An outbreak due to Shiga toxin-producing Escherichia coli O26:H11 (STEC) occurred at a nursery in southeastern Japan in 1997. Thirty-two children had watery or bloody diarrhoea but none of them suffered from haemolytic-uremic syndrome. All of the STEC O26 were isolated during the period from 23 July to 22 August from 24 children, 3 nurses, and 2 food samples. These organisms had stx1 and eae genes but none of the other genes for which we tested (stx2, bfp, and EAF plasmid). They also possessed multiple antimicrobial resistances, which were encoded by a transmissible plasmid, and showed mostly identical genomic pulsed-field gel electrophoretic patterns. The results of this investigation suggested that contaminated food was the main contributing factor to this multiple antimicrobial-resistant STEC O26 infection, and person-to-person transmission also contributed to the spread of this outbreak.

We developed a quick genetic approach to screen variants of the intimin gene (eae) by using a heteroduplex mobility assay (HMA) that targets the 5 conserved region of eae. The eae variants were categorized into 4 major HMA types and 10 minor subtypes.Enteropathogenic Escherichia coli (EPEC) and Shiga toxinproducing E. coli (STEC) produce characteristic attaching and effacing lesions using intimin encoded by eae (24). The eae genes of several strains have been cloned and sequenced and have a highly conserved 5Ј-terminal region but are variable in the 3Ј-terminal region (13). Allele-specific PCRs targeting eae genes in the variable 3Ј region have been employed to determine eae types (1,5,10,27,30,31,33) and subtypes in combination with restriction fragment length polymorphism (RFLP) (4, 27). Ramachandran et al. (29) designed universal PCR primers to amplify the Int280-encoding region and identified types by RFLP. Recently, methods using real-time PCR (25) and oligonucleotide microarray (14) have been developed. Eighteen types and nine subtypes of intimin, namely, ␣, ␣2, ␤1 to -3, ␥1, ␥2, ␦, ε, ε2 to

Enteroaggregative Escherichia coli (EAggEC) are an important cause of diarrhea. Four types of AAF have been identified; however, their prevalence and association with virulence properties remain unclear. E. coli strains carrying the aggR gene as EAggEC that were isolated in Japan and Thailand (n ¼ 90) were examined for AAF subunit genes, two toxin genes (pet/astA), and clump formation. The most prevalent AAF gene was hdaA (28%), followed by aafA (20%), aggA (12%), and agg3A (4%), as well as a putative new AAF sequence (25.6%). Retention status of the toxin genes and intensities of clump formation appeared to vary according to the AAF type.Key words aggregative adherence (AA), aggregative adherence fimbriae (AAF) type, aggR-positive Escherichia coli, toxin gene.Enteroaggregative Escherichia coli (EAggEC) comprise an emerging group of pathogens that cause pediatric and adult diarrhea worldwide and are known to be heterogeneous in their symptoms (1, 2) and patterns of retaining virulence genes (3-5). EAggEC exhibits characteristic AA at the surface of cultured cells by means of AAF (2). Four distinct AAF variants have been identified to date based on the sequences of fimbrial AAF subunits encoded by aggA (6), aafA (7), agg3A (8), and hdaA (alias agg4A) (9).Many putative virulence factors have been reported in addition to AAF, including EAST1, a 104-kDa cytotoxin designated as a Pet, and a global transcriptional regulator (AggR). AggR is the central regulator of virulence functions in EAggEC; therefore, the term 'typical EAEC List of Abbreviations: AA, aggregative adherence; AAF, aggregative adherence fimbriae; EAggEC, enteroaggregative Escherichia coli; EAST1, enteroaggregative E. coli heat stable enterotoxin 1; Pet, plasmid encoded toxin.

Enteropathogenic Escherichia coli (EPEC) strains produce a bundle-forming pilus (BFP) that mediates localized adherence (LA) to intestinal epithelial cells. The major structural subunit of the BFP is bundlin, which is encoded by the bfpA gene located on a large EAF plasmid. The perA gene has been shown to activate genes within the bfp operon. We analyzed perA gene polymorphism among typical (eaeand bfpA-positive) EPEC strains isolated from healthy and diarrheal persons in Japan (n = 27) and Thailand (n = 26) during the period 1995 to 2007 and compared this with virulence and phenotypic characteristics. Eight genotypes of perA were identified by heteroduplex mobility assay (HMA). The strains isolated in Thailand showed strong autoaggregation and had an intact perA, while most of those isolated in Japan showed weak or no autoaggregation, and had a truncated perA due to frameshift mutation. The degree of autoaggregation was well correlated with adherence to HEp-2 cells, contact hemolysis and BFP expression. Our results showed that functional deficiency due to frameshift mutation and subsequent nonsense mutation in perA reduced BFP expression in typical EPEC strains isolated in Japan.

A lactose slow-fermenting, non motile Escherichia coli strain was isolated from a diarrheal patient returning from Indonesia. Examination of virulence of the strain showed that it penetrated into tissue culture cells, and were positive in Serény test and enzyme-linked immunosorbent assay for detection of enteroinvasive E. coli. A large plasmid responsible for virulence was also detected. Thus the strain was confirmed to be a typical enteroinvasive E. coli. Analysis of O serotype using the antisera against E. coli O1-O170 revealed that the strain had O121 antigen, whose antiserum is not included in the commercial serotyping kit for the diagnosis of diarrheagenic E. coli.