A comparison among differently enriched rotifers (Brachionus plicatilis) and their effect on Atlantic cod (Gadus morhua) larvae early growth, survival and lipid composition

We evaluated the effect of differently enriched rotifers on the early growth, survival and lipid composition of Atlantic cod larvae (Gadus morhua). The enrichments tested were: (i) AlgaMac 2000^®; (ii) AquaGrow^® Advantage; and (iii) a combination of Pavlova sp. paste and AlgaMac 2000^®. Larvae from treatment 3 [1.50 ± 0.11 mg dry weight (dw) and 7.10 ± 0.14 dw specific growth rate (SGR)] were heavier (P = 0.006) and grew faster (P = 0.004) than larvae from treatment 2 (1.03 ± 0.04 mg dw and 6.29 ± 0.04 dw SGR). No significant differences were found in the final weight and SGR among larvae from treatment 1 (1.21 ± 0.07 mg dw and 6.58 ± 0.20 dw SGR) and larvae from treatments 2 and 3. The treatment 3 also resulted in the best survival at the end of the experimental period, estimated to be 3 on a scale from 1 to 5, whereas the survival estimates for the two other groups were 1–2. Larvae from the treatment 3 reached 37 days posthatch with levels of ω6DPA 32‐fold higher than newly hatched larvae. Differences in the larval enrichment of ω6DPA may explain the differences in growth and survival of the Atlantic cod larvae.     

Manipulation of the fatty acid composition of phosphatidylcholine and phosphatidylethanolamine in rotifers Brachionus Nevada and Brachionus Cayman

Rotifer cultures of Brachionus Nevada and Cayman were grown semi‐continuously, fed live Rhodmonas baltica algae, and cultures were diluted daily (20% day^?1). The enrichment diets Marol E (triacylglycerol‐rich) and PL (cod roe emulsion, phospholipid‐rich) were added to both cultures (Nevada 40 ng ind^?1 day^?1 and Cayman 25 ng ind^?1 day^?1, respectively). After 4 days of enrichment (dilution rate: 20% day^?1), Cayman rotifers–fed Marol E had comparable (P > 0.05) quantitative amounts of DHA (docosahexaenoic acid, 22:6n‐3) to natural harvested copepods, which were analysed for comparison, and the Cayman strain had generally more efficient incorporation of HUFAs (highly unsaturated fatty acids) than the Nevada strain. However, the percentage DHA of the total fatty acids in copepods were much higher than in enriched rotifers (P < 0.05). The PC‐DHA (DHA in phosphatidylcholine) and PE‐DHA (DHA in phosphatidylethanolamine) levels of copepods were up to 50%, which was highly unlikely for rotifers to reach. The increase of DHA in total phospholipids of rotifers was mainly due to an increase of DHA in PC, whereas the PE‐DHA was only weakly affected by dietary lipids. The per cent PC‐DHA levels in rotifers were positively related to the total dietary DHA levels (P < 0.0001), independent of the PC‐DHA content in the diets.     

Use of differently enriched rotifers,Brachionus plicatilis,during larviculture of haddock,Melanogrammus aeglefinus: effects on early growth,survival and body lipid composition

We evaluated the effects of enriched rotifers on growth, survival and on the lipid composition of haddock larvae. The treatments tested were (1) AlgaMac 2000^®, (2) AquaGrow^® Advantage and (3) Pavlova sp. paste and AlgaMac 2000^®. The treatments did not influence larval growth rate throughout the experimental period (P = 0.70). Larvae from all treatments grew approximately 8% of their dry weight per day between 1 and 29 days post hatch (dph). Treatment 3 resulted in the best survival, estimated to be 3 on a scale from 0 to 5, whereas for the two other groups the survival estimates were 0 and 2. Rotifers from treatment 1 had low sterol concentrations, high eicosapentaenoic acid/arachidonic acid ratio and their feeding resulted in high larval mortality. Rotifers enriched with Pavlova sp. had the lowest proportions of the sum of saturated fatty acids, docosahexaenoic acid and sum of ω3 and the highest proportions of the sum of monounsaturated fatty acids (ΣMUFA). This was partially reflected in larvae from treatment 3 in that they had the highest proportions of ΣMUFA and the lowest proportions of Σω3 (P < 0.0001 for both analyses). In addition, these larvae had the highest and lowest ΣC₂₀ and ΣC₂₂ polyunsaturated fatty acids (PUFA) respectively (P < 0.0001 for both analyses). We suggest that more research with ω3 and ω6 PUFA can lead to improvements in the rearing of haddock larvae produced in hatcheries.     

