Integrated Single-Cell Atlases Reveal an Oral SARS-CoV-2 Infection and Transmission Axis
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Abstract
Despite signs of infection, the involvement of the oral cavity in COVID-19 is poorly understood. To address this, single-cell RNA sequencing data-sets were integrated from human minor salivary glands and gingiva to identify 11 epithelial, 7 mesenchymal, and 15 immune cell clusters. Analysis of SARS-CoV-2 viral entry factor expression showed enrichment in epithelia including the ducts and acini of the salivary glands and the suprabasal cells of the mucosae. COVID-19 autopsy tissues confirmed in vivo SARS-CoV-2 infection in the salivary glands and mucosa. Saliva from SARS-CoV-2-infected individuals harbored epithelial cells exhibiting ACE2 expression and SARS-CoV-2 RNA. Matched nasopharyngeal and saliva samples found distinct viral shedding dynamics and viral burden in saliva correlated with COVID-19 symptoms including taste loss. Upon recovery, this cohort exhibited salivary antibodies against SARS-CoV-2 proteins. Collectively, the oral cavity represents a robust site for COVID-19 infection and implicates saliva in viral transmission.
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SciScore for 10.1101/2020.10.26.20219089: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement Consent: HUMAN MINOR SALIVARY GLAND SINGLE CELL RNA SEQUENCING: Tissue Dissociation: Minor SGs were collected from healthy volunteers who provided informed consent on NIH protocols 15-D-0051 or 94-D-0094. Randomization For integration across datasets, each individual dataset was first randomly down-sampled to having <=500 cells per cell type, then pooled raw expression values were re-processed following standard SCANPY procedure while using Harmony for correcting batch effects across datasets and samples (70). Blinding not detected. Power Analysis not detected. Sex as a biological variable HUMAN GINGIVAL SINGLE CELL RNA SEQUENCING: Clinical Protocol: Five subjects … SciScore for 10.1101/2020.10.26.20219089: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement Consent: HUMAN MINOR SALIVARY GLAND SINGLE CELL RNA SEQUENCING: Tissue Dissociation: Minor SGs were collected from healthy volunteers who provided informed consent on NIH protocols 15-D-0051 or 94-D-0094. Randomization For integration across datasets, each individual dataset was first randomly down-sampled to having <=500 cells per cell type, then pooled raw expression values were re-processed following standard SCANPY procedure while using Harmony for correcting batch effects across datasets and samples (70). Blinding not detected. Power Analysis not detected. Sex as a biological variable HUMAN GINGIVAL SINGLE CELL RNA SEQUENCING: Clinical Protocol: Five subjects were enrolled in this study conducted at the University of North Carolina Adams School of Dentistry (Chapel Hill, North Carolina), and 5 total healthy subjects (2 females and 3 males; aged 20 to 30 years) completed the experimental gingivitis protocol in October 2019. Table 2: Resources
Software and Algorithms Sentences Resources Illustration of the results were generated using python code around SCANPY. IHC AND ISH FOR HUMAN ORAL TISSUES AND SALIVA: CLINICAL STUDIES: PROCUREMENT AND TESTING OF AUTOPSY TISSUE SPECIMENS FOR SARS-COV-2: SALIVARY pythonsuggested: (IPython, RRID:SCR_001658)Results from OddPub: Thank you for sharing your code and data.
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:These percentages are slightly lower than reported for cells in the respiratory tract but may reflect limitations in the detection of low-expression genes by scRNAseq. Co-expression of these markers was more frequent using ISH and emphasized that SARS-CoV-2 is predicted to have tropism to the acini and ducts. While there has been a historical focus on major SG viral infections, broad expression of viral entry factors and significant infection of minor SG epithelia was confirmed. Minor SGs are widely distributed among the tongue, palate, and mucosae (60), sites now demonstrated to be hotspots for SARS-CoV-2 infection. The novel finding of SG and tongue infection helps explain acute onset and post-recovery COVID-19 symptoms, such as dry mouth and taste alterations (14); however, infection and productive replication in even a majority of these glands may produce no symptoms, possibly explaining the phenomenon of “silent spreaders” in COVID-19. One limitation of our scRNAseq approach was the cellular representation of the mucosal atlas which enriched for immune cells over suprabasal squamous epithelial cells (Figure 2; Supp. Figure 1). When compared to the SG, these results suggested that SARS-CoV-2 would have relatively little tropism for the mucosa based on the low expression and abundance of cells expressing ACE2 and TMPRRS2. However, integrated pan-oral mouse single cell data revealed a more robust description of the oral cavity due to increased numbers of suprabasal cells fr...
Results from TrialIdentifier: We found the following clinical trial numbers in your paper:
Identifier Status Title NCT04105569 Completed Atlas of Experimental Gingivitis in Humans NCT04348240 Recruiting Transmissibility and Viral Load of SARS-CoV-2 in Oral Secret… NCT02327884 Recruiting Characterization of Diseases With Salivary Gland Involvement Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
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.
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Ophir Klein, Irit Miller Zmora
Review 2: "Integrated Single-Cell Atlases Reveal an Oral SARS-CoV-2 Infection and Transmission Axis"
This pre-print uses single-cell RNA sequencing to investigate oral SARS-CoV-2 tropism and finds the mucosa represents a robust site of infection. Reviewers deem the study's claims reliable.
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Hongxiang Liu, Zhonghou Wang, Wenxin Yu, Mohamed Ishan
Review 1: "Integrated Single-Cell Atlases Reveal an Oral SARS-CoV-2 Infection and Transmission Axis"
This pre-print uses single-cell RNA sequencing to investigate oral SARS-CoV-2 tropism and finds the mucosa represents a robust site of infection. Reviewers deem the study's claims reliable.
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Strength of evidence
Reviewers: Hongxiang Liu (University of Georgia), Zhonghou Wang, Wenxin Yu, Mohamed Ishan | 📗📗📗📗◻️
Ophir Klein, Irit Miller Zmora (UCSF) | 📘📘📘📘📘 -
