Gilthead sea bream (Sparus aurata) is a species of paramount importance to the Mediterranean aquaculture industry, with an annual production exceeding 140,000 metric tons. Pasteurellosis due to the Gram-negative bacterium Photobacterium damselae subsp. piscicida (Phdp) causes significant mortality, especially during larval and juvenile stages, and poses a serious threat to bream production. Selective breeding for improved resistance to pasteurellosis is a promising avenue for disease control, and the use of genetic markers to predict breeding values can improve the accuracy of selection, and allow accurate calculation of estimated breeding values of nonchallenged animals. In the current study, a population of 825 sea bream juveniles, originating from a factorial cross between 67 broodfish (32 sires, 35 dams), were challenged by 30 min immersion with 1 × 105 CFU virulent Phdp. Mortalities and survivors were recorded and sampled for genotyping by sequencing. The restriction-site associated DNA sequencing approach, 2b-RAD, was used to generate genome-wide single nucleotide polymorphism (SNP) genotypes for all samples. A high-density linkage map containing 12,085 SNPs grouped into 24 linkage groups (consistent with the karyotype) was constructed. The heritability of surviving days (censored data) was 0.22 (95% highest density interval: 0.11-0.36) and 0.28 (95% highest density interval: 0.17-0.4) using the pedigree and the genomic relationship matrix respectively. A genome-wide association study did not reveal individual SNPs significantly associated with resistance at a genome-wide significance level. Genomic prediction approaches were tested to investigate the potential of the SNPs obtained by 2b-RAD for estimating breeding values for resistance. The accuracy of the genomic prediction models (r = 0.38-0.46) outperformed the traditional BLUP approach based on pedigree records (r = 0.30). Overall results suggest that major quantitative trait loci affecting resistance to pasteurellosis were not present in this population, but highlight the effectiveness of 2b-RAD genotyping by sequencing for genomic selection in a mass spawning fish species.

Food supply from the ocean is constrained by the shortage of domesticated and selected fish. Development of genomic models of economically important fishes should assist with the removal of this bottleneck. European sea bass Dicentrarchus labrax L. (Moronidae, Perciformes, Teleostei) is one of the most important fishes in European marine aquaculture; growing genomic resources put it on its way to serve as an economic model.

Sexual dimorphism is a fascinating subject in evolutionary biology and mostly results from sex-biased expression of genes, which have been shown to evolve faster in gonochoristic species. We report here genome and sex-specific transcriptome sequencing of Sparus aurata, a sequential hermaphrodite fish. Evolutionary comparative analysis reveals that sex-biased genes in S. aurata are similar in number and function, but evolved following strikingly divergent patterns compared with gonochoristic species, showing overall slower rates because of stronger functional constraints. Fast evolution is observed only for highly ovary-biased genes due to female-specific patterns of selection that are related to the peculiar reproduction mode of S. aurata, first maturing as male, then as female. To our knowledge, these findings represent the first genome-wide analysis on sex-biased loci in a hermaphrodite vertebrate species, demonstrating how having two sexes in the same individual profoundly affects the fate of a large set of evolutionarily relevant genes.

Teleost fish scales play important roles in animal protection and homeostasis. They can be targeted by endogenous estrogens and by environmental estrogenic endocrine disruptors. The phytoestrogen genistein is ubiquitous in the environment and in aquaculture feeds and is a disruptor of estrogenic processes in vertebrates. To test genistein disrupting actions in teleost fish we used a minimally invasive approach by analysing scales plucked from the skin of sea bass (Dicentrarchus labrax). Genistein transactivated all three fish nuclear estrogen receptors and was most potent with the Esr2, had the highest efficacy with Esr1, but reached, in all cases, transactivation levels lower than those of estradiol. RNA-seq revealed 254 responsive genes in the sea bass scales transcriptome with an FDR < 0.05 and more than 2-fold change in expression, 1 or 5 days after acute exposure to estradiol or to genistein. 65 genes were specifically responsive to estradiol and 106 by genistein while 83 genes were responsive to both compounds. Estradiol specifically regulated genes of protein/matrix turnover and genistein affected sterol biosynthesis and regeneration, while innate immune responses were affected by both compounds. This comprehensive study revealed the impact on the fish scale transcriptome of estradiol and genistein, providing a solid background to further develop fish scales as a practical screening tool for endocrine disrupting chemicals in teleosts.

Efforts towards utilisation of diets without fish meal (FM) or fish oil (FO) in finfish aquaculture have been being made for more than two decades. Metabolic responses to substitution of fishery products have been shown to impact growth performance and immune system of fish as well as their subsequent nutritional value, particularly in marine fish species, which exhibit low capacity for biosynthesis of long-chain poly-unsaturated fatty acids (LC-PUFA). The main objective of the present study was to analyse the effects of a plant-based diet on the hepatic transcriptome of European sea bass (Dicentrarchus labrax).