Evaluating bioencapsulation of florfenicol in rotifers (Brachionus plicatilis)

The rotifer Brachionus plicatilis is one of the most important live‐feed organisms used in the larval culture of marine fish. Bioencapsulation of florfenicol in rotifers was investigated by delivering it directly to the organisms as particles and the doses ranged from 50 to 1000 mg L^?1. Analysis of the florfenicol concentrations in rotifers was performed using high‐performance liquid chromatography. The uptake increased with the dose and the enrichment time, and the highest concentration achieved in this study, 1.25 ng rotifer^?1, was obtained using a dose of 1000 mg L^?1 and 150 min of enrichment. The optimal time for satisfactory enrichment of rotifers with florfenicol was found to be dose dependent and ranged from 30 to 120 min for doses between 400 and 800 mg L^?1. The parameters affecting the concentrations achieved in rotifers were the size of the florfenicol particles, enrichment time and dose.     

Examination of a practical method for copper enrichment of euryhaline rotifers (Brachionus plicatilis) as diet of Eriocheir sinensis zoea larvae

In this study, an effective method to enrich the rotifer Brachionus plicatilis with copper was developed as a feed for the Chinese mitten crab Eriocheir sinensis zoea larvae. Changes in the concentrations of other minerals in rotifers were also examined when copper was added for rotifer enrichment. The ability of Chlorella to absorb waterborne copper is much higher than that of rotifers, and hence, copper was preaccumulated in Chlorella before its ingestion by rotifers. The copper content in rotifers was comparable to the dietary copper requirement of the crab larvae when the rotifers were enriched with 0.1 mg Cu g⁻¹ Chlorella for 12 h. Further enrichment in rotifers with Cu‐enriched Chlorella and lipid emulsions did not significantly change the profile of major fatty acids and mineral composition in the rotifers. Evidence shows the feasibility of copper enrichment in rotifers using microalgae that can accumulate copper. This study indicates that copper in rotifers can be enriched by feeding copper‐enriched algae at a concentration of 0.1–0.2 mg Cu g⁻¹ Chlorella. The developmental rates of E. sinensis can be improved by feeding zoea larvae with copper‐enriched rotifers, but survival rates were not affected by dietary copper enrichment.     

不同脂肪源对褶皱臂尾轮虫脂类和脂肪酸组成的影

以褶皱臂尾轮虫（Brachionus plicatilis)为实验动物并设计不同脂肪源饵料，分别为：面包酵母Saccharomyces cerevisice（对照组），微绿球藻Nannochlorpisis oculata(A)，5%大豆磷脂 5%鱼油 90%面包酵母（C）。结果表明：（1）接受不同脂肪源的轮虫的脂肪酸组成显著不同，尤其是高度不饱和脂肪酸（HUFA，20C）含量存在显著差异，说明饵料中HUFA含量对轮虫体内相应脂肪酸的含量有显著影响。投喂饵料C使轮虫的脂类HUFA水平得到了强化，提高了轮虫的营养价值。（2）轮虫脂类的HUFA水平不仅由饵料中脂类的相应脂肪酸组成决定，而且轮虫脂类HUFA的提高和强化效果与饵料中HUFA的化学形态密切相关。甘油三酯型饵料HUFA的强化轮虫效果高于磷脂型的HUFA。（3）将富含HUFA的鱼油直接添加在干性饵料如面包酵母中，而不是通过对鲁油乳化后直接强化轮虫，同样能取得良好的强化效果，且操作简便、不易污染水质，适用于轮虫的规模生产。     

