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: PROXIMATE COMPOSITION, FUNCTIONAL PROPERTIES AND QUANTITATIVE ANALYSIS OF BENZOYL PEROXIDE AND BENZOIC ACID IN WHEAT FLOUR SAMPLES: EFFECT ON WHEAT FLOUR QUALITY
Review round: 1
Reviewer: 1

Output:
Basic reporting: Comment on language
Authors must check the English language and ensure that the paper is clearly written in standard, scientific English language, to make text easy to understand.
Example: lines 57-59 this paragraph is unclear, please consider rewriting, and how yellow color of WF can affect the dough??

suggestion:
All abbreviations must be clear in the first time in introduction
Line 94 : Please specify the genus and the species of wheat in the sample collection section even it is clear

Please change ml by mL in whole manuscript

Line 111 : Please change : described by Beuchat

Line 116 : check the form of reference : by Neto et al. (2001), the same in line 125 please check the form of all references as journal requirements.

Please check subscripts and superscripts in all calculation equation

Please include at the bottom of the Tables all the abbreviations included in the Tables

 Authors should add the signifiance difference in all tables
Experimental design: Methods are well described with sufficient detail in manuscript and with high technical standard

suggestion:
Authors were used samples from local supplier with additives and a control sample without additives under optimized field conditions. If WF is the control please specify in background.
Authors must use the same sample (seeds grown in the same conditions) with and without additives to estimate the effect of BP and BA.
The proximate composition included gluten, protein, fat, fiber was lesser for SF then WF but these results can be attributed to the different samples and not only to the traitement by BP and BA. Same comment for foaming capacity, gelation capacity and other analysis used.
How authors explain the choice of samples (control and samples with additives).
Validity of the findings: no comment on finding

Conclusion : lines 419-421 : repeated idea in introduction. The reviewer suggests that the authors conclude the current result importance on the food science and/or consumer’s health, as well as their future projection.
Additional comments: The authors evaluated nutritive composition and concentration of benzoyl peroxide and benzoic acid in the wheat flour samples. Apparently, there are no studies on the limite of BP and BA in wheat flour. Therefore, the results revealed superiority of WF over SF in  nutritive qualities and this is clear.

The manuscript is very concrete and with very complete information. Such study is important in the field of cereal science. Complete the study by estimate the dietary intake for BP and BA is very interesting and added more value to the manuscript. However, the reviewer wants to make some suggestions to improve its comprehension (cited above).


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: PROXIMATE COMPOSITION, FUNCTIONAL PROPERTIES AND QUANTITATIVE ANALYSIS OF BENZOYL PEROXIDE AND BENZOIC ACID IN WHEAT FLOUR SAMPLES: EFFECT ON WHEAT FLOUR QUALITY
Review round: 1
Reviewer: 2

Output:
Basic reporting: Line 41. Please add abbreviation (BA) after “Benzoic Acid” as first use
Line 75. … into BA, which is….
Line 81. Please put the tow references in the same brackets
Experimental design: Line 82-83. Explain why WF is not processed in the same way as SF, it would be highly interesting if the flours would have comparable treatments.
Please specify in “Sample collection” section that WF was whole wheat flour. Was the particle size of SF and WF characterized?
Line 117. Some instead certain
Line 118. Delete point after “min”, this is valid throughout the manuscript. Please specify the type of the used oil.
Line 131. Elsewhere, use h for hours (SI unit)
Line 141-142. Please give more details about crude fiber and fat determination
Line 189. Please add reference for Extraction Procedure for BPO and BA.
Statistics: you mention that the analysis were performed in triplicates. What is about the experimental setup, have the samples also been prepared in triplicates? Please indicate the significance level.
Validity of the findings: Authors state in materials and methods that Bulk density (BD) was measured and related results were showed in table 1, but not discussed.
Line 238. FC and FS of WF are more.
Table 1. Do not use % as a concentration unit - replace here by e.g. g/100 g or g/100 mL. Please use symbol “ρ” for bulk density instead BD.
Table 2. Please explain how gluten content was higher than crude protein content, knowing that gluten is an insoluble part of proteins.
Table 4. WF not WW
Additional comments: Manuscript titled " Proximate composition and quantitative analysis of benzoyl peroxide and benzoic acid in the wheat flour samples: wheat flour quality " investigates nutritive quality of commercial wheat flour (soft flour) by quantification of benzoyl peroxide added as bleaching agent in the SF, their proximate composition and functional properties.
There are many publications about benzoyl peroxide and benzoic acid in the wheat flour and their determination by HPLC, but topic is interesting and relevant for the field.


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

Input:
Title: PROXIMATE COMPOSITION, FUNCTIONAL PROPERTIES AND QUANTITATIVE ANALYSIS OF BENZOYL PEROXIDE AND BENZOIC ACID IN WHEAT FLOUR SAMPLES: EFFECT ON WHEAT FLOUR QUALITY
Review round: 1
Reviewer: 3

Output:
Basic reporting: The English language should be improved to ensure that an international audience can clearly understand the text. Some examples where the language could be improved include lines 44-46, 66, 78, 229-231, 251-253, 283, 338, 361, 386, 407, 408, 410.
Experimental design: The authors should pprovide more details in some methods (lines 139 and 141-142).
More details about statistical analyses should be provided.
Validity of the findings: The authors should state their conclusion about the dietry intake of BP and BA.
Additional comments: Lines1-3. We suggest to choice an other title for the manuscript.
Line 32. Please write “The objective”.
Line 37 and line 95. The authors write “commercial soft flour samples”. Please delete the word samples to avoid redundancy.
Line 37-40. In this section, authors should report all the tests performed in the current study.
Line 110. Please write “Water absorption capacity “.
Line 110. Please write “(WAC and OAC respectively)”.
Line 116. Please write “Emulsifying stability and emulsifying activity (ES and EA respectively)”.
Lines 137-172. This section should be reconsidered to avoid redundancy.
Lines 173-204. WE suggest changing the title to be “BP and BA quantification”. Also in this section, authors should indicate the references.
Line 207. Please indicate the gender and the age of the 200 subjects.
Line 214. What is the meaning of wt?.
Line 215. Which post-hoc test was performed?
Line 276. Please write “flour” instead of “floor”.
Line 318. Please write “parameter” instead of “component”.
Line 375. Please write “during” instead of “while”.
Table 1 and table 2. Please add standard deviation and letters to clearly identify differences between mean values.
Table 4 and table 5. Please add letters to clearly identify differences between mean values.


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: PROXIMATE COMPOSITION, FUNCTIONAL PROPERTIES AND QUANTITATIVE ANALYSIS OF BENZOYL PEROXIDE AND BENZOIC ACID IN WHEAT FLOUR SAMPLES: EFFECT ON WHEAT FLOUR QUALITY
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: PROXIMATE COMPOSITION, FUNCTIONAL PROPERTIES AND QUANTITATIVE ANALYSIS OF BENZOYL PEROXIDE AND BENZOIC ACID IN WHEAT FLOUR SAMPLES: EFFECT ON WHEAT FLOUR QUALITY
Review round: 2
Reviewer: 2

Output:
Basic reporting: The English language and style can be improved in the results section.
Experimental design: no comment
Validity of the findings: The conclusion should be reconsidered.
Additional comments: Lines 1-3. We suggest modifying the manuscript title to indicate the determination of functional properties of wheat flour in the current study.
Line 32. Please write “The objective” instead of “Objective”.
Line 37-44. In this section, authors should report all the tests performed in the current study (functional properties and proximate composition).
Line 46. Please delete the first sentence since it has been already reported in background section.
Line 64. Please write “the yellow colour”.
Line 95. Please delete “were”.
Line 114. Please delete “as”.
Lines 120-121. Please write “… were determined by the method described by Neto et al. (2001) with modifications”.
Line 130. Please write “the method reported by”.
Line 164. Please write “the method of”.
Line 183. Please write “cereal” instead of “clinical”.
Lines 199-200. Please rephrase this statement “About 8-10 g of sample in a sample bottle was added…”.
Line 206. Please write “BPO and BA were quantified in wheat flour as described by the method of Abe-Onishi et al. (2004).
Line 248. Please indicate the meaning of wt?.
Line 252. Please write “Tukey” instead of Turkey
Line 253. The authors reported a significant level of p <0.001 while others significant levels were reported in table 1 and table 2 (p < 0.05 and p < 0.01). Please indicate which significant level was used in statistical analysis.
Line 263. Please put “.” Instead of “:”.
Lines 264-339. The English language in the results section should be improved to ensure clear and professional English.
Line 400. Please write “Akintayo et al. (1999)”.
Line 451. Please write “mg/kg” instead od “mg.kg1”.
Lines 464-473. We suggest reconsidering the conclusion.
Table 1, 2, 3, 4 and 5. Please add letters to clearly identify differences between mean values.


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: PAN-CANCER SYSTEMATIC IDENTIFICATION OF LNCRNAS ASSOCIATED WITH CANCER PROGNOSIS
Review round: 1
Reviewer: 1

Output:
Basic reporting: reporting is generally good, some clarifications are needed.
Experimental design: the experimental design is sound and novel as it uses an ingenious way to use previously published microarray data.
Validity of the findings: the findings are interesting, albeit some clarifications are needed.
Additional comments: Review

The authors described a novel computational framework, in which they first applied the ARACNe framework to the TCGA pan-cancer RNA-seq data and calculated an essentially protein-coding-gene / lncRNA co-expression network based on mutual information theory (regulons). Next, they fit these regulons to the older microarray expression datasets and inferred the expression of lncRNAs in these microarray studies, which were not measure originally. By doing so, the authors were able to infer the lncRNA expression data for many of these older microarray-based studies, which also had longitudinal expression and clinical data for the patients.

