Current state-of-the-art in the use of plants for the production of recombinant vaccines against infectious bursal disease virus.

Infectious bursal disease is a widely spread threatening contagious viral infection of chickens that induces major damages to the Bursa of Fabricius and leads to severe immunosuppression in young birds causing significant economic losses for poultry farming. The etiological agent is the infectious bursal disease virus (IBDV), a non-enveloped virus belonging the family of Birnaviridae. At present, the treatment against the spread of this virus is represented by vaccination schedules mainly based on inactivated or live-attenuated viruses. However, these conventional vaccines present several drawbacks such as insufficient protection against very virulent strains and the impossibility to differentiate vaccinated animals from infected ones. To overcome these limitations, in the last years, several studies have explored the potentiality of recombinant subunit vaccines to provide an effective protection against IBDV infection. In this review, we will give an overview of these novel types of vaccines with special emphasis on current state-of-the-art in the use of plants as "biofactories" (plant molecular farming). In fact, plants have been thoroughly and successfully characterized as heterologous expression systems for the production of recombinant proteins for different applications showing several advantages compared with traditional expression systems (Escherichia coli, yeasts and insect cells) such as absence of animal pathogens in the production process, improved product quality and safety, reduction of manufacturing costs, and simplified scale-up.

Functional characterization of a plant-produced infectious bursal disease virus antigen fused to the constant region of avian IgY immunoglobulins.

Infectious bursal disease virus (IBDV) is the cause of an economically important highly contagious disease of poultry, and vaccines are regarded as the most beneficial interventions for its prevention. In this study, plants were used to produce a recombinant chimeric IBDV antigen for the formulation of an innovative subunit vaccine. The fusion protein (PD-FcY) was designed to combine the immunodominant projection domain (PD) of the viral structural protein VP2 with the constant region of avian IgY (FcY), which was selected to enhance antigen uptake by avian immune cells. The gene construct encoding the fusion protein was transiently expressed in Nicotiana benthamiana plants and an extraction/purification protocol was set up, allowing to reduce the contamination by undesired plant compounds/proteins. Mass spectrometry analysis of the purified protein revealed that the glycosylation pattern of the FcY portion was similar to that observed in native IgY, while in vitro assays demonstrated the ability of PD-FcY to bind to the avian immunoglobulin receptor CHIR-AB1. Preliminary immunization studies proved that PD-FcY was able to induce the production of protective anti-IBDV-VP2 antibodies in chickens. In conclusion, the proposed fusion strategy holds promises for the development of innovative low-cost subunit vaccines for the prevention of avian viral diseases.

The expression in plants of an engineered VP2 protein of Infectious Bursal Disease Virus induces formation of structurally heterogeneous particles that protect from a very virulent viral strain.

Infectious Bursal Disease Virus (IBDV), the etiological agent of Gumboro disease, causes mortality and immunosuppression in chickens and major losses to poultry industry worldwide. The IBDV major capsid protein VP2 is considered the best candidate for the production of novel subunit vaccines. This structural protein contains the major conformational epitopes responsible for the induction of IBDV neutralizing antibodies in chickens and has been demonstrated able to form supramolecular structures in yeast and insect cells. The aim of this study was to express an engineered version of the VP2 protein (His-pVP2) to verify its ability to self-assemble into virus-like particles in plants. The recombinant VP2 was transiently expressed by agroinfiltration in Nicotiana benthamiana and transmission electron microscopy of sucrose density gradient fractions revealed the presence of a mixed population of differently shaped particles ranging from spherical capsids, with a diameter between ~25 and ~70 nm, to tubular structures, with variable length (from 100 to 400 nm). The recombinant VP2-based particles when used for the intramuscular immunization of specific-pathogen-free chicks resulted able to induce the production of anti-IBDV specific antibodies at titers comparable to those induced by a commercial vaccine. Moreover, all the immunized birds survived to the challenge with a Moroccan very virulent IBDV strain with no major histomorphological alterations of the Bursa of Fabricius, similarly to what obtained with the commercial inactivated vaccine.