Researcher Profile and Settings

Master

Affiliation (Master)

• International Institute for Zoonosis Control　Division of Global Epidemiology

Affiliation (Master)

• International Institute for Zoonosis Control　Division of Global Epidemiology

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Profile and Settings

Degree

• (BLANK)（Hokkaido University）

Profile and Settings

• Name (Japanese)

  Takada

• Name (Kana)

  Ayato

• Name

  200901079814217420

Achievement

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

Research 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

Awards

• 2022/02 Hokkaido University 教育研究総長表彰
   

  受賞者: 髙田 礼人
• 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
  

Published Papers

• Characterization of human tibrovirus envelope glycoproteins

  Yannick Munyeku-Bazitama, Takeshi Saito, Takanari Hattori, Hiroko Miyamoto, Boniface Pongombo Lombe, Akina Mori-Kajihara, Masahiro Kajihara, Jean-Jacques Muyembe-Tamfum, Manabu Igarashi, Eun-sil Park, Shigeru Morikawa, Sheila Makiala-Mandanda, Ayato Takada

  Journal of Virology 0022-538X 2024/07/02 

  ABSTRACT Tibroviruses are novel rhabdoviruses detected in humans, cattle, and arthropods. Four tibroviruses are known to infect humans: Bas-Congo virus (BASV), Ekpoma virus 1 (EKV-1), Ekpoma virus 2, and Mundri virus. However, since none of them has been isolated, their biological properties are largely unknown. We aimed to characterize the human tibrovirus glycoprotein (G), which likely plays a pivotal role in viral tropism and pathogenicity. Human tibrovirus Gs were found to share some primary structures and display 14 conserved cysteine residues, although their overall amino acid homology was low (29%–48%). Multiple potential glycosylation sites were found on the G molecules, and endoglycosidase H- and peptide-N-glycosidase F-sensitive glycosylation was confirmed. AlphaFold-predicted three-dimensional (3D) structures of human tibrovirus Gs were overall similar. Membrane fusion mediated by these tibrovirus Gs was induced by acidic pH. The low pH-induced conformational change that triggers fusion was reversible. Virus-like particles (VLPs) were produced by transient expression of Gs in cultured cells and used to produce mouse antisera. Using vesicular stomatitis Indiana virus pseudotyped with Gs, we found that the antisera to the respective tibrovirus VLPs showed limited cross-neutralizing activity. It was also found that human C-type lectins and T-cell immunoglobulin mucin 1 acted as attachment factors for G-mediated entry into cells. Interestingly, BASV-G showed the highest ability to utilize these molecules. The viruses infected a wide range of cell lines with preferential tropism for human-derived cells whereas the preference of EKV-1 was unique compared with the other human tibroviruses. These findings provide fundamental information to understand the biological properties of the human tibroviruses. IMPORTANCE Human tibroviruses are poorly characterized emerging rhabdoviruses associated with either asymptomatic infection or severe disease with a case fatality rate as high as 60% in humans. However, the extent and burden of human infection as well as factors behind differences in infection outcomes are largely unknown. In this study, we characterized human tibrovirus glycoproteins, which play a key role in virus-host interactions, mainly focusing on their structural and antigenic differences and cellular tropism. Our results provide critical information for understanding the biological properties of these novel viruses and for developing appropriate preparedness interventions such as diagnostic tools, vaccines, and effective therapies.
• Detection of Human Adenovirus and Rotavirus in Wastewater in Lusaka, Zambia: Demonstrating the Utility of Environmental Surveillance for the Community

  Ngonda Saasa, Ethel M’kandawire, Joseph Ndebe, Mulenga Mwenda, Fred Chimpukutu, Andrew Nalishuwa Mukubesa, Fred Njobvu, Doreen Mainza Shempela, Jay Sikalima, Carol Chiyesu, Bruce Muvwanga, Sarah M. Nampokolwe, Clement Sulwe, Thokozile Khondiwa, Todd Jennings, Ameck Kamanga, Edgar Simulundu, Conceptor Mulube, Wizaso Mwasinga, Jalaimo Mumeka, John Simwanza, Patrick Sakubita, Otridah Kapona, Chilufya Susan Aneta Mulenga, Musole Chipoya, Kunda Musonda, Nathan Kapata, Nyambe Sinyange, Muzala Kapina, Joyce Siwila, Misheck Shawa, Masahiro Kajihara, Ayato Takada, Hirofumi Sawa, Simulyamana A. Choonga, Roma Chilengi, Earnest Muyunda, King S. Nalubamba, Bernard M. Hang’ombe

  Pathogens 13 (6) 486 - 486 2024/06/07 

  Enteric infections due to viral pathogens are a major public health concern. Detecting the risk areas requires a strong surveillance system for pathogenic viruses in sources such as wastewater. Towards building an environmental surveillance system in Zambia, we aimed to identify group A rotavirus (RVA) and human adenovirus (HAdV) in wastewater. Convenient sampling was conducted at four study sites every Tuesday for five consecutive weeks. The research team focused on three different methods of viral concentration to determine the suitability in terms of cost and applicability for a regular surveillance system: the bag-mediated filtration system (BMFS), polyethylene glycol-based (PEG) precipitation, and skimmed milk (SM) flocculation. We screened 20 wastewater samples for HAdV and RVA using quantitative polymerase chain reaction (qPCR) and conventional polymerase chain reaction (cPCR). Of the 20 samples tested using qPCR, 18/20 (90%) tested positive for HAdV and 14/20 (70%) tested positive for RVA. For the genetic sequencing, qPCR positives were subjected to cPCR, of which 12 positives were successfully amplified. The human adenovirus was identified with a nucleotide identity range of 98.48% to 99.53% compared with the reference genome from GenBank. The BMFS and SM flocculation were the most consistent viral concentration methods for HAdV and RVA, respectively. A statistical analysis of the positives showed that viral positivity differed by site (p < 0.001). SM and PEG may be the most appropriate options in resource-limited settings such as Zambia due to the lower costs associated with these concentration methods. The demonstration of HAdV and RVA detection in wastewater suggests the presence of the pathogens in the communities under study and the need to establish a routine wastewater surveillance system for the identification of pathogens.
• Molecular characterization of an avian rotavirus a strain detected from a large-billed crow (Corvus macrorhynchos) in Japan.

  Yuji Fujii, Tatsunori Masatani, Shoko Nishiyama, Tatsuki Takahashi, Misuzu Okajima, Fumiki Izumi, Yoshihiro Sakoda, Ayato Takada, Makoto Ozawa, Makoto Sugiyama, Naoto Ito

  Virology 596 110114 - 110114 2024/05/18 

  Avian rotaviruses A (RVAs) are occasionally transmitted to animals other than the original hosts across species barriers. Information on RVAs carried by various bird species is important for identifying the origin of such interspecies transmission. In this study, to facilitate an understanding of the ecology of RVAs from wild birds, we characterized all of the genes of an RVA strain, JC-105, that was detected in a fecal sample of a large-billed crow (Corvus macrorhynchos) in Japan. All of the genes of this strain except for the VP4 and VP7 genes, which were classified as novel genotypes (P[56] and G40, respectively), were closely related to those of the avian-like RVA strain detected from a raccoon, indicating the possibility that crows had been involved in the transmission of avian RVAs to raccoons. Our findings highlight the need for further viral investigations in wild birds and mammals to understand the mechanisms of avian-to-mammal RVA transmission.
• Seroprevalence of Bas-Congo virus in Mangala, Democratic Republic of the Congo: a population-based cross-sectional study

  Yannick Munyeku-Bazitama, Patient Okitale-Talunda, Takanari Hattori, Takeshi Saito, Boniface Pongombo Lombe, Hiroko Miyamoto, Akina Mori-Kajihara, Masahiro Kajihara, Agathe Bikupe Nkoy, Augustin Tshibwabwa Twabela, Justin Masumu, Steve Ahuka-Mundeke, Jean-Jacques Muyembe-Tamfum, Manabu Igarashi, Eun-sil Park, Shigeru Morikawa, Sheila Makiala-Mandanda, Ayato Takada

  The Lancet Microbe 2666-5247 2024/03
• Phylogenetic Analysis of Newcastle Disease Virus Isolated from Poultry in Live Bird Markets and Wild Waterfowl in Zambia.

  Annie Kalonda, Ngonda Saasa, Masahiro Kajihara, Naganori Nao, Ladslav Moonga, Joseph Ndebe, Akina Mori-Kajihara, Andrew Nalishuwa Mukubesa, Yoshihiro Sakoda, Hirofumi Sawa, Ayato Takada, Edgar Simulundu

  Microorganisms 12 (2) 2024/02/08 

  Poultry production is essential to the economy and livelihood of many rural Zambian households. However, the industry is threatened by infectious diseases, particularly Newcastle disease virus (NDV) infection. Therefore, this study employed next-generation sequencing to characterise six NDV isolates from poultry in Zambia's live bird markets (LBMs) and wild waterfowl. Four NDV isolates were detected from 410 faecal samples collected from chickens in LBMs in Lusaka and two from 2851 wild birds from Lochinvar National Park. Phylogenetic analysis revealed that the four NDVs from LBM clustered in genotype VII and sub-genotype VII.2 were closely related to viruses previously isolated in Zambia and other Southern African countries, suggesting possible local and regional transboundary circulation of the virus. In contrast, the two isolates from wild birds belonged to class I viruses, genotype 1, and were closely related to isolates from Europe and Asia, suggesting the possible introduction of these viruses from Eurasia, likely through wild bird migration. The fusion gene cleavage site motif for all LBM-associated isolates was 112RRQKR|F117, indicating that the viruses are virulent, while the isolates from wild waterfowl had the typical 112ERQER|L117 avirulent motif. This study demonstrates the circulation of virulent NDV strains in LBMs and has, for the first time, characterised NDV from wild birds in Zambia. The study further provides the first whole genomes of NDV sub-genotype VII.2 and genotype 1 from Zambia and stresses the importance of surveillance and molecular analysis for monitoring the circulation of NDV genotypes and viral evolution.
• Development of an Immunochromatography Assay to Detect Marburg Virus and Ravn Virus

  Katendi Changula, Masahiro Kajihara, Shino Muramatsu, Koji Hiraoka, Toru Yamaguchi, Yoko Yago, Daisuke Kato, Hiroko Miyamoto, Akina Mori-Kajihara, Asako Shigeno, Reiko Yoshida, Corey W. Henderson, Andrea Marzi, Ayato Takada

  Viruses 15 (12) 2349 - 2349 2023/11/29 

  The recent outbreaks of Marburg virus disease (MVD) in Guinea, Ghana, Equatorial Guinea, and Tanzania, none of which had reported previous outbreaks, imply increasing risks of spillover of the causative viruses, Marburg virus (MARV) and Ravn virus (RAVV), from their natural host animals. These outbreaks have emphasized the need for the development of rapid diagnostic tests for this disease. Using monoclonal antibodies specific to the viral nucleoprotein, we developed an immunochromatography (IC) assay for the rapid diagnosis of MVD. The IC assay was found to be capable of detecting approximately 102−4 50% tissue culture infectious dose (TCID50)/test of MARV and RAVV in the infected culture supernatants. We further confirmed that the IC assay could detect the MARV and RAVV antigens in the serum samples from experimentally infected nonhuman primates. These results indicate that the IC assay to detect MARV can be a useful tool for the rapid point-of-care diagnosis of MVD.
• Prevalence and Genomic Characterization of Rotavirus A from Domestic Pigs in Zambia: Evidence for Possible Porcine–Human Interspecies Transmission

  Joseph Ndebe, Hayato Harima, Herman Moses Chambaro, Michihito Sasaki, Junya Yamagishi, Annie Kalonda, Misheck Shawa, Yongjin Qiu, Masahiro Kajihara, Ayato Takada, Hirofumi Sawa, Ngonda Saasa, Edgar Simulundu

  Pathogens 12 (10) 1199 - 1199 2023/09/27 [Refereed]
   

  Rotavirus is a major cause of diarrhea globally in animals and young children under 5 years old. Here, molecular detection and genetic characterization of porcine rotavirus in smallholder and commercial pig farms in the Lusaka Province of Zambia were conducted. Screening of 148 stool samples by RT-PCR targeting the VP6 gene revealed a prevalence of 22.9% (34/148). Further testing of VP6-positive samples with VP7-specific primers produced 12 positives, which were then Sanger-sequenced. BLASTn of the VP7 positives showed sequence similarity to porcine and human rotavirus strains with identities ranging from 87.5% to 97.1%. By next-generation sequencing, the full-length genetic constellation of the representative strains RVA/pig-wt/ZMB/LSK0137 and RVA/pig-wt/ZMB/LSK0147 were determined. Genotyping of these strains revealed a known Wa-like genetic backbone, and their genetic constellations were G4-P[6]-I5-R1-C1-M1-A8-N1-T1-E1-H1 and G9-P[13]-I5-R1-C1-M1-A8-N1-T1-E1-H1, respectively. Phylogenetic analysis revealed that these two viruses might have their ancestral origin from pigs, though some of their gene segments were related to human strains. The study shows evidence of reassortment and possible interspecies transmission between pigs and humans in Zambia. Therefore, the “One Health” surveillance approach for rotavirus A in animals and humans is recommended to inform the design of effective control measures.
• ザンビアにおけるブタインフルエンザの疫学調査

  播磨 勇人, 奥谷 公亮, 梶原 将大, 小川 寛人, Simulundu Edgar, Bwalya Eugene, 邱 永晋, 迫田 義博, 西藤 岳彦, Hang' ombe Bernard, 澤 洋文, Mweene Aaron, 高田 礼人

  日本獣医学会学術集会講演要旨集 (公社)日本獣医学会 166回 154 - 154 1347-8621 2023/09
• Surveillance, Isolation, and Genetic Characterization of Bat Herpesviruses in Zambia.

  Hayato Harima, Yongjin Qiu, Junya Yamagishi, Masahiro Kajihara, Katendi Changula, Kosuke Okuya, Mao Isono, Tomoyuki Yamaguchi, Hirohito Ogawa, Naganori Nao, Michihito Sasaki, Edgar Simulundu, Aaron S Mweene, Hirofumi Sawa, Kanako Ishihara, Bernard M Hang'ombe, Ayato Takada

  Viruses 15 (6) 2023/06/13 

  Bats are of significant interest as reservoirs for various zoonotic viruses with high diversity. During the past two decades, many herpesviruses have been identified in various bats worldwide by genetic approaches, whereas there have been few reports on the isolation of infectious herpesviruses. Herein, we report the prevalence of herpesvirus infection of bats captured in Zambia and genetic characterization of novel gammaherpesviruses isolated from striped leaf-nosed bats (Macronycteris vittatus). By our PCR screening, herpesvirus DNA polymerase (DPOL) genes were detected in 29.2% (7/24) of Egyptian fruit bats (Rousettus aegyptiacus), 78.1% (82/105) of Macronycteris vittatus, and one Sundevall's roundleaf bat (Hipposideros caffer) in Zambia. Phylogenetic analyses of the detected partial DPOL genes revealed that the Zambian bat herpesviruses were divided into seven betaherpesvirus groups and five gammaherpesvirus groups. Two infectious strains of a novel gammaherpesvirus, tentatively named Macronycteris gammaherpesvirus 1 (MaGHV1), were successfully isolated from Macronycteris vittatus bats, and their complete genomes were sequenced. The genome of MaGHV1 encoded 79 open reading frames, and phylogenic analyses of the DNA polymerase and glycoprotein B demonstrated that MaGHV1 formed an independent lineage sharing a common origin with other bat-derived gammaherpesviruses. Our findings provide new information regarding the genetic diversity of herpesviruses maintained in African bats.
• Head-to-head comparison of diagnostic accuracy of four Ebola virus disease rapid diagnostic tests versus GeneXpert® in eastern Democratic Republic of the Congo outbreaks: a prospective observational study

  Daniel Mukadi-Bamuleka, Junior Bulabula-Penge, Bart K.M. Jacobs, Anja De Weggheleire, François Edidi-Atani, Fabrice Mambu-Mbika, Anaïs Legand, John D. Klena, Peter N. Fonjungo, Placide Mbala-Kingebeni, Sheila Makiala-Mandanda, Masahiro Kajihara, Ayato Takada, Joel M. Montgomery, Pierre Formenty, Jean-Jacques Muyembe-Tamfum, Kevin K. Ariën, Johan van Griensven, Steve Ahuka-Mundeke, Hgo Kavunga-Membo, Elie Ishara-Nshombo, Stijn Roge, Noella Mulopo-Mukanya, Espérance Tsiwedi-Tsilabia, Emile Muhindo-Milonde, Marie-Anne Kavira-Muhindo, Maria E. Morales-Betoulle, Antoine Nkuba-Ndaye

  eBioMedicine 91 104568 - 104568 2352-3964 2023/05
• Structural and energetic basis for differential binding of Ebola and Marburg virus glycoproteins to a bat-derived Niemann-Pick C1 protein.

  Manabu Igarashi, Takatsugu Hirokawa, Ayato Takada

  The Journal of infectious diseases 2023/04/29 

  Our previous study demonstrated that the fruit bat (Yaeyama flying fox)-derived cell line FBKT1 showed preferential susceptibility to Ebola virus (EBOV), whereas the human cell line HEK293T was similarly susceptible to EBOV and Marburg virus (MARV). This was due to three amino acid differences of the endosomal receptor Niemann-Pick C1 (NPC1) between this bat species and humans (i.e., TET and SGA, respectively, at positions 425-427 of NPC1), as well as two specific amino acid differences at positions 87 and 142 of the viral surface glycoprotein (GP) between EBOV and MARV. To understand the contribution of these amino acid differences to intermolecular interactions between NPC1 and GP, we performed molecular dynamics simulations and binding free energy calculations. The average binding free energies of human NPC1 (hNPC1) and its mutant having TET at positions 425-427 (hNPC1/TET) were similar for the interaction with EBOV GP. In contrast, hNPC1/TET had a weaker interaction with MARV GP than wildtype hNPC1. As expected, substitutions of amino acid residues at 87 or 142 in EBOV and MARV GPs converted the binding affinity to hNPC1/TET. Our data provide structural and energetic insights for understanding potential differences in the GP-NPC1 interaction, which could influence the host tropism of EBOV and MARV.
• Morphogenesis of Bullet-Shaped Rabies Virus Particles Regulated by TSG101.

  Yukari Itakura, Koshiro Tabata, Takeshi Saito, Kittiya Intaruck, Nijiho Kawaguchi, Mai Kishimoto, Shiho Torii, Shintaro Kobayashi, Naoto Ito, Michiko Harada, Satoshi Inoue, Ken Maeda, Ayato Takada, William W Hall, Yasuko Orba, Hirofumi Sawa, Michihito Sasaki

  Journal of virology 97 (5) e0043823  2023/04/12 

  Viral protein assembly and virion budding are tightly regulated to enable the proper formation of progeny virions. At this late stage in the virus life cycle, some enveloped viruses take advantage of the host endosomal sorting complex required for transport (ESCRT) machinery, which contributes to the physiological functions of membrane modulation and abscission. Bullet-shaped viral particles are unique morphological characteristics of rhabdoviruses; however, the involvement of host factors in rhabdovirus infection and, specifically, the molecular mechanisms underlying virion formation are not fully understood. In the present study, we used a small interfering RNA (siRNA) screening approach and found that the ESCRT-I component TSG101 contributes to the propagation of rabies virus (RABV). We demonstrated that the matrix protein (M) of RABV interacts with TSG101 via the late domain containing the PY and YL motifs, which are conserved in various viral proteins. Loss of the YL motif in the RABV M or the downregulation of host TSG101 expression resulted in the intracellular aggregation of viral proteins and abnormal virus particle formation, indicating a defect in the RABV assembly and budding processes. These results indicate that the interaction of the RABV M and TSG101 is pivotal for not only the efficient budding of progeny RABV from infected cells but also for the bullet-shaped virion morphology. IMPORTANCE Enveloped viruses bud from cells with the host lipid bilayer. Generally, the membrane modulation and abscission are mediated by host ESCRT complexes. Some enveloped viruses utilize their late (L-) domain to interact with ESCRTs, which promotes viral budding. Rhabdoviruses form characteristic bullet-shaped enveloped virions, but the underlying molecular mechanisms involved remain elusive. Here, we showed that TSG101, one of the ESCRT components, supports rabies virus (RABV) budding and proliferation. TSG101 interacted with RABV matrix protein via the L-domain, and the absence of this interaction resulted in intracellular virion accumulation and distortion of the morphology of progeny virions. Our study reveals that virion formation of RABV is highly regulated by TSG101 and the virus matrix protein.
• Surveillance and Phylogenetic Characterisation of Avian Influenza Viruses Isolated from Wild Waterfowl in Zambia in 2015, 2020, and 2021

  Annie Kalonda, Ngonda Saasa, Masahiro Kajihara, Naganori Nao, Ladislav Moonga, Joseph Ndebe, Akina Mori-Kajihara, Andrew Nalishuwa Mukubesa, Mulemba Samutela, Samuel Munjita, Yoshihiro Sakoda, Hirofumi Sawa, Ayato Takada, Edgar Simulundu

  Transboundary and Emerging Diseases 2023 1 - 16 1865-1674 2023/03/01 

  In recent years, the southern African region has experienced repeated incursions of highly pathogenic avian influenza viruses (HPAIVs), with wild migratory birds being implicated in the spread. To understand the profile of avian influenza viruses (AIVs) circulating in Zambia, we surveyed wild waterfowl for AIVs and phylogenetically characterised the isolates detected in 2015, 2020, and 2021. A total of 2,851 faecal samples of wild waterfowl were collected from Lochinvar National Park in the Southern Province of Zambia. During the study period, 85 (3.0%) low pathogenicity AIVs belonging to various subtypes were isolated, with H2N9, H8N4, and H10N8 being reported for the first time in avian species in Africa. The majority of the isolates were detected from glossy ibis (order Pelecaniformes) making it the first report of AIV from these birds in Zambia. Phylogenetic analysis of all eight gene segments of the 30 full genomes obtained in this study revealed that all the isolates belonged to the Eurasian lineage with their closest relatives being viruses isolated from wild and/or domestic birds in Bangladesh, Belgium, Egypt, Georgia, Mongolia, the Netherlands, and South Africa. Additionally, the Zambian viruses were grouped into distinct clusters based on the year of isolation. While no notifiable AIVs of the H5 or H7 subtypes were detected in wild birds in Zambia, viral internal protein genes of some viruses were closely related to H7 low pathogenicity AIVs. This study shows that periodically, a considerable diversity of AIV subtypes are introduced into the Zambian ecosystem by wild migratory waterfowl. The findings highlight the importance of continuous surveillance and monitoring of AIVs in wild waterfowl, including birds traditionally not considered to be major AIV reservoirs, for a better understanding of the eco-epidemiology and evolutionary dynamics of AIVs in Africa.
• Detection of Old and New World Relapsing Fever Borreliae in Ornithodoros Ticks Collected from Warthog Burrows in Zambia.

  Yongjin Qiu, Herman M Chambaro, Kozue Sato, David Squarre, Edgar Simulundu, Masahiro Kajihara, Katendi Changula, Manyando Simbotwe, Hayato Harima, Joseph Ndebe, Ladslav Moonga, Ryo Nakao, Ayato Takada, Bernard Mudenda Hang'ombe, Hirofumi Sawa, Hiroki Kawabata

  Microorganisms 11 (1) 2023/01/12 

  Relapsing fever (RF) is an arthropod-borne disease caused by Borrelia spirochete, which is one of the major public health concerns in endemic regions including Africa. However, information on Borrelia&nbsp;spirochetes is limited in Zambia. Here, we investigate the Borrelia&nbsp;spirochetes harbored by Ornithodoros ticks in Zambian National Parks. We analyzed 182 DNA samples pooled from 886 Ornithodoros ticks. Of these, 43 tested positive, and their sequence revealed that the ticks harbored both Old and New World RF borreliae. This research presents the first evidence of Old-World RF borreliae in Zambia. The New World RF borreliae detected herein differed from the Candidatus Borrelia fainii previously reported in Zambia and were closely related to the pathogenic Borrelia sp. VS4 identified in Tanzania. Additionally, Borrelia theileri was recently reported in Zambia. Hence, at least four different Borrelia species occur in Zambia, and the organisms causing relapsing fever there might be more complex than previously thought. We empirically confirmed that real-time PCR with TaqMan minor groove binder probes accurately and simultaneously detected both Old and New World RF. In this manner, they could facilitate quantitative analyses of both types of RF borreliae. Subsequent investigations should endeavor to isolate the aforementioned Borrelia spp. and perform serosurveys on patients with RF.
• Current status and molecular epidemiology of rabies virus from different hosts and regions in Malawi.

  Henson Kainga, Elisha Chatanga, Marvin Collen Phonera, John Pilate Kothowa, Precious Dzimbiri, Gladson Kamwendo, Malala Mulavu, Cynthia Sipho Khumalo, Katendi Changula, Herman Chambaro, Hayato Harima, Masahiro Kajihara, Kholiwe Mkandawire, Patrick Chikungwa, Julius Chulu, Gilson Njunga, Simbarashe Chitanga, Benjamin Mubemba, Michihito Sasaki, Yasuko Orba, Yongjin Qiu, Junya Yamagishi, Edgar Simulundu, Ayato Takada, Boniface Namangala, Hirofumi Sawa, Walter Muleya

  Archives of virology 168 (2) 61 - 61 2023/01/12 

  Although rabies is endemic in Malawi, there have been no studies in which rabies virus was systematically investigated and characterized in multiple animal hosts in that country. In order to provide molecular epidemiological data on rabies virus in Malawi, 683 suspected rabies case reports from 2008 to 2021 were examined, and 46 (dog = 40, cow = 5, and cat = 1) viable rabies-positive brain samples archived at the Central Veterinary Laboratory (CVL), Lilongwe, Malawi, were analyzed genetically. The results showed an increase in the submission of brain samples from 2008 to 2010, with the highest number of submissions observed in 2020. Of the 683 case reports analyzed for the period under review, 38.1% (260/683) (CI: 34.44 - 41.84) were confirmed by direct fluorescent antibody test. Among the confirmed cases, 65.4% (170/260) (CI: 59.23 - 71.09) were canine rabies. Further, phylogenetic analysis revealed that sequences from different animal hosts clustered together within the Africa 1b lineage, suggesting that the strains circulating in livestock are similar to those in domestic dogs. This finding supports the hypothesis that canine rabies is spilling over to livestock and emphasizes the need for further studies to provide data for effective control of rabies in Malawi.
• Isolation and Characterization of Distinct Rotavirus A in Bat and Rodent Hosts.

  Mai Kishimoto, Masahiro Kajihara, Koshiro Tabata, Yukari Itakura, Shinsuke Toba, Seiya Ozono, Yuko Sato, Tadaki Suzuki, Naoto Ito, Katendi Changula, Yongjin Qiu, Akina Mori-Kajihara, Yoshiki Eto, Hayato Harima, Daniel Mwizabi, Bernard M Hang'ombe, William W Hall, Ayato Takada, Yasuko Orba, Hirofumi Sawa, Michihito Sasaki

  Journal of virology 97 (1) e0145522  2023/01/12 

  Rotavirus A (RVA) causes diarrheal disease in humans and various animals. Recent studies have identified bat and rodent RVAs with evidence of zoonotic transmission and genome reassortment. However, the virological properties of bat and rodent RVAs with currently identified genotypes still need to be better clarified. Here, we performed virus isolation-based screening for RVA in animal specimens and isolated RVAs (representative strains: 16-06 and MpR12) from Egyptian fruit bat and Natal multimammate mouse collected in Zambia. Whole-genome sequencing and phylogenetic analysis revealed that the genotypes of bat RVA 16-06 were identical to that of RVA BATp39 strain from the Kenyan fruit bat, which has not yet been characterized. Moreover, all segments of rodent RVA MpR12 were highly divergent and assigned to novel genotypes, but RVA MpR12 was phylogenetically closer to bat RVAs than to other rodent RVAs, indicating a unique evolutionary history. We further investigated the virological properties of the isolated RVAs. In brief, we found that 16-06 entered cells by binding to sialic acids on the cell surface, while MpR12 entered in a sialic acid-independent manner. Experimental inoculation of suckling mice with 16-06 and MpR12 revealed that these RVAs are causative agents of diarrhea. Moreover, 16-06 and MpR12 demonstrated an ability to infect and replicate in a 3D-reconstructed primary human intestinal epithelium with comparable efficiency to the human RVA. Taken together, our results detail the unique genetic and virological features of bat and rodent RVAs and demonstrate the need for further investigation of their zoonotic potential. IMPORTANCE Recent advances in nucleotide sequence detection methods have enabled the detection of RVA genomes from various animals. These studies have discovered multiple divergent RVAs and have resulted in proposals for the genetic classification of novel genotypes. However, most of these RVAs have been identified via dsRNA viral genomes and not from infectious viruses, and their virological properties, such as cell/host tropisms, transmissibility, and pathogenicity, are unclear and remain to be clarified. Here, we successfully isolated RVAs with novel genome constellations from three bats and one rodent in Zambia. In addition to whole-genome sequencing, the isolated RVAs were characterized by glycan-binding affinity, pathogenicity in mice, and infectivity to the human gut using a 3D culture of primary intestinal epithelium. Our study reveals the first virological properties of bat and rodent RVAs with high genetic diversity and unique evolutional history and provides basic knowledge to begin estimating the potential of zoonotic transmission.
• Susceptibility of common dabbling and diving duck species to clade 2.3.2.1 H5N1 high pathogenicity avian influenza virus: an experimental infection study

  Kosuke SODA, Yukiko TOMIOKA, Tatsufumi USUI, Hiroichi OZAKI, Hiroshi ITO, Yasuko NAGAI, Naoki YAMAMOTO, Masatoshi OKAMATSU, Norikazu ISODA, Masahiro KAJIHARA, Yoshihiro SAKODA, Ayato TAKADA, Toshihiro ITO

  Journal of Veterinary Medical Science 85 (9) 942 - 949 0916-7250 2023
• Field-deployable multiplex detection method of SARS-CoV-2 and influenza virus using loop-mediated isothermal amplification and DNA chromatography.

  Kyoko Hayashida, Alejandro Garcia, Lavel Chinyama Moonga, Tatsuki Sugi, Kodera Takuya, Mitsuo Kawase, Fumihiro Kodama, Atsushi Nagasaka, Nobuhisa Ishiguro, Ayato Takada, Masahiro Kajihara, Naganori Nao, Masashi Shingai, Hiroshi Kida, Yasuhiko Suzuki, William W Hall, Hirofumi Sawa, Junya Yamagishi

  PloS one 18 (5) e0285861  2023 

  A novel multiplex loop-mediated isothermal amplification (LAMP) method combined with DNA chromatography was developed for the simultaneous detection of three important respiratory disease-causing viruses: severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza A virus, and influenza B virus. Amplification was performed at a constant temperature, and a positive result was confirmed by a visible colored band. An in-house drying protocol with trehalose was used to prepare the dried format multiplex LAMP test. Using this dried multiplex LAMP test, the analytical sensitivity was determined to be 100 copies for each viral target and 100-1000 copies for the simultaneous detection of mixed targets. The multiplex LAMP system was validated using clinical COVID-19 specimens and compared with the real-time qRT-PCR method as a reference test. The determined sensitivity of the multiplex LAMP system for SARS-CoV-2 was 71% (95% CI: 0.62-0.79) for cycle threshold (Ct) ≤ 35 samples and 61% (95% CI: 0.53-0.69) for Ct ≤40 samples. The specificity was 99% (95%CI: 0.92-1.00) for Ct ≤35 samples and 100% (95%CI: 0.92-1.00) for the Ct ≤40 samples. The developed simple, rapid, low-cost, and laboratory-free multiplex LAMP system for the two major important respiratory viral diseases, COVID-19 and influenza, is a promising field-deployable diagnosis tool for the possible future 'twindemic, ' especially in resource-limited settings.
• 2022 taxonomic update of phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales

  Jens H. Kuhn, Scott Adkins, Sergey V. Alkhovsky, Tatjana Avšič-Županc, María A. Ayllón, Justin Bahl, Anne Balkema-Buschmann, Matthew J. Ballinger, Martina Bandte, Martin Beer, Nicolas Bejerman, Éric Bergeron, Nadine Biedenkopf, Laurent Bigarré, Carol D. Blair, Kim R. Blasdell, Steven B. Bradfute, Thomas Briese, Paul A. Brown, Rémy Bruggmann, Ursula J. Buchholz, Michael J. Buchmeier, Alexander Bukreyev, Felicity Burt, Carmen Büttner, Charles H. Calisher, Thierry Candresse, Jeremy Carson, Inmaculada Casas, Kartik Chandran, Rémi N. Charrel, Yuya Chiaki, Anya Crane, Mark Crane, Laurent Dacheux, Elena Dal Bó, Juan Carlos de la Torre, Xavier de Lamballerie, William M. de Souza, Rik L. de Swart, Nolwenn M. Dheilly, Nicholas Di Paola, Francesco Di Serio, Ralf G. Dietzgen, Michele Digiaro, J. Felix Drexler, W. Paul Duprex, Ralf Dürrwald, Andrew J. Easton, Toufic Elbeaino, Koray Ergünay, Guozhong Feng, Claudette Feuvrier, Andrew E. Firth, Anthony R. Fooks, Pierre B. H. Formenty, Juliana Freitas-Astúa, Selma Gago-Zachert, María Laura García, Adolfo García-Sastre, Aura R. Garrison, Scott E. Godwin, Jean-Paul J. Gonzalez, Joëlle Goüy de Bellocq, Anthony Griffiths, Martin H. Groschup, Stephan Günther, John Hammond, Jussi Hepojoki, Melanie M. Hierweger, Seiji Hongō, Masayuki Horie, Hidenori Horikawa, Holly R. Hughes, Adam J. Hume, Timothy H. Hyndman, Dàohóng Jiāng, Gilda B. Jonson, Sandra Junglen, Fujio Kadono, David G. Karlin, Boris Klempa, Jonas Klingström, Michel C. Koch, Hideki Kondō, Eugene V. Koonin, Jarmila Krásová, Mart Krupovic, Kenji Kubota, Ivan V. Kuzmin, Lies Laenen, Amy J. Lambert, Jiànróng Lǐ, Jun-Min Li, François Lieffrig, Igor S. Lukashevich, Dongsheng Luo, Piet Maes, Marco Marklewitz, Sergio H. Marshall, Shin-Yi L. Marzano, John W. McCauley, Ali Mirazimi, Peter G. Mohr, Nick J. G. Moody, Yasuaki Morita, Richard N. Morrison, Elke Mühlberger, Rayapati Naidu, Tomohide Natsuaki, José A. Navarro, Yutaro Neriya, Sergey V. Netesov, Gabriele Neumann, Norbert Nowotny, Francisco M. Ochoa-Corona, Gustavo Palacios, Laurane Pallandre, Vicente Pallás, Anna Papa, Sofia Paraskevopoulou, Colin R. Parrish, Alex Pauvolid-Corrêa, Janusz T. Pawęska, Daniel R. Pérez, Florian Pfaff, Richard K. Plemper, Thomas S. Postler, Françoise Pozet, Sheli R. Radoshitzky, Pedro L. Ramos-González, Marius Rehanek, Renato O. Resende, Carina A. Reyes, Víctor Romanowski, Dennis Rubbenstroth, Luisa Rubino, Artemis Rumbou, Jonathan A. Runstadler, Melanie Rupp, Sead Sabanadzovic, Takahide Sasaya, Heike Schmidt-Posthaus, Martin Schwemmle, Torsten Seuberlich, Stephen R. Sharpe, Mang Shi, Manuela Sironi, Sophie Smither, Jin-Won Song, Kirsten M. Spann, Jessica R. Spengler, Mark D. Stenglein, Ayato Takada, Robert B. Tesh, Jana Těšíková, Natalie J. Thornburg, Nicole D. Tischler, Yasuhiro Tomitaka, Keizō Tomonaga, Noël Tordo, Kenta Tsunekawa, Massimo Turina, Ioannis E. Tzanetakis, Anna Maria Vaira, Bernadette van den Hoogen, Bert Vanmechelen, Nikos Vasilakis, Martin Verbeek, Susanne von Bargen, Jiro Wada, Victoria Wahl, Peter J. Walker, Anna E. Whitfield, John V. Williams, Yuri I. Wolf, Junki Yamasaki, Hironobu Yanagisawa, Gongyin Ye, Yong-Zhen Zhang, Arnfinn Lodden Økland

  Archives of Virology 167 (12) 2857 - 2906 0304-8608 2022/11/28 [Refereed]
   

  In March 2022, following the annual International Committee on Taxonomy of Viruses (ICTV) ratification vote on newly proposed taxa, the phylum Negarnaviricota was amended and emended. The phylum was expanded by two new families (bunyaviral Discoviridae and Tulasviridae), 41 new genera, and 98 new species. Three hundred forty-nine species were renamed and/or moved. The accidentally misspelled names of seven species were corrected. This article presents the updated taxonomy of Negarnaviricota as now accepted by the ICTV.
• Luna Virus and Helminths in Wild Mastomys natalensis in Two Contrasting Habitats in Zambia: Risk Factors and Evidence of Virus Dissemination in Semen.

  Samuel Munalula Munjita, Given Moonga, Andrew Nalishuwa Mukubesa, Joseph Ndebe, Benjamin Mubemba, Manu Vanaerschot, Cristina Tato, John Tembo, Nathan Kapata, Simbarashe Chitanga, Katendi Changula, Mashiro Kajihara, Walter Muleya, Ayato Takada, Elisabeth Fichet-Calvet, Alimuddin Zumla, Hirofumi Sawa, Matthew Bates, Sody Munsaka, Edgar Simulundu

  Pathogens (Basel, Switzerland) 11 (11) 2022/11/14 

  Transmission dynamics and the maintenance of mammarenaviruses in nature are poorly understood. Using metagenomic next-generation sequencing (mNGS) and RT-PCR, we investigated the presence of mammarenaviruses and co-infecting helminths in various tissues of 182 Mastomys natalensis rodents and 68 other small mammals in riverine and non-riverine habitats in Zambia. The Luna virus (LUAV) genome was the only mammarenavirus detected (7.7%; 14/182) from M. natalensis. Only one rodent from the non-riverine habitat was positive, while all six foetuses from one pregnant rodent carried LUAV. LUAV-specific mNGS reads were 24-fold higher in semen than in other tissues from males. Phylogenetically, the viruses were closely related to each other within the LUAV clade. Helminth infections were found in 11.5% (21/182) of M. natalensis. LUAV-helminth co-infections were observed in 50% (7/14) of virus-positive rodents. Juvenility (OR = 9.4; p = 0.018; 95% CI: 1.47-59.84), nematodes (OR = 15.5; p = 0.001; 95% CI: 3.11-76.70), cestodes (OR = 10.8; p = 0.025; 95% CI: 1.35-86.77), and being male (OR = 4.6; p = 0.036; 95% CI: 1.10-18.90) were associated with increased odds of LUAV RNA detection. The role of possible sexual and/or congenital transmission in the epidemiology of LUAV infections in rodents requires further study, along with the implications of possible helminth co-infection.
• Seroprevalence and Associated Risk Factors of Rift Valley Fever in Livestock from Three Ecological Zones of Malawi.

  Henson Kainga, Marvin Collen Phonera, Elisha Chatanga, Simegnew Adugna Kallu, Prudence Mpundu, Mulemba Samutela, Herman Moses Chambaro, Masahiro Kajihara, Doreen Mainza Shempela, Jay Sikalima, Walter Muleya, Misheck Shawa, Julius Chulu, Gilson Njunga, Martin Simuunza, Ayato Takada, Hirofumi Sawa, Edgar Simulundu, Ngonda Saasa

  Pathogens (Basel, Switzerland) 11 (11) 2022/11/14 

  The epidemiology of Rift Valley fever (RVF) is poorly understood in Malawi. Here, a cross-sectional study was conducted (March-June 2020) to investigate the seroprevalence and potential risk factors of RVF virus (RVFV) in cattle, goats, and sheep in three ecological zones of Malawi. A total of 1523 serum samples were tested for anti-RVFV IgG and IgM antibodies by ELISA. Additionally, a questionnaire survey was used to assess potential RVF risk factors. The overall seroprevalence was 17.14% (261/1523; 95% CI = 15.33-19.11) for individual livestock and 33.24% (120/361; 95% CI = 28.18-38.11) for the livestock herd. Seroprevalence was significantly high in sheep (25.68%, 95% CI = 19.31-33.26) compared with cattle (21.35%, 95% CI = 18.74-24.22) and goats (7.72%, 95% CI = 5.72-10.34), (p = 0.047). At the individual livestock level, the risk was elevated in female livestock (OR: 1.74, 95% CI = 1.08-12.82) (p = 0.016), while at the herd level, areas receiving approximately 1001-1500 mm of rainfall (OR: 2.47, 95% CI = 1.14-5.37) (p = 0.022), areas of rainfall amount greater than approximately 1600 mm (OR: 2.239, 95% CI = 1.07-8.82) (p = 0.023), and mixed species herds (OR: 10.410, 95% CI = 3.04-35.59) (p = 0.001), were significant risk factors. The detection of IgM antibodies confirmed active circulation of RVFV in Malawi. Therefore, monitoring of RVF in animals, humans, and vectors using a "One Health" approach, along with community sensitization among the high-risk populations, could help mitigate the threat posed by this zoonotic disease in Malawi.
• Experimental and natural infections of white-tailed sea eagles (Haliaeetus albicilla) with high pathogenicity avian influenza virus of H5 subtype

  Yoshikazu Fujimoto, Kohei Ogasawara, Norikazu Isoda, Hitoshi Hatai, Kosuke Okuya, Yukiko Watanabe, Ayato Takada, Yoshihiro Sakoda, Keisuke Saito, Makoto Ozawa

  Frontiers in Microbiology 13 2022/10/03 [Refereed][Not invited]
   

  White-tailed sea eagle (Haliaeetus albicilla), a regionally rare species of raptor, is threatened in several countries. To assess the risk of H5 high pathogenicity avian influenza (HPAI) viral infection in rare bird species, we performed experimental infections with a GS/GD96-lineage H5N6 HPAI virus of clade 2.3.4.4e in white-tailed sea eagles. Additionally, during the winter of 2020–2021 in Japan, we accidentally encountered a white-tailed sea eagle that had a fatal outcome due to natural infection with a GS/GD96-lineage H5N8 HPAI virus of clade 2.3.4.4b, allowing us to compare experimental and natural infections in the same rare raptor species. Our experiments demonstrated the susceptibility of white-tailed sea eagles to the GS/GD96-lineage H5 HPAI virus with efficient replication in systemic organs. The potential for the viruses to spread within the white-tailed sea eagle population through indirect transmission was also confirmed. Comprehensive comparisons of both viral distribution and histopathological observations between experimentally and naturally infected white-tailed sea eagles imply that viral replication in the brain is responsible for the disease severity and mortality in this species. These findings provide novel insights into the risk assessment of H5 HPAI viral infection in white-tailed sea eagles, proper diagnostic procedures, potential risks to artificially fed eagle populations and persons handling superficially healthy eagles, potential impact of intragastric infection on eagle outcomes, and possibility of severity of the disease being attributed to viral replication in the brain.
• Molecular characterisation of a novel avian rotavirus A strain detected from a gull species (Larus sp.).

