Research activity information

• In Vitro Selection and Characterization of Tecovirimat-resistant Mpox Virus.
  Kei Konishi; Haruka Maeda; Misato Shibazaki; Takao Shishido; Manabu Igarashi; William W Hall; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa; Akihiko Sato
  The Journal of infectious diseases, 18 May 2026, [International Magazine]
  English, Scientific journal, BACKGROUND: Mpox virus (MPXV) is the causative agent of mpox, a disease characterized by skin lesions, and remains a global threat since the recent 2022 outbreak. Tecovirimat is an antiviral agent that inhibits MPXV replication in vitro and in vivo by targeting the viral phospholipase VP37 protein and blocking viral envelope wrapping. Although tecovirimat has been administered to many mpox patients, the emergence of tecovirimat-resistant MPXVs carrying mutations in the VP37 protein has been reported, particularly in immunocompromised individuals. However, the impact of these respective mutations on drug susceptibility and viral fitness have not been fully characterized. METHODS: Tecovirimat was applied as a selective pressure in vitro to isolate MPXV variants carrying clinically observed VP37 substitutions. Drug susceptibility among the major VP37 mutations was compared, and the cellular fitness of the isolated viruses was evaluated. RESULTS: The VP37-N267D and -Y258C mutants showed markedly reduced susceptibility to tecovirimat compared with the VP37-I372N and -T245I (+I7L-A388S) mutants, while retaining susceptibility to an alternative anti-MPXV agent, brincidofovir. The VP37-I372N, T245I (+I7L-A388S), Y258C, and N267D mutants exhibited viral fitness comparable to that of the parental virus in A549 and Vero cells, as well as in MRC-5 and HaCaT cells. CONCLUSIONS: These findings provide insights into the phenotype of each VP37 mutation and emphasize the importance of developing antivirals with distinct mechanisms of action to effectively combat tecovirimat-resistant MPXV.
• The Evaluation of Zika virus susceptibility in Aedes (Ochlerotatus) japonicus from Hokkaido, Japan.
  Leo Uchida; Kana Iwamoto; Shino Kawae; Makoto Shimooka; Ronald Enrique Morales-Vargas; Yuki Eshita; Yasuko Orba; Hirofumi Sawa; Yasukazu Muramatsu
  Japanese journal of infectious diseases, 30 Apr. 2026, [Domestic magazines]
  English, Scientific journal, Zika virus (ZIKV) is a mosquito-borne orthoflavivirus that causes acute symptoms, such as fever and conjunctivitis. Aedes (Stegomyia) aegypti and Ae. (Stg.) albopictus are the primary and secondary vectors, respectively, although other Aedes species may also transmit ZIKV. Aedes (Ochlerotatus) japonicus, native to East Asia, has recently spread to North America and Europe; however, its potential in ZIKV transmission, particularly in Japanese populations, remains unclear. In our previous study, a low viral titer of ZIKV was detected in a single virus-challenged Ae. japonicus individual. In the present study, we increased the sample size and compared infection rates and viral titers among Ae. japonicus, Ae. aegypti, and Ae. albopictus following exposure to the ZIKV strain MR766. Cumulative infection rates of Ae. japonicus (18%) were lower than those of Ae. aegypti (71%) and Ae. albopictus (67%). Nonetheless, some Ae. japonicus individuals had higher viral titers than the challenge dose, indicating efficient replication. ZIKV RNA was also detected in 14% of the saliva samples at 10 days post-infection. These findings suggested that the Japanese population of Ae. japonicus could potentially serve as a vector.
• Antiviral effect of cetylpyridinium chloride on SARS-CoV-2 in patients with COVID-19: An Observational Study.
  Ryo Takeda; Hirofumi Sawa; Michihito Sasaki; Yasuko Orba; Nako Maishi; Takuya Tsumita; Yoichi Ohiro; Akira Hasebe; Hidehiko Sano; Yoshimasa Kitagawa; Kyoko Hida; Yasuhiro Hida
  Journal of oral biosciences, 68, 2, 100772, 100772, Apr. 2026, [International Magazine]
  English, Scientific journal, OBJECTIVES: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the viral agent responsible for the COVID-19 pandemic and primarily invades the body through the oral cavity. Cetylpyridinium chloride (CPC), a widely used component of oral care products, has demonstrated antiviral effects against SARS-CoV-2; however, the clinical evidence remains limited. This study investigated the virucidal effects of oral CPC in patients with COVID-19 and its impact on reverse transcription polymerase chain reaction (RT-PCR) positivity. METHODS: Thirty-four patients with COVID-19 were initially recruited after obtaining positive PCR test result. This study was conducted during a nationwide Delta-variant-driven epidemic characterized by high viral loads and enhanced transmissibility. Saliva was collected at seven predefined time points to assess the effects of CPC-containing tablets and mouthwash. Viral RNA levels and infectious viral titers were quantified using qRT-PCR and plaque assays. Sixteen patients with detectable SARS-CoV-2 RNA in baseline saliva samples who completed all scheduled samplings constituted the final analytical cohort. RESULTS: During the Delta-variant-predominant period, CPC-containing tablets reduced both SARS-CoV-2 RNA levels and viral infectivity in saliva. In contrast, CPC-containing mouthwashes did not significantly reduce viral RNA levels compared to baseline samples. The RT-PCR positivity rates after CPC use varied according to the applied cycle threshold (Ct) cutoff values. CONCLUSIONS: CPC-containing tablets transiently decrease the SARS-CoV-2 salivary viral load and infectivity and may help reduce the risk of transmission in public settings. When CPC formulations are used prior to saliva-based PCR testing, the Ct cut-off values may require adjustment. CLINICAL SIGNIFICANCE: Despite challenges in conducting saliva-based clinical research in COVID-19 patients, this study provides clinical evidence supporting the antiviral efficacy of CPC formulations on salivary SARS-CoV-2 during the Delta-variant pandemic.
• Circulation of an emerging neurotropic tick-borne phenuivirus in brown bears and wildlife in northern Japan
  Keita Matsuno; Anastasiia Kovba; Michito Shimozuru; Yuma Ohari; Mebuki Ito; Jingshu Li; Yume Mimura; Kohei Shinjo; Teruhiro Kanagawa; Masami Yamanaka; Kotaro Shimizu; Leo Uchida; Yasuko Orba; Michihito Sasaki; Nijiho Kawaguchi; Eri Fujii; Ryo Nakao; Mackenzie Kwak; Yurie Taya; Toshio Tsubota
  Springer Science and Business Media LLC, 18 Mar. 2026
  Abstract

  Tick-borne phenuiviruses are a group of zoonotic pathogens causing severe, sometimes lethal, diseases in humans. While several novel phenuiviruses have been identified in ticks, especially in East Asia, their mammalian infectivity largely remains uncharacterized. Here, we report the identification and successful isolation of an emerging tick-borne phenuivirus, Toyo virus (TOYV), from Hokkaido brown bears (Ursus arctos yesoensis) during wildlife surveillance in northern Japan. TOYV was isolated from bear samples using type I and II interferon receptor-knockout (AG129) mice and Hep3B cells. Intracerebral inoculation of suckling AG129 mice induced neurological signs, and viral replication was confirmed in mouse neuroblastoma cells. Detection of TOYV RNA in brown bears and Haemaphysalis species ticks, together with seropositivity in brown bears, raccoons, and sika deer, demonstrated active tick-borne circulation of TOYV among diverse wildlife hosts. This study provides the first evidence of tick-borne phenuivirus infections in an ursid species and the neuroinvasive and neurotropic potential of TOYV. Our findings underscore the critical role of wildlife surveillance in early detection of viruses with zoonotic potential and accelerating responses to emerging viral epidemics.

• Integrated in vivo and transcriptomic analyses of lethal Oropouche virus infection reveal suppression of pathogenic host responses by antiviral therapy
  Cássia Sousa Moraes; Gabriel Gonzalez; Akihiko Sato; Shigeru Miki; Atsuko Inoue; Koshiro Tabata; Joshua W. Kranrod; Chilekwa Frances Kabamba; Aiko Ohnuma; Keita Matsuno; Rio Harada; Shinji Saito; Michihito Sasaki; Yasuko Orba; William W. Hall; Hirofumi Sawa; Yukari Itakura
  16 Mar. 2026
• Application of the CPER reverse genetics system for genetic engineering of rabies virus.
  Yukari Itakura; Nijiho Kawaguchi; Koshiro Tabata; Gabriel Gonzalez; Kei Konishi; Aiko Ohnuma; Itsuko Furuta; Naoto Ito; Shinji Saito; William W Hall; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki
  Journal of virology, e0187225, 13 Mar. 2026, [International Magazine]
  English, Scientific journal, Reverse genetics (RG) systems are essential tools for basic virological studies and applied studies using engineered recombinant viruses in various research fields. While the circular polymerase extension reaction (CPER) has been widely applied to prepare a full-length infectious complementary DNA (cDNA) of positive-sense RNA viruses, its use for negative-sense RNA viruses (mononegaviruses) remains limited. Here, we report the first CPER-based RG system for rabies virus (RABV), a member of mononegaviruses. Infectious RABV was successfully rescued from cells transfected with helper plasmids and the CPER product, the assembled overlapping DNA fragments encoding the full-length viral genome cDNA and regulatory elements. Using this system, we generated wild-type, point-mutant, reporter-expressing, and chimeric RABVs, all of which retained their expected biological properties. Deep sequencing revealed that CPER-derived viruses occasionally harbor low-frequency mutations undetectable by Sanger sequencing, highlighting PCR-related artifacts as a limitation. In addition, CPER products with a pUC19 backbone could be directly applied for Escherichia coli transformation and cloning of RABV full-genome cDNA plasmids, offering a flexible, ligase-free cloning strategy for conventional RG. Our work establishes CPER as a versatile platform for engineering recombinant RABVs, facilitating rapid generation and genetic manipulation of RABV with potential applications for research on other mononegaviruses.IMPORTANCEReverse genetics systems allow researchers to generate recombinant viruses with precise genetic modifications, advancing studies of viral replication, pathogenicity, and vaccine development. However, constructing a full-length viral genome expressing plasmids is often time-consuming and technically demanding. To bypass the cloning process, a simple, cloning-free reverse genetics platform based on the circular polymerase extension reaction (CPER) has been applied for positive-sense RNA viruses. In this study, we applied the CPER-based reverse genetics system for rabies virus (RABV), a mononegavirus, enabling rapid and flexible generation of recombinant RABVs, including mutant, reporter-expressing, and chimeric clones. Our approach greatly facilitates genetic engineering of RABV and provides a versatile framework that can be extended to other mononegaviruses, thereby accelerating both basic and applied virology research.
• NS4B and NS5 mutations expand the temperature range permissive for the replication of dual-host affiliated insect-specific flaviviruses in mosquito cells.
  Hiroko Kobayashi; Koshiro Tabata; Michihito Sasaki; William W Hall; Hirofumi Sawa; Yasuko Orba
  Archives of virology, 171, 3, 26 Feb. 2026, [International Magazine]
  English, Scientific journal, The genus Orthoflavivirus within the family Flaviviridae includes mosquito-borne flaviviruses (MBFs) and dual-host affiliated insect-specific flaviviruses (dISFs). MBFs infect both vertebrates and mosquitoes, while dISFs are thought to infect only mosquitoes, despite their phylogenetic proximity to MBFs. Some dISFs have been shown to be capable of infecting mammalian cells in the setting of a suppressed immune response and subphysiological temperatures. Therefore, temperature sensitivity is considered one of the key factors restricting viral host tropism. To investigate the effect of temperature on dISFs propagation, we evaluated the growth of two dISFs, Psorophora flavivirus (PSFV) and Barkedji virus (BJV), in mosquito-derived C6/36 cells at 28 and 37 °C. While MBFs, including dengue virus (DENV) and Japanese encephalitis virus (JEV) could propagate efficiently at both temperatures, both PSFV and BJV failed to propagate at 37 °C. Serial passaging of PSFV with a gradual increase in temperature resulted in PSFV adaptation to 37 °C, and whole-genome sequencing revealed that the 37 °C-adapted PSFV acquired non-synonymous amino acid substitutions in the non-structural proteins, NS4B and NS5. We demonstrated that both NS4B and NS5 amino acid mutations in PFSV confer the ability to propagate at 37 °C, suggesting that the replication machinery contributes to the thermal restriction on dISFs. This study provides new insights into the temperature sensitivity of dISFs and their relationship to host tropism.
• Epidemiological, clinical, and virological characterizations of Yezo virus infections, an emerging tick-borne orthonairovirus disease in Japan.
  Hiroki Yamaguchi; Keita Mizuma; Kei Watari; Yuma Ohari; Kazuya Mitsuhashi; Kazuma Tamiya; Nijiho Kawaguchi; Yasuko Orba; Masayuki Saijo; Keita Matsuno
  The Journal of infectious diseases, 10 Feb. 2026, [International Magazine]
  English, Scientific journal, BACKGROUND: Febrile disease caused by infections of Yezo virus (YEZV), a tick-borne orthonairovirus, was first reported in Japan in 2021 and subsequently identified in China. While the recent studies suggest a broader endemic area of the emerging YEZV infections, its clinical and epidemiological characteristics remain poorly elucidated due to the limited number of reported cases. METHODS: We conducted retrospective and prospective surveillance of patients with suspected tick-borne febrile illnesses in Hokkaido, Japan, between 2013 and 2024. Serum samples were analyzed by RT-qPCR, IgM/IgG ELISA, and neutralization assay. Viral genome of isolates was sequenced and phylogenetically analyzed with publicly available YEZV genomes. RESULTS: A total of 22 patients with YEZV infections including 10 newly identified patients was analyzed. Most infections occurred between May and July, coinciding with the peak activity of Ixodes ticks in the study area. A case from southern Hokkaido was firstly identified. Fever, leukocytopenia, thrombocytopenia, elevated levels of liver enzymes, and ferritin were determined as common features observed in patients with YEZV infections. Viral RNA was typically detectable in the serum within 7 days after the disease onset. Serum antibodies became positive later and might persist up to at least 600 days. Phylogenetic analysis incorporating five new isolates showed the circulations of two genetic groups of YEZVs in Japan. CONCLUSIONS: Our results revealed characteristics of YEZV infections in patients and highlighted the importance of combination of molecular and serological diagnostics. The proposed case definitions and diagnostic framework may contribute future surveillance.
• Detection and genetic characterization of pigeon gammacoronaviruses
  Hiroko KOBAYASHI; Mai KISHIMOTO; Sakiho IMAI; Yasuko ORBA; Hirofumi SAWA; Masayuki HORIE
  Journal of Veterinary Medical Science, 88, 4, 654, 661, Japanese Society of Veterinary Science, 2026, [Domestic magazines]
  English, Scientific journal, Most members of the genus Gammacoronavirus infect avian hosts, but far fewer viruses have been characterized than in the other coronavirus genera, leaving their diversity largely unclear. Pigeon gammacoronaviruses were previously detected by consensus PCR, but only partial sequences were determined. In this study, we comprehensively analyzed public RNA-seq datasets and reconstructed two nearly complete genomes of pigeon gammacoronaviruses. Molecular evolutionary analyses showed pigeon gammacoronaviruses belong to the subgenus Igacovirus, and their pairwise distances to the members of this subgenus meet the International Committee on Taxonomy of Viruses (ICTV) species demarcation criteria, supporting their designation as a novel species in this subgenus. Notably, although nucleotide sequences are highly conserved among pigeon coronavirus variants, their 3'-genomic region exhibited differences in genome organization. These findings expand our knowledge of the diversity of gammacoronaviruses.
• Universal and quantitative detection of double-stranded RNAs as a signature of pan-virus infections using a luciferase-based biosensor.
  Michihito Sasaki; Eri Fujii; Satoko Sasaki; Takuma Ariizumi; Kei Konishi; Akihiko Sato; William W Hall; Hirofumi Sawa; Yasuko Orba
  Journal of biological engineering, 20, 1, 11, 11, 16 Dec. 2025, [International Magazine]
  English, Scientific journal, BACKGROUND: Infections with various RNA viruses and certain DNA viruses may produce double-stranded RNA (dsRNA) during replication, which trigger host innate immune responses. Immunoassays using anti-dsRNA antibodies have been widely employed to detect viral dsRNA. In this study, we used a luciferase-based dsRNA biosensor for viral dsRNA detection, which consists of protein kinase R (PKR)-derived dsRNA binding domains fused to split luciferase subunits and is available as part of a commercial system. RESULTS: We demonstrate the use of the dsRNA biosensor to measure viral dsRNA in RNA specimens extracted from cells infected with Japanese encephalitis virus (JEV). Moreover, the biosensor reacts to a broad-spectrum of dsRNAs from infection with representatives of various viral families including positive- and negative-sense single-stranded RNA (ssRNA) viruses, dsRNA viruses, and DNA viruses. We validated the specific interaction between the dsRNA biosensor and viral RNA including subgenomic flavivirus RNA (sfRNA) through RNA immunoprecipitation. Additionally, we observed luminescence signals directly from lysates of JEV-infected cells after cell lysis and phase separation with Triton X-114. Finally, we used the biosensor to assess the activity of antiviral compounds. CONCLUSIONS: Our results demonstrate that the luciferase-based dsRNA biosensor offers a simple, homogeneous, and high-throughput platform for quantifying viral replication, presenting a promising alternative to antibody-based dsRNA detection methods.
• Development of a novel plate reader-based antibody-dependent enhancement (ADE) assay for orthoflavivirus infections.
  Yannis Aklil; Hiroto Takeuchi; Gabriel Gonzalez; Atsuko Inoue; Kaito Maeda; Shintaro Kobayashi; Yukari Itakura; Shinji Saito; Anavaj Sakuntabhai; Michihito Sasaki; William W Hall; Hirofumi Sawa; Yasuko Orba; Koshiro Tabata
  Journal of virological methods, 338, 115210, 115210, Dec. 2025, [International Magazine]
  English, Scientific journal, Antibody-dependent enhancement (ADE) is one of the mechanisms associated with severe clinical outcomes in infections caused by certain viruses, including dengue virus (DENV). Several ADE assay systems have been established, including flow cytometric assays using live viruses, enzyme-linked immuno-sorbent assay (ELISA) for the detection of viral NS1, and luciferase reporter gene assays. Among these, the flow cytometric assay is the most commonly used to evaluate ADE activity; however, it has limitations such as high operational costs due to fixation and immunostaining procedures, as well as a long analysis time. Fluorescent protein-expressing single-round infectious particles (SRIPs) enables label-free detection of ADE activity, but the flow cytometric procedure still requires a long analysis time. In this study, to simplify and expedite the ADE assay using enhanced green fluorescent protein (EGFP)-expressing SRIPs, we developed a plate reader-based ADE assay as an alternative to the conventional flow cytometry-based method. To evaluate effectiveness of this assay, we measured ADE activities in K562 cells induced by pan-orthoflavivirus 4G2 and pan-dengue 4E11 monoclonal antibodies (mAbs) using both flow cytometric assays using live viruses and plate-reader-based EGFP-expressing SRIPs assays. The results showed strong correlations between the two different ADE assays with R² values of 0.92 for 4G2 mAb and 0.94 for 4E11 mAb (Pearson correlation coefficients). In summary, this newly established assay offers a high-throughput and cost-effective method for comprehensive characterization of the relationship between vaccine- or infection-induced antibodies and ADE in orthoflavivirus infections.
• Oita virus rediscovered after 50 years: isolation of genetically conserved strains from bats in Southern Japan
  Saiko Sawai; Kittiya Intaruck; Sho Sata; Mana Esaki; Kimitake Funakoshi; Shin Murakami; Keita Matsuno; Ryo Nakao; Kazunori Kimitsuki; Takaaki Yahiro; Akira Nishizono; Naganori Nao; Yasuko Orba; Ayato Takada; Masahiro Kajihara; Makoto Ozawa; Kosuke Okuya
  Microbiology Spectrum, 13, 12, e0315825, American Society for Microbiology, 11 Nov. 2025, [International Magazine]
  English, Scientific journal, ABSTRACT
  
  Bats are natural reservoirs of numerous viruses, including members of the Rhabdoviridae family. Oita virus (OITV), classified within the genus Ledantevirus , was first isolated from the blood of a Japanese horseshoe bat ( Rhinolophus cornutus ) in the Oita Prefecture, Japan, in 1972. However, since then, no other OITV isolates have been reported. In this study, we isolated two OITV strains—OITV 321/2022 and OITV 326/2022—from oral swab samples of R. cornutus collected in the Katano Cave, Kagoshima Prefecture, Japan. Genetic analysis revealed >98% nucleotide identity with the previous isolate, OITV 296/1972. Phylogenetic analysis confirmed their classification within the Ledantevirus subgroup C. We assessed viral growth kinetics using 17 cell lines from diverse animal species, including bats, rodents, humans, and arthropods. OITV 321/2022 showed efficient replication in primate-derived, bat-derived, and human-derived cells, but not in tick or mosquito cell lines. Experimental infection of BALB/c mice demonstrated that OITV 321/2022 induced no overt clinical signs via either the intracerebral or intranasal routes. However, systemic infection and viral replication in the brain and lungs were observed by intracerebral inoculation. Intranasal inoculation with OITV 321/2022 and OITV 296/1972 resulted in transient pulmonary infection without systemic dissemination. This study provides evidence of OITV circulation in wild bat populations over 50 years, highlighting the need for continued surveillance of bat-associated rhabdoviruses in Japan.
  
  IMPORTANCE
  
  Bats are known reservoirs of zoonotic viruses, and their proximity to humans raises concerns regarding zoonotic risks. We report the first isolation of the Oita virus (OITV), a bat-associated ledantevirus, that has circulated for over 50 years. Unlike a previous blood-derived isolate, our isolates were obtained from oral swabs, suggesting their potential for respiratory transmission. OITV could infect a wide range of mammalian cells, including human-derived cells, and induce systemic infection in mice without clinical symptoms. These findings indicate that OITV possesses a broad host tropism and may circulate among microbats through the respiratory tract. Although the pathogenicity of the newly isolated strain appears to be attenuated compared with that of a historical brain-passaged strain, its ability to replicate in human cells underscores its potential zoonotic relevance, necessitating active surveillance and functional characterization of bat-derived rhabdoviruses to better assess emerging infectious disease threats.
• Spatial gene expression analysis reveals pathological niches in Japanese encephalitis virus neuroinvasion.
  Yasuko Orba; Yukie Kashima; Koshiro Tabata; Yukari Itakura; Takuma Ariizumi; William W Hall; Hirofumi Sawa; Yutaka Suzuki; Michihito Sasaki
  Proceedings of the National Academy of Sciences of the United States of America, 122, 43, e2515006122, 28 Oct. 2025, [International Magazine]
  English, Scientific journal, Japanese encephalitis virus (JEV) infection causes encephalitis in humans and animals. Following intradermal infection, JEV crosses the blood-brain barrier (BBB) and reaches target cells in the brain parenchyma. However, the cellular dynamics and pathological niches involved in JEV neuroinvasion remain poorly understood. In this study, we investigated the early stages of JEV infection in the mouse brain employing a highly multiplexed spatial transcriptomics platform to map viral RNA and host gene expressions in intact brain sections at a single-cell resolution. Although JEV RNA was undetectable in brain sections at 1-day postinfection (dpi), innate immune responses were transiently activated across the brain. At 4 dpi, we detected limited viral RNA and mapped its spatial distribution, identifying glial cells surrounding microvessels as early targets of brain infection. We further characterized transcriptional changes in infected and surrounding bystander cells, revealing cell-type-specific antiviral responses. Notably, JEV neuroinvasion led to the downregulation of endothelial tight junction genes, indicative of an early event that precedes BBB impairment during subsequent disease progression. Our spatial transcriptomic analysis provides insights into cell-type- and region-specific responses to JEV infection, and highlights the early role of glial cells in shaping the immune response landscape of the brain. These findings greatly improve our understanding of JEV pathogenesis before the onset of clinical encephalitis.
• A cap-dependent endonuclease inhibitor acts as a potent antiviral agent against La Crosse virus infection.
  Kei Konishi; Yoshiyuki Taoda; Manabu Igarashi; Takao Shishido; Kazuya Yasuo; William W Hall; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki; Akihiko Sato
  Antimicrobial agents and chemotherapy, 69, 9, e0018625, 03 Sep. 2025, [International Magazine]
  English, Scientific journal, La Crosse virus (LACV) infection, the causative agent of La Crosse encephalitis, can lead to severe neurological symptoms and sequelae, particularly in children. Despite annual reports of neurologically symptomatic cases, no effective treatment has yet been established. Bunyaviruses, including LACV, utilize a cap-snatching mechanism for transcription, with a cap-dependent endonuclease (CEN) serving as a promising target for antiviral treatment. Specifically, we now demonstrate that a CEN inhibitor, carbamoyl pyridone carboxylic acid (CAPCA)-1, exhibits potent anti-LACV activity in vitro and in vivo. CAPCA-1 exhibited 50% effective concentration values below 1 µM in neuronal and non-neuronal cells, demonstrating a higher in vitro activity than the nucleoside analogs, ribavirin and favipiravir. Multiple passages of LACV in the presence of CAPCA-1 produced numerous amino acid mutations in the CEN active site. Notably, using a lethal infection model in mice, CAPCA-1 treatment reduced viral loads in the brain and extended the survival rate of LACV-infected mice. These findings highlight the potential of CEN inhibitors as treatment options for La Crosse encephalitis.
• スパルフロキサシンの多面的抗ウイルス作用によるRSウイルス複製の抑制効果
  小笠原 徳子; 山本 聡; 谷向 由佳; 吉田 有梨枝; 佐藤 彰彦; 澤 洋文; 大場 靖子; 吉田 圭太朗; 高野 賢一; 横田 伸一
  臨床とウイルス, 53, 3, 185, 185, 日本臨床ウイルス学会, Sep. 2025
  Japanese
• Selective TLR ligand stimulation enhances in vivo mosquito-borne flavivirus pathogenicity.
  Tatsuya Suzuki; Yuka Miyata; Saori Haga; Yumi Itoh; Tsukika Tanaka; Tomomi Hishinuma; Yasuko Orba; Yuki Eshita; Yusuke Sakai; Takeshi Kurosu; Shigeru Tajima; Chang-Kweng Lim; Masayuki Saijo; Atsushi Yamanaka; Thipruethai Phanitchat; Ronald Enrique Morales Vargas; Daisuke Okuzaki; Hirofumi Sawa; Takashi Satoh; Shizuo Akira; Yoshiharu Matsuura; Toru Okamoto
  Cell reports, 44, 9, 116210, 116210, 29 Aug. 2025, [International Magazine]
  English, Scientific journal, Mosquito saliva facilitates pathogen transmission and enhances the severity of diseases caused by mosquito-borne viruses; however, the underlying mechanisms are unknown. Here, we demonstrate that mosquito salivary gland extracts (SGEs) enhance flaviviral pathogenicity in vivo by activating innate immune responses following the accumulation of immune cells at the infection site. Among the innate immune signaling pathways, the TLR2 pathway enhances flaviviral pathogenicity in a manner similar to that of SGEs. TLR2 ligands and SGEs induce neutrophils to secrete chemokines that recruit virus-permissive monocytes and macrophages to infection sites. SGEs activate TLR2, and inhibition of TLR2 signaling markedly reduces mosquito-saliva-enhanced viral pathogenicity. Overall, this study provides important insights into vector-host interactions and suggests that TLR2 is a potential target for preventing mosquito-borne flaviviral infection.
• Synergistic antiviral activity of a cathepsin B/L inhibitor and a TMPRSS2 inhibitor against SARS-CoV-2 in vitro and in vivo.
  Shinsuke Toba; Kentaro Uemura; Takao Sanaki; Shinji Kusakabe; Kei Konishi; Shigeru Miki; Yuki Maruyama; Atsuhiro Iimuro; Takao Shishido; Michihito Sasaki; Yasuko Orba; William W Hall; Hirofumi Sawa; Akihiko Sato
  Virology, 612, 110661, 110661, 21 Aug. 2025, [International Magazine]
  English, Scientific journal, The spike (S) protein of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) binds to a host cell receptor which dictates the viral entry pathway. SARS-CoV-2 utilizes two different pathways for cellular entry mediated by both a host type II transmembrane serine protease (TMPRSS2) and cathepsin proteases. These host proteases cleave the viral S protein and initiate membrane fusion allowing viral infection. We previously isolated a SARS-CoV-2 mutant with deletion in the furin cleavage site of the S gene (del2) and revealed differences in cell tropism between wild-type (WT) and del2 viruses. Here, we evaluated the antiviral activities of cellular protease inhibitors against SARS-CoV-2 WT and del2 viruses using several different cell lines. The TMPRSS2 inhibitor, camostat, exhibited strong antiviral activity against WT virus but not del2, while the cathepsin B/L inhibitor, K11777, exhibited potent antiviral activity against the del2 virus. We isolated K11777-escape mutants of SARS-CoV-2 and SARS-CoV and demonstrated that these mutations facilitated S protein cleavage at the S2' site mediated by cathepsin L. Finally, we demonstrated that combination treatment of K11777 and camostat potently inhibited SARS-CoV-2 WT infection in vitro and in vivo, suggesting the usefulness of combination therapeutics targeting host TMPRSS2 and cathepsin proteases against coronavirus infection. In summary, our study characterized K11777 as an inhibitor of S2' cleavage by cathepsins, highlighting the critical role of the S2' site in SARS-CoV-2 cellular entry. This research sheds light on the infection process and has implications for potential therapeutic interventions for SARS-CoV-2 infection.
• SARS-CoV-2 uptake and inflammatory response in senescent endothelial cells are regulated by the BSG/VEGFR2 pathway.
  Yuya Sakurai; Yoichiro Fujioka; Nako Maishi; Ryo Takeda; Yusuke Ohba; Michihito Sasaki; Takahito Teshirogi; Wataru Ito; Yasuhiro Hida; Aya Matsuda; Kanta Kido; Yasuko Orba; Hirofumi Sawa; Kyoko Hida
  Proceedings of the National Academy of Sciences of the United States of America, 122, 31, e2502724122, 05 Aug. 2025, [International Magazine]
  English, Scientific journal, Aging is a risk factor for severe COVID-19, characterized by vascular endothelial dysfunction. Although possible susceptibility of vascular endothelial cells (ECs) to SARS-CoV-2 infection has been suggested, the details of entry into cells have not been clarified. Previously, we reported that in an aged mouse model of severe COVID-19, ECs show a massive viral uptake and inflammatory response. Here, we focused on the endocytic capacity of senescent ECs. We found that the senescent ECs showed high endocytic capacity and SARS-CoV-2 virus uptake. This triggers an nuclear factor-kappa B (NF-κB) pathway-mediated inflammatory response. Further, Basigin enhanced endocytosis in the senescent ECs by activating the intracellular vascular endothelial growth factor signaling. Thus, EC senescence is associated with enhanced SARS-CoV-2 endocytosis and subsequent vascular endothelial dysfunction. This could prove a potential target for treating severe COVID-19 in older adults.
• Mosquito-borne alphaviruses in Zambia: Isolation and characterization of Eilat and Sindbis viruses.
  Chadwic De'Sean Mears; Koshiro Tabata; Takuma Ariizumi; Bernard M Hang'ombe; Yongjin Qiu; Hayato Harima; Masahiro Kajihara; William W Hall; Michihito Sasaki; Hirofumi Sawa; Yasuko Orba
  Virus research, 358, 199604, 199604, Aug. 2025, [International Magazine]
  English, Scientific journal, Alphaviruses in the family Togaviridae include zoonotic arthropod-borne viruses, including Sindbis virus (SINV), chikungunya virus, as well as insect-specific viruses such as Eilat virus (EILV). Previous investigations of alphaviruses in Zambia have identified a novel insect-specific alphavirus, Mwinilunga alphavirus in mosquitoes. Further ongoing surveillance resulted in the isolation of EILV and SINV for the first time in Zambia. Here, these alphaviruses were characterized in terms of growth kinetics in cells, and molecular phylogenetic relatedness to other alphaviruses. Zambian EILV (strain zmq19_M44) exhibited a close phylogenetic relationship with other insect-specific alphaviruses and shared a close nucleotide identity to those of EILV isolate (90.4 %) and Mwinilunga alphavirus (75.5 %). EILV zmq19_M44 attained a saturating titer in C6/36 cells at 6-8-days post infection but was unable to replicate in mammalian cells. Phylogenetic analysis revealed the Zambian SINV (strain zmq17_M115) belongs in Clade D of SINV Genotype 1 along with the Kenyan isolate BONI 584 from Central Africa. The growth of the SINV zmq17_M115 was comparable to that of the prototype SINV strain AR339 in mammalian cells but was statistically different in insect cells. Our findings will contribute to public health measures for the control of alphaviral diseases in Zambia.
• SARS-CoV-2 infection promotes lung thrombosis by inducing integrinβ3 expression in vascular endothelial cells.
  Wataru Ito; Yuya Sakurai; Nako Maishi; Ryo Takeda; Takahito Teshirogi; Li Yu; Yasuhiro Hida; Michihito Sasaki; Yasuko Orba; Takuya Tsumita; Haruhisa Watanabe; Tadahiro Iimura; Terufumi Kubo; Shinsuke Toba; Akihiko Sato; Aya Matsuda; Daisuke Kyuno; Makoto Osanai; Yoichi Ohiro; Toshihiko Torigoe; Hirofumi Sawa; Kyoko Hida
  Scientific reports, 15, 1, 20447, 20447, 01 Jul. 2025, [International Magazine]
  English, Scientific journal, Severe COVID-19 shows a high incidence of pulmonary thrombosis. However, the molecular mechanism underlying this phenomenon remains unclear. We have performed RNA sequencing of isolated endothelial cells (ECs) from infected mid-aged and young mice. Compared to young mice, Integrinβ3 (ITGB3) expression levels were higher in ECs of mid-aged mice which showed thrombosis in lungs. SARS-CoV-2 exposure increased the number of adhered platelets on the EC monolayer in vitro. Knockdown of ITGB3 in ECs decreased platelet adhesion to them. Among the molecules known as SARS-CoV-2 receptors, Kringle-containing transmembrane protein 1 contributed to ITGB3 upregulation in ECs by SARS-CoV-2. Histological analysis showed that ITGB3-positive blood vessels were frequently detected not only in infected-mid-aged mouse lungs but also in COVID-19-affected human autopsy lungs. This study suggests that the induction of ITGB3 expression in ECs is one of the mechanisms of thrombosis in severe COVID-19 pneumonia.
• Structural and virological identification of neutralizing antibody footprint provides insights into therapeutic antibody design against SARS-CoV-2 variants.
  Yuki Anraku; Shunsuke Kita; Taishi Onodera; Akihiko Sato; Takashi Tadokoro; Shiori Ito; Yu Adachi; Ryutaro Kotaki; Tateki Suzuki; Jiei Sasaki; Nozomi Shiwa-Sudo; Naoko Iwata-Yoshikawa; Noriyo Nagata; Souta Kobayashi; Yasuhiro Kazuki; Mitsuo Oshimura; Takao Nomura; Michihito Sasaki; Yasuko Orba; Tadaki Suzuki; Hirofumi Sawa; Takao Hashiguchi; Hideo Fukuhara; Yoshimasa Takahashi; Katsumi Maenaka
  Communications biology, 8, 1, 483, 483, 22 Mar. 2025, [International Magazine]
  English, Scientific journal, Medical treatments using potent neutralizing SARS-CoV-2 antibodies have achieved remarkable improvements in clinical symptoms, changing the situation for the severity of COVID-19 patients. We previously reported an antibody, NT-108 with potent neutralizing activity. However, the structural and functional basis for the neutralizing activity of NT-108 has not yet been understood. Here, we demonstrated the therapeutic effects of NT-108 in a hamster model and its protective effects at low doses. Furthermore, we determined the cryo-EM structure of NT-108 in complex with SARS-CoV-2 spike. The single-chain Fv construction of NT-108 improved the cryo-EM maps because of the prevention of preferred orientations induced by Fab orientation. The footprints of NT-108 illuminated how escape mutations such as E484K evade from class 2 antibody recognition without ACE2 affinity attenuation. The functional and structural basis for the potent neutralizing activity of NT-108 provides insights into the rational design of therapeutic antibodies.
• Malabaricone C isolated from edible plants as a potential inhibitor of SARS-CoV-2 infection.
  Mutmainah; Yuta Murai; Ai Fujimoto; Rintaro Kawamura; Akira Kitamura; Sajeer Koolath; Seigo Usuki; Michihito Sasaki; Yasuko Orba; Yasuyuki Igarashi; Hirofumi Sawa; Akihiko Sato; Kenji Monde
  Scientific reports, 15, 1, 8518, 8518, 12 Mar. 2025, [International Magazine]
  English, Scientific journal, Although the SARS-CoV-2 epidemic worldwide has gradually decreased, in some areas, the situation has not yet been stamped and has become a global health emergency. It is quite possible that we could again be threatened by a new coronavirus. Therefore, new nucleotide analog drugs and vaccines or using drug repositioning for SARS-CoV-2 still has been developed, yet their safety and efficacy against COVID-19 remains underexplored. Malabaricone C is 2,6-dihydroxyphenyl acylphenol found in edible plants such as the mace spice of nutmeg derived from the seeds of Myristica fragrans. In this study, we identified malabaricone C as the first inhibitor of SARS-CoV-2 from natural food with a safe alternative for drugs. Malabaricone C and its chemical derivatives showed EC50 values of 1-1.5 μM against SARS-CoV-2 (WK-521, ancestral strain) and its variant strains in mammalian cells (HEK293T and Vero E6). In addition, we have successfully established novel evaluation system for the inhibition of SARS virus cell fusion by visualization for providing a versatile tool for study SARS-CoV-2 mediated fusion. Furthermore, our experiments suggested that malabaricone C could affect the distribution of sphingomyelin on the plasma membrane, which involves in viral infections. Thus, in light of the beneficial effect of malabaricone C on viral infection, the nontoxic malabaricone C is a suitable candidate as a drug that can be employed in the treatment and prevention of COVID-19. Moreover, it may potentially be used to treat acute infections of other enveloped viruses.
• 加齢に伴う血管内皮細胞SARS-CoV-2感染増強メカニズムの解析(Analysis of the mechanism of enhanced vascular endothelial cell SARS-CoV-2 infection with aging)
  桜井 優弥; 間石 奈湖; 藤岡 容一朗; 大場 雄介; 佐々木 道仁; 大場 靖子; 樋田 泰浩; 澤 洋文; 樋田 京子
  日本病理学会会誌, 114, 1, 328, 328, (一社)日本病理学会, Mar. 2025
  English
• Risk Mapping of African Swine Fever in Domestic Pigs and Wild Boars to Enhance Management and Surveillance in Asia.
  Nijiho Kawaguchi; Cecilia Aguilar-Vega; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa; José Manuel Sánchez-Vizcaíno; Norikazu Isoda; Jaime Bosch; Satoshi Ito
  Transboundary and emerging diseases, 2025, 8850856, 8850856, 2025, [International Magazine]
  English, Scientific journal, African swine fever (ASF) is a highly lethal disease affecting domestic pigs and wild boars, caused by the ASF virus (ASFV), which has rapidly spread across Asia in recent years. In this region, most reported ASF cases involve domestic pigs, while cases in wild boars remain notably lower except in a few countries. However, factors such as the high population of wild boars, limited wildlife surveillance, and inadequate farm biosecurity suggest that the prevalence and transmission of ASFV between these hosts may be underestimated. Therefore, we used a simplified multicriteria approach (SMCA) to identify vulnerable areas (VAs) for ASFV infection and validated the resulting VA maps with chi-square tests using reported ASF cases. The spatial SMCA revealed that VAs for ASFV infection in domestic pigs are concentrated in eastern China, while high-risk zones for ASFV infection in wild boars span Russia, eastern China, and Southeast Asia. Sensitity analysis showed that the variables that most influenced the risk of ASFV infection in domestic pigs and wild boars were anthropogenic factors and distribution of wild boars, respectively. Additionally, we predicted areas with significant transmission potential between domestic pigs and wild boars. High-risk regions for interspecies transmission include eastern China, southwestern Korea, and southern Japan. This study offers a standardized method to assess ASFV infection risk across Asia by integrating environmental and anthropogenic factors rather than relying solely on reported outbreaks. The findings highlight potential high-risk regions, including those without detected outbreaks, to improve surveillance and early detection strategies.
• First identification and whole genome characterization of rotavirus C in pigs in Zambia.
  Hayato Harima; Yongjin Qiu; Michihito Sasaki; Joseph Ndebe; Kapila Penjaninge; Edgar Simulundu; Masahiro Kajihara; Aiko Ohnuma; Keita Matsuno; Naganori Nao; Yasuko Orba; Ayato Takada; Kanako Ishihara; William W Hall; Bernard M Hang'ombe; Hirofumi Sawa
  Virology, 603, 110385, 110385, 31 Dec. 2024, [International Magazine]
  English, Scientific journal, Rotavirus C (RVC) causes acute gastroenteritis in neonatal piglets. Despite the clinical importance of RVC infection, the distribution and prevalence in pig populations in most African countries remains unknown. In this study, we identified RVC in Zambian pigs by metagenomic analysis. The full genome sequence of the RVC revealed two different VP4 sequences, implying that two different RVC strains (ZP18-77-c1 and ZP18-77-c2) were present in the same sample. Genetic analyses demonstrated that all segments of ZP18-77-c1 and ZP18-77-c2 showed high nucleotide sequence identities (87.7-94.5%) to known porcine RVC strains, and ZP18-77-c1 and ZP18-77-c2 strains were assigned to genotype constellations, G1-P[4]/P[14]-I13-R5-C5-M1-A7-N9-T10-E5-H1. We further screened RVC genomes among pig feces collected in Zambia (n = 147) by RT-qPCR, and 78 samples (53.1%) were positive. This study demonstrated the first full genome sequence of African RVC strains with a relatively high prevalence of RVC infection in the pig populations in Zambia.
• Neuron-associated retroelement-derived protein Arc/Arg3.1 assists in the early stages of alphaherpesvirus infection in human neuronal cells
  Hiroko Kobayashi; Mitsuki Yasukochi; Masayuki Horie; Yasuko Orba; Hirofumi Sawa; Kan Fujino; Satoshi Taharaguchi
  PLOS ONE, 19, 12, e0314980, 12 Dec. 2024, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Alphaherpesviruses, including herpes simplex virus type 1 (HSV-1) and pseudorabies virus (PRV), are neurotropic double-stranded DNA viruses. Alphaherpesviruses control the expression of various host factors to ensure efficient infection and propagation. Recently, HSV-1 was found to upregulate Arc/Arg3.1 (Arc) expression, which is a retroelement-derived domesticated gene. Arc is associated with learning and neuroplasticity in host neuronal cells, and its abnormal expression leads to neurological disorders. However, the detailed mechanisms underlying the upregulation of Arc and its physiological significance in viral infections remain unclear. In this study, we found that PRV infection upregulated Arc expression in vitro and identified ICP0 and EP0, the transcriptional regulatory genes of HSV-1 and PRV, as triggers for enhanced Arc expression. Mass spectrometry and co-immunoprecipitation assays identified VP5, the major capsid protein of PRV and HSV-1, as the viral factor that interacted with Arc. Arc knockdown delayed viral infection during the early stages of the viral life cycle, but did not impact the viral attachment and entry. In conclusion, we provide evidence that alphaherpesvirus ICP0 homologues control Arc expression. Additionally, we demonstrate that Arc interacts with the major capsid protein VP5 and plays an important role in the viral lifecycle after intracellular entry. This study advances our knowledge of herpesvirus and retroelement-derived Arc interactions, providing fundamental insights into the pathogenesis of retroelement-derived domesticated genes and herpesvirus-induced neurological diseases.
• X-206 exhibits broad-spectrum anti-β-coronavirus activity, covering SARS-CoV-2 variants and drug-resistant isolates.
  Jiei Sasaki; Akihiko Sato; Michihito Sasaki; Iori Okabe; Kota Kodama; Satoko Otsuguro; Kosuke Yasuda; Hirotatsu Kojima; Yasuko Orba; Hirofumi Sawa; Katsumi Maenaka; Yusuke Yanagi; Takao Hashiguchi
  Antiviral research, 232, 106039, 106039, 19 Nov. 2024, [International Magazine]
  English, Scientific journal, Coronaviruses such as the Middle East respiratory syndrome coronavirus (MERS-CoV), severe acute respiratory syndrome coronavirus (SARS-CoV), and SARS-CoV-2, causing MERS, SARS, and Coronavirus disease-19, respectively, are highly pathogenic to humans. Notably, several antiviral drugs against SARS-CoV-2, such as nirmatrelvir and remdesivir, have been approved. However, no approved vaccines or antiviral agents are available for other highly pathogenic β-coronaviruses. In this study, we identified two compounds, thapsigargin and X-206, that exhibit antiviral activities against SARS-CoV, MERS-CoV, and SARS-CoV-2. Notably, both compounds effectively inhibited the cell-to-cell fusion mediated by the Spike proteins of all three β-coronaviruses. X-206 exhibited antiviral activity against nirmatrelvir- and remdesivir-resistant SARS-CoV-2 isolates and SARS-CoV-2 variants, including Delta, BA.5, and XBB.1. Consequently, the mechanism of action of these compounds with anti-β-coronavirus activities may differ from that of the approved direct-acting drugs for SARS-CoV-2, thereby offering potential use as a cocktail with other antivirals, and serving as a chemical basis for developing therapeutic agents against β-coronaviruses in preparation for the next spillover and pandemic.
• African lineage 1a West Nile virus isolated from crocodiles exhibits low neuroinvasiveness in mice.
  Hiroko Kobayashi; Herman Chambaro; Koshiro Tabata; Takuma Ariizumi; Wallaya Phongphaew; Kunda Ndashe; Joseph Ndebe; Paul Fandamu; Shintaro Kobayashi; Naoto Ito; Michihito Sasaki; Bernard M Hang'ombe; Edgar Simulundu; Yasuko Orba; Hirofumi Sawa
  The Journal of general virology, 105, 11, Nov. 2024, [International Magazine]
  English, Scientific journal, West Nile virus (WNV) is a mosquito-borne flavivirus that causes encephalitis in humans and infects crocodiles, resulting in rashes and neurological signs. In Zambia, two distinct lineages of WNV have been detected in neighbouring areas: lineage 2 in mosquitoes and lineage 1a in farmed crocodiles. Considering the risk of direct or vector-mediated WNV transmission from crocodiles to mammals, it is necessary to elucidate the pathogenicity of WNV strains derived from crocodiles. In this study, WNV was successfully isolated from naturally infected farmed crocodiles (Croc110/2019/1/ZM, Croc110). We then investigated its proliferation and pathogenicity in mice in comparison with a WNV isolate from mosquitoes in Zambia (Zmq16) and two reference strains, including one highly pathogenic (NY99) and one low pathogenic (Eg101) strain. Although viral proliferation in Vero and mammalian neuronal cells was comparable among the strains, Croc110 exhibited low cell-to-cell transmission efficiency. In vivo, more than 70% of mice (C57BL/6) intracerebrally inoculated with Croc110 displayed neurological signs, and Croc110-infected mice exhibited similarly high mortality rates as NY99- and Zmq16-infected mice. Meanwhile, comparable virus growth was observed among the strains in the brain. However, the virulence of Croc110 was significantly lower than that of Zmq16 and NY99 following intradermal (ID) and intraperitoneal inoculation. Consistently, Croc110 displayed lower growth than Zmq16 and NY99 in the brain and peripheral tissues after ID inoculation. Our study revealed that the crocodile-derived WNV strain is less neuroinvasive in mice, and it exhibits distinct pathogenicity from the highly pathogenic mosquito-derived WNV strain circulating in Zambia.
• A first report of rotavirus B from Zambian pigs leading to the discovery of a novel VP4 genotype P[9].
  Hayato Harima; Yongjin Qiu; Michihito Sasaki; Joseph Ndebe; Kapila Penjaninge; Edgar Simulundu; Masahiro Kajihara; Aiko Ohnuma; Keita Matsuno; Naganori Nao; Yasuko Orba; Ayato Takada; Kanako Ishihara; William W Hall; Bernard M Hang'ombe; Hirofumi Sawa
  Virology journal, 21, 1, 263, 263, 24 Oct. 2024, [International Magazine]
  English, Scientific journal, BACKGROUND: Rotavirus B (RVB) causes diarrhea in humans and pigs. Although various RVB strains were identified in humans and various animals globally, little is known about the epidemiology RVB infection in Africa. In this study, we attempted to examine the prevalence of RVB infection in pig populations in Zambia. METHODS: Metagenomic analyses were conducted on pig feces collected in Zambia to detect double stranded RNA viruses, including RVB. To clarify the prevalence of RVB infection in pig populations in Zambia, 147 fecal samples were screened for the RVB detection by RT-qPCR. Full genome sequence of a detected RVB was determined by Sanger sequencing and genetically analyzed. RESULTS: The metagenomic analyses revealed that RVB sequence reads and contigs of RVB were detected from one fecal sample collected from pigs in Zambia. RT-qPCR screening detected RVB genomes in 36.7% (54/147) of fecal samples. Among 54 positive samples, 13 were positive in non-diarrheal samples (n = 48, 27.1%) and 41 in diarrheal samples (n = 99, 41.4%). Genetic analyses demonstrated that all the segments of ZP18-18, except for VP4, had high nucleotide sequence identities (80.6-92.6%) with all other known RVB strains detected in pigs. In contrast, the VP4 sequence of ZP18-18 was highly divergent from other RVB strains (< 64.6% identities) and formed a distinct lineage in the phylogenetic tree. Notably, the VP8 subunit of the VP4 showed remarkably low amino acid identities (33.3%) to those of known RVB strains, indicating that the VP8 subunit of ZP18-18 was unique among RVB strains. According to the whole genome classification for RVB, ZP18-18 was assigned to a genotype constellation, G18-P[9]-I12-R4-C4-M4-A8-N10-T5-E4-H7 with the newly established VP4 genotype P[9]. CONCLUSIONS: This current study updates the geographical distribution and the genetic diversity of RVB. Given the lack of information regarding RVB in Africa, further RVB surveillance is required to assess the potential risk to humans and animals.
• Characterization of a mammalian orthoreovirus isolated from the large flying fox, Pteropus vampyrus, in Indonesia.
  Kittiya Intaruck; Koshiro Tabata; Yukari Itakura; Nijiho Kawaguchi; Mai Kishimoto; Agus Setiyono; Ekowati Handharyani; Hayato Harima; Takashi Kimura; William W Hall; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki
  The Journal of general virology, 105, 9, Sep. 2024, [International Magazine]
  English, Scientific journal, Fruit bats serve as an important reservoir for many zoonotic pathogens, including Nipah virus, Hendra virus, Marburg virus and Lyssavirus. To gain a deeper insight into the virological characteristics, pathogenicity and zoonotic potential of bat-borne viruses, recovery of infectious viruses from field samples is important. Here, we report the isolation and characterization of a mammalian orthoreovirus (MRV) from a large flying fox (Pteropus vampyrus) in Indonesia, which is the first detection of MRV in Southeast Asia. MRV was recovered from faecal samples of three different P. vampyrus in Central Java. Nucleotide sequence analysis revealed that the genome of the three MRV isolates shared more than 99% nucleotide sequence identity. We tentatively named one isolated strain as MRV12-52 for further analysis and characterization. Among 10 genome segments, MRV12-52 S1 and S4, which encode the cell-attachment protein and outer capsid protein, had 93.6 and 95.1% nucleotide sequence identities with known MRV strains, respectively. Meanwhile, the remaining genome segments of MRV12-52 were divergent with 72.9-80.7 % nucleotide sequence identities. Based on the nucleotide sequence of the S1 segment, MRV12-52 was grouped into serotype 2, and phylogenetic analysis demonstrated evidence of past reassortment events. In vitro characterization of MRV12-52 showed that the virus efficiently replicated in BHK-21, HEK293T and A549 cells. In addition, experimental infection of laboratory mice with MRV12-52 caused severe pneumonia with 75% mortality. This study highlights the presence of pathogenic MRV in Indonesia, which could serve as a potential animal and public health concern.
• β-d-N4-hydroxycytidine, a metabolite of molnupiravir, exhibits in vitro antiviral activity against rabies virus.
  Kei Konishi; Shinji Kusakabe; Nijiho Kawaguchi; Takao Shishido; Naoto Ito; Michiko Harada; Satoshi Inoue; Ken Maeda; William W Hall; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki; Akihiko Sato
  Antiviral research, 229, 105977, 105977, Sep. 2024, [International Magazine]
  English, Scientific journal, Rabies is a fatal neurological disorder caused by rabies virus (RABV) infection. Approximately 60,000 patients die from rabies annually, and there are no effective treatments for this disease. Nucleoside analogs are employed as antiviral drugs based on their broad antiviral spectrum, and certain nucleoside analogs have been reported to exhibit anti-RABV activity. The nucleoside analog β-d-N4-hydroxycytidine (NHC) has antiviral effects against a range of RNA viruses. Molnupiravir (MPV), a prodrug of NHC, is clinically used as an oral antiviral drug for coronavirus infections. Despite its broad-spectrum activity, the antiviral activity of NHC against RABV remains unclear. In this study, we reveal that NHC exhibits comparable in vitro anti-RABV activity as ribavirin and favipiravir (also known as T-705) with a 90% effective concentration of 6 μM in mouse neuroblastoma cells. NHC reduced viral loads in neuronal and nonneuronal cells in a dose-dependent manner. Both laboratory and field RABVs (fixed and street strains, respectively) were susceptible to NHC. However, no increase in survival or reduction in viral titers in the brain was observed in RABV-infected mice treated prophylactically with MPV. These findings highlight the potential and challenges of NHC in the treatment of RABV infection.
• Increased production of orthoflavivirus single-round infectious particles produced in mammalian cells at a suboptimal culture temperature of 28°C
  Koshiro Tabata; Shintaro Kobayashi; Yukari Itakura; Gabriel Gonzalez; Chilekwa F. Kabamba; Shinji Saito; Michihito Sasaki; William W. Hall; Hirofumi Sawa; Yasuko Orba
  Journal of Virological Methods, 329, 115007, 115007, Elsevier BV, Sep. 2024, [International Magazine]
  English, Scientific journal, In the employment of serodiagnostic methods for the detection of orthoflavivirus infections, neutralization tests are known to be more accurate than measurements of antibody binding properties employing enzyme-linked immunosorbent assays. However, neutralization tests require infectious virus and laboratories with an appropriate level of biosafety. Single-round infectious particles (SRIPs), which encode a reporter gene instead of the viral structural protein genes, are replication incompetent and represent a safe and reliable alternative to the diagnosis of pathogenic viruses in neutralization tests. The orthoflavivirus SRIPs are produced by co-transfection of plasmids expressing virus-like particles and replicons into mammalian cell lines preferably with high transfection efficacy, such as HEK293T cells. However, certain orthoflavivirus SRIPs have limitations in their efficient expression at 37°C, which is the optimal temperature for mammalian cell growth, resulting in insufficient yields for neutralization tests. Here, we demonstrate that the production of orthoflavivirus SRIPs increases at the lower temperature of 28°C compared to 37°C. Moreover, infections with 28°C-cultured SRIPs in microneutralization tests were specifically inhibited in the presence of serum from mice infected with homologous viruses, suggesting that these SRIPs preserved their neutralizing epitopes for antibodies. Our method to produce high titer SRIPs is anticipated to promote efficient and safe SRIPs neutralization tests as a general serodiagnostic method for detecting virus-specific neutralizing antibodies against orthoflaviviruses.
• Reverse genetic approaches allowing the characterization of the rabies virus street strain belonging to the SEA4 subclade.
  Nijiho Kawaguchi; Yukari Itakura; Kittiya Intaruck; Takuma Ariizumi; Michiko Harada; Satoshi Inoue; Ken Maeda; Naoto Ito; William W Hall; Hirofumi Sawa; Yasuko Orba; Michihito Sasaki
  Scientific reports, 14, 1, 18509, 18509, 09 Aug. 2024, [International Magazine]
  English, Scientific journal, Rabies virus (RABV) is the causative agent of rabies, a lethal neurological disease in mammals. RABV strains can be classified into fixed strains (laboratory strains) and street strains (field/clinical strains), which have different properties including cell tropism and neuroinvasiveness. RABV Toyohashi strain is a street strain isolated in Japan from an imported case which had been bitten by rabid dog in the Philippines. In order to facilitate molecular studies of RABV, we established a reverse genetics (RG) system for the study of the Toyohashi strain. The recombinant virus was obtained from a cDNA clone of Toyohashi strain and exhibited similar growth efficiency as the original virus in cultured cell lines. Both the original and recombinant strains showed similar pathogenicity with high neuroinvasiveness in mice, and the infected mice developed a long and inconsistent incubation period, which is characteristic of street strains. We also generated a recombinant Toyohashi strain expressing viral phosphoprotein (P protein) fused with the fluorescent protein mCherry, and tracked the intracellular dynamics of the viral P protein using live-cell imaging. The presented reverse genetics system for Toyohashi strain will be a useful tool to explore the fundamental molecular mechanisms of the replication of RABV street strains.
• A wide distribution of Beiji nairoviruses and related viruses in Ixodes ticks in Japan.
  Mai Kishimoto; Yukari Itakura; Koshiro Tabata; Rika Komagome; Hiroki Yamaguchi; Kohei Ogasawara; Ryo Nakao; Yongjin Qiu; Kozue Sato; Hiroki Kawabata; Masahiro Kajihara; Naota Monma; Junji Seto; Asako Shigeno; Masayuki Horie; Michihito Sasaki; William W Hall; Hirofumi Sawa; Yasuko Orba; Keita Matsuno
  Ticks and tick-borne diseases, 15, 6, 102380, 102380, 11 Jul. 2024, [International Magazine]
  English, Scientific journal, Beiji nairovirus (BJNV), in the family Nairoviridae, the order Bunyavirales, was recently reported as a causative agent of an emerging tick-borne zoonotic infection in China. This study investigated the prevalence of BJNV in ticks in Japan. Screening of over 2,000 ticks from multiple regions revealed a widespread distribution of BJNV and BJNV-related viruses in Japan, particularly in the northern island, and in other high altitude areas with exclusive occurrence of Ixodes ticks. Phylogenetic analysis identified three distinct groups of nairoviruses in ticks in Japan: BJNV, Yichun nairovirus (YCNV) and a newly identified Mikuni nairovirus (MKNV). BJNV and YCNV variants identified in ticks in Japan exhibited high nucleotide sequence identities to those in China and Russia with evidence of non-monophyletic evolution among BJNVs, suggesting multiple cross-border transmission events of BJNV between the Eurasian continent and Japan. Whole genome sequencing of BJNV and MKNV revealed a unique GA-rich region in the S segment, the significance of which remains to be determined. In conclusion, the present study has shown a wide distribution and diversity of BJNV-related nairoviruses in Ixodes ticks in Japan and has identified unique genomic structures. The findings demonstrate the significance of BJNV as well as related viruses in Japan and highlight the necessity of monitoring emerging nairovirus infections and their potential risks to public health.
• Molecular evolution of Hokkaido virus, a genotype of Orthohantavirus puumalaense, among Myodes rodents.
  Duong Thi Ngoc Thuy; Michihito Sasaki; Yasuko Orba; Passawat Thammahakin; Keisuke Maezono; Shintaro Kobayashi; Hiroaki Kariwa
  Virology, 597, 110168, 110168, 03 Jul. 2024, [International Magazine]
  English, Scientific journal, Viruses in the genus Orthohantavirus within the family Hantaviridae cause human hantavirus infections and represent a threat to public health. Hokkaido virus (HOKV), a genotype of Orthohantavirus puumalaense (Puumala virus; PUUV), was first identified in Tobetsu, Hokkaido, Japan. Although it is genetically related to the prototype of PUUV, the evolutionary pathway of HOKV is unclear. We conducted a field survey in a forest in Tobetsu in 2022 and captured 44 rodents. Complete coding genome sequences of HOKVs were obtained from five viral-RNA-positive rodents (four Myodes rufocanus bedfordiae and one Apodemus speciosus). Phylogenetic analysis revealed a close relationship between the phylogenies and geographical origins of M. rufocanus-related orthohantaviruses. Comparison of the phylogenetic trees of the S segments of orthohantaviruses and the cytochrome b genes of Myodes species suggested that Myodes-related orthohantaviruses evolved in Myodes rodent species as a result of genetic isolation and host switching.
• Genes involved in the limited spread of SARS-CoV-2 in the lower respiratory airways of hamsters may be associated with adaptive evolution.
  Kosuke Takada; Yasuko Orba; Yurie Kida; Jiaqi Wu; Chikako Ono; Yoshiharu Matsuura; So Nakagawa; Hirofumi Sawa; Tokiko Watanabe
  Journal of virology, 98, 5, e0178423, 16 Apr. 2024, [International Magazine]
  English, Scientific journal, Novel respiratory viruses can cause a pandemic and then evolve to coexist with humans. The Omicron strain of severe acute respiratory syndrome coronavirus 2 has spread worldwide since its emergence in late 2021, and its sub-lineages are now established in human society. Compared to previous strains, Omicron is markedly less invasive in the lungs and causes less severe disease. One reason for this is that humans are acquiring immunity through previous infection and vaccination, but the nature of the virus itself is also changing. Using our newly established low-volume inoculation system, which reflects natural human infection, we show that the Omicron strain spreads less efficiently into the lungs of hamsters compared with an earlier Wuhan strain. Furthermore, by characterizing chimeric viruses with the Omicron gene in the Wuhan strain genetic background and vice versa, we found that viral genes downstream of ORF3a, but not the S gene, were responsible for the limited spread of the Omicron strain in the lower airways of the virus-infected hamsters. Moreover, molecular evolutionary analysis of SARS-CoV-2 revealed a positive selection of genes downstream of ORF3a (M and E genes). Our findings provide insight into the adaptive evolution of the virus in humans during the pandemic convergence phase.IMPORTANCEThe severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant has spread worldwide since its emergence in late 2021, and its sub-lineages are established in human society. Compared to previous strains, the Omicron strain is less invasive in the lower respiratory tract, including the lungs, and causes less severe disease; however, the mechanistic basis for its restricted replication in the lower airways is poorly understood. In this study, using a newly established low-volume inoculation system that reflects natural human infection, we demonstrated that the Omicron strain spreads less efficiently into the lungs of hamsters compared with an earlier Wuhan strain and found that viral genes downstream of ORF3a are responsible for replication restriction in the lower respiratory tract of Omicron-infected hamsters. Furthermore, we detected a positive selection of genes downstream of ORF3a (especially the M and E genes) in SARS-CoV-2, suggesting that these genes may undergo adaptive changes in humans.
• Establishment of a lethal mouse model of emerging tick-borne orthonairovirus infections.
  Takuma Ariizumi; Koshiro Tabata; Yukari Itakura; Hiroko Kobayashi; William W Hall; Michihito Sasaki; Hirofumi Sawa; Keita Matsuno; Yasuko Orba
  PLoS pathogens, 20, 3, e1012101, Mar. 2024, [International Magazine]
  English, Scientific journal, Emerging and reemerging tick-borne virus infections caused by orthonairoviruses (family Nairoviridae), which are genetically distinct from Crimean-Congo hemorrhagic fever virus, have been recently reported in East Asia. Here, we have established a mouse infection model using type-I/II interferon receptor-knockout mice (AG129 mice) both for a better understanding of the pathogenesis of these infections and validation of antiviral agents using Yezo virus (YEZV), a novel orthonairovirus causing febrile illnesses associated with tick bites in Japan and China. YEZV-inoculated AG129 mice developed hepatitis with body weight loss and died by 6 days post infection. Blood biochemistry tests showed elevated liver enzyme levels, similar to YEZV-infected human patients. AG129 mice treated with favipiravir survived lethal YEZV infection, demonstrating the anti-YEZV effect of this drug. The present mouse model will help us better understand the pathogenicity of the emerging tick-borne orthonairoviruses and the development of specific antiviral agents for their treatment.
• Immunogenicity and protective efficacy of a co-formulated two-in-one inactivated whole virus particle COVID-19/influenza vaccine.
  Chimuka Handabile; Marumi Ohno; Toshiki Sekiya; Naoki Nomura; Tomomi Kawakita; Mamiko Kawahara; Masafumi Endo; Tomohiro Nishimura; Minako Okumura; Shinsuke Toba; Michihito Sasaki; Yasuko Orba; Brendon Y Chua; Louise C Rowntree; Thi H O Nguyen; Masashi Shingai; Akihiko Sato; Hirofumi Sawa; Kazumasa Ogasawara; Katherine Kedzierska; Hiroshi Kida
  Scientific reports, 14, 1, 4204, 4204, 20 Feb. 2024, [International Magazine]
  English, Scientific journal, Due to the synchronous circulation of seasonal influenza viruses and severe acute respiratory coronavirus 2 (SARS-CoV-2) which causes coronavirus disease 2019 (COVID-19), there is need for routine vaccination for both COVID-19 and influenza to reduce disease severity. Here, we prepared individual WPVs composed of formalin-inactivated SARS-CoV-2 WK 521 (Ancestral strain; Co WPV) or influenza virus [A/California/07/2009 (X-179A) (H1N1) pdm; Flu WPV] to produce a two-in-one Co/Flu WPV. Serum analysis from vaccinated mice revealed that a single dose of Co/Flu WPV induced antigen-specific neutralizing antibodies against both viruses, similar to those induced by either type of WPV alone. Following infection with either virus, mice vaccinated with Co/Flu WPV showed no weight loss, reduced pneumonia and viral titers in the lung, and lower gene expression of proinflammatory cytokines, as observed with individual WPV-vaccinated. Furthermore, a pentavalent vaccine (Co/qFlu WPV) comprising of Co WPV and quadrivalent influenza vaccine (qFlu WPV) was immunogenic and protected animals from severe COVID-19. These results suggest that a single dose of the two-in-one WPV provides efficient protection against SARS-CoV-2 and influenza virus infections with no evidence of vaccine interference in mice. We propose that concomitant vaccination with the two-in-one WPV can be useful for controlling both diseases.
• 血管内皮細胞におけるSARS-CoV-2侵入機構の解析(Analysis of SARS-CoV-2 uptake mechanism in vascular endothelial cells)
  桜井 優弥; 間石 奈湖; 藤岡 容一郎; 大場 雄介; 武田 遼; 佐々木 道仁; 大場 靖子; 樋田 泰浩; 澤 洋文; 樋田 京子
  日本病理学会会誌, 113, 1, 369, 369, (一社)日本病理学会, Feb. 2024
  English
• Combination therapy with oral antiviral and anti-inflammatory drugs improves the efficacy of delayed treatment in a COVID-19 hamster model.
  Michihito Sasaki; Tatsuki Sugi; Shun Iida; Yuichiro Hirata; Shinji Kusakabe; Kei Konishi; Yukari Itakura; Koshiro Tabata; Mai Kishimoto; Hiroko Kobayashi; Takuma Ariizumi; Kittiya Intaruck; Haruaki Nobori; Shinsuke Toba; Akihiko Sato; Keita Matsuno; Junya Yamagishi; Tadaki Suzuki; William W Hall; Yasuko Orba; Hirofumi Sawa
  EBioMedicine, 99, 104950, 104950, 29 Dec. 2023, [International Magazine]
  English, Scientific journal, BACKGROUND: Pulmonary infection with SARS-CoV-2 stimulates host immune responses and can also result in the progression of dysregulated and critical inflammation. Throughout the pandemic, the management and treatment of COVID-19 has been continuously updated with a range of antiviral drugs and immunomodulators. Monotherapy with oral antivirals has proven to be effective in the treatment of COVID-19. However, treatment should be initiated in the early stages of infection to ensure beneficial therapeutic outcomes, and there is still room for further consideration on therapeutic strategies using antivirals. METHODS: We studied the therapeutic effects of monotherapy with the oral antiviral ensitrelvir or the anti-inflammatory corticosteroid methylprednisolone and combination therapy with ensitrelvir and methylprednisolone in a delayed dosing model of hamsters infected with SARS-CoV-2. FINDINGS: Combination therapy with ensitrelvir and methylprednisolone improved respiratory conditions and reduced the development of pneumonia in hamsters even when the treatment was started after 2 days post-infection. The combination therapy led to a differential histological and transcriptomic pattern in comparison to either of the monotherapies, with reduced lung damage and down-regulation of expression of genes involved in the inflammatory response. Furthermore, we found that the combination treatment is effective in case of infection with either the highly pathogenic delta or circulating omicron variants. INTERPRETATION: Our results demonstrate the advantage of combination therapy with antiviral and corticosteroid drugs in COVID-19 treatment from the perspective of lung pathology and host inflammatory responses. FUNDING: Funding bodies are described in the Acknowledgments section.
• Viral uptake and pathophysiology of the lung endothelial cells in age‐associated severe SARS‐CoV‐2 infection models
  Takuya Tsumita; Ryo Takeda; Nako Maishi; Yasuhiro Hida; Michihito Sasaki; Yasuko Orba; Akihiko Sato; Shinsuke Toba; Wataru Ito; Takahito Teshirogi; Yuya Sakurai; Tomohiro Iba; Hisamichi Naito; Hitoshi Ando; Haruhisa Watanabe; Amane Mizuno; Toshiki Nakanishi; Aya Matsuda; Ren Zixiao; Ji‐Won Lee; Tadahiro Iimura; Hirofumi Sawa; Kyoko Hida
  Aging Cell, e14050, 2, e14050, Wiley, 14 Dec. 2023, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Abstract
  
  Thrombosis is the major cause of death in severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) infection, and the pathology of vascular endothelial cells (ECs) has received much attention. Although there is evidence of the infection of ECs in human autopsy tissues, their detailed pathophysiology remains unclear due to the lack of animal model to study it. We used a mouse‐adapted SARS‐CoV‐2 virus strain in young and mid‐aged mice. Only mid‐aged mice developed fatal pneumonia with thrombosis. Pulmonary ECs were isolated from these infected mice and RNA‐Seq was performed. The pulmonary EC transcriptome revealed that significantly higher levels of viral genes were detected in ECs from mid‐aged mice with upregulation of viral response genes such as DDX58 and IRF7. In addition, the thrombogenesis‐related genes encoding PLAT, PF4, F3 PAI‐1, and P‐selectin were upregulated. In addition, the inflammation‐related molecules such as CXCL2 and CXCL10 were upregulated in the mid‐aged ECs upon viral infection. Our mouse model demonstrated that SARS‐CoV‐2 virus entry into aged vascular ECs upregulated thrombogenesis and inflammation‐related genes and led to fatal pneumonia with thrombosis. Current results of EC transcriptome showed that EC uptake virus and become thrombogenic by activating neutrophils and platelets in the aged mice, suggesting age‐associated EC response as a novel finding in human severe COVID‐19.
• Ubiquitin accumulation induced by the finger and palm sub-domains of NS5 modulates the replication of West Nile virus.
  Shintaro Kobayashi; Ryoko Kawakami; Chisaki Takeda; Keisuke Maezono; Passawat Thammahakin; Haruto Eguchi; Bernard M Hang'ombe; Yasuko Orba; Hirofumi Sawa; Kentaro Yoshii; Hiroaki Kariwa
  Virology, 588, 109902, 109902, Nov. 2023, [International Magazine]
  English, Scientific journal, West Nile virus (WNV) causes encephalitis in human and animals. WNV is phylogenetically classified into at least five distinct genetic lineages with different pathogenicity. The pathogenesis of West Nile encephalitis is affected by ubiquitin accumulation in infected cells, but the mechanism is unknown. In this study, the association between ubiquitin accumulation and WNV pathogenicity was investigated. Ubiquitin accumulation was detected in cells infected with NY99 strain belonging to lineage-1, but not FCG and Zmq16 strains belonging to lineage-2. Substitution of the Finger and Palm sub-domains of NS5 from lineage-1 to -2 decreased ubiquitin accumulation and viral replication. Furthermore, the survival rate was increased, and viral replication and ubiquitin accumulation in the brain were attenuated, in mice inoculated with the substituted WNV compared with lineage-1 WNV. Therefore, the intracellular ubiquitin accumulation induced by the Finger and Palm sub-domains of NS5 is linked to the differences in pathogenicity among WNV lineages.
• Expanding diversity of bunyaviruses identified in mosquitoes.
  Yasuko Orba; Yusuf Eshimutu Abu; Herman M Chambaro; Tapiwa Lundu; Walter Muleya; Yuki Eshita; Yongjin Qiu; Hayato Harima; Masahiro Kajihara; Akina Mori-Kajihara; Keita Matsuno; Michihito Sasaki; William W Hall; Bernard M Hang'ombe; Hirofumi Sawa
  Scientific reports, 13, 1, 18165, 18165, 24 Oct. 2023, [International Magazine]
  English, Scientific journal, Mosquitoes interact with various organisms in the environment, and female mosquitoes in particular serve as vectors that directly transmit a number of microorganisms to humans and animals by blood-sucking. Comprehensive analysis of mosquito-borne viruses has led to the understanding of the existence of diverse viral species and to the identification of zoonotic arboviruses responsible for significant outbreaks and epidemics. In the present study on mosquito-borne bunyaviruses we employed a broad-spectrum RT-PCR approach and identified eighteen different additional species in the Phenuiviridae family and also a number of related but unclassified bunyaviruses in mosquitoes collected in Zambia. The entire RNA genome segments of the newly identified viruses were further analyzed by RNA sequencing with a ribonuclease R (RNase R) treatment to reduce host-derived RNAs and enrich viral RNAs, taking advantage of the dsRNA panhandle structure of the bunyavirus genome. All three or four genome segments were identified in eight bunyavirus species. Furthermore, L segments of three different novel viruses related to the Leishbunyaviridae were found in mosquitoes together with genes from the suspected host, the Crithidia parasite. In summary, our virus detection approach using a combination of broad-spectrum RT-PCR and RNA sequencing analysis with a simple virus enrichment method allowed the discovery of novel bunyaviruses. The diversity of bunyaviruses is still expanding and studies on this will allow a better understanding of the ecology of hematophagous mosquitoes.
• 2-thiouridine is a broad-spectrum antiviral nucleoside analogue against positive-strand RNA viruses.
  Kentaro Uemura; Haruaki Nobori; Akihiko Sato; Shinsuke Toba; Shinji Kusakabe; Michihito Sasaki; Koshiro Tabata; Keita Matsuno; Naoyoshi Maeda; Shiori Ito; Mayu Tanaka; Yuki Anraku; Shunsuke Kita; Mayumi Ishii; Kayoko Kanamitsu; Yasuko Orba; Yoshiharu Matsuura; William W Hall; Hirofumi Sawa; Hiroshi Kida; Akira Matsuda; Katsumi Maenaka
  Proceedings of the National Academy of Sciences of the United States of America, 120, 42, e2304139120, 17 Oct. 2023, [International Magazine]
  English, Scientific journal, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections are causing significant morbidity and mortality worldwide. Furthermore, over 1 million cases of newly emerging or re-emerging viral infections, specifically dengue virus (DENV), are known to occur annually. Because no virus-specific and fully effective treatments against these or many other viruses have been approved, there is an urgent need for novel, effective therapeutic agents. Here, we identified 2-thiouridine (s2U) as a broad-spectrum antiviral ribonucleoside analogue that exhibited antiviral activity against several positive-sense single-stranded RNA (ssRNA+) viruses, such as DENV, SARS-CoV-2, and its variants of concern, including the currently circulating Omicron subvariants. s2U inhibits RNA synthesis catalyzed by viral RNA-dependent RNA polymerase, thereby reducing viral RNA replication, which improved the survival rate of mice infected with DENV2 or SARS-CoV-2 in our animal models. Our findings demonstrate that s2U is a potential broad-spectrum antiviral agent not only against DENV and SARS-CoV-2 but other ssRNA+ viruses.
• Development of flavivirus subviral particles with low cross-reactivity by mutations of a distinct antigenic domain.
  Koshiro Tabata; Yukari Itakura; Takuma Ariizumi; Manabu Igarashi; Hiroko Kobayashi; Kittiya Intaruck; Mai Kishimoto; Shintaro Kobayashi; William W Hall; Michihito Sasaki; Hirofumi Sawa; Yasuko Orba
  Applied microbiology and biotechnology, 107, 24, 7515, 7529, 13 Oct. 2023, [International Magazine]
  English, Scientific journal, The most conserved fusion loop (FL) domain present in the flavivirus envelope protein has been reported as a dominant epitope for cross-reactive antibodies to mosquito-borne flaviviruses (MBFVs). As a result, establishing accurate serodiagnosis for MBFV infections has been difficult as anti-FL antibodies are induced by both natural infection and following vaccination. In this study, we modified the most conserved FL domain to overcome this cross-reactivity. We showed that the FL domain of lineage I insect-specific flavivirus (ISFV) has differences in antigenicity from those of MBFVs and lineage II ISFV and determined the key amino acid residues (G106, L107, or F108), which contribute to the antigenic difference. These mutations were subsequently introduced into subviral particles (SVPs) of dengue virus type 2 (DENV2), Zika virus (ZIKV), Japanese encephalitis virus (JEV), and West Nile virus (WNV). In indirect enzyme-linked immunosorbent assays (ELISAs), these SVP mutants when used as antigens reduced the binding of cross-reactive IgG and total Ig induced by infection of ZIKV, JEV, and WNV in mice and enabled the sensitive detection of virus-specific antibodies. Furthermore, immunization of ZIKV or JEV SVP mutants provoked the production of antibodies with lower cross-reactivity to heterologous MBFV antigens compared to immunization with the wild-type SVPs in mice. This study highlights the effectiveness of introducing mutations in the FL domain in MBFV SVPs with lineage I ISFV-derived amino acids to produce SVP antigens with low cross-reactivity and demonstrates an improvement in the accuracy of indirect ELISA-based serodiagnosis for MBFV infections. KEY POINTS: • The FL domain of Lineage I ISFV has a different antigenicity from that of MBFVs. • Mutated SVPs reduce the binding of cross-reactive antibodies in indirect ELISAs. • Inoculation of mutated SVPs induces antibodies with low cross-reactivity.
• Unprecedented spike flexibility revealed by BSL3 Cryo-ET of active SARS-CoV-2 virions
  Hideo Fukuhara; Hisham M. Dokainish; Shunsuke Kita; Koshiro Tabata; Akira Takasu; Juha T. Huiskonen; Yuki Anraku; Toshiya Senda; David I. Stuart; Michihito Sasaki; Yasuko Orba; Yasuhiko Suzuki; Hirofumi Sawa; Katsumi Maenaka
  11 Oct. 2023
• Ecological Niche Modeling of Aedes and Culex Mosquitoes: A Risk Map for Chikungunya and West Nile Viruses in Zambia
  Rachel Milomba Velu; Geoffrey Kwenda; Prof. Samuel Bosomprah; Moses Ngongo Chisola; Michelo Simuyandi; Caroline Cleopatra Chisenga; Bumbangi Nsoni Flavien; Nicholus Chintu Sande; Limonty Simubali; Monicah Mirai Mburu; John Tembo; Matthew Bates; Roma Chilengi; Yasuko Orba; Hirofumi Sawa; Edgar Simulundu
  Viruses, 15, 9, Sep. 2023, [International Magazine]
  English, Scientific journal, The circulation of both West Nile Virus (WNV) and Chikungunya Virus (CHIKV) in humans and animals, coupled with a favorable tropical climate for mosquito proliferation in Zambia, call for the need for a better understanding of the ecological and epidemiological factors that govern their transmission dynamics in this region. This study aimed to examine the contribution of climatic variables to the distribution of Culex and Aedes mosquito species, which are potential vectors of CHIKV, WNV, and other arboviruses of public-health concern. Mosquitoes collected from Lusaka as well as from the Central and Southern provinces of Zambia were sorted by species within the Culex and Aedes genera, both of which have the potential to transmit viruses. The MaxEnt software was utilized to predict areas at risk of WNV and CHIKV based on the occurrence data on mosquitoes and environmental covariates. The model predictions show three distinct spatial hotspots, ranging from the high-probability regions to the medium- and low-probability regions. Regions along Lake Kariba, the Kafue River, and the Luangwa Rivers, as well as along the Mumbwa, Chibombo, Kapiri Mposhi, and Mpika districts were predicted to be suitable habitats for both species. The rainfall and temperature extremes were the most contributing variables in the predictive models.
• Morphogenesis of Bullet-Shaped Rabies Virus Particles Regulated by TSG101.
  Yukari Itakura; Koshiro Tabata; Takeshi Saito; Kittiya Intaruck; Nijiho Kawaguchi; Mai Kishimoto; Shiho Torii; Shintaro Kobayashi; Naoto Ito; Michiko Harada; Satoshi Inoue; Ken Maeda; Ayato Takada; William W Hall; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki
  Journal of virology, 97, 5, e0043823, 12 Apr. 2023, [International Magazine]
  English, Scientific journal, Viral protein assembly and virion budding are tightly regulated to enable the proper formation of progeny virions. At this late stage in the virus life cycle, some enveloped viruses take advantage of the host endosomal sorting complex required for transport (ESCRT) machinery, which contributes to the physiological functions of membrane modulation and abscission. Bullet-shaped viral particles are unique morphological characteristics of rhabdoviruses; however, the involvement of host factors in rhabdovirus infection and, specifically, the molecular mechanisms underlying virion formation are not fully understood. In the present study, we used a small interfering RNA (siRNA) screening approach and found that the ESCRT-I component TSG101 contributes to the propagation of rabies virus (RABV). We demonstrated that the matrix protein (M) of RABV interacts with TSG101 via the late domain containing the PY and YL motifs, which are conserved in various viral proteins. Loss of the YL motif in the RABV M or the downregulation of host TSG101 expression resulted in the intracellular aggregation of viral proteins and abnormal virus particle formation, indicating a defect in the RABV assembly and budding processes. These results indicate that the interaction of the RABV M and TSG101 is pivotal for not only the efficient budding of progeny RABV from infected cells but also for the bullet-shaped virion morphology. IMPORTANCE Enveloped viruses bud from cells with the host lipid bilayer. Generally, the membrane modulation and abscission are mediated by host ESCRT complexes. Some enveloped viruses utilize their late (L-) domain to interact with ESCRTs, which promotes viral budding. Rhabdoviruses form characteristic bullet-shaped enveloped virions, but the underlying molecular mechanisms involved remain elusive. Here, we showed that TSG101, one of the ESCRT components, supports rabies virus (RABV) budding and proliferation. TSG101 interacted with RABV matrix protein via the L-domain, and the absence of this interaction resulted in intracellular virion accumulation and distortion of the morphology of progeny virions. Our study reveals that virion formation of RABV is highly regulated by TSG101 and the virus matrix protein.
• Structure-activity relationship studies of anti-bunyaviral cap-dependent endonuclease inhibitors.
  Yoshiyuki Taoda; Akihiko Sato; Shinsuke Toba; Yuto Unoh; Makoto Kawai; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa
  Bioorganic & medicinal chemistry letters, 83, 129175, 129175, 01 Mar. 2023, [International Magazine]
  English, Scientific journal, Bunyaviruses, including the Lassa virus (LASV), are known to cause hemorrhagic fever and have a high fatality rate among hospitalized patients, as there are few effective treatments. We focused on the fact that bunyaviruses use cap-dependent endonuclease (CEN) for viral replication, which is similar to influenza viruses. This led us to screen carbamoyl pyridone bicycle (CAB) compounds, which compose a series of baloxavir acid (BXA) derivatives, against lymphocytic choriomeningitis virus (LCMV) and Junin virus (JUNV) among the bunyaviruses. This led to the discovery of 1c, which has potent anti-bunyaviral activities. In SAR studies, we found that a large lipophilic side chain is preferred for the 1-position of the CAB scaffold, similar to the influenza CEN inhibitor, and that a small alkyl group for the 3-position shows high activity. Moreover, the 7‑carboxyl group of the scaffold is essential for anti-bunyaviral activities, and the antiviral activity is reduced by conversion to various carboxylic acid bioisosteres. The SAR results are discussed using a binding model of 9d in the active center of the known LCMV CEN crystal structure. These compounds show promise as broad-spectrum anti-bunyavirus therapeutics, given their relatively favorable metabolic stability and PK profiles.
• Current status and molecular epidemiology of rabies virus from different hosts and regions in Malawi.
  Henson Kainga; Elisha Chatanga; Marvin Collen Phonera; John Pilate Kothowa; Precious Dzimbiri; Gladson Kamwendo; Malala Mulavu; Cynthia Sipho Khumalo; Katendi Changula; Herman Chambaro; Hayato Harima; Masahiro Kajihara; Kholiwe Mkandawire; Patrick Chikungwa; Julius Chulu; Gilson Njunga; Simbarashe Chitanga; Benjamin Mubemba; Michihito Sasaki; Yasuko Orba; Yongjin Qiu; Junya Yamagishi; Edgar Simulundu; Ayato Takada; Boniface Namangala; Hirofumi Sawa; Walter Muleya
  Archives of virology, 168, 2, 61, 61, 12 Jan. 2023, [International Magazine]
  English, Scientific journal, Although rabies is endemic in Malawi, there have been no studies in which rabies virus was systematically investigated and characterized in multiple animal hosts in that country. In order to provide molecular epidemiological data on rabies virus in Malawi, 683 suspected rabies case reports from 2008 to 2021 were examined, and 46 (dog = 40, cow = 5, and cat = 1) viable rabies-positive brain samples archived at the Central Veterinary Laboratory (CVL), Lilongwe, Malawi, were analyzed genetically. The results showed an increase in the submission of brain samples from 2008 to 2010, with the highest number of submissions observed in 2020. Of the 683 case reports analyzed for the period under review, 38.1% (260/683) (CI: 34.44 - 41.84) were confirmed by direct fluorescent antibody test. Among the confirmed cases, 65.4% (170/260) (CI: 59.23 - 71.09) were canine rabies. Further, phylogenetic analysis revealed that sequences from different animal hosts clustered together within the Africa 1b lineage, suggesting that the strains circulating in livestock are similar to those in domestic dogs. This finding supports the hypothesis that canine rabies is spilling over to livestock and emphasizes the need for further studies to provide data for effective control of rabies in Malawi.
• Isolation and Characterization of Distinct Rotavirus A in Bat and Rodent Hosts.
  Mai Kishimoto; Masahiro Kajihara; Koshiro Tabata; Yukari Itakura; Shinsuke Toba; Seiya Ozono; Yuko Sato; Tadaki Suzuki; Naoto Ito; Katendi Changula; Yongjin Qiu; Akina Mori-Kajihara; Yoshiki Eto; Hayato Harima; Daniel Mwizabi; Bernard M Hang'ombe; William W Hall; Ayato Takada; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki
  Journal of virology, 97, 1, e0145522, 12 Jan. 2023, [International Magazine]
  English, Scientific journal, Rotavirus A (RVA) causes diarrheal disease in humans and various animals. Recent studies have identified bat and rodent RVAs with evidence of zoonotic transmission and genome reassortment. However, the virological properties of bat and rodent RVAs with currently identified genotypes still need to be better clarified. Here, we performed virus isolation-based screening for RVA in animal specimens and isolated RVAs (representative strains: 16-06 and MpR12) from Egyptian fruit bat and Natal multimammate mouse collected in Zambia. Whole-genome sequencing and phylogenetic analysis revealed that the genotypes of bat RVA 16-06 were identical to that of RVA BATp39 strain from the Kenyan fruit bat, which has not yet been characterized. Moreover, all segments of rodent RVA MpR12 were highly divergent and assigned to novel genotypes, but RVA MpR12 was phylogenetically closer to bat RVAs than to other rodent RVAs, indicating a unique evolutionary history. We further investigated the virological properties of the isolated RVAs. In brief, we found that 16-06 entered cells by binding to sialic acids on the cell surface, while MpR12 entered in a sialic acid-independent manner. Experimental inoculation of suckling mice with 16-06 and MpR12 revealed that these RVAs are causative agents of diarrhea. Moreover, 16-06 and MpR12 demonstrated an ability to infect and replicate in a 3D-reconstructed primary human intestinal epithelium with comparable efficiency to the human RVA. Taken together, our results detail the unique genetic and virological features of bat and rodent RVAs and demonstrate the need for further investigation of their zoonotic potential. IMPORTANCE Recent advances in nucleotide sequence detection methods have enabled the detection of RVA genomes from various animals. These studies have discovered multiple divergent RVAs and have resulted in proposals for the genetic classification of novel genotypes. However, most of these RVAs have been identified via dsRNA viral genomes and not from infectious viruses, and their virological properties, such as cell/host tropisms, transmissibility, and pathogenicity, are unclear and remain to be clarified. Here, we successfully isolated RVAs with novel genome constellations from three bats and one rodent in Zambia. In addition to whole-genome sequencing, the isolated RVAs were characterized by glycan-binding affinity, pathogenicity in mice, and infectivity to the human gut using a 3D culture of primary intestinal epithelium. Our study reveals the first virological properties of bat and rodent RVAs with high genetic diversity and unique evolutional history and provides basic knowledge to begin estimating the potential of zoonotic transmission.
• S-217622, a SARS-CoV-2 main protease inhibitor, decreases viral load and ameliorates COVID-19 severity in hamsters.
  Michihito Sasaki; Koshiro Tabata; Mai Kishimoto; Yukari Itakura; Hiroko Kobayashi; Takuma Ariizumi; Kentaro Uemura; Shinsuke Toba; Shinji Kusakabe; Yuki Maruyama; Shun Iida; Noriko Nakajima; Tadaki Suzuki; Shinpei Yoshida; Haruaki Nobori; Takao Sanaki; Teruhisa Kato; Takao Shishido; William W Hall; Yasuko Orba; Akihiko Sato; Hirofumi Sawa
  Science translational medicine, 15, 679, eabq4064, 03 Nov. 2022, [International Magazine]
  English, Scientific journal, In parallel with vaccination, oral antiviral agents are highly anticipated to act as countermeasures for the treatment of the coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Oral antiviral medication demands not only high antiviral activity, but also target specificity, favorable oral bioavailability, and high metabolic stability. Although a large number of compounds have been identified as potential inhibitors of SARS-CoV-2 infection in vitro, few have proven to be effective in vivo. Here, we show that oral administration of S-217622 (ensitrelvir), an inhibitor of SARS-CoV-2 main protease (Mpro, also known as 3C-like protease), decreases viral load and ameliorates disease severity in SARS-CoV-2-infected hamsters. S-217622 inhibited viral proliferation at low nanomolar to sub-micromolar concentrations in cells. Oral administration of S-217622 demonstrated favorable pharmacokinetic properties and accelerated recovery from acute SARS-CoV-2 infection in hamster recipients. Moreover, S-217622 exerted antiviral activity against SARS-CoV-2 variants of concern (VOCs), including the highly pathogenic Delta variant and the recently emerged Omicron BA.5 and BA.2.75 variants. Overall, our study provides evidence that S-217622, an antiviral agent that is under evaluation in a phase 3 clinical trial (clinical trial registration no. jRCT2031210350), possesses remarkable antiviral potency and efficacy against SARS-CoV-2 and is a prospective oral therapeutic option for COVID-19.
• Discovery of Chlorofluoroacetamide-Based Covalent Inhibitors for Severe Acute Respiratory Syndrome Coronavirus 2 3CL Protease.
  Yuya Hirose; Naoya Shindo; Makiko Mori; Satsuki Onitsuka; Hikaru Isogai; Rui Hamada; Tadanari Hiramoto; Jinta Ochi; Daisuke Takahashi; Tadashi Ueda; Jose M M Caaveiro; Yuya Yoshida; Shigehiro Ohdo; Naoya Matsunaga; Shinsuke Toba; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa; Akihiko Sato; Eiji Kawanishi; Akio Ojida
  Journal of medicinal chemistry, 65, 20, 13852, 13865, 27 Oct. 2022, [International Magazine]
  English, Scientific journal, The coronavirus disease 2019 (COVID-19) pandemic has necessitated the development of antiviral agents against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). 3C-like protease (3CLpro) is a promising target for COVID-19 treatment. Here, we report a new class of covalent inhibitors of 3CLpro that possess chlorofluoroacetamide (CFA) as a cysteine-reactive warhead. Based on an aza-peptide scaffold, we synthesized a series of CFA derivatives in enantiopure form and evaluated their biochemical efficiency. The data revealed that 8a (YH-6) with the R configuration at the CFA unit strongly blocks SARS-CoV-2 replication in infected cells, and its potency is comparable to that of nirmatrelvir. X-ray structural analysis showed that YH-6 formed a covalent bond with Cys145 at the catalytic center of 3CLpro. The strong antiviral activity and favorable pharmacokinetic properties of YH-6 suggest its potential as a lead compound for the treatment of COVID-19.
• Dihydromaniwamycin E, a Heat-Shock Metabolite from Thermotolerant Streptomyces sp. JA74, Exhibiting Antiviral Activity against Influenza and SARS-CoV-2 Viruses.
  Shun Saito; Kayo Funayama; Wataru Kato; Mayu Okuda; Meiko Kawamoto; Teruhiko Matsubara; Toshinori Sato; Akihiko Sato; Satoko Otsuguro; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa; Katsumi Maenaka; Kazutoshi Shindo; Masaya Imoto; Midori A Arai
  Journal of natural products, 85, 11, 2583, 2591, 12 Oct. 2022, [International Magazine]
  English, Scientific journal, Dihydromaniwamycin E (1), a new maniwamycin derivative featuring an azoxy moiety, has been isolated from the culture extract of thermotolerant Streptomyces sp. JA74 along with the known analogue maniwamycin E (2). Compound 1 is produced only by cultivation of strain JA74 at 45 °C, and this type of compound has been previously designated a "heat shock metabolite (HSM)" by our research group. Compound 2 is detected as a production-enhanced metabolite at high temperature. Structures of 1 and 2 are elucidated by NMR and MS spectroscopic analyses. The absolute structure of 1 is determined after the total synthesis of four stereoisomers. Though the absolute structure of 2 has been proposed to be the same as the structure of maniwamycin D, the NMR and the optical rotation value of 2 are in agreement with those of maniwamycin E. Therefore, this study proposes a structural revision of maniwamycins D and E. Compounds 1 and 2 show inhibitory activity against the influenza (H1N1) virus infection of MDCK cells, demonstrating IC50 values of 25.7 and 63.2 μM, respectively. Notably, 1 and 2 display antiviral activity against SARS-CoV-2, the causative agent of COVID-19, when used to infect 293TA and VeroE6T cells, with 1 and 2 showing IC50 values (for infection of 293TA cells) of 19.7 and 9.7 μM, respectively. The two compounds do not exhibit cytotoxicity in these cell lines at those IC50 concentrations.
• Identification of cap-dependent endonuclease inhibitors with broad-spectrum activity against bunyaviruses.
  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 Kida
  Proceedings of the National Academy of Sciences of the United States of America, 119, 36, e2206104119, 06 Sep. 2022, [International Magazine]
  English, Scientific journal, 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., 33801613;32230793
• Antiviral effect of cetylpyridinium chloride in mouthwash on SARS-CoV-2
  Ryo Takeda; Hirofumi Sawa; Michihito Sasaki; Yasuko Orba; Nako Maishi; Takuya Tsumita; Natsumi Ushijima; Yasuhiro Hida; Hidehiko Sano; Yoshimasa Kitagawa; Kyoko Hida
  Scientific Reports, 12, 1, Springer Science and Business Media LLC, 18 Aug. 2022
  Scientific journal, Abstract
  
  Cetylpyridinium chloride (CPC), a quaternary ammonium compound, which is present in mouthwash, is effective against bacteria, fungi, and enveloped viruses. This study was conducted to explore the antiviral effect of CPC on SARS-CoV-2. There are few reports on the effect of CPC against wild-type SARS-CoV-2 at low concentrations such as 0.001%–0.005% (10–50 µg/mL). Interestingly, we found that low concentrations of CPC suppressed the infectivity of human isolated SARS-CoV-2 strains (Wuhan, Alpha, Beta, and Gamma) even in saliva. Furthermore, we demonstrated that CPC shows anti-SARS-CoV-2 effects without disrupting the virus envelope, using sucrose density analysis and electron microscopic examination. In conclusion, this study provided experimental evidence that CPC may inhibit SARS-CoV-2 infection even at lower concentrations.
• Isolation and characterization of an orthoreovirus from Indonesian fruit bats.
  Kittiya Intaruck; Yukari Itakura; Mai Kishimoto; Herman M Chambaro; Agus Setiyono; Ekowati Handharyani; Kentaro Uemura; Hayato Harima; Satoshi Taniguchi; Masayuki Saijo; Takashi Kimura; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki
  Virology, 575, 10, 19, 15 Aug. 2022, [International Magazine]
  English, Scientific journal, Nelson Bay orthoreovirus (NBV) is an emerging bat-borne virus and causes respiratory tract infections in humans sporadically. Over the last two decades, several strains genetically related to NBV were isolated from humans and various bat species, predominantly in Southeast Asia (SEA), suggesting a high prevalence of the NBV species in this region. In this study, an orthoreovirus (ORV) belonging to the NBV species was isolated from Indonesian fruit bats' feces, tentatively named Paguyaman orthoreovirus (PgORV). Serological studies revealed that 81.2% (108/133) of Indonesian fruit bats sera had neutralizing antibodies against PgORV. Whole-genome sequencing and phylogenetic analysis of PgORV suggested the occurrence of past reassortments with other NBV strains isolated in SEA, indicating the dispersal and circulation of NBV species among bats in this region. Intranasal PgORV inoculation of laboratory mice caused severe pneumonia. Our study characterized PgORV's unique genetic background and highlighted the potential risk of PgORV-related diseases in Indonesia.
• Serological characterization of lineage II insect-specific flaviviruses compared with pathogenic mosquito-borne flaviviruses.
  Koshiro Tabata; Yukari Itakura; Shinsuke Toba; Kentaro Uemura; Mai Kishimoto; Michihito Sasaki; Jessica J Harrison; Akihiko Sato; William W Hall; Roy A Hall; Hirofumi Sawa; Yasuko Orba
  Biochemical and biophysical research communications, 616, 115, 121, 06 Aug. 2022, [International Magazine]
  English, Scientific journal, The genus Flavivirus includes pathogenic tick- and mosquito-borne flaviviruses as well as non-pathogenic insect-specific flaviviruses (ISFVs). Phylogenetic analysis based on whole amino acid sequences has indicated that lineage II ISFVs have similarities to pathogenic flaviviruses. In this study, we used reactive analysis with immune serum against Psorophora flavivirus (PSFV) as a lineage IIa ISFV, and Barkeji virus (BJV) as a lineage IIb ISFV, to evaluate the antigenic similarity among lineage IIa and IIb ISFVs, and pathogenic mosquito-borne flaviviruses (MBFVs). Binding and antibody-dependent enhancement assays showed that anti-PSFV sera had broad cross-reactivity with MBFV antigens, while anti-BJV sera had low cross-reactivity. Both of the lineage II ISFV antisera were rarely observed to neutralize MBFVs. These results suggest that lineage IIa ISFV PSFV has more antigenic similarity to MBFVs than lineage IIb ISFV BJV., 32230793;31949323;33801644
• A high-affinity aptamer with base-appended base-modified DNA bound to isolated authentic SARS-CoV-2 strains wild-type and B.1.617.2 (delta variant).
  Hirotaka Minagawa; Hirofumi Sawa; Tomoko Fujita; Shintaro Kato; Asumi Inaguma; Miwako Hirose; Yasuko Orba; Michihito Sasaki; Koshiro Tabata; Naoki Nomura; Masashi Shingai; Yasuhiko Suzuki; Katsunori Horii
  Biochemical and biophysical research communications, 614, 207, 212, 23 Jul. 2022, [International Magazine]
  English, Scientific journal, Simple, highly sensitive detection technologies for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are crucial for the effective implementation of public health policies. We used the systematic evolution of ligands by exponential enrichment with a modified DNA library, including a base-appended base (uracil with a guanine base at its fifth position), to create an aptamer with a high affinity for the receptor-binding domain (RBD) of the SARS-CoV-2 spike glycoprotein. The aptamer had a dissociation constant of 1.2 and < 1 nM for the RBD and spike trimer, respectively. Furthermore, enzyme-linked aptamer assays confirmed that the aptamer binds to isolated authentic SARS-CoV-2 wild-type and B.1.617.2 (delta variant). The binding signal was larger that of commercially available anti-SARS-CoV-2 RBD antibody. Thus, this aptamer as a sensing element will enable the highly sensitive detection of SARS-CoV-2.
• Virological characteristics of the SARS-CoV-2 Omicron BA.2 spike.
  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 Sato
  Cell, 185, 12, 2103, 2115, 09 Jun. 2022, [International Magazine]
  English, Scientific journal, 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.
• An unusually long Rift valley fever inter-epizootic period in Zambia: Evidence for enzootic virus circulation and risk for disease outbreak.
  Herman M Chambaro; Kazuyo Hirose; Michihito Sasaki; Brigadier Libanda; Yona Sinkala; Paul Fandamu; Walter Muleya; Fredrick Banda; Joseph Chizimu; David Squarre; Misheck Shawa; Yongjin Qiu; Hayato Harima; Yuki Eshita; Edgar Simulundu; Hirofumi Sawa; Yasuko Orba
  PLoS neglected tropical diseases, 16, 6, e0010420, Jun. 2022, [International Magazine]
  English, Scientific journal, Rift valley fever (RVF) is a mosquito-borne disease of animals and humans. Although RVF outbreaks are usually reported at 5-15-year intervals in sub-Saharan Africa, Zambia has experienced an unusually long inter-epizootic/-epidemic period of more than three decades. However, serological evidence of RVF virus (RVFV) infection in domestic ruminants during this period underscores the need for comprehensive investigation of the mechanisms of virus perpetuation and disease emergence. Mosquitoes (n = 16,778) captured from eight of the ten provinces of Zambia between April 2014 and May 2019 were pooled (n = 961) and screened for RVFV genome by a pan-phlebo RT-PCR assay. Aedes mosquito pools (n = 85) were further screened by nested RT-PCR assay. Sera from sheep (n = 13), goats (n = 259) and wild ungulates (n = 285) were screened for RVFV antibodies by ELISA while genome detection in pooled sera (n = 276) from domestic (n = 248) and wild ungulates (n = 37) was performed by real-time RT-PCR assay. To examine the association between the long inter-epizootic period and climatic variables, we examined El Niño-Southern Oscillation indices, precipitation anomalies, and normalized difference vegetation index. We then derived RVF risk maps by exploring climatic variables that would favor emergence of primary RVFV vectors. While no RVFV genome could be detected in pooled mosquito and serum samples, seroprevalence was significantly high (OR = 8.13, 95% CI [4.63-14.25]) in wild ungulates (33.7%; 96/285) compared to domestic ruminants (5.6%; 16/272). Retrospective analysis of RVF epizootics in Zambia showed a positive correlation between anomalous precipitation (La Niña) and disease emergence. On risk mapping, whilst northern and eastern parts of the country were at high risk, domestic ruminant population density was low (< 21 animals/km2) in these areas compared to low risk areas (>21 animals/km2). Besides evidence of silent circulation of RVFV and the risk of disease emergence in some areas, wildlife may play a role in the maintenance of RVFV in Zambia., 32230793;31949323;24716624
• Glu333 in rabies virus glycoprotein is involved in virus attenuation through astrocyte infection and interferon responses.
  Yukari Itakura; Koshiro Tabata; Kohei Morimoto; Naoto Ito; Herman M Chambaro; Ryota Eguchi; Ken-Ichi Otsuguro; William W Hall; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki
  iScience, 25, 4, 104122, 104122, 15 Apr. 2022, [International Magazine]
  English, Scientific journal, The amino acid residue at position 333 of the rabies virus (RABV) glycoprotein (G333) is a major determinant of RABV pathogenicity. Virulent RABV strains possess Arg333, whereas the attenuated strain HEP-Flury (HEP) possesses Glu333. To investigate the potential attenuation mechanism dependent on a single amino acid at G333, comparative analysis was performed between HEP and HEP333R mutant with Arg333. We examined their respective tropism for astrocytes and the subsequent immune responses in astrocytes. Virus replication and subsequent interferon (IFN) responses in astrocytes infected with HEP were increased compared with HEP333R both in vitro and in vivo. Furthermore, involvement of IFN in the avirulency of HEP was demonstrated in IFN-receptor knockout mice. These results indicate that Glu333 contributes to RABV attenuation by determining the ability of the virus to infect astrocytes and stimulate subsequent IFN responses.
• Discovery of S-217622, a Noncovalent Oral SARS-CoV-2 3CL Protease Inhibitor Clinical Candidate for Treating COVID-19.
  Yuto Unoh; Shota Uehara; Kenji Nakahara; Haruaki Nobori; Yukiko Yamatsu; Shiho Yamamoto; Yuki Maruyama; Yoshiyuki Taoda; Koji Kasamatsu; Takahiro Suto; Kensuke Kouki; Atsufumi Nakahashi; Sho Kawashima; Takao Sanaki; Shinsuke Toba; Kentaro Uemura; Tohru Mizutare; Shigeru Ando; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa; Akihiko Sato; Takafumi Sato; Teruhisa Kato; Yuki Tachibana
  Journal of medicinal chemistry, 65, 9, 6499, 6512, 30 Mar. 2022, [International Magazine]
  English, Scientific journal, The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has resulted in millions of deaths and threatens public health and safety. Despite the rapid global spread of COVID-19 vaccines, effective oral antiviral drugs are urgently needed. Here, we describe the discovery of S-217622, the first oral noncovalent, nonpeptidic SARS-CoV-2 3CL protease inhibitor clinical candidate. S-217622 was discovered via virtual screening followed by biological screening of an in-house compound library, and optimization of the hit compound using a structure-based drug design strategy. S-217622 exhibited antiviral activity in vitro against current outbreaking SARS-CoV-2 variants and showed favorable pharmacokinetic profiles in vivo for once-daily oral dosing. Furthermore, S-217622 dose-dependently inhibited intrapulmonary replication of SARS-CoV-2 in mice, indicating that this novel noncovalent inhibitor could be a potential oral agent for treating COVID-19.
• Application of Acoustic Ejection MS System to High-Throughput Screening for SARS-CoV-2 3CL Protease Inhibitors.
  Tsukasa Hasegawa; Riyo M Imamura; Tateki Suzuki; Takao Hashiguchi; Takao Nomura; Satoko Otsuguro; Katsumi Maenaka; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa; Akihiko Sato; Takayoshi Okabe; Tetsuo Nagano; Hirotatsu Kojima
  Chemical & pharmaceutical bulletin, 70, 3, 199, 201, 01 Mar. 2022, [Domestic magazines]
  English, Scientific journal, MS is a powerful methodology for chemical screening to directly quantify substrates and products of enzymes, but its low throughput has been an issue. Recently, an acoustic liquid-handling apparatus (Echo®) used for rapid nano-dispensing has been coupled to a high-sensitivity mass spectrometer to create the Echo® MS system, and we applied this system to screening of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) 3CL protease inhibitors. Primary screening of 32033 chemical samples was completed in 12 h. Among the hits showing selective, dose-dependent 3CL-inhibitory activity, 8 compounds showed antiviral activity in cell-based assay.
• Synthesis and Anti-dengue Virus Activity of 5-Ethynylimidazole-4-carboxamide (EICA) Nucleotide Prodrugs.
  Motoki Nakamura; Kentaro Uemura; Noriko Saito-Tarashima; Akihiko Sato; Yasuko Orba; Hirofumi Sawa; Akira Matsuda; Katsumi Maenaka; Noriaki Minakawa
  Chemical & pharmaceutical bulletin, 70, 3, 220, 225, 01 Mar. 2022, [Domestic magazines]
  English, Scientific journal, We previously showed that 5-ethynyl-(1-β-D-ribofuranosyl)imidazole-4-carboxamide (1; EICAR) is a potent anti-dengue virus (DENV) compound but is cytotoxic to some cell lines, while its 4-thio derivative, 5-ethynyl-(4-thio-1-β-D-ribofuranosyl)imidazole-4-carboxamide (2; 4'-thioEICAR), has less cytotoxicity but also less anti-DENV activity. Based on the hypothesis that the lower anti-DENV activity of 2 is due to reduced susceptibility to phosphorylation by cellular kinase(s), we investigated whether a monophosphate prodrug of 2 can improve its activity. Here, we first prepared two types of prodrug of 1, which revealed that the S-acyl-2-thioethyl (SATE) prodrug had stronger anti-DENV activity than the aryloxyphosphoramidate (so-called ProTide) prodrug. Based on these findings, we next prepared the SATE prodrug of 4'-thioEICAR 18. As expected, the resulting 18 showed potent anti-DENV activity, which was comparable to that of 1; however, its cytotoxicity was also increased relative to 2. Our findings suggest that prodrugs of 4'-thioribonucleoside derivatives such as EICAR (1) represent an effective approach to developing potent biologically active compounds; however, the balance between antiviral activity and cytotoxicity remains to be addressed.
• Attenuated fusogenicity and pathogenicity of SARS-CoV-2 Omicron variant.
  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 Sato
  Nature, 603, 7902, 700, 705, 01 Feb. 2022, [International Magazine]
  English, Scientific journal, 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.
• Current knowledge of vector-borne zoonotic pathogens in Zambia: A clarion call to scaling-up "One Health" research in the wake of emerging and re-emerging infectious diseases.
  Benjamin Mubemba; Monicah M Mburu; Katendi Changula; Walter Muleya; Lavel C Moonga; Herman M Chambaro; Masahiro Kajihara; Yongjin Qiu; Yasuko Orba; Kyoko Hayashida; Catherine G Sutcliffe; Douglas E Norris; Philip E Thuma; Phillimon Ndubani; Simbarashe Chitanga; Hirofumi Sawa; Ayato Takada; Edgar Simulundu
  PLoS neglected tropical diseases, 16, 2, e0010193, Feb. 2022, [International Magazine]
  English, Scientific journal, BACKGROUND: Although vector-borne zoonotic diseases are a major public health threat globally, they are usually neglected, especially among resource-constrained countries, including those in sub-Saharan Africa. This scoping review examined the current knowledge and identified research gaps of vector-borne zoonotic pathogens in Zambia. METHODS AND FINDINGS: Major scientific databases (Web of Science, PubMed, Scopus, Google Scholar, CABI, Scientific Information Database (SID)) were searched for articles describing vector-borne (mosquitoes, ticks, fleas and tsetse flies) zoonotic pathogens in Zambia. Several mosquito-borne arboviruses have been reported including Yellow fever, Ntaya, Mayaro, Dengue, Zika, West Nile, Chikungunya, Sindbis, and Rift Valley fever viruses. Flea-borne zoonotic pathogens reported include Yersinia pestis and Rickettsia felis. Trypanosoma sp. was the only tsetse fly-borne pathogen identified. Further, tick-borne zoonotic pathogens reported included Crimean-Congo Haemorrhagic fever virus, Rickettsia sp., Anaplasma sp., Ehrlichia sp., Borrelia sp., and Coxiella burnetii. CONCLUSIONS: This study revealed the presence of many vector-borne zoonotic pathogens circulating in vectors and animals in Zambia. Though reports of human clinical cases were limited, several serological studies provided considerable evidence of zoonotic transmission of vector-borne pathogens in humans. However, the disease burden in humans attributable to vector-borne zoonotic infections could not be ascertained from the available reports and this precludes the formulation of national policies that could help in the control and mitigation of the impact of these diseases in Zambia. Therefore, there is an urgent need to scale-up "One Health" research in emerging and re-emerging infectious diseases to enable the country to prepare for future epidemics, including pandemics.
• Hepatitis E virus infection in pigs: a first report from Zambia.
  Herman M Chambaro; Michihito Sasaki; Walter Muleya; Masahiro Kajihara; Misheck Shawa; Kabemba E Mwape; Hayato Harima; Yongjin Qiu; William W Hall; Paul Fandamu; David Squarre; Edgar Simulundu; Hirofumi Sawa; Yasuko Orba
  Emerging microbes & infections, 10, 1, 2169, 2172, Dec. 2021, [International Magazine]
  English, While evidence suggests presence of HEV infection in humans in Zambia, currently, there is no information on its occurrence in domestic pigs. Here, we investigated the presence of HEV antibodies and genome in domestic pigs in Zambia. Sera (n = 484) from domestic pigs were screened for antibodies against HEV by ELISA while genome detection in fecal (n = 25) and liver (n = 100) samples from slaughter pigs was conducted using nested RT-PCR assay. Overall, seroprevalence was 47.7% (231/484) while zoonotic genotype 3 HEV RNA was detected in 16.0% (20/125) of slaughtered pigs. This is the first report to highlight occurrence of HEV infection in domestic pigs in Zambia. This finding suggests possible contamination of the pork supply chain. Moreover, there is a potential risk of zoonotic transmission of HEV to abattoir workers, pig farmers and handlers.
• SARS-CoV-2 inhibits induction of the MHC class I pathway by targeting the STAT1-IRF1-NLRC5 axis.
  Ji-Seung Yoo; Michihito Sasaki; Steven X Cho; Yusuke Kasuga; Baohui Zhu; Ryota Ouda; Yasuko Orba; Paul de Figueiredo; Hirofumi Sawa; Koichi S Kobayashi
  Nature communications, 12, 1, 6602, 6602, 15 Nov. 2021, [International Magazine]
  English, Scientific journal, The MHC class I-mediated antigen presentation pathway plays a critical role in antiviral immunity. Here we show that the MHC class I pathway is targeted by SARS-CoV-2. Analysis of the gene expression profile from COVID-19 patients as well as SARS-CoV-2 infected epithelial cell lines reveals that the induction of the MHC class I pathway is inhibited by SARS-CoV-2 infection. We show that NLRC5, an MHC class I transactivator, is suppressed both transcriptionally and functionally by the SARS-CoV-2 ORF6 protein, providing a mechanistic link. SARS-CoV-2 ORF6 hampers type II interferon-mediated STAT1 signaling, resulting in diminished upregulation of NLRC5 and IRF1 gene expression. Moreover, SARS-CoV-2 ORF6 inhibits NLRC5 function via blocking karyopherin complex-dependent nuclear import of NLRC5. Collectively, our study uncovers an immune evasion mechanism of SARS-CoV-2 that targets the function of key MHC class I transcriptional regulators, STAT1-IRF1-NLRC5.
• Abnormal Blood Coagulation and Kidney Damage in Aged Hamsters Infected with Severe Acute Respiratory Syndrome Coronavirus 2.
  Marumi Ohno; Michihito Sasaki; Yasuko Orba; Toshiki Sekiya; Md Abdul Masum; Osamu Ichii; Tatsuya Sawamura; Akemi Kakino; Yasuhiko Suzuki; Hiroshi Kida; Hirofumi Sawa; Masashi Shingai
  Viruses, 13, 11, 22 Oct. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Systemic symptoms have often been observed in patients with coronavirus disease 2019 (COVID-19) in addition to pneumonia, however, the details are still unclear due to the lack of an appropriate animal model. In this study, we investigated and compared blood coagulation abnormalities and tissue damage between male Syrian hamsters of 9 (young) and over 36 (aged) weeks old after intranasal infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Despite similar levels of viral replication and inflammatory responses in the lungs of both age groups, aged but not young hamsters showed significant prolongation of prothrombin time and prominent acute kidney damage. Moreover, aged hamsters demonstrated increased intravascular coagulation time-dependently in the lungs, suggesting that consumption of coagulation factors causes prothrombin time prolongation. Furthermore, proximal urinary tract damage and mesangial matrix expansion were observed in the kidneys of the aged hamsters at early and later disease stages, respectively. Given that the severity and mortality of COVID-19 are higher in elderly human patients, the effect of aging on pathogenesis needs to be understood and should be considered for the selection of animal models. We, thus, propose that the aged hamster is a good small animal model for COVID-19 research.
