Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: RAPID RELATIVE INCREASE OF CRUSTOSE CORALLINE ALGAE FOLLOWING HERBIVORE EXCLUSION IN A REEF OF EL SALVADOR
Review round: 1
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: The replication may be an issue and more detail on replication and how that relates to figure 2 is needed.
Validity of the findings: A concern would be the potential for another factor leading to the increase in coralline cover which would be the use of a different data recorder or surveyor for the later dataset leading to differences in cover due to human sampling (over-estimation by person sampling later) and not ecology.
Additional comments: I believe this manuscript to be within the scope of PeerJ. This study was generally well designed, depending on the number of “before” samples collected the study may be well replicated, and was conducted for 7 weeks to determine responses to experimental treatments. The overarching goal was to determine how fishing may affect Los Cóbanos reef. Researchers addressed this by investigating changes in the benthic community following the exclusion of piscine macroherbivores using cages over a period of seven weeks. The cages may have provided refuge for herbivorous invertebrates which graze fleshy macroalgae and complicate interpretation of the results. The paper generally reads well, is interesting, and should be of interest to both the research and applied ecology communities. My main concern would revolve around the interpretation of the results (similarly to the previous reviewer comments). My main concern would be the potential for another factor leading to the increase in coralline cover which would be the use of a different data recorder or surveyor for the later dataset leading to differences in cover due to human sampling (over-estimation by person sampling later) and not ecology. The number of replicates used for the caging experiment may be a concern. I did not see any before control points in figure 2? I was expecting to see four control points and four points where cages will be installed (randomly selected) before the experiment and then four control points and four cage points after 7 weeks. Im a little concerned about replication or maybe I missed something. If these concerns could be alleviated then there are only minor comments.

General comments.

1)	Line . 48-50 - Could state how fishing indirectly leads to the favor of “competitive fleshy algae and turf”. For example, “By removing herbivorous fishes that consume macroalgae, fishing leads to release of competitive fleshy …”. The connection between fishing and algae/turf seems direct as previously written.
2)	Line 152-155. Im unclear on what “resist” means. “CCA tends to resist lower light…”. I think generally CCAs perform well in lower light levels but I think youre making the case CCAs may be less common in lower light as they ”resist” lower light. Maybe you meant they can tolerate lower light or perform better under lower light? It looks like the next sentence supports CCAs may do well in low light so id clarify.
3)	Line 159-164 – These results for water flow and light differences within and outside of cages could be reported in the results section rather than the methods section. These results help with the interpretation of the caging effects.
4)	Line 163-164 – Id maybe reword this. “any” unwanted physical cage effects could include many other factors (alteration of sedimentation, structure stops drifting algae, structure attracts mobile fauna, etc.). Id consider rewording to “indicating that the physical structure of cages do not affect light or water flow“ and not “any unwanted physical effects”.
5)	Line 179 – I believe the test is Levene’s Test not Levene Test.
6)	Line 198 – I believe that with such low numbers of permutations (perms = 35) its recommended to conduct monte carlo simulations.
PERMANOVA+ for PRIMER: Guide to Software and Statistical Methods by Marti J. Anderson, Ray N. Gorley & K. Robert Clarke (2008) - “The last column of the PERMANOVA table (‘Unique perms’) indicates how many unique values of the test statistic were obtained under permutation. Recall that PERMANOVA does not systematically do all permutations, but rather draws a random subset of them. In the example (Fig. 1.10), this value is very large (9862) and close to the number of random permutations that were chosen to be done by the user (9999). This means that only a few repeated values of pseudo-F? were encountered under permutation, and the number of unique values is plenty enough to make reasonable inferences using the resulting permutation P-value, as shown. This information is important because, in some cases, the number of possible permutations is not large, and very few unique values of the test statistic are obtained. In such cases, a more meaningful (but approximate) P-value can be obtained by random sampling from the asymptotic permutation distribution instead (see the section Monte Carlo P-values below).”
7)	Line 204 – add “and” before “turf algae benthic cover (…”.
8)	Line 194 – 205 – I would think a concern may be that the conclusions could be due to factors other than dramatic increases in CCA cover that have been brought up before by reviewers. One factor that could yield the results would be different people collecting the data. Maybe I missed where this was stated but I would like to know if the same surveyor took the data as this is commonly the first thing of concern when taking community data. Are changes due to communities responding to time, space, or experimental treatment or are differences due to how the data were collected.
9)	Line 265 – No need for “benthic” cover. CCAs are benthic and just CCA cover would work.
10)	Line 186 – I think this should be high “nutrient” availability without the s.
11)	Line 295 – CCA tend to “resist” lower light. As stated above I think CCAs perform well at low light and may even perform optimally depending on which species and whether they are in polar versus tropical areas. I think there may be a better term than resist. If youre stating they perform better at lower light relative to turf algae maybe use “tolerate lower light”.
12)	Line 316 – After 51 days of “the” experiment or after 51 days of “experimentation”.
13)	Line 340-341 – I don’t quite get this statement. Id reword “…the CCA cover increased after 16 and 8 weeks already, respectively…”. Maybe …”After 8 and 16 weeks, respectively”?
14)	Line 354 – Maybe reword ”CCA have been found to survive overgrow by filamentous turfs over long periods of time” to “being overgrown” or “overgrowth”.
15)	Figure 2 – I don’t see any before control (black circle) points on the figure? I only see 2 points for the before cage (black square)? Was there only two before surveys for plots? I think I may have missed the replication. Line 138-139 – “Plots with a similar benthic community composition to the enclosed ones were selected as controls (70 x 70 cm2)”. How many plots? I don’t see a clear description of replicates for control/treatment for before and after. Figure 2 states "dotted lines" but I think that should be "dashed lines".
16)	Figure 4b – caption could be reworded. Relative benthic cover of turf algae in enclosed and control areas, before and after the experiment. Figure 4a should have a similar caption to 4b but lacks the statement “before and after the experiment”.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: RAPID RELATIVE INCREASE OF CRUSTOSE CORALLINE ALGAE FOLLOWING HERBIVORE EXCLUSION IN A REEF OF EL SALVADOR
Review round: 1
Reviewer: 3

Output:
Basic reporting: Ilosvay et al. survey an impacted coral reef off the coast of El Salvador and use experimental grazer exclusion to explore the effects of herbivorous fish and exclude these species to simulate over-fishing.  The most interesting part of the study (in my opinion) is the reaction of coralline algae and turf algal species to these cages.  The authors touch on this in the discussion, but I feel this is a story about turf reduction and not necessarily an increase in corallines.
Experimental design: Can you expand on the methods you used to measure percent cover before the cages were installed?  Were different layers not analyzed?  This topic is touched upon in the discussion, but possibly these results show a decrease in fleshy seaweed and not an increase in corallines.

While I understand the logistical limitations of open cages, I'm not sure a 24 h period measuring flow and light is enough to show that flow and light did not affect your results.  Do you feel the 24 h period measured was representative of the study period?

It seems that there were grazers around, but that your sampling methods did not catch them, although I think that you discussed this, I think non-fish grazers might be a big part of these results.

My other main concern is turf numbers dropped in the control plots as well, and I think this indicates an abiotic factor playing a role and then the biology magnifying this.  I think in your discussion you need to touch on multiple factors affecting your results.  Such as season and cage effects.
Validity of the findings: The findings of this study are valid and important.  But I do feel that potentially season and the cages played a role in the results.  In the discussion the topic of multiple stressors needs to be discussed.  The limitations of the experimental design need to be explained more as well.  Even with these limitations, I do think there are interesting patterns that emerged from this study.
Additional comments: See above text.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: RAPID RELATIVE INCREASE OF CRUSTOSE CORALLINE ALGAE FOLLOWING HERBIVORE EXCLUSION IN A REEF OF EL SALVADOR
Review round: 2
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: I think the authors did a great job at addressing the reviewer concerns and I am supportive of this manuscript being published. Some small comments were to potentially add a clarifying statement to line 202-204 that the jitter function was used to reveal points with the same community assemblage that were overlapping and line 331 changing "...period from dry to rainy season" to "...period from the dry to rainy season". The sentence from 437-440 was quite long. One option may be "It is most likely that the combination of grazing macroinvertebrates, increased nutrient concentration and turbidity as a result of seasonal river run-off, and potential effects of the cages, tipped the balance from turf algae to CCA".


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: RAPID RELATIVE INCREASE OF CRUSTOSE CORALLINE ALGAE FOLLOWING HERBIVORE EXCLUSION IN A REEF OF EL SALVADOR
Review round: 2
Reviewer: 2

Output:
Basic reporting: Pass
Experimental design: Pass
Validity of the findings: Pass
Additional comments: Thank you for your consideration of my comments.  Great work!


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: USING NATURAL HISTORY COLLECTIONS TO INVESTIGATE CHANGES IN PANGOLIN (PHOLIDOTA: MANIDAE) GEOGRAPHIC RANGES THROUGH TIME
Review round: 1
Reviewer: 1

Output:
Basic reporting: A generally well presented MS, but a number of specific points ought to be addressed subject to the trajectory of this MS.