This allowed the authors to conduct survival analysis on these lncRNAs and they identified a number of prognostic lncRNAs from these datasets. These lncRNAs were further grouped into “hazardous” and “protective”, according to the result of survival analysis. The authors then compared these lncRNAs with results from CRISPRi screen and found hazardous lncRNAs are enriched in “essential lncRNAs”.

Overall the work is quite interesting and the results are largely reasonable. I have the following comments that the authors need to address before the paper can be further considered for publication.

Figure 2B is shown as evidence that the regulon-based inference approach works in inferring the lncRNA expression level. It appears to me that the signal is stronger for certain cancer type than others. I would like to ask the authors to show the distributions individually for each cancer type, and comment on why the difference in the inference performance. Also, correlations are not the best indicator of accuracy, the correlations should be compared with random controls, i.e. what is the correlation between random pairs of predicted values and real measured values?

Figure 3A.
How is the p-value calculated (which is shown as a red horizontal bar) ? The red bar happens to intersect with the Y axis at about 0.05 too, is this a coincidence or did the authors actually meant Pearson rho cutoff at 0.05 ? The correlation for HOTAIR and XIST, 0.54 and 0.53. Are these high ? low ? compared to what ? I think the authors need to provide some perspective here.
Also, XIST is not the most appropriate lncRNA for such comparison since it is X-linked. Is the comparison only limited to female subjects ?

Figure 3B.
Again, if you take random pairs of predicted and experimentally measured lncRNAs and calculate Spearman correlation, are the correlations centered at 0.0 ?


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: PAN-CANCER SYSTEMATIC IDENTIFICATION OF LNCRNAS ASSOCIATED WITH CANCER PROGNOSIS
Review round: 1
Reviewer: 2

Output:
Basic reporting: Authors should mention that lncRNAs analyzed in the TCGA and microarray datasets are polyadenylated, and that there are many other lncRNAs which are non-polyadenylated, and  therefore, not analyzed in the present study. Indeed, majority of lncRNAs are non-polyadenylated.
Experimental design: Authors should perform general descriptions in the  Results sections about:

- ARACNe-AP algorithm
- BASE algorithm
- Cox regression z-scores
- Meta z-scores

Althought a deep description of this methods are explained in the M&M sections, brief explanations are necessary in the Results.
Validity of the findings: no comment
Additional comments: Manuscript describes a profound bioinformatic analysis in order to extract lncRNAs associated with poor outcome in different cancer types. I would like to see whether the lncRNAs with prognostic association are mapping into regulatory regions within the human genome. More specifically, how many of them are within enhancer regions? It would also be interesting to see whether the coding genes within lncRNA-regulons are located close to the corresponding lncRNA in the genome, so they could represent enhancer-promoter regulations, or sense (coding) to anti-sense (non-coding) interactions.


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: MASTICATORY MUSCULATURE OF THE AFRICAN MOLE-RATS (RODENTIA: BATHYERGIDAE)
Review round: 1
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: I have now read the paper by Cox and co-authors on the masticatory musculature of the African mole-rats (Rodentia: Bathyergidae) submitted to PeerJ. I found the paper interesting, the anatomical descriptions solid and the interpretation of the data meaningful and accurate. As such I mostly have minor suggestions to offer.

One thing I did find disappointing is that the authors did not report any data on muscle fibre lengths and pennation angles. This would make the manuscript much more useful for future studies interested in comparative functional anatomy or interested in creating biomechanical models which is one of the goals of the study. This is may only major comment on an otherwise excellent paper.

line 138 and elsewhere: the focus in the paper is on tooth digging which is fine. However, these animals are known to eat hard foods like roots and tubers which often are also of large size. As such having a large temporalis may also be of use in this context and may explain why the authors found no difference between the scratch digger and the tooth diggers. I would like to see this discussed/highlighted more in the manuscript.

line 326: I would like to have seen just a short comment on the fact that the authors had only a single specimen of each species in their data set. This prevents the authors from doing any statistical analyses testing for differences between tooth and scratch diggers and as such caution should be used in interpreting the differences.


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: MASTICATORY MUSCULATURE OF THE AFRICAN MOLE-RATS (RODENTIA: BATHYERGIDAE)
Review round: 1
Reviewer: 2

Output:
Basic reporting: No comment
Experimental design: No comment
Validity of the findings: No comment
Additional comments: This paper presents a detailed comparative study of the jaw adductor musculature of five extant African mole-rats (bathyergids), and the naked mole-rat, using diceCT. The relative size and attachments of the muscles were discussed in relation to chisel-tooth digging (4 of the 5 bathyergids plus the naked mole-rat) versus scratch-digging (1 species of bathyergid).
Overall, the paper is very well written, the descriptions of the muscles are very succinct, the figures are informative, and the discussion of the results is comprehensive. It is also good to see the scans are available on Morphosource. The discussion is hypothesis driven and clear, and the conclusions are supported. I particularly like the discussion of convergent forms between chisel-tooth digging mole-rats and other rodents like beavers that also use their incisors for demanding tree-felling, and the function of the rostral portion of the IOZM (or lack thereof) in bathyergids.
I really don’t have much to add for improvements; I found two minor grammar changes I would make, but that is more a preference (on Line 101 I’d remove “is they” and only have “…chisel-tooth diggers, that dig tunnels…”; and, on Line 232-233, I’d have “posterior to” instead of “posterior that of”).
In the methods, please also add which specimens were physically dissected on Line 178-179, and refer to these in the Results at Line 222 as well. If not all species were physically dissected, an explanation for this should also be given.
Otherwise, I’m happy to recommend this 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: CIRRIPECTES MATATAKARO, A NEW SPECIES OF COMBTOOTH BLENNY FROM THE CENTRAL PACIFIC, ILLUMINATES THE ORIGINS OF THE HAWAIIAN FISH FAUNA
Review round: 1
Reviewer: 1

Output:
Basic reporting: The paper is well written.
Experimental design: Solid.
Validity of the findings: Solid
Additional comments: It is always a pleasure to read a manuscript where your only role as a reviewer ends up being copyediting. I am completely convinced that this is a new species, and I agree with their morphological and molecular data.

I have a single substantive concern and maybe 56 copy-editing level concerns. My only real concern is that the type series is exclusively located in a single collection. This is frowned upon as all of the authors should know. This is particularly true given that the type material is exclusively near a country's capital. We don't have to look beyond Brazil to see that all material being located in a single collection is totally unacceptable (even if in a national collection). All journals, including PeerJ, should mandate that one of the type specimens is located in a second collection when the material is as common as seen in this study. It is difficult when someone describes a species including material from unaffiliated collections, but this study is submitted by the head collection manager of the relevant collection. I cannot understate how critical it is that the type series includes at least one specimen from a second collection; please transfer one specimen to a second institution.

Thanks again for the high-quality submission. Here are the copy-editing comments:

Throughout — the Oxford comma is not consistently used or not,

Title — should it say "the" Hawaiian fish fauna?

Second to last line of abstract, line 31 — combines should be combined

line 51 — sp. shouldn't be italicized.

line 54 — spp. shouldn't be italicized.

line 71 (most <100 mm SL) — should be followed by a comma

line 80 — should these numbers be written out?

line 83 — most references consider the common name to be a proper noun such that all words are capitalized.

line 117 — should this be fishes?

line 144 — comma should be removed

lines 149–152 — should this appear above the preceding paragraph because of the use of institutional codes in this preceding paragraph. Also why say NMNH here instead of USNM? Also, see line 155 where you given the National Museum as SECOND acronym. I get it, but how can the same place be abbreviated in two different ways. PICK ONE and use it consistently

lines 206–207, line 241, line 327, line 330 (x2), line 330–331, line 334, line 346 (x2), line 347, line 348 (x2), line 349 (x2), line 350 (x3), line 351, line 366, line 375, line 384  — normally ranges use an en-dash not a hyphen; please correct here and elsewhere

line 208, line 249, line 276, line 410, line 440, line 487, line 503, line 504 — it is usually frowned upon to use a scientific name as an adjective; consider a re-write

line 212 (first part) — here commas are used to separate the thousand levels, but they were not used in line 185 and 212 (second part); be consistent and follow the journal guidelines

line 302 – vessels are usually referred to as fishing vessels or research vessels

lines 320–323 — it is ALWAYS preferable to use a differential diagnosis where you provide the alternative states for the relevant taxa (congeners or congeners found in same location)

lines 339–340 — why use fin-rays or fin-spines here, but not on line 341?

line 354 — dorsal-fin spine (needs hyphen)

line 413 — why use a mononomial?

line 414 — needs a comma before the first and

line 416 — impractical is such a throw away; either delete or comment intelligently on its possibility or not using Sanger and high-throughput sequencing

line 445 — needs a comma before the and

line 451 — other species of what? Cirripectes, Blenniidae, Blennioidei, Percomorpha, Teleostei?

line 468–470 — needs a citation to support point

line 473 & line 483–486 — needs a comma before the but

line 504 — remove comma

line 526 — separate "and other recent studies" off by commas

Figure 2a lacks resolution in the version I can review

Figure 6 has filamentosus is misspelled

Figure 8 figure legend should say live not life

Again, I recommend acceptance after a specimen is transferred to a second institution and these copy-editing changes are made.


