Infants and children living with HIV-1 develop HIV-1 envelope (Env)-reactive broadly neutralizing antibodies (bnAbs) with a higher frequency and faster kinetics than adults, but the mechanisms of pediatric bnAb induction are not fully defined. Importantly, understanding the cellular and molecular mechanisms of bnAb induction in pediatric subjects will improve our understanding of pediatric HIV-1 immunity, and inform pediatric HIV-1 vaccine design to elicit bnAbs. We recently used a chimeric simian-human immunodeficiency virus (SHIV) to infect neonate RMs that generated heterologous HIV-1 nAbs over time as young RMs. Following neonatal SHIV infection in rhesus macaques (RMs), 8/13 (62%) animals generated plasma nAbs against heterologous HIV-1, with increased titers over time (6-24 months) in association with enhanced germinal center (GC) activity, including antigen (Ag)-specific GC B cells—a known mechanism for bnAb induction in humans. After neonatal rearing, the available dams were infected with the same SHIV and 3/11 (27%) dams generated heterologous HIV-1 nAbs with increased titers over time. Compared to neonate RMs, dams also had lower mean plasma viral load over time, and higher levels of CD4 T follicular regulatory cells (Tfr)—both of these mechanistic differences are known to impact bnAb development. Thus, we now have an appropriate model to define pediatric HIV-1 immunity associated with Env-reactive bnAb induction. In this proposal, we will define the unique specificities of pediatric immunity associated with HIV-1 bnAb induction using immune cell transcriptomics (Aim 1) combined with viral env evolution analyses (Aim 2) in young SHIV-infected RMs. In Aim 3, we will isolate and characterize blood and GC B cell-derived HIV-1 nAbs in young RMs following neonatal SHIV infection, with a focus on defining the maturation of pediatric nAbs that can develop into bnAb status. Finally, we will use env-nAb coevolutionary analyses to identify candidate Env immunogens to elicit HIV-1 bnAbs as previously described. The specific aims for this proposal are as follows: Aim 1. To define the permissive immunologic milieu for heterologous HIV-1 nAb induction in pediatric subjects. Aim 2. To identify env mutation patterns associated with neutralization breadth development following SHIV-infection in young and adult RMs. Aim 3. To define the characteristics and maturation pathways of heterologous HIV-1 nAbs elicited following neonatal SHIV infection.