  Yuji Fujii, Tatsunori Masatani, Shoko Nishiyama, Misuzu Okajima, Fumiki Izumi, Katsunori Okazaki, Yoshihiro Sakoda, Ayato Takada, Makoto Ozawa, Makoto Sugiyama, Naoto Ito

  The Journal of general virology 103 (10) 2022/10 

  A recent study demonstrated the possibility that migratory birds are responsible for the global spread of avian rotavirus A (RVA). However, little is known about what types of RVAs are retained in migratory birds. In this study, to obtain information on RVA strains in migratory birds, we characterised an RVA strain, Ho374, that was detected in a faecal sample from a gull species (Larus sp.). Genetic analysis revealed that all 11 genes of this strain were classified as new genotypes (G28-P[39]-I21-R14-C14-M13-A24-N14-T16-E21-H16). This clearly indicates that the genetic diversity of avian RVAs is greater than previously recognised. Our findings highlight the need for investigations of RVA strains retained in migratory birds, including gulls.
• Single Nucleotide Variants of the Human TIM-1 IgV Domain with Reduced Ability to Promote Viral Entry into Cells

  Takanari Hattori, Takeshi Saito, Hiroko Miyamoto, Masahiro Kajihara, Manabu Igarashi, Ayato Takada

  Viruses 14 (10) 2124 - 2124 2022/09/26 

  Human T-cell immunoglobulin mucin 1 (hTIM-1) is known to promote cellular entry of enveloped viruses. Previous studies suggested that the polymorphisms of hTIM-1 affected its function. Here, we analyzed single nucleotide variants (SNVs) of hTIM-1 to determine their ability to promote cellular entry of viruses using pseudotyped vesicular stomatitis Indiana virus (VSIV). We obtained hTIM-1 sequences from a public database (Ensembl genome browser) and identified 35 missense SNVs in 3 loops of the hTIM-1 immunoglobulin variable (IgV) domain, which had been reported to interact with the Ebola virus glycoprotein (GP) and phosphatidylserine (PS) in the viral envelope. HEK293T cells transiently expressing wildtype hTIM-1 or its SNV mutants were infected with VSIVs pseudotyped with filovirus or arenavirus GPs, and their infectivities were compared. Eleven of the thirty-five SNV substitutions reduced the efficiency of hTIM-1-mediated entry of pseudotyped VSIVs. These SNV substitutions were found not only around the PS-binding pocket but also in other regions of the molecule. Taken together, our findings suggest that some SNVs of the hTIM-1 IgV domain have impaired ability to interact with PS and/or viral GPs in the viral envelope, which may affect the hTIM-1 function to promote viral entry into cells.
• Detection of anti-ebolavirus antibodies in Ghanaian pigs.

  Hirohito Ogawa, Kenji Ohya, Raphael Ayizanga, Hiroko Miyamoto, Asako Shigeno, Masao Yamada, Yasuhiro Takashima, Miho Inoue-Murayama, Ayato Takada, Boniface Baboreka Kayang

  The Journal of veterinary medical science 2022/09/20 [Refereed]
   

  Some filoviruses such as ebolaviruses and marburgviruses, cause hemorrhagic fever in humans and nonhuman primates. Pigs are suggested to play a potential role in the filovirus ecology. We investigated the seroprevalence of filovirus infection in pigs in Ghana. Using a viral glycoprotein (GP)-based enzyme-linked immunosorbent assay, we detected filovirus-specific immunoglobulin G antibodies in 5 of 139 samples. These positive sera showed specificities to four different filovirus species. Particularly, two of the positive sera reacted to GPs of two African ebolaviruses (i.e., Ebola virus and Taï Forest virus) in Western blotting. Our results suggest that these Ghanaian pigs were exposed to multiple filoviruses and emphasize the importance of continuous monitoring of filovirus infection in pig populations in West African countries.
• ザンビアのコウモリにおけるネルソンベイオルソレオウイルスの保有調査および分離ウイルスの病原性比較解析

  播磨 勇人, 佐々木 道仁, 大場 靖子, 邱 永晋, Changula Katendi, 梶原 将大, Simulundu Edgar, 谷口 怜, 高田 礼人, 西條 政幸, Hang'ombe Bernard M., 澤 洋文

  日本獣医学会学術集会講演要旨集 (公社)日本獣医学会 165回 [F1P - 09] 1347-8621 2022/09
• Genomic Surveillance of SARS-CoV-2 in the Southern Province of Zambia: Detection and Characterization of Alpha, Beta, Delta, and Omicron Variants of Concern.

  Ben Katowa, Annie Kalonda, Benjamin Mubemba, Japhet Matoba, Doreen Mainza Shempela, Jay Sikalima, Boniface Kabungo, Katendi Changula, Simbarashe Chitanga, Mpanga Kasonde, Otridah Kapona, Nathan Kapata, Kunda Musonda, Mwaka Monze, John Tembo, Matthew Bates, Alimuddin Zumla, Catherine G Sutcliffe, Masahiro Kajihara, Junya Yamagishi, Ayato Takada, Hirofumi Sawa, Roma Chilengi, Victor Mukonka, Walter Muleya, Edgar Simulundu

  Viruses 14 (9) 2022/08/24 

  Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) have significantly impacted the global epidemiology of the pandemic. From December 2020 to April 2022, we conducted genomic surveillance of SARS-CoV-2 in the Southern Province of Zambia, a region that shares international borders with Botswana, Namibia, and Zimbabwe and is a major tourist destination. Genetic analysis of 40 SARS-CoV-2 whole genomes revealed the circulation of Alpha (B.1.1.7), Beta (B.1.351), Delta (AY.116), and multiple Omicron subvariants with the BA.1 subvariant being predominant. Whereas Beta, Delta, and Omicron variants were associated with the second, third, and fourth pandemic waves, respectively, the Alpha variant was not associated with any wave in the country. Phylogenetic analysis showed evidence of local transmission and possible multiple introductions of SARS-CoV-2 VOCs in Zambia from different European and African countries. Across the 40 genomes analysed, a total of 292 mutations were observed, including 182 missense mutations, 66 synonymous mutations, 23 deletions, 9 insertions, 1 stop codon, and 11 mutations in the non-coding region. This study stresses the need for the continued monitoring of SARS-CoV-2 circulation in Zambia, particularly in strategically positioned regions such as the Southern Province which could be at increased risk of introduction of novel VOCs.
• Human ACE2 Genetic Polymorphism Affecting SARS-CoV and SARS-CoV-2 Entry into Cells

  Takanari Hattori, Takeshi Saito, Kosuke Okuya, Yuji Takahashi, Hiroko Miyamoto, Masahiro Kajihara, Manabu Igarashi, Ayato Takada

  Microbiology Spectrum 10 (4) e0087022  2022/07/11 [Refereed]
   

  SARS-CoV and SARS-CoV-2 are known to cause severe pneumonia in humans. The S protein of these CoVs binds to the ACE2 molecule on the plasma membrane and mediates virus entry into cells.
• Field performance of three Ebola rapid diagnostic tests used during the 2018–20 outbreak in the eastern Democratic Republic of the Congo: a retrospective, multicentre observational study

  Daniel Mukadi-Bamuleka, Junior Bulabula-Penge, Anja De Weggheleire, Bart K M Jacobs, François Edidi-Atani, Fabrice Mambu-Mbika, Placide Mbala-Kingebeni, Sheila Makiala-Mandanda, Martin Faye, Cheick T Diagne, Moussa M Diagne, Oumar Faye, Masahiro Kajihara, Ousmane Faye, Ayato Takada, Amadou A Sall, Jean-Jacques Muyembe-Tamfum, Johan van Griensven, Kevin K Ariën, Steve Ahuka-Mundeke

  The Lancet Infectious Diseases 22 (6) 891 - 900 1473-3099 2022/06
• Influenza A Virus Infection in Domestic Ferrets.

  Yudai Kuroda, Ai Okada, Hiroshi Shimoda, Yasutsugu Miwa, Akiko Watamori, Hiroho Ishida, Shin Murakami, Ayato Takada, Taisuke Horimoto, Ken Maeda

  Japanese journal of infectious diseases 75 (3) 325 - 327 2022/05/24 [Refereed][Invited]
   

  Ferrets are animals that are known to be susceptible to influenza A virus (IAV) infection. To evaluate the risk of IAV transmission from diseased ferrets to humans, we performed a serosurvey to detect specific antibodies against the H1, H3, H5, and H7 subtypes of IAV. We found a high positive rate of the H1 (24.1%) and H3 (5.2%) subtypes in pet ferrets by using an enzyme-linked immunosorbent assay for hemagglutinin proteins. The results were confirmed by the virus-neutralization test for representative antibody-positive serum samples. We also detected hemagglutinin and neuraminidase genes in two ferrets showing acute respiratory illness and whose owner was diagnosed with IAV infection; a human H1N1pdm virus was isolated from one of these ferrets. Our findings suggest that attention should be paid for IAV infection from humans to ferrets, and vice versa.
• Detection of Tick-Borne Bacterial and Protozoan Pathogens in Ticks from the Zambia-Angola Border.

  Yongjin Qiu, Martin Simuunza, Masahiro Kajihara, Joseph Ndebe, Ngonda Saasa, Penjani Kapila, Hayato Furumoto, Alice C C Lau, Ryo Nakao, Ayato Takada, Hirofumi Sawa

  Pathogens (Basel, Switzerland) 11 (5) 2022/05/10 [Refereed][Not invited]
   

  Tick-borne diseases (TBDs), including emerging and re-emerging zoonoses, are of public health importance worldwide; however, TBDs tend to be overlooked, especially in countries with fewer resources, such as Zambia and Angola. Here, we investigated Rickettsia, Anaplasmataceae, and Apicomplexan pathogens in 59 and 96 adult ticks collected from dogs and cattle, respectively, in Shangombo, a town at the Zambia-Angola border. We detected Richkettsia africae and Rickettsia aeschilimannii in 15.6% of Amblyomma variegatum and 41.7% of Hyalomma truncatum ticks, respectively. Ehrlichia minasensis was detected in 18.8% of H. truncatum, and Candidatus Midichloria mitochondrii was determined in Hyalomma marginatum. We also detected Babesia caballi and Theileria velifera in A. variegatum ticks with a 4.4% and 6.7% prevalence, respectively. In addition, Hepatozoon canis was detected in 6.5% of Rhipicephalus lunulatus and 4.3% of Rhipicephalus sanguineus. Coinfection of R. aeshilimannii and E. minasensis were observed in 4.2% of H. truncatum. This is the first report of Ca. M. mitochondrii and E. minasensis, and the second report of B. caballi, in the country. Rickettsia africae and R. aeschlimannii are pathogenic to humans, and E. minasensis, B. caballi, T. velifera, and H. canis are pathogenic to animals. Therefore, individuals, clinicians, veterinarians, and pet owners should be aware of the distribution of these pathogens in the area.
• Gargling with povidone iodine has a short-term inhibitory effect on SARS-CoV-2 in patients with COVID-19

  T. Seikai, A. Takada, A. Hasebe, M. Kajihara, K. Okuya, T. Sekiguchi (Yamada), W. Kakuguchi, S. Konno, Y. Ohiro

  Journal of Hospital Infection 123 179 - 181 0195-6701 2022/05 [Refereed][Not invited]
  
• Multiple Routes of Antibody-Dependent Enhancement of SARS-CoV-2 Infection.

  Kosuke Okuya, Takanari Hattori, Takeshi Saito, Yoshihiro Takadate, Michihito Sasaki, Wakako Furuyama, Andrea Marzi, Yoichi Ohiro, Satoshi Konno, Takeshi Hattori, Ayato Takada

  Microbiology spectrum 10 (2) e0155321  2022/04/27 [Refereed]
   

  Antibody-dependent enhancement (ADE) of infection is generally known for many viruses. A potential risk of ADE in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has also been discussed since the beginning of the coronavirus disease 2019 (COVID-19) pandemic; however, clinical evidence of the presence of antibodies with ADE potential is limited. Here, we show that ADE antibodies are produced by SARS-CoV-2 infection and the ADE process can be mediated by at least two different host factors, Fcγ receptor (FcγR) and complement component C1q. Of 89 serum samples collected from acute or convalescent COVID-19 patients, 62.9% were found to be positive for SARS-CoV-2-specific IgG. FcγR- and/or C1q-mediated ADE were detected in 50% of the IgG-positive sera, whereas most of them showed neutralizing activity in the absence of FcγR and C1q. Importantly, ADE antibodies were found in 41.4% of the acute COVID-19 patients. Neutralizing activity was also detected in most of the IgG-positive sera, but it was counteracted by ADE in subneutralizing conditions in the presence of FcγR or C1q. Although the clinical importance of ADE needs to be further investigated with larger numbers of COVID-19 patient samples, our data suggest that SARS-CoV-2 utilizes multiple mechanisms of ADE. C1q-mediated ADE may particularly have a clinical impact since C1q is present at high concentrations in plasma and its receptors are ubiquitously expressed on the surfaces of many types of cells, including respiratory epithelial cells, which SARS-CoV-2 primarily infects. IMPORTANCE Potential risks of antibody-dependent enhancement (ADE) in the coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has been discussed and the proposed mechanism mostly depends on the Fc gamma receptor (FcγR). However, since FcγRs are exclusively expressed on immune cells, which are not primary targets of SARS-CoV-2, the clinical importance of ADE of SARS-CoV-2 infection remains controversial. Our study demonstrates that SARS-CoV-2 infection induces antibodies that increase SARS-CoV-2 infection through another ADE mechanism in which complement component C1q mediates the enhancement. Although neutralizing activity was also detected in the serum samples, it was counteracted by ADE in the presence of FcγR or C1q. Considering the ubiquity of C1q and its cellular receptors, C1q-mediated ADE may more likely occur in respiratory epithelial cells, which SARS-CoV-2 primarily infects. Our data highlight the importance of careful monitoring of the antibody properties in COVID-19 convalescent and vaccinated individuals.
• Susceptibility of herons (family: Ardeidae) to clade 2.3.2.1 H5N1 subtype high pathogenicity avian influenza virus.

  Kosuke Soda, Yukiko Tomioka, Tatsufumi Usui, Yukiko Uno, Yasuko Nagai, Hiroshi Ito, Takahiro Hiono, Tomokazu Tamura, Masatoshi Okamatsu, Masahiro Kajihara, Naganori Nao, Yoshihiro Sakoda, Ayato Takada, Toshihiro Ito

  Avian pathology : journal of the W.V.P.A 51 (2) 146 - 153 2022/04 [Refereed]
   

  The pathogenicity of the H5 subtype high pathogenicity avian influenza viruses (HPAIVs) in Ardeidae bird species has not been investigated yet, despite the increasing infections reported. Therefore, the present study aimed to examine the susceptibility of the Ardeidae species, which had already been reported to be susceptible to HPAIVs, to a clade 2.3.2.1 H5N1 HPAIV. Juvenile herons (four grey herons, one intermediate egret, two little egrets, and three black-crowned night herons) were intranasally inoculated with 106 50% egg infectious dose of the virus and observed for 10 days. Two of the four grey herons showed lethargy and conjunctivitis; among them, one died at 6 days post-inoculation (dpi). The viruses were transmitted to the other two cohoused naïve grey herons. Some little egrets and black-crowned night herons showing neurological disorders died at 4-5 dpi; these birds mainly shed the virus via the oral route. The viruses predominantly replicated in the brains of birds that died of infection. Seroconversion was observed in most surviving birds, except some black-crowned night herons. These results demonstrate that most Ardeidae species are susceptible to H5 HPAIVs, sometimes with lethal effects. Herons are mostly colonial and often share habitats with Anseriformes, natural hosts of influenza A viruses; therefore, the risks of cluster infection and contribution to viral dissemination should be continuously evaluated. RESEARCH HIGHLIGHTSClade 2.3.2.1 H5N1 HPAIV causes lethal infections in Ardeidae sp.Viruses are transmitted among grey herons.Some herons with HPAIV showed conjunctivitis or neurological symptoms.HPAIV systemically replicated in herons tissues.
• Mapping of Antibody Epitopes on the Crimean-Congo Hemorrhagic Fever Virus Nucleoprotein.

  Boniface Pongombo Lombe, Takeshi Saito, Hiroko Miyamoto, Akina Mori-Kajihara, Masahiro Kajihara, Masayuki Saijo, Justin Masumu, Takanari Hattori, Manabu Igarashi, Ayato Takada

  Viruses 14 (3) 2022/03/06 [Refereed]
   

  Crimean-Congo hemorrhagic fever virus (CCHFV), a nairovirus, is a tick-borne zoonotic virus that causes hemorrhagic fever in humans. The CCHFV nucleoprotein (NP) is the antigen most used for serological screening of CCHFV infection in animals and humans. To gain insights into antibody epitopes on the NP molecule, we produced recombinant chimeric NPs between CCHFV and Nairobi sheep disease virus (NSDV), which is another nairovirus, and tested rabbit and mouse antisera/immune ascites, anti-NP monoclonal antibodies, and CCHFV-infected animal/human sera for their reactivities to the NP antigens. We found that the amino acids at positions 161-320 might include dominant epitopes recognized by anti-CCHFV IgG antibodies, whereas cross-reactivity between anti-CCHFV and anti-NSDV antibodies was limited. Their binding capacities were further tested using a series of synthetic peptides whose sequences were derived from CCHFV NP. IgG antibodies in CCHFV-infected monkeys and patients were reactive to some of the synthetic peptide antigens (e.g., amino acid residues at positions 131-150 and 211-230). Only a few peptides were recognized by IgG antibodies in the anti-NSDV serum. These results provide useful information to improve NP-based antibody detection assays as well as antigen detection tests relying on anti-NP monoclonal antibodies.
• Molecular Mechanisms Underlying the Cellular Entry and Host Range Restriction of Lujo Virus.

  Takeshi Saito, Takanari Hattori, Kosuke Okuya, Rashid Manzoor, Hiroko Miyamoto, Masahiro Kajihara, Ayato Takada

  mBio 13 (1) e0306021  2022/02/15 [Refereed]
   

  Like other human-pathogenic arenaviruses, Lujo virus (LUJV) is a causative agent of viral hemorrhagic fever in humans. LUJV infects humans with high mortality rates, but the susceptibilities of other animal species and the molecular determinants of its host specificity remain unknown. We found that mouse- and hamster-derived cell lines (NIH 3T3 and BHK, respectively) were less susceptible to a replication-incompetent recombinant vesicular stomatitis virus (Indiana) pseudotyped with the LUJV glycoprotein (GP) (VSVΔG*-LUJV/GP) than were human-derived cell lines (HEK293T and Huh7). To determine the cellular factors involved in the differential susceptibilities between the human and mouse cell lines, we focused on the CD63 molecule, which is required for pH-activated GP-mediated membrane fusion during LUJV entry into host cells. The exogenous introduction of human CD63, but not mouse or hamster CD63, into BHK cells significantly increased susceptibility to VSVΔG*-LUJV/GP. Using chimeric human-mouse CD63 proteins, we found that the amino acid residues at positions 141 to 150 in the large extracellular loop (LEL) region of CD63 were important for the cellular entry of VSVΔG*-LUJV/GP. By site-directed mutagenesis, we further determined that a phenylalanine at position 143 in human CD63 was the key residue for efficient membrane fusion and VSVΔG*-LUJV/GP infection. Our data suggest that the interaction of LUJV GP with the LEL region of CD63 is essential for cell susceptibility to LUJV, thus providing new insights into the molecular mechanisms underlying the cellular entry of LUJV and the host range restriction of this virus. IMPORTANCE Lujo virus (LUJV) infects humans with high mortality rates, but the host range of LUJV remains unknown. We found that rodent-derived cell lines were less susceptible to LUJV infection than were human-derived cell lines, and the differential susceptibilities were determined by the difference of CD63, the intercellular receptor of LUJV. We further identified an amino acid residue on human CD63 important for efficient LUJV infection. These results suggest that the interaction between LUJV glycoprotein and CD63 is one of the important factors determining the host range of LUJV. Our findings on the CD63-regulated susceptibilities of the cell lines to LUJV infection provide important information for the development of anti-LUJV drugs as well as the identification of natural hosts of LUJV. Importantly, our data support a concept explaining the molecular mechanism underlying viral tropisms controlled by endosomal receptors.
• Current knowledge of vector-borne zoonotic pathogens in Zambia: A clarion call to scaling-up "One Health" research in the wake of emerging and re-emerging infectious diseases.

  Benjamin Mubemba, Monicah M Mburu, Katendi Changula, Walter Muleya, Lavel C Moonga, Herman M Chambaro, Masahiro Kajihara, Yongjin Qiu, Yasuko Orba, Kyoko Hayashida, Catherine G Sutcliffe, Douglas E Norris, Philip E Thuma, Phillimon Ndubani, Simbarashe Chitanga, Hirofumi Sawa, Ayato Takada, Edgar Simulundu

  PLoS neglected tropical diseases 16 (2) e0010193  2022/02 [Refereed]
   

  BACKGROUND: Although vector-borne zoonotic diseases are a major public health threat globally, they are usually neglected, especially among resource-constrained countries, including those in sub-Saharan Africa. This scoping review examined the current knowledge and identified research gaps of vector-borne zoonotic pathogens in Zambia. METHODS AND FINDINGS: Major scientific databases (Web of Science, PubMed, Scopus, Google Scholar, CABI, Scientific Information Database (SID)) were searched for articles describing vector-borne (mosquitoes, ticks, fleas and tsetse flies) zoonotic pathogens in Zambia. Several mosquito-borne arboviruses have been reported including Yellow fever, Ntaya, Mayaro, Dengue, Zika, West Nile, Chikungunya, Sindbis, and Rift Valley fever viruses. Flea-borne zoonotic pathogens reported include Yersinia pestis and Rickettsia felis. Trypanosoma sp. was the only tsetse fly-borne pathogen identified. Further, tick-borne zoonotic pathogens reported included Crimean-Congo Haemorrhagic fever virus, Rickettsia sp., Anaplasma sp., Ehrlichia sp., Borrelia sp., and Coxiella burnetii. CONCLUSIONS: This study revealed the presence of many vector-borne zoonotic pathogens circulating in vectors and animals in Zambia. Though reports of human clinical cases were limited, several serological studies provided considerable evidence of zoonotic transmission of vector-borne pathogens in humans. However, the disease burden in humans attributable to vector-borne zoonotic infections could not be ascertained from the available reports and this precludes the formulation of national policies that could help in the control and mitigation of the impact of these diseases in Zambia. Therefore, there is an urgent need to scale-up "One Health" research in emerging and re-emerging infectious diseases to enable the country to prepare for future epidemics, including pandemics.
• Seroprevalence and Risk Factors of Crimean-Congo Hemorrhagic Fever in Cattle of Smallholder Farmers in Central Malawi.

  Marvin Collen Phonera, Martin Chitolongo Simuunza, Henson Kainga, Joseph Ndebe, Mwelwa Chembensofu, Elisha Chatanga, Setiala Kanyanda, Katendi Changula, Walter Muleya, Benjamin Mubemba, Simbarashe Chitanga, Masahiro Kajihara, Hirofumi Sawa, Gilson Njunga, Ayato Takada, Edgar Simulundu

  Pathogens (Basel, Switzerland) 10 (12) 2021/12/10 [Refereed]
   

  Crimean-Congo hemorrhagic fever virus (CCHFV) is endemic in Africa, Asia, and Eastern Europe where it circulates among animals and ticks causing sporadic outbreaks in humans. Although CCHF is endemic in sub-Saharan Africa, epidemiological information is lacking in many countries, including Malawi. To assess the risk of CCHF in Malawi, we conducted an epidemiological study in cattle reared by smallholder livestock farmers in central Malawi. A cross-sectional study was conducted in April 2020 involving seven districts, four from Kasungu and three from Lilongwe Agriculture Development Divisions. A structured questionnaire was administered to farmers to obtain demographic, animal management, and ecological risk factors data. Sera were collected from randomly selected cattle and screened for CCHF virus (CCHFV) specific antibodies using a commercial ELISA kit. Ticks were collected from cattle and classified morphologically to species level. An overall CCHFV seropositivity rate of 46.9% (n = 416; 95% CI: 42.0-51.8%) was observed. The seropositivity was significantly associated with the age of cattle (p < 0.001), sex (p < 0.001), presence of ticks in herds (p = 0.01), district (p = 0.025), and type of grazing lands (p = 0.013). Five species of ticks were identified, including Hyalomma truncatum, a known vector of CCHFV. Ticks of the species Hyalomma truncatum were not detected in two districts with the highest seroprevalence for CCHF and vector competency must be further explored in the study area. To our knowledge, this is the first report of serologic evidence of the presence of CCHV among smallholder cattle in central Malawi. This study emphasizes the need for continued monitoring of CCHFV infection among livestock, ticks, and humans for the development of data-based risk mitigation strategies.
• Structural Requirements in the Hemagglutinin Cleavage Site-Coding RNA Region for the Generation of Highly Pathogenic Avian Influenza Virus.

  Yurie Kida, Kosuke Okuya, Takeshi Saito, Junya Yamagishi, Aiko Ohnuma, Takanari Hattori, Hiroko Miyamoto, Rashid Manzoor, Reiko Yoshida, Naganori Nao, Masahiro Kajihara, Tokiko Watanabe, Ayato Takada

  Pathogens (Basel, Switzerland) 10 (12) 2021/12/09 [Refereed]
   

  Highly pathogenic avian influenza viruses (HPAIVs) with H5 and H7 hemagglutinin (HA) subtypes are derived from their low pathogenic counterparts following the acquisition of multiple basic amino acids in their HA cleavage site. It has been suggested that consecutive adenine residues and a stem-loop structure in the viral RNA region that encodes the cleavage site are essential for the acquisition of the polybasic cleavage site. By using a reporter assay to detect non-templated nucleotide insertions, we found that insertions more frequently occurred in the RNA region (29 nucleotide-length) encoding the cleavage site of an H5 HA gene that was predicted to have a stem-loop structure containing consecutive adenines than in a mutated corresponding RNA region that had a disrupted loop structure with fewer adenines. In virus particles generated by using reverse genetics, nucleotide insertions that created additional codons for basic amino acids were found in the RNA region encoding the cleavage site of an H5 HA gene but not in the mutated RNA region. We confirmed the presence of virus clones with the ability to replicate without trypsin in a plaque assay and to cause lethal infection in chicks. These results demonstrate that the stem-loop structure containing consecutive adenines in HA genes is a key molecular determinant for the emergence of H5 HPAIVs.
• Influenza A and D Viruses in Non-Human Mammalian Hosts in Africa: A Systematic Review and Meta-Analysis.

  Annie Kalonda, Marvin Phonera, Ngonda Saasa, Masahiro Kajihara, Catherine G Sutcliffe, Hirofumi Sawa, Ayato Takada, Edgar Simulundu

  Viruses 13 (12) 2021/12/02 [Refereed]
   

  We conducted a systematic review and meta-analysis to investigate the prevalence and current knowledge of influenza A virus (IAV) and influenza D virus (IDV) in non-human mammalian hosts in Africa. PubMed, Google Scholar, Wiley Online Library and World Organisation for Animal Health (OIE-WAHIS) were searched for studies on IAV and IDV from 2000 to 2020. Pooled prevalence and seroprevalences were estimated using the quality effects meta-analysis model. The estimated pooled prevalence and seroprevalence of IAV in pigs in Africa was 1.6% (95% CI: 0-5%) and 14.9% (95% CI: 5-28%), respectively. The seroprevalence of IDV was 87.2% (95% CI: 24-100%) in camels, 9.3% (95% CI: 0-24%) in cattle, 2.2% (95% CI: 0-4%) in small ruminants and 0.0% (95% CI: 0-2%) in pigs. In pigs, H1N1 and H1N1pdm09 IAVs were commonly detected. Notably, the highly pathogenic H5N1 virus was also detected in pigs. Other subtypes detected serologically and/or virologically included H3N8 and H7N7 in equids, H1N1, and H3N8 and H5N1 in dogs and cats. Furthermore, various wildlife animals were exposed to different IAV subtypes. For prudent mitigation of influenza epizootics and possible human infections, influenza surveillance efforts in Africa should not neglect non-human mammalian hosts. The impact of IAV and IDV in non-human mammalian hosts in Africa deserves further investigation.
• Correction to: 2021 Taxonomic update of phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales.

  Jens H Kuhn, Scott Adkins, Bernard R Agwanda, Rim Al Kubrusli, Sergey V Alkhovsky, Gaya K Amarasinghe, Tatjana Avšič-Županc, María A Ayllón, Justin Bahl, Anne Balkema-Buschmann, Matthew J Ballinger, Christopher F Basler, Sina Bavari, Martin Beer, Nicolas Bejerman, Andrew J Bennett, Dennis A Bente, Éric Bergeron, Brian H Bird, Carol D Blair, Kim R Blasdell, Dag-Ragnar Blystad, Jamie Bojko, Wayne B Borth, Steven Bradfute, Rachel Breyta, Thomas Briese, Paul A Brown, Judith K Brown, Ursula J Buchholz, Michael J Buchmeier, Alexander Bukreyev, Felicity Burt, Carmen Büttner, Charles H Calisher, Mengji Cao, Inmaculada Casas, Kartik Chandran, Rémi N Charrel, Qi Cheng, Yuya Chiaki, Marco Chiapello, Il-Ryong Choi, Marina Ciuffo, J Christopher S Clegg, Ian Crozier, Elena Dal Bó, Juan Carlos de la Torre, Xavier de Lamballerie, Rik L de Swart, Humberto Debat, Nolwenn M Dheilly, Emiliano Di Cicco, Nicholas Di Paola, Francesco Di Serio, Ralf G Dietzgen, Michele Digiaro, Olga Dolnik, Michael A Drebot, J Felix Drexler, William G Dundon, W Paul Duprex, Ralf Dürrwald, John M Dye, Andrew J Easton, Hideki Ebihara, Toufic Elbeaino, Koray Ergünay, Hugh W Ferguson, Anthony R Fooks, Marco Forgia, Pierre B H Formenty, Jana Fránová, Juliana Freitas-Astúa, Jingjing Fu, Stephanie Fürl, Selma Gago-Zachert, George Fú Gāo, María Laura García, Adolfo García-Sastre, Aura R Garrison, Thomas Gaskin, Jean-Paul J Gonzalez, Anthony Griffiths, Tony L Goldberg, Martin H Groschup, Stephan Günther, Roy A Hall, John Hammond, Tong Han, Jussi Hepojoki, Roger Hewson, Jiang Hong, Ni Hong, Seiji Hongo, Masayuki Horie, John S Hu, Tao Hu, Holly R Hughes, Florian Hüttner, Timothy H Hyndman, M Ilyas, Risto Jalkanen, Dàohóng Jiāng, Gilda B Jonson, Sandra Junglen, Fujio Kadono, Karia H Kaukinen, Michael Kawate, Boris Klempa, Jonas Klingström, Gary Kobinger, Igor Koloniuk, Hideki Kondō, Eugene V Koonin, Mart Krupovic, Kenji Kubota, Gael Kurath, Lies Laenen, Amy J Lambert, Stanley L Langevin, Benhur Lee, Elliot J Lefkowitz, Eric M Leroy, Shaorong Li, Longhui Li, Jiànróng Lǐ, Huazhen Liu, Igor S Lukashevich, Piet Maes, William Marciel de Souza, Marco Marklewitz, Sergio H Marshall, Shin-Yi L Marzano, Sebastien Massart, John W McCauley, Michael Melzer, Nicole Mielke-Ehret, Kristina M Miller, Tobi J Ming, Ali Mirazimi, Gideon J Mordecai, Hans-Peter Mühlbach, Elke Mühlberger, Rayapati Naidu, Tomohide Natsuaki, José A Navarro, Sergey V Netesov, Gabriele Neumann, Norbert Nowotny, Márcio R T Nunes, Alejandro Olmedo-Velarde, Gustavo Palacios, Vicente Pallás, Bernadett Pályi, Anna Papa, Sofia Paraskevopoulou, Adam C Park, Colin R Parrish, David A Patterson, Alex Pauvolid-Corrêa, Janusz T Pawęska, Susan Payne, Carlotta Peracchio, Daniel R Pérez, Thomas S Postler, Liying Qi, Sheli R Radoshitzky, Renato O Resende, Carina A Reyes, Bertus K Rima, Gabriel Robles Luna, Víctor Romanowski, Paul Rota, Dennis Rubbenstroth, Luisa Rubino, Jonathan A Runstadler, Sead Sabanadzovic, Amadou Alpha Sall, Maria S Salvato, Rosemary Sang, Takahide Sasaya, Angela D Schulze, Martin Schwemmle, Mang Shi, Xiǎohóng Shí, Zhènglì Shí, Yoshifumi Shimomoto, Yukio Shirako, Stuart G Siddell, Peter Simmonds, Manuela Sironi, Guy Smagghe, Sophie Smither, Jin-Won Song, Kirsten Spann, Jessica R Spengler, Mark D Stenglein, David M Stone, Jari Sugano, Curtis A Suttle, Amy Tabata, Ayato Takada, Shigeharu Takeuchi, David P Tchouassi, Amy Teffer, Robert B Tesh, Natalie J Thornburg, Yasuhiro Tomitaka, Keizō Tomonaga, Noël Tordo, Baldwyn Torto, Jonathan S Towner, Shinya Tsuda, Changchun Tu, Massimo Turina, Ioannis E Tzanetakis, Janice Uchida, Tomio Usugi, Anna Maria Vaira, Marta Vallino, Bernadette van den Hoogen, Arvind Varsani, Nikos Vasilakis, Martin Verbeek, Susanne von Bargen, Jiro Wada, Victoria Wahl, Peter J Walker, Lin-Fa Wang, Guoping Wang, Yanxiang Wang, Yaqin Wang, Muhammad Waqas, Tàiyún Wèi, Shaohua Wen, Anna E Whitfield, John V Williams, Yuri I Wolf, Jiangxiang Wu, Lei Xu, Hironobu Yanagisawa, Caixia Yang, Zuokun Yang, F Murilo Zerbini, Lifeng Zhai, Yong-Zhen Zhang, Song Zhang, Jinguo Zhang, Zhe Zhang, Xueping Zhou

  Archives of virology 166 (12) 3567 - 3579 2021/12 [Refereed]
  
• Functional Importance of Hydrophobic Patches on the Ebola Virus VP35 IFN-Inhibitory Domain.

  Nodoka Kasajima, Keita Matsuno, Hiroko Miyamoto, Masahiro Kajihara, Manabu Igarashi, Ayato Takada

  Viruses 13 (11) 2021/11/20 [Refereed]
   

  Viral protein 35 (VP35) of Ebola virus (EBOV) is a multifunctional protein that mainly acts as a viral polymerase cofactor and an interferon antagonist. VP35 interacts with the viral nucleoprotein (NP) and double-stranded RNA for viral RNA transcription/replication and inhibition of type I interferon (IFN) production, respectively. The C-terminal portion of VP35, which is termed the IFN-inhibitory domain (IID), is important for both functions. To further identify critical regions in this domain, we analyzed the physical properties of the surface of VP35 IID, focusing on hydrophobic patches, which are expected to be functional sites that are involved in interactions with other molecules. Based on the known structural information of VP35 IID, three hydrophobic patches were identified on its surface and their biological importance was investigated using minigenome and IFN-β promoter-reporter assays. Site-directed mutagenesis revealed that some of the amino acid substitutions that were predicted to disrupt the hydrophobicity of the patches significantly decreased the efficiency of viral genome replication/transcription due to reduced interaction with NP, suggesting that the hydrophobic patches might be critical for the formation of a replication complex through the interaction with NP. It was also found that the hydrophobic patches were involved in the IFN-inhibitory function of VP35. These results highlight the importance of hydrophobic patches on the surface of EBOV VP35 IID and also indicate that patch analysis is useful for the identification of amino acid residues that directly contribute to protein functions.
• Serological and molecular epidemiological study on swine influenza in Zambia.

  Hayato Harima, Kosuke Okuya, Masahiro Kajihara, Hirohito Ogawa, Edgar Simulundu, Eugene Bwalya, Yongjin Qiu, Akina Mori-Kajihara, Musso Munyeme, Yoshihiro Sakoda, Takehiko Saito, Bernard M Hang'ombe, Hirofumi Sawa, Aaron S Mweene, Ayato Takada

  Transboundary and emerging diseases 69 (4) e931-e943  2021/11/01 [Refereed]
   

  Influenza A viruses (IAVs) cause highly contagious respiratory diseases in humans and animals. In 2009, a swine-origin pandemic H1N1 IAV, designated A(H1N1)pdm09 virus, spread worldwide, and has since frequently been introduced into pig populations. Since novel reassortant IAVs with pandemic potential may emerge in pigs, surveillance for IAV in pigs is therefore necessary not only for the pig industry but also for public health. However, epidemiological information on IAV infection of pigs in Africa remains sparse. In this study, we collected 246 serum and 605 nasal swab samples from pigs in Zambia during the years 2011-2018. Serological analyses revealed that 49% and 32% of the sera collected in 2011 were positive for hemagglutination-inhibition (HI) and neutralizing antibodies against A(H1N1)pdm09 virus, respectively, whereas less than 5.3% of sera collected during the following period (2012-2018) were positive in both serological tests. The positive rate and the neutralization titres to A(H1N1)pdm09 virus were higher than those to classical swine H1N1 and H1N2 IAVs. On the other hand, the positive rate for swine H3N2 IAV was very low in the pig population in Zambia in 2011-2018 (5.3% and 0% in HI and neutralization tests, respectively). From nasal swab samples, we isolated one H3N2 and eight H1N1 IAV strains with an isolation rate of 1.5%. Phylogenetic analyses of all eight gene segments revealed that the isolated IAVs were closely related to human IAV strains belonging to A(H1N1)pdm09 and seasonal H3N2 lineages. Our findings indicate that reverse zoonotic transmission from humans to pigs occurred during the study period in Zambia and highlight the need for continued surveillance to monitor the status of IAVs circulating in swine populations in Africa.
• 北海道の野鳥から検出されたロタウイルスA JC-105株の遺伝学的解析

  藤井 祐至, 正谷 達謄, 西山 祥子, 藤原 拓朗, 迫田 義博, 高田 礼人, 小澤 真, 杉山 誠, 伊藤 直人

  日本獣医学会学術集会講演要旨集 (公社)日本獣医学会 164回 [FO - 1] 1347-8621 2021/09
• Attenuated infection by a Pteropine orthoreovirus isolated from an Egyptian fruit bat in Zambia.

  Hayato Harima, Michihito Sasaki, Yasuko Orba, Kosuke Okuya, Yongjin Qiu, Christida E Wastika, Katendi Changula, Masahiro Kajihara, Edgar Simulundu, Tomoyuki Yamaguchi, Yoshiki Eto, Akina Mori-Kajihara, Akihiko Sato, Satoshi Taniguchi, Ayato Takada, Masayuki Saijo, Bernard M Hang'ombe, Hirofumi Sawa

  PLoS neglected tropical diseases 15 (9) e0009768  2021/09 [Refereed]
   

  BACKGROUND: Pteropine orthoreovirus (PRV) is an emerging bat-borne zoonotic virus that causes severe respiratory illness in humans. Although PRVs have been identified in fruit bats and humans in Australia and Asia, little is known about the prevalence of PRV infection in Africa. Therefore, this study performed an PRV surveillance in fruit bats in Zambia. METHODS: Egyptian fruit bats (Rousettus aegyptiacus, n = 47) and straw-colored fruit bats (Eidolon helvum, n = 33) captured in Zambia in 2017-2018 were screened for PRV infection using RT-PCR and serum neutralization tests. The complete genome sequence of an isolated PRV strain was determined by next generation sequencing and subjected to BLAST and phylogenetic analyses. Replication capacity and pathogenicity of the strain were investigated using Vero E6 cell cultures and BALB/c mice, respectively. RESULTS: An PRV strain, tentatively named Nachunsulwe-57, was isolated from one Egyptian fruit bat. Serological assays demonstrated that 98% of sera (69/70) collected from Egyptian fruit bats (n = 37) and straw-colored fruit bats (n = 33) had neutralizing antibodies against PRV. Genetic analyses revealed that all 10 genome segments of Nachunsulwe-57 were closely related to a bat-derived Kasama strain found in Uganda. Nachunsulwe-57 showed less efficiency in viral growth and lower pathogenicity in mice than another PRV strain, Miyazaki-Bali/2007, isolated from a patient. CONCLUSIONS: A high proportion of Egyptian fruit bats and straw-colored fruit bats were found to be seropositive to PRV in Zambia. Importantly, a new PRV strain (Nachunsulwe-57) was isolated from an Egyptian fruit bat in Zambia, which had relatively weak pathogenicity in mice. Taken together, our findings provide new epidemiological insights about PRV infection in bats and indicate the first isolation of an PRV strain that may have low pathogenicity to humans.
• 2021 Taxonomic update of phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales.

