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: ECOLOGICAL STOICHIOMETRIC CHARACTERISTICS AND INFLUENCING FACTORS OF CARBON, NITROGEN, AND PHOSPHORUS IN THE LEAVES OF SOPHORA ALOPECUROIDES L. IN THE YILI RIVER VALLEY, XINJIANG
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript #51868 submitted by Zhang and collaborators to Peer J is an original primary research contribution with a well-defined and relevant research question, with some not-so-well defined background and justifications to conduct the study.
The introduction needs to have a better background for the hypotheses. There are no assumptions in the introduction. In addition, the research significance of this article is not well explained. I am afraid that the introduction is not enough to make readers feel the value of research. In short, the introduction is poorly written and needs to be revised.
I am afraid the English used throughout was not professional. There are some grammatical and structural mistakes in the manuscript and I am not able to highlight/revise all of them. So, please revise the manuscript and improve it considerably in terms of grammatical and structural mistakes.
Experimental design: The experimental design appears to have followed a rigorous standard, I do not any concerns regarding the data collection.
Validity of the findings: All underlying data have been provided; they are robust, statistically sound.
But the discussion is confusing because it is based on losing hypotheses. Once the hypotheses are better formulated, the discussion should be easier to write.
Additional comments: 1.	For the study area map, please add the standard China map (complete borders and islands in the South China Sea). Many readers may not know the geographical location of Xinjiang.
2.	The significance P-value in the article is the same as the abbreviation P for phosphorus. To avoid confusion, please revise the whole research. Such as Fig. 2.
3.	How do the selected samples prove their representativeness? Please elaborate. It is necessary, otherwise, the conclusions cannot be supported.
4.	What are the criteria for dividing the four types of plots? How are plant samples collected? The number of sample replicates? These are not explained clearly.
5.	The experimental method is not clear. Please explain the test methods of C, N, P carefully, this is a very important step. For example, what instrument is used for N and P tests?
6.	L114, “0~10、10~20、20~30”, Is there a "、" in English? There are too many such small mistakes, please carefully modify the full text.
7.	L164, The unit of "470.09, 32.71," must be clearly written, don’t be lazy
8.	L217, Have you also considered the K content of the soil? Why is this indicator not seen elsewhere in the article?
9.	L268, I disagree with the conclusion of the sentence "The stoichiometric ratios of C, N and P in leaves of plants are relatively stable". C is relatively stable in plants, but N and P are not. Please read the relevant literature carefully.
10.	Discussion writing is really bad. The thoughts of the discussion were too confusing, and the logic of the discussion was very unclear.
11.	The local stoichiometric characteristics are usually compared with the global or domestic mean, which seems not mentioned in the study. For the discussion, didn't the first thing to compare Sophora alopecuroides with other research results?
12.	Fig. 2 needs to be adjusted, the image resolution is low and unsightly; Table 3 does not indicate the level of significance; Fig. 4 Different font sizes.
In brief, the biggest problem with this article is language and logic. In addition, the introduction of the article and experimental methods are incomplete, and the discussion is even more unclear. Therefore, this article needs to be revised very seriously and rigorously. Otherwise, the article will be difficult to publish in the current state.


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: ECOLOGICAL STOICHIOMETRIC CHARACTERISTICS AND INFLUENCING FACTORS OF CARBON, NITROGEN, AND PHOSPHORUS IN THE LEAVES OF SOPHORA ALOPECUROIDES L. IN THE YILI RIVER VALLEY, XINJIANG
Review round: 1
Reviewer: 2

Output:
Basic reporting: I  recommend the authors to have a second look at the English editing of the manuscript.
Formatting of citations in text were incorrect.
Experimental design: More details should be added in the text
Validity of the findings: Part of conclusions looks like results, and draws no general conclusions.
Additional comments: 1.Title:Ecological stoichiometry characteristics of carbon, nitrogen and phosphorus in leaves of Sophora alopecuroides in the Yili River Valley, Xinjiang
2.Abstract: authors should tell us what are the scientific issues in backgrounds,why? rather than "in order to....."
3.N and P contents should be concertrations
4.Authors must follow the rules of Latin names for plants in text.
5.The presentation of introduction is also not well-structured. why did authors conduct this experiment? what are the shortages of previously study? Authos should focus on the scientific issues. the current version is still insufficient and the logic is vague.
6.Three-layer soil samples were collected,Why? Where dose the majority of root distribution of plant? how the data of soil physical and chemical properties were used in the statistical analysis?
7.Sophora alopecuroides is a leguminous plants, nitrogen-fixing should be discussed and explain.
8.four habitats in the text are huge difference, such as: temperture, precipitation,fertilization in farmland,authors should explain in the manuscript.
9 Formatting of citations in text were incorrect.
10.Part of conclusions looks like results, and draws no general conclusions.
11.I  recommend the authors to have a second look at the English editing of 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: ECOLOGICAL STOICHIOMETRIC CHARACTERISTICS AND INFLUENCING FACTORS OF CARBON, NITROGEN, AND PHOSPHORUS IN THE LEAVES OF SOPHORA ALOPECUROIDES L. IN THE YILI RIVER VALLEY, XINJIANG
Review round: 1
Reviewer: 3

Output:
Basic reporting: no comment
Experimental design: The experimental design is problematic.
Validity of the findings: no comment
Additional comments: In recent decades, the C, N, P ecological stoichiometry characteristics of plant leaves at different time scales and spatial scales have been well studied. The authors study the differences in ecological stoichiometry characteristics of leaf carbon, nitrogen and phosphorus of Sophora alopecuroides in four habitats in the Yili River Valley and its influencing factors. This research is valuable, but there are many problems. The English presentation also needs careful polishing.

1. In the Abstract, the abbreviations of C, N, and P should be given in L29. Across the text, the Latin name of Sophora alopecuroides was not used standardizedly.
2. L46, that studies the changes rule? changes?
3. The content of influencing factors of plant leaf stoichiometry in the Introduction is too simple and should be described separately, and you should focus on your research object and focus on the discussion.
4. L72, what’s F resistance?
5. L83, carbon, nitrogen and phosphorus or C, N, and P?
6. A clear scientific question or hypothesis needs to be presented in the Introduction.
7. L93, 80°09′E—84°56′, 42°14′N—44°50′N？
8. L100, ℃?  Or °C?
9. L114-115, a total of 12 soil samples and 12 plant samples was collected in 4 plots. Only 12 samples? It’s very small for stoichiometric analysis. And your repeat is only three, also small. In total, your experimental design is problematic.
10. L115-116, Leaves were dried at 105 OC for 24 h. Generally, the temperature ranged from 60-75 °C. Your temperature is too high.
11. L156. what is LGA?
12. L164, Yili River Valley wetland, wetland?
13. L213, factors of soil physical and chemical or soil physical-chemical factors?
14. L234-242, these explanations for CCA should be placed in “Statistical analysis”.
15. L274, table 2? Table 2?
16. Sophora alopecuroides is a nitrogen fixing plant species, so it’s generally N-limited; however, it might be also not P-limited, due to it’s higher leaf P content (mean=1.53 mg/g) than 1.0 mg/g. Therefore, the rule for judging restrictive elements, i.e., 14 or 16, is not effective for nitrogen fixing plants to a large extent.
17. The quality of Figure 1 is poor.
18. In Table 2, are you sure the differences of leaf P among four habitats are right?
19. Please also check the multiple comparison results in Table 3.
20. Check all Tables and Figures to avoid errors.


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: GENE EXPRESSION DURING THP-1 DIFFERENTIATION IS INFLUENCED BY VITAMIN D3 AND NOT VIBRATIONAL MECHANOSTIMULATION
Review round: 1
Reviewer: 1

Output:
Basic reporting: This is an interesting but very complicated manuscript. It contained a great deal of information derived from an extensive set of experiments using immortalized cells and the authors tried very hard to organize it in a way that is readable, digestible, and relevant. There are 8 experimental variables (adherent + suspension) x (vibration + vitamin D + vibration and vitamin D + control) with 12 outcome measures belonging to three major categories: monocyte-macrophage differentiation, antigen presentation, and mechano-sensors. Together, there are 96 sets of results. Despite their efforts, this paper reports, and I quote Theodosius Dobzhansky, "a pile of sundry facts -- some of them interesting or curious but making no meaningful picture as a whole".
Experimental design: I do not like reductive experimental designs. Taken out of the local environment, with no supporting extracellular matrix and other cells, these in vitro, ex vivo studies cannot duplicate the complex adaptive system that is present in the tissues or organs. This study uses spontaneously immortalized cell line, THP-1, from a pediatric patient with monocytic leukemia. While this cell line has been extensively used in many laboratories, it is not the same as peripheral blood monocytes and certainly different from the precursors of yolk sac-derived tissue resident macrophages.

