Mild SARS-CoV-2 infection in rhesus macaques is associated with viral control prior to antigen-specific T cell responses in tissues

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) primarily replicates in mucosal sites, and more information is needed about immune responses in infected tissues. Here, we used rhesus macaques to model protective primary immune responses in tissues during mild coronavirus disease 2019 (COVID-19). Viral RNA levels were highest on days 1 to 2 after infection and fell precipitously thereafter. ¹⁸ F-fluorodeoxyglucose ( ¹⁸ FDG)–avid lung abnormalities and interferon (IFN)–activated monocytes and macrophages in the bronchoalveolar lavage (BAL) were found on days 3 to 4 after infection. Virus-specific effector CD8 ⁺ and CD4 ⁺ T cells became detectable in the BAL and lung tissue on days 7 to 10 after viral RNA, radiologic evidence of lung inflammation, and IFN-activated myeloid cells had substantially declined. SARS-CoV-2–specific T cells were not detectable in the nasal turbinates, salivary glands, and tonsils on day 10 after infection. Thus, SARS-CoV-2 replication wanes in the lungs, as well as the nasal and oral mucosa, of rhesus macaques before antigen-specific effector T cells arrive at those sites, suggesting that innate immunity efficiently restricts viral replication during mild COVID-19.

SciScore for 10.1101/2022.01.06.475282: (What is this?)

Please note, not all rigor criteria are appropriate for all manuscripts.

Table 1: Rigor

Ethics	IACUC: Study design: All animal experiments were approved by Animal Care and Use Committee (ACUC) and all methods were approved on animal safety protocol LPD-25E at the National Institute of Health.
Sex as a biological variable	Animals and infection: Six, male rhesus macaques aged 2.5 to 6 years, weighing 3-10 kg were infected with SARS-CoV-2/USA/WA-1 (Table 1).
Randomization	not detected.
Blinding	not detected.
Power Analysis	not detected.

Table 2: Resources

Antibodies
Sentences	Resources
After blocking, slides were incubated with primary antibodies against CD62P (clone EPR22850-190, Abcam, USA) and fibrin (clone 59D8, Millipore Sigma, USA) at a 1:500 and 1:200 concentration, respectively.	CD62P suggested:…

SciScore for 10.1101/2022.01.06.475282: (What is this?)

Please note, not all rigor criteria are appropriate for all manuscripts.

Table 1: Rigor

Ethics	IACUC: Study design: All animal experiments were approved by Animal Care and Use Committee (ACUC) and all methods were approved on animal safety protocol LPD-25E at the National Institute of Health.
Sex as a biological variable	Animals and infection: Six, male rhesus macaques aged 2.5 to 6 years, weighing 3-10 kg were infected with SARS-CoV-2/USA/WA-1 (Table 1).
Randomization	not detected.
Blinding	not detected.
Power Analysis	not detected.

Table 2: Resources

Antibodies
Sentences	Resources
After blocking, slides were incubated with primary antibodies against CD62P (clone EPR22850-190, Abcam, USA) and fibrin (clone 59D8, Millipore Sigma, USA) at a 1:500 and 1:200 concentration, respectively.	CD62P suggested: None fibrin suggested: (Millipore Cat# MABS2155, RRID:AB_2893306)
Software and Algorithms
Sentences	Resources
After processing, the spleen and lung were cleared of red blood cells by resuspending cell pellet in 2mL of ACK Lysing Buffer (Quality Biologicals Cat#118-156-101) for 2 minutes at room temperature, then stopping the reaction with 10-20mL of PBS + 1%FBS.	Quality Biologicals suggested: None
Differentially expressed genes between timepoints of a particular cluster were identified by running FindMarkers function with MAST and comparing one timepoint to all other timepoints or one timepoint to another in a pairwise manner.	MAST suggested: (MAST, RRID:SCR_016340)
Genes with a log fold change ≥ 0.5, percent of cells expressing the marker ≥ 0.25 and adjusted p value ≤ 0.01 were considered significant and these genes were hierarchically clustered and displayed as a heatmap using the ComplexHeatmap function in R.	ComplexHeatmap suggested: (ComplexHeatmap, RRID:SCR_017270)
Gene ontology enrichment analysis of genes upregulated at a particular timepoint was performed using clusterProfiler to identify biological processes (adjusted p value ≤ 0.05).	clusterProfiler suggested: (clusterProfiler, RRID:SCR_016884)
The AverageExpression function was used to calculate average gene expression of IFN and IFN stimulated genes across all cells over time and was visualized using pheatmap.	pheatmap suggested: (pheatmap, RRID:SCR_016418)

Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

Results from TrialIdentifier: No clinical trial numbers were referenced.

Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

Results from JetFighter: Please consider improving the rainbow (“jet”) colormap(s) used on pages 56 and 52. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

Results from rtransparent:

Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
No protocol registration statement was detected.

Results from scite Reference Check: We found no unreliable references.

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Mild SARS-CoV-2 infection in rhesus macaques is associated with viral control prior to antigen-specific T cell responses in tissues

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