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: EFFECTS OF PRESERVATION DURATION AT 4 °C ON THE QUALITY OF RNA IN RABBIT BLOOD SPECIMENS
Review round: 1
Reviewer: 2

Output:
Basic reporting: The paper by Song and Zhou is an investigation on the feasibility of using blood samples preserved for different times at 4°C for biobanking and subsequent gene expression studies.
The language is correct, the introduction is clear and literature–based, structure is in line with Peer J guidelines and figures are clear and of good quality.
Experimental design: The present research fits with the scope of the journal.
The research question is well defined, however it is a rather well-addressed question.
The investigation was performed with good technical and ethical standards. Methods are described with a sufficient degree of detail.
Validity of the findings: Most of the papers on a similar topic use room temperature blood preservation after withdrawal (the so-called bench time), and this is generally less than 8 h (see for example Hebels et al, Environmental Health Perspectives volume 121, 4, 2013). Can the authors justify the use of 4°C? Furthermore, considering how valuable are samples to be biobanked, it is difficult to imagine that a researcher would accept the risk to maintain them for longer than 8 h without freezing them. It would definitely be more acceptable to freeze them within 8 h from withdrawal (possibly after fractionation). Indeed, the Authors show that RNA purity and integrity are not significantly modified up to 3 d for whole blood and to 7 d for leukocyte RNA; however, they recognize that RNA integrity is a parameter related to ribosomal RNA, and they show a degree of variability of specific mRNAs with time. Thus, it is possible that small but relevant differences in mRNA integrity take place during the time at 4°C, that could have an impact on future gene expression studies.
It is doubtful that the observed differences in the various tested mRNAs is due to gene expression changes taking place at 4°C, as the Authors seem to suggest in the discussion (lines 295-316). It appears more likely that some mRNA degradation can take place during this time, so that the expression “mRNA levels” should be used in place of “gene expression” here.
Additional comments: No further comment


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

Input:
Title: EFFECTS OF PRESERVATION DURATION AT 4 °C ON THE QUALITY OF RNA IN RABBIT BLOOD SPECIMENS
Review round: 1
Reviewer: 3

Output:
Basic reporting: Introduction:  From line 74 the sentence should be excluded since it is not well formulated. It is necessary  to explain   what was the focus of the investigation and why that assessment was performed on rabbit blood instead of a human.

Discussion and conclusion: In lines 223 and 319, the sentences are not written appropriately. It should be reworded to be more understandable.
In line 324 the part of the sentence „but other studied genes were relatively stable“ should be replaced with „the expression of genes was stable“
Experimental design: Material and Methods:

Ethical approval is needed to be included in Material and Methods.
The authors have to explain why euthanasia methods are performed for the rabbits. Also, it is not clear if the same animal was used for successive blood sampling.
Validity of the findings: The paper describes diligent findings on information about effects of preservation duration at 4 °C for different preservation durations, from 1 hour to 7 days, on the quality of RNA in rabbit blood specimens. This paper addresses an interesting question: whether the quality of RNA from whole blood and/or leukocytes isolated from rabbit blood in regards to the effects of preservation duration at 4 °C could be a choice approach to valuable source of RNA for profiling gene expression.
Additional comments: Strong point:

The authors assessed that the purity of whole blood RNA and leukocyte RNA does not induce significant change after blood is stored at 4 °C for different preservation durations. Otherwise, the integrity of whole blood RNA and leukocyte RNA  is degraded after a preservation duration for more than 3 days at 4 °C . The gene expression, especially of whole blood RNA, can decrease over the increased preservation duration.


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 RAPID AND ACCURATE METHOD FOR THE DETECTION OF FOUR AMINOGLYCOSIDE MODIFYING ENZYME DRUG RESISTANCE GENE IN CLINICAL STRAINS OF ESCHERICHIA COLI BY A MULTIPLEX POLYMERASE CHAIN REACTION
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript requires significant editing by a native English speaker. As written, it is ambiguous and difficult to read. I have attempted to edit the abstract and introduction as an example of the extensive work that needs to be done.
Experimental design: The multiplex PCR strategy is reasonable. The data presented demonstrate that the approach is sounds and can identify four aminoglycoside resistance elements. What is not clear is why these particular genes were selected and wether this information will alter clinical practice as suggested in the abstract and introduction. Nevertheless, as a standard method to detect these genes, the work is sound.
Validity of the findings: No comment
Additional comments: The manuscript needs to be rewritten with improved English language use and clear reasons why these gens in particular were elected and how the information can be used. Technically, the work is sound.


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 RAPID AND ACCURATE METHOD FOR THE DETECTION OF FOUR AMINOGLYCOSIDE MODIFYING ENZYME DRUG RESISTANCE GENE IN CLINICAL STRAINS OF ESCHERICHIA COLI BY A MULTIPLEX POLYMERASE CHAIN REACTION
Review round: 1
Reviewer: 2

Output:
Basic reporting: See below
Experimental design: See below
Validity of the findings: See below
Additional comments: This article by Yaoqiang Shi et al. and entitled “  A rapid and accurate method for the detection of four aminoglycoside modifying enzymes drug resistance genes in clinical strains of Escherichia coli by the multiplex polymerase chain reaction ” is interesting and fits well with the idea outlined in the introduction and discussion, namely that a quick identification of the involved pathogen(s) and their drug susceptibility pattern is essential for a good antibiotic use.

Although already several groups published papers in this sense the article under focus here has it’s merit but, honestly, I think that it is not ready for publishing. I propose a major revision.

Main comments:

English style and/or phrasing is often not qualitatively good for the standard of the journal as well as for the scientific community. It is not specifically the wording or style but the fact that this could give misunderstandings or confusions. Therefore I propose that a native English speaking scientist helps the team to adjust.

I suggest an edit to the title:

Title:
A rapid and accurate method for the detection of four aminoglycoside modifying enzymes drug resistance genes in clinical strains of Escherichia coli by the multiplex polymerase chain reaction:

 suggestion:
A rapid and accurate method for the detection of four aminoglycoside modifying enzyme drug resistance genes in clinical strains of Escherichia coli by a multiplex polymerase chain reaction

I suggest edits to the Abstract:

Background. Antibiotics were and are (were the most) very effective drugs in the treatment of infectious diseases. Aminoglycoside antibiotics were one of the most common antibiotics in the treatment of bacterial infections, while the development (or emergence) of drug resistance against those medicines (of these antibiotics were very) serious (or maybe better significant).

AIM: develop an efficient, rapid, accurate and sensitive detection method, applicable in routine clinical use

Methods. (To establish an efficient, rapid, accurate, sensitive, and multiple drug
resistance detection method for clinical Escherichia coli.)
Escherichia coli (E. coli) was taken as model organism for the development of  a rapid, accurate, and reliable multiplex polymerase chain reaction (M-PCR) (system) for the detection (identification) of (the) four aminoglycoside modifying enzyme(s) (AMEs) resistance genes Aac(6')-I, Aac(3')-II, Ant(3'), and Aph(3') (was established).

(Also,) The M-PCR was used to detect the distribution of AMEs resistance genes in 236 clinical strains of E. coli. (And) The results were verified by the
simplex polymerase chain reaction (S-PCR).

Results. The results of M-PCR and S-PCR
showed that the detection rate of Aac(6')-I, Aac(3')-II, Ant(3'), and Aph(3') were 32.6%, 59.3%, 23.7%, and 16.9% in 236 clinical strains of E. coli respectively. Compared with the traditional methods for the detection and identification, (a) the rapid and accurate M-PCR detection method was established for the detection of AMEs drug resistance genes, which was significant to guide the reasonable(y) use of drugs, and prevent the spread of bacterial resistance.

…
Comments on the abstract:
Already in the abstract I find that for example the conclusion is certainly not clear in the sense that it says that the method developed “was significant to guide “reasonably” the antibiotherapy and prevent the spread of bacterial resistance…I find it to vague, also further in the paper which is technically and scientifically not at the required level of the journal.

Scientifically I find that there is too much room for subjectivity.  In order to be accurate and specific I propose to define what is meant by Multi Drug Resistance (MDR) since I have the impression MDR, here, is not well defined and could lead to confusions. A good paper related to this issue is the one of Magoriakos et al. (in Clin Microbiol Infect  2012:18:268-281) “ Multidrug-resistant, extensively drug-resistant and pandrug-resistant bacteria: an international expert proposal for interim standard definitions for acquired resistance” in which guidelines for MDR and definitions are discussed and proposed.

Since we only are concerned about aminoglycosides it would be useful to add following review by Krause et al. “Aminoglycosides: an overview” (Cold Spring Harbor Perspectives in Medicine, 2016; available at www.perspectivesinmedicine.org).

I would also suggest to shorten the paper by cutting in the introduction as well as in the discussion where we find several redundancies.

This will make the article much better and focussed without losing the importance of the message that the need is great for the development and daily use of more rapid pathogen identification and drug susceptibility tests in routine medical settings. In that frame this work has to be evaluated and every contribution into well described and evaluated technical approaches, especially the identification and validation of ‘good targets’ (genes in casu is) necessary and useful. This statement is still through although a lot of similar technical research and even commercially available assays is conducted and exist. An interesting and inspiring article related to this is the one of Chavada and Maley in the Open Microbiology Journal (2015, 9, 125-135) entitled “Evaluation of a Commercial Multiplex PCR for Rapid Detection of Multidrug Resistant Gram Negative Infections”.

No in-depth validation was performed and should be performed. Only 5 “MDR” were tested on the background of xxx  E. coli strains. Comments? Check!

Was it the aim only to target E. coli ? Why not other Enterobacteriaceae where the same ABRs are found? Could this also not be extrapolated to some non-fermenters like P. aeruginosa

Shortcomings of genome technologies, in casu PCR-technology: could not it be that a gene is present but not well expressed, meaning that the phenotype should give a certain susceptibility?

