人工ヌクレアーゼTALENを用いた遺伝子ノックアウトマダニ感染症モデルの作出

2015 Fiscal Year Annual Research Report

• Project/Area Number: 26660229
• Research Institution: Kagoshima University
• Principal Investigator: 田仲 哲也 鹿児島大学, 農水産獣医学域獣医学系, 准教授 (00322842)
• Project Period (FY): 2014-04-01 – 2016-03-31
• Keywords: マダニ / ゲノム / プロモーター領域 / アクチン / チュブリン / ディフェンシン / PGK

Outline of Annual Research Achievements

[全体構想] マダニの遺伝子機能の解析ではRNA干渉法による遺伝子破壊が用いられてきたが、ノックアウトの技術は導入されていない。そこで、申請者らは人工ヌクレアーゼに着目し、マダニの標的遺伝子ノックアウト技術の確立を試みることが本研究の主要目的である。[今まで得られた研究成果]平成27年度に同定したアクチンやフェリチンプロモ-ターを用いてルシフェラーゼ活性を調べたが、マダニやマダニ細胞で活性は低かった。しかし、PGKプロモーターを用いてマダニ細胞でのルシフェラーゼ活性を調べたところ、活性は高かった。以上の結果から、PGKプロモーターを利用して外来遺伝子を組み込めば、マダニ体内に外来遺伝子を導入できる可能性が示唆された。

