Response Of Gilthead Seabream Research Articles

Behavioural Monitoring Underlines Habituation to Repeated Stressor Stimuli in Farmed Gilthead Sea Bream (Sparus aurata) Reared at a High Stocking Density.

A confinement stress test with 75% tank space reduction and behavioural monitoring through tri-axial accelerometers externally attached to the operculum was designed. This procedure was validated by demonstrating the less pronounced stress response in gilthead sea bream than in European sea bass (950-1200 g). Our study aimed to assess habituation to high stocking densities with such procedure in gilthead sea bream. Animals (420-450 g) were reared (June-August) in a flow-through system at two stocking densities (CTRL: 10-15 kg/m3; HD: 18-24 kg/m3), with natural photoperiod and temperature (21-29 °C), and oxygen levels at 5.2-4.2 (CTRL) and 4.2-3.2 ppm (HD). At the end, blood and muscle were sampled for haematology and transcriptomic analyses, and external tissue damage was assessed by image-based scoring. Four days later, fish underwent a 45 min confinement stress test over two consecutive days. HD fish showed reduced feed intake, growth rates and haematopoietic activity. Muscle transcriptome changes indicated a shift from systemic to local growth regulation and a primed muscle regeneration over protein accretion in HD animals with slight external injuries. After stress testing, HD fish exhibited a decreased recovery time in activity and respiration rates, which was shorter after a second stressor exposure, confirming habituation to high densities.

Hepcidin and piscidin modulation and antibacterial response in gilthead seabream (Sparus aurata) infected with Vibrio harveyi.

Vibriosis is an infectious disease that generates large economic losses in Mediterranean aquaculture. Vibrio harveyi is one of the marine bacteria causing this disease, it is widespread in the Mediterranean Sea and causes ulcers on the skin of the fish it infects. In addition, the skin is a route of entry and colonization of this pathogen. In this study, one group of fish was injected intraperitoneally with phosphate buffered saline (control group) and another with V. harveyi (infected group). At 4h after injection, samples of skin mucus, blood, skin, head kidney, liver, and spleen were collected to study the immune response generated. Liver histology showed notable alterations in hepatocyte morphology, such as increased vacuolization. Bactericidal activity was measured in skin mucus and serum against V. harveyi and V. anguillarum, different changes in this activity were recorded depending on the bacteria target and sample (skin mucus or serum) used. Gene expression of genes encoding hepcidins and piscidins (antimicrobial peptides) was performed in the mentioned organs. The results indicated a different expression according to the type of AMP and the tissue studied. Hepcidin appeared involved in all tissues studied while piscidins were in the spleen. In this study we have integrated hepcidin-piscidin modulation with the effects of infection on skin mucosa, serum and hepatocyte morphology. Knowing the changes produced in all these parameters improves the understanding of the infection in the first hours in sea bream and could have applications in the diagnosis or treatment of vibriosis in fish farms.

Modulation of gut microbiota and intestinal immune response in gilthead seabream (Sparus aurata) by dietary bile salt supplementation.

Given their role in lipid digestion, feed supplementation with bile salts could be an economic and sustainable solution to alterations in adiposity and intestinal inflammation generated by some strategies currently used in aquaculture. An important part of the metabolism of bile salts takes place in the intestine, where the microbiota transforms them into more toxic forms. Consequently, we aimed to evaluate the gut immune response and microbial populations in gilthead seabream (Sparus aurata) fed a diet supplemented with a blend of bile salts with proven background as a regulator of lipid metabolism and fat content. After the 90-day feeding trial, a differential modulation of the microbiota between the anterior and posterior intestine was observed. While in the anterior intestine the relative abundance of Desulfobacterota doubled, in the posterior intestine, the levels of Firmicutes increased and Proteobacteria, Actinobacteriota, and Campylobacterota were reduced when supplementing the diet with bile salts. Even so, only in the anterior intestine, there was a decrease in estimated richness (Chao1 and ACE indices) in presence of dietary bile salts. No significant differences were displayed in alpha (Shannon and Simpson indices) nor beta-diversity, showing that bile sales did not have a great impact on the intestinal microbiota. Regarding the gene expression profile in 2 h postprandial-fish, several changes were observed in the analyzed biomarkers of epithelial integrity, nutrient transport, mucus production, interleukins, cell markers, immunoglobulin production and pathogen recognition receptors. These results may indicate the development of an intestinal immune-protective status to tackle future threats. This work also suggests that this immune response is not only regulated by the presence of the dietary bile salts in the intestine, but also by the microbial populations that are in turn modulated by bile salts. After a fasting period of 2 days, the overall gene expression profile was stabilized with respect to fish fed the unsupplemented diet, indicating that the effect of bile salts was transient after short periods of fasting. On the balance, bile salts can be used as a dietary supplement to enhance S. aurata farming and production without compromising their intestinal health.

