BRIG for multiple genome comparison. BRIG is a free tool [42, 43] that requires a local installation of BLAST (we used BRIG 0.95 on Mac OS X). The output is a static image. • Launch BRIG and set the reference sequence (EHEC EDL933 chromosome) and the location of other E. coli sequences for comparison. If you include reference sequences for the Stx2 phage and LEE pathogenicity island, it will be easy to see where these sequences are located. • Click ‘Next’ and specify the sequence data and colour for each ring to be displayed in comparison to the reference. • Click ‘Next’ and specify a title for the centre of the image and an output file, then click ‘Submit’ to run BRIG. • BRIG will create an output file containing a circular image like the one shown here. It is easy to see that the Stx2 phage is present in the EHEC chromosomes (purple) and the outbreak genome (black), but not the EAEC or EPEC chromosomes.