Final ReportSummary - ENRICH (Enrichement of Aquaculture Implants by Introduction of New Marine Species from the Wild to Breeding)
ENRICH project aims at optimising sea urchin Paracentrotus lividus rearing, in order to enrich pre-existing aquaculture SME farms (based on sea bass, bream, cod and/or bivalves) with this sea urchin species, which is not used to its full extent in culture. Sea urchin breeding will be performed starting from adult specimens captured from the wild.
The involved SMEs aim at obtaining the know-how and apply the knowledge acquired from the RTD performers to set up best rearing protocols for larvae, juveniles and adult sea urchins in order to set up sea and land-based implants for commercial exploitation.
The two main objectives are:
1. to insert sea urchin aquaculture in their activities, in order to enrich and diversify their production, that is already going on with different marine species: sea bass / bream and sponges (Italy, REHO), sea bass/bream and seaweed (Israel, ARDAG), sponges and bivalves (Croatia, MILU), cod (UK, VIKI).
2. to obtain an environmentally friendly expansion of their activity, in order to increase sustainability and prevent environmental impact.
Project context and objectives:
Sea urchin roe (gonads) is a luxury food product and is consumed worldwide. Yet, their taste is very sensitive to the adults' diet and this represents the major obstacle in aquaculturing sea urchins. Additionally, the requested taste and consistency of roe is different among the markets in different countries. As far as Europe, sea urchin roe is a largely appreciated culinary delicacy in France, Belgium, Italy, Greece, and its demand is satisfied by importation of different sea urchin species from USA and Chile and, most of all, by harvesting of wild specimens, often leading to a large overexploitation of the resource, where the wild populations are facing serious fishing pressure.
The development of a self-sustained aquaculture activity, independent from the supply of wild specimens, is envisaged from an industrial and conservation perspective, as for other valuable vertebrate and invertebrate species. Consequently, this project arises from the increasing demand for sea urchin roe suitable for European and international markets and it addresses the feasibility of sea urchin aquaculture, enhancing the competitiveness of involved SMEs.
The ENRICH project is envisaged as an effort for developing the technology for land- and sea-based integrated aquacultures of the Mediterranean sea urchin, Paracentrotus lividus, in order to provide high quality and high value products and as such fulfilling the demand of European markets, also in the perspective of helping in protecting an over-exploited species.
The project will explore the possibility to obtain environmentally friendly sea-based aquaculture, by integrated breeding of different species, including the sea urchin P. lividus. The project proposes to create integrated, environmentally friendly aquaculture systems, to substantiate the current knowledge on the reproduction and life cycle in aquaculture conditions of capture-based sea urchins, enhancing knowledge-base required for controlled development of eggs and larvae, and for the development of suitable and environmentally performing feeds. Comparison between land-based and sea-based aquaculture implants will be carried out.
To this aim, we have put together a consortium of excellence, taking together scientists with specific and complementary skills: some of the RTDs have coordinated EU projects in the specific field of aquaculture and environmentally friendly aquaculture.
The project will establish and/or consolidate interactions among RTDs and SMEs involved in fish farming in order to implement the number of commercially relevant marine species, improving SME competitiveness, and to optimise the economic exploitation of sea aquaculture resources.
In terms of conservation, the project will also contribute to the issue of preserving or restoring the natural environment, eventually impacted by the sea urchin sea-based aquaculture systems (diffusion of larvae from sea-based cultured sea urchins)
This project will:
1. improve competitiveness in the aquaculture SME industry;
2. contribute directly to ensuring the sustainability of capture based aquaculture;
3. contribute to conservation of the wild edible sea urchin, Paracentrotus lividus, at present endangered from overfishing and poaching in some Countries (France, Belgium, Southern Italy, Greece, etc.);
4. contribute to production of a high quality roe, suitable for exportation to high consumer countries;
5. contribute to widen the practice of environmentally friendly aquaculture, taking care of avoiding the spreading of adults and larvae from the aquaculture systems to the free environment, where excessive densities of these grazing organisms could have detrimental effects on algal assemblages, reducing their biomass, productivity and biodiversity.
RTDs will set up the theoretical bases and, through the SMEs, carry out pilot experiments for environmentally friendly aquaculture systems, by integrated breeding of different species, such as fish, sea urchins, and macroalgae. The algal growth will be enhanced by using waste nutrients from the aquaculture systems. The starting organisms will be captured from the wild and will be used for establishing the broodstock. Protocols will be produced for optimising broodstock management for gamete quality (WP1). Larvae and juveniles will be produced from the broodstock and rearing protocols will be optimised in order to achieve largest survival and fastest growth of both larvae and juveniles (WP2). Then, adult urchins will be grown out in land- and sea-based aquaculture systems, inserted in 'friendly' polyculture (WP3). The wastes from fish farming will be used to grow macroalgae (e.g. Ulva or Laminaria sp.) that, in addition to the cleaning effect on fish wastes will also constitute the food for adult sea urchins.
Concurrently, a comprehensive study of the physiological regulation of gametogenesis in the sea urchin, Paracentrotus lividus will be performed, comparing different artificial and natural foods (and their combinations) to provide possible tools for controlling gametogenesis in the broodstock and to enable fast gonadial growth rates, achieving marketable gonads in a short time, with high GSI percentage (about 16% of body weight), whose quality will however be close to that of wild sea urchins in colour, flavour, texture and firmness (WP4).
Finally, attention will be focused on reducing the potential environmental impact of such sea-based aquaculture systems, by avoiding the dispersal of larvae from the integrated systems (e.g. removing the individuals when mature, and providing scientific knowledge on the transplantation technique of high value natural macroalgal species (e.g. Cystoseira spp.) and setting up appropriate implantation experiments (WP5).
Project results:
WP1 - Securing seed supply in SME partner bases: Optimising broodstock management and diet for egg quality
Experiments have been run at SAMS, UNIGE, VIKI and UNILE.SAMS has run different trials testing:
1) four different oil sources incorporated into a standardised diet formulation;
2) six diets as in exp. one, using the same basic recipe. However, the lipid source was substituted for a defined fatty acid composition;
3) four diets as in exp. one, using the same basic recipe, but the percentage of lipid inclusion varied between the diets (0 %, 2 %, 4 %, 8 %);
4) four diets produced, that were either low in lipid and low in protein, low in lipid high in protein, high in protein low in lipid, high in protein high in lipid;
5) relationship between nutritional status and gonad development in broodstock urchins was tested in a starvation experiment;
6) animal based artificial diet (including cod liver oil) and vegetable based artificial diet (including flax oil).
VIKI has run experiments aimed at testing effects of commercial artificial diets with different lipid content compared with kelp, as a control (low in lipids). Those urchins fed on the kelp diet were significantly more fecund than the artificial diets, although gonad growth was more pronounced on the high lipid artificial diet. However, for female urchins the high lipid diets (treatments A&B) resulted in earlier maturation of the gonads compared to the other diets.
The fatty acid composition of the eggs in terms of the essential fatty acid content reflected that of the diet for the four experimental treatments.
