Mitochondrial D-loop sequence variation and maternal lineage in the endangered Cleveland Bay horse

Andy C. Dell; Mark C. Curry; Kelly M. Yarnell; Gareth R. Starbuck; Philippe B. Wilson

doi:10.1371/journal.pone.0243247

Peer Review History

Original SubmissionAugust 18, 2020
13 Oct 2020 Decision Letter - Chris Rogers, Editor PONE-D-20-24140 Mitochondrial D-loop sequence variation and maternal lineage in the endangered Cleveland Bay horse PLOS ONE Dear Dr. Wilson, 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. Both reviewers found it to be sound and well written. Some of the material is very horse and breed specific and this was the reason for differing opinions between the reviewers. I do have to agree with reviewer two that it would have been better to have combined your two papers on this study to provide one very robust manuscript. Please look at the nomenclature suggestion provided by reviewer one and provide a revised manuscript. Please submit your revised manuscript by Nov 27 2020 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: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols We look forward to receiving your revised manuscript. Kind regards, Chris Rogers Academic Editor PLOS ONE Additional Editor Comments: Thank you for your submission. Both reviewers found it to be sound and well written. Some of the material is very horse and breed specific and this was the reason for differing opinions between the reviewers. I do have to agree with reviewer two that it would have been better to have combined your two papers on this study to provide one very robust manuscript. Please look at the nomenclature suggestion provided by reviewer one and provide a revised manuscript. Journal Requirements: When submitting your revision, we need you to address these additional requirements. 1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf 2. Thank you for stating the following in the Acknowledgments Section of your manuscript: "University of Lincoln and The Rare Breeds Survival Trust" We note that you have provided funding information that is not currently declared in your Funding Statement. 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If you have no competing interests, please state "The authors have declared that no competing interests exist.", as detailed online in our guide for authors at http://journals.plos.org/plosone/s/submit-now This information should be included in your cover letter; we will change the online submission form on your behalf. Please know it is PLOS ONE policy for corresponding authors to declare, on behalf of all authors, all potential competing interests for the purposes of transparency. PLOS defines a competing interest as anything that interferes with, or could reasonably be perceived as interfering with, the full and objective presentation, peer review, editorial decision-making, or publication of research or non-research articles submitted to one of the journals. Competing interests can be financial or non-financial, professional, or personal. Competing interests can arise in relationship to an organization or another person. Please follow this link to our website for more details on competing interests: http://journals.plos.org/plosone/s/competing-interests 4. Your ethics statement should only appear in the Methods section of your manuscript. If your ethics statement is written in any section besides the Methods, please move it to the Methods section and delete it from any other section. Please ensure that your ethics statement is included in your manuscript, as the ethics statement entered into the online submission form will not be published alongside your manuscript. [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: Yes Reviewer #2: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: Yes ****** 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: Yes Reviewer #2: Yes ****** 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: Yes Reviewer #2: Yes ****** 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: This paper presents analysis of mtDNA typing of the Cleveland Bay horse, one of the world's most endangered horse breeds. The paper is well written and appropriate methods are used. I have only a couple of comments. One, the mtDNA haplotype nomenclature of Jansen has largely been replaced by that of Achilli et al. 2012. Using Jansen is ok but Achilli should also be used concurrently. The diversity of mtDNA in the CB is consistent with the low nuclear DNA variation that has recently been reported and the high inbreeding level within the breed. It is interesting how well the maternal lines predicted by pedigree analysis are aligned with the mtDNA haplotypes. I also am somewhat surprised by how high the proportion of haplotypes representing horses from the British Isles is. Within breed ancestry representations are often quite diverse representing past intermixing of horses prior to breed formation. I also will say that the diversity shown does not indicate that the Chapman horse is not the direct link to all parts of the CB ancestry as there is no knowledge of the make-up of that horse. My one negative comment is not actually a criticism of the paper. This paper has a great deal of very specific information about the make-up of the individuals that comprise the breed. This has almost no meaning to most potential readers but is valuable information for the specialist. The great detail of the specific maternal lines and groups within the breed make certain sections of this paper very difficult to follow. In fact the paper presents little that is new except in reference to the breed. The paper might be more appropriate for a different journal although I don't have a specific recommendation for one. Reviewer #2: The current study aimed to establish maternal lineages within the Cleveland Bay Horse breed and detect relationships with other domestic horse breeds. The Cleveland Bay Horse is a very important breed and known to have low genetic diversity. The authors used typical methods for the data analysis of part of the D-loop. I think sequencing the full D-loop would be more informative especially with the high level of similarity seen among tested samples. Although the results and discussion were well written, the approach and results are lacking novelty, very specific to horses and not applicable for novel discoveries in other species. The overall outcomes of the current study might be interesting and useful for a narrow group of readers. More importantly, I wish the authors have included the finding of this study with their previous paper. The story could be then much informative by integrating the autosomal genetics findings in the light of the mtDNA sequencing instead of having them separated in two manuscripts. ****** 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. 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: 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. https://doi.org/10.1371/journal.pone.0243247.r001
