Human colostrum and serum contain antibodies reactive to the intimin-binding region of the enteropathogenic Escherichia coli translocated intimin receptor.
BACKGROUND
In Brazil, enteropathogenic Escherichia coli (EPEC) diarrhoea is endemic in young infants. A characteristic feature of EPEC adhesion to host cells is intimate attachment leading to the formation of distinctive "attaching and effacing" (A/E) lesions on mammalian cells. Two genes directly involved in intimate adhesion, eae and tir, encode the adhesion molecule intimin and its translocated receptor Tir, respectively. The intimin-binding domain of Tir was recently mapped to the middle part of the polypeptide (Tir-M), and the amino (Tir-N) and carboxy (Tir-C) termini were found to be located within infected host cells. Recently, it was shown that colostrum samples from mothers living in Sao Paulo contain IgA-class antibodies reactive with a number of proteins associated with EPEC virulence. It has also been shown that patients infected with verocytotoxin-producing E. coli O157 can produce antibodies to Tir. In the current study antibody responses to the different Tir domains were analyzed in sera and colostrum samples collected in an EPEC-endemic area of Brazil.


METHODS
Recombinant Tir, Tir-N, Tir-M, and Tir-C were expressed as His-tagged protein in E. coli BL21a and purified on nickel columns. Western blot analysis was used to investigate colostrum IgA- and serum IgG-class antibodies reactive with the Tir fragments.


RESULTS
Anti-Tir IgG antibodies were detected in the serum of children, with (63%) or without (50%) diarrhoea. Anti-Tir IgA-class antibodies were detected in all the colostrum pools tested. With the use of both serum IgG- and colostrum IgA-class antibodies, an immunodominant domain of the Tir-polypeptide, Tir M, was identified.


CONCLUSIONS
The intimin-binding region of Tir (Tir-M) is the immunodominant region of the polypeptide in humans. Both serum IgG-class and colostrum IgA-class antibodies reacted predominantly with the Tir-M domain.
Sanches
M I
MI
Departamento de Microbiologia, Instituto de Ciencias Biomedicas, Sao Paulo, Brazil.
Keller
R
R
Hartland
E L
EL
Figueiredo
D M
DM
Batchelor
M
M
Martinez
M B
MB
Dougan
G
G
Careiro-Sampaio
M M
MM
Frankel
G
G
Trabulsi
L R
LR
D018829:Adhesins, Bacterial; D000907:Antibodies, Bacterial / Q000032:analysis* / Q000097:blood*; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D001426:Bacterial Proteins / Q000276:immunology; D001665:Binding Sites; D001938:Brazil; D002352:Carrier Proteins; D003126:Colostrum / Q000276:immunology*; D018604:Epitope Mapping; D004926:Escherichia coli / Q000276:immunology*; D019453:Escherichia coli O157 / Q000276:immunology; D029968:Escherichia coli Proteins; D005260:Female; D006801:Humans; D007070:Immunoglobulin A / Q000032:analysis; D007074:Immunoglobulin G / Q000097:blood; D010446:Peptide Fragments / Q000276:immunology; D011956:Receptors, Cell Surface / Q000276:immunology*; D011994:Recombinant Proteins / Q000276:immunology
D018829:Adhesins, Bacterial; D000907:Antibodies, Bacterial; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D007070:Immunoglobulin A; D007074:Immunoglobulin G; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; C109287:Tir protein, E coli; C073819:eaeA protein, E coli