José M. Yáñez

Genetics and genomics of disease resistance in salmonid species.

Infectious and parasitic diseases generate large economic losses in salmon farming. A feasible and sustainable alternative to prevent disease outbreaks may be represented by genetic improvement for disease resistance. To include disease resistance into the breeding goal, prior knowledge of the levels of genetic variation for these traits is required. Furthermore, the information from the genetic architecture and molecular factors involved in resistance against diseases may be used to accelerate the genetic progress for these traits. In this regard, marker assisted selection and genomic selection are approaches which incorporate molecular information to increase the accuracy when predicting the genetic merit of selection candidates. In this article we review and discuss key aspects related to disease resistance in salmonid species, from both a genetic and genomic perspective, with emphasis in the applicability of disease resistance traits into breeding programs in salmonids.

Genomewide single nucleotide polymorphism discovery in Atlantic salmon (Salmo salar ) : validation in wild and farmed American and European populations

A considerable number of single nucleotide polymorphisms (SNPs) are required to elucidate genotype–phenotype associations and determine the molecular basis of important traits. In this work, we carried out de novo SNP discovery accounting for both genome duplication and genetic variation from American and European salmon populations. A total of 9 736 473 nonredundant SNPs were identified across a set of 20 fish by whole‐genome sequencing. After applying six bioinformatic filtering steps, 200 K SNPs were selected to develop an Affymetrix Axiom® myDesign Custom Array. This array was used to genotype 480 fish representing wild and farmed salmon from Europe, North America and Chile. A total of 159 099 (79.6%) SNPs were validated as high quality based on clustering properties. A total of 151 509 validated SNPs showed a unique position in the genome. When comparing these SNPs against 238 572 markers currently available in two other Atlantic salmon arrays, only 4.6% of the SNP overlapped with the panel developed in this study. This novel high‐density SNP panel will be very useful for the dissection of economically and ecologically relevant traits, enhancing breeding programmes through genomic selection as well as supporting genetic studies in both wild and farmed populations of Atlantic salmon using high‐resolution genomewide information.

Genome-wide association analysis reveals loci associated with resistance against Piscirickettsia salmonis in two Atlantic salmon (Salmo salar L.) chromosomes

BackgroundPisciricketssia salmonis is the causal agent of Salmon Rickettsial Syndrome (SRS), which affects salmon species and causes severe economic losses. Selective breeding for disease resistance represents one approach for controlling SRS in farmed Atlantic salmon. Knowledge concerning the architecture of the resistance trait is needed before deciding on the most appropriate approach to enhance artificial selection for P. salmonis resistance in Atlantic salmon. The purpose of the study was to dissect the genetic variation in the resistance to this pathogen in Atlantic salmon.Methods2,601 Atlantic salmon smolts were experimentally challenged against P. salmonis by means of intra-peritoneal injection. These smolts were the progeny of 40 sires and 118 dams from a Chilean breeding population. Mortalities were recorded daily and the experiment ended at day 40 post-inoculation. Fish were genotyped using a 50K Affymetrix® Axiom® myDesignTM Single Nucleotide Polymorphism (SNP) Genotyping Array. A Genome Wide Association Analysis was performed on data from the challenged fish. Linear regression and logistic regression models were tested.ResultsGenome Wide Association Analysis indicated that resistance to P. salmonis is a moderately polygenic trait. There were five SNPs in chromosomes Ssa01 and Ssa17 significantly associated with the traits analysed. The proportion of the phenotypic variance explained by each marker is small, ranging from 0.007 to 0.045. Candidate genes including interleukin receptors and fucosyltransferase have been found to be physically linked with these genetic markers and may play an important role in the differential immune response against this pathogen.ConclusionsDue to the small amount of variance explained by each significant marker we conclude that genetic resistance to this pathogen can be more efficiently improved with the implementation of genetic evaluations incorporating genotype information from a dense SNP array.

