Molecular Characterization of a Serotype O121:H19 Clone, a Distinct Shiga Toxin-Producing Clone of PathogenicEscherichia coli

Abstract

Most illnesses caused by Shiga toxin-producing Escherichia coli (STEC) have been attributed to E. coli serotype O157:H7, but non-O157 STEC infections are now increasingly recognized as public health problems worldwide. The O121:H19 serotype is being isolated more frequently from clinical specimens and has been implicated in one waterborne outbreak. We used multilocus virulence gene profiling, a PCR-based assay, to characterize the virulence gene content of 24 isolates of serotype O121:H19 and nonmotile variants. We also performed multilocus enzyme electrophoresis and multilocus sequencing to establish the clonal relatedness of O121 isolates and to elucidate the relationship of O121 to common STEC clones. The 24 isolates were found to represent a single bacterial clone, as there was no allelic variation across 18 enzyme loci among the isolates. The complete nucleotide sequence of the intimin gene differed by four substitutions from that of the epsilon (Int-) allele of O103:H2 strain PMK5. The typical O121 virulence gene profile was similar to the profiles of enterohemorrhagic E. coli (EHEC) clones of E. coli: it included a Shiga toxin 2 gene (stx 2 ), two genes on the EHEC plasmid (toxB and ehxA), and the gene encoding intimin (eae). Despite the similarities, putative virulence genes distributed on O islands-large chromosomal DNA segments present in the O157:H7 genome-were useful for discriminating among STEC serotypes and the O121:H19 clone had a composite profile that was distinct from the profiles of the other major EHEC clones of pathogenic E. coli. On the basis of sequencing analysis with 13 housekeeping genes, the O121:H19 clone did not fall into any of the four classical EHEC and enteropathogenic E. coli groups but instead was closely related to two eae-negative STEC strains.

Characterization of Shiga Toxin-Producing Escherichia coli Strains Isolated from Human Patients in Germany over a 3-Year Period

“…EHEC hemolysin, which causes an enterohemolytic phenotype on blood agar, was detected in many STEC strains of different origins (12,13,28,29,69) and was found to be significantly associated with eae-positive STEC strains belonging to classical and emerging EHEC types (13,28,46,69,76). Similar findings were made in our study, where EHEC hemolysin was detected in 96.2% of the eae-positive STEC strains.…”

“…In this study, intimin ␤1 was detected in 123 STEC strains and 12 serotypes. Intimin ␤1 was found as a characteristic trait of all STEC strains with flagellar type H11 (Tables 1 and 3 Intimin ε was first described for EHEC O103:H2 strains (55) and more recently for emerging EHEC O121:H19 strains (31,76). Apart from EHEC O103 and O121 strains, we have detected intimin ε in two new groups of STEC O123:H2 and Ont:H2 strains (Table 1).…”

Detection and Characterization of Verocytotoxin-Producing Escherichia coli by Automated 5′ Nuclease PCR Assay

“…This panel can easily be extended to include other subtypes of specific genes in, for example, the LEE region or new factors when these are identified or when particular genes need to be detected for specific purposes. Our main interest has been to detect the VTEC virulence factors most important at present, but also to detect variants of some of these factors, as it has been shown that there is a strong correlation between, e.g., the eae variant, the serotype, and the presence of other virulence factors (33,45,52). Although some of the variants of the LEE genes are found only or primarily in EPEC strains, primers and probes for these variants are included to make the most complete characterization system.…”

Serotypes and Virulence Profiles of Non-O157 Shiga Toxin-Producing Escherichia coli Isolates from Bovine Farms

“…In addition to the stx genes, other virulence factors encoded by eaeA (intimin), tir (espE) (translocated intimin receptor), espA (EspA protein), and espB (EspB protein) are required for intimate attachment and the formation of the attaching and effacing (A/E) lesions characteristic of STEC infection (10,13,79). Other clinically significant virulence markers include hlyA (pO157 enterohemolysin releasing hemoglobin from red blood cells), katP (pO157 catalase peroxidase that defends the cell against oxidative damage), etpD (pO157, encodes part of a type II secretory pathway transporting proteins across the outer membrane) (73), lpf (chromosomal long polar fimbriae) (72), espP (pO157 extracellular serine protease autotransporter), saa (pO113 STEC agglutinating adhesion) (55), sab (pO113 STEC autotransporter) (33), toxB (pO157 adhesion), and iha (chromosomal iron-regulated gene A homolog adhesin) (68,70).…”