Multiple detection of zoonotic variegated squirrel bornavirus 1 in different squirrel species
A recently discovered novel putative zoonotic bornavirus, variegated squirrel bor-navirus 1 (VSBV-1), caused fatal encephalitides in three squirrel breeders and a zookeeper. Viral RNA was detected in brain samples from the patients as well as in organ samples of a variegated squirrel (Sciurus variegatoides) kept by one of the breeders. Screening of more than 750 squirrels of 18 different species (oral swab samples and in parts fecal pool samples) using the published VSBV-1 specific RT-qPCR revealed a positivity rate of 3.5%, including squirrels of the subfamilies Sciurinae (1.5%) and Callosciurinae (8.5%). These squirrels belonged to private holdings as well as zoo-logical gardens. VSBV-1 sequences of the 28 RNA positive squirrels were generated and phylogenetic analysis revealed a holding-related clustering of the sequences, independent from the squirrel species. We were able to co-cultivate infected primary squirrel cells with a permanent cell line and to isolate infectious virus from these passaged cells. Electron microscopy exhibited the typical structure of a bornavirus. The virus isolate was used to infect neonatal rats intracranial. After a few weeks, two of four rats were VSBV-1 genome positive and virus re-isolation from rat brain material was successful. In conclusion, the non-invasive sampling methods and the established workflow seems to be suitable for rapid screening of squirrels and revealed further infected animals of different squirrel species, representing a threat for humans handling squirrels. In addition, we were able to isolate the virus and passage it in rats, which is a precondition for further animal trials.