Motility of sea urchin Paracentrotus lividus spermatozoa in the post‐activation phase

The characterization of sperm motility patterns, particularly post‐activation changes, is the first step in setting up species‐specific protocols involving gamete management and embryo production, for both aquaculture and laboratory research purposes. This study is aimed at the characterization of the sperm motility pattern of the purple sea urchin Paracentrotus lividus. Semen samples were individually diluted in artificial sea water for sperm motility activation. They were then incubated at 18°C for up to 24 hr. Motility was evaluated on dilution, and 1 hr, 3 hr and 24 hr after activation, by computerized analyser. The semen fertilization capacity was also evaluated. Under our experimental conditions (dilution 1:1,000 in artificial sea water plus 0.05% BSA, 18°C, in the dark), P. lividus semen remained viable for up to 24 hr, as the total motile sperm and the fertilization percentages did not change significantly during the incubation time. In contrast, the mean curvilinear velocity and the subpopulation of rapid sperm (those having a curvilinear velocity > 100 µm/s) slightly but significantly decreased after 3 hr, thereafter remaining unchanged for up to 24 hr after activation. In conclusion, our results show that diluted P. lividus semen can be used for a longer period than that of most fish species, with no need for motility inhibition procedures, supporting its wider use in laboratory research. In addition, the development of artificial fertilization protocols for aquaculture production is simplified by long‐lasting sperm motility.     

The use of diatom Skeletonema costatum on aquaculture‐produced purple sea urchin (Paracentrotus lividus) larvae and post‐larvae diet

The quality of the microalgae provided on Paracentrotus lividus larvae rearing is a primordial factor having a direct (nutritional properties) and indirect (water quality) impact on growth, competence and survival. Skeletonema costatum is a diatom commonly used in the bivalve cultivation. However, the use of this diatom in P. lividus larval cultivations is poorly known. The Rhodomonas spp. is a microalgae commonly used in sea urchin larvae culture. Three different diets were tested on P. lividus larvae and post‐larvae cultivation (D1—Rhodomonas marina, D2—S. costatum, D3—mixture of both algae). Larvae fed with the D2 diet (55.8%) and D3 (39.9%) had a survival at 15 DAH higher than D1 (5.5%). The low survival in D1 could be due to the higher microbiological load on microalgae (Vibrio alginolyticus and V. pectenicide). Larvae fed with S. costatum (D2) showed a lower development than other diets. The competency index was lower for larvae fed with the D2. These results show that microalgae diversified diets contribute to a better development of P. lividus larvae. During the settlement and post‐settlement phase, there was also a lower growth of the sea urchin fed with the D2 and a higher survival for D3.     

Effects of on‐demand feeding on sea urchin larvae (Paracentrotus lividus; Lamarck, 1816), development,survival and microalgae utilization

This study compared the growth of sea urchin Paracentrotus lividus larvae cultured using two different rearing methods: a variable method based on a variable amount of feed (microalgae) and seawater exchange (30% or 50%) established according to the phytoplankton concentration in the larval cultures and a fixed method characterized by a fixed amount of feed and seawater exchange. Three microalgae diets, Isochrysis sp. (Tahitian strain, T‐Iso), Chaetoceros gracilis and a 50:50 mixed diet, were tested with both rearing methods. Larval development and survival were assessed at the 6‐arm pluteus stage (P6), competence (Cp) and metamorphosis (Mt). Data showed that the variable method reduced the requirements for phytoplankton and seawater exchange. Indeed, through the optimization of feed rations, it was possible to reduce the production of debris and settled phytoplankton, minimizing the need for water exchanges. Higher larval survival resulted at Cp and Mt stages for those reared with the variable method as opposed to the fixed one. Survival and development were also influenced by the tested dietary treatments: at Mt stage, the mixed diet resulted in a higher larval survival (63.3 ± 8.9%) than T‐Iso (19.7 ± 12.1%) and C. gracilis (23.4 ± 15.1%) (P < 0.05). These results suggest that the use of the variable method improves the larval survival and development and also it reduces resource consumption (phytoplankton, seawater use and work effort), which in turn could potentially improve the hatchery production of P. lividus.     

