean diameter of 0.66 mm could be induced to spawn in captivity using a total dose of 20 mg SPH/kg + 3000 IU HCG/kg given in two injections. Badly injured females did not respond readily to the hormone and required more than two injections to induce ovulation. Milkfish larvae were reared successfully to metamorphosis using only Chlorella -fed rotifers during the first 10 days. Survival rates were greatly improved when, aside from Chlorella and Chlorella -fed rotifers, Isochrysis galbana and Tetraselmis chuii were added to the rearing tanks.

In order to define the nutritional role of dietary phospholipids (PL) in larval fish, the PL classes and possible fatty acid combinations of phosphatidylcholine (PC) were investigated on the food organisms such as the micro-algae and rotifers Brachionus plicatilis, post-larval prawns (P-30) Penaeus japonicus, and larval fish. PC and phosphatidylethanolamine were the major PL classes in a marine Chlorella (CH), Tetraselmis (TS), rotifers cultured with either CH or TS, red sea bfeam (40-days old) Chrysophrys major, knife jaw (30 days-old) Oplegnathus fasciatus, Ayu (74-days old) Ptecoglossus altivelis, and prawn. The major PC in the larval fish and prawn were those having fatty acid combinations of C16C20 (mainly C16:0, C20:5ω3), C16C22 (mainly C16:0C22:6ω3), and C18C22 (mainly C18:0C22:6ω3 and/or C18:1ω9C22:6ω3). Because larval fish are incapable of de novo synthesis of ω3-and ω6-series of fatty acids and also possibly have a limited ability for the synthesis of some PC, a part of these PC occurring in the larval fish may be derived from food organisms such as the rotifers which contained PC with fatty acid combinations of C16C20 (mainly Cl6:0C20:5ω3).

Phytoplankton cultures of Chlorella sp., Tetraselmis sp. and Isochrysis galbana were used alone as feed to rear separate batches of newly caught milkfish ( Chanos chanos ) fry (about 21 days old). Ultrastructural studies of fry hepatocytes and pancreatic acinar cells indicated that they cannot directly utilize Chlorella , which has a rigid cell wall. The fry can directly utilize Tetraselmis and Isochrysis , but neither is nutritionally adequate for growth and survival if used as the only feed. Tetraselmis is nutritionally inferior to Isochrysis .

Different larval stages of Penaeus monodon were fed with increasing densities of Tetraselmis sp. to evaluate the relationship between food density, ingestion rates, development and growth of Penaeus monodon larvae. Ingestion showed a saturation response to food density. This served as a basis for determining maximum ingestion rates and incipient limiting levels (ILL), defined as the lowest food density to provide maximum ingestion rates (McMahon and Rigler, 1963). Both ILL and maximum ingestion rate increased as larval stage progressed, with maximum ingestion rate reaching a peak at mysis 3 and declining at postlarva 1. When protozoea 1 and protozoea 2 were given Tetraselmis sp. at levels equal to or above their respective ILL's, rate of development to protozoea 3 was enhanced. Growth rate, in terms of total length, was significantly enhanced in protozoea 3 and mysis 1 if earlier stages were given Tetraselmis sp. at levels equal to or above their respective ILL's.

The function of the gill of the mussel Mytilus edulis (L.) has been studied in intact animals and in animals with cut posterior adductor muscle, as well as on gill fragments and isolated gill filaments. Intact 3–4 cm long specimens, kept singly in glass jars containing 600 ml aerated sea-water at 1–2° or 14°C, were able to clear suspensions of various particles at high rates. The gills retained almost completely particles down to about 3–5 µm in diameter (yeast cells or latex spheres, and about half of particles 1–2 µm in diameter (Bacterium sp. or latex spheres). The immediate effect of cutting the adductor muscle was to decrease the retentiveness of the morphologically intact gills; later also the rate of water transport decreased. In intact, fully open mussels, 10−7-10−5 M dopamine (DA) or 10−7-10−6 M serotonin (5-HT) did not affect the rates at which the mussels cleared suspensions of yeast, whereas 10−5 M 5-HT strongly reduced clearances of suspensions of yeast and 18 µm Tetraselmis cells, t...

