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: WIN 55,212-2 SHOWS ANTI-INFLAMMATORY AND SURVIVAL PROPERTIES IN HUMAN IPSC-DERIVED CARDIOMYOCYTES INFECTED WITH SARS-COV-2
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript is well written and structured, but the authors would be appreciated if they clarified the following:

1.	Suggested title: WIN55,212-2 shows anti-inflammatory and survival properties in human…

2.	Line 85 : …elevated levels of creatine kinase and lactate dehydrogenase activity (LDH), which are injury biomarkers... To which organ/system?

3.	Line 88-90: The citations do not appear to support the claim of correlation between high expression of ACE2 and patients susceptibility to severe COVID 19.

4.	Line 106-108: Studies have been investigating… but only one example is given. Citations needed.
5.	Line 110-113: High affinity of WIN to CB receptors does not necessarily imply high functional activity or efficacy. Author should consider citation of functional data of WIN

6.	Line 113-122: The author has given good examples to support anti-inflammation by WIN, but these activities were not CB-mediated. Author should include studies such as PMID: 10734163, which are examples of CB mediated anti-inflammation. A statement highlighting that the anti-inflammatory activity of WIN may or may not be CB mediated will improve this section.
7.	Line 124-125: Citation needed.
8.	Line 170: Full meaning of MOI at first use.
9.	Line 275-276: Same as line 110. Reference to functional data will be more suitable.
Experimental design: 10.	Line 277: There is significant evidence that CB2 antibodies have poor selectivity and/or specificity (Atwood, Straiker, & Mackie (2012), Cécyre et al. (2013), Marchalant et al. (2014). I) Did you use a robust negative control (e.g. undifferentiated iPSC) in your western blot? Ii)How many biological replicates did you undertake? Iii) Did you validate the CB2 antibody? V) Could you please provide densitometry for CB receptor expression? Iv) Please include a figure legend in your supplementary data.

11.	Line 280: I may be mistaken, but Marchalant, Rosi & Wenk, 2007 used immunohistochemistry (IHC) to study CB1 expression. How do you compare the banding pattern of WB to IHC? Secondly, please cite the original article; Atwood & Mackie, 2010 is a review.

12.	There are several pro-inflammatory cytokines. What was the rationale for choosing these three?
Validity of the findings: The studies are robust except for the data which support the expression of CB receptors. The relevant clarifications have been raise in the experimental section above.
Additional comments: 13.	Line 328: …which is a hallmark for damage. Damage to what? Cell damage?

14.	Line 334: Discussion can be improved by elaborating upon the CB1 and CB2 expression on hiPSC-CMs and comparing to other literature. There is evidence that both receptors are expressed in murine embryonic stem cells (Jiang et al. 2007), but perhaps there is little information on hiPSC-CMs

15.	Line 368-369: If your data shows CB receptor expression. How does the absence of CB2 receptors explain the inactivity of WIN?

16.	Line 393-400: THC and WIN are CB1 and CB2 agonists, but their efficacy is significantly different (Soethoudt et al. 2017). So, they will not necessarily produce a similar effect through the cannabinoid receptors.

17.	Line 395-400: CB1 and CB2 receptors mediate cardioprotective properties (Liao et al. 2013; Batkai et al. 2007; Dueer et al. 2014; Netherland et al. 2010). Therefore cardioprotection by THC in neonatal murine cardiomyocytes could have been mediated through CB2 because CB1 receptors were not expressed.  Although the authors subsequently mention CB1,  I’ll suggest the inference in lines 399-400 should mention both receptors instead of highlighting CB2 receptors only.


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: WIN 55,212-2 SHOWS ANTI-INFLAMMATORY AND SURVIVAL PROPERTIES IN HUMAN IPSC-DERIVED CARDIOMYOCYTES INFECTED WITH SARS-COV-2
Review round: 1
Reviewer: 2

Output:
Basic reporting: I commend the authors for undertaking this interesting study on investigating the anti-inflammatory potential of synthetic cannabinoid, WIN55212-2, in human iPSC-derived cardiomyocytes (hiPSC-CMs) infected with SARS-CoV-2. The authors of the present manuscript addressed this issue by examining the in vitro effect of WIN on modulating the expression of ACE2; cellular staining and confocal microscopy to study the effect of WIN on SARS-CoV-2 infection and replication; and the effect of WIN in modulating the release of several pro-inflammatory cytokines - IL-6, IL-8, and TNF-α associated with the disease.

In general, the manuscript should have been read carefully before submission. The paper is generally well written and structured. However, in my opinion, the paper has some shortcomings in regard to sample size (n=3) and text, and I feel this unique dataset has not been utilized to its full extent. I have provided numerous remarks on texts, specific examples are given below.
Experimental design: The experimental design is appropriately described. The impetus for this series of studies is also defined in the Introduction. Although the sample size is low, I am unsure if enough sample size is considered for this study to be repeatable.
Validity of the findings: Based on what is provided, the data appears to be robust. That said, while the results are noteworthy, the study is somewhat descriptive, and additional investigation of the mechanism underlying the anti-inflammatory effect of WIN in cells infected with SARS-CoV-2 (involvement of CB1 or CB2) would greatly improve the rigor, reproducibility, and utility of this manuscript.
Additional comments: Line 101: well described effects of cannabinoids on cardiovascular disease - the authors should elaborate on this. I think that the cardiovascular effects of cannabis are not well known. In this sentence, it is not very clear whether the authors are talking about the therapeutic or adverse effects of cannabis consumption on cardiovascular events.

Line 103: The author should also mention the other phytocannabinoid - CBD (non-psychoactive component of the cannabis plant). It is mentioned later in the text but needs an introduction here.

Line 104-108: Please provide a reference for the following statement - studies investigating the potential of cannabinoids on SARS-Cov-2 infection.

Line 117-120: It is important to mention the composition of cannabis sativa arbel strain, that it contains CBD, THCV, CBG, and multiple terpenes, as the same study has looked at the differences in the anti-inflammatory effects of CBD by itself compared to F(CBD) strain in alveolar epithelial cells.

Line 149: Consider rewording as this is somewhat confusing to read.

Line 206: Please indicate the incubation time for WIN treatment for each experimental setup in the methods section.

Line 261 and following: Group size should be included in the data and statistical analysis section - three independent experiments, in your case? I am not sure if the sample size of 3-4 is statistically appropriate for estimating the P-value using unpaired-test or ANOVA.

Line 275: I would not say that WIN is a full agonist, there are other synthetic cannabinoids identified in the recreational market that activates the CB1 receptor with much higher potency and efficacy than that of WIN. Please refer to this paper https://doi.org/10.1111/bph.14829

Line 279: Fig S1, please label the band appropriately. It is not very clear what I am looking at.

Line 316 and following: The authors showed that WIN was able to reduce the inflammatory cytokines in cells infected with SARA-CoV-2. Was this effect mediated by the CB1/2 receptor - it could have been tested simply by pre-treating the cells with CB1 or CB2 antagonist.

 Line 330-333: This sentence should be rewritten for clarity as the subjects and descriptors may become confusing to the reader.

Line 368: This statement is a bit contradictory to what has been shown in the results section - Fig S1 indicates that hiPSC-CMs used in this study express both CB1 and CB2 receptors, so how does the ineffectiveness of WIN in reducing SARS-CoV-2 infection attributed to CB2 receptor?

Line 400 and following: there is a discussion on the involvement of CB2 receptors regarding the reduced levels of LDH by WIN. Due to lack of relevant data I find this discussion highly speculative, please rewrite the sentence, or omission is recommended.

Figure 3, It seems like asterisks representing statistical significance are missing from Figure 3D (72h incubation, SARS-Cov-2 ± WIN).

Textual remarks:
-	Abstract Line 57: replace “provoke” with “may cause”
-	Abstract Line 68: need internal consistency for SARS-CoV-2
-	Line 86: replace “which are injury biomarkers” by “a biomarker of cell/tissue injury”
-	Line 103: replace “agent” with “ingredient”
-	Line 121: need internal consistency for hyphenation (pre-treated or pretreated)
-	Line 125: need reference to support the statement
-	Line 138: solution instead of solutions
-	Line 172: …with or without treatment…instead of with treatments or not
-	Line 202: add respectively at the end of the sentence
-	Line 208: need consistency for hour or h
-	Line 211: ...overnight: anti-SARS-Cov-2 convalescent… add a colon, and remove the bracket
-	Line 226: don’t need to expand it every time - WIN is fine
-	Line 228-229: please provide the full form of an abbreviation when it is used first time in the paper, neutral red dye (NR) is an example
-	Line 230: …cells were washed again…
-	Line 235: after 24h treatment of hiPSC-CMs...with what?
-	Line 241: …and extracted samples…
-	Line 247: comma missing after bracket enclosing Sigma-Aldrich
-	Line 283: replace “regarding” with “for”
-	Line 328: …hallmark for cell/tissue damage…


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: IMPROVING SIZE SELECTIVITY OF ROUND POT FOR CHARYBDIS JAPONICA BY CONFIGURING ESCAPE VENTS IN THE YELLOW SEA, CHINA
Review round: 1
Reviewer: 1

Output:
Basic reporting: In general, the manuscript was well written. However, improvements on the following are suggested :

● The title can still be reduced. It is too long.

●It is better to indicate the word crab in the first line under Abstract so that those who not familiar (especially non-crab workers/researchers) with Charybdis would immediately know.

●It is better to show the figure of the conventional crab pot design beside the modified round pot.
Experimental design: The experimental design was described well.  However, information on the the basis/criteria in selecting the site for the deployment of crab pots should be included? How about the info on the crab population in the selected site?
Validity of the findings: In general, the data obtained were discussed well and the conclusions were based on the the acceptable discipline-specific repository.

There are some comments that need to be addressed though:

●Is there no MLS for male C. japonica in the decree of the Chinese Ministry of Agriculture? Is this the reason why the % males and females retained in the round pots with various sizes of escape vent was not determined? This could have been also a good information that can be useful in the future. In many fisheries ordinances, MLS for males and females are specified.

