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: BIOLOGICAL AGE FOR CHRONIC KIDNEY DISEASE PATIENTS USING INDEX MODEL
Review round: 2
Reviewer: 1

Output:
Basic reporting: The authors have taken into account the reviewers’ comments in the revision of the manuscript.
Therefore, I recommend its acceptance for publication.
Nonetheless, the English writing should be carefully revised throughout all the text.
Please also confirm  the numbering of the bibliographic references, namely those in Table 1. As it stands, there are no citations to references 24 to 28.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: Lines 111-112 - In the pdf file it is shown "Error! Reference source not found.".
Equation (9) and line 142 - Are the weights equal for all individuals? If so, they do not need the index x.
Line 156 - "BioAge" should be replaced by "BioAge for x" in the index of I
Line 199 - "biomarkers feature almost similar" should be replaced by "biomarkers feature have almost similar".
Line 238 - "to follow the normal distribution Z ~ N(0,1)." should be replaced by "to follow a normal distribution.". The difference between CA and BA do not have zero mean neither unitary standard deviation as stated: Z ~ N(0,1).
Line 278 - "enables a just comparison" should be replaced by "enables a fair comparison".


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: BIOLOGICAL AGE FOR CHRONIC KIDNEY DISEASE PATIENTS USING INDEX MODEL
Review round: 2
Reviewer: 2

Output:
Basic reporting: N/A
Experimental design: N/A
Validity of the findings: N/A
Additional comments: N/A


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: FIGHTING ABILITY, PERSONALITY AND MELANIN SIGNALLING IN FREE-LIVING EURASIAN TREE SPARROWS (PASSER MONTANUS)
Review round: 1
Reviewer: 1

Output:
Basic reporting: In this well written article, the authors find a link between fighting behaviour and physical traits, as well as a potential "honest signal" function in females, but not males.
I appreciate the thought and effort that went into this experiment to solve a question that to a layman seems easy to solve, but which, after reading the materials and methods is not straightforward.

Most of the relevant literature is present, although an extra ref here and there could be useful (see below). While most remarks are small, I have three remarks that I personally find important.

1)This is probably the shortest results-section I have ever seen. Maybe reconsidered the structure and call it Results and Discussion. An alternative would be to provide some additional summary statistics on exploration (e.g. on average individuals would visit x compartiments) etc.

2) Line 379: is this a likely explanation? How would food shortage promote smaller males being more successful? Have you considered population differences? Age-structural differences (i.e are smaller males young and fit males while bigger birds are old, fat non-fit birds)?

3)Line 408-428: As this is your main results I feel it might be a bit more expanded. I feel that there is a lot of focus on group characteristics influencing the results. However, parasite load, immunity and stress levels could easily differ between populations, explaining the observed results.

Other minor remarks:

Line 61: While it is clear how personality is linked to fighting ability, it is unclear what the “id est” means. Also the “see examples below” can be removed as the examples are listed immediately after.
Detail: the text contains a lot of “like”, to make the text a bit more fluent the author could replace some of the likes with synonyms.
Line 65: I think you can change “were” to “are”.
Line 77: expand a bit on the ultimate perspective. Are these processes really similar, and if so is it then correct to state that they “covary due to multiple reasons” as you mention only one. Even though this has to be clarified a bit, I do appreciate lines 67-77 which are nicely explained.
Line 88/Line 92-95: I would try to find another bird example (in addition to great tits), rather than distantly related species. If such examples do not exist (this is not my expertise), then mention that it is actually quite rare, and rephrase this section that while numerous throughout the animal kingdom, it is rare in birds.
Line 120: this section missed some of the key publications, including McGraw (2008) (“An update on the honesty of melanin-based color signals in birds”). Also, I do not agree with the term “ubiquitous”. Unlike carotenoids (for which ubiquitous could be correct), support for melanin as an honest signal is often uncertain or more complex (see e.g. D’Alba et al. Melanin-based color of plumage: role of condition and feathers’ microstructure). I would simply shift the focus that one potential signal associated with status is melanin-based colouration. Maybe also drop the conspicuous, since melanin is often considered inconspicuous compared to other colours present. Finally, this section is rather short compared to other sections – but this might be just a personal preference.
Figure 1-3, mention R² values in caption.
Line 337: is this not contrasting with Monus et al. 2017 you mention in the introduction?
Line 356: briefly explain what fission-fusion type of group dynamics is.
Line 365: is it  possible to measure this if not you can mention this.
Mention in the data accesibility that some data has already been published.
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: FIGHTING ABILITY, PERSONALITY AND MELANIN SIGNALLING IN FREE-LIVING EURASIAN TREE SPARROWS (PASSER MONTANUS)
Review round: 1
Reviewer: 2

Output:
Basic reporting: This manuscript reports the results from an experimental field study that aims to understand whether exploratory behaviour, an aspect of personality, and some other individual features (sex, bib size, etc), explain fighting ability when competing for food during winter in free-living Eurasian tree sparrows. The authors should be commended on a well-conceived and well-designed experiment that, in my opinion, is well balanced and with (at least) some replicates. In addition, I have found the manuscript very well constructed and easy to follow and understand. In particular, the authors have clearly introduced the importance of the study - which I find very interesting indeed - and discussed their results in a broad and general context. I have raised some major and some specific issues below that I hope would improve the manuscript though.
My major comment is about statistics. I think the response variable is a proportion (i.e. number of times an individual win related to the number of encounters). Indeed, this is what the Authors stated in the Abstract (Line 35). However, they analyzed this variable as a binomially distributed one (i.e. the number of fights won vs the number of fights lost) and built a generalized linear model. The number of fights won is in this case related to the number of fights lost because when an individual win a fight, it cannot lose it. In my opinion, it would be more appropriate to estimate the proportion of e.g. fights won. Actually, that was what I thought the Authors did when reading the manuscript. But I got confused by the model they built. This variable should be treated as a beta (that is, a proportion limited between 0 and 1 values) instead of as a binomially distributed one. In addition, some individuals took part in some encounters, but the number of times that the different individuals participated in different fights is different. I think they should include individual as a random factor to account for this unbalanced structure of the data. In this line, the experiment was done in two different years. To account for the potential confounding effect of the year, as the Authors indeed noticed in the Discussion section, year should be also included as a random factor. Finally, I am wondering whether the residuals of their model are temporally autocorrelated, as the experiment was conducted in consecutive (?) days. I would expect the behaviour of the individuals to be more similar on consecutive days than on days that are far away. So my suggestions are:
(1)	Build a generalized mixed (see point 2 below) model, but where the residuals of the response variable are beta instead of binomially distributed
(2)	individual and year are included as a random factor
(3)	check and, if necessary, include a factor to account for the fact that behaviours are temporally autocorrelated
Some specific comments:
Line 77: This is very interesting. Could the Authors expand their explanations about the possible negative frequency-dependent selection?
Line 82: unclear sentence
Line 166: Could the Authors provide the website of the provider?
Line 194: is 10 minutes enough? Has been this time selected based on some previous studies?
Line 375: 15 years earlier but the study was published in 2017. Is this correct, right? Just checking…
Line 378-385: I found it vague to argue that the difference could be only attributed to differences in wintering conditions. There might be many other factors, including experimental design, individual condition, etc
-	Could the Authors provide a fit-of-goodness (r-square or deviance explained) of the selected model?
Experimental design: As stated above, I think the experiment is well-conducted. But I need to honestly say here that I am not usually designing and performing experiments. My research is mainly based on observational studies.
Validity of the findings: The authors have clearly introduced the importance of the study - which I find very interesting indeed - and discussed their results in a broad and general context.
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: FIGHTING ABILITY, PERSONALITY AND MELANIN SIGNALLING IN FREE-LIVING EURASIAN TREE SPARROWS (PASSER MONTANUS)
Review round: 1
Reviewer: 3

Output:
Basic reporting: .
Experimental design: .
Validity of the findings: .
Additional comments: Attached my comments on the PDF.