UNIGE, in the second year run experiments comparing commercial artificial diets to a home made fresh diet based on spinach, carrots and eggs. The home made diet performed better at a temperature of 15 - 18 °C, producing 30 % of spawning (male and female equally) in one month. While mix and commercial pellet diets did not produce spawning specimens. At higher temperature (above 20 °C) the home made diet did not work so good, probably because of faster degradation. A mixed diet (50 % home made - 50 % commercial pellets) seems to have the same results of the home made diet, at a reduced amount (15 g / week / specimen instead of 30 g / week / specimen).
UNILE has assessed lipid profiles of a diet formulation produced by UNIGE.
Overall results have shown that diet quality has a direct link to gonad development, gonad size, and sexual maturation. As such, broodstock conditioning using selected and tailored diets will provide a useful tool for the industry to ensure maximum fecundity and control over the gametogentic cycles.
A variety of diets can be used for effective broodstock conditioning with lipid contents in the range of 6 - 8 %. However, there is strong evidence to suggest that the micro nutrients present in macroalgae may play a crucial role in fecundity and fertilisation success. For this reason, it is recommended that macro be alongside or incorporated into the broodstock conditioning diet.
Larval survival and development seems to be independent of broodstock conditioning suggesting that external nutrition for P. lividus larvae is more important than maternal provisioning. This has implications for the formulation of conditioning diets. P. lividus is capable of synthesising a number of 20 carbon chain polyunsaturated fatty acids from 18 carbon or shorter fatty acids. For this reason and the fact that maternal diets seems to have limited impact on larval survival and development, the inclusion of expensive fish oils in the broodstock conditioning diet is not necessary.
The gonad development in P. lividus responds to changes in diet very rapidly, often within weeks or months, as such conditioning diets need not be utilised for extended periods prior to spawning. A period of two to three months on the condition diet should be sufficient to ensure that the full benefits of the diet are observed.
WP2 - Optimisation of larval rearing, metamorphosis and juvenile growth
Experiments have been run at UNIGE, VIKI and SAMS, regarding:
1) Larval rearing conditions
Physical stress caused by water changes in larval tanks enhance larval mortality, thus higher survival has also been obtained reducing the number of water changes. The use of flow through systems and/or of bottles with internal smooth surface in order to reduce microfouling settling, proved appropriated for the purpose. Also the surface / volume ratio of the tanks was tested in order to have best performance of larvae: high S/V ratios turned to work better. Results obtained so far have allowed to significantly increase larval survival. This is particularly important to the aquaculture aims, as the larval stages are the most delicate and sensitive to possible crashes due to different causes, among which bacterial diseases or culture contamination by other invertebrates.
2) Larval feeding with natural diets
Experiments run at UNIGE and VIKI proved the better performance of the microalga Cricosphaera elongata, instead of the traditionally used Dunaliella tertiolecta. In particular an experiment was run at VIKI investigating the growth, survival and fatty acid composition of P. lividus larvae fed four microalgal diets: Cricosphaera elongata, Pleurochrysis carterae, Tetraselmis suecica and Dunaliella tertiolecta (control). 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 development 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), eicosapentaenoate (EPA; 20:5n-3) and arachidonate (ARA; 20:4n-6), either by assimilation and retention of dietary fatty acids, and/or synthesis from a-linolenic acid 18:3n-3 and linoleic acid 18:2n-6. Moreover, an accumulation of n-3 LC-PUFA and higher EPA/DHA and EPA/ARA ratios appeared to be associated with improved larval 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.
3) Larval feeding with artificial diets
Experiments have been run at SAMS. Although artificial diets have shown to allow completion of the larval stages of P. lividus under laboratory conditions, our studies have shown that under semi commercial and commercial scales of productions, artificial diets have a number of shortcomings that may prevent them from replacing microalgae as the industry standard for larval production. These include the following:
- Particle size: sub 32 micron particle produce the best results. However, microencapsulated particles of this size are difficult and expensive to manufacture.
- Availability of the artificial diets to the larvae within the culture vessel, because of negative buoyancy of commercial microencapsulated diets, that fast sink out of the water column and become unavailable to the larvae.
- Enrichment of sea urchin larvae with PUFA emulsions may provide a strategy to shorten larval production cycle and may provide direct benefit to the industry and would be a good target for further research and development.
4) Optimisation of larval settling and juvenile growth
Experiments were run at UNIGE. Metamorphosis on the mixed diatom bed ranges between 40 and 60 %, metamorphosis on marinated substrates (plastic panels) is about 50 %, while metamorphosis on vegetated stones from the wild is about 70 %. The safest stocking size seems to be over 1 cm.
Overall results have shown that:
Physical stress caused by water changes in larval tanks enhance larval mortality, thus higher survival has also been obtained reducing the number of water changes. The use of flow through systems and/or of bottles with internal smooth surface in order to reduce microfouling settling, proved appropriated for the purpose. Also the surface/volume ratio of the tanks was tested in order to have best performance of larvae: high S/V ratios turned to work better. Results obtained so far have allowed to significantly increase larval survival. This is particularly important to the aquaculture aims, as the larval stages are the most delicate and sensitive to possible crashes due to different causes, among which bacterial diseases or culture contamination by other invertebrates.
For every system, water should have a temperature of 18 ± 2 °C and should be sterilised. For commercial purpose, we suggest the use of 0.1 ml/L of hypochlorite (12 - 15 %). After 24 hours, it is needed to add 0.1 ml/L of sodium thiosulfate solution (154 g/L of distilled water).
Cricosphaera elongata resulted to be the best food to be used, instead of the traditionally used Dunaliella tertiolecta.
Although artificial diets have shown to allow completion of the larval stages of P. lividus under laboratory conditions, our studies have shown that under semi commercial and commercial scales of productions, artificial diets have a number of shortcomings that may prevent them from replacing microalgae as the industry standard for larval production. These include the following:
- Particle size: Sub 32 micron particle produce the best results. However, microencapsulated particles of this size are difficult and expensive to manufacture.
- Availability of the artificial diets to the larvae within the culture vessel, because of negative buoyancy of commercial microencapsulated diets, that fast sink out of the water column and become unavailable to the larvae.
- Enrichment of sea urchin larvae with PUFA emulsions may provide a strategy to shorten larval production cycle and may provide direct benefit to the industry and would be a good target for further research and development.
WP3 - Sea Urchin - juvenile to adult growth in integrated systems -pilot scale
Transplantation to farming conditions in three different systems:
O3.1 Land-based integrated systems with bass / bream seaweeds (Israel: IOLR and ARDAG)
O3.2 Farming at sea integrated systems in the Atlantic (Scotland)
a) cod and seaweeds (SAMS and VIKI)
O3.3 Farming at sea integrated systems in the Mediterranean
a) bass / bream seaweeds (Italy: UNILE/UNIGE and REHO/AQUA)
b) sponges and mussels (Croatia: IMP and MILU).
IOLR/ARDAG integrated multi-trophic system (IMTA)
Effluent water from the super-intensive fish farm ponds of ARDAG was drained into an Ulva biofilter and production unit. The enriched Ulva was used as a food source for the sea urchins which were grown in a separate compartment. The performance (growth rates, GSI, survival and gonad colour) of Paracentrotus lividus was measured during their culture at the IMTA for thirty months. During this period, five feeding and three density experiments were carried out in the sea urchins compartment of the system.