A recent paper amongst others is the one of Moran et al. (in J Antimicrob Chemother  2017; 72:700-704) “Prediction of antibiotic resistance from antibiotic resistance genes detected in antibiotic-resistant commensal Escherichia coli using PCR or WGS”. This aspect is not discussed neither just mentioned. I think this issue needs a certain attention in the discussion.

Another interesting paper is the article of Su et al. in J Clin Microbiol (2019, 57: 1405-1418) entitled “Genome-based prediction of antibiotic resistance”.

Both articles are very interesting and could be taken up in the global discussion as well as in the references.

The use of “etc. should absolutely be avoided in a scientific and technical paper! It occurs several times.

Also I would suggest that for example more rigour should be applied in for example:

See introduction line 49: …were mainly used to treat infections caused by facultative aerobic gram-negative bacteria, such as Escherichia coli, Klebsiella pneumoniae,  and non-fermenters like Pseudomonas aeruginosa,  etc 15,22,41.

Line 56: … there is an urgent need …


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 RAPID AND ACCURATE METHOD FOR THE DETECTION OF FOUR AMINOGLYCOSIDE MODIFYING ENZYME DRUG RESISTANCE GENE IN CLINICAL STRAINS OF ESCHERICHIA COLI BY A MULTIPLEX POLYMERASE CHAIN REACTION
Review round: 2
Reviewer: 1

Output:
Basic reporting: .
Experimental design: .
Validity of the findings: .
Additional comments: Title OK
Line 37-39:

This technique can be used for the clinical detection as well as the  surveillance and monitoring of the spread of those specific antibiotic resistance genes.

Line 47:
I would delete …”and super bacteria.” Since I find it redundant with “pan-drug-resistant”

37-51: OK with addition of the Magiorakis definitions.

Line 52: Addition of Krausse et al. OK

Line 53: please correct the typing error in the word “Gram”
Line 56: please correct However, bacterial can… into However, bacteria can…
Line 59: add into the sentence: … tolerance has become a global worldwide concern.
Line 93: I would change into … to timely guide the clinical antibiotic therapy.

Line 102:
Please change into or rephrase as suggested:
Given that the M-PCR can detect multiple genes simultaneously we aimed to develop a M-PCR system for the detection of the four most widely spread AME genes. The reaction system was verified in 237 clinical E. coli strains.
Lines 151 and further:

I would suggest to rephrase it as follows if I understood well the meaning:
Based on the drug sensitivity information of 237 clinical strains of E. coli, five isolates (…) previously tested for the Aac(6’)-Ib, Ant(3”)-Ia, Aph(3’)-Ia and Aac(3)-II by S-PCR were each tested for the M-PCR.

Line 164: 236 or 237?

Line 177: I think the …(data unknown)… means and should be …(data not shown)…?

Line 210:
I suggest to rephrase into
…, and the use of antibiotics or antibiotic therapy should be more standardized and technically, where possible, monitored.

Line 213:
Although WGS is gradually reaching maturity, while sequencing costs are drastically decreasing, it is however still relatively costly, complex in operation and requires for analysis specialised personnel like bioinformaticians


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: INHIBITION OF EXCESSIVE MITOPHAGY BY N-ACETYL-L-TRYPTOPHAN CONFERS HEPATOPROTECTION AGAINST ISCHEMIA-REPERFUSION INJURY IN RATS
Review round: 1
Reviewer: 1

Output:
Basic reporting: Literature background, figure structure and hypothesis are well established. English proficiency needs work.
Experimental design: Research question and experimental methodology well defined.
Validity of the findings: Conclusions and the data are well presented, needs few things to elaborate (see comments).
Additional comments: Comments on manuscript # 41725 " Inhibition of excessive mitophagy by N-acetyl-L-tryptophan confers hepatoprotection against Ischemia-Reperfusion Injury" by Li et. al.

The author studied the role of N-acetyl-L-tryptophan in hepatoprotection by inhibiting mitophagy against Ischemia-reperfusion injury in vitro and in-vivo.

Following things need to be addressed in the MS.
1.	Line 186-200- Author studied the autophagy markers in rat liver tissues after I/R or I/R+L-NAT Treatment-These are all nice studies. But, what is missing how different is I/R rat compared to L-NAT treated ones in terms of metabolism, body weight, whole liver morphology or pathology or even other symptoms, like energy loss in terms of movement for food or water or balance on a beam from one end to another. The animal study needs to be elaborated or explained further in MS.

2. Figure 2: H&E staining should be included to see the proliferation/tissue integrity of the tissues used for Immunofluorescence staining?

3.	Proofreading is missing all over the MS, e.g. line 116 and 150, Where is Dark? Should be in dark. Line 148: Living cells-should be live cells. Line 220-221: improve mitochondrial disfunction –that makes reverse of the result-should be: rescue mitochondrial dysfunction.

The paper needs nice English proficiency. Comments above- will make the MS better.


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: INHIBITION OF EXCESSIVE MITOPHAGY BY N-ACETYL-L-TRYPTOPHAN CONFERS HEPATOPROTECTION AGAINST ISCHEMIA-REPERFUSION INJURY IN RATS
Review round: 1
Reviewer: 2

Output:
Basic reporting: 1.	All acronyms should be defined (expanded form in parenthesis) whenever mentioned for the first time in the text, including abstract.
2.	English grammar and sentence structure need significant attention throughout the manuscript.
3.	Some citations are either missing or mentioned very late in the text, for example,  line 55-54 and 67-69. Also, Li et al 2013 (line 60) is missing from the reference section.
4.	Figure 1B and C- relative expression of beclin 1 compared to the housekeeping gene GAPDH in Fig 1C at 4 hours is higher than 1hr. However, in the Weston blot image that does not appear to be the case. Similarly, the ratio for 24 hours seems lower than 6 hours, but in the western blot image it appears to be same as 6 hours. This discrepancy should be resolved. Additionally, there are no error bars on Sham group. It is important to mention where biological replicates for the sham group were included in the experiment or not, to account for baseline variation.
5.	There is no error bar in sham group in any of the graphs. Does that mean no biological/experimental replicates were used?
6.	In figure 1A, there seem to be a marginal increase in the relative mRNA expression however, there is a two-fold induction in the same in figure 2A. It is important to address the inconsistency between the experiments. Also, the timeline that was used as end point in Figure 2 should be mentioned in the figure legend.
7.	In Figure 2 J, last panel, P62 is unusually high for sham control. What does the author think could be the reason? Similar observation for Figure 3J. It is important to include IgG controls in the panel for both Figure 2J and 3J to account for non-specific staining.
8.	In figure 4 and 5, quantitation of autophagosomes and damaged mitochondria (if there are multiple image fields) will be more informative than providing single images for different conditions.
9.	In line 221, the authors mistakenly mention mitochondrial function as dysfunction as it is implied that L-NAT can improve mitochondrial function.
10.	Figure 7- a quantitative graph for the percent colocalization of beclin 1 with the mito-tracker dye should be included.
11.	In Figure 8 the cell viability in L-NAT treated group does not seem to reach to sham level and is only marginally higher than the H2O2 group. Can the author speculate the reason behind it in context with protective effect of L-NAT. What dose of 3-MA was used to compare for beclin-1 expression in the mRNA and protein quantitation?
Experimental design: Biological/experimental replicates are missing from the control samples.
Validity of the findings: Some findings, as outlined in the basic reporting section, need to be re-evaluated carefully.
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: INHIBITION OF EXCESSIVE MITOPHAGY BY N-ACETYL-L-TRYPTOPHAN CONFERS HEPATOPROTECTION AGAINST ISCHEMIA-REPERFUSION INJURY IN RATS
Review round: 1
Reviewer: 3

Output:
Basic reporting: No comment.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: The paper entitled “Inhibition of Excessive Mitophagy by N-acetyl-L-tryptophan Confers Hepatoprotection against Ischemia-Reperfusion Injury” by Huiting Li et al investigated the effects of L-NAT in HIRI models both in vitro (BRL cells) and in vivo (Sprague-Dawley (SD) rats). The authors concluded that L-NAT could protect hepatocytes against HIRI by inhibiting excessive mitophagy.

Overall, conclusions are well stated and linked to original research question. Literature is relevant and well referenced.

However, the following concerns need to be addressed:

The authors claim that BRL cells and rats are under I/R injury after H202 or vessel clamp surgery treatment, and literature indicates that autophagy correlates with I/R injury. While the authors clearly showed the marker proteins for autophagy in all experiments, it is unclear to me what is the evaluation criteria for I/R injury. Are there markers to indicate that the treated rat or cells are correct I/R models? What are the evaluation factors to assess that rat and BRL cells are really injured after respective treatments?

The author concluded that L-NAT could protect hepatocytes against HIRI by inhibiting excessive mitophagy. Please explain differences between “normal mitophagy” and “excessive mitophagy” How did the authors differentiate between the two during the rat and cellular experiments?

Are all proteins in figure 2E from the same gel (membrane) but with different contrasts? It seems that P62 and GAPDH are from a different gel.

Please include more descriptions for the immunofluorescence result in Figure 2J. How did authors quantify the fold changes between I/R and I/R-LNAT group?  Please also use other color (eg. white) for “DAPI” characters on top.

The autophagosomes in figure 4 and the mitochondria in figure 5 are quite similar in shape and morphology. Since no specific antibody was used in transmission electron microscopy, the author should specifically explain how mitochondria and autophagosomes are differentiated under TEM. Otherwise specific bio-markers for mitochondria and autophagosome need to be used to distinguish between the two.

Please include more detailed description for Figure 4 and 5, including how did the volume changes calculated?

