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WOS: 000416855400001, Science Citation Index Expanded, The aim of this research was to gather preliminary data about the potential inhibitory effects of commercial diets on the protease activities of meagre larvae using in vitro techniques. Four commercial diets (Gemma Micro 150 (100-200 mu m), Caviar (200-300 mu m), Caviar (300-500 mu m) and Perla Larva Proactive 4.0 (300-500 mu m)) were tested in this study. The differences in the protease activities of meagre larvae during the sampling period were statistically significant (p, Unit of Scientific Research Projects, Suleyman Demirel University [SDUBAP 3453-D2-13], This study was supported by the Unit of Scientific Research Projects, Suleyman Demirel University (Gurkan Diken's PhD thesis, Suleyman Demirel University; project no. SDUBAP 3453-D2-13).

Larvae and early juveniles of fast-growing fish species show tremendous growth potential, leading to higher requirements for protein, high-unsaturated fatty acids, and other nutrients. Several commercial weaning diets with relatively high success have been developed for low/moderate-growing species; however, additional challenges are outlined to meet growth potential and energy requirements of such fast-growing species. The objective of the present study was to evaluate two commercial microdiets for marine fish, one having simultaneously a very high protein and high lipid contents (HIGH), and another (MOD) with a high protein and moderate lipid content, in a growth performance trial with greater amberjack post-larvae from 33 to 78 days after hatching. Moreover, histological preparations of liver, anterior and posterior intestine were assessed for hepatic and intestinal lipid inclusions quantification and gut epithelial brush height measurement. Activities of the digestive enzymes: pepsin, trypsin, chymotrypsin, lipase and amylase were also analyzed. Post-larvae fed HIGH microdiet exhibited higher final weight and lower feed conversion ratio that those fed on MOD microdiet. Liver displayed a higher level of lipid inclusions for the MOD diet than for HIGH diet. Moreover, enterocytes of posterior intestine presented a much higher level of supranuclear vacuoles for the HIGH diet compared to MOD diet. The lower Trypsin/Chymotrypsin ratio observed at the end of the experiment in larvae fed on MOD diet may indicate a deficiency in protein of this diet. Together, these results support that larvae of greater amberjack, and likely other fast-growing marine fish species, require high protein–high lipid microdiets. The use of microdiets developed targeting slower growing marine species may lead to sub-optimal performances in fast-growing larvae., This study was undertaken by Project WEANTUNA supported by Eurostars (E! 7994) and co-financed by FEDER through PO Algarve 21 in the framework of QREN 2007-2013 (30200).

In the present work, recent studies that contributed for improving dietary protein in practical microdiets for Senegalese sole larvae are revised, in an attempt to overcome some of the current problems in this species larviculture, such as the difficult early adaptation to inert diets and highly variable growth rates. Different diet formulation strategies were employed in an attempt to improve Senegalese sole larvae capacity to utilize and deposit protein throughout metamorphosis, to further maximize growth potential: (1) increasing indispensable amino acids (IAA) content (2) meeting the ideal IAA profile by adjusting the dietary AA profile to the larval body AA profile and (3) decreasing the complexity (molecular weight) of dietary protein to increase its digestibility. Either manipulating the quality or the complexity of dietary protein had impact on the larvae capacity to utilize protein and direct it for growth. Sole larvae seem to be able to adapt their digestive functions and metabolic capacity to dietary protein. Moreover, increasing the dietary IAA/DAA ratio and changing the degree of hydrolysis of dietary protein affected the expression pattern of muscle growth related genes, with consequences on muscle cellularity and potential for growth. The expression of DNA methyltransferases was altered in response to changes in dietary protein. The novelty of such information in fish may trigger further studies on the effect of dietary protein on the epigenetic regulation of growth. In conclusion, optimal protein quality for Senegalese sole seems to change during larval development. Whereas the inclusion of a moderately hydrolysed protein comes up as a promising way to improve growth in early larval stages, larger peptides and intact protein seem to be more suitable to sole post-larvae and young juveniles. Therefore, these results suggest that dietary protein fraction formulation of microdiets for Senegalese sole should be adapted to each developmental stage, what has important consequences for practical larval microdiets formulation and feeding protocols. POAlgarve 21 [13380] QREN [13380] European UnionEuropean Union (EU) [13380] FCT grant [SFRH/BD/82149/2011] FCT investigator grant - European Social Fund, the Operational Programme Human Potential [IF/00482/2014/CP1217/CT0005] FCT investigator grant - Foundation for Science and Technology of Portugal (FCT) [IF/00482/2014/CP1217/CT0005]

