Rotifer Culture Research Articles

Role of rotifer (Brachionus plicatilis) and Artemia (Artemia salina) nauplii in the horizontal transmission of a natural nervous necrosis virus (NNV) reassortant strain to Senegalese sole (Solea senegalensis) larvae

Background Marine invertebrates are provided as a first feed for marine fish larvae because of their strict nutritional requirements, despite also being a potential source of infectious agents. Aim To assess horizontal transmission of a nervous necrosis virus reassortant strain (NNV) to sole larvae via Artemia and rotifers. Materials and methods Rotifer (Brachionus plicatilis) and Artemia (Artemia salina) nauplii cultures were bath infected with a reassortant (RGNNV/SJNNV) NNV strain isolated from gilthead sea bream and viral internalisation was confirmed by IFA. Senegalese sole (Solea senegalensis) larvae were fed on infected Artemia and disease signs and mortality were recorded. In addition, NNV viability was checked in cultures of either unfed invertebrates or invertebrates fed on phytoplankton and in the supernatant of microalgae cultures. All samples were tested by RT-qPCR and inoculation in cell culture. Results Both rotifers and Artemia internalised NNV. Experimental transmission to sole larvae was achieved using infected Artemia and subsequently 60% mortality was recorded. At 24 h post-infection, orally infected individuals contained 9.34 × 104 copies of viral RNA, whereas the bath infection yielded 2.05 × 106 RNA copies larvae−1. Viral presence in both invertebrates was detected up to 8 days post infection but viral load decreased over time. Feeding with microalgae decreased viral detection even more and microalgae supernatants were demonstrated to significantly affect NNV viability. Conclusions Our results demonstrate that both invertebrates can bioaccumulate NNV and that Senegalese sole larvae fed on infected Artemia might develop viral encephalopathy and retinopathy and high mortality.

Dietary Value of Waste-Fed RotiferBrachionus rotundiformison the Larval Rearing of Japanese WhitingSillago japonica

Live food resources are useful for larval fish rearing. However, production of sufficient live food resources is expensive. This study employed a cost-effective rotifer culture technique using fish waste diet (FWD) and investigate the effect of the FWD-fed rotifer on larval rearing of the Japanese whiting,Sillago japonica. Fertilized eggs ofS. japonicawere hatched in polycarbonate tanks containing 100 l of artificial seawater at 10 eggs l-1with 50 ml min-1of aeration at room temperature. Two diets (i.e. FWD-fed rotifers and rotifers fed with super fresh Chlorella-V12 as control) were used with 10 rotifers ml-1for 10 days. Fish were sampled every two days for morphometric and gut content analysis. Fatty acid analysis was done for both rotifers and fish larvae. The fish larvae fed with FWD-rotifers had higher total length than those given control diet. There was no significant difference in survival rate, viability, dry weight, gut content, head length, eye diameter, and body depth between the two diets. The DHA recorded of total lipid for the fish given FWD-rotifer and control fish are3 5.2% and 18.2% respectively. The use of waste-fed rotifers is cost-effective method to enhance the production of larval fish rearing in hatcheries.

Enriching Rotifers with "Premium" Microalgae: Rhodomonas lens.

The nutritional value of the marine cryptophyte Rhodomonas lens for the filter feeder Brachionus plicatilis as well as its biotechnological potential as a source of phycoerythrin (PE) and polyunsaturated fatty acids (PUFA) were evaluated in semi-continuous cultures maintained with different daily renewal rates (RR), from 10% (R10) to 50% (R50) of the total volume. Steady-state cell density decreased from 22 to 7 × 106cellsmL-1 with increasing RR, with the maximum cell productivity, nearly 0.4gL-1day-1, observed with R40. PE cell content attained the highest values with the highest RR (circa 9pgcell-1). All treatments of R. lens maintained under nitrate-saturated conditions (R20-R50) showed a similar high content of PUFAs, > 60% of total fatty acids (FA), with linolenic acid (18:3n-3) and 18:4n-3, representing 12 and 29% of total FA respectively. The PUFA level in the nitrogen-limited R10 cultures was significantly lower (37%). R. lens promoted higher weight gain in the rotifer B. plicatilis than Tisochrysis lutea (T-ISO), a species commonly used for rotifer culture and enrichment. Significant differences were found in the protein content and in the ratio n-3/n-6 fatty acids among rotifers fed with R. lens from different RRs, with higher values being found in those fed with R. lens from higher RRs. The enrichment of the rotifers for short periods of 3h was sufficient to modify the biochemical composition of the rotifers, but it was evidenced as too short for the accumulation of PUFAs, when compared to long-term (24h) enrichment. The rotifers reflected the higher protein and PUFA content of R. lens cultivated with nutrient sufficient microalgae (R40) after only 3h of enrichment. These results demonstrate that semi-continuous culture of R. lens under appropriate conditions can strongly enhance the nutritional value of this species, being reflected in the growth and biochemical composition of the filter feeder, even in short exposure periods.