• LIGHTHOUSE illuminates therapeutics for a variety of diseases including COVID-19
  Hideyuki Shimizu; Manabu Kodama; Masaki Matsumoto; Yasuko Orba; Michihito Sasaki; Akihiko Sato; Hirofumi Sawa; Keiichi I. Nakayama
  Cold Spring Harbor Laboratory, 25 Sep. 2021
  Although numerous promising therapeutic targets for human diseases have been discovered, most have not been successfully translated into clinical practice¹. A bottleneck in the application of basic research findings to patients is the enormous cost, time, and effort required for high-throughput screening of potential drugs² for given therapeutic targets. Recent advances in 3D docking simulations have not solved this problem, given that 3D protein structures with sufficient resolution are not always available and that they are computationally expensive to obtain. Here we have developed LIGHTHOUSE, a graph-based deep learning approach for discovery of the hidden principles underlying the association of small-molecule compounds with target proteins, and we present its validation by identifying potential therapeutic compounds for various human diseases. Without any 3D structural information for proteins or chemicals, LIGHTHOUSE estimates protein-compound scores that incorporate known evolutionary relations and available experimental data. It identified novel therapeutics for cancer, lifestyle-related disease, and bacterial infection. Moreover, LIGHTHOUSE predicted ethoxzolamide as a therapeutic for coronavirus disease 2019 (COVID-19), and this agent was indeed effective against alpha, beta, gamma, and delta variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that are rampant worldwide. Given that ethoxzolamide is already approved for several diseases, it could be rapidly deployed for the treatment of patients with COVID-19. We envision that LIGHTHOUSE will bring about a paradigm shift in translational medicine, providing a bridge from bench side to bedside.
• A targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses.
  Patrick Reteng; Linh Nguyen Thuy; Tam Tran Thi Minh; Maria Angélica Monteiro de Mello Mares-Guia; Maria Celeste Torres; Ana Maria Bispo de Filippis; Yasuko Orba; Shintaro Kobayashi; Kyoko Hayashida; Hirofumi Sawa; William W Hall; Lan Anh Nguyen Thi; Junya Yamagishi
  Scientific reports, 11, 1, 19031, 19031, 24 Sep. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Nucleic acid test (NAT), most typically quantitative PCR, is one of the standard methods for species specific flavivirus diagnosis. Semi-comprehensive NATs such as pan-flavivirus PCR which covers genus Flavivirus are also available; however, further specification by sequencing is required for species level differentiation. In this study, a semi-comprehensive detection system that allows species differentiation of flaviviruses was developed by integration of the pan-flavivirus PCR and Nanopore sequencing. In addition, a multiplexing method was established by adding index sequences through the PCR with a streamlined bioinformatics pipeline. This enables defining cut-off values for observed read counts. In the laboratory setting, this approach allowed the detection of up to nine different flaviviruses. Using clinical samples collected in Vietnam and Brazil, seven different flaviviruses were also detected. When compared to a commercial NAT, the sensitivity and specificity of our system were 66.7% and 95.4%, respectively. Conversely, when compared to our system, the sensitivity and specificity of the commercial NAT were 57.1% and 96.9%, respectively. In addition, Nanopore sequencing detected more positive samples (n = 8) compared to the commercial NAT (n = 6). Collectively, our study has established a semi-comprehensive sequencing-based diagnostic system for the detection of flaviviruses at extremely affordable costs, considerable sensitivity, and only requires simple experimental methods.
• A novel nairovirus associated with acute febrile illness in Hokkaido, Japan.
  Fumihiro Kodama; Hiroki Yamaguchi; Eunsil Park; Kango Tatemoto; Mariko Sashika; Ryo Nakao; Yurino Terauchi; Keita Mizuma; Yasuko Orba; Hiroaki Kariwa; Katsuro Hagiwara; Katsunori Okazaki; Akiko Goto; Rika Komagome; Masahiro Miyoshi; Takuya Ito; Kimiaki Yamano; Kentaro Yoshii; Chiaki Funaki; Mariko Ishizuka; Asako Shigeno; Yukari Itakura; Lesley Bell-Sakyi; Shunji Edagawa; Atsushi Nagasaka; Yoshihiro Sakoda; Hirofumi Sawa; Ken Maeda; Masayuki Saijo; Keita Matsuno
  Nature communications, 12, 1, 5539, 5539, 20 Sep. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, The increasing burden of tick-borne orthonairovirus infections, such as Crimean-Congo hemorrhagic fever, is becoming a global concern for public health. In the present study, we identify a novel orthonairovirus, designated Yezo virus (YEZV), from two patients showing acute febrile illness with thrombocytopenia and leukopenia after tick bite in Hokkaido, Japan, in 2019 and 2020, respectively. YEZV is phylogenetically grouped with Sulina virus detected in Ixodes ricinus ticks in Romania. YEZV infection has been confirmed in seven patients from 2014-2020, four of whom were co-infected with Borrelia spp. Antibodies to YEZV are found in wild deer and raccoons, and YEZV RNAs have been detected in ticks from Hokkaido. In this work, we demonstrate that YEZV is highly likely to be the causative pathogen of febrile illness, representing the first report of an endemic infection associated with an orthonairovirus potentially transmitted by ticks in Japan., 32230793;24716623;24716624
• 5-Hydroxymethyltubercidin Exhibits Potent Antiviral Activity against Flaviviruses and Coronaviruses, including SARS-CoV-2.
  Kentaro Uemura; Haruaki Nobori; Akihiko Sato; Takao Sanaki; Shinsuke Toba; Michihito Sasaki; Akiho Murai; Noriko Saito-Tarashima; Noriaki Minakawa; Yasuko Orba; Hiroaki Kariwa; William W Hall; Hirofumi Sawa; Akira Matsuda; Katsumi Maenaka
  iScience, 24, 10, 103120, 103120, 11 Sep. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Newly emerging or re-emerging viral infections continue to cause significant morbidity and mortality every year worldwide, resulting in serious effects on both health and the global economy. Despite significant drug discovery research against dengue viruses (DENV) and severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), no fully effective and specific drugs directed against these viruses have been discovered. Here, we examined the anti-DENV activity of tubercidin derivatives from a compound library from Hokkaido University and demonstrated that 5-hydroxymethyltubercidin (HMTU, HUP1108) possessed both potent anti-flavivirus and anti-coronavirus activities at submicromolar levels without significant cytotoxicity. Furthermore, HMTU inhibited viral RNA replication and specifically inhibited replication at the late stages of the SARS-CoV-2 infection process. Finally, we demonstrated that HMTU 5'-triphosphate inhibited RNA extension catalyzed by the viral RNA-dependent RNA-polymerase. Our findings suggest that HMTU has the potential of serving as a lead compound for the development of a broad-spectrum of antiviral agents, including SARS-CoV-2.
• Air-liquid interphase culture confers SARS-CoV-2 susceptibility to A549 alveolar epithelial cells.
  Michihito Sasaki; Mai Kishimoto; Yukari Itakura; Koshiro Tabata; Kittiya Intaruck; Kentaro Uemura; Shinsuke Toba; Takao Sanaki; Akihiko Sato; William W Hall; Yasuko Orba; Hirofumi Sawa
  Biochemical and biophysical research communications, 577, 146, 151, 10 Sep. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, The human lung cell A549 is susceptible to infection with a number of respiratory viruses. However, A549 cells are resistant to Severe Acute Respiratory Syndrome-Coronavirus-2 (SARS-CoV-2) infection in conventional submerged culture, and this would appear to be due to low expression levels of the SARS-CoV-2 entry receptor: angiotensin-converting enzyme-2 (ACE2). Here, we examined SARS-CoV-2 susceptibility to A549 cells after adaptation to air-liquid interface (ALI) culture. A549 cells in ALI culture yielded a layer of mucus on their apical surface, exhibited decreased expression levels of the proliferation marker KI-67 and intriguingly became susceptible to SARS-CoV-2 infection. We found that A549 cells increased the endogenous expression levels of ACE2 and TMPRSS2 following adaptation to ALI culture conditions. Camostat, a TMPRSS2 inhibitor, reduced SARS-CoV-2 infection in ALI-cultured A549 cells. These findings indicate that ALI culture switches the phenotype of A549 cells from resistance to susceptibility to SARS-CoV-2 infection through upregulation of ACE2 and TMPRSS2., 32230793
• Molecular Survey of Babesia and Anaplasma Infection in Cattle in Bolivia.
  Shohei Ogata; Juan Antonio Cristian Pereira; Loza Vega Ariel Jhonny; Herbas Perez Gladys Carolina; Keita Matsuno; Yasuko Orba; Hirofumi Sawa; Fumihiko Kawamori; Nariaki Nonaka; Ryo Nakao
  Veterinary sciences, 8, 9, 07 Sep. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Latin American countries produce more than a quarter of the world's beef and are a major global supplier of livestock protein. Tick-borne diseases (TBDs) are a major constraint to the livestock industry worldwide, including in Latin America. The aim of this study was to detect and characterise tick-borne pathogens in cattle from Santa Cruz, Bolivia, where no detailed epidemiological data are available. Blood samples were collected from 104 cattle. Apicomplexan parasites were detected by nested PCR amplification of the 18S ribosomal RNA gene (rDNA), and Anaplasmataceae was screened by the PCR amplification of 16S rDNA, followed by characterisation based on the heat shock protein and citrate synthase gene sequences. Babesia infection was observed in nine cattle (one Babesia bovis and eight Babesia bigemina), while Anaplasmataceae infection was detected in thirty-two cattle. A sequencing analysis confirmed the presence of Anaplasma marginale and Anaplasma platys-like. These results provide the first molecular evidence for the four above-mentioned tick-borne pathogens in cattle in Bolivia. This information improves our understanding of the epidemiology of TBDs and will help in formulating appropriate and improved pathogen control strategies.
• Attenuated infection by a Pteropine orthoreovirus isolated from an Egyptian fruit bat in Zambia.
  Hayato Harima; Michihito Sasaki; Yasuko Orba; Kosuke Okuya; Yongjin Qiu; Christida E Wastika; Katendi Changula; Masahiro Kajihara; Edgar Simulundu; Tomoyuki Yamaguchi; Yoshiki Eto; Akina Mori-Kajihara; Akihiko Sato; Satoshi Taniguchi; Ayato Takada; Masayuki Saijo; Bernard M Hang'ombe; Hirofumi Sawa
  PLoS neglected tropical diseases, 15, 9, e0009768, 07 Sep. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, BACKGROUND: Pteropine orthoreovirus (PRV) is an emerging bat-borne zoonotic virus that causes severe respiratory illness in humans. Although PRVs have been identified in fruit bats and humans in Australia and Asia, little is known about the prevalence of PRV infection in Africa. Therefore, this study performed an PRV surveillance in fruit bats in Zambia. METHODS: Egyptian fruit bats (Rousettus aegyptiacus, n = 47) and straw-colored fruit bats (Eidolon helvum, n = 33) captured in Zambia in 2017-2018 were screened for PRV infection using RT-PCR and serum neutralization tests. The complete genome sequence of an isolated PRV strain was determined by next generation sequencing and subjected to BLAST and phylogenetic analyses. Replication capacity and pathogenicity of the strain were investigated using Vero E6 cell cultures and BALB/c mice, respectively. RESULTS: An PRV strain, tentatively named Nachunsulwe-57, was isolated from one Egyptian fruit bat. Serological assays demonstrated that 98% of sera (69/70) collected from Egyptian fruit bats (n = 37) and straw-colored fruit bats (n = 33) had neutralizing antibodies against PRV. Genetic analyses revealed that all 10 genome segments of Nachunsulwe-57 were closely related to a bat-derived Kasama strain found in Uganda. Nachunsulwe-57 showed less efficiency in viral growth and lower pathogenicity in mice than another PRV strain, Miyazaki-Bali/2007, isolated from a patient. CONCLUSIONS: A high proportion of Egyptian fruit bats and straw-colored fruit bats were found to be seropositive to PRV in Zambia. Importantly, a new PRV strain (Nachunsulwe-57) was isolated from an Egyptian fruit bat in Zambia, which had relatively weak pathogenicity in mice. Taken together, our findings provide new epidemiological insights about PRV infection in bats and indicate the first isolation of an PRV strain that may have low pathogenicity to humans.
• SARS-CoV-2 Bearing a Mutation at the S1/S2 Cleavage Site Exhibits Attenuated Virulence and Confers Protective Immunity.
  Michihito Sasaki; Shinsuke Toba; Yukari Itakura; Herman M Chambaro; Mai Kishimoto; Koshiro Tabata; Kittiya Intaruck; Kentaro Uemura; Takao Sanaki; Akihiko Sato; William W Hall; Yasuko Orba; Hirofumi Sawa
  mBio, 12, 4, e0141521, 31 Aug. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) possesses a discriminative polybasic cleavage motif in its spike protein that is recognized by the host furin protease. Proteolytic cleavage activates the spike protein, thereby affecting both the cellular entry pathway and cell tropism of SARS-CoV-2. Here, we investigated the impact of the furin cleavage site on viral growth and pathogenesis using a hamster animal model infected with SARS-CoV-2 variants bearing mutations at the furin cleavage site (S gene mutants). In the airway tissues of hamsters, the S gene mutants exhibited low growth properties. In contrast to parental pathogenic SARS-CoV-2, hamsters infected with the S gene mutants showed no body weight loss and only a mild inflammatory response, thereby indicating the attenuated variant nature of S gene mutants. This transient infection was sufficient for inducing protective neutralizing antibodies that cross-react with different SARS-CoV-2 lineages. Consequently, hamsters inoculated with S gene mutants showed resistance to subsequent infection with both the parental strain and the currently emerging SARS-CoV-2 variants belonging to lineages B.1.1.7 and P.1. Taken together, our findings revealed that the loss of the furin cleavage site causes attenuation in the airway tissues of hamsters and highlighted the potential benefits of S gene mutants as potential immunogens. IMPORTANCE SARS-CoV-2 uses its spike protein to enter target cells. The spike protein is cleaved by a host protease, and this event facilitates viral entry and broadens cell tropism. In this study, we employed SARS-CoV-2 mutants lacking the S protein cleavage site and characterized their growth and pathogenicity using hamsters, a laboratory animal model for SARS-CoV-2 infection. These mutants exerted low pathogenicity but induced sufficient levels of neutralizing antibodies in hamsters, which protected hamsters from rechallenge with pathogenic clinical SARS-CoV-2 strains. These virus mutants may be used as protective immunogens against SARS-CoV-2 infection., 32230793
• Dual Effect of Organogermanium Compound THGP on RIG-I-Mediated Viral Sensing and Viral Replication during Influenza a Virus Infection.
  Sunanda Baidya; Yoko Nishimoto; Seiichi Sato; Yasuhiro Shimada; Nozomi Sakurai; Hirotaka Nonaka; Koki Noguchi; Mizuki Kido; Satoshi Tadano; Kozo Ishikawa; Kai Li; Aoi Okubo; Taisho Yamada; Yasuko Orba; Michihito Sasaki; Hirofumi Sawa; Hiroko Miyamoto; Ayato Takada; Takashi Nakamura; Akinori Takaoka
  Viruses, 13, 9, 24 Aug. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, The interaction of viral nucleic acid with protein factors is a crucial process for initiating viral polymerase-mediated viral genome replication while activating pattern recognition receptor (PRR)-mediated innate immune responses. It has previously been reported that a hydrolysate of Ge-132, 3-(trihydroxygermyl) propanoic acid (THGP), shows a modulatory effect on microbial infections, inflammation, and immune responses. However, the detailed mechanism by which THGP can modify these processes during viral infections remained unknown. Here, we show that THGP can specifically downregulate type I interferon (IFN) production in response to stimulation with a cytosolic RNA sensor RIG-I ligand 5'-triphosphate RNA (3pRNA) but not double-stranded RNA, DNA, or lipopolysaccharide. Consistently, treatment with THGP resulted in the dose-dependent suppression of type I IFN induction upon infections with influenza virus (IAV) and vesicular stomatitis virus, which are known to be mainly sensed by RIG-I. Mechanistically, THGP directly binds to the 5'-triphosphate moiety of viral RNA and competes with RIG-I-mediated recognition. Furthermore, we found that THGP can directly counteract the replication of IAV but not EMCV (encephalitismyocarditis virus), by inhibiting the interaction of viral polymerase with RNA genome. Finally, IAV RNA levels were significantly reduced in the lung tissues of THGP-treated mice when compared with untreated mice. These results suggest a possible therapeutic implication of THGP and show direct antiviral action, together with the suppressive activity of innate inflammation.
• A SARS-CoV-2 antibody broadly neutralizes SARS-related coronaviruses and variants by coordinated recognition of a virus-vulnerable site.
  Taishi Onodera; Shunsuke Kita; Yu Adachi; Saya Moriyama; Akihiko Sato; Takao Nomura; Shuhei Sakakibara; Takeshi Inoue; Takashi Tadokoro; Yuki Anraku; Kohei Yumoto; Cong Tian; Hideo Fukuhara; Michihito Sasaki; Yasuko Orba; Nozomi Shiwa; Naoko Iwata; Noriyo Nagata; Tateki Suzuki; Jiei Sasaki; Tsuyoshi Sekizuka; Keisuke Tonouchi; Lin Sun; Shuetsu Fukushi; Hiroyuki Satofuka; Yasuhiro Kazuki; Mitsuo Oshimura; Tomohiro Kurosaki; Makoto Kuroda; Yoshiharu Matsuura; Tadaki Suzuki; Hirofumi Sawa; Takao Hashiguchi; Katsumi Maenaka; Yoshimasa Takahashi
  Immunity, 54, 10, 2385, 2398, 24 Aug. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Potent neutralizing SARS-CoV-2 antibodies often target the spike protein receptor-binding site (RBS), but the variability of RBS epitopes hampers broad neutralization of multiple sarbecoviruses and drifted viruses. Here, using humanized mice, we identified an RBS antibody with a germline VH gene that potently neutralized SARS-related coronaviruses, including SARS-CoV and SARS-CoV-2 variants. X-ray crystallography revealed coordinated recognition by the heavy chain of non-RBS conserved sites and the light chain of RBS with a binding angle mimicking the angiotensin-converting enzyme 2 (ACE2) receptor. The minimum footprints in the hypervariable region of RBS contributed to the breadth of neutralization, which was enhanced by immunoglobulin G3 (IgG3) class switching. The coordinated binding resulted in broad neutralization of SARS-CoV and emerging SARS-CoV-2 variants of concern. Low-dose therapeutic antibody treatment in hamsters reduced the virus titers and morbidity during SARS-CoV-2 challenge. The structural basis for broad neutralizing activity may inform the design of a broad spectrum of therapeutics and vaccines.
• Mosquito-Borne Viral Pathogens Detected in Zambia: A Systematic Review.
  Rachel Milomba Velu; Geoffrey Kwenda; Liyali Libonda; Caroline Cleopatra Chisenga; Bumbangi Nsoni Flavien; Obvious Nchimunya Chilyabanyama; Michelo Simunyandi; Samuel Bosomprah; Nicholus Chintu Sande; Katendi Changula; Walter Muleya; Monicah Mirai Mburu; Benjamin Mubemba; Simbarashe Chitanga; John Tembo; Matthew Bates; Nathan Kapata; Yasuko Orba; Masahiro Kajihara; Ayato Takada; Hirofumi Sawa; Roma Chilengi; Edgar Simulundu
  Pathogens (Basel, Switzerland), 10, 8, 10 Aug. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Emerging and re-emerging mosquito-borne viral diseases are a threat to global health. This systematic review aimed to investigate the available evidence of mosquito-borne viral pathogens reported in Zambia. A search of literature was conducted in PubMed and Google Scholar for articles published from 1 January 1930 to 30 June 2020 using a combination of keywords. Eight mosquito-borne viruses belonging to three families, Togaviridae, Flaviviridae and Phenuiviridae were reported. Three viruses (Chikungunya virus, Mayaro virus, Mwinilunga virus) were reported among the togaviruses whilst four (dengue virus, West Nile virus, yellow fever virus, Zika virus) were among the flavivirus and only one virus, Rift Valley fever virus, was reported in the Phenuiviridae family. The majority of these mosquito-borne viruses were reported in Western and North-Western provinces. Aedes and Culex species were the main mosquito-borne viral vectors reported. Farming, fishing, movement of people and rain patterns were among factors associated with mosquito-borne viral infection in Zambia. Better diagnostic methods, such as the use of molecular tools, to detect the viruses in potential vectors, humans, and animals, including the recognition of arboviral risk zones and how the viruses circulate, are important for improved surveillance and design of effective prevention and control measures.
• RIG-I triggers a signaling-abortive anti-SARS-CoV-2 defense in human lung cells.
  Taisho Yamada; Seiichi Sato; Yuki Sotoyama; Yasuko Orba; Hirofumi Sawa; Hajime Yamauchi; Michihito Sasaki; Akinori Takaoka
  Nature immunology, 22, 7, 820, 828, Jul. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Efficient immune responses against viral infection are determined by sufficient activation of nucleic acid sensor-mediated innate immunity1,2. Coronavirus disease 2019, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), remains an ongoing global pandemic. It is an urgent challenge to clarify the innate recognition mechanism to control this virus. Here we show that retinoic acid-inducible gene-I (RIG-I) sufficiently restrains SARS-CoV-2 replication in human lung cells in a type I/III interferon (IFN)-independent manner. RIG-I recognizes the 3' untranslated region of the SARS-CoV-2 RNA genome via the helicase domains, but not the C-terminal domain. This new mode of RIG-I recognition does not stimulate its ATPase, thereby aborting the activation of the conventional mitochondrial antiviral-signaling protein-dependent pathways, which is in accordance with lack of cytokine induction. Nevertheless, the interaction of RIG-I with the viral genome directly abrogates viral RNA-dependent RNA polymerase mediation of the first step of replication. Consistently, genetic ablation of RIG-I allows lung cells to produce viral particles that expressed the viral spike protein. By contrast, the anti-SARS-CoV-2 activity was restored by all-trans retinoic acid treatment through upregulation of RIG-I protein expression in primary lung cells derived from patients with chronic obstructive pulmonary disease. Thus, our findings demonstrate the distinctive role of RIG-I as a restraining factor in the early phase of SARS-CoV-2 infection in human lung cells.
• Serologic and molecular evidence for circulation of Crimean-Congo hemorrhagic fever virus in ticks and cattle in Zambia.
  Masahiro Kajihara; Martin Simuunza; Ngonda Saasa; George Dautu; Akina Mori-Kajihara; Yongjin Qiu; Ryo Nakao; Yoshiki Eto; Hayato Furumoto; Bernard M Hang'ombe; Yasuko Orba; Hirofumi Sawa; Edgar Simulundu; Shuetsu Fukushi; Shigeru Morikawa; Masayuki Saijo; Jiro Arikawa; Swithine Kabilika; Mwaka Monze; Victor Mukonka; Aaron Mweene; Ayato Takada; Kumiko Yoshimatsu
  PLoS neglected tropical diseases, 15, 6, e0009452, Jun. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, 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.
• Combinatorial approach with mass spectrometry and lectin microarray dissected glycoproteomic features of virion-derived spike protein of SARS-CoV-2
  Takahiro Hiono; Azusa Tomioka; Hiroyuki Kaji; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa; Atsushi Kuno
  Journal of proteome research, 23, 4, 1408, 1419, Cold Spring Harbor Laboratory, 12 Apr. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, The COVID-19 pandemic caused by the novel coronavirus, SARS-CoV-2, has a global impact on public health. Since glycosylation of the viral envelope glycoproteins is known to be deeply associated with their immunogenicity, intensive studies on the glycans of its major glycoprotein, S protein, have been conducted. Nevertheless, the detailed site-specific glycan compositions of virion-associated S protein have not yet been clarified. Here, we conducted intensive glycoproteomic analyses of SARS-CoV-2 S protein using a combinatorial approach with two different technologies: mass spectrometry (MS) and lectin microarray. Using our unique MS1-based glycoproteomic technique, Glyco-RIDGE, in addition to MS2-based Byonic search, we identified 1,759 site-specific glycan compositions. The most frequent was HexNAc:Hex:Fuc:NeuAc:NeuGc = 6:6:1:0:0, suggesting a tri-antennary N-glycan terminating with LacNAc and having bisecting GlcNAc and a core fucose, which was found in 20 of 22 glycosylated sites. The subsequent lectin microarray analysis emphasized intensive outer arm fucosylation of glycans, which efficiently complemented the glycoproteomic features. The present results illustrate the high-resolution glycoproteomic features of SARS-CoV-2 S protein and significantly contribute to vaccine design, as well as the understanding of viral protein synthesis.
• Host serine proteases TMPRSS2 and TMPRSS11D mediate proteolytic activation and trypsin-independent infection in group A rotaviruses.
  Michihito Sasaki; Yukari Itakura; Mai Kishimoto; Koshiro Tabata; Kentaro Uemura; Naoto Ito; Makoto Sugiyama; Christida E Wastika; Yasuko Orba; Hirofumi Sawa
  Journal of virology, 95, 11, 24 Mar. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Group A rotaviruses (RVAs) are representative enteric virus species and major causes of diarrhea in humans and animals. The RVA virion is a triple-layered particle, and the outermost layer consists of the glycoprotein VP7 and spike protein VP4. To increase the infectivity of RVA, VP4 is proteolytically cleaved into VP5* and VP8* subunits by trypsin; and these subunits form a rigid spike structure on the virion surface. In this study, we investigated the growth of RVAs in cells transduced with type II transmembrane serine proteases (TTSPs), which cleave fusion proteins and promote infection by respiratory viruses, such as influenza viruses, paramyxoviruses, and coronaviruses. We identified TMPRSS2 and TMPRSS11D as host TTSPs that mediate trypsin-independent and multi-cycle infection by human and animal RVA strains. In vitro cleavage assays revealed that recombinant TMPRSS11D cleaved RVA VP4. We also found that TMPRSS2 and TMPRSS11D promote the infectious entry of immature RVA virions, but they could not activate nascent progeny virions in the late phase of infection. This observation differed from the TTSP-mediated activation process of paramyxoviruses, revealing the existence of virus species-specific activation processes in TTSPs. Our study provides new insights into the interaction between RVAs and host factors, and TTSP-transduced cells offer potential advantages for RVA research and development.ImportanceProteolytic cleavage of the viral VP4 protein is essential for virion maturation and infectivity in group A rotaviruses (RVAs). In cell culture, RVAs are propagated in culture medium supplemented with the exogenous protease trypsin, which cleaves VP4 and induces the maturation of progeny RVA virions. In this study, we demonstrated that the host proteases TMPRSS2 and TMPRSS11D mediate the trypsin-independent infection and growth of RVA. Our data revealed that the proteolytic activation of RVAs by TMPRSS2 and TMPRSS11D occurs at the viral entry step. Because TMPRSS2 and TMPRSS11D gene expression induced similar or higher levels of RVA growth as trypsin-supplemented culture, this approach offers potential advantages for RVA research and development.
• MRC5 cells engineered to express ACE2 serve as a model system for the discovery of antivirals targeting SARS-CoV-2.
  Kentaro Uemura; Michihito Sasaki; Takao Sanaki; Shinsuke Toba; Yoshimasa Takahashi; Yasuko Orba; William W Hall; Katsumi Maenaka; Hirofumi Sawa; Akihiko Sato
  Scientific reports, 11, 1, 5376, 5376, 08 Mar. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Although the spread of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) has resulted in a worldwide pandemic, there are currently no virus-specific drugs that are fully effective against SARS-CoV-2. Only a limited number of human-derived cells are capable of supporting SARS-CoV-2 replication and the infectivity of SARS-CoV-2 in these cells remains poor. In contrast, monkey-derived Vero cells are highly susceptibility to infection with SARS-CoV-2, although they are not suitable for the study of antiviral effects by small molecules due to their limited capacity to metabolize drugs compared to human-derived cells. In this study, our goal was to generate a virus-susceptible human cell line that would be useful for the identification and testing of candidate drugs. Towards this end, we stably transfected human lung-derived MRC5 cells with a lentiviral vector encoding angiotensin-converting enzyme 2 (ACE2), the cellular receptor for SARS-CoV-2. Our results revealed that SARS-CoV-2 replicates efficiently in MRC5/ACE2 cells. Furthermore, viral RNA replication and progeny virus production were significantly reduced in response to administration of the replication inhibitor, remdesivir, in MRC5/ACE2 cells compared with Vero cells. We conclude that the MRC5/ACE2 cells will be important in developing specific anti-viral therapeutics and will assist in vaccine development to combat SARS-CoV-2 infections.
• An African tick flavivirus forming an independent clade exhibits unique exoribonuclease-resistant RNA structures in the genomic 3'-untranslated region.
  Hayato Harima; Yasuko Orba; Shiho Torii; Yongjin Qiu; Masahiro Kajihara; Yoshiki Eto; Naoya Matsuta; Bernard M Hang'ombe; Yuki Eshita; Kentaro Uemura; Keita Matsuno; Michihito Sasaki; Kentaro Yoshii; Ryo Nakao; William W Hall; Ayato Takada; Takashi Abe; Michael T Wolfinger; Martin Simuunza; Hirofumi Sawa
  Scientific reports, 11, 1, 4883, 4883, 01 Mar. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Tick-borne flaviviruses (TBFVs) infect mammalian hosts through tick bites and can cause various serious illnesses, such as encephalitis and hemorrhagic fevers, both in humans and animals. Despite their importance to public health, there is limited epidemiological information on TBFV infection in Africa. Herein, we report that a novel flavivirus, Mpulungu flavivirus (MPFV), was discovered in a Rhipicephalus muhsamae tick in Zambia. MPFV was found to be genetically related to Ngoye virus detected in ticks in Senegal, and these viruses formed a unique lineage in the genus Flavivirus. Analyses of dinucleotide contents of flaviviruses indicated that MPFV was similar to those of other TBFVs with a typical vertebrate genome signature, suggesting that MPFV may infect vertebrate hosts. Bioinformatic analyses of the secondary structures in the 3'-untranslated regions (UTRs) revealed that MPFV exhibited unique exoribonuclease-resistant RNA (xrRNA) structures. Utilizing biochemical approaches, we clarified that two xrRNA structures of MPFV in the 3'-UTR could prevent exoribonuclease activity. In summary, our findings provide new information regarding the geographical distribution of TBFV and xrRNA structures in the 3'-UTR of flaviviruses.
• TMPRSS11D and TMPRSS13 Activate the SARS-CoV-2 Spike Protein.
  Mai Kishimoto; Kentaro Uemura; Takao Sanaki; Akihiko Sato; William W Hall; Hiroaki Kariwa; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki
  Viruses, 13, 3, 28 Feb. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) utilizes host proteases, including a plasma membrane-associated transmembrane protease, serine 2 (TMPRSS2) to cleave and activate the virus spike protein to facilitate cellular entry. Although TMPRSS2 is a well-characterized type II transmembrane serine protease (TTSP), the role of other TTSPs on the replication of SARS-CoV-2 remains to be elucidated. Here, we have screened 12 TTSPs using human angiotensin-converting enzyme 2-expressing HEK293T (293T-ACE2) cells and Vero E6 cells and demonstrated that exogenous expression of TMPRSS11D and TMPRSS13 enhanced cellular uptake and subsequent replication of SARS-CoV-2. In addition, SARS-CoV-1 and SARS-CoV-2 share the same TTSPs in the viral entry process. Our study demonstrates the impact of host TTSPs on infection of SARS-CoV-2, which may have implications for cell and tissue tropism, for pathogenicity, and potentially for vaccine development.
• Mastomys natalensis is a possible natural rodent reservoir for encephalomyocarditis virus.
  Mai Kishimoto; Bernard M Hang'ombe; William W Hall; Yasuko Orba; Hirofumi Sawa; Michihito Sasaki
  The Journal of general virology, 102, 3, 03 Feb. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Encephalomyocarditis virus (EMCV) infects a wide range of hosts and can cause encephalitis, myocarditis, reproductive disorders and diabetes mellitus in selected mammalian species. As for humans, EMCV infection seems to occur by the contact with animals and can cause febrile illnesses in some infected patients. Here we isolated EMCV strain ZM12/14 from a natal multimammate mouse (Mastomys natalensis: M. natalensis) in Zambia. Pairwise sequence similarity of the ZM12/14 P1 region consisting of antigenic capsid proteins showed the highest similarity of nucleotide (80.7 %) and amino acid (96.2%) sequence with EMCV serotype 1 (EMCV-1). Phylogenetic analysis revealed that ZM12/14 clustered into EMCV-1 at the P1 and P3 regions but segregated from known EMCV strains at the P2 region, suggesting a unique evolutionary history. Reverse transcription PCR (RT-PCR) screening and neutralizing antibody assays for EMCV were performed using collected tissues and serum from various rodents (n=179) captured in different areas in Zambia. We detected the EMCV genome in 19 M. natalensis (19/179=10.6 %) and neutralizing antibody for EMCV in 33 M. natalensis (33/179=18.4 %). However, we did not detect either the genome or neutralizing antibody in other rodent species. High neutralizing antibody litres (≧320) were observed in both RT-PCR-negative and -positive animals. Inoculation of ZM12/14 caused asymptomatic persistent infection in BALB/c mice with high antibody titres and high viral loads in some organs, consistent with the above epidemiological results. This study is the first report of the isolation of EMCV in Zambia, suggesting that M. natalensis may play a role as a natural reservoir of infection.
• Diverse mosquito-specific flaviviruses in the Bolivian Amazon basin.
  Yasuko Orba; Keita Matsuno; Ryo Nakao; Kirill Kryukov; Yumi Saito; Fumihiko Kawamori; Ariel Loza Vega; Tokiko Watanabe; Tadashi Maemura; Michihito Sasaki; William W Hall; Roy A Hall; Juan Antonio Pereira; So Nakagawa; Hirofumi Sawa
  The Journal of general virology, 102, 3, 08 Jan. 2021, [Peer-reviewed], [Lead author, Corresponding author], [International Magazine]
  English, Scientific journal, The genus Flavivirus includes a range of mosquito-specific viruses in addition to well-known medically important arboviruses. Isolation and comprehensive genomic analyses of viruses in mosquitoes collected in Bolivia resulted in the identification of three novel flavivirus species. Psorophora flavivirus (PSFV) was isolated from Psorophora albigenu. The coding sequence of the PSFV polyprotein shares 60 % identity with that of the Aedes-associated lineage II insect-specific flavivirus (ISF), Marisma virus. Isolated PSFV replicates in both Aedes albopictus- and Aedes aegypti-derived cells, but not in mammalian Vero or BHK-21 cell lines. Two other flaviviruses, Ochlerotatus scapularis flavivirus (OSFV) and Mansonia flavivirus (MAFV), which were identified from Ochlerotatus scapularis and Mansonia titillans, respectively, group with the classical lineage I ISFs. The protein coding sequences of these viruses share only 60 and 40 % identity with the most closely related of known lineage I ISFs, including Xishuangbanna aedes flavivirus and Sabethes flavivirus, respectively. Phylogenetic analysis suggests that MAFV is clearly distinct from the groups of the current known Culicinae-associated lineage I ISFs. Interestingly, the predicted amino acid sequence of the MAFV capsid protein is approximately two times longer than that of any of the other known flaviviruses. Our results indicate that flaviviruses with distinct features can be found at the edge of the Bolivian Amazon basin at sites that are also home to dense populations of human-biting mosquitoes.
• First COVID-19 case in Zambia - Comparative phylogenomic analyses of SARS-CoV-2 detected in African countries.
  Edgar Simulundu; Francis Mupeta; Pascalina Chanda-Kapata; Ngonda Saasa; Katendi Changula; Walter Muleya; Simbarashe Chitanga; Miniva Mwanza; Paul Simusika; Herman Chambaro; Benjamin Mubemba; Masahiro Kajihara; Duncan Chanda; Lloyd Mulenga; Sombo Fwoloshi; Aaron Lunda Shibemba; Fred Kapaya; Paul Zulu; Kunda Musonda; Mwaka Monze; Nyambe Sinyange; Mazyanga L Mazaba; Muzala Kapin'a; Peter J Chipimo; Raymond Hamoonga; Davie Simwaba; William Ngosa; Albertina N Morales; Nkomba Kayeyi; John Tembo; Mathew Bates; Yasuko Orba; Hirofumi Sawa; Ayato Takada; King S Nalubamba; Kennedy Malama; Victor Mukonka; Alimuddin Zumla; Nathan Kapata
  International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases, 102, 455, 459, Jan. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Since its first discovery in December 2019 in Wuhan, China, COVID-19, caused by the novel coronavirus SARS-CoV-2, has spread rapidly worldwide. While African countries were relatively spared initially, the initial low incidence of COVID-19 cases was not sustained for long due to continuing travel links between China, Europe and Africa. In preparation, Zambia had applied a multisectoral national epidemic disease surveillance and response system resulting in the identification of the first case within 48 h of the individual entering the country by air travel from a trip to France. Contact tracing showed that SARS-CoV-2 infection was contained within the patient's household, with no further spread to attending health care workers or community members. Phylogenomic analysis of the patient's SARS-CoV-2 strain showed that it belonged to lineage B.1.1., sharing the last common ancestor with SARS-CoV-2 strains recovered from South Africa. At the African continental level, our analysis showed that B.1 and B.1.1 lineages appear to be predominant in Africa. Whole genome sequence analysis should be part of all surveillance and case detection activities in order to monitor the origin and evolution of SARS-CoV-2 lineages across Africa.
• SARS-CoV-2 variants with mutations at the S1/S2 cleavage site are generated in vitro during propagation in TMPRSS2-deficient cells.
  Michihito Sasaki; Kentaro Uemura; Akihiko Sato; Shinsuke Toba; Takao Sanaki; Katsumi Maenaka; William W Hall; Yasuko Orba; Hirofumi Sawa
  PLoS pathogens, 17, 1, e1009233, Jan. 2021, [Peer-reviewed], [International Magazine]
  English, Scientific journal, The spike (S) protein of Severe Acute Respiratory Syndrome-Coronavirus-2 (SARS-CoV-2) binds to a host cell receptor which facilitates viral entry. A polybasic motif detected at the cleavage site of the S protein has been shown to broaden the cell tropism and transmissibility of the virus. Here we examine the properties of SARS-CoV-2 variants with mutations at the S protein cleavage site that undergo inefficient proteolytic cleavage. Virus variants with S gene mutations generated smaller plaques and exhibited a more limited range of cell tropism compared to the wild-type strain. These alterations were shown to result from their inability to utilize the entry pathway involving direct fusion mediated by the host type II transmembrane serine protease, TMPRSS2. Notably, viruses with S gene mutations emerged rapidly and became the dominant SARS-CoV-2 variants in TMPRSS2-deficient cells including Vero cells. Our study demonstrated that the S protein polybasic cleavage motif is a critical factor underlying SARS-CoV-2 entry and cell tropism. As such, researchers should be alert to the possibility of de novo S gene mutations emerging in tissue-culture propagated virus strains.
• Identification of quinolone derivatives as effective anti-Dengue virus agents.