Mechanical perturbations are important as cells exist in a mechanical environment and are subjected to tensile, compressive, and shear forces, including, and especially, the tissue resident macrophages. The selection of cyclic loading is good as majority of our daily activities generate stresses that are not static or even quasi-static. However, the selection of 30 to 60 nm amplitude and 1000 Hz frequency demands biological relevance. What kind of daily activities can generate that kind of continuous "nano-kicks" for 3 days?

Regarding the calibration using laser interferometer, was this done within the incubator? How was the vibration administered? That is was it vertical (up-and-down) or horizontal (side-to-side)? Since the authors wish to find out the effect of mechanical forces on the THP-1 cells, what was the force profile with this vibration? How much cellular deformation can occur? a 5 micron (10E-5 m) monocyte with 50 nm (10E-8) deformation give a nominal maximum cellular strain in the 0.001 range, likely much less. What evidence can the authors provide to support that such strain range is physiologically relevant? Compounding the problem is the liquid medium. How much damping can take place? How does the viscosity of the fluid affect the force transmission?

Lastly, it would be interesting to check what cells become adherent out of these immortalized cells which are more like peripheral blood monocytes? How many remain in suspension and how many attached to the plate?
Validity of the findings: I like how the authors presented this set of experimental data, and I believe they are valid. It is the higher level synthesis that is missing. I encourage the authors to focus on the fact that vitamin D3 is the much stronger stimulus than this very artificial vibration protocol. These are largely non-adherent peripheral blood monocytes and as such, they probably  normally experience different kind of mechanical loading such as fluid shear, rather than tensile stress or bi-axial compression.
Additional comments: I learned a lot from reviewing your paper, for that I am grateful. One of my mentors from decades ago would ask me, after I proposed my experiments, "Let's fast forward a year or two and say that you have done these experiments and found this and that. So what? Then what?"

I would very much appreciate if the authors would answer the so what and then what.


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: GENE EXPRESSION DURING THP-1 DIFFERENTIATION IS INFLUENCED BY VITAMIN D3 AND NOT VIBRATIONAL MECHANOSTIMULATION
Review round: 1
Reviewer: 2

Output:
Basic reporting: The figure legend for Figure 4 should be revised. I believe this refers to adherent cell data yet suspension is written.
Figure legend for Table 2 should be revised. I believe this is comparison of stimulated adherent THP-1 cells vs unstimulated adherent cells

Improve English language and sentence structure throughout manuscript e.g., repetitive use of the phrase “the vitamin D3” can simply be replaced with “vitamin D3” for example line 288 “The vitamin D3 downregulated..” instead “Vitamin D3 downregulated..” may be used.

Improve sentence structure in lines 609-610 “This study had limitation because of the technology” – e.g., This study was limited by the technology which…
line 73-74; improve sentence structure. Maybe break the sentence in two
Experimental design: The methodology should be clearer. For example, over the 72-hour period, how was vitamin D3 administered? Is it given once in the medium and left for 3 days? Is it replenished after each day?

Did the authors look at PIEZO2 in THP-1 cells? It may be informative to include the sibling of PIEZO1 for comparative purposes. RT-qPCR analysis of this mechanosensor may be included across vitamin D3 treated, 1000Hz and combined conditions.
 Alternatively, the reason why it was not analysed should be presented e.g., future studies will do so; or based on previous data PIEZO2 is not highly expressed in THP1-cells (Reference).

Using only one vibrational parameter (1000Hz) may limit potential mechanosensory responses. For example, previous studies (Koizumi et al, 2014; Functional role for Piezo1 in stretch-evoked Ca2+influx and ATP release in urothelial cell cultures. J Biol Chem 289 (23):16565-16575) have shown the presence of a stretch stimulated threshold that must be attained before mechanosensor mediated changes in cell responses (ATP efflux) are observed. Similarly, the current study may yet to uncover a potential vibrational range that alters gene expression differently than reported. Is there a spectrum of gene expression changes that occurs by changing the range of vibration (Hz, time, cyclical stimulation?). Is there a range in which macrophage differentiation is promoted? Furthermore, the nature of mechanical stimuli (vibration vs stretch vs hydrostatic pressure changes) may alter THP-1 cell`s gene expression. The 1000Hz is a standard range used in THP-1 cells thus it is understandable why the current report did not alter it. However, commenting on these issues may help the reader understand the dynamic range of cell responses that have yet to be elucidated. For the current scope of this report additional experiments are not essential (specially as the authors explained the limitation; lines 609-610), however acknowledgement or dismissal of such potential changes using previously published data or inference from data presented by the study should be presented to the reader.
Validity of the findings: Conclusion should be added to abstract. Currently the abstract abruptly ends on Results. A clear “conclusion:” with a sentence or two to summarise the main findings should be included.

The authors summarise the data elegantly using tables (Tables 1, 2 and 3); however, the main text makes little to no reference of this. Guiding readers to use these tables as comparisons are made may help with the flow of data presentation and discussion points.
Additional comments: In this study Simakou et al, set out to examine the effects of vitamin D3 administration, vibrational stimulation and their combined effect on THP-1 cells with a focus on their differentiation potential into macrophages. A series of RT-qPCRs were conducted to measure the various gene expression changes associated in response to stimuli (vitamin D3 vs 1000Hz or both) and cell condition (suspended vs adherent). The data presented shows novelty and proficiency that should make this manuscript eligible for publication in PeerJ.

The data is efficiently summarised and presented well in both graph and table format which allows clear comparison and interpretation of the data. The authors show novel expression patterns such as PKD2 upregulation with vitamin D3 administration. With further testing in other cell types, this may serve as unique signature of vitamin D3 stimulation.
A comment on potential limitations of THP-1 immortalised cell line vs primary cell lines may be warranted in order to acknowledge the need for further research and confirm findings in the future. In order to enhance readability, the authors should significantly improve sentence structure and avoid grammatical errors. This will avoid breaking the flow of the text and hasten understanding of key points presented.

It would be informative to compare basal expression levels of key genes in suspended and adherent conditions. For example, do adherent cells express comparably higher levels of PIEZO1 than unstimulated suspended cells? Are the levels of CD14 and CD36 higher in adherent vs suspended cells?

Perhaps beyond the scope of the current study, but analysis of protein expression levels of key macrophage differentiation markers (CD14, CD36 etc.) through western blot may strengthen the claims made. The authors could state the need to confirm their key findings at the protein level to corroborate their RT-qPCR 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: GENE EXPRESSION DURING THP-1 DIFFERENTIATION IS INFLUENCED BY VITAMIN D3 AND NOT VIBRATIONAL MECHANOSTIMULATION
Review round: 1
Reviewer: 3

Output:
Basic reporting: The manuscript is well written with sufficient literature support.
The study is an observational study designed to investigate effects of VitD and/or vibration treatment on monocyte to macrophage differentiation.  The rationale of the investigation was however unclear; or could have improved by proposing some potential clinical application or translational value.
The discussion reads a bit long on the background support compared to the novel findings.  It would be easier to the reader if the focus of each paragraph is shifted more towards the novel findings.
Experimental design: The experimental design is straight forwards with sufficient description for replication.  The results are presented with good clarity.
Validity of the findings: The results are well presented and support the conclusions being made.
Additional comments: There are a large number of erroneous table/figure references in the results.  Please revisit to fix them.


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: GENE EXPRESSION DURING THP-1 DIFFERENTIATION IS INFLUENCED BY VITAMIN D3 AND NOT VIBRATIONAL MECHANOSTIMULATION
Review round: 2
Reviewer: 1

Output:
Basic reporting: The manuscript shows drastic improvements in legibility and flow of data presentation. References to tables have been added which improve the reader's ability to follow the claims made. Structurally the manuscript is well organised and self-contained in its discussion in light of the results presented.
Experimental design: Research questions are well designed and addressed efficiently in the experiments chosen and data analysis employed. Many of my original concerns have been addressed and supplementary figures added that further support this manuscript.
Methods are explained and previous issues have been addressed.
Validity of the findings: As stated previously, data presented in this manuscript shows novelty and proficiency. The role of Vitamin D3, vibrational stimulation and THP-1 differential potential is addressed well in this report. The findings should have cross-field relevancy in other cell types.
Conclusion is now referenced in abstract and the authors discuss their findings well. By including supplementary figure 2 the authors elegantly address previous concerns. Overall, the findings presented here and corrections applied should warrant the current manuscript novel and impactful for a wider readership.
Additional comments: An annotated pdf is not necessary. Thank you for addressing the concerns raised. The manuscript presents data to  the field of mechanosensation that will intrigue others.


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: GENE EXPRESSION DURING THP-1 DIFFERENTIATION IS INFLUENCED BY VITAMIN D3 AND NOT VIBRATIONAL MECHANOSTIMULATION
Review round: 2
Reviewer: 2

Output:
Basic reporting: Revision has addressed previous question and I have no further comment.
Experimental design: I have no further comment.
Validity of the findings: I have no further comment.
Additional comments: I have no further comment to the authors for improvement.


