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Comparative Study
. 2016 Mar 15;113(11):3048-53.
doi: 10.1073/pnas.1517719113. Epub 2016 Mar 14.

SARS-like WIV1-CoV Poised for Human Emergence

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Free PMC article
Comparative Study

SARS-like WIV1-CoV Poised for Human Emergence

Vineet D Menachery et al. Proc Natl Acad Sci U S A. .
Free PMC article

Abstract

Outbreaks from zoonotic sources represent a threat to both human disease as well as the global economy. Despite a wealth of metagenomics studies, methods to leverage these datasets to identify future threats are underdeveloped. In this study, we describe an approach that combines existing metagenomics data with reverse genetics to engineer reagents to evaluate emergence and pathogenic potential of circulating zoonotic viruses. Focusing on the severe acute respiratory syndrome (SARS)-like viruses, the results indicate that the WIV1-coronavirus (CoV) cluster has the ability to directly infect and may undergo limited transmission in human populations. However, in vivo attenuation suggests additional adaptation is required for epidemic disease. Importantly, available SARS monoclonal antibodies offered success in limiting viral infection absent from available vaccine approaches. Together, the data highlight the utility of a platform to identify and prioritize prepandemic strains harbored in animal reservoirs and document the threat posed by WIV1-CoV for emergence in human populations.

Keywords: CoV; SARS; Spike; WIV1; emergence.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Full-length and chimeric WIV1 infectious clones produce viruses that replicate in primary human airway epithelial cell cultures. (A) Spike amino acid residues that interact directly with human ACE2 from SARS-CoV, SARS-MA15, and WIV1-CoV spike proteins. Residue changes are highlighted by color. (B) Interaction between S1 domain of SARS-Urbani spike (black) and WIV1 spike (blue) with human ACE2 (gray). Contact residues highlighted with consensus amino acids (red) and differences (circled) between SARS and WIV1 spike proteins; human ACE2 contact residues are also highlighted (orange). (C) Viral replication of WIV1-CoV (blue), WIV1-MA15 (blue hatched), and SARS-CoV Urbani (black) following infection of Vero cells at a multiplicity of infection (MOI) of 0.01. (D) Well-differentiated air–liquid interface primary human airway epithelial cell cultures were infected with SARS-CoV Urbani (black), SARS-CoV MA15 (black hatched), WIV1-MA15 (blue-white hatched), and WIV-CoV (blue) at (E) MOI of 0.01 in cells from the same donor at an MOI of 0.01. Samples were collected at individual time points with biological replicates (n = 3) for all experiments for both C and D.
Fig. 2.
Fig. 2.
Viruses using WIV1 spike attenuated relative to SARS spike in vivo. (A and B) Ten-week-old BALB/c mice were infected with 104 pfu of either SARS-CoV MA15 (black) or WIV1-MA15 (blue hatched) via the i.n. route and examined over a 7-d time course. (A) Weight loss (n = 17 for WIV1-MA15, n = 9 for SARS-CoV MA15) and (B) lung titer (n = 3 for MA15, n = 4 for WIV1-MA15. (C and D) Ten-week-old BALB/c mice were infected with 1 × 105 pfu of either SARS-CoV Urbani (black), WIV1-CoV (blue), or SARS-CoV MA15 (gray) and examined over a 4-d time course. (C) Weight loss (n = 6 for WIV1-CoV, n = 6 for SARS-CoV Urbani) and (D) lung titer (n = 3 for WIV1-CoV, n = 3 for SARS-CoV Urbani) were examined. For each bar graph, center value is representative of group mean and error bars are defined by SEM. P values based on two-tailed Student’s t test of individual time points are marked as indicated: ***P < 0.001.
Fig. 3.
Fig. 3.
WIV1-Cov still attenuated despite human ACE2 expression in vivo. (A) Ten- to twenty-week-old HFH4 ACE2-expressing mice were infected with 105 pfu of SARS-CoV Urbani (black) or WIV1-CoV (blue) and examined over a 7-d time course for (A) survival and (B) day-2 lung titer (n = 3 for WIV1-CoV, n = 3 for SARS-CoV Urbani). (C and D) Upon reaching thresholds for humane sacrifice (>20% weight loss) or 7 d postinfection (DPI), endpoint titers were determined in the (C) lung and (D) brain following infection. P values based on two-tailed Student’s t test of individual time points are marked as indicated: *P < 0.05.
Fig. 4.
Fig. 4.
SARS-CoV monoclonal antibodies have robust neutralization against WIV1 spike-mediated infection. Neutralization efficacy was evaluated using percent neutralization assays against SARS-CoV Urbani (black) or WIV1-MA15 (blue) with a panel of monoclonal antibodies: (A) fm6, (B) 230.15, (C) 227.15, and (D) 109.8, all originally generated against epidemic SARS-CoV. Each data point is representative of two or more independent neutralization wells. (E and F) Twenty- to twenty-four-week-old HFH4 ACE2-expressing mice were injected with 200 μg of anti-SARS human antibody 227.15 (hatched line) or mock (solid line) 1 d before infection with 1 × 10^5 pfu of SARS-CoV Urbani (black) or WIV1-CoV (blue) and examined over a 7-d time course for (E) survival (n = 3 for both antibody-treated groups and mock PBS control WIV1-CoV, n = 2 for mock-treated SARS-CoV Urbani), (F) day-2 lung titer (n = 3 for all groups). ND signifies no titers detected. For each bar graph, center value is representative of group mean and error bars are defined by SEM.
Fig. 5.
Fig. 5.
Double-inactivated whole SARS-CoV vaccine fails to protect aged animals from chimeric WIV1-CoV infection. Twelve-month-old mice were vaccinated and boosted with DIV (dotted line) or PBS (solid line) and infected 21 d postboost with 104 pfu of WIV1-MA15 via the i.n. route. (A) Weight loss following WIV1-MA15 challenge and (B) viral replication in the lung 4 DPI. (C) Neutralization of WIV1-MA15 (blue) with serum from aged, DIV-vaccinated mice. (DH) Histopathology lung sections stained for H&E from DIV- and mock-vaccinated mice. (D) Eosinophil score (scale 0–4) following DIV or mock vaccination 4 DPI. (E and F) Representative H&E lung sections for (E) mock- and (F) DIV-vaccinated mice infected with WIV-MA15. Red arrows indicate individual eosinophil locations. P values based on two-tailed Student’s t test of individual time points are marked as indicated: **P < 0.01.

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