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Information on the nutritional requirements and preferred diets of sea cucumber juveniles is extremely limited and this has hindered development of hatchery culture methods. This study assessed ingestion, cell wall digestion and relative nutritional values of two live micro-algae (Isochrysis aff. galbana (TISO) and Chaetoceros muelleri) and six concentrated micro-algae products (Instant Algae®, Reed Mariculture Inc.) for early juveniles of sandfish, Holothuria scabra. Seven of the eight micro-algae tested were ingested by juveniles with the exception of live TISO. Faeces excretion times varied between ingested diets that passed through the juvenile gut in less than 1 h. The cell walls of five of the eight micro-algae tested were partially or mostly digested (Chaetoceros muelleri, TW1200®, Palova 1800®, Isochrysis 1800® and Shellfish 1800®), while the cell walls of Tetraselmis 3600® and 3H 1800® remained intact. Juvenile growth rates were significantly different between diet treatments over the duration of a 14 day growth trial. Mean (± SE) length of early juveniles at the end of the growth trial was highest for those fed live C. muelleri (4.10 ± 0.03 mm) followed by TW 1200® (3.49 ± 0.05 mm). Juvenile survival did not differ significantly between diet treatments and was highest for those fed C. muelleri (79.33 ± 6.11%) followed by TW1200® (78.33 ± 1.20%). Pearson's correlation tests were used to identify key correlations between the levels of specific nutrients and juvenile performance (growth and survival). A significant positive correlation between growth and dietary protein content (P < 0.05), and a highly significant positive correlation between growth and dietary EPA:DHA ratio (P < 0.01) provide new information to inform diet selection for juvenile sandfish that will help improve juvenile performance and support improved hatchery production. Use of commercially available micro-algae concentrates as a replacement for live micro-algae in sandfish hatcheries supports development of cheaper, simpler post settlement rearing protocols for this species.

Knowledge of ingestion and digestion of micro-algae by bivalve larvae is critical for provision of appropriate larval nutrition supporting maximal growth and survival. However, little is known about the ingestion and digestion of micro-algae by giant clam larvae. This study determined the rates of ingestion and digestion of commercially available micro-algae concentrates by Tridacna noae larvae of different ages using epifluorescence microscopy. The micro-algae used were Isochrysis sp. (Isochrysis 1800®), Pavlova sp. (Pavlova 1800®), Tetraselmis sp. (Tetraselmis 3600®) and Thalassiosira weissflogii (TW 1200®). None of the four micro-algal concentrates were ingested by T. noae larvae at 24 h post-fertilisation, but all were ingested at 48 h and 72 h post-fertilisation, at different frequencies. At 48 h post-fertilisation, Isochrysis sp. and Pavlova sp. were ingested by 77% and 70% of veligers, respectively, while T. weissflogii and Tetraselmis sp. were ingested by 10% and 30% of veligers, respectively. Similar rates of ingestion were observed for each micro-alga by larvae at 72 h post-fertilisation. Larvae capable of ingesting micro-algae concentrates were significantly larger than those that were empty and the minimum antero-posterior shell length of T. noae larvae capable of ingesting Pavlova sp. and Isochrysis sp. was 141 μm and 132 μm, respectively. Digestion of micro-algae by 48 h-veligers was observed 2 h after the start of feeding in 26.1% and 14.3% of larvae that had ingested Isochrysis sp. and Pavlova sp., respectively, but digestion of Tetraselmis sp. and T. weissflogii was not observed until 4 h and 8 h after the start of feeding, respectively. Complete digestion of Pavlova sp. and Isochrysis sp. took up to 12 h in larvae at both 48 h and 72 h post-fertilisation. Our results provide a basis for developing a more nutritionally informed approach to hatchery culture of T. noae.

Veliger larvae of giant clams are demonstrable planktotrophs and provision of a particulate food source is generally accepted to improve hatchery production. However, the appropriateness of continuing to administer particulate food after larvae settle and develop as pediveligers is unclear. This study evaluated whether larval (age and size) and microalgal (species and cell density) factors influenced the capacity for pediveligers of Tridacna noae to ingest and digest cells from commercially available microalgae concentrates. Pediveligers at 6 and 10-days post-fertilisation were offered one of four microalgae species (Isochrysis sp., Pavlova sp., Tetraselmis sp., Thalassiosira weissflogii) at two cell densities (4000 and 8000 cells mL−1) and subsequently examined using epifluorescence microscopy. The appropriateness of continuing to administer microalgae as a food source was found to diminish as pediveligers approached metamorphosis, with negligible ingestion occurring at 10-days post-fertilisation. Earlier in development, at 6-days post-fertilisation, ingestion was significantly influenced by larval size, microalgae species, and microalgae cell density. Considering capacity for both ingestion and digestion, the Isochrysis sp. and Tetraselmis sp. microalgae concentrates had the greatest potential as a particulate food source for pediveligers of T. noae. The results of this study provide a basis for further refinement of giant clam hatchery procedures utilising microalgae concentrates as a larval food source.

This study evaluated the survival, growth, and larval quality of the spotted rose snapper L. guttatus during a critical developmental period when mass mortalities often occur (at 15 to 24 days post-hatch, DPH), from eating live food to beginning of formulated feed consumption. At first feeding, a large batch of snapper larvae was fed a mixture of the microalgae Tetraselmis and Isochrysis, the rotifer B. plicatilis, and the copepod P. crassirostris. Thereafter (15 DPH), a total of 1800 larvae (5.3 ± 1.3 mm total length) were randomly selected from the large batch and assigned to three dietary treatment groups in 300 L triplicate tanks at a density of 200 larvae tank−1: Group 1 was fed the copepod Cyclopina sp. (CY) only, but they (the copepods) were an assortment of life stages and sizes. Group 2 was fed a mixture of both copepods (CY) and enriched Artemia nauplii (Mx), and Group 3 was fed only enriched Artemia sp. nauplii (AR). Larval quality was evaluated as the tolerance of 24 DPH larvae to a short-term (1.5 min) air exposure stress test. The percent survival of larvae was significantly (p 0.05), were observed in lengths measurements of the larvae fed the different diets. The higher survival estimates found in our studies were attributed primarily to the prominent jerky swimming movement of the copepods, and the wide variety of sizes in the copepod culture available for the snapper larvae to prey on, thus increasing capture of the copepods and reducing cannibalism.

