Identification and characterization of a novel orthoreovirus from American eels, Anguilla rostrata

Hemorrhagic gill necrosis disease (HGND) has resulted in substantial economic losses in the farming of American eels (Anguilla rostrata). Although prior research implicates viruses like American eel adomavirus (AEAdoV) and Anguillid herpesvirus (AngHV), the primary etiological agent remains elusive. In this study, a novel virus was isolated from the HGND samples of American eel using eel ovary (EO) cell lines. Following isolation, the virus was subjected to comprehensive characterization encompassing morphological features, genomic organization, phylogenetic relationships, pathogenic potential, and epidemiological prevalence. The findings revealed that the virions are non-enveloped, icosahedral symmetry with a double-layered capsid and containing a genome composed of 10 double-stranded RNA segments, thereby confirming its classification within the genus Orthoreovirus. Phylogenetic analyses positioned this orthoreovirus in a distinct clade alongside largemouth bass reovirus (LMBRV) and piscine reovirus (PRV), occupying an intermediate position between the Orthoreovirus and Aquareovirus genera, indicating a unique evolutionary lineage within the Spinareoviridae family. This virus has been designated as Anguillid orthoreovirus (AORV). Experimental intraperitoneal challenge of American eels with AORV did not lead to apparent clinical manifestations. Epidemiological investigation revealed that AORV is an endemic orthoreovirus with an infection prevalence of 32.3% in farmed American eels. In summary, this study reports the successful isolation and comprehensive characterization of a novel orthoreovirus, AORV, from HGND samples of American eel. These findings provide a foundation for further investigations into the biology of AORV and offer novel insights toward future taxonomic refinement within the family Spinareoviridae.IMPORTANCEA novel orthoreovirus, designated Anguillid orthoreovirus (AORV), was isolated from American eels with "hemorrhagic gill necrosis disease," and forms a unique evolutionary lineage within the Spinareoviridae family. Although experimental challenges with AORV did not result in observable pathogenicity, epidemiological surveys indicated that approximately one-third of the farmed American eels displaying clinical signs of "hemorrhagic gill necrosis disease" tested positive for the virus, suggesting that AORV may be secondary to another primary pathogen or environmental stressor.