Transcriptional Evaluation of Neuropeptides, Hormones, and Tissue Repair Modulators in the Skin of Gilthead Sea Bream (Sparus aurata L.) Subjected to Mechanical Damage.

The skin of bony fish is the first physical barrier and is responsible for maintaining the integrity of the fish. Lesions make the skin vulnerable to potential infection by pathogens present in the aquatic environment. In this way, wound repair has barely been studied in gilthead sea bream. Thus, this study investigated the modulation of peripheral neuro-endocrine and tissue repair markers at the transcriptional level in the skin of teleost fish subjected to mechanical damage above or below the lateral line (dorsal and ventral lesions, respectively). Samples were evaluated using RT-qPCR at 2-, 4-, and 20-days post-injury. Fish with a ventral lesion presented a trend of progressive increase in the expressions of corticotropin-releasing hormone (crh), pro-opiomelanocortin-A (pomca), proenkephalin-B (penkb), cholecystokinin (cck), oxytocin (oxt), angiotensinogen (agt), and (less pronounced) somatostatin-1B (sst1b). By contrast, fish with a dorsal lesion registered no significant increase or biological trend for the genes evaluated at the different sampling times. Collectively, the results show a rapid and more robust response of neuro-endocrine and tissue repair markers in the injuries below than above the lateral line, which could be attributable to their proximity to vital organs.

Serum proteinogram of gilthead seabream (Sparus aurata) and European sea bass (Dicentrarchus labrax) as a new useful approach for detecting loss of haemostasis.

Proteinograms, a semiquantitative analytical method that separates proteins into multiple bands, have not been explored in teleosts for diagnostic or prognostic purposes. This study aimed to establish reference values for proteinograms in the serum of gilthead seabream (Sparus aurata) and European sea bass (Dicentrarchus labrax), two important farmed fish species in the Mediterranean region. Serum proteins were studied using SDS-PAGE, electropherogram, and HPLC-mass spectrometry. SDS-PAGE analysis revealed four major bands of proteins around 11, 25, 70, and 100 kDa in the serum of gilthead seabream and European sea bass. Electropherogram results showed that a protein with a molecular weight of 76.8 kDa was the most abundant protein in the serum of gilthead seabream, while a peak of 75.5 kDa was the most abundant in European sea bass. HPLC-mass spectrometry detected 87 proteins and 119 proteins in the serum of gilthead seabream and European sea bass, respectively, including α1-globulins, α2-globulins, ß-globulins, and γ-globulins. Notably, the albumin sequence was not detected in either of the two species. These results help to characterize the serum protein profile and to establish reference proteinograms for these two fish species. They also provide a basis for the development of novel approaches for the rapid detection of loss of haemostasis due to stress, health disorders or disease in farmed fish.

Effects of dietary astaxanthin on immune status and lipid metabolism in gilthead seabream (Sparus aurata).

Astaxanthin (AX) is a carotenoid known to have one of the highest documented antioxidant capacities and has attracted considerable scientific and commercial interest. The incorporation of AX into aquaculture practices has been associated with improved pigmentation, modulation of the immune and endocrine systems, stress reduction, reproductive efficiency and general fish health. This study describes the effects of dietary AX (0, control, 20, 100 and 500 mg kg-1 AX per kg of diet) for 15 and 30 days on growth performance, immune and antioxidant status, histology and gene expression in gilthead seabream (Sparus aurata). Fish fed diets enriched with 500 mg kg-1 of AX for 15 days decreased in skin mucus peroxidase activity while at 30 days of trial, fish fed a diet supplemented with 20 mg kg-1 AX increased the peroxidase activity in serum. In addition, bactericidal activity against Vibrio harveyi increased in the skin mucus of fish fed any of the AX supplemented diets. Regarding antioxidant activities in the liver, catalase and glutathione reductase were decreased and increased, respectively, in fish fed a diet supplemented with 500 mg kg-1 of AX. Finally, although the expression of up to 21 inflammatory and lipid metabolism-related genes was analysed in visceral adipose tissue, only the expression of the interleukin 6 (il6) gene was up-regulated in fish fed a diet supplemented with 20 mg kg-1 of AX. The present results provide a detailed insight into the potent antioxidant properties of AX and its possible modulatory effects on the immune status and lipid metabolism of seabream, which may be of interest to the aquaculture sector.

