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Aquaculture routine practices may cause stress induction on the fish and compromise their welfare affecting the production. This experiment aimed to evaluate the potential links between handling during culture with stress responses and growth on Senegalese sole (Solea senegalensis). We worked with two fish cohorts in terms of initial body weight and culture stage: Trial 1 included specimens in the fattening stage (226 ± 4.96 g) and Trial 2 animals in the pre-fattening stage (27.20 ± 0.44 g). The tested culture protocol, which lasted 6 and 4 months for Trial 1 and 2, respectively, mainly reduced handling-derived stressors in the experimental tanks via lowering routine samplings to a minimum. This decrease of the handling-derived stress was reflected in both trials with lower concentration of circulating cortisol in blood plasma from the experimental fish when compared to controls. Moreover, the proposed protocol promoted higher growth in the fish cultured in the less disturbing protocol in Trial 2. Higher specific growth rates and mean body weight and length were reported. In order to further explore the potential beneficial effects of our protocol, we studied the musculoskeletal from Trial 2 gene expression of key genes regulating glucocorticoid signaling pathway and apoptosis: glucocorticoid receptors 1 and 2 (gr1, gr2), heat shock protein 90 AA (hsp90aa), and caspase 6 (casp6). In line with the cortisol reduced level in this trial, gr1, hsp90aa, and casp6 genes showed lower expression in the samples coming from the experimental group. The findings of this study provide valuable information to the aquaculture industry for the management of Solea senegalensis stress and welfare. [ABSTRACT FROM AUTHOR]

The extensive growth of intensive fish farming has led to a massive spread of infectious diseases. Nervous necrosis virus (NNV) is the causative agent of the viral encephalo‐ and retinopathy disease which has become a major threat for fish farming all over the globe. The devastating mortality rates recorded in disease outbreaks, especially when infected specimens are at early stages of development, have a high economic impact on the sector. Currently, vaccines are the most cost‐effective preventing tool in the fight against viruses. Inactivated vaccines have the advantage of simplicity in their development at the same time as present the antigen in a similar manner than the natural infection in the host. Nevertheless, they usually trigger weaker immune responses needing adjuvants to boost their effectiveness. In this work, we have intraperitoneally vaccinated Senegalese sole juveniles (Solea senegalensis) with a previously designed inactivated vaccine against NNV based on binary ethylenimine (BEI), mixed or not with an oil‐adjuvant. Our results demonstrated the potential activation of different immune pathways when the vaccine was administered alone compared to the oil‐adjuvanted vaccine, both resulting in an equivalent partial improvement in survival following a NNV challenge. However, whilst the vaccine alone led to a significant increase in specific antibodies, in the adjuvanted version those antibodies were kept basal although with a slight improvement in their neutralization capacity. At transcriptional level, neither vaccine (adjuvanted or not) triggered the immune system activation during the vaccination period. However, after NNV infection, the BEI‐inactivated vaccines alone and oil‐adjuvanted both elicited the stimulation of antiviral responsive genes (rtp3, herc4), antigen presentation molecules (mhcii) and T‐cell markers (cd8a) in the head‐kidney. Additionally, the oil‐adjuvanted vaccine appears to stimulate mediator cytokines (il6) and B‐cell markers (ight and ighm). Surprisingly, when the adjuvant was administered alone, fish showed the highest survival rates concomitantly with a lack of NNV‐IgM production, pointing to the possible induction of different immune pathways than the B‐cell responses via antibodies by the adjuvant. Since this combined vaccine did not succeed in the full extension of protection against the pathogen, further studies should be performed focusing on unravelling the molecular mechanisms through which adjuvants trigger the immune response, both independently and when added to a vaccine antigen. [ABSTRACT FROM AUTHOR]

Senegalese sole, Solea senegalensis, is a flatfish of high commercial value in the world. It has been identified as an interesting and promising species for marine commercial aquaculture diversification in Europe for at least four decades and was introduced to China in 2003. Early ontogenesis from embryo to juvenile stages in S. senegalensis was analysed under controlled laboratory conditions to provide morphological information for aquaculture. From 0 to 59 days post hatching (dph), 10–20 larvae were sampled and measured each day (0–17 dph) or every 2–6 days (17–59 dph). Morphological characteristics from the egg to the juvenile stage were described. The eggs were separate and spherical with multiple oil globules. After 3 dph, the yolk sac was completely absorbed, mouth and anus were open, a swim bladder appeared, and larvae began feeding on rotifers (Brachionus plicatilis). The larvae began metamorphosis as the notochord flexed upward and the left eye migrated upward after 10 dph. The left eye migrated to the dorsal midline at 15 dph. At 19 dph, the left eye was translocated to the right‐ocular side, and the juveniles adopted a benthic lifestyle. The swim bladder degenerated, and the juveniles completed metamorphosis at 23 dph. The growth patterns of some parameters (TL, SL, BH, BW) during larval and juvenile development stages were identified. The inflection points, which are slopes of growth changes, were calculated in growth curves. Three inflection points occurring in the growth curves of larvae and juveniles were found to be associated with metamorphosis, weaning, and transitions in feeding habits. The basic information of embryo development and ontogenesis in this study represents a valuable contribution to the S. senegalensis industry, especially in artificial breeding and rearing techniques. [ABSTRACT FROM AUTHOR]

Pleuronectiformes are flatfishes with high commercial value and a prominent example of successful marine adaptation through chromosomal evolution. Hence, the aim of this study was to analyze the 14 relative abundance of repetitive elements (satellite DNA and transposable elements (TE)) in the 15 genome of 10 fish species (8 flatfish) delving into the study of the species of special relevance, 16 Senegalese sole, Solea senegalensis. The results showed differences in the abundance of repetitive elements, with S. senegalensis exhibiting the highest frequency and coverage of these elements reaching the 40% of the genome and not at random distribution. It is noteworthy the presence of relevant peaks of Helitrons in centromeric/pericentromeric positions mainly in the bi-armed chromosomes 1, 2, 4, 6, 7, and 9. The position of the centromeres of this species determined through the genomic localization of the family of satellite DNA PvuII, and other repetitive sequences was obtained de novo. This allowed us to know the genomic position of the centromeres in 19 out of the 21 chromosomes of S. senegalensis. Helitrons showed an accumulation of tandem copies mainly in the pericentromeric positions of chromosomes 1 and 2, occupying a region, in the first case, of 600Kb of tandem repeats. That has only been previously described inmammals andplants.Divergence and copy number studies indicated the presence of active families in the species' genome and the existence of two important events of transposon activity (burst) in the genome of S. senegalensis, mainly accentuated in Helitrons. The results showed that only the families of DNA transposons exhibited a landscape with symmetrical bell-shaped distribution. The phylogenetic analysis of Helitron families revealed the presence of two large groups of families and the presence of four groups of sequences with heterogeneous distribution among chromosomes. Finally, the phylogenomic analysis of 8615 sequences belonging to Helitron insertions from 5 families of flatfish and two external species, allowed to classify the copies into nine groups of sequences with different levels of divergence and clusters, including some branches with distant phylogenetically species. The implications of this study will help to expand the knowledge of chromosome structure and evolution of these species. [ABSTRACT FROM AUTHOR]

Biotic samples from Algeciras Bay (South-west Europe) were studied to assess its ecological status, complementing the previous abiotic monitoring of trace metals in water and sediments. This bay is a densely populated area with intense port traffic and is highly industrialised with metal inputs. To study the impact of this, Zn, Cd, Pb, and Cu contents were determined in tissues of benthic (Solea senegalensis) and benthopelagic species (Scorpaena porcus, Trigloporus lastoviza, and Diplodus sargus sargus). Notable levels of Zn and Cu were found in the liver and gills of all fish species. Compared to international muscle guidelines, Pb sometimes exceeded the most restrictive values, outstanding S. porcus with 27% of samples above the permissible value. Metal pollution indexes revealed that the liver and gills of benthic species were more affected by metal pollution than benthopelagic species, especially in most industrialised sites. Particularly, S. senegalensis presented a higher accumulation factor from sediment of Cd and Cu in the liver (30.1 and 345.1), probably due to the close interaction as benthic species. Among the species studied, S. senegalensis and D. sargus sargus proved to be the best representative and useful bioindicators of metal-polluted environments as this bay. The results were consistent with the findings from the abiotic samples. [ABSTRACT FROM AUTHOR]

Simple Summary: Yellow mealworm is considered as one of the most promising protein sources for replacing fish meal in aquafeeds, among other things because is rich in protein, a good source of micronutrients and exhibits low carbon footprint and land usage. The aim of this study was to evaluate the effects of substituting partial plant or marine-derived ingredients with full-fat yellow mealworm meal at two different levels on the growth performance and fatty acid profiles of Senegalese sole. For this purpose, the study tested a control diet and four experimental diets. Two of the experimental diets substituted marine-derived ingredients for insect meal at two levels (5 and 10%), while the other two substituted components of vegetable origin for insect meal (10 and 15%). The addition of insect meal resulted in an increase in growth rate in both cases, whether replacing fish or plant meals. This led to a decrease in muscle total lipid while maintaining the relative levels of n-3 PUFA and DHA, ultimately improving the lipid health indices n-3: n-6. Tenebrio molitor (TM) is considered as one of the most promising protein sources for replacing fish meal in aquafeeds, among other things because it is rich in protein, a good source of micronutrients and has a low carbon footprint and land use. However, the main drawback of TM is its fatty acid profile, in particular its low content of n-3 PUFA. This study evaluates the effects of partially replacing plant or marine-derived with full-fat TM meal at two different levels on growth performance and lipid profiles of Senegalese sole (Solea senegalensis). For this purpose, a control diet (CTRL) and four experimental isoproteic (53%) and isolipidic (16%) diets were formulated containing 5 and 10% TM meal replacing mostly fish meal (FM5 and FM10), or 10 and 15% TM meal replacing mostly plant meal (PP10 and PP15). Fish (215 g) were fed at 1% of their body weight for 98 days. The final body weight of fish fed the experimental diets containing TM meal was not different from that of fish fed the CTRL diet (289 g). However, the inclusion of TM meal resulted in a gradual improvement in growth rate and feed efficiency in both cases (replacement of fish or plant meals), and significant differences in specific growth rate (SGR) were observed between fish fed the CTRL diet (SGR = 0.30% day−1) and those fed diets with the highest TM meal content (PP15; SGR = 0.35% day−1). The experimental groups did not show any differences in the protein content of the muscle (19.6% w/w). However, significant differences were observed in the total lipid content of the muscle, with the FM10, PP10, and PP15 groups having the lowest muscle lipid contents (2.2% ww). These fish also showed the lowest neutral lipid content in muscle (6.6% dw), but no differences were observed in the total phospholipid content (2.6% dw). Regarding the fatty acid profile, fish fed FM10, PP10 and PP15 had lower levels of linoleic acid (18:2n-6) and higher levels of oleic acid (18:1n-9) in liver and muscle compared to fish fed CTRL. However, no differences were found between fish fed CTRL and TM-based diets for docosahexaenoic acid (22:6n-3) and total n-3 PUFA in liver and muscle. In conclusion, our study demonstrated that full-fat TM inclusion up to 15% in S. senegalensis diets had no negative effects or even some positive effects on fish survival, growth performance, nutrient utilization and flesh quality. [ABSTRACT FROM AUTHOR]