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: AMBULACRARIAN INSULIN-RELATED PEPTIDES AND THEIR PUTATIVE RECEPTORS SUGGEST HOW INSULIN AND SIMILAR PEPTIDES MAY HAVE EVOLVED FROM INSULIN-LIKE GROWTH FACTOR
Review round: 1
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: This paper reports of a very well done analysis on the evolution of insulin-related peptides in ambulacraria. The paper is well written, the scientific context well introduced and the results are clearly presented and discussed.

I have only a few questions and suggestions that could help non expert readers to better understand the study:

The author uses for good reason "sequence similarity trees" approach analysis. The rationale for that is reported in the Materials and methods section where a very brief explanation of the approach is also reported. To help the non expert reader, I would give some general explanation in the Results section instead, and may be give more technical details in the Methods. For instance, it is stated that: "and Fasttree (Price, Dehal & Arkin, 2010) to construct trees and estimate probabilities", but no further details are given.

Similarly, there is quite a discussion in the sub sections  "Nomenclature",  "Sequence Analysis" and "Expression" of the Methods section, part of which would be more useful in the results, especially to orient the non expert reader who is only familiar with terms such insulin, IGF, etc.

Two other technical points are not clear to me and should be better specified:

1.  where the genomic information for the following species was taken from?
- Anneissia japonica
- Holothuria glaberrima
- Patiria miniata
- Ophiothrix spiculata
I would assume from Echinobase, but I did not find this reference. Have I have missed it?

2. in the file "peerj-60199-Suppl.Spreadsheet1" there are several genes with Genbank identifier: NEW in red; what does it mean? are those sequences for which an accession number will be provider after publication?


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: AMBULACRARIAN INSULIN-RELATED PEPTIDES AND THEIR PUTATIVE RECEPTORS SUGGEST HOW INSULIN AND SIMILAR PEPTIDES MAY HAVE EVOLVED FROM INSULIN-LIKE GROWTH FACTOR
Review round: 1
Reviewer: 2

Output:
Basic reporting: The peptide family described is quite complicated and the members have confusing names that make them easily mixed up. The terms "related" and "like" are not easily distinguishable. This makes it difficult to describe the situation clearly for non-experts. See my comments below. This is by no means the authors "fault", this is the way the peptides' names have gradually been assigned, making them quite challenging to describe. An introductory figure or table should help clarify the relationships. Likewise, a summarizing figure at the end can help describe the evolutionary scenario that the author proposes. I expect such clarifications would make the description easier to grasp for those who are not experts on this peptide and receptor family. I do not demand to see these clarifying additions, hence I have ticke "minor revision" rather than major, but I am prepared to do so if requested.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: The terminology for this peptide family is quite confusing for non-experts (as are several other peptide families). It is difficult to distinguish the peptides that are described as IGF-like/related versus those that are called insulin-like, especially as the latter are considered to have emerged from the former by gene duplication. On line 65, this entire family of peptides is called "insulin-related peptides (irps)". So the family of insulin-related peptides started with an IGF-like that subsequently gave rise to insulin-like, then the "neuroendocrine insect irps" arose from their IGF-like. So it seems like irp is both the overarching name and a subset of IGF-like called neuroendocrine irps. Is it possible to describe this with a figure in the introduction? The author writes about the naming confusion on lines 99-108, especially regarding the receptors.

Another thing that would be very nice to have is a duplication scheme that summerizes the scenario that the authors favours.

24: It is confusing to add that the first of the three hormones is IGF-like when it has just been said that all three are IGF-related. Then on line 26 it is said that the peptides are insulin-like. One step in the evolution is missing, namely how two peptides went form being IGF-like to become insulin-like. Could it be that "Arthropod IGF" is the oldest and was duplicated, and this duplicate shrunk and became "insulin-like", and then evolved preference for a GPCR, and then this insulin-like gene was duplicated to give the third member. The two latter are what we call dilp7 and Arthropod gonadulin. Indeed, on line 88 the author writes about "two successive local gene duplications". The likely order of events gradually becomes clear as one reads the manuscript, but it is necessary that the abstract can summarize this in a reasonably intelligible way.

25: Why suggest that the ancestor of the three peptides used both types of receptors? Isn't it equally possible that one of the duplicates evolved preference for a second, and different, type of receptor? dilp7 is said on lines 76-77 to act on an RTK, in addition to its GPCR, so this peptide could represent the transition stage from one to the other and not necessarily an ancestral state of dual receptor preference. Again, the reasoning becauses clearer on reading the entire manuscript, but if this shall be presented in the abstract, it should ideally be well motivated here.

39-40 The author writes "If IGF were to act through both a GPCR and an RTK it would suggest that GSS acts on only one…". I am afraid I do not follow the reasoning here. If IGF worked on both, and gave rise to GSS, it would be expected that GSS too would initially act on both types of receptors, wouldn't it? But referring to my comment above, couldn't it be that IGF worked only through an RTK already before it gave rise to GSS, then GSS too would act on the RTK.

Minor comments

In a couple of places the author writes "genes coding peptides" or similary. It should "encoding peptides" or "coding for peptides", as indeed written correctly on line 263. Please scan the manuscript for this.

61 It is advisable to delete "closely" as some ligands bind to receptor subtypes that arose by duplication some 500 million years ago or more and have diverged considerably. I suggest that it is emphasized that this discussion concerns peptide ligands by inserting the word "peptide" before ligand(s) in this paragraph.

66 Correct the spelling of "coupled"

72 The sentence says "One of these" which seems to refer to dilp7 in the preceding sentence, but it should be "One of the irps".

86-87 The part after the comma is confusing. It does not necessarily have anything to do with the fact that the IGF gene is part of a local triplet.

112 Should be: number of Amulacraria species.

128 For those unfamiliar with these species, please write: "transcriptome data from the three crinoid species Antedon…."

154 should be "orthologous"

178 shold be "cleave any secreted protein with appropriate cleavage site".

206-211 This peptide organization would be good to illustrate with a figure.

216 "other echinoderms" should be "other ambulacrarians"

223 should be "cleavage sites"

258 it looks grammatically odd to see "are single copy", I suggest "is a single copy gene", this cannot be misunderstood.

260 Why write genus name for one and species name for the other?

308 should be "shows"
310 should be "where"

316-317: This sounds like a circular argument. Of course the tree based on sequence alignment and similarity will reflect the way they "look". Please rephrase.

324: should be NCBI

335: should be "multiple"

338-9: should be "from time to time"

339: "assembled constructs" is probably not the correct term here, I believe "assembled sequeces" would be more suitable.

340: "both" makes this sentence a bit difficult, please rephrase.

343: should be and adverb: "particularly"

352: "subbranches" should be followed by a comma.

357: "it is interesting to see" comments on something that cannot be seen in this clade in Fig. 9. Please describe differently or refer to a supplementary tree that contains this information.

363: What is "The second branch"? Please describe in an unambiguous way.

371: "are" should be "is"

372: "it are the glycoprotein hormone and relaxin receptors that are identified" can be corrected to "the glycoprotein hormone and relaxin receptors are identified"

373: add comma after GRL101.

386: it is more common to say "echinoderms"

391: "This" has unclear refrence and the sentence is dangerously close to circular reasoning: "pressure is weak…because it is poorly maintained" - please rephrase.

394: should be "suggest"

427: "with" should be "by"

430: As the sentence says "a second insulin RTK gene", it will have to be: "none was found".

431: "neither" does not fit here, better to write "were unsuccessful"

435-436: why can't binding to a second receptor have arisen later in evolution? See my comments at the beginning of my referee report.

463: "it were the…that were identified" can be simplified to: "the…receptors were identified…"

492: "evolved" can be replaced by "diverged"

Fig. 6. It would be very useful to have lines showing the organization of the prepro-peptides in relation to these schematic mRNA/cDNA outlines, and not only the Cys positions. Probably is would suffice with the prepropeptide outlines for the top and bottom sequences.

Fig. S11: Is TM1 correctly marked? I am not totally familiar with LRR TM regions but I think it might be shifted right-wards. All six sequences have an R in the TM1 region and four of them have a K a few positions further upstream. This would be a bit unusual to have within a TM region, but the LRR:s are a bit unusual so it may be correct. Please double check the boundaries.


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: AMBULACRARIAN INSULIN-RELATED PEPTIDES AND THEIR PUTATIVE RECEPTORS SUGGEST HOW INSULIN AND SIMILAR PEPTIDES MAY HAVE EVOLVED FROM INSULIN-LIKE GROWTH FACTOR
Review round: 2
Reviewer: 1

Output:
Basic reporting: N/A
Experimental design: N/A
Validity of the findings: N/A
Additional comments: I am fully satisfied of the explanation/corrections proposed by the author. In my former review I didn't raise a problem of nomenclature for the insulin-related growth factors, because I didn't have a solution to propose, but I see that, thanks to the suggestion of the other reviewer, the author found a solution which makes the manuscript more easily readable by both specialist and not specialist readers.
Overall I think that the manuscript improved considerably and represent a significant contribution to the field.


