Infection of Norway lobster (Nephrops norvegicus) by the parasite Hematodinium sp.: insights from 30 years of field observations.

The Norway lobster, Nephrops norvegicus, is an important representative of the benthos and also supports valuable fisheries across Europe. Nephrops are susceptible to infection by Hematodinium sp., an endoparasitic dinoflagellate that causes morbidity and mortality. From an epizootiological perspective, the Clyde Sea Area (CSA; west of Scotland) is the best-studied Hematodinium-Nephrops pathosystem, with historical data available between 1988 and 2008. We have revisited this pathosystem by curating and updating prevalence values, differentiating host traits associated with disease exposure and progression, and comparing Hematodinium sp. disease dynamics in the CSA to other locations and to other decapod hosts (Cancer pagurus, Carcinus maenas). Prevalence from a 2018/2019 survey (involving 1739 lobsters) revealed Hematodinium sp. still mounts a synchronized patent infection in the CSA; hence this pathogen can be considered as enzootic in this location. We highlight for the first time that Nephrops size is associated with high severity infection, while females are more exposed to Hematodinium sp. More generally, regardless of the host (Norway lobster, brown and shore crabs) or the geographical area (Ireland, Wales, Scotland), Hematodinium sp. patent infections peak in spring/summer and reach their nadir during autumn. We contend that Hematodinium must be considered one of the most important pathogens of decapod crustaceans in temperate waters.

Circadian rhythm of preferred temperature in fish: Behavioural thermoregulation linked to daily photocycles in zebrafish and Nile tilapia.

Ectothermic vertebrates, e.g. fish, maintain their body temperature within a specific physiological range mainly through behavioural thermoregulation. Here, we characterise the presence of daily rhythms of thermal preference in two phylogenetically distant and well-studied fish species: the zebrafish (Danio rerio), an experimental model, and the Nile tilapia (Oreochromis niloticus), an aquaculture species. We created a non-continuous temperature gradient using multichambered tanks according to the natural environmental range for each species. Each species was allowed to freely choose their preferred temperature during the 24h cycle over a long-term period. Both species displayed strikingly consistent temporal daily rhythms of thermal preference with higher temperatures being selected during the second half of the light phase and lower temperatures at the end of the dark phase, with mean acrophases at Zeitgeber Time (ZT) 5.37 h (zebrafish) and ZT 12.5 h (tilapia). Interestingly, when moved to the experimental tank, only tilapia displayed consistent preference for higher temperatures and took longer time to establish the thermal rhythms. Our findings highlight the importance of integrating both light-driven daily rhythm and thermal choice to refine our understanding of fish biology and improve the management and welfare of the diversity of fish species used in research and food production.

Optimization of Low-Biomass Sample Collection and Quantitative PCR-Based Titration Impact 16S rRNA Microbiome Resolution.

The major aquatic interface between host and environment in teleost finfish species is the gill. The diversity of this infraclass, high complexity of the organ, and its direct exposure to the surrounding environment make it an ideal candidate for furthering our understanding of the intertwined relationships between host and microbiome. Capturing the structure and diversity of bacterial communities from this low-biomass, inhibitor-rich tissue can, however, prove challenging. Lessons learned in doing so are directly applicable to similar sample types in other areas of microbiology. Through the development of a quantitative PCR assay for both host material and 16S rRNA genes, we tested and developed a robust method for low-biomass sample collection which minimized host DNA contamination. Quantification of 16S rRNA facilitated not only the screening of samples prior to costly library construction and sequencing but also the production of equicopy libraries based on 16S rRNA gene copies. A significant increase in diversity of bacteria captured was achieved, providing greater information on the true structure of the microbial community. Such findings offer important information for determining functional processes. Results were confirmed across fresh, brackish, and marine environs with four different fish species, with results showing broad homology between samples, demonstrating the robustness of the approach. Evidence presented is widely applicable to samples similar in composition, such as sputum or mucus, or those that are challenging due to the inherent inclusion of inhibitors. IMPORTANCE The interaction between the fish gill and surrounding bacteria-rich water provides an intriguing model for examining the interaction between the fish, free-floating bacteria, and the bacterial microbiome on the gill surface. Samples that are inherently low in bacteria, or that have components that inhibit the ability to produce libraries that identify the components of microbial communities, present significant challenges. Gill samples present both of these types of challenges. We developed methods for quantifying both the bacterial and host DNA material and established a sampling method which both reduced inhibitor content and maximized bacterial diversity. By quantifying and normalizing bacteria prior to library construction, we showed significant improvements with regards to the fidelity of the final data. Our results support wide-ranging applications for analyzing samples of similar composition, such as mucus and sputum, in other microbiological spheres.

