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Abstract：
BACKGROUND: Although several SARS-CoV-2-related coronaviruses (SC2r-CoVs) were discovered in bats and pangolins, the differences in virological characteristics between SARS-CoV-2 and SC2r-CoVs remain poorly understood. Recently, BANAL-20-236 (B236) was isolated from a rectal swab of Malayan horseshoe bat and was found to lack a furin cleavage site (FCS) in the spike (S) protein. The comparison of its virological characteristics with FCS-deleted SARS-CoV-2 (SC2ΔFCS) has not been conducted yet.
METHODS: We prepared human induced pluripotent stem cell (iPSC)-derived airway and lung epithelial cells and colon organoids as human organ-relevant models. B236, SARS-CoV-2, and artificially generated SC2ΔFCS were used for viral experiments. To investigate the pathogenicity of B236 in vivo, we conducted intranasal infection experiments in hamsters.
RESULTS: In human iPSC-derived airway epithelial cells, the growth of B236 was significantly lower than that of the SC2ΔFCS. A fusion assay showed that the B236 and SC2ΔFCS S proteins were less fusogenic than the SARS-CoV-2 S protein. The infection experiment in hamsters showed that B236 was less pathogenic than SARS-CoV-2 and even SC2ΔFCS. Interestingly, in human colon organoids, the growth of B236 was significantly greater than that of SARS-CoV-2.
CONCLUSIONS: Compared to SARS-CoV-2, we demonstrated that B236 exhibited a tropism toward intestinal cells rather than respiratory cells. Our results are consistent with a previous report showing that B236 is enterotropic in macaques. Altogether, our report strengthens the assumption that SC2r-CoVs in horseshoe bats replicate primarily in the intestinal tissues rather than respiratory tissues.
BACKGROUND: This study was supported in part by AMED ASPIRE (JP23jf0126002, to Keita Matsuno, Kazuo Takayama, and Kei Sato); AMED SCARDA Japan Initiative for World-leading Vaccine Research and Development Centers \"UTOPIA\" (JP223fa627001, to Kei Sato), AMED SCARDA Program on R&D of new generation vaccine including new modality application (JP223fa727002, to Kei Sato); AMED SCARDA Hokkaido University Institute for Vaccine Research and Development (HU-IVReD) (JP223fa627005h0001, to Takasuke Fukuhara, and Keita Matsuno); AMED Research Program on Emerging and Re-emerging Infectious Diseases (JP21fk0108574, to Hesham Nasser; JP21fk0108493, to Takasuke Fukuhara; JP22fk0108617 to Takasuke Fukuhara; JP22fk0108146, to Kei Sato; JP21fk0108494 to G2P-Japan Consortium, Keita Matsuno, Shinya Tanaka, Terumasa Ikeda, Takasuke Fukuhara, and Kei Sato; JP21fk0108425, to Kazuo Takayama and Kei Sato; JP21fk0108432, to Kazuo Takayama, Takasuke Fukuhara and Kei Sato; JP22fk0108534, Terumasa Ikeda, and Kei Sato; JP22fk0108511, to Yuki Yamamoto, Terumasa Ikeda, Keita Matsuno, Shinya Tanaka, Kazuo Takayama, Takasuke Fukuhara, and Kei Sato; JP22fk0108506, to Kazuo Takayama and Kei Sato); AMED Research Program on HIV/AIDS (JP22fk0410055, to Terumasa Ikeda; and JP22fk0410039, to Kei Sato); AMED Japan Program for Infectious Diseases Research and Infrastructure (JP22wm0125008 to Keita Matsuno); AMED CREST (JP21gm1610005, to Kazuo Takayama; JP22gm1610008, to Takasuke Fukuhara; JST PRESTO (JPMJPR22R1, to Jumpei Ito); JST CREST (JPMJCR20H4, to Kei Sato); JSPS KAKENHI Fund for the Promotion of Joint International Research (International Leading Research) (JP23K20041, to G2P-Japan Consortium, Keita Matsuno, Takasuke Fukuhara and Kei Sato); JST SPRING (JPMJSP2108 to Shigeru Fujita); JSPS KAKENHI Grant-in-Aid for Scientific Research C (22K07103, to Terumasa Ikeda); JSPS KAKENHI Grant-in-Aid for Scientific Research B (21H02736, to Takasuke Fukuhara); JSPS KAKENHI Grant-in-Aid for Early-Career Scientists (22K16375, to Hesham Nasser; 20K15767, to Jumpei Ito); JSPS Core-to-Core Program (A. Advanced Research Networks) (JPJSCCA20190008, to Kei Sato); JSPS Research Fellow DC2 (22J11578, to Keiya Uriu); JSPS Research Fellow DC1 (23KJ0710, to Yusuke Kosugi); JSPS Leading Initiative for Excellent Young Researchers (LEADER) (to Terumasa Ikeda); World-leading Innovative and Smart Education (WISE) Program 1801 from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) (to Naganori Nao); Ministry of Health, Labour and Welfare (MHLW) under grant 23HA2010 (to Naganori Nao and Keita Matsuno); The Cooperative Research Program (Joint Usage/Research Center program) of Institute for Life and Medical Sciences, Kyoto University (to Kei Sato); International Joint Research Project of the Institute of Medical Science, the University of Tokyo (to Terumasa Ikeda and Takasuke Fukuhara); The Tokyo Biochemical Research Foundation (to Kei Sato); Takeda Science Foundation (to Terumasa Ikeda and Takasuke Fukuhara); Mochida Memorial Foundation for Medical and Pharmaceutical Research (to Terumasa Ikeda); The Naito Foundation (to Terumasa Ikeda); Hokuto Foundation for Bioscience (to Tomokazu Tamura); Hirose Foundation (to Tomokazu Tamura); and Mitsubishi Foundation (to Kei Sato).
