Analysis of hepatitis C virus sequence quasi species by NGS

2013 Fiscal Year Final Research Report

• Project/Area Number: 23659407
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Gastroenterology
• Research Institution: National Institute of Infectious Diseases
• Principal Investigator: WAKITA Takaji 国立感染症研究所, ウイルス第二部, 部長 (40280789)
• Project Period (FY): 2011 – 2012
• Keywords: 肝炎ウイルス / 次世代シーケンス / quasispecies

Research Abstract

Hepatitis C Virus (HCV) genome accumulates nucleotide substitutions because of low viral RNA polymerase fidelity and high viral replication ratio. This quasispecies nature has been considered to contribute for its pathogenesis or viral escape. We constructed a method to determine the full-length viral genome RNA species from patient serum combined next-generation sequencing and capillary sequencing. We found independent dominant species co-existed in one patient serum. Polymorphisms related to the favorable response of interferon sensitivity were gathered into one species. Sequential species were formed same clusters. We thus considered that both species might have been mixture and survived the bottleneck events when the patient was infected, and then evolved individually in the patient. In conclusion, our study demonstrated that considering each full-length species in patient serum is important for understanding HCV life cycle.

Research Products

• [Journal Article] Replication of hepatitis C virus genotype 3a in cultured cells (2013)
  • Author(s): Saeed M, Gondeau C, Hmwe S, Yokokawa H, Date T, Suzuki T, Kato T, Maurel P, Wakita T
  • Journal Title: Gastroenterology Volume: 144(1) Pages: 56-58.e7
  • DOI: 10.1053/j.gastro.2012.09.017.
• [Journal Article] Neutralizing antibodies induced by cell culture-derived hepatitis C virus protect against infection in mice (2013)
  • Author(s): Akazawa D, Moriyama M, Yokokawa H, Omi N, Watanabe N, Date T, Morikawa K, Aizaki H, Ishii K, Kato T, Mochizuki H, Nakamura N, Wakita T
  • Journal Title: Gastroenterology Volume: 145(2) Pages: 447-55.e1-4
  • DOI: 10.1053/j.gastro.2013.05.007.
• [Journal Article] Specific inhibition of hepatitis C virus entry into host hepatocytes by fungi-derived sulochrin and its derivatives (2013)
  • Author(s): Nakajima S, Watashi K, Kamisuki S, Tsukuda S, Takemoto K, Matsuda M, Suzuki R, Aizaki H, Sugawara F, Wakita T
  • Journal Title: Biochem Biophys Res Commun Volume: 440(4) Pages: 515-20
  • DOI: 10.1016/j.bbrc.2013.09.100.
• [Journal Article] Replication and infectivity of a novel genotype 1b hepatitis C virus clone (2012)
  • Author(s): Date T, Morikawa K, Tanaka Y, Tanaka-Kaneko K, Sata T, Mizokami M, Wakita T
  • Journal Title: Microbiol Immunol Volume: 56(5) Pages: 308-17
  • DOI: 10.1111/j.1348-0421.2012.00437.x.
• [Journal Article] Detergent-induced activation of the hepatitis C virus genotype 1b RNA polymerase (2012)
  • Author(s): Weng L, Kohara M, Wakita T, Shimotohno K, Toyoda T
  • Journal Title: Gene Volume: 496(2) Pages: 79-87
  • DOI: 10.1016/j.gene.2012.01.044.
• [Journal Article] Novel cell culture-adapted genotype 2a hepatitis C virus infectious clone (2012)
  • Author(s): Date T, Kato T, Kato J, Takahashi H, Morikawa K, Akazawa D, Murayama A,Tanaka-Kaneko K, Sata T, Tanaka Y, Mizokami M, Wakita T
  • Journal Title: J Virol Volume: 86(19) Pages: 10805-20
  • DOI: 10.1128/JVI.07235-11.