Proteomic analysis in the brain and liver of sea bream (Sparus aurata) exposed to the antibiotics ciprofloxacin, sulfadiazine, and trimethoprim.

Antibiotics, frequently detected in aquatic ecosystems, can negatively impact the health of resident organisms. Although the study on the possible effects of antibiotics on these organisms has been increasing, there is still little information available on the molecular effects on exposed non-target organisms. In our study we used a label free proteomic approach and sea bream, Sparus aurata, to evaluate the effects of exposure to environmentally relevant concentrations of the antibiotic compounds ciprofloxacin (CIP), sulfadiazine (SULF) and trimethoprim (TRIM) produced at the protein level. Individuals of sea bream were exposed to single compounds at 5.2 ± 2.1 µg L-1 of CIP, 3.8 ± 2.7 µg L-1 of SULF and 25.7 ± 10.8 µg L-1 of TRIM for 21 days. After exposure, the number of differentially expressed proteins in the liver was 39, 73 and 4 for CIP, SULF and TRIM respectively. In the brain, there was no alteration of proteins after CIP and TRIM treatment, while 9 proteins were impacted after SULF treatment. The differentially expressed proteins were involved in cellular biological, metabolic, developmental, growth and biological regulatory processes. Overall, our study evidences the vulnerability of Sparus aurata, after exposure to environmentally relevant concentrations of the major antibiotics CIP, SULF and TRIM and that their chronic exposure could lead to a stress situation, altering the proteomic profile of key organs such as brain and liver.

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.

Dietary Artemisia arborescens Supplementation Effects on Growth, Oxidative Status, and Immunity of Gilthead Seabream (Sparus aurata L.).

Fish infectious diseases are one of the main constraints of the aquaculture sector. The use of medicinal plants provides a sustainable way of protection using safe, eco-friendly compounds in a more cost-effective way of treatment, compared to antibiotics. The aim of the present study is the assessment of Artemisia arborescens (AA) feed-supplementation effects on gilthead seabream (Sparus aurata). Fish with an average initial body weight of 109.43 ± 3.81 g, were divided into two groups based on AA feed composition (A25 and A50). Following two months of ad libitum feeding, the effect of diets on fish weight and length were measured. Fish serum and mucus were analyzed for non-specific immune parameters (nitric oxide, lysozyme, myeloperoxidase, protease-/anti-protease activity, and complement), antibody responses, oxidative stress (cytochrome P450 1A1, metallothionein), and metabolism markers (total protein, alkaline phosphatase, and glucose). Expression levels of antioxidants (sod1, gpx1), cytokines (il-1b, il-10, tfgb1, and tnfa), hepcidin, and heat shock protein grp75 genes were measured in spleen samples. A results analysis indicated that A. arborescens use as a feed supplement has a compromised positive effect on the growth performance, immune response, and blood parameters of gilthead seabream. Overall, the suitability of A. arborescens as an efficient food supplement for gilthead seabream health improvement was investigated, setting the basis for its application assessment in Mediterranean aquaculture.

Mucosal affairs: glycosylation and expression changes of gill goblet cells and mucins in a fish-polyopisthocotylidan interaction.

