Trait variation in patchy landscapes: Morphology of spotted salamanders (Ambystoma maculatum) varies more within ponds than between ponds

Elizabeth T. Green; Anthony I. Dell; John A. Crawford; Elizabeth G. Biro; David R. Daversa

doi:10.1371/journal.pone.0299101

Peer Review History

Original SubmissionNovember 3, 2023
18 Dec 2023 Decision Letter - Janice L. Bossart, Editor PONE-D-23-36372Trait variation in patchy landscapes: morphology of spotted salamanders (Ambystoma maculatum) varies more within ponds than between pondsPLOS ONE Dear Dr. Daversa, 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 Feb 01 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. 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Thank you for stating the following in the Acknowledgments Section of your manuscript: "We thank members of the Tyson Research Station for their support of our field sampling, J. Grady for assistance with data analyses and visualization, J. Messier for assistance with the statistical analyses, and S. Trombulak for assistance with data analyses and comments that significantly improved this manuscript. This project was conducted in accordance with University of Illinois IACUC #16203 and funded, in part, by the NGRREC intern program. None of the authors experienced conflict of interest that could have influenced the objectivity of this study" 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. 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Please include captions for your Supporting Information files at the end of your manuscript, and update any in-text citations to match accordingly. Please see our Supporting Information guidelines for more information: http://journals.plos.org/plosone/s/supporting-information. 9. 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. Additional Editor Comments: Your manuscript was favorably received by both reviewers, with both indicating only minor revisions were necessary. Please carefully consider two significant points they raised regarding analysis and interpretation of between-pond variation: Consider whether you can use date as a covariate in your models as an effective way to control for temporal effects, which might then allow you to explicitly explore/include the role of environmental variables. Another important consideration is whether spatial autocorrelation exists among ponds or not, and how that might relate to factors contributing to between-pond variation. Some additional minor problems I noted when reading through your manuscript. Lines 62-63. This is very vertebrate focused text, given the preceding sentence is inclusive of all species, and generally not reflective of what might drive within and between patch variation in, e.g. herbivorous insects, where major drivers are the specific array of host species present and the relative quality and abundance of each. Line 297-298. Wording issue. Word(s) missing? Line 329. 'driving to' should be 'driving the' Line 336-338. Wording issue. Do you mean '...remains as to the specific...' ? Line 377. Change 'reasoning' to either 'reasons' or 'causes' [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: No ****** 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: The authors present the results of an investigation of morphological variation among salamanders in six ponds that is motivated by 1) the recognition that individual variation can have important ecological implications and, in a more applied context, 2) a question about whether artificial ponds that might be used in restoration might create trait variation or change the scaling of traits (i.e. allometric relationships). The study is limited to the questions addressed because, as the authors explain, geography and time were confounded such that explorations of mechanisms could not be attempted. Thus, the study is a detailed exploration of trait variance that partitions within- and between - pond effects for trait variation and allometric variation. Given the potential importance of individual variation, this project provides some insight into how trait variation is organized in these salamanders. The authors repeatedly refer to this as preliminary. I object to this language because it has negative connotations that perhaps highlight the limitations of the sampling. Perhaps that conveys the authors' assessment, but it distracts from the main point as written which is the description of individual variation. With respect to the limitations, would it be possible to simply use day of the year as a covariate to control for sampling date differences? This would assume that life history events are synchronized across the six ponds, but would facilitate a deeper exploration of environmental variables. minor comments: line 60: change "comprised of" to "comprising" lines 66-67, 76, 78, 126, 144, 179, 184: references here don't conform to the rest of the ms formatting lines 93-95: I wonder if the context for the study would be more clear if this question included the idea that the artificial ponds differing in age. That is one of the interesting results. lines 186-190: One could also use MacArthur's broken stick approach, or a similar inertia method, to assess how many PC axes are sufficient. line 204: I'm not sure why Figure 2 is referenced for this sentence about the data being normally distributed. line 211: I think this needs more explanation. How does this multiplication 1) facilitate convergence and, 2) "standardize the units with length measurements"? For example, why not center and scale mass and length? line 329: change "diving to" to "driving the" Reviewer #2: I enjoyed reading this manuscript. It’s well-written, and the motivation was well justified and explained, as were the methods. I particularly appreciated the explanations given for modelling choices throughout the methods section. I have no major concerns, only a minor comment: It seems the authors were careful to minimize the possibility that growth differences and temporal sampling contributed to major between-pond variation. I wonder if the authors looked at spatial autocorrelation among the sample sites. From the results it doesn’t seem obvious that there was spatial autocorrelation or potential for isolation by distance patterns in trait variation (with one site being markedly further away). But perhaps a lack of autocorrelation would also be interesting and support the idea that trait variation is more related to the pond-specific properties of salamander populations and environmental factors. In any case whether spatial autocorrelation was explored, or why the authors chose not to, should be noted in the Methods. L234: predator density? Is this supposed to be here? Fig S3: the confidence intervals are either hard to see or missing ****** 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.0299101.r001
