Novel Production of Bovine Papillomavirus Pseudovirions in Tobacco Plants.

Inga Hitzeroth,Inge Pietersen,Albertha Van Zyl,Edward Rybicki

doi:10.3390/pathogens9120996

Inga Hitzeroth, Inge Pietersen + Show 2 more

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https://doi.org/10.3390/pathogens9120996

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Abstract

Vaccine efficacy requires the production of neutralising antibodies which offer protection against the native virus. The current gold standard for determining the presence of neutralising antibodies is the pseudovirion-based neutralisation assay (PBNA). PBNAs utilise pseudovirions (PsVs), structures which mimic native virus capsids, but contain non-viral nucleic material. PsVs are currently produced in expensive cell culture systems, which limits their production, yet plant expression systems may offer cheaper, safer alternatives. Our aim was to determine whether plants could be used for the production of functional PsVs of bovine papillomavirus 1 (BPV1), an important causative agent of economically damaging bovine papillomas in cattle and equine sarcoids in horses and wild equids. BPV1 capsid proteins, L1 and L2, and a self-replicating reporter plasmid were transiently expressed in Nicotiana benthamiana to produce virus-like particles (VLPs) and PsVs. Strategies to enhance particle yields were investigated and optimised protocols were established. The PsVs’ ability to infect mammalian cells and express their encapsidated reporter genes in vitro was confirmed, and their functionality as reagents in PBNAs was demonstrated through their neutralisation by several different antibodies. This is the first report of BPV PsVs expressed in plants and demonstrates the potential for the development of therapeutic veterinary vaccines in planta.

Highlights

Bovine papillomaviruses (BPVs) are a group of epitheliotropic viruses which are of significant importance, both as etiological agents of veterinary diseases and as long-standing models for the study of human and other papillomaviruses (PVs) [1]
To first establish that bovine papillomavirus 1 (BPV1) capsid proteins expressed in N. benthamiana were able to self-assemble into higher order structures and survive purification with techniques developed for human papillomaviruses (HPVs) [50], we performed infiltrations of L1-only and L1 + L2 for the production of virus-like particles (VLPs)
BPV1 capsid proteins, L1 and L2, were codon-optimised for expression in N. benthamiana, as optimisation for tobacco was shown in previous studies to increase protein expression [47]

Summary

Introduction

Bovine papillomaviruses (BPVs) are a group of epitheliotropic viruses which are of significant importance, both as etiological agents of veterinary diseases and as long-standing models for the study of human and other papillomaviruses (PVs) [1]. Autogenous vaccines are still most commonly used for prophylaxis, and while a growing body of evidence indicates that these vaccines have some therapeutic effects in cattle [16,17,18] and equids [8,9,19], these vaccines carry risks of reversion/recombination and exposing the animal to blood-borne diseases [20]. They are further limited by the fact that they require large amounts of wart tissue and Pathogens 2020, 9, 996; doi:10.3390/pathogens9120996 www.mdpi.com/journal/pathogens

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