{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,20]],"date-time":"2026-06-20T09:07:16Z","timestamp":1781946436501,"version":"3.54.5"},"reference-count":69,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2020,9,29]],"date-time":"2020-09-29T00:00:00Z","timestamp":1601337600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BB\/R019576\/1"],"award-info":[{"award-number":["BB\/R019576\/1"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BBS\/E\/I\/00001977"],"award-info":[{"award-number":["BBS\/E\/I\/00001977"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BBS\/E\/I\/00007034"],"award-info":[{"award-number":["BBS\/E\/I\/00007034"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>The Gammacoronavirus infectious bronchitis virus (IBV) causes a highly contagious and economically important respiratory disease in poultry. In the laboratory, most IBV strains are restricted to replication in ex vivo organ cultures or in ovo and do not replicate in cell culture, making the study of their basic virology difficult. Entry of IBV into cells is facilitated by the large glycoprotein on the surface of the virion, the spike (S) protein, comprised of S1 and S2 subunits. Previous research showed that the S2\u2032 cleavage site is responsible for the extended tropism of the IBV Beaudette strain. This study aims to investigate whether protease treatment can extend the tropism of other IBV strains. Here we demonstrate that the addition of exogenous trypsin during IBV propagation in cell culture results in significantly increased viral titres. Using a panel of IBV strains, exhibiting varied tropisms, the effects of spike cleavage on entry and replication were assessed by serial passage cell culture in the presence of trypsin. Replication could be maintained over serial passages, indicating that the addition of exogenous protease is sufficient to overcome the barrier to infection. Mutations were identified in both S1 and S2 subunits following serial passage in cell culture. This work provides a proof of concept that exogenous proteases can remove the barrier to IBV replication in otherwise non-permissive cells, providing a platform for further study of elusive field strains and enabling sustainable vaccine production in vitro.<\/jats:p>","DOI":"10.3390\/v12101102","type":"journal-article","created":{"date-parts":[[2020,9,29]],"date-time":"2020-09-29T08:43:27Z","timestamp":1601369007000},"page":"1102","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":8,"title":["Treatment with Exogenous Trypsin Expands In Vitro Cellular Tropism of the Avian Coronavirus Infectious Bronchitis Virus"],"prefix":"10.3390","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-6509-3354","authenticated-orcid":false,"given":"Phoebe","family":"Stevenson-Leggett","sequence":"first","affiliation":[{"name":"The Pirbright Institute, Ash Road, Woking, Surrey GU24 0NF, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sarah","family":"Keep","sequence":"additional","affiliation":[{"name":"The Pirbright Institute, Ash Road, Woking, Surrey GU24 0NF, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4012-1283","authenticated-orcid":false,"given":"Erica","family":"Bickerton","sequence":"additional","affiliation":[{"name":"The Pirbright Institute, Ash Road, Woking, Surrey GU24 0NF, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,9,29]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1394","DOI":"10.1126\/science.1085952","article-title":"Characterization of a Novel Coronavirus Associated with Severe Acute Respiratory Syndrome","volume":"300","author":"Rota","year":"2003","journal-title":"Science"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"635","DOI":"10.1128\/MMBR.69.4.635-664.2005","article-title":"Coronavirus Pathogenesis and the Emerging Pathogen Severe Acute Respiratory Syndrome Coronavirus","volume":"69","author":"Weiss","year":"2005","journal-title":"Microbiol. Mol. Boil. Rev."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1814","DOI":"10.1056\/NEJMoa1211721","article-title":"Isolation of a Novel Coronavirus from a Man with Pneumonia in Saudi Arabia","volume":"367","author":"Zaki","year":"2012","journal-title":"N. Engl. J. Med."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"727","DOI":"10.1056\/NEJMoa2001017","article-title":"A Novel Coronavirus from Patients with Pneumonia in China, 2019","volume":"382","author":"Zhu","year":"2020","journal-title":"N. Engl. J. Med."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"349","DOI":"10.1016\/j.meegid.2016.02.015","article-title":"S1 gene-based phylogeny of infectious bronchitis virus: An attempt to harmonize virus classification","volume":"39","author":"Valastro","year":"2016","journal-title":"Infect. Genet. Evol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"281","DOI":"10.1051\/vetres:2006055","article-title":"Coronavirus avian infectious bronchitis virus","volume":"38","author":"Cavanagh","year":"2007","journal-title":"Veter Res."