Recognition of enteropathogenic Escherichia coli virulence determinants by human colostrum and serum antibodies.

Human colostra and sera collected from Mexican mothers and their children at birth and 6 months thereafter were studied for the presence of antibodies against the bundle-forming pilus and several chromosomal virulence gene products (intimin and secreted proteins EspA and EspB) of enteropathogenic Escherichia coli (EPEC). Among 21 colostrum samples studied, 76, 71.5, 57, and 47% of them contained immunoglobulin A (IgA) antibodies against EspA, intimin, EspB, and BfpA, respectively. Interestingly, there was a difference in IgG response to EPEC antigens between the sera from neonates and sera from the same children 6 months later. While the number of neonates reacting to Esps and intimin diminished when they reached 6 months of age, those reacting with BfpA increased from 9 to 71%. Intimin from an enterohemorrhagic E. coli strain was also recognized by most of the samples reacting with EPEC intimin. These data suggest that Bfp and Esps elicit an antibody response during the early days of life of neonates and support the value of breast-feeding in areas of the world where bacterial diarrheal infections are endemic.

Identification of enterotoxigenic Escherichia coli harboring longus type IV pilus gene by DNA amplification.

DNA amplification of lngA, the structural gene of longus type IV pilus produced by human enterotoxigenic Escherichia coli (ETEC) was achieved by the use of specific oligonucleotide primers designed from the nucleotide sequence of lngA. A 630-bp fragment representing the entire lngA gene was amplified in eight prototype strains previously characterized as longus positive. Five ETEC strains producing colonization factor antigen III (CFA III) (also a type IV pilus) were also positive by PCR, confirming the DNA homology between CFA III and longus. None of the non-ETEC and non-E. coli enteropathogens studied showed the 0.63-kbp amplicon. The procedure thus detected only ETEC strains harboring type IV pili genes with or without other colonization factors. Except for five lngA PCR-positive, probe-positive strains, all lngA PCR-positive strains produced the pilin as demonstrated by immunoblotting. To test the amplification procedure in a clinical setting, a collection of 264 fresh clinical E. coli strains isolated from 88 Mexican children with diarrhea was screened by PCR. Among 82 ETEC isolates found, 30 (36.5%) were lngA PCR-positive. Twenty-seven percent of the children shed ETEC that possessed lngA. In parallel with DNA probes or PCR protocols to detect enterotoxin genes, the lngA PCR method may prove useful for detection of ETEC harboring type IV pilus genes in epidemiological studies.

Antibody response of children with enteropathogenic Escherichia coli infection to the bundle-forming pilus and locus of enterocyte effacement-encoded virulence determinants.

Enteropathogenic Escherichia coli (EPEC) express a plasmid-encoded type IV pilus termed bundle-forming pilus, which is associated with the formation of bacterial microcolonies on cultured epithelial cells. Bacterial attachment and effacement of the enterocyte brush border membrane is attributed to a surface outer membrane protein adhesin termed intimin and EPEC-secreted proteins EspA, EspB, and EspD. Except for intimin, production in vivo or antibody response against these virulence determinants during natural EPEC infections in young children has not been demonstrated. Antibody responses against BfpA, intimin, EspA, and EspB were investigated in Brazilian children naturally infected with EPEC. Generally, IgG antibodies against BfpA and EspB were the most commonly found, followed by anti-EspA and intimin antibodies. Thus, bundle-forming pilus and locus of enterocyte attachment-encoded products are produced in vivo during natural EPEC infections and elicit an immune response against heterologous EPEC virulence determinants. These findings have important implications in the immunoprophylaxis against EPEC infections.

Longus pilus of enterotoxigenic Escherichia coli and its relatedness to other type-4 pili--a minireview.

Longus is a long pilus produced by human enterotoxigenic Escherichia coli (ETEC) which shares significant structural and biochemical features with class-B type-4 pili. These pili include the toxin-coregulated pilus (TCP) of Vibrio cholerae, the bundle-forming pilus (BFP) of enteropathogenic E. coli and both longus and the colonization factor antigen III (CFA/III) of ETEC. These pili are produced under defined growth conditions indicating that they are under the control of different regulatory elements. While TCP is chromosomally encoded, the remaining pili are encoded on large virulence plasmids. Longus and CFA/III are closely related pili although certain DNA and protein differences also exist between them. This may account for the differences in the regulation, surface presentation, antigenicity, and prevalence of these two pilins among ETEC. Neighboring lngA, a second open reading frame termed lngB was found which encodes a protein with significant homology to proteins which are part of a type-II secretory system such as XcpV, OutC, and PulO of Pseudomonas aeruginosa, Erwinia chrysanthemi, and Klebsiella pneumoniae, respectively. This suggests that lngB may be an accessory gene involved in biogenesis of longus.

Diffuse-adhering Escherichia coli (DAEC) as a putative cause of diarrhea in Mayan children in Mexico.

Diarrhea is a major cause of infantile morbidity and mortality in developing countries. A community-based, case control study was conducted in a southern Mexican Mayan village for 3 weeks during the peak diarrhea period to prospectively identify the infectious agents associated with childhood diarrheal disease. Several enteropathogens were isolated from stools of 34 of 58 cases, although none was significantly associated with diarrhea. For the 24 cases from which no enteropathogens were isolated, diffuse-adhering Escherichia coli (DAEC) strains were significantly associated with diarrheal disease (P less than .02; odds ratio = 6; 95% confidence limit, 1.08-99.0). DAEC were highly heterogeneous with respect to plasmid content and serotype. Three DNA probes designed to differentiate E. coli exhibiting localized, diffuse, or aggregative adherence were compared with results from a standard HeLa cell binding assay to assess the utility of these probes in the field. This study provides evidence for the potential pathogenic capacity of DAEC and underscores the variety of diarrheal agents operating within a community.