Revision 1
30 Jan 2024 Author Response Dear editors, reviewers and journal staff: Thank you for taking the time to review our manuscript and for the constructive feedback to improve the work. We have addressed the line edits in our revision, which you can see in the track revised manuscript with track changes. We have also revised the title page to match the PLOS ONE style requirements and removed the funding information from the Acknowledgements section. Our revised funding statement is below – “This study was funded in part by the National Great Rivers Research and Education Center Grant/Award Number: NGRREC-IP2016-5” We have also published our raw data and R scripts to the Figshare Data Repository. We have embargoed the raw data, and we will release the embargo upon acceptance of the manuscript. The R script for the data analyses and figures is available at: https://figshare.com/articles/dataset/Amac_Morphology_R_scripts/24996476 DOI: https://doi.org/10.6084/m9.figshare.24996476.v1 We provide responses to each of the review comments below: Additional Editor Comments: Your manuscript was favorably received by both reviewers, with both indicating only minor revisions were necessary. Please carefully consider two significant points they raised regarding analysis and interpretation of between-pond variation: Consider whether you can use date as a covariate in your models as an effective way to control for temporal effects, which might then allow you to explicitly explore/include the role of environmental variables. Another important consideration is whether spatial autocorrelation exists among ponds or not, and how that might relate to factors contributing to between-pond variation. Response: Thanks for taking the time to consider our manuscript. We have considered the two main comments pertaining to (a) the addition of day-of-year as a covariate in our statistical models and (b) the possibility of autocorrelation. Adding the day-of-year covariate did not affect model performance because the factor varied perfectly with our random effect – the ID of ponds. This is due to our sampling design, where we surveyed one pond/week, and a different pond each week; pond ID changed perfectly in tandem with day-of-year. In terms of spatial autocorrelation, we decided against a formal analysis because the results would have been unreliable with our small sample size of six ponds. Judging from the patterns of trait partitioning among ponds, we are doubtful that spatial autocorrelation was a factor. However, we do not have the breadth of data and sampling locations to reliably test this hypothesis, and we now state so in the manuscript (Line 244). Below, we elaborate on how we came to these decisions. Although in the end no changes to the analyses were made, we assure you that ample time was given to consider the options. Some additional minor problems I noted when reading through your manuscript. Lines 62-63. This is very vertebrate focused text, given the preceding sentence is inclusive of all species, and generally not reflective of what might drive within and between patch variation in, e.g. herbivorous insects, where major drivers are the specific array of host species present and the relative quality and abundance of each. Response: This is a good point. We have clarified the scope of this statement by adding ‘in vertebrates’ after ‘some notable examples...’ (Line 76). We considered adding examples pertaining to invertabrates, but because our study focuses on a vertebrate species, we decided for brevity to stick to vertebrate examples. Line 297-298. Wording issue. Word(s) missing? Response: Yes, the sentence was missing the word ‘in’ after ‘involved. We have corrected this error: “Spatial connectivity may also have been involved in the observed partitioning of morphological variation” (Line 366). Line 329. 'driving to' should be 'driving the' Response: Changed (Line 403) Line 336-338. Wording issue. Do you mean '...remains as to the specific...' ? Response: Yes, and changed (Line 412) Line 377. Change 'reasoning' to either 'reasons' or 'causes' Response: we changed to ‘reasons’ (Line 462) Reviewer #1: “The authors present the results of an investigation of morphological variation among salamanders in six ponds that is motivated by 1) the recognition that individual variation can have important ecological implications and, in a more applied context, 2) a question about whether artificial ponds that might be used in restoration might create trait variation or change the scaling of traits (i.e. allometric relationships). The study is limited to the questions addressed because, as the authors explain, geography and time were confounded such that explorations of mechanisms could not be attempted. Thus, the study is a detailed exploration of trait variance that partitions within- and between - pond effects for trait variation and allometric variation. Given the potential importance of individual variation, this project provides some insight into how trait variation is organized in these salamanders.” Response: We are glad to hear that the reviewer finds our project important despite the noted limitations. “The authors repeatedly refer to this as preliminary. I object to this language because it has negative connotations that perhaps highlight the limitations of the sampling. Perhaps that conveys the authors' assessment, but it distracts from the main point as written which is the description of individual variation.” Response: We referred to the analysis between “new” and “old” ponds as preliminary because of the low replication of ponds (N=6), however, we understand your objection and the negative connotation. To discuss our analysis, without overselling the statistical power, we have removed the “preliminary” language from the methods and results and instead describe the limitations in the discussion section, which we feel is appropriate and warranted. “With respect to the limitations, would it be possible to simply use day of the year as a covariate to control for sampling date differences? This would assume that life history events are synchronized across the six ponds, but would facilitate a deeper