Reproductive success and offspring survival until sexual maturity are essential traits both for fish fitness and aquaculture development. Variation in offspring’s survival among family results in unbalanced parental contributions to the next generation and may explain the loss of genetic diversity observed in some farmed populations. Therefore, we studied the variance in parental contributions to a progeny cohort, as well as the biological factors impacting offspring early survival in rainbow trout. The data consisted of 945 individual survival observations from fertilization to the juvenile stage from 135 full-sib families of the INRAE experimental synthetic line. Survival was assessed at eyed-egg stage, hatching, and 3 weeks after first feeding. We used a full-factorial mating design to partition phenotypic variance in early survival traits into maternal and additive genetic effects under threshold GBLUP models considering the inclusion of genomic information for 32,725 SNP. Average offspring survival proportions were 91.0% at the eyed-egg stage, 87.2% at hatching, and 84.4% three weeks after first feeding. Significant unbalanced dam contributions were observed at the eyed-egg and hatching stages. Low heritability was estimated for early survival traits (h²=0.20 ± 0.12 and 0.13 ± 0.09 for survival from egg-eyed stage and, respectively, hatching and first feeding), revealing that additive genetic variance was not significantly different from zero, while maternal effects explained a larger part (c²=0.37 ± 0.16 and 0.15 ± 0.07, respectively) of the phenotypic variances. There was no evidence of inbreeding depression on survival in our study. Phenotypically, offspring early survival was positively correlated with dam fecundity, while it was negatively correlated with dam post-spawning weight. Negative, but not significant association was observed between early survival and dam’s average egg weight. If a study of genetic correlations confirms these phenotypic trends, promoting high fecund females should help the breeders to increase offspring early survival and to maintain genetic diversity in breeding programs.

Global warming is expected to increase the frequency and intensity of heatwaves, resulting in more common combined acute hyperthermia and hypoxia conditions in fish farms. Such poor thermal and oxygenation conditions induce problems, including growth losses, increased pathogens pressure and mortality. Selective breeding is a promising solution to improve resistance to non-optimal water quality. Indeed, genetic variability to survive in acute hyperthermia or hypoxia conditions has been proved in fish. However, the characterization of these traits is not yet detailed enough to include them in a selection program. Here, we investigated the ranking stability of genotypes for acute hyperthermia or hypoxia resistances over age and between acute hyperthermia and acute hypoxia resistances. To this end, we established rankings of six isogenic lines of rainbow trout (Oncorhynchus mykiss) for their resistance to acute hyperthermia and hypoxia stress factors at 6 and 15 months. The experimental design was robust with more than a hundred fish per line and age. There were statistically significant resistance variations among lines confirming the potential of genetic selection for these traits. Hyperthermia and hypoxia resistance rankings were found stable 1 year apart for most genotypes. Therefore, it would be possible to select for resistance to hyperthermia and hypoxia at an early stage. No overall relationship was found between acute hyperthermia and hypoxia resistance traits: some lines were resistant to both stress factors while others were resistant to one but sensitive to the other. This indicates no strong antagonistic genetic effects between acute hyperthermia and hypoxia resistance traits in rainbow trout.

Infectious diseases are a major constraint on aquaculture. Genetic lines with different susceptibilities to diseases are useful models to identify resistance mechanisms to pathogens and to improve prophylaxis. Bacterial cold-water disease (BCWD) caused by Flavobacterium psychrophilum represents a major threat for freshwater salmonid farming worldwide. A collection of rainbow trout ( Oncorhynchus mykiss ) isogenic lines was previously produced from a French domestic population. Here, we compared BCWD resistance phenotypes using a subset of isogenic lines chosen for their contrasted susceptibilities to F. psychrophilum . We applied individual monitoring to document the infection process, including time-course quantification of bacteremia and innate immune response. Strikingly, BCWD resistance was correlated with a lower bacterial growth rate in blood. Several immune genes were expressed at higher levels in resistant fish regardless of infection: the Type II arginase ( arg2 ), a marker for M2 macrophages involved in anti-inflammatory responses and tissue repair, and two Toll-like receptors ( tlr2 / tlr7 ), responsible for pathogen detection and inflammatory responses. This study highlights the importance of innate and intrinsic defense mechanisms in determining the outcome of F. psychrophilum infections, and illustrates that non-lethal time-course blood sampling for individual monitoring of bacteremia is a powerful tool to resolve within-host pathogen behavior in bacterial fish diseases.