Increasing the stocking density in Paracentrotus lividus larviculture: Effects on survival and metamorphosis rates

The development of sustainable methods for sea urchin juvenile production is currently constrained by high mortality rates during larval growth and the high costs of larval rearing systems management. With the aim of developing a method for the production of juveniles of the purple sea urchin Paracentrotus lividus in a medium‐scale recirculating system, the present study focused on the effects of high stocking densities on larval growth. Plutei larvae were reared at three different densities (up to 7 ind/ml) in a semi‐static culture system. Larval survival and metamorphosis success were evaluated in order to identify the most effective density range. The highest metamorphosis rates (80%–95%) were obtained at 4 and 7 larvae/ml. These results are comparable with (and in some cases higher than) those reported for the same species at much lower larval densities. In conclusion, the rearing conditions tested here show for the first time that a significantly higher (4 ind/ml) stocking density than those of traditional P. lividus rearing methods (based on large volumes and low densities) can be adopted, thus supporting the feasibility of an increase in the final output of competent larvae with no increase in rearing volumes.     

Protein content and amino acid composition of eggs and endotrophic larvae from wild and captive fed sea urchin Paracentrotus lividus (Echinodermata: Echinoidea)

Knowledge on the biochemistry of proteins and amino acids (AA) of eggs and larvae of echinoids is scarce and the possibility to modify their profiles by diet manipulations is unknown. The protein content of eggs, prisms and pre‐plutei and the amino acid composition of eggs of Paracentrotus lividus from captive broodstock fed prepared diets were analysed and compared with the ones obtained from wild broodstock. Diets differed on protein source (fish or soy meals) and on protein content (10–40% dry weight – DW). Total and soluble protein content of both eggs and larvae was higher than 400 g kg^?1 DW and 200 g kg^?1 DW respectively. Glycine was the most abundant free AA. Very few differences were found among P. lividus eggs and larvae biochemical parameters, suggesting that their AA composition can hardly be changed by broodstock diet manipulations.     

Evaluation of dried macrophytes as an alternative diet for the rearing of the sea urchin Paracentrotus lividus (Lamarck, 1816)

The aim of this work was to assess the potential use of different dried macroalgae as food in the rearing of Paracentrotus lividus. Growth, consumption and food conversions were compared in adult sea urchins fed with fresh or dried thalli of four macroalgae species. Six experimental diets were tested: (a) fresh Palmaria palmata; (b) fresh Saccharina latissima; (c) dry P. palmata; (d) dry S. latissima; (e) dry Laminaria digitata and (f) dry Grateloupia turuturu. Linear growth rates were similar for all treatments. Specific growth rate was higher in sea urchins fed with fresh P. palmata, but no difference was found between animals fed with fresh S. latissima and those fed with dried diets. Regarding daily food consumption (DFC), sea urchins consumed the same amount of dried macroalgae as fresh but exhibited a higher food conversion efficiency (FCE) when fed with fresh P. palmata. However, this FCE was only significantly higher when compared to sea urchins fed with dry L. digitata. Dried G. turuturu is not a suitable diet due to its rapid degradation after rehydration. The results suggest that P. lividus adults can be reared on dried macroalgae thalli without detriment to their somatic growth, especially over short periods. The low cost of feeding sea urchins with this diet could help small shellfish farmers to diversify their production into echinoculture.     

Experimental rearing of the sea urchin Paracentrotus lividus fed with discards of the lettuce Lactuca sativa in a sea‐based system

Small‐sized specimens (test diameter: 16.64 ± 0.93 mm, mean ± SE) of the sea urchin Paracentrotus lividus were fed with fresh discards of the lettuce Lactuca sativa during a 24‐week experiment in a sea‐based system. At the end of the experiment, stock survival was high (88.8 ± 6.6%). Sea urchin test diameter and total weight grew by 35% and 56%, respectively, while gonad somatic index, after an initial decrease, increased by 3.2%. Moreover, more than 90% of specimens achieved the gonad colour that the market demands. These results support the exploitation of L. sativa as the main ingredient in a manufactured diet for echinoculture of P. lividus. Employing diets formulated with discard ingredients, combined with a low‐cost sea‐based rearing system, could be a suitable approach for sustainable management of echinoculture.     