Alternative prophylactic strategies to limit farm animal infection are needed in order to avoid the use of antibiotics. Anti-bacterial and immunostimulatory properties of bioactive compounds are of great interest in aquaculture. Marine derived polysaccharides, such as chitosan and ulvan, together with nanotechnology, have become the focus of attention in the scientific community due to their wide range of biological properties. In this work, chitosan and ulvan-loaded chitosan nanoparticles (referred as CS-TPP NPs and CS-UL-TPP NPs, respectively), obtained by the ionotropic gelation method, had round shape, and the mean sizes were 137.00 ± 5.44 and 325.50 ± 4.95 nm, respectively. No study about the anti-bacterial activity of both types of NPs against Photobacterium damselae subsp. piscicida, an important fish pathogen, has been reported so far. Furthermore, the potential immunostimulatory effects of CS-UL-TPP NPs after oral administration in fish have not yet been evaluated. The percentage of bacterial inhibition against P. damselae subsp. piscicida was determined through in vitro assays, and it was significantly higher in CS-UL-TPP NPs than in CS-TPP NPs at concentrations below 0.03 mg mL−1. The effects on the immune system of CS-TPP and CS-UL-TPP NPs were evaluated in Solea senegalensis juveniles at 30 days after oral administration. Lysozyme activity as well as gene expression levels of il1b, il6, hamp1, tf and c3 was significantly higher in CS-UL-TPP NP-treated groups than in the controls, and no significant differences were observed in CS-TPP NP-treated groups. Thus, ulvan extracted from the macroalgae Ulva ohnoi could improve anti-bacterial and immunostimulant properties of CS-TPP NPs thereby making them suitable to be used as vaccine adjuvant or as immunostimulant. [ABSTRACT FROM AUTHOR]

The isoflavones genistein and daidzein are flavonoid compounds mainly found in legumes, especially in soybeans and their derived products. These flavonoids can be present in agricultural, domestic and industrial wastewater effluents as a result of anthropogenic activities and may be discharged in the environment. Due to the large growth of the aquaculture sector in recent decades, new and cost‐effective fish feeds are being sought, but there is also a particular need to determine the effects of exposed flavonoids on fish in the aquatic environment, as this is the main route of exposure of organisms to endocrine disruptors. This study evaluated the possible effects of these isoflavones on juveniles of Solea senegalensis and Solea solea. After 48–96 h of exposure, the acetylcholinesterase activity in the sole head tissues was measured, and the cholinesterase activity in juveniles of common sole (S. solea) was determined. Experiments were carried out to determine the optimal pH, investigate the specificity of three substrates (acetylthiocholine, butyrylthiocholine, propionylthiocholine) on cholinesterase activity and determine the kinetic parameters (Vmax and Km). The results obtained showed that neither genistein nor daidzein exposure to S. senegalensis and S. solea inhibited the activity of acetylcholinesterase at the tested concentrations (genistein: 1.25, 2.5, 5, 10 and 20 mg/L; daidzein: 0.625, 1.25, 2.5, 5 and 10 mg/L). Possible effects of isoflavones on juveniles of different species of flatfish were evaluated. After 48–96 h of exposure with genistein and daidzein, the acetylcholinesterase activity in the sole head tissues did not showed significant differences with the control. Likewise, optimal pH, specificity of three substrates (acetylthiocholine, butyrylthiocholine, and propionylthiocholine) on cholinesterase activity and kinetic parameters (Vmax and km) were determined on Solea solea. [ABSTRACT FROM AUTHOR]

Pleuronectiformes are flatfishes with high commercial value and a prominent example of successful marine adaptation through chromosomal evolution. Hence, the aim of this study was to analyze the 14 relative abundance of repetitive elements (satellite DNA and transposable elements (TE)) in the 15 genome of 10 fish species (8 flatfish) delving into the study of the species of special relevance, 16 Senegalese sole, Solea senegalensis. The results showed differences in the abundance of repetitive elements, with S. senegalensis exhibiting the highest frequency and coverage of these elements reaching the 40% of the genome and not at random distribution. It is noteworthy the presence of relevant peaks of Helitrons in centromeric/pericentromeric positions mainly in the bi-armed chromosomes 1, 2, 4, 6, 7, and 9. The position of the centromeres of this species determined through the genomic localization of the family of satellite DNA PvuII, and other repetitive sequences was obtained de novo. This allowed us to know the genomic position of the centromeres in 19 out of the 21 chromosomes of S. senegalensis. Helitrons showed an accumulation of tandem copies mainly in the pericentromeric positions of chromosomes 1 and 2, occupying a region, in the first case, of 600Kb of tandem repeats. That has only been previously described in mammals and plants. Divergence and copy number studies indicated the presence of active families in the species’ genome and the existence of two important events of transposon activity (burst) in the genome of S. senegalensis, mainly accentuated in Helitrons. The results showed that only the families of DNA transposons exhibited a landscape with symmetrical bell-shaped distribution. The phylogenetic analysis of Helitron families revealed the presence of two large groups of families and the presence of four groups of sequences with heterogeneous distribution among chromosomes. Finally, the phylogenomic analysis of 8615 sequences belonging to Helitron insertions from 5 families of flatfish and two external species, allowed to classify the copies into nine groups of sequences with different levels of divergence and clusters, including some branches with distant phylogenetically species. The implications of this study will help to expand the knowledge of chromosome structure and evolution of these species.

Trace elements such as Cu, Fe, Mn and Zn are essential minerals in fish diets, especially important at early larval stages. The chemical speciation of these elements directly influences their uptake efficiency and metabolic utilization. In order to optimize the form of trace elements incorporated into larval feed, two experiments were conducted using two commercial fish species, European seabass (Dicentrarchus labrax) and Senegalese sole (Solea senegalensis), and two chemical forms (inorganic and glycinate chelates). Several fish performance parameters were measured, as well as bone status parameters to assess which form of mineral results in optimal fish biological performance. European seabass and Senegalese sole post-larvae were unresponsive (P > 0.05) to dietary treatments in terms of dry weight (DW), standard length (SL), relative growth rate (RGR) or feed conversion rates (FCR) when fed diets supplemented with chelated over inorganic trace minerals. This study suggests that replacing dietary inorganic mineral supplementation by their organic glycinate-chelated forms brings no beneficial effects on somatic growth and bone development in Senegalese sole and European seabass post-larvae fed high-quality commercial microdiets. Additionally, we show that mineral leaching from diets can be significant, but the use of chelated minerals can potentially mitigate this leaching phenomenon. Therefore, the selection of the dietary mineral form should take into account not only their economic value, but also their biological effect and environmental impact. Data generated in this trial provides new knowledge in trace mineral nutrition of early-stage marine fish. [ABSTRACT FROM AUTHOR]

Recirculating aquaculture systems (RAS) pose unique challenges in microbial community management since they rely on a stable community with key target groups, both in the RAS environment and in the host (in this case, Solea senegalensis). Our goal was to determine how much of the sole microbiome is inherited from the egg stage, and how much is acquired during the remainder of the sole life cycle in an aquaculture production batch, especially regarding potentially probiotic and pathogenic groups. Our work comprises sole tissue samples from 2 days before hatching and up to 146 days after hatching (-2 to 146 DAH), encompassing the egg, larval, weaning, and pre-ongrowing stages. Total DNA was isolated from the different sole tissues, as well as from live feed introduced in the first stages, and 16S rRNA gene was sequenced (V6-V8 region) using the Illumina MiSeq platform. The output was analysed with the DADA2 pipeline, and taxonomic attribution with SILVAngs version 138.1. Using the Bray-Curtis dissimilarity index, both age and life cycle stage appeared to be drivers of bacterial community dissimilarity. To try to distinguish the inherited (present since the egg stage) from the acquired community (detected at later stages), different tissues were analysed at 49, 119 and 146 DAH (gill, intestine, fin and mucus). Only a few genera were inherited, but those that were inherited accompany the sole microbiome throughout the life cycle. Two genera of potentially probiotic bacteria (Bacillus and Enterococcus) were already present in the eggs, while others were acquired later, in particularly, forty days after live feed was introduced. The potentially pathogenic genera Tenacibaculum and Vibrio were inherited from the eggs, while Photobacterium and Mycobacterium seemed to be acquired at 49 and 119 DAH, respectively. Significant co-occurrence was found between Tenacibaculum and both Photobacterium and Vibrio. On the other hand, significantly negative correlations were detected between Vibrio and Streptococcus, Bacillus, Limosilactobacillus and Gardnerella. Our work reinforces the importance of life cycle studies, which can contribute to improve production husbandry strategies. However, we still need more information on this topic as repetition of patterns in different settings is essential to confirm our findings. [ABSTRACT FROM AUTHOR]

Some of the characteristics of probiotics used in aquaculture are the antagonistic activity against pathogens and the activation of the immune response. Moreover, some probiotics with common antigens to pathogens can produce antibodies that react with these pathogens. One of those potential probiotics is Vibrio proteolyticus DCF12.2, a strain with the capacity to produce antibodies in soles (Solea senegalensis) that react with Photobacterium damselae subsp. piscicida and Vibrio harveyi antigens. Therefore, the objective of the work was to determine the capacity of DCF12.2 to activate the immune response in the sole, providing protection against both pathogens. For this purpose, DCF12.2 was administered to cultured soles by intraperitoneal injection, bath, and diet. The DCF12.2 strain activated the gene expression related to the fish immune response and increased the antibody production with cross-reaction to P. damselae subsp. piscicida and V. harveyi cells. Challenges with P. damselae subsp. piscicida resulted in a 30–40% relative percent survival (RPS) through DCF12.2 treatments. After challenge with V. harveyi, only intraperitoneal treatment protected the fish (67% RPS). In conclusion, the DCF12.2 strain showed great potential to be used to prevent diseases caused by both pathogens, and indicates a way to use probiotics as live vaccines. [ABSTRACT FROM AUTHOR]

