Rescue of bovine ephemeral fever virus through reverse genetics, but inability to propagate.

Abstract

Bovine ephemeral fever (BEF) is caused by BEF virus (BEFV) belonging to the Genus Ephemerovirus under the Family Rhabdoviridae. The BEFV carries a single-stranded, negative-sense RNA genome. Not much is known about the various aspects of BEFV replication, its interaction with cellular proteins or the cellular response to BEFV infection. Here, we report the rescue of BEFV through reverse genetics. A full-length cDNA copy of BEFV was assembled to be driven by the RNA polymerase I (PolI) promoter. Parallely, eukaryotic expression plasmids containing BEFV sequences encoding the helper proteins N, P and L, which form the replicase complex, were generated. The expression of N and P proteins were verified by using the in-house generated and purified polyclonal sera. Transfection of the full-length cDNA copy along with the helper plasmids rescued BEFV, as evaluated by transmission electron microscopy, reverse-transcription polymerase reaction, immunofluorescence and Western blotting. However, the virus did not produce a cytopathic effect and failed to be propagated beyond a certain number of passages. The results lay the foundation for establishment of reverse genetics for BEFV but also highlight the difficulties in studying this virus.