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: AMBULACRARIAN INSULIN-RELATED PEPTIDES AND THEIR PUTATIVE RECEPTORS SUGGEST HOW INSULIN AND SIMILAR PEPTIDES MAY HAVE EVOLVED FROM INSULIN-LIKE GROWTH FACTOR
Review round: 2
Reviewer: 2

Output:
Basic reporting: No comment.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: I feel that the renaming of the peptide groups now should make reading easier.

357: add missing 'o' in evolutionary

Fig. 1
This figure should help many readers. I just wonder if the D-domain has to start at the last residue of insulin, a single N. In insulin, this position is considered to be part of the A-chain. It would be difficult to argue that a single N constitutes a domain. But maybe there is a good reason to draw the boundary here that I am unaware of.

Previous Fig. 6, new Fig. 7
I had asked for prepropeptide outline to be displayed or marked, and the author wrote that this had been done, but the figure still looks the same to me. This is not a demand from me, I just thought it would be helpful.


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: CONVERSION OF ALPINE PASTURELAND TO ARTIFICIAL GRASSLAND ALTERED CO2 AND N2O EMISSIONS BY DECREASING C AND N IN DIFFERENT SOIL AGGREGATES
Review round: 1
Reviewer: 1

Output:
Basic reporting: Findings of this study provide a valuable insight into understanding the influences of land use change on soil aggregate structure, SOC and TN, and soil GHGs emissions in an alpine grassland ecosystem.
Experimental design: Methodology is good and clearly presented.
Validity of the findings: Result section in the abstract should be improved. Author wrote basic and general results, however results whit actual data would be good. moreover, the abstract should be reduced.
overall results are presented very well.
Additional comments: The manuscript entitled “Deterioration of aggregate structure and decrease of soil organic C and N caused by alpine pastureland converted to artificial grassland altered CO2 and N2O emissions pattern” is interesting and suitable for publication but several mistakes in the manuscript should be corrected before acceptance.
Specific comments
•	Authors should carefully recheck the manuscript for the typos and also write the meaning of all acronyms because there are some which are not presented.
•	All the tables and figures are clearly presented but authors must check all the figure pixels again. Some of them are not very clear.
•	I suggest that authors check the bibliographic references in the main text and revise the format of documents to meet the requirement of the Journal.
•	In abstract, please add more information about methods of the experiment.
•	In abstract, please don't write the same conclusion lines as they were written in the main conclusion section.
•	In abstract, line 36-41; it’s a general result, author should present data and justify it.
•	Introduction, Line 76: “use GHGs abbreviation”.
•	Introduction, Line 83: correct the spell of different.
•	Methodology Line 85: what is TP stands for???
•	Discussion should also be improved critically. Don't write the results in the discussion section, discus them with strong logic.
•	Please recheck whole manuscript very carefully, particularly English language and remove all the typo mistakes.
•	Some of the papers that would be interesting to compare and cite in this current study are, for example.

Shakoor, A., Shakoor, S., Rehman, A., Ashraf, F., Abdullah, M., Shahzad, S. M., ... & Altaf, M. A. (2020). Effect of animal manure, crop type, climate zone, and soil attributes on greenhouse gas emissions from agricultural soils—A global meta-analysis. Journal of Cleaner Production, 124019.

Shakoor, Awais, Muhammad Shahbaz, Taimoor Hassan Farooq, Najam E. Sahar, Sher Muhammad Shahzad, Muhammad Mohsin Altaf, and Muhammad Ashraf. "A global meta-analysis of greenhouse gases emission and crop yield under no-tillage as compared to conventional tillage." Science of The Total Environment 750 (2021): 142299


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: CONVERSION OF ALPINE PASTURELAND TO ARTIFICIAL GRASSLAND ALTERED CO2 AND N2O EMISSIONS BY DECREASING C AND N IN DIFFERENT SOIL AGGREGATES
Review round: 1
Reviewer: 2

Output:
Basic reporting: In this paper the authors have made an attempt to study the effect of conversion of alpine pasture land to artificial grassland on distribution of soil aggregates and aggregate associated C and N and also emissions of CO2 and N2O from these aggregates. This is an interesting study. This paper may be accepted for publication with major revisions as suggested in Specific comments and also on the body of the text.
Experimental design: Results
Page 16 Line 233: Percentage of SOC lost CO2 emissions may be computed.

Page 16 Line 256: Percentage of soil TN lost through N2O emissions may be computed. The data on cumulative N2O fluxes may be presented.

Discussion
Page 20 Line 328: C/N ratio in different aggregate fractions may be computed and correlated with the CO2 and N2O emissions. Because C/N ratio indicates the relative lability of SOC to microbial decomposition
Page 20 Line 334-336: Estimated carbon input under different treatments may be computed to support this hypothesis.

Table 1: SOC and TN of bulk soil may be presented.
Table 2: The number of data points used in correlation may be indicated. Correlation between CO2 and N2O emission with C/N ratio may be attempted.

Figure 3: The peak value of CO2  emissions may be discussed.
Figure 4: The peak value of N2O emissions may be discussed.
Validity of the findings: It is an interesting study and the conclusions are well stated, linked to original research questions and limited to supporting results.
Additional comments: Please address the Specific comments and also the comments made on the body of the 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: CONVERSION OF ALPINE PASTURELAND TO ARTIFICIAL GRASSLAND ALTERED CO2 AND N2O EMISSIONS BY DECREASING C AND N IN DIFFERENT SOIL AGGREGATES
Review round: 2
Reviewer: 1

Output:
Basic reporting: Clear
Experimental design: Clear
Validity of the findings: Good
Additional comments: Authors have answered all the concerns. Paper can be accepted for publication.


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: SCALING THE EFFECTS OF OCEAN ACIDIFICATION ON CORAL GROWTH AND CORAL–CORAL COMPETITION ON CORAL COMMUNITY RECOVERY
Review round: 1
Reviewer: 1

Output:
Basic reporting: The study tests the possibility to apply model simulation and projection for coral recovery under ocean acidification, which is very meaningful in the future coral conservation. While I still have some concerns listed below.

Line 154-161, authors claimed to use Evensen & Edmunds (2016) results for model fitting. In the previous results, both vertical and horizontal growth were measured, so which growth rates are applied to the model in this study? Please be more specific.

Line 174-176, the competitive hierarchies were determined according to previous research, while the three researches were conducted in different regions (Taiwan, Israel and France). It’s reasonable that coral communities may maintain different competition hierarchies in different places. Although I think the hierarchy is acceptable for the present model, more evidence is needed if you’re trying to clarify there is consistency in coral competition across the regions or please find researches conducted in similar regions to support this statement.

Fig.2(a) The experiments were conducted and published in previous study and not the main purpose of the present one. It’d be better to move it to supplementary material.
Experimental design: Line 180-182 I can easily understand the meaning of equation 1. While for equation 2 in Table 1, where does it come from? Do you have any reference or just build it from your data trend?

Line 248-251, all the analysis code, including this model simulation, which is a major part of the study, should be provided in the supplementary material.
Validity of the findings: Line 328:  The model assumed the growth rate of Porites was not affected by OA, this assumption is not justified. There are evidences that show a significant decline in growth and calcification of Porites under OA.

Line 377-382: Macroalgae is the main competitor in most disturbed reef, thus exclude this from the model could prevent its general application.
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: SCALING THE EFFECTS OF OCEAN ACIDIFICATION ON CORAL GROWTH AND CORAL–CORAL COMPETITION ON CORAL COMMUNITY RECOVERY
Review round: 1
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: Evensen et al addressed a very important while largely neglected topic in this area and combined empirical data and modelling approach to scale up the OA effects on the dynamics and recovery of coral communities. They found that when considering coral-coral competition in the model, the effect of OA on coral recovery was stronger. Generally, this is a timely and excellent study. The writing and logic flow is very good for easy reading. I only have one major concern and several specific comments
Although the authors did state that Macroalgae was not considered a major competitor for space in the present model and algal cover remain <5% in Moorea. According to figure 1, many fleshy macroalgae adjacent to corals can be clearly seen from the quadrat. How much do the authors think that this may influence your modelling approach and results? I think that it is better to relook at all the photoquadrats and see how many corals are surrounded by fleshy algae?
Specific comments:
Line 130-131 “colonies of all four genera were assumed to extend linearly in planar view by 1.5 cm every 6 months, representing their average growth over 6 months (Table S1).” After checking table S1 and calculation codes, I still cannot get how you can get the variable linear growth rate of all four genera into a uniform the planar extension rate of 1.5cm? This is very important but it was not clearly explained as the explanation about the coral-coral competition in figure 1.