●Likewise, information on the by-catch was lacking (other animals that can be trapped in that type of crab pots? Animals other than crabs that entered the pots could have also fed on the bait. This could have  affected the entry of crabs based on the availability of baits/lures (which serve as attractant).
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: IMPROVING SIZE SELECTIVITY OF ROUND POT FOR CHARYBDIS JAPONICA BY CONFIGURING ESCAPE VENTS IN THE YELLOW SEA, CHINA
Review round: 1
Reviewer: 2

Output:
Basic reporting: This manuscript reports the result from catch comparison analysis of standard round pots vs pots with fitted escape vents to improve size selectivity. The set of experiments is well planned, the collection of data is appropriate and the analysis is valid. I find this manuscript suitable for publication after very minor revision.
Experimental design: The research question id clearly defines, relevant and meaningful. The set of experiments is well planned, the collection of data is appropriate and the analysis is valid. I would advise the authors to extent and clarify the definition of relative fishing intensity (lines 139-40). ..."relative to what"?
Validity of the findings: Definitely this study systematically assess and document the effect of different escape vents on the size selectivity of crabs in the Yellow Sea. This manuscript applies adequate models to carry out catch comparisons. It uses state of the art bootstrapping technique to estimate confidence intervals and applies exploitation indicators to assess the capture efficiency.
Additional comments: Minor comments:
Line 13: I would suggest deleting the word "little"
Line 139: It is a bit difficult to understand the term "relative fishing intensity". Please, provide a definition. relative to what? what do you mean by "intensity"


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: IMPROVING SIZE SELECTIVITY OF ROUND POT FOR CHARYBDIS JAPONICA BY CONFIGURING ESCAPE VENTS IN THE YELLOW SEA, CHINA
Review round: 1
Reviewer: 3

Output:
Basic reporting: The author has data set, although I noted lack of details on the suitable analytical method for drawing conclusions.  In addition, this manuscript is clearly written in a professional manner
Experimental design: Research question well defined
Validity of the findings: The findings on the effectiveness of window trap sizes are well presented well, with photograph and tables.  However, there is a slight disturbance because I have not found out what method is used to conclude that there is a significant catch size between trap models with different window size
Still needed a clearer explanation about determining the best window trap size.  Need a more suitable analytical method
Additional comments: Abstract:
It is necessary to add the analytical method used to determine the significance of each window traps size in selecting the size of the crabs caught

Introduction: good
Materials & method.
It is necessary to add the analytical method used to determine the significance of each window traps size in selecting the size of the crabs caught

Results:  Crab catch information should be displayed in more detail
Preferably, the number of catches for each trip and each trap is displayed.  Because if you look at the number of crabs caught, only 704 crabs (line 195).  This number gives an indication that the crabs are very few.  The number of traps is 50 with 9 hauls, so that when averaged only 1,56 crabs per trap for each haul.

Discussion: What about the influence of ocean currents on crab movement and the effectiveness of traps and the window in catching and selecting crabs according ti their size.  This information will be very interesting and add value to this article

Conclusion: Good.  The conclusion has answered the research objectives and the impact of the results of this research in the future


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: ANALYSIS OF HOW THE SPATIAL AND TEMPORAL PATTERNS OF FIRE AND THEIR BIOCLIMATIC AND ANTHROPOGENIC DRIVERS VARY ACROSS THE AMAZON RAINFOREST IN EL NIÑO AND NON-EL NIÑO YEARS
Review round: 1
Reviewer: 1

Output:
Basic reporting: This manuscript is well written and used professional English. The most prominent literature has been cited in this paper.
Experimental design: The experiment is well designed. However, surface fires under the tree canopy are difficult to be detected through MODIS, which means the authors could have missed the surface fires. It deserves a discussion on how the missed surface fire could affect your results. Additionally, the Amazon region is experiencing frequent clouds, which could degrade remote sensing quality. How did you solve this issue?
Validity of the findings: Data quality needs a thorough discussion.
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: ANALYSIS OF HOW THE SPATIAL AND TEMPORAL PATTERNS OF FIRE AND THEIR BIOCLIMATIC AND ANTHROPOGENIC DRIVERS VARY ACROSS THE AMAZON RAINFOREST IN EL NIÑO AND NON-EL NIÑO YEARS
Review round: 1
Reviewer: 2

Output:
Basic reporting: This research topic is relevant and interesting. The Abstract could provide more information to the reader of why this particular study is important and needed. More context and careful interpretation of the literature are needed to improve both the Abstract and Introduction. Also, many variables and metrics were presented in the Abstract and Introduction setting this study to be a very ambitious one, however, without cohesion and focus it became very confusing to understand properly what was being assessed and how all the variables were connected. It was challenging to assess if the chosen data and statistical analysis used were appropriate for this study because the objectives of this investigation were not clear at all. The results seemed interesting and instigated that this study has potential, but it was impossible to assess if the results had hit the mark once I did not know what we were aiming at. I strongly encourage the authors to work on this manuscript focusing on improving the clarity and focus of the study to simplify the message using only what is needed to deliver it, and thus make it a stronger paper.
Experimental design: The research question seems to be relevant and meaningful, however it is not well defined and explained in this manuscript. The Methods can be improved but more importantly they need to be better connected with the Introduction. It was difficult to evaluate if the Data and Methods are appropriate without knowing clearly what was being investigated.
Validity of the findings: Currently it is impossible to review and assess if the findings and the discussion around them are relevant or well stated as I cannot see how they are well linked with the original research question.
Additional comments: This is a promising study, and it seems the authors have good knowledge of statistics to provide solid analysis. Unfortunately, it was impossible to dive in the analysis and truly assess the results and interpret the discussion as I could not understand what was being investigated. I strongly encourage the authors to improve the packaging of this study and try and make the message very clear. I will be happy to review this manuscript in the future once the whole study is more cohesive.


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: ANALYSIS OF HOW THE SPATIAL AND TEMPORAL PATTERNS OF FIRE AND THEIR BIOCLIMATIC AND ANTHROPOGENIC DRIVERS VARY ACROSS THE AMAZON RAINFOREST IN EL NIÑO AND NON-EL NIÑO YEARS
Review round: 1
Reviewer: 3

Output:
Basic reporting: i) The English seem adequate.
ii) I identify a lack of understanding of the drivers of Amazonian droughts. There are a vast literature about it, one chan check José Marengo papers.
iii) The study focus on an important issue and has the potential to contribute to the scientific discussion, but there are two major problems in their design, which is described in the next section.
Experimental design: The article is interesting but there are three major problems in the experiment design, described below.
ii) I understand that there is an insufficient literature background, which deeply affects the methods: The authors define El Niño x non-El Niño years. Although El Níno is an important drought driver in Amazonia, this is not the single driver acting during the time-series studied. Large-scale weather patterns affect the incidence of rain in the Amazon. The essential driver of the hydrological cycle is water from the oceans. When heated, in intra- and inter-annual and decennial cycles in an anomalous manner, they cause changes in the large-scale atmospheric circulation and may intensify or attenuate the rain patterns on the continent. A clear connection is observed in the relationship between the surface temperature of the oceans and the rain: When the Atlantic Ocean is warmer than normal climatological conditions, in the region of the Atlantic Multi-Decadal Oscillation (AMO index), there is a decrease in the rain in the southwestern quadrant of the Amazon; When the Pacific is hotter than normal, in the El Niño phenomenon (MEI index), the reduction of rain occurs in t the central, eastern and southern parts of the Amazon. Finally, when the Ocean in the Pacific Decadal Oscillation region (PDO index) is hotter than normal, rainfall reduction is concentrated in the central part of the Amazon. Not accounting for these interactions properly, affects the climatological analysis, which becomes an incorrect representation of the spatial and temporal processes associated with droughts at the paper results.
iii) The dry and wet season definition from August to October is simply not adequate - the dry season in the northern hemisphere starts in November and goes up to March, as well as the fire peaks. Therefore, the analysis is incorrect for the North part of the biome above the equator (Venezuela, part of Colombia, Guyana..). Even in Southern Amazon, the dry season varies.
iv) Depending on when, during the year, the drought occurred (drought during the dry season or drought during the wet season) will deeply affect the fire characteristics and the forest vulnerability to the fire. Thus, the fire "accountability" in the paper results may be incorrectly captured.
Validity of the findings: The problems identified in the section above mean that the results are compromised, and must be completely revisited. Experiment design must be changed.
Additional comments: This paper aims to bring an important contribution with a method that has some potential. However, the problems in the experiment design make the results unreliable.

In addition to accounting for the dry years process correctly, which currently it is not, you also need to take into account the dry season and wet season correctly; and also, it must be identified when the drought hit the region/forest. For example, if the drought occurs during the dry season, it may slightly exacerbate the forest vulnerability to fires, but if the drought hits during the wet season, it means that the forest will enter the dry season already in water deficit, exacerbating its fire vulnerability. This is clearly what happened in southwestern Amazonia during the 2015/16 El Nino. Perpahs, instead of rainfall, you can use Water Deficit to better capture this and thus disentangle the drivers, as you tried to perform. By looking at rainfall only, the western part of the Amazon practically has no dry season, but one needs to look at the water deficit, which is better related to the forest vulnerability to fires due to water stress.

Some must-read papers:

BERENGUER E, CARVALHO N, ANDERSON Liana O., ARAGÃO LEOC, FRANÇA F, BARLOW J. Improving the spatial‐temporal analysis of Amazonian fires. Global Change Biology, letter to the editor , https://doi.org/10.1111/gcb.15279.

