PONE-D-24-05394Isolation limits spring pollination in a UK fragmented landscapePLOS ONE

Dear Dr. Li,

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D. L was funded by the China Scholarship Council (grant no. 20186190011)

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Additional Editor Comments:

Dear Dr. Li,

After this first review round, we obtained two positive reviews and one negative one. I believe that if you are able to solve the issues raised by all reviewers, your manuscript will be suitable for publication in PLoS One.

Sincerely,

Daniel Silva

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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: Partly

Reviewer #3: Yes

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

Reviewer #1: Yes

Reviewer #2: No

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: Yes

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: Yes

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: In this study, the authors investigated the effect of habitat size and isolation on the plant-pollinator interaction. To do so, they conducted pollination experiments using potted plant phytometry, in which they placed English Bluebell. The work and experimental design attempt to separate the effect of habitat size and isolation on the plant-pollinator interaction. The authors conclude that the data suggest isolation of areas poses a greater threat to pollination than habitat size. It's a very elegant study with a great idea and well-written.

However, I have a few minor queries.

Although not explicitly stated, it appears that the authors placed the same number of English bluebell individuals in all patches, regardless of area size and isolation. First, I would like to request the authors to clarify whether the number of bluebell individuals was the same for all areas or not, in a clear manner. Second, if the number of plants was the same for all areas regardless of size and isolation, I have some doubts.

The authors correctly point out that larger areas should have a greater abundance of pollinators and that this could increase reproductive success in these areas. However, the data found show that the reproductive success of bluebells was not impacted by area size. The authors attribute this to insufficient pollinators, which could result in pollen limitation for the plants. I agree that the observed result is likely due to pollen limitation, but I have a different perspective on this. I wonder if areas of different sizes receiving the same number of bluebell individuals could affect these results? I say this because pollination is a process dependent on the density of pollinators (as the authors state in the text), but also on the density of conspecific plants. Larger areas should have a greater number of plant individuals, which would increase the chances of pollen transfer. I wonder if the experimental design masked this relationship. When I see the result that reproductive success was associated with proximity, I wonder if the closer areas are close enough to function like a larger area with more individuals?

Results

L 218-219 - How many individuals were flowering?

Reviewer #2: The authors evaluated the effect of the proximity and size of habitat patches on the reproductive success of a plant species largely pollinated by early spring bumblebee queens, as a phytometer plant.

The study has some limitations, which were carefully elucidated by the authors in the discussion, but which still represent an alternative for evaluating the effects of habitat fragmentation on pollination services in subsequent studies, considering a less local scale.

Below are some considerations about this study.

Line 49: of flowering plants rely on animal pollination

Line 85: In most cases

Line 101: foraging and nesting

Line 130: Do these flowering plant species share bee visitors with English bluebells?

Line 134: Could you improve the resolution quality of figure 1?

Line 168-170: During the experiment, 9 plants were placed in 51 patches, totaling 459 plants. Of these, only 130 plants were successful in survival and pollination, representing less than 30% of the number of plants used at the beginning of sampling.

Was there a total loss of plants in any of the sampling patches? If so, were these areas included or removed from subsequent analyses?

Line 212: The Kruskal-Wallis test is a non-parametric test used to compare the variation of a numerical variable with a categorical variable of at least 3 groups. According to the data presented by the authors, the numeric variables (i.e. the number of seeds, number of seed capsules, and number of flowers that failed to set seeds) were compared using two categories of plants: plants exposed to pollinators vs plants prevented from pollinators.

If this is the case, the analysis could be carried out using a more appropriate non-parametric test considering the nature of the data in the present study.

Line 226-230: See previous comment on the section "Data analysis"

Line 234: We found that both the number of seeds and seed capsules are positively related to the patch proximity. (Table 1, Fig. 4). However, no relationship was found between patch proximity and the size and quality of seeds

Reviewer #3: Overall, the authors have written a very clear and concise manuscript that provides interesting information on how habitat patch isolation impacts pollination within a relatively small study area in the spring. This information will help future studies that are looking to determine what factors are limiting pollination within various landscapes, including small fragmented habitats. Overall I don't have many comments and all my comments below are minor revisions.

Introduction

- It would be helpful to discuss the other factors outside of habitat fragmentation and isolation that might impact the different plant pollination metrics being measured, for example microclimatic variables such as temperature and light.

- Lines 107-108: Add references to what spring fragmentation studies have been completed to date and refer back to their results within the discussion.

Methods and materials

- It would be good to elaborate on how the plant surveys were conducted within the patches, for example how many transects/quadrats were used per area? Was number of flowering plants also measured or just species richness?

Collecting Seed Data

- Lines 187-188: Not clear how sub-samples were chosen, it would be good to include the number of subsamples analyzed per plant

Data Analysis

- The number of plant species was found to be correlated with area, but assessing whether the number of flowering plants in total was correlated with area would also be helpful. Was this data measured within the plant surveys? There may have been some patches will small amounts of area, but a relatively high number of flowering plants utilized by spring pollinators (for example flowering trees/shrubs) which may explain why area wasn’t significantly associated with pollination. If plant abundances are available, I would suggest investigating their correlations with area and possible the pollination metrics (e.g., seed set).

Results

- Line 221 – the p-value should be put within the brackets for this sentence

Discussion

- No line numbers

- A reference should be provided to prove that a significant positive relationship between total number of plant species and habitat area indicates that the woodland patches are a genuinely fragmentated system.

- It would be good to acknowledge that future studies could be improved by measuring pollinator visitation in addition to the metrics measured in this study so information on the relationships between pollinators and pollination could be further expanded on.

- The discussion should speak further to what other factors could reduce seed set and capsule development outside of reduced pollination, for example microclimatic variables, for example light and temperature

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Reviewer #1: No

Reviewer #2: No

Reviewer #3: No

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Attachments

Attachment

Submitted filename: Research artical - Li et. al Isolation.docx

Isolation limits spring pollination in a UK fragmented landscape

PONE-D-24-05394R1

Dear Dr. Li,

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.

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Kind regards,

Daniel de Paiva Silva, Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Dear Dr. Li,

I am pleased to accept you manuscript for publication in PLoS One!

Sincerely,

Daniel Silva

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: All comments have been addressed

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 #1: Yes

Reviewer #3: Yes

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

Reviewer #1: Yes

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 #1: 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: Yes

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 #1: (No Response)

Reviewer #3: The authors have addressed all my previous comments adequately and I have no further comments. Overall, this study provides interesting information on how habitat patch isolation impacts pollination within a relatively small study area in the spring. This information will help future studies that are looking to determine what factors are limiting pollination within different landscapes and provides helpful guidance on how future studies can improve upon the methods used within this study.

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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 #3: No

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