PONE-D-23-17864Population genetic diversity and structure of the endangered species  Tetracentron sinense  Oliver ( Tetracentraceae ) with SNPs based on RAD sequencingPLOS ONE

Dear Dr. Gan,

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The reviewers believe that the paper needs major revision in terms of English language, presenting the results in a palatable way and further analysis of population genetics parameter, which you find all in reviewers comments.

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Xiaohong Gan: Funding acquisition; Investigation; Project administration; Supervision; Visualization; Writing–review & editing."

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Dear authors,

The reviewers of the manuscript have raised issues on the language aspect and additional analysis on the manuscript and overall presentation of the result. The paper needs major revision. Please address the issues raised and send us the revised version.

Best regards,

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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: Yes

Reviewer #2: Yes

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

Reviewer #1: Yes

Reviewer #2: 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

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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

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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: Dear authors

The following modifications are required

Abstract

� In general, this section is poorly written. It is written simply. This section should include. As a result, this section should be improved.

� Before describing the goal, the authors must define the issue in a single line and explain why they chose this approach to study this review.

� Some scored data should be added

� In the final line of the abstract, the authors should present a decisive conclusion derived from the research and provide a single line of future prospects.

Keywords

� The content of keywords did not reflect the content of this manuscript and the words used for forming the title should not be used as the keywords. So, the structure of keywords should be changed.

Introduction

� This section is too long and should be shortened

� The authors should give some lines about the knowledge gap which their reviews have covered along with the hypothesis statement

� Also, the authors should provide a novelty statement at the end. What new things authors have done or correlated in this research compared to old ones?

� The general and specific aim should be specified

Materials and Methods

� All measurements should be supported by the references

� All abbreviations should be written in full name

Results and discussion

� In general, the figures are not presented clearly.

� The admixture status of population structure should be mentioned

� The authors should mention the scored data when they explain the maximum and minimum of the studied parameters.

� All captions should be improved, showing the contents of tables and figures

� A PCA plot should be detailed to better understand

� The discussion is weak. The authors should interpret all results obtained in this study by adding some information about the results obtained in their study. The authors should explain how all of the findings from this study relate to their own findings.

Conclusion

� The authors should summarize the most significant findings because they have written this section in an easy-to-read manner.

� Future works about this research should also include additional works

Reviewer #2: Population genetic diversity and structure of the endangered species Tetracentron

sinense Oliver (Tetracentraceae) with SNPs based on RAD sequencing.

Abstract

Line 25 to 28- The sentence is long and confusing. Break into two shorter clearer sentences.

Line 32 to 37 – Edit to remove numbers (1), (2) and (3).

Enrich the results section by adding in information on the additional results from the recommended analysis.

Line 38- Delete the word ‘Finally’.

Line 40 – Break the sentence into two so that the first sentence ends at geographic groups. Delete ‘and that'. Start new sentence with ‘Attention’.

Align the conclusion with the additional recommended analyses.

Introduction

A relevant introduction has been provided however the grammar needs to be improved. Some of the changes to be made are listed below:

Line 48 delete the word ‘very’

Line 51 delete the words 'endangered species particularly’. Replace ‘cause’ with causes.

Line 52 replace the word with ‘affect’ with affects.

Line 56 – delete the word ‘great’.

Line 57 delete ‘a’ before small

Line 59 italicise “Tetracentron”

Line 62 edit sentence to remove the abbreviation e.g.

Lien 67 add the words ‘have been done’ after techniques

Line 83 to 87 The sentence is long and confusing. Break into two shorter clearer sentences.

Line 96 edit last part of the sentence to read ‘low cost with high reliability’

Materials and methods

Appropriate methods were used. The section needs to be improved as detailed below.

In the methodology section, line 107-108 the statement “Samples of spaced about 50m apart were randomly selected…” does not make grammatical sense. What did the authors mean?

Line 131: It would be good to declare the type of alignment done after running the process_radtags programme. The authors are leaving the readers to assume that a de novo assembly was done based on the flow from ustacks to cstacks and sstacks.

Although its mentioned in the methodology as a filtering parameter, the proportion of SNPs with a minor allele frequency (MAF) ≥ 0.05 is not declared in the results. There is need to declare the proportion of these SNPs before and after filtering.

Population stratification has been explored. However, more analysis should be done to implore the spatial population structure using TESS (refer to Front Genet 2019 10:954, Fig. 1).