PESSÔA, A.C.M.; ANDERSON, Liana O.; CARVALHO, N.S.; CAMPANHARO, W.A.; JUNIOR, C.H.L.S.; ROSAN, T.M.; REIS, J.B.C.; PEREIRA, F.R.S.; ASSIS, M.; JACON, A.D.; OMETTO, J.P.; SHIMABUKURO, Y.E.; SILVA, C.V.J.; PONTES-LOPES, A.; MORELLO, T.F.; ARAGÃO, L.E.O.C. Intercomparison of Burned Area Products and Its Implication for Carbon Emission Estimations in the Amazon. Remote Sens. 2020, 12, 3864.   https://doi.org/10.3390/rs12233864

ANDERSON, L.O.; RIBEIRO NETO, G.; CUNHA, A.P.; FONSECA, M.G.; MENDES,  Y. M. ; DALAGNOL, R.; WAGNER, F.H.;  ARAGÃO, L.E.O.C.  Vulnerability of Amazonian forests to repeated droughts. Philosophical Transactions of the Royal Society B: Biological Sciences, 2018 373 20170411; doi:10.1098/rstb.2017.0411, 2018.

ARAGÃO, L.E.O.C.; ANDERSON, L.O.; FONSECA, M.G.; ROSAN, T.M.; VEDOVATO, L.; WAGNER, F.; SILVA, C.; JUNIOR, C.; ARAI, E.; AGUIAR. A.P.; BARLOW, J.; BERENGUER, E.; DEETER., M.; DOMINGUES, L.; GATTI, L.; GLOOR, M.; MALHI, Y.; MARENGO, J.; MILLER, J.; PHILLIPS, O.; SAATCHI, S. 21st Century drought-related fires counteract the decline of Amazon deforestation carbon emissions. Nature  Communications, 9, ( 536), doi:10.1038/s41467-017-02771-y

ARAGÃO, L. E. O. C.; MALHI, Y.; ROMAN-CUESTA, R. M.; SAATCHI, S.; ANDERSON, LIANA O.; SHIMABUKURO, Y. E. Spatial patterns and fire response of recent Amazonian droughts. Geophysical Research Letters, v. 34, p. doi:10.1029/200, 2007.

SILVA JUNIOR CHL; ARAGÃO L; ANDERSON Liana O.; FONSECA M; SHIMABUKURO Y; VANCUTSEM C; ACHARD F; BLEUCHE R; NUMATA I; SILVA C; MAEDA E; LONGO M; SAATCHI S. Persistent collapse of biomass in Amazonian forest edges following deforestation leads to unaccounted carbon losses. Science Advances  30 Sep 2020, Vol. 6, no. 40, eaaz8360, DOI: 10.1126/sciadv.aaz8360

SILVEIRA, M.V.F.; PETRI, C.A.; BROGGIO, I.S.; CHAGAS, G.O.; MACUL, M.S.; LEITE, C.C.S.S.; FERRARI, E.M.M.; AMIM, C.G.V.; FREITAS, A.L.R.; MOTTA, A.Z.V.; CARVALHO, L.M.E.; SILVA JUNIOR, C.H.L.; ANDERSON, Liana O.; ARAGÃO, L.E.O.C. Drivers of Fire Anomalies in the Brazilian Amazon: Lessons Learned from the 2019 Fire Crisis. Land  2020, 9, 516. https://doi.org/10.3390/land9120516

ARAGÃO, L.E.O.C.; ANDERSON, L.O.; FONSECA, M.G.; ROSAN, T.M.; VEDOVATO, L.; WAGNER, F.; SILVA, C.; JUNIOR, C.; ARAI, E.; AGUIAR. A.P.; BARLOW, J.; BERENGUER, E.; DEETER., M.; DOMINGUES, L.; GATTI, L.; GLOOR, M.; MALHI, Y.; MARENGO, J.; MILLER, J.; PHILLIPS, O.; SAATCHI, S. 21st Century drought-related fires counteract the decline of Amazon deforestation carbon emissions. Nature  Communications, 9, ( 536), doi:10.1038/s41467-017-02771-y

SILVA JUNIOR, C. H. L.;  ARAGÃO, L.E.O.C.; FONSECA, M.G.; ALMEIDA, C. T.; VEDOVATO, L. B.;  ANDERSON, L.O. Deforestation-Induced Fragmentation Increases Forest Fire Occurrence in Central Brazilian Amazonia. Forests 2018, 9(6), 305; https://doi.org/10.3390/f9060305.

The role of ENSO flavours and TNA on recent droughts over Amazon forests and the Northeast Brazil region JC Jimenez, JA Marengo, LM Alves, JC Sulca, K Takahashi, S Ferrett, ...
International Journal of Climatology

Extreme seasonal climate variations in the Amazon basin: droughts and floods
JA Marengo, ER Williams, LM Alves, WR Soares, DA Rodriguez
Interactions between biosphere, atmosphere and human land use in the Amazon …

The extreme 2014 flood in south-western Amazon basin: the role of tropical-subtropical South Atlantic SST gradient JC Espinoza, JA Marengo, J Ronchail, JM Carpio, LN Flores, JL Guyot
Environmental Research Letters 9 (12), 124007

Drought in Bolivia: The worst in the last 25 years JA Marengo, JC Espinoza, LM Alves, J Ronchail
https://doi. org/10.1175/2018BAMSStateoftheClimate. 1

Extreme seasonal droughts and floods in Amazonia: causes, trends and impacts
JA Marengo, JC Espinoza
International Journal of Climatology 36 (3), 1033-1050


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

Input:
Title: THE SOIL MICROBIOMICS OF INTACT, DEGRADED AND PARTIALLY-RESTORED SEMI-ARID SUCCULENT THICKET (ALBANY SUBTROPICAL THICKET)
Review round: 1
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: the fungal community should be also included in the study, especially it plays a very important role there.
Validity of the findings: no comment
Additional comments: The soil microbiomics of intact, degraded and partially-restored semi-arid succulent thicket (Albany Subtropical Thicket) by Schagen et al. reports the composition of soil microbiota between intact and degraded semi-arid succulent thicket. Albany Thicket biome is a typical ecosystem in South Africa and it is important to understand the soil microbiota that drive the nutrient cycling there.
Major comments:
1. Yes, soil microbiota includes bacterial and fungal communities. Why only bacterial community is examined in this study? If there is no specific reason for this, is it possible to add data of fungal communities?
2. As stated in lines 97-100 the aim of this study is to compare the composition of the soil microbiotas between the intact and degraded succulent thicket soils and identify environmental factors that could account for the observed changes in the microbial communities, the abstract and conclusion paragraph (lines 352-361) should include content of environmental factors.
3. lines 232-234 vegetation loss has no impact on the biodiversity. Could this be due to DNA extraction and PCR amplification biases? Was a “Mock community” included in the experiment?

Minor comments:
1. In the abstract, please state the aim of the study.
2. Please spread out the abbreviations “CAM” and “RDA” in their first appearance.
3. Data Analysis is poorly organized. Please describe each analysis step especially those in lines 143-149 in detail.
4. https://github.com/jasonbosch/The-soil-microbiomics-of-intact-degraded-and-partially-restored-semi-arid-succulent-thicket is not available. 16S rRNA data should be submitted to NCBI.
5. line 314 “A potential ecological implication of this observation are that niches”, “are” should be “is”?


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

Input:
Title: THE SOIL MICROBIOMICS OF INTACT, DEGRADED AND PARTIALLY-RESTORED SEMI-ARID SUCCULENT THICKET (ALBANY SUBTROPICAL THICKET)
Review round: 1
Reviewer: 2

Output:
Basic reporting: The manuscript entitled "The soil microbiomics of intact, degraded and partially-restored semi-arid succulent thicket (Albany Subtropical Thicket)" brings important considerations about the soil microbiota of impacted areas with pristine and reclaimed areas. I believe the manuscript raises interesting data. However I believe the authors can still value their results with some multivariate statistical analysis between abiotic factors (P and N) and microbial diversity (as suggested below)
Experimental design: 1) At what temperature was the soil transported to the laboratory? And the storage conditions during the period prior to DNA extraction should also be described in Methods.
2) Describe the protocol used for chemical soil analysis. Another option would be to provide the reference that describes the chemical analysis procedures.
3) Provide the sequences of the primers used in the amplifications or provide the references
4) How many sequencing were performed for each site? Please make this information clear here in Methodos, you in Results address data such as reads and others, but the sequencing number of each sample must appear here for other scientists to follow/compare your methodology.
5) Describe which software and methods were used for the multivariate analysis (PCA and RDA).
Validity of the findings: 1) In the section “Abiotic drivers of microbial community structure in both intact and degraded zones.” I believe that the authors should pay special attention to the levels of P and N as factors that influence diversity, these factors had influences in other studies. I strongly suggest that you read and cite the study: de Souza, L.C., Procópio, L. The profile of the soil microbiota in the Cerrado is influenced by land use. Appl Microbiol Biotechnol 105, 4791–4803 (2021).doi.org/10.1007/s00253-021-11377-w

2) Dear reviewers, I confess that I was unable to open your figures that would describe alpha-diversity. But anyway I believe that a picture describing comparing the abundance and diversity data could help in understanding the little difference between the two sites. It is common that when there is a loss of diversity, there is an increase in abundance. I believe their alpha diversity data could be further leveraged and discussed, including in the section "Unique to near-unique members of the core microbial community in each zone." And raise questions such as: these genera were more abundant in which place? The species of these genera were more diversified in which site?
Additional comments: The text is well written and the results are interesting and worthy of being discussed by the scientific community. Congratulations for the effort to study this important biome for humanity, although forgotten and highly degraded around the world


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

Input:
Title: THE SOIL MICROBIOMICS OF INTACT, DEGRADED AND PARTIALLY-RESTORED SEMI-ARID SUCCULENT THICKET (ALBANY SUBTROPICAL THICKET)
Review round: 1
Reviewer: 3

Output:
Basic reporting: Generally, good writing and structure. Some sections require major clarification  (see General comments for the author). Data is provided and the article is self contained.
Experimental design: Well defined aim and experimental design within the scope of the journal. Some parts of the methods require clarification (see General comments for the author).
Validity of the findings: The overall conclusions are well stated and not overly speculative. The novel case study provides seemingly robust findings, although some aspects require further evaluation  (see General comments for the author).
Additional comments: The comments I provide here are not general but mostly very specific.