As far as feeds, urchins were exposed to: raw Ulva from the biofilter compartment and various ratios of dried Ulva as a supplement to our traditional prepared (pelleted) diet. Pellet diets best contributed to gonad growth. FCR was around 12 in the algae diet (WW) and ranged 3 - 4 in the pelleted diet. Because the pellet diet performer best in GSI growth and the seaweed in gonad colour, to create the option for sea urchins to eat the pellet diet while also gaining the benefits of algae, we combined Ulva into the pellet feed formulation (up to 50 %).
As far as density effects, the effect of stocking densities was tested on juvenile urchins (0.3 - 3 gr). Ulva was used as a food for the urchins. Survival rate was much higher (18 %) in the lower density. Animals are density dependents in term of survival. Optimal suggested stocking densities are: 5000 ind m-2 up to 1gr 1000 ind. m-2 up to 3 gr, 250 ind m-2 up to 10 gr, and 125 ind m-2 to commercial size 30 - 50gr.
VIKI-SAMS:
As currently there are cod at sea in Scotland, work concentrated on designing, constructing and trialling an integrated onshore system for the production of sea urchins and kelp, using the effluent from on shore cod broodstock. The system was designed to allow a comparison between a system that utilises the effluent stream compared to one that did not. Preliminary results show that on shore cod production generates nutrient rich waters suitable for the production of sea urchins and macroalgae; Nitrogen utilisation within the raceway continues throughout the full 24 hour cycle; the poly-tunnel integrated aquaculture system may be a suitable model for the development of land-based sea urchin and macroalgae production in northern latitudes across Europe.
UNIGE, UNILE have been performing activities in off-shore implants. The first trials have been done in REHO facilities in the first year and at AQUA in the second one. At AQUA, special barrels where used to maintain specimens at 8 m depth hanging at the support ropes of the fish cages. We put in the cages adult specimens collected in the wild. The survival ratio, after 45 days was 95 %. Three feeding treatments have been set up (unfed except from fish wastes, fish pellets, macroalgae). The results were quite promising, particularly if the deployment time of the cage would have been since earlier in the project execution.
IMP-MILU:
experiments were set up both at sea (in Mali Ston Bay) and in an integrated system in-land. Experimental grow out at sea was performed according different feeds (Ulva or grazing biofilm), alongside mussel ropes or in cages, and at different depths. Best results were achieved at 3 m depth. Yet, best results in growth performance and GSI values were achieved in the in-land system, where urchins were fed enriched Ulva from the fish wastes.
WP4 - Gonad improvement (color and GSI) and control gametogenesis
The following experiments have been performed at IOLR:
1) effect of photoperiod manipulation on body growth and gonadal development;
2) diet effects on gonadal growth and gametogenesis;
3) effects of mammalian GnRH agonist on steroidogenesis in cultivated P. lividus populations;
4) effects of sex steroids on gonadal development in cultivated P. lividus populations.
Our study evaluated the effects of environmental conditions (i.e. photoperiod and diet) and those of exogenous hormones (i.e. exposure via water to GnRH agonist or via food to sex steroids), on gonadal development in cultivated P. lividus populations. Additionally, aiming at identifying endogenous factors that signify the onset of gametogenesis, we studied temporal profiles of:
(i) sex steroids;
(ii) transcript levels of dopamine receptors; and
(iii) transcript levels and enzymatic activity of cathepsins, the major proteases involved in yolk processing.
Exposure of adult P. lividus population to 24-h dark [D]or 24-h light [L], indicated no significant differences in terms of body growth or gonadosomatic index (GSI). Nevertheless, the manipulated photoperiod did affect gametogenesis. On one hand, the D regime induced reproductive development, giving rise to multiple reproductive cycles, which are relatively shorter compared to those of wild urchins. On the other hand, the L regime reduced the rates of gametogenesis. The latter effect was more pronounced in males than in females. Yet, on a long-term, acclimation of the urchins to the L regime is observed, emphasising the limitation of the manipulated photoperiod as means to control gametogenesis in P. lividus. Such a method is only effective in short-term applications.
The diet experiment shows that Ulva lactuca (U), which is considered to be a natural food resource for wild P. lividus stocks, significantly increased P. lividus body mass (BW) compared to the formulated dry pellets (P). Nevertheless, the P diet significantly increased the urchins' gonadal growth (GSI) as compared to the U diet. These contrasting effects draw attention to the advantage of using the combination of both diets. Hence, further optimisation of the dietary ratio between fresh enriched alga and formulated dry pellets, is expected to shorten the growth phase and give rise to high quality marketable product.
Our results also reveal sexual-dimorphic patterns, favouring female populations attributable to their higher GSI values. Hence, future research focused on implementing genetic manipulations in sea urchins, should aid in establishing the desired all-female P. lividus populations.
The studies with exogenous hormones indicate that despite the evolutionary distance separating echinoderms from vertebrates, both phyla share some conserved neuroendocrine elements, which are involved in the control of reproduction. Thus, we demonstrated that the vertebrate GnRH ligand can activate P. lividus GnRH receptor(s), and consequently elicit stimulated gonadal sex steroids (i.e. testosterone and estradiol). The obtained stage-dependent profiles further link sex steroid dopamine and cathepsins (mainly B and L) to the onset of gametogenesis. Beside their contribution to the understanding of mechanisms that controls the onset of gametogenesis in P. lividus, our data underscore aspects, which are oriented to aquaculture applicability. One such aspect could be the use of food supplements (approved for human consumption) in order to retard / inhibit steroidogenesis and/or yolk protein degradation by cathepsins. Once optimised, such diets are expected to hold back the onset of gametogenesis, enhance the somatic growth, and widen the period during which high quality roe is being preserved.
WP5 - Environmental quality assessment
Main objectives of WP5 refer to culture Ulva sp. in a biofiltering system using wastewaters and monitoring urchin population and environmental conditions in the ranching areas.
As far as Ulva as a biofilter, activities were fundamentally carried out at IOLR-ARDAG. The performance parameters of Ulva included growth rate, yield, biochemical composition and nutrient uptake. Effluent water from a semi-closed super-intensive fish pond system was drained continuously into Ulva reactors. Nitrogen removal dynamics in a batch flow system for seven days was evaluated. A gradual reduction of TDN from 600 µm to almost 0 within 6 days is observed. Three stages could be identified: rapid reduction of ammonia within 24 h, 48 h of lag with no nitrogen removal, and 36 hours reduction of nitrate. Ulva shows a preference for ammonium compared to nitrates because the nitrogen in the ammonium is in a reduced state that algae can directly utilise for production of amino acids. Nitrates start to be depleted from water after the complete removal of ammonium from the water. This shift takes around 48 hours, the time needed to form active nitrate reductase because active nitrate reductase is not formed in the presence of ammonium. The nitrate then must first be reduced to nitrite, and then ammonium.
