Detection of Plasmodium infections in macaques from areas endemic for brugian filariasis in Belitung District, Indonesia

Background Malaria caused by Plasmodium knowlesi and lymphatic filariasis caused by Brugia malayi are mosquito-borne infections with non-human primates as reservoirs. P. knowlesi has emerged as a significant cause of human malaria in Southeast Asia over the past two decades. Belitung district, Indonesia, was until recently assumed to have eliminated B. malayi until infections were detected in humans and long-tailed macaques. To investigate whether the local reservoir of B. malayi is also a reservoir for malaria we screened macaques from 4 areas in Belitung for malaria parasites. Methods and findings Blood samples from 163 long-tailed macaques (Macaca fascicularis) that had been tested for B. malayi were examined by quantitative PCR assays specific for Plasmodium spp., P. knowlesi, P. inui, P. coatneyi and P. cynomolgi. A total of 130 macaques (79.8%) tested positive in the pan-Plasmodium qPCR assay. Plasmodium inui was most prevalent (41.7%), followed by P. knowlesi (38.7%), P. coatneyi (24.5%) and P. cynomolgi (13.5%). Multiple species infections, with 2-3 Plasmodium species were detected in 37% of macaques. Notably, 20 (91%) of 22 B. malayi-positive macaques were co-infected with at least one Plasmodium species. We sequenced the complete mitochondrion from 9 samples diagnosed by qPCR as mono-infections. Phylogenetic analysis confirmed 7 as P. knowlesi, and the other two as P. inui and P. coatneyi. Phylogenetic and pairwise analysis revealed that P. knowlesi isolates from Belitung were closely related to each other and to P. knowlesi from humans and monkeys from Thailand, Malaysia and Indonesia. Conclusions Molecular evidence shows high prevalence of zoonotic malaria parasites in macaques from Belitung, emphasizing the risk of human transmission. Increased surveillance, improved diagnostics, and targeted interventions are needed to prevent zoonotic spillover of P. knowlesi as it has been observed for B. malayi in Belitung.