In I/R group of Figure 5, it looks like the majority mitochondria still have normal shape. Please explain.

The introduction needs more detail. I suggest that authors improve the description at lines 62 to provide more justification for how “mitophagy may be a new therapeutic target for HIRI”.

The English language should be improved to ensure that an international audience can clearly understand your text. Corrections including but not limited to:

Line 67, “And another study also shown that L-NAT, an antagonist of the neurokinin 1 receptor (NK-1R) known to disrupt the binding of Substance P  (SP) to NK-1R, significantly protective effect on neurodegenerative diseases.” may change to  “Another study also shown that L-NAT, an antagonist of the neurokinin 1 receptor (NK-1R) known to disrupt the binding of Substance P  (SP) to NK-1R, shows significantly protective effect on neurodegenerative diseases.”

Line 71, “but also alleviate the mice model of ALS and cerebral ischemia / hypoxia injury in vivo” change to “but also alleviate ALS and cerebral ischemia / hypoxia injury in mouse model in vivo ”

Line 73, “Recent work confirmed that hepatoprotective effects of…” change to “recent work confirmed the hepatoprotective effects of…””

Minor corrections:

Put more space between 2E and 2F, 3E and 3F


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: INHIBITION OF EXCESSIVE MITOPHAGY BY N-ACETYL-L-TRYPTOPHAN CONFERS HEPATOPROTECTION AGAINST ISCHEMIA-REPERFUSION INJURY IN RATS
Review round: 2
Reviewer: 1

Output:
Basic reporting: The authors have made significant revisions in the English language and have made most of the suggested changes in the manuscript. Therefore, at this point, the manuscript can be accepted for publication.
Experimental design: Satisfactory
Validity of the findings: satisfactory
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: INHIBITION OF EXCESSIVE MITOPHAGY BY N-ACETYL-L-TRYPTOPHAN CONFERS HEPATOPROTECTION AGAINST ISCHEMIA-REPERFUSION INJURY IN RATS
Review round: 2
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The author has answered all questions raised.


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: REDUCED FISH DIVERSITY DESPITE INCREASED FISH BIOMASS IN A GULF OF CALIFORNIA MARINE PROTECTED AREA
Review round: 1
Reviewer: 1

Output:
Basic reporting: Basic reporting:
English usage: English usage could use minor revision. For example, there are instances of disagreement between sentence subject and verbs (e.g., line 91: “…this group play important…”, line 284: “ CONAPESCA inspectors’ controls…”).
References/Background: Functional metrics are conceptually underdeveloped in the paper. The differences in inference and conclusions drawn between ‘traditional’ ecological metrics and functional metrics needs more background in the introduction and more discussion in discussion/conclusions. In its current form, functional metrics are introduced in the last two sentences of the introduction (line 97) and are only introduced to say they have recently been used. Besides the area of functional metrics, literature cited is adequate.
Structure/Figures/Data: The article structure is acceptable. Raw data provided is easy enough to navigate and appears robust. Figures are clean and well done. Many of the figure and table captions need to be reworked, as they inadequately describe the figure/table and lack necessary information to allow figure/table to be assessed independently. For example, the caption for Figure 1 needs to include that points on the map relate to the sampling locations (Also, PNZMAES versus PNAES in the figure). Further suggestions are included within the attached files.
Self-contained: The current manuscript needs to further motivate the questions and hypotheses/predictions for the study. As it is currently written, there are no or weakly developed questions and reasoning for the use of functional metrics. As such, it is unclear whether the data presented, and response variables chosen, address the questions or were chosen because they have been used in other publications. If correctly reframed in revisions, data would likely be adequate.
Experimental design: Experimental design
Aims & Scope: The research fits the scope of the journal.
Research question: Not clearly defined research question/s and hypotheses/prediction. The objective as stated is, “In the current study we aimed to evaluate how fish assemblages changed through a 13-year study period at PNZMAES”. This does not motivate the question about areas of different use, for example. See above and attached file for further comments.
Rigor/Methods: The methods, as written, lack the details to fully assess whether certain aspects of the study can be addressed. I highlight a few I find very important: 1) there is inadequate information about the underwater visual censuses to have an idea of the precision, bias, and power to detect changes. Were the censuses conducted by a single diver? Was there any attempt (e.g., training, repeat censuses, etc.) to estimate inter-diver bias amongst the, likely, many divers over the decade of sampling? 2) Looking at the raw data, there is a general decrease in sampling effort from the beginning of the sampling period to the last years (~400 m2 to 300 m2). Can the decrease in species richness be explained by the decrease in effort? Where are the researchers on the yield-effort curve? What effort is needed to adequately characterize the species pools? Is this something to be concerned or have they accounted for this? 3) Line 105-106: Does the study represent a 3- or 13-year period of “MPA effect”? This is important for how the study is framed and how much inference can be drawn about the impacts of a management action versus ‘natural’ ‘background’ patterns/trends. This should be made very explicit in the methods and in framing the study.
Validity of the findings: Validity of findings
Data: See above.
Conclusions: The discussion needs a large revision. There is inadequate discussion around the results of the manuscript in the last half of the discussion. Further, there are areas of speculation that need to be noted as such and sections that move beyond speculation, beyond tangential to the questions, and near the point of unsubstantiated accusations against an international non-profit that are not suited for this forum.
Additional comments: General overview: Ramírez-Ortiz and colleagues explore temporal trends in ecological and functional diversity of reef fish assemblages of a multi-use marine protected area within the Gulf of California. They claim a loss of species and certain functional diversity metrics and a lack of enhancement of commercial fish stock bring in to question whether the goals of this MPA are being met. The data the authors present is interesting and provides for the comparison of fish assemblage responses to management with relatively long-term data.
However, I cannot recommend publication of the manuscript in its current form and suggest that substantial revisions would be necessary. I have 5 general areas that I would like to highlight as especially important to address, and also provide more specific and line by line comments in attached documents. Important areas for improvement/revision are:
1)	The introduction doesn’t clearly state questions and hypotheses/predictions or motivate the use of functional metrics over ‘traditional’ ecological metrics. Why are functional metrics useful, how can they “…sensitively detect early changes in assemblage structure and instability, through redundancy and complementary processes”? and can predictions be made about how management of MPAs should impact these metrics? Is increasing functional dispersion (for example) a predicted positive outcome of no-take zones?
2)	The methodology surrounding the UVCs was inadequate to assess if errors, precision, biases were accounted for through the decade long study. This is especially important because there seems to be a decrease in the effort (transect area surveyed) through time. If conclusions about species richness are made, there needs to be assurances that richness measurements are not dependent upon effort. I admit this is slightly outside of my expertise, but if 300 m2 is well established to be adequate to characterize species richness in reefs and richness estimates are well saturated at this amount of effort a relevant citation would suffice. If this is not established, the authors should be able to show that richness measurements are well saturated and not dependent on effort so as to allow comparisons through time.
3)	It needs to be clearly stated how long management has been implemented in the park if conclusions about how management/MPA use zones are to be made. Are there 2-3 years of pre-MPA data (2005-2007) or ~9 years (2005-2014)? This is needs to be clear for readers to assess the possible implications for the trends observed.
4)	It seems like there is a clear result of shifting body size distribution towards larger body size individuals in the assemblage over time. This appears to lead to divergent and opposing temporal patterns in ecological and functional metrics between density and biomass. I suggest this is an important trend that cannot be dismissed, as loss of density may be, partially, attributable to body size shifts to larger species and individuals within a species. The current lack of comment on it anywhere in the manuscript is notable.
5)	The discussion needs a large revision, especially the last half. There is a large ‘digression’ that distracts from the findings in the paper and is not suited for this forum. Instead, more attention should be paid towards the results in an objective manner. Much of the discussion focuses on negative trends or losses of function. But depending on whether density or biomass is looked at the opposite observations can be made in many cases. Right now, it seems like there is a bias towards highlighting shortcomings. If explicit questions and hypotheses and predictions were clearly stated earlier, the reporting of the results and choice of what to highlight would seem less arbitrary.


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: REDUCED FISH DIVERSITY DESPITE INCREASED FISH BIOMASS IN A GULF OF CALIFORNIA MARINE PROTECTED AREA
Review round: 1
Reviewer: 2

Output:
Basic reporting: This short manuscript, untitled "Reduced fish diversity despite increased fish bioass in a Gulf of California marine protected area" presents interesting results due to the long-term database (13 years).
This study reaches the scope and aims of the revue but need corrections before publication.
My concerns are about the presentaton of the results, that are sometimes difficult to read (figures) or in supplementatry material.

Title : add a upper-case letter to "marine protected area"

L30-31 : « we detected a reduction in fish biodiversity… » In which zone ? This result should be detailed.

L32-33 : a part of this sentence is missing.

L34-35 : reword this sentence.

L55 : « it is rapidly incrasing », reword this sentence. For exampe, by « it is currently growing rapidly »

L80-88 : I found that information on anthropogenic impacts/influence or human activities in the MPA are missing in the introduction ?
Frequentation of the MPA, numbers of tourists per year ? In which area ? The quantity of fished organisms in the sustainable and artisanal areas ? As well as the effort of management in the MPA (number of monitor guards ? penalities to people that do not respect the rules ?)

Literature references are well cited and listed at the end of the manuscript.
Only one change to do :
L272 : change « Bellwood et al. 2011 » by « Bellwood, Hoey & Hughes, 2011 »

A lot of tables (statistical results) are given in supplementary materiel, that I think is a shame.

The raw data are shared and are clearly presented.
Experimental design: The methods is well presented and well isllustrated with scientific references.

Figure 1 : where are located the sustainable use areas ? Does the MPA encompasses the islands ? The limits of the MPA and the different zones are not clear in Figure 1.

