Metagenomic virus detection in clinical specimens using high-throughput sequencers

• Project/Area Number: 17KK0170
• Research Category: Fund for the Promotion of Joint International Research (Fostering Joint International Research)
• Allocation Type: Multi-year Fund
• Research Field: Epidemiology and preventive medicine
• Research Institution: Nagasaki University
• Principal Investigator: KUROSAKI Yohei 長崎大学, 感染症共同研究拠点, 准教授 (40415443)
• Project Period (FY): 2018 – 2019
• Project Status: Completed (Fiscal Year 2019)
• Budget Amount: ¥14,430,000 (Direct Cost: ¥11,100,000、Indirect Cost: ¥3,330,000)
• Keywords: ウイルス / 出血熱 / 次世代シークエンス / ナノポアシークエンス / メタゲノム / 診断 / 分子疫学 / 疫学 / ウイルス進化

Outline of Final Research Achievements

For a comprehensive detection of the viruses in clinical samples and virus surveillance in virus outbreak settings, we established a method of viral metagenomic analysis using high-throughput sequencing technologies including next-generation sequencer (NGS) and nanopore sequencer MinION. Using whole genome amplification and Illumina NGS, we determined a genome sequence of Measles virus in the Ebola-negative samples collected in the endemic of Ebola virus diseases in West Africa. We developed a rapid viral genome sequencing for Crimean-Congo hemorrhagic fever virus (CCHFV) using target genome amplification by tiling PCR, and nanopore sequencer MinION. We retrospectively determined the CCHFV sequences in the ticks and human blood samples collected in the virus endemic area in Tajikistan. Our methods can be used for rapid viral genome sequencing and surveillance of the virus circulating in settings of virus outbreaks.

Academic Significance and Societal Importance of the Research Achievements

新たなウイルス感染症のアウトブレイクにおいて、原因ウイルスをいち早く同定し、そのゲノム配列情報からウイルス学的性状を把握し、感染源を推定することは極めて重要である。ウイルス流行時において、個々の分離株のゲノム配列情報は、疫学情報から推定されるウイルス拡散経路を裏付け、より詳細な伝播モデルの推定を可能にする。今回開発したハイスループットシークエンサーによる網羅的なウイルス検出法、並びにナノポアシークエンスを用いたウイルスの迅速ゲノム配列決定法は、現在まさに流行している新型コロナウイルスのような新たなウイルス感染症の発生時にも活用できる技術であり、感染症対策に資することが期待できる。

Research Products

• [Int'l Joint Research] 英国公衆衛生局(英国) (2018)
• [Journal Article] エボラ診断薬とその現場での役割 (2019)
  • Author(s): 黒崎陽平
  • Journal Title: 最新医学 Volume: 74 Pages: 548-554
• [Journal Article] Development of an RT-LAMP assay for the detection of Lassa viruses in southeast and south-central Nigeria. (2019)
  • Author(s): Pemba CM, Kurosaki Y, Yoshikawa R, Oloniniyi OK, Urata S, Sueyoshi M, Zadeh VR, Nwafor I, Iroezindu MO, Ajayi NA, Chukwubike CM, Chika-Igwenyi NM, Ndu AC, Nwidi DU, Maehira Y, Unigwe US, Ojide CK, Onwe EO, Yasuda J.
  • Journal Title: Journal of Virological Methods Volume: 269 Pages: 30-37
  • DOI: 10.1016/j.jviromet.2019.04.010
  • Peer Reviewed / Int'l Joint Research
• [Presentation] Nanopore sequencing for genome surveillance of Crimean-Congo hemorrhagic fever virus (2019)
  • Author(s): Yohei Kurosaki, Babak Afrough, John Chamberlain, Roger Hewson, Jiro Yasuda
  • Organizer: 第67回日本ウイルス学会学術集会
• [Presentation] Development of an RT-LAMP assay for the detection of Lassa viruses in southeast and south-central Nigeria (2019)
  • Author(s): C. Pemba, Y. Kurosaki, R. Yoshikawa, I. Nwafor, O. K. Oloniniyi, S. Urata, M. Sueyoshi, V. R. Zadeh, M. O. Iroezindu, N. Ajayi, C. M. Chukwubike, N. M. Chika-Igwenyi, A. C. Ndu, D. U. Nwidi, Y. Maehira, U. S. Unigwe, C. K. Ojide, E. O. Onwe, J. Yasuda
  • Organizer: Lassa fever First International Conference
  • Int'l Joint Research
• [Presentation] DIFFERENT EFFECTS OF TWO MUTATIONS ON THE INFECTIVITY OF EBOLA VIRUS GLYCOPROTEIN IN NINE MAMMALIAN SPECIES (2018)
  • Author(s): Y. Kurosaki, M. T. Ueda, Y. Nakano, J. Yasuda, Y. Koyanagi, K. Sato, S. Nakagawa
  • Organizer: The 2018 Negative Strand RNA Virus meeting
  • Int'l Joint Research