6 pages, 1 table, 2 figures, We aimed to evaluate three defined monoalgal diets, Isochrysis galbana clone T-ISO, Tetraselmis suecica and Rhodomonas lens, as a food source for Mytilus galloprovincialis to ascertain which of the diets maximized the feeding, digestion and the assimilatory balance of nutrients and energy. Mussels fed with Rhodomonas yielded the highest clearance and ingestion rates (CR and IR), suggesting that the dimensions of Tetraselmis and T-ISO might have restricted their capture by the mussels’ gill. Absorption efficiency (AE), an indicator of digestibility, was significantly higher for Rhodomonas (69.6%) than for Tetraselmis (38.4%) or T-ISO (23.6%) diets. This could be explained by the greater proportion of refractive non-digestible material contained in Tetraselmis and T-ISO diets, together with the low digestibility of the cell wall of Tetraselmis. The Rhodomonas diet showed the highest protein content, which was reflected in the highest ingestion and absorption of proteins compared with the other diets. However, the amount of carbohydrates and lipids ingested did not match the amount absorbed, probably owing to inefficient carbohydrate digestion and lipids lost through metabolic fecal losses. The total energy absorbed was higher for Rhodomonas (34.5 Jh− 1) than for T-ISO (20.1 J h− 1) or Tetraselmis (13.9 J h− 1) diet. The optimal feeding and digestive behaviour obtained for mussels fed with Rhodomonas diet, coupled with its ideal size, volume, weight and biochemical composition, might provide a better coverage for the anabolic demands of proteins during the seasonal growth cycle, especially during shell formation, gametogenesis and byssogenesis. Statement of relevance We state that this manuscript is relevant for the field of bivalve aquaculture and hatchery. We recommend that the main criteria in selecting algae for culturing mussels should be based on choosing the suitable dimension (size, volume, weight) of algal cell depending on the feeding capacity of mussels, which might vary between species and life stages (larvae, adult). The second main criteria should be based on selecting algae with high digestibility and balanced biochemical composition, to obtain a high absorption of energy that covers the nutritional requirements of each life stage of the mussels., This work was funded by the project FIGEBIV (AGL2013-49144-C3-2-R). Jade Irisarri is supported by JAE-Predoc CSIC-FSE 2012–2015 scholarship.

We investigated the effect of the stimulation of bacterial growth on Artemia performance in combination with a standard and with a low algal feeding regime. In both regimes, organic carbon (supplied as sucrose or soluble potato starch) and 15N labeled inorganic nitrogen (supplied as NaNO3) were used to stimulate bacterial growth in the Artemia cultures at C/N ratio 10 and 50. After a culture period of 15 days, significantly improved biomass production was obtained in all treatments with the low algae feeding regime, supplemented by carbohydrate addition. In addition, results of 15N accumulation and fatty acid analysis in Artemia indicated that Artemia utilized more bacteria in algae-limited conditions. Our study shows that bacteria can be used as a nutrient source for Artemia compensating for suboptimal algae supply. In Artemia pond cultures, carbohydrate addition may hence potentially be used to stimulate the conversion of nitrogen waste into heterotrophic bacterial biomass. This can be converted into protein-rich Artemia biomass, especially when algae are in sub-optimal supply. These findings open perspectives for alternative Artemia pond production protocols, in addition to the present management procedures that exclusively focus on phytoplankton blooms as nutrient source to sustain dense Artemia populations.

article i nfo This study investigated the growth, survival and fatty acid composition of sea urchin Paracentrotus lividus lar- vae fed four microalgal diets: Cricosphaera elongata, Pleurochrysis carterae, Tetraselmis suecica and Dunaliella tertiolecta (control). Larvae were successfully raised to competence for metamorphosis when fed C. elongata, P. carterae and D. tertiolecta diets but significant differences were found in survival rate and development. Larvae fed C. elongata showed 3 times higher survival and 20% faster development than larvae fed the other two microalgae diets that supported development. In contrast, T. suecica failed to fully support devel- opment and larvae stalled at the four arms stage for more than 30 days. The urchin larvae could accumulate long-chain polyunsaturated fatty acids (LC-PUFA) such as docosahexaenoate (DHA; 22:6n-3), eicosapen- taenoate (EPA; 20:5n-3) and arachidonate (ARA; 20:4n-6), either by assimilation and retention of dietary fatty acids, and/or synthesis from α-linolenic acid 18:3n-3 and linoleic acid 18:2n-6. Moreover, an accumu- lation of n-3 LC-PUFA and higher EPA/DHA and EPA/ARA ratios appeared to be associated with improved lar- val performance. The results indicate that live microalgae species, with appropriate fatty acid profiles are able to improve P. lividus larval performance, ultimately increasing hatchery profitability.

The marine prasinophyte Tetraselmis suecica was grown in a chemostat culture system over a series of phosphate-limited growth rates ranging from 0.164 to 0.755 d − 1 . The relationship between phosphate concentration and growth rate was well described by the Monod equation with a half-saturation constant of 3.45 nM. The kinetics of short-term uptake of phosphate spikes were well described by the Michaelis–Menten equation, maximum uptake rates being 2–3 orders of magnitude higher than associated growth rates. The relationship between cellular carbon-to-phosphorus ratios and growth rates was consistent with the Droop formulation of nutrient-limited growth, and chlorophyll a /carbon ratios were linearly related to growth rate. The protein content of the cells inferred from calculated nitrogen-to-carbon ratios ranged from a minimum of ~ 7% at zero growth rate to a maximum of ~ 59% at a relative growth rate of 1.0. Control of the relative growth rate of T. suecica via selection of the phosphate concentration in the growth medium and dilution rate of the culture appears to be an effective and practical way to manipulate the biochemical composition of this species in order to maximize its nutritional value to invertebrate consumers in aquaculture systems.

