Enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli (EHEC) produce the characteristic "attaching and effacing" (A/E) lesion of the brush border. Intimin, an outer membrane protein encoded by eae, is responsible for the tight association of both pathogens with the host cell. Several eae have been cloned from different EPEC and EHEC strains isolated from humans and animals. These sequences are conserved in the N-terminal region but highly variable in the last C-terminal 280 amino acids (aa), where the cell binding activity is localized. Based on these considerations, we developed a panel of specific primers to investigate the eae heterogeneity of the variable 3 region by using PCR amplification. We then investigated the distribution of the known intimin types in a large collection of EPEC and EHEC strains isolated from humans and different animal species. The existence of a yet-unknown family of intimin was suspected because several EHEC strains, isolated from human and cattle, did not react with any of the specific primer pairs, although these strains were eae positive when primers amplifying the conserved 5 end were used. We then cloned and sequenced the eae present in one of these strains (EHEC of serotype O103:H2) and subsequently designed a PCR primer that recognizes in a specific manner the variable 3 region of this new intimin type. This intimin, referred to as "," was present in human and bovine EHEC strains of serogroups O8, O11, O45, O103, O121, and O165. Intimin is the largest intimin cloned to date (948 aa) and shares the greatest overall sequence identity with intimin ␤, although analysis of the last C-terminal 280 aa suggests a greater similarity with intimins ␣ and ␥.

SummaryIn this study, we identified and characterized a novel secreted protein, the extracellular serine protease EspP, which is encoded by the large plasmid of enterohaemorrhagic Escherichia coli (EHEC) O157:H7. The corresponding espP gene consists of a 3900 bp open reading frame that is able to encode a 1300-aminoacid protein. EspP is synthesized as a large precursor which is then processed at the N-and C-termini during secretion. It can be grouped into the autotransporter protein family. The deduced amino acid sequence of EspP showed homology to several secreted or surface-exposed proteins of pathogenic bacteria, in particular EspC of enteropathogenic E. coli and IgA1 proteases from Neisseria spp. and Haemophilus influenzae. Hybridization experiments and immunoblot analysis of clinical EHEC isolates showed that EspP is widespread among EHEC of the serogroup O157 and that it also exists in serogroup O26. A specific immune response against EspP was detected in sera from patients suffering from EHEC infections. Functional analysis showed that EspP is a protease capable of cleaving pepsin A and human coagulation factor V. Degradation of factor V could contribute to the mucosal haemorrhage observed in patients with haemorrhagic colitis.

Isolates of the most common O serogroups of enteropathogenic Escherichia coli (EPEC) associated with infant diarrhea (designated class I) adhere to Hep-2 cells; the genes for this adhesin, termed EPEC adherence factor (EAF), are located on plasmids 50-70 MDa in size. Volunteers ingested 10(10) organisms of an O127:H6 Hep-2-adhesive class I strain (E2348/69) or its plasmid-minus, nonadhesive derivative. Diarrhea occurred in nine of 10 volunteers who ingested the parent strain (mean, 1,178 ml) but in only two of nine who took the plasmid-minus variant (mean, 433 ml; P less than .006). All volunteers ill from strain E2348/69 mounted serum IgA and IgG responses to a 94-kDa plasmid-associated outer membrane protein of E2348/69; this protein was found in other class I EPEC but not in enterotoxigenic or meningitic strains. The 50-70-MDa EAF plasmid seems necessary for full expression of pathogenicity in EPEC that exhibit Hep-2 adhesiveness. EPEC isolates of certain other, less common, O serogroups (O44, O86, and O114) are rarely Hep-2 adhesive. These EPEC, designated class II, possess distinct 50-70 MDa plasmids lacking EAF genes. Diarrhea was caused by 10(8) or 10(10) organisms of an O114:H2 class II EPEC strain (mean, 1,156 ml) in six of 11 volunteers. This result confirmed that class II EPEC are pathogenic by a mechanism not involving Hep-2 adhesiveness.

Subtypes of GBS defined by preceding infections were only approximately associated with different patterns of clinical, neurophysiologic, and immunologic features. A single infectious agent caused more than one type of pathology in GBS, implying interaction with additional host factors. Most patients had no identified infection.