生物界の暗黒物質「未知アーキア」の解明―分離培養で開拓する多様な新生物機能―

• Project/Area Number: 15H02419
• Research Category: Grant-in-Aid for Scientific Research (A)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Biodiversity/Systematics
• Research Institution: Japan Agency for Marine-Earth Science and Technology
• Principal Investigator: 井町 寛之 国立研究開発法人海洋研究開発機構, 深海・地殻内生物圏研究分野, 主任研究員 (20361933)
• Co-Investigator(Kenkyū-buntansha): 玉木 秀幸 国立研究開発法人産業技術総合研究所, 生命工学領域, 主任研究員 (00421842) ; 高木 善弘 国立研究開発法人海洋研究開発機構, 深海・地殻内生物圏研究分野, 主任技術研究員 (10399561) ; 諸野 祐樹 国立研究開発法人海洋研究開発機構, 高知コア研究所, グループリーダー代理 (30421845)
• Project Period (FY): 2015-04-01 – 2019-03-31
• Project Status: Granted (Fiscal Year 2018)
• Budget Amount: ¥40,170,000 (Direct Cost: ¥30,900,000、Indirect Cost: ¥9,270,000); Fiscal Year 2018: ¥7,150,000 (Direct Cost: ¥5,500,000、Indirect Cost: ¥1,650,000); Fiscal Year 2017: ¥10,010,000 (Direct Cost: ¥7,700,000、Indirect Cost: ¥2,310,000); Fiscal Year 2016: ¥9,880,000 (Direct Cost: ¥7,600,000、Indirect Cost: ¥2,280,000); Fiscal Year 2015: ¥13,130,000 (Direct Cost: ¥10,100,000、Indirect Cost: ¥3,030,000)
• Keywords: 微生物 / アーキア / 生物多様性 / バイオリアクター / 古細菌

Outline of Annual Research Achievements

集積培養に成功している嫌気的メタン酸化反応に関与する未知アーキアの完全長ゲノム配列をメタゲノム解析により決定した。さらに、培養条件の改善ならびに大量培養を行うことにより、RNA-Seq法による代謝発現解析を行うことができた。得られたゲノム情報、mRNAの発現情報および培養条件から、本未知アーキアの代謝経路についての推定を行い、利用可能なエネルギー基質等の重要な情報を得ることに成功した。さらに本未知アーキアの詳細な細胞形態や構造を調査するために、電子顕微鏡による観察を行い、その細胞内構造を明らかにした。
下降流懸垂型スポンジリアクターを利用して海底堆積物や天然ガス鹹水に生息する未知アーキアの培養を初年度から引き続き行った。その結果、新たに海底堆積物中に優占化している別の未知アーキアを高度に集積培養をすることに成功した。この未知アーキアの16S rRNA遺伝子に特異的なプライマーセットを使った定量PCRにより、本未知アーキアの増殖の確認と増殖条件の検討を進めることができた。

Current Status of Research Progress

2: Research has progressed on the whole more than it was originally planned.

Reason

集積培養ができていた嫌気的メタン酸化反応に関与する未知アーキアの完全長ゲノム決定ならびにRNA-Seq法による代謝発現解析を行い、培養条件と組み合わせて考えることで、本未知アーキアの代謝経路を推定できたことは大きな前進であった。加えて、新たな未知アーキアの高度な集積培養にも成功できたことから、「おおむね順調に進展している」と評価した。

Strategy for Future Research Activity

新たに培養に成功した未知アーキアのゲノム解析や電子顕微鏡による細胞形態観察を用いて行う。また、初年度から運転を行っている下降流懸垂型スポンジリアクターには多種類の未知アーキアが集積培養できているので、それぞれの未知アーキアの分離作業ならびにゲノム配列の取得を行う。