Revision 1
30 Oct 2020 Author Response Reviewer #1: This paper presents analysis of mtDNA typing of the Cleveland Bay horse, one of the world's most endangered horse breeds. The paper is well written and appropriate methods are used. I have only a couple of comments. One, the mtDNA haplotype nomenclature of Jansen has largely been replaced by that of Achilli et al. 2012. Using Jansen is ok but Achilli should also be used concurrently. The nomenclature from Achilli has been incorporated within Table 2. The diversity of mtDNA in the CB is consistent with the low nuclear DNA variation that has recently been reported and the high inbreeding level within the breed. It is interesting how well the maternal lines predicted by pedigree analysis are aligned with the mtDNA haplotypes. I also am somewhat surprised by how high the proportion of haplotypes representing horses from the British Isles is. Within breed ancestry representations are often quite diverse representing past intermixing of horses prior to breed formation. I also will say that the diversity shown does not indicate that the Chapman horse is not the direct link to all parts of the CB ancestry as there is no knowledge of the make-up of that horse. My one negative comment is not actually a criticism of the paper. This paper has a great deal of very specific information about the make-up of the individuals that comprise the breed. This has almost no meaning to most potential readers but is valuable information for the specialist. The great detail of the specific maternal lines and groups within the breed make certain sections of this paper very difficult to follow. In fact the paper presents little that is new except in reference to the breed. The paper might be more appropriate for a different journal although I don't have a specific recommendation for one. Reviewer #2: The current study aimed to establish maternal lineages within the Cleveland Bay Horse breed and detect relationships with other domestic horse breeds. The Cleveland Bay Horse is a very important breed and known to have low genetic diversity. The authors used typical methods for the data analysis of part of the D-loop. I think sequencing the full D-loop would be more informative especially with the high level of similarity seen among tested samples. Although the results and discussion were well written, the approach and results are lacking novelty, very specific to horses and not applicable for novel discoveries in other species. The overall outcomes of the current study might be interesting and useful for a narrow group of readers. More importantly, I wish the authors have included the finding of this study with their previous paper. The story could be then much informative by integrating the autosomal genetics findings in the light of the mtDNA sequencing instead of having them separated in two manuscripts. We thank the reviewers for their comments, and their careful assessment of the manuscript. https://doi.org/10.1371/journal.pone.0243247.r002
10 Nov 2020 Decision Letter - Chris Rogers, Editor PONE-D-20-24140R1 Mitochondrial D-loop sequence variation and maternal lineage in the endangered Cleveland Bay horse PLOS ONE Dear Dr. Wilson, 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 look at the revised manuscript in the results section. In the copy uploaded there are a number of occurrences of the following Error! Reference source not found Please submit your revised manuscript by Dec 25 2020 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: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols We look forward to receiving your revised manuscript. Kind regards, Chris Rogers Academic Editor PLOS ONE Additional Editor Comments (if provided): Please look at the revised manuscript in the results section. In the copy uploaded there are a number of occurrences of the following Error! Reference source not found Please correct and upload a new version. [Note: HTML markup is below. Please do not edit.] [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. https://doi.org/10.1371/journal.pone.0243247.r003
Revision 2
10 Nov 2020 Author Response Dear Colleagues, Further to our recent submission, we have modified the main manuscript to refer to our first paper concerning the genetic analysis of the Cleveland Bay horse population through pedigree evaluations and molecular methods. Furthermore, we have reiterated the tutorial-based nature of our initial paper, which guides the reader through our framework and proposes its implementation in the decision-making process for conservation of endangered breeds and species. Whilst we recognise the points of the Editorial Office in terms of segmenting research projects, the authors would like to firmly stress that these pieces of work are not only standalone, but represent logical and careful developments of the narrative as part of the large project we continue to undertake in conserving the Cleveland Bay horse and the native equine breeds of the United Kingdom. Whilst we use these breeds as examples herein, our approaches are careful to be extendable not just to equines and livestock, but in wider conservation practices. Furthermore, according to the email dated 10th November 2020, we have amended the manuscript to remove the “Error! Reference source not found” therein. Updates to statements based on the requests from Editorial Office: 1) Thank you for updating your data availability statement. You note that your data are available within the Supporting Information files, but no such files have been included with your submission. At this time we ask that you please upload your minimal data set as a Supporting Information file, or to a public repository such as Figshare or Dryad. Please also ensure that when you upload your file you include separate captions for your supplementary files at the end of your manuscript. As soon as you confirm the location of the data underlying your findings, we will be able to proceed with the review of your submission. Authors: These data have been provided: 10.6084/m9.figshare.13154429. The statements have also been amended. 