Researcher Database

Ayato Takada
Research Center for Zoonosis Control Division of Global Epidemiology
Professor

Researcher Profile and Settings

Affiliation

  • Research Center for Zoonosis Control Division of Global Epidemiology

Job Title

  • Professor

Degree

  • (BLANK)(Hokkaido University)

J-Global ID

Research Interests

  • diagnostics   anti-virals   自然宿主   宿主域   ワクチン   疫学   レセプター   侵入機構   病原性   ウイルス   Pathogenesis   Vaccine   Immunity   Virus   

Research Areas

  • Life sciences / Virology
  • Life sciences / Veterinary medicine
  • Life sciences / Veterinary medicine
  • Life sciences / Veterinary medicine

Academic & Professional Experience

  • 2009/06 - Today Rocky Mountain Laboratories, Hamilton, Montana, USA. Special Volunteer
  • 2007/10 - Today School of Veterinary Medicine, University of Zambia Visiting Professor
  • 2006 - Today Kobe University
  • 2005/05 - Today Hokkaido University Research Center for Zoonosis Control Professor
  • 2000/12 - 2009 Canadian Science Centre for Human and Animal Health Visiting Scientist
  • 2000 - 2004 The University of Tokyo The Institute of Medical Science
  • 1997 - 2000 Hokkaido University Graduate School of Veterinary Medicine
  • 1997 - 2000 Hokkaido University,
  • Assistant Professor
  • Graduate School of Veterinary Medicine,

Education

  •        - 1996  Hokkaido University
  •        - 1996  Hokkaido University  Graduate School, Division of Veterinary Medicine
  •        - 1993  Hokkaido University  School of Veterinary Medicine
  •        - 1993  Hokkaido University  Faculty of Veterinary Medicine

Association Memberships

  • 日本ワクチン学会   日本ウィルス学会   日本獣医学会   

Research Activities

Published Papers

  • Kajihara M, Hang'ombe BM, Changula K, Harima H, Isono M, Okuya K, Yoshida R, Mori-Kajihara A, Eto Y, Orba Y, Ogawa H, Qiu Y, Sawa H, Simulundu E, Mwizabi D, Munyeme M, Squarre D, Mukonka V, Mweene A, Takada A
    Emerging infectious diseases 25 (8) 1080-6040 2019/08 [Refereed][Not invited]
  • Amarasinghe GK, Ayllón MA, Bào Y, Basler CF, Bavari S, Blasdell KR, Briese T, Brown PA, Bukreyev A, Balkema-Buschmann A, Buchholz UJ, Chabi-Jesus C, Chandran K, Chiapponi C, Crozier I, de, Swart RL, Dietzgen RG, Dolnik O, Drexler JF, Dürrwald R, Dundon WG, Duprex WP, Dye JM, Easton AJ, Fooks AR, Formenty PBH, Fouchier RAM, Freitas-Astúa J, Griffiths A, Hewson R, Horie M, Hyndman TH, Jiāng D, Kitajima EW, Kobinger GP, Kondō H, Kurath G, Kuzmin IV, Lamb RA, Lavazza A, Lee B, Lelli D, Leroy EM, Lǐ J, Maes P, Marzano SL, Moreno A, Mühlberger E, Netesov SV, Nowotny N, Nylund A, Økland AL, Palacios G, Pályi B, Pawęska JT, Payne SL, Prosperi A, Ramos-González PL, Rima BK, Rota P, Rubbenstroth D, Shī M, Simmonds P, Smither SJ, Sozzi E, Spann K, Stenglein MD, Stone DM, Takada A, Tesh RB, Tomonaga K, Tordo N, Towner JS, van den Hoogen B, Vasilakis N, Wahl V, Walker PJ, Wang LF, Whitfield AE, Williams JV, Zerbini FM, Zhāng T, Zhang YZ, Kuhn JH
    Archives of virology 164 (7) 1967 - 1980 0304-8608 2019/07 [Refereed][Not invited]
  • Yongjin Qiu, Ryo Nakao, Bernard Mudenda Hang'ombe, Kozue Sato, Masahiro Kajihara, Sharon Kanchela, Katendi Changula, Yoshiki Eto, Joseph Ndebe, Michihito Sasaki, May June Thu, Ayato Takada, Hirohumi Sawa, Chihiro Sugimoto, Hiroki Kawabata
    Clinical infectious diseases : an official publication of the Infectious Diseases Society of America 69 (1) 107 - 112 1058-4838 2019/06/18 [Refereed][Not invited]
     
    BACKGROUND: Relapsing fever is an infectious disease previously neglected in Africa, which imposes a large public health burden in the country. We aimed to investigate and report on a case of relapsing fever borreliosis in Zambia. METHODS: A previously unknown Borrelia species was isolated from the blood of a febrile patient. Investigations of the presumptive vector ticks and natural hosts for the Borrelia species were conducted by culture isolation and/or DNA detection by Borrelia-specific polymerase chain reaction. Using culture isolates from the patient and bat specimens, genetic characterization was performed by multilocus sequence analysis based on the draft genome sequences. RESULTS: The febrile patient was diagnosed with relapsing fever. The isolated Borrelia species was frequently detected in Ornithodoros faini (n = 20/50 [40%]) and bats (n = 64/237 [27%]). Multilocus sequence analysis based on a draft genome sequence revealed that the Borrelia species isolates from the patient and presumptive reservoir host (bats) formed a monophyletic lineage that clustered with relapsing fever borreliae found in the United States. CONCLUSIONS: A febrile illness caused by a Borrelia species that was treatable with erythromycin was identified in Zambia. This is the first study to report on relapsing fever Borrelia in Zambia and suggesting the likely natural reservoir hosts of the isolated Borrelia species. Interestingly, the isolated Borrelia species was more closely related to New World relapsing fever borreliae, despite being detected in the Afrotropic ecozone.
  • Kuhn JH, Amarasinghe GK, Basler CF, Bavari S, Bukreyev A, Chandran K, Crozier I, Dolnik O, Dye JM, Formenty PBH, Griffiths A, Hewson R, Kobinger GP, Leroy EM, Mühlberger E, Netesov Нетёсов Сергей Викторович SV, Palacios G, Pályi B, Pawęska JT, Smither SJ, Takada 高田礼人, Towner JS, Wahl V, Ictv Report, Consortium
    The Journal of general virology 100 (6) 911 - 912 0022-1317 2019/06 [Refereed][Not invited]
  • Maes P, Amarasinghe GK, Ayllón MA, Basler CF, Bavari S, Blasdell KR, Briese T, Brown PA, Bukreyev A, Balkema-Buschmann A, Buchholz UJ, Chandran K, Crozier I, de, Swart RL, Dietzgen RG, Dolnik O, Domier LL, Drexler JF, Dürrwald R, Dundon WG, Duprex WP, Dye JM, Easton AJ, Fooks AR, Formenty PBH, Fouchier RAM, Freitas-Astúa J, Ghedin E, Griffiths A, Hewson R, Horie M, Hurwitz JL, Hyndman TH, Jiāng D, Kobinger GP, Kondō H, Kurath G, Kuzmin IV, Lamb RA, Lee B, Leroy EM, Lǐ J, Marzano SL, Mühlberger E, Netesov SV, Nowotny N, Palacios G, Pályi B, Pawęska JT, Payne SL, Rima BK, Rota P, Rubbenstroth D, Simmonds P, Smither SJ, Song Q, Song T, Spann K, Stenglein MD, Stone DM, Takada A, Tesh RB, Tomonaga K, Tordo N, Towner JS, van den Hoogen B, Vasilakis N, Wahl V, Walker PJ, Wang D, Wang LF, Whitfield AE, Williams JV, Yè G, Zerbini FM, Zhang YZ, Kuhn JH
    Archives of virology 164 (4) 1233 - 1244 0304-8608 2019/04 [Refereed][Not invited]
  • Rijal P, Elias SC, Machado SR, Xiao J, Schimanski L, O'Dowd V, Baker T, Barry E, Mendelsohn SC, Cherry CJ, Jin J, Labbé GM, Donnellan FR, Rampling T, Dowall S, Rayner E, Findlay-Wilson S, Carroll M, Guo J, Xu XN, Huang KA, Takada A, Burgess G, McMillan D, Popplewell A, Lightwood DJ, Draper SJ, Townsend AR
    Cell reports 27 (1) 172 - 186.e7 2019/04 [Refereed][Not invited]
  • May June Thu, Yongjin Qiu, Keita Matsuno, Masahiro Kajihara, Akina Mori-Kajihara, Ryosuke Omori, Naota Monma, Kazuki Chiba, Junji Seto, Mutsuyo Gokuden, Masako Andoh, Hideo Oosako, Ken Katakura, Ayato Takada, Chihiro Sugimoto, Norikazu Isoda, Ryo Nakao
    Scientific reports 9 (1) 1500 - 1500 2019/02/06 [Refereed][Not invited]
     
    Spotted fever group (SFG) rickettsiae are obligate intracellular Gram-negative bacteria mainly associated with ticks. In Japan, several hundred cases of Japanese spotted fever, caused by Rickettsia japonica, are reported annually. Other Rickettsia species are also known to exist in ixodid ticks; however, their phylogenetic position and pathogenic potential are poorly understood. We conducted a nationwide cross-sectional survey on questing ticks to understand the overall diversity of SFG rickettsiae in Japan. Out of 2,189 individuals (19 tick species in 4 genera), 373 (17.0%) samples were positive for Rickettsia spp. as ascertained by real-time PCR amplification of the citrate synthase gene (gltA). Conventional PCR and sequencing analyses of gltA indicated the presence of 15 different genotypes of SFG rickettsiae. Based on the analysis of five additional genes, we characterised five Rickettsia species; R. asiatica, R. helvetica, R. monacensis (formerly reported as Rickettsia sp. In56 in Japan), R. tamurae, and Candidatus R. tarasevichiae and several unclassified SFG rickettsiae. We also found a strong association between rickettsial genotypes and their host tick species, while there was little association between rickettsial genotypes and their geographical origins. These observations suggested that most of the SFG rickettsiae have a limited host range and are maintained in certain tick species in the natural environment.
  • Masahiro Sato, Junki Maruyama, Tatsunari Kondoh, Naganori Nao, Hiroko Miyamoto, Yoshihiro Takadate, Wakako Furuyama, Masahiro Kajihara, Hirohito Ogawa, Rashid Manzoor, Reiko Yoshida, Manabu Igarashi, Ayato Takada
    Scientific reports 9 (1) 1158 - 1158 2019/02/04 [Refereed][Not invited]
     
    Two novel influenza A virus-like genomes were detected in fruit bats in Central and South America. However, the biological properties of these bat-derived influenza viruses (BatIVs) are still largely unknown since infectious viral particles have never been isolated from the infected host species. In this study, a reverse genetics approach was used to generate infectious BatIV particles entirely from plasmids encoding full-length sequences in eight gene segments. We inoculated BatIV particles into various cell cultures including bat-derived cell lines and found that BatIVs infected particular bat-derived cells efficiently but not the other cell lines tested. Reassortant viruses between the two BatIVs were also successfully generated and their replication in the susceptible bat cell lines was confirmed. These findings suggest a limited host range and reassortment potential of BatIVs in nature, providing fundamental information for understanding of the ecology of BatIVs.
  • Shiho Torii, Keita Matsuno, Yongjin Qiu, Akina Mori-Kajihara, Masahiro Kajihara, Ryo Nakao, Naganori Nao, Katsunori Okazaki, Mariko Sashika, Takahiro Hiono, Masatoshi Okamatsu, Yoshihiro Sakoda, Hideki Ebihara, Ayato Takada, Hirofumi Sawa
    Ticks and tick-borne diseases 10 (2) 328 - 335 1877-959X 2019/02 [Refereed][Not invited]
     
    Recent discoveries of tick-borne pathogens have raised public health concerns on tick-borne infectious diseases and emphasize the need to assess potential risks of unrecognized tick-borne pathogens. First, to determine the existence of tick-borne phleboviruses (TBPVs), genetic surveillance of phleboviruses in ticks was conducted mainly in Hokkaido, the northernmost island in Japan from 2013 to 2015. Genes of two TBPVs, previously reported as Mukawa virus (MKWV) and a newly identified relative of MKWV, Kuriyama virus (KURV), were detected and the viruses were isolated from Ixodes persulcatus collected in Hokkaido, but not in I. persulcatus collected from other areas of Japan. These viruses were phylogenetically and antigenically similar to each other. Next, to investigate the infection of MKWV in mammals, serum samples from wildlife captured in Hokkaido from 2007 to 2011 were used for serological screening. Neutralizing antibodies against MKWV were detected in both Yezo-deer (Cervus nippon yesoensis) (2/50) and raccoons (Procyon lotor) (16/64). However, no infectious MKWV was recovered from laboratory mice in experimental infections, though viral RNAs were detected in their tissues. Thus, MKWV and KURV may maintain tick-mammalian life cycles in Hokkaido, suggesting their potential as causative agents of tick-borne diseases in mammals.
  • Kapiya J, Nalubamba KS, Kaimoyo E, Changula K, Chidumayo N, Saasa N, Simuunza MC, Takada A, Mweene AS, Chitanga S, Simulundu E
    Archives of virology 164 (1) 303 - 307 0304-8608 2019/01 [Refereed][Not invited]
  • Kobayashi T, Matsugo H, Maruyama J, Kamiki H, Takada A, Maeda K, Takenaka-Uema A, Tohya Y, Murakami S, Horimoto T
    Scientific reports 9 (1) 573  2019/01 [Refereed][Not invited]
  • Moriyama M, Igarashi M, Koshiba T, Irie T, Takada A, Ichinohe T
    Journal of virology 92 (19) 0022-538X 2018/10 [Refereed][Not invited]
  • Sasaki M, Kajihara M, Changula K, Mori-Kajihara A, Ogawa H, Hang'ombe BM, Mweene AS, Simuunza M, Yoshida R, Carr M, Orba Y, Takada A, Sawa H
    Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases 63 104 - 109 1567-1348 2018/09 [Refereed][Not invited]
  • Keita Matsuno, Noriyuki Nonoue, Ayako Noda, Nodoka Kasajima, Keita Noguchi, Ai Takano, Hiroshi Shimoda, Yasuko Orba, Mieko Muramatsu, Yoshihiro Sakoda, Ayato Takada, Shinji Minami, Yumi Une, Shigeru Morikawa, Ken Maeda
    Emerging infectious diseases 24 (9) 1726 - 1729 1080-6040 2018/09 [Refereed][Not invited]
     
    Two captive cheetahs from a zoo in Japan died of a severe fever with thrombocytopenia syndrome-like illness. Severe fever with thrombocytopenia syndrome virus, an endemic tickborne phlebovirus, was detected systemically with secretion of infectious viruses into the saliva. These cases highlight the risk for exposure of captive animals to endemic arthropodborne pathogens.
  • Milligan JC, Parekh DV, Fuller KM, Igarashi M, Takada A, Saphire EO
    The Journal of infectious diseases 219 (3) 415 - 419 0022-1899 2018/09 [Refereed][Not invited]
  • Amarasinghe GK, Aréchiga Ceballos NG, Banyard AC, Basler CF, Bavari S, Bennett AJ, Blasdell KR, Briese T, Bukreyev A, Caì Y, Calisher CH, Campos Lawson C, Chandran K, Chapman CA, Chiu CY, Choi KS, Collins PL, Dietzgen RG, Dolja VV, Dolnik O, Domier LL, Dürrwald R, Dye JM, Easton AJ, Ebihara H, Echevarría JE, Fooks AR, Formenty PBH, Fouchier RAM, Freuling CM, Ghedin E, Goldberg TL, Hewson R, Horie M, Hyndman TH, Jiāng D, Kityo R, Kobinger GP, Kondō H, Koonin EV, Krupovic M, Kurath G, Lamb RA, Lee B, Leroy EM, Maes P, Maisner A, Marston DA, Mor SK, Müller T, Mühlberger E, Ramírez VMN, Netesov SV, Ng TFF, Nowotny N, Palacios G, Patterson JL, Pawęska JT, Payne SL, Prieto K, Rima BK, Rota P, Rubbenstroth D, Schwemmle M, Siddell S, Smither SJ, Song Q, Song T, Stenglein MD, Stone DM, Takada A, Tesh RB, Thomazelli LM, Tomonaga K, Tordo N, Towner JS, Vasilakis N, Vázquez-Morón S, Verdugo C, Volchkov VE, Wahl V, Walker PJ, Wang D, Wang LF, Wellehan JFX, Wiley MR, Whitfield AE, Wolf YI, Yè G, Zhāng YZ, Kuhn JH
    Archives of virology 163 (8) 2283 - 2294 0304-8608 2018/08 [Refereed][Not invited]
  • Saphire EO, Schendel SL, Fusco ML, Gangavarapu K, Gunn BM, Wec AZ, Halfmann PJ, Brannan JM, Herbert AS, Qiu X, Wagh K, He S, Giorgi EE, Theiler J, Pommert KBJ, Krause TB, Turner HL, Murin CD, Pallesen J, Davidson E, Ahmed R, Aman MJ, Bukreyev A, Burton DR, Crowe JE Jr, Davis CW, Georgiou G, Krammer F, Kyratsous CA, Lai JR, Nykiforuk C, Pauly MH, Rijal P, Takada A, Townsend AR, Volchkov V, Walker LM, Wang CI, Zeitlin L, Doranz BJ, Ward AB, Korber B, Kobinger GP, Andersen KG, Kawaoka Y, Alter G, Chandran K, Dye JM, Viral Hemorrhagic Fever, Immunotherapeutic Consortium
    Cell 174 (4) 938 - 952.e13 0092-8674 2018/08 [Refereed][Not invited]
  • Kondoh T, Letko M, Munster VJ, Manzoor R, Maruyama J, Furuyama W, Miyamoto H, Shigeno A, Fujikura D, Takadate Y, Yoshida R, Igarashi M, Feldmann H, Marzi A, Takada A
    The Journal of infectious diseases 218 (suppl_5) S397 - S402 0022-1899 2018/07 [Refereed][Not invited]
  • Keita Matsuno, Masahiro Kajihara, Ryo Nakao, Naganori Nao, Akina Mori-Kajihara, Mieko Muramatsu, Yongjin Qiu, Shiho Torii, Manabu Igarashi, Nodoka Kasajima, Keita Mizuma, Kentaro Yoshii, Hirofumi Sawa, Chihiro Sugimoto, Ayato Takada, Hideki Ebihara
    mSphere 3 (3) 2018/06/27 [Refereed][Not invited]
     
    The recent emergence of novel tick-borne RNA viruses has complicated the epidemiological landscape of tick-borne infectious diseases, posing a significant challenge to public health systems that seek to counteract tick-borne diseases. The identification of two novel tick-borne phleboviruses (TBPVs), severe fever with thrombocytopenia syndrome virus (SFTSV) and Heartland virus (HRTV), as causative agents of severe illness in humans has accelerated the investigation and discoveries of novel TBPVs. In the present study, we isolated a novel TBPV designated Mukawa virus (MKWV) from host-questing Ixodes persulcatus females captured in Japan. Genetic characterization revealed that MKWV is a member of the genus Phlebovirus in the family Phenuiviridae Interestingly, MKWV is genetically distinct from other known TBPVs and shares a most recent common ancestor with mosquito/sandfly-borne (insect-borne) phleboviruses. Despite its genetic similarity to insect-borne phleboviruses, the molecular footprints of its viral proteins and its biological characteristics define MKWV as a tick-borne virus that can be transmitted to mammals. A phylogenetic ancestral-state reconstruction for arthropod vectors of phleboviruses including MKWV based on viral L segment sequences indicated that ticks likely harbored ancestral phleboviruses that evolved into both the tick-borne and MKWV/insect-borne phlebovirus lineages. Overall, our findings suggest that most of the phlebovirus evolution has occurred in hard ticks to generate divergent viruses, which may provide a seminal foundation for understanding the mechanisms underlying the evolution and emergence of pathogenic phleboviruses, such as Rift Valley fever virus and SFTSV/HRTV.IMPORTANCE The emergence of novel tick-borne RNA viruses causing severe illness in humans has complicated the epidemiological landscape of tick-borne diseases, requiring further investigation to safeguard public health. In the present study, we discovered a novel tick-borne phlebovirus from Ixodes persulcatus ticks in Japan. While its viral RNA genome sequences were similar to those of mosquito/sandfly-borne viruses, molecular and biological footprints confirmed that this is a tick-borne virus. The unique evolutionary position of the virus allowed us to estimate the ancestral phlebovirus vector, which was likely a hard tick. Our findings may provide a better understanding of the evolution and emergence of phleboviruses associated with emerging infectious diseases, such as severe fever with thrombocytopenia syndrome (SFTS) and Heartland virus disease.
  • Mwenda R, Changula K, Hang'ombe BM, Chidumayo N, Mangani AS, Kaira T, Takada A, Mweene AS, Simulundu E
    Avian pathology : journal of the W.V.P.A 47 (3) 300 - 313 0307-9457 2018/06 [Refereed][Not invited]
  • Marzi A, Haddock E, Kajihara M, Feldmann H, Takada A
    The Journal of infectious diseases 218 (suppl_5) S662 - S665 0022-1899 2018/06 [Refereed][Not invited]
  • Changula K, Kajihara M, Mori-Kajihara A, Eto Y, Miyamoto H, Yoshida R, Shigeno A, Hang'ombe B, Qiu Y, Mwizabi D, Squarre D, Ndebe J, Ogawa H, Harima H, Simulundu E, Moonga L, Kapila P, Furuyama W, Kondoh T, Sato M, Takadate Y, Kaneko C, Nakao R, Mukonka V, Mweene A, Takada A
    The Journal of infectious diseases 218 (suppl_5) S312 - S317 0022-1899 2018/06 [Refereed][Not invited]
  • Simulundu E, Sinkala Y, Chambaro HM, Chinyemba A, Banda F, Mooya LE, Ndebe J, Chitanga S, Makungu C, Munthali G, Fandamu P, Takada A, Mweene AS
    The Onderstepoort journal of veterinary research 85 (1) e1 - e5 0030-2465 2018/06 [Refereed][Not invited]
  • Sasaki M, Anindita PD, Ito N, Sugiyama M, Carr M, Fukuhara H, Ose T, Maenaka K, Takada A, Hall WW, Orba Y, Sawa H
    The Journal of infectious diseases 217 (11) 1740 - 1749 0022-1899 2018/05 [Refereed][Not invited]
  • Qiu Y, Kaneko C, Kajihara M, Ngonda S, Simulundu E, Muleya W, Thu MJ, Hang'ombe MB, Katakura K, Takada A, Sawa H, Simuunza M, Nakao R
    Ticks and tick-borne diseases 9 (4) 988 - 995 1877-959X 2018/05 [Refereed][Not invited]
  • Saasa N, Kajihara M, Dautu G, Mori-Kajihara A, Fukushi S, Sinkala Y, Morikawa S, Mweene A, Takada A, Yoshimatsu K, Arikawa J
    Vector borne and zoonotic diseases (Larchmont, N.Y.) 18 (5) 273 - 277 1530-3667 2018/05 [Refereed][Not invited]
  • Fujihira H, Usami K, Matsuno K, Takeuchi H, Denda-Nagai K, Furukawa JI, Shinohara Y, Takada A, Kawaoka Y, Irimura T
    Scientific reports 8 (1) 5495  2018/04 [Refereed][Not invited]
  • Asuka Nanbo, Junki Maruyama, Masaki Imai, Michiko Ujie, Yoichiro Fujioka, Shinya Nishide, Ayato Takada, Yusuke Ohba, Yoshihiro Kawaoka
    PLoS pathogens 14 (1) e1006848  1553-7366 2018/01 [Refereed][Not invited]
     
    Cell surface receptors for phosphatidylserine contribute to the entry of Ebola virus (EBOV) particles, indicating that the presence of phosphatidylserine in the envelope of EBOV is important for the internalization of EBOV particles. Phosphatidylserine is typically distributed in the inner layer of the plasma membrane in normal cells. Progeny virions bud from the plasma membrane of infected cells, suggesting that phosphatidylserine is likely flipped to the outer leaflet of the plasma membrane in infected cells for EBOV virions to acquire it. Currently, the intracellular dynamics of phosphatidylserine during EBOV infection are poorly understood. Here, we explored the role of XK-related protein (Xkr) 8, which is a scramblase responsible for exposure of phosphatidylserine in the plasma membrane of apoptotic cells, to understand its significance in phosphatidylserine-dependent entry of EBOV. We found that Xkr8 and transiently expressed EBOV glycoprotein GP often co-localized in intracellular vesicles and the plasma membrane. We also found that co-expression of GP and viral major matrix protein VP40 promoted incorporation of Xkr8 into ebolavirus-like particles (VLPs) and exposure of phosphatidylserine on their surface, although only a limited amount of phosphatidylserine was exposed on the surface of the cells expressing GP and/or VP40. Downregulating Xkr8 or blocking caspase-mediated Xkr8 activation did not affect VLP production, but they reduced the amount of phosphatidylserine on the VLPs and their uptake in recipient cells. Taken together, our findings indicate that Xkr8 is trafficked to budding sites via GP-containing vesicles, is incorporated into VLPs, and then promote the entry of the released EBOV to cells in a phosphatidylserine-dependent manner.
  • Chitanga S, Simulundu E, Simuunza MC, Changula K, Qiu Y, Kajihara M, Nakao R, Syakalima M, Takada A, Mweene AS, Mukaratirwa S, Hang'ombe BM
    Parasites & vectors 11 (1) 40  2018/01 [Refereed][Not invited]
  • Pipina A. Vlahakis, Simbarashe Chitanga, Martin C. Simuunza, Edgar Simulundu, Yongjin Qiu, Katendi Changula, Herman M. Chambaro, Masahiro Kajihara, Ryo Nakao, Ayato Takada, Aaron S. Mweene
    TICKS AND TICK-BORNE DISEASES 9 (1) 39 - 43 1877-959X 2018/01 [Refereed][Not invited]
     
    Although tick-borne pathogens, Anaplasma platys and Anaplasma phagocytophilum are recognized as zoonotic agents associated with appreciable morbidity and mortality in dogs and humans worldwide, there is limited information on these infections in many African countries, including Zambia. The purpose of this study was to detect, identify and phylogenetically characterize Anaplasma species from dogs in Chilanga District in Lusaka Province, Zambia. A total of 301 blood samples were collected from apparently healthy and semi-confined dogs. Initial screening by polymerase chain reaction with specific primers targeting the 16S rRNA gene of Anaplasma species revealed that 9% (27/301) of our samples were positive. Subsequent sequence and phylogenetic analysis of a longer fragment of the 16S rRNA and citrate synthase (gltA) genes of four positive samples showed the presence of A. platys and an Anaplasma species, which was closely related to those detected in dogs in South Africa. This is the first report on molecular identification and characterization of canine-associated zoonotic Anaplasma species in Zambia.
  • Hirohito Ogawa, Masahiro Kajihara, Naganori Nao, Asako Shigeno, Daisuke Fujikura, Bernard M Hang'ombe, Aaron S Mweene, Alisheke Mutemwa, David Squarre, Masao Yamada, Hideaki Higashi, Hirofumi Sawa, Ayato Takada
    Viruses 9 (12) 2017/12/04 [Refereed][Not invited]
     
    Bats are important reservoirs for emerging zoonotic viruses. For extensive surveys of potential pathogens in straw-colored fruit bats (Eidolon helvum) in Zambia, a total of 107 spleen samples of E. helvum in 2006 were inoculated onto Vero E6 cells. The cell culture inoculated with one of the samples (ZFB06-106) exhibited remarkable cytopathic changes. Based on the ultrastructural property in negative staining and cross-reactivity in immunofluorescence assays, the virus was suspected to be an adenovirus, and tentatively named E. helvum adenovirus 06-106 (EhAdV 06-106). Analysis of the full-length genome of 30,134 bp, determined by next-generation sequencing, showed the presence of 28 open reading frames. Phylogenetic analyses confirmed that EhAdV 06-106 represented a novel bat adenovirus species in the genus Mastadenovirus. The virus shared similar characteristics of low G + C contents with recently isolated members of species Bat mastadenoviruses E, F and G, from which EhAdV 06-106 diverged by more than 15% based on the distance matrix analysis of DNA polymerase amino acid sequences. According to the taxonomic criteria, we propose the tentative new species name "Bat mastadenovirus H". Because EhAdV 06-106 exhibited a wide in vitro cell tropism, the virus might have a potential risk as an emerging virus through cross-species transmission.
  • Manzoor R, Igarashi M, Takada A
    International journal of molecular sciences 18 (12) 2017/12 [Refereed][Not invited]
  • Tatsunari Kondoh, Rashid Manzoor, Naganori Nao, Junki Maruyama, Wakako Furuyama, Hiroko Miyamoto, Asako Shigeno, Makoto Kuroda, Keita Matsuno, Daisuke Fujikura, Masahiro Kajihara, Reiko Yoshida, Manabu Igarashi, Ayato Takada
    PLOS ONE 12 (10) e0186450  1932-6203 2017/10 [Refereed][Not invited]
     
    It has been proposed that some non-retroviral RNA virus genes are integrated into vertebrate genomes. Endogenous filovirus-like elements (EFLs) have been discovered in some mammalian genomes. However, their potential roles in ebolavirus infection are unclear. A filovirus VP35-like element (mlEFL35) is found in the little brown bat (Myotis lucifugus) genome. Putative mlEFL35-derived protein (mlEFL35p) contains nearly full-length amino acid sequences corresponding to ebolavirus VP35. Ebola virus VP35 has been shown to bind double-stranded RNA, leading to inhibition of type I interferon (IFN) production, and is also known as a viral polymerase cofactor that is essential for viral RNA transcription/replication. In this study, we transiently expressed mlEFL35p in human kidney cells and investigated its biological functions. We first found that mlEFL35p was coimmunoprecipitated with itself and ebolavirus VP35s but not with the viral nucleoprotein. Then the biological functions of mlEFL35p were analyzed by comparing it to ebolavirus VP35s. We found that the expression of mlEFL35p significantly inhibited human IFN-beta promoter activity as well as VP35s. By contrast, expression of mlEFL35p did not support viral RNA transcription/replication and indeed slightly decrease the reporter gene expression in a minigenome assay. These results suggest that mlEFL35p potentially acts as an IFN antagonist but not a polymerase cofactor.
  • Edgar Simulundu, Caesar H. Lubaba, Juanita van Heerden, Masahiro Kajihara, Liywalii Mataa, Herman Moses Chambaro, Yona Sinkala, Samuel Munalula Munjita, Hetron Mweemba Munang'andu, King Shimumbo Nalubamba, Kenny Samui, Girja Shanker Pandey, Ayato Takada, Aaron S. Mweene
    VIRUSES-BASEL 9 (9) 1999-4915 2017/09 [Refereed][Not invited]
     
    African swine fever (ASF) is a highly contagious and deadly viral hemorrhagic disease of swine. In Zambia, ASF was first reported in 1912 in Eastern Province and is currently believed to be endemic in that province only. Strict quarantine measures implemented at the Luangwa River Bridge, the only surface outlet from Eastern Province, appeared to be successful in restricting the disease. However, in 1989, an outbreak occurred for the first time outside the endemic province. Sporadic outbreaks have since occurred almost throughout the country. These events have brought into acute focus our limited understanding of the epidemiology of ASF in Zambia. Here, we review the epidemiology of the disease in areas considered nonendemic from 1989 to 2015. Comprehensive sequence analysis conducted on genetic data of ASF viruses (ASFVs) detected in domestic pigs revealed that p72 genotypes I, II, VIII and XIV have been involved in causing ASF outbreaks in swine during the study period. With the exception of the 1989 outbreak, we found no concrete evidence of dissemination of ASFVs from Eastern Province to other parts of the country. Our analyses revealed a complex epidemiology of the disease with a possibility of sylvatic cycle involvement. Trade and/ or movement of pigs and their products, both within and across international borders, appear to have been the major factor in ASFV dissemination. Since ASFVs with the potential to cause countrywide and possibly regional outbreaks, could emerge from "nonendemic regions", the current ASF control policy in Zambia requires a dramatic shift to ensure a more sustainable pig industry.
  • Olamide K. Oloniniyi, Yohei Kurosaki, Hiroko Miyamoto, Ayato Takada, Jiro Yasuda
    JOURNAL OF VIROLOGICAL METHODS 246 8 - 14 0166-0934 2017/08 [Refereed][Not invited]
     
    Ebola virus disease (EVD), a highly virulent infectious disease caused by ebolaviruses, has a fatality rate of 25-90%. Without a licensed chemotherapeutic agent or vaccine for the treatment and prevention of EVD, control of outbreaks requires accurate and rapid diagnosis of cases. In this study, five sets of six oligonucleotide primers targeting the nucleoprotein gene were designed for specific identification of each of the five ebolavirus species using reverse transcription-loop mediated isothermal amplification (RT-LAMP) assay. The detection limits of the ebolavirus species-specific primer sets were evaluated using in vitro transcribed RNAs. The detection limit of species-specific RT-LAMP assays for Zaire ebolavirus, Sudan ebolavirus, Tat Forest ebolavirus, and Bundibugyo ebolavirus was 256 copies/reaction, while the detection limit for Reston ebolavirus was 64 copies/reaction, and the detection time for each of the RT-LAMP assays was 13.3 +/- 3.0, 19.8 +/- 4.6, 14.3 +/- 0.6, 16.1 +/- 4.7, and 19.8 +/- 2.4 min (mean +/- SD), respectively. The sensitivity of the species-specific RT-LAMP assays were similar to that of the establishedRT-PCR and quantitative RT-PCR assays for diagnosis of EVD and are suitable for field or point-of-care diagnosis. The RT-LAMP assays were specific for the detection of the respective species of ebolavirus with no cross reaction with other species of ebolavirus and other viral hemorrhagic fever viruses such as Marburg virus, Lassa fever virus, and Dengue virus. The species-specific RT-LAMP assays developed in this study are rapid, sensitive, and specific and could be useful in case of an EVD outbreak.
  • Edgar Simulundu, Nandi Mtine, Thoko F. Kapalamula, Masahiro Kajihara, Yongjin Qiu, James Ngoma, Victor Zulu, Geoffrey Kwenda, Chrispin Chisanga, Isaac K. Phiri, Ayato Takada, Aaron S. Mweene
    ARCHIVES OF VIROLOGY 162 (8) 2363 - 2367 0304-8608 2017/08 [Refereed][Not invited]
     
    Orf or contagious ecthyma is a neglected and economically important zoonotic disease caused by a dermatotropic parapoxvirus that commonly affects domestic small ruminants. Although orf is globally distributed, there is a paucity of information on the disease in many African countries. Here, a suspected severe outbreak of orf in goats at a farm in Lusaka was investigated. Orf virus (ORFV) infection was confirmed by PCR amplification of viral DNA (RNA polymerase, B2L and virus interferon-resistance genes) in clinical samples. Some detected genes were sequenced and phylogenetically analyzed. This is the first report on molecular characterization of ORFV in goats in Zambia.
  • Gaya K. Amarasinghe, Yiming Bao, Christopher F. Basler, Sina Bavari, Martin Beer, Nicolas Bejerman, Kim R. Blasdell, Alisa Bochnowski, Thomas Briese, Alexander Bukreyev, Charles H. Calisher, Kartik Chandran, Peter L. Collins, Ralf G. Dietzgen, Olga Dolnik, Ralf Duerrwald, John M. Dye, Andrew J. Easton, Hideki Ebihara, Qi Fang, Pierre Formenty, Ron A. M. Fouchier, Elodie Ghedin, Robert M. Harding, Roger Hewson, Colleen M. Higgins, Jian Hong, Masayuki Horie, Anthony P. James, Daohong JiAng, Gary P. Kobinger, Hideki Kondo, Gael Kurath, Robert A. Lamb, Benhur Lee, Eric M. Leroy, Ming Li, Andrea Maisner, Elke Muhlberger, Sergey V. Netesov, Norbert Nowotny, Jean L. Patterson, Susan L. Payne, Janusz T. Paweska, Michael N. Pearson, Rick E. Randall, Peter A. Revill, Bertus K. Rima, Paul Rota, Dennis Rubbenstroth, Martin Schwemmle, Sophie J. Smither, Qisheng Song, David M. Stone, Ayato Takada, Calogero Terregino, Robert B. Tesh, Keizo Tomonaga, Noel Tordo, Jonathan S. Towner, Nikos Vasilakis, Viktor E. Volchkov, Victoria Wahl-Jensen, Peter J. Walker, Beibei Wang, David Wang, Fei Wang, Lin-Fa Wang, John H. Werren, Anna E. Whitfield, Zhichao Yan, Gongyin Ye, Jens H. Kuhn
    ARCHIVES OF VIROLOGY 162 (8) 2493 - 2504 0304-8608 2017/08 [Refereed][Not invited]
     
    In 2017, the order Mononegavirales was expanded by the inclusion of a total of 69 novel species. Five new rhabdovirus genera and one new nyamivirus genus were established to harbor 41 of these species, whereas the remaining new species were assigned to already established genera. Furthermore, non-Latinized binomial species names replaced all paramyxovirus and pneumovirus species names, thereby accomplishing application of binomial species names throughout the entire order. This article presents the updated taxonomy of the order Mononegavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV).
  • Lam Thanh Nguyen, Kazunari Nakaishi, Keiko Motojima, Ayako Ohkawara, Erina Minato, Junki Maruyama, Takahiro Hiono, Keita Matsuno, Masatoshi Okamatsu, Takashi Kimura, Ayato Takada, Hiroshi Kida, Yoshihiro Sakoda
    PLOS ONE 12 (8) e0182228  1932-6203 2017/08 [Refereed][Not invited]
     
    Highly pathogenic avian influenza viruses (HPAIVs) of H5 subtype have persistently caused outbreaks in domestic poultry and wild birds worldwide and sporadically infected humans. Rapid and accurate diagnosis is one of the key strategies for the control of H5 HPAIVs. However, the sensitivity of the diagnosis of H5 HPAIVs has gradually reduced due to extensive antigenic variation during their evolution. Particularly, the previously developed immunochromatographic diagnosis kit for H5 viruses, Linjudge Flu A/H5, exhibits reduced detection of H5 HPAIVs isolated in recent years. In the present study, we established a new advanced H5 rapid immunochromatographic detection kit (New Linjudge Flu A/H5) by a combination of two anti-H5 hemagglutinin monoclonal antibodies, A64/1 previously applied in the Linjudge Flu A/H5 and A32/2, a novel monoclonal antibody generated from a clade 2.3.4.4 H5 HPAIV. The new kit broadly detected all classical and recent H5 influenza viruses and showed a higher specificity and sensitivity than the original Linjudge Flu A/H5 with recently circulating H5 HPAIVs. Furthermore, the applicability of the New Linjudge Flu A/H5 was demonstrated by detecting antigens from the swabs and tissue homogenates of naturally infected birds and experimentally infected chickens with H5N6 HPAIVs belonging to the genetic clade 2.3.4.4. Our study, therefore, can provide an effective point-of-care rapid antigen detection kit for the surveillance of H5 avian influenza viruses and as a prompt countermeasure against the current widespread of the clade 2.3.4.4 H5 HPAIVs in domestic and wild birds.
  • Thomas S. Postler, Anna N. Clawson, Gaya K. Amarasinghe, Christopher F. Basler, Sbina Bavari, Maria Benko, Kim R. Blasdell, Thomas Briese, Michael J. Buchmeier, Alexander Bukreyev, Charles H. Calisher, Kartik Chandran, Remi Charrel, Christopher S. Clegg, Peter L. Collins, Juan Carlos de la Torre, Joseph L. Derisi, Ralf G. Dietzgen, Olga Dolnik, Ralf Duerrwald, John M. Dye, Andrew J. Easton, Sebastian Emonet, Pierre Formenty, Ron A. M. Fouchier, Elodie Ghedin, Jean-Paul Gonzalez, Balazs Harrach, Roger Hewson, Masayuki Horie, Daohong Jiang, Gary Kobinger, Hideki Kondo, Andrew M. Kropinski, Mart Krupovic, Gael Kurath, Robert A. Lamb, Eric M. Leroy, Igor S. Lukashevich, Andrea Maisner, Arcady R. Mushegian, Sergey V. Netesov, Norbert Nowotny, Jean L. Patterson, Susan L. Payne, Janusz T. Paweska, Clarence J. Peters, Sheli R. Radoshitzky, Bertus K. Rima, Victor Romanowski, Dennis Rubbenstroth, Sead Sabanadzovic, Helene Sanfacon, Maria S. Salvato, Martin Schwemmle, Sophie J. Smither, Mark D. Stenglein, David M. Stone, Ayato Takada, Robert B. Tesh, Keizo Tomonaga, Noel Tordo, Jonathan S. Towner, Nikos Vasilakis, Viktor E. Volchkov, Victoria Wahl-Jensen, Peter J. Walker, Lin-Fa Wang, Arvind Varsani, Anna E. Whitfield, F. Murilo Zerbini, Jens H. Kuhn
    SYSTEMATIC BIOLOGY 66 (3) 463 - 473 1063-5157 2017/05 [Refereed][Not invited]
     
    Botanical, mycological, zoological, and prokaryotic species names follow the Linnaean format, consisting of an italicized Latinized binomen with a capitalized genus name and a lower case species epithet (e.g., Homo sapiens). Virus species names, however, do not follow a uniform format, and, even when binomial, are not Linnaean in style. In this thought exercise, we attempted to convert all currently official names of species included in the virus family Arenaviridae and the virus order Mononegavirales to Linnaean binomials, and to identify and address associated challenges and concerns. Surprisingly, this endeavor was not as complicated or time-consuming as even the authors of this article expected when conceiving the experiment.
  • Yiming Bao, Gaya K. Amarasinghe, Christopher F. Basler, Sina Bavari, Alexander Bukreyev, Kartik Chandran, Olga Dolnik, John M. Dye, Hideki Ebihara, Pierre Formenty, Roger Hewson, Gary P. Kobinger, Eric M. Leroy, Elke Muhlberger, Sergey V. Netesov, Jean L. Patterson, Janusz T. Paweska, Sophie J. Smither, Ayato Takada, Jonathan S. Towner, Viktor E. Volchkov, Victoria Wahl-Jensen, Jens H. Kuhn
    VIRUSES-BASEL 9 (5) 1999-4915 2017/05 [Refereed][Not invited]
     
    The mononegaviral family Filoviridae has eight members assigned to three genera and seven species. Until now, genus and species demarcation were based on arbitrarily chosen filovirus genome sequence divergence values (approximate to 50% for genera, approximate to 30% for species) and arbitrarily chosen phenotypic virus or virion characteristics. Here we report filovirus genome sequence-based taxon demarcation criteria using the publicly accessible PAirwise Sequencing Comparison (PASC) tool of the US National Center for Biotechnology Information (Bethesda, MD, USA). Comparison of all available filovirus genomes in GenBank using PASC revealed optimal genus demarcation at the 55-58% sequence diversity threshold range for genera and at the 23-36% sequence diversity threshold range for species. Because these thresholds do not change the current official filovirus classification, these values are now implemented as filovirus taxon demarcation criteria that may solely be used for filovirus classification in case additional data are absent. A near-complete, coding-complete, or complete filovirus genome sequence will now be required to allow official classification of any novel "filovirus." Classification of filoviruses into existing taxa or determining the need for novel taxa is now straightforward and could even become automated using a presented algorithm/flowchart rooted in RefSeq (type) sequences.
  • Girja S. Pandey, Edgar Simulundu, Danstan Mwiinga, Kenny L. Samui, Aaron S. Mweene, Masahiro Kajihara, Alfred Mangani, Racheal Mwenda, Joseph Ndebe, Satoru Konnai, Ayato Takada
    ARCHIVES OF VIROLOGY 162 (4) 1051 - 1056 0304-8608 2017/04 [Refereed][Not invited]
     
    Bovine leukemia virus (BLV) causes enzootic bovine leucosis (EBL) and is responsible for substantial economic losses in cattle globally. However, information in Africa on the disease is limited. Here, based on clinical, hematological, pathological and molecular analyses, two clinical cases of EBL were confirmed in a dairy cattle herd in Zambia. In contrast, proviral DNA was detected by PCR in five apparently healthy cows from the same herd, suggesting subclinical BLV infection. Phylogenetic analysis of the env gene showed that the identified BLV clustered with Eurasian genotype 4 strains. This is the first report of confirmed EBL in Zambia.
  • Ryo Nakao, Keita Matsuno, Yongjin Qiu, Junki Marilyama, Nao Eguchi, Naganori Nao, Masahiro Kajihara, Kentaro Yoshii, Hirofumi Sawa, Ayato Takada, Chihiro Sugimoto
    TICKS AND TICK-BORNE DISEASES 8 (1) 103 - 111 1877-959X 2017 [Refereed][Not invited]
     
    Ticks harbour various microorganisms, some of which act as pathogens of humans and animals. The recent advancement of genome sequencing technologies revealed that a wide range of previously unrecognised microorganisms exist in ticks. Continuous cell lines established from ticks could play a key role in the isolation of such microorganisms; however, tick cells themselves have been known to harbour symbiotic microorganisms. The present study aimed to characterise putative RNA viral sequences detected in the culture supernatant of one of the most frequently used tick cell lines, ISE6, which was derived from embryos of the blacklegged tick Ixodes scapularis. Viral particles purified from the culture supernatant were used for RNA extraction, followed by Illumina sequencing. The reads were de novo assembled and the resulting contigs were annotated by tBLASTx search. The results suggested that there were at least five putative viral sequences of four phylogenetically distinct lineages in ISE6 cells. The predominant viral sequence found in ISE6 cells, designated L scapularis iflavirus, was a member of the family Iflaviridae, which is an arthropod-infecting virus group. We also identified L and M segments of the family Bunyaviridae, which could not be classified into any of the five known genera, and a potential capsid protein related to Drosophila A virus. In addition to these previously unrecognised viruses, ISE6 was revealed to harbour a putative genome sequence of L scapularis-associated virus-1, which was reported in a recent metagenomic study of L scapularis itself. All the five putative viral sequences were detected by RT-PCR in both ISE6 cells and the culture supernatant. Electron microscopic analysis showed the existence of spherical virions with a varying diameter of 50-70 nm in the culture supernatant of ISE6 cells. Further studies are required to investigate the potential roles of ISE6-associated viruses in ticks. (C) 2016 Elsevier GmbH. All rights reserved.
  • Naganori Nao, Junya Yamagishi, Hiroko Miyamoto, Manabu Igarashi, Rashid Manzoor, Aiko Ohnuma, Yoshimi Tsuda, Wakako Furuyama, Asako Shigeno, Masahiro Kajihara, Noriko Kishida, Reiko Yoshida, Ayato Takada
    MBIO 8 (1) 2150-7511 2017/01 [Refereed][Not invited]
     
    Highly pathogenic avian influenza viruses with H5 and H7 hemagglutinin (HA) subtypes evolve from low-pathogenic precursors through the acquisition of multiple basic amino acid residues at the HA cleavage site. Although this mechanism has been observed to occur naturally only in these HA subtypes, little is known about the genetic basis for the acquisition of the polybasic HA cleavage site. Here we show that consecutive adenine residues and a stem-loop structure, which are frequently found in the viral RNA region encoding amino acids around the cleavage site of low-pathogenic H5 and H7 viruses isolated from waterfowl reservoirs, are important for nucleotide insertions into this RNA region. A reporter assay to detect nontemplated nucleotide insertions and deep-sequencing analysis of viral RNAs revealed that an increased number of adenine residues and enlarged stem-loop structure in the RNA region accelerated the multiple adenine and/or guanine insertions required to create codons for basic amino acids. Interestingly, nucleotide insertions associated with the HA cleavage site motif were not observed principally in the viral RNA of other subtypes tested (H1, H2, H3, and H4). Our findings suggest that the RNA editing-like activity is the key mechanism for nucleotide insertions, providing a clue as to why the acquisition of the polybasic HA cleavage site is restricted to the particular HA subtypes. IMPORTANCE Influenza A viruses are divided into subtypes based on the antigenicity of the viral surface glycoproteins hemagglutinin (HA) and neuraminidase. Of the 16 HA subtypes (H1 to -16) maintained in waterfowl reservoirs of influenza A viruses, H5 and H7 viruses often become highly pathogenic through the acquisition of multiple basic amino acid residues at the HA cleavage site. Although this mechanism has been known since the 1980s, the genetic basis for nucleotide insertions has remained unclear. This study shows the potential role of the viral RNA secondary structure for nucleotide insertions and demonstrates a key mechanism explaining why the acquisition of the polybasic HA cleavage site is restricted to particular HA subtypes in nature. Our findings will contribute to better understanding of the ecology of influenza A viruses and will also be useful for the development of genetically modified vaccines against H5 and H7 influenza A viruses with increased stability.
  • Furuyama W, Miyamoto H, Yoshida R, Takada A
    Methods in molecular biology (Clifton, N.J.) 1628 309 - 320 1064-3745 2017 [Refereed][Not invited]
  • Constantin Brinkmann, Inga Nehlmeier, Kerstin Walendy-Gnirss, Julia Nehls, Mariana Gonzalez Hernandez, Markus Hoffmann, Xiangguo Qiu, Ayato Takada, Michael Schindler, Stefan Poehlmann
    JOURNAL OF VIROLOGY 90 (24) 11075 - 11086 0022-538X 2016/12 [Refereed][Not invited]
     