  Jens H Kuhn, Scott Adkins, Bernard R Agwanda, Rim Al Kubrusli, Sergey V Alkhovsky Aльxoвcкий Cepгeй Bлaдимиpoвич, Gaya K Amarasinghe, Tatjana Avšič-Županc, María A Ayllón, Justin Bahl, Anne Balkema-Buschmann, Matthew J Ballinger, Christopher F Basler, Sina Bavari, Martin Beer, Nicolas Bejerman, Andrew J Bennett, Dennis A Bente, Éric Bergeron, Brian H Bird, Carol D Blair, Kim R Blasdell, Dag-Ragnar Blystad, Jamie Bojko, Wayne B Borth, Steven Bradfute, Rachel Breyta, Thomas Briese, Paul A Brown, Judith K Brown, Ursula J Buchholz, Michael J Buchmeier, Alexander Bukreyev, Felicity Burt, Carmen Büttner, Charles H Calisher, Mengji Cao 曹孟籍, Inmaculada Casas, Kartik Chandran, Rémi N Charrel, Qi Cheng, Yuya Chiaki 千秋祐也, Marco Chiapello, Il-Ryong Choi, Marina Ciuffo, J Christopher S Clegg, Ian Crozier, Elena Dal Bó, Juan Carlos de la Torre, Xavier de Lamballerie, Rik L de Swart, Humberto Debat, Nolwenn M Dheilly, Emiliano Di Cicco, Nicholas Di Paola, Francesco Di Serio, Ralf G Dietzgen, Michele Digiaro, Olga Dolnik, Michael A Drebot, J Felix Drexler, William G Dundon, W Paul Duprex, Ralf Dürrwald, John M Dye, Andrew J Easton, Hideki Ebihara 海老原秀喜, Toufic Elbeaino, Koray Ergünay, Hugh W Ferguson, Anthony R Fooks, Marco Forgia, Pierre B H Formenty, Jana Fránová, Juliana Freitas-Astúa, Jingjing Fu 付晶晶, Stephanie Fürl, Selma Gago-Zachert, George Fú Gāo 高福, María Laura García, Adolfo García-Sastre, Aura R Garrison, Thomas Gaskin, Jean-Paul J Gonzalez, Anthony Griffiths, Tony L Goldberg, Martin H Groschup, Stephan Günther, Roy A Hall, John Hammond, Tong Han 韩彤, Jussi Hepojoki, Roger Hewson, Jiang Hong 洪健, Ni Hong 洪霓, Seiji Hongo 本郷誠治, Masayuki Horie 堀江真行, John S Hu, Tao Hu, Holly R Hughes, Florian Hüttner, Timothy H Hyndman, M Ilyas, Risto Jalkanen, Dàohóng Jiāng 姜道宏, Gilda B Jonson, Sandra Junglen, Fujio Kadono 上遠野冨士夫, Karia H Kaukinen, Michael Kawate, Boris Klempa, Jonas Klingström, Gary Kobinger, Igor Koloniuk, Hideki Kondō 近藤秀樹, Eugene V Koonin, Mart Krupovic, Kenji Kubota 久保田健嗣, Gael Kurath, Lies Laenen, Amy J Lambert, Stanley L Langevin, Benhur Lee, Elliot J Lefkowitz, Eric M Leroy, Shaorong Li 李邵蓉, Longhui Li 李龙辉, Jiànróng Lǐ 李建荣, Huazhen Liu 刘华珍, Igor S Lukashevich, Piet Maes, William Marciel de Souza, Marco Marklewitz, Sergio H Marshall, Shin-Yi L Marzano, Sebastien Massart, John W McCauley, Michael Melzer, Nicole Mielke-Ehret, Kristina M Miller, Tobi J Ming, Ali Mirazimi, Gideon J Mordecai, Hans-Peter Mühlbach, Elke Mühlberger, Rayapati Naidu, Tomohide Natsuaki 夏秋知英, José A Navarro, Sergey V Netesov Heтёcoв Cepгeй Bиктopoвич, Gabriele Neumann, Norbert Nowotny, Márcio R T Nunes, Alejandro Olmedo-Velarde, Gustavo Palacios, Vicente Pallás, Bernadett Pályi, Anna Papa Άννα Παπά, Sofia Paraskevopoulou Σοφία Παρασκευοπούλου, Adam C Park, Colin R Parrish, David A Patterson, Alex Pauvolid-Corrêa, Janusz T Pawęska, Susan Payne, Carlotta Peracchio, Daniel R Pérez, Thomas S Postler, Liying Qi 亓立莹, Sheli R Radoshitzky, Renato O Resende, Carina A Reyes, Bertus K Rima, Gabriel Robles Luna, Víctor Romanowski, Paul Rota, Dennis Rubbenstroth, Luisa Rubino, Jonathan A Runstadler, Sead Sabanadzovic, Amadou Alpha Sall, Maria S Salvato, Rosemary Sang, Takahide Sasaya 笹谷孝英, Angela D Schulze, Martin Schwemmle, Mang Shi 施莽, Xiǎohóng Shí 石晓宏, Zhènglì Shí 石正丽, Yoshifumi Shimomoto 下元祥史, Yukio Shirako, Stuart G Siddell, Peter Simmonds, Manuela Sironi, Guy Smagghe, Sophie Smither, Jin-Won Song 송진원, Kirsten Spann, Jessica R Spengler, Mark D Stenglein, David M Stone, Jari Sugano, Curtis A Suttle, Amy Tabata, Ayato Takada 高田礼人, Shigeharu Takeuchi 竹内繁治, David P Tchouassi, Amy Teffer, Robert B Tesh, Natalie J Thornburg, Yasuhiro Tomitaka 冨高保弘, Keizō Tomonaga 朝長啓造, Noël Tordo, Baldwyn Torto, Jonathan S Towner, Shinya Tsuda 津田新哉, Changchun Tu 涂长春, Massimo Turina, Ioannis E Tzanetakis, Janice Uchida, Tomio Usugi 宇杉富雄, Anna Maria Vaira, Marta Vallino, Bernadette van den Hoogen, Arvind Varsani, Nikos Vasilakis Νίκος Βασιλάκης, Martin Verbeek, Susanne von Bargen, Jiro Wada 和田治郎, Victoria Wahl, Peter J Walker, Lin-Fa Wang 王林发, Guoping Wang 王国平, Yanxiang Wang 王雁翔, Yaqin Wang 王亚琴, Muhammad Waqas, Tàiyún Wèi 魏太云, Shaohua Wen 温少华, Anna E Whitfield, John V Williams, Yuri I Wolf, Jiangxiang Wu 吴建祥, Lei Xu 徐雷, Hironobu Yanagisawa 栁澤広宣, Caixia Yang 杨彩霞, Zuokun Yang 杨作坤, F Murilo Zerbini, Lifeng Zhai 翟立峰, Yong-Zhen Zhang 张永振, Song Zhang 张松, Jinguo Zhang 张靖国, Zhe Zhang 张哲, Xueping Zhou 周雪平

  Archives of virology 166 (12) 3513 - 3566 2021/08/31 [Refereed]
   

  In March 2021, following the annual International Committee on Taxonomy of Viruses (ICTV) ratification vote on newly proposed taxa, the phylum Negarnaviricota was amended and emended. The phylum was expanded by four families (Aliusviridae, Crepuscuviridae, Myriaviridae, and Natareviridae), three subfamilies (Alpharhabdovirinae, Betarhabdovirinae, and Gammarhabdovirinae), 42 genera, and 200 species. Thirty-nine species were renamed and/or moved and seven species were abolished. This article presents the updated taxonomy of Negarnaviricota as now accepted by the ICTV.
• Dual Effect of Organogermanium Compound THGP on RIG-I-Mediated Viral Sensing and Viral Replication during Influenza a Virus Infection.

  Sunanda Baidya, Yoko Nishimoto, Seiichi Sato, Yasuhiro Shimada, Nozomi Sakurai, Hirotaka Nonaka, Koki Noguchi, Mizuki Kido, Satoshi Tadano, Kozo Ishikawa, Kai Li, Aoi Okubo, Taisho Yamada, Yasuko Orba, Michihito Sasaki, Hirofumi Sawa, Hiroko Miyamoto, Ayato Takada, Takashi Nakamura, Akinori Takaoka

  Viruses 13 (9) 2021/08/24 [Refereed]
   

  The interaction of viral nucleic acid with protein factors is a crucial process for initiating viral polymerase-mediated viral genome replication while activating pattern recognition receptor (PRR)-mediated innate immune responses. It has previously been reported that a hydrolysate of Ge-132, 3-(trihydroxygermyl) propanoic acid (THGP), shows a modulatory effect on microbial infections, inflammation, and immune responses. However, the detailed mechanism by which THGP can modify these processes during viral infections remained unknown. Here, we show that THGP can specifically downregulate type I interferon (IFN) production in response to stimulation with a cytosolic RNA sensor RIG-I ligand 5'-triphosphate RNA (3pRNA) but not double-stranded RNA, DNA, or lipopolysaccharide. Consistently, treatment with THGP resulted in the dose-dependent suppression of type I IFN induction upon infections with influenza virus (IAV) and vesicular stomatitis virus, which are known to be mainly sensed by RIG-I. Mechanistically, THGP directly binds to the 5'-triphosphate moiety of viral RNA and competes with RIG-I-mediated recognition. Furthermore, we found that THGP can directly counteract the replication of IAV but not EMCV (encephalitismyocarditis virus), by inhibiting the interaction of viral polymerase with RNA genome. Finally, IAV RNA levels were significantly reduced in the lung tissues of THGP-treated mice when compared with untreated mice. These results suggest a possible therapeutic implication of THGP and show direct antiviral action, together with the suppressive activity of innate inflammation.
• Mosquito-Borne Viral Pathogens Detected in Zambia: A Systematic Review.

  Rachel Milomba Velu, Geoffrey Kwenda, Liyali Libonda, Caroline Cleopatra Chisenga, Bumbangi Nsoni Flavien, Obvious Nchimunya Chilyabanyama, Michelo Simunyandi, Samuel Bosomprah, Nicholus Chintu Sande, Katendi Changula, Walter Muleya, Monicah Mirai Mburu, Benjamin Mubemba, Simbarashe Chitanga, John Tembo, Matthew Bates, Nathan Kapata, Yasuko Orba, Masahiro Kajihara, Ayato Takada, Hirofumi Sawa, Roma Chilengi, Edgar Simulundu

  Pathogens (Basel, Switzerland) 10 (8) 2021/08/10 [Refereed]
   

  Emerging and re-emerging mosquito-borne viral diseases are a threat to global health. This systematic review aimed to investigate the available evidence of mosquito-borne viral pathogens reported in Zambia. A search of literature was conducted in PubMed and Google Scholar for articles published from 1 January 1930 to 30 June 2020 using a combination of keywords. Eight mosquito-borne viruses belonging to three families, Togaviridae, Flaviviridae and Phenuiviridae were reported. Three viruses (Chikungunya virus, Mayaro virus, Mwinilunga virus) were reported among the togaviruses whilst four (dengue virus, West Nile virus, yellow fever virus, Zika virus) were among the flavivirus and only one virus, Rift Valley fever virus, was reported in the Phenuiviridae family. The majority of these mosquito-borne viruses were reported in Western and North-Western provinces. Aedes and Culex species were the main mosquito-borne viral vectors reported. Farming, fishing, movement of people and rain patterns were among factors associated with mosquito-borne viral infection in Zambia. Better diagnostic methods, such as the use of molecular tools, to detect the viruses in potential vectors, humans, and animals, including the recognition of arboviral risk zones and how the viruses circulate, are important for improved surveillance and design of effective prevention and control measures.
• Screening of tick-borne pathogens in argasid ticks in Zambia: Expansion of the geographic distribution of Rickettsia lusitaniae and Rickettsia hoogstraalii and detection of putative novel Anaplasma species.

  Yongjin Qiu, Martin Simuunza, Masahiro Kajihara, Herman Chambaro, Hayato Harima, Yoshiki Eto, Edgar Simulundu, David Squarre, Shiho Torii, Ayato Takada, Bernard Mudenda Hang'ombe, Hirofumi Sawa, Chihiro Sugimoto, Ryo Nakao

  Ticks and tick-borne diseases 12 (4) 101720 - 101720 2021/07 [Refereed]
   

  Ticks (Ixodidae and Argasidae) are important arthropod vectors of various pathogens that cause human and animal infectious diseases. Many previously published studies on tick-borne pathogens focused on those transmitted by ixodid ticks. Although there are increasing reports of viral pathogens associated with argasid ticks, information on bacterial pathogens they transmit is scarce. The aim of this molecular study was to detect and characterize Rickettsia and Anaplasmataceae in three different argasid tick species, Ornithodoros faini, Ornithodoros moubata, and Argas walkerae collected in Zambia. Rickettsia hoogstraalii and Rickettsia lusitaniae were detected in 77 % (77/100) of Ar. walkerae and 10 % (5/50) of O. faini, respectively. All O. moubata pool samples (n = 124) were negative for rickettsial infections. Anaplasmataceae were detected in 63 % (63/100) of Ar. walkerae and in 82.2 % (102/124) of O. moubata pools, but not in O. faini. Phylogenetic analysis based on the concatenated sequences of 16S rRNA and groEL genes revealed that Anaplasma spp. detected in the present study were distinct from previously validated Anaplasma species, indicating that the current knowledge on the diversity and vector range of Anaplasma spp. is incomplete. Our findings highlight new geographical records of R. lusitaniae and R. hoogstraalii and confirm that the wide geographic distribution of these species includes the African continent. The data presented here increase our knowledge on argasid tick-borne bacteria and contribute toward understanding their epidemiology.
• Serological Evidence of Filovirus Infection in Nonhuman Primates in Zambia.

  Katendi Changula, Edgar Simulundu, Boniface Pongombo Lombe, Eri Nakayama, Hiroko Miyamoto, Yuji Takahashi, Hirofumi Sawa, Chuma Simukonda, Bernard M Hang'ombe, Ayato Takada

  Viruses 13 (7) 2021/06/30 [Refereed]
   

  Ebolaviruses and marburgviruses are filoviruses that are known to cause severe hemorrhagic fever in humans and nonhuman primates (NHPs). While some bat species are suspected to be natural reservoirs of these filoviruses, wild NHPs often act as intermediate hosts for viral transmission to humans. Using an enzyme-linked immunosorbent assay, we screened two NHP species, wild baboons and vervet monkeys captured in Zambia, for their serum IgG antibodies specific to the envelope glycoproteins of filoviruses. From 243 samples tested, 39 NHPs (16%) were found to be seropositive either for ebolaviruses or marburgviruses with endpoint antibody titers ranging from 100 to 25,600. Interestingly, antibodies reactive to Reston virus, which is found only in Asia, were detected in both NHP species. There was a significant difference in the seropositivity for the marburgvirus antigen between the two NHP species, with baboons having a higher positive rate. These results suggest that wild NHPs in Zambia might be nonlethally exposed to these filoviruses, and this emphasizes the need for continuous monitoring of filovirus infection in wild animals to better understand the ecology of filoviruses and to assess potential risks of outbreaks in humans in previously nonendemic countries.
• Molecular Detection and Genotyping of Coxiella-Like Endosymbionts in Ticks Collected from Animals and Vegetation in Zambia.

  Toshiya Kobayashi, Elisha Chatanga, Yongjin Qiu, Martin Simuunza, Masahiro Kajihara, Bernard Mudenda Hang'ombe, Yoshiki Eto, Ngonda Saasa, Akina Mori-Kajihara, Edgar Simulundu, Ayato Takada, Hirofumi Sawa, Ken Katakura, Nariaki Nonaka, Ryo Nakao

  Pathogens (Basel, Switzerland) 10 (6) 2021/06/21 [Refereed]
   

  Ticks are obligate ectoparasites as they require to feed on their host blood during some or all stages of their life cycle. In addition to the pathogens that ticks harbor and transmit to vertebrate hosts, they also harbor other seemingly nonpathogenic microorganisms including nutritional mutualistic symbionts. Tick nutritional mutualistic symbionts play important roles in the physiology of the host ticks as they are involved in tick reproduction and growth through the supply of B vitamins as well as in pathogen maintenance and propagation. Coxiella-like endosymbionts (CLEs) are the most widespread endosymbionts exclusively reported in ticks. Although CLEs have been investigated in ticks in other parts of the world, there is no report of their investigation in ticks in Zambia. To investigate the occurrence of CLEs, their maintenance, and association with host ticks in Zambia, 175 ticks belonging to six genera, namely Amblyomma, Argas, Haemaphysalis, Hyalomma, Ornithodoros, and Rhipicephalus, were screened for CLEs, followed by characterization of CLEs by multi-locus sequence typing of the five Coxiella housekeeping genes (dnaK, groEL, rpoB, 16S rRNA, and 23S rRNA). The results showed that 45.7% (n = 80) were positive for CLEs. The comparison of the tick 16S rDNA phylogenetic tree with that of the CLEs concatenated sequences showed that there was a strong correlation between the topology of the trees. The results suggest that most of the CLEs have evolved within tick species, supporting the vertical transmission phenomenon. However, the negative results for CLE in some ticks warrants further investigations of other endosymbionts that the ticks in Zambia may also harbor.
• Immunization Coverage and Antibody Retention against Rabies in Domestic Dogs in Lusaka District, Zambia.

  Chiho Kaneko, Michihito Sasaki, Ryosuke Omori, Ryo Nakao, Chikako Kataoka-Nakamura, Ladslav Moonga, Joseph Ndebe, Walter Muleya, Edgar Simulundu, Bernard M Hang'ombe, George Dautu, Masahiro Kajihara, Akina Mori-Kajihara, Yongjin Qiu, Naoto Ito, Herman M Chambaro, Chihiro Sugimoto, Hideaki Higashi, Ayato Takada, Hirofumi Sawa, Aaron S Mweene, Norikazu Isoda

  Pathogens (Basel, Switzerland) 10 (6) 2021/06/11 [Refereed]
   

  Rabies remains endemic in Zambia. Despite conducting canine vaccinations in Lusaka district, the vaccination coverage and actual seropositivity in the dog population in Lusaka district are rarely evaluated. This study estimated the seropositivity-based immunization coverage in the owned dog population in Lusaka district using the expanded program on immunization cluster survey method. The time-series trend of neutralizing antibodies against rabies in vaccinated dogs was also evaluated. Of 366 dogs in 200 dog-owning households in Lusaka district, blood samples were collected successfully from 251 dogs. In the sampled dogs, 42.2% (106/251) had an antibody titer ≥0.5 IU/mL. When the 115 dogs whose blood was not collected were assumed to be seronegative, the minimum immunization coverage in Lusaka district's owned dog population was estimated at 29.0% (95% confidence interval: 22.4-35.5). It was also found that a single vaccination with certified vaccines is capable of inducing protective levels of antibodies. In contrast, higher antibody titers were observed in multiple-vaccinated dogs than in single-vaccinated dogs, coupled with the observation of a decline in antibody titer over time. These results suggest the importance of continuous booster immunization to maintain herd immunity and provide useful information to plan mass vaccination against rabies in Zambia.
• Serologic and molecular evidence for circulation of Crimean-Congo hemorrhagic fever virus in ticks and cattle in Zambia.

  Masahiro Kajihara, Martin Simuunza, Ngonda Saasa, George Dautu, Akina Mori-Kajihara, Yongjin Qiu, Ryo Nakao, Yoshiki Eto, Hayato Furumoto, Bernard M Hang'ombe, Yasuko Orba, Hirofumi Sawa, Edgar Simulundu, Shuetsu Fukushi, Shigeru Morikawa, Masayuki Saijo, Jiro Arikawa, Swithine Kabilika, Mwaka Monze, Victor Mukonka, Aaron Mweene, Ayato Takada, Kumiko Yoshimatsu

  PLoS neglected tropical diseases 15 (6) e0009452  2021/06 [Refereed]
   

  Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne zoonosis with a high case fatality rate in humans. Although the disease is widely found in Africa, Europe, and Asia, the distribution and genetic diversity of CCHF virus (CCHFV) are poorly understood in African countries. To assess the risks of CCHF in Zambia, where CCHF has never been reported, epidemiologic studies in cattle and ticks were conducted. Through an indirect immunofluorescence assay, CCHFV nucleoprotein-specific serum IgG was detected in 8.4% (88/1,047) of cattle. Among 290 Hyalomma ticks, the principal vector of CCHFV, the viral genome was detected in 11 ticks. Phylogenetic analyses of the CCHFV S and M genome segments revealed that one of the detected viruses was a genetic reassortant between African and Asian strains. This study provides compelling evidence for the presence of CCHFV in Zambia and its transmission to vertebrate hosts.
• Structural Insights into the Interaction of Filovirus Glycoproteins with the Endosomal Receptor Niemann-Pick C1: A Computational Study.

  Manabu Igarashi, Takatsugu Hirokawa, Yoshihiro Takadate, Ayato Takada

  Viruses 13 (5) 2021/05/14 [Refereed]
   

  Filoviruses, including marburgviruses and ebolaviruses, have a single transmembrane glycoprotein (GP) that facilitates their entry into cells. During entry, GP needs to be cleaved by host proteases to expose the receptor-binding site that binds to the endosomal receptor Niemann-Pick C1 (NPC1) protein. The crystal structure analysis of the cleaved GP (GPcl) of Ebola virus (EBOV) in complex with human NPC1 has demonstrated that NPC1 has two protruding loops (loops 1 and 2), which engage a hydrophobic pocket on the head of EBOV GPcl. However, the molecular interactions between NPC1 and the GPcl of other filoviruses remain unexplored. In the present study, we performed molecular modeling and molecular dynamics simulations of NPC1 complexed with GPcls of two ebolaviruses, EBOV and Sudan virus (SUDV), and one marburgvirus, Ravn virus (RAVV). Similar binding structures were observed in the GPcl-NPC1 complexes of EBOV and SUDV, which differed from that of RAVV. Specifically, in the RAVV GPcl-NPC1 complex, the tip of loop 2 was closer to the pocket edge comprising residues at positions 79-88 of GPcl; the root of loop 1 was predicted to interact with P116 and Q144 of GPcl. Furthermore, in the SUDV GPcl-NPC1 complex, the tip of loop 2 was slightly closer to the residue at position 141 than those in the EBOV and RAVV GPcl-NPC1 complexes. These structural differences may affect the size and/or shape of the receptor-binding pocket of GPcl. Our structural models could provide useful information for improving our understanding the differences in host preference among filoviruses as well as contributing to structure-based drug design.
• A Fc engineering approach to define functional humoral correlates of immunity against Ebola virus.

  Bronwyn M Gunn, Richard Lu, Matthew D Slein, Philipp A Ilinykh, Kai Huang, Caroline Atyeo, Sharon L Schendel, Jiyoung Kim, Caitlin Cain, Vicky Roy, Todd J Suscovich, Ayato Takada, Peter J Halfmann, Yoshihiro Kawaoka, Matthias G Pauthner, Mambu Momoh, Augustine Goba, Lansana Kanneh, Kristian G Andersen, John S Schieffelin, Donald Grant, Robert F Garry, Erica Ollmann Saphire, Alexander Bukreyev, Galit Alter

  Immunity 54 (4) 815 - 828 2021/04/13 [Refereed]
   

  Protective Ebola virus (EBOV) antibodies have neutralizing activity and induction of antibody constant domain (Fc)-mediated innate immune effector functions. Efforts to enhance Fc effector functionality often focus on maximizing antibody-dependent cellular cytotoxicity, yet distinct combinations of functions could be critical for antibody-mediated protection. As neutralizing antibodies have been cloned from EBOV disease survivors, we sought to identify survivor Fc effector profiles to help guide Fc optimization strategies. Survivors developed a range of functional antibody responses, and we therefore applied a rapid, high-throughput Fc engineering platform to define the most protective profiles. We generated a library of Fc variants with identical antigen-binding fragments (Fabs) from an EBOV neutralizing antibody. Fc variants with antibody-mediated complement deposition and moderate natural killer (NK) cell activity demonstrated complete protective activity in a stringent in vivo mouse model. Our findings highlight the importance of specific effector functions in antibody-mediated protection, and the experimental platform presents a generalizable resource for identifying correlates of immunity to guide therapeutic antibody design.
• Effect of varying storage conditions on diagnostic test outcomes of SARS-CoV-2.

  Satoshi Oguri, Shinichi Fujisawa, Keisuke Kamada, Sho Nakakubo, Yu Yamashita, Junichi Nakamura, Hiroshi Horii, Kazuki Sato, Mutsumi Nishida, Takanori Teshima, Yoichi Ohiro, Ayato Takada, Satoshi Konno

  The Journal of infection 83 (1) 119 - 145 2021/04/03 [Refereed]
  
• Domestic dog demographics and estimates of canine vaccination coverage in a rural area of Zambia for the elimination of rabies.

  Chiho Kaneko, Ryosuke Omori, Michihito Sasaki, Chikako Kataoka-Nakamura, Edgar Simulundu, Walter Muleya, Ladslav Moonga, Joseph Ndebe, Bernard M Hang'ombe, George Dautu, Yongjin Qiu, Ryo Nakao, Masahiro Kajihara, Akina Mori-Kajihara, Herman M Chambaro, Hideaki Higashi, Chihiro Sugimoto, Hirofumi Sawa, Aaron S Mweene, Ayato Takada, Norikazu Isoda

  PLoS neglected tropical diseases 15 (4) e0009222  2021/04 [Refereed]
   

  BACKGROUND: An estimated 75% or more of the human rabies cases in Africa occur in rural settings, which underscores the importance of rabies control in these areas. Understanding dog demographics can help design strategies for rabies control and plan and conduct canine mass vaccination campaigns effectively in African countries. METHODOLOGY/PRINCIPAL FINDINGS: A cross-sectional survey was conducted to investigate domestic dog demographics in Kalambabakali, in the rural Mazabuka District of Zambia. The population of ownerless dogs and the total achievable vaccination coverage among the total dog population was estimated using the capture-recapture-based Bayesian model by conducting a canine mass vaccination campaign. This study revealed that 29% of the domestic dog population was under one year old, and 57.7% of those were under three months old and thus were not eligible for the canine rabies vaccination in Zambia. The population growth was estimated at 15% per annum based on the cross-sectional household survey. The population of ownerless dogs was estimated to be small, with an ownerless-to-owned-dog ratio of 0.01-0.06 in the target zones. The achieved overall vaccination coverage from the first mass vaccination was estimated 19.8-51.6%. This low coverage was principally attributed to the owners' lack of information, unavailability, and dog-handling difficulties. The follow-up mass vaccination campaign achieved an overall coverage of 54.8-76.2%. CONCLUSIONS/SIGNIFICANCE: This paper indicates the potential for controlling canine rabies through mass vaccination in rural Zambia. Rabies education and responsible dog ownership are required to achieve high and sustainable vaccination coverage. Our findings also propose including puppies below three months old in the target population for rabies vaccination and emphasize that securing an annual enforcement of canine mass vaccination that reaches 70% coverage in the dog population is necessary to maintain protective herd immunity.
• Prevalence and genetic diversity of Shibuyunji virus, a novel tick-borne phlebovirus identified in Zambia.

  Edgar Simulundu, Saidon Mbambara, Herman M Chambaro, Karen Sichibalo, Masahiro Kajihara, King S Nalubamba, Hirofumi Sawa, Ayato Takada, Katendi Changula, Simbarashe Chitanga

  Archives of virology 166 (3) 915 - 919 2021/03 [Refereed]
   

  Tick-borne pathogens are an emerging public health threat worldwide. However, information on tick-borne viruses is scanty in sub-Saharan Africa. Here, by RT-PCR, 363 ticks (Amblyomma, Hyalomma and Rhipicephalus) in the Namwala and Livingstone districts of Zambia were screened for tick-borne phleboviruses (TBPVs). TBPVs (L gene) were detected in 19 (5.2%) Rhipicephalus ticks in Namwala. All the detected TBPVs were Shibuyunji viruses. Phylogenetically, they were closely related to American dog tick phlebovirus. This study highlights the possible role of Rhipicephalus ticks as the main host of Shibuyunji virus and suggests that these viruses may be present outside the area where they were initially discovered.
• Variation in the HA antigenicity of A(H1N1)pdm09-related swine influenza viruses.

  Ahmed Magdy Khalil, Reiko Yoshida, Tatsunori Masatani, Ayato Takada, Makoto Ozawa

  The Journal of general virology 102 (3) 2021/03 [Refereed]
   

  Since the influenza pandemic in 2009, the causative agent 'A(H1N1)pdm09 virus', has been circulating in both human and swine populations. Although phylogenetic analyses of the haemagglutinin (HA) gene segment have revealed broader genetic diversity of A(H1N1)pdm09-related swine influenza A viruses (swIAVs) compared with human A(H1N1)pdm09 viruses, it remains unclear whether the genetic diversity reflects the antigenic differences in HA. To assess the impact of the diversity of the HA gene of A(H1N1)pdm09-related swIAVs on HA antigenicity, we characterized 12 swIAVs isolated in Japan from 2013 to 2018. We used a ferret antiserum and a panel of anti-HA mouse monoclonal antibodies (mAbs) raised against an early A(H1N1)pdm09 isolate. The neutralization assay with the ferret antiserum revealed that five of the 12 swIAVs were significantly different in their HA antigenicity from the early A(H1N1)pdm09 isolate. The mAbs also showed differential neutralization patterns depending on the swIAV strains. In addition, the single amino acid substitution at position 190 of HA, which was found in one of the five antigenically different swIAVs but not in human isolates, was shown to be one of the critical determinants for the antigenic difference of swIAV HAs. Two potential N-glycosylation sites at amino acid positions 185 and 276 of the HA molecule were identified in two antigenically different swIAVs. These results indicated that the genetic diversity of HA in the A(H1N1)pdm09-related swIAVs is associated with their HA antigenic variation. Our findings highlighted the need for surveillance to monitor the emergence of swIAV antigenic variants with public health importance.
• An African tick flavivirus forming an independent clade exhibits unique exoribonuclease-resistant RNA structures in the genomic 3'-untranslated region.

  Hayato Harima, Yasuko Orba, Shiho Torii, Yongjin Qiu, Masahiro Kajihara, Yoshiki Eto, Naoya Matsuta, Bernard M Hang'ombe, Yuki Eshita, Kentaro Uemura, Keita Matsuno, Michihito Sasaki, Kentaro Yoshii, Ryo Nakao, William W Hall, Ayato Takada, Takashi Abe, Michael T Wolfinger, Martin Simuunza, Hirofumi Sawa

  Scientific reports 11 (1) 4883 - 4883 2021/03/01 [Refereed]
   

  Tick-borne flaviviruses (TBFVs) infect mammalian hosts through tick bites and can cause various serious illnesses, such as encephalitis and hemorrhagic fevers, both in humans and animals. Despite their importance to public health, there is limited epidemiological information on TBFV infection in Africa. Herein, we report that a novel flavivirus, Mpulungu flavivirus (MPFV), was discovered in a Rhipicephalus muhsamae tick in Zambia. MPFV was found to be genetically related to Ngoye virus detected in ticks in Senegal, and these viruses formed a unique lineage in the genus Flavivirus. Analyses of dinucleotide contents of flaviviruses indicated that MPFV was similar to those of other TBFVs with a typical vertebrate genome signature, suggesting that MPFV may infect vertebrate hosts. Bioinformatic analyses of the secondary structures in the 3'-untranslated regions (UTRs) revealed that MPFV exhibited unique exoribonuclease-resistant RNA (xrRNA) structures. Utilizing biochemical approaches, we clarified that two xrRNA structures of MPFV in the 3'-UTR could prevent exoribonuclease activity. In summary, our findings provide new information regarding the geographical distribution of TBFV and xrRNA structures in the 3'-UTR of flaviviruses.
• Tim4 recognizes carbon nanotubes and mediates phagocytosis leading to granuloma formation.

  Satoshi Omori, Misato Tsugita, Yasuto Hoshikawa, Masanobu Morita, Fumiya Ito, Shin-Ichiro Yamaguchi, Qilin Xie, Osamu Noyori, Tomoya Yamaguchi, Ayato Takada, Tatsuya Saitoh, Shinya Toyokuni, Hisaya Akiba, Shigekazu Nagata, Kengo Kinoshita, Masafumi Nakayama

  Cell reports 34 (6) 108734 - 108734 2021/02/09 [Refereed]
   

  Macrophage recognition and phagocytosis of crystals is critical for the associated fibrosis and cancer. Of note, multi-walled carbon nanotubes (MWCNTs), the highly representative products of nanotechnology, induce macrophage NLRP3 inflammasome activation and cause asbestosis-like pathogenesis. However, it remains largely unknown how macrophages efficiently recognize MWCNTs on their cell surfaces. Here, we identify by a targeted screening of phagocyte receptors the phosphatidylserine receptors T cell immunoglobulin mucin 4 (Tim4) and Tim1 as the pattern-recognition receptors for carbon crystals. Docking simulation studies reveal spatiotemporally stable interfaces between aromatic residues in the extracellular IgV domain of Tim4 and one-dimensional carbon crystals. Further, CRISPR-Cas9-mediated deletion of Tim4 and Tim1 reveals that Tim4, but not Tim1, critically contributes to the recognition of MWCNTs by peritoneal macrophages and to granuloma development in a mouse model of direct mesothelium exposure to MWCNTs. These results suggest that Tim4 recognizes MWCNTs through aromatic interactions and mediates phagocytosis leading to granulomas.
• Purification of Crimean-Congo hemorrhagic fever virus nucleoprotein and its utility for serological diagnosis.

  Boniface Pongombo Lombe, Hiroko Miyamoto, Takeshi Saito, Reiko Yoshida, Rashid Manzoor, Masahiro Kajihara, Masayuki Shimojima, Shuetsu Fukushi, Shigeru Morikawa, Tomoki Yoshikawa, Takeshi Kurosu, Masayuki Saijo, Qing Tang, Justin Masumu, David Hawman, Heinz Feldmann, Ayato Takada

  Scientific reports 11 (1) 2324 - 2324 2021/01/27 [Refereed]
   

  Crimean-Congo hemorrhagic fever virus (CCHFV) causes a zoonotic disease, Crimean-Congo hemorrhagic fever (CCHF) endemic in Africa, Asia, the Middle East, and Southeastern Europe. However, the prevalence of CCHF is not monitored in most of the endemic countries due to limited availability of diagnostic assays and biosafety regulations required for handling infectious CCHFV. In this study, we established a protocol to purify the recombinant CCHFV nucleoprotein (NP), which is antigenically highly conserved among multiple lineages/clades of CCHFVs and investigated its utility in an enzyme-linked immunosorbent assay (ELISA) to detect CCHFV-specific antibodies. The NP gene was cloned into the pCAGGS mammalian expression plasmid and human embryonic kidney 293 T cells were transfected with the plasmid. The expressed NP molecule was purified from the cell lysate using cesium-chloride gradient centrifugation. Purified NP was used as the antigen for the ELISA to detect anti-CCHFV IgG. Using the CCHFV NP-based ELISA, we efficiently detected CCHFV-specific IgG in anti-NP rabbit antiserum and CCHFV-infected monkey serum. When compared to the commercially available Blackbox CCHFV IgG ELISA kit, our assay showed equivalent performance in detecting CCHFV-specific IgG in human sera. These results demonstrate the usefulness of our CCHFV NP-based ELISA for seroepidemiological studies.
• First COVID-19 case in Zambia - Comparative phylogenomic analyses of SARS-CoV-2 detected in African countries.

  Edgar Simulundu, Francis Mupeta, Pascalina Chanda-Kapata, Ngonda Saasa, Katendi Changula, Walter Muleya, Simbarashe Chitanga, Miniva Mwanza, Paul Simusika, Herman Chambaro, Benjamin Mubemba, Masahiro Kajihara, Duncan Chanda, Lloyd Mulenga, Sombo Fwoloshi, Aaron Lunda Shibemba, Fred Kapaya, Paul Zulu, Kunda Musonda, Mwaka Monze, Nyambe Sinyange, Mazyanga L Mazaba, Muzala Kapin'a, Peter J Chipimo, Raymond Hamoonga, Davie Simwaba, William Ngosa, Albertina N Morales, Nkomba Kayeyi, John Tembo, Mathew Bates, Yasuko Orba, Hirofumi Sawa, Ayato Takada, King S Nalubamba, Kennedy Malama, Victor Mukonka, Alimuddin Zumla, Nathan Kapata

  International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases 102 455 - 459 2021/01 [Refereed]
   

  Since its first discovery in December 2019 in Wuhan, China, COVID-19, caused by the novel coronavirus SARS-CoV-2, has spread rapidly worldwide. While African countries were relatively spared initially, the initial low incidence of COVID-19 cases was not sustained for long due to continuing travel links between China, Europe and Africa. In preparation, Zambia had applied a multisectoral national epidemic disease surveillance and response system resulting in the identification of the first case within 48 h of the individual entering the country by air travel from a trip to France. Contact tracing showed that SARS-CoV-2 infection was contained within the patient's household, with no further spread to attending health care workers or community members. Phylogenomic analysis of the patient's SARS-CoV-2 strain showed that it belonged to lineage B.1.1., sharing the last common ancestor with SARS-CoV-2 strains recovered from South Africa. At the African continental level, our analysis showed that B.1 and B.1.1 lineages appear to be predominant in Africa. Whole genome sequence analysis should be part of all surveillance and case detection activities in order to monitor the origin and evolution of SARS-CoV-2 lineages across Africa.
• A Novel Mechanism Underlying Antiviral Activity of an Influenza Virus M2-Specific Antibody.

  Rashid Manzoor, Nao Eguchi, Reiko Yoshida, Hiroichi Ozaki, Tatsunari Kondoh, Kosuke Okuya, Hiroko Miyamoto, Ayato Takada

  Journal of virology 95 (1) 2020/12/09 [Refereed]
   

  Protective immunity against influenza A viruses (IAVs) generally depends on antibodies to the major envelope glycoprotein, hemagglutinin (HA), whose antigenicity is distinctive among IAV subtypes. On the other hand, the matrix 2 (M2) protein is antigenically highly conserved and has been studied as an attractive vaccine antigen to confer cross-protective immunity against multiple subtypes of IAVs. However, antiviral mechanisms of M2-specific antibodies are not fully understood. Here, we report the molecular basis of antiviral activity of an M2-specific monoclonal antibody (MAb), rM2ss23. We first found that rM2ss23 inhibited A/Aichi/2/1968 (H3N2) (Aichi) but not A/PR/8/1934 (H1N1) (PR8) replication. rM2ss23 altered the cell surface distribution of M2, likely by cross-linking the molecules, and interfered with the colocalization of HA and M2, resulting in reduced budding of progeny viruses. However, these effects were not observed for another strain, PR8, despite the binding capacity of rM2ss23 to PR8 M2. Interestingly, HA was also involved in the resistance of PR8 to rM2ss23. We also found that two amino acid residues at positions 54 and 57 in the M2 cytoplasmic tail were critical for the insensitivity of PR8 to rM2ss2. These findings suggest that the disruption of the M2-HA colocalization on infected cells and subsequent reduction of virus budding is one of the principal mechanisms of antiviral activity of M2-specific antibodies and that anti-M2 antibody-sensitive and -resistant IAVs have different properties in the interaction between M2 and HA.IMPORTANCE Although the IAV HA is the major target of neutralizing antibodies, most of the antibodies are HA subtype specific, restricting the potential of HA-based vaccines. On the contrary, the IAV M2 protein has been studied as a vaccine antigen to confer cross-protective immunity against IAVs with multiple HA subtypes, since M2 is antigenically conserved. Although a number of studies highlight the protective role of anti-HA neutralizing and nonneutralizing antibodies, precise information on the molecular mechanism of action of M2-specific antibodies is still obscure. In this study, we found that an anti-M2 antibody interfered with the HA-M2 association, which is important for efficient budding of progeny virus particles from infected cells. The antiviral activity was IAV strain dependent despite the similar binding capacity of the antibody to M2, and, interestingly, HA was involved in susceptibility to the antibody. Our data provide a novel mechanism underlying antiviral activity of M2-specific antibodies.
• 2020 taxonomic update for phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales.

  Jens H Kuhn, Scott Adkins, Daniela Alioto, Sergey V Alkhovsky, Gaya K Amarasinghe, Simon J Anthony, Tatjana Avšič-Županc, María A Ayllón, Justin Bahl, Anne Balkema-Buschmann, Matthew J Ballinger, Tomáš Bartonička, Christopher Basler, Sina Bavari, Martin Beer, Dennis A Bente, Éric Bergeron, Brian H Bird, Carol Blair, Kim R Blasdell, Steven B Bradfute, Rachel Breyta, Thomas Briese, Paul A Brown, Ursula J Buchholz, Michael J Buchmeier, Alexander Bukreyev, Felicity Burt, Nihal Buzkan, Charles H Calisher, Mengji Cao, Inmaculada Casas, John Chamberlain, Kartik Chandran, Rémi N Charrel, Biao Chen, Michela Chiumenti, Il-Ryong Choi, J Christopher S Clegg, Ian Crozier, John V da Graça, Elena Dal Bó, Alberto M R Dávila, Juan Carlos de la Torre, Xavier de Lamballerie, Rik L de Swart, Patrick L Di Bello, Nicholas Di Paola, Francesco Di Serio, Ralf G Dietzgen, Michele Digiaro, Valerian V Dolja, Olga Dolnik, Michael A Drebot, Jan Felix Drexler, Ralf Dürrwald, Lucie Dufkova, William G Dundon, W Paul Duprex, John M Dye, Andrew J Easton, Hideki Ebihara, Toufic Elbeaino, Koray Ergünay, Jorlan Fernandes, Anthony R Fooks, Pierre B H Formenty, Leonie F Forth, Ron A M Fouchier, Juliana Freitas-Astúa, Selma Gago-Zachert, George Fú Gāo, María Laura García, Adolfo García-Sastre, Aura R Garrison, Aiah Gbakima, Tracey Goldstein, Jean-Paul J Gonzalez, Anthony Griffiths, Martin H Groschup, Stephan Günther, Alexandro Guterres, Roy A Hall, John Hammond, Mohamed Hassan, Jussi Hepojoki, Satu Hepojoki, Udo Hetzel, Roger Hewson, Bernd Hoffmann, Seiji Hongo, Dirk Höper, Masayuki Horie, Holly R Hughes, Timothy H Hyndman, Amara Jambai, Rodrigo Jardim, Dàohóng Jiāng, Qi Jin, Gilda B Jonson, Sandra Junglen, Serpil Karadağ, Karen E Keller, Boris Klempa, Jonas Klingström, Gary Kobinger, Hideki Kondō, Eugene V Koonin, Mart Krupovic, Gael Kurath, Ivan V Kuzmin, Lies Laenen, Robert A Lamb, Amy J Lambert, Stanley L Langevin, Benhur Lee, Elba R S Lemos, Eric M Leroy, Dexin Li, Jiànróng Lǐ, Mifang Liang, Wénwén Liú, Yàn Liú, Igor S Lukashevich, Piet Maes, William Marciel de Souza, Marco Marklewitz, Sergio H Marshall, Giovanni P Martelli, Robert R Martin, Shin-Yi L Marzano, Sébastien Massart, John W McCauley, Nicole Mielke-Ehret, Angelantonio Minafra, Maria Minutolo, Ali Mirazimi, Hans-Peter Mühlbach, Elke Mühlberger, Rayapati Naidu, Tomohide Natsuaki, Beatriz Navarro, José A Navarro, Sergey V Netesov, Gabriele Neumann, Norbert Nowotny, Márcio R T Nunes, Are Nylund, Arnfinn L Økland, Renata C Oliveira, Gustavo Palacios, Vicente Pallas, Bernadett Pályi, Anna Papa, Colin R Parrish, Alex Pauvolid-Corrêa, Janusz T Pawęska, Susan Payne, Daniel R Pérez, Florian Pfaff, Sheli R Radoshitzky, Aziz-Ul Rahman, Pedro L Ramos-González, Renato O Resende, Carina A Reyes, Bertus K Rima, Víctor Romanowski, Gabriel Robles Luna, Paul Rota, Dennis Rubbenstroth, Jonathan A Runstadler, Daniel Ruzek, Sead Sabanadzovic, Jiří Salát, Amadou Alpha Sall, Maria S Salvato, Kamil Sarpkaya, Takahide Sasaya, Martin Schwemmle, Muhammad Z Shabbir, Xiǎohóng Shí, Zhènglì Shí, Yukio Shirako, Peter Simmonds, Jana Širmarová, Manuela Sironi, Sophie Smither, Teemu Smura, Jin-Won Song, Kirsten M Spann, Jessica R Spengler, Mark D Stenglein, David M Stone, Petra Straková, Ayato Takada, Robert B Tesh, Natalie J Thornburg, Keizō Tomonaga, Noël Tordo, Jonathan S Towner, Massimo Turina, Ioannis Tzanetakis, Rainer G Ulrich, Anna Maria Vaira, Bernadette van den Hoogen, Arvind Varsani, Nikos Vasilakis, Martin Verbeek, Victoria Wahl, Peter J Walker, Hui Wang, Jianwei Wang, Xifeng Wang, Lin-Fa Wang, Tàiyún Wèi, Heather Wells, Anna E Whitfield, John V Williams, Yuri I Wolf, Zhìqiáng Wú, Xin Yang, Xīnglóu Yáng, Xuejie Yu, Natalya Yutin, F Murilo Zerbini, Tong Zhang, Yong-Zhen Zhang, Guohui Zhou, Xueping Zhou

  Archives of virology 165 (12) 3023 - 3072 2020/12 [Refereed]
   

  In March 2020, following the annual International Committee on Taxonomy of Viruses (ICTV) ratification vote on newly proposed taxa, the phylum Negarnaviricota was amended and emended. At the genus rank, 20 new genera were added, two were deleted, one was moved, and three were renamed. At the species rank, 160 species were added, four were deleted, ten were moved and renamed, and 30 species were renamed. This article presents the updated taxonomy of Negarnaviricota as now accepted by the ICTV.
• A biaryl sulfonamide derivative as a novel inhibitor of filovirus infection.

