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Antarctic fish belonging to Notothenioidei represent an extraordinary example of radiation in the cold. In addition to the absence of hemoglobin, icefish show a number of other striking peculiarities including large-diameter blood vessels, high vascular densities, mitochondria-rich muscle cells, and unusual mitochondrial architecture. In order to investigate the bases of icefish adaptation to the extreme Southern Ocean conditions we sequenced the complete genome of the icefish Chionodraco myersi. Comparative analyses of the icefish genome with those of other teleost species, including two additional white-blooded and five red-blooded notothenioids, provided a new perspective on the evolutionary loss of globin genes. Muscle transcriptome comparative analyses against red-blooded notothenioids as well as temperate fish revealed the peculiar regulation of genes involved in mitochondrial function in icefish. Gene duplication and promoter sequence divergence were identified as genome-wide patterns that likely contributed to the broad transcriptional program underlying the unique features of icefish mitochondria. Bargelloni et al. assemble the genome of the icefish Chionodraco myersi and use comparative analysis to examine the basis of adaptation to extreme Southern Ocean conditions. They show evidence for the evolutionary loss of globin genes and the peculiar regulation of genes involved in mitochondrial function. [ABSTRACT FROM AUTHOR]

An amendment to this paper has been published and can be accessed via a link at the top of the paper. [ABSTRACT FROM AUTHOR]

Yellowfin tuna, Thunnus albacares, is one of the most important seafood commodities in the world. Despite its great biological and economic importance, conflicting evidence arises from classical genetic and tagging studies concerning the yellowfin tuna population structure at local and global oceanic scales. Access to more powerful and cost effective genetic tools would represent the first step towards resolving the population structure of yellowfin tuna across its distribution range. Using a panel of 939 neutral Single Nucleotide Polymorphisms (SNPs), and the most comprehensive data set of yellowfin samples available so far, we found genetic differentiation among the Atlantic, Indian and Pacific oceans. The genetic stock structure analysis carried out with 33 outlier SNPs, putatively under selection, identified discrete populations within the Pacific Ocean and, for the first time, also within the Atlantic Ocean. Stock assessment approaches that consider genetic differences at neutral and adaptive genomic loci should be routinely implemented to check the status of the yellowfin tuna, prevent illegal trade, and develop more sustainable management measures. [ABSTRACT FROM AUTHOR]

Background: Photobacteriosis is an infectious disease developed by a Gram-negative bacterium Photobacterium damselae subsp. piscicida (Phdp), which may cause high mortalities (90–100%) in sea bream. Selection and breeding for resistance against infectious diseases is a highly valuable tool to help prevent or diminish disease outbreaks, and currently available advanced selection methods with the application of genomic information could improve the response to selection. An experimental group of sea bream juveniles was derived from a Ferme Marine de Douhet (FMD, Oléron Island, France) selected line using ~ 109 parents (~ 25 females and 84 males). This group of 1187 individuals represented 177 full-sib families with 1–49 sibs per family, which were challenged with virulent Phdp for a duration of 18 days, and mortalities were recorded within this duration. Tissue samples were collected from the parents and the recorded offspring for DNA extraction, library preparation using 2b-RAD and genotyping by sequencing. Genotypic data was used to develop a linkage map, genome wide association analysis and for the estimation of breeding values. Results: The analysis of genetic variation for resistance against Phdp revealed moderate genomic heritability with estimates of ~ 0.32. A genome-wide association analysis revealed a quantitative trait locus (QTL) including 11 SNPs at linkage group 17 presenting significant association to the trait with p-value crossing genome-wide Bonferroni corrected threshold P ≤ 2.22e-06. The proportion total genetic variance explained by the single top most significant SNP was ranging from 13.28–16.14% depending on the method used to compute the variance. The accuracies of predicting breeding values obtained using genomic vs. pedigree information displayed 19–24% increase when using genomic information. Conclusion: The current study demonstrates that SNPs-based genotyping of a sea bream population with 2b-RAD approach is effective at capturing the genetic variation for resistance against Phdp. Prediction accuracies obtained using genomic information were significantly higher than the accuracies obtained using pedigree information which highlights the importance and potential of genomic selection in commercial breeding programs. [ABSTRACT FROM AUTHOR]

Bivalves have colonized the interface between land and sea for the last 500 million years. Although bivalves lack an adaptive immune system, they are extraordinarily well adapted to adverse environmental conditions. Bivalves are valuable aquaculture resources worldwide and are used as sentinels for monitoring pollution. In this work, the immune transcriptomes of mussels (Mytilus galloprovincialis and edulis) and clam (Ruditapes decussatus) were sequenced. For comparative purposes, an already published transcriptome dataset of Ruditapes philippinarum was also included in the analyses. The 454 pyrosequencing of stimulated hemocytes resulted in more than 400,000 reads for each transcriptome. The percentage of annotated sequences ranges from 50% for mussels to 30-40% for clams. Considering the 28,061 non-redundant sequences from the four transcriptomes, the four species share 785 genes. Moreover, sequences related to different putative pathogens were found in the four bivalves. A high number of bivalve herpesvirus ORFs were found, which confirms the value of NGSs as tools to detect and quantify pathogen RNA. Based on an examination of the immune-enriched transcriptomes of these four species, we can conclude that bivalves present an immune system that differs from its conventional characterization as a simple innate immune response against invading pathogens. Enrichment analyses showed that species in the Mytilus genus, especially M. galloprovincialis, possesses a significantly higher number of sequences related to immune processes and killing molecules than species in the Ruditapes genus. This could be related to the broader ecological niche occupied by mussels and the scarcity of reported mussel mass mortalities compared to the high number of mass mortalities reported for clams. [ABSTRACT FROM AUTHOR]

Background: The reuse of dredged sediments in ports and lagoons is a big issue as it should not affect the quality and the equilibrium of ecosystems. In the lagoon of Venice, sediment management is of crucial importance as sediments are often utilized to built-up structures necessary to limit erosion. However, the impact of sediment reuse on organisms inhabiting this delicate area is poorly known. The Manila clam is a filter-feeding species of high economic and ecological value for the Venice lagoon experiencing a drastic decline in the last decades. In order to define the molecular mechanisms behind sediment toxicity, we exposed clams to sediments sampled from different sites within one of the Venice lagoon navigable canals close to the industrial area. Moreover, we investigated the impacts of dredged sediments on clam's microbial communities. Results: Concentrations of the trace elements and organic chemicals showed increasing concentrations from the city of Venice to sites close to the industrial area of Porto Marghera, where PCDD/Fs and PCBs concentrations were up to 120 times higher than the southern lagoon. While bioaccumulation of organic contaminants of industrial origin reflected sediments' chemical concentrations, metal bioaccumulation was not consistent with metal concentrations measured in sediments probably due to the activation of ABC transporters. At the transcriptional level, we found a persistent activation of the mTORC1 signalling pathway, which is central in the coordination of cellular responses to chemical stress. Microbiota characterization showed the over-representation of potential opportunistic pathogens following exposure to the most contaminated sediments, leading to host immune response activation. Despite the limited acquisition of new microbial species from sediments, the latter play an important role in shaping Manila clam microbial communities. Conclusions: Sediment management in the Venice lagoon will increase in the next years to maintain and create new canals as well as to allow the operation of the new mobile gates at the three Venice lagoon inlets. Our data reveal important transcriptional and microbial changes of Manila clams after exposure to sediments, therefore reuse of dredged sediments represents a potential risk for the conservation of this species and possibly for other organisms inhabiting the Venice lagoon. [ABSTRACT FROM AUTHOR]