Simple Summary: In contemporary dietary formulations for aquaculture, the amounts of fishmeal are being constantly reduced, and the inclusion of plant-protein sources is now a standard procedure. This approach may result in an unbalanced supply of selected nutrients, from which taurine was identified. Taurine is an amino acid that plays important physiological roles and is included in bile salts, which are essential for the emulsion, digestion, and absorption of dietary lipids and fat-soluble vitamins. In mammals, a hypolipidemic effect of taurine has been described. Senegalese sole (Solea senegalensis) is a marine fish species with increasing importance for aquaculture production in Southern European countries, with a high market value and low tolerance to dietary lipid levels. Thus, this study tested the effects of taurine supplementation to low-fishmeal diets on the physiological responses of Senegalese sole, with an emphasis on lipid metabolism. The results show that dietary inclusion of high levels of plant-protein sources to replace marine ingredients resulted in negative effects on lipid metabolism, due to the resultant low bile-acid concentration and/or the limited availability of taurine for bile-salt emulsification. Taurine supplementation mitigated part of the negative effects of plant-based diets, leading to better lipid utilisation. Taurine is a sulphur-containing amino acid with important physiological roles and a key compound for the synthesis of bile salts, which are essential for the emulsion and absorption of dietary lipids. This study aimed to evaluate the effects of taurine supplementation to low-fishmeal diets on the metabolism of taurine, bile acids, and lipids of Senegalese sole. A fishmeal (FM) and a plant-protein-based (PP0) diet were formulated, and the latter was supplemented with taurine at 0.5 and 1.5% (diets PP0.5 and PP1.5). Diets were assigned to triplicate tanks containing 35 fish (initial weight ~14 g) for 6 weeks. Fish from the PP0 treatment presented lower taurine and bile-acid concentrations compared with the FM treatment, and a downregulation of cyp7a1 and abcb11 was observed. Triolein catabolism decreased in PP0-fed fish, resulting in increased hepatic fat content and plasma triglycerides, while no effects on plasma cholesterol were observed. Taurine supplementation to plant-based diets resulted in a higher taurine accumulation in fish tissues, increased bile-acid concentration, and upregulation of cyp7a1 and abcb11. Hepatic fat content and plasma triglycerides decreased with increasing dietary taurine supplementation. Taurine supplementation mitigated part of the negative effects of plant-based diets, leading to better lipid utilisation. [ABSTRACT FROM AUTHOR]

Sex determination (SD) shows huge variation among fish and a high evolutionary rate, as illustrated by the Pleuronectiformes (flatfishes). This order is characterized by its adaptation to demersal life, compact genomes and diversity of SD mechanisms. Here, we assembled the Solea senegalensis genome, a flatfish of great commercial value, into 82 contigs (614 Mb) combining long‐ and short‐read sequencing, which were next scaffolded using a highly dense genetic map (28,838 markers, 21 linkage groups), representing 98.9% of the assembly. Further, we established the correspondence between the assembly and the 21 chromosomes by using BAC‐FISH. Whole genome resequencing of six males and six females enabled the identification of 41 single nucleotide polymorphism variants in the follicle stimulating hormone receptor (fshr) consistent with an XX/XY SD system. The observed sex association was validated in a broader independent sample, providing a novel molecular sexing tool. The fshr gene displayed differential expression between male and female gonads from 86 days post‐fertilization, when the gonad is still an undifferentiated primordium, concomitant with the activation of amh and cyp19a1a, testis and ovary marker genes, respectively, in males and females. The Y‐linked fshr allele, which included 24 nonsynonymous variants and showed a highly divergent 3D protein structure, was overexpressed in males compared to the X‐linked allele at all stages of gonadal differentiation. We hypothesize a mechanism hampering the action of the follicle stimulating hormone driving the undifferentiated gonad toward testis. [ABSTRACT FROM AUTHOR]

Simple Summary: Adding physical enrichment to the rearing conditions may promote fish welfare and reduce detrimental characteristics that fish develop in captivity. Senegalese sole (Solea senegalensis), an important species for European aquaculture, is reared in intensive conditions using fibreglass tanks. This species shows reproductive dysfunctions that do not allow it to complete its life cycle in production. Recently, dominance behaviour has been studied to try to solve this problem. The present study aimed to assess the effect of sand as an environmental enrichment in the dominance behaviour and brain gene expression of Senegalese sole juveniles. Several behaviours associated with feeding and territoriality were analysed by video recordings. In both environments, dominant soles were the first to feed, displayed a higher number of contact interactions and dominated the area close to the feeding point, where the events were reduced in fish maintained in the sand. The nr4a2 and fshra genes related to differentiation of dopamine neurons and regulation of maturation were significantly up-regulated in dominant fish maintained with sand compared to dominants maintained without sand. Using an enriched environment may affect Senegalese sole dominance, enhance welfare and advance future maturation. Physical complexity adds physical enrichment to rearing conditions. This enrichment promotes fish welfare and reduces detrimental characteristics that fish develop in captivity. Senegalese sole (Solea senegalensis) is an important species for European aquaculture, where it is reared in intensive conditions using fibreglass tanks. However, reproductive dysfunctions present in this species do not allow it to complete its life cycle in captivity. Recently, dominance behaviour has been studied to try to solve this problem. The present study aimed to assess the effect of sand as environmental enrichment in the dominance behaviour and brain mRNA abundance of Senegalese sole juveniles. Four tanks of sole (n = 48 fish in total) were established in two different environments (with and without sand). Juveniles were subjected to dominance tests of feeding and territoriality. Behaviours analysed by video recordings related to the distance from the food delivered and harassment behaviour towards other individuals (e.g., resting of the head on another individual). In both environments, dominant sole were the first to feed, displayed more head-resting behaviour and dominated the area close to the feeding point, where the events were reduced in fish maintained in the sand. mRNA expression related to differentiation of dopamine neurons (nr4a2) and regulation of maturation (fshra) were significantly upregulated in dominant fish in the sand environment compared to dominants maintained without sand. The use of an enriched environment may affect Senegalese sole dominance, enhance welfare and possibly advance future maturation. [ABSTRACT FROM AUTHOR]

Simple Summary: It is almost 50 years since amoeba-associated granulomatosis was first described in goldfish (Carassius auratus), but the aetiologic agent has never been identified. In this study, a new archamoeba species, Endolimax carassius, is characterised as the causative agent of systemic granulomatous disease in goldfish. Phylogenetic analyses determined the new goldfish parasite species as a genotype sister of Endolimax piscium, the causative agent of systemic granulomatous disease in Senegalese sole. The fish genotypes clustered with other available genotypes from mammals in a well-supported Endolimax clade within the Mastigamoebidae. Endolimax is a genus of intestinal amoebae which stands among the least known human protists. Previous studies on amoebic systemic granulomatosis of a marine fish (Solea senegalensis) resulted in the unexpected characterization of a new organism which was related to Endolimax and named E. piscium. The existence of multiple reports of systemic granulomatosis caused presumptively by unidentified amoebae in goldfish lead us to investigate the organism involved in goldfish disease. Analysed goldfish presented small whitish nodules in the kidney, which correspond to chronic granulomatous inflammatory reactions with a ring-layer of amoebae in the periphery. Amoebae were amitochondriate and were located in a parasitophorous vacuole within macrophages, as previous studies on this condition in goldfish and other freshwater fish pointed out. SSU rDNA characterization confirmed a new Endolimax lineage which appears closely related to E. piscium, but the molecular evidence, distinct pathological features and lack of ecological overlapping between the hosts support their assignment to a new species, E. carassius. The results support the existence of a considerable unexplored diversity of Endolimax spp. among fish, and their proper characterization can contribute to an understanding of Archamoebae evolution and pathogenic potential. [ABSTRACT FROM AUTHOR]

Tenebrio molitor (TM) is considered as one of the most promising protein sources for replacing fish meal in aquafeeds, among other things because it is rich in protein, a good source of micronutrients and has a low carbon footprint and land use. However, the main drawback of TM is its fatty acid profile, in particular its low content of n-3 PUFA. This study evaluates the effects of partially replacing plant or marine-derived with full-fat TM meal at two different levels on growth performance and lipid profiles of Senegalese sole (Solea senegalensis). For this purpose, a control diet (CTRL) and four experimental isoproteic (53%) and isolipidic (16%) diets were formulated containing 5 and 10% TM meal replacing mostly fish meal (FM5 and FM10), or 10 and 15% TM meal replacing mostly plant meal (PP10 and PP15). Fish (215 g) were fed at 1% of their body weight for 98 days. The final body weight of fish fed the experimental diets containing TM meal was not different from that of fish fed the CTRL diet (289 g). However, the inclusion of TM meal resulted in a gradual improvement in growth rate and feed efficiency in both cases (replacement of fish or plant meals), and significant differences in specific growth rate (SGR) were observed between fish fed the CTRL diet (SGR = 0.30% day−1) and those fed diets with the highest TM meal content (PP15; SGR = 0.35% day−1). The experimental groups did not show any differences in the protein content of the muscle (19.6% w/w). However, significant differences were observed in the total lipid content of the muscle, with the FM10, PP10, and PP15 groups having the lowest muscle lipid contents (2.2% ww). These fish also showed the lowest neutral lipid content in muscle (6.6% dw), but no differences were observed in the total phospholipid content (2.6% dw). Regarding the fatty acid profile, fish fed FM10, PP10 and PP15 had lower levels of linoleic acid (18:2n-6) and higher levels of oleic acid (18:1n-9) in liver and muscle compared to fish fed CTRL. However, no differences were found between fish fed CTRL and TM-based diets for docosahexaenoic acid (22:6n-3) and total n-3 PUFA in liver and muscle. In conclusion, our study demonstrated that full-fat TM inclusion up to 15% in S. senegalensis diets had no negative effects or even some positive effects on fish survival, growth performance, nutrient utilization and flesh quality.