A series of experiments was conducted to evaluate the suitability of ten microalgal diets, including 4 monoalgal, 5 binary and 1 tri-algal diet, for the culture of the tropical paracalanid copepod Bestiolina similis. The four monoalgal diets were the Tahitian strain of Isochrysis sp. (T-Iso), Pavlova 50 (Pav), Tetraselmis chuii (Tet) and the diatom Chaetoceros muelleri (Chaet), the 5 binary diets were Tet+T-Iso, Tet+Pav, T-Iso+Pav, Chaet+Tet and Chaet+Pav while the tri-algal diet was T-Iso+Tet+Pav. After feeding B. similis with 10 algal diets for 3 days, 24 h egg production rate (EPR, eggs female−1 day−1) was obtained for each diet treatment by averaging the egg output of 6 individual females (replaced daily) for 4 consecutive days. The tri-algal diet T-Iso+Tet+Pav produced the highest EPR (44.1 eggs female−1 day−1), which was significantly higher than any other diets tested (p

The microalgae species Chlorella minutissima and Tetraselmis chuii were bioencapsulated in Artemia metanauplii and used during the weaning of Senegalese sole, Solea senegalensis post-larvae. We evaluated the effect of microalgae on: (i) survival and growth of the Senegalese sole post-larvae and juveniles, (ii) expression of genes related to the non-specific immune system, and (iii) the microbial load of sole juveniles. The survival of sole juveniles at the end of the experiment was significantly higher in the treatments added microalgae in their diet compared with the control treatment ( P P Vibrio as determined by counts on TCBS. Four partial nucleotide sequences of genes related to the immune system were isolated from lymphoid tissues of S. senegalensis : natural resistance associated macrophage protein 1 (Nramp1), complement C3, transforming growth factor β1 (TGF-β1), and transferrin. The expression of these four genes was calculated at the end of the experiment in relation to the expression of β-actin , a housekeeping gene. No significant differences ( P > 0.05) were determined among the three treatments in relation to the expression of the four genes in homogenates of fish.

Identifying appropriate larval husbandry is a key to improve larval quality and shorten duration of larval stages, and culture costs of ornamental cleaner shrimps Lysmata amboinensis. Several feeding and zootechnical experiments were conducted under laboratory conditions to test the effect of food during the first day posthatch, the effect of feeding with microalgae on larval digestion after 24 h posthatch, the effect of enriched rotifers upon survival and the combined effect of stocking density and food concentration on growth and survival of young larvae. For the first time, oxygen consumption values, trypsin-like activity and metabolite content (protein and triacylglycerols) were determined for L. amboinensis early stage larvae. When starved during the first day of life, L. amboinensis larvae had the same oxygen consumption as fed larvae, indicating that food ingestion is not crucial during that period. Nevertheless, energy reserves such as TAG were significantly lower in 24 h starved larvae when compared with fed larvae indicating facultative primary lecitotrophy. Trypsin-like activity of digestive enzymes (U mg prot− 1) was low when compared with other decapod larvae. Larvae fed with Tetraselmis chuii showed a significant increase in enzyme activity after 24 h. Present results showed that enriched rotifers result in higher larval survival during the first days of life when compared with larvae fed with non-enriched rotifers and that survival is not dependent on the relation between larval density and food concentration. In addition, stocking densities of 10 larvae ml− 1 showed higher survival compared to that obtained at the stocking density of 20 larvae ml− 1.

The tropical calanoid copepod, Acartia sinjiensis, has good potential for mass culture as a live feed for tropical reef fish larvae. This study was carried out to investigate the effects of temperature and salinity on population growth and egg hatching rates of A. sinjiensis. At a salinity of 30 ± 1 psu, temperature effects on population growth of A. sinjiensis was determined after 8 day culture at eight temperatures of 10, 15, 20, 25, 28, 30, 34 and 38 °C. Adopting similar experimental protocols, effects of salinity on population growth of A. sinjiensis were investigated under nine salinities of 10, 15, 20, 25, 30, 35, 40, 45 and 50 psu, with culture temperature set at 30 ± 1 °C. In addition to population growth experiments, egg hatching rates (%) after 48 h of incubation at various temperature and salinity conditions were also examined. For all experiments, five replicates were set up for each treatment. A. sinjiensis were fed an identical binary algal diet of Tetraselmis chuii and the Tahitian strain of Isochrysis sp. (T-ISO), photoperiod was maintained at 12 L:12D. The results showed that temperature had significant effects on both population growth and egg hatching rates. Population growth at 30 °C (with an initial number of 10 adults/replicate, final mean population reached 677.2 ± 136.6) was significantly higher (p The results of this study suggest that for maximum population growth and egg hatching success, A. sinjiensis should be cultured at 30 °C with a salinity of 30 psu.

In aquaculture, large volumes of phytoplankton are often grown outdoors to reduce costs. However, growing microalgae in an environment not as well-controlled as in the laboratory can lead to unwanted phytoplankton, including cyanobacteria, contaminating a culture. A cyanobacterial contaminant was isolated from an outdoor culture of Tetraselmis chui (PLY429) at the Milford Laboratory. This study investigated the growth of PLY429 and the cyanobacterium in pure cultures and a mixed culture in a pH range of 6.5–9.5. The division of PLY429 was greater at a pH range of 7.0–8.0; whereas, for the cyanobacterium, higher growth was obtained at pH 8.0–9.0. Results from combined cultures of PLY429 and the cyanobacterium grown at various pHs indicated that maintaining pH near 7.1 yields higher growth of PLY429 than those of the cyanobacterium. These findings suggest that controlling pH may reduce the population of a cyanobacterial contaminant in an aquaculture feed culture.

