Ecological complexity of zoonotic malaria in macaque natural hosts

Macaques are important reservoirs of zoonotic malaria in Southeast Asia. Although cross-sectional malaria surveys have been conducted in macaques, little is known about intra-host infection dynamics and host variation in susceptibility to infection in these infectious reservoirs. We performed a longitudinal monitoring of Plasmodium and Hepatocystis infections by microscopy, species-specific polymerase chain reaction (PCR) and targeted amplicon deep sequencing (TADS) in three long-tailed macaques and 20 pig-tailed macaques in two districts of Narathiwat Province, southern Thailand. In total, 104 macaques' blood samples were obtained during 5 visits with sequential time intervals of 9, 4, 7 and 12 months. Transiently patent Plasmodium infections with low parasite density ( ≤ 1,050 parasites/µL) occurred in 7 pig-tailed macaques, while PCR and TADS diagnosed infections in 45 (43.27%) blood samples with one or more species of parasites, including Plasmodium knowlesi, P. cynomolgi, P. inui, P. fieldi, P. coatneyi, P. aff. coatneyi and Hepatocystis sp. in one long-tailed and 12 pig-tailed macaques. Compared with PCR, TADS additionally detected co-infecting species in 22 of 45 ( 48.89%) samples. Although living in close proximity to other infected macaques, seven macaques were free from infection during the 32-month period. Infections for 4 to 32 months with malaria parasites carrying identical complete mitochondrial genome sequences were reaffirmed in 10 macaques. Potentially new infections were detected transiently or over a long period during the course of the infections while competitive exclusion seemed to occur between Hepatocystis sp. and Plasmodium taxa. Macaques' Duffy phenotypes did not influence differential susceptibility to Plasmodium infections. These results suggest the ecological complexity of hemoparasite infections in natural reservoirs of zoonotic malaria. The long period of Plasmodium infections in macaques could affect the transmission and control of the disease.