PROJECT SUMMARY Dengue viruses (DENV) cause a significant and unchecked burden of human death and disease, with vaccine development hindered by critical gaps in our understanding of how multi-serotypic protection against DENV is generated, sustained, and subsequently identified in immunological assays. As the greatest risk for severe dengue illness occurs with secondary infection, DENV vaccines will need to generate protection against at least two serotypes simultaneously to maximize efficacy and safety. Our prior studies have demonstrated that durable, multi-typic immunity can be achieved naturally, through sequential exposures accumulated over time in hyperendemic areas for DENV transmission. Accordingly, our objective is to define the impacts of a child’s earliest flavivirus exposures in shaping DENV humoral immune phenotypes and clinical outcomes of subse- quent DENV exposures, generating important benchmarks for immune correlates of protection. To address this objective, we will leverage an ongoing long-term multigenerational family cohort study for DENV transmission in Kamphaeng Phet, Thailand. The cohort was established in 2015, leveraging NIH P01 and US DOD funds, and has enrolled over 3000 individuals within 500 families. 432 primary DENV infections have been identified among 814 DENV-naïve children to date, with more to be identified by the end of the study period in 2028 and marking 13 years of continuous surveillance. Incident infections are identified through quarterly sampling to detect seroconversions and through active surveillance for acute dengue illnesses. We will relate levels of maternally-transferred immunity, through placental transfer and breastfeeding, to risks of dengue illness with primary DENV infection in 750 mother-infant dyads (including 500 previously-enrolled and 250 newly-enrolled dyads) (Aim 1). Next, we will continue our long-term follow-up of DENV-naïve children and identify isotype- and antigen-specific DENV antibody phenotypes associated with protection from illness with post-primary DENV infection (Aim 2). Finally, we will relate non-DENV flavivirus exposures (Japanese enceph- alitis virus [JEV] vaccination, Zika virus infection, JEV infection) to risks of subsequent dengue illness, defining effects of time since exposure, pre-infection antibody phenotypes, and JEV vaccine type (Aim 3). These activities are consistent with NIAID’s mission to better understand, treat, and ultimately prevent infec- tious diseases. The application is innovative in using a custom multiplex panel for profiling DENV antibodies in saliva, permitting frequent longitudinal sampling, and in using advanced modeling techniques to reconstruct immune kinetics and identify subclinical infections. Successful completion of study aims will represent an im- portant advancement towards identifying immune correlates of durable, multi-serotypic protection against den- gue illness, providing critical benchmarks for diagnostics, triage, and DENV vaccines and immuno-therapies.

PROJECT NARRATIVE Dengue viruses (DENV) are the most significant arboviral cause of human death and disease, but vaccine development is challenged by our incomplete understanding of how lasting immunity to multiple DENV serotypes may be generated and how it may be identified in immunological assays. This study will leverage an established and uniquely long-term family cohort study for DENV in Thailand to define how the early immune environment to DENV, as shaped by maternally-transferred DENV antibodies and the timing and features of early DENV and non-DENV flavivirus exposures, shapes an individual’s future antibody-mediated immune profiles to DENV and subsequent risks of dengue illness. This proposed research is relevant for human health because it will identify features of exposure histories and pre-infection immunity associated with protection from dengue illness, ultimately supporting the identification of correlates of protection, which are critically needed for public health surveillance and countermeasure development.