Cited by 142SARS-CoV-2 and COVID-19 ResearchAnimal Virus Infections StudiesCOVID-19 Clinical Research Studies

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SARS-CoV-2 growth, furin-cleavage-site adaptation and neutralization using serum from acutely infected hospitalized COVID-19 patients

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Klimstra, William B., Tilston-Lunel, Natasha L., Nambulli, Sham, Boslett, James, McMillen, Cynthia M., Gilliland, Theron, Dunn, Matthew D., Sun, Chengun, Wheeler, Sarah E., Wells, Alan, Hartman, Amy L., McElroy, Anita K., Reed, Douglas S., Rennick, Linda J., Duprex, W. Paul·2020·Journal of General Virology

DOI 10.1099/jgv.0.001481PMC PMC7879561

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Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), emerged at the end of 2019 and by mid-June 2020 the virus had spread to at least 215 countries, caused more than 8 000 000 confirmed infections and over 450 000 deaths, and overwhelmed healthcare systems worldwide. Like severe acute respiratory syndrome coronavirus (SARS-CoV), which emerged in 2002 and caused a similar disease, SARS-CoV-2 is a betacoronavirus. Both viruses use human angiotensin-converting enzyme 2 (hACE2) as a receptor to enter cells. However, the SARS-CoV-2 spike (S) glycoprotein has a novel insertion that generates a putative furin cleavage signal and this has been postulated to expand the host range. Two low-passage (P) strains of SARS-CoV-2 (Wash1 : P4 and Munich : P1) were cultured twice in Vero E6 cells and characterized virologically. Sanger and MinION sequencing demonstrated significant deletions in the furin cleavage signal of Wash1 : P6 and minor variants in the Munich : P3 strain. Cleavage of the S glycoprotein in SARS-CoV-2-infected Vero E6 cell lysates was inefficient even when an intact furin cleavage signal was present. Indirect immunofluorescence demonstrated that the S glycoprotein reached the cell surface. Since the S protein is a major antigenic target for the development of neutralizing antibodies, we investigated the development of neutralizing antibody titres in serial serum samples obtained from COVID-19 human patients. These were comparable regardless of the presence of an intact or deleted furin cleavage signal. These studies illustrate the need to characterize virus stocks meticulously prior to performing either in vitro or in vivo pathogenesis studies.

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Authors: Klimstra, W. B., Tilston-Lunel, N. L., Nambulli, S., Boslett, J., McMillen, C. M., Gilliland, T., Dunn, M. D., Sun, C., Wheeler, S. E., Wells, A., Hartman, A. L., McElroy, A. K., Reed, D. S., Rennick, L. J., & Duprex, W. P.
Affiliations: University of Pittsburgh (US)
Department of Microbiology and Molecular Genetics, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
Corresponding author: Klimstra, William B.
Duprex, W. Paul
ORCID: Klimstra, William B.
Tilston-Lunel, Natasha L.
Nambulli, Sham
Boslett, James
McMillen, Cynthia M.
Gilliland, Theron
Wheeler, Sarah E.
Wells, Alan
Hartman, Amy L.
McElroy, Anita K.
Reed, Douglas S.
Rennick, Linda J.
Duprex, W. Paul
Venue: Journal of General Virology
Publisher: Microbiology Society
Published: 2020-11-01
Volume: 101
Issue: 11
Pages: 1156-1169
ISSN: 0022-1317 (ISSN-L)
1465-2099
DOI: 10.1099/jgv.0.001481
PMC: PMC7879561
OpenAlex: W3081204841
Cited by: 137
References: 69
Citations by year: 2026: 3 · 2025: 5 · 2024: 6 · 2023: 20 · 2022: 29 · 2021: 61 · 2020: 13
FWCI: 4.03
Type: journal article
Language: en
Open access: hybrid
doi.org
Licence: cc-by
creativecommons.org/licenses/by/4.0/ (unspecified)
Locations: Journal of General Virology (publishedVersion, cc-by)
bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
PubMed Central (publishedVersion, cc-by)
PubMed (publishedVersion)
PubMed Central (submittedVersion, cc-by-nc-nd)
Funders: Dr. Patrick Gallagher, Chancellor of the University of Pittsburgh
Center for Vaccine Research, University of Pittsburgh
University of Pittsburgh Clinical and Translational Science Institute
DSF Charitable Foundation
National Institutes of Health (AI060525)
Topics: Health Sciences › Medicine › Infectious Diseases › SARS-CoV-2 and COVID-19 Research
Life Sciences › Agricultural and Biological Sciences › Animal Science and Zoology › Animal Virus Infections Studies
Health Sciences › Medicine › Infectious Diseases › COVID-19 Clinical Research Studies
Keywords: SARS-CoV-2, COVID-19 pandemic, furin cleavage site, spike protein, viral growth, neutralizing antibodies, host range, SARS-CoV, angiotensin-converting enzyme 2

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SARS-CoV-2 growth, furin-cleavage-site adaptation and neutralization using serum from acutely infected hospitalized COVID-19 patients

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