Changes in circulating insulin-like growth factor-1 and its binding proteins in yearling rainbow trout during spring under natural and manipulated photoperiods and their relationships with gill Na+, K+-ATPase and body size.

Smoltification in salmonids occurs during spring in response to increasing photoperiod to prepare for marine life. Smoltification is associated with increased hypo-osmoregulatory ability and enhanced growth potential, mediated by growth hormone and insulin-like growth factor (IGF)-1. Rainbow trout is uniquely insensitive to the induction of smoltification-associated changes by photoperiod, such as the activation of gill Na+,K+-ATPase (NKA). We measured the circulating IGF-1 and IGF-binding protein (IGFBP)-2b levels in yearling rainbow trout exposed to natural and manipulated photoperiods during spring and correlated these with gill NKA activity and body size. Although the effect of photoperiod manipulation on body size and circulating IGF-1 and IGFBP-2b was negligible, they were positively correlated with gill NKA activity in fish under simulated natural photoperiod. We next pit-tagged yearling rainbow trout and fed them a restricted ration or to satiation under a natural photoperiod. In April, gill NKA activity was higher in the satiation group than in the restricted group and positively correlated with body size and growth rate. In addition, circulating IGFBP-2b was positively correlated with gill NKA, size and growth, whereas circulating IGF-1 was correlated only with size and growth. The relationship between circulating IGF-1 and growth intensified from May to June, suggesting that the IGF-1-growth relationship was disrupted in April when gill NKA was activated. Two additional IGFBPs were related to growth parameters but not to gill NKA activity. The present study suggests that circulating IGFBP-2b and IGF-1 mediate the size-dependent activation of gill NKA in yearling rainbow trout during spring.

Characterization of External Mucosal Microbiomes of Nile Tilapia and Grey Mullet Co-cultured in Semi-Intensive Pond Systems.

The external mucosal surfaces of the fish harbor complex microbial communities, which may play pivotal roles in the physiological, metabolic, and immunological status of the host. Currently, little is known about the composition and role of these communities, whether they are species and/or tissue specific and whether they reflect their surrounding environment. Co-culture of fish, a common practice in semi-intensive aquaculture, where different fish species cohabit in the same contained environment, is an easily accessible and informative model toward understanding such interactions. This study provides the first in-depth characterization of gill and skin microbiomes in co-cultured Nile tilapia (Oreochromis niloticus) and grey mullet (Mugil capito) in semi-intensive pond systems in Egypt using 16S rRNA gene-based amplicon sequencing. Results showed that the microbiome composition of the external surfaces of both species and pond water was dominated by the following bacterial phyla: Proteobacteria, Fusobacteriota, Firmicutes, Planctomycetota, Verrucomicrobiota, Bacteroidota, and Actinobacteriota. However, water microbial communities had the highest abundance and richness and significantly diverged from the external microbiome of both species; thus, the external autochthonous communities are not a passive reflection of their allochthonous communities. The autochthonous bacterial communities of the skin were distinct from those of the gill in both species, indicating that the external microbiome is likely organ specific. However, gill autochthonous communities were clearly species specific, whereas skin communities showed higher commonalities between both species. Core microbiome analysis identified the presence of shared core taxa between both species and pond water in addition to organ-specific taxa within and between the core community of each species. These core taxa included possibly beneficial genera such as Uncultured Pirellulaceae, Exiguobacterium, and Cetobacterium and opportunistic potential pathogens such as Aeromonas, Plesiomonas, and Vibrio. This study provides the first in-depth mapping of bacterial communities in this semi-intensive system that in turn provides a foundation for further studies toward enhancing the health and welfare of these cultured fish and ensuring sustainability.

Testing of NKA expression by mobile real time PCR is an efficient indicator of smoltification status of farmed Atlantic salmon.

