AN ESCHERICHIA-COLI CS31A FIBRILLUM CHIMERA CAPABLE OF INDUCING MEMORY ANTIBODIES IN OUTBRED MICE FOLLOWING BOOSTER IMMUNIZATION WITH THE ENTERO-PATHOGENIC CORONAVIRUS TRANSMISSIBLE GASTROENTERITIS VIRUS

CS31A fibrillae are thin, flexible, heteropolymeric proteinaceous appe ndages exposed as a capsule-like material around the cell surface of c ertain Escherichia coli strains. Two antigenic peptides of the S spike glycoprotein (TGEV-S) amino acids (aa) 363-371 and 521-531 of the tra nsmissible gastroenteritis virus (TGEV) were tandemly introduced in th e loop-structured variable region an 202-218 of the major ClpG subunit protein composing the bulk of CS3IA. The resulting hybrid fibrillae w ith a 25 aa heterologous peptide were produced at the cell surface. Us ing a monoclonal antibody, (Mab) specific for the TGEV epitopes, purif ied hybrid fibrillae were analysed in Western blotting tinder native c onditions, which showed that the two viral epitopes Mere recognized im munologically as an integral part of the hybrid fibrillae, and therefo re that they were antigenically active. The immunogenicity, of the fus ion construct was evaluated with live recombinant bacteria, purified h ybrid ClpG monomers, and purified chimeric CS31A polymers. Whatever th e form of hybrid used as antigen, intraperitoneally immunized outbred mice elicited serum anti-TGEV peptides antibodies (Abs) with significa nt titres and capable of recognizing native TGEV particles, indicating that the epitopes are exposed in an immunogenic conformation in all c ases. However, virus neutralization titres were only obtained after im munization with either purified polymers or monomers. Furthermore, 4 m onths after an ultimate immunization with 20 mu g of hybrid fibrillae mice developed a strong anamnestic Ab response against the two TGEV pe ptides following booster inoculation with virions. We conclude that CS 31A fibrillae carrying a combination of TGEV epitopes as insert can in duce an immunological memory in outbred animals infected with TGEV, an d therefore that hybrid CS31A fibrillae may prove efficient as compone nts of a subunit vaccine. Copyright (C) 1997 Elsevier Science Ltd.