Additional aspects of genetic diversity should be explored. Some important analysis such as linkage disequilibrium (LD) patterns and samples’ co-ancestry have not been done. The authors are encouraged to explore this option since it would provide information on the historical recombinations that could have occurred within samples as well as help in understanding the observed population structure.

The authors are also encouraged to explore using OutFLANK and PCAdapt approaches to identify the putative adaptive loci as well as the outlier SNPs.

Editing to remove grammatical errors to be done in whole section.

Results

Commendable effort has been made to report the results. The manuscript can be improved as follows.

It is recommended that the authors compute and include in the results section other genetic diversity indices such as the Hardy-Weinberg Equilibrium (HWE) test scores for the different T. sinense populations under study.

There are certain important summary statistics that could be included in the analysis such as the Watterson’s theta which would also help in understanding the TajimaD statistics.

Important population parameters such as site frequency spectra (SFS) to show the distribution of alleles in the different populations of T. sinense also need to be provided. This is important because it groups the SNPs into classes based on how many copies of the minor allele they possess and then calculates the size of each group. As such, this will provide the readers with information on the distribution of alleles in the different populations under study.

Results from the additional analyses should be inserted in the appropriate sections of the results.

Editing to remove grammatical errors will improve the section.

Discussion and conclusions

The authors have tried to discuss the results described in the manuscript. The discussion can be further improved.

Align the discussion and conclusions with the results from the recommended additional analysis.

The sections has too many grammar errors to enumerate. This needs to be diligently addressed.

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Reviewer #1: No

Reviewer #2: Yes:  Nancy L. M. Budambula and Joshua K. Muli . We are both from University of Embu, Kenya

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Attachment

Submitted filename: PLOS-review.docx

PONE-D-23-17864R1Population genetic diversity and structure of the endangered species  Tetracentron sinense  Oliver ( Tetracentraceae ) with SNPs based on RAD sequencingPLOS ONE

Dear Dr. Gan,

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 Jun 29 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,

Miquel Vall-llosera Camps

Senior Staff 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)

Reviewer #3: (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

Reviewer #3: Yes

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

Reviewer #1: N/A

Reviewer #2: Yes

Reviewer #3: 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: Yes

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 #1: No

Reviewer #2: Yes

Reviewer #3: No

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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: Dear authors

The following modifications are required:

Abstract

� Summarizing of all measured data (for example Fst) are not mentioned in this section

� All abbreviation should be written in full name

� Why did not the authors calculate the gene flow?

� Why did not the authors calculate the polymorphism percentage and private or unique band for each population?

Keywords

� The keywords did not accurately reflect the substance of this book, and the words used to make the title should not be utilized as keywords. As a result, the keyword structure needs to be changed

Introduction

� No information about the method of the study of genetic diversity is available

� Some information about the methods of the study of genetic diversity should be included

� Some information about the conservation of this plant species should be provided

� Along with the hypothesis statement, the authors should include a few paragraphs regarding the knowledge gap that their research addressed.

� Additionally, the authors should include a novelty statement at the end. What new things have the authors done or correlated in this study compared to previous ones?

� The general and specific aim should be inserted

Materials and Methods

� The number of samples should be greater than seven. Less than eight samples are not acceptable

� The voucher number should be added for all samples in all populations in Table 1

� L109: The distance between plants is too less

� The procedure of PCR should be added in detail

� The manufacture of all materials should be added

� Why did not the authors calculate the PIC?

� The procedure of population structure should be detailed by adding the information about the burn-in period and Markov chain Monte Carlo (MCMC) iterations

Results and discussion

� All abbreviations in the tables should be defined

� How did the authors determine the number of clusters? What was the criteria points?

� All captions should be detailed

� The figure of delta K should be inserted

� The number of clusters formed by AHC or PCA should be defined and labelled

� A mantel test should be measured among molecular data

� The location of quarters on the PCA plot should be defined

� The results of PCA are not presented in detail.

� There is little discussion. By providing some context for the findings of their own study, the authors should be able to interpret all of the conclusions of their investigation. It is important to understand properly the relationships between the many populations that are examined using multivariate methodologies and diversity indexes. What significance, for instance, does this study place on the examination of PIC, Fst, and other genetic diversity parameters? The authors should compare the results of clustering and AMOVA

Conclusion

� Additional studies on this topic should be included.

� The authors should describe the most noteworthy findings, as they have written this section in an easy-to-read format.

Reviewer #2: Address the following before release of the final draft.