L120 : «  we obtained 320 transects… ». it could be usefull to have the numbers of transects done per year (before or after the establishment of the MPA) and per area (no-take, sustainable, traditional).

L139-149 : the definitions of the three indices and the mode of calculation are not clear, and it is difficult to understand the meaning of these indices based on Figure 2.

Figure 3 : authors should add in the caption, the meaning of « (B) » and « (D) » in the legends axes.

Figure 3 : the caption of figure 3 should be « Fish diversity, density and biomass according to the levels of use in the PNZMAES » (same remark for Figure 4, this figure shows also the evolution of fish density and biomass, not only diversity).
Validity of the findings: Figure 3 : fish density is the only parameter that showed difference between the three zones areas of the MPA. Do densities are different between the three zones ou between « traditional use zone » and the two other ? It looks like black and blue boxplot are not different.

Figure 4 : The red and green circles are refering to which zone area ? It appears that specific richness increases in traditional use area, while a red circle is mentionned in the small graph.

Figure 4 : grey lines  (sustainable use) are not visible.

L184 : « species richness and density decreased while biomass increased.. » That means that mean fish size increased ? Why the fish sizes are not presented in the different figures ? Which species increase in size ?

L305 : isn’t there some information about benthic composition of the reef during the survey census period ? Evolution of coral cover or macroalgae cover ?

L339 : « habitat alteration, lowering in larval recruitment and global change » can also be included in the anthropogenix impacts (not as « natural factors »).
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: REDUCED FISH DIVERSITY DESPITE INCREASED FISH BIOMASS IN A GULF OF CALIFORNIA MARINE PROTECTED AREA
Review round: 2
Reviewer: 1

Output:
Basic reporting: The language of the text has been much improved. A few minor edits include:
Line 60: 'persist' to persists
Line 64: add 'to' following due...
Line 144: subtle changes at *the* community level
Line 191: 'of' not if
Line 364: 'increase' not increment
Experimental design: Revised manuscript clearly defines questions, addresses my concerns regarding changing effort surveys throughout the study, and greatly improves the description of what was done.
Validity of the findings: Fine.
Additional comments: Authors should be commended for their efforts to address my previous concerns in this revised article. It is much improved in my opinion.


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 PEBP GENES IN PEARS AND THE PUTATIVE ROLE OF PBFT IN FLOWER BUD DIFFERENTIATION
Review round: 1
Reviewer: 1

Output:
Basic reporting: .
Experimental design: .
Validity of the findings: .
Additional comments: Review PeerJ #44784

The authors tried to clarify the role of PEBP gene families, especially FT gene, and they suggested that pear FT genes was regulated by day length and circadian clock.  Moreover, transgenic arabidopsis over-expressing FT, suggested that FT played a role in not only promoting flower bud differentiation, but also regulating the balance between vegetative and reproductive growth.  The former parts on bioinformatics were also important but not so interesting; rather latter parts on physiological analysis should be more strengthen.  Accordingly, I felt, direct results were not provided in the current manuscript for supporting the authors’ conclusion.  The authors should provide the status during flower bud development including initiation/induction stages, along with the expression data.  Without the flower bud status, it was impossible to get a authors’ conclusion.
Other suggestions:
1) Provide the similar data to FT on TFL1.
2) In pear, the movement of FT from leaves to meristems may not occurr because the grafting of wild scions of European pear on the FT-over-expressed pear rootstocks did not cause the early flowering phenotypes of the grafted wild scion cultivars.  Therefore, the leaves as PCR materials for FT expressions may not be adequate.  I recommend the use of buds instead of leaves.


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 PEBP GENES IN PEARS AND THE PUTATIVE ROLE OF PBFT IN FLOWER BUD DIFFERENTIATION
Review round: 1
Reviewer: 2

Output:
Basic reporting: 1- The writing of the manuscript is quite coarse, and the authors may use professional editing service to improve the logic flow and accuracy of the manuscript. Please rewrite the Introduction section. Specifically, rearrange the logic between paragraphs, and add more new references (recent research progress).

2- Please rewrite the Abstract section. The highlights of the study were not highlighted.

3- Journals often require both the manufacturer's name and location (city, state, country) for specialized equipment, software, and reagents. .

4- Please ensure that all gene and protein names are formatted consistently throughout the manuscript and adhere to the appropriate conventions. E.g. ‘PEBP’ in PEBP genes needs italics.

5-Abbreviations and acronyms are often defined the first time they are used within the abstract and again in the main text and then used throughout the remainder of the manuscript. E.g. FT.
Experimental design: 1.The statistical analysis was not appropriate as it was not provided by the authors. How many treatments were used? How the data was analyzed?

2.How many replicates do you use for your transcriptome? The transcriptomic analysis is based on your replicates of samples among different treatments. You should highlight how many replicates do you test and show the repeatability error in Results section to make sure this experiment is reliable. The content of transcriptome sequencing needs to be described in detail in the method.
Validity of the findings: The overall concept of the research and the experimental design are correct.
Additional comments: I carefully read the submission titled ‘Genome-wide identification of the PEBP genes in
 pears and the putative role of PbFT in flower bud differentiation’. The manuscript would be of general interest; however, the some points must be addressed.
I do accept this paper for further revision after these topics are corrected/clarified.


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 PEBP GENES IN PEARS AND THE PUTATIVE ROLE OF PBFT IN FLOWER BUD DIFFERENTIATION
Review round: 2
Reviewer: 1

Output:
Basic reporting: see below
Experimental design: see below
Validity of the findings: see below
Additional comments: The authors tried to improve the manuscript, but I did not satisfy their revision as indicated the below reasons.
1) They did not provide the expression data on TFL1.  It was strange that the expression of TFL1 homologue did not change so much in Fig. 4A.  Usually, TFL1 expression could decrease before bud differentiation (=before the increase in FT expression).  Discuss the reason(s) for no-expression on TFL1 homologue; Pear TFL1 did not express in the leaves so that I asked to carry out expression analysis of FT and TFL1 in the apical buds during bud differentiation as pointed out in my first review.
2) In line 222, the authors used “maybe”, which means that there are no direct data showing the expression regulations of FT through day-length/circadian clock.  Therefore, provide direct evidences on their relationships.
3) Provide the reason for focusing the Pyrus bretchneideri from Figure 3.
4) In Fig. 4A, change to XA, XB････, to 30 DAF, 45 DAF.
5) In Table 1, distinguish the pear species by the horizontal lines.
6) In line 218, “The expression patterns with the qRT-PCR experiment were shown in Figure 2C”.   Fig. 2 is correct?


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 PEBP GENES IN PEARS AND THE PUTATIVE ROLE OF PBFT IN FLOWER BUD DIFFERENTIATION
Review round: 3
Reviewer: 1

Output:
Basic reporting: The author has done the revision better.
Experimental design: no comment
Validity of the findings: no comment
Additional comments: 1. Punctuation and formatting errors persist. Need to be perfected. E.g: ‘assaying samples. (Since samples harvested at 30DAB and 90DAB have been confirmed as not critical samples in our research (unsubmitted), both’.

2. What does XA, XB, XC, XD and XE stand for?

3.Table 1-Capitalize the first letter of ‘gene length’.

The format of the article needs further specification.


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 PEBP GENES IN PEARS AND THE PUTATIVE ROLE OF PBFT IN FLOWER BUD DIFFERENTIATION
Review round: 3
Reviewer: 2

Output:
Basic reporting: This MS entitled 'Genome-wide identification of the PEBP genes in pears and the putative role of PbFT in the flower bud differentiation' reported the isolation and the functional research of  PbFT in pear.  They found that PbFT might play a role in not only promoting flower bud differentiation, but also regulating the balance between vegetative and reproductive growth.
Experimental design: The experiments were well designed.
Validity of the findings: This research provides important data for phase transition in pears.
Additional comments: This MS entitled 'Genome-wide identification of the PEBP genes in pears and the putative role of PbFT in the flower bud differentiation' reported the isolation and the functional research of  PbFT in pear.  They found that PbFT might play a role in not only promoting flower bud differentiation, but also regulating the balance between vegetative and reproductive growth.
    It is qualified to be published in Peer J after minor revision:
    1 In the abstract, 'A trail of different treatments indicated that PbFT expressions in leaves were regulated by day-length and circadian clock, which was completely different from many other scholars’ findings' could be changed to ' PbFT expressions in leaves were firstly found to be regulated by day-length and circadian clock'.
    2 The format of the MS should be modified, there should be spaces  in the second paragragh of each parts.
    3 Discuss the possible role of PbFT in phase transition.


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 UNIFIED SIMULATION MODEL FOR UNDERSTANDING THE DIVERSITY OF CANCER EVOLUTION
Review round: 1
Reviewer: 1

Output:
Basic reporting: The article describes a simulation framework and software to model cancer evolution under different modes.

There are places where the English could be improved. More specifically, the introduction is fine, but in the Results section the tenses are inconsistent (e.g., switching from past to present in l. 83 and 84) and some sentences are misleading (e.g., l. 83 “each agent corresponds to each cell in a tumor”, it sounds like an agent represents several cells). There are also some formatting issues that need to be corrected. The citations are sometimes not well embedded in the text (e.g., l. 36, the author name should be between parentheses), which makes the reading more difficult. Please correct.

I find the introduction not sufficiently clear, lacking definitions of the basic phenomena described in the paper. In particular, the branching evolution model is never really defined. Also, the sentence l. 42 is misleading; “[...] intratumor heterogeneity (ITH) [...] results from branching evolution” makes it sound like ITH always results from branching evolution, which is in contradiction with the rest of the introduction, and the evolutionary literature.