Line 133 photoquadrat: What is the size of quadrat and are they equal to the cells in your modelling? are these quadrats fixed or not between 2010 and 2016?

Line 371 remove “the” before how little
Figure 2 caption: relationship between planar growth (as a proportion of the maximum growth rate). If all data are expressed in proportion to the max, they should be below 1 and I noticed that there are many datapoints beyond 1.


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: SCALING THE EFFECTS OF OCEAN ACIDIFICATION ON CORAL GROWTH AND CORAL–CORAL COMPETITION ON CORAL COMMUNITY RECOVERY
Review round: 2
Reviewer: 1

Output:
Basic reporting: My concerns have all been well addressed. I have no further comment.
Experimental design: No comment.
Validity of the findings: No comment.
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: EFFECT OF GENETIC BACKGROUND ON THE EVOLUTION OF VANCOMYCIN-INTERMEDIATE STAPHYLOCOCCUS AUREUS (VISA)
Review round: 1
Reviewer: 1

Output:
Basic reporting: No comment.
Experimental design: The research question is well defined, relevant and meaningful. My only comment on experimental design is that these kinds of experimental evolution experiments should ideally include a "media only" arm, where bacteria are passaged without antibiotic to allow for the identification of mutations that arose due to passaging in the growth media, rather than antibiotic selection. Alternately, the parent strain can be "pre-adapted" to the growth media before the selection experiment starts. If the authors haven't done either of these, then there needs to be more explicit mention in the Discussion of how mutations observed could be due to adaptation to experimental conditions, rather than antibiotic selection.
Validity of the findings: No comment.
Additional comments: The manuscript by Su et al. describes a thorough investigation into patterns of genetic adaptation that arise during the evolution of VISA. Besides my comment above about the experimental design, I have a few additional comments:

1. There should be more attention paid to the actual mutations detected in each of the genes of focus in this study. The mutations themselves should be summarized somewhere, such as a supplemental table. In reading the paper I got the sense that all mutations in a given gene were being treated equally by the authors, but is this really the case? Presumably mutations in different regions/domains of a given gene can have different downstream effects. While a variety of different mutations were observed in each gene, did they tend to cluster in particular regions/domains? This information is useful for thinking about how mutations in these genes exert an effect on VISA phenotype, and should therefore be included. Finally, I'm a bit confused about how conclusions regarding genetic background effects can be made when the actual mutations in the genes are different. Unless the authors are confident that two different non-synonymous SNPs in a particular gene have the same functional effect, I would suggest softening the conclusions regarding epistasis except in cases where allele-level parallelism (rather than just gene-level parallelism) is observed.

2. Are the genome sizes of the parent strains very different from one another? If so this could effect the number of mutations observed during the course of selection. Either way parental strain genome sizes should be reported.

3. Tables 1-5 should be moved to supplemental material, since indels were not a focus of the study.

4. It would be helpful to see the MICs of the parent strains indicated somehow on Figure 3.

5. What are the actual mutations in the isolates with the highest Daptomycin and Vancomycin MICs in Figure 4? Why do the authors think that these mutations have the greatest phenotypic effect?

6. Lines 146-148 in the methods should also be stated in the Figure legends for Figures 5 and 6.

7. Lines 25-27 in the abstract were confusing to me. I would suggest revising to "We used an iterative method to experimentally evolve three important MRSA strains to generate a library of 120 VISA isolates."


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: EFFECT OF GENETIC BACKGROUND ON THE EVOLUTION OF VANCOMYCIN-INTERMEDIATE STAPHYLOCOCCUS AUREUS (VISA)
Review round: 1
Reviewer: 2

Output:
Basic reporting: The manuscript it well written and provides a clear description of both methods and results. The introduction is well structured and provides a suitable context for the study. The review of the literature is appropriate for the topic of vancomycin resistance.

The structure is consistent with the scope of PeerJ. The figures are of good quality and clearly labelled. A figure 1 with a summary of the experimental evolution approach would be very helpful (see below).

There are two excel tables with raw data are on phenotypic testing of evolved isolates, however the authors should also provide a table with all mutations detected (eg breseq output in tabular form).
Experimental design: This is an original genomic study of experimentally evolved vancomycin-resistant S. aureus strains. It fits the scope of the journal. The research question (“What is the impact of the clonal background of MRSA on the acquisition of vancomycin resistance in terms of mutations, fitness cost, ability to become of resistance and co-resistance to other antibiotics?”) is clearly stated and fill a knowledge gap in the field.

The investigation is rigorous, and the methods applied in terms of resistance selection, genomic analysis and phenotypic testing are state-of-the art. More detail should be provided on the genomic analysis (eg QC of reads, were mutations filtered / manually validated?).

Regarding the statistical analysis, the authors combined different approaches to assess the impact of background, SNPs on the phenotypes: first, they used non-parametric test to compare backgrounds in terms of vancomycin MIC, growth rate, number of mutations and prevalence of mutations in the top mutated genes (the latter with Bonferroni correction); second, they fitted a binomial generalised linear model (logistic regression) to assess the impact of background on the prevalence of mutations in the top mutated genes. Finally, the fitted linear models where vancomycin MIC was the response variable and the top mutated genes were predictors (as binary variable mutated - non mutated).

Overall, the approach used seems appropriate, however, the statistical analysis is quite complex and I suggest providing a more structured description in the methods session, for example separating simple comparisons between groups and more complex models. It would also be good if the authors could share the code used, at least for the complex linear regression models.
Validity of the findings: The main findings of the study are :
1.	Vancomycin resistance selection in CC8 background appears to be more difficult with a higher number of selection steps, higher number of mutation and higher fitness cost
2.	While there are mutations in 151 genes among 107 evolved strains, six genes (walK, prs, rpoB, rpoC, vraS, yvqF/vraT) have a strong signature of convergence
These findings are supported by the data and are also consistent with what is known about the genetic determinants of VISA so far. However, this is not a mere replication study, since it explores the relationship between mutations and clonal background and also, by analysis 107 independently evolved strains, uses an approach that increases the chance of identifying convergent mutations and that allows to rank the importance of these mutations / mutated genes.’

The conclusions are well stated and there is a clear recognition that, while these findings might inform the use of genomics for the prediction of vancomycin resistance, data from experimental data alone are not sufficient for prediction of resistance.
Additional comments: 1.	While the six “universally mutated genes” captures more attention, I think that the authors should explore the remaining 145 mutated genes a little bit more. I’m impressed by this high number: how many of these have been previously linked to vancomycin / antibiotic resistance? Are there compensatory mutations?
2.	Did the authors perform an epistasis analysis of the 151 mutated genes? This should give a good opportunity to explore the issue of epistasis in antibiotic resistance as stated in the introduction
3.	Again to investigate epistasis and compensatory mutations a bit further, it might be useful to sequence strains obtained at an intermediate stage of the resistance selection
4.	Please add a figure 1 with an overview of the experimental approach: resistance selection steps, number of independent experiments, choice of strains for downstream phenotypic and genomic analyses
5.	Line 147: you mention the “average r of each strain”: were growth curves measures performed in replicates?
6.	Line 151: see comments above for the description of the statistical analysis.
7.	Line 219: truncations are often found in experimental evolution: what was the proportion here? Did you look at intergenic mutations to investigate whether they occurred in promoter regions of known VISA determinants?
8.	Line 227 and figure 2: there are a couple of outliers among the NRS70 and NRS384 mutants that have a very high number of SNPs (> 20). What is the reason for this? Were the alignments checked manually to exclude false positive calls (eg in repetitive regions)?
9.	Line 229: was there convergence among mutations / positions or mutated regions within the top mutated genes?
10.	Line 320: Which genes were most frequently mutated in evolved strains with co-resistance to daptomycin?
11.	Line 331: given the accumulation of mutations in some strains, it would be interesting to test whether re-sensitisation to beta-lactams was related to compensatory mutations
12.	Line 354: I appreciate the use of modelling to investigate the impact of single mutated genes and genetic background on the final vancomycin MIC. Here add a comment on stats approach
13.	Please add a table with the list of mutations / large deletions / insertions identified using breseq


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: EFFECT OF GENETIC BACKGROUND ON THE EVOLUTION OF VANCOMYCIN-INTERMEDIATE STAPHYLOCOCCUS AUREUS (VISA)
Review round: 2
Reviewer: 1

Output:
Basic reporting: No comment.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: The authors have addressed all of my comments in their revised 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: EFFECT OF GENETIC BACKGROUND ON THE EVOLUTION OF VANCOMYCIN-INTERMEDIATE STAPHYLOCOCCUS AUREUS (VISA)
Review round: 2
Reviewer: 2

Output:
Basic reporting: The authors have appropriately responded to the reviewer’s comments and the new version of the manuscript is now clearer and provides more information on statistical methods and on the details of the variants identified.
Experimental design: No new experiments have been performed for this revision; however, the authors have now performed a genome-wide epistasis analysis using SpydrPick, a tool for bacterial genome-wide epistasis studies (GWES). It is plausible that the dataset was too small for a GWES.