The manuscript provides a compelling case study for investigating the effects of restoration efforts on the soil microbiome in arid/semi-arid regions. The study investigates differences in physicochemical properties and the soil microbial community composition between intact, restored and degraded succulent thicket.
The study provides evidence of systematic shifts in bacterial community composition that could enable the detection of plant-associated indicator taxa. Furthermore, the restored regions and their microbiome seem to lie between the intact and degraded regions, suggesting a gradual transition.
Overall, the paper is well written, concise and easy to follow with a clear aim and experimental design. However, there are some aspects regarding methodology and interpretation of results that require clarification (see below). I can thus recommend publication after major revision.

1. It is not entirely clear how the ASV data was treated in the analysis. Was the data normalized? How? I think the differences in read numbers should be accounted for when comparing proportions across samples (1:10’000 reads is very different from 1:1’000’000). At least it should be reported or the authors could demonstrate that it does not affect their conclusions (e.g., regarding differences in diversity).
2. Several thresholds are used based on relative abundance. Again, this is difficult to interpret (see comment 1.). Also, I think the authors should show how different thresholds affect the outcome of the analysis.
4. I am not sure about the section on co-occurrence and the interpretation of interactions. It appears a bit out of the blue and does not substantially add to the conclusions of the manuscript. Maybe I missed something?

Comments as I went through the text (major comments marked with '*':
- L21: Maybe better state the years after replanting (instead of the year), since readers do not know when the study/sampling took place.
- L26: remove 'in'
- L27: "wide variety of habitats" does this refer to global (i.e. across continents and biomes: desert, forests,...) or regional habitats?
- L27-30: I suggest to split into two sentences and slightly adapt. "[...] threatened biome. It provides [...] function, concerning the restoration of degraded [...]"
- L50: Maybe break the sentence: "[...] 2013). These regions are referred to as [...]"
- L52: "[...] attributed to [...]"
- L77: please specify: "[...] the {region/ecosystem/soil?} remains [...]"
- L78-L82: The sentence could be written more clearly by first mentioning the attempts made ("replanting") and then (in a second sentence) detailing their success.
- L83: it is not clear what "these" refers to. I suggest to either specify or drop the word.
- L91-94: Here, I think also climate should be mentioned. Maybe this section could be expanded by providing some background for arid environments - e.g.: How is the soil/rhizosphere microbiome affected by water limitation? Relations of precipitation with pH, and organic matter contents?
- L98-100: Nice and clear!
- *L154-155: Why was this chosen? How about the other normalization methods (e.g., rarefy) that account for sampling depths?
- Fig. 1: The figure gives a good impression of the biome and nice overview of the sampled locations. Maybe add whitespace between the images. Also, the order is difficult to follow; maybe change to: (A) overview, (B + C) etc.
- Table S1: Please provide units for the measured values.
- *Fig. 2: The text is very small making the figure difficult to read (also some other figures). What was the proportion of variance explained in A?
- *L185-186: How was this tested? This should be clearly mentioned in the main text. Please, provide mean and standard deviation (also elsewhere in the text).
- L188: What "complex"?
- *L197-206: The significance test should be mentioned. Also, why were minimum/maximum values chosen for significance testing? This should be more explicit in the main text.
- L215: I think this suspicion could be tested by obtaining rainfall data (e.g., from satellite data https://disc.gsfc.nasa.gov/datasets).
- Fig. S5: It seems that the variability of diversity between samples was larger for the intact site compared to the degraded one. Maybe this is worth mentioning (also in context of the statistical tests used). Furthermore, if this pattern is consistent with the physicochemical measurements, it could indicate heterogeneity in the samples with presence of vegetation.

- L237: This is difficult to interpret. Was the data normalized (e.g., rarified)? Sequence reads might be high for a single sample with great sampling depth and could inflate the dominance of a particular species across samples.
- L237-238: "[...] three sampled zones [...]"
- *L235-246: It is not entirely clear to me how this was applied. Why was 95% of samples chosen? Others could be tested. Are those samples from the same group (e.g., 95% of intact samples)? I would expect that all samples are normalized (taking into account differing number of reads) and then the number of nonzero entries for each species is divided by the total number of samples to obtain the prevalence/ubiquity/occupancy.
- L251-252: How about the variation of these "rare" ASV? How did their proportion vary?
- *L268-270: It is a bit unclear what was used. E.g., are the variables soil density, soil moisture and temperature considered a "soil chemistry variable"? Also, maybe use "soil physicochemical property". Lastly, I could not follow where the 50% come from.

- *L276-279: Again, why were those thresholds chosen? Others should be tested. E.g., the authors could show how the number of ASV changes for differing thresholds.
- L288: Average pH value?

- *L303-308: I am not sure what this means in the context of network complexity. What metrics were compared to quantify the complexity (e.g., modularity, centrality, degree distribution)? How was the network constructed and what clustering method was used?
- *L309-315: The interpretation of interactions obtained from co-occurrences should be handled with care, especially in heterogeneous environments such as soil (see https://doi.org/10.1038/s41396-019-0463-3). This should be discussed.

- L338: I like this section for comparisons with the restoration. The intermediate position supports the robustness of findings.


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

Input:
Title: THE SOIL MICROBIOMICS OF INTACT, DEGRADED AND PARTIALLY-RESTORED SEMI-ARID SUCCULENT THICKET (ALBANY SUBTROPICAL THICKET)
Review round: 2
Reviewer: 1

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


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

Input:
Title: THE SOIL MICROBIOMICS OF INTACT, DEGRADED AND PARTIALLY-RESTORED SEMI-ARID SUCCULENT THICKET (ALBANY SUBTROPICAL THICKET)
Review round: 2
Reviewer: 2

Output:
Basic reporting: 132 / 5000
Resultados de tradução
All inquiries have been answered. Although the reference requested to be citata was not inserted in the final text, unfortunately
Experimental design: All done
Validity of the findings: All done
Additional comments: All done


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

Input:
Title: THE SOIL MICROBIOMICS OF INTACT, DEGRADED AND PARTIALLY-RESTORED SEMI-ARID SUCCULENT THICKET (ALBANY SUBTROPICAL THICKET)
Review round: 2
Reviewer: 3

Output:
Basic reporting: OK. Hypotheses could be more clearly stated.
Experimental design: OK. Methods need further writing.
Validity of the findings: OK
Additional comments: Overall, the manuscript improved and reads more fluently. However, there are still two issues that need to be resolved before I could recommend publication:
(i) The methods and the motivation behind them need to be clearly reported in the main text. Referring to computer code is not acceptable. It is not clear how data was treated. How exactly were temperature and humidity aggregated? Were proportions used or rarefied counts?
(ii) Different thresholds should be tested (as mentioned in my previous report) to show robustness of the findings. Commonly used thresholds are still arbitrary and thus not necessarily relevant. Showing how the outcome varies with different thresholds provides valuable insight into the robustness of analysis and distribution of the data (I am not suggesting to use multiple thresholds – only testing them).
Furthermore, some editing and proofreading is required.
Additional comments as I went through the text:
L88-90 delete 'elsewhere in the world'. What is meant here by ‘soil stability’?
Also, this hypothesis is always true and cannot be tested (i.e., they could always play a role). Change to 'Soil bacterial diversity is important for maintaining ...' and then introduce your own hypothesis: 'We hypothesize that increased diversity increases soil ... in the Albany Subtropical Thicket.', however, given the aim of the study (L103) it could be 'We hypothesize that changes in environmental factors due to the loss of succulent thicket vegetation explain shifts in soil microbiome composition.' (Ideally a hypothesis would also come with defined direction)
L96 The part on the 'feedback loop' is not relevant for this study and can be removed (including the reference Rocca et al. 2019 that introduced a database and no mechanism).
L116 delete 'as part of a restoration experiment'
L121 change '... and a "site" is the point ...'
L122 change '... soil chemical analysis ...'
L128-131 This sentence is unclear. First introduce the RDA and the motivation for using it. Then in a second sentence explain how the variables were aggregated (not 'generated' I hope...). What was the input? Mean and std? Why? Also, later there is mention of daily values. This needs to be cleared up. How were things aggregated? The authors refer to their code in the point by point response, but I do not think this is ok. Most readers do not have the time to read 1000 lines of code. Additionally, GitHub is not a proper archive and something like Zenodo (with DOI) should be used to guarantee future access.
L128 Why was such a high resolution chosen? Were the values filtered/smoothed? Was the raw data used to do the analysis? e.g., a single temperature/humidity peak within two hours could have been chosen as the maximum value used for further analysis (and would be considered representative for the sample)? Is this robust?
L162-164 Does this make sense? I suggest the authors repeat the alpha diversity analysis on normalized (rarefied) data. Otherwise I think it is neither representative nor interpretable. How big was the variation in reads (not written in the main text - 3x is not nothing and depends on the range e.g., 10000-30000)? How does the (expected) number of individuals (cells) compare to the number of reads?
L166 'agglomerated'?
L170 'The final model was tested and evaluated by ANOVA to ensure that all terms were statistically significant' what is the point of this (statistically speaking)? If a term is insignificant it should be kept in the model and it would not affect the outcome.
L176 please rephrase. Is it 'different in abundance' or 'differentially abundant'?
L182 replace '2' with 'two'
L188 what is 'shallow-subsurface'?
L201 How about the water retention due to organic matter?
L228 move 'content' before the bracket after 'NH4-N'
L299 'These results replicate previous findings for ...'
L246-248 Please rewrite. This sentence is too long and very difficult to follow. E.g., if everything is consistently aggregated to daily values (and reported in the methods) then it would not need to be stated again. Why should the maximum and minimum be compared here?
L253-254 Was this 'lens' observed at the same site? If not, can it be generalized? I suggest to be clear that this is speculation. In general organic matter would decrease bulk density and increase porosity and, thus, water infiltration.
L264 This would not be 'unexpected' if dispersal is not limiting (due to proximity of sites),; i.e., richness is constant and only abundance changes.
L283 The dominance of few species is well known in ecology and does not only apply to soil microorganisms.
L313 This suggests that the water balance is altered (e.g., http://www.nature.com/doifinder/10.1038/nature20139) and could affect the community structure (i.e., evenness). I suggest to reconsider interpretation (L316-318) since both components (humidity and temperature) are responsible for changes in water balance. Also please fix the typos and introduce the hypothesis at the beginning.
L399 remove 'tend to'


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: POTENTIAL SOURCES AND CHARACTERISTIC OCCURRENCE OF MOBILE COLISTIN RESISTANCE (MCR) GENE-HARBOURING BACTERIA RECOVERED FROM THE POULTRY SECTOR: A LITERATURE SYNTHESIS SPECIFIC TO HIGH-INCOME COUNTRIES
Review round: 1
Reviewer: 1

Output:
Basic reporting: No comment
Experimental design: No comment
Validity of the findings: No comment
Additional comments: Comments_peerj2020_1
The authors review the mcr-harboring bacteria of poultry origin in high-income countries. The manuscript is very long but loose and difficult to read. The sequence of the context is organized disorderly. Numerous numbers (of isolates, date, …) listed in the Tables are inconsistent with those described in the text. It is recommended that authors check the tables and the text for errors and arrange the order of countries or regions in the tables to match the narrative order of the text. The manuscript should be shortened by half to make it more focused and concise.

