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: THE RESPONSE OF GEOPHYTES TO CONTINUOUS HUMAN FORAGING ON THE CAPE SOUTH COAST, SOUTH AFRICA AND ITS IMPLICATIONS FOR EARLY HUNTER-GATHERER MOBILITY PATTERNS
Review round: 1
Reviewer: 2

Output:
Basic reporting: Source documents and information: Written consent was obtained and this is noted in manuscript, but there is no copy of form uploaded; if PeerJ required this, then the authors may attend to it. Similarly, the Metadata sheet on “DepletionForaging” workbook seems to not be fully filled out.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: This is a clearly-written presentation of a very useful study. There are not enough studies with this level of detail to actually test hypotheses that abound in the archaeological literature about foraging behavior. The authors make a strong case about the lack of quantitative data available for understanding the impacts of human foragers on plant yields, especially in the GCFR. The study integrates nicely into existing (and especially recent) literature, and it was absorbing and informative to read. It is an impressive study with an enormous dataset, and the findings are robust and offer useful data for future work. I find almost nothing to critique in the way the study was designed, the way it is contextualized in the literature, the way the analyses were conducted, or the way it was presented. The introductory material and discussion was engrossing and informative, especially considering how plant remains are under-researched in hunter-gatherer archaeology. The idea that the evolution of bet-hedging strategies during periods of exposed Palaeo-Agulhas Plain and increased herbivory, leading to fixation in the modern day when it is less necessary, is fascinating. If this is true, then the authors may be picking up on one of the key niche-constructive behaviors of our early ancestors.

I have only a few minor comments, below, and then I believe it is ready to be accepted.

1) 	Implied linkage of genetic ancestry and traditional knowledge: On Line 176 the authors write, “Many residents from these towns have genetic ancestry linking them to the Khoe-San (de Jongh 2012). The Khoe-San share descendants with the San, who were traditionally hunter-gatherers, and the Khoe-khoen, who were traditionally pastoralists (Crawhall 2006; Schlebusch 2010). Recent research suggests that these people were the direct descendants of Homo sapiens sapiens (Pickrell et al. 2012; Schlebusch et al. 2020; Veeramah et al. 2012) who lived on the Cape south coast since about 160 000 BP (Brown et al. 2009; Marean 2010).” My first critique is that the genetic ancestry of a population is irrelevant to the problem at hand, which is how quickly foragers will deplete a plot of USOs. The way the sentence is set up, it sounds like the authors are implying that because there is some genetic ancestry linking the individuals in their study to ancient foragers, these specific people should have particularly pertinent knowledge about the local environment and foraging opportunities in the present day. This may or may not be true that they do, but the genetic connection does not give any special reason on its own that it should be true. When the authors go back in time to point out that genetic studies broadly – and I do mean very broadly – link living Khoe-San and Khoe-khoen to ancient individuals associated with foraging material culture, and that such populations have likely been in place across southern Africa for more than 100,000 years, they are stretching this argument to its very limit. To me, the entire set of sentences should be removed because it is irrelevant to the point, and risks implying some kind of genetic-cultural relationship between genes and foraging knowledge. Furthermore, have these specific people in these specific towns actually had their genes linked to ancestral or living San? Or is this just an assumption based on what is more broadly known about the distribution of genetic ancestry across southern Africa and (undoubtedly) some assignment of ancestry based on phenotype? It is enough to just say the following: There has been evidence of early human foragers (why even go into the species and subspecies?) in the area going back at least 160,000 years (CITE). And if they want to talk about genetics, they could say something like: Genetic data indicate some continuity between living people across the southern African coast with ancient foragers, thus suggesting that cultural knowledge about USO harvesting in these ecosystems may also have had the potential to be passed down from very ancient times. In short, this section doesn’t add anything to the argument about foraging and risks opening up problems.

2) There are a few random places where some typos remain, but they are far and few between (e.g. no need for comma on line 321 after “region”, line 368 should read “an evergreen geophyte”, etc.). Overall, it is written very clearly and concisely and is very information rich, and would benefit from one last read-through.

3)  Line 42 – Is poor reproductive success the best wording? It seems more that they authors are describing the slow growth and difficulty in recruitment, which would only translate to poor reproductive success when there is high herbivory or other reasons for high extrinsic mortality. Perhaps the authors mean fertility?

4) Line 300: suggest adding the following reference, as that is where : Rector, Amy L., and Kaye E. Reed. "Middle and late Pleistocene faunas of Pinnacle Point and their paleoecological implications." Journal of Human Evolution 59.3-4 (2010): 340-357.

5) Lines 358-359 – spelling is “O’Connell

6) Line 410 – reference seems incorrect

7) Figure 1 – It would be useful if this figure showed the distribution of the various vegetation types within the GCFR on the main map. Also, on the tiny map inset it is difficult to discern the GCFR at all. I suggest zooming in on southern Africa – it’s still big enough that a reader can identify where it is. That way the entire GCFR can be shown, and possibly even some lines drawn to indicate the summer vs. winter rainfall in the GCFR for reference for readers less familiar with this specific rainfall configuration.

8) Figure 4 –The caption reads that we are looking at yields of species, but then at first I could see only total weights and years and not discern where the species were. After much effort (which is not really what you want in a figure, which is meant to provide clarity), I think I can see how to interpret it. However, are there some mistakes? For example, how can there be any black boxes in some years, like in Plot 20, but not in all years, if black represents a taxon that was harvested for all three years?

9) Figure 5 – It is almost impossible to discern the difference in the two greys that are used. This is, however, an incredibly useful figure.

10) I would find a single master figure to be very useful that shows the rainfall, year, and yield of each plot together in one place. Even just mapping rainfall onto Fig. 3 would be useful so we can see the relationship.


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: NETWORK SUBGRAPH-BASED APPROACH FOR ANALYZING AND COMPARING MOLECULAR NETWORKS
Review round: 1
Reviewer: 1

Output:
Basic reporting: The work describes an approach to determine network similarity in the contest of directed graphs. Overall the works is interesting and the description sufficiently clear.

The testing approach used by the author is acceptable although when exploring the function of genes embedded in certain motifs enrichment analysis should be used (including a measure of statistical validity) when making any claim on the functions of the genes. Perhaps, it would have been better to focus more on a single example (and only marginally mentions additional examples) rather than having a few rather unconnected examples that are only briefly described.

The approach used sounds conceptually very similar to motif analysis, and I'm a bit perplexed by the authors' effort to stress that it's very different. I see the analysis as complementary to classical motif analysis and I would encourage the author to include a more positive spin in the comparison rather than trying to stress the differences.
Experimental design: The testing approach sounds valid. I think some kind of flow diagram to describe the analysis would improve the readability but it's not necessary.
Validity of the findings: The algorithmic analysis sounds sound, but I'm a bit concerned about the validity of the biological findings. Picking one of the case studies and explore that more extensively would make the work more impactful.
Additional comments: The article is quite lengthy. A summary for the key findings would be definitely quite 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: NETWORK SUBGRAPH-BASED APPROACH FOR ANALYZING AND COMPARING MOLECULAR NETWORKS
Review round: 1
Reviewer: 2

Output:
Basic reporting: The greatest flaw currently is the lack of appropriate coverage to current literature, in particular, the work of the Algo Dyn Lab, for example:

- Algorithmic complexity of motifs clusters superfamilies of networks (https://ieeexplore.ieee.org/document/6732768)
- Evaluating network inference methods in terms of their ability to preserve the topology and complexity of genetic networks (https://www.sciencedirect.com/science/article/abs/pii/S108495211630012X?via%3Dihub)
- Quantifying loss of information in network-based dimensionality reduction techniques (https://academic.oup.com/comnet/article-abstract/4/3/342/1745204)

Arix versions are available in most cases in case the authors find some of the above behind a firewall.
Experimental design: The paper is clear and methodologically sound, although some of the measures proposed are weak.
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: NETWORK SUBGRAPH-BASED APPROACH FOR ANALYZING AND COMPARING MOLECULAR NETWORKS
Review round: 1
Reviewer: 3

Output:
Basic reporting: Table 1 is very wordy. Please consider it to be Supplementary Note.
Please reshape Tables 4-6 to make them clearer.
Please increase the resolution of Figures 2-5 to make the content clearer.
Experimental design: It is interesting that the authors propose a method for analyzing and comparing molecular networks with an information entropy measure.

PatternFinder [32] the authors developed is also based on the network subgraph approach. However, the authors do not mention about PatternFinder in "Introduction". For example, at line 83, the authors write "we propose the network subgraph-based approach ..." but they do not refer to [32] in this line. Moreover, lines 86-92 "We hypothesize that network subgraphs are the fundamental building blocks ..." mostly overlap the lines in the "Network subgraphs (N-node subgraphs) vs. network motifs" section in [32]. The authors need to mention that (at least a part of) their approach already proposed in [32] in "Introduction".

The authors also need to clarify the difference between the method proposed in this paper and PatternFinder described in [32]. The only difference seems to be that they adopt H_js as the network similarity metric. Please clarify if other differences exist or not.
Validity of the findings: In Table 2, the authors compare the results between PatternFinder, acc-Motif, and Loto. But the comparison seems to be similar as Table 2 in [32] except that the input data are different. If the authors would like to point out the superiority of PatternFinder, they can do so only by referring to Table 2 in [32]. If they address other points, they need to clarify the difference between them.

