研究者データベース
Last Updated :2024/09/08
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プロフィール情報
学位
プロフィール情報
姓
森松名
組子ID各種
200901099501828291
通称等の別名
- 吉松 組子
業績リスト
研究キーワード
- ブニヤウイルス 腎症候性出血熱 HFRS ワクチン 核蛋白 実験動物 診断法 マウス 抗原提示 診断抗原 カイコ CKD インテグリン 感染モデル 膜蛋白 抗原性 エピトープ 組換えバキュロウイルス 組み換え抗原 外皮蛋白 ペプチドワクチン 細胞融合 モノクローナル抗体 中和抗体 レセプター 精製法 感染防御 エンベロープ 血清学 齧歯類 人獣共通感染症 ハンタウイルス serology zoonosis hantavirus
研究分野
- ライフサイエンス / 衛生学、公衆衛生学分野：実験系を含まない
- ライフサイエンス / 衛生学、公衆衛生学分野：実験系を含む
- ライフサイエンス / 医療管理学、医療系社会学
- ライフサイエンス / ウイルス学
- ライフサイエンス / 応用分子細胞生物学
- ライフサイエンス / 獣医学
- ライフサイエンス / 実験動物学
経歴
- 2019年07月 - 現在 北海道大学遺伝子病制御研究所 附属動物実験施設 准教授
- 2011年05月 - 2019年06月 北海道大学 医学(系)研究科(研究院) 微生物学免疫学分野 准教授
- 1997年05月 - 2011年05月 北海道大学医学部 附属動物実験施設 助教
- 1989年 - 1997年 北海道大学免疫科学研究所 教務職員
学歴
論文
- Shelly Wulandari, Samuel Nyampong, Sithumini M.W. Lokupathirage, Kumiko Yoshimatsu, Hiroshi Shimoda, Daisuke HayasakaVirology 598 110170 - 110170 2024年10月
- Tomokazu Tamura, Takashi Irie, Sayaka Deguchi, Hisano Yajima, Masumi Tsuda, Hesham Nasser, Keita Mizuma, Arnon Plianchaisuk, Saori Suzuki, Keiya Uriu, Mst Monira Begum, Ryo Shimizu, Michael Jonathan, Rigel Suzuki, Takashi Kondo, Hayato Ito, Akifumi Kamiyama, Kumiko Yoshimatsu, Maya Shofa, Rina Hashimoto, Yuki Anraku, Kanako Terakado Kimura, Shunsuke Kita, Jiei Sasaki, Kaori Sasaki-Tabata, Katsumi Maenaka, Naganori Nao, Lei Wang, Yoshitaka Oda, Hirofumi Sawa, Ryoko Kawabata, Yukio Watanabe, Ayaka Sakamoto, Naoko Yasuhara, Tateki Suzuki, Yukari Nakajima, Zannatul Ferdous, Kenji Shishido, Yuka Mugita, Otowa Takahashi, Kimiko Ichihara, Yu Kaku, Naoko Misawa, Ziyi Guo, Alfredo Hinay, Yusuke Kosugi, Shigeru Fujita, Jarel M. Tolentino, Luo Chen, Lin Pan, Mai Suganami, Mika Chiba, Ryo Yoshimura, Kyoko Yasuda, Keiko Iida, Naomi Ohsumi, Adam P. Strange, Yuki Shibatani, Tomoko Nishiuchi, Shiho Tanaka, Olivia Putri, Gustav Joas, Yoonjin Kim, Daichi Yamasoba, Kazuhisa Yoshimura, Kenji Sadamasu, Mami Nagashima, Hiroyuki Asakura, Isao Yoshida, So Nakagawa, Akifumi Takaori-Kondo, Kotaro Shirakawa, Kayoko Nagata, Ryosuke Nomura, Yoshihito Horisawa, Yusuke Tashiro, Yugo Kawai, Takamasa Ueno, Chihiro Motozono, Mako Toyoda, Terumasa Ikeda, Akatsuki Saito, Keita Matsuno, Jumpei Ito, Shinya Tanaka, Kei Sato, Takao Hashiguchi, Kazuo Takayama, Takasuke FukuharaNature Communications 15 1 2024年02月08日Abstract Circulation of SARS-CoV-2 Omicron XBB has resulted in the emergence of XBB.1.5, a new Variant of Interest. Our phylogenetic analysis suggests that XBB.1.5 evolved from XBB.1 by acquiring the S486P spike (S) mutation, subsequent to the acquisition of a nonsense mutation in ORF8. Neutralization assays showed similar abilities of immune escape between XBB.1.5 and XBB.1. We determine the structural basis for the interaction between human ACE2 and the S protein of XBB.1.5, showing similar overall structures between the S proteins of XBB.1 and XBB.1.5. We provide the intrinsic pathogenicity of XBB.1 and XBB.1.5 in hamsters. Importantly, we find that the ORF8 nonsense mutation of XBB.1.5 resulted in impairment of MHC suppression. In vivo experiments using recombinant viruses reveal that the XBB.1.5 mutations are involved with reduced virulence of XBB.1.5. Together, our study identifies the two viral functions defined the difference between XBB.1 and XBB.1.5.
- Tomokazu Tamura, Hayato Ito, Shiho Torii, Lei Wang, Rigel Suzuki, Shuhei Tusjino, Akifumi Kamiyama, Yoshitaka Oda, Yuhei Morioka, Saori Suzuki, Kotaro Shirakawa, Kei Sato, Kumiko Yoshimatsu, Yoshiharu Matsuura, Satoshi Iwano, Shinya Tanaka, Takasuke Fukuhara2023年10月13日Summary Monitoringin vivoviral dynamics can improve our understanding of pathogenicity and tissue tropism. For positive-sense, single-stranded RNA viruses, several studies have attempted to monitor viral kineticsin vivousing reporter genomes. The application of such recombinant viruses can be limited by challenges in accommodating bioluminescent reporter genes in the viral genome. Conventional luminescence also exhibits relatively low tissue permeability and thus less sensitivity for visualizationin vivo. Here we show that unlike NanoLuc bioluminescence, the improved method, termed AkaBLI, allows visualization of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in Syrian hamsters. By successfully incorporating a codon-optimized Akaluc luciferase gene into the SARS-CoV-2 genome, we visualizedin vivoinfection, including the tissue-specific differences associated with particular variants. Additionally, we could evaluate the efficacy of neutralizing antibodies and mRNA vaccination by monitoring changes in Akaluc signals. Overall, AkaBLI is an effective technology for monitoring viral dynamics in live animals.
- Izumi Kimura, Daichi Yamasoba, Hesham Nasser, Hayato Ito, Jiri Zahradnik, Jiaqi Wu, Shigeru Fujita, Keiya Uriu, Jiei Sasaki, Tomokazu Tamura, Rigel Suzuki, Sayaka Deguchi, Arnon Plianchaisuk, Kumiko Yoshimatsu, Yasuhiro Kazuma, Shuya Mitoma, Gideon Schreiber, Hiroyuki Asakura, Mami Nagashima, Kenji Sadamasu, Kazuhisa Yoshimura, Akifumi Takaori-Kondo, Jumpei Ito, Kotaro Shirakawa, Kazuo Takayama, Takashi Irie, Takao Hashiguchi, So Nakagawa, Takasuke Fukuhara, Akatsuki Saito, Terumasa Ikeda, Kei SatoJournal of virology e0101123 2023年10月05日Previous studies on the Omicron BA.2 variant suggested that the virological characteristics of BA.2 are determined by the mutations in at least two different regions of the viral genome: in the BA.2 spike gene (enhancing viral fusogenicity and intrinsic pathogenicity) and the non-spike region of the BA.2 genome (leading to intrinsic pathogenicity attenuation). However, the mutations modulating the BA.2 virological properties remain elusive. In this study, we demonstrated that the L371F substitution in the BA.2 spike protein confers greater fusogenicity and intrinsic pathogenicity. Furthermore, we revealed that multiple mutations downstream of the spike gene in the BA.2 genome are responsible for attenuating intrinsic viral pathogenicity and replication capacity. As mutations in the SARS-CoV-2 variant spike proteins could modulate certain virological properties, such as immune evasion and infectivity, most studies have previously focused on spike protein mutations. Our results underpin the importance of non-spike protein-related mutations in SARS-CoV-2 variants. IMPORTANCE Most studies investigating the characteristics of emerging SARS-CoV-2 variants have been focusing on mutations in the spike proteins that affect viral infectivity, fusogenicity, and pathogenicity. However, few studies have addressed how naturally occurring mutations in the non-spike regions of the SARS-CoV-2 genome impact virological properties. In this study, we proved that multiple SARS-CoV-2 Omicron BA.2 mutations, one in the spike protein and another downstream of the spike gene, orchestrally characterize this variant, shedding light on the importance of Omicron BA.2 mutations out of the spike protein.
- Tomokazu Tamura, Daichi Yamasoba, Yoshitaka Oda, Jumpei Ito, Tomoko Kamasaki, Naganori Nao, Rina Hashimoto, Yoichiro Fujioka, Rigel Suzuki, Lei Wang, Hayato Ito, Yukie Kashima, Izumi Kimura, Mai Kishimoto, Masumi Tsuda, Hirofumi Sawa, Kumiko Yoshimatsu, Yuki Yamamoto, Tetsuharu Nagamoto, Jun Kanamune, Yutaka Suzuki, Yusuke Ohba, Isao Yokota, Keita Matsuno, Kazuo Takayama, Shinya Tanaka, Kei Sato, Takasuke FukuharaCommunications biology 6 1 772 - 772 2023年07月24日The unremitting emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants necessitates ongoing control measures. Given its rapid spread, the new Omicron subvariant BA.5 requires urgent characterization. Here, we comprehensively analyzed BA.5 with the other Omicron variants BA.1, BA.2, and ancestral B.1.1. Although in vitro growth kinetics of BA.5 was comparable among the Omicron subvariants, BA.5 was much more fusogenic than BA.1 and BA.2. Airway-on-a-chip analysis showed that, among Omicron subvariants, BA.5 had enhanced ability to disrupt the respiratory epithelial and endothelial barriers. Furthermore, in our hamster model, in vivo pathogenicity of BA.5 was slightly higher than that of the other Omicron variants and less than that of ancestral B.1.1. Notably, BA.5 gains efficient virus spread compared with BA.1 and BA.2, leading to prompt immune responses. Our findings suggest that BA.5 has low pathogenicity compared with the ancestral strain but enhanced virus spread /inflammation compared with earlier Omicron subvariants.
- Regina Amanda Fonseka, Pavani Senarathne, Devinda Shameera Muthusinghe, Nishantha Nanayakkara, Lishantha Gunaratne, Kumiko Yoshimatsu, Nobuo Koizumi, Chandika Damesh Gamage2023年06月01日
- Tomokazu Tamura, Jumpei Ito, Keiya Uriu, Jiri Zahradnik, Izumi Kida, Yuki Anraku, Hesham Nasser, Maya Shofa, Yoshitaka Oda, Spyros Lytras, Naganori Nao, Yukari Itakura, Sayaka Deguchi, Rigel Suzuki, Lei Wang, Mst Monira Begum, Shunsuke Kita, Hisano Yajima, Jiei Sasaki, Kaori Sasaki-Tabata, Ryo Shimizu, Masumi Tsuda, Yusuke Kosugi, Shigeru Fujita, Lin Pan, Daniel Sauter, Kumiko Yoshimatsu, Saori Suzuki, Hiroyuki Asakura, Mami Nagashima, Kenji Sadamasu, Kazuhisa Yoshimura, Yuki Yamamoto, Tetsuharu Nagamoto, Gideon Schreiber, Katsumi Maenaka, Takao Hashiguchi, Terumasa Ikeda, Takasuke Fukuhara, Akatsuki Saito, Shinya Tanaka, Keita Matsuno, Kazuo Takayama, Kei SatoNature communications 14 1 2800 - 2800 2023年05月16日 [査読有り]
In late 2022, SARS-CoV-2 Omicron subvariants have become highly diversified, and XBB is spreading rapidly around the world. Our phylogenetic analyses suggested that XBB emerged through the recombination of two cocirculating BA.2 lineages, BJ.1 and BM.1.1.1 (a progeny of BA.2.75), during the summer of 2022. XBB.1 is the variant most profoundly resistant to BA.2/5 breakthrough infection sera to date and is more fusogenic than BA.2.75. The recombination breakpoint is located in the receptor-binding domain of spike, and each region of the recombinant spike confers immune evasion and increases fusogenicity. We further provide the structural basis for the interaction between XBB.1 spike and human ACE2. Finally, the intrinsic pathogenicity of XBB.1 in male hamsters is comparable to or even lower than that of BA.2.75. Our multiscale investigation provides evidence suggesting that XBB is the first observed SARS-CoV-2 variant to increase its fitness through recombination rather than substitutions. - Jumpei Ito, Rigel Suzuki, Keiya Uriu, Yukari Itakura, Jiri Zahradnik, Kanako Terakado Kimura, Sayaka Deguchi, Lei Wang, Spyros Lytras, Tomokazu Tamura, Izumi Kida, Hesham Nasser, Maya Shofa, Mst Monira Begum, Masumi Tsuda, Yoshitaka Oda, Tateki Suzuki, Jiei Sasaki, Kaori Sasaki-Tabata, Shigeru Fujita, Kumiko Yoshimatsu, Hayato Ito, Naganori Nao, Hiroyuki Asakura, Mami Nagashima, Kenji Sadamasu, Kazuhisa Yoshimura, Yuki Yamamoto, Tetsuharu Nagamoto, Jin Kuramochi, Gideon Schreiber, Akatsuki Saito, Keita Matsuno, Kazuo Takayama, Takao Hashiguchi, Shinya Tanaka, Takasuke Fukuhara, Terumasa Ikeda, Kei SatoNature communications 14 1 2671 - 2671 2023年05月11日 [査読有り]
In late 2022, various Omicron subvariants emerged and cocirculated worldwide. These variants convergently acquired amino acid substitutions at critical residues in the spike protein, including residues R346, K444, L452, N460, and F486. Here, we characterize the convergent evolution of Omicron subvariants and the properties of one recent lineage of concern, BQ.1.1. Our phylogenetic analysis suggests that these five substitutions are recurrently acquired, particularly in younger Omicron lineages. Epidemic dynamics modelling suggests that the five substitutions increase viral fitness, and a large proportion of the fitness variation within Omicron lineages can be explained by these substitutions. Compared to BA.5, BQ.1.1 evades breakthrough BA.2 and BA.5 infection sera more efficiently, as demonstrated by neutralization assays. The pathogenicity of BQ.1.1 in hamsters is lower than that of BA.5. Our multiscale investigations illuminate the evolutionary rules governing the convergent evolution for known Omicron lineages as of 2022. - So Shinya, Devinda S. Muthusinghe, Nobuo Koizumi, Kumiko Yoshimatsu, Shouta M. M. Nakayama, Mayumi Ishizuka, Yoshinori IkenakaToxics 11 4 2023年04月11日Wild rodents are natural hosts of Leptospira spp. and are exposed to various pesticides, some of which are immunotoxic. Rodent urine is an important source of infection for humans and other animals. We evaluated the effects of pesticide exposure on Leptospira growth in mice. Diazinon, at doses of 0.2, 1, and 5 mg/kg/day, was orally administered continuously to mice infected with Leptospira interrogans serogroup Hebdomadis for 32 days. The numbers of L. interrogans in urine and kidney tissues were significantly lower in mice exposed to 5 mg/kg/day diazinon than in unexposed mice (p < 0.05). The urinary concentration of 2-isopropyl-6-methyl-4-pyrimidinol, the metabolite of diazinon, was comparable with the concentration at which viability of L. interrogans was decreased in in vitro assay, suggesting that it had toxic effects on L. interrogans in the proximal renal tubules. Diazinon exposure reinforced Leptospira-induced expression of inflammatory cytokine genes in kidney tissues, and an enhanced immune system might suppress Leptospira growth. These results suggest that diazinon exposure may not increase the risk of Leptospira transmission to humans through mice. This novel study evaluated the relationship between pesticide exposure and Leptospira infection in mice, and the results could be useful for risk assessment of leptospirosis.
- Chandika Damesh Gamage, Shanika Nanayakkara, Yomani D. Sarathkumara, Devinda S. Muthusinghe, Kenta Shimizu, Jiro Arikawa, Sithumini M. W. Lokupathirage, Nishantha Nanayakkara, Lishanthe Gunarathne, Rohana Chandrajith, Kouji H. Harada, Akio Koizumi, Kumiko YoshimatsuJournal of Medical Microbiology 71 12 2022年12月16日 [査読有り][通常論文]
- Akatsuki Saito, Tomokazu Tamura, Jiri Zahradnik, Sayaka Deguchi, Koshiro Tabata, Yuki Anraku, Izumi Kimura, Jumpei Ito, Daichi Yamasoba, Hesham Nasser, Mako Toyoda, Kayoko Nagata, Keiya Uriu, Yusuke Kosugi, Shigeru Fujita, Maya Shofa, Mst Monira Begum, Ryo Shimizu, Yoshitaka Oda, Rigel Suzuki, Hayato Ito, Naganori Nao, Lei Wang, Masumi Tsuda, Kumiko Yoshimatsu, Jin Kuramochi, Shunsuke Kita, Kaori Sasaki-Tabata, Hideo Fukuhara, Katsumi Maenaka, Yuki Yamamoto, Tetsuharu Nagamoto, Hiroyuki Asakura, Mami Nagashima, Kenji Sadamasu, Kazuhisa Yoshimura, Takamasa Ueno, Gideon Schreiber, Akifumi Takaori-Kondo, Kotaro Shirakawa, Hirofumi Sawa, Takashi Irie, Takao Hashiguchi, Kazuo Takayama, Keita Matsuno, Shinya Tanaka, Terumasa Ikeda, Takasuke Fukuhara, Kei SatoCell host & microbe 30 11 1540 - 1555 2022年11月09日The SARS-CoV-2 Omicron BA.2.75 variant emerged in May 2022. BA.2.75 is a BA.2 descendant but is phylogenetically distinct from BA.5, the currently predominant BA.2 descendant. Here, we show that BA.2.75 has a greater effective reproduction number and different immunogenicity profile than BA.5. We determined the sensitivity of BA.2.75 to vaccinee and convalescent sera as well as a panel of clinically available antiviral drugs and antibodies. Antiviral drugs largely retained potency, but antibody sensitivity varied depending on several key BA.2.75-specific substitutions. The BA.2.75 spike exhibited a profoundly higher affinity for its human receptor, ACE2. Additionally, the fusogenicity, growth efficiency in human alveolar epithelial cells, and intrinsic pathogenicity in hamsters of BA.2.75 were greater than those of BA.2. Our multilevel investigations suggest that BA.2.75 acquired virological properties independent of BA.5, and the potential risk of BA.2.75 to global health is greater than that of BA.5.
- Izumi Kimura, Daichi Yamasoba, Tomokazu Tamura, Naganori Nao, Tateki Suzuki, Yoshitaka Oda, Shuya Mitoma, Jumpei Ito, Hesham Nasser, Jiri Zahradnik, Keiya Uriu, Shigeru Fujita, Yusuke Kosugi, Lei Wang, Masumi Tsuda, Mai Kishimoto, Hayato Ito, Rigel Suzuki, Ryo Shimizu, Mst Monira Begum, Kumiko Yoshimatsu, Kanako Terakado Kimura, Jiei Sasaki, Kaori Sasaki-Tabata, Yuki Yamamoto, Tetsuharu Nagamoto, Jun Kanamune, Kouji Kobiyama, Hiroyuki Asakura, Mami Nagashima, Kenji Sadamasu, Kazuhisa Yoshimura, Kotaro Shirakawa, Akifumi Takaori-Kondo, Jin Kuramochi, Gideon Schreiber, Ken J Ishii, Takao Hashiguchi, Terumasa Ikeda, Akatsuki Saito, Takasuke Fukuhara, Shinya Tanaka, Keita Matsuno, Kei SatoCell 185 21 3992 - 4007 2022年10月13日 [査読有り]
After the global spread of the SARS-CoV-2 Omicron BA.2, some BA.2 subvariants, including BA.2.9.1, BA.2.11, BA.2.12.1, BA.4, and BA.5, emerged in multiple countries. Our statistical analysis showed that the effective reproduction numbers of these BA.2 subvariants are greater than that of the original BA.2. Neutralization experiments revealed that the immunity induced by BA.1/2 infections is less effective against BA.4/5. Cell culture experiments showed that BA.2.12.1 and BA.4/5 replicate more efficiently in human alveolar epithelial cells than BA.2, and particularly, BA.4/5 is more fusogenic than BA.2. We further provided the structure of the BA.4/5 spike receptor-binding domain that binds to human ACE2 and considered how the substitutions in the BA.4/5 spike play roles in ACE2 binding and immune evasion. Moreover, experiments using hamsters suggested that BA.4/5 is more pathogenic than BA.2. Our multiscale investigations suggest that the risk of BA.2 subvariants, particularly BA.4/5, to global health is greater than that of original BA.2. - Zhouoxing Wei, Kenta Shimizu, Rakiiya S. Sarii, Devinda S. Muthusinghe, Sithumini M. W. Lokupathirage, Junko Nio-Kobayashi, Kumiko YoshimatsuViruses 14 10 2247 - 2247 2022年10月13日 [査読有り]
Hantaan virus is the causative agent of hemorrhagic fever with renal syndrome (HFRS). The Hantaan virus strain, Korean hemorrhagic fever virus clone-5 (KHF5), causes weight loss and renal hemorrhage in laboratory mice. Clone-4 (KHF4), which has a single E417K amino acid change in its glycoprotein, is an avirulent variant. In this study, KHF4 and KHF5 were compared to evaluate pathological differences in mice in vitro and in vivo. The characteristics of the two glycoproteins were not significantly different in vitro. However, the virulent KHF5 strain targeted the lungs and caused pneumonia and edema in vivo. Both strains induced high infectivity levels in the liver and caused hepatitis; however, petechial hemorrhage and glycogen storage reduction were observed in KHF5-infected mice alone. Renal hemorrhage was observed using viral antigens in the tubular region of KHF5-infected mice. In addition, an increase in white blood cell levels and neutrophilia were found in KHF5-infected mice. Microarray analysis of liver cells showed that CD8+ T cell activation, acute-phase protein production, and neutrophil activation was induced by KHF5 infection. KHF5 infectivity was significantly increased in vivo and the histological and clinicopathological findings were similar to those in patients with HFRS. - Shinsuke Toba, Akihiko Sato, Makoto Kawai, Yoshiyuki Taoda, Yuto Unoh, Shinji Kusakabe, Haruaki Nobori, Shota Uehara, Kentaro Uemura, Keiichi Taniguchi, Masanori Kobayashi, Takeshi Noshi, Ryu Yoshida, Akira Naito, Takao Shishido, Junki Maruyama, Slobodan Paessler, Michael J Carr, William W Hall, Kumiko Yoshimatsu, Jiro Arikawa, Keita Matsuno, Yoshihiro Sakoda, Michihito Sasaki, Yasuko Orba, Hirofumi Sawa, Hiroshi KidaProceedings of the National Academy of Sciences of the United States of America 119 36 e2206104119 2022年09月06日 [査読有り]
Viral hemorrhagic fevers caused by members of the order Bunyavirales comprise endemic and emerging human infections that are significant public health concerns. Despite the disease severity, there are few therapeutic options available, and therefore effective antiviral drugs are urgently needed to reduce disease burdens. Bunyaviruses, like influenza viruses (IFVs), possess a cap-dependent endonuclease (CEN) that mediates the critical cap-snatching step of viral RNA transcription. We screened compounds from our CEN inhibitor (CENi) library and identified specific structural compounds that are 100 to 1,000 times more active in vitro than ribavirin against bunyaviruses, including Lassa virus, lymphocytic choriomeningitis virus (LCMV), and Junin virus. To investigate their inhibitory mechanism of action, drug-resistant viruses were selected in culture. Whole-genome sequencing revealed that amino acid substitutions in the CEN region of drug-resistant viruses were located in similar positions as those of the CEN α3-helix loop of IFVs derived under drug selection. Thus, our studies suggest that CENi compounds inhibit both bunyavirus and IFV replication in a mechanistically similar manner. Structural analysis revealed that the side chain of the carboxyl group at the seventh position of the main structure of the compound was essential for the high antiviral activity against bunyaviruses. In LCMV-infected mice, the compounds significantly decreased blood viral load, suppressed symptoms such as thrombocytopenia and hepatic dysfunction, and improved survival rates. These data suggest a potential broad-spectrum clinical utility of CENis for the treatment of both severe influenza and hemorrhagic diseases caused by bunyaviruses. - Daichi Yamasoba, Izumi Kimura, Hesham Nasser, Yuhei Morioka, Naganori Nao, Jumpei Ito, Keiya Uriu, Masumi Tsuda, Jiri Zahradnik, Kotaro Shirakawa, Rigel Suzuki, Mai Kishimoto, Yusuke Kosugi, Kouji Kobiyama, Teppei Hara, Mako Toyoda, Yuri L Tanaka, Erika P Butlertanaka, Ryo Shimizu, Hayato Ito, Lei Wang, Yoshitaka Oda, Yasuko Orba, Michihito Sasaki, Kayoko Nagata, Kumiko Yoshimatsu, Hiroyuki Asakura, Mami Nagashima, Kenji Sadamasu, Kazuhisa Yoshimura, Jin Kuramochi, Motoaki Seki, Ryoji Fujiki, Atsushi Kaneda, Tadanaga Shimada, Taka-Aki Nakada, Seiichiro Sakao, Takuji Suzuki, Takamasa Ueno, Akifumi Takaori-Kondo, Ken J Ishii, Gideon Schreiber, Hirofumi Sawa, Akatsuki Saito, Takashi Irie, Shinya Tanaka, Keita Matsuno, Takasuke Fukuhara, Terumasa Ikeda, Kei SatoCell 185 12 2103 - 2115 2022年06月09日 [査読有り]
Soon after the emergence and global spread of the SARS-CoV-2 Omicron lineage BA.1, another Omicron lineage, BA.2, began outcompeting BA.1. The results of statistical analysis showed that the effective reproduction number of BA.2 is 1.4-fold higher than that of BA.1. Neutralization experiments revealed that immunity induced by COVID vaccines widely administered to human populations is not effective against BA.2, similar to BA.1, and that the antigenicity of BA.2 is notably different from that of BA.1. Cell culture experiments showed that the BA.2 spike confers higher replication efficacy in human nasal epithelial cells and is more efficient in mediating syncytia formation than the BA.1 spike. Furthermore, infection experiments using hamsters indicated that the BA.2 spike-bearing virus is more pathogenic than the BA.1 spike-bearing virus. Altogether, the results of our multiscale investigations suggest that the risk of BA.2 to global health is potentially higher than that of BA.1. - Rigel Suzuki, Daichi Yamasoba, Izumi Kimura, Lei Wang, Mai Kishimoto, Jumpei Ito, Yuhei Morioka, Naganori Nao, Hesham Nasser, Keiya Uriu, Yusuke Kosugi, Masumi Tsuda, Yasuko Orba, Michihito Sasaki, Ryo Shimizu, Ryoko Kawabata, Kumiko Yoshimatsu, Hiroyuki Asakura, Mami Nagashima, Kenji Sadamasu, Kazuhisa Yoshimura, Hirofumi Sawa, Terumasa Ikeda, Takashi Irie, Keita Matsuno, Shinya Tanaka, Takasuke Fukuhara, Kei SatoNature 603 7902 700 - 705 2022年03月 [査読有り]
The emergence of the Omicron variant of SARS-CoV-2 is an urgent global health concern1. In this study, our statistical modelling suggests that Omicron has spread more rapidly than the Delta variant in several countries including South Africa. Cell culture experiments showed Omicron to be less fusogenic than Delta and than an ancestral strain of SARS-CoV-2. Although the spike (S) protein of Delta is efficiently cleaved into two subunits, which facilitates cell-cell fusion2,3, the Omicron S protein was less efficiently cleaved compared to the S proteins of Delta and ancestral SARS-CoV-2. Furthermore, in a hamster model, Omicron showed decreased lung infectivity and was less pathogenic compared to Delta and ancestral SARS-CoV-2. Our multiscale investigations reveal the virological characteristics of Omicron, including rapid growth in the human population, lower fusogenicity and attenuated pathogenicity. - Sithumini M W Lokupathirage, Yoshimi Tsuda, Kodai Ikegame, Kisho Noda, Devinda S Muthusinghe, Fumiya Kozawa, Rashid Manzoor, Kenta Shimizu, Kumiko YoshimatsuScientific reports 11 1 22977 - 22977 2021年11月26日 [査読有り]
Severe fever with thrombocytopenia syndrome virus (SFTSV) is an emerging bunyavirus that causes novel zoonotic diseases in Asian countries including China, Japan, South Korea, and Vietnam. In phleboviruses, viral proteins play a critical role in viral particle formation inside the host cells. Viral glycoproteins (GPs) and RNA-dependent RNA polymerase (RdRp) are colocalized in the Golgi apparatus and endoplasmic reticulum-Golgi intermediate compartment (ERGIC). The nucleocapsid (N) protein was widely expressed in the cytoplasm, even in cells coexpressing GP. However, the role of SFTSV N protein remains unclear. The subcellular localization of SFTSV structural proteins was investigated using a confocal microscope. Subsequently, minigenome and immunoprecipitation assays were carried out. The N protein interacts with viral RNA (vRNA) and further shows translational activity with RdRp which is L protein and localized in the ERGIC and Golgi apparatus when co-expressed with GP. On the other hand, mutant N protein did not interact with vRNA either localized in the ERGIC or Golgi apparatus. The interaction between the N protein of SFTSV and vRNA is important for the localization of viral proteins and viral assembly. This study provides useful insights into the life cycle of SFTSV, which will lead to the detection of antiviral targets. - Akatsuki Saito, Takashi Irie, Rigel Suzuki, Tadashi Maemura, Hesham Nasser, Keiya Uriu, Yusuke Kosugi, Kotaro Shirakawa, Kenji Sadamasu, Izumi Kimura, Jumpei Ito, Jiaqi Wu, Kiyoko Iwatsuki-Horimoto, Mutsumi Ito, Seiya Yamayoshi, Samantha Loeber, Masumi Tsuda, Lei Wang, Seiya Ozono, Erika P Butlertanaka, Yuri L Tanaka, Ryo Shimizu, Kenta Shimizu, Kumiko Yoshimatsu, Ryoko Kawabata, Takemasa Sakaguchi, Kenzo Tokunaga, Isao Yoshida, Hiroyuki Asakura, Mami Nagashima, Yasuhiro Kazuma, Ryosuke Nomura, Yoshihito Horisawa, Kazuhisa Yoshimura, Akifumi Takaori-Kondo, Masaki Imai, Shinya Tanaka, So Nakagawa, Terumasa Ikeda, Takasuke Fukuhara, Yoshihiro Kawaoka, Kei SatoNature 602 7896 300 - 306 2021年11月25日 [査読有り]
During the current SARS-CoV-2 pandemic, a variety of mutations have accumulated in the viral genome, and currently, four variants of concern (VOCs) are considered potentially hazardous to human society1. The recently emerged B.1.617.2/Delta VOC is closely associated with the COVID-19 surge that occurred in India in the spring of 20212. However, its virological properties remain unclear. Here, we show that the B.1.617.2/Delta variant is highly fusogenic and notably more pathogenic than prototypic SARS-CoV-2 in infected hamsters. The P681R mutation in the spike protein, which is highly conserved in this lineage, facilitates spike protein cleavage and enhances viral fusogenicity. Moreover, we demonstrate that the P681R-bearing virus exhibits higher pathogenicity than its parental virus. Our data suggest that the P681R mutation is a hallmark of the virological phenotype of the B.1.617.2/Delta variant and is associated with enhanced pathogenicity. - Devinda S. Muthusinghe, Kenta Shimizu, Sithumini M, W. Lokupathirage, Zhouoxing Wei, Yomani D. Sarathkumara, G. R. Am, a Fonseka, Pavani Senarathne, Nobuo Koizumi, Tomonori Kawakami, Akio Koizumi, Chaminda Wickramasinghe, Hideki Ebihara, Keita Matsuno, Yoshimi Tsuda, Jiro Arikawa, Chandika D. Gamage, Kumiko YoshimatsuViruses 13 10 1984 - 1984 2021年10月02日 [査読有り]
- 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 YoshimatsuPLoS neglected tropical diseases 15 6 e0009452 2021年06月01日 [査読有り]
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. - Shumpei P Yasuda, Kenta Shimizu, Takaaki Koma, Nguyen Thuy Hoa, Mai Quynh Le, Zhuoxing Wei, Devinda S Muthusinghe, Sithumini M W Lokupathirage, Futoshi Hasebe, Tetsu Yamashiro, Jiro Arikawa, Kumiko YoshimatsuViruses 13 4 2021年04月12日 [査読有り]
To clarify the mechanism of Seoul orthohantavirus (SEOV) persistence, we compared the humoral and cell-mediated immune responses to SEOV in experimentally and naturally infected brown rats. Rats that were experimentally infected by the intraperitoneal route showed transient immunoglobulin M (IgM) production, followed by an increased anti-SEOV immunoglobulin G (IgG) antibody response and maturation of IgG avidity. The level of SEOV-specific cytotoxic T lymphocytes (CTLs) peaked at 6 days after inoculation and the viral genome disappeared from serum. In contrast, naturally infected brown rats simultaneously had a high rate of SEOV-specific IgM and IgG antibodies (28/43). Most of the IgM-positive rats (24/27) had the SEOV genome in their lungs, suggesting that chronic SEOV infection was established in those rats. In female rats with IgG avidity maturation, the viral load in the lungs was decreased. On the other hand, there was no relationship between IgG avidity and viral load in the lungs in male rats. A CTL response was not detected in naturally infected rats. The difference between immune responses in the experimentally and naturally infected rats is associated with the establishment of chronic infection in natural hosts. - Kristina T Lu, Takuya Yamamoto, David McDonald, Wei Li, Marissa Tan, Meng Ling Moi, Eun-Chung Park, Kumiko Yoshimatsu, Marie Ricciardone, Allan Hildesheim, Yukari Totsuka, Asuka Nanbo, Opass Putcharoen, Gompol Suwanpimolkul, Watsamon Jantarabenjakul, Leilani Paitoonpong, F Gray Handley, K Gayle Bernabe, Masahiko Noda, Miwa Sonoda, Patrick Brennan, Diane E Griffin, Ichiro KuraneVirology 555 71 - 77 2021年03月This review summarizes the presentations given at the 22nd International conference on Emerging Infectious Diseases in the Pacific Rim. The purpose of this annual meeting is to foster international collaborations and address important public health issues in the Asia-Pacific region. This meeting was held in Bangkok in February 2020 and focused on emerging virus infections. Unexpectedly, the SARS-CoV-2 pandemic was in the initial stages leading to a special session on COVID-19 in addition to talks on dengue, influenza, hepatitis, AIDS, Zika, chikungunya, rabies, cervical cancer and nasopharyngeal carcinoma.
