-->PNTD-D-25-02109

Genetic background and immune response in paracoccidioidomycosis: a systematic review and meta-analysis of single nucleotide variants

PLOS Neglected Tropical Diseases

Dear Dr. Venturini,

Thank you for submitting your manuscript to PLOS Neglected Tropical Diseases. After careful consideration, we feel that it has merit but does not fully meet PLOS Neglected Tropical Diseases'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 within by Feb 24 2026 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 plosntds@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pntd/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

* A letter that responds to each point raised by the editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. This file does not need to include responses to any formatting updates and technical items listed in the 'Journal Requirements' section below.

* 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, competing interests statement, or data availability statement, please make these updates within the submission form at the time of resubmission. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

We look forward to receiving your revised manuscript.

Kind regards,

Joshua Nosanchuk, MD

Section Editor

PLOS Neglected Tropical Diseases

Shaden Kamhawi

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-4304-636XX

Paul Brindley

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-1765-0002

Journal Requirements:

1) We noticed that you used “unpublished data" in the manuscript. We do not allow these references, as the PLOS data access policy requires that all data be either published with the manuscript or made available in a publicly accessible database. Please amend the supplementary material to include the referenced data or remove the references.

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Reviewers' Comments:

Reviewer's Responses to Questions

Key Review Criteria Required for Acceptance?

As you describe the new analyses required for acceptance, please consider the following:

Methods:

-Are the objectives of the study clearly articulated with a clear testable hypothesis stated?

-Is the study design appropriate to address the stated objectives?

-Is the population clearly described and appropriate for the hypothesis being tested?

-Is the sample size sufficient to ensure adequate power to address the hypothesis being tested?

-Were correct statistical analysis used to support conclusions?

-Are there concerns about ethical or regulatory requirements being met?

Reviewer #1: (No Response)

Reviewer #2: The study presents clearly defined objectives and the methodology used is appropriate.

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Results

-Does the analysis presented match the analysis plan?

-Are the results clearly and completely presented?

-Are the figures (Tables, Images) of sufficient quality for clarity?

Reviewer #1: (No Response)

Reviewer #2: The analyses performed are generally consistent with the proposed analysis plan; however, the manuscript has limitations in data analysis that were not adequately addressed or discussed. The suggestions and notes have been included in the section "Summary and General Comments".

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Conclusions

-Are the conclusions supported by the data presented?

-Are the limitations of analysis clearly described?

-Do the authors discuss how these data can be helpful to advance our understanding of the topic under study?

-Is public health relevance addressed?

Reviewer #1: (No Response)

Reviewer #2: The conclusions are generally supported by the data presented. The manuscript acknowledges some limitations of the analyses; however, additional limitations should be more clearly addressed and discussed.

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Editorial and Data Presentation Modifications?

Use this section for editorial suggestions as well as relatively minor modifications of existing data that would enhance clarity. If the only modifications needed are minor and/or editorial, you may wish to recommend “Minor Revision” or “Accept”.

Reviewer #1: (No Response)

Reviewer #2: (No Response)

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Summary and General Comments

Use this section to provide overall comments, discuss strengths/weaknesses of the study, novelty, significance, general execution and scholarship. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. If requesting major revision, please articulate the new experiments that are needed.

Reviewer #1: The manuscript by Coelho et al., addresses an important topic: the genetic basis of susceptibility to paracoccidioidomycosis (PCM), a neglected tropical disease endemic to Latin America. The review and meta-analysis are relevant. However, the study faces methodological limitations that affect the robustness of its conclusions.

Concerns:

The meta-analysis includes only two studies per SNV, which severely limits statistical power. This should be emphasized more clearly in the abstract and discussion.

Funnel plot asymmetry tests were omitted due to insufficient studies, this limitation should be stated in the methods and discussion.

Only one study applied multiple testing correction. The absence of FDR or Bonferroni adjustments in most studies increases the risk of false positives. This limitation should be more prominently addressed.

The discussion is somewhat repetitive. It also should separate methodological limitations from biological interpretation.

Figure captions must be improved to make them self-explanatory.