I agree with the authors that an epistasis analysis would require further data and investigations that are beyond the scope of this manuscript.
Validity of the findings: No comments, since no new findings have been added
Additional comments: 1.	Please provide a description of the supplementary data. I’m not sure on what supplemental tables 2-6 represent


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: PEACH-MORCHELLA INTERCROPPING MODE AFFECTS SOIL PROPERTIES AND FUNGAL COMPOSITION
Review round: 1
Reviewer: 1

Output:
Basic reporting: The English language looks good, and some errors were existed in the text.
In the introduction, please focus on the advances of the soil microorganism and soil health caused by intercropping patterns.
Experimental design: The experimental design is ok.
Please provide the details of the methods of enzyme activity assays.
Validity of the findings: This manuscript was study the effects of peach-Morchella intercropping on the soil physicochemical properties, enzyme activities and fungal community in an orchard. The results indicated that intercropping reduced soil fungal diversity and improved soil structure and fertility, and suggested intercropping is the efficient planting practice for soil health and farmers’ income. The data is detailed, and the result part is also very sufficient.
Additional comments: Some amendments are suggested as follows:
Line 39, delete “.” After “activities”.
Lines 64-67, I do not understand the planting processes of Morchella from the description.
Line 72, the PM-CK was without peach-Morchella intercropping. Was it from the monoculture peach or Morchella?
Line 78, please descript the sites of soil sampling. How far was the sample from the tree?
Please check the cited format of the reference, such as “Gawryluk, Wyupek & Pawe” in the Line 89, “Huang, Li & Zhang” in the Line 94, and in the Lines 198-200, and so on.
I found the soil organic matter was sharply increased in 1 or 2 years. Generally, the increase of soil organic matter was slow, please explain it.
Lines 180-182, I do not think CK and PM1 were closer on the first axis, while PM1 and PM2 were closer on the second axis, and thus confirmed continuous intercropping result in the decline of the soil fungal diversity.
Please check the “sucrose” and “sucrase”.
Please increase the font size in the Figure 3.
In the discussion, the authors mentioned “Soil physical and chemical properties are closely related to each other”, “Organic matter has a great effect on soil enzyme activities”. I suggest the authors add the correlation analysis between soil physicochemical and biological properties, especially the association between the sol properties and fungal community in order to improve the quality of 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: PEACH-MORCHELLA INTERCROPPING MODE AFFECTS SOIL PROPERTIES AND FUNGAL COMPOSITION
Review round: 1
Reviewer: 2

Output:
Basic reporting: In general, the paper sent a clear message and demonstrated an interesting intercropping system of peach and Morchella mushroom, which yielded significant impact on both economic output and soil health. The article provided sufficient background and context regarding the relevance and scientific basis of intercropping systems on fruit crops. I commend the authors for sharing the detailed sequencing and crop yield raw data to support their claims.
Experimental design: The research questions regarding soil health, reflected in soil physico-chemical properties, crop yield, and economical benefits were clearly laid out in the introduction and well-addressed by the results.
Validity of the findings: The author clearly stated the experimental set up and the results are clearly supported by data.
Additional comments: In general, the paper sent a clear message and demonstrated an interesting intercropping system of peach and Morchella mushroom, which yielded significant impact on both economic output and soil health.

There are, however, several places need to be improved before this paper can be accepted for publication:

1.Line 70: Soil sample collection: please explain how far away from the peach trees the soil samples were collected. The difference between bulk soil and rhizosphere microbiome can influence the result and interpretation of the data collected in this study.

2.Line 74: Did you mean “from 2015 to 2017”?

3.Line 86: An brief explanation of what NYT 1121.4-2006 is should be included.

4.Line 100: “…employed to isolate fungal DNA”. I assume that the first step was to extract total DNA in the soil, not just fungal DNA. Only fungal ITS sequences were enriched from total soil DNA in the following step.

5.Table 1: I suggest the authors include standard deviations/standard errors when reporting statistics of the soil physico-chemical properties, and indicate specifically which statistical test was used to access differences between groups here (presumably, one-way ANOVA?)

6.Table 1 and throughout the manuscript: The author should replace “Pure Income” with a more accurate term, such as “Net income”, of “Profit” as this value was calculated as such.

7.Line 178: typo (should be “Fungal” instead of “Fugal”)

8.Line 273-274: “the fungal community is probably an important contributor to the higher fertility of peach orchard soils in this study”. It is surprising that the authors observed a reduction in fungal diversity, especially at year 2 of inter-cropping. I would like to see the authors’ elaboration on the correlation between fungal diversity and soil health, especially in the context that there are previous reports linking the positive correlation between fungal diversity and soil fertility.

9.Also, would the author expect that continuous peach-Morchella inter-cropping would make the soil fungal diversity worst in the long run?

10.Figure S1: it would be informative if the authors can show individual replicate and the hierarchical analysis to see if the same treatments would clusters together.

11.Please elaborate on how inter-cropping and the reduction of general fungal diversity would affect the abundance of pathogenic fungi often found in this system.


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: LONG-TERM DYNAMICS OF NOROVIRUS TRANSMISSION IN JAPAN, 2005–2019
Review round: 1
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The manuscript written by Misumi and Nishiura descriptively analyzed the norovirus outbreaks reported in public sectors in Japan, 2005-2019, and built statistical models to predict the risk of infection, asymptomatic ratio, and risk of norovirus positive outcome. The outbreak data was nicely collected and summarized well, providing interesting insights on temporal dynamics of transmissibility and asymptomatic ratio. Authors could expand more the method section to explain the robustness and/or limitation of the available dataset. More specific analyses/descriptions to the outbreak settings would be helpful to gain insights on norovirus infection patterns with more biologically relevant explanations/hypotheses.

1. The authors’ hypothesis is not very clear to this reviewer. Why the dominance of GII.4 implies increase transmissibility of this virus -resulting in larger outbreak size- over time? If that is true it is easily visible from the annual epidemic curves showing the increase of the infection cases, which seems not true in Fig. 1. I think it is not necessarily increase its transmissibility but could you please explain a little bit more the hypothesis? The dominance of this genotype has been partly explained by its epochal evolution and continuous changes of its antigenicity (Debbink et al. PLoS Pathog 2012; Parra et al. PLoS Pathog 2017; Tohma et al. MBio 2019), which have probably resulted in the increase of susceptible population to the new variants. That said, it should be very interesting if authors could estimate those parameters for GII.4 and non-GII.4 viruses, parsing those into corresponding variants circulated in each time period. Also, if possible it might be interesting to look over the outbreak size, number of outbreaks, genotype distribution, and parameters by outbreak settings, e.g. nursery, elderly facility, restaurants, hotels.

2. Please describe more detail about the dataset and data collection procedures; definition of outbreaks, sampling strategies (collected fecal samples from all the symptomatic patients or a subset of them? How about asymptomatic cases? What is the criteria for diagnosing cases as symptomatic? How did they define the exposure to the viruses?), and viral detection procedure (norovirus screening detected all three human genogroups? Are they genotyped by PCR, or genomes sequenced?) are not clearly written in the materials and methods section. If genotype information is available in each outbreak, it is helpful to show the prevalent genotypes in each of them or in each year. As I believe this data was not collected in cohort studies, The ratio of asymptomatic infections and data for the model fit largely relies on the sampling strategy from asymptomatic cases in each institute or outbreak settings.

3. Although authors claimed that findings in this manuscript was consistent with the study from Matsuyama et al. (lines 188-190), it is not reflecting the sharp drop of the reproduction numbers in 2011-2012 or increase in 2013 in their study. Besides, in Matsuyama’s study the best fit model presented steep rise of the reproduction number from 2006/07 to 2007/08 season, which was included in the year-range in this study, and seems not consistent with data here. Parameters estimated were different between these studies but it would be nice if authors could explain the differences between the studies.

4. In addition, authors associated the yearly variation in the asymptomatic ratio with the change of dominant genotype circulating in the corresponding winters (lines 190-192, 223-224), without showing any data of dominant genotypes in each winter season. Again, it is helpful if authors could provide such information and conduct additional correlation analyses to prove this hypothesis. Authors commented (without statistical significance in this study and Miura’s study) that high asymptomatic ratio could be attributed to the dominance of GII.4 virus (line 224); however, multiple birth cohort studies showed higher or similar asymptomatic ratio upon GII.4 infection as compared from that of other genotypes (Ballard PLoS ONE 2015; Burcado Infect Genet Evol 2017; Saito Clin Infect Dis 2014; Colston JID 2019; Bhavanam Microorganisms 2020). Also, waste water monitoring along with clinical surveillance indicated wide range of genotypes distributed in sewage samples, suggesting that clinical samples were skewed to symptomatic infections from GII.4 while more asymptomatic/mild infections with different genotypes circulated in the community (Kazama Water Res 2016; Fumian Environ Int 2019). Please clarify any inconsistencies between the modeling analyses and those cohort/monitoring studies.