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"239","DOI":"10.1080\/03079457.2012.680432","article-title":"The long view: 40 years of infectious bronchitis research","volume":"41","author":"Cook","year":"2012","journal-title":"Avian Pathol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"71","DOI":"10.1080\/03079450094108","article-title":"Detection of infectious bronchitis virus","volume":"29","year":"2000","journal-title":"Avian Pathol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1155\/2015\/424860","article-title":"Progress and Challenges toward the Development of Vaccines against Avian Infectious Bronchitis","volume":"2015","author":"Bande","year":"2015","journal-title":"J. Immunol. Res."},{"key":"ref_10","first-page":"112","article-title":"Modification of the avian coronavirus infectious bronchitis virus for vaccine development","volume":"3","author":"Britton","year":"2012","journal-title":"Bioeng. Bugs"},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Armesto, M., Evans, S., Cavanagh, D., Abu-Median, A.-B., Keep, S., and Britton, P. (2011). A Recombinant Avian Infectious Bronchitis Virus Expressing a Heterologous Spike Gene Belonging to the 4\/91 Serotype. PLoS ONE, 6.","DOI":"10.1371\/journal.pone.0024352"},{"key":"ref_12","first-page":"51","article-title":"Cultivation of the virus of infectious bronchitis","volume":"90","author":"Beaudette","year":"1937","journal-title":"J. Am. Vet. Med. Assoc."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1011","DOI":"10.3390\/v4061011","article-title":"Mechanisms of Coronavirus Cell Entry Mediated by the Viral Spike Protein","volume":"4","author":"Belouzard","year":"2012","journal-title":"Viruses"},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Shang, J., Zheng, Y., Yang, Y., Liu, C., Geng, Q., Luo, C., Zhang, W., and Li, F. (2018). Cryo-EM structure of infectious bronchitis coronavirus spike protein reveals structural and functional evolution of coronavirus spike proteins. PLoS Pathog., 14.","DOI":"10.2210\/pdb6cv0\/pdb"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"2452","DOI":"10.1128\/JVI.75.5.2452-2457.2001","article-title":"Murine Coronavirus Spike Protein Determines the Ability of the Virus To Replicate in the Liver and Cause Hepatitis","volume":"75","author":"Seo","year":"2001","journal-title":"J. Virol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"7607","DOI":"10.1128\/JVI.73.9.7607-7618.1999","article-title":"Targeted Recombination Demonstrates that the Spike Gene of Transmissible Gastroenteritis Coronavirus Is a Determinant of Its Enteric Tropism and Virulence","volume":"73","author":"Izeta","year":"1999","journal-title":"J. Virol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"14122","DOI":"10.1128\/JVI.79.22.14122-14130.2005","article-title":"Acquisition of Macrophage Tropism during the Pathogenesis of Feline Infectious Peritonitis Is Determined by Mutations in the Feline Coronavirus Spike Protein","volume":"79","author":"Rottier","year":"2005","journal-title":"J. Virol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1634","DOI":"10.1038\/sj.emboj.7600640","article-title":"Receptor and viral determinants of SARS-coronavirus adaptation to human ACE2","volume":"24","author":"Li","year":"2005","journal-title":"EMBO J."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"13804","DOI":"10.1128\/JVI.78.24.13804-13811.2004","article-title":"Recombinant Infectious Bronchitis Coronavirus Beaudette with the Spike Protein Gene of the Pathogenic M41 Strain Remains Attenuated but Induces Protective Immunity","volume":"78","author":"Hodgson","year":"2004","journal-title":"J. Virol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1681","DOI":"10.1099\/jgv.0.001167","article-title":"Recombinant infectious bronchitis viruses expressing heterologous S1 subunits: Potential for a new generation of vaccines that replicate in Vero cells","volume":"99","author":"Bickerton","year":"2018","journal-title":"J. Gen. Virol."