Other comments:
1.	Line 38: Since only very few studies characterized the virulence genes of isolates, it is suggested that the authors exclude this part (virulence) to make the review article more focused.
2.	Line 145: spelling error “Enterobacteriacae”
3.	Lines 59-60: should give a citation for this statement
4.	Line 211: Ninety-nine “strains”… Here, “isolates” rather than “strains” is more appropriate. For many places in the text, isolate(s) instead of strain(s) is more proper.
5.	Lines 211-212: “Salmonella enterica subspecies Typhimurium”  The full name should be Salmonella enterica subspecies enterica serovar (or serotype) Typhimurium (abbreviation: S. Typhimurium)
6.	Line 216: To replace “Salmonella enterica subspecies enterica Typhimurium” with the abbreviation “S. Typhimurium” and to use the abbreviation in the following text.
7.	Lines 366-367 (and several places in the text): …In Portugal, 57 strains (56 E. coli and 1 Salmonella enterica Typhimurium) were detected among 242 Enterobacteriaceae (23.6%) … It is recommended moving the percentage (23.6%) to the position after the number (57)… as “In Portugal, 57 (23.6%) isolates (56 E. coli and 1 S. Typhimurium) were detected among 242 Enterobacteriaceae isolates …
8.	Line 626: To replace “S. Paratyphi B dT+” with “ S. Paratyphi B var. Java” (in all the text and tables). D-tartaric acid “–“ is S. Partyphi B, d-tartaric acid “+” is S. Paratyphi B var. Java
9.	Line 673: “APEC isolated”  To spell out the full name of APEC as “avian pathogenic E. coli (APEC) isolates”
10.	Line 684: “mcr-“  Is mcr-1?
11.	Line 678 and the paragraph: mcr-9 has been shown not to offer resistance to colistin; it should not be important to this article and not be described as the first paragraph in the section of North America.
12.	Line 728: … in the High-income countries (HICs)… The full name of HICs should not be re-written here again.
13.	Line 735: both terms “farm-to-table” and “farm-to-plate (line 680)” are mentioned in the text. Should use only one?
14.	Table 1: the column title “Source of isolate (Number of birds)” However, no “number” appears in this column. South Korea: mcr-1 is detected in 10 E. coli isolates but 12 STs are identified for the isolates. Taiwan: Abbreviation “S. Typhimurium” is recommended; 3 STs are listed but which one(s) are for the S. Typhimurium isolates and which one(s) are for the E. coli isolates? It is recommended that virulence genes should not be discussed in this review article.
15.	Table 2: Netherlands: mcr-1 is detected in 14 E. coli and 13 Salmonella isolates and 10 STs are listed for the isolates; which STs are for the E. coli isolates, which one(s) are for the Salmonella isolates?


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

Input:
Title: POTENTIAL SOURCES AND CHARACTERISTIC OCCURRENCE OF MOBILE COLISTIN RESISTANCE (MCR) GENE-HARBOURING BACTERIA RECOVERED FROM THE POULTRY SECTOR: A LITERATURE SYNTHESIS SPECIFIC TO HIGH-INCOME COUNTRIES
Review round: 1
Reviewer: 2

Output:
Basic reporting: The review entitled "Potential sources and characteristic occurrence of mobile colistin resistance (mcr) gene-habouring bacteria recovered from the poultry sector: A literature synthesis specific to high-income countries" by Madubuike Umunna Anyanwu et al. aims to undestand the sources, prevalence, and genotypic characteristics of mcr-haroring bacteria in the poultry sector found in high-income countries. Overall, this review discusses the factor that could lead to a high prevalence of mcr-harboring bacteria and by comparing the countries between them show a general view of the current situation and action to reduce this prevalence. However, this review need to address a number of points in its current form. These are highlighted below.

Line 29: "Currently, 10 mcr genes (mcr-1 to mcr-10) are described". I sugget to change "are" by "have been".

Line 32: Escherichia coli, Klebsiella, Salmonella, and Aeromonas isolates. Klebsiella spp. or Klebsiella pneumoniae. The same for Salmonella.

Line 38: virulence factors. factors word should be added.

Line 70: "clinicians were forced to using COL for the treatment of". I sugget to change "using" by "use".

Line 80: "Plasmids are self-replicating DNA independent of chromosomes, and
80 they are highly mobile, carrying resistance and virulence-associated genes (VAGs)". This is not true at all, some plasmid are non-conjugative plasmid and need the machinary from another plasmid to be mobilized.

Line 112-117: "To reiterate the potential sources of plasmid-mediated colistin resistance (PMCR), particularly within the poultry sector, and to be specific, high-income countries (HICs) should be considered of very great importance. More importantly, to gather information about the characteristic occurrence of mcr gene-habouring bacteria (MGHB) recovered from the poultry sector specific to the high-income countries should also be considered of great importance". This part is repetitive and useless for the reader, it should be shorter. I suggest -> the potential source and the characteristic ocurrence of mcr-harboring bacteria (MGHB) particularly within the pultry sector specific to the high-income countries should be considered of great importance.

Line 189: "Nevertheless, some plasmids could increase in frequency and capture ARGs even in the absence of selective pressure (Lopatkin et al., 2016)".This is not correct and the reference used not say that the absence of exposition to antibiotics can increase the frequency of ARGs. I suggest to eliminate from the sentence -> "increase in frequency".

Line 259 and 262: I found reference missing at the end of the paper and names that were wrongly writed on the text. Please check again carefully all the references since it is a review and it is one the most important part due to the acquisition of the information it is from the reference. The same mistake were also found in Line 603,604,605,613 and 614,631,649.

Line 352: "transmission since the COL determinants were also detected". You should include resistance between COL and determinants.

Line 366: You should write -> 57 mcr-harboring strains.

Line 571: Sometimes the prevalence in percentage is given in comparison with all samples tested and in this part related to the COL-resistant isolates, creating the false impresion to the reader of a high prevalence of mcr-1. Considering the strains tested on the reference Allerberger et al. 2016 (164) and the mcr-1-positive isolates found. the prevalence is 1.8%.

Line 578: During all the text, you are using in excess the word "Unfortunately" that have sinonyms such as lamentably regrettably or dismally. By this way, the repetition could be avoided.

Line 649: Change mcr-1 by mcr-5.

Line 696: "However, the qseBC two-component system, which is thought to induce mcr-9 expression". Reference of the induction of mcr-9 expression should be included. DOI: 10.1128/AAC.00965-19.

Table 1: Remove "Number of birds" second column.

Table 2 (France): Instead of 24 is 275 in column 4. In column 5 is 3 and 23 instead of 2 and 21.

Table 2 (Austria): 168 instead of 4.
Experimental design: no comment
Validity of the findings: The conclusions should be more complete, I missed mainly a comparison in term of mcr genes prevalence between the different HICs countries and the factors that enhance this prevalence in comparison with other HICs such us, high level of food importation from developing countries, heavily use of COL in livestock, etc. This discussion between the different HIC countries would increased the quality of the conclusion.
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: POTENTIAL SOURCES AND CHARACTERISTIC OCCURRENCE OF MOBILE COLISTIN RESISTANCE (MCR) GENE-HARBOURING BACTERIA RECOVERED FROM THE POULTRY SECTOR: A LITERATURE SYNTHESIS SPECIFIC TO HIGH-INCOME COUNTRIES
Review round: 1
Reviewer: 3

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: Rapid dissemination of mobile colistin resistance (mcr) genes is particularly concerning and poses a major medical challenge. This manuscript described the dissemination of the mcr alleles(mcr-1 to mcr-10) among Enterobacteriaceae strains collected from food animals in high-income countries. The process of acquisition of mobile colistin resistance gene depends on various genomic background associated with insertion sequences. mcrs can be quickly transmitted among bacterial community through horizontal transfer. Also, the authors presented the other information of mcrs gene, including their prevalence, plasmids type and sequence type.

Limitations:
The authors collect information about all mcrs gene (mcr-1 to mcr-10) but did not listed the similarities and differences among these genes in different high-income countries. Overall, the logic of this manuscript was a little bit messy even though heading was used, which’s so hard to quickly get useful information without a clear structure. There are many grammatical mistakes needed to be corrected. Besides, the tables were not definite, terse, lacking clear, logical and comparative function.
I’m afraid this manuscript is more discursive than necessary and would benefit from some condensation. To be honest, I afraid that this manuscript is not well organized and would not be appropriate for PeerJ. It would also benefit from attention to the following points.

