This report focuses on a systematic search for Cry proteins in Bacillus spp. other than B. thuringiensis by analyzing reported Bacillus spp. genomes, using conserved sequences from the C-terminal half of reported Cry proteins in hidden Markov model profiles. A high-throughput model based on the use of HMMER and CD-HIT tools was designed, which identified Cry proteins. This model was used on 857 reported Bacillus spp. genomes, where 174 Cry protein sequences were identified, mostly, as expected, in B. thuringiensis genomes but, interestingly, 42 were identified on other species. Despite including 89 species of Bacillus in the HMMER analysis, Cry protein sequences were found only in genomes from species within the B. cereus group. According to the species registered at the NCBI database containing each genome, this group was formed by 18 non-B. thuringiensis strains. However, when sequences in those genomes were analyzed by multilocus sequence typing, the number of non-B. thuringiensis strains increased to 39, indicating that as many as 119 Cry protein sequences were found in four non-B. thuringiensis species. Therefore, dispersion of Cry proteins is much wider and frequent than previously thought, questioning its role in nature.

This chapter analyses the different strategies that have been used for the expression and production of heterologous proteins in plants and the impact they have had in the field of production of vaccines against the Human papillomavirus.

Rotavirus is the most common cause of severe diarrhea in infants and children worldwide and is responsible for about 215,000 deaths annually. Over 85% of these deaths originate in low-income/developing countries in Asia and Africa. Therefore, it is necessary to explore the development of vaccines that avoid the use of "living" viruses and furthermore, vaccines that have viral antigens capable of generating powerful heterotypic responses. Our strategy is based on the expression of the fusion of the anti-DEC205 single-chain variable fragment (scFv) coupled by an OLLAS tag to a viral protein (VP6) of Rotavirus in Nicotiana plants. It was possible to express transiently in N. benthamiana and N. sylvestris a recombinant protein consisting of the single chain variable fragment linked by an OLLAS tag to the VP6 protein. The presence of the recombinant protein, which had a molecular weight of approximately 75 kDa, was confirmed by immunodetection, in both plant species and in both cellular compartments (cytoplasm and apoplast) where it was expressed. In addition, the recombinant protein was modeled, and it was observed that some epitopes of interest are exposed on the surface, which could favor their immunogenic response.

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