  Mao Isono, Wakako Furuyama, Makoto Kuroda, Tatsunari Kondoh, Manabu Igarashi, Masahiro Kajihara, Reiko Yoshida, Rashid Manzoor, Kosuke Okuya, Hiroko Miyamoto, Heinz Feldmann, Andrea Marzi, Masahiro Sakaitani, Asuka Nanbo, Ayato Takada

  Antiviral research 183 104932 - 104932 2020/11 [Refereed]
   

  Ebolaviruses and marburgviruses, members of the family Filoviridae, are known to cause fatal diseases often associated with hemorrhagic fever. Recent outbreaks of Ebola virus disease in West African countries and the Democratic Republic of the Congo have made clear the urgent need for the development of therapeutics and vaccines against filoviruses. Using replication-incompetent vesicular stomatitis virus (VSV) pseudotyped with the Ebola virus (EBOV) envelope glycoprotein (GP), we screened a chemical compound library to obtain new drug candidates that inhibit filoviral entry into target cells. We discovered a biaryl sulfonamide derivative that suppressed in vitro infection mediated by GPs derived from all known human-pathogenic filoviruses. To determine the inhibitory mechanism of the compound, we monitored each entry step (attachment, internalization, and membrane fusion) using lipophilic tracer-labeled ebolavirus-like particles and found that the compound efficiently blocked fusion between the viral envelope and the endosomal membrane during cellular entry. However, the compound did not block the interaction of GP with the Niemann-Pick C1 protein, which is believed to be the receptor of filoviruses. Using replication-competent VSVs pseudotyped with EBOV GP, we selected escape mutants and identified two EBOV GP amino acid residues (positions 47 and 66) important for the interaction with this compound. Interestingly, these amino acid residues were located at the base region of the GP trimer, suggesting that the compound might interfere with the GP conformational change required for membrane fusion. These results suggest that this biaryl sulfonamide derivative is a novel fusion inhibitor and a possible drug candidate for the development of a pan-filovirus therapeutic.
• Evidence for exposure of asymptomatic domestic pigs to African swine fever virus during an inter-epidemic period in Zambia.

  Herman M Chambaro, Michihito Sasaki, Yona Sinkala, Gabriel Gonzalez, David Squarre, Paul Fandamu, Caesar Lubaba, Liywalii Mataa, Misheck Shawa, Kabemba E Mwape, Sarah Gabriël, Mwelwa Chembensofu, Michael J Carr, William W Hall, Yongjin Qiu, Masahiro Kajihara, Ayato Takada, Yasuko Orba, Edgar Simulundu, Hirofumi Sawa

  Transboundary and emerging diseases 67 (6) 2741 - 2752 2020/11 [Refereed][Not invited]
   

  African swine fever (ASF) causes persistent outbreaks in endemic and non-endemic regions in Zambia. However, the epidemiology of the disease is poorly understood, particularly during the inter-epidemic periods. We conducted surveillance for ASF in asymptomatic domestic pigs and soft ticks in selected Zambian provinces. While serum samples (n = 1,134) were collected from crossbred pigs from all study sites between 2014 and 2017, whole blood (n = 300) was collected from both crossbred and indigenous pigs in Eastern Province (EP) in 2017. Soft ticks were collected from Mosi-oa-Tunya National Park in Southern Province (SP) in 2019. Sera were screened for antibodies against ASF by ELISA while genome detection in whole blood and soft ticks was conducted by PCR. Ticks were identified morphologically and by phylogenetic analysis of the 16S rRNA gene. Seroprevalence was highest in EP (50.9%, 95% CI [47.0-54.9]) compared to significantly lower rates in SP (2.9%, 95% CI [1.6-5.1]). No antibodies to ASFV were detected in Lusaka Province. In EP, the prevalence of ASFV genome was 11.7% (35/300), significantly higher (OR = 6.2, 95% CI [2.4-16.6]) in indigenous pigs compared to crossbred pigs. The pooled prevalence of ASFV genome in ticks was 11.0%, 95% CI [8.5-13.9]. Free-range husbandry system was the only factor that was significantly associated with seropositive (p < .0001, OR = 39.3) and PCR-positive results (p < .001, OR = 5.7). Phylogenetically, based on the p72 gene, ASFV from Ornithodoros moubata ticks detected in this study belonged to genotype I, but they separated into two distinct clusters. Besides confirming ASF endemicity in EP and the presence of ASFV-infected ticks in SP, these results provide evidence for exposure of domestic pigs to ASFV in non-endemic regions during the inter-epidemic period.
• Receptor-Mediated Host Cell Preference of a Bat-Derived Filovirus, Lloviu Virus.

  Yoshihiro Takadate, Rashid Manzoor, Takeshi Saito, Yurie Kida, Junki Maruyama, Tatsunari Kondoh, Hiroko Miyamoto, Hirohito Ogawa, Masahiro Kajihara, Manabu Igarashi, Ayato Takada

  Microorganisms 8 (10) 2020/10/05 [Refereed]
   

  Lloviu virus (LLOV), a bat-derived filovirus that is phylogenetically distinct from human pathogenic filoviruses such as Ebola virus (EBOV) and Marburg virus (MARV), was discovered in Europe. However, since infectious LLOV has never been isolated, the biological properties of this virus remain poorly understood. We found that vesicular stomatitis virus (VSV) pseudotyped with the glycoprotein (GP) of LLOV (VSV-LLOV) showed higher infectivity in one bat (Miniopterus sp.)-derived cell line than in the other bat-derived cell lines tested, which was distinct from the tropism of VSV pseudotyped with EBOV (VSV-EBOV) and MARV GPs. We then focused on the interaction between GP and Niemann-Pick C1 (NPC1) protein, one of the cellular receptors of filoviruses. We introduced the Miniopterus bat and human NPC1 genes into NPC1-knockout Vero E6 cells and their susceptibilities to the viruses were compared. The cell line expressing the bat NPC1 showed higher susceptibility to VSV-LLOV than that expressing human NPC1, whereas the opposite preference was seen for VSV-EBOV. Using a site-directed mutagenesis approach, amino acid residues involved in the differential tropism were identified in the NPC1 and GP molecules. Our results suggest that the interaction between GP and NPC1 is an important factor in the tropism of LLOV to a particular bat species.
• Characterization of mammalian orthoreoviruses isolated from faeces of pigs in Zambia.

  Hayato Harima, Michihito Sasaki, Masahiro Kajihara, Gabriel Gonzalez, Edgar Simulundu, Eugene C Bwalya, Yongjin Qiu, Kosuke Okuya, Mao Isono, Yasuko Orba, Ayato Takada, Bernard M Hang'ombe, Aaron S Mweene, Hirofumi Sawa

  The Journal of general virology 101 (10) 1027 - 1036 2020/10 [Refereed][Not invited]
   

  Mammalian orthoreovirus (MRV) has been identified in humans, livestock and wild animals; this wide host range allows individual MRV to transmit into multiple species. Although several interspecies transmission and genetic reassortment events of MRVs among humans, livestock and wildlife have been reported, the genetic diversity and geographic distribution of MRVs in Africa are poorly understood. In this study, we report the first isolation and characterization of MRVs circulating in a pig population in Zambia. In our screening, MRV genomes were detected in 19.7 % (29/147) of faecal samples collected from pigs by reverse transcription PCR. Three infectious MRV strains (MRV-85, MRV-96 and MRV-117) were successfully isolated, and their complete genomes were sequenced. Recombination analyses based on the complete genome sequences of the isolated MRVs demonstrated that MRV-96 shared the S3 segment with a different MRV isolated from bats, and that the L1 and M3 segments of MRV-117 originated from bat and human MRVs, respectively. Our results suggest that the isolated MRVs emerged through genetic reassortment events with interspecies transmission. Given the lack of information regarding MRVs in Africa, further surveillance of MRVs circulating among humans, domestic animals and wildlife is required to assess potential risk for humans and animals.
• Avian Influenza Viruses Detected in Birds in Sub-Saharan Africa: A Systematic Review.

  Annie Kalonda, Ngonda Saasa, Panji Nkhoma, Masahiro Kajihara, Hirofumi Sawa, Ayato Takada, Edgar Simulundu

  Viruses 12 (9) 2020/09/07 [Refereed]
   

  In the recent past, sub-Saharan Africa has not escaped the devastating effects of avian influenza virus (AIV) in poultry and wild birds. This systematic review describes the prevalence, spatiotemporal distribution, and virus subtypes detected in domestic and wild birds for the past two decades (2000-2019). We collected data from three electronic databases, PubMed, SpringerLink electronic journals and African Journals Online, using the Preferred Reporting Items for Systematic reviews and Meta-Analyses protocol. A total of 1656 articles were reviewed, from which 68 were selected. An overall prevalence of 3.0% AIV in birds was observed. The prevalence varied between regions and ranged from 1.1% to 7.1%. The Kruskal-Wallis and Wilcoxon signed-rank sum test showed no significant difference in the prevalence of AIV across regions, χ2(3) = 5.237, p = 0.1553 and seasons, T = 820, z = -1.244, p = 0.2136. Nineteen hemagglutinin/neuraminidase subtype combinations were detected during the reviewed period, with southern Africa recording more diverse AIV subtypes than other regions. The most detected subtype was H5N1, followed by H9N2, H5N2, H5N8 and H6N2. Whilst these predominant subtypes were mostly detected in domestic poultry, H1N6, H3N6, H4N6, H4N8, H9N1 and H11N9 were exclusively detected in wild birds. Meanwhile, H5N1, H5N2 and H5N8 were detected in both wild and domestic birds suggesting circulation of these subtypes among wild and domestic birds. Our findings provide critical information on the eco-epidemiology of AIVs that can be used to improve surveillance strategies for the prevention and control of avian influenza in sub-Saharan Africa.
• A complement component C1q-mediated mechanism of antibody-dependent enhancement of Ebola virus infection.

  Wakako Furuyama, Asuka Nanbo, Junki Maruyama, Andrea Marzi, Ayato Takada

  PLoS neglected tropical diseases 14 (9) e0008602  2020/09 

  Besides the common Fc receptor (FcR)-mediated mechanism of antibody-dependent enhancement (ADE), Ebola virus (EBOV) is known to utilize the complement component C1q for ADE of infection. This mechanism is FcR-independent and mediated by cross-linking of virus-antibody-C1q complexes to cell surface C1q receptors, leading to enhanced viral entry into cells. Using confocal microscopy, we found that virus-like particles (VLPs) consisting of EBOV glycoprotein, nucleoprotein, and matrix protein attached to the surface of human kidney 293 cells more efficiently in the presence of an ADE monoclonal antibody and C1q than with the antibody or C1q alone, and that there was no significant difference in the efficiency of VLP uptake into endosomes between the C1q-mediated ADE and non-ADE entry. Accordingly, both ADE and non-ADE infection were similarly decreased by inhibitors of the signaling pathways known to be required for endocytosis. These results suggest that C1q-mediated ADE of EBOV infection is simply caused by increased attachment of virus particles to the cell surface, which is distinct from the mechanism of FcR-mediated ADE requiring intracellular signaling to promote phagocytosis/macropinocytosis.
• Modeling the efficiency of filovirus entry into cells in vitro: Effects of SNP mutations in the receptor molecule.

  Kwang Su Kim, Tatsunari Kondoh, Yusuke Asai, Ayato Takada, Shingo Iwami

  PLoS computational biology 16 (9) e1007612  2020/09 [Refereed]
   

  Interaction between filovirus glycoprotein (GP) and the Niemann-Pick C1 (NPC1) protein is essential for membrane fusion during virus entry. Some single-nucleotide polymorphism (SNPs) in two surface-exposed loops of NPC1 are known to reduce viral infectivity. However, the dependence of differences in entry efficiency on SNPs remains unclear. Using vesicular stomatitis virus pseudotyped with Ebola and Marburg virus GPs, we investigated the cell-to-cell spread of viruses in cultured cells expressing NPC1 or SNP derivatives. Eclipse and virus-producing phases were assessed by in vitro infection experiments, and we developed a mathematical model describing spatial-temporal virus spread. This mathematical model fit the plaque radius data well from day 2 to day 6. Based on the estimated parameters, we found that SNPs causing the P424A and D508N substitutions in NPC1 most effectively reduced the entry efficiency of Ebola and Marburg viruses, respectively. Our novel approach could be broadly applied to other virus plaque assays.
• A Surrogate Animal Model for Screening of Ebola and Marburg Glycoprotein-Targeting Drugs Using Pseudotyped Vesicular Stomatitis Viruses.

  Takeshi Saito, Junki Maruyama, Noriyo Nagata, Mao Isono, Kosuke Okuya, Yoshihiro Takadate, Yurie Kida, Hiroko Miyamoto, Akina Mori-Kajihara, Takanari Hattori, Wakako Furuyama, Shinya Ogawa, Shigeru Iida, Ayato Takada

  Viruses 12 (9) 2020/08/22 [Refereed]
   

  Filoviruses, including Ebola virus (EBOV) and Marburg virus (MARV), cause severe hemorrhagic fever in humans and nonhuman primates with high mortality rates. There is no approved therapy against these deadly viruses. Antiviral drug development has been hampered by the requirement of a biosafety level (BSL)-4 facility to handle infectious EBOV and MARV because of their high pathogenicity to humans. In this study, we aimed to establish a surrogate animal model that can be used for anti-EBOV and -MARV drug screening under BSL-2 conditions by focusing on the replication-competent recombinant vesicular stomatitis virus (rVSV) pseudotyped with the envelope glycoprotein (GP) of EBOV (rVSV/EBOV) and MARV (rVSV/MARV), which has been investigated as vaccine candidates and thus widely used in BSL-2 laboratories. We first inoculated mice, rats, and hamsters intraperitoneally with rVSV/EBOV and found that only hamsters showed disease signs and succumbed within 4 days post-infection. Infection with rVSV/MARV also caused lethal infection in hamsters. Both rVSV/EBOV and rVSV/MARV were detected at high titers in multiple organs including the liver, spleen, kidney, and lungs of infected hamsters, indicating acute and systemic infection resulting in fatal outcomes. Therapeutic effects of passive immunization with an anti-EBOV neutralizing antibody were specifically observed in rVSV/EBOV-infected hamsters. Thus, this animal model is expected to be a useful tool to facilitate in vivo screening of anti-filovirus drugs targeting the GP molecule.
• Genetic and antigenic characterization of H5 and H7 avian influenza viruses isolated from migratory waterfowl in Mongolia from 2017 to 2019.

  Ankhanbaatar Ulaankhuu, Enkhbold Bazarragchaa, Masatoshi Okamatsu, Takahiro Hiono, Khishgee Bodisaikhan, Tsolmon Amartuvshin, Jargalsaikhan Tserenjav, Tsogtbaatar Urangoo, Khanui Buyantogtokh, Keita Matsuno, Takanari Hattori, Tatsunari Kondoh, Masahiro Sato, Yoshihiro Takadate, Shiho Torii, Mao Isono, Kosuke Okuya, Takeshi Saito, Nodoka Kasajima, Yurie Kida, Junki Maruyama, Manabu Igarashi, Ayato Takada, Hiroshi Kida, Damdinjav Batchuluun, Yoshihiro Sakoda

  Virus genes 56 (4) 472 - 479 2020/08 [Refereed][Not invited]
   

  The circulation of highly pathogenic avian influenza viruses (HPAIVs) of various subtypes (e.g., H5N1, H5N6, H5N8, and H7N9) in poultry remains a global concern for animal and public health. Migratory waterfowls play important roles in the transmission of these viruses across countries. To monitor virus spread by wild birds, active surveillance for avian influenza in migratory waterfowl was conducted in Mongolia from 2015 to 2019. In total, 5000 fecal samples were collected from lakesides in central Mongolia, and 167 influenza A viruses were isolated. Two H5N3, four H7N3, and two H7N7 viruses were characterized in this study. The amino acid sequence at hemagglutinin (HA) cleavage site of those isolates suggested low pathogenicity in chickens. Phylogenetic analysis revealed that all H5 and H7 viruses were closely related to recent H5 and H7 low pathogenic avian influenza viruses (LPAIVs) isolated from wild birds in Asia and Europe. Antigenicity of H7Nx was similar to those of typical non-pathogenic avian influenza viruses (AIVs). While HPAIVs or A/Anhui/1/2013 (H7N9)-related LPAIVs were not detected in migratory waterfowl in Mongolia, sporadic introductions of AIVs including H5 and H7 viruses into Mongolia through the wild bird migration were identified. Thus, continued monitoring of H5 and H7 AIVs in both domestic and wild birds is needed for the early detection of HPAIVs spread into the country.
• Comparative Analyses of the Antiviral Activities of IgG and IgA Antibodies to Influenza A Virus M2 Protein.

  Kosuke Okuya, Nao Eguchi, Rashid Manzoor, Reiko Yoshida, Shinji Saito, Tadaki Suzuki, Michihito Sasaki, Takeshi Saito, Yurie Kida, Akina Mori-Kajihara, Hiroko Miyamoto, Osamu Ichii, Masahiro Kajihara, Hideaki Higashi, Ayato Takada

  Viruses 12 (7) 2020/07/20 [Refereed][Not invited]
   

  The influenza A virus (IAV) matrix-2 (M2) protein is an antigenically conserved viral envelope protein that plays an important role in virus budding together with another envelope protein, hemagglutinin (HA). An M2-specific mouse monoclonal IgG antibody, rM2ss23, which binds to the ectodomain of the M2 protein, has been shown to be a non-neutralizing antibody, but inhibits plaque formation of IAV strains. In this study, we generated chimeric rM2ss23 (ch-rM2ss23) IgG and IgA antibodies with the same variable region and compared their antiviral activities. Using gel chromatography, ch-rM2ss23 IgA were divided into three antibody subsets: monomeric IgA (m-IgA), dimeric IgA (d-IgA), and trimeric and tetrameric IgA (t/q-IgA). We found that t/q-IgA had a significantly higher capacity to reduce the plaque size of IAVs than IgG and m-IgA, most likely due to the decreased number of progeny virus particles produced from infected cells. Interestingly, HA-M2 colocalization was remarkably reduced on the infected cell surface in the presence of ch-rM2ss23 antibodies. These results indicate that anti-M2 polymeric IgA restricts IAV budding more efficiently than IgG and suggest a role of anti-M2 IgA in cross-protective immunity to IAVs.
• Isolation of Candidatus Bartonella rousetti and Other Bat-associated Bartonellae from Bats and Their Flies in Zambia.

  Yongjin Qiu, Masahiro Kajihara, Ryo Nakao, Evans Mulenga, Hayato Harima, Bernard Mudenda Hang'ombe, Yoshiki Eto, Katendi Changula, Daniel Mwizabi, Hirofumi Sawa, Hideaki Higashi, Aaron Mweene, Ayato Takada, Martin Simuunza, Chihiro Sugimoto

  Pathogens (Basel, Switzerland) 9 (6) 2020/06/13 [Refereed][Not invited]
   

  Bat-associated bartonellae, including Bartonella mayotimonensis and Candidatus Bartonella rousetti, were recently identified as emerging and potential zoonotic agents, respectively. However, there is no report of bat-associated bartonellae in Zambia. Thus, we aimed to isolate and characterize Bartonella spp. from bats and bat flies captured in Zambia by culturing and PCR. Overall, Bartonella spp. were isolated from six out of 36 bats (16.7%), while Bartonella DNA was detected in nine out of 19 bat flies (47.3%). Subsequent characterization using a sequence of five different genes revealed that three isolates obtained from Egyptian fruit bats (Rousettus aegyptiacus) were Ca. B. rousetti. The isolates obtained from insectivorous bats (Macronycteris vittatus) were divided into two previously unclassified bat-associated bartonellae. A phylogenetic analysis of the six genotypes of Bartonella gltA sequences from nine pathogen-positive bat flies revealed that three genotypes belonged to the same clades as bat-associated bartonellae, including Ca. B. rousetti. The other three genotypes represented arthropod-associated bartonellae, which have previously been isolated only from ectoparasites. We demonstrated that Ca. B. rousetti is maintained between bats (R. aegyptiacus) and bat flies in Zambia. Continuous surveillance of Bartonella spp. in bats and serological surveys in humans in Africa are warranted to evaluate the public health importance of bat-associated bartonellae.
• Host ESCRT factors are recruited during chikungunya virus infection and are required for the intracellular viral replication cycle.

  Shiho Torii, Yasuko Orba, Michihito Sasaki, Koshiro Tabata, Yuji Wada, Michael Carr, Jody Hobson-Peters, Roy A Hall, Ayato Takada, Takasuke Fukuhara, Yoshiharu Matsuura, William W Hall, Hirofumi Sawa

  The Journal of biological chemistry 295 (23) 7941 - 7957 2020/06/05 [Refereed][Not invited]
   

  Chikungunya fever is a re-emerging zoonotic disease caused by chikungunya virus (CHIKV), a member of the Alphavirus genus in the Togaviridae family. Only a few studies have reported on the host factors required for intracellular CHIKV trafficking. Here, we conducted an imaging-based siRNA screen to identify human host factors for intracellular trafficking that are involved in CHIKV infection, examined their interactions with CHIKV proteins, and investigated the contributions of these proteins to CHIKV infection. The results of the siRNA screen revealed that host endosomal sorting complexes required for transport (ESCRT) proteins are recruited during CHIKV infection. Co-immunoprecipitation analyses revealed that both structural and nonstructural CHIKV proteins interact with hepatocyte growth factor-regulated tyrosine kinase substrate (HGS), a component of the ESCRT-0 complex. We also observed that HGS co-localizes with the E2 protein of CHIKV and with dsRNA, a marker of the replicated CHIKV genome. Results from gene knockdown analyses indicated that, along with other ESCRT factors, HGS facilitates both genome replication and post-translational steps during CHIKV infection. Moreover, we show that ESCRT factors are also required for infections with other alphaviruses. We conclude that during CHIKV infection, several ESCRT factors are recruited via HGS and are involved in viral genome replication and post-translational processing of viral proteins.
• Prevalence and genotypic characterization of Giardia duodenalis isolates from asymptomatic school-going children in Lusaka, Zambia.

  Shadreck J Tembo, Mable M Mutengo, Lungowe Sitali, Katendi Changula, Ayato Takada, Aaron S Mweene, Edgar Simulundu, Simbarashe Chitanga

  Food and waterborne parasitology 19 e00072  2020/06 [Refereed]
   

  Giardia duodenalis is one of the most common causes of diarrhea in humans with about 250-300 million cases per year. It is considered to be a species complex comprising of eight genetic assemblages (A to H), with assemblages A and B being the major causes of human infections. In this study we carried out genotypic characterization of G. duodenalis isolates detected in asymptomatic school-going children aged 3-16 years. Between May and September 2017, a total of 329 fecal samples were collected from school-going children from Chawama compound of Lusaka City and were screened for Giardia by microscopic examination. All microscopically positive fecal samples were analyzed by semi-nested polymerase chain reaction (PCR) targeting the glutamate dehydrogenase (gdh) gene. Genotyping of amplified PCR products was conducted by restriction fragment length polymorphism (RFLP) and DNA sequence analysis. Microscopically, Giardia was found in 10% (33/329) of fecal samples. The PCR-RFLP analysis of the gdh gene revealed assemblages A and B in 27.3% (9/33) and 72.7% (24/33), respectively. Furthermore, analysis with restriction enzymes identified sub-assemblages AII (27.3%, 9/33), BIII (12.1%, 4/33), BIV (51.5%, 17/33) and mixed infections of BIII and BIV (9.1%, 3/33). Phylogenetic analysis showed the clustering of 27.6% (8/29) and 72.4% (21/29) of Zambian Giardia gdh gene sequences into assemblages A and B, respectively. This study has revealed the presence of both assemblage A and B and that spread of G. duodenalis in school-going children appears to be mostly through anthroponotic transmission. To our knowledge, this is the first report of genotypic characterization of G. duodenalis identified in Zambia.
• Potential Role of Nonneutralizing IgA Antibodies in Cross-Protective Immunity against Influenza A Viruses of Multiple Hemagglutinin Subtypes.

  Kosuke Okuya, Reiko Yoshida, Rashid Manzoor, Shinji Saito, Tadaki Suzuki, Michihito Sasaki, Takeshi Saito, Yurie Kida, Akina Mori-Kajihara, Tatsunari Kondoh, Masahiro Sato, Masahiro Kajihara, Hiroko Miyamoto, Osamu Ichii, Hideaki Higashi, Ayato Takada

  Journal of virology 94 (12) 2020/06/01 [Refereed][Not invited]
   

  IgA antibodies on mucosal surfaces are known to play an important role in protection from influenza A virus (IAV) infection and are believed to be more potent than IgG for cross-protective immunity against IAVs of multiple hemagglutinin (HA) subtypes. However, in general, neutralizing antibodies specific to HA are principally HA subtype specific. Here, we focus on nonneutralizing but broadly cross-reactive HA-specific IgA antibodies. Recombinant IgG, monomeric IgA (mIgA), and polymeric secretory IgA (pSIgA) antibodies were generated based on the sequence of a mouse anti-HA monoclonal antibody (MAb) 5A5 that had no neutralizing activity but showed broad binding capacity to multiple HA subtypes. While confirming that there was no neutralizing activity of the recombinant MAbs against IAV strains A/Puerto Rico/8/1934 (H1N1), A/Adachi/2/1957 (H2N2), A/Hong Kong/483/1997 (H5N1), A/shearwater/South Australia/1/1972 (H6N5), A/duck/England/1/1956 (H11N6), and A/duck/Alberta/60/1976 (H12N5), we found that pSIgA, but not mIgA and IgG, significantly reduced budding and release of most of the viruses from infected cells. Electron microscopy demonstrated that pSIgA deposited newly produced virus particles on the surfaces of infected cells, most likely due to tethering of virus particles. Furthermore, we found that pSIgA showed significantly higher activity to reduce plaque sizes of the viruses than IgG and mIgA. These results suggest that nonneutralizing pSIgA reactive to multiple HA subtypes may play a role in intersubtype cross-protective immunity against IAVs.IMPORTANCE Mucosal immunity represented by pSIgA plays important roles in protection from IAV infection. Furthermore, IAV HA-specific pSIgA antibodies are thought to contribute to cross-protective immunity against multiple IAV subtypes. However, the mechanisms by which pSIgA exerts such versatile antiviral activity are not fully understood. In this study, we generated broadly cross-reactive recombinant IgG and pSIgA having the same antigen-recognition site and compared their antiviral activities in vitro These recombinant antibodies did not show "classical" neutralizing activity, whereas pSIgA, but not IgG, significantly inhibited the production of progeny virus particles from infected cells. Plaque formation was also significantly reduced by pSIgA, but not IgG. These effects were seen in infection with IAVs of several different HA subtypes. Based on our findings, we propose an antibody-mediated host defense mechanism by which mucosal immunity may contribute to broad cross-protection from IAVs of multiple HA subtypes, including viruses with pandemic potential.
• West Nile Virus in Farmed Crocodiles, Zambia, 2019.

  Edgar Simulundu, Kunda Ndashe, Herman M Chambaro, David Squarre, Paul Michael Reilly, Simbarashe Chitanga, Katendi Changula, Andrew N Mukubesa, Joseph Ndebe, John Tembo, Nathan Kapata, Matthew Bates, Yona Sinkala, Bernard M Hang'ombe, King S Nalubamba, Masahiro Kajihara, Michihito Sasaki, Yasuko Orba, Ayato Takada, Hirofumi Sawa

  Emerging infectious diseases 26 (4) 811 - 814 2020/04 [Refereed]
   

  We detected West Nile virus (WNV) nucleic acid in crocodiles (Crocodylus niloticus) in Zambia. Phylogenetically, the virus belonged to lineage 1a, which is predominant in the Northern Hemisphere. These data provide evidence that WNV is circulating in crocodiles in Africa and increases the risk for animal and human transmission.
• エボラウイルスエンベロープへのホスファチジルセリン集積に関与する宿主スクランブラーゼの同定

  南保 明日香, 丸山 隼輝, 今井 正樹, 氏江 美智子, 高田 礼人, 大場 雄介, 河岡 義裕

  日本女性科学者の会学術誌 (一社)日本女性科学者の会 20 (1) 65 - 66 1349-4449 2020/03
• Genetic and Biological Diversity of Porcine Sapeloviruses Prevailing in Zambia.

  Hayato Harima, Masahiro Kajihara, Edgar Simulundu, Eugene Bwalya, Yongjin Qiu, Mao Isono, Kosuke Okuya, Gabriel Gonzalez, Junya Yamagishi, Bernard M Hang'ombe, Hirofumi Sawa, Aaron S Mweene, Ayato Takada

  Viruses 12 (2) 2020/02/05 [Refereed][Not invited]
   

  Porcine sapelovirus (PSV) has been detected worldwide in pig populations. Although PSV causes various symptoms such as encephalomyelitis, diarrhea, and pneumonia in pigs, the economic impact of PSV infection remains to be determined. However, information on the distribution and genetic diversity of PSV is quite limited, particularly in Africa. In this study, we investigated the prevalence of PSV infection in Zambia and characterized the isolated PSVs genetically and biologically. We screened 147 fecal samples collected in 2018 and found that the prevalences of PSV infection in suckling pigs and fattening pigs were high (36.2% and 94.0%, respectively). Phylogenetic analyses revealed that the Zambian PSVs were divided into three different lineages (Lineages 1-3) in the clade consisting of Chinese strains. The Zambian PSVs belonging to Lineages 2 and 3 replicated more efficiently than those belonging to Lineage 1 in Vero E6 and BHK cells. Bioinformatic analyses revealed that genetic recombination events had occurred and the recombination breakpoints were located in the L and 2A genes. Our results indicated that at least two biologically distinct PSVs could be circulating in the Zambian pig population and that genetic recombination played a role in the evolution of PSVs.
• Detection of novel orthoreovirus genomes in shrew (Crocidura hirta) and fruit bat (Rousettus aegyptiacus).

  Hayato Harima, Michihito Sasaki, Masahiro Kajihara, Akina Mori-Kajihara, Bernard M Hang'ombe, Katendi Changula, Yasuko Orba, Hirohito Ogawa, Martin Simuunza, Reiko Yoshida, Aaron Mweene, Ayato Takada, Hirofumi Sawa

  The Journal of veterinary medical science 82 (2) 162 - 167 2020/02/04 [Refereed][Not invited]
   

  Orthoreoviruses have been indentified in several mammals, however, there is no information about orthoreoviruses in shrews. In this study, we screened wild animals in Zambia, including shrews, rodents, and bats for the detection of orthoreoviruses. Two orthoreovirus RNA genomes were detected from a shrew intestinal-contents (1/24) and a bat colon (1/96) sample by reverse-transcription (RT)-PCR targeting the RNA-dependent RNA polymerase gene of orthoreoviruses. Phylogenetic analyses revealed that each of the identified orthoreoviruses formed a distinct branch among members of the Orthoreovirus genus. This is the first report that shrews are susceptible to orthoreovirus infection. Our results suggest the existence of undiscovered orthoreoviruses in shrews and provide important information about the genetic diversity of orthoreoviruses.
• Niemann-Pick C1 Heterogeneity of Bat Cells Controls Filovirus Tropism.

  Yoshihiro Takadate, Tatsunari Kondoh, Manabu Igarashi, Junki Maruyama, Rashid Manzoor, Hirohito Ogawa, Masahiro Kajihara, Wakako Furuyama, Masahiro Sato, Hiroko Miyamoto, Reiko Yoshida, Terence E Hill, Alexander N Freiberg, Heinz Feldmann, Andrea Marzi, Ayato Takada

  Cell reports 30 (2) 308 - 319 2020/01/14 [Refereed][Not invited]
   

  Fruit bats are suspected to be natural hosts of filoviruses, including Ebola virus (EBOV) and Marburg virus (MARV). Interestingly, however, previous studies suggest that these viruses have different tropisms depending on the bat species. Here, we show a molecular basis underlying the host-range restriction of filoviruses. We find that bat-derived cell lines FBKT1 and ZFBK13-76E show preferential susceptibility to EBOV and MARV, respectively, whereas the other bat cell lines tested are similarly infected with both viruses. In FBKT1 and ZFBK13-76E, unique amino acid (aa) sequences are found in the Niemann-Pick C1 (NPC1) protein, one of the cellular receptors interacting with the filovirus glycoprotein (GP). These aa residues, as well as a few aa differences between EBOV and MARV GPs, are crucial for the differential susceptibility to filoviruses. Taken together, our findings indicate that the heterogeneity of bat NPC1 orthologs is an important factor controlling filovirus species-specific host tropism.
• Seroprevalence of Rift Valley fever in cattle of smallholder farmers in Kwilu Province in the Democratic Republic of Congo.

  Abdel-Amir Dib Halawi, Ngonda Saasa, Boniface Lombe Pongombo, Masahiro Kajihara, Herman Moses Chambaro, Mutambel Hity, Hirofumi Sawa, Ayato Takada, Aaron S Mweene, Luamba Lua Nsembo, Edgar Simulundu

  Tropical animal health and production 51 (8) 2619 - 2627 2019/11 

  Rift Valley fever (RVF) is a zoonotic mosquito-borne disease caused by RVF virus (RVFV) that causes abortions and high mortalities in livestock and is also associated with acute and fatal disease in humans. In the Democratic Republic of Congo (DRC), information on the epidemiology of RVF is limited, particularly among cattle reared by smallholder farmers. This cross-sectional study was conducted to investigate the seroprevalence of RVF in cattle raised by smallholder farmers in Kwilu Province of DRC, which has not yet reported an RVF epidemic. A total of 677 cattle sera were collected from four territories and tested for anti-RVFV antibodies using immunofluorescent assay and enzyme-linked immunosorbent assay. The overall seroprevalence of anti-RVFV IgG was 6.5% (44/677) (95% CI 4.81-8.7). There was a statistically significant difference in the seroprevalence among the territories (χ2 = 28.79, p < 0.001). Territory seroprevalences were as follows: Idiofa 14.08% (95% CI 9.78-19.76), Bulungu 4.14% (95% CI 1.83-8.68), Gungu 3.21% (95% CI 1.41-6.78), and Masi-Manimba 1.19% (95% CI 0.06-7.37). Seroprevalence differed significantly among age categories (p = 0.0017) and ecosystem (p < 0.001). The seroprevalence of animals aged between 1 and 2 years was 20.0% (95% CI 8.4-39.13) and was higher than group aged <1 year, between 2 and 3 years, and > 3 years. Forest area (18.92% (95% CI 12.35-27.7)) had higher seropositivity than savannah area (4.06% (95% CI 2.65-6.12)). Sex difference was not significant (χ2 = 0.14, p = 0.704). These findings indicate that cattle in Kwilu Province had been exposed to RVFV, which represents a significant risk for both livestock and human health.
• Molecular detection and characterization of genotype 1 bovine leukemia virus from beef cattle in the traditional sector in Zambia.

  Mundia M Phiri, Evans Kaimoyo, Katendi Changula, Isaac Silwamba, Herman M Chambaro, Penjaninge Kapila, Masahiro Kajihara, Martin Simuunza, John Bwalya Muma, Girja S Pandey, Ayato Takada, Aaron S Mweene, Simbarashe Chitanga, Edgar Simulundu

  Archives of virology 164 (10) 2531 - 2536 2019/10 

  Whilst bovine leukemia virus (BLV) causes considerable economic losses to the dairy industry worldwide, information on its molecular epidemiology and economic impact in beef cattle is limited. Here, blood from 880 animals from Zambia's major cattle-rearing provinces was screened for BLV by nested PCR. Positive pools were sequenced and phylogenetically analyzed. The estimated pooled prevalence was 2.1%. All strains belonged to genotype 1 and formed a distinct phylogenetic cluster. The study suggests circulation of genotype 1 BLV in beef cattle in these regions. This is the first report on molecular detection and characterization of BLV from beef cattle in Africa.
• Genetic diversity of rabies virus in different host species and geographic regions of Zambia and Zimbabwe.

  Muleya W, Chambaro HM, Sasaki M, Gwenhure LF, Mwenechanya R, Kajihara M, Saasa N, Mupila Z, Mori-Kajihara A, Qiu Y, Kangwa E, Mweene A, Namangala B, Takada A, Sawa H

  Virus genes 55 (5) 713 - 719 0920-8569 2019/10 [Refereed][Not invited]
   

  Rabies is endemic in Zambia and Zimbabwe. The previously investigated strains of rabies virus in central Zambia belong to the Africa 1b lineage, with similar circulating virus strains found in the various tested hosts and regions. However, prior work assessed only limited regions and host species. Thus, this study aimed to more comprehensively determine the genetic diversity of rabies virus across regions of Zambia and Zimbabwe. RNA (n = 76) was extracted from positive direct fluorescent antibody test brain tissues from dog, cow, goat, cat, pig, human, and jackal collected from Zambia and Zimbabwe. The amplicons of the nucleoprotein and glycoprotein genes were obtained from all examined samples by nested RT-PCR and subsequently sequenced. A phylogenetic analysis of the N gene confirmed that all the endemic strains of rabies virus in Zambia and Zimbabwe belong to the Africa 1b lineage. The obtained viral gene sequences were phylogenetically divided into two clusters. Cluster II comprised only Zambian strains. In contrast, cluster I comprised both Zambia and Zimbabwe strains, with strains from Zimbabwe forming a distinct lineage from Zambian strains, implying viral genetic divergence due to geographical barriers. However, no evidence of clustering based on host or region was observed, implying the circulation of similar virus strains occurs in different hosts and regions of Zambia and Zimbabwe. The clustering of rabies virus strains from jackals with those from domestic animals provides evidence of similar virus strains circulating in both wildlife and domestic animals, and that the jackal might be one of the potential reservoirs of rabies virus infection. In this study, no strains circulating in Zimbabwe were detected in Zambia.
• A versatile platform technology for recombinant vaccines using non-propagative human parainfluenza virus type 2 vector.

  Junpei Ohtsuka, Masayuki Fukumura, Wakako Furuyama, Shujie Wang, Kenichiro Hara, Mitsuyo Maeda, Masato Tsurudome, Hiroko Miyamoto, Aika Kaito, Nobuyuki Tsuda, Yosky Kataoka, Akira Mizoguchi, Ayato Takada, Tetsuya Nosaka

  Scientific reports 9 (1) 12901 - 12901 2019/09/09 

  Ectopic protein with proper steric structure was efficiently loaded onto the envelope of the F gene-defective BC-PIV vector derived from human parainfluenza virus type 2 (hPIV2) by a reverse genetics method of recombinant virus production. Further, ectopic antigenic peptide was successfully loaded either outside, inside, or at both sides of the envelope of the vector. The BC-PIV vector harboring the Ebola virus GP gene was able to elicit neutralizing antibodies in mice. In addition, BC-PIV with antigenic epitopes of both melanoma gp100 and WT1 tumor antigen induced a CD8+ T-cell-mediated response in tumor-transplanted syngeneic mice. Considering the low pathogenicity and recurrent infections of parental hPIV2, BC-PIV can be used as a versatile vector with high safety for recombinant vaccine development, addressing unmet medical needs.
• 重症熱性血小板減少症候群ウイルスの核タンパク質Nと結合する宿主RNAの機能解明

  水間 奎太, 松野 啓太, 岡松 正敏, 高田 礼人, 迫田 義博

  日本獣医学会学術集会講演要旨集 (公社)日本獣医学会 162回 390 - 390 1347-8621 2019/08
• マダニ中のフレボウイルスの遺伝子系統解析に基づく性状推定

  松野 啓太, 中尾 亮, 梶原 将大, 下田 宙, 海老原 秀喜, 高田 礼人, 前田 健, 岡松 正敏, 迫田 義博

  日本獣医学会学術集会講演要旨集 (公社)日本獣医学会 162回 400 - 400 1347-8621 2019/08
• Marburgvirus in Egyptian Fruit Bats, Zambia.

  Masahiro Kajihara, Bernard M Hang'ombe, Katendi Changula, Hayato Harima, Mao Isono, Kosuke Okuya, Reiko Yoshida, Akina Mori-Kajihara, Yoshiki Eto, Yasuko Orba, Hirohito Ogawa, Yongjin Qiu, Hirofumi Sawa, Edgar Simulundu, Daniel Mwizabi, Musso Munyeme, David Squarre, Victor Mukonka, Aaron Mweene, Ayato Takada

  Emerging infectious diseases 25 (8) 1577 - 1580 1080-6040 2019/08 [Refereed][Not invited]
   

  We detected Marburg virus genome in Egyptian fruit bats (Rousettus aegyptiacus) captured in Zambia in September 2018. The virus was closely related phylogenetically to the viruses that previously caused Marburg outbreaks in the Democratic Republic of the Congo. This finding demonstrates that Zambia is at risk for Marburg virus disease.
• Taxonomy of the order Mononegavirales: update 2019.