Detail points:
Line 72: It is not appropriate to use ccrB, it would be crrAB. Also, the related references should be used here.
Line 86: It would be “lipopolysaccharide of lipid A” rather than“lipid A lipopolysaccharide”.
Line 87-88: Authors should correct the describe of the mcr-1-mediated colistin mechanism.
Line185-188: It is not appropriate to say that China services as a source for dissemination of plasmid-mediated colistin resistance. Authors should be careful to use the word of causal relationship. Recent literatures in Lancet Infect Dis have been showed that the decreasing use of colistin in agriculture have had a significant effect on reducing the prevalence of mcr-1 in China.
Additionally, it would be better to list actions taken by authorities in response to the mcrs gene and summary the prevalence of mcrs gene after the intervention. For example, the European Medicines Agency updated advice on the use of colistin in European veterinary practices and recommended that colistin should be included in category 2 of Antimicrobial Advice ad hoc Expert Group.
For Figure1, the summary was partial, like mcr-1, mcr-3, and mcr-5 have been detected in E. coli derived from diseased pigs in Japan.
For the tables, authors should better highlight the important information.


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: POTENTIAL SOURCES AND CHARACTERISTIC OCCURRENCE OF MOBILE COLISTIN RESISTANCE (MCR) GENE-HARBOURING BACTERIA RECOVERED FROM THE POULTRY SECTOR: A LITERATURE SYNTHESIS SPECIFIC TO HIGH-INCOME COUNTRIES
Review round: 2
Reviewer: 1

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


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

Input:
Title: POTENTIAL OF ARTIFICIAL INTELLIGENCE TO ACCELERATE DIAGNOSIS AND DRUG DISCOVERY FOR COVID-19
Review round: 1
Reviewer: 1

Output:
Basic reporting: The author reviewed this MS by how Artificial Intelligence can be utilized not only in disease identification but also in drug discovery. However, your article is inadequately presented. Besides, there are numerous grammatical mistakes and spelling mistakes, as well.
Although the article has scientific rigor, several major issues need to be changed before publication.
Experimental design: 1. The current manuscript will advance no new information/recommendation for the readers.
2. Many non-scientific and incorrect/wrong information/sentences are there, which may mislead the readers.
3. The authors just have addressed several points randomly. Many sentences/information throughout the manuscript have serious flaws that withdrawn my thought from it.
4. Every section of the manuscript must be written scientifically according to the possible literature with relevant references.
5. The language needs to be improved considerably, mostly in the introduction and abstract. There are several errors that make it difficult to understand the manuscript.
6. The work seems preliminary. The study problem is not apparent. Can the authors explain the importance of Artificial Intelligence in the introduction section?
7. The article title is very vague. The authors need to simplify the title. It looks like a research MS by sighted the title.
8. The authors make errors in the naming of COVID-19 and SARS-CoV-2. SARS-CoV-2 is a virus that causes COVID-19 (Coronavirus Disease-19). Please make sure that this is correct.
9. The abstract section is inacceptable—no focus point in the abstract section.
10. The introduction section is inapplicable. Need to change the introduction considerably. Try to include the existing barriers also, how the present review unties those limits.
11. English is feeble. The authors need to improve their writing style. The whole manuscript needs to be checked by native English speakers.
12. Authors are suggested to replace \'Covid-19\' with \'COVID-19\' throughout the manuscript.
13. The research questions could be defined more precisely. This is probably due to language concerns.
14. The author should discuss more literature survey with references.
15. What is the need for 'COVID-19’ detection methods?
Validity of the findings: 1. ‘Artificial Intelligence in disease identification and drug discovery’ should be the main focus point. The authors need to elaborate on this section in detail.
2. Very few references are cited here. This is a review article. The authors should increase the number of references.
3. The conclusion needs to address future perspectives.
Additional comments: Conclusively, there are likewise problems to address. I found this manuscript very foundation and do not recommend to publish in the present form. I also find the present manuscript lacks for its originality. Fundamental changes are obligatory before go for the final version.


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

Input:
Title: POTENTIAL OF ARTIFICIAL INTELLIGENCE TO ACCELERATE DIAGNOSIS AND DRUG DISCOVERY FOR COVID-19
Review round: 1
Reviewer: 2

Output:
Basic reporting: The present review targeted potential area that is vital in disseminating the role of AI in COVID-19 patients data in disease identification and drug discovery approach. However, authors have not gone in depth of machine learning, IoT and role big data analytics in a way that meets the standard of PeerJ guidelines.   The following drawbacks are noted
1. lack of professional English
2. repeated use of similar references (Literature Survey)
3. Though, authors tried to pull literature from latest published literature, there is scope for improvement in structure of article.
Experimental design: The study design is very much focused on survey of recent literature and lacks coverage from global resources (databases) like WHO on COVID -19 from UK, US, Brazil, INDIA which are worst hit and how they are doing data analytics using Big data and machine learning. There is vast amount of literature recently deposited globally on the subject under discussion in the review paper. This is major drawback of the review paper where it lacks professional approach.
Validity of the findings: Though,  authors have developed and supported argument that meets the goals set out in the introduction. However, authors have not focused on role of AI in drug discovery in detail. this leads to inconsistency in the whole approach of Review paper.

Conclusion identifies future scope of the work on AI.
Additional comments: The review sheds light of AI and drug discovery which is critical area of research in COVID-19 in diagnosing disease and control authors have a not fully met the standards of PeerJ journal which is having wide audience.


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: POTENTIAL OF ARTIFICIAL INTELLIGENCE TO ACCELERATE DIAGNOSIS AND DRUG DISCOVERY FOR COVID-19
Review round: 1
Reviewer: 3

Output:
Basic reporting: The article is well written. The content included is quite good and is useful to the scientific community working for COVID-19. The concepts are well mentioned and the manuscript is also organized neatly.
However, the references mentioned in the manuscript are not in the same format. Please change the references to this journal format.
Experimental design: no comment
Validity of the findings: no comment
Additional comments: To conclude on this article, change all the references as per this journal format.


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: POTENTIAL OF ARTIFICIAL INTELLIGENCE TO ACCELERATE DIAGNOSIS AND DRUG DISCOVERY FOR COVID-19
Review round: 2
Reviewer: 1

Output:
Basic reporting: The author reviewed this manuscript by how Artificial Intelligence can be utilized not only in disease identification but also in drug discovery. However, your article is inadequately presented.
Although the article has scientific rigor, several minor issues need to be changed before publication.
Experimental design: 1. The language still needs to be improved, mostly in the introduction section. There are several errors that make it difficult to understand the manuscript.
2. The introduction section is inapplicable. Need to change the introduction considerably. Try to include the existing barriers also, how the present review unties those limits.
3. Please add a section named AI IN DRUG DISCOVERY.
Validity of the findings: 1. Please add a section named AI IN DRUG DISCOVERY.
2. Figures 1 and 2 need to illustrative, not just presenting in a text form.
Additional comments: Though the authors addressed some of my comments still, I found this manuscript not suitable for publication. Minor changes are obligatory before go for the final version.


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

Input:
Title: POTENTIAL OF ARTIFICIAL INTELLIGENCE TO ACCELERATE DIAGNOSIS AND DRUG DISCOVERY FOR COVID-19
Review round: 3
Reviewer: 1

Output:
Basic reporting: Title: Potential of artificial intelligence to accelerate 1 diagnosis and drug discovery for COVID-19 (#51371)

Journal: PeerJ

The author reviewed this manuscript by how Artificial Intelligence can be utilized not only in disease identification but also in drug discovery. However, your article is inadequately presented.
Although the article has scientific rigor, several minor issues need to be changed before publication.
Experimental design: 1. Check the references. Many inappropriate (format and style) are in there.
2. Check the Figure 1 legend.
Validity of the findings: 1. Figures 2 need to illustrative, not just presenting in a text form.
Additional comments: Title: Potential of artificial intelligence to accelerate 1 diagnosis and drug discovery for COVID-19 (#51371)
Journal: PeerJ

BASIC REPORTING

The author reviewed this manuscript by how Artificial Intelligence can be utilized not only in disease identification but also in drug discovery. However, your article is inadequately presented.
Although the article has scientific rigor, several minor issues need to be changed before publication.

EXPERIMENTAL DESIGN

1. Check the references. Many inappropriate (format and style) are in there.
2. Check the Figure 1 legend.

VALIDITY OF THE FINDINGS

1. Figures 2 need to illustrative, not just presenting in a text form.


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: POTENTIAL OF ARTIFICIAL INTELLIGENCE TO ACCELERATE DIAGNOSIS AND DRUG DISCOVERY FOR COVID-19
Review round: 3
Reviewer: 2

Output:
Basic reporting: After revision, manuscript looks professional with literature.
Experimental design: Article content is within the Aims and Scope of the journal.
Validity of the findings: The conclusion address future needs of medicine and health and supports with recommendation.
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: POTENTIAL OF ARTIFICIAL INTELLIGENCE TO ACCELERATE DIAGNOSIS AND DRUG DISCOVERY FOR COVID-19
Review round: 4
Reviewer: 1

Output:
Basic reporting: The author reviewed this manuscript by how Artificial Intelligence can be utilized not only in disease identification but also in drug discovery. However, your article is inadequately presented.
The authors addressed my comments accordingly. Just some minor spell checks are required (corona virus will be coronavirus, etc.).
The revisions look good to me. The manuscript is good enough to publish in the PeerJ journal.
The authors did an excellent job.
Congratulations!!!
Experimental design: The revisions look good to me. The manuscript is good enough to publish in the PeerJ journal.
Validity of the findings: The revisions look good to me.
Additional comments: Looks okay now.


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: CHARACTERIZATIONS OF NOVEL PESTICIDE-DEGRADING BACTERIAL STRAINS FROM INDUSTRIAL WASTES FOUND IN THE INDUSTRIAL CITIES OF PAKISTAN AND THEIR BIODEGRADATION POTENTIAL
Review round: 1
Reviewer: 1

Output:
Basic reporting: Overall paper is good; there are few minor changes that should be made.
Experimental design: Kindly give the reference of the Cetyl trimethyl ammonium bromide (CTAB) method, which is mentioned in the methodology.

Mention which version of the MEGA X software was used.

