中华医学杂志（英文版）2005，Vol.118Issue(6)：451-459,9.
Severe acute respiratory syndrome coronavirus persistence in Vero cells
Severe acute respiratory syndrome coronavirus persistence in Vero cells
摘要
Abstract
Background Several coronaviruses establish persistent infections in vitro and in vivo, however it is unknown whether persistence is a feature of the severe acute respiratory syndorme coronavirus (SARS-CoV) life cycle. This study was conducted to investigate viral persistence.Methods We inoculated confluent monolayers of Vero cells with SARS-CoV at a multiplicity of infection of 0.1 TCID50 and passaged the remaining cells every 4 to 8 days for a total of 11 passages. Virus was titrated at each passage by limited dilution assay and nucleocapsid antigen was detected by Western blot and immunofluoresence assays. The presence of viral particles in passage 11 cells was assessed by electron microscopy. Changes in viral genomic sequences during persistent infection were examined by DNA sequencing. Results Cytopathic effect was extensive after initial inoculation but diminished with serial passages. Infectious virus was detected after each passage and viral growth curves were identical for parental virus stock and virus obtained from passage 11 cells. Nucleocapsid antigen was detected in the majority of cells after initial inoculation but in only 10%-40% of cells at passages 2-11. Electron microscopy confirmed the presence of viral particles in passage 11 cells. Sequence analysis at passage 11 revealed fixed mutations in the spike (S) gene and ORFs 7a-8b but not in the nucleocapsid (N) gene. Conclusions SARS-CoV can establish a persistent infection in vitro. The mechanism for viral persistence is consistent with the formation of a carrier culture whereby a limited number of cells are infected with each round of virus replication and release. Persistence is associated with selected mutations in the SARS-CoV genome. This model may provide insight into SARS-related lung pathology and mechanisms by which humans and animals can serve as reservoirs for infection.关键词
severe acute respiratory syndrome/coronavirus/infection/microbiological techniquesKey words
severe acute respiratory syndrome/coronavirus/infection/microbiological techniques分类
医药卫生引用本文复制引用
Gustavo Palacios,Omar Jabado,Neil Renwick,Thomas Briese,W. Ian Lipkin..Severe acute respiratory syndrome coronavirus persistence in Vero cells[J].中华医学杂志（英文版）,2005,118(6):451-459,9.基金项目
This work was supported by grants from the National Institutes of Health (No. AI 51292), and the Ellison Medical Foundation (No. ID-SS-0030). （No. AI 51292）
