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Methionine and taurine are amino acids (AA) that are usually deficient when fish meal is replaced by plant proteins. In this study, three diets were tested in juvenile meagre (initial weight: 13.4 g) for 8 weeks. The D1 diet had 0.2% methionine and 1% taurine supplementation; the D2 and D3 diets had 0.6% methionine and 1% and 2% taurine supplementation, respectively. The results showed that meagre fed the D1 diet had lower specific growth rate (2.2 to 2.5), lower feed efficiency (0.9 to 1.2) and higher food conversion rate (FCR, 1.1 to 0.8) as well as a lower activity of the alanine aminotransferase (ALAT) enzyme. Furthermore, a higher recruitment of muscle fibres (46% compared to 36%) as well as a higher fibre density was observed (1019 compared to 870 fibres mm(-2)). This study shows that meagre requires a sufficient quantity of methionine in plant-based diets to avoid a reduction in fish performance. Furthermore, taurine supplementation in the D1 diet was not able to mitigate the effects of methionine deficiency. A higher taurine supplementation did not improve meagre performance. MAR02.01.01 FEAMP-0175 info:eu-repo/semantics/publishedVersion

The sustainability of the Aquaculture industry relies on optimising diets to promote nitrogen retention and maximise fish growth. The aim of this study was to assess how different dietary formulations influence the bioavailability and metabolic fate of distinct amino acids in gilthead seabream juveniles. Amino acids (lysine, tryptophan, and methionine) were selected based on their ketogenic and/or glucogenic nature. Seabream were fed practical diets with different protein (44 and 40%) and lipid contents (21 and 18%): 44P21L, 44P18L, 40P21L, and 40P18L. After three weeks of feeding, the fish were tube-fed the correspondent diet labelled with 14C-lysine, 14C-tryptophan, or 14C-methionine. The amino acid utilisation was determined based on the evacuation, retention in gut, liver, and muscle, and the catabolism of the tracer. The metabolic fate of amino acids was mainly determined by their nature. Tryptophan was significantly more evacuated than lysine or methionine, indicating a lower availability for metabolic purposes. Methionine was more retained in muscle, indicating its higher availability. Lysine was mainly catabolised, suggesting that catabolism is preferentially ketogenic, even when this amino acid is deficient in diets. This study underpins the importance of optimising diets considering the amino acids’ bioavailability and metabolic fate to maximise protein retention in fish. info:eu-repo/semantics/publishedVersion

Commercial diets for tilapia juveniles contain high levels of plant protein sources. Soybean meal has been utilised due to its high protein content; however, soy-based diets are limited in methionine (Met) and require its supplementation to fulfil fish requirements. dl-Methinone (dl-Met) and Ca bis-methionine hydroxyl analogue (MHA-Ca) are synthetic Met sources supplemented in aquafeeds, which may differ in biological efficiency due to structural differences. The present study evaluated the effect of both methionine sources on metabolism and growth of Nile tilapia. A growth trial was performed using three isonitrogenous and isoenergetic diets, containing plant ingredients as protein sources: DLM and MHA diets were supplemented on equimolar levels of Met, while REF diet was not supplemented. Hepatic free Met and one-carbon metabolites were determined in fish fed for 57 d. Metabolism of dl-Met and MHA was analysed by an in vivo time-course trial using 14C-labelled tracers. Only dl-Met supplementation significantly increased final body weight and improved feed conversion and protein efficiency ratios compared with the REF diet. Our findings indicate that Met in DLM fed fish follows the transsulphuration pathway, while in fish fed MHA and REF diets it is remethylated. The in vivo trial revealed that 14C-dl-Met is absorbed faster and more retained than 14C-MHA, resulting in a greater availability of free Met in the tissues when fish is fed with DLM diet. Our study indicates that dietary dl-Met supplementation improves growth performance and N retention, and that Met absorption and utilisation are influenced by the dietary source in tilapia juveniles.

