Fig 1.
Detection and isolation of SARS-CoV-2 in swabs.
(A) Representative virus isolations confirmed by immunofluorescence using anti-SARS2 spike Ab with phalloidin as background. (B) Isolation results from swabs. Number in each square represents the # of plaques obtained from isolation. (C-F) vRNA in swabs measured by q-RT-PCR. AGM infected by aerosol (closed symbols/solid lines; n = 4) or multi-route mucosal (open symbols/dashed lines; n = 2). In B-F, Red symbols highlight representative IFA images shown in (A).
Fig 2.
PET/CT imaging of SARS-CoV-2-infected AGM.
(A) and (B) Region-of-interest analysis on PET images of animals infected with SARS-CoV-2. (A) Measurement of total lung inflammation via FDG uptake over time. (B) Measurement of average lymph node inflammation over time. Aerosol (closed symbols/solid lines; n = 3); multi-route mucosal (open symbols/dashed lines; n = 2). Animal A2 is not included in (A) and (B) because only CT scans were performed. (C) For AGM A4 (aerosol), only CT scan was obtained pre-infection; PET/CT at was obtained at 4 dpi and 11 dpi. (D) For AGM M1 (multi-route mucosal), only CT scan was obtained pre-infection; PET/CT at was obtained at 4 dpi and 11 dpi.. Pulmonary infection (yellow arrows); thoracic lymph nodes (green arrows). Cyan arrow highlights new focal area of disease visible on 11 dpi. PET color scale is from 0–15 SUV.
Fig 3.
Seroconversion and neutralization in SARS-CoV-2 infected AGMs.
Serial plasma samples were assayed for virus specific antibodies or neutralization capacity. (A) virus-specific IgG and IgM were measured using a SARS-CoV-2 spike receptor binding domain-based ELISA. (B) neutralization titers were determined by a plaque-reduction neutralization 80% assay (PRNT80). The horizontal dotted line represents the limit of detection of each assay.
Fig 4.
Blood cell populations in SARS-CoV-2 infected AGMs.
(A) Classical monocytes (CD14+CD16-), (B) Ki-16+ classical monocytes, (C) CD4+ T cells, (D) CD4+Ki-67+ cells, (E) CD8+ T cells, (F) CD8+Ki-67+ cells, (G) B cells with high CD20 expression, (H) B cells with low CD20 expression (plasmablasts), (I) Grouped expression of Ki-67 on CD20hi and CD20lo (plasmablast) populations. 2-way ANOVA with multiple comparisons was used to determine statistical significance compared to 0 dpi and is indicated by asterisks above each time point. N.s. in the lower right corner indicates no significant differences.
Fig 5.
Cytokinemia during SARS-CoV-2 infection.
Cytokines were measured in longitudinal plasma samples using the Cytokine & Chemokine 30-Plex NHP ProcartaPlex Panel from Invitrogen. (A) MCP-1; (B) IL-1RA; (C) IP-10; (D) ITAC/CXCL11/IP-9; (E) BLC/CXCL13; (F) Eotaxin; (G) SDF-1. Parameters not shown were below the limit of detection across all animals and time points. Aerosol (closed symbols/solid lines; n = 4); multi-route mucosal (open symbols/dashed lines; n = 2). A 2-way ANOVA with multiple comparisons was used to determine statistical significance compared to 0 dpi and is indicated by asterisks above each time point. N.s. in the lower right corner indicates no significant differences.
Fig 6.
Viral RNA within tissues of SARS-CoV-2 infected AGM.
Animals underwent necropsy at 28 dpi (35 dpi for A1 and A2) and the indicated tissues were extracted and tested for viral RNA by q-RT-PCR. Heat map shows the log-transformed vRNA copies per 100 mg of tissue. X indicates the sample was not available or not tested.
Fig 7.
Histopathology detected at necropsy.
(A) and (B) Lung (20X) reveals pulmonary foci of mild infiltration and interstitial expansion by lymphocytic and mixed inflammatory cells. (C) Peyer’s patch (20X) shows multiple, syncytialized cells (white arrows). Inset of Peyer’s patch illustrates smudgy syncytia. Histopathology interpreted by a board-certified veterinary pathologist.