Since aeration of the Ulva compartment constitutes about 85 % of the operating cost (energy), we tried to reduce cost by reducing aeration from 100 % to 50 % or 25 % of the time. We evaluated Ulva performance when exposed to these three aeration regimes. A significant increase in Ulva yield is observed between the 50 % and the 100 % aeration regimes. 25 % aeration is the optimal aeration regime for maximal nitrogen removal efficiency (3. 5 g N M-2 d-1), and relatively low energy cost. 100 % aeration regime is less economic but produces maximal yield (178±25 g WW m-2 d-1).
Ulva rearing experiments were performed also at IMP and UNIGE-AQUA. As far as the former, attention was given to the biochemical composition of reared Ulva compared to the wild one, stressing, even in short term experiments, a large enrichment of reared Ulva in terms of organic content compared to ash and moisture. As far as UNIGE-AQUA algae were cultured according to two different treatments, plastic panles and nets, hanging along the support ropes of the AQUA fish cages, at around 1.5 m depth. Nets provided highest macroalgal yield compared to plastic panels, showing biomass values doubled, in spring time, in a month time span.
As far as the natural populations around sea based implants, surveys have been carried out from IMP-MILU and UNIGE-UNILE-AQUA (in the second year, while UNIGE-UNILE-REHO in the first one). Mali Ston Bay is a shallow bay characterised by numerous underwater freshwater springs, which make it very suitable for mariculture. The whole bay is very populated with Paracentrotus lividus, with densities locally exceeding 15 ind/m2. Most abundant size class is 3 - 4.5 cm, average test diameter is 3.7 ± 0.6 cm. According to such population parameters, Mali Ston populations exhibit large size, although a slower growth rate, compared to other Mediterranean areas. GSI varies through the year, with one value peak during April, when spawning occurs, confirmed by histological surveys.
Results from REHO site have been reported in the first reporting report. As far as AQUA, four sampling sites have been chosen, two south of the implants and two north of it. At each site urchins have been visually sampled at the start of experimental culturing and at the end (around two months), in terms of density and size structure, while specimens have been collected at each site on a fortnight base for gonad condition analyses, carried out in terms of GSI and histology (UNIGE) and lipid profile (UNILE) and compared to cultured specimens. Environmental features at the AQUA implant have been regularly monitored.
Potential impact:
P. lividus roe (the red gonads) are very prized, but at the present time in Europe there is an important exploitation in France and Belgium, a moderate one in Italy and Spain and a non-significant, but fast growing one in the rest of Mediterranean Europe, where fishery assumes a seasonal, tourist role. However, the world demand for urchin roe remains very high and searching for new markets. Such a great exploitation has made necessary protection by policy organs:
Already listed within species whose exploitation is regulated in specially protected areas (MAP Phase II, Monaco 24/11/1996), Paracentrotus lividus has now been included among the protected fauna species (Council of Europe, Strasbourg, 3/1998, Appendix III).
In recent years, a new aquaculture activity has developed directed at highly appreciated species for which the current level of production by commercial fisheries is not enough to meet consumers' demand and where the wild populations are facing serious fishing pressure.
ENRICH will provide an essential knowledge base for enrichment of aquaculture farms by the introduction of the European sea urchin, Paracentrotus lividus. In addition the project will develop and test pilot experiments in order to verify the possible commercial exploitation of this valuable species.
The project aimed at substantiating the current knowledge on the reproduction of capture based aquaculture species in captivity, through objectives of WP1 and WP2, establishing knowledge-base required for controlled development of eggs and larvae, and for the development of suitable and environmentally performing feeds. ENRICH fulfils this demand, as the major innovation of this proposal is be improvement of eggs and consequently of larval development, through the controlled quality of food to be supplied in order to optimise broodstock.
The development of a self-sustained aquaculture activity, independent from the supply of wild fish, is thus required both from an industrial and conservation perspective. Thus, the relevance of the introduction of this species into aquaculture farms, either land-based or sea-based, as envisaged in WP3, is at present very high.
WP4 has provided fundamental tools for controlling gametogenesis both for optimising the gamete production of the broodstock and for optimising the gonads of cultured individuals for satisfying the market requirements.
In addition, through WP5, attention has been given to produce macroalgae in integrated systems, both for 'cleaning' effluent waters form fish farming and for feeding urchins with a protein enriched diet. On the other side, WP5 has gathered information on wild stocks, comparing their gonad conditions to cultured specimens and paid attention on possible impacts on the environment in the areas surrounding the sea-based aquaculture systems, contributing directly to ensure the sustainability of capture based aquaculture.
The involved partnership is important to cover the extent of P. lividus distribution in the Mediterranean and the north Atlantic, encompassing a large range of environmental conditions, thus testing feeding and culturing systems in different environmental conditions.
All of the partners bring to the consortium excellence in the field of aquaculture. ENRICH will consequently create competitiveness in the aquaculture of sea urchins.
The SME participants will profit of this work for improving and amplifying the field of work, and explore new markets. Two of them have started the insertion of sea urchins in the aquaculture ranching (AQUA and MILU), and two of them already have experience and their aim is to maximise results in terms of production and quality (VIKI and ARDAG). In Europe, relevant markets are represented by France, Belgium, Holland and also Southern Italy, Greece, Spain.
Local and exportation products will be represented by living samples, but also canned or frozen roe, prepared recipes to be used for particular appetisers and first courses, by cooperation with other differentiated enterprises. Some of the SMEs , in particular VIKI, are interested to produce seeds (sea urchins up to 1 cm diameter), to be delivered to European aquaculturers (Spain, France, Italy, Belgium, Greece) for further growth, and to protection agencies for recovery of depleted areas. All the above will enlarge the field and the occupation of operators.
The SMEs will also profit of the new knowledge to sell it to other enterprises, via the production of CDs covered by copyright (mainly for broodstock improvement), thus increasing European competitiveness, as at present the major exportation to eastern Countries is from California and Chile.
In prospective, following the project completion, touristic exploitation is also planned, as tourists visit to the aquaculture sites, followed by product degustation will be inserted among the touristic attraction of the respective regions.
The project will also have indirect positive effects in terms of environmental protection, as the possibility of successfully rear larvae and juveniles on a large scale will provide a tool for possible restocking of overexploited populations throughout the P. lividus distribution range.
Dissemination activities have been performed at different levels:
1) publication on scientific journals and on journals particularly addressed to the aquaculture world;
2) participation to a wide range of conferences (particularly to aquaculture conferences, in Europe and in other extra-european countries) through oral and poster presentations;
3) participation to specific workshops on P. lividus resource management;
4) organisation of a final workshop of the project in the context of a large scale event, such as the SLOWFISH event in Genova (29 May 2011), gathering scientists from Europe and USA and showing to the wide public the results of the project through digital objects;
5) setting up of a web site and a flyer showing main targets of the project;
6) organising a number of dissemination activities for schools, visiting the implants;
7) compiling a large number of bachelor and master theses and participating to the fulfillment of two PhD thesis;
8) a complete list of the dissemination activities is provided below in the list of dissemination activities.
Exploitation of the results of the project may been fundamentally summarised as follows:
1) establishment of broodstocks for the different SMEs involved in the projects;
2) production of different protocols for optimising the different phases of sea urchin culturing, from the management of the broodstock, to the larval and juvenile phases and a summary of results achieved for the grow out phase, both inland and at sea, in sheltered and exposed areas, also with the support of the results from the gametogenic control.