The dietary protein to energy ratio (P/E) has proven to influence protein utilization and/or growth in several fish species. This study intended to unravel the bioavailability and metabolic fate of lysine and methionine in gilthead seabream (Sparus aurata) juveniles fed plant diets with different P/E ratios. Seabream juveniles were fed two isonitrogenous diets (45% crude protein) differing in crude lipids (20 and 14%): LowP/E ( P / E ratio = 20.0 mg protein k J − 1 ) and HighP/E ( P / E ratio = 21.4 mg protein k J − 1 ). After three weeks, fish ( 11.6 ± 4.3 g ) were tube-fed the respective diet labelled with 14C-protein (L-amino acid mixture), 14C-lysine, or 14C-methionine. Protein, lysine, and methionine utilization were determined based on the proportion of 14C-amino acid evacuated, retained in the free or protein-bound fraction of liver and muscle, or catabolized. This study revealed that a decrease in P/E ratio resulted in lower amino acid evacuation (p p < 0.05 ) due to a simultaneous higher retention of lysine and methionine, without affecting the overall protein retention. Methionine catabolism was significantly lower than lysine or protein independently of the P/E ratio ( p < 0.05 ), reinforcing that this amino acid is preferentially spared for metabolic functions and not used as energy source. In contrast, increasing the dietary P/E ratio decreased lysine catabolism and increased its availability for growth. The bioavailability and metabolism of individual amino acids should be considered when optimizing P/E ratios in diets for gilthead seabream juveniles. Formulating diets with optimum P/E ratios will improve diet utilization and fish performance.

To support the expected increase in aquaculture production during the next years, a wider range of alternative ingredients to fishmeal is needed, towards contributing to an increase in production sustainability. This study aimed to test diets formulated with non‐conventional feed ingredients on gilthead seabream (Sparus aurata) growth performance, feed utilization, apparent digestibility of nutrients and nutrient outputs to the environment. Four isonitrogenous and isoenergetic diets were formulated: a control diet (CTRL) similar to a commercial feed and three experimental diets containing, as main protein sources, plant by‐products, glutens and concentrates (PLANT); processed animal proteins (PAP); or micro/macroalgae, insect meals and yeast (EMERG). Diets were tested in triplicate during 80 days. The EMERG treatment resulted in lower fish growth performance, higher FCR and lower nutrient and energy retentions than the other treatments. The lowest protein digestibility was found for the EMERG diet, which caused increased nitrogen losses. The PLANT and PAP treatments resulted in better fish growth performance, higher nutrient and energy retentions, and lower FCR than the CTRL treatment. The significant improvement in FCR found for fish fed PLANT and PAP diets and the high protein digestibility of these diets contribute towards minimizing the environmental impacts of seabream production. FCT: UID/Multi/04326/2019/ PTDC/MAR‐BIO/3034/2014/ DL57/2016/CP1361/CT0033. info:eu-repo/semantics/publishedVersion

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Gilthead seabream (Sparus aurata) is vulnerable to low water temperature, which may occur in the Southern Europe and Mediterranean region during Winter. Fish are poikilothermic animals, therefore feed intake, digestion, metabolism and ultimately growth are affected by water temperature. This study aimed to evaluate growth performance, feed utilisation, nutrient apparent digestibility, and nitrogen losses to the environment in gilthead seabream juveniles reared under low temperature (similar to 13 degrees C). Three isolipid and isoenergetic diets were formulated: a diet similar to a commercial feed (COM) that contained 44% crude protein and 27.5% fishmeal, and two experimental diets with a lower protein content of 42% (ECO and ECOSup). In both ECO diets fishmeal inclusion was reduced (10% in ECO and 7.5% in ECOSup diet) and 15% poultry meal was included. Additionally, the ECOSup diet was supplemented with a mix of feed additives intended to promote fish growth performance and feed intake. The ECO diets presented lower production costs than the COM diet, whilst incorporating more sustainable ingredients. Gilthead seabream juveniles (+/- 154.5 g initial body weight) were randomly assigned to triplicate tanks and fed the diets for 84 days. Fish fed the ECOSup diet attained a similar final body weight than fish fed the COM diet, significantly higher than fish fed the ECO diet. ECOSup fed fish presented significantly higher hepatosomatic index than COM fed fish, most likely due to higher hepatic glycogen reserves. The viscerosomatic index of ECOSup fed fish were significantly lower compared to COM fed fish, which is a positive achievement from a consumer's point of view. ECOSup diet exhibited similar nutrient digestibility than the COM diet. Moreover, feeding fish with the ECO diets resulted in lower faecal nitrogen losses when compared to COM fed fish. The results suggest that feeding gilthead seabream with an eco-friendly diet with a mix of feed additives such as the ECOSup diet, promoted growth and minimised nitrogen losses to the environment. Nutritional strategies that ultimately promote feed intake and diet utilisation are valuable tools that may help conditioning fish to sustain growth even under low temperatures. European UnionEuropean Commission [727315]; Portuguese Foundation for Science and Technology (Ministry of Science and Higher Education, Portugal) [DL 57/2016/CP1361/CT0033]

