Phylogenetic analysis of Shiga toxin 1 and Shiga toxin 2 genes associated with disease outbreaks

Abstract

Background: Shiga toxins 1 and 2 (Stx1 and Stx2) are bacteriophage-encoded proteins that have been associated with hemorrhagic colitis, hemolytic uremic syndrome and other severe disease conditions. Stx1 and Stx2 are genetically and immunologically distinct but share the same compound toxin structure, method of entry and enzymatic function.

Results: Phylogenetic analysis was performed using Stx1 and Stx2 amino acid and nucleotide sequences from 41 strains of Escherichia coli, along with known stx sequences available from GenBank. The analysis confirmed the Stx1 and Stx2 divergence, and showed that there is generally more sequence variation among stx2 genes than stx1. The phylograms showed generally flat topologies among our strains' stx1 and stx2 genes. In the stx2 gene, 39.5% of the amino acid sites display very low nonsynonymous to synonymous substitution ratios.

Conclusion: The stx1 and stx2 genes used in this phylogenetic study show sequence conservation with no significant divergence with respect to place or time. These data could indicate that Shiga toxins are experiencing purifying selection.

Figure 1

Stx1 A and B Subunit Amino Acid Alignment. Sequences that are identical are on the same line and are pipe-delimited.

Figure 2

stx1 Maximum Likelihood Nucleotide Phylogenetic Tree (unrooted). Bootstrap values over 50% are displayed. Pipe-delimited strain names indicate identical sequences. Dates and locations of the isolates are shown if available. The horizontal bar shows 0.001 nucleotide substitutions per site.

Figure 3

Stx1 Distance Amino Acid Phylogenetic Tree (unrooted). Bootstrap values over 50% are displayed. Pipe-delimited strain names indicate identical sequences. Dates and locations of the isolates are shown if available. The horizontal bar shows 0.001 amino acid substitutions per site.

Figure 4

Stx2 A and B Subunit Amino Acid Alignment. Sequences that are identical are on the same line and are pipe-delimited.

Figure 5

Color Coded Selecton Results for stx2 Sequences. Nonsynonymous:synonymous (d_N:d_S) ratios using the output from the Selecton program. Shades of yellow (1 and 2) indicate a d_N:d_S ratio of > 1 or positive selection. Any shade of bordeaux (4–7) indicates a d_N:d_S ratio of < 1 or purifying selection.

Figure 6

stx2 Maximum Likelihood Nucleotide Phylogenetic Tree (unrooted). Bootstrap values over 50% displayed. Pipe-delimited strain names indicate identical sequences. Dates and locations of the isolates are shown if available. The horizontal bar shows 0.1 nucleotide substitutions per site.

Figure 7

Stx2 Distance Amino Acid Phylogenetic Tree (unrooted). Bootstrap values over 50% are displayed. Pipe-delimited strain names indicate identical sequences. Dates and locations of the isolates are shown if available. The horizontal bar shows 0.1 amino acid substitutions per site.

Figure 8

Unrooted Maximum Likelihood Nucleotide Phylogenetic Tree of all Shiga Toxin Sequences. The horizontal bar shows 0.1 nucleotide substitutions per site.