Dear Professor Na-Bangchang,

Thank you very much for submitting your manuscript "Racial disparities in the promoter region G-308A polymorphism (rs1800629) of the tumor necrosis factor-α (TNF-α) gene associated with susceptibility to severe Plasmodium falciparum malaria: a meta-analysis" for consideration at PLOS Neglected Tropical Diseases. As with all papers reviewed by the journal, your manuscript was reviewed by members of the editorial board and by several independent reviewers. The reviewers appreciated the attention to an important topic. Based on the reviews, we are likely to accept this manuscript for publication, providing that you modify the manuscript according to the review recommendations.

Please prepare and submit your revised manuscript within 30 days. If you anticipate any delay, please let us know the expected resubmission date by replying to this email.

When you are ready to resubmit, please upload the following:

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Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out

[2] Two versions of the revised manuscript: one with either highlights or tracked changes denoting where the text has been changed; the other a clean version (uploaded as the manuscript file).

Important additional instructions are given below your reviewer comments.

Thank you again for your submission to our journal. We hope that our editorial process has been constructive so far, and we welcome your feedback at any time. Please don't hesitate to contact us if you have any questions or comments.

Sincerely,

Walderez O. Dutra, PhD.

Section Editor

PLOS Neglected Tropical Diseases

Walderez Dutra

Section Editor

PLOS Neglected Tropical Diseases

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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: The methodology is sound and proper statistical analysis methods were used for this meta-analysis study.

Reviewer #2: -Are the objectives of the study clearly articulated with a clear testable hypothesis stated? YES

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

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

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

-Were correct statistical analysis used to support conclusions? NO

-Are there concerns about ethical or regulatory requirements being met? NO

Reviewer #3: The objectives of the study are specifically and clearly to achieve through the study. For the study design is appropriate becuase authors select specifically research articles by using PRISMA guidelines and create excellently inclusion and exclusion criteria leading to have finally the valuable articles (8 out of 13,087). Statistic analysis for this study is appropriate and support the conclusions.

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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: The results are appropriately presented.

Reviewer #2: -Does the analysis presented match the analysis plan? YES

-Are the results clearly and completely presented? YES

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

Reviewer #3: For the results, authors show the clearly and completely information that they have. They try to explain step by step about association between TNF alpha polymorphism and severity of malaria. Furthermore, they try to go deeply into the subgroup of subjects such as the ethnicity. Lastly, the figures and tables are sufficient quality for clarity. It is easy to understand the points or informations of authors with in a second.

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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: The conclusions are supported by the data presented. The limitations of the study are well described. However, the authors do not describe how their findings ccontribute to the understanding and contribution of this polymorphism in complicated malaria among the Asians.

Reviewer #2: -Are the conclusions supported by the data presented? PARTIALLY

-Are the limitations of analysis clearly described? YES

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

-Is public health relevance addressed? NO

Reviewer #3: For discussion part, authors try to clearify about association between TNF alpha gene polymorphism and severity of malaria in each ethnic group. They also discuss clearly about limitation and strengths of this study.The conclusion is support clearly the objectives. They try to show the association of severe malaria and Asian population. This point will be knowledge for physician having awareness of severe malaia in Asian population who have TNF alpha polymorphism. So this study have public health relevance addressed.

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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 recommendation

Reviewer #2: (No Response)

Reviewer #3: Typo is shown in result's part of manuscript (line 271, page 12): "...... the statistical power ranged from 18.7 to 80.1% , in which .................." should be "...... the statistical power ranged from 12.4 to 80.1% , in which .................." .

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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: This is a nice work of metanalysis and the authors concluded that the polymorphism TNFa -308 is associated with susceptibility among the Asians but not among the Africans. However, my main concern in this study is the minor allele frequency among the Asians. There is a great disparity ranging from 0.04 to 0.26 for the minor allele A among the Asians. Of note. there are two studies from India with completely different minor allele frequency (0.07 and 0.26). It will be interesting for the authors to discuss this disparity and how this could have affected the outcome of the analysis.

Reviewer #2: Major and minor comments follow:

MAJOR

Please modify the title, as it is it does not reflect what the authors have studied, since the association with ethnicity is based on the stratification studies, no proper analysis for racial disparities between Africans and Asians were compared.

It is advised to maintain OR estimates and CIs, leave to one decimal case, more than that is not relevant to understand the effect estimate, and it pollutes the manuscript with numbering.

In Methods, line 183, what does criteria vii refers to as duplicated genotype data? Please clarify.

In Methods, line 197, what does corrections for multiplicity refer to? Is it not referring to multiple comparisons’ correction? Please clarify.