- Zhouoxing Wei, Kenta Shimizu, Kumpei Nishigami, Yoshimi Tsuda, Yomani Sarathukumara, Devinda S Muthusinghe, Chandika D Gamage, Lishanta Granathne, Sithumini M W Lokupathirage, Nishanta Nanayakkara, Jiro Arikawa, Fuka Kikuchi, Keiko Tanaka-Taya, Motoi Suzuki, Shigeru Morikawa, Satoru Arai, Kumiko YoshimatsuArchives of virology 166 1 275 - 280 2021年01月 [査読有り]
The infectivity of shrew-borne hantaviruses to humans is still unclear because of the lack of a serodiagnosis method for these viruses. In this study, we prepared recombinant nucleocapsid (rN) proteins of Seewis orthohantavirus, Altai orthohantavirus (ALTV), Thottapalayam thottimvirus (TPMV), and Asama orthohantavirus. Using monospecific rabbit sera, no antigenic cross-reactivity was observed. In a serosurvey of 104 samples from renal patients and 271 samples from heathy controls from Sri Lanka, one patient serum and two healthy control sera reacted with rN proteins of ALTV and TPMV, respectively. The novel assays should be applied to investigate potential infectivity of shrew-borne hantaviruses to humans. - Kisho Noda, Yoshimi Tsuda, Fumiya Kozawa, Manabu Igarashi, Kenta Shimizu, Jiro Arikawa, Kumiko YoshimatsuViruses 13 1 2020年12月27日 [査読有り]
Severe fever with thrombocytopenia syndrome virus subclone B7 shows strong plaque formation and cytopathic effect induction compared with other subclones and the parental strain YG1. Compared to YG1 and the other subclones, only B7 possesses a single substitution in the L protein at the amino acid position 1891, in which N is changed to K (N1891K). In this study, we evaluate the effects of this mutation on L protein activity via a cell-based minigenome assay. Substitutions of N with basic amino acids (K or R) enhanced polymerase activity, while substitutions with an acidic amino acid (E) decreased this activity. Mutation to other neutral amino acids showed no significant effect on activity. These results suggest that the characteristic of the amino acid at position 1891 of the L protein are critical for its function, especially with respect to the charge status. Our data indicate that this C-terminal domain of the L protein may be crucial to its functions in genome transcription and viral replication. - Noriko Tosa, Tomoko Ishida, Kumiko Yoshimatsu, Nobuhito Hayashimoto, Kanae Shiokawa, Akira Takakura, Jiro ArikawaExperimental animals 2020年11月12日 [査読有り]
Rapid and simple serologic tests that require only a small amount of blood without the euthanization of animals are valuable for microbial control in colonies of laboratory animals. In this study, we developed a multiplex immunochromatographic assay (ICA) for detection of antibodies to Sendai virus (also known as hemagglutinating virus of Japan), hantavirus, and sialodacryoadenitis virus, which are causative agents of major infectious diseases in rats. For this assay, an ICA strip was placed into a microtube containing 150 μl PBS and either 0.75 μl of rat serum or 1.5 μl of whole blood. Binding antibodies were visualized by using anti-rat IgG antibody-conjugated colloidal gold. Under these conditions, the multiplex ICA simultaneously and specifically detected antibodies to multiple antigens. Positive serum samples for each infectious disease were used to evaluate the sensitivity and specificity of the multiplex ICA. The sensitivities of the multiplex ICA for Sendai virus, hantavirus, and sialodacryoadenitis virus were 100%, 100%, and 81%, respectively. No nonspecific reactions were observed in any of the 52 positive sera against heterologous antigens. In addition, 10 samples of uninfected sera did not show any bands except for the control line. These observations indicate high specificity of the multiplex ICA. Moreover, the multiplex ICA could be applied to diluted blood. These results indicate that the multiplex ICA is appropriate for rapid and simple serological testing of laboratory rats. - 抗核蛋白質抗体を用いた各種ハンタウイルス抗原の新規検出法の確立三橋 健斗, 小林 進太郎, 好井 健太朗, 吉松 組子, 苅和 宏明日本獣医学会学術集会講演要旨集 163回 253 - 253 (公社)日本獣医学会 2020年10月
- Lilani Karunanayake, Chandika D Gamage, Chandima P Gunasekara, Sajiv De Silva, Hidemasa Izumiya, Masatomo Morita, Devinda S Muthusinghe, Kumiko Yoshimatsu, Roshan Niloofa, Panduka Karunanayake, Wimalasiri Uluwattage, Makoto Ohnishi, Nobuo KoizumiPLoS neglected tropical diseases 14 8 e0008573 2020年08月 [査読有り][通常論文]
BACKGROUND: Leptospirosis has gained much attention in Sri Lanka since its large outbreak in 2008. However, most of the cases were clinically diagnosed and information on Leptospira genotypes and serotypes currently prevailing in the country is lacking. METHODOLOGY/PRINCIPAL FINDINGS: We retrospectively analyzed 24 Leptospira strains from human patients as well as isolated and characterized three Leptospira strains from black rats using the microscopic agglutination test with antisera for 19 serovars and multilocus sequence typing. The isolates were identified as Leptospira borgpetersenii sequence types (STs) 143 and 144; L. interrogans STs 30, 34, 43, 44, 74, 75, 80, 308, 313, 314, 316, and 317; and L. kirschneri ST318. Six of the 15 STs were identified for the first time in this study. Five serogroups such as Autumnalis, Grippotyphosa, Hebdomadis, Javanica, and Pyrogenes were detected among the isolates. Contrary to previous studies, various genotypes including novel STs were isolated during an outbreak in Southern Province. L. borgpetersenii serogroup Javanica ST143 was isolated both from a human and black rat. CONCLUSIONS/SIGNIFICANCE: This study revealed that genetically diverse Leptospira strains currently circulate in Sri Lanka: some genotypes have been circulating and others have emerged recently, which may explain the recent surge of leptospirosis patients with varying clinical manifestations and frequent outbreaks of leptospirosis. Black rats were identified as the source of infection for humans, but reservoir animals for other genotypes remain unknown. - Satoko Mori, Thidalay Thwe, Wai Min Thu, Shumpei P. Yasuda, Saw Bawm, Kimiyuki Tsuchiya, Ken Katakura, Satoru Arai, Kumiko Yoshimatsu, Hitoshi SuzukiMammal Research 65 3 493 - 502 2020年07月 [査読有り][通常論文]
- [Hantavirus infection as a risk for chronic kidney disease of unknown etiology (CKDu) in Sri Lanka].Kumiko YoshimatsuUirusu 70 2 175 - 184 2020年 [査読無し]
Chronic kidney disease of unknown etiology (CKDu) has emerged in endemic areas of Sri Lanka since the 1990s. The disease is a chronic but fatal disease. Until now, heavy metals and agrochemicals have been suspected as the cause of CKDu, but it has been still unknown. Recently, we have found a high seroprevalence to hantavirus in CKDu patients and reported that hantavirus infection is a risk of CKDu. Hantaviruses are rodent-borne zoonotic viruses. Here, I would like to introduce a story of the research from sero-epidemiology to the search for host animals. - Y D Sarathkumara, Chandika Damesh Gamage, Sithumini Lokupathirage, Devinda S. Muthusinghe, Nishantha Nanayakkara, Lishanthe Gunarathne, Kenta Shimizu, Yoshimi Tsuda, Jiro Arikawa, YoshimatsuViruses 11 8 2019年12月11日 [査読有り][通常論文]
- Noriko Tosa, Tomoko Ishida, Kumiko Yoshimatsu, Nobuhito Hayashimoto, Kanae Shiokawa, Akira Takakura, Jiro ArikawaJournal of the American Association for Laboratory Animal Science : JAALAS 58 6 790 - 795 2019年11月01日 [査読有り]
Serologic monitoring of infectious diseases is important for microbial control in colonies of laboratory mice. Rapid and simple tests that do not require killing animals are valuable for this purpose. In this study, we developed a multiplex immunochromatographic assay (ICA) for detection of antibodies to mouse hepatitis virus (MHV), Sendai virus (also known as hemagglutinating virus of Japan [HVJ]), and Clostridium piliforme (The pathogen that causes Tyzzer disease), which are major infectious diseases in mice. For this assay, an ICA strip was put into a microtube containing 150 μL PBS and either 0.75 μL mouse serum or 1.5 μL whole blood. Binding antibodies were visualized by using protein A-conjugated colloidal gold. Under these conditions, multiplex ICA simultaneously and specifically detected antibodies to multiple antigens. To evaluate the sensitivity and specificity of multiplex ICA, positive serum samples for each infectious disease were used. Sensitivities of the multiplex ICA test for MHV, HVJ, and C. piliforme were 100%, 100%, and 90%, respectively. No nonspecific reaction was observed in any of the 30 positive sera. In addition, 10 samples of uninfected sera did not show any bands except for the control line. These observations indicate high specificity of the multiplex ICA test. Moreover, the multiplex ICA could be applied to diluted blood. These results indicate that the multiplex ICA is appropriate for rapid, simple, and safe serologic testing of laboratory mice. - Lokupathirage, S.M.W., Muthusinghe, D.S., Shimizu, K., Nishigami, K., Noda, K., Tsuda, Y., Sarathkumara, Y.D., Gunawardana, S., Arikawa, J., Gamage, C.D., Yoshimatsu, K.Vector-Borne and Zoonotic Diseases 19 11 2019年07月 [査読有り][通常論文]
- Development of a multiplex immunochromatographic test for serological diagnosis of major infectious diseases in laboratory miceTosa N, Ishida T, Yoshimatsu K, Hayashimoto N, Kanae S, Takakura A, Arikawa JJ Am Assoc Lab Anim Sci 2019年06月 [査読有り][通常論文]
- Yoshimatsu, K., Gamage, C.D., Sarathkumara, Y.D., Kulendiran, T., Muthusinghe, D.S., Nanayakkara, N., Gunarathne, L., Shimizu, K., Tsuda, Y., Arikawa, J.Archives of Virology 164 1 267 - 271 2019年01月04日 [査読有り][通常論文]
- 土佐紀子Biomedical Research 40 1 9 - 15 2019年 [査読有り][通常論文]
Can f 1 belongs to the lipocalin superfamily and is considered to be an animal allergen. The immune response induced by Can f 1 in mice was compared with that induced by ovalbumin (OVA), a typical food allergen. Female BALB/c and C57BL/6 mice (6 weeks of age) were subcutaneously injected with Can f 1 or OVA with or without aluminum hydroxide (Alum) three times with intervals of two weeks. Serum levels of total IgE or antigen-specific IgE and production of IL13 and IFNγ from splenocytes were analyzed. Immunization with Can f 1 or OVA increased serum levels of both total IgE and antigen-specific IgE significantly irrespective of Alum. These results indicate that Can f 1 and OVA were able to induce allergic sensitization in mice. Splenocyte production of IL13 in mice immunized with Can f 1 or OVA with and without Alum were significantly increased after stimulation with each antigen. However, IL13 levels in the mice immunized with Can f 1 with Alum were significantly lower than those immunized without Alum. Increases in IFNγ levels after stimulation with Can f 1 or OVA were not remarkable. No influence of genetic backgrounds of BALB/c and C57BL/6 mice was found. Although Can f 1 induced Th2 type immune responses as was also the case for immunization with OVA, an inhibitory effect of Alum on induction of IL13 was observed only in mice immunized with Can f 1. These results suggest that the immune mechanism for allergic sensitization with Can f 1 is different from that with OVA. - Gamage, C., Sarathkumara, Y., Weerakkodi, V., Shiokawa, K., Yoshimatsu, K., Arikawa, J.Journal of Immunological Methods 463 134 - 136 2018年09月 [査読有り][通常論文]
- Kenta Shimizu, Kumiko Yoshimatsu, Midori Taruishi, Yoshimi Tsuda, Jiro ArikawaArchives of Virology 163 6 1577 - 1584 2018年06月01日 [査読有り][通常論文]
Hemorrhagic fever with renal syndrome (HFRS) is caused by hantavirus infection. Although host immunity is thought to be involved in the pathogenesis of HFRS, the mechanism remains to be elucidated. A mouse model of HFRS, which showed renal hemorrhage similar to that seen in patients, has been developed previously. In this study, we aimed to clarify whether CD4+ and CD8+ T cells are involved in the development of renal hemorrhage in the mouse model. At 2 days before virus inoculation, CD4+ or CD8+ T cells in 6-week-old BALB/c mice were depleted by administration of antibodies. The CD4+ T cell-depleted mice developed signs of disease such as transient weight loss, ruffled fur and renal hemorrhage as in non-depleted mice. In contrast, the CD8+ T cell-depleted mice showed no signs of disease. After determination of CTL epitopes on the viral glycoprotein in BALB/c mice, the quantity of virus-specific CTLs was analyzed using an MHC tetramer. The quantity of virus-specific CTLs markedly increased in spleens and kidneys of virus-infected mice. However, the quantity in high-pathogenic clone-infected mice was comparable to that in low-pathogenic clone-infected mice. We previously reported that the high-pathogenic clone propagated more efficiently than the low-pathogenic clone in kidneys of mice during the course of infection. Therefore, there is a possibility that the balance between quantities of the target and effector is important for disease outcome. In conclusion, this study showed that CD8+ T cells are involved in the development of renal hemorrhage in a mouse model of HFRS. - Ngonda Saasa, Masahiro Kajihara, George Dautu, Akina Mori-Kajihara, Shuetsu Fukushi, Yona Sinkala, Shigeru Morikawa, Aaron Mweene, Ayato Takada, Kumiko Yoshimatsu, Jiro ArikawaVector-Borne and Zoonotic Diseases 18 5 273 - 277 2018年05月01日 [査読有り][通常論文]
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. - Tapiwa LUNDU, Yoshimi TSUDA, Ryo ITO, Kenta SHIMIZU, Shintaro KOBAYASHI, Kentaro YOSHII, Kumiko YOSHIMATSU, Jiro ARIKAWA, Hiroaki KARIWABiomedical Research 39 1 27 - 38 2018年 [査読有り][通常論文]
- Sanae Nishio, Yoshimi Tsuda, Ryo Ito, Kenta Shimizu, Kumiko Yoshimatsu, Jiro ArikawaJAPANESE JOURNAL OF INFECTIOUS DISEASES 70 4 388 - 393 2017年07月 [査読有り][通常論文]
The first clinical case of the YG1 strain of the severe fever with thrombocytopenia syndrome virus (SFTSV) has been isolated in Japan. We found that only some of the cells underwent low pH-dependent cell fusion, although all of the cells were confirmed to have been infected with the virus. This suggested that the YG1 strain consists of a heterogeneous mixture of related viruses. Here, we established 3 subclones (termed E3, A4, and B7) from the YG1 strain, using the limiting dilution method with the pH-dependent cell fusion activity. Subclone E3 showed weak fusion activity and cytopathic effects (CPE) in Vero E6 cells. The amino acid sequence of E3 was identical to the published sequence for the YG1 strain, and it likely comprises a subpopulation of the YG1 strain. Subclone A4 displayed strong fusion activity under acidic conditions. In contrast, subclone B7 showed strong fusion activity and CPE under neutral and acidic conditions. Two amino acid differences shared between B7 and A4 were found in the envelope glycoproteins. In addition, an amino acid variant of the RNA-dependent RNA polymerase was found only in B7. These subclones will be valuable tools to elucidate cell fusion mechanisms of SFTSV and the relationship between viral proteins and their functions. - Kenta Shimizu, Rie Isozumi, Kazutoshi Takami, Isao Kimata, Kanae Shiokawa, Kumiko Yoshimatsu, Yoshimi Tsuda, Sanae Nishio, Jiro ArikawaJOURNAL OF VETERINARY MEDICAL SCIENCE 79 7 1261 - 1263 2017年07月 [査読有り][通常論文]
We examined 33 rodents captured in an urban area of Osaka City, Japan for IgG antibodies against Seoul virus, severe fever with thrombocytopenia syndrome virus, hepatitis E virus, Leptospira interrogans, Yersinia pestis, spotted fever, typhus and scrub typhus group rickettsiae. We found that 3 (9.1%) and 1 (3.0%) of the 33 rodents had antibodies against L. interrogans and spotted fever group rickettsiae, respectively. DNAs of leptospires were detected from 2 of the 3 seropositive rodents, but DNA of rickettsia was not detected. Phylogenetic analysis and multiple locus sequence typing revealed that the 2 leptospires were L. interrogans belonging to a novel sequence type. There is a potential risk for acquiring rodent-borne zoonotic pathogens even in cities in developed countries. - Chandika D. Gamage, Kumiko Yoshimatsu, Yomani D. Sarathkumara, Thiviyaaluxmi Kulendiran, Nishantha Nanayakkara, Jiro ArikawaINTERNATIONAL JOURNAL OF INFECTIOUS DISEASES 57 77 - 78 2017年04月 [査読有り][通常論文]
- Kumiko Yoshimatsu, Satoru Arai, Kenta Shimizu, Yoshimi Tsuda, Bazartseren Boldgiv, Bazartseren Boldbaatar, Erdenebaatar Sergelen, Dagvatseren Ariunzaya, Orsoo Enkhmandal, Sukhbaatar Tuvshintugs, Shigeru Morikawa, Jiro ArikawaJAPANESE JOURNAL OF VETERINARY RESEARCH 65 1 39 - 44 2017年02月 [査読有り][通常論文]
Seroepizootiological surveys among wild rodents were carried out on the east side of Lake Khovsgol in Mongolia in 2010 and 2011. A total of 76 voles belonging to the genera Myodes and Microtus were captured. Most of the voles that were seropositive to Tula virus antigen were Middendorf's voles (Microtus middendorffii (6/31)). Two of the 18 Myodes voles were also seropositive to Tula virus antigen. On the other hand, only one vole was seropositive to Puumala virus antigen. The results suggest that Tula virus was maintained in Middendorf's vole. This is the first report of detection of anti-Tula virus antibody in the central part of the Eurasia continent. - Sanae Nishio, Yoshimi Tsuda, Ryo Ito, Kenta Shimizu, Kumiko Yoshimatsu, Jiro ArikawaJapanese Journal of Infectious Diseases 70 4 388 - 393 2017年 [査読有り][通常論文]
SUMMARY: The first clinical case of the YG1 strain of the severe fever with thrombocytopenia syndrome virus (SFTSV) has been isolated in Japan. We found that only some of the cells underwent low pH-dependent cell fusion, although all of the cells were confirmed to have been infected with the virus. This suggested that the YG1 strain consists of a heterogeneous mixture of related viruses. Here, we established 3 subclones (termed E3, A4, and B7) from the YG1 strain, using the limiting dilution method with the pH-dependent cell fusion activity. Subclone E3 showed weak fusion activity and cytopathic effects (CPE) in Vero E6 cells. The amino acid sequence of E3 was identical to the published sequence for the YG1 strain, and it likely comprises a subpopulation of the YG1 strain. Subclone A4 displayed strong fusion activity under acidic conditions. In contrast, subclone B7 showed strong fusion activity and CPE under neutral and acidic conditions. Two amino acid differences shared between B7 and A4 were found in the envelope glycoproteins. In addition, an amino acid variant of the RNA-dependent RNA polymerase was found only in B7. These subclones will be valuable tools to elucidate cell fusion mechanisms of SFTSV and the relationship between viral proteins and their functions. - Satomu Obana, Kenta Shimizu, Kumiko Yoshimatsu, Futoshi Hasebe, Kozue Hotta, Rie Isozumi, Hoa Thuy Nguyen, Mai Quynh Le, Tetsu Yamashiro, Yoshimi Tsuda, Jiro ArikawaJOURNAL OF VETERINARY MEDICAL SCIENCE 79 1 76 - 81 2017年01月 [査読有り][通常論文]
There is concern about the zoonotic potential of rodent-borne hepatitis E virus, designated as HEV-C1. However, epizootiological information about HEV-C1 is limited. To address this issue, serum samples from 443 small mammals captured at 5 sites in Hanoi, Vietnam, were examined for anti-HEV-Cl IgG antibodies. In addition, livers of seropositive animals were examined for viral RNA. Anti-HEV-C1 antibodies were detected in 57 (12.9%) of the 443 serum samples. Seropositive animals were found in all of the sites (4.7% to 22.2%). Anti-HEV-C1 antibodies were detected from 48 (12.3%) of 389 Rattus norvegicus and 9 (19.6%) of 46 R. tanezumi, but were not detected from 8 Suncus murinus. Viral RNAs were detected from 13 (22.8%) of the 57 seropositive rodents. The detectioh rate of viral RNA in seropositive R. tanezumi (66.7%, 6/9) was significantly higher than that in seropositive R. norvegicus (14.6%, 7/48). The results suggest that R. tanezumi is more susceptible than R. norvegicus to HEV-Cl infection. Phylogenetic analysis revealed that Vietnamese strains were divided into 3 clusters in genetic group 2 of HEV-C1. Multiple clusters of viruses were detected at several sites without species specificity, suggesting that 3 clusters of HEV-C1 co-circulate in Hanoi, Vietnam. - Kenta Shimizu, Takaaki Koma, Kumiko Yoshimatsu, Yoshimi Tsuda, Yuji Isegawa, Jiro ArikawaVIROLOGY JOURNAL 14 1 13 2017年01月 [査読有り][通常論文]
Background: Hemorrhagic fever with renal syndrome (HFRS) caused by hantavirus infection is characterized by fever, renal dysfunction and hemorrhage. An animal model mimicking symptoms of HFRS remains to be established. In this study, we evaluated the pathogenicity of an HFRS patient-derived Hantaan virus (HTNV) in adult mice. Methods: Five clones of HTNV strain KHF 83-61 BL (KHFV) that was derived from blood of an HFRS patient were obtained by plaque cloning. The pathogenicity of the virus clones was evaluated by using 6-week-old female BALB/c mice. Sequence analysis of the viral genome was performed by conventional methods. Results: All of the mice intravenously inoculated with KHFV clone (cl)-1, -2, -3 and -5 showed signs of disease such as transient body weight loss, ruffled fur, reduced activity and remarkably prominent hemorrhage in the renal medulla at 6 to 9 days post-inoculation (dpi) and then recovered. In contrast, mice intravenously inoculated with KHFV cl-4 did not show any signs of disease. We selected KHFV cl-5 and cl-4 as representative of high-pathogenic and low-pathogenic clones, respectively. Quantities of viral RNA in kidneys of KHFV cl-5-infected mice were larger than those in KHFV cl-4-infected mice at any time point examined (3, 6, 9 and 12 dpi). The quantities of viral RNA of KHFV cl-5 and cl-4 peaked at 3 dpi, which was before the onset of disease. Sequence analysis revealed that the amino acid at position 417 in the glycoprotein Gn was the sole difference in viral proteins between KHFV cl-5 and cl-4. The result suggests that amino acid at position 417 in Gn is related to the difference in pathogenicity between KHFV cl-5 and cl-4. When the inoculum of KHFV cl-5 was pretreated with a neutralizing antibody against HTNV strain 76-118, which belongs to the same serotype as KHFV clones, mice did not show any signs of disease, confirming that the disease was caused by KHFV infection. Conclusion: We found that an HFRS patient-derived HTNV caused renal hemorrhage in adult mice. We anticipate that this infection model will be a valuable tool for understanding the pathogenesis of HFRS. - Yoshimi Tsuda, Manabu Igarashi, Ryo Ito, Sanae Nishio, Kenta Shimizu, Kumiko Yoshimatsu, Jiro ArikawaBIOMEDICAL RESEARCH-TOKYO 38 2 89 - 97 2017年 [査読有り][通常論文]
Severe fever with thrombocytopenia syndrome virus (SFTSV) is a novel phlebovirus responsible for causing an emerging zoonotic disease. We previously established subclones from SFTSV strain YG1 based on differences in low-pH-dependent cell fusion activities and found two amino acid substitutions, Y328H and R624W, in the envelope glycoprotein (GP) of high fusion subclones. In this study, we show that transiently expressed GP with the R624W mutation, but not the Y328H mutation, induced cell fusion under acidic conditions. GP possessing either tryptophan, serine, glycine or aspartic acid at position 624 induced cell fusion, whereas GP possessing basic amino acids such as arginine or lysine did not induce cell fusion. These results indicated that the amino acid at position 624 has an important role for inducing low-pH-dependent cell fusion. - Kenta Shimizu, Sugihiro Hamaguchi, Cuong Chi Ngo, Tian-Cheng Li, Shuji Ando, Kumiko Yoshimatsu, Shumpei P. Yasuda, Takaaki Koma, Rie Isozumi, Yoshimi Tsuda, Hiromi Fujita, Thuy Thanh Pham, Mai Quynh Le, Anh Duc Dang, Tuan Quang Nguyen, Lay-Myint Yoshida, Koya Ariyoshi, Jiro ArikawaJOURNAL OF VETERINARY MEDICAL SCIENCE 78 11 1677 - 1681 2016年11月 [査読有り][通常論文]
Zoonotic potential of a rat-derived hepatitis E virus (HEV), designated as HEV-Cl, remains unknown. To evaluate the risk for HEV-Cl infection in humans, paired sera of 208 hospitalized febrile patients collected from 2001 to 2003 in Hanoi, Vietnam, were examined for IgG antibodies to HEV-Cl and genotype 1 HEV (HEV-1), which is common in humans. IgG antibodies to virus-like particles (VLPs) of HEV-Cl and/or HEV-1 were detected from 99 of the 208 convalescent sera in enzyme-linked immunosorbent assay (ELISA). IgG antibody titers to HEV-Cl antigen in 3 of the 99 sera were more than 8-fold higher than those to HEV-1 antigen. IgM antibodies to HEV-Cl antigen were detected in acute sera from 2 of the 3 patients in ELISA and Western blotting. However, no HEV genome was detected. Clinical information was available for 1 of the 2 patients. Hepatic enzymes, aspartate aminotransferase and alanine aminotransferase, were mildly elevated (156 IU/l and 68 IU/l, respectively), and hepatomegaly was detected by ultrasonography. The patient recovered from the illness after 17 days. These results indicated that HEV-Cl or its variants infect humans in Vietnam and may cause acute febrile illness with mild liver dysfunction. - Yu Koarashi, Abraham G. Caceres, Florencia Margarita Zuniga Saca, Elsa Elvira Palacios Flores, Adela Celis Trujillo, Jose Luis Abanto Alvares, Kumiko Yoshimatsu, Jiro Arikawa, Ken Katakura, Yoshihisa Hashiguchi, Hirotomo KatoACTA TROPICA 158 83 - 87 2016年06月 [査読有り][通常論文]
A PCR-Restriction Fragment Length Polymorphism (RFLP) targeting the mannose phosphate isomerase gene was established to differentiate Leishmania species distributed near the Department of Huanuco, Peru. The technique was applied to 267 DNA samples extracted from Giemsa-stained smears of cutaneous lesions taken from patients suspected for cutaneous leishmaniasis in the area, and the present status of causative Leishmania species was identified. Of 114 PCR-amplified samples, 22, 19, 24 and 49 samples were identified to be infected by Leishmania (Viannia) braziliensis, L. (V.) peruviana, L. (V.) guyanensis, and a hybrid of L. (V.) braziliensis/L. (V.) peruviana, respectively, and the validity of PCR-RFLP was confirmed by sequence analysis. Since PCR-RFLP is simple and rapid, the technique will be a useful tool for the epidemiological study of leishmaniasis. (C) 2016 Elsevier B.V. All rights reserved. - Kanae Shiokawa, Chandika D. Gamage, Nobuo Koizumi, Yoshihiro Sakoda, Kenta Shimizu, Yoshimi Tsuda, Kumiko Yoshimatsu, Jiro ArikawaJOURNAL OF VETERINARY MEDICAL SCIENCE 78 2 221 - 230 2016年02月 [査読有り][通常論文]
The applicability of the recombinant LipL32 for serodiagnosis of leptospiral infection in field rodents was assessed in this study. An immunodominant region of LipL32 was determined by monoclonal antibodies, and then, truncated LipL32 (tLipL32) was designed to contain the region (87-188th amino acid). The tLipL32 was compared between two recombinant expression hosts Escherichia coli and Pichia pastoris in ELISA. With field rat sera, tLipL32 expressed by P. pastoris (tLipL32p) had high antigenicity without background reactions, while tLipL32 expressed by E. coli (tLipL32e) showed high background reactions, which were reduced by pre-adsorption of sera with E. coli. To evaluate tLipL32-ELISA, field rat sera were tentatively divided into a Leptospira infection positive (12 sera) and a negative group (12 sera) based on the results from flaB gene PCR of kidney samples and WB with whole Leptospira cell. Consequently, the sensitivity of tLipL32p-ELISA for field rat sera was 83%. A similar result was obtained from tLipL32e-ELISA with adsorbed sera, (92%). However, sensitivity of tLipL32e-ELISA using sera without an adsorption treatment was 50%. Regardless of the expression host, tLipL32-ELISA had 100% specificity and sensitivity in experimentally infected laboratory rats. These results suggest that recombinant LipL32 expressed by P pastoris is more applicable for serodiagnosis in field rats due to a lack of background reaction. - Nobuo Koizumi, Hidemasa Izumiya, Jung-Jung Mu, Zbigniew Arent, Shou Okano, Chie Nakajima, Yasuhiko Suzuki, Maki Mizutani Muto, Tsutomu Tanikawa, Kyle R. Taylor, Noriyuki Komatsu, Kumiko Yoshimatsu, Hoang Thi Thu Ha, Makoto OhnishiINFECTION GENETICS AND EVOLUTION 36 434 - 440 2015年12月 [査読有り][通常論文]
Leptospira spp. are the causative agents of a worldwide zoonosis, leptospirosis, maintained by various mammals. Each Leptospira serovar is frequently associated with a particular maintenance host, and recently, Leptospira genotype-host association has also been suggested to limit serovars to restricted areas. We investigated the molecular characteristics of L. interrogans and L. borgpetersenii which were isolated from small feral and wild animals in four East Asian states using multiple-locus variable-number tandem repeat analysis (MLVA). MLVA using 11 loci was performed on 110 L. interrogans serogroups from Japan (79 strains of 5 serogroups from 3 animal species), Philippines (21; 3; 2), Taiwan (7; 2; 3), and Vietnam (3; 1; 1). A MLVA method using 4 loci for L. borgpetersenii was established and performed on 52 isolates from Japan (26; 3; 7), Philippines (13; 1; 2), and Taiwan (13; 1; 3). In L. interrogans, serogroups Autumnalis and Hebdomadis appeared more genetically diverse than serogroups Bataviae, Grippotyphosa, Icterohaemorrhagiae, Pomona, or Pyrogenes. The former serogroup strains with the exception of one Hebdomadis strain were isolated from Apodemus speciosus while all the latter serogroup strains with the exception of Grippotyphosa were isolated from Rattus norvegicus. L. borgpetersenii was isolated from at least 11 animal species while L. interrogans was isolated from five species, which might suggest a wider host range for L. borgpetersenii. Broad host preference in a single genotype was also observed, which colonized not only different species of the same genera but also multiple animal genera. This study demonstrates that there may be variability in the range of genetic diversity among different Leptospira serogroups, which may be attributed to maintenance host animals and environmental factors. (C) 2015 The Authors. Published by Elsevier B.V. - Jenny Verner-Carlsson, Mare Lõhmus, Karin Sundström, Tanja M. Strand, Monique Verkerk, Chantal Reusken, Kumiko Yoshimatsu, Jiro Arikawa, Frank van de Goot, Åke LundkvistAfrican Journal of Disability 5 1 27215 2015年 [査読有り][通常論文]
We report the first detection of Seoul hantavirus (SEOV)-specific antibodies in the wild brown rat population in the Netherlands. SEOV-reactive antibodies were found in three rats out of 16 in a repeated series of tests including immunofluorescence assay, immunoblot, and enzyme-linked immunosorbent assay. Focus reduction neutralization test confirmed the presence of SEOV-specific antibodies, and reverse-transcription polymerase chain reaction (RT-PCR) confirmed the presence of hantaviral RNA. This discovery follows the recent findings of SEOV infections in wild and pet brown rats and humans in England, Wales, France, Belgium, and Sweden, indicating an even higher importance of this hantavirus for public health in large areas of Europe. - Shumpei P. Yasuda, Chandika D. Gamage, Nobuo Koizumi, Sanae Nishio, Rie Isozumi, Kenta Shimizu, Takaaki Koma, Takako Amada, Hitoshi Suzuki, Kumiko Yoshimatsu, Jiro ArikawaGenes and Genetic Systems 89 2 71 - 80 2014年09月01日 [査読有り][通常論文]
We examined genetic variation in black rats (the Rattus rattus complex) from Kandy District, Sri Lanka using mitochondrial cytochrome b (cytb, 1140 bp) and nuclear melanocortin 1 receptor (Mc1r, 954 bp) gene sequences together with database sequences. We confirmed the existence of two divergent mitochondrial lineages in Sri Lankan black rats, with genetic distance of 2.2% and estimated divergence time of 0.3 million years ago. Because one lineage is unique to the island and the other is closely related to R. rattus populations on the Indian subcontinent, two migration events of R. rattus from the subcontinent are inferred, one ancient and one recent. Mc1r analyses revealed 12 haplotypes among the Sri Lankan black rats. A median-joining network together with other availablesequences separated the 12 haplotypes into two groups, one unique to the island and the other related to previously reported R. rattus sequences. Notably, most individuals possessed various combinations of both haplotype groups which had no association with the cytb clades. These results imply that old and new R. rattuslineages are now intermingled as a result of hybridization in Sri Lanka. Specimensof the lesser bandicoot rat (Bandicota bengalensis) collected from Sri Lanka (n = 24) were shown to have no genetic variability in the cytb sequence. Our results indicate that the two most abundant groups of commensal rats in Sri Lanka, black rats and lesser bandicoot rats, are the product of contrasting evolutionary histories on different timescales. - Kumiko Yoshimatsu, Jiro ArikawaVIRUSES-BASEL 6 8 3097 - 3109 2014年08月 [査読有り][通常論文]
Hantavirus causes two important rodent-borne viral zoonoses, hemorrhagic fever with renal syndrome (HFRS) in Eurasia and hantavirus pulmonary syndrome (HPS) in North and South America. Twenty-four species that represent sero-and genotypes have been registered within the genus Hantavirus by the International Committee on Taxonomy of Viruses (ICTV). Among the viral proteins, nucleocapsid (N) protein possesses an immunodominant antigen. The antigenicitiy of N protein is conserved compared with that of envelope glycoproteins. Therefore, N protein has been used for serological diagnoses and seroepidemiological studies. An understanding of the antigenic properties of N protein is important for the interpretation of results from serological tests using N antigen. N protein consists of about 430 amino acids and possesses various epitopes. The N-terminal quarter of N protein bears linear and immunodominant epitopes. However, a serotype-specific and multimerization-dependent antigenic site was found in the C-terminal half of N protein. In this paper, the structure, function, and antigenicity of N protein are reviewed. - Kumiko Yoshimatsu, Jiro ArikawaVIRUS RESEARCH 187 77 - 83 2014年07月 [査読有り][通常論文]
Hantaviruses are causative agents of two rodent-borne zoonoses, hemorrhagic fever with renal syndrome (HERS) and nephropathia epidemica (NE) in the Old World and hantavirus pulmonary syndrome (HPS) in the New World. Serological examinations to detect hantavirus antibodies have been most widely used for surveillance among humans and rodent reservoirs. Here, we will review antigenic structure of nucleocapsid (N) protein of hantaviruses and application of recombinant N protein as diagnostic antigen for screening and serotyping. (C) 2014 The Authors. Published by Elsevier B.V. All rights reserved. - Takaaki Koma, Kumiko Yoshimatsu, Noriyo Nagata, Yuko Sato, Kenta Shimizu, Shumpei P. Yasuda, Takako Amada, Sanae Nishio, Hideki Hasegawa, Jiro ArikawaJOURNAL OF VIROLOGY 88 13 7178 - 7188 2014年07月 [査読有り][通常論文]
Hantavirus infections are characterized by vascular hyperpermeability and neutrophilia. However, the pathogenesis of this disease is poorly understood. Here, we demonstrate for the first time that pulmonary vascular permeability is increased by Hantaan virus infection and results in the development of pulmonary edema in C. B-17 severe combined immunodeficiency ( SCID) mice lacking functional T cells and B cells. Increases in neutrophils in the lung and blood were observed when pulmonary edema began to be observed in the infected SCID mice. The occurrence of pulmonary edema was inhibited by neutrophil depletion. Moreover, the pulmonary vascular permeability was also significantly suppressed by neutrophil depletion in the infected mice. Taken together, the results suggest that neutrophils play an important role in pulmonary vascular hyperpermeability and the occurrence of pulmonary edema after hantavirus infection in SCID mice. - Takako Amada, Kumiko Yoshimatsu, Takaaki Koma, Kenta Shimizu, Chandika D. Gamage, Kanae Shiokawa, Sanae Nishio, Clas Ahlm, Jiro ArikawaVIROLOGY JOURNAL 11 87 2014年05月 [査読有り][通常論文]
Background: Hantaviruses are causative agents of hemorrhagic fever with renal syndrome (HFRS) and nephropathia epidemica (NE) in the Old World and hantavirus pulmonary syndrome (HPS) in the New World. There is a need for time-saving diagnostic methods. In the present study, recombinant N antigens were used as antigens in an immunochromatography strip (ICG) test to detect specific IgG antibodies. Methods: The N-terminal 103 amino acids (aa) of Hantaan virus (HTNV), Puumala virus (PUUV) and Andes virus (ANDV) nucleocapsid (N) protein were expressed in E. coli as representative antigens of three groups (HFRS, NE and HPS-causing viruses) of hantavirus. Five different types of ICG test strips, one antigen line on one strip for each of the three selected hantaviruses (HTNV, PUUV and ANDV), three antigen lines on one strip and a mixed antigen line on one strip, were developed and sensitivities were compared. Results: A total of 87 convalescent-phase patient sera, including sera from 35 HFRS patients, 36 NE patients and 16 HPS patients, and 25 sera from healthy seronegative people as negative controls were used to evaluate the ICG test. Sensitivities of the three-line strip and mixed-line strip were similar to those of the single antigen strip (97.2 to 100%). On the other hand, all of the ICG test strips showed high specificities to healthy donors. Conclusion: These results indicated that the ICG test with the three representative antigens is an effective serodiagnostic tool for screening and typing of hantavirus infection in humans. - Kenta Shimizu, Kumiko Yoshimatsu, Takaaki Koma, Shumpei P. Yasuda, Jiro ArikawaVirus Research 178 2 349 - 356 2013年12月26日 [査読有り][通常論文]
To understand the role of nucleocapsid protein (NP) of hantaviruses in viral assembly, the effect of NP on intracellular traffic of viral glycoproteins Gn and Gc was investigated. Double staining of viral and host proteins in Hantaan virus (HTNV)-infected Vero E6 cells showed that Gn and Gc were localized to cis-Golgi, in which virus particles are thought to be formed. When HTNV Gn and Gc were expressed by a plasmid encoding glycoprotein precursor (GPC), which is posttranslationaly cleaved into Gn and Gc, Gn was localized to cis-Golgi, whereas Gc showed diffuse distribution in the cytoplasm in 32.9% of Gc-positive cells. The ratio of the diffused Gc-positive cells was significantly decreased to 15.0% by co-expression of HTNV NP. Co-expression of HTNV GPC with NPs of other hantaviruses, such as Seoul virus, Puumala virus and Sin Nombre virus, also reduced the ratios of diffused Gc-positive cells to 13.5%, 25.2%, and 11.6%, respectively. Among amino- and carboxyl-terminally truncated HTNV NPs, NP75-429, NP116-429, NP1-333, NP1-233, and NP1-155 possessed activity to reduce the ratio of diffused Gc-positive cells, while NP155-429 and NP1-116 did not. NP30-429 has partial activity. These results indicate that amino acid region 116-155 of NP is important for the activity, although amino acid region 1-30 is partially related. Truncation of the HTNV Gc cytoplasmic tail caused an increase in diffused Gc-positive cells. In addition, the effect of coexpression of HTNV NP was weakened. These results suggest that HTNV NP has a role to promote Golgi localization of Gc through a mechanism possibly mediated by the Gc cytoplasmic tail. © 2013 Elsevier B.V. - Takako Arnada, Kumiko Yoshimatsu, Shumpei P. Yasuda, Kenta Shimizu, Takaaki Koma, Nobuhito Hayashimoto, Chandika D. Gamage, Sanae Nishio, Akira Takakura, Jiro ArikawaJOURNAL OF VIROLOGICAL METHODS 193 1 42 - 49 2013年10月 [査読有り][通常論文]
Hantavirus is a causative agent of rodent-borne viral zoonoses, hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome. Seoul virus (SEOV) is a causative agent of urban and laboratory rat-associated HFRS worldwide. Surveillance of rodents has been done mainly by serological detection of hantavirus-specific antibodies by enzyme linked immunosorbent assay (ELISA) and immunofluorescent antibody assay (IFA). An immunochromatographic (ICG) test was developed with the N-terminal 103 amino acids of nucleocapsid protein of Hantaan virus expressed by Escherichia coli as an antigen to detect IgG antibody specific to hantavirus in sera from Rattus sp. animals. Antibody-detecting sensitivity of the ICG test was the same as that of ELISA and about 100-times higher than that of IFA. Overall sensitivities and specificities of the ICG test in comparison to ELISA and IFA for sera from 192 urban rats and 123 laboratory rats were 99.3% and 100%, respectively. Diluted whole blood samples without separation could be used for the ICG test. The ICG test enabled detection of antibodies to SEOV, Hantaan, Dobrava/Belgrade, and Thailand viruses, which are causative agents of HFRS throughout Eurasia. The ICG test is a rapid, simple and safe method for diagnosis of SEOV infection in rats. (c) 2013 Elsevier B.V. All rights reserved. - T. Koma, K. Yoshimatsu, S. P. Yasuda, T. Li, T. Amada, K. Shimizu, R. Isozumi, L. T.Q. Mai, N. T. Hoa, V. Nguyen, T. Yamashiro, F. Hasebe, J. ArikawaEpidemiology and Infection 141 9 1876 - 1884 2013年09月 [査読有り][通常論文]
To examine the prevalence of human pathogens carried by rats in urban areas in Hanoi and Hai Phong, Vietnam, we live-trapped 100 rats in January 2011 and screened them for a panel of bacteria and viruses. Antibodies against Leptospira interrogans (22·0%), Seoul virus (14·0%) and rat hepatitis E virus (23·0%) were detected in rats, but antibodies against Yersinia pestis were not detected. Antibodies against L. interrogans and Seoul virus were found only in adult rats. In contrast, antibodies to rat hepatitis E virus were also found in juvenile and sub-adult rats, indicating that the transmission mode of rat hepatitis E virus is different from that of L. interrogans and Seoul virus. Moreover, phylogenetic analyses of the S and M segments of Seoul viruses found in Rattus norvegicus showed that Seoul viruses from Hai Phong and Hanoi formed different clades. Human exposure to these pathogens has become a significant public health concern. Copyright © Cambridge University Press 2012. - T. Koma, K. Yoshimatsu, S. P. Yasuda, T. Li, T. Amada, K. Shimizu, R. Isozumi, L. T.Q. Mai, N. T. Hoa, V. Nguyen, T. Yamashiro, F. Hasebe, J. ArikawaEpidemiology and Infection 141 9 1876 - 1884 2013年09月 [査読有り][通常論文]