Feeding gilthead sea bream with increasing dietary bacterial single cell protein level: Implication on growth, plasma biochemistry, gut histology, and gut microbiota

Bacterial single cell protein (SCP) is considered a promising circular protein ingredient for aquafeed, due to the high protein content, and for the possibilities to grow them on different substrates such as organic waste thus leading to low environmental footprint and affordable production costs. Their use as raw material has been assessed in several farmed species, however, research on Mediterranean ones is still scarce. Hence, a study was undertaken to evaluate growth, plasma biochemistry, gut histology and gut microbiota (GM) response of gilthead sea bream (Sparus aurata) fed diets with increasing levels of bacterial SCP in comparison to a control without SCP. Three isonitrogenous and isolipidic extruded diets (44% protein; 19% lipid) were formulated with different bacterial SCP (derived from Corinebacterium glutamicum) level (10% SCP, SPC10; 15% SCP, SCP15; 20% SCP, SCP20) to replace vegetable protein ingredients (total replacement of soy protein concentrate and partial replacement of corn gluten), while a control diet (CTRL) was formulated without SCP. Fish groups of 45 individuals (initial weight: 75 g) were fed to visual satiation over 108 days. At the end of the trial there were no significant differences on growth, feed intake, feed conversion rate, protein efficiency ratio and protein apparent digestibility. Most of plasma parameters were found to be equal for all treatments, except for those related to nucleic acids molecules degradation such as phosphorus and urea which were higher in SCP10 and SCP20 compared to CTRL, respectively. No morphological alterations were found in the intestines of any fish analysed. Different responses of the overall GM structure in relation to the bacterial SCP inclusion level were detected. Specifically, SCP exerted a positive effect on GM internal diversity which increased at increasing dietary SCP inclusion level. In addition, SCP inclusion lead to increase in the abundance of Bacillus spp., taxa which can potentially support nutrition, immune system, and disease resistance. In conclusion, it seems feasible to include up to 20% of SCP dietary level for gilthead sea bream without compromising growth, feed efficiency and health parameters.

Transcriptomic and metabolomic integration to assess the response of gilthead sea bream (Sparus aurata) exposed to the most used insect repellent: DEET.

DEET is one of the most frequently detected insect repellents in the environment reaching concentrations of several μg L−1 in surface water. There is scarce information available regarding its mode of action in non-target organisms. Here, we have used an integrated metabolomic and transcriptomic approach to elucidate the possible adverse effects of DEET exposure in the marine fish gilthead sea bream (Sparus aurata). Individuals were exposed at an environmentally relevant concentration of DEET (10 μg L−1) for 22 days in a continuous flow-through system. Transcriptomic analysis revealed 250 differentially expressed genes in liver, while metabolomic analysis identified 190 differentially modulated features in liver and 98 in plasma. Multi-omic data integration and visualization allowed elucidation of the modes of action of DEET exposure, including: energy depletion through the disruption of carbohydrate and amino acids metabolisms, oxidative stress leading to DNA damage, lipid peroxidation, and damage to cell membrane and apoptosis. Activation of xenobiotic pathway as well as the inmune-inflammatory reaction was evidenced in the present work.

