title: Evaluation of biological safety in vitro and immunogenicity in vivo of recombinant Escherichia coli Shiga toxoids as candidate vaccines in cattle.
abstract: Cattle are the most important reservoir for enterohemorrhagic Escherichia coli (EHEC), a subset of shigatoxigenic E. coli (STEC) capable of causing life-threatening infectious diseases in humans. In cattle, Shiga toxins (Stx) suppress the immune system thereby promoting long-term STEC shedding. First infections of animals at calves' age coincide with the lack of Stx-specific antibodies. We hypothesize that vaccination of calves against Shiga toxins prior to STEC infection may help to prevent the establishment of a persistent type of infection. The objectives of this study were to generate recombinant Shiga toxoids (rStx1mut & rStx2mut) by site-directed mutagenesis and to assess their immunomodulatory, antigenic, and immunogenic properties. Cultures of bovine primary immune cells were used as test systems. In ileal intraepithelial lymphocytes both, recombinant wild type Stx1 (rStx1WT) and rStx2WT significantly induced transcription of IL-4 mRNA. rStx1WT and rStx2WT reduced the expression of Stx-receptor CD77 (syn. Globotriaosylceramide, Gb3) on B and T cells from peripheral blood and of CD14 on monocyte-derived macrophages. At the same concentrations, rStx1mut and rStx2mut exhibited neither of these effects. Antibodies in sera of cattle naturally infected with STEC recognized the rStxmut toxoids equally well as the recombinant wild type toxins. Immunization of calves with rStx1mut plus rStx2mut led to induction of antibodies neutralizing Stx1 and Stx2. While keeping their antigenicity and immunogenicity recombinant Shiga toxoids are devoid of the immunosuppressive properties of the corresponding wild type toxins in cattle and candidate vaccines to mitigate long-term STEC shedding by the reservoir host.
authors.affiliation: Institute of Hygiene and Infectious Diseases of Animals, Justus Liebig University, Frankfurter Str. 85-89, 35392, Giessen, Germany. katharina.kerner@vetmed.uni-giessen.de.
authors.forename: Katharina
authors.initials: K
authors.lastname: Kerner
authors.affiliation: Institute of Hygiene and Infectious Diseases of Animals, Justus Liebig University, Frankfurter Str. 85-89, 35392, Giessen, Germany. psbridger@googlemail.com.
authors.forename: Philip S
authors.initials: PS
authors.lastname: Bridger
authors.affiliation: Institute of Hygiene and Infectious Diseases of Animals, Justus Liebig University, Frankfurter Str. 85-89, 35392, Giessen, Germany. gabriele_koepf@yahoo.com.
authors.forename: Gabriele
authors.initials: G
authors.lastname: Köpf
authors.affiliation: Institute of Hygiene and Infectious Diseases of Animals, Justus Liebig University, Frankfurter Str. 85-89, 35392, Giessen, Germany. jule.froehlich@arcor.de.
authors.forename: Julia
authors.initials: J
authors.lastname: Fröhlich
authors.affiliation: Institute of Hygiene and Infectious Diseases of Animals, Justus Liebig University, Frankfurter Str. 85-89, 35392, Giessen, Germany. stefanie.barth@fli.bund.de.|Current Address: Friedrich-Loeffler-Institut, Institute of Molecular Pathogenesis, Naumburger Str. 96a, 07743, Jena, Germany. stefanie.barth@fli.bund.de.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Barth
authors.affiliation: Clinic for Ruminants and Swine (Internal Medicine & Surgery), Justus Liebig University, Giessen, Germany. hermann.willems@vetmed.uni-giessen.de.
authors.forename: Hermann
authors.initials: H
authors.lastname: Willems
authors.affiliation: Institute of Hygiene and Infectious Diseases of Animals, Justus Liebig University, Frankfurter Str. 85-89, 35392, Giessen, Germany. rolf.bauerfeind@vetmed.uni-giessen.de.
authors.forename: Rolf
authors.initials: R
authors.lastname: Bauerfeind
authors.affiliation: Institute of Hygiene and Infectious Diseases of Animals, Justus Liebig University, Frankfurter Str. 85-89, 35392, Giessen, Germany. georg.baljer@vetmed.uni-giessen.de.
authors.forename: Georg
authors.initials: G
authors.lastname: Baljer
authors.affiliation: Institute of Hygiene and Infectious Diseases of Animals, Justus Liebig University, Frankfurter Str. 85-89, 35392, Giessen, Germany. christian.menge@fli.bund.de.|Current Address: Friedrich-Loeffler-Institut, Institute of Molecular Pathogenesis, Naumburger Str. 96a, 07743, Jena, Germany. christian.menge@fli.bund.de.
authors.forename: Christian
authors.initials: C
authors.lastname: Menge
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002417:Cattle; D002418:Cattle Diseases / Q000276:immunology* / Q000382:microbiology; D004927:Escherichia coli Infections / Q000276:immunology / Q000382:microbiology / Q000662:veterinary*; D022361:Escherichia coli Vaccines / Q000009:adverse effects / Q000276:immunology*; D008297:Male; D016297:Mutagenesis, Site-Directed / Q000662:veterinary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D054323:Shiga-Toxigenic Escherichia coli / Q000276:immunology*; D014121:Toxoids / Q000494:pharmacology*; D014614:Vaccines, Synthetic / Q000009:adverse effects / Q000276:immunology
chemical_list: D001426:Bacterial Proteins; D022361:Escherichia coli Vaccines; D011994:Recombinant Proteins; D014121:Toxoids; D014614:Vaccines, Synthetic