Specific points:
L42 – most species are nocturnal. Exception is the P. tetradactyla. This should be caveated.
L43-45 -  There is debate on whether pangolins are the most trafficked mammals. Consider revising to ‘are considered to be…’ You may need to amend references for this point.
L44-49 – I think you are referring to international trade reported to CITES in this sentence. It cannot be said based on available evidence and the literature that most ‘trade’ has involved the Asian species. African species – like the Asian pangolins – have been traded and consumed at a local level through history. You could amend this to international trade reported to CITES and cite Challender et al. 2015 and/or Heinrich et al. 2016.
L46 – I would state large populations collapses in parts of Asia. Available evidence indicates the Chinese, Sunda and Philippine pangolin have been exploited at a higher rate than the Indian pangolin.
L46 – Need to change to apparent increases in exploitation of African pangolins if referring to international trade – legal or illegal – because while it appears this is the case, we don’t have unbiased data indicating that this is so. If you want to cite increases in exploitation referring to local use as well in Africa cite Ingram et al. 2018.
L47 – I would remove reference to Aisher. It is redundant here.
L50-51 – there are issues with captive breeding, but they don’t prevent such programmes from exiting or breeding pangolins. Taipei zoo breeds Chinese pangolins for example. Suggest revising this wording.
L52-54 – The Red List assessments are based on time periods looking forwards and backwards. Please check whether this paragraph reflects this or needs amending to reflect this. I suspect the latter.
L56. Illegal trafficking as opposed to legal trafficking. This requires editing.
L57- what does regional mean in this context? Within a country, or countries collectively (e.g., W Africa).
L59 – what specifically do you mean by stricter regulations in recent years? Do you mean amended laws? Movement of pangolins to CITES App I?
L60 – I would remove reference to Chin and Pantel 2009. This is now eleven years old and not the best source for this. See this document which discusses point explicitly: https://cites.org/sites/default/files/eng/com/sc/69/E-SC69-57-A.pdf
L61 – the actual extent of illegal trade is unknown. This is not unique to pangolins of course but applies to virtually all trafficked wildlife derivatives and other commodities. I disagree that species distributions are unknown. They are being honed cumulatively based on new, local data. Updated distributions were published in the IUCN RL in 2019. This requires editing.
L90 – typo
L105-106 – I’m not sure Mittermeier and Wilson (2011) is the best source for this information. New authoritative sources have been published on pangolins which contradict this table for white-bellied pangolin. See https://www.sciencedirect.com/science/article/pii/B9780128155073000095.
L138 – what resolution?
L154- population size of land-use variable, but not both, correct?
L165-169 – MS reverts to alternative font.
L170 – how many specimens/record did this rule result in being omitted?
L233-234 – Makes me wonder why all these records are not in GBIF!
L256 – why highlight this specimen specifically here?
L256-259 – Anything else to say about these results? It reads like you have given them short shrift.
L263 – extinction risk. No need for ‘status’ here. Or, say IUCN RL status.
L264 – occur in?
L271-273 – This oversimplifies things a little. See previous comments on trade characterisation. There appears to be increasing pressure on African populations. See Ingram et al. 2018. Aisher et al. 2016 is again redundant here.
L275 - There are two species of Smutsia - are you referring to them both here?
L278-280 – I think you are referring to Asia here, but I’m guessing because it is not clear. In Asia the meat is considered a delicacy by some in countries including Vietnam and China, but is also consumed locally around protected areas too. Again Aisher et al. 2016 doesn’t add any value here.
L309-311 - This is possible – though it is worth noting here that the maps were updated in 2019, which you will need to recognise here.
L323-330 – Although these measures (and the omission of certain data points) were designed to deal with error as far as possible.
L333 – I think you mean threatened species here, no just endangered (=Endangered) here?
L340-342 – I’m not sure this is a fitting final sentence for reasons similar to the above point. Suggest reviewing.
Fig. 2 – I suggest zooming into the Palawan faunal region for the Philippine pangolin. Currently, this map is not particularly useful but it is hard to see where the points are in the species range.
Fig. 3 – New RL maps available from the RL version 2019-3.
Experimental design: I have some specific thoughts/questions on experiment design:
L74-77 – why weren’t other museum records included in the analyses? Why was this limited to GBIF+NHM London?
L103-104 – Why did you use range maps from 2014, and RL status from 2020? Why not use the range maps published with the new Red List assessments in December 2019? This would make the analyses as up to date as possible.
L103-104 – The map for Manis crassicaudata is incorrect and should not reflect distribution in Myanmar. This is an error on the IUCN Red List account for that species. It is not an insignificant error in my opinion and ought to be corrected.
L148 – I suggest the present day be 2019 using updated RL maps. As this paper – if accepted – would be published in 2019, using 2014 (i.e., old maps) need justification.
L284-293. I think this slightly mischaracterises habitat use and occupancy of pangolins in modified landscapes. While habitat loss does pose a threat, e.g., in SE Asia, the Sunda pangolin, like the white-bellied pangolin (and others) is able to adapt to modified habitats e.g., oil palm plantations, gardens and rubber plantations etc. This is documented in the literature (see here https://www.sciencedirect.com/science/article/pii/B978012815507300006X?via%3Dihub and the RL assessment for this species). However, there is little detailed knowledge on things like relative densities between tropical forest and oil palm plantations. I make this point because one could have perhaps been able to predict – based in part on this – that this would have been the outcome of this research. I suggest this and similar insights be captured in the discussion in Lines 290-319.
L334 – Fine, but most of these 437 were not used in the analyses, correct?
Validity of the findings: Some specific points:
L277-278 – Doesn’t this seriously undermine the framework being proposed in the paper? To be honest, I’d be surprised if you had found a mechanism using these data.
L337 – I think this would have been a remarkable finding if it were. In fact, I’m not sure that hypothesis that it would have been would have been a sound one.
L338 – 340 – This feels like a limp ending. It reads as though, despite your efforts, you are saying that the real value in using museum data is to use the old records because they are not available anywhere else. The MS needs another couple of sentences that really underlines the potential of this research to inform contemporary conservation.
Overall - I think the paper is flawed because it does not place the findings of the modelling work in a realistic context for pangolins, i.e., relating the findings adequately to the species ecology. The study would be have been much stronger, I suspect, had the authors collaborated with researchers that understand pangolins and their ecology.
Fig. 2 – The range map for the Indian pangolin is incorrect. I recommend downloading the corrected version from the IUCN RL 2019-3 and using that (alongside new maps for all species).
Additional comments: This is an interesting study but I am not convinced in its current form the MS meets the requirements for publication. The MS proposes use of framework but doesn't really present a framework. It uses a novel method but seems to conclude that the best use of museum data is as older locality records. Nor does it place the findings of the modelling work in a realistic context for pangolins.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: USING NATURAL HISTORY COLLECTIONS TO INVESTIGATE CHANGES IN PANGOLIN (PHOLIDOTA: MANIDAE) GEOGRAPHIC RANGES THROUGH TIME
Review round: 1
Reviewer: 2

Output:
Basic reporting: The paper is well written, the analyses carefully done and clearly explained, and the figures and tables clear and well documented. There are some minor errors in the literature cited.The data are also available for reanalysis.
Experimental design: I think that some additional analyses would make the paper much more sound scientifically. It is not a question of an incorrect approach as stands, just that there is an opportunity to enhance the results.
Validity of the findings: Nothing really to add here, but I will add my comments and suggestions relevant to this point below.
Additional comments: I have made a number of comments on the pdf attached. These tend to be more detailed comments and suggestions. I have two additional broader suggestion.

The Introduction seems to end with a presentation of results, even some discussion. This does not fit well, and it would be better to tie the introduction into the literature on evaluation of threats, threat assessments, and the Red List process. This links the paper more clearly to the process of improving threat assessments, especially in a spatial context. The ending of the introduction needs to be revised to address this.There are a number of more recent papers the discuss the improvement of threat evaluation (e.g. some of the work of Lucas Joppa).

In the section lines 300-320, there is the statement "solution would be to use IUCN range maps along with information on habitat suitability" however a group from IUCN recently published a paper with explicit instructions on how to do this, developing a "Area of Habitat" map. here is the citation.

Brooks, T.M., Pimm, S.L., Akçakaya, H.R., Buchanan, G.M., Butchart, S.H.M., Foden, W., Hilton-Taylor, C., Hoffmann, M., Jenkins, C.N., Joppa, L., Li, B.V., Menon, V., Ocampo-Peñuela, N., Rondinini, C., 2019. Measuring Terrestrial Area of Habitat (AOH) and Its Utility for the IUCN Red List. Trends in Ecology & Evolution 34, 977–986. https://doi.org/10.1016/j.tree.2019.06.009

I think that the quality of the analysis, and its relevance to Red List assessments, would be greatly enhanced by calculating AOH from the Red List maps used in the paper, and rerunning the analyses. I have done AOH recently in a paper submitted, and it is not complicated. The paper as it stands is fine, but with a lot of caveats. I think this would greatly improve the conservation values of this analysis but updated the maps to deal with the authors expressed concerns about problems with the IUCN polygons. AOH is far less complicated than running SDMs, but with a similar impact in improving the results.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: USING NATURAL HISTORY COLLECTIONS TO INVESTIGATE CHANGES IN PANGOLIN (PHOLIDOTA: MANIDAE) GEOGRAPHIC RANGES THROUGH TIME
Review round: 1
Reviewer: 3