7 Figuras.-- 3 Tablas, Given its complex metamorphosis and digestive system ontogeny, Senegalese sole larvae capacity to digest and utilize dietary protein is likely to change throughout development. In the present study, we hypothesized that the manipulation of dietary protein complexity may affect Senegalese sole larvae capacity to digest, absorb and retain protein during metamorphosis, as well as the mRNA expression of genes encoding for the precursors of proteolytic enzymes of the digestive tract and the enterocyte peptide transporter PepT1, which may have further impact on somatic growth. Three diets were formulated using approximately the same practical ingredients, except for the main protein source. The Intact diet protein content was mostly based on intact plant protein where the target peptide molecular weight (MW) would be > 70 kDa. The PartH diet protein fraction was mostly based on a protein hydrolysate with a high content of 5–70 kDa peptides. The HighH diet protein fraction was mostly based on a protein hydrolysate with a high content of 5 kDa peptides. A growth trial was performed with larvae reared at 19 °C under a co-feeding regime from mouth opening. The transcription of pga, tryp1c, ialp, ampn and pepT1 (encoding respectively for PepsinogenA, Trypsinogen1C, Intestinal alkaline phosphatase, Aminopeptidase N and for the enterocyte peptide transporter 1) was quantified by qPCR, during the metamorphosis climax (16 DAH) and after the metamorphosis was completed (28 DAH). An in vivo method of controlled tube-feeding was used to assess the effect on the larvae capacity to utilize polypeptides with different MW (1.0 and 7.2 kDa) representing a typical peptide MW of each of the hydrolysates included in the diets. The PartH diet stimulated growth in metamorphosing larvae (16 DAH), whereas the Intact diet stimulated growth after 36 DAH. The Intact diet stimulated the larvae absorption capacity for 1.0 kDa peptides at 16 DAH, which may have contributed for enhanced growth in later stages. The PartH diet stimulated the transcription of tryp1c and pept1 at 28 DAH, which seemed to reflect on increased post-larvae capacity to retain dietary 7.2 kDa polypeptides. That may indicate a possible strategy to optimize the digestion and utilisation of the PartH dietary protein, though it did not reflect into increased growth. The Intact diet promoted the transcription of pepsinogenA, which may reflect a reduced gastrointestinal transit time, which could have enhanced the dietary nutrients assimilation, ultimately improving growth. The present results suggest that, whereas pre-metamorphic sole larvae utilize better dietary protein with a moderate degree of hydrolysis, post-metamorphic sole make a greater use of intact protein., This work was funded by Project EPISOLE (FCT) [PTDC/MAR/110547/2009], through project CCMAR/Multi/04326/2013 (Portugal) from FCT (Portugal), and by project MICALA — I&DT Co-Promoção I&DT Co-Promoção No. 13380 (Portugal, supported by POAlgarve 21, QREN and European Union). P. Canada was supported by FCT grant SFRH/BD/82149/2011. Sofia Engrola was supported by FCT investigator grant IF/00482/2014/CP1217/CT0005 funded by the European Social Fund, the Operational Programme Human Potential and the Foundation for Science and Technology of Portugal (FCT). The authors acknowledge the collaboration of Wilson Pinto and André Santos (SPAROS Lda.), the Aquagroup team, (CCMAR) and Hernâni Gerós for revising the manuscript.