It is difficult to understand Figure 1 and Table 3 because very similar topologies (sometimes the same topologies) of the networks are classified into different classes in Figure 1 but the classification accuracies are 100% in Table 3. For example, the "scale-free graph" in (a) is the same as the "Aging-Random Graph 2%". Please explain about Figure 1 and Table 3 in detail.

The authors state that Figures 2-3 and Figures 4-5 are embedded in 3-node and 4-node subgraphs, respectively. However, it is difficult to identify 3-node or 4-node subgraphs in the figures. It may be useful to show the locations of the subgraph "id"s of Table 4 in Figures 2-5.
Additional comments: no comment.


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

Input:
Title: NETWORK SUBGRAPH-BASED APPROACH FOR ANALYZING AND COMPARING MOLECULAR NETWORKS
Review round: 2
Reviewer: 1

Output:
Basic reporting: The paper has been revised according to reviewers' comments. The reviewer has no further comments on the revised version of the paper.
Experimental design: No further comments.
Validity of the findings: No further 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: GENOME-WIDE IDENTIFICATION OF THE SWEET GENE FAMILY MEDIATING THE COLD STRESS RESPONSE IN PRUNUS MUME
Review round: 1
Reviewer: 1

Output:
Basic reporting: Zhenying Wen and coauthors present data on sequences with homology to SWEET proteins from the tree species Prunus mume and other Rosaceae species. The authors conduct phylogenetic analysis and expression analysis based on RNAseq and RT-qPCR analysis. In general, the comparison between a cold-sensitive and cold-tolerant accession of this interesting species opens the possibility for identifying factors responsible for cold tolerance. However, the presented study fails in the aspect to connect cold stress response and SWEET proteins and displays technical errors probably due to insufficient curation and presentation of used sequence material.
Experimental design: I think the major issue of the manuscript is that it is unclear if phylogenetic analysis of PmSWEETs have been done with complete and accurately curated sequences. Inspection of table 1 leaves the reader helpless to judge the quality of the used sequence material. After all, only six SWEETs possess seven transmembrane helices (TMHs), all other (except SWEET3) possess less. For all these SWEET types, seven TMHs have been reported in other species. Does this mean they are pseudogenes, unfunctional proteins or just incomplete sequences in Prunus mume? If so, any conclusion drawn from differential gene expression for a role of the respective transporter for cold tolerance would be not supported by their function.
Validity of the findings: I really think authors should manually inspect the extracted sequences and check for completeness and accuracy of the used sequence material. In this respect, I find it of paramount importance that the sequences of all analysed PmSWEETs been made available to the reader in a supplementary sheet and that both, coding sequences and TMHs be highlighted in these sequences. Without provision of the sequences, it impossible for the reader to comprehend the conducted analyses.

Some more comments:
The overall choice of SWEET protein family seems a bit arbitrary and needs better explanation and introduction already in the abstract since there is no real connection between SWEET activity and cold tolerance given. For example, other transporters mainly from the MST (monosaccharide transporter family) are much heavier involved in cold stress response and cold tolerance than SWEETs and would have probably been an equally important choice of in-depth analysis.
The statement in the abstract that SWEETs from different plant species divide into four group is not new (see Eom et al., 2015 Curr Op Plant Biology for Review) and does not explain the situation in P. mume.
Why do the authors classify “circadian control” as stress?
From the abstract, it is unclear why the mentioned accessions “Songchun” and “Zaolve” were chosen for comparative transcription analysis.
Additional comments: No additional comments


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

Input:
Title: GENOME-WIDE IDENTIFICATION OF THE SWEET GENE FAMILY MEDIATING THE COLD STRESS RESPONSE IN PRUNUS MUME
Review round: 1
Reviewer: 2

Output:
Basic reporting: Wen et al. investigated the SWEET family in the Prunus m. and studied the transcript expression in different conditions. This work was well designed and conducted while I have a few comments, hopefully, to improve this manuscript.
Although the writing is smooth, I would recommend the authors check the grammar with Naïve speakers. In addition, probably change the title to “Genome-wide identification of the SWEET gene family mediating the cold stress response in Prunus mume”
1. Line 18-20: the sentence may be revised as “and medicinal values; however, the distribution…..by low temperature.” And delete the “there were no…..about SWEET gene family in P. mune”. BTW, please make sure to use Italic when referred the gene name.
2. Line 34-38/Line 337-343: is “composited” in … In addition, the product of photosynthesis is sucrose (part of sugars) as well as involved in the phenotypical-metabolic changes and interaction with plant hormones, so I would recommend to authors to include few references and revise the sentence.
Lastdrager, J., Hanson, J. & Smeekens, S. Sugar signals and the control of plant growth and development. J. Exp. Bot. 65, 799–807 (2014); Ruan, Y. L. Sucrose metabolism: gateway to diverse carbon use and sugar signaling. Annu. Rev. Plant Biol. 65, 33–67 (2014); Wang, J.Y., Alseekh, S., Xiao, T. et al. Multi-omics approaches explain the growth-promoting effect of the apocarotenoid growth regulator zaxinone in rice. Commun Biol 4, 1222 (2021). https://doi.org/10.1038/s42003-021-02740-8; Sulpice, R. & McKeown, P. C. Moving toward a comprehensive map of central plant metabolism. Annu. Rev. Plant Biol. 66, 187–210 (2015). Barbier, F. F., Dun, E. A., Kerr, S. C., Chabikwa, T. G. & Beveridge, C. A. An update on the signals controlling shoot branching. Trends Plant Sci. 24, 220–236 (2019).
3. Line 276-312: the authors analyzed the promotors in the SWEET genes while the data indicated the cold temperature did not affect most of the genes; however, in figure 7, it looks like the temperature influenced the genes pattern, which might need a link or explanation. Moreover, the author claimed a similar expression pattern in the same period of different regions (fig 7C), but the data presented here did not fit. Please check it carefully.
4. In the qRTPCR figure, there is no statistical analysis that makes it difficult to assess the claim. Also, probably use two colors to distinguish the two genotypes.
5. Line 337-349/Line 359-366, this section was introduced at the beginning of the text (or at least similar), which looked wordy. Please revise it or shorten it.
6. I felt the discussion part is too dense to read which might make the reader confused. I would suggest the authors shorten and focus on “main” findings.
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: CLIMATE CHANGE INFLUENCES ON THE GEOGRAPHIC DISTRIBUTIONAL POTENTIAL OF THE SPOTTED FEVER VECTORS AMBLYOMMA MACULATUM AND DERMACENTOR ANDERSONI
Review round: 1
Reviewer: 1

Output:
Basic reporting: This is a good writing manuscript, that employed niche and distributional model techniques to estimate potential distributions of two vectors of spotted fevers under future climate. The manuscript incorporated recent advances in model development and validation, with climate change effects were estimated under different scenarios, and forecast associated uncertainty were explored …I think it merits acceptance and a fast publication.
I have some minor comments on the writing and style issue for further improvement.

Abstract:
Line 20: if Rickettsias is a genus name, it should be italicized. In the middle of this line, R. parkeri was first mentioned, but we don’t know whether the genus name is Rickettsias, as Rickettsias was not italicized, please check this out.
Line 28-32: I think it is not necessary to mention these states in the Abstract, they are too many and would distract reader, people not live in the U.S. might not familiar where are these states.

Introduction:
Line 47: pay attention where you first mention Rickettsia, and should be full name, after that it can be abbreviated.
Line 59-62: You are talking about distribution of these two vectors here, so it would good to cite Figure 1 here, see also my below comment on how to improve Figure 1.

Methods:
Line 86. I think it is better to put some information on (0,0), I know they are long and lat, but people might feel wield about these two “0” in brackets…or simply remove them?
Results:
Line 176-183: as I mentioned before, people not in the U.S. would not know where are these states, what if put abbreviations of these state name on Figure 1?

Discussion
I think it would be good to incorporate some discussion on common limitation of ENM in estimating vectors’ potential distribution, like climate niche conservatism, biotic interaction, and dispersal issue. For example, those blue areas identified as suitable habitat loss, but the tick might also survive there in future because of microclimate effect or adaption….my colleagues always complain that even we identified habitat unsuitable, but they still find the insect there, we should not underestimate insect evolve potential especially invasive species.

Comments on Figures:
Figure 1: I think it would be good to insert a scatter plot panel in this map, with the distribution (i.e. occurrence associate variables) of these two vectors in temperature and precipitations dimensions, with x axis is annual mean temperature and y is annual precipitation. It would demonstrate what you are talking the ecological preference of these two vectors in introduction in Line 59-62.
Figure 4: It would be good to insert a small overview panel in a corner of figure 4, to show reader where this map is showing…. It would be also good to mark “Illinois” and “Connecticut” states on this map, so people not in the US would know where these two states.

gengping.zhu@wsu.edu
Experimental design: Good, the manuscript incorporated recent advances in model development and validation, with many candidate models were calibrated.
Validity of the findings: Results are based on models that incorporated recent advances in niche and distributional modelling, they are robust and statistically sound.
Additional comments: None


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: CLIMATE CHANGE INFLUENCES ON THE GEOGRAPHIC DISTRIBUTIONAL POTENTIAL OF THE SPOTTED FEVER VECTORS AMBLYOMMA MACULATUM AND DERMACENTOR ANDERSONI
Review round: 1
Reviewer: 2

Output:
Basic reporting: In the case of English, I reserve comments. Two references would be missing that could notably improve the manuscript Pascoe et al. 2019 and Fernando Cuervo et al. 2021. In the case of the hypothesis, it is not well reflected in the text.
Experimental design: no comment. It seems to me that they meet the requirements.
Validity of the findings: no comment. These medically important species provide evidence to make better use of control measures and the information provided is very relevant.
Additional comments: Summary
In the Coltivirus species, add spp. I think that in the summary, they do not mention anything about the distribution of these species in Mexico.