Histological techniques are a traditional and essential tool for adequately comprehending cellular morphology and physiology in several scientific fields, including ecotoxicological studies. Although bioassays generally recommend using small test organisms, based on advantages such as low maintenance costs and the possibility of a higher number of replicates, the histological process of such organisms is still challenging due to the difficulty of handling them. The present study proposes an accessible protocol for performing the histology of a biological sample of 8 mm maximum length and using histopathological biomarkers in Senegal sole larvae for ecotoxicological tests. Eight-day old larvae of Solea senegalensis obtained from breeders' natural spawns in cultivation conditions were submitted to histological processing by an adjusted protocol comprising fixation, agarose array pre-embedding, dehydration, paraffin embedding, slides' preparation, and image analysis by light optical microscope coupled to a digital camera. Low-cost and easy-to-use alternatives were applied in the technique execution. The histological steps were detailed, and S. senegalensis morphological features were presented. This can be considered valuable background information for ecotoxicological studies where changes in cell morphology and physiology are expected. [ABSTRACT FROM AUTHOR]

Microplastics (MP) are contaminants able to cause adverse effects on organisms. MPs are capable to interact with other environmental contaminants, including pesticides, altering their toxicity. The objective of the study was to research the sublethal effects (enzymatic activity) of pesticides alone and in combination with MPs. Cholinesterase enzymes are used as biomarkers to determine and evaluate the effects produced in organisms after exposure to pollutants. This study showed the acetylcholinesterase (AChE) enzymatic activity in the tissue of Solea senegalensis exposed to two environmental pollutants, the insecticide chlorpyrifos (CPF) and antibacterial triclosan (TCS) with and without microplastics (MPs). Solea senegalensis was chosen because it is a species in high demand because of its high economic value in southern Europe, as well as the use of this species in ecotoxicology and its increasing use as sentinel species, which justify using it to assess biological effects of pollutants. Toxicity tests were performed in organisms exposed to concentrations of between 5 and 80 μg/L CPF and 0.1 and 0.4 mg/L TCS for 96 h. In addition, each test incorporated MPs that were added at different concentrations in order to evaluate their role as a possible enhancer of the effects caused by the previous pollutants. In the case of CPF, the head and muscle tissue cholinesterase activity was inhibited from a concentration of 5 μg/L both without and with MPs, and the AChE enzymatic activity for the treatment with MPs was approximately half the activity for the treatment without MPs in the tissues studied. Besides, TCS inhibited the cholinesterase activity at a concentration of 0.3 mg/L in the muscle of S. senegalensis. In contrast, no significant differences were observed in the TCS + MP treatment compared to the controls. These results showed the importance of studies in assessing the anticholinesterase effects of pesticides combined with microplastics due to the abundance of these contaminants in the marine environment and the role of cholinesterase activity (biomarker) in the neurotransmission of key physiological processes. [ABSTRACT FROM AUTHOR]

The regression of fisheries in the Gulf of Cadiz is evident since current fish catches are 33% of that 30 years before. Consequently, some initiatives for the replenishment of exhausted wild stocks are welcome. The objective of the present work is to describe and analyse the results coming from the first flatfish stock enhancements in Andalusia. A total of 3189 fish from three flatfish species: Senegal sole (Solea senegalensis Kaup), wedge sole (Dicologlossa cuneata Moreau), and brill (Scophthalmus rhombus Linnaeus) were tagged and released. Several variables were calculated through the data analysis of recovered fish. Some variables were calculated only for Senegal soles since wedge sole and brill recaptures were not significant. The Senegal sole recapture rate was 2.71 ± 0.72, similar to other published data, the recapture rates for bigger fish being higher though not significant. No significant differences were detected for distance, time, growth, or recapture rate amongst initial Senegal sole sizes. Around 80% of recaptures were registered within 15 weeks after release. The results show that it is possible for the release and recovery of tagged Senegal soles in the Gulf of Cadiz. Future long-term programmes on stock enhancement could help to determine the effects on fisheries and recover stocks. [ABSTRACT FROM AUTHOR]

Simple Summary: The Senegalese sole (Solea senegalensis, Kaup 1858), a marine flatfish species, is of commercial interest for both fisheries and aquaculture. In aquaculture, currently there are several production bottlenecks, mainly in larvae culture and rearing. The hox genes participate in cell differentiation and structuring of the anterior–posterior axis during embryonic development. In this work, using cytogenetic and genomic techniques, hox genes were isolated and sequenced to study the sequence and cluster organization, together with the cytogenetic localization. Results were analyzed from an evolutionary perspective by comparing with other species. Our findings should represent a useful starting point for further research in order to gain more knowledge of the development of Senegalese sole for planning functional analyses focused on the main concerns that characterize the development phase. The Senegalese sole (Solea senegalensis, Kaup 1858), a marine flatfish, belongs to the Pleuronectiformes order. It is a commercially important species for fisheries and aquaculture. However, in aquaculture, several production bottlenecks have still to be resolved, including skeletal deformities and high mortality during the larval and juvenile phase. The study aims to characterize the hox gene clusters in S. senegalensis to understand better the developmental and metamorphosis process in this species. Using a BAC library, the clones that contain hox genes were isolated, sequenced by NGS and used as BAC-FISH probes. Subsequently the hox clusters were studied by sequence analysis, comparative genomics, and cytogenetic and phylogenetic analysis. Cytogenetic analysis demonstrated the localization of four BAC clones on chromosome pairs 4, 12, 13, and 16 of the Senegalese sole cytogenomic map. Comparative and phylogenetic analysis showed a highly conserved organization in each cluster and different phylogenetic clustering in each hox cluster. Analysis of structural and repetitive sequences revealed accumulations of polymorphisms mediated by repetitive elements in the hoxba cluster, mainly retroelements. Therefore, a possible loss of the hoxb7a gene can be established in the Pleuronectiformes lineage. This work allows the organization and regulation of hox clusters to be understood, and is a good base for further studies of expression patterns. [ABSTRACT FROM AUTHOR]

Simple Summary: Vitamin K (VK), and particularly phylloquinone (VK1), is an essential micronutrient whose stability in aquafeeds has not been extensively evaluated. Losing stability can lead to nutritional deficiency, which is known to hamper fish development and physiology. Gut microbiota also plays a key role in host health through the interaction with several biological processes. The present study evaluated the best storing conditions of aquafeeds when supplemented in VK1 and intestinal microbiota modulation in Senegalese sole (Solea senegalensis) juveniles. Aquafeeds with a high level of VK1 supplementation required storage at −20 °C for short-term (up to 7 days) and at −80 °C for long-term (up to three months) to ensure optimal preservation. Furthermore, gut bacterial communities of Senegalese sole specimens fed with a commercial feed supplemented with VK1 showed a better-balanced population of microorganisms in the intestine, which might improve Senegalese sole health during the ongrowing phase. These results provide the practical guidelines for the proper storing of aquafeeds in the industry when supplemented with VK1 and highlight the potential benefits of dietary VK1 supplementation for a balanced intestinal microbiota and overall fish health. Vitamin K (VK) is a fat-soluble vitamin necessary for fish metabolism and health. VK stability as dietary component during aquafeed storage and its potential effect on intestinal microbiome in fish have not yet been completely elucidated. The convenient storage conditions of aquafeeds when supplemented with phylloquinone (VK1), as well as its potential effects on the gut microbiota of Senegalese sole (Solea senegalensis) juveniles, have been explored. Experimental feeds were formulated to contain 0, 250 and 1250 mg kg−1 of VK1 and were stored at different temperatures (4, −20 or −80 °C). VK stability was superior at −20 °C for short-term (7 days) storage, while storing at −80 °C was best suited for long-term storage (up to 3 months). A comparison of bacterial communities from Senegalese sole fed diets containing 0 or 1250 mg kg−1 of VK1 showed that VK1 supplementation decreased the abundance of the Vibrio, Pseudoalteromonas, and Rhodobacterace families. All these microorganisms were previously associated with poor health status in aquatic organisms. These results contribute not only to a greater understanding of the physiological effects of vitamin K, particularly through fish intestinal microbiome, but also establish practical guidelines in the industry for proper aquafeed storage when supplemented with VK1. [ABSTRACT FROM AUTHOR]

Within the efforts in the search for novel feed ingredients in aquaculture, macroalgae are being widely explored due to their diversity of bioactive compounds with considerable prophylactic and therapeutic potential. Plocamium cartilagineum and Sargassum vulgare , have the potential to positively impact fish health, mainly due to their antioxidant properties. The production of Senegalese sole (Solea senegalensis) still faces some constraints due to the low sperm quantity and quality of males from captivity, which is highly affected by oxidative stress. We hypothesized that the scavenger ability of P. cartilagineum and S. vulgare could ameliorate the reproductive performance of Senegalese sole males, once incorporated in the broodstock feeds. In this experiment, broodstock groups were fed with three different formulated experimental diets for 60 days: Control (Ctrl), a diet supplemented with 5% P. cartilagineum (Pc), and a diet supplemented with 5% S. vulgare (Sv). Fish were sampled twice to evaluate sperm quality using different techniques: sperm motility by Computer Assisted Sperm Analysis (CASA) software, and cell viability and Reactive Oxygen Species (ROS) by flow cytometer. At the final sampling, 6 fish per treatment were sacrificed and gonads were extracted to determine the relative expression of anti- and pro-apoptotic genes (bcl2b, bcl-x, mcl1b, badb, bax, boka), and genes involved in fish antioxidant system (sod3, cat, gpx, hsp70). Sex steroids were measured on blood plasma by ELISA. Specific growth rate (SGR) and feed efficiency were also evaluated at the end of the experiment. Senegalese sole breeders fed with Pc diet for 60 days showed enhanced sperm total motility and cell viability at the end of the experiment, when compared with the Ctrl diet. On the other hand, fish fed with Sv diet had lower spermatozoa intracellular ROS and upregulated gpx in the gonads. No differences were obtained in levels of sex steroids and gonadal apoptotic system after 60 days of feeding, however, the macroalgae species and the percentages of incorporation used proved to positively modulate different spermatozoa traits. • Macroalgae-supplemented diets modulate different Senegalese sole sperm traits. • 5% Plocamium cartilagineum meal improved spermatozoa total motility and viability. • 5% Sargassum vulgare meal improved gonadal and sperm antioxidant system. • The macroalgae antioxidant proprieties can mitigate fish sperm oxidative processes. [ABSTRACT FROM AUTHOR]