The enrichment and retention of ascorbic acid in rotifers fed microalgal diets

The enrichment and retention of ascorbic acid (AA) was investigated in rotifers Brachionus plicatilis fed on microalgae ( Nannochloropsis oculata and Isochrysis sp. (T.ISO)) and baker's yeast Saccharomyces cerevisiae . The concentrations of AA of the rotifer diets used in the study differed significantly: 4200 μg g⁻¹ of dry weight in Isochrysis sp. (T.ISO), 2600 μg g⁻¹ in N. oculata and only 77 μg g⁻¹ in S. cerevisiae . Rotifers contained 620 μg AA g⁻¹ prior to the experimental feeding. When subsequently fed for 3 h on microalgae at a ration of 0.13 mg dry microalgae per 10⁶ rotifers rapidly and efficiently increased their content of AA: Isochrysis sp.-fed rotifers contained 1600 μg AA g⁻¹ and N. oculata -fed rotifers contained 1100 μg AA g⁻¹. Concentrations were boosted by a further feeding of a second ration of algae at three times the initial feeding ration; 21 h later, Isochrysis sp.-fed rotifers contained 2500 μg AA g⁻¹ and N. oculata -fed rotifers contained 1700 μg AA g⁻¹. This represented a 180% and 310% increase in the pre-feeding vitamin concentrations in Isochrysis sp. and N. oculata -fed rotifers, respectively. There were no significant changes in AA concentration in rotifers fed a similar ration of yeast throughout the feeding period (520-620 μg AA g⁻¹). Rotifers retained AA during a subsequent 24 h non-feeding period, with no significant changes in the concentrations in any of the rotifer groups. The production of rotifers rich in AA may be particularly valuable for the culture of fish larvae that have a high requirement for the vitamin.     

Biochemical composition and fatty acid profile in a strain of the lineage ‘Nevada’, belonging to the Brachionus plicatilis (Rotifera) complex,fed different diets

A strain of the lineage Brachionus ‘Nevada’ was batch cultured with two diets, differing in biochemical composition: baker's yeast (treatment 1), which has higher protein:lipid ratio compared with CULTURE SELCO (treatment 2). The biochemical composition (DNA, RNA, lipid, protein content) and fatty acid profile of rotifers of both treatments was analysed and related to previously published population structure data. CULTURE SELCO‐fed rotifers showed higher DNA, RNA, lipid, n‐3 highly unsaturated fatty acids (HUFA) average content, compared with yeast‐fed rotifers, which had higher protein content. Rotifer lipid content showed significant diurnal variation in yeast‐fed rotifers. Rotifer lipid and n‐3 HUFA content was associated with reproductive output. DNA and RNA content was related to embryonic development while protein content, to somatic growth and mixis. The saturated and monounsaturated fatty acid rotifer content was stable irrespective of feed, in contrast to eicosapentanoic acid (EPA) and DHA. The levels of AA were similar in both rotifer populations, but those of EPA and docosahexaenoic acid (DHA) were about half in yeast‐fed compared with CULTURE SELCO‐fed rotifers. CULTURE SELCO resulted in a temporally stable rotifer lipid profile and a better enriched parthenogenetic population.     

Examination of a practical method for zinc enrichment of euryhaline rotifers (Brachionus plicatilis)

Larval growth and survival of marine finfish in mass seed production are affected by the nutritional value of live feeds such as rotifers and Artemia. Thus far, many studies have been conducted to develop effective methods for the enrichment of live feeds with essential fatty acids and vitamins. In this study, a practical method for enrichment of rotifers with zinc was investigated. Changes in the concentrations of other minerals when zinc was added to the rotifer-enrichment tanks were also studied. The mineral composition of rotifers and Chlorella after zinc enrichment revealed that the direct addition of zinc to the culture media was not effective because rotifers cannot efficiently accumulate waterborne zinc. The ability of Chlorella to absorb waterborne zinc is much higher than that of rotifers, and hence, zinc was pre-accumulated in Chlorella, which was then fed to the rotifers. The maximum zinc content of the rotifers was 585.0 μg g^? 1 (dry matter) when the rotifers were enriched with zinc alone. This zinc concentration is comparable to that found in natural zooplankton. In rotifers simultaneously enriched with zinc and n?3 highly unsaturated fatty acids (HUFAs), the zinc content increased, but the n?3 HUFA content did not. Therefore, separate enrichment with zinc and fatty acids was adopted. The zinc content of rotifers fed zinc-enriched Chlorella was significantly higher than that of rotifers fed unenriched Chlorella. After zinc enrichment, rotifers were enriched with fatty acids, and the docosahexaenoic acid (DHA) and n?3 HUFA levels in rotifers were higher than the levels obtained after simultaneous enrichment with zinc and fatty acids. With regard to the concentration of other minerals in rotifers after zinc enrichment, the manganese content tended to decrease when the zinc content increased.The results of this study demonstrated that zinc enrichment of rotifers was successfully performed by using microalgae that had accumulated zinc, and the enrichment of rotifers with fatty acids was also achieved after the completion of zinc enrichment and before feeding the larvae. This method could be utilized for the enrichment of zooplankton with other minerals as well.     

