The Fine Structure of Replicating Simian Adenoviruses in LLC-MK2

Abstract

The simian adenoviruses SV20, SV30 and SA7 have been found to be oncogenic in the Syrian hamster. The growth characteristics and replicative cycle of these viruses in tissue culture therefore appeared appropriate to investigate. Cesium chloride purified simian adenovirus with an infectivity titer of 100 TCID50, was inoculated into monolayers of LLC-MK2 cells. Cells were fixed in osmium tetroxide and embedded for ultrastructural studies at 1, 3, 6, 9, 18, 24, 48, 72, 120 and 192 hours post-infection.At the first hour post-infection, virus particles were adsorbed to the plasmalemma and found within the peripheral cytoplasm of many LLC-MK2 cells (Fig. 1). Although the first detection of infectious virus occurred at 14 hours and infectivity titers did not reach a maximum until 30 hours, intranuclear virus particles were observed by 3 hours in typical adenovirus crystalline array (Fig. 2) by means of electron microscopy. These typical honeycomb arrayed virus particles at 3 hours provided evidence of significant replication in approximately 5 percent of tissue culture cells examined. Simultaneously, a classical nuclear inclusion manifested by peripheral condensation of nuclear chromatin was evident by light microscopy. As early at 6 to 9 hours, unusual intranuclear concentric membranes formed “tubes” which contained linear arranged virus particles (Fig. 3). In transverse or tangential sections, these “tubes” appeared cochlear-like in shape. In longitudinal section, these intranuclear tubular structures contained individual virus particles at various stages of maturation in a linear arranged order. This arrangement resembled “peas in a pod”.