Introduction: Secreted mucins are highly O-glycosylated glycoproteins produced by goblet cells in mucosal epithelia. They constitute the protective viscous gel layer overlying the epithelia and are involved in pathogen recognition, adhesion and expulsion. The gill polyopisthocotylidan ectoparasite Sparicotyle chrysophrii, feeds on gilthead seabream (Sparus aurata) blood eliciting severe anemia. Methods: Control unexposed and recipient (R) gill samples of gilthead seabream experimentally infected with S. chrysophrii were obtained at six consecutive times (0, 11, 20, 32, 41, and 61 days post-exposure (dpe)). In histological samples, goblet cell numbers and their intensity of lectin labelling was registered. Expression of nine mucin genes (muc2, muc2a, muc2b, muc5a/c, muc4, muc13, muc18, muc19, imuc) and three regulatory factors involved in goblet cell differentiation (hes1, elf3, agr2) was studied by qPCR. In addition, differential expression of glycosyltransferases and glycosidases was analyzed in silico from previously obtained RNAseq datasets of S. chrysophrii-infected gilthead seabream gills with two different infection intensities. Results and Discussion: Increased goblet cell differentiation (up-regulated elf3 and agr2) leading to neutral goblet cell hyperplasia on gill lamellae of R fish gills was found from 32 dpe on, when adult parasite stages were first detected. At this time point, acute increased expression of both secreted (muc2a, muc2b, muc5a/c) and membrane-bound mucins (imuc, muc4, muc18) occurred in R gills. Mucins did not acidify during the course of infection, but their glycosylation pattern varied towards more complex glycoconjugates with sialylated, fucosylated and branched structures, according to lectin labelling and the shift of glycosyltransferase expression patterns. Gilthead seabream gill mucosal response against S. chrysophrii involved neutral mucus hypersecretion, which could contribute to worm expulsion and facilitate gas exchange to counterbalance parasite-induced hypoxia. Stress induced by the sparicotylosis condition seems to lead to changes in glycosylation characteristic of more structurally complex mucins.

Identification and florfenicol-treatment of pseudomonas putida infection in gilthead seabream (Sparus aurata) fed on tilapia-trash-feed.

The present study aimed to determine the major cause of the high mortality affecting farmed gilthead seabream (Sparus aurata) and controlling this disease condition. Fifteen diseased S. aurata were sampled from a private fish farm located at Eldeba Triangle, Damietta, fish showed external skin hemorrhages, and ulceration. Bacterial isolates retrieved from the diseased fish were identified biochemically as Pseudomonas putida and then confirmed by phylogenetic analysis of the 16 S rRNA gene sequence. P. putida was also isolated from three batches of tilapia-trash feed given to S. aurata. Biofilm and hemolytic assay indicated that all P. putida isolates produced biofilm, but 61.11% can haemolyse red blood cells. Based on the antibiotic susceptibility test results, P. putida was sensitive to florfenicol with minimum inhibitory concentrations ranging between 0.25 and 1.0 µg mL- 1, but all isolates were resistant to ampicillin and sulfamethoxazole-trimethoprim. Pathogenicity test revealed that P. putida isolate (recovered from the tilapia-trash feed) was virulent for S. aurata with LD50 equal to 4.67 × 107 colony forming unit (CFU) fish- 1. After intraperitoneal (IP) challenge, fish treated with 10 mg kg- 1 of florfenicol showed 16.7% mortality, while no mortality was recorded for the fish group that received 20 mg kg- 1. The non-treated fish group showed 46.7% mortality after bacterial challenge. HPLC analysis of serum florfenicol levels reached 1.07 and 2.52 µg mL- 1 at the 5th -day post-drug administration in the fish groups received 10 and 20 mg kg- 1, respectively. In conclusion, P. putida was responsible for the high mortality affecting cultured S. aurata, in-feed administration of florfenicol (20 mg kg- 1) effectively protected the challenged fish.

In vitro immune-depression and anti-inflammatory activities of cantharidin on gilthead seabream (Sparus aurata) leucocytes activated by λ-carrageenan.

