Mapping of B-cell epitopes on the fusion protein of human respiratory syncytial virus

HRSV is the leading cause of lower respiratory tract (LTR) infection in the new-bom and during early childhood, accounting for approximately 90% of the reported cases of bronchiolitis in infancy. The WHO estimates that of the 12.2 million annual deaths in children under five years of age, a third is due to acute infections of the LRT. RSV may also be the cause of up to 2.4 % community-acquired LRT infections in adults of less than 60 years of age and in elderly people. The fusion (F) and attachment (G) proteins are the major protective antigens in RSV. Almost all the available F-specific monoclonal antibodies efficiently neutralize RSV in vitro and monoclonal antibodies have been successfully used for in vivo treatment of human RSV infection. Since the F-protein is genetically more stable than the G-protein it represents a better candidate for vaccine development. Immunity to RSV includes serum antibody, secretory antibody, and cytotoxic T-cell responses. Secretory antibodies are mainly effective in the upper respiratory tract, while serum antibodies are the key for resolution of LRT infection. However, immunity to RSV is incomplete and re-infections occur throughout life. The work described in this thesis was designed to: A: Identify both linear and conformational B-cell epitopes of the fusion protein of human RSV as a part of a programme of work to develop an epitope based vaccine against RSV. 55 overlapping peptides covering the whole of the fusion protein of RSV were synthesized by solid phase synthesis using the RAMPs system these peptides were used to screen sera from RSV-infectcd individuals (9 adults and 49 infants) for reactivity with linear epitopes using an Enzyme Linked Immunosorbent Assay (ELISA). In addition, a human monoclonal antibody (RF2) that recognizes a conformational epitope on the fusion protein of HRSV was used to screen a combinatorial peptide library to identify mimotopes of this conformational epitope. The linear epitopes and the mimotopes identified in this way were used in vivo experiments to assess their ability to induce anti-peptide antibodies that can cross- react with RSV. B. In an attempt to throw light on the question of how RSV can re-infect the same individual, serial serum samples were obtained from infants prior to exposure to RSV at 6 months of age, following primary exposure at 9-12 months and following second exposure at 12-18 months. The reactivity of these sera with the 55 overlapping peptides from the F protein was assessed by ELISA to determine if there were differences in the pattern of peptide recognition at different stages of exposure to RSV.