Characterization of virulence factor intracellular spread A (IcsA, or VirG) functional epitopes against 2a and invasion and adherence.

There are no licensed vaccines against , a leading cause of children's diarrhea and travelers' diarrhea. To develop a cross-protective vaccine against heterogeneous species and serotypes, we attempted to apply an epitope- and structure-based vaccinology platform, multiepitope fusion antigen, to construct an optimal polyvalent chimeric immunogen with functional epitopes from the key virulence determinants. With invasion plasmid antigens B and D functional epitopes identified in recent studies, in this study, we focused on intracellular spread protein A (IcsA; also known as virulence gene G, VirG), a multifunctional virulence factor that plays roles in bacterial adherence, invasion, and particularly intracellular and intercellular spread. We predicted continuous B-cell immunodominant epitopes from the IcsA functional passenger domain, and we fused each epitope to a non-homologous carrier protein, adhesin CS4 subunit CsaB, for epitope fusions. We then immunized mice with each epitope fusion and examined antibody functions against bacterial invasion and adherence. Eleven IcsA B-cell epitopes were identified and fused to CsaB. Mice intramuscularly immunized with each epitope fusion developed IgG responses. Mouse serum antibodies significantly reduced or 2a bacterial invasion or adherence , particularly from the fusion with epitope #2 (GDNNDGNSCGGNG), #3 (GGSGADHNGDGGE), #7 (YIISGKEDDGTQNV), #9 (WNDTDGDSHG), or #10 (TILADNLSHHNIN). These IcsA epitopes are more effective in inducing functional antibodies, suggesting their antigenic utility in constructing an optimal polyvalent immunogen and developing a cross-protective vaccine, and perhaps in better understanding their role in IcsA biogenesis.IMPORTANCEAn effective vaccine is urgently needed to reduce -associated diarrhea and dysentery. IcsA (or VirG) plays a crucial role in bacterial pathogenesis and is conserved across species and serotypes, thus making IcsA an excellent antigen target in vaccine development. Identification of the functional epitopes of the IcsA passenger domain from this study enables us to construct an optimal epitope- and structure-based polyvalent immunogen for a cross-protective multivalent vaccine. Unlike the lipopolysaccharide-based whole-cell or subunit vaccine candidates that are serotype-specific, a multivalent vaccine carrying functional epitopes of conserved virulence determinants would be cross-protective against the heterogeneous species and serotypes, effectively preventing shigellosis and dysentery. Data from this study may also provide insightful information for a better understanding of IcsA biogenesis.