Cantharidin is a natural compound with known therapeutic applications in humans. The aim of this study was to investigate the in vitro effects of cantharidin on gilthead seabream (Sparus aurata) head kidney leucocytes (HKL) stimulated with λ-carrageenan. HKLs were incubated for 24 h with cantharidin (0, 2.5 and 5 µg mL-1) and λ-carrageenan (0 and 1000 µg mL-1). The results showed that HKL viability only decreased by 15.2% after incubated with 5 µg mL-1 of cantharidin and λ-carrageenan. Cantharidin increased the peroxidase activity of HKLs only when incubated in combination with λ-carrageenan. Besides this, cantharidin inhibited the respiratory burst and phagocytic activities. Furthermore, cantharidin induced morphological changes in HKLs (apoptotic and vacuolization signs) that were enhanced when incubated with λ-carrageenan. Considering the analysis of the selected gene expression studied in HKLs [NF-κB subunits (rela, relb, crel, nfkb1, nfkb2), proinflammatory cytokines (il1b, tnfa), anti-inflammatory cytokines (il10, tgfb) and caspases (casp1, casp3, casp8, casp9)], although λ-carrageenan up-regulated the expression of the proinflammatory gene il1b, λ-carrageenan and cantharidin down-regulated its expression in HKLs. In addition, cantharidin up-regulated casp3 and casp9 expression. The casp3 and casp9 gene expression was down-regulated while casp1 gene expression was up-regulated in HKLs incubated with both cantharidin and λ-carrageenan. All the effects of cantharidin are related to its inhibitory effect on protein phosphatases, which induce apoptosis at long exposure times, and minimize the effects of λ-carrageenan. The present results provide detailed insight into the immune-depressive and anti-inflammatory properties of cantharidin on immune cells, which could be of interest to the aquaculture sector.

Participation of Hepcidins in the Inflammatory Response Triggered by λ-Carrageenin in Gilthead Seabream (Sparus aurata).

The role of hepcidins, antimicrobial peptides involved in iron metabolism, immunity, and inflammation, is studied. First, gilthead seabream (Sparus aurata L.) head-kidney leucocytes (HKLs) were incubated with λ-carrageenin to study the expression of hepcidin and iron metabolism-related genes. While the expression of most of the genes studied was upregulated, the expression of ferroportin gene (slc40a) was downregulated. In the second part of the study, seabream specimens were injected intramuscularly with λ-carrageenin or buffer (control). The expression of the same genes was evaluated in the head kidney, liver, and skin at different time points after injection. The expression of Hamp1m, ferritin b, and ferroportin genes (hamp1, fthb, and slc40a) was upregulated in the head kidney of fish from the λ-carrageenin-injected group, while the expression of Hamp2C and Hamp2E genes (hamp2.3 and hamp2.7) was downregulated. In the liver, the expression of hamp1, ferritin a (ftha), slc40a, Hamp2J, and Hamp2D (hamp2.5/6) genes was downregulated in the λ-carrageenin-injected group. In the skin, the expression of hamp1 and (Hamp2A Hamp2C) hamp2.1/3/4 genes was upregulated in the λ-carrageenin-injected group. A bioinformatic analysis was performed to predict the presence of transcription factor binding sites in the promoter region of hepcidins. The primary sequence of hepcidin was conserved among the different mature peptides, although changes in specific amino acid residues were identified. These changes affected the charge, hydrophobicity, and probability of hepcidins being antimicrobial peptides. This study sheds light on the poorly understood roles of hepcidins in fish. The results provide insight into the regulatory mechanisms of inflammation in fish and could contribute to the development of new strategies for treat inflammation in farm animals.

Black soldier fly larvae meal as a potential modulator of immune, inflammatory, and antioxidant status in gilthead seabream juveniles.