As natural diets of fish larvae, a number of calanoid copepod species are being investigated for use as live prey in aquaculture hatcheries. One of these, the tropical calanoid copepod, Acartia sinjiensis, has good potential as a live feed for tropical reef fish larvae. However, the rearing techniques for A. sinjiensis require further development to improve productivity. This study was carried out to investigate the population growth and egg hatching success of A. sinjiensis when fed a range of mono-and binary algal diets, including algae in the form of frozen paste. For the population growth experiment, the final A. sinjiensis population, including eggs, nauplii, copepodites and adults, was determined after feeding eight algal diets (two frozen algae, four live monoalgal and two live binary algal diets) for 8 days at temperature 28 ± 1 °C; salinity 34 ± 1 psu and photoperiod 12 L:12 D. Five replicates, with an initial 12 adult A. sinjiensis per replicate, were set up for each treatment. In a separate experiment, effects of diets on egg hatching success were examined after 48 h incubation of eggs produced by A. sinjiensis fed the same eight diets. The results showed that diet significantly affected both population growth and hatching success of A. sinjiensis. Of the diets tested, the binary algal diets were more successful than monoalgal diets, while the frozen algae had little dietary value. The highest population growth was recorded on A. sinjiensis fed a binary diet of Tetraselmis chuii and the Tahitian strain of Isochrysis sp. (T-ISO) (final population: 1091 ± 80), which was significantly higher (P < 0.001) than all other diets tested except for the other binary diet of Nannochloropsis sp. and T-ISO (final population: 897 ± 123). Diet also had a significant effect (P < 0.001) on egg hatching rate, though the highest hatch rate was recorded with eggs produced by A. sinjiensis fed binary diet Nannochloropsis sp. and T-ISO (88.1 ± 2.1%), this was not significantly different from that of eggs produced by A. sinjiensis fed either T-ISO alone (88.0 ± 1.7%) or the binary algal diet of T. chuii and T-ISO (76.4 ± 7.1%). The results of this study suggest that among the diets tested, a combination of live T. chuii and T-ISO was the best for the culture of A. sinjiensis.

The Caribbean scallops Argopecten nucleus and Nodipecten nodosus are currently being placed into mass culture in Colombia. The limited availability of wild seed upon which to base these cultures has promoted research into the development of artificial production of this seed in hatcheries. In support of this effort, we studied the effects of different diets on the physiology of the two scallop species in order to determine the optimal feeding regimes for maintenance of adult specimens in the laboratory. Seven monoalgal diets were tested, including Chaetoceros calcitrans , Chaetoceros muelleri , Isochrysis galbana (Ig), Nannochloris oculata (No), Phaeodactylum tricornutum (Pt), Tetraselmis chui (Tc) and Tetraselmis tetrahele (Tt). Four mixed diets were also tested, including I. galbana + C. calcitrans (Ig + Cc), I. galbana + N. oculata (Ig + No), I. galbana + T. tetrahele (Ig + Tt), and I. galbana + lipid emulsion of docosahexanoic acid DHA (Ig + lip). All the dietary trials were carried out under uniform conditions of temperature (25 °C), salinity (36‰) and algal concentration (0.45 mg L − 1 ). Physiological variables measured in association with each diet included feeding rates (clearance, ingestion and absorption), oxygen consumption and ammonium excretion rates as well as their scope for growth. The results showed that the best scope for growth for both scallops was obtained with diet Ig since this diet induced the highest feeding rates, accompanied by the lowest oxygen consumption and ammonium excretion. The feeding rates and scope for growth of A. nucleus were greater than those of N. nodosus for the majority of the diets, which was attributed to a higher rate of water pumping by the former species. Greater capacity for branchial food retention by A. nucleus was discarded as a possibility since N. nodosus had a greater branchial surface area per unit dry weight than A. nucleus . In spite of these differences, the oxygen consumption and the excretion rates were similar between the two scallops which suggested that A. nucleus was more efficient in its use of oxygen and retention of body proteins for physiological functioning. Mixed diets or addition of DHA did not permit increases in scope for growth in either of the scallops over that observed using monoalgal diet Ig , which suggest that biologically and economically this diet is optimal for the feeding of adult scallops in the laboratory.

The microbial conditions and antimicrobial activity were determined in batch cultures of two microalgae species, Tetraselmis chuii and Chlorella minutissima . The number of bacteria associated with the microalgae cultures showed an exponential growth 2, 10, and 16 days after inoculation, and they were higher in T. chuii in all three sampling points compared with C. minutissima . No presumptive Vibrio strains were observed in any of the samples, as measured by the growth on TCBS agar. A total of 17 and 30 bacterial strains were isolated from C. minutissima and T. chuii , respectively. A high percentage of Gram-positive strains was detected among the bacterial strains isolated, as Gram-positive strains constituted 82% (14 / 17) and 73% (22 / 30) of the total numbers of isolates in C. minutissima and T. chuii , respectively. The isolated bacteria were screened in vitro for inhibition against two pathogenic strains, and nine of the 34 strains tested (26%) showed inhibition in vitro against either Photobacterium damselae , susp. piscicida or Vibrio anguillarum . Incubation of enriched Artemia in cultures of the two microalgae for 30 min resulted in a significant decrease of the bacterial load in Artemia ( P , and a significant decrease of the level of presumptive Vibrio in Artemia homogenates ( P Vibrio among the bacteria associated with enriched Artemia .

Of the calanoid copepods, species of the genus Acartia are most commonly used for larval diets of tropical marine finfish. It is sourced from adventitious harvesting of wild populations or less frequently, cultured specifically as a live feed. However, little work is published on optimising diets for the aquaculture of copepods. In this study, A. sinjiensis was cultured on a range of mono- and binary algal diets and the effect on development assessed. There were significant differences in the rate of copepod development between the mono-algal diets. Rhodomonas was confirmed as an excellent algal diet for Acartia , but it is often unpredictable in mass culture. The cryptophyte, Cryptomonad sp. (CS-412) was shown to support an equally rapid development rate with the advantage of being more stable in mass culture. The algal feed concentration for maximal copepod development rate was dependent on the algal feed species. For the cryptophytes Rhodomonas sp. or Cryptomonad sp. , 1.13 μg AFDW ml − 1 was the minimal feed concentration required to support maximal copepod development at 28 °C. Mono-algal diets of the prymnesiophyte Isochrysis sp. (T.ISO) or Pavlova salina supported a lower level of copepod development than an equivalent biomass of either cryptophyte. Also, a higher feed concentration was required to achieve these lower development rates: 2.02 and 1.86 μg AFDW ml − 1 for Isochrysis sp. (T.ISO) and P. salina , respectively. Copepod nauplii fed the prasinophyte Tetraselmis sp., developed as fast as those fed an equal biomass of cryptophyte. However, development did not proceed beyond copepodite stage C1. Although survival was high, almost all of these C1 copepodites exhibited signs of a nutritional irregularity. The eyespot was frequently completely absent, greatly reduced in size or fragmented and setae were twisted. For rapid development, a mono-algal diet of Cryptomonad sp. could be used for the culture of A. sinjiensis . However, inclusion of a lipid-rich prymnesiophyte is likely to be beneficial to maximise egg production, a lipid dependent process.