The application of an electrochemical (impedance) tool for monitoring Escherichia coli contamination in shellfish was evaluated after 13 months of observation. The primary aim of the present study was to compare the standard most probable number (MPN) and µ-trac 4200 (log imped/100 g) for the assessment of E. coli contamination (log MPN/100 g) in non-depurated bivalve mollusks (BM) from five sampling areas of the Veneto-Emilian coast (Italy) (118 samples). The secondary aim was to evaluate the correlation between E. coli concentrations in BM and environmental factors on a large data set (690). The methods showed a moderate, positive correlation (0.60 and 0.69 Pearson and Spearman coefficients, respectively; P<0.01) in Ruditapes philippinarum. The McNemar test indicated analogous sample classification between methods, and the impedance method overestimated the most contaminated class (P=0.03; >4,600 MPN/100 g). The results highlighted the suitability of the impedance method for a faster evaluation and routine use especially in clams, while in Mytilus it seemed less effective. Different models built by multivariate permutational variance analysis and multinomial logistic regression selected the suitable environmental features able to predict the E. coli load. Overall, salinity and season affected the E. coli contamination, whereas locally it was mainly influenced by hydrometry and salinity. The application of the impedance method coupled with environmental data analysis could help purification phase management to adhere to legal limits and could represent an advantage for local control authorities to define actions, considering extreme meteorological events' effects as a proactive reaction to climate change. [ABSTRACT FROM AUTHOR]

Ongoing ocean warming due to climate change poses new challenges for marine life and its exploitation. We have used transcriptomics to find genetically based responses to increased temperature in natural populations of the marine clam Ruditapes decussatus, which lives along parallel thermal gradients in southern Europe. Clams of the Atlantic and West Mediterranean races were collected in northern (cool) and a southern (warm) localities. The animals were kept in running seawater in the warm, southern Atlantic locality for a 15-week period. During this period, water temperature was raised to typical southern European summer values. After this period, an expression profile was obtained for a total of 34 clams and 11,025 probes by means of an oligonucleotide microarray. We found distinct transcriptional patterns for each population based on a total of 552 differentially expressed genes (DEGs), indicating innate differences which probably have a genetic basis. Race and latitude contributed significantly to gene expression differences, with very different sets of DEGs. A gene ontology analysis showed that races differed mainly in the genes involved in ribosomal function and protein biosynthesis, while genes related to glutathione metabolism and ATP synthesis in the mitochondria were the most outstanding with respect to north/south transcriptional differences. [ABSTRACT FROM AUTHOR]

Simple Summary: The clinical importance and the great economic impact of equine asthma (EA), classified as mild (MEA) and severe (SEA), have led to several studies to better understand its features; however, the fragmented information available does not allow researchers to fully describe the pathophysiology of this disease. Transcriptomic analysis can help in understanding the biological mechanisms underlying equine asthma. To date, the great majority of gene expression studies on equine airway diseases have focused on the analysis of only a few candidate genes, hampering robust conclusions about EA pathogenesis. In the present study, the employment of an immune-specific horse array on SEA and MEA-affected horses helped to comprehensively define their transcriptomic fingerprints. For both diseases, findings suggest that the increased amount of mucus in the airways is due to mucociliary clearance reduction rather than mucus hypersecretion. In addition, excessive complement activation might be responsible for tissue injury. Other biological pathways, mainly those involved in the perpetuation of chronic inflammation and tissue remodeling similar to human asthma, are characteristic of SEA. Background: Mild equine asthma (MEA) and severe equine asthma (SEA) are two of the most frequent equine airway inflammatory diseases, but knowledge about their pathogenesis is limited. The goal of this study was to investigate gene expression differences in the respiratory tract of MEA- and SEA-affected horses and their relationship with clinical signs. Methods: Clinical examination and endoscopy were performed in 8 SEA- and 10 MEA-affected horses and 7 healthy controls. Cytological and microbiological analyses of bronchoalveolar lavage (BAL) fluid were performed. Gene expression profiling of BAL fluid was performed by means of a custom oligo-DNA microarray. Results: In both MEA and SEA, genes involved in the genesis, length, and motility of respiratory epithelium cilia were downregulated. In MEA, a significant overexpression for genes encoding inflammatory mediators was observed. In SEA, transcripts involved in bronchoconstriction, apoptosis, and hypoxia pathways were significantly upregulated, while genes involved in the formation of the protective muco-protein film were underexpressed. The SEA group also showed enrichment of gene networks activated during human asthma. Conclusions: The present study provides new insight into equine asthma pathogenesis, representing the first step in transcriptomic analysis to improve diagnostic and therapeutic approaches for this respiratory disease. [ABSTRACT FROM AUTHOR]

The molecular factors and gene regulation involved in sex determination and gonad differentiation in bivalve molluscs are unknown. It has been suggested that doubly uniparental inheritance (DUI) of mitochondria may be involved in these processes in species such as the ubiquitous and commercially relevant Manila clam, Ruditapes philippinarum. We present the first long-read-based de novo genome assembly of a Manila clam, and a RNA-Seq multi-tissue analysis of 15 females and 15 males. The highly contiguous genome assembly was used as reference to investigate gene expression, alternative splicing, sequence evolution, tissue-specific co-expression networks, and sexual contrasting SNPs. Differential expression (DE) and differential splicing (DS) analyses revealed sex-specific transcriptional regulation in gonads, but not in somatic tissues. Co-expression networks revealed complex gene regulation in gonads, and genes in gonad-associated modules showed high tissue specificity. However, male gonad-associated modules showed contrasting patterns of sequence evolution and tissue specificity. One gene set was related to the structural organization of male gametes and presented slow sequence evolution but high pleiotropy, whereas another gene set was enriched in reproduction-related processes and characterized by fast sequence evolution and tissue specificity. Sexual contrasting SNPs were found in genes overrepresented in mitochondrial-related functions, providing new candidates for investigating the relationship between mitochondria and sex in DUI species. Together, these results increase our understanding of the role of DE, DS, and sequence evolution of sex-specific genes in an understudied taxon. We also provide resourceful genomic data for studies regarding sex diagnosis and breeding in bivalves. [ABSTRACT FROM AUTHOR]

