Growth of Japanese encephalitis virus in Culex tritaeniorhynchus cell cultures

Ovarian cells from the mosquito Culex tritaeniorhynchus established in continuous tissue cultures have been found to support the growth of Japanese encephalitis virus with titers reaching as high as 10(7.36) MICLD50 per 0.03 ml. Virus-cell cultures were serially subcultured 60 times over 300 days and the virulence of the virus was determined after each passage. A gradual loss in virus titer was observed, with titers always higher in the medium than in the cells. Cytopathogenic effects were not observed in the infected cell cultures, either in primary or subsequent passages. However, the cell growth rate suggested that the cells may have been metabolically or mechanically damaged. The chromosome complement of the cells remained unchanged. Direct-fluorescent antibody studies during early and late passages revealed a loss in virus infectivity from chronically infected cells which may have been due to a gradual decrease in virus replication as the passages increased. The infectivity of virus particles, however, returned to previous levels after passage into fresh cell cultures. The loss of virus particles during serial passage may be correlated with the condition of the cells.