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: COMPARISON OF THE FECAL MICROBIOTA OF TWO FREE-RANGING CHINESE SUBSPECIES OF THE LEOPARD (PANTHERA PARDUS) USING HIGH-THROUGHPUT SEQUENCING
Review round: 2
Reviewer: 1

Output:
Basic reporting: note: numbering of lines applies to the reviewed manuscript without track-changes

INTRODUCTION
L56-57: Rephrase sentence "North Chinese leopard originally distributed North and Central China while lost as much as 98% of their historic range." to clarify. While? -> but
L62: North Chinses -> North Chinese
L64: supported -> supporting
L65: Amur Leopards -> Amur leopards; north Chinese -> North Chinese
L69-L70: Rephrase sentence "Gut microbial diversity analyses based on leopard fecal samples should also be taken into account conservation efforts" to clarify. Should conservation efforts take into account microbial diversity analyses or vice versa?! I believe you refer to the first option but that does not come from this sentence.
L77: remove"richness" as it is encompassed in the term "diversity"
L78: characterized with -> under

METHODS
L121: traces like footprints -> footprint traces
L121: each transects -> each transect
L122: intervals -> interval
L123: was contributed to -> contributed to
L176: to detecting -> to detect

RESULTS
L193-194: were also contributed to -> contributed also to
L198: remove 'also'
L209: The Boxplots -> The boxplots
L218: remove 'accurately' and 'here'
L221-L224: Rephrase full sentence starting with "According to PcoA..." to clarify. What do you want to say exactly? What do you mean by scattered within groups? Due to the use of different visual representation of data, the key message tends to get a bit lost.

DISCUSSION
L228-230: Please rewrite the sentence, with attention for use and tense of verbs. Grammar in that sentence is not correct. "Amur and North Chinese leopards are endangered flagship species..."
L230: were -> are
L259: north Chinese. Since -> North Chinese, since
L261: North Chinses -> North Chinese
L271: include -> including
L275: north Chinese -> North Chinese
L275: (Lubbs et al., 2009) -> include ref in a sentence without brackets when used as a subject, so Lubbs et al. (2009) suggest ... (check author guidelines for in-text use of references)
L276: domestic cat -> domestic cats
L277: the C. perfringens -> C. perfringens
L288: were -> are
L293: Please rephrase, it is not sure that these diseases cause the dysbiosis or are the result of a dysbiosis.
L296: add a verb for the second part of the sentence
L299: plays -> play
L299-L301: Adds unnecessary information to the discussion. The role of Proteobacteria for lignin digestion is not relevant for a strict carnivore.
L301: remove "plus", not a scientific phrasing
L303: which phylum do you refer to with "this phylum"? Proteobacteria? Moreover, this sentence is too long and confuses since C. difficile belongs again to Firmicutes. Please rephrase more clearly and shorten.
L305: samplings -> sampling
L306: remove 'certain' because this is too vague
L307: refine your statement from "wild species" to "some" wild species, because for some species there is already a lot of information available.
L308: the role of bacteria in gut health/function or host health? Please remove "gut microbiology"
L309: remove 'was'
L310-L311: Remove redundant sentence " the relative abundance of..." because this is just a bulk sentence with no novel information or specific information
L327: Please rephrase and avoid the verb 'related to'. Simply by just mentioning the functions of Bacteroides species.
L329-L331: Please rephrase end of this sentence. Grammar is not correct. "anaerobic bacteria which beneficial the host"?
L333-L334: Rephrase "above interferences" and I suggest you reconsider the statement that more fecal samples are required. Consider more the type of information you can get from a 16S diversity study compared to more functional analyses or metagenomic analyses! You don't need necessarily more samples to find out more about the microbial community but a different type of analyses. Especially when you reference work that points out functions of bacterial species.
L349-L352: I don't think your analyses can support this statement. Please consider carefully. Diversity analyses have been mainly discussed at the phylum level, the dietary impact might manifest at lower taxonomic levels and/or only be assessed on functional analyses. Moreover, specific dietary regimes and nutrient intake for the animals included in the study have not been recorded.
L356-L358: Many of the reported variables (sex e.g.) are not unmeasurable! They have just not been recorded for the animals included in your study and or require a more defined set of descriptors (e.g. biomarkers for health, age ranges based on certain anatomical or physical or physiological characteristics, ...)
L358: Is it the number of samples that is the limiting factor? Consider the analyses performed. Will you gather necessarily more useful and "in-depth" information with just more samples?
L361: Please remove "in the hope of acquiring" ... It is evident that scientific studies have as an outcome "to acquire more knowledge by gathering new and/or more data".
L361-L364: Please consider this statement carefully and check the most recent literature. High-throughput sequencing technology can already provide much more in-depth information than provided in this study and the technology in itself should not be scrutinized as such.
L365: Please consider the use of "normal" as it puts forward the question highly discussed "what is a normal host-associated microbiota?!"
L369: remove "using high-throughput sequencing" and "after analyzing 21 wild feces samples'; this is redundant info
L373-L374: specify the taxonomic level for this statement
L378: remove "into the prospective research direction"
L378: include -> including
Experimental design: Comments on experimental design are incorporated in the 'Basic Reporting' section.
Validity of the findings: While the introduction and discussion section have improved, the authors still lack to incorporate a critical reflection of the results in the light of leopard conservation. Which aspects of leopard conservation can benefit from this study? How can leopard conservation be addressed via the study of the gut microbiota? e.g. Likely functional analyses and dietary analyses will provide more inside. But the introduction also highlights the impact of habitat fragmentation on gut microbial communities of species living in the wild but this is not mentioned again in the discussion. Wouldn't studies considering both gut microbial diversity and ecological and anthropogenic factors benefit in-situ conservation? Which type of conservation (ex-situ or in-situ) can benefit from microbial analyses? Diet-gut microbiota relations (as suggested in the discussion by mentioning prey type) might be of bigger importance in captive settings (ex-situ conservation) where a diet is an important management tool.