    The glycoprotein of Ebola virus (EBOV GP), a member of the family Filoviridae, facilitates viral entry into target cells. In addition, EBOV GP antagonizes the antiviral activity of the host cell protein tetherin, which may otherwise restrict EBOV release from infected cells. However, it is unclear how EBOV GP antagonizes tetherin, and it is unknown whether the GP of Lloviu virus (LLOV), a filovirus found in dead bats in Northern Spain, also counteracts tetherin. Here, we show that LLOV GP antagonizes tetherin, indicating that tetherin may not impede LLOV spread in human cells. Moreover, we demonstrate that appropriate processing of N-glycans in tetherin/GP-coexpressing cells is required for tetherin counteraction by EBOV GP. Furthermore, we show that an intact receptor-binding domain (RBD) in the GP1 subunit of EBOV GP is a prerequisite for tetherin counteraction. In contrast, blockade of Niemann-Pick disease type C1 (NPC1), a cellular binding partner of the RBD, did not interfere with tetherin antagonism. Finally, we provide evidence that an antibody directed against GP1, which protects mice from a lethal EBOV challenge, may block GP-dependent tetherin antagonism. Our data, in conjunction with previous reports, indicate that tetherin antagonism is conserved among the GPs of all known filoviruses and demonstrate that the GP1 subunit of EBOV GP plays a central role in tetherin antagonism. IMPORTANCE Filoviruses are reemerging pathogens that constitute a public health threat. Understanding how Ebola virus (EBOV), a highly pathogenic filovirus responsible for the 2013-2016 Ebola virus disease epidemic in western Africa, counteracts antiviral effectors of the innate immune system might help to define novel targets for antiviral intervention. Similarly, determining whether Lloviu virus (LLOV), a filovirus detected in bats in northern Spain, is inhibited by innate antiviral effectors in human cells might help to determine whether the virus constitutes a threat to humans. The present study shows that LLOV, like EBOV, counteracts the antiviral effector protein tetherin via its glycoprotein (GP), suggesting that tetherin does not pose a defense against LLOV spread in humans. Moreover, our work identifies the GP1 subunit of EBOV GP, in particular an intact receptor-binding domain, as critical for tetherin counteraction and provides evidence that antibodies directed against GP1 can interfere with tetherin counteraction.
  • Wakako Furuyama, Andrea Marzi, Aaron B. Carmody, Junki Maruyama, Makoto Kuroda, Hiroko Miyamoto, Asuka Nanbo, Rashid Manzoor, Reiko Yoshida, Manabu Igarashi, Heinz Feldmann, Ayato Takada
    PLOS PATHOGENS 12 (12) e1006139  1553-7366 2016/12 [Refereed][Not invited]
     
    Antibody-dependent enhancement (ADE) of Ebola virus (EBOV) infection has been demonstrated in vitro, raising concerns about the detrimental potential of some anti-EBOV antibodies. ADE has been described for many viruses and mostly depends on the cross-linking of virus-antibody complexes to cell surface Fc receptors, leading to enhanced infection. However, little is known about the molecular mechanisms underlying this phenomenon. Here we show that Fc gamma-receptor IIa (Fc gamma RIIa)-mediated intracellular signaling through Src family protein tyrosine kinases (PTKs) is required for ADE of EBOV infection. We found that deletion of the Fc gamma RIIa cytoplasmic tail abolished EBOV ADE due to decreased virus uptake into cellular endosomes. Furthermore, EBOV ADE, but not non-ADE infection, was significantly reduced by inhibition of the Src family protein PTK pathway, which was also found to be important to promote phagocytosis/macropinocytosis for viral uptake into endosomes. We further confirmed a significant increase of the Src phosphorylation mediated by ADE. These data suggest that antibody-EBOV complexes bound to the cell surface Fc gamma RIIa activate the Src signaling pathway that leads to enhanced viral entry into cells, providing a novel perspective for the general understanding of ADE of virus infection.
  • Edgar Simulundu, Aaron S. Mweene, Katendi Changula, Mwaka Monze, Elizabeth Chizema, Peter Mwaba, Ayato Takada, Guiseppe Ippolito, Francis Kasolo, Alimuddin Zumla, Matthew Bates
    REVIEWS IN MEDICAL VIROLOGY 26 (6) 446 - 454 1052-9276 2016/11 [Refereed][Not invited]
     
    Lujo virus is a novel Old World arenavirus identified in Southern Africa in 2008 as the cause of a viral hemorrhagic fever (VHF) characterized by nosocomial transmission with a high case fatality rate of 80% (4/5 cases). Whereas this outbreak was limited, the unprecedented Ebola virus disease outbreak in West Africa, and recent Zika virus disease epidemic in the Americas, has brought into acute focus the need for preparedness to respond to rare but potentially highly pathogenic outbreaks of zoonotic or arthropod-borne viral infections. A key determinant for effective control of a VHF outbreak is the time between primary infection and diagnosis of the index case. Here, we review the Lujo VHF outbreak of 2008 and discuss how preparatory measures with respect to developing diagnostic capacity might be effectively embedded into existing national disease control networks, such as those for human immunodeficiency virus, tuberculosis, and malaria.
  • Hiromichi Mitake, Yuji Fujii, Makoto Nagai, Naoto Ito, Kota Okadera, Kazuma Okada, Kento Nakagawa, Mai Kishimoto, Tetsuya Mizutani, Katsunori Okazaki, Yoshihiro Sakoda, Ayato Takada, Makoto Sugiyama
    JOURNAL OF GENERAL VIROLOGY 97 (8) 1818 - 1822 0022-1317 2016/08 [Refereed][Not invited]
     
    Ljungan virus (LV) has been isolated/detected from rodents in a limited area including European countries and the USA. In this study, we isolated an LV strain from faecal samples of wild birds that had been collected in Japan, and determined the nearly complete sequence of the genome. Sequence analyses showed that the isolate possesses an LV-like genomic organization: 5UTR-VP0-VP3-VP1-2A1-2A2-2B-2C-3A-3B-3C-3D-3UTR. Phylogenetic and similarity analyses based on the VP1 region indicated that the strain constitutes a novel genotype within LV. In addition, we identified species origin of the faeces as gull species by using the DNA barcoding technique. These data suggested that the novel LV strain infected a gull species, in which the virus had not been identified. Taken together, this study has provided the first evidence of the presence of a novel LV in Japan, highlighting the possibility of LV infection in birds.
  • Claudio L. Afonso, Gaya K. Amarasinghe, Krisztian Banyai, Yiming Bao, Christopher F. Basler, Sina Bavari, Nicolas Bejerman, Kim R. Blasdell, Francois-Xavier Briand, Thomas Briese, Alexander Bukreyev, Charles H. Calisher, Kartik Chandran, Jiasen Cheng, Anna N. Clawson, Peter L. Collins, Ralf G. Dietzgen, Olga Dolnik, Leslie L. Domier, Ralf Duerrwald, John M. Dye, Andrew J. Easton, Hideki Ebihara, Szilvia L. Farkas, Juliana Freitas-Astua, Pierre Formenty, Ron A. M. Fouchier, Yanping Fu, Elodie Ghedin, Michael M. Goodin, Roger Hewson, Masayuki Horie, Timothy H. Hyndman, Daohong Jiang, Elliot W. Kitajima, Gary P. Kobinger, Hideki Kondo, Gael Kurath, Robert A. Lamb, Sergio Lenardon, Eric M. Leroy, Ci-Xiu Li, Xian-Dan Lin, Lijiang Liu, Ben Longdon, Szilvia Marton, Andrea Maisner, Elke Muhlberger, Sergey V. Netesov, Norbert Nowotny, Jean L. Patterson, Susan L. Payne, Janusz T. Paweska, Rick E. Randall, Bertus K. Rima, Paul Rota, Dennis Rubbenstroth, Martin Schwemmle, Mang Shi, Sophie J. Smither, Mark D. Stenglein, David M. Stone, Ayato Takada, Calogero Terregino, Robert B. Tesh, Jun-Hua Tian, Keizo Tomonaga, Noel Tordo, Jonathan S. Towner, Nikos Vasilakis, Martin Verbeek, Viktor E. Volchkov, Victoria Wahl-Jensen, John A. Walsh, Peter J. Walker, David Wang, Lin-Fa Wang, Thierry Wetzel, Anna E. Whitfield, Jiatao Xie, Kwok-Yung Yuen, Yong-Zhen Zhang, Jens H. Kuhn
    ARCHIVES OF VIROLOGY 161 (8) 2351 - 2360 0304-8608 2016/08 [Refereed][Not invited]
     
    In 2016, the order Mononegavirales was emended through the addition of two new families (Mymonaviridae and Sunviridae), the elevation of the paramyxoviral subfamily Pneumovirinae to family status (Pneumoviridae), the addition of five free-floating genera (Anphevirus, Arlivirus, Chengtivirus, Crustavirus, and Wastrivirus), and several other changes at the genus and species levels. This article presents the updated taxonomy of the order Mononegavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV).
  • Taisho Yamada, Hiromasa Horimoto, Takeshi Kameyama, Sumio Hayakawa, Hiroaki Yamato, Masayoshi Dazai, Ayato Takada, Hiroshi Kida, Debbie Bott, Angela C. Zhou, David Hutin, Tania H. Watts, Masahiro Asaka, Jason Matthews, Akinori Takaoka
    NATURE IMMUNOLOGY 17 (6) 687 - + 1529-2908 2016/06 [Refereed][Not invited]
     
    Aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that mediates the toxic activity of many environmental xenobiotics. However, its role in innate immune responses during viral infection is not fully understood. Here we demonstrate that constitutive AHR signaling negatively regulates the type I interferon (IFN-I) response during infection with various types of virus. Virus-induced IFN-beta production was enhanced in AHR-deficient cells and mice and resulted in restricted viral replication. We found that AHR upregulates expression of the ADP-ribosylase TIPARP, which in turn causes downregulation of the IFN-I response. Mechanistically, TIPARP interacted with the kinase TBK1 and suppressed its activity by ADP-ribosylation. Thus, this study reveals the physiological importance of endogenous activation of AHR signaling in shaping the IFN-I-mediated innate response and, further, suggests that the AHR-TIPARP axis is a potential therapeutic target for enhancing antiviral responses.
  • Jonas Thoromo, Edgar Simulundu, Herman M. Chambaro, Liywalii Mataa, Caesar H. Lubaba, Girja S. Pandey, Ayato Takada, Gerald Misinzo, Aaron S. Mweene
    ONDERSTEPOORT JOURNAL OF VETERINARY RESEARCH 83 (1) a1095  0030-2465 2016/04 [Refereed][Not invited]
     
    In early 2015, a highly fatal haemorrhagic disease of domestic pigs resembling African swine fever (ASF) occurred in North Western, Copperbelt, and Lusaka provinces of Zambia. Molecular diagnosis by polymerase chain reaction targeting specific amplification of p72 (B646L) gene of ASF virus (ASFV) was conducted. Fourteen out of 16 domestic pigs from the affected provinces were found to be positive for ASFV. Phylogenetic analyses based on part of the p72 and the complete p54 (E183L) genes revealed that all the ASFVs detected belonged to genotypes I and Id, respectively. Additionally, epidemiological data suggest that the same ASFV spread from Lusaka to other provinces possibly through uncontrolled and/or illegal pig movements. Although the origin of the ASFV that caused outbreaks in domestic pigs in Zambia could not be ascertained, it appears likely that the virus may have emerged from within the country or region, probably from a sylvatic cycle. It is recommended that surveillance of ASF, strict biosecurity, and quarantine measures be imposed in order to prevent further spread and emergence of new ASF outbreaks in Zambia.
  • Kunda Ndashe, Edgar Simulundu, Bernard M. Hang'ombe, Ladslav Moonga, Hirohito Ogawa, Ayato Takada, Aaron S. Mweene
    ARCHIVES OF VIROLOGY 161 (3) 513 - 519 0304-8608 2016/03 [Refereed][Not invited]
     
    Infectious bursal disease (IBD) is an acute, highly contagious, and immunosuppressive viral disease of young chickens and remains one of the economically most important diseases threatening the poultry industry worldwide. In this study, 16 and 11 nucleotide sequences of the VP2 hypervariable region (VP2-HVR) and part of VP1, respectively, of IBD virus (IBDV) detected in vaccinated broiler chickens in Lusaka in 2012 were determined. Phylogenetic analysis revealed that these Zambian IBDVs separated into three genotypes of very virulent (VV) IBDVs. Although the majority of these viruses belonged to the African VV type (VV1), which consisted of viruses from West Africa, South Africa and Zambia, one virus belonged to the East African VV type (VV2). Interestingly, a Zambian IBDV belonging to the VV3 genotype (composed of viruses from several continents) clustered with attenuated vaccine strains. Although sequence analysis of VP2-HVR showed that all detected Zambian IBDVs had conserved putative virulence marker amino acids (i.e., 222A, 242I, 256I, 294I and 299S), one virus had two unique amino acid substitutions, N280S and E300A. This study demonstrates the diversity of Zambian IBDVs and documents for the first time the possible involvement of attenuated vaccine strains in the epidemiology of IBD in Zambia. Strict biosecurity of poultry farms, monitoring of live vaccine use in the field, surveillance and characterization of IBDV in poultry and development of a vaccine from local or regional IBDV field strains are recommended for improved IBD control in Zambia.
  • Wakako Furuyama, Andrea Marzi, Asuka Nanbo, Elaine Haddock, Junki Maruyama, Hiroko Miyamoto, Manabu Igarashi, Reiko Yoshida, Osamu Noyori, Heinz Feldmann, Ayato Takada
    SCIENTIFIC REPORTS 6 20514  2045-2322 2016/02 [Refereed][Not invited]
     
    During the latest outbreak of Ebola virus disease in West Africa, monoclonal antibody therapy (e.g., ZMapp) was utilized to treat patients. However, due to the antigenic differences among the five ebolavirus species, the current therapeutic monoclonal antibodies are only effective against viruses of the species Zaire ebolavirus. Although this particular species has indeed caused the majority of human infections in Central and, recently, West Africa, other ebolavirus species (e.g., Sudan ebolavirus and Bundibugyo ebolavirus) have also repeatedly caused outbreaks in Central Africa and thus should not be neglected in the development of countermeasures against ebolaviruses. Here we report the generation of an ebolavirus glycoprotein-specific monoclonal antibody that effectively inhibits cellular entry of representative isolates of all known ebolavirus species in vitro and show its protective efficacy in mouse models of ebolavirus infections. This novel neutralizing monoclonal antibody targets a highly conserved internal fusion loop in the glycoprotein molecule and prevents membrane fusion of the viral envelope with cellular membranes. The discovery of this highly cross-neutralizing antibody provides a promising option for broad-acting ebolavirus antibody therapy and will accelerate the design of improved vaccines that can selectively elicit cross-neutralizing antibodies against multiple species of ebolaviruses.
  • Kurosaki Y, Magassouba N, Oloniniyi OK, Cherif MS, Sakabe S, Takada A, Hirayama K, Yasuda J
    PLoS neglected tropical diseases 10 (2) e0004472  1935-2727 2016/02 [Refereed][Not invited]
  • Junki Maruyama, Naganori Nao, Hiroko Miyamoto, Ken Maeda, Hirohito Ogawa, Reiko Yoshida, Manabu Igarashi, Ayato Takada
    VIROLOGY 488 43 - 50 0042-6822 2016/01 [Refereed][Not invited]
     
    Recently found bat-derived influenza viruses (BatlVs) have hemagglutinin (HA) and neuraminidase (NA) gene segments distinct from those of previously known influenza A viruses. However, pathogenicities of these BatlVs remain unknown since infectious virus strains have not been isolated yet. To gain insight into the biological properties of BatlVs, we generated vesicular stomatitis viruses (VSVs) pseudotyped with the BatIV HA and NA. We found that VSVs pseudotyped with BatIV HAs and NAs efficiently infected particular bat cell lines but not those derived from primates, and that proteolytic cleavage with a trypsinlike protease was necessary for HA-mediated virus entry. Treatment of the susceptible bat cells with some enzymes and inhibitors revealed that BatIV HAs might recognize some cellular glycoproteins as receptors rather than the sialic acids used for the other known influenza viruses. These data provide fundamental information on the mechanisms underlying the cellular entry and host restriction of BatIVs. (C) 2015 Elsevier Inc. All rights reserved.
  • Florian Wrensch, Christina B. Karsten, Kerstin Gnirss, Markus Hoffmann, Kai Lu, Ayato Takada, Michael Winkler, Graham Simmons, Stefan Poehlmann
    JOURNAL OF INFECTIOUS DISEASES 212 S210 - S218 0022-1899 2015/10 [Refereed][Not invited]
     
    Ebolaviruses are highly pathogenic in humans and nonhuman primates and pose a severe threat to public health. The interferon-induced transmembrane (IFITM) proteins can restrict entry of ebolaviruses, influenza A viruses, and other enveloped viruses. However, the breadth and mechanism of the antiviral activity of IFITM proteins are incompletely understood. Here, we employed ebolavirus glycoprotein-pseudotyped vectors and ebolavirus-like particles to address this question. We show that IFITM proteins inhibit the cellular entry of diverse ebolaviruses and demonstrate that type I interferon induces IFITM protein expression in macrophages, major viral targets. Moreover, we show that IFITM proteins block entry of influenza A viruses and ebolaviruses by different mechanisms and provide evidence that antibodies and IFITM proteins can synergistically inhibit cellular entry of ebolaviruses. These results provide insights into the role of IFITM proteins in infection by ebolaviruses and suggest a mechanism by which antibodies, though poorly neutralizing in vitro, might contribute to viral control in vivo.
  • Hirohito Ogawa, Hiroko Miyamoto, Eri Nakayama, Reiko Yoshida, Ichiro Nakamura, Hirofumi Sawa, Akihiro Ishii, Yuka Thomas, Emiko Nakagawa, Keita Matsuno, Masahiro Kajihara, Junki Maruyama, Naganori Nao, Mieko Muramatsu, Makoto Kuroda, Edgar Simulundu, Katendi Changula, Bernard Hang'ombe, Boniface Namangala, Andrew Nambota, Jackson Katampi, Manabu Igarashi, Kimihito Ito, Heinz Feldmann, Chihiro Sugimoto, Ladslav Moonga, Aaron Mweene, Ayato Takada
    JOURNAL OF INFECTIOUS DISEASES 212 S101 - S108 0022-1899 2015/10 [Refereed][Not invited]
     
    Fruit bats are suspected to be a natural reservoir of filoviruses, including Ebola and Marburg viruses. Using an enzyme-linked immunosorbent assay based on the viral glycoprotein antigens, we detected filovirus-specific immunoglobulin G antibodies in 71 of 748 serum samples collected from migratory fruit bats (Eidolon helvum) in Zambia during 2006-2013. Although antibodies to African filoviruses (eg, Zaire ebolavirus) were most prevalent, some serum samples showed distinct specificity for Reston ebolavirus, which that has thus far been found only in Asia. Interestingly, the transition of filovirus species causing outbreaks in Central and West Africa during 2005-2014 seemed to be synchronized with the change of the serologically dominant virus species in these bats. These data suggest the introduction of multiple species of filoviruses in the migratory bat population and point to the need for continued surveillance of filovirus infection of wild animals in sub-Saharan Africa, including hitherto nonendemic countries.
  • Naganori Nao, Masahiro Kajihara, Rashid Manzoor, Junki Maruyama, Reiko Yoshida, Mieko Muramatsu, Hiroko Miyamoto, Manabu Igarashi, Nao Eguchi, Masahiro Sato, Tatsunari Kondoh, Masatoshi Okamatsu, Yoshihiro Sakoda, Hiroshi Kida, Ayato Takada
    PLOS ONE 10 (9) e0137989  1932-6203 2015/09 [Refereed][Not invited]
     
    Two highly pathogenic avian influenza virus strains, A/duck/Hokkaido/WZ83/2010 ( H5N1) (WZ83) and A/duck/Hokkaido/WZ101/2010 (H5N1) (WZ101), which were isolated from wild ducks in Japan, were found to be genetically similar, with only two amino acid differences in their M1 and PB1 proteins at positions 43 and 317, respectively. We found that both WZ83 and WZ101 caused lethal infection in chickens but WZ101 killed them more rapidly than WZ83. Interestingly, ducks experimentally infected with WZ83 showed no or only mild clinical symptoms, whereas WZ101 was highly lethal. We then generated reassortants between these viruses and found that exchange of the M gene segment completely switched the pathogenic phenotype in both chickens and ducks, indicating that the difference in the pathogenicity for these avian species between WZ83 and WZ101 was determined by only a single amino acid in the M1 protein. It was also found that WZ101 showed higher pathogenicity than WZ83 in mice and that WZ83, whose M gene was replaced with that of WZ101, showed higher pathogenicity than wild-type WZ83, although this reassortant virus was not fully pathogenic compared to wild-type WZ101. These results suggest that the amino acid at position 43 of the M1 protein is one of the factors contributing to the pathogenicity of H5N1 highly pathogenic avian influenza viruses in both avian and mammalian hosts.
  • Takahiro Hiono, Ayako Ohkawara, Kohei Ogasawara, Masatoshi Okamatsu, Tomokazu Tamura, Duc-Huy Chu, Mizuho Suzuki, Saya Kuribayashi, Shintaro Shichinohe, Ayato Takada, Hirohito Ogawa, Reiko Yoshida, Hiroko Miyamoto, Naganori Nao, Wakako Furuyama, Junki Maruyama, Nao Eguchi, Gerelmaa Ulziibat, Bazarragchaa Enkhbold, Munkhduuren Shatar, Tserenjav Jargalsaikhan, Selenge Byambadorj, Batchuluun Damdinjav, Yoshihiro Sakoda, Hiroshi Kida
    VIRUS GENES 51 (1) 57 - 68 0920-8569 2015/08 [Refereed][Not invited]
     
    Migratory water birds are the natural reservoir of influenza A viruses. H5 and H7 influenza viruses are isolated over the world and also circulate among poultry in Asia. In 2010, two H5N1 highly pathogenic avian influenza viruses (HPAIVs) were isolated from fecal samples of water birds on the flyway of migration from Siberia, Russia to the south in Hokkaido, Japan. H7N9 viruses are sporadically isolated from humans and circulate in poultry in China. To monitor whether these viruses have spread in the wild bird population, we conducted virological surveillance of avian influenza in migratory water birds in autumn from 2010 to 2014. A total of 8103 fecal samples from migratory water birds were collected in Japan and Mongolia, and 350 influenza viruses including 13 H5 and 19 H7 influenza viruses were isolated. A phylogenetic analysis revealed that all isolates are genetically closely related to viruses circulating among wild water birds. The results of the antigenic analysis indicated that the antigenicity of viruses in wild water birds is highly stable despite their nucleotide sequence diversity but is distinct from that of HPAIVs recently isolated in Asia. The present results suggest that HPAIVs and Chinese H7N9 viruses were not predominantly circulating in migratory water birds; however, continued monitoring of H5 and H7 influenza viruses both in domestic and wild birds is recommended for the control of avian influenza.
  • Hirohito Ogawa, Nobuo Koizumi, Aiko Ohnuma, Alisheke Mutemwa, Bernard M. Hang'ombe, Aaron S. Mweene, Ayato Takada, Chihiro Sugimoto, Yasuhiko Suzuki, Hiroshi Kida, Hirofumi Sawa
    INFECTION GENETICS AND EVOLUTION 32 143 - 147 1567-1348 2015/06 [Refereed][Not invited]
     
    The role played by bats as a potential source of transmission of Leptospira spp. to humans is poorly understood, despite various pathogenic Leptospira spp. being identified in these mammals. Here, we investigated the prevalence and diversity of pathogenic Leptospira spp. that infect the straw-colored fruit bat (Eidolon helvum). We captured this bat species, which is widely distributed in Africa, in Zambia during 2008-2013. We detected the flagellin B gene (flaB) from pathogenic Leptospira spp. in kidney samples from 79 of 529 E. helvum (14.9%) bats. Phylogenetic analysis of 70 flaB fragments amplified from E. helvum samples and previously reported sequences, revealed that 12 of the fragments grouped with Leptospira borgpetersenii and Leptospira kirschneri; however, the remaining 58 flaB fragments appeared not to be associated with any reported species. Additionally, the 16S ribosomal RNA gene (rrs) amplified from 27 randomly chosen flaB-positive samples was compared with previously reported sequences, including bat-derived Leptospira spp. All 27 rrs fragments clustered into a pathogenic group. Eight fragments were located in unique branches, the other 19 fragments were closely related to Leptospira spp. detected in bats. These results show that rrs sequences in bats are genetically related to each other without regional variation, suggesting that Leptospira are evolutionarily well-adapted to bats and have uniquely evolved in the bat population. Our study indicates that pathogenic Leptospira spp. in E. helvum in Zambia have unique genotypes. (C) 2015 The Authors. Published by Elsevier B.V.
  • Makoto Kuroda, Daisuke Fujikura, Asuka Nanbo, Andrea Marzi, Osamu Noyori, Masahiro Kajihara, Junki Maruyama, Keita Matsuno, Hiroko Miyamoto, Reiko Yoshida, Heinz Feldmann, Ayato Takada
    JOURNAL OF VIROLOGY 89 (12) 6481 - 6493 0022-538X 2015/06 [Refereed][Not invited]
     
    Multiple host molecules are known to be involved in the cellular entry of filoviruses, including Ebola virus (EBOV); T-cell immunoglobulin and mucin domain 1 (TIM-1) and Niemann-Pick C1 (NPC1) have been identified as attachment and fusion receptors, respectively. However, the molecular mechanisms underlying the entry process have not been fully understood. We found that TIM-1 and NPC1 colocalized and interacted in the intracellular vesicles where EBOV glycoprotein (GP)-mediated membrane fusion occurred. Interestingly, a TIM-1-specific monoclonal antibody (MAb), M224/1, prevented GP-mediated membrane fusion and also interfered with the binding of TIM-1 to NPC1, suggesting that the interaction between TIM-1 and NPC1 is important for filovirus membrane fusion. Moreover, MAb M224/1 efficiently inhibited the cellular entry of viruses from all known filovirus species. These data suggest a novel mechanism underlying filovirus membrane fusion and provide a potential cellular target for antiviral compounds that can be universally used against filovirus infections.
  • Keita Matsuno, Carla Weisend, Masahiro Kajihara, Colette Matysiak, Brandi N. Williamson, Martin Simuunza, Aaron S. Mweene, Ayato Takada, Robert B. Tesh, Hideki Ebihara
    JOURNAL OF VIROLOGY 89 (1) 594 - 604 0022-538X 2015/01 [Refereed][Not invited]
     
    Until the recent emergence of two human-pathogenic tick-borne phleboviruses (TBPVs) (severe fever with thrombocytopenia syndrome virus [SFTSV] and Heartland virus), TBPVs have been neglected as causative agents of human disease. In particular, no studies have addressed the global distribution of TBPVs, and consequently, our understanding of the mechanism(s) underlying their evolution and emergence remains poor. In order to provide a useful tool for the ecological and epidemiological study of TBPVs, we have established a simple system that can detect all known TBPVs, based on conventional reverse transcription-PCR (RT-PCR) with degenerate primer sets targeting conserved regions of the viral L genome segment. Using this system, we have determined that several viruses that had been isolated from ticks decades ago but had not been taxonomically identified are novel TBPVs. Full-genome sequencing of these viruses revealed a novel fourth TBPV cluster distinct from the three known TBPV clusters (i.e., the SETS, Bhanja, and Uukuniemi groups) and from the mosquito/sandfly-borne phleboviruses. Furthermore, by using tick samples collected in Zambia, we confirmed that our system had enough sensitivity to detect a new TBPV in a single tick homogenate. This virus, tentatively designated Shibuyunji virus after the region of tick collection, grouped into a novel fourth TBPV cluster. These results indicate that our system can be used as a first-line screening approach for TBPVs and that this kind of work will undoubtedly lead to the discovery of additional novel tick viruses and will expand our knowledge of the evolution and epidemiology of TBPVs. IMPORTANCE Tick-borne phleboviruses (TBPVs) have been largely neglected until the recent emergence of two virulent viruses, severe fever with thrombocytopenia syndrome virus and Heartland virus. Little is known about the global distribution of TBPVs or how these viruses evolved and emerged. A major hurdle to study the distribution of TBPVs is the lack of tools to detect these genetically divergent phleboviruses. In order to address this issue, we have developed a simple, rapid, and cheap RT-PCR system that can detect all known TBPVs and which led to the identification of several novel phleboviruses from previously uncharacterized tick-associated virus isolates. Our system can detect virus in a single tick sample and novel TBPVs that are genetically distinct from any of the known TBPVs. These results indicate that our system will be a useful tool for the surveillance of TBPVs and will facilitate understanding of the ecology of TBPVs.
  • Makoto Kuroda, Daisuke Fujikura, Osamu Noyori, Masahiro Kajihara, Junki Maruyama, Hiroko Miyamoto, Reiko Yoshida, Ayato Takada
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 455 (3-4) 223 - 228 0006-291X 2014/12 [Refereed][Not invited]
     
    Filoviruses, including Ebola and Marburg viruses, cause severe hemorrhagic fever in humans and nonhuman primates with mortality rates of up to 90%. Human T-cell immunoglobulin and mucin domain 1 (TIM-I) is one of the host proteins that have been shown to promote filovirus entry into cells. In this study, we cloned TIM-1 genes from three different African green monkey kidney cell lines (Vero E6, COS-I, and BSC-1) and found that TIM-1 of Vero E6 had a 23-amino acid deletion and 6 amino acid substitutions compared with those of COS-1 and BSC-1. Interestingly, Vero E6 TIM-I had a greater ability to promote the infectivity of vesicular stomatitis viruses pseudotyped with filovirus glycoproteins than COS-1-derived TIM-I. We further found that the increased ability of Vero E6 TIM-1 to promote virus infectivity was most likely due to a single amino acid difference between these TIM-1s. These results suggest that a polymorphism of the TIM-I molecules is one of the factors that influence cell susceptibility to filovirus infection, providing a new insight into the molecular basis for the filovirus host range. (C) 2014 Elsevier Inc. All rights reserved.
  • Edgar Simulundu, Naganori Nao, John Yabe, Nilton A. Muto, Thami Sithebe, Hirofumi Sawa, Rashid Manzoor, Masahiro Kajihara, Mieko Muramatsu, Akihiro Ishii, Hirohito Ogawa, Aaron S. Mweene, Ayato Takada
    ARCHIVES OF VIROLOGY 159 (10) 2633 - 2640 0304-8608 2014/10 [Refereed][Not invited]
     
    Whilst remarkable progress in elucidating the mechanisms governing interspecies transmission and pathogenicity of highly pathogenic avian influenza viruses (AIVs) has been made, similar studies focusing on low-pathogenic AIVs isolated from the wild waterfowl reservoir are limited. We previously reported that two AIV strains (subtypes H6N2 and H3N8) isolated from wild waterfowl in Zambia harbored some amino acid residues preferentially associated with human influenza virus proteins (so-called human signatures) and replicated better in the lungs of infected mice and caused more morbidity than a strain lacking such residues. To further substantiate these observations, we infected chickens and mice intranasally with AIV strains of various subtypes (H3N6, H3N8, H4N6, H6N2, H9N1 and H11N9) isolated from wild waterfowl in Zambia. Although some strains induced seroconversion, all of the tested strains replicated poorly and were nonpathogenic for chickens. In contrast, most of the strains having human signatures replicated well in the lungs of mice, and one of these strains caused severe illness in mice and induced lung injury that was characterized by a severe accumulation of polymorphonuclear leukocytes. These results suggest that some strains tested in this study may have the potential to infect mammalian hosts directly without adaptation, which might possibly be associated with the possession of human signature residues. Close monitoring and evaluation of host-associated signatures may help to elucidate the prevalence and emergence of AIVs with potential for causing zoonotic infections.
  • Jens H. Kuhn, Kristian G. Andersen, Yiming Bao, Sina Bavari, Stephan Becker, Richard S. Bennett, Nicholas H. Bergman, Olga Blinkova, Steven Bradfute, J. Rodney Brister, Alexander Bukreyev, Kartik Chandran, Alexander A. Chepurnov, Robert A. Davey, Ralf G. Dietzgen, Norman A. Doggett, Olga Dolnik, John M. Dye, Sven Enterlein, Paul W. Fenimore, Pierre Formenty, Alexander N. Freiberg, Robert F. Garry, Nicole L. Garza, Stephen K. Gire, Jean-Paul Gonzalez, Anthony Griffiths, Christian T. Happi, Lisa E. Hensley, Andrew S. Herbert, Michael C. Hevey, Thomas Hoenen, Anna N. Honko, Georgy M. Ignatyev, Peter B. Jahrling, Joshua C. Johnson, Karl M. Johnson, Jason Kindrachuk, Hans-Dieter Klenk, Gary Kobinger, Tadeusz J. Kochel, Matthew G. Lackemeyer, Daniel F. Lackner, Eric M. Leroy, Mark S. Lever, Elke Muehlberger, Sergey V. Netesov, Gene G. Olinger, Sunday A. Omilabu, Gustavo Palacios, Rekha G. Panchal, Daniel J. Park, Jean L. Patterson, Janusz T. Paweska, Clarence J. Peters, James Pettitt, Louise Pitt, Sheli R. Radoshitzky, Elena I. Ryabchikova, Erica Ollmann Saphire, Pardis C. Sabeti, Rachel Sealfon, Aleksandr M. Shestopalov, Sophie J. Smither, Nancy J. Sullivan, Robert Swanepoel, Ayato Takada, Jonathan S. Towner, Guido van der Groen, Viktor E. Volchkov, Valentina A. Volchkova, Victoria Wahl-Jensen, Travis K. Warren, Kelly L. Warfield, Manfred Weidmann, Stuart T. Nichol
    VIRUSES-BASEL 6 (9) 3663 - 3682 1999-4915 2014/09 [Refereed][Not invited]
     
    Sequence determination of complete or coding-complete genomes of viruses is becoming common practice for supporting the work of epidemiologists, ecologists, virologists, and taxonomists. Sequencing duration and costs are rapidly decreasing, sequencing hardware is under modification for use by non-experts, and software is constantly being improved to simplify sequence data management and analysis. Thus, analysis of virus disease outbreaks on the molecular level is now feasible, including characterization of the evolution of individual virus populations in single patients over time. The increasing accumulation of sequencing data creates a management problem for the curators of commonly used sequence databases and an entry retrieval problem for end users. Therefore, utilizing the data to their fullest potential will require setting nomenclature and annotation standards for virus isolates and associated genomic sequences. The National Center for Biotechnology Information's (NCBI's) RefSeq is a non-redundant, curated database for reference (or type) nucleotide sequence records that supplies source data to numerous other databases. Building on recently proposed templates for filovirus variant naming [<virus name> (<strain>)/<isolation host-suffix>/<country of sampling>/<year of sampling>/<genetic variant designation>-<isolate designation>], we report consensus decisions from a majority of past and currently active filovirus experts on the eight filovirus type variants and isolates to be represented in RefSeq, their final designations, and their associated sequences.
  • Yasushi Itoh, Reiko Yoshida, Shintaro Shichinohe, Megumi Higuchi, Hirohito Ishigaki, Misako Nakayama, Van Loi Pham, Hideaki Ishida, Mitsutaka Kitano, Masahiko Arikata, Naoko Kitagawa, Yachiyo Mitsuishi, Kazumasa Ogasawara, Hideaki Tsuchiya, Takahiro Hiono, Masatoshi Okamatsu, Yoshihiro Sakoda, Hiroshi Kida, Mutsumi Ito, Le Quynh Mai, Yoshihiro Kawaoka, Hiroko Miyamoto, Mari Ishijima, Manabu Igarashi, Yasuhiko Suzuki, Ayato Takada
    PLOS PATHOGENS 10 (6) e1004192  1553-7366 2014/06 [Refereed][Not invited]
     
    Highly pathogenic avian influenza (HPAI) viruses of the H5N1 subtype often cause severe pneumonia and multiple organ failure in humans, with reported case fatality rates of more than 60%. To develop a clinical antibody therapy, we generated a human-mouse chimeric monoclonal antibody (MAb) ch61 that showed strong neutralizing activity against H5N1 HPAI viruses isolated from humans and evaluated its protective potential in mouse and nonhuman primate models of H5N1 HPAI virus infections. Passive immunization with MAb ch61 one day before or after challenge with a lethal dose of the virus completely protected mice, and partial protection was achieved when mice were treated 3 days after the challenge. In a cynomolgus macaque model, reduced viral loads and partial protection against lethal infection were observed in macaques treated with MAb ch61 intravenously one and three days after challenge. Protective effects were also noted in macaques under immunosuppression. Though mutant viruses escaping from neutralization by MAb ch61 were recovered from macaques treated with this MAb alone, combined treatment with MAb ch61 and peramivir reduced the emergence of escape mutants. Our results indicate that antibody therapy might be beneficial in reducing viral loads and delaying disease progression during H5N1 HPAI virus infection in clinical cases and combined treatment with other antiviral compounds should improve the protective effects of antibody therapy against H5N1 HPAI virus infection.
  • Jens H. Kuhn, Yiming Bao, Sina Bavari, Stephan Becker, Steven Bradfute, Kristina Brauburger, J. Rodney Brister, Alexander A. Bukreyev, Yingyun Cai, Kartik Chandran, Robert A. Davey, Olga Dolnik, John M. Dye, Sven Enterlein, Jean-Paul Gonzalez, Pierre Formenty, Alexander N. Freiberg, Lisa E. Hensley, Thomas Hoenen, Anna N. Honko, Georgy M. Ignatyev, Peter B. Jahrling, Karl M. Johnson, Hans-Dieter Klenk, Gary Kobinger, Matthew G. Lackemeyer, Eric M. Leroy, Mark S. Lever, Elke Muehlberger, Sergey V. Netesov, Gene G. Olinger, Gustavo Palacios, Jean L. Patterson, Janusz T. Paweska, Louise Pitt, Sheli R. Radoshitzky, Elena I. Ryabchikova, Erica Ollmann Saphire, Aleksandr M. Shestopalov, Sophie J. Smither, Nancy J. Sullivan, Robert Swanepoel, Ayato Takada, Jonathan S. Towner, Guido van der Groen, Viktor E. Volchkov, Valentina A. Volchkova, Victoria Wahl-Jensen, Travis K. Warren, Kelly L. Warfield, Manfred Weidmann, Stuart T. Nichol
    ARCHIVES OF VIROLOGY 159 (5) 1229 - 1237 0304-8608 2014/05 [Refereed][Not invited]
     
    Specific alterations (mutations, deletions, insertions) of virus genomes are crucial for the functional characterization of their regulatory elements and their expression products, as well as a prerequisite for the creation of attenuated viruses that could serve as vaccine candidates. Virus genome tailoring can be performed either by using traditionally cloned genomes as starting materials, followed by site-directed mutagenesis, or by de novo synthesis of modified virus genomes or parts thereof. A systematic nomenclature for such recombinant viruses is necessary to set them apart from wild-type and laboratory-adapted viruses, and to improve communication and collaborations among researchers who may want to use recombinant viruses or create novel viruses based on them. A large group of filovirus experts has recently proposed nomenclatures for natural and laboratory animal-adapted filoviruses that aim to simplify the retrieval of sequence data from electronic databases. Here, this work is extended to include nomenclature for filoviruses obtained in the laboratory via reverse genetics systems. The previously developed template for natural filovirus genetic variant naming, < virus name > (< strain >/)< isolation host-suffix >/< country of sampling >/< year of sampling >/< genetic variant designation >-< isolate designation >, is retained, but we propose to adapt the type of information added to each field for cDNA clone-derived filoviruses. For instance, the full-length designation of an Ebola virus Kikwit variant rescued from a plasmid developed at the US Centers for Disease Control and Prevention could be akin to "Ebola virus H.sapiens-rec/COD/1995/Kikwit-abc1" (with the suffix "rec" identifying the recombinant nature of the virus and "abc1" being a placeholder for any meaningful isolate designator). Such a full-length designation should be used in databases and the methods section of publications. Shortened designations (such as "EBOV H.sap/COD/95/Kik-abc1") and abbreviations (such as "EBOV/Kik-abc1") could be used in the remainder of the text, depending on how critical it is to convey information contained in the full-length name. "EBOV" would suffice if only one EBOV strain/variant/isolate is addressed.
  • Hiroki Yamaguchi, Shintaro Kobayashi, Junki Maruyama, Michihito Sasaki, Ayato Takada, Takashi Kimura, Hirofumi Sawa, Yasuko Orba
    JOURNAL OF VETERINARY MEDICAL SCIENCE 76 (5) 637 - 644 0916-7250 2014/05 [Refereed][Not invited]
     
    Recently, we detected novel vervet monkey polyomavirus 1 (VmPyV) in a vervet monkey. Among amino acid sequences of major capsid protein VP1s of other polyomaviruses, VmPyV VP1 is the longest with additional amino acid residues in the C-terminal region. To examine the role of VmPyV VP1 in virion formation, we generated virus-like particles (VLPs) of VmPyV VP1, because VLP is a useful tool for the investigation of the morphological characters of polyomavirus virions. After the full-length VmPyV VP1 was subcloned into a mammalian expression plasmid, the plasmid was transfected into human embryonic kidney 293T (HEK293T) cells. Thereafter, VmPyV VLPs were purified from the cell lysates of the transfected cells via sucrose gradient sedimentation. Electron microscopic analyses revealed that VmPyV VP1 forms VLPs with a diameter of approximately 50 nm that are exclusively localized in cell nuclei. Furthermore, we generated VLPs consisting of the deletion mutant VmPyV VP1 (Delta C VP1) lacking the C-terminal 116 amino acid residues and compared its VLP formation efficiency and morphology to those of VLPs from wild-type VmPyV VP1 (WT VP1). WT and Delta C VP1 VLPs were similar in size, but the number of Delta C VP1 VLPs was much lower than that of WT VP1 VLPs in VP1-expressing HEK293T cells. These results suggest that the length of VP1 is unrelated to virion morphology; however, the C-terminal region of VmPyV VP1 affects the efficiency of its VLP formation.
  • Alexander A. Bukreyev, Kartik Chandran, Olga Dolnik, John M. Dye, Hideki Ebihara, Eric M. Leroy, Elke Muehlberger, Sergey V. Netesov, Jean L. Patterson, Janusz T. Paweska, Erica Ollmann Saphire, Sophie J. Smither, Ayato Takada, Jonathan S. Towner, Viktor E. Volchkov, Travis K. Warren, Jens H. Kuhn
    ARCHIVES OF VIROLOGY 159 (4) 821 - 830 0304-8608 2014/04 [Refereed][Not invited]
     
    The International Committee on Taxonomy of Viruses (ICTV) Filoviridae Study Group prepares proposals on the classification and nomenclature of filoviruses to reflect current knowledge or to correct disagreements with the International Code of Virus Classification and Nomenclature (ICVCN). In recent years, filovirus taxonomy has been corrected and updated, but parts of it remain controversial, and several topics remain to be debated. This article summarizes the decisions and discussion of the currently acting ICTV Filoviridae Study Group since its inauguration in January 2012.
  • Walter Muleya, Michihito Sasaki, Yasuko Orba, Akihiro Ishii, Yuka Thomas, Emiko Nakagawa, Hirohito Ogawa, Bernard Hang'ombe, Boniface Namangala, Aaron Mweene, Ayato Takada, Takashi Kimura, Hirofumi Sawa
    JOURNAL OF VETERINARY MEDICAL SCIENCE 76 (4) 611 - 614 0916-7250 2014/04 [Refereed][Not invited]
     
    In this study, we describe the detection of novel paramyxoviruses from the Eidolon helvum species of fruit bats. We extracted RNA from 312 spleen samples from bats captured in Zambia over a period of 4 years (2008-2011). Semi-nested RT-PCR detected a total of 25 (8%) positive samples for paramyxoviruses which were then directly sequenced and analyzed using phylogenetic analysis. Among the positive samples, seven novel paramyxoviruses were detected. Five viruses were closely related to the genus Henipavirus, while two viruses were related to the unclassified Bat paramyxoviruses from Ghana and Congo Brazzaville. Our study identified novel Henipavirus-related and unrelated viruses using RT-PCR in fruit bats from Kansaka National Park and indicated the presence of similar Bat paramyxoviruses originating from wide geographic areas, suggesting the ability of bats to harbor and transmit viruses. The presence of these viruses in fruit bats might pose a public health risk.
  • Rashid Manzoor, Kazumichi Kuroda, Reiko Yoshida, Yoshimi Tsuda, Daisuke Fujikura, Hiroko Miyamoto, Masahiro Kajihara, Hiroshi Kida, Ayato Takada
    JOURNAL OF BIOLOGICAL CHEMISTRY 289 (11) 7599 - 7614 0021-9258 2014/03 [Refereed][Not invited]
     
    Background: It has been shown that heat shock protein 70 (Hsp70) plays a role in influenza A virus replication. Results: A correlation between viral replication/transcription activities and nuclear/cytoplasmic shuttling of Hsp70 was observed. Conclusion: Hsp70 modulates the influenza A virus polymerase activity. Significance: This study, for the first time, suggests that Hsp70 may actually assist in influenza A virus replication. The role of heat shock protein 70 (Hsp70) in virus replication has been discussed for many viruses. The known suppressive role of Hsp70 in influenza virus replication is based on studies conducted in cells with various Hsp70 expression levels. In this study, we determined the role of Hsp70 in influenza virus replication in HeLa and HEK293T cells, which express Hsp70 constitutively. Co-immunoprecipitation and immunofluorescence studies revealed that Hsp70 interacted with PB2 or PB1 monomers and PB2/PB1 heterodimer but not with the PB1/PA heterodimer or PB2/PB1/PA heterotrimer and translocated into the nucleus with PB2 monomers or PB2/PB1 heterodimers. Knocking down Hsp70 resulted in reduced virus transcription and replication activities. Reporter gene assay, immunofluorescence assay, and Western blot analysis of nuclear and cytoplasmic fractions from infected cells demonstrated that the increase in viral polymerase activity during the heat shock phase was accompanied with an increase in Hsp70 and viral polymerases levels in the nuclei, where influenza virus replication takes place, whereas a reduction in viral polymerase activity was accompanied with an increase in cytoplasmic relocation of Hsp70 along with viral polymerases. Moreover, significantly higher levels of viral genomic RNA (vRNA) were observed during the heat shock phase than during the recovery phase. Overall, for the first time, these findings suggest that Hsp70 may act as a chaperone for influenza virus polymerase, and the modulatory effect of Hsp70 appears to be a sequel of shuttling of Hsp70 between nuclear and cytoplasmic compartments.
  • Junki Maruyama, Hiroko Miyamoto, Masahiro Kajihara, Hirohito Ogawa, Ken Maeda, Yoshihiro Sakoda, Reiko Yoshida, Ayato Takada
    JOURNAL OF VIROLOGY 88 (1) 99 - 109 0022-538X 2014/01 [Refereed][Not invited]
     
    Lloviu virus (LLOV), a novel filovirus detected in bats, is phylogenetically distinct from viruses in the genera Ebolavirus and Marburgvirus in the family Filoviridae. While filoviruses are known to cause severe hemorrhagic fever in humans and/or nonhuman primates, LLOV is biologically uncharacterized, since infectious LLOV has never been isolated. To examine the properties of LLOV, we characterized its envelope glycoprotein (GP), which likely plays a key role in viral tropism and pathogenicity. We first found that LLOV GP principally has the same primary structure as the other filovirus GPs. Similar to the other filoviruses, virus-like particles (VLPs) produced by transient expression of LLOV GP, matrix protein, and nucleoprotein in 293T cells had densely arrayed GP spikes on a filamentous particle. Mouse antiserum to LLOV VLP was barely cross-reactive to viruses of the other genera, indicating that LLOV is serologically distinct from the other known filoviruses. For functional study of LLOV GP, we utilized a vesicular stomatitis virus (VSV) pseudotype system and found that LLOV GP requires low endosomal pH and cathepsin L, and that human C-type lectins act as attachment factors for LLOV entry into cells. Interestingly, LLOV GP-pseudotyped VSV infected particular bat cell lines more efficiently than viruses bearing other filovirus GPs. These results suggest that LLOV GP mediates cellular entry in a manner similar to that of the other filoviruses while showing preferential tropism for some bat cells.
  • Mieko Muramatsu, Reiko Yoshida, Ayaka Yokoyama, Hiroko Miyamoto, Masahiro Kajihara, Junki Maruyama, Naganori Nao, Rashid Manzoor, Ayato Takada
    PLOS ONE 9 (1) e85582  1932-6203 2014/01 [Refereed][Not invited]
     