Individual differences in physiological and behavioural responses to stressors are increasingly recognised as adaptive variation and thus raw material for evolution and fish farming improvements including selective breeding. Such individual variation has been evolutionarily conserved and is present in all vertebrate taxa including fish. In farmed animals, the interest in consistent trait associations, that is coping styles, has increased dramatically over the last years because many studies have demonstrated links to performance traits, health and disease susceptibility and welfare. This study will review (i) the main behavioural, neuroendocrine, cognitive and emotional differences between reactive and proactive coping styles in farmed fish; (ii) the methodological approaches used to identify coping styles in farmed fish, including individual (group) mass-screening tests; and (iii) how knowledge on coping styles may contribute to improved sustainability of the aquaculture industry, including welfare and performance of farmed fish. Moreover, we will suggest areas for future research, where genetic basis (heritability/epigene tic) of coping styles, and the neuroendocrine mechanisms behind consistent as well as flexible behavioural patterns are pinpointed as central themes. In addition, the ontogeny of coping styles and the influence of age, social context and environmental change in coping styles will also be discussed. European Commission under the 7th Framework Programme FP7-KBBE [265957]

Fish hatcheries must adapt larval feeding protocols to feeding behavior and metabolism patterns to obtain more efficient feed utilization. Fish larvae exhibit daily ingesting rhythms rather than ingesting food continuously throughout the day. The aim of this study was to determine the daily patterns of feed intake, protein digestibility, protein retention and catabolism in Senegalese sole post-larvae (Solea senegalensis; 33 days post-hatching) using 14C-labeled Artemia protein and incubation in metabolic chambers. Sole post-larvae were fed at 09:00, 15:00, 21:00, 03:00 and 09:00+1 day; and those fed at 09:00, 21:00, 03:00 and 09:00+1 day showed significantly higher feed intake than post-larvae fed at 15:00 h (P=0.000). Digestibility and evacuation rate of ingested protein did not change during the whole cycle (P=0.114); however, post-larvae fed at 21:00 and 03:00 h showed the significantly highest protein retention efficiency and lowest catabolism (P=0.002). Therefore, results confirm the existence of daily rhythmicity in feeding activity and in the utilization of the ingested nutrients in Senegalese sole post-larvae., This work received national funds from Fundação para a Ciência e a Tecnologia (FCT; Portugal) through projects CCMAR/Multi/04326/2013 and SOLEAWIN (310305/FEP/71). It was partially supported by the PROMAR Program with funds from Fundo Regional para a Ciência e Tecnologia (FEDER), and also through the Ministerio de Economía y Competitividad (MINECO; Spain) by project EFISHDIGEST (AGL2014-52888-R) with FEDER/European Region Development Fund (ERDF) contribution granted to M.Y. (Spain). Thanks are also due to European Cooperation in Science and Technology (Cost Action) FA0801 (Larvanet). C.N.-G. was supported by a doctoral fellowship (BES-2012-051956) from Ministerio de Economía y Competitividad (Spain). S.E. acknowledges a FCT investigator grant IF/00482/2014/CP1217/CT0005 funded by Fundação para a Ciência e a Tecnologia (FCT; Portugal) and the European Social Fund under the Operational Programme for the Enhancement of Human Potential.

International audience; To sustain eel aquaculture, development of reproduction in captivity is vital. The aim of this review is to assess our current knowledge on the nutrition of broodstock eels in order to improve the quality of broodstock under farming conditions, drawing information from wild adult eels and other marine pelagic spawners. Freshwater eels spawn marine pelagic eggs with an oil droplet (type II), and with a large perivitelline space. Compared with other marine fish eggs, eel eggs are at the extreme end of the spectrum in terms of egg composition, even within this type II group. Eel eggs contain a large amount of total lipids, and a shortage of neutral lipids has been implied a cause for reduced survival of larvae. Eel eggs have higher ARA but lower EPA and DHA levels than in other fish. Too high levels of ARA negatively affected reproduction in the Japanese eel, although high levels of 18:2n-6 in the eggs of farmed eels were not detrimental. The total free amino acid amount and profile of eel eggs appears much different from other marine pelagic spawners. Nutritional intervention to influence egg composition seems feasible, but responsiveness of farmed eels to induced maturation might also require environmental manipulation. The challenge remains to succeed in raising European eel broodstock with formulated feeds and to enable the procurement of viable eggs and larvae, once adequate protocols for induced maturation have been developed