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: CIRRIPECTES MATATAKARO, A NEW SPECIES OF COMBTOOTH BLENNY FROM THE CENTRAL PACIFIC, ILLUMINATES THE ORIGINS OF THE HAWAIIAN FISH FAUNA
Review round: 1
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The authors of “Cirripectes matatakaro, a new species of combtooth blenny from the Central Pacific, illuminates the origins of Hawaiian fish fauna” present a well supported species description and add interesting phylogeographic comments. The authors do a nice job of setting up the interesting biogeography of Hawaii in the introduction. I am very impressed at the amount of work that went into this species description. There is a lot of research, knowledge and data gathering necessary to tease apart the relationship of a cryptic species, let alone within an already difficult group to identify (Cirripectes). A lot of care went into describing subtle differences among traits! I found the section on depth segregation rather interesting and wonder how common this is within closely related blenny species. I feel that the phylogenetic analyses might benefit from some updates (see species comments). One possible contentious point: the two fossils. Blenny fossils are exceedingly rare due to  habitat preferences and other factors. In my opinion there has been no reliable time calibrated blenniid phylogeny. There are several relatively whole body fossils (see below), but most are known from just otoliths. I am not a fossil expert, but I find it hard to believe that an otolith can be placed into a genus. Especially a small, relatively simple shaped otolith. I feel that the authors should remove the dating aspect since it is barely discussed and possibly inaccurate. If the time calibrated phylogeny is included I suggest a sentence describing the fossils.

Bannikov, A. F. (1998). New blennioid fishes of the familes Blenniidae and Clinidae (Perciformes) from the
Miocene of the Caucasus and Moldova. Paleontological Journal, 32(4), 385–389.
Yabumoto, Y., & Uyeno, T. (2007). Tottoriblennius hiraoi, a new genus and species of Miocene blennioid fish
from Tottori Prefecture, Japan. Bulletin of the National Museum of Nature and Science. Series C, Geology &
Paleontology, 33, 81–87.
L60-65: This section needs to be reworded and updated. I think Brandl recognized more families as
cryptobenthic… currently reads like there are 4. Use of “They” is a bit awkward. To avoid confusion refer to
blenniids as Combtooth Blennies and thereafter blennies.
L69: Perciformes is confusing and inaccurate. Please choose a different clade. Ovalentaria? See Betancur-R et
al 2017 or Near et al 2013
L72: Citation for Cirripectes as herbivores?
L77: Canines are on Premaxillary and dentary bones
L78-81: Williamsichthys is a named clade for Cirripectes sister genera
L104-114: I feel this can be moved to acknowledgments. NMNH abbreviation is used as before it is spelled out
L135: “In addition” used consecutively. Change one
L136: define nuchal flap
L146: BPBM abbreviations is used as before it is spelled out
L172: More accurately Exallias + Ophioblennius + Scartichthys is the sister clade to Cirripectes.
L176-187: was this data partitioned by codon? If not it probably should be, if so seems unlikely that
jmodeltest2 would select the same model for codon position 1,2,3 - especially a complex model. A poorly fit
model may be contributing to the number of poorly supported nodes in Figure 6.
L195: citation for pegas
L197-221: How many sequences (individuals) were used to represent each tip in BEAST analysis? Is it 1? IS this
because BEAST requires species designation? Seems like authors throwing a lot of data to the side, especially
since topology in Fig 6 is slightly different from Fig 7. Not sure how node at C. stigmaticus, variolosus,
casatneus in Fig. 6 is so highly supported. Is there a zero length branch at that node in Fig. 6?
L 320 – phylogenetic?
L334 (8): clarify if color in life?
L465-684: Critical point!
L580: Not sure if this citation is correct. I was unaware of this fossil and found a different citation… but not
sure which is correct.
L585, L663, L669: some parts of citation are in all caps
Figure 1: I think the figure would benefit from having the phylogeny (even cartoon version w/ fewer tips)
color/shape. Would help readers not as familiar with localities see where individuals/species were found.
Figure 2A: Photo color looks odd, but may be just a low resolution for this review version.
Figure 5B: It might help to label which species is lineage A and B


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: CIRRIPECTES MATATAKARO, A NEW SPECIES OF COMBTOOTH BLENNY FROM THE CENTRAL PACIFIC, ILLUMINATES THE ORIGINS OF THE HAWAIIAN FISH FAUNA
Review round: 1
Reviewer: 3

Output:
Basic reporting: This is a very well-written paper describing a new species of blenny from the central/south Pacific and placing it in an evolutionary context. Even though I am suggesting several changes in the comments to author box, these are all minor, with the possible exception of the removal of the phylogenetic analysis and correction of the molecular clock.
Experimental design: Research questions are clear and methods are appropriate, except for the phylogenetic analysis, which I think should be removed. In general, the data strongly supports the conclusions.
Validity of the findings: The authors make a strong case for recognition of Cirripectes matatakaro as a new species.
Additional comments: Minor suggestions:

Sentence in lines 35-36 (and line 514) suggests that Hawaii is the most isolated island in the world. I would change the sentence from "is the most isolated island chain in the world" to "is one of the most isolated island groups in the world". I know there are no other more isolated CHAINS but the readers might not know the difference between a chain and and archipelago and assume that you are saying that Hawaii is the most isolated archipelago in the world, which is not the case.

Line 37: Replace "Hawai'i hosts a highly endemic marine fauna, with the proportion of endemism among marine fishes having been recorded at 25%" with "a high rate endemism is found among the marine fauna of Hawai'i, with the proportion of endemics among fishes recorded at 25%".

Consistency: "Hawai'i" or "Hawaii". I don't have an opinion to which way to do it, but it's spelled as "Hawaii" in some places (line 25, 32, 552 are the ones I found), maybe pick one and use find and replace all.

Line 67: spell out eight (and every numeral that is not a count under 10).

On line 68, "The genus Cirripectes" should be the start of a new paragraph.

Lines 73, 74 need a citations for Cirripectes biology.

Replace "sister" with "closely related" on line 78. There should be only one sister genus, but may be several closely related. Same for line 182, Exallias is either "the" sister genus or "a" closely related genus.

Lines 154-173 could be reduced. If you are citing papers where we got the methods from, you don't need to specify the methods (i.e.: no need to write out primers and PCR conditions if they follow methods in Baldwin et al 2009, etc).

Lines 201-203, the 2% per million year rate presented by Bowen et al and Reece et al is for cytochrome B, not COI. The rate for COI is different, you have to use appropriate citations here and the appropriate rate for your marker. And the Bowen et al paper was published in 2001, not 2009. That's wrong on both the text and literature cited (which contain many other errors that I hope are corrected during the editorial process).

Lines 223-233, is this really necessary? I've never seen this statement in any other paper, including online journals.

Figure 5: please split the lineages by species (not lineage A and B). If they are split in the tree on Figure 6, they should also be split in the haplotype network by species (not lineage A and B under Cirripectes "variolosus").

I would delete lines 266-292 and analyses related to them. This adds very little to the manuscript and support for it is uncertain. First, the phylogenetic analysis are almost entirely absent from the discussion and conclusions, and second you can't really trust a phylogeny based on just one mitochondrial DNA marker, and this whole section does not contribute to the main subject of the paper (the new species description).

Paratypes: please deposit one or two paratypes in other institutions. Having them all in one collection (USNM) decreases access and increases risk.

Diagnosis: "based upon molecular phylogenic position" is not really a diagnostic character. Actually, I would delete lines 320-323 and leave the good diagnosis from lines 325-336.

First part of line 339 should be in methods.

Description: pectoral fin ray counts?

Color in life: Figure 8 shows a big difference between male and female colors, this should be discussed here.


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: CIRRIPECTES MATATAKARO, A NEW SPECIES OF COMBTOOTH BLENNY FROM THE CENTRAL PACIFIC, ILLUMINATES THE ORIGINS OF THE HAWAIIAN FISH FAUNA
Review round: 2
Reviewer: 1

Output:
Basic reporting: I just read through the changes, and I have no problems with the paper now.
Experimental design: I just read through the changes, and I have no problems with the paper now.
Validity of the findings: I just read through the changes, and I have no problems with the paper now.
Additional comments: I just read through the changes, and I have no problems with the paper 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: CIRRIPECTES MATATAKARO, A NEW SPECIES OF COMBTOOTH BLENNY FROM THE CENTRAL PACIFIC, ILLUMINATES THE ORIGINS OF THE HAWAIIAN FISH FAUNA
Review round: 2
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: I really enjoyed reviewing this paper and feel that it is a nice contribution to our understanding of Hawaiian fish fauna. I only have two comments for this round of reviews:
line 75 – herbivorous is misleading, unless defined as including detritus
Line 112 – define CRIOBE

Thanks and best wishes.


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: CIRRIPECTES MATATAKARO, A NEW SPECIES OF COMBTOOTH BLENNY FROM THE CENTRAL PACIFIC, ILLUMINATES THE ORIGINS OF THE HAWAIIAN FISH FAUNA
Review round: 2
Reviewer: 3

Output:
Basic reporting: This is a greatly improved version, the authors addressed all of my suggestions and I think this is ready to go.
Experimental design: Experimental design and methods are appropriate.
Validity of the findings: The new species is valid, and this is an important finding highlighting how little we know about even this widespread species (that was really two).
Additional comments: I think this is ready to be published after final checks. I agree with the authors that differential diagnosis should be in a comparisons section.


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: PREVALENCE AND TYPE DISTRIBUTION OF HUMAN PAPILLOMAVIRUS IN A CHINESE URBAN POPULATION BETWEEN 2014 AND 2018: A RETROSPECTIVE STUDY
Review round: 1
Reviewer: 1

Output:
Basic reporting: Please see my comment in "General comments for the author".
Experimental design: Please see my comment in "General comments for the author".
Validity of the findings: Please see my comment in "General comments for the author".
Additional comments: Dr. Lu and the research group investigated HPV infection over 5 years from a population of women living in an urban area of Beijing, China. A large sample size was assessed (>5000), with results indicating high prevalence of certain high risk HPV types (such as HPV52, 58, 16) in the surveyed area. As an epidemiological study, however, the study design, data presentation and analyses need to be improved before this manuscript could be accepted for publication.

1. The authors should have a diagram showing detailed subjective recruitment, such as criteria of inclusion and exclusion, total women recruited by year, follow-up size and the status of follow-up by year (e.g., drop out, re-visit...), available demographic information.

2. The authors should clearly indicate in abstract or title that serum but not cervicovaginal cellular swabs were used for HPV screening. To me, it is very important since HPV prevalence or type distribution is dramatically different between variable sample types. Meantime, discussion should be addressed for the difference in the manuscript since the authors compared prevalence and type distribution with other studies.

3. As shown in supplemental table 1 and the part of "outcome measurement", the authors used two different kits for HPV DNA screening. I realized, for the latest 3 years (2016-2018),  the majority of samples were tested with "fluorescence quantitative PCR" that covers 9 LR-HPV types while "PCR reverse dot blot" used for the first 3 years includes 3 LR types only. It concludes me that the increasing rate of LR HPV infection by year (Figure 2A) was probably not true due to in part the bias of detection methods for LR HPV types. Alternatively, the authors may focus on HPV6/11 alone to compare LR-HPV prevalence.

4. HPV is a kind of sexual transmitted infection which is closely related to life style, sex history, and marriage status. I don't have enough knowledge on the relationship between HPV infection and diabetes and/or hypertension, so the authors are recommended to justify the reason for such kind of statistical test, while the lack of key demographic information could largely impair the significance of this work.

5. I don't how to read Table 1. For example, Age (2014) - 43 is mean age? if so, what's meaning of 105 and 9.1 of the first part of diabets for example. How to define positive and negative status of diabetes and hypertension, and what's the proportion among the surveyed population?

6. Table 2, do authors have separated data between high-risk and low-risk HPV infection?

7. For Figure 3, the reviewer suggests to combine 2014-2018 with bar columns grouped by HPV types.

8. Please use "type" to replace "subtype" through the whole manuscript.

9. Line 40 - "All the HPV subtypes were detected". Please do not over-estimate HPV detection since only 12 HR and 10 LR HPV types were covered by the screening kits used in this study.

10. The English writing should be improved.

11. I feel confused about Supplemental tables 3 and 4.


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: PREVALENCE AND TYPE DISTRIBUTION OF HUMAN PAPILLOMAVIRUS IN A CHINESE URBAN POPULATION BETWEEN 2014 AND 2018: A RETROSPECTIVE STUDY
Review round: 1
Reviewer: 2

Output:
Basic reporting: The manuscript describes the prevalence and type-specific distribution of HPV infection in 5880 women over a 5-year interval. Authors also investigate persistence rate and coinfection in the population. It is quite interesting to see type-specific HPV distribution in this population, especially dominance of HPV52, while HPV18 is not even in the top five. Although the present manuscript has some major shortcomings, it would contribute to the current literature and thereby should be published.
1.	First, manuscript requires editing by professional/native English medical writer. Apart from grammatical mistakes throughout the manuscript, it is not always clear what authors are trying to say. In addition, I have some major concerns regarding the methodological approach and interpretation of the results/conclusions.
2.	The methodology of the HPV detection kits used is poorly described, could you please provide more details. I could not find information by the online reference provided. Furthermore, one of the links provided is accessible only in the Chinese language.
3.	According to the Supplemental Table 1, first 3 years mainly PCR reverse dot blot method was used, which was stopped in 2017 and replaced by fluorescence quantitative PCR. The first method detects 12 high-risk and three low-risk HPV types, while the second method detects 12 high-risk and 9 low-risk. It seems to me that the authors did not take into account this difference when interpreting the results. Significant increase in low-risk HPV detection appears when fluorescence quantitative PCR test was used frequently. You would expect this increase given that the first method is limited to detection of 15 HPV types, while the second could detect 21. It is necessary to adjust all analyses and results according to the detection abilities of the methods.  To identify the change in HPV prevalence you would need to limit the HPV detection to genotypes detectable by both methods. Similarly, decrease in HPV high-risk prevalence might be related to changes in HPV detection kits, even though both methods are able to detect 12 high-risk genotypes. I would like to draw the attention to the authors to tackle this in a detailed manner to limit the spurious findings that might arise with the methodology presented. In addition, these major limitations are not mentioned in the discussion.
4.	In regards to statistical analysis, the association between demographic and clinical variables cannot be studied with t-test or Chi2. These test just show if the groups are statistically different from each other, therefore more complex analysis would be necessary to study an association. Although, it is not clear to me how diabetes or hypertension might affect HPV infection incidence.
5.	HPV persistence and clearance is not well defined as well as how coinfection proportion. Could you please make it clearer?
6.	Could you please calculate median clearance and median persistence per type? Results presented in Table 2 are a bit confusing. In case you do not have enough data to show clearance/persistence by type, I would suggest grouping, for example according to phylogenetic distance (e.g. alpha 7, alpha 9).

Minor issues:
Abstract: Line 40 authors stated, “all the HPV subtypes were detected”. What do you mean by that?
Does “subtype” mean HPV type, or could you detect subtype/lineage of particular HPV type?
Please, use word type or genotype when referring to the HPV types.

Introduction:
Line 60-61: There is no evidence of an increased risk of cancer when infected with multiple HPV genotypes.
Line 68-71: 12 high-risk genotypes are mentioned, but only 13 are listed. HPV66 is classified as possibly carcinogenic by IARC.
Methods:
Line 97: Did you also exclude women with cervical cancer? If that is the case, how many women were excluded? Since this is HPV prevalence study, I would suggest keeping women with cervical cancer cases. Did participants receive a pap test? It would be interesting to see the impact of HPV incidence/persistence on pap-test results.
Line 123: Was any other clinical or demographic information collected?
What is the reason for women coming to the hospital? Do they have a health issue at admission or it is just a check-up? Were women invited for the examinations?
Do you have any information on HPV vaccination of young women participating in this study?
Why did you switch to fluorescence quantitative PCR test?
How was hypertension defined?
What was the median follow-up time?
How many participants received more than one examination?
Results: Line148-154, Please see comment 3 above. Did you take into account multiple testing when reporting HPV prevalence?  Re-testing HPV positive women would have an impact on reported incidence.
Line 155-160: Again, see comment 3 above. The first kit can detect 15 genotypes, while the second can detect 21. You would need to limit these results to 15 genotypes detectable by both methods, or report separate results for each kit used.
How was the coinfection proportion calculated? Please define in methods.
What was HPV acquisition rate? The proportion of participants negative at baseline and positive at follow-up?
Line 167-172: It is a bit difficult to understand HPV persistence results. May be reporting median clearance and persistence time in months would make it easier for the reader.
Line 169:
Discussion.
Line 181: Confounders cannot be studied with the statistical tests described in methods, see comment 4.
Line 184-193: See comment 3 above.
Line 241: I have doubts regarding the generalizability of the results of the current study. It is not clear to me why these women are attending the hospital, are they invited, or they attend due to previous health issues? If they are invited, what is the participation rate? Moreover, as you mentioned above in the discussion, HPV distribution is largely affected by many confounders and varies across the areas.
Line 253: You need to address the issue of changes of the detection methods in more details and how this would impact the results.
Since the first method was not used from 2017, I assume there are some participants tested with the first method and re-tested with the second. Could you please discuss this fact? Would this affect HPV persistence results?
Experimental design: See above
Validity of the findings: See above
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: PREVALENCE AND TYPE DISTRIBUTION OF HUMAN PAPILLOMAVIRUS IN A CHINESE URBAN POPULATION BETWEEN 2014 AND 2018: A RETROSPECTIVE STUDY
Review round: 1
Reviewer: 3

Output:
Basic reporting: - The authors described the prevalence of HPV DNA and their subtypes in a population of China with very robust number of specimens. It is important to mention that HPV prevalence still a concerning since there is no HPV vaccine which covers all HPVs associated with cervical cancer.
- I really recommend updating the introduction with respect to IARC classification. It is important to mention that some HPVs are also classified as possible or probably HR, which still are necessary evidences of their oncogenesis (based on molecular or epidemiological evidences). The author says 12 HR-HPVs but 13 was written, please, re-check the literature (PMID: 30963559 and others)
- There is no changing in HPV DNA prevalence during the years, and this should be better addressed in the discussion.
- The English grammar is good.
Experimental design: - Regarding the study design and participants, I strongly recommend rewriting it. Please, explain better the number of participants included, and the samples. How was collected and stored?
- The authors say that Viral DNA was isolated from serum samples, however, only few studies have reported presence of HPV DNA into the bloodstream of patients. This must be justified and clarified and very well discussed since could strongly impact the results. Due to this fact, I recommend changing the Title, including the type of specimen used.
- Should be clarified in the text and compared the HPV prevalence with studies that used the same technique and the limitations of this study.
Validity of the findings: The descriptive results are well addressed.
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: PREVALENCE AND TYPE DISTRIBUTION OF HUMAN PAPILLOMAVIRUS IN A CHINESE URBAN POPULATION BETWEEN 2014 AND 2018: A RETROSPECTIVE STUDY
Review round: 3
Reviewer: 1

Output:
Basic reporting: I have no further comment.
Experimental design: I have no further comment.
Validity of the findings: I have no further comment.
Additional comments: I have no further comment.