Welfare in aquaculture is a pressing topic. One of the main measures to improve the life of farmed fish is implementing environmental enrichment, which can include the addition of structural covers to rearing tanks. Here we test the effects of tank covers (fully covered, 50 % covered and uncovered) in an all-male population (N = 75 tagged individuals: 5 individuals per tank, 5 tanks per treatment) of farmed Nile tilapia using behavioural and physiological welfare indicators. Video recordings were performed over 21 days to evaluate overall swimming (swim bursts, freezing on the bottom or hovering in the water column) and aggression events (bites and chases). At the end of the experiment, blood was collected to assess circulating cortisol. Results showed a clearly lower welfare in the partially covered tanks regarding behavioural and physiological variables. The present study highlights the need to correctly study 1) the ethology of the farmed species and 2) the enrichment strategy to be implemented, before any measure is taken. While environmental enrichment is undoubtedly a much needed area of research and a promising solution to improve the welfare of farmed fish, a proper ethological assessment must be made a priori to avoid detrimental effects of poorly devised approaches. UID/Multi/04326/2020; IF/00482/2014/CP1217/CT0005; DL 57/2016/CP1361/CT0033 info:eu-repo/semantics/publishedVersion

Aquaculture is one of the most thriving animal production sectors, and Nile tilapia (Oreochromis niloticus) farming represents 8% of total finfish culture. However, the industry sustainability depends on the development of cost‐effective and environmental friendly feeds. This study aimed to reduce dietary protein levels in diets for juvenile tilapia and to minimize diet environmental impact while maximizing biological efficiency. A growth trial was performed using five isoenergetic plant protein‐based diets with decreasing levels of crude protein: 360, 340, 320, 300 and 280 g/kg diet (D360, D340, D320, D300 and D280, respectively). Dietary protein utilization was assessed by metabolic trials using a radiolabelled amino acid mixture. Tilapia in all treatments showed similar growth performance and feed intake. Feed conversion ratio was significantly higher in fish fed the D280 than the D360 diet, while no differences were found for other treatments. Protein retention was significantly higher in tilapia fed the D300 than the D360 diet. Amino acid catabolism increased in fish fed the D360 diet, though without significant differences in muscle amino acid retention. This study demonstrates that dietary protein levels can be reduced to 300 g/kg diet without hindering tilapia growth and feed conversion ratio, while reducing environmental nitrogen losses. FCT: UID/Multi/04326/2019/ IF/00482/2014/CP1217/CT0005/ DL57/2016/CP1361/CT0033. info:eu-repo/semantics/publishedVersion

The regulatory role of taurocholic acid (TC) and taurolithocholic acid (TLC) in ion transport was investigated with the short-circuit current (Isc) technique in the intestine of Senegalese sole (Solea senegalensis). Under voltage clamp conditions, Isc was stable for over 3 h and averaged −15.42 ± 2.75 μA/cm2 and −65.28 + 7.69 μA/cm2 in the anterior and posterior intestine, respectively. Only bumetanide (200 μM), a loop diuretic that inhibits Na+/K+/2Cl− co-transporters, inhibited the absorptive Isc, and apical application of NPPB (200 μM), EIPA (200 μM), glibenclamide (200 μM) and DIDS (200 μM) were without effect in the absorptive current. Apical application of TC and TLC induced rapid (

