To understand the distribution of Japanese encephalitis antibodies among preschool children in rural Taiwan, all three- to six-year-old children residing in Hsinchu, Taichung, Pingtung and Hualien Counties were selected for seroepidemiolonical investigation. The plaque reduction neurtralization test (PRNT) was used with a total of 2,281 children. The total positive rate of Japanese encephalitis neutralization antibodies found was 67% (1,520/2,281). After a series of three doses of Japanese encephalitis vaccine, the four-year-old children of all four counties showed the lowest neutralization antibody positive rate: these were 45% (56/125) in Hsinchu; 51% (57/111) in Taichung; 30% (39/128) in Pingtung and 66% (67/102) in Hualien. These figures are significantly lower than those for the other age group of the children immunized in each county. (p=0.0001, Chi-square test). Whether or not children had been immunized against Japanese encephalitis, the neutralization antibody positive rate went up after age five, reaching the highest rate (around 82 to 91%) at age six. This shows that in the areas of study, spontaneous infection with Japanese encephalitis generally existed. The antibody positive rate of the 47 non-immunized children also went up with age. The neutralization antibody positive rate was 67% (1,122/1,665) for children who had had three doses of vaccine: 66% (64/97) for children with two doses; 33% (4/12) for children with only dose and 40% (19/47) for children not immunized. The latter three groups had significantly lower rates than the ones who had had three doses of the vaccines (p <0.0001, Chi-square test for trend). This shows that the spontaneous infection rate was relatively high. However, children with two or more doses of vaccine had higher protection. It appears that vaccines still play a very important role in the current prevention and control program against Japanese encephalitis.