The European clam, Ruditapes decussatus is a species with a high commercial importance in Portugal and other Southern European countries. Its production is almost exclusively based on natural recruitment, which is subject to high annual fluctuations. Increased knowledge of the natural reproductive cycle of R. decussatus and its molecular mechanisms would be particularly important in providing new highly valuable genomic information for better understanding the regulation of reproduction in this economically important aquaculture species. In this study, the transcriptomic bases of R. decussatus reproduction have been analysed using a custom oligonucleotide microarray representing 51,678 assembled contigs. Microarray analyses were performed in four gonadal maturation stages from two different Portuguese wild populations, characterized by different responses to spawning induction when used as progenitors in hatchery. A comparison between the two populations elucidated a specific pathway involved in the recognition signals and binding between the oocyte and components of the sperm plasma membrane. We suggest that this pathway can explain part of the differences in terms of spawning induction success between the two populations. In addition, sexes and reproductive stages were compared and a correlation between mRNA levels and gonadal area was investigated. The lists of differentially expressed genes revealed that sex explains most of the variance in gonadal gene expression. Additionally, genes like Foxl2, vitellogenin, condensing 2, mitotic apparatus protein p62, Cep57, sperm associated antigens 6, 16 and 17, motile sperm domain containing protein 2, sperm surface protein Sp17, sperm flagellar proteins 1 and 2 and dpy-30, were identified as being correlated with the gonad area and therefore supposedly with the number and/or the size of the gametes produced. [ABSTRACT FROM AUTHOR]

Background: 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. Results: Global analysis of gene expression identified 4,223 and 4,147 genes differentially expressed (fold change - FC > 1.5) during in vitro mineralization of VSa13 (pre-chondrocyte) and VSa16 (pre-osteoblast) cells, respectively. Comparative analysis indicated that nearly 45% of these genes are common to both cell lines and gene ontology (GO) classification is also similar for both cell types. Up-regulated genes (FC > 10) were mainly associated with transport, matrix/membrane, metabolism and signaling, while down-regulated genes were mainly associated with metabolism, calcium binding, transport and signaling. Analysis of gene expression in proliferative and mineralizing cells exposed to vanadate revealed 1,779 and 1,136 differentially expressed genes, respectively. Of these genes, 67 exhibited reverse patterns of expression upon vanadate treatment during proliferation or mineralization. Conclusions: Comparative analysis of expression data from fish and data available in the literature for mammalian cell systems (bone-derived cells undergoing differentiation) indicate that the same type of genes, and in some cases the same orthologs, are involved in mechanisms of in vitro mineralization, suggesting their conservation throughout vertebrate evolution and across cell types. Array technology also allowed identification of genes differentially expressed upon exposure of fish cell lines to vanadate and likely involved in its anti-mineralogenic activity. Many were found to be unknown or they were never associated to bone homeostasis previously, thus providing a set of potential candidates whose study will likely bring insights into the complex mechanisms of tissue mineralization and bone formation. [ABSTRACT FROM AUTHOR]

Genomics is increasingly applied in breeding programmes for farmed fish and shellfish species around the world. However, current applications do not include information on genome functional activity, which can enhance opportunities to predict relationships between genotypes and phenotypes and hence increase the accuracy of selection. Here, we review prospects for improving aquaculture breeding practises through the uptake of functional genomics data in light of the EU Horizon 2020 project AQUA-FAANG: ' Advancing European Aquaculture by Genome Functional Annotation'. This consortium targeted the six major farmed fish species in European aquaculture, producing thousands of functional genomic datasets from samples representing embryos to mature adults of both sexes, and following immunological stimulation. This data was used to catalogue functional activity across the genome of each species, revealing transcribed regions, distinct chromatin states and regulatory elements impacting gene expression. These functional annotations were shared as open data through the Ensembl genome browser using the latest reference genomes for each species. AQUA-FAANG data offers novel opportunities to identify and prioritize causative genetic variants responsible for diverse traits including disease resistance, which can be exploited to enhance selective breeding. Such knowledge and associated resources have the potential to improve sustainability and boost production in aquaculture by accelerating genetic gain for health and robustness to infection, whilst reducing the requirement for animal testing. We further outline directions to advance and leverage genome functional annotation beyond the AQUA-FAANG project. Given the diversity of aquaculture sectors and businesses, the incorporation of functional genomic information into breeding decisions will depend on technological readiness level and scale of operation, with cost-benefit analysis necessary to determine the most profitable approach for each species and production system. • The AQUAFAANG project has produced improved and richly functionally annotated genomes for six fish species. • The prospects for utilising functional genomic information in breeding decisions is critically reviewed. • Future research opportunities and the need to determine the cost benefit of different genomic technologies is highlighted. [ABSTRACT FROM AUTHOR]

The Mediterranean mussel Mytilus galloprovincialis represents one of the most important aquaculture species in the Mediterranean Sea. The Venice lagoon is an historically important mussels farming area, contributing 25% of the total mussel production of the Veneto region. This area represents a vulnerable ecosystem, subject to significant spatio-temporal variations due to climate change and anthropogenic interventions as for the recent commissioning of MoSE (Experimental Electromechanical Module), a system of mobile barriers aimed at protecting Venice from high tides. In this study, we monitored for two consecutive production cycles three mussels farming sites located in Chioggia at different distance from the southern Venice lagoon inlet. During each season of the two productive cycles, mussels were collected in the three farming sites and site- and seasonal-specific variations of growth, chemical composition and fatty acid profile were investigated. In addition, we also characterized whole gene expression profiles of mussels grown in two investigated farming sites. Biometric traits and gene expression results highlighted narrow differences between rearing sites, inconsistent between the two production cycles. Conversely, seasonality led to important changes in mussel transcriptional profiles, fatty acid composition and condition index due to mussel reproductive cycle and seasonal conditions of the Venice lagoon (e.g. temperature, food availability). Fatty acid differences were primarily linked to dietary changes, rather than stress-related or physiological differences. Regardless of the rearing site, the lowest condition index was observed in October, probably due to intensified metabolic expenditure and depletion of energy reserves to cope with summer stressful environmental conditions. This hypothesis is corroborated by gene expression profiling and by the reduction in the mussel lipid content observed between July and October. The differences between mussels farmed in different sites were mainly related to the hydrodynamics which influences water physico-chemical parameters and feed availability for mussels. Overall biometric traits (i.e. growth) were more favourable in mussels farmed in the most external site characterized by higher hydrodynamics and more favourable water conditions compared to the other two sites. The most favourable condition of mussels farmed close to Chioggia inlet was also measured by gene expression profiles suggesting stressful conditions in inner site at the end of the second production cycle. Despite comparisons between farming sites at transcriptional level showed different results between production cycles, mussels exhibited comparable growth patterns during corresponding months or seasons, suggesting limited and not yet noticeable effects of MoSE commissioning in mussel production. • Venice lagoon mussels farming sites were characterized. • Mussels show site- and seasonal-specific variations of growth, fatty acid and transcriptional profiles. • Fatty acid differences between mussels were primarily linked to dietary changes. • Summer stressful environmental conditions lead to metabolic expenditure and depletion of energy reserves. • Stressful conditions were mainly observed in inner site. [ABSTRACT FROM AUTHOR]

