Memory B cells and long-lived plasma cells are responsible for producing neutralizing antibodies that can effectively eliminate a pathogen. Understanding the function of these cells in response to infection and how they can be induced and maintained by vaccination is therefore critical to eradicating diseases that are global health burdens. Malaria, caused by Plasmodium spp, is a major global health burden that is in urgent need of a vaccine. Over fifty years ago it was shown that transfer of human immune serum can neutralize Plasmodium parasites during the blood stage of infection. Little is known however about the Plasmodium-specific B cells that produce these antibodies due to the difficulties of studying low frequency antigen-specific B cells. Additionally, it is not understood how recently described populations of heterogeneous memory B cell (MBC) subsets induced by protein immunization form or function in response to infection. To clarify functional roles for distinct MBC subsets during malaria infection, tetramers were generated that identify Plasmodium-specific MBCs in both humans and mice. Multiparameter flow cytometry and single-cell B cell receptor sequencing revealed that long- lived murine Plasmodium-specific MBCs consisted of three populations: somatically hypermutated, classically defined IgG+ (IgG+), a previously unrecognized population of somatically hypermutated IgM+ (IgMhighIgDlow) MBCs and an unmutated IgD+ (IgMlowIgDhigh) MBC population. Surprisingly, Plasmodium-specific IgM+ antibody dominated the early response to a secondary infection. Further analyses revealed that upon rechallenge, IgM+ MBCs rapidly form two antibody-secreting populations: T cell-independent plasma cells and T-dependent IgM+ and IgG+ plasmablasts. IgM+ MBCs are therefore rapid, plastic, first responders to Plasmodium rechallenge and should be targeted by vaccine strategies. We are now poised to further characterize these and other Plasmodium-specific B cell populations to determine their unique contributions to protection against malaria in both humans and relevant murine models. The central hypothesis of this application is that the development of functionally heterogeneous yet synergistic populations of memory B cells will be required for vaccine-mediated protection to Plasmodium. The goals of this proposal are to identify the molecular and cellular mechanisms that lead to the formation of these distinct MBC subsets and to determine how these cells contribute to protection against malaria in mice and humans. This innovative approach could provide the information required to develop the first effective vaccine against malaria.