Electron Microscope Study of the Development of Japanese B Encephalitis Virus in PS Cells

Abstract

The ultrastructure and developmental process of Japanese B encephalitis virus were studied by electron microscopy in tissue culture of porcine kidney stable (PS) cells infected with the Mukai strain of Japanese B encephalitis virus. The virus and PS cells were kindly supplied by Dr. Y. Kanda Inoue at the Virus Institue of Kyoto University. PS cells were cultivated in bottles containing 10% calf serum and 0.5% lactalbumin hydrolyzate in Earle's balanced solution. Titration of cell-associated virus was estimated by cytopathic effect. At various intervals following infection, the cells were fixed in buffered 1% osmium tetroxide solution, embedded in methacrylates and cut on a Leitz ultramicrotome. After the sections were stained in saturated uranyl acetate solution, they were observed in the Hitachi type HU 11 electron microscope.
Japanese B encephalitis virus particles were hexagonal in thin sections and approximately 40 mil in the longest diameter, composed of the outer membrane, 30 A in thickness, viroplasm, 30 A in thickness and an electron-dense nucleoid, 25 mit in diameter. After the virus particles developed on the wall of the cytoplasmic vacuole, they were densely packed in the vacuole usually in random arrangement and rarely in crystalline arrays. The vacuole containing the virus particles gradually migrated to the cell surface and liberated the particles to the exterior of the cell through a narrow canaliculus which was formed between the vacuole and cell surface.