Generation of a recombinant Akabane virus expressing enhanced green fluorescent protein

Akiko Takenaka-Uema, Yousuke Murata, Fumihiro Gen, Yukari Ishihara-Saeki, Ken Ichi Watanabe, Kazuyuki Uchida, Kentaro Kato, Shin Murakami, Takeshi Haga, Hiroomi Akashi, Taisuke Horimoto

Research output: Contribution to journalArticlepeer-review

3 Citations (Scopus)

Abstract

We generated a recombinant Akabane virus (AKAV) expressing enhanced green fluorescence protein (eGFP-AKAV) by using reverse genetics. We artificially constructed an ambisense AKAV S genome encoding N/NSs on the negative-sense strand, and eGFP on the positive-sense strand with an intergenic region (IGR) derived from the Rift Valley fever virus (RVFV) S genome. The recombinant virus exhibited eGFP fluorescence and had a cytopathic effect in cell cultures, even after several passages. These results indicate that the gene encoding eGFP in the ambisense RNA could be stably maintained. Transcription of N/NSs and eGFP mRNAs of eGFP-AKAV was terminated within the IGR. The mechanism responsible for this appears to be different from that in RVFV, where the termination sites for N and NSs are determined by a defined signal sequence. We inoculated suckling mice intraperitoneally with eGFP-AKAV, which resulted in neurological signs and lethality equivalent to those seen for the parent AKAV. Fluorescence from eGFP in frozen brain slices from the eGFP-AKAV-infected mice was localized to the cerebellum, pons, and medulla oblongata. Our approach to producing a fluorescent virus, using an ambisense genome, helped obtain eGFPAKAV, a fluorescent bunyavirus whose viral genes are intact and which can be easily visualized.

Original languageEnglish
Pages (from-to)9477-9484
Number of pages8
JournalJournal of virology
Volume89
Issue number18
DOIs
Publication statusPublished - 2015
Externally publishedYes

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

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