},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Bickerton, E., Maier, H.J., Stevenson-Leggett, P., Armesto, M., and Britton, P. (2018). The S2 Subunit of Infectious Bronchitis Virus Beaudette Is a Determinant of Cellular Tropism. J. Virol., 92.","DOI":"10.1128\/JVI.01044-18"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"9084","DOI":"10.1128\/JVI.77.16.9084-9089.2003","article-title":"Recombinant Avian Infectious Bronchitis Virus Expressing a Heterologous Spike Gene Demonstrates that the Spike Protein Is a Determinant of Cell Tropism","volume":"77","author":"Casais","year":"2003","journal-title":"J. Virol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"12359","DOI":"10.1128\/JVI.75.24.12359-12369.2001","article-title":"Reverse Genetics System for the Avian Coronavirus Infectious Bronchitis Virus","volume":"75","author":"Casais","year":"2001","journal-title":"J. Virol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"114","DOI":"10.1038\/nature16988","article-title":"Cryo-electron microscopy structure of a coronavirus spike glycoprotein trimer","volume":"531","author":"Walls","year":"2016","journal-title":"Nature"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"118","DOI":"10.1038\/nature17200","article-title":"Pre-fusion structure of a human coronavirus spike protein","volume":"531","author":"Kirchdoerfer","year":"2016","journal-title":"Nature"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"2281","DOI":"10.1099\/0022-1317-65-12-2281","article-title":"Monoclonal Antibodies to the S1 Spike and Membrane Proteins of Avian Infectious Bronchitis Coronavirus Strain Massachusetts M41","volume":"65","author":"Mockett","year":"1984","journal-title":"J. Gen. Virol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"133","DOI":"10.1016\/0168-1702(86)90037-7","article-title":"Coronavirus IBV: Partial amino terminal sequencing of spike polypeptide S2 identifies the sequence Arg-Arg-Phe-Arg-Arg at the cleavage site of the spike precursor propolypeptide of IBV strains Beaudette and M41","volume":"4","author":"Cavanagh","year":"1986","journal-title":"Virus Res."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"313","DOI":"10.1080\/03079459508419072","article-title":"Immune responses to structural proteins of avian infectious bronchitis virus","volume":"24","author":"Ignjatovic","year":"1995","journal-title":"Avian Pathol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"11157","DOI":"10.1073\/pnas.1708727114","article-title":"Tectonic conformational changes of a coronavirus spike glycoprotein promote membrane fusion","volume":"114","author":"Walls","year":"2017","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Yamada, Y., Liu, X.B., Fang, S.G., Tay, F.P.L., and Liu, D.X. (2009). Acquisition of Cell\u2013Cell Fusion Activity by Amino Acid Substitutions in Spike Protein Determines the Infectivity of a Coronavirus in Cultured Cells. PLoS ONE, 4.","DOI":"10.1371\/journal.pone.0006130"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"12262","DOI":"10.1073\/pnas.1608147113","article-title":"Proteolytic processing of Middle East respiratory syndrome coronavirus spikes expands virus tropism","volume":"113","author":"Park","year":"2016","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"779","DOI":"10.1016\/j.molcel.2020.04.022","article-title":"A Multibasic Cleavage Site in the Spike Protein of SARS-CoV-2 Is Essential for Infection of Human Lung Cells","volume":"78","author":"Hoffmann","year":"2020","journal-title":"Mol. Cell"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"8744","DOI":"10.1128\/JVI.00613-09","article-title":"Proteolytic Activation of the Spike Protein at a Novel RRRR\/S Motif Is Implicated in Furin-Dependent Entry, Syncytium Formation, and Infectivity of Coronavirus Infectious Bronchitis Virus in Cultured Cells","volume":"83","author":"Yamada","year":"2009","journal-title":"J. Virol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"5871","DOI":"10.1073\/pnas.0809524106","article-title":"Activation of the SARS coronavirus spike protein via sequential proteolytic cleavage at two distinct sites","volume":"106","author":"Belouzard","year":"2009","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"15214","DOI":"10.1073\/pnas.1407087111","article-title":"Host cell entry of Middle East respiratory syndrome coronavirus after two-step, furin-mediated activation of the spike protein","volume":"111","author":"Millet","year":"2014","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"421","DOI":"10.1016\/j.virol.2012.08.037","article-title":"Characterization of cellular furin content as a potential factor determining the susceptibility of cultured human and animal cells to coronavirus infectious bronchitis virus infection","volume":"433","author":"Tay","year":"2012","journal-title":"Virology"},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Menachery, V.D., Dinnon, K.H., Yount, B.L., McAnarney, E.T., Gralinski, L.E., Hale, A., Graham, R.L., Scobey, T., Anthony, S.J., and Wang, L. (2019). Trypsin Treatment Unlocks Barrier for Zoonotic Bat Coronavirus Infection. J. Virol., 94.","DOI":"10.1101\/768663"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"2235","DOI":"10.1128\/jcm.26.11.2235-2239.1988","article-title":"Propagation of the virus of porcine epidemic diarrhea in cell culture","volume":"26","author":"Hofmann","year":"1988","journal-title":"J. Clin. Microbiol."