To examine the prevalence of human pathogens carried by rats in urban areas in Hanoi and Hai Phong, Vietnam, we live-trapped 100 rats in January 2011 and screened them for a panel of bacteria and viruses. Antibodies against Leptospira interrogans (22·0%), Seoul virus (14·0%) and rat hepatitis E virus (23·0%) were detected in rats, but antibodies against Yersinia pestis were not detected. Antibodies against L. interrogans and Seoul virus were found only in adult rats. In contrast, antibodies to rat hepatitis E virus were also found in juvenile and sub-adult rats, indicating that the transmission mode of rat hepatitis E virus is different from that of L. interrogans and Seoul virus. Moreover, phylogenetic analyses of the S and M segments of Seoul viruses found in Rattus norvegicus showed that Seoul viruses from Hai Phong and Hanoi formed different clades. Human exposure to these pathogens has become a significant public health concern. Copyright © Cambridge University Press 2012. - Ima-Nurisa Ibrahim, Kenta Shimizu, Kumiko Yoshimatsu, Andre Yunianto, Ervi Salwati, Shumpei P. Yasuda, Takaaki Koma, Rika Endo, Jiro ArikawaJOURNAL OF VETERINARY MEDICAL SCIENCE 75 8 1003 - 1008 2013年08月 [査読有り][通常論文]
Hemorrhagic fever with renal syndrome (HFRS) is a rodent-borne zoonotic disease caused by hantavirus infection. Many HFRS cases have been reported in East Asia and North Europe, while the situation in Southeast Asia remains unclear. In this study, the prevalence of hantavirus infection in rodents and humans in Thousand Islands regency, which is close to the port of Jakarta, one of the largest historic ports in Indonesia, was investigated. A total of 170 rodents were captured in 2005, and 27 (15.9%) of the rodents were antibody-positive against Hantaan virus antigen in an immunofluorescence assay (IFA) and Western blotting. Despite the high prevalence in rodents, human sera collected from 31 patients with fever of -unknown origin and 20 healthy volunteers in the islands in 2009 did not show positive reaction to the antigen in IFA. To identify the virus in rodents genetically, a total of 59 rodents were captured in 2009. Sera from the rodents were screened for antibody by ELISA, and lung tissues were subjected to RT-PCR. 20 (33.9%) of the 59 rodents were antibody-positive, and 3 of those 20 rodents were positive for S and M genome segments of hantaviruses. Genetic analysis showed that the viruses belonged to Seoul virus and formed a cluster with those in Vietnam and Singapore. These results suggest that a unique group of Seoul viruses has spread widely in Southeast Asia. - 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 ArikawaJOURNAL OF VETERINARY MEDICAL SCIENCE 75 6 819 - 825 2013年06月 [査読有り][通常論文]
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. - Tian-Cheng Li, Yasushi Ami, Yuriko Suzaki, Shumpei P. Yasuda, Kumiko Yoshimatsu, Jiro Arikawa, Naokazu Takeda, Wakita TakajiEMERGING INFECTIOUS DISEASES 19 1 115 - 118 2013年01月 [査読有り][通常論文]
We amplified the complete genome of the rat hepatitis E virus (HEV) Vietnam strain (V-105) and analyzed the nucleotide and amino acid sequences. The entire genome of V-105 shared only 76.8%-76.9% nucleotide sequence identities with rat HEV strains from Germany, which suggests that V-105 is a new genotype of rat HEV. - Tian-Cheng Li, Sayaka Yoshizaki, Yasushi Ami, Yuriko Suzaki, Shumpei P. Yasuda, Kumiko Yoshimatsu, Jiro Arikawa, Naokazu Takeda, Takaji WakitaVeterinary Microbiology 163 1-2 54 - 61 2013年 [査読有り][通常論文]
To determine whether or not rats are susceptible to hepatitis E virus (HEV) infection, each of group containing three laboratory rats (Wistar) were experimentally inoculated with genotypes 1, 3, 4 and rat HEV by intravenous injection. Serum and stool samples were collected and used to detect HEV RNA and anti-HEV antibodies by RT-PCR and ELISA, respectively. The virus infection was monitored up to 3 months after inoculation. None of the serum or stool samples collected from the rats inoculated with G1, G3, or G4 HEV indicated positive sign for virus replication. Although no alteration was observed in ALT level, rat HEV RNA was detected in stools from both of the rats inoculated with rat HEV, and both rats were positive for anti-rat HEV IgG and IgM from 3 weeks after inoculation. These results demonstrated that rats are susceptible to rat HEV but not to G1, G3, and G4 HEV. We also confirm that the nude rats were useful for obtaining a large amount of rat HEV and that the rat HEV was transmitted by the fecal-oral route. © 2012 Elsevier B.V. - 神戸 嘉一, 鈴木 莊介, 矢部 辰男, 中田 勝士, 前園 泰徳, 阿部 愼太郎, 石田 健, 谷川 力, 橋本 琢磨, 武田 美加子, 土屋 公幸, 吉松 組子, 鈴木 仁哺乳類科学 53 2 289 - 299 The Mammal Society of Japan 2013年クマネズミはthe Rattus rattus species complexとも称され，複数の種からなる種複合体である．日本には古くに移入した東アジア地域起源のRattus tanezumi（2n＝42）に加え，新規に移入したインド地域が起源のR. rattus（2n＝38）の2系統が存在する．本研究ではこれらクマネズミ系統の日本列島における分布および移入の歴史を把握することを試みた．毛色関連遺伝子Mc1r（954 bp）をマーカーとし，奄美大島産を含む36個体の塩基配列データを新規に収集し，既存の配列データと合わせ，日本列島の17地点，さらに比較対象として用いたパキスタン産を含め，総計133個体のデータを基に系統学的解析を行った．その結果，小樽，小笠原諸島および東京の3地域でR. rattus型が認められ，これらの地点ではR. tanezumi型とのヘテロ接合体も存在した．これらの結果から，既存系統への浸透交雑が一部の市街部，港湾部および離島で進行している実態が明示された．一方，琉球列島の自然林では，R. rattus型のMc1rハプロタイプは認められなかった．これは，新たな外来系統R. rattusの定着や浸透交雑を起こさない何らかの要因が存在する可能性を示唆する．琉球列島には独自のMc1r配列の存在も認められ，他地域とは遺伝的に分化した集団として位置づけられる可能性も示唆された．
- 新世界ハンタウイルス感染のためのハンタウイルス組み換え核蛋白を用いた血清型鑑別ELISA法の開発駒 貴明, 吉松 組子, 垂石 みどり, 宮下 大輔, 遠藤 理香, 清水 健太, 安田 俊平, 天田 貴子, 瀬戸 隆, 村田 亮, 吉田 喜香, 苅和 宏明, 高島 郁夫, 有川 二郎北海道医学雑誌 88 1 35 - 35 2013年01月 [査読無し][通常論文]
- Variation in the bacterial conditions inside cages is correlated with intracage humidity and ammonia levels.Tosa N, Yoshimatsu K, Tadasuke Tsukiyama, Hatakeyama S, Arikawa JLab Animal and Environ 21 2 87 - 98 2013年 [査読有り][通常論文]
- 吉松組子, 有川二郎ウイルス 62 2 239 - 250 日本ウイルス学会 2012年12月 [査読無し][通常論文]
ブニヤウイルスは，ブニヤウイルス科オルソブニヤウイルス属，ハンタウイルス属，ナイロウイルス属，フレボウイルス属およびトスポウイルス属に分類されるウイルスの総称である．植物に病原性を有するトスポウイルス属以外は，脊椎動物に感染し，人や動物に重篤な疾患を引き起こす．いずれも，医学・獣医学・農学領域で重要な疾病であり，その多くが人獣共通感染症である（図1）．ハンタウイルス属以外は，節足動物をベクターとするアルボウイルスであるが，自然界における感染環には属間で相違がある．近年，ハンタウイルスの自然宿主としてげっ歯目以外にトガリネズミ目の動物が重要な役割を担っていることが明らかになった．また，フレボウイルス属のウイルスを原因とし，血小板減少と発熱を特徴とする重篤な疾患が中国で新たに出現し1,2），その後，米国でも存在が確認され新興感染症として注目されている． - Mathias Schlegel, Erdenesaikhan Tegshduuren, Kumiko Yoshimatsu, Rasa Petraityte, Kestutis Sasnauskas, Baerbel Hammerschmidt, Robert Friedrich, Marc Mertens, Martin H. Groschup, Satoru Arai, Rika Endo, Kenta Shimizu, Takaaki Koma, Shumpei Yasuda, Chiaki Ishihara, Rainer G. Ulrich, Jiro Arikawa, Bernd KoellnerARCHIVES OF VIROLOGY 157 11 2179 - 2187 2012年11月 [査読有り][通常論文]
We developed serological tools for the detection of hantavirus-specific antibodies and hantavirus antigens in shrews. The work was focussed to generate Thottapalayam virus (TPMV)-specific monoclonal antibodies (mAbs) and anti-shrew immunoglobulin G (IgG) antibodies. The mAbs against TPMV nucleocapsid (N) protein were produced after immunization of BALB/c mice with recombinant TPMV N proteins expressed in Escherichia coli, baculovirus and Saccharomyces cerevisiae-mediated expression systems. In total, six TPMV N-protein-specific mAbs were generated that showed a characteristic fluorescent pattern in indirect immunofluorescence assay (IFA) using TPMV-infected Vero cells. Out of the six mAbs tested, five showed no cross-reaction to rodent-associated hantaviruses (Hantaan, Seoul, Puumala, Tula, Dobrava-Belgrade and Sin Nombre viruses) in IFA and enzyme-linked immunosorbent assay (ELISA), although one mAb reacted to Sin Nombre virus in IFA. None of the mAbs cross-reacted with an amino-terminal segment of the shrew-borne Asama virus N protein. Anti-shrew-IgG sera were prepared after immunization of rabbits and BALB/c-mice with protein-G-purified shrew IgG. TPMV-N-protein-specific sera were raised by immunisation of Asian house shrews (Suncus murinus) with purified yeast-expressed TPMV N protein. Using these tools, an indirect ELISA was developed to detect TPMV-N-protein-specific antibodies in the sera of shrews. Using an established serological assay, high TPMV N protein specific antibody titres were measured in the sera of TPMV-N-protein-immunized and experimentally TPMV-infected shrews, whereas no cross-reactivity to other hantavirus N proteins was found. Therefore, the generated mAbs and the established ELISA system represent useful serological tools to detect TPMV, TPMV-related virus antigens or hantavirus-specific antibodies in hantavirus-infected shrews. - Takaaki Koma, Kumiko Yoshimatsu, Midori Taruishi, Daisuke Miyashita, Rika Endo, Kenta Shimizu, Shumpei P. Yasuda, Takako Amada, Takahiro Seto, Ryo Murata, Haruka Yoshida, Hiroaki Kariwa, Ikuo Takashima, Jiro ArikawaJOURNAL OF VIROLOGICAL METHODS 185 1 74 - 81 2012年10月 [査読有り][通常論文]
New World hantaviruses were divided into five groups based on the amino acid sequence variability of the internal variable region (around 230-302 amino acids) of hantavirus nucleocapsid protein (NP). Sin Nombre virus (SNV), Andes virus, Black Creek Canal virus (BCCV). Carrizal virus (CARV) and Cano Delgadito virus belong to groups 1, 2, 3,4 and 5, respectively. Patient and rodent sera were serotyped successfully by an enzyme-linked immunosorbent assay (ELISA) with recombinant truncated NP lacking 99 N-terminal amino acids (trNP100) of SNV, CARV and BCCV. The trNP100 of BCCV showed lower reactivity to heterologous sera. In contrast, whole recombinant NP antigens detected both homologous and heterologous antibodies equally. The results together with results of a previous study suggest that trNP100 can distinguish infections among viruses in groups 1, 2, 3 and 4 of New World hantaviruses. The serotyping ELISA with trNP100 is useful for epidemiological surveillance in humans and rodents. (C) 2012 Elsevier BM. All rights reserved. - Takahiro Sanada, Takahiro Seto, Yuka Ozaki, Ngonda Saasa, Kumiko Yoshimatsu, Jiro Arikawa, Kentaro Yoshii, Hiroaki KariwaJOURNAL OF GENERAL VIROLOGY 93 10 2237 - 2246 2012年10月 [査読有り][通常論文]
Hantaviruses belong to the family Bunyaviridae and are maintained in wild rodents. Although Vero E6 cells, which originate from African green monkey kidney, are used widely in hantavirus research, isolation of hantaviruses from this cell line is difficult. To develop an efficient method of propagation and isolation of hantaviruses we established a novel cell line, MRK101, derived from the kidney of the grey red-backed vole (Myodes rufocanus bedfordiae), the natural host of Hokkaido virus (HOKV). The MRK101 cells showed a significantly higher susceptibility to Puumala virus (PUUV) hosted by Myodes glareolus than Vero E6 cells. Viral nucleocapsid protein in PUUV-infected MRK101 cells was detected earlier than in Vero E6 cells, and the viral titre in the culture fluid of MRK101 cells was higher than that of Vero E6 cells during the early phase of infection. In contrast, MRK101 cells showed no susceptibility to Hantaan virus. HOKV, which has not been isolated to date, was isolated successfully using MRK101 cells. Moreover, the newly isolated HOKV was successfully propagated in MRK101, but not Vero E6, cells. Phylogenic analyses of the S (small), M (medium) and L (large) segment sequences revealed that HOKV is related most closely to PUUV, but is distinct from other hantaviruses. These data suggest that the MRK101 cell line is a useful tool for the isolation and propagation of hantaviruses. Moreover, this is (to our knowledge) the first report of hantavirus isolation in a cell line that originated from the natural host. - Takahiro Sanada, Hiroaki Kariwa, Ngonda Saasa, Keisuke Yoshikawa, Takahiro Seto, Vyacheslav G. Morozov, Evgeniy A. Tkachenko, Leonid I. Ivanov, Kumiko Yoshimatsu, Jiro Arikawa, Kentaro Yoshii, Ikuo TakashimaJOURNAL OF VETERINARY MEDICAL SCIENCE 74 9 1237 - 1242 2012年09月 [査読有り][通常論文]
Antigenic diversity among different hantaviruses requires a variety of reagents for diagnosis of hantavirus infection. To develop a diagnostic method applicable to various hantavirus infections with a single set of reagents, we developed an enzyme-linked immuno-sorbent assay (ELISA) using recombinant nucleocapsid proteins of three hantaviruses, Amur, Hokkaido, and Sin Nombre viruses. This novel cocktail antigen-based ELISA enabled detection of antibodies against Hantaan, Seoul, Amur, Puumala, and Sin Nombre viruses in immunized laboratory animals. In wild rodent species, including Apodemus, Ratios, and Myodes, our ELISA detected antibodies against hantaviruses with high sensitivity and specificity. These data suggest that our novel diagnostic ELISA is a useful tool for screening hantavirus infections and could be effectively utilized for serological surveillance and quarantine purposes. - Vu Dinh Luan, Kumiko Yoshimatsu, Rika Endo, Midori Taruishi, Vo Thi Huong, Dang Tuan Dat, Pham Cong Tien, Kenta Shimzu, Takaaki Koma, Shumpei P. Yasuda, Le Nhi, Vu Thi Que Huong, Jiro ArikawaJOURNAL OF VETERINARY MEDICAL SCIENCE 74 9 1155 - 1162 2012年09月 [査読有り][通常論文]
To investigate the distribution of hantaviruses among animals in Southern and Central Highland area of Vietnam, a total of 1311 serum samples were obtained from rats and Asian house shrews (Suncus murinus) captured at 11 locations between 2006 and 2009. A total of 1066 serum samples from rats were examined for IgG antibodies against Hantaan virus, and there were 30 antibody-positive serum samples from rats that had been captured mainly in a port area and urban area in Ho Chi Minh City (HCMC) (2.8%). All of the antibody-positive rats were Rattus norvegicus, and they had Seoul virus (SEOV) genome in their lungs. SEOV sequences detected from rats captured in Southern Vietnam belonged to the same lineage as those from rats captured at Haiphong Port and a market area in Hanoi City. SEOV strain CSG5 was isolated from a rat captured at Saigon Harbor. Strain CSG5 showed a cross-neutralization pattern almost the same as that of a representative strain of SEOV. A total of 245 Asian house shrews were captured in the Central Highland area and near HCMC. Sera were examined for IgG antibodies against Thottapalayam virus (TPMV), and 32 (13.1%) of the antibody-positive shrews were mainly from the Central Highland area and showed a neutralizing antibody against TPMV. These results indicated that SEOV is distributed among R. norvegicus inhabiting harbor and urban areas of Southern Vietnam and that TPMV or an antigenically related virus is distributed among Asian house shrews in Central Highland area. - Ngonda Saasa, Cornelio Sanchez-Hernandez, Maria de Lourdes Romero-Almaraz, Ezequiel Guerrero-Ibarra, Alberto Almazan-Catalan, Haruka Yoshida, Daisuke Miyashita, Mariko Ishizuka, Takahiro Sanada, Takahiro Seto, Kentaro Yoshii, Celso Ramos, Kumiko Yoshimatsu, Jiro Arikawa, Ikuo Takashima, Hiroaki KariwaVIRUS RESEARCH 168 1-2 88 - 96 2012年09月 [査読有り][通常論文]
In our recent epidemiological survey conducted in Mexico for hantavirus infection, we identified three distinct viruses circulating in Mexican wild rodents, namely Montano virus (MTNV), Huitzilac virus (HUIV), and Carrizal virus (CARV). To gain a detailed understanding of hantavirus epidemiology and its associated hosts, 410 rodents were captured at eight collecting points in Morelos and Guerrero, Mexico, and examined for hantavirus seroprevalence, the presence of viral RNA, and rodent host species identification using cytochrome b gene sequences. Of the 32 species captured, seven species were positive for hantavirus: Peromyscus beatae (31/127; 24.4%), Reithrodontomys sumichrasti (6/15; 40%), Reithrodontomys megalotis (2/25; 8%), Peromyscus aztecus evides (1/1; 100%), Peromyscus megalops (1/41; 2.4%), Megadontomys thomasi (1/9; 11.1%), and Neotoma picta (1/6; 16.7%), with an overall prevalence of 10.5%; virus genome persisted in the majority of seropositive rodents. Nucleotide sequence and phylogenetic analysis showed that the viruses belonged mainly to the three lineages previously identified. The data showed that MTNV and CARV were primarily carried by P. beatae and R. sumichrasti, respectively. In addition, the data revealed an apparent complex interaction between hantaviruses and their hosts, suggesting active transmission and/or spillover infections within sympatric rodent species. (c) 2012 Elsevier B.V. All rights reserved. - ベトナムのラットおよびヒトにおけるラットE型肝炎ウイルスの感染状況の調査清水 健太, 李 天成, 安田 俊平, 吉松 組子, 駒 貴明, 長谷部 太, 山城 哲, Hoa Nguyen Thuy, Mai Le Thi Quynh, 有川 二郎日本獣医学会学術集会講演要旨集 154回 265 - 265 (公社)日本獣医学会 2012年08月
- 新たに分離されたHokkaidoウイルスの性状解析真田 崇弘, 尾崎 由佳, 瀬戸 隆弘, 中尾 桃子, Saasa Ngonda, 吉松 組子, 有川 二郎, 好井 健太朗, 苅和 宏明日本獣医学会学術集会講演要旨集 154回 265 - 265 (公社)日本獣医学会 2012年08月 [査読有り][通常論文]
- Rie Isozumi, Kumiko Yoshimatsu, Tetsu Yamashiro, Futoshi Hasebe, Binh Minh Nguyen, Tuan Cuong Ngo, Shumpei P. Yasuda, Takaaki Koma, Kenta Shimizu, Jiro ArikawaEMERGING INFECTIOUS DISEASES 18 8 1383 - 1385 2012年08月 [査読有り][通常論文]
- Ngonda Saasa, Haruka Yoshida, Kenta Shimizu, Cornelio Sanchez-Hernandez, Maria de Lourdes Romero-Almaraz, Takaaki Koma, Takahiro Sanada, Takahiro Seto, Kentaro Yoshii, Celso Ramos, Kumiko Yoshimatsu, Jiro Arikawa, Ikuo Takashima, Hiroaki KariwaVIROLOGY 428 1 48 - 57 2012年06月 [査読有り][通常論文]
The hantavirus nucleocapsid (N) protein is an important immunogen that stimulates a strong and cross-reactive immune response in humans and rodents. A large proportion of the response to N protein has been found to target its N-terminus. However, the exact nature of this bias towards the N-terminus is not yet fully understood. We characterized six monoclonal antibodies (mAbs) against the N protein of Montano virus (MTNV), a Mexican hantavirus. Five of these mAbs recognized eight American hantaviruses and six European and Asian hantaviruses, but not the Soricomorpha-borne Thottapalayam hantavirus. The N protein-reactive binding regions of the five mAbs were mapped to discontinuous epitopes within the N-terminal 13-51 amino acid residues, while a single serotype-specific mAb was mapped to residues 1-25 and 49-75. Our findings suggest that discontinuous epitopes at the N-terminus are conserved, at least in rodent-borne hantaviruses, and that they contribute considerably to N protein cross-reactivity. (C) 2012 Elsevier Inc. All rights reserved. - 腎症候性出血熱の動物モデルの開発清水 健太, 吉松 組子, 駒 貴明, 伊勢川 裕二, 安田 俊平, 天田 貴子, 五十棲 理恵, 有川 二郎日本獣医学会学術集会講演要旨集 153回 247 - 247 (公社)日本獣医学会 2012年03月
- イムノクロマト法による抗ハンタウイルス抗体の迅速抗体検出法の開発天田 貴子, 吉松 組子, 安田 俊平, 駒 貴明, 清水 健太, 五十棲 理恵, 林元 展人, 高倉 彰, 有川 二郎日本獣医学会学術集会講演要旨集 153回 273 - 273 (公社)日本獣医学会 2012年03月
- Hiroaki Kariwa, Keisuke Yoshikawa, Yoichi Tanikawa, Takahiro Seto, Takahiro Sanada, Ngonda Saasa, Leonid I. Ivanov, Raisa Slonova, Tatyana A. Zakharycheva, Ichiro Nakamura, Kumiko Yoshimatsu, Jiro Arikawa, Kentaro Yoshii, Ikuo TakashimaAMERICAN JOURNAL OF TROPICAL MEDICINE AND HYGIENE 86 3 545 - 553 2012年03月 [査読有り][通常論文]
Hemorrhagic fever with renal syndrome (HFRS) is a serious public health issue in Far East Russia. Two different hantaviruses were isolated from rodents captured in the Khabarovsk region: Amur virus (AMRV; Khekhtsir/AP209/2005 strain from Apodemus peninsulae) and Hantaan virus (HTNV; Galkino/AA57/2002 strain from A. agrarius). Genetic analysis of the new isolates revealed that the M and L segments were apparently different between AMRV and HTNV, but S segments of the two viruses were closer. The antigenicities of AMRV, HTNV, and Seoul virus (SEOV) were differentiated by cross-neutralization. Serological differential diagnoses of 67 HFRS patients in the Prymorsky and Khabarovsk regions of Far East Russia were conducted using a neutralization test. The results revealed that the major cause of HFRS varied with location in Far East Russia: SEOV for Vladivostok city in the Prymorsky region, AMRV in rural areas of the Primorsky region, and probably HTNV for the Khabarovsk region. - Shumpei P. Yasuda, Kumiko Yoshimatsu, Takaaki Koma, Kenta Shimizu, Rika Endo, Rie Isozumi, Jiro ArikawaJOURNAL OF VETERINARY MEDICAL SCIENCE 74 2 215 - 219 2012年02月 [査読有り][通常論文]
Truncated recombinant nucleocapsid proteins (trNs) that lack N-terminally located cross-reactive epitopes of four Murinae rodent-associated hantaviruses, Seoul virus (SEOV). Thailand virus, Hantaan virus (HTNV) and Dobrava-Belgrade virus, were produced by using a baculovirus expression system. ELISA with the trNs as antigens enabled serotyping of immune sera from rats experimentally inoculated with the corresponding hantaviruses with cut-off OD values of 60% of those of whole N of HTNV. The trN-based ELISA could serotype 12 out of 13 sera obtained from wild rodents (Ratios norvegicus) naturally infected with SEOV using the 60% cut-off value. These results indicate that screening with whole N followed by serotyping with trNs using a cut-off OD value of 60% of that of whole N is a useful method for serological surveillance of Murinae-associated hantavirus infection among rodents. - Hiroaki Kariwa, Haruka Yoshida, Cornelio Sanchez-Hernandez, Maria de Lourdes Romero-Almaraz, Jose Alberto Almazan-Catalan, Celso Ramos, Daisuke Miyashita, Takahiro Seto, Ayako Takano, Masashi Totani, Ryo Murata, Ngonda Saasa, Mariko Ishizuka, Takahiro Sanada, Kentaro Yoshii, Kumiko Yoshimatsu, Jiro Arikawa, Ikuo TakashimaVIRUS RESEARCH 163 2 486 - 494 2012年02月 [査読有り][通常論文]
A variety of hantaviruses are harbored by rodents in North and South America, some of which can cause hantavirus pulmonary syndrome. To obtain greater evolutionary insight into hantaviruses in the Americas, a total of 211 rodents were captured in the Mexican states of Guerrero and Morelos in 2006. Anti-hantavirus antibodies were detected in 27 of 211 serum samples (12.8%) by ELISA. The distribution of seropositive rodents was: 17 Peromyscus beatae, 1 Megadontomys thomasi,1 Neotoma picta, 6 Reithrodontomys sumichrasti, and 2 Reithrodontomys megalotis. The hantavirus small (S), medium (M), and large (L) genome segments from P. beatae, R. sumichrasti, and R. megalotis were amplified and the sequences covering the open reading frames were determined. The hantaviruses from P. beatae, R. sumichrasti, and R. megalotis were provisionally designated Montano (MTN), Carrizal (CAR), and Huitzilac (HUI), respectively. The M segment amino acid identities among the Mexican hantaviruses were 80.8-93.0%. When these M segments were compared to those of known hantaviruses, MTN virus was most closely related to Limestone Canyon (LSC) virus (88.9% amino acid identity), while the CAR and HUI viruses were most closely related to El Moro Canyon (ELMC) virus (90-91% identity). Phylogenetic analysis revealed that the MTN, CAR, and HUI viruses occupy a monophyletic clade with the LSC, ELMC, and Rio Segundo viruses, which are harbored by Peromyscus boylii, R. megalotis, and Reithrodontomys mexicanus, respectively. The data obtained in this study provide important information for understanding the evolution of hantaviruses in the Americas. (C) 2011 Elsevier B.V. All rights reserved. - Al-shere T. Amilasan, Mugen Ujiie, Motoi Suzuki, Eumelia Salva, Maria Cecilia P. Belo, Nobuo Koizumi, Kumiko Yoshimatsu, Wolf-Peter Schmidt, Shane Marte, Efren M. Dimaano, Jose Benito Villarama, Koya AriyoshiEMERGING INFECTIOUS DISEASES 18 1 91 - 94 2012年01月 [査読有り][通常論文]
After a typhoon in September 2009, an outbreak of leptospirosis occurred in Metro Manila, the Philippines; 471 patients were hospitalized and 51(10.8%) died. A hospital-based investigation found risk factors associated with fatal infection to be older age, hemoptysis, anuria, jaundice, and delayed treatment with antimicrobial drugs. - Tian-Cheng Li, Kumiko Yoshimatsu, Shumpei P. Yasuda, Jiro Arikawa, Takaaki Koma, Michiyo Kataoka, Yasushi Ami, Yuriko Suzaki, Le Thi Quynh Mai, Nguyen Thuy Hoa, Tetsu Yamashiro, Futoshi Hasebe, Naokazu Takeda, Takaji WakitaJOURNAL OF GENERAL VIROLOGY 92 Pt 12 2830 - 2837 2011年12月 [査読有り][通常論文]
Hepatitis E virus (HEV) is a causative agent of hepatitis E. Recently, a novel hepatitis E-like virus was isolated from Norway rats in Germany. However, the antigenicity, pathogenicity and epidemiology of this virus are unclear because of the lack of a cell-culture system in which to grow it. In this study, an N-terminally truncated ORF2 protein was expressed in insect Tn5 cells using a recombinant baculovirus expression system and a large amount of 53 kDa protein was expressed and efficiently released into the supernatant. Electron microscopic analyses of the purified 53 kDa protein revealed that the protein self-assembled into two types of empty HEV-like particles (rat HEVLPs). The smaller rat HEVLPs were estimated to be 24 nm in diameter, which is similar to the size of genotype G1, G3 and G4 HEVLPs. The larger rat HEVLPs were estimated to measure 35 nm in diameter, which is similar to the size of native rat HEV particles. An ELISA to detect antibodies was established using rat HEVLPs as the antigens, which demonstrated that rat HEVLPs were cross-reactive with G1, G3 and G4 HEVs. Detection of IgG and IgM antibodies was performed by examination of 139 serum samples from wild rats trapped in Vietnam, and it was found that 20.9 % (29/139) and 3.6 % (5/139) of the samples were positive for IgG and IgM, respectively. In addition, rat HEV RNA was detected in one rat serum sample that was positive for IgM. These results indicated that rat HEV is widespread and is transmitted among wild rats. - Effect of environmental enrichment after the occurrence of wet bedding created by mice and abnormal fur in mice.Tosa N, Yoshimatsu K, Arikawa JJ Am Assoc Lab Anim Sci 50 5 779 - 780 2011年09月 [査読有り][通常論文]
- Takahiro Sanada, Hiroaki Kariwa, Noriyo Nagata, Yoichi Tanikawa, Takahiro Seto, Kumiko Yoshimatsu, Jiro Arikawa, Kentaro Yoshii, Ikuo TakashimaVIRUS RESEARCH 160 1-2 108 - 119 2011年09月 [査読有り][通常論文]
The mechanism of hantavirus persistent infection in natural hosts is poorly understood due to a lack of laboratory animal models. Herein, we report that Syrian hamsters (Mesocricetus auratus) infected with Puumala virus (PUUV) at 4 weeks old show persistent infection without clinical symptoms for more than 2 months. IgG and IgM antibodies against the viral nucleocapsid protein and neutralizing antibody were first detectable at 14 days postinoculation (dpi) and maintained through 70 dpi. Viral RNA was first detected from 3 dpi in lungs and blood clots, and was detected in all tissues tested at 7 dpi. The viral RNA persisted for at least 70 days in the lungs, kidney, spleen, heart, and brain. The highest level of RNA copies was observed at 14 dpi in the lungs. Slight inflammatory reactions were observed in the lungs, adrenal glands, and brain. Immunohistochemical analysis revealed that PUUV antigen persisted until 56 dpi in the kidneys and adrenal glands. Infected hamsters showed no body weight loss or clinical signs. These results indicate that PUUV infection in hamsters is quite similar to the hantavirus infection of natural host rodents. (C) 2011 Elsevier B.V. All rights reserved. - ハンタウイルス糖蛋白質の細胞内輸送に関与するウイルス蛋白質とその機能領域清水 健太, 吉松 組子, 駒 貴明, 安田 俊平, 有川 二郎日本獣医学会学術集会講演要旨集 152回 236 - 236 (公社)日本獣医学会 2011年08月
- 日本のげっ歯類のハンタウイルス感染状況に関する血清疫学調査とエゾヤチネズミが保有するHokkaidoウイルスの遺伝的多様性尾崎 由佳, 萩谷 友洋, 真田 崇弘, 瀬戸 隆弘, Kyle Taylor, 吉川 佳佑, Leonid Ivanov, 好井 健太朗, 坪田 敏男, 池中 良徳, 石塚 真由美, 吉松 組子, 有川 二郎, 苅和 宏明日本獣医学会学術集会講演要旨集 152回 256 - 256 (公社)日本獣医学会 2011年08月 [査読有り][通常論文]
- 尾崎 由佳, 萩谷 友洋, 真田 崇弘, 瀬戸 隆弘, Taylor Kyle, 吉川 佳佑, Ivanov Leonid I., 好井 健太朗, 坪田 敏男, 池中 良徳, 石塚 真由美, 吉松 組子, 有川 二郎, 苅和 宏明北海道獣医師会雑誌 55 8 415 - 415 (公社)北海道獣医師会 2011年08月 [査読無し][通常論文]
- Valine 1532 of human BRC repeat 4 plays an important role in the interaction between BRCA2 and RAD51Kazuhiko Ochiai, Yasunaga Yoshikawa, Kumiko Yoshimatsu, Toshina Oonuma, Yukiko Tomioka, Eichi Takeda, Jiro Arikawa, Katsumi Mominoki, Toshinori Omi, Kazuyoshi Hashizume, Masami MorimatsuFEBS LETTERS 585 12 1771 - 1777 2011年06月 [査読有り][通常論文]
The breast cancer susceptibility protein BRCA2 is essential for recombinational DNA repair. BRCA2 specifically binds to RAD51 via eight BRC repeat motifs and delivers RAD51 to double-stranded DNA breaks. In this study, a mammalian two-hybrid assay and competitive ELISA showed that the interaction between BRC repeat 4 (BRC4) and RAD51 was strengthened by the substitution of a single BRC4 amino acid from valine to isoleucine (V1532I). However, the cancer-associated V1532F mutant exhibited very weak interaction with RAD51. This study used a comparative analysis of BRC4 between animal species to identify V1532 as an important residue that interacts with RAD51. Structured summary of protein interactions: cRAD51 physically interacts with cRAD51 by two hybrid (View interaction) fBRC4 physically interacts with cRAD51 by two hybrid (View interaction) cBRC4 physically interacts with cRAD51 by two hybrid (View interaction) hBRC4 physically interacts with hBRC4 and hRAD51 by competition binding (View Interaction 1, 2) hBRC4 physically interacts with cRAD51 by two hybrid (View interaction) hBRC4 binds to hRAD51 by enzyme linked immunosorbent assay (View interaction) (C) 2011 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved. - Takahiro Seto, Evgeniy A. Tkachenko, Vyacheslav G. Morozov, Yoichi Tanikawa, Sergey I. Kolominov, Sergey N. Belov, Ichiro Nakamura, Nobuo Hashimoto, Yasuhiro Kon, Alexander E. Balakiev, Tamara K. Dzagurnova, Olga A. Medvedkina, Mina Nakauchi, Mariko Ishizuka, Kentaro Yoshii, Kumiko Yoshimatsu, Leonid V. Ivanov, Jiro Arikawa, Ikuo Takashima, Hiroaki KariwaJOURNAL OF VIROLOGICAL METHODS 173 1 17 - 23 2011年04月 [査読有り][通常論文]
Puumala virus (PUUV) and other Arvicolinae-borne hantaviruses are difficult to cultivate in cell culture. To isolate these hantaviruses efficiently, hantavirus nucleocapsid protein (NP)-positive but seronegative wild rodents were selected by NP-detection ELISA. Three of 68 Myodes glareolus captured in Samara, Russia, were NP-positive and seronegative. Syrian hamsters were inoculated with lung homogenates from NP-positive rodents for virus propagation. Virus isolation in vitro was carried out by inoculation of lung homogenates of NP-positive hamsters to Vero E6 cell monolayers. Two PUUV strains (Samara49/CG/2005 and Samara94/CG/2005) from M. glareolus were isolated in Vero E6 cells. Nucleotide and amino acid sequence identities of the S segment of these isolates to those of PUUV F-s808 from a fatal HFRS patient in Samara region were 96.7-99.3% and 99.3-100.0%, respectively. Morphologic features of Vero E6 cells infected with PUUV strain Samara49/CG/2005 were quite similar to those of Hantaan virus-infected cells. Isolation of Hokkaido virus from Myodes rufocanus captured in Hokkaido, Japan, was also performed. Hokkaido virus NP and RNA were recovered and maintained in hamsters. These results suggest that inoculation of Syrian hamsters with rodent samples is an efficient method for the isolation and maintenance of PUUV and other Arvicolinae-borne hantaviruses. (C) 2011 Elsevier B.V. All rights reserved. - Chandika D. Gamage, Shumpei P. Yasuda, Sanae Nishio, Senanayake A. Kularatne, Kosala Weerakoon, Jayanthe Rajapakse, Chinyere Nwafor-Okoli, Romeo B. Lee, Yoshi Obayashi, Kumiko Yoshimatsu, Jiro Arikawa, Hiko TamashiroJAPANESE JOURNAL OF INFECTIOUS DISEASES 64 1 72 - 75 2011年01月 [査読無し][通常論文]
A cross-sectional study was undertaken to determine the current prevalence of leptospirosis and hantaviral infections, and the socio-demographic characteristics and risk factors of infected patients, in Kandy, Sri Lanka. This report discusses the serological evidence of hantavirus infections among 105 suspected leptospirosis patients, 8 of whom had hantavirus antibodies. Serotyping ELISA showed that these 8 patients had high optical density values for Thailand virus. Most of the sera showed that the focus reduction neutralization test titer against Thailand virus was higher than that against Seoul virus, thereby suggesting that the hantaviral antibodies found in Sri Lanka are different from Seoul virus but closely related to Thailand virus. These findings imply that the hantaviral infection found in Kandy, Sri Lanka appears to be due to a virus similar to Thailand virus. Epidemiological analysis revealed that the association between hantavirus infection and socio-demographic characteristics was not statistically significant. - Chandika D. Gamage, Shumpei P. Yasuda, Sanae Nishio, Senanayake A. Kularatne, Kosala Weerakoon, Jayanthe Rajapakse, Chinyere Nwafor-Okoli, Romeo B. Lee, Yoshi Obayashi, Kumiko Yoshimatsu, Jiro Arikawa, Hiko TamashiroJAPANESE JOURNAL OF INFECTIOUS DISEASES 64 1 72 - 75 2011年01月 [査読有り][通常論文]
A cross-sectional study was undertaken to determine the current prevalence of leptospirosis and hantaviral infections, and the socio-demographic characteristics and risk factors of infected patients, in Kandy, Sri Lanka. This report discusses the serological evidence of hantavirus infections among 105 suspected leptospirosis patients, 8 of whom had hantavirus antibodies. Serotyping ELISA showed that these 8 patients had high optical density values for Thailand virus. Most of the sera showed that the focus reduction neutralization test titer against Thailand virus was higher than that against Seoul virus, thereby suggesting that the hantaviral antibodies found in Sri Lanka are different from Seoul virus but closely related to Thailand virus. These findings imply that the hantaviral infection found in Kandy, Sri Lanka appears to be due to a virus similar to Thailand virus. Epidemiological analysis revealed that the association between hantavirus infection and socio-demographic characteristics was not statistically significant. - Erdenesaikhan Tegshduuren, Kumiko Yoshimatsu, Midori Taruishi, Rika Endo, Kenta Shimizu, Takaaki Koma, Shumpei P. Yasuda, Hiroaki Kariwa, Jiro Arikawa, Chiaki IshiharaCOMPARATIVE IMMUNOLOGY MICROBIOLOGY AND INFECTIOUS DISEASES 33 6 E67 - E73 2010年12月 [査読有り][通常論文]
Tula virus (TULV) and Puumala virus (PUUV) are hantaviruses carried by the bank vole (Myodes glareolus) and European common vole (Microtus arvalis), respectively. PUUV is a causative agent of hemorrhagic fever with renal syndrome (HFRS), while TULV is thought to be apathogenic to humans. The N-terminal regions of the N proteins from TULV and PUUV were expressed and applied as enzyme-linked immunosorbent assay (ELISA) antigens. Colonized Japanese grass voles (Microtus montebelli) and BALB/c mice were used for experimental inoculation of the vole-borne hantaviruses TULV and PUUV. Voles and mice showed significant antibody production toward both viruses, but these antisera showed little cross-reactivity between TULV and PUUV in the immunofluorescence antibody assay and ELISA. In contrast, sera from patients with HFRS caused by PUUV exhibited high cross-reactivity against the TULV antigen, and sera from a natural rodent reservoir showed moderate cross-reactivity against the heterologous antigen, indicating that the antigenic cross-reactivity between TULV and PUUV differs in sera from rodents and humans. (C) 2010 Elsevier Ltd. All rights reserved. - ハンタウイルスNucleocapsid proteinはGlycoprotein Gcのシスゴルジへの局在を促進する清水 健太, 吉松 組子, 駒 貴明, 遠藤 理香, 安田 俊平, Erdenesaikhan Tegshduuren, 有川 二郎日本獣医学会学術集会講演要旨集 150回 232 - 232 (公社)日本獣医学会 2010年09月
- メキシコ由来のハンタウイルスに対するモノクローナル抗体の作出と各種ハンタウイルスに対する反応性の検討吉田 喜香, 苅和 宏明, 真田 崇弘, Saasa Ngonda, 瀬戸 隆弘, 吉松 組子, 有川 二郎, 好井 健太朗, 高島 郁夫日本獣医学会学術集会講演要旨集 150回 249 - 249 (公社)日本獣医学会 2010年09月 [査読有り][通常論文]
- Puumalaウイルスを感染させたシリアンハムスター(Mesocricetus auratus)の感染動態の解析真田 崇弘, 苅和 宏明, 谷川 洋一, Nur Hardy Abu Daud, 瀬戸 隆弘, 永田 典代, 吉松 組子, 有川 二郎, 好井 健太朗, 高島 郁夫日本獣医学会学術集会講演要旨集 150回 252 - 252 (公社)日本獣医学会 2010年09月 [査読有り][通常論文]
- Takaaki Koma, Kumiko Yoshimatsu, Noemi Pini, David Safronetz, Midori Taruishi, Silvana Levis, Rika Endo, Kenta Shimizu, Shumpei P. Yasuda, Hideki Ebihara, Heinz Feldmann, Delia Enria, Jiro ArikawaJOURNAL OF CLINICAL MICROBIOLOGY 48 5 1635 - 1642 2010年05月 [査読有り][通常論文]
Sin Nombre virus (SNV), Andes virus (ANDV), and Laguna Negra virus (LANV) have been known as the dominant causative agents of hantavirus pulmonary syndrome (HPS). ANDV and LANV, with different patterns of pathogenicity, exist in a sympatric relationship. Moreover, there is documented evidence of person-to-person transmission of ANDV. Therefore, it is important in clinical medicine and epidemiology to know the serotype of a hantavirus causing infection. Truncated SNV, ANDV, and LANV recombinant nucleocapsid proteins (trNs) missing 99 N-terminal amino acids (trN100) were expressed using a baculovirus system, and their applicability for serotyping SNV, ANDV, and LANV infection by the use of enzyme-linked immunosorbent assays (ELISA) was examined. HPS patient sera and natural-reservoir rodent sera infected with SNV, ANDV, and LANV showed the highest optical density (OD) values for homologous trN100 antigens. Since even patient sera with lower IgM and IgG antibody titers were serotyped, the trN100s are therefore considered useful for serotyping with early-acute-phase sera. In contrast, assays testing whole recombinant nucleocapsid protein antigens of SNV, ANDV, and LANV expressed in Escherichia coli detected homologous and heterologous antibodies equally. These results indicated that a screening ELISA using an E. coli-expressed antigen followed by a serotyping ELISA using trN100s is useful for epidemiological surveillance in regions where two or more hantavirus species cocirculate. - Vu Thi Que Huong, Kumiko Yoshimatsu, Vu Dinh Luan, Le Van Tuan, Le Nhi, Jiro Arikawa, Tran Minh Nhu NguyenEMERGING INFECTIOUS DISEASES 16 2 363 - 365 2010年02月 [査読有り][通常論文]
- Jonas Schmidt-Chanasit, Sandra Essbauer, Rasa Petraityte, Kumiko Yoshimatsu, Kirsten Tackmann, Franz J. Conraths, Kestutis Sasnauskas, Jiro Arikawa, Astrid Thomas, Martin Pfeffer, Jerrold J. Scharninghausen, Wolf Splettstoesser, Matthias Wenk, Gerald Heckel, Rainer G. UlrichJOURNAL OF VIROLOGY 84 1 459 - 474 2010年01月 [査読有り][通常論文]
To examine the host association of Tula virus (TULV), a hantavirus present in large parts of Europe, we investigated a total of 791 rodents representing 469 Microtus arvalis and 322 Microtus agrestis animals from northeast, northwest, and southeast Germany, including geographical regions with sympatric occurrence of both vole species, for the presence of TULV infections. Based on serological investigation, reverse transcriptase PCR, and subsequent sequence analysis of partial small (S) and medium (M) segments, we herein show that TULV is carried not only by its commonly known host M. arvalis but also frequently by M. agrestis in different regions of Germany for a prolonged time period. At one trapping site, TULV was exclusively detected in M. agrestis, suggesting an isolated transmission cycle in this rodent reservoir separate from spillover infections of TULV-carrying M. arvalis. Phylogenetic analysis of the S and M segment sequences demonstrated geographical clustering of the TULV sequences irrespective of the host, M. arvalis or M. agrestis. The novel TULV lineages from northeast, northwest, and southeast Germany described here are clearly separated from each other and from other German, European, or Asian lineages, suggesting their stable geographical localization and fast sequence evolution. In conclusion, these results demonstrate that TULV represents a promiscuous hantavirus with a large panel of susceptible hosts. In addition, this may suggest an alternative evolution mode, other than a strict coevolution, for this virus in its Microtus hosts, which should be proven in further large-scale investigations on sympatric Microtus hosts. - Hiroaki Kariwa, Evgeniy A. Tkachenko, Vyacheslav G. Morozov, Takahiro Seto, Yoichi Tanikawa, Sergey I. Kolominov, Sergey N. Belov, Ichiro Nakamura, Nobuo Hashimoto, Alexander E. Balakiev, Tamara K. Dzagurnova, Nur Hardy bin Abu Daud, Daisuke Miyashita, Olga A. Medvedkina, Mina Nakauchi, Mariko Ishizuka, Kentaro Yoshii, Kumiko Yoshimatsu, Jiro Arikawa, Ikuo TakashimaJOURNAL OF VETERINARY MEDICAL SCIENCE 71 12 1569 - 1578 2009年12月 [査読有り][通常論文]
European Russia is a highly endemic area of hemorrhagic fever with renal syndrome (HFRS), a rodent-borne zoonotic disease, caused by hantaviruses. In total, 145 small mammals of four species (Myodes glareolus, Apodemus flavicollis, A. agrarius, and A. uralensis) were trapped in the Samara region of European Russia in August 2005 and examined for the presence of hantavirus (HV). Anti-HV antibodies were found in six of 68 (8.8%) M. glareolus and in one of 19 (5.3%) A. flavicollis by indirect immunofluorescent antibody assay (IFA). The Puumala virus (PUUV), which is one of the hantavirus species, was detected in the lungs of seven M. glareolus by RT-PCR. The virus S-segment was extremely similar (96.2% to 99.3%) to the sequence found in a fatal case of HFRS in the Samara region. Phylogenetic analyses of S and M segments showed that the Samara PUUVs form a cluster within the Russian Volga lineage and apparently differ from other European PUUVs. Anti-PUUV antibodies were found in blood sera from seven HFRS patients and from one undiagnosed patient from the Samara region, using IFA and an enzyme-linked immunosorbent assay (ELISA). These data Suggest that the bank vole M. glareolus is a primary natural reservoir and vector for PUUV, which is the main causative agent of HFRS in humans in the Samara region. - Thua Thang Truong, Kumiko Yoshimatsu, Koichi Araki, Byoung-Hee Lee, Ichiro Nakamura, Rika Endo, Kenta Shimizu, Shumpei P. Yasuda, Takaaki Koma, Midori Taruishi, Megumi Okumura, Uyen Ninh Truong, Jiro ArikawaJOURNAL OF VETERINARY MEDICAL SCIENCE 71 10 1357 - 1363 2009年10月 [査読有り][通常論文]