  Haruaki Nobori; Kentaro Uemura; Shinsuke Toba; Takao Sanaki; Takao Shishido; William W Hall; Yasuko Orba; Hirofumi Sawa; Akihiko Sato
  Antiviral research, 184, 104969, 104969, Dec. 2020, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Dengue virus (DENV) infection is one of the most important infectious diseases in tropical and subtropical regions around the world. Previously, we performed an initial phenotypic screening of 7000 compounds using DENV type 2 (DENV2)-infected BHK-21 cells to identify small molecules which could inhibit virus replication. In this study, we describe two novel compounds with anti-DENV2 activity, tentatively named Compound-X and Compound-Y. Both compounds possess a quinolone skeleton, and the EC50s of Compound-X and Compound-Y against DENV2 were 3.9 μM and 9.2 μM, respectively. Based on a DENV replicon assay, it was suggested that these compounds have anti-DENV2 activity by inhibition of a step in virus replication. Furthermore, using mutational analysis we obtained compounds-resistant to DENV2 infection and identified a mutation, V130A in the NS5 methyltransferase (MTase) domain. However, these compounds did not inhibit MTase activity. In addition, incorporation of an additional NS1 N246D mutation with the NS5 V130A mutation in DENV2 resulted in recovery of viral replication and a further reduction of the sensitivity to the quinolone compounds by an unknown mechanism. Therefore further investigations are required to clarify the antiviral mechanisms of these quinolone compounds.
• The Lethal(2)-Essential-for-Life [L(2)EFL] Gene Family Modulates Dengue Virus Infection in Aedes aegypti.
  Lucky R Runtuwene; Shuichi Kawashima; Victor D Pijoh; Josef S B Tuda; Kyoko Hayashida; Junya Yamagishi; Chihiro Sugimoto; Shoko Nishiyama; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa; Tomohiko Takasaki; Anthony A James; Takashi Kobayashi; Yuki Eshita
  International journal of molecular sciences, 21, 20, 12 Oct. 2020, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Efforts to determine the mosquito genes that affect dengue virus replication have identified a number of candidates that positively or negatively modify amplification in the invertebrate host. We used deep sequencing to compare the differential transcript abundances in Aedes aegypti 14 days post dengue infection to those of uninfected A. aegypti. The gene lethal(2)-essential-for-life [l(2)efl], which encodes a member of the heat shock 20 protein (HSP20) family, was upregulated following dengue virus type 2 (DENV-2) infection in vivo. The transcripts of this gene did not exhibit differential accumulation in mosquitoes exposed to insecticides or pollutants. The induction and overexpression of l(2)efl gene products using poly(I:C) resulted in decreased DENV-2 replication in the cell line. In contrast, the RNAi-mediated suppression of l(2)efl gene products resulted in enhanced DENV-2 replication, but this enhancement occurred only if multiple l(2)efl genes were suppressed. l(2)efl homologs induce the phosphorylation of eukaryotic initiation factor 2α (eIF2α) in the fruit fly Drosophila melanogaster, and we confirmed this finding in the cell line. However, the mechanism by which l(2)efl phosphorylates eIF2α remains unclear. We conclude that l(2)efl encodes a potential anti-dengue protein in the vector mosquito.
• Discoveries of Exoribonuclease-Resistant Structures of Insect-Specific Flaviviruses Isolated in Zambia.
  Christida E Wastika; Hayato Harima; Michihito Sasaki; Bernard M Hang'ombe; Yuki Eshita; Yongjin Qiu; William W Hall; Michael T Wolfinger; Hirofumi Sawa; Yasuko Orba
  Viruses, 12, 9, 11 Sep. 2020, [Peer-reviewed], [Corresponding author], [International Magazine]
  English, Scientific journal, To monitor the arthropod-borne virus transmission in mosquitoes, we have attempted both to detect and isolate viruses from 3304 wild-caught female mosquitoes in the Livingstone (Southern Province) and Mongu (Western Province) regions in Zambia in 2017. A pan-flavivirus RT-PCR assay was performed to identify flavivirus genomes in total RNA extracted from mosquito lysates, followed by virus isolation and full genome sequence analysis using next-generation sequencing and rapid amplification of cDNA ends. We isolated a newly identified Barkedji virus (BJV Zambia) (10,899 nt) and a novel flavivirus, tentatively termed Barkedji-like virus (BJLV) (10,885 nt) from Culex spp. mosquitoes which shared 96% and 75% nucleotide identity with BJV which has been isolated in Israel, respectively. These viruses could replicate in C6/36 cells but not in mammalian and avian cell lines. In parallel, a comparative genomics screening was conducted to study evolutionary traits of the 5'- and 3'-untranslated regions (UTRs) of isolated viruses. Bioinformatic analyses of the secondary structures in the UTRs of both viruses revealed that the 5'-UTRs exhibit canonical stem-loop structures, while the 3'-UTRs contain structural homologs to exoribonuclease-resistant RNAs (xrRNAs), SL-III, dumbbell, and terminal stem-loop (3'SL) structures. The function of predicted xrRNA structures to stop RNA degradation by Xrn1 exoribonuclease was further proved by the in vitro Xrn1 resistance assay.
• Co-Circulation of Multiple Serotypes of Bluetongue Virus in Zambia.
  Herman M Chambaro; Michihito Sasaki; Edgar Simulundu; Isaac Silwamba; Yona Sinkala; Gabriel Gonzalez; David Squarre; Paul Fandamu; Caesar H Lubaba; Musso Munyeme; Alikhadio Maseko; Choopa Chimvwele; Liywalii Mataa; Lynnfield E Mooya; Andrew N Mukubesa; Hayato Harima; Kenny L Samui; Hetron M Munang'andu; Martin Simuunza; King S Nalubamba; Yongjin Qiu; Michael J Carr; William W Hall; Yuki Eshita; Hirofumi Sawa; Yasuko Orba
  Viruses, 12, 9, 31 Aug. 2020, [Peer-reviewed], [Corresponding author], [International Magazine]
  English, Scientific journal, Bluetongue (BT) is an arthropod-borne viral disease of ruminants with serious trade and socio-economic implications. Although the disease has been reported in a number of countries in sub-Saharan Africa, there is currently no information on circulating serotypes and disease distribution in Zambia. Following surveillance for BT in domestic and wild ruminants in Zambia, BT virus (BTV) nucleic acid and antibodies were detected in eight of the 10 provinces of the country. About 40% (87/215) of pooled blood samples from cattle and goats were positive for BTV nucleic acid, while one hartebeest pool (1/43) was positive among wildlife samples. Sequence analysis of segment 2 revealed presence of serotypes 3, 5, 7, 12 and 15, with five nucleotypes (B, E, F, G and J) being identified. Segment 10 phylogeny showed Zambian BTV sequences clustering with Western topotype strains from South Africa, intimating likely transboundary spread of BTV in Southern Africa. Interestingly, two Zambian viruses and one isolate from Israel formed a novel clade, which we designated as Western topotype 4. The high seroprevalence (96.2%) in cattle from Lusaka and Central provinces and co-circulation of multiple serotypes showed that BT is widespread, underscoring the need for prevention and control strategies.
• Characterization of mammalian orthoreoviruses isolated from faeces of pigs in Zambia.
  Hayato Harima; Michihito Sasaki; Masahiro Kajihara; Gabriel Gonzalez; Edgar Simulundu; Eugene C Bwalya; Yongjin Qiu; Kosuke Okuya; Mao Isono; Yasuko Orba; Ayato Takada; Bernard M Hang'ombe; Aaron S Mweene; Hirofumi Sawa
  The Journal of general virology, 101, 10, 1027, 1036, 24 Jul. 2020, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Mammalian orthoreovirus (MRV) has been identified in humans, livestock and wild animals; this wide host range allows individual MRV to transmit into multiple species. Although several interspecies transmission and genetic reassortment events of MRVs among humans, livestock and wildlife have been reported, the genetic diversity and geographic distribution of MRVs in Africa are poorly understood. In this study, we report the first isolation and characterization of MRVs circulating in a pig population in Zambia. In our screening, MRV genomes were detected in 19.7 % (29/147) of faecal samples collected from pigs by reverse transcription PCR. Three infectious MRV strains (MRV-85, MRV-96 and MRV-117) were successfully isolated, and their complete genomes were sequenced. Recombination analyses based on the complete genome sequences of the isolated MRVs demonstrated that MRV-96 shared the S3 segment with a different MRV isolated from bats, and that the L1 and M3 segments of MRV-117 originated from bat and human MRVs, respectively. Our results suggest that the isolated MRVs emerged through genetic reassortment events with interspecies transmission. Given the lack of information regarding MRVs in Africa, further surveillance of MRVs circulating among humans, domestic animals and wildlife is required to assess potential risk for humans and animals.
• Host ESCRT factors are recruited during chikungunya virus infection and are required for the intracellular viral replication cycle.
  Shiho Torii; Yasuko Orba; Michihito Sasaki; Koshiro Tabata; Yuji Wada; Michael Carr; Jody Hobson-Peters; Roy A Hall; Ayato Takada; Takasuke Fukuhara; Yoshiharu Matsuura; William W Hall; Hirofumi Sawa
  The Journal of biological chemistry, 295, 23, 7941, 7957, 05 Jun. 2020, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Chikungunya fever is a re-emerging zoonotic disease caused by chikungunya virus (CHIKV), a member of the Alphavirus genus in the Togaviridae family. Only a few studies have reported on the host factors required for intracellular CHIKV trafficking. Here, we conducted an imaging-based siRNA screen to identify human host factors for intracellular trafficking that are involved in CHIKV infection, examined their interactions with CHIKV proteins, and investigated the contributions of these proteins to CHIKV infection. The results of the siRNA screen revealed that host endosomal sorting complexes required for transport (ESCRT) proteins are recruited during CHIKV infection. Co-immunoprecipitation analyses revealed that both structural and nonstructural CHIKV proteins interact with hepatocyte growth factor-regulated tyrosine kinase substrate (HGS), a component of the ESCRT-0 complex. We also observed that HGS co-localizes with the E2 protein of CHIKV and with dsRNA, a marker of the replicated CHIKV genome. Results from gene knockdown analyses indicated that, along with other ESCRT factors, HGS facilitates both genome replication and post-translational steps during CHIKV infection. Moreover, we show that ESCRT factors are also required for infections with other alphaviruses. We conclude that during CHIKV infection, several ESCRT factors are recruited via HGS and are involved in viral genome replication and post-translational processing of viral proteins.
• Evidence for exposure of asymptomatic domestic pigs to African swine fever virus during an inter-epidemic period in Zambia.
  Herman M Chambaro; Michihito Sasaki; Yona Sinkala; Gabriel Gonzalez; David Squarre; Paul Fandamu; Caesar Lubaba; Liywalii Mataa; Misheck Shawa; Kabemba E Mwape; Sarah Gabriël; Mwelwa Chembensofu; Michael J Carr; William W Hall; Yongjin Qiu; Masahiro Kajihara; Ayato Takada; Yasuko Orba; Edgar Simulundu; Hirofumi Sawa
  Transboundary and emerging diseases, 67, 6, 2741, 2752, 20 May 2020, [Peer-reviewed], [International Magazine]
  English, Scientific journal, African swine fever (ASF) causes persistent outbreaks in endemic and non-endemic regions in Zambia. However, the epidemiology of the disease is poorly understood, particularly during the inter-epidemic periods. We conducted surveillance for ASF in asymptomatic domestic pigs and soft ticks in selected Zambian provinces. While serum samples (n = 1,134) were collected from crossbred pigs from all study sites between 2014 and 2017, whole blood (n = 300) was collected from both crossbred and indigenous pigs in Eastern Province (EP) in 2017. Soft ticks were collected from Mosi-oa-Tunya National Park in Southern Province (SP) in 2019. Sera were screened for antibodies against ASF by ELISA while genome detection in whole blood and soft ticks was conducted by PCR. Ticks were identified morphologically and by phylogenetic analysis of the 16S rRNA gene. Seroprevalence was highest in EP (50.9%, 95% CI [47.0-54.9]) compared to significantly lower rates in SP (2.9%, 95% CI [1.6-5.1]). No antibodies to ASFV were detected in Lusaka Province. In EP, the prevalence of ASFV genome was 11.7% (35/300), significantly higher (OR = 6.2, 95% CI [2.4-16.6]) in indigenous pigs compared to crossbred pigs. The pooled prevalence of ASFV genome in ticks was 11.0%, 95% CI [8.5-13.9]. Free-range husbandry system was the only factor that was significantly associated with seropositive (p < .0001, OR = 39.3) and PCR-positive results (p < .001, OR = 5.7). Phylogenetically, based on the p72 gene, ASFV from Ornithodoros moubata ticks detected in this study belonged to genotype I, but they separated into two distinct clusters. Besides confirming ASF endemicity in EP and the presence of ASFV-infected ticks in SP, these results provide evidence for exposure of domestic pigs to ASFV in non-endemic regions during the inter-epidemic period.
• West Nile Virus in Farmed Crocodiles, Zambia, 2019.
  Edgar Simulundu; Kunda Ndashe; Herman M Chambaro; David Squarre; Paul Michael Reilly; Simbarashe Chitanga; Katendi Changula; Andrew N Mukubesa; Joseph Ndebe; John Tembo; Nathan Kapata; Matthew Bates; Yona Sinkala; Bernard M Hang'ombe; King S Nalubamba; Masahiro Kajihara; Michihito Sasaki; Yasuko Orba; Ayato Takada; Hirofumi Sawa
  Emerging infectious diseases, 26, 4, 811, 814, Apr. 2020, [Peer-reviewed], [International Magazine]
  English, Scientific journal, We detected West Nile virus (WNV) nucleic acid in crocodiles (Crocodylus niloticus) in Zambia. Phylogenetically, the virus belonged to lineage 1a, which is predominant in the Northern Hemisphere. These data provide evidence that WNV is circulating in crocodiles in Africa and increases the risk for animal and human transmission.
• Detection of novel orthoreovirus genomes in shrew (Crocidura hirta) and fruit bat (Rousettus aegyptiacus).
  Hayato Harima; Michihito Sasaki; Masahiro Kajihara; Akina Mori-Kajihara; Bernard M Hang'ombe; Katendi Changula; Yasuko Orba; Hirohito Ogawa; Martin Simuunza; Reiko Yoshida; Aaron Mweene; Ayato Takada; Hirofumi Sawa
  The Journal of veterinary medical science, 82, 2, 162, 167, 04 Feb. 2020, [Peer-reviewed], [Domestic magazines]
  English, Scientific journal, Orthoreoviruses have been indentified in several mammals, however, there is no information about orthoreoviruses in shrews. In this study, we screened wild animals in Zambia, including shrews, rodents, and bats for the detection of orthoreoviruses. Two orthoreovirus RNA genomes were detected from a shrew intestinal-contents (1/24) and a bat colon (1/96) sample by reverse-transcription (RT)-PCR targeting the RNA-dependent RNA polymerase gene of orthoreoviruses. Phylogenetic analyses revealed that each of the identified orthoreoviruses formed a distinct branch among members of the Orthoreovirus genus. This is the first report that shrews are susceptible to orthoreovirus infection. Our results suggest the existence of undiscovered orthoreoviruses in shrews and provide important information about the genetic diversity of orthoreoviruses.
• West Nile virus capsid protein inhibits autophagy by AMP-activated protein kinase degradation in neurological disease development.
  Shintaro Kobayashi; Kentaro Yoshii; Wallaya Phongphaew; Memi Muto; Minato Hirano; Yasuko Orba; Hirofumi Sawa; Hiroaki Kariwa
  PLoS pathogens, 16, 1, e1008238, Jan. 2020, [Peer-reviewed], [International Magazine]
  English, Scientific journal, West Nile virus (WNV) belongs to the Flaviviridae family and has emerged as a significant cause of viral encephalitis in birds and animals including humans. WNV replication directly induces neuronal injury, followed by neuronal cell death. We previously showed that accumulation of ubiquitinated protein aggregates was involved in neuronal cell death in the WNV-infected mouse brain. In this study, we attempted to elucidate the mechanisms of the accumulation of protein aggregates in the WNV-infected cells. To identify the viral factor inducing the accumulation of ubiquitinated proteins, intracellular accumulation of ubiquitinated proteins was examined in the cells expressing the viral protein. Expression of capsid (C) protein induced the accumulation, while mutations at residues L51 and A52 in C protein abrogated the accumulation. Wild-type (WT) or mutant WNV in which mutations were introduced into the residues was inoculated into human neuroblastoma cells. The expression levels of LC3-II, an autophagy-related protein, and AMP-activated protein kinase (AMPK), an autophagy inducer, were reduced in the cells infected with WT WNV, while the reduction was not observed in the cells infected with WNV with the mutations in C protein. Similarly, ubiquitination and degradation of AMPK were only observed in the cells infected with WT WNV. In the cells expressing C protein, AMPK was co-precipitated with C protein and mutations in L51 and A52 reduced the interaction. Although the viral replication was not affected, the accumulation of ubiquitinated proteins in brain and neurological symptoms were attenuated in the mouse inoculated with WNV with the mutations in C protein as compared with that with WT WNV. Taken together, ubiquitination and degradation of AMPK by C protein resulted in the inhibition of autophagy and the accumulation of protein aggregates, which contributes to the development of neurological disease.
• Discovery and genetic characterization of diverse smacoviruses in Zambian non-human primates
  Paulina D. Anindita; Michihito Sasaki; Gabriel Gonzalez; Wallaya Phongphaew; Michael Carr; Bernard M. Hang’ombe; Aaron S. Mweene; Kimihito Ito; Yasuko Orba; Hirofumi Sawa
  Scientific Reports, 9, 1, Nature Publishing Group, 01 Dec. 2019
  English, Scientific journal
• Upregulated expression of the antioxidant sestrin 2 identified by transcriptomic analysis of Japanese encephalitis virus-infected SH-SY5Y neuroblastoma cells.
  Carr M; Gonzalez G; Martinelli A; Wastika CE; Ito K; Orba Y; Sasaki M; Hall WW; Sawa H
  Virus genes, 55, 5, 630, 642, Oct. 2019, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Japanese encephalitis virus (JEV) exerts a profound burden of viral encephalitis. We have investigated the differentially expressed transcripts in the neuronal transcriptome during JEV infection by RNA sequencing (RNA-Seq) of virus-infected SH-SY5Y human neuroblastoma cells. Gene ontology analysis revealed significant enrichment from two main pathways: endoplasmic reticulum (ER)-nucleus signaling (P value: 5.75E-18; false discovery rate [FDR] 3.11E-15) and the ER unfolded protein response (P value: 7.58E-18; FDR 3.11E-15). qPCR validation showed significant upregulation and differential expression (P < 0.01) of ER stress-signaling transcripts (SESN2, TRIB3, DDIT3, DDIT4, XBP1, and ATF4) at 24 h post-infection for both low (LN) and high (HN) neurovirulence JEV strains. Immunoblot analysis following JEV infection of SH-SY5Y cells showed an increase in levels of SESN2 protein following JEV infection. Similarly, Zika virus (MR766) infection of SH-SY5Y showed a titer-dependent increase in ER stress-signaling transcripts; however, this was absent or diminished for DDIT4 and ATF4, respectively, suggestive of differences in the induction of stress-response transcripts between flaviviruses. Interestingly, SLC7A11 and SLC3A2 mRNA were also both deregulated in JEV-infected SH-SY5Y cells and encode the two constituent subunits of the plasma membrane xCT amino acid antiporter that relieves oxidative stress by export of glutamate and import of cystine. Infection of SH-SY5Y and HEK293T cells by the JEV HN strain Sw/Mie/40/2004 lead to significant upregulation of the SLC7A11 mRNA to levels comparable to DDIT3. Our findings suggest upregulation of antioxidants including SESN2 and, also, the xCT antiporter occurs to counteract the oxidative stress elicited by JEV infection.
• Autopsy findings in the early stage of amyotrophic lateral sclerosis with "dropped head" syndrome.
  Satoshi Tanikawa; Mishie Tanino; Lei Wang; Marin Ishikawa; Masaya Miyazaki; Masumi Tsuda; Yasuko Orba; Hirofumi Sawa; Kotarou Matoba; Nishio Nakamura; Kazuo Nagashima; William W Hall; Shinya Tanaka
  Neuropathology : official journal of the Japanese Society of Neuropathology, 39, 5, 374, 377, Oct. 2019, [Peer-reviewed], [International Magazine]
  English, Dropped head syndrome (DHS) has been rarely observed in amyotrophic lateral sclerosis (ALS), and the neuropathological findings of this condition have almost never been described. The identification of transactivation response DNA-binding protein 43 kDa (TDP-43), which binds to RNA/DNA has provided a new method for studying ALS and frontotemporal lobar degeneration (FTLD). Post-mortem examination of an adult sudden death case of a 71-year-old patient who complained of DHS exhibited severe loss of anterior motor neurons in the cervical cord (C4-6). Loss of nerve fibers of the anterior roots was striking compared with posterior roots, together with marked neurogenic atrophy of posterior muscles semispinalis cervicis. Bunina bodies were found in large neurons of Betz giant cells, but not in the motor neurons of spinal cords, or neurons of bulbar regions. Phosphorylated TDP-43 (p-TDP-43)-positive structures were detected in the residual neurons of the cervical, thoracic and lumber cords, hypoglossal nucleus, cerebellar dentate nucleus and parahippocampal cortex, together with ubiquitin-positive inclusions. Phosphorylated Tau positive structures in neuronal cytoplasm were found in the amygdala, entorhinal cortex and parahippocampal cortex, some of which co-expressed p-TDP-43. The medial zone of cervical cords may be the first onset site, and that is the cause of head drop in the early stage of ALS. In spite of detailed examination, the direct cause of sudden death was not verified. This autopsy report revealed the relation of DHS which is a rare clinical manifestation of ALS, and neuropathological findings.
• Serological evidence of Zika virus infection in non-human primates in Zambia.
  Christida E Wastika; Michihito Sasaki; Kentaro Yoshii; Paulina D Anindita; Bernard M Hang'ombe; Aaron S Mweene; Shintaro Kobayashi; Hiroaki Kariwa; Michael J Carr; William W Hall; Yuki Eshita; Yasuko Orba; Hirofumi Sawa
  Archives of virology, 164, 8, 2165, 2170, Aug. 2019, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Zika virus (ZIKV) circulation occurs between non-human primates (NHPs) in a sylvatic transmission cycle. To investigate evidence of flavivirus infection in NHPs in Zambia, we performed a plaque reduction neutralization test (PRNT) to quantify neutralizing antibodies. PRNT revealed that sera from NHPs (African green monkeys and baboons) exhibited neutralizing activity against ZIKV (34.4%; 33/96), whereas a PRNT for yellow fever virus using NHP sera showed no neutralization activity. ZIKV genomic RNA was not detected in splenic tissues from NHPs, suggesting that the presence of anti-ZIKV neutralizing antibodies represented resolved infections. Our evidence suggests that ZIKV is maintained in NHP reservoirs in Zambia.
• Marburgvirus in Egyptian Fruit Bats, Zambia.
  Masahiro Kajihara; Bernard M Hang'ombe; Katendi Changula; Hayato Harima; Mao Isono; Kosuke Okuya; Reiko Yoshida; Akina Mori-Kajihara; Yoshiki Eto; Yasuko Orba; Hirohito Ogawa; Yongjin Qiu; Hirofumi Sawa; Edgar Simulundu; Daniel Mwizabi; Musso Munyeme; David Squarre; Victor Mukonka; Aaron Mweene; Ayato Takada
  Emerging infectious diseases, 25, 8, 1577, 1580, Aug. 2019, [Peer-reviewed], [International Magazine]
  English, Scientific journal, We detected Marburg virus genome in Egyptian fruit bats (Rousettus aegyptiacus) captured in Zambia in September 2018. The virus was closely related phylogenetically to the viruses that previously caused Marburg outbreaks in the Democratic Republic of the Congo. This finding demonstrates that Zambia is at risk for Marburg virus disease.
• Publisher Correction: Discovery and genetic characterization of diverse smacoviruses in Zambian non-human primates.
  Anindita PD; Sasaki M; Gonzalez G; Phongphaew W; Carr M; Hang'ombe BM; Mweene AS; Ito K; Orba Y; Sawa H
  Scientific reports, 9, 1, 8502, 8502, Jun. 2019, [Peer-reviewed], [International Magazine]
  English, A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.
• Field diagnosis and genotyping of chikungunya virus using a dried reverse transcription loop-mediated isothermal amplification (LAMP) assay and MinION sequencing.
  Kyoko Hayashida; Yasuko Orba; Patricia C Sequeira; Chihiro Sugimoto; William W Hall; Yuki Eshita; Yutaka Suzuki; Lucky Runtuwene; Patricia Brasil; Guilherme Calvet; Cintia D S Rodrigues; Carolina C Dos Santos; Maria A M Mares-Guia; Junya Yamagishi; Ana M B de Filippis; Hirofumi Sawa
  PLoS neglected tropical diseases, 13, 6, e0007480, Jun. 2019, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Detection and sequencing of chikungunya virus (CHIKV) genome was performed using a combination of a modified reverse transcription loop-mediated isothermal amplification (RT-LAMP) method and a MinION sequencer. We developed the protocol for drying all the reagents for the RT-LAMP in a single reaction tube. Using this system, the CHIKV genome was effectively amplified under isothermal conditions, and used as a template for MinION sequencing with a laptop computer. Our in-house RT-LAMP method and MinION sequencing system were also validated with RNAs and serum samples from recent outbreaks of CHIKV patients in Brazil. The obtained sequence data confirmed the CHIKV outbreaks and identified the genotype. In summary, our established inexpensive on-site genome detection and sequencing system is applicable for both diagnosis of CHIKV infected patients and genotyping of the CHIKV virus in future outbreak in remote areas.
• Identification of Mouse and Human Antibody Repertoires by Next-Generation Sequencing.
  Lin Sun; Naoko Kono; Hiroyuki Toh; Hanbing Xue; Kaori Sano; Tadaki Suzuki; Akira Ainai; Yasuko Orba; Junya Yamagishi; Hideki Hasegawa; Yoshimasa Takahashi; Shigeyuki Itamura; Kazuo Ohnishi
  Journal of visualized experiments : JoVE, 145, 15 Mar. 2019, [Peer-reviewed], [International Magazine]
  English, Scientific journal, The immense adaptability of antigen recognition by antibodies is the basis of the acquired immune system. Despite our understanding of the molecular mechanisms underlying the production of the vast repertoire of antibodies by the acquired immune systems, it has not yet been possible to arrive at a global view of a complete antibody repertoire. In particular, B cell repertoires have been regarded as a black box because of their astronomical number of antibody clones. However, next-generation sequencing technologies are enabling breakthroughs to increase our understanding of the B cell repertoire. In this report, we describe a simple and efficient method to visualize and analyze whole individual mouse and human antibody repertoires. From the immune organs, representatively from spleen in mice and peripheral blood mononuclear cells in humans, total RNA was prepared, reverse transcribed, and amplified using the 5'-RACE method. Using a universal forward primer and antisense primers for the antibody class-specific constant domains, antibody mRNAs were uniformly amplified in proportions reflecting their frequencies in the antibody populations. The amplicons were sequenced by next-generation sequencing (NGS), yielding more than 105 antibody sequences per immunological sample. We describe the protocols for antibody sequence analyses including V(D)J-gene-segment annotation, a bird's-eye view of the antibody repertoire, and our computational methods.
• "Integrated diagnosis" of pilocytic astrocytoma: Molecular diagnostic procedure for an unusual case.
  Ishida Y; Tsuda M; Sawamura Y; Fujii K; Murai H; Horiuchi N; Orba Y; Sawa H; Hall WW; Nagashima K; Tanaka S
  Pathology international, 68, 12, 694, 699, Dec. 2018, [Peer-reviewed], [International Magazine]
  English, Scientific journal, A 24 year-old female presented with a mass lesion in the right temporal lobe. This case was difficult to diagnose using histological and immunological methods and therefore molecular analyses were applied to provide a definitive diagnosis. The tumor was well-demarcated, partially cystic, and irregularly-enhanced on gadolinium-enhanced T1-weighted magnetic resonance images. Pathologically, a large part of the tumor consisted of cells with fine cytoplasmic processes on a myxoid and mucinous background. Cells formed a microcystic structure around the mucinous tissue. Numerous eosinophilic granular bodies, but not Rosenthal fibers, were present. The solid and compact regions of the tumor were composed of fasciculation of dense fibrous glial tissues and occasional multinucleated giant cells. Tumor cells and their fragmented cytoplasmic processes were positively stained with GFAP, while eosinophilic granular bodies were both positive and negative. Xanthomatous changes were not detected and the reticulin fibers were restricted to vascular tissues. The MIB1 index was scored as approximately 10%. In molecular analyses of BRAF, the KIAA1549-BRAF (K16-B9) fusion gene was detected in all tumor regions, whereas BRAF V600E mutation was not detected by either conventional Sanger sequencing or the Eprobe-PCR method. Based on the results of the molecular analyses, this case was diagnosed as pilocytic astrocytoma.
• Identification of group A rotaviruses from Zambian fruit bats provides evidence for long-distance dispersal events in Africa
  Michihito Sasaki; Masahiro Kajihara; Katendi Changula; Akina Mori-Kajihara; Hirohito Ogawa; Bernard M. Hang'ombe; Aaron S. Mweene; Martin Simuunza; Reiko Yoshida; Michael Carr; Yasuko Orba; Ayato Takada; Hirofumi Sawa
  Infection, Genetics and Evolution, 63, 104, 109, Elsevier B.V., 01 Sep. 2018, [Peer-reviewed]
  English, Scientific journal
• Fatal Tickborne Phlebovirus Infection in Captive Cheetahs, Japan.
  Keita Matsuno; Noriyuki Nonoue; Ayako Noda; Nodoka Kasajima; Keita Noguchi; Ai Takano; Hiroshi Shimoda; Yasuko Orba; Mieko Muramatsu; Yoshihiro Sakoda; Ayato Takada; Shinji Minami; Yumi Une; Shigeru Morikawa; Ken Maeda
  Emerging infectious diseases, 24, 9, 1726, 1729, Sep. 2018, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Two captive cheetahs from a zoo in Japan died of a severe fever with thrombocytopenia syndrome-like illness. Severe fever with thrombocytopenia syndrome virus, an endemic tickborne phlebovirus, was detected systemically with secretion of infectious viruses into the saliva. These cases highlight the risk for exposure of captive animals to endemic arthropodborne pathogens.
• First isolation of West Nile virus in Zambia from mosquitoes.
  Orba Y; Hang'ombe BM; Mweene AS; Wada Y; Anindita PD; Phongphaew W; Qiu Y; Kajihara M; Mori-Kajihara A; Eto Y; Sasaki M; Hall WW; Eshita Y; Sawa H
  Transboundary and emerging diseases, 65, 4, 933, 938, Aug. 2018, [Peer-reviewed], [Lead author, Corresponding author]
• Identification of Compound-B, a novel anti-dengue virus agent targeting the non-structural protein 4A
  Haruaki Nobori; Shinsuke Toba; Ryu Yoshida; William W. Hall; Yasuko Orba; Hirofumi Sawa; Akihiko Sato
  Antiviral Research, 155, 60, 66, Elsevier B.V., 01 Jul. 2018, [Peer-reviewed]
  English, Scientific journal
• Single amino acid mutation in dengue virus NS4B protein has opposing effects on viral proliferation in mammalian and mosquito cells.
  Fujiki J; Nobori H; Sato A; Sasaki M; Carr M; Hall WW; Orba Y; Sawa H
  Japanese journal of infectious diseases, 71, 6, 448, 454, Jul. 2018, [Peer-reviewed], [Domestic magazines]
  English, Scientific journal, Dengue virus (DENV) has a considerable impact on the global health and is known to cause morbidity and mortality every year. By passaging DENV2 in baby hamster kidney (BHK)-21 cells, we isolated a mutant clone of DENV2 that shows rapid cytopathic effects in BHK-21 cells as compared with that showed by the parent strain. To investigate the relationship between amino acid mutations and proliferation activity of the isolated DENV2 clone, we performed full genome sequencing and identified 3 amino acid mutations in the coding region, the envelope T120K, NS4A M85T, and NS4B G124A. Genetically modified recombinant DENV2 (rDENV2) carrying the NS4A M85T and NS4B G124A mutations produced higher titers of progeny virus in BHK-21, Vero, and Huh-7 cells than in the wild-type (WT) rDENV2. rDENV2 with mutations at NS4A M85T and NS4B G124A failed to produce any plaques in C6/36 mosquito cell lines. Furthermore, rDENV2 possessing only the NS4B G124A mutation showed no plaque production in C6/36 cells but had higher viral titers in Vero and Huh-7 cells than the WT rDENV2 had. Our results clearly showed that the DENV2 NS4B G124A mutation has opposing effects on the virus proliferation in mosquito and certain mammalian cell lines.
• Ribavirin-related compounds exert in vitro inhibitory effects toward rabies virus.
  Anindita PD; Sasaki M; Okada K; Ito N; Sugiyama M; Saito-Tarashima N; Minakawa N; Shuto S; Otsuguro S; Ichikawa S; Matsuda A; Maenaka K; Orba Y; Sawa H
  Antiviral research, 154, 1, 9, Jun. 2018, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Rabies remains an invariably fatal neurological disease despite the availability of a preventive vaccination and post-exposure prophylaxis that must be immediately administered to the exposed individual before symptom onset. There is no effective medication for treatment during the symptomatic phase. Ribavirin, a guanine nucleoside analog, is a potent inhibitor of rabies virus (RABV) replication in vitro but lacks clinical efficacy. Therefore, we attempted to identify potential ribavirin analogs with comparable or superior anti-RABV activity. Antiviral activity and cytotoxicity of the compounds were initially examined in human neuroblastoma cells. Among the tested compounds, two exhibited a 5- to 27-fold higher anti-RABV activity than ribavirin. Examination of the anti-RABV mechanisms of action of the compounds using time-of-addition and minigenome assays revealed that they inhibited viral genome replication and transcription. Addition of exogenous guanosine to RABV-infected cells diminished the antiviral activity of the compounds, suggesting that they are involved in guanosine triphosphate (GTP) pool depletion by inhibiting inosine monophosphate dehydrogenase (IMPDH). Taken together, our findings underline the potency of nucleoside analogs as a class of antiviral compounds for the development of novel agents against RABV.
• Discovery of Mwinilunga alphavirus: A novel alphavirus in Culex mosquitoes in Zambia.
  Shiho Torii; Yasuko Orba; Bernard M Hang'ombe; Aaron S Mweene; Yuji Wada; Paulina D Anindita; Wallaya Phongphaew; Yongjin Qiu; Masahiro Kajihara; Akina Mori-Kajihara; Yoshiki Eto; Hayato Harima; Michihito Sasaki; Michael Carr; William W Hall; Yuki Eshita; Takashi Abe; Hirofumi Sawa
  Virus research, 250, 31, 36, 02 May 2018, [Peer-reviewed], [Corresponding author], [International Magazine]
  English, Scientific journal, Mosquito-borne alphaviruses are disseminated globally and cause febrile illness in humans and animals. Since the prevalence and diversity of alphaviruses has not been previously investigated in Zambia, reverse transcription PCR was employed as a broad-spectrum approach for the detection of alphaviruses in mosquitoes. From 552 mosquito pools, a novel alphavirus, tentatively named Mwinilunga alphavirus (MWAV), was discovered from a single Culex quinquefasciatus mosquito pool. The full genome of MWAV was subsequently determined, and pairwise comparisons demonstrated that MWAV represented a new alphavirus species. Phylogenetic analyses and a linear discriminant analysis based on the dinucleotide ratios in various virus sequences indicated that MWAV is related to a mosquito-specific alphavirus distinct from other known mosquito-borne alphaviruses due to its inability to replicate in vertebrate cell lines. Further analyses of these novel alphaviruses will help to facilitate a greater understanding of the molecular determinants of host range restriction and the evolutionary relationships of alphaviruses.
• The Role of Heparan Sulfate Proteoglycans as an Attachment Factor for Rabies Virus Entry and Infection.
  Sasaki M; Anindita PD; Ito N; Sugiyama M; Carr M; Fukuhara H; Ose T; Maenaka K; Takada A; Hall WW; Orba Y; Sawa H
  The Journal of infectious diseases, 217, 11, 1740, 1749, May 2018, [Peer-reviewed], [International Magazine]
  English, Scientific journal, Rabies virus (RABV) is the causative agent of fatal neurological disease. Cellular attachment is the initial and essential step for viral infections. Although extensive studies have demonstrated that RABV uses various target cell molecules to mediate infection, no specific molecule has been identified as an attachment factor for RABV infection. Here we demonstrate that cellular heparan sulfate (HS) supports RABV adhesion and subsequent entry into target cells. Enzymatic removal of HS reduced cellular susceptibility to RABV infection, and heparin, a highly sulfated form of HS, blocked viral adhesion and infection. The direct binding between RABV glycoprotein and heparin was demonstrated, and this interaction was shown to require HS N- and 6-O-sulfation. We also revealed that basic amino acids in the ectodomain of RABV glycoprotein serve as major determinants for the RABV-HS interaction. Collectively, our study highlights a previously undescribed role of HS as an attachment factor for RABV infection.
