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Severe acute respiratory syndrome coronavirus persistence in Vero cells.
Chin Med J (Engl). 2005 Mar 20; 118(6):451-9.CM

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 syndrome 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.

Authors+Show Affiliations

Greene Infectious Disease Laboratory, Mailman School of Public Health and Department of Epidemiology, Columbia University, NY 10032, USA.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

15788125

Citation

Palacios, Gustavo, et al. "Severe Acute Respiratory Syndrome Coronavirus Persistence in Vero Cells." Chinese Medical Journal, vol. 118, no. 6, 2005, pp. 451-9.
Palacios G, Jabado O, Renwick N, et al. Severe acute respiratory syndrome coronavirus persistence in Vero cells. Chin Med J (Engl). 2005;118(6):451-9.
Palacios, G., Jabado, O., Renwick, N., Briese, T., & Lipkin, W. I. (2005). Severe acute respiratory syndrome coronavirus persistence in Vero cells. Chinese Medical Journal, 118(6), 451-9.
Palacios G, et al. Severe Acute Respiratory Syndrome Coronavirus Persistence in Vero Cells. Chin Med J (Engl). 2005 Mar 20;118(6):451-9. PubMed PMID: 15788125.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Severe acute respiratory syndrome coronavirus persistence in Vero cells. AU - Palacios,Gustavo, AU - Jabado,Omar, AU - Renwick,Neil, AU - Briese,Thomas, AU - Lipkin,W Ian, PY - 2005/3/25/pubmed PY - 2005/5/5/medline PY - 2005/3/25/entrez SP - 451 EP - 9 JF - Chinese medical journal JO - Chin Med J (Engl) VL - 118 IS - 6 N2 - 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 syndrome 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. SN - 0366-6999 UR - https://www.unboundmedicine.com/medline/citation/15788125/Severe_acute_respiratory_syndrome_coronavirus_persistence_in_Vero_cells_ DB - PRIME DP - Unbound Medicine ER -