The tight connection between immunity and reproduction has been studied for decades. However, basic knowledge at the molecular level of the effect of mating on immune function is still lacking in many taxa. Determining whether and how the immune system is engaged after mating is a crucial step in understanding post‐mating mechanisms of reproduction and sexual selection. Here, we study the transcriptional changes in immunity‐related genes caused by the ejaculate in the female reproductive tract using a model species for sexual selection studies, the guppy Poecilia reticulata. To study changes triggered by the ejaculate only, rather than caused by mating, we used artificial inseminations to transfer ejaculate into females. We then compared gene expression in the reproductive tract (gonoduct and ovary) of females artificially inseminated either with ejaculate or with a control solution, after 1 hr and after 6 hr. Overall, contact with ejaculate caused short‐term changes in the expression of immune‐related genes in the female reproductive tract, with a complex pattern of up‐ and down‐regulation of immune‐related pathways, but with clear indication of a marked down‐regulation of the immune system shortly after ejaculate contact. This suggests a link between immune function and processes occurring between mating and fertilization in this species. [ABSTRACT FROM AUTHOR]

High‐throughput sequencing has revolutionized population and conservation genetics. RAD sequencing methods, such as 2b‐RAD, can be used on species lacking a reference genome. However, transferring protocols across taxa can potentially lead to poor results. We tested two different IIB enzymes (AlfI and CspCI) on two species with different genome sizes (the loggerhead turtle Caretta caretta and the sharpsnout seabream Diplodus puntazzo) to build a set of guidelines to improve 2b‐RAD protocols on non‐model organisms while optimising costs. Good results were obtained even with degraded samples, showing the value of 2b‐RAD in studies with poor DNA quality. However, library quality was found to be a critical parameter on the number of reads and loci obtained for genotyping. Resampling analyses with different number of reads per individual showed a trade‐off between number of loci and number of reads per sample. The resulting accumulation curves can be used as a tool to calculate the number of sequences per individual needed to reach a mean depth ≥20 reads to acquire good genotyping results. Finally, we demonstrated that selective‐base ligation does not affect genomic differentiation between individuals, indicating that this technique can be used in species with large genome sizes to adjust the number of loci to the study scope, to reduce sequencing costs and to maintain suitable sequencing depth for a reliable genotyping without compromising the results. Here, we provide a set of guidelines to improve 2b‐RAD protocols on non‐model organisms with different genome sizes, helping decision‐making for a reliable and cost‐effective genotyping. [ABSTRACT FROM AUTHOR]

Mass mortalities due to disease outbreaks have recently affected a number of major taxa in marine ecosystems. Climate‐ and pollution‐induced stress may compromise host immune defenses, increasing the risk of opportunistic diseases. Despite growing evidence that mass mortality events affecting marine species worldwide are strongly influenced by the interplay of numerous environmental factors, the reductionist approaches most frequently used to investigate these factors hindered the interpretation of these multifactorial pathologies. In this study, we propose a broader approach based on the combination of RNA‐sequencing and 16S microbiota analyses to decipher the factors underlying mass mortality in the striped venus clam, Chamelea gallina, along the Adriatic coast. On one hand, gene expression profiling and functional analyses of microbial communities showed the over‐expression of several genes and molecular pathways involved in xenobiotic metabolism, suggesting potential chemical contamination in mortality sites. On the other hand, the down‐regulation of several genes involved in immune and stress response, and the over‐representation of opportunistic pathogens such as Vibrio and Photobacterium spp. indicates that these microbial species may take advantage of compromised host immune pathways and defense mechanisms that are potentially affected by chemical exposure, resulting in periodic mortality events. We propose the application of our approach to interpret and anticipate the risks inherent in the combined effects of pollutants and microbes on marine animals in today's rapidly changing environment. [ABSTRACT FROM AUTHOR]

Highlights • There are potential risks associated with consumption of seafood. • NGS-generated microbiome data can be used as place-of-origin marker in seafood. • To trace the clam place of origin combining microbiome with machine-learning. Abstract Developing reliable tools to trace food origin represents a major goal for producers and control authorities. Here, we test the hypothesis whether NGS-generated data could provide a reliable tool to ensure seafood traceability. As a test case, we used the Manila clam Ruditapes philippinarum , a bivalve mollusk of high commercial interest with worldwide distribution, collected in the Venice lagoon sites subjected to prohibition of clam harvesting because of chemical contamination as well as in authorized clam harvesting areas. The results obtained demonstrated that the geographic origin of Manila clam may be more accurately determined basing on microbiome data than single nucleotide polymorphisms. In particular, combining microbiome data with machine-learning techniques, we provide the experimental evidence that it is possible to trace the clam place of origin at high spatial resolution. Considering its low cost and portability, NGS-analysis of microbiome data might represent a cost-effective, high-resolution tool for reliable food traceability. [ABSTRACT FROM AUTHOR]

Among reared bivalves, some “novel species”, such as the great scallop, Pecten maximus , have experienced more difficulty with routine reproduction due to their high sensitivity to biological, chemical, and physical stress during stages of early development. Working with high larval densities requires the use of aeration systems to provide optimal larval suspension and feed distribution. The high susceptibility of the great scallop to aeration in small-volume systems may impose an important limitation in hatchery-based practices. The present study aimed to investigate the processes impacted by aeration in P. maximus veliger larvae exposed to continuous aeration in small-volume tanks (10 L). Aeration appeared as major stressor that was responsible for early mortality among exposed animals (at 96 h after aeration started, haa) when exposure started 13 days after fertilization. Exposed larvae and controls were collected at 12, 24 and 72 haa, and a total of 18 cDNA libraries, each representing a pool of approximately 7,500 larvae, were sequenced, obtaining 358,817,016 raw reads. RNA-seq data were first used to build a de novo transcriptome assembly, and differential transcript abundance was assessed in exposed and control groups; thus, the molecular mechanisms involved in high sensitivity to aeration were deciphered. More than 2,000 transcripts were differentially expressed between exposed and control larvae across the entire time series (logFC > 1, FDR < 5%). Functional analysis revealed that transcriptional changes in larvae exposed to aeration mainly involved the genes that regulate digestive activity and energy metabolism, immune defense, inflammation, apoptosis, larval growth, and development. The results of this study demonstrate that, overall, aeration affects the feeding capacity and energy metabolism of larvae, with expected consequences on the animal's fitness, including its swimming efficiency. Aeration also triggered immune responses and apoptosis, which then increased through opportunistic infections. Notably, infections may be a consequence of a bacterial bloom triggered by the first mortality events that occurred in the culture. This study provides insights into the interactions between environmental variables and great scallop larvae physiology, and the results may contribute to the development of strategies for improving larval rearing practices and ensuring long-term sustainability of P. maximus aquaculture. [ABSTRACT FROM AUTHOR]