Use of flubendazole as a therapeutic agent against rotifers (Brachionus plicatilis) in intensive cultures of the harpacticoid copepod Tisbe holothuriae

Copepods are well known to be the optimal live feed for most species of marine fish larvae. Still copepods are rarely used in marine hatcheries worldwide. Lack of efficient production techniques are among the reasons for this. Consequently, Artemia and rotifers are utilized in commercial settings. One problem in intensive production of copepods is contamination with rotifers. Rotifers have higher growth rates than copepods and consequently will compete out the copepods when accidentally introduced to the copepod production systems. Once contamination has occurred, the only cure has been to shut down production and subsequently use a therapeutic agent to eliminate all zooplankton in the system before restart with a stock culture free of rotifers. We tested flubendazole as a mean of controlling rotifers (Brachionus plicatilis) in intensive laboratory cultures of the harpacticoid copepod (Tisbe holothuria). Flubendazole was lethal to rotifers in concentrations as low as 0.05 mg L^?1. There was no significant effect on the concentration of copepods, even at the highest concentration tested, i.e. 5.0 mg L^?1 flubendazole. We conclude that flubendazole is an effective drug for control of B. plicatilis in T. holothuriae batch cultures.     

Semi‐continuous mass culture of rotifers (Brachionus plicatilis) using an automatic feeder

This study describes the rotifer mass culture protocol developed at the Institute of Aquaculture (HCMR) in Crete. Rotifers were cultured semi‐continuously at a density of 257.6 ± 2.6 ind. mL^?1, with a daily dilution rate of 25%. An automatic feeder was used to administer dry feed (Selco Sparkle^®) every 10 min. A number of factors were investigated so as to provide an in‐depth analysis of the advantages and disadvantages of the two systems, especially when compared with traditional protocols such as batch culture. The use of these two systems resulted in a culture of considerable duration (41 days) and daily yield (0.21 ± 0.01 ind. 10⁶ day^?1 L^?1), using exclusively dry feed. Both systems can be easily applied in hatcheries, requiring few modifications on existing protocols and contribute to a stable and predictable production. Further improvements include manipulation of the dilution rate, food ration, culture volume and duration, as well as choice of a phytoplankton‐based diet.     

Growth and fatty acid composition of juvenile Cerastoderma edule (L.) fed live microalgae diets with different fatty acid profiles

The importance of dietary 20:5n‐3 (EPA), 22:6n‐3 (DHA) and 20:4n‐6 (ARA) for growth, survival and fatty acid composition of juvenile cockles (Cerastoderma edule) was investigated. Cockles of 6.24 ± 0.04 mm and 66.14 ± 0.34 mg (live weight) were distributed into three treatments where live microalgae diets were fed constantly below the pseudofaeces production threshold, for three weeks. Diets had distinct fatty acid profiles: high EPA (53% Chaetoceros muelleri + 47% Pyramimonas parkeae), no DHA (47% Brachiomonas submarina + 53% Tetraselmis suecica) and low ARA concentrations (73% P. parkeae + 27% Phaeodactylum tricornutum). Growth was positively affected by high EPA and low ARA diets, whereas no significant growth was observed for the no DHA diet. High mortality of cockles fed no DHA diet raises questions about its suitability for cockles. In balanced diets with EPA and DHA, lower concentrations of ARA do not limit growth. The impact of dietary fatty acids was evident in the fatty acids of neutral and polar lipids of cockles. In polar lipids of all cockles, there was a decrease in EPA, in contrast to an increase in DHA. The combination of EPA and DHA in a live microalgae diet was beneficial for the growth and survival of juvenile cockles.     

