A comparison of health care worker-collected foam and polyester nasal swabs in convalescent COVID-19 patients

Brian Hart; Yuan-Po Tu; Rachel Jennings; Prateek Verma; Leah R. Padgett; Douglas Rains; Deneen Vojta; Ethan M. Berke

doi:10.1371/journal.pone.0241100

Peer Review History

Original SubmissionMay 12, 2020
2 Sep 2020 Decision Letter - Eric HY Lau, Editor PONE-D-20-13951 A comparison of health care worker-collected foam and polyester nasal swabs in convalescent COVID-19 patients PLOS ONE Dear Dr. Hart, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. The Authors are expected to address all the criticisms by all Reviewers. In particular, please discuss if different volume of VTM or saline used, or normalization of Ct values would affect the results (Reviewer #1) and provide basic clinical information of the two cohorts (Reviewer #2). In additional to the above comments, please address, Please present the number of samples collected by day of visit group Tables 1-3 have pooled samples collected at different days after diagnosis. However, viral shedding decreased over time and hence test sensitivity is unlikely constant over sampling dates. Please further assess the sensitivity by the time of sampling. The authors have assessed sensitivity of different sampling methods. Could the authors explain why specificity was not assessed? Please submit your revised manuscript by Oct 17 2020 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. Please include the following items when submitting your revised manuscript: A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'. An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'. If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter. If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols We look forward to receiving your revised manuscript. Kind regards, Eric HY Lau, Ph.D. Academic Editor PLOS ONE Additional Editor Comments: The Authors are expected to address all the criticisms by all Reviewers. In particular, please discuss if different volume of VTM or saline used, or normalization of Ct values would affect the results (Reviewer #1) and provide basic clinical information of the two cohorts (Reviewer #2). In additional to the above comments, please address, 1. Please present the number of samples collected by day of visit group 2. Tables 1-3 have pooled samples collected at different days after diagnosis. However, viral shedding decreased over time and hence test sensitivity is unlikely constant over sampling dates. Please further assess the sensitivity by the time of sampling. 3. The authors have assessed sensitivity of different sampling methods. Could the authors explain why specificity was not assessed? Journal Requirements: When submitting your revision, we need you to address these additional requirements. 1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf 2. Thank you for stating the following in the Financial Disclosure section: "Laboratory testing was conducted with financial support from Thermo Fisher Scientific." We note that one or more of the authors are employed by a commercial company: UnitedHealth Group and Quantigen Biosciences. 2.1. 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The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #2: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: Yes ****** 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes Reviewer #2: Yes ****** 4. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes Reviewer #2: Yes ****** 5. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: A timely trial of polyester vs foam nasal swabs stored and/or tested using VTM, saline, or dry tubes with buffered saline. Comment only: It is unfortunate that you could not have treated the samples more similarly. I’m not sure why you chose to use different amount of VTM and saline, for instance, at the lab. Correcting for/normalizing the Ct values is adequate but not as convincing. Did you try to correct for the first/second collection of swabs in the analysis? It is not necessary, just a thought. Swabs do not detect virus, but rather “collect” them. Consider changing the wording. Discussion: line 186, please add, “in times of shortages” or something to that effect. Reviewer #2: In the context of the COVID-19 pandemic, the simple, convenient and effective detection methods are worth promoting. The polyester swabs stored in VTM or saline may be a viable sample collection method for COVID-19. But in the RT-PCR part, the author should show more details. In addition, the basic clinical information of the two cohorts should be presented, such as sex, BMI, age, symptoms and so on. ****** 6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review?** For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No Reviewer #2: No [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". 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Revision 1
16 Sep 2020 Author Response The authors would like to thank the reviewers and editor for their careful review and consideration of our manuscript titled “A comparison of health care worker-collected foam and polyester nasal swabs in convalescent COVID-19 patients”. Below we provide a point by point response to the reviewer comments below: • Please discuss if different volume of VTM or saline used, or normalization of Ct values would affect the results (Reviewer #1). Yes, using different sample volumes and applying mathematical corrections is not ideal and could affect the results. However, considering the urgent need for these studies, the standard 1-3 mL volumes used for clinical sample collection, and the valid mathematical approach for performing fold change calculations on qPCR, we felt normalization of the data was the best approach. For clarification, we have added text to the methods section and to the discussion. Methods: “Because of the volumetric differences among the three types of collection and/or elution media, mathematical adjustments to some Ct values were made in order to account for the differences in dilution of the various swab samples. By scaling (i.e., normalizing) Cts for both the 2 mL saline samples and the 3 mL VTM samples to match the 1 mL sample volume (VTM or 1 mL PBS dry swab elution), the sensitivity comparisons among the three sample collection volumes are more legitimate than using unnormalized data. Values were normalized as follows: to account for the 2-fold dilution difference between saline (2 mL) and dry polyester (1 mL PBS) or foam (1 mL VTM), 1 Ct was subtracted from all qPCR results for the former; similarly, to account for the 3-fold sample volume difference between the 3 mL VTM polyester swab samples and dry polyester (1 mL PBS) or foam (1 mL VTM), 1.585 Ct was subtracted from the former. Note that the specific Ct adjustments follow the formula 2-dCt=F, where dCt is the necessary Ct adjustment and F is the fold increase in volume. This formula assumes the assays perform at approximately 100% PCR efficiency during the geometric phase of amplification.” Discussion: “Although final Ct values were adjusted for varying amounts of transport media, imprecision in these adjustments could confound data interpretation; specifically, comparisons between the mathematically corrected samples (2 mL and 3 mL) and the non-corrected samples (1 mL) present the greatest risk for error.” • Provide basic clinical information of the two cohorts (Reviewer #2). Thank you for the suggestion. We have added in a new table one with basic clinical/demographic information available on the two cohorts. • Please