Development of a highly sensitive method for detection of viruses using the metagenome analysis

• Project/Area Number: 16K21140
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Infectious disease medicine; Laboratory medicine
• Research Institution: Osaka University
• Principal Investigator: Motooka Daisuke 大阪大学, 微生物病研究所, 特任助教(常勤) (10636830)
• Project Period (FY): 2016-04-01 – 2019-03-31
• Project Status: Completed (Fiscal Year 2018)
• Budget Amount: ¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000); Fiscal Year 2017: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000); Fiscal Year 2016: ¥2,470,000 (Direct Cost: ¥1,900,000、Indirect Cost: ¥570,000)
• Keywords: メタゲノム / 感染症診断

Outline of Final Research Achievements

Clinical metagenomics using high-throughput sequencing is widely applied to comprehensively detect pathogens in infectious diseases. The pathogen identification approach is based on a similarity search of known nucleotides along with taxonomic assignment of sequence reads produced by high-throughput sequencing. However, because the relative abundance of pathogens in the sequence reads is very low (generally < 0.1%), we need to sequence massive numbers of reads to detect pathogens. Consequently, it is difficult to improve sensitivity, cut cost, and accelerate data analysis. To resolve this problem, we developed a highly sensitive method to detect pathogenic viruses from metagenomics data. We performed experiments to concentrate and purify several types of viruses (e.g., influenza virus) from culture supernatants or clinical samples using a hydroxyapatite column and succeeded in obtaining a maximum of 256-fold concentration of the viruses.

Academic Significance and Societal Importance of the Research Achievements

感染症対策における臨床検体中の病原体を探索する方法として、次世代シークエンサーを用いたメタゲノム解析法が利用されている。本研究では、臨床検体中のウイルスの濃縮法開発に取り組み、カラム精製法や遊離核酸除去法を検討し、100倍以上のウイルス濃縮に成功した。高感度、迅速かつ安価な病原体検出法の確立に必須の技術であり、網羅的な病原体探索法の臨床応用にとって有用な成果が得られた。

Research Products

• [Journal Article] Urinary tract infection due to anaerobic bacteria in a two-month-old infant. (2019)
  • Author(s): Tao Chiaki、Kinoshita Noriko、Shoji Kensuke、Motooka Daisuke、Nakamura Shota、Eura Rumiko、Ueoka Katsuhiko、Kubota Mitsuru、Ishiguro Akira、Miyairi Isao
  • Journal Title: J Infect Chemother Volume: 25 Pages: 368-370
  • DOI: 10.1016/j.jiac.2018.11.016
  • Peer Reviewed / Open Access
• [Journal Article] WU polyomavirus detected in children with severe respiratory failure. (2018)
  • Author(s): Uda Kazuhiro、Koyama-Wakai Chitose、Shoji Kensuke、Iwase Noriyasu、Motooka Daisuke、Nakamura Shota、Miyairi Isao
  • Journal Title: J Clin Virol. Volume: 107 Pages: 25-28
  • DOI: 10.1016/j.jcv.2018.08.003
  • Peer Reviewed / Open Access
• [Journal Article] The clinical and phylogenetic investigation for a nosocomial outbreak of respiratory syncytial virus infection in an adult hemato-oncology unit (2017)
  • Author(s): Nabeya D, Kinjo T, Parrott GL, Uehara A, Motooka D, Nakamura S, Nahar S, Nakachi S, Nakamatsu M, Maeshiro S, Haranaga S, Tateyama M, Tomoyose T, Masuzaki H, Horii T, Fujita J
  • Journal Title: J Med Virol Volume: 印刷中
  • DOI: 10.1002/jmv.24800
  • Peer Reviewed
• [Journal Article] Sensitivity of Next-Generation Sequencing Metagenomic Analysis for Detection of RNA and DNA Viruses in Cerebrospinal Fluid: The Confounding Effect of Background Contamination (2017)
  • Author(s): Bukowska-Osko I, Perlejewski K, Nakamura S, Motooka D, Stokowy T, Kosinska J, Popiel M, Ploski R, Horban A, Lipowski D, Caraballo Cortes K, Pawelczyk A, Demkow U, Stepien A, Radkowski M, Laskus T
  • Journal Title: Adv Exp Med Biol Volume: 944 Pages: 53-62
  • Peer Reviewed / Int'l Joint Research
• [Book] 実験医学別冊 今すぐ始める！メタゲノム解析 実験プロトコール (2016)
  • Author(s): 元岡大祐、中村昇太、飯田哲也、堀井俊宏
  • Total Pages: 8
  • Publisher: 羊土社