Output:
Basic reporting: This paper is well written but needs more description of the taxa in the text for readers to be drawn into the project.  Some of the analyses could be described with a few additional details to help readers not familiar with how this science is done.
Experimental design: No comment
Validity of the findings: This is a timely topic to be looking into.  The authors have clearly done an extensive amount of work georeferencing the specimens in the NHM collections which is valuable.  The way they assess the accuracy of historical records is appropriate.  Effort was put into incorporating different types of data to add to their comparisons about the analyses and testing hypotheses with the available data (which unfortunately are few).
	Having read through this, however, I feel that the authors may have missed the most important aspect of what their data show and why.  The specific question being asked was whether or not there was a change in presence/absence over time (and the answer is "not that they could detect change"), but the true problem is not with the historical data, it is with the modern data (Note, that I feel that this was at least an indirect message given by Boakes et al. in their work).  We basically do not have suitable modern collections (or data points on presence/absence) for pangolins to say much of anything about changes in distribution in the present during the current intense harvesting pressure which they note.  This is, of course, stunning given how many have been caught for trade.  As I read the paper, the authors are assuming "present day" data in the form of IUCN range maps are somehow updated and "modern" and I highly doubt they are.  I didn't see anywhere in this manuscript where the way that IUCN ranges were determined (and the broad geographic scale is an important factor) is described to any extent.  I am guessing that experts were involved, but I am also guessing that a good portion of the same historical records was incorporated (or at least basic knowledge of those data) into generating the IUCN range maps.
	I have been criticized by reviewers for using shape files of ranges in my own research and I believe they actually reflect well most of the time, ranges, but that is because well-constructed polygons of ranges include historical data, which makes them useful for some research questions and not for others.  I think these present data and their analyses can be published (and should be), but the message should be how critical it is for there to be surveys conducted regionally whereby data is gathered about the modern status of these species within these polygons.  Personally, I believe this should include getting modern specimens to, in part large part because I think it is likely there is substantial population structure within many of these species.  These populations may be differentially affected. Thus, although the historic specimens tell you about historical patterns, only with similar modern-day data can you truly assess change over time (and this has to be investigated across different scales.  For instance, my guess is that the range of and of these species on an island in SE Asia is smaller because of habitat loss, but I don't this study was designed at all to capture that.  What the authors should advocate for is the need for modern data (and specimens), because I think there will be ample evidence for areas where they no longer occur (and that those areas will related to human activities.

The scale at which this study was done is not overly interesting because the current range maps are likely based on roughly the same historical data.

Line 71.  delete "can," these records to provide data from before exploitation was so widespread.
Line 76.  What are present day range maps based on?
Line 97.  Give specific download information that GBIF provides.
Line 137.  Grid cell is brought up for the first time here, give more information on the term.  Grid cell size would seem to me to have potentially huge influence of interpretations of pattern.
Line 171.  Do you think anything more could be done with data on multiple specimens in terms historical abundances?
Line 196.  Would using decade to bin records have increased power?
How were the IUCN range maps made?

Help readers out.  Introduce pangolins by taxonomy, natural history, and current threat status at the beginning of the manuscript.
Line 256.  I am very interested in these kinds of projects, but I have know idea what the significance of this sample is.  Help readers out here.  How could there possibly not have been those data for this locality?

Line 274.  Fix this sentence to clarify whether you are referring to the genus in general or one species in particular here.

Line 314.  The statement about IUCN range maps illustrates a huge challenge for this paper.  It seems highly likely to me that these maps were made using relying extensively on the historical records, so there is complete circularity here.
Additional comments: This paper would best be restructured to highlight that historical records reflect percieved present distributions of these taxa well, but we are in great need of better modern data.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: USING NATURAL HISTORY COLLECTIONS TO INVESTIGATE CHANGES IN PANGOLIN (PHOLIDOTA: MANIDAE) GEOGRAPHIC RANGES THROUGH TIME
Review round: 2
Reviewer: 1

Output:
Basic reporting: This is the revision of a previous version. I have reviewed them both and this version is significantly improved. It is clearly written, well cited using the appropriate literature, uses solid statistical and spatial analytical approaches with maximal reduction in potential biases, and provides access to data and code. The figures are also clear and easy to interpret.
Experimental design: Well conceived and carefully explained in the Methods. This is much improved over the previous version of the manuscript. It asks important questions about understanding factors responsible for the decline in pangolin populations and how we might better track these using historical records.
Validity of the findings: The findings are supported by the data and analyses and the authors present the appropriate cautions in interpreting the results, given possible biases and errors in the raw data (not the responsibility of the authors).The paper does highlight the important value of museum specimen records. This has the additional importance of presenting the role of natural history museums when so many are at threat.
Additional comments: I am quite please with the responses to all of the prior comments and suggestions. The inclusion of AOH has greatly improved the manuscript.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: USING NATURAL HISTORY COLLECTIONS TO INVESTIGATE CHANGES IN PANGOLIN (PHOLIDOTA: MANIDAE) GEOGRAPHIC RANGES THROUGH TIME
Review round: 2
Reviewer: 2

Output:
Basic reporting: I commend the authors for responding to the comments with changes that have greatly improved the manuscript.
Experimental design: This is fine
Validity of the findings: This is fine
Additional comments: I continue to be concerned that they have not really used their results to address the primary concern going forward for documenting pangolin distribution change and that is gathering more than what seems to be quite limited data on the presence/absence of these species across their distributions.  The AOH maps are still based on limited knowledge (that is not really quantified in any documented way).  This an opportunity to discuss that (even if the crux of the paper is to use the historical records).  One potential way forward would essentially be e-mammal, which would be the creation of a citizen science database along the lines of e-bird.  This doesn't obviate the need for specimens because specimens harbor data that go far beyond a sighting (something not really brought up anywhere in the manuscript), but if one is simply trying to assess potential changes in distribution through time, these approaches will gather data that can be applied across the broad geographic (and subsequently temporal) scales being considered here to answer important questions about how pangolins are responding to landscape change through time which is a start.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: AROUSAL MODULATES THE MOTOR INTERFERENCE EFFECT STIMULATED BY PICTURES OF THREATENING ANIMALS
Review round: 1
Reviewer: 1

Output:
Basic reporting: I find the concept of this paper clear, the writing is straightforward and the authors use professional English language. The article structure is good, figures and tables are informative, raw data files are well prepared. The paper is self-contained with relevant results to hypotheses.
My only concern here is that I did not found a sufficient background/context provided, that is the authors did not consider several papers that would be relevant to mention here as these studies also deal with the effect of emotional arousal regarding threatening objects. Especially since in the latter two of these papers, the arousal – motor interference effect is also relevant. Furthermore, since these studies showed very similar results regarding threatening stimuli at low vs high arousal levels. These should also be included in the discussion and the authors could reflect to their results in light of the arousal stimulation effect.

Zsido, A. N., Deak, A., & Bernath, L. (2019). Is a snake scarier than a gun? The ontogenetic–phylogenetic dispute from a new perspective: The role of arousal. Emotion, 19(4), 726.

Zsido, A. N., Bernath, L., Labadi, B., & Deak, A. (2020). Count on arousal: introducing a new method for investigating the effects of emotional valence and arousal on visual search performance. Psychological research, 84(1), 1-14.

Zsido, A. N., Matuz, A., Inhof, O., Darnai, G., Budai, T., Bandi, S., & Csatho, A. (2019). Disentangling the facilitating and hindering effects of threat‐related stimuli–A visual search study. British Journal of Psychology.
Experimental design: The paper introduces a cleverly designed experimental setting that clearly measures what the authors intended to. The research questions are well-defined, relevant and meaningful. The investigation performed is rigorous, the sample size and trial repetition is great, and the authors did a really great job checking the emotional variables of the stimuli used. The research presented here definitely fills a gap. Methods are described with sufficient details and information so that further studies can replicate it.
Validity of the findings: Please report the full statistical results for the post-hoc tests (not just p values).
Some of the analyses (e.g. error rates) showed only small to medium effect sizes. This, with the high sample size and p values close to .05 could signal false positive results. This should be explicitly mentioned in the discussion.
The studies I referred to earlier should be mentioned in the discussion since they showed very similar results.
I really liked the occupational-applied framework of this research paper and I think that is a very important thing to put basic research in such a perspective. Yet, as PeerJ reviewer suggestion highlight: Speculation is welcome, but should be identified as such. So please make sure this is true for the discussion of the results.
Additional comments: No further comment.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: AROUSAL MODULATES THE MOTOR INTERFERENCE EFFECT STIMULATED BY PICTURES OF THREATENING ANIMALS
Review round: 1
Reviewer: 2

