Sissel Kjøglum

Confirmation and fine-mapping of a major QTL for resistance to infectious pancreatic necrosis in Atlantic salmon (Salmo salar): population-level associations between markers and trait.

BackgroundInfectious pancreatic necrosis (IPN) is one of the most prevalent and economically devastating diseases in Atlantic salmon (Salmo salar) farming worldwide. The disease causes large mortalities at both the fry- and post-smolt stages. Family selection for increased IPN resistance is performed through the use of controlled challenge tests, where survival rates of sib-groups are recorded. However, since challenge-tested animals cannot be used as breeding candidates, within-family selection is not performed and only half of the genetic variation for IPN resistance is being exploited. DNA markers linked to quantitative trait loci (QTL) affecting IPN resistance would therefore be a powerful selection tool. The aim of this study was to identify and fine-map QTL for IPN-resistance in Atlantic salmon, for use in marker-assisted selection to increase the rate of genetic improvement for this trait.ResultsA genome scan was carried out using 10 large full-sib families of challenge-tested Atlantic salmon post-smolts and microsatellite markers distributed across the genome. One major QTL for IPN-resistance was detected, explaining 29% and 83% of the phenotypic and genetic variances, respectively. This QTL mapped to the same location as a QTL recently detected in a Scottish Atlantic salmon population. The QTL was found to be segregating in 10 out of 20 mapping parents, and subsequent fine-mapping with additional markers narrowed the QTL peak to a 4 cM region on linkage group 21. Challenge-tested fry were used to show that the QTL had the same effect on fry as on post-smolt, with the confidence interval for QTL position in fry overlapping the confidence interval found in post-smolts. A total of 178 parents were tested for segregation of the QTL, identifying 72 QTL-heterozygous parents. Genotypes at QTL-heterozygous parents were used to determine linkage phases between alleles at the underlying DNA polymorphism and alleles at single markers or multi-marker haplotypes. One four-marker haplotype was found to be the best predictor of QTL alleles, and was successfully used to deduce genotypes of the underlying polymorphism in 72% of the parents of the next generation within a breeding nucleus. A highly significant population-level correlation was found between deduced alleles at the underlying polymorphism and survival of offspring groups in the fry challenge test, parents with the three deduced genotypes (QQ, Qq, qq) having mean offspring mortality rates of 0.13, 0.32, and 0.49, respectively. The frequency of the high-resistance allele (Q) in the population was estimated to be 0.30. Apart from this major QTL, one other experiment-wise significant QTL for IPN-resistance was detected, located on linkage group 4.ConclusionThe QTL confirmed in this study represents a case of a major gene explaining the bulk of genetic variation for a presumed complex trait. QTL genotypes were deduced within most parents of the 2005 generation of a major breeding company, providing a solid framework for linkage-based MAS within the whole population in subsequent generations. Since haplotype-trait associations valid at the population level were found, there is also a potential for MAS based on linkage disequilibrium (LD). However, in order to use MAS across many generations without reassessment of linkage phases between markers and the underlying polymorphism, the QTL needs to be positioned with even greater accuracy. This will require higher marker densities than are currently available.

Epithelial Cadherin Determines Resistance to Infectious Pancreatic Necrosis Virus in Atlantic Salmon

Infectious pancreatic necrosis virus (IPNV) is the cause of one of the most prevalent diseases in farmed Atlantic salmon (Salmo salar). A quantitative trait locus (QTL) has been found to be responsible for most of the genetic variation in resistance to the virus. Here we describe how a linkage disequilibrium-based test for deducing the QTL allele was developed, and how it was used to produce IPN-resistant salmon, leading to a 75% decrease in the number of IPN outbreaks in the salmon farming industry. Furthermore, we describe how whole-genome sequencing of individuals with deduced QTL genotypes was used to map the QTL down to a region containing an epithelial cadherin (cdh1) gene. In a coimmunoprecipitation assay, the Cdh1 protein was found to bind to IPNV virions, strongly indicating that the protein is part of the machinery used by the virus for internalization. Immunofluorescence revealed that the virus colocalizes with IPNV in the endosomes of homozygous susceptible individuals but not in the endosomes of homozygous resistant individuals. A putative causal single nucleotide polymorphism was found within the full-length cdh1 gene, in phase with the QTL in all observed haplotypes except one; the absence of a single, all-explaining DNA polymorphism indicates that an additional causative polymorphism may contribute to the observed QTL genotype patterns. Cdh1 has earlier been shown to be necessary for the internalization of certain bacteria and fungi, but this is the first time the protein is implicated in internalization of a virus.