PRODUCTIVITY OF ROTIFER Brachionus plicatilis CULTURED UNDER BATCH SYSTEM BY USING SUGAR BEET MOLASSES AS A SUBSTITUTE MEDIUM

This study deals with the determination of availability of sugar beet molasses as a nutritional enrichment factor for rotifer cultures. A series of experiments under varying conditions such as ratio of sugar beet molasses in the nutrient complex were carried out in order to determine the effectiveness of sugar beet molasses as a carbohydrate and mineral source for the purpose of aquacultural operations. The culture water of Brachionus plicatilis with the salinity of 38% was treated through mechanical filtration and UV sterilization. 1 T cylindro-conical tanks are utilized for rotifer culture. Parameters of culture water were stabilized at values as following; Oxygen concentration:5 to 10 ppm, temperature: 27 0 C and pH:6.8 to 7.4. Rotifers were counted by using Sedgewick Rafter Cell on a daily basis in order to determine increase of number of individuals. The study was conducted as 3 different trial groups, each consisting three separate tanks, so triplicate determinations were obtained. For the Group I, 10% molasses by weight of Culture Selco Plus (200 gr Selco Plus+20 gr molasses per tank) was added 6 times a day with 4 hours’ intervals. For the Group II, 20% molasses by weight of Culture Selco Plus (200gr Selco Plus+40gr molasses per tank) was added and lastly for the control group, Group III, no molasses has been added and 200 gr Culture Selco Plus was only added per tank. The addition of molasses to nutritive complex has a decreasing effect on the number of rotifer and increasing effect on the number of ciliate. From the study, it is clear that sugar beet molasses should not be used as an additional nutritional factor for rotifer cultures due to its decreasing effect on the number rotifers and stimulating effect on the ciliate booming. Addition of 10% and 20% molasses resulted in 31.4% and 23.3% decrease in number of rotifers at the time of T3, respectively. On the other hand, it was observed that addition of molasses in the nutritive complex has not affected on the percentage of rotifers bearing eggs.

Effects of different harvesting and processing methods on Nannochloropsis oculata concentrates and their application on rotifer Brachionus sp. cultures

The effects of the cell separation method and the biomass processing of Nannochloropsis oculata were evaluated with respect to the cultivation performance of rotifers (Brachionus sp.). The algal culture was harvested by centrifugation or flocculation, and the biomass separation efficiency and the concentration factor were determined. The biomass was processed by chilling, freezing, heat-drying, or freeze-drying; and the cell integrity, resuspension percentage, and sedimentation profile of the diets were evaluated. The zootechnical performance (maximum rotifer density, growth rate, maximum egg production, fecundity, and feed ingestion rate) of rotifer cultures that fed the concentrated diets was compared with feeding the fresh culture of N. oculata (control treatment). Centrifugation resulted in 97% separation efficiency and a concentration factor of 578 and flocculation followed by centrifugation showed an efficiency of ~ 100% and concentration factor of 444. The harvesting technique did not have a significant influence on the characteristics of the diets and also on the zootechnical performance of the rotifers. The characteristics of the chilled concentrates (cell integrity, resuspension percentage, and sedimentation profiles) were similar to the control treatment, and the rotifers fed on those diets had the highest growth performances. Rotifers fed on frozen diets presented the worst performances. The heat-dried diets presented the lowest cell viability percentages, lowest resuspension percentages, and higher sedimentation rates. The feed ingestion rates of these diets were the lowest, resulting on inferior growth performance of the rotifers. In summary, both harvesting methods could be used, and chilling is the most recommended method for processing N. oculata biomass.

Diversity and function of the microbial community under strong selective pressure of rotifers.