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: FIGHTING ABILITY, PERSONALITY AND MELANIN SIGNALLING IN FREE-LIVING EURASIAN TREE SPARROWS (PASSER MONTANUS)
Review round: 2
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: no comment


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

Input:
Title: FIGHTING ABILITY, PERSONALITY AND MELANIN SIGNALLING IN FREE-LIVING EURASIAN TREE SPARROWS (PASSER MONTANUS)
Review round: 2
Reviewer: 2

Output:
Basic reporting: Dear Authors,

I congratulate you on the wonderful revision you made to your manuscript. I really thank you for following my advices on the statistical methods. I think the manuscript has greatly improved its quality. I hope you agree with me. I do not have further comments. I am really impressed by the work you have done on this revision.

Best,
maria
Experimental design: no comment
Validity of the findings: no comment
Additional comments: no comment


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

Input:
Title: FIGHTING ABILITY, PERSONALITY AND MELANIN SIGNALLING IN FREE-LIVING EURASIAN TREE SPARROWS (PASSER MONTANUS)
Review round: 2
Reviewer: 3

Output:
Basic reporting: Introduction and Discussion overlong compared to content
Experimental design: Reply to previous concerns not satisfactory
Validity of the findings: Reply to previous concerns not satisfactory
Additional comments: The reply to my previous concerns was not entirely satisfactory. Although the authors have somewhat improved the presentation of their results, there is ample room for improvement. I have provided more details in the attached annotated PDF file. I think there are several issues with the analyses that still needs to be addressed, with special reference to contextual effects of foraging group size and temperature. Also, issues of collinearity between sex, body size and bib size were not properly addressed and solved, in my opinion (at least not explicitly).


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: BLOOD GLUCOSE, INSULIN AND GLYCOGEN PROFILES IN SPRAGUE-DAWLEY RATS CO-INFECTED WITH PLASMODIUM BERGHEI ANKA AND TRICHINELLA ZIMBABWENSIS
Review round: 1
Reviewer: 1

Output:
Basic reporting: 1. The problem statement was not stated clearly, hence not explicit in the intro section.
2. Some literature references were not properly cited across the text, some cited references were missing in the ref. list, while often, most of the cited references were too old. It is understandable that there are not too many literature on Tz, but other spp of Trichinella or tissue-dwelling nematodes could be cited.
Experimental design: Some vital information about inoculation of the parasites and animal handling & sacrifice were not provided. The author should provide detailed info about these.
Validity of the findings: 1. Any evidence of permission by the journal or publisher of the Fig. 1&2 to be for publication in PeerJ?
2. The use of 2-way ANOVA for table 1. was not appropriate. Also, posthoc test should have been adopted to do the ranking for differences in means.
Additional comments: 1. The Title is too long. Suggestion has been provided in the text
2. Footnote should be provided for the table
3. There should be consistency in the references list.


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: BLOOD GLUCOSE, INSULIN AND GLYCOGEN PROFILES IN SPRAGUE-DAWLEY RATS CO-INFECTED WITH PLASMODIUM BERGHEI ANKA AND TRICHINELLA ZIMBABWENSIS
Review round: 1
Reviewer: 2

Output:
Basic reporting: The article is well written and meets the standard presentation of this journal.
Authors introduced their subject with a clear research question referring to relevant literature to give the reader context. Raw data was supplied as supplementary files and due process was taken to give recognition to authors whose findings are available already in literature to complete the methodology used in this paper. Unfortunately, the reference of this paper, Murambiwa et. al. 2020 is missing in the bibliography.
The figures are clear and of good quality, each presenting important aspects of their findings. They shared their raw data for verification of results and points of discussion.
Experimental design: Research question was clearly stated and the gap in knowledge argued from literature. The design is appropriate to answer the research question.
The experimental design is straightforward and sound, to discriminate between treatment (co-infection) and non-treatment (Pb mono-infection) effects. The 42-day trial period allowed only 14 days period of co-infection, since Tz took 28 days to establish itself in muscle tissue. As much as the authors clearly state in the supplementary files and methods that day 0 is day 28 (when Pb co-infection took place, giving them 14 days to D42) I suggest in the annotated comments on the pdf, to always refer to the groups as mono-infection (Pb only) group and co-infection (Pb + Tz) group, the Tz only group poses no confusion.

A rigorous investigation was performed and high ethical and technical standards were upheld.
Validity of the findings: Their findings are corroborated by the results of this investigation and they took their liberty to use results from literature, which is actually research done in their laboratory. The effects of Pb and Tz co-infection on insulin levels, glycogen concentration and glucose concentration will still need further investigation, but these conclusions will form part of the basis or reference point for comparison. The experiments were appropriately controlled.
Additional comments: The paper is well written and the English is easy to understand. I suggested a few grammatical corrections on the pdf. If they can better distinguish the mono-infected and co-infected groups as stated under experimental design comments and on the pdf anotations, then the confusing element will be obviated. It was interesting to engage with the findings of this research.


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: BLOOD GLUCOSE, INSULIN AND GLYCOGEN PROFILES IN SPRAGUE-DAWLEY RATS CO-INFECTED WITH PLASMODIUM BERGHEI ANKA AND TRICHINELLA ZIMBABWENSIS
Review round: 2
Reviewer: 1

Output:
Basic reporting: Satisfactory
Experimental design: Comprehensive and adequate
Validity of the findings: All data were provided and valid
Additional comments: Overall, the queries have been answered.


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: LNCRNA/MIRNA/MRNA CERNA NETWORK ANALYSIS IN SPINAL CORD INJURY RAT WITH PHYSICAL EXERCISE THERAPY
Review round: 1
Reviewer: 1

Output:
Basic reporting: It is a  professional English language used throughout.
The background  shows context, it is relevant for the research, but not so more informations about the physical exercise in the context.
The structure conforms to Peerj standards.
Figures are well labelled.
Experimental design: It is an original research and a novelty in field of analysis the role of physical exercises.
It is not clear how is the relationship between mRNA and spinal cord injury. Whys this approach?Please explain.
What means rats functional status?Please explain
The research is rigorous and respect  high technical & ethical standard
Methods described  have sufficient detail but are some question about line 119-120, `` If  multiple arguments are vectors a 120 matrix or array of results is returned. values for alpha and power and…..`seems to lack some informations.
It is not clear the rats groups, it is not a complet description, seems it is a group which has spine section and a group which participate to the training? Or both participate?
Line 130- you speak about 2 groups…which?
Any other inflammatory markers, used for quantifiy the anti-inflamator effects of physical exercise?
Validity of the findings: Anti-inflamatory effects seems to develop after few month of physical exercises. How did you proceed regarding this?
Conclusions are well stated, linked to original research question
Additional comments: 4.General comments
The papers is very interesting and well argumentated but a little bit hard to follow and for this reason I suggest to reorganize the results and present more clear the role of physical exercises.


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: LNCRNA/MIRNA/MRNA CERNA NETWORK ANALYSIS IN SPINAL CORD INJURY RAT WITH PHYSICAL EXERCISE THERAPY
Review round: 1
Reviewer: 2

