Molecular diversity of Bacillus anthracis in the Netherlands: investigating the relationship to the worldwide population using whole-genome SNP discovery.

Bacillus anthracis, the causative agent of anthrax, has been widely described as a clonal species. Here we report the use of both canonical SNP analysis and whole-genome sequencing to characterize the phylogenetic lineages of B. anthracis from the Netherlands. Eleven strains isolated over a 25-years period (1968-1993) were paired-end sequenced using parallel sequencing technology. Five canSNP groups or lineages, i.e. A.Br.001/002 (n=6), A.Br.Aust94 (n=2), A.Br.008/011 (n=1), A.Br.011/009 (n=1) and A.Br.Vollum (n=1) were identified. Comparative analyses, with a focus on SNPs discovery, were carried out using a total of 52 B. anthracis genomes. A phylogeographic "Dutch" cluster within the dominant A.Br.001/002 group was discovered, involving isolates from a single outbreak. Diagnostic SNPs specific to the newly identified sub-groups were developed into high-resolution melting SNP discriminative assays for the purpose of rapid molecular epidemiology. Phylogenetic relationships with strains from other parts of the world are discussed.