This study aimed to evaluate the effects of fish meal (FM) replacement with defatted Hermetia illucens larvae meal (HM) on the hematological profile, immune parameters, intestinal inflammatory status, and antioxidant response in gilthead seabream juveniles. Four diets were formulated, replacing FM with HM at 0%, 22%, 60%, and 100% levels, corresponding to an inclusion level of 15 (diet HM15), 30 (diet HM30), and 45% (diet HM45), respectively. Over 67 days, fish were fed these diets until apparent visual satiation. Results showed no significant differences in immune parameters or hematological profiles, except for a decrease in hemoglobin and hematocrit levels. In the liver, glucose-6-phosphate dehydrogenase and glutathione peroxidase decreased linearly with HM content, especially at 100% replacement. Glutathione reductase activity was also reduced with HM inclusion, being lower in fish fed diet HM30 compared to the control. Fish fed diet HM15 showed lower hepatic superoxide dismutase activity, while catalase activity and lipid peroxidation remained unaffected. In the intestine, antioxidant enzyme activity was not influenced by HM, but lipid peroxidation linearly decreased with HM inclusion, being lower in the HM30 diet compared to the control. The inclusion of HM reduced the expression of intestinal pro-inflammatory genes (interleukin-1ß and cyclooxygenase-2) while the expression of transforming growth factor ß was higher in fish fed diet HM30 compared to the control and HM45 diets. In conclusion, up to 45% dietary inclusion of HM showed no adverse effects, improving liver antioxidant status, reducing intestinal oxidative stress, and regulating inflammatory gene expression.

MassArray Genotyping as a Selection Tool for Extending the Shelf-Life of Fresh Gilthead Sea Bream and European Seabass.

In modern aquaculture, genomics-driven breeding programs have emerged as powerful tools for optimizing fish quality. This study focused on two emblematic Mediterranean fish species, the European seabass (Dicentrarchus labrax) and the gilthead sea bream (Sparus aurata), with a primary aim of exploring the genetic basis of white muscle/fillet degradation in fresh fish following harvest. We identified 57 and 44 missense SNPs in gilthead sea bream and European seabass, respectively, located within genes encoding for endogenous proteases responsible for fillet quality. These SNPs were cherry-picked based on their strategic location within the catalytic/regulatory domains of endogenous proteases that are expressed in the white muscle. Using MassArray technology, we successfully associated differentiated enzymatic activity of those endogenous proteases post-harvest as a phenotypic trait with genetic polymorphism of six SNPs in gilthead sea bream and nine in European seabass. These findings can be valuable attributes in selective breeding programs toward the extension of freshness and shelf life of these species. The integration of MassArray technology into breeding programs offers a cost-effective strategy for harnessing the potential of these genetic variants to enhance the overall quality of the final product. Recognizing that fresh fish perishability is a challenge, extending shelf-life is pivotal in reducing losses and production costs.

Oxidative stress, gene expression and histopathology of cultured gilthead sea bream (Sparus aurata) naturally co-infected with Ergasilus sieboldi and Vibrio alginolyticus.

BACKGROUND: Parasitic and bacterial co-infections have been associated with increasing fish mortalities and severe economic losses in aquaculture through the past three decades. The aim of this study was to evaluate the oxidative stress, histopathology, and immune gene expression profile of gilthead sea bream (Sparus aurata) co-infected with Ergasilus sieboldi and Vibrio alginolyticus. RESULTS: Vibrio alginolyticus and Ergasilus sieboldi were identified using 16 S rRNA and 28 S rRNA sequencing, respectively. The collagenase virulence gene was found in all Vibrio alginolyticus isolates, and the multiple antimicrobial resistance index ranged from 0.286 to 0.857. Oxidant-antioxidant parameters in the gills, skin, and muscles of naturally infected fish revealed increased lipid peroxidation levels and a decrease in catalase and glutathione antioxidant activities. Moreover, naturally co-infected gilthead sea bream exhibited substantial up-regulation of il-1ß, tnf-α, and cyp1a1. Ergasilus sieboldi encircled gill lamellae with its second antennae, exhibited severe gill architectural deformation with extensive eosinophilic granular cell infiltration. Vibrio alginolyticus infection caused skin and muscle necrosis in gilthead sea bream. CONCLUSION: This study described some details about the gill, skin and muscle tissue defense mechanisms of gilthead sea bream against Ergasilus sieboldi and Vibrio alginolyticus co-infections. The prevalence of co-infections was 100%, and no resistant fish were detected. These co-infections imbalance the health status of the fish by hampering the oxidant-antioxidant mechanisms and proinflammatory/inflammatory immune genes to a more detrimental side. Our results suggest that simultaneous screening for bacterial and parasitic pathogens should be considered.