• [Journal Article] Trans-complemented hepatitis C virus particles as a versatile tool forstudy of virus assembly and infection (2012)
  • Author(s): Suzuki R, Saito K, Kato T, Shirakura M, Akazawa D, Ishii K, Aizaki H, Kanegae Y, Matsuura Y, Saito I, Wakita T, Suzuki T
  • Journal Title: Virology Volume: 432(1) Pages: 29-38
  • DOI: 10.1016/j.virol.2012.05.033.
• [Journal Article] Visualization and measurement of ATP levels in living cells replicating hepatitis C virus genome RNA (2012)
  • Author(s): Ando T, Imamura H, Suzuki R, Aizaki H, Watanabe T, Wakita T, Suzuki T
  • Journal Title: PLoS Pathog Volume: 8(3) Pages: e1002561
  • DOI: 10.1371/journal.ppat.1002561.
• [Journal Article] 25-Hydroxyvitamin D3 suppresses hepatitis C virus production (2012)
  • Author(s): Matsumura T, Kato T, Sugiyama N, Tasaka-Fujita M, Murayama A, Masaki T, Wakita T, Imawari M
  • Journal Title: Hepatology Volume: 56(4) Pages: 1231-9
  • DOI: 10.1002/hep.25763.
• [Journal Article] Potent and selective inhibition of hepatitis C virus replication by novel phenanthridinone derivatives (2011)
  • Author(s): Salim MT, Aoyama H, Sugita K, Watashi K, Wakita T, Hamasaki T, Okamoto M, Urata Y, Hashimoto Y, Baba M
  • Journal Title: Biochem Biophys Res Commun Volume: 415(4) Pages: 714-9
  • DOI: 10.1016/j.bbrc.2011.10.153.
• [Journal Article] Genetic Variation of the IL-28B Promoter Affecting Gene Expression (2011)
  • Author(s): Sugiyama M, Tanaka Y, Wakita T, Nakanishi M, Mizokami M
  • Journal Title: PLoS One Volume: 6(10) Pages: e26620
  • DOI: 10.1371/journal.pone.0026620.
• [Journal Article] Hepatitis C virus reveals a novel early control in acute immune response (2011)
  • Author(s): Arnaud N, Dabo S, Akazawa D, Fukasawa M, Shinkai-Ouchi F, Hugon J, Wakita T, Meurs EF
  • Journal Title: PLoS Pathog Volume: 7(10) Pages: e1002289
  • DOI: 10.1371/journal.ppat.1002289.
• [Journal Article] Role of the endoplasmic reticulum-associated degradation ERAD) pathway in degradation of hepatitis C virus envelope proteins and production of virus particles (2011)
  • Author(s): Saeed M, Suzuki R, Watanabe N, Masaki T, Tomonaga M, Muhammad A, Kato T, Matsuura Y, Watanabe H, Wakita T, Suzuki T
  • Journal Title: J Biol Chem Volume: 286(43) Pages: 37264-73
  • DOI: 10.1074/jbc.M111.259085.
• [Journal Article] Development of recombinant hepatitis C virus with NS5A from strains of genotypes 1 and 2 (2011)
  • Author(s): Okamoto Y, Masaki T, Murayama A, Munakata T, Nomoto A, Nakamoto S, Yokosuka O, Watanabe H, Wakita T, Kato T
  • Journal Title: Biochem Biophys Res Commun Volume: 410(3) Pages: 404-9
  • DOI: 10.1016/j.bbrc.2011.05.144.
• [Journal Article] Structural requirements of virion-associated cholesterol for infectivity, buoyant density and apolipoprotein association of hepatitis C virus (2011)
  • Author(s): Yamamoto M, Aizaki H, Fukasawa M, Teraoka T, Miyamura T, Wakita T, Suzuki T
  • Journal Title: J Gen Virol Volume: 92(Pt 9) Pages: 2082-7
  • DOI: 10.1099/vir.0.032391-0.