Assessment of seawater readiness of freshwater salmon smolts is a crucial husbandry step with economic implications in salmon aquaculture but current methods rely on delayed centralised enzymic activity measurement. The efficiency of a qRT-PCR assay for sodium potassium ATPase (NKA) α1a mRNA was tested in a 3-year study on 19 hatcheries across Scotland incorporating environmental factors such as temperature and metal contamination. The NKA qRT-PCR assay was transferred to a mobile laboratory and on-site testing was carried out at 3 hatchery sites. For the first two years standard enzymatic and gene expression assays had similar success rates in detecting smoltification (NKA activity 60%, qRT-PCR 57%). In the third year, all but one site were determined as sea water ready by qRT-PCR but only at 4 by enzymatic testing. On site testing with mobile qRT-PCR was successfully performed on four farm sites. Altogether, high sensitivity was shown for the in lab (98.9%, SE 0.24) and mobile (93.43%, SE 0.119) assays when tested using a quantitative RNA standard. Some indication for obscured smoltification assay results due to environmental increased heavy metal contamination was observed. Our results prove it is possible to test a smoltification marker on site and provide results on the day of testing during the smolt period allowing for informed decisions on seawater transfer.

Endoplasmic reticulum stress as a key mechanism in stunted growth of seawater rainbow trout (Oncorhynchus mykiss).

BACKGROUND: Rainbow trout (Oncorhynchus mykiss) is a salmonid species with a complex life-history. Wild populations are naturally divided into freshwater residents and sea-run migrants. Migrants undergo an energy-demanding adaptation for life in seawater, known as smoltification, while freshwater residents display these changes in an attenuated magnitude and rate. Despite this, in seawater rainbow trout farming all fish are transferred to seawater. Under these circumstances, weeks after seawater transfer, a significant portion of the fish die (around 10%) or experience growth stunting (GS; around 10%), which represents an important profitability and welfare issue. The underlying causes leading to GS in seawater-transferred rainbow trout remain unknown. In this study, we aimed at characterising the GS phenotype in seawater-transferred rainbow trout using untargeted and targeted approaches. To this end, the liver proteome (LC-MS/MS) and lipidome (LC-MS) of GS and fast-growing phenotypes were profiled to identify molecules and processes that are characteristic of the GS phenotype. Moreover, the transcription, abundance or activity of key proteins and hormones related to osmoregulation (Gill Na+, K + -ATPase activity), growth (plasma IGF-I, and liver igf1, igfbp1b, ghr1 and ctsl) and stress (plasma cortisol) were measured using targeted approaches. RESULTS: No differences in Gill Na+, K + -ATPase activity and plasma cortisol were detected between the two groups. However, a significant downregulation in plasma IGF-I and liver igf1 transcription pointed at this growth factor as an important pathomechanism for GS. Changes in the liver proteome revealed reactive-oxygen-species-mediated endoplasmic reticulum stress as a key mechanism underlying the GS phenotype. From the lipidomic analysis, key observations include a reduction in triacylglycerols and elevated amounts of cardiolipins, a characteristic lipid class associated with oxidative stress, in GS phenotype. CONCLUSION: While the triggers to the activation of endoplasmic reticulum stress are still unknown, data from this study point towards a nutritional deficiency as an underlying driver of this phenotype.

Investigating the Effect of an Oxytetracycline Treatment on the Gut Microbiome and Antimicrobial Resistance Gene Dynamics in Nile Tilapia (Oreochromis niloticus).

Antibiotics play a vital role in aquaculture where they are commonly used to treat bacterial diseases. However, the impact of antibiotic treatment on the gut microbiome and the development of antimicrobial resistance in Nile tilapia (Oreochromis niloticus) over time remains to be fully understood. In this study, fish were fed a single treatment of oxytetracycline (100 mg/kg/day) for eight days, followed by a 14-day withdrawal period. Changes in the distal gut microbiome were measured using 16S rRNA sequencing. In addition, the abundance of antimicrobial resistance genes was quantified using real-time qPCR methods. Overall, the gut microbiome community diversity and structure of Nile tilapia was resilient to oxytetracycline treatment. However, antibiotic treatment was associated with an enrichment in Plesiomonas, accompanied by a decline in other bacteria taxa. Oxytetracycline treatment increased the proportion of tetA in the distal gut of fish and tank biofilms of the treated group. Furthermore, the abundance of tetA along with other tetracycline resistance genes was strongly correlated with a number of microbiome members, including Plesiomonas. The findings from this study demonstrate that antibiotic treatment can exert selective pressures on the gut microbiome of fish in favour of resistant populations, which may have long-term impacts on fish health.