    Both IgA and IgG antibodies are known to play important roles in protection against influenza virus infection. While IgG is the major isotype induced systemically, IgA is predominant in mucosal tissues, including the upper respiratory tract. Although IgA antibodies are believed to have unique advantages in mucosal immunity, information on direct comparisons of the in vitro antiviral activities of IgA and IgG antibodies recognizing the same epitope is limited. In this study, we demonstrate differences in antiviral activities between these isotypes using monoclonal IgA and IgG antibodies obtained from hybridomas of the same origin. Polymeric IgA-producing hybridoma cells were successfully subcloned from those originally producing monoclonal antibody S139/1, a hemaggulutinin (HA)-specific IgG that was generated against an influenza A virus strain of the H3 subtype but had cross-neutralizing activities against the H1, H2, H13, and H16 subtypes. These monoclonal S139/1 IgA and IgG antibodies were assumed to recognize the same epitope and thus used to compare their antiviral activities. We found that both S139/1 IgA and IgG antibodies strongly bound to the homologous H3 virus in an enzyme-linked immunosorbent assay, and there were no significant differences in their hemagglutination-inhibiting and neutralizing activities against the H3 virus. In contrast, S139/1 IgA showed remarkably higher cross-binding to and antiviral activities against H1, H2, and H13 viruses than S139/1 IgG. It was also noted that S139/1 IgA, but not IgG, drastically suppressed the extracellular release of the viruses from infected cells. Electron microscopy revealed that S139/1 IgA deposited newly produced viral particles on the cell surface, most likely by tethering the particles. These results suggest that anti-HA IgA has greater potential to prevent influenza A virus infection than IgG antibodies, likely due to increased avidity conferred by its multivalency, and that this advantage may be particularly important for heterosubtypic immunity.
  • Ozaki H, Guan Y, Peiris M, Webster R, Takada A, Webby R
    Influenza research and treatment 2014 547839  2090-1380 2014 [Refereed][Not invited]
  • Osamu Noyori, Keita Matsuno, Masahiro Kajihara, Eri Nakayama, Manabu Igarashi, Makoto Kuroda, Norikazu Isoda, Reiko Yoshida, Ayato Takada
    VIROLOGY 446 (1-2) 152 - 161 0042-6822 2013/11 [Refereed][Not invited]
     
    The viral envelope glycoprotein (GP) is thought to play important roles in the pathogenesis of filovirus infection. It is known that GP expressed on the cell surface forms a steric shield over host proteins such as major histocompatibility complex class I and integrin pi, which may result in the disorder of cell-to-cell contacts and/or inhibition of the immune response. However, it is not clarified whether this phenomenon contributes to the pathogenicity of filoviruses. In this study, we found that the steric shielding efficiency differed among filovirus strains and was correlated with the difference in their relative pathogenicities. While the highly glycosylated mucin-like region of GP was indispensable, the differential shielding efficiency did not necessarily depend on the primary structure of the mucin-like region, suggesting the importance of the overall properties (e.g., flexibility and stability) of the GP molecule for efficient shielding of host proteins. (C) 2013 Elsevier Inc. All rights reserved.
  • Noyori O, Nakayama E, Maruyama J, Yoshida R, Takada A
    Biochemical and biophysical research communications 441 (4) 994 - 998 0006-291X 2013/11 [Refereed][Not invited]
  • Masatoshi Okamatsu, Tatsuya Nishi, Naoki Nomura, Naoki Yamamoto, Yoshihiro Sakoda, Kenji Sakurai, Huy Duc Chu, Long Pham Thanh, Long Van Nguyen, Nam Van Hoang, Tien Ngoc Tien, Reiko Yoshida, Ayato Takada, Hiroshi Kida
    VIRUS GENES 47 (2) 317 - 329 0920-8569 2013/10 [Refereed][Not invited]
     
    To estimate the prevalence of avian influenza virus infection in Vietnam, surveillance was conducted in domestic and wild birds from households, live-bird markets, slaughtering sites, and bird sanctuaries in Vietnam between October 2010 and October 2012. Of the 4,550 samples collected, 226 influenza A virus isolates were obtained from domestic ducks, muscovy ducks, and chickens. Of these, 25 and 22 H5N1 highly pathogenic avian influenza viruses (HPAIVs) were isolated from apparently healthy domestic ducks in live-bird markets and slaughtering sites in northern and southern Vietnam, respectively. The HA genes of H5 viruses isolated from birds in northern Vietnam phylogenetically belonged to the genetic clade 2.3.2.1 and those in southern Vietnam belonged to the genetic clade 1.1. In addition, 39 H3, 12 H4, 1 H5, 93 H6, 2 H7, 18 H9, 3 H10, and 11 H11 viruses were isolated. Phylogenetic and antigenic analyses of the H6 and H9 viruses revealed that they were closely related to the isolates obtained from domestic poultry in China. Phylogenetic analyses of internal gene segments of these isolates revealed that these viruses were circulating in both domestic and wild birds in Asia and reassortment events had occurred frequently. Therefore, it will be important to continue the surveillance and strict controls over the movement and trade of poultry and poultry products in order to eradicate H5N1 HPAIV from Asia.
  • Changula K, Yoshida R, Noyori O, Marzi A, Miyamoto H, Ishijima M, Yokoyama A, Kajihara M, Feldmann H, Mweene AS, Takada A
    Virus research 176 (1-2) 83 - 90 0168-1702 2013/09 [Refereed][Not invited]
  • Thomas Hoenen, Allison Groseth, Julie Callison, Ayato Takada, Heinz Feldmann
    ANTIVIRAL RESEARCH 99 (3) 207 - 213 0166-3542 2013/09 [Refereed][Not invited]
     
    Ebola virus (EBOV) causes a severe hemorrhagic fever with case fatality rates of up to 90%, for which no antiviral therapies are available. Antiviral screening is hampered by the fact that development of cytopathic effect, the easiest means to detect infection with wild-type EBOV, is relatively slow. To overcome this problem we generated a recombinant EBOV carrying a luciferase reporter. Using this virus we show that EBOV entry is rapid, with viral protein expression detectable within 2 h after infection. Further, luminescence-based assays were developed to allow highly sensitive titer determination within 48 h. As a proof-of-concept for its utility in antiviral screening we used this virus to assess neutralizing antibodies and siRNAs, with significantly faster screening times than currently available wild-type or recombinant viruses. The availability of this recombinant virus will allow for more rapid and quantitative evaluation of antivirals against EBOV, as well as the study of details of the EBOV life cycle. Published by Elsevier B.V.
  • Mieko Muramatsu, Reiko Yoshida, Hiroko Miyamoto, Daisuke Tomabechi, Masahiro Kajihara, Junki Maruyama, Takashi Kimura, Rashid Manzoor, Kimihito Ito, Ayato Takada
    PLOS ONE 8 (8) e71534  1932-6203 2013/08 [Refereed][Not invited]
     
    Influenza A virus subtypes are classified on the basis of the antigenicity of their envelope glycoproteins, hemagglutinin (HA; H1-H17) and neuraminidase. Since HA-specific neutralizing antibodies are predominantly specific for a single HA subtype, the contribution of antibodies to the heterosubtypic immunity is not fully understood. In this study, mice were immunized intranasally or subcutaneously with viruses having the H1, H3, H5, H7, H9, or H13 HA subtype, and cross-reactivities of induced IgG and IgA antibodies to recombinant HAs of the H1-H16 subtypes were analyzed. We found that both subcutaneous and intranasal immunizations induced antibody responses to multiple HAs of different subtypes, whereas IgA was not detected remarkably in mice immunized subcutaneously. Using serum, nasal wash, and trachea-lung wash samples of H9 virus-immunized mice, neutralizing activities of cross-reactive antibodies were then evaluated by plaque-reduction assays. As expected, no heterosubtypic neutralizing activity was detected by a standard neutralization test in which viruses were mixed with antibodies prior to inoculation into cultured cells. Interestingly, however, a remarkable reduction of plaque formation and extracellular release of the H12 virus, which was bound by the H9-induced cross-reactive antibodies, was observed when infected cells were subsequently cultured with the samples containing HA-specific cross-reactive IgA. This heterosubtypic plaque reduction was interfered when the samples were pretreated with anti-mouse IgA polyclonal serum. These results suggest that the majority of HA-specific cross-reactive IgG and IgA antibodies produced by immunization do not block cellular entry of viruses, but cross-reactive IgA may have the potential to inhibit viral egress from infected cells and thus to play a role in heterosubtypic immunity against influenza A viruses.
  • Jens H. Kuhn, Yiming Bao, Sina Bavari, Stephan Becker, Steven Bradfute, J. Rodney Brister, Alexander A. Bukreyev, Yingyun Cai, Kartik Chandran, Robert A. Davey, Olga Dolnik, John M. Dye, Sven Enterlein, Jean-Paul Gonzalez, Pierre Formenty, Alexander N. Freiberg, Lisa E. Hensley, Anna N. Honko, Georgy M. Ignatyev, Peter B. Jahrling, Karl M. Johnson, Hans-Dieter Klenk, Gary Kobinger, Matthew G. Lackemeyer, Eric M. Leroy, Mark S. Lever, Loreen L. Lofts, Elke Muehlberger, Sergey V. Netesov, Gene G. Olinger, Gustavo Palacios, Jean L. Patterson, Janusz T. Paweska, Louise Pitt, Sheli R. Radoshitzky, Elena I. Ryabchikova, Erica Ollmann Saphire, Aleksandr M. Shestopalov, Sophie J. Smither, Nancy J. Sullivan, Robert Swanepoel, Ayato Takada, Jonathan S. Towner, Guido van der Groen, Viktor E. Volchkov, Victoria Wahl-Jensen, Travis K. Warren, Kelly L. Warfield, Manfred Weidmann, Stuart T. Nichol
    ARCHIVES OF VIROLOGY 158 (6) 1425 - 1432 0304-8608 2013/06 [Refereed][Not invited]
     
    The International Committee on Taxonomy of Viruses (ICTV) organizes the classification of viruses into taxa, but is not responsible for the nomenclature for taxa members. International experts groups, such as the ICTV Study Groups, recommend the classification and naming of viruses and their strains, variants, and isolates. The ICTV Filoviridae Study Group has recently introduced an updated classification and nomenclature for filoviruses. Subsequently, and together with numerous other filovirus experts, a consistent nomenclature for their natural genetic variants and isolates was developed that aims at simplifying the retrieval of sequence data from electronic databases. This is a first important step toward a viral genome annotation standard as sought by the US National Center for Biotechnology Information (NCBI). Here, this work is extended to include filoviruses obtained in the laboratory by artificial selection through passage in laboratory hosts. The previously developed template for natural filovirus genetic variant naming (< virus name > < isolation host-suffix >/< country of sampling >/< year of sampling >/< genetic variant designation >-< isolate designation >) is retained, but it is proposed to adapt the type of information added to each field for laboratory animal-adapted variants. For instance, the full-length designation of an Ebola virus Mayinga variant adapted at the State Research Center for Virology and Biotechnology "Vector" to cause disease in guinea pigs after seven passages would be akin to "Ebola virus VECTOR/C.porcellus-lab/COD/1976/Mayinga-GPA-P7". As was proposed for the names of natural filovirus variants, we suggest using the full-length designation in databases, as well as in the method section of publications. Shortened designations (such as "EBOV VECTOR/C.por/COD/76/May-GPA-P7") and abbreviations (such as "EBOV/May-GPA-P7") could be used in the remainder of the text depending on how critical it is to convey information contained in the full-length name. "EBOV" would suffice if only one EBOV strain/variant/isolate is addressed.
  • Ichiro Nakamura, Bernard Mudenda Hang'ombe, Hirofumi Sawa, Shintaro Kobayashi, Yasuko Orba, Akihiro Ishii, Yuka Thomas, Rie Isozumi, Kumiko Yoshimatsu, Aaron S. Mweene, Ayato Takada, Chihiro Sugimoto, Jiro Arikawa
    JOURNAL OF VETERINARY MEDICAL SCIENCE 75 (6) 819 - 825 0916-7250 2013/06 [Refereed][Not invited]
     
    A total of 466 rodents were captured in the Republic of Zambia from 2006 to 2010. Based on morphological observations and phylogenetic analyses of mitochondrial gene sequences, rodents were divided into 10 groups consisting of 39 Rattus rodents, 263 multimammate rats, 18 other Murinae rodents, 95 gerbils, 11 pouched mice, 1 giant-pouched rat, 38 fat mice and 1 dormouse. Rodent antibodies except that from Rattus were examined for their cross-reactivity to commercially available antibody detection reagents. Anti-mouse immunoglobulin G (IgG) was most cross-reactive to heterologous antibodies including multimammate rat, gerbil, pouched mouse and fat mouse. Thus, anti-mouse IgG would be a useful detection tool in serological examination of the Zambian rodent population. Preliminary sero-surveillance for plague, leptospirosis and hantavirus infection was performed by ELISA.
  • Masahiro Kajihara, Eri Nakayama, Andrea Marzi, Manabu Igarashi, Heinz Feldmann, Ayato Takada
    JOURNAL OF GENERAL VIROLOGY 94 (Pt 4) 876 - 883 0022-1317 2013/04 [Refereed][Not invited]
     
    Marburg virus (MARV) and Ebola virus, members of the family Filoviridae, cause lethal haemorrhagic fever in humans and non-human primates. Although the outbreaks are concentrated mainly in Central Africa, these viruses are potential agents of imported infectious diseases and bioterrorism in non-African countries. Recent studies demonstrated that non-human primates passively immunized with virus-specific antibodies were successfully protected against fatal filovirus infection, highlighting the important role of antibodies in protective immunity for this disease. However, the mechanisms underlying potential evasion from antibody mediated immune pressure are not well understood. To analyse possible mutations involved in immune evasion in the MARV envelope glycoprotein (GP) which is the major target of protective antibodies, we selected escape mutants of recombinant vesicular stomatitis virus (rVSV) expressing MARV GP (rVSV Delta G/MARVGP) by using two GP-specific mAbs, AGP127-8 and MGP72-17, which have been previously shown to inhibit MARV budding. Interestingly, several rVSV Delta G/MARVGP variants escaping from the mAb pressure-acquired amino acid substitutions in the furin-cleavage site rather than in the mAb-specific epitopes, suggesting that these epitopes are recessed, not exposed on the uncleaved GP molecule, and therefore inaccessible to the mAbs. More surprisingly, some variants escaping mAb MGP72-17 lacked a large proportion of the mucin-like region of GP, indicating that these mutants efficiently escaped the selective pressure by deleting the mucin-like region including the mAb-specific epitope. Our data demonstrate that MARV GP possesses the potential to evade antibody mediated immune pressure due to extraordinary structural flexibility and variability.
  • Masahiro Kajihara, Yoshihiro Sakoda, Kosuke Soda, Kenji Minari, Masatoshi Okamatsu, Ayato Takada, Hiroshi Kida
    VIROLOGY JOURNAL 10 45  1743-422X 2013/02 [Refereed][Not invited]
     
    Background: Wild ducks are the natural hosts of influenza A viruses. Duck influenza, therefore, has been believed inapparent infection with influenza A viruses, including highly pathogenic avian influenza viruses (HPAIVs) in chickens. In fact, ducks experimentally infected with an HPAIV strain, A/Hong Kong/483/1997 (H5N1) (HK483), did not show any clinical signs. Another HPAIV strain, A/whooper swan/Mongolia/3/2005 (H5N1) (MON3) isolated from a dead swan, however, caused neurological dysfunction and death in ducks. Method: To understand the mechanism whereby MON3 shows high pathogenicity in ducks, HK483, MON3, and twenty-four reassortants generated between these two H5N1 viruses were compared for their pathogenicity in domestic ducks. Results: None of the ducks infected with MON3-based single-gene reassortants bearing the PB2, NP, or NS gene segment of HK483 died, and HK483-based single-gene reassortants bearing PB2, NP, or NS genes of MON3 were not pathogenic in ducks, suggesting that multiple gene segments contribute to the pathogenicity of MON3 in ducks. All the ducks infected with the reassortant bearing PB2, PA, HA, NP, and NS gene segments of MON3 died within five days post-inoculation, as did those infected with MON3. Each of the viruses was assessed for replication in ducks three days post-inoculation. MON3 and multi-gene reassortants pathogenic in ducks were recovered from all of the tissues examined and replicated with high titers in the brains and lungs. Conclusion: The present results indicate that multigenic factors are responsible for efficient replication of MON3 in ducks. In particular, virus growth in the brain might correlate with neurological dysfunction and the disease severity.
  • Yonezawa K, Igarashi M, Ueno K, Takada A, Ito K
    PloS one 2 8 (2) e57684  2013 [Refereed][Not invited]
  • Ryo Takano, Maki Kiso, Manabu Igarashi, Quynh Mai Le, Masakazu Sekijima, Kimihito Ito, Ayato Takada, Yoshihiro Kawaoka
    JOURNAL OF INFECTIOUS DISEASES 207 (1) 89 - 97 0022-1899 2013/01 [Refereed][Not invited]
     
    Background. The neuraminidase (NA) inhibitor oseltamivir is widely used to treat patients infected with influenza viruses. An Ile-to-Val change at position 117 in influenza A virus subtype H5N1 NA (NA-I117V) confers a reduction in susceptibility to oseltamivir carboxylate. However, the in vivo relevance and molecular basis of the decreased sensitivity mediated by this mutation are poorly understood. Methods. We created single-point-mutant viruses with 3 genetically different backgrounds (ie, 1 belonging to clade 1 and 2 belonging to clade 2.3.4) and evaluated the effects of the I117V mutation on oseltamivir susceptibility in vitro, in vivo, and in silico. Results. The NA-I117V mutation conferred a slight reduction in susceptibility to oseltamivir in vitro (1.3- to 6.3-fold changes), although it did not substantially compromise NA enzymatic activity. Mice infected with I117V virus exhibited reduced susceptibility to oseltamivir and decreased survival in 2 of 3 virus pairs tested. Molecular dynamics simulations revealed that I117V caused the loss of hydrogen bonds between an arginine at position 118 and the carboxyl group of oseltamivir, leading to a lower binding affinity for oseltamivir. Conclusions. Our findings provide new insight into the mechanism of NA-I117V-mediated oseltamivir resistance in highly pathogenic H5N1 avian influenza viruses.
  • Masahiro Kajihara, Andrea Marzi, Eri Nakayama, Takeshi Noda, Makoto Kuroda, Rashid Manzoor, Keita Matsuno, Heinz Feldmann, Reiko Yoshida, Yoshihiro Kawaoka, Ayato Takada
    JOURNAL OF VIROLOGY 86 (24) 13467 - 13474 0022-538X 2012/12 [Refereed][Not invited]
     
    The envelope glycoprotein (GP) of Marburg virus (MARV) and Ebola virus (EBOV) is responsible for virus entry into host cells and is known as the only target of neutralizing antibodies. While knowledge about EBOV-neutralizing antibodies and the mechanism for the neutralization of infectivity is being accumulated gradually, little is known about antibodies that can efficiently regulate MARV infectivity. Here we show that MARV GP-specific monoclonal antibodies AGP127-8 (IgG1) and MGP72-17 (IgM), which do not inhibit the GP-mediated entry of MARV into host cells, drastically reduced the budding and release of progeny viruses from infected cells. These antibodies similarly inhibited the formation of virus-like particles (VLPs) consisting of GP, the viral matrix protein, and nucleoprotein, whereas the Fab fragment of AGP127-8 showed no inhibitory effect. Morphological analyses revealed that filamentous VLPs were bunched on the surface of VLP-producing cells cultured in the presence of the antibodies. These results demonstrate a novel mechanism of the antibody-mediated inhibition of MARV budding, in which antibodies arrest unformed virus particles on the cell surface. Our data lead to the idea that such antibodies, like classical neutralizing antibodies, contribute to protective immunity against MARV and that the "classical" neutralizing activity is not the only indicator of a protective antibody that may be available for prophylactic and therapeutic use.
  • Akihiro Ishii, Yuka Thomas, Ladslav Moonga, Ichiro Nakamura, Aiko Ohnuma, Bernard M. Hang'ombe, Ayato Takada, Aaron S. Mweene, Hirofumi Sawa
    JOURNAL OF GENERAL VIROLOGY 93 (10) 2247 - 2251 0022-1317 2012/10 [Refereed][Not invited]
     
    In order to survey arenaviruses in the Republic of Zambia, we captured 335 rodents from three cities between 2010 and 2011. Eighteen Luna virus (LUNV) and one lymphocytic choriomeningitis virus (LCMV)-related virus RNAs were detected by one-step RT-PCR from Mastomys natalensis and Mus minutoides, respectively. Four LUNV strains and one LCMV-related virus were isolated, and the whole genome nucleotide sequence was determined by pyrosequencing. Phylogenetic analyses revealed that the LUNV clade consists of two branches that are distinguished by geographical location and that the LCMV-related virus belongs to the LCMV clade, but diverges from the typical LCMVs. Comparison of nucleoprotein amino acid sequences indicated that the LCMV-related virus could be designated a novel arenavirus, which was tentatively named as the Lunk virus. Amino acid sequences of the GP, NP, Z and L proteins showed poor similarity among the three Zambian arenavirus strains, i.e. Luna, Lunk and Lujo virus.
  • Peter S. Lee, Reiko Yoshida, Damian C. Ekiert, Naoki Sakai, Yasuhiko Suzuki, Ayato Takada, Ian A. Wilson
    PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 109 (42) 17040 - 17045 0027-8424 2012/10 [Refereed][Not invited]
     
    Continual and rapid mutation of seasonal influenza viruses by antigenic drift necessitates the almost annual reformulation of flu vaccines, which may offer little protection if the match to the dominant circulating strain is poor. S139/1 is a cross-reactive antibody that neutralizes multiple HA strains and subtypes, including those from H1N1 and H3N2 viruses that currently infect humans. The crystal structure of the S139/1 Fab in complex with the HA from the A/Victoria/3/1975 (H3N2) virus reveals that the antibody targets highly conserved residues in the receptor binding site and contacts antigenic sites A, B, and D. Binding and plaque reduction assays show that the monovalent Fab alone can protect against H3 strains, but the enhanced avidity from binding of bivalent IgG increases the breadth of neutralization to additional strains from the H1, H2, H13, and H16 subtypes. Thus, antibodies making relatively low affinity Fab interactions with the receptor binding site can have significant antiviral activity when enhanced by avidity through bivalent interactions of the IgG, thereby extending the breadth of binding and neutralization to highly divergent influenza virus strains and subtypes.
  • Gary Wong, Jason S. Richardson, Stephane Pillet, Ami Patel, Xiangguo Qiu, Judie Alimonti, Jeff Hogan, Yi Zhang, Ayato Takada, Heinz Feldmann, Gary P. Kobinger
    SCIENCE TRANSLATIONAL MEDICINE 4 (158) 158ra146  1946-6234 2012/10 [Refereed][Not invited]
     
    Ebola virus causes severe hemorrhagic fever in susceptible hosts. Currently, no licensed vaccines or treatments are available; however, several experimental vaccines have been successful in protecting rodents and nonhuman primates (NHPs) from the lethal Zaire ebolavirus (ZEBOV) infection. The objective of this study was to evaluate immune responses correlating with survival in these animals after lethal challenge with ZEBOV. Knockout mice with impaired ability to generate normal T and/or B cell responses were vaccinated and challenged with ZEBOV. Vaccine-induced protection in mice was mainly mediated by B cells and CD4(+) T cells. Vaccinated, outbred guinea pigs and NHPs demonstrated the highest correlation between survival and levels of total immunoglobulin G (IgG) specific to the ZEBOV glycoprotein (ZGP). These results highlight the relevance of total ZGP-specific IgG levels as a meaningful correlate of protection against ZEBOV exposure.
  • Inhibitory effects of an M2-specific monoclonal antibody on different strains of influenza A virus
    Nilton Akio Muto, Reiko Yoshida, Tadaki Suzuki, Shintaro Kobayashi, Hiroichi Ozaki, Daisuke Fujikura, Rashid Manzoor, Mieko Muramatsu, Ayato Takada, Takashi Kimura, Hirofumi Sawa
    JAPANESE JOURNAL OF VETERINARY RESEARCH 60 (2-3) 71 - 83 0047-1917 2012/08 [Refereed][Not invited]
     
    New approaches to the treatment of influenza have been designed based on the highly conserved antigenicity of the M2 envelope protein among influenza A virus strains. The present study examined the anti-viral activities of an anti-M2 ectodomain (M2e) monoclonal antibody (clone rM2ss23), which binds to the M2 proteins of the influenza A virus strains A/Aichi/2/68 (H3N2) (Aichi) and A/PR/8/34 (H1N1) (PR8). The results showed that rM2ss23 bound to both Aichi and PR8 M2 proteins expressed on the cell surface. While the antibody did not prevent virus entry into cells, it significantly inhibited plaque formation by the Aichi strain in a dose-dependent manner when infected cells were cultured in the presence of the antibody. By contrast, the growth of PR8 (H1N1) was not affected by the antibody. A reverse genetics approach revealed that the inhibitory effect of rM2ss23 on the Aichi virus was abolished by replacing the genes encoding the HA and/or M proteins with those of the PR8 strain. These results suggest that rM2ss23 prevents virus release from infected cells and further suggest that the mechanisms underlying the virus budding mediates by HA and M2 proteins might differ between the Aichi and PR8 strains.
  • Chairul A. Nidom, Eri Nakayama, Reviany V. Nidom, Mohamad Y. Alamudi, Syafril Daulay, Indi N. L. P. Dharmayanti, Yoes P. Dachlan, Mohamad Amin, Manabu Igarashi, Hiroko Miyamoto, Reiko Yoshida, Ayato Takada
    PLOS ONE 7 (7) e40740  1932-6203 2012/07 [Refereed][Not invited]
     
    Ebola virus (EBOV) and Marburg virus (MARV) belong to the family Filoviridae and cause severe hemorrhagic fever in humans and nonhuman primates. Despite the discovery of EBOV (Reston virus) in nonhuman primates and domestic pigs in the Philippines and the serological evidence for its infection of humans and fruit bats, information on the reservoirs and potential amplifying hosts for filoviruses in Asia is lacking. In this study, serum samples collected from 353 healthy Bornean orangutans (Pongo pygmaeus) in Kalimantan Island, Indonesia, during the period from December 2005 to December 2006 were screened for filovirus-specific IgG antibodies using a highly sensitive enzyme-linked immunosorbent assay (ELISA) with recombinant viral surface glycoprotein (GP) antigens derived from multiple species of filoviruses (5 EBOV and 1 MARV species). Here we show that 18.4% (65/353) and 1.7% (6/353) of the samples were seropositive for EBOV and MARV, respectively, with little cross-reactivity among EBOV and MARV antigens. In these positive samples, IgG antibodies to viral internal proteins were also detected by immunoblotting. Interestingly, while the specificity for Reston virus, which has been recognized as an Asian filovirus, was the highest in only 1.4% (5/353) of the serum samples, the majority of EBOV-positive sera showed specificity to Zaire, Sudan, Cote d'Ivoire, or Bundibugyo viruses, all of which have been found so far only in Africa. These results suggest the existence of multiple species of filoviruses or unknown filovirus-related viruses in Indonesia, some of which are serologically similar to African EBOVs, and transmission of the viruses from yet unidentified reservoir hosts into the orangutan populations. Our findings point to the need for risk assessment and continued surveillance of filovirus infection of human and nonhuman primates, as well as wild and domestic animals, in Asia.
  • Masahiko Arikata, Yasushi Itoh, Masatoshi Okamatsu, Toshinaga Maeda, Takashi Shiina, Keiko Tanaka, Shingo Suzuki, Misako Nakayama, Yoshihiro Sakoda, Hirohito Ishigaki, Ayato Takada, Hideaki Ishida, Kosuke Soda, Van Loi Pham, Hideaki Tsuchiya, Shinichiro Nakamura, Ryuzo Torii, Takeshi Shimizu, Hidetoshi Inoko, Iwao Ohkubo, Hiroshi Kida, Kazumasa Ogasawara
    PLOS ONE 7 (5) e37220  1932-6203 2012/05 [Refereed][Not invited]
     
    We made an H1N1 vaccine candidate from a virus library consisting of 144 (= 16 HAx9 NA) non-pathogenic influenza A viruses and examined its protective effects against a pandemic (2009) H1N1 strain using immunologically naive cynomolgus macaques to exclude preexisting immunity and to employ a preclinical study since preexisting immunity in humans previously vaccinated or infected with influenza virus might make comparison of vaccine efficacy difficult. Furthermore, macaques carrying a major histocompatibility complex class I molecule, Mafa-A1*052:02, were used to analyze peptide-specific CD8(+) T cell responses. Sera of macaques immunized with an inactivated whole particle formulation without addition of an adjuvant showed higher neutralization titers against the vaccine strain A/Hokkaido/2/1981 (H1N1) than did sera of macaques immunized with a split formulation. Neutralization activities against the pandemic strain A/Narita/1/2009 (H1N1) in sera of macaques immunized twice with the split vaccine reached levels similar to those in sera of macaques immunized once with the whole particle vaccine. After inoculation with the pandemic virus, the virus was detected in nasal samples of unvaccinated macaques for 6 days after infection and for 2.67 days and 5.33 days on average in macaques vaccinated with the whole particle vaccine and the split vaccine, respectively. After the challenge infection, recall neutralizing antibody responses against the pandemic virus and CD8(+) T cell responses specific for nucleoprotein peptide NP262-270 bound to Mafa-A1*052:02 in macaques vaccinated with the whole particle vaccine were observed more promptly or more vigorously than those in macaques vaccinated with the split vaccine. These findings demonstrated that the vaccine derived from our virus library was effective for pandemic virus infection in macaques and that the whole particle vaccine conferred more effective memory and broader cross-reactive immune responses to macaques against pandemic influenza virus infection than did the split vaccine.
  • Andrea Marzi, Reiko Yoshida, Hiroko Miyamoto, Mari Ishijima, Yasuhiko Suzuki, Megumi Higuchi, Yukie Matsuyama, Manabu Igarashi, Eri Nakayama, Makoto Kuroda, Masayuki Saijo, Friederike Feldmann, Douglas Brining, Heinz Feldmann, Ayato Takada
    PLOS ONE 7 (4) e36192  1932-6203 2012/04 [Refereed][Not invited]
     
    Ebola virus (EBOV) is the causative agent of severe hemorrhagic fever in primates, with human case fatality rates up to 90%. Today, there is neither a licensed vaccine nor a treatment available for Ebola hemorrhagic fever (EHF). Single monoclonal antibodies (MAbs) specific for Zaire ebolavirus (ZEBOV) have been successfully used in passive immunization experiments in rodent models, but have failed to protect nonhuman primates from lethal disease. In this study, we used two clones of human-mouse chimeric MAbs (ch133 and ch226) with strong neutralizing activity against ZEBOV and evaluated their protective potential in a rhesus macaque model of EHF. Reduced viral loads and partial protection were observed in animals given MAbs ch133 and ch226 combined intravenously at 24 hours before and 24 and 72 hours after challenge. MAbs circulated in the blood of a surviving animal until virus-induced IgG responses were detected. In contrast, serum MAb concentrations decreased to undetectable levels at terminal stages of disease in animals that succumbed to infection, indicating substantial consumption of these antibodies due to virus replication. Accordingly, the rapid decrease of serum MAbs was clearly associated with increased viremia in non-survivors. Our results indicate that EBOV neutralizing antibodies, particularly in combination with other therapeutic strategies, might be beneficial in reducing viral loads and prolonging disease progression during EHF.
  • Yoshihiro Sakoda, Hiroshi Ito, Yuko Uchida, Masatoshi Okamatsu, Naoki Yamamoto, Kosuke Soda, Naoki Nomura, Saya Kuribayashi, Shintaro Shichinohe, Yuji Sunden, Takashi Umemura, Tatsufumi Usui, Hiroichi Ozaki, Tsuyoshi Yamaguchi, Toshiyuki Murase, Toshihiro Ito, Takehiko Saito, Ayato Takada, Hiroshi Kida
    JOURNAL OF GENERAL VIROLOGY 93 (3) 541 - 550 0022-1317 2012/03 [Refereed][Not invited]
     
    H5N1 highly pathogenic avian influenza virus (HPAIV) was reintroduced and caused outbreaks in chickens in the 2010-2011 winter season in Japan, which had been free from highly pathogenic avian influenza (HPAI) since 2007 when HPAI outbreaks occurred and were controlled. On 14 October 2010 at Lake Ohnuma, Wakkanai, the northernmost part of Hokkaido, Japan, H5N1 HPAIVs were isolated from faecal samples of ducks flying from their nesting lakes in Siberia. Since then, in Japan, H5N1 HPAIVs have been isolated from 63 wild birds in 17 prefectures and caused HPAI outbreaks in 24 chicken farms in nine prefectures by the end of March in 2011. Each of these isolates was genetically closely related to the HPAIV isolates at Lake Ohnuma, and those in China, Mongolia, Russia and Korea, belonging to genetic clade 2.3.2.1. In addition, these isolates were genetically classified into three groups, suggesting that the viruses were transmitted by migratory water birds through at least three different routes from their northern territory to Japan. These isolates were antigenic variants, which is consistent with selection in poultry under the immunological pressure induced by vaccination. To prevent the perpetuation of viruses in the lakes where water birds nest in summer in Siberia, prompt eradication of HPAIVs in poultry is urgently needed in Asian countries where HPAI has not been controlled.
  • Shinya Yamada, Kyoko Shinya, Ayato Takada, Toshihiro Ito, Takashi Suzuki, Yasuo Suzuki, Quynh Mai Le, Masahito Ebina, Noriyuki Kasai, Hiroshi Kida, Taisuke Horimoto, Pierre Rivailler, Li Mei Chen, Ruben O. Donis, Yoshihiro Kawaoka
    JOURNAL OF VIROLOGY 86 (3) 1411 - 1420 0022-538X 2012/02 [Refereed][Not invited]
     
    Quail are thought to serve as intermediate hosts of influenza A viruses between aquatic birds and terrestrial birds, such as chickens, due to their high susceptibility to aquatic-bird viruses, which then adapt to replicate efficiently in their new hosts. However, does replication of aquatic-bird influenza viruses in quail similarly result in their efficient replication in humans? Using sialic acid-galactose linkage-specific lectins, we found both avian (sialic acid-alpha 2-3-galactose [Sia alpha 2-3Gal] linkages on sialyloligosaccharides)- and human (Sia alpha 2-6Gal)-type receptors on the tracheal cells of quail, consistent with previous reports. We also passaged a duck H3N2 virus in quail 19 times. Sequence analysis revealed that eight mutations accumulated in hemagglutinin (HA) during these passages. Interestingly, many of the altered HA amino acids found in the adapted virus are present in human seasonal viruses, but not in duck viruses. We also found that stepwise stalk deletion of neuraminidase occurred during passages, resulting in reduced neuraminidase function. Despite some hemagglutinin mutations near the receptor binding pocket, appreciable changes in receptor specificity were not detected. However, reverse-genetics-generated viruses that possessed the hemagglutinin and neuraminidase of the quail-passaged virus replicated significantly better than the virus possessing the parent HA and neuraminidase in normal human bronchial epithelial cells, whereas no significant difference in replication between the two viruses was observed in duck cells. Further, the quail-passaged but not the original duck virus replicated in human bronchial epithelial cells. These data indicate that quail can serve as intermediate hosts for aquatic-bird influenza viruses to be transmitted to humans.
  • Kayoko Sato, Atsushi Iwai, Yosuke Nakayama, Junko Morimoto, Ayato Takada, Mitsuo Maruyama, Hiroshi Kida, Toshimitsu Uede, Tadaaki Miyazaki
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 417 (1) 274 - 279 0006-291X 2012/01 [Refereed][Not invited]
     
    Osteopontin (OPN) is involved in exacerbating various inflammatory diseases. A severe pulmonary inflammation is frequently found in lethal influenza A virus (IAV) infection. However, the function of OPN against the infection was poorly understood. Here, we demonstrate an importance of OPN on immune response and disease severity after IAV infection. We found that the expression level of OPN was increased in mice infected with IAV. The OPN knockout (KO) mice exhibited a severe pathological phenotype and the survival rate decreased after the lethal IAV infection, compared to the wild type mice, while the survival rate increased in OPN transgenic (Tg) mice. The population of natural killer (NK) cells significantly decreased in OPN KO mice at day 5 after the infection, whereas, it increased in OPN Tg mice. These results suggest that OPN plays an important role in host defense against IAV infection through the regulation of NK cell population. (C) 2011 Elsevier Inc. All rights reserved.
  • Ayato Takada
    FRONTIERS IN MICROBIOLOGY 3 34  1664-302X 2012 [Refereed][Invited]
     
    In human and non-human primates, filoviruses (Ebola and Marburg viruses) cause severe hemorrhagic fever. Recently, other animals such as pigs and some species of fruit bats have also been shown to be susceptible to these viruses. While having a preference for some cell types such as hepatocytes, endothelial cells, dendritic cells, monocytes, and macrophages, filoviruses are known to be pantropic in infection of primates. The envelope glycoprotein (GP) is responsible for both receptor binding and fusion of the virus envelope with the host cell membrane. It has been demonstrated that filovirus GP interacts with multiple molecules for entry into host cells, whereas none of the cellular molecules so far identified as a receptor/co-receptor fully explains filovirus tissue tropism and host range. Available data suggest that the mucin-like region (MLR) on GP plays an important role in attachment to the preferred target cells, whose infection is likely involved in filovirus pathogenesis, whereas the MLR is not essential for the fundamental function of the GP in viral entry into cells in vitro. Further studies elucidating the mechanisms of cellular entry of filoviruses may shed light on the development of strategies for prophylaxis and treatment of Ebola and Marburg hemorrhagic fevers.
  • Darryl Falzarano, Friederike Feldmann, Allen Grolla, Anders Leung, Hideki Ebihara, James E. Strong, Andrea Marzi, Ayato Takada, Shane Jones, Jason Gren, Joan Geisbert, Steven M. Jones, Thomas W. Geisbert, Heinz Feldmann
    JOURNAL OF INFECTIOUS DISEASES 204 S1082 - S1089 0022-1899 2011/11 [Refereed][Not invited]
     
    The recombinant vesicular stomatitis virus (rVSV) vector-based monovalent vaccine platform expressing a filovirus glycoprotein has been demonstrated to provide protection from lethal challenge with Ebola (EBOV) and Marburg (MARV) viruses both prophylactically and after exposure. This platform provides protection between heterologous strains within a species; however, protection from lethal challenge between species has been largely unsuccessful. To determine whether the rVSV-EBOV vaccines have the potential to provide protection against a newly emerging, phylogenetically related species, cynomolgus macaques were vaccinated with an rVSV vaccine expressing either the glycoprotein of Zaire ebolavirus (ZEBOV) or Cote d'Ivoire ebolavirus (CIEBOV) and then challenged with Bundibugyo ebolavirus (BEBOV), which was recently proposed as a new EBOV species following an outbreak in Uganda in 2007. A single vaccination with the ZEBOV-specific vaccine provided cross-protection (75% survival) against subsequent BEBOV challenge, whereas vaccination with the CIEBOV-specific vaccine resulted in an outcome similar to mock-immunized animals (33% and 25% survival, respectively). This demonstrates that monovalent rVSV-based vaccines may be useful against a newly emerging species; however, heterologous protection across species remains challenging and may depend on enhancing the immune responses either through booster immunizations or through the inclusion of multiple immunogens.
  • Eri Nakayama, Daisuke Tomabechi, Keita Matsuno, Noriko Kishida, Reiko Yoshida, Heinz Feldmann, Ayato Takada
    JOURNAL OF INFECTIOUS DISEASES 204 S978 - S985 0022-1899 2011/11 [Refereed][Not invited]
     
    Methods. To investigate this antibody-dependent enhancement (ADE) in MARV infection, we produced mouse antisera and monoclonal antibodies (mAbs) to the GPs of MARV strains Angola and Musoke. Results. The infectivity of vesicular stomatitis virus pseudotyped with Angola GP in K562 cells was significantly enhanced in the presence of Angola GP antisera, whereas only minimal ADE activity was seen with Musoke GP antisera. This difference correlated with the percentage of hybridoma clones producing infectivity-enhancing mAbs. Using mAbs to MARV GP, we identified 3 distinct ADE epitopes in the mucinlike region on Angola GP. Interestingly, some of these antibodies bound to both Angola and Musoke GPs but showed significantly higher ADE activity for strain Angola. ADE activity depended on epitopes in the mucinlike region and glycine at amino acid position 547, present in the Angola but absent in the Musoke GP. Conclusions. These results suggest a possible link between ADE and MARV pathogenicity and provide new insights into the mechanisms underlying ADE entry of filoviruses.
  • Akihiro Ishii, Yuka Thomas, Ladslav Moonga, Ichiro Nakamura, Aiko Ohnuma, Bernard Hang'ombe, Ayato Takada, Aaron Mweene, Hirofumi Sawa
    EMERGING INFECTIOUS DISEASES 17 (10) 1921 - 1924 1080-6040 2011/10 [Refereed][Not invited]
     
    To investigate arenavirus in Zambia, we characterized virus from the kidneys of 5 arenavirus RNA-positive rodents (Mastomys natalensis) among 263 captured. Full-genome sequences of the viruses suggested that they were new strains similar to Lassa virus-related arenaviruses. Analyzing samples from additional rodents and other species can elucidate epizootiologic aspects of arenaviruses.
  • Kimihito Ito, Manabu Igarashi, Yutaka Miyazaki, Teiji Murakami, Syaka Iida, Hiroshi Kida, Ayato Takada
    PLOS ONE 6 (10) e25953  1932-6203 2011/10 [Refereed][Not invited]
     
    Human influenza A viruses undergo antigenic changes with gradual accumulation of amino acid substitutions on the hemagglutinin (HA) molecule. A strong antigenic mismatch between vaccine and epidemic strains often requires the replacement of influenza vaccines worldwide. To establish a practical model enabling us to predict the future direction of the influenza virus evolution, relative distances of amino acid sequences among past epidemic strains were analyzed by multidimensional scaling (MDS). We found that human influenza viruses have evolved along a gnarled evolutionary pathway with an approximately constant curvature in the MDS-constructed 3D space. The gnarled pathway indicated that evolution on the trunk favored multiple substitutions at the same amino acid positions on HA. The constant curvature was reasonably explained by assuming that the rate of amino acid substitutions varied from one position to another according to a gamma distribution. Furthermore, we utilized the estimated parameters of the gamma distribution to predict the amino acid substitutions on HA in subsequent years. Retrospective prediction tests for 12 years from 1997 to 2009 showed that 70% of actual amino acid substitutions were correctly predicted, and that 45% of predicted amino acid substitutions have been actually observed. Although it remains unsolved how to predict the exact timing of antigenic changes, the present results suggest that our model may have the potential to recognize emerging epidemic strains.
  • Toru Ichihashi, Reiko Yoshida, Chihiro Sugimoto, Ayato Takada, Kiichi Kajino
    PLOS ONE 6 (9) e24626  1932-6203 2011/09 [Refereed][Not invited]
     
    Background: The virus-specific cytotoxic T lymphocyte (CTL) induction is an important target for the development of a broadly protective human influenza vaccine, since most CTL epitopes are found on internal viral proteins and relatively conserved. In this study, the possibility of developing a strain/subtype-independent human influenza vaccine was explored by taking a bioinformatics approach to establish an immunogenic HLA-A24 restricted CTL epitope screening system in HLA-transgenic mice. Methodology/Principal Findings: HLA-A24 restricted CTL epitope peptides derived from internal proteins of the H5N1 highly pathogenic avian influenza A virus were predicted by CTL epitope peptide prediction programs. Of 35 predicted peptides, six peptides exhibited remarkable cytotoxic activity in vivo. More than half of the mice which were subcutaneously vaccinated with the three most immunogenic and highly conserved epitopes among three different influenza A virus subtypes (H1N1, H3N2 and H5N1) survived lethal influenza virus challenge during both effector and memory CTL phases. Furthermore, mice that were intranasally vaccinated with these peptides remained free of clinical signs after lethal virus challenge during the effector phase. Conclusions/Significance: This CTL epitope peptide selection system can be used as an effective tool for the development of a cross-protective human influenza vaccine. Furthermore this vaccine strategy can be applicable to the development of all intracellular pathogens vaccines to induce epitope-specific CTL that effectively eliminate infected cells.
  • An H5N1 highly pathogenic avian influenza virus that invaded Japan through waterfowl migration
    Masahiro Kajihara, Keita Matsuno, Edgar Simulundu, Mieko Muramatsu, Osamu Noyori, Rashid Manzoor, Eri Nakayama, Manabu Igarashi, Daisuke Tomabechi, Reiko Yoshida, Masatoshi Okamatsu, Yoshihiro Sakoda, Kimihito Ito, Hiroshi Kida, Ayato Takada
    JAPANESE JOURNAL OF VETERINARY RESEARCH 59 (2-3) 89 - 100 0047-1917 2011/08 [Refereed][Not invited]
     
    In 2010, an H5N1 highly pathogenic avian influenza virus (HPAIV) was isolated from feces of apparently healthy ducks migrating southward in Hokkaido, the northernmost prefecture of Japan. The H5N1 HPAIVs were subsequently detected in domestic and wild birds at multiple sites corresponding to the flyway of the waterfowl having stopovers in the Japanese archipelago. The Hokkaido isolate was genetically nearly identical to H5N1 HPAIVs isolated from swans in the spring of 2009 and 2010 in Mongolia, but less pathogenic in experimentally infected ducks than the 2009 Mongolian isolate. These findings suggest that H5N1 HPAIVs with relatively mild pathogenicity might be selected and harbored in the waterfowl population during the 2009-2010 migration seasons. Our data provide "early warning" signals for preparedness against the unprecedented situation in which the waterfowl reservoirs serve as perpetual sources and disseminators of HPAIVs.
  • Edgar Simulundu, Akihiro Ishii, Manabu Igarashi, Aaron S. Mweene, Yuka Suzuki, Bernard M. Hang'ombe, Boniface Namangala, Ladslav Moonga, Rashid Manzoor, Kimihito Ito, Ichiro Nakamura, Hirofumi Sawa, Chihiro Sugimoto, Hiroshi Kida, Chuma Simukonda, Wilbroad Chansa, Jack Chulu, Ayato Takada
    JOURNAL OF GENERAL VIROLOGY 92 (6) 1416 - 1427 0022-1317 2011/06 [Refereed][Not invited]
     
    Although the quest to clarify the role of wild birds in the spread of the highly pathogenic H5N1 avian influenza virus (AIV) has yielded considerable data on AIVs in wild birds worldwide, information regarding the ecology and epidemiology of AIVs in African wild birds is still very limited. During AIV surveillance in Zambia (2008-2009), 12 viruses of distinct subtypes (H3N8, H4N6, H6N2, H9N1 and H11N9) were isolated from wild waterfowl. Phylogenetic analyses demonstrated that all the isolates were of the Eurasian lineage. Whilst some genes were closely related to those of AIVs isolated from wild and domestic birds in South Africa, intimating possible AIV exchange between wild birds and poultry in southern Africa, some gene segments were closely related to those of AIVs isolated in Europe and Asia, thus confirming the inter-regional AIV gene flow among these continents. Analysis of the deduced amino acid sequences of internal proteins revealed that several isolates harboured particular residues predominantly observed in human influenza viruses. Interestingly, the isolates with human-associated residues exhibited higher levels of virus replication in the lungs of infected mice and caused more morbidity as measured by weight loss than an isolate lacking such residues. This study stresses the need for continued monitoring of AIVs in wild and domestic birds in southern Africa to gain a better understanding of the emergence of strains with the potential to infect mammals.
  • Katsuaki Usami, Keita Matsuno, Manabu Igarashi, Kaori Denda-Nagai, Ayato Takada, Tatsuro Irimura
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 407 (1) 74 - 78 0006-291X 2011/04 [Refereed][Not invited]
     
    Ebola virus (EBOV) infection is initiated by the interaction of the viral surface envelope glycoprotein (GP) with the binding sites on target cells. Differences in the mortality among different species of the Ebola viruses, i.e., Zaire ebolavirus (ZEBOV) and Reston ebolavirus (REBOV), correspond to the in vitro infectivity of the pseudo-typed virus constructed with the GPs in cells expressing macrophage galactose-type calcium-type lectin (MGL/CD301). Through mutagenesis of GP2, the transmembrane-anchored subunit of GP, we found that residues 502-527 of the GP2 sequence determined the different infectivity between VSV-ZEBOV GP and -REBOV GP in MGL/CD301-expressing cells and a histidine residue at position 516 of ZEBOV GP2 appeared essential in the differential infectivity. These findings may provide a clue to clarify a molecular basis of different pathogenicity among EBOV species. (C) 2011 Elsevier Inc. All rights reserved.
  • Takuya Shiozaki, Atsushi Iwai, Yoshihiro Kawaoka, Ayato Takada, Hiroshi Kida, Tadaaki Miyazaki
    JOURNAL OF GENERAL VIROLOGY 92 (2) 315 - 325 0022-1317 2011/02 [Refereed][Not invited]
     