Output:
Basic reporting: In this study, the author performed RNA-seq and microRNA-seq to examine the differentially expressed circRNAs, lncRNAs, miRNAs, and genes between SCI and TMT groups. The author also established the ceRNAs network using interactions between miRNAs and mRNAs, as well as between miRNAs and lncRNAs/circRNAs. These findings may support that exercise promotes efficient signaling and neuronal plasticity in the spinal cord. Overall, the study is meaningful. I have some minor comments as follows:
The English writing needs to be improved to ensure clear understanding. In the methods part, the writing of the manuscript is not standardized. I suggest the author improve this part.
1) Line 120: repeated punctuation
2) Line 122: “value of .5” abnormal punctuation
3) Line124: “samples sizes” The first word of a sentence should be capitalized.
4) Line 127: “value though https” Here should be “through”.
5) Line 115: “CV” Uncommon abbreviations should be spelled out the first time.
6) Line 85 and Line 116: Please use one space following periods.
7) Line 112: “being compared. within groups” abnormal punctuation here
8) Line 130 “TMTgroup” no space between two words
9) Line 88 and Line 99: “Bladder care”, “all Rats” The capital letter is not necessary here.
10) Line 218: “mode” Here should be “model”.
11) Line 57 and Line 62: The font of in-text citations here is different from other parts.
Experimental design: 1) How does the author establish the ceRNAs network？ Although the author used the software or database, the interaction is based on what kind of condition? Expression level or correlation coefficient? LncRNAs, miRNAs, and mRNAs, are they positively correlated or negatively correlated?
2) Line 210: “The means ± standard deviations (mean ± SD) were used to express all of the data.” However, in the Figure 1 legend, “Data are expressed as mean ± SEM.” It is not consistent.
3) Figure 2 (B-D): The size of each group name is too small. It is not clear.
4) Line 34: “circRNAs (DECs), lncRNAs (DELs), miRNAs (DEMs), and genes (DEGs) between SCI and normal controls” Here the “normal controls” means “Sham group” or “TMT group”? But in Figure 1 (B-D), the heatmaps showed the gene patterns of SCI groups and TMT groups. Please explain it clearly.
Validity of the findings: no comment
Additional comments: no comment


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

Input:
Title: LNCRNA/MIRNA/MRNA CERNA NETWORK ANALYSIS IN SPINAL CORD INJURY RAT WITH PHYSICAL EXERCISE THERAPY
Review round: 1
Reviewer: 3

Output:
Basic reporting: The manuscript – LncRNA-miRNA-mRNA ceRNA network analysis in Spinal cord injury rat  with Physical exercise therapy investigated the effects of exercise therapy on the long noncoding RNA expression. Overall, the background and methods they used were clear and straightforward. Certain RT-PCR of potential inhibitory effect of exercise therapy on inflammatory response are quite significant. They provide a resource of differntitial expressed circRNA , lncRNA miRNA and differential expressed genes between spinal cord injury and normal rates.
Experimental design: The Experimental design and bioinformatic analysis were well defined. They proposed a question how the effect of exercise therapy influence the expression changes on long noncoding RNA, microRNA and mRNA in rate with spinal cord injure. They established a spinal cord injury model and traced the effects. Further lncRNA-seq miRNA-seq and mRNA-seq were perfomed to detected the molecular changes. GO analysis, KEGG and ceRNA network were further analyzed. Addtionally, they performed the qRT-PCR to validate the suppression of the post-SCI inflammatory response to physical exercise therapy.
Validity of the findings: They used the qRT-PCR validation to validate the inhibitory effect of exercise therapy on inflammatory response after SCI.
Additional comments: Minor comments

Figure 1A the figure seemed incomplete

Figure 2: Please describe in detail how your compared the DEG, DELs and DEMs?

Figure 3B The x axis is not clear

Table 4: I am wondering why you have so few DEGs, and why using the Pvalue in the table not FDR described in your method?


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: LNCRNA/MIRNA/MRNA CERNA NETWORK ANALYSIS IN SPINAL CORD INJURY RAT WITH PHYSICAL EXERCISE THERAPY
Review round: 2
Reviewer: 1

Output:
Basic reporting: No comments
Experimental design: No comments
Validity of the findings: No comments
Additional comments: All remaining issues were adequately addressed.
There are two minor points about the format of the manuscript.
1. Figure 6. There were no red and green colors here. The colors that I saw were blue and brown.
2. Abnormal space in Line 382 and 383.


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: ALGAE FROM AIPTASIA EGESTA ARE ROBUST REPRESENTATIONS OF SYMBIODINIACEAE IN THE FREE-LIVING STATE
Review round: 1
Reviewer: 1

Output:
Basic reporting: To be honest, the purpose of this study was not clearly understood. Based on the assumption that the origin of symbiotic algae acquired by animals is not a free-living population in the environment, but that they are expelled from other animals, the authors analyzed physiology and gene expression of the symbiotic algae in egesta of Aiptasia in detail, and based on the results, they seemed to propose using the symbiotic algae in the egesta as a standard host-symbiosis study. What would be that study? Given that Aiptasia acquires only a specific Symbiodiniaceae (Breviolum), it may be suggested that the results of this study be used as an experimental standard in the acquisition of symbiotic algae by Aiptasia, but this is not clearly stated. In view of various reports that symbiotic algae become a flagellated swimming stage in a free-living state, and that this swimming ability is necessary for attraction to corals, the results of this study, which did not observe whether they change to a swimming stage, do not seem to be applicable to coral research. So, if the authors are going to propose this as a model for how animals acquire symbiotic algae, they need to limit it to Aiptasia first.
    I, too, do not believe that symbiotic algae live solitary in the oligotrophic reef waters, and, like the authors, I believe that expelled symbionts from other animals are the symbiont source for the next animals. However, some researchers, for example Littleman et al. (2008), say that “Free-living Symbiodiniaceae” is a symbiotic source, and we don't know the truth yet, so I think Line 62 is saying too much.
   So, my suggestion is to rewrite the introduction as to focus on Aiptasia-symbionts only, not to broaden the story as to cover general of animals including corals, and to incorporate various aspects for the origin of symbiont sources in coral reef environment. Also, sentences describing inappropriateness of using culture strains (line 84) are unclear. There are numbers of studies using culture strains isolated from certain hosts for symbiosis establishment test on the same host (e.g. Umeki et al 2020). If the description was for Aiptasia study only, please mention so.
Experimental design: Line 126. It seems the tested animal was initially symbiosed with a culture strain of Breviolum. Isn’t this conflicting with the argument from Line 80 saying culture strain is not representative of what found in actual hosts?

Line 129 Was artificial seawater used to prepare culture medium F/2? If so, there are many reports arguing most microalgae are not maintainable in an artificial seawater-based medium, means that physiological results of this study were under abnormally unfavorable condition for the culture strain. It might be just a filtered seawater (FSW), wasn’t it?

Line 193 “Dark-adapted photosynthetic efficiency” should be “maximum quantum yield of the photosystem II”

Line 217. This seems to be common to many infection experiments, but is the cell density to be inoculated too high? It's a personal feeling, but if you give them so much symbiotic algae, they'll probably accidentally take it in, and do they really represent what's going on in the field?　This is fine this time, but I　recommend testing at a lower density next time to highlight egesta's contribution to symbiosis formation.
Validity of the findings: I did not understand the purpose of the description of gene expression in culture and in the host (from line 93). Are the authors trying to tell us the difference between the nutrient environment in egesta and a culture that is saturated with various nutrients? Or are they introducing AMT, NRT, NR, etc., to emphasize that the enzymes involved in nutrient uptake are not indicative, and that SWEET1 alone is a good indicator of the difference between conditions in the host and conditions away from the host? This fuzzy feeling is felt similarly in the text from Line 414 of the discussion. Genes for nutrient uptake remained the same in egesta and host and are much higher expressed than those in culture. This seems to indicate that the symbiotic state is not as present in the hypertrophic environment as in culture. If so, why the authors is “Challenging the hypothesis that the host is active limiting nutrient transport to the symbionts”? The implications of L 428 for high SWEET 1 expression are also unclear. Do they want to argue that this gene is not upregulated by host as suggested by Xiang et al 2020, and this is what they want to emphasize in this study?
Additional comments: Result
Fig. 1 No black arrows in D or E. Any specific reason for using 633 nm for chlorophyll excitation, not the blue one?

Line 308. Numbers of the expelled egesta were shown. However, we do not know the regular size of egesta, size variation, and average symbionts density in an egesta, which make us difficult to imagine whether the symbiont releases were large or small.

Line 315 “Photosynthetic health”. FV/Fm is just a quantum yield of the most PSII upstream and does not indicate the healthiness of overall photosynthesis.

Discussion
Line 394. This argument is highly agreeable, but probably limited to giant clams (see Umeki) and Aiptasia. In our experiments, the symbiotic algae released by corals died within 24 hours. In both Aiptasia and giant clams, the expelled symbiotic alga remain active (in egesta or feces) for a relatively long time, which may be advantageous to supply symbiotic source for others.