Outbreak of hirame rhabdovirus infection in cultured spotted sea bass Lateolabrax maculatus on the western coast of Korea

In this study, we determined the cause of a disease outbreak in spotted sea bass, Lateolabrax maculatus reared in culture cages on the western coast of Korea in 2013. The major signs in the diseased fish exhibited were haemorrhaging on the membranes of the abdomen, gastrointestinal organs and opercular gills, as well as an enlarged spleen. No external morphological signs of infection were visible, except for a darkening in colour. No parasites or pathological bacteria were isolated from the diseased fish; however, epithelioma papulosum cyprini (EPC) cells inoculated with tissue homogenates from the diseased fish showed cytopathic effects (CPEs). Virus particles in the EPC cells were bullet‐shaped, 185–225 nm long and 70–80 nm wide, characteristic of Rhabdoviridae. Polymerase chain reaction analyses of homogenized tissues from the diseased fish and supernatants of cell cultures with CPEs indicated specific, 553‐bp‐long fragments corresponding to the matrix protein gene of the hirame rhabdovirus (HIRRV). Phylogenetically, the HIRRV phosphoprotein gene of spotted sea bass was more closely related to phosphoproteins from Chinese and Polish HIRRV strains than from other Korean strains. To our knowledge, this is the first report of HIRRV infection in cultured spotted sea bass.     

Effects of dietary n‐3 to n‐6 fatty acid ratios on spawning performance and larval quality in tongue sole Cynoglossus semilaevis

This study investigated the effect of n‐3 to n‐6 fatty acid ratios in broodstock diets on reproduction performance, fatty acid composition of eggs and gonads of tongue sole Cynoglossus semilaevis. Broodstock were fed five isonitrogenous and isoenergetic diets for 60 days. The supplemented lipids were prepared by a combination of fish oil and soybean oil inclusion FO (fish oil); FSO1 (fish oil: soybean oil = 7:1); FSO2 (fish oil: soybean oil = 2.2:1); FSO3 (fish oil: soybean oil = 1:1); FSO4 (fish oil: soybean oil = 1:4.3) as lipid sources with different n‐3 to n‐6 fatty acid ratios 10.40, 5.21, 2.81, 1.71 and 0.87. Results showed that relative fecundity, fertilization rate and survival rate of larvae at 7 days posthatching were all higher in broodstock fed FSO1 and FSO2 diet and significantly (P < 0.05) decreased in groups fed FSO3 and FSO4 diets. The best result in starvation tolerance test was obtained in FSO2 diet. The present study suggests that n‐3 and n‐6 PUFA ratio in broodstock diet has a considerable effect on spawning performance, egg and larval quality for C. semilaevis.     

The development of a formulated feed containing Ulva (Chlorophyta) to promote rapid growth and enhanced production of high quality roe in the sea urchin Tripneustes gratilla (Linnaeus)

This study investigated growth and gonad production of Tripneustes gratilla fed four protein‐rich artificial diets supplemented with varying amounts (0%, 5%, 15% and 20% weight/weight (w/w); designated 0, 5, 15 and 20U, respectively) of the macroalga Ulva over a 12‐week period. Gonad size, texture, colour and a number of production performance parameters were quantified and compared with urchins fed fresh Ulva (FU) and a combination of FU and artificial feed (FB). All artificial diets significantly (P ≤ 0.05) increased gonad somatic indices (GSI) compared with the FU treatment. The 20U treatment increased GSI by 205% by week 9, compared with a 57% increase in the GSI of urchins fed FU. Gonad colour was calculated using three colour parameters, namely L* (lightness), a* (redness) and b* (yellowness). Whereas L* and a* values did not differ significantly between treatments, b* values for all treatments, with the exception of the 20U and FB treatments, were significantly (P ≤ 0.05) lower or less yellow than the FU treatment by week 12. These results show that we have successfully formulated a feed (20U) which can produce commercially acceptable gonads in terms of both size and colour, indicating the potential for this artificial feed to support commercial echinoculture.     

The effect of fresh seaweed and a formulated diet supplemented with seaweed on the growth and gonad quality of the collector sea urchin,Tripneustes gratilla,under farm conditions

This gonad enhancement study investigates the effect of different fresh and formulated feeds and feeding regimes on the growth and gonad quality of wild‐collected adult sea urchin, Tripneustes gratilla, under farm conditions for over 18 weeks. In the first 12 weeks (phase 1), urchins were fed fresh Ulva rigida (U); a 50:50 mixture of fresh U. rigida and Gracilaria gracilis (UG); fresh G. gracilis (G) and a formulated diet 20U (containing 20% U. rigida), and in the final 6 weeks (phase 2) of the study, diet was changed to a formulated feed (20U diet). By the end of phase 1, urchins fed the 20U diet produced gonads (50.72 ± 5.4 g) that were significantly heavier (p < .001) than the gonads of urchins fed the fresh seaweed diets (U, UG & G). By the end of phase 2, gonad weight of urchins in treatment groups UG‐20U and G‐20U were similar to those fed the 20U‐20U diet. Gonad colour of urchins in the G‐20U treatment became significantly lighter (ANOVA, p = .029) and poorer quality, compared with urchins in the U‐20U group. This gonad enhancement study, conducted on wild‐collected adult T. gratilla, has shown that a formulated feed (20U diet) can enhance gonad growth and produce commercially acceptable gonads. This farm‐based study supports previous findings from aquarium‐based studies by our group and indicates that short‐term sea urchin gonad enhancement can be carried out under farm conditions in South Africa.     