Lutein enrichment of the rotifer Brachionus sp. using freeze‐dried Muriellopsis sp. cells

Rotifers, used in aquaculture as feed for early larval stages, are poor in carotenoids compared with copepods, the natural most common prey item of marine fish larvae. A minimal sufficient level of carotenoids in fish larvae may be essential for a correct development. The freshwater microalga Muriellopsis sp. has been identified as a potential source of the carotenoid lutein with possible application in aquaculture. Lutein is a potent antioxidant, with potentially beneficial effects as a carotenoid for fish larvae. This study describes the lutein enrichment of the rotifer Brachionus sp. when incubated with homogeneously dispersed suspensions of freeze‐dried, lutein‐rich Muriellopsis sp. in seawater. A short enrichment time (2 or 2.5 h) resulted in a higher lutein content of the rotifers than a long enrichment time (13.5 or 24 h). Although the concentration of lutein in enriched rotifers, 0.06–0.16 mg (g dry weight)^?1 was low compared with its content in Muriellopsis sp. [about 3 mg (g dry weight)^?1], it was in the range reported for natural copepod populations. Therefore, we conclude that freeze‐dried biomass of Muriellopsis sp. can be used to enrich rotifers with lutein, providing an adequate level of this carotenoid for marine fish larvae.     

Essential fatty acid requirements of cultured marine fish larvae

Feeding of marine fish larvae is, in most cases, limited to the administration of two species of live prey. This reduction in the range of food available for the cultured larvae may occasionally lead to nutritional imbalances or deficiencies. A large amount of research has been recently devoted to the study of the essential fatty acid requirements of marine fish larvae. Studies on the biochemical composition of developing eggs and larvae, as well as the comparison of the patterns of loss and conservation during starvation, pointed out the importance of n-3 HUFA and arachidonic acid as essential fatty acids for larvae of marine fish. The biochemical composition of marine fish larvae, in terms of lipid content and fatty acid composition of total and polar lipids, is modified by dietary levels of essential fatty acids. Larval growth, survival and activity have also been reported to be affected by dietary levels of essential fatty acids. In addition, some pathological signs, such as hydrops or abnormal pigmentation, have been related to essential fatty acid deficiency in these fish. Based on these effects, the essential fatty acid requirements of marine larval fish have been reported to range between 0.3 and 55 g kg^?1 n-3 HUFA on a dry weight basis, suggesting that quantitative requirements of fish larvae may differ from those of juveniles or adults. But quantitative requirements for larvae of the same species reported by various authors are often contradictory. These differences are discussed in relation to the dietary lipid content, ratio 20:5n-3/22:6n-3 and culture conditions used.     

The enrichment of rotifers (Brachionus plicatilis) and Artemia franciscana with taurine liposomes and their subsequent effects on the larval development of California yellowtail (Seriola lalandi)

Taurine is an essential or conditionally essential nutrient for many species of marine fish, especially during early development. There is growing evidence that marine fish larvae benefit from taurine‐enriched rotifers; however, it is unknown if larvae benefit from taurine‐enriched Artemia. We investigated the effects of taurine‐enriched rotifers (Brachionus plicatilis) and Artemia franciscana on the growth and whole‐body taurine concentrations of California yellowtail (Seriola lalandi; CYT) larvae. The approach used in this study was to encapsulate taurine within microparticles (liposomes), which were then fed to rotifers and Artemia. We found that feeding taurine liposomes to rotifers and Artemia resulted in taurine concentrations in these prey species that were similar to or above those previously reported in copepods. At the end of the rotifer phase, CYT larvae fed taurine‐enriched rotifers showed increased growth (final dry weights; DW) and had higher whole body taurine concentrations when compared to larvae fed unenriched rotifers. At the end of the Artemia phase, CYT whole body taurine concentrations varied among dietary treatments. Larval lengths and DWs were not significantly different among treatments at the end of the Artemia phase, suggesting that the taurine concentrations of unenriched Artemia were sufficient to support the growth of CYT larvae.     