Abstract Background In response to major challenges regarding the supply and sustainability of marine ingredients in aquafeeds, the aquaculture industry has made a large-scale shift toward plant-based substitutions for fish oil and fish meal. But, this also led to lower levels of healthful n−3 long-chain polyunsaturated fatty acids (PUFAs)—especially eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids—in flesh. One potential solution is to select fish with better abilities to retain or synthesise PUFAs, to increase the efficiency of aquaculture and promote the production of healthier fish products. To this end, we aimed i) to estimate the genetic variability in fatty acid (FA) composition in visceral fat quantified by Raman spectroscopy, with respect to both individual FAs and groups under a feeding regime with limited n-3 PUFAs; ii) to study the genetic and phenotypic correlations between FAs and processing yields- and fat-related traits; iii) to detect QTLs associated with FA composition and identify candidate genes; and iv) to assess the efficiency of genomic selection compared to pedigree-based BLUP selection. Results Proportions of the various FAs in fish were indirectly estimated using Raman scattering spectroscopy. Fish were genotyped using the 57 K SNP Axiom™ Trout Genotyping Array. Following quality control, the final analysis contained 29,652 SNPs from 1382 fish. Heritability estimates for traits ranged from 0.03 ± 0.03 (n-3 PUFAs) to 0.24 ± 0.05 (n-6 PUFAs), confirming the potential for genomic selection. n-3 PUFAs are positively correlated to a decrease in fat deposition in the fillet and in the viscera but negatively correlated to body weight. This highlights the potential interest to combine selection on FA and against fat deposition to improve nutritional merit of aquaculture products. Several QTLs were identified for FA composition, containing multiple candidate genes with indirect links to FA metabolism. In particular, one region on Omy1 was associated with n-6 PUFAs, monounsaturated FAs, linoleic acid, and EPA, while a region on Omy7 had effects on n-6 PUFAs, EPA, and linoleic acid. When we compared the effectiveness of breeding programmes based on genomic selection (using a reference population of 1000 individuals related to selection candidates) or on pedigree-based selection, we found that the former yielded increases in selection accuracy of 12 to 120% depending on the FA trait. Conclusion This study reveals the polygenic genetic architecture for FA composition in rainbow trout and confirms that genomic selection has potential to improve EPA and DHA proportions in aquaculture species.

One of the top priorities of the aquaculture industry is the genetic improvement of economically important traits in fish, such as those related to processing and quality. However, the accuracy of genetic evaluations has been hindered by a lack of data on such traits from a sufficiently large population of animals. The objectives of this study were thus threefold: (i) to estimate genetic parameters of growth-, yield-, and quality-related traits in rainbow trout (Oncorhynchus mykiss) using three different phenotyping technologies [invasive and non-invasive: microwave-based, digital image analysis, and magnetic resonance imaging (MRI)], (ii) to detect quantitative trait loci (QTLs) associated with these traits, and (iii) to identify candidate genes present within these QTL regions. Our study collected data from 1,379 fish on growth, yield-related traits (body weight, condition coefficient, head yield, carcass yield, headless gutted carcass yield), and quality-related traits (total fat, percentage of fat in subcutaneous adipose tissue, percentage of fat in flesh, flesh colour); genotypic data were then obtained for all fish using the 57K SNP Axiom® Trout Genotyping array. Heritability estimates for most of the 14 traits examined were moderate to strong, varying from 0.12 to 0.67. Most traits were clearly polygenic, but our genome-wide association studies (GWASs) identified two genomic regions on chromosome 8 that explained up to 10% of the genetic variance (cumulative effects of two QTLs) for several traits (weight, condition coefficient, subcutaneous and total fat content, carcass and headless gutted carcass yields). For flesh colour traits, six QTLs explained 1–4% of the genetic variance. Within these regions, we identified several genes (htr1, gnpat, ephx1, bcmo1, and cyp2x) that have been implicated in adipogenesis or carotenoid metabolism, and thus represent good candidates for further functional validation. Finally, of the three techniques used for phenotyping, MRI demonstrated particular promise for measurements of fat content and distribution, while the digital image analysis-based approach was very useful in quantifying colour-related traits. This work provides new insights that may aid the development of commercial breeding programmes in rainbow trout, specifically with regard to the genetic improvement of yield and flesh-quality traits as well as the use of invasive and/or non-invasive technologies to predict such traits.