Fish scales are mineralized structures that play important roles in protection and mineral homeostasis. This tissue expresses multiple estrogen receptor subtypes and can be targeted by estrogens or estrogenic endocrine-disrupting compounds, but their effects are poorly explored. The transcriptome data here presented support the findings reported in the research article "Genistein and estradiol have common and specific impacts on the sea bass (Dicentrarchus labrax) skin-scale barrier" [1]. Juvenile sea bass were exposed to estradiol and the phytoestrogen genistein for 1 and 5 days, by intraperitoneal injections, and the effects on scale transcript expression were analysed by RNA-seq using an Illumina Hi-seq 1500. The raw reads of the 30 libraries produced have been deposited in the NCBI-SRA database with the project accession number SRP102504. Mapping of RNA-seq reads against the sea bass reference genome using the Cufflinks/TopHat package identified 371 genes that had significant (FDR<0.05) differential expression with the estradiol or genistein treatments in relation to the control scales at each exposure time, 254 of which presented more than a 2-fold change in expression. The identity of the differentially expressed genes was obtained using both automatic and manual annotations against multiple public sequence databases and they were grouped according to their patterns of expression using hierarchical clustering and heat-maps. The biological processes and KEGG pathways most significantly affected by the estradiol and/or genistein treatments were identified using Cytoscape/ClueGO enrichment analyses.

Epigenetic deregulation is a hallmark of cancer characterized by frequent acquisition of new DNA methylation in CpG islands. To gain insight into the methylation changes of canine DLBCL, we investigated the DNA methylome in primary DLBCLs in comparison with control lymph nodes by genome-wide CpG microarray. We identified 1,194 target loci showing different methylation levels in tumors compared with controls. The hypermethylated CpG loci included promoter, 5'-UTRs, upstream and exonic regions. Interestingly, targets of polycomb repressive complex in stem cells were mostly affected suggesting that DLBCL shares a stem cell-like epigenetic pattern. Functional analysis highlighted biological processes strongly related to embryonic development, tissue morphogenesis and cellular differentiation, including HOX, BMP and WNT. In addition, the analysis of epigenetic patterns and genome-wide methylation variability identified cDLBCL subgroups. Some of these epigenetic subtypes showed a concordance with the clinical outcome supporting the hypothesis that the accumulation of aberrant epigenetic changes results in a more aggressive behavior of the tumor. Collectively, our results suggest an important role of DNA methylation in DLBCL where aberrancies in transcription factors were frequently observed, suggesting an involvement during tumorigenesis. These findings warrant further investigation to improve cDLBCL prognostic classification and provide new insights on tumor aggressiveness.

The European sea bass (Dicentrarchus labrax) is a temperate-zone euryhaline teleost of prime importance for aquaculture and fisheries. This species is subdivided into two naturally hybridizing lineages, one inhabiting the north-eastern Atlantic Ocean and the other the Mediterranean and Black seas. Here we provide a high-quality chromosome-scale assembly of its genome that shows a high degree of synteny with the more highly derived teleosts. We find expansions of gene families specifically associated with ion and water regulation, highlighting adaptation to variation in salinity. We further generate a genome-wide variation map through RAD-sequencing of Atlantic and Mediterranean populations. We show that variation in local recombination rates strongly influences the genomic landscape of diversity within and differentiation between lineages. Comparing predictions of alternative demographic models to the joint allele-frequency spectrum indicates that genomic islands of differentiation between sea bass lineages were generated by varying rates of introgression across the genome following a period of geographical isolation.