Keywords, maybe we could add North America

Line 43. Must say are three of the ticks
Line 88. Only explain in better detail, the reason why they chose the distance of 50 km, (dispersion of the tick? Or some other behavioral reason?).
Line 91. In what way did you divide the data 50-50, did you use any procedure?
Line 99. Justify the use of the 200km buffer?
Line 104. The pixel size is very large. Is there a difference between using the 5 minute size and the 10 minute size?
Line 116. In the last update of March 2020, the new CMIP6 scenarios were made available, which difference exists between SSPs and RCPs; can we observe the same results?
Line 176. Although the geographic location in the United States is mentioned, the stable ecological niche in Mexico is not mentioned.
Line 232. I think that although Fernando Cuervo et al. 2021, does not show the results of the models, but it is very important to mention in relation to the dispersion of niche conservatism that occurs with A. triste and A. trigrinum; therefore, the part of migration would be important in the North American region.


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: CLIMATE CHANGE INFLUENCES ON THE GEOGRAPHIC DISTRIBUTIONAL POTENTIAL OF THE SPOTTED FEVER VECTORS AMBLYOMMA MACULATUM AND DERMACENTOR ANDERSONI
Review round: 2
Reviewer: 1

Output:
Basic reporting: The authors have addressed all my concerns, current revisions look good to me, indeed the manuscript is much improved in this revision, either in writing or figures, which clear demonstrated ecological dimensions and potential distribution, at present under future climate scenarios.
I think it can be considered for publication.
Experimental design: None
Validity of the findings: None
Additional comments: None


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: CLIMATE CHANGE INFLUENCES ON THE GEOGRAPHIC DISTRIBUTIONAL POTENTIAL OF THE SPOTTED FEVER VECTORS AMBLYOMMA MACULATUM AND DERMACENTOR ANDERSONI
Review round: 2
Reviewer: 2

Output:
Basic reporting: The manuscript has many improvements throughout the document. A few small comments are added to the review. The manuscript is ready to be accepted in its form with the suggested modifications.

Line 97. I understand the tick dispersal part and the host part, but I would like you to add the reference to support that argument.

Line 182: From the southeast of Mexico, only mention the state of Quintana Roo

Line 203: match the word west ern

Fig1. The word Dermacentor is misspelled.
Experimental design: The manuscript has many improvements throughout the document. A few small comments are added to the review. The manuscript is ready to be accepted in its form with the suggested modifications.

Line 97. I understand the tick dispersal part and the host part, but I would like you to add the reference to support that argument.

Line 182: From the southeast of Mexico, only mention the state of Quintana Roo

Line 203: match the word west ern

Fig1. The word Dermacentor is misspelled.
Validity of the findings: The manuscript has many improvements throughout the document. A few small comments are added to the review. The manuscript is ready to be accepted in its form with the suggested modifications.

Line 97. I understand the tick dispersal part and the host part, but I would like you to add the reference to support that argument.

Line 182: From the southeast of Mexico, only mention the state of Quintana Roo

Line 203: match the word west ern

Fig1. The word Dermacentor is misspelled.
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: THE ASSOCIATION OF PLASMA OSTEOPROTEGERIN LEVELS AND FUNCTIONAL OUTCOMES POST ENDOVASCULAR THROMBECTOMY IN ACUTE ISCHEMIC STROKE PATIENTS: A RETROSPECTIVE OBSERVATIONAL STUDY
Review round: 1
Reviewer: 1

Output:
Basic reporting: The paper is clearly written and well organized.
Experimental design: 1.	In general, this primary research is within the aims and scope of the journal. The experimental design was clearly written in a structured manner.
2.	However, I suggest the authors describing the assessment of outcome variable, i.e., mRS score (Line 125), e.g., by whom (inter-rater reliability if there is more than one assessor), using what platform (face-to-face visit, or through teleconsultation). This description is closely related to the potential information bias in the study.
3.	More information is needed to show that this research was conducted in conformity with the prevailing ethical standards in the field, e.g., the issue of informed consent during the data collection process, impact of withdrawal from participation, methods to ensure privacy and confidentiality in the stage of data management.
Validity of the findings: 1.	The data analysis is statistically sound, however insufficient information to report the specific selection of test (Table 1). I suggest the authors using footnote to represent which p-value was obtained from chi-square test, independent t-test, etc.
2.	Also, the information about the normality of the continuous variables is contradicting. Line 142-143 (continuous variables in our dataset were normally distributed) contradicts with the presentation of NIHSS scores in median and interquartile range (Line 169 and Table 2).
3.	Parameters showing model fitness e.g., area under ROC curve might be useful for model comparison in multivariable binary logistic regression analysis (Table 2).
4.	It is interesting to include the original logistic regression analysis with mRS (the original mRS score without dichotomization) as a dependent variable, perhaps the authors could present that result (line 186).
5.	The conclusions are not comprehensive, in which it could be linked to the research gap highlighted (Line 57-58) and how the study contributes to filing that gap e.g., in the current clinical practice of in-hospital stroke management with EVT.
Additional comments: This is a well-conducted research, and its contribution is desirable for future work.


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

Input:
Title: THE ASSOCIATION OF PLASMA OSTEOPROTEGERIN LEVELS AND FUNCTIONAL OUTCOMES POST ENDOVASCULAR THROMBECTOMY IN ACUTE ISCHEMIC STROKE PATIENTS: A RETROSPECTIVE OBSERVATIONAL STUDY
Review round: 1
Reviewer: 2

Output:
Basic reporting: Title of the study: Rewrite the title as it already stated the finding rather than the hypothesis of the study. It is suggested to modify it into “Association of Plasma Osteoprotegerin levels with Functional Outcomes in Acute Ischemic Stroke Patients who have undergone Endovascular Thrombectomy”

Abstract: Simplified the result and strengthen the methodology input.

Introduction: The information aims to justify the rationale of conducting the study. However, we notice that the info is inadequate. Readers want to know more about the association between OPG and stroke, especially during the acute phase. How these conditions can be associated with and determined the prognosis of the stroke patient? Does the level of plasma OPG change during EVT? What are the conditions that cofounded the level of OPG?
Experimental design: Method and tool: This section needs to be elaborated in detail and relevant to the topic. The authors need to show the robustness of the methodology conducted. Information related to study design, study location, study criteria, selection of the sample, and sample size determination is not mentioned in the manuscript. Please simplify the info under the subheading “Covariates” so that the readers could understand it better. Please elaborate on how the outcome of the study was measured. We notice that there is a discrepancy of information return in the text and in Figure 1. Please choose the correct one.
Validity of the findings: Result: Please explain in the legend what you mean by three different models in Table 2 and 3. How did you determine the fitness of those models?

Discussion: You do it well the discussion by citing the up-to-date references. However, we could not find any recommendation derive from your finding. Please provide significant recommendations that might help in managing acute stroke cases in the future. We also encourage you to give acknowledgment to those involved either directly or indirectly in the study.
Additional comments: Thank you for giving me the opportunity to review this interesting work. Your aim is to look at the association between plasma oesteoprotegrin level and functional outcome in stroke patients.
You did the writing very well but still, there are spaces 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: THE ASSOCIATION OF PLASMA OSTEOPROTEGERIN LEVELS AND FUNCTIONAL OUTCOMES POST ENDOVASCULAR THROMBECTOMY IN ACUTE ISCHEMIC STROKE PATIENTS: A RETROSPECTIVE OBSERVATIONAL STUDY
Review round: 1
Reviewer: 3

Output:
Basic reporting: Adequate
Experimental design: Acceptable
Validity of the findings: Good findings
Additional comments: -	Study design not mentioned in the title. Instead of writing the positive association in the title, suggest to revise the title to mention the objective that was mentioned in the last paragraph in the Introduction section
-	Abstract is acceptable
-	Introduction is adequate
-	In Methods, please mention the type of study (Cross sectional)
-	In Methods, to detail out steps taken to reduce sources of bias
-	In Methods, to explain how study size was arrived at
-	Results and Tables are adequate
-	In Discussion, summarize the key findings in the first paragraph in reference to the study objectives
-	To put in the Ethics clearance for this study and funding (if any)


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: THE ASSOCIATION OF PLASMA OSTEOPROTEGERIN LEVELS AND FUNCTIONAL OUTCOMES POST ENDOVASCULAR THROMBECTOMY IN ACUTE ISCHEMIC STROKE PATIENTS: A RETROSPECTIVE OBSERVATIONAL STUDY
Review round: 2
Reviewer: 1

Output:
Basic reporting: No comment.
Experimental design: The methodology is very much improved, however there is room for improvement.
1. Objective - suggest to use the word 'associated/association' instead of 'positively correlated' (Line 82).
2. Study design - cross-sectional study isn't appropriate as the exposure (OPG) preceds the outcome (3-month mRS).
3. Subjects - information on sampling method is lacking (Line 95) and suggest to add the definition or diagosis of acute ischemic stroke; e.g., ICD -10 coding, or clinical diagnosis or radiological diagnosis (Line 108)
4. Subgroup analysis based on door-to-puncture time, suggest to justify the rational of conducting this subgroup analysis, preferably with supporting evidence - this should be mentioned under the analysis of subheading methods.
5. Results - suggest to report the standard deviation of age (Line 191), and you may consider baseline mrs as the covariate in the modelling.
6. 'positively associated' (Line 247) isn't appropriate, suggest to remove the word 'positively'
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: THE ASSOCIATION OF PLASMA OSTEOPROTEGERIN LEVELS AND FUNCTIONAL OUTCOMES POST ENDOVASCULAR THROMBECTOMY IN ACUTE ISCHEMIC STROKE PATIENTS: A RETROSPECTIVE OBSERVATIONAL STUDY
Review round: 2
Reviewer: 2

Output:
Basic reporting: Literature references, sufficient field background/context provided.
Experimental design: Methods described with sufficient detail & information to replicate. I am satisfied with all of the changes made.
Validity of the findings: All underlying data have been provided; they are robust, statistically sound, & controlled
Additional comments: The authors clarified all of the remarks, which I found to be satisfactory and appropriate.