The sharp decrease in the cost of RNA-sequencing and the rapid improvement in computational analysis of eco-toxicogenomic data have brought new insights into the adverse effects of chemicals on aquatic organisms. Yet, transcriptomics is generally applied qualitatively in environmental risk assessments, hampering more effective exploitation of this evidence through multidisciplinary studies. In view of this limitation, a methodology is here presented to quantitatively elaborate transcriptional data in support to environmental risk assessment. The proposed methodology makes use of results from the application of Gene Set Enrichment Analysis to recent studies investigating the response of Mytilus galloprovincialis and Ruditapes philippinarum exposed to contaminants of emerging concern. The degree of changes in gene sets and the relevance of physiological reactions are integrated in the calculation of a hazard index. The outcome is then classified according to five hazard classes (from absent to severe), providing an evaluation of whole-transcriptome effects of chemical exposure. The application to experimental and simulated datasets proved that the method can effectively discriminate different levels of altered transcriptomic responses when compared to expert judgement (Spearman correlation coefficient of 0.96). A further application to data collected in two independent studies of Salmo trutta and Xenopus tropicalis exposed to contaminants confirmed the potential extension of the methodology to other aquatic species. This methodology can serve as a proof of concept for the integration of "genomic tools" in environmental risk assessment based on multidisciplinary investigations. To this end, the proposed transcriptomic hazard index can now be incorporated into quantitative Weight of Evidence approaches and weighed, with results from other types of analysis, to elucidate the role of chemicals in adverse ecological effects. [Display omitted] • A methodology is proposed to integrate transcriptomics into WoE approaches for ERA. • The level of transcriptional changes was translated into a hazard index. • Hazard index correlates well with the expert judgement of transcriptional responses. • Index flexibility allows for application of the methodology to different aquatic species. [ABSTRACT FROM AUTHOR]

Human pharmaceuticals such as Acetaminophen, Atenolol and Carbamazepine are pseudo persistent aquatic pollutants with yet unknown sub-lethal effects at environmentally relevant concentrations. Gilthead seabream ( Sparus aurata ) were exposed to Acetaminophen: 31.90 ± 11.07 μg L −1 ; Atenolol: 0.95 ± 0.38 μg L −1 and Carbamazepine: 6.95 ± 0.13 μg L −1 in a 28 day flow through experiment to (1) determine whether exposure to low concentrations in the μg·L −1 range of the pharmaceuticals alters the brain transcriptome and, (2) identify different expression profiles and treatment specific modes of action and pathways. Despite low exposure concentrations, 411, 7 and 612 differently expressed transcripts were identified in the individual treatments with Acetaminophen, Atenolol and Carbamazepine, respectively. Functional analyses of differentially expressed genes revealed a significant over representation of several biological processes, cellular compartment features and molecular functions for both Acetaminophen and Carbamazepine treatments. Overall, the results obtained in seabream brain suggest similar physiological responses to those observed in humans also at environmental concentrations, as well as the existence of treatment specific processes that may be useful for the development of biomarkers of contamination. [ABSTRACT FROM AUTHOR]

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 Gramnegative 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 x 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. [ABSTRACT FROM AUTHOR]

The use of selective breeding is still relatively limited in aquaculture species. Information on such activities is sparse, hindering an overall evaluation of their success. Here, we report on the results of an online survey of the major aquaculture breeding companies operating in Europe. Six main reared fish species were targeted. A total of 31 respondents contributed to the survey, representing 75 % of European breeding organizations. Family-based breeding schemes were predominant, but individual selection was more frequently applied in marine species. Artificial fertilization is the preferred means of reproduction; however, mass spawning is often used as a fallback method. The most frequently selected trait is growth performance, but the number of selected traits has been increasing over the years through the addition of traits such as disease resistance or product quality. The use of molecular tools is now common in all programs, mainly for pedigree traceability. An increasing number of programs use either genomic or marker-assisted selection. Results related to the seed production market confirmed that for Atlantic salmon there are a few dominant players at the European level, with 30-50 % market share. Only part of the European fish aquaculture industry today fully exploits selective breeding to the best advantage. A larger impact assessment still needs to be made by the remainder, particularly on the market share of fish seed (eggs, larvae or juveniles) and its consequences for hatchery stability. [ABSTRACT FROM AUTHOR]

Notothenioidei are typical Antarctic teleosts evolved to adapt to the very low temperatures of the Antarctic seas. Aim of the present paper is to investigate sequence and structure of C3, the third component of the complement system of the notothenioid Trematomus bernacchii and Chionodraco hamatus . We determined the complete nucleotide sequence of two C3 isoforms of T. bernacchii and a single C3 isoform of C. hamatus . These sequences were aligned against other homologous teleost sequences to check for the presence of diversifying selection. Evidence for positive selection was observed in the evolutionary lineage of Antarctic teleost C3 sequences, especially in that of C. hamatus , the most recently diverged species. Adaptive selection affected numerous amino acid positions including three residues located in the anaphylatoxin domain. In an attempt to evaluate the link between sequence variants and specific structural features, we constructed molecular models of Antarctic teleost C3s, of their proteolytic fragments C3b and C3a, and of the corresponding molecules of the phylogenetically related temperate species Epinephelus coioides , using human crystallographic structures as templates. Subsequently, we compared dynamic features of these models by molecular dynamics simulations and found that the Antarctic C3s models show higher flexibility, which likely allows for more pronounced movements of both the TED domain in C3b and the carboxyl-terminal region of C3a. As such dynamic features are associated to positively selected sites, it appears that Antarctic teleost C3 molecules positively evolved toward an increased flexibility, to cope with low kinetic energy levels of the Antarctic marine environment. [ABSTRACT FROM AUTHOR]

Parasites of the genus Perkinsus cause high mortality and economic losses in bivalves commonly produced in global aquaculture. Although the immune responses of oysters and clams naturally infected with Perkinsus marinus or Perkinsus olseni have been extensively studied, there is not much information on host response at the early stages of infection. In this study, we analysed how P . olseni influences the gene expression profiles of haemocytes from the Manila clam ( Venerupis philippinarum ) using temporal experimental infections and an immune-enriched microarray. We identified an early phase of infection that was characterised by no mortality and by the increased expression of genes associated with pathogen recognition, production of nitrogen radicals and antimicrobial activity. Cellular processes such as inhibition of serine proteases and proliferation were also involved in this early response. This phase was followed by an intermediate stage, when the pathogen was most likely multiplying and infecting new areas of the body, and animals began to die. In this stage, many genes related to cell movement were over-expressed. Thirty days after infection metabolic pathway genes were the most affected. Apoptosis appears to be important during pathogenesis. Our results provide novel observations of the broader innate immune response triggered by P . olseni at different infection stages. [ABSTRACT FROM AUTHOR]

Abstract: Pecten maximus, the king scallop, is a bivalve species with important commercial value for both fisheries and aquaculture, traditionally consumed in several European countries. Major problems in larval rearing, however, still limit hatchery-based seed production. High mortalities during early larval stages, likely related to bacterial pathogens, represent the most relevant bottleneck. To address this issue, understanding host defense mechanisms against microbes is extremely important. In this study next-generation RNA-sequencing was carried on scallop hemocytes. To enrich for immune-related transcripts, cDNA libraries from hemocytes challenged in vivo with inactivated-Vibrio anguillarum and in vitro with pathogen-associated molecular patterns, as well as unchallenged controls, were sequenced yielding 216,444,674 sequence reads. De novo assembly of the scallop hemocyte transcriptome consisted of 73,732 contigs (31% annotated). A total of 934 contigs encoded proteins with a known immune function, grouped into several functional categories. Particular attention was reserved to Toll-like receptors (TLRs), a family of pattern recognition receptors (PRRs) involved in non-self recognition. Through mining the scallop hemocyte transcriptome, at least four TLRs could be identified. The organization of canonical TLR domains demonstrated that single cysteine cluster and multiple cysteine cluster TLRs co-exist in this species. In addition, preliminary data concerning their mRNA level following bacterial challenge suggested that different members of this family could exhibit opposite responses to pathogenic stimuli. Finally, a global analysis of differential expression comparing gene-expression levels in in vitro and in vivo stimulated hemocytes against controls provided evidence on a large set of transcripts involved in the great scallop immune response. [Copyright &y& Elsevier]