Light intensity, day length, and nutrient concentrations are important factors regulating the growth of phytoplankton. To reduce culturing costs, natural sunlight and greenhouses can be used to minimize the amount of artificial light needed for algal growth. However, with natural sunlight there is much more variation in the light intensity and the day length than what would be found in a controlled laboratory environment. This study investigated how different light intensities and day lengths affect the growth and nutrient uptake of Tetraselmis chui (strain PLY429)—an algal strain used widely as an aquaculture feed. PLY429 was grown aseptically for 28 days under three different light intensities (220, 110, and 73 μEinst. m −2 s −1 ) and four different light:dark cycles (24:0, 16:8; 12:12; 8:16). Growth and net nutrient-uptake rates for PLY429 were calculated for each treatment. Longer day length and higher light intensities resulted in higher biomass production and complete utilization of nitrate and phosphate in less time, as compared with shorter days and lower intensities. PLY429 cultures that were exposed to only 8 h of light had the slowest growth and utilization of nutrients. These findings suggest that day length is important in determining growth and nutrient uptake in PLY429; at a latitude of 41°N, artificial light will need to be added to algal cultures in a greenhouse to increase both day length and total daily light input.

In an attempt to provide cost-effective alternative diets and to simplify hatchery–nursery procedures, substitutes for live microalgae in mariculture were tested and among them, concentrated microalgae have so far appeared to be the best alternative products for bivalves, though there are few experimental studies. The food quality of fresh and preserved Tetraselmis suecica for umboned Crassostrea gigas larvae was studied here and its potential as an algal feed for remote production was evaluated. Because it leads to lower growth and higher mortalities than the control diets (>60%), the use of T. suecica OR (strain Orbetello) for C. gigas larvae is not recommended as a monospecific diet. However, a binary diet that includes T. suecica OR is as efficient as the standard ternary PTC diet for C. gigas larvae. On the other hand, even though microalgal utilisation was not clearly affected by the initial size of the larvae, it seemed that old umboned larvae took full advantage of T. suecica OR. Its usefulness in C. gigas larval rearing is therefore limited, but its potential for oyster spat feeding has to be considered. Lastly, preservation at low temperature is an efficient method to maintain the food value of T. suecica OR.

The accumulation of bacteria in Brachionus plicatilis and Artemia franciscana during a short-term incubation was quantified using immunocolony blot (ICB) and an enzyme-linked immunosorbent assay (ELISA). Four bacterial strains, isolated from turbot and halibut, were grazed effectively by both species when given at high concentrations (≥5×10 7 bacteria ml −1 ). B. plicatilis accumulated 21–63×10 3 bacteria per rotifer and A. franciscana up to 45×10 3 bacteria per metanauplius after 20–60 min of grazing. The composition of the bacterial microflora of the live food organisms changed drastically, as the bioencapsulated strains comprised up to 100% of the total count of colony-forming units. After incubation in the bacterial suspensions, B. plicatilis and A. franciscana were transferred to seawater with added microalgae ( Tetraselmis sp., 2 mg C l −1 ), to evaluate the persistence of the changed bacterial composition in conditions similar to those present in a first feeding tank. The bioencapsulated bacteria decreased in numbers, but in most cases remained present in both live food organisms after 24 h. It is possible, after a short-term incubation, to replace opportunistic (r-selected) bacteria present in the live food cultures with other bacteria, which persist as a dominant part of the bacterial flora of the live food for a relatively long period of time (4–24 h).

The copepod Pseudodiaptomus euryhalinus was fed 320 cells μL− 1 of monoalgal cultures of Chaetoceros muelleri, Nannochloropsis oculata, Isochrysis galbana, Tetraselmis suecica, or a commercial frozen concentrate of Tetraselmis sp., and the diet which gave the best production was compared in a second experiment to three mixed diets: C. muelleri:I. galbana supplied in 1:1 and 2:1 cell ratios and C. muelleri:I. galbana:frozen Tetraselmis sp. in 2:2:1 ratio. These gave better results than the cultures of N. oculata, I. galbana, T. suecica and the frozen Tetraselmis concentrate, but the production was similar to that obtained with C. muelleri supplied as a monoalgal diet, showing that the addition of C. muelleri may improve the performance of other monoalgal diets, whereas the addition of other microalgae is unlikely to improve the results obtained when C. muelleri is supplied as a monoalgal diet.

Calanoid copepods of the genus Acartia have proven to be important in the diet of first feeding larvae of golden snapper, Lutjanus johnii , and mangrove jack, Lutjanus argentimaculatus , but it is difficult to reliably harvest these copepods from the wild. To overcome this problem, we present a simple method for the mass culture of Acartia spp. in the hatchery. This reduces the reliance on wild copepod populations and improves the coordination of larval rearing operations. Cultures of copepods were maintained in 100 and 1000 l tanks and fed daily with a mixed ration of algae, Rhodomonas sp., Tetraselmis sp. and Isochrysis sp., at a total algal cell density of 20,000 cells/ml. The cultures were lightly aerated and the salinity and temperature were maintained between 30–34 ppt and 28–32°C, respectively. The cultures were screened every 8 days to harvest adults and late-stage copepodids and to reduce contamination by rotifers and other undesirable zooplankton. The harvested copepods were used to restart the cultures. Using this method, more than 1000 adult copepods and late-stage copepodids per litre were produced from the 8-day culture cycle. This method has maintained productive cultures for more than 6 months and has been used to produce several batches of L. johnii fingerlings with survival rates of up to 40% at 21 days post-hatch. Adult copepods harvested from the cultures were stocked directly into larval rearing tanks. The copepods continued to breed in the larval rearing tanks and the resulting blooms of nauplii (up to 1.0/ml) were fed on directly by the larvae, eliminating the need to handle the easily damaged copepod nauplii.