Recent studies reported positive effects of taurine supplementation to plant protein-based diets on fish growth performance and feed conversion ratio but the mechanisms involved remain unclear. The objective of this work was to provide a first insight on the importance of taurine on dietary protein and lipid absorption and utilisation, in Senegalese sole (Solea senegalensis) juveniles fed plant-based diets. Seven practical diets were tested: a fishmeal-based diet (FM) and a plant protein-based formula (PP85) to which taurine was added in graded levels: 0.2% (TAU1), 0.3% (TAU2), 0.4% (TAUS), 0.7% (TAU4) and 1.5% (TAU5). Short-term metabolic trials were performed by tube-feeding the different experimental diets labelled with different tracers: C-14-taurine, C-14-triolein (as tracer of dietary triacylglycerol) and a mixture of C-14-amino acids. Feeding sole juveniles with the PP85 diet affected taurine metabolism, since a higher proportion of dietary taurine was directed to gallbladder and probably used for bile salt synthesis. Consistent with this, fish receiving the plant-based low taurine-content diets (PP85 and TAU1) displayed a poor triacyglycerol digestion capacity. A higher proportion of C-14-amino acids was retained in fish muscle when fed the TAU5 compared to the PP85 diet. These short-term trials suggested that taurine supplementation to a plant-based diet might be beneficial to sole performance by enhancing bile salt synthesis and triacylglycerol digestion as well as muscle amino acid retention. However, this dietary supplementation should be tested in long-term trials to verify the effect on fish growth and to assess the beneficial effects on protein and lipid metabolism suggested here. Statement of relevance in the general field of aquaculture (limited to 60 characters): Adding taurine to a plant-based diet is beneficial for sole (C) 2016 Elsevier B.V. All rights reserved. National Funds through Fundacao para a Ciencia e Tecnologia [PTDC/MAR/117047/2010] Fundacao para a Ciencia e Tecnologia, Portugal [SFRH/BDP/65578/2009] info:eu-repo/semantics/acceptedVersion

Broodstock rearing conditions and handling procedures should be optimized in aquaculture species in order to benefit fish welfare and guarantee optimal conditions for spawning. In teleosts, basal cortisol levels display daily rhythms, oscillating along the 24 h of the day. In this sense, handling fish at different moments of the day may lead to different stress responses. The present study aimed at investigating the optimal rearing conditions for Senegalese sole broodstock, considering ambient light intensity and handling time. The optimal light intensity (50, 100 or 200 lx) was investigated by measuring fish cortisol levels and monitoring locomotor activity rhythms under each intensity tested. Results showed a significant increase in cortisol levels of fish exposed to 200 lx, when compared to values obtained under 100 lx, accompanied by changes in locomotor activity rhythms in both tanks under study. These results suggested that 200 lx may be too high as light intensity for this species, whereas 100 lx seems to be more adequate. Also, daily rhythms of stress response were investigated in breeders from different origins (Wild and first generation, G1). Basal cortisol levels and cortisol stress response after an acute stressor (air exposure) were monitored at two distinct moments of the day (Mid-Light and Mid-Dark). Basal levels were higher during the day in the wild group, while G1 fish seemed to have lost the daily fluctuations in basal cortisol plasma levels, as well as their daily rhythms of locomotor activity. Both groups showed lower stress responses during night-time, an indication that this is an adequate period of the day to handle this species. Senegalese sole breeders born in captivity presented more pronounced stress responses when compared to wild fish, reflecting their different life history in terms of stress challenges. FP7/SME/2008/1; UID/Multi/04326/2019; DL 57/2016/CP1361/CT0007; DL 57/2016/CP1361/CT0033 info:eu-repo/semantics/publishedVersion

Nutrition shapes the individual physiological pathways prior to hatching, resulting in long-term effects on postnatal growth and physiological functions. The environment, including nutrition, determines the rate of myogenesis, the number and size of muscle fibres, the composition of sub-cellular organelles, the patterns of gene expression, influences protein turnover and the efficiency of protein deposition, among others. Moreover, protein retention efficiency and fish adaptive fitness have been said to be negatively correlated. In addition, high mortalities are normally observed in the marine larval stages in optimized farming conditions (70–80%) affecting fish production. The consequences and implications of early nutrition for aquaculture production is an important challenge for the future. Likewise, optimized feeds need to be provided in sufficient quantities and in a manner adequate to feeding behaviour, to fully express growth potential, while avoiding deterioration of water quality or disease problems caused by excessive feeding. Understanding the mechanisms that control early development and growth and their relation with nutrition are critical for the identification of time windows in development that introduce growth variation, impact growth potential, and affect viability and quality of juveniles.