Genomics in aquaculture to better understand species biology and accelerate genetic progress.

The production of fish and shellfish through aquaculture is an increasingly important source of high-quality animal protein, with a worldwide production of 66.6 million tons in 2012 (FAO, 2014). Considering the continuously growing global human population and increasing demand for fish products, improvements in the scale, efficiency, and sustainability of aquaculture are essential. To achieve this, several challenges facing the culture of fish and shellfish species need to be overcome. These relate to the diverse biology of the cultured species and their interaction with environmental factors. Examples include outbreaks of infectious diseases, control of sexual maturation, sustainable feed for carnivorous species, and tolerance of diverse and changing environments. This “Frontiers in Livestock Genomics” Research Topic highlights the opportunities offered by recent developments in the field of genomics, and in particular high-throughput sequencing, to contribute to addressing these challenges, with a focus on selective breeding programmes. The use of selective breeding as a tool to improve the biological efficiency of production in aquaculture generally lags behind plant and farm animal industries, and less than 10% of aquaculture production is based on genetically-improved stocks (Gjedrem et al., 2012). Encouragingly, annual genetic gains reported for aquatic species are in general substantially higher than that of terrestrial farm animals (Gjedrem et al., 2012) and there is considerable scope for achieving significant positive economic impact via improved breeding schemes. However, the status of breeding programs and the level of technology used for aquatic species production are wide-ranging, from use of wild seed stocks through to family-based selection incorporating genomic tools. Family selection and genomic tools can be applied to improve traits that are expensive or difficult to measure on the selection candidates themselves including disease resistance (Yanez et al., 2014; Odegard et al., 2014), flesh color (Colihueque and Araneda, 2014; Odegard et al., 2014) and other appearance traits such as body shape and skin pigmentation (Colihueque and Araneda, 2014) in finfish species. In contrast, despite the global importance of mollusc species for aquaculture, few selective breeding programmes exist and the state of genomic tools and knowledge for these species is typically lacking (Astorga, 2014). Genomics resources such as whole genome reference sequences, high-density SNP genotyping arrays and genotyping-by-sequencing are in development for several aquaculture species. Fuller characterisation of these resources is underway and is resulting in improved fundamental knowledge of the genome structure and biology, highlighted in this issue by the analysis of repeat elements in the Asian sea bass genome (Kuznetsova et al., 2014). These resources will provide powerful tools for the research community and will aid in the determination of the genetic factors involved in the regulation of complex traits. For example, high-throughput RNA sequencing can give a holistic view of the host response to infectious diseases, and help identify the important genes and pathways defining genetic resistance, as demonstrated in this issue for rainbow trout (Ali et al., 2014; Marancik et al., 2014) and panaeid shrimp (Santos et al., 2014). Sequencing technology has also facilitated the development of abundant genetic markers that have multi-faceted applications for selective breeding of aquatic species, including parentage assignment in mixed-family environments, providing greater control over family representation and inbreeding (Vandeputte and Haffray, 2014). Medium or high-density SNP arrays can be used to predict genomic breeding values for economically-important traits in well-developed breeding programmes, such as Atlantic salmon (Odegard et al., 2014). For instance, based on simulations of a Pacific white shrimp breeding program, genetic progress of disease resistance traits is faster with genomic-enabled selection compared to conventional phenotype-based selection due to higher accuracy (Castillo-Juarez et al., 2015). Incorporation of genetic marker information can also be a useful asset to optimize genetic diversity and future genetic gain when establishing base populations for breeding programmes (Fernandez et al., 2014). Furthermore, these genomic tools can be applied to investigate putative genomic signatures of selection during the domestication process of farmed fish species, thus potentially identifying genomic regions underlying variation in relevant phenotypes in wild and domestic fish populations (Lopez et al., 2015). Aquaculture species typically have several common features, for example high fecundity and external fertilization, plus a short evolutionary distance from their wild ancestors. The reproductive features enable flexible mating structures to be used for breeding programmes, and can provide a powerful resource for genetic studies of complex traits, such as disease resistance (Yanez et al., 2014). However, the diversity between these species is enormous and often necessitates the establishment of species-specific reproduction and breeding programmes. For example, there is a remarkable variety of sex-determination systems within aquatic farmed species, and the study of Martinez et al. (2014) highlights various methods of controlling sex ratio with aquaculture breeding programmes. This species diversity also presents an issue for choosing suitable model organisms to inform on the biology of the farmed species of interest. Model finfish species, such as zebrafish, have been well-characterized and Ulloa et al. (2014) highlight their utility for the evaluation of the response to alternative diets. However, due to the vast evolutionary distance between certain farmed aquatic and model species, it is clear that direct research on the species of interest can often be the most feasible and informative. The aquaculture industry has often been innovative and visionary in their application of new technologies to improve production. Genomics present another major opportunity, and the research published in this special issue provides several excellent examples of their potential or realized application. Using genomic tools to more effectively utilize genetic variation in economically-important traits via sustainable breeding programmes is paramount to the continued successful growth and stability of aquaculture production.