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: THE ASSOCIATION OF PLASMA OSTEOPROTEGERIN LEVELS AND FUNCTIONAL OUTCOMES POST ENDOVASCULAR THROMBECTOMY IN ACUTE ISCHEMIC STROKE PATIENTS: A RETROSPECTIVE OBSERVATIONAL STUDY
Review round: 2
Reviewer: 3

Output:
Basic reporting: Clear and corrections have been made
Experimental design: All issues commented in the previous review have been addressed
Validity of the findings: Valid and statistically sound
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: MIR-9-5P PROMOTES MYOGENIC DIFFERENTIATION VIA THE DLX3/MYF5 AXIS
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript entitled “miR-9-5p promotes myogenic differentiation via Dlx3/Myf5 axis” by Liying Dong et al. identified a novel pathway that miR-9-5p induced myogenic differentiation by targeting Dlx3 3-UTR to inhibit its binding to Myf5 promoter. The study is of clinical significance and provides potential therapeutic targets.
Experimental design: 1.	Authors well studied the miR-9-5p/DLX3/Myf5 axis in mouse myoblast-derived C2C12 cell line. Major experiments should be reproduced in a second cell line.
2.	In Figure 2C, miR-9-5p mimics increased myotube numbers compared to NC mimics while miR-9-5p inhibitor decreased myotube numbers compared to NC inhibitor. Why were myotube numbers so different between two controls, NC mimics (about 20) and NC inhibitor (about 60) groups? Authors should present these four groups in one panel to compare myotube numbers.
3.	In Figure 4C, siDLX3 alone inhibited expression of DLX3. Why did co-transfection with siMyf5 increase DLX3 level?
4.	In Methods, Statistical analysis should be elaborated. It is too simply just stating “We used one-way ANOVA analysis or t-test to determine statistical significance”. For example, in Figure 1B and Figure 4B, C and E, the statistical method should be properly described. One-way ANOVA is used to test whether it is statistically different among all compared groups. Multiple comparison is followed to test which two groups are different.
Validity of the findings: In Figure 1B, miR-9-5p was increased on day 2 and then back to the initial level on days 4 and 6. What is the interpretation? The data does not support the conclusion that the increased miR-9-5p expression participates the myogenic differentiation.
Additional comments: None.


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: MIR-9-5P PROMOTES MYOGENIC DIFFERENTIATION VIA THE DLX3/MYF5 AXIS
Review round: 1
Reviewer: 2

Output:
Basic reporting: In this study, Dong et al proposed that the myogenic differentiation of C2C12 cells was regulated by miR-9-5p/Dlx3/Myf5 axis. They found Dlx3 was decreased, whereas miR-905p was increased, during myogenic differentiation. And miR-9-5p promoted myotube formation via decreasing Dlx3. In addition, the suppression of the myogenic differentiation in C2C12 cells were mediated by Dlx3, and that was associated with its capacity in reducing Myf5. This is a straightforward study. It is easy to follow. Following are the comments:
1. Language editing is needed. I would suggest the authors to seek help from a native English speaker.
2. The statistical method used in this study is improper. ANOVA only tells whether there are significant differences between all compared groups. Post-hoc test is needed to further clarify where the differences are. In addition, please specify which type of t-test was used in this study, paired or unpaired.
3. Immunofluorescence staining of muscle protein markers during indicated time points is recommended. For instance, the authors can use alpha-actinin or MHC as markers
4. The biggest drawback of this study is that the changes of protein levels in western blot were not as robust as which were described in the quantification plots. For example, the change of Myod1 was barely visible after miR-9-5p inhibitor treatment in Figure 2B. However, the authors indicated there was significant decrease in their quantification plot. I would strongly suggest the authors to reverify there western blot results. Almost every WB data has this issue. In addition, the Myf5 bands in Figure 2B are hazy.
5. According to the data, Myod1 was also changed after miR-9-5p mimic and inhibitor treatment, and after knockdown or overexpressing Dlx3.  Why did the authors were more interested in Myf5?
6. I would suggest the authors to prove if modulating the expression of Myf5 alone is enough to delay or facilitate the myogenic differentiation. Otherwise, the proposed regulatory flux will be compromised.
Experimental design: Please see details in Basic reporting.
Validity of the findings: Please see details in Basic reporting.
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: MIR-9-5P PROMOTES MYOGENIC DIFFERENTIATION VIA THE DLX3/MYF5 AXIS
Review round: 1
Reviewer: 3

Output:
Basic reporting: MiR-9-5p is a popular microRNA and well studied in human cancers, fibrosis development and other diseases. However, its function in myogenic differentiation is rarely explored. The manuscript entitled “miR-9-5p promotes myogenic differentiation via Dlx3/Myf5 axis” unveiled miR-9-5p prompts bone formation through suppress Dlx3 mRNA expression, while increases Myf5 which is an essential transcript factor of myogenic differentiation. All the experiments are well designed and convincing. However, several question should be illustrated and minor modified to improve the paper before publication.

1.	In figure 1A, the bar value should be noted, so do figure2C and figure 3C&F. And in figure 1B&C should notify mRNA level fold change in Y axis.
2.	Fig2B, why the western blot of Myf5 is so obscure, I couldn’t tell the difference. Please run this experiment again.
3.	In fig2C, why NC mimics and NC inhibitors are so dramatically different? Is because the cells are at different time point or something else, please demonstrate this in the paper?
4.	All the western blot results should be quantified.
Experimental design: no comment
Validity of the findings: no comment
Additional comments: no 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: MIR-9-5P PROMOTES MYOGENIC DIFFERENTIATION VIA THE DLX3/MYF5 AXIS
Review round: 2
Reviewer: 1

Output:
Basic reporting: Authors have successfully responded to my comments. My only concern is the lack of the second cell line to valid their key findings. This drawback will significantly decrease the impact and validity of the manuscript. It is a poor justification that "Most papers only used C2C12 in their studies".
Experimental design: None.
Validity of the findings: None.
Additional comments: None.


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: MIR-9-5P PROMOTES MYOGENIC DIFFERENTIATION VIA THE DLX3/MYF5 AXIS
Review round: 2
Reviewer: 2

Output:
Basic reporting: The authors have improved their manuscript according to my comments. If it is applicable, this manuscript can be considered for publication in this current version.
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: MIR-9-5P PROMOTES MYOGENIC DIFFERENTIATION VIA THE DLX3/MYF5 AXIS
Review round: 2
Reviewer: 3

Output:
Basic reporting: All the questions are clearly answered and well revised. I do recommend this manuscript could be accepted.
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: A NEW COMPREHENSIVE APPROACH FOR REGIONAL DROUGHT MONITORING
Review round: 1
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: The authors in this study proposed a Regional Intensive Continuous Drought Probability Monitoring System (RICDPMS) method to obtain quick and comprehensive information regarding the drought probability and the temporal evolution of the droughts at the regional level, which was accomplished by using Monte Carlo Feature Selection (MCFS), steady-state probabilities, and copula-based bivariate analysis for different drought intensities.
Validity of the findings: The paper showed through its application in northern area of Pakistan that the RICDPMS could monitor the region's drought and provide a better quantitative way to analyze drought with varying drought intensities in the region. Therefore, this study has a certain meaning in practical applications and it is recommend accepting this manuscript.
Additional comments: Yet I still have several specific concerns for authors to improve their manuscripts.
1) The keywords included “drought risk monitoring”. However, there was no description of drought risk in the introduction and methods. Can you add relevant research in the introduction and explain the specific calculation method of drought risk in this study?
2) Lines 90-93 introduced that the precipitation in the study area was concentrated from January to June. Therefore, the drought in this period should be the lowest frequency and lowest intensity throughout the year. However, this paper selected only the rainy season for drought monitoring and analysis, please explain why, or does it have more practical significance and value to extend the research time series to the whole year?
3) In this paper, the MCFS method was used to represent the data of a certain station in the region to the entire research area. The intention was to reduce time and capital investment while maintaining the accuracy of the results. However, whether the analysis results of the comprehensive data obtained after using this method were the same or similar as the results obtained by the complete data using the traditional method? I hope the authors to add this comparison to verify the reliability and accuracy of using this method.
4) Please explain the reasons for choosing SPI-6 instead of other time scales as the drought indicator.
5) The title of Figure 1 is “Geographical locations of the six selected stations of the region of Pakistan”. However, the specific location of the meteorological stations were not marked, please add it to the map.
6) Line 119: Please check the title format and delete extra punctuation.
7) Line 226-228: Please check whether the sentence expression is correct.
8) Line 250: Please elaborate on the meaning of "climatological precipitation". Is it the average rainfall for many years or the rainfall threshold or something else?