We unearthed some interesting microecological discoveries while selecting for the most beneficial bacterial strains to be used as probiotics in Lecane inermis rotifer mass culture. For 3 years, we maintained the cultures of L. inermis, with selection for the highest growth rate and resistance to potential contamination. Then, we conducted further selection and isolation in two groups: rotifers inoculated with the bacterial consortium isolated from the rotifer cultures, and rotifers fed with a commercial bioproduct. Selection was conducted in demanding conditions, with particulate matter suspended in spring water as a substrate, without aeration and under strong consumer pressure, and led to selection of two cultivable strains isolated from the optimal rotifers culture. According to molecular analysis, these strains were Aeromonas veronii and Pseudomonas mosselii. Biolog® ECO plate tests showed that both investigated bacterial communities metabolized wide but similar range of substrates. Therefore, intensely selective conditions led to considerable reduction in bacterial community regarding taxonomy, but not in metabolic activity, showing a functional composition decoupling. Aside from this result, our novel selection method dedicated to the sustainable culture of two trophic levels, a directed selection procedure (DSC), could potentially lead to the development of biotechnologically valuable strains with high metabolic activity and the ability to metabolize different sorts of substrate without harmful impact on higher trophic levels.

Utilization of <i> Schizochytrium mangrovei </i> PQ6 as feed for rotifer <i> Brachionus plicatilis </i> in rearing black sleeper’s larvae (<i>Bostrichthys sinensis</i>, Lacepede, 1881)

Schizochytrium mangrovei PQ6 contains many important polyunsaturated fatty acids, such as docosahexaenoic acid (DHA, C22: 6w-3), eicosahexaenoic acid (EPA, C20: 5w-3) and docosapentaenoic acid (DPA, C22: 5w-6). These fatty acids are essential for survival and growth of many marine fish larvae. In this paper, fresh biomass of S. mangrovei PQ6 was used for culturing rotifer Brachionus plicatilis. In the first formula (L1), rotifers were fed on baker’s yeast; in second formula (L2), rotifers were fed on mixed two microalgae, Nannochloropsis oculata and Chaetoceros gracilis; in the third formula (L3), rotifers were fed on S. mangrovei PQ6 biomass. The results indicated that S. mangrovei PQ6 biomass can replace baker’s yeast and autotrophic microalgae in feeding rotifers. Total lipid, total fatty acid and polyunsaturated fatty acid contents of rotifers fed on formula 3 (S. mangrovei PQ6) were higher than those within others. High polyunsaturated fatty acid (C ≥ 20) in omega-3 and 6 groups; DHA and DPA contents of rotifers fed on S. mangrovei PQ6 were the highest, accounting for 39.81% of total fatty acid; 41.95% and 8.24% polyunsaturated fatty acids, corresponding to 20.52; 12.15 and 2.4 mg/g of dried weight, respectively.The survival rate of black sleeper’s larvae was highest when they were fed on rotifers grown on L3 formula (51.20 ± 0.89%), followed by L2 formula (48.70 ± 2.67%) and the lowest with L1 formula (43.44 ± 1.54%) (P < 0.05). However, among three formulas, no significant difference was found in the growth of black sleeper’s larvae (P > 0.05). These results suggest that biomass of S. mangrovei PQ6 can replace traditional feed like autotrophic microalgae or baker’s yeast for biomass culture of the Brachionus plicatilis rotifer for rearing black sleeper’s larvae to enhance seed quality to meet demands of market.

Bacterial communities protect the alga Microchloropsis salina from grazing by the rotifer Brachionus plicatilis