Reviewer #2: This manuscript by Coelho and colleagues addresses an important and underexplored topic. It looks at how host genetic differences affect susceptibility and clinical outcomes in paracoccidioidomycosis (PCM), which is a neglected disease relevant to public health in Latin America. The authors conducted a systematic review and reanalysis of published studies that focus on single nucleotide variants (SNVs) in immune-related genes and carried out a limited meta-analysis. Their effort to critically reassess published data, including Hardy-Weinberg equilibrium (HWE), inheritance models, and effect sizes, is commendable and meets the standards expected by this journal. However, despite its strengths, the manuscript has some problems that significantly limit its impact and may bias the conclusions. The main issues are discussed below.

Major concerns

1. A central limitation of the manuscript is the assumption that PCM susceptibility and severity are driven mainly by host genetic and immunological factors. While the introduction and discussion emphasize host immune responses, such as the Th1/Th2 balance, cytokine polymorphisms, PRRs, and VDR signaling, they largely overlook the role of the fungal pathogen itself. PCM is defined by host-pathogen interaction. There are well-documented differences in virulence, immunomodulatory capacity, thermotolerance, antigen expression, and cell wall composition among Paracoccidioides species and strains, such as P. brasiliensis and P. lutzii. These differences have significant effects on disease outcomes. The manuscript fails to adequately discuss how fungal genetic diversity, virulence factors, or strain-specific traits may influence host genetic associations. This omission risks presenting an oversimplified and potentially misleading view that PCM is “primarily a host-driven disease.” In reality, disease development stems from a dynamic interaction between host susceptibility and fungal pathogenicity. Additionally, it should be noted that the studies cited in references 7, 8, 9, 10, and 11 exclusively used the P. brasiliensis Pb18 isolate. Prolonged in vitro cultivation of pathogenic fungi is known to lead to significant changes in the expression of virulence-related components, often resulting in attenuated phenotypes. This laboratory adaptation can affect cell wall composition, secretion of enzymes, host-pathogen interactions, and stress response pathways. Therefore, relying on a single isolate that may have undergone changes due to long-term culture presents an additional limitation of these studies. This should be considered when interpreting conclusions related to fungal virulence and host-pathogen interactions. The authors should acknowledge this limitation and frame their conclusions within a host-pathogen interaction model instead of a host-centric view.

2. Though the manuscript correctly highlights associations involving IL10 and VDR SNVs, the biological interpretation remains shallow. Most of the variants mentioned are found in promoter, intronic, or untranslated regions, yet there is little discussion regarding whether these SNVs have any functional impact, such as altered cytokine production, receptor expression, or signaling efficiency in PCM or related fungal infections.

3. Another significant weakness is the inconsistency in defining “exposure” and “controls” across studies. Only two studies included controls with confirmed exposure to Paracoccidioides, while the others depended on apparently healthy individuals whose exposure status was unknown. Given the high prevalence of subclinical infection in endemic areas, this poses a critical confounding factor. The manuscript notes that none of the significant associations were convincingly tied to clinical form or disease severity, but it doesn’t integrate this finding into its conclusions effectively. As a result, the conclusions about “susceptibility” are vague and may be confused with exposure, infection, and progression.

Minor concerns

a) The introduction focuses heavily on immunology and could benefit from a brief section that recognizes fungal diversity and virulence as important factors in PCM.

b) Some repetition occurs in the Discussion concerning HWE and multiple testing corrections; this could be streamlined.

c) The authors might consider including a reformulated version of Supplementary Table 1S, retaining its main points of the table, in the main text, as this could improve clarity and strengthen the presentation of the results.

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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.]

Figure resubmission:

While revising your submission, we strongly recommend that you use PLOS’s NAAS tool (https://ngplosjournals.pagemajik.ai/artanalysis) to test your figure files. NAAS can convert your figure files to the TIFF file type and meet basic requirements (such as print size, resolution), or provide you with a report on issues that do not meet our requirements and that NAAS cannot fix.

