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The behavioural effects of confinement of sardine Sardina pilchardus in a purse seine were evaluated through three laboratory experiments simulating the final stages of purse seining; the process of slipping (deliberately allowing fishes to escape) and subsequent exposure to potential predators. Effects of holding time (the time S. pilchardus were held or entangled in the simulation apparatus) and S. pilchardus density were investigated. Experiment 1 compared the effect of a mild fishing stressor (20min in the net and low S. pilchardus density) with a control (fishing not simulated) while the second and third experiments compared the mild stressor with a severe stressor (40min in the net and high S. pilchardus density). In all cases, sea bass Dicentrarchus labrax were used as potential predators. Results indicated a significant effect of crowding time and density on the survival and behaviour of slipped S. pilchardus. After simulated fishing, S. pilchardus showed significant behavioural changes including lower swimming speed, closer approaches to predators and higher nearest-neighbour distances (wider school area) than controls, regardless of stressor severity. These results suggest that, in addition to the delayed and unobserved mortality caused by factors related to fishing operations, slipped pelagic fishes can suffer behavioural impairments that may increase vulnerability to predation. Possible sub-lethal effects of behavioural impairment on fitness are discussed, with suggestions on how stock assessment might be modified to account for both unobserved mortality and sub-lethal effects, and possible approaches to provide better estimates of unobserved mortality in the field are provided. (C) 2013 The Fisheries Society of the British Isles

The larval development of the dusky grouper Epinephelus marginatus up to the benthic juvenile stage is described in detail to establish a reference for their larval identification. Development is described in terms of ontogenetic changes in morphology, growth, pigmentation, fin structure and skeletal structure. Larvae were reared in mesocosms at a mean temperature of 24·3◦ C, salinity of 36·5, dissolved oxygen of 6·4mg l−1 and pH of 8·2. Newly hatched larvae had an estimated total length (LT) of 2·3 mm. On the second day post hatching the yolk was almost fully absorbed with traces of the oil globule still present, the eyes were already pigmented and mouth and gut functional. At this stage the cranial skeletal elements for feeding and breathing (mouth and gills) and the pectoral-fin support were already present. About 50% of the observed larvae had food in their guts. Pigmentation was very characteristic, consisting of two large chromatophores visible on the edge of the primordial fin, close to the midpoint of the post-anal region of the body and over the midgut and hindgut and post-anal portion of the body. At 2·9mm LT the emergence of the second dorsal-fin spine, characteristic of the Epinephilinae, was clearly visible. The pre-flexion stage started in larva of 3·2mm LT. At 5·5mm LT the larvae possessed posterior preopercular angle spines, and the dorsal and pelvic spines presented serrated edges and were pigmented. The water surface-tension-related death of the yolk sac and pre-flexion larvae described in the rearing of several other grouper species did not occur during E. marginatus culture. Notochord flexion, with initial ossification of the caudal-fin supporting elements, started at 6·6mm LT. At this stage the major melanophores, preopercular, dorsal and pelvic spines and mandibular teeth were already present. Transformation of larvae into juveniles occurred when larvae averaged 13·8mm LT. Juveniles with a mean LT of 20·1mm started to settle and most of them were benthic with a mean LT of 26·8 mm.

Lordosis and kyphosis are curvatures of the vertebral column that commonly affect Senegalese sole (Solea senegalensis) under intensive rearing conditions, constituting a problem for the aquaculture industry by increasing labor and operation costs and consequently reducing profits. However, no studies have yet addressed microanatomical changes in vertebral column curvature in this species. We have conducted histological analyses in order to determine the alterations at tissue and cellular levels in these deformities. Development of an ectopic cartilage-like tissue within bone matrix at the growth regions (contact area between two vertebrae) of affected vertebrae was observed in lordotic-kyphotic fish. As previously suggested, the increase in strain supported by deformed vertebrae may trigger the onset of metaplastic tissue formation through an osteochondroprogenitor precursor from the condensation of mesenchymal cells or by trans-differentiation of (pre-)osteoblastic cells to a chondrocytic lineage. The possibility of a notochord contribution, as well as the molecular events related to bone stress and bone types in terms of cellularity, are also discussed.