• Detection of novel gammaherpesviruses from fruit bats in Indonesia
  Yuji Wada; Michihito Sasaki; Agus Setiyono; Ekowati Handharyani; Ibenu Rahmadani; Siswatiana Taha; Sri Adiani; Munira Latief; Zainal Abidin Kholilullah; Mawar Subangkit; Shintaro Kobayashi; Ichiro Nakamura; Takashi Kimura; Yasuko Orba; Hirofumi Sawa
  Journal of Medical Microbiology, 67, 3, 415, 422, Microbiology Society, 01 Mar. 2018, [Peer-reviewed]
  English, Scientific journal
• Shape-dependent adjuvanticity of nanoparticle-conjugated RNA adjuvants for intranasal inactivated influenza vaccines
  Taiyu Tazaki; Koshiro Tabata; Akira Ainai; Yuki Ohara; Shintaro Kobayashi; Takafumi Ninomiya; Yasuko Orba; Hideyuki Mitomo; Tetsuo Nakano; Hideki Hasegawa; Kuniharu Ijiro; Hirofumi Sawa; Tadaki Suzuki; Kenichi Niikura
  RSC Advances, 8, 30, 16527, 16536, 2018, [Peer-reviewed]
  English, Scientific journal
• Development of a rapid and quantitative method for the analysis of viral entry and release using a NanoLuc luciferase complementation assay
  Michihito Sasaki; Paulina D. Anindita; Wallaya Phongphaew; Michael Carr; Shintaro Kobayashi; Yasuko Orba; Hirofumi Sawa
  VIRUS RESEARCH, 243, 69, 74, Jan. 2018, [Peer-reviewed]
  English, Scientific journal
• Identification of the same polyomavirus species in different African horseshoe bat species is indicative of short-range host-switching events
  Michael Carr; Gabriel Gonzalez; Michihito Sasaki; Serena E. Dool; Kimihito Ito; Akihiro Ishii; Bernard M. Hang'ombe; Aaron S. Mweene; Emma C. Teeling; William W. Hall; Yasuko Orba; Hirofumi Sawa
  JOURNAL OF GENERAL VIROLOGY, 98, 11, 2771, 2785, Nov. 2017, [Peer-reviewed]
  English, Scientific journal
• Discovery of novel cyclic peptide inhibitors of dengue virus NS2B-NS3 protease with antiviral activity
  Youhei Takagi; Kouhei Matsui; Haruaki Nobori; Haruka Maeda; Akihiko Sato; Takeshi Kurosu; Yasuko Orba; Hirofumi Sawa; Kazunari Hattori; Kenichi Higashino; Yoshito Numata; Yutaka Yoshida
  BIOORGANIC & MEDICINAL CHEMISTRY LETTERS, 27, 15, 3586, 3590, Aug. 2017, [Peer-reviewed]
  English, Scientific journal
• Discovery of a novel antiviral agent targeting the nonstructural protein 4 (nsP4) of chikungunya virus
  Yuji Wada; Yasuko Orba; Michihito Sasaki; Shintaro Kobayashi; Michael J. Carr; Haruaki Nobori; Akihiko Sato; William W. Hall; Hirofumi Sawa
  VIROLOGY, 505, 102, 112, May 2017, [Peer-reviewed]
  English, Scientific journal
• Discovery of African bat polyomaviruses and infrequent recombination in the large T antigen in the Polyomaviridae
  Michael Carr; Gabriel Gonzalez; Michihito Sasaki; Kimihito Ito; Akihiro Ishii; Bernard M. Hang'ombe; Aaron S. Mweene; Yasuko Orba; Hirofumi Sawa
  JOURNAL OF GENERAL VIROLOGY, 98, 4, 726, 738, Apr. 2017, [Peer-reviewed]
  English, Scientific journal
• Isolation of a simian immunodeficiency virus from a malbrouck (Chlorocebus cynosuros)
  Michael Carr; Akira Kawaguchi; Michihito Sasaki; Gabriel Gonzalez; Kimihito Ito; Yuka Thomas; Bernard M. Hang'ombe; Aaron S. Mweene; Guoyan Zhao; David Wang; Yasuko Orba; Akihiro Ishii; Hirofumi Sawa
  ARCHIVES OF VIROLOGY, 162, 2, 543, 548, Feb. 2017, [Peer-reviewed]
  English, Scientific journal
• Valosin-containing protein (VCP/p97) plays a role in the replication of West Nile virus
  Wallaya Phongphaew; Shintaro Kobayashi; Michihito Sasaki; Michael Carr; William W. Hall; Yasuko Orba; Hirofumi Sawa
  VIRUS RESEARCH, 228, 114, 123, Jan. 2017, [Peer-reviewed]
  English, Scientific journal
• Animal Models of Emerging Tick-Borne Phleboviruses: Determining Target Cells in a Lethal Model of SFTSV Infection
  Keita Matsuno; Yasuko Orba; Kimberly Maede-White; Dana Scott; Friederike Feldmann; Mifang Liang; Hideki Ebihara
  FRONTIERS IN MICROBIOLOGY, 8, 104, Jan. 2017, [Peer-reviewed]
  English, Scientific journal
• Multi-reassortant G3P[3] group A rotavirus in a horseshoe bat in Zambia
  Michihito Sasaki; Yasuko Orba; Satoko Sasaki; Gabriel Gonzalez; Akihiro Ishii; Bernard M. Hang'ombe; Aaron S. Mweene; Kimihito Ito; Hirofumi Sawa
  JOURNAL OF GENERAL VIROLOGY, 97, 10, 2488, 2493, Oct. 2016, [Peer-reviewed]
  English, Scientific journal
• Generation of recombinant rabies viruses encoding NanoLuc luciferase for antiviral activity assays
  Paulina Duhita Anindita; Michihito Sasaki; Haruaki Nobori; Akihiko Sato; Michael Carr; Naoto Ito; Makoto Sugiyama; Yasuko Orba; Hirofumi Sawa
  VIRUS RESEARCH, 215, 121, 128, Apr. 2016, [Peer-reviewed]
  English, Scientific journal
• Divergent bufavirus harboured in megabats represents a new lineage of parvoviruses
  Michihito Sasaki; Gabriel Gonzalez; Yuji Wada; Agus Setiyono; Ekowati Handharyani; Ibenu Rahmadani; Siswatiana Taha; Sri Adiani; Munira Latief; Zainal Abidin Kholilullah; Mawar Subangkit; Shintaro Kobayashi; Ichiro Nakamura; Takashi Kimura; Yasuko Orba; Kimihito Ito; Hirofumi Sawa
  SCIENTIFIC REPORTS, 6, 24257, Apr. 2016, [Peer-reviewed]
  English, Scientific journal
• Rab8b Regulates Transport of West Nile Virus Particles from Recycling Endosomes
  Shintaro Kobayashi; Tadaki Suzuki; Akira Kawaguchi; Wallaya Phongphaew; Kentaro Yoshii; Tomohiko Iwano; Akihiro Harada; Hiroaki Kariwa; Yasuko Orba; Hirofumi Sawa
  JOURNAL OF BIOLOGICAL CHEMISTRY, 291, 12, 6559, 6568, Mar. 2016, [Peer-reviewed]
  English, Scientific journal
• Distinct Lineages of Bufavirus in Wild Shrews and Nonhuman Primates
  Michihito Sasaki; Yasuko Orba; Paulina D. Anindita; Akihiro Ishii; Keisuke Ueno; Bernard M. Hang'ombe; Aaron S. Mweeile; Kimihito Ito; Hirofumi Sawa
  EMERGING INFECTIOUS DISEASES, 21, 7, 1230, 1233, Jul. 2015, [Peer-reviewed]
  English, Scientific journal
• Detection of coronavirus genomes in Moluccan naked-backed fruit bats in Indonesia
  Paulina Duhita Anindita; Michihito Sasaki; Agus Setiyono; Ekowati Handharyani; Yasuko Orba; Shintaro Kobayashi; Ibnu Rahmadani; Siswatiana Taha; Sri Adiani; Mawar Subangkit; Ichiro Nakamura; Hirofumi Sawa; Takashi Kimura
  ARCHIVES OF VIROLOGY, 160, 4, 1113, 1118, Apr. 2015, [Peer-reviewed]
  English, Scientific journal
• Detection of novel polyomaviruses in fruit bats in Indonesia
  Shintaro Kobayashi; Michihito Sasaki; Ryo Nakao; Agus Setiyono; Ekowati Handharyani; Yasuko Orba; Ibnu Rahmadani; Siswatiana Taha; Sri Adiani; Mawar Subangkit; Ichiro Nakamura; Takashi Kimura; Hirofumi Sawa
  ARCHIVES OF VIROLOGY, 160, 4, 1075, 1082, Apr. 2015, [Peer-reviewed]
  English, Scientific journal
• Orthopoxvirus infection among wildlife in Zambia
  Yasuko Orba; Michihito Sasaki; Hiroki Yamaguchi; Akihiro Ishii; Yuka Thomas; Hirohito Ogawa; Bernard M. Hang'ombe; Aaron S. Mweene; Shigeru Morikawa; Masayuki Saijo; Hirofumi Sawa
  JOURNAL OF GENERAL VIROLOGY, 96, 390, 394, Feb. 2015, [Peer-reviewed]
  English, Scientific journal
• Metagenomic analysis of the shrew enteric virome reveals novel viruses related to human stool-associated viruses
  Michihito Sasaki; Yasuko Orba; Keisuke Ueno; Akihiro Ishii; Ladslav Moonga; Bernard M. Hang'ombe; Aaron S. Mweene; Kimihito Ito; Hirofumi Sawa
  JOURNAL OF GENERAL VIROLOGY, 96, 440, 452, Feb. 2015, [Peer-reviewed]
  English, Scientific journal
• A nairovirus isolated from African bats causes haemorrhagic gastroenteritis and severe hepatic disease in mice
  Akihiro Ishii; Keisuke Ueno; Yasuko Orba; Michihito Sasaki; Ladslav Moonga; Bernard M. Hang'ombe; Aaron S. Mweene; Takashi Umemura; Kimihito Ito; William W. Hall; Hirofumi Sawa
  NATURE COMMUNICATIONS, 5, 5651, Dec. 2014, [Peer-reviewed]
  English, Scientific journal
• Autophagy inhibits viral genome replication and gene expression stages in West Nile virus infection
  Shintaro Kobayashi; Yasuko Orba; Hiroki Yamaguchi; Kenta Takahashi; Michihito Sasaki; Rie Hasebe; Takashi Kimura; Hirofumi Sawa
  VIRUS RESEARCH, 191, 83, 91, Oct. 2014, [Peer-reviewed]
  English, Scientific journal
• Detection and characterization of zoonotic pathogens of free-ranging non-human primates from Zambia
  Jesca Nakayima; Kyoko Hayashida; Ryo Nakao; Akihiro Ishii; Hirohito Ogawa; Ichiro Nakamura; Ladslav Moonga; Bernard M. Hang'ombe; Aaron S. Mweene; Yuka Thomas; Yasuko Orba; Hirofumi Sawa; Chihiro Sugimoto
  PARASITES & VECTORS, 7, 490, Oct. 2014, [Peer-reviewed]
  English, Scientific journal
• Role of the C-Terminal Region of Vervet Monkey Polyomavirus 1 VP1 in Virion Formation
  Hiroki Yamaguchi; Shintaro Kobayashi; Junki Maruyama; Michihito Sasaki; Ayato Takada; Takashi Kimura; Hirofumi Sawa; Yasuko Orba
  JOURNAL OF VETERINARY MEDICAL SCIENCE, 76, 5, 637, 644, May 2014, [Peer-reviewed]
  English, Scientific journal
• Molecular Epidemiology of Paramyxoviruses in Frugivorous Eidolon helvum Bats in Zambia
  Walter Muleya; Michihito Sasaki; Yasuko Orba; Akihiro Ishii; Yuka Thomas; Emiko Nakagawa; Hirohito Ogawa; Bernard Hang'ombe; Boniface Namangala; Aaron Mweene; Ayato Takada; Takashi Kimura; Hirofumi Sawa
  JOURNAL OF VETERINARY MEDICAL SCIENCE, 76, 4, 611, 614, Apr. 2014, [Peer-reviewed]
  English, Scientific journal
• Molecular epidemiology of paramyxoviruses in Zambian wild rodents and shrews
  Michihito Sasaki; Walter Muleya; Akihiro Ishii; Yasuko Orba; Bernard M. Hang'ombe; Aaron S. Mweene; Ladslav Moonga; Yuka Thomas; Takashi Kimura; Hirofumi Sawa
  JOURNAL OF GENERAL VIROLOGY, 95, 325, 330, Feb. 2014, [Peer-reviewed]
  English, Scientific journal
• Epidemiological and basic research activity targeting polyomaviruses.
  Sawa H; Kobayashi S; Suzuki T; Orba Y
  Uirusu, 64, 25, 34, 1, 2014, [Peer-reviewed]
• Viroporin activity of the JC polyomavirus is regulated by interactions with the adaptor protein complex 3
  Tadaki Suzuki; Yasuko Orba; Yoshinori Makino; Yuki Okada; Yuji Sunden; Hideki Hasegawa; William W. Hall; Hirofumi Sawa
  PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 110, 46, 18668, 18673, Nov. 2013, [Peer-reviewed]
  English, Scientific journal
• Cysteine Residues in the Major Capsid Protein, Vp1, of the JC Virus Are Important for Protein Stability and Oligomer Formation
  Shintaro Kobayashi; Tadaki Suzuki; Manabu Igarashi; Yasuko Orba; Noriko Ohtake; Keita Nagakawa; Kenichi Niikura; Takashi Kimura; Harumi Kasamatsu; Hirofumi Sawa
  PLoS ONE, 8, 10, e76668, 10, 09 Oct. 2013, [Peer-reviewed]
  English, Scientific journal
• Characterization of Japanese encephalitis virus infection in an immortalized mesencephalic cell line, CSM14.1
  Takashi Kimura; Megumi Okumura; Eunmi Kim; Michihito Sasaki; Yasuko Orba; Hirofumi Sawa
  MICROBIOLOGY AND IMMUNOLOGY, 57, 10, 723, 731, Oct. 2013, [Peer-reviewed]
  English, Scientific journal
• Human Parainfluenza Virus Type 3 in Wild Nonhuman Primates, Zambia
  Michihito Sasaki; Akihiro Ishii; Yasuko Orba; Yuka Thomas; Bernard M. Hang'ombe; Ladslav Moonga; Aaron S. Mweene; Hirohito Ogawa; Ichiro Nakamura; Takashi Kimura; Hirofumi Sawa
  EMERGING INFECTIOUS DISEASES, 19, 9, 1500, 1503, Sep. 2013, [Peer-reviewed]
  English, Scientific journal
• ラット中脳由来神経細胞株CSM14.1における日本脳炎ウイルス感染様式
  木村 享史; 奥村 恵; 金 恩美; 佐々木 道仁; 大場 靖子; 澤 洋文
  日本獣医学会学術集会講演要旨集, 156回, 214, 214, (公社)日本獣医学会, Aug. 2013
  Japanese
• オートファジーによるウエストナイルウイルス増殖抑制機構の解明
  小林 進太郎; 大場 靖子; 山口 宏樹; 佐々木 道仁; 長谷部 理絵; 木村 享史; 澤 洋文
  日本獣医学会学術集会講演要旨集, 156回, 267, 267, (公社)日本獣医学会, Aug. 2013
  Japanese
• Cross-Reactivity of Secondary Antibodies against African Rodents and Application for Sero-Surveillance
  Ichiro Nakamura; Bernard Mudenda Hang'ombe; Hirofumi Sawa; Shintaro Kobayashi; Yasuko Orba; Akihiro Ishii; Yuka Thomas; Rie Isozumi; Kumiko Yoshimatsu; Aaron S. Mweene; Ayato Takada; Chihiro Sugimoto; Jiro Arikawa
  JOURNAL OF VETERINARY MEDICAL SCIENCE, 75, 6, 819, 825, Jun. 2013, [Peer-reviewed]
  English, Scientific journal
• Identification of a novel polyomavirus from vervet monkeys in Zambia
  Hiroki Yamaguchi; Shintaro Kobayashi; Akihiro Ishii; Hirohito Ogawa; Ichiro Nakamura; Ladslav Moonga; Bernard M. Hang'ombe; Aaron S. Mweene; Yuka Thomas; Takashi Kimura; Hirofumi Sawa; Yasuko Orba
  Journal of General Virology, 94, 6, 1357, 1364, Pt 6, 01 Jun. 2013, [Peer-reviewed]
  English, Scientific journal
• Gold Nanoparticles as a Vaccine Platform: Influence of Size and Shape on Immunological Responses in Vitro and in Vivo
  Kenichi Niikura; Tatsuya Matsunaga; Tadaki Suzuki; Shintaro Kobayashi; Hiroki Yamaguchi; Yasuko Orba; Akira Kawaguchi; Hideki Hasegawa; Kiichi Kajino; Takafumi Ninomiya; Kuniharu Ijiro; Hirofumi Sawa
  ACS NANO, 7, 5, 3926, 3938, May 2013, [Peer-reviewed]
  English, Scientific journal
• Relationship between Methyl CpG Binding Protein 2 and JC Viral Proteins
  Kenta Takahashi; Yasuko Orba; Taichi Kimura; Lei Wang; Shinji Kohsaka; Masumi Tsuda; Mishie Tanino; Hiroshi Nishihara; Kazuo Nagashima; Hirofumi Sawa; Shinya Tanaka
  JAPANESE JOURNAL OF INFECTIOUS DISEASES, 66, 2, 126, 132, Mar. 2013, [Peer-reviewed]
  English, Scientific journal
• Role of JC virus agnoprotein in virion formation
  Tadaki Suzuki; Shingo Semba; Yuji Sunden; Yasuko Orba; Shintaro Kobayashi; Kazuo Nagashima; Takashi Kimura; Hideki Hasegawa; Hirofumi Sawa
  MICROBIOLOGY AND IMMUNOLOGY, 56, 9, 639, 646, Sep. 2012, [Peer-reviewed]
  English, Scientific journal
• Accumulation of ubiquitinated proteins is related to West Nile virus-induced neuronal apoptosis
  Shintaro Kobayashi; Yasuko Orba; Hiroki Yamaguchi; Takashi Kimura; Hirofumi Sawa
  NEUROPATHOLOGY, 32, 4, 398, 405, Aug. 2012, [Peer-reviewed]
  English, Scientific journal
• The sphingosine-1-phosphate transporter Spns2 expressed on endothelial cells regulates lymphocyte trafficking in mice
  Shigetomo Fukuhara; Szandor Simmons; Shunsuke Kawamura; Asuka Inoue; Yasuko Orba; Takeshi Tokudome; Yuji Sunden; Yuji Arai; Kazumasa Moriwaki; Junji Ishida; Akiyoshi Uemura; Hiroshi Kiyonari; Takaya Abe; Akiyoshi Fukamizu; Masanori Hirashima; Hirofumi Sawa; Junken Aoki; Masaru Ishii; Naoki Mochizuki
  JOURNAL OF CLINICAL INVESTIGATION, 122, 4, 1416, 1426, Apr. 2012, [Peer-reviewed]
  English, Scientific journal
• Detection and characterization of a novel polyomavirus in wild rodents
  Yasuko Orba; Shintaro Kobayashi; Ichiro Nakamura; Akihiro Ishii; Bernard M. Hang'ombe; Aaron S. Mweene; Yuka Thomas; Takashi Kimura; Hirofumi Sawa
  JOURNAL OF GENERAL VIROLOGY, 92, 4, 789, 795, Apr. 2011, [Peer-reviewed]
  English, Scientific journal
• Natalizumab Has No Direct Biological Effect on JC Virus Infectivity in Permissive Human Neural Cell Lines
  Tadaki Suzuki; Satoko Yamanouchi; Yuji Sunden; Yasuko Orba; Takashi Kimura; Hirofumi Sawa
  JOURNAL OF MEDICAL VIROLOGY, 82, 7, 1229, 1235, Jul. 2010, [Peer-reviewed]
  English, Scientific journal
• The Human Polyoma JC Virus Agnoprotein Acts as a Viroporin
  Tadaki Suzuki; Yasuko Orba; Yuki Okada; Yuji Sunden; Takashi Kimura; Shinya Tanaka; Kazuo Nagashima; William W. Hall; Hirofumi Sawa
  PLOS PATHOGENS, 6, 3, e1000801, Mar. 2010, [Peer-reviewed]
  English, Scientific journal
• Large T Antigen Promotes JC Virus Replication in G(2)-arrested Cells by Inducing ATM- and ATR-mediated G(2) Checkpoint Signaling
  Yasuko Orba; Tadaki Suzuki; Yoshinori Makino; Kanako Kubota; Shinya Tanaka; Takashi Kimura; Hirofumi Sawa
  JOURNAL OF BIOLOGICAL CHEMISTRY, 285, 2, 1544, 1554, Jan. 2010, [Peer-reviewed]
  English, Scientific journal
• Inhibition of the SDF-1 alpha-CXCR4 axis by the CXCR4 antagonist AMD3100 suppresses the migration of cultured cells from ATL patients and murine lymphoblastoid cells from HTLV-I Tax transgenic mice
  Akira Kawaguchi; Yasuko Orba; Takashi Kimura; Hidekatsu Iha; Masao Ogata; Takahiro Tsuji; Akira Ainai; Tetsutaro Sata; Takashi Okamoto; William W. Hall; Hirofumi Sawa; Hideki Hasegawa
  BLOOD, 114, 14, 2961, 2968, Oct. 2009, [Peer-reviewed]
  English, Scientific journal
• An siRNA against JC virus (JCV) agnoprotein inhibits JCV infection in JCV-producing cells inoculated in nude mice
  Tomoko Matoba; Yasuko Orba; Tadaki Suzuki; Yoshinori Makino; Hideo Shichinohe; Satoshi Kuroda; Takahiro Ochiya; Hiroshi Itoh; Shinya Tanaka; Kazuo Nagashima; Hirofumi Sawa
  NEUROPATHOLOGY, 28, 3, 286, 294, Jun. 2008, [Peer-reviewed]
  English, Scientific journal
• Progressive multifocal leukoencephalopathy and CD4+T-lymphocytopenia in a patient with Sjogren syndrome
  Yuichi Hayashi; Akio Kimura; Shimon Kato; Akihiro Koumura; Takeo Sakurai; Yuji Tanaka; Isao Hozumi; Yuji Sunden; Yasuko Orba; Hirofumi Sawa; Hitoshi Takahashi; Takashi Inuzuka
  JOURNAL OF THE NEUROLOGICAL SCIENCES, 268, 1-2, 195, 198, May 2008, [Peer-reviewed]
  English, Scientific journal
• Forkhead transcription factor FoxO1 in adipose tissue regulates energy storage and expenditure
  Jun Nakae; Yongheng Cao; Miyo Oki; Yasuko Orba; Hirofumi Sawa; Hiroshi Kiyonari; Kristy Iskandar; Koji Suga; Marc Lombes; Yoshitake Hayashi
  DIABETES, 57, 3, 563, 576, Mar. 2008, [Peer-reviewed]
  English, Scientific journal
• Pharmacological cdk inhibitor R-Roscovitine suppresses JC virus proliferation
  Yasuko Orba; Yuji Sunden; Tadaki Suzuki; Kazuo Nagashima; Takashi Kimura; Shinya Tanaka; Hirofumi Sawa
  VIROLOGY, 370, 1, 173, 183, Jan. 2008, [Peer-reviewed]
  English, Scientific journal
• Recent research on the JC virus
  Hirofumi Sawa; Tadaki Suzuki; Yasuko Orba; Yuji Sunden; Kazuo Nagashima
  Brain and Nerve, 59, 2, 101, 108, 2, Feb. 2007, [Peer-reviewed]
  Japanese, Scientific journal
• [Recent research on the JC virus].
  Sawa H; Suzuki T; Orba Y; Sunden Y; Nagashima K
  Brain and nerve = Shinkei kenkyu no shinpo, 59, 101, 108, 2, Feb. 2007, [Peer-reviewed]
• Identification of DDX1 as a JC virus transcriptional control region-binding protein
  Yuji Sunden; Shingo Semba; Tadaki Suzuki; Yuki Okada; Yasuko Orba; Kazuo Nagashima; Takashi Umemura; Hirofumi Sawa
  MICROBIOLOGY AND IMMUNOLOGY, 51, 3, 327, 337, 2007, [Peer-reviewed]
  English, Scientific journal
• DDX1 promotes proliferation of the JC virus through transactivation of its promoter
  Yuji Sunden; Shingo Semba; Tadaki Suzuki; Yuki Okada; Yasuko Orba; Kazuo Nagashima; Takashi Umemura; Hirofumi Sawa
  MICROBIOLOGY AND IMMUNOLOGY, 51, 3, 339, 347, 2007, [Peer-reviewed]
  English, Scientific journal
• A ubiquitin ligase HRD1 promotes the degradation of Pael receptor, a substrate of Parkin
  Tomohiro Omura; Masayuki Kaneko; Yasunobu Okuma; Yasuko Orba; Kazuo Nagashima; Ryosuke Takahashi; Noboru Fujitani; Satoshi Matsumura; Akihisa Hata; Kyoko Kubota; Karin Murahashi; Takashi Uehara; Yasuyuki Nomura
  JOURNAL OF NEUROCHEMISTRY, 99, 6, 1456, 1469, Dec. 2006, [Peer-reviewed]
  English, Scientific journal
• Characterization and application of polyclonal antibodies that specifically recognize JC virus large T antigen
  Y Sunden; T Suzuki; Y Orba; T Umemura; M Asamoto; K Nagashima; S Tanaka; H Sawa
  ACTA NEUROPATHOLOGICA, 111, 4, 379, 387, Apr. 2006, [Peer-reviewed]
  English, Scientific journal
• Thymus-derived leukemia-lymphoma in mice transgenic for the Tax gene of human T-lymphotropic virus type I
  H Hasegawa; H Sawa; MJ Lewis; Y Orba; N Sheehy; Y Yamamoto; T Ichinohe; Y Tsunetsugu-Yokota; H Katano; H Takahashi; J Matsuda; T Sata; T Kurata; K Nagashima; WW Hall
  NATURE MEDICINE, 12, 4, 466, 472, Apr. 2006, [Peer-reviewed]
  English, Scientific journal
• Identification of FEZ1 as a protein that interacts with JC virus agnoprotein and microtubules - Role of agnoprotein-induced dissociation of FEZ1 from microtubules in viral propagation
  T Suzuki; Y Okada; S Semba; Y Orba; S Yamanouchi; S Endo; S Tanaka; T Fujita; S Kuroda; K Nagashima; H Sawa
  JOURNAL OF BIOLOGICAL CHEMISTRY, 280, 26, 24948, 24956, Jul. 2005, [Peer-reviewed]
  English, Scientific journal
• Dissociation of heterochromatin protein 1 from lamin B receptor induced by human polyomavirus agnoprotein: role in nuclear egress of viral particles
  Y Okada; T Suzuki; Y Sunden; Y Orba; S Kose; N Imamoto; H Takahashi; S Tanaka; WW Hall; K Nagashima; H Sawa
  EMBO REPORTS, 6, 5, 452, 457, May 2005, [Peer-reviewed]
  English, Scientific journal
• Establishment of an immunoscreening system using recombinant VP1 protein for the isolation of a monoclonal antibody that blocks JC virus infection
  C Henmi; H Sawa; H Iwata; Y Orba; S Tanaka; K Nagashima
  BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 327, 1, 242, 251, Feb. 2005, [Peer-reviewed]
  English, Scientific journal
• Triglyceride accumulation and altered composition of triglyceride-associated fatty acids in the skin of tenascin-X-deficient mice
  K Matsumoto; T Sato; S Oka; Y Orba; H Sawa; K Kabayama; J Inokuchi; H Ariga
  GENES TO CELLS, 9, 8, 737, 748, Aug. 2004, [Peer-reviewed]
  English, Scientific journal
• Inhibition of virus production in JC virus-infected cells by postinfection RNA interference
  Y Orba; H Sawa; H Iwata; S Tanaka; K Nagashima
  JOURNAL OF VIROLOGY, 78, 13, 7270, 7273, Jul. 2004, [Peer-reviewed]
  English, Scientific journal
• Induction of matrix metalloproteinase-2 by tenascin-X deficiency is mediated through the c-Jun N-terminal kinase and protein tyrosine kinase phosphorylation pathway
  K Matsumoto; T Minamitani; Y Orba; M Sato; H Sawa; H Ariga
  EXPERIMENTAL CELL RESEARCH, 297, 2, 404, 414, Jul. 2004, [Peer-reviewed]
  English, Scientific journal
• Anti-Hu paraneoplastic syndrome presenting with brainstem-cerebellar symptoms and Lambert-Eaton myasthenic syndrome
  T Nagashima; Y Mizutani; H Kawahara; S Maguchi; Y Terayama; T Shinohara; Y Orba; T Chuma; Y Mano; T Itoh; H Sawa; K Sakai; M Motomura; K Nagashima
  NEUROPATHOLOGY, 23, 3, 230, 238, Sep. 2003, [Peer-reviewed]
  English, Scientific journal
• Application of laser capture microdissection to cytologic specimens for the detection of immunoglobulin heavy chain gene rearrangement in patients with malignant lymphoma
  Y Orba; S Tanaka; H Nishihara; N Kawamura; T Itoh; M Shimizu; H Sawa; K Nagashima
  CANCER CYTOPATHOLOGY, 99, 4, 198, 204, Aug. 2003, [Peer-reviewed]
  English, Scientific journal
• [JC virus infection (progressive multifocal leukoencephalopathy)].
  Nakagawa T; Orba Y; Nagashima K
  Nihon rinsho. Japanese journal of clinical medicine, 61 Suppl 2, 122, 127, Feb. 2003, [Peer-reviewed]
• JC virus agnoprotein colocalizes with tubulin
  S Endo; Y Okada; Y Orba; H Nishibara; S Tanaka; K Nagashima; H Sawa
  JOURNAL OF NEUROVIROLOGY, 9, 10, 14, 2003, [Peer-reviewed]
  English, Scientific journal
• Clinicopathological studies of peripheral neuropathy in Churg-Strauss syndrome
  T Nagashima; BZ Cao; N Takeuchi; T Chuma; Y Mano; M Fujimoto; M Nunomura; T Oshikiri; K Miyazaki; M Dohke; N Kashimura; T Shinohara; Y Orba; S Ishizawa; K Nagashima
  NEUROPATHOLOGY, 22, 4, 299, 307, Dec. 2002, [Peer-reviewed]
  English, Scientific journal
• Distribution of extracellular matrix tenascin-X in sciatic nerves
  K Matsumoto; H Sawa; M Sato; Y Orba; K Nagashima; H Ariga
  ACTA NEUROPATHOLOGICA, 104, 5, 448, 454, Nov. 2002, [Peer-reviewed]
  English, Scientific journal
• Expression of JC virus agnoprotein in progressive multifocal leukoencephalopathy brain
  Y Okada; H Sawa; S Endo; Y Orba; T Umemura; H Nishihara; AC Stan; S Tanaka; H Takahashi; K Nagashima
  ACTA NEUROPATHOLOGICA, 104, 2, 130, 136, Aug. 2002, [Peer-reviewed]
  English, Scientific journal
• Chronic demyelinating polyneuropathy in graft-versus-host disease following allogeneic bone marrow transplantation
  T Nagashima; F Sato; T Chuma; Y Mano; Sasaki, I; M Mori; T Higa; N Masauji; M Kasai; Y Orba; T Shinohara; K Nagashima
  NEUROPATHOLOGY, 22, 1, 1, 8, Mar. 2002, [Peer-reviewed]
  English, Scientific journal
• Immunohistochemical detection of hepatocellular carcinoma in the setting of ongoing necrosis after radiofrequency ablation
  T Itoh; Y Orba; H Takei; Y Ishida; M Saitoh; H Nakamura; T Meguro; S Horita; M Fujita; K Nagashima
  MODERN PATHOLOGY, 15, 2, 110, 115, Feb. 2002, [Peer-reviewed]
  English, Scientific journal
• Molecular neuropathology of JC virus
  Yasuko Orba; Hirofumi Sawa; Kazuo Nagashima
  Brain and Nerve, 54, 2, 101, 109, 2, 2002, [Peer-reviewed]
  Japanese

• Hunting zoonotic RNA viruses in wildlife
  大場靖子; 佐々木道仁; 澤洋文; 大場靖子; 佐々木道仁; 澤洋文, 実験医学, 43, 13, 2025
• 加齢に伴う血管内皮細胞SARS-CoV-2感染増強メカニズムの解析
  桜井優弥; 桜井優弥; 間石奈湖; 藤岡容一朗; 大場雄介; 佐々木道仁; 佐々木道仁; 大場靖子; 大場靖子; 大場靖子; 樋田泰浩; 城戸幹太; 澤洋文; 澤洋文; 澤洋文; 澤洋文; 樋田京子, 日本歯科医師会雑誌, 78, 4, 2025
• 人獣共通感染症対策に資する活動
  澤洋文; 澤洋文; 澤洋文; 田畑耕史郎; 板倉友香里; 梶原将大; 林田京子; 松野啓太; 直亨則; 大場靖子; 大場靖子; 佐々木道仁, 日本化学療法学会雑誌, 72, 5, 2024
• コウモリ由来オルソレオウイルスの病原性発現機構の解明
  播磨勇人; 佐々木道仁; 有泉拓馬; 川口虹穂; 小林広子; KITTIYA Intaruck; 小林剛; 川岸崇裕; 梶原将大; 大場靖子; 伊藤直人; 石原加奈子; HANG’OMBE Bernard; 澤洋文, 日本獣医学会学術集会講演要旨集, 167th, 2024
• 新規オルソナイロウイルス(エゾウイルス)感染症の病理学的解析および治療薬探索を目的とした致死感染マウスモデルの確立
  有泉拓馬; 松野啓太; 松野啓太; 田畑耕史郎; 板倉友香里; 小林広子; 佐々木道仁; 澤洋文; 大場靖子; 澤洋文; 大場靖子, 日本獣医学会学術集会講演要旨集, 167th, 2024
• Nsp5遺伝子に変異を有するSARS-CoV-2の抗ウイルス薬感受性とウイルス性状の解析
  佐々木道仁; 佐々木道仁; 板倉友香里; 田畑耕史郎; 日下部伸治; 日下部伸治; 小西慧; 小西慧; 杉達紀; 山岸潤也; 山岸潤也; 川口虹穂; 藤井英里; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦; 澤洋文; 澤洋文; 澤洋文; 大場靖子; 大場靖子; 大場靖子, 日本分子生物学会年会プログラム・要旨集(Web), 47th, 2024
• 日本のマダニから検出されたBeiji nairovirusおよび近縁ウイルスの解析
  岸本麻衣; 岸本麻衣; 岸本麻衣; 板倉友香里; 板倉友香里; 田畑耕史郎; 田畑耕史郎; 駒込理佳; 山口宏樹; 中尾亮; 邱永晋; 澤洋文; 大場靖子; 松野啓太; 澤洋文; 大場靖子; 松野啓太, 日本獣医学会学術集会講演要旨集, 167th, 2024
• Epidemiological and genetical study of Hokkaido virus, a genotype of Orthohantavirus puumalaense, among Myodes rodents
  DUONG Thuy Thi Ngoc; DUONG Thuy Thi Ngoc; SASAKI Michihito; ORBA Yasuko; AOSHIMA Keisuke; THAMMAHAKIN Passawat; MAEZONO Keisuke; KOBAYASHI Shintaro; KARIWA Hiroaki, 日本獣医学会学術集会講演要旨集, 167th, 2024
• Molnupiravir代謝活性体のβ-D-N⁴-hydroxycitidine(NHC)はin vitroで抗狂犬病ウイルス活性を示す
  小西慧; 小西慧; 日下部伸治; 日下部伸治; 川口虹穂; 宍戸貴雄; 伊藤直人; 大場靖子; 大場靖子; 大場靖子; 澤洋文; 澤洋文; 澤洋文; 佐々木道仁; 佐々木道仁; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦, 日本分子生物学会年会プログラム・要旨集(Web), 47th, 2024
• ウエストナイルウイルスサブユニットワクチンにおけるAS03様アジュバントの免疫賦活効果の検証
  井上敦子; 井上敦子; 板倉友香里; 齋藤慎二; 前仲勝実; 前仲勝実; 前仲勝実; 前仲勝実; 前仲勝実; 澤洋文; 澤洋文; 澤洋文; 大場靖子; 大場靖子; 田畑耕史郎, 日本分子生物学会年会プログラム・要旨集(Web), 47th, 2024
• 空間的遺伝子発現解析により明らかとなる日本脳炎ウイルス脳内感染初期イベント
  大場靖子; 大場靖子; 大場靖子; 鹿島幸恵; 田畑耕史郎; 板倉友香里; 澤洋文; 澤洋文; 佐々木道仁; 佐々木道仁; 鈴木穣, 日本分子生物学会年会プログラム・要旨集(Web), 47th, 2024
• 狂犬病ウイルス街上毒のリバースジェネティクスシステムの確立とイメージング解析への応用
  川口虹穂; 板倉友香里; 伊藤直人; 澤洋文; 澤洋文; 大場靖子; 大場靖子; 佐々木道仁; 佐々木道仁, 日本分子生物学会年会プログラム・要旨集(Web), 47th, 2024
• Mechanism of IgA tetramer formation
  田畑耕史郎; 田畑耕史郎; 福原秀雄; 佐々木道仁; 板倉友香里; 板倉友香里; 齊藤慎二; 澤洋文; 澤洋文; 澤洋文; 鈴木忠樹; 大場靖子, 日本生体防御学会学術総会講演抄録集, 35th (Web), 2024
• Establishment of antiviral neutralizing antibody targeting intracellular process of viral replication
  板倉友香里; 板倉友香里; 田畑耕史郎; 田畑耕史郎; 大場靖子; 大場靖子; 澤洋文; 澤洋文; 澤洋文; 佐々木道仁; 佐々木道仁, 日本生体防御学会学術総会講演抄録集, 35th (Web), 2024
• Insect-specific flaviviruses interfere with the replication of mosquito-borne flaviviruses
  森健人; 田村友和; 田村友和; 田村友和; 鈴木理滋; 鈴木理滋; 鈴木紗織; 鈴木紗織; 大場靖子; 大場靖子; 大場靖子; 福原崇介; 福原崇介; 福原崇介; 福原崇介, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024
• Identification of novel virulence factors of nelson bay orthoreovirus by genetic comparison between wild-type strains
  播磨勇人; 佐々木道仁; 有泉拓馬; 川口虹穂; 小林広子; KITTIYA Intaruck; 小林剛; 川岸崇裕; 金井祐太; 直亨則; 邱永晋; 梶原将大; 大場靖子; 伊藤直人; 石原加奈子; HANGOMBE Bernard M; 澤洋文, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024
• Development of MPXV therapeutics
  佐藤彰彦; 佐藤彰彦; 佐藤彰彦; 日下部伸治; 日下部伸治; 小西慧; 小西慧; 宍戸貴雄; 佐々木道仁; 佐々木道仁; 大場靖子; 大場靖子; 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), 71st, 2024
• Spatial analysis of early events in Japanese encephalitis virus brain invasion
  大場靖子; 大場靖子; 大場靖子; 鹿島幸恵; 田畑耕史郎; 板倉友香里; 澤洋文; 澤洋文; 佐々木道仁; 佐々木道仁; 鈴木穣, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024
• Reverse genetics of the negative single-stranded RNA virus by using CPER method
  板倉友香里; 板倉友香里; 川口虹穂; 田畑耕史郎; 田畑耕史郎; 伊藤直人; 大場靖子; 大場靖子; 大場靖子; 澤洋文; 澤洋文; 澤洋文; 佐々木道仁; 佐々木道仁, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024
• Virological characterization of SFTSV mutant adapted to specific cell lines and its application to develop antiviral compounds
  日下部伸治; 宍戸貴雄; 松野啓太; 松野啓太; 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; 佐藤彰彦; 佐藤彰彦, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024
• Characterization of SARS-CoV-2 with mutations in viral nsp5 gene
  佐々木道仁; 佐々木道仁; 板倉友香里; 田畑耕史郎; 日下部伸治; 日下部伸治; 小西慧; 小西慧; 杉達紀; 山岸潤也; 山岸潤也; 小林広子; 有泉拓馬; 川口虹穂; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦; 澤洋文; 澤洋文; 澤洋文; 大場靖子; 大場靖子; 大場靖子, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024
• Genetic characterization of a pigeon-derived gammacoronavirus using public RNA-seq data
  小林広子; 岸本麻衣; 岸本麻衣; 岸本麻衣; 今井咲帆; 大場靖子; 大場靖子; 大場靖子; 大場靖子; 澤洋文; 澤洋文; 澤洋文; 堀江真行; 堀江真行; 堀江真行, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024
• Search for respiratory syncytial virus therapeutic targets through phenotypic screening from drug repurposing in pediatric drugs
  小笠原徳子; 小笠原徳子; 山本聡; 谷向由佳; 吉田有梨枝; 杉田征彦; 佐藤彰彦; 佐藤彰彦; 澤洋文; 大場靖子; 勝山彬; 勝山彬; 勝山彬; 市川聡; 市川聡; 市川聡; 吉田圭太朗; 高野賢一; 横田伸一, 日本ウイルス学会学術集会プログラム・予稿集(Web), 71st, 2024
• Structural analysis of SARS-CoV-2 using BSL3 cryo-EM
  福原秀雄; DOKAINISH Hisham; 喜多俊介; 田畑耕史郎; 田畑耕史郎; 高巣晃; HUISKONEN Juha; 安楽佑樹; 千田俊哉; 千田俊哉; 千田俊哉; STUART David; 佐々木道仁; 大場靖子; 鈴木定彦; 鈴木定彦; 澤洋文; 澤洋文; 前仲勝実; 前仲勝実; 前仲勝実, 日本生体防御学会学術総会講演抄録集, 35th (Web), 2024
• 【One Healthのために,獣医学上おさえたいウイルス感染症】蚊媒介性ウイルス感染症とOne Health
  大場 靖子, 臨床とウイルス, 51, 4, 168, 172, Oct. 2023
  日本臨床ウイルス学会, Japanese
• 【新型コロナウイルス感染症時代に忘れてはならないウイルス感染症】RNAウイルス フラビウイルス科/トガウイルス科
  江下 優樹; 大場 靖子, 臨床と微生物, 50, 4, 363, 368, Jul. 2023
  (株)近代出版, Japanese
• IgA抗体の自己重合反応による四量体形成とその応用
  田畑耕史郎; 田畑耕史郎; 田畑耕史郎; 安楽佑樹; 多賀祐喜; 佐々木道仁; 佐々木道仁; 板倉友香里; 板倉友香里; 後藤希代子; 長谷川秀樹; 前仲勝実; 前仲勝実; 澤洋文; 澤洋文; 澤洋文; 福原秀雄; 大場靖子; 大場靖子; 鈴木忠樹, 日本抗体学会学術大会プログラム・抄録集(Web), 2nd, 2023
• Generation of severe COVID-19 mouse model by treatment with Cyclophosphamide
  日下部伸治; 日下部伸治; 鳥羽晋輔; 鳥羽晋輔; 井澤政明; 丸山優樹; 丸山優樹; 上村健太朗; 上村健太朗; 上村健太朗; 宍戸貴雄; 佐々木道仁; 大場靖子; 澤洋文; 澤洋文; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦, 日本ウイルス学会学術集会プログラム・予稿集(Web), 70th, 2023
• Establishment of antiviral neutralizing antibody targeting intracellular process of viral replication
  板倉友香里; 板倉友香里; 田畑耕史郎; 田畑耕史郎; 大場靖子; 大場靖子; 大場靖子; 澤洋文; 澤洋文; 澤洋文; 佐々木道仁; 佐々木道仁, 日本ウイルス学会学術集会プログラム・予稿集(Web), 70th, 2023
• Mechanism of severe COVID-19 regulated by amino acid mutations in non-structural proteins
  岸本麻衣; 岸本麻衣; 鳥羽晋輔; 鳥羽晋輔; 上村健太朗; 上村健太朗; 上村健太朗; 丸山優樹; 丸山優樹; 日下部伸治; 日下部伸治; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦; 大場靖子; 澤洋文; 澤洋文; 佐々木道仁, 日本ウイルス学会学術集会プログラム・予稿集(Web), 70th, 2023
• Broad-spectrum search for influenza virus CEN (cap endonuclease) inhibitors
  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
• Pathogenicity analysis of two strains of West Nile virus newly isolated from mosquito and crocodile in Zambia
  小林広子; 田畑耕史郎; 有泉拓馬; CHAMBARO Herman; PHONGPHAEW Wallaya; SIMULUNDU Edgar; 佐々木道仁; 澤洋文; 澤洋文; 澤洋文; 大場靖子; 大場靖子; 大場靖子, 日本ウイルス学会学術集会プログラム・予稿集(Web), 70th, 2023
• Investigation of anti-Nairovirus compounds
  鳥羽晋輔; 鳥羽晋輔; 平野港; 垰田善之; 日下部伸治; 日下部伸治; 小西慧; 小西慧; 丸山優樹; 宍戸貴雄; 好井健太朗; 佐々木道仁; 大場靖子; CHANGULA Katendi; MUDENDA Hang’ombe Bernard; 澤洋文; 澤洋文; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦, 日本ウイルス学会学術集会プログラム・予稿集(Web), 70th, 2023
• Comparison of the viral infectivity between the fixed and street strains of Rabies virus in peripheral tissues
  川口虹穂; 板倉友香里; 井上智; 前田健; 伊藤直人; 澤洋文; 澤洋文; 大場靖子; 大場靖子; 佐々木道仁; 佐々木道仁, 日本分子生物学会年会プログラム・要旨集(Web), 46th, 2023
• Discovery history of novel oral drug S-217622 (ensitrelvir) for COVID-19
  加藤輝久; 登治謙; 川島生; 丸山優樹; 鳥羽晋輔; 鳥羽晋輔; 佐名木孝央; 宇納佑斗; 中原健二; 垰田善之; 上原彰太; 山本志保; 水垂亨; 笠松幸司; 幸木謙典; 中橋徳文; 須藤貴弘; 佐々木道仁; 大場靖子; 澤洋文; 澤洋文; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦; 佐藤剛章; 立花裕樹, 日本薬学会年会要旨集(Web), 143rd, 2023
• Temperature-dependent regulatory mechanism of infectivity on Insect-Specific Flavivirus
  小林広子; 田畑耕史郎; 佐々木道仁; 佐々木道仁; 澤洋文; 澤洋文; 澤洋文; 大場靖子; 大場靖子; 大場靖子, 日本分子生物学会年会プログラム・要旨集(Web), 46th, 2023
• Discovery of a broad-spectrum antiviral nucleoside analogue against positive-strand RNA viruses
  上村健太朗; 上村健太朗; 上村健太朗; 上村健太朗; 上村健太朗; 登治謙; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦; 鳥羽晋輔; 鳥羽晋輔; 日下部伸治; 日下部伸治; 佐々木道仁; 田畑耕史郎; 大場靖子; 澤洋文; 澤洋文; 松浦善治; 松浦善治; 松田彰; 前仲勝実; 前仲勝実; 前仲勝実, 日本薬学会年会要旨集(Web), 143rd, 2023
• 血管内皮細胞におけるSARS-CoV-2侵入機構の解析
  桜井優弥; 間石奈湖; 藤岡容一朗; 大場雄介; 武田遼; 佐々木道仁; 大場靖子; 樋田泰浩; 澤洋文; 澤洋文; 樋田京子, 日本血管生物医学会学術集会プログラム・抄録集, 31st (CD-ROM), 2023
• X-206 exhibits broad-spectrum and robust inhibition against infections by several drug-resistant isolates and variants of SARS-CoV-2.