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

Input:
Title: A NEW COMPREHENSIVE APPROACH FOR REGIONAL DROUGHT MONITORING
Review round: 1
Reviewer: 2

Output:
Basic reporting: The authors proposed a Regional Intensive Continuous Drought Probability Monitoring System (RICDPMS) method to obtain quick and comprehensive information regarding the drought probability and the temporal evolution of the droughts at the regional level, as a study of the Pakistan.
The topic is of great regional importance and interesting results are shown in the paper, which could help readers to monitor the drought in a regional level. However, a few things need to be clarified before being considered for publishing. My further comments below contain some suggestions for extensions to the current manuscript.
Experimental design: Your manuscript structure needs clearer: “3. Results. 4. Discussion.”
Validity of the findings: no comment
Additional comments: 1. The abstract needs improvement. You should be sure that the abstract provides a concise problem statement and briefly explains the main findings and their significance.
2. The figures needs more detail in this manuscript. A figure is worth a thousand words. I suggest that you improve the figures in this manuscript to provide more visual information of the drought monitoring.
3. RICDPMS is the modification of the CDPMS (Pontes Filho et al., 2019). What are the advantages and improvements in RICDPMS? I suggest that you add the discussion to provide more justiﬁcation for your study, speciﬁcally, you should compare the drought monitoring effects of the two methods.


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

Input:
Title: A NEW COMPREHENSIVE APPROACH FOR REGIONAL DROUGHT MONITORING
Review round: 2
Reviewer: 1

Output:
Basic reporting: ok
Experimental design: ok
Validity of the findings: ok
Additional comments: The authors have explained the questions I raised in detail and revised the unreasonable parts accordingly. Now the structure of the article is clarity and the application validated that the Regional Intensive Continuous Drought Probability Monitoring System has a certain meaning in practical applications. Therefore, it is recommend accepting this manuscript in its current version.


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

Input:
Title: A NEW COMPREHENSIVE APPROACH FOR REGIONAL DROUGHT MONITORING
Review round: 2
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The authors have addressed all my comments. I do not have any further suggestion.


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: SPATIOTEMPORAL CHANGES OF EUTROPHICATION AND HEAVY METAL POLLUTION IN THE INFLOW RIVER SYSTEM OF BAIYANGDIAN AFTER THE ESTABLISHMENT OF XIONGAN NEW AREA
Review round: 1
Reviewer: 1

Output:
Basic reporting: Titile. The “water pollution” is too vast, however, the water quality of eutrophication and heavy metals is studied in this manuscript. The title should be clarified and specified.

Section Introductin. The water pollutants, including chemical oxygen demand (COD), ammonia nitrogen (NH4+-N), total phosphorus (TP), plumbum (Pb), cadmium (Cd) and chromium (Cr) should be described and emphasize in Introduction section to clarify the purpose of this research.

Lines220-221. “In spring, NH4+-N in F6 and COD in F6, 11 were worse than Class Ⅴwater standard”.What is the meaning of “worse”? The concentration of WQI parameter may be higher/lower than the WQI guideline/standard.

The pie chart in the Fig.2 is meaningless without the water discharge flux of Fuhe river.
Experimental design: Lines 151-152. “River water were sampled in three seasons: spring (May, headstream was not collected), summer (July) and autumn (October) of 2020, respectively”. Please clarify the reseaons to sample water in the three seasons. Why choose May, July and October? Maybe the “headstream” is not the “head” of Fuhe.

The water transfer from water reservoirs in the Mountain area to Baiyanddian should not be eliminated in the FRS.
Validity of the findings: This research in meaningful for water quality assessment in Baiyangdian lake.
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: SPATIOTEMPORAL CHANGES OF EUTROPHICATION AND HEAVY METAL POLLUTION IN THE INFLOW RIVER SYSTEM OF BAIYANGDIAN AFTER THE ESTABLISHMENT OF XIONGAN NEW AREA
Review round: 1
Reviewer: 2

Output:
Basic reporting: Clear and unambiguous, professional English used throughout.
Literature references, sufficient field background/context provided.
Professional article structure, figures, tables. Raw data shared.
Self-contained with relevant results to hypotheses.
Experimental design: Original primary research within Aims and Scope of the journal.
Research question well defined, relevant & meaningful.
Why only 3 heavy metals, lead, cadmium and chromium are studied, and it is recommended to further review the pollution of other heavy metals
Sampling shall be conducted by the corresponding sampling method of China Standard (GB3838-2002), or by the standard method of EPA
Validity of the findings: There are many literature on the water environment and water ecology of Baiyangdian Lake, and it is suggested to further deepen the review
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: SPATIOTEMPORAL CHANGES OF EUTROPHICATION AND HEAVY METAL POLLUTION IN THE INFLOW RIVER SYSTEM OF BAIYANGDIAN AFTER THE ESTABLISHMENT OF XIONGAN NEW AREA
Review round: 1
Reviewer: 3

Output:
Basic reporting: line 68: change to "due to the measures not taking serious consideration"

line 93: change to "In recent years", and please also clarify the sentence on line 95.

line 107: change to "critically"

line 125: change to "We aimed to"

line 151: change to "River water was"

line 180: change to "due to it being"

line 193: change to "classify". On the next line, please clarify this sentence.

line 198: delete the second "of"

line 243: change to "flowed", also on line 248.

line 283: change to "is needed"

line 294: change "were" to "was"

line 301: change to "threats"

line 309: is the word "processive" here correct?

line 318: change to "showed that the pollution"

line 320: delete "were"

line 323: change to "Greece"

line 329: change to "than what we found in"

line 337: change to "contaminate". On the next line, change to "ameliorate"

line 359: change to "In contrast". Also on line 398.

line 362-363: I do not understand this sentence.

line 367: change "form" to "from"

line 410: change to "recent decades" or "the last decade"
Experimental design: line 342: There were no replicate seasons sampled (i.e. spring, summer and autumn were only sampled during one year), so an experimental design with this limitation cannot be used to understand with consistency any information about seasonal variation. For this reason, I think the word "seasonal" here and else where (e.g. line 345, etc) should be changed to "temporal". I would say that a section of text should be included discussing how there may be seasonal patterns, but until further research has been done with seasonal replication (at least two or three summers, two or three autumns etc) it cannot be known whether the temporal variation seen here is associated with different seasons, or alternatively it may be associated with many other factors that also change across different times.
Validity of the findings: line 198: you should check the assumption of homogeneity of variances for the ANOVAs using Levene's or Cochran's tests. If this assumption is not met, then there may be some transformation of the data required that may change analysis results and affect the validity of the findings.
Additional comments: line 22: it would be good to have some explanation of what this "New Area" is. Is it a newly developed area, or a newly rehabilitated area? etc.

Line 258-261: I do not understand the relevance of correlating different heavy metals and eutrophic parameters with PC1 and PC2. How does this provide useful information? Normally when I do PCA analyses, I use the ordination along the PC axes to correlate different types of variables, e.g. the water quality parameters with samples from different locations, or something like that.

Line 389-392: could you please explain a bit more clearly how exactly the association of Pb with different principal components allowed the conclusion that "Pb in FRS could be attributed to the sources of traffic pollution, agricultural practices and historical polluted sediments rather than industrial wastewater discharges".

Figure 3: it would be good for these graphs to have some indication of whether differences among the sampling areas and/or sampling times are significantly different.

Figure 5: the horizontal dotted line is difficult to see.


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: SPATIOTEMPORAL CHANGES OF EUTROPHICATION AND HEAVY METAL POLLUTION IN THE INFLOW RIVER SYSTEM OF BAIYANGDIAN AFTER THE ESTABLISHMENT OF XIONGAN NEW AREA
Review round: 2
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The revision was sufficiently revised according to the reviewers' comments. I think the revision is acceptable 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: SPATIOTEMPORAL CHANGES OF EUTROPHICATION AND HEAVY METAL POLLUTION IN THE INFLOW RIVER SYSTEM OF BAIYANGDIAN AFTER THE ESTABLISHMENT OF XIONGAN NEW AREA
Review round: 2
Reviewer: 2

Output:
Basic reporting: There were just a few more changes needed to fine-tune the use of English language:

line 206: change to something like: "After the data of pollutant concentrations in three seasons were combined with the GPS positioning of each sampling site, Arc-GIS was used"

line 232: change to "are shown in Fig. 2"

line 357: change to "We did not find any previous study that has shown"

line 411: change to "confirming"

line 447: change to "In addition, dramatic spatiotemporal changes of pollution in FRS were found in this study, allowing the conclusions:"
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: INTEGRATED LIPIDOMICS AND RNA SEQUENCING ANALYSIS REVEAL NOVEL CHANGES DURING 3T3-L1 CELL ADIPOGENESIS
Review round: 1
Reviewer: 1

Output:
Basic reporting: Authors carried out in-depth study focusing on the lipidomic and transcriptional changes during maturation of 3T3-L1 cells. The study is straightforward yet important, deepening our understanding of adipogenesis, and could thus stimulate future studies to unveil obesity related disease in human. Most of my concern is regarding the formatting of the manuscript. Authors should try to address the following comments and check thoroughly for any similar errors.
Experimental design: no comments
Validity of the findings: no comment
Additional comments: 57： “The medium containing with DMEM” does not make sense. I suppose authors used DMEM supplemented with FBS et ac. Please change. Also please specify the manufacturer of materials used in the study.
68: microscope manufacturer.
70: could authors specify how many cells were used for each sample?
112: “by using” is a redundant expression. Remove by or using
116: what is MS2 metabolites?
127: MS2 or M2S?
128: “301 metabolites were up”: readers will understand what the authors mean, but this is not a proper expression. Same as the rest of this paragraph. Authors could use increased and decreased instead of up and down.
201: please define slc2a4 where it appears the first time in the experiment. Please check the whole ms for errors like this.
257: please state the full name of the cell line.
261: Authors should be more confident about their study. It will provide basic knowledge for interested readers in related subjects for sure. Lease remove “may”.
Discussion: authors should discuss briefly how the present study might benefit future studies in relation to obesity development, prevention, and treatment of obesity-induced diseases.
Figure1: scale bar should be added to each image.