The meagre (Argyrosomus regius) is a new species for aquaculture diversification, presenting a high economic value and a high growth rate, reaching 1 kg year)1. The occurrence of deformities, which generally appear in cultured larvae at early stages, is problematic for aquaculture since they reduce fish fitness and profitability. Knowledge about the stages of skeletal formation is essential to accurately assess deformities in hatchery produced fish. Here, we describe the osteology of the axial skeleton and unpaired fins of the meagre. Based on this data, the osteological development of meagre larvae reared at the IPIMAR and IRTA-SCR, under different rearing conditions, was compared. Although little literature is available on the osteology of teleosts from the same taxonomic group, the skeletal composition of the meagre is comparable to that of other Sciaenidae. All skeletal structures considered appeared at earlier stages in larvae reared at IPIMAR, which presented a more advanced skeletal development in comparison to larvae of comparable length reared at IRTA-SCR that were submitted to lower temperature and light intensity, and reduced photoperiod regimes.

In order to investigate the effects of dietary protein hydrolysates (PH) on larval growth performance, skeleton quality and proteome expression, triplicate groups of white seabream (Diplodus sargus) larvae were co-fed from first-feeding with live feed and three microencapsulated diets differing in the molecular weight of their PH fraction (Control – inclusion of CPSP-90; H – inclusion of a high amount in 0.5–30 kDa hydrolysates; L – inclusion of a high amount in

In order to investigate the effects of dietary protein hydrolysates (PH) on larval growth performance, skeleton quality and proteome expression, triplicate groups of white seabream (Diplodus sargus) larvae were co-fed from first-feeding with live feed and three microencapsulated diets differing in the molecular weight of their PH fraction (Control – inclusion of CPSP-90; H – inclusion of a high amount in 0.5–30 kDa hydrolysates; L – inclusion of a high amount in, This study was partially supported by the projects SAARGO – POCI/MAR/6123/2004 and HYDRAA – PTDC/MAR/71685/2006, granted by Fundação para a Ciência e Tecnologia, Portugal. NR, M de V and TSS acknowledge financial support from Fundação para a Ciência e Tecnologia, Portugal, through grants SFRH/BDP/34888/2007, SFRH/BD/40698/2007 and SFRH/BD/41392/2007 respectively.

Despite the low incidence of morpho-anatomical abnormalities occurring in natural populations, this pose a serious problem for cultivated fish causing increased mortality, reduced growth and conversion rates, thus limiting production and profits. In this study we made an osteological evaluation in the white seabream (Diplodus sargus) and in the common two-banded seabream (Diplodus vulgaris), two sparid species currently under adaptation to intensive aquaculture. Commission of the European Communities - 2007-2013 Atlantic Area Transnational Programme; Project “SEAFARE - Sustainable and environmentally friendly aquaculture for the Atlantic region of Europe”

The Senegalese sole (Solea senegalensis) is characteristic from southern Europe and Mediterranean. It has recently been adapted for aquaculture production since it is well accepted by consumers and reaches high commercial values. After the initial description of the ontogenic events of skeletogenesis, systematic evaluation of the incidence of skeletal malformations by several groups revealed high levels of skeletal deformities reaching up to 90% of the population.

Trabajo presentado en el V Workshop The cultivation of the Soles, celebrado en Faro (Portugal) del 5 al 7 de abril de 2011., Low sperm volume and quality is pointed out as a major reason for the low fertilization rates observed in Senegalese sole (Solea senegalensis) broodstocks both wild and F1. R ecent studies by our group have shown that the poor and variable sperm quality is related to the high sensibility of plasma membrane to seawater during fertilization. Diet enrichment in fatty acids and cholesterol could change the composition of the sperm membrane and the addition of antioxidants would protect sperm to peroxidation.Therefore, in the present work we tested specific diets enriched with DHA, selenium and Vit. E, aiming to improve sperm sole quality., J. Beirão is supported by PhD grant (SFRH/BD/31990/2006) from the Portuguese Ministry of Science and Technology.

Trabajo presentad en el World Aquaculture 2011, celebrado en Natal (Brasil) del 6 al 10 de junio de 2011., Currently, several species of grouper are already being produced in Asia for commercial purposes. Several individuals are kept as broodstock in research facilities throughout the Mediterranean.

Quality control in fish hatcheries is of paramount importance to achieve the desired characteristics of larvae and fry, either for the market or release to the wild. Quality programmes are applied in a day to day basis having direct implications for production management and final costs of product. Most common criteria used for larval quality assessment are morphometrics, condition factor, histometrical indices, lipid analysis, nucleic acid ratios, enzyme activity and stress tests.