Stressor controllability modulates the stress response in fish.

BACKGROUND: In humans the stress response is known to be modulated to a great extent by psychological factors, particularly by the predictability and the perceived control that the subject has of the stressor. This psychological dimension of the stress response has also been demonstrated in animals phylogenetically closer to humans (i.e. mammals). However, its occurrence in fish, which represent a divergent vertebrate evolutionary lineage from that of mammals, has not been established yet, and, if present, would indicate a deep evolutionary origin of these mechanisms across vertebrates. Moreover, the fact that psychological modulation of stress is implemented in mammals by a brain cortical top-down inhibitory control over subcortical stress-responsive structures, and the absence of a brain cortex in fish, has been used as an argument against the possibility of psychological stress in fish, with implications for the assessment of fish sentience and welfare. Here, we have investigated the occurrence of psychological stress in fish by assessing how stressor controllability modulates the stress response in European seabass (Dicentrarchus labrax). RESULTS: Fish were exposed to either a controllable or an uncontrollable stressor (i.e. possibility or impossibility to escape a signaled stressor). The effect of loss of control (possibility to escape followed by impossibility to escape) was also assessed. Both behavioral and circulating cortisol data indicates that the perception of control reduces the response to the stressor, when compared to the uncontrollable situation. Losing control had the most detrimental effect. The brain activity of the teleost homologues to the sensory cortex (Dld) and hippocampus (Dlv) parallels the uncontrolled and loss of control stressors, respectively, whereas the activity of the lateral septum (Vv) homologue responds in different ways depending on the gene marker of brain activity used. CONCLUSIONS: These results suggest the psychological modulation of the stress response to be evolutionary conserved across vertebrates, despite being implemented by different brain circuits in mammals (pre-frontal cortex) and fish (Dld-Dlv).

Analysis across diverse fish species highlights no conserved transcriptome signature for proactive behaviour.

BACKGROUND: Consistent individual differences in behaviour, known as animal personalities, have been demonstrated within and across species. In fish, studies applying an animal personality approach have been used to resolve variation in physiological and molecular data suggesting a linkage, genotype-phenotype, between behaviour and transcriptome regulation. In this study, using three fish species (zebrafish; Danio rerio, Atlantic salmon; Salmo salar and European sea bass; Dicentrarchus labrax), we firstly address whether personality-specific mRNA transcript abundances are transferrable across distantly-related fish species and secondly whether a proactive transcriptome signature is conserved across all three species. RESULTS: Previous zebrafish transcriptome data was used as a foundation to produce a curated list of mRNA transcripts related to animal personality across all three species. mRNA transcript copy numbers for selected gene targets show that differential mRNA transcript abundance in the brain appears to be partially conserved across species relative to personality type. Secondly, we performed RNA-Seq using whole brains from S. salar and D. labrax scoring positively for both behavioural and molecular assays for proactive behaviour. We further enriched this dataset by incorporating a zebrafish brain transcriptome dataset specific to the proactive phenotype. Our results indicate that cross-species molecular signatures related to proactive behaviour are functionally conserved where shared functional pathways suggest that evolutionary convergence may be more important than individual mRNAs. CONCLUSIONS: Our data supports the proposition that highly polygenic clusters of genes, with small additive effects, likely support the underpinning molecular variation related to the animal personalities in the fish used in this study. The polygenic nature of the proactive brain transcriptome across all three species questions the existence of specific molecular signatures for proactive behaviour, at least at the granularity of specific regulatory gene modules, level of genes, gene networks and molecular functions.

Innovation in Nucleotide-Binding Oligomerization-Like Receptor and Toll-Like Receptor Sensing Drives the Major Histocompatibility Complex-II Free Atlantic Cod Immune System.