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: INTEGRATED LIPIDOMICS AND RNA SEQUENCING ANALYSIS REVEAL NOVEL CHANGES DURING 3T3-L1 CELL ADIPOGENESIS
Review round: 1
Reviewer: 2

Output:
Basic reporting: The manuscript 'Integrated lipidomics and RNA sequencing analysis reveal novel changes during 3T3-L1 cell adipogenesis' from Yangli Pei et al systematically analyzed the molecular changes from preadipocyets to mature adipocytes. The finings of this manuscript are foundamentally important to reveal the the underlying mechanisms for adipogenesis. I have several comments mainly for the figures presented.
Experimental design: Good experiment design to address a basic but essential question regarding adipogenesis.
Validity of the findings: 1. Figure 1: Scale bars are missing from cell culture pictures. And did the  D8 and D10 pictures had the same magnitude?
2. According to Figure 4, the biochemical pathway enrichment for differently expressed lipids indicates minor effects on regulation of lipolysis in adipocytes. But in contrast, authors wrote from line 145-148 that differently regulated lipids molecules in adipocytes related to" regulation of lipolysis in adipocytes". If lipolysis process indeed changes significantly in mature adipocytes, did the authors quantify the lipolysis between preadipocytes and mature adipocytes? or the genes expression for lipolysis?
3. Figure 6 was confusing to distinguish qPCR data from RNAseq data. In addition, statistical analysis is missing from this Figure.
4. Figure 7: KEGG enrichment results shows the pathway enrichment showed the thermogenesis pathway. This is very interesting, did the author checked the KEGG, what caused the significant increases in thermogenesis?
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: INTEGRATED LIPIDOMICS AND RNA SEQUENCING ANALYSIS REVEAL NOVEL CHANGES DURING 3T3-L1 CELL ADIPOGENESIS
Review round: 1
Reviewer: 3

Output:
Basic reporting: Actually，there are studies on the lipidome and transcriptome profile alterations during the adipocyte adipogenesis Zhang et al.(2018, Int J Mol Sci), Sun et al. (2020, Front Mol Biosci) and Popkova et al. (2020, Analytical and Bioanalytical Chemistry). However, it is meaningful to analyze the molecular mechanism of adipocyte differentiation by integrating these two omics.
Experimental design: 1.There are some obvious mistakes that could have been eliminated by a simple revision of the manuscript. For example, spacing after the number (page9, 3.3, line155), number before the title (Materials and Methods 2.4-2.6), etc.
2.The author needs to provide the reference related to the the criteria for screening differential lipid molecules.
3. Because the number of samples in transcriptome and lipidomes analysis is different, it is important for the author to explain clearly how to carry out correlation analysis in section 2.6
Validity of the findings: 1.line 116-line127, MS2 and M2S mean? Please confirm.
2.It is suggested to provide the magnification or scale of Oil red O staining pictures in Figure 1.
Additional comments: 1.Put table1 in supplemental material.
2.It is suggested that the authors compare their results with other publised lipidome and transcriptome data of 3T3-L1 cells, and put them into the discussion.


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: UNIFIED SYSTEM DESCRIBING FACTORS RELATED TO THE ERADICATION OF AN ALIEN PLANT SPECIES
Review round: 1
Reviewer: 1

Output:
Basic reporting: Regarding basic reporting, I feel the article needs revision before it can be published.

The English writing needs to be improved to improve readability and reduce ambiguity.

I think the article will need to be reframed. I feel that standardising the terms used for factors that may influence eradication should not be the primary focus. What is more important is developing a standardised approach to scoring eradication success that this easy to use. I think the authors make good progress with this, but more specificity is also needed for each variable to allow for standardisation and comparison between sites.

I think the framing and objectives of this work need to be more clearly thought out and stated. (I think standardising terminology, should not be a core objective, but rather standardising socring).

The authors need to should how their work builds on previous studies e.g. Dodd, Gardnerner (a suggested new reference)
Experimental design: I think the review of factors is good. A little more is needed to show how the author's systems build on that of previous work especially Dodd et al.

I feel the new approach needs to be tested for more eradication projects globally. I think this will help to develop methods and metrics to improve standardisation to allow for comparison globally.

I think that for each factor standardisation is needed, and different catalogers to choose from should be offered for each one. Linking to this a confidence score should also be added for each.
Validity of the findings: I think the findings and introduction and conclusion do not link so well and there is a need for some reframing (more about standardised scoring). Based on this I feel the authors should be reading and getting inspiration from other scoring and reporting frameworks (not just those based on eradication). Some examples include:

Volery, L., Bacher, S., Blackburn, T.M., Bertolino, S., Evans, T., Genovesi, P., Kumschick, S., Roy, H.E. and Smith, K.G., 2020. Improving the Environmental Impact Classification for Alien Taxa (EICAT): a summary of revisions to the framework and guidelines.

Bacher, S., Blackburn, T.M., Essl, F., Genovesi, P., Heikkilä, J., Jeschke, J.M., Jones, G., Keller, R., Kenis, M., Kueffer, C. and Martinou, A.F., 2018. Socio‐economic impact classification of alien taxa (SEICAT). Methods in Ecology and Evolution, 9(1), pp.159-168.

Blackburn, T.M., Essl, F., Evans, T., Hulme, P.E., Jeschke, J.M., Kühn, I., Kumschick, S., Marková, Z., Mrugała, A., Nentwig, W. and Pergl, J., 2014. A unified classification of alien species based on the magnitude of their environmental impacts. PLoS biology, 12(5), p.e1001850.

Wilson, J.R., Faulkner, K.T., Rahlao, S.J., Richardson, D.M., Zengeya, T.A. and Van Wilgen, B.W., 2018. Indicators for monitoring biological invasions at a national level. Journal of Applied Ecology, 55(6), pp.2612-2620.

Pheloung, P.C., Williams, P.A. and Halloy, S.R., 1999. A weed risk assessment model for use as a biosecurity tool evaluating plant introductions. Journal of environmental management, 57(4), pp.239-251.

Shackleton, R.T., Bertzky, B., Wood, L.E., Bunbury, N., Jäger, H., van Merm, R., Sevilla, C., Smith, K., Wilson, J.R., Witt, A.B. and Richardson, D.M., 2020. Biological invasions in World Heritage Sites: current status and a proposed monitoring and reporting framework. Biodiversity and Conservation, 29(11), pp.3327-3347.
Additional comments: I have reviewed the paper looking at a unified system for describing factors related to the deracination of plant species. I think the paper has merit but I have some reservations at the moment and believe work is needed to improve the manuscript. I think the study is not so well framed and makes it a bit hard to follow and see the value of the work. I also feel it needs to be standardized more (a choice of categories for each factor, and better tested using more case studies globally).

Firstly, you make an argument for a unified system of terminology; however, I am not if there is any controversy or many issues with the use of the terms included in table one. Yes, people call certain things differently which is ok, there are just variables for analysis of the potential success of an eradication campaign. I am certain the lack of standardization is not causing any major problems.  There is more contention over core definitions like the word “eradication” itself that is an issue.  I am not sure “unifying” the names of factors that could affect eradications is that necessary or important. Previous “unifying” studies have tried to merge different frameworks or standardize key terms that are used massively like “invasion” “eradication”, “naturalization” “management”, “pathway” etc.  I would rather frame this article as a review and consolidation of factors that influence the eradication feasibility of plants. I would also go further and make it a detailed scoring tool see below). I think this is where the interest will be and less so on terminology standardization. Linking to this comment and list of variables, the authors cite Dodd et al. (2015) who identify 22 factors. Their final set of variables proposed here is 24. Based on this I am not sure how much this work actually adds in this regard either. The needs to be elaborated on and your contributions to science need to be better highlighted. Linking to this as well in your article you choose not to define the values or categories for the factors, however, to collect comparable data this is essential and I believe that through the literature review you did you could come up with meaningful categories. If this is not done, the data collected using the factors you propose will be highly subjective and incomparable.

Secondly, the English writing needs improvement, I know the authors are not first language English speakers and I know how difficult it must be to write in a “foreign” language but the current manuscript is hard to understand at times and the authors need to improve it. I do highlight some key changes needed by there are many more minor ones.

Thirdly, I also feel that the new set of proposed standardized variables to assess eradication success or failure should be tested on more than just three examples. This is not enough to test how robust the system is. I believe that many more case study examples should be tested and compared in your study. I would even go as far as testing 30 or more and running some statistical analyses on the data. Maybe focusing on a region (artic islands, or Europe etc.) could work (e.g. see the study you cite by Pluess et al 2012). The best thing is if your list could be used to predict the success an eradication campaign might have.