The citations are insufficient (less than 30 citations is quite small for a methodological paper grounded on a rich literature from evolutionary biology and cancer). Firstly, I think citations are lacking to backup some statements. l. 37, the “linear evolution” hypothesis is stated without reference; I can suggest citing Nowell, P. C. The clonal evolution of tumor cell populations. Science 194, 23–28 (1976), but the authors might have another one in mind. Secondly, and more importantly, in general I think more credit must be given to early work or at the very least, reviews of such work. For example, regarding the usefulness of next generation sequencing l.40 to study cancer evolution, I think credit should be given to earlier studies or at least earlier reviews such as Yates, L. R., & Campbell, P. J. (2012). Evolution of the cancer genome. Nature Reviews Genetics, 13(11), 795. Also, because the punctuated equilibrium hypothesis dates from the 1970s, it sounds weird to only cite very recent work. Finally, I think more studies of mathematical and computational models of cancer evolution should be described, and the previous results should be discussed (e.g., what of Beerenwinkel, N., Schwarz, R. F., Gerstung, M., & Markowetz, F. (2014). Cancer evolution: mathematical models and computational inference. Systematic biology, 64(1), e1-e25.)

The figures are of good quality. I think the axes labels on Fig. 2 are currently barely readable and should be increased though. The sharing of all the code on github is commendable. I believe the papers follows very good standards in terms of reproducibility.

The article is self-contained.
Experimental design: In my opinion, the research topic is quite interesting and in the scope of the journal.
The research question is well defined. A gap in the current knowledge is clearly stated, but although I agree that there is indeed a need for a simulation software to tackle the issues mentioned in the introduction, I think that there is not enough references to the literature on mathematical modelling of cancer evolution to justify why we also need an additional mathematical model in addition to the ones already developed.

I have concerns regarding the methods. I do not agree with the fundamental assumptions of the model. In the driver model, each mutation increases the cell division rate. Although such mutations accelerating the rate of cell division probably exist, this is not what is commonly referred to as Darwinian selection in the evolutionary literature, where a positively selected mutation leads to more “offsprings” due to several phenomena, among which the most common is increased survival. An increased cell division rate indeed would lead to more offsprings per unit of time, and thus can be considered as a fitness increase and one possible type of drivers, but reducing all drivers to such a model is a very strong assumption that would need to be backed up either by very strong empirical data  in the cancer literature. For my part, I believe that this assumption fatally reduces the scope of the model; for example, it excludes resistance mechanisms against important therapeutic strategies such as immune checkpoint blockade, where driver events leading to the evolution of an immune escape phenotype actually increased survival of cells by reducing predation from the immune system, without ever acting on the cell division rate. Also, I think the assumption that most tumors undergo numerous events increasing the cell division rate needs to be justified.

I find that the results of the driver and neutral models do not add much to the literature, so emphasis on them should be reduced and proper literature should be cited (this basic effect of mutation and selection has been long described in population genetics, and can already be simulated forward in time for example using python package Simupop). The model with stem cell hierarchy is the true novelty of the model.

The punctuated evolution model is also problematic, as what it actually models (sudden changes in carrying capacity) do not correspond to what is classically understood as punctuated evolution.

I find that the sensitivity analysis is good but too qualitative. For example, l. 209-220, 6 qualitative behaviors are stated but without any comment on the range of parameters that lead to these behaviors (what is a small md, exactly?). Also, it ignores the issue of the transitions between these behaviors: what happens when f is intermediate? how sharp is the transition between the small and large f behaviors? I think this is especially important to describe because most of the behaviors described are expected and thus the input of the simulations proposed is to get actual quantitative measures.
Validity of the findings: Because of the concerns raised raised above, I do not think that the results are sufficiently sound. Indeed, statements like "under the assumption of sufficiently
high mutation rates, branching evolution occurs with strong driver mutations" are misleading, because "strong driver mutations" refer to the very specific model presented here, and thus is an extrapolation of the results. Also, apart from the results from the neutral-s model and the punctuated model, the results constitute a repetition of textbook population genetic results (e.g., the joint effect of mutation and selection are well-known), so I suggest again reducing the emphasis on them.
Additional comments: I think that the authors should revise the fundamental assumptions of their model to make it more general; as it stands, I think its description is misleading as it sounds very general while actually describing a very specific scenario, and probably not the most common. Also, although novelty is not a publication criterion here, more citations are needed to give credit to the abundant evolutionary literature and other attempts at modelling cancer evolution, and to justify the assumptions of the model.


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 UNIFIED SIMULATION MODEL FOR UNDERSTANDING THE DIVERSITY OF CANCER EVOLUTION
Review round: 1
Reviewer: 2

Output:
Basic reporting: Manuscript presents a driver model to study cancer evolution, mainly analysing clonality and branching structures of oncogenesis. The idea is well defined and executed. However, this strategy is not novel (as stated in the abstract), as models with the same basic rationale were already published (MInussi, Henz and Oliveira, Cancer Research, 2019). However, the technicalities of this model and the model presented in the present manuscript are different, which makes the present model worth publishing. It would be interesting if the key findings of this work could be confirmed by the previously published model.
In general the role of cell death is undermined, as in most tumours, cells proliferation coexists with cell death. In addition, immune surveillance also adds growth pressure, mainly mediated by cell death. Please discuss the role of cell death more clearly in all 4 models of cancer evolution.

The Figure models of evolutionary trees presented throughout the manuscript makes it very difficult to read. Consider other forms of depicting the profiles (Fig 1D), or at least make the lines of the clustering stronger. Additionally, identifying what would be considered a clone in these profiles would improve clarity of the figure.
Heat maps would be easier to visualize if the legends instead of symbols had also a description of their meanings.

Minor: There are some typos, especially in the abstract.
Experimental design: no comment
Validity of the findings: Findings are important, but are limited by the limited data available of cancer evolution. This limitations are, however, clearly stated in the discussion.
Additional comments: no comment


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

Input:
Title: A UNIFIED SIMULATION MODEL FOR UNDERSTANDING THE DIVERSITY OF CANCER EVOLUTION
Review round: 2
Reviewer: 1

Output:
Basic reporting: I am very pleased with the additional references and improvements in English in this revised version.
Experimental design: I am pleased with the clarifications of the research question and models, and the addition of new models.
Validity of the findings: I find the additional models, and the additional information about hypotheses of the model satisfactory.
Additional comments: I find the manuscript much improved. The efforts of the authors to rethink the manuscript and models are very welcome. The framework developed by the authors could be quite useful to explore evolutionary scenarios.


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: MONITORING THE INFLUX OF NEW SPECIES THROUGH CITIZEN SCIENCE: THE FIRST INTRODUCED ANT IN DENMARK
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript is well written and other than a very few instances where sentence structure seems a bit ambiguous (and they are noted in the specific comments), the English is excellent.

The manuscript examines:
	-the use of citizen collectors to more broadly sample the environment for ants and compares those efforts to those of biologists.
	-the appearance of a new species (T. immigrans) to Denmark and the role of citizen science in making that discovery
	-the habitat and climatic niche differences between two species of Tetramorium in Europe.

The literature is well and appropriately referenced.

The structure is professional.

The Figures and raw data seem appropriate.  I note in general comments that the quality of Figure 3 b was not great in my copy. I also note that I am not sure how Figure 1 b contributes to the manuscript.  It is a fascinating and technically interesting image, but does not seem to advance the manuscript intent.

As a general comment, I did find (and this could arise from my initial quick reading) that the distinction between Danish (essentially presence/absence data for T. immigrans as it was only in one location) and the broader European dataset was not clear, at least for me. It wasn’t until I read that T. immigrans was found in only one location in Denmark that Denmark wasn’t really contributing to the overall niche analysis for the species. If this single discovery was noted earlier and clearly, it would be easier to understand the limited role of the Denmark T. immigrans data in the European analysis.
Experimental design: The manuscript meets my reading of the ‘Aims and Scope’ of Peer J.

The research questions are clear (although note my last comment under Basic Reporting).

The technical aspects are excellent.

With respect to the ability to replicate the research, I would ask for more details about the protocol given to students for collections. It might be also noted how the schools were approached to participate.
Validity of the findings: The results arise from a well designed methods. I would only mention the need to mention earlier the single finding of T. immigrans in Denmark.

Conclusions appropriate to the results
Additional comments: Some editorial comments

Line 23: Do you mean,  “Here we report on the discovery of Tetramorium immigrans, the first established exotic ant taxon in Denmark through The Ant Hunt,” or ‘Here we report on the discovery of Tetramorium immigrans, the first exotic ant taxon in Denmark established through The Ant Hunt.’

Line 65: What about the Hypoponera?

Line 86 (Biological data).
-It would be useful to know the age range of students participating
-It would be useful to know what six foods were used on the bait cards (I see this in Table 1 but this is likely to be placed in ‘Results’ which is distant from where the protocol in introduced)
-Is this project only ant focused or part of a larger citizen science project
-Is this protocol part of an established monitoring protocol. If so, what protocol (reference?)

Line 110 (Paragraph): I am not sure of the purpose of this imaging as only one specimen was used. Imaging of multiple specimens of the two cryptic species would seem of use for taxonomists, but I am uncertain if the one image is of value and should be included.

Line 122: Did you have students directly place specimens into 96% ethanol?

Line 147: you don’t explicitly mention the use of PCA in these methods but provide that in Figure 3

Line 154: some statement regarding confidence in correct IDs might help here given the noted difficulties in distinguishing between the two species.

Line 170-175:  I will just note that while many editors do not like explanations of methodological analyses, many readers find them quite useful.

Para 240 and 245:  First paragraph (Line 240) notes you can distinguish differences in land cover types which suggests those cover types are about to be noted (but do not seem to be). The next paragraph (Line 245), states “The two species also ….” which suggests you are moving onto different aspects of land cover.  Am I missing something?

Line 238: “…more than expected.”  More than expected from what? Random sampling?