Deposition of conjugated linoleic acid in market size sea bass (Dicentrarchus labrax) and its effects on performance,composition and fillet sensory and texture attributes

The effect conjugated linoleic acid (CLA) accumulation in fish fillet on sensory and texture attributes has been evaluated. Fish were fed diets with graded levels of CLA, which were administered for different durations prior to harvest. Fish were sampled for fillet proximate composition and fatty acid profile for sensory and texture parameters at the end of the trial. Growth performance, feed conversion ratio and muscle proximate composition were not affected by dietary treatments. Muscle fatty acid profile was not affected by dietary treatments despite the increase in saturated fatty acids in fish fed diet with 10 g kg⁻¹ of CLA for 16 weeks before harvest. Besides, the deposition levels of CLA isomers increased gradually depending on the CLA treatment, reaching its maximum level in fish fed 10 g kg⁻¹ CLA diet for 20 weeks before harvest. Fish fed 10 g kg⁻¹ were juicier (P < 0.05) compared with control group, whereas fish fed 5 g kg⁻¹ CLA diet were less (P < 0.05) chewy than fish fed control diet regarding sensory attributes. Texture parameters were not significantly affected by dietary CLA. These results suggest that market size European sea bass could successfully incorporate CLA isomers with some minor positive effects on sensory analyses.     

Effects of dietary fish oil and soya bean lecithin on gonad index,colour and biochemical composition of the purple sea urchin,Strongylocentrotus purpuratus (Stimpson 1857)

The effects of increasing soya bean lecithin (SL) levels (0%, 2% and 4% of diet dry matter, DM) at two fish oil (FO) levels (0% and 3% DM) on gonad index, colour and biochemical composition of Strongylocentrotus purpuratus were evaluated using a 3 × 2 factorial design with six iso‐nitrogenous formulated diets. All diets generated an increase in gonad index of more than 9%. Dietary FO had a positive effect on diet digestibility and accumulation of proteins in the gonad. Soy lecithin did not affect gonad composition, but improved its coloration, decreased gonad feed conversion ratio and increased gonad protein efficiency. There was a clear effect of diets on sea urchins gonad protein, ash and fatty acid content, in particular, decreasing the n‐3/n‐6 ratio associated with a decrease in 20:5n‐3 when SL levels increased. Nevertheless, 22:6n‐3 levels increased by week 12 in all treatments, and diets with SL increased the content of 20:4n‐6 in gonads. A decrease in bitter‐tasting amino acids such as histidine, cysteine, valine and methionine was observed in all treatments with a concomitant increase in isoleucine levels. We recommend using a diet containing 3% of FO and 2% of SL to increase food consumption, diet digestibility, improving marketable gonad colour and an increasing gonad n‐6 fatty acids and C22:6n‐3. In addition, the present study paves the way for future research in the use of FO and SL as an additive in diets for S. purpuratus towards the goal of increasing gonad size and nutritional quality.     

An artificial oyster‐shell reef for the culture and stock enhancement of sea cucumber,Apostichopus japonicus,in shallow seawater

Because of its high nutritional value and health benefits, aquaculture production of Apostichopus japonicus in China is the largest of any single species. Therefore, the development of new farming methods is of considerable significance. In this study, discarded oyster shells have been used to create an artificial reef for the culture of this species. The results have shown that from 6th March 2009 to 26th November 2009, the wet weight of sea cucumber increased from 49.57 ± 1.16 to 79.87 ± 1.46 g ind^?1. Between 16th July and 18th October, the specific growth rate and daily weight gain of A. japonicus differed significantly from other periods. Population density was higher within the reef compared with outside the reef area, and the difference was significant (P < 0.01). The maximum distance between A. japonicus individuals within the reef area on 3rd March and 16th July was 65.0 ± 3.3 and 62.9 ± 4.4 cm, respectively, but the difference was not significant (P > 0.05). The diatom species attached to the oyster shells were similar to those found in the stomach content of A. japonicus. In conclusion, the oyster‐shell reef provides a suitable habitat and shelter for the culture of the sea cucumber, A. japonicus.     