In the discussion section, it would be better to write short names of the microbes.
Validity of the findings: No comments
Additional comments: No additional comments


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

Input:
Title: CHARACTERIZATIONS OF NOVEL PESTICIDE-DEGRADING BACTERIAL STRAINS FROM INDUSTRIAL WASTES FOUND IN THE INDUSTRIAL CITIES OF PAKISTAN AND THEIR BIODEGRADATION POTENTIAL
Review round: 1
Reviewer: 2

Output:
Basic reporting: The paper is well written and grammatically correct. The figures and the rest of the things are according to the journal format.
Could you replace this reference  with an updated one;
Saitou N, N. M. The Neighbor-Joining Method: A New Method for Reconstructing Phylogenetic Trees. Mol. Biol. Evol. 1987. https://doi.org/10.1093/oxfordjournals.molbev.a040454.
Experimental design: The methodology is quite convincing, but my questions are about future perspectives. In conclusion, describe the future perspective, at which stage microbes degrade the DDT and other pesticides. Can the damaged land be recovered?
Validity of the findings: The statistical analysis is acceptable for me.
Additional comments: In acknowledgement, National Institute of Biotechnology and Genetic Engineering (NIBGE) Faisalabad-Pakistan,  name should be mentioned .


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: CHARACTERIZATIONS OF NOVEL PESTICIDE-DEGRADING BACTERIAL STRAINS FROM INDUSTRIAL WASTES FOUND IN THE INDUSTRIAL CITIES OF PAKISTAN AND THEIR BIODEGRADATION POTENTIAL
Review round: 1
Reviewer: 3

Output:
Basic reporting: The study seems interesting, and reporting the novel pesticides degrading strains seems interesting. The paper is well written scientifically. No major grammatical error was found.
Yet there are a few suggestions which could be followed:
•	Remove the full stop at the end of the title.
•	Remove the colon (:) after the heading “contribution”.
•	The heading “Abstract” and the description should not be in the same line.
•	Kindly write the manufacturer’s name of the thermocycler used in the methodology.
•	After the conclusion's heading, remove the (:) and start with the new line.
•	After the Keyword heading, remove (:) and starts with the new line.
Experimental design: I am convinced with the experimental design. The authors have thoroughly explained the research question.
Validity of the findings: The novelty if the finding were fully addressed and informative, and it seems it would help the researchers a lot in the future. The statistical result is quite clear.
Additional comments: The writing style is satisfactory.


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: CHARACTERIZATIONS OF NOVEL PESTICIDE-DEGRADING BACTERIAL STRAINS FROM INDUSTRIAL WASTES FOUND IN THE INDUSTRIAL CITIES OF PAKISTAN AND THEIR BIODEGRADATION POTENTIAL
Review round: 1
Reviewer: 4

Output:
Basic reporting: The manuscript "Characterizations of novel pesticide degrading bacterial strains from industrial wastes found in the industrial cities of Pakistan and their biodegradation potential" by Noreen Asim and colleagues describes the isolation of 6 different bacterial strains that are able to grow in DDT, aldrin or malathion as only carbon source from different environmental samples. This study provided strong evidence for utilizing these strains to remove persistent residual pesticides; thus, it gives potential for soil treatment and restoration.
Experimental design: The presentation and details are not sufficient.
Validity of the findings: The poor discussion of the results.
Additional comments: The manuscript "Characterizations of novel pesticide degrading bacterial strains from industrial wastes found in the industrial cities of Pakistan and their biodegradation potential" by Noreen Asim and colleagues describes the isolation of 6 different bacterial strains that are able to grow in DDT, aldrin or malathion as only carbon source from different environmental samples. This study provided strong evidence for utilizing these strains to remove persistent residual pesticides; thus, it gives potential for soil treatment and restoration. Overall, this work is interesting but the presentation and details are not sufficient. Some comments and suggestions are listed as follows for improvement.

Major comments：
1- The poor discussion of the results. Author just shows the great amount of results that they have achieved, but they did not use them to develop an interesting discussion which could supplement to earlier studies on biodegradation processes carried out by pure cell cultures.
2- Errors in grammar and language editing. Authors are responsible for preparing their papers in correct English language. The manuscript requires substantial grammatical revisions in its present form and it should not be accepted for publication unless both the technical and grammatical revisions have been made successfully and the English has been polished.
3-Many of the references have been superceded. More modern ones are required.

Specific comments:
1.The word "novel" is used in the title of this manuscript. How do you reflect it?
2.Keywords lack of representativeness and need to be re-summarized.
3.In the Introduction section, it is too long and verbose, and needs further refinement. In addition, many of the references have been superceded. The authors should add more recent references.
4.“The current study encompasses the isolation of organochlorine (DDT and Aldrin) and organophosphorus (malathion) degrading bacteria from the vicinity of industries involved in producing biochemical pesticides.” This conclusion is confusing. Please revise it.
5.Acinetobacter baumannii has been reported to degrade other pesticide in previous study (doi: 10.3389/fmicb.2018.00098). Why the authors did not compare degrading efficiency of strain 5E with that of other strains of A. baumannii?
6.Line 36, “Aldrin” should be changed to “aldrin”. “Malathion” should be “malathion”. Please check the format throughout the paper.
7.The unit format should be uniform, such as line 50 “mg/ml”, line 180 “mg/mL” and line 410 “ mg L−1” and so on. Please carefully check it throughout the manuscript.
8.Line 135, “could use pesticides as their nutritional source”. Please explain clearly which pesticides can be used.
9.37℃ is too hot for most environmental bacteria. Why use 37 ℃ to screen bacteria in line 187?
10.Can Figure 4 be replaced with a high-definition electron microscope image？
11.What is the mean about “do” in Table 1
12.What is the optimum pH and rotation rate for these bacteria?
13.Line 172 and 179, ‘100 r/min’, ‘150 rpm’, The expression of the unit needs to be unified.
14.Line 186, Are you sure it's 1 ml culture?
15.Line 189, ‘48 hrs’, For standardization, it is recommended to use ‘hours’ or ‘h’.
16.Line 233, 15, 30 and 40oC.
17.In the Materials and methods section, please provide methods and equipment for the determination of three pesticides (DDT, aldrin, malathion).
18.Figure 1, 2, 3 and 6, need to increase the resolution of these pictures. I suggest that the author make these figures into broken-line figures to better express the results of the experiment.
19.Figure 4, need to increase the resolution of these pictures.
20.Figure 5, phylogenetic trees need to be redrawn to be more standard.
21.Line 412, [32][33]???
22.Line 421, [18]?
23.References: Many of the references have been superceded and more modern ones are required. The following papers belong to this topic, I hope they are useful for the authors. doi: 10.1016/j.chemosphere.2016.12.129; doi: 10.1016/j.jhazmat.2020.124927; doi: 10.1080/07388551.2020.1853032; doi:10.1016/j.chemosphere.2020.128827; doi: 10.1016/j.envres.2020.110660; doi: 10.3389/fbioe.2021.632059; doi: 10.1186/s12934-021-01556-9; doi:10.3389/fmicb.2019.01453; doi:10.1016/j.biortech.2017.12.007.


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: CHARACTERIZATIONS OF NOVEL PESTICIDE-DEGRADING BACTERIAL STRAINS FROM INDUSTRIAL WASTES FOUND IN THE INDUSTRIAL CITIES OF PAKISTAN AND THEIR BIODEGRADATION POTENTIAL
Review round: 2
Reviewer: 1

Output:
Basic reporting: This article is according to the standards of journal. I recommend to publish this article.
Experimental design: It is original, relevant and sufficient details are provided.
Validity of the findings: Conclusion are well stated and linked to original research.
Additional comments: I have found this article suitable and relevant for publication.


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

Input:
Title: CHARACTERIZATIONS OF NOVEL PESTICIDE-DEGRADING BACTERIAL STRAINS FROM INDUSTRIAL WASTES FOUND IN THE INDUSTRIAL CITIES OF PAKISTAN AND THEIR BIODEGRADATION POTENTIAL
Review round: 2
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: After going thoroughly through paper, I am completely satisfied with basic reporting, experimental design, and validity of findings. Moreover, my previous comments have also been addressed properly. So, I have no further comments to add up and should be accepted.


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: CHARACTERIZATIONS OF NOVEL PESTICIDE-DEGRADING BACTERIAL STRAINS FROM INDUSTRIAL WASTES FOUND IN THE INDUSTRIAL CITIES OF PAKISTAN AND THEIR BIODEGRADATION POTENTIAL
Review round: 2
Reviewer: 3

Output:
Basic reporting: .
Experimental design: .
Validity of the findings: .
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: CHARACTERIZATIONS OF NOVEL PESTICIDE-DEGRADING BACTERIAL STRAINS FROM INDUSTRIAL WASTES FOUND IN THE INDUSTRIAL CITIES OF PAKISTAN AND THEIR BIODEGRADATION POTENTIAL
Review round: 2
Reviewer: 4

Output:
Basic reporting: The authors have considered all comments raised by the reviewers and revised the manuscript accordingly based on these comments. The revision is fine and can be accepted for publication in current form.
Experimental design: The experimental design is good.
Validity of the findings: The findings is interesting.
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: INTEGRATIVE GROWTH PHYSIOLOGY AND TRANSCRIPTOME PROFILING OF PROBIOTIC LIMOSILACTOBACILLUS REUTERI KUB-AC5
Review round: 1
Reviewer: 1

Output:
Basic reporting: 1. For the “abstract” part, too many unnecessary details are mentioned. Mainly findings and conclusions should be listed here other than detailed experimental method and the process;
2. The counting unit should be unified of the whole paper, such as line 49-51, both for the description of KUB-AC5, CFU/g and CFU/ml are listed.
3.The language writing of the whole paper should be better polished.
Experimental design: Some explanations are missed which should be more cleared. eg. In line 88, why you start with the concentration at about 109 CFU/ml? Any supported reference or past experience?
Validity of the findings: Some details of the findings should explain more clearly.eg. In fig.1B, as mentioned, sucrose-based culture showed higher inhibition effect than glucose-based culture;
It seems there is a big turnover in the curve of “glucose at L-phase” at the time of 18-24h, is there any reason for this change? And, if the experiments last for a longer time, will the curve continue to go down? How about the other three?
Additional comments: The paper investigated how the transcriptome change after they found the phenotypes change by cultured with various carbon source. Further, they explored metabolic network and three main functional gene categories which were believed to be the reason of this change.
It is an interesting story standing on the inhibition effect difference of KUB-AC5. Still, here are some questions and comments:
1. Based on the findings of different phenotypes, an investigation of transcriptional change is announced. However, this logical progression is far-fetched. The change of transcriptome is a complex process which determined by various facts.
2. Three functional DEG categories were tested to explain the how the transcriptome changes and how it gives an impact on the phenotypes. Besides these three, have you considered any other macroscopic may also influence its gene expression? Are these three sites enough to result in this change?
3. There are too much quoted contents show in the “results discussion” part which is not common. They are supposed to be described in the “introduction” or “experiments design” parts.
4. Too much repeat description if “results” and “discussion”, the paper is lack of further discussion of how the phenotypes change results in transcriptome change and how is their feedback regulation between them.
5. Too much keywords listed, some of them are meaningless.