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1007\/BF02123572","article-title":"Plaque assay and primary isolation of influenza a viruses in an established line of canine kidney cells (MDCK) in the presence of trypsin","volume":"162","author":"Tobita","year":"1975","journal-title":"Med Microbiol. Immunol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"7952","DOI":"10.1128\/JVI.00297-14","article-title":"Proteolytic Activation of the Porcine Epidemic Diarrhea Coronavirus Spike Fusion Protein by Trypsin in Cell Culture","volume":"88","author":"Wicht","year":"2014","journal-title":"J. Virol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"607","DOI":"10.1586\/erv.09.19","article-title":"Vero cell platform in vaccine production: Moving towards cell culture-based viral vaccines","volume":"8","author":"Barrett","year":"2009","journal-title":"Expert Rev. Vaccines"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"295","DOI":"10.1006\/viro.1998.9290","article-title":"The DF-1 Chicken Fibroblast Cell Line: Transformation Induced by Diverse Oncogenes and Cell Death Resulting from Infection by Avian Leukosis Viruses","volume":"248","author":"Himly","year":"1998","journal-title":"Virology"},{"key":"ref_43","doi-asserted-by":"crossref","unstructured":"Maier, H., Bickerton, E., and Britton, P. (2015). The preparation of chicken kidney cell cultures for virus propagation. Coronaviruses, Humana Press.","DOI":"10.1007\/978-1-4939-2438-7"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"227","DOI":"10.1007\/BF01318057","article-title":"Taxonomic studies on strains of avian infectious bronchitis virus using neutralisation tests in tracheal organ cultures","volume":"61","author":"Darbyshire","year":"1979","journal-title":"Arch Virol."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"203","DOI":"10.1016\/j.jviromet.2004.09.017","article-title":"Generation of a recombinant avian coronavirus infectious bronchitis virus using transient dominant selection","volume":"123","author":"Britton","year":"2005","journal-title":"J. Virol. Methods"},{"key":"ref_46","doi-asserted-by":"crossref","unstructured":"Cavanagh, D. (2008). SARS- and Other Coronaviruses. Methods in Molecular Biology, Humana Press.","DOI":"10.1007\/978-1-59745-181-9"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"223","DOI":"10.1080\/03079457.2011.566260","article-title":"Infectious bronchitis virus variants: A review of the history, current situation and control measures","volume":"40","author":"Cook","year":"2011","journal-title":"Avian Pathol."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"20","DOI":"10.1128\/JVI.01358-06","article-title":"A Contemporary View of Coronavirus Transcription","volume":"81","author":"Sawicki","year":"2006","journal-title":"J. Virol."},{"key":"ref_49","doi-asserted-by":"crossref","unstructured":"Knoops, K., Kikkert, M., Worm, S.H.E.V.D., Zevenhoven-Dobbe, J.C., Van Der Meer, Y., Koster, A.J., Mommaas, A.M., and Snijder, E.J. (2008). SARS-Coronavirus Replication Is Supported by a Reticulovesicular Network of Modified Endoplasmic Reticulum. PLoS Boil., 6.","DOI":"10.1371\/journal.pbio.0060226"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"5808","DOI":"10.1128\/JVI.07207-11","article-title":"Visualizing Coronavirus RNA Synthesis in Time by Using Click Chemistry","volume":"86","author":"Hagemeijer","year":"2012","journal-title":"J. Virol."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"625","DOI":"10.1080\/03079459708419239","article-title":"Molecular analysis of the 793\/B serotype of infectious bronchitis virus in Great Britain","volume":"26","author":"Adzhar","year":"1997","journal-title":"Avian Pathol."},{"key":"ref_52","doi-asserted-by":"crossref","unstructured":"Armesto, M., Cavanagh, D., and Britton, P. (2009). The Replicase Gene of Avian Coronavirus Infectious Bronchitis Virus Is a Determinant of Pathogenicity. PLoS ONE, 4.","DOI":"10.1371\/journal.pone.0007384"},{"key":"ref_53","doi-asserted-by":"crossref","unstructured":"Kim, Y., Oh, C., Shivanna, V., Hesse, R., and Chang, K.-O. (2017). Trypsin-independent porcine epidemic diarrhea virus US strain with altered virus entry mechanism. BMC Veter Res., 13.","DOI":"10.1186\/s12917-017-1283-1"},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"26","DOI":"10.1016\/j.virol.2013.09.018","article-title":"Mapping of the receptor-binding domain and amino acids critical for attachment in the spike protein of avian coronavirus infectious bronchitis virus","volume":"448","author":"Promkuntod","year":"2014","journal-title":"Virology"},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"1209","DOI":"10.1099\/vir.0.81651-0","article-title":"Sialic acid is a receptor determinant for infection of cells by avian Infectious bronchitis virus","volume":"87","author":"Winter","year":"2006","journal-title":"J. Gen. Virol."