The absence of MHC class II antigen presentation and multiple pathogen recognition receptors in the Atlantic cod has not impaired its immune response however how underlying mechanisms have adapted remains largely unknown. In this study, ex vivo cod macrophages were challenged with various bacterial and viral microbe-associated molecular patterns (MAMP) to identify major response pathways. Cytosolic MAMP-PRR pathways based upon the NOD-like receptors (NLRs) and RIG-I-like receptors (RLRs) were identified as the critical response pathways. Our analyses suggest that internalization of exogenous ligands through scavenger receptors drives both pathways activating transcription factors like NF-kB (Nuclear factor-kappa B) and interferon regulatory factors (IRFs). Further, ligand-dependent differential expression of a unique TLR25 isoform and multiple NLR paralogues suggests (sub)neofunctionalization toward specific immune defensive strategies. Our results further demonstrate that the unique immune system of the Atlantic cod provides an unprecedented opportunity to explore the evolutionary history of PRR-based signaling in vertebrate immunity.

Polarized Trout Epithelial Cells Regulate Transepithelial Electrical Resistance, Gene Expression, and the Phosphoproteome in Response to Viral Infection.

The burden of disease is a major challenge in aquaculture production. The fish gill characterized with a large surface area and short route to the bloodstream is a major environmental interface and a significant portal of entry for pathogens. To investigate gill responses to viral infection the salmonid gill cell line RTgill-W1 was stimulated with synthetic dsRNA and the salmonid alphavirus subtype 2 (SAV-2). Epithelial integrity in polarized cells can be measured as transepithelial electrical resistance (TEER) which is defined as the electrical resistance across a cell monolayer. TEER is a widely accepted quantitative measure of cellular integrity of a cell monolayer. TEER increased immediately after stimulation with the synthetic dsRNA, polyinosinic:polycytidylic acid (poly(I:C)). In parallel, tight junction and gene expression of innate immune activation markers was modulated in response to poly(I:C). The SAV-2 virus was found to replicate at a low level in RTgill-W1 cells where TEER was disturbed at an early stage of infection, however, gene expression related to tight junction regulation was not modulated. A strong poly(I:C)-driven antiviral response was observed including increases of Rig-like receptors (RLRs) and interferon stimulating genes (ISGs) mRNAs. At the level of signal transduction, poly(I:C) stimulation was accompanied by the phosphorylation of 671 proteins, of which 390 were activated solely in response to the presence of poly(I:C). According to motif analysis, kinases in this group included MAPKs, Ca2+/calmodulin-dependent kinase (CaMK) and cAMP-dependent protein kinase (PKA), all reported to be activated in response to viral infection in mammals. Results also highlighted an activation of the cytoskeletal organization that could be mediated by members of the integrin family. While further work is needed to validate these results, our data indicate that salmonid gill epithelia has the ability to mount a significant response to viral infection which might be important in disease progression. In vitro cell culture can facilitate both a deeper understanding of the anti-viral response in fish and open novel therapeutic avenues for fish health management in aquaculture.

Physiological responses during acute stress recovery depend on stress coping style in European sea bass, Dicentrarchus labrax.

Individual stress coping style (reactive, intermediate and proactive) was determined in 3 groups of 120 pit tagged European seabass using the hypoxia avoidance test. The same three groups (no change in social composition) were then reared according to the standards recommended for this species. Then, 127 days later, individuals initially characterized as reactive, intermediate or proactive were submitted to an acute confinement stress for 30 min. Blood samples were taken to measure plasma cortisol levels 30 min (Stress30) or 150 min (Stress150) after the end of the confinement stress. Individuals were then sacrificed to sample the telencephalon in order to measure the main monoamines and their catabolites (at Stress30 only). Individuals from Stress150 were sampled for whole brain for a transcriptomic analysis. The main results showed that reactive individuals had a lower body mass than intermediate individuals which did not differ from proactive individuals. The physiological cortisol response did not differ between coping style at Stress30 but at Stress150 when intermediate and proactive individuals had recovered pre stress levels, reactive individuals showed a significant higher level illustrating a modulation of stress recovery by coping style. Serotonin turnover ratio was higher in proactive and reactive individuals compared to intermediate individuals and a significant positive correlation was observed with cortisol levels whatever the coping style. Further, the confinement stress led to a general increase in the serotonin turnover comparable between coping styles. Stress150 had a significant effect on target mRNA copy number (Gapdh mRNA copy number decreased while ifrd1 mRNA copy number increased) and such changes tended to depend upon coping style.