Dietary novel oils modulate neural function and preserve locomotor response in gilthead sea bream (Sparus aurata) juveniles by regulating synthesis and contents of fatty acids in brain

N-3 long-chain polyunsaturated fatty acids (LC-PUFA), particularly docosahexaenoic acid (22:6n-3, DHA), are the most important fatty acids with physiological significance in brain function of all vertebrates. Recently, novel lipid sources are available for the industry, including products obtained from microorganisms. Dietary fatty acid profiles may affect tissue composition and cell functioning. To determine the effect of novel lipid sources on behaviour and neural function in gilthead sea bream (Sparus aurata), juveniles were fed three isoproteic and isoenergetic diets containing either (FO), a microalgae oil combined with poultry oil (DD) or only poultry oil (PO). Behaviour response, brain fatty acid composition and relative expression of neurogenesis and neural activity related-genes in telencephalon were evaluated during 5 months of feeding with the experimental diets. Brain of sea bream fed diet PO showed the highest DHA content as well as increased desaturation and elongation products, contrary to the dietary pattern. This, together with an increased telencephalic fads2 expression denoted the effective activation of LC-PUFA synthesis and retention as a compensatory mechanism for a dietary deficiency. Furthermore, neurod6, bdnf and nos1 were proportionally upregulated in relation to the neural DHA content. Locomotor performance during cruising and escape responses was not affected by the experimental diets. However, there was a tendency for escape latency to be longest in fish fed PO and DD diets, and a relationship between escape latency and brain n-6 docosapentaenoic acid (DPA) contents was suggested. Indeed, the relation of n-6 DPA with behavioural deficits was shown previously in mammals and might deserve further attention and corroboration in fish as well in the future.

Multi-omic approach to evaluate the response of gilt-head sea bream (Sparus aurata) exposed to the UV filter sulisobenzone

Sulisobenzone (BP-4) is one of the benzophenone type UV filters most frequently detected in aquatic ecosystems. As a suspected endocrine disrupting compound, scarce information is available yet about other molecular effects and its mechanism of action. Here, we used an integrated transcriptomic and metabolomic approach to improve the current understanding on the toxicity of BP-4 towards aquatic species. Gilt-head sea bream individuals were exposed at environmentally relevant concentrations (10 μg L-1) for 22 days. Transcriptomic analysis revealed 371 differentially expressed genes in liver while metabolomic analysis identified 123 differentially modulated features in plasma and 118 in liver. Integration of transcriptomic and metabolomic data showed disruption of the energy metabolism (>10 pathways related to the metabolism of amino acids and carbohydrates were impacted) and lipid metabolism (5 glycerophospholipids and the expression of 3 enzymes were affected), suggesting oxidative stress. We also observed, for the first time in vivo and at environmental relevant concentrations, the disruption of several enzymes involved in the steroid and thyroid hormones biosynthesis. DNA and RNA synthesis was also impacted by changes in the purine and pyrimidine metabolisms. Overall, the multiomic workflow presented here increases the evidence on suspected effects of BP-4 exposure and identifies additional modes of action of the compounds that could have been overlooked by using single omic approaches.

In silico and gene expression analysis of the acute inflammatory response of gilthead seabream (Sparus aurata) after subcutaneous administration of carrageenin

Inflammation is one of the main causes of loss of homeostasis at both the systemic and molecular levels. The aim of this study was to investigate in silico the conservation of inflammation-related proteins in the gilthead seabream (Sparus aurata L.). Open reading frames of the selected genes were used as input in the STRING database for protein–protein interaction network analysis, comparing them with other teleost protein sequences. Proteins of the large yellow croaker (Larimichthys crocea L.) presented the highest percentages of identity with the gilthead seabream protein sequence. The gene expression profile of these proteins was then studied in gilthead seabream specimens subcutaneously injected with carrageenin (1%) or phosphate-buffered saline (control) by analyzing skin samples from the injected zone 12 and 24 h after injection. Gene expression analysis indicated that the mechanisms necessary to terminate the inflammatory response to carrageenin and recover skin homeostasis were activated between 12 and 24 h after injection (at the tested dose). The gene analysis performed in this study could contribute to the identification of the main mechanisms of acute inflammatory response and validate the use of carrageenin as an inflammation model to elucidate these mechanisms in fish.