Table 1 title:  Would the term ‘sampling block’ be better than ‘experiment?’ I suggest ‘block’ given that a variety of foods were used. The term ‘sampling block’ may not be quite right, but experiment suggests a protocol with a control.

Table 1: It would be good to indicate which other species are known exotics (e.g., Hypoponera).
Figure 2: “b) 28S rDNA alignment showing a characteristic insertion of T. immigrans …”
Might be better as
“b) 28S rDNA alignment (T.immigrans) showing a characteristic insertion of (C) …”

Figure 3: the quality of Figure 3 b is not very good in the copy I have received.

Figure 3: the meaning of the shading in c and d are not noted in the title


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: MONITORING THE INFLUX OF NEW SPECIES THROUGH CITIZEN SCIENCE: THE FIRST INTRODUCED ANT IN DENMARK
Review round: 1
Reviewer: 2

Output:
Basic reporting: I put all my comments for the editor and the authors under "General comments for the author".

Just briefly:
English is great.
article structure, figures, tables are fine (same small mistakes like missing spaces in legends).
some literature missing (details see below).
Raw data: morphometric data of Tetramorium immigrans workers from Denmark should be shared!
Experimental design: I put all my comments for the editor and the authors under "General comments for the author".

Just briefly:
Original primary research within Aims and Scope of the journal: yes.
Research question well defined, relevant & meaningful: yes.
Methods well described.
Validity of the findings: I put all my comments for the editor and the authors under "General comments for the author".

Just briefly:
The huge difference of the climate model and the real distribution of Tetramorium immigrans is a problem. Maybe the model should be made new to be meaningful (details below).

The study has the potential to show one important novelty better: an overview where in Europe Tetramorium immigrans might be native and where it is introduced.
Additional comments: Dear Julie K. Sheard et al.,

It is very interesting to investigate the invasive potential of Tetramorium immigrans. There are several other authors who already speculated about that aspect (as you have correctly cited in the manuscript).

However, I think there are some aspects in the manuscript which should be improved.

1)	You have written: “… may not be native to all of Europe and is being accidentally introduced by humans“
In this aspect I see the chance to show something very interesting new. It would be great to give an overview where in Europe it might be native and where it is introduced. For the situation in Austria, for example, T. immigrans occurs in natural and seminatural habitats around Vienna (or in the east and south of Vienna), but not in natural habitats in Styria (around Graz), where it is only semisynanthropic (WAGNER 2011). The situation is similar around Lyon (France) or Poland, where it might be a non-native (GIPPET & al. 2017, BOROWIEC & SALATA 2018, CORDONNIER & al. 2019). I would guess that T. immigrans is native at least for the Balkans and the Pannonia. It is a mentioned to be a Neozoon for Germany (SEIFERT 2018). Maybe based on published habitat information (e.g., WAGNER & al. 2017) you can work out some information about its native range. You should mention somewhere that the highest COI variability is in Anatolia and Caucausus region (WAGNER & al. 2017). Thus, most likely there is its origin.

2)	You have written: “However, an even more recent study demonstrated that T. immigrans and T. caespitum widely share the same gene pool when occurring in sympatry (Cordonnier et al., 2019), challenging this idea.”
Hybridization does not challenging species delimitation between T. immigrans and T. caespitum! They are good species. Please take a look into papers of modern species concepts (DE QUEIROZ 2007, SEIFERT 2014) to consider that hybridization is allowed between species and Mayr’s old concept (MAYR 1942) is just an idea far away from species reality.

3)	It is often underestimated how difficult the identification of Tetramorium is. The lack of morphometric data of the T. immigrans nest in Denmark is a weakness and with your data we cannot fully exclude the possibility of a hybrid. The combination of COI and 28S is enough to separate pure caespitum from pure immigrans. But the question regarding a putative hybrid nest is still open and thus an alternative hypothesis to the occurrence of immigrans. Interspecific hybridization rates are higher in anthropogenic influenced habitats (SEIFERT 1999), where T. immigrans in Western/Central(/Northern) Europe often occurs. Especially in an area so far in north, where immigrans might be very rare (in relation to caespitum), the force do hybridize interspecifically might be high for an immigrans gyne. What I see on https://vimeo.com/user86430849 shows me that with POTCos of around 11 (only unilateral) probability of dealing with T. immigrans and not with T. caespitum is higher (but I cannot fully exclude hybrids). At the moment nobody is fully able to identify such hybrids (the Kaufmann group did that genetically). Seifert did that a few times morphometrically or tried to do that, but he mentioned that hybrids are similar to T. staerckei (which would not consider a problem in this study, since nobody would suggest that non-invasive T. staerckei occurs in Denmark). You should have used the key of WAGNER et al. (2017) and/or SEIFERT (2018) in addition to genetics, which demands morphometric data. Put all these morphometric data into the publication. If you are dealing with a hybrid, maybe future scientists will be able to proof that by your published morphometric data.

4)	“Niche overlap was calculated using Schoener’s overlap metric…“
Whenever you write about niche overlap in ants you have to cite Seifert’s monograph of ant ecology, which also deals with the calculation of ecological niches in ants (SEIFERT 2017).

5)	You should mention that the TAS (mean temperature from May to August) was before published for all 10 European species of the caespitum complex (WAGNER & al. 2017, Tab. 4) and the values for immigrans and caespitum are very similar with your “Mean temp. of warmest quarter”.

6)	The fact that the ant material was collected via baiting by schools and not via hand-collecting from nests by myrmecologists is not perfect for the scientific aspect of your paper. You could have individuals from different nests in the same sample. Usually you should work with nest samples in Tetramorium.

7)	“… making citizen science an excellent tool for exotic species monitoring”
I do not agree with that. Citizens might be useful to detect birds etc., but myrmecology needs accuracy. I rather consider it as problematic that citizens will, for example, confuse samples (which is also a big problem for myrmecologists). So I suggest toning down this message and tell the reader about possible risks. By the way: concerning several details about citizen scientists at all: I don’t think that these things are worth to be published that detailed in PeerJ (but that should be decided by the editor and not by me).

8)	I am not expert for climate niche modeling; however, the suggested niche of T. immigrans in Fig. 3 d) does not convince me. According this niche, eastern Germany must be THE hot spot of T. immigrans. However, not a single record is known from this well-investigated area. This “hot spot” might be an artifact based on the use of urban records of T. immigrans. In cities the temperature is higher than in the surroundings. Shouldn’t such records be omitted to model a climate niche to mean anything? On the other hand, Mediterranean areas in, for example, Greece or Croatia (were T. immigrans is very common at the coast) are white in your map, which does not reflecting the reality.
In that content, you only have written: “It has already been hypothesized that T. immigrans may not be native in most of Europe (Gippet et al., 2017; Borowiec & Salata, 2018; Cordonnier et al., 2019). If true, its’ current observed distributional focus in Southern Europe could reflect invasion history rather than climatic preference. This would explain why T. immigrans is presently less widespread in northern Europe than environmental niche modelling would predict (Fig. 3d).”
I think this is not enough to discuss to huge difference between the model and the real distribution.

And a few small things:

1)	Line 99: “T. albipes“: use full genus name.

2)	“T. caespitum/impurum complex“ is the old name for the complex, you should use the new name “T. caespitum complex”.

3)	Fig. 2a: “lineage E“ not in italics. And what does “~” mean?

4)	Fig. 2b is very interesting!!

5)	Go through the whole manuscript for detecting sloppinesses like missing spaces, missing full stops etc. Here an example: Table 1: You must never write “sp.“ in italics.

6)	Legend of Tab. 2: Tell the reader also in the legend from which source you have taken this data.

I hope you consider my critics as constructive and that you will improve the manuscript.

Literature:
BOROWIEC, L. & SALATA, S. 2018: Tetramorium immigrans Santschi, 1927 (Hymenoptera: Formicidae) nowy gatunek potencjalnie inwazyjnej mrówki w Polsce. – Acta entomologica silesiana 26: 1–5.
CORDONNIER, M., GAYET, T., ESCARGUEL, G. & KAUFMANN, B. 2019: From hybridization to introgression between two closely related sympatric ant species. – Journal of Zoological Systematics and Evolutionary Research.
DE QUEIROZ, K. 2007: Species concepts and species delimitation. – Systematic Biology 56: 879-886.
GIPPET, J.M.W., MONDY, N., DIALLO-DUDEK, J., BELLEC, A., DUMET, A., MISTLER, L. & KAUFMANN, B. 2017: I’m not like everybody else: urbanization factors shaping spatial distribution of native and invasive ants are species-specific. – Urban Ecosystems 20: 157-169.
MAYR, E. 1942: Systematics and the origin of species from the viewpoint of a zoologist. – Harvard University Press, Cambridge, 334 pp.
SEIFERT, B. 2014: A pragmatic species concept applicable to all eukaryotic organisms independent from their mode of reproduction or evolutionary history. – Soil Organisms 86: 85-93.
SEIFERT, B. 1999: Interspecific hybridisations in natural populations of ants by example of a regional fauna (Hymenoptera, Formicidae). – Insectes Sociaux 46: 45-52.
SEIFERT, B. 2018: The ants of Central and North Europe. – lutra Verlags- und Vertriebsgesellschaft, Tauer, 408 pp.
SEIFERT, B. 2017: The ecology of Central European non-arboreal ants – 37 years of a broad-spectrum analysis under permanent taxonomic control. – Soil Organisms 89: 1-67.
WAGNER, H.C. 2011: Tag der Artenvielfalt – Die Ameisen (Hymenoptera: Formicidae) im Botanischen Garten Graz. – Mitteilungen des naturwissenschaftlichen Vereines für Steiermark 141: 235–240.
WAGNER, H.C., ARTHOFER, W., SEIFERT, B., MUSTER, C., STEINER, F.M. & SCHLICK-STEINER, B.C. 2017: Light at the end of the tunnel: integrative taxonomy delimits cryptic species in the Tetramorium caespitum complex (Hymenoptera: Formicidae). – Myrmecological News 25: 95-130.


