PONE-D-24-28309Using DNA metabarcoding and direct behavioural observations to identify the diet of proboscis monkeys (Nasalis larvatus) in the Kinabatangan FloodplainPLOS ONE

Dear Dr. Boom,

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  few minor points raised during the review process.

Please submit your revised manuscript by Oct 07 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,

Wolfgang Blenau

Academic Editor

PLOS ONE

Journal requirements: 1. When submitting your revision, we need you to address these additional requirements. 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 financial disclosure:  [FNRS (Fonds de la Recherche Scientifique) Fonds Léopold III – pour l’Exploration et la Conservation de la Nature asbl. FNRS Gustave Boël-Sofina Fellowship].  Please state what role the funders took in the study.  If the funders had no role, please state: ""The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript."" If this statement is not correct you must amend it as needed. Please include this amended Role of Funder statement in your cover letter; we will change the online submission form on your behalf. 3. Thank you for stating the following in the Acknowledgments Section of your manuscript: [We thank the Sabah Biodiversity Centre and the Sabah Wildlife Department for their permission and support to this research. We also thank the Forest Research Centre (FRC) members, especially Dr. Joan Pererra, for the identification of many plant samples. We thank all the financial supporters: the FNRS (Fonds de la Recherche Scientifique), the Fonds Léopold III – pour l’Exploration et la Conservation de la Nature asbl., and the FNRS Gustave Boël-Sofina Fellowship 2016. We gratefully thank all research assistants and students at Danau Girang Field Centre (DGFC) for their precious help in the field and Dr Senthilvel KSS Nathan for his contribution. Special thank you to Oriana Bhasin for assisting us in the early-morning faeces sampling. We extend our gratitude to Laurent Grumiau, Esra Kaymak and Jérémy Migliore for their assistance during the laboratory work, as well as Latifa Karim for technical support for the MiSeq sequencing (GIGA-Genomics platform).]We note that you have provided funding information that is not currently declared in your Funding Statement. However, funding information should not appear in the Acknowledgments section or other areas of your manuscript. We will only publish funding information present in the Funding Statement section of the online submission form. Please remove any funding-related text from the manuscript and let us know how you would like to update your Funding Statement. Currently, your Funding Statement reads as follows:   [FNRS (Fonds de la Recherche Scientifique) Fonds Léopold III – pour l’Exploration et la Conservation de la Nature asbl. FNRS Gustave Boël-Sofina Fellowship]. Please include your amended statements within your cover letter; we will change the online submission form on your behalf. 4. Please provide a complete Data Availability Statement in the submission form, ensuring you include all necessary access information or a reason for why you are unable to make your data freely accessible. If your research concerns only data provided within your submission, please write "All data are in the manuscript and/or supporting information files" as your Data Availability Statement. 5. Please include your full ethics statement in the ‘Methods’ section of your manuscript file. In your statement, please include the full name of the IRB or ethics committee who approved or waived your study, as well as whether or not you obtained informed written or verbal consent. If consent was waived for your study, please include this information in your statement as well. 

6. 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.

[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: No

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: Using DNA metabarcoding and direct behavioural observations to identify the diet of proboscis monkeys (Nasalis larvatus) in the Kinabatangan Floodplain

Thiry et al.

reviewed by M. Clauss Zurich (does not do anonymous reviews)

This is a very good manuscript - a pleasure to read, and proof of a thorough and critical approach. How the different mtethods are compared, and the weaknesses of each outlined, is very clear and insightful. Thank you.

I have only minor comments - some references seem to have been mixed up (see attached word file). E.g., the Nijboer thesis 2006 is cited - but it consists either of individual chapters that should be cited seaprately, or of published papers that should be cited. Some citation numbering seems to be confused.

I also have a very few suggestions for grammar or spelling corrections in there.

I recommend to add 'seed' in the subheading 'correspondence with manual analyses' - see attached.

There are statements that fruit feeding differs by season, and for this, p-values are given but not the magnitude of the measurement. Note I think this paper is brlliant so this criticism is smartassing: it is common fashion in biology to value a p-value higher than an indication of magnitude, but it is the latter that is important biologically, the p-value is just thr prerogative that we can talk about the magnitude. So many people forget the magnitudes ... so if this is 5% fruit feeding vs. 6% fruit feeding, even if significant - a completely different interpretation than if it is 2% compared to 22% ... Please add that. Real biologists care about actual magnitudes, not about significance per se :-)

in l. 57 NIRS is mentioned - I thought this was a method to estimate nutrient content, not botanical composition - but I might be wrong. If you cite something on botanical composition here, forget my comment. If you cite this for nutrient content, I recommend to delete the NIRS from the list.

When reading for the first time, I was sceptical about the first sentence of the abstract, but the way this is not hyped in the main manuscript makes it ok, I think.

I could not find the GenBank entries, but that might have been my mistake. I recommend that for the final version, the access is ensured.

Thank you for having me review such a nice and concise manuscript!

sincerely. mclauss

Reviewer #2: I have also added comments in the manuscript.

Add one statement to indicate problem statement why Proboscis monkey and why in LKWS

The written manuscript is very good especially to involve endangered species. I would suggest to add more examples of research been done in Malaysia, or Asia regarding the diet intake using metabarcoding approach.

One section in introduction sound like or supposed to be in material and method section.

Add date/period of barcoding date base is generated

Add details on fecal samples, group information, etc in a Table

It would be more interesting to add details on metabarcoding data based on group, age and gender.

Revise the conclusion section

**********

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: Yes: Marcus Clauss

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.

Attachments

Attachment

Submitted filename: Manuscript_Proboscis_monkey_mc.docx

Attachment

Submitted filename: PONE-D-24-28309 reviewed.pdf

Using DNA metabarcoding and direct behavioural observations to identify the diet of proboscis monkeys (Nasalis larvatus) in the Kinabatangan Floodplain, Sabah

PONE-D-24-28309R1

Dear Dr. Boom,

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 will be generated when your article is formally accepted. Please note, if your institution has a publishing partnership with PLOS and your article meets the relevant criteria, all or part of your publication costs will be covered. Please make sure your user information is up-to-date by logging into Editorial Manager at Editorial Manager® and clicking the ‘Update My Information' link at the top of the page. If you have any questions relating to publication charges, 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,

Wolfgang Blenau

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments: