ETT2 is a second cryptic type III secretion system in Escherichia coli which was first discovered through the analysis of genome sequences of enterohemorrhagic E. coli O157:H7. Comparative analyses of Escherichia and Shigella genome sequences revealed that the ETT2 gene cluster is larger than was previously thought, encompassing homologues of genes from the Spi-1, Spi-2, and Spi-3 Salmonella pathogenicity islands. ETT2-associated genes, including regulators and chaperones, were found at the same chromosomal location in the majority of genome-sequenced strains, including the laboratory strain K-12. Using a PCR-based approach, we constructed a complete tiling path through the ETT2 gene cluster for 79 strains, including the well-characterized E. coli reference collection supplemented with additional pathotypes. The ETT2 gene cluster was found to be present in whole or in part in the majority of E. coli strains, whether pathogenic or commensal, with patterns of distribution and deletion mirroring the known phylogenetic structure of the species. In almost all strains, including enterohemorrhagic E. coli O157:H7, ETT2 has been subjected to varying degrees of mutational attrition that render it unable to encode a functioning secretion system. A second type III secretion systemassociated locus that likely encodes the ETT2 translocation apparatus was found in some E. coli strains. Intact versions of both ETT2-related clusters are apparently present in enteroaggregative E. coli strain O42.The species Escherichia coli contains a wide range of commensal strains and pathogenic varieties (pathotypes) in addition to the model laboratory organism, E. coli K-12 (16). At least six pathotypes are associated with human intestinal disease: they are enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC), enterohemorrhagic E. coli (EHEC), enteroinvasive E. coli, enteroaggregative E. coli (EAEC), and diffusely adherent E. coli. Two pathotypes are associated with extraintestinal disease in humans, namely uropathogenic E. coli (UPEC) and neonatal meningitic E. coli (NMEC). In addition, it is now clear that on phylogenetic grounds, all members of the genus Shigella belong within the species of E. coli (50). Furthermore, this dazzling phenotypic variety is matched by remarkable variations in genome size, with the largest E. coli genomes possessing more than a megabase more DNA than the smallest ones (43).Initial studies of UPEC, and later of other pathotypes, suggested that E. coli strains often acquire new complex pathogenic phenotypes in a single step by the acquisition of pathogenicity islands, which contain virulence genes clustered on the chromosome and which are acquired en bloc by horizontal gene transfer (21). Similar studies with the related bacterium Salmonella enterica have delineated several Salmonella pathogenicity islands (Spi-1, Spi-2, Spi-3, etc.) (2, 22). The horizontal transfer of DNA by mobile elements such as bacteriophages and plasmids is also known to play a role in the evolution of virulence in E. coli and S...

Escherichia coli K-12 possesses two adjacent, divergent, promoterless flagellar genes, fhiA-mbhA, that are absent from Salmonella enterica. Through bioinformatics analysis, we found that these genes are remnants of an ancestral 44-gene cluster and are capable of encoding a novel flagellar system, Flag-2. In enteroaggregative E. coli strain 042, there is a frameshift in lfgC that is likely to have inactivated the system in this strain. Tiling path PCR studies showed that the Flag-2 cluster is present in 15 of 72 of the well-characterized ECOR strains. The Flag-2 system resembles the lateral flagellar systems of Aeromonas and Vibrio, particularly in its apparent dependence on RpoN. Unlike the conventional Flag-1 flagellin, the Flag-2 flagellin shows a remarkable lack of sequence polymorphism. The Flag-2 gene cluster encodes a flagellar type III secretion system (including a dedicated flagellar sigma-antisigma combination), thus raising the number of distinct type III secretion systems in Escherichia/Shigella to five. The presence of the Flag-2 cluster at identical sites in E. coli and its close relative Citrobacter rodentium, combined with its absence from S. enterica, suggests that it was acquired by horizontal gene transfer after the former two species diverged from Salmonella. The presence of Flag-2-like gene clusters in Yersinia pestis, Yersinia pseudotuberculosis, and Chromobacterium violaceum suggests that coexistence of two flagellar systems within the same species is more common than previously suspected. The fact that the Flag-2 gene cluster was not discovered in the first 10 Escherichia/Shigella genome sequences studied emphasizes the importance of maintaining an energetic program of genome sequencing for this important taxonomic group.The motile gamma-proteobacterium Escherichia coli has been widely accepted in biology as a model organism, an opinion typified by quotations such as "all cell biologists have two cells of interest: the one they are studying and Escherichia coli" (34) or Jacques Monod's famous dictum "Tout ce qui est vrai pour le Colibacille est vrai pour l'éléphant" ("What is true for E. coli is also true of the elephant") (26). However, within this single model species, which now encompasses the shigellas, there are remarkable variations in genome size, and the largest E. coli genomes possess more than 1 Mb more DNA than the smallest E. coli genomes (42). Comfortingly, the most commonly used laboratory strain, K-12, has one of the smallest E. coli genomes, leading to the often unwitting assumption that this model strain represents the ancestral or archetypical state of the species.Curiously, one area of bacteriology in which E. coli K-12 has been eclipsed as a model organism is the study of flagellar biosynthesis, assembly, and regulation. In this area, Salmonella enterica serovar Typhimurium strain LT2 has been the most commonly used model organism (34,35). Nonetheless, it has been assumed that the genetics and physiology of flagellar systems are essentially the same in E. coli and S. en...