Probing the Viromic Frontiers

Modern molecular technology, and particularly high-throughput sequencing (HTS), has revolutionized virus discov- ery and expanded the depth and breadth of the virome. Recent HTS was used to identify and discover a previously undescribed member of the familyFlaviviridae that has genomic features characteristic of both hepaciviruses and pegiviruses. This virus, designated human hepegivirus-1 (HHpgV-1), may represent a previously undescribed new genus in the Flaviviridae family with implications for public health and blood supply safety. Detecting uncharacterized viruses such as HHpgV-1 in clinical samples requires an unbiased screening method that is as sensitive as PCR, while simultaneously detecting multiple rare viral sequences. The virome-capture-sequencing platform for vertebrate viruses (VirCapSeq-VERT) uses positive-selection oligonucleotide cap- ture to sensitively detect sequences from every known vertebrate virus, even in high-background specimens with low-abundance viruses. VirCapSeq-VERT can also detect uncharacterized viruses with sequence homology to known viruses, enabling a new paradigm for virus detection. M odern molecular biology has accelerated the pace of virus discovery, allowing pathogen identification based on ge- nome characterization to proceed much more rapidly than tradi- tional virologic, microscopic, or serologic methods. In particular, high-throughput sequencing (HTS) has revolutionized the unbi- aseddiscoveryofnovelviruses,ofteninunexpectedplacesthatare highly relevant to human health. This is exemplified by a recent article by Kapoor and colleagues from W. Ian Lipkin's research laboratory (1), in which a novel human virus was found in serum samples from blood transfusion recipients and hemophilia pa- tients treated with plasma-derived clotting factors. Further analy- sisindicatedthatthisviruscombinesfeaturesofhepacivirusesand pegiviruses, two distinct genera within the family Flaviviridae. This virus, named human hepegivirus-1 (HHpgV-1), may repre- sent a new genus within the Flaviviridae and a potential human pathogen. Previous work from the Lipkin laboratory has enhanced our understanding of hepacivirus and pegivirus diversity in the wild. Hepatitis C virus (HCV) was long thought to be the sole member of the genus Hepacivirus, until the discovery of a novel canine hepacivirus (2) suggested that there may be greater hepacivirus diversity in nonhuman mammalian species. Since then, numer- oushepaciviruseshavebeencharacterizedinhorses,bats,rodents, and nonhuman primates (3-7). A similar host range diversity has been observed for pegiviruses (8). As HTS-based methods in- crease for viromics study applications, we also expect to discover additional novel members of Flaviviridae and develop a more comprehensive understanding of flavivirus biology. The HHpgV-1 sequence shows that it has a type IV internal ribosome entry site (IRES) in its 5= untranslated region (UTR) withconsiderablestructuralhomologytothatfoundinHCV.This suggests that like hepaciviruses, translation of the viral polypro- tein occurs via a common internal initiation mechanism. HHpgV-1 also contained a distinguishing sequence in the pseu- doknot domain that is highly conserved across the genus Hepaci- virus. However, analysis of two relatively conserved nonstructural genes, the viral helicase (NS3) and RNA-dependent RNA poly- merase (NS5B), demonstrated that HHpgV-1 is phylogenetically grouped as a distinct branch within the pegivirus clade. Thus, HHpgV-1sharescharacteristicsofbothgenera,suggestingeithera