Immunodominant B-cell epitopes for precision surveillance of Plasmodium vivax malaria

Asymptomatic Plasmodium vivax infections sustain transmission and derail elimination efforts, yet these hidden reservoirs evade current diagnostics. Here, we integrated computational immunology with empirical serology to discover high-performance B-cell epitopes. From a library of 142 P. vivax proteins, we prioritized six antigens (AMA1, CSP, DBP, MSP1, MSP9, and P12), synthesized 64 epitopes, and screened them against serum from P. vivax-infected cases from the China-Myanmar border (CMB) via ELISA, with multiplex immunoassay validation across Brazil, the Solomon Islands, and Papua New Guinea samples. Rigorous assessment of specificity, sensitivity, and asymptomatic discrimination, along with antigenic conservation analyses in CMB clinical isolates and global homologues, yielded ten immunodominant epitopes. Four exhibited outstanding performance, namely, MSP9_{_603-609} (90.4% sensitivity, AUC = 0.980), AMA1_{_457-466} (AUC = 0.910), MSP9_{_458-464}(AUC = 0.802), and MSP9_{_425-433} (AUC = 0.898), all with absolute specificity against P. falciparum (P < 0.0001). Crucially, the MSP9_{_384-389} epitope distinguished asymptomatic infections (AUC = 0.756, P < 0.0005). Despite variable detection rates across regions (2.2-22.2%), with the highest rates in Brazil, all the epitopes remained 99-100% conserved globally, reflecting strong functional immunological constraints. These epitopes enable unprecedented detection of both symptomatic and asymptomatic P. vivax infections - addressing a critical surveillance gap - while their scalability and species specificity make them ideal for elimination campaigns, particularly in low-resource settings. This work lays the foundation for next-generation serological tools to accelerate vivax malaria eradication.