Shiga Toxin 2-Converting Bacteriophages Associated with Clonal Variability in Escherichia coli O157:H7 Strains of Human Origin Isolated from a Single Outbreak

Escherichia coli Shiga toxin-producing (STEC) strains are responsible for hemorrhagic diarrhea and hemolytic-uremic syndrome, which leads to acute renal failure in children (19, 22). E. coli O157:H7 is the most common virulent serogroup, but other serogroups have been described, such as O18, O26, O111, O128, and O138 (1). STEC is mainly transmitted by consumption of contaminated food and water and by personal contact (6, 17, 19). The number of outbreaks caused by STEC has increased drastically in all developed countries (19, 22). Among other virulence factors, the pathogenicity of STEC is related to the production of Shiga toxin 1 (Stx1), Stx2, or variants of Stx2 (7, 22). The genes encoding Shiga toxins (stx) are located in the genomes of temperate lambdoid bacteriophages (21). Stx2-converting phages are heterogeneous because they have wide ranges of DNA structures, restriction patterns, host spectra, and morphologies (10, 26, 30, 31). Phages carry stx1 (20), stx2 (21), and stx2 variants (18, 29). They carry not only the Shiga toxin genes but also some regulatory genes; hence, they are responsible for the production of toxin and its regulation (31, 32). Intraintestinal phage transmission between different bacteria has been described for stx1 (2) and in vitro for stx2 (26). Additionally, treatment with inhibitors of DNA synthesis, such as trimethoprim-sulfamethoxazole and mitomycin C, results in phage induction and an increase in toxin production in various STEC strains (12). The mobility of the stx2 characteristic mediated by phages suggests that a wide variety of E. coli strains (and other enterobacteria) are susceptible to infection by stx phages. Recent studies have indicated that strains of the most virulent enterohemorrhagic E. coli serotypes (O157:H7, O103:H2, O111:H−) belong to a few clone complexes (3, 11). However, the presence of variable DNA, which can be readily transferred by horizontal genetic exchange, has hampered our understanding of the phylogeny and pathogenesis of STEC. Moreover, genetic exchange has been demonstrated in lambdoid phages, and it has been proposed that these phages have a common gene pool from which DNA can be exchanged (9, 18). This exchange could influence the host strain, as has been described for E. coli O157:H7 (34), Shigella dysenteriae (15), and Shigella sonnei, (28), producing variations in toxin production and thus in the virulence of the host bacteria. This study of stx2-converting phages was carried out with 49 strains of E. coli isolated from a recent outbreak of STEC infection reported in Spain, which occurred in the Barcelona area between 19 September and 5 November 2000 and affected three schools. Infections in two of the schools were caused by contaminated food supplied by the same caterers. In the third school the transmission was attributed to personal contact. The whole outbreak involved more than 200 patients, most of whom were children under the age of 10 years. Six children developed hemolytic-uremic syndrome. The outbreak was associated with ingestion of E. coli O157:H7 via contaminated sausages (4), although bacteria were not isolated from the suspected food. Although all bacterial of the isolates were E. coli O157:H7 stx1− stx2+ and thus were assumed to be clonal, we were interested in examining the presence of stx2-converting phages in these bacteria. The presence and expression of stx2 varied from one isolate to another due to certain variability observed in the stx2-converting bacteriophages.