Minor comments:

5. Lines 54-55: Reference for the ’18 particles’ is incorrect. It should be Teunis et al. J Med Virol 2008.

6. Line 176: “systematically analyzing reports of 1728 outbreaks by published the prefectural institutes” should be “systematically analyzing reports of 1728 outbreaks published by the prefectural institutes”?


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: LONG-TERM DYNAMICS OF NOROVIRUS TRANSMISSION IN JAPAN, 2005–2019
Review round: 1
Reviewer: 2

Output:
Basic reporting: The English is clear. Reference also good. The background information is adequate. However, the author did not share any information about the model.
Experimental design: There is no experiment design specifically for the paper. The author has predicted the risk of infection.
Validity of the findings: no comment
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: LONG-TERM DYNAMICS OF NOROVIRUS TRANSMISSION IN JAPAN, 2005–2019
Review round: 1
Reviewer: 3

Output:
Basic reporting: Long-term dynamics of Norovirus transmission in 2005 – 2019 in Japan.
Megumi Misumi, Hiroshi Nishiura.

Norovirus infections are a major cause of gastroenteritis outbreaks. Foodborne transmission is an important route for the spread of virus and globalization of the food industry enables norovirus outbreaks on an international scale. The rapid rate of genetic and antigenic evolution of noroviruses makes it difficult to develop vaccines or therapies. Thus, it is really important to determine the dynamics of transmission of noroviruses over many years and also at the level of different genogroups. In this study, Misumi et al., address the dynamics of norovirus transmission from 2005 -2019 in Japan and also examined the predominant strain that caused the majority of clinical cases. The authors investigated the annual reports of 1728 outbreaks from the data obtained from 78 institutes of health in Japan. They reported geographic distribution of outbreaks in seven regions. They found that Kinki region had the highest number of outbreaks. In addition, they examined yearly and monthly variations in number of outbreaks. With data available only from Nara and Tokyo, they also reported yearly outbreaks of norovirus by type of outbreak settings. In Figure 2, they reported the seasonal (2A) and yearly (2B) variation of number of outbreaks from 2005-2019. They found large variation in outbreaks in winter compared to months of summer.  Misumi et al., also found that noroviruses of the genotype GII.4 has been a predominant strain circulating in Japan (Table 1 and 2). Their study led to an interesting conclusion that the number of outbreaks of norovirus has not changed significantly since 2005 in Japan. It will be interesting to know the transmissibility of noroviruses on an international scale because of ever increasing globalization of food industry.

Overall, it is an important and essential study reporting the changes in outbreaks of noroviruses nationally since 2005. It is a nice paper with carefully acquired data, it is well written. In my view, this manuscript is appropriate for publication in PeerJ.
Experimental design: Yes, the research question is well defined, investigated rigorously. Methods described with sufficient detail.
Validity of the findings: Their data is statistically sound and controlled. Conclusions well stated.
Additional comments: Very minor issue:

Please label both x and y axis in Figure 1D and E.


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: LONG-TERM DYNAMICS OF NOROVIRUS TRANSMISSION IN JAPAN, 2005–2019
Review round: 2
Reviewer: 1

Output:
Basic reporting: The revised version of this manuscript addressed all of the concerns and comments from this reviewer.
Experimental design: no comment
Validity of the findings: no comment
Additional comments: I have just a few minor comments on figures and tables.

1. Where are Nara and Tokyo in the map? It would be helpful if the location of those is indicated in the map in Fig.1A.

2. Is there any reason the order of each group in the graph is different between Fig.1D and 1E? I mean, in the Tokyo graph, the order of the groups in the bar plot and legend is different (oppositely ordered; Other>Eldery>Elementary>Nursery from top to bottom). Also, y-axis title is missing in Fig.1D.

3. Table 2: What does “GI+GII” mean? Co-infection/detection of GI and GII or a dataset that included all the outbreaks caused by GI or GII? If the latter is the case, why risk of asymptomatic infection is much lower than those with GI or GII alone?


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: LONG-TERM DYNAMICS OF NOROVIRUS TRANSMISSION IN JAPAN, 2005–2019
Review round: 2
Reviewer: 2

Output:
Basic reporting: The author has addressed the comments
Experimental design: The author has addressed the comments
Validity of the findings: The author has addressed the comments
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: LONG-TERM DYNAMICS OF NOROVIRUS TRANSMISSION IN JAPAN, 2005–2019
Review round: 2
Reviewer: 3

Output:
Basic reporting: The authors addressed most of our concerns and made required changes to the manuscript . Thus, the manuscript is appropriate for publication in PeerJ Journal.
Experimental design: No comments.
Validity of the findings: No comments.
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: INTER- AND INTRASPECIFIC VARIATION IN THE ARTIBEUS SPECIES COMPLEX DEMONSTRATES SIZE AND SHAPE PARTITIONING AMONG SPECIES
Review round: 1
Reviewer: 1

Output:
Basic reporting: The paper is well written. However, it is primarily focused on the system. Therefore it might be a better fit for a taxon-specific journal. I recommend providing more background information in the Introduction relevant to the question(s) instead of focusing exclusively on the system. By making these changes, the paper will be more attractive for a wider audience and possibly more suitable for PeerJ readers. Also,  I wonder if the author could make hypotheses that go beyond testing previous findings. Thus, the author would focus on the novelty of the results presenting in the paper, their new hypotheses and interpretations.
Experimental design: The study aims to assess the extent of the inter- and intra-specific variation in the skull’s size and shape of Artibeus. Although the author did analyze the data to test for the primary goal of the study, some information is required to validate the accuracy of data collected. In geometric morphometrics, it is well known, the need to account for measurement error, which is regularly accounted for by digitizing and photographing images at least twice. It is not mentioned that this was accounted for, so it is important to mention if it was (or not) the case. Also, in morphometrics, evaluating the degree of differences between sexes is traditionally recommended to be assessed before conducting any further analysis, particularly in a group for which previous studies have reported sexual dimorphism. The author did evaluate for differences between sexes. Still, it is unclear why the author did not partition the sample or run analysis separately by sex after finding statistical significance between males and females in the Procrustes ANOVA. One possibility for not doing this is the limited sample size in each of the two datasets analyzed. If so, it has to be mentioned.
Validity of the findings: The author overestimates the scope of the data used to conduct the analyses by referring to it as a “large sample.” Given the wide distribution of most of the species included in the analyses, it is unclear how the data could reflect the scope of the inter- and intra-specific variation in the group. More importantly, analyzing the level of intraspecific variation within a single species in the genus, at what scale, can be meaningful to understand overall patterns of variation in the group. Finally, the Discussion section might need to provide a more extended exploration of how the findings can be used to make inferences about patterns of variation in the group beyond the actual distribution of the species in the morphospace. Therefore, there is a need to link the findings to mechanistic explanations associated with environmental, behavioral, or phylogenetic clustering that could be used to interpret the Results.
Additional comments: -   	It is recommended to expand the justification to conduct the study in the Introduction. Otherwise, the whole motivation is primarily focused on the system and not necessarily on the processes that can be driven differences in shape among species and within them. For instance, why do we expect differences in shape and size to arise? How has morphology been reported to show “higher” variation between species that occupy large geographic ranges? How can ecology play a role in this?
-   	Line 129: In Figure 1, the author shows a phylogeny based on data presented by Rojast et al. (2016), which, although it does not reflect the most recent revision in the group (see Hoofer et al. 2008; Larsen et al. 2015; Redondo et al. 2008; York et al. 2019), it should be properly cited and explained on the legend to understand better how the phylogeny was generated and add the methods used to plot it in the corresponding section.
-   	Line 35. The author refers to the localities included in the intraspecific analysis, plotted in Figure 2. However, it is unclear by just seen the map the location of the sites where the specimens examined where collected. It is recommended to mark each of the unique locality points included in the study, instead of highlighting countries that are just geographic constructs that in nature might not reflect the actual distribution of species (or subspecies in the case of the intraspecific dataset). Also, it would be good to see the location of the sites for the interspecific analyses too, and ultimately providing a graphical representation of the geographic distribution of each of the species analyzed will allow assessing better how the sampling can capture the scope of the variation across the geographic range of the species, instead of potentially increasing the geographic differences among sites just by an effect of geography.
-   	Line 167. The author mentions that due to “sample size limitations,” sexual dimorphism in Artibeus species was assessed as a whole, instead of within species as it was done in A. lituratus. How can this influence the outcomes?
-   	Line 175. The author mentions that a phylogenetic Procrustes ANOVA was not conducted because of the need to summarize all the variation within each group using mean values and this would “loss” differences accounted by sex. Traditionally, when sexual dimorphism is evaluated, and there is statistical support to validate it, further analyses to evaluate the scope of variation in the sample must be conducted separating males and females. Did you consider doing that? It might be important to do this, especially if one of your goals focuses on testing this potential source of variation, which you found evidence for in your Procrustes ANOVA (line 253), but it seems that the author disregarded it by combining the data for the rest of the analyses.
- Line 260. The author found that differences among species in the morphospace are accounted for by phylogeny. However, this is not discussed later in the manuscript. How can this be interpreted? What kind of inferences can be made based on these results?
-   	Figure 6. How do you explain the differences in the arrangement of localities in the UPGMA cluster?
-   	Line 296. It is stated in the paper that the “overarching goal of this study was to understand better the relationship between intraspecific and interspecific variation and morphological differentiation based on a large sample.” How can intraspecific patterns of variation in A. lituratus reflect the scope of variation in the group within species? Given the wide range of distribution of most of the species included in the analysis, how the limited sample size to evaluate inter- and intra-specific variation can influence the outcomes?
-   	In the Discussion section, the author should avoid being too descriptive with the findings and instead focuses on their interpretation and possible inferences.
-   	How the differences accounted for by sex can be explained? How could geography play a role in the scope of variation reported between species and within them?
   	How can changes in the shape and size of the skull impact species' fitness in this group?
-   	What is the interpretation of the level of variation observed between species?
-   	Line 387. Is the intraspecific arrangement of populations in A. lituratus associated with the currently recognized subspecies in the group?


