PONE-D-24-10038Using diagnostic data from veterinary diagnostic laboratories to unravel macroepidemiological aspects of porcine circoviruses 2 and 3 in the United States from 2002-2023.PLOS ONE

Dear Dr. Trevisan,

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Additional Editor Comments:

The manuscript has two critical points reported by the system that should be addressed before the system goes for review. The first point concerns the title of the manuscript, which is very similar (although concerning a different pathogen) to the title of a recent manuscript by the same author. The second point concerns the number of authors. It is unthinkable that almost 40 authors have worked on a manuscript for which we invite the authors to include only and exclusively those who have made a significant contribution to the manuscript.

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PONE-D-24-10038R1Using diagnostic data from veterinary diagnostic laboratories to unravel macroepidemiological aspects of porcine circoviruses 2 and 3 in the United States from 2002-2023.PLOS ONE

Dear Dr. Trevisan,

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 Jul 08 2024 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,

Gianmarco Ferrara, PhD, MVD

Academic Editor

PLOS ONE

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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: (No Response)

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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: Partly

Reviewer #2: Yes

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3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: I Don't Know

Reviewer #2: Yes

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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: No

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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

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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: Guillerme et al.'s study primarily provides a descriptive analysis of the epidemiology of PCV2 and PCV3 in the U.S. swine population over time. Overall, it sounds like the paper has the potential to make a significant contribution to the field.

To improve the paper, it would be advantageous to incorporate more comprehensive details regarding the diagnostic tests employed and the interpretation of the results. This would enhance the clarity and depth of the study's findings. Additionally, it's important for the paper to clearly outline its limitations. This could include discussing potential biases introduced by the use of data from multiple VDLs, variations in laboratory protocols, and any other factors that may have influenced the results or interpretation of the findings. Providing a comprehensive overview of the study's limitations will help readers better understand the scope and implications of the research.

Please incorporate details regarding the diagnostic testing process. Given the existence of various protocols, it's unclear whether all samples underwent testing via real-time PCR TaqMan-based, Sybrgreen, or conventional PCR methodologies, especially during the initial years.

Have the VDLs ever conducted quantitative real-time PCR using the PCV2/PCV3 protocols? It would provide insight into the virus's quantity, as indicated by the Ct values. This information could prove immensely valuable and should be incorporated.

Incorporating PCV2 subtyping data would greatly enhance the value of the paper if it's accessible.

Were submissions from other countries excluded?

Please incorporate the study's limitations concerning variations in protocols across different laboratories, challenges in interpreting Ct values, and any other issues identified during the analysis.

Please ensure all references align accurately with the corresponding text.

The figures appear blurry.

Figure 1 employs the same color code for both negative and non-informed data points."

In lines 58-60, it would be more precise to specify that PCV1 is a non-pathogenic virus."

Lines 64-66: The term Porcine circovirus-associated diseases (PCVAD) typically refer to PCV2. The references should be reviewed accordingly. Additionally, the references regarding economic losses require correction.

Please ensure all references align accurately with the corresponding text.

In lines 109-110, it should be revised to 'PCV2 and PCV3 real-time PCR cycle threshold (Ct).

Regarding line 192, include more detail in the samples not included.

For line 188, please provide more details about the confirmed tissue disease. Was it confirmed via ISH or IHC?

171-173. CT values obtained from real-time PCR assays indirectly reflect the amount of virus present in a sample. Due to the nature of PCR amplification, these values often follow a logarithmic rather than a linear scale. Consequently, caution should be exercised when averaging CT values, as direct arithmetic means may not accurately represent the results.

Reviewer #2: The manuscript is well written and clear, the introduction adequate, methods and results well described and commented, and the amount of data used is impressive. However, I have some major concerns about the effect that secular trends in terms of veterinary perception, knowledge, diagnostic test availability, and changes in used matrices might have. These factors could be even more relevant than the underlying biological phenomena, biasing the results too much and preventing any useful conclusions.

Additionally, why was a Ct cut-off used instead of copy number? Ct values are highly variable among laboratories and assays, while copy numbers can be more easily standardized using a quantified reference sample and a standard curve. Was the reproducibility among laboratories assessed? Even if the Ct values were comparable among the laboratories involved in the study, the provided cut-off is not useful for other laboratories. On the contrary, providing such data could be misleading and lead to misdiagnoses.

More detailed comments and suggestions are reported in the attached PDF

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Reviewer #1: Yes: Victor Neira

Reviewer #2: No

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Attachments

Attachment

Submitted filename: PONE-D-24-10038_R1_reviewer.pdf

Using diagnostic data from veterinary diagnostic laboratories to unravel macroepidemiological aspects of porcine circoviruses 2 and 3 in the United States from 2002-2023.

PONE-D-24-10038R2

Dear Dr. Trevisan,

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.

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Kind regards,

Gianmarco Ferrara, PhD, MVD

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

All comments have been addressed

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 #3: All comments have been addressed

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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 #3: Yes

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3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #3: 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 #3: Yes

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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 #3: Yes

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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 #3: (No Response)

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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 #3: No

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