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: PREVALENCE AND TYPE DISTRIBUTION OF HUMAN PAPILLOMAVIRUS IN A CHINESE URBAN POPULATION BETWEEN 2014 AND 2018: A RETROSPECTIVE STUDY
Review round: 3
Reviewer: 2

Output:
Basic reporting: The manuscript has significantly improved, although I still have some concerns which have not been addressed by the authors.
As I mentioned in my previous review the prevalence of low risk genotypes has increased due to the change of the genotyping assays, it is not an increase in prevalence but simply increase in detection due to the broader spectrum assay used. Therefore, authors should carefully revise the interpretation of their results and discussion.
For example line 169. The statement regarding the significant difference of lr-hpv positivity rate between 2014 and 2018 is misleading and should be removed. This difference is expected due to the dissimilarity of the assays. The comparison would make sense if authors show the prevalence of low risk genotypes covered by both methods such as HPV6 and 11.
Similarly, lines 180-182 authors state that HPV positivity was zero in 2014 for some low risk HPV and increased over time. One would expect such results given that the reverse dot blot method can detect only HPV6 and 11. Please revise or remove the sentence.
Line 210: I would suggest explaining in details that study results showed increased detection rate of lr-hpv rather than increased prevalence, since the fluorescence quantitative PCR targets 9 lr-HPV types compared to 3 lr-HPV with reverse dot blot method.
The discussion would benefit if authors justify why they investigate an association between diabetes/hypertension with HPV infection and interpret results.
Minor changes:
Could authors please specify target region/gene of the assays.
Please briefly mention in abstract detection methods used.
Please address minor grammatical errors throughout the manuscript e.g:  line 71, line 91, line 162.
Line 112 and 121 please indicate years. Authors should clearly explain in methods which test was used during which period.
It is still confusing for me how was the persistence rate calculated. Could authors please revise the paragraph.
Some sentences still remain unclear especially in the discussion part e.g. lines 245-247.
Tables and figures:
In table one please indicate in a footnote that the numbers are median and frequencies as in reply to reviewer one comment 5.
Experimental design: see above
Validity of the findings: see above
Additional comments: see above


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: PREVALENCE AND TYPE DISTRIBUTION OF HUMAN PAPILLOMAVIRUS IN A CHINESE URBAN POPULATION BETWEEN 2014 AND 2018: A RETROSPECTIVE STUDY
Review round: 3
Reviewer: 3

Output:
Basic reporting: The authors improved the paper with minor revision regarding english grammar.
Experimental design: The authors compared risk factors of diabetes and hypertension with hpv presence. Would be great if the authors could explain why the compared with these variables, and what the results means. It can be explained in the discussion or results.
Validity of the findings: The results are important in order to keep the screen of hpv in a population where lack these informations.
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: PREVALENCE AND TYPE DISTRIBUTION OF HUMAN PAPILLOMAVIRUS IN A CHINESE URBAN POPULATION BETWEEN 2014 AND 2018: A RETROSPECTIVE STUDY
Review round: 4
Reviewer: 1

Output:
Basic reporting: The manuscript has been improved significantly.
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: PIGLEG: PREDICTION OF SWINE PHENOTYPE USING MACHINE LEARNING
Review round: 1
Reviewer: 1

Output:
Basic reporting: This manuscript, to the reviewer, is well-organized and easy to read. The use of English is acceptable. Literature review is also enough and relevant.
Experimental design: This study fits the journal well. The research question is well-defined. The data collection process needs more explanations.
Validity of the findings: The proposed model might be overfitting. This should be solved.
Additional comments: This study is interesting for adopting machine learning approaches to predict leg weakness and lameness of pigs. Their findings can have important practical implications. Several suggestions are provided for the authors.

1.‘Materials and Methods’ section: How predictors are selected for building the model should be explained.
2.‘Materials and Methods’ section: It is suggested to present the operational definitions and measurement scale of predictors used in this study. Further, how these predictors are collected should be delineated as well. If data collection involves human judgment, what are the qualifications of those raters? And also how to ensure the reliability of those collected data?
3.‘Materials and Methods’ section: Is there any particular considerations for choosing these nine machine learning algorithms?
4.‘Data Preparation’ section: Regarding class imbalance issue, it is suggested to explicitly express what method (under- or over-sampling) is used to solve the issue instead of saying ‘…using the ROSE package…’
5.‘Model training’ section: This study randomly chose 10% of the sample for the final assessment, nonetheless no assessment results was demonstrated in this study.
6.‘Results’ section:  The proposed model is suffered from ‘overfitting’ because the prediction accuracy of train data is higher than that of test data (lines 238-239). This issue should be solved or the proposed model is unable to predict new data accurately.


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: PIGLEG: PREDICTION OF SWINE PHENOTYPE USING MACHINE LEARNING
Review round: 1
Reviewer: 2

Output:
Basic reporting: See below
Experimental design: See below
Validity of the findings: See below
Additional comments: The paper can be accepted but it needs to be corrected and updated. The comments are:

a)       There are many grammatical mistakes such as:

Pork is the most widely consumed meats in the world

b)      Description of the data balancing techniques used is not given

c)       These also exist some classifiers whose performances are least affected by class imbalance. Comments on the that is also required.

d)      The reason of using a method for filling the missing values needs to be quoted. And also it needs to mentioned that who effective their approach was?

e)      The significance of Kappa value (Table 1), Friedman rank (Table 2) along with the other metrics needs to be mentioned. Also it needs to be mentioned the relevance of these tests w.r.t. these results.

f)        The description of the data used needs more details

g)       Figure 1 is confusing w.r.t. RF and KNN classifiers

h)      Figure 2 and figure 3 are not readable, quality of these figures need to be improved

i)        More emphasis can be given on the proposed methodology and the conclusions of this work


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

Input:
Title: PIGLEG: PREDICTION OF SWINE PHENOTYPE USING MACHINE LEARNING
Review round: 2
Reviewer: 1

Output:
Basic reporting: No comment.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: The authors have addressed reviewers' 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: SUBSTRATE RUGOSITY AND TEMPERATURE MATTERS: PATTERNS OF BENTHIC DIVERSITY AT TROPICAL INTERTIDAL REEFS IN THE SW ATLANTIC
Review round: 1
Reviewer: 1

Output:
Basic reporting: Overall this manuscript suffers from some lack of clarity in writing. However, the biggest issue with this manuscript is that I am not convinced the findings are novel in any way. The fact that sea surface temperatures and rugosity impact benthic biodiversity and community structure has been demonstrated for many ecosystems from the intertidal to the deep sea. The data appear robust and the study design is sound. I simply encourage the authors to find a more compelling question to answer. For example, can their information be used to establish predictions of baseline community structure, diversity, or richness predictions based on rugosity and SST patterns (or extremes, or anything else) that managers could then use to determine if another stressor is impacting the community? Why did the authors first want to test if lateritic reefs were structured by different abiotic factors- is there a reason they thought they might be?
Experimental design: The study design seems relatively sound. However, the spatial scale over which the explanatory variables were averaged is unclear. Also, the unit of replication at each site is unclear; at one point the authors state that each 0.5 m2 quadrat was used as a replicate but later in the same paragraph they say each transect was used as a replicate. Either way, but particularly if the individual quadrats were used as the unit of replication, the data from within a site at each sampling event should be tested for spatial autocorrelation. Finally, I wonder why the authors chose to assess rugosity in one 2 m2 patch instead of over each transect or at each quadrat. Finally, the analysis presented in the methods does not perfectly match the response variables reported in the results. This should be addressed.
Validity of the findings: With the caveats of methodological details highlighted in the previous section I do not have any reason to doubt the validity of the findings. I do doubt their novelty and impact on the field. The discussion of the paper focusses mostly on what is already known about rugosity and SST shaping benthic communities and does not do a good job of highlighting what this paper contributes to this body of knowledge. The authors claim, in the first paragraph of the discussion, to have developed ecosystem-based indicators for this system. They did not do this. An ecosystem based indicator is a value that can either be measured or calculated from a set of measurements in the ecosystem that, when tracked over time, tells you information about the health or state of the ecosystem. The authors did not develop a new ecosystem-level indicator nor did they, as the work is currently presented, assess the validity of a proposed indicator.
Additional comments: The authors have gathered a large amount of, what appears to be high quality data. I encourage them to rethink their approach and use this data to address a question that is more interesting to a larger 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: SUBSTRATE RUGOSITY AND TEMPERATURE MATTERS: PATTERNS OF BENTHIC DIVERSITY AT TROPICAL INTERTIDAL REEFS IN THE SW ATLANTIC
Review round: 1
Reviewer: 2

Output:
Basic reporting: 1. Consider results of GUICHARD, F.; BOURGET, E.; ROBERT, J. L. Scaling the influence of topography heterogeneity on intertidal benthic communities: alternate trajectories mediated by hidrodinamics and shading. Marine Ecology Progress Series, 217, 27-41. 2001.