present the number of samples collected by day of visit group These values can now be found in the expanded version of Tables 2 (previously Table 1). • Tables 1-3 have pooled samples collected at different days after diagnosis. However, viral shedding decreased over time and hence test sensitivity is unlikely constant over sampling dates. Please further assess the sensitivity by the time of sampling. Table 2 (previously called Table 1) has been expanded to breakout the results by the three different visits. Note that Tables 3 and 4 contain results which were collected at a single visit, so no similar table breakout was necessary/possible. • The authors have assessed sensitivity of different sampling methods. Could the authors explain why specificity was not assessed? When estimating sensitivity we assume that all positives were true positives. The primary advantage of this approach is that it allows for the estimation of sensitivity without assuming a single method is a gold standard “truth”. The disadvantage of our approach is that assuming all positives are true positives implies that all methods have a specificity of 100%. One could alternatively treat a single method (e.g. foam VTM) as the truth and compare the other methods when estimating sensitivity. This approach also involves strong assumptions and, while it allows for the estimation of both sensitivity and specificity of the non-reference methods, it implies 100% sensitivity and specificity of the reference method. Based on these tradeoffs, we chose to treat any positive as a true positive even though it precludes accurate specificity estimation. In addition to the edits made in response to reviewer comments we have made some additional changes to the description of the procedures. These changes were made so that the manuscript accurately describes how the study was carried out in practice. The sample collection procedures are now more accurately and clearly described in the manuscript. We have also changed the method used to calculate the 95% confidence intervals from the Normal approximation to the Wilson Score method, which has been shown to have better coverage. This change was motivated by the need for more stable confidence interval estimates given the low sample sizes, especially when broken out by visit. Attachments Attachment Submitted filename: Response to Reviewers_9_15.docx https://doi.org/10.1371/journal.pone.0241100.r002
8 Oct 2020 Decision Letter - Eric HY Lau, Editor PONE-D-20-13951R1 A comparison of health care worker-collected foam and polyester nasal swabs in convalescent COVID-19 patients PLOS ONE Dear Dr. Hart, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. The Authors have addressed most of the reviewers’ comments satisfactorily, but are expected to address the remaining criticisms by Reviewer #1. Please submit your revised manuscript by Nov 22 2020 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. Please include the following items when submitting your revised manuscript: A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'. An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'. If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter. If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols We look forward to receiving your revised manuscript. Kind regards, Eric HY Lau, Ph.D. Academic Editor PLOS ONE Additional Editor Comments (if provided): The Authors have addressed most of the reviewers’ comments satisfactorily, but are expected to address the remaining criticisms by Reviewer #1. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: (No Response) Reviewer #2: All comments have been addressed ******** 2. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #2: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: No Reviewer #2: Yes ****** 4. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: No Reviewer #2: Yes ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes Reviewer #2: Yes ****** 6. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: Thank you for the considerable revision you made. This revision is substantially clearer and more complete. A few comments: line 249-250 in the discussion: there is a sentence fragment that needs to be removed or completed. Although I will not "require" it, it would be helpful to provide p-values for comparisons across swab types and/or storage. Reviewer #2: 1. The study presents the results of original research. 2. Results reported have not been published elsewhere. 3. Experiments, statistics, and other analyses are performed to a high technical standard and are described in sufficient detail. 4. Conclusions are presented in an appropriate fashion and are supported by the data. 5. The article is presented in an intelligible fashion and is written in standard English. 6. The research meets all applicable standards for the ethics of experimentation and research integrity. 7. The article adheres to appropriate reporting guidelines and community standards for data availability. ****** 7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review?** For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No Reviewer #2: No [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.] While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0241100.r003
Revision 2
8 Oct 2020 Author Response The authors would like to thank the reviewers and editor for their careful review and consideration of our manuscript titled “A comparison of health care worker-collected foam and polyester nasal swabs in convalescent COVID-19 patients”. Below we provide a point by point response to the reviewer comments below: • line 249-250 in the discussion: there is a sentence fragment that needs to be removed or completed. Thank you for catching the error. The sentence fragment has been removed in the resubmitted manuscript. • Although I will not "require" it, it would be helpful to provide p-values for comparisons across swab types and/or storage. We have now provided p-values from a Fisher’s Exact test for the three polyester versus foam sensitivity comparisons. The p-values are included in the main text and in Tables 2-4. In Table 2, p-values were not included for the visit specific sensitivity comparisons due to small sample sizes. Two sentences have been added to the Statistical analysis section to explain methods used for p-value calculations. No additional changes were made to the manuscript. We again thank the reviewers for their constructive feedback which has led to a much improved manuscript. Attachments Attachment Submitted filename: Response to Reviewers Round 2.docx https://doi.org/10.1371/journal.pone.0241100.r004
9 Oct 2020 Decision Letter - Eric HY Lau, Editor A comparison of health care worker-collected foam and polyester nasal swabs in convalescent COVID-19 patients PONE-D-20-13951R2 Dear Dr. Hart, We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements. Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication. An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org. If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. Kind regards, Eric HY Lau, Ph.D. Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0241100.r005
Formally Accepted
16 Oct 2020 Acceptance Letter - Eric HY Lau, Editor PONE-D-20-13951R2 A comparison of health care worker-collected foam and polyester nasal swabs in convalescent COVID-19 patients Dear Dr. Hart: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Eric HY Lau Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0241100.r006

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