Finally, I am still a bit sceptical about this framework and its lack of specificity. I think it needs more testing. I also think the authors should look at similar assessment or scoring tools like weed risk assents and impact scoring tools to guide and improve their work.

Minor comments

Line 25-27 and Figure 1: I think calling it “site context” instead of “location context” would be best. Similarly, “social” context instead of human context would be better. Again “replay” is not the correct word to use. How about “reinvasion”, I do see later you discuss why you call it “replay” but it’s a bit colloquial and I am not sure if this is the correct word to use. Maybe “reassessment” would work but without “context” afterwards.

Line 25-25: This is hard to understand.

Line 28: I am not sure what is meant here.

Line 39-43: I would remove the quote, I don’t think it is really relevant for this paper.

Line 45. Delete

Line 46: “Biological invasions” is not a field, it is a process or phenomenon. Invasion science or invasion biology is the field. This needs to be changed.

Line 47: Reword this sentence. In rewording this sentence also change “prevention and combat” to “management”.

Line 58 -: I don’t think terminology is such an issue. It is more about identifying factors that contribute to or hinder the success of eradication. There are major terminology debates around what eradication means, but I am not sure if this fits in the scope of this paper. I think you should argue more that it is important to identify a standardized set of factors to analyze eradications, not to standardize the use of terms for these.

Line 67-73: This section is much better and focused on the study.  I would personally start your introduction here, and cut out most of what was written above. Small parts of it can be incorporated below.

 Line 77: I think a key paper that is missing is: Gardener, M.R., Atkinson, R. and Rentería, J.L., 2010. Eradications and people: lessons from the plant eradication program in Galapagos. Restoration Ecology, 18, pp.20-29.

Line 81: The name of your system does not make so much sense. I would rather call it “a system (or framework) for assessing eradication feasibility”.

Line 85-87: I have said this many times now, but to me, standardization of these terms are not important rather what is important is identifying and testing a full suite of factors that affect eradication campaign successes. I think the paper should be framed in this manner.
Line 89-90: I am not so sure what this means.

Line 91: “is now more …. to globalization” – delete this.

Lines 92-94: A little repetitive now.

Line 97: change “check” to “search”

Line 103: Please provide the citations for all the papers reviewed in a supplementary material file.
Methods: Please list all the exact details that were extracted from each of the reviewed papers. How did you go about collecting and compiling the data from the papers?

Line 149: I disagree with the authors here. The “number and distribution of propagules” relates to the size of the invasion – the associated term you put here relate to dispersal traits. They are not really comparable or the same in my mind. Maybe if it was termed number and dispersal it would be ok?

Line 176-179: I disagree, if you are doing it on a global level this needs to be done as precisely as possible otherwise you will be collecting different data based on people’s different subjectivities. Even with a high level of standardization personal understanding, subjectivities will still be an issue. I think you need to develop clear guidelines for this and things need to be as standardized as possible.  This can include ranges. Similar approaches like EICAT that analyze and compare impacts of various invasive species in different systems globally. Without making standardizing values this approach will end up comparing apples with oranges. Based on the literate you should be able to identify some ranges to categories values e.g. infestation size - < 1 m2  ; 1 – 10 m2 ; 10 – 50 m2 ; 50 – 100 m2 ; 100 – 500 m2 ; 500- 1km2 ; 1 – 5 km2 ; 5-10 km2 ; 10-50 km2  >50 km2 . I think setting broad scales that account for differences in social and ecological systems is needed. Land uses, could be protected,  natural, semi-natural, agricultural, urban, etc.

Line 196: For eradication, there needs to be an elimination of the total population (0 for at least a certain length of time) otherwise if it is just lowering it, that would be classified as containment or asset protection, not eradication.

Line 274: This might need to be broader than just climate? Soils, fire regimes etc.?

Line 288:Needs clarification.

Line 295: I would stick with “reinvasion”.

Line 301. Is it not more for scientists who would collect and analyze available information?

Line 304: I would not say this, you put doubt in the reader's mind about what your work contributes.

Line 305: Not without some form of standardizations for each factor.

Line 320-325: Wouldn’t this be best to identify using statistical approaches. You could assess the importance of factors within a given context using multiple regression? Again, this scoring system lacks guidelines and so maybe too subjective.

Lin 336: I think it needs clearer guidelines.

Line 530 onwards: As mentioned I think you should use a lot more cases to test this to see how robust and standardized and useful the proposed set of factors can be. Alternatively, it can be limited to fewer case studies but it might be useful to get multiple people to score it to see if there are similarities or differences in how it is applied based on different assessors. To check how robust the scoring is it might be useful to add a confidence score for each of the factors.

Figure 1:
 Point 6: “land use” and “ownership” should be separated into two different variables. They are not mutually exclusive.
Pint 8: What about adaptation to other things: should this be adaptations to the local environmental context, not just climate change.
I think there should be some predefined categories to choose from for each of the 24 factors.


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: UNIFIED SYSTEM DESCRIBING FACTORS RELATED TO THE ERADICATION OF AN ALIEN PLANT SPECIES
Review round: 1
Reviewer: 2

Output:
Basic reporting: This paper proposes a simple system describing factors related to the eradication of an alien species. This is an interesting paper. It is well written, well-structured and brings new information.
Experimental design: (a)	One of the problem is that some the factors are of categorical nature, while some other are continuous. In this ms, all factors are treated as categorical. Does this not reduce the power of the method?
(b)	The last context, “replay context”, which contains only one factor, “reinvasion possibility (factor 24)”, should not be produced mathematically by the previous ones?
(c)	The authors should make it clear that this method can be applied at a local scale. For example, this cannot be applied at a national scale, where the same factor (e.g. monitoring area size) may be labeled differently in one location (an island population) than another for the same species (a mainland population).
Validity of the findings: no comment
Additional comments: (d)	I think that the Table S1 should be part of the main text.

Minor comment
(e)	Survey methodology: authors have to give the time-period of publication of papers they searched for. In addition, we need a criterion for the final selection of 32 papers.


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

Input:
Title: A NEW SPATIOTEMPORAL TWO-STAGE STANDARDIZED WEIGHTED PROCEDURE FOR REGIONAL DROUGHT ANALYSIS
Review round: 1
Reviewer: 1

Output:
Basic reporting: The current effort with the title “A novel spatio-temporal two-stage standardized
weighting scheme for regional drought analysis” applies a composite indicator (SDI), which combines meteorological (SPI) and Agricultural (SPEI) aspects. The article is well structured and technically correct in general, with the overarching aim of showing that the Index can be used in areas with similar conditions. In addition, there is an issue with the methodology. It is underlined, as also pertinent research has recommended, to avoid time scales of 3-month or shorter for the indices estimation, since Pakistan exhibits characteristics with extremely low precipitation during summers. Thus, it is suggested to apply the time scales of 6 and 12 months to characterize more accurately droughts and the analysis results to be comparable across the continent.
Experimental design: - Please improve the introduction with the following terms composite indicators, drought indicators, and the statistical weighting methods (e.g. OECD, European Commission (ed) (2008) Handbook on constructing composite indicators: methodology and user guide. Organisation for Economic Co-operation and Development Publishing, Paris).
- It is proposed to use a hill shade background with transparency to improve the quality of the map. In addition, it will be helpful for the potential readers a base map of the whole area.
Validity of the findings: The quality of the figures and maps should be improved.
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: A NEW SPATIOTEMPORAL TWO-STAGE STANDARDIZED WEIGHTED PROCEDURE FOR REGIONAL DROUGHT ANALYSIS
Review round: 1
Reviewer: 2

Output:
Basic reporting: .
Experimental design: .
Validity of the findings: .
Additional comments: Manuscript No: #65175

Title: A novel spatio-temporal two-stage standardized weighting scheme for regional drought analysis.

1.       Pg. 8: Line 82-83: How has this index been validated. Plz explain in detail.

2.       Pg.10: Line 112: Change "Moreover, in a Markov process, it can be more explicitly defined as the probabilities are approached the steady-state probabilities .......

Change to:

       Moreover, in a Markov process, it can be more explicitly defined as the probabilities approach the steady-state probabilities.......

3.       Pg 13: Line 200-202: Multiple distributions have been used to the one-month SPI and one-month SPEI at different sites. How has it been ascertained that these scheme is statistically consistent ?

4.       Pg 13: Line 200-202: How has the weights obtained from STTSSWS been used to derive the various drought classes ?

5.       Pg. 14 Line 200-212: What could be the reason that one month-SPI and one-month SPEI has been used in this analysis ? Why the 3-month, 6-month and 12-month timescale SPI's not used in the analysis ?

6.       Has the indicators levels (weights) used for deciding the various classes of drought levels been fine-tuned with the actual field conditions during drought periods ?

7.       How sensitive are these weights for various drought levels ?

8.       The resolution of all the figures needs improvement.

9.       As compared to Figure 1, the study area depicted in Figure 7 & 8 seems to be elongated and is not in proportion (to scale). This may be rectified.


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

Input:
Title: A NEW SPATIOTEMPORAL TWO-STAGE STANDARDIZED WEIGHTED PROCEDURE FOR REGIONAL DROUGHT ANALYSIS
Review round: 1
Reviewer: 3

Output:
Basic reporting: The authors propose a method to ponderate the drought classes obtained from standardized drought indices.

