Edwige Quillet

The rainbow trout genome provides novel insights into evolution after whole-genome duplication in vertebrates

Vertebrate evolution has been shaped by several rounds of whole-genome duplications (WGDs) that are often suggested to be associated with adaptive radiations and evolutionary innovations. Due to an additional round of WGD, the rainbow trout genome offers a unique opportunity to investigate the early evolutionary fate of a duplicated vertebrate genome. Here we show that after 100 million years of evolution the two ancestral subgenomes have remained extremely collinear, despite the loss of half of the duplicated protein-coding genes, mostly through pseudogenization. In striking contrast is the fate of miRNA genes that have almost all been retained as duplicated copies. The slow and stepwise rediploidization process characterized here challenges the current hypothesis that WGD is followed by massive and rapid genomic reorganizations and gene deletions.

A Type I and Type II microsatellite linkage map of Rainbow trout (Oncorhynchus mykiss) with presumptive coverage of all chromosome arms

BackgroundThe development of large genomic resources has become a prerequisite to elucidate the wide-scale evolution of genomes and the molecular basis of complex traits. Linkage maps represent a first level of integration and utilization of such resources and the primary framework for molecular analyses of quantitative traits. Previously published linkage maps have already outlined the main peculiarities of the rainbow trout meiosis and a correspondance between linkage groups and chromosome arms has been recently established using fluorescent in situ hybridization. The number of chromosome arms which were covered by these maps remained unknown.ResultsWe report an updated linkage map based on segregation analysis of more than nine hundred microsatellite markers in two doubled haploid gynogenetic lines. These markers segregated into 31 linkage groups spanning an approximate total map length of 2750 cM. Centromeres were mapped for all the linkage groups using meiogenetic lines. For each of the 31 linkage groups, the meta or acrocentric structure infered from centromere mapping was identical with those recently found with fluorescent in situ hybridization results. The present map is therefore assumed to cover the 52 chromosome arms which constitute the rainbow trout karyotype. Our data confirm the occurrence of a high interference level in this species. Homeologous regions were identified in eleven linkage groups, reflecting the tetraploid nature of the salmonid genome. The data supported the assumption that gene orders are conserved between duplicated groups and that each group is located on a single chromosome arm. Overall, a high congruence with already published rainbow trout linkage maps was found for both gene syntenies and orders.ConclusionThis new map is likely to cover the whole set of chromosome arms and should provide a useful framework to integrate existing or forthcoming rainbow trout linkage maps and other genomic resources. Since very large numbers of EST containing microsatellite sequences are available in databases, it becomes feasible to construct high-density linkage maps localizing known genes. This will facilitate comparative mapping and, eventually, identification of candidate genes in QTL studies.

The sexually dimorphic on the Y-chromosome gene (sdY) is a conserved male-specific Y-chromosome sequence in many salmonids

All salmonid species investigated to date have been characterized with a male heterogametic sex‐determination system. However, as these species do not share any Y‐chromosome conserved synteny, there remains a debate on whether they share a common master sex‐determining gene. In this study, we investigated the extent of conservation and evolution of the rainbow trout (Oncorhynchus mykiss) master sex‐determining gene, sdY (sexually dimorphic on the Y‐chromosome), in 15 different species of salmonids. We found that the sdY sequence is highly conserved in all salmonids and that sdY is a male‐specific Y‐chromosome gene in the majority of these species. These findings demonstrate that most salmonids share a conserved sex‐determining locus and also strongly suggest that sdY may be this conserved master sex‐determining gene. However, in two whitefish species (subfamily Coregoninae), sdY was found both in males and females, suggesting that alternative sex‐determination systems may have also evolved in this family. Based on the wide conservation of sdY as a male‐specific Y‐chromosome gene, efficient and easy molecular sexing techniques can now be developed that will be of great interest for studying these economically and environmentally important species.