Effects of mannan oligosaccharide and synbiotic supplementation on growth performance and immune response of Gilthead Sea Bream ( Sparus aurata ) before and after thermal stress

In this study, the effects of mannan oligosaccharides (MOS) and a commercial synbiotic (Biomin) supplemented diets on growth and survival performance of gilthead sea bream after eight weeks culture at 27ºC and then ten days at 34ºC were evaluated. Before heat stress, the Biomin supplemented diet significantly increased the number of red blood cells (RBC) and eosinophils of the fish. The MOS supplemented diet increased the haematocrit, haemoglobin, lymphocytes, white blood cells (WBC) and monocytes significantly. Total immunoglobulin was significantly higher in both supplemented diets in contrast to the control group. After thermal stress, experimental diets increased the monocyte counts, lysozyme, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) of the fish significantly, also the lowest concentration of cortisol was observed in the fish fed with Biomin supplemented diet. Gene expression profile showed a significantly higher expression of innate immune genes (il-1β, ghr-1, tnf-α and igf-1) in fish fed with Biomin and MOS supplemented diets. However, after thermal challenge, inflammatory response was significantly lower the fish fed with the supplemented diets. According to our results, MOS and Biomin supplemented diets enhance the immune health status of the gilthead sea bream during thermal stress.

Dietary supplementation with Gracilaria sp. by-products modulates stress response, antioxidant and immune systems of gilthead seabream (Sparus aurata) exposed to crowding

By-products obtained from seaweed processing appear as a promissory source of bioactive compounds that could be included in aquafeed to improve the sustainability of aquaculture. Therefore, the present study aimed to evaluate the effects of dietary supplementation with by-products obtained from the seaweed Gracilaria sp. on the stress response, antioxidant, and immune systems of gilthead seabream (Sparus aurata) exposed to an acute crowding event. For this, fish were fed six experimental diets: (i) a diet containing a commercial antioxidant (CTR), (ii) a diet without antioxidant addition (NO-AOX), (iii) a NO-AOX diet with 0.5% ethanolic extract (0.5% EE), (iv) a NO-AOX diet with 5% ethanolic waste (5% EEW), (v) a NO-AOX diet with 5% agar waste (5% AW), and (vi) a NO-AOX diet with 2.5% agar waste (2.5% AW) for 59 days. After this period, fish were exposed to 1 h of crowding (100 kg m−3) and sampled immediately after 0 and at 24 h post-crowding. Our results showed that the fish-fed AW and EEW diets showed lower plasma cortisol levels than those fed the CTR diet, suggesting an improved stress response to crowding on these groups. No differences were detected on hepatic lipid peroxidation levels between dietary groups after crowding. However, the fish fed 0.5% EE diet showed lower hepatic glutathione reductase (GR) activity than fish fed the NO-AOX diet. Additionally, glutathione peroxidase (GPx) activity was decreased in the liver of fish fed 2.5% and 5% AW diets compared with the other dietary groups. Concerning immune parameters, the plasma peroxidase activity was higher in fish fed the 2.5% AW diet than fish fed NO-AOX or 5% EEW diets, suggesting a modulatory effect of dietary AW at 2.5% in the humoral immune response in crowded fish. Results suggest that dietary supplementation with AW, particularly when supplemented at 2.5%, could be used as a dietary tool to mitigate oxidative stress when improving immune response in gilthead seabream exposed to a stressor as crowding.

Immune gene expression in gilthead seabream (Sparus aurata) after Lymphocystis disease virus (LCDV-Sa) challenge resulting in asymptomatic infection.