2. Figure 3 have some too small letters.
Experimental design: 1.	The authors sad that “sites are similar in their presence of lateritic substrate, easiness of access, geographical orientation, exposure to wave action, geomorphological features, and anthropogenic pressure”. Provide some data or reference on that.
2.	Although the studied area is in a marine protected area, there are populated areas that could disturb the marine community under study. The authors should make clear whether the reefs are really comparable or whether different degrees of impact between reefs could cause some of the differences found.

3.	How the transect were positioned on each reef? Randomly?
4.	“A representative intertidal area of 2m2 in each site was photographed” – how these representative area were chosen?
Validity of the findings: no comment
Additional comments: The study brings relevant information on marine biodiversity at tropical reefs in the South Atlantic and its relationship with reef rugosity and sea surface temperatures. Some minor questions are pointed out.


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: SUBSTRATE RUGOSITY AND TEMPERATURE MATTERS: PATTERNS OF BENTHIC DIVERSITY AT TROPICAL INTERTIDAL REEFS IN THE SW ATLANTIC
Review round: 2
Reviewer: 1

Output:
Basic reporting: This paper still needs to be streamlined and better supported. I still feel the introduction does not set up the study well- by the end of it I still have no idea why the authors did the study. There are a couple instances of ambiguity in the language used and many instances where the language is general and vague where specifics would be helpful. I have pointed these out in the detailed comments I have provided with this review. Additionally, in the discussion the authors make several claims of causation where their data only support correlation. Again, these are pointed out in the comments I have provided. Finally, the authors only state two hypotheses, but there are 6 tables and 6 figures. This seems excessive. Particularly as some data is presented in both table and figure form.
Experimental design: Experimental design is sufficient.
Validity of the findings: The authors do not do a good job of putting their specific findings in the context of what is known. In the discussion there are very few instances where specifics of the studies findings are mentioned. When they are statements implying causation are used where only correlation is supported by the study.
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: SUBSTRATE RUGOSITY AND TEMPERATURE MATTERS: PATTERNS OF BENTHIC DIVERSITY AT TROPICAL INTERTIDAL REEFS IN THE SW ATLANTIC
Review round: 3
Reviewer: 1

Output:
Basic reporting: Overall the paper is greatly improved. There are still some issues with clarity of writing- largely in the introduction.
Experimental design: It is fine.
Validity of the findings: Much improved.
Additional comments: No additional comments


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

Input:
Title: COMPARATIVE FUNCTIONAL GENOMIC ANALYSIS OF ALZHEIMER’S AFFECTED AND NATURALLY AGING BRAINS
Review round: 1
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: This study describes transcriptomic changes that occur with aging and correlates these changes with AD pathogenesis. Using available microarray databases, Peng et al. mined the AD and Aging microarray results using a comprehensive method to analyze genes involved in AD and aging. Regional differences in differentially expressed gene categories were observed in aging/AD brain. Of interest, the authors observe perturbations in genes related in microbial infection in aging and AD. It is highlighted that the authors performed interacting gene network (IGN) analysis and the extended IGNs to establish a larger view of the transcriptomic landscape in aging/AD.

As written, the manuscript requires some revision. More specifically, the data/figures need to be reorganized and presented to enhance clarity, and the data requires further interpretation and analysis to reinforce the main conclusions in this study.

Major issues:

1. As the authors also mentioned in the “limitation” section, mining results from published microarray data is potentially informative. However, the manuscript presents an abundance of data/figures, but draws few or no conclusions from the analysis. Further clarification, interpretation would enhance the submission.

2. Providing the DEG, IGN, and xIGN in each dataset would provide a good resource to the research community. However, some data refinement may be beneficial: for example, the authors only consider the significance/FDR cutoffs, but fold change was not used to filter the data. Most of the results were generated based on these filtered gene sets, where fold change has not been factored into the results.

3. As described in the methods and Supplementary Figure 2, and Supplementary table 1, the authors selectively retrieved data from the original data sets, which may introduce some bias in the study. It is known that heterogeneity of individuals is a big concern, artificial omission of data in this study requires careful and a detailed explanation.

Minor issues:

1. Line 132 and 136, what is the criteria to selectively analyze the top 106 genes from the AlzGene gene set and top 109 genes in AlzBase gene set?

2. Some of the results were very vague as described. For instance, in line 203-204: “The overall ratio of downregulated to upregulated genes was about 3:1”, which is confusing. The groups compared need to be described.

3. Supplementary Table 1. GSE5281 and GSE53890 should be switched.

4. It is recommended to use several Venn diagrams to show overlap [in categories] rather than overlapping number of genes among DEG, IGN, and xIGN, and between these 6 gene sets (5 AD regions and 1 Aging case).

5. Please carefully check all the citations. Several references were missed, for instance, the source of the aging data set is from GSE53890 (PMID: 24670762) from Lu et al., 2014 have not been cited. The same for GSE5281 (PMID: 17077275, 18332434, 29937276). Since these studies also used the same microarray data, their results need to be compared to the current study. It is nice work to show the connection of OXPHOS and AD, which is one of the highlights in this manuscript. Please discuss more OXPHOS in AD (PMID: 20217279, 31257151), these two articles include the role of OXPHOS in tau pathology and in microglia. Line 400-401, NDUFS3 is also associated with Parkinson’s disease (PMID: 28062948).


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: COMPARATIVE FUNCTIONAL GENOMIC ANALYSIS OF ALZHEIMER’S AFFECTED AND NATURALLY AGING BRAINS
Review round: 1
Reviewer: 2

Output:
Basic reporting: The data and background information is outdated.
Experimental design: Too small dataset size and current practices are geared towards reporting the biological processes affected.  Individual genes are not that interesting from an in-silico perspective.
Validity of the findings: Nothing new in the findings.  Reporting individual genes without wet-lab validation on such outdated datasets that have been heavily worked on is just re-inventing the wheel without anything substantially new to reveal.
Additional comments: Please consider using new datasets with greater sample size or implementing techniques to increase the power of small datasets or new methodologies.  The 21 novel genes refer to biological pathways already indicated to be affected in AD.  I do not see exactly what is the contribution of this research.


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

Input:
Title: ICARIIN AFFECTS CELL CYCLE PROGRESSION AND PROLIFERATION OF HUMAN RETINAL PIGMENT EPITHELIAL CELLS VIA ENHANCING EXPRESSION OF H19
Review round: 1
Reviewer: 1

Output:
Basic reporting: In our opinion, language is acceptable, and the manuscript properly refers to the literature. We request some modifications, as follows:

Please, avoid using the terminology “dose” while working with cells.

Figure 1. The authors claim a concentration-dependent antiproliferative effect of ICA. However, there is no evidence of a statistical difference between each concentration used. Also, Fig 1 A and B must be cited in the results section, though they refer to the same description in the text.

Figure 4 legend merits revision. It neither matches the figure nor the results described on page 11, lines 160-164.
Experimental design: This study falls within the scope of the journal, and the methods are described in sufficient detail. Critical modifications to improve the quality of this study and to increase the potential readership by the journal audience are as follows:

The study does not describe the kind of cell death ICA induces. In our opinion, the antiproliferative assay must be associated with further experiments to indicate the cell death pattern. (for instance, please see DOI:10.1016/j.ejphar.2018.11.001).

What morphological alterations does ICA induce? Does H19 reverse them?
Validity of the findings: There is no citation/description of Figure 2 in the text. Regarding the same figure, the authors indicate that ICA suppressed the cell cycle progression. However, there is no evidence of such a result. Cells on G0/G1, S or G2 phases seem to be quantitatively the same despite the treatment. Are RPE cells susceptible to ICA? What is the duplication time of those cells? We believe the results do not support the conclusions concerning the cell cycle. An example of an altered cell cycle is found in DOI: 10.2174/1566524017666171106115655.
Additional comments: This study investigated the inhibitory effects of the flavonoid icariin (ICA) on retinal pigment epithelial cells against platelet-derived growth factor (PDGF)-BB-induced cell proliferation. Additionally, it explores the influence of H19 on ICA-induced antiproliferative effect. In our opinion, the manuscript is descriptive. How ICA induces cell death pattern merits investigation. We point out some concerns.


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: ICARIIN AFFECTS CELL CYCLE PROGRESSION AND PROLIFERATION OF HUMAN RETINAL PIGMENT EPITHELIAL CELLS VIA ENHANCING EXPRESSION OF H19
Review round: 1
Reviewer: 2

Output:
Basic reporting: - The article was written in clear, unambiguous, technically correct text.
- Literature references was appropriately referenced.
- The structure of the article was conform to the journal acceptable format.
- The results were relevant and clarify the hypothesis, but some terms have been possibly used inappropriately.
Experimental design: - The methods were described with sufficient detail and information to replicate and were appropriately chosen to respond to the research hypotheses.
Validity of the findings: No comment
Additional comments: 1. Interestingly, the IC50 values must be shown in Fig. 1.
2. Was the MTS assay performed to evaluate cytotoxicity against ARPE-9 without estimulation of PDGF-BB? What is the IC50 value?
3. The cytotoxicity of ICA should be evaluated against other cell lines in order to evaluate its selectivity.
4. It is interesting to clarify if ICA induced a cytotoxic or cytostatic effect against ARPE-9 cell line. MTS is not the most appropriate method to clarify this. I suggest that you use SRB assay if possible. It is important to evaluate a possible cell cycle arrest relationship with induction of cell death. It would be interesting?
5. In addition, the statistics need to be added to cell cycle data in Fig. 2 and Fig. 5. Could the raw data be sent?
6. In the Line 158 – ICA increased or decreased CDK4, CDK6 and Cyclin D1 protein levels? In Fig. 3 B-C a decrease in CDK4, CDK6 and Cyclin D1 protein levels by ICA was observed.
7. The effect of ICA on PDGF-BB-stimulated RPE cells always returns to basal levels of unstimulated RPE cells. Would it be appropriate to use the terms "upregulate" or "down-regulate" when it comes to the effects of ICA on PDGF-BB-stimulated RPE cells?