  Gaya K Amarasinghe, María A Ayllón, Yīmíng Bào, Christopher F Basler, Sina Bavari, Kim R Blasdell, Thomas Briese, Paul A Brown, Alexander Bukreyev, Anne Balkema-Buschmann, Ursula J Buchholz, Camila Chabi-Jesus, Kartik Chandran, Chiara Chiapponi, Ian Crozier, Rik L de Swart, Ralf G Dietzgen, Olga Dolnik, Jan F Drexler, Ralf Dürrwald, William G Dundon, W Paul Duprex, John M Dye, Andrew J Easton, Anthony R Fooks, Pierre B H Formenty, Ron A M Fouchier, Juliana Freitas-Astúa, Anthony Griffiths, Roger Hewson, Masayuki Horie, Timothy H Hyndman, Dàohóng Jiāng, Elliott W Kitajima, Gary P Kobinger, Hideki Kondō, Gael Kurath, Ivan V Kuzmin, Robert A Lamb, Antonio Lavazza, Benhur Lee, Davide Lelli, Eric M Leroy, Jiànróng Lǐ, Piet Maes, Shin-Yi L Marzano, Ana Moreno, Elke Mühlberger, Sergey V Netesov, Norbert Nowotny, Are Nylund, Arnfinn L Økland, Gustavo Palacios, Bernadett Pályi, Janusz T Pawęska, Susan L Payne, Alice Prosperi, Pedro Luis Ramos-González, Bertus K Rima, Paul Rota, Dennis Rubbenstroth, Mǎng Shī, Peter Simmonds, Sophie J Smither, Enrica Sozzi, Kirsten Spann, Mark D Stenglein, David M Stone, Ayato Takada, Robert B Tesh, Keizō Tomonaga, Noël Tordo, Jonathan S Towner, Bernadette van den Hoogen, Nikos Vasilakis, Victoria Wahl, Peter J Walker, Lin-Fa Wang, Anna E Whitfield, John V Williams, F Murilo Zerbini, Tāo Zhāng, Yong-Zhen Zhang, Jens H Kuhn

  Archives of virology 164 (7) 1967 - 1980 0304-8608 2019/07 [Refereed][Not invited]
   

  In February 2019, following the annual taxon ratification vote, the order Mononegavirales was amended by the addition of four new subfamilies and 12 new genera and the creation of 28 novel species. This article presents the updated taxonomy of the order Mononegavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV).
• Clinical Evaluation of QuickNaviTM-Ebola in the 2018 Outbreak of Ebola Virus Disease in the Democratic Republic of the Congo.

  Sheila Makiala, Daniel Mukadi, Anja De Weggheleire, Shino Muramatsu, Daisuke Kato, Koichi Inano, Fumio Gondaira, Masahiro Kajihara, Reiko Yoshida, Katendi Changula, Aaron Mweene, Placide Mbala-Kingebeni, Jean-Jacques Muyembe-Tamfum, Justin Masumu, Steve Ahuka, Ayato Takada

  Viruses 11 (7) 2019/06/28 

  The recent large outbreaks of Ebola virus disease (EVD) in West Africa and the Democratic Republic of the Congo (DRC) have highlighted the need for rapid diagnostic tests to control this disease. In this study, we clinically evaluated a previously developed immunochromatography-based kit, QuickNaviTM-Ebola. During the 2018 outbreaks in DRC, 928 blood samples from EVD-suspected cases were tested with QuickNaviTM-Ebola and the WHO-approved GeneXpert. The sensitivity and specificity of QuickNaviTM-Ebola, estimated by comparing it to GeneXpert-confirmed cases, were 85% (68/80) and 99.8% (846/848), respectively. These results indicate the practical reliability of QuickNaviTM-Ebola for point-of-care diagnosis of EVD.
• Human Borreliosis Caused by a New World Relapsing Fever Borrelia-like Organism in the Old World.

  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.
• ICTV Virus Taxonomy Profile: Filoviridae.

  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]
  
• Molecular characterization and phylogenetic analysis of Trypanosoma spp. isolated from striped leaf-nosed bat (Hipposideros vittatus) in Zambia.

  Qiu Y, Simuunza M, Mwizabi D, Changula K, Harima H, Takada A, Kajihara M, Takadate Y, Nakao R, Kawabata H, Sugimoto C, Mweene A, Sawa H, Eto Y, Mudenda Hang’ombe B, Hayashida K, Yoshida R, Mori-Kajihara A, Ndebe J

  International Journal for Parasitology 9 234 - 238 2019/04 [Refereed][Not invited]
  
• Taxonomy of the order Mononegavirales: second update 2018.

  Piet Maes, Gaya K Amarasinghe, María A Ayllón, Christopher F Basler, Sina Bavari, Kim R Blasdell, Thomas Briese, Paul A Brown, Alexander Bukreyev, Anne Balkema-Buschmann, Ursula J Buchholz, Kartik Chandran, Ian Crozier, Rik L de Swart, Ralf G Dietzgen, Olga Dolnik, Leslie L Domier, Jan F Drexler, Ralf Dürrwald, William G Dundon, W Paul Duprex, John M Dye, Andrew J Easton, Anthony R Fooks, Pierre B H Formenty, Ron A M Fouchier, Juliana Freitas-Astúa, Elodie Ghedin, Anthony Griffiths, Roger Hewson, Masayuki Horie, Julia L Hurwitz, Timothy H Hyndman, Dàohóng Jiāng, Gary P Kobinger, Hideki Kondō, Gael Kurath, Ivan V Kuzmin, Robert A Lamb, Benhur Lee, Eric M Leroy, Jiànróng Lǐ, Shin-Yi L Marzano, Elke Mühlberger, Sergey V Netesov, Norbert Nowotny, Gustavo Palacios, Bernadett Pályi, Janusz T Pawęska, Susan L Payne, Bertus K Rima, Paul Rota, Dennis Rubbenstroth, Peter Simmonds, Sophie J Smither, Qisheng Song, Timothy Song, Kirsten Spann, Mark D Stenglein, David M Stone, Ayato Takada, Robert B Tesh, Keizō Tomonaga, Noël Tordo, Jonathan S Towner, Bernadette van den Hoogen, Nikos Vasilakis, Victoria Wahl, Peter J Walker, David Wang, Lin-Fa Wang, Anna E Whitfield, John V Williams, Gōngyín Yè, F Murilo Zerbini, Yong-Zhen Zhang, Jens H Kuhn

  Archives of virology 164 (4) 1233 - 1244 0304-8608 2019/04 [Refereed][Not invited]
   

  In October 2018, the order Mononegavirales was amended by the establishment of three new families and three new genera, abolishment of two genera, and creation of 28 novel species. This article presents the updated taxonomy of the order Mononegavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV).
• Therapeutic Monoclonal Antibodies for Ebola Virus Infection Derived from Vaccinated Humans.

  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]
  
• Diversity of spotted fever group rickettsiae and their association with host ticks in Japan.

  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.
• Generation of bat-derived influenza viruses and their reassortants.

  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.
• Infection of newly identified phleboviruses in ticks and wild animals in Hokkaido, Japan indicating tick-borne life cycles.

  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.
• First genetic detection and characterization of canine parvovirus from diarrheic dogs in Zambia.

  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]
  
• Characterization of a novel species of adenovirus from Japanese microbat and role of CXADR as its entry factor.

  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 - 573 2019/01 [Refereed][Not invited]
   

  Recently, bat adenoviruses (BtAdVs) of genus Mastadenovirus have been isolated from various bat species, some of them displaying a wide host range in cell culture. In this study, we isolated two BtAdVs from Japanese wild microbats. While one isolate was classified as Bat mastadenovirus A, the other was phylogenetically independent of other BtAdVs. It was rather related to, but serologically different from, canine adenoviruses. We propose that the latter, isolated from Asian parti-colored bat, should be assigned to a novel species of Bat mastadenovirus. Both isolates replicated in various mammalian cell lines, implying their wide cell tropism. To gain insight into cell tropism of these BtAdVs, we investigated the coxsackievirus and adenovirus receptor (CXADR) for virus entry to the cells. We prepared CXADR-knockout canine kidney cells and found that replication of BtAdVs was significantly hampered in these cells. For confirmation, their replication in canine CXADR-addback cells was rescued to the levels with the original cells. We also found that viral replication was corrected in human or bat CXADR-transduced cells to similar levels as in canine CXADR-addback cells. These results suggest that BtAdVs were able to use several mammalian-derived CXADRs as entry factors.
• A Critical Domain of Ebolavirus Envelope Glycoprotein Determines Glycoform and Infectivity

  Haruhiko Fujihira, Katsuaki Usami, Keita Matsuno, Hideyuki Takeuchi, Kaori Denda-Nagai, Jun-Ichi Furukawa, Yasuro Shinohara, Ayato Takada, Yoshihiro Kawaoka, Tatsuro Irimura

  Scientific Reports 8 (1) 5495  2045-2322 2018/12/01 [Refereed][Not invited]
   

  Ebolaviruses comprises 5 species that exert varying degrees of mortality/infectivity in humans with Reston ebolaviruses (REBOV) showing the lowest and Zaire ebolaviruses (ZEBOV) showing the highest. However, the molecular basis of this differential mortality/infectivity remains unclear. Here, we report that the structural features of ebolavirus envelope glycoproteins (GPS) and one of their counter receptors, macrophage galactose-Type calcium-Type lectin (MGL/CD301), play crucial roles in determining viral infectivity. The low infectivity of REBOV mediated by the interaction between GPS and MGL/CD301 dramatically increased when the N-Terminal 18 amino acids (33rd through 50th) of GPS were replaced with that of ZEBOV. Furthermore, structural analysis of glycans of GPS revealed that N-glycans were more extended in REBOV than in ZEBOV. N-glycan extension was reversed by the replacement of aforementioned N-Terminal 18 amino acid residues. Therefore, these data strongly suggest that extended N-glycans on GPS reduce MGL/CD301-mediated viral infectivity by hindering the interaction between GPS and MGL/CD301 preferentially binds O-glycans.
• Seroprevalence of Filovirus Infection of Rousettus aegyptiacus Bats in Zambia.

  Katendi Changula, Masahiro Kajihara, Akina Mori-Kajihara, Yoshiki Eto, Hiroko Miyamoto, Reiko Yoshida, Asako Shigeno, Bernard Hang'ombe, Yongjin Qiu, Daniel Mwizabi, David Squarre, Joseph Ndebe, Hirohito Ogawa, Hayato Harima, Edgar Simulundu, Ladslav Moonga, Penjaninge Kapila, Wakako Furuyama, Tatsunari Kondoh, Masahiro Sato, Yoshihiro Takadate, Chiho Kaneko, Ryo Nakao, Victor Mukonka, Aaron Mweene, Ayato Takada

  The Journal of infectious diseases 218 (suppl_5) S312-S317 - S317 0022-1899 2018/11/22 [Refereed][Not invited]
   

  Bats are suspected to play important roles in the ecology of filoviruses, including ebolaviruses and marburgviruses. A cave-dwelling fruit bat, Rousettus aegyptiacus, has been shown to be a reservoir of marburgviruses. Using an enzyme-linked immunosorbent assay with the viral glycoprotein antigen, we detected immunoglobulin G antibodies specific to multiple filoviruses in 158 of 290 serum samples of R aegyptiacus bats captured in Zambia during the years 2014-2017. In particular, 43.8% of the bats were seropositive to marburgvirus, supporting the notion that this bat species continuously maintains marburgviruses as a reservoir. Of note, distinct peaks of seropositive rates were repeatedly observed at the beginning of rainy seasons, suggesting seasonality of the presence of newly infected individuals in this bat population. These data highlight the need for continued monitoring of filovirus infection in this bat species even in countries where filovirus diseases have not been reported.
• Single-Nucleotide Polymorphisms in Human NPC1 Influence Filovirus Entry Into Cells.

  Tatsunari Kondoh, Michael Letko, Vincent J Munster, Rashid Manzoor, Junki Maruyama, Wakako Furuyama, Hiroko Miyamoto, Asako Shigeno, Daisuke Fujikura, Yoshihiro Takadate, Reiko Yoshida, Manabu Igarashi, Heinz Feldmann, Andrea Marzi, Ayato Takada

  The Journal of infectious diseases 218 (suppl_5) S397-S402 - S402 0022-1899 2018/11/22 [Refereed][Not invited]
   

  Niemann-Pick C1 (NPC1), a host receptor involved in the envelope glycoprotein (GP)-mediated entry of filoviruses into cells, is believed to be a major determinant of cell susceptibility to filovirus infection. It is known that proteolytically digested Ebola virus (EBOV) GP interacts with 2 protruding loops in domain C of NPC1. Using previously published structural data and the National Center for Biotechnology Information Single-Nucleotide Polymorphism (SNP) database, we identified 10 naturally occurring missense SNPs in human NPC1. To investigate whether these SNPs affect cell susceptibility to filovirus infection, we generated Vero E6 cell lines stably expressing NPC1 with SNP substitutions and compared their susceptibility to vesicular stomatitis virus pseudotyped with filovirus GPs and infectious EBOV. We found that some of the substitutions resulted in reduced susceptibility to filoviruses, as indicated by the lower titers and smaller plaque/focus sizes of the viruses. Our data suggest that human NPC1 SNPs may likely affect host susceptibility to filoviruses.
• Two Conserved Amino Acids within the NSs of Severe Fever with Thrombocytopenia Syndrome Phlebovirus Are Essential for Anti-interferon Activity.

  Moriyama M, Igarashi M, Koshiba T, Irie T, Takada A, Ichinohe T

  Journal of virology 92 (19) 0022-538X 2018/10 [Refereed][Not invited]
   

  The nonstructural protein (NSs) of severe fever with thrombocytopenia syndrome phlebovirus (SFTSV) sequesters TANK-binding kinase 1 (TBK1) into NSs-induced cytoplasmic structures to inhibit the phosphorylation and nuclear translocation of interferon (IFN) regulatory factor 3 (IRF3) and subsequent interferon beta (IFN-β) production. Although the C-terminal region of SFTSV NSs (NSs66-249) has been linked to the formation of NSs-induced cytoplasmic structures and inhibition of host IFN-β responses, the role of the N-terminal region in antagonizing host antiviral responses remains to be defined. Here, we demonstrate that two conserved amino acids at positions 21 and 23 in the SFTSV and heartland virus (HRTV) NSs are essential for suppression of IRF3 phosphorylation and IFN-β mRNA expression following infection with SFTSV or recombinant influenza virus lacking the NS1 gene. Surprisingly, formation of SFTSV/HRTV NSs-induced cytoplasmic structures is not essential for inhibition of host antiviral responses. Rather, an association between SFTSV/HRTV NSs and TBK1 is required for suppression of mitochondrial antiviral signaling protein (MAVS)-mediated activation of IFN-β promoter activity. Although SFTSV NSs did not prevent the ubiquitination of TBK1, it associates with TBK1 through its N-terminal kinase domain (residues 1 to 307) to block the autophosphorylation of TBK1. Furthermore, we found that both wild-type NSs and the 21/23A mutant (NSs in which residues at positions 21 and 23 were replaced with alanine) of SFTSV suppressed NLRP3 inflammasome-dependent interleukin-1β (IL-1β) secretion, suggesting that the importance of these residues is restricted to TBK1-dependent IFN signaling. Together, our findings strongly implicate the two conserved amino acids at positions 21 and 23 of SFTSV/HRTV NSs in the inhibition of host interferon responses.IMPORTANCE Recognition of viruses by host innate immune systems plays a critical role not only in providing resistance to viral infection but also in the initiation of antigen-specific adaptive immune responses against viruses. Severe fever with thrombocytopenia syndrome (SFTS) is a newly emerging infectious disease caused by the SFTS phlebovirus (SFTSV), a highly pathogenic tick-borne phlebovirus. The 294-amino-acid nonstructural protein (NSs) of SFTSV associates with TANK-binding kinase 1 (TBK1), a key regulator of host innate antiviral immunity, to inhibit interferon beta (IFN-β) production and enhance viral replication. Here, we demonstrate that two conserved amino acids at positions 21 and 23 in the NSs of SFTSV and heartland virus, another tick-borne phlebovirus, are essential for association with TBK1 and suppression of IFN-β production. Our results provide important insight into the molecular mechanisms by which SFTSV NSs helps to counteract host antiviral strategies.
• Identification of group A rotaviruses from Zambian fruit bats provides evidence for long-distance dispersal events in Africa

  Michihito Sasaki, Masahiro Kajihara, Katendi Changula, Akina Mori-Kajihara, Hirohito Ogawa, Bernard M. Hang'ombe, Aaron S. Mweene, Martin Simuunza, Reiko Yoshida, Michael Carr, Yasuko Orba, Ayato Takada, Hirofumi Sawa

  Infection, Genetics and Evolution 63 104 - 109 1567-7257 2018/09/01 [Refereed][Not invited]
   

  Group A rotavirus (RVA) is a major cause of diarrhea in children worldwide. Although RVA infects many animals, little is known about RVA in bats. The present study investigated the genetic diversity of RVA in Zambian bats. We identified RVA from two straw-colored fruit bats (Eidolon helvum) and an Egyptian fruit bat (Rousettus aegyptiacus), and analyzed the genome sequences of these strains. Genome segments of the RVA strains from Zambian E. helvum showed 97%–99% nucleotide sequence identity with those of other RVA strains from E. helvum in Cameroon, which is 2800 km from the sampling locations. These findings suggest that migratory straw-colored fruit bat species, distributed across sub-Saharan Africa, have the potential to disseminate RVA across long distances. By contrast, the RVA strain from Zambian R. aegyptiacus carried highly divergent NSP2 and NSP4 genes, leading us to propose novel genotypes N21 and E27, respectively. Notably, this RVA strain also shared the same genotype for VP6 and NSP3 with the RVA strains from Zambian E. helvum, suggesting interspecies transmission and genetic reassortment may have occurred between these two bat species in the past. Our study has important implications for RVA dispersal in bat populations, and expands our knowledge of the ecology, diversity and evolutionary relationships of RVA.
• Fatal Tickborne Phlebovirus Infection in Captive Cheetahs, Japan.

  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.
• Structural Characterization of Pan-ebolavirus Antibody 6D6 Targeting the Fusion Peptide of the Surface Glycoprotein.

  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]
  
• Taxonomy of the order Mononegavirales: update 2018.

  Gaya K Amarasinghe, Nidia G Aréchiga Ceballos, Ashley C Banyard, Christopher F Basler, Sina Bavari, Andrew J Bennett, Kim R Blasdell, Thomas Briese, Alexander Bukreyev, Yíngyún Caì, Charles H Calisher, Cristine Campos Lawson, Kartik Chandran, Colin A Chapman, Charles Y Chiu, Kang-Seuk Choi, Peter L Collins, Ralf G Dietzgen, Valerian V Dolja, Olga Dolnik, Leslie L Domier, Ralf Dürrwald, John M Dye, Andrew J Easton, Hideki Ebihara, Juan E Echevarría, Anthony R Fooks, Pierre B H Formenty, Ron A M Fouchier, Conrad M Freuling, Elodie Ghedin, Tony L Goldberg, Roger Hewson, Masayuki Horie, Timothy H Hyndman, Dàohóng Jiāng, Robert Kityo, Gary P Kobinger, Hideki Kondō, Eugene V Koonin, Mart Krupovic, Gael Kurath, Robert A Lamb, Benhur Lee, Eric M Leroy, Piet Maes, Andrea Maisner, Denise A Marston, Sunil Kumar Mor, Thomas Müller, Elke Mühlberger, Víctor Manuel Neira Ramírez, Sergey V Netesov, Terry Fei Fan Ng, Norbert Nowotny, Gustavo Palacios, Jean L Patterson, Janusz T Pawęska, Susan L Payne, Karla Prieto, Bertus K Rima, Paul Rota, Dennis Rubbenstroth, Martin Schwemmle, Stuart Siddell, Sophie J Smither, Qisheng Song, Timothy Song, Mark D Stenglein, David M Stone, Ayato Takada, Robert B Tesh, Luciano Matsumiya Thomazelli, Keizō Tomonaga, Noël Tordo, Jonathan S Towner, Nikos Vasilakis, Sonia Vázquez-Morón, Claudio Verdugo, Viktor E Volchkov, Victoria Wahl, Peter J Walker, David Wang, Lin-Fa Wang, James F X Wellehan, Michael R Wiley, Anna E Whitfield, Yuri I Wolf, Gōngyín Yè, Yǒng-Zhèn Zhāng, Jens H Kuhn

  Archives of virology 163 (8) 2283 - 2294 0304-8608 2018/08 [Refereed][Not invited]
   

  In 2018, the order Mononegavirales was expanded by inclusion of 1 new genus and 12 novel species. This article presents the updated taxonomy of the order Mononegavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV) and summarizes additional taxonomic proposals that may affect the order in the near future.
• Systematic Analysis of Monoclonal Antibodies against Ebola Virus GP Defines Features that Contribute to Protection.

  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]
  
• The Unique Phylogenetic Position of a Novel Tick-Borne Phlebovirus Ensures an Ixodid Origin of the Genus Phlebovirus.

  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.
• Monoclonal Antibody Cocktail Protects Hamsters From Lethal Marburg Virus Infection.

  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]
   

  Marburg virus (MARV), family Filoviridae, causes Marburg hemorrhagic fever (MHF) in humans and nonhuman primates with case fatality rates of up to 90%. There is no approved therapeutic for MHF, yet several experimental approaches have been evaluated in preclinical studies including small interfering RNA and monoclonal antibody (mAb) treatment. In this study we attempted to improve the therapeutic efficacy of the neutralizing mAb M4 by combining treatment with 1 or 2 of blocking but nonneutralizing mAbs 126-15 and 127-8. We found that single-dose treatment early after infection with the neutralizing mAb M4 or any of the mAb combinations resulted in similar protection in the MARV hamster model. However, a single-dose treatment with the cocktail of all 3 mAbs provided the best protection in delayed treatment, with 67%-100% of the animals surviving a lethal challenge depending on the time of treatment. This study identified a new promising mAb cocktail as a therapeutic option for MHF.
• Genetic characterisation of African swine fever virus from 2017 outbreaks in Zambia: Identification of p72 genotype II variants in domestic pigs.

  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]
  
• Characterization of field infectious bursal disease viruses in Zambia: evidence of co-circulation of multiple genotypes with predominance of very virulent strains

  Racheal Mwenda, Katendi Changula, Bernard M. Hang’ombe, Nozyechi Chidumayo, Alfred S. Mangani, Titus Kaira, Ayato Takada, Aaron S. Mweene, Edgar Simulundu

  Avian Pathology 47 (3) 300 - 313 1465-3338 2018/05/04 [Refereed][Not invited]
   

  Infectious bursal disease (IBD) is a highly contagious, immunosuppressive disease of chickens and causes substantial economic losses to the poultry industry globally. This study investigated the genetic characteristics and pathological lesions induced by IBD viruses (IBDVs) that were associated with 60 suspected outbreaks in chickens during 2015–2016 in Lusaka Province, Zambia. Nucleotide sequences of VP2 hypervariable region (VP2-HVR) (n = 38) and part of VP1 (n = 37) of Zambian IBDVs were phylogenetically analysed. Phylogenetic analysis of the VP2-HVR and VP1 revealed that most viruses (n = 31 of each genome segment) clustered with the very virulent (vv) strains. The rest of the viruses clustered with the classical strains, with two of the viruses being closely related to attenuated vaccine isolates. Two of the viruses that belonged to the vv genotype had a unique amino acid (aa) substitution Q324L whereas one virus had two unique changes, N280S and E300A in the VP2-HVR aa sequence. Although Zambian strains with a vv genotype possessed virulence marker aa within VP1 at 145T, 146D and 147N, two viruses showed unique substitutions, with one virus having 147T while the other had 147H. Pathologically, it was noted that only viruses with a vv genotype appeared to be associated with inducing pathological lesions in non-lymphoid organs (proventriculus and gizzard). Whilst documenting for the first time the presence of classical virulent IBDVs, this study demonstrates the involvement of multiple genotypes, with predominance of vvIBDVs in the epidemiology of IBD in Zambia.
• The Role of Heparan Sulfate Proteoglycans as an Attachment Factor for Rabies Virus Entry and Infection.

  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]
   

  Rabies virus (RABV) is the causative agent of fatal neurological disease. Cellular attachment is the initial and essential step for viral infections. Although extensive studies have demonstrated that RABV uses various target cell molecules to mediate infection, no specific molecule has been identified as an attachment factor for RABV infection. Here we demonstrate that cellular heparan sulfate (HS) supports RABV adhesion and subsequent entry into target cells. Enzymatic removal of HS reduced cellular susceptibility to RABV infection, and heparin, a highly sulfated form of HS, blocked viral adhesion and infection. The direct binding between RABV glycoprotein and heparin was demonstrated, and this interaction was shown to require HS N- and 6-O-sulfation. We also revealed that basic amino acids in the ectodomain of RABV glycoprotein serve as major determinants for the RABV-HS interaction. Collectively, our study highlights a previously undescribed role of HS as an attachment factor for RABV infection.
• Tick-borne haemoparasites and Anaplasmataceae in domestic dogs in Zambia.

  Yongjin Qiu, Chiho Kaneko, Masahiro Kajihara, Saasa Ngonda, Edgar Simulundu, Walter Muleya, May June Thu, Mudenda Bernard Hang'ombe, Ken Katakura, Ayato Takada, Hirofumi Sawa, Martin Simuunza, Ryo Nakao

  Ticks and tick-borne diseases 9 (4) 988 - 995 1877-959X 2018/05 [Refereed][Not invited]
   

  Tick-borne diseases (TBDs), including emerging and re-emerging infectious diseases, are important threats to human and animal health worldwide. Indeed, the number of reported human and animal infectious cases of novel TBD agents has increased in recent decades. However, TBDs tend to be neglected, especially in resource-limited countries that often have limited diagnostic capacity. The aim of this molecular survey was to detect and characterise tick-borne pathogens (Babesia, Theileria, and Hepatozoon parasites and Anaplasmataceae bacteria) in domestic dogs in Zambia. In total, 247 canine peripheral blood samples were collected in Lusaka, Mazabuka, Monze, and Shangombo. Conventional PCR to detect the selected pathogens was performed using DNA extracted from canine blood. One hundred eleven samples were positive for protozoa and 5 were positive for Anaplasmataceae. Sequencing of thirty-five randomly selected protozoa-positive samples revealed the presence of Babesia rossi, Babesia vogeli, and Hepatozoon canis 18S rDNA. Based on these sequences, a multiplex PCR system was developed to yield PCR products with different amplicons, the size of which depended on the parasite species; thus, each species could be identified without the need for sequence analysis. Approximately 40% of dogs were positive for H. canis. In particular, the positive rate (75.2%) of H. canis infection was significantly higher in Shangombo than in other sampling sites. Multiplex PCR assay detected B. rossi and B. vogeli infections in five and seven dogs, respectively, indicating that this approach is useful for detecting parasites with low prevalence. Sequencing analysis of gltA and groEL genes of Anaplasmataceae revealed that two and one dogs in Lusaka were infected with Anaplasma platys and Ehrlichia canis, respectively. The data indicated that Zambian dogs were infected with multiple tick-borne pathogens such as H. canis, B. rossi, B. vogeli, A. platys, E. canis and uncharacterized Ehrlichia sp. Since some of these parasites are zoonotic, concerted efforts are needed to raise awareness of, and control, these tick-borne pathogens.
• Expression of a Recombinant Nucleocapsid Protein of Rift Valley Fever Virus in Vero Cells as an Immunofluorescence Antigen and Its Use for Serosurveillance in Traditional Cattle Herds in Zambia

  Ngonda Saasa, Masahiro Kajihara, George Dautu, Akina Mori-Kajihara, Shuetsu Fukushi, Yona Sinkala, Shigeru Morikawa, Aaron Mweene, Ayato Takada, Kumiko Yoshimatsu, Jiro Arikawa

  Vector-Borne and Zoonotic Diseases 18 (5) 273 - 277 1557-7759 2018/05/01 [Refereed][Not invited]
   

  The open reading frame of the nucleocapsid protein (NP) of Rift Valley fever virus (RVFV) strain MP12 was cloned and expressed in Vero E6 cells. The recombinant NP (rNP)-expressing cells were used as antigens for an indirect immunofluorescent antibody assay (IFA). The rNP-based IFA and RVFV-infected Vero E6 cell (authentic antigen)-based IFA showed similar IFA profiles with immune rabbit serum, which was prepared by immunization with rNP expressed using a baculovirus vector. A total of 942 traditional cattle sera obtained in five districts in Central, Southern, and Western provinces of Zambia were screened for anti-RVFV antibodies by the authentic antigen-based and rNP-based IFAs. Significant agreement was obtained between the two IFAs. The findings show that the rNP-based IFA is a safe and useful diagnostic tool as an alternative to the authentic antigen-based IFA. The antibody titers given by the rNP-based IFA were higher than those by the authentic antigen-based IFA. Therefore, the rNP-based IFA might be useful for serosurveillance of RVFV infection among cattle. Antibody prevalence rates in the five districts were 1.3% to 13.5% in the authentic antigen-based IFA and 6.0% to 21.4% in the rNP-based IFA. The results indicated that despite no reports of active cases of RVF in these provinces of Zambia, the virus is circulating among cattle herds.
• Co-circulation of multiple genotypes of African swine fever viruses among domestic pigs in Zambia (2013-2015).

  Simulundu E, Chambaro HM, Sinkala Y, Kajihara M, Ogawa H, Mori A, Ndebe J, Dautu G, Mataa L, Lubaba CH, Simuntala C, Fandamu P, Simuunza M, Pandey GS, Samui KL, Misinzo G, Takada A, Mweene AS

  Transboundary and emerging diseases 65 (1) 114 - 122 1865-1674 2018/02 [Refereed][Not invited]
  
• First molecular detection and genetic characterization of Coxiella burnetii in Zambian dogs and rodents.

  Simbarashe Chitanga, Edgar Simulundu, Martin C Simuunza, Katendi Changula, Yongjin Qiu, Masahiro Kajihara, Ryo Nakao, Michelo Syakalima, Ayato Takada, Aaron S Mweene, Samson Mukaratirwa, Bernard M Hang'ombe

  Parasites & vectors 11 (1) 40 - 40 2018/01/17 [Refereed][Not invited]
   

  Coxiella burnetii, the causative agent of Q fever, is a zoonotic pathogen associated with sylvatic or domestic transmission cycles, with rodents being suspected to link the two transmission cycles. Infection and subsequent disease in humans has historically been associated with contact with infected livestock, especially sheep. However, recently there have been reports of Q fever outbreaks associated with contact with infected rodents and dogs. Studies exploring the potential role of these animal hosts in the epidemiology of Q fever in many developing countries in Africa are very limited. This study aimed to determine the potential role of rodents and dogs in the epidemiological cycle of C. burnetti in Zambia. Using pathogen-specific polymerase chain reaction assays targeting the 16S rRNA gene, C. burnetii was detected for the first time in 45% of rodents (9/20), in one shrew and in 10% of domestic dogs (15/150) screened in Zambia. Phylogenetic characterization of six samples based on the isocitrate synthase gene revealed that the strains were similar to a group of isolates from chronic human Q fever patients, goats and rodents reported in multiple continents. Considering the close proximity of domestic dogs and rodents to humans, especially in resource-limited communities, the presence of C. burnetii in these animals could be of significant public health importance. It is thus important to determine the burden of Q fever in humans in such resource-limited communities where there is close contact between humans, rodents and dogs.
• Ebola virus requires a host scramblase for externalization of phosphatidylserine on the surface of viral particles.

  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.
• Molecular detection and characterization of zoonotic Anaplasma species in domestic dogs in Lusaka, Zambia

  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.
• Influenza A virus M2 protein: Roles from ingress to egress

  Rashid Manzoor, Manabu Igarashi, Ayato Takada

  International Journal of Molecular Sciences 18 (12) E2649  1422-0067 2017/12/07 [Refereed][Not invited]
   

  Influenza A virus (IAV) matrix protein 2 (M2) is among the smallest bona fide, hence extensively studied, ion channel proteins. The M2 ion channel activity is not only essential for virus replication, but also involved in modulation of cellular homeostasis in a variety of ways. It is also the target for ion channel inhibitors, i.e., anti-influenza drugs. Thus far, several studies have been conducted to elucidate its biophysical characteristics, structure-function relationships of the ion channel, and the M2-host interactome. In this review, we discuss M2 protein synthesis and assembly into an ion channel, its roles in IAV replication, and the pathophysiological impact on the host cell.
• Characterization of a Novel Bat Adenovirus Isolated from Straw-Colored Fruit Bat (Eidolon helvum).

  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.
• Putative endogenous filovirus VP35-like protein potentially functions as an IFN antagonist but not a polymerase cofactor

  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.
• The Epidemiology of African Swine Fever in "Nonendemic" Regions of Zambia (1989-2015): Implications for Disease Prevention and Control

  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.
• Rapid detection of all known ebolavirus species by reverse transcription-loop-mediated isothermal amplification (RT-LAMP)

  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.
• Genetic characterization of orf virus associated with an outbreak of severe orf in goats at a farm in Lusaka, Zambia (2015)

  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.
• Taxonomy of the order Mononegavirales: update 2017

  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).
• Rapid and broad detection of H5 hemagglutinin by an immunochromatographic kit using novel monoclonal antibody against highly pathogenic avian influenza virus belonging to the genetic clade 2.3.4.4

  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.
• Possibility and Challenges of Conversion of Current Virus Species Names to Linnaean Binomials

  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.
• Implementation of Objective PASC-Derived Taxon Demarcation Criteria for Official Classification of Filoviruses

  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.
• Clinical and subclinical bovine leukemia virus infection in a dairy cattle herd in Zambia

  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.
• エボラウイルスの糖タンパク質を標的化する大環状ペプチドの開発(Development of Macrocyclic Peptide Targeting Ebola Glycoprotein)

  Umemoto Shiori, Maruyama Junki, Takada Ayato, Suga Hiroaki

  日本薬学会年会要旨集 137年会 (1) 67 - 67 0918-9823 2017/03
• Putative RNA viral sequences detected in an Ixodes scapularis-derived cell line

  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.
• Genetic Predisposition To Acquire a Polybasic Cleavage Site for Highly Pathogenic Avian Influenza Virus Hemagglutinin

  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.
• Quantification of filovirus glycoprotein-specific antibodies

  Wakako Furuyama, Hiroko Miyamoto, Reiko Yoshida, Ayato Takada

  Methods in Molecular Biology 1628 309 - 320 1064-3745 2017 [Refereed][Not invited]
   

  Serological methods such as the enzyme-linked immunosorbent assay (ELISA) and virus neutralization test are fundamental tools used in diagnosis, seroepidemiological studies of filovirus transmission/prevalence, and the evaluation of vaccine immunogenicity and potential therapeutic antibodies. Filoviruses have a single transmembrane glycoprotein (GP), which is the only known target of neutralizing antibodies. Here we describe serological methods to quantify filovirus GP-specific antibodies.
• The Tetherin Antagonism of the Ebola Virus Glycoprotein Requires an Intact Receptor-Binding Domain and Can Be Blocked by GP1-Specific Antibodies

  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.
• Fc gamma-receptor IIa-mediated Src Signaling Pathway Is Essential for the Antibody-Dependent Enhancement of Ebola Virus Infection

  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.
• Lujo viral hemorrhagic fever: considering diagnostic capacity and preparedness in the wake of recent Ebola and Zika virus outbreaks

  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.
• Development of an Immunochromatography Assay (QuickNavi-Ebola) to Detect Multiple Species of Ebolaviruses.

  Yoshida R, Muramatsu S, Akita H, Saito Y, Kuwahara M, Kato D, Changula K, Miyamoto H, Kajihara M, Manzoor R, Furuyama W, Marzi A, Feldmann H, Mweene A, Masumu J, Kapeteshi J, Muyembe-Tamfum JJ, Takada A

  The Journal of infectious diseases 214 (suppl 3) S185 - S191 0022-1899 2016/10 [Refereed][Not invited]
   

  The latest outbreak of Ebola virus disease (EVD) in West Africa has highlighted the urgent need for the development of rapid and reliable diagnostic assays. We used monoclonal antibodies specific to the ebolavirus nucleoprotein to develop an immunochromatography (IC) assay (QuickNavi-Ebola) for rapid diagnosis of EVD. The IC assay was first evaluated with tissue culture supernatants of infected Vero E6 cells and found to be capable of detecting 103-104 focus-forming units/mL of ebolaviruses. Using serum samples from experimentally infected nonhuman primates, we confirmed that the assay could detect the viral antigen shortly after disease onset. It was also noted that multiple species of ebolaviruses could be detected by the IC assay. Owing to the simplicity of the assay procedure and absence of requirements for special equipment and training, QuickNavi-Ebola is expected to be a useful tool for rapid diagnosis of EVD.
• Isolation of a sp nov Ljungan virus from wild birds in Japan

  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.
• Taxonomy of the order Mononegavirales: update 2016

  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).
• Constitutive aryl hydrocarbon receptor signaling constrains type I interferon-mediated antiviral innate defense

  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.
• Diagnosis and genotyping of African swine fever viruses from 2015 outbreaks in Zambia

  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.
• Molecular characterization of infectious bursal disease viruses detected in vaccinated commercial broiler flocks in Lusaka, 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.
• Development and Evaluation of Reverse Transcription-Loop-Mediated Isothermal Amplification (RT-LAMP) Assay Coupled with a Portable Device for Rapid Diagnosis of Ebola Virus Disease in Guinea

  Yohei Kurosaki, N’Faly Magassouba, Olamide K. Oloniniyi, Mahamoud S. Cherif, Saori Sakabe, Ayato Takada, Kenji Hirayama, Jiro Yasuda

  PLoS Neglected Tropical Diseases 10 (2) e0004472  1935-2735 2016/02/22 [Refereed][Not invited]
   

  Given the current absence of specific drugs or vaccines for Ebola virus disease (EVD), rapid, sensitive, and reliable diagnostic methods are required to stem the transmission chain of the disease. We have developed a rapid detection assay for Zaire ebolavirus based on reverse transcription-loop-mediated isothermal amplification (RT-LAMP) and coupled with a novel portable isothermal amplification and detection platform. The RT-LAMP assay is based on primer sets that target the untranscribed trailer region or nucleoprotein coding region of the viral RNA. The test could specifically detect viral RNAs of Central and West African Ebola virus strains within 15 minutes with no cross-reactivity to other hemorrhagic fever viruses and arboviruses, which cause febrile disease. The assay was evaluated using a total of 100 clinical specimens (serum, n = 44 oral swab, n = 56) collected from suspected EVD cases in Guinea. The specificity of this diagnostic test was 100% for both primer sets, while the sensitivity was 100% and 97.9% for the trailer and nucleoprotein primer sets, respectively, compared with a reference standard RT-PCR test. These observations suggest that our diagnostic assay is useful for identifying EVD cases, especially in the field or in settings with insufficient infrastructure.
• Development and Evaluation of Reverse Transcription-Loop-Mediated Isothermal Amplification (RT-LAMP) Assay Coupled with a Portable Device for Rapid Diagnosis of Ebola Virus Disease in Guinea

  Yohei Kurosaki, N'Faly Magassouba, Olamide K. Oloniniyi, Mahamoud S. Cherif, Saori Sakabe, Ayato Takada, Kenji Hirayama, Jiro Yasuda

  PLOS NEGLECTED TROPICAL DISEASES 10 (2) 1935-2735 2016/02 [Refereed][Not invited]
   

  Given the current absence of specific drugs or vaccines for Ebola virus disease (EVD), rapid, sensitive, and reliable diagnostic methods are required to stem the transmission chain of the disease. We have developed a rapid detection assay for Zaire ebolavirus based on reverse transcription-loop-mediated isothermal amplification (RT-LAMP) and coupled with a novel portable isothermal amplification and detection platform. The RT-LAMP assay is based on primer sets that target the untranscribed trailer region or nucleoprotein coding region of the viral RNA. The test could specifically detect viral RNAs of Central and West African Ebola virus strains within 15 minutes with no cross-reactivity to other hemorrhagic fever viruses and arboviruses, which cause febrile disease. The assay was evaluated using a total of 100 clinical specimens (serum, n = 44; oral swab, n = 56) collected from suspected EVD cases in Guinea. The specificity of this diagnostic test was 100% for both primer sets, while the sensitivity was 100% and 97.9% for the trailer and nucleoprotein primer sets, respectively, compared with a reference standard RT-PCR test. These observations suggest that our diagnostic assay is useful for identifying EVD cases, especially in the field or in settings with insufficient infrastructure.
• Discovery of an antibody for pan-ebolavirus therapy

  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.
• Characterization of the glycoproteins of bat-derived influenza viruses

  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.
• Interferon-Induced Transmembrane Protein-Mediated Inhibition of Host Cell Entry of Ebolaviruses

  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.
• Seroepidemiological Prevalence of Multiple Species of Filoviruses in Fruit Bats (Eidolon helvum) Migrating in Africa

  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.
• A Single Amino Acid in the M1 Protein Responsible for the Different Pathogenic Potentials of H5N1 Highly Pathogenic Avian Influenza Virus Strains

  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.
• Genetic and antigenic characterization of H5 and H7 influenza viruses isolated from migratory water birds in Hokkaido, Japan and Mongolia from 2010 to 2014

  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.
• Molecular epidemiology of pathogenic Leptospira spp. in the straw-colored fruit bat (Eidolon helvum) migrating to Zambia from the Democratic Republic of Congo

  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.
• Interaction between TIM-1 and NPC1 Is Important for Cellular Entry of Ebola Virus

  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.
• Pathological and molecular diagnosis of the 2013 African swine fever outbreak in Lusaka, Zambia.

  Yabe J, Hamambulu P, Simulundu E, Ogawa H, Kajihara M, Mori-Kajihara A, Changula-Chitanga K, Mwase M, Mweemba-Muwowo M, Chambaro HM, Mataa L, Hang'ombe B, Namangala B, Fandamu P, Sawa H, Takada A, Higashi H, Mweene AS

  Tropical animal health and production 47 (2) 459 - 463 0049-4747 2015/02 [Refereed][Not invited]
   

  African swine fever (ASF) is a highly contagious and fatal hemorrhagic viral disease of domestic pigs. The disease is widespread in sub-Saharan Africa and has repeatedly been introduced into other continents. The current study describes the diagnostic investigations of a hemorrhagic disease that was reported in pigs in Lusaka (October 2013), Zambia. Necropsy, histopathology, and molecular diagnosis using polymerase chain reaction and sequence analysis confirmed the disease to be ASF. The sequences obtained showed high similarity to previously isolated ASF viruses. Consistent surveillance and rapid diagnosis of the disease is recommended to prevent future outbreaks and economic losses as there is currently no vaccine against the disease.
• Comprehensive Molecular Detection of Tick-Borne Phleboviruses Leads to the Retrospective Identification of Taxonomically Unassigned Bunyaviruses and the Discovery of a Novel Member of the Genus Phlebovirus

  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.
• A polymorphism of the TIM-1 IgV domain: Implications for the susceptibility to filovirus infection

  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.
• The zoonotic potential of avian influenza viruses isolated from wild waterfowl in Zambia

  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.
• Filovirus RefSeq Entries: Evaluation and Selection of Filovirus Type Variants, Type Sequences, and Names

  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.
• Protective Efficacy of Passive Immunization with Monoclonal Antibodies in Animal Models of H5N1 Highly Pathogenic Avian Influenza Virus Infection

  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.
• Virus nomenclature below the species level: a standardized nomenclature for filovirus strains and variants rescued from cDNA

  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.
• Role of the C-Terminal Region of Vervet Monkey Polyomavirus 1 VP1 in Virion Formation

  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.
• Discussions and decisions of the 2012-2014 International Committee on Taxonomy of Viruses (ICTV) Filoviridae Study Group, January 2012-June 2013

  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.
• Molecular Epidemiology of Paramyxoviruses in Frugivorous Eidolon helvum Bats in Zambia

  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.
• Heat Shock Protein 70 Modulates Influenza A Virus Polymerase Activity

  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.
• Characterization of the Envelope Glycoprotein of a Novel Filovirus, Lloviu Virus

  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.
• Comparison of Antiviral Activity between IgA and IgG Specific to Influenza Virus Hemagglutinin: Increased Potential of IgA for Heterosubtypic Immunity

  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.
• Effect of the PB2 and M Genes on the Replication of H6 Influenza Virus in Chickens.

