VHSV Single Amino Acid Polymorphisms (SAPs) Associated With Virulence in Rainbow Trout

Abstract

The Viral Hemorrhagic Septicemia Virus (VHSV) is an OIE notifiable pathogen widespread in the Northern Hemisphere that encompasses four genotypes and nine subtypes. In Europe, subtype Ia impairs predominantly the rainbow trout industry causing severe rates of mortality, while other VHSV genotypes and subtypes affect a number of marine and freshwater species, both farmed and wild. VHSV has repeatedly proved to be able to jump to rainbow trout from the marine reservoir, causing mortality episodes. The molecular mechanisms regulating VHSV virulence and host tropism are not fully understood, mainly due to the scarce availability of complete genome sequences and information on the virulence phenotype. With the scope of identifying in silico molecular markers for VHSV virulence, we generated an extensive dataset of 55 viral genomes and related mortality data obtained from rainbow trout experimental challenges. Using statistical association analyses that combined genetic and mortality data, we found 38 single amino acid polymorphisms scattered throughout the complete coding regions of the viral genome that were putatively involved in virulence of VHSV in trout. Specific amino acid signatures were recognized as being associated with either low or high virulence phenotypes. The phylogenetic analysis of VHSV coding regions supported the evolution toward greater virulence in rainbow trout within subtype Ia, and identified several other subtypes which may be prone to be virulent for this species. This study sheds light on the molecular basis for VHSV virulence, and provides an extensive list of putative virulence markers for their subsequent validation.

The Viral Hemorrhagic Septicemia Virus (VHSV) Markers of Virulence in Rainbow Trout (Oncorhynchus mykiss)

“…A previous study, based on the visual inspection of a limited panel of VHSV isolates, demonstrated the reliability of this approach in pointing out a single amino acid change in the NV protein that was clearly associated with virulence in rainbow trout (Baillon et al, 2017). In the present study, 17 out of the 38 SAPs previously identified by association analysis (Panzarin et al, 2020) were evaluated, alone or in combination, by reverse genetics using the French VHSV strain 23-75 as a backbone. This virus was isolated in France in 1975 from a brown trout, and showed a highly-virulent phenotype with devastating effects on farmed rainbow trout (de Kinkelin and Le Berre, 1977;Gaudin et al, 1999).…”

“…Recombinant viruses were named according to the viral protein together with the position and the amino acid change. Specific primers were designed based on the exact codon found in variant strain sequences (Panzarin et al, 2020) and are available upon request. The mutagenesis reaction was performed on VHSV fragments subcloned in pJET1.2 plasmid (CloneJET PCR cloning kit; Fermentas).…”

“…The phylogenetic tree used in this study was previously developed by Panzarin et al (2020) in a study aiming at identifying candidate VHSV virulence markers by statistical association analysis. In brief, 55 VHSV genomes were aligned using MAFFT v7.388 (Katoh et al, 2002) with default settings.…”

“…In a recent study by Panzarin et al (2020), 55 VHSV isolates, covering all European genotypes and isolated from a large panel of marine and freshwater fish species, were characterized for their degree of virulence in rainbow trout and fully-sequenced. Strains were assigned to different virulence groups (high, moderate, and low), based on the cumulative mortality observed.…”

“…In a recent study by Panzarin et al (2020), 55 VHSV isolates covering all European genotypes were selected based on field records and host of origin, and were characterized for their virulence phenotype in juvenile rainbow trout by bath immersion. All viral isolates were also subject to whole genome sequencing (WGS) and phylogenetic reconstruction based on protein coding regions.…”