Simple Summary: Selective breeding programs based on genomic data are still not a common practice in aquaculture, although genomic selection has been widely demonstrated to be advantageous when trait phenotyping is a difficult task. In this study, we investigated the accuracy of predicting the phenotype and the estimated breeding value (EBV) of three Bayesian models and a Random Forest algorithm exploiting the information of a genome-wide SNP panel for European sea bass. The genomic predictions were developed for mortality caused by viral nervous necrosis, post-stress cortisol concentration, antibody titer against nervous necrosis virus and body weight. Selective breeding based on genomic data is a possible option for improving these traits while overcoming difficulties related to individual phenotyping of the investigated traits. Our results evidenced that the EBV used as a pseudo-phenotype enhances the predictive performances of genomic models, and that EBV can be predicted with satisfactory accuracy. The genomic prediction of the EBV for mortality might also be used to classify the phenotype for the same trait. In European sea bass (Dicentrarchus labrax L.), the viral nervous necrosis mortality (MORT), post-stress cortisol concentration (HC), antibody titer (AT) against nervous necrosis virus and body weight (BW) show significant heritability, which makes selective breeding a possible option for their improvement. An experimental population (N = 650) generated by a commercial broodstock was phenotyped for the aforementioned traits and genotyped with a genome-wide SNP panel (16,075 markers). We compared the predictive accuracies of three Bayesian models (Bayes B, Bayes C and Bayesian Ridge Regression) and a machine-learning method (Random Forest). The prediction accuracy of the EBV for MORT was approximately 0.90, whereas the prediction accuracies of the EBV and the phenotype were 0.86 and 0.21 for HC, 0.79 and 0.26 for AT and 0.71 and 0.38 for BW. The genomic prediction of the EBV for MORT used to classify the phenotype for the same trait showed moderate classification performance. Genome-wide association studies confirmed the polygenic nature of MORT and demonstrated a complex genetic structure for HC and AT. Genomic predictions of the EBV for MORT could potentially be used to classify the phenotype of the same trait, though further investigations on a larger experimental population are needed. [ABSTRACT FROM AUTHOR]

Abstract: Numerous studies have demonstrated that environmental parameters affect bivalve immunomarkers. In the present study, we tested the hypothesis that clams (Venerupis philippinarum) collected in sites with different environmental conditions respond differently to experimental contaminant exposure. Clams were collected at two sites within the Lagoon of Venice that are influenced differently by both anthropogenic impact and natural conditions: Marghera, which is characterised by relatively high contamination levels and restricted clam fishing, and Chioggia, which is inside a licensed clam culture area that is characterised by lower contamination levels. Total haemocyte count, haemocyte diameter and volume, lysozyme activity in both haemocyte lysate and cell-free haemolymph, superoxide dismutase and catalase activities in gills and digestive glands were measured at time 0 (clam sampling time), after 7 days of acclimation in the laboratory and after 1, 3 and 7 days of copper exposure. Interestingly, statistical analyses (three-way ANOVA and Canonical Correlation Analysis) revealed persistent differences in the biological responses of clams from the two sampling sites before and after copper exposure. Conversely, the influence of copper on cellular and biochemical parameters was negligible. Overall, the results obtained indicated that animals with a different ecological history respond differently to experimental contaminant exposure. In addition, this study suggested that immunomarkers and other biomarkers might be used to determine the origin of fishing products. [Copyright &y& Elsevier]

Abstract: The halophilic bacterium Photobacterium damselae subsp. piscicida (Phdp) represents a substantial health problem for several fish species in aquaculture. Bacteria that reside free and inside phagocytes cause acute and chronic forms of photobacteriosis. Infections of juveniles rapidly kill up to 90–100% fish. Factors underlying failure of the immune protection against bacteria remain largely unknown. The reported study used a transcriptomic approach to address this issue. Juvenile sea breams (0.5 g) were challenged by immersion in salt water containing 2.89 × 108 CFU of a virulent Phdp and the head kidney was sampled after 24- and 48-h. Analyses were performed using the second version of a 44 k oligonucleotide DNA microarray that represents 19,734 sea bream unique transcripts and covers diverse immune pathways. Expression changes of selected immune genes were validated with qPCR. Results suggested rapid recognition of the pathogen, as testified by up-regulation of lectins and antibacterial proteins (bactericidal permeability-increasing protein lectins, lysozyme, intracellular and extracellular proteases), chemokines and chemokine receptors. Increased expression of proteins involved in iron and heme metabolism also could be a response against bacteria that are dependent on iron. However, negative regulators of immune/inflammatory response were preponderant among the up-regulated genes. A remarkable finding was the increased expression of IL-10 in concert with up-regulation of arginase I and II and proteins of the polyamine biosynthesis pathway that diverts the arginine flux from the production of reactive nitrogen species. Such expression changes are characteristic for alternatively activated macrophages that do not develop acute inflammatory responses. Immune suppression can be induced by the host to reduce tissue damages or by the pathogen to evade host response. [Copyright &y& Elsevier]

Abstract: In the present study we address the investigation of TLR2 evolutionary selection in two Antarctic teleosts, Trematomus bernacchii (Nototheniidae) and Chionodraco hamatus (Channichthyidae). The nucleotide sequence of TLR2 has been determined in both species, encoding 20 leucine-rich repeats (LRRs) in the extracellular region and a classical Toll/IL-1R (TIR) domain in the intracellular region. High expression level of T. bernacchii TLR2 was found in spleen and skin. Using different methods we identified six codons that underwent Darwinian selection while 20 were found to be negatively selected. Molecular models of C. hamatus and T. bernacchii TLR2 ectodomain as well as of the TIR domain were built by Homology Modeling. Molecular Dynamics simulations were performed in water for 15 ns. The sites under positive selection were residing on the convex side of the solenoid, four out of six were in a 35-residue-long region including the central/N-terminal domain boundary: two in the external loop of LRR11 and the other two in the LRR12 loop. This region has been demonstrated to be the functional site of ligand interaction in human TLR2 structure. Antarctic TLR2 models showed more flexibility than TLR2 from the temperate species Gasterosteus aculeatus. These results suggest that the selective pressure has shaped TLR2 molecule in such a way that increased its activity under the peculiar Antarctic environmental conditions. [Copyright &y& Elsevier]

