Generation of a GFP Reporter Akabane Virus with Enhanced Fluorescence Intensity by Modification of Artificial Ambisense S Genome
Akiko Takenaka-Uema,
Shin Murakami,
Nanako Ushio,
Tomoya Kobayashi-Kitamura,
Masashi Uema,
Kazuyuki Uchida,
Taisuke Horimoto
Affiliations
Akiko Takenaka-Uema
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
Shin Murakami
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
Nanako Ushio
Department of Veterinary Pathology, Graduate School of Agriculture and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
Tomoya Kobayashi-Kitamura
Department of Veterinary Pathology, Graduate School of Agriculture and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
Masashi Uema
Division of Biomedical Food Research, National Institute of Health Sciences, 3-25-26 Tonomachi, Kawasaki-ku, Kawasaki 210-9501, Japan
Kazuyuki Uchida
Department of Veterinary Pathology, Graduate School of Agriculture and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
Taisuke Horimoto
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
We previously generated a recombinant reporter Akabane virus expressing enhanced green fluorescence protein (eGFP-AKAV), with an artificial S genome encoding eGFP in the ambisense RNA. Although the eGFP-AKAV was able to detect infected cells in in vivo histopathological study, its fluorescent signal was too weak to apply to in vivo imaging study. Here, we successfully generated a modified reporter, eGFP/38-AKAV, with 38-nucleotide deletion of the internal region of the 5′ untranslated region of S RNA. The eGFP/38-AKAV expressed higher intensity of eGFP fluorescence both in vitro and in vivo than the original eGFP-AKAV did. In addition, eGFP/38-AKAV was pathogenic in mice at a comparable level to that in wild-type AKAV. In the mice infected with eGFP/38-AKAV, the fluorescent signals, i.e., the virus-infected cells, were detected in the central nervous system using the whole-organ imaging. Our findings indicate that eGFP/38-AKAV could be used as a powerful tool to help elucidate the dynamics of AKAV in vivo.
