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Development of a potential challenge model and plant-produced vaccine candidate for beak and feather disease virus
Psittacine beak and feather disease (PBFD), the most prevalent viral disease affecting psittacines, is caused by beak and feather disease virus (BFDV). An outbreak of the disease has been reported in wild endangered Cape parrots (Poicephalus robustus), which is endemic to South Africa. No treatment...
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| Format: | Thesis |
| Language: | English |
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Department of Molecular and Cell Biology
2015
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| _version_ | 1867613206628794368 |
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| access_status_str | Open Access |
| author | Regnard, Guy Louis |
| author2 | Hitzeroth, Inga |
| author_browse | Hitzeroth, Inga Regnard, Guy Louis |
| author_facet | Hitzeroth, Inga Regnard, Guy Louis |
| author_sort | Regnard, Guy Louis |
| collection | Thesis |
| description | Psittacine beak and feather disease (PBFD), the most prevalent viral disease affecting psittacines, is caused by beak and feather disease virus (BFDV). An outbreak of the disease has been reported in wild endangered Cape parrots (Poicephalus robustus), which is endemic to South Africa. No treatment or vaccine is commercially available. In this study, an investigation into the outbreak was undertaken. BFDV diversity was assessed and viral load and clinical signs correlated. A plant-produced BFDV subunit vaccine was produced in parallel with a corresponding challenge model. Cape parrots were assessed and 53 blood samples collected. Viral load was determined using quantitative real-time PCR (qPCR), and 22 BFDV full-length genome sequences acquired to infer phylogenetic relatedness. The capsid gene (cp) was optimised for transient Agrobacterium-mediated expression in whole-plant Nicotiana benthamiana (N. benthamiana). Virus-like particles (VLPs) were purified and analysed using transmission electron microscopy. Virions from a Palm cockatoo (Probosciger aterrimus) were purified and a BFDV dsDNA molecular clone was synthesised and replication assessed in 293TT mammalian cells and N. benthamiana using rolling circle replication and qPCR. Two distinct BFDV phylogenetic clusters were reported for Cape parrots, and a direct correlation was seen between viral load in the blood and clinical signs in PBFD-afflicted birds. The CP was successfully expressed in N. benthamiana, and increased through optimisation of Agrobacterium infiltration density and the inclusion of the NSs silencing suppressor. The CP formed VLPs, which were shown to be morphologically similar to infectious virions. The dsDNA molecular clone was shown to replicate autonomously in mammalian 293TT cells, and in plants with the assistance of the Bean yellow dwarf virus replication associated protein (Rep). BFDV genetic diversity in Cape parrots highlights the importance of ensuring new strains are not inadvertently introduced into the wild. This is the first systematic investigation of virus diversity in Cape parrots and assessment of BFDV viral load in a wild psittacine population. The CP was successfully produced in planta and presence of VLPs suggests the possibility of developing pseudovirions. This is the first reported replication of BFDV in tissue culture, and will greatly expand the scope of available research. |
| format | Thesis |
| id | oai:open.uct.ac.za:11427/15690 |
| institution | University of Cape Town (South Africa) |
| language | eng |
| last_indexed | 2026-06-10T12:32:27.580Z |
| license_str | Not specified — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UCTD — University of Cape Town Open Access Repository |
| publishDate | 2015 |
| publishDateRange | 2015 |
| publishDateSort | 2015 |
| publisher | Department of Molecular and Cell Biology |
| publisherStr | Department of Molecular and Cell Biology |
| record_format | dspace |
| source_str | UCTD — University of Cape Town Open Access Repository |
| spelling | oai:open.uct.ac.za:11427/15690 Development of a potential challenge model and plant-produced vaccine candidate for beak and feather disease virus Regnard, Guy Louis Hitzeroth, Inga Rybicki, Edward P Molecular and Cell Biology Psittacine beak and feather disease (PBFD), the most prevalent viral disease affecting psittacines, is caused by beak and feather disease virus (BFDV). An outbreak of the disease has been reported in wild endangered Cape parrots (Poicephalus robustus), which is endemic to South Africa. No treatment or vaccine is commercially available. In this study, an investigation into the outbreak was undertaken. BFDV diversity was assessed and viral load and clinical signs correlated. A plant-produced BFDV subunit vaccine was produced in parallel with a corresponding challenge model. Cape parrots were assessed and 53 blood samples collected. Viral load was determined using quantitative real-time PCR (qPCR), and 22 BFDV full-length genome sequences acquired to infer phylogenetic relatedness. The capsid gene (cp) was optimised for transient Agrobacterium-mediated expression in whole-plant Nicotiana benthamiana (N. benthamiana). Virus-like particles (VLPs) were purified and analysed using transmission electron microscopy. Virions from a Palm cockatoo (Probosciger aterrimus) were purified and a BFDV dsDNA molecular clone was synthesised and replication assessed in 293TT mammalian cells and N. benthamiana using rolling circle replication and qPCR. Two distinct BFDV phylogenetic clusters were reported for Cape parrots, and a direct correlation was seen between viral load in the blood and clinical signs in PBFD-afflicted birds. The CP was successfully expressed in N. benthamiana, and increased through optimisation of Agrobacterium infiltration density and the inclusion of the NSs silencing suppressor. The CP formed VLPs, which were shown to be morphologically similar to infectious virions. The dsDNA molecular clone was shown to replicate autonomously in mammalian 293TT cells, and in plants with the assistance of the Bean yellow dwarf virus replication associated protein (Rep). BFDV genetic diversity in Cape parrots highlights the importance of ensuring new strains are not inadvertently introduced into the wild. This is the first systematic investigation of virus diversity in Cape parrots and assessment of BFDV viral load in a wild psittacine population. The CP was successfully produced in planta and presence of VLPs suggests the possibility of developing pseudovirions. This is the first reported replication of BFDV in tissue culture, and will greatly expand the scope of available research. 2015-12-08T11:42:54Z 2015-12-08T11:42:54Z 2015 Doctoral Thesis Doctoral PhD http://hdl.handle.net/11427/15690 eng application/pdf Department of Molecular and Cell Biology Faculty of Science University of Cape Town |
| spellingShingle | Molecular and Cell Biology Regnard, Guy Louis Development of a potential challenge model and plant-produced vaccine candidate for beak and feather disease virus |
| thesis_degree_str | Doctoral |
| title | Development of a potential challenge model and plant-produced vaccine candidate for beak and feather disease virus |
| title_full | Development of a potential challenge model and plant-produced vaccine candidate for beak and feather disease virus |
| title_fullStr | Development of a potential challenge model and plant-produced vaccine candidate for beak and feather disease virus |
| title_full_unstemmed | Development of a potential challenge model and plant-produced vaccine candidate for beak and feather disease virus |
| title_short | Development of a potential challenge model and plant-produced vaccine candidate for beak and feather disease virus |
| title_sort | development of a potential challenge model and plant produced vaccine candidate for beak and feather disease virus |
| topic | Molecular and Cell Biology |
| url | http://hdl.handle.net/11427/15690 |
| work_keys_str_mv | AT regnardguylouis developmentofapotentialchallengemodelandplantproducedvaccinecandidateforbeakandfeatherdiseasevirus |