The distribution of anti-hantavirus antibodies in humans and rodents in northern Vietnam was examined. In total, 837 seruml samples from healthy humans (617) and patients with fever (220), living in six different areas were screened for IgG antibodies against Hantaan or Seoul virus (SEOV) by ELISA, IFA, and Western blot analysis. Antibody-positive sera were identified ill 7/617 (1.1%) healthy donors, 51150 port workers in the port of Hai Phong, and 2/185 residents of Ha Nam Province. In comparison, positive sera were detected in 5/220 (2.3%) fever patients ill the provinces of Ha Nam (1/58) and Thanh Hoa (4/146). Antibody-positive Rattus norvegicus were found in the provinces of Ha Nam (7/52) and Thanh Hoa (1/67), in Haibatrung District (7/43) in Hanoi, and in Hai Phong" Port (21/62), while antibody-positive R. rattus (2/17) were found in Hai Phong Port. Part of the Gc region from the viral genome was amplified by RT-PCR using lung tissue samples front R. norvegicus in Haibatrung (2/7) and Hai Phong Port (7/9), bill not from R. rattus (0/2). Viral sequences were located in the SEOV clade and formed a single lineage with Indonesian SEOV. suggesting that Vietnamese SEOV is part of a distinct lineage among Asian SEOVs. - S. Chandy, M. Okumura, K. Yoshimatsu, R. G. Ulrich, G. T. John, P. Abraham, J. Arikawa, G. SridharanINDIAN JOURNAL OF MEDICAL MICROBIOLOGY 27 4 348 - 350 2009年10月 [査読無し][通常論文]
Hantaviruses cause hemorrhagic fever with renal syndrome in Europe and Asia. There are about 20 documented hantavirus species and newer species are being described worldwide, especially in non-rodent reservoirs, i.e shrews. Focus reduction neutralization test is the classical serotyping technique for hantavirus. However, this study employs a previously established serotyping ELISA, to retrospectively analyze known hantavirus IgG reactive samples for infecting serotypes. The result suggests presence of Thailand virus- like and Hantaan virus -like strains in India. - メキシコの野生げっ歯類が保有するハンタウイルスの遺伝子解析吉田 喜香, 苅和 宏明, 高野 絢子, 戸谷 理詩, 宮下 大輔, Ngonda Saasa, 瀬戸 隆弘, 真田 崇弘, 吉川 佳佑, 好井 健太朗, 吉松 組子, 有川 次郎, 高島 郁夫日本獣医学会学術集会講演要旨集 148回 237 - 237 (公社)日本獣医学会 2009年09月 [査読有り][通常論文]
- 極東ロシアの野鼠からのハンタウイルスの分離と人におけるウイルス感染状況の調査吉川 佳佑, 苅和 宏明, 瀬戸 隆弘, 真田 崇弘, 石塚 万里子, 吉松 組子, 有川 二郎, Ivanov Leonid, Slonova Raisa, 好井 健太朗, 高島 郁夫北海道獣医師会雑誌 53 8 112 - 112 (公社)北海道獣医師会 2009年08月 [査読有り][通常論文]
- Marc Mertens, Roman Woelfel, Katrin Ullrich, Kumiko Yoshimatsu, Jana Blumhardt, Ina Roemer, Jutta Esser, Jonas Schmidt-Chanasit, Martin H. Groschup, Gerhard Dobler, Sandra S. Essbauer, Rainer G. UlrichMEDICAL MICROBIOLOGY AND IMMUNOLOGY 198 2 83 - 91 2009年05月 [査読有り][通常論文]
Puumala virus (PUUV) is the cause of the majority of haemorrhagic fever with renal syndrome cases in Germany. In 2004, a nephropathia epidemica outbreak was recorded in Lower Bavaria, South-East Germany. For a seroepidemiological study in this region including the resident population at four locations (n = 178) and soldiers from one location (n = 208) indirect immunoglobulin M (IgM) and immunoglobulin G (IgG) enzyme-linked immunosorbent assays (ELISAs) and immunoblot tests based on a yeast-expressed PUUV nucleocapsid protein were established. The validation using human serum panels originating from Germany revealed a diagnostic sensitivity and specificity of 98/100% for the IgM ELISA, 99/99% for the IgG ELISA, 99/100% for the IgM immunoblot test and 100/96% for the IgG immunoblot test. Using the novel IgG assays as well as a commercial IgG ELISA and an immunofluorescence assay for the resident population an average prevalence of 6.7% (12 of 178) with a range of 0% (0 of 21) to 11.9% (7 of 59) was observed. Positive serological results were equally distributed between males and females with an average age of 63 for males and 52 for females. The seroprevalence in the soldier group was found to be about 1% with one positive male of 203 (age 46 years) and one positive female of five (age 47 years). In conclusion, the PUUV seroprevalence in the residents of the outbreak region in Lower Bavaria was found to be up to fivefold higher than the average hantavirus seroprevalence of the German population. - Sara Chandy, Kumiko Yoshimatsu, Hari Kishan Boorugu, Anugrah Chrispal, Kurien Thomas, Abraham Peedicayil, Priya Abraham, Jiro Arikawa, Gopalan SridharanTRANSACTIONS OF THE ROYAL SOCIETY OF TROPICAL MEDICINE AND HYGIENE 103 4 407 - 412 2009年04月 [査読有り][通常論文]
Study of hantavirus infections in India is in its early stages. As early symptoms of hantavirus disease can be non-specific and the diagnosis confirmed only by laboratory testing, use of appropriate diagnostic toots is important. To improve the diagnosis of hantavirus infections in India, commercial ELISA systems followed by indirect immunofluorescence assays were used to detect anti-hantavirus IgM and IgG in samples from patients with acute febrile illness. Of 347 patients tested, 5.2% showed serological evidence of hantavirus infection. Sequences obtained from patients showing molecular evidence of hantavirus infection were related to Hantaan virus. In the absence of mu-capture ELISA, we recommend the use of combination testing systems in areas non-endemic for hantavirus infections. In India there is an increased risk of rodent-borne infections and the differential diagnosis of undifferentiated febrile illness should include hantavirus infection. (C) 2009 Royal Society of Tropical Medicine and Hygiene. Published by Elsevier Ltd. All rights reserved. - 極東ロシアのげっ歯類からのハンタウイルスの分離とウイルス遺伝子の性状解析吉川 佳佑, 苅和 宏明, 瀬戸 隆弘, 真田 崇弘, 石塚 万里子, 吉松 組子, 有川 二郎, Ivanov Lenid, 好井 健太朗, 高島 郁夫日本獣医学会学術集会講演要旨集 147回 260 - 260 (公社)日本獣医学会 2009年03月 [査読有り][通常論文]
- Nur Hardy Abu Daud, Hiroaki Kariwa, Evgeniy Tkachenko, Tamara Dzagurnova, Olga Medvedkina, Petr Tkachenko, Mariko Ishizuka, Takahiro Seto, Daisuke Miyashita, Takahiro Sanada, Mina Nakauchi, Kentaro Yoshii, Akihiko Maeda, Kumiko Yoshimatsu, Jiro Arikawa, Ikuo TakashimaJAPANESE JOURNAL OF VETERINARY RESEARCH 56 3 151 - 165 2008年11月 [査読有り][通常論文]
Puumala virus (PUUV), a causative agent of hemorrhagic fever with renal syndrome (HFRS), is prevalent in Europe and European Russia. No vaccine has been developed for PUUV-associated HFRS, primarily because of the low viral yield in cultured cells. A PUUV strain known as DTK/Ufa-97 was isolated in Russia and adapted for growth in Vero E6 cells maintained in serum-free medium. The DTK/Ufa-97 strain produced a higher viral titer in serum-free medium, suggesting that it may prove useful in the development of an HFRS vaccine. When PUUV-infected Vero E6 cells were grown in serum-free medium, the DTK/Ufa-97 strain yielded more copies of intracellular viral RNA and a higher viral titer in the culture fluid than did the Sotkamo strain. Phylogenetic analysis revealed that PUUVs can be classified into multiple lineages according to geographical origin, and that the DTK/Ufa-97 strain is a member of the Bashkiria-Saratov lineage. The deduced amino acid sequences of the small, medium, and large segments of the DTK/Ufa-97 strain were 99.2% to 100%, 99.3% to 99.8%, and 99.8% identical, respectively, to those of the Bashkirian PUUV strains and 96.9%, 92.6%, and 97.4% identical, respectively, to those of the Sotkamo strain, indicating that the PUUVs are genetically diverse. However, DTK/Ufa-97 and other strains of PUUV exhibited similar patterns of binding to a panel of monoclonal antibodies against Hantaan virus. In addition, diluted antisera (i.e., ranging from 1:160 to 1:640) specific to three strains of PUUV neutralized both homologous and heterologous viruses. These results suggest that the DTK/Ufa-97 strain is capable of extensive growth and is antigenically similar to genetically distant strains of PUUV. - 多種類のハンタウイルス血清型の検出が可能な抗原検出法の開発真田 崇弘, 苅和 宏明, 瀬戸 隆弘, 谷川 洋一, 宮下 大輔, 吉松 組子, 有川 二郎, 好井 健太朗, 高島 郁夫日本獣医学会学術集会講演要旨集 146回 225 - 225 (公社)日本獣医学会 2008年09月 [査読有り][通常論文]
- Midori Taruishi, Kumiko Yoshimatsu, Rei Hatsuse, Megumi Okumura, Ichiro Nakamura, Jiro ArikawaARCHIVES OF VIROLOGY 153 8 1605 - 1609 2008年08月 [査読有り][通常論文]
It is unclear how the hantaviruses are transferred from infected to uninfected rodents. We studied the status of persistently infected laboratory mice and examined the frequency of viral transmission to their offspring. Expression of Hantaan virus nucleocapsid protein was detected in the lungs of persistently infected dams. None of the progeny displayed viral antigen, although they were strongly positive for IgG antibodies against hantavirus. There was neither hantavirus RNA nor virus-specific IgM antibodies or virus-specific CD8(+) T cells in the progeny. These results did not show any indication for a vertical transmission of hantaviruses, at least in the laboratory mouse model studied. - Ichiro Nakamura, Kumiko Yoshimatsu, Byoung-Hee Lee, Megumi Okumura, Midori Taruishi, Koichi Araki, Hiroaki Kariwa, Ikuo Takashima, Jiro ArikawaARCHIVES OF VIROLOGY 153 8 1537 - 1542 2008年08月 [査読有り][通常論文]
To distinguish Thailand virus infection from infections with other hantaviruses, we established an ELISA serotyping system using a truncated nucleocapsid protein of Thailand virus lacking 49 amino acids at the N-terminus. In evaluations using patient and rodent sera, Thailand virus infection was readily distinguished from Hantaan and Seoul virus infections. Therefore, this ELISA system is an effective alternative to neutralization tests. - Arch VirolNakamura, I, Yoshimatsu, K, Lee, B. H, Okumura, M, Taruishi, M, Araki, K, Kariwa, H, Takashima, I, Arikawa, JDevelopment of a serotyping ELISA system for Thailand virus infection. 153 1537 1542 2008年 [査読無し][通常論文]
- Sara Chandy, Kumiko Yoshimatsu, Rainer G. Ulrich, Marc Mertens, Megumi Okumura, P. Rajendran, George T. John, Vinohar Balraj, Jayaprakash Muliyil, Joy Mammen, Priya Abraham, Jiro Arikawa, Gopalan SridharanTRANSACTIONS OF THE ROYAL SOCIETY OF TROPICAL MEDICINE AND HYGIENE 102 1 70 - 74 2008年01月 [査読有り][通常論文]
Hantaviruses are etiological agents of hemorrhagic fever with renal syndrome in many parts of Asia and Europe. There has been no documented case of hantavirus disease from India, although serological evidence exists. We investigated the prevalence of hantavirus in the Indian population and tried to identify potential risk groups for hantavirus infections. The presence of hantavirus-specific IgG antibodies was prospectively evaluated in 661 subjects belonging to different groups, i.e. patients with chronic renal disease, warehouse workers and tribal members engaged in rodent trapping. Healthy volunteer Wood donors were included as a control group. Thirty-eight seropositive samples were found using a combination of a commercial ELISA followed by an indirect immunofluorescence assay. Western blot using recombinant Hantaan virus nucleocapsid antigen confirmed the presence of anti-hantavirus IgG in 28 (74%) of the 38 sera tested. This study confirms the presence of hantaviruses in India and warrants increasing awareness of the problems of emerging pathogens and the threats they may pose to the public health system. (c) 2007 Royal Society of Tropical Medicine and Hygiene. Published by Elsevier Ltd. All rights reserved. - Midori Taruishi, Kumiko Yoshimatsu, Koichi Araki, Megumi Okumura, Ichiro Nakamura, Kilchl Kajino, Jiro ArikawaVIROLOGY 365 2 292 - 301 2007年09月 [査読有り][通常論文]
The major histocompatibility complex (MHC) class-I restricted epitope of Hantaan virus nucleocapsid protein (N) was identified using overlapping peptides and BALB/c mice. Using the MHC tetramer derived from the epitope, we found that the level of N-specific CD8(+) T cells increased to approximately 20% of all antigen-specific CD8(+) T cells in a mouse model of transient infection. However, N-specific CD8(+) T cells were undetectable in a mouse model of persistent infection, both in the persistently infected phase and in the convalescent phase. Levels of CD8(+) T cells producing interferon-gamma were weak in both the acute and convalescent phases in the persistently infected model. These results indicate that hantavirus strongly suppresses the production of N-specific CD8(+) T cells throughout the course of infection in persistently infected mice. Moreover, N-specific CD8(+) T cells were not effective in recovering persistently infected mice, despite the existence of abundant N antigen in vivo. (C) 2007 Elsevier Inc. All rights reserved. - Hiroaki Kariwa, Kumiko Yoshimatsu, Jiro ArikawaCOMPARATIVE IMMUNOLOGY MICROBIOLOGY AND INFECTIOUS DISEASES 30 5-6 341 - 356 2007年09月 [査読有り][通常論文]
Hantaviruses are enveloped RNA viruses that belong to the Hantavirus genus of the family Bunyaviridae. These viruses persistently infect their rodent reservoirs without causing disease. The virus is transmitted to humans via the inhalation of infectious aerosols generated from contaminated animal secretions or through the contaminated saliva of animal bites. Hantaviruses cause haemorrhagic fever with renal syndrome in Euro-Asia, and hantavirus pulmonary syndrome (HPS) in North and South America. Here, we review the epidemiology and epizootiology of hantavirus infection in Asian countries. (C) 2007 Elsevier Ltd. All rights reserved. - Arikawa J, Yoshimatsu K, Truong UT, Troung UNTrop Med Health 35 2 55 - 59 Japanese Society of Tropical Medicine 2007年09月 [査読有り][通常論文]
- Y. Matsuura, M. Suzuki, K. Yoshimatsu, J. Arikawa, I. Takashima, M. Yokoyama, H. Igota, K. Yamauchi, S. Ishida, D. Fukui, G. Bando, M. Kosuge, H. Tsunemitsu, C. Koshimoto, K. Sakae, M. Chikahira, S. Ogawa, T. Miyamura, N. Takeda, T. C. LiARCHIVES OF VIROLOGY 152 7 1375 - 1381 2007年07月 [査読無し][通常論文]
We examined 976 sika deer serum samples, 159 liver tissue samples and 88 stool samples collected from 16 prefectures in Japan, and performed ELISA and RT-PCR assays to detect antibodies to HEV and HEV RNA, respectively. Although 25 (2.6%) of 976 samples were positive for anti-HEV IgG, the antibody titers were very low. The OD values ranged between 0.018 and 0.486, forming a single distribution rather than a bimodal distribution, suggesting that the antibody detected in this study was not induced by HEV infection, or that deer have low sensitivity to HEV. HEV RNA was not detected in these samples, also suggesting that deer may not play a role as an HEV reservoir. - Megumi Okumura, Kumiko Yoshimatsu, Sanit Kumperasart, Ichiro Nakamura, Michiko Ogino, Midori Taruishi, Araya Sungdee, Sirima Pattamadilok, Ima Nurisa Ibrahim, Sri Erlina, Takashi Agui, Richard Yanagihara, Jiro ArikawaCLINICAL AND VACCINE IMMUNOLOGY 14 2 173 - 181 2007年02月 [査読有り][通常論文]
Thottapalayam virus (TPMV), a member of the genus Hantavirus in the family Bunyaviridae, was isolated from an insectivore, Suncus murinus (musk shrew), captured in southern India in 1964. While the isolation of TPMV predates the discovery of the prototype Hantaan virus, little is known about its genetics and biology. To date, preliminary evidence suggests that TPMV differs significantly, both antigenically and genetically, from all known rodent-borne hantaviruses. However, since detailed epizootiological studies have not been conducted, it is unclear if TPMV is naturally harbored by an insectivore host or if TPMV represents a "spillover" from its natural rodent reservoir host. Moreover, to what extent TPMV causes infection and/or disease in humans is not known. To address these issues, we first studied the antigenic profile of TPMV using monoclonal antibodies against Hantaan and Seoul viruses and pollyclonal immune sera against Puumala virus and TPMV. Armed with this newfound information, we developed an enzyme-linked immunosorbent assay system for the diagnosis of TPMV infections in shrews and humans, using a recombinant TPMV N antigen manipulated to have an E5/G6 epitope to be captured by monoclonal antibody clone E5/G6. Using this assay, we found anti-TPMV antibodies in sera from a patient with high fever of unknown etiology in Thailand and from two shrews captured in Indonesia. Seropositivity was verified by the indirect immunofluorescence antibody test, Western blotting analysis, and focus reduction neutralization test. Collectively, our data indicate that TPMV is harbored by Suncus murinus as its host in nature and is capable of infecting humans. - Hiroaki Kariwa, Kumari Lokugamage, Nandadeva Lokugamage, Hironobu Miyamoto, Kentaro Yoshii, Mina Nakauchi, Kumiko Yoshimatsu, Jiro Arikawa, Leonid I. Ivanov, Takuya Iwasaki, Ikuo TakashimaJAPANESE JOURNAL OF VETERINARY RESEARCH 54 4 145 - 161 2007年02月 [査読有り][通常論文]
Hantaviruses are causative agents of some severe human illnesses, including hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). The viruses are maintained by rodent hosts, and humans acquire infection by inhaling virus-contaminated excreta from infected animals. To examine the epidemiology of hantavirus infections in Japan and Far East Russia, we conducted epidemiological surveys in these regions. In Japan, anti-hantavirus antibodies were found in four rodent species, Clethrionomys rufocanus, Rattus norvegicus, R. rattus, and Apodemus speciosus. Although no new HFRS cases have been officially reported over the past 20 years in Japan, one member of the Japan Ground Self-Defense Force did test positive for hantavirus antibody. Repeated surveys in Far East Russia have revealed that two distinct hantavirus types cause severe HFRS in this region. Hantavirus sequences identified from A. peninsulae, fetal HFRS cases in Vladivostok, and Amur virus are highly similar to each other (> 92% identity), but they are less similar (similar to 84% identity) to the prototypical Hantaan virus, which is carried by A. agrarius. Phylogenetic analysis also indicates that Amur and A. peninsulae-associated viruses are distinct from Hantaan virus, suggesting that A. peninsulae is the reservoir animal for Amur virus, which causes severe HFRS. From HFRS patients in the Khabarovsk region, we identified viruses with nucleotide sequences that are more similar to Far East virus (> 96% identity) than to the Hantaan (88 - 89% identity) or Amur (81 - 83% identity) viruses. Phylogenetic analysis also indicates that the viruses from Khabarovsk HFRS patients are closely related to the Far East virus, and distinct from Amur virus. - Nur Hardy Abu Daud, Hiroaki Kariwa, Yoich Tanikawa, Ichiro Nakamura, Takahiro Seto, Daisuke Miyashita, Kentaro Yoshii, Mina Nakauchi, Kumiko Yoshimatsu, Jiro Arikawa, Ikuo TakashimaMICROBIOLOGY AND IMMUNOLOGY 51 11 1081 - 1090 2007年 [査読有り][通常論文]
Hokkaido virus (HOKV) is a member of the genus Hantavirus, in the family Bunyaviridae. To investigate HOKV infection in the host Myodes rufocanus, the grey red-backed vole, 199 animals were captured at Tobetsu (October 2004 and July 2005) and Nakagawa (October 2004) in Hokkaido, Japan, for detection of antibody, antigen, and viral RNA. In the surveys in Tobetsu (2004) and Nakagawa (2004), seropositive animals were detected at a frequency of 6.0% (5/84) and 10.4% (5/48), respectively. No seropositive animals were detected in Tobetsu in 2005. Seroprevalence in males in Tobetsu and Nakagawa in 2004 was 25 % (1/4) and 45.5 % (5111), respectively, which was higher than in females, at 5.0 % (4/80) and 0% (0/37), respectively (P<0.01). These results suggest that male animals play an important role in the maintenance of HOKV in M. rufocanus. Two females were seronegative but viral RNA-positive, indicating that these animals had acute infections before antibody was produced. Another five infected animals in Nakagawa were all male and had high levels of antibodies and viral RNA, suggesting that they had persistent infections. Viral RNA copies in organs of infected animals in Nakagawa were quantified by real-time polymerase chain reaction. Two acutely infected animals had >= 10 times the number of RNA copies in their lungs compared to those of persistently infected animals. In most cases, lungs or spleen had the highest RNA copy number, regardless of infection status. - Sirima Pattamadilok, Byoung-Hee Lee, Sanit Kumperasart, Kumiko Yoshimatsu, Megumi Okumura, Ichiro Nakamura, Koichi Araki, Yuvaluk Khoprasert, Prayadh Dangsupa, Pornpitak Panlar, Burkhard Jandrig, Detlev H. Krueger, Boris Klempa, Thomas Jaekel, Jonas Schmidt, Rainer Ulrich, Hiroaki Kariwa, Jiro ArikawaAMERICAN JOURNAL OF TROPICAL MEDICINE AND HYGIENE 75 5 994 - 1002 2006年11月 [査読有り][通常論文]
Phylogenetic investigations, sequence comparisons, and antigenic cross-re activity studies confirmed the classification of Thailand virus (THAIV) as a distinct hantavirus species. The examination of sera from 402 rodents trapped in 19 provinces of Thailand revealed that five greater bandicoot rats (Bandicota indica) and one lesser bandicoot rat (B. savilei) from four provinces were focus reduction neutralization test (FRNT) antibody-positive for THAIV. One of 260 patients from Surin province in Thailand (initially suspected of having contracted leptospirosis, but found to be negative) showed symptoms compatible with hemorrhagic fever with renal syndrome (HFRS). The serum of this patient showed high titers of hantavirus-reactive IgM and IgG. FRNT investigations confirmed virus-neutralizing antibodies against THAIV. These observations suggest that THAIV or THAI-like viruses occur throughout Indochina and may represent an additional causative agent of HFRS. - BH Lee, K Yoshimatsu, K Araki, M Okumura, Nakamura, I, J ArikawaVACCINE 24 15 2928 - 2934 2006年04月 [査読無し][通常論文]
We examined whether a vesicular stomatitis virus (VSV) pseudotype bearing the hantavirus envelope glycoproteins (GPs) G1 and G2 (VSV Delta G*HTN) could be used as a safe and effective alternative to native hantavirus. Mice were immunized with purified particles of VSV Delta G*HTN. After the second immunization, all mice produced anti-GP antibody as detected in ELISA and a neutralization test. After the third immunization, the mice were challenged with Hantaan virus. Neither anti-NP antibody production nor Hantaan virus-specific CD8 T-cell reactions were detected in these mice. The present study demonstrated the potential of using a pseudotype VSV system as a tool for developing a hantavirus vaccine. (c) 2005 Elsevier Ltd. All rights reserved. - BH Lee, K Yoshimatsu, K Araki, M Okumura, Nakamura, I, J ArikawaVACCINE 24 15 2928 - 2934 2006年04月 [査読有り][通常論文]
We examined whether a vesicular stomatitis virus (VSV) pseudotype bearing the hantavirus envelope glycoproteins (GPs) G1 and G2 (VSV Delta G*HTN) could be used as a safe and effective alternative to native hantavirus. Mice were immunized with purified particles of VSV Delta G*HTN. After the second immunization, all mice produced anti-GP antibody as detected in ELISA and a neutralization test. After the third immunization, the mice were challenged with Hantaan virus. Neither anti-NP antibody production nor Hantaan virus-specific CD8 T-cell reactions were detected in these mice. The present study demonstrated the potential of using a pseudotype VSV system as a tool for developing a hantavirus vaccine. (c) 2005 Elsevier Ltd. All rights reserved. - LJ Baek, H Kariwa, K Lokugamage, K Yoshimatsu, J Arikawa, Takashima, I, JI Kang, SS Moon, SY Chung, EJ Kim, HJ Kang, KJ Song, TA Klein, R Yanagihara, JW SongJOURNAL OF MEDICAL VIROLOGY 78 2 290 - 297 2006年02月 [査読有り][通常論文]
Hantaan (HTN) virus, the etiologic agent of clinically severe hemorrhagic fever with renal syndrome (HFRS), was first isolated in 1976 from lung tissue of a striped-field mouse (Apodemus agrarius) captured in Songnae-ri, Gyeonggi Province, Republic of Korea. Found primarily in mountainous areas, the Korean field mouse (A. peninsulae) is the second-most dominant field rodent species found throughout Korea. A new hantavirus, designated Soochong (SOO), was isolated in Vero E6 cells from four A. peninsulae captured in August 1997 at Mt. Gyebang in Hongcheon-gun, Mt. Gachil, Inje-gun, Gangwon Province, and in September 1998 at Mt. Deogyu, Muju-gun, Jeollabuk Province. The entire S, M, and L genomic segments of SOO virus, amplified by RT-PCR from lung tissues of seropositive A. peninsulae and from virus-infected Vero E6 cells, diverged from HTN virus (strain 76-118) by 15.6%, 22.8%, and 21.7% at the nucleotide level and 3.5%, 9.5%, and 4.6% at the amino acid level, respectively. Phylogenetic analyses of the nucleotide and deduced amino acid sequences, using the maximum parsimony and neighbor-joining methods, indicated that SOO virus was distinct from A. agrarius-borne HTN virus. SOO virus shared a common ancestry with Amur virus from Far East Russia, as well as with H5 and B78 hantaviruses, previously isolated from HFRS patients in China. Cross-focus-reduction neutralizating antibody tests showed that SOO virus which is the first hantavirus isolated in cell culture from A. peninsulae, could be classified as a new hantavirus serotype. - S Chandy, S Mitra, N Sathish, TS Vijayakumar, OC Abraham, MV Jesudason, P Abraham, K Yoshimatsu, J Arikawa, G SridharanINDIAN JOURNAL OF MEDICAL RESEARCH 122 3 211 - 215 2005年09月 [査読有り][通常論文]
Background & objectives: Hantaviruses are rodent-borne viruses of the family Bunyaviridae that have been identified as aetiological agents of two human diseases, haemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). There are no reports of hantavirus infections in humans from India, hence this pilot study was undertaken to provide the serological evidence of hantavirus infections in humans in south India. Methods: Serum samples were obtained from individuals with acute febrile illness and from voluntary blood donors, majority of whom were from south India. Serum samples were tested for anti-hantavirus IgM using a commercial enzyme immunoassay (EIA). Samples found positive by the EIA were tested by an indirect immunofluorescence assay (IFA) using slides coated with Seoul virus (SEOV) infected cells as substrate. Results: Of the 152 serum samples from individuals with pyrexic illness, 23 (14.7%) were positive for anti-hantavirus IgM by EIA. In contrast, only 5.7 per cent of healthy blood donors were positive by this assay. Eighteen of the 22 (82%) EIA-positive samples from patients were positive by the IFA assay. In contrast, only 2 of the 5 (40%) blood donor EIA positive samples were positive in the IFA assay. Interpretation & conclusion: The finding of this study indicated the possible presence of hantavirus infections in the human population of India presenting both as asymptomatic and symptomatic infections. Further studies need to be done to confirm the findings on a larger sample using molecular techniques. - 野生げっ歯類からのハンタウイルス抗原検出用ELISAの開発谷川 洋一, 苅和 宏明, 萩谷 友洋, Lokugamage Nandadeva, Lokugamage Kumari, 舘 敦史, 好井 健太朗, 吉松 組子, 有川 二郎, 高島 郁夫日本獣医学会学術集会講演要旨集 139回 185 - 185 (公社)日本獣医学会 2005年03月 [査読有り][通常論文]
- Jonas Schmidt, Burkhard Jandrig, Boris Klempa, Kumiko Yoshimatsu, Jiro Arikawa, Helga Meisel, Matthias Niedrig, Christian Pitra, Detlev H. Krüger, Rainer UlrichVirus Gene 30 1 37 - 48 2005年01月 [査読無し][通常論文]
Seoul virus (SEOV) is a hantavirus causing a mild to moderate form of hemorrhagic fever with renal syndrome that is distributed mainly in Asia. The nucleocapsid (N) protein-encoding sequence of SEOV (strain 80-39) was RT-PCR-amplified and cloned into a yeast expression vector containing a galactose-inducible promoter. A survey of the pattern of synonymous codon preferences for a total of 22 N protein-encoding hantavirus genes including 13 of SEOV strains revealed that there is minor variation in codon usage by the same gene in different viral genomes. Introduction of the expression plasmid into yeast Saccharomyces cerevisiae resulted in the high-level expression of a hexahistidine-tagged N protein derivative. The nickel-chelation chromatography purified, yeast-expressed SEOV N protein reacted in the immunoblot with a SEOV-specific monoclonal antibody and certain HTNV- and PUUV-cross-reactive monoclonal antibodies. The immunization of a rabbit with the recombinant N protein resulted in the induction of a high-titered antibody response. In ELISA studies, the N protein was able to detect antibodies in sera of experimentally infected laboratory rats and in human anti-hantavirus-positive sera or serum pools of patients from different geographical origin. The yeast-expressed SEOV N protein represents a promising antigen for development of diagnostic tools in serology, sero prevalence studies and vaccine development. © 2005 Springer Science+Business Media, Inc. - J Schmidt, B Jandrig, B Klempa, K Yoshimatsu, J Arikawa, H Meisel, M Niedrig, C Pitra, DH Kruger, R UlrichVIRUS GENES 30 1 37 - 48 2005年01月 [査読有り][通常論文]
Seoul virus (SEOV) is a hantavirus causing a mild to moderate form of hemorrhagic fever with renal syndrome that is distributed mainly in Asia. The nucleocapsid (N) protein-encoding sequence of SEOV (strain 80-39) was RT-PCR-amplified and cloned into a yeast expression vector containing a galactose-inducible promoter. A survey of the pattern of synonymous codon preferences for a total of 22 N protein-encoding hantavirus genes including 13 of SEOV strains revealed that there is minor variation in codon usage by the same gene in different viral genomes. Introduction of the expression plasmid into yeast Saccharomyces cerevisiae resulted in the high-level expression of a hexahistidine-tagged N protein derivative. The nickel-chelation chromatography purified, yeast-expressed SEOV N protein reacted in the immunoblot with a SEOV-specific monoclonal antibody and certain HTNV- and PUUV-cross-reactive monoclonal antibodies. The immunization of a rabbit with the recombinant N protein resulted in the induction of a high-titered antibody response. In ELISA studies, the N protein was able to detect antibodies in sera of experimentally infected laboratory rats and in human anti-hantavirus-positive sera or serum pools of patients from different geographical origin. The yeast-expressed SEOV N protein represents a promising antigen for development of diagnostic tools in serology, sero prevalence studies and vaccine development. - A pilot study for serological evidence of hantavirus infection in human population in south IndiaChandy, S, Mitra, S, Sathish, N, Vijayakumar, T. S, Abraham, O. C, Jesudason, M. V, Abraham, P, Yoshimatsu, K, Arikawa, J, Sridharan, GIndian J Med Res 122 211 215 2005年 [査読無し][通常論文]
- M. Okumura, K. Yoshimatsu, K. Araki, B. H. Lee, A. Asano, T. Agui, J. ArikawaArchives of Virology 149 12 2427 - 2434 2004年12月 [査読無し][通常論文]
Monoclonal antibody E5/G6 recognized a linear epitope common to hantavirus nucleocapsid proteins. Using synthetic peptides, we identified epitope E5/G6 as the 9 mer YEDVNGIRK (NP 165-173), in which D167, G170, I171, and R172 are indispensable. Furthermore, all the peptides synthesized using various hantavirus sequences bound MAb E5/G6 consistently, despite the existence of several amino acid variations in this region. These results indicate that MAb E5/G6 is a useful tool for detecting hantavirus antigen in rodent or patient tissues using Western blotting or other immunohistochemical assays. © Springer-Verlag 2004. - Michiko Ogino, Kumiko Yoshimatsu, Hideki Ebihara, Koichi Araki, Byoung-Hee Lee, Megumi Okumura, Jiro ArikawaJournal of Virology 78 19 10776 - 10782 2004年10月 [査読無し][通常論文]
Hantaan virus (HTNV)-infected Vero E6 cells undergo cell fusion with both infected and uninfected cells under low-pH conditions. Flow cytometry and fluorescence microscopy of HTNV-infected Vero E6 cells showed that envelope glycoproteins (GPs) were located both on the cell surface and in the cytoplasm. Neutralizing monoclonal antibodies (MAbs) against the G1 and G2 envelope GPs inhibited cell fusion, whereas nonneutralizing MAbs against G1 or G2 and MAbs against the nucleocapsid protein (NP) did not. Transfected Vero E6 cells that expressed GPs but not those that expressed NP fused and formed syncytia. These results indicate that HTNV GPs act as fusogens at the cell surface. No fusion activity was observed either in infected Vero cells that were passaged more than 150 times or in BHK-21 cells, although GPs appeared to localize to the cell surface. This variability in fusion induction suggests the involvement of host cell factors in the process of cell membrane fusion. - M Ogino, K Yoshimatsu, H Ebihara, K Araki, BH Lee, M Okumura, J ArikawaJOURNAL OF VIROLOGY 78 19 10776 - 10782 2004年10月 [査読有り][通常論文]
Hantaan virus (HTNV)-infected Vero E6 cells undergo cell fusion with both infected and uninfected cells under low-pH conditions. Flow cytometry and fluorescence microscopy of HTNV-infected Vero E6 cells showed that envelope glycoproteins (GPs) were located both on the cell surface and in the cytoplasm. Neutralizing monoclonal antibodies (MAbs) against the G1 and G2 envelope GPs inhibited cell fusion, whereas nonneutralizing MAbs against G1 or G2 and MAbs against the nucleocapsid protein (NP) did not. Transfected Vero E6 cells that expressed GPs but not those that expressed NP fused and formed syncytia. These results indicate that HTNV GPs act as fusogens at the cell surface. No fusion activity was observed either in infected Vero cells that were passaged more than 150 times or in BHK-21 cells, although GPs appeared to localize to the cell surface. This variability in fusion induction suggests the involvement of host cell factors in the process of cell membrane fusion. - 日本におけるハンタウイルス感染の動物伝染病学的及び疫学的研究(Epizootiological and epidemiological study of hantavirus infection in Japan)Lokugamage Nandadeva, 苅和 宏明, 好井 健太朗, 舘 敦史, 安藤 秀二, 福島 博, 土屋 公幸, 岩崎 琢也, 吉松 組子, 有川 二郎, 高島 郁夫日本獣医学会学術集会講演要旨集 138回 130 - 130 2004年08月 [査読有り][通常論文]
- Amur型ハンタウイルス糖蛋白の哺乳類細胞での発現と抗原性解析舘 敦史, 苅和 宏明, Lokugamage Kumari, Lokugamage Nandadeva, 谷川 洋一, 好井 健太朗, 吉松 組子, 有川 二郎, 高島 郁夫日本獣医学会学術集会講演要旨集 138回 131 - 131 (公社)日本獣医学会 2004年08月 [査読有り][通常論文]
- K Araki, K Yoshimatsu, BH Lee, M Okumura, H Kariwa, Takashima, I, J ArikawaArchives of Virology 149 7 1373 - 1382 2004年07月 [査読無し][通常論文]
To investigate age-dependent differences in hantavirus-specific CD8(+) T-cell responses, mice were inoculated with 0.1 50% newborn mouse lethal dose of Hantaan virus (HTNV) at 0, 3, 7, 14, or 35 days after birth. HTNV-specific CD8(+) T cells producing gamma interferon (IFN-gamma) were measured on day 30 after HTNV inoculation. Although no IFN-gamma-producing HTNV-specific CD8(+) T cells were detected in most of the mice inoculated with HTNV on day 0 after birth, most mice inoculated at 3, 7, 14, or 35 days had HTNV-specific CD8(+) T cells. The production of tumor necrosis factor alpha (TNF-alpha) by IFN-gamma-producing CD8(+) T cells and the cytotoxic activity against HTNV-infected target cells were similar in immature and adult mice. However, the number of IFN-gamma-producing HTNV-specific CD8(+) T cells was significantly less in mice inoculated with HTNV at 3 days than in older mice. In addition, a strong correlation between HTNV persistence and a lack of HTNV-specific CD8(+) T cells was observed. These results suggest that mice over 7 days old have the ability to induce functional HTNV-specific CD8(+) T-cell responses that are indistinguishable from the responses of adult mice, and that HTNV-specific CD8(+) T cells are important for clearance of HTNV. - K Lokugamage, H Kariwa, N Lokugamage, H Miyamoto, M Iwasa, T Hagiya, K Araki, A Tachi, T Mizutani, K Yoshimatsu, J Arikawa, Takashima, IVIRUS RESEARCH 101 2 127 - 134 2004年05月 [査読有り][通常論文]
The genetic and antigenic characteristics of the Amur (AMR) and Far East (FE) virus lineages, which are both within the genus Hantavirus, were studied. Representative viruses, H5 and B78 for AMR and Bao, 14 for FE, were used. The entire small (S) and medium (M) segments, except for the 3'- and 5'-ends, were sequenced. The deduced amino acid sequences of AMR had 96.7 and 92.0-92.2% identities with the Hantaan (HTN) virus in the S and M segments, respectively. The amino acid sequences of FE had 99.1 and 97.9% identities in the S and M segments, respectively. The three viral strains and HTN virus had similar binding patterns to a panel of monoclonal antibodies (MAbs), except that one MAb did not bind AMR. However, sera from Apodemus peninsulae, naturally infected with AMR virus, neutralized homologous viruses at 1: 160 to 1:320 dilutions and HTN at 1:20 to 1:40 dilutions. The anti-AMR serum neutralized homologous viruses at a 1:80 dilution and HTN at a 1:40 dilution. The anti-HTN serum did not neutralize AMR (< 1:40 dilution), although it had a high neutralizing titer (1:320) against the homologous virus. Therefore, we suggest that AMR virus may constitute a distinct serotype within the genus Hantavirus. (C) 2004 Elsevier B.V. All rights reserved. - K Araki, K Yoshimatsu, BH Lee, H Kariwa, Takashima, I, J ArikawaVIROLOGY 322 2 318 - 327 2004年05月 [査読有り][通常論文]
We established a viral persistence model that involves the adoptive transfer of spleen cells from immunocompetent mice (H-2(d)) into Hantaan virus (HTNV)-infected severe combined immunodeficient (SCID, H-2(d)) mice. The infection is maintained despite the presence of neutralizing antibodies, without apparent signs of disease, and there is a correlation between HTNV persistence and the lack of HTNV-specific CD8(+) T cells. In addition, disseminated HTNV infection before the initiation of immune responses appears to be important for virus persistence. The suppression of HTNV-specific CD8(+) T cells in the present model appears to occur at the periphery. The present study also demonstrates that CD8(+) T cells contribute to the clearance of HTNV Thus, it seems that HTNV-specific CD8(+) T cells play a key role in HTNV persistence in mice. This model of viral persistence is useful for studies of immune responses and immunocytotherapy against viral infection. (C) 2004 Elsevier Inc. All rights reserved. - K Lokugamage, H Kariwa, N Lokugamage, M Iwasa, T Hagiya, K Araki, A Tachi, T Mizutani, K Yoshimatsu, J Arikawa, T Iwasaki, Takashima, IJAPANESE JOURNAL OF VETERINARY RESEARCH 51 3-4 143 - 149 2004年02月 [査読有り][通常論文]
The virulence of hantaviruses that are antigenically related but have different genetic characteristics from the prototype of hantavirus, Hantaan (HTN) virus, was examined in newborn mice. The H5 and B78 strains of the Amur (AMR) genotype, the Bao14 strain of the Far East (FE) genotype, and the 76 - 118 strain of HTN virus were inoculated subcutaneously ( 1 focus-forming unit; FFU) into newborn mice. All of the AMR and FE genotype viruses inoculated mice were died by 16 days post-infection (dpi) and 21 dpi, respectively, while 50% of the HTN virus inoculated mice survived until 30 dpi. The AMR and FE genotype viruses inoculated mice had high viral titers in the lung (1.3x10(6) to 1.3x10(8) FFU/gram [g] tissue), brain (2.1x10(7) to 1.2x10(9) FFU/g tissue), and kidney(2.5x10(5) to 1.6x10(7) FFU/g tissue), and showed a detectable level of antibodies (titers 1 : 16 -1 : 32) at 14 dpi. In contrast, the HTN virus infected mice had viruses only in the lungs at low titers (1.1-5.3x10(5) FFU/g tissue). Observations of body-weight changes revealed that the AMR and FE genotype viruses inoculated mice had lower growth rates than the HTN virus inoculated mice. These data suggest that the AMR and FE genotype viruses are more virulent than the HTN virus in newborn mice. - Araki, K., Yoshimatsu, K., Lee, B.-H., Kariwa, H., Takashima, I., Arikawa, J.Virology 322 2 2004年 [査読無し][通常論文]
- N Lokugamage, H Kariwa, K Lokugamage, MA Iwasa, T Hagiya, K Yoshii, A Tachi, S Ando, H Fukushima, K Tsuchiya, T Iwasaki, K Araki, K Yoshimatsu, J Arikawa, T Mizutani, K Osawa, H Sato, K TakashimaMICROBIOLOGY AND IMMUNOLOGY 48 11 843 - 851 2004年 [査読有り][通常論文]
Epizootiological surveys on hantavirus infections in rodents were carried out in various areas of Japan, including the four major islands of Hokkaido, Honshu, Shikoku, and Kyushu from 2000 to 2003. A total of 1,221 rodents and insectivores were captured. Seropositive animals were found in Apodemus (A.) speciosus (5/482, 1.0%), Rattus (R.) norvegicus (4/364, 1.1%), R. rattus (3/45, 6.7%), and Clethrionomys (C.) rufocanus (7/197, 3.6%). The partial S segment was amplified from one seropositive R. rattus captured at Hakodate. The nucleotide sequence showed 96% identity with the Seoul virus (SEOV) prototype strain SR-11. In addition, we conducted an epidemiological survey on human hantavirus infection in a high-risk population, the personnel of the Japan Ground Self-defense Force on Hokkaido. One out of 207 human blood samples was positive for anti-hantavirus antibody by IFA, ELISA, and WB analysis. The result of the serotype specific ELISA indicates that this individual acquired SEOV infection. This study indicates that A. speciosus, R. norvegicus, R. rattus, and C. rufocanus carry hantaviruses as the reservoir animals in Japan. Infected R. rattus and R. norvegicus in port areas could be the sources of human SEOV infection and a threat to travelers and individuals working in seaports. - BH Lee, K Yoshimatsu, A Maeda, K Ochiai, M Morimatsu, K Araki, M Ogino, S Morikawa, J ArikawaVirus Res 98 1 83 - 91 2003年12月 [査読無し][通常論文]
We performed yeast two-hybrid screening of a human kidney cell cDNA library to study the biological role of the hantavirus nucleocapsid protein (NP). We found that Seoul virus (SEOV) and Hantaan virus (HTNV) NPs were associated with small ubiquitin-like modifier (SUMO)-1-interacting proteins PIAS 1, PIASxbeta, HIPK2, CHD3, and TTRAP, which interacted with the SUMO-1 conjugating enzyme (Ubc-9) and SUMO-1 in the yeast two-hybrid assay. Interactions between the HIPK2, CHD3, and TTRAP proteins and SEOV NP were also shown in a mammalian two-hybrid assay. However, there was no interaction between PIAS proteins and NP, which was probably due to the inhibitory effect of PIAS on transcription in the mammalian two-hybrid assay. Nevertheless, a co-expression experiment suggested the existence of a PIAS-NP interaction in the cytoplasm. The region spanning amino acids 100-125 of SEOV NP, which represents a critical region for NP-NP polymerization. was found to be responsible for the interaction with SUMO-1-related molecules in both the yeast and mammalian two-hybrid assays. These results add to the information on interactions of hantavirus NP and host cellular proteins. (C) 2003 Elsevier B.V. All rights reserved. - BH Lee, K Yoshimatsu, A Maeda, K Ochiai, M Morimatsu, K Araki, M Ogino, S Morikawa, J ArikawaVIRUS RESEARCH 98 1 83 - 91 2003年12月 [査読有り][通常論文]