Output:
Basic reporting: The manuscript is well written and clear in each part (intro, method, results, and discussion). The literature cited in the introduction is appropriate, but in some cases, unfortunately, misinterpreted and misguiding (see more comments on this in the next sections). The figures are relevant to support authors claims, even though the supplemental material does not appear to be complete or easy to access for everyone. It would have been useful, for example, to attach a translated version on the consent form or the approval documentation. As for the ERPs, I appreciate the inclusion of the average amplitude for each subject, but a text file containing the time course of the ERPs would have been probably more useful because it allows to replicate the figures reported in the manuscript and the average of a certain time window on interest can be easily calculated from there.
Experimental design: The experimental design is conceptually clever, and the details well described. However, the orthogonal manipulation of valence and arousal is not supported by the literature and there are several evidences in this regard:
1.  The authors are correctly referring many times to Lang’s studies, but in that model arousal and valence are NOT orthogonal. As the authors described in the introduction, valence indicates the motivational system that is supposed to be activated in a given situation (appetitive or defensive), and arousal the level of activation. Arousal and valence are expected to covary: as valence ratings increase, arousal ratings increase as well. This is also the reason of the so called ‘boomerang shape’ for the affective space defined by the IAPS.
And in the affective space defined by the IAPS, there are no neutral stimuli high in arousal. It simply cannot happen. In fact, the authors basically selected stimuli that would be considered NEUTRAL in almost every study. In a scale that varies from 1 to 9 for arousal, the authors selected stimuli with average scores of 4.5 or 5.5 (values reported in Table 2). The fact that there is a significant difference between these values it does not mean that the manipulation was successful. High arousing stimuli are considered pictures with scores of 6.5-7 and above, and low arousing stimuli are considered pictures with a score of ~2.
2. There is no proof of positivity offset (advantage of positive stimuli at low arousal) and negativity bias (advantage of negative stimuli over positive stimuli at high arousal) in ERPs literature. This is only a conceptual model developed by Cacioppo in the ’90 that has been largely dis-proofed (see Weinberg & Hajcak, 2010; Beyond Good and Evil: The Time-Course of Neural Activity Elicited by Specific Picture Content). Over 20 years of ERPs research found that, when stimuli are balanced for arousal, there is no negative bias in P3-LPP component.
3. The authors misinterpreted, in part, the concepts of positivity offset and negativity bias. These concepts does not necessarily decries differences between positivity and negativity in terms of absolute amplitude. Instead, Cacioppo (Cacioppo, Gardener, Berntson, 1997; Beyond Bipolar Conceptualizations and Measures: The Case of Attitudes and Evaluative Space) conceptualized the negativity bias, from Miller studies on approach avoidance gradients from the ’50, as a difference in the transfer functions of positivity and negativity (from the article cited above, at page 13): “ With each unit of activation, the change in negative motivational output is larger than the change in positive motivational output”. In short, the difference between positivity and negativity is essentially in the slope, when activation (arousal) varies from low to high).
Validity of the findings: All the methodological procedures are perfect and well described. From a methodological point of view, the results are perfect. However, the validity of the findings is unfortunately questioned by the conceptual limitations. The orthogonal manipulation of valence and arousal would not be positively accepted in the field, together with the problematic conceptualization of the negativity bias. An additional issue is that the authors found their effects in Fz and Cz, while all the ERPs studies cited in the introduction in reference to picture viewing (e.g., Cuthbert et al., 2000; Keil et al., 2002; Polich, 2007; Sabatinelli et al., 2007; Schupp et al., 2003) are in Pz or cluster of sensors around Pz. This is another point that would probably not be well received by experts in the field.
Additional comments: Despite being a well-written article with a good methodological approach for ERP analysis, there are major conceptual issues that does not make the study suitable for publication and cannot be addressed in a revision. I would invite the authors to re-consider the orthogonal manipulation of valence and arousal and eventually replicate the same study by keeping in mind how the negativity bias was originally conceptualized.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: AROUSAL MODULATES THE MOTOR INTERFERENCE EFFECT STIMULATED BY PICTURES OF THREATENING ANIMALS
Review round: 1
Reviewer: 3

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The present study utilized the P3 component, reaction time and error rate to examine whether arousal and/or valence drives the allocation of attentional resources. The authors found that the parietal P3 during the negative valence condition was increased in the high arousal condition compared to the low arousal condition. In the positive valence condition, the parietal P3 was reduced in the high arousal condition compared to the low arousal condition. Similar results were obtained at the frontal and central electrode sites. The question the study tries to answer seems important but the paper needs some clarity to have a bigger impact. Overall, the paper is difficult to follow.

1) The authors provide a lot of detail in the introduction which is good and I appreciate the authors grounding their hypotheses in theory. In doing so, the hypotheses are numerous and difficult to obtain. I think the introduction would be clearer if it was re-ordered and the hypotheses were made more obvious. Also, the authors should make it more obvious why this study is needed. This information was hard to obtain from a first reading of the introduction. I suspect readers would have similar difficulties.

2) The authors mentioned that large artifacts were manually deleted. What was entailed in this process? Who was in charge of cleaning the data?

3) There a mistake in line 415. It states positive > neutral > neutral.

4) In the discussion, line 508, the authors say "In the emotional field..". Please clarify that.

5) Please tie in the current study more explicitly with the previous study (Liu et al 2017) in the discussion. The discussion could also use some reorganizing.

6) I don't believe the authors are testing the hypothesis that valence has a bigger effect on frontal/central sites than parietal sites. I don't see any comparisons of the P3 within the valence conditions are different electrodes. I could be wrong. The information was hard to obtain.

7) The abstract doesn't mention anything about the p3 finding, except the finding in the Liu paper. Authors should mention it in the abstract. Or alternatively, it might make sense to only look at the P3 at Pz since that is what the Liu et al study seemed to have done.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: AROUSAL MODULATES THE MOTOR INTERFERENCE EFFECT STIMULATED BY PICTURES OF THREATENING ANIMALS
Review round: 2
Reviewer: 1

Output:
Basic reporting: Language is clear and professional, sufficient background provided, the structure is good.
Experimental design: The experimental design is sound, hypotheses/research questions are well defined, details are sufficient.
Validity of the findings: The findings are novel and interesting, the data support the claims the authors make, the conclusions are well stated. I think that the results will have a great impact on the field and attract many citations in the future.
Additional comments: The authors did a really great job responding to my comments. I wholeheartedly recommend this manuscript to be accepted without further modifications. I am looking forward to reading more papers form the authors investigating this topic.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: AROUSAL MODULATES THE MOTOR INTERFERENCE EFFECT STIMULATED BY PICTURES OF THREATENING ANIMALS
Review round: 2
Reviewer: 2

Output:
Basic reporting: No comment
Experimental design: No comment
Validity of the findings: No comment
Additional comments: I would like to thank the authors for their work on changing the manuscript according to my comments. I appreciate their efforts. I have just two small comments below.

1) Please replace insignificant with non-significant in the method and results section.

2) The pooling of 21 channels to create a centroparietal P3 seems excessive. The authors point to figure 1 as their justification; however, the activity is maximal in a smaller number of channels than the authors used. I guess I would suggest that the authors check the prior literature to see what makes sense to use for their pooling average. Looking at figure 1, I'd only pool at the sites very near Pz (including Pz).


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: CHARACTERIZATION AND COMPLETE GENOME SEQUENCE OF PRIVATEER, A HIGHLY PROLATE PROTEUS MIRABILIS PODOPHAGE
Review round: 1
Reviewer: 1

Output:
Basic reporting: Basic Reporting
The submission is a comprehensive, well-written report of the characterization of a new podophage with potential clinical applications. It originates from a research group with extensive experience in bacteriophage characterization. The article structure is appropriate and the methods well-described. The introduction spends time discussing a group of articles using polyvalent phage preparations with insufficient discussion of the host being studies and the biological importance of phage in the gut, their role in the microbiome and the sewage source. The discussion is similarly rather sparse.
Experimental design: Experimental Design
The experimental design is appropriate and comprehensive, characterizing the phage titer, infectivity characteristics, genome features and proteins amenable to mass spectrometry. The methods are presented in admirable detail (with a few minor suggestions below). The methods appear to be well-established methods of phage analysis.
Validity of the findings: Validity of the Findings
The findings appear credible and internally consistent. The figures are well-presented and mostly well-done.
Additional comments: Specific Comments with line numbers

Overall admirably detailed methods
86 Phage Storage conditions?
80 nuclease supplier?
104  Typo: 	Hernanded
140-PhageTerm in Galaxy? Or BASH?
148-Cutoff? Rho-independent termination scan not mentioned again….were none found?
The Genbank has record noting that frameshift was verified by MS.  Nice!
Both GENBank and ShortReadArchive records appear complete
173 Morphology also reminiscent of kuraviruses
179 Appreciate that the software for the figures were acknowledged.  A lot of people ignore that intellectual property.
197 Dried sample storage? Was it reconstituted in methanol or……….?
200 The MS analysis should be cited.  There are a number of papers on spectral counting statistics for peptide identification would be appropriate since there is some correlation implied between the SDS-PAGE gel band density and MS spectral count quantification. Is the MASCOT procedure proprietary? Also, explanation of the distinctions between the total spectrum count and unique peptide count would be a good addition.
221 infective characterization?  Odd phrase
222- Delete citation; it’s in the methods
227 –“reproduction” replace with ‘infection kinetics’
238 Any discussion of P. mirabilis B446? It is strikingly different.  Any characterization of P. mirabilis B446 that one could speculate about differences? Host range is not mentioned in the discussion.
299 “Assigned predicted functions are here discussed for both the
300 expected genetic repertoire, as well as several unusual genome features”. I understand what is being said here but even in passive voice, that can be better expressed.
307 You could search the non-Kuraviruses for DTRs, yes?  Was that done?
315 location of Kuravirus holin and endolysin?
331 “The specific role of ter proteins expressed by phage
331 genomes has not yet been explored” Well, it has been explored but not established.
341 How much is the similarity between Privateer thioredoxin and T7? Or is this just raw speculation?

Fig 2B nice looking curve; burst size of about 100 is as to be expected
Would prefer reporting of the three values not a representative curve for 2A and 2B
Can’t find this reference anywhere.  Is it “In Press” or something? If status is just “accepted”, that should be noted.
Ramsey J, Rasche H, Maughmer C, Criscione A, Mijalis E, Liu M, Hu JC, Young R, Gill JJ. 2020. Galaxy and Apollo as a biologist-friendly interface for high-quality cooperative phage genome annotation. PLOS Computational Biology.