Genomic prediction in an admixed population of Atlantic salmon (Salmo salar)

Reliability of genomic selection (GS) models was tested in an admixed population of Atlantic salmon, originating from crossing of several wild subpopulations. The models included ordinary genomic BLUP models (GBLUP), using genome-wide SNP markers of varying densities (1–220 k), a genomic identity-by-descent model (IBD-GS), using linkage analysis of sparse genome-wide markers, as well as a classical pedigree-based model. Reliabilities of the models were compared through 5-fold cross-validation. The traits studied were salmon lice (Lepeophtheirus salmonis) resistance (LR), measured as (log) density on the skin and fillet color (FC), with respective estimated heritabilities of 0.14 and 0.43. All genomic models outperformed the classical pedigree-based model, for both traits and at all marker densities. However, the relative improvement differed considerably between traits, models and marker densities. For the highly heritable FC, the IBD-GS had similar reliability as GBLUP at high marker densities (>22 k). In contrast, for the lowly heritable LR, IBD-GS was clearly inferior to GBLUP, irrespective of marker density. Hence, GBLUP was robust to marker density for the lowly heritable LR, but sensitive to marker density for the highly heritable FC. We hypothesize that this phenomenon may be explained by historical admixture of different founder populations, expected to reduce short-range lice density (LD) and induce long-range LD. The relative importance of LD/relationship information is expected to decrease/increase with increasing heritability of the trait. Still, using the ordinary GBLUP, the typical long-range LD of an admixed population may be effectively captured by sparse markers, while efficient utilization of relationship information may require denser markers (e.g., 22 k or more).

Difference in skin immune responses to infection with salmon louse (Lepeophtheirus salmonis) in Atlantic salmon (Salmo salar L.) of families selected for resistance and susceptibility.

Atlantic salmon is susceptible to the salmon louse (Lepeophtheirus salmonis) and the variation in susceptibility within the species can be exploited in selective breeding programs for louse resistant fish. In this study, lice counts were completed on 3000 siblings from 150 families of Atlantic salmon identified as high resistant (HR) and low resistant (LR) families in two independent challenge trials. Skin samples behind the dorsal fin (nearby lice attachment) were collected from ten extreme families (HR or LR) and analyzed by qPCR for the expression of 32 selected genes, including a number of genes involved in T helper cell (Th) mediated immune responses, which have been previously implied to play important roles during salmon louse infections. Most genes showed lower expression patterns in the LR than in HR fish, suggesting an immunosuppressed state in LR families. The average number of lice (chalimi) was 9 in HR and 15 in LR fish. Large variation in lice counts was seen both within resistant and susceptible families, which enabled us to subdivide the groups into HR < 10 and HR > 10, and LR < 10 and LR > 10 to better understand the effect of lice burden per se. As expected, expression patterns were influenced both by genetic background and the number of attached parasites. Higher number of lice (>10) negatively affected gene expression in both HR and LR families. In general, strongest down-regulation was seen in LR > 10 and lesser down-regulation in HR < 10. HR in general and especially HR < 10 fish were better at resisting suppression of expression of both Th1 and Th2 genes. However, the best inverse correlation with infection level was seen for the prototypical Th1 genes, including several members from the interferon pathways. In addition, skin histomorphometry suggests that infected LR salmon had thicker epidermis in the area behind the dorsal fin and larger mucous cell size compared to infected HR fish, however marginally significant (p = 0.08). This histomorphometric finding was in line with the immune response being skewed in LR towards the Th2 rather than a Th1 profile. Our findings suggest that the ability to resist lice infection depends on the ability to avoid immunosuppression and not as much on the physical tissue barrier functions.

The Effect of Specific MHC Class I and Class II Combinations on Resistance to Furunculosis in Atlantic Salmon (Salmo salar)

The aim was to evaluate the performance of selected individual major histocompatibility complex (MHC) class I and class II alleles and combinations of these on disease resistance against furunculosis. The material, consisting of 2,145 Atlantic salmon (Salmo salar) from seven families, contained four MHC class I (UBA) alleles and two MHC class II alpha (DAA) alleles. The included alleles were previously shown to influence resistance towards furunculosis. The fish were challenged with the bacterium Aeromonas salmonicidae, causing furunculosis. The total mortality in the material was 84.0%, and both dead and surviving fish were genotyped for UBA and DAA. For MHC class II fish with DAA*0301 were more resistant than fish with DAA*0201, which were significantly more susceptible (P < 0.05). For MHC class I, the ranking of performance of the alleles differed from previous results, and fish with allele UBA*0601 were found to be most resistant while fish with UBA*0501, UBA*0201 and UBA*0301 were more susceptible. The analysis of combined MHC class I and class II genotype detected that fish with genotypes UBA*0501/*0501;DAA*0301/*0301, UBA*0601/*0601;DAA*0301/*0301 and UBA*0601/*0601;DAA*0201/*0201 were most resistant (P < 0.05), while the genotypes associated with increased susceptibility were UBA*0501/*0501; DAA*0201/*0301 and UBA*0501/*0501;DAA*0201/*0201 (P < 0.05). In addition, Cox regression analysis show that both family and MHC alleles contributed significantly to survival. Thus, in addition to reproducing the effect on resistance and susceptibility of many MHC class I and class II alleles individually, we also detected an effect of certain MHC class I and class II combinations suggestive of cross‐talk between the class I and class II pathways.