The effect of the rotifer, Brachionus plicatilis , grown on unialgal cultures of Isochrysis galbana, Tetraselmis sp. and Chlorella sp. on the hepatocytes of milkfish fry was evaluated by electron microscopy. Rotifers grown on the three different species of phytoplankton brought about different ultrastructural features in milkfish fry hepatocytes. Best results were obtained from fry reared on Isochrysis -fed rotifers. The use of marine Chlorella -fed rotifers as feed for fry resulted in the poorest hepatocyte ultrastructure, indicating that this was nutritionally the least adequate cultured food. None of the three diets, however, produced an optimal hepatocyte ultrastructure similar to that obtained by feeding the fry with a mixture of artificial feed and newly hatched Artemia nauplii.

Uric acid is an excellent nitrogen source for the growth of cultures ofPlatymonas convolutae, the symbiotic alga fromConvoluta roscoffensis, and relatedPlatymonasandTetraselmisspecies. Nitrate-grown cells ofP. convolutaeandT. tetrathelehave two uptake systems for uric acid, which conform to Michaelis–Menten kinetics; a high-affinity system operating in the concentration range 0·2–4·5 μM a nd a low-affinity system operating at higher concentrations of uric acid. Uric acid uptake byP. convolutaeis abolished by uncouplers of phosphorylation. In darkness, intact cells ofP. convolutaemetabolize [2-14C]uric acid to [14C]carbon dioxide. These results are consistent with the proposal that the algal symbionts ofC. roscoffensisutilize uric acid, received from the host, as a nitrogen source.Aposymbiotic juvenile and symbiotic adultC. roscoffensisunder standard culture conditions contain uric acid. The solid uric acid content of juveniles declines on establishment of symbiosis withP. convolutaeand the endogenous uric acid is utilized in the adult symbiosis under conditions of nitrogen demand. However, adultC. roscoffensisdo not utilize exogenous uric acid. The growth of adult but not juvenileC. roscoffensisis dependent on nitrogen enrichment of the medium, and it is proposed that uric acid utilization is of significance to the growth of the developing symbiosis in a nitrogen-poor environment.

The effect of salinity on resting egg formation and hatching in the marine rotifer Brachionus plicatilis rotundiformis (S-type Hawaiian stock) was examined in 1986. B. plicatilis was cultured at three salinity levels. The average number of resting eggs produced at 8, 16 and 32 ppt ranged from 22 to 615 per 10 000 rotifers, or 29 to 413 per 1.0x108 Tetraselmis cells. Data on the rate of mictic female appearance, fertilization and fecundity of fertilized mictic females support these findings. Results indicated that the sexual reproduction of L- and S-type rotifers have opposing responses to salinity. After preservation for 2 mo at 5°C, simultaneous hatching of resting eggs was observed at six salinity levels at 30°C. Development time was extended and hatching rate decreased when eggs formed at 8 ppt were incubated at extremely high (40 ppt) or low (1 ppt) salinities. Hatching rates varied from 69 to 93%, and were influenced by the salinity present at the time of egg formation and incubation.

Survival, metamorphosis and growth of four larval shrimp species, Penaeus aztecus, P. setiferus, P. vannamei and P. stylirostris , were compared after feeding six food combinations. The food combinations consisted of the diatoms, Skeletonema costatum and Chaetoceros gracilis , or the phytoflagellates, Isochrysis sp. and Tetraselmis chuii , alone or in combination with live Artemia nauplii. Post-larval survival, within each algal food type, was not improved by feeding Artemia beginning at the second protozoeal versus the first mysis substage. In addition, larvae fed only algae survived as well as their counterparts fed Artemia, P. stylirostris was the only species tested which survived significantly better to the post-larval stage when fed diatoms rather than phytoflagellates, regardless of treatment. Except for P. stylirostris fed phytoflagellates, metamorphic rates to the post-larval stage were not improved by feeding Artemia beginning at the second protozoeal versus the first mysis substage. All larval species fed phytoflagellates only rather than diatoms only were delayed metamorphically. Only for P. stylirostris did all the diatom treatments result in better metamorphic rates when compared with their phytoflagellate counterparts. Post-larval dry weights, for the four shrimp species studied, were improved when fed Artemia at the second protozoeal substage regardless of algal food type. Except for P. aztecus , growth was significantly better for all species fed the diatom treatments versus their phytoflagellate counterparts. Larval growth, in terms of biomass rather than survival and/or metamorphic rates, is a better measurement of the nutritional value of a larval diet.