Epigenetic modifications are important early events during carcinogenesis. In particular, hypermethylation of CpG islands in the promoter region of tumor suppressor genes is a well-known mechanism of gene silencing that contributes to cancer development and progression. Tissue factor pathway inhibitor 2 (TFPI-2) is a tumor suppressor involved in invasiveness inhibition. Although TFPI-2 transcriptional silencing, through promoter hypermethylation, has been widely reported in several human malignancies, it has never been explored in lymphoma. In the present study TFPI-2 methylation and gene expression have been investigated in canine Diffuse Large B-cell lymphomas (cDLBCL). The methylation level of 23 CpGs located within the TFPI-2 promoter was investigated by bisulfite-specific PCR and next generation amplicon deep sequencing (GS Junior 454, Roche) in 22 cDLBCLs and 9 controls. For the same specimens, TFPI-2 gene expression was assessed by means of Real-time RT-PCR. Sequence analysis clearly demonstrated that TFPI2 is frequently hypermethylated in cDLBCL. Hypermethylation of the TFPI-2 promoter was found in 77% of DLBCLs (17 out of 22) and in one normal lymph node. Globally, dogs with DLBCL showed a mean methylation level significantly increased compared to controls (p<0.01) and analysis of hypermethylation by site identified 19 loci out of 23 (82%) with mean methylation levels from 2- to 120-fold higher in cDLBCL. Gene expression analysis confirmed a significant down-regulation of TFPI-2 (p<0.05) in DLBCLs compared with normal lymph nodes, suggesting that TFPI-2 hypermethylation negatively regulates its transcription. In addition, a significant positive correlation (p<0.01) was found between TFPI-2 methylation levels and age providing the first indication of age-associated epigenetic modifications in canine DLBCL. To conclude, our findings demonstrated that epigenetic dysregulation of TFPI-2, leading to its reduced expression, is frequently detected in canine DLBCL. In the next future, the aberrant TFPI-2 promoter hypermethylation may be considered in association with prognosis and therapy.

Water temperature greatly influences the physiology and behaviour of teleost fish as other aquatic organisms. While fish are able to cope with seasonal temperature variations, thermal excursions outside their normal thermal range might exceed their ability to respond leading to severe diseases and death.Profound differences exist in thermal tolerance across fish species living in the same geographical areas, promoting for investigating the molecular mechanisms involved in susceptibility and resistance to low and high temperatures toward a better understanding of adaptation to environmental challenges. The gilthead sea bream, Sparus aurata, is particularly sensitive to cold and the prolonged exposure to low temperatures may lead to the "winter disease", a metabolic disorder that significantly affects the aquaculture productions along the Northern Mediterranean coasts during winter-spring season. While sea bream susceptibility to low temperatures has been extensively investigated, the cascade of molecular events under such stressful condition is not fully elucidated.

The goldfish (Carassius auratus) uses steroids and prostaglandins as pheromone cues at different stages of the reproductive cycle to facilitate spawning synchronization. Steroid progestin pheromone binding has been detected in goldfish olfactory membranes but the receptors responsible for this specific binding remain unknown. In order to shed some light on the olfactory epithelium transcriptome and search for possible receptor candidates a large set of EST from this tissue were analysed and compared to and combined with a similar zebrafish (Danio rerio) resource.

We identified that the genes heat shock transcription factor 5 (hsf5) and ring finger protein 43 (rnf43) happened fusion in Nile tilapia (Oreochromis niloticus), called hsf5-rnf43, and provided the characteristic and functional analysis of hsf5-rnf43 gene in fish for the first time. Analysis of spatiotemporal expression showed that hsf5-rnf43 was specifically expressed in the testis and located in primary spermatocytes of adult Nile tilapia and gradually increased during testis development from 5 to 180 days after hatching. We also found DNA methylation regulated sex-biased expression of hsf5-rnf43 in the early development of Nile tilapia, and was affected by high temperature during the thermosensitive period of Nile tilapia sex differentiation. Therefore, we first reported that the fusion gene hsf5-rnf43 was sex-biased expressed in the testis regulated by DNA methylation and affected by high temperature, which may be involved in the maintenance of testis function and sex differentiation of Nile tilapia.

It is widely expected that physiological and behavioral stress responses will be integrated within divergent stress-coping styles (SCS) and that these may represent opposite ends of a continuously varying reactive-proactive axis. If such a model is valid, then stress response traits should be repeatable and physiological and behavioral responses should also change in an integrated manner along a major axis of among-individual variation. While there is some evidence of association between endocrine and behavioral stress response traits, few studies incorporate repeated observations of both. To test this model, we use a multivariate, repeated measures approach in a captive-bred population of Xiphophorus birchmanni. We quantify among-individual variation in behavioral stress response to an open field trial (OFT) with simulated predator attack (SPA) and measure waterborne steroid hormone levels (cortisol, 11-ketotestosterone) before and after exposure. Under the mild stress stimulus (OFT), (multivariate) behavioral variation among individuals was consistent with a strong axis of personality (shy-bold) or coping style (reactive-proactive) variation. However, behavioral responses to a moderate stressor (SPA) were less repeatable, and robust statistical support for repeatable endocrine state over the full sampling period was limited to 11-ketotestosterone. Although post hoc analysis suggested cortisol expression was repeatable over short time periods, qualitative relationships between behavior and glucocorticoid levels were counter to our a priori expectations. Thus, while our results clearly show among-individual differences in behavioral and endocrine traits associated with stress response, the correlation structure between these is not consistent with a simple proactive-reactive axis of integrated stress-coping style. Additionally, the low repeatability of cortisol suggests caution is warranted if single observations (or indeed repeat measures over short sampling periods) of glucocorticoid traits are used in ecological or evolutionary studies focussed at the individual level.