We performed yeast two-hybrid screening of a human kidney cell cDNA library to study the biological role of the hantavirus nucleocapsid protein (NP). We found that Seoul virus (SEOV) and Hantaan virus (HTNV) NPs were associated with small ubiquitin-like modifier (SUMO)-1-interacting proteins PIAS 1, PIASxbeta, HIPK2, CHD3, and TTRAP, which interacted with the SUMO-1 conjugating enzyme (Ubc-9) and SUMO-1 in the yeast two-hybrid assay. Interactions between the HIPK2, CHD3, and TTRAP proteins and SEOV NP were also shown in a mammalian two-hybrid assay. However, there was no interaction between PIAS proteins and NP, which was probably due to the inhibitory effect of PIAS on transcription in the mammalian two-hybrid assay. Nevertheless, a co-expression experiment suggested the existence of a PIAS-NP interaction in the cytoplasm. The region spanning amino acids 100-125 of SEOV NP, which represents a critical region for NP-NP polymerization. was found to be responsible for the interaction with SUMO-1-related molecules in both the yeast and mammalian two-hybrid assays. These results add to the information on interactions of hantavirus NP and host cellular proteins. (C) 2003 Elsevier B.V. All rights reserved. - N Lokugamage, H Kariwa, K Lokugamage, T Hagiya, H Miyamoto, MA Iwasa, K Araki, K Yoshimatsu, J Arikawa, T Mizutani, Takashima, IJOURNAL OF VETERINARY MEDICAL SCIENCE 65 11 1189 - 1194 2003年11月 [査読無し][通常論文]
Puumala (PUU) virus and PUU-related viruses are difficult to isolate in cell culture. To determine whether animal inoculation would be a better alternative for virus recovery, the Sotkamo strain of PUU virus was inoculated into several animal species. Newborn Mongolian gerbils (MGs), mice, and rats were infected with the Sotkamo strain by intracerebral (ic), intraperitoneal (ip), and subcutaneous (sc) inoculation. Antibodies to PUU appeared in MGs at 30 days post-infection (dpi), and in mice and rats at 15 dpi. Interestingly, virus appeared at 7 dpi in lung and brain of MGs inoculated via ic and ip routes. Virus was detected in all tested tissues of MGs at 15 dpi, with a peak level of 1.36 x 10(5) focus forming units (FFU)/g in brain tissue. The virus titer declined with the onset of the antibody response and became undetectable by 75 dpi, when the antibody titer reached the maximum level. The appearance of the virus in mice and rats was delayed as compared to MGs, and the virus titer was apparently lower, at approximately 4 to 8 x 10(3) FFU/g, at 15 dpi. In addition, lung homogenates of antibody-positive Clethrionomys (C) rufocanus (captured in Tobetsu, Hokkaido, Japan) were inoculated into MGs by the ic route. PUU-related viral RNA was detected at 16 dpi in the brains of MG inoculated with the lung homogenate, and antibodies were detected at 45 dpi. These findings indicate that newborn MG inoculation is an efficient method to recover PUU and PUU-related viruses. - N Lokugamage, H Kariwa, K Lokugamage, T Hagiya, H Miyamoto, MA Iwasa, K Araki, K Yoshimatsu, J Arikawa, T Mizutani, Takashima, IJOURNAL OF VETERINARY MEDICAL SCIENCE 65 11 1189 - 1194 2003年11月 [査読有り][通常論文]
Puumala (PUU) virus and PUU-related viruses are difficult to isolate in cell culture. To determine whether animal inoculation would be a better alternative for virus recovery, the Sotkamo strain of PUU virus was inoculated into several animal species. Newborn Mongolian gerbils (MGs), mice, and rats were infected with the Sotkamo strain by intracerebral (ic), intraperitoneal (ip), and subcutaneous (sc) inoculation. Antibodies to PUU appeared in MGs at 30 days post-infection (dpi), and in mice and rats at 15 dpi. Interestingly, virus appeared at 7 dpi in lung and brain of MGs inoculated via ic and ip routes. Virus was detected in all tested tissues of MGs at 15 dpi, with a peak level of 1.36 x 10(5) focus forming units (FFU)/g in brain tissue. The virus titer declined with the onset of the antibody response and became undetectable by 75 dpi, when the antibody titer reached the maximum level. The appearance of the virus in mice and rats was delayed as compared to MGs, and the virus titer was apparently lower, at approximately 4 to 8 x 10(3) FFU/g, at 15 dpi. In addition, lung homogenates of antibody-positive Clethrionomys (C) rufocanus (captured in Tobetsu, Hokkaido, Japan) were inoculated into MGs by the ic route. PUU-related viral RNA was detected at 16 dpi in the brains of MG inoculated with the lung homogenate, and antibodies were detected at 45 dpi. These findings indicate that newborn MG inoculation is an efficient method to recover PUU and PUU-related viruses. - Detection of antibody for the serodiagnosis of hantavirus infection in different rodent species.(共著）BH Lee, K Yoshimatsu, K Araki, M Ogino, M Okumura, K Tsuchiya, H Kariwa, J ArikawaArch Virol 148 10 1885 - 1897 2003年10月 [査読無し][通常論文]
Peroxidase-labeled staphylococcal protein A, streptococcal protein G, and antibodies directed against Mus musculus (mouse), Rattus norvegicus (rat), Mesocretus auratus (hamster), and Peromyscus leucopus were examined for their reactivity with immunoglobulin G (IgG) from various rodent species. The purpose of this study was to identify the optimal secondary antibodies or reagents for specific serodiagnosis of hantavirus infection in various rodent species. Using ELISA, a total of 65 sera from 29 rodent species of the family Muridae and one serum sample from family Octodontidae were compared for IgG reactivity with the six different reagents. The results demonstrate that the reactivities of the secondary antibodies and reagents to the sera varied, even among sera from rodents of the same genus. Hantavirus-specific antibody ELISA revealed that hantavirus-infected rodent sera obtained from M. musculus, R. norvegicus, Apodemus agrarius, A. peninsulae, and Bandicota indica bound to the six different conjugates in a similar pattern as that detected in IgG ELISA. These results indicate that the applicability of secondary antibodies and protein A and G should be carefully evaluated before use for serodiagnosis in different rodent species. - H Kariwa, H Tanabe, T Mizutani, Y Kon, K Lokugamage, N Lokugamage, MA Iwasa, T Hagiya, K Araki, K Yoshimatsu, J Arikawa, Takashima, IArchives of Virology 148 9 1671 - 1685 2003年09月 [査読無し][通常論文]
Seoul virus is a hantavirus that causes hemorrhagic fever with renal syndrome (HFRS). The virion has a tripartite (S, M, and L) negative-stranded RNA genome, which is characteristic of the family Bunyaviridae. However, the molecular basis of virus replication is not well known. We established a Northern blot hybridization (NB) procedure using digoxygenin-labeled RNA probes, to quantitate the hantaviral plus- and minus-strand RNAs separately. Virus RNA replication was analyzed in infected Vero E6 cells. When the Vero E6 cells were infected with Seoul virus strain KI-83-262 (KI) at m.o.i.=0.25, the plus-strand RNA was detected within 1 h post-infection (hpi), and the minus-strand RNA was detected subsequently. Using laser confocal microscopy, the nucleocapsid protein (NP) was detected within 2 hpi, and accumulated as scattered granules in the cytoplasm until 24 hpi. In contrast, the G2 protein first appeared at 8 hpi, was immediately transported to the Golgi, and accumulated in the Golgi until 24 hpi. Infectious virus particles were released into the medium at 24 h hpi. These findings indicate that hantavirus RNA replication starts with the appearance of NP at 2 hpi, glycoproteins then accumulate gradually in the Golgi, and virion formation is initiated once the viral RNAs and proteins have accumulated. - H Miyamoto, H Kariwa, K Araki, K Lokugamage, D Hayasaka, BZ Cui, N Lokugamage, LI Ivanov, T Mizutani, MA Iwasa, K Yoshimatsu, J Arikawa, Takashima, IArch Virol 148 8 1543 - 1556 2003年08月 [査読無し][通常論文]
Hemorrhagic fever with renal syndrome (HFRS) is endemic in East Asia and Europe. The disease is caused by several viruses belonging to the genus Hantavirus, including the Hantaan virus (HTNV), Seoul virus (SEOV), Dobrava Belgrade virus (DOBV), and Puumala virus (PUUV). Recently, HTNV-related viruses, Amur (AMR) and Far East (FE) genotypes were identified as causative agents of HFRS in Far Eastern Russia. To investigate the epidemiology of HFRS and virus transmission, we collected sera from 17 acute and 32 convalescent patients who were clinically diagnosed with HFRS in the Khabarovsk region of Far Eastern Russia, and detected anti-hantavirus antibodies using an ELISA that can differentiate the infected virus serotype using truncated hantavirus nucleocapsid protein antigen. Sixteen of the 17 acute phase patients had antibodies to hantavirus, and all the positive sera had higher optical densities for HTNV-specific antigen than for SEOV-, DOBV-, or PUUV-specific antigens. The partial M segment of the viral genome was amplified from blood clots from three acute patients by PCR. The nucleotide sequences had closer identities to the FE genotype (> 96%) than to the prototype HTNV (88 to 89%) or AMR genotype (81 to 83%). A phylogenetic analysis found that the virus sequences from the patients clustered with the FE type, and were distinct from the AMR type. Thirty-one of 32 convalescent patient sera had antibodies to HTNV-specific antigen. These data suggest ELISA system can detect HTNV-specific antibodies to the FE type, which may be responsible for most of the HFRS in Khabarovsk. - Hantavirus-specific CD8(+)-T-cell responses in newborn mice persistently infected with Hantaan virusK Araki, K Yoshimatsu, BH Lee, H Kariwa, Takashima, I, J ArikawaJOURNAL OF VIROLOGY 77 15 8408 - 8417 2003年08月 [査読有り][通常論文]
The relationship between virus-specific CD8(+)-T-cell responses and viral persistence was studied in mice by using Hantaan virus (HTNA). We first established a simple method for measuring levels of virus-specific CD8(+) T cells by How cytometry. Next, to produce a mouse model of persistent HTNV infection, newborn mice were inoculated subcutaneously within 24 h of birth with 1 or 0.150% newborn mouse lethal dose of HTNV. All mice that escaped lethal infection were persistently infected with HTNV until at least 30 days after virus inoculation and had no virus-specific CD8(+) T cells producing gamma interferon (IFN-gamma). Subsequently, the virus was eliminated from some of the mice, depending on the appearance of functional virus-specific CD8(+) T cells, which have the ability to produce IFN-gamma and tumor necrosis factor alpha (TNF-alpha) and have cytotoxic activity. Neutralizing antibodies were detected in all mice, regardless of the presence or absence of virus. In the acute phase, which occurs within 30 days of infection, IFN-gamma-producing HTNV-specific CD8(+) T cells were detected on day 15 after virus inoculation. However, TNF-alpha production and the cytotoxic activity of these specific CD8(+) T cells were impaired and HTNV was not removed. Almost all of these specific CD8(+) T cells disappeared by day 18. These results suggest that functional HTNV-specific CD8(+) T cells are important for clearance of HTNV. - K Yoshii, D Hayasaka, A Goto, M Obara, K Araki, K Yoshimatsu, J Arikawa, L Ivanov, T Mizutani, H Kariwa, Takashima, IJOURNAL OF VIROLOGICAL METHODS 108 2 171 - 179 2003年03月 [査読有り][通常論文]
A recombinant plasmid that expresses the tick-borne encephalitis (TBE) virus premembrane (prM) and envelope (E) proteins in mammalian cells was constructed. Recombinant proteins retained antigenic and conformational structures similar to those of native virus proteins, and transfected cells released virus-like particles (VLPs), which were 1.13-1.14 g/ml in density and 20-30 nm in diameter, into the culture medium. Recombinant E proteins were used for the development of an enzyme-linked immunosorbent assay (ELISA) to detect TBE virus-specific IgM and IgG antibodies in serum. The results of this ELISA correlated well with the results of commercial ELISA, when tested with 95 serum samples from clinically TBE-suspected patients. In addition, ELISA using recombinant antigens showed no cross-reactivity against serum from Japanese encephalitis (JE) patients, despite the cross-reactivity shown by commercial ELISA systems. These observations indicated that this newly developed ELISA system could distinguish tick-borne encephalitis from Japanese encephalitis infection, and that it constitutes a useful and safe alternative to conventional ELISA systems. (C) 2002 Published by Elsevier Science B.V. - Araki, K., Yoshimatsu, K., Lee, B.-H., Kariwa, H., Takashima, I., Arikawa, J.Journal of Virology 77 15 2003年 [査読無し][通常論文]
- Kentarou Yoshii, Daisuke Hayasaka, Akiko Goto, Mayumi Obara, Koichi Araki, Kumiko Yoshimatsu, Jiro Arikawa, Leonoid Ivanov, Tetsuya Mizutani, Hiroaki Kariwa, Ikuo TakashimaJournal of Virological Methods 108 2 171 - 179 2003年 [査読無し][通常論文]
A recombinant plasmid that expresses the tick-borne encephalitis (TBE) virus premembrane (prM) and envelope (E) proteins in mammalian cells was constructed. Recombinant proteins retained antigenic and conformational structures similar to those of native virus proteins, and transfected cells released virus-like particles (VLPs), which were 1.13-1.14 g/ml in density and 20-30 nm in diameter, into the culture medium. Recombinant E proteins were used for the development of an enzyme-linked immunosorbent assay (ELISA) to detect TBE virus-specific IgM and IgG antibodies in serum. The results of this ELISA correlated well with the results of commercial ELISA, when tested with 95 serum samples from clinically TBE-suspected patients. In addition, ELISA using recombinant antigens showed no cross-reactivity against serum from Japanese encephalitis (JE) patients, despite the cross-reactivity shown by commercial ELISA systems. These observations indicated that this newly developed ELISA system could distinguish tick-borne encephalitis from Japanese encephalitis infection, and that it constitutes a useful and safe alternative to conventional ELISA systems. © 2002 Published by Elsevier Science B.V. - M Ogino, H Ebihara, BH Lee, K Araki, A Lundkvist, Y Kawaoka, K Yoshimatsu, J ArikawaClinical and diagnostic laboratory immunology. 10 1 154 - 160 2003年01月 [査読無し][通常論文]
A vesicular stomatitis virus (VSV) pseudotype bearing hantavirus envelope glycoproteins was produced and used in a neutralization test as a substitute for native hantavirus. The recombinant VSV, in which the enveloped protein gene (G) was replaced by the green fluorescent protein gene and complemented with G protein expressed in trans (VSVDeltaG*G), was kindly provided by M. A. Whitt. 293T cells were transfected with plasmids for the expression of envelope glycoproteins (G1 and G2) of HTNV or SEOV and were then infected with VSVDeltaG*G. Pseudotype VSV with the Hantaan (VSVDeltaG*-HTN) or Seoul (VSVDeltaG*-SEO) envelope glycoproteins were harvested from the culture supernatant. The number of infectious units (IU) of the pseudotype VSVs ranged from 10(5) to 10(6)/ml. The infectivity of VSVDeltaG*-HTN and VSVDeltaG*-SEO was neutralized with monoclonal antibodies, immune rabbit sera, and sera from patients with hemorrhagic fever with renal syndrome, and the neutralizing titers were similar to those obtained with native hantaviruses. These results show that VSVDeltaG*-HTN and -SEO can be used as a rapid, specific, and safe neutralization test for detecting hantavirus-neutralizing antibodies as an effective substitute for the use of native hantaviruses. Furthermore, the IU of VSVDeltaG*-HTN and -SEO did not decrease by more than 10-fold when stored at 4degreesC for up to 30 days. The stability of the pseudotype viruses allows distribution of the material to remote areas by using conventional cooling boxes for use as a diagnostic reagent. - Journal of Virology J. Virol. 77(2),943-952 2003年 [査読無し][通常論文]
- A Maeda, BH Lee, K Yoshimatsu, M Saijo, Kurane, I, J Arikawa, S MorikawaVirology 305 2 288 - 297 2003年01月 [査読無し][通常論文]
Small ubiquitin-like modifier-1 (SUMO-1) conjugating enzyme 9 (Ubc9) conjugates SUMO-1 to target proteins and modulates cellular processes such as signal transduction, transcription regulation, and cell growth regulation. We demonstrated here that the nucleocapsid protein (NP) of Hantaan virus (HTNV) was associated with Ubc9 and SUMO-1 in vivo. Analysis of the interaction between the truncated NPs and Ubc9 revealed that the amino acid residues at the positions between 101 and 238 in the NP were responsible for the interaction. Furthermore, a consensus binding motif of Ubc9 and SUMO-1, MKAE, within this region, especially the second amino acid of the motif, K residue, was crucial for the interaction, and the interaction was essential for the NP to be localized in the perinuclear region. These results indicate that the assembly of the HTNV-NP is regulated by the interaction between the NP and Ubc9. This is the first report to demonstrate the interaction of Ubc9 with a structural protein of negative-strand RNA viruses. (C) 2003 Elsevier Science (USA). - K Yoshimatsu, BH Lee, K Araki, M Morimatsu, M Ogino, H Ebihara, J ArikawaJOURNAL OF VIROLOGY 77 2 943 - 952 2003年01月 [査読有り][通常論文]
Multimerization of the Hantaan virus nucleocapsid protein (NP) in Hantaan virus-infected Vero E6 cells was observed in a competitive enzyme-linked immunosorbent assay (ELISA). Recombinant and truncated NPs of Hantaan, Seoul, and Dobrava viruses lacking the N-terminal 49 amino acids were also detected as multimers. Although truncated NPs of Hantaan virus lacking the N-terminal 154 amino acids existed as a monomer, those of Seoul and Dobrava formed multimers. The multimerized truncated NP antigens of Seoul and Dobrava viruses could detect serotype-specific antibodies, whereas the monomeric truncated NP antigen of Hantaan virus lacking the N-terminal 154 amino acids could not, suggesting that a hantavirus serotype-specific epitope on the NP results in multimerization. The NP-NP interaction was also detected by using a yeast two-hybrid assay. Two regions, amino acids 100 to 125 (region 1) and amino acids 404 to 429 (region 2), were essential for the NP-NP interaction in yeast. The NP of Seoul virus in which the tryptophan at amino acid number 119 was replaced by alanine (W119A mutation) did not multimerize in the yeast two-hybrid assay, indicating that tryptophan 119 in region 1 is important for the NP-NP interaction in yeast. However, W119A mutants expressed in mammalian cells were detected as the multimer by using competitive ELISA. Similarly, the truncated NP of Seoul virus expressing amino acids 155 to 429 showed a homologous interaction in a competitive ELISA but not in the yeast two-hybrid assay, indicating that the C-terminal region is important for the multimerization detected by competitive ELISA. Combined, the results indicate that several steps and regions are involved in multimerization of hantavirus NP. - M Ogino, H Ebihara, BH Lee, K Araki, A Lundkvist, Y Kawaoka, K Yoshimatsu, J ArikawaCLINICAL AND DIAGNOSTIC LABORATORY IMMUNOLOGY 10 1 154 - 160 2003年01月 [査読有り][通常論文]
A vesicular stomatitis virus (VSV) pseudotype bearing hantavirus envelope glycoproteins was produced and used in a neutralization test as a substitute for native hantavirus. The recombinant VSV, in which the enveloped protein gene (G) was replaced by the green fluorescent protein gene and complemented with G protein expressed in trans (VSVDeltaG*G), was kindly provided by M. A. Whitt. 293T cells were transfected with plasmids for the expression of envelope glycoproteins (G1 and G2) of HTNV or SEOV and were then infected with VSVDeltaG*G. Pseudotype VSV with the Hantaan (VSVDeltaG*-HTN) or Seoul (VSVDeltaG*-SEO) envelope glycoproteins were harvested from the culture supernatant. The number of infectious units (IU) of the pseudotype VSVs ranged from 10(5) to 10(6)/ml. The infectivity of VSVDeltaG*-HTN and VSVDeltaG*-SEO was neutralized with monoclonal antibodies, immune rabbit sera, and sera from patients with hemorrhagic fever with renal syndrome, and the neutralizing titers were similar to those obtained with native hantaviruses. These results show that VSVDeltaG*-HTN and -SEO can be used as a rapid, specific, and safe neutralization test for detecting hantavirus-neutralizing antibodies as an effective substitute for the use of native hantaviruses. Furthermore, the IU of VSVDeltaG*-HTN and -SEO did not decrease by more than 10-fold when stored at 4degreesC for up to 30 days. The stability of the pseudotype viruses allows distribution of the material to remote areas by using conventional cooling boxes for use as a diagnostic reagent. - A Maeda, BH Lee, K Yoshimatsu, M Saijo, Kurane, I, J Arikawa, S MorikawaVIROLOGY 305 2 288 - 297 2003年01月 [査読有り][通常論文]
Small ubiquitin-like modifier-1 (SUMO-1) conjugating enzyme 9 (Ubc9) conjugates SUMO-1 to target proteins and modulates cellular processes such as signal transduction, transcription regulation, and cell growth regulation. We demonstrated here that the nucleocapsid protein (NP) of Hantaan virus (HTNV) was associated with Ubc9 and SUMO-1 in vivo. Analysis of the interaction between the truncated NPs and Ubc9 revealed that the amino acid residues at the positions between 101 and 238 in the NP were responsible for the interaction. Furthermore, a consensus binding motif of Ubc9 and SUMO-1, MKAE, within this region, especially the second amino acid of the motif, K residue, was crucial for the interaction, and the interaction was essential for the NP to be localized in the perinuclear region. These results indicate that the assembly of the HTNV-NP is regulated by the interaction between the NP and Ubc9. This is the first report to demonstrate the interaction of Ubc9 with a structural protein of negative-strand RNA viruses. (C) 2003 Elsevier Science (USA). - A Takakura, K Goto, T Itoh, K Yoshimatsu, Takashima, I, J ArikawaEXPERIMENTAL ANIMALS 52 1 25 - 30 2003年01月 [査読有り][通常論文]
A recombinant nucleocapsid protein of Hantaan virus (HTN) 76-118 strain expressed in E. coli was applied as a serodiagnostic antigen in an enzyme-linked immunosorbent assay (rHTN-ELISA) for detection of hantavirus antibody in rat sera. The sensitivity and specificity of the rHTN-ELISA were compared with those of the indirect immunofluoresent assay (IFA) using virus-infected cells. The sensitivity of rHTN-ELISA was similar to that of the IFA both in experimentally SR-11 infected rat and naturally infected rat sera. Sera showing a low antibody titer in IFA and suspected to be negative by other methods were also found to be negative in rHTN-ELISA. These results indicate that rHTN-ELISA is effective as a screening method for serodiagnosis of hantaviruses, because of its high sensitivity, specificity, safety and suitability for processing large number of samples. - Kumiko Yoshimatsu, Byoung-Hee Lee, Koichi Araki, Masami Morimatsu, Michiko Ogino, Hideki Ebihara, Jiro ArikawaJournal of Virology 77 2 943 - 952 2003年01月 [査読有り][通常論文]
Multimerization of the Hantaan virus nucleocapsid protein (NP) in Hantaan virus-infected Vero E6 cells was observed in a competitive enzyme-linked immunosorbent assay (ELISA). Recombinant and truncated NPs of Hantaan, Seoul, and Dobrava viruses lacking the N-terminal 49 amino acids were also detected as multimers. Although truncated NPs of Hantaan virus lacking the N-terminal 154 amino acids existed as a monomer, those of Seoul and Dobrava formed multimers. The multimerized truncated NP antigens of Seoul and Dobrava viruses could detect serotype-specific antibodies, whereas the monomeric truncated NP antigen of Hantaan virus lacking the N-terminal 154 amino acids could not, suggesting that a hantavirus serotype-specific epitope on the NP results in multimerization. The NP-NP interaction was also detected by using a yeast two-hybrid assay. Two regions, amino acids 100 to 125 (region 1) and amino acids 404 to 429 (region 2), were essential for the NP-NP interaction in yeast. The NP of Seoul virus in which the tryptophan at amino acid number 119 was replaced by alanine (W119A mutation) did not multimerize in the yeast two-hybrid assay, indicating that tryptophan 119 in region 1 is important for the NP-NP interaction in yeast. However, W119A mutants expressed in mammalian cells were detected as the multimer by using competitive ELISA. Similarly, the truncated NP of Seoul virus expressing amino acids 155 to 429 showed a homologous interaction in a competitive ELISA but not in the yeast two-hybrid assay, indicating that the C-terminal region is important for the multimerization detected by competitive ELISA. Combined, the results indicate that several steps and regions are involved in multimerization of hantavirus NP. - K Lokugamage, H Kariwa, D Hayasaka, BZ Cui, T Iwasaki, N Lokugamage, LI Ivanov, Volkov, VI, VA Demenev, R Slonova, G Kompanets, T Kushnaryova, T Kurata, K Maeda, K Araki, T Mizutani, K Yoshimatsu, J Arikawa, Takashima, IEMERGING INFECTIOUS DISEASES 8 8 768 - 776 2002年08月 [査読有り][通常論文]
In an epizootiologic survey of 122 rodents captured in Vladivostok, Russia, antibodies positive for hantavirus were found in Apodemus peninsulae (4/70), A. agrarius (1/39), and Clethrionomys rufocanus (1/8). The hantavirus sequences identified in two seropositive A. peninsulae and two patients with hemorrhagic fever with renal syndrome (HFRS) from the Primorye region of Far East Russia were designated as Solovey and Primorye, respectively. The nucleotide sequences of the Solovey, Primorye, and Amur (obtained through GenBank) sequences were closely related (>92% identity). Solovey and Primorye sequences shared 84% nucleotide identity with the prototype Hantaan 76-118. Phylogenetic analysis also indicated a close relationship between Solovey, Primorye, Amur, and other viruses identified in Russia, China, and Korea. Our findings suggest that the Korean field mouse (A. peninsulae) is the reservoir for a hantavirus that causes HFRS over a vast area of east Asia, including Far East Russia. - Kumari Lokugamage, Hiroaki Kariwa, Daisuke Hayasaka, Bai Zhong Cui, Takuya Iwasaki, Nandadeva Lokugamage, Leonid I. Ivanov, Vladimir I. Volkov, Vladimir A. Demenev, Raisa Slonova, Galina Kompanets, Tatyana Kushnaryova, Takeshi Kurata, Kenji Maeda, Koichi Araki, Tetsuya Mizutani, Kumiko Yoshimatsu, Jiro Arikawa, Ikuo TakashimaEmerging Infections Diseases 8 8 768 - 776 2002年 [査読無し][通常論文]
In an epizootiologic survey of 122 rodents captured in Vladivostok, Russia, antibodies positive for hantavirus were found in Apodemus peninsulae (4/70), A. agrarius (1/39), and Clethrionomys rufocanus (1/8). The hantavirus sequences identified in two seropositive A. peninsulae and two patients with hemorrhagic fever with renal syndrome (HFRS) from the Primorye region of Far East Russia were designated as Solovey and Primorye, respectively. The nucleotide sequences of the Solovey, Primorye, and Amur (obtained through GenBank) sequences were closely related (> 92% identity). Solovey and Primorye sequences shared 84% nucleotide identity with the prototype Hantaan 76-118. Phylogenetic analysis also indicated a close relationship between Solovey, Primorye, Amur, and other viruses identified in Russia, China, and Korea. Our findings suggest that the Korean field mouse (A. peninsulae) is the reservoir for a hantavirus that causes HFRS over a vast area of east Asia, including Far East Russia. - Epizootiological survey of hantavirus among rodent species in Ningxia Hui Autonomous Province, ChinaH Kariwa, CB Zhong, K Araki, K Yoshimatsu, K Lokugamage, N Lokugamage, ME Murphy, T Mizutani, J Arikawa, H Fukushima, H Xiong, JHC Chen, Takashima, IJAPANESE JOURNAL OF VETERINARY RESEARCH 49 2 105 - 114 2001年08月 [査読無し][通常論文]
Hantaviral antibodies were detected in the sera from Apodemus (A.) agrarius and A. peninsulae captured in Ningxia province, China by several different serological diagnostic methods. A total of 409 sera from rodent and insectivore species were collected in 1999 and examined by indirect immunofluorescent antibody assay (IFA). Among them, 19 of 191 (9.9%) sera of A. agrarius and 1 of 13 (7.7%) sera of A. peninsulae were positive for hantaviral antibodies. The other species (Rattus norvegicus, Mus musculus, Cricetulus triton, and Sorex cylindricauda) were negative. The reaction pattern of positive serum was characterized as scattered and granular virus antigens in the cytoplasm of hantavirus infected Vero E6 cells. Some of the A. agrarius sera positive for hantavirus were further examined by Western blotting (WB), enzyme-linked immunosorbent assay (ELISA), and the focus reduction neutralization test (FRNT). By WB, positive sera showed the same specific reaction pattern of baculovirus-expressed recombinant hantaviral nucleocapsid protein, as shown in hantavirus-immune serum. By ELISA, IFA-positive sera showed significantly higher optical densities (around 1.0) than the negative A. agrarius sera. Hantaan type hantavirus was neutralized with the positive sera. These results suggest that A. agrarius have hantavirus infection and may play a role as a reservoir animal for hantavirus in Ningxia Hui Autonomous Province, China. - MURPHY M E, KARIWA H, MIZUTANI T, TANABE H, YOSHIMATSU K, ARIKAWA J, TAKASHIMA IJournal of Veterinary Medical Science 63 6 637 - 645 2001年06月 [査読無し][通常論文]
Mechanisms of anti-hantaviral activities of bovine lactoferrin (LF) and ribavirin (Rbv) were investigated. Hantavirus focus formation at 48 hr was 1546 of the control in cells treated with 400 mug/ml LF for 1 hr at 37 degreesC prior to viral infection. Post infection treatment with 100 mug/ml Rbv also inhibited the focus formation to 2.5% of the control. Combined LF pre- and Rbv post-infection treatment completely inhibited focus formation. Viral glycoprotein (G2) and nucleocapsid protein (NP) syntheses were delayed in LF pre treated cells up to 24 hr post infection (hpi) but became comparable to the control by 48 hpi. Further, LF inhibited viral shedding at 24 hpi but did not inhibit shedding after 48 hpi. However, Rbv was able to inhibit synthesis of viral proteins, (+) and (-) strand RNAs also inhibited viral shedding after 24 hr. These results suggest that LF inhibits viral adsorption to cells, while Rbv inhibits viral RNA synthesis. For in vivo trials of LF and Rbv, LF pre- and Rbv post-treatment were evaluated in suckling mice infected with hantavirus, of which 7% survived. LF concentrations of 40 and 160 mg/cg administered prior to viral challenge improved survival rates to 15% and 70%, respectively for single administration and 85% and 94%. respectively, for double administration. Rbv concentrations of 25 and 50 mg/kg gave survival rates of 68% and 81%, respectively. This suggests that both LF and Rbv are efficacious in hantavirus infection in vivo. - K. Araki, K. Yoshimatsu, M. Ogino, H. Ebihara, A. Lundkvist, H. Kariwa, I. Takashima, J. ArikawaJournal of Clinical Microbiology 39 7 2397 - 2404 2001年 [査読無し][通常論文]
Truncated recombinant nucleocapsid proteins (rNPs) of Hantaan virus (HTNV), Seoul virus (SEOV), and Dobrava virus (DOBV) were expressed by a baculovirus system. The truncated rNPs, which lacked 49 (rNP50) or 154 (rNP155) N-terminal amino acids of the NPs of HTNV, SEOV, and DOBV, were able to differentiate HTNV-, SEOV-, and DOBV-specific immune sera. Recombinant NP50s retained higher reactivities than rNP155s and were proven useful for enzyme-linked immunosorbent assay (ELISA). The ELISAs based on the rNP50s of HTNV, SEOV, and DOBV successfully differentiated three groups of patient sera, previously defined by neutralization tests: 17 with HTNV infection, 12 with SEOV infection, and 20 with DOBV infection. The entire rNP of Puumala virus (PUUV) distinguished PUUV infection from the other types of hantavirus infection. Serotyping with these rNP50s can be recommended as a rapid and efficient system for hantavirus diagnosis. - Jin, H.K., Yoshimatsu, K., Takada, A., Ogino, M., Asano, A., Arikawa, J., Watanabe, T.Archives of Virology 146 1 2001年 [査読無し][通常論文]
- H Ebihara, K Yoshimatsu, M Ogino, K Araki, Y Ami, H Kariwa, Takashima, I, DX Li, J ArikawaJournal of Virology 74 19 9245 - 9255 2000年10月 [査読無し][通常論文]
Two Hantaan virus strains, clone 1 (cl-1), which is virulent in newborn mice, and its attenuated mutant (mul1E10), were used to examine the pathogenesis of Hantaan virus infection in a mouse model and identify virus factors relating to virulence. After subcutaneous inoculation of newborn BALB/c mice, cl-1 caused fatal disease with high viral multiplication in peripheral organs, but mul1E10 produced nonfatal infection with a low level of virus multiplication. Intracerebral inoculation of either strain caused fatal disease. Histopathological changes in the dead animals were prominent in the brain, indicating that the brain is the target organ and produces the fatal outcome. These results indicate that mul1E10 has a generally less virulent phenotype, and because of decreased multiplication in peripheral tissues, neuroinvasiveness is also decreased. An experiment with genetic reassortant viruses showed that in newborn mice the M segment is the most related to virulence and the L segment is partly related. Sequence comparison detected a single deduced amino acid change (cl-l Ile to mul1E10 Thr) at amino acid number 515 in glycoprotein G1. One nucleotide change, but no amino acid substitution, was observed in the noncoding region of the L segment. In mouse brain microvascular endothelial cells in vitro, viruses possessing a cl-1-derived M segment grew more rapidly than viruses containing a mul1E10 derived M segment. These results suggest that the single amino acid change in the glycoprotein alters peripheral growth, which affects invasion of the central nervous system in mice. - MURPHY M E, KARIWA H, MIZUTANI T, YOSHIMATSU K, ARIKAWA J, TAKASHIMA IArchives of Virology 145 8 1571 - 1582 2000年 [査読無し][通常論文]
Bovine lactoferrin (LF) and ribavirin (Rbv) were tested as antiviral agents against Seoul type hantavirus (SR-11 strain) in vitro. Hantaviral foci number in Vero E6 cells infected with SR-11 was reduced with LF treatment by 5 days post infection to obtain a 50% effective dose (ED50) Of 2500 mu g/ml, while pretreatment with LF was highly efficacious having an ED50 of 39 mu g/ml. Conversely, 1 h pretreatment with Rbv revealed no inhibition of viral focus formation but could significantly reduce the number of viral foci (ED50: 10 mu g/ml) when used from the time of viral infection. One hour pre-treatment of the cell monolayer with LF and subsequent addition of Rbv revealed a synergistic anti-hantaviral effect against SR-11, <20 FFU/ml as compared to 10(5) foci/ml in the control. One hour treatment of SR-11 with LF prior to cell inoculation gave an ED50 of 312.5 mu g/ml. Whereas, washing the LF-pretreated cell monolayer with PBS demonstrated minimal focus reduction, suggesting LF lightly adheres to cells. These results indicate that LF has anti-hantaviral activity in vitro and inhibition of virus adsorption to cells which play an important role in revealing the anti-hantaviral activity of LF. This paper reports for the first time the anti-hantaviral effect of LF. - Hua Wang, Kumiko Yoshimatsu, Hideki Ebihara, Michiko Ogino, Koichi Araki, Hiroaki Kariwa, Zhaoxiao Wang, Zhaozhuang Luo, Dexin Li, Changshou Hang, Jiro ArikawaVirology 278 2 332 - 345 2000年 [査読無し][通常論文]
The antigenic and genetic properties of 46 hantaviruses from China, 13 from patients, 23 from rodents, and 10 from unknown hosts, were compared with those of other hantaviruses. The viruses were classified as either Hantaan (HTN) or Seoul (SEO) viruses. A phylogenetic analysis of the partial M (300 bp) and S (around 485 bp) genomes of HTN viruses identified nine distinct genetic subtypes, one consisting of isolates from Korea. The SEO viruses were divided into five genetic subtypes, although they had less variability than the HTN subtypes. There was a correlation between the subtype and province of origin for four subtypes of HTN viruses, confirming geographical clustering. Hantaan virus NC167 isolated from Niviventer confucianus and SEO virus Gou3 isolated from Rattus rattus were the basal clades in each virus. The phylogenetic trees constructed from the entire S and M segments suggested that NC167 was introduced to N. confucianus in a host-switching event. The reactivity of a panel of 35 monoclonal antibodies was almost exactly the same in NC167 and a representative HTN virus and in Gou3 and a representative SEO virus. However, there was a one-way cross-neutralization between them. These results confirm the varied nature of Murinae-associated hantaviruses in China. © 2000 Academic Press. - H Kariwa, K Yoshimatsu, K Araki, K Chayama, H Kumada, M Ogino, H Ebihara, ME Murphy, T Mizutani, Takashima, I, J ArikawaMicrobiology and Immunology 44 5 357 - 362 2000年 [査読無し][通常論文]
Hantaviral antibodies were detected in the sera from patients with hepatic disease of unknown etiology in Japan by several different serological diagnostic methods. A total of 105 sera from diseased patients which were negative to A-G hepatitis virus infections in the Tokyo area were tested. Among them, 3 out of 73 sera from patients with chronic hepatic disease were positive to hantaviral antibody by enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescent antibody assay (IFA) and Western blot analysis (WB), Neutralizing antibody titers of the 3 sera to Seoul virus (SEO) were 4 to 8 times higher than those to Hantaan virus (HTN), However, all of the 32 sera from patients with acute hepatitis were negative for hantaviral antibody, Among the 60 patients with chronic hepatitis in Hokkaido which were serologically negative to B and C hepatitis virus infection, one was positive for hantaviral antibody by ELISA and WE. In contrast, the sera from healthy adults in Japan, 550 from the Honshu and Kyushu regions, and 1,000 from the Hokkaido region, were negative for hantavirus antibody. These results show that hantaviral antibodies are more frequently detected in patients with hepatic disease than in healthy adults. However, the observation that no positive sera were detected from patients with acute hepatitis implies that hantavirus might not be directly related to hepatitis. - 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 16 33 - 33 公益社団法人 日本実験動物学会 2000年
- K Tsujimura, T Mizutani, H Kariwa, K Yoshimatsu, M Ogino, Y Morii, H Inagaki, J Arikawa, Takashima, IJournal of Veterinary Medical Science 61 2 113 - 117 1999年02月 [査読無し][通常論文]
For a serological diagnostic test for Borna disease (BD), we developed a capture ELISA with specificity and sensitivity based on detection of antibodies against BD virus (BDV) p40 protein. Using our capture ELISA system, the antibody response of rats inoculated intracerebrally with BDV at 4 weeks after birth showed a sharp increase from 1 to 4 weeks postinoculation (p.i.) and a steady level after 5 weeks p.i. To investigate prevalence of BDV infection among wild rats, we examined sera of Rattus norvegicus in Kami-iso town, Oshima district, Hokkaido, suggesting that rats in this area had not been infected by BDV. - H Kariwa, K Yoshimatsu, J Sawabe, E Yokota, J Arikawa, Takashima, I, H Fukushima, A Lundkvist, FN Shubin, LM Isachkova, RA Slonova, GN Leonova, N HashimotoVirus Research 59 2 219 - 228 1999年02月 [査読無し][通常論文]
Seroepizootiologic surveys among wild rodents were carried out in Japan and Far East Russia in 1995 and 1996. Seropositive animals were only identified in Clethrionomys rufocanus (23/134) in Hokkaido, Japan. On the other hand, seropositives were identified in C. rufocanus (1/8), Apodemus argrarius (2/66), Apodemus spp. (2/26) and Microtus fortis (3/22) in Vladivostok, Far East Russia. Total RNA was isolated from lungs of seropositive animals and the S genome segments were amplified by PCR, cloned and sequenced. The S and M genomes of hantavirus, derived from Japanese C. rufocanus (Tobetsu genotype), were most closely related with Puumala viruses (76-79% nucleotide and 95% amino acid identities for S genome, 70-78% nucleotide and 87-92% amino acid identities for M genome). The recombinant nucleocapsid protein of Tobetsu genotype was antigenically quite similar with that of Sotkamo. These suggest that the virus endemic in Japanese C. rufocanus belongs to Puumala virus. Phylogenetic analysis indicates that the genotype forms a distinct lineage within Puumala viruses. Partial S segment (1-1251 nt), derived from seropositive M. fortis in Vladivostok, was sequenced and analyzed. The S genome segment, which was designated Vladivostok genotype, was most closely related with Khabarovsk virus (79% nucleotide and 90% amino acid identities) which was isolated from M. fortis. (C) 1999 Elsevier Science B.V. All rights reserved. - Koji Tsujimura, Tetsuya Mizutani, Hiroaki Kariwa, Kumiko Yoshimatsu, Michiko Ogino, Yuko Morii, Hisae Inagaki, Jiro Arikawa, Ikuo TakashimaJournal of Veterinary Medical Science 61 2 113 - 117 1999年02月 [査読有り][通常論文]
For a serological diagnostic test for Borna disease (BD), we developed a capture ELISA with specificity and sensitivity based on detection of antibodies against BD virus (BDV) p40 protein. Using our capture ELISA system, the antibody response of rats inoculated intracerebrally with BDV at 4 weeks after birth showed a sharp increase from 1 to 4 weeks postinoculation (p.i.) and a steady level after 5 weeks p.i. To investigate prevalence of BDV infection among wild rats, we examined sera of Rattus norvegicus in Kami-iso town, Oshima district, Hokkaido, suggesting that rats in this area had not been infected by BDV. - Hantavirus infection of SD rats reared in domestic laboratory animal facilities.（共著）Korean Journal of the laboratory Animal Science 15(1),49-52 1999年 [査読無し][通常論文]
- M Ogino, K Yoshimatsu, H Ebihara, J ArikawaArchives of Virology 144 9 1765 - 1777 1999年 [査読無し][通常論文]