Rainbow trout has a male heterogametic (XY) sex determination system controlled by a major sex-determining gene, sdY. Unexpectedly, a few phenotypically masculinised fish are regularly observed in all-female farmed trout stocks. To better understand the genetic determinism underlying spontaneous maleness in XX-rainbow trout, we recorded the phenotypic sex of 20,210 XX-rainbow trout from a French farm population at 10 and 15 months post-hatching. The overall masculinisation rate was 1.45%. We performed two genome-wide association studies (GWAS) on a subsample of 1139 individuals classified as females, intersex or males using either medium-throughput genotyping (31,811 SNPs) or whole-genome sequencing (WGS, 8.7 million SNPs). The genomic heritability of maleness ranged between 0.48 and 0.62 depending on the method and the number of SNPs used for the estimation. At the 31K SNPs level, we detected four QTL on three chromosomes (Omy1, Omy12 and Omy20). Using WGS information, we narrowed down the positions of the two QTL detected on Omy1 to 96 kb and 347 kb respectively, with the second QTL explaining up to 14% of the total genetic variance of maleness. Within this QTL, we detected three putative candidate genes, fgfa8, cyp17a1 and an uncharacterised protein (LOC110527930), which might be involved in spontaneous maleness of XX-female rainbow trout.

Phenotypic plasticity is a key component of the ability of organisms to respond to changing environmental conditions. In this study, we aimed to study the establishment of DNA methylation marks in response to an environmental stress in rainbow trout and to assess whether these marks depend on the genetic background. The environmental stress chosen here was temperature, a known induction factor of epigenetic marks in fish. To disentangle the role of epigenetic mechanisms such as DNA methylation in generating phenotypic variations, nine rainbow trout isogenic lines with no genetic variability within a line were used. For each line, half of the eggs were incubated at standard temperature (11°C) and the other half at high temperature (16°C), from eyed-stage to hatching. In order to gain a first insight into the establishment of DNA methylation marks in response to an early temperature regime (control 11°C vs. heated 16°C), we have studied the expression of 8 dnmt3 (DNA methyltransferase) genes, potentially involved in de novo methylation, and analysed global DNA methylation in the different rainbow trout isogenic lines using LUMA (LUminometric Methylation Assay). Finally, finer investigation of genome-wide methylation patterns was performed using EpiRADseq, a reduced-representation library approach based on the ddRADseq (Double Digest Restriction Associated DNA) protocol, for six rainbow trout isogenic lines. We have demonstrated that thermal history during embryonic development alters patterns of DNA methylation, but to a greater or lesser extent depending on the genetic background.

Infectious diseases represent a major threat for the sustainable development of fish farming. Efficient vaccines are not available against all diseases, and growing antibiotics resistance limits the use of antimicrobial drugs in aquaculture. It is therefore important to understand the basis of fish natural resistance to infections to help genetic selection and to develop new approaches against infectious diseases. However, the identification of the main mechanisms determining the resistance or susceptibility of a host to a pathogenic microbe is challenging, integrating the complexity of the variation of host genetics, the variability of pathogens, and their capacity of fast evolution and adaptation. Multiple approaches have been used for this purpose: (i) genetic approaches, QTL (quantitative trait loci) mapping or GWAS (genome-wide association study) analysis, to dissect the genetic architecture of disease resistance, and (ii) transcriptomics and functional assays to link the genetic constitution of a fish to the molecular mechanisms involved in its interactions with pathogens. To date, many studies in a wide range of fish species have investigated the genetic determinism of resistance to many diseases using QTL mapping or GWAS analyses. A few of these studies pointed mainly toward adaptive mechanisms of resistance/susceptibility to infections; others pointed toward innate or intrinsic mechanisms. However, in the majority of studies, underlying mechanisms remain unknown. By comparing gene expression profiles between resistant and susceptible genetic backgrounds, transcriptomics studies have contributed to build a framework of gene pathways determining fish responsiveness to a number of pathogens. Adding functional assays to expression and genetic approaches has led to a better understanding of resistance mechanisms in some cases. The development of knock-out approaches will complement these analyses and help to validate putative candidate genes critical for resistance to infections. In this review, we highlight fish isogenic lines as a unique biological material to unravel the complexity of host response to different pathogens. In the future, combining multiple approaches will lead to a better understanding of the dynamics of interaction between the pathogen and the host immune response, and contribute to the identification of potential targets of selection for improved resistance.