Deoxynivalenol (DON), a Fusarium mycotoxin, is one of the most prevalent mycotoxins in aquafeeds. The toxicokinetics of DON are rarely studied in aquatic species. The present study used juvenile rainbow trout (Oncorhynchus mykiss) with a mean initial body weight of 7.72 ± 1.42 g in order to evaluate the pharmacokinetic behaviour and the metabolization of radiolabelled DON ([3H]-DON). In a first trial, 30 fish were tube-fed with four pellets containing a total of 125 ± 0.019 ng of [3H]-DON. At different sampling time points after feeding (1 h, 3 h, 6 h, 12 h or 24 h), the tissue distribution of the [3H]-DON was assessed by liquid scintillation counting. In a second trial, five fish were tube-fed four pellets containing a total of 663 ng of unlabelled-DON. Twenty-four hours after feeding, metabolites of DON excreted into the water were analysed by LC-MS/MS. [3H]-DON was detected in fish liver one hour after tube-feeding, indicating a rapid absorption of DON. In the first hour, [3H]-DON achieved its maximum in the gastro-intestinal tract (GIT) (20.56 ± 8.30 ng). However, 6.19 ± 0.83 ng of [3H]-DON was also detected in the water at this sampling time point. The fast excretion of [3H]-DON (above the average gastric emptying time of trout) might be related to its high-water solubility and consequent excretion with the fluid phase of the chyme. The amount of [3H]-DON in the GIT was stable during the first six hours. Such long transit time of DON through the GIT increases the potential for damage and absorption. The period between six and twelve hours seems to be the turning point in terms of DON excretion. Twelve hours after tube-feeding, the trout excreted 50.71 ± 22.17% of the tube-fed DON amount into water, while at the previous sampling time point (six hours) only 11.03 ± 6.09% were detected. These data suggest that an effective method for gastrointestinal DON detoxification in trout requires a period of action lower than six hours. In the present trial, no DON metabolites were detected in water.

Apparent digestibility coefficients (ADCs) of processed agro-food by-products were assessed in European seabass (Dicentrarchus labrax). Each experimental diet was obtained by replacing 300 g/kg of a commercial-based diet used as reference (REF) with a test ingredient: wheat germ (GERM), okara meal (OKA), poultry by-product meal (POULT), steam hydrolysed (FeHY) and enzyme-treated feather meal (FeENZ), beta-lactoglobulin (β-Lg) and peptide fractions >3,000 Da obtained from brewer's yeast (YeastP) and fish by-products (FishP). Dry matter ADC was highest in β-Lg (95%) and lowest in OKA (40%). Protein ADCs were high in β-Lg, FishP, GERM and POULT (>93%); intermediate in FeHY and FeENZ (85%–88%); and moderate in OKA and YeastP (70%–78%). The essential amino acids' ADC mean was above 91% in POULT, β-Lg, GERM and FishP, 84%–89% in FeHY and FeENZ and 73%–76% in YeastP and OKA. Energy ADC was highest in POULT and β-Lg (89%–95%) and lowest in YeastP and OKA (61%–64%). Lipid ADC was highest for POULT and GERM (100%). Phosphorus ADC was lowest in GERM (19%) and highest in β-Lg (88%). FeHY, FeENZ, POULT, GERM, FishP and β-Lg are highlighted as protein sources for European seabass.

In Zambia, every year some parts of the maize fields are abandoned post-planting. Reasons for this are not clearly known. In this paper, we examine the influence of soil and climatic factors on crop abandonment using a six-year (2007–2012) panel data by modeling the planted-to-harvested ratio (a good indicator of crop abandonment) using a fractional and linear approach. Therefore, for the first time, our study appropriately (as supported by the model specification tests that favour fractional probit over linear) models the fractional nature of crop abandonment. Regression results, which are not very different between the two specifications, indicate that, more than anything, high rainfall immediately after planting and inadequate fertilizer are the leading determinants of crop abandonment. In the agro-ecological region where dry planting takes place, low temperature during planting months negatively affects the harvested area. The results have implications on the sustainability of farming systems in the face of a changing climate.