N-acetylgalactosamine (GalNAc)-specific lectins, Dolichos biflorus agglutinin (DBA), and soybean agglutinin (SBA), enhanced Hantaan (HTN) virus infections in Vero E6 and P388D1 cells. Treatment of Vero E6 cells with the lectins either before or during, but not after, virus inoculation resulted in lectin-mediated enhancement of infection (LME), indicating that GalNAc-specific lectin affects an early stage of the infection. Lectin blot and FAGS analysis showed that the ability of HTN virus envelope glycoproteins and cell surface molecules to bind DBA and SEA was essential for LME. GalNAc clearly inhibited LME, indicating that the lectins bind with their specific carbohydrate-binding site. These results suggest that a lectin cross-link between the virus and the cell surface is the most plausible mechanism for inducing infection enhancement. - M Morii, K Yoshimatsu, J Arikawa, GZ Zhou, H Kariwa, Takashima, IJournal of clinical Microbiology 36 9 2514 - 2521 1998年09月 [査読無し][通常論文]
Hantaan virus (HTN) and Seoul virus (SEO) are members of the genus Hantavirus in the family Bunyaviridae and are causative agents of hemorrhagic fever with renal syndrome. The complete and truncated nucleocapsid proteins (NP) of HTN and SEO were expressed by a recombinant baculovirus system, Antigenic characterization of the NP using monoclonal antibodies (Mabs)indicated that the binding sites For the serotype-specific MAbs were located between amino acids (aa) 155 and 429, a Western blot assay indicated that the serotype-specific epitopes were conformation dependent. An indirect immunofluorescence antibody (IFA) assay with the truncated NP (aa 155 to 429) was able to distinguish convalescent-phase sera from HTN and SEO patients. However, the antibody liters with the truncated NP were lower than those with the whole NP, The truncated NP of SEO (aa 155 to 429) could be used as an enzyme-linked immunosorbent assay (ELISA) antigen, but the truncated NP from HTN lost its reactivity when used for ELISA. The LFA assay using baculovirus-expressed truncated NP as an antigen is a rapid, simple, and safe test for distinguishing between HTN and SEO infections by serotype. - M Ogino, K Yoshimatsu, K Tsujimura, T Mizutani, J Arikawa, Takashima, IJournal of Veterinary Medical Sciense 60 4 531 - 534 1998年04月 [査読無し][通常論文]
We produced two recombinant Borna disease virus (BDV) proteins, p40 and p24, by using a baculovirus vector as a diagnostic antigen; Antigenicities of these recombinant proteins were evaluated by immune rabbit sera; Recombinant p40 was a more sensitive antigen than p24 for the detection of antibodies in infected rats. Rats inoculated with BDV within 24 hi after birth showed higher detection rates of viral RNA and viral proteins from the brain than rats inoculated at 4 weeks-old. Depending on the age of infection and the time postinfection, the detection of BDV RNA, protein, or anti-BDV antibody did not always correlate in individuals, We suggest both serological and molecular biological methods are needed in the diagnosis of BDV infection. - YC Yoo, K Yoshimatsu, Y Koike, R Hatsuse, K Yamanishi, O Tanishita, J Arikawa, Azuma, IVaccine 16 2-3 216 - 224 1998年01月 [査読無し][通常論文]
The adjuvant effect of two lipophilic derivatives of muramyl dipeptide (MDP), B30-MDP and MDP-Lys(L18), on the ability of an inactivated vaccine of B-I virus (B-l vaccine) to induce immune response against Hantavirus causing hemorrhagic fever with renal syndrome (HFRS) was examined, When mice were immunized subcutaneously (s.c.) twice at 2-week intervals with B-I vaccine admixed with or without 100 mu g mouse(-1) of B30-MDP (B-1/B30-MDP) or MDP-Lys(L18) [B-1/MDP-Lys(L18)], mice immunized with B-1/B30-MDP as well as B-1/MDP-Lys(L18) showed Significantly higher indirect fluorescent antibody (IFA) titers against HFRS virus than mice immunized with B-I vaccine alone, Both mice treated with B-1/B30-MDP and B-1/MDP-Lys(L18) also exhibited significantly higher neutralizing antibody titers against HFRS virus than mice immunized with B-I ,vaccine alone during 3-9 weeks after-the primary immunization. The evaluation of antibody-producing cells by enzyme-linked immunospot (ELISPOT) assay on week 4 revealed that both MDP derivatives,es enhanced the number of HFRS virus-specific IgG1 and IgM antibody-producing cells. Furthermore, mice treated with B-1/B30-MDP as well as B-1/MDP-Lys(L18) showed a higher level of Th-2 type cytokines, IL-4 and IL-6, in sera than mice treated with B-l alone. In an in-vitro analysis of T lymphocyte proliferation to baculovirus-expressed recombinant nucleocapsid pi protein (rNP) of Hantaan 76-118 sa strain, the splenocytes of mice treated with B-1/B30-MDP and B-1/MDP-Lys(L18) on week 4 showed a Significantly higher proliferating activity than those treated with B-1 vaccine alone, In addition when mice were immunized once with B-l vaccine admixed with or without B30-MDP and MDP-Lys(L18) and followed by intrafootpad (i.f.) injection of B-I vaccine on day 7, mice immunized with B-1/B30-MDP and B-1/MDP-Lys(LIS) induced a higher delayed-type hypersensitivity (DTH) reaction than mice immunized with B-I vaccine alone, These results suggest that B30-MDP and MDP-Lys(L18) are useful immunoadjuvants to enhance the ability of inactivated B-l vaccine to induce a humoral and cellular response to HFRS virus, (C) 1997 Elsevier Science Ltd, All rights reserved. - M Kikuchi, K Yoshimatsu, J Arikawa, R Yoshida, YC Yoo, Y Isegawa, K Yamanishi, S Tono-oka, Azuma, IArchives of Virology 143 1 73 - 83 1998年 [査読無し][通常論文]
Neutralizing monoclonal antibody (MAb) escape mutants of Hantaan virus were generated using MAbs to envelope protein G1 (16D2) and G2 (11E10). The mutant viruses (mu16D2 and mu11E10), lacked reactivity only to the selecting MAb, or a MAb belonging to the same antigenic site. Both mutants had a single amino acid (a.a.) substitution. The a.a. substitution, found in mu16D2, was different from that found in another mutant selected with the same MAb (16D2). Although MAb 11E10 immunoprecipitated G2 protein, a deduced a.a. substitution was located in the G1 region. These results suggest that antigenic sites defined by neutralizing MAbs are composed of discontinuous epitopes over the G1 and G2 proteins. Mutant 11E10 showed a significant decrease in virulence in suckling mice. A virulence revertant of mu11E10, selected through passages in suckling mice brain, showed exactly the same deduced a.a. sequence as mu11E10 and still was not neutralized by MAb 11E10. Since mutant 16D2 was virulent for suckling mice, neutralization related epitopes found with MAbs 11E10 and 16D2 were independent of pathogenicity in BALB/c mice. - H Kariwa, M Fujiki, K Yoshimatsu, J Arikawa, Takashima, I, N HashimotoArchives of Virology 143 1 15 - 24 1998年 [査読無し][通常論文]
To understand the mode of transmission of Seoul type hantavirus in Wistar rats, we examined the shedding of the virus and antibody production in infected rats. When 1-day-old rats were inoculated with the KI-83-262 strain of Seoul virus, S segment of the viral genome was detected in lungs, clots, urine, saliva, submaxillary glands, rectums, and kidneys by nested reverse transcriptase PCR. On the other hand, when 8-week-old rats were infected with the virus, viral genome was detected only in the lungs and rectum. In newborn rats intranasally administered urine from infected newborn rats, four of six rats shed the virus into their urine. In addition, three of eight rats kept in the same cage with infected animals also shed the virus into urine. Moreover, the virus genome was detected in the urine of urban rats (Rattus norvegicus) in an enzootic focus. These findings suggest that the urine containing virus from infected rats is an actual source of the Seoul virus infection. - H Kariwa, M Fujiki, K Yoshimatsu, J Arikawa, Takashima, I, N HashimotoARCHIVES OF VIROLOGY 143 2 365 - 374 1998年 [査読有り][通常論文]
To understand the mode of transmission of Seoul type hantavirus in Wistar rats, we examined the shedding of the virus and antibody production in infected rats. When 1-day-old rats were inoculated with KI-83-262 strain of Seoul virus, the S segment of the viral genome was detected in lungs, clots, urine, saliva, submaxillary glands, rectums, and-kidneys by nested reverse transcriptase PCR. On the other hand, when 8-week-old rats were infected with the virus, viral genome was detected only in the lungs and rectum. In uninfected newborn rats intranasally administered urine from infected newborn rats, four of six rats shed the virus into their urine. In addition, three of eight rats kept in a same cage with infected animals also shed the virus into urine. Moreover, the virus genome was detected in the urine of urban rats (Rattus norvegicus) in an enzootic focus. These findings suggest that the urine containing virus from infected rats is an actual source of Seoul virus infection. - K Yoshimatsu, J Arikawa, S Ohbora, C ItakuraJournal of Veterinary Medical Science 59 10 863 - 868 1997年10月 [査読無し][通常論文]
Severe combined immunodeficiency (SCID) mice were inoculated with Hantaan virus strain 76-118 (HTN) or Seoul virus strain SR-11 (SR) of hantaviruses. Susceptibility of SCID mice was compared with those of immunocompetent adult mice, newborn mice and nude mice. SCID mice inoculated with HTN or SR died 32 to 35 days after infection. Unlike newborn mice which also died of hantavirus infection, SCID mice survived longer than newborn mice and showed typical wasting symptoms rather than nervous symptoms. Immunohistochemical staining and virus isolation indicated that both HTN and SR inoculated SCID and SR inoculated nude mice showed systemic infection, but nude mice inoculated with SR survived for longer than 8 weeks after inoculation. Passive transfer of spleen cells from immunocompetent BALB/c mice conferred protection on SCID mice within 2 weeks of HTN infection. Immune mediated pathologic mechanism was examined by transferring the spleen cells to SCID mice inoculated with HTN virus 3 weeks before the cell transfer. The recipient SCID mice showed an increase of serum BUN level coinciding with the appearance of serum antibody to HTN virus, suggesting the immune mediated pathogenicity. - Takashima, I, K Morita, M Chiba, D Hayasaka, T Sato, C Takezawa, A Igarashi, H Kariwa, K Yoshimatsu, J Arikawa, N HashimotoJournal of Clinical Microbiology 35 8 1943 - 1947 1997年08月 [査読無し][通常論文]
A case of tick-borne encephalitis (TEE) has not been reported for many years in Japan, although a serological survey of sera from domestic animals suggested the presence of TEE foci in Hokkaido, the northern island of Japan. Studies were conducted to prove the presence of an endemic focus of TEE virus in Japan by means of serology and virus isolation. In October 1993 in Hokkaido, a severe case of encephalitis in a dairy farmer's nife was diagnosed as TEE. Serological examination of paired serum specimens showed a rise in the neutralization antibody titer to Russian spring summer encephalitis virus. A seroepizootiological survey of dogs showed that the TEE-related virus was prevalent in the area. Three virus isolates were obtained from the blood of sentinel dogs, and antigenic analysis grouped the isolates into TEE-related viruses. Sequence analysis of the envelope protein gene identified one of the isolates as being of the same subtype as the Russian spring summer encephalitis (Far Eastern TEE) virus. The results provide evidence that TEE is endemic in a certain area of Japan. - K Matsuzawa, YC Yoo, A Fukushima, K Yoshimatsu, J Arikawa, Azuma, IVaccine 15 1 85 - 89 1997年01月 [査読無し][通常論文]
We investigated the protection confered by the mucosal administration of recombinant human macrophage colony-stimulating factor (rhM-CSF) against mucosal infection of Sendai virus in mice. In an experimental infection model using Sendai virus, an intranasal (i.n.) administration of rhM-CSF (20 mu g per mouse) 2 days before infection induced significant protection against a lethal infection of this virus. Also, its antiviral activity was dependent upon the dose of rhM-CSF. However; a subcutaneous (s.c.) administration of rhM-CSF with an effective close (20 mu g per mouse) i.n. did not confer protection. In a time course analysis of virus growth in the lungs, mice given rhM-CSF in. significantly inhibited the early period of infection, compared with the untreated mice. Moreover, the level of interferon-gamma (IFN-gamma) in lung wash fluids from the rhM-CSF-treated mice was higher than that of the untreated mice. These results suggested that the mucosal (i.n.), but not the systemic (s.c.) administration of rhM-CSF augments host resistance against mucosal infection with Sendai virus, and that its prophylactic activity is related to growth inhibition of the virus and enhanced IFN-gamma secretion in the lungs. Copyright (C) 1997 Elsevier Science Ltd. - A Fukushima, YC Yoo, K Yoshimatsu, K Matsuzawa, M Tamura, S Tonooka, K Taniguchi, S Urasawa, J Arikawa, Azuma, IVaccine 14 6 485 - 491 1996年04月 [査読無し][通常論文]
To examine the effect of MDP-Lys(L18), a derivative of muramyl dipeptide (MDP), as a mucosal immunoadjuvant, we investigated its activity to augment host resistance against mucosal infections by Sendai virus and rotavirus in mice. lit an experimental infection model that suckling mice (10-day-old) were inoculated perorally (p.o.) with 1.5 x 10(6) p.f.u. mouse(-1) of rotavirus strain SA11, intrarectal (i.r.) as well as p.o., administration of MDP-Lys(L18) (50 mu g mouse(-1)) prior to virus infection markedly reduced rotavirus-induced diarrhea. Furthermore, when MDP-Lys(L18) was administered p.o. (1 mg mouse(-1)), i.r. (300 mu g mouse(-1)) or intranasally (i.n., 100 mu g mouse(-1)) various days before Sendai virus infection (2.6 x 10(4) HAD mouse(-1)), all the mucosal administration of MDP-Lys(L18) significantly protected a lethal infection of Sendai virus, showing a dose-dependent manner. However, the efficacy of MDP-Lys(L18) to induce the prophylactic activity against the viruses somewhat varied according to the administration route and timing. In time course analysis of virus isolation in vivo, the mice administered with MDP-Lys(L18) exhibited a significant reduction of both viruses in the lungs for Sendai virus and in the bowels for rotavirus. These results suggest that MDP-Lys(L18) is a potent mucosal immunoadjuvant to enhance nonspecific host resistance against two mucosal infectious viruses, Sendai virus and rotavirus. Copyright (C) 1996 Elsevier Science Ltd. - K Yoshimatsu, J Arikawa, M Tamura, R Yoshida, A Lundkvist, B Niklasson, H Kariwa, Azuma, IJournal of General Virology 77 695 - 704 1996年04月 [査読無し][通常論文]
We characterized the antigenic sites on the nucleocapsid protein (NP) of Hantaan virus (HTN) using 10 monoclonal antibodies (MAbs). At least seven antigenic sites were revealed by a competitive binding assay and divided into three partially overlapping antigenic regions (I, II and III). Regions I [amino acids (aa) 1-103], II (aa 104-204) and III (aa 205-402) were mapped on NP by examinincr the reactivity of truncated gene products. Those that corresponded to region I reacted with immune mouse serum, indicating that the region contained major linear epitopes as reported with Four corners virus (FCV) and Puumala virus (PUU) NP. At least one MAb to each region inhibited viral growth when they were introduced into cells by scrape-loading. In addition, they conferred protection from a lethal HTN challenge to newborn mice. A PEPSCAN assay localized the epitope of MAb E5/G6 between aa 166-175. Since E5/G6, which had the highest inhibitory effect both in cells and in mice, showed no virus neutralization activity by ordinary neutralization test, this region is suggested to be important for the virus growth after entry into the cells. - H Kariwa, M Kimura, S Yoshizumi, J Arikawa, K Yoshimatsu, Takashima, I, N HashimotoArchives of Virology 141 12 2327 - 2338 1996年 [査読無し][通常論文]
To understand the mode of persistent infection of Seoul virus in rodents, we examined the distribution of the virus genome and antibody production in infected rats. When 1-day-old rats were inoculated with the KI-83-262 strain, the S segment of viral genome was detected in sera, clots, lungs and kidneys from 3 to 184 days post inoculation (d.p.i.) by nested reverse transcriptase PCR. On the other hand, when 7-week-old rats were infected with this virus, viral genome was detected only in the lungs from 3 to 50 d.p.i. The neutralizing antibody titers of rats inoculated at 1-day of age were higher than those of rats inoculated at 7 weeks of age. In both a ge groups, however, the IgG avidity of antibody increased along with the course of infection. We found that urban rats (Rattus norvegicus) infected early in life harbored the virus for more than 6 months. - K Yoshimatsu, J Arikawa, H Li, H Kariwa, N HashimotoJournal of Veterinary Medical Science 58 1 71 - 74 1996年01月 [査読無し][通常論文]
Recombinant Hantaan virus nucleocapsid protein expressed in silkworm larvae was applied as a serological diagnostic antigen in Western blots (WE) of human sera. The sensitivity of this method was similar to that of the IFA test. Hemorrhagic fever with renal syndrome (HFRS) and nephropathia epidemica diagnosed by their cross-reactivity in WE. The specificity of this method was higher than that of IFA test because the background was low. Sera that exhibited high background staining in the IFA test were readily diagnosed with this method. We recommended WE using recombinant Hantaan virus nucleocapsid antigen as a confirmatory procedure for the serodiagnosis of hantavirus. - K Yoshimatsu, J Arikawa, R Yoshida, H Li, YC Yoo, H KariwaLaboratory Animal Science 45 6 641 - 646 1995年12月 [査読無し][通常論文]
The recombinant nucleocapsid protein (rNP) of Hantaan virus was expressed by a baculovirus vector in silkworm hemolymph and was used as an antigen in western blotting (WB). The rNP is expressed in insoluble form in hemolymph; therefore simple washing of the insoluble fraction with phosphate-buffered saline by low-speed centrifugation allowed preparation of purified antigen for WB. The rNP had strain-specific and hantavirus-common epitopes similar to the authentic NP antigen of hantavirus and was stable after transfer to membrane. For detection of antibody in serially obtained sera from experimentally infected rats, WB enabled detection of IgM antibodies 3 days after infection, which was at least 2 days earlier than detection by the indirect immunofluorescent antibody test (IFA). Thus WB had a higher sensitivity than the IFA for detection of hantavirus antibody in the serum of experimentally infected rats. The WB-determined IgG antibody titer was about 10 times higher than that determined by the IFA. No background staining was observed by WB even at a 1:10 dilution of serum. The selected rat sera with strong background staining or confusing staining patterns by LFA, but not focus reduction neutralization test titers, could be interpreted as test-negative because they did not have a specific reaction to virus antigen by WB. Thus the specificity of WB was higher than that of the IFA. Moreover, WB can distinguish specific from nonspecific reactions by the detection of the specific antigen on the WB membrane. Therefore the IFA or enzyme-linked immunosorbent assay followed by WB is recommended for serologic confirmation of hantavirus infection. - YC YOO, K YOSHIMATSU, R HATSUSE, M TAMURA, R YOSHIDA, S TONOOKA, J ARIKAWA, AZUMA, IVaccine 13 14 1300 - 1305 1995年10月 [査読無し][通常論文]
We examined the effect of MDP-Lys(L18), a lipophilic derivative of muramyl dipeptide, on the enhancement of host resistance against virus infection in newborn mice Newborn mice were inoculated with 4 LD(50)/mouse of Hantaan virus strain 76-118 (HTN) one day after birth. Mice given 100 mu g/mouse of MDP-Lys(L18) before infection exhibited significantly higher survival rates than those of non-treated mice. The effect of MDP-Lys(L18) was also restorative when given to the mice 4 or 7 days after infection. The titers of virus isolated from the lungs and spleens 12 days after infection, were about 30-times lower in MDP-Lys(L18)-treated (lung: 1.0 x 10(3) FFU; spleen: 6.8 x 10(1) FFU/mouse), than those of non-treated mice (lung: 3.4 x 10(4) FFU; spleen: 1.9 x 10(3) FFU/mouse). Furthermore the virus was undetectable in the brains of MDP-Lys(L18)-treated mice, whereas viruses were isolated from 3 of 6 non-treated mice MDP-Lys(L18) augmented the number of peripheral leukocytes and splenocytes, as well as mitogenic responses of the cells from bone marrow and spleen of newborn mice. These results suggest that MDP-Lys(L18) enhanced the resistance of newborn mice against HTN virus in a systemic infection model, and that this mechanism is involved in the enhancement of hematopoiesis and responsiveness of immune-related cells to mitogens. - Evidence for the existence of Puumala-related virus Clethrionomys rufocanus in Hokkaido, Japan. （共著）H KARIWA, S YOSHIZUMI, J ARIKAWA, K YOSHIMATSU, K TAKAHASHI, TAKASHIMA, I, N HASHIMOTOAmerican Journal of Tropical Medicine and Hygene 53 3 222 - 227 1995年09月 [査読無し][通常論文]
We conducted field surveys of indigenous rodent species in Hokkaido, Japan from 1980 to 1993. Serum samples were collected from 663 rodents, including Clethrionomys rufocanus, Apodemus speciosus, A. argenteus, and C. rutilus. Antibody to hantavirus was determined by the protein G antibody assay. Positive C. rufocanus were detected in seven of eight collection sites, but no antibody was detected in the remaining rodent species. To reveal the serotype of the circulating virus in C. rufocanus, antibody titers to Hantaan, Seoul, Puumala, and Prospect Hill viruses were compared by means of the focus reduction neutralization test. The titers in positive sera were extremely high to the Sotkamo strain of Puumala virus. Results were confirmed by the reverse transcriptase-polymerase chain reaction, and suggested that Puumala-related viruses are in circulation among C. rufocanus populations in Hokkaido. - M TAMURA, YC YOO, K YOSHIMATSU, R YOSHIDA, T OKA, K OHKUMA, J ARIKAWA, AZUMA, IVaccine 13 1 77 - 82 1995年01月 [査読無し][通常論文]
The ability of two muramyl dipeptide (MDP) derivatives, B30-MDP and MDP-Lys (L18), to enhance the immunogenicity of recombinant hepatitis B surface antigen (rHBsAg) was examined. When mice were immunized intraperitoneally with rHBsAg together with each MDP derivative, the antibody titres were higher than those in mice immunized with alum-adsorbed rHBsAg, which is a commercially available hepatitis B vaccine. When mice were given a subcutaneous or intramuscular injection of rHBsAg and either MDP derivative, the antibody titres were the same as those in mice given alum-adsorbed rHBsAg. These results indicate the usefulness of MDP derivatives as immunoadjuvants for a new-generation vaccine. - H KARIWA, M KAMIMURA, J ARIKAWA, K YOSHIMATSU, TAKASHIMA, I, N HASHIMOTOMicrobiology and Immunology 39 1 35 - 41 1995年 [査読無し][通常論文]
A polymerase chain reaction (PCR) for the detection of hantavirus genome was established and applied to analyze the mode of infection of Hantaan virus in adult ICR mice. The cDNA for the S genome segment of Hantaan virus was reverse-transcribed from the total RNA of organs of the infected mice. The sequence in the S genome segment of Hantaan virus was successfully amplified by reverse transcriptase (RT)-PCR followed by nested PCR. In 5-week-old ICR mice inoculated intraperitoneally with Hantaan virus, strain 76-118 (1.3 x 10(5) FFU/mouse), the virus was detected in clots and lungs from 3 to 10 days post-inoculation (p.i.) by nested PCR and virus-isolation techniques. No virus was detected in any specimens collected on 1 day and after 28 days p.i., and in spleens and brains through the observation period by both methods, The antibody which was measured by indirect immunofluorescence antibody assay (IFA) appeared at 7 days p.i. and the geometric mean titer was elevated to its maximum level of 1 : 203 at 10 days p.i., maintaining the same level until 35 days p.i. These results suggest that adult mice are transiently infected with Hantaan virus. - R YOSHIDA, K SATO, YC YOO, K YOSHIMATSU, M TAMURA, C ISHIHARA, J ARIKAWA, AZUMA, IImmunopharmacology 28 2 153 - 161 1994年09月 [査読無し][通常論文]
DT-5461 enhanced host resistance to Sendai virus infection in mice. Intranasal (i.n.) administration of 200 mu g of DT-5461 per mouse 3 days before infection was the most effective administration route, dose and timing. DT-5461 enhanced the cytotoxicity of murine natural killer (NK) cells. In addition, DT-5461 activated murine peritoneal macrophages, resulted in augmented of cytotoxicity and the induction of tumor necrosis factor-alpha (TNF-alpha). Therefore, these immunomodulating activities presented by DT-5461 caused protection against Sendai virus infection. - AZUMA, I, YC YOO, M TAMURA, R YOSHIDA, K YOSHIMATSU, J ARIKAWA, K YAMANISHIIMMUNOTHERAPY OF INFECTIONS 15 191 - 203 1994年 [査読有り][通常論文]
- K YOSHIMATSU, J ARIKAWA, H KARIWAJournal of Veterinary Medical Science 55 6 1047 - 1050 1993年12月 [査読無し][通常論文]
Recombinant baculovirus-infected insect cells which expressed recombinant protein, analogous to the nucleocapsid protein (NP) of Seoul type hantavirus, strain SR-11 (rNP-SR-Sf9) were applied to the indirect immunofluorescent antibody (LFA) test. The rNP-SR-Sf9 reacted with anti NP MAb clones which recognized strain specific or hantavirus common epitopes in the IFA test. The recombinant antigen was detected by antibodies to 3 serotypes of hantaviruses (Hantaan 76-118, SR-11, Puumala). Antibody titers of a group of experimentally infected mouse and urban rat sera using rNP-SR-Sf9 and strain SR-11-infected Vero cell antigens were mutual correlative. These results indicated that the rNP-SR-Sf9 were an effective and safety substitute for Vero E6 cells in the IFA test for serosurveys of hantavirus infection. - Y YUYAMA, K YOSHIMATSU, E ONO, M SAITO, M NAIKIJournal of Biochemistry (Tokyo) 113 4 488 - 492 1993年04月 [査読無し][通常論文]
Enterotoxigenic Escherichia coli possessing K99 fimbriae (E. coli K99) causes diarrhea in piglets of less than 1 week old. The first stage of the bacterial infection is adhesion by the fimbriae on the small intestinal mucosa and the adhesion is followed by colony formation. K99 fimbriae bind specifically to N-glycolylneuraminyl-lactosyl-ceramide, GM3(NeuGc) [Ono, E. et al. (1989) Infect. Immun. 57, 907-911]. We examined the postnatal change of the content and the molecular species of GM3(NeuGc) in the small intestinal mucosa of 0- to 14-day-old piglets and adult pigs. GM3(NeuGc) was a major ganglioside of piglet intestinal mucosa. GM3(NeuGc) content was maximal at birth and gradually decreased to 1/16 in adult animals (5 months old). The ceramide moiety of piglet intestinal GM3(NeuGc) was characterized by the presence of 2-hydroxylated palmitic acid. I-125-labeled bacteria strongly bound to GM3(NeuGc) containing 2-hydroxylated palmitic acid and phytosphingosine compared with GM3(NeuGc) containing any other ceramide moiety. The time when this particular GM3(NeuGc) appears coincides with the time that the infection occurs, and it may explain the susceptibility of newborn piglets to E. coli K99 infection. - C ISHIHARA, K YOSHIMATSU, M TSUJI, J ARIKAWA, SAIKI, I, S TOKURA, AZUMA, IVaccine 11 6 670 - 674 1993年04月 [査読無し][通常論文]
Sulfated chitin derivatives, selected for their low toxicity and high inhibitory activity of melanoma metastasis, were examined for anti-viral activity against Friend murine leukaemia, herpes simplex type-1 (HSV) and Sendai viruses. Carboxymethyl chitin with a 7.66% degree of sulfation (SCM-chitin III) showed a significant inhibition of Friend murine leukaemia helper virus (F-MuLV) and HSV, but not of Sendai virus growth in vitro. Sulfated N-deacetylated chitin had a significant but weak activity against F-MuLV and HSV infections. Carboxymethyl chitin showed no effect on these infections in vitro. SCM-chitin III also exhibited anti-viral activity in vivo by suppressing the splenomegaly which was caused by prior infection of mice with FV, a complex of F-MuLV and spleen focus-forming virus. - K YOSHIMATSU, YC YOO, R YOSHIDA, C ISHIHARA, AZUMA, I, J ARIKAWAArchives of Virology 130 3-4 365 - 376 1993年 [査読無し][通常論文]
Recombinant Hantaan virus nucleocapsid protein (rNP) and recombinant envelope (rEnv) proteins were prepared using a baculovirus expression system to examine the role of Hantaan virus structural proteins in protective immunity. Passive transfer of spleen cells from mice immunized with rNP conferred partial protection or prolongation of time to death from fatal Hantaan virus infection in suckling mice which were challenged with Hantaan virus at 40 LD50 (survival rate: 43%) or 4 LD50 (survival rate: 43%). The T cell-enriched fraction protected one mouse from lethal infection but the B cell-enriched fraction had no such effect on fatal HTN infection. The protective effects of the antibody against HTN challenge were examined by passive immunization. The monoclonal antibody ECO 2 directed to NP also conferred partial survival and significant difference in time to death. Although rEnv antigen failed to induce neutralizing antibody, both immune spleen cells and immune serum to rEnv conferred partial protection upon suckling mice. These results indicate that both nucleocapsid and envelope proteins of Hantaan virus were responsible for induction of cell mediated protective immunity. Vero E 6 cells infected with Hantaan virus expressed envelope protein on the surface, as determined by flow cytometry. However, there was only negligible expression of nucleocapsid protein. - YC YOO, K YOSHIMATSU, R YOSHIDA, M TAMURA, AZUMA, I, J ARIKAWAMicrobiology and Immunology 37 7 557 - 562 1993年 [査読無し][通常論文]
Virulence of hantavirus strain of SR-11 Seoul virus and Hantaan 76-118 (HTN) of Hantaan virus were compared. Infections of both strains were lethal in newborn mice. However, inoculum required to cause lethal infection was about 4,000 times higher for strain HTN (1.65 x 10(3) PFU/mouse/LD50) than for strain SR-11 (0.36 PFU). Thus, both strains were considered pathogenic to newborn mice but they possessed different levels of virulence. The assay system used for these strains in newborn mice proved to be useful in the study of hantavirus vilurence. Growth curves of the two strains in CV-7 cell cultures were compared. Strain SR-11 was shown to have higher activity of virus replication and virus release into the culture fluids than strain HTN. The possibility of a relationship between replication activity and high levels of virulence in mice was suggested. - JS YAO, J ARIKAWA, H KARIWA, K YOSHIMATSU, TAKASHIMA, I, N HASHIMOTOJAPANESE JOURNAL OF VETERINARY RESEARCH 40 2-3 87 - 97 1992年09月 [査読有り][通常論文]
The effect of neutralizing monoclonal antibodies (N MAbs) on Hantaan virus infection of macrophages was investigated using P388D1 cells, a murine macrophage cell line. MAbs to the G1 protein (G1b) and the G2 protein (G2a and G2c) neutralized viral infectivity in P388D1 cells. N MAbs to G1b showed much higher neutralizing potency than those to G2a and G2c. With each N MAbs, two distinct effects were observed: neutralization of viral infectivity occurring at high concentrations and enhancement of that at low concentrations. Non-neutralizing MAbs, on the other hand, showed only enhancement of viral infectivity even at high concentrations without any inhibitory effects. The Fab fragments of N MAbs showed neither neutralizing nor enhancing activities. However, when the virus-Fab complexes were reacted with the anti-Fab antibodies, both neutralization and enhancement of viral infectivity were restored depending on the dose of Fab fragments. These results indicate that Hantaan virus infection of P388D1 cells is mediated by the Fc portion of the antibodies and neutralization is dependent on the concentration of N antibodies bound bivalently to the neutralization site on the virion. - A WATANABE, M MORIMATSU, K YOSHIMATSU, S YAMAMOTO, A TERAO, K TSUKAZAKI, M SAITO, M NAIKIJOURNAL OF SMALL ANIMAL PRACTICE 33 2 71 - 77 1992年02月 [査読有り][通常論文]
C-reactive protein (CRP) was isolated from sera from healthy cats by calcium-dependent affinity chromatography on phosphorylcholine derivatives of bovine albumin-coupled Toyopearl, followed by an anion-exchange chromatography using DEAE cellulose. It was identified as CRP by its immunochemical cross reactivity with human CRP. The molecular weight of cat CRP was approximately 100 kilodaltons (kDa) and composed of two glycosylated subunits (23 kDa) and three non-glycosylated subunits (20 kDa) with non-covalent association. Under electron microscopic examination, cat CRP had a pentameric disc-like configuration which is characteristic of CRP. Immunoelectrophoresis and isoelectric focusing showed that cat CRP is an acidic alpha-1-globulin (pI 4.1 to 4.3). Serum concentrations of CRP in cats and kittens were measured by single radial immunodiffusion. In 12 healthy cats from various sources, values ranged from 38 to 168-mu-g/ml. In kittens, serum CRP levels also showed a wide distribution, 81 per cent of them were less than 40-mu-g/ml. - Wakamiya, N., Okuno, Y., Sasao, F., Ueda, S., Yoshimatsu, K., Naiki, M., Kurimura, T.Biochemical and Biophysical Research Communications 187 3 1992年
- AZUMA, I, C ISHIHARA, J IIDA, YC YOO, K YOSHIMATSU, J ARIKAWAMICROBIAL INFECTIONS 319 253 - 263 1992年 [査読有り][通常論文]
- JS YAO, H KARIWA, TAKASHIMA, I, K YOSHIMATSU, J ARIKAWA, N HASHIMOTOARCHIVES OF VIROLOGY 122 1-2 107 - 118 1992年 [査読有り][通常論文]
Antibody-dependent enhancement (ADE) of hantavirus infections (strains Hantaan 76-118 and SR-11) was studied using macrophage-like cell lines (J774.1, P388D 1, and U937). Significantly higher virus titers (1,000 to 4,000 FFU/ml) were obtained by pretreatment of the virus with immune serum as compared to normal serum (< 20 FFU/ml). Monoclonal antibodies (MAbs) to strain Hantaan 76-118 were employed to determine the antigenic determinants responsible for the ADE activity. ADE of the infection occurred with MAbs to both G1 and G2 envelope glycoproteins, but not with MAbs to nucleocapsid protein. Antigenic determinants related to haemagglutination or virus neutralization were found to cause ADE of the infection. - J ARIKAWA, JS YAO, K YOSHIMATSU, TAKASHIMA, I, N HASHIMOTOARCHIVES OF VIROLOGY 126 1-4 271 - 281 1992年 [査読有り][通常論文]
To investigate the role of Hantaan virus envelope glycoprotein in infection, a panel of monoclonal antibodies (MAbs) was examined in vitro with several serological tests and in vivo by passive transfer experiments in mice. An antigenic site, specific for the inhibition of infected cell focus was detected with the focus inhibition neutralization test (FINT), in addition to the neutralization related antigenic sites, which were revealed by the ordinary focus reduction neutralization test (FRNT). Suckling mice were given the MAbs by passive transfer followed by lethal Hantaan virus challenge. All neutralizing MAbs detected by either FRNT or FINT protected all mice from lethal infection, confirming the importance of the antigenic sites as a protective antigen. Mice given non-neutralizing MAbs by passive transfer, however, began to die earlier than the control group; mean time to death (18.2 +/- 2.1 to 21.5 +/- 2.8 days) being significantly shorter than that of the control group (25.8 +/- 1.8, p < 0.01, Mann-Whitney, U probability test). Virus titers in brains of mice which died early, were about 10 times higher than those of control mice. These results indicated the early death phenomenon of mice which was mediated by the anti-virus antibody. - M MORIMATSU, A WATANABE, K YOSHIMATSU, T FUJINAGA, M OKUBO, M NAIKIJOURNAL OF DAIRY RESEARCH 58 3 257 - 261 1991年08月 [査読有り][通常論文]
C-reactive protein (CRP) and serum amyloid P component (SAP), which are known to increase in sera from humans and many other animals with acute inflammation caused by infection, toxic drug administration or injury, were previously purified from bovine serum. These serum levels were determined by enzyme-linked immunosorbent assay (ELISA) using specific antiserum to bovine CRP or SAP which was prepared by immunizing rabbits and goats with each purified protein. Among 68 healthy Holstein cows, 45 non-lactating cows had levels of CRP and SAP of 20.6 +/- 1.4 and 27.6 +/- 1.3-mu-g/ml respectively; 23 lactating cows had higher levels of CRP and SAP (76.0 +/- 13.6 and 38.3 +/- 5.5-mu-g/ml respectively). In the latter group, there was a high correlation between milk yield and serum CRP levels (P < 0.001). From these observations, it was assumed that lactation might stimulate CRP synthesis rather than SAP synthesis in bovine liver as an acute phase reaction, and that CRP might be called a lactation-associated protein. - ISOLATION AND CHARACTERIZATION OF C-REACTIVE PROTEIN AND SERUM AMYLOID-P COMPONENT FROM BOVINE SERUMM MORIMATSU, H SAKAI, K YOSHIMATSU, O MINOWA, S YAMAMOTO, K YATOMI, T FUJINAGA, M NAIKIJAPANESE JOURNAL OF VETERINARY SCIENCE 51 4 723 - 732 1989年08月 [査読有り][通常論文]
MISC
- 佐藤彰彦, 佐藤彰彦, 佐藤彰彦, 鳥羽晋輔, 鳥羽晋輔, 垰田善之, 日下部伸治, 日下部伸治, 小西慧, 小西慧, 松野啓太, 松野啓太, 吉松組子, 有川二郎, 佐々木道仁, 佐々木道仁, 大場靖子, 大場靖子, 大場靖子, 澤洋文, 澤洋文, 澤洋文 日本ウイルス学会学術集会プログラム・予稿集(Web) 70th 2023年
- 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年
- Yomani D. Sarathkumara, Chandika D. Gamage, Sithumini Lokupathirage, Devinda S. Muthusinghe, Nishantha Nanayakkara, Lishanthe Gunarathne, Kenta Shimizu, Yoshimi Tsuda, Jiro Arikawa, Kumiko Yoshimatsu Viruses 11 (12) 2019年12月11日
- 小泉信夫, 泉谷秀昌, 慕蓉蓉, 岡野祥, 中島千絵, 鈴木定彦, 谷川力, 小松謙之, 吉松組子, 武藤(水谷)麻紀, 武藤(水谷)麻紀, 大西真 レプトスピラ・シンポジウム 53rd 2016年
- イムノクロマトグラフィー法によるハンタウイルスの迅速診断法の開発小山 芽以, 吉松 組子, 好井 健太朗, 有川 二郎, 苅和 宏明 日本獣医学会学術集会講演要旨集 158回 373 -373 2015年08月 [査読無し][通常論文]
- 吉松 組子, 有川 二郎 医学のあゆみ 253 (1) 63 -67 2015年04月04日 [査読無し][通常論文]
- 清水健太, 吉松組子, 駒貴明, 有川二郎 日本実験動物学会総会講演要旨集 61st 2014年
- 苅和 宏明, 尾崎 由佳, 真田 崇弘, 池中 良徳, 石塚 真由美, 坪田 敏男, 好井 健太朗, 吉松 組子, 有川 二郎, 高島 郁夫 獣医畜産新報 66 (4) 262 -264 2013年04月 [査読無し][通常論文]
ハンタウイルスはげっ歯類媒介性の人獣共通感染症の病原体で、腎症候性出血熱(HFRS)またはハンタウイルス肺症候群(HPS)を引き起こす。近年の日本におけるげっ歯類のハンタウイルス感染状況を明らかにするために、1994年から2010年にかけて国内の様々な地域で捕獲された1658匹のげっ歯類の血清について、抗ハンタウイルス抗体の検出を行った。840例のRattus属げっ歯類(ドブネズミとクマネズミ)は全て抗体陰性であった。北海道以外の地域で捕獲された野生げっ歯類113例はいずれも抗体陰性であったのに対し、北海道で捕獲された705例の野性げっ歯類のうち、エゾヤチネズミの7.4%(26/352)とアカネズミの1.2%(2/168)が抗体を保有していた。(著者抄録) - 駒 貴明, 吉松 組子, 垂石 みどり, 宮下 大輔, 遠藤 理香, 清水 健太, 安田 俊平, 天田 貴子, 瀬戸 隆弘, 村田 亮, 吉田 喜香, 苅和 宏明, 高島 郁夫, 有川 二郎 北海道醫學雜誌 = Acta medica Hokkaidonensia 88 (1) 35 -35 2013年01月01日 [査読無し][通常論文]
- 土佐 紀子, 吉松 組子, 有川 二郎 実験動物と環境 = Laboratory animal and environment 20 (1) 1 -8 2012年04月01日
- 清水健太, 吉松組子, 駒貴明, 伊勢川裕二, 安田俊平, 天田貴子, 五十棲理恵, 有川二郎 日本獣医学会学術集会講演要旨集 153rd 2012年
- 清水健太, 李天成, 安田俊平, 吉松組子, 駒貴明, 長谷部太, 山城哲, HOA Nguyen Thuy, MAI Le Thi Quynh, 有川二郎 日本獣医学会学術集会講演要旨集 154th 2012年
- 駒貴明, 永田典代, 吉松組子, 佐藤由子, 清水健太, 天田貴子, 西尾佐奈恵, 長谷川秀樹, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 60th 2012年
- 天田貴子, 吉松組子, 安田俊平, 清水健太, 駒貴明, 林元展人, 高倉彰, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 60th 2012年
- 天田貴子, 吉松組子, 安田俊平, 駒貴明, 清水健太, 五十棲理恵, 林元展人, 高倉彰, 有川二郎 日本獣医学会学術集会講演要旨集 153rd 2012年
- 清水健太, 吉松組子, 駒貴明, 天田貴子, 西尾佐奈恵, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 60th 2012年
- 有川二郎, 天田貴子, 林元展人, 吉松組子, 安田俊平, 清水健太, 駒貴明, 五十棲理恵, 高倉彰 日本実験動物学会総会講演要旨集 59th 2012年
- 真田崇弘, 瀬戸隆弘, 尾崎由佳, NGONDA Saasa, 吉松組子, 有川二郎, 好井健太朗, 苅和宏明 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 2012年
- 清水健太, 吉松組子, 駒貴明, 安田俊平, 有川二郎 日本獣医学会学術集会講演要旨集 152nd 2011年
- 駒貴明, 吉松組子, 永田典代, 清水健太, 安田俊平, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 58th 2010年
- 安田俊平, 吉松組子, 遠藤理香, 清水健太, 駒貴明, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 58th 2010年
- 清水健太, 吉松組子, 駒貴明, 安田俊平, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 58th 2010年
- 清水健太, 吉松組子, 駒貴明, 遠藤理香, 安田俊平, ERDENESAIKHAN Tegshduuren, 有川二郎 日本獣医学会学術集会講演要旨集 150th 2010年
- 吉田喜香, 苅和宏明, 真田崇弘, NGONDA Saasa, 瀬戸隆弘, 吉松組子, 有川二郎, 好井健太朗, 高島郁夫 日本ウイルス学会学術集会プログラム・抄録集 58th 2010年
- 清水健太, イブラヒムイマヌリサ, 吉松組子, 遠藤理香, 安田俊平, 駒貴明, エルテネサイハンテグシドーレン, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 57th 2009年
- エルテネサイハンテグシドーレン, 清水健太, 吉松組子, 遠藤理香, 駒貴明, 安田俊平, 有川二郎, 石原智明 日本ウイルス学会学術集会プログラム・抄録集 57th 2009年
- 安田俊平, 吉松組子, 遠藤理香, 清水健太, 駒貴明, TEGSHDUUREN Erdenesaikhan, 垂石みどり, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 57th 2009年
- 遠藤理香, 吉松組子, 駒貴明, 清水健太, 安田俊平, TEGSHDUUREN Erdenesaihan, 垂石みどり, 海老原秀喜, 宮下大輔, 瀬戸隆弘, HERNANDEZ Cornelio S., ALMARAZ Maria L.R., RAMOS Celso, 苅和宏明, 高島郁夫, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 57th 2009年
- 駒貴明, 吉松組子, 垂石みどり, 遠藤理香, 清水健太, 安田俊平, エルテネサイハンテグシドーレン, 海老原秀喜, HERNANDEZ Cornelio S., ALMARAZ Maria L.R., RAMOS Celso, 宮下大輔, 瀬戸隆弘, 苅和宏明, 高島郁夫, ENRIA Delia, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 57th 2009年
- 駒貴明, 吉松組子, 垂石みどり, 遠藤理香, 清水健太, 安田俊平, テグシドーレン エルデネサイハン, 海老原秀喜, HERNANDEZ Cornelio S., ALMARAZ Maria L.R., RAMOS Celso, 宮下大輔, 瀬戸隆弘, 苅和宏明, 高島郁夫, ENRIA Delia, 有川二郎 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 43rd 2009年
- 真田崇弘, 苅和宏明, 瀬戸隆弘, 谷川洋一, 宮下大輔, 吉松組子, 有川二郎, 好井健太朗, 高島郁夫 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 43rd 2009年
- S. Chandy, K. Yoshimatsu, R. G. Ulrich, M. Schlegel, D. J. Prakash, V. Balraj, P. Abraham, J. Arikawa, G. Sridharan INTERNATIONAL JOURNAL OF INFECTIOUS DISEASES 12 E131 -E131 2008年12月 [査読無し][通常論文]
- 駒貴明, 吉松組子, 垂石みどり, 遠藤理香, 海老原秀喜, 有川二郎 日本ウイルス学会学術集会プログラム・抄録集 56th 2008年
- 瀬戸隆弘, 苅和宏明, 谷川洋一, 吉松組子, 中村一郎, 宮下大輔, 中内美名, 好井健太朗, 有川二郎, 高島郁夫 日本ウイルス学会学術集会プログラム・抄録集 55th 2007年
- 宮下大輔, 苅和宏明, 瀬戸隆弘, 好井健太郎, 吉松組子, 有川二郎, 高島郁夫 日本ウイルス学会学術集会プログラム・抄録集 55th 2007年
- 有川 二郎, 吉松 組子, Kumperasart Sanit 獣医畜産新報 58 (4) 326 -328 2005年04月
- Jonas Schmidt, Burkhard Jandrig, Boris Klempa, Kumiko Yoshimatsu, Jiro Arikawa, Helga Meisel, Matthias Niedrig, Christian Pitra, Detlev H. Krüger, Rainer Ulrich Virus Genes 30 (1) 37 -48 2005年01月 [査読無し][通常論文]
