-->PONE-D-25-13754-->-->Investigation of mobile genetic elements and their association with antibiotic resistance genes in clinical pathogens worldwide-->-->PLOS ONE

Dear Dr. Johansson,

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.

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We look forward to receiving your revised manuscript.

Kind regards,

Mabel Kamweli Aworh, DVM, MPH, PhD. FCVSN

Academic Editor

PLOS ONE

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Additional Editor Comments:

In revising your manuscript, please ensure the following key points are addressed in addition to responding to all reviewers’ comments:

1. Limitations:
   Please include a dedicated paragraph in the final part of your Discussion section clearly outlining the limitations of your study. This reflection should help contextualize your findings and guide interpretation.
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   Add a Conclusion section at the end of the manuscript that succinctly summarizes the main findings and their implications.
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   Within the Conclusion section, please include recommendations derived from your key findings and/or suggest potential future research directions that could build on your work.

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Reviewers' comments:

Reviewer's Responses to Questions

-->Comments to the Author

1. 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

Reviewer #3: Yes

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-->2. Has the statistical analysis been performed appropriately and rigorously? -->

Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: Yes

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-->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: No

Reviewer #2: Yes

Reviewer #3: Yes

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-->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: No

Reviewer #2: Yes

Reviewer #3: Yes

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-->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: Comments

General comments

• The research topic is very interesting, and the subject is important. However, the presentation needs to be improved. The presentation of the methodologies could be improved. The findings should be well discussed because of the literature and not opinions.

• 74 authors are too many (though there is no cut off for the number of authors for a manuscript). However, it is clear from the author's contribution that the list from lines 734-738 are the primary authors of this research. It is suggested to the authors that all other members be acknowledged for their contribution to the paper.

Abstract

• Line 122: Are 59 diagnostic units enough to represent a snapshot of clinical infections worldwide? The methodology states 35 countries. What is the justification that 59 diagnostic units from 35 countries are enough to give a clinical representation worldwide?

• Line 142: Discussion on the identified 21 genes must be expanded

Introduction

1. Line 161: Why exclude important global pathogens like Salmonella, Campylobacter, and Listeria spp.

2. Lines requiring citations

a. Lines 165-167

b. Lines 172-174.

c. Many such sentences without appropriate citations exist in the article

3. Lines 185-187: Sentence is too long

4. Line 190: The collection of samples in a single year reflects a one-time period sampling, and this must be reflected in the abstract or title

5. Line 191: Statement is inappropriate. Symptoms of disease caused by a pathogen are more appropriate to say rather than symptoms of a pathogen

Methods

6. Line 200: Is TWIW a name or an abbreviation? Please clarify. If it is an abbreviation, the full name should be given. All such abbreviations must be defined

7. Line 211: What comprises these seven regions is undefined

8. Line 217: QC filtering may not be needed as it is deemed part of read processing

Results

9. Line 330: “(Error! Reference source not found)”. This must be corrected by supplying all appropriate citations. The same issue is found in other parts of the manuscript

10. Lines 331-337: The nomenclature of Beta-lactamase genes must be consistent following default patterns like blaCTX-M

DISCUSSION

11. Generally, this portion must be improved to really discuss the results in the context of appropriate literature providing references

12. Line 549: For abbreviation, the name should appear first, followed by the abbreviation. Example Escherichia coli (E. coli)

REFERENCES

13. The date of assessment of all Web pages and other documents without doi(s) must be provided in addition to the links

FIGURES

14. Figures all well prepared

Reviewer #2: This is an important study which aim to unravel the role of mobile genetic elements in antimicrobial resistance. I have attached some minor revisions below:

General

There are several instances of “Error reference source not found”, please review and update.

Abstract

The word count of the abstract exceeded the PLOS ONE requirement of 300 words maximum.

Introduction

The authors should include the implications of this study and how it contributes towards antimicrobial resistance solutions in public health.

Methods

Line 211: clarify what this means “and is NOT referred to as the “primary” dataset”.

Results

Line 373-375: The appropriate figure for reference is Figure S8. See the Supporting information list and caption too

Line 375-378: The appropriate figure to reference is Figure S9. See the Supporting information list and caption too.

Supporting information

The list (title and caption) for figures in the supporting information at the end of the manuscript does not match the order of their respective figures starting from Figure S3 up to S9. I will suggest a review of the figures referenced in the result section too.

Reviewer #3: Very good work and presentation however, there suggestions for improvement

1. In the Abstract, results were presented more like generalized and this was seen in the Results section of the manuscript. The authors would need to be more specific in describing that results in terms of number and/or percentages with possible denominators where necessary. Using generalized terms like "Many, Most, Some" does not give a comprehensive idea of the magnitude of ARGs, iMGEs including their diversity in the species of interest. I could see that the charts and table show some quantitative results but it will be good to include them in the results description as well as put them in context.

2. The missing references sources should to fixed by the authors. It makes the article easier to follow should the review wants to check the cited/referenced articles.

3. The section on "Controlling for high relatedness in the dataset" can be moved to methods because it is not actually describing the results. It describes more of methods. Same as the opening paragraph in the results except the authors consider rephrasing them to actually mean the results if they must stay in the result section.

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-->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.

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Reviewer #1: No

Reviewer #2: Yes:  Damilola Odumade

Reviewer #3: No

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[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.

-->PONE-D-25-13754R1-->-->Investigation of mobile genetic elements and their association with antibiotic resistance genes in clinical pathogens worldwide-->-->PLOS ONE

Dear Dr. Johansson,

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 Aug 15 2025 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,

Mabel Kamweli Aworh, DVM, MPH, PhD. FCVSN

Academic Editor

PLOS ONE

Journal Requirements:

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

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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: All comments have been addressed

Reviewer #2: All comments have been addressed

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 #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

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

Reviewer #1: Yes

Reviewer #2: N/A

Reviewer #3: N/A

**********

-->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: Yes

Reviewer #2: Yes

Reviewer #3: 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

Reviewer #3: 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:  All comments have been addressed by the authors and I recommend accepting the manuscript for publication

Reviewer #2:  Thank you for addressing the previous comments. However, there are a few additional issues that need to be resolved:

Figure 1 – Geographic distribution of β-lactamase gene families in E. coli

The current map in Figure 1 does not include the African continent, which creates an inconsistency (compare with Figure 2) and Africa is explicitly referenced in Line 309-310. I recommend updating the figure to include Africa in order to accurately reflect the geographic scope discussed in the text.

Line 354 – Inconsistent Figure Referencing

The current reference style in this line — “S8 Fig S8” — appears inconsistent. I suggest the authors adopt a consistent and standard format for referencing all figures throughout the manuscript (e.g., “S8 Fig” or “Fig S8,” not both), and apply this uniformly across the document.

Reviewer #3:  The authors have made all corrections that I recommended that the editors review for guidelines and then publish.

**********

-->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: Yes:  Damilola Odumade

Reviewer #3: 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.-->

Investigation of mobile genetic elements and their association with antibiotic resistance genes in clinical pathogens worldwide

PONE-D-25-13754R2

Dear Dr. Johansson,

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,

Mabel Kamweli Aworh, DVM, MPH, PhD. FCVSN

Academic Editor

PLOS ONE

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