Comparative genomics of Bacillus anthracis A and B-clades reveals genetic variation in genes responsible for spore germination

Bacillus anthracis, the causative agent of anthrax, is composed of three genetic clades (A, B, and C). Clade-A is the most common and distributed worldwide, B-clade has a narrow geographic distribution, and C-clade is rare. South Africa's Kruger National Park (KNP) has high diversity of B. anthracis, with strains from A and B clades described from its northernmost region, Pafuri. We employed whole genome sequencing to investigate the genomic diversity of B. anthracis strains isolated from animal carcasses (n = 34) during the 2012-2015 outbreaks. Whole-genome single-nucleotide polymorphism (wgSNP) analysis assigned the 2012-2015 B. anthracis genomes to the A-clade branch, distributed across the branch's two minor sub-clades A.Br.005/006. Additionally, pan-genomic analysis distinguished the A- and B-clade genomes, identifying unique accessory genes. Notable genetic differences include the biosynthetic spore cell wall genes; long-chain fatty acid CoA ligases (FaD13), Bacillus collagen-like protein of anthracis (BclA) involved in the exosporium germination, as well as a truncated murein DD-endopeptidase (mepH) found in the pXO2 plasmid of the B-clade strains. The tryptophan synthase subunit alpha gene (trpA), which results in a pseudogene in B-clade genomes separates the A- and B-clade genomes. These differences in biosynthetic cell wall genes suggest variation in adaptability or cell growth of the B-clade strains in the environment, further influenced by the truncation of the trpA gene involved in spore germination. The A.Br.005/006-clade strains in KNP exhibit higher genetic diversity, which may enhance their resilience to environmental stressors. In contrast, the KNP B-clade (B.Br.001/002) strains show limited genetic variation, potentially reducing their adaptability. This pattern is evident through whole-genome SNP analysis and pan-genomics investigating the evolution of B. anthracis.