To determine the immune gene expression response of gilthead seabream (Sparus aurata) that is experimentally infected with the lymphocystivirus LCDV-Sa. Viral DNA and transcripts were detected by qPCR in all samples from fish injected with LCDV-Sa, demonstrating that the virus establish a systemic and asymptomatic infection. The expression of 23 immune-related genes was also analysed by RT-qPCR in the head kidney (HK) and intestine at several times post-infection (dpi). In HK, the expression of five type I interferon (IFN)-related genes (ifn, irf3, mx2, mx3 and isg15), il10 and ck10 was upregulated at 1-3 dpi, while genes related to the inflammation process (tnfα, il1ß, il6, casp1) were not differentially expressed or even downregulated. The expression profile in the intestine was different regarding type I INF-related genes. An upregulated c3 and ighm expression was observed in both HK and intestine at 3-8 dpi. Finally, the transcription of nccrp1 and mhcIIα was induced in HK, whereas tcrβ expression was downregulated in both organs. LCDV-Sa seems to trigger an immune response in gilthead seabream characterized by a partial activation of type I IFN system and a lack of systemic inflammatory response which may be related to viral persistence. The immune response observed in gilthead seabream infected by LCDV-Sa could be implicated in the establishment of an asymptomatic persistent infection.

Effects of tetra (Cotinus coggygria) and common mallow (Malva sylvestris) plant extracts on growth performance and immune response in Gilthead Sea bream (Sparus aurata) and European Sea bass (Dicentrarchus labrax)

In this study, effects of tetra (T) (Cotinus coggygria) and common mallow (CM) (Malva sylvestris) aqueous methanolic extracts on growth performance and immune response in gilthead sea bream, Sparus aurata (19.92 ± 0.40 g) and European sea bass, Dicentrarchus labrax (18.66 ± 0.86 g) were investigated. The experiment was designed as 5 treatment groups: control (0 mg/kg), 500 mg kg−1 (CM500, T500) and 1000 mg kg−1 (CM1000, T1000) for each fish species. Fish were fed the diet supplemented with tetra and common mallow extracts for 60 days and blood samples were collected on the 30th and 60th days of the study. Immunological parameters, such as respiratory burst, phagocytic, lysozyme, myeloperoxidase and bacterial killing activities were determined. At the end of feeding trial, fish were challenged with Vibrio anguillarum infection and survival rate was recorded. Final weight of the both experimental fishes were significantly increased in CM treatment groups compared to control (P < .05). No difference was observed in FCR values in sea bass (P > .05), whereas a decrease in FCR was noticed in CM treatment groups of sea bream (P < .05). Moreover, oxidative radical production was significantly increased in sea bass in all treatment groups at all sampling times. In sea bream, it increased in T500 and CM500 groups on 30th day, and only in CM groups on 60th day compared to control (P < .05). Phagocytic activity was significantly increased in sea bass received only tetra extract on the 30th day compared to control (P < .05) and in all treatment groups except T1000 on 60th day. In sea bream, phagocytic activity was enhanced in all treatment groups on 30th day and only in CM500 and T1000 groups on 60th day. Lysozyme activity was elevated in all treatment groups at all sampling times in sea bass, and in T500 and T1000 groups of sea bream. Myeloperoxidase activity increased in both the fish species at any sampling time compared to that of control (P < .05). Survival rate was significantly higher in treatment groups compared to control of both fish species. These results suggest that both tetra and common mallow could be used as immunostimulants for European sea bass and sea bream with an added advantage of common mallow as a growth promoter.

Hints on T cell responses in a fish-parasite model: Enteromyxum leei induces differential expression of T cell signature molecules depending on the organ and the infection status