Experiments to grow Tetraselmis chuii (a marine alga) and Daphnia magna in the presence of inorganic arsenate are described. The algae incorporate arsenic rather efficiently and form a lipid-soluble organic arsenic compound. T. chuii has been successfully mass cultured in a medium containing 10 ppm arsenic as arsenate. Daphnia magna was cultured in a medium containing 74As-labeled H3AsO4 and 1 ppm Na2HAsO4 expressed as arsenic. The arsenic metabolites were extracted with a chloroform-methanol solution and isolated by using column and thin-layer chromatography. TLC analysis of the metabolites revealed the presence of a 74As-containing product which migrated with phosphatidylethanolamine. This product was hydrolyzed with the phospholipases A, C, and D. The experimental results are not inconsistent with the presence of an arsenocholine moiety in the lipids. Arsenocholine, arsenobetaine, and acetylarsenocholine have been synthesized and will serve as reference substances in the chromatography experiments. The preparation of arsenocholine-containing lipids is in progress.

In recent studies, we developed a combined nutrient removal-marine aquaculture process for the tertiary treatment of wastewater and the production of commercially important shellfish. Part of this process consists of an outdoor mass cultivation system for marine algae. During our previous experiments we observed that marine diatoms almost exclusively are the dominant algal species in our outdoor cultures. To better understand this phenomenon of diatom dominance we grew 16 species of marine algae in continuous monoculture under laboratory conditions simulating to some degree the conditions prevailing in our outdoor experiments. Species such as Skeletonema costatum, Monochrysis lutheri and Tetraselmis sp., which were never dominant in our outdoor cultures, grew as well in monoculture, as Phaeodactylum tricornutum, frequently, the prevalent species outdoors. However, when monocultures of Dunaliella tertiolecta and Thalassiosira pseudonana (3H) were purposely contaminated with P. tricornutum, the latter species quickly became dominant. It is suggested that a complex interaction of environmental factors is usually responsible for the dominance of a particular species; one such factor may be the nitrogen source in the growth media. Under conditions of virtually, complete nitrogen assimilation, the carbon: nitrogen ratio in the algae was high (7 to 8) when NO 3 - −N was the source of nitrogen, and low (4 to 6) when NH 4 + −N was the prime form of nitrogen. When algal growth was low, resulting in a large inorganic nitrogen residue, the carbon:nitrogen ratio was low regardless of whether NO 3 - −N or NH 4 + −N was the main nitrogen source.

Chaetoceros calcitrans (Paulsen) Takano, Tetraselmis suecica (Kylin) Butch. and Isochrysis galbana Parke were grown in artificial seawater containing either normal (9·8 mg atoms litre−1) or reduced (0·613 mg atoms litre−1) nitrogen. When grown in the nitrogen-deficient medium protein decreased and carbohydrate increased in all three species; lipid, however, increased in Chaetoceros and Isochrysis but decreased in Tetraselmis. The biochemical composition of Tetraselmis and Isochrysis sampled during the post-exponential phase of growth from normal medium was similar to that of cells growing exponentially in nitrogen-deficient medium. Salinities of 10–35‰ did not affect protein levels in Tetraselmis or Isochrysis but carbohydrate increased and lipid decreased in Isochrysis over the salinity range. Temperatures of 18–25 °C had little effect on the cell composition of either species. The energy potentially available from a unit volume of Chaetoceros and Isochrysis cells from normal medium was similar to that from cells grown in nitrogen-deficient conditions in both species. Tetraselmis had 34% more energy available when grown in normal compared to nitrogen-deficient medium.

Axenic cultures of Chlorella sp., Dunaliella primolecta Butcher, Platymonas subcordiformis (Willie) Hazen, Platymonas sp., Porphyridium cruentum (Ag.) Nag., and Tetraselmis chuii Stein were grown in artificial sea water containing various concentrations of sulfate, selenate, and selenite. In the absence of selenium, most algae were sensitive to reductions in sulfate levels below those concentrations normally found in sea water. When selenium was added, low selenate (≈0.01 ppm Se) concentrations restored the rate of growth previously diminished by low sulfate concentrations, whereas high selenate concentrations (≈10 ppm Se) were lethal. No lethal effects were caused by selenite, even at concentrations as high as 100 ppm. The effects of selenite were somewhat conditioned by the sulfate concentration of the medium. Taken in toto, these experiments suggest that the effects of selenium on marine algae depend on the species of the alga, the concentration and oxidation state of the element, and the concentration of sulfate. Uptake studies indicate that selenite and selenate are incorporated into the algal cells.

