Weining Wu, London School of Hygiene and Tropical Medicine

Weining Wu1, Ulrike Laugks2,3, Kay Grünewald2,3,4 and Polly Roy1 1. Department of Infection Biology, Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, London, United Kingdom 2. Centre for Structural System Biology, 22607 Hamburg, Germany 3. Department of Chemistry, University of Hamburg, 20146 Hamburg, Germany 4. Leibniz Institute of Virology, 20251 Hamburg, Germany

Virus Replication: Entry, Exit and Everything in Between

Bluetongue virus (BTV) is one of the most economically relevant orbiviruses and is the only example of a large complex, non-enveloped arbovirus. In addition to cell lysis, BTV is known to employ a ‘budding’ process similar to that used by enveloped viruses for cell exit, in which the viral NS3 protein plays a key role. Recent reports have demonstrated that BTV can also induce non-lytic release via extracellular vesicles (EVs), however, details of the type of EV used and the role of NS3 in the process remain incompletely understood. In this study we undertook a comprehensive microscopic analysis using electron cryo-tomography and transmission electron microscopy with immunogold labelling, together with biochemical studies on the non-lytic release of BTV particles in different forms of EV from different types of host cell. We found that BTV particles use both large EVs (LEVs) and smaller size EVs (SEVs) for non-lytic release and that, in each cell type studied, NS3 was particularly enriched in the SEV fraction. Non-enveloped BTV particles released in these ER-derived SEVs were highly infectious and central for efficient cell-to-cell transmission. Our findings highlight the complexity of the underlying mechanisms utilized by a non-enveloped arbovirus for egress and the significance of different types of EVs in this process.