The application of microsatellite markers to breeding programmes in the sea bass, Dicentrarchus labrax

Microsatellite markers isolated from a genomic library of sea bass, Dicentrarchus labrax, were used to assess parentage in a breeding programme. The progenies were obtained from a complete factorial design between three males and three females, and mass-reared in a single tank. Three samples of 329, 97 and 360 larvae respectively aged 11, 40 and 116 days-old, were genotyped with microsatellite loci. The parentage of all individuals was determined unambiguously with only two microsatellite loci. A genetic analysis of quantitative characters (survival, body weight, length and malformation, condition coefficient) was realized on the larvae identified in this way. Each of the families appeared significantly different for the analyzed traits. We argue that microsatellites will be a valuable tool in: (i) breed identification (ii) family selection programs in which genetic tagging will allow different genotypes to be reared together, thus greatly reducing the impact of environmental variance and the number of replicate needed ponds.

Phenotypic and Functional Similarity of Gut Intraepithelial and Systemic T Cells in a Teleost Fish

Gut-associated lymphocytes were described in fish, but their involvement in immune responses is still unknown. In rainbow trout, intraepithelial lymphocytes (IELs) are scattered between gut epithelial cells, but neither Peyer’s patches nor mesenteric lymph nodes were identified. Rainbow trout IELs contain mainly T cells, because they expressed transcripts of T cell marker homologs of CD8, CD4, CD28, CD3ε, TCRζ, TCRγ, and TCRβ and lacked IgM. However, trout IELs did not show specific homing to the gut mucosa, which in mammals defines IELs as a distinctive mucosal population. A detailed analysis of the TCRβ repertoire of rainbow trout IELs was performed in both naive and virus-infected animals. TCRβ transcripts of rainbow trout IELs were highly diverse and polyclonal in adult naive individuals, in sharp contrast with the restricted diversity of IEL oligoclonal repertoires described in birds and mammals. Significant modifications of the trout IEL TCRβ repertoire were observed after a systemic infection with a fish rhabdovirus and were especially marked for Vβ4-bearing receptors as previously reported for spleen cells. Thus, we could not find any specific properties of the trout IEL TCRβ repertoire compared with the spleen and pronephros TCRβ repertoire, which questions the reality of a distinct IEL compartment in teleosts. Our findings suggest that a highly diversified αβ ΤCR repertoire is maintained in fish IELs in the absence of Peyer’s patches and mesenteric lymph nodes, whereas the restricted diversity of mouse αβ IELs is attributed to multiple cycles of activation and recirculation, allowing a progressive narrowing of the repertoire.

Teleost fish mount complex clonal IgM and IgT responses in spleen upon systemic viral infection

Upon infection, B-lymphocytes expressing antibodies specific for the intruding pathogen develop clonal responses triggered by pathogen recognition via the B-cell receptor. The constant region of antibodies produced by such responding clones dictates their functional properties. In teleost fish, the clonal structure of B-cell responses and the respective contribution of the three isotypes IgM, IgD and IgT remain unknown. The expression of IgM and IgT are mutually exclusive, leading to the existence of two B-cell subsets expressing either both IgM and IgD or only IgT. Here, we undertook a comprehensive analysis of the variable heavy chain (VH) domain repertoires of the IgM, IgD and IgT in spleen of homozygous isogenic rainbow trout (Onchorhynchus mykiss) before, and after challenge with a rhabdovirus, the Viral Hemorrhagic Septicemia Virus (VHSV), using CDR3-length spectratyping and pyrosequencing of immunoglobulin (Ig) transcripts. In healthy fish, we observed distinct repertoires for IgM, IgD and IgT, respectively, with a few amplified μ and τ junctions, suggesting the presence of IgM- and IgT-secreting cells in the spleen. In infected animals, we detected complex and highly diverse IgM responses involving all VH subgroups, and dominated by a few large public and private clones. A lower number of robust clonal responses involving only a few VH were detected for the mucosal IgT, indicating that both IgM(+) and IgT(+) spleen B cells responded to systemic infection but at different degrees. In contrast, the IgD response to the infection was faint. Although fish IgD and IgT present different structural features and evolutionary origin compared to mammalian IgD and IgA, respectively, their implication in the B-cell response evokes these mouse and human counterparts. Thus, it appears that the general properties of antibody responses were already in place in common ancestors of fish and mammals, and were globally conserved during evolution with possible functional convergences.

