This study investigated the process of virulence acquisition in a Shiga toxin-producing Escherichia coli O104:H4 strain, TY-2482, by comparing the sequence of stx2 (Shiga toxin 2) prophage with the sequences of 1421 complete bacterial genomes and analyzing the evolutionary relationships of the stx2 prophage-like regions identified. The results showed a strong positive correlation between the evolutionary distance and the distance to switch sites of GC skew, suggesting that the loci near these sites are "hotspots" for insertions. These findings represent a potential new method to identify virulence factors in bacterial genomes.