Temporal trends in Plasmodium vivax diversity in eastern Cambodia evidence declining transmission

Elimination of P. vivax is challenging due to its dormant liver stages (hypnozoites), which can reactivate weeks or months after the primary infection, causing relapses and ongoing transmission of the parasite. In 2019, Cambodia introduced primaquine for the radical cure of P. vivax malaria in the effort to accelerate elimination of this species. We used parasite genotyping to assess the impact of primaquine intervention on transmission reduction. Genotyping was conducted on 182 symptomatic P. vivax isolates collected in eastern Cambodia in 2014, 2015, 2019 and 2023. A panel of 93 microhaplotype markers (vivaxGEN panel) was genotyped using Illumina sequencing. Population genetic measures were applied to determine infection diversity and relatedness (identity-by-descent (IBD)) in each year. The genetic results correlated well with clinical cases for the study years.The percentage of polyclonal infections was 5% in 2023 compared to 22-48% in earlier years (p<0.05) suggesting substantial reduction in superinfection. The number of cases in 2023 also had the highest percentage of infections with IBD >0.95 with one or more other infections (81.4% versus 8.9-10.8% in 2014-2019) indicative of inbreeding following population bottlenecking. In 2019 there was a spike in polyclonal infections (48%) and population diversity following local interruption to critical malaria control services. Our findings illustrate the potential of microhaplotype genotyping to inform on P. vivax transmission to assess intervention efficacy. In eastern Cambodia, the data provides evidence to support of widespread use of radical cure for patients with P. vivax malaria.