Investigating the entry mechanisms of a new member of the Orthobunyavirus genus.

Abstract

Recently, a novel member of the Orthobunyavirus genus of segmented negative sense RNA viruses was identified in two immunocompromised patients with fatal encephalitis. Whilst the infectious virus remains to be isolated, phylogenetic sequence analysis places it within the Turlock serogroup, closely related to Umbre virus (UMBV). Termed Cristoli virus (CRIV), its natural reservoir, tropism, and life cycle remain uncharacterised. Given the association of increased geographical range of vectors and Orthobunya-reassortments with outbreaks of haemorrhagic fever, the characterisation of the life cycles of novel viruses is crucial in anticipation of potential future outbreaks. In the absence of isolated virus and specific antibodies, we utilised pseudotyped lentiviruses and tagged proteins were utilised to characterise CRIV. We observed the infection of multiple cell types, and widespread Golgi disruption upon M-segment expression, suggesting a potential role in viral factory formation. To further study CRIV, we isolated the whole genome of UMBV, determining the sequence of its terminal regions using 3’ and 5’ RACE, and rescued infectious virus from cDNAs. A rescued GFP-recombinant version of UMBV was used confirming the tropism results obtained using the highly related CRIV pseudoviruses. Generation of NP antibodies for use as markers of virus multiplication have facilitated the characterisation of the general life cycle of rUMBV, as well as the potential immune evasion mechanisms of UMBV/CRIV. Further research using the rescued virus obtained from this research on UMBV/CRIV will illuminate viral and host components necessary for replication, potentially paving the way for future antiviral or host-directed therapies.