Impact of Hydroxytyrosol-Rich Extract Supplementation in a High-Fat Diet on Gilthead Sea Bream (Sparus aurata) Lipid Metabolism.

High-fat diets (HFDs) enhance fish growth by optimizing nutrient utilization (i.e., protein-sparing effect); however, their potential negative effects have also encouraged the search for feed additives. This work has investigated the effects of an extract rich in a polyphenolic antioxidant, hydroxytyrosol (HT), supplemented (0.52 g HT/kg feed) in a HFD (24% lipid) in gilthead sea bream (Sparus aurata). Fish received the diet at two ration levels, standard (3% of total fish weight) or restricted (40% reduction) for 8 weeks. Animals fed the supplemented diet at a standard ration had the lowest levels of plasma free fatty acids (4.28 ± 0.23 mg/dL versus 6.42 ± 0.47 in the non-supplemented group) and downregulated hepatic mRNA levels of lipid metabolism markers (ppara, pparb, lpl, fatp1, fabp1, acox1, lipe and lipa), supporting potential fat-lowering properties of this compound in the liver. Moreover, the same animals showed increased muscle lipid content and peroxidation (1.58- and 1.22-fold, respectively, compared to the fish without HT), suggesting the modulation of body adiposity distribution and an enhanced lipid oxidation rate in that tissue. Our findings emphasize the importance of considering this phytocompound as an optimal additive in HFDs for gilthead sea bream to improve overall fish health and condition.

Repeated hypoxic episodes allow hematological and physiological habituation in rainbow trout.

Introduction: Under climate change, the increase in temperature in aquatic environments may induce oxygen depletion. In extreme cases, low oxygen may become a limiting factor for fish, thus generating stress. In addition, consecutive hypoxic episodes may complicate the recovery of individuals and hinder their ability to modulate physiological and biochemical responses to maintain homeostasis. Thus, the aim of this study was to determine the hematological and physiological responses of rainbow trout under a condition of repeated hypoxic and manipulation stresses at three different time points. Methods: Every hypoxic episode consisted of exposing the fish to low dissolved oxygen concentrations (2 mgO2/L for 1 h). Following the exposure, the fish were allowed to recover for 1 h, after which they were sampled to investigate hematological and physiological parameters. Results and discussion: The results showed a pattern of habituation reflected by values of hematocrit, hemoglobin, and mean corpuscular volume, indicating a certain ability of rainbow trout to resist this type of repeated hypoxic events, provided that the fish can have some recovery time between the exposures.

Ontogeny of the digestive enzyme activity of the pikeperch (Sander lucioperca) under culture condition.

The pikeperch (Sander lucioperca) is a species with a high potential for aquaculture and a valuable food with high market acceptance. The aim of the study was to evaluate the functional ontogeny of digestive enzyme of pikeperch from hatching to 45 days-post fertilization, 777 degree-day (DPF, dd) under culture condition. The average total length (TL) of larvae measured at hatching was 3.6 ± 0.4 mm (5 DPF; 67 dd) and at the end of experiment (45 DPF, 777 dd) was 27.1 ± 1.1 mm. The survival rate was 80-90% during the experiment period. Inhibition zimography reveals the presence of nine bands with proteolytic activity in the digestive tract of juvenile pikeperch. Zimography results during the ontogeny revealed that in larvae at 8 DPF (108 dd) and 13 DPF (189 dd), three bands were presented. The variations observed in the enzymatic activity reflected a high amount of total protease activity at 10 DPF (133.5 dd). Regarding pepsin, its activity was observed for the first time at 26 DPF (378.9 dd). Lipase activity remained constant from hatching to 26 DPF (378.9 dd). The highest amount of α-amylase activity was detected at 15 DPF (211.5 dd) and 45 DPF (777 dd). The low lipase enzyme activity suggested that live feeds with low lipid were more suitable than diets containing high lipid levels; larvae had also early capability to digest nutrient-dense diet that was high in protein. According to results the pikeperch larvae possess after the exogenous feeding, a functional digestive system with high activities that indicated the gradual development of the digestive system.

Cysteamine improves growth and the GH/IGF axis in gilthead sea bream (Sparus aurata): in vivo and in vitro approaches.

Aquaculture is the fastest-growing food production sector and nowadays provides more food than extractive fishing. Studies focused on the understanding of how teleost growth is regulated are essential to improve fish production. Cysteamine (CSH) is a novel feed additive that can improve growth through the modulation of the GH/IGF axis; however, the underlying mechanisms and the interaction between tissues are not well understood. This study aimed to investigate the effects of CSH inclusion in diets at 1.65 g/kg of feed for 9 weeks and 1.65 g/kg or 3.3 g/kg for 9 weeks more, on growth performance and the GH/IGF-1 axis in plasma, liver, stomach, and white muscle in gilthead sea bream (Sparus aurata) fingerlings (1.8 ± 0.03 g) and juveniles (14.46 ± 0.68 g). Additionally, the effects of CSH stimulation in primary cultured muscle cells for 4 days on cell viability and GH/IGF axis relative gene expression were evaluated. Results showed that CSH-1.65 improved growth performance by 16% and 26.7% after 9 and 18 weeks, respectively, while CSH-3.3 improved 32.3% after 18 weeks compared to control diet (0 g/kg). However, no significant differences were found between both experimental doses. CSH reduced the plasma levels of GH after 18 weeks and increased the IGF-1 ones after 9 and 18 weeks. Gene expression analysis revealed a significant upregulation of the ghr-1, different igf-1 splice variants, igf-2 and the downregulation of the igf-1ra and b, depending on the tissue and dose. Myocytes stimulated with 200 µM of CSH showed higher cell viability and mRNA levels of ghr1, igf-1b, igf-2 and igf-1rb compared to control (0 µM) in a similar way to white muscle. Overall, CSH improves growth and modulates the GH/IGF-1 axis in vivo and in vitro toward an anabolic status through different synergic ways, revealing CSH as a feasible candidate to be included in fish feed.