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After uploading your figures to PLOS’s NAAS tool - https://ngplosjournals.pagemajik.ai/artanalysis, NAAS will process the files provided and display the results in the "Uploaded Files" section of the page as the processing is complete. If the uploaded figures meet our requirements (or NAAS is able to fix the files to meet our requirements), the figure will be marked as "fixed" above. If NAAS is unable to fix the files, a red "failed" label will appear above. When NAAS has confirmed that the figure files meet our requirements, please download the file via the download option, and include these NAAS processed figure files when submitting your revised manuscript.-->

Reproducibility:

To enhance the reproducibility of your results, we recommend that authors of applicable studies deposit laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. Additionally, PLOS ONE offers an option to publish peer-reviewed clinical study protocols. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols

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-->PNTD-D-25-02109R1

Genetic background and immune response in paracoccidioidomycosis: a systematic review and meta-analysis of single nucleotide variants

PLOS Neglected Tropical Diseases

Dear Dr. Venturini,

Thank you for submitting your manuscript to PLOS Neglected Tropical Diseases. Although it is clear that thoughtful revisions have been made by the authors, the response to reviewers document does not provide a point by point response to individual comments from the two prior reviewers, which is required in order to effectively assess whether the changes in the revision are sufficient to address ALL the issues raised during review. 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 within by Mar 18 2026 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 plosntds@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pntd/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

* A letter that responds to each point raised by the editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. This file does not need to include responses to any formatting updates and technical items listed in the 'Journal Requirements' section below.

* 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, competing interests statement, or data availability statement, please make these updates within the submission form at the time of resubmission. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

We look forward to receiving your revised manuscript.

Kind regards,

Joshua Nosanchuk

Section Editor

PLOS Neglected Tropical Diseases

Shaden Kamhawi

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-4304-636XX

Paul Brindley

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-1765-0002

Reviewers' Comments:

Need to provide formal point by point responses to reviewer comments to the original submission.

[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.]

Figure resubmission:

-->While revising your submission, we strongly recommend that you use PLOS’s NAAS tool (https://ngplosjournals.pagemajik.ai/artanalysis) to test your figure files. NAAS can convert your figure files to the TIFF file type and meet basic requirements (such as print size, resolution), or provide you with a report on issues that do not meet our requirements and that NAAS cannot fix.-->-->

After uploading your figures to PLOS’s NAAS tool - https://ngplosjournals.pagemajik.ai/artanalysis, NAAS will process the files provided and display the results in the "Uploaded Files" section of the page as the processing is complete. If the uploaded figures meet our requirements (or NAAS is able to fix the files to meet our requirements), the figure will be marked as "fixed" above. If NAAS is unable to fix the files, a red "failed" label will appear above. When NAAS has confirmed that the figure files meet our requirements, please download the file via the download option, and include these NAAS processed figure files when submitting your revised manuscript.-->

Reproducibility:

To enhance the reproducibility of your results, we recommend that authors of applicable studies deposit laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. Additionally, PLOS ONE offers an option to publish peer-reviewed clinical study protocols. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols

-->

Dear Dr. Venturini,

The authors are applauded for their rigorous response to the feedback on their work. We are pleased to inform you that your manuscript 'Genetic background and immune response in paracoccidioidomycosis: a systematic review and meta-analysis of single nucleotide variants' has been provisionally accepted for publication in PLOS Neglected Tropical Diseases.

Before your manuscript can be formally accepted you will need to complete some formatting changes, which you will receive in a follow up email. A member of our team will be in touch with a set of requests.

Please note that your manuscript will not be scheduled for publication until you have made the required changes, so a swift response is appreciated.

IMPORTANT: The editorial review process is now complete. PLOS will only permit corrections to spelling, formatting or significant scientific errors from this point onwards. Requests for major changes, or any which affect the scientific understanding of your work, will cause delays to the publication date of your manuscript.

Should you, your institution's press office or the journal office choose to press release your paper, you will automatically be opted out of early publication. We ask that you notify us now if you or your institution is planning to press release the article. All press must be co-ordinated with PLOS.

Thank you again for supporting Open Access publishing; we are looking forward to publishing your work in PLOS Neglected Tropical Diseases.

Best regards,

Joshua Nosanchuk

Section Editor

PLOS Neglected Tropical Diseases

Shaden Kamhawi

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-4304-636XX

Paul Brindley

co-Editor-in-Chief

PLOS Neglected Tropical Diseases

orcid.org/0000-0003-1765-0002

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