Cutthroat trout virus (CTV) was first isolated in 1988 in spawning trout from the West Coast of the United states. Due to its similarities with hepatitis E virus it has been officially classified as the only aquatic member of the Hepeviridae family. Since the discovery of CTV, surveillance of salmon and trout in Eastern and Western Canada isolated and sequenced numerous viruses resembling CTV. Phylogenetic analysis of eight full genomes delineated the Canadian CTV isolates into two genotypes (CTV-1 and CTV-2). Hepevirus genomes typically have three open reading frames but an ORF3 counterpart was not predicted in the Canadian CTV isolates. In vitro replication of a CTV-2 isolate produced cytopathic effects in the CHSE-214 cell line with similar amplification efficiency as CTV. Likewise the morphology of the CTV-2 isolate resembled CTV, yet viral replication caused dilation of the endoplasmic reticulum lumen which has not been previously observed (Fig 1). Controlled laboratory studies exposing Sockeye (Oncorhynchus nerka), Pink (O. gorbuscha), and Chinook Salmon (O. tshawytscha) to CTV-2 resulted in persistent infections without disease and mortality. Infected Atlantic Salmon (Salmo salar) and Chinook Salmon served as a host and potential reservoir of CTV-2. The data presented herein provides the first in vitro and in vivo characterization of CTV-2 and reveals greater diversity of piscihepeviruses extending the known host range and geographic distribution of CTV viruses