BackgroundSelective breeding is a relatively recent practice in aquaculture species compared to terrestrial livestock. Nevertheless, the genetic variability of farmed salmonid lines, which have been selected for several generations, should be assessed. Indeed, a significant decrease in genetic variability due to high selection intensity could have occurred, potentially jeopardizing the long-term genetic progress as well as the adaptive capacities of populations facing change(s) in the environment. Thus, it is important to evaluate the impact of selection practices on genetic diversity to limit future inbreeding. The current study presents an analysis of genetic diversity within and between six French rainbow trout (Oncorhynchus mykiss) experimental or commercial lines based on a medium-density single nucleotide polymorphism (SNP) chip and various molecular genetic indicators: fixation index (F-ST), linkage disequilibrium (LD), effective population size (N-e) and inbreeding coefficient derived from runs of homozygosity (ROH).ResultsOur results showed a moderate level of genetic differentiation between selected lines (F-ST ranging from 0.08 to 0.15). LD declined rapidly over the first 100kb, but then remained quite high at long distances, leading to low estimates of N-e in the last generation ranging from 24 to 68 depending on the line and methodology considered. These results were consistent with inbreeding estimates that varied from 10.0% in an unselected experimental line to 19.5% in a commercial line, and which are clearly higher than corresponding estimates in ruminants or pigs. In addition, strong variations in LD and inbreeding were observed along the genome that may be due to differences in local rates of recombination or due to key genes that tended to have fixed favorable alleles for domestication or production.ConclusionsThis is the first report on ROH for any aquaculture species. Inbreeding appeared to be moderate to high in the six French rainbow trout lines, due to founder effects at the start of the breeding programs, but also likely to sweepstakes reproductive success in addition to selection for the selected lines. Efficient management of inbreeding is a major goal in breeding programs to ensure that populations can adapt to future breeding objectives and SNP information can be used to manage the rate at which inbreeding builds up in the fish genome.

We aim to investigate the potential role of epigenetic marks in the expression of phenotypes and their variability in fish, in particular genetic variability of epigenetic marks in response to an environmental stress. In this context, rainbow trout isogenic lines are the material of choice. Within each line, all fish have the same genome i.e. there is no genetic variability. This allows the comparison of epigenetic marks among several individuals with the same genotype. The environmental stress chosen here is temperature because several studies have reported genetic determinism of thermotolerance. Moreover, rainbow trout isogenic lines have recently been characterized for their response to temperature and the existence of a high between-line variability was shown. The aim of this study is to contribute to the understanding of why certain lines are more tolerant to temperature stress than others, by investigating the implication of epigenetic mechanisms in the variability of the response to temperature. More specifically, the objective of this study was to test whether temperature regime experienced during early development leads to epigenetic modifications within and between lines. Nine rainbow trout isogenic lines were chosen. For each line, half of the eggs were incubated at standard temperature (12°C) and the other half at high temperature (16°C), from eyed-stage to hatching. At eyed-stage just before hatching, analysis of global DNA methylation was performed with LUMA (LUminometric Methylation Assay) on 3 pools of 5 eggs per line and per incubation temperature. LUMA analysis revealed significant differences between lines but little or no effect of incubation temperature. Also, for 6 out of the 9 isogenic lines, genomewide patterns of methylation were analysed by EpiRADseq on the same biological material. EpiRADseq is a reduced-representation library-based approach which has been recently developed and tested on a single clone of water fleas. The protocol was here modified to account for genetic variability and allow both within and between-lines comparisons. Globally, very few loci were differentially methylated between the two incubation temperatures. Thus, in the future, the impact of a longer exposure to high temperatures during early development will be tested.