Vertebral deformities are frequent and constitute a major drawback for many aquaculture produced species. Lordosis and kyphosis, which are severe curvatures of the vertebral column, are particularly problematic, compromising fish marketing and profitability. Senegalese sole is a species with increasing interest in this industry and, like many others, presents high frequencies of vertebral deformities under rearing conditions. project SEACASE (Nº 044483; FP6-2005-SSP5A) and by project SPECIAL_K (PTDC-MAR-105152-2008)

Trabajo presentado en el 3rd International Workshop on Biology of Fish Gametes, celebrado en Budapest del 7 al 9 de septiembre de 2011., Cryopreservation of testicular germ cells offers a tool for the conservation and production of species with potential for aquaculture, due to the capacity of spermatogonia differentiating into gametes. These cellshave the ability to proliferate in a host individual after transplantation producing, in their gonad, functionalgametes depending on the host sex determination (Yoshizaki et al., 2011). Cryopreservation andtransplantation of testicular germ cells have been attempted in several mammalian species (Redden et al., 2009) and more recently in some fish (Takeuchi et al . 2009; Yoshizaki et al., 2011). In the present studycryopreservation protocols were developed for testicular germ cells from Senegalese sole ( Solea senegalensis ), a species with several reproductive problems, with the aim to transplant into newly hatched larvae and testtheir efficiency., This research was supported by the Spanish Ministry of Science and Innovation through project PGCFISH(AGL2008-02172) and ASSEMBLE call; E. Cabrita was supported by a research contract (RYC-2007-01650); T. Pacchiarini was supported by a JAE-DOC fellow (CSIC).

The meagre (Argyrosomus regius) is a new species for aquaculture with high economical value, presenting growth rates of 1 kg/year. Skeletal deformities generally appear in cultured larvae at early stages, reducing the value of the fish. The present study describes the osteological development of the vertebral column and fins in meagre larvae. Meagre larvae were obtained from wild broodstocks, kept at IRTA and IPIMAR, and induced to spawn using GnRHa injection. AQUACOR 31-03-05-FEP-0003; INIA predoctoral fellowship

The European project Sustainable extensive and semi-intensive coastal aquaculture in Southern Europe (SEACASE) included the proposal of 6 Codes of Conduct for the aquaculture systems studied during the project work. These case studies are typical of the countries involved, namely semi-intensive polyculture (Portugal and Spain), extensive polyculture in esteros (Spain), integrated systems (France), valliculture (Italy), nursery ponds (Greece, Portugal, France and Italy) and eel ponds (France). This article was written having as objective the providing of a concise summary of the characteristics that SEACASE collaborators found to be central to build a complete and trustable Certification system for products of sustainable non-intensive aquaculture (for “certification of sustainability”, see Ojeda, 2008). The 6 Codes of Conduct are the basic information to build such a system in the future., This study has been carried out with the financial support from the Commission of the European Communities, specific RTD programme “Specific Support to Policies”, SSP-2005-44483 “SEACASE - Sustainable extensive and semi-intensive coastal aquaculture in Southern Europe”, and does not necessarily reflect the European Commission views and in no way anticipates the Commission’s future policy in this area.

Earthen ponds are the main production system for seabass and seabream in Portugal and in Southern Spain, in particular in Cádiz province. Different farms use various levels of stocking densities and pond sizes, but in general these are semi-intensive systems covering large areas with ponds ranging from one to several hectares and production levels from 0.5 to 6 Kg/m3 (mostly below 2 kg/m3) at the end of the production cycle., This research has been carried out with the financial support from the Commission of the European Communities, specific RTD programme “Specific Support to Policies”, SSP-2005-44483 “SEACASE - Sustainable extensive and semi-intensive coastal aquaculture in Southern Europe”, and does not necessarily reflect the European Commission views and in no way anticipates the Commission’s future policy in this area.