METHODS: We prepared human induced pluripotent stem cell (iPSC)-derived airway and lung epithelial cells and colon organoids as human organ-relevant models. B236, SARS-CoV-2, and artificially generated SC2ΔFCS were used for viral experiments. To investigate the pathogenicity of B236 in vivo, we conducted intranasal infection experiments in hamsters.
RESULTS: In human iPSC-derived airway epithelial cells, the growth of B236 was significantly lower than that of the SC2ΔFCS. A fusion assay showed that the B236 and SC2ΔFCS S proteins were less fusogenic than the SARS-CoV-2 S protein. The infection experiment in hamsters showed that B236 was less pathogenic than SARS-CoV-2 and even SC2ΔFCS. Interestingly, in human colon organoids, the growth of B236 was significantly greater than that of SARS-CoV-2.
CONCLUSIONS: Compared to SARS-CoV-2, we demonstrated that B236 exhibited a tropism toward intestinal cells rather than respiratory cells. Our results are consistent with a previous report showing that B236 is enterotropic in macaques. Altogether, our report strengthens the assumption that SC2r-CoVs in horseshoe bats replicate primarily in the intestinal tissues rather than respiratory tissues.
BACKGROUND: This study was supported in part by AMED ASPIRE (JP23jf0126002, to Keita Matsuno, Kazuo Takayama, and Kei Sato); AMED SCARDA Japan Initiative for World-leading Vaccine Research and Development Centers \"UTOPIA\" (JP223fa627001, to Kei Sato), AMED SCARDA Program on R&D of new generation vaccine including new modality application (JP223fa727002, to Kei Sato); AMED SCARDA Hokkaido University Institute for Vaccine Research and Development (HU-IVReD) (JP223fa627005h0001, to Takasuke Fukuhara, and Keita Matsuno); AMED Research Program on Emerging and Re-emerging Infectious Diseases (JP21fk0108574, to Hesham Nasser; JP21fk0108493, to Takasuke Fukuhara; JP22fk0108617 to Takasuke Fukuhara; JP22fk0108146, to Kei Sato; JP21fk0108494 to G2P-Japan Consortium, Keita Matsuno, Shinya Tanaka, Terumasa Ikeda, Takasuke Fukuhara, and Kei Sato; JP21fk0108425, to Kazuo Takayama and Kei Sato; JP21fk0108432, to Kazuo Takayama, Takasuke Fukuhara and Kei Sato; JP22fk0108534, Terumasa Ikeda, and Kei Sato; JP22fk0108511, to Yuki Yamamoto, Terumasa Ikeda, Keita Matsuno, Shinya Tanaka, Kazuo Takayama, Takasuke Fukuhara, and Kei Sato; JP22fk0108506, to Kazuo Takayama and Kei Sato); AMED Research Program on HIV/AIDS (JP22fk0410055, to Terumasa Ikeda; and JP22fk0410039, to Kei Sato); AMED Japan Program for Infectious Diseases Research