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: MONITORING THE INFLUX OF NEW SPECIES THROUGH CITIZEN SCIENCE: THE FIRST INTRODUCED ANT IN DENMARK
Review round: 1
Reviewer: 3

Output:
Basic reporting: Presented research is supported by solid analyses, and I strongly recommend its publication. The manuscript is written in clear and professional English. However, the introduction is, in some places, inconsistent and misleading. Its greater part deals with introduced and invasive species. But the Authors didn't point out differences between those two groups. It is also not clear to which of those group Tetramorium immigrans is assigned, and why? In further parts of the ms, the Authors started also using the term “exotic.” Is it possible to explain the relationship between those three terms? In my opinion, “exotic” usually refers to species originated from the biogeographic realm different from the one in which it was collected. All evidence suggests that Tetramorium immigrans is a Palearctic species, thus describing it as “exotic” can be confusing.
Additionally, information referring to the species is, in my opinion, very scarce, and it lacks background data of distribution and biology of the whole caespitum group (Wagner et al., 2017 published comprehensive revision of the entire group). The Authors stated that Tetramorium immigrans “is thought to have a more southern and urban distribution than T. caespitum,” but they failed to describe the distribution range of T. caespitum. I think that, in this case, distribution ranges of both taxa should be discussed in detail to show discussed differences.
Lines 100-103 very clearly point out differences between T. immigrans and T. caespitum. However, the mentioned characters are not visible on photographs 1a and 1b. Especially photograph 1a is blurry and indistinct. To visualize discussed differences, the Authors should also include photographs of T. caespitum.
Experimental design: In lines 148-149, the Authors stated that the geographical distribution of T. immigrans and T. caespitum in Europe was implemented from AntMaps. In my opinion, this approach is quite risky. On the cited website is stated that “There are errors and uncertainties that are inherited from the data sources and others that might have been introduced during the databasing process. Distributional data in GABI should be viewed critically before forming the basis of scientific investigation.” Did the Authors check if used records have been published or verified by experts? If so, they should cite the original sources of those records.
Line 308: the statement that the “species may not be native to all of Europe” needs clarification. What does it mean? If not to all, then which part can be considered as native, and why?
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: MONITORING THE INFLUX OF NEW SPECIES THROUGH CITIZEN SCIENCE: THE FIRST INTRODUCED ANT IN DENMARK
Review round: 2
Reviewer: 1

Output:
Basic reporting: The English is better than mine.

Literature references are sufficient.

Ad raw data: It should be written in the manuscript that traditional morphometric data of the T. immigrans workers in Denmark have NOT been collected. Based on that, every reader can decide by himself if he or she trusts in the identification. By the way, all in all, for me the identification is convincing. (I do not know if anybody is able to transfer a CT-scan video into absolute traditional morphometric values. So that video probably does not help a lot.)

Ad figures: I have to complain that I am still not very happy with the climate niche model of T. immigrans (Fig. 3d), although the authors have mentioned the aspects of city data (which is welcome!). I have mentioned the problem with eastern Germany in the first review just as one unlucky example. But the model is also inappropriate for other parts of Europe. What is about England, for example? Phil Attewell is interested in finding new English Tetramorium species for years, but he has not found T. immigrans. According this model (Fig. 3d), eastern England shoud be and ideal area for the species. But, empirically, that is not the case, probably because of climatic reasons, temperature and/or precipitation. I have no good idea how to improve the problematic aspect that the model does not fit with the realitiy. The authors should think once again if they made any mistake in the data analyses and how they can explain such unexpected results. Is the city aspect enough?
Experimental design: Original primary research: Yes.

Research question well defined, relevant & meaningful: Yes.

Rigorous investigation performed to a high technical & ethical standard: Unfortunately, I consider one big problem which I have not recognized during the first review (and which has been already considered by Rev. 3): The Tetramorium caespitum data of antmaps.org are partly not convincing and probably in parts wrong. I do not believe that Tetramorium caespitum occurs on southern Greek islands as well as in some low-altitude parts of southern Spain near the coast. This records are most likely identification errors and the data should not be used for the analyses. Who did the determination of these samples? Usually, Tetramorium caespitum is replaced by other species in the very south of Europe at low altitudes. The authors should think more critical about the published data they use and omit those which are not convincing and maybe do the analyses new.
Validity of the findings: All fine (but see 1. and 2.).
Additional comments: Dear Julie K. Sheard et al.,

Thank you for your detailed answers to all my comments! I consider that you thought about them all. I like your answers and many of them convince me.

I am sorry that I still see some problems in the current version of the manuscript and I hope you can rather solve the problems or convince me or the editor that the problems do not exist.

Here some small aspects:

Line 59: One „.“ is missing.

Line 83: Shouldn’t there be a comma after „Anatolia“? As I have heard a comma before „and“ shoud always be there in a list of three or more words.

Line 168: Put „immigrans“ in lower case.

Line 308: I do not understand the followig sentence: „This would also explain the discrepancies between its’ current distribution and modelled distribution based on its climatic niche, which suggests the species should be present in eastern Germany (Fig. 3d).“ I do not understand the explanation, what is „This“?

If you are interested in my record of Tetramorium immigrans in the Botanical Garden of Graz, look here:
https://www.zobodat.at/pdf/MittNatVerSt_141_0235-0240.pdf
I called the species "hemisynanthropic".

Take care and best wishes,
Heri


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: URBAN–RURAL DIFFERENCES IN PERCEPTION OF TREES DESCRIBED BY PARENTS BRINGING UP CHILDREN IN WARSAW AND JEDLIŃSK, POLAND
Review round: 1
Reviewer: 1

Output:
Basic reporting: The article is in general written in clear, professional English but please check these two sentences:

Page 4, line 74-75, sentence beginning with "Trees conflict with the road infrastructure is one of the main reasons..." Is there something wrong with this sentence? My (non-native) ear says that it ought to be either "Trees' conflict" or "Trees conflicting". Please check this.

Page 12, line 293, sentence "It is evident that city dwellers are more inclined to say that the cost of maintaining the trees exceeds their benefits." Is this in conflict with the findings? Should it be "city dwellers are less inclined"? Please check this.

The discussion is somewhat unusually long and includes quite a lot of new literature. Some of this literature might have been included already in the introduction, but it is not necessary to make any changes.
Experimental design: This is a well designed study. My only comment is that I wish to see a reason for choosing these respondents in chapter 2.1. Why were parents of primary school children the target group, and why were these two schools picked?
Validity of the findings: The conclusions are otherwise well stated, but the word "much" on line 547 (city respondents much more often agreed) seems exaggerated compared to the findings. After all, the main conclusion is that there was no great difference between the two groups.
Additional comments: Thank you for the chance of reading this clear and interesting study.


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: URBAN–RURAL DIFFERENCES IN PERCEPTION OF TREES DESCRIBED BY PARENTS BRINGING UP CHILDREN IN WARSAW AND JEDLIŃSK, POLAND
Review round: 1
Reviewer: 2