Open algal ponds are likely to succumb to unpredictable, devastating crashes by one or several deleterious species. Developing methodology to mitigate or prevent pond crashes will increase algal biomass production, drive down costs for algae farmers, and reduce the risk involved with algae cultivation, making it more favorable for investment by entrepreneurs and biotechnology companies. Here, we show that specific algal-bacterial co-cultures grown with the green alga Microchloropsis salina prevented grazing by the marine rotifer, Brachionus plicatilis. We obtained seven algal-bacterial co-cultures from crashed rotifer cultures, maintained them in co-culture with Microchloropsis salina, and used a microalgal survival assay to determine that algae present in each co-culture were protected from rotifer grazing and culture crash. After months of routinely diluting and maintaining these seven algal-bacterial co-cultures, we repeated the assay and found the opposite result: none of the seven bacterial communities protected the microalgae from rotifer grazing. We performed 16S rRNA gene amplicon sequencing on the protective and nonprotective co-culture samples and identified substantial differences in the makeup of the bacterial communities. Protective bacterial communities consisted primarily of Alphaproteobacteria (Rhodobacteraceae) and Gammaproteobacteria (Marinobacter, Pseudomonas, Methylophaga) while nonprotective bacterial communities were less diverse and missing many putatively crucial members. We compared the seven protective communities with the seven nonprotective communities and we correlated specific bacterial amplicon sequence variants with algal protection. With these data, our future work will aim to define and develop an engineered-microbiome that can stabilize industrial Microchloropsis salina cultures by protecting against grazer-induced pond crashes.

Effect of probiotic Pediococcus acidilactici on growth, reproductive and bacterial count of marine rotifer Brachionus plicatilis

This study investigated the effects of different concentrations of probiotic Pediococcus acidilactici on the total number, specific growth rate, attached egg number and microbial count (lactic acid and total aerobic bacteria) in culture water and body of marine rotifer Brachionus plicatilis . The experiment was conducted as a completely randomized design in four groups: first group fed probiotic at 0.5×10 6 CFU ml -1 (P1), the second group fed probiotic at 1×10 6 CFU ml -1 (P2), the third group fed probiotic at 1.5×10 6 CFU ml -1 (P3), and the last group fed without probiotic (NP, control) . Rotifers were cultured in standard conditions at an initial density of 30 ind.ml -1 using a 2.5×10 6 cell mL -1 of Nanochloropsis oculta . Based on the results, the maximum number (215±4.91 ind.ml -1 ) of rotifers were significantly obtained after ten days in the third group in comparison to other treatments ( P 0.05). Moreover, it was revealed that the total aerobic bacteria was significantly related to the 3rd group that found be 1.80×10 3 CFU rotifer -1 and 34.0×10 4 CFU ml -1 in rotifer body and culture water,respectively ( P <0.05). Nonetheless, lactic acid bacteria of rotifer body and culture water was not concentration-dependent and the highest number of lactic acid bacteria in rotifer body (50.58±6.08 CFU rotifer -1 ) and rotifer culture water (4.43±3.28×10 3 CFU ml -1 ) was obtained in the treatments of P1 and P2, respectively. In conclusion, the present study showed that the bacteria P. acidilactici with N. oculta algae provide a higher growth rate and total aerobic bacteria in rotifer B. plicatilis.

Effect of two commercial probiotic products on population growth of rotifer Brachionus rotundiformis Tschugunoff

The application of probiotics has varying effects on rotifer culture. These effects can either be directly on their growth and reproduction or indirectly by affecting the culture environment. The role played by specific probiotic candidates is still indistinct and further research is required to ascertain their significance in rotifer culture. In this study, euryhaline rotifer Brachionus rotundiformis (SS-type) was used to evaluate the effect of two commercial probiotic products on rotifer density, reproductive parameters and unionized ammonia concentration for a twelve-day culture period. To estimate these parameters, rotifers were cultured in glass bottles containing dilute sterilized sea water with the following five diet treatments: Nannochloropsis oculata (control), Igsign (PB1), Toaraze Aqua (PB2) or the combination of N. oculata with either PB1 or PB2. The rotifer density was significantly affected by the addition of probiotic. N. oculata + PB2 treatment resulted in higher rotifer density and specific growth rate compared to the control treatment. Mixis induction was unaffected by N. oculata + PB2 while it was repressed by N. oculata + PB1. Ammonia concentration was not significantly affected with the addition of probiotics. These results indicate that probiotics have synergetic effects with microalgae diet that resulted in the highest rotifer densities.

