PROJECT SUMMARY/ABSTRACT There is a pressing need for sensitive, field-deployable malaria surveillance tools to guide the optimal deployment of interventions in elimination zones, where transmission is focal and malaria risk heterogeneous. Ultrasensitive molecular methods have revealed that most malaria infections in these settings are subclinical, low-density infections, which are not detectable by standard diagnostic tests, serving as a silent reservoir for malaria transmission. However, ultrasensitive molecular methods are expensive, time-consuming, require a well- equipped laboratory, and thus cannot provide point-of-contact results, limiting their utility in remote malaria- endemic areas. An ideal surveillance tool would not only measure parasite prevalence, but also estimate recent malaria exposure, providing a more robust characterization of malaria in a population. Antibody biomarkers are promising targets for malaria surveillance in this setting because they can indicate cumulative exposure and are easily integrated into existing point-of-care platforms. However, serological markers remain underutilized due to a lack of well-defined targets or consensus on how to interpret results. Many of the most seroreactive malaria antigens are also the most polymorphic, confounding results based on reactivity to reference-strain proteins. This work proposes to leverage this diversity to identify informative antibody biomarkers to estimate malaria exposure. Based on our preliminary data, the central hypothesis of this proposal is that individuals exposed to low-density malaria infections have unique serological profiles that can be used for rapid evaluation of recent and current exposure. This hypothesis will be tested with the following specific aims: 1) Identify antibody biomarkers for current and recent (6 months) subclinical Plasmodium falciparum and Plasmodium vivax exposure, and 2) validate novel peptide targets against malaria antigens using a multiplexed bead-based immunoassay format. In Aim 1, antibody biomarkers will be identified by measuring seroreactivity of matched exposed and unexposed individuals on novel ultra-dense peptide arrays populated with sequences from Plasmodium falciparum and Plasmodium vivax antigen variants from both reference strains and geographically- relevant field isolates. In Aim 2, down-selected peptides will be synthesized in bulk and conjugated to barcoded magnetic beads for a multiplexed fluorescence-based immunoassay. Seroreactivity to peptide targets will be measured on well-characterized samples from individuals with known malaria exposure as well as endemic controls. The proposed work is the first step toward developing a robust point-of-contact test that can be used to characterize malaria transmission and risk, ultimately enabling better and more precise targeting of interventions.