Thirteen species of tropical Australian microalgae (both benthic and planktonic) were compared in terms of survival and growth of Artemia and enrichment of polyunsaturated fatty acids (PUFAs), including eicosapenaenoic acid (EPA; 20:5n−3) and docosahexaenoic acid (DHA; 22:6n−3) in Artemia. Artemia nauplii showed enhanced growth beyond the unfed control value after 24 h feeding with all species of microalgae except Isochrysis NT14. However, when the growth was compared against the commercial northern hemisphere species Isochrysis galbana TISO, only Artemia fed Cryptomonas CRFI01 showed better growth (significance at P

The genus Strombus is widely distributed in the Caribbean. Six species are of commercial importance: S. gigas, S. raninus, S. costatus, S. alatus, S. gallus and S. pugilis. Economically, the Queen conch, S. gigas is the most important and consequently the most widely studied. However, since 1970 a decline of S. gigas populations due to over-fishing has been observed. Many authors have studied S. gigas hatchery rearing techniques in order to address this problem; however, for these hatchery techniques to be successful, an adequate diet must be provided for the larvae. Some information of the nutritional requirements of S. gigas larvae have been reported since 1965, but a nutritionally complete diet is still not available. In this work we summarize the different diets that have been used for S. gigas, S. costatus and S. pugilis larvae rearing. Twenty one different algae species have been used: Amphidinium carteri, Chaetoceros gracilis, Dunaliella tertiolecta, Emillania huxleyi, Heterocapsa pygmacea, Isochrysis (Caicos), Isochrysis (Tahiti), Isochrysis sp., Monochrysis sp., Nannochloris, Nitzchia, Platymonas sp., P. tetraselmis, Prorocentrum minimun, Rhodomonas sp., Skeletonema costatus, Tetraselmis chuii, Tetraselmis sp., T. suecica, Thalassiosira fluviatilis and T. weissflogii. There are other diets that have seldom been studied with Strombus veliger larvae, that could be a potential food source for these gastropods. The type and concentration of algae, larval rearing conditions are summarized along with the results attained in larval growth, metamorphosis, survival, ingestion and digestion rates.

Four Australian microalgal isolates ( Pavlova pinguis , Rhodomonas salina , Tetraselmis sp. CS-362 and Nannochloropsis -like sp. CS-246) were compared to an overseas isolate ( Isochrysis sp. (T.ISO)) to assess their potential as food for bivalve aquaculture. When fed to Pacific oyster spat, the best growth was observed with a mixed diet comprising the four Australian species plus Isochrysis sp. (T.ISO) (161% increase in organic weight over 21 days). The best growth using the monospecific diets was produced by P. pinguis (111% increase), followed by Isochrysis sp. (T.ISO) and R. salina (85% and 82%); then Tetraselmis sp. CS-362 (51%), and Nannochloropsis -like sp. CS-246 (35%). The growth rates (based on an increase in organic weight) of oysters fed the monospecific diets were correlated to the amounts of protein and carbohydrate they ingested. The gross biochemical composition of the microalgae grown in semicontinuous culture differed significantly. Protein ranged from 48% of the dry weight ( R. salina ) to 17% ( Nannochloropsis -like sp. CS-246); carbohydrate ranged from 41% ( P. pinguis ) to 22% ( R. salina ); and lipid from 16% ( Tetraselmis sp. CS-362) to 26% ( Nannochloropsis -like sp. CS-246). The instantaneous growth rate ( μ ) of Isochrysis sp. (T.ISO) ( μ =0.48 day −1 ) was greater than the Australian strains, the best of which was R. salina ( μ =0.37).

This paper reports on two experiments in which three species of tropical microalgae, Isochrysis (T-ISO), Pavlova salina and Chaetoceros simplex, dried Tetraselmis suecica and a commercial yeast-based diet (`L-10', Microfeast®) were assessed for their nutritional value for larvae of the blacklip pearl oyster, Pinctada margaritifera. In the first experiment, T-ISO supported significantly greater growth of D-stage larvae than either Pav. salina, C. simplex or a ternary algal diet (TAD) composed of an equal mixture of the three species. Fifty percent of the mixed algal diet could be replaced with dried Tetraselmis without significantly affecting larval growth rate or survival. Indeed, growth of larvae fed a 1:1 mixture of TAD and dried Tetraselmis was significantly greater than that of larvae fed the three species of microalgae alone or the TAD. In the second experiment, umbo larvae were fed either the TAD, dried Tetraselmis, `L-10' or 1:1 combinations of these diets. Survival of unfed larvae was significantly lower than that of fed larvae and larvae receiving food had significantly greater antero-posterior shell length than unfed controls. Larvae fed dried Tetraselmis were significantly larger than those fed `L-10' but did not differ significantly in size from those fed the TAD. Larvae fed the 1:1 mixture of dried Tetraselmis and `L-10' were significantly larger than those fed `L-10' alone but significantly smaller than those fed dried Tetraselmis alone. The largest larvae at the end of Experiment 2 were those fed a 1:1 mixture of TAD and `L-10' which were significantly larger than larvae fed the TAD.

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

Nine microalgal strains from the prasinophyte genus Tetraselmis that were chosen for high total lipid content, and one marine strain of the chlorophyte Chlamydomonas , were compared with Isochrysis sp., strain T-ISO, as diets for oyster spat in a controlled laboratory feeding experiment. Five of these high-lipid Tetraselmis strains supported significantly faster oyster growth than an equivalent ration of T-ISO. Doubling times for oyster weight, volume, and shell height were in the range of 1.5–3 weeks on single daily feedings of the best diets. Tetraselmis strains yielding the most rapid oyster growth contained higher contents of the essential fatty acid 20:5 n − 3 and of the sterols 24-methylcholesterol and/or 24-methylenecholesterol. These data are consistent with our earlier findings that these compounds appear to be deficient in most phytoplankton for optimal growth of Crassostrea virginica spat.

