Yoshikatsu Ozaki

Intranuclear crystal formation of poliovirus: Electron microscopic observations

Abstract Poliovirus-infected FL cells, incubated at 28°C, were examined by electron microscopy. The immunofluorescent antibody technique was employed for detection of the intracellular viral antigen. Electron micrographs of cells from infected cultures showed both extensive cytoplasmic and nuclear alterations. Virus crystals were observed not only in the cytoplasm but also in the nucleus of cells 14–20 hours after infection. Intracytoplasmic crystals consisted of dense particles, approximately 27 mμ in diameter, suggesting mature virus particles, while intranuclear crystals were composed of empty particles with a less dense central area. Nuclei were stained specifically by fluorescent antibodies as small brilliant foci, suggesting that the intranuclear crystals possess the same antigenicity as the viral antigen. As judged by their size, structure and antigenicity, the intranuclear virus-like particles probably represent an incomplete form of the virus.

Studies on the Neutralization of Japanese Encephalitis Virus

The neutralization reaction of Japanese encephalitis virus with early serum was compared with that with late serum. The analysis of antiserum by Sephadex G200 gel filtration indicated that the neutralizing activity in early serum was present only in the IgM fraction, while that in late serum was present only in the IgG fraction. The antibody dose response curves in early serum were characterized by the early and high appearance of a persistent fraction. This fraction was found to consist of an infectious virus antibody complex (sensitized virus) which was neutralized by anti-IgM serum. The amount of virus neutralized by anti-IgM serum varied with the concentration of antiviral antibody employed for the sensitization. In contrast, it was a characteristic of the neutralization by late serum that the residual infectivity was inversely related to the concentration of antibody in the serum, resulting in a low level of a non-neutralized virus fraction. Therefore, the maximal reduction of residual infectivity by anti-IgG serum was attained under an optimal ratio of antibody to virus. Virus sensitized with early serum had a blocking effect against a high concentration of late serum antibody, but was neutralized by anti-IgM serum. Virus sensitized with an insufficient amount of late serum antibody was neutralized not only by high concentrations of late serum antibody, but also was supersensitized by early serum antibody. Since the sensitized virus which had been adsorbed on host cells was still neutralizable by anti-γ-globulin, aggregation seemed to be excluded as the main factor in the mechanism of neutralization by anti-γ-globulin serum.

Studies on neutralization of Japanese encephalitis virus. IV. Effect of anti-cellular serum on the neutralization of sensitized virus by anti-rabbit IgG serum.

SummaryThe effect of anti-cellular rabbit serum (ACRS) on the neutralization of sensitized Japanese encephalitis virus (JEV) by anti-rabbit IgG serum was examined to elucidate the interaction between virus-antibody complex and the surface of the host cells during the process of neutralization.ACRS had no effect on the adsorption of either sensitized or non-sensitized virus, but was able to restore the lost infectivity of sensitized virus which occurred during the process of neutralization by anti-rabbit IgG serum. This restoration of infectivity was found to take place not only by the addition of ACRS to the reaction mixtures (virus-antibody, anti-rabbit IgG complex) but also by pretreatment of the host cells with ACRS. Although the restoration of lost infectivity varied in magnitude with the concentration of ACRS used, it never exceeded the infectivity titer of the sensitized virus before incubation with anti-rabbit IgG serum. This result suggests that ACRS has no ability to reverse the neutralization by anti-viral serum. Since the ACRS reacted only with anti-rabbit IgG serum treated sensitized virus, resulting in an increase of the number of infectious centers, the restoration of lost infectivity was explained as being due to the enhancement of adsorption of sensitized virus to the host cells by bridge formation of anti-rabbit IgG antibody between them.

ELECTRON MICROSCOPIC STUDIES ON THE YOSHIDA SARCOMA CELLS INFECTED WITH ECTROMELIA VIRUS

The morphological changes of Yoshida sarcoma cells infected with ectromelia virus were studied with the electron microscope using the thin sections and the phase contrast microscope.Phase contrast microscopic observations: The inclusion bodies were clearly found in the cytoplasm at 72 hours after inoculation. They were frequently revealed as an accumulation of elementary bodies and varied in size. The serious necrotic changes were seen in the cytoplasm at the later stage, and the diffuse distribution of elementary bodies was observed in them.Electron microscopic observations: The inclusion bodies and matrix regions were found in the infected cytoplasm as they were in the case of other tissues. The inclusion bodies were sharply outlined from the cytoplasm. The elementary bodies embedded within the inclusions showed polymorphological appearances and their density was less than that in mature particles. Though empty circles were found in matrix region, it was not proved that the relationship was present between these bodies and mature virus particles.