Research Products

• [Int'l Joint Research] University of Bremen(Germany)
• [Int'l Joint Research] National Taiwan University(Taiwan)
• [Int'l Joint Research] California Institute of Technology/Craig J. Vender Institute/University of Illinoi(United States of America)
• [Int'l Joint Research] University of Bremen(Germany)
• [Int'l Joint Research] National Taiwan University/National Cheng Kung University(Taiwan)
• [Int'l Joint Research] California Institute of Technology/Craig J. Vender Institute/University of South California(United States of America)
• [Journal Article] Production of biogenic manganese oxides coupled with methane oxidation in a bioreactor for removing metals from wastewater (2018)
  • Author(s): Matsushita Shuji、Komizo Daisuke、Cao Linh Thi Thuy、Aoi Yoshiteru、Kindaichi Tomonori、Ozaki Noriatsu、Imachi Hiroyuki、Ohashi Akiyoshi
  • Journal Title: Water Research Volume: 130 Pages: 224-233
  • DOI: 10.1016/j.watres.2017.11.063
  • Peer Reviewed
• [Journal Article] Fractionation of nitrogen isotopes during amino acid metabolism in heterotrophic and chemolithoautotrophic microbes across Eukarya, Bacteria and Archaea: Effects of nitrogen sources and metabolic pathways (2017)
  • Author(s): Yamaguchi Yasuhiko T.、Chikaraishi Yoshito、Takano Yoshinori、Ogawa Nanako O.、Imachi Hiroyuki、Yokoyama Yusuke、Ohkouchi Naohiko
  • Journal Title: Organic Geochemistry Volume: 111 Pages: 101-112
  • DOI: 10.1016/j.orggeochem.2017.04.004
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Biotic manganese oxidation coupled with methane oxidation using a continuous-flow bioreactor system under marine conditions (2017)
  • Author(s): Kato Shingo、Miyazaki Masayuki、Kikuchi Sakiko、Kashiwabara Teruhiko、Saito Yumi、Tasumi Eiji、Suzuki Katsuhiko、Takai Ken、Cao Linh Thi Thuy、Ohashi Akiyoshi、Imachi Hiroyuki
  • Journal Title: Water Science and Technology Volume: 76 Pages: 1781-1795
  • DOI: 10.2166/wst.2017.365
  • Peer Reviewed
• [Journal Article] Microbial Community Composition and Functional Capacity in a Terrestrial Ferruginous, Sulfate-Depleted Mud Volcano (2017)
  • Author(s): Tu Tzu-Hsuan、Wu Li-Wei、Lin Yu-Shih、Imachi Hiroyuki、Lin Li-Hung、Wang Pei-Ling
  • Journal Title: Frontiers in Microbiology Volume: 8
  • DOI: 10.3389/fmicb.2017.02137
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Variance and potential niche separation of microbial communities in subseafloor sediments off Shimokita Peninsula, Japan (2016)
  • Author(s): Takuro Nunoura, Yoshihiro Takaki, Shigeru Shimamura, Jungo Kakuta, Hiromi Kazama, Miho Hirai, Noriaki Masui, Hitoshi Tomaru, Yuki Morono, Hiroyuki Imachi, Fumio Inagaki, Ken Takai
  • Journal Title: Environmental Microbiology Volume: 18 Pages: 1-16
  • DOI: 10.1111/1462-2920.13096
  • Peer Reviewed
• [Journal Article] Sedimentibacter acidaminivorans sp. nov., an anaerobic, amino acids-utilizing bacterium isolated from marine subsurface sediment (2016)
  • Author(s): Imachi H, Sakai S, Kubota K, Miyazaki M, Saito Y and Takai K
  • Journal Title: International Journal of Systematic and Evoltionary Microbiology Volume: 66 Pages: 1293-1300
  • DOI: 10.1099/ijsem.0.000878
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Hydrogen and carbon isotope systematics in hydrogenotrophic methanogenesis under H2-limited and -enriched conditions: Implications for the origin of methane and its isotopic diagnosis (2016)
  • Author(s): Okumura T., Kawagucci S., Saito Y., Matsui Y., Takai K., Imachi H.
  • Journal Title: Progress in Earth and Planetary Science Volume: 3 Pages: 14-14
  • DOI: 10.1186/s40645-016-0088-3
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Nitrogen and oxygen isotope effects of ammonia oxidation by thermophilic Thaumarchaeota from a geothermal water stream (2016)
  • Author(s): Nishizawa M, Sakai S, Konno U, Nakahara N, Takaki Y, Saito Y, Imachi H, Tasumi E, Makabe A, Koba K and Takai K
  • Journal Title: Applied and Environmental Microbiology Volume: 82 Pages: 4492-4504
  • DOI: 10.1128/aem.00250-16
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Genomic composition and dynamics among Methanomicrobiales predict adaptation to contrasting environments (2016)
  • Author(s): Browne P, Tamaki H, Kyrpides N, Woyke T, Goodwin L, Imachi H, Brauer, S Yavitt J B, Liu W-T, Zinder S and Cadillo-Quiroz H
  • Journal Title: The ISME J Volume: 11 Pages: 87-99
  • DOI: 10.1038/ismej.2016.104