Simple Summary: The ability to obtain in vivo information on characteristics related to fish composition is necessary for aquaculture. In addition, there is growing interest in production traits, such as growth, feed efficiency or fillet weight, but it remains difficult to precisely record in vivo individual fish traits that report to these production traits, which can increase edible fish meat production and decrease the environmental impact. In the present study, we performed an ultrasound approach for the in vivo prediction of fillet volume of the Senegalese sole (Solea senegalensis), a species considered a promising flatfish species for marine fish farming. The results show that models based on ultrasound fillet volume measurements explain above 95% of the variation observed in fillet volume. However, for fillet yield estimation, the results were modest. Therefore, further studies are necessary to better understand the potential of the ultrasound approach to this trait. Nevertheless, this work allows us to conclude that the approach with ultrasound is promising for measuring in vivo fish composition traits. Senegalese sole (Solea senegalensis) has been considered a promising new flatfish species for Mediterranean marine fish farming. Accurate prediction of fillet traits in live animals may allow for more efficient control of muscle deposition in fish. In this sense, this study was undertaken to develop a non-invasive method to predict in vivo fish fillet volume and yield using real-time ultrasonography (RTU). The trial was conducted with 44 market weight Senegalese sole (298.54 ± 87.30 g). Fish were scanned with an Aloka SSD 500V with a 7.5 MHz probe. Ten RTU cross-sectional images were taken from the operculum to the caudal fin at regular intervals. These images were analyzed using Fiji software. These data were then used to estimate the partial volumes of the fillet. Actual fillet volume was determined using Archimedes' principle. Simple and stepwise multiple regression analyses were then used to develop prediction models of fillet volume and yield. The most cranial RTU sections of the fish fillet were the best single predictors of both fillet volume and fillet yield and were the ones included in the best stepwise models. The best RTU slice area explained 82% of the variation observed in fillet volume, but the other RTU slice areas used as predictors of fillet volume showed poor to moderate accuracy (0.035 ≤ R2 ≤ 0.615). Single RTU partial volumes showed poor to very high accuracy (0.395 ≤ R2 ≤ 0.970) as predictors of fillet volume. The best stepwise model based on the RTU slice areas included three independent variables and explained 88.3% of the observed variation. The best stepwise models based on RTU partial volumes (single volumes and/or combinations of single volumes) explained about 97% of the variation observed in fillet volume. Two RTU volume traits, V1–5 + V6–9, and V1+()+9, showed to be practically direct predictors of the actual fillet volume, explaining, respectively, 97% and 96% of the variation observed in the actual fillet volume. The fillet yields show lower correlations with slice areas (r between 0.044 and 0.601) than with volumes (r between 0.288 and 0.637). While further studies are clearly necessary to better understand the potential of RTU for the estimation of fillet yield in fish in general and Senegalese sole in particular, the present results showed that RTU traits can be very good predictors of Senegalese sole's fillet volume, either used in regression models or as direct predictors. [ABSTRACT FROM AUTHOR]

Melatonin is a powerful antioxidant present in fish seminal plasma. This study aimed to understand melatonin's endogenous and exogenous effects on first-generation Senegalese sole sperm quality for sperm management applications. In the first experiment, samples were collected at mid-light (ML) and mid-dark (MD) daytimes, to evaluate the effects on sperm motility. In a second experiment, using confocal microscopy and melatonin-FITC, spermatozoa permeability to melatonin was evaluated and, after showing that it enters the nucleus and mitochondria by passive diffusion, exogenous melatonin toxicity and antioxidant potential during a cryopreservation assay were performed. The toxicity assay tested different melatonin concentrations (0.01, 0.1, 1, and 10 mM) and exposure times (3, 5, 15 and 30 min), and sperm motility parameters were measured (TM, PM, VCL, VSL, LIN) using CASA system. The best conditions (0.1 and 10 mM) were selected for the cryopreservation assay, and a set of post-thaw sperm quality analyses were performed (motility, viability, reactive oxygen species, lipid peroxidation, and DNA fragmentation). The motility analyzed at ML and MD showed significant differences in all parameters, mainly on velocities (VCL, VSL, VAP), that were significantly higher at MD. Supplemented melatonin did not influence spermatozoa motility, MDA content or DNA fragmentation, although a lower percentage of viable cells was obtained on the 10 mM treatment. Altogether, Senegalese sole spermatozoa motility was enhanced at night, putatively by endogenous melatonin through direct or indirect mechanisms, whereas supplemented melatonin did not confer extra protection during cryopreservation., Competing Interests: Declaration of competing interest The authors have nothing to declare., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

• Reared breeders of Solea senegalensis show plasticity in observational learning. • Housing conditions from the juvenile stage contribute to reproductive behaviour success. • Social learning from experienced congeners offers opportunities for courtship success. • A lack of consistency in reproductive behaviour and spawning occurs over time. • A system of stimulus reinforcement may be necessary for reactivation of the behaviour. The reproductive failure of Senegalese sole (Solea senegalensis) cultured males (reared entirely in captivity from egg through to adult) that do not participate in reproductive behaviours to fertilise spawns, results in a problem to achieve reproductive control in captivity. However, cohabitation with wild males has led to an increase in the involvement of cultured males in reproductive behaviour, although their contribution to fertilised spawning is still lower than that of wild breeders. This study aimed to examine the effect of different social conditions, on the reproductive behaviour and spawning success of cultured breeders over three reproductive seasons. Before starting this study, different social learning opportunities were provided to the breeders from the juvenile to the pubertal stages of the individuals. Behaviour and spawning were evaluated in four experimental groups of cultured breeders: two groups (W1 and W2) that prior to this study were reared during the juvenile stage with wild breeders that fertilized spawns, a Culture breeder group (CB) that was previously reared with cultured breeders that spawned unfertile eggs, and a negative control group (CN) that was reared in isolation from adult fish. During the three reproductive seasons, spawning was obtained from all groups. Generally, the first year had the highest egg production and the third year the lowest. However, fertilised eggs were only obtained from W1 in the first year. A total of eight fertilised spawns were collected with a fertilisation rate of 28.02 ± 13.80 % and a hatching rate of 15.04 ± 10.40 %. The mean number of larvae obtained per spawn was 7,683 ± 5,947 and the total number of larvae from all eight spawns was 61,468. The paternity analysis assigned 64.3 % of larvae to a single couple of breeders, while 34.3 % of larvae were not assigned to any single family, but inconclusively to more than three parents. The highest locomotor activity was observed in W1, while no significant differences were observed in the number of movements within W2, CB and CN. In all groups, during the peak of locomotor activity (19 h 00-20 h 00), the main reproductive behaviours observed were Rest the Head and Follow , while the Guardian behaviour was low and Coupled behaviour was only observed in W1. Over time, the reproductive behaviours decreased, except for Follow. The social learning opportunities provided by cohabitation with wild fish during juvenile stages prior to spawning in W1, increased activity and fertilised spawning. However, the number of successful spawns was low and over time stopped in association with a decrease in reproductive behaviour. This suggests that other mechanisms of behavioural learning could be involved in reproductive success, such as reproductive dominance, environmental conditions or hormonal interactions that could affect physiological processes in the reproduction of captive breeders. [ABSTRACT FROM AUTHOR]

The maturation of the intestinal digestive and absorptive functions might limit the amount of absorbed nutrients to fulfil the high requirements of the fast-growing marine fish larva. Glutamine (Gln) has been described to improve intestinal epithelium functions, due to its involvement in energy metabolism and protein synthesis. The purpose of this study was to evaluate dietary 0.2% Gln supplementation on aspects of intestinal physiology, protein metabolism and growth-related genes expression in Senegalese sole larvae. Experiment was carried out between 12 and 33 days post hatching (DPH) and fish were divided into two experimental groups, one fed Artemia spp. (CTRL) and the other fed Artemia spp. supplemented with Gln (GLN). GLN diet had two times more Gln than the CTRL diet. Samples were collected at 15, 19, 26 and 33 DPH for biometry, histology, and digestive enzymes activity, and at 33 DPH for gene expression, protein metabolism and AA content determination. Growth was significantly higher for Senegalese sole fed GLN diet, supported by differences on protein metabolism and growth-related gene expression. Slight differences were observed between treatments regarding the intestinal physiology. Overall, GLN diet seems to be directed to enhance protein metabolism leading to higher larval growth. [Display omitted] • Dietary glutamine supplementation benefits Senegalese sole larval growth. • Glutamine enriched live feed diet enhanced protein metabolism of sole larvae. • Growth-related genes expression of sole larvae was higher for fish fed glutamine-supplemented diet. • Glutamine supplementation could be an early nutritional strategy to boost Solea senegalensis production in aquaculture. [ABSTRACT FROM AUTHOR]

Taurine is a sulphur-containing amino acid with important physiological roles and a key compound for the synthesis of bile salts, which are essential for the emulsion and absorption of dietary lipids. This study aimed to evaluate the effects of taurine supplementation to low-fishmeal diets on the metabolism of taurine, bile acids, and lipids of Senegalese sole. A fishmeal (FM) and a plant-protein-based (PP0) diet were formulated, and the latter was supplemented with taurine at 0.5 and 1.5% (diets PP0.5 and PP1.5). Diets were assigned to triplicate tanks containing 35 fish (initial weight ~14 g) for 6 weeks. Fish from the PP0 treatment presented lower taurine and bile-acid concentrations compared with the FM treatment, and a downregulation of cyp7a1 and abcb11 was observed. Triolein catabolism decreased in PP0-fed fish, resulting in increased hepatic fat content and plasma triglycerides, while no effects on plasma cholesterol were observed. Taurine supplementation to plant-based diets resulted in a higher taurine accumulation in fish tissues, increased bile-acid concentration, and upregulation of cyp7a1 and abcb11. Hepatic fat content and plasma triglycerides decreased with increasing dietary taurine supplementation. Taurine supplementation mitigated part of the negative effects of plant-based diets, leading to better lipid utilisation.