A survey of the algal genus Tetraselmis (Chlorophyta, Prasinophyceae) was conducted using starch gel enzyme electrophoresis. Thirteen enzyme loci were resolved and scored, of which nine were invariant among strains. Phenotypes at the four variable loci, glyceraldehyde-3-phosphate dehydrogenase-1, glutamate–oxaloacetic transaminase-2, malate dehydrogenase-1, and peptidase-3, were strongly linked. Two groups, to which 58 of 66 strains were clearly assignable, could be constructed on the basis of the electrophoretic results. These groups do not appear to correlate with ecological, geographic or physiological characters and may have taxonomic significance. A number of strains had electrophoretic phenotypes characteristic of heterozygotes, and presumably were therefore diploid. This could indicate the occurrence of sexual reproduction, heretofore unknown in the class Prasinophyceae.

SUMMARY The relationship between assimilation of nitrogen and animating activities of glutamine synthetase (GS), NADH-glutamate dehydrogenase (GDH) and NADPH-GDH in ammonium-grown Chlamydomonas pulsatilla Wollen-weber and ammonium- and nitrate-grown Brachiomonas submarina Bohlin and Tetraselmis succica (Kylin) Butcher was investigated. The three flagellates show minor changes (15–60%) in enzyme activities when ammonium is replaced by nitrate as the nitrogen source or the concentration of nitrogen is lowered from 2 to 0·2 mM. The activities of NADH-GDH and NADPH-GDH in the three flagellates do not appear to be high enough to make a significant contribution to ammonium assimilation in the presence of a highly active GS. Fast protein liquid chromatography (FPLC) of GS revealed the presence of two distinct molecular forms, designated as GS1 and GS2, in C. pulsatilla and T. suecica, whereas only a single GS peak designated as GS2 was detected in B. submarina. Both GS1 and GS2 are stabilized by sorbitol and glycinebetaine in media employed for enzyme extraction and FPLC fractionation. The presence of thiol reagents is essential for GS2 stability, but inhibits GS1 activity. GS2 is by far the more active of the two isoenzymes in C. pulsatilla and T. suecica. This isoenzyme, therefore, appears to be the major port of entry of ammonium in all three flagellates.

Egg capsules collected from wild colonies and laboratory cultures of Convoluta roscoffensis carried a variety of organisms. However, algae were not found on capsules laid by animals that had previously been washed free of exogenous algae. It is concluded that symbiotic algae are not transmitted during reproduction of laboratory-maintained C. roscoffensis and that all the algae associated with the capsules are free-living. Cells of all Platymonas and Tetraselmis species tested aggregated around and adhered to the capsules of C. roscoffensis. No evidence for chemotaxis of P. convolutae and T. tetrathele towards the capsules was found.

Females of Siganus guttatus reared to sexual maturity in canvas tanks were induced to spawn by using human chorionic gonadotropin (HCG, Ayerst) at 500 IU/fish or about 2 IU/g body weight. The amount of HCG used depended on the initial mean egg diameter; the smaller the diameter, the more HCG was used. Fish with oocytes characterized by germinal vesicle migration (mean egg diameter ≥ 0.47 mm) spawned without HCG injection. Fertilization and hatching rates for both treated and untreated fish were more than 90%. The larvae were reared to metamorphosis using rotifers from day 2–17, rotifers + newly hatched Artemia nauplii from day 18–20 and rotifers + newly hatched Artemia nauplii + artificial feed from day 21–35. In addition, Isochrysis galbana was introduced to the rearing tanks from day 1–10 and Chlorella sp. and/or Tetraselmis sp. from day 1–35. Survival rates of larvae tended to be lower as the broodstock became older.

Rotifers were fed on a laboratory-cultured alga, Platymonas (= Tetraselmis) suecica , and/or a diet composed of commercial single-cell proteins. They were given to sea-bass larvae up to day 15–20 after hatching, while Artemia nauplii were supplied from day 9–13. At day 21, the survival rate of fish obtained with rotifers fed on compound diet ranged between 32 and 82%, and their mean weight between 5.0 and 7.8 mg. When rotifers were fed either only on algae or on a mixture of 33% algae and 67% compound diet, the growth and survival rates of sea-bass were not clearly different to those obtained with rotifers fed on compound diet. No significant difference was observed when these rotifers were enriched with nutrients just before distributing them to fish. However, these rates appeared to be quite high in comparison to those obtained by several other authors. We can therefore recommend rotifers fed on compound diet as being, even without enrichment, convenient and low-cost food for sea-bass larvae.