    Infection with influenza A virus causes acute respiratory tract infections in humans and may lead to lethal diseases including pneumonia. Identifying host factors that are involved in the severity of infectious diseases caused by influenza A virus is considered important for the prevention and treatment of these viral infections. This report demonstrated that Siva-1 is crucial for the induction of apoptosis caused by infection with influenza A virus and is involved in virus replication. Susceptibility to apoptosis induced by influenza A virus infection was increased in human lung-derived A549 cells, which stably express Siva-1. In addition, induction of apoptosis after influenza A virus infection was strongly inhibited by knockdown of Siva-1 expression. Furthermore, the replication of influenza A virus was significantly suppressed in A549 cells in which Siva-1 expression was inhibited and the effect of Siva-1 knockdown was eliminated by treatment with Z-VAD-FMK. These findings suggest that the caspase-dependent pathway for induction of apoptosis is involved in Siva-1-mediated influenza A virus replication.
  • A vaccine prepared from a non-pathogenic H5N1 influenza virus strain from the influenza virus library conferred protective immunity to chickens against the challenge with antigenically drifted highly pathogenic avian influenza virus
    Rozanah Asmah Abdul Samad, Naoki Nomura, Yoshimi Tsuda, Rashid Manzoor, Masahiro Kajihara, Daisuke Tomabechi, Takashi Sasaki, Norihide Kokumai, Toshiaki Ohgitani, Masatoshi Okamatsu, Ayato Takada, Yoshihiro Sakoda, Hiroshi Kida
    JAPANESE JOURNAL OF VETERINARY RESEARCH 59 (1) 23 - 29 0047-1917 2011/02 [Refereed][Not invited]
     
    Inactivated influenza virus vaccine prepared from a non-pathogenic influenza virus strain A/duck/Hokkaido/Vac-1/2004 (H5N1) from the virus library conferred protective immunity to chickens against the challenge of antigenically drifted highly pathogenic avian influenza virus (HPAIV), A/whooper swan/Hokkaido/1/2008 (H5N1). The efficacy of the vaccine was comparable to that prepared from genetically modified HPAIV strain Delta RRRRK rg-A/whooper swan/Mongolia/3/2005 (H5N1), which is more antigenically related to the challenge virus strain, in chickens.
  • Hirohito Ogawa, Hiroko Miyamoto, Hideki Ebihara, Kimihito Ito, Shigeru Morikawa, Heinz Feldmann, Ayato Takada
    JOURNAL OF VIROLOGICAL METHODS 171 (1) 310 - 313 0166-0934 2011/01 [Refereed][Not invited]
     
    The filoviruses, Marburg virus (MARV) and Ebola virus (EBOV), are causative agents of severe hemorrhagic fever with high mortality rates in humans and non-human primates. Sporadic outbreaks of filovirus infection have occurred in Central Africa and parts of Asia. Identification of the natural reservoir animals that are unknown yet and epidemiological investigations are current challenges to forestall outbreaks of filovirus diseases. The filovirus species identified currently include one in the MARV group and five in the EBOV group, with large genetic variations found among the species. Therefore, it has been difficult to develop a single sensitive assay to detect all filovirus species, which would advance laboratory diagnosis greatly in endemic areas. In this study, a highly sensitive universal RT-PCR assay targeting the nucleoprotein (NP) gene of filoviruses was developed. The genomic RNAs of all known MARV and EBOV species were detected by using an NP-specific primer set. In addition, this RT-PCR procedure was verified further for its application to detect viral RNAs in tissue samples of animals infected experimentally and blood specimens of infected patients. This assay will be a useful method for diagnostics and epidemiological studies of filovirus infections. (C) 2010 Elsevier B.V. All rights reserved.
  • Yuko Uchida, Katsushi Kanehira, Masaji Mase, Nobuhiro Takemae, Chiaki Watanabe, Tatsufumi Usui, Yoshikazu Fujimoto, Toshihiro Ito, Manabu Igarashi, Kimihito Ito, Ayato Takada, Yoshihiro Sakoda, Masatoshi Okamatsu, Yu Yamamoto, Kikuyasu Nakamura, Hiroshi Kida, Yasuaki Hiromoto, Tomoyuki Tsuda, Takehiko Saito
    VETERINARY MICROBIOLOGY 147 (1-2) 1 - 10 0378-1135 2011/01 [Refereed][Not invited]
     
    From February to March 2009, six strains of H7N6 subtype avian influenza virus were isolated from quails in three farms in Aichi prefecture in Japan. The isolates were shown to be low pathogenic for chicken by the examination performed using the "Manual of Standards for Diagnostic Tests and Vaccines" by World organisation for Animal Health (OIE). The deduced amino acid sequence at the cleavage site was PE (I/Q/L) PKRR (nucleotide sequences were cct gaa (a/c) (t/a) a cc (a/g) aaa aga aga), suggesting persistence in domestic poultry for some time. The direct putative ancestor strain could not be elucidated by phylogenetic analysis of all genome segments of the quail isolates. Diverged date from a putative common ancestor in a non-rooted phylogenetic tree among quail viruses was estimated between March 2002 and July 2004. Three putative N-linked glycosylation sites resided in the vicinity of the receptor binding pocket of HA1 region. They are considered to decrease the reactivity of neutralizing antibody against the virus. Experiments for the infectivity and pathogenicity of a quail strain to poultry indicated that the quail isolate had higher infectivity to quails than chickens and ducks. Direct and dust-borne and/or droplet-borne transmissions among quail were proven in quails with and without direct contact with experimentally infected quails. The virus is seldom transmitted among chickens either directly or indirectly, and indirect transmission from infected quails to chickens was not observed. The pathogenicity of the quail strain for mammalian, pig and mouse was low, although it could replicate in those animals. (C) 2010 Elsevier B.V. All rights reserved.
  • Masatoshi Okamatsu, Tomohisa Tanaka, Naoki Yamamoto, Yoshihiro Sakoda, Takashi Sasaki, Yoshimi Tsuda, Norikazu Isoda, Norihide Kokumai, Ayato Takada, Takashi Umemura, Hiroshi Kida
    VIRUS GENES 41 (3) 351 - 357 0920-8569 2010/12 [Refereed][Not invited]
     
    In April and May 2008, whooper swans (Cygnus cygnus) were found dead in Hokkaido in Japan. In this study, an adult whooper swan found dead beside Lake Saroma was pathologically examined and the identified H5N1 influenza virus isolates were genetically and antigenically analyzed. Pathological findings indicate that the swan died of severe congestive edema in the lungs. Phylogenetic analysis of the HA genes of the isolates revealed that they are the progeny viruses of isolates from poultry and wild birds in China, Russia, Korea, and Hong Kong. Antigenic analyses indicated that the viruses are distinguished from the H5N1 viruses isolated from wild birds and poultry before 2007. The chickens vaccinated with A/duck/Hokkaido/Vac-1/2004 (H5N1) survived for 14 days after challenge with A/whooper swan/Hokkaido/1/2008 (H5N1), although a small amount of the challenge virus was recovered from the tissues of the birds. These findings indicate that H5N1 highly pathogenic avian influenza viruses are circulating in wild birds in addition to domestic poultry in Asia and exhibit antigenic variation that may be due to vaccination.
  • Keita Matsuno, Eri Nakayama, Osamu Noyori, Andrea Marzi, Hideki Ebihara, Tatsuro Irimura, Heinz Feldmann, Ayato Takada
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 403 (1) 144 - 148 0006-291X 2010/12 [Refereed][Not invited]
     
    Cellular C-type lectins have been reported to facilitate filovirus infection by binding to glycans on filovirus glycoprotein (GP). However, it is not clearly known whether interaction between C-type lectins and GP mediates all the steps of virus entry (i.e., attachment, internalization, and membrane fusion). In this study, we generated vesicular stomatitis viruses pseudotyped with mutant GPs that have impaired structures of the putative receptor binding regions and thus reduced ability to infect the monkey kidney cells that are routinely used for virus propagation. We found that infectivities of viruses with the mutant GPs dropped in C-type lectin-expressing cells, parallel with those in the monkey kidney cells, whereas binding activities of these GPs to the C-type lectins were not correlated with the reduced infectivities. These results suggest that C-type lectin-mediated entry of filoviruses requires other cellular molecule(s) that may be involved in virion internalization or membrane fusion. (C) 2010 Elsevier Inc. All rights reserved.
  • Eri Nakayama, Ayaka Yokoyama, Hiroko Miyamoto, Manabu Igarashi, Noriko Kishida, Keita Matsuno, Andrea Marzi, Heinz Feldmann, Kimihito Ito, Masayuki Saijo, Ayato Takada
    CLINICAL AND VACCINE IMMUNOLOGY 17 (11) 1723 - 1728 1556-6811 2010/11 [Refereed][Not invited]
     
    Several enzyme-linked immunosorbent assays (ELISAs) for the detection of filovirus-specific antibodies have been developed. However, diagnostic methods to distinguish antibodies specific to the respective species of filoviruses, which provide the basis for serological classification, are not readily available. We established an ELISA using His-tagged secreted forms of the transmembrane glycoproteins (GPs) of five different Ebola virus (EBOV) species and one Marburg virus (MARV) strain as antigens for the detection of filovirus species-specific antibodies. The GP-based ELISA was evaluated by testing antisera collected from mice immunized with virus-like particles as well as from humans and nonhuman primates infected with EBOV or MARV. In our ELISA, little cross-reactivity of IgG antibodies was observed in most of the mouse antisera. Although sera and plasma from some patients and monkeys showed notable cross-reactivity with the GPs from multiple filovirus species, the highest reactions of IgG were uniformly detected against the GP antigen homologous to the virus species that infected individuals. We further confirmed that MARV-specific IgM antibodies were specifically detected in specimens collected from patients during the acute phase of infection. These results demonstrate the usefulness of our ELISA for diagnostics as well as ecological and serosurvey studies.
  • Yoshihiro Sakoda, Sengee Sugar, Damdinjav Batchluun, Tseren-Ochir Erdene-Ochir, Masatoshi Okamatsu, Norikazu Isoda, Kosuke Soda, Hiroki Takakuwa, Yoshimi Tsuda, Naoki Yamamoto, Noriko Kishida, Keita Matsuno, Eri Nakayama, Masahiro Kajihara, Ayaka Yokoyama, Ayato Takada, Ruuragchaa Sodnomdarjaa, Hiroshi Kida
    VIROLOGY 406 (1) 88 - 94 0042-6822 2010/10 [Refereed][Not invited]
     
    H5N1 highly pathogenic avian influenza (HPAI) viruses were isolated from dead wild waterfowl at Khunt, Erkhel, Doityn Tsagaan, Doroo, and Ganga Lakes in Mongolia in July 2005, May 2006, May 2009, July 2009, and May 2010, respectively. The isolates in 2005 and 2006 were classified into genetic clade 2.2, and those in 2009 and 2010 into clade 2.3.2. A/whooper swan/Mongolia/6/2009 (H5N1) experimentally infected ducks and replicated systemically with higher mortality than that of the isolates in 2005 and 2006. Intensive surveillance of avian influenza in migratory waterfowl flying from their nesting lakes in Siberia to Mongolia in every autumn indicate that HPAI viruses have not perpetuated at their nesting lakes until 2009. The present results demonstrate that wild waterfowl were sporadically infected with H5N1 HPAI viruses prevailing in domestic poultry in the southern Asia and died in Mongolia on the way back to their northern territory in spring. (C) 2010 Elsevier Inc. All rights reserved.
  • JiuPian Yang, Reiko Yoshida, Yuki Kariya, Xu Zhang, Shuhei Hashiguchi, Toshihiro Nakashima, Yasuo Suda, Ayato Takada, Yuji Ito, Kazuhisa Sugimura
    JOURNAL OF BIOCHEMISTRY 148 (4) 507 - 515 0021-924X 2010/10 [Refereed][Not invited]
     
    The development of new therapeutic targets and strategies to control highly pathogenic avian influenza (HPAI) H5N1 virus infection in humans is urgently needed. Neutralizing recombinant human antibodies would provide important agents for immunotherapy on human H5N1 virus infection and definition of the critical mimotope for vaccine development. In this study, we have characterized an anti-H5-specific scFv clone, 3D1 from the human-scFv-displaying phage library. 3D1 blocked the binding of H5-Fc to MDCK cells in flow cytometry and neutralized H5N1 subtype influenza A viruses in a microneutralization assay. Employing a peptide-displaying phage library, Ph.D-12, the mimotope was determined to be at #128-131 and #204-211 of H5, which are silic acid-binding regions. In consistency with this result, 3D1 binds the recombinant sugar-binding domain (#50G-#272E) produced by a baculovirus vector. The 3D1 antibody employs the germline gene VH1-23. As this antibody is the first human anti-H5 scFv clearly defined on the sugar-binding epitope, it allows us to investigate the influence of amino acid substitutions in this region on the determination of the binding specificity to either sialic acid alpha 2,6-galactose (SA alpha 2,6Gal) or sialic acid alpha 2,3-galactose (SA alpha 2,3Gal) providing new insight for the development of effective H5N1 pandemic vaccines.
  • Atsushi Iwai, Takuya Shiozaki, Taro Kawai, Shizuo Akira, Yoshihiro Kawaoka, Ayato Takada, Hiroshi Kida, Tadaaki Miyazaki
    JOURNAL OF BIOLOGICAL CHEMISTRY 285 (42) 32064 - 32074 0021-9258 2010/10 [Refereed][Not invited]
     
    Type I interferons (IFNs) are known to be critical factors in the activation of host antiviral responses and are also important in protection from influenza A virus infection. Especially, the RIG-I- and IPS-1-mediated intracellular type I IFN-inducing pathway is essential in the activation of antiviral responses in cells infected by influenza A virus. Previously, it has been reported that influenza A virus NS1 is involved in the inhibition of this pathway. We show in this report that the influenza A virus utilizes another critical inhibitory mechanism in this pathway. In fact, the viral polymerase complex exhibited an inhibitory activity on IFN beta promoter activation mediated by RIG-I and IPS-1, and this activity was not competitive with the function of NS1. Co-immunoprecipitation analysis revealed that each polymerase subunit bound to IPS-1 in mammalian cells, and each subunit inhibited the activation of IFN beta promoter by IPS-1 independently. In addition, by a combinational expression of each polymerase subunit, IPS-1-induced activation of IFN beta promoter was more efficiently inhibited by the expression of PB2 or PB2-containing complex. Moreover, the expression of PB2 inhibited the transcription of the endogenous IFN beta gene induced after influenza A virus infection. These findings demonstrate that the viral polymerase plays an important role for regulating host anti-viral response through the binding to IPS-1 and inhibition of IFN beta production.
  • Keita Matsuno, Noriko Kishida, Katsuaki Usami, Manabu Igarashi, Reiko Yoshida, Eri Nakayama, Masayuki Shimojima, Heinz Feldmann, Tatsuro Irimura, Yoshihiro Kawaoka, Ayato Takada
    JOURNAL OF VIROLOGY 84 (10) 5140 - 5147 0022-538X 2010/05 [Refereed][Not invited]
     
    The glycoproteins (GPs) of filoviruses are responsible for virus entry into cells. It is known that GP interacts with cellular C-type lectins for virus attachment to cells. Since primary target cells of filoviruses express C-type lectins, C-type lectin-mediated entry is thought to be a possible determinant of virus tropism and pathogenesis. We compared the efficiency of C-type lectin-mediated entry between Marburg virus strains Angola and Musoke by using a vesicular stomatitis virus (VSV) pseudotype system. VSV pseudotyped with Angola GP (VSV-Angola) infected K562 cells expressing the C-type lectin, human macrophage galactose-type C-type lectin (hMGL), or dendritic cell-specific ICAM-3-grabbing nonintegrin (DC-SIGN) more efficiently than VSV pseudotyped with Musoke GP (VSV-Musoke). Unexpectedly, the binding affinity of the C-type lectins to the carbohydrates on GPs did not correlate with the different efficiency of C-type lectin-mediated entry. Site-directed mutagenesis identified the amino acid at position 547, which switched the efficiency of C-type lectin-mediated entry. In a three-dimensional model of GP, this amino acid was in close proximity to the putative site of cathepsin processing. Interestingly, the cathepsin inhibitors reduced the infectivity of VSV-Angola less efficiently than that of VSV-Musoke in C-type lectin-expressing K562 cells, whereas only a limited difference was found in control cells. The amino acid at position 547 was critical for the different effects of the inhibitors on the virus infectivities. These results suggest that the efficiency of C-type lectin-mediated entry of filoviruses is controlled not only by binding affinity between C-type lectins and GP but also by mechanisms underlying endosomal entry, such as proteolytic processing by the cathepsins.
  • Manabu Igarashi, Kimihito Ito, Reiko Yoshida, Daisuke Tomabechi, Hiroshi Kida, Ayato Takada
    PLOS ONE 5 (1) e8553  1932-6203 2010/01 [Refereed][Not invited]
     
    The pandemic influenza virus (2009 H1N1) was recently introduced into the human population. The hemagglutinin (HA) gene of 2009 H1N1 is derived from "classical swine H1N1'' virus, which likely shares a common ancestor with the human H1N1 virus that caused the pandemic in 1918, whose descendant viruses are still circulating in the human population with highly altered antigenicity of HA. However, information on the structural basis to compare the HA antigenicity among 2009 H1N1, the 1918 pandemic, and seasonal human H1N1 viruses has been lacking. By homology modeling of the HA structure, here we show that HAs of 2009 H1N1 and the 1918 pandemic virus share a significant number of amino acid residues in known antigenic sites, suggesting the existence of common epitopes for neutralizing antibodies cross-reactive to both HAs. It was noted that the early human H1N1 viruses isolated in the 1930s-1940s still harbored some of the original epitopes that are also found in 2009 H1N1. Interestingly, while 2009 H1N1 HA lacks the multiple N-glycosylations that have been found to be associated with an antigenic change of the human H1N1 virus during the early epidemic of this virus, 2009 H1N1 HA still retains unique three-codon motifs, some of which became N-glycosylation sites via a single nucleotide mutation in the human H1N1 virus. We thus hypothesize that the 2009 H1N1 HA antigenic sites involving the conserved amino acids will soon be targeted by antibody-mediated selection pressure in humans. Indeed, amino acid substitutions predicted here are occurring in the recent 2009 H1N1 variants. The present study suggests that antibodies elicited by natural infection with the 1918 pandemic or its early descendant viruses play a role in specific immunity against 2009 H1N1, and provides an insight into future likely antigenic changes in the evolutionary process of 2009 H1N1 in the human population.
  • Edgar Simulundu, Aaron S. Mweene, Daisuke Tomabechi, Bernard M. Hang'ombe, Akihiro Ishii, Yuka Suzuki, Ichiro Nakamura, Hirofumi Sawa, Chihiro Sugimoto, Kimihito Ito, Hiroshi Kida, Lewis Saiwana, Ayato Takada
    ARCHIVES OF VIROLOGY 154 (9) 1517 - 1522 0304-8608 2009/09 [Refereed][Not invited]
     
    We characterized an influenza virus isolated from a great white pelican in Zambia. Phylogenetic analysis showed that all of its gene segments belonged to the Eurasian lineage and that they appear to have evolved in distinct geographical regions in Europe, Asia, and Africa, suggesting reassortment of virus genes maintained in wild aquatic birds whose flyways overlap across these continents. It is notable that this virus might possess some genes of the same origin as those of highly pathogenic H7 and H5 viruses isolated in Eurasia. The present study underscores the need for continued monitoring of avian influenza viruses in Eurasia and Africa.
  • Reiko Yoshida, Manabu Igarashi, Hiroichi Ozaki, Noriko Kishida, Daisuke Tomabechi, Hiroshi Kida, Kimihito Ito, Ayato Takada
    PLOS PATHOGENS 5 (3) e1000350  1553-7366 2009/03 [Refereed][Not invited]
     
    The hemagglutinin (HA) of influenza A viruses has been classified into sixteen distinct subtypes (H1-H16) to date. The HA subtypes of influenza A viruses are principally defined as serotypes determined by neutralization or hemagglutination inhibition tests using polyclonal antisera to the respective HA subtypes, which have little cross-reactivity to the other HA subtypes. Thus, it is generally believed that the neutralizing antibodies are not broadly cross-reactive among HA subtypes. In this study, we generated a novel monoclonal antibody (MAb) specific to HA, designated MAb S139/1, which showed heterosubtypic cross-reactive neutralization and hemagglutination inhibition of influenza A viruses. This MAb was found to have broad reactivity to many other viruses (H1, H2, H3, H5, H9, and H13 subtypes) in enzyme-linked immunosorbent assays. We further found that MAb S139/1 showed neutralization and hemagglutination-inhibition activities against particular strains of H1, H2, H3, and H13 subtypes of influenza A viruses. Mutant viruses that escaped neutralization by MAb S139/1 were selected from the A/Aichi/2/68 (H3N2), A/Adachi/2/57 (H2N2), and A/WSN/33 (H1N1) strains, and sequence analysis of the HA genes of these escape mutants revealed amino acid substitutions at positions 156, 158, and 193 (H3 numbering). A molecular modeling study showed that these amino acids were located on the globular head of the HA and formed a novel conformational epitope adjacent to the receptor-binding domain of HA. Furthermore, passive immunization of mice with MAb S139/1 provided heterosubtypic protection. These results demonstrate that MAb S139/1 binds to a common antigenic site shared among a variety of HA subtypes and neutralizes viral infectivity in vitro and in vivo by affecting viral attachment to cells. The present study supports the notion that cross-reactive antibodies play some roles in heterosubtypic immunity against influenza A virus infection, and underscores the potential therapeutic utility of cross-reactive antibodies against influenza.
  • Shin Murakami, Ayaka Iwasa, Kiyoko Iwatsuki-Horimoto, Mutsumi Ito, Maki Kiso, Hiroshi Kida, Ayato Takada, Chairul A. Nidom, Le Quynh Mai, Shinya Yamada, Hirotaka Imai, Yuko Sakai-Tagawa, Yoshihiro Kawaoka, Taisuke Horimoto
    VACCINE 26 (50) 6398 - 6404 0264-410X 2008/11 [Refereed][Not invited]
     
    H5N1 highly pathogenic avian influenza viruses evolved into several clades, leading to appreciably distinct antigenicities of their hemagglutinins. As such, candidate H5N1 pre-pandemic vaccines for human use should be sought. Here, to evaluate fundamental immunogenic variations between H5N1 vaccines, we prepared four inactivated H5N1 test vaccines from different phylogenetic Glades (clade 1, 2.1, 2.2, and 2.3.4) in accordance with the WHO recommendation, and tested their cross-Glade immunity in a mouse model by vaccination followed by challenge with heterologous virulent viruses. All H5N1 vaccines tested provided full or partial cross-clade protective immunity, except one Glade 2.2-based vaccine, which did not protect mice from clade 2.3.4 virus challenge. Among the test vaccines, a clade 2.1-based vaccine possessed the broadest-spectrum cross-immunity. These results suggest that currently stockpiled pre-pandemic vaccines, especially clade 2.1-based vaccines, will likely be useful as backup vaccines in a pandemic situation, even one involving antigenic-drifted viruses. (c) 2008 Elsevier Ltd. All rights reserved.
  • Kosuke Soda, Hiroichi Ozaki, Yoshihiro Sakoda, Norikazu Isoda, Yoshinari Haraguchi, Saori Sakabe, Noritaka Kuboki, Noriko Kishida, Ayato Takada, Hiroshi Kida
    ARCHIVES OF VIROLOGY 153 (11) 2041 - 2048 0304-8608 2008/11 [Refereed][Not invited]
     
    In order to prepare H5N1 influenza virus vaccine, the hemagglutinins (HAs) of 14 H5 virus isolates from water birds in Asia were antigenically and genetically analyzed. Phylogenetic analysis of the H5 HA genes revealed that 13 isolates belong to Eurasian and the other one to North American lineages. Each of the deduced amino acid sequences of the HAs indicated a non-pathogenic profile. Antigenic analysis using a panel of monoclonal antibodies recognizing six different epitopes on the HA of A/duck/Pennsylvania/10218/1984 (H5N2) and chicken antiserum to an H5N1 reassortant strain generated between A/duck/Mongolia/54/2001 (H5N2) and A/duck/Mongolia/47/2001 (H7N1), [R(Dk/Mong-Dk/Mong) (H5N1)] showed that the HAs of highly pathogenic avian influenza (HPAI) viruses currently circulating in Asia were antigenically closely related to those of the present isolates from water birds. Mice subcutaneously injected with formalin-inactivated R(Dk/Mong-Dk/Mong) were protected from challenge with 100 mouse lethal dose of A/Viet Nam/1194/2004 (H5N1). The present results support the notion that the H5 isolates and the reassortant H5N1 strain should be useful for vaccine preparation.
  • Noriko Kishida, Yoshihiro Sakoda, Mai Shiromoto, Gui-Rong Bai, Norikazu Isoda, Ayato Takada, Graeme Laver, Hiroshi Kida
    VIRUS GENES 37 (1) 16 - 21 0920-8569 2008/08 [Refereed][Not invited]
     
    To investigate the prevalence of influenza viruses in feral water birds in the Southern Hemisphere, fecal samples of terns were collected on Heron Island, Australia, in December 2004. Six H2N5 influenza viruses were isolated. This is the first report of the isolation of the H2 subtype from shore birds in Australia. Phylogenetic analysis revealed that the M gene belonged to the American lineage of avian influenza viruses and the other genes belonged to the Eurasian lineages, indicating that genetic reassortment occurs between viruses of Eurasian and American lineages in free flying birds in nature.
  • Manabu Igarashi, Kimihito Ito, Hiroshi Kida, Ayato Takada
    VIROLOGY 376 (2) 323 - 329 0042-6822 2008/07 [Refereed][Not invited]
     
    The addition of oligosaccharide side chains to influenza virus hemagglutinin (HA) is believed to facilitate viral escape from immune pressure in the human population. To determine the implicit potentials for acquisition of N-linked glycosylation, we analyzed the genetic background of 16 subtypes of avian influenza virus, some of which may be potential pandemic viruses in the future. We found a significant difference among HA subtypes in their genomic sequences to produce N-glycosylation sites. Notably, recently circulating avian influenza viruses of the H5 and H9 subtypes may have rather greater capacities to undergo mutations associated with glycosylation of HA than past pandemic viruses. We hypothesize that influenza viruses maintained in natural reservoirs could have different potentials for sustained circulation, depending on their HA subtypes, if introduced into the human population. (C) 2008 Elsevier Inc. All rights reserved.
  • Ayato Takada, Hideki Ebihara, Heinz Feldmann, Thomas W. Geisbert, Yoshihiro Kawaoka
    JOURNAL OF INFECTIOUS DISEASES 196 S347 - S356 0022-1899 2007/11 [Refereed][Not invited]
     
    We have shown that antibody-dependent enhancement (ADE) of infection with Zaire Ebola virus (ZEBOV) is mediated by interaction of virus-specific antibodies with Fc receptors or complement component C1q and its receptors in vitro. ADE activities of the antisera to the viral glycoprotein (GP) were virus species specific and were primarily correlated with immunoglobulin (Ig) G2a and IgM levels but not with IgG1 levels. Interestingly, compared with ZEBOV, Reston Ebola virus (REBOV) had substantially weaker potential to induce ADE antibodies. Using monoclonal antibodies, we identified ZEBOV-specific ADE epitopes. To confirm epitope specificity, we constructed a chimeric ZEBOV GP, the ADE epitopes of which were replaced with the corresponding regions of REBOV GP. We found that mouse antisera to the chimeric ZEBOV GP showed less potential to induce ADE activity than did mouse antisera to wild-type ZEBOV GP, although they retained neutralizing activity. These data suggest that GP lacking the ADE-inducing epitopes may increase the potential of GP as a vaccine antigen.
  • Yohei Kurosaki, Ayato Takada, Hideki Ebihara, Allen Grolla, Naoki Kamo, Heinz Feldmann, Yoshihiro Kawacka, Jiro Yasuda
    JOURNAL OF VIROLOGICAL METHODS 141 (1) 78 - 83 0166-0934 2007/04 [Refereed][Not invited]
     
    Ebola virus (EBOV) causes severe hemorrhagic fever in humans and nonhuman primates with high mortality rates. Rapid identification of the virus is required to prevent spread of the infection. In this study, we developed and evaluated a one-step simple reverse transcription-loop mediated isothermal amplification (RT-LAMP) assay for the rapid detection of Zaire ebolavirus (ZEBOV), the most virulent species of EBOV, targeting the trailer region of the viral genome. The assay could detect 20 copies of the artificial ZEBOV RNA in 26 min with a real time-monitoring detection, and also detect 10(-3) FFU of the cell-culture propagated viruses. The reaction time needed to detect 104 FFU of ZEBOV was only 20 min. In addition, the assay was highly specific for ZEBOV. The RT-LAMP assay developed in this study is rapid, simple, highly specific, and sensitive for the detection of ZEBOV, and so may be an effective diagnostic tool. Furthermore, as this technique does not require sophisticated instrumentation, it seems very suitable for diagnosis in the field or laboratories in Ebola outbreak areas such as Central Africa. (c) 2006 Elsevier B.V. All rights reserved.
  • Makoto Ozawa, Ken Fujii, Yukiko Muramoto, Shinya Yamada, Seiya Yamayoshi, Ayato Takada, Hideo Goto, Taisuke Horimoto, Yoshihiro Kawaoka
    JOURNAL OF VIROLOGY 81 (1) 30 - 41 0022-538X 2007/01 [Refereed][Not invited]
     
    The RNA genome of influenza A virus, which forms viral ribonucleoprotein complexes (vRNPs) with viral polymerase subunit proteins (PA, PB1, and PB2) and nucleoprotein (NP), is transcribed and replicated in the nucleus. NP, the major component of vRNPs, has at least two amino acid sequences that serve as nuclear localization signals (NLSs): an unconventional NLS (residues 3 to 13; NLS1) and a bipartite NLS (residues 198 to 216; NLS2). Although both NLSs are known to play a role in nuclear transport, their relative contributions to viral replication are poorly understood. We therefore investigated their contributions to NP subcellular/sulmuclear localization, viral RNA (vRNA) transcription, and viral replication. Abolishing the unconventional NLS caused NP to localize predominantly to the cytoplasm and affected its activity in vRNA transcription. However, we were able to create a virus whose NP contained amino acid substitutions in NLS1 known to abolish its nuclear localization function, although this virus was highly attenuated. These results indicate that while the unconventional NLS is not essential for viral replication, it is necessary for efficient viral mRNA synthesis. On the other hand, the bipartite NLS, whose contribution to the nuclear transport of NP is limited, was essential for vRNA transcription and NP's nucleolar accumulation. A virus with nonfunctional NLS2 could not be generated. Thus, the bipartite NLS, but not the unconventional NLS, of NP is essential for influenza A virus replication.
  • Ayato Takada, Hideki Ebihara, Sueven Jones, Heinz Feldmann, Yoshihiro Kawaoka
    VACCINE 25 (6) 993 - 999 0264-410X 2007/01 [Refereed][Not invited]
     
    Ebola virus causes lethal hemorrhagic fever in humans and nonhuman primates, but no effective antiviral compounds are available for the treatment of this infection. The surface glycoprotein (GP) of Ebola virus is an important target of neutralizing antibodies. Although passive transfer of GP-specific antibodies has been evaluated in mouse and guinea. pig models, protection was achieved only by treatment shortly before or after virus challenge. Using these animal models, we evaluated the protective efficacy of two monoclonal antibodies whose epitopes are distinct from those of the antibodies tested by others. Treatment of mice with these antibodies 2 days after challenge completely protected most of the animals; even treatment 3 or 4 days after challenge was partially effective. Although antibody treatment in the guinea pig model was not as effective as in the mouse model, single-dose treatment of guinea pigs I day before, or I or 2 days after challenge did protect some animals. Interestingly, the protective effects seen in these animal models did not correlate with the in vitro neutralizing activity of the antibodies, suggesting different mechanisms of the neutralization by these antibodies. These results underscore the potential therapeutic utility of monoclonal antibodies for postexposure treatment of Ebola virus infections. (c) 2006 Elsevier Ltd. All rights reserved.
  • Masayuki Shimojima, Ayato Takada, Hideki Ebihara, Gabriele Neumann, Kouki Fujioka, Tatsuro Irimura, Steven Jones, Heinz Feldmann, Yoshihiro Kawaoka
    JOURNAL OF VIROLOGY 80 (20) 10109 - 10116 0022-538X 2006/10 [Refereed][Not invited]
     
    Filoviruses, represented by the genera Ebolavirus and Marburgvirus, cause a lethal hemorrhagic fever in humans and in nonhuman primates. Although filovirus can replicate in various tissues or cell types in these animals, the molecular mechanisms of its broad tropism remain poorly understood. Here we show the involvement of members of the Tyro3 receptor tyrosine kinase family - Axl, Dtk, and Mer-in cell entry of filoviruses. Ectopic expression of these family members in lymphoid cells, which otherwise are highly resistant to filovirus infection, enhanced infection by pseudotype viruses carrying filovirus glycoproteins on their envelopes. This enhancement was reduced by antibodies to Tyro3 family members, Gas6 ligand, or soluble ectodomains of the members. Live Ebola viruses infected both Axl- and Dtk-expressing cells more efficiently than control cells. Antibody to Axl inhibited infection of pseudotype viruses in a number of Axl-positive cell lines. These results implicate each Tyro3 family member as a cell entry factor in filovirus infection.
  • Takeshi Noda, Hideki Ebihara, Yukiko Muramoto, Ken Fujii, Ayato Takada, Hiroshi Sagara, Jin Hyun Kim, Hiroshi Kida, Heinz Feldmann, Yoshihiro Kawaoka
    PLOS PATHOGENS 2 (9) 864 - 872 1553-7366 2006/09 [Refereed][Not invited]
     
    Ebolavirus is responsible for highly lethal hemorrhagic fever. Like all viruses, it must reproduce its various components and assemble them in cells in order to reproduce infectious virions and perpetuate itself. To generate infectious Ebolavirus, a viral genome-protein complex called the nucleocapsid (NC) must be produced and transported to the cell surface, incorporated into virions, and then released from cells. To further our understanding of the Ebolavirus life cycle, we expressed the various viral proteins in mammalian cells and examined them ultrastructurally and biochemically. Expression of nucleoprotein alone led to the formation of helical tubes, which likely serve as a core for the NC. The matrix protein VP40 was found to be critical for transport of NCs to the cell surface and for the incorporation of NCs into virions, where interaction between nucleoprotein and the matrix protein VP40 is likely essential for these processes. Examination of virus-infected cells revealed that virions containing NCs mainly emerge horizontally from the cell surface, whereas empty virions mainly bud vertically, suggesting that horizontal budding is the major mode of Ebolavirus budding. These data form a foundation for the identification and development of potential antiviral agents to combat the devastating disease caused by this virus.
  • Hideki Ebihara, Ayato Takada, Darwyn Kobasa, Steven Jones, Gabriele Neumann, Steven Theriault, Mike Bray, Heinz Feldmann, Yoshihiro Kawaoka
    PLOS PATHOGENS 2 (7) 705 - 711 1553-7366 2006/07 [Refereed][Not invited]
     
    Zaire ebolavirus (ZEBOV) causes severe hemorrhagic fever in humans and nonhuman primates, with fatality rates in humans of up to 90%. The molecular basis for the extreme virulence of ZEBOV remains elusive. While adult mice resist ZEBOV infection, the Mayinga strain of the virus has been adapted to cause lethal infection in these animals. To understand the pathogenesis underlying the extreme virulence of Ebola virus ( EBOV), here we identified the mutations responsible for the acquisition of the high virulence of the adapted Mayinga strain in mice, by using reverse genetics. We found that mutations in viral protein 24 and in the nucleoprotein were primarily responsible for the acquisition of high virulence. Moreover, the role of these proteins in virulence correlated with their ability to evade type I interferon-stimulated antiviral responses. These findings suggest a critical role for overcoming the interferon-induced antiviral state in the pathogenicity of EBOV and offer new insights into the pathogenesis of EBOV infection.
  • Hetron M. Munang'andu, Victor M. Siamudaala, Andrew Nambota, John M. Bwalya, Musso Munyeme, Aaron S. Mweene, Ayato Takada, Hiroshi Kida
    JAPANESE JOURNAL OF VETERINARY RESEARCH 54 (1) 3 - 13 0047-1917 2006/05 [Refereed][Not invited]
     
    Eco-tourism depending on wildlife is becoming increasingly profitable and landowners are beginning to favor game farming and ecotourism. In these areas, large-scale translocation of wildlife involves a diversity of species and large populations. The African buffalo (Syncerus caffer) is one of the major tourist attractions in Zambia. It accounts for 8.7% and 12.4% of the total animal species hunted in the Game Management Areas and the total hunting revenue earned in Zambia, respectively. It is ecologically an important animal species essential for the purpose of habitat control and facilitating the provision of suitable grazing pastures. However, the rearing of the African buffalo on game ranches has been hampered by its carrier state of the Southern Africa Terroritory (SAT) serotypes of foot and mouth disease virus (FMD). The African buffalo is also known to be a carrier of Theileria parva lawrencei, the causative agent of corridor disease (CD) that continues to have devastating effects on the livestock industry in Zambia. In addition, the importation of buffaloes from countries with populations endemic to bovine tuberculosis is highly restricted. Veterinary regulations in Zambia, strongly advocate against the translocation of buffaloes from protected areas to private ranches for disease control purposes thereby mounting a considerable constraint on the economic and ecological viability of the industry. It is hoped that this review will motivate the relevant government authorities in exploiting ways in which this animal species play a central role in eco-tourism.
  • Ecology and epidemiology of anthrax in cattle and humans in Zambia
    Victor M. Siamudaala, John M. Bwalya, Hetron M. Munang'andu, Peter G. Sinyangwe, Fred Banda, Aaron S. Mweene, Ayato Takada, Hiroshi Kida
    JAPANESE JOURNAL OF VETERINARY RESEARCH 54 (1) 15 - 23 0047-1917 2006/05 [Refereed][Not invited]
     
    Anthrax is endemic in Western and North-western Provinces of Zambia. The disease occurs throughout the year and impacts negatively on the economy of the livestock industry and public health in Zambia. During 1989-1995, there were 1, 626 suspected cases of anthrax in cattle in Western province and of these 51 were confirmed. There were 220 cases of human anthrax cases in 1990 alone and 248 cases during 1991-1998 with 19.1% and 7.7% case fatality rates, respectively. Interplay of the ecology of affected areas and anthropogenic factors seem to trigger anthrax epidemics. Anthrax has drawn considerable attention in recent years due to its potential use as a biological weapon. In this paper, the history, current status and approaches towards the control of the disease in Zambia are discussed. Quarantine measures restrict trade of livestock and exchange of animals for draught power resulting in poor food security at household levels. Challenges of anthrax control are complex and comprise of socio-political, economical, environmental and cultural factors. Inadequate funding, lack of innovative disease control strategies and lack of cooperation from stakeholders are the major constraints to the control of the disease. It is hoped that the information provided here will stimulate continued awareness for the veterinary and medical authorities to maintain their surveillance and capabilities against the disease. This may lead to a culminating positive impact on livestock and human health in the southern African region.
  • T Horimoto, T Ayato, K Fujii, H Goto, M Hatta, S Watanabe, K Iwatsuki-Horimoto, M Ito, Y Tagawa-Sakai, S Yamada, H Ito, T Ito, M Imai, S Itamura, T Odagiri, M Tashiro, W Lim, Y Guan, M Peiris, Y Kawaoka
    VACCINE 24 (17) 3669 - 3676 0264-410X 2006/04 [Refereed][Not invited]
     
    The pandemic threat posed by highly pathogenic H5N1 influenza A viruses has created ail Urgent need for vaccines to protect against H5 virus infection. Because pathogenic viruses grow poorly in chicken eggs and their virulence poses a biohazard to vaccine producers, avirulent viruses produced by reverse genetics have become the preferred basis for vaccine production. Here, we investigated two key characteristics of potential H5 vaccine candidates: the hemaggutinin (HA) cleavage site sequence and its modification to attenuate virulence and the choice of background virus to provide a high-growth rate. We produced recombinant (6:2 reassortant) viruses that possessed a series of modified avirulent-type HA and neuraminidase genes, both of which were derived from an H5N1 human isolate. The other genes of these recombinant viruses were derived from donor virus strains known to grow well in eggs: the human strain A/Puerto Rico/8/34 (PR8) or an avian strain. All of the recombinant viruses grew well in eggs, were avirulent in chicks, and protected animals against infection with a wild-type virus. However, one of the recombinant viruses with an avian virus background acquired a Mutation in the HA cleavage site sequence that conferred virulence potential to this virus. Moreover, vaccine candidates with the avian virus background were more virulent than those with the human virus background. We conclude that 6:2 recombinant viruses with a PR8 background are more suitable than those with an avian virus background for vaccine development and that the HA cleavage site sequence must be modified to minimize the potential for a vaccine virus to convert to a virulent form. (c) 2006 Elsevier Ltd. All rights reserved.
  • Y Muramoto, A Takada, K Fujii, T Noda, K Iwatsuki-Horimoto, S Watanabe, T Horimoto, H Kida, Y Kawaoka
    JOURNAL OF VIROLOGY 80 (5) 2318 - 2325 0022-538X 2006/03 [Refereed][Not invited]
     
    The genome of influenza A viruses comprises eight negative-strand RNA segments. Although all eight segments must be present in cells for efficient viral replication, the mechanism(s) by which these viral RNA (vRNA) segments are incorporated into virions is not fully understood. We recently found that sequences at both ends of the coding regions of the HA, NA, and NS vRNA segments of A/WSN/33 play important roles in the incorporation of these vRNAs into virions. In order to similarly identify the regions of the PB2, PB1, and PA vRNAs of this strain that are critical for their incorporation, we generated a series of mutant vRNAs that possessed the green fluorescent protein gene flanked by portions of the coding and noncoding regions of the respective segments. For all three polymerase segments, deletions at the ends of their coding regions decreased their virion incorporation efficiencies. More importantly, these regions not only affected the incorporation of the segment in which they reside, but were also important for the incorporation of other segments. This effect was most prominent with the PB2 vRNA. These findings suggest a hierarchy among vRNA segments for virion incorporation and may imply intersegment association of vRNAs during virus assembly.
  • S Bar, A Takada, Y Kawaoka, M Alizon
    JOURNAL OF VIROLOGY 80 (6) 2815 - 2822 0022-538X 2006/03 [Refereed][Not invited]
     
    Ebola viruses (EboV) are enveloped RNA viruses infecting cells by a pH-dependent process mediated by viral glycoproteins (GP) involving endocytosis of virions and their rooting into acidic endosomes. As with well-characterized pH-dependent viral entry proteins, in particular influenza virus hemagglutinin.. it is thought that EboV GP require activation by low pH in order to mediate fusion of the viral envelope with the membrane of endosomes. However, it has not yet been possible to confirm the direct role of EboV GP in membrane fusion and the requirement for low-pH activation. It was in particular not possible to induce formation of syncytia by exposing cells expressing EboV GP to acidic medium. Here, we have used an assay based on the induction of a beta-galactosidase (lacZ) reporter gene in target cells to detect cytoplasmic exchanges, indicating membrane fusion, with cells expressing EboV GP (Zaire species). Acidic activation of GP-expressing cells was required for efficient fusion with target cells. The direct role of EboV GP in this process is indicated by its inhibition by anti-GP antibodies and by the lack of activity of mutant GP normally expressed at the cell surface but defective for virus entry. Fusion was not observed when target cells underwent acidic treatment, for example, when they were placed in coculture with GP-expressing cells before the activation step. This unexpected feature, possibly related to the nature of the EboV receptor, could explain the impossibility of inducing formation of syncytia among GIP-expressing cells.
  • K Okazaki, S Fujii, A Takada, H Kida
    VIRUS RESEARCH 115 (2) 105 - 111 0168-1702 2006/02 [Refereed][Not invited]
     
    In order to address the neutralization epitope on bovine herpesvirus 1 (BHV 1) glycoprotein B (gB), a panel of monoclonal antibodies (MAbs), a series of truncation forms of the glycoprotein and an MAb-escape mutant were used in this study. Immunocytochemistry on the truncations using MAbs against the glycoprotein revealed that the neutralization epitopes recognized by the MAbs lay between residues 1 and 52 of mature gB. Comparison of the sequences among the mutant, parent, and revertant viruses demonstrated that the amino-terminal residue of mature gB of the escape mutant was changed from Arg to Gln. These findings indicate that the amino-terminal residue of gB is critical for neutralization of BHV1. (c) 2005 Elsevier B.V. All rights reserved.
  • Y Muramoto, H Ozaki, A Takada, CH Park, Y Sunden, T Umemura, Y Kawaoka, H Matsuda, H Kida
    MICROBIOLOGY AND IMMUNOLOGY 50 (1) 73 - 81 0385-5600 2006 [Refereed][Not invited]
     
    Severe hemorrhage at multiple organs is frequently observed in chickens infected with highly pathogenic avian influenza (HPAI) A viruses. In this study we examined whether HPAI virus infection leads to coagulation disorder in chickens. Pathological examinations showed that the fibrin thrombi were formed in arterioles at the lung, associated with the viral antigens in endothelial cells of chickens infected intravenously with HPAI virus. Hematological analyses of peripheral blood collected from the chickens revealed that coagulopathy was initiated at early stage of infection when viral antigens were detected only in the endothelial cells and monocytes/macrophages. Furthermore, gene expression of the tissue factor, the main initiator of blood coagulation, was upregulated in the spleen, lung, and brain of HPAI virus-infected chickens. These results suggest that dysfunction of endothelial cells and monocytes/macrophages upon HPAI virus infection may induce hemostasis abnormalities represented by the excessive blood coagulation and consumptive coagulopathy in chickens.
  • T Noda, H Sagara, A Yen, A Takada, H Kida, RH Cheng, Y Kawaoka
    NATURE 439 (7075) 490 - 492 0028-0836 2006/01 [Refereed][Not invited]
     
    In viruses, as in eukaryotes, elaborate mechanisms have evolved to protect the genome and to ensure its timely replication and reliable transmission to progeny. Influenza A viruses are enveloped, spherical or filamentous structures, ranging from 80 to 120 nm in diameter(1). Inside each envelope is a viral genome consisting of eight single-stranded negative- sense RNA segments of 890 to 2,341 nucleotides each(1). These segments are associated with nucleoprotein and three polymerase subunits, designated PA, PB1 and PB2; the resultant ribonucleoprotein complexes (RNPs) resemble a twisted rod ( 10 - 15 nm in width and 30 - 120 nm in length) that is folded back and coiled on itself(2-4). Late in viral infection, newly synthesized RNPs are transported from the nucleus to the plasma membrane, where they are incorporated into progeny virions capable of infecting other cells. Here we show, by transmission electron microscopy of serially sectioned virions, that the RNPs of influenza A virus are organized in a distinct pattern ( seven segments of different lengths surrounding a central segment). The individual RNPs are suspended from the interior of the viral envelope at the distal end of the budding virion and are oriented perpendicular to the budding tip. This finding argues against random incorporation of RNPs into virions(5), supporting instead a model in which each segment contains specific incorporation signals that enable the RNPs to be recruited and packaged as a complete set(6-12). A selective mechanism of RNP incorporation into virions and the unique organization of the eight RNP segments may be crucial to maintaining the integrity of the viral genome during repeated cycles of replication.
  • QM Le, M Kiso, K Someya, YT Sakai, TH Nguyen, KHL Nguyen, ND Pham, HH Ngyen, S Yamada, Y Muramoto, T Horimoto, A Takada, H Goto, T Suzuki, Y Suzuki, Y Kawaoka
    NATURE 437 (7062) 1108 - 1108 0028-0836 2005/10 [Refereed][Not invited]
  • K Shinya, M Hatta, S Yamada, A Takada, S Watanabe, P Halfmann, T Horimoto, G Neumann, JH Kim, W Lim, Y Guan, M Peiris, M Kiso, T Suzuki, Y Suzuki, Y Kawaoka
    JOURNAL OF VIROLOGY 79 (15) 9926 - 9932 0022-538X 2005/08 [Refereed][Not invited]
     
    In 2003, H5N1 avian influenza virus infections were diagnosed in two Hong Kong residents who had visited the Fujian province in mainland China, affording us the opportunity to characterize one of the viral isolates, A/Hong Kong/213/03 (HK213; H5N1). In contrast to H5N1 viruses isolated from humans during the 1997 outbreak in Hong Kong, HK213 retained several features of aquatic bird viruses, including the lack of a deletion in the neuraminidase stalk and the absence of additional oligosaccharide chains at the globular head of the hemagglutinin molecule. It demonstrated weak pathogenicity in mice and ferrets but caused lethal infection in chickens. The original isolate failed to produce disease in ducks but became more pathogenic after five passages. Taken together, these findings portray the HK213 isolate as an aquatic avian influenza A virus without the molecular changes associated with the replication of H5N1 avian viruses in land-based poultry such as chickens. This case challenges the view that adaptation to land-based poultry is a prerequisite for the replication of aquatic avian influenza A viruses in humans.
  • G Neumann, H Ebihara, A Takada, T Noda, D Kobasa, LD Jasenosky, S Watanabe, JH Kim, H Feldmann, Y Kawaoka
    JOURNAL OF VIROLOGY 79 (16) 10300 - 10307 0022-538X 2005/08 [Refereed][Not invited]
     