The Pleuronectiformes order, which includes several commercially-important species, has undergone extensive chromosome evolution. One of these species is Solea senegalensis, a flatfish with 2n = 42 chromosomes. In this study, a cytogenomics approach and integration with previous maps was applied to characterize the karyotype of the species. Synteny analysis of S. senegalensis was carried out using two flatfish as a reference: Cynoglossus semilaevis and Scophthalmus maximus. Most S. senegalensis chromosomes (or chromosome arms for metacentrics and submetacentrics) showed a one-to-one macrosyntenic pattern with the other two species. In addition, we studied how repetitive sequences could have played a role in the evolution of S. senegalensis bi-armed (3, and 5–9) and acrocentric (11, 12 and 16) chromosomes, which showed the highest rearrangements compared with the reference species. A higher abundance of TEs (Transposable Elements) and other repeated elements was observed adjacent to telomeric regions on chromosomes 3, 7, 9 and 16. However, on chromosome 11, a greater abundance of DNA transposons was detected in interstitial BACs. This chromosome is syntenic with several chromosomes of the other two flatfish species, suggesting rearrangements during its evolution. A similar situation was also found on chromosome 16 (for microsatellites and low complexity sequences), but not for TEs (retroelements and DNA transposons). These differences in the distribution and abundance of repetitive elements in chromosomes that have undergone remodeling processes during the course of evolution also suggest a possible role for simple repeat sequences in rearranged regions. [ABSTRACT FROM AUTHOR]

The inclusion of macroalgae in the diets of farmed fish offers the opportunity for an added-value dietary ingredient to the nutraceutical feed. The composition of algae varies greatly among species. Several Ulva species have been considered in aquafeed formulations for different farmed fish, and Ulva ohnoi is being applied recently. However, the effects of seaweed dietary inclusion on the host must be evaluated. Considering the important role of the host intestinal microbiota, the potential effects of U. ohnoi dietary inclusion need to be studied. In this study, the characterization of the intestinal microbiome of Solea senegalensis , a flatfish with high potential for aquaculture in South Europe, receiving U. ohnoi (5%)-supplemented diet for 90 days has been carried out. In addition, the functional profiles of bacterial communities have been determined by using PICRUSt, a computational approach to predict the functional composition of a metagenome by using marker gene data and a database of reference genomes. The results show that long-term dietary administration of U. ohnoi (5%)-supplemented feed modulates S. senegalensis intestinal microbiota, especially in the posterior intestinal section. Increased relative abundance of Vibrio jointly with decreased Stenotrophomonas genus has been detected in fish receiving Ulva diet compared to control-fed fish. The influence of the diet on the intestinal functionality of S. senegalensis has been studied for the first time. Changes in bacterial composition were accompanied by differences in predicted microbiota functionality. Increased abundance of predicted genes involved in xenobiotic biodegradation and metabolism were observed in the microbiota when U. ohnoi diet was used. On the contrary, predicted percentages of genes associated to penicillin and cephalosporin biosynthesis as well as beta-lactam resistance were reduced after feeding with Ulva diet. [ABSTRACT FROM AUTHOR]

A mechanistic model based on Dynamic Energy Budget (DEB) theory was developed to predict the combined effects of ocean warming, acidification and decreased food availability on growth and reproduction of three commercially important marine fish species: white seabream (Diplodus sargus), zebra seabream (Diplodus cervinus) and Senegalese sole (Solea senegalensis). Model simulations used a parameter set for each species, estimated by the Add-my-Pet method using data from laboratory experiments complemented with bibliographic sources. An acidification stress factor was added as a modifier of the somatic maintenance costs and estimated for each species to quantify the effect of a decrease in pH from 8.0 to 7.4 (white seabream) or 7.7 (zebra seabream and Senegalese sole). The model was used to project total length of individuals along their usual lifespan and number of eggs produced by an adult individual within one year, under different climate change scenarios for the end of the 21st century. For the Intergovernmental Panel on Climate Change SSP5–8.5, ocean warming led to higher growth rates during the first years of development, as well as an increase of 32–34% in egg production, for the three species. Ocean acidification contributed to reduced growth for white seabream and Senegalese sole and a small increase for zebra seabream, as well as a decrease in egg production of 48–52% and 14–33% for white seabream and Senegalese sole, respectively, and an increase of 4–5% for zebra seabream. The combined effect of ocean warming and acidification is strongly dependent on the decrease of food availability, which leads to significant reduction in growth and egg production. This is the first study to assess the combined effects of ocean warming and acidification using DEB models on fish, therefore, further research is needed for a better understanding of these climate change-related effects among different taxonomic groups and species. [ABSTRACT FROM AUTHOR]

Intensive fish farming at high densities results in a wide range of adverse consequences on fish welfare, including pathogen spreading, stress and increased mortality rates. In this work, we have assessed whether the survival of Senegalese sole infected with the nervous necrosis virus (NNV), a pathogen responsible for severe disease outbreaks, is affected by rearing density. Based on the different fish ratios per surface area (g cm−2) and water volume (g L−1), our research showed an earlier mortality onset in the tanks containing NNV‐infected fish reared at medium density (MD: 0.071 g cm−2/5 g L−1) and high density (HD: 0.142 g cm−2/10 g L−1), as well as higher cumulative mortality values. However, transcription analysis of hsp70, gr1 and pepck genes, well‐known stress biomarkers, seems to indicate that none of the challenged fish were under high stress conditions. NNV load was slightly higher both in dead and in sampled fish from MD and HD groups, and especially in the rearing water from these groups, where peaks in mortality seemed to correlate with increasing NNV load in the water. In conclusion, our results suggest that rearing NNV‐infected Senegalese sole at high densities resulted in an earlier mortality onset and higher cumulative values and viral load. [ABSTRACT FROM AUTHOR]

Background Marine invertebrates are provided as a first feed for marine fish larvae because of their strict nutritional requirements, despite also being a potential source of infectious agents. Aim To assess horizontal transmission of a nervous necrosis virus reassortant strain (NNV) to sole larvae via Artemia and rotifers. Materials and methods Rotifer (Brachionus plicatilis) and Artemia (Artemia salina) nauplii cultures were bath infected with a reassortant (RGNNV/SJNNV) NNV strain isolated from gilthead sea bream and viral internalisation was confirmed by IFA. Senegalese sole (Solea senegalensis) larvae were fed on infected Artemia and disease signs and mortality were recorded. In addition, NNV viability was checked in cultures of either unfed invertebrates or invertebrates fed on phytoplankton and in the supernatant of microalgae cultures. All samples were tested by RT-qPCR and inoculation in cell culture. Results Both rotifers and Artemia internalised NNV. Experimental transmission to sole larvae was achieved using infected Artemia and subsequently 60% mortality was recorded. At 24 h post-infection, orally infected individuals contained 9.34 × 104 copies of viral RNA, whereas the bath infection yielded 2.05 × 106 RNA copies larvae−1. Viral presence in both invertebrates was detected up to 8 days post infection but viral load decreased over time. Feeding with microalgae decreased viral detection even more and microalgae supernatants were demonstrated to significantly affect NNV viability. Conclusions Our results demonstrate that both invertebrates can bioaccumulate NNV and that Senegalese sole larvae fed on infected Artemia might develop viral encephalopathy and retinopathy and high mortality.

The inclusion of macroalgae in the diets of farmed fish offers the opportunity for an added-value dietary ingredient to the nutraceutical feed. The composition of algae varies greatly among species. Several Ulva species have been considered in aquafeed formulations for different farmed fish, and Ulva ohnoi is being applied recently. However, the effects of seaweed dietary inclusion on the host must be evaluated. Considering the important role of the host intestinal microbiota, the potential effects of U. ohnoi dietary inclusion need to be studied. In this study, the characterization of the intestinal microbiome of Solea senegalensis, a flatfish with high potential for aquaculture in South Europe, receiving U. ohnoi (5%)-supplemented diet for 90 days has been carried out. In addition, the functional profiles of bacterial communities have been determined by using PICRUSt, a computational approach to predict the functional composition of a metagenome by using marker gene data and a database of reference genomes. The results show that long-term dietary administration of U. ohnoi (5%)-supplemented feed modulates S. senegalensis intestinal microbiota, especially in the posterior intestinal section. Increased relative abundance of Vibrio jointly with decreased Stenotrophomonas genus has been detected in fish receiving Ulva diet compared to control-fed fish. The influence of the diet on the intestinal functionality of S. senegalensis has been studied for the first time. Changes in bacterial composition were accompanied by differences in predicted microbiota functionality. Increased abundance of predicted genes involved in xenobiotic biodegradation and metabolism were observed in the microbiota when U. ohnoi diet was used. On the contrary, predicted percentages of genes associated to penicillin and cephalosporin biosynthesis as well as beta-lactam resistance were reduced after feeding with Ulva diet.

The Senegalese sole (Solea senegalensis, Kaup 1858), a marine flatfish, belongs to the Pleuronectiformes order. It is a commercially important species for fisheries and aquaculture. However, in aquaculture, several production bottlenecks have still to be resolved, including skeletal deformities and high mortality during the larval and juvenile phase. The study aims to characterize the hox gene clusters in S. senegalensis to understand better the developmental and metamorphosis process in this species. Using a BAC library, the clones that contain hox genes were isolated, sequenced by NGS and used as BAC-FISH probes. Subsequently the hox clusters were studied by sequence analysis, comparative genomics, and cytogenetic and phylogenetic analysis. Cytogenetic analysis demonstrated the localization of four BAC clones on chromosome pairs 4, 12, 13, and 16 of the Senegalese sole cytogenomic map. Comparative and phylogenetic analysis showed a highly conserved organization in each cluster and different phylogenetic clustering in each hox cluster. Analysis of structural and repetitive sequences revealed accumulations of polymorphisms mediated by repetitive elements in the hoxba cluster, mainly retroelements. Therefore, a possible loss of the hoxb7a gene can be established in the Pleuronectiformes lineage. This work allows the organization and regulation of hox clusters to be understood, and is a good base for further studies of expression patterns.