The effects of dissolved oxygen concentration combined with sublethal doses of un-ionised ammonia nitrogen (UIA-N) on growth and physiology were investigated in juvenile turbot. Fish with a mean ± SD initial weight of 19.1 ± 4.1 g, were reared in replicate at normoxic (80–85% O 2 saturation) and hyperoxic (110–115% O 2 saturation) conditions, combined with either control or two sublethal concentrations of un-ionised ammonia (0, 0.17 and 0.31 mg UIA-N l − 1 ) added for 64 days at 18 °C and pH 8.04 in a flow-through system. The subsequent effect of fixed O 2 and UIA concentrations on growth and blood physiology was measured every 22 days. The results show that UIA-N concentrations only had a negative effect on growth under normoxic conditions, suggesting that hyperoxic conditions may increase tolerance to un-ionised ammonia in juvenile turbot. Only minor effects on the acid-base balance were found. A generally higher blood CO 2 and P CO 2 , along with an accumulation of HCO 3 − resulting in an increased blood pH was found in the hyperoxic groups towards the end of the experiment. We conclude that, under normoxic conditions, a chronic exposure of ammonia above 0.17 mg UIA-N l − 1 will have a negative effect on growth and that hyper-oxygenation can be a possible solution to counteract this harmful effect.

Despite considerable progress in recent years, many questions regarding fish larval nutrition remain largely unanswered, and several research avenues remain open. A holistic understanding of the supply line of nutrients is important for developing diets for use in larval culture and for the adaptation of rearing conditions that meet the larval requirements for the optimal presentation of food organisms and/or microdiets. The aim of the present review is to revise the state of the art and to pinpoint the gaps in knowledge regarding larval nutritional requirements, the nutritional value of live feeds and challenges and opportunities in the development of formulated larval diets. © 2013 Wiley Publishing Asia Pty Ltd., This study benefited from participation in LARVANET – COST action FA0801 (EU RTD framework programme. K. Hamre received funding from the Norwegian Ministry of Fisheries and the Research Council of Norway (CODE-199482). M. Yúfera received financial support from the Spanish Ministry of Science and Innovation MICINN + FEDER/ERDF (projects AGL2007-64450-C02-01 and Consolider Ingenio 2010 Program-project Aquagenomics CSD2007-0002). L. Conceição participated in this review in the framework of project HYDRAA – PTDC/MAR/71685/2006, granted by Fundação para a Ciência e a Tecnologia (FCT), Portugal, with the support of FEDER. I. Rønnestad received funding from the Research Council of Norway (CODE- 199482 and GutFeeling- 190019) and the EC FP7 (LIFECYCLE- 222719).

The presence of skeletal anomalies in farmed teleost fish is currently a major problem in aquaculture, entailing economical, biological and ethical issues. The common occurrence of skeletal abnormalities in farmed fish and the absence of effective solutions for avoiding their onset or definitely culling out the affected individuals as early as possible from the productive cycle, highlight the need to improve our knowledge on the basic processes regulating fish skeletogenesis and skeletal tissues differentiation, modelling and remodelling. Severe skeletal anomalies may actually occur throughout the entire life cycle of fish, but their development often begins with slight aberrations of the internal elements. Comprehensive investigation efforts conducted on reared larvae and juveniles could provide a great contribution in filling the gap in knowledge, as skeletogenesis and skeletal tissue differentiation occur during these early life stages. The aim of this review is to provide a synthetic but comprehensive picture of the actual knowledge on the ontogeny, typologies and occurrence of skeletal anomalies, and on the proposed causative factors for their onset in larvae and juveniles of European farmed fish. The state-of-art of knowledge of these issues is analysed critically intending to individualize the main gaps of knowledge that require to be filled, in order to optimize the morphological quality of farmed juveniles. LARVANET COST action [FA0801]; Italian Ministry for Agriculture, Food and Forestry Policy [5C49, 6C82, 6C100, 6C138, 6C144]; Commission of the European Communities [SSP-2005-44483]; NSRF of the Ministry of Education, Lifelong Learning and Religious Affairs, Greece [09SYN-24-619]; Portuguese Foundation for Science and Technology - FCT [PDTC-MAR-105152-2008]; integrated action CRUP-MICINN [AI-E-54]; European Community [222719-LIFECYCLE]; Spanish Ministry of Science and Innovation (MICIIN) [AGL2008-03897-C04-01, AIB2010-PT-00313] info:eu-repo/semantics/publishedVersion