  佐々木慈英; 岡部伊織; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦; 児玉耕太; 乙黒聡子; 佐々木道仁; 大場靖子; 澤洋文; 澤洋文; 前仲勝実; 柳雄介; 橋口隆生, 日本ウイルス学会学術集会プログラム・予稿集(Web), 70th, 2023
• Combination therapy with oral antiviral and anti-inflammatory treatments improves the delayed-therapeutic effect on COVID-19
  佐々木道仁; 杉達紀; 飯田俊; 平田雄一郎; 日下部伸治; 日下部伸治; 小西慧; 小西慧; 板倉友香里; 板倉友香里; 田畑耕史郎; 田畑耕史郎; 岸本麻衣; 岸本麻衣; 小林広子; 有泉拓馬; KITTIYA Intaruck; 登治謙; 鳥羽晋輔; 鳥羽晋輔; 佐藤彰彦; 佐藤彰彦; 佐藤彰彦; 松野啓太; 松野啓太; 山岸潤也; 山岸潤也; 鈴木忠樹; 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
• ザンビアのエジプトルーセットオオコウモリおよびマストミスから分離されたロタウイルスAの性状解析
  岸本麻衣; 岸本麻衣; 梶原将大; 高田礼人; 大園誠也; 鈴木忠樹; 伊藤直人; 大場靖子; 澤洋文; 澤洋文; 佐々木道仁, 日本獣医学会学術集会講演要旨集, 166th, 2023
• Genes involved in spread of SARS-CoV-2 into the lower respiratory tract may be under adaptive selection.
  高田光輔; 大場靖子; 木田裕里恵; 呉佳齊; 小野慎子; 小野慎子; 松浦善治; 松浦善治; 中川草; 澤洋文; 渡辺登喜子; 渡辺登喜子, 日本進化学会大会プログラム・講演要旨集(Web), 25th, 2023
• 3種のヤブカ体内において選択されたジカウイルスのゲノム変異評価
  下岡 誠; 三木 要; 江下 優樹; 大場 靖子; 澤 洋文; 村松 康和; 内田 玲麻, 日本獣医学会学術集会講演要旨集, 165回, [F1P, 06], Sep. 2022
  (公社)日本獣医学会, Japanese
• ザンビアのコウモリにおけるネルソンベイオルソレオウイルスの保有調査および分離ウイルスの病原性比較解析
  播磨 勇人; 佐々木 道仁; 大場 靖子; 邱 永晋; Changula Katendi; 梶原 将大; Simulundu Edgar; 谷口 怜; 高田 礼人; 西條 政幸; Hang'ombe Bernard M.; 澤 洋文, 日本獣医学会学術集会講演要旨集, 165回, [F1P, 09], Sep. 2022
  (公社)日本獣医学会, Japanese
• 5-Ethynylimidazole-4-carboxamide(EICA)ヌクレオチドプロドラッグの合成と抗デングウイルス活性
  日野谷 直人; 中村 元紀; 田良島 典子; 大場 靖子; 澤 洋文; 松田 彰; 前仲 勝実; 南川 典昭, 日本薬学会年会要旨集, 142年会, 28S, pm03S, Mar. 2022
  (公社)日本薬学会, Japanese
• 5-Ethynylimidazole-4-carboxamide(EICA)ヌクレオチドプロドラッグの合成と抗デングウイルス活性
  日野谷 直人; 中村 元紀; 田良島 典子; 大場 靖子; 澤 洋文; 松田 彰; 前仲 勝実; 南川 典昭, 日本薬学会年会要旨集, 142年会, 28S, pm03S, Mar. 2022
  (公社)日本薬学会, Japanese
• 蚊特異的フラビウイルスを用いた組換えフラビウイルスの作出と種特異的増殖機構の解析
  小林広子; 田畑耕史郎; 佐々木道仁; 澤洋文; 澤洋文; 大場靖子, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• 狂犬病ウイルスの弾丸状粒子形成におけるM蛋白質とESCRT-1構成因子TSG101の相互作用の意義
  板倉友香里; 田畑耕史郎; KITTIYA Intaruck; 岸本麻衣; 伊藤直人; 大場靖子; 澤洋文; 佐々木道仁, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• プラス鎖RNAウイルスに対して広域的に抗ウイルス活性を示すヌクレオシドアナログの探索
  上村健太朗; 上村健太朗; 上村健太朗; 上村健太朗; 登治謙; 佐藤彰彦; 佐藤彰彦; 鳥羽晋輔; 鳥羽晋輔; 日下部伸治; 日下部伸治; 佐々木道仁; 田畑耕史郎; 大場靖子; 澤洋文; 松浦善治; 松浦善治; 松田彰; 前仲勝実; 前仲勝実, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• 蚊特異的フラビウイルスの蚊成体における感染動態の解析
  田畑耕史郎; 江下優樹; 佐々木道仁; 澤洋文; 澤洋文; 大場靖子, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• ウイルス分離先行型スクリーニングにより分離された野生動物由来ロタウイルスAの性状解析
  岸本麻衣; 梶原将大; 高田礼人; 伊藤直人; 大場靖子; 澤洋文; 佐々木道仁, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• 北海道で発熱患者から分離された新規オルソナイロウイルス(エゾウイルス)のマウス感染モデルの作製および病態解析
  有泉拓馬; 大場靖子; 田畑耕史朗; 板倉友香里; 小林広子; 佐々木道仁; 松野啓太; 澤洋文, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• 継代飼育ヒトスジシマカに存在する昆虫特異的ウイルスの単離とアルボウイルス感染への影響
  大場靖子; 田畑耕史郎; 佐々木道仁; 澤洋文; 江下優樹, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• Comparative analysis of Vertebrate-Infected Flaviviruses and Insect-Specific Flaviviruses to explore the mechanisms of Flavivirus host tropism
  小林広子; 田畑耕史郎; 佐々木道仁; 澤洋文; 澤洋文; 澤洋文; 大場靖子, 日本分子生物学会年会プログラム・要旨集(Web), 45th, 2022
• Antiviral efficacy of Ensitrelvir, an oral antiviral agent for COVID-19, in hamsters infected with SARS-CoV-2
  佐々木道仁; 田畑耕史郎; 岸本麻衣; 板倉友香里; 小林広子; 有泉拓馬; 上村健太朗; 上村健太朗; 上村健太朗; 鳥羽晋輔; 鳥羽晋輔; 日下部伸治; 日下部伸治; 丸山優樹; 丸山優樹; 飯田俊; 中島典子; 鈴木忠樹; 吉田晋平; 登治謙; 佐名木孝央; 加藤輝久; 宍戸貴雄; HALL William; HALL William; 大場靖子; 佐藤彰彦; 佐藤彰彦; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 45th, 2022
• Pathological analyses of Yezo virus infection in a mouse model
  有泉拓馬; 大場靖子; 田畑耕史郎; 板倉友香里; 小林広子; 佐々木道仁; 松野啓太; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 45th, 2022
• Virus-host interaction in bullet-shaped virion formation of rabies virus
  板倉友香里; 田畑耕史郎; 齋藤健; KITTIYA Intaruck; 川口虹穂; 岸本麻衣; 伊藤直人; 高田礼人; 大場靖子; 澤洋文; 佐々木道仁, 日本分子生物学会年会プログラム・要旨集(Web), 45th, 2022
• Molecular mechanism of severe COVID-19 regulated by non-structural proteins of SARS-CoV-2
  岸本麻衣; 鳥羽晋輔; 鳥羽晋輔; 上村健太朗; 上村健太朗; 上村健太朗; 丸山優樹; 丸山優樹; 日下部伸治; 日下部伸治; 佐藤彰彦; 佐藤彰彦; 大場靖子; 澤洋文; 佐々木道仁, 日本分子生物学会年会プログラム・要旨集(Web), 45th, 2022
• コロナウイルスを用いた効率の良い薬剤escapeウイルス分離法の開発
  佐藤彰彦; 佐藤彰彦; 上村健太朗; 上村健太朗; 上村健太朗; 鳥羽晋輔; 鳥羽晋輔; 日下部伸治; 日下部伸治; 丸山優樹; 丸山優樹; 宍戸貴雄; 佐々木道仁; 大場靖子; 澤洋文, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• 分泌型IgA抗体の四量体形成分子機構の生化学的・構造生物学的解析
  田畑耕史郎; 田畑耕史郎; 福原秀雄; 福原秀雄; 安楽佑樹; 多賀祐喜; 佐々木道仁; 板倉友香里; 佐野芳; 上野朗; 相内章; 後藤希代子; 長谷川秀樹; 前仲勝実; 前仲勝実; 澤洋文; 澤洋文; 大場靖子; 鈴木忠樹, 日本ワクチン学会学術集会プログラム・抄録集, 26th, 2022
• SARS-CoV-2感染ハムスターにおけるCOVID-19経口治療薬S-217622の抗ウイルス効果
  佐々木道仁; 田畑耕史郎; 岸本麻衣; 板倉友香里; 小林広子; 有泉拓馬; 上村健太朗; 上村健太朗; 上村健太朗; 鳥羽晋輔; 鳥羽晋輔; 日下部伸治; 日下部伸治; 丸山優樹; 丸山優樹; 飯田俊; 中島典子; 鈴木忠樹; 吉田晋平; 登治謙; 佐名木孝央; 加藤輝久; 宍戸貴雄; HALL William; HALL William; 大場靖子; 佐藤彰彦; 佐藤彰彦; 澤洋文, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• 強力な中和抗体NT-108によるSARS-CoV-2スパイクタンパク質認識の構造的基盤
  安楽佑樹; 小野寺大志; 喜多俊介; 安達悠; 森山彩野; 佐藤彰彦; 佐藤彰彦; 野村尚生; 田所高志; 田所高志; 湯本航平; 湯本航平; 伊東詩織; 田聡; 福原秀雄; 福原秀雄; 佐々木道仁; 大場靖子; 志和希; 岩田奈織子; 永田典代; 鈴木干城; 佐々木慈英; 関塚剛史; 登内奎介; 福士秀悦; 里深博幸; 香月康宏; 孫琳; 押村光雄; 黒田誠; 鈴木忠樹; 澤洋文; 橋口隆生; 高橋宜聖; 前仲勝実, 日本ウイルス学会学術集会プログラム・予稿集(Web), 69th, 2022
• SARS-CoV-2感染マウスを用いた肺血管内皮細胞の重症化関連遺伝子の解析
  武田遼; 武田遼; 伊藤航; 手代木孝仁; 佐々木道仁; 大場靖子; 大場靖子; 間石奈湖; 松田彩; 佐藤彰彦; 鳥羽晋輔; 李智媛; 飯村忠浩; 樋田泰浩; 北川善政; 澤洋文; 澤洋文; 澤洋文; 樋田京子, 日本血管生物医学会学術集会プログラム・抄録集, 30th (CD-ROM), 1, 219, 219, 2022
  (一社)日本病理学会, Japanese
• S-217622, a Novel SARS-CoV-2 3C-Like Protease Inhibitor Clinical Candidate, As a Potential Oral Therapeutic Agent for COVID-19
  登治謙; 立花裕樹; 山津維子; 川島生; 丸山優樹; 鳥羽晋輔; 鳥羽晋輔; 宇納佑斗; 中原健二; 垰田善之; 上原彰太; 山本志保; 水垂亨; 笠松幸司; 幸木謙典; 中橋徳文; 須藤貴弘; 佐名木孝央; 上村健太朗; 上村健太朗; 佐藤剛章; 澤洋文; 大場靖子; 佐々木道仁; 佐藤彰彦; 佐藤彰彦; 加藤輝久, 日本ウイルス学会学術集会プログラム・予稿集(Web), 68th, 2021
• Construction of the screening system for development of anti-coronavirus drugs
  佐藤彰彦; 佐藤彰彦; 上村健太朗; 上村健太朗; 鳥羽晋輔; 鳥羽晋輔; 丸山優樹; 丸山優樹; 日下部伸治; 日下部伸治; 佐名木孝夫; 宍戸貴雄; 佐々木道仁; 大場靖子; 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), 68th, 2021
• Reactive analyses of anti-mosquito-specific flavivirus antibodies against vertebrate-infecting flaviviruses.
  田畑耕史郎; 板倉友香里; 岸本麻衣; 佐々木道仁; 澤洋文; 澤洋文; 大場靖子, 日本ウイルス学会学術集会プログラム・予稿集(Web), 68th, 2021
• The role of TSG101 in RABV life cycle
  板倉友香里; 伊藤直人; 大場靖子; 澤洋文; 澤洋文; 佐々木道仁, 日本ウイルス学会学術集会プログラム・予稿集(Web), 68th, 2021
• Interaction between SARS-CoV-2 and host type II transmembrane serine proteases (TTSP)
  岸本麻衣; 上村健太朗; 上村健太朗; 佐名木孝央; 佐名木孝央; 佐藤彰彦; 佐藤彰彦; 大場靖子; 澤洋文; 佐々木道仁, 日本ウイルス学会学術集会プログラム・予稿集(Web), 68th, 2021
• A Natural Compound in Edible Food with the Potential for Inhibition of Corona Virus Infection
  村井勇太; 村井勇太; MUTMAINAH; KOOLATH Sajeer; 臼杵靖剛; 湯山耕平; 中岡慎治; 佐々木道仁; 大場靖子; 佐藤彰彦; 佐藤彰彦; 五十嵐靖之; 澤洋文; 門出健次; 門出健次, 天然有機化合物討論会講演要旨集(Web), 63rd, 2021
• Characterization of a Nelson Bay orthoreovirus isolated from an Egyptian fruit bat in Zambia
  播磨勇人; 佐々木道仁; 大場靖子; 邱永晋; チャングラカテンディ; 梶原将大; シムルンドゥエドガー; 谷口怜; 高田礼人; 西條政幸; ハンゴンべバーナード; 澤洋文, 日本ウイルス学会学術集会プログラム・予稿集(Web), 68th, 2021
• Glycoprotein Analyses by Mass Spectrometry
  梶裕之; 富岡あづさ; 日尾野隆大; 日尾野隆大; 佐々木道仁; 大場靖子; 澤洋文; 久野敦, 質量分析総合討論会講演要旨集, 69th, 2021
• RIG-Iはヒト肺細胞においてシグナル伝達非依存的な抗SARS-CoV-2防御を誘導する
  山田大翔; 山田大翔; 佐藤精一; 佐藤精一; 外山雄貴; 外山雄貴; 大場靖子; 澤洋文; 山内肇; 佐々木道仁; 高岡晃教; 高岡晃教, 日本インターフェロン・サイトカイン学会学術集会抄録集, 85th (CD-ROM), 2021
• G蛋白質333位のアミノ酸残基が規定する狂犬病ウイルスの病原性発現機構の解析
  板倉友香里; 田畑耕史郎; 伊藤直人; 大場靖子; 澤洋文; 佐々木道仁, 日本獣医学会学術集会講演要旨集, 164th (CD-ROM), [FO, 8], 2021
  (公社)日本獣医学会, Japanese
• ボリビア多民族国サンタクルス県の牛におけるBabesiaおよびAnaplasmaの感染状況
  小方昌平; PEREIRA Juan Antonio; 松野啓太; 大場靖子; 澤洋文; 川森文彦; 野中成晃; 中尾亮, 日本獣医学会学術集会講演要旨集, 164th (CD-ROM), [CO, 40], 2021
  (公社)日本獣医学会, Japanese
• 北海道産ヤマトヤブカにおけるジカウイルス感受性の評価
  川江詩乃; 岩本夏奈; 江下優樹; 大場靖子; 澤洋文; 村松康和; 内田玲麻, 日本獣医学会学術集会講演要旨集, 164th (CD-ROM), [FO, 5], 2021
  (公社)日本獣医学会, Japanese
• ヤマトヤブカ体内におけるジカウイルスのQuasispecies評価
  三木要; 岩本夏奈; 江下優樹; 大場靖子; 澤洋文; 村松康和; 内田玲麻, 日本獣医学会学術集会講演要旨集, 164th (CD-ROM), [FO, 6], 2021
  (公社)日本獣医学会, Japanese
• ザンビアに生息する野生げっ歯類動物における脳心筋炎ウイルス(EMCV)の疫学調査
  岸本麻衣; HANG’OMBE Bernard M.; 大場靖子; 澤洋文; 佐々木道仁, 日本獣医学会学術集会講演要旨集, 164th (CD-ROM), [DVO, 38], 2021
  (公社)日本獣医学会, Japanese
• ザンビアのブタにおける消化管ウイルスの疫学調査
  播磨勇人; 梶原将大; 佐々木道仁; SIMULUNDU Edgar; BWALYA Eugene; 邱永晋; 奥谷公亮; 大場靖子; HANG’OMBE Bernard M; MWEENE Aaron S; 高田礼人; 澤洋文, 日本獣医学会学術集会講演要旨集, 164th (CD-ROM), [DVO, 33], 2021
  (公社)日本獣医学会, Japanese
• 質量分析とレクチンアレイを組み合わせたSARS-CoV-2スパイクタンパク質の高精度糖鎖構造解析
  日尾野隆大; 日尾野隆大; 富岡あづさ; 梶裕之; 佐々木道仁; 大場靖子; 澤洋文; 久野敦, 日本獣医学会学術集会講演要旨集, 164th (CD-ROM), [DVO, 47], 2021
  (公社)日本獣医学会, Japanese
• Characterization of a newly emerging orthonairovirus in Hokkaido, Japan
  有泉拓馬; 山口宏樹; 澤洋文; 大場靖子; 松野啓太, 日本分子生物学会年会プログラム・要旨集(Web), 44th, 2021
• Evaluation of cell-penetrating peptides for the production of intracellular invasive antiviral neutralizing antibody
  板倉友香里; 田畑耕史郎; 大場靖子; 澤洋文; 澤洋文; 佐々木道仁, 日本分子生物学会年会プログラム・要旨集(Web), 44th, 2021
• A mechanism of Ig variable region-independent oligomerization of secretory IgA antibody accelerated by polymeric Ig receptor.
  田畑耕史郎; 田畑耕史郎; 鈴木忠樹; 佐野芳; 齊藤慎二; 相内章; 佐々木道仁; 長谷川秀樹; 大場靖子; 澤洋文; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 44th, 2021
• SARS-CoV-2 variants are generated during propagation in Vero cells
  佐々木道仁; 上村健太朗; 上村健太朗; 鳥羽晋輔; 鳥羽晋輔; 佐藤彰彦; 佐藤彰彦; 板倉友香里; INTARUCK Kittiya; 岸本麻衣; 田畑耕史郎; CHAMBARO Herman; 大場靖子; 大場靖子; 澤洋文; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 44th, 2021
• Examination of the accumulation of TDP-43 observed in neuronal cells infected with West Nile virus
  小林進太郎; 好井健太朗; 松野啓太; 大場靖子; 澤洋文; 苅和宏明, 日本分子生物学会年会プログラム・要旨集(Web), 44th, 2021
• Investigation of an orthoreovirus isolated from a fruit bat in Zambia
  播磨勇人; 佐々木道仁; 大場靖子; QIU Yongjin; 梶原将大; 谷口怜; 高田礼人; 西條政幸; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 44th, 2021
• Identification of host type II transmembrane serine proteases which promote SARS-CoV-2 infection
  岸本麻衣; 上村健太朗; 上村健太朗; 佐名木孝央; 佐名木孝央; 佐藤彰彦; 佐藤彰彦; 大場靖子; 澤洋文; 佐々木道仁, 日本分子生物学会年会プログラム・要旨集(Web), 44th, 2021
• 食品成分およびその誘導体群によるコロナウイルス感染抑制効果
  村井勇太; 村井勇太; ムト マイナー; クーラス サジール; 臼杵靖剛; 湯山耕平; 中岡慎治; 佐々木道仁; 大場靖子; 佐藤彰彦; 佐藤彰彦; 五十嵐靖之; 澤洋文; 門出健次; 門出健次, 日本生化学会大会(Web), 94th, [P, 943], 2021
  (公社)日本生化学会, Japanese
• Bat- and arthropod-borne viruses raising potential public health concern.
  梶原将大; 大場靖子; 高田礼人, Pharma Medica, 39, 1, 2021
• [Arthropod-borne viruses (arboviruses)].
  Yasuko Orba; Hirofumi Sawa; Keita Matsuno, Uirusu, 70, 1, 3, 14, 2020, [Domestic magazines]
  "Arbovirus" is a term for a virus transmitted to mammals by hematophagous arthropods; arboviruses; replicate in both mammals and arthropods. Since the life cycle of arboviruses is highly dependent on arthropods, control of the arthropods (vectors) is generally considered important for the control of arbovirus infection. Various pathogens that cause diseases in the medical and veterinary fields are grouped into arboviruses with a history of their discoveries since the early 20th century. Furthermore, because of recent advances in sequencing technology, new arboviruses have been discovered one after another. Here we would like to overview the known arboviruses and their infections., Japanese
• In vitroにおける四量体分泌型IgA抗体形成機構の解明
  田畑耕史郎; 田畑耕史郎; 鈴木忠樹; 佐野芳; 齊藤慎二; 相内章; 大場靖子; 長谷川秀樹; 長谷川秀樹; 澤洋文; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 42nd, 2019
• インターフェロン受容体欠損マウスにおける狂犬病ウイルス病原性発現機構
  板倉友香里; 大場靖子; 澤洋文; 澤洋文; 佐々木道仁, 日本分子生物学会年会プログラム・要旨集(Web), 42nd, 2019
• ウエストナイルウイルス感染で起こるAMP-activated protein kinaseの分解およびオートファジーの抑制による神経病態形成への影響
  小林進太郎; 好井健太朗; PHONGPAEW Wallaya; 武藤芽未; 平野港; 大場靖子; 澤洋文; 苅和宏明, 日本分子生物学会年会プログラム・要旨集(Web), 42nd, 2019
• ロタウイルス感染を促進させる宿主プロテアーゼの同定
  佐々木道仁; 伊藤直人; 杉山誠; 大場靖子; 澤洋文; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 42nd, 2019
• ESCRTタンパク質HGSはチクングニアウイルスのコードするタンパク質と相互作用し,細胞内増殖の2過程を制御する
  鳥居志保; 鳥居志保; 大場靖子; 佐々木道仁; 和田雄治; HOBSON-PETERS Jody; HALL Roy A.; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 42nd, 2019
• 蚊のゲノムに内在する多様なフラビウイルスエレメント
  大場靖子; HANG’OMBE Bernard M.; MWEENE Aaron S.; 佐々木道仁; 江下優樹; 澤洋文; 澤洋文; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 42nd, 2019
• ESCRT蛋白質HGSはチクングニアウイルスと相互作用し,細胞内増殖2過程を制御する
  鳥居志保; 鳥居志保; 大場靖子; 佐々木道仁; 和田雄治; HOBSON-PETERS Jody; HALL Roy; 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), 67th, 2019
• 宿主タンパク質YB-1は狂犬病ウイルスmRNAに結合し遺伝子発現を促進する
  佐々木道仁; 伊藤直人; 杉山誠; GONZALEZ Gabriel; 伊藤公人; 伊藤公人; 大場靖子; 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), 67th, 2019
• 抗デングウイルス活性をもつ低分子化合物の探索
  佐藤彰彦; 佐藤彰彦; 登治謙; 鳥羽晋輔; 鳥羽晋輔; 上村健太朗; 上村健太朗; 吉田立; 宍戸貴雄; 大場靖子; 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), 67th, 2019
• 多種の蚊に内在するフラビウイルスエレメント
  大場靖子; HANG’OMBE Bernard M.; MWEENE Aaron S.; 佐々木道仁; 江下優樹; 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), 67th, 2019
• ザンビアのエジプトルーセットオオコウモリから検出されたマールブルグウイルスの遺伝学的解析
  梶原将大; CHANGULA Katendi; HANG’OMBE Bernard; 播磨勇人; 宮本洋子; 衛藤芳樹; 奥谷公亮; 磯野真央; 吉田玲子; 邱永晋; 森(梶原)亜紀奈; 大場靖子; 澤洋文; 澤洋文; 小川寛仁; SIMULUNDU Edgar; SQUARRE David; MUKONKA Victor; MWEENE Aaron; 高田礼人; 高田礼人, 日本ウイルス学会学術集会プログラム・予稿集(Web), 67th, 2019
• ザンビアのダニで発見した新規フラビウイルスはフラビウイルス属の中で独特な系統を形成するNgoye virusに遺伝的に密接に関係している
  播磨勇人; 鳥居志保; 邱永晋; 梶原将大; 衛藤芳樹; 大場靖子; 江下優樹; ハンゴンベ バーナード; 好井健太朗; シムンザ マーティン; 高田礼人; 澤洋文, 日本ウイルス学会学術集会プログラム・予稿集(Web), 67th, 2019
• 狂犬病ウイルスに対して強い抗ウイルス活性を示す低分子化合物の同定
  上村健太朗; 上村健太朗; 鳥羽晋輔; 鳥羽晋輔; 登治謙; 吉田立; 宍戸貴雄; 佐々木道仁; 伊藤直人; 大場靖子; 澤洋文; 澤洋文; 佐藤彰彦; 佐藤彰彦, 日本ウイルス学会学術集会プログラム・予稿集(Web), 67th, 2019
• LCMV感染マウスモデルを用いたCompound Xのin vivo PK/PD解析
  鳥羽晋輔; 鳥羽晋輔; 登治謙; 上村健太朗; 上村健太朗; 垰田善之; 佐藤健司; 吉田立; 宍戸貴雄; 大場靖子; 澤洋文; 澤洋文; 佐藤彰彦; 佐藤彰彦, 日本ウイルス学会学術集会プログラム・予稿集(Web), 67th, 2019
• マダニ中の多様なフレボウイルスは何者か?