Chitosan-Based Sustained Expression of Sterol Regulatory Element-Binding Protein 1a Stimulates Hepatic Glucose Oxidation and Growth in Sparus aurata.

The administration of a single dose of chitosan nanoparticles driving the expression of sterol regulatory element-binding protein 1a (SREBP1a) was recently associated with the enhanced conversion of carbohydrates into lipids. To address the effects of the long-lasting expression of SREBP1a on the growth and liver intermediary metabolism of carnivorous fish, chitosan-tripolyphosphate (TPP) nanoparticles complexed with a plasmid expressing the N terminal active domain of hamster SREBP1a (pSG5-SREBP1a) were injected intraperitoneally every 4 weeks (three doses in total) to gilthead sea bream (Sparus aurata) fed high-protein-low-carbohydrate and low-protein-high-carbohydrate diets. Following 70 days of treatment, chitosan-TPP-pSG5-SREBP1a nanoparticles led to the sustained upregulation of SREBP1a in the liver of S. aurata. Independently of the diet, SREBP1a overexpression significantly increased their weight gain, specific growth rate, and protein efficiency ratio but decreased their feed conversion ratio. In agreement with an improved conversion of dietary carbohydrates into lipids, SREBP1a expression increased serum triglycerides and cholesterol as well as hepatic glucose oxidation via glycolysis and the pentose phosphate pathway, while not affecting gluconeogenesis and transamination. Our findings support that the periodical administration of chitosan-TPP-DNA nanoparticles to overexpress SREBP1a in the liver enhanced the growth performance of S. aurata through a mechanism that enabled protein sparing by enhancing dietary carbohydrate metabolisation.

Short-term swimming up-regulates pro-inflammatory mediators and cytokines in gilthead seabream (Sparus aurata).

Aerobic swimming exercise in fish has been shown to improve robustness of some species. However, the optimal conditions to be applied and the mechanisms underlying remain unknown. We investigated the effects of 6 h of induced swimming on the immune response of gilthead seabream (Sparus aurata), by analysing markers related to immune status in plasma, skin mucus, gills, heart and head-kidney. Forty fish were individually exercised in swim tunnels by applying different water currents: steady low (SL, 0.8 body lengths (BL) s-1), steady high (SH, 2.3 BL s-1), oscillating low (OL, 0.2/0.8 BL s-1) and oscillating high (OH, 0.8/2.3 BL s-1) velocities, including a non-exercised group with minimal water flow (MF, <0.1 BL s-1). Swimming conditions did not trigger a stress response or anaerobic metabolism, suggested by similar levels of cortisol, lactate, and glucose in plasma among groups. Blood haemoglobin and innate immune parameters in plasma and skin mucus also remained unaltered. However, decreased blood haematocrit was observed in fish swimming on the OL condition. Interestingly, gene expression analysis revealed that the OL condition led to the up-regulation of pro-inflammatory mediators (nfκb1 and mapk3) and cytokines (tnfα, il1ß and il6) in gills. A similar response occurred in heart, with an up-regulation of nfκb1, tnfα, il6 and cox2 in the OL condition. Gene expression of these cytokines was unaltered in the head-kidney. The inflammatory response in gills and heart of gilthead seabream triggered by the OL condition highlights the importance of establishing suitable rearing conditions to improve welfare of cultured fish.

Disruption of the skin, gill, and gut mucosae microbiome of gilthead seabream fingerlings after bacterial infection and antibiotic treatment.

