Aerosol Exposure of Cynomolgus Macaques to SARS-CoV-2 Results in More Severe Pathology than Existing Models
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Abstract
The emergence of SARS-CoV-2 pandemic has highlighted the need for animal models that faithfully recapitulate the salient features of COVID-19 disease in humans; these models are necessary for the rapid down-selection, testing, and evaluation of medical countermeasures. Here we performed a direct comparison of two distinct routes of SARS-CoV-2 exposure, combined intratracheal/intranasal and small particle aerosol, in two nonhuman primate species: rhesus and cynomolgus macaques. While all four experimental groups displayed very few outward clinical signs, evidence of mild to moderate respiratory disease was present on radiographs and at the time of necropsy. Cynomolgus macaques exposed via the aerosol route also developed the most consistent fever responses and had the most severe respiratory disease and pathology. This study demonstrates that while all four models were suitable representations of mild COVID-like illness, aerosol exposure of cynomolgus macaques to SARS-CoV-2 produced the most severe disease, which may provide additional clinical endpoints for evaluating therapeutics and vaccines.
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SciScore for 10.1101/2021.04.27.441510: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics Field Sample Permit: Study approval: Research was conducted under an IACUC-approved protocol in compliance with the Animal Welfare Act, PHS Policy, and other Federal statutes and regulations relating to animals and experiments involving animals. Sex as a biological variable NHPs: The study used eight healthy cynomolgus macaques and eight healthy rhesus macaques with equal distribution of sex among the groups (n=8 males and n=8 females; n=2 of each per group). Randomization Virus exposure: Animals were randomized to exposure route balanced by sex and weight. Blinding not detected. Power Analysis not detected. Cell Line Authentication Contamination: The production stock underwent additional testing to … SciScore for 10.1101/2021.04.27.441510: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics Field Sample Permit: Study approval: Research was conducted under an IACUC-approved protocol in compliance with the Animal Welfare Act, PHS Policy, and other Federal statutes and regulations relating to animals and experiments involving animals. Sex as a biological variable NHPs: The study used eight healthy cynomolgus macaques and eight healthy rhesus macaques with equal distribution of sex among the groups (n=8 males and n=8 females; n=2 of each per group). Randomization Virus exposure: Animals were randomized to exposure route balanced by sex and weight. Blinding not detected. Power Analysis not detected. Cell Line Authentication Contamination: The production stock underwent additional testing to evaluate sterility, mycoplasma and endotoxin levels, as well as a number of real-time reverse transcriptase polymerase chain reaction (RT-PCR) assays for exclusivity and inclusivity, to include two specific for SARS-CoV-2 virus. Table 2: Resources
Antibodies Sentences Resources Then the sections were incubated with primary antibodies: rabbit anti-SARS-CoV Spike (1:200, 40150-T62-COV2, Sino Biological, Chesterbrook, PA, USA) anti-SARS-CoVsuggested: (Rockland Cat# 200-401-A51, RRID:AB_828457), rabbit anti-CD3 (1:200, A045229-2, Dako Agilent Pathology Solutions, Carpinteria, CA, USA), rabbit anti-MPO (1:200, A039829-2, Dako Agilent Pathology Solutions, Carpinteria, CA, USA), rabbit anti-CD68 (1:200, ab125047, Abcam, Cambridge, MA, USA), mouse anti-CD68 (1:100, M081401-2, Dako Agilent Pathology Solutions) anti-MPOsuggested: Noneanti-CD68suggested: (Abcam Cat# ab125047, RRID:AB_10971844), Thermo Fisher Scientific) and goat anti-mouse Alexa Fluor 568 (red, 1:500, Thermo Fisher Scientific) antibodies, for 1 hour at room temperature. anti-mousesuggested: NoneExperimental Models: Cell Lines Sentences Resources The CDC isolate had been passaged three times in CCL-81 cells prior to receipt at the USAMRIID lab. CCL-81suggested: NonePlaque assay: A neutral red plaque assay using ATCC Vero 76 cells was performed on the virus stock, AGIs from the AE-exposed animals, and nasopharyngeal and oropharyngeal swab lysates as previously described (13). Vero 76suggested: IZSLER Cat# BS CL 101, RRID:CVCL_0603)Software and Algorithms Sentences Resources Images were captured on a Zeiss LSM 880 confocal system (Zeiss, Oberkochen, Germany) and processed using ImageJ software (National Institutes of Health, Bethesda, ImageJsuggested: (ImageJ, RRID:SCR_003070)Statistics: All analyses were performed using GraphPad Prism 9. GraphPad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)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 page 24. 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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