In Methods, line 215, please justify the rationale for applicating the Laplace correction in this scenario? rs1800629 has an overall MAF less than 10%, even reaching to 2% in EAS populations according to ENSEMBL. Adding a +1 to genotype counts which are probably to AA genotype is a highly sensitive modification in terms of a SNP that already has a low MAF. Also, having 0 genotype counts doesn’t make inviably the performance and calculation neither of the most relevant genetic models (dominant, codominant), or even the OR.

Please justify in detail.

In Results Line 321, the results from Asians and Africans cannot be compared between them, the results for Africans were not significant, so further description that favour UM lack the basis of statistically significant and should not be interpreted. Please remove.

In Results if the study by Ubalee, 2001 deviated in HWE for the control group there should have been a separate analysis in which the study was excluded from summary statistics, as it has been clearly stated how HWE deviations imply in potential genotyping errors and include it in Supplementary material if the association stands after the exclusion of this study.

MINOR

In abstract and please review in the rest of the text, when referring to gene and SNP nomenclature modify to TNFA G>A polymorphism or TNFA gene or polymorphism when required.

In Introduction, line 151 what does UM stands for?

In Materials and Methods please modify the subtitle (line 164) Case-control definition and remove lines 167 -169 that indicate that the meta-analysis followed PRISMA guidelines to include them as the initial line for Publication Search and Study selection in line 172.

Minor Typing and concordance in English redaction errors should be revised. E.g. line 221 in methods correct for heterogeneity.

Line 456. Remove bunch and change for other term.

Reviewer #3: This manuscript has a good composition, harmonization and clarification of writing. Value of meta-analysisof to this study is the usefulness of the association of TNF alpha gene polymorphism and severity of malaria in Asian population. It is valuable information for physician to have more concerned about severe malaria in Asian than other ethnic group.

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Reviewer #1: No

Reviewer #2: Yes: Lucia E. Alvarado-Arnez

Reviewer #3: No

Figure Files:

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References

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.

Dear Professor Na-Bangchang,

Thank you very much for submitting your manuscript "Racial disparities in the promoter region G-308A polymorphism (rs1800629) of the tumor necrosis factor-α (TNF-α) gene associated with susceptibility to severe Plasmodium falciparum malaria: a meta-analysis" for consideration at PLOS Neglected Tropical Diseases. As with all papers reviewed by the journal, your manuscript was reviewed by members of the editorial board and by several independent reviewers. In light of the reviews (below this email), we would like to invite the resubmission of a significantly-revised version that takes into account the reviewers' comments.

We cannot make any decision about publication until we have seen the revised manuscript and your response to the reviewers' comments. Your revised manuscript is also likely to be sent to reviewers for further evaluation.

When you are ready to resubmit, please upload the following:

[1] A letter containing a detailed list of your responses to the review comments and a description of the changes you have made in the manuscript. Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out.

[2] Two versions of the revised manuscript: one with either highlights or tracked changes denoting where the text has been changed; the other a clean version (uploaded as the manuscript file).

Important additional instructions are given below your reviewer comments.

Please prepare and submit your revised manuscript within 60 days. If you anticipate any delay, please let us know the expected resubmission date by replying to this email. Please note that revised manuscripts received after the 60-day due date may require evaluation and peer review similar to newly submitted manuscripts.

Thank you again for your submission. We hope that our editorial process has been constructive so far, and we welcome your feedback at any time. Please don't hesitate to contact us if you have any questions or comments.

Sincerely,

Abhay R Satoskar

Section Editor

PLOS Neglected Tropical Diseases

Walderez Dutra

Section Editor

PLOS Neglected Tropical Diseases

***********************

Figure 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. 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 us at figures@plos.org.

Data Requirements:

Please note that, as a condition of publication, PLOS' data policy requires that you make available all data used to draw the conclusions outlined in your manuscript. Data must be deposited in an appropriate repository, included within the body of the manuscript, or uploaded as supporting information. This includes all numerical values that were used to generate graphs, histograms etc.. For an example see here: http://www.plosbiology.org/article/info%3Adoi%2F10.1371%2Fjournal.pbio.1001908#s5.

Reproducibility:

To enhance the reproducibility of your results, we recommend that you deposit your 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 Professor Na-Bangchang,

We are pleased to inform you that your manuscript 'Tumor necrosis factor-α (TNF-α) -308G >A promoter polymorphism (rs1800629) promotes Asians in susceptibility to Plasmodium  falciparum severe malaria: a meta-analysis' 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,

Abhay R Satoskar

Section Editor

PLOS Neglected Tropical Diseases

Walderez Dutra

Section Editor

PLOS Neglected Tropical Diseases

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