Results MLST analysis We have previously reported that aEPEC isolates obtained during a water quality study were heterogeneous in terms of serotype, intimin type and patterns of adherence to HEp-2 cells [20]. This overall heterogeneity was confirmed by MLST analysis, which showed that 56 of the 79 aEPEC strains PARP activation of human origin STI571 investigated in the study belonged to one of 11 different clades and that 23 strains could not allocated to a clade (Figure 1). As observed with phylogenetic analyses of A/E strains of E. coli in general, there was a tendency for each clade to contain strains with the same intimin and flagellar type. Five of 11 clades which contained aEPEC strains in this

study were clades that include either tEPEC or STEC strains, whereas six clades were apparently distinct for aEPEC. These six clades comprised one which contained three strains with intimin-β and H7 (and O-antigens, O25 or O153); one clade with seven strains with intimin-ν and H19 (all O-nontypable [nt]); one clade with six strains with intimin-θ and H21 (O119 and Ont); a clade with five strains with GSI-IX intimin-ι and H8 or H- (O98, O107 or O177); one with four strains with intimin-κ and H10 or H- (O49, O88, and O153), and one with 13 strains with intimin-α and H6 or H34 (O71, O125, O126, and Ont). The last-mentioned clade was

closely related to a tEPEC clade (EPEC-1), which also comprises strains with intimin-α and flagellar antigen, H6. Figure 1 Phylogenetic relationships of sequence types of 95 strains of attaching-effacing E. coli. An unrooted phylogenetic

tree was constructed by the neighbour-joining algorithm based on the Kimura two-parameter model of nucleotide substitution. Bootstrap values greater than 50% based on 500 replications are given at the internal nodes. Strain names highlighted in pink are reference strains of typical EPEC or STEC; those highlighted in pale blue and yellow were originally isolated from cattle and rabbits, respectively, those highlighted in green were from children with persistent diarrhoea, and those highlighted in grey were from humans without diarrhoea. The right hand column indicates distinctive Urease aEPEC clades in boldface type. The intimin and flagella type is shown for each clade. Abbreviations: nt, non-typable; int, intimin; ND, not determined. Of the strains that clustered with known clades of tEPEC or EHEC, three (all intimin-γ and O55:H7) belonged to the EHEC-1 clade, which also includes the pandemic, prototypical O157:H7 EHEC clone. Five aEPEC isolates grouped within the EPEC-2 clade which includes pEAF/BFP-positive strains with intimin-β and H2. The aEPEC serotypes in this clade included O15:Hnt, O114:H2, O117:H2, O128:H2, and Ont:H2. This clade also contained the prototypical aEPEC strain, E128012 [12], two rabbit-specific EPEC (REPEC) strains, 84/110/1 and E22 (both of which carry intimin-β and are serotype O103:H2), and a calf isolate, also O103:H2, but with intimin-ε.