Overall, very solid paper. My main criticism is that there is insufficient discussion of the host range, yet an attempt to extrapolate the host range to Proteus spp. and further extrapolate that to clinical applications.  All justifiable, but a few lines to delineate those ties are needed.
The use of Privateer against CAUTIs is mentioned in the introduction and conclusion, but it seems pro forma for funding purposes. There is room for more discussion that would justify that connection. Although the host range doesn’t provide strong evidence that the isolated host is close to the primary host of the phage, the infection and burst data clearly demonstrate that it is capable as acting as an anti-bacterial for relatives of P. mirabilis. The Milo, Ujmajuridze and Maszewska papers cited use polyvalent phage preparations, so there is some advantage to characterizing individual phages. Discussion of the host range of Cronobacter phage vB_CsaP_GAP52, in particular should be discussed. What is the level of similarity between endolysin of those two phages? (Holin in Cronobacter phage vB_CsaP_GAP52 has not been identified, either).


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: CHARACTERIZATION AND COMPLETE GENOME SEQUENCE OF PRIVATEER, A HIGHLY PROLATE PROTEUS MIRABILIS PODOPHAGE
Review round: 1
Reviewer: 2

Output:
Basic reporting: Very nicely written.
Experimental design: The research plan was well described, and methods were easy to follow.
Validity of the findings: Overall the data and conclusions were valid and well described. A few comments are listed to be addressed to clarify a few of the results and figures.
Additional comments: Specific Comments for the authors to address:
line 51: A space is needed between lacking and (

line 179 : Citation for software that has no manuscript typically has a link to download. This is not listed in the methods or the reference list for Inkscape.

line 121-122: Please add reference to table 1 with the host range strains of interest.

line 228-235: Figure 2 and the methods indicates three replicates of adsorption curves, one-step growth curves, and lysis curves. In the results, figure, and raw data for figure 2 only the representative image is shown. Please provide statistical analysis for each of these metrics. If necessary please put the other replicates data in supplemental material.

line 259-261: Many of the phages listed in the text which have protein similarity to Privateer are not shown in the comparative genomics figure 4. Please update text to reflect those that are described in figure 4 versus those that are only described in the phylogenetic tree in figure 6.

In the description of comparative genomic analysis (lines 258-265) with other phages proteins, which types protein are conserved (structural or throughout genome)?

Figure 1A: If possible increase resolution on this image.

Figure 4: Please indicate the species of each virus in the legend since these phages come from a diverse group of bacteria. Does the grey represent a certain percentage homology between each protein? If so can you indicate what percent homology that is in the legend.

Figure 5: Can you add rational for the shift to a higher molecular mass for the major and minor capsid band? The molecular mass listed in the table is 35kD and 53kD but appears to be closer to 38kD and 60kD in the gel.

Table 1: Please add EOP data for each strain, some this data is mentioned in the text on line 240 but is not replicated in the table.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: CHARACTERIZATION AND COMPLETE GENOME SEQUENCE OF PRIVATEER, A HIGHLY PROLATE PROTEUS MIRABILIS PODOPHAGE
Review round: 1
Reviewer: 3

Output:
Basic reporting: In this article, Corban et al. isolated and characterized a novel prolate podophage, Privateer, that infects a relatively narrow range of Proteus mirabilis strains. The article very well written, detailed, and easy to follow. I am extremely impressed by the meticulous discussion on putative phage encoded proteins. For example, instead of just reporting that Privateer encodes a potential thioredoxin, the authors explained the role of thioredoxin in bacterial stress response and how it has been shown to influence the primer binding affinity of phage T7 DNA polymerase. I have some minor comments/suggestions for the authors:

Line 50: Introduce space between “lacking(”
Line 145: I could not find Ramsey et al., 2020 and the reference does not have a DOI. Is the article in press? I could not find a BioRXiv version of this reference either. This will be a very useful reference for the entire phage community.
Line 228: From Fig.2A, seems like it should be “nearly 80%” instead of “80% of the phages were adsorbed to host’.
 Line 307: Can the authors expand on “Privateer is predicted to have novel end types”?
Experimental design: Line 69-72: Please provide a detailed protocol for the phage isolation technique here.
Validity of the findings: No comment
Additional comments: No comment


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: A VALIDATION OF ILLUMINA EPIC ARRAY SYSTEM WITH BISULFITE-BASED AMPLICON SEQUENCING
Review round: 1
Reviewer: 1

Output:
Basic reporting: The structure of the paper is apporpriate
Experimental design: The authors report a comparison between Illumina EPIC arrays and bisulfite amplicon sequencing.  They identified sites associated with smoking using the EPIC array and then attempted to validate 15 of the top sites using bisulfite amplicon sequencing.  The results are presented in Figure 1 which shows a scatter plot of the two techniques and Fig 2 which shows a Bland Altman plot.

My primary criticism is that the data as presented in unconvincing.  The BSAS approach was carried out on 82 individuals, yet we only see the data for the means of each of the 15 sites in the two groups in Figs 1 and 2.  It would be good to show the entire dataset so we can see how reproducible data is across individuals and not just across means.

Fig 2 is problematic  The x- axis should be the mean of the measurement of the two approaches but has negative values.  Moreover, as the authors state, AHRR has a mean difference of 0.23 in the EPIC array and only 0.04 in BSAS, yet none of the differences in fig 2 are greater than 0.1

Thus it is impossible to determine whether BSAS is actually working or whether the 40 cycles of PCR that are needed to generate sufficient amplicons are significantly biasing the methylation estimates (which could easily happen if the converted and unconverted CpGs have slightly different amplification biases.  Moreover, there is no mention of the coverage distribution of each amplicon across individuals.  The minimum coverage of 8 seems remarkably low as it limits the resolution of a methylation estimate

Some minor concerns are:

1) In the introduction it is stated that most CpGs are in promoters, which is not correct.

2) When the other methods for targeted DNA methylation profiling are presented, hybridization capture approaches are not mentioned.

3) It is unclear how edge r was used in the analysis, as all we see are figures 1 and 2 that do not involve any estimates of the significance of differential methylation
Validity of the findings: Other bisulfite amplicon approaches are not discussed so it is not clear if there is any novelty
Additional comments: See above


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: A VALIDATION OF ILLUMINA EPIC ARRAY SYSTEM WITH BISULFITE-BASED AMPLICON SEQUENCING
Review round: 1
Reviewer: 2

Output:
Basic reporting: Language, literature references etc. are fine.
Experimental design: The experimental design is ok. However, I miss in depth validation of the BSAS procedure - as mentioned by the authors in line  273 and 274 in the discussion part, they need to investigate, which differences can reliably be detected. that should be done in that manuscript to improve the quality.
Validity of the findings: findings are valid and nice, but I expect more details on the analysis. why did the authors just investigate the correlation between BSAS and the EPIC array for the individual groups and not for the complete data set? why did they not report on results of group comparison etc.
the result section is very weak and should be matter of major revision.
Additional comments: The provided manuscript by Noble et al. aims to investigate an interesting topic, as validation of the EPIC array is of great interest for the community. However the manuscript needs to provide more results tho prove the evidence of the findings. Why did the authors use different sample groups (cannabis/tobacco users vs. non-smokers) and provide just information on the correlation of the individual groups between EPIC and BSAS? they should use this data as a showcas and report also on the biological findings.

Moreover, I guess the authors missed to report the data of all CpGs they have investigated. I am quite sure that they did start with more than 15 CpGs and just picked out the results of the best one.

The manuscript needs more information on the capacity of BSAS. How many samples and CpGs can be investigated e.g. using a Illumina HiSeq or NovaSeq machine?


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: A VALIDATION OF ILLUMINA EPIC ARRAY SYSTEM WITH BISULFITE-BASED AMPLICON SEQUENCING
Review round: 2
Reviewer: 1

Output:
Basic reporting: Adequate
Experimental design: Adequate
Validity of the findings: Adequate
Additional comments: I still do not find the figures acceptable.

In figure 1 the x axis is labeled as mean, but presumably represents the log of the mean.  Why take the log of the mean in t he first place and if it is deemed necessary then at leas label the figure as log(mean).

In fig 2 it is very difficult to distinguish the results from the epic array and the amplicon data.  The different symbols are barely distinguishable.  Maybe present them side by instead of showing the on top of each other.  It would also be good to show the mean or median of each distribution


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: COMPARATIVE ANALYSIS OF THE COMPLETE PLASTID GENOMES OF MANGIFERA SPECIES AND GENE TRANSFER BETWEEN PLASTID AND MITOCHONDRIAL GENOMES
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript deals with the importance of chloroplasts and mitochondria genomes to identify mango species from the genus Mangifera spp.  The genomes from five mango species were described and compared. Some specific DNA fragments were identified as transferred from the chloroplast to mitochondrial genomes. The whole chloroplast genome phylogenetic analysis is suggested as a powerful tool to solve evolutionary relationships and discriminate among mango species.

Comments and suggestions:
The authors must review and correct the English language of the whole manuscript as it contains some grammatical, syntax, and spelling errors.
It is highly recommended to use passive voice through the manuscript; by using passive voice, readers will focus on the action and the results of the study. It is not really important for a reader to know who performed any action for the development of this study.
Some corrections:
-The excess of commas should be corrected.
-Lanes 33. No comma after “flavor”
-Lane 65. Published by Li et al. (2013)
-Lane 70. The illumina HiSeq 2500 system (XXXX) was used to sequence DNA samples in the paired-end…
-Lane 74. The Trimmomatic 0.38 tool was used
	-Lane 93. Circos… software/ web tool?

