Abstract
Rotavirus is the leading cause of severe diarrheal disease among newborns. Plant-based rotavirus vaccines have been developed in recent years and have been proven to be effective in animal models. In the present study, we report a bivalent vaccine candidate expressing rotavirus subunits VP6 and NSP4 fused with the adjuvant subunit B of E. coli heat-labile enterotoxin (LTB) in maize seeds. The RT-PCR and Western blot results showed that VP6 and LTB-NSP4 antigens were expressed and accumulated in maize seeds. The expression levels were as high as 0.35 and 0.20% of the total soluble protein for VP6 and LTB-NSP4, respectively. Oral administration of transgenic maize seeds successfully stimulated systemic and mucosal responses, with high titers of serum IgG and mucosal IgA antibodies, even after long-term storage. This study is the first to use maize seeds as efficient generators for the development of a bivalent vaccine against rotavirus.
Highlights
As transgenic technology advanced over the past decades, plant-based expression systems have experienced rapid development
A GPGP hinge region was fused between the LTB and non-structural protein 4 (NSP4) peptide to improve the flexibility of the protein (Figure 1A)
VP6 and LTB-NSP4 proteins accumulated in the seeds and were hardly detected in the other organs, while the internal control β-actin was ubiquitously expressed (Figure 4B). These results demonstrate that VP6 and LTB-NSP4 genes are expressed efficiently and are localized in the transgenic maize seeds
Summary
As transgenic technology advanced over the past decades, plant-based expression systems have experienced rapid development. Antibodies, cytokines and other functional proteins and growth factors have been expressed in tobacco, potatoes, tomatoes, maize and rice (da Cunha et al, 2014; Streatfield et al, 2015). Viral subunits produced by plants are promising vaccine candidates due to the following advantages: large amounts of antigen can be produced at relatively low cost and are scaled up to field levels; they are safe and are not contaminated with human viruses; post-translational modification is efficient; and oral administration of edible plantbased vaccines can induce mucosal immune responses, which is the primary barrier against viral infection, and systemic immune responses (Goldstein and Thomas, 2004). The first plantbased vaccine was developed in 1992 by Mason and colleagues, who expressed the hepatitis B surface antigen (HBsAg) in tobacco leaves (Mason et al, 1992).
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