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: ICARIIN AFFECTS CELL CYCLE PROGRESSION AND PROLIFERATION OF HUMAN RETINAL PIGMENT EPITHELIAL CELLS VIA ENHANCING EXPRESSION OF H19
Review round: 1
Reviewer: 3

Output:
Basic reporting: 1. The work presented is important and well written, however with a few textual inconsistencies and some minor errors in English, which should be thoroughly reviewed.
2. Throughout the text the term dose could be replaced by concentration.
Experimental design: 1. I would like to know about the choice of concentrations used (10, 20 and 40µM).
2. Flow cytometry tests did not show the amount of events acquired.
3. Western blot and PCR tests also did not show the amount of sample loaded.
Validity of the findings: Figures 2 and 5. The authors do not show the values or standard deviation bars, as well as the asterisks that symbolizes statistical differences in cell cycle evaluation.
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: ICARIIN AFFECTS CELL CYCLE PROGRESSION AND PROLIFERATION OF HUMAN RETINAL PIGMENT EPITHELIAL CELLS VIA ENHANCING EXPRESSION OF H19
Review round: 2
Reviewer: 1

Output:
Basic reporting: No further comments.
Experimental design: No further comments.
Validity of the findings: No further comments.
Additional comments: Thank you for the rebuttal to the comments we raised.


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

Input:
Title: ICARIIN AFFECTS CELL CYCLE PROGRESSION AND PROLIFERATION OF HUMAN RETINAL PIGMENT EPITHELIAL CELLS VIA ENHANCING EXPRESSION OF H19
Review round: 2
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The authors answered all the questions satisfactorily


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 COMBINED EFFECTS OF TEMPERATURE AND AROMATASE INHIBITOR ON METAMORPHOSIS, GROWTH, LOCOMOTION, AND SEX RATIO OF TIGER FROG (HOPLOBATRACHUS RUGULOSUS) TADPOLES
Review round: 1
Reviewer: 1

Output:
Basic reporting: The paper is generally well-written and understandable.  However, there are problems with tense and there are areas where the language is awkward making it difficult to read smoothly.  For example the first line of the Introduction, line 46, there should be a modifier before ‘attention’, such as recent or much etc. and Line 89, ‘restrain the transformation’ ‘prevent’ would be a better word.  The use of ‘E.g’ is problematic; the information would be better incorporated into a sentence.

Line 79: Whereas it is true that there has been quite a lot of work with steroidal hormones and amphibians it is ALSO true that there has been quite a bit of attention to the effects of aromatase inhibitors – contrary to what the authors state there is quite a bit of work with Xenopus and Silurana e.g. Olmstead et al. 2009 https://doi.org/10.1016/j.aquatox.2008.07.018, and Duarte-Gutterman et al. 2009  https://doi.org/10.1159/000280586

There appears to be an emphasis on Asian authors in the cited literature.  Which is reasonable when referring specifically to the study organism, but may be limiting the breadth of the available information on the topic of aromatase inhibitor effects on phenotypic sex in general.

The authors should tell the reader why they chose letrozole instead of the more commonly known aromatase inhibitors (e.g. tamoxifen).  It is particularly interesting that letrozole has a very specific mode of action (https://link.springer.com/article/10.1007/s10549-007-9696-3) and is more potent and specific for AI and is non steroidal.

Lines 91-95: Consistent with the title, the authors state clearly what the study entails.  However, the last line of the Introduction leaves me wondering what the connection is between the two.  I think this linkage of the effects of exposure to such a potent AI and conservation are a bit obscure and need explanation.  Alternatively, remove the sentence.

Figure 1 has no SE; and shouldn’t it be + not just +?  Legends for Figs 2 and 3 are switched.  Would be helpful if there are no significant differences to state same.  Are survival and % male means?  Check language in legends.
Experimental design: There is no replication of the experimental units which is unfortunate and makes the results less rigorous.  This should be addressed in the discussion.

Line 100 -104: The use of plastic in which to conduct the experiments is of concern to me.  Recall that the topic of endocrine disrupting chemicals was in part ‘discovered’ by Ana Soto who observed her breast cells growing abnormally in plastic culture dishes.

Line 105: It is not clear how 1.8 tadpole density was maintained through metamorphosis…were mortalities replaced?

Line 111: “the feed..was only 100 mL …ethanol” needs to be reworded.

Line 118:  how was the weighing of the tadpoles performed?  Every X days they weighed 90 tadpoles in each treatment?  Removed them to a tared beaker?

Line 123: It is not clear to me what happens with each tadpole that metamorphosed.  Were the froglets put into individual lidded bowls or grouped?  How many of the 90 froglets/tadpoles were tested for jumping; just 10? Just the first 10? It appears that your results are for 10 but there are data for 20 in your supplemental data.  I’m confused.  Please address this. Moreover, the supplemental data for the jumping results are only the means?  Can you please provide the 3 distance measurements?

Line 128: is there a protocol that can be cited for the jumping test? If not, could it be explained in some more detail?  It is not clear whether the froglets are jumping within the bowl or whether they are removed.  Also, are they returned to a start position for the three jumps or are they probed to jump from where they landed from the previous jump?

Line 130: new paragraph at “The froglets were reared…”

Line 135:  what is an appropriate amount of food?  So, were froglets placed in pond after metamorphosis directly from jumping experiment? From exposure vessel? When did 90 days start? From beginning of exposure in plastic vessels; it says 90 days of feeding, I don’t understand how you could ensure 90 d for each froglet if they metamorphosed at different times?  What happened at the end of the experiment? Were the ponds drained and the frogs recovered?  How did they ensure all were recovered?  Were the ponds enclosed to ensure that no frogs could escape?

Line 144: Define metamorphose rate?  Did you move the tadpoles that had not metamorphosed into the pond when you moved others that had?  I cannot follow the sequence of events.

How do the temperatures of 29c and 34c compare to the temperatures the frogs are naturally exposed to in their native habitat?  Defend/explain why these were chosen.
Validity of the findings: The authors state that the exposure treatment did not affect metamorphosis rate nor survival rate.  But I am not sure that this is accurate. I believe the authors have determined that there is no difference in metamorphosis rate nor survival rate AMONG the treatments.  However, did they actually test to determine if the rates were different from 100% metamorphosis or survival?  If they did the latter, then I can believe that there was no effect of temperatura or letrozole on survival or metamorphosis rate. (also see Line 188)

Could the authors please address the cause of the high mortality? The high mortality could have compromised the results.  Is there any relationship between those tadpoles that did not metamorphose and those that died?  What sex were the tadpoles that died?  Could there have been a bias such that only or more females than males died?

Line 186: The authors indicate that their results show indirect support for sex hormones not involved in growth or development.  This needs to be put into context within this early developmental window and at physiological concentrations.  Outside of this window and at exposure to concentrations of estrogens or androgens greater than physiological concentrations there may in fact be effects on growth and development.

Line 195: The authors implicitly suggest they anticipated an AI to increase testosterone concentrations, which would increase the proportion of males to females and muscle development so that the frogs would jump further. On line 146 the authors considered SVL relative to jumping distance and did not observe a relationship.  However, they did not consider mass or show the results of the relationship to jumping and mass.  Lighter frogs may have been able to jump further, and there were effects of temperature on size.

Also, did the authors consider measuring testosterone and estradiol in the frogs?

Line 222:  instead of accumulation, use additive. And instead of overlap use interact.  Replication would have helped with defining this better.

Line 227: Statements such as ‘considered for the first time’ are unnecessarily grandiose when in the context of doing an experiment such as this where the AI effects on phenotypic sex of an amphibian generically have been explored and the only novelty is really the use of a different species  I would strongly encourage the authors to leave this statement out so as to not turn off the reader.

Line: 232: “letrozole can induce a male bias in the tadpoles of H. rugulosus, and this effect is increased at high temperature” in Lines 170-175 I do not see that the proportion of males at letrozole and 34C are significantly higher than letrozole and 29C.  Is this the case?  Or is it that there was a sig difference in proportion between 29 and 34. And within temperatures sig diff between 0 letrozole and 0.1 and 1 treatments but not between temp x letrozole treatments?  Do the authors know when the thermosensitive period is for this species?
Additional comments: No additional comments


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

Input:
Title: THE COMBINED EFFECTS OF TEMPERATURE AND AROMATASE INHIBITOR ON METAMORPHOSIS, GROWTH, LOCOMOTION, AND SEX RATIO OF TIGER FROG (HOPLOBATRACHUS RUGULOSUS) TADPOLES
Review round: 1
Reviewer: 2

Output:
Basic reporting: .
Experimental design: .
Validity of the findings: .
Additional comments: “Effects of temperature and aromatase inhibitors on metamorphosis, growth, locomotion, and sex ratio in Hoplobatrachus rugulosus tadpoles”
General comments:
This is an original study focusing on the combined effect of temperature and an aromatase inhibitor over several aspects of growth, development, sex determination and performance of tadpoles and metamorphs of the anuran Hoplobatrachus rugulosus.