  Ozaki H, Guan Y, Peiris M, Webster R, Takada A, Webby R

  Influenza research and treatment 2014 547839  2090-1380 2014 [Refereed][Not invited]
  
• Suppression of Fas-mediated apoptosis via steric shielding by filovirus glycoproteins

  Osamu Noyori, Eri Nakayama, Junki Maruyama, Reiko Yoshida, Ayato Takada

  Biochemical and Biophysical Research Communications 441 (4) 994 - 998 0006-291X 2013/11/29 [Refereed][Not invited]
   

  Apoptotic death of virus-infected cells is generally thought to be a defense mechanism to limit the spread of infectious virions by eliminating virus-producing cells in host animals. On the other hand, several viruses have been shown to have anti-apoptotic mechanisms to facilitate efficient viral replication and transmission. In this study, we found that the filovirus glycoprotein (GP) expressed on cell surfaces formed a steric shield over the Fas molecule and that GP-expressing cells showed resistance to cell death induced by a Fas agonistic antibody. These results suggest that filovirus GP-mediated steric shielding may interfere with the Fas-induced apoptotic signal transduction in infected cells and serve as an immune evasion mechanism for filoviruses. © 2013 Elsevier Inc.
• Differential potential for envelope glycoprotein-mediated steric shielding of host cell surface proteins among filoviruses

  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.
• The genetic and antigenic diversity of avian influenza viruses isolated from domestic ducks, muscovy ducks, and chickens in northern and southern Vietnam, 2010-2012

  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.
• Mapping of conserved and species-specific antibody epitopes on the Ebola virus nucleoprotein

  Katendi Changula, Reiko Yoshida, Osamu Noyori, Andrea Marzi, Hiroko Miyamoto, Mari Ishijima, Ayaka Yokoyama, Masahiro Kajihara, Heinz Feldmann, Aaron S. Mweene, Ayato Takada

  Virus Research 176 (1-2) 83 - 90 0168-1702 2013/09 [Refereed][Not invited]
   

  Filoviruses (viruses in the genus Ebolavirus and Marburgvirus in the family Filoviridae) cause severe haemorrhagic fever in humans and nonhuman primates. Rapid, highly sensitive, and reliable filovirus-specific assays are required for diagnostics and outbreak control. Characterisation of antigenic sites in viral proteins can aid in the development of viral antigen detection assays such immunochromatography-based rapid diagnosis. We generated a panel of mouse monoclonal antibodies (mAbs) to the nucleoprotein (NP) of Ebola virus belonging to the species Zaire ebolavirus. The mAbs were divided into seven groups based on the profiles of their specificity and cross-reactivity to other species in the Ebolavirus genus. Using synthetic peptides corresponding to the Ebola virus NP sequence, the mAb binding sites were mapped to seven antigenic regions in the C-terminal half of the NP, including two highly conserved regions among all five Ebolavirus species currently known. Furthermore, we successfully produced species-specific rabbit antisera to synthetic peptides predicted to represent unique filovirus B-cell epitopes. Our data provide useful information for the development of Ebola virus antigen detection assays. © 2013 Elsevier B.V.
• A novel Ebola virus expressing luciferase allows for rapid and quantitative testing of antivirals

  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.
• Heterosubtypic Antiviral Activity of Hemagglutinin-Specific Antibodies Induced by Intranasal Immunization with Inactivated Influenza Viruses in Mice

  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.
• Virus nomenclature below the species level: a standardized nomenclature for laboratory animal-adapted strains and variants of viruses assigned to the family Filoviridae

  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.
• Cross-Reactivity of Secondary Antibodies against African Rodents and Application for Sero-Surveillance

  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.
• Novel mutations in Marburg virus glycoprotein associated with viral evasion from antibody mediated immune pressure

  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.
• Resampling Nucleotide Sequences with Closest-Neighbor Trimming and Its Comparison to Other Methods

  Kouki Yonezawa, Manabu Igarashi, Keisuke Ueno, Ayato Takada, Kimihito Ito

  PLoS ONE 8 (2) e57684  1932-6203 2013/02/27 [Refereed][Not invited]
   

  A large number of nucleotide sequences of various pathogens are available in public databases. The growth of the datasets has resulted in an enormous increase in computational costs. Moreover, due to differences in surveillance activities, the number of sequences found in databases varies from one country to another and from year to year. Therefore, it is important to study resampling methods to reduce the sampling bias. A novel algorithm-called the closest-neighbor trimming method-that resamples a given number of sequences from a large nucleotide sequence dataset was proposed. The performance of the proposed algorithm was compared with other algorithms by using the nucleotide sequences of human H3N2 influenza viruses. We compared the closest-neighbor trimming method with the naive hierarchical clustering algorithm and k-medoids clustering algorithm. Genetic information accumulated in public databases contains sampling bias. The closest-neighbor trimming method can thin out densely sampled sequences from a given dataset. Since nucleotide sequences are among the most widely used materials for life sciences, we anticipate that our algorithm to various datasets will result in reducing sampling bias. © 2013 Yonezawa et al.
• The PB2, PA, HA, NP, and NS genes of a highly pathogenic avian influenza virus A/whooper swan/Mongolia/3/2005 (H5N1) are responsible for pathogenicity in ducks

  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.
• Molecular Mechanisms Underlying Oseltamivir Resistance Mediated by an I117V Substitution in the Neuraminidase of Subtype H5N1 Avian Influenza A Viruses

  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.
• Inhibition of Marburg Virus Budding by Nonneutralizing Antibodies to the Envelope Glycoprotein

  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.
• エボラウイルス表層糖タンパク質に対する糖鎖修飾制御メカニズムの解明

  藤平 陽彦, 宇佐美 克明, 伝田 香里, 松野 啓太, 篠原 康郎, 佐藤 あやの, 高田 礼人, 入村 達郎

  日本生化学会大会プログラム・講演要旨集 (公社)日本生化学会 85回 3T15 - 08 2012/12
• Molecular surveillance and phylogenetic analysis of Old World arenaviruses in Zambia

  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.
• Heterosubtypic antibody recognition of the influenza virus hemagglutinin receptor binding site enhanced by avidity

  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.
• Immune Parameters Correlate with Protection Against Ebola Virus Infection in Rodents and Nonhuman Primates

  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.
• Serological Evidence of Ebola Virus Infection in Indonesian Orangutans

  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.
• Memory Immune Responses against Pandemic (H1N1) 2009 Influenza Virus Induced by a Whole Particle Vaccine in Cynomolgus Monkeys Carrying Mafa-A1*052:02

  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.
• Protective Efficacy of Neutralizing Monoclonal Antibodies in a Nonhuman Primate Model of Ebola Hemorrhagic Fever

  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.
• Reintroduction of H5N1 highly pathogenic avian influenza virus by migratory water birds, causing poultry outbreaks in the 2010-2011 winter season in Japan

  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.
• Adaptation of a Duck Influenza A Virus in Quail

  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.
• Osteopontin is critical to determine symptom severity of influenza through the regulation of NK cell population

  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.
• Filovirus tropism: cellular molecules for viral entry

  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.
• Single Immunization With a Monovalent Vesicular Stomatitis Virus-Based Vaccine Protects Nonhuman Primates Against Heterologous Challenge With Bundibugyo ebolavirus

  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.
• Antibody-Dependent Enhancement of Marburg Virus Infection

  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.
• Novel Arenavirus, Zambia

  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.
• Gnarled-Trunk Evolutionary Model of Influenza A Virus Hemagglutinin

  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.
• Cross-Protective Peptide Vaccine against Influenza A Viruses Developed in HLA-A*2402 Human Immunity Model

  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.
• Characterization of influenza A viruses isolated from wild waterfowl in Zambia

  Edgar Simulundu, Akihiro Ishii, Manabu Igarashi, Aaron S. Mweene, Yuka Suzuki, Bernard M. Hang&apos, 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.
• Prediction of the antigenic changes of the pandemic (H1N1) 2009 influenza virus hemagglutinin

  Manabu Igarashi, Kimihito Ito, Reiko Yoshida, Daisuke Tomabechi, Hiroshi Kida, Ayato Takada

  INFLUENZA AND OTHER RESPIRATORY VIRUSES 5 402 - 404 1750-2640 2011/05 [Refereed][Not invited]
  
• Involvement of viral envelope GP2 in Ebola virus entry into cells expressing the macrophage galactose-type C-type lectin

  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.
• Requirement for Siva-1 for replication of influenza A virus through apoptosis induction

  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.
• Detection of all known filovirus species by reverse transcription-polymerase chain reaction using a primer set specific for the viral nucleoprotein gene

  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.
• Genetic characterization and susceptibility on poultry and mammal of H7N6 subtype avian influenza virus isolated in Japan in 2009

  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.
• Antigenic, genetic, and pathogenic characterization of H5N1 highly pathogenic avian influenza viruses isolated from dead whooper swans (Cygnus cygnus) found in northern Japan in 2008

  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.
• C-type lectins do not act as functional receptors for filovirus entry into cells

  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.
• Enzyme-Linked Immunosorbent Assay for Detection of Filovirus Species-Specific Antibodies

  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.
• Characterization of H5N1 highly pathogenic avian influenza virus strains isolated from migratory waterfowl in Mongolia on the way back from the southern Asia to their northern territory

  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.
• Characterization of human single-chain antibodies against highly pathogenic avian influenza H5N1 viruses: mimotope and neutralizing activity

  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.
• Influenza A Virus Polymerase Inhibits Type I Interferon Induction by Binding to Interferon beta Promoter Stimulator 1

  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.
• Different Potential of C-Type Lectin-Mediated Entry between Marburg Virus Strains

  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.
• Predicting the Antigenic Structure of the Pandemic (H1N1) 2009 Influenza Virus Hemagglutinin

  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.
• Characterization of H3N6 avian influenza virus isolated from a wild white pelican in Zambia

  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.
• MGL/CD301を介するエボラウイルス感染増強に関わる表層糖蛋白質の構造的特徴(Structural features of Ebola viral envelope glycoproteins responsible for the enhancement of infection through the interaction with MGL/CD301)

  宇佐美 克明, 松野 啓太, 五十嵐 学, 伝田 香里, 高田 礼人, 入村 達郎

  日本生化学会大会プログラム・講演要旨集 82回 4T20p - 16 2009/09
• Cross-Protective Potential of a Novel Monoclonal Antibody Directed against Antigenic Site B of the Hemagglutinin of Influenza A Viruses

  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.
• Cross-Glade protective immunity of H5N1 influenza vaccines in a mouse model

  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.
• Antigenic and genetic analysis of H5 influenza viruses isolated from water birds for the purpose of vaccine use

  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.
• H2N5 influenza virus isolates from terns in Australia: genetic reassortants between those of the Eurasian and American lineages

  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.
• Genetically destined potentials for N-linked glycosylation of influenza virus hemagglutinin

  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.
• Epitopes required for antibody-dependent enhancement of Ebola virus infection

  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.
• Rapid and simple detection of Ebola virus by reverse transcription-loop-mediated isothermal amplification

  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.
• Changes in Cellular Composition and Induction of Cell Death in the Spleen Are Correlated to the Disease Symptoms in Response to Influenza A Virus

  Sato Kayoko, Fujikura Daisuke, Takada Ayato, Kida Hiroshi, Miyazaki Tadaaki

  JOURNAL OF IMMUNOLOGY 178 0022-1767 2007/04/01 [Refereed][Not invited]
  
• Contributions of two nuclear localization signals of influenza A virus nucleoprotein to viral replication

  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.
• Protective efficacy of neutralizing antibodies against Ebola virus infection

  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.
• Bovine and water buffalo Mx2 genes: Polymorphism and antiviral activity

  H. A.E. Babiker, Y. Nakatsu, K. Yamada, A. Yoneda, A. Takada, J. Ueda, H. Hata, T. Watanabe

  Immunogenetics 59 (1) 59 - 67 0093-7711 2007/01 [Refereed][Not invited]
   

  Millennia-long selective pressure of single-strand RNA viruses on the bovine Mx locus has increased the advantages of using the bovine Mx protein to evaluate the ultimate significance of the antiviral role of Mx proteins. The conclusions of research based only on the bovine Mx1 protein showed the need for comprehensive studies that demonstrate the role of all isoforms, individually or together, especially in the presence of a second isoform, the bovine Mx2 gene. This study provides information about bovine and water buffalo Mx2 genes, as well as their allelic polymorphism and basic antiviral potential. Observation of an Mx2 cDNA sequence (2,381 bp) obtained from 15 animals from 11 breeds using primers based on a previous sequence (NCBI accession no. AF335147) revealed several nucleotide substitutions, with eight different alleles and two amino acid exchanges: Gly to Ser at position 302 and Ile to Val at position 354, though the latter was found only in the NCBI database. A water buffalo Mx2 cDNA sequence was identified for the first time, revealing 46 nucleotide substitutions with 12 amino acid variations, in addition to a 9-bp insertion in the 5′ untranslated region UTR, compared with the bovine Mx2 cDNA. Transfected 3T3 cells expressing bovine Mx2 mRNAs coding Gly or Ser at position 302, water buffalo Mx2 mRNA, positive control bovine Mx1 mRNA-expressing cells, and negative control parental 3T3 were subjected to infection with recombinant vesicular stomatitis virus (VSVΔG*-G), as were empty pCI-neo vector-transfected cells. The positive control and all cells expressing Mx2 mRNAs displayed significantly higher levels of antiviral activity against VSVΔG*-G (P < 0.01) than did the negative controls. © 2006 Springer-Verlag.
• Tyro3 family-mediated cell entry of Ebola and Marburg viruses

  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.
• Assembly and budding of Ebolavirus

  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.
• Molecular determinants of Ebola virus virulence in mice

  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.
• Data Mining in Amino Acid Sequences of H3N2 Influenza Viruses Isolated during 1968 to 2006.

  Ito K, Igarashi M, Takada A

  Knowledge Media Technologies TU Ilmenau 21 154 - 158 1617-9048 2006/07 [Refereed][Not invited]
   

  The hemagglutinin (HA) of influenza viruses undergoes antigenic drift to escape from antibody-mediated immune pressure. In order to predict possible structural changes of the HA molecules in future, it is important to understand the patterns of amino acid mutations and structural changes in the past. We performed a retrospective and comprehensive analysis of structural changes in H3 hemagglutinins of human influenza viruses isolated during 1968 to 2006. Amino acid sequence data of more than 2000 strains have been collected from NCBI Influenza virus resources. Information theoretic analysis of the collected sequences revealed a number of simultaneous mutations of amino acids at two or more different positions (correlated mutations). We also calculated the net charge of the HA1 subunit, based on thenumber of charged amino acid residues, and found that the net charge increased linearly from 1968 to 1984 and, after then, has been saturated. This level of the net charge may be an upper limit for H3 HA to be functional. It is noted that "correlated mutations" with the conversion of acidic and basic amino acid residues between two different positions were frequently found after 1984, suggesting that these mutations contributed to counterbalancing effect to keep the net charge of the HA . These approaches may open the way to find the direction of future antigenic drift of influenza viruses.
• Disease constraints for utilization of the African buffalo (Syncerus caffer) on game ranches in Zambia

  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.
• The development and characterization of H5 influenza virus vaccines derived from a 2003 human isolate

  T Horimoto, A Takada, 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.
• Hierarchy among viral RNA (vRNA) segments in their role in vRNA incorporation into influenza A virions

  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.
• Detection of cell-cell fusion mediated by Ebola virus glycoproteins

  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.
• The amino-terminal residue of glycoprotein B is critical for neutralization of bovine herpesvirus 1

  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.
• Highly pathogenic H5N1 influenza virus causes coagulopathy in chickens

  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.
• Architecture of ribonucleoprotein complexes in influenza A virus particles

  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.
• Avian flu - Isolation of drug-resistant H5N1 virus

  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]
  
• Characterization of a human H5N1 influenza A virus isolated in 2003

  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.
• Ebola virus VP40 late domains are not essential for viral replication in cell culture

  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.
• Live bivalent vaccine for parainfluenza and influenza virus infections

  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.
• Importance of both the coding and the segment-specific noncoding regions of the influenza a virus NS segment for its efficient incorporation into virions

  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.
• Enhanced virulence of influenza A viruses with the haemagglutinin of the 1918 pandemic virus

  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).
• A protective immune response in mice to viral components other than hemagglutinin in a live influenza A virus vaccine model

  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.
• Genetic polymorphisms and antiviral activity in the bovine MX1 gene

  Y. Nakatsu, K. Yamada, J. Ueda, A. Onogi, G. P. Ables, M. Nishibori, H. Hata, A. Takada, K. Sawai, Y. Tanabe, M. Morita, M. Daikohara, T. Watanabe

  Animal Genetics 35 (3) 182 - 187 0268-9146 2004/06 [Refereed][Not invited]
   

  Bovine MX1 cDNAs consisting of 2280 bp from 11 animals of five breeds and from a cultured cell line were sequenced and compared with previously reported data. Ten nucleotide substitutions were synonymous mutations, and a single nucleotide substitution at 458 resulted in an amino acid exchange of Ile (ATT) and Met (ATG). A 13-bp deletion-insertion mutation was also found in the 3′-UTR. Based on the nucleotide substitutions found in this study, bovine MX1 cDNA was classified into 11 genotypes. A phylogenetic tree of the 11 genotypes suggested that the genotypes observed in Brahman were a great genetic distance from other genotypes. An 18-bp deletion-insertion variation at position 171 was found to be the result of alternative splicing. The 18-bp deletion-insertion is located at the boundary between exon 3 and intron 3. Permanently transfected 3T3 cell lines expressing bovine MX1 mRNA were established to analyse the antiviral potential against VSV ΔG*-G infection. Transfected cell clones expressing bovine MX1 mRNA showed a significantly smaller number of cells infected with VSV ΔG*-G compared with the control cells. These results indicate that the bovine MX1 protein has potent antiviral activity.
• Native antiviral specificity of chicken Mx protein depends on amino acid variation at position 631

  J. H. Ko, A. Takada, T. Mitsuhashi, T. Agui, T. Watanabe

  Animal Genetics 35 (2) 119 - 122 0268-9146 2004/04 [Refereed][Not invited]
   

  An attempt was made to determine whether amino acid variation at position 631 in the chicken Mx protein definitely influences antiviral specificity, using an artificial mutation technique by which a single amino acid was reciprocally substituted between Ser (AGT) and Asn (AAT) at position 631 of the negative and positive chicken Mx, respectively. Using permanently transfected 3T3 cell lines, the antiviral potential of chicken Mx against vesicular stomatitis virus infection was analysed. The results indicated that the phenotype of antiviral activity depends on the amino acid difference at position 631 that is, the genotype coding Asn at position 631 corresponds to the positive antiviral phenotype, and the genotype coding Ser corresponds to the negative phenotype. The present study has confirmed that the antiviral specificity of chicken Mx protein is determined by an amino acid substitution at the carboxy terminus.
• A型インフルエンザウイルスPB2遺伝子分節のパッケージングと粒子形成に必要な領域

  村本 裕紀子, 高田 礼人, 藤井 健, 堀本 研子, 渡辺 真治, 喜田 宏, 河岡 義裕

  日本獣医学会学術集会講演要旨集 (公社)日本獣医学会 137回 91 - 91 1347-8621 2004/03
• Human macrophage C-type lectin specific for galactose and N-acetylgalactosamine promotes filovirus entry

  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.
• Biological significance of the U residue at the -3 position of the mRNA sequences of influenza A viral segments PB1 and NA

  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.
• Production of novel Ebola virus-like particles from cDNAs: an alternative to Ebola virus generation by reverse genetics

  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.
• Induction of interferon-inducible protein-10 and monokine induced by interferon-gamma from human endothelial cells infected with Influenza A virus

  N Ishiguro, A Takada, M Yoshioka, Ma, X, H Kikuta, H Kida, K Kobayashi

  ARCHIVES OF VIROLOGY 149 (1) 17 - 34 0304-8608 2004/01 [Refereed][Not invited]
   

  Primary human umbilical vein endothelial cells (HUVECs) were infected with Influenza virus A/Aichi/2/68 (H3N2) in order to determine the role of endothelial cells in mediating inflammation induced upon virus infection. Structural proteins of the virus and mRNA of the M2 protein were detected in the infected cells, indicating that virus infection had occurred in HUVECs. The Influenza A virus-infected HUVECs showed elevated levels of gene expression of interferon (IFN)-inducible protein (IP)-10 and monokine induced by IFN-gamma (Mig), while heat-, formalin- and diethyl ether-inactivated viruses did not enhance the IP-10 and Mig gene expression. The results thus indicate that infection of live Influenza A virus is responsible for elevation of IP-10 and Mig gene expression. The elevation of IP-10 and Mig gene expression in infected HUVECs was not accompanied by the elevation of IFN-gamma gene expression, indicating that the elevation of IP-10 and Mig gene expression was independent of the IFN-gamma pathway.
• Intranasal immunization with formalin-inactivated virus vaccine induces a broad spectrum of heterosubtypic immunity against influenza virus infection in mice

  A Takada, S Matsushita, A Ninomiya, Y Kawaoka, H Kida

  OPTIONS FOR THE CONTROL OF INFLUENZA V 1263 618 - 621 0531-5131 2004 [Refereed][Not invited]
   

  It has been known that the influenza A virus infection induces a cross-protective immunity against infection by viruses with different subtypes of viral envelope proteins, hemagglutinin and neuraminidase. 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 ethersplit 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) 2004 Elsevier B.V. All rights reserved.
• Potential for use of influenza A viruses with chimeric (type A/B) hemagglutinins as vaccines.

  Horimoto T, Takada A, Iwatsuki-Horimoto K, Hatta M, Goto H, Kawaoka Y

  Options for the Control of Influenza V ICS1263 622 - 625 2004 [Refereed][Not invited]
  
• Preparation of a panel of avian influenza viruses of different subtypes for vaccine strains against future pandemics.

  Sakoda Y, Okazaki K, Takada A, Ito Y, Tamai K, Okamatsu M, Ito T, Shortridge KF, Webster RG, Kida H

  Options for the Control of Influenza V, International congress series 1263 674 - 677 2004 [Refereed][Not invited]
  
• Persistence of viral RNA segments in the central nervous system of mice after recovery from acute influenza A virus infection

  Park CH, Matsuda K, Sunden Y, Ninomiya A, Takada A, Ito H, Kimura T, Ochiai K, Kida H, Umemura T

  Veterinary Microbiology 97 259 - 268 2003/12 [Refereed][Not invited]
  
• Neurotropism of the 1997 Hong Kong H5N1 influenza virus in mice

  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.
• Intranasal immunization with formalin-inactivated virus vaccine induces a broad spectrum of heterosubtypic immunity against influenza A virus infection in mice

  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.
• Antibody-dependent enhancement of Ebola virus infection

  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.
• Generation of influenza A viruses with chimeric (type A/B) hemagglutinins

  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.
• 強毒インフルエンザウイルスは血液凝固不全を引き起こす

  村本 裕紀子, 尾崎 弘一, 高田 礼人, 朴 天鎬, 寸田 祐嗣, 梅村 孝司, 河岡 義裕, 喜田 宏

  日本獣医学会学術集会講演要旨集 (公社)日本獣医学会 135回 128 - 128 1347-8621 2003/03
• Identification of protective epitopes on Ebola virus glycoprotein at the single amino acid level by using recombinant vesicular stomatitis viruses

  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.
• Seroepidemiological evidence of avian H4, H5, and H9 influenza A virus transmission to pigs in southeastern China

  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.
• Intranasal administration of a synthetic peptide vaccine encapsulated in liposome together with an anti-CD40 antibody induces protective immunity against influenza A virus in mice

  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.
• Ebola virus VP40 drives the formation of virus-like filamentous particles along with GP

  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.
• Polymorphisms and the differential antiviral activity of the chicken Mx gene

  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.
• The invasion routes of neurovirulent A Hong Kong 483/97 (H5N1) influenza virus into the central nervous system after respiratory infection in mice

  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.
• The loss of enzyme activities by a single amino acid substitution of a newly cloned rabbit CYP2D isozyme, CYP2D24

  Mayumi Ishizuka, Yukio Yamamoto, Ayato Takada, Akio Kazusaka, Shoichi Fujita

  International Congress Series 1233 (C) 121 - 126 0531-5131 2002 [Refereed][Not invited]
   

  In the previous study, we cloned two novel cytochrome P450 (CYP) cDNAs (CYP2D23 and CYP2D24) from a rabbit liver cDNA library. CYP2D23 and CYP2D24 were heterogeneously expressed in 293T cells. CYP2D24 effectively catalyzed the oxidation of bufuralol and bunitrolol, the archetypal substrates of the CYP2D subfamily, while CYP2D23 exhibited catalytic activity only toward bufuralol. Furthermore, in this study, we isolated the variant of CYP2D24, in which 473threonine was substituted with valine, and mutant CYP2D24 forms, in which 76lysine was replaced by proline. We determined the catalytic activities of the CYP2D24 forms and found that the mutants showed no activity related to drug metabolism. The computer-assisted analysis of the primary sequences of the CYP2D24 variant and mutant forms suggested that proline substitution results in alterations in the secondary structure and hydrophobicity of beta-Sheet 1, which was closed to Helix A, B and SRS-1, and was the proposed substrate-binding sites of P450. © 2002 Elsevier Science B.V.
• Plasminogen-binding activity of neuraminidase determines the pathogenicity of influenza A virus

  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.
• Primary target cells of virulent strains of type A influenza virus in chicken embryos

  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.
• Infectivity-enhancing antibodies to ebola virus glycoprotein

  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.
• Ebola virus glycoprotein: Proteolytic processing, acylation, cell tropism, and detection of neutralizing 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.
• Mucosal vaccination against influenza: Protection of pigs immunized with inactivated virus and ether-split vaccine

  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.
• Mouse Mx2 protein inhibits hantavirus but not influenza virus replication

  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.
• Global surveillance of animal influenza for the control of future pandemics

  H Kida, K Okazaki, A Takada, H Ozaki, M Tashiro, DK Lvov, KF Shortridge, RG Webster

  OPTIONS FOR THE CONTROL OF INFLUENZA IV 1219 169 - 171 0531-5131 2001 [Refereed][Not invited]
   

  Influenza viruses of different subtypes were isolated from fecal samples of ducks in their nesting areas in Siberia in summer from 1996 to 1999. Phylogenetic analysis of the NP genes of the isolates in Siberia and Japan on their flyway of migration to the south indicate that influenza viruses perpetuated in ducks nesting in Siberia should have contributed genes in the emergence of the H5N1 virus in Hong Kong. Inactivated vaccine prepared from an avirulent H5N4 virus isolated from a migratory duck in Japan showed enough potency to induce protective immunity against the pathogenic H5N1 virus in mice. To expand surveillance for influenza viruses in migratory and domestic ducks and geese, chickens, quail, and pigs of the world and be informed about what influenza viruses are dominant in the animal reservoirs, we have started the "Programme of Excellence in Influenza". Influenza virus strains isolated from animal species in the surveillance should be thoroughly characterized, stored, and be provided for the use of diagnosis and vaccine preparation for the control of future pandemics. (C) 2001 Elsevier Science B.V. All rights reserved.
• Downregulation of beta 1 integrins by Ebola virus glycoprotein: Implication for virus entry

  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.
• Functional importance of the coiled-coil of the Ebola virus glycoprotein

  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.
• Functional importance of the coiled-coil of the Ebola virus glycoprotein

  S Watanabe, A Takada, T Watanabe, H Ito, H Kida, Y Kawaoka

  JOURNAL OF VIROLOGY 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.
• Recognition of N-glycolylneuraminic acid linked to galactose by the alpha 2,3 linkage is associated with intestinal replication of influenza A virus in ducks

  T Ito, Y Suzuki, T Suzuki, A Takada, 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.
• 26.マウスのインターフェロン誘導性抗ウイルス遺伝子Mxに関する研究

  陳 喜慶, 高田 礼人, 森松 組子, 荻野 倫子, 有川 二郎, 渡辺 智正

  日本疾患モデル学会記録 公益社団法人 日本実験動物学会 16 33 - 33 0918-8991 2000
• Avian influenza virus intranasally inoculated infects the central nervous system of mice through the general visceral afferent nerve

  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.
• Precursor genes of future pandemic influenza viruses are perpetuated in ducks nesting in Siberia

  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.
• Structure of the infected cell protein 0 gene of canine herpesvirus

  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.
• Transcriptional activation of JC virus (JCV) by human T-lymphotropic virus type I (HTLV-I) tax protein in human neuronal cell lines.

  Okada Y, Sawa H, Tanaka S, Takada A, Suzuki S, Hasegawa H, Umemura T, Fujisawa J-i, Tanaka Y, Hall WW, Nagashima K

  J Biol Chem 275 (22) 17016 - 17023 0021-9258 2000 [Refereed][Not invited]
   

  Okada Y, Sawa H*, Tanaka S, Takada A, Suzuki S, Hasegawa H, Umemura T, Fujisawa J-i, Tanaka Y, Hall WW, Nagashima1 K: Transcriptional activation of JC virus (JCV) by human T-lymphotropic virus type I (HTLV-I) tax protein in human neuronal cell lines. J Biol Chem, 275, 17016-17023, 2000 (* corresponding author)*
• 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 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.
• Induction of antibodies in mice by a recombinant baculovirus expressing pseudorabies virus glycoprotein B in mammalian cells

  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.
• Detection of canine herpesvirus DNA in the ganglionic neurons and the lymph node lymphocytes of latently infected dogs

  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.
• Detection of Canine Herpesvirus DNA in the Ganglionic Neurons and the Lymph Node Lymphocytes of Latently Infected Dogs

  Masahiro Miyoshi, Yuki Ishii, Mitsuyoshi Takiguchi, Ayato Takada, Jun Yasuda, Akira Hashimoto, Katsunori Okazaki, Hiroshi Kida

  Journal of Veterinary Medical Science 61 (4) 375 - 379 1782-155X 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.
• Detection of Canine Herpesvirus DNA in the Ganglionic Neurons and the Lymph Node Lymphocytes of Latently Infected Dogs

  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.
• Antigenic and genetic analyses of H5 influenza viruses isolated from ducks in Asia

  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.
• Structure of the immediate early gene of canine herpesvirus - Brief report

  M Miyoshi, M Takiguchi, J Yasuda, A Hashimoto, A Takada, K Okazaki, H Kida

  ARCHIVES OF VIROLOGY 144 (2) 407 - 420 0304-8608 1999 [Refereed][Not invited]
   

  The nucleotide sequence of the immediate early (IE) gene of canine herpesvirus was determined. This gene was located in the inverted repeat regions, encoding a polypeptide of 1,383 amino acids. The predicted amino acid sequence was most closely related to that of the feline herpesvirus 1 IE protein among those of other alphaherpesviruses. DNA binding and transcriptional activation domains were found in the IE protein. A spliced region of the IE gene transcript was determined in its 5' non-coding region.
• Characterization of avian H5N1 influenza viruses from poultry in Hong Kong

  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.
• Changes in H3 influenza A virus receptor specificity during replication in humans

  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.
• An attenuation mechanism of Newcastle disease vaccine strain TCND

  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.
• Cloning, tissue distribution, and functional expression of two novel rabbit cytochrome P450 isozymes, CYP2D23 and CYP2D24

  Yukio Yamamoto, Mayumi Ishizuka, Ayato Takada, Shoichi Fujita

  Journal of Biochemistry 124 (3) 503 - 508 0021-924X 1998 [Refereed][Not invited]
   

  We cloned two novel cytochrome P450 cDNAs (CYP2D23 and CYP2D24) from a rabbit liver cDNA library. The open-reading frames of these cDNAs encode proteins that are each composed of 500 amino acids. The amino acid sequence identity of CYP2D23 with CYP2D24 is 91.6%, and the homology of these two isozymes with other known mammalian CYPs in the CYP2D subfamily range from 64.9 to 79.8%. Using RT-PCR, we determined the distribution of these two isozymes in 9 major organs, including brain tissue sections. CYP2D23 mRNA was abundantly expressed in the liver and small intestine, but only slightly in the brain sections, whereas CYP2D24 mRNA was expressed in the liver, small intestine, and stomach, CYP2D23 and CYP2D24 were heterogeneously expressed in 293T cells. CYP2D24 effectively catalyzed the oxidation of bufuralol and bunitrolol, the archetypal substrates of the CYP2D subfamily, while CYP2D23 exhibited catalytic activity only toward bufuralol. The results of this first study on rabbit CYP2D isozymes indicate that CYP2D23 and CYP2D24 are functionally expressed in rabbits, and have different organ distributions and metabolic properties.
• A system for functional analysis of Ebola virus glycoprotein

  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.
• The cysteine residues of the M2 protein are not required for influenza A virus replication

  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.
• Protective immune response of chickens against Newcastle disease, induced by the intranasal vaccination with inactivated virus

  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.
• PERPETUATION OF INFLUENZA-A VIRUSES IN ALASKAN WATERFOWL RESERVOIRS

  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.
• INDUCTION OF PROTECTIVE ANTIBODY-RESPONSES AGAINST PSEUDORABIES VIRUS BY INTRANASAL VACCINATION WITH GLYCOPROTEIN-B IN MICE

  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.
• PROTECTION OF MICE AGAINST AUJESZKYS-DISEASE VIRUS-INFECTION BY INTRANASAL VACCINATION WITH INACTIVATED VIRUS

  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.
• Potentially virulent Newcastle disease viruses are maintained in migratory waterfowl populations.

  Takakuwa H, Ito T, Takada A, Okazaki K, Kida H

  Japanese Journal of Veterinary Research 45 207 - 215 [Refereed][Not invited]
  

MISC

• Genetic characterization of various herpesviruses in bats in Zambia

  播磨勇人, 邱永晋, 山岸潤也, 梶原将大, CHANGULA Katendi, 佐々木道仁, MWEENE Aaron S, 澤洋文, 石原加奈子, HANG’OMBE Bernard M, 高田礼人  日本ウイルス学会学術集会プログラム・予稿集(Web)  70th-  2023
• High Seroprevalence among Mangala Residents 14 Years after the Bas-Congo virus-associated acute hemorrhagic fever outbreak in DR. Congo

  MUNYEKU Yannick Bazitama, MUNYEKU Yannick Bazitama, MUNYEKU Yannick Bazitama, HATTORI Takanari, SAITO Takeshi, OKITALE Patient Talunda, OKITALE Patient Talunda, MIYAMOTO Hiroko, KAJIHARA Masahiro, MASUMU Justin, MASUMU Justin, MASUMU Justin, AHUKA Steve Mundeke, AHUKA Steve Mundeke, MUYEMBE Jean-Jacques Tamfum, MUYEMBE Jean-Jacques Tamfum, MAKIALA Sheila Mandanda, MAKIALA Sheila Mandanda, IGARASHI Manabu, TAKADA Ayato  日本ウイルス学会学術集会プログラム・予稿集(Web)  70th-  2023
• ザンビアのエジプトルーセットオオコウモリおよびマストミスから分離されたロタウイルスAの性状解析

  岸本麻衣, 岸本麻衣, 梶原将大, 高田礼人, 大園誠也, 鈴木忠樹, 伊藤直人, 大場靖子, 澤洋文, 澤洋文, 佐々木道仁  日本獣医学会学術集会講演要旨集  166th-  2023
• Role of Rab11-dependent endocytic pathway in Marburg virus particle formation

  山田健斗, 古山若呼, 服部貴成, 坂口美亜子, 高田礼人, 南保明日香  日本ウイルス学会学術集会プログラム・予稿集(Web)  70th-  2023
• ウイルス学者の独り言

  ウェンディ  2022/07  [Not refereed][Invited]
  
• ザンビアにおけるクリミア・コンゴ出血熱の疫学調査

  梶原将大, SIMUUNZA Martin, SAASA Ngonda, DAUTU George, 森亜紀奈, 邱永晋, 中尾亮, HANG’OMBE Bernard, HANG’OMBE Bernard, 澤洋文, 澤洋文, SIMULUNDU Edgar, 福士秀悦, 森川茂, 西條政幸, 有川二郎, 高田礼人, 高田礼人, 吉松組子  日本ウイルス学会学術集会プログラム・予稿集(Web)  69th-  2022
• T-cell immunoglobulin mucin1の一塩基変異がウイルスの細胞侵入へ与える影響

  服部貴成, 斎藤健, 高館佳弘, 笠島和, 梶原亜紀奈, 宮本洋子, 梶原将大, 五十嵐学, 高田礼人  日本ウイルス学会学術集会プログラム・予稿集(Web)  69th-  2022
• カモメ属の鳥種(Larus sp.)から検出された新型鳥類ロタウイルスA株の遺伝学的解析

  藤井祐至, 正谷達謄, 正谷達謄, 西山祥子, 岡島美鈴, 泉郁輝, 岡崎克則, 迫田義博, 高田礼人, 高田礼人, 小澤真, 杉山誠, 伊藤直人, 伊藤直人  日本ウイルス学会学術集会プログラム・予稿集(Web)  69th-  2022
• 今、君たちに伝えたいこと ウイルス学者の視点あれこれ

  髙田 礼人  道徳と特別行動  38-  (9)  58  -59  2021/11  [Not refereed][Invited]
  
• エボラウイルス増殖機構と病原性発現メカニズム

  創薬研究者がこれだけは知っておきたい最新のウイルス学  90  -103  2021/08  [Not refereed][Invited]
  
• エボラおよびマールブルグウイルス －予防・治療薬開発の現状－

  高田礼人  オベリスク  26-  (2)  18  -26  2021/06  [Not refereed][Invited]
  
• Characterization of a Nelson Bay orthoreovirus isolated from an Egyptian fruit bat in Zambia

  播磨勇人, 佐々木道仁, 大場靖子, 邱永晋, チャングラカテンディ, 梶原将大, シムルンドゥエドガー, 谷口怜, 高田礼人, 西條政幸, ハンゴンべバーナード, 澤洋文  日本ウイルス学会学術集会プログラム・予稿集(Web)  68th-  2021
• Single nucleotide polymorphisms in human angiotensin-converting enzyme 2 receptor and cell susceptibility to SARS-CoV-2

  HATTORI Takanari, SAITO Takeshi, TAKADATE Yoshihiro, OKUYA Kosuke, KASAJIMA Nodoka, KAJIHARA Akina, MIYAMOTO Hiroko, KAJIHARA Masahiro, TAKADA Ayato  日本ウイルス学会学術集会プログラム・予稿集(Web)  68th-  2021
• Inhibition of filovirus entry by cyclic peptides derived from the Niemann-Pick C1 loop structure

  高橋侑嗣, 高舘佳弘, 宮本洋子, 梶原将大, 五十嵐学, 高田礼人  日本ウイルス学会学術集会プログラム・予稿集(Web)  68th-  2021
• 海洋生物に含まれる抗新型コロナウイルス物質の探索

  住谷朋之丞, 辺浩美, 酒井隆一, 田中良和, 高田礼人  日本水産学会大会講演要旨集(CD-ROM)  2021-  2021
• ザンビアのブタにおける消化管ウイルスの疫学調査

  播磨勇人, 梶原将大, 佐々木道仁, SIMULUNDU Edgar, BWALYA Eugene, 邱永晋, 奥谷公亮, 大場靖子, HANG’OMBE Bernard M, MWEENE Aaron S, 高田礼人, 澤洋文  日本獣医学会学術集会講演要旨集  164th (CD-ROM)-  2021
• Investigation of an orthoreovirus isolated from a fruit bat in Zambia

  播磨勇人, 佐々木道仁, 大場靖子, QIU Yongjin, 梶原将大, 谷口怜, 高田礼人, 西條政幸, 澤洋文  日本分子生物学会年会プログラム・要旨集(Web)  44th-  2021
• Bat- and arthropod-borne viruses raising potential public health concern.