Abstract.
-Lane´s 25-26 sentence gives an incomplete idea. A higher resolution for Magifera species identification?
Experimental design: Methods and Material.
-Basic and essential information is missing about materials and method.
-First paragraph. Authors should include how many, and the scientific names of those Mangifera species that were sampled.
- The method used for DNA isolation should be mentioned in the methods section.
- The authors should mention and describe the kit or reagents used to construct libraries.
- In the methods section, software or web tools are not clearly distinguished. The software may include the complete name and version; web tools may include electronic address.
- Phylogenetic analysis. Authors should describe better the analysis including:
¿how many other available chloroplast genomes were used to compare their Mangifera data?; ¿how, and why those sequences were selected?; ¿how authors evaluated the reliability of the tree topologies?; ¿Which were the optimized parameters for each selected method?
Validity of the findings: Results and Discussion
- Lane 106. The approximated data numbers indicate as units ¿nucleotides? Or ¿sequences? It should be mentioned.
- This section is a general results description, but the in-depth analysis of what these results mean should be improved.
- Why were Sapindales selected for comparison purposes?
- Why variations or differences among sequences are in the non-coding region of the chloroplast genomes? What does this mean?
- The identified hypervariable regions were analyzed in deep? What about any region that may function as a molecular marker allowing to distinguish among Mangifera species?
- Figure 4. The legend colors should be found in the little colored bars from the out/ in sides of the circle, however. the figure quality does not allow to clearly distinguish each color, so it makes difficult to understand the genes identity and location.
- Besides considering the assembly errors that mitochondrial genes transfer may produce in the chloroplast genome sequencing, authors should analyze in deep what do these transfers mean or promote into the chloroplast genome? ¿Are always the same genes transferred from mitochondria to the chloroplast in other species genomes? ¿Any of these transferred genes could be a specific marker from one Mangifera species?
The results from four main objectives of the study were obtained and described, but not explained.
Additional comments: In this manuscript, several original data were obtained. Some of them deserve to be better analyzed and discussed. The main differences among the sequenced genomes should be explained, and those differences with other genus species should also be considered in the discussion section.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: COMPARATIVE ANALYSIS OF THE COMPLETE PLASTID GENOMES OF MANGIFERA SPECIES AND GENE TRANSFER BETWEEN PLASTID AND MITOCHONDRIAL GENOMES
Review round: 1
Reviewer: 2

Output:
Basic reporting: The context is poorly provided in the Introduction. The first and second paragraph lack a clear connection. The knowledge gap is not described in the introduction.
The results need to be better described, especially the contrasting results.
Experimental design: - Provide more details about how these species were identified. Are they part of a botanical collection? Who identified these species?
- Please provide the names of kits used for library preparation. 5-10ug of DNA is a high amount of DNA. Were all of this amount used for library preps?
- The methods used for phylogenetic inference need more detail, for ex. was the substitution model choosed based on what?
- The phylogenetic inference was performed using species from 5 families. Theses species shows any structural rearrangements? how was the sequence alignment done?
Validity of the findings: - The sequencing details, as the plastome depth of coverage needs to be described.
- The phylogenetic inference methodology needs to be revised and the results and discussion on this topic rewritten.
Additional comments: - Please use "plastid genome" or "plastome" when referring to the DNA content of chloroplasts.
- Line 65. Revise Li et al. citation.
- Line 106. Provide the unit of measure for raw data.
- Line 108-109. "GenBank database"
- Lines 115-121. Please describe in detail the gene content variation.
- Lines 141-142. The variable sites were identified using only Mangifera plastomes. The statement that this region could be "ideal molecular markers to distinguish Mangifera from other genera" has no support.
Line. 155. Similarity or identity?
Line 159. Adjust, infer of confirm?


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: COMPARATIVE ANALYSIS OF THE COMPLETE PLASTID GENOMES OF MANGIFERA SPECIES AND GENE TRANSFER BETWEEN PLASTID AND MITOCHONDRIAL GENOMES
Review round: 1
Reviewer: 3

Output:
Basic reporting: This work is interesting in terms of chloroplast genomes analysis between Mangifera species. Overall, the results are novel and presented in a logical order.  There are a few points that have to be addressed by the authors towards improvement of the manuscript.
-	Abstract should be rewritten to set a clear scope of the study, and not just report numbers and sizes.
-	Authors should give info concerning the different Mangifera species used for chloroplast genome analysis.
Why did they select these species? Is there any special physiological or morphological characteristic or relation to cultivated mango? Authors should incorporate photographs from these plants describing their phenotype and/or diversity. There is a lot of space in Suppl. Figs.
-	It would be beneficial to add plastome variation by region between the Mangifera species. SNPs and INDELs across coding gene regions, intergenic regions and introns could be described. Please add a graph about all the above.
-	Since authors discovered possible “Divergence Hotspot Regions to be ideal molecular markers to distinguish Mangifera from other genera”, these markers should be experimentally verified and presented, otherwise please delete it.
Experimental design: Please see above.
Validity of the findings: Please see above.
Additional comments: Please see above.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: COMPARATIVE ANALYSIS OF THE COMPLETE PLASTID GENOMES OF MANGIFERA SPECIES AND GENE TRANSFER BETWEEN PLASTID AND MITOCHONDRIAL GENOMES
Review round: 2
Reviewer: 1

Output:
Basic reporting: After the second revision, no fails were found!
Experimental design: After the second revision, no fails were found!
Validity of the findings: After the second revision, no fails were found!
Additional comments: Manuscript ID:
PeerJ-51712
Title: Comparative analysis of the complete chloroplast genomes of Mangifera species and gene transfer between chloroplast and mitochondrial genomes

The manuscript describes, compares, and discusses the chloroplast´s genome from various mango species. After revision, most corrections were done, and writing was improved. The previously missing and confusing data are precise now.

Minor comments:
 Table 2. Column 1. Photosynthesis (P capitalized). Column 2. Cytochrome Synthesis ( capitalized and an “e” is missing).


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: GENOME SIZE VERSUS GEOGRAPHIC RANGE SIZE IN BIRDS
Review round: 1
Reviewer: 1

Output:
Basic reporting: 1.	As more avian genomes become available, new opportunities arise for analyzing global patterns of genetic factors underlying the geographical variation in diversity of life on Earth. This paper examines how genome size (as available on a genome size database) correlates with how widespread species are, and also examines the relationship with number of chromosomes, body size and brain-size (as proxy for behavioural flexibility). This is a novel and interesting topic, but in view of the many historical and ecological factors that affect range-size, the interpretation of possible correlations may be challenging. The paper is clear and well formulated, in professional English.
Experimental design: 2.	The introduction provides a clear logical framework, and the structure of the MS, and logics of the analysis, is fine, over all. I assume that brain-size (in various places in the text) refers to residual brain-size. The figures are relevant and clear. I could not see to what extent the raw data is made available
Validity of the findings: 3.	While I see no problems with the data analysis that was done, the authors are missing some important macroecological facts and have overlooked some confounding factors. These need to be taken into account when interpreting the data.
One premise of the study is that birds can fly, and therefore should be little affected by physical barriers (lines 65-66). This is true for high-latitude environments, but note that only few avian lineages are widely distributed (thus, only 5 of 145 families of passerine birds are represented on all continents). Most lineages have diversified only within a rather restricted part of the land surface, and many tropical species are (albeit able to fly) highly reluctant to cross unfamiliar habitat. The dispersal of birds are highly dependent on their life stragegies of the particular lineages! With respect to the ability to adapt to a broad range of climates, birds are often assumed to be constrained by niche conservatism. But it is important to note that niche conservatism appears to be assymetrical: ancient tropical groups cannot easily adapt to, and expand into, cold/harsh climates, but groups that evolved at high latitudes and are able to tolerate cold are actually thermally flexible and easily adapt to new climates, and are often founders of breeding populations (and new proliferations of species) within the tropics (e.g., Smith et al. Ecol. Letters 15(2012): 1218-25; Khaliq et al. J. Biogeogr. 42(2015): 2187-96; and see also Winger et al. in Biol. Reviews 94(2018): 737-753).  For these reasons, the relationships that this study sets out to explore will probably differ greatly between northern and tropical (+austral) biota. The result may therefore depend much on the geographical sampling, and in the interpretation of the results, it is essential that latitudinal differences are taken into account and also that attention is given to how well the sample of species (with genome data) represents the global avian fauna.
The authors refer much to the studies of range-size variation by Kevin Gaston, some 20 years ago. Gaston’s papers used range-size data of exceedingly poor quality (very generalized range-maps from a fieldguide), which means that these studies greatly underestimated how many species that have really tiny distributions (and even the data from BirdLife provide rather crude range-size estimates). One of the great challenges in recent studies of macroecological patterns has been how to explain the highly aggregated geographical distribution of species with tiny geographical ranges in certain tropical areas (oroclines or locations near the margins of continental plates, and in archipelagos (see, for instance Rahbek et al. in Proc. Royal Soc. B 274(2007): 165-174 and Science 365(2019): 1108-1113, and for a review of diversification in archipelagos see Jønsson et al. in Ann. Rev. Evol. Evol. Syst. 48(2017): 231-253). The 50% of avian species with the smallest geographical ranges are found almost entirely at <30 degrees latitude (resulting in very high residual values in comparison with ecosystem productivity in certain areas). Unfortunately, the 50% of  avian species with the smallest geographical ranges are very poorly represented in the genome data (thus, only one of the 65 species in Fig. 1B! [Serinus canaria]). In other words: this study is highly biased towards species with large to moderately small geographical distributions, and seriously neglects the large number of tropical species with really tiny distributions (this bias probably explains the negative association between range size and body size). These are of two kinds: some old relict species, and paraspecies within some lineages that underwent rapid recent radiation within certain oroclines and archipelagos.
Additional comments: 4.	As long as small-ranged species is so strongly underrepresented in the genome dataset, there is not much to do about this bias (other than waiting for more genomes to be available, especially from the b10k project), but it must at least me mentioned as a shortcoming of the study, and it would probably help the interpretation if tropical species (centroid of range at <30 degrees lat.) are conspicuously marked with colour symbol in Fig. 4. Another bias (or complicating factor) may be that high flying powers (aerial foraging) has already been found to correlate with small genome size (Zhang et al. 2014), but flying powers also affects range-size.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: GENOME SIZE VERSUS GEOGRAPHIC RANGE SIZE IN BIRDS
Review round: 1
Reviewer: 2