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: INTER- AND INTRASPECIFIC VARIATION IN THE ARTIBEUS SPECIES COMPLEX DEMONSTRATES SIZE AND SHAPE PARTITIONING AMONG SPECIES
Review round: 1
Reviewer: 2

Output:
Basic reporting: Basic reporting is all fine.  This is an interesting and well written manuscript.
Experimental design: The only concern that I have with the experimental design are the identities of the specimens.  I do not believe that the bats in the LSU mammal collection have been curated since the 1970's.  The Taxonomy of Artibeus has been very fluid since then.  Did the author accept the species names on the tags of the specimens or did they independently verify the ID of each individual?  If they did not independently ID specimens, when was the last time that the Artibeus collection was curated?  Based on the data provided by the author, there is no way to establish even where the specimens come from.  I would like to see a specimens examined section added to the manuscript and in that speciments examined, the location of each specimen be disclosed.
Validity of the findings: All underlying data have not been provided.  The authors need to provide the raw landmark data.  It appears that they make the derived data (PC's) available, but not the raw landmarks.  As stated in the Intro, Artibeus is not the most species rich genus of Phyllostomidae...Sturnira is.
Additional comments: Nice manuscript, just concerned about the specimen ID's.


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: INTER- AND INTRASPECIFIC VARIATION IN THE ARTIBEUS SPECIES COMPLEX DEMONSTRATES SIZE AND SHAPE PARTITIONING AMONG SPECIES
Review round: 1
Reviewer: 3

Output:
Basic reporting: Clarity of hypotheses and gap in knowledge: This work is described (L95-100) as a re-assessment of interspecific variation in this same group, with a different analysis method. Could you expand on why you might expect different results with this new method or limitations of previous methods? This would also help define a clear hypothesis, which is currently lacking, though predictions are laid out in lines 106-225. The manuscript currently doesn't define a clear gap in knowledge.

Presentation and visualization of data (see also comments under #3): (1) I appreciate that there were 2 different sets of color for inter-and intraspecific analyses. However, I found myself trying to find colors from Fig 1 in the map in Fig 2 and wondering, e.g., if the light green colors were supposed to match. A few suggestions to try to distinguish these two analyses more: Perhaps different symbols could be used in figures 4-6? How about moving Figure 2 to directly before Figure 6? (2) Rather than a boxplot, using a violin plot, adding individual points to the boxplot, or using another alternative that shows the distribution of data points would avoid known shortcomings of a basic box plot and more fully detail your data in Figure 5. (3) Have color sets used been tested regarding readers with red-green colorblindness or those printing in black and white?

Raw data: Stated as provided to the journal. Perhaps it would be more accessible if posted in dryad or other repository that have clear policies about archiving as well?
Experimental design: Effect of size: (1) I appreciate that the allometric trend in this data is assessed, particularly given the size differences between specimens and statistical association of size on shape. Is there a reason size correction was not done to generate allometry free shape principal component measures, a function that is built into geomorph, rather than assessing via ANOVA and common allometric component? Was it possible to assess overall length of animals, or were only crania available? This may be particularly important as PC1 explains both size and shape. Shape differences (which seemed like the key question in the analysis) may be more distinct once size is removed from the variation. (2) What are size differences in intraspecific analysis in Figure 6? Based on ANOVA analyses in L280 and L283), it sounds like this is a contributing factor to variation amongst countries, but this is not addressed within the presented data in Figure 6.

Choice of species and phylogenetic corrections: (1) While I appreciate wanting to preserve the sexual dimorphism component (L174-177), is there a reason phylogenetic correction Procrustes ANOVAs were not performed to verify species-level signals? That is, one analysis to verify this was not causing a bias in conclusions about species, and another analysis to more specifically look at the influence of sex on shape? (2) Are the 11 species chosen representative of Artibeus cranial variation?
Validity of the findings: Illustration and inclusion of data to assist readers in reaching conclusions: (1) I found myself wishing for more visualization of the data in Figures 4-5. For instance, a warp of mean shape for each species, a violin or other plotting of range of shape by species or species and sex, or even outlines on the morphospace to visualize distinctions between species. These types of figures would help readers make their own assessments of distinctions between shape in small and large Artibeus groups (results L201-203), and other descriptions of data in the results section. As is, this is difficult for the reader to parse easily, without matching the full species name with abbreviations in the figures, then searching for the colors in the morphospace. (2) What are the shape differences on PC2? This warp is not illustrated. This is especially interesting as species in the small Artibeus group were separated in PC2 scores. Additionally, warps of shape variation are not provided at all for intraspecific analyses of A. lituratus. (3) Sex was stated as an important variable (L174-177, results in L252-259, based on statistical analyses, and a L359-385 in discussion), but this is not illustrated in figures. Perhaps this could be done with symbol size or filled/unfilled in Figures 4-6? Alternatively (and perhaps preferably for clarity), should there be a figure comparing sexes if this is an important conclusion?

Relation of shape changes to niche and foraging: (1) A primary discussion point was how these shape changes related to feeding niche (L24-25 and L26-31 of abstract, L312 “strongly suggests niche partitioning of species,” and diets are discussed in L346-357). I believe there is extensive information about what these species eat. Could this be incorporated and tested? Alternatively, this is presented as a supported conclusion in the abstract, rather than speculation, as it appears in the discussion. This discrepancy needs to be addressed. (2) Literature is cited in L380-381 about the relationship of skull shape to fruit hardness. Given these and other literature related to function of different skull shapes, can more details about feeding be addressed in the discussion? That is, rather than concluding that there are “different niches” could there be a discussion of the impact of shape on feeding mechanics? This would require further detailing of shapes in the results, as the shapes detailed are currently pretty scant. E.g. “shorter rostrum” in L210, or no visualization of the shapes between males and females or warps for intraspecific analyses.

Alternative explanations not fully discussed: Foraging is presented as the explanation for all shape variation. Are there additional speculations about the functional impacts or driving forces that cranial variation? For instance: (1) What is the relationship between geography and phylogeny across species, and more specifically within A. lituratus? That is, is there an alternative explanation (genetic variation) that could explain the differences in morphology, rather than diet? (2) For intraspecific analyses, it is mentioned that there are no patterns of geographical variation (L307-308). However, it does not appear that these designations are included in figures or Table S5 to support that conclusion. That is, can a non-specialist who doesn’t know the various collection locations also reach this conclusion? (3) Could sexual selection explain sexual dimorphism rather than sexes choosing fruits of different hardness?

Other minor comments:
--What is shape on the Y axis in Figure 5? PC1 score?
--L232 Should “dorsal PCA” be “lateral PCA”
--Are descriptions of shape differences being minor in interspecific analyses (L 273-274 and L278-279) based on qualitative assessments or can this be quantitatively or statistically assessed?
--L305 Should this read “…contrary to previous studies…”
Additional comments: This study assesses both inter- and intraspecific variation in cranial morphology within the adaptive radiation of bats. While clearly written and methodically sound, additional analyses, specifically size and phylogenetic corrections, as well as testing of foraging speculations, would greatly strengthen conclusions or allow current speculations within the abstract to be appropriately assessed. Further illustration and visualization of data as I have suggested above should be improved upon before acceptance and would not require additional review. However, additional analyses such as size correction or diet analysis may alter conclusions drawn or allow a new line of more detailed conclusions to be drawn, which may necessitate another review.