The activity of the microbiome of fish mucosae provides functions related to immune response, digestion, or metabolism. Several biotic and abiotic factors help maintaining microbial homeostasis, with disruptions leading to dysbiosis. Diseases and antibiotic administration are known to cause dysbiosis in farmed fish. Pathogen infections greatly affect the production of gilthead seabream, and antibiotic treatment is still frequently required. Here, we employed a 16S rRNA high-throughput metataxonomics approach to characterize changes in the gut, skin, and gill microbiomes occurring due to infection with Photobacterium damselae subsp. piscicida and subsequent antibiotic treatment with oxytetracycline (OTC), as well as during recovery. Although microbiota response differed between studied tissues, overall changes in composition, diversity, structure, and predicted function were observed in all mucosae. The skin and gill microbiomes of diseased fish became largely dominated by taxa that have been frequently linked to secondary infections, whereas in the gut the genus Vibrio, known to include pathogenic bacteria, increased with OTC treatment. The study highlights the negative impacts of disease and antibiotic treatment on the microbiome of farmed fish. Our results also suggest that fish transportation operations may have profound effects on the fish microbiome, but further studies are needed to accurately evaluate their impact.

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.

Proximate Composition and Fatty Acid Profile of Gilthead Seabream (Sparus aurata) Fed with Pelvetia canaliculata-Supplemented Diets: An Insight towards the Valorization of Seaweed Biomass.

Seaweeds are a sustainable source of protein and lipids that may be used to replace fish by-products in aquaculture feed. This study aimed at using the macroalgae Pelvetia canaliculata as an ingredient in gilthead seabream (Sparus aurata) feed, either as freeze-dried powder or as algae residue (waste) that was obtained after the supplementation of sunflower oil. The formulated diets and the fish muscle were analyzed concerning the proximate composition and the fatty acid profile. The health lipid indices hypocholesterolemic/hypercholesterolemic (h/H), atherogenic (AI), thrombogenic (TI), as well as n-3/n-6 and polyunsaturated fatty acid/saturated fatty acid (PUFA/SFA) ratios were calculated. Additionally, the peroxidizability index (PI) was determined. No differences were observed in the proximate composition of fish muscle regardless of the diet used. Fish fed a diet supplemented with 10% of algae waste (W10) stand out for the highest content in oleic acid (C18:1 n-9), and the lowest in both linoleic (C18:2 n-6) and palmitic (C16:0) fatty acids. All fish samples showed values of health lipid indices within the limits recommend for a nutritional balanced diet. These results highlight that fish fed diets supplemented with P. canaliculata are sources of healthy lipids that might be consumed on a regular basis to prevent cardiovascular diseases.

Long-Term Effects of a Short Juvenile Feeding Period with Diets Enriched with the Microalgae Nannochloropsis gaditana on the Subsequent Body and Muscle Growth of Gilthead Seabream, Sparus aurata L.

Currently, microalgae are used in fish diets, but their long-term growth effect is unknown. In this experiment, juvenile seabream specimens were fed with microalgae-enriched diets for three months, and then transferred to a microalgae-free diet for 10 months to assess long-term effects up to commercial size (≈27 cm and ≈300 g). The juvenile diets contained Nannochloropsis gaditana at 2.5 or 5% inclusion levels, either raw (R2.5 and R5 groups) or cellulose-hydrolyzed (H2.5 and H5 groups). The body length and weight were measured in 75 fish group-1 at commercial stage. The size, number, and fibrillar density of white muscle fibers and the white muscle transverse area were measured in nine fish group-1 at commercial stage. The results showed the highest body weight in H5 at commercial stage. The white muscle transverse area and the white fibres hyperplasia and density also showed the highest values in H5, followed by H2.5. In contrast, the highest hypertrophy was observed in C and R2.5, being associated with the lowest muscle growth in both groups. These results showed a microalgae concentration-dependent effect in hydrolyzed diets as well as an advantageous effect of the hydrolyzed versus raw diets on the long-term growth of Sparus aurata.