1. Methodology: Line 128, The authors should declare explicitly that they did a de novo assembly for the benefit of the readers who are not familiar with the flow of events in stacks software. This had been raised earlier

2. Using the GPS coordinates of the sampling sites together with the SNP data, the authors could explore ancestry estimation for spatial population genetic analysis using TESS as earlier suggested. This will enable the evaluation on whether T. sinense populations in central and south China have distinct or mixed origins and enrich the ADMIXTURE results. The generated map(s) can then be added to the results.

3. Results: It is commendable that the authors explored the linkage disequilibrium (LD) pattern of the SNP markers in the population. However, an LD scatter plot would be more informative to show the decay rate as explained in lines 251-257. I suggest replacing Table 6 with an LD scatter plot.

4. The grammar and writing style also improved. However, the final draft will need proof reading and we recommend that the authors get an English-speaking colleague to check for grammar and language use.

Reviewer #3: Dear editor,

Thank you so much for sending this paper for me to review. I think the paper needs more corrections before the final acceptance and publishing.Here are my comments:

- The introduction is too long.

- The aims of this study must be explained clearly.

- I request authors to explain why the individual number differed among the studied populations?

- The language of paper must be corrected, specially in material and method section.

-The numbers after the decimal point must be reduced into 2 numbers.

- I could not find the gene flow rate. If the parameter hasn't be calculated I request to calculate it.

All the best

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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 #1: Yes:  Nawroz Tahir

Reviewer #2: Yes:  Nancy L.M. Budambula and Joshua K. Muli

Reviewer #3: No

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Attachments

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Submitted filename: Revision Comments.docx

PONE-D-23-17864R2Population genetic diversity and structure of the endangered species  Tetracentron sinense  Oliver ( Tetracentraceae ) with SNPs based on RAD sequencingPLOS ONE

Dear Dr. Gan,

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.

Reviewers have recognized a significant improve in the clarity and quality of the manuscript. However, They are still concerned  about some issues which need to be addressed.

Please submit your revised manuscript by Oct 31 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'.
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Vicente Martínez López

Academic Editor

PLOS ONE

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Reviewer #2: (No Response)

Reviewer #3: (No Response)

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Reviewer #2: Yes

Reviewer #3: Yes

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Reviewer #2: Yes

Reviewer #3: Yes

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Reviewer #2: Yes

Reviewer #3: Yes

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Reviewer #2: Yes

Reviewer #3: Yes

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Reviewer #2: The comments below which were raised on Revision 1 in March 2024 have not been addressed. Kindly the authors address them.

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Dear Editor March 2024

Overall, the authors have made good effort to address the concerns we raised on the previous manuscript draft. I draw your attention to the following:

1. Methodology: Line 128, The authors should declare explicitly that they did a de novo assembly for the benefit of the readers who are not familiar with the flow of events in stacks software. This had been raised earlier

2. Using the GPS coordinates of the sampling sites together with the SNP data, the authors could explore ancestry estimation for spatial population genetic analysis using TESS as earlier suggested. This will enable the evaluation on whether T. sinense populations in central and south China have distinct or mixed origins and enrich the ADMIXTURE results. The generated map(s) can then be added to the results.

3. Results: It is commendable that the authors explored the linkage disequilibrium (LD) pattern of the SNP markers in the population. However, an LD scatter plot would be more informative to show the decay rate as explained in lines 251-257. I suggest replacing Table 6 with an LD scatter plot.

4. The grammar and writing style also improved. However, the final draft will need proof reading and we recommend that the authors get an English-speaking colleague to check for grammar and language use.

Reviewer #3: Dear editor,

Thank you so much for sending the paper fpr me to review. I checked the paper and found it needs some minor corrections before the final acceptance. Here are my comments which helps to improve the paper quality.

-Authors must clearly explain the aims of their study.

-The gene flow amount (Nm) must be calculated among the populations.

-The species and genera names must be in italic form.

All the best

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Reviewer #2: Yes:  Nancy L.M. Budambula and Joshua Kiilu Muli

Reviewer #3: Yes:  Seyed Mehdi Talebi

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Attachments

Attachment

Submitted filename: Revision Comments.docx

Population genetic diversity and structure of the endangered species  Tetracentron sinense  Oliver ( Tetracentraceae ) with SNPs based on RAD sequencing

PONE-D-23-17864R3

Dear Dr. Gan,

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,

Vicente Martínez López

Academic Editor

PLOS ONE

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