Seoul virus (SEOV) is a hantavirus causing a mild to moderate form of hemorrhagic fever with renal syndrome that is distributed mainly in Asia. The nucleocapsid (N) protein-encoding sequence of SEOV (strain 80-39) was RT-PCR-amplified and cloned into a yeast expression vector containing a galactose-inducible promoter. A survey of the pattern of synonymous codon preferences for a total of 22 N protein-encoding hantavirus genes including 13 of SEOV strains revealed that there is minor variation in codon usage by the same gene in different viral genomes. Introduction of the expression plasmid into yeast Saccharomyces cerevisiae resulted in the high-level expression of a hexahistidine-tagged N protein derivative. The nickel-chelation chromatography purified, yeast-expressed SEOV N protein reacted in the immunoblot with a SEOV-specific monoclonal antibody and certain HTNV- and PUUV-cross-reactive monoclonal antibodies. The immunization of a rabbit with the recombinant N protein resulted in the induction of a high-titered antibody response. In ELISA studies, the N protein was able to detect antibodies in sera of experimentally infected laboratory rats and in human anti-hantavirus-positive sera or serum pools of patients from different geographical origin. The yeast-expressed SEOV N protein represents a promising antigen for development of diagnostic tools in serology, sero prevalence studies and vaccine development. © 2005 Springer Science+Business Media, Inc. - M Okumura, K Yoshimatsu, K Araki, BH Lee, A Asano, T Agui, J Arikawa ARCHIVES OF VIROLOGY 149 (12) 2427 -2434 2004年12月 [査読無し][通常論文]
Monoclonal antibody E5/G6 recognized a linear epitope common to hantavirus nucleocapsid proteins. Using synthetic peptides, we identified epitope E5/G6 as the 9 merYEDVNGIRK (NP 165 - 173), in which D167, G170, I171, and R172 are indispensable. Furthermore, all the peptides synthesized using various hantavirus sequences bound MAb E5/G6 consistently, despite the existence of several amino acid variations in this region. These results indicate that MAb E5/G6 is a useful tool for detecting hantavirus antigen in rodent or patient tissues using Western blotting or other immunohistochemical assays. - Michiko Ogino, Kumiko Yoshimatsu, Hideki Ebihara, Koichi Araki, Byoung-Hee Lee, Megumi Okumura, Jiro Arikawa Journal of Virology 78 (19) 10776 -10782 2004年10月 [査読無し][通常論文]
Hantaan virus (HTNV)-infected Vero E6 cells undergo cell fusion with both infected and uninfected cells under low-pH conditions. Flow cytometry and fluorescence microscopy of HTNV-infected Vero E6 cells showed that envelope glycoproteins (GPs) were located both on the cell surface and in the cytoplasm. Neutralizing monoclonal antibodies (MAbs) against the G1 and G2 envelope GPs inhibited cell fusion, whereas nonneutralizing MAbs against G1 or G2 and MAbs against the nucleocapsid protein (NP) did not. Transfected Vero E6 cells that expressed GPs but not those that expressed NP fused and formed syncytia. These results indicate that HTNV GPs act as fusogens at the cell surface. No fusion activity was observed either in infected Vero cells that were passaged more than 150 times or in BHK-21 cells, although GPs appeared to localize to the cell surface. This variability in fusion induction suggests the involvement of host cell factors in the process of cell membrane fusion. - K Araki, K Yoshimatsu, BH Lee, M Okumura, H Kariwa, Takashima, I, J Arikawa ARCHIVES OF VIROLOGY 149 (7) 1373 -1382 2004年07月 [査読無し][通常論文]
To investigate age-dependent differences in hantavirus-specific CD8(+) T-cell responses, mice were inoculated with 0.1 50% newborn mouse lethal dose of Hantaan virus (HTNV) at 0, 3, 7, 14, or 35 days after birth. HTNV-specific CD8(+) T cells producing gamma interferon (IFN-gamma) were measured on day 30 after HTNV inoculation. Although no IFN-gamma-producing HTNV-specific CD8(+) T cells were detected in most of the mice inoculated with HTNV on day 0 after birth, most mice inoculated at 3, 7, 14, or 35 days had HTNV-specific CD8(+) T cells. The production of tumor necrosis factor alpha (TNF-alpha) by IFN-gamma-producing CD8(+) T cells and the cytotoxic activity against HTNV-infected target cells were similar in immature and adult mice. However, the number of IFN-gamma-producing HTNV-specific CD8(+) T cells was significantly less in mice inoculated with HTNV at 3 days than in older mice. In addition, a strong correlation between HTNV persistence and a lack of HTNV-specific CD8(+) T cells was observed. These results suggest that mice over 7 days old have the ability to induce functional HTNV-specific CD8(+) T-cell responses that are indistinguishable from the responses of adult mice, and that HTNV-specific CD8(+) T cells are important for clearance of HTNV. - J Arikawa, K Yoshimatsu, H Wang, H Kariwa, K Lokugamage, N Lokugamage, Takashima, I JOURNAL OF CLINICAL VIROLOGY 28 S28 -S28 2003年12月 [査読無し][通常論文]
- BH Lee, K Yoshimatsu, A Maeda, K Ochiai, M Morimatsu, K Araki, M Ogino, S Morikawa, J Arikawa VIRUS RESEARCH 98 (1) 83 -91 2003年12月 [査読無し][通常論文]
We performed yeast two-hybrid screening of a human kidney cell cDNA library to study the biological role of the hantavirus nucleocapsid protein (NP). We found that Seoul virus (SEOV) and Hantaan virus (HTNV) NPs were associated with small ubiquitin-like modifier (SUMO)-1-interacting proteins PIAS 1, PIASxbeta, HIPK2, CHD3, and TTRAP, which interacted with the SUMO-1 conjugating enzyme (Ubc-9) and SUMO-1 in the yeast two-hybrid assay. Interactions between the HIPK2, CHD3, and TTRAP proteins and SEOV NP were also shown in a mammalian two-hybrid assay. However, there was no interaction between PIAS proteins and NP, which was probably due to the inhibitory effect of PIAS on transcription in the mammalian two-hybrid assay. Nevertheless, a co-expression experiment suggested the existence of a PIAS-NP interaction in the cytoplasm. The region spanning amino acids 100-125 of SEOV NP, which represents a critical region for NP-NP polymerization. was found to be responsible for the interaction with SUMO-1-related molecules in both the yeast and mammalian two-hybrid assays. These results add to the information on interactions of hantavirus NP and host cellular proteins. (C) 2003 Elsevier B.V. All rights reserved. - N Lokugamage, H Kariwa, K Lokugamage, T Hagiya, H Miyamoto, MA Iwasa, K Araki, K Yoshimatsu, J Arikawa, T Mizutani, Takashima, I JOURNAL OF VETERINARY MEDICAL SCIENCE 65 (11) 1189 -1194 2003年11月 [査読無し][通常論文]
Puumala (PUU) virus and PUU-related viruses are difficult to isolate in cell culture. To determine whether animal inoculation would be a better alternative for virus recovery, the Sotkamo strain of PUU virus was inoculated into several animal species. Newborn Mongolian gerbils (MGs), mice, and rats were infected with the Sotkamo strain by intracerebral (ic), intraperitoneal (ip), and subcutaneous (sc) inoculation. Antibodies to PUU appeared in MGs at 30 days post-infection (dpi), and in mice and rats at 15 dpi. Interestingly, virus appeared at 7 dpi in lung and brain of MGs inoculated via ic and ip routes. Virus was detected in all tested tissues of MGs at 15 dpi, with a peak level of 1.36 x 10(5) focus forming units (FFU)/g in brain tissue. The virus titer declined with the onset of the antibody response and became undetectable by 75 dpi, when the antibody titer reached the maximum level. The appearance of the virus in mice and rats was delayed as compared to MGs, and the virus titer was apparently lower, at approximately 4 to 8 x 10(3) FFU/g, at 15 dpi. In addition, lung homogenates of antibody-positive Clethrionomys (C) rufocanus (captured in Tobetsu, Hokkaido, Japan) were inoculated into MGs by the ic route. PUU-related viral RNA was detected at 16 dpi in the brains of MG inoculated with the lung homogenate, and antibodies were detected at 45 dpi. These findings indicate that newborn MG inoculation is an efficient method to recover PUU and PUU-related viruses. - BH Lee, K Yoshimatsu, K Araki, M Ogino, M Okumura, K Tsuchiya, H Kariwa, J Arikawa ARCHIVES OF VIROLOGY 148 (10) 1885 -1897 2003年10月 [査読無し][通常論文]
Peroxidase-labeled staphylococcal protein A, streptococcal protein G, and antibodies directed against Mus musculus (mouse), Rattus norvegicus (rat), Mesocretus auratus (hamster), and Peromyscus leucopus were examined for their reactivity with immunoglobulin G (IgG) from various rodent species. The purpose of this study was to identify the optimal secondary antibodies or reagents for specific serodiagnosis of hantavirus infection in various rodent species. Using ELISA, a total of 65 sera from 29 rodent species of the family Muridae and one serum sample from family Octodontidae were compared for IgG reactivity with the six different reagents. The results demonstrate that the reactivities of the secondary antibodies and reagents to the sera varied, even among sera from rodents of the same genus. Hantavirus-specific antibody ELISA revealed that hantavirus-infected rodent sera obtained from M. musculus, R. norvegicus, Apodemus agrarius, A. peninsulae, and Bandicota indica bound to the six different conjugates in a similar pattern as that detected in IgG ELISA. These results indicate that the applicability of secondary antibodies and protein A and G should be carefully evaluated before use for serodiagnosis in different rodent species. - H Kariwa, H Tanabe, T Mizutani, Y Kon, K Lokugamage, N Lokugamage, MA Iwasa, T Hagiya, K Araki, K Yoshimatsu, J Arikawa, Takashima, I ARCHIVES OF VIROLOGY 148 (9) 1671 -1685 2003年09月 [査読無し][通常論文]
Seoul virus is a hantavirus that causes hemorrhagic fever with renal syndrome (HFRS). The virion has a tripartite (S, M, and L) negative-stranded RNA genome, which is characteristic of the family Bunyaviridae. However, the molecular basis of virus replication is not well known. We established a Northern blot hybridization (NB) procedure using digoxygenin-labeled RNA probes, to quantitate the hantaviral plus- and minus-strand RNAs separately. Virus RNA replication was analyzed in infected Vero E6 cells. When the Vero E6 cells were infected with Seoul virus strain KI-83-262 (KI) at m.o.i.=0.25, the plus-strand RNA was detected within 1 h post-infection (hpi), and the minus-strand RNA was detected subsequently. Using laser confocal microscopy, the nucleocapsid protein (NP) was detected within 2 hpi, and accumulated as scattered granules in the cytoplasm until 24 hpi. In contrast, the G2 protein first appeared at 8 hpi, was immediately transported to the Golgi, and accumulated in the Golgi until 24 hpi. Infectious virus particles were released into the medium at 24 h hpi. These findings indicate that hantavirus RNA replication starts with the appearance of NP at 2 hpi, glycoproteins then accumulate gradually in the Golgi, and virion formation is initiated once the viral RNAs and proteins have accumulated. - H Miyamoto, H Kariwa, K Araki, K Lokugamage, D Hayasaka, BZ Cui, N Lokugamage, LI Ivanov, T Mizutani, MA Iwasa, K Yoshimatsu, J Arikawa, Takashima, I ARCHIVES OF VIROLOGY 148 (8) 1543 -1556 2003年08月 [査読無し][通常論文]
Hemorrhagic fever with renal syndrome (HFRS) is endemic in East Asia and Europe. The disease is caused by several viruses belonging to the genus Hantavirus, including the Hantaan virus (HTNV), Seoul virus (SEOV), Dobrava Belgrade virus (DOBV), and Puumala virus (PUUV). Recently, HTNV-related viruses, Amur (AMR) and Far East (FE) genotypes were identified as causative agents of HFRS in Far Eastern Russia. To investigate the epidemiology of HFRS and virus transmission, we collected sera from 17 acute and 32 convalescent patients who were clinically diagnosed with HFRS in the Khabarovsk region of Far Eastern Russia, and detected anti-hantavirus antibodies using an ELISA that can differentiate the infected virus serotype using truncated hantavirus nucleocapsid protein antigen. Sixteen of the 17 acute phase patients had antibodies to hantavirus, and all the positive sera had higher optical densities for HTNV-specific antigen than for SEOV-, DOBV-, or PUUV-specific antigens. The partial M segment of the viral genome was amplified from blood clots from three acute patients by PCR. The nucleotide sequences had closer identities to the FE genotype (> 96%) than to the prototype HTNV (88 to 89%) or AMR genotype (81 to 83%). A phylogenetic analysis found that the virus sequences from the patients clustered with the FE type, and were distinct from the AMR type. Thirty-one of 32 convalescent patient sera had antibodies to HTNV-specific antigen. These data suggest ELISA system can detect HTNV-specific antibodies to the FE type, which may be responsible for most of the HFRS in Khabarovsk. - Hantavirus-specific CD8(+)-T-cell responses in newborn mice persistently infected with Hantaan virusK Araki, K Yoshimatsu, BH Lee, H Kariwa, Takashima, I, J Arikawa JOURNAL OF VIROLOGY 77 (15) 8408 -8417 2003年08月 [査読無し][通常論文]
The relationship between virus-specific CD8(+)-T-cell responses and viral persistence was studied in mice by using Hantaan virus (HTNA). We first established a simple method for measuring levels of virus-specific CD8(+) T cells by How cytometry. Next, to produce a mouse model of persistent HTNV infection, newborn mice were inoculated subcutaneously within 24 h of birth with 1 or 0.150% newborn mouse lethal dose of HTNV. All mice that escaped lethal infection were persistently infected with HTNV until at least 30 days after virus inoculation and had no virus-specific CD8(+) T cells producing gamma interferon (IFN-gamma). Subsequently, the virus was eliminated from some of the mice, depending on the appearance of functional virus-specific CD8(+) T cells, which have the ability to produce IFN-gamma and tumor necrosis factor alpha (TNF-alpha) and have cytotoxic activity. Neutralizing antibodies were detected in all mice, regardless of the presence or absence of virus. In the acute phase, which occurs within 30 days of infection, IFN-gamma-producing HTNV-specific CD8(+) T cells were detected on day 15 after virus inoculation. However, TNF-alpha production and the cytotoxic activity of these specific CD8(+) T cells were impaired and HTNV was not removed. Almost all of these specific CD8(+) T cells disappeared by day 18. These results suggest that functional HTNV-specific CD8(+) T cells are important for clearance of HTNV. - K Yoshii, D Hayasaka, A Goto, M Obara, K Araki, K Yoshimatsu, J Arikawa, L Ivanov, T Mizutani, H Kariwa, Takashima, I JOURNAL OF VIROLOGICAL METHODS 108 (2) 171 -179 2003年03月 [査読無し][通常論文]
A recombinant plasmid that expresses the tick-borne encephalitis (TBE) virus premembrane (prM) and envelope (E) proteins in mammalian cells was constructed. Recombinant proteins retained antigenic and conformational structures similar to those of native virus proteins, and transfected cells released virus-like particles (VLPs), which were 1.13-1.14 g/ml in density and 20-30 nm in diameter, into the culture medium. Recombinant E proteins were used for the development of an enzyme-linked immunosorbent assay (ELISA) to detect TBE virus-specific IgM and IgG antibodies in serum. The results of this ELISA correlated well with the results of commercial ELISA, when tested with 95 serum samples from clinically TBE-suspected patients. In addition, ELISA using recombinant antigens showed no cross-reactivity against serum from Japanese encephalitis (JE) patients, despite the cross-reactivity shown by commercial ELISA systems. These observations indicated that this newly developed ELISA system could distinguish tick-borne encephalitis from Japanese encephalitis infection, and that it constitutes a useful and safe alternative to conventional ELISA systems. (C) 2002 Published by Elsevier Science B.V. - A Takakura, K Goto, T Itoh, K Yoshimatsu, Takashima, I, J Arikawa EXPERIMENTAL ANIMALS 52 (1) 25 -30 2003年01月 [査読無し][通常論文]
A recombinant nucleocapsid protein of Hantaan virus (HTN) 76-118 strain expressed in E. coli was applied as a serodiagnostic antigen in an enzyme-linked immunosorbent assay (rHTN-ELISA) for detection of hantavirus antibody in rat sera. The sensitivity and specificity of the rHTN-ELISA were compared with those of the indirect immunofluoresent assay (IFA) using virus-infected cells. The sensitivity of rHTN-ELISA was similar to that of the IFA both in experimentally SR-11 infected rat and naturally infected rat sera. Sera showing a low antibody titer in IFA and suspected to be negative by other methods were also found to be negative in rHTN-ELISA. These results indicate that rHTN-ELISA is effective as a screening method for serodiagnosis of hantaviruses, because of its high sensitivity, specificity, safety and suitability for processing large number of samples. - M Ogino, H Ebihara, BH Lee, K Araki, A Lundkvist, Y Kawaoka, K Yoshimatsu, J Arikawa CLINICAL AND DIAGNOSTIC LABORATORY IMMUNOLOGY 10 (1) 154 -160 2003年01月 [査読無し][通常論文]
A vesicular stomatitis virus (VSV) pseudotype bearing hantavirus envelope glycoproteins was produced and used in a neutralization test as a substitute for native hantavirus. The recombinant VSV, in which the enveloped protein gene (G) was replaced by the green fluorescent protein gene and complemented with G protein expressed in trans (VSVDeltaG*G), was kindly provided by M. A. Whitt. 293T cells were transfected with plasmids for the expression of envelope glycoproteins (G1 and G2) of HTNV or SEOV and were then infected with VSVDeltaG*G. Pseudotype VSV with the Hantaan (VSVDeltaG*-HTN) or Seoul (VSVDeltaG*-SEO) envelope glycoproteins were harvested from the culture supernatant. The number of infectious units (IU) of the pseudotype VSVs ranged from 10(5) to 10(6)/ml. The infectivity of VSVDeltaG*-HTN and VSVDeltaG*-SEO was neutralized with monoclonal antibodies, immune rabbit sera, and sera from patients with hemorrhagic fever with renal syndrome, and the neutralizing titers were similar to those obtained with native hantaviruses. These results show that VSVDeltaG*-HTN and -SEO can be used as a rapid, specific, and safe neutralization test for detecting hantavirus-neutralizing antibodies as an effective substitute for the use of native hantaviruses. Furthermore, the IU of VSVDeltaG*-HTN and -SEO did not decrease by more than 10-fold when stored at 4degreesC for up to 30 days. The stability of the pseudotype viruses allows distribution of the material to remote areas by using conventional cooling boxes for use as a diagnostic reagent. - Kumiko Yoshimatsu, Byoung-Hee Lee, Koichi Araki, Masami Morimatsu, Michiko Ogino, Hideki Ebihara, Jiro Arikawa Journal of Virology 77 (2) 943 -952 2003年01月 [査読無し][通常論文]
Multimerization of the Hantaan virus nucleocapsid protein (NP) in Hantaan virus-infected Vero E6 cells was observed in a competitive enzyme-linked immunosorbent assay (ELISA). Recombinant and truncated NPs of Hantaan, Seoul, and Dobrava viruses lacking the N-terminal 49 amino acids were also detected as multimers. Although truncated NPs of Hantaan virus lacking the N-terminal 154 amino acids existed as a monomer, those of Seoul and Dobrava formed multimers. The multimerized truncated NP antigens of Seoul and Dobrava viruses could detect serotype-specific antibodies, whereas the monomeric truncated NP antigen of Hantaan virus lacking the N-terminal 154 amino acids could not, suggesting that a hantavirus serotype-specific epitope on the NP results in multimerization. The NP-NP interaction was also detected by using a yeast two-hybrid assay. Two regions, amino acids 100 to 125 (region 1) and amino acids 404 to 429 (region 2), were essential for the NP-NP interaction in yeast. The NP of Seoul virus in which the tryptophan at amino acid number 119 was replaced by alanine (W119A mutation) did not multimerize in the yeast two-hybrid assay, indicating that tryptophan 119 in region 1 is important for the NP-NP interaction in yeast. However, W119A mutants expressed in mammalian cells were detected as the multimer by using competitive ELISA. Similarly, the truncated NP of Seoul virus expressing amino acids 155 to 429 showed a homologous interaction in a competitive ELISA but not in the yeast two-hybrid assay, indicating that the C-terminal region is important for the multimerization detected by competitive ELISA. Combined, the results indicate that several steps and regions are involved in multimerization of hantavirus NP. - K Lokugamage, H Kariwa, D Hayasaka, BZ Cui, T Iwasaki, N Lokugamage, LI Ivanov, Volkov, VI, VA Demenev, R Slonova, G Kompanets, T Kushnaryova, T Kurata, K Maeda, K Araki, T Mizutani, K Yoshimatsu, J Arikawa, Takashima, I EMERGING INFECTIOUS DISEASES 8 (8) 768 -776 2002年08月 [査読無し][通常論文]
In an epizootiologic survey of 122 rodents captured in Vladivostok, Russia, antibodies positive for hantavirus were found in Apodemus peninsulae (4/70), A. agrarius (1/39), and Clethrionomys rufocanus (1/8). The hantavirus sequences identified in two seropositive A. peninsulae and two patients with hemorrhagic fever with renal syndrome (HFRS) from the Primorye region of Far East Russia were designated as Solovey and Primorye, respectively. The nucleotide sequences of the Solovey, Primorye, and Amur (obtained through GenBank) sequences were closely related (>92% identity). Solovey and Primorye sequences shared 84% nucleotide identity with the prototype Hantaan 76-118. Phylogenetic analysis also indicated a close relationship between Solovey, Primorye, Amur, and other viruses identified in Russia, China, and Korea. Our findings suggest that the Korean field mouse (A. peninsulae) is the reservoir for a hantavirus that causes HFRS over a vast area of east Asia, including Far East Russia. - Epizootiological survey of hantavirus among rodent species in Ningxia Hui Autonomous Province, ChinaH Kariwa, CB Zhong, K Araki, K Yoshimatsu, K Lokugamage, N Lokugamage, ME Murphy, T Mizutani, J Arikawa, H Fukushima, H Xiong, JHC Chen, Takashima, I JAPANESE JOURNAL OF VETERINARY RESEARCH 49 (2) 105 -114 2001年08月 [査読無し][通常論文]
Hantaviral antibodies were detected in the sera from Apodemus (A.) agrarius and A. peninsulae captured in Ningxia province, China by several different serological diagnostic methods. A total of 409 sera from rodent and insectivore species were collected in 1999 and examined by indirect immunofluorescent antibody assay (IFA). Among them, 19 of 191 (9.9%) sera of A. agrarius and 1 of 13 (7.7%) sera of A. peninsulae were positive for hantaviral antibodies. The other species (Rattus norvegicus, Mus musculus, Cricetulus triton, and Sorex cylindricauda) were negative. The reaction pattern of positive serum was characterized as scattered and granular virus antigens in the cytoplasm of hantavirus infected Vero E6 cells. Some of the A. agrarius sera positive for hantavirus were further examined by Western blotting (WB), enzyme-linked immunosorbent assay (ELISA), and the focus reduction neutralization test (FRNT). By WB, positive sera showed the same specific reaction pattern of baculovirus-expressed recombinant hantaviral nucleocapsid protein, as shown in hantavirus-immune serum. By ELISA, IFA-positive sera showed significantly higher optical densities (around 1.0) than the negative A. agrarius sera. Hantaan type hantavirus was neutralized with the positive sera. These results suggest that A. agrarius have hantavirus infection and may play a role as a reservoir animal for hantavirus in Ningxia Hui Autonomous Province, China. - K Araki, K Yoshimatsu, M Ogino, H Ebihara, A Lundkvist, H Kariwa, Takashima, I, J Arikawa JOURNAL OF CLINICAL MICROBIOLOGY 39 (7) 2397 -2404 2001年07月 [査読無し][通常論文]
Truncated recombinant nucleocapsid proteins (rNPs) of Hantaan virus (HTNV), Seoul virus (SEOV), and Dobrava virus (DOBV) were expressed by a baculovirus system. The truncated rNPs, which lacked 49 (rNP50) or 154 (rNP155) N-terminal amino acids of the NPs of HTNV, SEOV, and DOBV, were able to differentiate HTNV-, SEOV-, and DOBV-specific immune sera. Recombinant NP50s retained higher reactivities than rNP155s and were proven useful for enzyme-linked immunosorbent assay (ELISA). The ELISAs based on the rNP50s of HTNV, SEOV, and DOBV successfully differentiated three groups of patient sera, previously defined by neutralization tests: 17 with HTNV infection, 12 with SEOV infection, and 20 with DOBV infection. The entire rNP of Puumala virus (PUUV) distinguished PUUV infection from the other types of hantavirus infection. Serotyping with these rNP50s can be recommended as a rapid and efficient system for hantavirus diagnosis. - ME Murphy, H Kariwa, T Mizutani, H Tanabe, K Yoshimatsu, J Arikawa, Takashima, I JOURNAL OF VETERINARY MEDICAL SCIENCE 63 (6) 637 -645 2001年06月 [査読無し][通常論文]
Mechanisms of anti-hantaviral activities of bovine lactoferrin (LF) and ribavirin (Rbv) were investigated. Hantavirus focus formation at 48 hr was 1546 of the control in cells treated with 400 mug/ml LF for 1 hr at 37 degreesC prior to viral infection. Post infection treatment with 100 mug/ml Rbv also inhibited the focus formation to 2.5% of the control. Combined LF pre- and Rbv post-infection treatment completely inhibited focus formation. Viral glycoprotein (G2) and nucleocapsid protein (NP) syntheses were delayed in LF pre treated cells up to 24 hr post infection (hpi) but became comparable to the control by 48 hpi. Further, LF inhibited viral shedding at 24 hpi but did not inhibit shedding after 48 hpi. However, Rbv was able to inhibit synthesis of viral proteins, (+) and (-) strand RNAs also inhibited viral shedding after 24 hr. These results suggest that LF inhibits viral adsorption to cells, while Rbv inhibits viral RNA synthesis. For in vivo trials of LF and Rbv, LF pre- and Rbv post-treatment were evaluated in suckling mice infected with hantavirus, of which 7% survived. LF concentrations of 40 and 160 mg/cg administered prior to viral challenge improved survival rates to 15% and 70%, respectively for single administration and 85% and 94%. respectively, for double administration. Rbv concentrations of 25 and 50 mg/kg gave survival rates of 68% and 81%, respectively. This suggests that both LF and Rbv are efficacious in hantavirus infection in vivo. - J Arikawa, K Yoshimatsu, H Kariwa JAPANESE JOURNAL OF INFECTIOUS DISEASES 54 (3) 95 -102 2001年06月 [査読無し][通常論文]
Hantaviruses cause two severe human diseases: hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). Various rodent species act as animal reservoirs for hantavirus. In Japan, urban rat- (Rattus norvegicus) and laboratory rat-derived human infections were reported during the 1960s and 1970s-1984, respectively. Although no human cases of infection have been reported since 1984, infected urban rats have been found throughout Japan, and infected grey red-backed voles (Clethrionomys rufocanus) have been identified in Hokkaido. These carriers can be considered to be potential sources of human infection. This review examines the epidemiology and epizootiology of this important zoonosis in Japan. - HK Jin, K Yoshimatsu, A Takada, M Ogino, A Asano, J Arikawa, T Watanabe ARCHIVES OF VIROLOGY 146 (1) 41 -49 2001年 [査読無し][通常論文]
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. - H Wang, K Yoshimatsu, H Ebihara, M Ogino, K Araki, H Kariwa, ZX Wang, ZZ Luo, DX Li, CS Hang, J Arikawa VIROLOGY 278 (2) 332 -345 2000年12月 [査読無し][通常論文]
The antigenic and genetic properties of 46 hantaviruses from China, 13 from patients, 23 from rodents, and 10 from unknown hosts, were compared with those of other hantaviruses. The viruses were classified as either Hantaan (HTN) or Seoul (SEO) viruses. A phylogenetic analysis of the partial M (300 bp) and S (around 485 bp) genomes of HTN viruses identified nine distinct genetic subtypes, one consisting of isolates from Korea. The SEC viruses were divided into five genetic subtypes, although they had less variability than the HTN subtypes. There was a correlation between the subtype and province of origin for four subtypes of HTN viruses, confirming geographical clustering. Hantaan virus NC167 isolated from Niviventer confucianus and SEO virus Gou3 isolated from Rattus rattus were the basal clades in each virus. The phylogenetic trees constructed from the entire S and M segments suggested that NC167 was introduced to N. confucianus in a host-switching event. The reactivity of a panel of 35 monoclonal antibodies was almost exactly the same in NC167 and a representative HTN virus and in Gou3 and a representative SEC virus. However. there was a one-way cross-neutralization between them. These results confirm the varied nature of Murinae-associated hantaviruses in China. (C) 2000 Academic Press. - H Ebihara, K Yoshimatsu, M Ogino, K Araki, Y Ami, H Kariwa, Takashima, I, DX Li, J Arikawa JOURNAL OF VIROLOGY 74 (19) 9245 -9255 2000年10月 [査読無し][通常論文]
Two Hantaan virus strains, clone 1 (cl-1), which is virulent in newborn mice, and its attenuated mutant (mul1E10), were used to examine the pathogenesis of Hantaan virus infection in a mouse model and identify virus factors relating to virulence. After subcutaneous inoculation of newborn BALB/c mice, cl-1 caused fatal disease with high viral multiplication in peripheral organs, but mul1E10 produced nonfatal infection with a low level of virus multiplication. Intracerebral inoculation of either strain caused fatal disease. Histopathological changes in the dead animals were prominent in the brain, indicating that the brain is the target organ and produces the fatal outcome. These results indicate that mul1E10 has a generally less virulent phenotype, and because of decreased multiplication in peripheral tissues, neuroinvasiveness is also decreased. An experiment with genetic reassortant viruses showed that in newborn mice the M segment is the most related to virulence and the L segment is partly related. Sequence comparison detected a single deduced amino acid change (cl-l Ile to mul1E10 Thr) at amino acid number 515 in glycoprotein G1. One nucleotide change, but no amino acid substitution, was observed in the noncoding region of the L segment. In mouse brain microvascular endothelial cells in vitro, viruses possessing a cl-1-derived M segment grew more rapidly than viruses containing a mul1E10 derived M segment. These results suggest that the single amino acid change in the glycoprotein alters peripheral growth, which affects invasion of the central nervous system in mice. - ME Murphy, H Kariwa, T Mizutani, K Yoshimatsu, J Arikawa, Takashima, I ARCHIVES OF VIROLOGY 145 (8) 1571 -1582 2000年 [査読無し][通常論文]
Bovine lactoferrin (LF) and ribavirin (Rbv) were tested as antiviral agents against Seoul type hantavirus (SR-11 strain) in vitro. Hantaviral foci number in Vero E6 cells infected with SR-11 was reduced with LF treatment by 5 days post infection to obtain a 50% effective dose (ED50) Of 2500 mu g/ml, while pretreatment with LF was highly efficacious having an ED50 of 39 mu g/ml. Conversely, 1 h pretreatment with Rbv revealed no inhibition of viral focus formation but could significantly reduce the number of viral foci (ED50: 10 mu g/ml) when used from the time of viral infection. One hour pre-treatment of the cell monolayer with LF and subsequent addition of Rbv revealed a synergistic anti-hantaviral effect against SR-11, <20 FFU/ml as compared to 10(5) foci/ml in the control. One hour treatment of SR-11 with LF prior to cell inoculation gave an ED50 of 312.5 mu g/ml. Whereas, washing the LF-pretreated cell monolayer with PBS demonstrated minimal focus reduction, suggesting LF lightly adheres to cells. These results indicate that LF has anti-hantaviral activity in vitro and inhibition of virus adsorption to cells which play an important role in revealing the anti-hantaviral activity of LF. This paper reports for the first time the anti-hantaviral effect of LF. - H Kariwa, K Yoshimatsu, K Araki, K Chayama, H Kumada, M Ogino, H Ebihara, ME Murphy, T Mizutani, Takashima, I, J Arikawa MICROBIOLOGY AND IMMUNOLOGY 44 (5) 357 -362 2000年 [査読無し][通常論文]
Hantaviral antibodies were detected in the sera from patients with hepatic disease of unknown etiology in Japan by several different serological diagnostic methods. A total of 105 sera from diseased patients which were negative to A-G hepatitis virus infections in the Tokyo area were tested. Among them, 3 out of 73 sera from patients with chronic hepatic disease were positive to hantaviral antibody by enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescent antibody assay (IFA) and Western blot analysis (WB), Neutralizing antibody titers of the 3 sera to Seoul virus (SEO) were 4 to 8 times higher than those to Hantaan virus (HTN), However, all of the 32 sera from patients with acute hepatitis were negative for hantaviral antibody, Among the 60 patients with chronic hepatitis in Hokkaido which were serologically negative to B and C hepatitis virus infection, one was positive for hantaviral antibody by ELISA and WE. In contrast, the sera from healthy adults in Japan, 550 from the Honshu and Kyushu regions, and 1,000 from the Hokkaido region, were negative for hantavirus antibody. These results show that hantaviral antibodies are more frequently detected in patients with hepatic disease than in healthy adults. However, the observation that no positive sera were detected from patients with acute hepatitis implies that hantavirus might not be directly related to hepatitis. - K Tsujimura, T Mizutani, H Kariwa, K Yoshimatsu, M Ogino, Y Morii, H Inagaki, J Arikawa, Takashima, I JOURNAL OF VETERINARY MEDICAL SCIENCE 61 (2) 113 -117 1999年02月 [査読無し][通常論文]
For a serological diagnostic test for Borna disease (BD), we developed a capture ELISA with specificity and sensitivity based on detection of antibodies against BD virus (BDV) p40 protein. Using our capture ELISA system, the antibody response of rats inoculated intracerebrally with BDV at 4 weeks after birth showed a sharp increase from 1 to 4 weeks postinoculation (p.i.) and a steady level after 5 weeks p.i. To investigate prevalence of BDV infection among wild rats, we examined sera of Rattus norvegicus in Kami-iso town, Oshima district, Hokkaido, suggesting that rats in this area had not been infected by BDV. - H Kariwa, K Yoshimatsu, J Sawabe, E Yokota, J Arikawa, Takashima, I, H Fukushima, A Lundkvist, FN Shubin, LM Isachkova, RA Slonova, GN Leonova, N Hashimoto VIRUS RESEARCH 59 (2) 219 -228 1999年02月 [査読無し][通常論文]
Seroepizootiologic surveys among wild rodents were carried out in Japan and Far East Russia in 1995 and 1996. Seropositive animals were only identified in Clethrionomys rufocanus (23/134) in Hokkaido, Japan. On the other hand, seropositives were identified in C. rufocanus (1/8), Apodemus argrarius (2/66), Apodemus spp. (2/26) and Microtus fortis (3/22) in Vladivostok, Far East Russia. Total RNA was isolated from lungs of seropositive animals and the S genome segments were amplified by PCR, cloned and sequenced. The S and M genomes of hantavirus, derived from Japanese C. rufocanus (Tobetsu genotype), were most closely related with Puumala viruses (76-79% nucleotide and 95% amino acid identities for S genome, 70-78% nucleotide and 87-92% amino acid identities for M genome). The recombinant nucleocapsid protein of Tobetsu genotype was antigenically quite similar with that of Sotkamo. These suggest that the virus endemic in Japanese C. rufocanus belongs to Puumala virus. Phylogenetic analysis indicates that the genotype forms a distinct lineage within Puumala viruses. Partial S segment (1-1251 nt), derived from seropositive M. fortis in Vladivostok, was sequenced and analyzed. The S genome segment, which was designated Vladivostok genotype, was most closely related with Khabarovsk virus (79% nucleotide and 90% amino acid identities) which was isolated from M. fortis. (C) 1999 Elsevier Science B.V. All rights reserved. - M Ogino, K Yoshimatsu, H Ebihara, J Arikawa ARCHIVES OF VIROLOGY 144 (9) 1765 -1777 1999年 [査読無し][通常論文]
N-acetylgalactosamine (GalNAc)-specific lectins, Dolichos biflorus agglutinin (DBA), and soybean agglutinin (SBA), enhanced Hantaan (HTN) virus infections in Vero E6 and P388D1 cells. Treatment of Vero E6 cells with the lectins either before or during, but not after, virus inoculation resulted in lectin-mediated enhancement of infection (LME), indicating that GalNAc-specific lectin affects an early stage of the infection. Lectin blot and FAGS analysis showed that the ability of HTN virus envelope glycoproteins and cell surface molecules to bind DBA and SEA was essential for LME. GalNAc clearly inhibited LME, indicating that the lectins bind with their specific carbohydrate-binding site. These results suggest that a lectin cross-link between the virus and the cell surface is the most plausible mechanism for inducing infection enhancement. - M Ogino, K Yoshimatsu, K Tsujimura, T Mizutani, J Arikawa, Takashima, I JOURNAL OF VETERINARY MEDICAL SCIENCE 60 (4) 531 -534 1998年04月 [査読無し][通常論文]
We produced two recombinant Borna disease virus (BDV) proteins, p40 and p24, by using a baculovirus vector as a diagnostic antigen; Antigenicities of these recombinant proteins were evaluated by immune rabbit sera; Recombinant p40 was a more sensitive antigen than p24 for the detection of antibodies in infected rats. Rats inoculated with BDV within 24 hi after birth showed higher detection rates of viral RNA and viral proteins from the brain than rats inoculated at 4 weeks-old. Depending on the age of infection and the time postinfection, the detection of BDV RNA, protein, or anti-BDV antibody did not always correlate in individuals, We suggest both serological and molecular biological methods are needed in the diagnosis of BDV infection. - Antigenic characterization of Hantaan and Seoul virus nucleocapsid proteins expressed by recombinant baculovirus. application of a truncated protein, lacking an antigenic region common to the two virus."jointly worked"Journal of clinical Microbiology 36(9 ),2514-2521 1998年 [査読無し][通常論文]
- YC Yoo, K Yoshimatsu, Y Koike, R Hatsuse, K Yamanishi, O Tanishita, J Arikawa, Azuma, I VACCINE 16 (2-3) 216 -224 1998年01月 [査読無し][通常論文]
The adjuvant effect of two lipophilic derivatives of muramyl dipeptide (MDP), B30-MDP and MDP-Lys(L18), on the ability of an inactivated vaccine of B-I virus (B-l vaccine) to induce immune response against Hantavirus causing hemorrhagic fever with renal syndrome (HFRS) was examined, When mice were immunized subcutaneously (s.c.) twice at 2-week intervals with B-I vaccine admixed with or without 100 mu g mouse(-1) of B30-MDP (B-1/B30-MDP) or MDP-Lys(L18) [B-1/MDP-Lys(L18)], mice immunized with B-1/B30-MDP as well as B-1/MDP-Lys(L18) showed Significantly higher indirect fluorescent antibody (IFA) titers against HFRS virus than mice immunized with B-I vaccine alone, Both mice treated with B-1/B30-MDP and B-1/MDP-Lys(L18) also exhibited significantly higher neutralizing antibody titers against HFRS virus than mice immunized with B-I ,vaccine alone during 3-9 weeks after-the primary immunization. The evaluation of antibody-producing cells by enzyme-linked immunospot (ELISPOT) assay on week 4 revealed that both MDP derivatives,es enhanced the number of HFRS virus-specific IgG1 and IgM antibody-producing cells. Furthermore, mice treated with B-1/B30-MDP as well as B-1/MDP-Lys(L18) showed a higher level of Th-2 type cytokines, IL-4 and IL-6, in sera than mice treated with B-l alone. In an in-vitro analysis of T lymphocyte proliferation to baculovirus-expressed recombinant nucleocapsid pi protein (rNP) of Hantaan 76-118 sa strain, the splenocytes of mice treated with B-1/B30-MDP and B-1/MDP-Lys(L18) on week 4 showed a Significantly higher proliferating activity than those treated with B-1 vaccine alone, In addition when mice were immunized once with B-l vaccine admixed with or without B30-MDP and MDP-Lys(L18) and followed by intrafootpad (i.f.) injection of B-I vaccine on day 7, mice immunized with B-1/B30-MDP and B-1/MDP-Lys(LIS) induced a higher delayed-type hypersensitivity (DTH) reaction than mice immunized with B-I vaccine alone, These results suggest that B30-MDP and MDP-Lys(L18) are useful immunoadjuvants to enhance the ability of inactivated B-l vaccine to induce a humoral and cellular response to HFRS virus, (C) 1997 Elsevier Science Ltd, All rights reserved. - M Kikuchi, K Yoshimatsu, J Arikawa, R Yoshida, YC Yoo, Y Isegawa, K Yamanishi, S Tono-oka, Azuma, I ARCHIVES OF VIROLOGY 143 (1) 73 -83 1998年 [査読無し][通常論文]
Neutralizing monoclonal antibody (MAb) escape mutants of Hantaan virus were generated using MAbs to envelope protein G1 (16D2) and G2 (11E10). The mutant viruses (mu16D2 and mu11E10), lacked reactivity only to the selecting MAb, or a MAb belonging to the same antigenic site. Both mutants had a single amino acid (a.a.) substitution. The a.a. substitution, found in mu16D2, was different from that found in another mutant selected with the same MAb (16D2). Although MAb 11E10 immunoprecipitated G2 protein, a deduced a.a. substitution was located in the G1 region. These results suggest that antigenic sites defined by neutralizing MAbs are composed of discontinuous epitopes over the G1 and G2 proteins. Mutant 11E10 showed a significant decrease in virulence in suckling mice. A virulence revertant of mu11E10, selected through passages in suckling mice brain, showed exactly the same deduced a.a. sequence as mu11E10 and still was not neutralized by MAb 11E10. Since mutant 16D2 was virulent for suckling mice, neutralization related epitopes found with MAbs 11E10 and 16D2 were independent of pathogenicity in BALB/c mice. - H Kariwa, M Fujiki, K Yoshimatsu, J Arikawa, Takashima, I, N Hashimoto ARCHIVES OF VIROLOGY 143 (1) 15 -24 1998年 [査読無し][通常論文]
To understand the mode of transmission of Seoul type hantavirus in Wistar rats, we examined the shedding of the virus and antibody production in infected rats. When 1-day-old rats were inoculated with the KI-83-262 strain of Seoul virus, S segment of the viral genome was detected in lungs, clots, urine, saliva, submaxillary glands, rectums, and kidneys by nested reverse transcriptase PCR. On the other hand, when 8-week-old rats were infected with the virus, viral genome was detected only in the lungs and rectum. In newborn rats intranasally administered urine from infected newborn rats, four of six rats shed the virus into their urine. In addition, three of eight rats kept in the same cage with infected animals also shed the virus into urine. Moreover, the virus genome was detected in the urine of urban rats (Rattus norvegicus) in an enzootic focus. These findings suggest that the urine containing virus from infected rats is an actual source of the Seoul virus infection. - K Yoshimatsu, J Arikawa, S Ohbora, C Itakura JOURNAL OF VETERINARY MEDICAL SCIENCE 59 (10) 863 -868 1997年10月 [査読無し][通常論文]
