Characterization of a quasi-enveloped, fast replicating hepevirus from fish and its use as hepatitis E virus surrogate

• Published in: Journal of virological methods Vol. 263; pp. 111 - 119
• Main Authors: Bochud, Maëlle, Schäfer, Wolfram, Roth, Nathan J., Ros, Carlos
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.01.2019
• Subjects: Animals; Cell Line; Cutthroat trout virus; Estradiol - pharmacology; Ethanol - pharmacology; Hepatitis E virus; Hepevirus; Hepevirus - drug effects; Hepevirus - physiology; Hepevirus - ultrastructure; HEV; Interferons - pharmacology; Models, Biological; Oncorhynchus mykiss; Pasteurization; Quasi-enveloped; RNA, Viral - analysis; Viral Envelope Proteins - metabolism; Virus Inactivation - drug effects; Virus model; Virus Replication - drug effects; HEV; Quasi-enveloped; Hepevirus; Virus model; Cutthroat trout virus
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2018.11.002

•The lack of an efficient cell culture system hampers the study of human HEV.•CTV is a fish HEV, which can replicate efficiently in cell culture to high titers.•There are substantial similarities between the human and the fish HEV.•CTV is a practical model for virus clearance studies and to gain new insights into the understudied biology of hepeviruses. Hepatitis E virus (HEV) is an emerging concern for the safety of plasma-derived medicinal products. The lack of an efficient cell culture system hampers the studies on HEV biology as well as validation studies to test the capacity of virus reduction steps to clear HEV. Hence, a surrogate hepevirus that can efficiently replicate in cell culture is needed. Cutthroat trout virus (CTV) is a non-pathogenic fish hepevirus, which can replicate in cell culture to high titers. Under interferon inhibition, CTV replication reached up to 5 × 107 genome equivalents per μL in 4-5 days. The intracellular CTV progeny was already lipid-associated, suggesting that the envelope is acquired from intracellular membranes. Transmission electron microscopy of purified quasi-enveloped virus revealed exosome-like structures with an average size of 40 nm, in contrast to 27–34 nm for the non-enveloped virus. The quasi-enveloped virus was significantly less infectious than the non-enveloped virus. Assays based on quantitative RT-PCR, immunofluorescence and immunocytochemistry were established to evaluate virus inactivation. Cold ethanol fractionation removed 3.0 log of CTV and pasteurization of human albumin inactivated more than 3.7 log to below the limit of detection. Similar to HEV, virus replication was promoted in the presence of 17β-estradiol, an effect that can contribute to the understanding of the exacerbated virulence of HEV in pregnant women. These results together reveal substantial similarities between the human and fish HEV and validate CTV as a practical virus model to use in some applications for evaluating the HEV reduction capacity of biological manufacturing process steps.