Growth of Japanese encephalitis virus in tissue cultures

The development of Japanese encephalitis virus (JaGAr 01 strain) in the cell cultures was investigated by the methods of fluorescent antibody technic and electron microscopy. The results obtained were as follows: 1) Cytopathic effect after virus inoculation was observed in BHK-21 and HK cells at 48 hr, and Vero and HmLu cells at 72 hr, respectively. The highest titers of infectivity in the media were demonstrated at 46 to 96 hr after virus inoculation and significant correlation was ovserved between infective titers and HA. 2) In infective cell cultures, viral specific fluorescences were observed in BHK-21 cells at 10 hr, in HmLu cells at 14 hr, in Vero and HK cells at 16 hr, and in CE cells at 18 hr, respectively. At initial stage of infection, the fluorescence appeared around the perinuclear area of a single cell and developed into cytoplasmic area with granular or radial form. These cells formed gradually into a mass. At 24 hr after virus inoculation, a mass of 10 to 20 cells was predominantly observed. 3) In electron microscopy, viral particle was approximately 40nm in diameter. It was composed of an outer membrane, viroplasm, and an electron dense uncleoid which was 25 nm in diameter. Viral particles, isolated or clustered, were observed at extracellular space near the cell membrane, in intracellular vacuole, in phagosome, and in enlarged endoplasmic reticulum fulled with granular matrix, but not in nucleus.