Expression of Influenza nucleoprotein in microalgal chloroplasts. Application for generation of edible vaccines

Abstract

The World Health Organization (WHO), the National Institutes of Health, and UNESCO have stressed the need for a new generation of low-cost vaccines to promote vaccination programs in the poorest regions of the world. They especially emphasized the need for heat-stable vaccines to avoid the considerable expense in maintaining the cold-chain during production and distribution, and needle-free formulations to reduce the risk of opportunistic contamination as well as the need for qualified personnel. Influenza is one of the most significant diseases worldwide, being implicated in 4 major pandemics and the causative agent of ongoing yearly epidemics. Every year Influenza A viruses initiate outbreaks of respiratory tract infection resulting in unacceptable mortality especially in the immuno-compromised persons and the +65's, and morbidity that impacts significantly in workplace economies. Influenza H1N1 nucleoprotein (NP) sequence, codon optimized for C.reinhardtii chloroplast-specific regulatory sequences was designed as an epitope to induce immunity against Influenza. Wild type C. reinhardtii chloroplast genomes were transformed by NP transgene expressed under control of chloroplast-specific regulatory sequences, promotors and terminators. A large region of the tscA gene was used for integration into the chloroplast genome via homological recombination. Integration and expression of NP epitope containing transgene in C. reinhardtii chloroplast genomes was confirmed phenotypically by antibiotic spectinomycin selection and using a set of molecular biology techiques such as PCR, RT-PCR, DNA sequencing and ELISA. It is considered further research would be advantageous to confirm vaccine candidacy