Recombinant expression of beak and feather disease virus capsid protein and assembly of virus-like particles in Nicotiana benthamiana

dc.contributor.authorRegnard, Guy L
dc.contributor.authorRybicki, Edward P
dc.contributor.authorHitzeroth, Inga I
dc.date.accessioned2017-09-18T08:12:56Z
dc.date.available2017-09-18T08:12:56Z
dc.date.issued2017-09-11
dc.date.updated2017-09-17T03:22:19Z
dc.description.abstractBackground: Beak and feather disease virus (BFDV) is an important disease causing agent affecting psittacines. BFDV is highly infectious and can present as acute, chronic or subclinical disease. The virus causes immunodeficiency and is often associated with secondary infections. No commercial vaccine is available and yields of recombinant BFDV capsid protein (CP) expressed in insect cells and bacteria are yet to be seen as commercially viable, although both systems produced BFDV CP that could successfully assemble into virus-like particles (VLPs). Plants as expression systems are increasingly becoming favourable for the production of region-specific and niche market products. The aim of this study was to investigate the formation and potential for purification of BFDV VLPs in Nicotiana benthamiana. Methods: The BFDV CP was transiently expressed in N. benthamiana using an Agrobacterium-mediated system and plant expression vectors that included a bean yellow dwarf virus (BeYDV)-based replicating DNA vector. Plant-produced BFDV CP was detected using immunoblotting. VLPs were purified using sucrose cushion and CsCl density gradient centrifugation and visualised using transmission electron microscopy. Results: In this study we demonstrate that the BFDV CP can be successfully expressed in N. benthamiana, albeit at relatively low yield. Using a purification strategy based on centrifugation we demonstrated that the expressed CP can self-assemble into VLPs that can be detected using electron microscopy. These plant-produced BFDV VLPs resemble those produced in established recombinant expression systems and infectious virions. It is possible that the VLPs are spontaneously incorporating amplicon DNA produced from the replicating BeYDV plant vector. Conclusions: This is the first report of plant-made full-length BFDV CP assembling into VLPs. The putative pseudovirions could be used to further the efficacy of vaccines against BFDV.
dc.identifier.apacitationRegnard, G. L., Rybicki, E. P., & Hitzeroth, I. I. (2017). Recombinant expression of beak and feather disease virus capsid protein and assembly of virus-like particles in Nicotiana benthamiana. <i>Virology Journal</i>, http://hdl.handle.net/11427/25228en_ZA
dc.identifier.chicagocitationRegnard, Guy L, Edward P Rybicki, and Inga I Hitzeroth "Recombinant expression of beak and feather disease virus capsid protein and assembly of virus-like particles in Nicotiana benthamiana." <i>Virology Journal</i> (2017) http://hdl.handle.net/11427/25228en_ZA
dc.identifier.citationRegnard, G. L., Rybicki, E. P., & Hitzeroth, I. I. (2017). Recombinant expression of beak and feather disease virus capsid protein and assembly of virus-like particles in Nicotiana benthamiana. Virology Journal, 14(1), 174. DOI: 10.1186/s12985-017-0847-9.
dc.identifier.ris TY - Journal Article AU - Regnard, Guy L AU - Rybicki, Edward P AU - Hitzeroth, Inga I AB - Background: Beak and feather disease virus (BFDV) is an important disease causing agent affecting psittacines. BFDV is highly infectious and can present as acute, chronic or subclinical disease. The virus causes immunodeficiency and is often associated with secondary infections. No commercial vaccine is available and yields of recombinant BFDV capsid protein (CP) expressed in insect cells and bacteria are yet to be seen as commercially viable, although both systems produced BFDV CP that could successfully assemble into virus-like particles (VLPs). Plants as expression systems are increasingly becoming favourable for the production of region-specific and niche market products. The aim of this study was to investigate the formation and potential for purification of BFDV VLPs in Nicotiana benthamiana. Methods: The BFDV CP was transiently expressed in N. benthamiana using an Agrobacterium-mediated system and plant expression vectors that included a bean yellow dwarf virus (BeYDV)-based replicating DNA vector. Plant-produced BFDV CP was detected using immunoblotting. VLPs were purified using sucrose cushion and CsCl density gradient centrifugation and visualised using transmission electron microscopy. Results: In this study we demonstrate that the BFDV CP can be successfully expressed in N. benthamiana, albeit at relatively low yield. Using a purification strategy based on centrifugation we demonstrated that the expressed CP can self-assemble into VLPs that can be detected using electron microscopy. These plant-produced BFDV VLPs resemble those produced in established recombinant expression systems and infectious virions. It is possible that the VLPs are spontaneously incorporating amplicon DNA produced from the replicating BeYDV plant vector. Conclusions: This is the first report of plant-made full-length BFDV CP assembling into VLPs. The putative pseudovirions could be used to further the efficacy of vaccines against BFDV. DA - 2017-09-11 DB - OpenUCT DO - 10.1186/s12985-017-0847-9 DP - University of Cape Town J1 - Virology Journal LK - https://open.uct.ac.za PB - University of Cape Town PY - 2017 T1 - Recombinant expression of beak and feather disease virus capsid protein and assembly of virus-like particles in Nicotiana benthamiana TI - Recombinant expression of beak and feather disease virus capsid protein and assembly of virus-like particles in Nicotiana benthamiana UR - http://hdl.handle.net/11427/25228 ER - en_ZA
dc.identifier.urihttp://dx.doi.org/10.1186/s12985-017-0847-9
dc.identifier.urihttp://hdl.handle.net/11427/25228
dc.identifier.vancouvercitationRegnard GL, Rybicki EP, Hitzeroth II. Recombinant expression of beak and feather disease virus capsid protein and assembly of virus-like particles in Nicotiana benthamiana. Virology Journal. 2017; http://hdl.handle.net/11427/25228.en_ZA
dc.language.isoen
dc.publisherBioMed Central
dc.publisher.departmentDepartment of Molecular and Cell Biologyen_ZA
dc.publisher.facultyFaculty of Scienceen_ZA
dc.publisher.institutionUniversity of Cape Town
dc.rights.holderThe Author(s).
dc.rights.urihttps://creativecommons.org/licenses/by/4.0/
dc.sourceVirology Journal
dc.source.urihttps://virologyj.biomedcentral.com/
dc.subject.otherPlant-made pharmaceutical
dc.subject.otherBeak and feather disease virus
dc.subject.otherVirus-like particle
dc.subject.otherCircovirus
dc.subject.otherSubunit vaccine
dc.titleRecombinant expression of beak and feather disease virus capsid protein and assembly of virus-like particles in Nicotiana benthamiana
dc.typeJournal Article
uct.type.filetypeText
uct.type.filetypeImage
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