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

Input:
Title: ALGAE FROM AIPTASIA EGESTA ARE ROBUST REPRESENTATIONS OF SYMBIODINIACEAE IN THE FREE-LIVING STATE
Review round: 1
Reviewer: 2

Output:
Basic reporting: I enjoyed reading the submission by Maruyama and colleagues. Overall, this ms adheres to PeerJ professional and structural guidelines. My three primary concerns with regards to basic reporting are described below.

1.	The taxonomic nomenclature of the focal study species within the submission reflects outdated taxonomy. Replace “Aiptasia” with “Exaiptasia” throughout and cite Grajales & Rodriguez 2014 Zootaxa https://www.mapress.com/zootaxa/2014/f/z03826p100f.pdf .
2.	I often felt that the flow of the results and discussion were not tight enough and could be improved. Within these sections there were several brief (2-3 sentence) paragraphs (L313, L322, L394, L437). Flushing out the transitions in and out of these segments ought to help make the story more succinct.
3.	Information on the ex situ culture experiment need to be described in more detail (see comment on L263 of methods). Cell densities from the duration of the culture experiment do not appear to be reported in the supplement
Experimental design: The clever and creative experimental design are among the submission’s major strengths. The ms does an excellent job detailing the importance of the knowledge gap and difficulties associated with studying environmental Symbiodiniaceae populations. As such, using Symbiodiniaceae from anemone egesta as a proxy for studying the photophysiology, cell cycle, and gene expression of environmental conterparts is creative. I imagine this will inspire future research in the study system. I also wish to commend the authors on their successful incorporation of undergraduate research-based teaching and inclusion course members as co-authors.
Validity of the findings: The ms describes the proper controls and provides most necessary raw data. For the most part, the conclusions are justified by the results at hand (point by point comments below).
Additional comments: Thank you, Maruyama and colleagues, for your interesting and creative submission to PeerJ! My point by point comments are below.

Abstract
L31-32: omit from abstract to improve brevity
L33: define egesta

Introduction
L58-63: I think these statements over-speculate about the transient nature of environmental populations of Symbiodiniaceae. I also recommend citing Grupstra et al 2021 Animal microbiome which documents an abundance of viable Cladocopium and Durusdinium in the feces of corallivorous fishes. Some additional papers about environmental Symbiodiniaceae populations are also listen in my comments on the discussion.
L91-93: two sentences in a row are started with the phrase “using algae.” Consider revising
L96-97: Add the citations for the data papers evaluating nutrient availability, gene expression, and cell division rates
L121: finish off this paragraph with an additional sentence tying together how this paper demonstrates that expelled algae are an effective proxy for studying environmental populations of Symbiodiniaceae,

Methods
L138: how frequently were the anemones checked for egesta released?
L237: is there a reference for the Cyclophilin housekeeping gene?
L263: Major concern -- more detail is needed on the details of harvesting samples from the ex hospite culturing experiments. The ms needs to verify the growth stage of each flask to ensure that samples were generated during comparable physiological states (ideally exponential growth). Here are some examples of papers demonstrating the physiological & transcriptomic variability of microalgal in between growth stages: Droop 1975 “The nutrient status of algal cells in batch culture”; Mansour et al 2003 “The effect of growth phase on the lipid class, fatty acid and sterol composition in the marine dinoflagellate, Gymnodinium sp. in batch culture”; Yoon et al 2002 “Combined transcriptome and proteome analysis of Escherichia coli during high cell density culture.”

Results
L311: is this data included in the supplement?
Paragraph at L328: include results from output of statistical analyses.

Discussion
L366: cite the original data papers evaluating nutrient availability and cell division rates
L370: swap “cooperative” for “homologous”
L373: I think this statement also over-reaches with speculation. Consider revising or removing
L375-377: I agree! I also think a benefit of the Exaiptasia-Symbiodiniaceae system is its capacity to work with several genera (and therefore species) of Symbiodiniaceae. Consider suggesting comparative approaches along these lines.
Paragraph at L379: this could also be due to elevated rates of stony coral feeding at night?
L394: is the FSW sterile? Perhaps bacterial symbionts are missing
L397: I think this statement overstates the conclusions and as it challenges our understanding of Symbiodiniaceae population genetics. For example, if the same genotype of D. trenchii is dominant throughout the Caribbean, how was its rapid, invasive population spread possible given the stated hypothesis about the transient nature of environmental populations (i.e., Pettay et al 2015 PNAS)? Here are some papers reporting population dynamics of Symbiodiniaceae in the water column: Decelle et al 2020 (https://www.sciencedirect.com/science/article/pii/S0960982218312193) and Raina et al 2022 (extended data 1: https://www.nature.com/articles/s41586-022-04614-3)
Paragraph at L413: I am surprised that the utility of proteomes in phenotyping (a la Maryuma & Weis 2021) is not mentioned here.
L422: Nice! I agree!
L441-442: I think this statement articulates why this submission represents a considerable breakthrough in the field. Consider alluding to earlier on in the text.
L442: replace col2lected with collected
L450-452: would perturbing anemone oral disks with a glass Pasteur pipette induce egesta release (without adding confounding variables)?


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: ALGAE FROM AIPTASIA EGESTA ARE ROBUST REPRESENTATIONS OF SYMBIODINIACEAE IN THE FREE-LIVING STATE
Review round: 1
Reviewer: 3

Output:
Basic reporting: The manuscript “Algae from Aiptasia egesta are competent symbionts and more closely represent the free-living state of Symbiodiniaceae in nature than do algae in culture” by Maruyama et al. explores the physiology of aiptasia egesta and compares it to an algal culture. I commend the authors for doing this work. However, the overall manuscript needs significant revision and streamlining of the results and discussion. This study could be interesting for the aiptasia community, but its current form should be revised.

More broad concerns/critiques
1. I felt the manuscript could have been written as a description of the biology of egesta. The authors should have streamlined the claims and only discussed what their data tells us.

2. The overall justification and rationale of the study are somehow weak, and there are points in the discussion that don’t align with it.
Experimental design: Methods:
L207: Please mention and specify the light intensity used.

Did the authors use negative control (seawater only) for the inoculation experiment? This should be included in the method section so we can tell that the four-day exposure to L:D was not the reason we are seeing algal signals in aposymbiotic aiptasia.

Primer design. The authors used Cyclophilin as a housekeeping gene. This should be justified. For example, show expression that the gene does not change in expression over time or cite a study that used the same gene.
Validity of the findings: Results

The authors reported no significant difference between egesta and in hospite gene expression using seven genes. How can we tell that the signal was not confounded by non-intact/dead cells incorporated in the extracted egesta?
A supplemental figure showing the ratio of intact and non-intact cells could be added to the supplementary materials.
Additional comments: Discussion

In Line 340-341, the authors mentioned about a variety of microbes. However, the current methods employed in the study to support this is not sufficient. More specifically, microscopy resolution compared to 16S sequencing is very different.

In Line 364-365, the authors mentioned that a higher mitotic index of egesta than in hospite could be due to nutrient availability. This statement doesn’t align with the justification above that egesta can be a representation of algal symbiont lifestyle than that of the cultured algae.

In lines 394-397, the authors mentioned that egesta is short-lived, which could be a limitation if we suggest its use for comparative study instead of algal culture. It’s also challenging to employ this in coral research when we have to wait at nighttime to have enough expelled symbionts samples.

Line 402-404 is speculative, and this statement should be removed unless the authors can cite a study. The results from this study don’t support this.

The whole section of 412-413 used select genes, and the statement that “egesta to represent the ex hospite state can remove nutrition as confounding variable” could not strongly support this claim.