Research Products

• [Journal Article] Course of induced infection by Eimeria krijgsmanni in immunocompetent and immunodeficient mice (2016)
  • Author(s): Ono, U., Matsubayashi, M., Tsujio, M., Mizuno, M., Tanaka, T., Masatani, T., Matsui, T. and Matsuo, T.
  • Journal Title: Parasitol. Res. Volume: 115 (1) Pages: 211-215
  • DOI: 10.1007/s00436-015-4737-z.
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Role of the tumor necrosis factor receptor-associated factor-type zinc finger domain containing protein 1 (TRAFD1) from the hard tick Haemaphysalis longicornis in immunity against bacterial infection (2016)
  • Author(s): Takechi, R., Galay, R. L., Maeda, H., Matsuo, T., Kusakisako, K., Talactac, M. R., Mochizuki, M., Fujisaki, K. and Tanaka, T.
  • Journal Title: Ticks Tick Borne Dis. Volume: 7 (1) Pages: 36-45
  • DOI: 10.1016/j.ttbdis.2015.08.002.
  • Peer Reviewed / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Functional analysis of recombinant 2-Cys peroxiredoxin from the hard tick Haemaphysalis longicornis (2016)
  • Author(s): Kusakisako, K., Masatani, T., Miyata, T., Galay, R. L., Maeda, H., Talactac, M. R., Tsuji, N., Mochizuki, M., Fujisaki, K. and Tanaka, T.
  • Journal Title: Insect Mol. Biol. Volume: 25 (1) Pages: 16-23
  • DOI: 10.1111/imb.12193.
  • Peer Reviewed / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] A novel C-type lectin with triple carbohydrate recognition domains has critical roles for the hard tick Haemaphysalis longicornis against Gram-negative bacteria (2016)
  • Author(s): Maeda, H., Miyata, T., Kusakisako, K., Galay, R. L., Talactac, M. R., Umemiya-Shirafuji, R., Mochizuki, M., Fujisaki, K. and Tanaka, T.
  • Journal Title: Dev. Comp. Immunol. Volume: 57 Pages: 38-47
  • DOI: 10.1016/j.dci.2015.12.015.
  • Peer Reviewed / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Impaired cellular immune response to injected bacteria after knockdown of ferritin genes in the hard tick Haemaphysalis longicornis (2016)
  • Author(s): Galay, R. L., Takechi, R., Umemiya-Shirafuji, R., Talactac, M. R., Maeda, H., Kusakisako, K., Mochizuki, M., Fujisaki, K. and Tanaka, T.
  • Journal Title: Parasitol. Int. Volume: 65 (3) Pages: 251-257
  • DOI: 10.1016/j.parint.2016.01.007.
  • Peer Reviewed / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Virucidal activity of Haemaphysalis longicornis longicin P4 peptide against tick-borne encephalitis virus surrogate Langat virus (2016)
  • Author(s): Talactac, M. R., Yoshii, K., Maeda, H., Kusakisako, K., Hernandez, E. P., Tsuji, N., Fujisaki, K., Galay, R. L., Tanaka, T. and Mochizuki, M.
  • Journal Title: Parasit. Vectors Volume: 9 (1) Pages: 59
  • DOI: 10.1186/s13071-016-1344-5.
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Improvement of the cryopreservation method for the Babesia gibsoni parasite by using commercial freezing media (2016)
  • Author(s): Kusakisako, K., Masatani, T., Yada Y., Talactac, M. R., Hernandez, E. P., Maeda, H., Mochizuki, M. and Tanaka, T.
  • Journal Title: Parasitol. Int. Volume: In press Pages: In press
  • DOI: 10.1016/j.parint.2016.02.012.
  • Peer Reviewed / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Induction of gene silencing in Haemaphysalis longicornis ticks through immersion in double-stranded RNA (2016)
  • Author(s): Galay, R. L., Hernandez, E. P., Talactac, M. R., Maeda, H., Kusakisako, K., Umemiya-Shirafuji, R., Mochizuki, M., Fujisaki, K. and Tanaka, T.
  • Journal Title: Ticks Tick Borne Dis. Volume: In press Pages: In press
  • DOI: 10.1016/j.ttbdis.2016.03.018.
  • Peer Reviewed / Int'l Joint Research / Acknowledgement Compliant
• [Presentation] 病原微生物感染に関与するマダニ生物活性分子について (2015)
  • Author(s): 田仲哲也
  • Organizer: 日本衛生動物学会・日本ダニ学会共催 公開シンポジウム
  • Place of Presentation: 法政大学(千代田区、東京都)
  • Year and Date: 2015-09-11 – 2015-09-13
  • Invited
• [Presentation] Virucidal activity of Haemaphysalis longicornis longicin P4 peptide against tick-borne encephalitis virus surrogate Langat virus (2015)
  • Author(s): Melbourne Rio Talactac, 好井健太朗, 辻 尚利, 藤崎幸蔵, 田仲哲也, 望月雅美
  • Organizer: 第158回日本獣医学会学術集会
  • Place of Presentation: 北里大学(八戸市、青森)
  • Year and Date: 2015-09-07 – 2015-09-09
• [Presentation] Knockdown of ferritin genes increases the susceptibility of the hard tick Haemaphysalis longicornis to bacterial infection (2015)
  • Author(s): Tetsuya Tanaka, Remil Linggatong Galay, Rie Takechi, Rika Umemiya-Shirafuji, Melbourne Rio Talactac, Hiroki Maeda, Kodai Kusakisako, Masami Mochizuki, Kozo Fujisaki
  • Organizer: World Association for the Advancement of Veterinary Parasitology Congress 2015
  • Place of Presentation: The Arena and Convention Centre(Liverpool, UK)
  • Year and Date: 2015-08-16 – 2015-08-20
  • Int'l Joint Research
• [Presentation] Decreased resistance of Haemaphysalis longicornis ticks to bacterialchallenge after knockdown of ferritin molecules (2015)
  • Author(s): Remil Linggatong Galay, 武智理恵, 白藤(梅宮)梨可, Melbourne Rio Talactac, 前田大輝, 草木迫浩大, 望月雅美, 藤崎幸蔵, 田仲哲也
  • Organizer: 第23回SADI
  • Place of Presentation: 名取市文化会館(名取市、宮城)
  • Year and Date: 2015-06-26 – 2015-06-28
• [Presentation] フタトゲチマダニ由来ロイシンリッチリピート保有タンパク質(HlLRR)の発現動態とバベシア原虫に及ぼす影響 (2015)
  • Author(s): 田仲哲也, 栗巣孔士, 前田大輝, 宮田 健, 草木迫浩大, Remil Linggatong Galay, Melbourne Rio Talactac, 望月雅美, 藤崎幸蔵
  • Organizer: 第23回SADI
  • Place of Presentation: 名取市文化会館(名取市、宮城)
  • Year and Date: 2015-06-26 – 2015-06-28
• [Book] Vaccine Design: Methods and Protocols (2016)
  • Author(s): Galay, R. L., Miyata, T., Umemiya-Shirafuji, R., Mochizuki, M., Fujisaki, K. and Tanaka, T.
  • Total Pages: 848
  • Publisher: Springer New York
• [Book] RNA Interference (2016)
  • Author(s): Galay, R. L., Miyata, T., Umemiya-Shirafuji, R., Mochizuki, M., Fujisaki, K. and Tanaka, T.
  • Total Pages: 460
  • Publisher: InTech-Open Access Publisher
• [Remarks] 鹿児島大学 共同獣医学部獣医学科 病態予防獣医学講座 感染症学分野
  • URL: http://www.vet.kagoshima-u.ac.jp/kadai/V-infection/sinkoukansen/top.html