14 páginas, 5 figuras, 2 tablas., The response of marine microalgae to different cryopreservation methods was described. Of the six species evaluated, only Chaetoceros gracilis depended on faster cooling rates to increase its postthaw viability (7.2% at 0.25 °C min−1, 29.3% at 4 °C min−1) when using 15% dimethyl sulphoxide in 36 p.p.t. salinity seawater. Tetraselmis chuii, Nannochloris atomus and Nannochloropsis gaditana were the most tolerant species to biological freezing, achieving mean viabilities of 97.9%, 80.5% and 61.6% respectively. Rhodomonas baltica and Isochrysis galbana, T-ISO strain, showed the lowest viability (means of 7.3% and 15.1% respectively) after cryopreservation under the same conditions of salinity, cryoprotectant concentration and cooling rates. Avoidance of undercooling by inducing ice nucleation when reaching the freezing point did not change viability in comparison to all the procedures that did not include seeding in any of the tested species. Five species showed similar viabilities when a single controlled cooling step to −50 °C procedure was compared to a two-step cooling process, in which algae were plunged into liquid nitrogen (LN) after the first step. Isochrysis galbana represented an exception. A mean viability of 25.8% was achieved when cooled to −50 °C, whereas viability decreased to 4.4% when the second cooling step to −196 °C was used. Replacing the special biological freezing equipment by a −20 °C freezer to perform the first cooling step resulted in a steady cooling rate after the commencement of ice formation. This was due to the fact that samples reached that temperature in a liquid state. Solidification occurred spontaneously at variable times once −20 °C was reached. A cooling rate of −14 °C min−1 during the change from liquid to solid state was achieved when a −80 °C freezer was used to perform the first cooling step. The performance of the first cooling step in both type of freezers resulted in similar viabilities after thawing from liquid nitrogen, in comparison to the use of special equipment for controlling cooling rates in T. chuii (90.8% for −20 °C and 89.7% for −80 °C), N. gaditana (44.6% for −20 °C and 42.6% for −80 °C) and N. atomus (85.9% for −20 °C and 85.6% for −80 °C). Lower viabilities were recorded for R. baltica and Ch. gracilis cooled to −20 °C and in LN (2.2% and 2.9% respectively) but no difference were found with respect to the first technique, when both species were cooled to −80 °C and in LN (6.3% and 19.9% respectively). I. galbana showed no viability when cooled to −80 °C., This work was supported by the Regional Agriculture and Fisheries Council (Junta de Andalucia) through a collaboration programme with the National Council Research (CSIC).

Ascorbic acid (AA) is an essential nutrient in particle and live aquafeeds. In order to better assess the needs for this nutrient during larviculture the AA content of algae, rotifers and Artemia was studied with respect to their suitability at startfeeding. In general, the microalgae evaluated were rich in AA (1000-4000 µgAA/gDW), but showed a considerable variability among the different species: e.g. the concentration in Isochrysis andChlorella reached values 3-to 4-fold the percentage of Tetraselmis (0.11% of DW). Brachionus routinely cultured on Chlorella contained 2300 µg AA/gDW. Cysts of various batches and strains of Artemia differed considerably in ascorbic acid-2-sulphate (AAS) concentration (296-517 µgAA/gDW). The amount of AA available in the freshly-hatched nauplii reflected exactly the AAS reserve present in the cysts, what evidences the complete conversion of AAS to free AA during completion of embryonic development into nauplii. Boosting techniques both for Brachionus and Artemia using ascorbyl palmitate (AP) as the vitamin C source were established. The addition of 20% AP in the diet of Brachionus enhanced their AA content 10-fold over 3 days of culture. Supplementation of the enrichment emulsion for Artemia with 20% AP increased the AA content up to 2000 µg/gDW after 24h enrichment. This lipophilic derivative of AA appeared to be a stable form of vitamin C for enhancing AA levels in the live diets during culture and/or enrichment. This bioencapsulated method provides a tool for hatcheries to build up high AA concentrations in the live prey administered to first feeding larvae of aquaculture organisms in case of specific requirements (e.g. with respect to handling stress, deformities). A survey of commercial hatcheries indicated that a wide range of products is used for the cultivation and boosting of rotifers, which consequently affect their AA levels. In general, the AA content in the algae and, consequently, the algal enrichment of Brachionus tended to score lower in the hatcheries than under lab conditions.

This paper is the first to present a basic feeding schedule for Penaeus setiferus larvae based on diatoms ( Chaetoceros ceratosporum ), flagellates ( Tetraselmis chuii ) and Artemia nauplii. Growth rate (μm/day), rate of metamorphosis and survival of larvae fed 5 different concentrations of diatoms, flagellates and Artemia nauplii were measured. Survival associated with sudden changes in salinity (quality index QI) of the postlarvae harvested from each treatment was measured. A performance index was calculated from the growth rate, survival and QI. Considering all responses, a feeding schedule for P. setiferus larvae is recommended including a progressive increase in the concentration of diatoms (between protozoea and mysis; 30 and 40 × 10 3 cells/ml, respectively), flagellates (between protozoea and mysis; 2 × 10 3 cells/ml, respectively) and Artemia nauplii (0.5, 1.0 and 1.5 between PZ 3 -M 1 , M 1 -M 2 and M 2 -M 3 , respectively). The increase in concentration of algae and Artemia indicates a pattern of development of the digestive system associated with development of the larvae. This pattern must be considered to improve the production of larvae of this species under controlled conditions.

13 páginas, 4 tablas., Lipid class and fatty acid contents, survival and nutritional status of Penaeus kerathurus larvae, reared on a mixture of the marine microalgae Tetraselmis chuii and Isochrysis galbana (clone T-ISO), the rotifer Brachionus plicatilis and Artemia (parthenogenetic strain from Cádiz, S.W. Spain), were studied throughout development. Dry weight and free sterol contents increased and correlated positively (r = 0.91, P < 0.05) during development, whereas the ratio of triacylglycerol/free sterol (an indicator of the nutritional status of the larvae) decreased as did survival. Fatty acid contents at different larval stages reflected fatty acid content of foods. The content of docosahexaenoic acid (22:6n - 3) remained constant during the mysis and first postlarval stage and a low bioconversion rate from its precursor, eicosapentaenoic acid (20:5n - 3) (very abundant in Artemia food) was observed., This work was supported by the Comisión Interministerial de Ciencia y Tecnología, Spain (CICYT projects MAR90-0767-C04-02 and AGF93-0173).

