Mapping human norovirus antigens during infection reveals the breadth of the humoral immune response

Human noroviruses (HuNoV) are the leading cause of acute gastroenteritis worldwide. The humoral immune response plays an important role in clearing HuNoV infections and elucidating the antigenic landscape of HuNoV during an infection can shed light on antibody targets to inform vaccine design. Here, we utilized Jun-Fos-assisted phage display of a HuNoV genogroup GI.1 genomic library and deep sequencing to simultaneously map the epitopes of serum antibodies of six individuals infected with GI.1 HuNoV. We found both unique and common epitopes that were widely distributed among both nonstructural proteins and the major capsid protein. Recurring epitope profiles suggest immunodominant antibody footprints among these individuals. Analysis of sera collected longitudinally from three individuals showed the presence of existing epitopes in the pre-infection sera, suggesting these individuals had prior HuNoV infections. Nevertheless, newly recognized epitopes surfaced seven days post-infection. These new epitope signals persisted by 180 days post-infection along with the pre-infection epitopes, suggesting a persistent production of antibodies recognizing epitopes from previous and new infections. Lastly, analysis of a GII.4 genotype genomic phage display library with sera of three persons infected with GII.4 virus revealed epitopes that overlapped with those identified in GI.1 affinity selections, suggesting the presence of GI.1/GII.4 cross-reactive antibodies. The results demonstrate that genomic phage display coupled with deep sequencing can characterize HuNoV antigenic landscapes from complex polyclonal human sera to reveal the timing and breadth of the human humoral immune response to infection.