Output:
Basic reporting: In my view, the authors have to work better the argument about the bigger the genome, the better adapted an organism is to deal with multiple and more diverse environmental pressures. Put it another way, is there previous evidence that at least some sort of selection could lead to bigger genome sizes? They do not mention any empirical evidence in that direction.

Also, If they are stating the hypothesis of a positive correlation between genome size and geographic range for the first time, then they have to make it explicit. If not, then cite other works which have explicitly discussed that genome size co-varies with range size...if they are making this claim for the first time, then it is important to think about at least one  alternative hypothesis / outcome, such as genome regulation being more important to adaptation that genome size per se. I am sure the literature is full of discussions like that, but the authors did not mention then here.
Experimental design: It is worth mentioning that the topology of the tree upon which their best explanatory model was based (Fig. 1B), is very different from the "best" trees achieved with more powerful genomic datasets published recently (see Jarvis et al. 2014: 10.1126/science.1253451)....so, we know the topology of Fig. 1B is likely inaccurate, which raises the issue of whether the correlations found were spurious, particularly considering the very low explanatory power of the models selected, and the slight strength of the correlation observed.

I did not check how the topology of Fig. 1A, compares with that of Jarvis et al. 2014, but Jetz et al. phylogeny is indeed fraught with problems and cannot be compared with estimates based on more powerful genomic datasets.
Validity of the findings: The models with better explanatory power still accounted to a maximum of 17% of the total variation, which is still very little, particularly considering that this was achieved with the smallest dataset, which was the least representative for the entire Aves class. Again, this raises the issue of whether the correlations found were spurious, particularly considering the very low explanatory power of the models selected, and the slight strength of the correlation observed between genome size and geographic range.
Additional comments: I have made several detailed comments directly onto the text which I hope will be useful to the authors.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: GENOME SIZE VERSUS GEOGRAPHIC RANGE SIZE IN BIRDS
Review round: 1
Reviewer: 3

Output:
Basic reporting: The manuscript “Genome size and geographic range size in birds” by Grzywacz and Skórka addresses an interesting and potentially important question that links the genome to biogeography. I would like to start by disclosing that I would not have undertaken this study because I would have predicted that it would almost certainly yield a null result. Since unexpected results are intrinsically exciting, I think this paper could have a high impact on the field and prove to be influential. However, this excitement regarding the results is contingent on the results being correct; I am not (yet) convinced that this is the case.

I would like to break my review of the manuscript into two major sections:

1.	The nature of the question (and the authors framing of the question).

I have two reasons why I would have expected a null outcome from this study: 1) there is simply not that much variation in avian genome sizes; and 2) there is actually a lot of variation in avian karyotypes. In other words, when considered relative to most tetrapod groups almost all birds have the same amount of DNA per haploid genome but its organization into chromosomes varies quite a bit (in large part reflecting the large number of microchromosomes in birds). I expect the impact of genome size on whole organism phenotypes to be subtle (which, to be fair, is precisely what the authors find) so I would have expected the minimal variation in genome size to be swamped out by other factors. On the other hand, I would have expected to high degree of variation in chromosome number to simply lead to noise. This is actually why I find the study interesting!

With that said, the authors really do not acknowledge the limited variation in avian. genome sizes, something readily apparent from, for example, the Andrews et al. (2009) paper they cite (cf. Fig. 2 of Andrews et al., note that the scale of that figure is from 1.15 pg of DNA per haploid genome to 1.45 pg – i.e., not much variation). In the second paragraph, the authors cite a paper discussing genome size-phenotype linkages that highlights dinoflagellates (Hou and Lin 2009), a group that is known for wide variation in genome size and unusual chromosomal organization (relative to other eukaryotes). I am not opposed to citing Hou and Lin. Indeed, “unusual” taxa sometimes emphasize general patterns that may be much more subtle in less unusual taxa. But the authors should acknowledge the nature of the variation in genome size.

In practical terms, the authors should provide a figure similar to Fig. 2 of Andrews et al. (2009), updated with the new data they have incorporated. They should also include the chromosome number information they used and, ideally, this would be plotted on a large-scale phylogeny. An excellent example of such a figure can be found in Fig. 6 of Kretschmer et al. (2018). Although I don’t want to be too specific, I would not recommend a tree with all tips; I think it would be a more readable figure would collapse groups down to major lineages (e.g., to orders) and put a bar charts with error bars for the standard deviation (or interquartile range) or just numbers adjacent to the tips. This would give readers an idea of the scale of the variation in genome size within birds.

With all of that said, I’d like to applaud the authors for trying this. If true, I’m very interested in the effect they see. Indeed, the authors make the point that “…the explained variation is low, thus statistical significance does not necessary[sic] mean biological meaning” (their lines 198-199). Given the limited variation in genome size I would say that that finding any relationship at all – even a weak one – has the potential to be interesting and meaningful.

2.	Inadequate estimates of phylogeny used for comparative methods

Adequate phylogenetic controls are critical for this project, but the tree they use is poorly documented and very old. Specifically, they state that they use the Jetz et al. (2012) tree, but the Jetz tree is actually two different sets of trees, one based on constraints from Ericson et al. (2006) and one based on constraints from Hackett et al. (2008). It appears to me (based on the authors Fig. 1) that the authors used the Ericson constraint set. I strongly recommend that the authors use the Hackett tree set; this is what most users of the Jetz tree use. Moreover, the Hackett et al. (2008). tree was based on an analysis of 19 loci whereas the Ericson et al. (2006) tree was based on five loci (and the constraints Jetz used were actually based on one of the supporting figures in Ericson et al. that included a mere four loci). Regardless, it is critical that the authors are clear regarding the trees they used.

With all of that said, the Jetz tree is now showing its age. The analysis Jetz and colleagues used was constrained by the Ericson and Hackett “backbone” trees and by taxonomy. The use of taxonomy allowed Jetz et al. to include taxa for which there was no actual data, but that practice led to some very problematic results (e.g., Wang et al 2017). Moreover, there are parts of the Jetz topology for taxa with no data that have 100% support (Hosner et al. 2015; this probably reflects the difficulty of running the MCMC chain long enough to adequately sample the posterior distribution when so many taxa are included). I am not opposed to including analyses using the Jetz tree because it is still the only really easy to use avian megaphylogeny, though it should be used with caution.

I stated that the Jetz tree is the only really easy to use avian megaphylogeny, but there is now a “replacement” that isn’t that hard to use: the tree used by Ksepka et al. (2020). It is based on a reanalysis of the Burleigh et al. (2015) supermatrix with constraints from the Jarvis et al. (2014) tree, which reflects analyses of about 40 Mbp of aligned data and includes more than 10,000 loci. The tree is available as supporting data from the publication, although it may be a bit of a pain to use. Specifically, there is a dated version trimmed to a subset of taxa (supplementary file S5 of Ksepka et al.) and a version without rate smoothing that can be converted to an ultrametric form (supplementary file S4, which it has fossil calibrations readable by the program r8s [available from http://ceiba.biosci.arizona.edu/r8s/index.html] included in the file but the tree needs to be converted to an ultrametric format). I think that, at a minimum, the authors should see how many of their taxa overlap with the file S5 tree and conduct an analysis of their data using that tree (after trimming to their taxa). This is why I could see retaining analyses based on Jetz et al. – the authors could do an analysis using the Jetz tree and all taxa and then trim down to taxa that overlap with the Ksepka file S5 tree and do an analysis using fewer taxa and that tree (or course,  this would be irrelevant if all of their taxa are present in the Ksepka file S5 tree).

I wish there was a truly up to date avian megaphylogeny with dates out there; such a tree does not exist at this point. Unfortunately, the Jetz tree is definitely showing its age at this point and the Ericson backbone (14 years old at this point) is definitely problematic.