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: ALGAE FROM AIPTASIA EGESTA ARE ROBUST REPRESENTATIONS OF SYMBIODINIACEAE IN THE FREE-LIVING STATE
Review round: 2
Reviewer: 1

Output:
Basic reporting: I still wonder whether the symbiont of egesta signifies one of their life stages, i.e., a free-living state. It seems to me that the symbionts in the egesta were simply dragging the environment within the host. In addition, it is difficult for me to imagine that the cellular environment in the host cells is oligotrophic even if the host animals live in the oligotrophic seawater, and therefore, the logical development that the nutrient-rich culture environment does not represent the actual situation is uncertain. However, it is valuable in the sense that the symbionts kept photosynthetic activity for long time of 48 hours even if they left from the host, and in addition, the current observation gives information to the hot discussion on what Symbiodiniaceae as a symbiotic source of the coral derives from.
Experimental design: The authors provide new information on the matters pointed out previously, and the result of live or death discrimination by Evans Blue is also interesting. The low rate of dead cells was surprising.
Validity of the findings: The observation gives another important information on how animals including corals acquire symbiotic algae and what they are derived from, and is worthy of publication.
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: ALGAE FROM AIPTASIA EGESTA ARE ROBUST REPRESENTATIONS OF SYMBIODINIACEAE IN THE FREE-LIVING STATE
Review round: 2
Reviewer: 2

Output:
Basic reporting: see additional comments
Experimental design: see additional comments
Validity of the findings: see additional comments
Additional comments: I thank the authors for their efforts restructuring the ms. I believe it is an exciting contribution to the fields of symbiosis biology and coral reef ecology. My remaining comments are minor and primarily responding to the rebuttal.

1.	Photosynthetic health metrics. I agree with reviewer 1, Fv/Fm indicates PSII photochemical efficiency, which is important for phenotyping, but is not suitable as a sole indicator of photosynthetic health. I encourage the ms to reconsider revising their use of the phrase “photosynthetic health” to more accurately depict the contents of their data and embrace the nuances associated with photobiology (i.e., something along the lines of ‘algal photophysiology’). The utility of Fv/Fm and chla fluorescence are further described  in Suggett et al 2009 MEPS https://doi.org/10.3354/meps07830 and Kalaji et al 2017 Photosynthetic Research https://doi.org/10.1007/s11120-016-0318-y
2.	Anemone taxonomy. I respectfully disagree with the rebuttal on Exaiptasia taxonomy. Many papers include both the updated taxonomy and informal name (see Dungan et al 2020 Symbiosis; Strumpen et al 2022 symbiosis; Siddiqui et al 2015 Aquatic Toxicology). I see this as a critical component to basic reporting, promoting reproducible research, and avoiding confusion for new students/researchers. I appreciate that the formal binomial was added to the abstract and introduction and request that it be added to the first mention in the methods (L125)
3.	Algal culturing methods. Typically, cell growth calculations are displayed as maximum growth rate (mu), but here are reported as cell yield. The ms now reports the age at which cultures were harvested (L370). The strain of B. minutum could be in either exponential or stationary phase by day 21 – this distinction would greatly impacts its physiology + nutritional status and the broader conclusions of the ms. I recognized it is likely a) not feasible to run additional experiments and b) additional experiments would induce batch effects in qpcr data; but request that this caveat is incorporated in the discussion sections at L422 & L442 (see references from first round of review).
4. Reviewer 1 expressed concern with regards to usage of artificial seawater (ASW) in algal cultural media and potential for unintentionally inflicting stress on the microalgae. I agree with the author’s response about ASW as an effective culturing option. The nutrient concentrations in common algal culture recipes are replete (see Krueger 2020 https://www.researchgate.net/publication/339788469_Media_recipes_commonly_used_for_Symbiodiniaceae_cultivation). As such, ASW culturing is unlikely to induce treatment artifacts.

L359: change “sebae” to “sea”


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: BASELINE SCENARIOS OF HEAT-RELATED AMBULANCE TRANSPORTATIONS UNDER CLIMATE CHANGE IN TOKYO, JAPAN
Review round: 1
Reviewer: 1

Output:
Basic reporting: Overall interesting results are presented that demonstrate extreme heat impacts on hospital transportation and how this is projected to change.
I would move the section about Tokyo starting from line 274 into the introduction as this is the region this study focuses on.
Line 49 - this figure quoted is for 3 heatwaves over the summer of 2019.

Line 88- the WBGT is not known for being easy to calculate (i.e. see Kong and Huber, 2021)

The conclusion could be restructured to better highlight the importance of these results and the wider importance expanded in the discussion.
Experimental design: WBGT is a useful metric for this type of study for many reasons some of which are touched upon in this manuscript, however it would be beneficial if the authors could comment on the accuracy of their choosen approximation (line 158).

Line 162 is Solar Radiation, could you provide more details on how this is obtained. Other WBGT approximations make use of total sky radiation proportion and downward solar radiation for example.

How and why were the WBGT thresholds used in this study choosen, and are they designed for this approximation of WBGT?

A simple bootstrapping method is used, could you add in more detail why this particular method was choosen.
Validity of the findings: Your findings are very interesting and well presented, to me I find the study similar to the one referenced in line 385 but with a different methodology which is something that could be changed on line 244.

As in section 1, there is scope for more discussion for example line 235 the elderly in ambulances lower than the overall population and the comment on degree days in line 269 to highlight 2 instances.

I also think focusing on what are the key questions your study answers moreso in the conclusion would be beneficial, especially because this is a very important study in terms of projecting heat impact.
Additional comments: I've recommended minor revisions, and would like to thank the authors for their important research on this area. I think just a little more clarification as I state above and a stronger conclusion is needed before 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: BASELINE SCENARIOS OF HEAT-RELATED AMBULANCE TRANSPORTATIONS UNDER CLIMATE CHANGE IN TOKYO, JAPAN
Review round: 1
Reviewer: 2

Output:
Basic reporting: The manuscript is well written with clear explanations.
Experimental design: The data that is used in the study are not clearly stated. Suggest stating the sample size, name of parameters, etc. The data can be supplemented as supplementary information.
Validity of the findings: Lack of comparing the findings with the literature/theory. Suggest to include discussion and comparison with other literature.
Additional comments: The study discussed more mathematical models rather than the concept of heat transfer and its effect.


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: BASELINE SCENARIOS OF HEAT-RELATED AMBULANCE TRANSPORTATIONS UNDER CLIMATE CHANGE IN TOKYO, JAPAN
Review round: 2
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: I thank the authors for addressing my concerns and I have no further comments to make.


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: BASELINE SCENARIOS OF HEAT-RELATED AMBULANCE TRANSPORTATIONS UNDER CLIMATE CHANGE IN TOKYO, JAPAN
Review round: 2
Reviewer: 2

Output:
Basic reporting: The manuscript has been revised by taking into account the feedback from reviewers.
Experimental design: All is good now.
Validity of the findings: Yes, this is performed.
Additional comments: The manuscript is revised and suitable to be published.


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: FUEL TRAIT EFFECTS ON FLAMMABILITY OF NATIVE AND INVASIVE ALIEN SHRUBS IN COASTAL FYNBOS AND THICKET (CAPE FLORISTIC REGION)
Review round: 1
Reviewer: 1

Output:
Basic reporting: In both the introduction and abstract, you talk about the 2017 fires but then never mention them again in the discussion. The discussion should either touch on them again, or remove mention of them from the paper. Conceptually it seems like they are the inspiration of the paper, but do not play any sort of critical part in the research.

Throughout the abstract and paper, rather than saying that [habitat] had [fire trait], say species from [habitat] had [fire trait] e.g. “[species from] Fynbos and IAPs generally exhibited greater flammability” (line 31) as you assessed everything at the species, rather than ecosystem level. You also only sampled 10 species from each habitat type and generalizing to all species from that community does not seem to be  supported by the data.

Supplementary 1 can be expanded as it is missing several recent experimental studies of plant flammability at the branch level. References to the following may also be appropriate in the main text. References cited in these papers also include more recent ecosystem level studies which are likely also relevant.