The SMEs partners of the project will make use of such protocols in order to enhance their production.
Project website: http://www.enrichnet.eu
The involved SMEs aim at obtaining the know-how and apply the knowledge acquired from the RTD performers to set up best rearing protocols for larvae, juveniles and adult sea urchins in order to set up sea and land-based implants for commercial exploitation.
The two main objectives are:
1. to insert sea urchin aquaculture in their activities, in order to enrich and diversify their production, that is already going on with different marine species: sea bass / bream and sponges (Italy, REHO), sea bass/bream and seaweed (Israel, ARDAG), sponges and bivalves (Croatia, MILU), cod (UK, VIKI).
2. to obtain an environmentally friendly expansion of their activity, in order to increase sustainability and prevent environmental impact.
Project context and objectives:
Sea urchin roe (gonads) is a luxury food product and is consumed worldwide. Yet, their taste is very sensitive to the adults' diet and this represents the major obstacle in aquaculturing sea urchins. Additionally, the requested taste and consistency of roe is different among the markets in different countries. As far as Europe, sea urchin roe is a largely appreciated culinary delicacy in France, Belgium, Italy, Greece, and its demand is satisfied by importation of different sea urchin species from USA and Chile and, most of all, by harvesting of wild specimens, often leading to a large overexploitation of the resource, where the wild populations are facing serious fishing pressure.
The development of a self-sustained aquaculture activity, independent from the supply of wild specimens, is envisaged from an industrial and conservation perspective, as for other valuable vertebrate and invertebrate species. Consequently, this project arises from the increasing demand for sea urchin roe suitable for European and international markets and it addresses the feasibility of sea urchin aquaculture, enhancing the competitiveness of involved SMEs.
The ENRICH project is envisaged as an effort for developing the technology for land- and sea-based integrated aquacultures of the Mediterranean sea urchin, Paracentrotus lividus, in order to provide high quality and high value products and as such fulfilling the demand of European markets, also in the perspective of helping in protecting an over-exploited species.
The project will explore the possibility to obtain environmentally friendly sea-based aquaculture, by integrated breeding of different species, including the sea urchin P. lividus. The project proposes to create integrated, environmentally friendly aquaculture systems, to substantiate the current knowledge on the reproduction and life cycle in aquaculture conditions of capture-based sea urchins, enhancing knowledge-base required for controlled development of eggs and larvae, and for the development of suitable and environmentally performing feeds. Comparison between land-based and sea-based aquaculture implants will be carried out.
To this aim, we have put together a consortium of excellence, taking together scientists with specific and complementary skills: some of the RTDs have coordinated EU projects in the specific field of aquaculture and environmentally friendly aquaculture.
The project will establish and/or consolidate interactions among RTDs and SMEs involved in fish farming in order to implement the number of commercially relevant marine species, improving SME competitiveness, and to optimise the economic exploitation of sea aquaculture resources.
In terms of conservation, the project will also contribute to the issue of preserving or restoring the natural environment, eventually impacted by the sea urchin sea-based aquaculture systems (diffusion of larvae from sea-based cultured sea urchins)
This project will:
1. improve competitiveness in the aquaculture SME industry;
2. contribute directly to ensuring the sustainability of capture based aquaculture;
3. contribute to conservation of the wild edible sea urchin, Paracentrotus lividus, at present endangered from overfishing and poaching in some Countries (France, Belgium, Southern Italy, Greece, etc.);
4. contribute to production of a high quality roe, suitable for exportation to high consumer countries;
5. contribute to widen the practice of environmentally friendly aquaculture, taking care of avoiding the spreading of adults and larvae from the aquaculture systems to the free environment, where excessive densities of these grazing organisms could have detrimental effects on algal assemblages, reducing their biomass, productivity and biodiversity.
RTDs will set up the theoretical bases and, through the SMEs, carry out pilot experiments for environmentally friendly aquaculture systems, by integrated breeding of different species, such as fish, sea urchins, and macroalgae. The algal growth will be enhanced by using waste nutrients from the aquaculture systems. The starting organisms will be captured from the wild and will be used for establishing the broodstock. Protocols will be produced for optimising broodstock management for gamete quality (WP1). Larvae and juveniles will be produced from the broodstock and rearing protocols will be optimised in order to achieve largest survival and fastest growth of both larvae and juveniles (WP2). Then, adult urchins will be grown out in land- and sea-based aquaculture systems, inserted in 'friendly' polyculture (WP3). The wastes from fish farming will be used to grow macroalgae (e.g. Ulva or Laminaria sp.) that, in addition to the cleaning effect on fish wastes will also constitute the food for adult sea urchins.
Concurrently, a comprehensive study of the physiological regulation of gametogenesis in the sea urchin, Paracentrotus lividus will be performed, comparing different artificial and natural foods (and their combinations) to provide possible tools for controlling gametogenesis in the broodstock and to enable fast gonadial growth rates, achieving marketable gonads in a short time, with high GSI percentage (about 16% of body weight), whose quality will however be close to that of wild sea urchins in colour, flavour, texture and firmness (WP4).
Finally, attention will be focused on reducing the potential environmental impact of such sea-based aquaculture systems, by avoiding the dispersal of larvae from the integrated systems (e.g. removing the individuals when mature, and providing scientific knowledge on the transplantation technique of high value natural macroalgal species (e.g. Cystoseira spp.) and setting up appropriate implantation experiments (WP5).
Project results:
WP1 - Securing seed supply in SME partner bases: Optimising broodstock management and diet for egg quality
Experiments have been run at SAMS, UNIGE, VIKI and UNILE.SAMS has run different trials testing:
1) four different oil sources incorporated into a standardised diet formulation;
2) six diets as in exp. one, using the same basic recipe. However, the lipid source was substituted for a defined fatty acid composition;
3) four diets as in exp. one, using the same basic recipe, but the percentage of lipid inclusion varied between the diets (0 %, 2 %, 4 %, 8 %);
4) four diets produced, that were either low in lipid and low in protein, low in lipid high in protein, high in protein low in lipid, high in protein high in lipid;
5) relationship between nutritional status and gonad development in broodstock urchins was tested in a starvation experiment;
6) animal based artificial diet (including cod liver oil) and vegetable based artificial diet (including flax oil).
VIKI has run experiments aimed at testing effects of commercial artificial diets with different lipid content compared with kelp, as a control (low in lipids). Those urchins fed on the kelp diet were significantly more fecund than the artificial diets, although gonad growth was more pronounced on the high lipid artificial diet. However, for female urchins the high lipid diets (treatments A&B) resulted in earlier maturation of the gonads compared to the other diets.
The fatty acid composition of the eggs in terms of the essential fatty acid content reflected that of the diet for the four experimental treatments.
UNIGE, in the second year run experiments comparing commercial artificial diets to a home made fresh diet based on spinach, carrots and eggs. The home made diet performed better at a temperature of 15 - 18 °C, producing 30 % of spawning (male and female equally) in one month. While mix and commercial pellet diets did not produce spawning specimens. At higher temperature (above 20 °C) the home made diet did not work so good, probably because of faster degradation. A mixed diet (50 % home made - 50 % commercial pellets) seems to have the same results of the home made diet, at a reduced amount (15 g / week / specimen instead of 30 g / week / specimen).
