Abstract
The sole member of the Piscihepevirus genus (family Hepeviridae) is cutthroat trout virus (CTV) but recent metatranscriptomic studies have identified numerous fish hepevirus sequences including CTV-2. In the current study, viruses with sequences resembling both CTV and CTV-2 were isolated from salmonids in eastern and western Canada. Phylogenetic analysis of eight full genomes delineated the Canadian CTV isolates into two genotypes (CTV-1 and CTV-2) within the Piscihepevirus genus. 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 was not previously observed. 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 hosts and potential reservoirs 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.
Highlights
Pacific Biological Station, Fisheries and Oceans Canada, Nanaimo, BC V9T 6N7, Canada; Gulf Fisheries Centre, Fisheries and Oceans Canada, Moncton, NB E1C 5K4, Canada; New Jersey Division of Fish and Wildlife, Office of Fish & Wildlife Health & Forensics, Abstract: The sole member of the Piscihepevirus genus is cutthroat trout virus (CTV) but recent metatranscriptomic studies have identified numerous fish hepevirus sequences including CTV-2
In addition to differing in host range, the two genera are phylogenetically distant with cutthroat trout virus (CTV), the sole member of the Piscihepevirus genus, being the most distant virus of the Hepeviridae family sharing less than 46% nucleotide identity with the Orthohepeviruses [2]
As CTV was found to replicate in an embryo derived cell line, it remains the only hepevirus that can efficiently replicate in cell culture and has shown promise as an Hepatitis E virus (HEV) surrogate [7,8]
Summary
Farmed Atlantic salmon, rainbow trout (Oncorhynchus mykiss), and Arctic char (Salvelinus alpinus) were collected from 2010 to 2017 for research purposes, mortality investigations, or as routine fish health screening as part of the Fish Health Protection. Tissue homogenates of gill from individual fish or kidney, spleen, pyloric caeca-pancreas, and gills from 5 fish, were inoculated in duplicate onto chinook salmon embryo cells (CHSE-214; ATCC CRL-1681) in 24-well tissue culture plates. Plates were incubated at 15 ◦ C and regularly monitored for cytopathic effect (CPE) for 21 d post-inoculation. If wells showed presumptive CPE, the supernatant and cells were removed from the well, passed through a 0.45 μm filter, and re-inoculated Wells displaying CPE were harvested and archived at −80 ◦ C. The virus isolate used for in vivo and in vitro studies, identified as CA/BC/2017-111/AtS, was harvested from CHSE-214 cells 21 days post-inoculum and quantified using the TCID50 method as described by Reed and Muench [13]
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