Infectious diseases are one of the main causes of social and economical losses in world aquaculture. Senegalese sole (Solea senegalensis) is an important species for aquaculture in southern Europe, whose production is affected by the appearance of bacterial diseases such as photobacteriosis, a septicemia caused by Photobacterium damselae subsp. piscicida (Phdp). The aim of this study was to obtain an oral DNA nanovaccine and to evaluate its efficacy against Phdp in S. senegalensis juveniles. For this purpose, the amplified product corresponding to the protein inosine-5′-monophophate dehydrogenase (IMPDH) from Phdp, was cloned into the expression vector pcDNA™6.2/C-EmGFP-GW obtaining the DNA vaccine named as pPDPimpdh. The correct transcription and protein expression was verified at 48 h post tansfection in HEK293 cells. Chitosan nanoparticles (CS-TPP NPs) were prepared by ionotropic gelation and their features were appropriate for use as oral delivery system. Therefore, pPDPimpdh was protected with chitosan CS-TPP NPs throughout complex coacervation method giving as a result a DNA nanovaccine referred as CS-TPP+pPDPimpdh NPs. Sole juveniles were vaccinated orally with CS-TPP NPs, pPDPimpdh and CS-TPP+pPDPimpdh NPs followed by a challenge with Phdp at 30 days post vaccination (dpv). The relative percentage survival (RPS) for pPDPimpdh vaccinated groups was 6.25%, probably due to its degradation in the digestive tract. RPS value obtained for CS-TPP NPs and CS-TPP+pPDPimpdh NPs was 40% and antibodies were observed in both cases. However, a delay in mortality was observed in sole juveniles vaccinated orally with CS-TPP+pPDPimpdh NPs. In fact, an upregulation of tf , mhcII , cd8a and igm in the posterior gut and c3, hamp1, tf and cd4 in spleen was observed in juveniles vaccinated with CS-TPP+pPDPimpdh NPs. After challenge, a modulation of cd8a and cd4 expression levels in the posterior gut and c3 , tf, lyg , cd4 , igm and igt expression levels in spleen was observed. Moreover, the concentration of lysozyme in skin mucus significantly increased in fish vaccinated orally with CS-TPP+pPDPimpdh NPs at 11 dpc. These data indicate that oral vaccination with CS-TPP+pPDPimpdh NPs could be acting through the non-specific immune responses as well as the specific humoral and cell mediated immunity and provide the first step toward a development of an oral DNA nanovaccine against Phdp in sole. • Oral vaccination of naked pPDPimpdh leads to a weak response and low efficacy in the protection against Phdp in sole. • CS-TPP NPs prepared in this study are suitable for oral vaccination of pPDPimpdh. • CS-TPP NPs and CS-TPP+pPDPimpdh NPs are able to generate antibodies and a moderate protection against Phdp in sole. • CS-TPP+pPDPimpdh NPs could boost non-specific, humoral and cellular immune responses in Senegalese sole juveniles. [ABSTRACT FROM AUTHOR]

Many personal care products integrate UV-filters, such as 4-methylbenzylidene camphor (4-MBC), a compound frequently detected in aquatic habitats, including coastal areas. However, the potential effects of 4-MBC to saltwater species have been poorly studied. Therefore, the main objective of this work is to study the effects of 4-MBC exposure on Solea senegalensis during metamorphosis, a sensitive life stage of this flatfish. To achieve this, fish were exposed to 4-MBC (0.2–2.0 mg L-1) for 48 h at the beginning of metamorphosis (13 days after hatching, dah). After this period, the fish were transferred to a clean medium. They were fed and maintained until more than 80% of individuals in the control group completed the metamorphosis (24 dah). Mortality, malformations, and metamorphic progression were studied daily. Growth, behavior, and biochemical markers of neurotransmission (acetylcholinesterase, AChE), oxidative stress (catalase, CAT; lipid peroxidation, LPO), detoxification (glutathione S-transferase, GST), and anaerobic metabolism (lactate dehydrogenase, LDH) were also determined at the end of the experiment. An acceleration of metamorphosis progression was observed during and 2 days after the 4-MBC exposure in all concentrations tested. In addition, reduced length, inhibition of CAT activity, and induction of oxidative damage were observed (lowest observed effect concentration, LOEC = 0.928 mg L−1 4-MBC for length, CAT, and LPO). Short-term exposure to 4-MBC at the onset of metamorphosis affected S. senegalensis at several levels of organization, even after 9 days in a clean medium, including growth and metamorphic progression, suggesting possible long-term adverse effects in this species. [ABSTRACT FROM AUTHOR]

The toxicity of malathion to Solea senegalensis was studied in a static renewal bioassay during its first month of larval life (between 4 and 30 dph). Through the use of different biomarkers and biochemical, cellular and molecular approaches (inhibition of cholinesterases [ChEs], changes in cytochrome P450‐1A [CYP1A] and the study of histopathological alterations), the effects of three concentrations of malathion (1.56, 3.12, and 6.25 μg/L) have been analyzed. In subacute exposure, malathion inhibited cholinesterase activities (AChE, BChE, CbE) in a dose‐ and time‐dependent manner, ranging the inhibition percentage from 20% to 90%. However, the expression levels of CYP1A and AChE transcripts or proteins were not modified. Additionally, exposure to malathion provoked histopathological alterations in several organ systems of Senegalese sole in a time‐ and dose dependent way, namely disruption of parenchymal architecture in the liver, epithelial desquamation, pyknotic nuclei and steatosis in the intestine, disorganization of supporting cartilage, and sings of hyperplasia and hypertrophy in the gills and degeneration of the epithelial cells from the renal tubules. Malathion exposure also provoked strong disorganization of cardiac fibers from the heart. The findings provide evidence that exposure to sublethal concentrations of malathion that provoked serious injury to the fish S. senegalensis, were below the expected environmental concentrations reported in many other ecosystems and different fish species,revealing a higher sensitivity for Solea senegalensis to malathion exposure, thus reinforcing its use as sentinel species for environmental pollution in coastal and estuarine environments. [ABSTRACT FROM AUTHOR]

Vitamin K (VK) is a fat-soluble vitamin necessary for fish metabolism and health. VK stability as dietary component during aquafeed storage and its potential effect on intestinal microbiome in fish have not yet been completely elucidated. The convenient storage conditions of aquafeeds when supplemented with phylloquinone (VK1), as well as its potential effects on the gut microbiota of Senegalese sole (Solea senegalensis) juveniles, have been explored. Experimental feeds were formulated to contain 0, 250 and 1250 mg kg−1 of VK1 and were stored at different temperatures (4, −20 or −80 °C). VK stability was superior at −20 °C for short-term (7 days) storage, while storing at −80 °C was best suited for long-term storage (up to 3 months). A comparison of bacterial communities from Senegalese sole fed diets containing 0 or 1250 mg kg−1 of VK1 showed that VK1 supplementation decreased the abundance of the Vibrio, Pseudoalteromonas, and Rhodobacterace families. All these microorganisms were previously associated with poor health status in aquatic organisms. These results contribute not only to a greater understanding of the physiological effects of vitamin K, particularly through fish intestinal microbiome, but also establish practical guidelines in the industry for proper aquafeed storage when supplemented with VK1.

Senegalese sole (Solea senegalensis) has been considered a promising new flatfish species for Mediterranean marine fish farming. Accurate prediction of fillet traits in live animals may allow for more efficient control of muscle deposition in fish. In this sense, this study was undertaken to develop a non-invasive method to predict in vivo fish fillet volume and yield using real-time ultrasonography (RTU). The trial was conducted with 44 market weight Senegalese sole (298.54 ± 87.30 g). Fish were scanned with an Aloka SSD 500V with a 7.5 MHz probe. Ten RTU cross-sectional images were taken from the operculum to the caudal fin at regular intervals. These images were analyzed using Fiji software. These data were then used to estimate the partial volumes of the fillet. Actual fillet volume was determined using Archimedes’ principle. Simple and stepwise multiple regression analyses were then used to develop prediction models of fillet volume and yield. The most cranial RTU sections of the fish fillet were the best single predictors of both fillet volume and fillet yield and were the ones included in the best stepwise models. The best RTU slice area explained 82% of the variation observed in fillet volume, but the other RTU slice areas used as predictors of fillet volume showed poor to moderate accuracy (0.035 ≤ R2 ≤ 0.615). Single RTU partial volumes showed poor to very high accuracy (0.395 ≤ R2 ≤ 0.970) as predictors of fillet volume. The best stepwise model based on the RTU slice areas included three independent variables and explained 88.3% of the observed variation. The best stepwise models based on RTU partial volumes (single volumes and/or combinations of single volumes) explained about 97% of the variation observed in fillet volume. Two RTU volume traits, V1–5 + V6–9, and V1+()+9, showed to be practically direct predictors of the actual fillet volume, explaining, respectively, 97% and 96% of the variation observed in the actual fillet volume. The fillet yields show lower correlations with slice areas (r between 0.044 and 0.601) than with volumes (r between 0.288 and 0.637). While further studies are clearly necessary to better understand the potential of RTU for the estimation of fillet yield in fish in general and Senegalese sole in particular, the present results showed that RTU traits can be very good predictors of Senegalese sole’s fillet volume, either used in regression models or as direct predictors.

Understanding the daily digestive function could lead to more efficient feeding practices in the farmed fish species. In this work, the effect of several daily feeding protocols was assessed on the daily expression of the genes related to the circadian system (arntl1, clock, per1, and cry1), appetite (cck), nutrients hydrolysis (try1, ctrb1, pga, atp4a, cel, and amy2a), peptides transport (slc15a1), pH, and activity of key digestive (trypsin, chymotrypsin, amylase, 4C- and 7C-like lipases) and absorptive (alkaline phosphatase) enzymes in the gastrointestinal tract (GIT) of Senegalese sole (Solea senegalensis) juveniles. Some of the tested feeding protocols demonstrated a significant influence on the entrainment of the peripheral oscillator. The synchrony of slc15a1 and some digestive proenzymes with per1 in some of the experimental treatments could underline the role of clock genes in the regulation of these biological functions. Unlike gene expression of the digestive proenzymes wherein feeding protocol occasionally affected the temporal organization, the influence of this factor was determinant on the daily average production and/or circadian pattern of their activities. Daily pH was maintained at approximately neutral levels in all the GIT segments, and no pepsin activity was detected. Fish fed a single diurnal meal showed the lowest activity for all the studied enzymes, except for amylase, suggesting an insufficient utilization of the ingested nutrients. Higher amylase activity in this group, on the other hand, implies the possible activation of alternative compensatory metabolic pathways. Overall, our results pointed to the suitability of higher feeding frequencies, especially during dark hours, for the aquaculture production of this species.