• [Journal Article] Production and characterization of HCV particles from serum-free culture (2011)
  • Author(s): Akazawa D, Morikawa K, Omi N, Takahashi H, Nakamura N, Mochizuki H, Date T, Ishii K, Suzuki T, Wakita T
  • Journal Title: Vaccine Volume: 29(29-30) Pages: 4821-8
  • DOI: 10.1016/j.vaccine.2011.04.069.
• [Journal Article] In vivo adaptation of hepatitis C virus in chimpanzees for efficient virus production and evasion of apoptosis (2011)
  • Author(s): Saeed M, Shiina M, Date T, Akazawa D, Watanabe N, Murayama A, Suzuki T, Watanabe H, Hiraga N, Imamura M, Chayama K, Choi Y, Krawczynski K, Liang TJ, Wakita T, Kato T
  • Journal Title: Hepatology Volume: 54(2) Pages: 425-33
  • DOI: 10.1002/hep.24399.
• [Journal Article] Hokuriku Liver Study Group. Malnutrition impairs interferon signaling through mTOR and FoxO pathways in patients with chronic hepatitis C (2011)
  • Author(s): Honda M, Takehana K, Sakai A, Tagata Y, Shirasaki T, Nishitani S, Muramatsu T, Yamashita T, Nakamoto Y, Mizukoshi E, Sakai Y, Yamashita T, Nakamura M, Shimakami T, Yi M, Lemon SM, Suzuki T, Wakita T, Kaneko S
  • Journal Title: Gastroenterology Volume: 141(1) Pages: 128-40, 140.e1-2
  • DOI: 10.1053/j.gastro.2011.03.051.
• [Journal Article] Development of mouse hepatocyte lines permissive for hepatitis C virus (HCV) (2011)
  • Author(s): Aly HH, Oshiumi H, Shime H, Matsumoto M, Wakita T, Shimotohno K, Seya T
  • Journal Title: PLoS One Volume: 6(6) Pages: e21284
  • DOI: 10.1371/journal.pone.0021284.
• [Presentation] Molecular Biology and Experimental Models of HCV (2013)
  • Author(s): T Wakita
  • Organizer: 19th Annual KASL Meeting
  • Place of Presentation: Sheraton Grande Walkerhill Hotel, Seoul, Korea
  • Year and Date: 20130613-15
• [Presentation] 患者血清中におけるHCV ゲノム多種性の存在形式 (2013)
  • Author(s): 脇田隆字、安東友美、林和彦、杉山真也、石上雅敏、片野義明、後藤秀実、溝上雅史、黒田誠、相崎英樹
  • Organizer: 第49回日本肝臓学会総会
  • Place of Presentation: 京王プラザホテル、ワークショップ3「ウイルス肝炎の新潮流」
  • Year and Date: 20130606-07
• [Presentation] Independent Evolution of Multi-dominant Viral Genome Species of Hepatitis C Virus (2012)
  • Author(s): T Wakita
  • Organizer: The 10th JSH Single Topic Conference, "Hepatitis C : Best Practice Based on Science"
  • Place of Presentation: Keio Plaza Hotel, Tokyo, Japan
  • Year and Date: 20121121-22
• [Presentation] Independent evolution of multi-dominant viral genome species observed in a single HCV carrier (2012)
  • Author(s): T Ando, H Aizaki, M Sugiyama, M Mizokami, M Kuroda, T Wakita
  • Organizer: 19th International Meeting on Hepatitis C and Related Viruses
  • Place of Presentation: Palazzo del Cinema, Lido-Venice, Italy
  • Year and Date: 20121005-09
• [Presentation] Hepatitis C virus replication models and anti-viral development (2012)
  • Author(s): T Wakita
  • Organizer: The 1^<st> International Symposium on Latent TGF-beta Activation Reaction
  • Place of Presentation: RIKEN Kobe Inst, Ctr. For Delop Biol, Auditorium, Kobe
  • Year and Date: 2012-02-25
• [Presentation] Hepatitis C virus replication in vitro and persistent infection in vivo : mechanistic analysis and antiviral development (2012)
