Alexander Falkenhagen, German Federal Institute for Risk Assessment

Alexander Falkenhagen, German Federal Institute for Risk Assessment Roman Valusenko-Mehrkens, German Federal Institute for Risk Assessment Reimar Johne, German Federal Institute for Risk Assessment

Combatting and Exploiting dsRNA viruses

Despite the widespread use of vaccines, rotavirus disease burden remains high in Sub-Saharan Africa. As the antigenic profile of rotaviruses circulating in Sub-Saharan Africa can substantially vary from other regions, we explored different reverse genetics strategies for the generation of vaccine strains based on simian rotavirus SA11 carrying VP4, VP6 and VP7 antigens from African rotaviruses. While generation of VP6 or VP7 reassortants was easily achieved, production of VP4 reassortants proved more challenging, but could be improved by substituting the putative receptor-binding region or combination with homologous VP6 and VP7. Long-term passaging of reassortants in cell culture further improved replication and resulted in the identification of multiple mutations in different genome segments. A common problem of available sequences from field strains is the lack of sequence information for the untranslated regions (UTRs). We therefore examined the generation of reassortants with a chimeric genome segment comprised of a human rotavirus VP7 open reading frame (ORF) flanked by simian rotavirus SA11 UTRs, showing that the simian rotavirus UTRs supported replication of these reassortants. Finally, we investigated attenuation by introducing NSP1 deletions of 559, 1110 or 1248 base pairs across different NSP1 regions. While replication was unaffected in African green monkey kidney-derived MA-104 cells, replication in human colon-derived HT-29 cells depended on the deletion and was 1-3 log10 lower compared to the non-truncated virus. Taken together, we have shown different approaches to improve replication, broaden the antigenic repertoire, and achieve attenuation, which could help in the development of a next-generation vaccine.