A detection survey for malaria infection by routine microscopy and polymerase chain reaction (PCR) assay was conducted on 336 inhabitants of two villages in Khammouane Province, Lao People's Democratic Republic (Lao PDR), in July 1997. Malaria infection was demonstrated in 58 (17.3%) subjects by microscopy and in 117 (34.8%) by PCR assay. Specimens positive by both methods were frequent in young villagers, suggesting the presence of many subclinical infections in older persons. The most common species of malaria parasite was Plasmodium falciparum (82.9%). Polymerase chain reaction assay detected mixed infections with 2-4 species in 27 specimens (23.1%). The results demonstrate that there are many subclinical malaria infections with low parasite level and infection with all four human malaria species in Lao PDR.