Improving methods, proposing indices, and improving approaches to monitoring drought is important for its better management. However, the document has several methodological shortcomings that make it unsuitable for publication. The paper structure needs an improvement too.
Experimental design: -The methodology is not clear. The approach makes use of two meteorological drought indices (SPI and SPEI), their methodology must be included, even in a general way, since the method to weight the type of classes is based on this type of indices.
-It is difficult for readers to keep track of how the weights are calculated.
-It is also necessary to describe the type of drought classes according to the type of index used.
-The methodology for calculating STTSSWS is not clear. I suggest describing the steps that are necessary to calculate it.
-What is the justification for weighting by drought classes?
-Why not directly use the values of the indices to calculate the weights?
-What is the difference between this method and the use of a threshold directly applied to the values of the hydrometeorological variable?
-In a practical case, what is the significance of the weights obtained by the proposed method?
-In general, the occurrence of the Extreme class is less than the Severe, which is, in turn, less than the Moderate one (Median as mentioned in the document), why then weight by classes?
-Why not monitor drought directly by type of drought (i.e. Extreme, Severe, Moderate)?
Validity of the findings: The Discussion section requires improvement.
Some points to discuss are:
-The advantages of the method (compared to other approaches)
-Disadvantages, see the previous section.
-Since this paper is more for drought monitoring, it would be interesting to provide the reader with a guide to interpret the results obtained.
-Difference / Similarity with respect to other approaches.
-Difference / Similarity with respect to other studies applied in the study area.
Additional comments: I'm not sure if this method is novel, the use of drought classes (Extreme, Severe, and Moderate) is widely used and discussed, and also the treatment of this type of information.

Introduction Section.
What about the methods that address drought as an object that changes in space and time?
Corzo Perez et al. (2011), Diaz et al. (2020a, 2020b), van Huijgevoort et al. (2013), Vernieuwe et al. (2020) and Zscheischler et al. (2013)

It is not clear the motivation.

Line 61 “drought monitoring”?
Lines 65-69 The sentences are not clear. SPEI uses also PET
Lines 71 What issues? Indicate some of them.
Line 73 what does mean complicated phenomenon?
Line 73 what does mean complex occurrence?
Line 74 now, drought monitoring policies? What this term means
Line 2.1 Where is the standardized drought index? This is an introduction of SPI and SPEI, but the methodology is not described.
Line 155 what does mean imperative knowledge?
Line 103-157 STTSSWS methodology is not clear. It’s difficult to follow the logic of the procedure.
Line 169 this was already shown in the Introduction sect.
Line 171-772 this is not the place for this sentence.
In Application (I assume is the Experimental setup), the experiment has to be described, what specifications other colleagues have to follow in order to reproduce your experiment.
Line 184 SDI? How this drought index was calculated?
Line 189-194 Where this procedure was described? Where Weibull, 4p Beta Johnson SB distribution were described?
Line 235 BIC criteria?
Line 241-242 How “the class” reflects for consideration in the region?
Line 242-246 this is not the place for this text. This is not a discussion.
Line 246 It is not clear how the introduced index or method provides more appropriate spatiotemporal information. What do you mean by “appropriate spatiotemporal information”?
Line 253-257 this text is not part of your conclusions, this part is for the introduction.
Line 258-259 See comments on Line 242-246

References
Corzo Perez, G.A., van Huijgevoort, M.H.J., Voß, F., and van Lanen, H.A.J. (2011). On the spatio-temporal analysis of hydrological droughts from global hydrological models. Hydrology and Earth System Sciences, 15(9), 2963–2978. https://doi.org/10.5194/hess-15-2963-2011

Diaz, V., Corzo Perez, G.A., Van Lanen, H.A.J., Solomatine, D., and Varouchakis, E.A. (2020a). An approach to characterise spatio-temporal drought dynamics. Advances in Water Resources, 137(January), 103512. https://doi.org/10.1016/j.advwatres.2020.103512

Diaz, V., Corzo Perez, G.A., Van Lanen, H.A.J., Solomatine, D., and Varouchakis, E.A. (2020b). Characterisation of the dynamics of past droughts. Science of The Total Environment, 718, 134588. https://doi.org/10.1016/j.scitotenv.2019.134588

van Huijgevoort, M.H.J., Hazenberg, P., van Lanen, H.A.J., Teuling, A.J., Clark, D.B., Folwell, S., Gosling, S.N., Hanasaki, N., Heinke, J., Koirala, S., Stacke, T., Voss, F., Sheffield, J., and Uijlenhoet, R. (2013). Global Multimodel Analysis of Drought in Runoff for the Second Half of the Twentieth Century. Journal of Hydrometeorology, 14(5), 1535–1552. https://doi.org/10.1175/JHM-D-12-0186.1

Vernieuwe, H., De Baets, B., and Verhoest, N.E.C. (2020). A mathematical morphology approach for a qualitative exploration of drought events in space and time. International Journal of Climatology, 40(1), 530–543. https://doi.org/10.1002/joc.6226

Zscheischler, J., Mahecha, M.D., Harmeling, S., and Reichstein, M. (2013). Detection and attribution of large spatiotemporal extreme events in Earth observation data. Ecological Informatics, 15, 66–73. https://doi.org/10.1016/j.ecoinf.2013.03.004


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: FUNGAL COMMUNITIES REPRESENT THE MAJORITY OF ROOT-SPECIFIC TRANSCRIPTS IN THE TRANSCRIPTOMES OF AGAVE PLANTS GROWN IN SEMIARID REGIONS
Review round: 1
Reviewer: 1

Output:
Basic reporting: The article is mostly well written. However, some sentences are too long and should be shortened for clarity. Some examples are:
Lines 118-121, 184-187, 197-199, etc.

Some references related to the fungal part of the microbiome associated with Agave species and their effects are missing, such as:
doi: 10.1111/1462-2920.15395.
https://doi.org/10.1016/B978-0-08-102268-9.00012-4
10.1111/j.1469-8137.1992.tb00092.x
https://doi.org/10.1023/A:1024046925472
http://www.scielo.org.mx/scielo.php?script=sci_abstract&pid=S0187-71512009000400005&lng=es&nrm=iso&tlng=es
http://www.scielo.cl/scielo.php?script=sci_abstract&pid=S0718-95162016000400016&lng=es&nrm=iso&tlng=en
http://www.ajol.info/index.php/ajb/article/view/95729

Raw transcriptomic reads have been shared, as two papers based on the RNA-Seq analyses have been published already (plant transcripts and viruses). I identified however, that only 17 libraries are associated with this accession number, although I expected 27 or 18 (3 Agave cultivars, 3 plant compartments (roots, stem and leaves) and 3/2 biological replicas).

Authors should include quantitative data about the size of the Agave plants sampled, the properties of the soils (if possible), and also about the precipitation patterns. They argue these plants have suffered irregular precipitation periods, but no evidence of this is provided.
Experimental design: The research question was well defined. However, the methods are not fully and clearly described to allow replication.

For instance, how total RNA was obtained from collected plant samples is not fully described. Did authors used a kit to deplete ribosomal RNAs? Did they use PolyA tails to recover only eukaryotic mRNA sequences?  If total RNA was sequenced, can authors also provide a general view of the prokaryotic mRNAs in the leaves, roots and stems of the three Agave cultivars investigated too? how many.. I realized that authors have already published the analyses on the plant transcripts and also on the viruses! That is great! However, this paper on fungal transcripts is insufficiently documented to be clearly followed.

To my knowledge and expertise, ribosomal RNA depletion efficiency is always below 100% and thus, rRNAs are normally sequenced and can be used for taxonomic identification. Can authors recover and report fungal rRNAs sequences? That would link their functional annotations to more specific lineages. This information would also allow the mapping of the fungal transcripts to "specific" fungal genomes recovered from these agaves.

One of the major aspects missing is the analysis of the distribution and identity of fungal transcripts in the three plant niches or compartments (root, stem and leaves) for each Agave species. Why only root associated fungal transcripts are described and discussed? I strongly suggest authors to give an overview of the differences between plant niches and cultivars.

The methods used for the annotation of  the fungal transcripts are not sufficiently documented.

Ideally, authors should make the in-house transcripts publicly available in a platform such as github.

I could not find clear references to the fungal genomes used in this study outside "Ascomycota" and "Basidiomycota" (nor in the main Manuscript nor in the Supplemental Material). Did authors fully neglected AMF or other Mucoromycota? to date there are at least 10 genome assemblies of AMF in public databases, and they are important fungal symbionts of plants, including agaves.
Validity of the findings: One of the major points that need revision is the lack of analyses of the different plant compartments sampled, that is the roots, stem and leaves of the 3 Agave cultivars. Authors only discussed and presented detailed data about fungal root transcripts, but an overview of the fungal transcripts in the other plant niches is missing.

The findings mostly describe the expression of certain fungal genes in the plant samples, but these expression can not be directly linked to mechanisms or causality of drought resistance and/or pathogen defense. So, authors should revise the discussion of their results to reflect this limitation. Further experiments or analyses would be needed for that.

In general, authors could compare in more detail their results to the fungal phylogenetic profiles generated already for other Agave species (line 393-396). Indeed from these reports, no differences in fungal diversity associated with the roots of cultivated and wild Agave species was observed. Contrary to the claim authors make in line 396-400.
Additional comments: No additional comments