UNILE has assessed lipid profiles of a diet formulation produced by UNIGE.
Overall results have shown that diet quality has a direct link to gonad development, gonad size, and sexual maturation. As such, broodstock conditioning using selected and tailored diets will provide a useful tool for the industry to ensure maximum fecundity and control over the gametogentic cycles.
A variety of diets can be used for effective broodstock conditioning with lipid contents in the range of 6 - 8 %. However, there is strong evidence to suggest that the micro nutrients present in macroalgae may play a crucial role in fecundity and fertilisation success. For this reason, it is recommended that macro be alongside or incorporated into the broodstock conditioning diet.
Larval survival and development seems to be independent of broodstock conditioning suggesting that external nutrition for P. lividus larvae is more important than maternal provisioning. This has implications for the formulation of conditioning diets. P. lividus is capable of synthesising a number of 20 carbon chain polyunsaturated fatty acids from 18 carbon or shorter fatty acids. For this reason and the fact that maternal diets seems to have limited impact on larval survival and development, the inclusion of expensive fish oils in the broodstock conditioning diet is not necessary.
The gonad development in P. lividus responds to changes in diet very rapidly, often within weeks or months, as such conditioning diets need not be utilised for extended periods prior to spawning. A period of two to three months on the condition diet should be sufficient to ensure that the full benefits of the diet are observed.
WP2 - Optimisation of larval rearing, metamorphosis and juvenile growth
Experiments have been run at UNIGE, VIKI and SAMS, regarding:
1) Larval rearing conditions
Physical stress caused by water changes in larval tanks enhance larval mortality, thus higher survival has also been obtained reducing the number of water changes. The use of flow through systems and/or of bottles with internal smooth surface in order to reduce microfouling settling, proved appropriated for the purpose. Also the surface / volume ratio of the tanks was tested in order to have best performance of larvae: high S/V ratios turned to work better. Results obtained so far have allowed to significantly increase larval survival. This is particularly important to the aquaculture aims, as the larval stages are the most delicate and sensitive to possible crashes due to different causes, among which bacterial diseases or culture contamination by other invertebrates.
2) Larval feeding with natural diets
Experiments run at UNIGE and VIKI proved the better performance of the microalga Cricosphaera elongata, instead of the traditionally used Dunaliella tertiolecta. In particular an experiment was run at VIKI investigating the growth, survival and fatty acid composition of P. lividus larvae fed four microalgal diets: Cricosphaera elongata, Pleurochrysis carterae, Tetraselmis suecica and Dunaliella tertiolecta (control). 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 development 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), eicosapentaenoate (EPA; 20:5n-3) and arachidonate (ARA; 20:4n-6), either by assimilation and retention of dietary fatty acids, and/or synthesis from a-linolenic acid 18:3n-3 and linoleic acid 18:2n-6. Moreover, an accumulation of n-3 LC-PUFA and higher EPA/DHA and EPA/ARA ratios appeared to be associated with improved larval 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.
3) Larval feeding with artificial diets
Experiments have been run at SAMS. Although artificial diets have shown to allow completion of the larval stages of P. lividus under laboratory conditions, our studies have shown that under semi commercial and commercial scales of productions, artificial diets have a number of shortcomings that may prevent them from replacing microalgae as the industry standard for larval production. These include the following:
- Particle size: sub 32 micron particle produce the best results. However, microencapsulated particles of this size are difficult and expensive to manufacture.
- Availability of the artificial diets to the larvae within the culture vessel, because of negative buoyancy of commercial microencapsulated diets, that fast sink out of the water column and become unavailable to the larvae.
- Enrichment of sea urchin larvae with PUFA emulsions may provide a strategy to shorten larval production cycle and may provide direct benefit to the industry and would be a good target for further research and development.
4) Optimisation of larval settling and juvenile growth
Experiments were run at UNIGE. Metamorphosis on the mixed diatom bed ranges between 40 and 60 %, metamorphosis on marinated substrates (plastic panels) is about 50 %, while metamorphosis on vegetated stones from the wild is about 70 %. The safest stocking size seems to be over 1 cm.
Overall results have shown that:
Physical stress caused by water changes in larval tanks enhance larval mortality, thus higher survival has also been obtained reducing the number of water changes. The use of flow through systems and/or of bottles with internal smooth surface in order to reduce microfouling settling, proved appropriated for the purpose. Also the surface/volume ratio of the tanks was tested in order to have best performance of larvae: high S/V ratios turned to work better. Results obtained so far have allowed to significantly increase larval survival. This is particularly important to the aquaculture aims, as the larval stages are the most delicate and sensitive to possible crashes due to different causes, among which bacterial diseases or culture contamination by other invertebrates.
For every system, water should have a temperature of 18 ± 2 °C and should be sterilised. For commercial purpose, we suggest the use of 0.1 ml/L of hypochlorite (12 - 15 %). After 24 hours, it is needed to add 0.1 ml/L of sodium thiosulfate solution (154 g/L of distilled water).
Cricosphaera elongata resulted to be the best food to be used, instead of the traditionally used Dunaliella tertiolecta.
Although artificial diets have shown to allow completion of the larval stages of P. lividus under laboratory conditions, our studies have shown that under semi commercial and commercial scales of productions, artificial diets have a number of shortcomings that may prevent them from replacing microalgae as the industry standard for larval production. These include the following:
- Particle size: Sub 32 micron particle produce the best results. However, microencapsulated particles of this size are difficult and expensive to manufacture.
- Availability of the artificial diets to the larvae within the culture vessel, because of negative buoyancy of commercial microencapsulated diets, that fast sink out of the water column and become unavailable to the larvae.
- Enrichment of sea urchin larvae with PUFA emulsions may provide a strategy to shorten larval production cycle and may provide direct benefit to the industry and would be a good target for further research and development.
WP3 - Sea Urchin - juvenile to adult growth in integrated systems -pilot scale
Transplantation to farming conditions in three different systems:
O3.1 Land-based integrated systems with bass / bream seaweeds (Israel: IOLR and ARDAG)
O3.2 Farming at sea integrated systems in the Atlantic (Scotland)
a) cod and seaweeds (SAMS and VIKI)
O3.3 Farming at sea integrated systems in the Mediterranean
a) bass / bream seaweeds (Italy: UNILE/UNIGE and REHO/AQUA)
b) sponges and mussels (Croatia: IMP and MILU).
IOLR/ARDAG integrated multi-trophic system (IMTA)
Effluent water from the super-intensive fish farm ponds of ARDAG was drained into an Ulva biofilter and production unit. The enriched Ulva was used as a food source for the sea urchins which were grown in a separate compartment. The performance (growth rates, GSI, survival and gonad colour) of Paracentrotus lividus was measured during their culture at the IMTA for thirty months. During this period, five feeding and three density experiments were carried out in the sea urchins compartment of the system.