BackgroudEnteromyxum leei is a myxozoan parasite that produces a slow-progressing intestinal disease. This parasite invades the paracellular space of the intestinal epithelium and progresses from the posterior to the anterior intestine. The aim of the present study was to gain insights into fish T cell responses in the gilthead sea bream-E. leei infection model using a PCR-array with 30 signature molecules for different leukocyte responses in head kidney, spleen, anterior and posterior intestine.ResultsThe PCR-array results suggest that E. leei induced migration of T cells from head kidney to intestines where TH1, CTL and TH17 profiles were activated and kept in balance by the upregulation of regulatory cytokines. These results were partially validated by the use of cross-reacting antibodies and BrdU immunostaining to monitor proliferation. Zap70 immunostaining supported the increased number of T cells in the anterior intestine detected by gene expression, but double staining with BrdU did not show active proliferation of this cell type at a local level, supporting the migration from lymphohaematopoietic tissues to the site of infection. Global analyses of the expression profiles revealed a clear separation between infected and exposed, but non-infected fish, more evident in the target organ. Exposed, non-infected animals showed an intermediate phenotype closer to the control fish.ConclusionsThese results evidence a clear modulation of the T cell response of gilthead sea bream upon E. leei infection. The effects occurred both at local and systemic levels, but the response was stronger and more specific at the site of infection, the intestine. Altogether, this research poses a promising basis to understand the response against this important parasite and establish effective preventive or palliative measures.

A time course study of glucose levels and innate immune response in gilthead seabream (Sparus aurata L.) after exposure to clove oil-eugenol derived anaesthetic

Clove oil is used as an anaesthetic for many species of fish worldwide; however, relatively few studies have assessed its effects on the innate immune response on these species. The present work aimed to investigate the effects of clove oil-eugenol derived anaesthetic on some humoral and cellular immune response in gilthead seabream (Sparus aurata L.). To compare with an unexposed control group, fish were exposed to 55 ppm clove oil for 5 min, before being sampled at 1, 24 and 48 h post-exposure. Serum glucose level was also measured to obtain information on the fish physiological response after clove oil anaesthesia. One hour after exposure the haemolytic complement activity of fish was lower than in the unexposed group. By contrast, the leucocyte peroxidase activity in head-kidney was significantly stimulated 24 h after exposure to clove oil-eugenol. The rest of innate immune parameters evaluated and the glucose levels not were affected by clove oil exposure at any sampling point. Overall, the use of clove oil at 55 ppm as anaesthetic did not seem to alter the innate immune response and neither did it trigger a stress response. The use of clove oil-eugenol derived had become common practice in aquaculture, and its use with gilthead seabream can be considered safe as it does not cause immunodepression in anesthetized fish.

Aluminum adjuvant potentiates gilthead seabream immune responses but induces toxicity in splenic melanomacrophage centers

A key goal of a successful vaccine formulation is the strong induction of persistent protective immune responses without producing side-effects. Adjuvants have been proved to be successful in several species at inducing increased immune responses against poorly immunogenic antigens. Fish are not the exception and promising results of adjuvanted vaccine formulations in many species are needed. In this study, over a period of 300 days, we characterized the apparent damage and immune response in gilthead seabream immunized by intraperitoneal injection with the model antigen keyhole limpet hemocyanin (KLH) alone or formulated with Montanide ISA water-in-oil (761 or 763), or Imject™ aluminum hydroxide (aluminium), as adjuvants. Throughout the trial, external tissue damage was examined visually, but no change was observed. Internally, severe adhesions, increased fat tissue, and hepatomegaly were recorded, but, without impairing animal health. At 120 days post priming (dpp), histopathological evaluations of head-kidney, spleen and liver revealed the presence of altered melanomacrophage centers (MMC) in HK and spleen, but not in liver. Surprisingly, in all aluminium treated fish, classical stains unmasked a toxic effect on splenic-MMC, unequivocally characterized by a strong cell depletion. Furthermore, at 170 dpp transmission electron microscopy confirmed this data. Paradoxically, at the same time powerful immune responses were recorded in most vaccinated groups, including the aluminium treatment. Whatever the case, despite the observed adhesions and MMC depletion, fish physiology was not affected, and most side-effects were resolved after 300 dpp. Therefore, our data support adjuvant inclusion, but strongly suggest that use of aluminium must be further explored in detail before it might benefit the rational design of new vaccination strategies in aquaculture.