and Infrastructure (JP22wm0125008 to Keita Matsuno); AMED CREST (JP21gm1610005, to Kazuo Takayama; JP22gm1610008, to Takasuke Fukuhara; JST PRESTO (JPMJPR22R1, to Jumpei Ito); JST CREST (JPMJCR20H4, to Kei Sato); JSPS KAKENHI Fund for the Promotion of Joint International Research (International Leading Research) (JP23K20041, to G2P-Japan Consortium, Keita Matsuno, Takasuke Fukuhara and Kei Sato); JST SPRING (JPMJSP2108 to Shigeru Fujita); JSPS KAKENHI Grant-in-Aid for Scientific Research C (22K07103, to Terumasa Ikeda); JSPS KAKENHI Grant-in-Aid for Scientific Research B (21H02736, to Takasuke Fukuhara); JSPS KAKENHI Grant-in-Aid for Early-Career Scientists (22K16375, to Hesham Nasser; 20K15767, to Jumpei Ito); JSPS Core-to-Core Program (A. Advanced Research Networks) (JPJSCCA20190008, to Kei Sato); JSPS Research Fellow DC2 (22J11578, to Keiya Uriu); JSPS Research Fellow DC1 (23KJ0710, to Yusuke Kosugi); JSPS Leading Initiative for Excellent Young Researchers (LEADER) (to Terumasa Ikeda); World-leading Innovative and Smart Education (WISE) Program 1801 from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) (to Naganori Nao); Ministry of Health, Labour and Welfare (MHLW) under grant 23HA2010 (to Naganori Nao and Keita Matsuno); The Cooperative Research Program (Joint Usage/Research Center program) of Institute for Life and Medical Sciences, Kyoto University (to Kei Sato); International Joint Research Project of the Institute of Medical Science, the University of Tokyo (to Terumasa Ikeda and Takasuke Fukuhara); The Tokyo Biochemical Research Foundation (to Kei Sato); Takeda Science Foundation (to Terumasa Ikeda and Takasuke Fukuhara); Mochida Memorial Foundation for Medical and Pharmaceutical Research (to Terumasa Ikeda); The Naito Foundation (to Terumasa Ikeda); Hokuto Foundation for Bioscience (to Tomokazu Tamura); Hirose Foundation (to Tomokazu Tamura); and Mitsubishi Foundation (to Kei Sato).
摘要：
背景：尽管在蝙蝠和穿山甲中发现了几种与SARS-CoV-2相关的冠状病毒（SC2r-CoV），SARS-CoV-2和SC2r-CoV之间的病毒学特征差异仍然知之甚少。最近,BANAL-20-236（B236）从马来亚马蹄蝙蝠的直肠拭子中分离出来，发现刺突（S）蛋白中缺乏弗林蛋白酶切割位点（FCS）。尚未进行其病毒学特征与FCS缺失的SARS-CoV-2（SC2ΔFCS）的比较。
方法：我们制备了人诱导多能干细胞（iPSC）来源的气道和肺上皮细胞以及结肠类器官作为人体器官相关模型。B236，SARS-CoV-2和人工产生的SC2ΔFCS用于病毒实验。研究B236在体内的致病性，我们在仓鼠中进行了鼻内感染实验。
结果：在人iPSC来源的气道上皮细胞中，B236的生长明显低于SC2ΔFCS。融合分析表明，B236和SC2ΔFCSS蛋白的融合性低于SARS-CoV-2S蛋白。仓鼠感染实验表明，B236的致病性低于SARS-CoV-2，甚至低于SC2ΔFCS。有趣的是,在人类结肠类器官中，B236的增长明显大于SARS-CoV-2。
结论：与SARS-CoV-2相比，我们证明B236表现出对肠道细胞而不是呼吸道细胞的嗜性。我们的结果与先前的报告一致，该报告显示B236在猕猴中具有肠促功能。总之,我们的报告强化了以下假设:马蹄蝙蝠中的SC2r-CoV主要在肠道组织而不是呼吸道组织中复制.