The introduction of Next Generation Sequencing (NGS) has revolutionised population genetics, providing studies of non- model species with unprecedented genomic coverage, allowing evolutionary biologists to address questions previously far beyond the reach of available resources. Furthermore, the simple mutation model of Single Nucleotide Polymorphisms (SNPs) permits cost-effective high-throughput genotyping in thousands of individuals simultaneously. Genomic resources are scarce for the Atlantic herring (Clupea harengus), a small pelagic species that sustains high revenue fisheries. This paper details the development of 578 SNPs using a combined NGS and high-throughput genotyping approach. Eight individuals covering the species distribution in the eastern Atlantic were bar- coded and multiplexed into a single cDNA library and sequenced using the 454 GS FLX platform. SNP discovery was performed by de novo sequence clustering and contig assembly, followed by the mapping of reads against consensus contig sequences. Selection of candidate SNPs for genotyping was conducted using an in silico approach. SNP validation and genotyping were performed simultaneously using an Illumina 1,536 GoldenGate assay. Although the conversion rate of candidate SNPs in the genotyping assay cannot be predicted in advance, this approach has the potential to maximise cost and time efficiencies by avoiding expensive and time-consuming laboratory stages of SNP validation. Additionally, the in silico approach leads to lower ascertainment bias in the resulting SNP panel as marker selection is based only on the ability to design primers and the predicted presence of intron-exon boundaries. Consequently SNPs with a wider spectrum of minor allele frequencies (MAFs) will be genotyped in the final panel. The genomic resources presented here represent a valuable multi-purpose resource for developing informative marker panels for population discrimination, microarray development and for population genomic studies in the wild. [ABSTRACT FROM AUTHOR]

Background: 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. Results: Quantitative determination of Dexamethasone (DEX) residues in the urine collected at the slaughterhouse was performed by Liquid Chromatography-Mass Spectrometry (LC-MS). DNA-microarray technology was used to obtain transcriptomic profiles of skeletal muscle in commercial samples and negative controls. LC-MS confirmed the presence of low level of DEX residues in the urine of the commercial samples suspect for histological classification. Principal Component Analysis (PCA) on microarray data identified two clusters of samples. One cluster included negative controls and a subset of commercial samples, while a second cluster included part of the specimens collected at the slaughterhouse together with positives for corticosteroid treatment based on thymus histology and LC-MS. Functional analysis of the differentially expressed genes (3961) between the two groups provided further evidence that animals clustering with positive samples might have been treated with corticosteroids. These suspect samples could be reliably classified with a specific classification tool (Prediction Analysis of Microarray) using just two genes. Conclusions: Despite broad variation observed in gene expression profiles, the present study showed that DNA-microarrays can be used to find transcriptomic signatures of putative anabolic treatments and that gene expression markers could represent a useful screening tool. [ABSTRACT FROM AUTHOR]

Background: Genomic and transcriptomic approaches have the potential for unveiling the genome-wide response to environmental perturbations. The abundance of the catadromous European eel (Anguilla anguilla) stock has been declining since the 1980s probably due to a combination of anthropogenic and climatic factors. In this paper, we explore the transcriptomic dynamics between individuals from high (river Tiber, Italy) and low pollution (lake Bolsena, Italy) environments, which were measured for 36 PCBs, several organochlorine pesticides and brominated flame retardants and nine metals. Results: To this end, we first (i) updated the European eel transcriptome using deep sequencing data with a total of 640,040 reads assembled into 44,896 contigs (Eeelbase release 2.0), and (ii) developed a transcriptomic platform for global gene expression profiling in the critically endangered European eel of about 15,000 annotated contigs, which was applied to detect differentially expressed genes between polluted sites. Several detoxification genes related to metabolism of pollutants were upregulated in the highly polluted site, including genes that take part in phase I of the xenobiotic metabolism (CYP3A), phase II (glutathione-S-transferase) and oxidative stress (glutathione peroxidase). In addition, key genes in the mitochondrial respiratory chain and oxidative phosphorylation were down-regulated at the Tiber site relative to the Bolsena site. Conclusions: Together with the induced high expression of detoxification genes, the suggested lowered expression of genes supposedly involved in metabolism suggests that pollution may also be associated with decreased respiratory and energy production. [ABSTRACT FROM AUTHOR]

Background: The Manila clam, Ruditapes philippinarum, is one of the major aquaculture species in the world and a potential sentinel organism for monitoring the status of marine ecosystems. However, genomic resources for R. philippinarum are still extremely limited. Global analysis of gene expression profiles is increasingly used to evaluate the biological effects of various environmental stressors on aquatic animals under either artificial conditions or in the wild. Here, we report on the development of a transcriptomic platform for global gene expression profiling in the Manila clam. Results: A normalized cDNA library representing a mixture of adult tissues was sequenced using a ultra high-throughput sequencing technology (Roche 454). A database consisting of 32,606 unique transcripts was constructed, 9,747 (30%) of which could be annotated by similarity. An oligo-DNA microarray platform was designed and applied to profile gene expression of digestive gland and gills. Functional annotation of differentially expressed genes between different tissues was performed by enrichment analysis. Expression of Natural Antisense Transcripts (NAT) analysis was also performed and bi-directional transcription appears a common phenomenon in the R. philippinarum transcriptome. A preliminary study on clam samples collected in a highly polluted area of the Venice Lagoon demonstrated the applicability of genomic tools to environmental monitoring. Conclusions: The transcriptomic platform developed for the Manila clam confirmed the high level of reproducibility of current microarray technology. Next-generation sequencing provided a good representation of the clam transcriptome. Despite the known limitations in transcript annotation and sequence coverage for non model species, sufficient information was obtained to identify a large set of genes potentially involved in cellular response to environmental stress. [ABSTRACT FROM AUTHOR]

The article presents a study on the effect of global warming on Notothenioids, a taxonomic group that dominates Antarctic fish. It notes that the genetic constraints of this group limits their adaptation to climate change specifically the scenario in Southern Ocean. An observation on Trematomus bernacchii, an Antarctic fish, responding to heat stress suggests a lack of specific heat-shock response. A study of macro- and micro-evolutionary processes of Notothenioids is said to be essential to understand better how they can cope with climate change.