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Methanomicrobium antiquum sp. nov., a hydrogenotrophic methanogen isolated from deep sedimentary aquifers in a natural gas field (2016)
  • Author(s): Mochimaru H, Tamaki H, Katayama T, Imachi H, Sakata S and Kamagata Y
  • Journal Title: Int. J. Syst. Evol. Microbiol. Volume: 66 Pages: 4873-4877
  • DOI: 10.1099/ijsem.0.001444
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Exploring deep microbial life in coal-bearing sediments down to ~2.5km below the oceanfloor (2015)
  • Author(s): INAGAKI FUMIO, Kai-Uwe Hinrichs,KUBO YUSUKE,Marshall W. Bowles,Verena B. Heuer,Wei-Li Hong,HOSHINO TATSUHIKO,IJIRI AKIRA,IMACHI HIROYUKI,ITO MOTOO,KANEKO MASANORI,Mark A. Lever,Lin Yu-Shin,Barbara A. Methe,Morita Sumito,MORONO YUKI,et al.
  • Journal Title: Science Volume: 349 Pages: 420-424
  • DOI: 10.1126/science.aaa6882
  • Peer Reviewed / Int'l Joint Research / Acknowledgement Compliant
• [Presentation] 微生物暗黒物質は暗黒ではない-海底下生命圏からの未培養微生物の培養を例として- (2017)
  • Author(s): 井町寛之
  • Organizer: 環境微生物系学会合同大会2017
  • Invited
• [Presentation] Cultivation of uncultured subseafloor microbes using down-flow hanging sponge (DHS) bioreactors (2017)
  • Author(s): Imachi H
  • Organizer: Deep Life Cultivation workshop
  • Int'l Joint Research / Invited
• [Presentation] Cultivation of microbial dark matters from marine sediments: novel bacteria affiliated with uncultured groups of the phylum Chloroflexi. (2017)
  • Author(s): Nakahara N, Yoshida N, Tasumi E, Nobu M, Miyazaki M, Takaki Y, Sakai S, Tamaki H, Yamaguchi T, Takai K and Imachi H
  • Organizer: JpGU-AGU Joint Meeting 2017
  • Int'l Joint Research / Invited
• [Presentation] 海底堆積物から分離した新規Chlorolexi門細菌の生理代謝機能の解明 (2017)
  • Author(s): 中原望・吉田奈央子・田角栄二・高木善弘・酒井早苗・宮崎征行・山口隆司・高井研・井町寛之
  • Organizer: 環境微生物系学会合同大会2017
• [Presentation] 海底堆積物に生息する微生物ダークマターChloroflexi門細菌の分離・培養 (2016)
  • Author(s): 中原望・小西優・高木善弘・酒井早苗・玉木秀幸・山口隆司・高井研・井町寛之
  • Organizer: 第31回日本微生物生態学会大会
  • Place of Presentation: 横須賀市文化会館 (横須賀、神奈川)
  • Year and Date: 2016-10-23
• [Presentation] Cultivation of uncultured Chloroflexi bacteria, a microbial dark matter in subseafloor biosphere (2016)
  • Author(s): Nakahara N, Takai Y, Konishi M, Tamaki H, Yamaguchi T, Takai K and Imachi H
  • Organizer: The 8th Asian Symposium on Microbial Ecology
  • Place of Presentation: National Taiwan University (Taipei, Taiwan)
  • Year and Date: 2016-10-01
  • Int'l Joint Research
• [Presentation] Identifying methanogenic community members obtained from 2-km deep coalbed (2016)
  • Author(s): Identifying methanogenic community members obtained from 2-km deep coalbed
  • Organizer: International Symposium on Microbial Ecology (ISME) 2016
  • Place of Presentation: Palais des congres de Montreal (Montreal, Canada)
  • Year and Date: 2016-08-23
  • Int'l Joint Research
• [Presentation] Cultivation of previously uncultured Chloroflexi, a bacterial dark matter in subseafloor biosphere (2016)
  • Author(s): Nakahara N, Takai Y, Konishi M, Tamaki H, Yamaguchi T, Takai K and Imachi H
  • Organizer: International Symposium on Microbial Ecology (ISME) 2016
  • Place of Presentation: Palais des congres de Montreal (Montreal, Canada)
  • Year and Date: 2016-08-23
  • Int'l Joint Research
• [Presentation] Identifying 2 km-deep methanogenic community members using a long-term bioreactor cultivation (2016)
  • Author(s): Imachi H, Tasumi E, Tu T-H, Ijiri A, Morono Y, Ishii S, Methe BA, Takai K and Inagaki F
  • Organizer: Goldschimidt2016
  • Place of Presentation: Pacifico Yokohama (Yokohama, Japan)
  • Year and Date: 2016-06-29
  • Int'l Joint Research
• [Book] Methanocellales. In Bergey’s Manual of Systematics of Archaea and Bacteria (2016)
  • Author(s): Sakai S and Imachi H
  • Total Pages: 1
  • Publisher: John Wiley & Sons, Inc.
• [Book] Methanocellaceae. In Bergey’s Manual of Systematics of Archaea and Bacteria (2016)
  • Author(s): Sakai S and Imachi H
  • Total Pages: 1
  • Publisher: John Wiley & Sons, Inc.
• [Book] Methanocella. In Bergey’s Manual of Systematics of Archaea and Bacteria (2016)
  • Author(s): Sakai S and Imachi H
  • Total Pages: 6
  • Publisher: John Wiley & Sons, Inc.
• [Book] Methanoregulaceae. In Bergey’s Manual of Systematics of Archaea and Bacteria (2016)
  • Author(s): Imachi H and Sakai S
  • Total Pages: 4
  • Publisher: John Wiley & Sons, Inc.
• [Book] Methanolinea. In Bergey’s Manual of Systematics of Archaea and Bacteria (2016)
  • Author(s): Imachi H and Sakai S
  • Total Pages: 4
  • Publisher: John Wiley & Sons, Inc.