Abstract
BackgroundBeak 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.MethodsThe 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.ResultsIn 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.ConclusionsThis 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.
Highlights
Beak and feather disease virus (BFDV) is an important disease causing agent affecting psittacines
Optimisation of BFDV capsid protein (CP) expression in N. benthamiana To determine the best conditions for expression, various densities of Agrobacterium suspensions were tested by plant syringe infiltration, and leaves were harvested on days 1, 3, 5 and 7
Gene amplification had a mixed effect on expression: accumulation in the cytoplasm and chloroplasts appeared to be improved; accumulation in the ER decreased, and there was no change when the CP was secreted to the apoplast
Summary
Beak and feather disease virus (BFDV) is an important disease causing agent affecting psittacines. Beak and feather disease virus (BFDV; family Circoviridae, genus Circovirus) is one of the most common disease agents to infect psittacines [1]. The virus infects both wild and captive birds, and has been detected in at least 10% of psittacine species [2,3,4]. It is the causative agent of psittacine beak and feather disease (PBFD), which can present. The likelihood that a single-strain vaccine would be successful is high, due to the absence of obvious serotypes which indicates conservation of major epitopes between isolates: there is a high probability of crossisolate protection [17, 18]
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