title: A naturally occurring substitution in the E2 protein of Salmonid alphavirus subtype 3 changes viral fitness.
abstract: Phylogenetic analyses of the Salmonid alphavirus subtype 3 (SAV3) epizootic have suggested that a substitution from proline to serine in the receptor binding protein E2 position 206 has occurred after the introduction of virus from a wild reservoir to farmed salmonid fish in Norway. We modelled the 3D structure of P62, the uncleaved E3-E2 precursor, of SAVH20/03 based on its sequence homology to the Chikungunya virus (CHIKV), and studied in vitro and in vivo effects of the mutation using reverse genetics. E2(206) is located on the surface of the B-domain of E2, which is associated with receptor attachment in alphaviruses. Recombinant virus expressing the E2(206S) codon replicated slower and produced significantly less genomic copies than virus expressing the ancestral E2(206P) codon in vitro in Bluegill Fry (BF2) cells. The E2(206S) mutant was out-competed by the E2(206P) mutant after 5 passages in an in vitro competition assay, confirming that the substitution negatively affects the efficacy of virus multiplication in cell culture. Both mutants were highly infectious to Atlantic salmon (Salmo salar), produced similar viral RNA loads in gills, heart, kidney and brain, and induced similar histopathologic changes in these organs. The E2(206S) mutant produced a less persistent infection in salmon and was shed more rapidly to water than the E2(206P) mutant. Reduced generation time through more rapid shedding could therefore explain why a serine in this position became dominant in the viral population after SAV3 was introduced to farmed salmon from the wild reservoir.
authors.affiliation: Department of Biology, University of Bergen, Thor Møhlens gt 55, 5020 Bergen, Norway. Electronic address: marius.karlsen@pharmaq.no.
authors.forename: Marius
authors.initials: M
authors.lastname: Karlsen
authors.affiliation: Department of Biology, University of Bergen, Thor Møhlens gt 55, 5020 Bergen, Norway.
authors.forename: Linda
authors.initials: L
authors.lastname: Andersen
authors.affiliation: Department of Biology, University of Bergen, Thor Møhlens gt 55, 5020 Bergen, Norway.
authors.forename: Steffen H
authors.initials: SH
authors.lastname: Blindheim
authors.affiliation: Department of Food Safety and Infection Biology, Faculty of Veterinary Medicine and Biosciences, Norwegian University of Life Sciences, P.O. 8146 Dep, 0033 Oslo, Norway.
authors.forename: Espen
authors.initials: E
authors.lastname: Rimstad
authors.affiliation: Department of Biology, University of Bergen, Thor Møhlens gt 55, 5020 Bergen, Norway.
authors.forename: Are
authors.initials: A
authors.lastname: Nylund
mesh_terms: D000524:Alphavirus / Q000235:genetics*; D019943:Amino Acid Substitution; D000818:Animals; D004252:DNA Mutational Analysis; D005393:Fish Diseases / Q000821:virology; D056084:Genetic Fitness; D008958:Models, Molecular; D009154:Mutation; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D012367:RNA, Viral / Q000235:genetics; D020317:Salmo salar / Q000821:virology; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D014779:Virus Replication; D017201:Virus Shedding
chemical_list: D012367:RNA, Viral; D014764:Viral Proteins
keywords: Adaptation; E2; Evolution; SAV3; Salmo salar; Salmonid alphavirus