  • Author(s): T Wakita
  • Organizer: Challenges to overcome Emerging Infectious Diseases in South-eastern Asia
  • Place of Presentation: Siran Kaikan, Kyoto Uninversity. Kyoto
  • Year and Date: 2012-01-13
• [Presentation] Hepatitis C virus replication in vitro and persistent infection in vivo : mechanistic analysis and antiviral development (2011)
  • Author(s): T Wakita
  • Organizer: Singapore-Japan Forum on Emerging Concepts in Microbiology
  • Place of Presentation: National University of Singapore, Singapore
  • Year and Date: 20111115-16
• [Presentation] HCV JFH-1キメラ株を用いたNS5a阻害剤の株特異的抗ウイルス活性の評価 (2011)
  • Author(s): 加藤孝宣、政木隆博、脇田隆字
  • Organizer: 第15回日本肝臓学会大会
  • Place of Presentation: 福岡国際会議場、シンポジウム1「C 型肝炎治療の新たな展開」
  • Year and Date: 20111020-21
• [Presentation] Strain Specific Susceptibility to The Hepatitis C Virus NS5A Inhibitor (2011)
  • Author(s): Y Okamoto, T Masaki, A Murayama, A Nomoto, T Wakita, T Kato
  • Organizer: XV International Congress of Virology
  • Place of Presentation: Sapporo Convention Center, Sapporo, Japan
  • Year and Date: 20110911-16
• [Presentation] Identification of lipid droplet-associated membrane proteins that are involved in HCV production (2011)
  • Author(s): H Aizaki, Y Matsumoto, K Goto, K Watashi, R Suzuki, M Fukasawa, K Hanada, S Sato, N Takahashi, Y Matsuura, K Motojima, T Miyamura, T Suzuki , T Wakita
  • Organizer: XV International Congress of Virology
  • Place of Presentation: Sapporo Convention Center, Sapporo, Japan
  • Year and Date: 20110911-16
• [Presentation] Antiviral activity of glycyrrhizic acid against hepatitis C virus in vitro (2011)
  • Author(s): Y Matsumoto, K Watashi, R Suzuki, T Matsuura, T Suzuki, T Miyamura, K Wake, T Wakita , H Aizaki
  • Organizer: XV International Congress of Virology
  • Place of Presentation: Sapporo Convention Center, Sapporo, Japan
  • Year and Date: 20110911-16
• [Presentation] Identification of small molecules affecting late steps of hepatitis C virus life cycle (2011)
  • Author(s): K Watashi, N Uchida, R Suzuki, H Aizaki, T Wakita
  • Organizer: XV International Congress of Virology
  • Place of Presentation: Sapporo Convention Center, Sapporo, Japan
  • Year and Date: 20110911-16
• [Presentation] Identification of novel NS5A-associated proteins in the host-cell membrane fraction and their role in HCV life cycle (2011)
  • Author(s): K Goto, T Kimura, K Watashi, R Suzuki, S Yamagoe, T Miyamura, K Moriya, H Yotsuyanagi, K Koike, T Suzuki, T Wakita, H Aizaki
  • Organizer: 18th International Meeting on Hepatitis C and Related Viruses
  • Place of Presentation: Seattle Sheraton Hotel, Seattle, USA
  • Year and Date: 20110908-12
• [Presentation] Does a vaccine derived from a single HCV strain elicit broadly cross-neutralising antibodies in humans? (2011)
  • Author(s): J Law, D Hockman, S Frey,R Khoshy, T Wakita, J Bukh, C Rice, M Houghton
  • Organizer: 18th International Meeting on Hepatitis C and Related Viruses
  • Place of Presentation: Seattle Sheraton Hotel, Seattle, USA