exploration of environmental variables.” Response: We could not include day of year as a covariate because of its collinearity with the pond IDs. We sampled one pond/week, and a different pond each week. With this design, the day of year varies equally with the pond identification where we sampled. Because our models already include ‘pond ID’ as a random effect, adding day of year would not add further explanatory power. Just as a check of this, we went back to a subset of our models and added the day of year as a covariate. The result was that the day of year variable was automatically dropped due to singularity, meaning essentially that the two variables vary equally. minor comments: lines 66-67, 76, 78, 126, 144, 179, 184: references here don't conform to the rest of the ms formatting Response: We updated the references so that they now follow the numbered format used by this journal. Thanks for pointing this out. lines 186-190: One could also use MacArthur's broken stick approach, or a similar inertia method, to assess how many PC axes are sufficient. Response: In response to your suggestion, we ran a broken stick model on the decomposed principal component values. For the whole body shape the cut off was found to be the third component, while for the head, body, and tail we confirmed the cutoff at the second component. Since the cutoff was found to be the third component in only one of the shape metrics, we kept our previously decided cutoff at the second PC axis to allow for comparison between shapes. line 211: I think this needs more explanation. How does this multiplication 1) facilitate convergence and, 2) "standardize the units with length measurements"? For example, why not center and scale mass and length? Response: We had originally explored multiplication of the response variables as a technique for getting the mixed effects models to converge. However, we were able to reach convergence without taking this measure, and instead we log transformed both the response and explanatory variables. We agree that scaling and centering mass and length measurements is good practice for running mixed effects models. We repeated the analyses using the scaled variables, and the results did not qualitatively change. We offer both versions of the models in the R script for data analysis: https://doi.org/10.6084/m9.figshare.24996476.v2. We have deleted the sentence of concern and now specify that we scaled and log-transformed the variables. Reviewer #2: “I enjoyed reading this manuscript. It’s well-written, and the motivation was well justified and explained, as were the methods. I particularly appreciated the explanations given for modelling choices throughout the methods section. I have no major concerns, only a minor comment:” Response: We are pleased that the reviewer enjoyed the manuscript and found our motivations justified and modelling choices well explained. “It seems the authors were careful to minimize the possibility that growth differences and temporal sampling contributed to major between-pond variation. I wonder if the authors looked at spatial autocorrelation among the sample sites. From the results it doesn’t seem obvious that there was spatial autocorrelation or potential for isolation by distance patterns in trait variation (with one site being markedly further away). But perhaps a lack of autocorrelation would also be interesting and support the idea that trait variation is more related to the pond-specific properties of salamander populations and environmental factors. In any case whether spatial autocorrelation was explored, or why the authors chose not to, should be noted in the Methods.” Response: We agree that an analysis of spatial autocorrelation would be interesting even if no trend were found. However, given the small sample size of ponds in this study, we were concerned that any result of a positive/negative trend, or lack of a trend, would not be reliable. Our study aims did not specifically center around spatial autocorrelation and so our sampling design did not generate data sufficient for such analyses. Further, sampling enough ponds to reliably assess spatial autocorrelation, while minimizing temporal effects on morphology, would not have been feasible. We think the question of spatial autocorrelation is valuable though and would be a nice extension to this study’s report on between-pond variation. We will consider this in future projects that build off of this work. We have added a statement regarding this to our methods (Line 212): “Our sample size did not permit reliable examination into the role of spatial autocorrelation in explaining between-pond variation”. Attachments Attachment Submitted filename: Response to Reviewers_DD.docx https://doi.org/10.1371/journal.pone.0299101.r002
6 Feb 2024 Decision Letter - Janice L. Bossart, Editor Trait variation in patchy landscapes: morphology of spotted salamanders (Ambystoma maculatum) varies more within ponds than between ponds PONE-D-23-36372R1 Dear Dr. Daversa, 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. Congratulations! 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, Dr. Janice L. Bossart Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0299101.r003
Formally Accepted
26 Mar 2024 Acceptance Letter - Janice L. Bossart, Editor PONE-D-23-36372R1 PLOS ONE Dear Dr. Daversa, I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now being handed over to our production team. At this stage, our production department will prepare your paper for publication. This includes ensuring the following: * All references, tables, and figures are properly cited * All relevant supporting information is included in the manuscript submission, * There are no issues that prevent the paper from being properly typeset If revisions are needed, the production department will contact you directly to resolve them. If no revisions are needed, you will receive an email when the publication date has been set. At this time, we do not offer pre-publication proofs to authors during production of the accepted work. Please keep in mind that we are working through a large volume of accepted articles, so please give us a few weeks to review your paper and let you know the next and final steps. Lastly, 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 customercare@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. Janice L. Bossart Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0299101.r004

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