Infection phenotypes of CpG-recoded RNA viruses in animal models with immature immunity

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The long-term goal of the proposed research program is to understand better how increasing the number of CpG (cytosine-phosphate-guanine) dinucleotides in viral genomes affects infection and host immune responses and harness this knowledge to improve vaccination in the veterinary field. We are particularly interested in vaccine technology that will be safe to apply during pregnancy and in newborn and young animals with immature immunity. Increasing CpG dinucleotide numbers in RNA viral genomes, while preserving natural amino acid composition of proteins and antigenic profile, attenuates infection. Attenuated infection, however, leads to robust immune responses. It has been demonstrated that host immunity more efficiently targets CpG-recoded mutants than wild-type viruses. Despite significant progress, there is no full understanding of the mechanism(s) through which CpG composition influences virulence. In addition to exciting fundamental questions of virus-host interactions, CpG recoding potentially provides a cutting-edge technology for safer live vaccines, which will cause attenuated infection, no pathology, but robust immune responses. So far, one CpG vaccine candidate has been characterized in adult mice, showing promising results. However, safety margins of the CpG recoding technology in general, and particularly in animals with an immature immune system, are not defined. We chose to work with Zika virus (ZIKV) as a model because it affects fetuses and neonates. Our research applies a novel reverse genetics system to manipulate CpG frequencies in viral RNA genomes, a neonatal mouse model, and the first fetal pig model for ZIKV infection in the world. To better understand how increasing of CpG dinucleotides in an RNA viral genome affects infection and to determine if increasing of CpG dinucleotides leads to attenuated infection phenotypes in neonates and fetuses, we will address four objectives: First, we will evaluate a mechanistic relationship between the number of CpG dinucleotides in an RNA viral genome and infection phenotypes in different cell lines. Second, we will compare infection outcomes (clinical signs/mortality) and immune responses in newborn animals infected with wild and CpG-recoded ZIKV. Finally, we will address in utero infection kinetics, pathology, and immune responses to wild and CpG-recoded viruses in fetuses at the early stage and mid-stage of development. These experiments will improve our understanding of how CpG recoding in viral genomes affects infection and host immunity, and safety margins of the CpG-recoded viruses in animals with immature immunity. This basic knowledge will influence theories and application of the CpG vaccine technology particularly in the veterinary field benefiting Canadians and global community. Significantly, the program will also provide outstanding multidisciplinary training for young researchers.