Functional Annotation of All Salmonid Genomes (FAASG): An international initiative supporting future salmonid research, conservation and aquaculture

We describe an emerging initiative - the ‘Functional Annotation of All Salmonid Genomes’ (FAASG), which will leverage the extensive trait diversity that has evolved since a whole genome duplication event in the salmonid ancestor, to develop an integrative understanding of the functional genomic basis of phenotypic variation. The outcomes of FAASG will have diverse applications, ranging from improved understanding of genome evolution, to improving the efficiency and sustainability of aquaculture production, supporting the future of fundamental and applied research in an iconic fish lineage of major societal importance.

Evidence of recent signatures of selection during domestication in an Atlantic salmon population

Selective breeding practices in Atlantic salmon aquaculture have been carried out intensively since the 1970s. Along with the phenotypic improvement of fish, we expect to observe genomic regions showing evidence of selection for traits related to growth and age at sexual maturation, as well as traits involved in the domestication process. This is mainly linked to the increase in the frequency of favourable alleles at loci that affect the traits of interest in the breeding population. In this study we searched for signatures of selection in the Cermaq Atlantic salmon broodstock, a Mowi strain, which was derived from wild Norwegian populations, and is now farmed in British Columbia, Canada. A 6.5K SNP array was used to genotype 202 fish from the Cermaq population, and the genotypes were compared with four wild populations from Norway. We used three methods based on FST values to detect signatures of selection. Forty four markers showing divergence in allele frequency were identified as outliers by the three detection methods, suggesting the presence of signatures of selection in the Cermaq population relative to their wild counterparts. Markers identified as outliers are associated with molecular functions that could be related to the selection for economically important traits (e.g., growth) as well as the domestication process (e.g., response to pathogens and environmental stressors). Of particular interest were three outlier markers that had been previously associated with grilsing (i.e., early sexual maturation) an undesirable trait, which has been heavily selected against in Atlantic salmon aquaculture. This study provides clear evidence of the presence of signatures of selection and domestication in a farmed Atlantic salmon population.

Assessing footprints of selection in commercial Atlantic salmon populations using microsatellite data

Relatively large rates of response to traits of economic importance have been observed in different selection experiments in salmon. Several QTL have been mapped in the salmon genome, explaining unprecedented levels of phenotypic variation. Owing to the relatively large selection intensity, individual loci may be indirectly selected, leaving molecular footprints of selection, together with increased inbreeding, as its likely relatives will share the selected loci. We used population differentiation and levels of linkage disequilibrium in chromosomes known to be harbouring QTL for body weight, infectious pancreatic necrosis resistance and infectious salmon anaemia resistance to assess the recent selection history at the genomic level in Atlantic salmon. The results clearly suggest that the marker SSA0343BSFU on chromosome 3 (body weight QTL) showed strong evidence of directional selection. It is intriguing that this marker is physically mapped to a region near the coding sequence of DVL2 , making it an ideal candidate gene to explain the rapid evolutionary response of this chromosome to selection for growth in Salmo salar. Weak evidence of diversifying selection was observed in the QTL associated with infectious pancreatic necrosis and infectious salmon anaemia resistance. Overall, this study showed that artificial selection has produced important changes in the Atlantic salmon genome, validating QTL in commercial salmon populations used for production purposes according to the recent selection history.

