Liu-Chiu islet, a relatively isolated ecosystem that is free of rice cultivation, has long considered free of Japanese encephalitis (JE). However, a new strain (T1P1) of JE virus was isolated from the mosquito, Armigeres subalbatus, and a rather broad distribution of neutralizing antibody has been reported on the islet, suggesting that the circulating virus could be an attenuated strain. In an assessment on 219 blood samples obtained from residents of Liu-Chiu islet, the positive rate of JEV-specific IgM antibodies decreased with age while that of neutralizing antibodies increased with age. Both antibodies were mainly responsive to the T1P1 strain since higher positive rates and titers of specific neutralizing antibodies are shown in this investigation. Importantly, the T1P1 strain is herein characterized as being broader in neutralizing virus strains, stable in genetic traits, and productive in Vero cells. Taken together, the JE virus strain endemically circulating on Liu-Chiu islet may have served as a natural form of a live-attenuated vaccine. As a result, it possibly can be utilized as a new and effective vaccine candidate in the future.