No evidence of SARS-CoV-2 in hospitalized patients with severe acute respiratory syndrome in five Italian hospitals from 1st November 2019 to 29th February 2020

Donatella Panatto; Andrea Orsi; Beatrice Marina Pennati; Piero Luigi Lai; Stefano Mosca; Bianca Bruzzone; Patrizia Caligiuri; Christian Napoli; Enrico Bertamino; Giovanni Battista Orsi; Ilaria Manini; Daniela Loconsole; Francesca Centrone; Elisabetta Pandolfi; Marta Luisa Ciofi Degli Atti; Carlo Concato; Giulia Linardos; Andrea Onetti Muda; Massimiliano Raponi; Livia Piccioni; Caterina Rizzo; Maria Chironna; Giancarlo Icardi

doi:10.1371/journal.pone.0260947

Peer Review History

Original SubmissionApril 8, 2021
24 May 2021 Decision Letter - Wenbin Tan, Editor PONE-D-21-10636 No evidence of SARS-CoV-2 in hospitalized patients with severe acute respiratory syndrome in five Italian hospitals from 1st November 2019 to 29th February 2020 PLOS ONE Dear Dr. Panatto, 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. Please submit your revised manuscript by Jun 27 2021 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'. Please refer to the detailed comments from Reviewer 1 in the attachment for your revision. 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. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols. We look forward to receiving your revised manuscript. Kind regards, Wenbin Tan Academic Editor PLOS ONE 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 including your ethics statement: "The study protocol was approved by the Ethics Committee of the Liguria Region (Genoa, Italy) (n° 245/2019)." a) Please provide additional details regarding participant consent. In the ethics statement in the Methods and online submission information, please ensure that you have specified (1) whether consent was informed and (2) what type you obtained (for instance, written or verbal, and if verbal, how it was documented and witnessed). If your study included minors, state whether you obtained consent from parents or guardians. If the need for consent was waived by the ethics committee, please include this information. If you are reporting a retrospective study of medical records or archived samples, please ensure that you have discussed whether all data were fully anonymized before you accessed them and/or whether the IRB or ethics committee waived the requirement for informed consent. If patients provided informed written consent to have data from their medical records used in research, please include this information. Once you have amended this/these statement(s) in the Methods section of the manuscript, please add the same text to the “Ethics Statement” field of the submission form (via “Edit Submission”). For additional information about PLOS ONE ethical requirements for human subjects research, please refer to http://journals.plos.org/plosone/s/submission-guidelines#loc-human-subjects-research. 3) In the Methods section and the online ethics statement. Please provide some clarification whether the current ethics statement provided is for the DRIVE study or the current study reported. 4) Please include in your Methods section (or in Supplementary Information files) the participating hospitals/institutions. Reviewers' comments: Reviewer #1: All the sections of the manuscript need to be rewritten or to be rephrased with more focus on the aim of the work and the value of the results. the sections need to be better connected to show the importance of the findings. There are some drawbacks in the sampling and storing. These drawbacks should be discussed to clearly show their effects on the results and their reliability. Reviewer #2: Please address the following points: 1. Were the tests for all the viruses done simultaneously? The authors have to rule out the possibility that RNA in the samples was not degraded to yield a positive test for SARS-CoV-2. 2. Are the authors sure that the RT-PCR tests were real time? 3. Which of the four cities are located in Northern Italy (the worst affected region)? 4. ECDC? European Center for Disease Control? 5. Table 1: What does N/A mean? Explain. 6. Write figure legends or at least their titles. Attachments Attachment Submitted filename: PONE-D-21-10636_reviewer_corrected.pdf https://doi.org/10.1371/journal.pone.0260947.r001
Revision 1
2 Sep 2021 Author Response Journal requirements 1) The manuscript meets PLOS ONE’s style requirements. 2) The sentence “The study protocol was approved by the Ethics Committee of the Liguria Region (Genoa, Italy) (n◦ 245/2019) as coordinator center and subsequently approved by all local the Ethics Committees” has been added. a) In the manuscript, the paragraph “Ethics Statement”, containing the EC approval code and the detail on informed consent, has been added. 3) The sentence “Informed written consent was obtained from each patient, as required by the DRIVE study protocol [15]” has been added. 4) In the Methods section, the description of the hospitals has been added. Reviewers’ comments Reviewer #1: All the sections of the manuscript need to be rewritten or to be rephrased with more focus on the aim of the work and the value of the results. the sections need to be better connected to show the importance of the findings. There are some drawbacks in the sampling and storing. These drawbacks should be discussed to clearly show their effects on the results and their reliability. Done As required by reviewer 1, all sections have been rewritten in order to better focus on the aims and results of the study. All revisions are well visible in the revised manuscript with track change. Reviewer #2: 1) Were the tests for all the viruses done simultaneously? The authors have to rule out the possibility that RNA in the samples was not degraded to yield a positive test for SARS-CoV-2. The text has been modified in order to better explain that the molecular analyses for influenza and SARS-CoV-2 were not performed simultaneously. The molecular test for influenza was performed before the test for SARS-CoV-2. The study biases have been better explained. 