Effects of dietary n‐3 PUFA supplements on composition of n‐3 PUFA and expression of fatty acid elongase 5 (AJELOVL5) in sea cucumber,Apostichopus japonicus

This is the first comprehensive study on the effect of dietary polyunsaturated fatty acid (PUFA) levels on the expression of fatty acid elongase 5 (AJELOVL5), PUFA composition, and growth in juvenile sea cucumbers. The specific growth rate (SGR_w) was improved in n‐3 PUFA‐rich diets compared to low n‐3 PUFA diets. AJELOVL5 expression was apparently upregulated in juveniles fed lower PUFA diets relative to higher PUFA diets, with higher expression in the body wall and respiratory tree of juveniles fed diets without ɑ‐linolenic acid (ALA, 18:3n‐3) compared to juveniles fed higher ALA level diets; similar results were also detected in juveniles fed diets with lower eicosapentaenoic acid (EPA, 20:5n‐3), docosahexaenoic acid (DHA, 22:6n‐3), and none of ALA, EPA, or DHA respectively. The concentrations of ALA, EPA, and DHA in tissues were positively related to the content of dietary corresponding PUFA, with higher ALA content in juveniles fed diet ALA_12.71 than in the ALA_7.46 and ALA₀ groups. Similar results were also obtained in sea cucumber fed diets enriched with either EPA or DHA. Interestingly, considerable levels of EPA and DHA were found in the tissues of juveniles fed diets of CK₀ and DHA₀, with no specific input of EPA or DHA, showing that the sea cucumber was capable of biosynthesizing EPA and DHA from their corresponding precursors as ALA and linoleic acid (LA, 18:2n‐6).     

The use of Kluyveromyces marxianus in the diet of Red‐Stirling tilapia (Oreochromis niloticus,Linnaeus) exposed to natural climatic variation: effects on growth performance,fatty acids,and protein deposition

The purpose of this study was to evaluate the use of single cell proteins (Kluyveromyces marxianus) as fishmeal substitutes for Oreochromis niloticus during long‐term feeding under natural climatic conditions. Juveniles (150) were divided into two groups: the control group, fed with conventional fish diet (T1); and the experimental group, fed with the alternative diet (T2). T1 animals had a higher final weight and per cent weight gain than T2 animals, which we mainly observed in winter and autumn. The lipid content ranged from 1.9% to 3.8% in liver and from 0.2% to 1% in white muscle for both groups. T2 animals had a higher protein concentration in the fillet and the lipid content in this tissue was not altered by diet. Liver and plasma protein content in both groups and the muscle content in T1 decreased during growth. Specific fatty acids (FAs) were selectively retained in fillet phospholipids (PLs), primarily as C22:6n3, which was found at high levels in the PLs of both groups during all seasons. Despite the differences observed, the percentage of polyunsaturated FAs in the fillet was similar throughout the experiment for both diets, reaching approximately 50% of the total FA in the PLs.     

Effect of feeding various levels of poultry by‐product meal on the blood parameters,filet composition and structure of female tenches (Tinca tinca)

Homogeneous background (age, sex, genetic lineage, culture conditions) was created to clearly demonstrate the impact of the tested dietary treatments. No feeds optimized for the rearing of the tench (Tinca tinca L. 1758) are available. Feeds are formulated to increase the growth rate or eliminate skeletal deformations. With the increasing prices of the basic components, fish meal (FM) and fish oil, poultry by‐product meal (PBM) can be used. The aim of the study was to evaluate the effects of substitution of FM with PBM on the tench blood parameters, body composition and structure of skeletal muscles. Cage‐reared female tenches (325 ± 18 g) were fed for 86 days with five types of feeds with 0% (control), 25.7%, 48.6%, 71.4% and 100% substitution of FM with PBM. No significant differences between the formulations were reported for weight, total length, fillet weight, visceral, liver, gonado‐somatic, proximate composition and biochemical blood parameters. However, significant differences were found in the fillet profiles of fatty acids—an increase in the PBM content correlated with an increase in saturated and monounsaturated fatty acids and a decrease in n‐3 PUFA and, generally, n‐6 PUFA. Significant differences were also observed in, for example, the content of intramuscular fatty tissue and the level of organ lipidosis between the control variant (0% PBM) and that with 100% substitution. A sensory assessment indicated a higher gustatory value of the fillets in the case of feeds with 48.6% and 71.4% substitution.