Recurrent emergence of SARS-CoV-2 spike deletion H69/V70 and its role in the Alpha variant B.1.1.7
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SciScore for 10.1101/2020.12.14.422555: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IRB: Controls with COVID-19 were enrolled to the NIHR BioResource Centre Cambridge under ethics review board (17/EE/0025). Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication Contamination: All cells are regularly tested and are mycoplasma free. Table 2: Resources
Antibodies Sentences Resources Membranes were blocked for 1 hour in 5% non-fat milk in PBS + 0.1% Tween-20 (PBST) at room temperature with agitation, incubated in primary antibody (anti-SARS-CoV-2 Spike, which detects the S2 subunit of SARS-CoV-2 S (Invitrogen, PA1-41165)) diluted in 5% non-fat milk in PBST for 2 hours at … SciScore for 10.1101/2020.12.14.422555: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IRB: Controls with COVID-19 were enrolled to the NIHR BioResource Centre Cambridge under ethics review board (17/EE/0025). Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Cell Line Authentication Contamination: All cells are regularly tested and are mycoplasma free. Table 2: Resources
Antibodies Sentences Resources Membranes were blocked for 1 hour in 5% non-fat milk in PBS + 0.1% Tween-20 (PBST) at room temperature with agitation, incubated in primary antibody (anti-SARS-CoV-2 Spike, which detects the S2 subunit of SARS-CoV-2 S (Invitrogen, PA1-41165)) diluted in 5% non-fat milk in PBST for 2 hours at 4°C with agitation, washed four times in PBST for 5 minutes at room temperature with agitation and incubated in secondary antibody (anti-rabbit HRP conjugate, Invitrogen 31462), anti-bactin HRP (sc-47778) diluted in 5% non-fat milk in PBST for 1 hour with agitation at room temperature. anti-SARS-CoV-2suggested: NonePA1-41165suggested: (Thermo Fisher Scientific Cat# PA1-41165, RRID:AB_1087210)anti-rabbitsuggested: Noneanti-bactin HRPsuggested: Nonesc-47778suggested: NoneExperimental Models: Cell Lines Sentences Resources Briefly, Vero cells and 293T cells were seeded at 80% confluency in a 24 multiwell plate. Verosuggested: None293T cells were then detached 5 hours post transfection, mixed together with the green-labelled Vero cells, and plated in a 12 multiwell plate. 293Tsuggested: NoneHEK 293T CRL-3216 HEK 293Tsuggested: ATCC Cat# CRL-3216, RRID:CVCL_0063), Vero CCL-81 were purchased from ATCC and maintained in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal calf serum (FCS), 100 U/ml penicillin, and 100μg/ml streptomycin. Vero CCL-81suggested: NoneSoftware and Algorithms Sentences Resources All sequences were realigned to the SARS-CoV-2 reference strain MN908947.3, using MAFFT v7.475 with automatic flavour selection and the --keeplength --addfragments options36. MAFFTsuggested: (MAFFT, RRID:SCR_011811)Maximum likelihood phylogenetic trees were produced using the above curated dataset using IQ-TREE v2.1.237. IQ-TREEsuggested: (IQ-TREE, RRID:SCR_017254)All trees were visualised with Figtree v. Figtreesuggested: (FigTree, RRID:SCR_008515)Alignment visualisation was done using BioEdit42. BioEdit42suggested: NoneFigures prepared with PyMOL using PDBs 7C2L, 6M0J46, 6ZGE28 and 6ZGG47. PyMOLsuggested: (PyMOL, RRID:SCR_000305)Graphs were generated using Prism 8 software. Prismsuggested: (PRISM, RRID:SCR_005375)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: We detected the following sentences addressing limitations in the study:Given the emergence of multiple clusters of variants carrying RBD mutations and the ΔH69/V70 deletion, limitation of transmission takes on a renewed urgency. Continued emphasis on testing/tracing, social distancing and mask wearing are essential, with investment in other novel methods to limit transmission34. In concert, comprehensive vaccination efforts in the UK and globally should be accelerated in order to further limit transmission and acquisition of further mutations. Finally, detection of the deletion and other key mutations by rapid diagnostics should be a research priority as such tests could be used as a proxy for antibody escape mutations to inform surveillance at global scale. Limitations: The laboratory virology aspects of this study were conducted with pseudoviruses rather than replication competent viruses.
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: 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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SciScore for 10.1101/2020.12.14.422555: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
NIH rigor criteria are not applicable to paper type.Table 2: Resources
Experimental Models: Cell Lines Sentences Resources 1.4.4 Pseudotype virus preparation Viral vectors were prepared by transfection of 293T cells by using Fugene HD transfection reagent ( 293Tsuggested: NCBI_Iran Cat# C498, RRID:CVCL_0063)Software and Algorithms Sentences Resources Duplicate and low-quality sequences Maximum likelihood phylogenetic trees were produced using the above curated dataset using IQ-TREE v2.1.2 ModelFinder bootstrap replicates20. IQ-TREE…SciScore for 10.1101/2020.12.14.422555: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
NIH rigor criteria are not applicable to paper type.Table 2: Resources
Experimental Models: Cell Lines Sentences Resources 1.4.4 Pseudotype virus preparation Viral vectors were prepared by transfection of 293T cells by using Fugene HD transfection reagent ( 293Tsuggested: NCBI_Iran Cat# C498, RRID:CVCL_0063)Software and Algorithms Sentences Resources Duplicate and low-quality sequences Maximum likelihood phylogenetic trees were produced using the above curated dataset using IQ-TREE v2.1.2 ModelFinder bootstrap replicates20. IQ-TREEsuggested: (IQ-TREE, RRID:SCR_017254)Evolutionary model selection for trees were inferred using and trees were estimated using the GTR+F+I model with 1000 ultrafast All trees were visualised with Figtree v. Figtreesuggested: (FigTree, RRID:SCR_008515)Figures prepared with PyMOL (Schrödinger) using PDBs 7C2L, 6ZGE28 and 6ZGG . PyMOLsuggested: (PyMOL, RRID:SCR_000305)All sequences carrying the double-deletion were downloaded from the GISAID database and aligned to the Wuhan-Hu-1 reference strain using MAFFT. MAFFTsuggested: (MAFFT, RRID:SCR_011811)LK- B3A ope /202 0|E /V |EP -19 /So 9|20 606sk_ A f r i c and /MI hC oV hC oV- 19/ 1-08 |Eur 14| 00 IC a B5B D5F PI_ ISL _64 I_I SL uth _A sk_202 rope 4|20 20-1 0-1 1-1 V-1 9/E ngl and /AL DPsk_ |2 _6 fri /202 0|E _65 10-2 9|Eu 3|A fric ngl 2|2 ca DPsk_suggested: NoneResults 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: We detected the following sentences addressing limitations in the study:
Given the emergence of multiple clusters of variants carrying RBD mutations and the ઢH69/ઢV70 deletion, limitation of transmission takes on a renewed urgency. Concerted global vaccination efforts with wide coverage should be accelerated. Continued emphasis on testing/tracing, social distancing and mask wearing are essential, with investment in other novel methods to limit transmission15. Detection of the deletion by rapid diagnostics should be a research priority as such tests could be used as a proxy for antibody escape mutations to inform surveillance at global scale.
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: We did not find any issues relating to colormaps.
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