-->PONE-D-25-35418-->-->Complete Molecular Spectrum of β-Globin Gene Mutations via Direct Sequencing Identifies Seven Novel Variants in β-Thalassemia Major-->-->PLOS ONE

Dear Dr. Othman,

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Nejat Mahdieh

Academic Editor

PLOS ONE

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Additional Editor Comments (if provided):

Reviewer #1:

Reviewer #2:

Reviewer #3:

[Note: HTML markup is below. Please do not edit.]

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: No

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: The author should indicate the scoring system used for pathogenicity classification. The ACMG 2015 guideline is the most widely used one in the field. Terms such as "benign mutations" do not conform to current, standard terminology. Novel variants should be reported according to standard terminology so they can be checked in related databases. For example, "NM number, nucleotide change" or "genome version, chromosome position."

Reviewer #2: This is a highly relevant and clinically important study. The use of full-gene sequencing has successfully uncovered a significant number of novel variants, providing a valuable update to the mutational spectrum of β-thalassemia in the Kurdish population of Iraq. The findings have direct implications for improving local genetic diagnostics and counselling. The manuscript is generally well-structured, but several key issues need to be addressed to strengthen the validity and clarity of the conclusions:

1. Methods: Mentioning the ACMG/AMP guidelines is good. Briefly state which specific criteria (e.g., PVS1, PM2) were used to classify the novel variants as pathogenic/likely pathogenic.

2.Results: The study describes finding many mutations but does not clearly report the actual genotypes of the patients. How many were compound heterozygous? How many were homozygous? For instance, which specific combinations of the pathogenic mutations were found in the transfusion-dependent patients? Please add a table or a summary in the Results section. This directly links your molecular findings to the clinical presentation (Transfusion-Dependent Thalassemia).

3. The article requires proofreading and structural revision (e.g., concertation in line 127 and some inconsistencies in font size).

Reviewer #3: The article provides a comprehensive overview of β-thalassemia, including genetic background, clinical classification, prevalence, and regional context (particularly Iraq and the Kurdish population). It is well-structured and demonstrates a clear rationale for the study. However, a few points need improvement:

The background is lengthy and sometimes repetitive. Condensing these parts would make it more focused.

Some sentences are overly long or contain minor grammatical issues (e.g., “several studies have been partially addressed…” should be “several studies have partially addressed…”). Editing for readability would improve impact.

Ensure consistency in formatting (spacing around years, punctuation). Some references (e.g., “(Huang et al., 2022)..”) have typographical errors.

Some sentences in the method section are long or awkwardly phrased. For example:

“Informed consent were obtained…” → should be “Informed consent was obtained…”

“concertation and purity” → should be “concentration and purity.”

Minor inconsistencies (spacing, capitalization, punctuation in citations).

The table footnotes should be expanded to provide complete explanations of all abbreviations used.

It would be better to include sequencing chromatogram figures of the newly identified mutations and to provide more detailed bioinformatic analyses regarding their pathogenicity.

In the table, references should be cited for the mutations that are reported as pathogenic.

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Reviewer #1: No

Reviewer #2: No

Reviewer #3: Yes:  Mahdieh Soveizi

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Complete Molecular Spectrum of β-Globin Gene Mutations via Direct Sequencing Identifies Seven Novel Variants in β-Thalassemia Major

PONE-D-25-35418R1

Dear Dr. Othman,

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.

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Kind regards,

Nejat Mahdieh

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

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

**********

-->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: The authors have carefully and satisfactorily responded to comments. The revised version demonstrates significant improvement in both structure and content. I find the manuscript ready for publication in its present form.

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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: Yes:  Mahdieh Soveizi

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