Response of Gilthead Seabream (Sparus aurata L., 1758) Larvae to Nursery Odor Cues as Described by a New Set of Behavioral Indexes

Temperate marine fish larvae use a series of environmental cues (e.g., olfactory, hearing, visual) to mediate the selection of nursery habitats. However, habitat selection may vary according to individuals' physiological condition. Therefore, this study aimed to determine the ability of gilthead seabream (Sparus aurata L., 1758) larvae to utilize natural odor cues to locate nursery habitats along ontogeny and to examine how it varies with individual's physiological condition. The hypothesis being tested is that S. aurata larvae prefer coastal rocky reefs as nursery areas, but they might use coastal lagoons as nursery grounds—ecosystems known for their productivity—if under starvation conditions, as a compensatory mechanism to avoid slow growth or even death. A choice-chamber experiment was used to investigate the behavioral responses of satiated and starved laboratory-reared S. aurata larvae, along ontogeny (pre-flexion, flexion, post-flexion), to water collected in a coastal artificial rocky reef and a coastal lagoon. The physiological condition of S. aurata larvae was determined by analyzing several biochemical condition indices. Complementarily, a new set of four preference indexes were developed—Choice-Chamber Preference Indexes—and discussed to provide a clear measure of the behavioral changes of a species along ontogeny by balancing all the behavioral choices made during the experimental trials, including the unresponsive behavior. A developmental threshold was identified at 24 days post-hatching, before which insufficient swimming capability disabled responsive behavior. Beyond this threshold, post-flexion larvae preferred rocky coastal water over lagoon water, even if under starvation conditions or poor physiological condition, despite the fact that the unresponsive behavior was largely predominant. S. aurata larvae displayed a cautionary behavioral strategy, so the compensatory mechanisms to ensure metapopulation stability and resilience have to rely on their feeding plasticity and on being a batch-spawning species (i.e., diversified bet-hedging strategy) to compensate the lack of apparent behavioral plasticity.

Effects of graded dietary levels of soy protein concentrate supplemented with methionine and phosphate on the immune and antioxidant responses of gilthead sea bream (Sparus aurata L.)

The effects of a dietary soy protein concentrate (SPC) as a fish meal (FM) substitute, on selected innate immune responses, the oxidative status, hepatic and intestinal morphology of gilthead sea bream, Sparus aurata, were evaluated after a three-month feeding trial. Isonitrogenous (45% crude protein) and isoenergetic (23 kJ/g gross energy) diets with 20% (SPC20), 40% (SPC40) and 60% (SPC60) of SPC inclusion, supplemented with methionine and phosphate, were evaluated against a diet containing FM as the sole protein source. Diets were allocated in triplicate groups of 26-g fish (8 kg m−3/tank) and administered for three months. Immune responses were evaluated by performing immunological assays in blood (respiratory burst activity) and serum (myeloperoxidase content, bacteriolytic and lysozyme activity), as well as by gene expression analysis of immune-associated genes (MHCIIα, β2m, CSF-1R, NCCRP-1, TGF-β1, HSP70) in the head kidney and distal intestine. In addition, oxidative stress was evaluated by measuring the activity of liver enzymes associated with the antioxidant system. The respiratory burst activity of blood was significantly decreased in the SPC40 group, while serum myeloperoxidase content and bacteriolytic and lysozyme activities were affected. Significantly higher expression levels of NCCRP-1 and HSP70 were found in SPC60 head kidneys, while increased intestinal MHCIIα and NCCRP-1 transcripts were observed in SPC40. Hepatic antioxidant enzyme activity of glutathione reductase and glutathione-S-transferase was significantly enhanced in the SPC40 and SPC60 groups, while superoxide dismutase activity was increased only in the SPC40 group. Moreover, increased lipid accumulation in the enterocytes of the distal intestine was observed in the SPC60 group. Overall, a three-month feeding period with diets over 40% of dietary SPC inclusion as a FM substitute, indicated increases on immune and antioxidant enzyme responses, suggesting the dietary SPC levels that gilthead sea bream can tolerate.