  松野啓太; 松野啓太; 中尾亮; 草木迫浩大; 大場靖子; 澤洋文; 澤洋文, 日本獣医学会学術集会講演要旨集, 162nd, 2019
• ウエストナイルウイルス感染で起こるオートファジーの抑制と神経病態形成の関係
  小林 進太郎; 好井 健太朗; Wallaya Phongpaew; 武藤 芽未; 平野 港; 大場 靖子; 澤 洋文; 苅和 宏明, 日本獣医学会学術集会講演要旨集, 161回, 395, 395, Aug. 2018
  (公社)日本獣医学会, Japanese
• 日本産ヒトスジシマカのジカウイルス媒介能
  江下優樹; 江下優樹; 江下優樹; 江下優樹; 和田雄治; 林田京子; 山岸潤也; 杉本千尋; 佐々木道仁; 大場靖子; 澤洋文; 飛弾野真也; 神山長慶; 小林隆志; 高崎智彦; 加藤文博; 田島茂; 倉根一郎; LIM Chang Kweng, 衛生動物, 69, Suppl., 51, 51, Mar. 2018
  日本衛生動物学会, Japanese
• 狂犬病ウイルスの遺伝子発現に関与する宿主因子の同定と機能解析
  佐々木道仁; GONZALEZ Gabriel; 伊藤直人; 杉山誠; 伊藤公人; 伊藤公人; 大場靖子; 澤洋文; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 41st, 2018
• チクングニアウイルスの増殖を制御する宿主因子の同定と機能解析
  鳥居志保; 和田雄二; 佐々木道仁; HOBSON-PETERS Jody; HALL Roy A.; 大場靖子; 澤洋文; 澤洋文; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 41st, 2018
• ウエストナイルウイルスのカプシドタンパク質によるオートファジーの抑制による変性タンパク質の蓄積と神経病態形成についての解析
  小林 進太郎; 好井 健太朗; Phonqphaew Wallaya; 武藤 芽未; 平野 港; 大場 靖子; 澤 洋文; 苅和 宏明, 生命科学系学会合同年次大会, 2017年度, [3P, 1110], Dec. 2017
  生命科学系学会合同年次大会運営事務局, Japanese
• ウイルス研究の最前線 2.ザンビア,インドネシアにおける野生動物が保有するDNAウイルスの探索
  澤洋文; 澤洋文; 佐々木道仁; 大場靖子, ウイルス, 67, 2, 151‐160, 160, Dec. 2017
  日本ウイルス学会, Japanese
• In vitroでの狂犬病ウイルスに対する5-エチニル-1-リボフラノシルイミダゾール-4-カルボキサミド(EICAR)の抗ウイルス活性に関する検討(Examination of antiviral activity of 5-ethynyl-1-ribofuranosylimidazole-4-carboxamide(EICAR) against rabies virus in vitro)
  Anindita Paulina Duhita; 佐々木 道仁; 伊藤 直人; 杉山 誠; 南川 典昭; 周東 智; 乙黒 聡子; 市川 聡; 松田 彰; 前仲 勝実; 大場 靖子; 澤 洋文, 日本獣医学会学術集会講演要旨集, 160回, 390, 390, Aug. 2017
  (公社)日本獣医学会, English
• Examination of antiviral activity of 5-ethynyl-1-ribofuranosylimidazole-4-carboxamide(EICAR) against rabies virus in vitro(和訳中)
  Anindita Paulina Duhita; 佐々木 道仁; 伊藤 直人; 杉山 誠; 南川 典昭; 周東 智; 乙黒 聡子; 市川 聡; 松田 彰; 前仲 勝実; 大場 靖子; 澤 洋文, 日本獣医学会学術集会講演要旨集, 160回, 390, 390, Aug. 2017
  (公社)日本獣医学会, English
• 【One Healthの視点からみた感染症の現状と対策】 野生動物が保有するウイルスを対象とする研究
  佐々木 道仁; 大場 靖子; 澤 洋文, 最新医学, 72, 4, 508, 512, Apr. 2017
  (株)最新医学社, Japanese
• これからの感染症とその対策―国際化社会および2020年,東京五輪への対応 診療科別のトピックス 蚊媒介性ウイルス
  江下優樹; SRISAWAT Raweewan; RUNTUWENE Lucky Ronald; 飛弾野真也; 林田京子; 大場靖子, 臨床と微生物, 43, 6, 701‐707, 707, Nov. 2016
  (株)近代出版, Japanese
• ウエストナイルウイルスの粒子放出過程におけるRab8bタンパク質の役割
  小林 進太郎; 鈴木 忠樹; 川口 晶; Phongphaew Wallaya; 好井 健太朗; 苅和 宏明; 大場 靖子; 澤 洋文, 日本獣医学会学術集会講演要旨集, 159回, 409, 409, Aug. 2016
  (公社)日本獣医学会, Japanese
• ヘパラン硫酸を介した狂犬病ウイルスの細胞吸着機構の解析
  佐々木道仁; アニンディタ パウリナ; 伊藤直人; 杉山誠; 福原秀雄; 尾瀬農之; 前仲勝実; 大場靖子; 澤洋文; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 39th, ROMBUNNO.2P‐0314 (WEB ONLY), 2016
  Japanese
• ウエストナイルウイルス感染による変性タンパク質蓄積機構の解析
  小林 進太郎; Phongphaew Wallaya; 好井 健太朗; 平野 港; 武藤 芽未; 大場 靖子; 澤 洋文; 苅和 宏明, 日本生化学会大会・日本分子生物学会年会合同大会講演要旨集, 88回・38回, [1P1135], [1P1135], Dec. 2015
  (公社)日本生化学会, English
• プリオン病及び遅発性ウイルス感染症の分子病態解明・治療法開発に関する研究 進行性多巣性白質脳症(PML)におけるJCウイルスタンパク質の発現様式の免疫組織学的検討
  澤洋文; 鈴木忠樹; 大場靖子, プリオン病及び遅発性ウイルス感染症の分子病態解明・治療法開発に関する研究 平成26年度 委託業務成果報告書, 67, 70, 2015
  Japanese
• ポリオーマウイルスの疫学研究と基礎研究
  澤 洋文; 小林 進太郎; 鈴木 忠樹; 大場 靖子, ウイルス, 64, 1, 25, 34, Jun. 2014
  日本ウイルス学会, Japanese
• Involvement of EphA2-mediated tyrosine phosphorylation of Shp2 in Shp2-regulated activation of extracellular signal-regulated kinase
  K. Miura; Y. Wakayama; M. Tanino; Y. Orba; H. Sawa; M. Hatakeyama; S. Tanaka; H. Sabe; N. Mochizuki, Oncogene, 32, 45, 5292, 5301, 07 Nov. 2013
  English
• 進行性多巣性白質脳症(PML)の病態解明への分子アプローチ
  鈴木忠樹; 澤洋文; 大場靖子; 片野晴隆; 佐藤由子; 長谷川秀樹; 長嶋和郎, 日本病理学会会誌, 102, 1, 207, 26 Apr. 2013
  Japanese
• 金ナノ粒子ワクチンによる形状依存的なサイトカイン産生及び抗体誘導の評価
  松永達也; 新倉謙一; 鈴木忠樹; 小林進太郎; 山口宏樹; 大場靖子; 梶野喜一; 二宮孝文; 澤洋文; 居城邦治, 日本化学会講演予稿集, 93rd, 3, 2013
• 金ナノ粒子ワクチンのin vivo及びin vitroにおける形状依存的ワクチン活性の評価
  松永達也; 新倉謙一; 鈴木忠樹; 小林進太郎; 山口宏樹; 大場靖子; 川口晶; 長谷川; 梶野喜一; 二宮孝文; 澤洋文; 居城邦治, 高分子学会北海道支部研究発表会講演要旨集, 47th, 2013
• In vivoおよびin vitroにおける金ナノ粒子ワクチンの形状依存的な活性
  新倉謙一; 松永達也; 鈴木忠樹; 小林進太郎; 小林進太郎; 山口宏樹; 大場靖子; 川口晶; 長谷川秀樹; 梶野喜一; 二宮孝文; 澤洋文; 居城邦治, バイオ・高分子シンポジウム講演要旨集, 23rd, 2013
• ナノ粒子ワクチンのワクチン活性に及ぼす粒子形状およびサイズ効果
  新倉謙一; 松永達也; 鈴木忠樹; 小林進太郎; 山口宏樹; 大場靖子; 川口晶; 長谷川秀樹; 梶野喜一; 二宮孝文; 澤洋文; 居城邦治, 高分子学会予稿集(CD-ROM), 62, 2, 2013
• オートファジーはウエストナイルウイルスの増殖を抑制する
  小林進太郎; 大場靖子; 山口宏樹; 佐々木道仁; 長谷部理絵; 木村享史; 澤洋文, 日本ウイルス学会学術集会プログラム・抄録集, 61st, 2013
• 霊長類動物からの新規ポリオーマウイルスの検出
  山口宏樹; 小林進太郎; 石井秋宏; 小川寛人; 木村享史; 澤洋文; 大場靖子, 日本分子生物学会年会プログラム・要旨集(Web), 36th, 2013
• ザンビアの野生動物におけるオルソポックスウイルス感染の疫学調査
  大場靖子; 石井秋宏; トーマス 由佳; 小川寛人; 中村一郎; 木村享史; 森川茂; 西條政幸; 澤洋文, 日本ウイルス学会学術集会プログラム・抄録集, 60th, 443, 31 Oct. 2012
  Japanese
• ウエストナイルウイルス感染神経細胞におけるユビキチン化タンパク質の蓄積
  小林進太郎; 小林進太郎; 大場靖子; 山口宏樹; 木村享史; 澤洋文; 澤洋文, 日本ウイルス学会学術集会プログラム・抄録集, 60th, 2012
• 霊長類動物におけるパラインフルエンザウイルスの疫学調査
  佐々木道仁; 石井秋宏; 大場靖子; トーマス由佳; 小川寛人; 中村一郎; 木村享史; 澤洋文; 澤洋文, 日本ウイルス学会学術集会プログラム・抄録集, 60th, 2012
• ザンビアにおける霊長類動物からの新規ポリオーマウイルスの検出
  山口宏樹; 山口宏樹; 大場靖子; 小林進太郎; 小林進太郎; 石井秋宏; 小川寛人; 中村一郎; THOMAS由佳; 木村享史; 澤洋文; 澤洋文, 日本ウイルス学会学術集会プログラム・抄録集, 60th, 2012
• 霊長類動物におけるパラインフルエンザウイルス3型の疫学調査
  佐々木道仁; 石井秋宏; 大場靖子; THOMAS Yuka; 小川寛人; 中村一郎; 木村享史; 澤洋文; 澤洋文, 日本分子生物学会年会プログラム・要旨集(Web), 35th, 2012
• ザンビア共和国の霊長類動物におけるポリオーマウイルスの疫学調査
  山口宏樹; 小林進太郎; 大場靖子; 石井秋宏; 小川寛人; Thomas由佳; 木村享史; 澤洋文, 日本獣医学会学術集会講演要旨集, 152nd, 2011
• 野生齧歯動物からの新規ポリオーマウイルスの検出
  大場靖子; 小林進太郎; 中村一郎; 石井秋宏; 木村享史; 澤洋文, 日本ウイルス学会学術集会プログラム・抄録集, 58th, 445, 15 Oct. 2010
  Japanese
• Disruption of Intracellular Vesicular Trafficking by Agnoprotein is Essential for Viroporin Activity and JC Virus Replication
  Tadaki Suzuki; Yasuko Orba; Yoshinori Malcino; Yuki Okada; Yuji Sunden; Takashi Kimura; Hideki Hasegawa; Tetsutaro Sata; William W. Hall; Hirofumi Sawa, JOURNAL OF NEUROVIROLOGY, 16, 84, 84, Oct. 2010
  English, Summary international conference
• Large T antigen promotes JC virus replication in G2 arrest by inducing G2 checkpoint signaling
  Yasuko Orba; Tadaki Suzuki; Takashi Kimura; Hirofumi Sawa, JOURNAL OF NEUROVIROLOGY, 16, 64, 65, Oct. 2010
  English, Summary international conference
• Inhibition of the SDF-1F?-CXCR4 Axis by the CXCR4 Antagonist AMD3100 Suppresses the Migration of Human ATLL Cells and Murine Lymphoblastoid Cells from HTLV-I Tax Transgenic Mice
  A. Kawaguichi; Hidekatsu, I; H. Hasegawa; H. Sawa; M. Ogata; T. Tsuji; T. Kimura; T. Sata; W. W. Hall; Y. Orba, AIDS RESEARCH AND HUMAN RETROVIRUSES, 25, 11, 1212, 1212, Nov. 2009
  English, Summary international conference
• CXCR4アンタゴニスト(AMD3100)を用いたSCIDマウスにおけるTaxトランスジェニックマウス由来腫瘍細胞の組織浸潤抑制
  川口晶; 辻隆裕; 大場靖子; 木村享史; 相内章; 伊波英克; 緒方正男; 佐多徹太郎; 澤洋文; 長谷川秀樹, 日本ウイルス学会学術集会プログラム・抄録集, 57th, 445, 01 Oct. 2009
  Japanese
• ラット神経由来CSM14.1細胞株における日本脳炎ウイルス感受性
  奥村恵; 木村享史; 大場靖子; 大場靖子; 澤洋文; 澤洋文, 日本ウイルス学会学術集会プログラム・抄録集, 57th, 2009
• Viroporin activity of JCV agnoprotein
  Tadaki Suzuki; Yasuko Orba; Yuji Sunden; Takashi Kimura; Hirofumi Sawa, JOURNAL OF NEUROVIROLOGY, 15, 96, 97, 2009
  English, Summary international conference
• ヒトATL細胞及びTaxトランスジェニックマウス由来腫瘍細胞のSDF‐1αに対する走化性
  川口晶; 大場靖子; 木村享史; 伊波英克; 緒方正男; 辻隆裕; 佐多徹太郎; 澤洋文; 長谷川秀樹, 日本ウイルス学会学術集会プログラム・抄録集, 56th, 247, 01 Oct. 2008
  Japanese
• ATL腫瘍細胞の走化性におけるCXCR4を介したシグナル経路の解析
  川口晶; 大場靖子; 木村享史; 伊波英克; 緒方正男; 佐多徹太郎; 澤洋文; 長谷川秀樹, 補体シンポジウム講演集, 45th, 179, Jul. 2008
  Japanese
• 【進行性多巣性白質脳症の新しい展開 PMLが治る時代へ向けて】 JCウイルスの最近の基礎的知見
  澤 洋文; 鈴木 忠樹; 大場 靖子; 寸田 祐嗣; 長嶋 和郎, BRAIN and NERVE: 神経研究の進歩, 59, 2, 101, 108, Feb. 2007
  (株)医学書院, Japanese
• JC virus agnoprotein modifies plasma membrane permeability and facilitates
  Tadaki Suzuki; Yuki Okada; Yasuko Orba; Yuji Sunden; Takashi Kimura; Kazuo Nagashima; Hirofumi Sawa, JOURNAL OF NEUROVIROLOGY, 13, 128, 128, 2007
  English, Summary international conference
• Pharmacological cdk inhibitor suppresses JC virus proliferation
  Yasuko Orba; Yuji Sunden; Tadaki Suzuki; Takashi Kimura; Shinya Tanaka; Kazuo Nagashima; Hirofumi Sawa, JOURNAL OF NEUROVIROLOGY, 13, 109, 110, 2007
  English, Summary international conference
• Biological effects of Natalizurnab on JC virus infectivity in its permissive human neural cell lines in vitro
  Tadaki Suzuki; Satoko Yamanouchi; Yuji Sunden; Yasuko Orba; Shinya Tanaka; Takashi Kimura; Hirofumi Sawa, JOURNAL OF NEUROVIROLOGY, 13, 127, 127, 2007
  English, Summary international conference
• Development of a model of adult T-cell leukemia/lymphoma (Atll)
  H Hasegawa; H Sawa; M Lewis; Y Orba; T Sata; T Kurata; K Nagashima; N Sheehy; WW Hall, AIDS RESEARCH AND HUMAN RETROVIRUSES, 21, 5, 454, 455, May 2005
  English, Summary international conference
• Agnoprotein plays a role in intracellular trafficking of JC virus
  Hirofumi Sawa; Tadaki Suzuki; Yuki Okada; Yasuko Orba; Yuji Sunden; Shingo Semba; Kazuo Nagashima, JOURNAL OF NEUROVIROLOGY, 14, 63, 63, 2005
  English, Summary international conference
• 6 頭蓋内悪性黒色腫の一例(脳・頭頸部3)
  渡辺 麻那美; 大場 靖子; 近井 佳奈子; 村田 純一; 田中 伸哉; 長嶋 和郎, 日本臨床細胞学会雑誌, 43, 2, 405, 405, 22 Sep. 2004
  特定非営利活動法人日本臨床細胞学会, Japanese
• JC virus agnoprotein facilitates viral propagation by removing FEZ1 from microtubules
  Tadaki Suzuki; Hirofumi Sawa; Yuki Okada; Yasuko Orba; Shingo Semba; Kazuo Nagashima, JOURNAL OF NEUROVIROLOGY, 10, 131, 131, 2004
  English, Summary international conference
• Identification of cell surface molecule as a candidate receptor for JC virus
  C. Henmi; H. Sawa; Y. Orba; S. Tanaka; K. Nagashima, JOURNAL OF NEUROVIROLOGY, 14, 42, 42, 2004
  English, Summary international conference
• Human polyomavirus agnoprotein disrupts the interaction between HP1 alpha and LBR
  H. Sawa; Y. Okada; T. Suzuki; Y. Orba; Y. Sunden; C. Henmi; S. Semba; H. Takahashi; S. Tanaka; K. Nagashima, JOURNAL OF NEUROVIROLOGY, 14, 33, 33, 2004
  English, Summary international conference
• 除負荷がin situ凍結解凍処理膝蓋腱のサイトカイン発現に与える効果
  内田 尚哉; 遠山 晴一; 松本 秀男; 戸山 芳昭; 大場 靖子; 長嶋 和郎; 安田 和則, 膝, 28, 2, 72, 75, Dec. 2003
  日本膝関節学会, Japanese
• 除負荷後の再負荷が膝蓋腱の力学的特性と線維芽細胞におけるサイトカイン発現に与える効果
  内田 尚哉; 遠山 晴一; 松本 秀男; 戸山 芳昭; 大場 靖子; 長嶋 和郎; 安田 和則, 日本臨床バイオメカニクス学会誌, 24, 77, 82, Nov. 2003
  ラット50匹を用い,除負荷後の再負荷が膝蓋腱の力学的特性およびサイトカイン発現に与える効果を検討した.2週間の除負荷後に再負荷を4週間施行した群は,6週間の除負荷を施行した群に比し,IL-1βおよびTNF-α発現では有意差を認めなかった.また,TGF-β発現は有意な低下を認めたもののsham群に比し高値を維持していた.一方,力学的特性に関しては,除負荷によりもたらされた膝蓋腱の弾性率の低下を再負荷は軽度ではあるものの増加させていた, 日本臨床バイオメカニクス学会, Japanese
• 307.術中細胞診が有用であった異型髄膜腫の一例(脳・頭頸部7)(第42回日本臨床細胞学会秋期大会)
  渡辺 麻那美; 大場 靖子; 田中 伸哉; 吉本 哲之; 長嶋 和郎, 日本臨床細胞学会雑誌, 42, 2, 495, 495, 22 Sep. 2003
  特定非営利活動法人日本臨床細胞学会, Japanese
• JC virusの神経親和性を規定する転写因子
  仙葉 愼吾; 長嶋 和郎; 大場 靖子; 澤 洋文, Neuropathology : official journal the Japanese Society of Neuropathology, 23, 202, 202, 01 May 2003
  Japanese
• siRNAを用いたJC virus関連蛋白質の発現抑制
  大場 靖子; 澤 洋文; 長嶋 和郎, Neuropathology : official journal the Japanese Society of Neuropathology, 23, 201, 201, 01 May 2003
  Japanese
• 非典型的な画像を呈し術中細胞診が有用であった膠肉腫の一例
  渡辺 麻那美; 大場 靖子; 清水 和子; 吉本 哲之; 田中 伸哉; 長嶋 和郎, 日本臨床細胞学会雑誌, 42, 1, 219, 219, 22 Mar. 2003
  特定非営利活動法人日本臨床細胞学会, Japanese
• 【新世紀の感染症学 ゲノム・グローバル時代の感染症アップデート】 グローバル時代の感染症学 ウイルス感染症 中枢神経系疾患 JCウイルス感染症(進行性多巣性白質脳症)
  中川 智子; 大場 靖子; 長嶋 和郎, 日本臨床, 61, 増刊2 新世紀の感染症学(上), 122, 127, Feb. 2003
  (株)日本臨床社, Japanese
• Expression of JC virus agnoprotein in progressive multifocal leukoencephalopathy brain
  AC Stan; Y Okada; H Sawa; S Endo; Y Orba; T Umemura; H Nishihara; S Tanaka; H Takahashi; K Nagashima, ACTA NEUROPATHOLOGICA, 104, 5, 570, 570, Nov. 2002
  English, Summary international conference
• 神経症状から診断されたSjogren症候群の臨床病理学的検討
  長嶋 淑子; 滝沢 昌彦; 間口 四郎; 佐々木 勲; 篠原 敏也; 大場 靖子; 中馬 孝容; 真野 行生; 澤 洋文; 長嶋 和郎; 酒井 宏一郎, Neuropathology : official journal the Japanese Society of Neuropathology, 22, 50, 50, 01 May 2002
  Japanese
• 進行性多巣性白質脳症(PML)におけるJC virus Agnoproteinの発現と局在
  大場 靖子; 岡田 由紀; 遠藤 秀一; 澤 洋文; 長嶋 和郎, Neuropathology : official journal the Japanese Society of Neuropathology, 22, 49, 49, 01 May 2002
  Japanese
• Laser capture microdissection法を用いた胆汁細胞診標本からの細胞採取とK‐ras遺伝子変異の検索
  大場靖子; 西原広史; 田中伸哉; 沢洋文; 伊藤智雄; 長嶋和郎, 日本病理学会会誌, 91, 1, 298, 28 Feb. 2002
  Japanese, Others
• JC virusの分子神経病理学
  大場 靖子; 澤 洋文; 長嶋 和郎, 脳と神経, 54, 2, 101, 109, Feb. 2002
  (株)医学書院, Japanese
• Molecular Neuropathology of JC Virus
  大場 靖子; 澤 洋文; 長嶋 和郎, Brain and nerve : 神経研究の進歩, 54, 2, 101, 109, Feb. 2002
  医学書院, Japanese
• 除負荷がラット膝蓋腱の力学的特性とTransforming Growth Factor-βの発現に与える効果に関する解析
  内田 尚哉; 遠山 晴一; 安田 和則; 松本 秀男; 戸山 芳昭; 大場 靖子; 長嶋 和郎, 膝, 26, 184, 188, Dec. 2001
  日本膝関節学会, Japanese
• 除負荷が膝蓋腱の力学的特性とサイトカイン発現に与える効果
  内田 尚哉; 遠山 晴一; 安田 和則; 大場 靖子; 長嶋 和郎; 松本 秀男; 戸山 芳昭, 日本整形外科学会雑誌, 75, 8, S1111, S1111, Aug. 2001
  (公社)日本整形外科学会, Japanese
• Broad distribution of the JC virus receptor contrasts with a marked cellular restriction of virus replication
  S Suzuki; H Sawa; R Komagome; Y Orba; M Yamada; Y Okada; Y Ishida; H Nishihara; S Tanaka; K Nagashima, VIROLOGY, 286, 1, 100, 112, Jul. 2001
  English
• JC virus(JCV) agnoproteinの発現と機能に関する研究
  岡田由紀; 沢洋文; 遠藤秀一; 大場靖子; 西原広史; 田中伸哉; 長嶋和郎, 日本病理学会会誌, 90, 1, 349, 01 Mar. 2001
  Japanese, Others
• Laser capture microdissection法を用いた細胞診標本からの遺伝子診断
  大場靖子; 西原広史; 伊藤智雄; 清水道生; 藤田美とし; 長嶋和郎, 日本臨床細胞学会雑誌, 39, 2, 327, 327, 22 Sep. 2000
  特定非営利活動法人日本臨床細胞学会, Japanese, Others
• Expression of JC virus early and late proteins in various cell lines and PML brains
  H Sawa; Y Okada; S Tanaka; Y Orba; S Endo; M Sasada; Y Hara; M Shintaku; K Nagashima, BRAIN PATHOLOGY, 10, 4, 757, 757, Sep. 2000
  English, Summary international conference
• JC virus 構成蛋白の発現と機能に関する研究
  岡田 由紀; 遠藤 秀一; 原 由紀子; 大場 靖子; 駒込 理佳; 田中 伸哉; 澤 洋文; 長嶋 和郎, Neuropathology : official journal the Japanese Society of Neuropathology, 20, 59, 59, 01 Jun. 2000
  Japanese
• 自家移植膝蓋腱へ浸潤する外来性細胞は生理的力学環境下においてもIII型コラーゲンを産生する
  遠山 晴一; 安田 和則; 東 裕隆; 大場 靖子; 長嶋 和郎, 日本整形外科学会雑誌, 73, 8, S1846, S1846, Aug. 1999
  (公社)日本整形外科学会, Japanese
• 新発色基質であるHNPPの神経疾患解析への応用 : FISH法、免疫染色法、in situ hybridization法の検討
  佐藤 真実; 高橋 文誉; 大場 靖子; 近藤 恭光; 鍵山 直人; 高橋 礼典; 澤 洋文; 長嶋 和郎, Neuropathology : official journal the Japanese Society of Neuropathology, 16, 203, 203, 01 May 1996
  Japanese
• B-16 有機水銀中毒症における小脳変性とアポトーシス
  藤井 幸子; 清水 和子; 中瀬 健一; 佐藤 真実; 大場 靖子; 奴久妻 聡一; 塚本 哲; 長嶋 和郎, 日本組織細胞化学会総会プログラムおよび抄録集, 36, 46, 46, 1995
  日本組織細胞化学会, Japanese

• ネオウイルス学
  河岡, 義裕
  集英社, Mar. 2021, 9784087211597, 315p, Japanese
• グローバル時代のウイルス感染症
  澤 洋文; 江下 優樹; 大場靖子, リフトバレー熱フレボウイルス感染症
  日本医事新報社, Jan. 2019, [Joint work]
• 蚊のはなし -病気との関わり-
  大場靖子, 蚊のうつす病気
  朝倉書店, Aug. 2017, [Joint work]

• 地球規模で広がる蚊媒介ウイルス感染症の対策に向けて
  大場 靖子
  2021年度 山口大学 中高温微生物研究センター発酵微生物部門シンポジウム「未来を拓くバイオSDGs」, 04 Mar. 2022, Nominated symposium
  [Invited]
• 多種の蚊に内在するフラビウイルスエレメント
  大場靖子
  第67回ウイルス学会, 30 Oct. 2019, English, Poster presentation
  [Domestic Conference]
• Research activities of Hokkaido Univ. CZC in Zambia
  Yasuko Orba
  Tsukuba Conference 2019, 02 Oct. 2019, English, Nominated symposium
  [Invited], [International presentation]
• Evidence of West Nile virus circulation in Zambia
  Yasuko Orba
  第66回日本ウイルス学会学術集会, 29 Oct. 2018, Japanese, Poster presentation
  [Domestic Conference]
• 節足動物媒介性ウイルス
  Yasuko Orba
  熱研サマースクール2018, 05 Aug. 2018, Japanese, Public discourse
  [Invited], [Domestic Conference]
• 蚊に潜むウイルス
  Yasuko Orba
  LOVE LABO 2018, 03 Aug. 2018, Japanese, Public discourse
  [Invited], [Domestic Conference]
• 病原体を見える化？
  Yasuko Orba
  WinterSchool2017＠微研, 25 Dec. 2017, Japanese, Public discourse
  [Invited], [Domestic Conference]
• An RNA virus enrichment approach for viral metagenomics using Ribonuclease R
  Yasuko Orba
  ConBio2017, 06 Dec. 2017, English, Poster presentation
  [Domestic Conference]
• Discovery of diverse mosquito-borne bunyaviruses in field-collected mosquitoes
  Yasuko Orba
  第65回日本ウイルス学会, 25 Oct. 2017, Japanese, Poster presentation
  [Domestic Conference]

• 感染病理学特論, 2024年, 博士後期課程, 国際感染症学院
• 人獣共通感染症対策専門特論, 2024年, 博士後期課程, 国際感染症学院
• 英語演習, 2024年, 学士課程, 全学教育

• THE MOLECULAR BIOLOGY SOCIETY OF JAPAN
• THE JAPANESE SOCIETY FOR VIROLOGY

• 非病原性フラビウイルスを用いた広域交差性エピトープの解析とワクチン開発
  科学研究費助成事業
  01 Apr. 2023 - 31 Mar. 2027
  大場 靖子; 福原 秀雄
  ISFV-IIに対するモノクローナル抗体作製のためのマウス免疫用の抗原として、ISFV-IIであるPsorophora virus（PSFV）を蚊培養細胞であるC6/36細胞を用いて増殖させ、20％ショ糖クッション法にてウイルスの濃縮を行い抗原とした。続いて、抗原を免疫したマウスの脾臓から、抗原特異的B細胞受容体を発現している胚中心由来B細胞をソーティングにより単離するためのプローブの作出を試みた。PSFVのCapsid（C）、precursor membrane（prM）並びにEnvelope（E）タンパク質から構成されるウイルス様粒子（VLP）の発現コンストラクトを作出し、哺乳類由来培養細胞を用いて発現させたVLPに蛍光標識することでプローブとした。また、胚中心由来B細胞をソーティングするための方法の最適化、また単離したB細胞の単細胞培養の方法を最適化した。フラビウイルスの中和試験並びにADE試験を迅速且つ高感度に判定するために、ウイルスを用いた試験法の代替試験法として、一回感染粒子（SRIP）を用いた試験の構築を試みた。SRIP作製のために、West Nile virus（WNV）由来のRepliconを発現するプラスミド、WNV由来のCタンパク質、並びに各フラビウイルスのprMEを発現するプラスミドを作出した。これらのプラスミドを哺乳類培養細胞に共発現させることでSRIPを作出し、中和試験法、並びにADE試験法を構築した。また、クライオ電子顕微鏡を用いてISFV-IIの構造解析を実施するためのウイルスを精製した。ショ糖クッション法にて濃縮したPSFVを密度勾配遠心法にて密度別にフラクションを回収し、PSFV抗原陽性だったフラクションを限外濾過膜にて濃縮した。PSFV抗原用性フラクションをさらにショ糖密度勾配遠心法によって精製したサンプルについて現在解析を進めている。
  日本学術振興会, 基盤研究(B), 北海道大学, 23K27831
• 非病原性フラビウイルスを用いた広域交差性エピトープの解析とワクチン開発
  科学研究費助成事業 基盤研究(B)
  01 Apr. 2023 - 31 Mar. 2027
  大場 靖子
  日本学術振興会, 基盤研究(B), 北海道大学, 23H03141
• Pathological method development for congenital infections and unexplained illness of possible infectious etiology
  新興・再興感染症に対する革新的医薬品等開発推進研究事業
  Apr. 2022 - Mar. 2025
  鈴木忠樹; 片野晴隆; 山下和男; 大場靖子; 林紀乃; 相澤志保子
  Japan Agency for Medical Research and Development, National Institute of Infectious Diseases, 22fk0108637
• VIrome analysis of bats in Indonesia
  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)
  01 Apr. 2021 - 31 Mar. 2024
  澤 洋文; 大場 靖子; 佐々木 道仁
  コウモリは人に致死的な肺炎、出血熱、脳炎、狂犬病 等の自然宿主と考えられている。現在の国際社会のCOVID-19の感染状況において、飛行機等による交通は制限されており、出入国の際には検疫が必要である。斯かる状況において、私達が海外において疫学活動を実施し、コウモリの検体を入手すること、および、海外からコウモリの検体を日本に輸送することは極めて困難である。
  そこで本研究計画では、申請者の所属する研究施設に保管されているインドネシアのコウモリから採集した検体を使用することによって、海外に渡航する困難を克服できる。コウモリは3つの島 （スマトラ、ジャワ、スラウェシ）から採集しており、インドネシア地域でのウイルスの多様性を確認することが可能である。多くのウイルスの自然宿主であるコウモリにおいて、新規・既知のウイルスの存在を明らかにすることを計画する。公衆衛生学的に重要と考えられるコウモリ由来のウイルスの存在を明らかにすることを本研究の目的とする。
  令和3年度においては我々の研究グループがアフリカのザンビアのコウモリから単離したオルソレオウイルスについて（Harima et al. PLos Negl Trop Dis, 2021)、アジアとアフリカでの地域差を確認するために、2010年から2014年にかけて、スマトラ、ジャワ、スラウェシで採集したコウモリを対象とし133検体の血清を用いて以下の実験を実施した。
  オーストラリアのコウモリから単離されたオルソレオウイルスである、Nelson Bayオルソレオウイルスを用いて、インドネシアのコウモリの血清の中和活性を測定した結果、133検体中、118検体（89.5%）に中和抗体価を確認した。
  Japan Society for the Promotion of Science, Grant-in-Aid for Scientific Research (C), Hokkaido University, 21K05951
• ウイルス-人体相互作用ネットワークの理解と制御
  MOONSHOT RESEARCH & DEVEROPMENT PROGRAM
  Dec. 2020 - Mar. 2024
  Yasuko Orba
  Japan Science and Technology Agency (JST), Coinvestigator, Competitive research funding, JPMJMS2025
• 昆虫特異的ウイルスによるアルボウイルス制御
  科学研究費助成事業 挑戦的研究(萌芽)
  30 Jul. 2020 - 31 Mar. 2023
  大場 靖子
  本研究では、蚊と共存し病原性の無い昆虫特異的ウイルスが、蚊に同時に感染したアルボウイルスの増殖に及ぼす影響を明らかにし、昆虫特異的ウイルス感染蚊によるアルボウイルス制御の可能性を示すことにより、蚊媒介性ウイルス感染症の予防策に繋げることを目的としている。
  本年度は、ボリビアの生息蚊から単離したネジェウイルス群のロレトウイルス(LORV)の成虫蚊、又は幼虫での増殖形態を明らかにするため、in vivo感染実験を実施した。Aedes albopictus, Aedes aegypti, Culex pipiensの成虫継代蚊にLORVを人口吸血により感染させ、2週間後の各蚊個体でのLORV感染をqRT-PCRで解析した結果、Ae. albopictusおよびAe. aegyptiで感染率が高く、ウイルスコピー数は多くの個体で10^8コピー以上であった。また、Ae. aegypti幼虫へのLORV感染実験の結果、幼虫への感染はほとんど認められず、LORVがシマカ成虫で効率的に増殖することが判明した。Ae. albopictusを用いたLORV感染モデルを用いて、LORVが蚊成体内で、デングウイルス２（DENV2）と共感染した際のウイルス増殖への影響を検討した、その結果、LORVとの共感染によるDENV2の感染率、および共感染した個体のDENV2ゲノム量は、DENV2感染単独の蚊と比較し顕著な差が認められなかった。このことから、昨年度に得られた蚊由来細胞でのLORV共感染によるDENV2増殖抑制の結果と異なり、蚊成体内では、双方のウイルスが共存し増殖し得ることが明らかとなった。
  日本学術振興会, 挑戦的研究(萌芽), 北海道大学, 20K21298
• Global profiling of pathogens in ticks
  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
  01 Apr. 2019 - 31 Mar. 2023
  松野 啓太; 大場 靖子; 梶原 将大
  本年度は主に、昨年度までに検出あるいは分離培養に成功したマダニ由来ウイルスについての性状解析を実施した。さらに、昨年度に報告したマダニ咬傷後に発熱を呈した患者から検出されたオルソナイロウイルス（Yezoウイルス)が分布していると考え られる北海道において、このウイルスを対象とした性状解析ならびに野外調査を実施した。なお、Yezoウイルスの調査については、他研究費との連携により効率的に展開した。また、マダニからオルソナイロウイルスを検出する系を樹立し、実際に北海道のマダニから複数のオルソナイロウイルスを検出した。本年度は新型コロナウ イルス感染拡大の影響で、海外でのマダニ採集が引き続き困難になったため、研究協力者からマダニを送付してもらうなどの代替策を進め、採集地域が限定されることを防いだ。さらに、実験室内で人為的にウイルスを接種したマダニを用いた実験モデルからのウイルス検出・分離培養方法検証を進め、状況が改善した際に野外調査を精力的に進められるよう準備を行った。 マダニから分離培養された各微生物については、遺伝情報解読と病原性解析の2つの性状比較解析を実施し、マダニから検出された各微生物は、サンガーシークエンサーと次世代シークエンサーを併用して遺伝子配列を解読した。さらに、これまでに収集あるいは分離培養したウイルスについて、実験動物を用いて病原性の検討を実施した。これらの結果を用いて、微生物のプロファイリングに用いることができる基礎的データを収集し、各微生物について塩基配列およびアミノ酸配列に基づいて病原性を推定する方法の検討を開始した。
  Japan Society for the Promotion of Science, Grant-in-Aid for Scientific Research (B), Hokkaido University, 19H03112
• Development of small compounds for pan-arenavirus therapy
  地球規模保健課題解決推進のための研究事業 日米医学協力計画の若手・女性育成のための日米共同研究公募
  Oct. 2021 - Mar. 2023
  AMED, 北海道大学, Principal investigator
• Study of comprehensive evaluation methods to determine immune modulators in flavivirus protection for the identification of high-risk groups and vaccination strategies
  新興・再興感染症研究基盤創生事業（海外拠点活用研究領域）
  Jul. 2020 - Mar. 2023
  AMED, 北海道大学, Principal investigator
• Development of pathological diagnostic methods for congenital infections and unexplained illness of possible infectious etiology.
  新興・再興感染症に対する革新的医薬品等開発推進研究事業
  Apr. 2019 - Mar. 2022
  鈴木忠樹; 長谷川秀樹; 有吉紅也; 片野晴隆; 山下和男; 大場靖子
  Japan Agency for Medical Research and Development, Competitive research funding, 21fk0108104
• Comprehensive investigation and functional analysis of endogenous viruses and endogenous viral elements in hematophagous arthropods and their host animals
  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)
  30 Jun. 2016 - 31 Mar. 2021
  Sawa Hirofumi
  Blood-sucking arthropods, such as mosquitoes and ticks, have been studied as vectors of pathogens, including zoonotic viruses.
  In this study, we examined especially endogenous viruses in ticks and mosquitoes and investigated their roles in nature under the “dance floor” hypothesis; in blood-sucking arthropods, their indigenous microorganisms and those from human or animal blood are communicating each other.
  As a result, we have discovered new endogenous viruses in the mosquitoes and ticks collected worldwide and found that these viruses may have co-evolved with their hosts.
  Japan Society for the Promotion of Science, Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area), Hokkaido University, 16H06431
• Investigation of latent pathogenic viruses in Latin America and South-eastern Asia by virome analysis
  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
  01 Apr. 2016 - 31 Mar. 2020
  Sawa Hirofumi
  We obtained results based on three projects that aimed at; 1) Implementing epidemiological research in Latin America and South-eastern Asia, 2) Analysis of viral genome using “Virome methods”, 3) Investigation of distribution, evolution and the infection cycle of viruses.
  During screening of mosquito and fruit bat samples collected in Latin America and South-eastern Asia, respectively, we detected novel viruses i.e. flaviviruses from mosquitoes and gammaherpesviruses from fruit bats. Furthermore, we performed Virome analysis using pooled fecal samples obtained from 50 non-human primates, from which we isolated a new small circular replication-associated protein (Rep) encoding single-stranded (CRESS) DNA viruses. Additionally, genome analysis of the polyomaviruses isolated from several different horseshoe bats indicated evidence of ‘host-switching’ during evolution in horseshoe bats.
  Japan Society for the Promotion of Science, Grant-in-Aid for Scientific Research (B), Hokkaido University, 16H05805
• Investigation of pathogenesis of SFTS using comprehensive immunoglobulin gene repertoire analysis
  医療研究開発推進事業費補助金（感染症研究革新イニシアティブ）
  Aug. 2017 - Mar. 2020
  鈴木忠樹; 大西和夫; 大場靖子
  Japan Agency for Medical Research and Development, Competitive research funding, 19fm0208002
• Comprehensive detection methods for mosquito-borne viruses
  Grants-in-Aid for Scientific Research
  2015 - 2018
  OBA YASUKO
  In order to identify arboviruses in mosquitoes, we have established the one-step RT-PCR methods to detect broad arboviruses by using degenerated primers against flaviviruses, alphaviruses, phleboviruses respectively. Using these comprehensive methods, we have identified many mosquito-borne viruses including previously unidentified viruses and West Nile virus from field-collected mosquitoes in Zambia. These methods were applied to the detection of arboviruses in samples from febrile human cases in Zambia.
  Our results using established methods provide the first evidence of the circulation of West Nile virus in Zambia and suggest there should be an increased awareness of possible associated human and animal diseases in that country.
  Japan Society for the Promotion of Science, Grant-in-Aid for Scientific Research (C), Hokkaido University, Principal investigator, Competitive research funding, 15K07715
• Surveillance of zoonotic viral diseases by an efficient method for detection of viral genomes
  Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
  01 Apr. 2012 - 31 Mar. 2016
  Sawa Hirofumi; ORBA Yasuko; ISHII Akihiro
  In this research, we have succeeded to establish the enrichment of the viral genomes from wildlife intestinal contents.
  Thereafter, we performed a deep sequencing using isolated and enriched viral nucleic acids. We have finally detected a novel bufavirus named as Mpulungu bufavirus from shrew feces. We have determined the almost whole genome sequences of the novel bufavirus which has a homology with that of human bufavirus (more than 51-53% of amino acid identity).
  Based on the obtained sequences, we established a nested PCR method for screening of bufaviruses in spleen samples from wildlife. Finally, we obtained the relatively high positive rate of detection of the viral genome in shrew (17/49=34.7%). Our established nested PCR method is useful for screening of the bufaviruses in mammals.
  Japan Society for the Promotion of Science, Grant-in-Aid for Scientific Research (B), Hokkaido University, 24405043
• Regulation of JC virus late gene expression
  Grants-in-Aid for Scientific Research
  2011 - 2013
  ORBA Yasuko
  JC virus(JCV) late transcripts, which produce agnoprotein, VP1, VP2, and VP3, are polycistronic mRNAs and processed by alternative splicing. We found that JCV late transcripts are processed into mature mRNAs using a unique splicing mechanism. Analysis of virus infection using JCV genomes with splice site mutations indicated that the identified splicing mechanism is necessary for efficient production of late proteins and viral infection.
  Japan Society for the Promotion of Science, Grant-in-Aid for Young Scientists (B), Hokkaido University, Principal investigator, Competitive research funding, 23790429
• Cell-specific regulation of the JC virus late gene
  Grants-in-Aid for Scientific Research(若手研究(スタートアップ))
  2008 - 2009
  Yasuko ORBA
  (1) Large T antigen of JC virus activates various promoters including JC virus late promoter. The enhancement of promoter activity by large T antigen is prominent in JC virus permissive cells. The cell-specific transcriptional regulation by T antigen is one of potential mechanisms of cell specificity for JC virus infection.(2) We found that caffeine had a pronounced antiviral effect in cells infected with JC virus. Caffeine suppressed expression of viral early proteins and viral DNA replication.
  Ministry of Education, Culture, Sports, Science and Technology, 若手研究(スタートアップ), 北海道大学, Principal investigator, Competitive research funding, 20890005
• ATLモデルマウスを用いたT-cell腫瘍化機構の解明と治療法の開発
  Grants-in-Aid for Scientific Research Grant-in-Aid for JSPS Fellows
  2005 - 2007
  大場 靖子
  HTLV-1 Taxトランスジェニックマウスで発症するT-cell腫瘍は、ATL患者由来の腫瘍細胞と同様にアポトーシス抑制因子であるsurvivinの発現が顕著に亢進していることから、これまでにsurvivinを標的とした腫瘍細胞の増殖抑制の検討を行ってきた。HMG-CoA還元酵素阻害剤であるスタチン系薬剤は高脂血症治療薬として臨床で広く用いられているが、白血病細胞をはじめとした多くの腫瘍細胞でapoptosisを誘導することが報告されている。スタチンの一つであるロバスタチンはsurvivinの発現を抑制効果、apoptosis誘導効果を検討した。本マウス由来のT-cell腫瘍細胞を培養し、ロバスタチン(5,10,20μM)またはDMSO(control)を培養液中に加え、48時間後に細胞を回収しapoptosisの測定を行った。その結果、ロバスタチンの濃度依存性にsurvivinのタンパク質発現量が低下し、それに伴いcaspase3の活性化が見られ、Annexin-Vに陽性のapoptosis細胞が増加することが明らかとなった。また、Tax発現細胞でのsurvivin発現増加はNF-kB経路の活性化を介していることから、IkBのリン酸化をimmunoblotにより検討した結果、ロバスタチンの濃度依存性にIkBのリン酸化が低下していた。このことから、ロバスタチンはNF-kBの活性化を抑制し、survivinの発現を低下させる可能性が示唆された。今後、腫瘍細胞を腹腔内接種したSCIDマウスまたはTaxトランスジェニックマウスを用いて、ロバスタチンのin vivoにおける効果を検討する予定である。
  Japan Society for the Promotion of Science, Grant-in-Aid for JSPS Fellows, Hokkaido University, 05J09123