Genomic Prediction Accuracy for Resistance Against Piscirickettsia salmonis in Farmed Rainbow Trout

Salmonid rickettsial syndrome (SRS), caused by the intracellular bacterium Piscirickettsia salmonis, is one of the main diseases affecting rainbow trout (Oncorhynchus mykiss) farming. To accelerate genetic progress, genomic selection methods can be used as an effective approach to control the disease. The aims of this study were: (i) to compare the accuracy of estimated breeding values using pedigree-based best linear unbiased prediction (PBLUP) with genomic BLUP (GBLUP), single-step GBLUP (ssGBLUP), Bayes C, and Bayesian Lasso (LASSO); and (ii) to test the accuracy of genomic prediction and PBLUP using different marker densities (0.5, 3, 10, 20, and 27 K) for resistance against P. salmonis in rainbow trout. Phenotypes were recorded as number of days to death (DD) and binary survival (BS) from 2416 fish challenged with P. salmonis. A total of 1934 fish were genotyped using a 57 K single-nucleotide polymorphism (SNP) array. All genomic prediction methods achieved higher accuracies than PBLUP. The relative increase in accuracy for different genomic models ranged from 28 to 41% for both DD and BS at 27 K SNP. Between different genomic models, the highest relative increase in accuracy was obtained with Bayes C (∼40%), where 3 K SNP was enough to achieve a similar accuracy to that of the 27 K SNP for both traits. For resistance against P. salmonis in rainbow trout, we showed that genomic predictions using GBLUP, ssGBLUP, Bayes C, and LASSO can increase accuracy compared with PBLUP. Moreover, it is possible to use relatively low-density SNP panels for genomic prediction without compromising accuracy predictions for resistance against P. salmonis in rainbow trout.

Applications in the search for genomic selection signatures in fish

Selection signatures are genomic regions harboring DNA sequences functionally involved in the genetic variation of traits subject to selection. Selection signatures have been intensively studied in recent years because of their relevance to evolutionary biology and their potential association with genes that control phenotypes of interest in wild and domestic populations. Selection signature research in fish has been confined to a smaller scale, due in part to the relatively recent domestication of fish species and limited genomic resources such as molecular markers, genetic mapping, DNA sequences, and reference genomes. However, recent genomic technology advances are paving the way for more studies that may contribute to the knowledge of genomic regions underlying phenotypes of biological and productive interest in fish.

From the viral perspective: Infectious salmon anemia virus (ISAV) transcriptome during the infective process in Atlantic salmon (Salmo salar)

The infectious salmon anemia virus (ISAV) is a severe disease that mainly affects the Atlantic salmon (Salmo salar) aquaculture industry. Although several transcriptional studies have aimed to understand Salmon-ISAV interaction through the evaluation of host-gene transcription, none of them has focused their attention upon the viral transcriptional dynamics. For this purpose, RNA-Seq and RT-qPCR analyses were conducted in gills, liver and head-kidney of S. salar challenged by cohabitation with ISAV. Results evidence the time and tissue transcript patterns involved in the viral expression and how the transcription levels of ISAV segments are directly linked with the protein abundance found in other virus of the Orthomyxoviridae family. In addition, RT-qPCR result evidenced that quantification of ISAV through amplification of segment 3 would result in a more sensitive approach for detection and quantification of ISAV. This study offers a more comprehensive approach regarding the ISAV infective process and gives novel knowledge for its molecular detection.