Rotavirus is an important cause of diarrheal morbidity and mortality in infants and children, particularly in developing countries. The rotavirus outer capsid is composed of VP4 and VP7 proteins, which facilitate viral attachment and entry and are primary targets of neutralizing antibodies. Rotaviruses exhibit narrow species and cell tropism, with human rotaviruses growing poorly in most cell lines and animal models. This phenotype, to which VP4 contributes significantly, has hampered studies of human rotavirus molecular biology and vaccine development. Following rotavirus vaccine introduction, there has been a significant decrease in severe rotavirus disease but concurrent emergence and spread of previously uncommon rotavirus serotypes and continued evolution of common serotypes. Differential patterns of circulating rotavirus serotypes that correlate with the specific vaccine implemented in a geographic region have been reported, suggesting vaccine- mediated serotype selection. While the antigenic diversity of contemporary and emerging human rotaviruses suggests antibodies elicited by current vaccines may neutralize them inequivalently, neutralization has not yet been empirically tested. The recent development of a plasmid-based reverse genetics system for a simian rotavirus and the use of human intestinal enteroids to culture human rotaviruses present new opportunities to elucidate outcomes of interactions between rotavirus outer-capsid antigens and vaccine-elicited antibodies. To test the hypothesis that that outer-capsid antigenic diversity, vaccine, and cell type influence serum neutralization of human rotaviruses, we propose two integrated Subaims. In Subaim 1, we will engineer chimeric simian rotaviruses incorporating outer-capsid genes from contemporary and emerging pathogenic human strains or vaccine strains using reverse genetics. We will quantify the capacity of serum antibodies elicited in rabbits or infants by different rotavirus vaccines to bind and neutralize the engineered viruses using standard assays, to permit comparison with previous studies. By isolating a human G or P type in an otherwise isogenic background, these studies will provide insight into antigen-specific functions of VP7 and VP4 in neutralization. In Subaim 2, we will perform replication and neutralization assays in human cell lines and human intestinal enteroids, using a panel of chimeric rotaviruses and differentially vaccinated rabbit and infant sera. These studies will provide biologically relevant insights into serum antibody responses elicited by different vaccines and roles of VP4 and VP7 in neutralization, which may support models that differ from those proposed based on animal rotaviruses or experiments in animal cell lines. Together, these studies will reveal individual rotavirus outer-capsid antigen functions in neutralization and differences in the breadth and specificity of vaccine-elicited serum antibody responses, provide insight into immunological pressures influencing rotavirus population dynamics, and enable development of improved rotavirus neutralization assays and platforms to test immune responses to current globally circulating rotaviruses.
The proposed research will significantly improve our understanding of the interactions of specific outer-capsid proteins of rotavirus, an important cause of infant and child diarrheal disease, with the immune system and host cells. This work will inform us about the capacity of current vaccines to elicit antibodies that bind to different genotypes of rotavirus and prevent them from infecting cells. Such information will enable development of more effective tests of rotavirus neutralization, which may improve vaccine evaluation.
