title: Development of mucosal protection against the heat-stable enterotoxin (ST) of Escherichia coli by oral immunization with a genetic fusion delivered by a bacterial vector.
abstract: An LT-B-ST (LT-B/ST) fusion peptide was constructed by genetically joining the 5' terminus of a synthetic gene coding for the heat-stable enterotoxin (ST) of Escherichia coli to the 3' terminus of the gene coding for the binding subunit of the heat-labile enterotoxin (LT-B) of E. coli. An eight-amino-acid, proline-containing linker was included between the LT-B and ST moieties. An aroA mutant of Salmonella dublin transformed with a plasmid carrying this genetic construct was shown to express a fusion peptide with antigenic determinants of both LT-B and ST. Mice were immunized orally with this strain or with a control strain expressing just LT-B from the same plasmid. Sera and mucosal secretions were obtained and analyzed for the presence of serum immunoglobulin G and mucosal immunoglobulin A that were able to recognize LT-B and ST by enzyme-linked immunosorbent assay (ELISA) and, more importantly, were able to neutralize native ST in the suckling mouse assay. Sera and mucosal secretions from animals immunized with the strain expressing the LT-B/ST fusion exhibited detectable ELISA reactivity against LT-B but not against native ST. However, even in the absence of detectable ELISA reactivity, both sera and mucosal secretions from these animals were able to neutralize the biological activity of native ST in the suckling mouse assay. These findings are important because they demonstrate the development of mucosal protection against ST by oral immunization with a genetic fusion delivered by a bacterial vector.
authors.affiliation: Department of Microbiology and Immunology, Tulane University School of Medicine, New Orleans, Louisiana 70112.
authors.forename: L
authors.initials: L
authors.lastname: Cárdenas
authors.forename: J D
authors.initials: JD
authors.lastname: Clements
mesh_terms: D000284:Administration, Oral; D000595:Amino Acid Sequence; D000818:Animals; D001427:Bacterial Toxins / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D004768:Enterotoxins / Q000235:genetics / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D029968:Escherichia coli Proteins; D005260:Female; D005822:Genetic Vectors; D007114:Immunization; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009092:Mucous Membrane / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology*; D012475:Salmonella / Q000235:genetics
chemical_list: D001427:Bacterial Toxins; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; C034151:heat stable toxin (E coli); C027538:heat-labile enterotoxin, E coli