A final comment about presentation of trees: the authors clearly use an older taxonomy in their figure 1. They should update their ordinal names based on a modern taxonomy (i.e., the IOC World bird list; Gill et al. 2020). The use of the much older circumscription of e.g., Falconiformes serves to obscure the tree. Even in the early large-scale bird trees of Ericson et al. and Hackett et al. it was clear that Accipitriformes and Falconiformes sensu stricto (i.e., limiting extant Falconiformes to Falconidae) were not a clade. This is only amplified by more modern trees like Jarvis et al. Using Falconiformes in the old-fashioned manner obscures rather than illuminates.

And now one final comment about the comparative methods: the authors state that “…non-linear methods do not include phylogenetic relations among species” (lines 202-203) of their manuscript. This is not strictly true, Quader et al. (2004) discuss the issue of non-linearity. This paragraph is an excellent one, but it should be tweaked to incorporate Quader et al.  I think the bigger point that there is no easy way to incorporate non-linearity is true. On the other hand, I’m not convince that is the biggest limitation of available methods.

Some minor issues:

The authors generally use excellent English but there are a few glitches. For example, I used [sic] above and the authors should change “necessary” to “necessarily” in that case. I feel a little bad for even saying this because it is always hard to write in a language other than your native tongue and I would not be able to do anywhere as well as the authors if I were writing in a language other than English. However, the authors should be conscious regarding the issue of language and they might consider asking a native English-speaker colleague to help them with a read through. In that context, they begin some paragraphs with “Interestingly, …” I dislike this, since it does not highlight anything in particular.  After all, everything in a paper should make the bar of being interesting enough to include! Unless a sentence begins either with a contrasting or with an amplifying word/phrase (e.g., “In contrast,” “In fact,” etc.) it is probably best to avoid constructions similar to “Interestingly, …”. This is even more true for the beginnings of paragraphs.

Finally, I’m not sure what to make of the authors’ finding that “…body mass in the multivariate models and path analyses was negatively correlated with the range size” (their lines 183-184). I like that they point this out. Perhaps a little expansion of the reasons why this is not expected are warranted.

Disclosures:

A general policy for PeerJ is to make sure authors are aware that they should not be pushed into adding citations for papers written by reviewers or editors. I think this is a good policy. Therefore, I would like to disclose that I am a co-author on several papers cited below (e.g., Hackett et al., Jarvis et al., Ksepka et al.). In some cases, I think citation of those works is critical; for example, I think using the Ksepka et al. tree is really important. In other cases, my citation within this review was merely to illustrate a point (e.g., Hosner et al., Wang et al. illustrate issues with Jetz et al.) and I certainly don’t want to push for citations of my work beyond that which is appropriate given standard scientific ethics. In that context, I do not expect everything I cited in this review to be cited by the authors (moreover, some of the papers I cite are already cited by the authors).

I also want to say that like to sign reviews, regardless of whether they are positive or negative. I feel signing reviews helps the reviewer stay focused on providing constructive feedback. My overall evaluation of this manuscript is positive – I will say that “I want to believe” that the authors have found the relationship they say they have found. Of course, I also want them to convince me (and other readers!) that their results are robust. They’ve already convinced me that their results are interesting.

Edward L. Braun
Professor of Biology
University of Florida
Gainesville, FL USA

References for this review:

Andrews CB, Mackenzie SA, Gregory TR. 2009. Genome size and wing parameters in passerine birds. Proceedings of the Royal Society B. 276: 55–61 DOI:10.1098/rspb.2008.1012.

Burleigh, J. G., Kimball, R. T., & Braun, E. L. (2015). Building the avian tree of life using a large-scale, sparse supermatrix. Molecular phylogenetics and evolution, 84, 53-63.

Ericson, P. G. P., Anderson, C. L., Britton, T., Elzanowski, A., Johansson, U. S., Källersjö, M., et al. (2006). Diversification of Neoaves: integration of molecular sequence data and fossils. Biology letters, 2(4), 543-547.

Gill F, D Donsker & P Rasmussen (Eds). 2020. IOC World Bird List (v10.2). doi: 10.14344/IOC.ML.10.2. https://www.worldbirdnames.org/

Hackett, S. J., Kimball, R. T., Reddy, S., Bowie, R. C., Braun, E. L., Braun, M. J., et al. (2008). A phylogenomic study of birds reveals their evolutionary history. Science, 320(5884), 1763-1768.

Hosner, P. A., Braun, E. L., & Kimball, R. T. (2015). Land connectivity changes and global cooling shaped the colonization history and diversification of New World quail (Aves: Galliformes: Odontophoridae). Journal of Biogeography, 42(10), 1883-1895.

Hou Y, Lin S. 2009. Distinct Gene Number-Genome Size Relationships for Eukaryotes and Non- Eukaryotes: Gene Content Estimation for Dinoflagellate Genomes. PLoS ONE 4(9): e697 DOI:10.1371/journal.pone.0006978.

Jarvis, E. D., Mirarab, S., Aberer, A. J., Li, B., Houde, P., Li, C., et al. (2014). Whole-genome analyses resolve early branches in the tree of life of modern birds. Science, 346(6215), 1320-1331.

Jetz W, Thomas GH, Joy JB, Hartmann K, Mooers AO. 2012. The global diversity of birds in space and time. Nature 491: 444–448 DOI:10.1038/nature11631.

Ksepka, D. T., Balanoff, A. M., Smith, N. A., Bever, G. S., Bhullar, B. A. S., Bourdon, E., et al. Tempo and Pattern of Avian Brain Size Evolution. Current Biology.
volume 30, issue 11: 8 June 2020, Pages 2026-2036.e3

Kretschmer, R.; Ferguson-Smith, M.A.; De Oliveira, E.H.C. Karyotype Evolution in Birds: From Conventional Staining to Chromosome Painting. Genes 2018, 9, 181.

Quader, S., Isvaran, K., Hale, R. E., Miner, B. G., & Seavy, N. E. (2004). Nonlinear relationships and phylogenetically independent contrasts. Journal of Evolutionary Biology, 17(3), 709-715.

Wang, N., Kimball, R. T., Braun, E. L., Liang, B., & Zhang, Z. (2017). Ancestral range reconstruction of Galliformes: the effects of topology and taxon sampling. Journal of Biogeography, 44(1), 122-135.
Experimental design: As detailed in my complete review (the "basic reporting"), my primary concern with the experimental design is the use of a older phylogeny for their comparative methods.
Validity of the findings: As detailed in my complete review (the "basic reporting"), I think the findings could be valid but I feel the authors like to do a bit more to present a convincing case
Additional comments: No additional comments


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: GENOME SIZE VERSUS GEOGRAPHIC RANGE SIZE IN BIRDS
Review round: 2
Reviewer: 1

Output:
Basic reporting: The paper has been successfully revised following the first review. The work is well presented, in clear English, and has a clear focus and interesting results
Experimental design: By using two alternative sets of phylogenetic data and analysis a much more robust result has been obtained. The analyses are well done and I see no weak points.
Validity of the findings: The results, as presented in lines 214-237, are well supported and presented
Additional comments: The Authors have followed well the comments by reviewers, especially by taking into account other phylogenetic data (Ksepka et al.) and by pointing out various confounding factors and the undersampling of species with very localized distributions (lines 290-323). The MS is now much more robust and well formulated, and I have only a few minor extra points:
The aspect of how avian flight has affected genome size is mentioned on line 247, but it seems relevant also to mention in the Introduction (maybe after line 90), and in line 302) how avian flight has required massive changes (for light weight and increased energy efficiency) of all aspects, including the size of the genome. I assume that Zhang et al. 2014 would be a useful reference here.
Further details: In line 40, I think it would be better to change ‘may’ to ‘could’ (twice).
Lines 275-277: Gaston used data from a field guide from his first studies of range-size variation, but actually the data source was different in the study of waterfowl by Gaston & Blackburn. But even here, the range size data are extremely crude as they used presence-absence data in a grid of 10x10 geographical degrees.


Instruction: You are one of the reviewers, your task is to write a review for the article. You will be given the title of the article, the number of the round in which the article is located, and your order among the reviewers.

Input:
Title: GENOME SIZE VERSUS GEOGRAPHIC RANGE SIZE IN BIRDS
Review round: 2
Reviewer: 2

Output:
Basic reporting: All standards met.
Experimental design: All standards met.
Validity of the findings: All standards met.
Additional comments: The authors have made tremendous progress with this newer version of the manuscript and I am happy with what they have accomplished at this point: a somewhat daring, yet, sober (i.e., acknowledging all or most of the study´s limitations) analysis of the correlation between genome and ranges sizes, based on the best available evidence published so far. With that said, it is quite fortunate (or not!) that a recently published study based on 363 whole genomes covering from 92.4% of bird families (https://doi.org/10.1038/s41586-020-2873-9) offers quite a window of opportunity to be used in this paper as well. Interestingly, many of the shortcomings carefully addressed by the authors in this second version are due to problems that could be circumvented if this newer phylogeny had been used. For instance overall taxonomic coverage, as well as that of tropical taxa, is much better now at the genomic level. So,while I have no doubt that even a separate more "controlled" analysis focused just on this new tree would be a tremendous addition to this paper (while keeping the current presented results), making it probably even more cited, I realized that these analyses could potentially never end, given the wealth of data that is becoming available at a faster pace now (see also, this mind-boggling phylogenomic study focused at birds as well: https://academic.oup.com/mbe/advance-article/doi/10.1093/molbev/msaa191/5891114). So, while I am recommending acceptance as is, I would strongly recommend them to make an effort to include at least the whole genome more complete tree that just came out. I realize it is a lot of additional work, but in my view it would just add some more value to their contribution.