Validation of serological markers of recent exposure to Plasmodium vivax in Cambodia: a longitudinal cohort and cross-sectional study.

BACKGROUND: Asymptomatic Plasmodium vivax infections undermine malaria control by sustaining transmission. This study aimed to validate an eight-marker serological panel (selected from 14 antigens) of P. vivax exposure to detect individuals likely harbouring hypnozoites in rural Cambodia, and to compare it with molecular surveillance for identifying at-risk populations in low-transmission settings. METHODS: Two studies were conducted in Mondulkiri, Cambodia. In a longitudinal cohort (N = 471), participants were tested monthly for one year for P. vivax using qPCR. At 12 months, IgG responses to 14 antigens were assessed, and Random Forest models ('Cambodia-specific' and 'global') were used to classify previous infection, with PCR as the reference standard. In a cross-sectional survey (N = 3538), risk factors for sero- and PCR positivity were assessed using generalized linear mixed-effects models. FINDINGS: PCR detected P. vivax in 29.9% (141/471) of participants over the 12-month follow-ups, while 31.8% (150/471) were classified as seropositive at month 12. Of 141 PCR-confirmed cases, 95 were seropositive (67.4%) and 46 seronegative (32.6%) under the 8-antigen panel. The 'global' model achieved an AUC of 0.81, with 83.3% specificity and 69.5% sensitivity for infections within nine months, increasing to 82% sensitivity for last-month infections. In the cross-sectional study, serology identified more positives than PCR, reflecting detection of exposure not captured by PCR. INTERPRETATION: This study provides the first validation of serological markers of P. vivax exposure in Cambodia, showing that such tools can identify recent and asymptomatic infections, though sensitivity was below target. Serology-based surveillance and strategies such as PvSeroTAT could accelerate malaria elimination. FUNDING: NIH/NIAID, ICEMR Asia-Pacific U19AI129392.