Stress effects of amyloodiniosis in gilthead sea bream Sparus aurata.

Amyloodiniosis is a parasitological disease caused by one of the most common and important parasitic dinoflagellates in fish, Amyloodinium ocellatum (Brown), that represents a major bottleneck for gilthead seabream semi-intensive aquaculture in Southern Europe. In this experiment, we analyzed some metabolic, osmoregulatory and stress indicators to elucidate some of the physiological responses of gilthead sea bream when exposed to an A. ocellatum outbreak. We observed significant differences between Control and Infection groups in the cortisol, lactate and gill Na+/K+-ATPase (NKA) activity levels but that glucose, osmolarity, pH and total protein did not present such differences. This could indicate that the presence of the parasite induced a stress response, possibly enhancing the metabolization of glucose and subsequently lactate to cope with the higher energy requirements of the organism. There was also a decrease in gill NKA activity possibly due to severe epithelial damage and increased mucus production caused by the parasite A. ocellatum, which could induce anoxia and osmoregulatory impairment in the organism. However, further works must be performed to fully understand the physiological reactions of fish for A. ocellatum outbreaks.

Molecular details on gilthead sea bream (Sparus aurata) sensitivity to low water temperatures from 1H NMR metabolomics

Biometric and metabolic responses of gilthead sea bream to cold challenge are described following a growth trial divided into three water temperature steps, namely cooling, cold maintenance and recovery. Experimental data provide a useful description of fish response to thermal stress at both zootechnical and molecular level. Although no mortality has been observed, Nuclear Magnetic Resonance-based metabolomics confirms the marked sensitivity of this fish species to low water temperature, and explains some key molecular events associated to fish response to cold. Increase in hepatosomatic index is associated to liver fat accumulation, as a consequence of lipid mobilization from muscle and other extrahepatic tissues, and metabolic rearrangements linked to homeoviscous adaptation of cellular membranes are observed. Following primary responses to descending temperature from 18°C to 11°C, the energetic metabolism (insulin signaling, glycolysis) is first clearly affected; then, at constant low water temperature (11°C), the most perturbed metabolic pathways are related to methionine cycle in liver, while osmoregulatory function is exerted by TMAO in muscle. Water temperature recovery from 11°C to 18°C stimulates gluconeogenesis and glycogen synthesis activities at hepatic level, although the rate of a thermo-compensatory response seems to be slower than that of the cooling phase. The obtained results are intended to guide novel high-performance feed formulations for gilthead sea bream reared during winter.

Vessel noise pollution as a human threat to fish: assessment of the stress response in gilthead sea bream (Sparus aurata, Linnaeus 1758).

This study examined the effects of boat noise pollution on the stress indices of gilthead sea bream (Sparus aurata, Linnaeus 1758). To assess the stress response in these fish, biometric values and plasma parameters such as ACTH, cortisol, glucose, lactate, haematocrit, Hsp70, total protein, cholesterol, triglycerides and osmolarity were analysed. After acclimatization of the animals, the experiment was carried out in a tank fitted with underwater speakers where the fish were exposed to sound treatments (in duplicate) consisting of: 10 days of no sound (control treatment; the animals were only exposed to the experimental tank's background noise) and 10 days of noise derived from original recordings of motor boats, including recreational boats, hydrofoil, fishing boat and ferry boat (vessel noise treatment). The exposure to noise produced significant variations in almost all the plasma parameters assessed, but no differences were observed in weights and fork lengths. A PERMANOVA analysis highlighted significantly increased values (p < 0.05) of ACTH, cortisol, glucose, lactate, haematocrit, Hsp70, cholesterol, triglycerides and osmolarity in the fish exposed to vessel noise for 10 days. This study clearly highlights that anthropogenic noise negatively affects fish, and they are valuable targets for detailed investigations into the effects of this global pollutant. Finally, these experimental studies could represent part of the science that is able to improve the quality of the policies related to management plans for maritime spaces (Marine Strategy Framework Directive 56/2008 CE) that are aimed at stemming this pollutant phenomenon.