Ms Molina defended her PhD thesis, The plastidial transformation of tobacco for the production of an amphigenic vaccine against parvovirus canino at the Public University of Navarra.

The agricultural engineer Andrea Molina Azcona has developed a vaccine in the leaves of the tobacco plant that combats parvovirus canino, a virus that affects dog pups. It involves a vaccine produced by the transformation of the chloroplast in the plant, given the high yield – from just one plant some 500-vaccine units can be produced and so may well be commercially viable. Ms Molina defended her PhD thesis, The plastidial transformation of tobacco for the production of an amphigenic vaccine against parvovirus canino at the Public University of Navarra. An alternative vaccine Parvovirus is a disease that provokes haemorrhaged gastro-enteritis and may result in myocarditis. The youngest pups are those most prone to shock and death due to the infirmity; it may be only a question of days after diagnosis. Specifically in Navarre, there are usually one or two outbreaks annually, one just after Christmas, coinciding with the relocation of pups from one place to another. The high morbidity and death rates associated with this disease may be considerably reduced with prophylactic treatment and vaccines. These vaccines are mostly based on attenuated viruses and, so that they are not so virulent, they are subjected to a treatment with heat or some chemical product. Nevertheless, the vaccine developed by Andrea Molina was made with proteins produced in genetically modified crops, specifically in the tobacco plant, which is one that is most easily transformed. Moreover, for the production of the necessary proteins, instead of modifying the DNA of the nucleus of the plant cells, it is the chloroplast DNA that is transformed, by introducing the gene that has the desired protein. The transformation of the plastidial genome, instead of the more usual nuclear genome, has a series of important advantages for biotechnological applications. First, much more protein is obtained – a yield of some 31%, i.e. of every 100 proteins produced, 31 are of the type we are interested in, which is a very high proportion. Moreover, it has other, environmental advantages such as, for example, the fact that tobacco pollen does not contain chloroplasts, so that, when the tobacco produces flowers, the dispersion of the genetic modification is avoided. This research specifically developed peptide 2L21 in the chloroplasts of the tobacco plant, a compound that confers protection to dogs against virulent canine parvovirus (CPV), fused with two co-proteins: the B sub unit of the cholera toxin (CTB) and the green fluorescent protein (GFP). Thus, apart from obtaining high yield levels of the fusion protein, it was observed that the two fusion proteins were stable both in young leaves as well as in old and that the estimated average life, as determined by marking, was greater than 48 hours in both cases. Marketing possibilities The lyophilisation of the pulverised leaves was a simple method of post-harvest treatment, after seven months of storage at ambient temperature, and which did not produce significant loss of the recombinant protein. As regards the marketing of the product, the high yield of the amphigenes in the chloroplast achieved may reduce the quantity of vegetable matter required for vaccination (100 mg for one 500 mg dose of antigens) and thus enables the encapsulation of the lyophilised material or the production of pills. Vaccines based on chloroplastic transformation in plants may well be commercially viable. For the moment, the vaccine has been used in trials on mice, with good results with intraperitoneal administration. For next year it is proposed to vaccinate pups with the collaboration of the Veterinary Faculty at the Universidad Complutense in Madrid.

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