  梶原将大, 大場靖子, 高田礼人  Pharma Medica  39-  (1)  2021
• Viral pathogenicity

  高田礼人  実験医学  39-  (2)  2021
• Ebola virus requires a host scramblase for its efficient entry into target cells Ebola virus requires a host scramblase for its efficient entry into target cells

  南保明日香, 丸山隼輝, 今井正樹, 氏江美智子, 高田礼人, 大場雄介, 河岡義裕  日本女性科学者の会学術誌(Web)  20-  (1)  2020
• インタビュー エボラ出血熱の治療薬開発を目指して 人類史上、最も危険なウイルスに挑む

  タカダ アヤト  Harvard business review = Diamondハーバード・ビジネス・レビュー  44-  (8)  62  -73  2019/08
• 高病原性病原体による感染症対策―BSL‐4施設により変わる研究―医薬品開発 エボラワクチン

  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]
  
• シュードタイプ水胞性口炎ウイルスを用いたフィロウイルス感染代替動物モデル

  齋藤健, 磯野真央, 木田祐里恵, 丸山隼輝, 高田礼人  日本ウイルス学会学術集会プログラム・予稿集(Web)  67th-  2019
• エボラウイルスの細胞侵入阻害薬の探索

  磯野真央, 古山若呼, 黒田誠, 近藤達成, 五十嵐学, 梶原将大, 吉田玲子, マンズール ラシッド, 奥谷公亮, 宮本洋子, フェルドマン ハインツ, マルジ アンドレア, 堺谷政弘, 高田礼人  日本ウイルス学会学術集会プログラム・予稿集(Web)  67th-  2019
• A型インフルエンザウイルスに対する交差感染防御免疫への非中和IgA抗体の役割

  奥谷公亮, 吉田玲子, マンズール ラシッド, 齊藤慎二, 鈴木忠樹, 市居修, 東秀明, 高田礼人  日本ウイルス学会学術集会プログラム・予稿集(Web)  67th-  2019
• ザンビアのエジプトルーセットオオコウモリから検出されたマールブルグウイルスの遺伝学的解析

  梶原将大, CHANGULA Katendi, HANG’OMBE Bernard, 播磨勇人, 宮本洋子, 衛藤芳樹, 奥谷公亮, 磯野真央, 吉田玲子, 邱永晋, 森(梶原)亜紀奈, 大場靖子, 澤洋文, 澤洋文, 小川寛仁, SIMULUNDU Edgar, SQUARRE David, MUKONKA Victor, MWEENE Aaron, 高田礼人, 高田礼人  日本ウイルス学会学術集会プログラム・予稿集(Web)  67th-  2019
• ザンビアのダニで発見した新規フラビウイルスはフラビウイルス属の中で独特な系統を形成するNgoye virusに遺伝的に密接に関係している

  播磨勇人, 鳥居志保, 邱永晋, 梶原将大, 衛藤芳樹, 大場靖子, 江下優樹, ハンゴンベ バーナード, 好井健太朗, シムンザ マーティン, 高田礼人, 澤洋文  日本ウイルス学会学術集会プログラム・予稿集(Web)  67th-  2019
• 動物におけるA型インフルエンザウイルスの疫学調査

  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]
  
• 渡り鳥におけるロタウイルスAの分子疫学的研究

  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  161回-  396  -396  2018/08  [Not refereed][Not invited]
  
• エボラウイルス研究最前線

  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]
  
• 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  2018/03/25  [Not refereed][Not invited]
  
• Differential filovirus tropism controlled by the polymorphism of bat Niemann-Pick C1 proteins

  TAKADATE Yoshihiro, KONDOH Tatsunari, MARUYAMA Junki, MARUYAMA Junki, MANZOOR Rashid, KURODA Makoto, KURODA Makoto, OGAWA Hirohito, OGAWA Hirohito, SATO Masahiro, FURUYAMA Wakako, FURUYAMA Wakako, YOSHIDA Reiko, IGARASHI Manabu, TAKADA Ayato, TAKADA Ayato  日本ウイルス学会学術集会プログラム・抄録集  66th-  2018
• グローバル化・温暖化と感染症対策 II.各論 3.新興・再興感染症 3)エボラ出血熱

  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]
  
• 人獣共通感染症―インフルエンザとエボラ出血熱の疫学―

  高田礼人  日本内科学会雑誌  106-  (10)  2237‐2245  2017/10/10  [Not refereed][Not invited]
  
• エボラウイルス感染症研究における最近の知見

  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]
  
• 国内のコウモリから分離された2種類のアデノウイルスの性状解析

  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]
  
• 2.3.4.4高病原性鳥インフルエンザウイルスに対する新規モノクローナル抗体を用いた免疫クロマトグラフィーキットによるH5ヘマグルチニンの迅速かつ広範囲検出(Rapid and broad detection of H5 hemagglutinin by an immunochromatographic kit using novel monoclonal antibody against a 2.3.4.4 highly pathogenic avian influenza virus)

  Nguyen Thanh-Lam, 中石 和成, 本島 桂子, 大河原 彩子, 日尾野 隆大, 松野 啓太, 岡松 正敏, 高田 礼人, 喜田 宏, 迫田 義博  日本獣医学会学術集会講演要旨集  160回-  384  -384  2017/08  [Not refereed][Not invited]
  
• Rapid and broad detection of H5 hemagglutinin by an immunochromatographic kit using novel monoclonal antibody against a 2.3.4.4 highly pathogenic avian influenza virus(和訳中)

  Nguyen Thanh-Lam, 中石 和成, 本島 桂子, 大河原 彩子, 日尾野 隆大, 松野 啓太, 岡松 正敏, 高田 礼人, 喜田 宏, 迫田 義博  日本獣医学会学術集会講演要旨集  160回-  384  -384  2017/08  [Not refereed][Not invited]
  
• 私達の研究(170)エボラ出血熱に対する治療法および診断法の開発

  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]
  
• Ecology of Filoviruses and Transmission Routes to Humans

  高田 礼人  最新醫學 = The medical frontline  72-  (4)  534  -540  2017/04  [Not refereed][Not invited]
  
• 臨床ウイルス分野の海外協力 アフリカを舞台にしたフィロウイルス研究―自然宿主探索と診断法開発―

  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]
  
• 人獣共通感染症としてのインフルエンザ

  高田礼人  月刊臨床と研究  94-  (3)  384  2017/03/20  [Not refereed][Not invited]
  
• ウイルスの増殖・病態発現機構に関する最新の知見 エボラウイルスの増殖・病原性発現機構の最先端の知見

  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]
  
• 新興・再興感染症 IV.特論 エボラ出血熱の診断・治療法開発―モノクローナル抗体の活用―

  高田礼人  日本臨床  74-  (12)  2080‐2085  2016/12/01  [Not refereed][Not invited]
  
• ザンビア共和国に飛来するストローオオコウモリ(Eidolon helvum)から分離されたアデノウイルスの性状解析と分子疫学的解析

  小川寛人, 小川寛人, 小川寛人, 梶原将大, 直亨則, 丸山隼輝, 上野恵介, 石井秋宏, 石井秋宏, 藤倉大輔, HANG’OMBE Bernard, AARON Mweene, 山田雅夫, 東秀明, 東秀明, 澤洋文, 澤洋文, 高田礼人, 高田礼人  日本獣医学会学術集会講演要旨集  159th-  376  -376  2016/08/30  [Not refereed][Not invited]
  
• グローバル感染症最前線―NTDsの先へ Vol.3 エボラウイルス研究の現状と最新の知見

  高田礼人  医学のあゆみ  258-  (7/8)  803‐810  -810  2016/08/20  [Not refereed][Not invited]
  
• エボラ出血熱との戦いの総括 3.エボラ出血熱の予防・治療・診断法開発の現状

  古山若呼, 高田礼人  ウイルス  66-  (1)  63‐72  -72  2016/06  [Not refereed][Not invited]
   

  Ebolaviruses, members of the family Filoviridae, cause severe hemorrhagic fever in humans and nonhuman primates, with human case fatality rates of up to 90%. No effective prophylaxis or treatment for Ebola virus disease (EVD) is yet commercially available. During the latest outbreak of EVD in West Africa, several unapproved drugs were used for the treatment of patients. This outbreak has indeed accelerated efforts to develop antiviral strategies and some of the vaccine and drug candidates have undergone clinical trials. This article reviews previous researches and recent advances on the development of vaccine, therapeutics, and diagnostics for EVD.
• エボラウイルス研究の最前線

  高田礼人  感染症学雑誌  90-  125  2016/03/20  [Not refereed][Not invited]
  
• 北海道で分離された新規ダニ媒介性フレボウイルスの性状解析

  鳥居志保, 鳥居志保, 松野啓太, 松野啓太, 中尾亮, 邱永晋, 梶原将大, 直亨則, 村松美笑子, 海老原秀喜, 高田礼人, 高田礼人  日本分子生物学会年会プログラム・要旨集(Web)  39th-  ROMBUNNO.1LBA‐083 (WEB ONLY)  2016  [Not refereed][Not invited]
  
• エボラウイルスの診断・治療法開発の最前線

  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]
  
• RT‐PCR法による新規ダニ媒介性フレボウイルス遺伝子の探索

  松野啓太, 松野啓太, 下田宙, 鳥居志保, 邱永晋, 中尾亮, 梶原将大, 岡松正敏, 迫田義博, 迫田義博, 奥村敦, 奥村敦, 高田礼人, 高田礼人, 前田健, 海老原秀喜  日本獣医学会学術集会講演要旨集  158th-  344  -344  2015/08/30  [Not refereed][Not invited]
  
• 新興・再興感染症と感染症対策 A.ウイルス フィロウイルス感染症

  KAJIHARA MASAHIRO, TAKADA AYATO, TAKADA AYATO, TAKADA AYATO  生体の科学  66-  (4)  296  -300  2015/08/15  [Not refereed][Not invited]
  
• 渡り鳥から検出された新型ロタウイルスAの遺伝学的解析

  藤井 祐至, 岡寺 康太, 三竹 博道, 伊藤 直人, 岡田 和真, 中川 賢人, 岡崎 克則, 迫田 義博, 高田 礼人, 小澤 真, 正谷 達謄, 杉山 誠  日本獣医学会学術集会講演要旨集  158回-  371  -371  2015/08  [Not refereed][Not invited]
  
• Filovirus disease

  梶原 将大, 高田 礼人  生体の科学  66-  (4)  296  -300  2015/07  [Not refereed][Not invited]
  
• エボラ出血熱 3.エボラウイルスに対するワクチンおよび治療法

  TAKADA AYATO  ウイルス  65-  (1)  61  -70  2015/06  [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.
• エボラ出血熱 適切な対応で感染は防げる 風邪のような症状からはじまり免疫細胞を破壊

  TAKADA AYATO  いつでも元気  (283)  16  -19  2015/05/01  [Not refereed][Not invited]
  
• Filovirology

  高田 礼人  医学のあゆみ  253-  (1)  5  -11  2015/04/04  [Not refereed][Not invited]
  
• いま注目のウイルス感染症【エボラ出血熱とマーブルグ病】フィロウイルスのウイルス学

  TAKADA AYATO  医学のあゆみ  253-  (1)  5  -11  2015/04/04  [Not refereed][Invited]
  
• 最恐ウイルスにどう立ち向かうか―エボラとマールブルグウイルス―

  TAKADA AYATO  アニムス  20-  (2)  19  -22  2015/04/01  [Not refereed][Invited]
  
• エボラ出血熱にどう立ち向かうか エボラ出血熱とはどのような病気か

  TAKADA AYATO  生活と環境  60-  (3)  11  -15  2015/03/01  [Not refereed][Invited]
  
• エボラ出血熱とはどのような病気か (特集 エボラ出血熱にどう立ち向かうか)

  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]
  
• Ebola vaccine and treatment

  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.
• The clinical potential of passive immunization with therapeutic antibodies: focus on highly pathogenic avian influenza virus infection

  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.
• フィロウイルスの細胞侵入におけるTIM‐1とNPC1相互作用の解析

  黒田誠, 藤倉大輔, 南保明日香, 野依修, 梶原将大, 丸山隼輝, 宮本洋子, 吉田玲子, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  62nd-  174  2014/10/31  [Not refereed][Not invited]
  
• H5N1インフルエンザウイルスに対する中和抗体を用いた受動免疫の感染防御効果

  吉田玲子, 伊藤靖, 七戸新太郎, 小笠原一誠, 日尾野隆大, 岡松正敏, 迫田義博, 喜田宏, LE Mai Quynh, 河岡義裕, 五十嵐学, 鈴木定彦, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  62nd-  234  2014/10/31  [Not refereed][Not invited]
  
• エボラ出血熱

  TAKADA AYATO  現代化学  (523)  16  -18  2014/10/01  [Not refereed][Invited]
  
• 科学の目 エボラ出血熱

  髙田 礼人  現代化学  (523)  16  -18  2014/10  [Not refereed][Invited]
  
• Ebola and Marburg virus diseases in Africa: Increased risk of outbreaks in previously unaffected areas?

  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.
• 北海道産マダニからの新規フレボウイルスの検出

  NAKAO RYO, KAJIWARA MASAHIRO, KYU EISHIN, MORI AKINA, NAO NAGANORI, MURAMATSU MIEKO, YOSHII KENTARO, KARIWA HIROAKI, SAWA HIROFUMI, SUGIMOTO CHIHIRO, TAKADA AYATO  日本獣医学会学術集会講演要旨集  157th-  451  2014/08/11  [Not refereed][Not invited]
  
• 次世代型感染症ワクチン エボラ出血熱ワクチン・炭疽ワクチン

  ONISHI NAOMI, HIGASHI HIDEAKI, TAKADA AYATO  最新医学  69-  (4)  865  -871  2014/04  [Not refereed][Not invited]
  
• Ebola Hemorrhagic Fever Vaccine ・ Anthrax Vaccine

  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]
  
• A型インフルエンザウイルスヘマグルチニン特異的IgA抗体の亜型間交差抗ウイルス活性

  村松美笑子, 吉田玲子, 横山文香, 宮本洋子, 梶原将大, 丸山隼輝, 直享則, MANZOOR Rashid, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  62nd-  2014
• エボラウイルスの抗体依存性感染増強現象におけるFcレセプターを介したシグナル伝達経路の解析

  古山若呼, 黒田誠, 丸山隼輝, 宮本洋子, 吉田玲子, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  62nd-  2014
• マダニ媒介性フレボウイルスの検出

  中尾亮, 梶原将大, 松野啓太, 邱永晋, 森亜紀奈, 海老原秀喜, 高田礼人, 杉本千尋  大原綜合病院年報  53-  69  2013/12/15  [Not refereed][Not invited]
  
• インフルエンザに立ち向かう 人獣共通感染症としてのインフルエンザ

  ISODA NORIKAZU, TAKADA AYATO  Mebio  30-  (12)  32  -41  2013/12/10  [Not refereed][Not invited]
  
• 感染症“死の病原体”に前線で挑むサイエンス フィロウイルスの生態

  KAJIHARA MASAHIRO, OGAWA HIROHITO, TAKADA AYATO  実験医学  31-  (19)  3054  -3060  2013/12/01  [Not refereed][Invited]
  
• フィロウイルス感受性に及ぼすTIM‐1多型の影響

  黒田誠, 藤倉大輔, 野依修, 中山絵里, 梶原将大, 丸山隼輝, 宮本洋子, 吉田玲子, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  61st-  163  2013/10/29  [Not refereed][Not invited]
  
• 2010年に稚内で分離された2株の高病原性鳥インフルエンザウイルスの病原性比較

  直亨則, 梶原将大, 丸山隼輝, 木村享史, MANZOOR Rashid, 村松美笑子, 岡松正敏, 宮本洋子, 吉田玲子, 迫田義博, 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  61st-  150  2013/10/29  [Not refereed][Not invited]
  
• Lloviu virusの表面糖タンパク質GPの性状解析

  丸山隼輝, 宮本洋子, 吉田玲子, 前田健, 小川寛人, 迫田義博, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  61st-  164  2013/10/29  [Not refereed][Not invited]
  
• 計算科学手法を用いたインフルエンザウイルスヘマグルチニンと亜型間交差反応性抗体との分子間相互作用の解析

  五十嵐学, 吉田玲子, 関嶋政和, 常盤広明, 喜田宏, 伊藤公人, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  61st-  239  2013/10/29  [Not refereed][Not invited]
  
• Do therapeutic antibodies hold the key to an effective treatment for Ebola hemorrhagic fever?

  Ayato Takada  Immunotherapy  5-  (5)  441  -443  2013/05  [Refereed][Invited]
  
• 2010‐11シーズンに国内の野鳥斃死体から分離されたH5N1高病原性鳥インフルエンザウイルスのカモ科およびサギ科鳥類に対する病原性と伝播能の解析

  曽田公輔, 笛吹達史, 宇野有紀子, 永井泰子, 尾崎弘一, 伊藤啓史, 山本直樹, 田村友和, 日尾野隆大, 岡松正敏, 迫田義博, 高田礼人, 村瀬敏之, 山口剛士, 伊藤壽啓  日本獣医学会学術集会講演要旨集  155th-  83  2013/03/04  [Not refereed][Not invited]
  
• フィロウイルス糖蛋白質によるアポトーシスシグナル阻害作用

  野依修, 中山絵里, 丸山隼輝, 吉田玲子, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  61st-  2013
• バイオテロに使用される可能性のある病原体等の新規検出法と標準化に関する研究 新興アレナウイルスに対応可能なウイルス抗原検出法の開発に関する研究

  森川茂, 福士秀悦, 吉河智城, 谷英樹, 谷口怜, 伊波興一朗, 下島昌幸, 西條政幸, 高田礼人  バイオテロに使用される可能性のある病原体等の新規検出法と標準化に関する研究 平成24年度 総括・分担研究報告書  19  -25  2013  [Not refereed][Not invited]
  
• Negative Strand RNA Virusのウイルス学 3.フィロウイルス

  TAKADA AYATO  ウイルス  62-  (2)  197  -208  2012/12  [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.
• Filoviruses

  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.
• フィロウイルス糖蛋白質による宿主細胞表面分子の立体的遮蔽現象の解析

  野依修, 松野啓太, 梶原将大, 中山絵里, 五十嵐学, 磯田典和, 吉田玲子, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  60th-  198  2012/10/31  [Not refereed][Not invited]
  
• 近隣剪定法によるウイルス遺伝子配列のリサンプリング

  米澤弘毅, 五十嵐学, 高田礼人, 伊藤公人  日本ウイルス学会学術集会プログラム・抄録集  60th-  321  2012/10/31  [Not refereed][Not invited]
  
• アカゲザルモデルを用いたエボラ出血熱に対する中和モノクローナル抗体の防御効果

  吉田玲子, MARZI Andrea, 宮本洋子, 石島麻理, 鈴木定彦, 五十嵐学, 中山絵里, 黒田誠, FELDMANN Heinz, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  60th-  199  2012/10/31  [Not refereed][Not invited]
  
• H3N2インフルエンザウイルスHAへの糖鎖付加と周辺アミノ酸残基にかかる選択圧の経詩的変化

  五十嵐学, 中尾亮, 喜田宏, 喜田宏, 高田礼人, 伊藤公人  日本ウイルス学会学術集会プログラム・抄録集  60th-  318  2012/10/31  [Not refereed][Not invited]
  
• Protecting Against the Zombie Apocalypse

  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)  2012/10  [Not refereed][Not invited]
  
• いきものの不思議 ウイルスはどうやって生き延びているのか? : インフルエンザウイルスの存続様式と進化

  伊藤 公人, 高田 礼人  遺伝 : 生物の科学  66-  (4)  358  -364  2012/07  [Not refereed][Invited]
  
• ウイルスはどうやって生き延びているのか?—インフルエンザウイルスの存続様式と進化

  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]
  
• 新生児に致死的肺炎を引き起こしたアデノウイルスの解析

  直亨則, 津曲俊太郎, 吉岡幹郎, 梶原将大, 鈴木昭, 高橋豊, 高田礼人  日本ウイルス学会北海道支部夏季シンポジウム講演抄録  46th-  10  2012  [Not refereed][Not invited]
  
• Ebola and Marburg Hemorrhagic Fever

  髙田 礼人  最新医学  66-  (12)  2668  -2675  2011/12  [Not refereed][Invited]
  
• 人獣共通感染症の克服に向けて エボラ・マールブルグ出血熱

  高田礼人  最新医学  66-  (12)  2668-2675  -2675  2011/12  [Not refereed][Invited]
  
• 2009年のウズラ由来H7N6亜型高病原性鳥インフルエンザウイルスの感染経路の推定

  内田 裕子, 金平 克史, 真瀬 昌司, 竹前 喜洋, 渡辺 千晶, 笛吹 達史, 藤本 佳万, 伊藤 壽啓, 五十嵐 学, 伊藤 公人, 高田 礼人, 迫田 義博, 岡松 正敏, 山本 祐, 中村 菊保, 喜田 宏, 廣本 靖明, 津田 知幸, 西藤 岳彦  動物衛生研究成果情報  (10)  15  -16  2011/09  [Not refereed][Not invited]
  
• トップランナーに聞く--最先端の医療に挑む若手研究者への直撃インタビュー(2)人獣共通感染症の克服を目指して

  高田 礼人  最新医学  66-  (2)  304  -307  2011/02  [Not refereed][Invited]
  
• Structural characteristic of Ebola viral GP1 involved in attachment and infection.

  FUJIHIRA HARUHIKO, USAMI KATSUAKI, DENDA KAORI, YAMADA KEITA, MATSUNO KEITA, SHINOHARA YASURO, TAKADA AYATO, KAKEHI KAZUAKI, IRIMURA TATSURO  生化学  84回-  ROMBUNNO.3T8A-14  -14  2011  [Not refereed][Not invited]
  
• Ebola and Marburg viruses

  Eri Nakayama, Ayato Takada  Journal of Disaster Research  6-  (4)  381  -389  2011  [Refereed][Invited]
   

  Ebola and Marburg viruses, members of the filovirus family, cause severe hemorrhagic fever in human and nonhuman primates and are classified as biosafety level 4 agents. No effective filovirus-specific prophylaxis or treatment is yet commercially available. Filovirus species vary genetically, with one in the Marburg virus group and five in the Ebola virus group. Epidemiological efforts to prevent outbreaks lie mainly in identifying natural animal reservoirs. Increasingly frequent outbreaks in Africa and concerns about bioterrorism and imported cases in nonendemic areas point to the importance of public health in two ways - finding strategies to control disease outbreak and developing effective vaccines and drugs.
• Strain-Dependent Glycosylation of Ebola Viral Envelope Glycoproteins Determines their Infectivity

  Katsuaki Usami, Haruhiko Fujihira, Kaori Denda-Nagai, Keita Yamada, Keita Matsuno, Ayato Takada, Kazuaki Kakehi, Tatsuro Irimura  GLYCOBIOLOGY  20-  (11)  1529  -1530  2010/11  [Not refereed][Not invited]
  
• フィロウイルスの宿主域

  TAKADA AYATO  感染症  236-  (6)  220  -225  2010/11  [Not refereed][Invited]
  
• H3N2インフルエンザウイルスHAに付加された糖鎖は周辺エピトープを覆い隠していたか?〜糖鎖付加部位周辺アミノ酸残基の多様性変化の解析〜

  五十嵐学, 高田礼人, 喜田宏, 喜田宏, 喜田宏, 伊藤公人, 伊藤公人  日本ウイルス学会学術集会プログラム・抄録集  58th-  227  2010/10/15  [Not refereed][Not invited]
  
• フィロウイルス種特異抗体検出ELISA法の開発

  中山絵里, 横山文香, 宮本洋子, 岸田典子, 松野啓太, 五十嵐学, MARZI Andrea, FELDMANN Heinz, 伊藤公人, 高田礼人  日本ウイルス学会学術集会プログラム・抄録集  58th-  286  2010/10/15  [Not refereed][Not invited]
  
• パンデミックインフルエンザAウイルス(H1N1)ヘマグルチニンに対するモノクローナル抗体の性状解析

  吉田玲子, 苫米地大輔, 五十嵐学, 宮本洋子, 加瀬哲男, 喜田宏, 喜田宏, 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  58th-  219  2010/10/15  [Not refereed][Not invited]
  
• Immunity against influenza virus infection

  高田 礼人  Japanese journal of clinical medicine  68-  (9)  1625  -1630  2010/09  [Not refereed][Invited]
  
• パンデミックインフルエンザ インフルエンザウイルス研究の進歩 インフルエンザウイルスに対する免疫応答・感染防御機構

  高田礼人  日本臨床  68-  (9)  1625-1630  -1630  2010/09  [Not refereed][Invited]
  
• 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

  高田礼人, 吉田玲子, 五十嵐学, 伊藤公人  日本薬学会年会要旨集  130th-  (1)  339  2010/03/05  [Not refereed][Not invited]
  
• 種の壁を越えるウイルス感染症—EpidemiologyとEpizootiology フィロウイルス

  高田礼人  臨床と微生物  37-  (2)  125-131  -131  2010/03  [Not refereed][Invited]
  
• 人獣共通感染症としてのインフルエンザ

  TAKADA AYATO  Campus Health  47-  (1)  67  -71  2010/02  [Not refereed][Not invited]
  
• 新型インフルエンザ教養講座 Q&Aインフルエンザウイルスの基礎

  TAKADA AYATO  化学  64-  (11)  18  -24  2009/11  [Not refereed][Invited]
  
• 新型インフルエンザウイルス 4.インフルエンザウイルスの抗原変異予測とワクチン

  伊藤公人, 高田礼人  Virus Rep  6-  (2)  60-68  -68  2009/11  [Not refereed][Invited]
  
• Q&A インフルエンザウイルスの基礎 (特集 新型インフルエンザ教養講座--化学者として知っておくべき基礎知識)

  高田 礼人  化学  64-  (11)  18  -24  2009/11  [Not refereed][Not invited]
  
• Special Interview インフルエンザウイルスの抗体ライブラリー構築とは何か?

  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]
  
• 対談 荒俣宏の〈万物に叡智あり〉 ゲスト 北海道大学人獣共通感染症リサーチセンター副センター長 高田礼人さん 変わり者だけが勝ち残る 世にも奇妙なウイルスの生存戦略を見よ

  荒俣 宏, 高田 礼人  フォーレ  0-  (86)  32  -37  2009/11  [Not refereed][Invited]
  
• 愛知県でウズラから分離されたH7N6亜型鳥インフルエンザウイルスの遺伝子解析

  内田裕子, 真瀬昌司, 竹前喜洋, 廣本靖明, 五十嵐学, 伊藤公人, 高田礼人, 西藤岳彦, 津田知幸, 山口成夫  日本ウイルス学会学術集会プログラム・抄録集  57th-  346  2009/10/01  [Not refereed][Not invited]
  
• ホモロジーモデリング法による新型H1N1インフルエンザウイルスのヘマグルチニンの抗原構造の解析

  五十嵐学, 伊藤公人, 吉田玲子, 喜田宏, 喜田宏, 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  57th-  180  2009/10/01  [Not refereed][Not invited]
  
• インフルエンザウイルスのヘマグルチニンのMDS解析と変異予測への応用

  伊藤公人, 五十嵐学, 村上悌治, 喜田宏, 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  57th-  181  2009/10/01  [Not refereed][Not invited]
  
• Characterization of H3N6 avian influenza virus isolated from a wild white pelican in Zambia

  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.
• ウイルスと宿主の相互作用 フィロウイルスの感染機構と宿主因子

  高田礼人  蛋白質 核酸 酵素  54-  (8)  913-919  -919  2009/06/10  [Not refereed][Invited]
  
• Filovirus infection and host factors

  高田 礼人  Protein, nucleic acid and enzyme  54-  (8)  913  -919  2009/06  [Not refereed][Invited]
  
• ウイルスハンター--ウイルスの生きざまを見つめて (特集 ウイルス--その多様性と謎に満ちた存在)

  高田 礼人  ミルシル  2-  (3)  16  -19  2009/05  [Not refereed][Invited]
  
• 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 エボラウイルスの診断法と抗ウイルス薬の検討

  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]
  
• Prediction of antigenic change of influenza virus

  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]
  
• Ebola hemorrhagic fever

  高田 礼人  綜合臨床  57-  (11)  2673  -2679  2008/11  [Not refereed][Invited]
  
• 変貌する感染症—人類の備えは十分か?エボラ出血熱

  高田礼人  綜合臨床  57-  (11)  2673-2679  -2679  2008/11/01  [Not refereed][Not invited]
  
• マールブルグウイルスの病原性とC型レクチン介在性細胞侵入効率との相関

  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]
  
• H5N1不活化インフルエンザワクチンのクレード間交差防御免疫能の解析

  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]
  
• インフルエンザウイルスHA遺伝子の塩基配列上に潜在する糖鎖付加ポテンシャルの解析

  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]
  
• 2008年北海道で発見された斃死オオハクチョウから分離したH5N1高病原性鳥インフルエンザウイルスの性状

  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]
  
• H5N1インフルエンザウイルスのPB2に会合する宿主細胞因子の同定とその機能解析

  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]
  
• 2008年北海道で発見された斃死オオハクチョウから分離したH5N1高病原性鳥インフルエンザウイルスの性状

  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]
  
• 人獣共通感染症の先回り予防戦略

  高田礼人  補体シンポジウム講演集  45th-  51  2008/07  [Not refereed][Not invited]
  
• インフルエンザウイルスHA遺伝子の塩基配列上に潜在する糖鎖付加ポテンシャルの解析

  五十嵐学, 伊藤公人, 喜田宏, 喜田宏, 喜田宏, 高田礼人  日本ウイルス学会北海道支部夏季シンポジウム講演抄録  42nd-  12  2008  [Not refereed][Not invited]
  
• 第2章 空気の通り道における感染・免疫機構 6.インフルエンザに対する感染防御免疫

  YOSHIDA REIKO, TAKADA AYATO  実験医学  25-  (20)  3176  -3182  2007/12  [Not refereed][Invited]
  
• Epitopes required for antibody-dependent enhancement of Ebola virus infection

  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.
• Antibody therapy as a future treatment option for Ebola virus infection

  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.
• 動物インフルエンザのグローバルサーベイランスと全ての亜型ウイルスライブラリーの構築

  梶原将大, 梶原将大, 迫田義博, 迫田義博, 伊藤壽啓, 高田礼人, 岸田典子, 五十嵐学, 磯田典和, 磯田典和, 曽田公輔, 曽田公輔, 喜田宏, 喜田宏, 喜田宏  日本ウイルス学会学術集会プログラム・抄録集  55th-  216  2007/10/01  [Not refereed][Not invited]
  
• 動物インフルエンザのグローバルサーベイランスと全ての亜型ウイルスライブラリーの構築

  梶原将大, 梶原将大, 迫田義博, 迫田義博, 伊藤壽啓, 高田礼人, 伊藤公人, 岸田典子, 五十嵐学, 磯田典和, 磯田典和, 曽田公輔, 曽田公輔, WEBSTER R.G, 喜田宏, 喜田宏, 喜田宏  日本獣医学会学術集会講演要旨集  144th-  91  2007/08/27  [Not refereed][Not invited]
  
• Filovirus infection: pathogenesis and host specificity

  高田 礼人  Protein, nucleic acid and enzyme  52-  (10)  1242  -1247  2007/08  [Not refereed][Not invited]
  
• ウイルス病原性発現の分子機構 フィロウイルスの病原性と宿主域

  高田礼人  蛋白質 核酸 酵素  52-  (10)  1242-1247  -1247  2007/08  [Not refereed][Invited]
  
• Importance of interaction between macrophage galactose-type C-type lectin(MGL/CD301) and viral envelope glycoprotein O-glycans in the infectivity of Ebola virus

  USAMI Katsuaki, TAKEUCHI Hideyuki, FUJIOKA Kouki, TAKADA Ayato, TAKADA Ayato, KAWAOKA Yoshihiro, KAWAOKA Yoshihiro, IRIMURA Tatsuro  生化学  80回・30回-  4P-0022  -7  2007  [Not refereed][Not invited]
  
• Protective efficacy of neutralizing antibodies against Ebola virus infection

  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.
• Glaciological experiments in the bar

  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]
  
• An Analysis of Correlated Mutations in the Hemagglutinin of Influenza Viruses

  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.
• An Analysis of Correlated Mutations in the Hemagglutinin of Influenza Viruses

  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.
• An Analysis of Correlated Mutations in the Hemagglutinin of Influenza Viruses

  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.
• An Analysis of Correlated Mutations in the Hemagglutinin of Influenza Viruses

  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.
• インフルエンザウイルス亜型間交差中和活性を有するモノクローナル抗体の解析

  吉田玲子, 五十嵐学, 喜田宏, 喜田宏, 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  54th-  347  2006/11/01  [Not refereed][Not invited]
  
• Tyro3 family-mediated cell entry of Ebola and Marburg viruses

  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.
• Entry and Egress of Enveloped Viruses

  TAKADA AYATO  膜  31-  (5)  243-247  -247  2006/09/01  [Not refereed][Not invited]
  
• エボラウイルス感染におけるマクロファージレクチンの糖鎖認識特性の重要性

  宇佐美克明, 竹内英之, 藤岡宏樹, 高田礼人, 河岡義裕, 河岡義裕, 入村達郎  日本糖質学会年会要旨集  26th-  35  2006/07/28  [Not refereed][Not invited]
  
• フィロウイルス感染症

  高田 礼人  臨床と微生物 = Clinical microbiology  33-  (4)  337  -343  2006/07/25  [Not refereed][Invited]
  
• Molecular determinants of Ebola virus virulence in mice

  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.
• 感染症:最近の世界の現状 フィロウイルス感染症

  高田礼人  臨床と微生物  33-  (4)  337-343  -343  2006/07  [Not refereed][Invited]
  
• 北海道大学人獣共通感染症リサーチセンター国際疫学部門

  高田 礼人  ウイルス  56-  (1)  125  -126  2006/06/26  [Not refereed][Invited]
  
• エボラウイルス表面糖蛋白質の機能解析

  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.
• Disease constraints for utilization of the African buffalo (Syncerus caffer) on game ranches in Zambia

  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.
• Glycans in Ebola viral glycoprotein recognized by macrophage galactose-type C-type lectin (MGL/CD301)

  USAMI Katsuaki, TAKEUCHI Hideyuki, FUJIOKA Kouki, TAKADA Ayato, KAWAOKA Yoshihiro, KAWAOKA Yoshihiro, IRIMURA Tatsuro  生化学  A14472(1P-A-168)  2006  [Not refereed][Not invited]
  
• Avian flu: Isolation of drug-resistant H5N1 virus (vol 437, pg 1108, 2005)

  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  438-  (7069)  754  -754  2005/12  [Not refereed][Not invited]
  
• エボラウイルス受容体

  下島昌幸, 下島昌幸, 高田礼人, 高田礼人, 海老原秀樹, 海老原秀樹, 藤岡宏樹, 入村達郎, FELDMANN Heinz, 河岡義裕, 河岡義裕, 河岡義裕  日本分子生物学会年会講演要旨集  28th-  89  2005/11/25  [Not refereed][Not invited]
  
• Interaction between Filovirus Proteins and Host Cell Membrane

  TAKADA AYATO, NODA TAKESHI, KAWAOKA YOSHIHIRO  膜  30-  (2)  68-72  -72  2005/03/01  [Not refereed][Invited]
  
• フィロウイルスの細胞への侵入 (特集 ウイルス増殖に関わる宿主細胞分子)

  高田 礼人, 河岡 義裕  細胞工学  24-  (2)  141  -144  2005/02  [Not refereed][Not invited]
  
• Entry of Filoviruses into Host Cells

  TAKADA AYATO, KAWAOKA YOSHIHIRO  細胞工学  24-  (2)  141-144  -144  2005/01  [Not refereed][Invited]
  
• Interaction of Ebola virus glycoprotein with human macrophage galactose-type C-type lectin

  K Usami, H Takeuchi, K Fujioka, A Takada, Y Kawaoka, T Irimura  GLYCOBIOLOGY  14-  (11)  1101  -1101  2004/11  [Not refereed][Not invited]
  
• Enhanced virulence of influenza A viruses with the haemagglutinin of the 1918 pandemic virus

  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).
• Interaction of Ebola virus glycoprotein with human macrophage galactose-type C-type lectin

  USAMI K, TAKEUCHI H, FUJIOKA K, TAKADA A, KAWAOKA Y, IRIMURA T  生化学  76-  (8)  1014  2004/08/25  [Not refereed][Not invited]
  
• A Human Macrophage Galactose-Type C-Type Lectin (MGL) Promotes Filovirus Entry

  FUJIOKA K, TAKADA A, TSUIJI M, MORIKAWA A, HIGASHI N, EBIHARA H, KOBASA D,FELDMANN H, KAWAOKA Y  Trends Glycoscience Glycotechnology  16-  (Supplement)  S55  2004/05/02  [Not refereed][Not invited]
  
• Human macrophage C-type lectin specific for galactose and N-acetylgalactosamine promotes filovirus entry

  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.
• インフルエンザ研究の最前線 インフルエンザのウイルス学 病原性の機序

  堀本泰介, 高田礼人  最新医学  59-  (2)  223-230  -230  2004/02  [Not refereed][Invited]
  
• エボラウイルス感染における抗体依存性感染増強現象

  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]
  
• Antibody-dependent enhancement of viral infection: molecular mechanisms and in vivo implications

  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.
• Antibody-dependent enhancement of Ebola virus infection

  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.
• Intranasal immunization with formalin-inactivated virus vaccine induces a broad spectrum of heterosubtypic immunity against influenza A virus infection in mice

  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.
• 鼎談 これからのインフルエンザワクチン

  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]
  
• イヌMxタンパク質の機能的解析

  中村鉄平, 浅野淳, 高在弘, 昆泰寛, 高田礼人, 渡辺智正, 安居院高志  日本獣医学会学術集会講演要旨集  135th-  2003
• Identification of protective epitopes on Ebola virus glycoprotein at the single amino acid level by using recombinant vesicular stomatitis viruses

  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.
• ウイルスの病原性発現における糖タンパク質の役割 (免疫病と感染症)

  堀本 泰介, 五藤 秀男, 高田 礼人  免疫  2003-  212  -226  2003  [Not refereed][Not invited]
  
• Intranasal administration of a synthetic peptide vaccine encapsulated in liposome together with an anti-CD40 antibody induces protective immunity against influenza A virus in mice

  A Ninomiya, K Ogasawara, K Kajino, A Takada, H Kida  VACCINE  20-  (25-26)  3123  -3129  2002/08  [Not 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.
• Seroepidemiological evidence of avian H4, H5, and H9 influenza A virus transmission to pigs in southeastern China

  A Ninomiya, A Takada, K Okazaki, KF Shortridge, H Kida  VETERINARY MICROBIOLOGY  88-  (2)  107  -114  2002/08  [Not 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.
• Ebola virus VP40 drives the formation of virus-like filamentous particles along with GP

  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.
• Polymorphisms and the differential antiviral activity of the chicken Mx gene

  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  2002/04  [Not 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.
• The invasion routes of neurovirulent A Hong Kong 483/97 (H5N1) influenza virus into the central nervous system after respiratory infection in mice

  CH Park, M Ishinaka, A Takada, H Kida, T Kimura, K Ochiai, T Umemura  ARCHIVES OF VIROLOGY  147-  (7)  1425  -1436  2002  [Not 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.
• The pathogenesis of Ebola hemorrhagic fever

  A Takada, Y Kawaoka  TRENDS IN MICROBIOLOGY  9-  (10)  506  -511  2001/10  [Refereed][Invited]
   

  Ebola virus causes lethal hemorrhagic disease in humans, yet there are sti I I no satisfactory biological explanations to account for its extreme virulence. This review focuses on recent findings relevant to understanding the pathogenesis of Ebola virus infection and developing vaccines and effective therapy. The available data suggest that the envelope glycoprotein and the interaction of some viral proteins with the immune system are likely to play important roles in the extraordinary pathogenicity of this virus. There are also indications that genetically engineered vaccines, including plasmid DNA and viral vectors expressing Ebola virus proteins, and passive transfer of neutralizing antibodies could be feasible options for the control of Ebola virus-associated disease.
• The pathogenesis of Ebola hemorrhagic fever

  A Takada, Y Kawaoka  TRENDS IN MICROBIOLOGY  9-  (10)  506  -511  2001/10  [Not refereed][Not invited]
   

  Ebola virus causes lethal hemorrhagic disease in humans, yet there are sti I I no satisfactory biological explanations to account for its extreme virulence. This review focuses on recent findings relevant to understanding the pathogenesis of Ebola virus infection and developing vaccines and effective therapy. The available data suggest that the envelope glycoprotein and the interaction of some viral proteins with the immune system are likely to play important roles in the extraordinary pathogenicity of this virus. There are also indications that genetically engineered vaccines, including plasmid DNA and viral vectors expressing Ebola virus proteins, and passive transfer of neutralizing antibodies could be feasible options for the control of Ebola virus-associated disease.
• Primary target cells of virulent strains of type A influenza virus in chicken embryos

  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.
• Mucosal vaccination against influenza: Protection of pigs immunized with inactivated virus and ether-split vaccine

  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.
• Ebola virus glycoprotein: Proteolytic processing, acylation, cell tropism, and detection of neutralizing antibodies

  H Ito, S Watanabe, A Takada, Y Kawaoka  JOURNAL OF VIROLOGY  75-  (3)  1576  -1580  2001/02  [Not 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.
• Plasminogen-binding activity of neuraminidase determines the pathogenicity of influenza a virus（共著）

  Journal of Virology  75, 9297-9301-  2001  [Not refereed][Not invited]
  
• Mouse Mx2 protein inhibits hantavirus but not influenza virus replication

  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.
• Downregulation of beta 1 integrins by Ebola virus glycoprotein: Implication for virus entry

  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.
• Recognition of N-glycolylneuraminic acid linked to galactose by the alpha 2,3 linkage is associated with intestinal replication of influenza A virus in ducks

  T Ito, Y Suzuki, T Suzuki, A Takada, 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.
• Recognition of N-glycolylneuraminic acid linked to galactose by the alpha 2,3 linkage is associated with intestinal replication of influenza A virus in ducks

  T Ito, Y Suzuki, T Suzuki, A Takada, 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.
• Interspecies transmission of influenza viruses: H5N1 virus and a Hong Kong SAR perspective

  Kennedy F. Shortridge, Peng Gao, Yi Guan, Toshihiro Ito, Yoshihiro Kawaoka, Deborah Markwell, Ayato Takada, Robert G. Webster  Veterinary Microbiology  74-  (1-2)  141  -147  2000/05/22  [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. Copyright (C) 2000 Elsevier Science B.V.
• Interspecies transmission of influenza viruses: H5N1 virus and a Hong Kong SAR perspective

  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.
• ウシヘルペスウイルス1のリセプター特異性

  岡崎克則, 高田礼人, 喜田宏  日本獣医学会学術集会講演要旨集  129th-  86  2000/03/01  [Not refereed][Not invited]
  
• Structure of the infected cell protein 0 gene of canine herpesvirus

  M Miyoshi, M Takiguchi, J Yasuda, A Hashimoto, A Takada, K Okazaki, H Kida  ARCHIVES OF VIROLOGY  145-  (8)  1715  -1723  2000  [Not 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.
• Precursor genes of future pandemic influenza viruses are perpetuated in ducks nesting in Siberia

  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  2000  [Not 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.
• Avian influenza virus intranasally inoculated infects the central nervous system of mice through the general visceral afferent nerve

  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  2000  [Not 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.
• 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.
• Protective effects of intranasal vaccination with plasmid encoding pseudorabies virus glycoprotein B in mice

  Ayato Takada, Katsunori Okazaki, Hiroshi Kida  Japanese Journal of Veterinary Research  47-  (1-2)  25  -33  1999/08  [Not 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.
• Induction of antibodies in mice by a recombinant baculovirus expressing pseudorabies virus glycoprotein B in mammalian cells

  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.
• Detection of canine herpesvirus DNA in the ganglionic neurons and the lymph node lymphocytes of latently infected dogs

  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  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.
• Antigenic and genetic analyses of H5 influenza viruses isolated from ducks in Asia

  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.
• Structure of the immediate early gene of canine herpesvirus - Brief report

  M Miyoshi, M Takiguchi, J Yasuda, A Hashimoto, A Takada, K Okazaki, H Kida  ARCHIVES OF VIROLOGY  144-  (2)  407  -420  1999  [Not refereed][Not invited]
   

  The nucleotide sequence of the immediate early (IE) gene of canine herpesvirus was determined. This gene was located in the inverted repeat regions, encoding a polypeptide of 1,383 amino acids. The predicted amino acid sequence was most closely related to that of the feline herpesvirus 1 IE protein among those of other alphaherpesviruses. DNA binding and transcriptional activation domains were found in the IE protein. A spliced region of the IE gene transcript was determined in its 5' non-coding region.
• Characterization of avian H5N1 influenza viruses from poultry in Hong Kong

  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  1998/12  [Not 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.
• Detection of antibody for hemagglutinin of avian influenza virus in pig of China.

  NINOMIYA AI, YASUDA JIRO, TAKADA AYATO, OKAZAKI KATSUNORI, SHORTRIDGE K F, KIDA HIROSHI  日本獣医学会学術集会講演要旨集  126th-  142  1998/08  [Not refereed][Not invited]
  
• Changes in H3 influenza A virus receptor specificity during replication in humans

  K Ryan-Poirier, Y Suzuki, WJ Bean, D Kobasa, A Takada, T Ito, Y Kawaoka  VIRUS RESEARCH  56-  (2)  169  -176  1998/08  [Not 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.
• Pathogenesis of Ebola virus infection: recent insights

  A Takada, Y Kawaoka  TRENDS IN MICROBIOLOGY  6-  (7)  258  -259  1998/07  [Refereed][Invited]
  
• Identification of the latent infected part of canine herpesvirus.

