Susceptibility of well-differentiated airway epithelial cell cultures from domestic and wildlife animals to SARS-CoV-2

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   ScreenIT

   Nov 11, 2020

   SciScore for 10.1101/2020.11.10.374587: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	not detected.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   For the detection of SARS-CoV-2, fixed animal AEC cultures were incubated with a rabbit polyclonal antibody against SARS-CoV Nucleocapsid protein (Rockland, 200-401-A50), which has previously been shown to cross-react with SARS-CoV-2 20.	antibody against SARS-CoV Nucleocapsid protein suggested: None
   To visualize the distribution of ACE2 in the AEC cultures, a rabbit polyclonal antibody against ACE2 (ab15348, Abcam) was used.	ACE2 suggested: (Abcam Cat# …

   SciScore for 10.1101/2020.11.10.374587: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	not detected.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   For the detection of SARS-CoV-2, fixed animal AEC cultures were incubated with a rabbit polyclonal antibody against SARS-CoV Nucleocapsid protein (Rockland, 200-401-A50), which has previously been shown to cross-react with SARS-CoV-2 20.	antibody against SARS-CoV Nucleocapsid protein suggested: None
   To visualize the distribution of ACE2 in the AEC cultures, a rabbit polyclonal antibody against ACE2 (ab15348, Abcam) was used.	ACE2 suggested: (Abcam Cat# ab15348, RRID:AB_301861)
   Alexa Fluor® 488-labeled donkey anti-Rabbit and -mouse IgG (H + L) (Jackson Immunoresearch) were used as secondary antibodies.	anti-Rabbit suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Conventional cell culture: Vero E6 cells (kindly provided by Doreen Muth, Marcel Müller, and Christian Drosten, Charité, Berlin, Germany) were cultured in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FBS), 1 mM sodium pyruvate, 1x GlutaMAX, 100 μg/ml streptomycin, 100 IU/ml penicillin, 1% (v/v) non-essential amino acids, and 15 mM HEPES (Gibco).	Vero E6 suggested: RRID:CVCL_XD71)
   The viral titer was determined by TCID50 assay on MDCK-II cells as described previously 41,42.	MDCK-II suggested: None
   Virus stocks were propagated in the human rectal tumor cell line HRT-18G (CRL11663, ATCC) for 96 hours in the infection medium, with the adjustment of using 0,25 μg/ml of trypsin.	HRT-18G suggested: ATCC Cat# CRL-11663, RRID:CVCL_2515)
   Software and Algorithms
   Sentences	Resources
   All images were processed using Fiji software packages 44.	Fiji suggested: (Fiji, RRID:SCR_002285)
   The protein alignment was performed using ClustalW in Geneious 11.1.5.	ClustalW suggested: (ClustalW, RRID:SCR_017277)
   Whole-Genome Sequencing using Oxford Nanopore (MinION): Sequencing was performed on viral RNA isolated from the SARS-CoV-2 stock and the 96 hpi apical washes of SARS-CoV-2-infected monkey and cat AEC cultures according to the ARTIC platform nCoV19 protocols 48,49.	Oxford Nanopore suggested: (Oxford Nanopore Technologies, RRID:SCR_003756)
   Sequencing libraries were generated using the Native Barcoding Expansion 96 kit (EXP-NBD196, Oxford Nanopore Technologies) and sequenced on a MinION R9.4.1 flowcell according to the manufacturer's instructions for a duration of 48 hours.	MinION suggested: (MinION, RRID:SCR_017985)
   Consensus sequences were aligned and further analyzed in Geneious 11.1.5.	Geneious suggested: (Geneious, RRID:SCR_010519)

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   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).

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   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.

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   Results from TrialIdentifier: No clinical trial numbers were referenced.

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   Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

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   Results from JetFighter: Please consider improving the rainbow (“jet”) colormap(s) used on page 18. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

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   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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   Version published to 10.1101/2020.11.10.374587 on bioRxiv

   Nov 10, 2020