Fish scales are an important reservoir of calcium and phosphorus and together with the skin function as an integrated barrier against environmental changes and external aggressors. Histological studies have revealed that the skin and scales regenerate rapidly in fish when they are lost or damaged. In the present manuscript the histological and molecular changes underlying skin and scale regeneration in fed and fasted sea bream (Sparus auratus) were studied using a microarray 3 and 7 days after scale removal to provide a comprehensive molecular understanding of the early stages of these processes.

In the last decade, a new gene family encoding non-rearranging receptors, called novel immune-type receptors (NITRs), has been discovered in teleost fish. NITRs belong to the immunoglobulin superfamily and represent an extraordinarily divergent and rapidly evolving gene complex. Genomic analysis of a region spanning 270 kb led to the discovery of a NITR gene cluster in the European sea bass (Dicentrarchus labrax). In total, 27 NITR genes and three putative pseudogenes, organised in a tandemly arrayed cluster, were identified. Sea bass NITR genes maintain the three major genomic organisations that appear to be essentially conserved among fish species along with new features presumably involving processes of intron loss, exon deletion and acquisition of new exons. Comparative and evolutionary analyses suggest that these receptors have evolved following a "birth-and-death" model of gene evolution in which duplication events together with lineage-specific gain and loss of individual members contributed to the rapid diversification of individual gene families. In this study, we demonstrate that species-specific gene expansions provide the raw material for diversifying, positive Darwinian selection favouring the evolution of a highly diverse array of molecules.

Fish has been deemed suitable to study the complex mechanisms of vertebrate skeletogenesis and gilthead seabream (Sparus aurata), a marine teleost with acellular bone, has been successfully used in recent years to study the function and regulation of bone and cartilage related genes during development and in adult animals. Tools recently developed for gilthead seabream, e.g. mineralogenic cell lines and a 4 × 44K Agilent oligo-array, were used to identify molecular determinants of in vitro mineralization and genes involved in anti-mineralogenic action of vanadate.

Aquaculture represents the most sustainable alternative of seafood supply to substitute for the declining marine fisheries, but severe production bottlenecks remain to be solved. The application of genomic technologies offers much promise to rapidly increase our knowledge on biological processes in farmed species and overcome such bottlenecks. Here we present an integrated platform for mRNA expression profiling in the gilthead sea bream (Sparus aurata), a marine teleost of great importance for aquaculture.

Steroid P450 11beta-hydroxylase, encoded by the CYP11B gene, is a key mitochondrial enzyme for the production of 11-oxygenated androgens, which have been shown to be potent masculinising steroids in several fish species. In this study we have isolated a CYP11B cDNA of 1903 base pairs from the testis of European sea bass (Dicentrarchus labrax) encoding a predicted protein of 552 amino acids. The amino acid identities to other vertebrate 11beta-hydroxylase proteins ranged from 66% to 72% to other fish; 45% to amphibian and 35-39% to mammalian. Southern blot indicated that a single CYP11B gene is present. Northern blot analysis detected two transcripts in testis and head kidney, one of the same size of the isolated clone and the other of 3.9 kb. Reverse transcriptase-polymerase chain reaction showed abundant mRNA expression only in testis and head kidney, being residual in a range of other tissues. Expression of CYP11B and CYP19A (which encodes for ovarian aromatase) was detected from at least 4 days post-hatching and did not appear to be affected by rearing temperature (15 and 20 degrees C) during the first 60 days, a period in which high temperatures promote masculinisation in European sea bass. Throughout, gonadogenesis (60-300 dph), a highly dimorphic pattern of CYP11B expression was consistent with a role of this gene in testicular development.

The common sole (Solea solea) is a promising candidate for European aquaculture; however, the limited knowledge of the physiological mechanisms underlying larval development in this species has hampered the establishment of successful flatfish aquaculture. Although the fact that genomic tools and resources are available for some flatfish species, common sole genomics remains a mostly unexplored field. Here, we report, for the first time, the sequencing and characterisation of the transcriptome of S. solea and its application for the study of molecular mechanisms underlying physiological and morphological changes during larval-to-juvenile transition.

The use of growth-promoters in beef cattle, despite the EU ban, remains a frequent practice. The use of transcriptomic markers has already proposed to identify indirect evidence of anabolic hormone treatment. So far, such approach has been tested in experimentally treated animals. Here, for the first time commercial samples were analyzed.