As far as feeds, urchins were exposed to: raw Ulva from the biofilter compartment and various ratios of dried Ulva as a supplement to our traditional prepared (pelleted) diet. Pellet diets best contributed to gonad growth. FCR was around 12 in the algae diet (WW) and ranged 3 - 4 in the pelleted diet. Because the pellet diet performer best in GSI growth and the seaweed in gonad colour, to create the option for sea urchins to eat the pellet diet while also gaining the benefits of algae, we combined Ulva into the pellet feed formulation (up to 50 %).
As far as density effects, the effect of stocking densities was tested on juvenile urchins (0.3 - 3 gr). Ulva was used as a food for the urchins. Survival rate was much higher (18 %) in the lower density. Animals are density dependents in term of survival. Optimal suggested stocking densities are: 5000 ind m-2 up to 1gr 1000 ind. m-2 up to 3 gr, 250 ind m-2 up to 10 gr, and 125 ind m-2 to commercial size 30 - 50gr.
VIKI-SAMS:
As currently there are cod at sea in Scotland, work concentrated on designing, constructing and trialling an integrated onshore system for the production of sea urchins and kelp, using the effluent from on shore cod broodstock. The system was designed to allow a comparison between a system that utilises the effluent stream compared to one that did not. Preliminary results show that on shore cod production generates nutrient rich waters suitable for the production of sea urchins and macroalgae; Nitrogen utilisation within the raceway continues throughout the full 24 hour cycle; the poly-tunnel integrated aquaculture system may be a suitable model for the development of land-based sea urchin and macroalgae production in northern latitudes across Europe.
UNIGE, UNILE have been performing activities in off-shore implants. The first trials have been done in REHO facilities in the first year and at AQUA in the second one. At AQUA, special barrels where used to maintain specimens at 8 m depth hanging at the support ropes of the fish cages. We put in the cages adult specimens collected in the wild. The survival ratio, after 45 days was 95 %. Three feeding treatments have been set up (unfed except from fish wastes, fish pellets, macroalgae). The results were quite promising, particularly if the deployment time of the cage would have been since earlier in the project execution.
IMP-MILU:
experiments were set up both at sea (in Mali Ston Bay) and in an integrated system in-land. Experimental grow out at sea was performed according different feeds (Ulva or grazing biofilm), alongside mussel ropes or in cages, and at different depths. Best results were achieved at 3 m depth. Yet, best results in growth performance and GSI values were achieved in the in-land system, where urchins were fed enriched Ulva from the fish wastes.
WP4 - Gonad improvement (color and GSI) and control gametogenesis
The following experiments have been performed at IOLR:
1) effect of photoperiod manipulation on body growth and gonadal development;
2) diet effects on gonadal growth and gametogenesis;
3) effects of mammalian GnRH agonist on steroidogenesis in cultivated P. lividus populations;
4) effects of sex steroids on gonadal development in cultivated P. lividus populations.
Our study evaluated the effects of environmental conditions (i.e. photoperiod and diet) and those of exogenous hormones (i.e. exposure via water to GnRH agonist or via food to sex steroids), on gonadal development in cultivated P. lividus populations. Additionally, aiming at identifying endogenous factors that signify the onset of gametogenesis, we studied temporal profiles of:
(i) sex steroids;
(ii) transcript levels of dopamine receptors; and
(iii) transcript levels and enzymatic activity of cathepsins, the major proteases involved in yolk processing.
Exposure of adult P. lividus population to 24-h dark [D]or 24-h light [L], indicated no significant differences in terms of body growth or gonadosomatic index (GSI). Nevertheless, the manipulated photoperiod did affect gametogenesis. On one hand, the D regime induced reproductive development, giving rise to multiple reproductive cycles, which are relatively shorter compared to those of wild urchins. On the other hand, the L regime reduced the rates of gametogenesis. The latter effect was more pronounced in males than in females. Yet, on a long-term, acclimation of the urchins to the L regime is observed, emphasising the limitation of the manipulated photoperiod as means to control gametogenesis in P. lividus. Such a method is only effective in short-term applications.
The diet experiment shows that Ulva lactuca (U), which is considered to be a natural food resource for wild P. lividus stocks, significantly increased P. lividus body mass (BW) compared to the formulated dry pellets (P). Nevertheless, the P diet significantly increased the urchins' gonadal growth (GSI) as compared to the U diet. These contrasting effects draw attention to the advantage of using the combination of both diets. Hence, further optimisation of the dietary ratio between fresh enriched alga and formulated dry pellets, is expected to shorten the growth phase and give rise to high quality marketable product.
Our results also reveal sexual-dimorphic patterns, favouring female populations attributable to their higher GSI values. Hence, future research focused on implementing genetic manipulations in sea urchins, should aid in establishing the desired all-female P. lividus populations.
The studies with exogenous hormones indicate that despite the evolutionary distance separating echinoderms from vertebrates, both phyla share some conserved neuroendocrine elements, which are involved in the control of reproduction. Thus, we demonstrated that the vertebrate GnRH ligand can activate P. lividus GnRH receptor(s), and consequently elicit stimulated gonadal sex steroids (i.e. testosterone and estradiol). The obtained stage-dependent profiles further link sex steroid dopamine and cathepsins (mainly B and L) to the onset of gametogenesis. Beside their contribution to the understanding of mechanisms that controls the onset of gametogenesis in P. lividus, our data underscore aspects, which are oriented to aquaculture applicability. One such aspect could be the use of food supplements (approved for human consumption) in order to retard / inhibit steroidogenesis and/or yolk protein degradation by cathepsins. Once optimised, such diets are expected to hold back the onset of gametogenesis, enhance the somatic growth, and widen the period during which high quality roe is being preserved.
WP5 - Environmental quality assessment
Main objectives of WP5 refer to culture Ulva sp. in a biofiltering system using wastewaters and monitoring urchin population and environmental conditions in the ranching areas.
As far as Ulva as a biofilter, activities were fundamentally carried out at IOLR-ARDAG. The performance parameters of Ulva included growth rate, yield, biochemical composition and nutrient uptake. Effluent water from a semi-closed super-intensive fish pond system was drained continuously into Ulva reactors. Nitrogen removal dynamics in a batch flow system for seven days was evaluated. A gradual reduction of TDN from 600 µm to almost 0 within 6 days is observed. Three stages could be identified: rapid reduction of ammonia within 24 h, 48 h of lag with no nitrogen removal, and 36 hours reduction of nitrate. Ulva shows a preference for ammonium compared to nitrates because the nitrogen in the ammonium is in a reduced state that algae can directly utilise for production of amino acids. Nitrates start to be depleted from water after the complete removal of ammonium from the water. This shift takes around 48 hours, the time needed to form active nitrate reductase because active nitrate reductase is not formed in the presence of ammonium. The nitrate then must first be reduced to nitrite, and then ammonium.
Since aeration of the Ulva compartment constitutes about 85 % of the operating cost (energy), we tried to reduce cost by reducing aeration from 100 % to 50 % or 25 % of the time. We evaluated Ulva performance when exposed to these three aeration regimes. A significant increase in Ulva yield is observed between the 50 % and the 100 % aeration regimes. 25 % aeration is the optimal aeration regime for maximal nitrogen removal efficiency (3. 5 g N M-2 d-1), and relatively low energy cost. 100 % aeration regime is less economic but produces maximal yield (178±25 g WW m-2 d-1).
