Infectious pancreatic necrosis viruses (IPNVs) belonging to the family Birnaviridae display a high degree of antigenic variability, pathogenicity, and differences in outbreak mortality in salmonid species. To determine if virus isolates of Sp serotype differ in virulence, fry of Atlantic salmon (Salmo salar L.) were challenged with nine different field strains. These viruses caused either high mortality and severe pathological changes or low mortality and no lesions. To study the molecular basis for the variation in virulence of IPNV, complete nucleotide sequences of segment A of all these strains as well as segment B of three selected strains were determined. All viruses tested had a unique genome sequence. Only minor differences were noted in the genes encoding VP1, VP3, and VP4 proteins, whereas most changes were observed in the gene encoding the VP2 protein. A high level of variation was found in the small open reading frame (ORF), which encodes a 15-kDa nonstructural (NS) polypeptide also known as VP5. One of the strains lacked the initiation codon for this protein, whereas the other four could encode a truncated version of the NS protein. Additional data obtained by sequencing of the NS and VP2 genes directly from diseased fish demonstrated changes in the VP2 gene after two passages in cell culture, which could possibly be associated with attenuation. Comparison of the deduced amino acid sequences of the NS and VP2 genes reveals that the virulent strains possess a 12-kDa coding NS gene and have residues Thr, Ala, Thr/Ala, and Tyr/His at positions 217, 221, 247, and 500 of the VP2 gene, respectively-the motifs identified in this study to be involved in the virulence of IPNV.

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.

Infectious pancreatic necrosis viruses (IPNVs) exhibit a wide range of virulence in salmonid species. In previous studies, we have shown that the amino acid residues at positions 217 and 221 in VP2 are implicated in virulence. To pinpoint the molecular determinants of virulence in IPNV, we generated recombinant IPNV strains using the cRNA-based reverse-genetics system. In two virulent strains, residues at positions 217 and 247 were replaced by the corresponding amino acids of a low-virulence strain. The growth characteristics of the recovered chimeric strains in cell culture were similar to the low-virulence strains, and these viruses induced significantly lower mortality in Atlantic salmon fry than the parent strains did in in vivo challenge studies. Furthermore, the virulent strain was serially passaged in CHSE-214 cells 10 times and was completely characterized by nucleotide sequencing. Deduced amino acid sequence analyses revealed a single amino acid substitution of Ala to Thr at position 221 in VP2 of this virus, which became highly attenuated and induced 15% cumulative mortality in Atlantic salmon fry, compared to 68% mortality induced by the virulent parent strain. The attenuated strain grows to higher titers in CHSE cells and can be distinguished antigenically from the wild-type virus by use of a monoclonal antibody. However, the virulent strain passaged 10 times in RTG-2 cells was stable, and it retained its antigenicity and virulence. Our results indicate that residues Thr at position 217 (Thr217) and Ala221 of VP2 are the major determinants of virulence in IPNV of the Sp serotype. Highly virulent isolates possess residues Thr217 and Ala221; moderate-to low-virulence strains have Pro217 and Ala221; and strains containing Thr221 are almost avirulent, irrespective of the residue at position 217.

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).

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Infectious pancreatic necrosis virus (IPNV) is the causative agent of infectious pancreatic necrosis (IPN)disease

The development of ectothermic embryos is strongly affected by incubation temperature, and thermal imprinting of body growth and muscle phenotype has been reported in various teleost fishes. The complex epigenetic regulation of muscle development in vertebrates involves DNA methylation of the myogenin promoter. Body growth is a heritable and highly variable trait among fish populations that allows for local adaptations, but also for selective breeding. Here we studied the epigenetic effects of embryonic temperature and genetic background on body growth, muscle cellularity and myogenin expression in farmed Atlantic salmon (Salmo salar). Eggs from salmon families with either high or low estimated breeding values for body growth, referred to as Fast and Slow genotypes, were incubated at 8°C or 4°C until the embryonic ‘eyed-stage’ followed by rearing at the production temperature of 8°C. Rearing temperature strongly affected the growth rates, and the 8°C fish were about twice as heavy as the 4°C fish in the order Fast8>Slow8>Fast4>Slow4 prior to seawater transfer. Fast8 was the largest fish also at harvest despite strong growth compensation in the low temperature groups. Larval myogenin expression was approximately 4–6 fold higher in the Fast8 group than in the other groups and was associated with relative low DNA methylation levels, but was positively correlated with the expression levels of the DNA methyltransferase genes dnmt1, dnmt3a and dnmt3b. Juvenile Fast8 fish displayed thicker white muscle fibres than Fast4 fish, while Slow 8 and Slow 4 showed no difference in muscle cellularity. The impact of genetic background on the thermal imprinting of body growth and muscle development in Atlantic salmon suggests that epigenetic variation might play a significant role in the local adaptation to fluctuating temperatures over short evolutionary time.

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