Use of essential oils extracted from three Origanum species for disinfection of cultured rotifers (Brachionus plicatilis)

Essential oils extracted from three plant species of the genus Origanum, Origanum vulgare subsp. hirtum, Origanum onites L. and Origanum marjorana L., were assayed as potential antibacterial agents for disinfection of rotifers Brachionus plicatilis. Survival of rotifers in seawater with added oregano essential oils was significantly higher than in the control treatment (P < 0.05). When rotifers were incubated in seawater with added oregano essential oils at a final concentration of 10 mg L^?1 for 4 h, the numbers of total culturable bacteria were not significantly lower than in the control treatment (P > 0.05), while the numbers of presumptive Vibrio in rotifers were significantly reduced compared with the control treatment (P < 0.05).     

Essential fatty acid requirement of juvenile red drum (Sciaenops ocellatus)

Feeding experiments and laboratory analyses were conducted to establish the essential fatty acid (EFA) requirement of red drum (Sciaenops ocellatus). Juvenile red drum were maintained in aquaria containing brackish water (5 ± 2‰ total dissolved solids) for two 6-week experiments. Semipurified diets contained a total of 70% lipid consisting of different combinations of tristearin [predominantly 18:0] and the following fatty acid ethyl esters: oleate, linoleate, linolenate, and a mixture of highly unsaturated fatty acids (HUFA) containing approximately 60% eicosapentaenoate plus docosahexaenoate. EFA-deficient diets (containing only tristearin or oleate) rapidly reduced fish growth and feed efficiency, and increased mortality. Fin erosion and a “shock syndrome” also occurred in association with EFA deficiency. Of the diets containing fatty acid ethyl esters, those with 0.5–1% (n-3) HUFA (0.3–0.6% eicosapentaenoate plus docosahexaenoate) promoted the best growth, survival, and feed efficiency; however, the control diet containing 7% menhaden fish oil provided the best performance. Excess (n-3) HUFA suppressed fish weight gain; suppression became evident at 1.5% (n-3) HUFA, and was pronounced at 2.5%. Fatty acid compositions of whole-body, muscle and liver tissues from red drum fed the various diets generally reflected dietary fatty acids, but modifications of these patterns also were evident. Levels of saturated fatty acids appeared to be regulated independent of diet. In fish fed EFA-deficient diets (containing only tristearin or oleate), monoenes increased and (n-3) HUFA were preferentially conserved in polar lipid fractions. Eicosatrienoic acid [20:3(n-9)] was not elevated in EFA-deficient red drum, apparently due to their limited ability to transform fatty acids. Red drum exhibited some limited ability to elongate and desaturate linoleic acid [18:2(n-6)] and linolenic acid [18:3(n-3)]; however, metabolism of 18:3(n-3) did not generally result in increased tissue levels of (n-3) HUFA. Based on these responses, the red drum required approximately 0.5% (n-3) HUFA in the diet (approximately 7% of dietary lipid) for proper growth and health.     

Effects of yeast and algal diets on the growth and biochemical composition of the rotifer Brachionus plicatilis (Müller) in culture

Abstract. Growth rates and fecundity of Brachionus plicatilis fed baker's yeast, a diatom, and three different flagellates, were monitored during a 15-day feeding trial. Identical growing conditions and equivalent food levels for the different diets were maintained during rotifer culture. The highest growth rate and mean fecundity were exhibited by rotifers fed Chlorella saccharophila , followed in decreasing order by those fed Isochrysis galbana (T-iso), Tetraselmis suecica, Saccharomyces cerevisiae and Thalassiosira pseudonana . Significant differences in contents of neutral lipids, polar lipids, sugars, glycogen, protein, ash and corresponding energy reserves in the resultant five groups of rotifers were diet dependent. Total available energy, about 15 kJ/g in rotifers fed S. cerevisiae and I. galbana (T-iso) was significantly different from the 14kJ/g in rotifers fed T. suecica and C saccharophila, and in turn from the 13kJ/g in rotifers fed T. pseudonana .