Thermal induction of gynogenesis and triploidy in Atlantic salmon (Salmo salar) and their potential interest for aquaculture

Abstract Triploid and gynogenetic progenies were produced in Atlantic salmon with moderate thermal shocks (26–29°C) applied soon after fertilization. Yields of gynogenetics were high (60–70% of control at start feeding); gynogenetics proved to be all female and could be helpful in the establishment of populations of sex-reversed females to produce all-female livestock in that species. All-triploid groups were also obtained with a wide range of thermal treatments and good survival (66–89% of control at start feeding). Nevertheless, at the age of 2.5 years, triploids displayed lower survival and growth in weight than their control, which may limit their practical interest for aquaculture.

A synthetic rainbow trout linkage map provides new insights into the salmonid whole genome duplication and the conservation of synteny among teleosts

BackgroundRainbow trout is an economically important fish and a suitable experimental organism in many fields of biology including genome evolution, owing to the occurrence of a salmonid specific whole-genome duplication (4th WGD). Rainbow trout is among some of the most studied teleosts and has benefited from substantial efforts to develop genomic resources (e.g., linkage maps. Here, we first generated a synthetic map by merging segregation data files derived from three independent linkage maps. Then, we used it to evaluate genome conservation between rainbow trout and three teleost models, medaka, stickleback and zebrafish and to further investigate the extent of the 4th WGD in trout genome.ResultsThe INRA linkage map was updated by adding 211 new markers. After standardization of marker names, consistency of marker assignment to linkage groups and marker orders was checked across the three different data sets and only loci showing consistent location over all or almost all of the data sets were kept. This resulted in a synthetic map consisting of 2226 markers and 29 linkage groups spanning over 3600 cM. Blastn searches against medaka, stickleback, and zebrafish genomic databases resulted in 778, 824 and 730 significant hits respectively while blastx searches yielded 505, 513 and 510 significant hits. Homology search results revealed that, for most rainbow trout chromosomes, large syntenic regions encompassing nearly whole chromosome arms have been conserved between rainbow trout and its closest models, medaka and stickleback. Large conserved syntenies were also found between the genomes of rainbow trout and the reconstructed teleost ancestor. These syntenies consolidated the known homeologous affinities between rainbow trout chromosomes due to the 4th WGD and suggested new ones.ConclusionsThe synthetic map constructed herein further highlights the stability of the teleost genome over long evolutionary time scales. This map can be easily extended by incorporating new data sets and should help future rainbow trout whole genome sequence assembly. Finally, the persistence of large conserved syntenies across teleosts should facilitate the identification of candidate genes through comparative mapping, even if the occurrence of intra-chromosomal micro-rearrangement may hinder the accurate prediction their genomic location.

Production of viable hybrids in salmonids by triploidization

Rainbow trout eggs were fertilized with milt collected from various salmanids (brown trout Salmo trutta ; brook trout Salvelinus lontinaliv ; coho salmon Oncorhynchus kisutch) and heat-shocked few minutes later in order to prevent the second polar body extrusion. This operation resulted in high survival rates after 161 days : respectively 49.2 p. 100, 66.2 p. 100 and 12.5 p. 100 of the diploid rainbow trout control (basis 100 p. 100) ; the triploid hybrid constitution of these fish was ascertained by karyological and biochemical investigations.

Early development and survival in brown trout (Salmo trutta fario L.): indirect effects of selection for growth rate and estimation of genetic parameters

Abstract Some components of the response to selection for growth in the PROSPER program on brown trout (Salmo trutta fario L.) (+30% in three generations) were investigated. A correlated response on growth during yolk-sac resorption was looked for, and we estimated the genetic parameters on growth-related traits at the end of endogenous feeding. For this purpose, a diallel cross between selected and control fish was designed, including 200 full-sib families (20 males×10 females), each of them being represented by four individually recorded fish. A positive-correlated effect of selection was seen on survival and percent weight gain at swim-up, and a negative one was shown on dry weight and dry matter content. No effect was seen on final wet weight. A putative interpretation is that selected individuals have a higher development rate, inducing a higher consumption of yolk reserves by the end of the experiment, and therefore, a decrease in wet weight, dry weight and dry matter content. Significant sire heritabilities were seen for final wet weight and percent weight gain (hs2=0.28±0.13 in both cases). For all traits except percent weight gain, maternal effects were strong, and an important part of them was related to the initial weight of eggs. Significant dominance effects were reported for wet weight, dry weight and percent weight gain.