Human B cell immunity in the context of Zika and dengue virus infections

Dengue virus, consisting of four distinct serotypes (DENV1-4), is a mosquito-borne flavivirus that causes over a hundred million human infections annually. DENV infections can lead to a range of outcomes, from asymptomatic infection to classical dengue fever (DF) to dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS). B cell immune responses are critical for clearing flavivirus infections through serum antibodies from long-lived plasma cells (LLPCs) and the activation of memory B cells (MBCs). Conversely, B cells and antibodies from a previous infection can be associated with severity in a subsequent secondary DENV infection. The recent explosive epidemic of Zika in the Americas and the co-circulation of Zika virus (ZIKV) with the phylogenetically related DENV underscored the relevance of defining the role of MBC and antibody responses in these antigenically related viruses that overlap or succeed one another in space and time. In this dissertation, the effect of pre-existing DENV immunity in the development of B cell immune responses after a ZIKV infection and the analysis of the polyclonal MBC response to the four DENV serotypes plus ZIKV during DENV infection are studied./nAs a means to study MBCs at a polyclonal level, we developed, optimized and implemented a Multi-Color FluoroSpot (MCF) to enable the analysis of type-specific versus cross-reactive B cell populations at a single-cell level for all four DENV serotypes and ZIKV. This novel assay permitted the analysis of unique human samples from long-term cohort and hospital studies of pediatric DENV and ZIKV infections in Nicaragua./nWe then analyzed long-term MBC alongside serum antibody responses after ZIKV infection in patients who had experienced prior DENV infections or were DENV-naïve. We showed that ZIKV infection generates strong type-specific MBC responses and that the majority of late convalescent anti-ZIKV serum neutralizing activity was attributable to ZIKV-specific antibodies, despite prior DENV immunity. Moreover, the initial high levels of ZIKV and DENV cross-reactive MBC responses declined over time in patients with prior DENV immunity. Overall, our results suggest that despite the phylogenetic and antigenic relatedness between DENV and ZIKV, immunity to DENV only modestly shapes the breadth of B cell responses to ZIKV./nThe development of polyclonal MBCs to the four DENV serotypes and ZIKV during DENV infection is not fully understood. Here, we analyzed polyclonal MBCs from primary and secondary DENV infections over time using the MCF assay. We showed that DENV elicits robust type-specific MBC responses after primary and secondary DENV infection, with cross-reactive responses always being significantly higher. Despite the comparable number and proportion of type-specific and cross-reactive MBCs between primary and secondary DENV infections, we found that the breadth of the cross-reactive MBC response was greater after a secondary DENV infection, with more MBCs recognizing multiple DENV serotypes as well as ZIKV. Overall, this shows that the breadth of DENV MBCs enables broader recognition of not-yet encountered DENV serotypes, as well as the related flavivirus ZIKV, in subsequent infections./nTaken together, the work included in this dissertation has made significant contributions to the understanding of the development of B cell immunity after primary and secondary DENV and ZIKV infections in patients from a dengue-endemic setting, with important implications for vaccine development.

Advisor: Harris, Eva