Epifluorescence microscopy was used to observe the ingestion and digestion processes of eight algae diets in 18-day-old Strombus gigas larvae. The eight diets used were: Isochrysis aff. galbana, Tetraselmis chuii, Tetraselmis suecica, Dunaliella tertiolecta, Chlamydomonas coccoides, Thalassiosira fluviatilis, Chlorella sp. and Chaetoceros sp. Observations were made at 0.5, 1, 2, 3, 4, 5, 6, 7, 8 and 24 h after feeding. Ingestion index was faster with T. chuii, Chaetoceros sp. and Chlorella sp. than with I. aff. galbana, D. tertiolecta, C. coccoides, T. fluviatilis and T. suecica . Digestion began at the second hour, being faster with Chaetoceros sp. and Chlorella sp. and slower with D. tertiolecta and T. fluviatilis . Values of ingestion and digestion indices were specific for each alga. This research was performed to provide helpful information in selecting the best diet for each larval stage, according to culture conditions.

The effect which microalgal addition and rotifer enrichment with algae had on survival, growth rate and fry viability during first-feeding were examined for turbot (Scophthalmus maximus L.). Addition of the microalgae Isochrysis galbana or Tetraselmis sp. together with rotifers Brachionus plicatilis, grown on yeast and oil emulsion, and Artemia greatly improved rearing success, whereas short-term enrichment of the rotifers with Tetraselmis sp. gave only improved viability. The algae modified the relative fatty acid composition of rotifers. Rotifers with I. galbana exhibited and increased level of 22:6n-3 and a lower level of 20:5n-3, whereas the opposite change occurred when Tetraselmis sp. was used. Addition of either of the two algal species at 1 mg cl−1 to the larval tanks resulted in a constant lipid level and high egg ratio of the rotifers, and thereby also high individual biomass content. Without algal addition, the lipid content of the rotifers decreased by 20% day−1 in the early phase, and their egg ratio became close to zero. In addition to improved nutritional conditions of the larvae, some other factor, e.g. some trigger mechanism or changed microbial or light conditions, also can operate to explain the increased early appetite of larvae with microalgae added and must be taken into consideration.

Commercially important members of the genus Tetraselmis were cryopreserved with high levels of post-thaw viability. Levels of post-thaw viability > 50% for T. suecica (Kylin) Butcher were achieved by employing a simple protocol using glycerol as a cryoprotectant; samples were placed in a coolingbath preset at −30°C for 30 min, followed by plunging into liquid nitrogen. This was found to be less satisfactory for T. chui Butcher. The highest levels of viability were obtained using cultures grown at 20°C, preincubated in 10% (v/v) glycerol, followed by a two-stage cooling process, using a cooling rate of 1°C min−1 to −30°C and then plunging the samples into liquid nitrogen. Viability levels were higher in all the strains tested when they were grown under ambient conditions (15–20°C) and were species-dependent, with T. suecica (Kylin) Butcher strains having higher levels of viability (> 70%) than T. chui Butcher (28–40%).

Atlantic halibut (Hippoglossus hippoglossus L.) were offered rotifers (Brachionus sp.) and algae (Tetraselmis sp.) at different ages, and feeding frequency and survival through a 48-h feeding period were measured The highest frequency of larvae with algae in the gut was found when larvae were first offered food between 44 and 55 days after hatching (mean temperature 4.5 °C). The highest frequency of larvae with rotifers in the gut was found when larvae were offered food between days 55 and 69. The highest number of rotifers in the gut was found in the groups offered food between days 55 and 59. Larvae offered food as late as day 75 died before sampling. The results indicate that larvae held at low temperature (4.5 °C) should be offered food between 44 and 59 days after hatching (200–265 d °).

A study was conducted to determine a suitable algal diet for the window-pane oyster, Placuna placenta Linnaeus, during gonad development. Sexually immature P. placenta (70 mm shell height) were fed Isochrysis galbana Parke, Tetraselmis tetrahele (G.S. West) Butch, or their 1:1 combination, at 100 000 cells per ml divided into two rations per day. Temperature and salinity ranged from 27 to 29 °C and 28 to 32 ppt, respectively. Monthly gonad histological examination showed a rapid gonad development in P. placenta fed the combination of I. galbana and T. tetrahele, attaining sexual maturity in the third month of rearing, a month ahead of those fed single algal species.

Epifluorescence microscopy was used to observe the ingestion and digestion of unicellular algae in gastropod larvae. Larvae started feeding 8 h after hatching. Three kinds of diet were tested during the period of this study: Isochrysis aff. galbana, Tetraselmis chuii and Isochrysis-Tetraselmis mixture. All the experiments were conducted at 29°C. It was shown that ingestion was faster with Tetraselmis than with Isochrysis. Within 10 min 43% of the larvae had ingested Tetraselmis and no larvae had ingested any Isochrysis. Within 30 min, 60% of the larvae had ingested Tetraselmis while only 25% had ingested Isochrysis. Using a scale based on the quality of fluorescence, it was possible to determine the time interval to digest a diet, which is shorter with Tetraselmis than with Isochrysis. After 8 h the digestion index reached 45% for larvae fed with Tetraselmis while it was only 28% for larvae fed with Isochrysis. Using a quantitative scale based on measurements of the fluorescent surface in each larva, it was demonstrated that the amount of diet ingested is higher with Tetraselmis than with Isochrysis. These results indicate that Tetraselmis chuii forms an adequate diet for young veligers of Strombus gigas.

Camacho, F., Mohna, E., Martinez, Ma. E., Sanchez, S. and Garcia, F., 1990. Continuous culture of the marine? microalga Tetraselmis sp. - productivity analysis. Aquaculture, 90: 75-84. An outdoor experimental system for continuous culture of the marine microalga Tetraselmis sp. is described. The influence of dilution rate, initial biomass concentration and the intensity of solar radiation on the amount of biomass obtained was examined. The average rate of production of cells, G,. is a parabolic function of the dilution rate and reached a maximum value at D=O.O9 h-‘. The intensity of light, I,, influences productivity up to approximately 100 W/m*, above which saturation was observed. The experimental results G,-I, were adjusted to an exponential model for light-restricted growth. The average rate of production of cells, versus the initial biomass concentration, C,, reached a maximum value. G,,, decreased as C,, increased to high cell density values due to light limitation and when C, decreased to low cell density values due to decreased efficiency in the use of incident solar radiation.

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 .

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.

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 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.

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.

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.