Development of an improved reverse genetics system for Akabane bunyavirus

Akiko Takenaka-Uema; Keita Sugiura; Norasuthi Bangphoomi; Chieko Shioda; Kazuyuki Uchida; Kentaro Kato; Takeshi Haga; Shin Murakami; Hiroomi Akashi; Taisuke Horimoto

doi:10.1016/j.jviromet.2015.12.014

Development of an improved reverse genetics system for Akabane bunyavirus

J Virol Methods. 2016 Jun:232:16-20. doi: 10.1016/j.jviromet.2015.12.014. Epub 2016 Feb 27.

Authors

Affiliations

¹ Department of Veterinary Microbiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan; Department of Infection Control and Disease Prevention, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: atakiko@mail.ecc.u-tokyo.ac.jp.
² Department of Veterinary Microbiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: sugiura_keita@kaken.co.jp.
³ Department of Veterinary Microbiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan; Department of Preclinical and Applied Animal Science, Faculty of Veterinary Science, Mahidol University, 999 Phutthamonthon Sai 4 Road Salaya, Phutthamonthon Nakhonpathom 73170, Thailand. Electronic address: norasuthi.ban@mahidol.ac.th.
⁴ Department of Veterinary Pathology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: chieko_shioda@nm.maff.go.jp.
⁵ Department of Veterinary Pathology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: auchidak@mail.ecc.u-tokyo.ac.jp.
⁶ Department of Veterinary Microbiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan; Research Unit for Global Infection Control, National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada-cho, Obihiro, Hokkaido 080-8555, Japan. Electronic address: kkato@obihiro.ac.jp.
⁷ Department of Infection Control and Disease Prevention, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: ahaga@mail.ecc.u-tokyo.ac.jp.
⁸ Department of Veterinary Microbiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: amurakam@mail.ecc.u-tokyo.ac.jp.
⁹ Department of Veterinary Microbiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: akashih@mail.ecc.u-tokyo.ac.jp.
¹⁰ Department of Veterinary Microbiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address: ahorimo@mail.ecc.u-tokyo.ac.jp.

PMID: 26927704
DOI: 10.1016/j.jviromet.2015.12.014

Abstract

Akabane disease, caused by the insect-transmitted Akabane virus (AKAV), affects livestock by causing life-threatening deformities or mortality of fetuses. Therefore, Akabane disease has led to notable economic losses in numerous countries, including Japan. In this short communication, a new T7 RNA polymerase-based AKAV reverse genetics system was developed. Using this system, in which three plasmids transcribing antigenomic RNAs were transfected into cells stably expressing T7 polymerase, we successfully reconstituted the live attenuated vaccine TS-C2 strain (named rTTT), and also generated a mutant AKAV (rTTTΔNSs) that lacked the gene encoding the nonstructural NSs protein, which is regarded as a virulence factor. Analysis of growth kinetics revealed that rTTTΔNSs grew at a much slower rate than the rTTT and TS-C2 virus. These results suggest that our established reverse genetics system is a powerful tool that can be used for AKAV vaccine studies with gene-manipulated viruses.

Keywords: Akabane virus; Reverse genetics; Vaccine.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Bunyaviridae Infections / veterinary
Bunyaviridae Infections / virology
Cell Line
Japan
Mutation
Orthobunyavirus / genetics*
Orthobunyavirus / growth & development
Orthobunyavirus / pathogenicity
Orthobunyavirus / physiology*
Reverse Genetics / methods*
Virulence Factors / genetics

Substances

Virulence Factors