Earthen ponds are the main production system for seabass and seabream in Portugal and in Southern Spain, in particular in Cádiz province. Different farms use various levels of stocking densities and pond sizes, but in general these are semi-intensive systems covering large areas with ponds ranging from one to several hectares and production levels from 0.5 to 6 Kg/m3 (mostly below 2 kg/m3) at the end of the production cycle. Although seabass and seabream are traditionally the target species produced in such ponds, there is commonly natural recruitment of wild larvae of other fish species, including Senegalese sole. Previous attempts at ongrowing sole in ponds with seabass and seabream as added value gave varied results in terms of number of individuals recovered, but sole growth rates were promising. Polyculture of seabass and seabream is already an established practice, with proportions of 4:1 normally used. In seabass dominated ponds seabream is used to control growth of macroalgae and to clean the ponds. When seabream production dominates, the carnivorous nature of seabass is used to control populations of smaller fish which enter the ponds naturally and might compete for feed. Polyculture of species from different trophic levels has also been considered an efficient and environmentally sound strategy to minimise the impacts of aquaculture systems, since an important fraction of dissolved nutrients and organic matter is recycled within the pond (Buschmann et al. 1996, Sorgeloos 2001, World Bank 2006)., This research has been carried out with the financial support from the Commission of the European Communities, specific RTD programme “Specific Support to Policies”, SSP-2005-44483 “SEACASE - Sustainable extensive and semi-intensive coastal aquaculture in Southern Europe”, and does not necessarily reflect the European Commission views and in no way anticipates the Commission’s future policy in this area.

Senegalese sole is distributed along the Mediterranean and southwestern Europe, reaching a market price of 20-25 €/kg - Promising species for aquaculture diversification - Control and intensification of culture is reasonably achieved - The high incidence of skeletal anomalies in intensively reared fish represents a constraint to commercial production - Samples from Portugal and Greece were analyzed towards identifying skeletal differences due to rearing conditions

The meagre (Argyrosomus regius) is a marine teleost with high growth rates and excellent biological characteristics for rearing, making it a proper candidate for diversification of aquaculture in Southern Europe. The description of skeletal development is essential to establish chronometric relationships between skeletal developments, types of ossification, processes and different development stages, contributing to the improvement of rearing conditions. POTC-MAR-10515 2-2008; PTDC/MARl0S313(2008)

One of the major obstacles to propagating dusky grouper, Epinephelus marginatus, is the difficulty of rearing the early larvae. We have successfully raised dusky grouper larvae in mesocosms using a mixed diet of endogenous plankton grown in the rearing tank and an exogenous supply of Brachionus plicatilis and Artemia sp. Newly hatched larvae at an initial density of 1.3 ind.l-1 were stocked in partially shaded 3 m3 circular outdoor tanks during the summers of 2007 and 2008. Before introducing newly hatched larvae, the water was left for six days to promote plankton growth. Larval growth occurred at two different rates: i) a faster rate from first feeding to the beginning of metamorphosis, and ii) a slower rate at transformation. Survival at the beginning of metamorphosis was less than 10% (33 DPH) in 2007 and between 25 and 50% (25 DPH) in 2008. High mortalities were observed during larval transformation. The estimated minimum food requirement per grouper larvae increased more than 300% from the beginning of the notochord flexion to the beginning of metamorphosis. To meet such a high feeding requirement the number of larger prey organisms/copepods in the mesocosm should be eight times higher during this time period.

The appearance of skeletal deformities is a major problem associated to intensive, large-scale culture of fish species, affecting growth, development and ultimately, survival of produced fish. Although the mechanisms working behind it are still unclear, it is known that environmental, genetic and nutritional factors play fundamental roles. Given its importance as a vertebrate model to study skeleton-related diseases, the selection of an appropriate dietary protocol is a critical step in order to diminish the nutritional effect on bone and cartilage development. To better understand the effect of diet on skeletal development and deformities, as well as on larvae and juvenile global performance, we have tested four distinct dietary regimes, composed of rotifers Brachionus sp., Artemia nauplii and dry, formulated food.

Species diversification is presently a major concern amongst aquaculture researchers and producers trying to find new profitable species for commercial production and restocking. White seabream Diplodus sargus (Linnaeus, 1758) and common two-banded seabream Diplodus vulgaris (Saint-Hillaire, 1817) are two new promising sparid species for aquaculture production, as an alternative to gilthead seabream Sparus aurata and sea bass Dicenthrarchus labrax, since they present different spawning seasons and good growth ratios.