Background Bacterial cold-water disease, which is caused by Flavobacterium psychrophilum , is one of the major diseases that affect rainbow trout ( Oncorhynchus mykiss ) and a primary concern for trout farming. Better knowledge of the genetic basis of resistance to F. psychrophilum would help to implement this trait in selection schemes and to investigate the immune mechanisms associated with resistance. Various studies have revealed that skin and mucus may contribute to response to infection. However, previous quantitative trait loci (QTL) studies were conducted by using injection as the route of infection. Immersion challenge, which is assumed to mimic natural infection by F. psychrophilum more closely, may reveal different defence mechanisms. Results Two isogenic lines of rainbow trout with contrasting susceptibilities to F. psychrophilum were crossed to produce doubled haploid F2 progeny. Fish were infected with F. psychrophilum either by intramuscular injection (115 individuals) or by immersion (195 individuals), and genotyped for 9654 markers using RAD-sequencing. Fifteen QTL associated with resistance traits were detected and only three QTL were common between the injection and immersion. Using a model that accounted for epistatic interactions between QTL, two main types of interactions were revealed. A “compensation-like” effect was detected between several pairs of QTL for the two modes of infection. An “enhancing-like” interaction effect was detected between four pairs of QTL. Integration of the QTL results with results of a previous transcriptomic analysis of response to F. psychrophilum infection resulted in a list of potential candidate immune genes that belong to four relevant functional categories (bacterial sensors, effectors of antibacterial immunity, inflammatory factors and interferon-stimulated genes). Conclusions These results provide new insights into the genetic determinism of rainbow trout resistance to F. psychrophilum and confirm that some QTL with large effects are involved in this trait. For the first time, the role of epistatic interactions between resistance-associated QTL was evidenced. We found that the infection protocol used had an effect on the modulation of defence mechanisms and also identified relevant immune functional candidate genes.

Background: The viral hemorrhagic septicemia virus (VHSV) is a major threat for salmonid farming and for wild fish populations worldwide. Previous studies have highlighted the importance of innate factors regulated by a major quantitative trait locus (QTL) for the natural resistance to waterborne VHSV infection in rainbow trout. The aim of this study was to analyze the early transcriptomic response to VHSV inoculation in cell lines derived from previously described resistant and susceptible homozygous isogenic lines of rainbow trout to obtain insights into the molecular mechanisms responsible for the resistance to the viral infection. Results: We first confirmed the presence of the major QTL in a backcross involving a highly resistant fish isogenic line (B57) and a highly susceptible one (A22), and were able to define the confidence interval of the QTL and to identify its precise position. We extended the definition of the QTL since it controls not only resistance to waterborne infection but also the kinetics of mortality after intra-peritoneal injection. Deep sequencing of the transcriptome of B57 and A22 derived cell lines exposed to inactivated VHSV showed a stronger response to virus inoculation in the resistant background. In line with our previous observations, an early and strong induction of interferon and interferonstimulated genes was correlated with the resistance to VHSV, highlighting the major role of innate immune factors in natural trout resistance to the virus. Interestingly, major factors of the antiviral innate immunity were much more expressed in naive B57 cells compared to naive A22 cells, which likely contributes to the ability of B57 to mount a fast antiviral response after viral infection. These observations were further extended by the identification of several innate immune-related genes localized close to the QTL area on the rainbow trout genome. Conclusions: Taken together, our results improve our knowledge in virus-host interactions in vertebrates and provide novel insights in the molecular mechanisms explaining the resistance to VHSV in rainbow trout. Our data also provide a collection of potential markers for resistance and susceptibility of rainbow trout to VHSV infection.

Better understanding of the mechanisms underlying interindividual variation in stress responses and their links with production traits is a key issue for sustainable animal breeding. In this study, we searched for quantitative trait loci (QTL) controlling the magnitude of the plasma cortisol stress response and compared them to body size traits in five F2 full-sib families issued from two rainbow trout lines divergently selected for high or low post-confinement plasma cortisol level. Approximately 1000 F2 individuals were individually tagged and exposed to two successive acute confinement challenges (1 month interval). Post-stress plasma cortisol concentrations were determined for each fish. A medium density genome scan was carried out (268 markers, overall marker spacing less than 10 cM). QTL detection was performed using qtlmap software, based on an interval mapping method (http://www.inra.fr/qtlmap). Overall, QTL of medium individual effects on cortisol responsiveness (<10% of phenotypic variance) were detected on 18 chromosomes, strongly supporting the hypothesis that control of the trait is polygenic. Although a core array of QTL controlled cortisol concentrations at both challenges, several QTL seemed challenge specific, suggesting that responses to the first and to a subsequent exposure to the confinement stressor are distinct traits sharing only part of their genetic control. Chromosomal location of the steroidogenic acute regulatory protein (STAR) makes it a good potential candidate gene for one of the QTL. Finally, comparison of body size traits QTL (weight, length and body conformation) with cortisol-associated QTL did not support evidence for negative genetic relationships between the two types of traits.