Abstract
The use of transgenic plants to produce novel products has great biotechnological potential as the relatively inexpensive inputs of light, water, and nutrients are utilised in return for potentially valuable bioactive metabolites, diagnostic proteins and vaccines. Extensive research is ongoing in this area internationally with the aim of producing plant-made vaccines of importance for both animals and humans. Vaccine purification is generally regarded as being integral to the preparation of safe and effective vaccines for use in humans. However, the use of crude plant extracts for animal immunisation may enable plant-made vaccines to become a cost-effective and efficacious approach to safely immunise large numbers of farm animals against diseases such as avian influenza. Since the technology associated with genetic transformation and large-scale propagation is very well established in Nicotiana, the genus has attributes well-suited for the production of plant-made vaccines. However the presence of potentially toxic alkaloids in Nicotiana extracts impedes their use as crude vaccine preparations. In the current study we describe a Nicotiana tabacum and N. glauca hybrid that expresses the HA glycoprotein of influenza A in its leaves but does not synthesize alkaloids. We demonstrate that injection with crude leaf extracts from these interspecific hybrid plants is a safe and effective approach for immunising mice. Moreover, this antigen-producing alkaloid-free, transgenic interspecific hybrid is vigorous, with a high capacity for vegetative shoot regeneration after harvesting. These plants are easily propagated by vegetative cuttings and have the added benefit of not producing viable pollen, thus reducing potential problems associated with bio-containment. Hence, these Nicotiana hybrids provide an advantageous production platform for partially purified, plant-made vaccines which may be particularly well suited for use in veterinary immunization programs.
Highlights
Transgenic plants are gaining acceptance as a platform for the production of affordable recombinant proteins in the pharmaceutical industries [1,2]
The mean HA content for each construct was calculated from these independent Nt LAFC-HA lines from the enzymelinked immunosorbant assay (ELISA) analysis, and demonstrated that the Nt LAFC-HA plants containing the 4OCS-DMas promoter construct, pDAB4493, produced significantly higher HA levels (p,0.05) compared to those produced by plants harbouring the cassava vein mosaic virus (CsVMV) promoter construct, pDAB4492 (Figure 2a)
The potential for a pandemic outbreak has led to classification of H5N1 AIV in List A of the l’Office International des Epizooties (OIE; World Organization for Animal Health)
Summary
Transgenic plants are gaining acceptance as a platform for the production of affordable recombinant proteins in the pharmaceutical industries [1,2]. Advantages associated with using a plant expression system include the ability to utilize gene splicing to produce multi-antigen vaccines and the decreased risk of product contamination with human or animal pathogens. Plant-made heat stable vaccines can be shipped and stored without refrigeration and have the potential to be produced in edible plant organs and delivered orally without the requirement for recombinant protein purification. Avian influenza virus (AIV) can infect a variety of avian and mammalian species including domestic poultry and humans, and poses a serious international pandemic threat [8]. The first plant-made vaccine (PMV) to be commercially licensed was a partially purified, injectable poultry vaccine against Newcastle Disease Virus (NDV) [9]. As current regulations allow crude or partially purified veterinary vaccine formulations to be administered by injection
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