Alam MA, Wyse SV, Buckley HL, Perry GL, Sullivan JJ, Mason NW, Buxton R, Richardson SJ, Curran TJ (2020) Shoot flammability is decoupled from leaf flammability, but controlled by leaf functional traits. J Ecol 108:641–653. https://doi.org/10.1111/1365-2745.13289

Burton JE, Cawson JG, Filkov AI, Penman TD (2020) Leaf traits predict global patterns in the structure and flammability of forest litter beds. J of Ecol 109:1344–1355. https://doi.org/10.1111/1365-2745.13561

Cornwell WK, Elvira A, van Kempen L, van Logtestijn RS, Aptroot A, Cornelissen JHC (2015) Flammability across the gymnosperm phylogeny: the importance of litter particle size. New Phytol 206:672–681. https://doi.org/10.1111/nph.13317

Cui X, Paterson AM, Wyse SV, Alam MA, Maurin KJ, Pieper R, Cubino JP, O’Connell DM, Donkers D, Bréda J, Buckley HL (2020) Shoot flammability of vascular plants is phylogenetically conserved and related to habitat fire-proneness and growth form. Nat Plants 6:355–359. https://doi.org/10.1038/s41477-020-0635-1

Santacruz-García AC, Bravo S, del Corro F, Ojeda F (2019) A comparative assessment of plant flammability through a functional approach: the case of woody species from Argentine Chaco region. Austral Ecol 44:1416–1429. https://doi.org/10.1111/aec.12815

Wyse SV, Perry GL, O’Connell DM, Holland PS, Wright MJ, Hosted CL, Whitelock SL, Geary IJ, Maurin KJ, Curran TJ (2016) A quantitative assessment of shoot flammability for 60 tree and shrub species supports rankings based on expert opinion. Int J Wildland Fire 25:466–477. https://doi.org/10.1071/WF15047

Wyse SV, Perry GL, Curran TJ (2018) Shoot-level flammability of species mixtures is driven by the most flammable species: implications for vegetation-fire feedback favouring invasive species. Ecosystems 2:886–900. https://doi.org/10.1007/s10021-017-0195-z

Supplementary 2 should be made as table 1 as the selection of species will be of interest to many readers and is important to make the species identities clear considering only 10 species were sampled per vegetation group. It would also be of value to add the family for each species and sort it by the family within the vegetation groups. It is unclear how the species are currently ordered. “Gymnosporia buxifolia (L.)” and “Scolopia zeyheri (Nees)” are not correct names per ICN as they are missing the revising author.

The description of Table 1 appears cut off at the end.

Table 1 was also not intuitive to me and I had to look at it side-by side with supp. 3 to make sense of it. I suggest switching the rows and columns as it is derived from a glm which is typically written from left to right.

In figure 1, you have enough space to include the entire word “thicket” and “fynbos” and I suggest you use that instead of the abbreviation. Same in the legend of Figure 2.

In figure 1 I suggest changing the colors to those which are better for those who are red/green colorblind. Making the points different colors and shapes would also be beneficial in case this ever gets printed as black and white copies.

In figure 2 you also have enough room to not abbreviate vegetation group.

In your discussion, it would be useful to break up your 2nd paragraph into two or more, one comparing your results to studies which assessed flammability at the shoot level like your study, and others that assessed it at a plot or ecosystem level. While shoot level flammability is generally correlated with ecosystem level of flammability, there are differences and opportunities for discussion about those differences (like you have already done for dead material). Incorporation of the citations given above would also be beneficial.

Hypotheses are clear and the data test them directly.
Experimental design: Line 143: Please elaborate in the text how the 30 species were chosen. What were the criteria you used for selection?

Line 155: “samples that did not spontaneously ignite within 120 seconds of pre-heating were ignited with a blow torch and assigned an arbitrary time-to-ignition of 200 seconds.” Why did you use this number specifically?

Line 162: “The dry mass of samples was regarded to be a proxy for the fuel load that samples presented.” How do you justify this? Under most definitions fuel load is the quantity of fuels contributed by a species divided by a certain area of habitat, therefore rare species would contribute a smaller fuel load than common species. Your definition of this entirely ignores the area component. Methodologically, this is the weakest part of the paper and unless you have a very good justification, I suggest you reanalyze these data but either recalculate fuel load with different data, rename it as “sample biomass”, or omit it entirely from the analysis.

Line 174: “ Each of these samples was separated based on stem diameter into fine fuels (< 3 mm) and coarse fuels (> 6 mm).” Why did you not consider 3-6 mm stems? Burger & Bond consider them as a separate category.

Line 178: “calculated as the canopy volume (based on the formula for the volume of a cone)” How do you justify this? Did you specifically choose samples which were approximately conical? Can you add clarification to the methods? I could see an argument for a cylinder (or even a rectangular prism) being a better representation of the volume depending on the sample.

Line 189: “A combined dataset was created for further analyses containing, for all the variables, the averages per species, with the 30 species thus comprising replicates.” I don’t think replicate is the right word to use here since each species is different. Maybe replace replicates with “unique data points”?

Line 193: “Multiple regression models were used” Can you clarify what function you used in the text? lm()?

For the analysis shown in figure 2, did you consider using the unaveraged results between each species (e.g. in a-c analyzing the 21 replicates for each species). As it currently stands, your data is taking an average of an average which affects your error bars and significance tests. For analyzing the unaveraged data you could try using a glmm or gam (depending on normality) with species as a random factor and habitat as the fixed factor.
Validity of the findings: Data are provided in the supplementary material. Providing the R code would also be beneficial.

All analyses and discussion make sense and are supported by the data with the exception of fuel load per my above comments.

Conclusions are clear and supported by the data.
Additional comments: Overall, this is a well written paper with good data and analysis. More recent research in this area needs to be reviewed and the citations above should hopefully be helpful in that regard.

The main reason I suggest major revisions is due to the fuel load issue.


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: FUEL TRAIT EFFECTS ON FLAMMABILITY OF NATIVE AND INVASIVE ALIEN SHRUBS IN COASTAL FYNBOS AND THICKET (CAPE FLORISTIC REGION)
Review round: 1
Reviewer: 2

Output:
Basic reporting: This manuscript presents data from flammability trials on 30 shrub species from the southern Cape of South Africa. The authors sampled species from two vegetation types and also included a set of invasive alien species. This area of work is relatively undeveloped, therefore I think there is scope for exploratory analyses such as these. I did find that the introduction and framing did not provide much explanation or justification for the study. For example, is there field-based evidence that these vegetation types have different fire behavior? From my own knowledge of fynbos and thicket, I would expect higher flammability among typical fynbos species (but with plenty of exceptions). The authors, however, never lay out any background on this.

This comparison across vegetation types is one of two main goals of the manuscript and should be better justified. I also found the category of invasives confusing. Is there theory predicting that non native invasive species should have a particular flammability? The second purpose of the manuscript was to determine important traits that predict flammability. But I found the justification here also a bit confusing. Why were those particular traits measured? The paragraph that starts on line 55 seems to be setting up a justification but it is muddled and ends with a rather weak one: "fuel bed porosity and proportion of fine and dead fuels have received less attention". Perhaps with good reason? Is there any proposed link to the plant ecology that would justify that attention? Or is the goal to find something easier to measure that will predict flammability without the need for trials? What is the actual goal of the trait portion of the study?

There are some references in the text that are missing from the literature cited.