Severe combined immunodeficiency (SCID) mice were inoculated with Hantaan virus strain 76-118 (HTN) or Seoul virus strain SR-11 (SR) of hantaviruses. Susceptibility of SCID mice was compared with those of immunocompetent adult mice, newborn mice and nude mice. SCID mice inoculated with HTN or SR died 32 to 35 days after infection. Unlike newborn mice which also died of hantavirus infection, SCID mice survived longer than newborn mice and showed typical wasting symptoms rather than nervous symptoms. Immunohistochemical staining and virus isolation indicated that both HTN and SR inoculated SCID and SR inoculated nude mice showed systemic infection, but nude mice inoculated with SR survived for longer than 8 weeks after inoculation. Passive transfer of spleen cells from immunocompetent BALB/c mice conferred protection on SCID mice within 2 weeks of HTN infection. Immune mediated pathologic mechanism was examined by transferring the spleen cells to SCID mice inoculated with HTN virus 3 weeks before the cell transfer. The recipient SCID mice showed an increase of serum BUN level coinciding with the appearance of serum antibody to HTN virus, suggesting the immune mediated pathogenicity. - Takashima, I, K Morita, M Chiba, D Hayasaka, T Sato, C Takezawa, A Igarashi, H Kariwa, K Yoshimatsu, J Arikawa, N Hashimoto JOURNAL OF CLINICAL MICROBIOLOGY 35 (8) 1943 -1947 1997年08月 [査読無し][通常論文]
A case of tick-borne encephalitis (TEE) has not been reported for many years in Japan, although a serological survey of sera from domestic animals suggested the presence of TEE foci in Hokkaido, the northern island of Japan. Studies were conducted to prove the presence of an endemic focus of TEE virus in Japan by means of serology and virus isolation. In October 1993 in Hokkaido, a severe case of encephalitis in a dairy farmer's nife was diagnosed as TEE. Serological examination of paired serum specimens showed a rise in the neutralization antibody titer to Russian spring summer encephalitis virus. A seroepizootiological survey of dogs showed that the TEE-related virus was prevalent in the area. Three virus isolates were obtained from the blood of sentinel dogs, and antigenic analysis grouped the isolates into TEE-related viruses. Sequence analysis of the envelope protein gene identified one of the isolates as being of the same subtype as the Russian spring summer encephalitis (Far Eastern TEE) virus. The results provide evidence that TEE is endemic in a certain area of Japan. - Kaori Matsuzawa, Yung Choon Yoo, Akihisa Fukushima, Kumiko Yoshimatsu, Jiro Arikawa, Ichiro Azuma. Collected papers from the Institute of Immunological Science Hokkaido University 20 171 -175 1997年
- K Matsuzawa, YC Yoo, A Fukushima, K Yoshimatsu, J Arikawa, Azuma, I VACCINE 15 (1) 85 -89 1997年01月 [査読無し][通常論文]
We investigated the protection confered by the mucosal administration of recombinant human macrophage colony-stimulating factor (rhM-CSF) against mucosal infection of Sendai virus in mice. In an experimental infection model using Sendai virus, an intranasal (i.n.) administration of rhM-CSF (20 mu g per mouse) 2 days before infection induced significant protection against a lethal infection of this virus. Also, its antiviral activity was dependent upon the dose of rhM-CSF. However; a subcutaneous (s.c.) administration of rhM-CSF with an effective close (20 mu g per mouse) i.n. did not confer protection. In a time course analysis of virus growth in the lungs, mice given rhM-CSF in. significantly inhibited the early period of infection, compared with the untreated mice. Moreover, the level of interferon-gamma (IFN-gamma) in lung wash fluids from the rhM-CSF-treated mice was higher than that of the untreated mice. These results suggested that the mucosal (i.n.), but not the systemic (s.c.) administration of rhM-CSF augments host resistance against mucosal infection with Sendai virus, and that its prophylactic activity is related to growth inhibition of the virus and enhanced IFN-gamma secretion in the lungs. Copyright (C) 1997 Elsevier Science Ltd. - A Fukushima, YC Yoo, K Yoshimatsu, K Matsuzawa, M Tamura, S Tonooka, K Taniguchi, S Urasawa, J Arikawa, Azuma, I VACCINE 14 (6) 485 -491 1996年04月 [査読無し][通常論文]
To examine the effect of MDP-Lys(L18), a derivative of muramyl dipeptide (MDP), as a mucosal immunoadjuvant, we investigated its activity to augment host resistance against mucosal infections by Sendai virus and rotavirus in mice. lit an experimental infection model that suckling mice (10-day-old) were inoculated perorally (p.o.) with 1.5 x 10(6) p.f.u. mouse(-1) of rotavirus strain SA11, intrarectal (i.r.) as well as p.o., administration of MDP-Lys(L18) (50 mu g mouse(-1)) prior to virus infection markedly reduced rotavirus-induced diarrhea. Furthermore, when MDP-Lys(L18) was administered p.o. (1 mg mouse(-1)), i.r. (300 mu g mouse(-1)) or intranasally (i.n., 100 mu g mouse(-1)) various days before Sendai virus infection (2.6 x 10(4) HAD mouse(-1)), all the mucosal administration of MDP-Lys(L18) significantly protected a lethal infection of Sendai virus, showing a dose-dependent manner. However, the efficacy of MDP-Lys(L18) to induce the prophylactic activity against the viruses somewhat varied according to the administration route and timing. In time course analysis of virus isolation in vivo, the mice administered with MDP-Lys(L18) exhibited a significant reduction of both viruses in the lungs for Sendai virus and in the bowels for rotavirus. These results suggest that MDP-Lys(L18) is a potent mucosal immunoadjuvant to enhance nonspecific host resistance against two mucosal infectious viruses, Sendai virus and rotavirus. Copyright (C) 1996 Elsevier Science Ltd. - K Yoshimatsu, J Arikawa, M Tamura, R Yoshida, A Lundkvist, B Niklasson, H Kariwa, Azuma, I JOURNAL OF GENERAL VIROLOGY 77 695 -704 1996年04月 [査読無し][通常論文]
We characterized the antigenic sites on the nucleocapsid protein (NP) of Hantaan virus (HTN) using 10 monoclonal antibodies (MAbs). At least seven antigenic sites were revealed by a competitive binding assay and divided into three partially overlapping antigenic regions (I, II and III). Regions I [amino acids (aa) 1-103], II (aa 104-204) and III (aa 205-402) were mapped on NP by examinincr the reactivity of truncated gene products. Those that corresponded to region I reacted with immune mouse serum, indicating that the region contained major linear epitopes as reported with Four corners virus (FCV) and Puumala virus (PUU) NP. At least one MAb to each region inhibited viral growth when they were introduced into cells by scrape-loading. In addition, they conferred protection from a lethal HTN challenge to newborn mice. A PEPSCAN assay localized the epitope of MAb E5/G6 between aa 166-175. Since E5/G6, which had the highest inhibitory effect both in cells and in mice, showed no virus neutralization activity by ordinary neutralization test, this region is suggested to be important for the virus growth after entry into the cells. - Akihisa Fukushima, Yung Choon Yoo, Kumiko Yoshimatsu, Kaori Matsuzawa, Mizuho Tamura, Shuichi Tono-oka, Koki Taniguchi, Shozo Urasawa, Jiro Arikawa, Ichiro Azuma. Collected papers from the Institute of Immunological Science Hokkaido University 19 153 -159 1996年
- Kumiko Yoshimatsu, Jiro Arikawa, Mizuho Tamura, Ryu Yoshida, Lundkvist, B. Niklasson, H. Kariwa, Ichiro Azuma. Collected papers from the Institute of Immunological Science Hokkaido University 19 143 -152 1996年
- H Kariwa, M Kimura, S Yoshizumi, J Arikawa, K Yoshimatsu, Takashima, I, N Hashimoto ARCHIVES OF VIROLOGY 141 (12) 2327 -2338 1996年 [査読無し][通常論文]
To understand the mode of persistent infection of Seoul virus in rodents, we examined the distribution of the virus genome and antibody production in infected rats. When 1-day-old rats were inoculated with the KI-83-262 strain, the S segment of viral genome was detected in sera, clots, lungs and kidneys from 3 to 184 days post inoculation (d.p.i.) by nested reverse transcriptase PCR. On the other hand, when 7-week-old rats were infected with this virus, viral genome was detected only in the lungs from 3 to 50 d.p.i. The neutralizing antibody titers of rats inoculated at 1-day of age were higher than those of rats inoculated at 7 weeks of age. In both a ge groups, however, the IgG avidity of antibody increased along with the course of infection. We found that urban rats (Rattus norvegicus) infected early in life harbored the virus for more than 6 months. - K Yoshimatsu, J Arikawa, H Li, H Kariwa, N Hashimoto JOURNAL OF VETERINARY MEDICAL SCIENCE 58 (1) 71 -74 1996年01月 [査読無し][通常論文]
Recombinant Hantaan virus nucleocapsid protein expressed in silkworm larvae was applied as a serological diagnostic antigen in Western blots (WE) of human sera. The sensitivity of this method was similar to that of the IFA test. Hemorrhagic fever with renal syndrome (HFRS) and nephropathia epidemica diagnosed by their cross-reactivity in WE. The specificity of this method was higher than that of IFA test because the background was low. Sera that exhibited high background staining in the IFA test were readily diagnosed with this method. We recommended WE using recombinant Hantaan virus nucleocapsid antigen as a confirmatory procedure for the serodiagnosis of hantavirus. - K Yoshimatsu, J Arikawa, R Yoshida, H Li, YC Yoo, H Kariwa LABORATORY ANIMAL SCIENCE 45 (6) 641 -646 1995年12月 [査読無し][通常論文]
The recombinant nucleocapsid protein (rNP) of Hantaan virus was expressed by a baculovirus vector in silkworm hemolymph and was used as an antigen in western blotting (WB). The rNP is expressed in insoluble form in hemolymph; therefore simple washing of the insoluble fraction with phosphate-buffered saline by low-speed centrifugation allowed preparation of purified antigen for WB. The rNP had strain-specific and hantavirus-common epitopes similar to the authentic NP antigen of hantavirus and was stable after transfer to membrane. For detection of antibody in serially obtained sera from experimentally infected rats, WB enabled detection of IgM antibodies 3 days after infection, which was at least 2 days earlier than detection by the indirect immunofluorescent antibody test (IFA). Thus WB had a higher sensitivity than the IFA for detection of hantavirus antibody in the serum of experimentally infected rats. The WB-determined IgG antibody titer was about 10 times higher than that determined by the IFA. No background staining was observed by WB even at a 1:10 dilution of serum. The selected rat sera with strong background staining or confusing staining patterns by LFA, but not focus reduction neutralization test titers, could be interpreted as test-negative because they did not have a specific reaction to virus antigen by WB. Thus the specificity of WB was higher than that of the IFA. Moreover, WB can distinguish specific from nonspecific reactions by the detection of the specific antigen on the WB membrane. Therefore the IFA or enzyme-linked immunosorbent assay followed by WB is recommended for serologic confirmation of hantavirus infection. - YC YOO, K YOSHIMATSU, R HATSUSE, M TAMURA, R YOSHIDA, S TONOOKA, J ARIKAWA, AZUMA, I VACCINE 13 (14) 1300 -1305 1995年10月 [査読無し][通常論文]
We examined the effect of MDP-Lys(L18), a lipophilic derivative of muramyl dipeptide, on the enhancement of host resistance against virus infection in newborn mice Newborn mice were inoculated with 4 LD(50)/mouse of Hantaan virus strain 76-118 (HTN) one day after birth. Mice given 100 mu g/mouse of MDP-Lys(L18) before infection exhibited significantly higher survival rates than those of non-treated mice. The effect of MDP-Lys(L18) was also restorative when given to the mice 4 or 7 days after infection. The titers of virus isolated from the lungs and spleens 12 days after infection, were about 30-times lower in MDP-Lys(L18)-treated (lung: 1.0 x 10(3) FFU; spleen: 6.8 x 10(1) FFU/mouse), than those of non-treated mice (lung: 3.4 x 10(4) FFU; spleen: 1.9 x 10(3) FFU/mouse). Furthermore the virus was undetectable in the brains of MDP-Lys(L18)-treated mice, whereas viruses were isolated from 3 of 6 non-treated mice MDP-Lys(L18) augmented the number of peripheral leukocytes and splenocytes, as well as mitogenic responses of the cells from bone marrow and spleen of newborn mice. These results suggest that MDP-Lys(L18) enhanced the resistance of newborn mice against HTN virus in a systemic infection model, and that this mechanism is involved in the enhancement of hematopoiesis and responsiveness of immune-related cells to mitogens. - EVIDENCE FOR THE EXISTENCE OF PUUMULA-RELATED VIRUS AMONG CLETHRIONOMYS RUFOCANUS IN HOKKAIDO, JAPANH KARIWA, S YOSHIZUMI, J ARIKAWA, K YOSHIMATSU, K TAKAHASHI, TAKASHIMA, I, N HASHIMOTO AMERICAN JOURNAL OF TROPICAL MEDICINE AND HYGIENE 53 (3) 222 -227 1995年09月 [査読無し][通常論文]
We conducted field surveys of indigenous rodent species in Hokkaido, Japan from 1980 to 1993. Serum samples were collected from 663 rodents, including Clethrionomys rufocanus, Apodemus speciosus, A. argenteus, and C. rutilus. Antibody to hantavirus was determined by the protein G antibody assay. Positive C. rufocanus were detected in seven of eight collection sites, but no antibody was detected in the remaining rodent species. To reveal the serotype of the circulating virus in C. rufocanus, antibody titers to Hantaan, Seoul, Puumala, and Prospect Hill viruses were compared by means of the focus reduction neutralization test. The titers in positive sera were extremely high to the Sotkamo strain of Puumala virus. Results were confirmed by the reverse transcriptase-polymerase chain reaction, and suggested that Puumala-related viruses are in circulation among C. rufocanus populations in Hokkaido. - Mizuho Tamura, Yung Choon Yoo, Kumiko Yoshimatsu, Ryu Yoshida, Tetsuya Oka, Kunio Ohkuma, Jiro arikawa, Ichiro Azuma Collected papers from the Institute of Immunological Science Hokkaido University 18 155 -160 1995年
- Yung Choon Yoo, Kumiko Yoshimatsu, Rei Hatsuse, Mizuho Tamura, Ryu Yoshida, Shuichi Tono-oka, Jiro Arikawa, Ichiro Azuma. Collected papers from the Institute of Immunological Science Hokkaido University 18 203 -208 1995年
- M TAMURA, YC YOO, K YOSHIMATSU, R YOSHIDA, T OKA, K OHKUMA, J ARIKAWA, AZUMA, I VACCINE 13 (1) 77 -82 1995年01月 [査読無し][通常論文]
The ability of two muramyl dipeptide (MDP) derivatives, B30-MDP and MDP-Lys (L18), to enhance the immunogenicity of recombinant hepatitis B surface antigen (rHBsAg) was examined. When mice were immunized intraperitoneally with rHBsAg together with each MDP derivative, the antibody titres were higher than those in mice immunized with alum-adsorbed rHBsAg, which is a commercially available hepatitis B vaccine. When mice were given a subcutaneous or intramuscular injection of rHBsAg and either MDP derivative, the antibody titres were the same as those in mice given alum-adsorbed rHBsAg. These results indicate the usefulness of MDP derivatives as immunoadjuvants for a new-generation vaccine. - H KARIWA, M KAMIMURA, J ARIKAWA, K YOSHIMATSU, TAKASHIMA, I, N HASHIMOTO MICROBIOLOGY AND IMMUNOLOGY 39 (1) 35 -41 1995年 [査読無し][通常論文]
A polymerase chain reaction (PCR) for the detection of hantavirus genome was established and applied to analyze the mode of infection of Hantaan virus in adult ICR mice. The cDNA for the S genome segment of Hantaan virus was reverse-transcribed from the total RNA of organs of the infected mice. The sequence in the S genome segment of Hantaan virus was successfully amplified by reverse transcriptase (RT)-PCR followed by nested PCR. In 5-week-old ICR mice inoculated intraperitoneally with Hantaan virus, strain 76-118 (1.3 x 10(5) FFU/mouse), the virus was detected in clots and lungs from 3 to 10 days post-inoculation (p.i.) by nested PCR and virus-isolation techniques. No virus was detected in any specimens collected on 1 day and after 28 days p.i., and in spleens and brains through the observation period by both methods, The antibody which was measured by indirect immunofluorescence antibody assay (IFA) appeared at 7 days p.i. and the geometric mean titer was elevated to its maximum level of 1 : 203 at 10 days p.i., maintaining the same level until 35 days p.i. These results suggest that adult mice are transiently infected with Hantaan virus. - R YOSHIDA, K SATO, YC YOO, K YOSHIMATSU, M TAMURA, C ISHIHARA, J ARIKAWA, AZUMA, I IMMUNOPHARMACOLOGY 28 (2) 153 -161 1994年09月 [査読無し][通常論文]
DT-5461 enhanced host resistance to Sendai virus infection in mice. Intranasal (i.n.) administration of 200 mu g of DT-5461 per mouse 3 days before infection was the most effective administration route, dose and timing. DT-5461 enhanced the cytotoxicity of murine natural killer (NK) cells. In addition, DT-5461 activated murine peritoneal macrophages, resulted in augmented of cytotoxicity and the induction of tumor necrosis factor-alpha (TNF-alpha). Therefore, these immunomodulating activities presented by DT-5461 caused protection against Sendai virus infection. - Y YUYAMA, K YOSHIMATSU, E ONO, M SAITO, M NAIKI JOURNAL OF BIOCHEMISTRY 113 (4) 488 -492 1993年04月 [査読無し][通常論文]
Enterotoxigenic Escherichia coli possessing K99 fimbriae (E. coli K99) causes diarrhea in piglets of less than 1 week old. The first stage of the bacterial infection is adhesion by the fimbriae on the small intestinal mucosa and the adhesion is followed by colony formation. K99 fimbriae bind specifically to N-glycolylneuraminyl-lactosyl-ceramide, GM3(NeuGc) [Ono, E. et al. (1989) Infect. Immun. 57, 907-911]. We examined the postnatal change of the content and the molecular species of GM3(NeuGc) in the small intestinal mucosa of 0- to 14-day-old piglets and adult pigs. GM3(NeuGc) was a major ganglioside of piglet intestinal mucosa. GM3(NeuGc) content was maximal at birth and gradually decreased to 1/16 in adult animals (5 months old). The ceramide moiety of piglet intestinal GM3(NeuGc) was characterized by the presence of 2-hydroxylated palmitic acid. I-125-labeled bacteria strongly bound to GM3(NeuGc) containing 2-hydroxylated palmitic acid and phytosphingosine compared with GM3(NeuGc) containing any other ceramide moiety. The time when this particular GM3(NeuGc) appears coincides with the time that the infection occurs, and it may explain the susceptibility of newborn piglets to E. coli K99 infection. - C ISHIHARA, K YOSHIMATSU, M TSUJI, J ARIKAWA, SAIKI, I, S TOKURA, AZUMA, I VACCINE 11 (6) 670 -674 1993年04月 [査読無し][通常論文]
Sulfated chitin derivatives, selected for their low toxicity and high inhibitory activity of melanoma metastasis, were examined for anti-viral activity against Friend murine leukaemia, herpes simplex type-1 (HSV) and Sendai viruses. Carboxymethyl chitin with a 7.66% degree of sulfation (SCM-chitin III) showed a significant inhibition of Friend murine leukaemia helper virus (F-MuLV) and HSV, but not of Sendai virus growth in vitro. Sulfated N-deacetylated chitin had a significant but weak activity against F-MuLV and HSV infections. Carboxymethyl chitin showed no effect on these infections in vitro. SCM-chitin III also exhibited anti-viral activity in vivo by suppressing the splenomegaly which was caused by prior infection of mice with FV, a complex of F-MuLV and spleen focus-forming virus. - Journal of Veterinary Medical Science 55(6),1047-1050 1993年 [査読無し][通常論文]
- YC YOO, K YOSHIMATSU, R YOSHIDA, M TAMURA, AZUMA, I, J ARIKAWA MICROBIOLOGY AND IMMUNOLOGY 37 (7) 557 -562 1993年 [査読無し][通常論文]
Virulence of hantavirus strain of SR-11 Seoul virus and Hantaan 76-118 (HTN) of Hantaan virus were compared. Infections of both strains were lethal in newborn mice. However, inoculum required to cause lethal infection was about 4,000 times higher for strain HTN (1.65 x 10(3) PFU/mouse/LD50) than for strain SR-11 (0.36 PFU). Thus, both strains were considered pathogenic to newborn mice but they possessed different levels of virulence. The assay system used for these strains in newborn mice proved to be useful in the study of hantavirus vilurence. Growth curves of the two strains in CV-7 cell cultures were compared. Strain SR-11 was shown to have higher activity of virus replication and virus release into the culture fluids than strain HTN. The possibility of a relationship between replication activity and high levels of virulence in mice was suggested. - K YOSHIMATSU, YC YOO, R YOSHIDA, C ISHIHARA, AZUMA, I, J ARIKAWA ARCHIVES OF VIROLOGY 130 (3-4) 365 -376 1993年 [査読無し][通常論文]
Recombinant Hantaan virus nucleocapsid protein (rNP) and recombinant envelope (rEnv) proteins were prepared using a baculovirus expression system to examine the role of Hantaan virus structural proteins in protective immunity. Passive transfer of spleen cells from mice immunized with rNP conferred partial protection or prolongation of time to death from fatal Hantaan virus infection in suckling mice which were challenged with Hantaan virus at 40 LD50 (survival rate: 43%) or 4 LD50 (survival rate: 43%). The T cell-enriched fraction protected one mouse from lethal infection but the B cell-enriched fraction had no such effect on fatal HTN infection. The protective effects of the antibody against HTN challenge were examined by passive immunization. The monoclonal antibody ECO 2 directed to NP also conferred partial survival and significant difference in time to death. Although rEnv antigen failed to induce neutralizing antibody, both immune spleen cells and immune serum to rEnv conferred partial protection upon suckling mice. These results indicate that both nucleocapsid and envelope proteins of Hantaan virus were responsible for induction of cell mediated protective immunity. Vero E 6 cells infected with Hantaan virus expressed envelope protein on the surface, as determined by flow cytometry. However, there was only negligible expression of nucleocapsid protein. - K YOSHIMATSU, Y YUYAMA, E ONO, B SYUTO, M NAIKI JOURNAL OF VETERINARY MEDICAL SCIENCE 53 (6) 1119 -1121 1991年12月 [査読無し][通常論文]
書籍等出版物
担当経験のある科目（授業）
所属学協会
Works（作品等）
- スリランカにおける原因不明慢性腎臓病(CKDu)とハンタウイルス感染症の関連に関する研究森松(吉松) 組子 2016年 - 現在
- 腎症候性出血熱とハンタウイルス肺症候群の中和代替試験法による鑑別診断法の開発1996年 -2008年
- 組換え可溶性外皮蛋白を用いたハンタウイルスワクチンおよび迅速診断キットの開発2001年 -2003年
- 自然免疫情報伝達系による生体防御と胎児発生の制御～新規NFкB抑制タンパク質ファミリー分子欠損マウスの解析を中心に2003年
- モノクローナル抗体および細胞内抗体導入（スクレイプローディング）法を用いたハンタウイルス核蛋白の機能の解析1995年 -1996年
- 蚕バキュロウイルス発現ハンタウイルス核蛋白の精製法の確立と診断法への応用1993年 -1993年
- 免疫不全マウス(SCID、ヌード)での腎症候性出血熱ウイルスによる腎症候性出血熱障害の解析1992年 -1992年
- 実験マウスにおける組換え腎症候性出血熱ウイルス蛋白を用いた免疫応答の解析－パキュロウイルスベクター発現系の新しい応用1991年 -1991年
共同研究・競争的資金等の研究課題
- 日本学術振興会：科学研究費助成事業 基盤研究(C)研究期間 : 2020年04月 -2023年03月代表者 : 土佐 紀子, 林元 展人, 石田 智子, 森松 組子学術・科学技術の研究に供する動物の微生物モニタリングは、研究結果の信頼性の担保、および動物福祉の観点から必要不可欠である。近年、多系統少数個体維持の増加や個別換気型飼育機の普及等により、従来の微生物モニタリング方法、すなわち「おとり動物」の感染症の検出を指標とする間接的な方法では感染症を摘発することが困難となっている。この問題を解決するためには、動物を安楽死させることなく微量全血を用い、マウス、ラットの主要感染症を同時に且つ迅速・簡便に検出できる個体別血清診断法の確立が必要である。その方法として有望と考えられる「多項目イムノクロマト(ICG)法」の開発について申請者らは取り組んで来た。 本研究ではこれまでの成果を基に、１）多項目ICG法の検出条件が確定したマウス・ラットの感染症（マウス：マウス肝炎ウイルス感染症、センダイウイルス感染症、ティザー病。ラット：唾液腺涙腺炎ウイルス感染症、センダイウイルス感染症、腎症候性出血熱）の検出における本法の実用性を検証し、２）上記感染症に加え、主要感染症であるマイコプラズマ感染症（マウス、ラット）とティザー病（ラット）のICG法を確立することを目的としている。２年度（令和３年度）においては、以下の研究を実施した。 １．多項目ICGスティックの最適な保管条件・最長保管期間を明らかにする実験において、保管期間６ヶ月および１２ヶ月の多項目ICGスティックの各保管条件における検出・感度特異性への影響の結果を得た。 ２．乾燥血液を用いた時のICG法の検出・感度特異性を明らかにするため、乾燥血液を作製するための条件を検討した。 ３．多項目ICGスティックの輸送条件を確定するための実験設備の準備を行った。 ４．Myco（マウス、ラット）とTyzzer（ラット）のICG法を確立するため、抗原の準備を行い、ICGメンブレンに塗布する抗原処理方法の実験に着手した。
- 日本学術振興会：科学研究費助成事業 基盤研究(C)研究期間 : 2019年04月 -2022年03月代表者 : 津田 祥美, 森松 組子本課題ではエボラウイルスの主要標的細胞とされているマクロファージや肝細胞でのウイルス増殖がエボラウイルス感染症で観察される病態にどのように関与しているのか、ウイルスの組織指向性や主要標的細胞での増殖能力と致死的病態の関連性を明らかにすることを目的とする。これまでの解析によりマクロファージや樹状細胞で主に発現しているmicroRNAのターゲット配列をウイルス遺伝子に組込むことでマクロファージや樹状細胞でのウイルス増殖を抑制した組換えエボラウイルス作出可能であることを示した。また作出したウイルスは著しく病原性が減弱したことから、感染初期におけるマクロファージや樹状細胞でのウイルス増殖の重要性を示した。本研究ではクロファージや樹状細胞で増殖したウイルスが次にターゲットとし、重要な宿主応答に関与していると考えられる肝細胞でのウイルス増殖を抑制したウイルスの作成を試みた。これまでのC型肝炎ウイルスなどの研究により、肝臓で発現するいくつかのmicroRNAの重要性が報告されている。そこで本研究ではmicroRNA122をまずターゲットとして選び、microRNA122に対応する遺伝子配列を組み込んだ組換えウイルスの作出を試みた。これまでの研究と同様にマウスを用いた病原性解析を目的として、親株であるエボラウイルスとマウス馴化株に遺伝子組換えを行ったところ、ともにウイルスの作出を確認した。今後は作出したウイルスの細胞での増殖、マウスモデルを用いた病原性の解析を行う予定である。
- 日本学術振興会：科学研究費助成事業 基盤研究(C)研究期間 : 2019年04月 -2022年03月代表者 : 森松 組子, 清水 健太, 津田 祥美慢性腎臓病(CKD)は2002年に米国腎臓財団によって提唱された概念である。先進国では成人人口の13％を占めるとされ、そのほとんどが糖尿病や高血圧等の生活習慣病を原疾患とする。一方で、このような原疾患を持たない原因不明のCKDがスリランカ、インドの一部、中南米から報告されており、CKDuと呼ばれている。特にスリランカでは重症型のCKDuが健康な労働年齢の男性に頻発し、国家的な問題となっている。近年申請者らは、CKDu患者の約半数が抗ハンタウイルス抗体陽性であることを見いだし、ハンタウイルス感染症が慢性腎臓病の発症に関与していることを疫学的解析により示した。さらに、CKDuの流行地のげっ歯類からハンタウイルスを検出し、このウイルスが感染源となっている可能性をみいだした。本研究では、スリランカ固有の新規ハンタウイルスを解析し、この情報を基礎に血清診断法、分子生物学的診断法を開発する。さらにこれらの診断法を用いて、ヒトおよびげっ歯類におけるハンタウイルスの感染状況およびCKDu発症機序の解明を試みることを最終的な目的とする。 今年度は特に、スリランカ固有のハンタウイルスの特定と遺伝子解析を目標としていた。結果的に、存在がすでに確認されていたランカウイルスに加えてもう一つのタイランド型ハンタウイルスであるアンゾロベウイルスを見いだし、それぞれの宿主がインドハツカネズミおよびクマネズミであることを明らかにした。さらにそれぞれのウイルスの遺伝子配列を決定し、ランカウイルスは最も進化的に離れたタイランド型ハンタウイルスであること、クマネズミ由来のウイルスはマダガスカルに棲息するクマネズミ由来のアンゾロベウイルスに極めて近縁であることが明らかとなった。また、遺伝子配列を基に血清診断法および遺伝子診断法の開発を行った。
- スリランカに頻発する原因不明慢性腎臓病に関連するハンタウイルスの分子生物学的解析日本医療研究開発機構：日米医学協力計画若手・女性育成のための日米共同研究研究期間 : 2018年04月 -2019年03月代表者 : 森松(吉松) 組子
- 重症熱性血小板減少症候群（SF TS）に対する診断・治療・予防 法の開発厚生労働省：感染症実用化研究事業研究期間 : 2016年04月 -2019年03月代表者 : 西條 政幸
- 日本学術振興会：科学研究費助成事業 基盤研究(C)研究期間 : 2015年04月 -2018年03月代表者 : 森松 組子レプトスピラおよびハンタウイルス感染の様式と宿主を明らかにすることを目的とした。ラットおよびトガリネズミから病原性レプトスピラが分離された。抗ハンタウイルス血清抗体保有率は水田地帯で19%と高く、スリランカ固有クマネズミがタイランド型関連ハンタウイルスの宿主であると考えられた。全島横断的なヒトの抗体調査の結果、原因不明慢性腎臓病患者(CKDu)の約50%がハンタウイルス抗体陽性であり、一方CKDu非流行地の健常人では5%であった。 本研究より、スリランカでは固有種のクマネズミを感染源として、レプトスピラおよびハンタウイルス感染が高頻度で起こり、さらにCKDu発症のリスクとなる可能性が示された。
- スリランカにおけるレプトスピラ症とハンタウイルス感 染症の媒介げっ歯類に関する研究文部科学省：科学研究費補助金(基盤研究(C))研究期間 : 2015年 -2018年03月代表者 : 森松(吉松) 組子
- 日本学術振興会：科学研究費助成事業 基盤研究(B)研究期間 : 2013年04月 -2017年03月代表者 : 苅和 宏明, 有川 二郎, 好井 健太朗, 森松 組子本研究では、近年国内外で問題となっているウイルス性人獣共通感染症のうち、重篤化しやすく致死率も高いハンタウイルス感染症、ダニ媒介性脳炎、およびウエストナイル熱について簡便な診断法の開発を試みた。ハンタウイルスの核蛋白質抗原とし、様々なげっ歯類の抗体検出が可能なELISAとイムノクロマトグラフィー法を開発した。IgGもしくはStrep-tagを付加したダニ媒介性脳炎ウイルスのウイルス様粒子 (SPs) を用いた抗体検出用のELISAを開発した。相同組換えを利用した、安定性の高いウエストナイルウイルスのリバースジェネティック法を開発した。
- 日本学術振興会：科学研究費助成事業 基盤研究(C)研究期間 : 2011年 -2013年代表者 : 森松 組子, 有川 二郎, 清水 健太これまでに申請者は東南アジアおよび南アジア諸国においてハンタウイルス関連疾患に関する調査を行ってきた。その結果、多くの不明熱症例は未診断のまま残されていることが明らかとなった。本研究では南アジア・東南アジアにおける熱性疾患の実態を明らかとすることを目的とした。予め、デング熱およびインフルエンザ患者ではないことが明らかな熱性疾患患者血清およびキャリアとなるげっ歯類の調査をベトナムおよびスリランカにおいて実施した。その結果、特定に至らない病原体の一部として、E型肝炎、レプトスピラ、およびハンタウイルスが含まれることが示唆された。げっ歯類は上記病原体を高率に保有し、感染源となりうることが示された。
- 日本学術振興会：科学研究費助成事業 基盤研究(B)研究期間 : 2009年 -2012年代表者 : 苅和 宏明, 有川 二郎, 好井 健太朗, 森松 組子, 高島 郁夫メキシコのげっ歯類から3種類の新規ハンタウイルスを検出した。各種ハンタウイルス感染の検査に応用可能な抗体検出法を開発した。メキシコのハンタウイルスのヌクレオキャプシド(NP)に対して作出されたモノクローナル抗体6種類のうち5種類はげっ歯類を宿主とするハンタウイルスに対して広く反応した。ウエストナイルウイルスのE蛋白上の糖鎖の有無がニワトリのヒナにおける病原性発現に大きな影響を与えることが判明した。
- 日本学術振興会：科学研究費助成事業 基盤研究(B)研究期間 : 2006年 -2008年代表者 : 有川 二郎, 森松 組子, 遠藤 理香, 苅和 宏明, 森松 組子, 苅和 宏明, 垂石 みどりハンタウイルスは持続感染齧歯類が感染源となる人獣共通感染症でヒトに腎症候性出血熱やハンタウイルス肺症候群という重篤な疾病を引き起こす。齧歯類は血中に高い中和抗体を保有しつつ不顕性にウイルスを排泄し続けるというきわめて特徴的な持続感染を成立させている。本研究では齧歯類におけるウイルス特異的CTL抑制を中心とする免疫抑制を介したハンタウイルス持続感染成立機構の詳細を明らかにすることを目的とする。
- 文部科学省：科学研究費補助金(基盤研究(B))研究期間 : 2004年 -2007年代表者 : 苅和 宏明, 高島 郁夫, 有川 二郎, 吉松 組子本研究では、日本国内に侵入または流行の危険性の高いウエストナイル熱とハンタウイルス感染症について、診断法を開発するとともに、疫学調査を実施した。1)ウエストナイル熱の診断法開発:ウエストナイルウイルス感染の日本への侵入を監視するため、抗ウエストナイルウイルス抗体の検出法として微量中和法や阻害ELISA法を開発した。2)ロシア国内で捕獲された鳥類におけるウエストナイルウイルスの感染調査:ロシアウラジオストック近郊のハンカ湖とハバロフスク北部のアニュイ川河畔で捕獲された98例の鳥類の腎臓からReal-time PCRによりウエストナイルウイルスの遺伝子検出を行ったが、いずれの検体からもウイルス遺伝子は検出されなかった。しかし、微量中和法により血清の得られた91例中うち15例から抗ウエストナイルウイルス抗体が検出され、極東ロシアで野鳥においてウエストナイルウイルスの感染のあることが示唆された。極東ロシアからは日本に渡り鳥が多数飛来することから、今後わが国におけるウエストナイル熱の防疫体制の強化が強く望まれる。3)メキシコのげっ歯類におけるハンタウイルス感染症の疫学調査:北アメリカ産ハンタウイルスのS遺伝子を標的としたRT-PCRによる遺伝子検出法を開発した。そこでメキシコ国内で捕獲されたげっ歯類213例の肺からRNAを抽出し、RT-PCRによりハンタウイルス遺伝子の検出を試みたとこ...
- Studies on hantavirus nucleocapsid protein and envelope glycoproteinsGrant-in-Aid for Scientific Research研究期間 : 2001年 -2007年
- 日本学術振興会：科学研究費助成事業 基盤研究(C)研究期間 : 2005年 -2006年代表者 : 森松 組子ハンタウイルスは腎症候性出血熱(HFRS)とハンタウイルス肺症候(HPS)の原因ウイルスである。不顕性に持続感染したげっ歯類を媒介動物とする重要な人獣共通感染症である。申請者はハンタウイルスエンベロープ蛋白が細胞融合能を持つことを報告し、さらに細胞融合活性に関する部位は中和抗体のエピトープに対応することを発見した。これはハンタウイルスの中和の本質が細胞融合阻止であることを示すワクチン作成に貢献する重要な情報である。本研究では感染成立におけるハンタウイルスエンベロープ蛋白の役割について分子レベルで解明することを試みた。現在レセプター候補とされているαVβ3インテグリンヘテロダイマーの発現ベクターを構築し、非感受性細胞MDBK細胞に導入し、感染性への影響を調べたところ、低導入効率のため判定はやや困難であるが、感染性の回復は認められなかった。また、感受性細胞A549のαVインテグリンのRNAiによるKOもまた感染性へ影響しなかった。以上の結果からαVβ3インテグリンの感染成立への関与は補助的なものであると考えられた。次に糖とハンタウイルスエンベロープ蛋白との相互作用を網羅的にスクリーニングするため、感受性細胞から糖脂質画分を抽出し2次元展開後PVDF膜に転写し、GPを外套したシュードタイプウイルス等をプローブとして結合を検討した。有意な結合が検出されなかったことから、糖鎖が関与する可能性は低いと考えられた。以上の結果から、ハンタウイルスのレセプター分子はおそらくαVβ3インテグリン以外の蛋白質分子であると考えられた。そこで次に、高感受性細胞由来のcDNAライブラリを作製し増幅した。これを導入するためのMDBK細胞をクローニングし、比較的遺伝子導入効率の高いクローンを選別し導入法を決定した。本研究ではレセプター分子の特定には至らなかったものの、基礎研究を完了し、おおむね当初の目的を達成することができた。
- アジアのヒトおよび動物におけるハンタウイルスの血清疫学的および分子疫学的研究科学技術振興調整費による中核的研究拠点（ＣＯＥ）育成研究期間 : 2006年アジアにおける疫学的解析を通して、ハンタウイルス感染の浸淫状況を明らかにし、新規ハンタウイルスを検索する。
- Serological and molecular biological stdies on hantavirus infection among human and animals in AsiaSCF System for Establishment and Support of Center's of Excellence研究期間 : 2006年Serological surveylance of human and animal sera and hantavirus genome analysis by using molecular biological methods
- 文部科学省：科学研究費補助金(基盤研究(B))研究期間 : 2003年 -2005年代表者 : 森松 正美, 吉松 組子, 山本 欣郎自然免疫(innate immunity)は、各種病原体に共通して存在する構造物(リポ多糖体(LPS)など)を認識する生体防御反応であり、これらの構造物がToll様受容体(TLR)を介してNF-kBを活性化する細胞内情報伝達系により支えられている。最近、研究代表者らはLPS刺激によって発現誘導される新規分子MAILを発見した。この分子を中心に研究を実施した。1.MAIL破壊マウスの解析MAIL破壊マウスの約9割は胎生期に死亡することが判明した。胎子を摘出して調べたところ、肝臓の形成不全が認められ、これが胎生期致死の主な要因であると考えられた。MAIL破壊マウスの約1割は出生したが、これらはすべてアトピー性皮膚炎様の病態を示した。2.MAIL遺伝子発現調節の解析MAIL遺伝子のプロモータをクローニングしてそれより各種変異体を作製して機能解析を行った。NF-kBがMAIL遺伝子の発現を促進し、特に転写開始点近傍に存在するkBモチーフ配列が重要であることが判明した。MAIL遺伝子はNF-kBによって発現誘導され、転写翻訳によりできたMAILタンパク質はNF-kBをフィードバック制御することが示唆された。3.BRCA2遺伝子産物の解析NF-kB標的分子のひとつであるBRCA2は、相同組換えによるDNA損傷修復や細胞周期の調節を通じて細胞増殖や胎子発生に重要な役割を果たしている。BRC...
- ハンタウイルス核蛋白およびエンベロープ蛋白の構造と機能に関する研究文部科学省：科学研究費補助金研究期間 : 2001年04月 -2004年03月代表者 : 森松(吉松)組子組換え蛋白を用いて粒子形成、出芽、細胞への侵入の感染サイクルにおける構造蛋白の役割を解析する
- 日本学術振興会：科学研究費助成事業 基盤研究(B)研究期間 : 2001年 -2003年代表者 : 森松 組子, 小野 恵利子, 森松 正美, 苅和 宏明, 有川 二郎1.CAG promotorを用いて発現させた組み換えハンタウイルスエンベロープ蛋白が細胞融合活性を持つことを明らかにした。ハンタウイルスの中和エピトープの中央に細胞融合関連エピトープが位置することから、この組み換え蛋白が中和エピトープを保持していることが示された。また、同時に核蛋白抗原もCAG promotorを用いて発現させることに成功し、エンベロープ蛋白との相互作用について検討した。その結果核蛋白はエンベロープ蛋白の発現と輸送を押さえる可能性があること。この二つの蛋白のみではウイルス様粒子は形成されないことが示された。 2.CAG promotorを用いて発現させた組み換えハンタウイルスエンベロープ蛋白を用いて、外皮のみがハンタウイルスエンベロープ蛋白であり、内部が不稔のvesicular stomatitis virus (VSV)であるPseudotype VSV (VSVΔG^*HTN)を作製し、迅速で安全な中和抗体測定システムを構築した。また、分泌型の組み換えハンタウイルスエンベロープ蛋白を抗原としてマウスへ免疫し、中和抗体の誘導の可否を検討した。その結果、中和抗体は誘導されなかった。そのため、次にVSVΔG^*HTN粒子を抗原としてワクチネーションを試みた。その結果、免疫されたマウスにはエンベロープ蛋白に対する抗体の上昇が確認された。さらにこのマウスに攻撃接種したところ、核蛋白に対する抗体の上昇、および活性はほとんど見られず、感染が防御されたと考えられた。以上のことから、エンベロープ蛋白をウイルス様にパッケージングすることで、感染防御活性を有する抗体応答が得られることが分かった。これらの結果から、この組み換え蛋白を用いた安全なワクチン開発の可能性が示された。
- 文部科学省：科学研究費補助金(基盤研究(B))研究期間 : 1999年 -2001年代表者 : 森松 正美, 吉松 組子, 小川 和重, 首藤 文榮, 吉村 佳典, 三好 一郎本研究の目的は,マウスのジーンターゲティングにおいて,ノックインにより哺乳動物に存在しない特異的な目印(エピトープタグ)を標的分子に導入し,生体反応におけるその動向を抗エピトープタグ抗体で調べる方法を開発するための基礎研究を行うことである。それに向けて、以下のごとく数種類のモデル遺伝子を標的とすべく基礎研究を行った。また、導入するエピトープタグについての解析を行った。1.内毒素誘導性新規遺伝子(MAIL)を標的とした検討MAILは、内毒素による顕著な発現調節を受け`遺伝子産物は速やかに核に移行するため、標的に適している。この遺伝子のゲノムDNAを分離し,塩基配列を決定してその構造を明らかにした。これを用いてジーンターゲティングを行った。2.キチナーゼ遺伝子を標的とした検討キチナーゼは特徴的な組織特異的発現調節を示す遺伝子であるため、これを標的とすることを試みた。この遺伝子のクローニング、構造解所、および発現解析を行い、ターゲティングを実施するための基盤を築いた。3.他の標的遺伝子候補の検索上記の他誘導性遺伝子であるB-13や、組織特異的遺伝子であるGlucose transporter、Leptin等、疾病に関わると考えられる遺伝子を選んで標的の候補として検索した。4.エピトープタグの解析新規エピトープの候補であるハンタウイルス由来タンパク質について、抗体の反応性や、細胞に与...
- 日本学術振興会：科学研究費助成事業 基盤研究(B)研究期間 : 1999年 -2001年代表者 : 有川 二郎, 森松 組子(吉松 組子), 苅和 宏明, 佐藤 浩, 高倉 彰1.ハンタウイルス抗体測定用の抗原として、大腸菌とバキュロウイルスを発現ベクターとしてハンターン型、ソウル型およびドブラバ型のそれぞれの核蛋白抗原の発現に成功し、いずれも、ELISA法診断用抗原として利用が可能である活性と量が得られることが判明した。 2.バキュロウイルス発現ベクターによって、核蛋白のN末端50アミノ酸を削除した抗原を発現させてELISAへ応用すると、血清型鑑別抗原として応用可能であることが、患者血清および感染げっ歯類血清を用いて確認された。 3.マウス肝炎ウイルス(MHV)核蛋白コード遺伝子をイーストの系で発現することに成功した。培養上清中の発現のピークは培養後72時間であった。しかし、ELISAにおける反応性が低く、発現系および核蛋白内の発現部位の再検討が今後の課題である。 4.リンパ球性脈絡髄膜炎(LCM)ウイルス国内分離株及びプロトタイプ株の核蛋白コード領域遺伝子をクローニングし、全長、C末領域および中間領域の3つの領域についてそれぞれCOS細胞での発現に成功した。それぞれの両域抗原の診断的有用性の検討が今後必要である。 5.LCMウイルス核蛋白発現バキュロウイルスを国立感染症研究所から分与を受け、その抗原性を検討した。その結果、感染Sf9細胞はIFA抗原として、また、Tm5細胞にて増殖させたウイルスはELISA抗原に適していることが判明した。本抗原と両診断法を用いて、わが国のマウス動物実験施設、延べ1,117施設由来9,840検体を対象に血清のスクリーニングを行った結果、ELISAは大量検体処理のためのスクリーニング試験に、IFAはその結果の確認試験に有用であることが示された。
- 日本学術振興会：科学研究費助成事業 奨励研究(A)研究期間 : 1999年 -2000年代表者 : 森松 組子本研究はハンタウイルスの侵入を阻止するために齧歯類における診断法、特にHPS感染を摘発する方法を確立することを目的とする。ハンタウイルスの抗原性は大きく3つに分けられる.ネズミ亜科(Murinae)、ハタネズミ亜科(Microtinae/Arvicolinae)、アメリカネズミ亜科(Sigmodontinae)それぞれに属する齧歯類を宿主とするウイルスである.本年度はさらに応用範囲を拡大し、さらに鑑別診断系の評価を行った。 1.平成11年度に引き続き野鼠血清の収集を行った。本年度はHPSのreservoir動物となるアメリカネズミ亜科のげっ歯類の血清および抗血清を入手することができた。さらにより多くのネズミ亜科、ハタネズミ亜科のげっ歯類血清合計31種について適切な抗体検出系を決定することができた。この結果からより広範な抗体摘発体制を取ることが可能となった。 2.平成11年度に引き続き、Hantaan,Puumala,SinNombreハンタウイルスに対する標準抗血清の作成を行った。 3.3種類のハンタウイルス、Hantaan,Puumala,SinNombreハンタウイルス核蛋白の抗原性の交差は少ないことが明らかとなった。 4.ネズミ亜科のウイルス3種類(Hantaan,Seoul,Dobrava)の核蛋白をコードするcDNAから主要共通抗原領域をコードする部分を削除し、バキュロウイルスベクターを用いて発現させることで調整した抗原を用いた鑑別診断法の感度は約95%、特異性は100%であり、十分な感度と特異性を示した。 以上の結果から、ハンタウイルスの侵入を阻止するために、広範な野鼠の抗体検出法、適切な抗原を供給することが可能となった。さらにネズミ亜科のハンタウイルスについては詳細なウイルス型別も可能な診断系を提供できることが明らかとなった。
- 日本学術振興会：科学研究費助成事業 基盤研究(B)研究期間 : 1999年 -2000年代表者 : 有川 二郎, 森松 組子, 苅和 宏明, 高島 郁夫, 森川 茂1.中国側研究分担者の所属研究機関を訪問し中国では地域的に強毒と弱毒のハンタウイルスが流行している可能性が示唆された。また、中国におけるワクチン開発の現状と有効性について情報を得た。さらに各国流行株と中国株の遺伝子の相互関係の解析ならびに血清型鑑別診断法の技術交流を行った。 2.スロバキア研究分担者の研究所を訪問し、欧州におけるハンタウイルス遺伝子解析に関する情報交換と技術習得を行った。 3.日本側研究協力者を米国側研究分担者の研究所に派遣しレセプター解析と遺伝子再集合ウイルス作製のための情報交換と技術交流を行った。 4.日本側研究協力者を米国Wisconsin大学獣医学部に派遣しcDNAから人工的にハンタウイルスを合成するための基礎的技術の習得を行った。 5.ヘルシンキ大学、Vaheri博士から北欧流行ハンタウイルスの病原性規定因子についての研究打ち合わせを行うと同時に、当該研究分野の他の研究者と共に情報交換を行った。 6.東南アジア流行ウイルスとの比較を目的にウイルス株および患者と感染動物の血清の取得の可能性について情報交換を行った。 7.ハンタウイルスの感染がGalNacを特異的に認識するレクチンであるDBAとSBAによって増強されること、ならびに感染過程でのウイルスと細胞膜の融合には細胞側の因子が関与することを明らかにした。 8.マウスへの病原性が末梢組織での増殖効率が高いことが関連し、エンベロープ蛋白の単一アミノ酸の変異が規定していることを明らかにした。 9.遺伝子再集合ウイルス作成のためにウイルス遺伝子の発現をMとS遺伝子について完了した。また、転写、複製機構解明のためのミニゲノムを作成した。今後、それら遺伝子発現蛋白とミニゲノムの発現の確認が課題である。
- 日本学術振興会：科学研究費助成事業 基盤研究(A)研究期間 : 1998年 -2000年代表者 : 有川 二郎, 森川 茂, 苅和 宏明, 森松 組子1.ハンタウイルスの感染効率が、N-アセチルガラクトサミン結合性レクチン(DBAとSBA)を添加することにより約10倍増強され、細胞表面上のレセプター又はその近傍にあるN-アセチルガラクトサミンとウイルスをレクチンが架橋することにより吸着効率が増強するためと考えられた。 2.ハンタウイルス感染後、細胞融合を惹起する細胞株と起こさない細胞株の確立に成功し、ハンタウイルスエンベロープ蛋白が引き起こす感染細胞融合には細胞側因子も関与していることを明らかにした。 3.強毒株感染マウスでは脳へのウイルスの到達が弱毒株に比べ4-5日早いことがその後の全身感染の拡大と致死的転帰につながるものと考えられた。 4.強毒株は脳微小管内皮細胞と腹腔マクロファージにおいて弱毒株よりも高い増殖率を示したが、神経芽腫細胞では両ウイルスに差は認められなかったことからウイルスの病原性には末梢の標的細胞での増殖率が関連していると考えられた。 5.強毒と弱毒ハンタンウイルス間で遺伝子分節の交換された遺伝子再集合ウイルスを作成し、病原性と関連するウイルス側の要因を解析した結果、Mセグメント遺伝子中のアミノ酸変異(515番目アミノ酸、IleからThr)が病原性に関連し、Lセグメントも部分的に関連することが明らかになった。 6.強毒株感染SCIDマウスに、感染3週後のBALB/cマウス由来脾臓細胞を移入したところ、感染3週目に移入した場合、移入4日目から著しい体重減少が一過性に認められ、免疫関連の病原性発現機構が確認された。 7.ハンタンウイルスの核蛋白遺伝子(S分節)とエンベロープ蛋白遺伝子(M分節)をクローニングし、哺乳動物細胞(293T細胞)で高率に発現させることに成功した。 8.Lセグメントの発現系の確立を目的にLセグメントRNA全長のクローニングに成功した。今後、この発現の有無について検討を加える予定である。
- 文部科学省：科学研究費補助金(基盤研究(C))研究期間 : 1998年 -1999年代表者 : 苅和 宏明, 吉松 組子, 水谷 哲也, 有川 二郎, 高島 郁夫我が国や極東口シアにおいてハンタウイルスの疫学調査を行い、以下のことが明らかになった。1.実験的に作出した感染ラットの尿中にウイルスが排泄され、この尿を感受性ラットに経鼻的に滴下することにより感染が成立すること、さらに尿接種によって感染したラットの尿からもウイルスが検出されたことから、尿を介した経鼻伝播で本ウイルスが自然界で維持されうることが明らかになった。2.パキュロウイルスで様々なハンタウイルスの核蛋白を発現させ、これらを抗原に用いたELISA法を開発した。本法により、抗体保有が判定できるばかりでなく、血清を異なった抗原に反応させることで、感染したハンタウイルスの型までも類推することが可能となった。3.ハンタウイルス肺症候群(HPS)患者の体内のウイルス量を測定するために定量的PCR法を開発した。このPCR法により、HPSの急性期には高力価のウイルスが血中に存在するが、感染の回復期にはウイルスはもはや血中から消失することが明らかにされた。4.複数の血清診断法を用いて、我が国のヒトにおける大規模なハンタウイルスの血清疫学調査を行った。健常者では抗体保有者は全く存在しないにもかかわらず、原因不明の肝炎患者においては約2〜3%の患者がハンタウイルス抗体を保有することが判明した。これにより、本邦でも一般の市民にハンタウイルスの感染があることが明らかになった。5.極東口シアのウラジ...
- 文部科学省：科学研究費補助金(奨励研究(A))研究期間 : 1997年 -1998年代表者 : 森松 組子ハンタウイルスは腎症候性出血熱の原因ウイルスで、ブニヤウイルス科に属するRNAウイルスである。この研究ではハンタウイルス内部蛋白の特定のエピトープを標的抗原とする新しいタイプのペプチドワクチンの開発を目的として、マウス実験感染モデルを用いて単クローン抗体E5/G6エピトープペプチドの誘導する免疫応答とその感染防御活性について検討している。平成10年度は、カセット配列(主要組織適合抗原結合領域)を付加した,E5/G6エピトープのペプチド抗原による免疫応答の解析を明らかにするために、ペプチドを合成しマウスに免疫し免疫応答を測定することを試みた。1. ペプチドデザインPeptide 1 YEGF-EDVNGIRKP-KAKGITPeptide 2 YEGFS-EDVNGIR---KAKGITをMAP及び通常のペプチドとして合成した。これはH-2bのカセット配列に、E5G6エピトープ配列EDVNGIRKPKを挿入したものである。2. 免疫MAPのペプチド抗原を100ug/mouse complete adjuvantとともにC57BL6マウスに免疫し、経時的に採血し、抗体価の上昇を調べた。3. T細胞応答の測定系の確立牌臓細胞をX線照射によって不活化し、抗原提示細胞として用いることを試みた。昨年度の研究結果から非RI測定系を確立するため代替法として、市販のテトラゾリウム塩類、酵素を比...
- 文部科学省：科学研究費補助金(国際学術研究)研究期間 : 1996年 -1997年代表者 : 有川 二郎, 宋 干, 李 徳新, Antti Vaheri, Bo Niklasson, 網 康至, 伊勢川 裕二, 五十嵐 章, Clarence Pet, 杉山 和良, 高島 郁夫, 森松 組子, 王 華, Clarence Peters1. 研究分担者、杉山和良らは中国、北京市と西安市近郊で野生げっ歯類の捕獲調査を実施し、2地区より100例のげっ歯類が得られ抗体および抗原の分布を明らかにした。2. 中国側研究分担者、李徳新を3カ月間、日本側研究機関に招聘し、得られた材料の解析を行い、中国由来2株のウイルスの遺伝子配列が一部決定され、日本側流行株よりもむしろ韓国由来株に近縁であることが明らかになった。3. 研究代表者、有川二郎と研究分担者、森松組子は米国側研究分担者、Petersを訪問し、世界各地での本ウイルス流行状況と遺伝的解析方法についての最新情報を得た。4. 研究分担者、Vaheri(フィンランド)をわが国に招聘し、北欧地域調査と北欧由来およびアジア由来ウイルスの相互比較の可能性について情報交換と将来計画を検討した。5. 研究代表者有川と研究分担者、高島は、英国、オーストリアおよびスロバキア側研究分担者の所属研究機関を訪問し、欧州におけるハンタウイルス感染症流行地域拡大に関する情報を得た。病原性の高い血清型(Dobrava型)である可能性についても情報を得た。6. 研究分担者、森松、苅和は英国側研究分担者の研究所を訪問し、遺伝子再集合ウイルス作製法ならびにReverse genetics法に関する最新の情報を得た。7. 研究代表者、有川および研究分担者、森松、高島、苅和は、韓国側研究分担者、李鎬汪の研究...
- 日本学術振興会：科学研究費助成事業 基盤研究(A)研究期間 : 1994年 -1996年代表者 : 有川 二郎, 高倉 彰, 杉山 和良, 吉松 組子, 苅和 宏明, 橋本 信夫1.バキュロウイルス発現ハンタウイルス組み換え核蛋白(Bac.HTN-NP)を用いたELISAとWestern blotting法による感度と特性を感染細胞を抗原とする従来のIFA法と比較・検討した。ラット、ヒト血清いずれにおいても感度はIFA法と同等であった。しかし、陰性血清におけるバックグランドの発色のレベルが比較的高く、発色の程度から肉眼判定するには問題点が残った。 2.上記1.で明らかとなったバックグランド発色は、抗原の精製の不十分さが原因と考えられた。しかし、Bac.HTN-NPは、ELISA抗原として使用するための穏やかな条件下では可溶化されづらくこれ以上の精製が困難でることが判明した。そこで、次に、大腸菌中に同様のHTN-NPを発現させる系を構築した。この系では発現蛋白にビオチンが融合蛋白として結合しているためにアビジンを用いてcapture ELISAを確立した。 3.本capture-ELISAの実験諸条件を検討し、至適抗原固相化時間ならびに抗原濃度を決定した。この条件下で本ELISAはIFAに比べ安全で同等の検出感度と特異性を有することが示され実験用ラット等を対象とする標準スクリーニングにも応用可能であると考えられた。 4.Bac.HTN-NPをWestern blotting法に応用した場合、感度はラット血清、ヒト血清いずれにおいてもIFA法とほぼ同等であった。しかし、陰性血清群を用いた特異性の検討では、IFA法で非特異反応や紛らわしい蛍光像を示す検体でもWestern blotting法では非特異反応が全く認められなかった。すなわち、Western blotting法は、IFA法に比べ特異性が高いことが判明し、確定診断法として有用であると考えられた。 5.今後、本法のキット化のための基礎的条件の決定が必要である。
- 文部科学省：科学研究費補助金(奨励研究(A))研究期間 : 1995年 -1995年代表者 : 吉松 組子ハンタウイルスは腎症候性出血熱の原因ウイルスで、ブニヤウイルス科に属するRNAウイルスである。この研究ではハンタウイルス核蛋白の誘導する液性抗体の感染防御について明らかにすることを目標として感染防御に有効なエピトープ領域の決定を行なった。1次構造上の抗原領域の決定; 大腸菌を使ってハンタウイルスHantaan 76-118株の核蛋白の様々な部分を組換え蛋白として発現させ、単クローン抗体との反応性を調べ、単クローン抗体の認識部位を明らかにした。リニアエピトープを認識する単クローン抗体E5/G6についてはcDNAから予想されるアミノ酸配列を元にペプチドを合成し反応性を確認し、エピトープの特定を行なったところ,アミノ酸166-175の配列,EDVNGIRKPKであることが明らかとなった。またこの実験に付随して,ハンタウイルスの核蛋白のアミノ酸1から103がimunodominant領域であることが分かった.すなわち,人及び動物がハンタウイルスに感染したとき,核蛋白に対する抗体が強く誘導されるが,そのほとんどのリニアエピトープはこの領域に集まっていることが明らかとなり,診断抗原として有用な領域であることが分かった。また、立体構造依存性のエピトープについては,バキュロウイルスでの同様の実験から,この1-103の領域と200-300の領域にもにも存在することが示唆された。スクレープロ-デ...
- 文部科学省：科学研究費補助金(国際学術研究)研究期間 : 1993年 -1995年代表者 : 有川 二郎, Bo Niklasson, 宋 干, 杉山 和良, 橋本 信夫, Niklasson Bo, 周 桂珍, 森松 組子, Ake Lundkvis, Bo Niklasson1.スウェーデン側研究分担者(Dr.BoNiklasson)の所属研究機関(Swedish Institute for Infectious Disease Control)およびフィンランド、ヘルシンキ大学ウイルス研究所(教授、Dc・Antti Vaheri)を、有川(研究代表者)と橋本が平成5年度に、有川および森松、苅和(研究分担者)が平成7年度に訪問し、スウェーデンとフィンランドにおける腎症候性出血熱流行の現状ならびに調査・研究に関する情報を収集し、わが国の研究成績とともに情報交換した。スウェーデン側研究分担者とは、患者血清ならびに臨床材料収集・提供のシステムについて討議し、スウェーデン側からは患者血清と免疫血清を、日本側からは単クローン抗体を相互提供することとした。また、英国、グラスゴ-大学の研究者とも情報交換を行い、ヨーロッパ全域における腎症候性出血熱流行についての情報を得た。2.中国側研究分担者(Dr.Song Can)と研究協力者(Dr.Han Changshou)を平成5年度に、スウェーデン側研究分担者(Dr.BoNiklasson)と研究協力者(Dr.AkeLundkvist)を平成7年度にわが国に短期招聘し、日本側各研究分担者と情報交換するとともに共同研究の実施について討論した。3.わが国研究分担者(森松組子、北海道大学)をスウェーデン側研究分担者(Dr....
- 文部科学省：科学研究費補助金(奨励研究(A))研究期間 : 1994年 -1994年代表者 : 吉松 組子これまでに、私は蚕のバキュロウイルス系を用いて、ハンタウイルスの核蛋白を大量に発現するシステムを開発してきた.今回の研究では、この組換え蛋白を用いて、RNPトランスフェクション系を確立する事を目的とした.ハンタウイルス核蛋白を蚕のヘモリンフ中に発現させた場合、不溶性となる.この不溶性蛋白は、尿素及びグアニジン塩酸塩で可溶化され、特に尿素存在下で陰イオン交換クロマトグラフィーを行った場合、可溶性の蛋白として精製が可能である.しかしながら、この蛋白は消化を受け、フラグメント化されている場合が多い.これをさらに精製し、全長の核蛋白のみを精製するため、核蛋白に対するモノクローナル抗体を使ったAffinity精製を試みた.結果は収率は著しく低いものの、全長を含む核蛋白も精製が可能であった.しかしながら、これらの核蛋白に対するモノクローナル抗体を用いて精製した場合、それぞれの抗体の認識部位によって精製されるフラグメントが異なる.このため、これらの抗体の一次構造上の認識部位を決定しておく必要が生じた.また、Derai et alは大腸菌による発現組換え蛋白がRNA結合能を保持していたことを報告している.このため、大腸菌の発現系を利用し、抗原部位の決定を試みた.結論は、比較的効率よく全長の組換え蛋白を精製できるモノクローナル抗体は、立体構造認識の一部の単クローン抗体と、N末端を認識する抗体で...
- 文部科学省：科学研究費補助金(奨励研究(A))研究期間 : 1993年 -1993年代表者 : 森松 組子, 吉松 組子1.感染蚕ヘモリンフ中の不溶性画分を遠心洗浄することによって、組換え蛋白を部分精製、濃縮を行った。この不溶性画分は、弱アルカリ性(pH8.0)、還元条件下で、4M Guanidin Hydrochrolide, 6M Urea,あるいは1%SDS等によって可溶化された。ここから可溶化のための物質を取り除くと再び不溶性の沈殿となった。尿素存在下の陰イオン交換クロマトグラフィーを行なったところ、0.15-0.2MのNaClで核蛋白は溶出され、この画分は50mM Tris-HCl pH8.0、または水に透析後も再沈殿することはなかった。しかしながらこの時、リン酸を含む緩衝液に透析すると沈殿を生じることが分かったので、以下Tris系の緩衝液を用いることとした。この可溶化核蛋白を含む画分は室温で取り扱うと断片化が生じ、これはセリンプロテアーゼ阻害剤であるPMSFを加えて4℃で取り扱うことによって妨げることが明かとなった。2.可溶化され、濃縮された組換え核蛋白は単クローン抗体を結合させたアフィニティーカラムを用いてさらに精製することが可能であった。この抗原をHigh-density Particleに結合させ凝集抗原の作製を試みた。しかしながら抗原を感作させるだけでParticleは凝集した。塩析では10%飽和硫安で沈殿し、しばしば不溶化した。これらの結果は、アミノ酸配列からも予想されて...
- 文部科学省：科学研究費補助金(試験研究(B))研究期間 : 1991年 -1993年代表者 : 有川 二郎, 伊勢川 裕二, 石原 智明, 吉松 組子, 板倉 智敏, 橋本 信夫1.腎症候性出血熱ウイルス(Hantaan virus)を接種された重症複合型免疫不全(CB-17 scid)マウスでは、成熟例(4〜6週齢)においても感染後35日目に発症・死亡することが明らかになった。2.それら発症scidマウスはいずれも、腎臓をふくむ全身諸臓器中にウイルス抗原が分布していることが酵素抗体法による免疫組織学的検索によって確認された。また、腎臓の機能障害を示す血液尿素窒素(BUN)値のわずかな上昇も観察されたことから、人と同様の腎症状を伴う疾患モデルとなり得る可能性が示唆された。3.感染動物臓器から抽出されるRNAを鋳型として合成したcDNAを用いるRT-PCR法にNested-PCR法を組み合わせることによって、臓器中のウイルスゲノムを高感度に検出する方法を確立した。本法は、ウイルス分離法に比べ血中の中和抗体の存在に影響されないことから、持続感染マウスの病態の解析に有効な解析方法であると考えられた。4.正常BALB/cマウス由来の脾臓細胞を移入された感染scidマウスでは、細胞移入4日目以降、抗ウイルス抗体の急激な出現と同時に腎臓の機能障害の指標である血中BUN値の顕著な上昇が認められ、感染腎臓組織への免疫反応が機能障害を引き起こしていると考えられた。しかし、明瞭な病理組織学的変性の出現は確認されていない。免疫複合体の臓器中への沈着は確認されていないがさら...
- 文部科学省：科学研究費補助金(一般研究(C))研究期間 : 1990年 -1992年代表者 : 有川 二郎, 吉松 組子, 高島 郁夫, 橋本 信夫1.エンベロープ蛋白上の中和抗体の誘導に関与する抗原は同時に感染増強抗体をも誘導することが単クローン性抗体を用いた研究によって明らかになった。さらに、感染の中和と増強は、作用する抗体の濃度に依存し、高濃度では感染を中和し、低濃度では感染を増強した。すなわち、ある特定のエピトープのみを含む抗原をワクチンとして用いても、感染中和抗体のみを誘導することは困難であると判断される。2.高い病原性をもつウイルスの構成蛋白をより安全かつ効率的に精製することを目的に、バキュロウイルスを発現ベクターとして組み換えエンベロープ蛋白の調製を試みた。発現蛋白の抗原性は単クローン性抗体の結合能で判定する限り、本来のウイルスのものと区別することは出来なかった。しかし、中和抗体の誘導能は極めて低く、機能的な抗体の誘導には問題があることが明らかになった。3.以上のような限界の存在下では、組み換え蛋白とアジュバントの組合せによって高レベルの中和抗体を持続させる試みが今後必要であると考えている。また、組み換えEnv.蛋白や組み換え核蛋白で免疫された脾臓細胞の移入によっても感染からの回復に有効であることがマウスを用いた実験感染で明らかになった。このため、細胞性の免疫の誘導も考慮したワクチンの開発が今後期待される。4.本疾患の発症メカニズムとして感染個体の免疫応答の可能性が示唆されているものの未だ不明な点が多く残さ...
- 腎症候性出血熱ウイルス(ハンタウイルス)の病原性に関する研究科学研究費補助金
- Study on Virulency and pathogenicity of hantavirus, which causes hemorrhagic fever with renal syndromGrant-in-Aid for Scientific Research
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- 特許第3809502号:ハンタウィルス抗原蛋白質およびモノクローナル抗体 2006年06月02日吉松 組子, 有川 二郎, 橋本 信夫, 三谷 勝男 有川 二郎, 橋本 信夫, 株式会社エイアンドティー 200903028180319252