Based on the reactivities of monoclonal antibodies, we have reported previously that 25 Japanese encephalitis virus (JEV) strains fell into five antigenic groups: Nakayama, Beijing-1, Kamiyama, 691004 and Muar. In the present study, we compared the nucleotide and deduced amino acid sequences of the structural proteins of these strains. Four strains, Nakayama-RFVL, Beijing-1, Kamiyama and 691004, shared > 96.4% identity in the nucleotide sequence and > 94.7% identity in the amino acid sequence. However, the Muar strain had only an 80.3-87.7% homology for nucleotides and an 85.7-93.5% homology for amino acids, compared with the other four strains. The following strain-specific amino acid differences were found in the envelope protein: two in Nakayama-RFVL; three in Beijing-1; one in Kamiyama; five in 691004; 42 in Muar. These findings showed that the Muar strain was the most structurally different from the other four strains, and that it would be possible to compare antigenic differences among the JEV strains by using monoclonal antibodies which showed different specificities.