    Ebola virus particle formation and budding are mediated by the VP40 protein, which possesses overlapping PTAP and PPXY late domain motifs (7-PTAPPXY-13). These late domain motifs have also been found in the Gag proteins of retroviruses and the matrix proteins of rhabdo- and arenaviruses. While in vitro studies suggest a critical role for late domain motifs in the budding of these viruses, including Ebola virus, it remains unclear as to whether the VP40 late domains play a role in Ebola virus replication. Alteration of both late domain motifs drastically reduced VP40 particle formation in vitro. However, using reverse genetics, we were able to generate recombinant Ebola virus containing mutations in either or both of the late domains. Viruses containing mutations in one or both of their late domain motifs were attenuated by one log unit. Transmission and scanning electron microscopy did not reveal appreciable differences between the mutant and wild-type viruses released from infected cells. These findings indicate that the Ebola VP40 late domain motifs enhance virus replication but are not absolutely required for virus replication in cell culture.
  • Y Maeda, M Hatta, A Takada, T Watanabe, H Goto, G Neumann, Y Kawaoka
    JOURNAL OF VIROLOGY 79 (11) 6674 - 6679 0022-538X 2005/06 [Refereed][Not invited]
     
    Influenza and human parainfluenza virus infections are of both medical and economical importance. Currently, inactivated vaccines provide suboptimal protection against influenza, and vaccines for human parainfluenza virus infection are not available, underscoring the need for new vaccines against these respiratory diseases. Furthermore, to reduce the burden of vaccination, the development of multivalent vaccines is highly desirable. Thus, to devise a single vaccine that would elicit immune responses against both influenza and parainfluenza viruses, we used reverse genetics to generate an influenza A virus that possesses the coding region for the hemagglutinin/neuraminidase ectodomain of parainfluenza virus instead of the influenza virus neuraminidase. The recombinant virus grew efficiently in eggs but was attenuated in mice. When intranasally immunized with the recombinant vaccine, all mice developed antibodies against both influenza and parainfluenza viruses and survived an otherwise lethal challenge with either of these viruses. This live bivalent vaccine has obvious advantages over combination vaccines, and its method of generation could, in principle, be applied in the development of a "cocktail" vaccine with efficacy against several different infectious diseases.
  • K Fujii, Y Fujii, T Noda, Y Muramoto, T Watanabe, A Takada, H Goto, T Horimoto, Y Kawaoka
    JOURNAL OF VIROLOGY 79 (6) 3766 - 3774 0022-538X 2005/03 [Refereed][Not invited]
     
    The genome of influenza A virus consists of eight single-strand negative-sense RNA segments, each comprised of a coding region and a noncoding region. The noncoding region of the NS segment is thought to provide the signal for packaging; however, we recently showed that the coding regions located at both ends of the hemagglutinin and neuraminidase segments were important for their incorporation into virions. In an effort to improve our understanding of the mechanism of influenza virus genome packaging, we sought to identify the regions of NS viral RNA (vRNA) that are required for its efficient incorporation into virions. Deletion analysis showed that the first 30 nucleotides of the 3' coding region are critical for efficient NS vRNA incorporation and that deletion of the 3' segment-specific noncoding region drastically reduces NS vRNA incorporation into virions. Furthermore, silent mutations in the first 30 nucleotides of the 3' NS coding region reduced the incorporation efficiency of the NS segment and affected virus replication. These results suggested that segment-specific noncoding regions together with adjacent coding regions (especially at the 3' end) form a structure that is required for efficient influenza A virus vRNA packaging.
  • D Kobasa, A Takada, K Shinya, M Hatta, P Halfmann, S Theriault, H Suzuki, H Nishimura, K Mitamura, N Sugaya, T Usui, T Murata, Y Maeda, S Watanabe, M Suresh, T Suzuki, Y Suzuki, H Feldmann, Y Kawaoka
    NATURE 431 (7009) 703 - 707 0028-0836 2004/10 [Refereed][Not invited]
     
    The 'Spanish' influenza pandemic of 1918-19 was the most devastating outbreak of infectious disease in recorded history. At least 20 million people(1) died from their illness, which was characterized by an unusually severe and rapid clinical course. The complete sequencing of several genes of the 1918 influenza virus has made it possible to study the functions of the proteins encoded by these genes in viruses generated by reverse genetics, a technique that permits the generation of infectious viruses entirely from cloned complementary DNA. Thus, to identify properties of the 1918 pandemic influenza A strain that might be related to its extraordinary virulence, viruses were produced containing the viral haemagglutinin(2) (HA) and neuraminidase(3) (NA) genes of the 1918 strain. The HA of this strain supports the pathogenicity of a mouse-adapted virus in this animal(4,5). Here we demonstrate that the HA of the 1918 virus confers enhanced pathogenicity in mice to recent human viruses that are otherwise non-pathogenic in this host. Moreover, these highly virulent recombinant viruses expressing the 1918 viral HA could infect the entire lung and induce high levels of macrophage-derived chemokines and cytokines, which resulted in infiltration of inflammatory cells and severe haemorrhage, hallmarks of the illness produced during the original pandemic(6).
  • T Horimoto, A Takada, K Iwatsuki-Horimoto, Y Kawaoka
    VACCINE 22 (17-18) 2244 - 2247 0264-410X 2004/06 [Refereed][Not invited]
     
    Previously, we generated influenza A viruses that possess chimeric type (A/B) hemagglutinins (HA), in which immunogenic regions of type A HA were replaced with those of type B HA, and showed that these viruses were attenuated in mice (J. Virol. 77 (2003) 803 1). Here, we intranasally immunized mice with these viruses and then challenged them with a wild-type A virus to assess a protective immune response to viral components other than HA in the form of a live virus. All immunized mice survived challenge with a lethal dose of wild-type virus; none or a limited amount of virus, if any, was recovered from nasal turbinates or lungs of the mice 3 days post-challenge. These results provide direct evidence that immune responses to viral components other than HA confer protection against influenza A virus infection in a mouse model, suggesting the usefulness of live vaccines for viruses that have undergone antigenic drift with respect to HA, or for viruses with heterosubtypic HAs. (C) 2003 Elsevier Ltd. All rights reserved.
  • A Takada, K Fujioka, M Tsuiji, A Morikawa, N Higashi, H Ebihara, D Kobasa, H Feldmann, T Irimura, Y Kawaoka
    JOURNAL OF VIROLOGY 78 (6) 2943 - 2947 0022-538X 2004/03 [Refereed][Not invited]
     
    Filoviruses cause lethal hemorrhagic disease in humans and nonhuman primates. An initial target of filovirus infection is the mononuclear phagocytic cell. Calcium-dependent (C-type) lectins such as dendritic cell- or liver/lymph node-specific ICAM-3 grabbing nonintegrin (DC-SIGN or L-SIGN, respectively), as well as the hepatic asialoglycoprotein receptor, bind to Ebola or Marburg virus glycoprotein (GP) and enhance the infectivity of these viruses in vitro. Here, we demonstrate that a recently identified human macrophage galactose- and N-acetylgalactosamine- specific C-type lectin (hMGL), whose ligand specificity differs from DC-SIGN and L-SIGN, also enhances the infectivity of filoviruses. This enhancement was substantially weaker for the Reston and Marburg viruses than for the highly pathogenic Zaire virus. We also show that the heavily glycosylated, mucin-like domain on the filovirus GP is required for efficient interaction with this lectin. Furthermore, hMGL, like DC-SIGN and L-SIGN, is present on cells known to be major targets of filoviruses (i.e., macrophages and dendritic cells), suggesting a role for these C-type lectins in viral replication in vivo. We propose that filoviruses use different C-type lectins to gain cellular entry, depending on the cell type, and promote efficient viral replication.
  • Y Maeda, H Goto, T Horimoto, A Takada, Y Kawaoka
    VIRUS RESEARCH 100 (2) 153 - 157 0168-1702 2004/03 [Refereed][Not invited]
     
    The levels of viral proteins in infected cells are thought to be regulated by a variety of mechanisms. The initiation codons for the PB1 and NA proteins of A/WSN/33 (H1N1) influenza virus are in a suboptimal Kozak sequence for translation. To determine the significance of these suboptimal Kozak sequences, model vRNAs, whose coding regions were replaced with the reporter SEAP gene (for secreted alkaline phosphatase) and recombinant viruses with optimal Kozak sequences for PB1 and NA were constructed. Conversion of the upstream sequence of the PB 1 and NA initiation codon to an optimal Kozak sequence was reflected in the level of reporter protein expression, but not the level of PB1 and NA protein expression. The recombinant viruses that had optimal Kozak sequences for PB1, NA, or both genes had similar replicative properties, both in cell culture and in mice, to those of the wild-type virus. These results suggest that expression of the PB1 and NA proteins is regulated by a mechanism other than that controlling the initiation of translation of these proteins. (C) 2003 Elsevier B.V. All rights reserved.
  • S Watanabe, T Watanabe, T Noda, A Takada, H Feldmann, LD Jasenosky, Y Kawaoka
    JOURNAL OF VIROLOGY 78 (2) 999 - 1005 0022-538X 2004/01 [Refereed][Not invited]
     
    We established a plasmid-based system for generating infectious Ebola virus-like particles (VLPs), which contain an Ebola virus-like minigenome consisting of a negative-sense copy of the green fluorescent protein gene. This system produced nearly 10(3) infectious particles per ml of supernatant, equivalent to the titer of Ebola virus generated by a reverse genetics system. Interestingly, infectious Ebola VLPs were generated, even without expression of VP24. Transmission and scanning electron microscopic analyses showed that the morphology of the Ebola VLPs was indistinguishable from that of authentic Ebola virus. Thus, this system allows us to study Ebola virus entry, replication, and assembly without biosafety level 4 containment. Furthermore, it may be useful in vaccine production against this highly pathogenic agent.
  • H Tanaka, CH Park, A Ninomiya, H Ozaki, A Takada, T Umemura, H Kida
    VETERINARY MICROBIOLOGY 95 (1-2) 1 - 13 0378-1135 2003/08 [Refereed][Not invited]
     
    The direct transmission of H5N1 influenza A viruses from chickens to humans in Hong Kong in 1997 emphasized the need to have information on the pathogenesis of avian influenza virus infection in mammals. H5N1 influenza viruses isolated from patients during the incident killed experimentally infected mice. The principal lesions of the mice were broncho-interstitial pneumonia and nonsuppurative encephalitis. Infectious viruses and/or viral antigens were detected in the brain as well as in the trigeminal and vagal ganglia but not in the blood of the mice. These findings suggest that the virus reached the brain through the vagus and/or trigeminal nerves following replication in the respiratory mucosa. The results imply that neurotropism of the H5N1 virus in mice is a novel characteristic in the pathogenesis of infection by human influenza virus isolates. (C) 2003 Elsevier B.V. All rights reserved.
  • A Takada, S Matsushita, A Ninomiya, Y Kawaoka, H Kida
    VACCINE 21 (23) 3212 - 3218 0264-410X 2003/07 [Refereed][Not invited]
     
    It has been known that influenza A virus infection induces a cross-protective immunity against infection by viruses with different subtypes of viral envelope proteins, hemagglutinin (HA) and neuraminidase (NA). This heterosubtypic immunity is generally mediated by cytotoxic T lymphocytes (CTL) reactive to specific epitopes in the viral internal proteins, such as nucleoprotein and matrix protein. By contrast, immunization with inactivated virus antigens has been thought to be unable to generate heterosubtypic immunity, since inactivated antigens do not usually induce CTL responses. However, we show that intranasal immunization with formalin-inactivated intact virus, but not ether-split vaccines, induced a broad spectrum of heterosubtypic protective immunity in mice. The protection may be mediated by the mucosal immune response, most likely secretory IgA antibodies to the viral proteins. This approach may overcome limitations in the efficacy of inactivated influenza vaccines and confer potent immunity to humans against viruses with new pandemic potential. (C) 2003 Elsevier Science Ltd. All rights reserved.
  • A Takada, H Feldmann, TG Ksiazek, Y Kawaoka
    JOURNAL OF VIROLOGY 77 (13) 7539 - 7544 0022-538X 2003/07 [Refereed][Not invited]
     
    Most strains of Ebola virus cause a rapidly fatal hemorrhagic disease in humans, yet there are still no biologic explanations that adequately account for the extreme virulence of these emerging pathogens. Here we show that Ebola Zaire virus infection in humans induces antibodies that enhance viral infectivity. Plasma or serum from convalescing patients enhanced the infection of primate kidney cells by the Zaire virus, and this enhancement was mediated by antibodies to the viral glycoprotein and by complement component C1q. Our results suggest a novel mechanism of antibody-dependent enhancement of Ebola virus infection, one that would account for the dire outcome of Ebola outbreaks in human populations.
  • T Horimoto, A Takada, K Iwatsuki-Horimoto, M Hatta, H Goto, Y Kawaoka
    JOURNAL OF VIROLOGY 77 (14) 8031 - 8038 0022-538X 2003/07 [Refereed][Not invited]
     
    To gain insight into the intertypic incompatibility between type A and B influenza viruses, we focused on the hemagglutinin (HA) gene, systematically studying the compatibility of chimeric (type A/B) HAs with a type A genetic background. An attempt to generate a reassortant containing an intact type B HA segment in a type A virus background by reverse genetics was unsuccessful despite transcription of the type B HA segment by the type A polymerase complex. Although a type A virus with a chimeric HA segment comprising the entire coding sequence of the type B HA flanked by the noncoding sequence of the type A HA was viable, it replicated only marginally. Other chimeric viruses contained type A/B HAs possessing the type A noncoding region together with either the signal peptide or transmembrane/cytoplasmic region of type A virus or both, with the remaining regions derived from the type B HA. Each of these viruses grew to median tissue culture infectious doses of more than 10(5) per ml, but those with more type A HA regions replicated better, suggesting protein-protein interactions or increased HA segment incorporation into virions as contributing factors in the efficient growth of this series of viruses. All of these chimeric (A/B) HA viruses were attenuated in mice compared with wild-type A or B viruses. All animals intranasally immunized with a chimeric virus survived upon challenge with a lethal dose of wild-type type B virus. These results suggest a framework for the design of a novel live vaccine virus.
  • A Takada, H Feldmann, U Stroeher, M Bray, S Watanabe, H Ito, M McGregor, Y Kawaoka
    JOURNAL OF VIROLOGY 77 (2) 1069 - 1074 0022-538X 2003/01 [Refereed][Not invited]
     
    Ebola virus causes lethal hemorrhagic fever in humans, but currently there are no effective vaccines or antiviral compounds for this infectious disease. Passive transfer of monoclonal antibodies (MAbs) protects mice from lethal Ebola virus infection (J. A. Wilson, M. Hevey, R. Bakken, S. Guest, M. Bray, A. L. Schmaljohn, and M. K. Hart, Science 287:1664-1666, 2000). However, the epitopes responsible for neutralization have been only partially characterized because some of the MAbs do not recognize the short synthetic peptides used for epitope mapping. To identify the amino acids recognized by neutralizing and protective antibodies, we generated a recombinant vesicular stomatitis virus (VSV) containing the Ebola virus glycoprotein-encoding gene instead of the VSV G protein-encoding gene and used it to select escape variants by growing it in the presence of a MAb (133/3.16 or 226/8.1) that neutralizes the infectivity of the virus. All three variants selected by MAb 133/3.16 contained a single amino acid substitution at amino acid position 549 in the GP2 subunit. By contrast, MAb 226/8.1 selected three different variants containing substitutions at positions 134, 194, and 199 in the GP1 subunit, suggesting that this antibody recognized a conformational epitope. Passive transfer of each of these MAbs completely protected mice from a lethal Ebola virus infection. These data indicate that neutralizing antibody cocktails for passive prophylaxis and therapy of Ebola hemorrhagic fever can reduce the possibility of the emergence of antigenic variants in infected individuals.
  • A Ninomiya, A Takada, K Okazaki, KF Shortridge, H Kida
    VETERINARY MICROBIOLOGY 88 (2) 107 - 114 0378-1135 2002/08 [Refereed][Not invited]
     
    Pig serum samples collected in southeastern China were examined for antibodies to influenza A viruses. Since the hemagglutination inhibition (HI) test does not accurately detect antibodies to the hemagglutinins (HAs) of "avian" influenza viruses, we utilized the neutralization (NT) test to detect subtype-specific antibodies to the HA of avian viruses in pig sera. Neutralizing antibodies to H1, H3, H4, and H5 influenza viruses were detected in the serum samples collected in 1977-1982 and 1998, suggesting that pigs in China have been sporadically infected with avian H4 and H5 viruses in addition to swine and human HI and H3 viruses. Antibodies to H9 virus, on the other hand, were found only in the sera collected in 1998, not in those collected in 1977-1982, correlating with the recent spread in poultry and subsequent isolation of H9N2 viruses from pigs and humans in 1998. The present results indicate that avian influenza viruses have been transmitted to pig populations in southeastern China. (C) 2002 Elsevier Science B.V. All rights reserved.
  • A Ninomiya, K Ogasawara, K Kajino, A Takada, H Kida
    VACCINE 20 (25-26) 3123 - 3129 0264-410X 2002/08 [Refereed][Not invited]
     
    Mucosal immunity is critical for protection from viral infections. We attempted to activate mucosal cytotoxic T lymphocytes (CTLs) specific for influenza A virus nucleoprotem (NP) which play an important role in protective immunity. It has been shown that dendritic cells (DCs) activated by signaling via CD40-CD40 ligand (CD40L) interaction are required for the differentiation of naive CD8(+) T cells into antigen-specific CTLs in a non-mucosal environment. We herein inoculated mice intranasally with an anti-CD40 monoclonal antibody (anti-CD40 mAb) and NP366-374 peptide, corresponding to a CTL epitope on NP, encapsulated in liposome (liposomal NP366-374) to induce protective CTL responses against influenza A virus. Intranasal but not subcutaneous immunization with liposomal NP366-374 effectively induced mucosal immunity to reduce virus replication in the lung, suggesting that anti-CD40 mAb also functioned as a mucosal adjuvant. Interestingly, neither MHC class I- nor class II-deficient mice immunized intranasally with these materials were resistant to the infection. Since anti-CD40 mAb was considered to help replace CD4(+) T cells, another help of CD4(+) T cells are presumably required for the induction of CTL activity in the lung. This approach may prove promising for developing vaccines to induce mucosal CTL responses, and seems to highlight differences between mucosal and non-mucosal immunity. (C) 2002 Elsevier Science Ltd. All rights reserved.
  • T Noda, H Sagara, E Suzuki, A Takada, H Kida, Y Kawaoka
    JOURNAL OF VIROLOGY 76 (10) 4855 - 4865 0022-538X 2002/05 [Refereed][Not invited]
     
    Using biochemical assays, it has been demonstrated that expression of Ebola virus VP40 alone in mammalian cells induced production of particles with a density similar to that of virions. To determine the morphological properties of these particles, cells expressing VP40 and the particles released from the cells were examined by electron microscopy. VP40 induced budding from the plasma membrane of filamentous particles, which differed in length but had uniform diameters of approximately 65 nm. When the Ebola virus glycoprotein (GP) responsible for receptor binding and membrane fusion was expressed in cells, we found pleomorphic particles budding from the plasma membrane. By contrast, when GP was coexpressed with VP40, GP was found on the filamentous particles induced by VP40. These results demonstrated the central role of VP40 in formation of the filamentous structure of Ebola virions and may suggest an interaction between VP40 and GP in morphogenesis.
  • JH Ko, HK Jin, A Asano, A Takada, A Ninomiya, H Kida, H Hokiyama, M Ohara, M Tsuzuki, M Nishibori, M Mizutani, T Watanabe
    GENOME RESEARCH 12 (4) 595 - 601 1088-9051 2002/04 [Refereed][Not invited]
     
    The nucleotide sequence of chicken Mx cDNA was reported earlier using the White Leghorn breed in Germany, but it showed no enhanced resistance to viruses. In this study, the nucleotide sequences of chicken Mx cDNA were determined in many breeds. A total of 25 nucleotide substitutions, of which 14 were deduced to cause amino acid exchanges, were detected, Suggesting that the chicken Mx gene is very polymorphic. Transfected cell clones expressing chicken Mx mRNA were established after the Mx cDNA was constructed with an expression vector and introduced into mouse 3T3 cells, and the Mx genes from some breeds were demonstrated to confer positive antiviral responses to influenza virus and vesicular stomatitis virus. On the basis of the comparison among the antiviral activities associated with many Mx variations, a specific amino acid substitution at position 631 (Ser to Asn) was considered to determine the antivirally positive or negative Mx gene. Thus, a single amino acid Substitution influences the antiviral activity of Mx in domesticated chickens.
  • CH Park, M Ishinaka, A Takada, H Kida, T Kimura, K Ochiai, T Umemura
    ARCHIVES OF VIROLOGY 147 (7) 1425 - 1436 0304-8608 2002 [Refereed][Not invited]
     
    A/Hong Kong/483/97 (H5N1) influenza virus (HK483) isolated from the third patient during the outbreak of chicken and human influenza in Hong Kong in 1997 was shown to be neurovirulent in mice. HK483 was inoculated intranasally to mice, and the invasion routes of the virus in the central nervous system (CNS) were investigated by immunohistochemical and in situ hybridization. The pathological changes consisted of bronchopneumonia, ganglionitis, and nonpurulent encephalomyelitis of the brain stem and the anterior part of the thoracic cord. Viral antigens and viral nucleic acids (RNA and mRNA) were demonstrated in the pterygopalatine, trigeminal and superior ganglions prior to or simultaneously with their detection in the CNS. The antigens and nucleic acids were also observed in the olfactory bulb from an early stage of the infection. In the spinal cord, virus-infected cells were first demonstrated in the grey matter of the thoracic cord. The virus, which primarily replicated in the lungs, was considered to invade the thoracic cord via cardiopulmonary splanchnic nerves and sympathetic nerves. These findings indicate that the virus reached the CNS through afferent fibers of the olfactory, vagal, trigeminal, and sympathetic nerves following replication in the respiratory mucosa.
  • H Goto, K Wells, A Takada, Y Kawaoka
    JOURNAL OF VIROLOGY 75 (19) 9297 - 9301 0022-538X 2001/10 [Refereed][Not invited]
     
    When expressed in vitro, the neuraminidase (NA) of A/WSN/33 (WSN) virus binds and sequesters plasminogen on the cell surface, leading to enhanced cleavage of the viral hemagglutinin. To obtain direct evidence that the plasminogen-binding activity of the NA enhances the pathogenicity of WSN virus, we generated mutant viruses whose NAs lacked plasminogen-binding activity because of a mutation at the C terminus, from Lys to Arg or Leu. In the presence of trypsin, these mutant viruses replicated similarly to wild-type virus in cell culture. By contrast, in the presence of plasminogen, the mutant viruses failed to undergo multiple cycles of replication while the wild-type virus grew normally. The mutant viruses showed attenuated growth in mice and failed to grow at all in the brain. Furthermore, another mutant WSN virus, possessing an NA with a glycosylation site at position 130 (146 in N2 numbering), leading to the loss of neurovirulence, failed to grow in cell culture in the presence of plasminogen. We conclude that the plasminogen-binding activity of the WSN NA determines its pathogenicity in mice.
  • CH Park, H Ozaki, A Takada, H Kida, K Ochiai, T Umemura
    AVIAN PATHOLOGY 30 (3) 269 - 272 0307-9457 2001/06 [Refereed][Not invited]
     
    Virulent or avirulent strains of type A influenza virus were inoculated into the allantoic cavities of chicken embryos. The antigens of virulent strains appeared initially in the surface epithelium of the allantoic membrane, then in vascular endothelial cells of the chorioallantoic membrane and visceral organs of the embryos, and then spread to parenchymal cells of many organs. In contrast, the antigens of the avirulent strain were confined to the allantoic membrane. These observations indicate that the primary target of virulent influenza viruses in chicken embryos is vascular endothelial cells, and that the embryos died after systemic viral infection.
  • A Takada, S Watanabe, K Okazaki, H Kida, Y Kawaoka
    JOURNAL OF VIROLOGY 75 (5) 2324 - 2330 0022-538X 2001/03 [Not refereed][Not invited]
     
    Ebola virus causes severe hemorrhagic fever in primates, resulting in mortality rates of up to 100%, yet there are no satisfactory biologic explanations for this extreme virulence. Here we show that antisera produced by DNA immunization with a plasmid encoding the surface glycoprotein (GP) of the Zaire strain of Ebola virus enhances the infectivity of vesicular stomatitis virus pseudotyped with the GP. Substantially weaker enhancement was observed with antiserum to the GP of the Reston strain, which is much less pathogenic In humans than the Ebola Zaire and Sudan viruses. The enhancing activity was abolished by heat but was increased in the presence of complement system inhibitors, suggesting that heat-labile factors other than the complement system are required for this effect. We also generated an anti-Zaire GP monoclonal antibody that enhanced viral infectivity and another that neutralized it, indicating the presence of distinct epitopes for these properties. Our findings suggest that antibody-dependent enhancement of infectivity may account for the extreme virulence of the virus. They also raise issues about the development of Ebola virus vaccines and the use of passive prophylaxis or therapy with Ebola virus GP antibodies.
  • H Ito, S Watanabe, A Takada, Y Kawaoka
    JOURNAL OF VIROLOGY 75 (3) 1576 - 1580 0022-538X 2001/02 [Refereed][Not invited]
     
    Using the vesicular stomatitis virus (VSV) pseudotype system, we studied the functional properties of the Ebola virus glycoprotein (GP). Amino acid substitutions at the GP cleavage site, which reduce glycoprotein cleavability and viral infectivity in some viruses, did not appreciably change the infectivity of VSV pseudotyped dth GP. Likewise, removal of two acylated cysteine residues in the transmembrane region of GP showed no discernible effects on infectivity. Although most filoviruses are believed to target endothelial cells and hepatocytes preferentially, the GP-carrying VSV shelved greater affinity for epithelial cells than for either of these cell types, indicating that Ebola virus GP does not necessarily have strong tropism toward endothelial cells and hepatocytes. Finally, when it was used to screen for neutralizing antibodies against Ebola virus GP, the VSV pseudotype system allowed us to detect strain-specific neutralizing activity that was inhibited by secretory GP (SGP). This finding provides evidence of shared neutralizing epitopes on GP and SGP molecules and indicates the potential of SGP to serve as a decoy for neutralizing antibodies.
  • YK Lim, A Takada, T Tanizaki, H Ozaki, K Okazaki, H Kida
    JAPANESE JOURNAL OF VETERINARY RESEARCH 48 (4) 197 - 203 0047-1917 2001/02 [Refereed][Not invited]
     
    Effective vaccinations against swine influenza reduce the economic loss of pig industries, and also may minimize the possibility of emergence of new pandemic viruses, since pigs are intermediate hosts to generate reassortant viruses among avian and mammalian influenza viruses. In this study, we showed that intranasal immunization of pigs with formalin-inactivated or ether-split influenza vaccine (A/Aichi/2/68) induced virus-specific IgG, IgM, and IgA antibodies in their nasal secretions and sera, resulting in complete protection from virus challenge. Antibody response to the challenge virus was not observed in the immunized pigs, suggesting that the replication of the virus in the primary targets, respiratory epithelial cells, was inhibited. The present results indicate that intranasal immunization of pigs with inactivated vaccines is effective to control swine influenza, and also provide a good model, as well as a mouse model, to evaluate an intranasal application of influenza vaccine for humans.
  • HK Jin, K Yoshimatsu, A Takada, M Ogino, A Asano, J Arikawa, T Watanabe
    ARCHIVES OF VIROLOGY 146 (1) 41 - 49 0304-8608 2001 [Refereed][Not invited]
     
    The antiviral potential of Mx2 protein remains unknown, because the Mx2 gene in commonly used strains of laboratory mice is nonfunctional. Our previous study showed that functional Mx2 protein in some feral-origin strains was induced upon interferon treatment, was localized in the cytoplasm, and inhibited vesicular stomatitis virus replication. In the present study, we have demonstrated chat the embryonic fibroblastic cells from a feral-origin strain (SPR) expressed 74 kDa Mx2 protein, which prevented the accumulation of viral transcripts and proteins of hantaviruses when the Mx2 gene was constitutively expressed in transfected Vero cells. Furthermore, the cells showed significantly lower titers of the virus than control cells. In contrast, influenza virus replication was not affected by the expression of Mx2 protein in the Vero cells.
  • A Takada, S Watanabe, H Ito, K Okazaki, H Kida, Y Kawaoka
    VIROLOGY 278 (1) 20 - 26 0042-6822 2000/12 [Refereed][Not invited]
     
    Filoviruses, including Ebola virus, are cytotoxic. To investigate the role of the Ebola virus glycoprotein (GP) in this cytopathic effect, we transiently expressed the GP in human kidney 293T cells. Expression of wild-type GP, but not the secretory form of the molecule lacking a membrane anchor, induced rounding and detachment of the cells, as did a chimeric GP containing its ectodomain and influenza virus hemagglutinin transmembrane-cytoplasmic domain. These results indicate that the GP ectodomain and its anchorage to the membrane are required for GP-induced morphologic changes in host cells. Since cell rounding and detachment could be associated with reduced levels of cell adhesion molecules, we also studied the expression of integrins, which are major molecules for adhesion to extracellular matrices, and found that the beta1 integrin group is downregulated by the GP. This result was further extended by experiments in which anti-beta1 monoclonal antibodies or purified integrins inhibited the infectivity of vesicular stomatitis virus pseudotyped with the GP. We suggest that integrins, especially the beta1 group, might interact with the GP and perhaps be involved in Ebola virus entry into cells, (C) 2000 Academic Press.
  • S Watanabe, A Takada, T Watanabe, H Ito, H Kida, Y Kawaoka
    JOURNAL OF VIROLOGY 74 (21) 10194 - 10201 0022-538X 2000/11 [Refereed][Not invited]
     
    Ebola virus contains a single glycoprotein (GP) that is responsible for receptor binding and membrane fusion and is proteolytically cleaved into disulfide-linked GP1 and GP2 subunits. The GP2 subunit possesses a coiled-coil motif, which plays an important role in the oligomerization and fusion activity of other viral GPs. To determine the functional significance of the coiled-coil motif of GP2, we examined the effects of peptides corresponding to the coiled-coil motif of GP2 on the infectivity of a mutant vesicular stomatitis virus (lacking the receptor-binding/fusion protein) pseudotyped with the Ebola virus GP. A peptide corresponding to the C-terminal helix reduced the infectivity of the pseudotyped virus. We next introduced alanine substitutions into hydrophobic residues in the coiled-coil motif to identify residues important for GP function. None of the substitutions affected GP oligomerization, but some mutations, two in the N-terminal helix and all in the C-terminal helix, reduced the ability of GP to confer infectivity to the mutant vesicular stomatitis virus without affecting the transport of GP to the cell surface, its incorporation into virions, and the production of virus particles. These results indicate that the coiled-coil motif of GP2 plays an important role in facilitating the entry of Ebola virus into host cells and that peptides corresponding to this region could act as efficient antiviral agents.
  • Watanabe Shinji, Takada Ayato, Watanabe Tokiko, Ito Hiroshi, Kida Hiroshi, Kawaoka Yoshihiro
    Journal of Virology American Society for Microbiology 74 (21) 10194 - 10201 0022-538X 2000/11 [Not refereed][Not invited]
     
    Ebola virus contains a single glycoprotein (GP) that is responsible for receptor binding and membrane fusion and is proteolytically cleaved into disulfide-linked GP1 and GP2 subunits. The GP2 subunit possesses a coiled-coil motif, which plays an important role in the oligomerization and fusion activity of other viral GPs. To determine the functional significance of the coiled-coil motif of GP2, we examined the effects of peptides corresponding to the coiled-coil motif of GP2 on the infectivity of a mutant vesicular stomatitis virus (lacking the receptor-binding/fusion protein) pseudotyped with the Ebola virus GP. A peptide corresponding to the C-terminal helix reduced the infectivity of the pseudotyped virus. We next introduced alanine substitutions into hydrophobic residues in the coiled-coil motif to identify residues important for GP function. None of the substitutions affected GP oligomerization, but some mutations, two in the N-terminal helix and all in the C-terminal helix, reduced the ability of GP to confer infectivity to the mutant vesicular stomatitis virus without affecting the transport of GP to the cell surface, its incorporation into virions, and the production of virus particles. These results indicate that the coiled-coil motif of GP2 plays an important role in facilitating the entry of Ebola virus into host cells and that peptides corresponding to this region could act as efficient antiviral agents.
  • T Ito, Y Suzuki, T Suzuki, A Takda, T Horimoto, K Wells, H Kida, K Otsuki, M Kiso, H Ishida, Y Kawaoka
    JOURNAL OF VIROLOGY 74 (19) 9300 - 9305 0022-538X 2000/10 [Refereed][Not invited]
     
    The hemagglutinin (IU) of H3 human influenza viruses does not support viral replication in duck intestine despite its avian origin. A Leu-to-Gin mutation at position 226 and a Ser-to-Gly mutation at position 228 in the HA of human A/Udorn/307/72 (H3N2) permit a reassortant virus [human Udorn HA, with all other genes from A/mallard/New York/6750/78 (H2N2)] to replicate in ducks. To understand the molecular basis of this change in host range restriction, we investigated the receptor specificity of duck influenza viruses as well as of human-duck virus reassortants. The results indicate that the recognition of a glycoconjugate moiety possessing N-glycolneuramic acid (NeuGc) linked to galactose by the alpha 2,3 linkage (NeuGc alpha 2,3Gal) is associated with viral replication in duck intestine. Immunofluorescence assays with NeuGc alpha 2,3Gal-specific antiserum detected this moiety primarily on the crypt epithelial cells of duck colon. Such recognition, together with biochemical evidence of NeuGc in crypt cells, correlated exactly with the ability of the virus to replicate in duck colon. These results suggest that recognition of the NeuGc alpha 2,3-Gal moiety plays an important role in the enterotropism of avian influenza viruses.
  • K Shinya, A Shimada, T Ito, K Otsuki, T Morita, H Tanaka, A Takada, H Kida, T Umemura
    ARCHIVES OF VIROLOGY 145 (1) 187 - 195 0304-8608 2000 [Refereed][Not invited]
     
    To define the route of influenza virus invasion into the central nervous system (CNS), an avian influenza A (H5N3) virus was inoculated into mice intranasally or intravenously. Only the intranasal infection group mice showed depression and retention of gas in the digestive system. Pathological findings in the animals were bronchointerstitial pneumonia and non-suppurative encephalitis restricted to the brain stem. The nerve nucleus primarily affected was the nucleus of solitary tract. Prior to the development of the CNS lesions, viral antigen was detected in vagal and trigeminal ganglia. These results suggest that the primarily replicated virus in the respiratory mucosa ascended to the CNS via sensory nerve routes, inducing lesions in the brain stem, and then spread trans-synaptically in the CNS.
  • K Okazaki, A Takada, T Ito, M Imai, H Takakuwa, M Hatta, H Ozaki, T Tanizaki, T Nagano, A Ninomiya, VA Demenev, MM Tyaptirganov, TD Karatayeva, SS Yamnikova, DK Lvov, H Kida
    ARCHIVES OF VIROLOGY 145 (5) 885 - 893 0304-8608 2000 [Refereed][Not invited]
     
    Influenza A viruses of different subtypes were isolated from fecal samples of ducks in their nesting areas in Siberia in summer from 1996 to 1998. Phylogenetic analysis of the NP genes of the isolates in Siberia and those in Hokkaido, Japan on their flyway of migration from Siberia to the south in autumn revealed that they belong to the Eurasian lineage of avian influenza viruses. It is noted that the genes of the isolates in Siberia are closely related to those of H5N1 influenza virus strains isolated from chickens and humans in Hong Kong in 1997 as well as to those of isolates from domestic birds in southern China. The results indicate that influenza viruses perpetuated in ducks nesting in Siberia should have contributed genes in the emergence of the H5N1 virus in Hong Kong. Vaccine prepared from avirulent A/duck/Hokkaido/4/96 (H5N3) influenza virus was potent enough to protect mice from challenge with lethal dose of the pathogenic H5N1 virus [19]. Intensive surveillance study of aquatic birds especially in Siberia is, therefore, stressed to provide information on the future pandemic influenza virus strains and for vaccine preparation.
  • M Miyoshi, M Takiguchi, J Yasuda, A Hashimoto, A Takada, K Okazaki, H Kida
    ARCHIVES OF VIROLOGY 145 (8) 1715 - 1723 0304-8608 2000 [Refereed][Not invited]
     
    The canine herpesvirus infected cell protein 0 (CICP0) gene was sequenced. The CICP0 gene was transcribed as a 1.4 kb mRNA from the end of the unique long region nearby the internal repeat during early phase of productive infection of the virus. An open reading frame of the gene encodes a polypeptide of 333 amino acids. The RING finger domain and acidic transcriptional activation domain were found at the N-terminus and within the middle region in the deduced amino acid sequence, respectively, suggesting that the CICP0, like the ICP0 of herpes simplex virus 1, is a transactivating protein.
  • A Takada, N Kuboki, K Okazaki, A Ninomiya, H Tanaka, H Ozaki, S Itamura, H Nishimura, M Enami, M Tashiro, KF Shortridge, H Kida
    JOURNAL OF VIROLOGY 73 (10) 8303 - 8307 0022-538X 1999/10 [Not refereed][Not invited]
     
    In the influenza H5N1 virus incident in Hong Kong in 1997, viruses that are closely related to H5N1 viruses initially isolated in a severe outbreak of avian influenza in chickens were isolated from humans, signaling the possibility of an incipient pandemic. However, it was not possible to prepare a vaccine against the virus in the conventional embryonated egg system because of the lethality of the virus for chicken embryos and the high level of biosafety therefore required for vaccine production. Alternative approaches, including an avirulent H5N4 virus isolated from a migratory duck as a surrogate virus, H5N1 virus as a reassortant with avian virus H3N1 and an avirulent recombinant H5N1 virus generated by reverse genetics, have been explored. All vaccines were formalin inactivated. Intraperitoneal immunization of mice with each of vaccines elicited the production of hemagglutination-inhibiting and virus-neutralizing antibodies, while intranasal vaccination without adjuvant induced both mucosal and systemic antibody responses that protected the mice from lethal H5N1 virus challenge. Surveillance of birds and animals, particularly aquatic birds, for viruses to provide vaccine strains, especially surrogate viruses, for a future pandemic is stressed.
  • H Aoki, Y Sakoda, K Jukuroki, A Takada, H Kida, A Fukusho
    VETERINARY MICROBIOLOGY 68 (3-4) 197 - 207 0378-1135 1999/08 [Refereed][Not invited]
     
    The glycoprotein gB of pseudorabies virus (PrV) was expressed in various mammalian cells by a recombinant baculovirus carrying the PrV gB gene under the control of the CAG promoter. When the recombinant baculovirus was inoculated into the stable porcine kidney cell line CPK, expression of PrV gB was detected by immunofluorescent antibody analysis and a 155 kDa of protein, which has the same molecular mass as the native PrV gB, was detected by Western blotting. High levels of expression of PrV gB were observed in BHK-21, HmLu-1 and SK-K cell Lines. Furthermore, anti-PrV gB-specific antibodies against PrV gB protein were detected by the enzyme-linked immunosorbent assay in mice inoculated the recombinant baculovirus. The recombinant baculovirus containing the PrV glycoprotein gB gene under the CAG promoter could be a candidate for a pseudorabies vaccine. (C) 1999 Elsevier Science B.V. All rights reserved.
  • Protective effects of intranasal vaccination with plasmid encoding pseudorabies virus glycoprotein B in mice
    A Takada, K Okazaki, H Kida
    JAPANESE JOURNAL OF VETERINARY RESEARCH 47 (1-2) 25 - 33 0047-1917 1999/08 [Refereed][Not invited]
     
    Intranasal administration of plasmid DNA encoding glycoprotein B of pseudorabies virus into mice induced both serum and secretory antibody responses. These mice resisted intranasal challenge with lethal dose of the virus, but did not intraperitoneal challenge. On the other hand, intramuscular injection of the plasmid induced less secretory and higher serum antibody responses than those of intranasally vaccinated mice. None of them was protected from virus challenge. The present results suggest that administration of plasmid DNA encoding glycoprotein B by respiratory mucosal route generates local secretory antibodies which. serve to protect animals from pseudorabies virus infection.
  • Identification of the murine Mx2 gene: Interferon-induced expression of the Mx2 protein from the feral mouse gene confers resistance to vesicular stomatitis virus
    HK Jin, A Takada, Y Kon, O Haller, T Watanabe
    JOURNAL OF VIROLOGY 73 (6) 4925 - 4930 0022-538X 1999/06 [Refereed][Not invited]
     
    The mouse genome contains two related interferon-regulated genes, Mx1 and Mx2, Whereas Mx1 codes for the nuclear 72-kDa protein that interferes with influenza virus replication after interferon treatment, the Mx2 gene is nonfunctional in all laboratory mouse strains examined, since its open reading frame (ORF) is interrupted by an insertional mutation and a subsequent frameshift mutation. In the present study, we demonstrate that Mx2 mRNA of cells from feral mouse strains NJL (Mus musculus musculus) and SPR (Mus spretus) differs from that of the laboratory mouse strains tested. The Mx2 mRNA of the feral strains contains a single long ORF consisting of 656 amino acids. We further show that Mx2 protein in the feral strains is expressed upon interferon treatment and localizes to the cytoplasm much like the rat Mx2 protein, which inhibits vesicular stomatitis virus replication. Furthermore, transfected 3T3 cell lines of laboratory mouse origin expressing Mx2 from feral strains acquire slight resistance to vesicular stomatitis virus.
  • M Miyoshi, Y Ishii, M Takiguchi, A Takada, J Yasuda, A Hashimoto, K Okazaki, H Kida
    JOURNAL OF VETERINARY MEDICAL SCIENCE 61 (4) 375 - 379 0916-7250 1999/04 [Refereed][Not invited]
     
    TO determine the site of latent infection of canine herpesvirus (CHV), tissues from dogs convalescent from acute infection with CHV were examined for the presence of viral genome DNA by the nested polymerase chain reaction. CHV DNA was detected in the trigeminal ganglia and the retropharyngeal lymph nodes. In situ hybridization study of the tissues revealed that CHV genome persisted in the nuclei of ganglionic neurons and lymphocytes.
  • MIYOSHI Masahiro, ISHII Yuki, TAKIGUCHI Mitsuyoshi, TAKADA Ayato, YASUDA Jun, HASHIMOTO Akira, OKAZAKI Katsunori, KIDA Hiroshi
    The Journal of veterinary medical science 社団法人日本獣医学会 61 (4) 375 - 379 0916-7250 1999/04 [Not refereed][Not invited]
     
    To determine the site of latent infection of canine herpesvirus (CHV), tissues from dogs convalescent from acute infection with CHV were examined for the presence of viral genome DNA by the nested polymerase chain reaction. CHV DNA was detected in the trigeminal ganglia and the retropharyngeal lymph nodes. In situ hybridization study of the tissues revealed that CHV genome persisted in the nuclei of ganglionic neurons and lymphocytes.
  • M Imai, A Takada, K Okazaki, H Kida
    JAPANESE JOURNAL OF VETERINARY RESEARCH 46 (4) 171 - 177 0047-1917 1999/02 [Not refereed][Not invited]
     
    The hemagglutinin (HA) of six H5 influenza virus strains isolated from ducks in Japan and China in 1976 to 1996 were analyzed antigenically and genetically. Antigenic analysis using a panel of monoclonal antibodies revealed that the HA of H5 influenza viruses isolated from ducks are antigenically closely related to each other. Phylogenetic analysis indicates that the isolates from ducks in Hokkaido were derived from an ancestor common with the highly pathogenic isolates from chickens and humans in Hong Kong in 1997.
  • Miyoshi M, Takiguchi M, Yasuda J, Hashimoto A, Takada A, Okazaki K, Kida H
    Archives of Virology 144 (2) 407 - 420 0304-8608 1999 [Refereed][Not invited]
  • KF Shortridge, NN Zhou, Y Guan, P Gao, T Ito, Y Kawaoka, S Kodihalli, S Krauss, D Markwell, KG Murti, M Norwood, D Senne, L Sims, A Takada, RG Webster
    VIROLOGY 252 (2) 331 - 342 0042-6822 1998/12 [Refereed][Not invited]
     
    The transmission of avian H5N1 influenza viruses to 18 humans in Hong Kong in 1997 with six deaths established that avian influenza viruses can transmit to and cause lethal infection in humans. This report characterizes the antigenic and biological properties of the H5N1 influenza viruses isolated from chickens, ducks, and geese from farms and poultry markets in Hong Kong during 1997 and compares them with those of virus isolated from the index human case. Each of the H5N1 viruses from Hong Kong poultry markets that were tested were lethal in chickens, possessed polybasic amino acids at the carboxy-terminus of HAI, and by definition were highly pathogenic in poultry. The available nonpathogenic H5 influenza viruses and the pathogenic H5N1 virus from Hong Kong were analyzed with monoclonal antibodies prepared to A/chicken/Pennsylvania/1370/83 (H5N2). The analysis revealed limited antigenic drift in 15 years and established that monoclonal antibodies are useful reagents for identification and antigenic analysis of avian strains that may transmit to humans in the future. One of the monoclonal antibodies permitted separation of the H5N1 influenza viruses from poultry into two groups that correlated with the presence or absence of a carbohydrate at residue 158 adjacent to the receptor binding site on HA. The H5N1 viruses examined replicated in geese, pigs, rats, and mice, but to only a very limited extent in ducks. It is noteworthy that all infected geese shed virus and that the H5N1 viruses caused disease signs and death in a portion (3 of 16) of the geese, with evidence of systemic spread to the brain. The tropism for geese is unusual and may provide insight into the origin of these viruses. In mice, the H5N1 virus caused lethal pneumonia and spread systemically to the brain. Mice would thus provide an ideal model system for studying immune responses and pathogenesis. Transmission experiments in chickens revealed that the H5N1 viruses are spread by fecal-oral transmission rather than by aerosol, and that the viruses are inactivated by drying of feces at ambient temperature. However, infectivity is maintained for at least 4 days in wet feces at 25 degrees C. There were differences in the morphology of the H5N1 viruses isolated from birds and humans. The perpetuation of H5N1 influenza viruses in the poultry markets in Hong Kong and the transmission of these viruses to humans emphasize the importance of these markets in the epidemiology of influenza. The poultry markets are of critical importance in the perpetuation and transmission of influenza viruses to other avian species and to mammals, including humans. (C) 1998 Academic Press.
  • K Ryan-Poirier, Y Suzuki, WJ Bean, D Kobasa, A Takada, T Ito, Y Kawaoka
    VIRUS RESEARCH 56 (2) 169 - 176 0168-1702 1998/08 [Refereed][Not invited]
     
    Influenza A viruses of the H3 subtype caused the 1968 Hong Kong pandemic, the hemagglutinin (HA) gene being introduced into humans following a reassortment event with an avian virus. Receptor specificity and serum inhibitor sensitivity of the HA of influenza A viruses are linked to the host species. Human Hi viruses preferentially recognize N-acetyl sialic acid linked to galactose by alpha 2,6 linkages (Neu5Ac alpha 2,6Gal) and are sensitive to serum inhibitors, whereas avian and equine viruses preferentially recognize Neu5Ac alpha 2,3Gal linkages and are resistant to serum inhibitors. We have examined the receptor specificity and serum inhibitor sensitivity of H3 human influenza A viruses from the time they were introduced into the human population to gain insight into the mechanism of viral molecular evolution and host tropism. All of the viruses were sensitive to neutralization and hemagglutination inhibition by horse serum. Early H3 viruses were resistant to pig and rabbit serum inhibitors. Viruses isolated after 1977 were uniformly sensitive to inhibition by pig and rabbit sera.. The recognition of Neu5Ac alpha 2,3Gal or Neu5Ac alpha 2,6Gal linkages was not correlated with the serum sensitivity. These data showed that the receptor specificity of HA, measured as inhibitor sensitivity, has changed during replication in humans since its introduction from an avian virus. (C) 1998 Published by Elsevier Science B.V. All rights reserved.
  • H Takakuwa, T Ito, A Takada, K Okazaki, H Kida
    ARCHIVES OF VIROLOGY 143 (6) 1129 - 1143 0304-8608 1998 [Refereed][Not invited]
     