Effect of nutritional status and concentration of Nannochloropsis gaditana as enrichment diet for the marine rotifer Brachionus sp

The development of strategies to improve the quality and to optimize the costs of rotifer production for fish larval feeding, particularly those related to microalgal supply, are highly desirable. In the present work, three different rations (80,000, 130,000 or 200,000 cells rotifer−1) of Nannochloropsis gaditana were assessed as enrichment diets for the marine rotifer Brachionus sp. The microalga was cultured in semi-continuous regime with two different daily renewal rates, 10% and 40%, which provided nutrient-depleted or nutrient-sufficient conditions respectively and resulted in strong differences in biochemical composition. Protein and arachidonic acid (ARA) content were twice higher and eicosapentaenoic acid (EPA) was three times higher in N. gaditana from nutrient-sufficient cultures. After 24-h enrichment, rotifers fed N. gaditana cultured in nutrient-sufficient conditions showed higher dry weight, as well as higher contents of protein, lipids and carbohydrates. Total polyunsaturated fatty acids (PUFAs) and ARA contents increased >50%, whereas EPA content doubled in rotifers fed N. gaditana from nutrient-sufficient conditions. As for feed ration, the increase from 80,000 to 130,000 cells individual−1 enhanced the accumulation of protein, lipids, carbohydrates, total PUFAs, ARA and EPA, but a further increase to 200,000 cells individual−1 only enhanced the accumulation of lipid content. Our results show that the nutritional quality of microalgae, rather than concentration, is the most important factor influencing biomass productivity and biochemical composition of rotifer cultures. Since microalgal biomass is in general a limited resource for aquaculture, the improvement of its nutritional value through the optimization of culture conditions should be considered as an adequate and cost-efficient strategy to produce high-quality live feed.

Effects of the biochemical composition of three microalgae on the life history of the rotifer Brachionus plicatilis (Alvarado strain): an assessment

The biochemical composition of microalgae used as food is essential for aquatic species in commercial production systems, such as rotifers and microcrustaceans. Life table bioassays with the rotifer Brachionus sp. “Alvarado” strain were performed using three microalgae (Nannochloropsis oculata, Dunaliella salina and Isochrysis sp.) as food. Microalgae growth rate, dry weight and biochemical composition (protein, lipid, carbohydrate) and pigments (chlorophyll and carotenoid) were determined. The microalgae showed significant differences in their biochemical composition. N. oculata showed the highest growth rate, while D. salina showed the slowest growth rate, but instead, it displayed a higher content of proteins, lipids, carbohydrates, chlorophyll, and carotenoids per cell. Rotifer life table analysis showed no significant differences among any of the microalgae as food bioassays. However, Isochrysis sp. had a higher effect on the net reproductive rate of the rotifer Brachionus sp. “Alvarado” followed by D. salina, while N. oculata showed a higher effect on life expectancy and generation time. In conclusion, the three microalgae are found to be useful to support rotifer cultures; however, both, D. salina and Isochrysis sp., might improve the rotifer culture due to better growth and reproduction in short time. This information is useful to implement the culture of this tropical strain of Brachionus plicatilis complex in order to obtain high population densities, making rotifers available for several applications such as the establishment of larviculture in hatcheries, bioassays for ecological studies or to assess its sensitivity through toxicity tests.

Growth performance of the very small rotifer Proales similis is more dependent on proliferating bacterial community than the bigger rotifer Brachionus rotundiformis

Proales similis and Brachionus rotundiformis are commercially cultured species, which predominantly feed on microalgae. However, the importance of the bacterial community on growth performance of P. similis and B. rotundiformis culture is still unknown. In this study, the effect of limiting the bacterial growth and, as a second aim, the effect of the addition of a small amount of live or dead bacteria on the growth performance and microbial community (MC) of P. similis or B. rotundiformis culture was evaluated for a 10day culture period. A rotifer culture with non-manipulated MC and fed autoclaved algae was used as the first control, and a culture started in autoclaved sea water and fed autoclaved algae was used as the second control. In order to test a feed effect, probiotics and other bacteria present in the culture system were killed (inhibited to grow) through the addition of an antibiotic mixture (AB) and the rotifer culture performance was compared to that of a culture to which live probionts were added. In the presence of the live probiotic mixture, both rotifers species showed a better growth performance than those without the presence of probiotic mixture or those with the AB added. In addition, the growth performance of the rotifer P. similis is more dependent on proliferating bacterial community than the rotifer B. rotundiformis. The supplementation of these probiotic bacteria not only increased the production of the rotifers, but also had a regulating effect on the microbiota. The bacterial density was below detection limit in TCBS, MRS agar and MA in all treatments using AB during the culture period for both rotifer species.

Performance of Debaryomyces hansenii as a Diet for Rotifers for Feeding Zebrafish Larvae.

