Characteristics of emerging human-pathogenic Escherichia coli O26:H11 strains isolated in France between 2010 and 2013 and carrying the stx2d gene only

Abstract

Strains of Escherichia coli O26:H11 that were positive for stx2 alone (n = 23), which were not epidemiologically related or part of an outbreak, were isolated from pediatric patients in France between 2010 and 2013. We were interested in comparing these strains with the new highly virulent stx2a-positive E. coli O26 clone sequence type 29 (ST29) that has emerged recently in Europe, and we tested them by multilocus sequence typing (MLST), stx2 subtyping, clustered regularly interspaced short palindromic repeat (CRISPR) sequencing, and plasmid (ehxA, katP, espP, and etpD) and chromosomal (Z2098, espK, and espV) virulence gene profiling. We showed that 16 of the 23 strains appeared to correspond to this new clone, but the characteristics of 12 strains differed significantly from the previously described characteristics, with negative results for both plasmid and chromosomal genetic markers. These 12 strains exhibited a ST29 genotype and related CRISPR arrays (CRISPR2a alleles 67 or 71), suggesting that they evolved in a common environment. This finding was corroborated by the presence of stx2d in 7 of the 12 ST29 strains. This is the first time that E. coli O26:H11 carrying stx2d has been isolated from humans. This is additional evidence of the continuing evolution of virulent Shiga toxin-producing E. coli (STEC) O26 strains. A new O26:H11 CRISPR PCR assay, SP_O26_E, has been developed for detection of these 12 particular ST29 strains of E. coli O26:H11. This test is useful to better characterize the stx2-positive O26:H11 clinical isolates, which are associated with severe clinical outcomes such as bloody diarrhea and hemolytic uremic syndrome.

FIG 1

MLST and CRISPR typing of the O26:H11 clinical isolates. In the graphic representation of spacer arrangements in CRISPR1 and CRISPR2a, each unique spacer is represented by a unique combination of the center shape and background color. The shape in the center indicates the spacer length (□, 32 bp; Ø, 1,293 bp). Gaps were introduced to improve the alignment of similar CRISPR arrays. CRISPR1 and CRISPR2a allele numbers are shown in the respective columns to the left of the CRISPR arrays. Each unique combination of CRISPR1 and CRISPR2a alleles was assigned a CRISPR type (CT) number. *, stx_2d-positive strains.

FIG 2

Alignment with reference strain 5905 of the amino acid sequences of the C-terminal ends of the A subunits and the N-terminal ends of the B subunits of the 7 stx_2a-negative and stx_2d-positive strains. Numbers in parentheses indicate the reading frames used for amino acid translation. Asterisks indicate the separation of the amino acid sequences of subunits A and B of Stx2.