During 1975 and 1976 it was found that lower growth and therefore productivity from 20 l semi-continuous cultures of Isochrysis galbana Parke and Tetraselmis suecica (Kylin) Butch. was related to an increase in salinity from 31 to 36‰ of the natural sea water used in medium preparation. This effect was removed in 1977 by lowering salinity to 25‰ prior to medium preparation. Small-scale trials with Isochrysis in 1977 showed that growth was improved when the salinity was reduced to 28‰. Optimal salinity ranges for growth in medium prepared from artificial sea water were found to be 15–25‰ for Isochrysis and 25–30‰ for Tetraselmis.

Standard autoradiographic techniques have been used to describe the temporal pattern in the transfer of radioactive carbon from the chlorophyte Tetraselmis suesica (Kylin) to females of the neritic copepod Temora longicornis (Muller, 1792). After 4 and 6 h exposure to labelled phytoplankton, silver grains were deposited only in the cells of the gut-wall. After 24 h exposure, grains were deposited in most tissues, with highest concentrations in cells of the gut-wall and in oocytes within the ovary. Experiments with Centropages typicus (Kroyer), Centropages hamatus (Lilljeborg), and Labidocera aestiva Wheeler demonstrated that much of the assimilated label was not turning over rapidly and that the amount of label assimilated was correlated with known food preferences of these species.

Diets of Chaetoceros calcitrans (Paulsen) Takano and Tetraselmis suecica (Kylin) Butch containing high or low levels of protein were fed to Crassostrea gigas Thunberg larvae and spat. The 4-day increment in D-larvae shell length was not related to differences in the ash-free dry weight or protein content of the diets at cell concentrations of 75–125 Chaetoceros μl −1 . At concentrations of 2–100 Chaetoceros μl −1 the largest growth increment was found when 0.267 μg ash-free dry weight larva −1 d −1 (equivalent to 3.35 Joules · 10 −3 larva −1 d −1 ) was filtered at a temperature of 28.1°C. Percentage settlement of C. gigas increased as protein in Chaetoceros , fed at 75–125 cells μl −1 ( P Tetraselmis , fed at 5–15 cells μl −1 ( P Tetraselmis diet ( P Chaetoceros diets were not significant.

Yields and efficiencies of light utilization were determined for the Chrysophyte alga, Isochrysis (Tahitian strain), and the Xanthophyte, Monallantus salina . Under conditions of nitrogen sufficiency the maximum yield of the former alga was 47·9 g m −2 day −1 with an efficiency of light utilization based on PAR supplied (400–700 nm) of 11·6%. Nitrogen deficiency decreased these values to 7·6 g m −2 day −1 and 1·9% respectively. For Monallantus (N-sufficient) the maximum values were 13·9 g m −2 day −1 and 6·4% respectively; N deficiency reduced these values to 7·5 g m −2 day −1 and 3·7%, respectively. Deficiency reduced protein levels in both species and in Isochrysis metabolism was shunted towards the formation of carbohydrate. Carbohydrate yield was doubled by deficiency. In Monallantus , lipid content was not increased by deficiency; this is in contrast to previous reports. Rather an appreciable amount of some fraction ‘other’ than protein, carbohydrate, lipid or ash was formed. The results of all five marine microalgae studied in this series are compared. Phaedactylum gave the highest yield and efficiency, but similar values were obtained by Tetraselmis and Isochrysis . To produce the maximum amount of a high-energy component such as lipid, one would choose Isochrysis , and for high protein yield, Phaeodactylum or Tetraselmis would be chosen. The highest carbohydrate yield would be attained by Dunaliella, Tetraselmis , or Isochrysis under N-deficient conditions. These comparative results may be useful in choosing species and conditions for maximizing various biomass products in outdoor cultures.

