Because of the inconsistent efficacy of the current TB vaccine (BCG), the development of improved TB vaccines and TB immunization strategies is an international research priority. Oral vaccines should permit the design of more convenient immunization strategies and allow the implementation of universal vaccination programs throughout the world. Recent advances in plant molecular biology has strongly suggested that oral vaccines, including an oral tuberculosis preparation, can be produced in edible plants. To develop anti-tuberculosis edible vaccines, we have modified two plant expression vectors. The genes encoding the immunogenic proteins - MPT64 and Esat-6 - have been cloned into these vectors and the recombinants have been transformed into the plant pathogen Agrobacterium tumefaciens. In addition, we have generated Agrobacterium transformants with vectors that encode cholera toxin subunit B - MPT64 fusion proteins. PCR analyses has demonstrated that infection with the recombinant Agrobacterium produced transformed Arabidopsis (model plant system) and tomato plants. Moreover, low levels of expression of both the MPT64 and the cholera toxin fusion proteins were detected in a few tomato plants. However, although mice fed tomatoes spiked with cholera toxin subunit B elicited an anti-cholera toxin response, immune responses to MPT64 were not detected in mice fed recombinant tomatoes expressing the mycobacterial protein. We are currently evaluating procedures to increase the expression of mycobacterial antigens in plant cells. Apparently higher foreign protein concentrations within transformed plants will be needed to induce appropriate immune responses after oral immunization.