2) Are the authors sure that the RT-PCR tests were real time? Yes, all molecular tests were performed by means of the real-time method. 3) Which of the four cities are located in Northern Italy (the worst affected region)? Genoa is the only city located in Northern Italy 4) ECDC? European Center for Disease Control? Yes 5) Table 1: What does N/A mean? Explain. The notation N/A (not available) has been explained 6) Write figure legends or at least their titles. Done References have been updated and renumbered. Figure 2 has been improved. Attachments Attachment Submitted filename: Response_to_reviewers_PONE-D-21-10636.docx https://doi.org/10.1371/journal.pone.0260947.r002
23 Sep 2021 Decision Letter - Wenbin Tan, Editor PONE-D-21-10636R1No evidence of SARS-CoV-2 in hospitalized patients with severe acute respiratory syndrome in five Italian hospitals from 1st November 2019 to 29th February 2020PLOS ONE Dear Dr. Panatto, 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. Please submit your revised manuscript by Nov 07 2021 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: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols. We look forward to receiving your revised manuscript. Kind regards, Wenbin Tan Academic Editor PLOS ONE Journal Requirements: Review Comments to the Author Reviewer #1: All the comments have been addressed. I recommend the authors to rewrite the conclusion section Reviewer #2: The authors have indicated that the detection for SARS-CoV2 was done on archived samples that were previously tested for Influenza viruses. They found that all the samples were found negative for the coronavirus. The onus is on the researchers to show that the RNA in the samples was not degraded. They should have used some positive controls for these samples. For example test again for the already detected Influenza virus, for example. Furthermore, they should indicate whether they re-extracted RNA from the samples or used already extracted RNA from these samples. These are extremely important points. For this reason, major revision is indicated. https://doi.org/10.1371/journal.pone.0260947.r003
Revision 2
9 Nov 2021 Author Response Genoa, 4 November 2021 Dear Academic Editor PLOS ONE, Thank you for giving us the opportunity to resubmit our manuscript “No evidence of SARS-CoV-2 in hospitalized patients with severe acute respiratory syndrome in five Italian hospitals from 1st November 2019 to 29th February 2020” (PONE-D-21-10636) for publication in your journal. The reviewers' comments were insightful and pertinent. Below, you will find the reviewers' reports with point-by-point replies and explanations of the modifications made. The reviewers’ comments are in bold, while our replies are in italics. The revised manuscript has been uploaded to the system in two versions: one with “track changes” and the other without “track changes”. Sincerely, Prof. Donatella Panatto Reviewers’ comments Reviewer #1: All the comments have been addressed. I recommend the authors to rewrite the conclusion section Done Reviewer #2: The authors have indicated that the detection for SARS-CoV2 was done on archived samples that were previously tested for Influenza viruses. They found that all the samples were found negative for the coronavirus. The onus is on the researchers to show that the RNA in the samples was not degraded. They should have used some positive controls for these samples. For example test again for the already detected Influenza virus. Furthermore, they should indicate whether they re-extracted RNA from the samples or used already extracted RNA from these samples. These are extremely important points. For this reason, major revision is indicated. As required by reviewer 2, we have better specified, in the Materials and Methods section and Discussion section, the method of verifying the correct re-extraction of the genetic material from the sample and the integrity of the viral RNA. Attachments Attachment Submitted filename: Response_to_reviewers_PONE-D-21-10636_4112021.docx https://doi.org/10.1371/journal.pone.0260947.r004
22 Nov 2021 Decision Letter - Wenbin Tan, Editor No evidence of SARS-CoV-2 in hospitalized patients with severe acute respiratory syndrome in five Italian hospitals from 1st November 2019 to 29th February 2020 PONE-D-21-10636R2 Dear Dr. Panatto, 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, Wenbin Tan Academic Editor PLOS ONE Additional Editor Comments: The 2nd reviewer's comment was well addressed. https://doi.org/10.1371/journal.pone.0260947.r005
Formally Accepted
25 Nov 2021 Acceptance Letter - Wenbin Tan, Editor PONE-D-21-10636R2 No evidence of SARS-CoV-2 in hospitalized patients with severe acute respiratory syndrome in five Italian hospitals from 1^st November 2019 to 29^th February 2020 Dear Dr. Panatto: 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. Wenbin Tan Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0260947.r006

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