The incorporation of tritiated ([3H]-) thymidine into DNA by axenic laboratory cultures of a common marine blue-green algae and two marine eukaryotic algae was measured. Tritiated thymidine was incorporated into the cold TCA fraction of the cyanobacterium and eukaryote algae. However, only in the culture of cyanobacterium was the thymidine consistently incorporated into DNA. Considering the usual algal densities in natural habitats, thymidine concentrations and incubation times, our data do not preclude the use of thymidine incorporation in bacterial production studies in marine environments.

Assimilation efficiencies reported for bivalves vary, but different experimental conditions, in particular the concentration of the suspension at which the animals are fed, may affect the values found. Widdows & Bayne (1971) determined the assimilation efficiency forMytilus eduliswhen fedTetraselmisand they found that efficiencies decreased with increasing concentration of cells. Allen (1962) fed radioactively labelledPhaeodactylumtoMya arenariaandVenus striatulaand suggested that the amount of P recovered in the faeces was proportional to the amount of algae ingested rather than to concentration of suspension directly. However, Winter (1969) concluded that forArctica islandicaandModiolus modiolusthere was no relation between assimilation efficiency and concentration of algae.

Autotomy of the flagella in Chlamydomonas (Chlorophyceae) and Tetraselmis (Prasinophyceae) may take less than I s to complete. In both genera it follows through similar stages. The flagellar membrane immediately distal to the transition zone detaches from the axoneme and forms a sort of brim or sleeve. At the same level, inside the axial cylinder a section of the core appears to be digested and the outer microtubules are ruptured. The flagellar membrane pinches in, detaching the flagellar shaft, and then seals over the stump as a transverse septum. At the same time, inside the base of the flagellum the upper beaker-shaped portion of the transition body changes. The mouth of the beaker narrows as the sides converge distally. These changes seem to be general, though details differ among genera and species. Underlying physiological responses and biochemical changes remain to be elucidated.

MONOSPECIFIC AXENIC CULTURE OF MARINE PLANKTONIC ALGAE FROM SEWAGE EFFLUENTS Having care for economic aspects, the authors have studied the nutritional value of raw domestic wastewaters, sterilized by U.V. irradiation, precipitated or stored, for the monospecific and axenic culture of marine planktonic algae (Asterionella japonica, Tetraselmis. maculata). Contamination by heavy metals (Hg,Cd,Pb ,Zn,Cu) was controled over all experiments. The fertilizing property of sewages was compared to that of macerations of digested sludges.

The possible roles of algae (i.e. “green water”) in facilitating Macrobrachium rosenbergii larviculture have been investigated. Larvae were incubated with 14 C uniformly labeled Tetraselmis sp. and Phaeodactylum sp. No counts were incorporated by the larvae indicating that negligible quantities of algal cell material are ingested directly. In contrast, Artemia nauplii assimilated and digested appreciable quantities of algae under similar conditions. The algae rapidly removed ammonia that had been added to the larviculture medium even starting at levels toxic to the larvae (0.6 ppm). However, very high densities of Artemia (20/ml) caused ammonia accumulation even in the presence of the algae due to their own increased metabolic rate when fed upon algae and due to the reduced number of assimilating algae. Removal of the Artemia under these conditions was followed by rapid disappearance of the ammonia. The beneficial effects of Tilapia as a larviculture associate, reported elsewhere, was observed only in the presence of algae. Larval mortality actually increased in water that had supported the growth of Tilapia if algae were not present. It is concluded that algae facilitate the growth of Macrobrachium larvae only indirectly by removing toxic material, notably ammonia, from the medium. Under conditions where the Artemia are grown before contact with the Macrobrachium larvae and where other means of ammonia removal are employed, larval growth and development are not affected by the presence of algae and the latter can be dispensed with.

Two algal flagellates, Dunaliella tertiolecta Butcher and Tetraselmis maculata Butcher, harvested in the stationary phase from a semi-continuous carboy culture apparatus, were analyzed for dry weight, total carbohydrate, total protein, and total lipid. Each species was cultured in three different growth media. The growth response of D. tertiolecta was similar in all three formulations but populations of T. maculata were considerably limited in the reduced-nutrient medium, X1. Both algal species cultured in the X1 medium had significantly greater dry weights and contained more carbohydrate and less protein than cells cultured in the standard formulation (E). A third formulation (N/P), in which all medium components were reduced except nitrate and phosphate, produced algae with reduced carbohydrate and increased protein as compared with E medium. The total lipid content of D. tertiolecta was significantly less than that of T. maculata regardless of the culture medium.Algae cultured in the three formulations...