  • Year and Date: 20110908-12
• [Presentation] Isolation of a highly infectious hepatitis C virus with adaptive mutations (2011)
  • Author(s): M Fukasawa, Y Shirasago, K Saito, Y Murakami, H Fukazawa, T Suzuki, R Suzuki, T Wakita, K Hanada, J Chiba
  • Organizer: 18th International Meeting on Hepatitis C and Related Viruses
  • Place of Presentation: Seattle Sheraton Hotel, Seattle, USA
  • Year and Date: 20110908-12
• [Presentation] Development of purification method for HCV particles using chromatographic technique (2011)
  • Author(s): H Yokokawa, D Akazawa, M Moriyama, N Nakamura, T Kato, K Ishii, T Wakita
  • Organizer: 18th International Meeting on Hepatitis C and Related Viruses
  • Place of Presentation: Seattle Sheraton Hotel, Seattle, USA
  • Year and Date: 20110908-12
• [Presentation] Serum and trypsin inhibitors inhibit the early step of hepatitis C virus infection (2011)
  • Author(s): M Esumi, S Kikuta, H Yamaguchi, S Nakajima, M Ishibashi, T Wakita
  • Organizer: 18th International Meeting on Hepatitis C and Related Viruses
  • Place of Presentation: Seattle Sheraton Hotel, Seattle, USA
  • Year and Date: 20110908-12
• [Presentation] Immunological memory response to induce neutralizing immunoglobulin in HCV particles-immunized mice (2011)
  • Author(s): M Moriyama, D Akazawa, H Yokokawa, K Nishimura, N Nakamura, H Mochizuki, T Kato, K Ishii, T Wakita
  • Organizer: 18th International Meeting on Hepatitis C and Related Viruses
  • Place of Presentation: Seattle Sheraton Hotel, Seattle, USA
  • Year and Date: 20110908-12
• [Presentation] Identification of small molecules affecting late steps of hepatitis C virus life cycle (2011)
  • Author(s): K Watashi, N Uchida, R Suzuki, H Aizaki, T Wakita
  • Organizer: 18th International Meeting on Hepatitis C and Related Viruses
  • Place of Presentation: Seattle Sheraton Hotel, Seattle, USA
  • Year and Date: 20110908-12
• [Presentation] Halopemide inhibited a post-assembly step in hepatitis C virus life cycle (2011)
  • Author(s): N Uchida, K Watashi, R Suzuki, H Aizaki, J Chiba, T Wakita
  • Organizer: 18th International Meeting on Hepatitis C and Related Viruses
  • Place of Presentation: Seattle Sheraton Hotel, Seattle, USA
  • Year and Date: 20110908-12
• [Presentation] 2 種類のヒト肝細胞株を用いたHCV 感染レポーターアッセイ系の開発 (2011)
  • Author(s): 武田緑、池田正徳、有海康雄、脇田隆字、加藤宜之
  • Organizer: 第47回日本肝臓学会総会
  • Place of Presentation: ホテルグランパシフィック
  • Year and Date: 20110602-03
• [Presentation] 「新型シーケンサで展開するウイルスゲノム研究」、ランチョンセミナー (2011)
  • Author(s): 脇田隆字
  • Organizer: 第34回日本分子生物学会年会
  • Place of Presentation: パシフィコ横浜
  • Year and Date: 2011-12-14
• [Presentation] HCV RNA replication and drug development (2011)
  • Author(s): T Wakita
  • Organizer: The 8th APASL Single Topic Conference Beijing
  • Place of Presentation: China
  • Year and Date: 2011-10-07
• [Presentation] Hepatitis C Virus Infection and Replication, annual meeting of Prof (2011)
  • Author(s): T Wakita
  • Organizer: Juei-Low Sung's Research Foundation
  • Place of Presentation: Taipei, Taiwan
  • Year and Date: 2011-08-06