    To provide information on the mechanism of attenuation of a Newcastle disease vaccine strain, TCND, we compared it with the parental virulent strain California 11914 (CAL) biologically and genetically. It was found that TCND bore the fusion protein of virulent type, consisting of a pair of dibasic amino acid residues at the cleavage site and was a temperature sensitive (ts) mutant restricted to grow at 41.5 degrees C. Revertants were obtained by prolonged incubation of chicken embryos inoculated with TCND at the nonpermissive temperature. In cultured cells, viral gene transcription and protein synthesis of TCND occurred similarly to those of CAL and the revertants at 41.5 degrees C. Hemadsorption and immunofluorescence assays revealed that cell surface expression of functional hemagglutinin-neuraminidase (HN) of TCND at 41.5 degrees C was lower than that at 35 degrees C. The revertants exhibited lower activity in fusion assay than CAL and recovered virulence to chicken only in part. The results indicate that the ts mutation of TCND in association with the defect of HN glycoprotein transport is a mechanism of the attenuation, and in addition, some other factors such as fusion activity should be involved in the loss of virulence of CAL to chickens.
  • A Takada, C Robison, H Goto, A Sanchez, KG Murti, MA Whitt, Y Kawaoka
    PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 94 (26) 14764 - 14769 0027-8424 1997/12 [Refereed][Not invited]
     
    Ebola virus causes hemorrhagic fever in humans and nonhuman primates, resulting in mortality rates of up to 90%. Studies of this virus have been hampered by its extraordinary pathogenicity, which requires biosafety level 4 containment. To circumvent this problem, we developed a novel complementation system for functional analysis of Ebola virus glycoproteins, It relies on a recombinant vesicular stomatitis virus (VSV) that contains the green fluorescent protein gene instead of the receptor-binding G protein gene (VSV Delta G*). Herein we show that Ebola Reston virus glycoprotein (ResGP) is efficiently incorporated into VSV particles. This recombinant VSV with integrated ResGP (VSV Delta G*-ResGP) infected primate cells more efficiently than any of the other mammalian or avian cells examined, in a manner consistent with the host range tropism of Ebola virus, whereas VSV Delta G* complemented with VSV G protein (VSV Delta G*-G) efficiently infected the majority of the cells tested. We also tested the utility of this system for investigating the cellular receptors for Ebola virus. Chemical modification of cells to alter their surface proteins markedly reduced their susceptibility to VSV Delta G*-ResGP but not to VSV Delta G*-G. These findings suggest that cell surface glycoproteins with N-linked oligosaccharide chains contribute to the entry of Ebola viruses, presumably acting as a specific receptor and/or cofactor for virus entry. Thus, our VSV system should be useful for investigating the functions of glycoproteins from highly pathogenic viruses or those incapable of being cultured in vitro.
  • MR Castrucci, M Hughes, L Calzoletti, Donatelli, I, K Wells, A Takada, Y Kawaoka
    VIROLOGY 238 (1) 128 - 134 0042-6822 1997/11 [Refereed][Not invited]
     
    The M2 protein of influenza A virus functions as an ion channel. It contains three cysteine residues: cysteines 17 and 19, which form disulfide bonds in the ectodomain, and cysteine 50, which is acylated. To understand the role of these cysteine residues in virus replication, we used reverse genetics to create influenza viruses in which the individual cysteines were mutated and a virus in which all three cysteines were changed to serine. The M2 cysteine mutants that lacked either of the cysteine residues in the ectodomain and the mutant that lacked all three residues had appreciably lower amounts of M2 oligomers than did the wild-type virus when examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. None of the mutants, however, were defective in replication, either in vitro or in ferrets and mice; These findings demonstrate that noncovalent interactions are sufficient for the M2 protein to form functional oligomers for virus replication and that its cysteine residues are dispensable for influenza virus replication in vitro and in vivo. (C) 1997 Academic Press.
  • Differences in sialic acid-galactose linkages in the chicken egg amnion and allantois influence human influenza virus receptor specificity and variant selection
    T Ito, Y Suzuki, A Takada, A Kawamoto, K Otsuki, H Masuda, M Yamada, T Suzuki, H Kida, Y Kawaoka
    JOURNAL OF VIROLOGY 71 (4) 3357 - 3362 0022-538X 1997/04 [Refereed][Not invited]
     
    Human influenza viruses are more efficiently isolated by inoculating patient samples into the amniotic rather than the allantoic cavity of embryonated chicken eggs, This type of cultivation selects virus variants with mutations around the hemagglutinin (HA) receptor binding site. To understand the molecular basis of these phenomena, we investigated the abundances of sialic acid (SA) linked to galactose (Gal) by the alpha-2,3 linkage (SA alpha 2,3Gal) and SA alpha 2,6Gal in egg amniotic and allantoic cells and in Madin-Darby canine kidney (MDCK) cells, Using SA-Gal linkage-specific lectins (Maackia amurensis agglutinin specific for SA alpha 2,6Gal and Sambucus nigra agglutinin specific for SA alpha 2,3Gal), we found SA alpha 2,3Gal in both allantoic and amniotic cells and SA alpha 2,6Gal in only the amniotic cells, MDCK; cells contained both linkages, To investigate how this difference in abundances of SA alpha 2,3Gal and SA alpha 2,6Gal in allantoic and amniotic cells affects the appearance of host cell variants in eggs, we determined the receptor specificities and HA amino acid sequences of two different patient viruses which were isolated and passaged in the amnion or in the allantois and which were compared with MDCK cell grown viruses, We found that the viruses maintained high SA alpha 2,6Gal specificities when grown in MDCK cells or following up to two amniotic passages; however, further passages in either the amnion or allantois resulted in the acquisition of, or a complete shift to, SA alpha 2,3Gal specificity, depending on the virus strain. This change in receptor specificity was accompanied by the appearance of variants in the population with Leu-to Gln mutations at position 226 in their HA, These findings suggest that lack of SA alpha 2,6Gal linkages in the allantois of chicken eggs is a selective pressure for the appearance of host cell variants with altered receptor specificities and amino acid changes at position 226.
  • A Takada, H Kida
    VETERINARY MICROBIOLOGY 50 (1-2) 17 - 25 0378-1135 1996/05 [Refereed][Not invited]
     
    Intranasal vaccination of chickens with inactivated Newcastle disease virus (NDV) induced both local and systemic antibody responses, resulting in protection against intranasal challenge with a lethal dose of a virulent NDV strain. The immune response was enhanced by the use of cholera toxin B subunit (CTB) as an adjuvant and only small amounts of the challenge virus were recovered from the birds vaccinated together with CTB. On the other hand, subcutaneous vaccination with the same antigen induced only a serum antibody response in chickens, allowing the challenge virus to replicate in the sinus. The present results indicate that secretory antibodies induced on the respiratory mucosal surface by intranasal vaccination with inactivated NDV protected chickens from lethal infection by inhibiting virus replication at the portal of entry for the virus.
  • T ITO, K OKAZAKI, Y KAWAOKA, A TAKADA, RG WEBSTER, H KIDA
    ARCHIVES OF VIROLOGY 140 (7) 1163 - 1172 0304-8608 1995 [Refereed][Not invited]
     
    To provide information on the mechanism of perpetuation of influenza viruses among waterfowl reservoirs in nature, virological surveillance was carried out in Alaska during their breeding season in summer from 1991 to 1994. Influenza viruses were isolated mainly from fecal samples of dabbling ducks in their nesting places in central Alaska. The numbers of subtypes of 108 influenza virus isolates were 1 H2N3, 37 H3N8, 55 H4N6, 1 H7N3, 1 H8N2, 1 H10N2, 11 H10N7, and H10 N9. influenza viruses were also isolated from water samples of the lakes where they nest. Even in September of 1994 when the most ducks had left for migration to south, viruses were still isolated from the lake water. Phylogenetic analysis of the NP genes of the representative isolates showed that they belong to the North American lineage of avian influenza viruses, suggesting that the majority of the waterfowls breeding in central Alaska migrate to North America and not to Asia. The present results support the notion that influenza viruses have been maintained in waterfowl population by water-borne transmission and revealed the mechanism of year-by-year perpetuation of the viruses in the lakes where they breed.
  • A TAKADA, H KIDA
    ARCHIVES OF VIROLOGY 140 (9) 1629 - 1635 0304-8608 1995 [Refereed][Not invited]
     
    Intranasal vaccination of mice with glycoprotein B (gB) of pseudorabies virus (PRV) induced specific IgA and IgG antibody responses in the secretion of the respiratory tract, resulting in protection of the animals against intranasal challenge with a lethal dose of virulent PRV. The immune response was enhanced by the use of cholera toxin B subunit as an adjuvant. The present results indicate that local vaccination with gB is a promising strategy to confer protective immunity on animals against PRV infection by inducing secretory antibodies on their mucosal surfaces where the primary replication of the virus occurs.
  • ACQUISITION OF PATHOGENICITY OF A NEWCASTLE-DISEASE VIRUS ISOLATED FROM A JAPANESE-QUAIL BY INTRACEREBRAL PASSAGE IN CHICKENS
    MA ISLAM, T ITO, H TAKAKUWA, A TAKADA, C ITAKURA, H KIDA
    JAPANESE JOURNAL OF VETERINARY RESEARCH 42 (3-4) 147 - 156 0047-1917 1994/12 [Refereed][Not invited]
     
    Newcastle disease virus (NDV) was isolated from a Japanese quail (Cotornix cotornix japonica). The effect of intracerebral and intranasal passages of the NDV in chickens on the pathogenicity was studied. Pathogenicity of the viruses of different passages levels was compared with that of the original isolate by the mean death time with the minimum lethal dose in chicken embryos, intracerebral pathogenicity index in day-old chicks, intravenous pathogenicity index with 6-week-old chickens and the mortality rates of chickens and quails inoculated intravenously or intransally. The original isolate from the quail did not kill chickens but only embryos and some one-day-old chicks, exhibiting a mesogenic character. Pathogenicity of the virus of the 10th intranasal passage was not different from that of the original isolate. The viruses passaged intracerebrally, on the other hand, killed chickens of all ages by either route of inoculation, showing a velogenic property. Virus recovery from the blood and the brain was positive only in the chickens infected with brain-passaged viruses by any route of inoculation. Virus titers in the tissues of chickens infected with the brain-passaged viruses were higher than those with the original isolate and the virus of the 10th intranasal passage. These results indicate that the enhanced pathogenicity of the mesogenic NDV isolate from the quail for chickens was induced by acquiring the properties of neurotropism and pantropism through intracerebral passage in chickens.
  • A TAKADA, Y SHIMIZU, H KIDA
    JOURNAL OF VETERINARY MEDICAL SCIENCE 56 (4) 633 - 637 0916-7250 1994/08 [Refereed][Not invited]
     
    Intranasal vaccination of mice with inactivated Aujeszky's disease virus (ADV) induced IgA and IgG antibody responses to the virus in the secretion of the respiratory tract, resulting in complete protection of the animals against intranasal challenge with virulent ADV. The immune response was enhanced by the use of the cholera toxin B subunit (CTB) as an adjuvant. On the other hand, subcutaneous vaccination of mice with inactivated ADV, even together with CTB, scarcely stimulated secretory antibody responses, resulting in only partial protection. The present results suggest that development of a vaccination procedure to stimulate the mucosal immune response should improve the protective effects of the inactivated herpesvirus vaccines, and thereby make it possible to control the infections by prohibiting virus replication at the site where primary infection takes place, as well as inhibiting subsequent latency and reactivation of the virus.
  • Takakuwa H, Ito T, Takada A, Okazaki K, Kida H
    Japanese Journal of Veterinary Research 45 207 - 215 [Refereed][Not invited]

Books etc

Conference Activities & Talks

  • フィロウイルスに対する細胞の感受性とウイルスレセプターの多形  [Invited]
    高田 礼人
    「感染・免疫・がん・炎症」シンポジウム  2019/03
  • Toward the Control of Zoonoses  [Invited]
    TAKADA Ayato
    Nagasaki University WISE Symposium  2019/03
  • Toward the control of viral zoonoses: Our SATREPS activity in Africa  [Invited]
    TAKADA Ayato
    The 59th Annual Meeting for the Japanese Society of Tropical Medicine  2018/11
  • Update Ebola Virus Research  [Invited]
    TAKADA Ayato
    Studium Generale 3, Disease Control of the Wildlife-Livestock-Human Interface: Series  2018/07
  • エボラウイルス研究最前線  [Invited]
    高田 礼人
    第20回日本医療マネジメント学会学術総会  2018/06
  • Recent Topics from Ebola virus Research  [Invited]
    TAKADA Ayato
    Inner Mongolia Agriculture University Seminar  2018/04
  • Filoviruses  [Invited]
    TAKADA Ayato
    第91回日本細菌学会総会 シンポジウム  2018/03
  • エボラウイルス感染症における新知見  [Invited]
    高田 礼人
    第66回日本感染症学会東日本地方会学術集会  2017/11
  • 鳥インフルエンザの基礎とグローバルサーベイランス  [Invited]
    高田 礼人
    テニュアトラック推進機構主催セミナー 鳥インフルエンザと渡り鳥の関わり  2017/10
  • 鳥インフルエンザウイルスHA開裂部位への塩基性アミノ酸挿入機構  [Invited]
    高田 礼人
    第31回インフルエンザ研究者交流の会シンポジウム  2017/06
  • Ebolavirus --Ecology and antiviral strategies--  [Invited]
    TAKADA Ayato
    11th Annual Meeting of Korean Society of Zoonosis  2017/05
  • エボラウイルス研究の最前線  [Invited]
    高田 礼人
    第32回日本環境感染学会総会学術集会 シンポジウム16 エボラウイルス病の最前線  2017/01
  • エボラウイルス研究の最前線  [Invited]
    高田 礼人
    ICD講習会  2016/10
  • Neutralization and Antibody-Dependent Enhancement of Ebolavirus  [Invited]
    TAKADA Ayato
    8th International Global Virus Network Meeting  2016/10
  • グローバルヘルスを支える生化学 エボラウイルスの診断・治療法開発の最前線  [Invited]
    高田 礼人
    第89回日本生化学会大会  2016/09
  • Ebolavirus Entry into Cells --- Neutralization and Antibody-Dependent Enhancement---  [Invited]
    高田 礼人
    The 15th Awaji International Forum on Infection and Immunity  2016/09
  • 人獣共通感染症としてのインフルエンザ  [Invited]
    高田 礼人
    第24回呼吸器疾患・感染症研究会  2016/07
  • 人獣共通感染症の研究最前線 -エボラ出血熱-  [Invited]
    高田 礼人
    日本麻酔科学会第63回学術集会  2016/04
  • エボラウイルス研究の最前線  [Invited]
    高田 礼人
    第90回日本感染症学会総会・学術講演会  2016/04
  • 人獣共通感染症研究最前線 -エボラ出血熱とインフルエンザ-  [Invited]
    高田 礼人
    第45回野依フォーラム例会  2016/04
  • ウイルスの生態  [Invited]
    高田 礼人
    人間文化研究機構広領域連携型基幹研究プロジェクト キックオフ・シンポジウム  2016/03
  • A Monoclonal Antibody Neutralizing All Known Ebola Viruses  [Invited]
    TAKADA Ayato
    2016 US–Japan Annual Medical Biodefense Research Symposium Theme: “Ebola And Emerging Pathogens”  2016/01
  • フィロウイルスの細胞侵入機構  [Invited]
    高田 礼人
    平成27年度 遺伝子病制御研究所研究集会 感染・免疫・炎症・発癌  2015/12
  • Ebolavirus: ecology and antiviral strategies  [Invited]
    TAKADA Ayato
    3rd GRF One Health Summit 2015  2015/10
  • エボラウイルスとは  [Invited]
    高田 礼人
    第15回日本ハ-イオセーフティ学会総会  2015/09
  • エボラウイルス研究の現状と展望  [Invited]
    高田 礼人
    第32回医学情報サービス研究大会  2015/07
  • エボラ出血熱  [Invited]
    高田 礼人
    第62回日本実験動物学会総会  2015/05
  • フィロウイルス研究の現状と展望  [Invited]
    高田 礼人
    第33回川内カンファレンス  2015/04
  • エボラ出血熱:現状と研究の最前線  [Invited]
    高田 礼人
    岐阜大学市民講座  2015/03
  • 人獣共通感染症 特にエボラ出血熱について  [Invited]
    高田 礼人
    平成26年度 鳥取県獣医師会東部支部総会  2015/02
  • エボラウイルスに迫る  [Invited]
    高田 礼人
    北海道大学 創成研究機構第 12 回 創成シンポジウム 感染症研究の最前線 ― エボラ・結核を例に―  2015/02
  • Comparison of antiviral activity between IgA and IgG specific to influenza virus hemagglutinin: Increased potential of IgA for heterosubtypic immunity  [Invited]
    TAKADA Ayato
    "Improving Efficacy of Vaccines for ARI", Acute Respiratory Infections (ARI) Panel Meeting, 17th International Conference on Emerging Infectious Diseases (EID) in the Pacific Rim Emerging Viral Diseases, United States–Japan Cooperative Medical Sciences Pr  2015/01
  • エボラウイルス研究とBSL4  [Invited]
    高田 礼人
    第十二回 日本防護服研究会 学術総会  2015/01
  • エボラおよびマールブルグウイルスによる感染症  [Not invited]
    高田 礼人
    平成26年度新型インフルエンザ等新興・再興感染症研究推進事業シンポジウム  2015/01
  • エボラウイルスの基礎と最新知見  [Invited]
    高田 礼人
    第11回 群馬感染症研究会学術講演会  2014/12
  • エボラウイルス -研究の現状と展望-  [Invited]
    高田 礼人
    第200回 医学セミナー  2014/10
  • Infection and pathogenicity of Ebola virus and Biosafety  [Invited]
    TAKADA Ayato
    Laboratory Biosafety and Infection Control of Emerging Infectious Diseases  2014/09
  • 北海道大学ザンビア拠点での取り組み  [Invited]
    高田 礼人
    J-GRID市民公開講演会「いま話題の感染症-SFTS、MERS、エボラ出血熱」  2014/08
  • エボラ出血熱とBSL-4  [Invited]
    高田 礼人
    長崎大学熱帯医学研究所 「市民公開特別講座」  2014/08
  • ホットゾーン的ウイルス研究とバイオセーフティーレベル4  [Invited]
    高田 礼人
    第8回細菌学若手コロッセウム  2014/08
  • Comparison of antiviral activity between IgA and IgG  [Not invited]
    TAKADA Ayato
    XVI International Congress of Virology  2014/07
  • 鳥インフルエンザの基礎  [Not invited]
    高田 礼人
    第12回インフルエンザ夏季セミナー  2014/07
  • フィロウイルス感染症に対する防御免疫における抗体の役割  [Invited]
    高田 礼人
    第79回インターフェロン・サイトカイン学会  2014/06
  • Role of antibodies in protective immunity against filovirus infection  [Invited]
    TAKADA Ayato
    6th International Symposium on Filoviruses  2014/04
  • H7N9株やH5N1株を含めた新たなインフルエンザパンデミックの可能性について  [Invited]
    高田 礼人
    化血研KIKUCHIバイオセミナー「三風会」  2014/01
  • 鳥インフルエンザ  [Invited]
    高田 礼人
    平成25年度家畜保健衛生所病性鑑定技術検討会  2013/12
  • フィロウイルス感染症  [Invited]
    高田 礼人
    第61回日本ウイルス学会学術集会  2013/11
  • Ecology of Avian Influenza Viruses: Topics from surveillance in Asia and Africa  [Invited]
    TAKADA Ayato
    1st Kyoto International Symposium on Virus-Host Coevolution  2013/11
  • フィロウイルスの病原性と宿主域  [Invited]
    高田 礼人
    市民講座 やさしく学ぶ話題の感染症 -動物から人へうつる新たな病気-  2013/10
  • Recent advances in filovirus research: Epidemiology and antiviral strategies  [Invited]
    TAKADA Ayato
    The 12th Awaji International Forum on Infection and Immunity  2013/08
  • 「エボラ出血熱ウイルス」…糸のような形の、やさしそうで怖いウイルスの話  [Invited]
    高田 礼人
    みちのくウイルス塾  2013/07
  • Recent Topics of Filovirus Research  [Invited]
    TAKADA Ayato
    International Conference in Medicine and Public Health 2013 (ICMPH2013) “Healthy Society beyond Frontiers”  2013/06
  • Heterosubtypic antiviral activity of influenza virus hemagglutinin-specific antibodies  [Not invited]
    TAKADA Ayato
    Fifteenth International Conference on Negative Strand Viruses  2013/06
  • Activities of Hokkaido University Research Center for Zoonosis Control  [Invited]
    TAKADA Ayato
    Center of Excellence for Emerging and Zoonotic Animal Diseases (CEEZAD) 2013 Annual Meeting  2013/04
  • Serological Survey of Antibodies to Filoviruses  [Invited]
    TAKADA Ayato
    6th U.S. - Japan Medical Biodefense Research Symposium, “New Frontiers in Medical Biodefense Research Between the United States and Japan”  2013/02
  • Serological survey of antibodies to filoviruses in wild animals  [Invited]
    TAKADA Ayato
    Asian-African Research Forum on Emerging and Reemerging Infections 2013  2013/01
  • 海外のBSL4 施設での実験の状況  [Invited]
    高田 礼人
    日本学術会議 公開シンポジウム  2012/12
  • エボラおよびマールブルグウイルス  [Invited]
    高田 礼人
    第60回日本ウイルス学会学術集会 シンポジウム1 人獣共通感染症  2012/11
  • 人獣共通感染症としてのインフルエンザ  [Invited]
    高田 礼人
    第21回先端科学移動大学2012  2012/11
  • H5N1高病原性鳥インフルエンザウイルスと私との関わり  [Invited]
    高田 礼人
    第2回宮崎大学 鳥インフルエンザシンポジウム  2012/10
  • Serological evidence of filovirus infection in Indonesian orangutans  [Invited]
    TAKADA Ayato
    46th Joint Working Conference on Viral Diseases US-Japan Cooperative, Medical Science Program  2012/06
  • Ebola and Marburg viruses  [Invited]
    TAKADA Ayato
    4th International Symposium of Animal Global Health  2012/05
  • ウイルス感染と粘膜免疫  [Invited]
    高田 礼人
    日本薬学会第132年会 一般シンポジウムS29 鼻腔内投与による脳機能治療の可能性  2012/03
  • Ebola and Marburg viruses  [Invited]
    TAKADA Ayato
    Nagasaki Symposium on Emerging Viral Diseases  2012/02
  • エボラおよびマールブルグウイルス  [Invited]
    高田 礼人
    第7回霊長類医科学フォーラム  2011/11
  • Influenza and Ebola viruses “Pathogenicity and Host Range”  [Invited]
    TAKADA Ayato
    ヤンセンファーマ株式会社 社内勉強会  2011/09
  • Antibody-dependent enhancement of Marburg virus infection  [Invited]
    TAKADA Ayato
    45th Joint Working Conference on Viral Diseases US-Japan Cooperative, Medical Science Program  2011/06
  • フィロウイルスの細胞侵入機構  [Invited]
    高田 礼人
    第58回日本ウイルス学会学術集会 シンポジウム10 ウイルスの細胞侵入  2010/11
  • 人獣共通感染症としてのインフルエンザ  [Invited]
    高田 礼人
    第19回先端科学移動大学2010  2010/11
  • 「人獣共通感染症」ウイルスはどうやって行きのびているのか  [Invited]
    高田 礼人
    長崎大学熱帯医学研究所市民公開特別講座  2010/10
  • ウイルスの病原性と宿主域-人獣共通感染症としてのインフルエンザ  [Invited]
    高田 礼人
    第75回日本民族衛生学会  2010/09
  • 人獣共通感染症としてのインフルエンザ  [Invited]
    高田 礼人
    第84回日本数理生物学会大会  2010/09
  • Enzyme-linked immunosorbent assay for the detection of filovirus species-specific antibodies  [Invited]
    TAKADA Ayato
    44th Joint Working Conference on Viral Diseases US-Japan Cooperative, Medical Science Program  2010/06
  • インフルエンザウイルスヘマグルチニン亜型間交差反応性抗体の解析  [Invited]
    高田 礼人
    第10回日本蛋白質科学会年会 ウィルス感染の分子機構とその制御  2010/06
  • インフルエンザウイルスの抗原変異予測と亜型間交差感染防制  [Invited]
    高田 礼人
    日本薬学会第13年回 一般シンポジウムS49 インフルエンザの制圧に向けた新しい挑戦  2010/03
  • エボラウイルスとインフルエンザウイルスを追いかけて  [Invited]
    高田 礼人
    公開シンポジウム 感染症の国際疫学研究の最前線―日本をとりまく感染症の現状―  2009/12
  • Mechanisms of filovirus entry  [Invited]
    TAKADA Ayato
    Workshop Taiwan-Japan "Host-Pathogen Interaction"  2009/11
  • 人獣共通感染症としてのインフルエンザ  [Invited]
    高田 礼人
    第4回抗菌化学療法研究会  2009/11
  • 人獣共通感染症としてのインフルエンザ  [Invited]
    高田 礼人
    第47回全国大学保健管理研究集会  2009/09
  • Different efficiency of C-type lectin-mediated entry between filoviruses  [Invited]
    TAKADA Ayato
    43rd Joint Working Conference on Viral Diseases, US-Japan Cooperative Medical Science Program  2009/07
  • 人獣共通感染症の先回り戦略  [Invited]
    高田 礼人
    第55回日本ウイルス学会学術集会  2008/10
  • インフルエンザウイルスの病原性と宿主域  [Invited]
    高田 礼人
    第11回北海道ウイルス感染症セミナーの会  2008/09
  • インフルエンザウイルスの病原性と宿主域  [Invited]
    高田 礼人
    自衛隊北部防衛衛生学会  2008/01
  • Mission of Research Center for Zoonosis Control  [Invited]
    TAKADA Ayato
    日本の人獣共通感染症研究動向=AIとBSE=  2007/10
  • エボラウイルスの病原性発現の分子基盤  [Invited]
    高田 礼人
    第27回日本医学会総会  2007/04
  • Epitopes required for antibody-dependent enhancement of Ebola virus infection  [Not invited]
    Takada A, Ebihara H, Feldmann H, Kawaoka Y
    FILOVIRUSES:Recent Advances and Future Challenges  2006/09
  • エボラウイルスの感染、免疫、疫学  [Not invited]
    高田 礼人
    人獣共通感染症 教育公開フォーラム  2006/07
  • Mechanisms of filovirus entry into cells  [Not invited]
    Ayato Takada, Heinz Feldmann, Yoshihiro Kawaoka
    XIII International Conference on Negative Strand Viruses  2006/06
  • インフルエンザという人獣共通感染症  [Invited]
    TAKADA Ayato
    International Symposium in Fukuoka - Coming Health Emergency: How Nurses Respond to Unknown Emerging Diseases  2006/03
  • 平成17年度杉浦奨励賞受賞講演 「エボラウイルス表面糖蛋白質の機能解析」  [Invited]
    高田 礼人
    第53回日本ウイルス学会総会  2005/11
  • Two different routes of antibody-dependent enhancement of Ebola virus infection  [Invited]
    TAKADA Ayato
    German-Japanese Symposium on Emerging and Reemerging Viruses  2005/05
  • エボラウイルスの抗体依存性感染増強現象とエピトープ  [Not invited]
    高田礼人, 海老原秀喜, 河岡義裕
    第52回日本ウイルス学会総会  2004/11
  • エボラウイルスの感染初期課程:抗体およびレクチンによる感染増強現象  [Not invited]
    高田礼人, 河岡義裕
    第51回日本ウイルス学会総会  2003/10
  • エボラウイルスに対する中和抗体のエピトープの同定と受動免疫の感染防御効果  [Not invited]
    高田礼人, Heinz Feldmann, 河岡義裕
    第50回日本ウイルス学会総会  2002/10
  • Antibody-dependent enhancement of Ebola virus infection  [Invited]
    TAKADA Ayato
    Awaji International Forum on Infection and Immunity  2002/08
  • エボラウイルスにおける抗体依存性感染増強現象  [Not invited]
    高田礼人, Heinz Feldmann, 渡辺真治, 河岡義裕
    第49回日本ウイルス学会総会  2001/10
  • エボラウイルス表面糖蛋白の発現による細胞変性効果とインテグリンの発現抑制  [Not invited]
    高田礼人, 岡崎克則, 喜田宏, 渡辺真治, 伊藤啓史, 河岡義裕
    第48回日本ウイルス学会総会  2000/10
  • エボラウイルス表面糖蛋白もつG蛋白欠損VSVの作出と応用  [Not invited]
    高田礼人, Anthony Sanchez, Michael A. Whitt, 喜田宏, 岡崎克則, 河岡義裕
    第45回日本ウイルス学会総会  1997/09
  • エボラウイルス表面糖蛋白の機能解析 プラスミドによる発現とG蛋白欠失VSVの利用  [Not invited]
    高田礼人, SANCHEZ A, WHITT M A, 喜田宏, 岡崎克則, 河岡義裕
    第124回日本獣医学会学術集会講演要旨集  1997
  • オーエスキー病ウイルスgII蛋白の点鼻接種によるマウスの感染防御  [Not invited]
    高田礼人, 喜田宏
    第119回日本獣医学会学術集会  1995
  • 不活化ウイルスの鼻腔内接種によって誘導される抗体応答およびその感染防御効果  [Not invited]
    高田礼人, 伊藤寿啓, 清水悠紀臣, 喜田宏
    第118回日本獣医学会学術集会  1994

MISC

  • 古山若呼, 高田礼人  最新医学  74-  (4)  530‐538  2019/04/10  [Not refereed][Not invited]
  • 感染症 現状の問題点と未来への展望 エボラ出血熱
    高田 礼人  臨床と微生物  45-  (2)  165  -169  2019/03  [Not refereed][Invited]
  • 岡田愛, 鍬田龍星, 下田宙, 村上晋, 堀本泰介, 宇根有美, 相馬武久, 高田礼人, 前田健, 鈴木和男  日本獣医学会学術集会講演要旨集  161st-  346  2018/08/21  [Not refereed][Not invited]
  • 佐野豊, 伊藤直人, 伊藤直人, 西山祥子, 西山祥子, 岡田和真, 高橋龍樹, 岡崎克則, 高田礼人, 迫田義博, 小澤真, 正谷達謄, 杉山誠, 杉山誠  日本獣医学会学術集会講演要旨集  161st-  396  2018/08/21  [Not refereed][Not invited]
  • 高田礼人  日本医療マネジメント学会雑誌  19-  (Supplement)  100  2018/05/07  [Not refereed][Not invited]
  • 高田礼人  臨床と微生物  45-  (2)  165‐169  2018/03/25  [Not refereed][Not invited]
  • 高田礼人  小児科臨床  70-  2308‐2317  2017/12/20  [Not refereed][Not invited]
  • エボラ出血熱
    高田 礼人  小児科臨床  70-  (増刊号)  2308  -2317  2017/12  [Not refereed][Invited]
  • 高田礼人  日本内科学会雑誌  106-  (10)  2237‐2245  2017/10/10  [Not refereed][Not invited]
  • 高田礼人  日本感染症学会東日本地方会学術集会・日本化学療法学会東日本支部総会合同学会プログラム・抄録集  66th-64th-  70  2017/09  [Not refereed][Not invited]
  • 小林知也, 丸山隼輝, 松郷宙倫, 前田健, 高田礼人, 村上晋, 堀本泰介  日本獣医学会学術集会講演要旨集  160th-  378  2017/08/30  [Not refereed][Not invited]
  • Influenza A Virus M2 Protein: Roles from Ingress to Egress
    TAKADA Ayato  Int J Mol Sci  18-  (12)  E2649  2017/07  [Refereed][Invited]
  • 古山若呼, 吉田玲子, 五十嵐学, 高田礼人  化学療法の領域  33-  (5)  1098‐1105  2017/04/25  [Not refereed][Not invited]
  • 高田 礼人  最新醫學 = The medical frontline  72-  (4)  534  -540  2017/04  [Not refereed][Not invited]
  • 梶原将大, 高田礼人, 高田礼人  臨床とウイルス  45-  (1)  32‐40  2017/03/31  [Not refereed][Not invited]
  • 高田礼人  月刊臨床と研究  94-  (3)  384  2017/03/20  [Not refereed][Not invited]
  • 丸山隼輝, 高田礼人  医学のあゆみ  260-  (6)  491‐497  2017/02/11  [Not refereed][Not invited]
  • 高田礼人  日本臨床  74-  (12)  2080‐2085  2016/12/01  [Not refereed][Not invited]
  • 高田礼人  医学のあゆみ  258-  (7/8)  803‐810  -810  2016/08/20  [Not refereed][Not invited]
  • 古山若呼, 高田礼人  ウイルス  66-  (1)  63‐72  2016/06  [Not refereed][Not invited]
  • 高田礼人  感染症学雑誌  90-  125  2016/03/20  [Not refereed][Not invited]
  • 高田礼人  日本生化学会大会(Web)  89th-  ROMBUNNO.1F05‐2 (WEB ONLY)  2016  [Not refereed][Not invited]
  • エボラ出血熱 -研究の現状と展望-
    高田 礼人  學士會会報  915-  89  -94  2015/11  [Not refereed][Invited]
  • KAJIHARA MASAHIRO, TAKADA AYATO, TAKADA AYATO, TAKADA AYATO  生体の科学  66-  (4)  296  -300  2015/08/15  [Not refereed][Not invited]
  • 梶原 将大, 高田 礼人  生体の科学  66-  (4)  296  -300  2015/07  [Not refereed][Not invited]
  • TAKADA AYATO  ウイルス  65-  (1)  61  -70  2015/06  [Not refereed][Not invited]
  • TAKADA AYATO  いつでも元気  (283)  16  -19  2015/05/01  [Not refereed][Not invited]
  • 高田 礼人  医学のあゆみ  253-  (1)  5  -11  2015/04/04  [Not refereed][Not invited]
  • TAKADA AYATO  医学のあゆみ  253-  (1)  5  -11  2015/04/04  [Not refereed][Invited]
  • TAKADA AYATO  アニムス  20-  (2)  19  -22  2015/04/01  [Not refereed][Invited]
  • TAKADA AYATO  生活と環境  60-  (3)  11  -15  2015/03/01  [Not refereed][Invited]
  • 高田 礼人  生活と環境  60-  (3)  11  -15  2015/03  [Not refereed][Invited]
  • Host cell factors involved in filovirus infection
    TAKADA Ayato  Current Tropical Medicine Reports  2-  30  -40  2015/02  [Refereed][Invited]
  • TAKADA Ayato  Uirusu  65-  (1)  61  -70  2015  [Not refereed][Not invited]
     
    Filoviruses (Ebola and Marburg viruses) cause severe hemorrhagic fever in humans and nonhuman primates. No effective prophylaxis or treatment for filovirus diseases is yet commercially available. The recent outbreak of Ebola virus disease in West Africa has accelerated efforts to develop anti-Ebola virus prophylaxis and treatment, and unapproved drugs were indeed used for the treatment of patients during the outbreak. This article reviews previous researches and the latest topics on vaccine and therapy for Ebola virus disease.
  • Ayato Takada  FUTURE VIROLOGY  10-  (5)  491  -496  2015  [Refereed][Invited]
     
    The H5N1 highly pathogenic avian influenza (HPAI) virus has been reported to infect humans and posing a significant pandemic threat. Although neuraminidase inhibitors are currently available for the treatment of human influenza, alternative strategies need to be developed for the treatment of H5N1 HPAI virus infection in humans due to the appearance of drug-resistant viruses. Recently, passive immunization with virus-specific monoclonal antibodies has been tested for H5N1 HPAI virus infection in animal models, providing evidence that antibody therapy may be a promising option for prophylaxis or treatment of this infectious disease. Here we discuss potential benefits and limitations of antibody therapy in clinical cases of H5N1 virus infection in humans.
  • TAKADA AYATO  現代化学  (523)  16  -18  2014/10/01  [Not refereed][Invited]
  • 髙田 礼人  現代化学  (523)  16  -18  2014/10  [Not refereed][Invited]
  • Katendi Changula, Masahiro Kajihara, Aaron S. Mweene, Ayato Takada  MICROBIOLOGY AND IMMUNOLOGY  58-  (9)  483  -491  2014/09  [Refereed][Not invited]
     
    Filoviral hemorrhagic fever (FHF) is caused by ebolaviruses and marburgviruses, which both belong to the family Filoviridae. Egyptian fruit bats (Rousettus aegyptiacus) are the most likely natural reservoir for marburgviruses and entry into caves and mines that they stay in has often been associated with outbreaks of MVD. On the other hand, the natural reservoir for ebola viruses remains elusive; however, handling of wild animal carcasses has been associated with some outbreaks of EVD. In the last two decades, there has been an increase in the incidence of FHF outbreaks in Africa, some being caused by a newly found virus and some occurring in previously unaffected areas such as Guinea, Liberia and Sierra Leone, in which the most recent EVD outbreak occurred in 2014. Indeed, the predicted geographic distribution of filoviruses and their potential reservoirs in Africa includes many countries in which FHF has not been reported. To minimize the risk of virus dissemination in previously unaffected areas, there is a need for increased investment in health infrastructure in African countries, policies to facilitate collaboration between health authorities from different countries, implementation of outbreak control measures by relevant multi-disciplinary teams and education of the populations at risk.
  • ONISHI NAOMI, HIGASHI HIDEAKI, TAKADA AYATO  最新医学  69-  (4)  865  -871  2014/04  [Not refereed][Not invited]
  • 大西 なおみ, 東 秀明, 高田 礼人  最新医学  69-  (4)  865  -871  2014/04  [Not refereed][Invited]
  • ISODA NORIKAZU, TAKADA AYATO  Mebio  30-  (12)  32  -41  2013/12/10  [Not refereed][Not invited]
  • KAJIHARA MASAHIRO, OGAWA HIROHITO, TAKADA AYATO  実験医学  31-  (19)  3054  -3060  2013/12/01  [Not refereed][Invited]
  • Takada A  Immunotherapy  5-  (5)  441  -443  2013/05  [Refereed][Invited]
  • TAKADA AYATO  ウイルス  62-  (2)  197  -208  2012/12  [Not refereed][Invited]
  • TAKADA Ayato  VIRUS  62-  (2)  197  -208  2012/12/01  [Not refereed][Invited]
     
    Filoviruses (Ebola and Marburg viruses) cause severe hemorrhagic fever in humans and nonhuman primates. No effective prophylaxis or treatment for filovirus diseases is yet commercially available. Recent studies have advanced our knowledge of filovirus protein functions and interaction between viral and host factors in the replication cycle. Current findings on the ecology of filoviruses (i.e., natural infection of nonprimate animals and discovery of a new member of filoviruses in Europe) have also provided new insights into the epidemiology of Ebola and Marburg hemorrhagic fever. This article reviews the fundamental aspects of filovirus biology and the latest topics on filovirus research.
  • 伊藤 公人, 高田 礼人  遺伝 : 生物の科学  66-  (4)  358  -364  2012/07  [Not refereed][Invited]
  • 伊藤公人, 高田礼人  生物の科学 遺伝  66-  (4)  358-364  -364  2012/07  [Not refereed][Invited]
  • 髙田 礼人  最新医学  66-  (12)  2668  -2675  2011/12  [Not refereed][Invited]
  • 高田礼人  最新医学  66-  (12)  2668-2675  -2675  2011/12  [Not refereed][Invited]
  • FUJIHIRA HARUHIKO, USAMI KATSUAKI, DENDA KAORI, YAMADA KEITA, MATSUNO KEITA, SHINOHARA YASURO, TAKADA AYATO, KAKEHI KAZUAKI, IRIMURA TATSURO  生化学  ROMBUNNO.3T8A-14  2011  [Not refereed][Not invited]
  • Ebola and Marburg viruses.
    Nakayama E, Takada A  Journal of Disaster Reseach  6-  (4)  381  -389  2011  [Refereed][Invited]
  • TAKADA AYATO  感染症  236-  (6)  220  -225  2010/11  [Not refereed][Invited]
  • 高田 礼人  Japanese journal of clinical medicine  68-  (9)  1625  -1630  2010/09  [Not refereed][Invited]
  • 高田礼人  臨床と微生物  37-  (2)  125-131  -131  2010/03  [Not refereed][Invited]
  • TAKADA AYATO  Campus Health  47-  (1)  67  -71  2010/02  [Not refereed][Not invited]
  • TAKADA AYATO  化学  64-  (11)  18  -24  2009/11  [Not refereed][Invited]
  • 伊藤公人, 高田礼人  Virus Rep  6-  (2)  60-68  -68  2009/11  [Not refereed][Invited]
  • 高田 礼人, 斉藤 勝司  メディカルバイオ  6-  (6)  6  -8  2009/11  [Not refereed][Invited]
  • Edgar Simulundu, Aaron S. Mweene, Daisuke Tomabechi, Bernard M. Hang'ombe, Akihiro Ishii, Yuka Suzuki, Ichiro Nakamura, Hirofumi Sawa, Chihiro Sugimoto, Kimihito Ito, Hiroshi Kida, Lewis Saiwana, Ayato Takada  ARCHIVES OF VIROLOGY  154-  (9)  1517  -1522  2009/09  [Not refereed][Not invited]
     
    We characterized an influenza virus isolated from a great white pelican in Zambia. Phylogenetic analysis showed that all of its gene segments belonged to the Eurasian lineage and that they appear to have evolved in distinct geographical regions in Europe, Asia, and Africa, suggesting reassortment of virus genes maintained in wild aquatic birds whose flyways overlap across these continents. It is notable that this virus might possess some genes of the same origin as those of highly pathogenic H7 and H5 viruses isolated in Eurasia. The present study underscores the need for continued monitoring of avian influenza viruses in Eurasia and Africa.
  • 高田礼人  蛋白質 核酸 酵素  54-  (8)  913-919  -919  2009/06/10  [Not refereed][Invited]
  • 高田 礼人  Protein, nucleic acid and enzyme  54-  (8)  913  -919  2009/06  [Not refereed][Invited]
  • Reiko Yoshida, Manabu Igarashi, Hiroichi Ozaki, Noriko Kishida, Daisuke Tomabechi, Hiroshi Kida, Kimihito Ito, Ayato Takada  PLOS PATHOGENS  5-  (3)  e1000350  2009/03  [Not refereed][Not invited]
     
    The hemagglutinin (HA) of influenza A viruses has been classified into sixteen distinct subtypes (H1-H16) to date. The HA subtypes of influenza A viruses are principally defined as serotypes determined by neutralization or hemagglutination inhibition tests using polyclonal antisera to the respective HA subtypes, which have little cross-reactivity to the other HA subtypes. Thus, it is generally believed that the neutralizing antibodies are not broadly cross-reactive among HA subtypes. In this study, we generated a novel monoclonal antibody (MAb) specific to HA, designated MAb S139/1, which showed heterosubtypic cross-reactive neutralization and hemagglutination inhibition of influenza A viruses. This MAb was found to have broad reactivity to many other viruses (H1, H2, H3, H5, H9, and H13 subtypes) in enzyme-linked immunosorbent assays. We further found that MAb S139/1 showed neutralization and hemagglutination-inhibition activities against particular strains of H1, H2, H3, and H13 subtypes of influenza A viruses. Mutant viruses that escaped neutralization by MAb S139/1 were selected from the A/Aichi/2/68 (H3N2), A/Adachi/2/57 (H2N2), and A/WSN/33 (H1N1) strains, and sequence analysis of the HA genes of these escape mutants revealed amino acid substitutions at positions 156, 158, and 193 (H3 numbering). A molecular modeling study showed that these amino acids were located on the globular head of the HA and formed a novel conformational epitope adjacent to the receptor-binding domain of HA. Furthermore, passive immunization of mice with MAb S139/1 provided heterosubtypic protection. These results demonstrate that MAb S139/1 binds to a common antigenic site shared among a variety of HA subtypes and neutralizes viral infectivity in vitro and in vivo by affecting viral attachment to cells. The present study supports the notion that cross-reactive antibodies play some roles in heterosubtypic immunity against influenza A virus infection, and underscores the potential therapeutic utility of cross-reactive antibodies against influenza.
  • 高田礼人  防疫上緊急を要するウイルス性出血熱等に対する病原体診断法の確立及び予防・治療法の開発に関する研究 平成20年度 総括・分担研究報告書  27  -30  2009  [Not refereed][Not invited]
  • 伊藤 公人, 高田 礼人  Virus report  6-  (2)  60  -68  2009  [Not refereed][Invited]
  • 将来のインフルエンザ対策
    TAKADA Ayato  BIO Clinica  24-  (9)  13  -13  2009  [Not refereed][Invited]
  • 高田 礼人  綜合臨床  57-  (11)  2673  -2679  2008/11  [Not refereed][Invited]
  • 高田礼人  綜合臨床  57-  (11)  2673-2679  -2679  2008/11/01  [Not refereed][Not invited]
  • 松野啓太, 岸田典子, 宇佐美克明, 入村達郎, 下島昌幸, 河岡義裕, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  56th-  162  2008/10/01  [Not refereed][Not invited]
  • 村上晋, 岩佐彩香, 岩附(堀本)研子, 伊藤睦美, 木曽真紀, 喜田宏, 高田礼人, NIDOM Chairul A, LE QUYNH Mai, 山田晋弥, 今井博貴, 坂井(田川)優子, 河岡義裕, 堀本泰介  日本ウイルス学会学術集会プログラム・抄録集  56th-  224  2008/10/01  [Not refereed][Not invited]
  • 五十嵐学, 伊藤公人, 喜田宏, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  56th-  144  2008/10/01  [Not refereed][Not invited]
  • 岡松正敏, 迫田義博, 吉田裕美, 田中智久, 津田祥美, 磯田典和, 中山絵里, 苫米地大輔, 松野啓太, 梅村孝司, 高田礼人, 喜田宏  日本ウイルス学会学術集会プログラム・抄録集  56th-  263  2008/10/01  [Not refereed][Not invited]
  • 塩崎拓也, 岩井淳, 河岡義裕, 高田礼人, 喜田宏, 宮崎忠昭  日本ウイルス学会学術集会プログラム・抄録集  56th-  379  2008/10/01  [Not refereed][Not invited]
  • 岡松正敏, 迫田義博, 吉田裕美, 田中智久, 津田祥美, 磯田典和, 中山絵里, 苫米地大輔, 松野啓太, 梅村孝司, 高田礼人, 喜田宏  日本獣医学会学術集会講演要旨集  146th-  193  2008/09/05  [Not refereed][Not invited]
  • 高田礼人  補体シンポジウム講演集  45th-  51  2008/07  [Not refereed][Not invited]
  • Igarashi, M, Ito, K, Kida, H, Takada, A  Virology  376-  (2)  323  -329  2008  [Not refereed][Not invited]
  • YOSHIDA REIKO, TAKADA AYATO  実験医学  25-  (20)  3176  -3182  2007/12  [Not refereed][Invited]
  • Ayato Takada, Hideki Ebihara, Heinz Feldmann, Thomas W. Geisbert, Yoshihiro Kawaoka  JOURNAL OF INFECTIOUS DISEASES  196-  S347  -S356  2007/11  [Not refereed][Not invited]
     
    We have shown that antibody-dependent enhancement (ADE) of infection with Zaire Ebola virus (ZEBOV) is mediated by interaction of virus-specific antibodies with Fc receptors or complement component C1q and its receptors in vitro. ADE activities of the antisera to the viral glycoprotein (GP) were virus species specific and were primarily correlated with immunoglobulin (Ig) G2a and IgM levels but not with IgG1 levels. Interestingly, compared with ZEBOV, Reston Ebola virus (REBOV) had substantially weaker potential to induce ADE antibodies. Using monoclonal antibodies, we identified ZEBOV-specific ADE epitopes. To confirm epitope specificity, we constructed a chimeric ZEBOV GP, the ADE epitopes of which were replaced with the corresponding regions of REBOV GP. We found that mouse antisera to the chimeric ZEBOV GP showed less potential to induce ADE activity than did mouse antisera to wild-type ZEBOV GP, although they retained neutralizing activity. These data suggest that GP lacking the ADE-inducing epitopes may increase the potential of GP as a vaccine antigen.
  • Keita Matsuno, Ayato Takada  FUTURE VIROLOGY  2-  (6)  607  -614  2007/11  [Refereed][Invited]
     
    Ebola virus causes lethal hemorrhagic fever in human and nonhuman primates. Effective prophylaxis and treatment for this disease are not yet available. Antisera and monoclonal antibodies specific to Ebola virus proteins have been tested for passive immunization in experimental animal models and clinical cases, and shown to be effective in mice and guinea pigs, whereas the evidence of protective efficacy in primates, including humans, remains elusive. In this review, we focus on research relevant to prophylaxis and treatment by passive immunization, and discuss the potential use of antibody therapy for Ebola virus infection. Nevertheless, there is no doubt that a comprehensive understanding of Ebola virus pathogenesis will aid in the development of therapeutic strategies against Ebola hemorrhagic fever.
  • 高田 礼人  Protein, nucleic acid and enzyme  52-  (10)  1242  -1247  2007/08  [Not refereed][Not invited]
  • 高田礼人  蛋白質 核酸 酵素  52-  (10)  1242-1247  -1247  2007/08  [Not refereed][Invited]
  • Ayato Takada, Hideki Ebihara, Sueven Jones, Heinz Feldmann, Yoshihiro Kawaoka  VACCINE  25-  (6)  993  -999  2007/01  [Not refereed][Not invited]
     