2) Your ethics statement should only appear in the Methods section of your manuscript. If your ethics statement is written in any section besides the Methods, please move it to the Methods section and delete it from any other section. Please ensure that your ethics statement is included in your manuscript, as the ethics statement entered into the online submission form will not be published alongside your manuscript. Authors: This has been moved. 3) It is important that you include a cover letter with your manuscript. Please ensure that this letter is addressed specifically to PLoS ONE. Please also include * why this manuscript is suitable for publication in PLoS ONE. * how does your paper provide a worthwhile addition to the scientific literature? * how does your paper relate to previously published work? * which types of scientists do you believe will be most interested in your study? Authors: This serves as the document. 4) Thank you for stating the following financial disclosure: NO At this time, please address the following queries: a) Please clarify the sources of funding (financial or material support) for your study. List the grants or organizations that supported your study, including funding received from your institution. b) State what role the funders took in the study. If the funders had no role in your study, please state: “The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.” c) If any authors received a salary from any of your funders, please state which authors and which funders. d) If you did not receive any funding for this study, please state: “The authors received no specific funding for this work.” Authors: The authors received no specific funding for this work. Please include your amended statements within your cover letter; we will change the online submission form on your behalf. 5) Thank you for stating the following in your Competing Interests section: "No" Please complete your Competing Interests on the online submission form to state any Competing Interests. If you have no competing interests, please state "The authors have declared that no competing interests exist.", as detailed online in our guide for authors at https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Fjournals.plos.org%2Fplosone%2Fs%2Fsubmit-now&data=04%7C01%7Cphilippe.wilson%40ntu.ac.uk%7Cb7c2ef235df54ce631fe08d87b5be85f%7C8acbc2c5c8ed42c78169ba438a0dbe2f%7C0%7C0%7C637394981962863769%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C1000&sdata=ex8IGrShDq%2B7e39fAIsGyALzEK6Z7TDaR0HTS5u2E4c%3D&reserved=0 This information should be included in your cover letter; we will change the online submission form on your behalf. Authors: The authors have declared that no competing interests exist Original letter: Herein, we consider the genetic diversity within the Cleveland Bay horse population, one of the oldest and most foundationally integral equine breeds. Indeed, until now, there was little convincing evidence concerning the loss of genetic variation within the breed, across the generations, leading to a status as endangered as that today. We therefore approach the problem of rigorously developing breed management plans for a critically endangered population such as the Cleveland Bay horse by first considering the genealogical and molecular data pertaining to the generational genetic history of the breed. The Cleveland Bay horse is a historic breed of horse native to the UK which has been widely used throughout the equine world for generations. However, since the move to mechanization, its uses on land, in the field became limited and the population threatened. Now critically endangered, the breed must be subject to strict management based on genetic factors in order to increase diversity therein and safeguard the Cleveland Bay for future generations. Herein, we demonstrate that only four original female lines are represented in the current population, and consider the evolution of the breed from the now-extinct Chapman Horse. We present the methodology as a powerful tool for conservation, both livestock and in applications to wild species in situ. Furthermore, we dispel previous assumptions regarding the lineage and ancestry of the modern Cleveland Bay population. Whilst being evidently pertinent to livestock and native breed conservation biology, our approach is inherently translatable to in situ conservation of species more globally. Indeed, our follow-up paper on this work, develops this study and describes 16 years of breed management programmes, and their effect on the genetic diversity within the population; work which could not have taken place without the robust and necessary first principles established within the present study. In addition to a robustly-researched and thoroughly described methodology, we present the logical development of the work and fundamental theory throughout in order for this work to be recognised as a valuable standalone resource for biological conservationists. We suggest the following colleagues as reviewers for this manuscript: Prof Stephen Hall: stHall@lincoln.ac.uk Prof Tim Morris: Tim.Morris@nottingham.ac.uk Dr Tim Bray: tbray@bristolzoo.org.uk We hope you will consider this manuscript with the attention it deserves, and look forward to receiving your feedback. With kind regards, Prof Philippe B. Wilson MChem(Hons) PhD PGCertHE MRSC FRSB FLS FHEA Attachments Attachment Submitted filename: R1ResponseToReviewersmtDNACBH.docx https://doi.org/10.1371/journal.pone.0243247.r004
18 Nov 2020 Decision Letter - Chris Rogers, Editor Mitochondrial D-loop sequence variation and maternal lineage in the endangered Cleveland Bay horse PONE-D-20-24140R2 Dear Dr. Wilson, 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. An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org. If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. Kind regards, Chris Rogers Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0243247.r005
Formally Accepted
20 Nov 2020 Acceptance Letter - Chris Rogers, Editor PONE-D-20-24140R2 Mitochondrial D-loop sequence variation and maternal lineage in the endangered Cleveland Bay horse Dear Dr. Wilson: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Chris Rogers Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0243247.r006

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