Output:
Basic reporting: In my opinion, the manuscript would benefit from proofreading by a native speaker. Your Introduction is well written and comprehensive. However, in my opinion, you should update your references (some research you cite is from the 80s and more recently published papers are available). For example:
•	Trees reduce urban heat stress:
Potchter, O., Cohen, P., Bitan, A., 2006. Climatic behavior of various urban parks during hot and humid summer in the Mediterranean City of Tel Aviv, Israel. Int. J. Climatol. 26, 1695–1711, http://dx.doi.org/10.1002/joc
Bowler, D.E., Buyung-Ali, L., Knight, T.M., Pullin, A.S., 2010. Urban greening to cool towns and cities: a systematic review of the empirical evidence. Landsc. Urban Plan. 97 (3), 147–155, http://dx.doi.org/10.1016/j.landurbplan.2010.05.006
Shashua-Bar, L., Potchter, O., Bitan, A., Yaakov, Y., 2010. Microclimate modelling of street tree species effects within the varied urban morphology in the Mediterranean city of Tel Aviv, Israel. Int. J. Climatol. 30, 44–57, http://dx.doi.org/10.1002/joc
McCarthy, M.P., Best, M.J., Betts, R.A., 2010. Climate change in cities due to global warming and urban effects. Geophys. Res. Lett. 37, 1–5, http://dx.doi.org/10. 1029/2010GL042845
•	Trees reduces building energy use:
Akbari, H., Pomerantz, M., Taha, H., 2001. Cool surfaces and shade trees to reduce energy use and improve air quality in urban areas. Sol. Energy 70 (3), 295–310, http://dx.doi.org/10.1016/S0038-092X(00)00089-X
Nowak, D.J., Dwyer, J.F., 2007. Understanding the benefits and costs of urban forest ecosystems. In: Kuser, J.E. (Ed.), Urban and Community Forestry in the Northeast (2nd). Springer Netherlands, Dordrecht, pp. 25–46, http://dx.doi.org/10.1007/978-1-4020-4289-82
Escobedo, F.J., Kroeger, T., Wagner, J.E., 2011. Urban forests and pollution mitigation: analyzing ecosystem services and disservices. Environ. Pollut. 159 (8–9), 2078–2087, http://dx.doi.org/10.1016/j.envpol.2011.01.010
•	Trees reduce atmospheric carbon dioxide:
Nowak, D.J., Dwyer, J.F., 2007. Understanding the benefits and costs of urban forest ecosystems. In: Kuser, J.E. (Ed.), Urban and Community Forestry in the Northeast (2nd). Springer Netherlands, Dordrecht, pp. 25–46, http://dx.doi.org/10.1007/978-1-4020-4289-82
Escobedo, F.J., Kroeger, T., Wagner, J.E., 2011. Urban forests and pollution mitigation: analyzing ecosystem services and disservices. Environ. Pollut. 159 (8–9), 2078–2087, http://dx.doi.org/10.1016/j.envpol.2011.01.010
•	Trees reduce rainfall runoff, flooding, noise levels, and wind speed:
Tyrväinen, L., Pauleit, S., Seeland, K., de Vries, S., 2005. Benefits and uses of urban forests and trees. In: Konijnendijk, C.C., Nilsson, K., Randrup, T.B., Schipperijn, J. (Eds.), Urban Forests and Trees. Springer, Berlin, pp. 81–114.
Nowak, D.J., Dwyer, J.F., 2007. Understanding the benefits and costs of urban forest ecosystems. In: Kuser, J.E. (Ed.), Urban and Community Forestry in the Northeast (2nd). Springer Netherlands, Dordrecht, pp. 25–46, http://dx.doi.org/10.1007/978-1-4020-4289-82
•	Trees remove air pollutants:
Nowak, D.J., Hirabayashi, S., Greenfield, E., 2014. Tree and forest effects on air quality and human health in the United States. Environ. Pollut. 193, 119–129, http://dx.doi.org/10.1016/j.envpol.2014.05.028
Nowak, D.J., Dwyer, J.F., 2007. Understanding the benefits and costs of urban forest ecosystems. In: Kuser, J.E. (Ed.), Urban and Community Forestry in the Northeast (2nd). Springer Netherlands, Dordrecht, pp. 25–46, http://dx.doi.org/10.1007/978-1-4020-4289-82
Escobedo, F.J., Kroeger, T., Wagner, J.E., 2011. Urban forests and pollution mitigation: analyzing ecosystem services and disservices. Environ. Pollut. 159 (8–9), 2078–2087, http://dx.doi.org/10.1016/j.envpol.2011.01.010
•	Aesthetic value of trees:
Lohr, V.I., Pearson-Mims, C.H., 2006. Responses to scenes with spreading, rounded, and conical tree forms. Environ. Behav. 38 (5), 667–688, http://dx.doi.org/10.1177/0013916506287355
Schroeder, H.W., Flannigan, J., Coles, R., 2006. Residents’ attitudes toward street trees in the UK and U.S. communities. Arboricult. Urban For. 32 (5), 236–246.
•	Relief from stress when being surrounded by nature:
Jiang, B., Chang, C.-Y., Sullivan, W.C., 2014a. A dose of nature: tree cover, stress reduction, and gender differences. Landsc. Urban Plan. 132, 26–36, http://dx.doi.org/10.1016/j.landurbplan.2014.08.005
Jiang, B., Li, D., Larsen, L., Sullivan, W.C., 2014b. A dose–response curve describing the relationship between urban tree cover density and self-reported stress recovery. Environ. Behav., 1–23, http://dx.doi.org/10.1177/0013916514552321
Van den Berg, A.E., Jorgensen, A., Wilson, E.R., 2014. Evaluating restoration in urban green spaces: does setting type make a difference? Landsc. Urban Plan. 127, 173–181, http://dx.doi.org/10.1016/j.landurbplan.2014.04.012
•	The presence of nearby nature increases property values:
Wolch, J. R., Byrne, J., & Newell, J. P. (2014). Urban green space, public health, and environmental justice: The challenge of making cities ’just green enough’. Landscape and Urban Planning, 125, 234–244, http://dx.doi/10.1016/j.landurbplan.2014.01.017
•	Trees contribute to urban neighbourhoods’ aesthetic quality and enhance human mental and physical health and well-being:
Tyrväinen, L., Pauleit, S., Seeland, K., de Vries, S., 2005. Benefits and uses of urban forests and trees. In: Konijnendijk, C.C., Nilsson, K., Randrup, T.B., Schipperijn, J. (Eds.), Urban Forests and Trees. Springer, Berlin, pp. 81–114.
Nilsson, K., Sangster, M., Konijnendijk, C.C., 2011. Forests trees and human health and well-being: introduction. In: Nilsson, K., Sangster, M., Gallis, C., Hartig, T., de Vries, S., Seeland, K., Schipperijn, J. (Eds.), Forests, Trees and Human Health. Springer, Dordrecht, pp. 1–19.
De Vries, S., Verheij, R.A., Groenewegen, P.P., Spreeuwenberg, P., 2003. Natural environments – healthy environments? An exploratory analysis of the relationship between greenspace and health. Environ. Plan. 35 (10), 1717–1731, http://dx.doi.org/10.1068/a35111
Maas, J., Verheij, R.A., Groenewegen, P.P., de Vries, S., Spreeuwenberg, P., 2006. Green space, urbanity, and health: how strong is the relation? J. Epidemiol. Community Health 60 (7), 587–592, http://dx.doi.org/10.1136/jech.2005.043125
Maas, J., Verheij, R.A., de Vries, S., Spreeuwenberg, P., Schellevis, F.G., Groenewegen, P.P., 2009. Morbidity is related to a green living environment. J. Epidemiol. Community Health 63 (12), 967–973, http://dx.doi.org/10.1136/jech.2008.079038
Mitchell, R., Popham, F., 2007. Greenspace, urbanity and health: Relationships in England. J. Epidemiol. Community Health 61 (8), 681–683, http://dx.doi.org/10.1136/jech.2006.053553
Taylor, M.S., Wheeler, B.W., White, M.P., Economou, T., Osborne, N.J., 2015. Research note: urban street tree density and antidepressant prescription rates – a cross-sectional study in London, UK. Landsc. Urban Plan. 136, 174–179, http://dx.doi.org/10.1016/j.landurbplan.2014.12.005
Donovan, G.H., Michael, Y.L., Butry, D.T., Sullivan, A.D., Chase, J.M., 2011. Urban trees and the risk of poor birth outcomes. Health Place 17 (1), 390–393, http://dx.doi.org/10.1016/j.healthplace.2010.11.004
Hansmann, R., Hug, S.-M., Seeland, K., 2007. Restoration and stress relief through physical activities in forests and parks. Urban For. Urban Green. 6 (4), 213–225, http://dx.doi.org/10.1016/j.ufug.2007.08.004
Korpela, K., Borodulin, K., Neuvonen, M., Paronen, O., Tyrväinen, L., 2014. Analyzing the mediators between nature-based outdoor recreation and emotional well-being. J. Environ. Psychol. 37, 1–7, http://dx.doi.org/10.1016/j.jenvp.2013. 11.003
Berman, M.G., Jonides, J., Kaplan, S., 2008. The cognitive benefits of interacting with nature. Psychol. Sci. 19 (12), 1207–1212, http://dx.doi.org/10.1111/j.1467-9280.2008.02225.x
Tyrväinen, L., Ojala, A., Korpela, K., Lanki, T., Tsunetsugu, Y., Kagawa, T., 2014. The influence of urban green environments on stress relief measures: a field experiment. J. Environ. Psychol. 38, 1–9, http://dx.doi.org/10.1016/j.jenvp.2013.12.005
•	Trees and nature are associated with social cohesion:
Holtan, M.T., Dieterlen, S.L., Sullivan, W.C., 2014. Social life under cover: tree canopy and social capital in Baltimore, Maryland. Environ. Behav. 46 (6), 1–24, http://dx.doi.org/10.1177/0013916513518064
•	Review articles on positive effects of nearby urban forest resources on human health and well-being:
Keniger, L.E., Gaston, K.J., Irvine, K.N., Fuller, R.A., 2013. What are the benefits of interacting with nature? Int. J. Environ. Res. Public Health 10 (3), 913–935, http://dx.doi.org/10.3390/ijerph10030913
Turner‐Skoff, J. B., & Cavender, N. (2019). The benefits of trees for livable and sustainable communities. Plants, People, Planet. https://doi.org/10.1002/ppp3.39
Kondo, M. C., Fluehr, J. M., McKeon, T., & Branas, C. C. (2018). Urban green space and its impact on human health. International Journal of Environmental Research and Public Health, 15(3). https://doi.org/10.3390/ijerph15030445
Nilsson, K., Bentsen, P., Grahn, P., & Mygind, L. (2019). De quelles preuves scientifiques disposons-nous concernant les effets des forêts et des arbres sur la santé et le bien-être humains? Santé Publique, HS(S1), 219–240. https://www.cairn.info/revue-sante-publique-2019-HS-page-219.htm

I suggest restructuring the manuscript: E.g., large parts of the discussion should be part of the results in terms of a categorisation of previous findings (see my recommendations that I uploaded in the attachment).

In my opinion, more work is needed to make the manuscript self-contained (see my uploaded file).
Experimental design: Please see my comments in the file I uploaded. In my opinion, more work is needed with respect to defining the research question and describing why this knowledge is important.
Validity of the findings: In my opinion, the conclusions could be stated more clearly (please see the file I uploaded).
Additional comments: I like your research. However, in my opinion, a lot more work is needed to make the most out of it (see my comments in the attached file).


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: URBAN–RURAL DIFFERENCES IN PERCEPTION OF TREES DESCRIBED BY PARENTS BRINGING UP CHILDREN IN WARSAW AND JEDLIŃSK, POLAND
Review round: 2
Reviewer: 1

Output:
Basic reporting: The structure and language have been improved by the changes made. Basic reporting is OK as such.
Experimental design: Questions concerning the data and methods have been clarified and are now presented in an acceptable manner.
Validity of the findings: The discussion relates the study to previous research and gives depth to the findings. Conclusions are clear and such that can be drawn from the results.
Additional comments: I find the article greatly improved. Well done!