    Ebola virus causes lethal hemorrhagic fever in humans and nonhuman primates, but no effective antiviral compounds are available for the treatment of this infection. The surface glycoprotein (GP) of Ebola virus is an important target of neutralizing antibodies. Although passive transfer of GP-specific antibodies has been evaluated in mouse and guinea. pig models, protection was achieved only by treatment shortly before or after virus challenge. Using these animal models, we evaluated the protective efficacy of two monoclonal antibodies whose epitopes are distinct from those of the antibodies tested by others. Treatment of mice with these antibodies 2 days after challenge completely protected most of the animals; even treatment 3 or 4 days after challenge was partially effective. Although antibody treatment in the guinea pig model was not as effective as in the mouse model, single-dose treatment of guinea pigs I day before, or I or 2 days after challenge did protect some animals. Interestingly, the protective effects seen in these animal models did not correlate with the in vitro neutralizing activity of the antibodies, suggesting different mechanisms of the neutralization by these antibodies. These results underscore the potential therapeutic utility of monoclonal antibodies for postexposure treatment of Ebola virus infections. (c) 2006 Elsevier Ltd. All rights reserved.
  • Yamaguchi Satoru, Iwamoto Katsushi, Takada Ayato  Preprints of the Annual Conference, Japanese Society of Snow and Ice  2007-  (0)  157  -157  2007  [Not refereed][Not invited]
  • インフルエンザに対する感染防御免疫
    吉田 玲子, 高田 礼人  実験医学  25-  (20)  98  -104  2007  [Not refereed][Invited]
  • 谷口 剛, 伊藤公人, 五十嵐 学, 村上 悌治, 高田 礼人, 原口 誠  情報処理学会研究報告バイオ情報学(BIO)  2006-  (135)  185  -192  2006/12/22  [Not refereed][Not invited]
     
    インフルエンザウイルスにおける抗原変異の規則性を発見するために,アミノ酸残基の共変異を解析する共変異とは,タンパク質を構成するアミノ酸残基のうち〆複数の位置のアミノ酸が共に置換する現象である.従来からアミノ酸残基の共変異を解析する手法がいくつか提案されていたが,それらの手法では進化の過程における分岐や時間的関係が考慮されていなかったそこで,これらの問題を解決するために,進化系統解析によって得られる系統樹を利用する手法を提案する.過去40年間のH3N2亜型インフルエンザウイルスのHAタンパク質を対象とし,共変異の検出を行い,その結果を示す,また,共変異は時代と共に変化するため,共変異の変化を検出するための手法を提案する.The influenza viruses undergo antigenic drift to escape from antibody-mediated immune pressure. In order to predict possible structural changes of their molecules in future, it is important to analyze the patterns of amino acid substitutions in the past. In this paper, we present a new method to extract the sets of residue positions which were involved in correlated mutations. We also discuss a method to detect changes of correlation among co-evolving residues.
  • 谷口 剛, 伊藤公人, 五十嵐 学, 村上 悌治, 高田 礼人, 原口 誠  情報処理学会研究報告数理モデル化と問題解決(MPS)  2006-  (135)  185  -192  2006/12/22  [Not refereed][Not invited]
     
    インフルエンザウイルスにおける抗原変異の規則性を発見するために,アミノ酸残基の共変異を解析する共変異とは,タンパク質を構成するアミノ酸残基のうち〆複数の位置のアミノ酸が共に置換する現象である.従来からアミノ酸残基の共変異を解析する手法がいくつか提案されていたが,それらの手法では進化の過程における分岐や時間的関係が考慮されていなかったそこで,これらの問題を解決するために,進化系統解析によって得られる系統樹を利用する手法を提案する.過去40年間のH3N2亜型インフルエンザウイルスのHAタンパク質を対象とし,共変異の検出を行い,その結果を示す,また,共変異は時代と共に変化するため,共変異の変化を検出するための手法を提案する.The influenza viruses undergo antigenic drift to escape from antibody-mediated immune pressure. In order to predict possible structural changes of their molecules in future, it is important to analyze the patterns of amino acid substitutions in the past. In this paper, we present a new method to extract the sets of residue positions which were involved in correlated mutations. We also discuss a method to detect changes of correlation among co-evolving residues.
  • TANIGUCHI Tsuyoshi, ITO Kimihito, IGARASHI Manabu, MURAKAMI Teiji, TAKADA Ayato, HARAGUCHI Makoto  IPSJ SIG technical reports  2006-  (135)  185  -192  2006/12/21  [Not refereed][Not invited]
     
    The influenza viruses undergo antigenic drift to escape from antibody-mediated immune pressure. In order to predict possible structural changes of their molecules in future, it is important to analyze the patterns of amino acid substitutions in the past. In this paper, we present a new method to extract the sets of residue positions which were involved in correlated mutations. We also discuss a method to detect changes of correlation among co-evolving residues.
  • TANIGUCHI Tsuyoshi, ITO Kimihito, IGARASHI Manabu, MURAKAMI Teiji, TAKADA Ayato, HARAGUCHI Makoto  IPSJ SIG Notes  2006-  (135)  185  -192  2006/12/21  [Not refereed][Not invited]
     
    The influenza viruses undergo antigenic drift to escape from antibody-mediated immune pressure. In order to predict possible structural changes of their molecules in future, it is important to analyze the patterns of amino acid substitutions in the past. In this paper, we present a new method to extract the sets of residue positions which were involved in correlated mutations. We also discuss a method to detect changes of correlation among co-evolving residues.
  • Masayuki Shimojima, Ayato Takada, Hideki Ebihara, Gabriele Neumann, Kouki Fujioka, Tatsuro Irimura, Steven Jones, Heinz Feldmann, Yoshihiro Kawaoka  JOURNAL OF VIROLOGY  80-  (20)  10109  -10116  2006/10  [Not refereed][Not invited]
     
    Filoviruses, represented by the genera Ebolavirus and Marburgvirus, cause a lethal hemorrhagic fever in humans and in nonhuman primates. Although filovirus can replicate in various tissues or cell types in these animals, the molecular mechanisms of its broad tropism remain poorly understood. Here we show the involvement of members of the Tyro3 receptor tyrosine kinase family - Axl, Dtk, and Mer-in cell entry of filoviruses. Ectopic expression of these family members in lymphoid cells, which otherwise are highly resistant to filovirus infection, enhanced infection by pseudotype viruses carrying filovirus glycoproteins on their envelopes. This enhancement was reduced by antibodies to Tyro3 family members, Gas6 ligand, or soluble ectodomains of the members. Live Ebola viruses infected both Axl- and Dtk-expressing cells more efficiently than control cells. Antibody to Axl inhibited infection of pseudotype viruses in a number of Axl-positive cell lines. These results implicate each Tyro3 family member as a cell entry factor in filovirus infection.
  • TAKADA AYATO  膜  31-  (5)  243-247  -247  2006/09/01  [Not refereed][Not invited]
  • 高田 礼人  臨床と微生物 = Clinical microbiology  33-  (4)  337  -343  2006/07/25  [Not refereed][Invited]
  • Hideki Ebihara, Ayato Takada, Darwyn Kobasa, Steven Jones, Gabriele Neumann, Steven Theriault, Mike Bray, Heinz Feldmann, Yoshihiro Kawaoka  PLOS PATHOGENS  2-  (7)  705  -711  2006/07  [Not refereed][Not invited]
     
    Zaire ebolavirus (ZEBOV) causes severe hemorrhagic fever in humans and nonhuman primates, with fatality rates in humans of up to 90%. The molecular basis for the extreme virulence of ZEBOV remains elusive. While adult mice resist ZEBOV infection, the Mayinga strain of the virus has been adapted to cause lethal infection in these animals. To understand the pathogenesis underlying the extreme virulence of Ebola virus ( EBOV), here we identified the mutations responsible for the acquisition of the high virulence of the adapted Mayinga strain in mice, by using reverse genetics. We found that mutations in viral protein 24 and in the nucleoprotein were primarily responsible for the acquisition of high virulence. Moreover, the role of these proteins in virulence correlated with their ability to evade type I interferon-stimulated antiviral responses. These findings suggest a critical role for overcoming the interferon-induced antiviral state in the pathogenicity of EBOV and offer new insights into the pathogenesis of EBOV infection.
  • 高田礼人  臨床と微生物  33-  (4)  337-343  -343  2006/07  [Not refereed][Invited]
  • 高田 礼人  ウイルス  56-  (1)  125  -126  2006/06/26  [Not refereed][Invited]
  • 高田礼人  ウイルス  56-  (1)  117  -124  2006/06/26  [Not refereed][Invited]
     
    In central and west Africa, Ebola virus, a member of the filovirus group, has produced sporadic outbreaks of lethal disease. This virus causes hemorrhagic fever in humans and nonhuman primates, resulting in mortality rates of up to 90%. Although there are no satisfactory biologic explanations for this extreme virulence, it has been suggested that functions of the envelope glycoprotein are likely to play important roles in the pathogenicity of Ebola virus.
  • Hetron M. Munang'andu, Victor M. Siamudaala, Andrew Nambota, John M. Bwalya, Musso Munyeme, Aaron S. Mweene, Ayato Takada, Hiroshi Kida  JAPANESE JOURNAL OF VETERINARY RESEARCH  54-  (1)  3  -13  2006/05  [Refereed][Not invited]
     
    Eco-tourism depending on wildlife is becoming increasingly profitable and landowners are beginning to favor game farming and ecotourism. In these areas, large-scale translocation of wildlife involves a diversity of species and large populations. The African buffalo (Syncerus caffer) is one of the major tourist attractions in Zambia. It accounts for 8.7% and 12.4% of the total animal species hunted in the Game Management Areas and the total hunting revenue earned in Zambia, respectively. It is ecologically an important animal species essential for the purpose of habitat control and facilitating the provision of suitable grazing pastures. However, the rearing of the African buffalo on game ranches has been hampered by its carrier state of the Southern Africa Terroritory (SAT) serotypes of foot and mouth disease virus (FMD). The African buffalo is also known to be a carrier of Theileria parva lawrencei, the causative agent of corridor disease (CD) that continues to have devastating effects on the livestock industry in Zambia. In addition, the importation of buffaloes from countries with populations endemic to bovine tuberculosis is highly restricted. Veterinary regulations in Zambia, strongly advocate against the translocation of buffaloes from protected areas to private ranches for disease control purposes thereby mounting a considerable constraint on the economic and ecological viability of the industry. It is hoped that this review will motivate the relevant government authorities in exploiting ways in which this animal species play a central role in eco-tourism.
  • TAKADA AYATO, NODA TAKESHI, KAWAOKA YOSHIHIRO  膜  30-  (2)  68-72  -72  2005/03/01  [Not refereed][Invited]
  • 高田 礼人, 河岡 義裕  細胞工学  24-  (2)  141  -144  2005/02  [Not refereed][Not invited]
  • TAKADA AYATO, KAWAOKA YOSHIHIRO  細胞工学  24-  (2)  141-144  -144  2005/01  [Not refereed][Invited]
  • D Kobasa, A Takada, K Shinya, M Hatta, P Halfmann, S Theriault, H Suzuki, H Nishimura, K Mitamura, N Sugaya, T Usui, T Murata, Y Maeda, S Watanabe, M Suresh, T Suzuki, Y Suzuki, H Feldmann, Y Kawaoka  NATURE  431-  (7009)  703  -707  2004/10  [Not refereed][Not invited]
     
    The 'Spanish' influenza pandemic of 1918-19 was the most devastating outbreak of infectious disease in recorded history. At least 20 million people(1) died from their illness, which was characterized by an unusually severe and rapid clinical course. The complete sequencing of several genes of the 1918 influenza virus has made it possible to study the functions of the proteins encoded by these genes in viruses generated by reverse genetics, a technique that permits the generation of infectious viruses entirely from cloned complementary DNA. Thus, to identify properties of the 1918 pandemic influenza A strain that might be related to its extraordinary virulence, viruses were produced containing the viral haemagglutinin(2) (HA) and neuraminidase(3) (NA) genes of the 1918 strain. The HA of this strain supports the pathogenicity of a mouse-adapted virus in this animal(4,5). Here we demonstrate that the HA of the 1918 virus confers enhanced pathogenicity in mice to recent human viruses that are otherwise non-pathogenic in this host. Moreover, these highly virulent recombinant viruses expressing the 1918 viral HA could infect the entire lung and induce high levels of macrophage-derived chemokines and cytokines, which resulted in infiltration of inflammatory cells and severe haemorrhage, hallmarks of the illness produced during the original pandemic(6).
  • A Takada, K Fujioka, M Tsuiji, A Morikawa, N Higashi, H Ebihara, D Kobasa, H Feldmann, T Irimura, Y Kawaoka  JOURNAL OF VIROLOGY  78-  (6)  2943  -2947  2004/03  [Not refereed][Not invited]
     
    Filoviruses cause lethal hemorrhagic disease in humans and nonhuman primates. An initial target of filovirus infection is the mononuclear phagocytic cell. Calcium-dependent (C-type) lectins such as dendritic cell- or liver/lymph node-specific ICAM-3 grabbing nonintegrin (DC-SIGN or L-SIGN, respectively), as well as the hepatic asialoglycoprotein receptor, bind to Ebola or Marburg virus glycoprotein (GP) and enhance the infectivity of these viruses in vitro. Here, we demonstrate that a recently identified human macrophage galactose- and N-acetylgalactosamine- specific C-type lectin (hMGL), whose ligand specificity differs from DC-SIGN and L-SIGN, also enhances the infectivity of filoviruses. This enhancement was substantially weaker for the Reston and Marburg viruses than for the highly pathogenic Zaire virus. We also show that the heavily glycosylated, mucin-like domain on the filovirus GP is required for efficient interaction with this lectin. Furthermore, hMGL, like DC-SIGN and L-SIGN, is present on cells known to be major targets of filoviruses (i.e., macrophages and dendritic cells), suggesting a role for these C-type lectins in viral replication in vivo. We propose that filoviruses use different C-type lectins to gain cellular entry, depending on the cell type, and promote efficient viral replication.
  • 堀本泰介, 高田礼人  最新医学  59-  (2)  223-230  -230  2004/02  [Not refereed][Invited]
  • 高田礼人, 河岡義裕  週刊医学のあゆみ  208-  (4)  222-223  -223  2004/01/24  [Not refereed][Not invited]
  • A Takada, Y Kawaoka  REVIEWS IN MEDICAL VIROLOGY  13-  (6)  387  -398  2003/11  [Refereed][Not invited]
     
    Besides the common receptor/coreceptor-dependent mechanism of cellular attachment, some viruses rely on antiviral antibodies for their efficient entry into target cells. This mechanism, known as antibody-dependent enhancement (ADE) of viral infection, depends on the cross-linking of complexes of virus-antibody or virus-activated complement components through interaction with cellular molecules such as Fc receptors or complement receptors, leading to enhanced infection of susceptible cells. Recent studies have suggested that additional mechanisms underlie ADE: involvement of complement component C1q and its receptor (Ebola virus), antibody-mediated modulation of the interaction between viral protein and its coreceptor (human immunodeficiency virus) and suppression of cellular antiviral genes by the replication of viruses entering cells via ADE (Ross River virus). Since ADE is exploited by a variety of viruses and has been associated with disease exacerbation, it may have broad relevance to the pathogenesis of viral infection and antiviral strategies. Copyright (C) 2003 John Wiley Sons, Ltd.
  • A Takada, H Feldmann, TG Ksiazek, Y Kawaoka  JOURNAL OF VIROLOGY  77-  (13)  7539  -7544  2003/07  [Not refereed][Not invited]
     
    Most strains of Ebola virus cause a rapidly fatal hemorrhagic disease in humans, yet there are still no biologic explanations that adequately account for the extreme virulence of these emerging pathogens. Here we show that Ebola Zaire virus infection in humans induces antibodies that enhance viral infectivity. Plasma or serum from convalescing patients enhanced the infection of primate kidney cells by the Zaire virus, and this enhancement was mediated by antibodies to the viral glycoprotein and by complement component C1q. Our results suggest a novel mechanism of antibody-dependent enhancement of Ebola virus infection, one that would account for the dire outcome of Ebola outbreaks in human populations.
  • A Takada, S Matsushita, A Ninomiya, Y Kawaoka, H Kida  VACCINE  21-  (23)  3212  -3218  2003/07  [Not refereed][Not invited]
     
    It has been known that influenza A virus infection induces a cross-protective immunity against infection by viruses with different subtypes of viral envelope proteins, hemagglutinin (HA) and neuraminidase (NA). This heterosubtypic immunity is generally mediated by cytotoxic T lymphocytes (CTL) reactive to specific epitopes in the viral internal proteins, such as nucleoprotein and matrix protein. By contrast, immunization with inactivated virus antigens has been thought to be unable to generate heterosubtypic immunity, since inactivated antigens do not usually induce CTL responses. However, we show that intranasal immunization with formalin-inactivated intact virus, but not ether-split vaccines, induced a broad spectrum of heterosubtypic protective immunity in mice. The protection may be mediated by the mucosal immune response, most likely secretory IgA antibodies to the viral proteins. This approach may overcome limitations in the efficacy of inactivated influenza vaccines and confer potent immunity to humans against viruses with new pandemic potential. (C) 2003 Elsevier Science Ltd. All rights reserved.
  • 田村 慎一, 高田 礼人, 河岡 義裕  インフルエンザ  4-  (2)  105  -118  2003/04  [Not refereed][Invited]
  • A Takada, H Feldmann, U Stroeher, M Bray, S Watanabe, H Ito, M McGregor, Y Kawaoka  JOURNAL OF VIROLOGY  77-  (2)  1069  -1074  2003/01  [Not refereed][Not invited]
     
    Ebola virus causes lethal hemorrhagic fever in humans, but currently there are no effective vaccines or antiviral compounds for this infectious disease. Passive transfer of monoclonal antibodies (MAbs) protects mice from lethal Ebola virus infection (J. A. Wilson, M. Hevey, R. Bakken, S. Guest, M. Bray, A. L. Schmaljohn, and M. K. Hart, Science 287:1664-1666, 2000). However, the epitopes responsible for neutralization have been only partially characterized because some of the MAbs do not recognize the short synthetic peptides used for epitope mapping. To identify the amino acids recognized by neutralizing and protective antibodies, we generated a recombinant vesicular stomatitis virus (VSV) containing the Ebola virus glycoprotein-encoding gene instead of the VSV G protein-encoding gene and used it to select escape variants by growing it in the presence of a MAb (133/3.16 or 226/8.1) that neutralizes the infectivity of the virus. All three variants selected by MAb 133/3.16 contained a single amino acid substitution at amino acid position 549 in the GP2 subunit. By contrast, MAb 226/8.1 selected three different variants containing substitutions at positions 134, 194, and 199 in the GP1 subunit, suggesting that this antibody recognized a conformational epitope. Passive transfer of each of these MAbs completely protected mice from a lethal Ebola virus infection. These data indicate that neutralizing antibody cocktails for passive prophylaxis and therapy of Ebola hemorrhagic fever can reduce the possibility of the emergence of antigenic variants in infected individuals.
  • 堀本 泰介, 五藤 秀男, 高田 礼人  免疫  2003-  212  -226  2003  [Not refereed][Not invited]
  • T Noda, H Sagara, E Suzuki, A Takada, H Kida, Y Kawaoka  JOURNAL OF VIROLOGY  76-  (10)  4855  -4865  2002/05  [Not refereed][Not invited]
     
    Using biochemical assays, it has been demonstrated that expression of Ebola virus VP40 alone in mammalian cells induced production of particles with a density similar to that of virions. To determine the morphological properties of these particles, cells expressing VP40 and the particles released from the cells were examined by electron microscopy. VP40 induced budding from the plasma membrane of filamentous particles, which differed in length but had uniform diameters of approximately 65 nm. When the Ebola virus glycoprotein (GP) responsible for receptor binding and membrane fusion was expressed in cells, we found pleomorphic particles budding from the plasma membrane. By contrast, when GP was coexpressed with VP40, GP was found on the filamentous particles induced by VP40. These results demonstrated the central role of VP40 in formation of the filamentous structure of Ebola virions and may suggest an interaction between VP40 and GP in morphogenesis.
  • The invasion routes of neurovirulent A/Hong Kong/483/97(H5N1) influenza virus into the central nervous system after respiratory infection in mice(共著)
    Archives of Virology  147, 1425-1436-  2002  [Not refereed][Not invited]
  • Genome Research  12,595-601-  2002  [Not refereed][Not invited]
  • CH Park, H Ozaki, A Takada, H Kida, K Ochiai, T Umemura  AVIAN PATHOLOGY  30-  (3)  269  -272  2001/06  [Not refereed][Not invited]
     
    Virulent or avirulent strains of type A influenza virus were inoculated into the allantoic cavities of chicken embryos. The antigens of virulent strains appeared initially in the surface epithelium of the allantoic membrane, then in vascular endothelial cells of the chorioallantoic membrane and visceral organs of the embryos, and then spread to parenchymal cells of many organs. In contrast, the antigens of the avirulent strain were confined to the allantoic membrane. These observations indicate that the primary target of virulent influenza viruses in chicken embryos is vascular endothelial cells, and that the embryos died after systemic viral infection.
  • YK Lim, A Takada, T Tanizaki, H Ozaki, K Okazaki, H Kida  JAPANESE JOURNAL OF VETERINARY RESEARCH  48-  (4)  197  -203  2001/02  [Not refereed][Not invited]
     
    Effective vaccinations against swine influenza reduce the economic loss of pig industries, and also may minimize the possibility of emergence of new pandemic viruses, since pigs are intermediate hosts to generate reassortant viruses among avian and mammalian influenza viruses. In this study, we showed that intranasal immunization of pigs with formalin-inactivated or ether-split influenza vaccine (A/Aichi/2/68) induced virus-specific IgG, IgM, and IgA antibodies in their nasal secretions and sera, resulting in complete protection from virus challenge. Antibody response to the challenge virus was not observed in the immunized pigs, suggesting that the replication of the virus in the primary targets, respiratory epithelial cells, was inhibited. The present results indicate that intranasal immunization of pigs with inactivated vaccines is effective to control swine influenza, and also provide a good model, as well as a mouse model, to evaluate an intranasal application of influenza vaccine for humans.
  • Trends in Microbiology  9(10), 506-511-  2001  [Refereed][Invited]
  • Journal of Virology  75, 9297-9301-  2001  [Not refereed][Not invited]
  • Trends in Microbiology  9(10), 506-511-  2001  [Not refereed][Not invited]
  • HK Jin, K Yoshimatsu, A Takada, M Ogino, A Asano, J Arikawa, T Watanabe  ARCHIVES OF VIROLOGY  146-  (1)  41  -49  2001  [Not refereed][Not invited]
     
    The antiviral potential of Mx2 protein remains unknown, because the Mx2 gene in commonly used strains of laboratory mice is nonfunctional. Our previous study showed that functional Mx2 protein in some feral-origin strains was induced upon interferon treatment, was localized in the cytoplasm, and inhibited vesicular stomatitis virus replication. In the present study, we have demonstrated chat the embryonic fibroblastic cells from a feral-origin strain (SPR) expressed 74 kDa Mx2 protein, which prevented the accumulation of viral transcripts and proteins of hantaviruses when the Mx2 gene was constitutively expressed in transfected Vero cells. Furthermore, the cells showed significantly lower titers of the virus than control cells. In contrast, influenza virus replication was not affected by the expression of Mx2 protein in the Vero cells.
  • Shortridge KF, Gao P, Guan Y, Ito T, Kawaoka Y, Markwell D, Takada A, Webster RG  Veterinary Microbiology  74-  141  -147  2001  [Refereed][Not invited]
  • A Takada, S Watanabe, H Ito, K Okazaki, H Kida, Y Kawaoka  VIROLOGY  278-  (1)  20  -26  2000/12  [Not refereed][Not invited]
     
    Filoviruses, including Ebola virus, are cytotoxic. To investigate the role of the Ebola virus glycoprotein (GP) in this cytopathic effect, we transiently expressed the GP in human kidney 293T cells. Expression of wild-type GP, but not the secretory form of the molecule lacking a membrane anchor, induced rounding and detachment of the cells, as did a chimeric GP containing its ectodomain and influenza virus hemagglutinin transmembrane-cytoplasmic domain. These results indicate that the GP ectodomain and its anchorage to the membrane are required for GP-induced morphologic changes in host cells. Since cell rounding and detachment could be associated with reduced levels of cell adhesion molecules, we also studied the expression of integrins, which are major molecules for adhesion to extracellular matrices, and found that the beta1 integrin group is downregulated by the GP. This result was further extended by experiments in which anti-beta1 monoclonal antibodies or purified integrins inhibited the infectivity of vesicular stomatitis virus pseudotyped with the GP. We suggest that integrins, especially the beta1 group, might interact with the GP and perhaps be involved in Ebola virus entry into cells, (C) 2000 Academic Press.
  • T Ito, Y Suzuki, T Suzuki, A Takda, T Horimoto, K Wells, H Kida, K Otsuki, M Kiso, H Ishida, Y Kawaoka  JOURNAL OF VIROLOGY  74-  (19)  9300  -9305  2000/10  [Not refereed][Not invited]
     
    The hemagglutinin (IU) of H3 human influenza viruses does not support viral replication in duck intestine despite its avian origin. A Leu-to-Gin mutation at position 226 and a Ser-to-Gly mutation at position 228 in the HA of human A/Udorn/307/72 (H3N2) permit a reassortant virus [human Udorn HA, with all other genes from A/mallard/New York/6750/78 (H2N2)] to replicate in ducks. To understand the molecular basis of this change in host range restriction, we investigated the receptor specificity of duck influenza viruses as well as of human-duck virus reassortants. The results indicate that the recognition of a glycoconjugate moiety possessing N-glycolneuramic acid (NeuGc) linked to galactose by the alpha 2,3 linkage (NeuGc alpha 2,3Gal) is associated with viral replication in duck intestine. Immunofluorescence assays with NeuGc alpha 2,3Gal-specific antiserum detected this moiety primarily on the crypt epithelial cells of duck colon. Such recognition, together with biochemical evidence of NeuGc in crypt cells, correlated exactly with the ability of the virus to replicate in duck colon. These results suggest that recognition of the NeuGc alpha 2,3-Gal moiety plays an important role in the enterotropism of avian influenza viruses.
  • T Ito, Y Suzuki, T Suzuki, A Takda, T Horimoto, K Wells, H Kida, K Otsuki, M Kiso, H Ishida, Y Kawaoka  JOURNAL OF VIROLOGY  74-  (19)  9300  -9305  2000/10  [Not refereed][Not invited]
     
    The hemagglutinin (IU) of H3 human influenza viruses does not support viral replication in duck intestine despite its avian origin. A Leu-to-Gin mutation at position 226 and a Ser-to-Gly mutation at position 228 in the HA of human A/Udorn/307/72 (H3N2) permit a reassortant virus [human Udorn HA, with all other genes from A/mallard/New York/6750/78 (H2N2)] to replicate in ducks. To understand the molecular basis of this change in host range restriction, we investigated the receptor specificity of duck influenza viruses as well as of human-duck virus reassortants. The results indicate that the recognition of a glycoconjugate moiety possessing N-glycolneuramic acid (NeuGc) linked to galactose by the alpha 2,3 linkage (NeuGc alpha 2,3Gal) is associated with viral replication in duck intestine. Immunofluorescence assays with NeuGc alpha 2,3Gal-specific antiserum detected this moiety primarily on the crypt epithelial cells of duck colon. Such recognition, together with biochemical evidence of NeuGc in crypt cells, correlated exactly with the ability of the virus to replicate in duck colon. These results suggest that recognition of the NeuGc alpha 2,3-Gal moiety plays an important role in the enterotropism of avian influenza viruses.
  • KF Shortridge, P Gao, Y Guan, T Ito, Y Kawaoka, D Markwell, A Takada, RG Webster  VETERINARY MICROBIOLOGY  74-  (1-2)  141  -147  2000/05  [Not refereed][Not invited]
     
    This account takes stock of events and involvements, particularly on the avian side of the influenza H5N1 'bird flu' incident in Hong Kong SAR in 1997, It highlights the role of the chicken in the many live poultry markets as the source of the virus for humans. The slaughter of chicken and other poultry across the SAR seemingly averted an influenza pandemic. This perspective from Hong Kong SAR marks the: coming-of-age of acceptance of the role of avian hosts as a source of pandemic human influenza viruses and offers the prospect of providing a good baseline for influenza pandemic preparedness in the future. Improved surveillance is the key. This is illustrated through the H9N2 virus which appears to have provided the 'replicating' genes for the H5N1 virus and which has since been isolated in the SAR from poultry, pigs and humans highlighting its propensity for interspecies transmission. (C) 2000 Elsevier Science B.V. All rights reserved.
  • Archives of Virology  145-  (8)  1715  -1723  2000  [Not refereed][Not invited]
  • Archives of Virology  145-  (5)  885  -893  2000  [Not refereed][Not invited]
  • Avirulent avian influenza virus as a vaccine strain against a potential human pandemic
    A Takada, N Kuboki, K Okazaki, A Ninomiya, H Tanaka, H Ozaki, S Itamura, H Nishimura, M Enami, M Tashiro, KF Shortridge, H Kida  JOURNAL OF VIROLOGY  73-  (10)  8303  -8307  1999/10  [Not refereed][Not invited]
     
    In the influenza H5N1 virus incident in Hong Kong in 1997, viruses that are closely related to H5N1 viruses initially isolated in a severe outbreak of avian influenza in chickens were isolated from humans, signaling the possibility of an incipient pandemic. However, it was not possible to prepare a vaccine against the virus in the conventional embryonated egg system because of the lethality of the virus for chicken embryos and the high level of biosafety therefore required for vaccine production. Alternative approaches, including an avirulent H5N4 virus isolated from a migratory duck as a surrogate virus, H5N1 virus as a reassortant with avian virus H3N1 and an avirulent recombinant H5N1 virus generated by reverse genetics, have been explored. All vaccines were formalin inactivated. Intraperitoneal immunization of mice with each of vaccines elicited the production of hemagglutination-inhibiting and virus-neutralizing antibodies, while intranasal vaccination without adjuvant induced both mucosal and systemic antibody responses that protected the mice from lethal H5N1 virus challenge. Surveillance of birds and animals, particularly aquatic birds, for viruses to provide vaccine strains, especially surrogate viruses, for a future pandemic is stressed.
  • H Aoki, Y Sakoda, K Jukuroki, A Takada, H Kida, A Fukusho  VETERINARY MICROBIOLOGY  68-  (3-4)  197  -207  1999/08  [Not refereed][Not invited]
     
    The glycoprotein gB of pseudorabies virus (PrV) was expressed in various mammalian cells by a recombinant baculovirus carrying the PrV gB gene under the control of the CAG promoter. When the recombinant baculovirus was inoculated into the stable porcine kidney cell line CPK, expression of PrV gB was detected by immunofluorescent antibody analysis and a 155 kDa of protein, which has the same molecular mass as the native PrV gB, was detected by Western blotting. High levels of expression of PrV gB were observed in BHK-21, HmLu-1 and SK-K cell Lines. Furthermore, anti-PrV gB-specific antibodies against PrV gB protein were detected by the enzyme-linked immunosorbent assay in mice inoculated the recombinant baculovirus. The recombinant baculovirus containing the PrV glycoprotein gB gene under the CAG promoter could be a candidate for a pseudorabies vaccine. (C) 1999 Elsevier Science B.V. All rights reserved.
  • Identification of the murine Mx2 gene: Interferon-induced expression of the Mx2 protein from the feral mouse gene confers resistance to vesicular stomatitis virus
    HK Jin, A Takada, Y Kon, O Haller, T Watanabe  JOURNAL OF VIROLOGY  73-  (6)  4925  -4930  1999/06  [Not refereed][Not invited]
     
    The mouse genome contains two related interferon-regulated genes, Mx1 and Mx2, Whereas Mx1 codes for the nuclear 72-kDa protein that interferes with influenza virus replication after interferon treatment, the Mx2 gene is nonfunctional in all laboratory mouse strains examined, since its open reading frame (ORF) is interrupted by an insertional mutation and a subsequent frameshift mutation. In the present study, we demonstrate that Mx2 mRNA of cells from feral mouse strains NJL (Mus musculus musculus) and SPR (Mus spretus) differs from that of the laboratory mouse strains tested. The Mx2 mRNA of the feral strains contains a single long ORF consisting of 656 amino acids. We further show that Mx2 protein in the feral strains is expressed upon interferon treatment and localizes to the cytoplasm much like the rat Mx2 protein, which inhibits vesicular stomatitis virus replication. Furthermore, transfected 3T3 cell lines of laboratory mouse origin expressing Mx2 from feral strains acquire slight resistance to vesicular stomatitis virus.
  • M Imai, A Takada, K Okazaki, H Kida  JAPANESE JOURNAL OF VETERINARY RESEARCH  46-  (4)  171  -177  1999/02  [Not refereed][Not invited]
     
    The hemagglutinin (HA) of six H5 influenza virus strains isolated from ducks in Japan and China in 1976 to 1996 were analyzed antigenically and genetically. Antigenic analysis using a panel of monoclonal antibodies revealed that the HA of H5 influenza viruses isolated from ducks are antigenically closely related to each other. Phylogenetic analysis indicates that the isolates from ducks in Hokkaido were derived from an ancestor common with the highly pathogenic isolates from chickens and humans in Hong Kong in 1997.
  • Protective effects of intranasal vaccination with plasmid encoding pseudorabies virus glycoprotein B in mice
    Japanese Journal of Veterinary Research  47-  25  -33  1999  [Not refereed][Not invited]
  • Masahiro Miyoshi, Yuki Ishii, Mitsuyoshi Takiguchi, Ayato Takada, Jun Yasuda, Akira Hashimoto, Katsunori Okazaki, Hiroshi Kida  Jornal of Veterinary Medical Science  61-  (4)  375  -379  1999  [Not refereed][Not invited]
  • Archives of Virology  144-  (2)  407  -420  1999  [Not refereed][Not invited]
  • A Takada, Y Kawaoka  TRENDS IN MICROBIOLOGY  6-  (7)  258  -259  1998/07  [Refereed][Invited]
  • Virology  252-  (2)  331  -342  1998  [Not refereed][Not invited]
  • Virus Research  56-  (2)  169  -176  1998  [Not refereed][Not invited]
  • Tends in Microbiology  6-  (7)  258  1998  [Not refereed][Not invited]
  • Japanese Journal of Veterinary Research  45-  207  1998  [Not refereed][Not invited]
  • Archives of Virology  143-  (6)  1129  -1143  1998  [Not refereed][Not invited]
  • A Takada, C Robison, H Goto, A Sanchez, KG Murti, MA Whitt, Y Kawaoka  PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA  94-  (26)  14764  -14769  1997/12  [Not refereed][Not invited]
     
    Ebola virus causes hemorrhagic fever in humans and nonhuman primates, resulting in mortality rates of up to 90%. Studies of this virus have been hampered by its extraordinary pathogenicity, which requires biosafety level 4 containment. To circumvent this problem, we developed a novel complementation system for functional analysis of Ebola virus glycoproteins, It relies on a recombinant vesicular stomatitis virus (VSV) that contains the green fluorescent protein gene instead of the receptor-binding G protein gene (VSV Delta G*). Herein we show that Ebola Reston virus glycoprotein (ResGP) is efficiently incorporated into VSV particles. This recombinant VSV with integrated ResGP (VSV Delta G*-ResGP) infected primate cells more efficiently than any of the other mammalian or avian cells examined, in a manner consistent with the host range tropism of Ebola virus, whereas VSV Delta G* complemented with VSV G protein (VSV Delta G*-G) efficiently infected the majority of the cells tested. We also tested the utility of this system for investigating the cellular receptors for Ebola virus. Chemical modification of cells to alter their surface proteins markedly reduced their susceptibility to VSV Delta G*-ResGP but not to VSV Delta G*-G. These findings suggest that cell surface glycoproteins with N-linked oligosaccharide chains contribute to the entry of Ebola viruses, presumably acting as a specific receptor and/or cofactor for virus entry. Thus, our VSV system should be useful for investigating the functions of glycoproteins from highly pathogenic viruses or those incapable of being cultured in vitro.
  • KAWAOKA YOSHIHIRO, TAKADA AYATO, WHITT M A  実験医学  15-  (19)  2389-2393  -2393  1997/12  [Not refereed][Not invited]
  • KIDA Hiroshi, OKAZAKI Katsunori, TAKADA Ayato  Japanese journal of veterinary research  45-  (1)  32  -33  1997/05/30  [Not refereed][Not invited]
  • Virology  238-  (1)  128  -134  1997  [Not refereed][Not invited]
  • Differences in sialic acid-galactose linkages in the chicken egg amnion and allantois influence human influenza virus receptor specificity and variant selection(共著)
    Journal of Virology  71-  (4)  3357  1997  [Not refereed][Not invited]
  • ウイルスの病原性発現における糖タンパク質の役割
    堀本泰介, 五藤秀男, 高田礼人, 河岡義裕  Molecular Medicine  39-  212  -226  1997  [Not refereed][Invited]
  • ISLAM Mohammed A, ITO Toshihiro, TAKAKUWA Hiroki, TAKADA Ayato, ITAKURA Chitoshi, KIDA Hiroshi  Japanese journal of veterinary research  42-  (3)  147  -156  1995/01/31  [Not refereed][Not invited]
     
    Newcastle disease virus (NDV) was isolated from a Japanese quail (Cotornix cotornix japonica). The effect of intracerebral and intranasal passages of the NDV in chickens on the pathogenicity was studied. Pathogenicity of the viruses of different passage levels was compared with that of the original isolate by the mean death time with the minimum lethal dose in chicken embryos, intracerebral pathogenicity index in day-old chicks, intravenous pathogenicity index with 6-week-old chickens and the mortality rates of chickens and quails inoculated intravenously or intranasally. The original isolate from the quail did not kill chickens but only embryos and some one-day-old chicks, exhibiting a mesogenic character. Pathogenicity of the virus of the 10th intranasal passage was not different from that of the original isolate. The viruses passaged intracerebrally, on the other hand, killed chickens of all ages by either route of inoculation, showing a velogenic property. Virus recovery from the blood and the brain was positive only in the chickens infected with brain-passaged viruses by any route of inoculation. Virus titers in the tissues of chickens infected with the brain-passaged viruses were higher than those with the original isolate and the virus of the 10th intranasal passage. These results indicate that the enhanced pathogenicity of the mesogenic NDV isolate from the quail for chickens was induced by acquiring the properties of neurotropism and pantropism through intracerebral passage in chickens.
  • Archives of Virology  140-  (7)  1163  -1172  1995  [Not refereed][Not invited]
  • Archives of Virology  140-  (9)  1629  -1635  1995  [Not refereed][Not invited]
  • A TAKADA, Y SHIMIZU, H KIDA  JOURNAL OF VETERINARY MEDICAL SCIENCE  56-  (4)  633  -637  1994/08  [Not refereed][Not invited]
     
    Intranasal vaccination of mice with inactivated Aujeszky's disease virus (ADV) induced IgA and IgG antibody responses to the virus in the secretion of the respiratory tract, resulting in complete protection of the animals against intranasal challenge with virulent ADV. The immune response was enhanced by the use of the cholera toxin B subunit (CTB) as an adjuvant. On the other hand, subcutaneous vaccination of mice with inactivated ADV, even together with CTB, scarcely stimulated secretory antibody responses, resulting in only partial protection. The present results suggest that development of a vaccination procedure to stimulate the mucosal immune response should improve the protective effects of the inactivated herpesvirus vaccines, and thereby make it possible to control the infections by prohibiting virus replication at the site where primary infection takes place, as well as inhibiting subsequent latency and reactivation of the virus.
  • Acquisition of pathogenicity of a Newcastle disease virus isolated from a Japanese quail by intracerebral passage in chickens(共著)
    Japanese Journal of Veterinary Research  42-  147  1994  [Not refereed][Not invited]

Industrial Property Rights

  • 特願2017-193292:抗ウイルス性ペプチドおよびその応用  2017年/10/03
    吉田徹彦, 高田礼人, ベイリー小林菜穂子
  • 特願2017-071729:ィロウイルスの細胞侵入阻害活性を有するビアリールスルフォンアミド誘導体  2017年/03/31
    高田礼人, 堺谷政弘, 古山若呼
  • 特願2016-216753:免疫誘導剤  2016年/11/11
    増田健一, 齊藤隆, 石井保之, 高田礼人, 五十嵐学, 丸山隼輝, 齊藤祐介, 奈良拓也
  • 特願2016-188897:エボラウイルスワクチン  2016年/09/27
    増田健一, 齊藤隆, 石井保之, 高田礼人, 五十嵐学, 丸山隼輝, 齊藤祐介, 奈良拓也
  • 特願2015-161567:全てのエボラウイルス種の感染性を中和するモノクローナル抗体  2015年/08/19
    高田礼人, 吉田玲子, 古山若呼, 宮本洋子, 丸山隼輝, 飯田茂, 小川進也
  • 特願2014-210419:フィロウイルス感染阻害剤のスクリーニング法  2014年/10/15
    高田礼人, 南保明日香, 黒田誠, 藤倉大輔
  • 入村 達郎, 河岡 義裕, 高田 礼人, 藤岡 宏樹  国立大学法人 東京大学  200903062567546269
  • 入村 達郎, 河岡 義裕, 高田 礼人, 藤岡 宏樹  国立大学法人 東京大学  200903072722971678

Awards & Honors

  • 2017/01 北海道大学 北海道大学研究総長賞 奨励賞
     
    受賞者: 高田 礼人
  • 2016/02 Hokkaido University Hokkaido University President's Award for Outstanding Research
     
    受賞者: TAKADA Ayato
  • 2015/02 Hokkaido University Hokkaido University President's Award for Outstanding Research
     
    受賞者: TAKADA Ayato
  • 2005 日本ウイルス学会 杉浦奨励賞

Research Grants & Projects

  • フィロウイルスの宿主域・病原性・生態の解明と予防・治療・診断法の開発
    日本医療研究開発機構(AMED):感染症研究革新イニシアティブ(J-PRIDE)
    Date (from‐to) : 2017/09 -2025/03 
    Author : 高田 礼人
  • Epidemiology of zoonotic virus infections in Africa
    Japan Agency for Medical Research and Development:Science and Technology Research Partnership for Sustainable Development
    Date (from‐to) : 2018/04 -2024/03 
    Author : TAKADA Ayato
  • エボラ出血熱に対する治療法の開発
    文部科学省:科学研究費補助金(基盤(A))
    Date (from‐to) : 2016/04 -2021 
    Author : 高田 礼人
  • Surveillance of Viral Zoonoses in Africa
    Japan Science and Technology Agency:Science and Technology Research Partnership for Sustainable Development
    Date (from‐to) : 2012/06 -2017/03 
    Author : TAKADA Ayato
  • 文部科学省:科学研究費補助金(挑戦的萌芽研究)
    Date (from‐to) : 2015 -2016 
    Author : 高田 礼人
  • 文部科学省:科学研究費補助金(新学術領域研究(研究領域提案型))
    Date (from‐to) : 2015 -2016 
    Author : 高田 礼人
  • 文部科学省:科学研究費補助金(基盤研究(B))
    Date (from‐to) : 2012 -2015 
    Author : 高田 礼人
     
    インフルエンザAウイルスはウイルス表面糖蛋白質であるヘマグルチニン(HA)およびノイラミダーゼ(NA)の抗原性によって複数の亜型に分けられる。ウイルス中和抗体の標的は主にHAであり、通常の中和試験において抗ウイルス血清は亜型間で交差反応性を示さないことから、亜型間交差感染防御免疫における抗体の役割に関する知見は限られていた。しかし、これまでに申請者らは亜型間交差反応性のHA特異的モノクローナル抗体の作出に成功した。本研究では、これらのモノクローナル抗体を用いて、HA亜型間共通エピトープを探索し、亜型間交差感染防御免疫における抗体の関与を明らかにする。また、亜型間交差反応性モノクローナル抗体とHAの結合構造を詳細に解析し、HA分子と相互作用する抗体分子上のアミノ酸を改変することによって、抗体の亜型間交差反応性を操作する技術の確立を試みる。これまでに、H1、H2、H3およびH13亜型のウイルスに対して亜型間交差中和活1生を示すモノクローナル抗体(S139/1)の作出に成功した。S139/1とHAの共結晶構造のX線解析によって、この抗体はHAのglobular head領域のHAのレセプター結合近傍のアミノ酸を広く覆い、HCDR2がレセプター結合ポケットの中に深く入り込む結合様式であることが分かった。また、不活化ウイルスでマウスを免疫すると、通常の中和活性は持たないが複数の異なる亜型のウイルスに対する抗体が誘導されることを明らかにした。本研究は、亜型間交差感染防御免疫における抗体の役割を実証し、交差反応性抗体を用いたインフルエンザに対する抗体療法の可能性を示唆するものである。
  • 文部科学省:科学研究費補助金(新学術領域研究(研究領域提案型))
    Date (from‐to) : 2013 -2014 
    Author : 高田 礼人
  • 文部科学省:科学研究費補助金(挑戦的萌芽研究)
    Date (from‐to) : 2012 -2013 
    Author : 高田 礼人
     
    通常のウイルス感染性中和試験とは、培養細胞外でウイルスと抗体を混合することによって、ウイルスに結合した抗体がウイルスの細胞侵入を阻害する「細胞外中和活性」を検出する方法である。したがって、ウイルス感染性中和と感染防御抗体の従来の概念は、「細胞外中和」の有無によって形成されてきた。しかし、これまでに申請者は通常のウイルス中和試験では活性が殆ど認められない抗体でも、ウイルス感染(吸着・侵入)後に培養細胞の上清に抗体を添加しておくと、ウイルスの増殖を著しく減少させる現象を見出した。本研究は、フィロウイルス感染モデルを用いて、抗体が感染細胞表面でウイルスの出芽・粒子形成過程を抑える「細胞表面」中和の可能性を探り、抗体による新たな中和原理の発見を目指す。これまでに作出したフィロウイルス(エボラおよびマールブルグウイルス)の表面糖蛋白質(GP)に対するマウスモノクローナル抗体について、非増殖性シュードタイプVesicular Stomatitis Virus (VSV)および増殖性シュードタイプVSVを用いて、通常の中和試験および細胞表面の中和試験を行った。その結果、通常の中和活性(細胞外中和)の無い抗体(マールブルグウイルスに対する抗体)でも、単層培養したVero E6細胞に増殖性シュードタイプVSVを感染させた後に、抗体を含む培地で培養すると、抗体が細胞表面に発現したGPに結合してウイルスの出芽が抑えられ、プラック形成抑制効果を有することが明らかとなった。
  • 文部科学省:科学研究費補助金(基盤研究(B))
    Date (from‐to) : 2009 -2011 
    Author : 高田 礼人
     
    インフルエンザAウイルスはヒトを含む多くの哺乳類および鳥類に感染する病原体であり、インフルエンザは重要な人獣共通感染症の一つである。現行の注射による不活化ワクチンは、中和抗体の標的であるヘマグルチニン(HA)およびノイラミニダーゼ(NA)、に対する血中抗体の産生を誘導し、重症化を防ぐが、粘膜組織でのウイルスの初感染を阻止できない。また、現在ヒトに流行しているインフルエンザウイルスとは異なるHA亜型のウイルスによる「新型インフルエンザ」に対しては、現行のワクチンは全く無効である。本研究は、それら諸問題を解決し、インフルエンザに対する新規予防・治療法を開発するための基盤研究である。BALB/cマウスの鼻腔内または皮下にフォルマリン不活化インフルエンザウイルスを2-3週間間隔で2または3回接種し、ヘマグルチニンに特異的な血清中の抗体量をELISAで測定した結果、複数の亜型のウイルスに対して交差反応性を示す抗体が誘導されることが明らかになった。また、この交差反応性の範囲は、進化系統学的な分類と必ずしも一致しない事が明らかになった。さらに、免疫したマウスの脾臓を用いて、定法に従いハイブリドーマを作出し、ウイルス蛋白質特異抗体を産生するハイブリドーマをスクリーニングし、ウイルスの亜型間で交差反応性を示すモノクローナル抗体を複数得た。それらの抗体はウイルスの感染性を中和するものと、結合はす...
  • 文部科学省:科学研究費補助金(挑戦的萌芽研究)
    Date (from‐to) : 2009 -2010 
    Author : 伊藤 公人, 高田 礼人
     
