The interplay between Group A Streptococcus virulence factors and a functional antibody response

Abstract

Background: Group A Streptococcus (GAS) is a serious human pathogen and effective vaccines are lacking. Two major vaccine targets are the M-protein and T-antigen, however there is a gap in understanding as to whether expression of either antigen impacts a protective immune response to the other during infection. This is of particular importance for M-proteins as members of this protein family are capable of interfering with antibody function. Accordingly, the aim of this project was to investigate how M-protein and T-antigen expression levels and functional capabilities influence opsonophagocytosis and killing of GAS in the presence of M-protein and T-antigen specific antisera. Methods: The project first characterised the rabbit derived antisera using ELISAs to determine the titre of antibodies specific for the antigens of interest. The characterised rabbit sera was then utilised to optimise a flow cytometry protocol to measure M-protein and T-antigen expression on GAS strains that express M proteins with differing immune evasion properties (emm1 and emm6 strains). Opsonophagocytic killing assays that utilise human neutrophil-like cells (HL-60 cells) were used to investigate the killing capacity of the M- and T-specific antisera alone and in combination against the emm1 and emm6 strains. Results: Serum characterisation established the specificity of the rabbit antisera against the GAS antigens and confirmed the ability for the M1 protein, but not the M6 protein, to non-specifically bind serum antibodies. The flow cytometry showed variation in the expression levels of M-protein and T- antigen on both emm1 and emm6 strains. The results of the opsonophagocytic killing assays with the rabbit antisera suggested that the level of M-protein expression, as well as the presence of an antibody binding motif within the M-protein, influenced T-antigen specific killing. In particular, a high level of M1 expression was associated with reduced killing by T1 antisera, and the M6/T6 strains were associated with negligible T-antigen expression and opsonophagocytic killing by T-antigen specific antisera. Conclusion: There is marked differences in opsonophagocytic killing of GAS between strains with different M-protein and T-antigen profiles. This highlights the need for careful consideration of antigen expression and characterisation when selecting strains for vaccine development and assessment.