An Electron Microscope Study of the Development of a Mouse Hepatitis Virus in Tissue Culture Cells

Abstract

Samples taken at different intervals of time from suspension cultures of the NCTC 1469 line of mouse liver—derived (ML) cells infected with a mouse hepatitis virus have been studied with the electron microscope. The experiments revealed that the viruses are incorporated into the cells by viropexis within 1 hour after being added to the culture. An increasing number of particles are found later inside dense cytoplasmic corpuscles similar to lysosomes. In the cytoplasm of the cells from the samples taken 7 hours after inoculation, two organized structures generally associated and never seen in the controls are observed: one consists of dense material arranged in a reticular disposition (reticular inclusion); the other is formed by small tubules organized in a complex pattern (tubular body). No evidence has been found concerning their origin. Their significance is discussed. With the progression of the infection a system of membrane-bounded tubules and cisternae is differentiated in the cytoplasm of the ML cells. In the lumen of these tubules or cisternae, which are occupied by a dense material, numerous virus particles are observed. The virus particles which originate in association with the limiting membranes of tubules and cisternae are released into their lumen by a "budding" process. The virus particles are 75 mµ in diameter and possess a nucleoid constituted of dense particles or rods limiting an electron transparent core. The virus limiting membrane is sometimes covered by an outer layer of a dense material. In the cells from the samples taken 14 to 20 hours after inoculation, larger zones of the cell cytoplasm are occupied by inclusion bodies formed by channels or cisternae with their lumens containing numerous virus particles. In the samples taken 20 hours or more after the inoculation numerous cells show evident signs of degeneration.

Ultrastructural and biochemical analyses of hepatitis C virus-associated host cell membranes

“…The same SARS model demonstrated that viral replication occurs preferentially in circular structures known as convoluted membranes (Knoops et al, 2008) which may be precursors of DMVs. These structures were found to be identical to the 'reticular inclusions' first observed in cells infected with mouse hepatitis coronavirus (MHV) more than 40 years ago (David-Ferreira & Manaker, 1965) and subsequently referred to as 'clusters of tubular cisternal elements' (Krijnse-Locker et al, 1994). The small circular units observed in HCV RNA-replicating cells in our study (Fig.…”

Identification of GBF1 as a Cellular Factor Required for Hepatitis C Virus RNA Replication

“…Like HCV, both of them induce membranous replication complexes, and MHV replication complexes are associated with ER membranes (12). In contrast, Sindbis virus, another plus-strand RNA virus, which assembles replication complexes on membranes derived from lysosomes (21), is much less sensitive to BFA (reference 58 and our unpublished observation), suggesting that GBF1 is not required for its replication.…”

Several Rat Cell Lines Share a Common Defect in Their Inability to Internalize Murine Coronaviruses Efficiently

“…Subsequent to adsorption, the murine coronaviruses are presumably internalized into the host cell by absorptive endocytosis (David-Ferreira & Manaker, 1965). To examine the internalization of JHM by the various cell types, JHM-adsorbed cells were incubated at 37 °C for various time periods and the amount of virus internalized was determined as described in the legend to Fig.…”