    本研究では、公共データベースに蓄積された大量のウイルスゲノムを計算機を用いて解析することにより、RNA分節の選択的集合を説明し得る塩基対の構造を網羅的に探索することを目的とする。平成22年度は、下記項目に関して研究を行った。1.RNA分節において、塩基が極度に保存されている位置を明らかにする従来研究の成績からパッケージングシグナルは、各RNA分節のから5'末端および3'末端付近の領域に存在することが示唆されている。パンデミック2009インフルエンザウイルス(H1N1)の2285株分の塩基配列をNCBIから取得し、平均情報量を指標に各RNA分節において塩基が極度に保存されている領域を検索した。その結果、ほとんど全てのRNA分節において、3'末端付近の領域に塩基置換が見られた。2.二つのRNA分節で同時に変異する傾向にある塩基対があるか否かを確認する二つのRNA分節で同時に変異する傾向にある塩基対を高速に検索するために、同時エントロピーの比を用いる手法を開発した。同法を用いて、パンデミック2009インフルエンザウイルス(H1N1)のRNA分節間で同時に変異している塩基を探索した。その結果、PB2の95番目とNPの1530番目、PB2の573番目とNPの44番目、PB1の288番目とHAの1715番目、PAの64番目とNPの417番目、PAの64番目とNPの1419番目、NPの32...
  • 文部科学省:科学研究費補助金(萌芽研究, 挑戦的萌芽研究)
    Date (from‐to) : 2008 -2009 
    Author : 高田 礼人
     
    ウイルス感染症の対策として、ワクチン接種による予防が最も一般的であり効果が期待される。ワクチンは生ワクチンと不活化ワクチンに分類されるが、生ワクチンは安全性の問題、不活化ワクチンは免疫原性の弱さの問題が、それぞれ短所として挙げられ、双方の短所を克服した安全で効果的なワクチンの開発は困難である。特に、生ワクチンは短期間で作出することが不可能であるため、発生頻度が増している新興感染症に対して迅速に対応するためには、不活化ワクチンの効率的な開発および接種法の改良が求められる。本研究では、ウイルス感染に広く認められる抗体依存性感染増強現象(ADE)に着目した。この現象は、ウイルスが抗体を利用してマクロファージや樹状細胞等の抗原提示細胞に効率よく感染するためのメカニズムであると考えられている。本研究では、この抗体の特性を利用して抗原提示細胞に目的の不活化ワクチン抗原を効率よく取り込ませるための手法を開発し、液性免疫および細胞性免疫の両方を誘導する安全なアジュバントとしての抗体の可能性を探る。不活化ウイルス抗原として、ショ糖密度勾配遠心によって精製したエボラウイルスのウイルス様粒子をウイルス表面糖蛋白質に特異的なADE抗体、中和抗体およびどちらの活性も示さないモノクローナル抗体とそれぞれ混合し、マウスの皮下または腹腔内接種して、経時的に血清中の抗体応答を解析し、異なる性質を持つこれらの抗...
  • Ministry of Education, Culture, Sports, Science and Technology:Grants-in-Aid for Scientific Research(基盤研究(B))
    Date (from‐to) : 2007 -2009 
    Author : Kimihito ITO, Ayato TAKADA, Shin-ichi MINATO
     
    Human influenza viruses mutate from time to time, causing annual epidemics worldwide. Given the high mutation rate of the viral gene, it is difficult to select an effective vaccine strain prior to each influenza season. In order to elucidate the pattern of viral evolution, we introduce a bioinformatics technology that analyzes a large number of epidemic strains in a multidimensional space. We found that the relative sequence distances among past epidemic strains could be predicted by a mathematical model. Retrospective tests for 12 years showed that the model could predict the direction of ...
  • 文部科学省:科学研究費補助金(特定領域研究)
    Date (from‐to) : 2007 -2008 
    Author : 高田 礼人
     
    フィロウイルスは霊長類に重篤な出血熱を引き起こし、その致死率は90%近くに及ぶこともある。マクロファージや樹状細胞および肝細胞はウイルスの生体内における標的細胞であり、これらの細胞への感染が病原性発現に関与していると考えられている。フィロウイルスの表面糖蛋白質GPはこれらの細胞に発現しているC型レクチンと結合してウイルスの侵入効率を上げることが知られている。本研究では、フィロウイルスのプロトタイプであるマールブルグウイルス(MARV)のC型レクチン介在性細胞侵入機構について、病原性の異なる2つの株を用いて解析を行った。MARVのGPを持つシュードタイプウイルスを、ヒトのガラクトース型C型レクチンhMGLまたは樹状細胞に発現するC型レクチンDC-SIGNを発現させたK562細胞に接種し、フローサイトメーターを用いてGFP陽性のウイルス感染細胞数から各細胞に対する感染価を測定した。霊長類に強い病原性を示すAngola株のGPを持つシュードタイプウイルスは、比較的弱い病原性のMusoke株のGPを持つものよりもレクチン発現細胞への感染性が高いことが判明した。また、547番目のアミノ酸(Angola株 : グリシン、Musoke株 : バリン)がレクチン介在性の細胞侵入効率を決定していることが明らかとなった。MARVの病原性とレクチン発現細胞への感染性との間に相関が見られたことから、...
  • 文部科学省:科学研究費補助金(萌芽研究)
    Date (from‐to) : 2007 -2008 
    Author : 吉田 玲子, 高田 礼人, 鈴木 定彦
     
    ウイルス感染症やがんの有効な遺伝子治療法を開発するために、モノクローナル抗体を利用した標的細胞に効率よく目的遺伝子を導入できるターゲティングウイルス(標的指向性ウイルス)の作製を目指した。本研究ではインフルエンザ感染に対するターゲティングウイルスを作製することを目的に、インフルエンザウイルスHAに対するモノクローナル抗体のFab領域を発現するプラスミド、および膜融合タンパク質としてセンダイウイルスFタンパク質を発現するプラスミドを構築した。293T細胞にプラスミドを導入したところ、タンパク質の単独および共発現が確認できた。さらに、抗体と融合タンパク質を細胞に発現させ、G遺伝子をGFPに置換したVSVを感染させて、増殖したウイルスを回収した。作出したウイルス粒子の構造を電子顕微鏡で観察するとともに、抗体とFタンパク質のウイルス粒子内への取り込みを、SDS-PAGEとWestern blotで解析したところ、抗体タンパク質のウイルス粒子への取り込み効率が極めて低いことが判明した。また、ターゲティングウイルスとしての活性も認められなかった。そこで、抗体タンパク質のウイルス取り込み効率を上昇させるために、VSV-Gタンパク質およびFタンパク質が三量体であることから、抗体タンパク質を三量体で発現させることを試みた。抗体遺伝子の3'側にVSV-Gタンパク質またはFタンパク質の細胞外領域4...
  • Ministry of Education, Culture, Sports, Science and Technology:Grants-in-Aid for Scientific Research(基盤研究(B))
    Date (from‐to) : 2005 -2008 
    Author : Taisuke HORIMOTO, Ayato TAKADA, Hideo GOTOH, Jiro YASUDA, Masayuki SHIMOJIMA, Ayato TAKADA, Jiro YASUDA, Masayuki SHIMOJIMA
     
    人畜共通新興再興感染症は人類の脅威である。特に、H5N1 高病原性鳥インフルエンザの世界的な蔓延とヒトへの感染は、インフルエンザの新たな世界的大流行(パンデミック) を危惧させている。本研究では、こういった世界情勢を鑑み、H5N1 ワクチン開発のための基礎研究を実施した。その結果、不活化ワクチン製造のためのシードウイルスの発育鶏卵ならびにMDCK 細胞における増殖基盤を明らかにし、その知見をもとに高増殖性シードウイルスの作出に成功した。本成果は、今後のインフルエンザワクチン開発におおいに貢献することが期待される。
  • 文部科学省:科学研究費補助金(萌芽研究)
    Date (from‐to) : 2004 -2006 
    Author : 高田 礼人, 堀本 泰介
     
    インフルエンザウイルスは、表面糖蛋白質ヘマグルチニン(HA)およびノイラミニダーゼ(NA)の抗原性によって様々な亜型に分けられる。現在まで、インフルエンザワクチンは主にウイルスHAに対する血中抗体を誘導することを目的としてきた。しかし、現行の不活化インフルエンザワクチンは抗原性が異なるHA亜型のウイルスには全く効果がない。本研究の目的はこれを克服し、全てのA型インフルエンザに有効な免疫法を検討する事である。これまでに、ホルマリンで不活化したインフルエンザワクチンをマウスの鼻腔内に投与すると、様々な亜型のウイルスに対して交差感染防御が成立することを明らかにした。これにはウイルス表面糖蛋白質に対する亜型特異的中和抗体以外の免疫応答が関与していると考えられた。免疫したマウスのB細胞を用いてハイブリドーマを作出した結果、ウイルス蛋白質に対するIgAおよびIgG抗体を産生するハイブリドーマクローンが多数得られる事が判った。これらの中には様々な亜型のウイルスに交差反応性を示す抗体があった。H1、H2、H3およびH13亜型のHAをもつウイルスに交差反応性を示す中和抗体が得られたため、そのエピトープを同定した結果、このモノクローナル抗体はHAがレセプターに結合する領域の近傍を認識する事が明らかとなった。この領域の構造は亜型に関わらず類似性が高いため、交差反応性を示すことが推測された。これらの...
  • 文部科学省:科学研究費補助金(若手研究(A))
    Date (from‐to) : 2004 -2006 
    Author : 高田 礼人
     
    これまでに、エボラウイルスなどの新興感染症は世界の限られた地域でしか認められていないが、昨今の急激な国際化による人の移動および動植物の輸出入に伴い、それらの疾病の原因病原体が他国に拡散する可能性が高まっている。さらに近年、エボラウイルスのような致死率の高い出血熱ウイルスがバイオテロリズムの手段として使用される危険性が高まっており、対策を講じる必要がある。しかし、ウイルス性出血熱に対する効果的な医療手段はほとんどなく、予防・治療法を開発する事が急務となってきた。これまでに申請者は、エボラウイルスZaire株の表面糖蛋白質(GP)分子はウイルスの感染性を中和する抗体および増強する抗体の両方の標的である事を証明した。そこで、エボラウイルスの病原性の強さと感染増強抗体との関わりを調べるために、病原性の非常に強いZaireウイルスと病原性の比較的弱いRestonウイルスの間で、感染増強抗体誘導能を比較した。ZaireウイルスとRestonウイルスGPに対するマウスの抗血清をそれぞれ作成し、血清中の感染増強活性および中和活性を調べたところ、中和活性には殆ど差が無かったのに対して、感染増強活性はZaireウイルスの血清の方が優位に高かった。これは、感染増強抗体が結合できる抗原決定基の種類と数が両ウイルスの間で異なることを示している。また、この感染増強活性は主に血清中のIgG2a抗体によるも...
  • 文部科学省:科学研究費補助金(特定領域研究)
    Date (from‐to) : 2004 -2005 
    Author : 高田 礼人
     
    これまでに、エボラウイルスなどの新興感染症は世界の限られた地域でしか認められていないが、昨今の急激な国際化による人の移動および動植物の輸出入に伴い、それらの疾病の原因病原体が他国に拡散する可能性が高まっている。さらに近年、エボラウイルスのような致死率の高い出血熱ウイルスがバイオテロリズムの手段として使用される危険性が高まっており、対策を講じる必要がある。しかし、ウイルス性出血熱に対する効果的な医療手段はほとんどなく、予防・治療法を開発する事が急務となってきた。しかし、エボラウイルスの強い病原性に関わる因子は解明されていない。本ウイルスに対して効果的な予防・治療法を開発するためには、病原性発現のメカニズムを個体、細胞および分子レベルに至るまで明らかにする事が必要である。これまでに申請者は、エボラウイルスZaire株の表面糖蛋白質GP分子はウイルスの感染性を中和する抗体および増強する抗体の両方の標的である事を証明した。そこで、病原性の強いZaireウイルスと病原性の弱いRestonウイルスに対する抗血清を作出し、感染増強活性を比較したところ、補体成分C1qを介した感染増強効果およびFcレセプターを介した感染増強効果のいずれもZaireウイルスに対する抗血清の方が高かった。一方、中和活性に差は認められなかった。以上の成績は、抗体依存性感染増強現象がエボラウイルスの病原性発現に関わっ...
  • 文部科学省:科学研究費補助金(特定領域研究)
    Date (from‐to) : 2003 -2003 
    Author : 高田 礼人
     
    エボラウイルスの強い病原性に関わる因子は解明されていない。本ウイルスに対して効果的な予防・治療法を開発するためには、病原性発現のメカニズムを個体、細胞および分子レベルに至るまで明らかにする事が必要である。エボラウイルスZaire株の表面糖蛋白質(GP)に対する抗体の中には、マクロファージ等の細胞で抗体依存性感染増強現象を引き起こすものがある。合成ペプチドおよびキメラGPを用いて、これらの抗体が認識するエピトープの同定を行った結果、少なくとも4箇所の異なるエピトープの存在が明らかになった。マクロファージや未成熟樹状細胞に発現し、galactoseおよびN-acetylgalactosamineに特異的に結合するC-typeレクチン(hMGL)を培養細胞に発現させると、エボラウイルスの感染性が上昇する事を発見した。hMGLはエボラウイルスGP分子上のO-linkの糖鎖を認識している事が示唆された。マクロファージおよび樹状細胞等の抗原提示細胞にエボラウイルスが感染する事によって起こる免疫応答の異常が本ウイルス感染の病原性に深く関わっていると考えられている。本ウイルスはこれらの細胞に効率よく感染するために、抗体、補体あるいはレクチンを利用し、ウイルスレセプターへの吸着効率を上げると考えられる。従って、これらの因子が本ウイルスの病原性発現において重要な役割を演じている事が推察される。マ...
  • Ministry of Education, Culture, Sports, Science and Technology:Grants-in-Aid for Scientific Research(基盤研究(C))
    Date (from‐to) : 2002 -2003 
    Author : Taisuke HORIMOTO, 高田 礼人, 五藤 秀男
     
    Progeny viruses are rapidly produced from the cells infected with influenza A viruses for a short period. A several cellular factors regulate virus replication in cells, interacting with viral proteins or RNAs directly or indirectly. In this study, we sought to identify and analyze cellular factors responsible for virus replication. It is hopeful that such information would contribute one to develop novel anti-influenza drugs in the future. To this end, we attempted to establish and use new methodology to identify cellular factors ; that is, "self-motivating" recombinant virus system. In th...
  • Ministry of Education, Culture, Sports, Science and Technology:Grants-in-Aid for Scientific Research(基盤研究(B))
    Date (from‐to) : 2001 -2003 
    Author : Hideo GOTO, 高田 礼人, 堀本 泰介, 河岡 義裕, 近藤 高志
     
    Equine influenza is an important contagious respiratory disease of horses. In the field of veterinary medicine, equine influenza is recognized as a serious problem because it causes economical loss. Inactivated vaccine for equine influenza has been used to control infection; however its efficacy is not satisfactory.The aim of this study is to develop a live attenuated vaccine for equine influenza infection. A live attenuated vaccine for influenza virus can be generated by insertion of a gene encoding target protein into genome of an attenuated virus. To this end we established a strategy to...
  • 文部科学省:科学研究費補助金(特定領域研究)
    Date (from‐to) : 2002 -2002 
    Author : 高田 礼人
     
    エボラウイルスの強い病原性に関わる因子は解明されていない。本ウイルスに対して効果的な予防・治療法を開発するためには、病原性発現のメカニズムを個体、細胞および分子レベルに至るまで明らかにする事が必要である。エボラウイルスの表面糖蛋白質(GP)に対して誘導される抗体の中には、ウイルスの感染を増強するものが存在する事をこれまでに明らかにした。感染患者回復期血清を調べると、約半数に感染増強活性が認められたので、実際の感染でも感染増強抗体の産生が誘導されている事が明らかになった。GPに対するモノクローナル抗体(MAb)のうち、ウイルスを中和する活性を示すものに関して、エピトープの同定を行った。VSVのG蛋白質遺伝子をZaire株GP遺伝子に置換したリコンビナントVSVを中和抗体存在下で増殖させてエスケープミュータントを選択し、GPのアミノ酸変異を同定したところ、MAb133/3.16は膜融合ペブチド、MAb226/8.1はレセプター結合領域付近のエピトープを認識すると考えられた。これらの抗体の受動免疫による感染防御効果をマウスモデルを用いて調べた。抗体をウイルス攻撃前あるいは後に腹腔内に投与したマウスは致死量のウイルス感染に対して無症状で耐化した。ウイルス感染後に抗体価の上昇が全く認められなかった事から、ウイルスはマウス体内で殆ど増殖しなかったと考えられる。これらの結果は、中和抗体のみ...
  • Ministry of Education, Culture, Sports, Science and Technology:Grants-in-Aid for Scientific Research(基盤研究(A))
    Date (from‐to) : 2000 -2002 
    Author : Hiroshi KIDA, 高田 礼人, 河岡 義裕, 岡崎 克則, 伊藤 壽啓, 迫田 義博
     
    To provide information on the precursor genes of the next pandemic influenza, epidemiological surveillance of animal and avian influenza viruses was performed. In total, 3,987 fecal samples of migratory waterfowls were collected in Russia, Mongolia, China and Holdcaido, Japan. From these samples, 212 influenza A viruses were isolated H1N2 swine influenza viruses were isolated from pigs with respiratory syndrome in Miyagi Prefecture, Japan. Twenty-five strains of H9N2 viruses were exchanged between 5 centers of global animal influenza network and stated analyzing these viruses.Genetic analys...
  • 文部科学省:科学研究費補助金(特定領域研究(C))
    Date (from‐to) : 2001 -2001 
    Author : 高田 礼人
     
    エボラウイルスの高病原性に関わる因子は解明されていない。本ウイルスに対して効果的な予防・治療法を開発するためには、病原性発現のメカニズムを個体、細胞および分子レベルに至るまで明らかにする事が必要である。本研究では、申請者らが既に確立したVSVシステムおよびウイルスをプラスミドDNAから作出するリバースジェネテイクス法を用いて、病原性に関わる可能性のある蛋白質について変異ウイルスを作出し、ウイルス増殖および病原性発現に関わるウイルス蛋白質の同定を行う。また、マウス実験感染モデルを用いて、エボラウイルスの病原性についてウイルス側のみならず宿主因子をも含めた総合的な解析を行う。GPに対するモノクローナル抗体を作出した。いくつかの抗体は、正常マウス血清存在下でウイルスの感染性を増強した・正常血清の代わりに補体成分C1qを加えると同様の感染増強効果が認められることから、感染増強のメカニズムはGPと抗体の複合物にC1qが結合し標的細胞表面に存在するC1qレセプターを介してウイルスと細胞を架橋し、効率よくウイルスレセプターに結合させる事、あるいはC1qレセプターを介した細胞内シグナルによって細胞の食作用の活性化が起こる事によると考えられた。エボラウイルスのGPは蛋白質分解酵素によって、GP1とGP2に開裂される。ある種のウイルスでは表面糖蛋白質の開裂はウイルスが感染性を獲得するために必要で...
  • 文部科学省:科学研究費補助金(萌芽的研究)
    Date (from‐to) : 2001 -2001 
    Author : 河岡 義裕, 高田 礼人, 五藤 秀男, 堀本 泰介
     
    申請者らが開発したリバースジェネティクス法を用いて、安全性の高い不活化ワクチンと、侵入門戸の感染防御に有効な弱毒化生ワクチン双方の長所を兼ね備えたワクチン、すなわち細胞に一度だけ感染してウイルス蛋白質を発現し、粘膜免疫および細胞性免疫は誘導するが新たな感染性粒子は産生しない、"半生"ワクチンの開発を試みた。本研究では、インフルエンザウイルスの8本のRNA分節のうち、NS遺伝子に着目した。NS遺伝子がコードするNS2蛋白質を発現しないように変異を導入し、これを用いてNS2欠損ウイルスを作製した。NS2欠損ウイルスを培養細胞に感染させると、主要ウイルス抗原蛋白質であるNP, HA, NA, M1の発現は確認されたが、新たな感染性ウイルス粒子の産生は認められなかった。このウイルスをワクチンとして、3週間おきに3回マウスに経鼻接種して、血中と鼻腔および肺洗浄液中の抗体価を調べたところ、血中からはIgG抗体が、肺洗浄液中からはIgA抗体が検出された。続いて最終免疫から1ヶ月後あるいは3ヶ月後に、致死量の100倍のウイルスでマウスを攻撃した。最終免疫から1ヵ月後に攻撃した場合、対照群のマウスは全て死亡したのに対し、半生ワクチン接種マウスは全て生残した。さらに、最終免疫から3ヵ月後に攻撃した場合でも、半生ワクチン接種マウスは9匹中8匹が生残した。以上の結果は、増殖能を欠く"半生"ワクチンは...
  • 文部科学省:科学研究費補助金(奨励研究(A))
    Date (from‐to) : 2000 -2001 
    Author : 高田 礼人
     
    現在までに多くのウイルス感染症は予防、制圧されてきたが、エイズ、ヘルベス、インフルエンザあるいはエボラ出血熱等の感染症に対する効果的な治療法は未だ確立されていない。本研究の目的は特定のアミノ酸配列をもつ合成ベプチドによってウイルスの感染を選択的に阻害する方法を開発する事である。pSKANファージディスプレイシステムを用いて、現在までにインフルエンザウイルス蛋白質に特異的に結合するファージを選択した。また、エボラウイルスの表面糖蛋白をプラスミドから発現させ、精製する事に成功した。これを用いて、この糖蛋白に特異的に結合するファージを選択した。さらに、エボラウイルス糖蛋白の幹部に存在する螺旋状部位が機能的に重要である事が判明したので、その部位と同様のアミノ酸配列を有する合成ペプチドを作成し、エボラウイルス表面糖蛋白でシュードタイプした豚水泡性口炎ウイルスを用いてウイルス感染性中和試験を行った。その結果、約80%のウイルスの侵入が阻止された。この成績は、この合成ペプチドがエボラウイルス表面糖蛋白の立体構造の変化をさまたげ、その侵入を特異的に阻害したためと考えられる。また、エボラウイルスの糖蛋白に対して誘導される抗体の中には、ウイルスの感染性を増強するものがあり、エボラウイルスの強い病原性に関わっている事が示唆された。したがって、この抗体が認識するエピトープを解析し、そのアミノ酸配列を...
  • 文部科学省:科学研究費補助金(奨励研究(A))
    Date (from‐to) : 1998 -1999 
    Author : 高田 礼人
     
    オーエスキー病ウイルスの表面糖蛋白gBを発現するプラスミドを構築した。これに市販のリポソームとコレラトキシンBサブユニットをアジュバントとして加えたワクチンをマウスに接種し、抗体応答および感染防御効果を調べた。鼻腔内にワクチンを接種したマウスの呼吸器分泌液中にgB特異IgA抗体が血清中にIgG抗体が検出された。これらのマウスの40%が致死量のウイルス攻撃に対して生残した。また、死亡したマウスでも生残時間の延長が認められた。一方、筋肉内にワクチンを接種したマウスでは、血清中にIgG抗体が検出されたが、呼吸器分泌液中には抗体が検出されなかった。これらのマウス全てがウイルス攻撃後、コントロールのマウス同様の経過で死亡した。gB遺伝子を組み込んだリコンビナントバキュロウイルスをマウスの鼻腔内に接種すると同様の抗体応答を誘導することが判明した。これらの成績はgBを動物の粘膜細胞に発現させるワクチンがオーエスキー病の予防に有効である事を示唆している。インフルエンザウイルス(A/Aichi/2/68)のヘマグルチニンを発現するプラスミドを構築した。これを用いてDNAワクチンの粘膜投与における抗CD40抗体または多重膜リボソームのアジュバント効果を検討した。CD40分子は抗原提示細胞の表面糖蛋白であり、これを刺激する事によって免疫応答を増強する事が期待された。抗CD40抗体を混合したワクチン...
  • Ministry of Education, Culture, Sports, Science and Technology:Grants-in-Aid for Scientific Research(国際学術研究, 基盤研究(A))
    Date (from‐to) : 1998 -1999 
    Author : Hiroshi KIDA, 河岡 義裕, 伊藤 壽啓, 高田 礼人, 岡崎 克則
     
    Influenza A viruses of different subtypes were isolated from fecal samples of ducks in their nesting areas in Siberia. Phylogenetic analysis of the NP genes of the isolates in Siberia and those in Hokkaido, Japan on their flyway of migration from Siberia to the south in autumn revealed that they belong to the Eurasian lineage of avian influenza viruses. It is noted that the genes of the isolates in Siberia are closely related to those of H5N1 influenza virus strains isolated from chickens and humans in Hong Kong in 1997 as well as to those of isolates from domestic birds in southern China. ...
  • 文部科学省:科学研究費補助金(萌芽的研究)
    Date (from‐to) : 1998 -1999 
    Author : 岡崎 克則, 高田 礼人, 喜田 宏
     
    ヘルペスウイルス主要糖蛋白の機能を明らかにするためウシヘルペスウイルス1(BHV1)の糖蛋白gBあるいはgCを発現する株化細胞を樹立し、これらにG蛋白遺伝子を欠失した水疱性口炎ウイルス(VSVdelG-G)を感染させて表現型混合ウイルスの回収を試みた。培養上清中に放出されたウイルス粒子をMDBK細胞に接種し、感染性を獲得したか否かをGFPの発現によって調べた。それぞれの糖蛋白を単独で導入した場合あるいはトランスフェクションによってgB、gC、gDの3種を導入した場合にもVSVdelG-Gは感染性を獲得しなかった。したがって、ヘルペスウイルスが宿主細胞に感染する際にはgB、gCおよびgD以外のウイルス蛋白を必要とすることが考えられる。ヘルペスウイルス表面糖蛋白のリセプター特異性を明らかにするため、BHV1gCおよびgBを単独で発現して各種グルコサミノグリカン分子との結合活性を調べた。両糖蛋白はヘパリン、ヘパラン硫酸およびコンドロイチン硫酸Bと結合した。ヘパリン、ヘパラン硫酸およびコンドロイチン硫酸Bはウロン酸としてイヅロン酸を含み、コンドロイチン硫酸AおよびCはグルクロン酸のみを含む。したがって、gCおよびgBは[イヅロン酸-硫酸化アミノ糖]の繰り返し構造を認識するものと考えられる。また、ヘパリンおよびヘパラン硫酸のアミノ糖はグルコサミンであるのに対し、コンドロイチン硫酸群のそ...
  • Ministry of Education, Culture, Sports, Science and Technology:Grants-in-Aid for Scientific Research(基盤研究(A))
    Date (from‐to) : 1996 -1998 
    Author : Hiroshi KIDA, 中里 幸和, 板倉 智敏, 伊藤 壽啓, 岡崎 克則, 前出 吉光, 藤田 正一, 高田 礼人, 梅村 孝司, 中里 幸和
     
    Intravenous injection into chickens with the virus-free extracts from the allantoic membrane of chicken embryos infected with influenza viruses resulted in sudden death due to blood coagulation in the vessels. The chickens injected intravenously with heparin prior to the injection with the extract survived without any signs of disease. The results suggest that some coagulation factors exist in the tissues where virus replicated and that they showed toxiciry to the chickens.To provide information on the pathogenically of fowl plague virus, virological and histopathological comparison was car...
  • Ministry of Education, Culture, Sports, Science and Technology:Grants-in-Aid for Scientific Research(基盤研究(B))
    Date (from‐to) : 1996 -1997 
    Author : Katsunori OKAZAKI, 喜田 宏, 高田 礼人
     
    An escape mutant of bovine herpesvirus 1 (BHV1) was obtained in the presence of monoclonal antibody against glycoprotein B (gB) , which required complement for virus neutralization. The mutant was also resistant to other 3 monoclonal antibodies against gB,which exhibited complement-independent neutralizing activity. Amino acid substitution of gB of the mutant is now being analyzed. The cytotoxic activity of peripheral blood lymphocytes from BHV1-infected cattle is also being investigated using the mutant-infected primary culture as a target to study whether the amino acid substitution affec...
  • Ministry of Education, Culture, Sports, Science and Technology:Grants-in-Aid for Scientific Research(国際学術研究)
    Date (from‐to) : 1995 -1997 
    Author : Hiroshi KIDA, 伊藤 壽啓, 河岡 義裕, 岡崎 克則, デメネフ V, ヤムニコバ S, ルボフ D.K, Svetrana Yam, 高田 礼人, Yamnikova Sv
     
    In each summer of 1995-1997,3,000 fecal materials of waterfowls and lake-water samples were collected in Kamchatka, Khabarovsk, and Saha Republic. In 1996, out of 900 fecal samples collected in Kobyaysky area of the basin of the River Lena, 60゚30'N,19 H4N6,1 H4N9,1 H11N1,2 H11N6, and 8 H11N9 influenza viruses were isolated. Although no virus was isolated from the samples collected in "40 islands" area of the River Lena (65゚00'-64゚34'N) , 1 H4N6 and 5 H3N8 viruses were isolated from 120 samples collected in Kobyaysky area and 72 samples in Yakutsk (62゚05'N) , respectively, in 1997. No influe...
  • エボラウイルスの感染機構の解明
  • ウィルス感染症に対するワクチンの研究
  • Mechanism of Ebola Virus Replication
  • Study on Mucosal Vaccination of Animals against Viral Infections

Educational Activities

Teaching Experience

  • 感染症学特別研究Ⅰ
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 国際感染症学院
  • Microbiology
    開講年度 : 2018
    課程区分 : 学士課程
    開講学部 : 医学部
    キーワード : 細菌、ウイルス、グラム陽性菌、グラム陰性菌、抗酸性菌、感染症、ワクチン、プリオン、寄生虫、原虫、マイコプラズマ、クラミジア、スピロヘータ、真菌
  • 感染症学特別演習
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 国際感染症学院
  • Zoonotic Science
    開講年度 : 2018
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 人獣共通感染症、新興・再興感染症、疫学、自然宿主、媒介動物、感染環、ウイルス、細菌、原虫、寄生虫 
  • 人獣共通感染症対策専門特論
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 国際感染症学院
  • 感染症学特別研究ⅡA
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 国際感染症学院
  • 感染症学特別研究ⅡB
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 国際感染症学院
  • 獣医科学・感染症学基礎科目 微生物学特論
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 国際感染症学院
  • 国際感染症学コア科目Ⅰ 人獣共通感染症制御学特論
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 国際感染症学院
  • 獣医科学特別研究
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 専門獣医科学特論
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 人獣共通感染症対策専門特論
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 獣医科学特論演習
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 獣医科学基礎科目B 獣医微生物学特論
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 獣医科学基礎科目A 人獣共通感染症制御学特論
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 獣医科学基礎科目 微生物学特論
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 獣医学院
  • 専門獣医科学特論
    開講年度 : 2018
    課程区分 : 博士後期課程
    開講学部 : 獣医学院

Campus Position History

  • 2017年4月1日 
    2019年3月31日 
    大学院国際感染症学院副学院長
  • 2019年4月1日 
    2021年3月31日 
    大学院国際感染症学院副学院長

Position History

  • 2017年4月1日 
    2019年3月31日 
    大学院国際感染症学院副学院長
  • 2019年4月1日 
    2021年3月31日 
    大学院国際感染症学院副学院長

Social Contribution

Social Contribution

Social Contribution

  • JICAの現場から 日本の医療技術に高評価
    Date (from-to) : 2018/12/21
    Role : Informant
    Event, Program, Title : 日刊工業新聞
  • デンカ生研、エボラウイルス迅速診断キット(クイックナビシリーズ)試作品をコンゴ民主共和国へ提供
    Date (from-to) : 2018/05/28
    Role : Others
    Sponser, Organizer, Publisher  : 日本経済新聞
    Event, Program, Title : 日本経済新聞(電子版)
  • 医師10万人アンケートでわかった!
    Date (from-to) : 2018/01/27
    Role : Advisor
    Sponser, Organizer, Publisher  : 日本テレビ
    Event, Program, Title : 世界一受けたい授業
  • 人獣共通感染症的視点から 鳥インフルエンザとエボラ出血熱を例に
    Date (from-to) : 2017/11/02
    Role : Lecturer
    Sponser, Organizer, Publisher  : 神津教育研究会
    Event, Program, Title : 神津教育研究会「日本の国際協力の現状と成果・課題」
  • 人間が“ゾンビ”になることはあるのか?
    Date (from-to) : 2017/09/05
    Role : Advisor
    Sponser, Organizer, Publisher  : NHK
    Event, Program, Title : モーガン・フリーマン 時空を超えて
  • アフリカにおけるウイルス性 人獣共通感染症の調査研究
    Date (from-to) : 2017/08/28
    Role : Lecturer
    Sponser, Organizer, Publisher  : SATREPS
    Event, Program, Title : SATREPS 科学と開発をつなぐブリッジ・ワークショップ
  • エボラ出血熱など、人獣共通感染症ウイルスとどう向き合うか
    Date (from-to) : 2017/07/12
    Role : Appearance
    Sponser, Organizer, Publisher  : 文化放送
    Event, Program, Title : ブンナビ薬学特別企画2017
  • デンカ生研と北大、コンゴ民主共和国へエボラウイルス迅速診断キットを提供
    Date (from-to) : 2017/05/30
    Role : Informant
    Sponser, Organizer, Publisher  : 日本経済新聞
    Event, Program, Title : 日本経済新聞(電子版)
  • 人獣共通感染症研究「ウイルス研究の最前線」
    Date (from-to) : 2016/11/17
    Role : Lecturer
    Sponser, Organizer, Publisher  : とわの森三愛高等学校
    Event, Program, Title : とわの森三愛高等学校 獣医進学コース 講義
  • エボラ出血熱への挑戦
    Date (from-to) : 2016/10/24
    Role : Appearance
    Sponser, Organizer, Publisher  : 文化放送
    Event, Program, Title : ブンナビ薬学特別企画2016
  • ウイルス研究の最前線 -インフルエンザとエボラ出血熱の話-
    Date (from-to) : 2016/08/04
    Role : Lecturer
    Sponser, Organizer, Publisher  : 北海道ハイテクノロジー専門学校
    Event, Program, Title : 北海道ハイテクノロジー専門学校 平成28年度高校教員バイオ講習会
  • 人獣共通感染症 -インフルエンザとエボラ出血熱の話-
    Date (from-to) : 2016/08/02
    Role : Lecturer
    Event, Program, Title : 北海道大学 平成遠友夜学校
  • ウイルスの生態
    Date (from-to) : 2016/06/04
    Role : Lecturer
    Sponser, Organizer, Publisher  : 北海道大学
    Event, Program, Title : 第58回北大祭公開講座
  • エボラ熱の抗体発見 ウイルス全5種抑制
    Date (from-to) : 2016/04/05
    Role : Informant
    Sponser, Organizer, Publisher  : 北海道新聞
    Event, Program, Title : 北海道新聞
  • ウイルスの生態
    Date (from-to) : 2016/03/19
    Role : Panelist
    Sponser, Organizer, Publisher  : 人間文化研究機構
    Event, Program, Title : 人間文化研究機構広領域連携型基幹研究プロジェクト キックオフ・シンポジウム
  • ウイルスって何?-エボラとインフルエンザの話-
    Date (from-to) : 2016/03/08
    Role : Lecturer
    Sponser, Organizer, Publisher  : 北海道旭川西高等学校
    Event, Program, Title : 平成27年度北海道旭川西高等学校「SSH講演会」
  • エボラウイルスの抗体発見 北大チーム
    Date (from-to) : 2016/02/15
    Role : Informant
    Sponser, Organizer, Publisher  : 読売新聞社
    Event, Program, Title : 読売新聞
  • ウイルスはどうやって生き残っているのか
    Date (from-to) : 2015/11/18
    Role : Lecturer
    Sponser, Organizer, Publisher  : 慶應学術事業会
    Event, Program, Title : 慶應丸の内シティキャンパス定例講演会『夕学五十講』
  • エボラウイルス
    Date (from-to) : 2015/11/13
    Role : Lecturer
    Sponser, Organizer, Publisher  : 宮崎大学
    Event, Program, Title : 宮崎大学 第5回国際シンポジウム
  • 人獣共通感染症の研究最前線 ~エボラ出血熱とインフルエンザ~
    Date (from-to) : 2015/10/18
    Role : Lecturer
    Sponser, Organizer, Publisher  : サイエンステクノフロンティアフォーラム
    Event, Program, Title : 第91回サイテックサロン
  • 人獣共通感染症って?-エボラ出血熱とインフルエンザの話-
    Date (from-to) : 2015/09/29
    Role : Lecturer
    Sponser, Organizer, Publisher  : 札幌啓成高等学校
    Event, Program, Title : 平成27年度スーパー・サイエンス・ハイスクール特別科学講演会
  • 協和発酵キリン 北大とエボラ抗体薬開発 ウイルス全種に効果期待
    Date (from-to) : 2015/09/16
    Role : Informant
    Sponser, Organizer, Publisher  : 化学工業日報社
    Event, Program, Title : 化学工業日報
  • Recent Advance in Ebolavirus Research: Epidemiology & Virology
    Date (from-to) : 2015/08/12
    Role : Lecturer
    Sponser, Organizer, Publisher  : Airlangga University
    Event, Program, Title : AIRC International Seminar: Bioterrorism and Agroterrorism in Indonesia
  • エボラウイルス ー研究の現状と展望ー
    Date (from-to) : 2015/08/05
    Role : Lecturer
    Sponser, Organizer, Publisher  : 日本学術会議第二部(生命科学分野)部会
    Event, Program, Title : 公開学術講演会
  • エボラ出血熱への挑戦
    Date (from-to) : 2015/07/28
    Role : Appearance
    Sponser, Organizer, Publisher  : 文化放送キャリアパートナーズ 帝京大学
    Event, Program, Title : ブンナビ薬学特別企画2015
  • エボラウイルス研究の最前線
    Date (from-to) : 2015/07/20
    Role : Panelist
    Sponser, Organizer, Publisher  : 北海道大学
    Event, Program, Title : 平成27年度北海道大学公開講座
  • エボラ感染 15分で判定 北大教授ら キット開発、実用化へ
    Date (from-to) : 2015/04/01
    Role : Informant
    Sponser, Organizer, Publisher  : 北海道新聞
    Event, Program, Title : 北海道新聞
  • エボラ感染 15分で検査 北大教授ら キット開発
    Date (from-to) : 2015/04/01
    Role : Informant
    Sponser, Organizer, Publisher  : 読売新聞
    Event, Program, Title : 読売新聞
  • エボラ出血熱:現状と研究の最前線
    Date (from-to) : 2015/03/02
    Role : Lecturer
    Sponser, Organizer, Publisher  : 岐阜大学
    Event, Program, Title : 岐阜大学市民講座
  • エボラウイルスに迫る
    Date (from-to) : 2015/02/24
    Role : Panelist
    Sponser, Organizer, Publisher  : 北海道大学
    Event, Program, Title : 北海道大学 創成研究機構第 12 回 創成シンポジウム 感染症研究の最前線 ― エボラ・結核を例に―
  • エボラおよびマールブルグウイルスによる感染症
    Date (from-to) : 2015/01/15
    Role : Panelist
    Sponser, Organizer, Publisher  : 厚生労働省
    Event, Program, Title : 平成26年度新型インフルエンザ等新興・再興感染症研究推進事業シンポジウム
  • エボラ出血熱とBSL-4
    Date (from-to) : 2014/09/21
    Role : Lecturer
    Sponser, Organizer, Publisher  : 長崎大学熱帯医学研究所
    Event, Program, Title : 長崎大学熱帯医学研究所 「市民公開特別講座」
  • 北海道大学ザンビア拠点での取り組み
    Date (from-to) : 2014/08/23
    Role : Panelist
    Sponser, Organizer, Publisher  : J-GRID
    Event, Program, Title : J-GRID市民公開講演会「いま話題の感染症-SFTS、MERS、エボラ出血熱」
  • 鳥インフルエンザ
    Date (from-to) : 2013/12/18
    Role : Lecturer
    Sponser, Organizer, Publisher  : 石狩家畜保健衛生所
    Event, Program, Title : 平成25年度家畜保健衛生所病性鑑定技術検討会
  • 「エボラ出血熱ウイルス」…糸のような形の、やさしそうで怖いウイルスの話
    Date (from-to) : 2013/06/13
    Role : Lecturer
    Sponser, Organizer, Publisher  : 日本ウイルス学会
    Event, Program, Title : みちのくウイルス塾
  • 海外のBSL4 施設での実験の状況
    Date (from-to) : 2012/11/14
    Role : Lecturer
    Sponser, Organizer, Publisher  : 日本学術会議
    Event, Program, Title : 日本学術会議 公開シンポジウム
  • 人獣共通感染症としてのインフルエンザ
    Date (from-to) : 2012/11/09
    Role : Lecturer
    Sponser, Organizer, Publisher  : 北海道青少年科学文化財団
    Event, Program, Title : 第21回先端科学移動大学2012
  • H5N1高病原性鳥インフルエンザウイルスと私との関わり
    Date (from-to) : 2012/10/27
    Role : Lecturer
    Sponser, Organizer, Publisher  : 宮崎大学
    Event, Program, Title : 第2回宮崎大学 鳥インフルエンザシンポジウム
  • 人獣共通感染症としてのインフルエンザ
    Date (from-to) : 2010/11/13
    Role : Lecturer
    Sponser, Organizer, Publisher  : 北海道青少年科学文化財団
    Event, Program, Title : 第19回先端科学移動大学2010
  • 人獣共通感染症としてのインフルエンザ
    Date (from-to) : 2009/09/16
    Role : Lecturer
    Sponser, Organizer, Publisher  : IUMS
    Event, Program, Title : 市民公開講座 微生物をよく知ろう
  • インフルエンザウイルスの病原性と宿主域
    Date (from-to) : 2008/01/22
    Role : Lecturer
    Sponser, Organizer, Publisher  : 自衛隊
    Event, Program, Title : 自衛隊北部防衛衛生学会
  • エボラウイルスの感染、免疫、疫学
    Date (from-to) : 2006/07/22
    Role : Lecturer
    Sponser, Organizer, Publisher  : 宮崎大学
    Event, Program, Title : 人獣共通感染症 教育公開フォーラム
  • Ayato Takada and the Ebola Virus
    Role : Others
    Sponser, Organizer, Publisher  : 三修社
    Event, Program, Title : 英語教科書

Media Coverage

  • 「鳥インフルエンザ 新たな脅威」
    Date : 2019/11/04
    Publisher, broadcasting station: NHK
    Program, newspaper magazine: サイエンスZERO
    Media report
  • 死に至る病「エボラ」から世界を救う…日本人ウイルス学者の奮闘記
    Date : 2019/01/15
    Program, newspaper magazine: 現代ビジネス
    Internet
  • ウイルスは悪者か
    Date : 2018/12/09
    Publisher, broadcasting station: NHK
    Program, newspaper magazine: 「マイあさラジオ」
    Media report
  • 「風疹、インフルエンザ、HIV、エボラ出血熱~ウイルス学者・高田礼人さんが語る~ウイルスとはいったい何なのか?」
    Date : 2018/11/28
    Publisher, broadcasting station: TBS
    Program, newspaper magazine: Session22
    Media report
  • The Battle Against Ebola
    Date : 2018/07/02
    Publisher, broadcasting station: NHK
    Program, newspaper magazine: Direct Talk
    Internet
  • 「伊集院光とらじおと」
    Date : 2018/06/04
    Publisher, broadcasting station: TBS
    Program, newspaper magazine: 「伊集院光とらじおと」 TBS
    Media report
  • A型インフルエンザウイルス—ウイルス学の視点から
    Date : 2018
    Publisher, broadcasting station: メディカルノート
    Program, newspaper magazine: メディカルノート
    Internet
  • 加計学園問題と獣医師の現状
    Date : 2017/06/10
    Publisher, broadcasting station: TBS
    Program, newspaper magazine: 報道特集
    Media report
  • スクラムトーク 日本はもっと人道で貢献できる ウイルスとの戦いに「世界市民」への道 哲学ある人材を大学から輩出したい
    Date : 2016/03/20
    Publisher, broadcasting station: 聖教新聞社
    Program, newspaper magazine: 聖教新聞
    Paper
  • 『緊急!池上彰と考える 今年の細菌・ウイルス大疑問』
    Date : 2016/01/27
    Publisher, broadcasting station: TBS
    Program, newspaper magazine: テレビ未来遺産
    Media report
  • エボラ出血熱への挑戦
    Date : 2015/10/25
    Publisher, broadcasting station: 文化放送キャリアパートナーズ 高崎健康福祉大学
    Program, newspaper magazine: ブンナビ薬学特別企画2015
    Pr
  • Ebola spurs creation of Japan's first maximum-security biolab
    Date : 2015/08/13
    Publisher, broadcasting station: Nature
    Program, newspaper magazine: Nature
    Paper
  • エボラ出血熱 適切な対応で感染は防げる
    Date : 2015/05/01
    Publisher, broadcasting station: 民医連
    Program, newspaper magazine: 民医連 いつでも元気
    Paper
  • エボラウイルス解明に挑戦
    Date : 2015/05/01
    Publisher, broadcasting station: 北海道建設新聞
    Program, newspaper magazine: 真砂徳子の起ーパーソン ~風をおこす人々~
    Paper
  • 未踏の世界へ エボラに効く抗体を追う
    Date : 2015/04/30
    Publisher, broadcasting station: 毎日新聞社
    Program, newspaper magazine: 毎日新聞
    Paper
  • Ebola Antibodies in Zambia Bats Match West African Virus
    Date : 2015/03/27
    Program, newspaper magazine: Bloomberg News
    Internet
  • ウイルスの専門家に聞く感染症とエボラウイルスの真実
    Date : 2015/03/13
    Publisher, broadcasting station: 洋泉社
    Program, newspaper magazine: 感染症クライシス
    Others
  • 日曜Navi ほっかいどう知究人 エボラ出血熱解明に挑む
    Date : 2015/03/08
    Publisher, broadcasting station: 北海道新聞
    Program, newspaper magazine: 北海道新聞
  • ウイルスと感染症
    Date : 2015/02/10
    Publisher, broadcasting station: 株式会社 ニュートンプレス
    Program, newspaper magazine: Newton 別冊
    Paper
  • エボラ出血熱、高病原性鳥インフルエンザ……人獣共通感染症を研究 エボラウイルスの抗体もーー高田礼人さんの挑戦
    Date : 2015/02/01
    Publisher, broadcasting station: 株式会社ビッグイシュー日本版
    Program, newspaper magazine: ビッグイシュー日本版
    Paper
  • ウイルス
    Date : 2015/01/31
    Publisher, broadcasting station: 日本テレビ
    Program, newspaper magazine: 世界一受けたい授業
    Media report
  • 脅威のエボラ、英知をかけて挑む ウイルス学者・高田礼人
    Date : 2015/01/05
    Publisher, broadcasting station: NHK
    Program, newspaper magazine: NHK プロフェッショナル~仕事の流儀
    Media report
  • 挑む Front Runner アフリカの森林でエボラウイルスを追う
    Date : 2015/01/01
    Publisher, broadcasting station: 日経サイエンス社
    Program, newspaper magazine: 日経サイエンス
    Paper
  • エボラ…日本の課題
    Date : 2014/11/29
    Program, newspaper magazine: 報道特集
    Media report
  • The Ebola Questions
    Date : 2014/10/30
    Publisher, broadcasting station: Nature
    Program, newspaper magazine: Nature
    Paper
  • 猛威を振るう出血熱ウイルス 感染ルートや生態は未解明 野生動物に寄生、撲滅困難
    Date : 2014/09/01
    Publisher, broadcasting station: 産経新聞
    Program, newspaper magazine: 産経新聞
    Paper
  • 北大、エボラ熱阻止支援 ザンビア 感染の疑い診断
    Date : 2014/08/17
    Publisher, broadcasting station: 北海道新聞
    Program, newspaper magazine: 北海道新聞
    Paper
  • エボラ早期診断 北大が一役
    Date : 2014/08/14
    Publisher, broadcasting station: 朝日新聞
    Program, newspaper magazine: 朝日新聞
    Paper
  • ウイルス学魂
    Date : 2013/12/22
    Publisher, broadcasting station: BS日テレ
    Program, newspaper magazine: 加藤浩次の本気対談!『コージ魂!!』
    Media report
  • Orang-utans infected by mystery Ebola-like virus
    Date : 2012/11/06
    Program, newspaper magazine: New Scientist
    Internet
  • 獣医師たちのたたかい こうもりからウイルスを探す
    Date : 2012/01/15
    Publisher, broadcasting station: 朝日新聞
    Program, newspaper magazine: 朝日新聞グローブ
    Paper
  • ウィルス学者・高田礼人
    Date : 2010/02/28
    Publisher, broadcasting station: 毎日放送
    Program, newspaper magazine: 情熱大陸
    Media report
  • インフルエンザ21世紀
    Date : 2009/12/20
    Publisher, broadcasting station: 文春新書
    Program, newspaper magazine: インフルエンザ21世紀
    Others
  • 新型インフルエンザ 専門家が予想…脅威の感染力
    Date : 2008/12/16
    Publisher, broadcasting station: 日本テレビ
    Program, newspaper magazine: スッキリ
    Media report
  • ウイルス その奇妙な生き方〜ウイルス学 高田礼人〜
    Date : 2008/06/12
    Publisher, broadcasting station: NHK
    Program, newspaper magazine: 爆笑問題のニッポンの教養
    Media report
  • シリーズ ミクロの生命体1 新型インフルエンザを阻止せよ!
    Date : 2005/11/06
    Publisher, broadcasting station: テレビ朝日
    Program, newspaper magazine: 素敵な宇宙船地球号
    Media report


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