Evidence for Novel Hepaciviruses in Rodents

Drexler, Jan Felix; Corman, Victor Max; Müller, Marcel Alexander; Lukashev, Alexander N.; Gmyl, Anatoly; Coutard, Bruno; Adam, Alexander; Ritz, Daniel; Leijten, Lonneke M.; van Riel, Debby; Kallies, Rene; Klose, Stefan M.; Gloza-Rausch, Florian; Binger, Tabea; Annan, Augustina; Adu-Sarkodie, Yaw; Oppong, Samuel; Bourgarel, Mathieu; Rupp, Daniel; Hoffmann, Bernd; Schlegel, Mathias; Kümmerer, Beate M.; Krüger, Detlev H.; Schmidt-Chanasit, Jonas; Setién, Alvaro Aguilar; Cottontail, Veronika M.; Hemachudha, Thiravat; Wacharapluesadee, Supaporn; Osterrieder, Klaus; Bartenschlager, Ralf; Matthee, Sonja; Beer, Martin; Kuiken, Thijs; Reusken, Chantal; Leroy, Eric M.; Ulrich, Rainer G.; Drosten, Christian

The genome ends of representatives of all three rodent viruses were determined, including virus RMU10-3382 belonging to M. glareolus clade 1, NLR-AP-70 belonging to M. glareolus clade 2, and virus SAR-46 belonging to the R. pumilio hepacivirus clade. Figure 5 and Supplementary Figure S2A show that the 59genome terminus of RMU10-3382 contained structural elements typical of both pegi-and HCV-like internal ribosomal entry sites (IRESs). Predicted structural similarities with the HCV-like IRES included the first stem-loop element (termed Ia and highlighted in orange in Figure 5 ) and one of two sites involved in miRNA122 binding [51] , while most of the remaining stem-loop elements (termed 3, 4 and 5 and highlighted in blue in Figure 5 ) were more closely related to a pegivirus-like IRES. The 59-end of AP-70 was identical in structure to RMU10-3382 and contained only a few nucleotide exchanges. SAR-46 contained the typical HCV-like IRES structures including the characteristic stem-loop III ( Figure 5 and Supplementary Figure S2B) . The observed structural similarity between the first stem-loop of all rodent viruses described here and the prototype hepaciviruses HCV and GBV-B consisted of a hairpin with a six-nucleotide stem and fourfive nucleotide loop. The equine/canine hepaciviruses contained a similar structural element located as their second predicted IRES domain, instead of the most 59-position this domain occupied in all other hepaciviruses. The RMU10-3382 and NLR-365 translation initiation sites contained a cytosine immediately following the putative start codon at position +4, which is suboptimal in the original Kozak sequence context (ACCATGG) but should not block initiation [52] . The 39-ends of RMU10-3382 and SAR-46 contained three highly ordered stem-loop elements. In RMU10-3382, these RNA elements did not resemble any known 39noncoding sequence RNA structure. In SAR-46, the 39-terminal stem-loop structure, but not the preceding structures, resembled that of the HCV X-tail ( Figure 5 and Supplementary Figure S3) . A similar 39-terminal structure could be predicted for GBV-B, but not for the genetically related pegiviruses ( Figure 5 and Supplementary Figure S4) . The 39-end of NLR-AP-70 could not be determined. Contrary to HCV and GBV-B, no poly-uracil stretch was observed in the rodent hepaciviruses.