The zebrafish larval stage is a critical moment due to high mortality rates associated with inadequate supplies of nutritional requirements. Larval feeding has important challenges associated with such factors as small mouth gape (≈100 μm), the low activity of digestive enzymes, and the intake of live food. A common zebrafish live food at the onset of exogenous feeding is rotifers, mainly Brachionus plicatilis. These rotifers should be fed with other microorganisms such as microalgae or yeast, mostly from the Saccharomyces genus. In the laboratory, the culture of microalgae is more expensive than the culture of yeast. The aim of this study was to evaluate the performance of Debaryomyces hansenii as a diet for rotifers in comparison to a microalgae-based diet (Rotigrow®). To achieve this aim, we assessed the rotifer total protein content, the rotifers fatty acid profile, zebrafish larval growth performance, the expression of key growth, and endocrine appetite regulation genes. The total protein and fatty acids content were similar in both rotifer cultures, averaging 35% of dry matter (DM) and 18% of DM, respectively. Interestingly, the fatty acids profile showed differences between the two rotifer cultures: omega-3 fatty acids were only observed in the Microalgae/rotifer, whereas, omega-6 fatty acids presented similar levels in both rotifer cultures. No differences were observed in the larval body length distribution or mortalities between the rotifer cultures. However, gh, igf-1, and cck gene expression showed significantly higher upregulation in zebrafish fed the Microalgae/rotifer diet compared with those fed the Debaryomyces/rotifer diet. In conclusion, D. hansenii could be an alternative diet for rotifer used as a live food in zebrafish larvae at the onset of exogenous feeding. The gene responses observed in this work open up the opportunity to study the effect of omega-3 supply on growth regulation in zebrafish.

Treatment of seawater for rotifer culture uses applying adsorption and advanced oxidation processes

The use of live feed is common in aquaculture processes due to its nutritional advantages, and rotifers comprise one of the most used feeds in finfish aquaculture. However, bacteria present in water used for rotifer culture are a significant source of infection and consequently diseases could cause mortality in fishes. Advanced oxidation process (AOPs) are methods that may be useful to eliminate bacterial infections present in aquaculture wastewater and may allow the reuse of this water. Therefore, the prospect of using different advanced oxidation processes to inactivate Vibrio spp. and total bacteria cultures (TCB) present in this kind of water was evaluated. In particular, UVc/O 3 /H 2 O 2 , UVc/H 2 O 2 and O 3 /H 2 O 2 and the adsorption process were applied to seawater coming from a fish farming tank. The results obtained were highly promising because it was possible to inactivate 100% of bacteria present in the seawater when UVc/ozone/hydrogen peroxide and UVc/hydrogen peroxide were applied, needing around 20 and 80 min for Vibrio spp. and total culturable bacteria inactivation, respectively. These results are justified due to the strong action between H 2 O 2 in the presence of UVc, which may produce a large amount of hydroxyl radicals. Nevertheless, when ozone/hydrogen peroxide was applied, total inactivation of bacteria did not occur. Also, an interesting relationship between bacteria inactivation and oxide reduction potential of seawater was observed during the process. Rotifers put in water after the treatment did not suffer behavior changes (swimming) and no mortalities were observed.

Technical and economic feasibility of adding pure oxygen for rotifer production (Brachionus plicatilis) spotted snapper (Lutjanus guttatus) larvae feeding

A study to technically and economically evaluate the addition of pure oxygen in a culture of rotifers for feeding marine fish in the Pacific Marine Park, Puntarenas, Costa Rica was performed. The results showed no significant differences (ANOVA P&amp;gt; 0.05) between the cultivation with addition of oxygen treatment and control, regarding the instantaneous growth rate, crop yield, doubling time, rotifers length and maximum densities reached. Concerning the economic analysis, control culture was 53% less expensive compared to the addition of oxygen system.