Ulva rearing experiments were performed also at IMP and UNIGE-AQUA. As far as the former, attention was given to the biochemical composition of reared Ulva compared to the wild one, stressing, even in short term experiments, a large enrichment of reared Ulva in terms of organic content compared to ash and moisture. As far as UNIGE-AQUA algae were cultured according to two different treatments, plastic panles and nets, hanging along the support ropes of the AQUA fish cages, at around 1.5 m depth. Nets provided highest macroalgal yield compared to plastic panels, showing biomass values doubled, in spring time, in a month time span.
As far as the natural populations around sea based implants, surveys have been carried out from IMP-MILU and UNIGE-UNILE-AQUA (in the second year, while UNIGE-UNILE-REHO in the first one). Mali Ston Bay is a shallow bay characterised by numerous underwater freshwater springs, which make it very suitable for mariculture. The whole bay is very populated with Paracentrotus lividus, with densities locally exceeding 15 ind/m2. Most abundant size class is 3 - 4.5 cm, average test diameter is 3.7 ± 0.6 cm. According to such population parameters, Mali Ston populations exhibit large size, although a slower growth rate, compared to other Mediterranean areas. GSI varies through the year, with one value peak during April, when spawning occurs, confirmed by histological surveys.
Results from REHO site have been reported in the first reporting report. As far as AQUA, four sampling sites have been chosen, two south of the implants and two north of it. At each site urchins have been visually sampled at the start of experimental culturing and at the end (around two months), in terms of density and size structure, while specimens have been collected at each site on a fortnight base for gonad condition analyses, carried out in terms of GSI and histology (UNIGE) and lipid profile (UNILE) and compared to cultured specimens. Environmental features at the AQUA implant have been regularly monitored.
Potential impact:
P. lividus roe (the red gonads) are very prized, but at the present time in Europe there is an important exploitation in France and Belgium, a moderate one in Italy and Spain and a non-significant, but fast growing one in the rest of Mediterranean Europe, where fishery assumes a seasonal, tourist role. However, the world demand for urchin roe remains very high and searching for new markets. Such a great exploitation has made necessary protection by policy organs:
Already listed within species whose exploitation is regulated in specially protected areas (MAP Phase II, Monaco 24/11/1996), Paracentrotus lividus has now been included among the protected fauna species (Council of Europe, Strasbourg, 3/1998, Appendix III).
In recent years, a new aquaculture activity has developed directed at highly appreciated species for which the current level of production by commercial fisheries is not enough to meet consumers' demand and where the wild populations are facing serious fishing pressure.
ENRICH will provide an essential knowledge base for enrichment of aquaculture farms by the introduction of the European sea urchin, Paracentrotus lividus. In addition the project will develop and test pilot experiments in order to verify the possible commercial exploitation of this valuable species.
The project aimed at substantiating the current knowledge on the reproduction of capture based aquaculture species in captivity, through objectives of WP1 and WP2, establishing knowledge-base required for controlled development of eggs and larvae, and for the development of suitable and environmentally performing feeds. ENRICH fulfils this demand, as the major innovation of this proposal is be improvement of eggs and consequently of larval development, through the controlled quality of food to be supplied in order to optimise broodstock.
The development of a self-sustained aquaculture activity, independent from the supply of wild fish, is thus required both from an industrial and conservation perspective. Thus, the relevance of the introduction of this species into aquaculture farms, either land-based or sea-based, as envisaged in WP3, is at present very high.
WP4 has provided fundamental tools for controlling gametogenesis both for optimising the gamete production of the broodstock and for optimising the gonads of cultured individuals for satisfying the market requirements.
In addition, through WP5, attention has been given to produce macroalgae in integrated systems, both for 'cleaning' effluent waters form fish farming and for feeding urchins with a protein enriched diet. On the other side, WP5 has gathered information on wild stocks, comparing their gonad conditions to cultured specimens and paid attention on possible impacts on the environment in the areas surrounding the sea-based aquaculture systems, contributing directly to ensure the sustainability of capture based aquaculture.
The involved partnership is important to cover the extent of P. lividus distribution in the Mediterranean and the north Atlantic, encompassing a large range of environmental conditions, thus testing feeding and culturing systems in different environmental conditions.
All of the partners bring to the consortium excellence in the field of aquaculture. ENRICH will consequently create competitiveness in the aquaculture of sea urchins.
The SME participants will profit of this work for improving and amplifying the field of work, and explore new markets. Two of them have started the insertion of sea urchins in the aquaculture ranching (AQUA and MILU), and two of them already have experience and their aim is to maximise results in terms of production and quality (VIKI and ARDAG). In Europe, relevant markets are represented by France, Belgium, Holland and also Southern Italy, Greece, Spain.
Local and exportation products will be represented by living samples, but also canned or frozen roe, prepared recipes to be used for particular appetisers and first courses, by cooperation with other differentiated enterprises. Some of the SMEs , in particular VIKI, are interested to produce seeds (sea urchins up to 1 cm diameter), to be delivered to European aquaculturers (Spain, France, Italy, Belgium, Greece) for further growth, and to protection agencies for recovery of depleted areas. All the above will enlarge the field and the occupation of operators.
The SMEs will also profit of the new knowledge to sell it to other enterprises, via the production of CDs covered by copyright (mainly for broodstock improvement), thus increasing European competitiveness, as at present the major exportation to eastern Countries is from California and Chile.
In prospective, following the project completion, touristic exploitation is also planned, as tourists visit to the aquaculture sites, followed by product degustation will be inserted among the touristic attraction of the respective regions.
The project will also have indirect positive effects in terms of environmental protection, as the possibility of successfully rear larvae and juveniles on a large scale will provide a tool for possible restocking of overexploited populations throughout the P. lividus distribution range.
Dissemination activities have been performed at different levels:
1) publication on scientific journals and on journals particularly addressed to the aquaculture world;
2) participation to a wide range of conferences (particularly to aquaculture conferences, in Europe and in other extra-european countries) through oral and poster presentations;
3) participation to specific workshops on P. lividus resource management;
4) organisation of a final workshop of the project in the context of a large scale event, such as the SLOWFISH event in Genova (29 May 2011), gathering scientists from Europe and USA and showing to the wide public the results of the project through digital objects;
5) setting up of a web site and a flyer showing main targets of the project;
6) organising a number of dissemination activities for schools, visiting the implants;
7) compiling a large number of bachelor and master theses and participating to the fulfillment of two PhD thesis;
8) a complete list of the dissemination activities is provided below in the list of dissemination activities.
Exploitation of the results of the project may been fundamentally summarised as follows:
1) establishment of broodstocks for the different SMEs involved in the projects;
2) production of different protocols for optimising the different phases of sea urchin culturing, from the management of the broodstock, to the larval and juvenile phases and a summary of results achieved for the grow out phase, both inland and at sea, in sheltered and exposed areas, also with the support of the results from the gametogenic control.
The SMEs partners of the project will make use of such protocols in order to enhance their production.
Project website: http://www.enrichnet.eu