Early events of SARS coronavirus infection in vero cells

Abstract

An isolate from a patient in the recent severe acute respiratory syndrome (SARS) outbreak in Singapore was used to infect Vero E6 cells. This study concentrated on the first 30 min of infection. It was discovered that the SARS coronavirus attached, entered, and uncoated the nucleocapsids, all within a 30-min period. At 5 min after infection, several virus particles lined the Vero cell plasma membrane. Virus particles were at various stages of fusion at the cell surface, since entry was not a synchronised process. After entry (10 and 15 min), spherical core particles moved into the cytoplasm within large vacuoles. Quite surprising at such early stages of infection (20 min), a virus-induced change in the infected cells was evident. The induction of myelin-like membrane whorls was obvious within the same vacuoles as the core particles. The significance of this virus-induced change is unknown at this stage. By 25-30 min postinfection (p.i.), the spherical core particles appeared to be disassociating and, in their place, doughnut-shaped electron-dense structures were observed. These could be the virus genomes together with the helical nucleocapsids. They were no longer in large vacuoles but packaged into smaller vacuoles in the cytoplasm, and occasionally in small groups.

Implication of proprotein convertases in the processing and spread of severe acute respiratory syndrome coronavirus

“…This result raised the possibility that the kinetics of SARS-CoV syncytia formation or cell to cell spread is affected by proS processing. Taken together, our work strongly suggests that proS processing and viral spread are interconnected, and that processed proS may be required for direct cell-cell spread and/or for CPE initiation resulting in enhanced secretion of mature virions [4]. Similarly, in our experiments, dec-RVKR-cmk directly or indirectly reduced the appearance of the $80 kDa protein, which in turn may have resulted in the abrogation of the cytopathicity associated with virus spread.…”

Establishment of Vero E6 cell clones persistently infected with severe acute respiratory syndrome coronavirus

“…This culture seems to accord with the histopathologic findings in intestinal samples from patients [13,14]. In sharp contrast to the LoVo cell line, the Vero E6 cell line undergoes a lytic infection with characteristic refractile rounding CPE [16,17] that seem to correspond to the features of multinucleated pneumocytes resulting from local replication of SARS-CoV in the lungs of patients [11,12].…”

Molecular biology of severe acute respiratory syndrome coronavirus

“…Following S protein-mediated fusion of the viral envelope with the host cell membrane (see Update) and release of the viral genome RNA into the cytoplasm of the infected cell [23], SARS-CoV genome expression begins with the (cap-dependent) translation of the genomic RNA (mRNA 1; Figure 1). The translation product that is encoded by ORF1a is a protein of 4382 amino acid residues and is called polyprotein 1a (pp1a).…”