OBJECTIVE: To investigate the change of viral titers under different conditions in cultured cells persistently-infected with different strains of Japanese encephalitis virus (JEV) and find out the factors that influence viral multiplication. METHODS: JEV JaGAr-01 and Nakayama wild strains were used to infect human hepatoma cell line KN73 respectively, and the persistent infection model was established. Viral titers were examined by plaque methods using BHK cells. Human nerve fibroblastoma cell line IMR-32 was infected with the strains of the virus that can cause persistent infection, and the thermal sensitivity of the viral strains was measured at 30 degrees C and 37 degrees C. KN73 cells persistently infected with JEV were infected with two JEV strains respectively, and viral superinfection was studied. To explore the replication of the persistently-infected viruses, KN73 and IMR-32 cells were infected with the viral strains. RESULTS: Two persistently infected viral strains did not show any thermal difference. The results of superinfection were that the viral titers of JaGAr-01 strains were 1.3 and 8.8 percent of the control, respectively, and the viral titers of Nakayama strain were 80 and 1.7 percent of the control, respectively. JaGAr-01 wild strains, Nakayama wild strains and their persistently-infected strains infected KN73 and IMR-32 respectively. The replication of the persistently-infected strains was obviously lower than the wild strains in KN73 cells, however, in IMR-32 cells their replication was similar. CONCLUSIONS: The two strains of JEV were not found to be temperature-mutant. It is possible that mutant viruses containing DI particles exist in JEV persistently-infected strains. In different cells there may be different host factors hindering the replication of the persistently-infected strains.