Plasma proteome profiling of freshwater and seawater life stages of rainbow trout (Oncorhynchus mykiss).

The sea-run phenotype of rainbow trout (Oncorhynchus mykiss), like other anadromous salmonids, present a juvenile stage fully adapted to life in freshwater known as parr. Development in freshwater is followed by the smolt stage, where preadaptations needed for seawater life are developed making fish ready to migrate to the ocean, after which event they become post-smolts. While these three life stages have been studied using a variety of approaches, proteomics has never been used for such purpose. The present study characterised the blood plasma proteome of parr, smolt and post-smolt rainbow trout using a gel electrophoresis liquid chromatography tandem mass spectrometry approach alone or in combination with low-abundant protein enrichment technology (combinatorial peptide ligand library). In total, 1,822 proteins were quantified, 17.95% of them being detected only in plasma post enrichment. Across all life stages, the most abundant proteins were ankyrin-2, DNA primase large subunit, actin, serum albumin, apolipoproteins, hemoglobin subunits, hemopexin-like proteins and complement C3. When comparing the different life stages, 17 proteins involved in mechanisms to cope with hyperosmotic stress and retinal changes, as well as the downregulation of nonessential processes in smolts, were significantly different between parr and smolt samples. On the other hand, 11 proteins related to increased growth in post-smolts, and also related to coping with hyperosmotic stress and to retinal changes, were significantly different between smolt and post-smolt samples. Overall, this study presents a series of proteins with the potential to complement current seawater-readiness assessment tests in rainbow trout, which can be measured non-lethally in an easily accessible biofluid. Furthermore, this study represents a first in-depth characterisation of the rainbow trout blood plasma proteome, having considered three life stages of the fish and used both fractionation alone or in combination with enrichment methods to increase protein detection.

Linking stress coping styles with brain mRNA abundance of selected transcripts for Senegalese sole (Solea senegalensis) juveniles.

In fish, proactive and reactive individual stress copying styles (SCS) have been used to resolve variation in molecular expression data. Stress coping styles have been previously described in several stages of Solea senegalensis by validating for the species the use of standard behavioural screening tests. The present study aimed to link behavioural SCS tests with brain transcript abundance in early Senegalese sole juveniles in order to observe the natural variation in a molecular pathway in this species. A total of 50 juveniles were subjected to three individual behavioural (Restraining, New environment and Confinement) and one group (Risk-taking) screening tests. The fish were classified in SCS categories by applying a hierarchical cluster to the variable "Total activity" (the total activity time that the fish was moving in each individual test). Three categories were defined, proactive, intermediate and reactive sole. Six transcripts were chosen and tested, one related to basic metabolism (gapdh-2), three to feeding behaviour (per1, igf-Ia, pparß) and two to the stress response (crh-BP and hsp90aa) in 30 juveniles (10 individuals per SCS category) using rt-qPCR to observe differences in the abundance of those transcripts among SCS. Four transcripts were differentially expressed (DETs) among them. The transcript gapdh-2 showed up-regulation for proactive and intermediate SCS sole while reactive individuals showed down-regulation. Target mRNAs per1, igf-Ia and pparß, showed different levels of up-regulation for proactive and reactive fish while intermediates were highly down-regulated. Surprisingly no differences in stress related transcripts were observed. Correlations were found between variation in coping styles and variation in the abundance of mRNAs involved in important biological functions in Senegalese sole. These results are the first evidence of the relationship between the behavioural individual variation and the fluctuation in brain transcripts abundance in Senegalese sole.

'Love at first sight': The effect of personality and colouration patterns in the reproductive success of zebrafish (Danio rerio).

