Single nucleotide polymorphisms (SNPs) in stable genome regions provide durable measurements of species evolution. We systematically identified each SNP in concatenations of all backbone ORFs in 7 newly or previously sequenced evolutionarily instructive pathogenic Escherichia coli O157:H7, O157:H ؊ , and O55:H7. The 1,113 synonymous SNPs demonstrate emergence of the largest cluster of this pathogen only in the last millennium. Unexpectedly, shared SNPs within circumscribed clusters of organisms suggest severely restricted survival and limited effective population sizes of pathogenic O157:H7, tenuous survival of these organisms in nature, source-sink evolutionary dynamics, or, possibly, a limited number of mutations that confer selective advantage. A single large segment spanning the rfb-gnd gene cluster is the only backbone region convincingly acquired by recombination as O157 emerged from O55. This concatenomic analysis also supports using SNPs to differentiate closely related pathogens for infection control and forensic purposes. However, constrained radiations raise the possibility of making false associations between isolates.E. coli ͉ evolution ͉ SNPs

Escherichia coli O157:H7, a zoonotic human pathogen for which domestic cattle are a reservoir host, produces a Shiga toxin(s) (Stx) encoded by bacteriophages. Chromosomal insertion sites of these bacteriophages define three principal genotypes (clusters 1 to 3) among clinical isolates of E. coli O157:H7. Stxencoding bacteriophage insertion site genotypes of 282 clinical and 80 bovine isolates were evaluated. A total of 268 (95.0%) of the clinical isolates, but only 41 (51.3%) of the bovine isolates, belonged to cluster 1, 2, or 3 (P < 0.001). Thirteen additional genotypes were identified in isolates from both cattle and humans (four genotypes), from only cattle (seven genotypes), or from only humans (two genotypes). Two other markers previously associated with isolates from cattle or with clinical isolates showed similar associations with genotype groups within bovine isolates; the tir allele sp-1 and the Q 933W allele were under-and overrepresented, respectively, among cluster 1 to 3 genotypes. Stx-encoding bacteriophage insertion site typing demonstrated that there is broad genetic diversity of E. coli O157:H7 in the bovine reservoir and that numerous genotypes are significantly underrepresented among clinical isolates, consistent with the possibility that there is reduced virulence or transmissibility to humans of some bovine E. coli O157:H7 genotypes.Escherichia coli O157:H7 is an important food-and waterborne zoonotic pathogen that expresses two cardinal virulence factors: the ability to produce one or more Shiga toxins (Stx) encoded by genes located in lambdoid bacteriophages (8,33) and the ability to attach intimately to epithelial cells via expression of a pathogenicity island termed the locus of enterocyte effacement (LEE) (31). Feng et al. described a plausible series of evolutionary events leading to the evolution of the dominant non-sorbitol-fermenting, ␤-glucuronidase-negative E. coli O157:H7 clade from a nontoxigenic progenitor, E. coli O55:H7 (5). Shaikh and Tarr (27) assayed clinical isolates of this dominant E. coli O157:H7 clade for the Stx-encoding bacteriophage insertion sites defined in the strains that have been sequenced (7,24). Three principal groups of isolates sharing Stx bacteriophage insertion site genotypes were identified: isolates with an Stx2-encoding bacteriophage inserted at a location other than wrbA and with yehV occupied by a centrally truncated bacteriophage (cluster 1), isolates with an Stx2-encoding bacteriophage inserted into wrbA and with yehV occupied by a truncated bacteriophage as in cluster 1 (cluster 2), and isolates with a complete Stx1-encoding bacteriophage inserted into yehV and with an Stx2-encoding bacteriophage inserted into wrbA (cluster 3).Association of the genetic diversity within the sorbitolnegative, ␤-glucuronidase-negative E. coli O157:H7 clade with prophage insertions has been demonstrated previously. Kim et al. (14) identified two major lineages within this clade using octamer-based genomic scanning and also found prophage sequences in some of the ...

Fim operons were examined to illuminate the emergence of Escherichia coli O157:H7 from the less-virulent E. coli O55:H7. A fim invertible element deletion occurred only after O157:H7 descended from O55:H7, and after sorbitol nonfermenting O157 diverged. Type 1 pili nonexpression correlates with this deletion in all enterohemorrhagic E. coli (EHEC) tested. An N135K FimH mutation in the two most evolved O157:H7 clusters is not found in other EHEC. These data refine the evolutionary history of an emerging pathogen.

The sfp cluster, encoding Sfp fimbriae and located in the large plasmid of sorbitol-fermenting (SF) enterohemorrhagic Escherichia coli (EHEC) O157 (pSFO157), has been considered a unique characteristic of this organism. We discovered and then characterized the sfp cluster in EHEC O165:H25/NM (nonmotile) isolates of human and bovine origin. All seven strains investigated harbored a complete sfp cluster (carrying sfpA, sfpH, sfpC, sfpD, sfpJ, sfpF, and sfpG) of 6,838 bp with >99% nucleotide sequence homology to the sfp cluster of SF EHEC O157:NM. The sfp cluster in EHEC O165:H25/NM strains was located in an ϳ80-kb (six strains) or ϳ120-kb (one strain) plasmid which differed in structure, virulence genes, and sfp flanks from pSFO157. All O165:H25/NM strains belonged to the same multilocus sequence type (ST119) and were only distantly phylogenetically related to SF EHEC O157:NM (ST11). The highly conserved sfp cluster in different clonal backgrounds suggests that this segment was acquired independently by EHEC O165:H25 and SF EHEC O157:NM. Its presence in an additional EHEC serotype extends the diagnostic utility of PCR targeting sfpA as an easy and efficient approach to seek EHEC in patients' stools. The reasons for the convergence of pathogenic EHEC strains on a suite of virulence loci remain unknown.

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