Some nonhuman primate Plasmodium species including P. knowlesi and P. cynomolgi can cross-transmit from macaque natural hosts to humans under natural infection. This study aims to retrospectively explore other simian Plasmodium species in the blood samples of symptomatic malaria patients in Thailand. A total of 5271 blood samples from acute febrile patients from 5 malaria endemic provinces and 1015 blood samples from long-tailed and pig-tailed macaques from 3 locations were examined for Plasmodium species by microscopy and species-specific polymerase chain reaction. The Plasmodium mitochondrial cytochrome oxidase 1 (COX1) gene was analyzed by amplicon deep sequencing as well as Sanger sequencing from recombinant plasmid clones to reaffirm and characterize P. inui and P. fieldi. Besides human malaria, P. knowlesi, P. cynomolgi, P. inui and P. fieldi infections were diagnosed in 15, 21, 19, and 3 patients, respectively. Most P. inui and all P. fieldi infected patients had simultaneous infections with other Plasmodium species, and seemed to be responsive to chloroquine or artemisinin-mefloquine. P. inui was the most prevalent species among macaque populations. Phylogenetic analysis of the COX1 sequences from human and macaque isolates reveals the genetic diversity of P. inui and suggests that multiple parasite strains have been incriminated in human infections. Both P. inui and P. fieldi could establish infection in humans under natural transmission. Despite occurring at a low prevalence and mostly co-existing with other Plasmodium species, P. inui infections in humans have a wide distribution in Thailand.