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: INTEGRATIVE GROWTH PHYSIOLOGY AND TRANSCRIPTOME PROFILING OF PROBIOTIC LIMOSILACTOBACILLUS REUTERI KUB-AC5
Review round: 1
Reviewer: 2

Output:
Basic reporting: See below
Experimental design: See below
Validity of the findings: See below
Additional comments: This article titled “Integrative growth physiology and transcriptome profiling of probiotic Limosilactobacillus reuteri KUB-AC5” reported the growth phases of KUB-AC5 strain in medium with two different carbon sources. The authors carried out a transcriptome analysis combined with growth physiology to predict global metabolic responses using metabolic network at a genome-scale of this strain. The subject is interesting and the data mining is proper. It can be of significance for the use of bioinformatics and omics to analyze the potential beneficial activity of probiotics. However, in this manuscript, I think there are still some problems about the inhibition effects of this strain to be solved.

1. The authors says “Samples were taken at different time points (i.e. 0, 3, 6, 9, 12, 15, 18, 21 and 24 h) for further viable cell count and pH measurement.” (line 91-92). But the pH changing with time is not mentioned in the result. This is very important, because the decrease of pH will also inhibit the growth of some pathogens.

2. The authors took the harvested cells to perform RNA-seq at 6h for L-phase and 12h for S-phase (line 113). Please explain why choose 12h for S-phase? The inhibition effects against S. Enteritidis S003 showed the largest difference appeared at 18h between Sucrose phase-S and Glucose phase-S or between Sucrose phase-L and Glucose phase-L (Fig.1B), while the differences between groups at 12h were less. Samples at this time point may result in a smaller set of differentially expressed genes, and may miss out on important candidate genes associated with the effect of S. Enteritidis S003 inhibition.

3. There is no statistical test in the results of KUB-AC5 strain growth characteristics and S. Enteritidis S003 inhibition (line 164-182). The corresponding P-value should be supplemented in the paragraphs and in Fig 1.

4. The authors used “cell-free culture supernatants (CFS) of L. reuteri KUB-AC5 from sucrose or glucose cultures at L- and S-phases were grown in nutrient broth for investigating the growth of S. Enteritidis S003” (line174-176). How does the S. Enteritidis S003 grow in ordinary NB medium?

5. The first paragraph of Discussion describes the research progress of probiotics in bacteriostasis. These contents should be moved into the Introduction part.


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: ROLES OF H2S AND NO IN REGULATING THE ANTIOXIDANT SYSTEM OF VIBRIO ALGINOLYTICUS UNDER NORFLOXACIN STRESS
Review round: 1
Reviewer: 1

Output:
Basic reporting: H2S and NO are important second messengers in organisms, which take part in many physiological processes. The manuscript (59645) reported that roles of H2S and NO in regulating the antioxidant system of vibrio alginolyticus under norfloxacin stress, which is import to understand the signaling role of H2S and NO. However, the following comments should be considered.
(1)	The abbreviations should be given the full names at the first occurring place in the text.
(2)	The H2S donor NaHS is a alkaline chemical, which might be affect the pH of the treatment solution, how to maintain the stability of pH.
(3)	How to define the units of enzymes.
(4)	Antioxidant enzymes usually are large family, which includes many members. In the text, the gene expression that is a member or a family for every enzyme is unclear.
(5)	Figs.2, 3 should be labeled significance or no significance.
(6)	Language should be carefully improved.
Experimental design: H2S and NO are important second messengers in organisms, which take part in many physiological processes. The manuscript (59645) reported that roles of H2S and NO in regulating the antioxidant system of vibrio alginolyticus under norfloxacin stress, which is import to understand the signaling role of H2S and NO. However, the following comments should be considered.
(1)	The abbreviations should be given the full names at the first occurring place in the text.
(2)	The H2S donor NaHS is a alkaline chemical, which might be affect the pH of the treatment solution, how to maintain the stability of pH.
(3)	How to define the units of enzymes.
(4)	Antioxidant enzymes usually are large family, which includes many members. In the text, the gene expression that is a member or a family for every enzyme is unclear.
(5)	Figs.2, 3 should be labeled significance or no significance.
(6)	Language should be carefully improved.
Validity of the findings: H2S and NO are important second messengers in organisms, which take part in many physiological processes. The manuscript (59645) reported that roles of H2S and NO in regulating the antioxidant system of vibrio alginolyticus under norfloxacin stress, which is import to understand the signaling role of H2S and NO. However, the following comments should be considered.
(1)	The abbreviations should be given the full names at the first occurring place in the text.
(2)	The H2S donor NaHS is a alkaline chemical, which might be affect the pH of the treatment solution, how to maintain the stability of pH.
(3)	How to define the units of enzymes.
(4)	Antioxidant enzymes usually are large family, which includes many members. In the text, the gene expression that is a member or a family for every enzyme is unclear.
(5)	Figs.2, 3 should be labeled significance or no significance.
(6)	Language should be carefully improved.
Additional comments: H2S and NO are important second messengers in organisms, which take part in many physiological processes. The manuscript (59645) reported that roles of H2S and NO in regulating the antioxidant system of vibrio alginolyticus under norfloxacin stress, which is import to understand the signaling role of H2S and NO. However, the following comments should be considered.
(1)	The abbreviations should be given the full names at the first occurring place in the text.
(2)	The H2S donor NaHS is a alkaline chemical, which might be affect the pH of the treatment solution, how to maintain the stability of pH.
(3)	How to define the units of enzymes.
(4)	Antioxidant enzymes usually are large family, which includes many members. In the text, the gene expression that is a member or a family for every enzyme is unclear.
(5)	Figs.2, 3 should be labeled significance or no significance.
(6)	Language should be carefully improved.


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: ROLES OF H2S AND NO IN REGULATING THE ANTIOXIDANT SYSTEM OF VIBRIO ALGINOLYTICUS UNDER NORFLOXACIN STRESS
Review round: 1
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: In this work, the authors evaluated the roles of H2S and NO in regulating the antioxidant system of Vibrio alginolyticus under norfloxacin stress. They revealed that anti-oxidative abilityin Vibrio alginolyticus was enhanced via the regulation of H2S and NO under norfloxacin (NOR)-induced stress, and H2S and NO may act synergistically to regulate the antioxidant system of Vibrio alginolyticus treated with NOR. The work is of interest of this journal readership and can be published.
And should address some comments as listed below:
1. The abstract section describes too much results, which should be reorganized the description structure.
2. The paper was poorly written and needs much editing to improve interpretation.
3. In the experimental part, the author did not cite any literature. What was the basis of the experimental operation?
4. The mechanism of H2S and NO on the antioxidant system of Vibrio alginolyticus should be illustrated.
5. The abstract should be rewritten to summarize the main research content.
6. To determine the interaction of H2S and NO, the author used SNP, PTIO and so on. How was the concentration and processing time determined? What was the influence of different solvent concentration and treatment time on the experimental results?
7. Please give the full name of the abbreviation for the first time use, such as MIC, SOD.
8. The following related papers on this topic maybe referenced for possible improving this manuscript:
Proteomic analysis reveals the protective role of exogenous hydrogen sulfide …., NITRIC OXIDE-BIOLOGY AND CHEMISTRY  Volume:  111   Pages:  14-30   Published:  JUN 1 2021
The vasodilatory mechanism of nitric oxide and hydrogen sulfide ….., EXPERIMENTAL AND THERAPEUTIC MEDICINE  Volume:  21   Issue:  3     Article Number: 214   Published:  MAR 2021
Imaging of anti-inflammatory effects of HNO via a near-infrared fluorescent probe ….., JOURNAL OF MATERIALS CHEMISTRY B  Volume:  7   Issue:  2   Pages:  305-313   Published:  JAN 14 2019
A highly selective turn-on near-infrared fluorescent probe for hydrogen sulfide ….., CHEMICAL COMMUNICATIONS  Volume:  48   Issue:  96   Pages:  11757-11759   Published: 2012
Changes in nitric oxide, carbon monoxide, hydrogen sulfide ….., FREE RADICAL BIOLOGY AND MEDICINE  Volume:  162   Pages:  353-366   Published:  JAN 2021
Nitric Oxide and Hydrogen Sulfide Coordinately Reduce ……, ANTIOXIDANTS  Volume:  10   Issue:  1     Article Number: 108   Published:  JAN 2021
Nitric oxide and hydrogen sulfide protect plasma ……, PLANT PHYSIOLOGY AND BIOCHEMISTRY  Volume:  157   Pages:  244-255   Published:  DEC 2020
Hydrogen sulfide (H2S) and nitric oxide (NO) alleviate cobalt toxicity ……, JOURNAL OF HAZARDOUS MATERIALS  Volume:  388     Article Number: 122061   Published:  APR 15 2020