  MIYOSHI MASAHIRO, ISHII YUKI, TAKIGUCHI MITSUYOSHI, YASUDA JUN, HASHIMOTO AKIRA, TAKADA AYATO, OKAZAKI KATSUNORI, KIDA HIROSHI  日本獣医学会学術集会講演要旨集  125th-  140  1998/03  [Not refereed][Not invited]
  
• Potentially virulent Newcastle disease viruses are maintained in migratory waterfowl populations

  H Takakuwa, T Ito, A Takada, K Okazaki, H Kida  JAPANESE JOURNAL OF VETERINARY RESEARCH  45-  (4)  207  -215  1998/02  [Not refereed][Not invited]
   

  Forty-seven Newcastle disease virus (NDV) strains isolated from fecal samples of waterfowls in Alaska and Siberia from 1991 to 1996 were analyzed for their virulence. None of the viruses formed plaques on MDBK cells in the absence of trypsin. Of these, 29 strains showed virulent character by the mean death time with the minimum lethal dose in chicken embryos comparable to velogenic NDV strains. Of the 29 strains, 11 were sequenced for their fusion protein (F) gene. The results showed that 5 of them contained a pair of dibasic amino acids at the cleavage site of the F, which is of a virulent type. The present results suggest that potentially virulent strains of NDV are maintained in migratory waterfowl populations in nature, and that some of those may be transmitted to domestic poultry and acquire pathogenicity during passages in chicken population.
• Pathogenesis of Ebola Virus Infection

  Tends in Microbiology  6-  (7)  258  1998  [Not refereed][Not invited]
  
• An Attenuation Mechanism of Newcastle Disease Vaccine Strain TCND. (共著)

  Archives of Virology  143-  (6)  1129  -1143  1998  [Not refereed][Not invited]
  
• A system for functional analysis of Ebola virus glycoprotein

  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.
• Use of a mutant VSV for analysis of viral proteins.

  KAWAOKA YOSHIHIRO, TAKADA AYATO, WHITT M A  実験医学  15-  (19)  2389-2393  -2393  1997/12  [Not refereed][Not invited]
  
• The cysteine residues of the M2 protein are not required for influenza A virus replication

  MR Castrucci, M Hughes, L Calzoletti, Donatelli, I, K Wells, A Takada, Y Kawaoka  VIROLOGY  238-  (1)  128  -134  1997/11  [Not 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.
• Functional analysis of Ebola virus surface glycoprotein. Expression by plasmid and utilization of G protein deleted VSV.

  TAKADA AYATO, SANCHEZ A, WHITT M A, YOSHIDA HIROSHI, OKAZAKI KATSUNORI, KAWAOKA YOSHIHIRO  日本獣医学会学術集会講演要旨集  124th-  147  1997/09  [Not refereed][Not invited]
  
• Sialyl-linkage mediated selection for the appearance of host cell variants of influenza A viruses

  Y Suzuki, T Ito, H Masuda, A Takada, A Kawamoto, K Otsuki, D Miyamoto, T Suzuki, H Kida, Y Kawaoka  FASEB JOURNAL  11-  (9)  A1443  -A1443  1997/07  [Not refereed][Not invited]
  
• Laboratory of Microbiology

  KIDA Hiroshi, OKAZAKI Katsunori, TAKADA Ayato  Japanese journal of veterinary research  45-  (1)  32  -33  1997/05/30  [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

  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  1997/04  [Not 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.
• Expression of recombinant protein in mammal cell with baculovirus as a vector.

  AOKI HIROFUMI, SAKODA YOSHIHIRO, TSUCHIYA YOSHINORI, SHOJI IKUO, MATSUURA ZENJI, TAKADA AYATO, KIDA HIROSHI, SUGIYAMA KIMIHIRO, FUKUSHO AKIO  日本獣医学会学術集会講演要旨集  123rd-  153  1997/03  [Not refereed][Not invited]
  
• N-acetyl neuraminic acid-2,6-galactoselinkage is abundantin amnion but not in allantois of chicken eggs: Implication on efficient isolation of human influenza viruses in the former compartment and selection of egg variants

  Ito, T, Suzuki, T, Takada, A, Kawamoto, A, Otsuki, K, Masuda, H, Yamada, M, Suzuki, Y, Kida, H, Kawaoka, Y  Journal of Virology  71-  3357  -3362  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]
  
• Protective immune response of chickens against Newcastle disease, induced by the intrarasal vaccination with inactivated virus

  Veterinary Microbiology  50-  17  1996  [Not refereed][Not invited]
  
• ACQUISITION OF PATHOGENICITY OF A NEWCASTLE DISEASE VIRUS ISOLATED FROM A JAPANESE QUAIL BY INTRACEREBRAL PASSAGE IN CHICKENS

  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.
• Perpetuation of influenza A viruses in Alaskan waterfowl reservoirs(共著)

  Archives of Virology  140-  (7)  1163  -1172  1995  [Not refereed][Not invited]
  
• Induction of Protective antibody responses against pseudorabies virus by intranasal vaccination with glycoprotein B in mice

  Archives of Virology  140-  (9)  1629  -1635  1995  [Not refereed][Not invited]
  
• 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  1994/12  [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 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.
• PROTECTION OF MICE AGAINST AUJESZKYS-DISEASE VIRUS-INFECTION BY INTRANASAL VACCINATION WITH INACTIVATED VIRUS

  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.
• EXPERIMENTAL STUDY ON THE MUCOSAL IMMUNIZATION OF ANIMALS USING INACTIVATED VIRUSES

  TAKADA Ayato  Japanese journal of veterinary research  41-  (1)  49  -49  1993/05/27

Books etc

• Encyclopedia of Infection and Immunity

  Ayato Takada (ContributorEbolavirus and Other Filoviruses)
  Elsevier Science 2022/04
• 創薬研究者がこれだけは知っておきたい最新のウイルス学

  技術情報協会 (Contributorエボラウイルス増殖機構と病原性発現メカニズム)
  技術情報協会 2021/08 (ISBN: 9784861048555) 602p
• ザンビアを知るための55章

  島田周平, 大山修一, 髙田 礼人ほか (Contributorエボラおよびマールブルグウイルス－ザンビア大学獣医学部との共同研究－)
  明石書店 2020/08
• 新型コロナはいつ終わるのか

  岩田健太郎, 髙田礼人 ほか (Contributor無症状感染と「集団免疫」の基礎知識)
  宝島社 2020/06
• 新型コロナ 19氏の意見 われわれはどこにいて、どこにむかうのか

  内山節, 山本太郎, 髙田礼人 ほか (Contributorウイルスとは何かを知れば、向き合い方が見えてくる)
  農山漁村文化協会 2020/05
• 新型コロナ感染爆発と隠された中国の罪

  五味洋治、高橋洋一、高田礼人 ほか (Contributor水際では止められない不顕性感染の新型コロナウイルス)
  宝島社 2020/04 (ISBN: 9784299004376) 249p
• 新興ウイルス感染症に対する抗体療法の開発

  高田 礼人 (Contributor)
  日本医事新報社 2019/01
• ウイルスは悪者か : お侍先生のウイルス学講義

  高田 礼人 
  亜紀書房 2018 (ISBN: 9784750515595)
• ボクが逆さに生きる理由 誤解だらけのこうもり (Natsume-sha Science)

  中島宏章, 福井大, 高田礼人 (Supervisor)
  ナツメ社 2017/10 (ISBN: 4816363459) 248

  
• 人獣共通感染症 改訂3版

  高田 礼人 (Contributorマールブルグ出血熱、エボラ出血熱)
  医薬ジャーナル社 2016/02
• 感染症から知るウイルス・細菌〈3〉感染症の予防と研究最前線!―病原体とのたたかいから利用へ

  西條 政幸, 高田 礼人, 高橋 幸裕 (Joint work)
  学研教育出版 2011/02 (ISBN: 4055007981) 44

  
• 感染症から知るウイルス・細菌〈2〉細菌とウイルスの正体を知ろう!―ウイルスは生物?

  西條 政幸, 高田 礼人, 高橋 幸裕 (Joint work)
  学研教育出版 2011/02 (ISBN: 4055007973) 44

  
• 感染症から知るウイルス・細菌〈1〉感染症の原因を知ろう!―なぜかかる?なぜうつる?

  西條 政幸, 高田 礼人, 高橋 幸裕 (Joint work)
  学研教育出版 2011/02 (ISBN: 4055007965) 44

  
• 獣医微生物学第3版

  高田 礼人 (Contributorフィロウイルスと感染症)
  文永堂出版 2011
• Ebola and Marburg Viruses: Molecular and Cellular Biology.

  Neumann G, Noda T, Takada A, Jasenosky LD, Kawaoka Y (Joint workRoles of filoviral matrix- and glycoproteins in the viral life cycle.)
  Horizon 2004

Presentations

• Niemann-Pick C1 heterogeneity controlling filovirus tropism to bat cells  [Invited]

  Ayato Takada

  10th International Filovirus Symposium  2022/09
• 何故 H5 および H7 亜型のウイルスだけが高病原性鳥インフルエンザウイルスになるのか  [Invited]

  髙田 礼人

  令和3年度「感染・免疫・がん・炎症」全国共同研究拠点シンポジウム  2022/03
• The importance and issues of collaboration during R&D period for infectious disease control in Africa  [Invited]

  Ayato Takada

  AMED International Joint Symposium 2021  2021/11
• ウイルスは悪者か  [Invited]

  髙田 礼人

  経団連フォーラム21  2021/10
• Viral Zoonosis and Global Surveillance  [Invited]

  Ayato Takada

  Third Country Training Program (Online) PREPARE for Emerging and Re-emerging Infectious Diseases in Southern Africa Region  2021/10
• JICA SATREPS ザンビア・コンゴにおける人獣共通感染症対策の成果と課題  [Invited]

  髙田礼人

  環境省タンザニア気候変動適応事業設計  2021/05
• Filovirus research: VSV Pseudotype and antibody  [Invited]

  Ayato Takada

  J-Rapid/eAsia Joint Symposium Power of Natural Products  2021/03
• ウイルス性新興感染症の制御を目指して  [Invited]

  髙田礼人

  第16回北海道HIV/AIDS歯科医療研究会  2021/02
• ウイルスは悪者か  [Invited]

  髙田礼人

  ブンナビ薬学特別企画 対談シリーズ  2020/12
• フィロウイルス研究における最近の知見  [Invited]

  髙田礼人

  第162回日本獣医学会学術集会  2019/09
• エボラおよびマールブルグ出血熱  [Invited]

  髙田礼人

  第8回日本精神科医学会学術集会  2019/07
• 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

Teaching Experience

Advanced Lecture on MicrobiologyAdvanced Lecture on Microbiology Hokkaido University
-- Hokkaido University
Advanced and Comprehensive Studies on Zoonosis ControlAdvanced and Comprehensive Studies on Zoonosis Control Hokkaido University
-- Hokkaido University

Association Memberships

• 日本ワクチン学会   日本ウィルス学会   日本獣医学会   American Society for Microbiology   

Research Projects

• 細胞の動的応答を制御する海洋天然物の探索

  日本学術振興会：科学研究費助成事業

  Date (from‐to) : 2022/04 -2025/03 

  Author : 酒井 隆一, 高田 礼人, 田中 良和, 松本 健, 八代田 陽子, 荒木 真理人
• Structure-based functionalization of novel sugar-binding proteins derived from marine natural products for corona drug discovery

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research

  Date (from‐to) : 2022/04 -2025/03 

  Author : 田中 良和, 高田 礼人, 酒井 隆一, 横山 武司
• 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
• マールブルグ出血熱に対する抗体医薬開発

  日本学術振興会：科学研究費助成事業 基盤研究(B)

  Date (from‐to) : 2021/04 -2024/03 

  Author : 高田 礼人

   

  エボラおよびマールブルグウイルスは、ヒトまたはサルに急性で致死率の高い感染症（エボラ出血熱およびマールブルグ出血熱）を引き起こす。現在までに承認された抗体医薬はエボラウイルス1種のみに特異的であり、マールブルグウイルスには全く効果が無い。本研究は、これまでエボラウイルスに対する抗体に関して得られた知見を基にして、ウイルスの細胞侵入に関わる特定のエピトープに着目することによって、マールブルグ出血熱に対して有効な抗体医薬の開発に資する事を目的とする。 マールブルグウイルス（Angola株）の表面糖蛋白質(GP)の遺伝子をゲノム内に組み込んだ増殖性の水疱性口炎ウイルス（シュードタイプウイルス）に感染させ回復したマウスに、マールブルグウイルス（Angola株）のGPを持つウイルス様粒子を腹腔内投与してブーストした。そのマウスから脾臓を採取し、脾細胞とミエローマ細胞を融合させ、通常の方法でハイブリドーマの作出を試みた。その結果、調べたマールブルグウイルス株（Angola、MusokeおよびRavn）全てに交差中和活性を示すモノクローナルIgG抗体およびAngolaとMusoke株にのみ中和活性を示すIgG抗体が複数得られた。 フィロウイルスのレセプターであるNPC1分子上のドメインC内に存在するloop構造（loop1およびloop2）を模した合成ペプチドをヒトNPC1のアミノ酸配列を基に設計した。これらのペプチドの中和活性を、非増殖性シュードタイプ水疱性口炎ウイルスを用いて解析した結果、阻害薬候補となるアミノ酸配列を見出した。
• 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
• Study for the development of Ebola hemorrhagic fever treatment

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research

  Date (from‐to) : 2016/04 -2021/03 

  Author : Takada Ayato

   

  Mouse monoclonal antibodies with cross-neutralizing activity against multiple ebolavirus species were generated. Detailed analyses of the epitopes recognized by the antibodies suggested that that some of the antibodies bind to the side region of the viral surface glycoprotein (GP) molecule and inhibit the membrane fusion between the cellular membrane and the virus envelope. Electron microscopic structural analysis of the complex of the GP molecule and Mab 6D6, one of the cross-reactive antibodies, revealed that this antibody recognizes the fusion loop region of GP. In addition, a novel compound having the ability to reduce the infectivity of all known ebolaviruses were obtained from an existing compound library. It was suggested that this compound binds to the side surface region of the GP molecule and inhibits the membrane fusion.
• Development of universal antibody therapy for Ebola hemorrhagic fever

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Exploratory Research

  Date (from‐to) : 2015/04 -2018/03 

  Author : Takada Ayato

   

  Ebolaviruses cause severe hemorrhagic fever (Ebola hemorrhagic fever) in humans and nonhuman primates. Five phylogenetically distinct species are currently known in the genus Ebolavirus and the antigenicity among these ebolavirus species is also largely different. This study attempted to establish procedures to induce neutralizing antibodies cross-reactive to all known ebolavirus species. Our data suggest the possibility that cross-reactive neutralizing antibodies can be induced more efficiently in mice immunized with virus-like particles using an immunosuppressive agent (rapamycin) or mice infected with replication competent vesicular stomatitis virus pseudotyped with ebolavirus glycoprotein than in mice immunized with virus-like particles using a standard adjuvant.
• フィロウイルスの細胞侵入メカニズムと宿主域に関する研究

  日本学術振興会：科学研究費助成事業 新学術領域研究(研究領域提案型)

  Date (from‐to) : 2015/04 -2017/03 

  Author : 高田 礼人

   

  ヒトNPC1の多様性とフィロウイルスに対する感受性：ヒト由来Jurkat TおよびRamos B細胞はいずれもフィロウイルスに対して非感受性である事が知られている。面白いことに、Jurkat T細胞は、フィロウイルスの細胞吸着因子の一つであるC型レクチンを発現させることによって感受性を獲得したのに対して、Ramos B細胞は非感受性のままであった。そこで、Jurkat TおよびRamos B細胞のNPC1遺伝子をクローニングし、アミノ酸配列を比較したところ、2箇所（200番目と215番目）のアミノ酸が異なる事が分かった。これらのNPC1をクローニングし強制発現させた細胞を確立し、Vesicular Stomatitis Virus (VSV)のシュードタイプシステムを用いて、フィロウイルスに対する感受性の違いを解析したが、違いは認められなかった。
  コウモリ由来細胞のフィロウイルスに対する感受性を決定する因子の探索：既知のフィロウイルス全てについて、GPを持つシュードタイプウイルスの感染性を様々なコウモリ由来の培養細胞で比較した結果、エボラウイルスは感染するがマールブルグウイルスは感染しないコウモリ細胞（FBKT1）が存在する事が明らかとなり、レセプター（NPC1）の違いによるものと推定された。FBKT1にヒト由来のNPC1を導入するとマールブルグウイルスに対する感受性を獲得した。マールブルグウイルス感受性細胞とFBKT1の間で、NPC1のアミノ酸配列を比較した結果、GPとの結合に重要と考えられている領域のアミノ酸配列に違いが認められた。また、NPC1との結合に重要と考えられているGP上のアミノ酸配列にもエボラウイルスとマールブルグウイルスの間で違いが認められた。これらのアミノ酸を入れ替えると感染性が逆転した。以上より、GPとNPC1の結合がフィロウイルスの宿主域に関与している事が示唆された。
• Computational analysis of a conformational epitope of a broadly neutralizing antibody in influenza A virus hemagglutinin

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

  Date (from‐to) : 2013/04 -2017/03 

  Author : Igarashi Manabu, TAKADA AYATO

   

  The HA of influenza A viruses is classified into 16 subtypes (H1-H16). We have previously reported a cross-reactive antibody, designated S139/1, which neutralizes H1, H2, H3, H13, and H16 subtypes. In this study, we characterized the S139/1 recognition sites on different HAs by computational methods such as molecular modeling and dynamics simulations. We investigated the contribution of individual residues on each HA to the interaction with S139/1, and found that amino acids at 10 positions on HA strongly contributed to S139/1 binding as for the strains neutralized by S139/1. Analysis of hydrogen bond interactions emphasized that the residues at positions 156, 158 and 193 were the most important for S139/1 binding. Indeed, amino acid substitutions at these three positions were experimentally observed in the mutant viruses escaping from neutralization by S139/1. Thus, our computational methods identified the amino acid residues critical for the cross-neutralizing activity of S139/1.
• Cross-reactive antibodies to influenza virus hemagglutinins and their epitopes

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

  Date (from‐to) : 2012/04 -2017/03 

  Author : Takada Ayato, ITO Kimihito, IGARASHI Manabu, YOSHIDA Reiko, MURAMATSU Mieko, MIYAMOTO Hiroko, Wilson Ian

   

  By focusing on a novel antiviral function (budding inhibition), we found potential roles of anti-hemagglutinin antibodies in cross-protective immunity against influenza A viruses with multiple HA subtypes. It was particularly noted that the comparison between IgG and IgA antibodies recognizing the same epitope on the HA molecule clearly demonstrated the greater potential of HA-specific IgA than IgG antibodies in antiviral activities. It was also attempted to modify antibody specificity by structure-based molecular simulation and production of recombinant antibodies.
• 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
• Predicting antigenic changes of influenza viruses using text data assimilation.

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

  Date (from‐to) : 2013/04 -2016/03 

  Author : Kimihito Ito, TAKADA Ayato, IGARASHI Manabu

   

  Human influenza viruses mutate from time to time, causing annual epidemics worldwide. Given the high mutation rate of the viral gene, influenza vaccine strains need to be updated every two to three years. In order to prepare an effective vaccine strain prior to each influenza season, we investigated text data assimilation methods that allow us to predict the evolution of viral amino acid sequences. As a result, we developed a mathematical model consisting of human herd immunity, the number of infections, and population of circulating vires. The model provides a mathematical basis of bioinformatics methods to prepare an effective vaccine strain prior to influenza seasons.
• Mechanism of IgA-mediated cross protection induced by mucosal immunization

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

  Date (from‐to) : 2013/04 -2016/03 

  Author : YOSHIDA Reiko, TAKADA Ayato

   

  Mucosal immunization with influenza virus vaccine confers cross-protection against infection with influenza virus antigenic variants as well as multiple subtypes of influenza A viruses. In this study, we found that many cross-reactive but non-neutralizing anti-HA IgA antibodies are produced in the upper respiratory tract of immunized mice. These anti-HA IgA antibodies showed stronger antiviral activities against influenza A viruses than IgG antibodies. Particularly, the results from this study suggest that the inhibition of virus particles release from the infected cell surface may be one of the IgA-mediated protective mechanisms important for heterosubtypic immunity.
• フィロウイルスの宿主域と受容体に関する研究

  日本学術振興会：科学研究費助成事業 新学術領域研究(研究領域提案型)

  Date (from‐to) : 2013/04 -2015/03 

  Author : 高田 礼人

   

  フィロウイルス科に属するエボラおよびマールブルグウイルスはヒトを含む霊長類に重篤な出血熱をひきおこす病原体として知られている。しかし、人獣共通感染症病原体であるフィロウイルスの宿主特異性あるいは宿主域を決定する分子基盤に関する知見は殆どない。本研究では、ウイルスのtropism決定の第一因子である細胞侵入過程（吸着・膜融合）に着目し、フィロウイルスの宿主域決定メカニズムに関する基礎的知見を得ることを目的とする。 フィロウイルスの細胞吸着に関与する宿主分子の一つであるT-cell immunoglobulin and mucin domain 1 (TIM-1)に対するマウスモノクローナル抗体M224/1がフィロウイルスの細胞侵入を阻害する事を見出した。蛍光標識したウイルス様粒子（VLP）を用いて、M224/1存在下および非存在下で吸着・取り込み・膜融合を定量的に比較解析した結果、M224/1存在下でのVLPの吸着阻害効果は僅かであるが、VLPとエンドソーム膜との膜融合が著しく阻害された。興味深い事に、TIM-1はNiemann-Pick C1 (NPC1：フィロウイルスの膜融合に必須な宿主分子)はVero E6細胞内小胞において共局在し、両者は小胞内で結合していた。さらに、細胞内のTIM-1/NPC1複合体と膜融合を起こしているVLPの局在は一致した。以上の結果は、フィロウイルスはTIM-1を介して細胞表面に吸着した後、TIM-1と共に細胞内に取り込まれ、後期エンドソーム/リソソームにおいて、TIM-1とNPC1との結合を経て膜融合が起こることを示唆する。さらに、M224/1はTIM-1とNPC1との相互作用を阻害することによって、膜融合を阻害することが示された。
• Antibody-mediated inhibition of filovirus budding

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Exploratory Research

  Date (from‐to) : 2012/04 -2014/03 

  Author : TAKADA Ayato, ITO Kimihito, IGARASHI Manabu

   

  Monoclonal antibodies (MAbs) specific to Marburg virus glycoprotein (GP) were produced to analyze their activity to suppress the virus budding. We found that some of the MAbs, which did not have a classical neutralizing activity, efficiently inhibited the budding and release of MARV particles from infected cells. A couple of escape variants selected by these MAbs had deletions of the mucin-like region in GP or single amino acid substitutions in the furin-recognition motif in GP. These results demonstrate a novel mechanism of antibody-mediated inhibition of MARV budding and the extraordinary structural flexibility and variability of MARV GP to escape from immunological pressure of antibodies.
• インフルエンザウイルスの異種宿主間伝播に関与する細胞因子の研究

  日本学術振興会：科学研究費助成事業 特別研究員奨励費

  Date (from‐to) : 2010 -2012 

  Author : 高田 礼人, MANZOOR Rashid

   

  オルソミクスウイルス科に属するA型インフルエンザウイルスは様々な鳥類および哺乳類に感染する人獣共通感染症病原体である。その宿主域を決定する因子についてはこれまでに数多くの報告がなされている。それら因子のうちのいくつかは、宿主域のみならずウイルスの病原性をも決定することが知られている。しかしながら、ウイルスが宿主細胞へ侵入した後、それら因子がどのような宿主因子と相互作用し、どのようにウイルスの宿主域や病原性に関与しているのかを説明する知見はほとんど見られない。本研究は、これまで解明できなかったインフルエンザウイルスポリメラーゼ蛋白質と宿主因子の相互作用に関する様々な新しい知見を得ることを目的とする。 FLAGタグをC末端あるいはN末端に付した組換えインフルエンザウイルス(HK483およびHK486株)ポリメラーゼ蛋白質(PB2、PB1およびPA蛋白質)を単独で発現するプラスミドを293T細胞に導入し、可溶化後、抗FLAGタグ抗体ビーズを用いて免疫沈降した。SDS-PAGEで展開した結果、幾つかの宿主由来蛋白質バンドが確認された。これらのバンドを資料分析によって解析した結果、宿主蛋白質であるHsp70およびhnRNP等であることが判明した。これらの蛋白質はポリメラーゼと相互作用する因子である可能性が高い。共焦点顕微鏡を用いた観察によって、ポリメラーゼとこれらの宿主蛋白質は細胞内で共局在することが明らかとなった。HK483およびHK486株以外のインフルエンザAウイルスでの同様の現象が認められた。特にPB2蛋白質との相互作用が重要であった。さらに、siRNAを用いてHsp70をノックダウンした細胞(293TおよびHera細胞)では、インフルエンザウイルスの増殖が抑えられることを見出した。また、各種ストレスによって細胞内に核内に誘導されるHsp70発現とウイルス増殖効率の間に相関があることを見出した。 以上の結果は、Hsp70はインフルエンザAウイルスの増殖効率を高める宿主因子であることを示唆している。
• Antibody therapy against novel influenza virus infection using anImmunosuppressed non-human primate model

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

  Date (from‐to) : 2010 -2012 

  Author : ITOH Yasushi, OGASAWARA Kazumasa, ITO Mutsumi, TAKADA Ayato

   

  The purpose of this study is development of anti-influenza virus drugs that are effective under the immunocompromised condition.We developed human-mouse chimeric anti-influenza virus hemagglutinin antibody with neutralizing activity. Administration of the antibody to immunosuppressed cynomolgus macaques improved survival rates in highly pathogenic avian influenza virus infection, whereas immunosuppressed macaques without the treatment showed severe symptoms. It is expected that the therapy with the developed anti-hemagglutinin antibody would be effective in immunocompromised influenza patients as confirmed in the immunosuppressed macaquemodel.
• Studies on cross-reactive antibodies to multiple subtypes of influenza virus henagglutinin induced by mucosal immunization

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

  Date (from‐to) : 2009 -2011 

  Author : TAKADA Ayato, SUZUKI Yasuhiko, ITO Kimihito

   

  Currently used parenteral influenza vaccines induce serum IgG antibody response against hemagglutinin (HA), but do not protect primary infection in the respiratory tissues. In addition, vaccine-induced antibodies are not cross-reactive to viruses with different HA subtypes from that of vaccine strains. In this study, we found that intranasal immunization of mice with inactivated influenza viruses induced antibodies cross-reactive to multiple HA subtypes. Furthermore, it was suggested that IgA antibodies associated with mucosal immunity played an important role in heterosubtypic inhibition of virus replication.
• インフルエンザウイルスのRNA分節間に形成される塩基対の検索

  日本学術振興会：科学研究費助成事業 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...
• 抗体依存性感染増強現象を利用したアジュバントの開発

  日本学術振興会：科学研究費助成事業 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...
• Predicting Antigenic Changes of Influenza Viruses by Discovering Patterns of Viral Evolution

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

  Date (from‐to) : 2007 -2009 

  Author : ITO Kimihito, TAKADA Ayato, MINATO Shin-ichi

   

  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 ...
• フィロウイルスの細胞侵入機構と病原性発現に関する研究

  日本学術振興会：科学研究費助成事業 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、...
• 抗体の特異性を利用したターゲティングウイルスの開発

  日本学術振興会：科学研究費助成事業 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...
• A novel therapeutic strategy for zoonotic emerging and reemerging infections

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

  Date (from‐to) : 2005 -2008 

  Author : HORIMOTO Taisuke, GOTOH Hideo, TAKADA Ayato, YASUDA Jiro, SHIMOJIMA Masayuki, TAKADA Ayato, YASUDA Jiro, SHIMOJIMA Masayuki

   

  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。
• すべてのA型インフルエンザに有効な免疫法の開発

  日本学術振興会：科学研究費助成事業 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...
• フィロウイルスの病原性制御に関する研究

  日本学術振興会：科学研究費助成事業 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...
• フィロウイルスの病原性発現とその分子基盤に関する研究

  日本学術振興会：科学研究費助成事業 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...
• フィロウイルスの病原性発現とその分子基盤に関する研究

  日本学術振興会：科学研究費助成事業 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...
• Analysis on cellular factors regulating influenza virus infection using "self-motivating" recombinant virus

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

  Date (from‐to) : 2002 -2003 

  Author : HORIMOTO Taisuke, GOTO Hideo, TAKADA Ayato

   

  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...
• Study on development of a live attenuated vaccine for equine influenza infection using reverse genetics

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

  Date (from‐to) : 2001 -2003 

  Author : GOTO Hideo, TAKADA Ayato, HORIMOTO Taisuke, KAWAOKA Yoshihiro, KONDO Takashi

   

  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...
• フィロウイルスの病原性発現とその分子基盤に関する研究

  日本学術振興会：科学研究費助成事業 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...
• Development of influenza live vaccine by reverse genetics

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

  Date (from‐to) : 2001 -2002 

  Author : KAWAOKA Yoshihiro, HORIMOTO Taisuke, GOTO Hideo, TAKADA Ayato, OSUMI Kunio

   

  Influenza virus is a highly infectious respiratory pathogen of birds and mammals, including human. It often produces significant mobidity and mortality in humans. Current methods of immunization against influenza virus include parental administration of inactivated influenza virus vaccines, which reduces the incidence of clinical illness in healthy subjects. But in some cases, immunization with inactivated vaccines is associated with very low protection rates or short protection periods. Another promising approach to vaccination is the use of live-attenuated influenza viruses. Theses vaccines have shown considerable promise in ongoing clinical trials. However, it has been concerned that the pathogenesis happened to recovered during their replication. We propose an alternative approach to the design of live virus vaccines, one that relies on alteration of the influenza A virus M2 protein which is responsible for ion channel activity. It did not show appreciable growth defect in cell culture, although its growth was attenuated in mice. It has shown that this M2 ion channel defective mutant virus protected mice against challenge with lethal doses of influenza virus, indicating the potential or incorporating this M2 alteration in a live influenza vaccine as one of the attenuating mutations.
• Ecological study of animal influenza viruses : To prepare for the emergence of pandemic influenza

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (A)

  Date (from‐to) : 2000 -2002 

  Author : KIDA Hiroshi, KATSUNORI Okazaki, YOSHIHIRO Sakoda, YOSHIHIRO Kawaoka, AYATO Takada, TOSHIHIRO Ito

   

  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...
• フィロウイルスの病原性発現機構に関する研究

  日本学術振興会：科学研究費助成事業 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...
• 半生インフルエンザウイルスベクターの開発

  日本学術振興会：科学研究費助成事業 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...
• 合成ペプチドを用いた抗ウイルス薬の開発

  日本学術振興会：科学研究費助成事業 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...
• 粘膜免疫応答を誘導するDNAワクチンの開発

  日本学術振興会：科学研究費助成事業 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...
• Molecular mechanisms of infection and pathogenesis of viruses

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (A).

  Date (from‐to) : 1998 -1999 

  Author : KIDA Hiroshi, ITO Toshihiro, TAKADA Ayato, OKAZAKL Katsunori

   

  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. ...
• G蛋白欠失VSVを用いたウイルス表面糖蛋白の機能解析

  日本学術振興会：科学研究費助成事業 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...
• Molecular mechanisms of infection and pathogenesis of viruses

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (A)

  Date (from‐to) : 1996 -1998 

  Author : KIDA Hiroshi, ITO Toshihiro, UMEMURA Takashi, FUJITA Shoichi, MAEDA Yoshimitsu, NAKAZATO Yoshikazu

   

  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...
• Studies on the T-cell epitopes of herpesviruses

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

  Date (from‐to) : 1996 -1997 

  Author : OKAZAKI Katsunori, TAKADA Ayato, KIDA Hiroshi

   

  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...
• Prediction of new pandemic influenza viruses -Distribution of the viruses in waterfowls in their nesting places in Siberia

  Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for international Scientific Research

  Date (from‐to) : 1995 -1997 

  Author : KIDA Hiroshi, YAMNIKOVA Svetrana, KAWAOKA Yoshihiro, TAKADA Ayato, OKAZAKI Katsunori

   

  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

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
• 特開2005-187428:抗MGL抗体によるフィロウイルス治療薬  2005/07/14

  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
• 特開2005-185235:ウイルス感染に対する感受性診断のための方法およびキット。  2005/07/14

  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

Social Contribution

• 新感染症時代を生きる⑤〜アフリカの今…新型コロナと世界の未来は

  Date (from-to) : 2022/04/24

  Role : Appearance

  Sponser, Organizer, Publisher  : UHB北海道文化放送

  Event, Program, Title : 松本裕子の病を知る
• 新感染症時代を生きる③〜オミクロン株の次は？新型コロナ変異の行方

  Date (from-to) : 2022/04/10

  Role : Appearance

  Sponser, Organizer, Publisher  : UHB北海道文化放送

  Event, Program, Title : 松本裕子の病を知る
• コロナ下の羅針盤 ウイルスと共生 グローバル化の宿命

  Date (from-to) : 2022/01/10

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 北海道新聞

  Event, Program, Title : 北海道新聞
• 新感染症時代を生きる③〜未知のウイルスと闘う…人獣共通感染症研究所からの警告（後編)

  Date (from-to) : 2021/06/27

  Role : Appearance

  Sponser, Organizer, Publisher  : UHB北海道文化放送

  Event, Program, Title : 松本裕子の病を知る
• 新感染症時代を生きる②〜未知のウイルスと闘う…人獣共通感染症研究所からの警告

  Date (from-to) : 2021/06

  Role : Appearance

  Sponser, Organizer, Publisher  : UHB北海道文化放送

  Event, Program, Title : 松本裕子の病を知る
• まなびのひろば ぐんぐん 伸びゆく君へ

  Date (from-to) : 2021/05/25

  Role : Contribution

  Sponser, Organizer, Publisher  : 北海道新聞

  Event, Program, Title : 北海道新聞
• 「なぜ？」解き明かす探求心を

  Date (from-to) : 2021/05/25

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 北海道新聞

  Event, Program, Title : まなびのひろば ぐんぐん 伸びゆく君へ
• 野生のマウンテンゴリラの保護が地球を救う!? 世界の子ども応援SDGs 3時間SP

  Date (from-to) : 2021/05/05

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : TBS

  Event, Program, Title : 世界くらべてみたら
• 「コロナ第４波」拡大のウラで警戒すべき、次に来そうな「人獣共通感染症」の脅威

  Date (from-to) : 2021/04/30

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 講談社

  Event, Program, Title : 現代ビジネス
• 科学者 野口英世

  Date (from-to) : 2021/01

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : NHK

  Event, Program, Title : フランケンシュタインの誘惑 科学史 闇の事件簿
• パンデミックウイルス インフルエンザと新型コロナ

  Date (from-to) : 2020/10/14

  Role : Lecturer

  Sponser, Organizer, Publisher  : 朝日新聞

  Event, Program, Title : 朝日カルチャーセンター公開講座
• ウイルスとは何か

  Date (from-to) : 2020/09/30

  Role : Lecturer

  Sponser, Organizer, Publisher  : 朝日新聞

  Event, Program, Title : 朝日カルチャーセンター公開講座
• ウイルス対策は粘り強さが必要

  Date (from-to) : 2020/08/24

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 北海道新聞

  Event, Program, Title : 北海道新聞
• 新型コロナウイルスと共存するには 差別、偏見の課題

  Date (from-to) : 2020/08/19

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 読売新聞

  Event, Program, Title : 読売新聞医療健康サイト「ヨミドクター」
• 日本感染症研究に遅れ 規制・インフラ不足が壁

  Date (from-to) : 2020/08/16

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 日本経済新聞

  Event, Program, Title : 日本経済新聞
• ウイルスと感染症

  Date (from-to) : 2020/08/07

  Role : Lecturer

  Sponser, Organizer, Publisher  : 成田ロータリークラブ

  Event, Program, Title : 成田ロータリークラブ
• 感染症と闘う 新型コロナ 夏を乗り切るマスクの工夫

  Date (from-to) : 2020/08/05

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 毎日新聞

  Event, Program, Title : 毎日新聞
• コロナ撲滅ほぼ不可能

  Date (from-to) : 2020/07/30

  Role : Coverage cooperation

  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) : 2020/07/29

  Role : Lecturer

  Sponser, Organizer, Publisher  : 北海道新聞

  Event, Program, Title : 北海道政経懇話会
• ウイルスを知る

  Date (from-to) : 2020/07/22

  Role : Lecturer

  Sponser, Organizer, Publisher  : 北海道新聞

  Event, Program, Title : 苫小牧政経文化懇話会
• 新型コロナウイルスの新しい”宿主”ヒト やがて季節かせのようなウイルスになって定着化か

  Date (from-to) : 2020/06/15

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : The Big Issue

  Event, Program, Title : The Big Issue
• 新型コロナ 夏になれば感染減るの？

  Date (from-to) : 2020/06/10

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 朝日新聞

  Event, Program, Title : 朝日新聞
• 新型コロナのパンデミック 乗り越える鍵は皆が地球規模で考えること

  Date (from-to) : 2020/05/27

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 聖教新聞

  Event, Program, Title : 聖教新聞
• 聞く語る 野生動物と静かに共生 人類との遭遇増え脅威に

  Date (from-to) : 2020/05/15

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 北海道新聞

  Event, Program, Title : 北海道新聞
• ウイルスとは何か、その姿に迫る

  Date (from-to) : 2020/04/13

  Role : Lecturer

  Sponser, Organizer, Publisher  : TechnionJapan

  Event, Program, Title : 丸ノ内 square academy
• 教えて 新型コロナウイルス

  Date (from-to) : 2020/03/26

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 十勝毎日新聞

  Event, Program, Title : 十勝毎日新聞
• 聞きたい「新型コロナウイルス」予防の基本は薄める

  Date (from-to) : 2020/03/03

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 北海道新聞

  Event, Program, Title : 北海道新聞
• ネット情報にデマ

  Date (from-to) : 2020/02/28

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : 北海道新聞

  Event, Program, Title : 北海道新聞
• 動物から人へうつる病気のこと知って

  Date (from-to) : 2020/02/16

  Role : Lecturer

  Sponser, Organizer, Publisher  : 朝日小学生新聞

  Event, Program, Title : 朝日小学生新聞
• 謎のウィルス・パニック エボラ出血熱

  Date (from-to) : 2020/02

  Role : Coverage cooperation

  Sponser, Organizer, Publisher  : TBS

  Event, Program, Title : ワールド極限ミステリー
• エボラ治療薬開発への取り組み

  Date (from-to) : 2020/01/24

  Role : Lecturer

  Sponser, Organizer, Publisher  : 宮崎大学

  Event, Program, Title : 宮崎大学 感染症公開セミナー
• ウイルスは悪者か？

  Date (from-to) : 2019/09/10

  Role : Lecturer

  Sponser, Organizer, Publisher  : 読売新聞

  Event, Program, Title : サイエンス読書カフェ
• エボラウイルスを見つける／診断する／治療する

  Date (from-to) : 2019/06/17

  Role : Lecturer

  Sponser, Organizer, Publisher  : JICA

  Event, Program, Title : TICAD関連セミナー
• インハンド

  Date (from-to) : 2019/06

  Role : Advisor

  Sponser, Organizer, Publisher  : TBS

  Event, Program, Title : 金曜ドラマ
• 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

• 報道ランナー

  Date : 2022/07/26

  Publisher, broadcasting station: 関西テレビ

  Program, newspaper magazine: 報道ランナー
• エボラ10分で判定 北大など開発診断キット 国内製造販売承認 感染地域普及目指す

  Date : 2021/04/07

  Program, newspaper magazine: 北海道新聞

  Paper
• デンカと北大、エボラウイルス抗原迅速診断キットの国内製造販売承認を取得

  Date : 2021/03/23

  Program, newspaper magazine: 日本経済新聞

  Paper
• 世界の果てまでも追い続ける 「ウイルスハンター」と呼ばれる２人の日本人研究者

  Date : 2020/09/16

  Publisher, broadcasting station: 朝日新聞

  Program, newspaper magazine: 朝日新聞GLOBE+

  Internet
• 「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
• エボラ熱、日本勢も対策 コンゴで感染拡大 北海道大学検査キット試作 アンジェス新薬の開発進む

  Date : 2019/08/12

  Publisher, broadcasting station: 日本経済新聞

  Program, newspaper magazine: 日本経済新聞

  Paper
• エボラ出血熱の治療薬開発を目指して 人類史上、最も危険なウイルスに挑む

  Date : 2019/08/01

  Program, newspaper magazine: DIAMONDハーバード・ビジネス・レビュー

  Paper
• 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
• Speedy Ebola tests help contain Africa's latest outbreak

  Date : 2018/06/11

  Publisher, broadcasting station: Nature

  Program, newspaper magazine: Nature

  Paper
• 「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

Academic Contribution

• PLoS Neglected Tropical Diseases

  Date (from-to) :2020-Today

  Role: Planning etc

  Type: Peer review etc
• Scientific Reports

  Date (from-to) :2019-Today

  Role: Planning etc

  Type: Peer review etc
• インフルエンザ研究者交流の会

  Date (from-to) :2018-Today

  Role: Planning etc

  Type: Academic society etc
• Viruses

  Date (from-to) :2018-Today

  Role: Planning etc

  Type: Peer review etc
• Vector-Borne and Zoonotic Diseases

  Date (from-to) :2014-Today

  Role: Planning etc

  Type: Peer review etc
• Archives of Virology

  Date (from-to) :2011-Today

  Role: Planning etc

  Type: Peer review etc
• Journal of General Virology

  Date (from-to) :2015-2018

  Role: Planning etc

  Type: Peer review etc
• Influenza Research and Treatment

  Date (from-to) :2009-2017

  Role: Planning etc

  Type: Peer review etc
• Microbiology and Immunology

  Date (from-to) :2005-2007

  Role: Planning etc

  Type: Peer review etc

  Organizer, responsible person: 日本ウイルス学会
• Science Translational Medicine

  Role: Peer review

  Type: Peer review etc
• PLoS Pathogens

  Role: Peer review

  Type: Peer review etc
• Nature Medicine

  Role: Peer review

  Type: Peer review etc
• Nature Communications

  Role: Peer review

  Type: Peer review etc
• Lancet Infectious Diseases

  Role: Peer review

  Type: Peer review etc
• Journal of Virology

  Role: Peer review

  Type: Peer review etc
• Journal of Infectious Diseases

  Role: Peer review

  Type: Peer review etc
• Emerging Infectious Diseases

  Role: Peer review

  Type: Peer review etc
• Cell Reports

  Role: Peer review

  Type: Peer review etc
• Cell

  Role: Peer review

  Type: Peer review etc