Abstract: Fish pasteurellosis is an infectious disease that affects several fish species living in marine temperate waters. Its causative agent is the Gram-negative bacterium Photobacterium damselae subsp. piscicida (Phdp). Fish pasteurellosis represents a serious health problem for the majority of intensive sea bream hatcheries, with 90–100% mortality during disease outbreaks. Larvae and juveniles are the most susceptible stages. A potential strategy to prevent fish pasteurellosis is to select for animals that are genetically resistant to it. The aim of this work was to evaluate the genetic variation of disease resistance and growth in the gilthead sea bream (Sparus aurata L.). An experimental population, approximately 3500 animals originating from mass spawning of four broodstocks, was experimentally infected with a highly virulent strain of Phdp. Mortality was monitored daily for 19days. Upon completion of the challenge experiment, genetic profiles at nine microsatellite loci were obtained for 1753 animals and for all (256) broodstock fish, for parentage assignment. A subset of families defined by means of genetic analysis was used to calculate heritability for survival to infection. For body length (measured post-mortem), heritability was 0.38±0.07. The heritability for disease resistance varied depending on how resistance was defined. Heritability was 0.12±0.04 for days of survival post challenge, defined as a continuous trait, while it ranged from 0.45±0.04 to 0.18±0.08 for the binary trait dead/alive at a specific day. The genetic correlation between body length and survival was positive and significant (r =0.61±0.16). These results confirm the existence of genetic variation for growth and resistance against Phpd and highlight also the potential for selective breeding programs to improve these traits. [Copyright &y& Elsevier]

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. [ABSTRACT FROM AUTHOR]

The identification of the genetic basis of domestication in fish species is of timely importance for the aquaculture industry in order to increase productivity, quality, and the welfare of farmed fish. The goal of this study is to investigate the largely unknown aquaculture-induced evolution in gilthead seabream, which is one of the most important farmed fish in the Mediterranean region. We used a panel of 1159 genome-wide SNPs, and genotyped 956 fish from 23 wild populations of Mediterranean-wide distribution and 362 farmed fish from five Greek hatcheries. We assessed the genetic diversity of the sampled populations and contrasted the results of four different approaches of outlier detection methods. We recognized one very strong candidate and two good candidate SNPs with evidence for aquaculture-induced evolution in gilthead seabream. The annotation of these SNPs revealed neighboring genes with biological roles from stress tolerance and disease resistance to sexual maturation that may explain our observations. In conclusion, we demonstrate that the genome of gilthead seabream, despite the fact that the species is often suggested to be in the early stages of the domestication process, shows evidence of aquaculture-induced evolution. We report on a list of genes that may explain our observations and that may be investigated further. We anticipate that our findings will stimulate additional research with the use of SNP panels of higher density that can elucidate the genomic architecture of domestication in this species of high aquacultural interest. [ABSTRACT FROM AUTHOR]

C6O4 (difluoro{[2,2,4,5-tetrafluoro-5-(trifluoromethoxy)-1,3-dioxolan-4-yl]oxy}acetic acid) is a new surfactant and emulsifier used as a substitute of perfluorooctanoic acid (PFOA). Recently, C6O4 has been detected in aquatic environments, but, at present, no information concerning the effects of C6O4 on aquatic species, such as bivalves, are available in the literature. Therefore, in this study we evaluated for the first time the effects of C6O4 (0.1 and 1 µg/L) and PFOA (1 µg/L) to the clam Ruditapes philippinarum. Short-term (7 days) and long-term (21 days) exposures of clams to the two compounds were carried out and numerous biomarkers were measured in haemocytes/haemolymph, as well as in gills and digestive gland. The MANOVA analysis demonstrated statistically significant effects of the independent variables "treatment", "time" and "treatment-time interaction" on the whole dataset of biomarker responses. The two-way ANOVA analysis performed for each biomarker response indicated that the two compounds affected most of the cellular and tissue parameters measured. Despite preliminary, the results obtained suggested that C6O4 - similarly to PFOA - can affect both cellular and biochemical parameters of clams. [Display omitted] • C6O4 and PFOA effects to clams were evaluated for the first time. • 1 μg/L of C6O4 or PFOA affected THC and cell proliferation after 7 days. • 1 μg/L of C6O4 or PFOA affected cell volume and diameter after 7 days. • C6O4 or PFOA affected antioxidants after 7 and 21 days. • C6O4 and PFOA seemed to act in a similar way. [ABSTRACT FROM AUTHOR]

Abstract: Toll-Like Receptors (TLRs) have recently emerged as key sensors of invading microbes, acting through recognition of pathogen-associated molecular patterns. It has been demonstrated that TLR9 is involved in the recognition of unmethylated CpG motifs in mice, humans, and pigs. We report here the full-length sequence of TLR9 cDNA in the gilthead sea bream (Sparus aurata L.). The predicted protein (1063 amino acids) was similar to mammalian TLR9s, showing 21 leucine-rich repeats in the extracellular region and a typical Toll/IL-1R (TIR) domain in the intracellular region. Comparative analysis of TLR9 sequences indicated that critical residues for ligand-binding are conserved across vertebrate lineages, although evidence of functional divergence was observed. Analysis of the genomic structure of sea bream TLR9 gene revealed the presence of two intervening sequences. Retention of the second intron produced an alternatively spliced mRNA (TLR9B) showing differential expression among tissues or developmental stages compared to the wild-type isoform (TLR9A). RT-PCR analysis indicated a broad expression of TLR9A, especially in immune-related organs (spleen, head-kidney) and mucosal–epithelial barriers (gills, gut, skin). Using quantitative Real-Time RT-PCR, no statistically significant variation was observed for TLR9 mRNAs expression in the spleen of experimentally infected animals compared to healthy controls. Comparing sequence and expression profile of sea bream TLR9 with mammalian TLR9s suggested that the main function of TLR9 might be conserved across vertebrates, although species-specific features are present (modulation of ligand-binding specificity, alternative splicing). [Copyright &y& Elsevier]

There is growing concern for the wide use of perfluorooctanoic acid (PFOA) because of its toxic effects on the environment and on human health. A new compound – the so called C6O4 (perfluoro ([5-methoxy-1,3-dioxolan-4-yl]oxy) acetic acid) - was recently introduced as one of the alternative to traditional PFOA, however this was done without any scientific evidence of the effects of C6O4 when dispersed into the environment. Recently, the Regional Agency for the Protection of the Environment of Veneto (Italy) detected high levels of C6O4 in groundwater and in the Po river, increasing the alarm for the potential effects of this chemical into the natural environment. The present study investigates for the first time the effects of C6O4 on the Manila clam Ruditapes philippinarum exposed to environmental realistic concentrations of C6O4 (0.1 µg/L and 1 µg/L) for 7 and 21 days. Furthermore, in order to better understand if C6O4 is a valid and less hazardous alternative to its substitute, microbial and transcriptomic alterations were also investigated in clams exposed to 1 µg/L of PFOA. Results indicate that C6O4 may cause significant perturbations to the digestive gland microbiota, likely determining the impairment of host physiological homeostasis. Despite chemical analyses suggest a 5 times lower accumulation potential of C604 as compared to PFOA in clam soft tissues, transcriptional analyses reveal several alterations of gene expression profile. A large part of the altered pathways, including immune response, apoptosis regulation, nervous system development, lipid metabolism and cell membrane is the same in C6O4 and PFOA exposed clams. In addition, clams exposed to C6O4 showed dose-dependent responses as well as possible narcotic or neurotoxic effects and reduced activation of genes involved in xenobiotic metabolism. Overall, the present study suggests that the potential risks for marine organism following environmental contamination are not reduced by replacing PFOA with C6O4. In addition, the detection of both C6O4 and PFOA into tissues of clams inhabiting the Lagoon of Venice - where there are no point sources of either compounds - recommends a similar capacity to spread throughout the environment. These results prompt the urgent need to re-evaluate the use of C6O4 as it may represent not only an environmental hazard but also a potential risk for human health. [ABSTRACT FROM AUTHOR]