Genetic diversity in the protective antigen gene of Bacillus anthracis

Bacillus anthracis is a gram-positive spore-forming bacterium that causes t he disease anthrax, The anthrax toxin contains three components, including the protective antigen (PA), which binds to eucaryotic cell surface recepto rs and mediates the transport of toxins into the cell. In this study, the e ntire 2,294-nucleotide protective antigen gene (pag) was sequenced from 26 of the most diverse B. anthracis strains to identify potential variation in the toxin and to further our understanding of B. anthracis evolution. Five point mutations, three synonymous and two missense, were identified. These differences correspond to six different haploid types, which translate int o three different amino acid sequences. The two amino acid changes were sho wn to be located in an area near a highly antigenic region critical to leth al factor binding. Nested primers were used to amplify and sequence this sa me region of pag from necropsy samples taken from victims of the 1979 Sverd lovsk incident. This investigation uncovered five different alleles among t he strains present in the tissues, including two not seen in the 26-sample survey. One of these two alleles included a novel missense mutation, again located just adjacent to the highly antigenic region. Phylogenetic (cladist ic) analysis of the pag corresponded with previous strain grouping based on chromosomal variation, suggesting that plasmid evolution in B, anthracis h as occurred with little or no horizontal transfer between the different str ains.