Supplementary table 1 is a good start but is woefully incomplete. Why that particular selection of studies? There are many other important studies omitted. Off hand I can think of several from New Zealand, several from North America, and several missed European studies. The missed literature includes studies on leaf litter flammability in forests, and others on canopy flammability across shrubs and grasses. I also do not agree with some of the characterization of "fuels traits" as listed in that table. The authors don't distinguish canopy from litter flammability throughout despite these fuel types involving very different vegetation-fire feedbacks.
Experimental design: My main concern was that the flammability trials were not well described. I had trouble figuring out how exactly the authors measured the main flammability parameter reported ("burn intensity"). On line 151, the authors cite Keeley 2009 implying that the temperature measure used follows from that citation. Keeley (2009), however does not define the term "burn intensity" but rather discusses uses of the term "fire intensity". However, even broad definitions of fire intensity include the idea of rate of heat release/transfer which is not the same as a peak temperature measured on an unknown material. What do the authors mean by "the maximum temperature reached by a sample during burning"? Temperature of what? gas temperature? Temperature of some object of known thermal properties? I realize that heat transfer is difficult to estimate in flammability trials and I am sympathetic to using proxy measures, but the reader needs to know what exactly is being measured and the justification. Was this number taken from a time series from thermocouples? The original Jaureguiberry et al (2011) paper describes an IR gun aimed at the grill? I'm not sure what that is measuring. I am ok with well-described proxies and have found that this area is an example of where some things work better in practice than in theory: for example, temperature time series summaries from thermocouples often correlate well with other measures of fire behavior. However, the authors should demonstrate that their temperature proxy captures something real about fire behavior. If the authors measured the maximum temperature reached by some object, then I am willing to consider that as a proxy for rate of heat transfer/fire intensity. The figures should be labeled accurately, however, as "peak temperature". I was unconvinced that the "burn intensity" measure was relevant. Is there any field truthing of this measurement? Also note that Jaureguiberry et al (2011) is missing from the literature cited.

How were the samples treated prior to flammability trials? Cut and bagged on ice? Any dry down period prior to burning?  This is all important and the various labs using similar plant "barbecues" all use slight variations.

For the comparisons across species groups, it seems here that the authors used species means. That is acceptable but I would have expected a mixed model with species as a random effect --- I have no problem with the approach but was curious.
Validity of the findings: This paper includes an a priori test of flammability differences across three species groups (fynbos, thicket, and invasives). That is fine, although I would like to see some explanation of how species were chosen and how sampling was randomized. I'm not sure why "invasives" would be thought to have a particular flammability. Perhaps the authors can discuss the types of invasive species (traits, lineages?) found in these communities? I don't believe that the realm of inference for the "IAP" group is really all invasive alien plants and the authors recognize this as they state in the Discussion on lines 304-306! Across the literature, there are examples of invasive species increasing fire spread rates and occurrence (eg grass fire cycle) and invasives decreasing ignitability and spread (some shrub encroachment examples).

I'm not sure what the journal's exact requirements are for data and code. I see that summarized species means are provided, but there is not sufficient data nor any code with which to reproduce the analyses.
Additional comments: In summary: the manuscript would benefit from 1) clearer goals, 2) clearer justification for the exploratory vs the hypothesis testing portions, 3) better description of the flammability trials including some attention to the physics.

Minor comments

- The acronym IAP is not needed. Acronyms hinder understanding and don't really save space.
- This manuscript has both exploratory statistics (the trait-flammability model selection) and hypothesis testing (tests of flammability differences across the species groups). A better explanation of why each approach was used would improve the manuscript. I am also skeptical of any stepwise model selection and there are plenty of good criticisms of stepwise selection in the literature. Why not use the full model? Or represent specific hypotheses with separate models? Or even use model averaging if this is truly just exploratory?
- Line 55: I think this terminology is terrible! Is arrangement of leaves really "extrinsic"? This is an idiosyncratic terminology that is not well accepted.
- Line 73: But some studies explicitly exclude total mass and don't treat fuel load as a "flammability trait" -- presumably because it is so plastic. The framing here cites work from both studies of how leaf litter behaves as fuel and how entire plants (or portions of live plants) behave as fuel. These are different in some important ways but I found the Introduction  was not terribly careful about distinguishing them.
- Line 97. Yes, but do you expect some consistent effects or are these likely to be idiosyncratic?
- Lines 257-260. Yes, an issue with treating a single live branch as a random sample of the canopy is that it is likely to misrepresent live vs dead tissue.


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: FUEL TRAIT EFFECTS ON FLAMMABILITY OF NATIVE AND INVASIVE ALIEN SHRUBS IN COASTAL FYNBOS AND THICKET (CAPE FLORISTIC REGION)
Review round: 1
Reviewer: 3

Output:
Basic reporting: Good. There were a few places where more explanation was needed.
Experimental design: Good, except the experimental equipment should have been described
Validity of the findings: No issues
Additional comments: This is a really useful study of flammability and its drivers for a south African region. The paper is well conceived, clearly and concisely written and the conclusions are valid. I have only minor comments, mostly about adding some additional explanation of the methods.
Also, I would prefer if the supplementary information was included as tables in the main text, especially 1 and 2. However, this might be a style issue with the journal.
While I am no expert on flammability studies, my understanding is that the use of different experimental set-ups could have a large influence on results, and therefore this is an important discussion point that was left out.

Specific comments/
L48. Or the leaf scale.
L144. Can you state whether these were the most common shrub species in each vegetation type? It matters because you compare the fuel characteristics of the species among veg types.
L148. You need to briefly explain the experimental set-up, even if it is more fully explained in Msweli 2020. You refer to a grill. What is that?
L299. You need to back this statement up. IAPs were only slightly more intense and complete than fynbos and did not have the shortest time to ignition. So why do you say they were high flammability?


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: FUEL TRAIT EFFECTS ON FLAMMABILITY OF NATIVE AND INVASIVE ALIEN SHRUBS IN COASTAL FYNBOS AND THICKET (CAPE FLORISTIC REGION)
Review round: 2
Reviewer: 1

Output:
Basic reporting: Everything looks good, all reviewer comments were adequately addressed.
Experimental design: Everything looks good now that the methods have been clarified.
Validity of the findings: Thank you for attaching the R code
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: FUEL TRAIT EFFECTS ON FLAMMABILITY OF NATIVE AND INVASIVE ALIEN SHRUBS IN COASTAL FYNBOS AND THICKET (CAPE FLORISTIC REGION)
Review round: 2
Reviewer: 2

Output:
Basic reporting: Overall, the manuscript was improved. I have no major outstanding issues here.
Experimental design: It is still not clear exactly what material is represented by the maximum temperature measurement. There is no standard thermal properties associated with whatever object the IR gun is seeing. But that seems to be a flaw in the design of the original barbecue method (Jaureguiberry et al 2011). Can the authors make clear exactly what material is the temperature reported?

The authors did improve their description of how species were selected and what the purpose of the groups was.
Validity of the findings: I think the statistics are fine.

In regards to the authors query: "We thus do not understand the discrepancy made by the reviewer between exploratory and hypothesis testing approaches". My answer: If you run a model and report p-values you are doing hypothesis testing.

Regarding biomass/fuel load: Yes, one must include these values but often studies "correct for biomass or total fuel." I think this is dealt with adequately in this paper. The authors effectively standardize for shoot length, therefore mass differences across species arise from canopy traits.
Additional comments: My only real outstanding criticism is the authors still don't really distinguish between canopy fire/shoot flammability and litter driven fire. This does not need to be a huge issue, but the current draft confuses this. I think the authors should simply make very clear their interest in shoot flammability which is relevant given the system and also has a shorter history of study via flammability trials. This should be a relatively minor change to the main text

The authors write in their response letter that "Whether flammability pertains to the canopy or litter is only relevant in plot-scale experiments." This is not true. Past work demonstrates that the same leaf traits can have opposing effects on flammability depending on whether the fire is burning in intact live fuels or in leaf litter. Litter fire vs crown fire also represent quite different natural fire behavior and the relative importance of live fuels vs litter varies by ecosystem and fire regime (litter driven fire is less important globally but is sometimes the dominant fire type in dry forests). The authors are concerned with live fuels and that makes sense given the ecosystem. Simply be clear about this. For example, on line 100, the authors state that fine fuels "enhance flammability". This is true for live fuels but absolutely not for litter fuels where oxygen limitation dominates (eg Scarff and Westoby 2006, Magalhaes and Schwilk 2012).

The supplementary Table 1 focuses on shoot flammability studies but does not state this clearly and also throws in a couple of leaf litter flammability examples and a few studies conducted on individual leaves.
Unless the authors significantly expand this table, perhaps they should focus it only on shoot flammability trials? As it is, they include a few litter flammability trials but only studies with small scale trials that don't produce realistic flame spread rates (but include large number of species). An entire other literature is ignored. Eg work by Kreye, Varner, Schwilk or Kane in the western and eastern US, Ganteaume in France, Australian litter flammability work by Scarff and Westoby, etc. To be clear: I think it is fine to just focus on shoot flammability and canopy fire.  That makes sense given the system the authors are working in. But that must be explicit. Why cite Cornwell et al and Grootemaat et al in that case, however? If those are to be included, it makes no sense to omit earlier work from which that work derived.

