Kazuya Akatsuka

Electron microscopic study on vaccinia virus release

SummaryFL cells infected with the IHD-W strain of vaccinia virus were studied by scanning and transmission electron microscopy. A large number of naked virus particles were found to accumulate beneath the host cell plasma membrane and to protrude from the cell surface. It was seen in some cases that naked viral particles were released by budding not only from the cell surface but also from the surface of cytoplasmic packets which were seen along the cell periphery.

Tubular Structures Detected in the Nuclei of Human Embryonal Lung Fibroblasts Infected with Human Cytomegalovirus

Various viruses with icosahedral capsids have been reported to produce cylindrical or tubular structures in cells infected with these viruses. For example, some viruses belonging to the papovavirus family (9, 18, 27) and many viruses of the herpesvirus family are known to form such structures in infected nuclei (1, 2, 4, 5, 11, 15, 21, 25, 31-33). The structures are regarded as aberrant forms of these viruses, resulting from mistakes in the assembly of capsid subunits. This report is concerned with findings concerning tubular structures in cells infected with a freshly isolated strain of human cytomegalovirus. The strain of human cytomegalovirus (HCMV) used in this experiment was isolated in 1978 by one of the authors (S.N.) from the urine of a patient with fever and signs of liver dysfunction 40 days after receiving a renal transplant, and was named OU-1. The AD-169 strain was used as a reference strain. Human embryonal lung fibroblast (HEL) cells used for isolation and subsequent passages of the OU-1 strain were kindly supplied by Dr. M. Takahashi, Research Institute for Microbial Diseases, Osaka University. These cells were cultivated in Eagles minimum essential medium (MEM) supplemented with 10% fetal bovine serum. HCMV infected cells were usually prepared for electron microscopic observations as follows. Normal HEL cell suspensions and those of severely infected cells were mixed at ratios of 5:1 to 10:1, diluted with fresh medium so as to contain approximately 106 cells/ml and inoculated into fresh culture bottles. When most of the cells in the monolayer cultures developed cytopathic effects, the cells were scraped into MEM and centrifuged at low speed. The resulting pellet was prefixed for 1 hr in 1% glutaraldehyde in 0.1 M cacodylate buffer (pH 7.2), washed well, postfixed with 1% osmium tetroxide in 0.1 M s-collidine buffer, dehydrated and embedded in epoxy resin. Sections made with a diamond knife were stained with uranyl acetate and lead citrate and observed under a Hitachi H-700H electron microscope. With respect to the ultrastructure of HCMV infected cells, several characteristic morphological changes distinct from those observed in other human herpesvirus infected cells have been presented (6, 20, 25), and these changes could easily be

Ultrastructures of Leuco-Adsorption on Monolayers Infected with Influenza Virus

Leuco‐adsorption occurring in influenza virus infected‐cell cultures was studied morphologically to clarify the mechanisms of adsorption of leukocytes. Among the various types of chicken leukocytes studied, such as lymphocytes, monocytes, granulocytes, and thrombocytes, all were found to adhere to the virus‐infected cells. The adsorption seems to occur through at least two processes, one is mediated by microvilli (microvillus‐attachment), and the other is direct adherence of both cells (cell‐to‐cell‐attachment). In the former, the leukocytes are bound to the microvilli protruding from the infected MDCK cells and in the latter both cell membranes attach directly. In the cell‐to‐cell‐attachment, there was an electron‐lucent gap of about 12 nm in width in the intermembranous space of the junctional regions. This region was similar morphologically to the gap junction. As a result of leuco‐adsorption no cytolytic effects occurred in the MDCK cells under the experimental conditions.

Unusual production of Gross murine leukemia virus (GrMuLV) in murine lymphoblasts as studied by electron microscopy.

The production of murine leukemia virus (GrMuLV) in a clonal line of Gross virus-induced murine lymphoblasts was studied by scanning and transmission electron microscopy (SEM and TEM, respectively). Most virions were found distributed randomly over the cell surface membrane as single particles. In rare cases, a large cluster of virions had accumulated on the cell surface membrane. SEM showed that, in such clusters, many virions with diameters ranging from 88 to 140 nm were tightly interconnected with one another, some appearing to bnd directly from the cell surface membrane. TEM revealed an abnormally large virion (165 nm in diameter) present in a cluster. The core structure of this virion was an open circle in which both free ends were further circled within the circle. The core length was longer (485 nm) than normal (220 nm). These observations suggest that GrMuLV can be produced in a cluster accompanying simultaneous production of an abnormally large virion with an atypical core structure.