Laboratory and Epidemiology Communications Genotypic Analysis of Japanese Encephalitis Virus Strains Isolated from Swine in Tokyo, Japan

Japanese encephalitis (JE) is a serious viral encephalitis caused by a mosquito-borne flavivirus, the Japanese en- cephalitis virus (JEV). In the metropolitan area of Tokyo, the number of JE cases has decreased dramatically. The number of JEV isolates obtained from swine serum samples and the number of Culex tritaeniorhynchus isolates collected at sentinel points also have been decreased. Furthermore, in recent years, both the number of JEV-infected swine and the number of households that breed swine have decreased sharply. Because of these facts, there is a frequent misperception that JEV is no longer passed between mosquitoes and swine in metropolitan Tokyo. However, swine are still seroconverted to JEV every year, suggesting that JEV still exists in Tokyo, and thus that JEV still has the potential to be a serious cause of illness. In the present study, we analyzed JEV strains isolated in the metropolitan region of Tokyo from 1965 to 2000. Eighty- two JEV isolates were obtained from swine serum samples from 1965 to 2000 (Table 1). The swine blood samples were collected in the Tama region of Tokyo and the serum was separated. The virus was isolated by inoculating the swine serum into the brains of suckling mice. Of the 82 isolates, 13 isolates were selected based on the differences in their hemagglutination patterns with goose blood cells at VAD 6.0 - 7.0, and the differences in their responses to anti-JEV antibodies. A list of the 13 JEV strains used in the analysis are shown along with other 6 strains in Table 2. RNA was extracted from JEV-infected mice brain by SepaGene RV-R (Sanko-Junyaku Co., Ltd., Tokyo, Japan). The JEV genome was amplified by RT-PCR with a pair of primers, JEV#5 (5´-CTGGG AATGG GCAAT CGTGA C-3´ (987-1007)) and JEV#6 (5´-TTTGA GGGTT ATCGA AGGAG CAT-3´ (1514-1492)). RT-PCR amplicons were