A 65-day experiment was conducted to determine survival, growth, feed utilization and oogenesis of Nereis diversicolor (O. F. Müller, 1776), fed with a commercial dry diet developed for gilthead seabream Sparus auratus L., 1758. Worms were reared in three replicate tanks with 50 worms per tank (255 indiv · m_2). Tetramin®, a commonly used diet for ornamental fish and polychaete laboratory experiments, was used as a control. Survival was high (95.3 %) for both treatments, and not significantly affected by diet. Specific growth rates and feed efficiency ratios did not differ significantly between worms fed with either diets, nor were there significant differences in the proportion of individuals in the oocyte size classes., La supervivencia, el crecimiento, la utilización del alimento y la oogénesis de Nereis diversicolor (O. F. Müller, 1776) alimentado con pienso de dorada Sparus auratus L., 1758 fueron estudiados durante 65 días. Los poliquetos se cultivaron en tres tanques replicados con 50 individuos en cada uno (255 indiv · m_2). Como control se utilizó Tetramin®, una dieta común para peces ornamentales y en experimentos de laboratorio con poliquetos. La supervivencia fue alta (95,3 %) en ambos tratamientos y no se vio afectada significativamente por la dieta. El aumento de peso y los cocientes de eficiencia alimentaria no variaron significativamente entre los poliquetos alimentados con ambas dietas. Entre los grupos experimentales no hubo diferencia significativa en la proporción de individuos por clases de tamaño de los oocitos., Instituto Español de Oceanografía

The growth and survival of the polychaete Nereis diversicolor (O. F. Müller, 1776) fed on faeces of the carpet shell clam Ruditapes decussatus (L., 1758) were studied under controlled temperature and salinity conditions. Juvenile worms were fed with clam R. decussatus faeces in the presence or absence of sediment during 65 days. Starvation treatments were tested as a control. The results of the present study show that the worms were able to grow using the clam faeces in the presence or absence of sediment. However, the worms from the trials with sediment showed a higher survival rate and biomass production than those from the trials in which no sediment was used. These findings suggest that a suspension of bivalve faeces can be used in the rearing of juvenile N. diversicolor in an integrated polyculture system., El crecimiento y la supervivencia del poliqueto Nereis diversicolor (O. F. Müller, 1776) alimentado con heces de almeja Ruditapes decussatus (L., 1758) fueron estimados bajo condiciones controladas de temperatura y salinidad. Los poliquetos juveniles fueron alimentados con heces en presencia y ausencia de sedimento durante 65 días. Como control se aplicaron tratamientos de ayuno. Los resultados sugieren que los poliquetos crecieron usando las heces como única dieta en presencia y en ausencia de sedimento. Es más: los poliquetos de los ensayos con sedimiento mostraron supervivencia y producción de biomasa mayores que aquéllos en que no se utilizó sedimiento. Los resultados sugieren que se puede usar una suspensión de heces procedentes de bivalvos en el cultivo de juveniles de N. diversicolor en un sistema de policultivo integrado., Instituto Español de Oceanografía

The growth and survival of the polychaete Nereis diversicolor (O. F. Müller, 1776) fed on faeces of the carpet shell clam Ruditapes decussatus (L., 1758) were studied under controlled temperature and salinity conditions. Juvenile worms were fed with clam R. decussatus faeces in the presence or absence of sediment during 65 days. Starvation treatments were tested as a control. The results of the present study show that the worms were able to grow using the clam faeces in the presence or absence of sediment. However, the worms from the trials with sediment showed a higher survival rate and biomass production than those from the trials in which no sediment was used. These findings suggest that a suspension of bivalve faeces can be used in the rearing of juvenile N. diversicolor in an integrated polyculture system. El crecimiento y la supervivencia del poliqueto Nereis diversicolor (O. F. Müller, 1776) alimentado con heces de almeja Ruditapes decussatus (L., 1758) fueron estimados bajo condiciones controladas de temperatura y salinidad. Los poliquetos juveniles fueron alimentados con heces en presencia y ausencia de sedimento durante 65 días. Como control se aplicaron tratamientos de ayuno. Los resultados sugieren que los poliquetos crecieron usando las heces como única dieta en presencia y en ausencia de sedimento. Es más: los poliquetos de los ensayos con sedimiento mostraron supervivencia y producción de biomasa mayores que aquéllos en que no se utilizó sedimiento. Los resultados sugieren que se puede usar una suspensión de heces procedentes de bivalvos en el cultivo de juveniles de N. diversicolor en un sistema de policultivo integrado. Instituto Español de Oceanografía

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