The study is well presented, however I prompt the authors to include more information linking why to study the effect of these two stressors (Temperature and aromatase inhibitor) in the context of this species (that is at same degree of risk). Also, considering the proposed framework they need to define more clearly the objectives of the study. For example, ending the Introduction I suggest authors to better present the objectives. Why to study the effect of aromatase inhibitor (letrozol) and temperature? How is the connection with conservation strategies in this species of high commercial value?
The experimental design is appropriate, it result clear and the analysis is well done.
Respect to the validity of the results, they are conclusive and clear.
Discussion in general is correct, but again, I suggest to focus and discuss after the objectives were clearly proposed.
Finally, be careful with grammar and punctuation.

Particular comments:
Introduction.
In the first paragraph, if authors are talking about amphibians in general, would be adequate to cite more general literature related to the effect of temperature and hormones on life history responses in amphibians.
Ending the Introduction, authors need to define the objectives of the study and link them with appropriate framework of study. Then, based on previous studies and the question/s authors propose here I suggest to better explain what is the purpose to use letrozol, Is it a common compound? Can it emulate responses in typical hormones? And how common is the use of this kid of substances in these studies? Then, it is not clear (both in the framework presented in Introduction and Discussion) why to study the effect of this aromatase inhibitor (letrozol) and temperature. As I suggested before, the connection between the effect of these stressors and the conservation status of this species need to be better explained.
In particular, experimental design: What was the criteria used for the set of experimental temperatures (29° and 34°)? These temperatures are common in nature? Or, It is related to settings at amphibian hatcheries? It must be better explained.
Discussion
Lines 186-188. This paragraph merely needs have connection with objectives. For example, authors suggest that letrozol results could be extrapolated to the effect of steroid and testosterone hormone as is observed in other studies, however, again I think it is needed to present the idea in the Introduction to connect with objectives and to be suitable discussed.
Lines 190-194. To discuss this result (discrepancy about the temperature range employed in this study) it is necessary to understand how the set of experimental temperature was determined.

Figures and legends.
There is a confusion between Fig. 2 and Fig. 3 and their legends. Please correct it.


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 COMBINED EFFECTS OF TEMPERATURE AND AROMATASE INHIBITOR ON METAMORPHOSIS, GROWTH, LOCOMOTION, AND SEX RATIO OF TIGER FROG (HOPLOBATRACHUS RUGULOSUS) TADPOLES
Review round: 2
Reviewer: 1

Output:
Basic reporting: revision is acceptable
Experimental design: revision is acceptable
Validity of the findings: revision is acceptable
Additional comments: revision is acceptable


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 COMBINED EFFECTS OF TEMPERATURE AND AROMATASE INHIBITOR ON METAMORPHOSIS, GROWTH, LOCOMOTION, AND SEX RATIO OF TIGER FROG (HOPLOBATRACHUS RUGULOSUS) TADPOLES
Review round: 3
Reviewer: 1

Output:
Basic reporting: This is an original study focusing on the combined effect of temperature and an aromatase inhibitor (letrozole) on several aspects of growth, development, sex determination and performance of tadpoles and metamorphs of the anuran Hoplobatrachus rugulosus.

My main concerns are focused on the objectives and how the discussion was addressed.
Experimental design: My main concern is on the scientific questions of the study. It is not clear why the use of litrozole, Is it a common substance found in nature? or Is it commonly used in the industry of frog-farms? Then, I could not find the link with temperature and the interest of the study.
Validity of the findings: My suggestion is that authors need to explain how your results relate to expectations (Was letrozol a disruptor of life history patterns? Is there an interaction with temperature? Can you explain the importance of the results on the conservation status of the species? And for the economic purpose?), to the literature (including more cites) and how they are important in the field of conservation of the species.
Additional comments: Review
“Effects of temperature and aromatase inhibitors on metamorphosis, growth, locomotion, and sex ratio in Hoplobatrachus rugulosus tadpoles”

General comments:
This is an original study focusing on the combined effect of temperature and an aromatase inhibitor (letrozole) on several aspects of growth, development, sex determination and performance of tadpoles and metamorphs of the anuran Hoplobatrachus rugulosus.
My main concern is on the scientific questions of the study. It is not clear why the use of litrozole, Is it a common substance found in nature? or Is it commonly used in the industry of frog-farms? Then, I could not find the link with temperature and the interest of the study.

Then, my main concerns are focused on the objectives and how the discussion was addressed.

Particular comments

Line23-25: Please correct the text to:
In the present study, H. rugulosus tadpoles were subjected to two water temperatures (29 °C and 34 °C) and three letrozole concentrations in feed (0 mg/g, 0.1 mg/g, and 1 mg/g) to examine the effects of …

Line 31. Please, Could you specify the difference between body size and condition? I do not find a diference.

Line 34. Again, what is a condition factor? There some words and statements (“condition,” “condition values” and “condition factor”) that need to be explained. In the section Data measurement “condition factor” is well explained, then I suggest to include here a kind of similar explanation to put the reader in context.

Introduction
Lines 45-53 I suggest that authors need to connect the use of letrozole in the questions that they are proposing. Is letrozole common in nature? It is a problem to this protected species?

Lines 53-54. Please include citations.

Line 66, 105. Please correct, the cite is. Hayes, 1997

Lines 119-124. I suggest that authors need to connect the use of letrozol in the questions that they are proposing. Is letrozole common in nature? It is a problem to this protected species?
Lines 250-252. I think this conclusion must be taken with caution.

Discussion
In the first paragraph authors present the relationship between temperature and life history responses, and I found that literature cited is insuficient. I suggest to include more general literature as e.g Stearns work (Stearns, S. C. 1992. The evolution of life histories.
Stearns, S. C. and J. C. Koella. 1986. Evolution and some others). There is a lot of literature demonstrating that temperature increase development and result in smaller froglets. Then, I suggest authors to consider include more references.

In general Discussion needs to be reorder. Organize the Discussion from the specific to the general.
In the Introduction authors pay attention on the economic value of this anuran species having in account the context of their conservation status.
I think the main problem with this manuscript is that objectives are not clear enough and there is no relation between the results and there is no a clear what are the implications of the findings.

Authors need to clarify what is the objective to use letrozole and why temperature is also important in the interaction with this aromatase inhibitor. Is letrozole common in nature? The use in the study has the purpose to mimic other aromatase inhibitors? Is it commonly used in frog-farms?
Once the objectives are clear try to connect the results in this sense to answer the questions posed in the Introduction. I recommend to explain how the results support the objectives (once they are clarified) and answers proposed and, how these answers fit in with existing knowledge on the topic. By the way, because of the abundant published work related to this topic I suggest to include more general cites (not just recent and regional research) to discuss the results.
Also, authors need to explain how your results relate to expectations (Was letrozol a disruptor of life history patterns? Is there an interaction with temperature? Can you explain the importance of the results on the conservation status of the species? And for the economic purpose?), to the literature (including more cites) and how they are important in the field of conservation of the species.


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: ALTERED PROTEOME OF A BURKHOLDERIA PSEUDOMALLEI MUTANT DEFECTIVE IN SHORT-CHAIN DEHYDROGENASE AFFECTS CELL ADHESION, BIOFILM FORMATION AND HEAT STRESS TOLERANCE
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript is relatively well written, the figures are well presented . There are no significant omissions of data.
1. There is a utilisation of Et al. which is not italicised throughout the text and should be corrected.
2. lines 86-87 this sentence should be re drafted as it seems incomplete.
3. Line 92. differential proteins should be replaced with ' differentially expressed proteins'
4. Line 94. the word 'conditions' should be removed.
5. Line 112. dissolved to be replaced with resuspended.
6. Lines 126-127. 'The percentage volume was used for quantification'. Not sure what the authors mean by this?
7. Line 150. Remove the word conventional. Could replace with standard.
Experimental design: The study describe phenotypic assays carried out to assess the results of knocking out BPSS2242, coding for a short-chain dehydrogenase.
Proteomic analysis by Q-TOF MS/MS of the knock out and the wild type indicated 5 differentially abundant proteins. This is a straight forward phenotypic characterisation study so the main question appears to be in establishing if the selected assays adequately demonstrate and support the conclusions.
Figure 1 shows the proteomic profile of a WT and SDO mutant strain. It could be argued that more accurate quantitative approaches such as iTRAQ could have yielded more data but this is not necessarily a cheap technique.
Figure 2. Indicates fold changes in gene expression. This figure is adequate, however, I note that a complement of SDO was made. So why is this not included?
Figure 3. A small set of assays to compare wt, mutant and complement.

Overall, the experiments have been carried out to an acceptable standard , although the expression data from the SDO complement should be shown.
Validity of the findings: The findings are acceptable and not overstated. The conclusions that SDO modulates the expression of several proteins are supported by both proteomic and genetic assays. The authors describe that SDO has been shown to be an important virulence determinant in P. aeruginosa also. However, has any linkage between SDO and regulation of ANY other gene ever been demonstrated in other bacteria?
Also, the finding that expression of the Hsp70 family protein was down-regulated is interesting, but what is the relevance of testing resistance to a temperature as high as 50C, this seems too extreme.
Additional comments: A well written and presented study. The work is robust and the data supports the conclusions.
In my opinion there is just 1 omission that must be further explored. The gene expression data in the SDO complement of the 5 downregulated genes must to be demonstrated.