Effects of Dietary Vegetable Oil Mixtures including Soybean Oil on Intestinal Oxidative Stress in Gilthead Sea Bream (Sparus aurata).

Fish oil is commonly replaced by vegetable oils in sea bream diets, but little is known about their effects on intestinal health regarding oxidative stress biomarkers. The negative effects of lipid peroxidation on digestive mucosa could have consequences in animal nutrition and welfare. In this study, five isonitrogenous (46%) and isolipidic (22%) diets with 75% of vegetable oils inclusion were evaluated: soybean oil (S) alone or different mixtures containing soybean oil with linseed (SL), linseed and rapeseed (SLR), linseed and palm (SLP), and linseed, rapeseed, and palm (SLRP). Gilthead sea bream juveniles were fed twice a day for 18 weeks. Pyloric caeca and proximal intestine samples were collected 24 h post feeding for lipid peroxidation (LPO), antioxidant enzyme activities (SOD, CAT, GPx, GST, and GR) and gene expression analyses. Pyloric caeca presented larger unhealthy changes in oxidative status than proximal intestine. Although SL-fed fish showed the highest antioxidant activities, they were unable to cope with LPO that in pyloric caeca was 31.4 times higher than in the other groups. Instead, SLP fish presented the best oxidative status, with low LPO levels, antioxidant enzyme activities, and gene expression. In summary, between the vegetable oils dietary mixtures tested, SPL would maintain better intestinal health.

Rainbow trout integrated response after recovery from short-term acute hypoxia.

Overcoming a stress situation, such as hypoxia episodes, which involve an allostatic load, will depend on the ability of fish to modulate physiological and biochemical systems to maintain homeostasis. The aim of the study was to determine the integrated stress response after acute hypoxia of the rainbow trout considering the different elements and areas of the stress response: systemic and mucosal, local and global, and from the systemic hypothalamic-pituitary-interrenal axis to skin mucosa. For this purpose, trout were subjected to acute hypoxia (dissolved O2 down to 2 mg/L) for 1 h and then recovered and sampled at 1, 6, and 24 h after reoxygenation. Physiological responses were significantly affected by hypoxic stress and their interaction with time after the challenge, being significant for plasma lactate and cortisol levels, in both plasma and skin mucus. At the central brain level, only trh expression was modulated 1 h after hypoxia which indicates that brain function is not heavily affected by this particular stress. Unlike the brain, the head kidney and skin were more affected by hypoxia and reoxygenation. In the head kidney, an upregulation in the expression of most of the genes studied (gr, il1ß, il6, tgfß1, lysozyme, caspase 3, enolase, hif-1, myoglobin, sod2, gpx, gst, and gsr) took place 6 h after recovery, whereas only hsp70 and il10 were upregulated after 1 h. On the contrary, in the skin, most of the analyzed genes showed a higher upregulation during 1 h after stress suggesting that, in the skin, a local response took place as soon as the stressor was detected, thus indicating the importance of the skin in the building of a stress response, whereas the interrenal tissue participated in a later time point to help prevent further alteration at the central level. The present results also show that, even though the stressor is a physical/environmental stressor, all components of the biological systems participate in the regulation of the response process and the recovery process, including neuroendocrine, metabolism, and immunity.

Hydroxytyrosol-rich extract from olive juice as an additive in gilthead sea bream juveniles fed a high-fat diet: Regulation of somatic growth.

The dietary inclusion of plant-based products in fish feeds formulation is required for the sustainable development of aquaculture. Moreover, considering functional diets, hydroxytyrosol, one of the major phenolic compounds found in olives (Olea europaea), has been identified as a potential candidate to be used in the aquafeeds industry due to its health promoting abilities. The aim of this study was to evaluate the effects of the inclusion of an olive juice extract rich in hydroxytyrosol as an additive (0.52 g HT/kg feed) in a high-fat (24% lipids) diet in gilthead sea bream (Sparus aurata) juveniles. Moreover, the experimental diets, with or without the extract, were administered daily at a standard (3% of total biomass in the tank) or restricted ration (40% reduction) for 8-9 weeks. Growth and biometric parameters, insulin-like growth factor 1 (IGF-1) plasma levels and growth hormone/IGF axis-, myogenic- and osteogenic-related genes expression in liver, white muscle and/or bone were analyzed. Moreover, in vitro cultures of vertebra bone-derived cells from fish fed the diets at a standard ration were performed at weeks 3 and 9 to explore the effects of hydroxytyrosol on osteoblasts development. Although neither body weight or any other biometric parameter were affected by diet composition after 4 or 8 weeks, the addition of the hydroxytyrosol-rich extract to the diet increased IGF-1 plasma levels, regardless of the ration regime, suggesting an anabolic condition. In muscle, the higher mRNA levels of the binding protein igfbp-5b and the myoblast fusion marker dock5 in fish fed with the hydroxytyrosol-rich diet suggested that this compound may have a role in muscle, inducing development and a better muscular condition. Furthermore in bone, increased osteogenic potential while delayed matrix mineralization after addition to the diet of the olive juice extract was supported by the upregulated expression of igf-1 and bmp4 and reduced transcript levels of osteopontin. Overall, this study provides new insights into the beneficial use of hydroxytyrosol as a dietary additive in gilthead sea bream functional diets to improve muscle-skeletal condition and, the aquaculture industry.