Dietary fatty acid supply can affect stress response in fish during early development. Although knowledge on the mechanisms involved in fatty acid regulation of stress tolerance is scarce, it has often been hypothesised that eicosanoid profiles can influence cortisol production. Genomic cortisol actions are mediated by cytosolic receptors which may respond to cellular fatty acid signalling. An experiment was designed to test the effects of feeding gilthead sea-bream larvae with four microdiets, containing graded arachidonic acid (ARA) levels (0·4, 0·8, 1·5 and 3·0 %), on the expression of genes involved in stress response (steroidogenic acute regulatory protein, glucocorticoid receptor and phosphoenolpyruvate carboxykinase), lipid and, particularly, eicosanoid metabolism (hormone-sensitive lipase, PPARα, phospholipase A2, cyclo-oxygenase-2 and 5-lipoxygenase), as determined by real-time quantitative PCR. Fish fatty acid phenotypes reflected dietary fatty acid profiles. Growth performance, survival after acute stress and similar whole-body basal cortisol levels suggested that sea-bream larvae could tolerate a wide range of dietary ARA levels. Transcription of all genes analysed was significantly reduced at dietary ARA levels above 0·4 %. Nonetheless, despite practical suppression of phospholipase A2 transcription, higher leukotriene B4 levels were detected in larvae fed 3·0 % ARA, whereas a similar trend was observed regarding PGE2 production. The present study demonstrates that adaptation to a wide range of dietary ARA levels in gilthead sea-bream larvae involves the modulation of the expression of genes related to eicosanoid synthesis, lipid metabolism and stress response. The roles of ARA, other polyunsaturates and eicosanoids as signals in this process are discussed., This study was supported by ‘Fundação para a Ciência e a Tecnologia’, (FCT, Portugal; EFARFish – ‘A new method for the study of essential fatty acid requirements in fish larvae’ PTDC/MAR/67017/2006); ‘Consolider – Ingenio 2010’ programme (Plan Nacional I+D+I+FEDER, Spain; ‘Aquagenomics – Improvement of aquaculture production by the use of biotechnological tools’ CSD2007-00002); MCYT+FEDER (Plan Nacional I+D+I, Spain; ‘Mejora del aprovechamiento del alimento en primeras edades de peces marinos: funcionalidad del tubo digestivo y valoración de la utilización de macronutrientes’ AGL2007-64450-C02-01); POCTEP Programme (0251-ECOAQUA-5-E). D. A. M. was supported by grant SFRH/BPD/32469/2006 (FCT, Portugal). The study also benefited from the LARVANET grants COST-STSM-FA0801-4496 and COST-STSM-FA0801-5429.

A number of scientific studies have investigated aspects of soles ( Solea solea and S. senegalensis) ecology, population genetics and biology in their natural environment, and the species have been extensively studied in captivity during the last decade. Studies on the genetic population structure of sole indicate that several distinct breeding populations exist within its distributional range in European waters. Recent studies suggest a phylogenetic relatedness of S. solea and S. senegalensis, being found as closest sister lineages in most reconstructions. However, studies on molecular genetics and morphological traits give diagnostic differences that consistently lead to their taxonomic separation at the specific rank. Studies show that sole spawn readily in captivity, and the buoyant, fertilized eggs are easily collected. Stocking density during maturation should be 1-1.5 kg/m(2), and temperature should be kept above 16degreesC ( S. senegalensis) or between 8 and 12degreesC ( S. solea). In nature, the onset of spawning is related to a rise in temperature occurring during spring ( March-June). Salinity should be kept constant around 33-35& and the fish reared under simulated natural photoperiod ( LDN). In other cultured flatfish species, a change in the photoperiod is the key environmental signal used to manipulate and control maturation, but at present time there are no published work that verifies or contradicts this for either S. senegalensis or S. solea. Studies indicate that a mixture of inert and live food may increase the weaning success of sole fry, and this can be further enhanced by using attractants in the dry feed. Future experiments are needed to determine the ideal time to commence weaning and determine the minimum duration of this period. Studies on alternative feeding strategies are also required. The effect of temperature and photoperiod on juvenile growth has not been studied systematically in neither of the two species and the relative importance of a direct photoperiod effect on growth in sole therefore remains to be defined.