背景：这项研究部分得到了AMEDASPIRE的支持（JP23jf0126002，给KeitaMatsuno，高山和夫,和KeiSato）；AMEDSCARDA日本世界领先的疫苗研发中心“UTOPIA”倡议（JP223fa627001，至KeiSato），AMEDSCARDA新一代疫苗研发计划，包括新模式应用（JP223fa727002，致KeiSato）；AMEDSCARDA北海道大学疫苗研究与开发研究所（HU-IVReD）（JP223fa627005h0001，致福原高介，和KeitaMatsuno）；关于新兴和再新兴传染病的AMED研究计划（JP21fk0108574，给HeshamNasser；JP21fk0108493，给福原高介；JP22fk0108617给福原高介；JP22fk0108146，给日本KekiSato0G194KeitaMatsuno,田中新也,池田Terumasa,福原高介,和KeiSato；JP21fk0108425，给高山一夫和和KeiSato；JP21fk0108432，给高山一夫，福原高介和佐藤Kei；JP22fk0108534，池田英正，和KeiSato；JP22fk0108511，到YamamotoYuki，池田Terumasa,KeitaMatsuno,田中新也,高山和夫,福原高介,和KeiSato；JP22fk0108506，给KazuoTaka和KeiSato联合研究，给Takuama和Keto）；AMED艾滋病毒/艾滋病研究计划（JP22fk0410055，给TerumasaIkeda；JP22fk0410039，给KeiSato）；日本传染病研究和基础设施计划（JP22wm012500KeitaMatsuno,FukuharaTakasukeandKeiSato);高级研究网络）（JPJSCCA20190008，给KeiSato）；JSPS研究员DC2（22J11578，给KeiyaUriu）；JSPS研究员DC1（23KJ0710，给YusukeKosugi）；JSPS优秀青年研究人员领导计划（LEADER）（给TerumasaIkeda）；世界领先的创新与智能教育计划（ISW01文化,体育,科学技术（MEXT）（至NaganoriNao）；卫生部，劳工和福利（MHLW）授予23HA2010（给NaganoriNao和KeitaMatsuno）；生命与医学科学研究所的合作研究计划（联合使用/研究中心计划），京都大学（toKeiSato）；医学科学研究所国际联合研究项目，东京大学（至池田英正和福原高介）；东京生化研究基金会（至佐藤基业）；武田科学基金会（至池田英正和福原高介）；摩田医学和药物研究纪念基金会（至池田英正）；田村恒业基金会（至三菱公司）（至佐田广园）；
方法：我们制备了人诱导多能干细胞（iPSC）来源的气道和肺上皮细胞以及结肠类器官作为人体器官相关模型。B236，SARS-CoV-2和人工产生的SC2ΔFCS用于病毒实验。研究B236在体内的致病性，我们在仓鼠中进行了鼻内感染实验。
结果：在人iPSC来源的气道上皮细胞中，B236的生长明显低于SC2ΔFCS。融合分析表明，B236和SC2ΔFCSS蛋白的融合性低于SARS-CoV-2S蛋白。仓鼠感染实验表明，B236的致病性低于SARS-CoV-2，甚至低于SC2ΔFCS。有趣的是,在人类结肠类器官中，B236的增长明显大于SARS-CoV-2。
结论：与SARS-CoV-2相比，我们证明B236表现出对肠道细胞而不是呼吸道细胞的嗜性。我们的结果与先前的报告一致，该报告显示B236在猕猴中具有肠促功能。总之,我们的报告强化了以下假设:马蹄蝙蝠中的SC2r-CoV主要在肠道组织而不是呼吸道组织中复制.
背景：这项研究部分得到了AMEDASPIRE的支持（JP23jf0126002，给KeitaMatsuno，高山和夫,和KeiSato）；AMEDSCARDA日本世界领先的疫苗研发中心“UTOPIA”倡议（JP223fa627001，至KeiSato），AMEDSCARDA新一代疫苗研发计划，包括新模式应用（JP223fa727002，致KeiSato）；AMEDSCARDA北海道大学疫苗研究与开发研究所（HU-IVReD）（JP223fa627005h0001，致福原高介，和KeitaMatsuno）；关于新兴和再新兴传染病的AMED研究计划（JP21fk0108574，给HeshamNasser；JP21fk0108493，给福原高介；JP22fk0108617给福原高介；JP22fk0108146，给日本KekiSato0G194KeitaMatsuno,田中新也,池田Terumasa,福原高介,和KeiSato；JP21fk0108425，给高山一夫和和KeiSato；JP21fk0108432，给高山一夫，福原高介和佐藤Kei；JP22fk0108534，池田英正，和KeiSato；JP22fk0108511，到YamamotoYuki，池田Terumasa,KeitaMatsuno,田中新也,高山和夫,福原高介,和KeiSato；JP22fk0108506，给KazuoTaka和KeiSato联合研究，给Takuama和Keto）；AMED艾滋病毒/艾滋病研究计划（JP22fk0410055，给TerumasaIkeda；JP22fk0410039，给KeiSato）；日本传染病研究和基础设施计划（JP22wm012500KeitaMatsuno,FukuharaTakasukeandKeiSato);高级研究网络）（JPJSCCA20190008，给KeiSato）；JSPS研究员DC2（22J11578，给KeiyaUriu）；JSPS研究员DC1（23KJ0710，给YusukeKosugi）；JSPS优秀青年研究人员领导计划（LEADER）（给TerumasaIkeda）；世界领先的创新与智能教育计划（ISW01文化,体育,科学技术（MEXT）（至NaganoriNao）；卫生部，劳工和福利（MHLW）授予23HA2010（给NaganoriNao和KeitaMatsuno）；生命与医学科学研究所的合作研究计划（联合使用/研究中心计划），京都大学（toKeiSato）；医学科学研究所国际联合研究项目，东京大学（至池田英正和福原高介）；东京生化研究基金会（至佐藤基业）；武田科学基金会（至池田英正和福原高介）；摩田医学和药物研究纪念基金会（至池田英正）；田村恒业基金会（至三菱公司）（至佐田广园）；