A few specific points by line

- lines 330-331: There are more studies than this one that show that finer particles result in reduced flammability of litterbeds as a result of oxygen limitation. The pattern is not limited to gymnosperms and was known before the study cited. See Scarff and Westoby 2006, Kane et al 2008, de Magalhaes and Schwilk 2012 for earlier studies than the Cornwell et al one highlighted. There is also all of the litter bed flammability work by Ganteaume focused on field-collected intact litter beds and going back over a decade. There is also work by western US fire ecologists that was very applied in nature that shows this result (eg Miller and Urban 1999). But oxygen limitation is a litter bed phenomenon, not really applicable to shoot flammability (Schwilk 2015).

- Line 368: I don't think one can subsume litter driven fire vs crown fire into "scale". It seems the authors did not understand my previous comments on this point.

- Line 375: Some studies have examined mixtures of species and looked at non additivity and interactions, at least for litter fuels. See de Magalhaes and Schwilk 2012, van Altena et al 2012, Gormley et al 2020, de Magalhaes and Schwilk 2021.

Some litter flammability studies. These are not necessary to cite because the authors' focus is on canopy flammability. However, the authors seem intent on including litter flammability trials in their table and discussion. I include this here as a start on this literature if the authors do want to include a table that includes both shoot and litter flammability trials.

van Altena C, van Logtestijn RSP, Cornwell WK, Cornelissen JHC. 2012. Species composition and fire: non-additive mixture effects on ground fuel flammability. Frontiers in Plant Science 3: 1–10.

Ganteaume, A., Marielle, J., Corinne, L.M., Thomas, C. and Laurent, B., 2011. Effects of vegetation type and fire regime on flammability of undisturbed litter in Southeastern France. Forest Ecology and Management, 261(12), pp.2223-2231.

Ganteaume, A., 2018. Does plant flammability differ between leaf and litter bed scale? Role of fuel characteristics and consequences for flammability assessment. International Journal of Wildland Fire, 27(5), pp.342-352.

Kane JM, Varner JM, Hiers JK. 2008. The burning characteristics of southeastern oaks: discriminating fire facilitators from fire impeders. Forest Ecology and Management 256: 2039–2045.

Kane, J.M., Kreye, J.K., Barajas-Ramirez, R. and Varner, J.M., 2021. Litter trait driven dampening of flammability following deciduous forest community shifts in eastern North America. Forest Ecology and Management, 489, p.119100.

Kane, J.M., Gallagher, M.R., Varner, J.M. and Skowronski, N.S., 2022. Evidence of local adaptation in litter flammability of a widespread fire‐adaptive pine. Journal of Ecology, 110(5), pp.1138-1148.

Kreye, J.K., Varner, J.M., Hiers, J.K. and Mola, J., 2013. Toward a mechanism for eastern North American forest mesophication: differential litter drying across 17 species. Ecological Applications, 23(8), pp.1976-1986.

Miller, C. & Urban, D. (1999) Interactions between forest heterogeneity and surface fire regimes in the southern Sierra Nevada. Canadian Journal of Forest Research, 29, 202–212.

Scarff FR, Westoby M. 2006. Leaf litter flammability in some semi-arid Australian woodlands. Functional Ecology 20: 745–752.

de Magalhaes RMQ, Schwilk DW. 2012. Leaf traits and litter flammability: evidence for non-additive mixture effects in a temperate forest. Journal of Ecology 100: 1153–1163.

An older review paper on the subject:

Varner, J.M., Kane, J.M., Kreye, J.K. and Engber, E., 2015. The flammability of forest and woodland litter: a synthesis. Current Forestry Reports, 1(2), pp.91-99.

A paper that has a short discussion of litter vs canopy fuels:

Schwilk, D.W. 2015. Dimensions of plant flammability. New Phytologist. 206:286-288


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: FUEL TRAIT EFFECTS ON FLAMMABILITY OF NATIVE AND INVASIVE ALIEN SHRUBS IN COASTAL FYNBOS AND THICKET (CAPE FLORISTIC REGION)
Review round: 2
Reviewer: 3

Output:
Basic reporting: As mentioned in the original review, this was all fine.
Experimental design: As mentioned in the original review, this was all fine
Validity of the findings: As mentioned in the original review, this is all fine
Additional comments: The authors have addressed all of my concerns. Well done.


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

Input:
Title: MAIZE-PEANUT ROTATIONAL STRIP INTERCROPPING IMPROVES PEANUT GROWTH AND SOIL PROPERTIES BY OPTIMIZING MICROBIAL COMMUNITY DIVERSITY
Review round: 1
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The authors collected extensive data through years of field trial investigations of strip rotational intercropping of maize and peanut. And the logic of the manuscript is clear, especially the correlation analysis of bacterial and fungal communities in rhizosphere soil is necessary. However, the section on the results needs to be supplemented with specific values. Also, authors need to check the text format of the manuscript and correct it before accepting it.
Line220: What is the significance P value please? Below is the same.
Line 221-222, Line 230-231, Line232-233: Please specify the increased or decreased value.
Line260, Line263:chang the P to p
Line295-295: The analysis of bacterial and fungal correlations is novel and necessary, and we suggest that the analysis in this part should be more specific, using numerical values to illustrate.
Line300-308: We recommend that the reasons for analyzing yield increases should be discussed in detail, eg, agronomic traits can also have an impact on yield.
Line337: flowering and needle stages
Line375-383: Conclusion Part of the logical relationship is scattered, we suggest rearranging, such as rotational strip intercropping of maize and peanut was increased yield, and then separately explain the reasons for the increase in yield.


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: MAIZE-PEANUT ROTATIONAL STRIP INTERCROPPING IMPROVES PEANUT GROWTH AND SOIL PROPERTIES BY OPTIMIZING MICROBIAL COMMUNITY DIVERSITY
Review round: 1
Reviewer: 2

Output:
Basic reporting: The submitted manuscript aims to understand the effect of rotational strip intercropping of maize and peanut on soil microbial community and crop growth. The manuscript is clearly written, well structured and presents interesting results. I have minor comments below:

- Overall throughout, authors have often interpreted correlation as causation. Please check these statements in the manuscript (for example Line 28 - 30 in the abstract) and rewrite.

- Could the authors please include full forms at the beginning of each section when they first mention a terminology such as PR or PC.

- Line 229 & 239: if there is a significant interaction, the authors must first state what this overall means for that particular result.

- ‘Reasonably’ used in lines 235 and 226 could be replaced with another more relevant word/words.

- Line 256-259: In addition to mentioning that Shannon index increased, authors should also mention what this means for a specific variable to improve the readibility of the results.

- Line 260 – 262: Negative correlation does not indicate the opposite effects of different treatments. This should be rephrased.
Experimental design: - Experimental design and statistical method is unclear. Authors should provide clearer description of variables included in the statistics? Was cropping method the only variable. Did they include year or cropping cycle or Block in the statistical analysis? Were there any random effects?

- Figure legend description does not match the figure in Figure 1
Validity of the findings: The conclusion and end of discussion section are more like a repetition of results. The authors could also mention what is the relevance of their findings and potentially discuss how the results relate back to the origina research questions. The authors could also discuss the limitations of their work.
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: QUALITY OF CARE AND PRESCRIPTION PATTERNS AMONG PATIENTS WITH DIABETIC KIDNEY DISEASE—A LARGE-SCALE COHORT STUDY FROM TAIWANESE CLINICS
Review round: 1
Reviewer: 1

Output:
Basic reporting: Please check the PDF for my comments
Experimental design: Please check the PDF for my comments
Validity of the findings: Please check the PDF for my comments
Additional comments: Please check the PDF for my comments