Microalgae are one of the most promising groups of functional ingredients for inclusion in microdiets for Senegalese sole (Solea senegalensis) larvae, supplying components with anti-oxidant effects that may be important during an accelerated stage of development where excessive reactive oxygen substance production leading to oxidative stress may occur. The diatom Phaeodactylum tricornutum complies with such potential since it is rich in fucoxanthin, a carotenoid with demonstrated antioxidant activity. This study aimed at determining the effect of dietary P. tricornutum supplementation on the growth performance, survival and levels of oxidative stress of Senegalese sole postlarvae. Four dietary treatments were tested: a commercial microdiet, used as control, and three experimental variants where P. tricornutum was included (1) whole cells at 5%—WP5, (2) broken cells at 5%—BP5 and (3) broken cells at 10%—BP10. At the end of the trial, no significant differences were found in growth performance of larvae fed diets where P. tricornutum was included at 5% (WP5 and BP5) and the commercial control. However, larvae from the BP10 treatment had a significantly higher dry weight than larvae from the remaining treatments. Moreover, the experimental diets had no significant effects on oxidative stress. Nonetheless, lipid peroxidation levels were significantly higher in smaller larvae fed diets where P. tricornutum whole cells were included, relatively to the commercial control. The results of the present study demonstrate that P. tricornutum broken cell biomass has the potential for inclusion in weaning microdiets for Senegalese sole, increasing larval growth performance at an inclusion level of 10 %. [ABSTRACT FROM AUTHOR]

The implementation of marine protected areas (MPAs) has been widely used as a tool to manage and conserve marine resources and services. Yet, to date, the reserve effect is still weakly evaluated, particularly for soft‐sediment habitats.The Arrábida MPA, considered as a biodiversity hotspot, was fully established in 2009 on the west coast of Portugal and is characterized by large expanses of soft‐sediment habitats. This MPA was established to protect biodiversity and to ensure the livelihood of the local small‐scale fisheries community.Beyond before–after‐control–impact (BACI) analyses were carried out on catch data (abundance and biomass) of 351 trammel net sets from experimental fishing campaigns (2007–2019) to study the reserve effect on demersal fish populations.The results show a declining trend in abundance inside and outside the protected area, with significant positive effects only found for undersize commercial species and Solea senegalensis, and a general increase in fish species total length. Despite the lack of any obvious reserve effect, the increase in fish length can be considered as a first sign.Anthropogenic pressures are generalized around the area, and transgressions of the restrictions are frequently observed inside the no‐take zone. These can, among other aspects, delay or prevent the expected reserve effects. Therefore, enforcement and/or modification of the spatial plan must be incentivized to achieve the goals of the MPA. [ABSTRACT FROM AUTHOR]

Immunonutrition is a promising and viable strategy for the development of prophylactic measures in aquaculture. Ulvan, a sulphated marine polysaccharide from green seaweeds, has many biological activities including the immunomodulatory ones. The aim of this study was to assess the short and long-term effects of an ulvan-rich extract obtained from U. ohnoi as immunonutrient in Senegalese sole juveniles. In this work, an ulvan-rich extract from Ulva ohnoi has been obtained by the hot water method and isolated by ethanol precipitation. The FTIR analysis revealed that the ulvan-rich extact had very similar characteristics to previously published ulvan spectra. The total sulfate and protein content was 24.85 ± 3.98 and 0.91 ± 0.04 %, respectively. In vitro assays performed in Senegalese sole (Solea senegalensis) macrophages showed that the ulvan obtained in this study did not compromise the cell viability at concentrations up to 1 mg ml−1 and expression levels of lyg , irf1 , il6 , il10 , c7 , tf and txn were significantly upregulated in a concentration dependent-manner. Finally, S. senegalensis juveniles were fed basal diets and diets supplemented with the ulvan-rich extract at ratios 1 and 2 % for 30 days and then, challenged with Photobacterium damselae subsp. piscicida (Phdp). Thereafter, ulvan was withdrawn from the diet and all juveniles were fed the basal diet for 30 days. At 30 days post withdrawal (dpw), juveniles were challenged with Phdp. The expression profiles of a set of genes related to the immune system in spleen were evaluated as well as the lysozyme, peroxidase and bactericidal activity in plasma. Dietary effects of 1 % ulvan resulted in a boost of the immune response and increased disease resistance at short-term whereas juveniles fed diets supplemented with 2 % ulvan showed a significant decrease in the bactericidal activity and lack of protection against Phdp. At long-term (30 days after the withdrawal of ulvan), an improved response was observed in juveniles previously fed 1 % ulvan. • The ulvan-rich extract obtained shows suitable features to be use as immunonutrient. • The ulvan-rich extract shows immunomodulatory effects in a concentration dependent-manner. • Dietary administration of 1 % ulvan enhance immune response and disease resistance in Solea senegalensis juveniles. • Dietary administration of 2 % ulvan resulted in lack of protection against Phdp. [ABSTRACT FROM AUTHOR]

Understanding vertebral bone development is essential to prevent skeletal malformations in farmed fish related to genetic and environmental factors. This is an important issue in Solea senegalensis , with special impact of spinal anomalies in postlarval and juvenile stages. Vertebral bone transcriptomics in farmed fish mainly comes from coding genes, and barely on miRNA expression. Here, we used RNA-seq of spinal samples to obtain the first comprehensive coding and miRNA transcriptomic repertoire for postlarval and juvenile vertebral bone, covering different vertebral phenotypes and egg-incubation temperatures related to skeleton health in S. senegalensis. Coding genes, miRNA and pathways regulating bone development and growth were identified. Differential transcriptomic profiles and suggestive mRNA-miRNA interactions were found between postlarvae and juveniles. Bone-related genes and functions were associated with the extracellular matrix, development and regulatory processes, calcium binding, retinol and lipid metabolism or response to stimulus, including those revealed by the miRNA targets related to signaling, cellular and metabolic processes, growth, cell proliferation and biological adhesion. Pathway enrichment associated with fish skeleton were identified when comparing postlarvae and juveniles: growth and bone development functions in postlarvae, while actin cytoskeleton, focal adhesion and proteasome related to bone remodeling in juveniles. The transcriptome data disclosed candidate coding and miRNA gene markers related to bone cell processes, references for functional studies of the anosteocytic bone of S. senegalensis. This study establishes a broad transcriptomic foundation to study healthy and anomalous spines under early thermal conditions across life-stages in S. senegalensis , and for comparative analysis of skeleton homeostasis and pathology in fish and vertebrates. • Novel mRNA- and miRNA-sequencing was obtained from Senegalese sole vertebrae. • Postlarval and juvenile transcriptomes under early thermal conditions were profiled. • A transcriptome baseline was set up for studying spinal anomalies in Senegalese sole. • New skeleton transcriptome data useful for comparative studies in fish and vertebrates. [ABSTRACT FROM AUTHOR]

Essential dietary trace elements, such as zinc (Zn) and manganese (Mn), critically influence a wide range of physiological, metabolic, and hormonal processes in fish larvae and post-larvae. Despite their importance for normal fish growth and skeletal development, trace mineral nutrition has not been extensively studied in the early stages of development of fish. Post-larvae of an emergent aquaculture species, Senegalese sole (Solea senegalensis), were the subject of this study in order to better understand the effects of diet supplementation of trace minerals upon fish larval development and performance. Sole post-larvae were fed a combination of organic Mn (45 and 90 mg kg−1 feed) and organic Zn (100 and 130 mg kg−1 feed) and survival, growth, mineral deposition rates, and vertebral bone status were assessed. Our results showed that although no significant effect was found on the growth performance of Senegalese sole post-larvae, Mn and Zn supplementation to a commercial microdiet for marine fish larvae at higher dietary levels (Mn at 90 mg kg−1 and Zn at 130 mg kg−1) improved larval survival, decreased the severity of vertebral malformations, and increased the deposition of Mn in bone. [ABSTRACT FROM AUTHOR]

Aquaculture is a fundamental sector of the food industry nowadays. However, to become a sustainable and more profitable industry, it is necessary to monitor several associated parameters, such as temperature, salinity, ammonia, potential of hydrogen, nitrogen dioxide, bromine, among others. Their regular and simultaneous monitoring is expected to predict and avoid catastrophes, such as abnormal fish mortality rates. In this paper, we propose a novel anomaly detection approach for the early prediction of high fish mortality based on a multivariate Gaussian probability model. The goal of this approach is to determine the correlation between the number of daily registered physicochemical parameters of the fish tank water and the fish mortality. The proposed machine learning model was fitted with data from the weaning and pre-fattening phases of Senegalese sole (Solea senegalensis) collected over 2018, 2019, and 2020. This approach is suitable for real-time tracking and successful prediction of up to 80% of the high fish mortality rates. To the best of our knowledge, the proposed anomaly detection approach is the first time studied and applied in the framework of the aquaculture industry. [ABSTRACT FROM AUTHOR]

The Senegalese sole (Solea senegalensis) is an economically important flatfish species. In this study, a genome draft was analyzed to identify microsatellite (SSR) markers for whole-genome genotyping. A subset of 224 contigs containing SSRs were preselected and validated by using a de novo female hybrid assembly. Overall, the SSR density in the genome was 886.7 markers per megabase of genomic sequences and the dinucleotide motif was the most abundant (52.4%). In silico comparison identified a set of 108 SSRs (with di-, tetra- or pentanucleotide motifs) widely distributed in the genome and suitable for primer design. A total of 106 markers were structured in thirteen multiplex PCR assays (with up to 10-plex) and the amplification conditions were optimized with a high-quality score. Main genetic diversity statistics and genotyping reliability were assessed. A subset of 40 high polymorphic markers were selected to optimize four supermultiplex PCRs (with up to 11-plex) for pedigree analysis. Theoretical exclusion probabilities and real parentage allocation tests using parent–offspring information confirmed their robustness and effectiveness for parental assignment. These new SSR markers were combined with previously published SSRs (in total 229 makers) to construct a new and improved integrated genetic map containing 21 linkage groups that matched with the expected number of chromosomes. Synteny analysis with respect to C. semilaevis provided new clues on chromosome evolution in flatfish and the formation of metacentric and submetacentric chromosomes in Senegalese sole. [ABSTRACT FROM AUTHOR]

Probiotics can be used to reduce disease outbreaks in aquaculture. Some of them are characterised by their antagonistic activity against pathogens or the stimulation of the fish immune response, including the production of specific antibodies. If a probiotic has common antigens with a determined pathogen, it could produce antibodies with a cross-reaction to that pathogen. Thus, a probiotic with these characteristics could be used in a similar way to a live vaccine. The aim of this study was to select bacteria with antigenic similarity and antagonistic activity against the pathogens Photobacterium damselae subsp. piscicida and Vibrio harveyi, and to determine their ability to stimulate the production of antibodies in sole (Solea senegalensis, Kaup) with cross-reaction against these pathogens.Dot blot was used to detect strains with cross-reaction using sera immunized against P. damselae subsp. piscicida Lg41.01 and V. harveyi Lg16.00. A total of 138 strains were selected from 718 strains, based on the intensity of the dot blot reaction. A second selection was performed to detect their ability to inhibit the pathogens growth. Five strains inhibited the growth of P. damselae subsp. piscicida, four strains inhibited the growth of V. harveyi, while two strains inhibited both pathogens. A Western blot confirmed the cross-reactions of the selected strains with the pathogens.Selected strains were subsequently inoculated into sole specimens by intraperitoneal injection. Four strains produced antibodies with cross-reaction against the pathogens. None mortality was observed in the inoculated fish. Further research demonstrated the storage capability of the selected strains in saline solution and feed, their growth at low pH, and identified their enzymatic characteristics. In conclusion, the selected strains showed antimicrobial activity and capacity to activate a specific immune response against fish pathogens.