We studied the effect of changes in dietary methionine (Met) supply in broodstock and first-feeding rainbow trout fry (offspring). Three plant-based diets differing in Met level (deficient, adequate or in excess of the established requirement) were fed to the broodstock (male and female) for 6 months prior to spawning (diets BD, BA and BE, respectively). The offspring from the parental Met-groups was then challenged in turn with the different Met fry-diets (FD, FA and FE, respectively) for 3 weeks from first-feeding. At spawning, females fed diet BD had significantly higher plasma total and LDL-cholesterol and slightly lower plasma triacylglycerol. Diet BD reduced female (but not male) growth, weight of spawn and egg size, but had no effect on relative fecundity. The free amino acid profile of oocytes was modified, with levels of Met and Cys correlating positively with the Met-levels of broodstock diets. SAM and SAH levels in oocytes followed the same pattern, as opposed to SAM/SAH ratio. At the swim-up stage, no significant effect of parental diet on fry weight was noted, whereas survival was the highest in fry from BE-broodstock. The subsequent 21-day fry feeding with different Met levels highly affected the daily growth index with a significant interaction between the parental-diet and fry-diet effects. The expression of a number of genes regulating sulfur amino acid metabolism was modified either directly by the dietary Met supply in both broodstock liver and in whole fry (e.g. BHMT1, GR, GST pi, MsrA1) or indirectly by the parental Met intakes as seen in the swim-up fry (e.g. BHMT1, MTR, GST pi, MsrA1). Importantly, long-lasting parental effects linked to broodstock Met-intake were seen in the fry, 21-days after first-feeding and irrespective of the fry diet (CTH, MsrA1, MsrB2, SOD2). Similarly, parental effects were noted on the gene expression of both NPY and POMC feeding peptides in fry prior to exogenous feeding which persisted for POMC in the 21-day fry. Parental effects were also demonstrated on the key myogenic gene Myog, on fMHC and GDH in swim-up fry, which persisted for GDH in 21-day fry. In summary, our results demonstrate that dietary Met levels of rainbow trout broodstock affect various traits in the offspring, some of which persisted during the first weeks of exogenous feeding. Further studies need to evaluate the long-term persistence of the parental effects over time and to elucidate the mechanisms, whether epigenetic or not. Statement of relevance: Determining the multiple effects of dietary methionine levels on reproductive, growth performance and metabolism in offspring will help improve formulations of low fish meal feeds for rainbow trout at sensitive life cycle stages. (C) 2016 Elsevier B.V. All rights reserved. EU [288925] F.C.T., Portugal - Fundo Social Europeu (POCH) and National Funds [SFRH/BPD/108389/2015] info:eu-repo/semantics/publishedVersion

Senegalese sole was one of the earliest identified candidate species with high potential for aquaculture diversification in the south of Europe.íts culture has been possible, and commercially attempted, for several decades, but intensive production has been slow to take off. This has been explained mostly by serious disease problems, high mortality at weaning, variable growth and poor juvenile quality. However, a strong and sustained research investment that started in the eighties has led to a better understanding of the requirements and particularities of this species. More recently, better management and technical improvements have been introduced, which have led to important progress in productivity and given a new impetus to the cultivation of Senegalese sole. As a result, the last 5 years have marked a probable turning point in the culture of sole towards the development of a knowledge-driven, competitive and sustainable industry. This review will focus on the main technical improvements and advances in the state of knowledge that have been made in the last decade in areas as diverse as reproductive biology, behaviour, physiology, nutritional requirements, modulation of the immune system in response to environmental parameters and stress, and characterization and mitigation of the main disease threats.ít is now clear that Senegalese sole has important particularities that differentiate it from other current and candidate marine aquaculture species, which bring about important challenges, some still unsolved, but also notable opportunities (e.g. a nutritional physiology that is better adapted to dietary vegetable ingredients), as will be discussed here. © 2014 Wiley Publishing Asia Pty Ltd., SM holds a Ramón y Cajal postdoctoral contract from the Spanish Ministry of Economy and Competitiveness (MINECO) and is supported by the European Commission Marie Curie Actions (FP7-PEOPLE-2010-RG, Project No. 274184). BC and SE are supported by Fundação para a Ciência e a Tecnologia, Portugal (SFRH/BPD/77210/2011 and SFRH/BPD/49051/2008, respectively).