Serum specimens were collected from 125 pigs on Miyako Island, 112 pigs on Ishigaki Island, and 42 pigs on Kume Island from 2005 to 2007, and 54 pigs on Yonaguni Island from 2006 to 2007. Their sera were tested for Japanese encephalitis virus (JEV) antibody by hemagglutination inhibition (HI) assay. Five serum samples (4.5%) from Ishigaki Island were positive for HI antibody, and 4 of the 5 samples were positive for 2-mercaptoethanol- sensitive antibody (IgM Ab). All samples from Miyako, Kume, and Yonaguni Islands were negative for HI antibody. Our results indicate that JEV transmission activity was extremely low on Miyako, Ishigaki, Kume, and Yonaguni Islands. The JEV genome (JEV-RNA) was detected from the sera of one pig on Ishigaki Island. The partial gene of the E region (151 nt) was analyzed phylogenetically. The analysis showed that the new JEV-RNA belonged to genotype 3 and was closely related to JEV strains isolated in Taiwan from 1985 to 1996. It was suggested that JEV previously introduced from Taiwan had been maintained on Ishigaki Island.

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