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English

Cod Larviculture Using High‐density Rotifer Production with Different Enrichments

AbstractHigh‐density (HD) rotifer culture systems have been recently commercialized, but are not commonly used by the aquaculture industry. The aim of this study was to determine if HD systems could be used in cod hatcheries. An enrichment strategy using the commercial products, ArteMac and Protein Selco Plus (Com), was compared with the manufacturer's suggested enrichment, using Pavlova‐DHA (Pav). The Pav enrichment increased the eicosapentaenoic acid (EPA) levels in rotifers, but reduced the docosahexaenoic acid (DHA) and 22:5n‐6 (n‐6DPA, docosapentaenoic acid) levels. Larvae EPA levels in both polar and neutral lipids were relatively stable in the larvae fed with Com‐rotifers; while they were higher in early stages, they were progressively reduced through ontogeny in the Pav‐rotifers fed‐larvae. DHA levels in polar lipids decreased in larvae, particularly when fed with HD‐Pav rotifers. In all larvae, arachidonic acid (ARA) levels increased in the polar and neutral lipids, regardless of treatments. In both lipid fractions, the levels of ARA were quite stable in time, but still higher in larvae fed with Com‐rotifers. Bacterial load was lower in larvae fed with Pav‐rotifers. Denaturing gradient gel electrophoresis (DGGE) bacterial profiles of larvae and rotifers were all similar. This study shows the potential of using HD systems to produce rotifers, but highlights the necessity of adjusting the nutritional composition of rotifers prior to being fed to larvae.

Application of a new real-time polymerase chain reaction assay for surveillance studies of lymphocystis disease virus in farmed gilthead seabream.

BackgroundLymphocystis disease (LCD) is the main viral infection reported to affect cultured gilthead seabream (Sparus aurata) in Europe. The existence of subclinical Lymphocystis disease virus (LCDV) infection in this fish species has been recognised by using polymerase chain reaction (PCR)-based methods. Nevertheless, these methods do not provide quantitative results that can be useful in epidemiological and pathological studies. Moreover, carrier fish have been involved in viral transmission, therefore the use of specific and sensitive diagnostic methods to detect LCDV will be relevant for LCD prevention.ResultsWe have developed a real-time PCR (qPCR) assay to detect and quantify LCDV. The assay was evaluated for viral diagnosis in surveillance studies in gilthead seabream farms, and also to identify viral reservoirs in a hatchery. The prevalence of LCDV infection in the asymptomatic gilthead seabream populations tested varied from 30 to 100 %, including data from one farm without previous records of LCD. Estimated viral load in caudal fin of subclinically infected fish was two to five orders of magnitude lower than in diseased fish. The qPCR assay allowed the detection of carrier fish in broodstock from a farm with a history of clinical LCD in juvenile fish. In addition, the quantitative detection of LCDV was achieved in all samples collected in the hatchery, including fertilized eggs, larvae and fingerlings, and also rotifer cultures and artemia metanauplii and cysts used for larval rearing.ConclusionsThe qPCR assay developed in this study has proved to be a rapid, sensitive, and reliable method for LCDV diagnosis, which could be valuable to identify LCDV reservoirs or to study viral replication in gilthead seabream.

The Performance of Continuous Rotifer (Brachionus Plicatilis) Culture System for Ornamental Fish Production

Rotifers are considered as one of the most important prey organisms in the culture of altricial fish larvae. However, high density rotifer culture is often problematic due to water quality problems which results in frequent crashes. In the present study, the performance of a small-scale, continuous system was evaluated for culturing rotifers, Brachionus plicatilis, using concentrated nonviable green algae, Nannochloropsis oculeataas feed in a 160 L tank for a period of 90 days. The system configuration was simple and major components consisted of a protein skimmer and a pure oxygen delivery system. Although egg ratio increased from 3% on day 1 to 21.8% and 39.3% on days 7 and 9, respectively, rotifer growth was slow at start up and resulted in fluctuations in total number of rotifers between days 19-41. Rotifer densities remained < 400 until day 51 but increased at higher rates reaching 900 individuals/mL on day 55, 1,620 on day 60 and 2,127 on day 70. Rotifer density reached a maximum of 2,188 individuals/mL on day 85. Once the rotifer density exceeded 1,500 individuals/mL (day 60), periodical harvesting (a total of 16 harvest events) produced a total of 369,920,000 rotifers corresponding to a daily production of 12,330,667 individuals/day during the next 30 days until the experiment was terminated at day 90. As a result of periodic harvesting, water makeup and continuous protein skimming, total settleable solids and NH3-N levels remained low and ranged between 4-22 mL/L and 0.4-2.2 mg/L, respectively. The authors' findings indicated that this inexpensive culture system can be successfully used for small-scale marine or freshwater ornamental fish production. Further work is required to minimize lag period at start-up and increase the production potential and yield by better management of suspended solids.