Individual differences in animal personality and external appearance such as colouration patterns have both been extensively studied separately. A significant body of research has explored many of pertinent ecological and biological aspects that can be affected by them and their impact upon fitness. Currently little is known about how both factors interact and their effect on reproductive success. In this study, we evaluated two major parameters contributing to the fitness of the species: reproduction and offspring survival. We selected two different phenotypes of the zebrafish (Danio rerio) selected by their colouration patterns: phenotype 1) named Wild type, UAB line (WT-UAB) with a homogeneous colouration pattern (clear and defined lateral stripes) and phenotype 2) Wild type indefinite (WT-I) had a heterogeneous colouration pattern and different degrees of lateral stripe definition. All animals were also screened for personality. We then compared their reproductive success (spawning rate) and offspring survival at different stages, from egg to larvae, and for 2 successive generations (parental generation was G0; First and second generations were G1 and G2 respectively). Our results show that personality traits were the main source of variability between the fitness components measured: both personalities had similar total numbers of eggs spawned but proactive animals, for both colour phenotypes, had higher reproductive success. This was reflected in a higher percentage of spawning viability at 1day post fertilization (dpf), higher total survival and growth rates at larval stages. Proactive phenotypes from WT-UAB population had a higher overall fitness in comparison to the other phenotypes studied. Our findings imply that fitness of this species when kept under similar husbandry conditions is significantly influenced by parental personality and not by their external appearance. Under these conditions the reproductive success is enhanced. The implications of this study are important for zebrafish breeding and husbandry in captivity and are relevant toward understanding the underlying drivers of trait selection in natural environments.

Behavioral Fever Drives Epigenetic Modulation of the Immune Response in Fish.

Ectotherms choose the best thermal conditions to mount a successful immune response, a phenomenon known as behavioral fever. The cumulative evidence suggests that behavioral fever impacts positively upon lymphocyte proliferation, inflammatory cytokine expression, and other immune functions. In this study, we have explored how thermal choice during infection impacts upon underpinning molecular processes and how temperature increase is coupled to the immune response. Our results show that behavioral fever results in a widespread, plastic imprint on gene regulation, and lymphocyte proliferation. We further explored the possible contribution of histone modification and identified global associations between temperature and histone changes that suggest epigenetic remodeling as a result of behavioral fever. Together, these results highlight the critical importance of thermal choice in mobile ectotherms, particularly in response to an infection, and demonstrate the key role of epigenetic modification to orchestrate the thermocoupling of the immune response during behavioral fever.

The expression of TRPV channels, prostaglandin E2 and pro-inflammatory cytokines during behavioural fever in fish.

A fever, or increased body temperature, is a symptom of inflammation, which is a complex defence reaction of the organism to pathogenic infections. After pathogens enter the body, immune cells secrete a number of agents, the functions of which stimulate the body to develop a functional immune and fever response. In mammals it is known that PGE2 is the principal mediator of fever. The extent to which PGE2 and other pro-inflammatory cytokines such as TNF-α, IL-6, or IL-1ß could be involved in the induction of behavioural fever in fish remains to be clarified. Several members of the transient receptor potential (TRP) family of ion channels have been implicated as transducers of thermal stimuli, including TRPV1 and TRPV2, which are activated by heat. Here we show that members of the TRP family, TRPV1 and TRPV4, may participate in the coordination of temperature sensing during the behavioural fever. To examine the behavioral fever mechanism in Salmo salar an infection with IPNV, infectious pancreatic necrosis virus, was carried out by an immersion challenge with 10 × 105 PFU/mL-1 of IPNV. Behavioural fever impacted upon the expression levels of both TRPV1 and TRPV4 mRNAs after the viral challenge and revealed a juxtaposed regulation of TRPV channels. Our results suggest that an increase in the mRNA abundance of TRPV1 is tightly correlated with a significant elevation in the expression of pro-inflammatory cytokines (IL-1ß, IL-6, TNF-α and PGE2) in the Pre-Optic Area (POA) and cytokine release in plasma. Together, these data indicate that the reduction of TRPV4 expression during behavioural fever may contribute to the onset of behavioural fever influencing movement toward higher water temperatures. Our data also suggest an effect of TRPV channels in the regulation of behavioural fever through activation of EP3 receptors in the central nervous system by PGE2 induced by plasma-borne cytokines. These results highlight for first time in mobile ectotherms the key role of pro-inflammatory cytokines and TRPV channels in behavioural fever that likely involves a complex integration of prostaglandin induction, cytokine recognition and temperature sensing.