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"45","DOI":"10.1637\/0005-2086(2007)051[0045:HSIASA]2.0.CO;2","article-title":"Heparan Sulfate Is a Selective Attachment Factor for the Avian Coronavirus Infectious Bronchitis Virus Beaudette","volume":"51","author":"Madu","year":"2007","journal-title":"Avian Dis."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"8903","DOI":"10.1128\/JVI.05112-11","article-title":"Binding of Avian Coronavirus Spike Proteins to Host Factors Reflects Virus Tropism and Pathogenicity","volume":"85","author":"Wickramasinghe","year":"2011","journal-title":"J. Virol."},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"12552","DOI":"10.1128\/JVI.01890-13","article-title":"Middle East Respiratory Syndrome Coronavirus Infection Mediated by the Transmembrane Serine Protease TMPRSS2","volume":"87","author":"Shirato","year":"2013","journal-title":"J. Virol."},{"key":"ref_59","unstructured":"Saif, Y.M., Barnes, H.J., Glisson, J.R., Fadly, A.M., McDougald, L.R., and Swayne, D.E. (2003). Infectious bronchitis. Diseases of Poultry, Iowa State University Press."},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"218","DOI":"10.1016\/j.virol.2016.08.030","article-title":"Polymorphisms in the S1 spike glycoprotein of Arkansas-type infectious bronchitis virus (IBV) show differential binding to host tissues and altered antigenicity","volume":"498","author":"Leyson","year":"2016","journal-title":"Virology"},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"2700","DOI":"10.1128\/jvi.69.4.2700-2703.1995","article-title":"Impairment of multicycle influenza virus growth in Vero (WHO) cells by loss of trypsin activity","volume":"69","author":"Kaverin","year":"1995","journal-title":"J. Virol."},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"12","DOI":"10.1016\/j.virol.2019.04.014","article-title":"S gene and 5a accessory gene are responsible for the attenuation of virulent infectious bronchitis coronavirus","volume":"533","author":"Zhao","year":"2019","journal-title":"Virology"},{"key":"ref_63","doi-asserted-by":"crossref","first-page":"1749","DOI":"10.1007\/s00705-011-1044-6","article-title":"Receptor-bound porcine epidemic diarrhea virus spike protein cleaved by trypsin induces membrane fusion","volume":"156","author":"Park","year":"2011","journal-title":"Arch. Virol."},{"key":"ref_64","doi-asserted-by":"crossref","first-page":"7872","DOI":"10.1128\/JVI.00464-11","article-title":"Role of Proteases in the Release of Porcine Epidemic Diarrhea Virus from Infected Cells","volume":"85","author":"Shirato","year":"2011","journal-title":"J. Virol."},{"key":"ref_65","doi-asserted-by":"crossref","unstructured":"Cheng, J., Zhao, Y., Xu, G., Zhang, K., Jia, W., Sun, Y., Zhao, J., Xue, J., Hu, Y., and Zhang, G.-Z. (2019). The S2 Subunit of QX-type Infectious Bronchitis Coronavirus Spike Protein Is an Essential Determinant of Neurotropism. Viruses, 11.","DOI":"10.3390\/v11100972"},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"87","DOI":"10.1186\/s12985-016-0544-0","article-title":"Middle East Respiratory Syndrome Coronavirus (MERS-CoV) origin and animal reservoir","volume":"13","author":"Mohd","year":"2016","journal-title":"Virol. J."},{"key":"ref_67","doi-asserted-by":"crossref","first-page":"4211","DOI":"10.1128\/JVI.80.9.4211-4219.2006","article-title":"Animal Origins of the Severe Acute Respiratory Syndrome Coronavirus: Insight from ACE2-S-Protein Interactions","volume":"80","author":"Li","year":"2006","journal-title":"J. Virol."},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"255","DOI":"10.1038\/s41586-018-0010-9","article-title":"Fatal swine acute diarrhoea syndrome caused by an HKU2-related coronavirus of bat origin","volume":"556","author":"Zhou","year":"2018","journal-title":"Nature"},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"601","DOI":"10.1007\/s00253-011-3569-8","article-title":"Trypsin promotes efficient influenza vaccine production in MDCK cells by interfering with the antiviral host response","volume":"93","author":"Seitz","year":"2012","journal-title":"Appl. Microbiol. Biotechnol."}],"container-title":["Viruses"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4915\/12\/10\/1102\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T10:14:43Z","timestamp":1760177683000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4915\/12\/10\/1102"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,9,29]]},"references-count":69,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2020,10]]}},"alternative-id":["v12101102"],"URL":"https:\/\/doi.org\/10.3390\/v12101102","relation":{},"ISSN":["1999-4915"],"issn-type":[{"value":"1999-4915","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,9,29]]}}}