Relative abundance levels at the phylum level between leopards and other species have been discussed, but the authors should finalize this manuscript with a reflection of these data in the light of conservation.
Additional comments: Thank you for your re-submission and addressing in detail most of the comments and suggestions from the first review. The manuscript has improved. Please go carefully over this second manuscript review which includes some remaining grammatical comments and critical remarks about some of the statements made.
Please consider the advice in section 3 (Validity of findings). The discussion section could be shorter in the comparative description about relative abundances of phyla and include more interpretation of these results in light of conservation of leopards.


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: COMPARISON OF THE FECAL MICROBIOTA OF TWO FREE-RANGING CHINESE SUBSPECIES OF THE LEOPARD (PANTHERA PARDUS) USING HIGH-THROUGHPUT SEQUENCING
Review round: 2
Reviewer: 2

Output:
Basic reporting: The manuscript “Comparison of the fecal microbiota of two free-ranging Chinese subspecies of the leopard (Panthera pardus) using high-throughput sequencing” by Han et al. is a resubmission of a manuscript I reviewed previously. It describes a comparison of the gut microbiota of two subspecies of leopards using faecal samples as a surrogate. The gut microbiome of carnivores is still not well understood today and the current study adds to that limited knowledge. The study has some merit and the resubmitted version is already much improved in terms of quality of the figures and clarity of the text. Grammatical issues remain in the text but do not impact clarity. I would like to see the work published after my remaining issues with the figures are clarified.
Introduction:
The introduction is now better tailored to the results of the study.
Methods:
Lines 157—162: What version of the Silva database was used? How were the phylogenetic trees inferred after aligning the sequences with Muscle?
Results:
I still think it would be beneficial if the authors provided their annotated OTU table as supplementary information so that readers can investigate taxa of interest.
The results section feels disconnected from the figures that it should be describing.

Figures:
Figure quality was improved overall.
Figure 1:
As previously mentioned, the legend describes what rarefaction and rank abundance curves are. This is not relevant. The figure legend should briefly summarize the findings. This is also true for some of the other figures.
Figure 3:
a)	The figure or the legend should state that the values shown in the heatmap are z-scores. The authors explained this in their rebuttal letter but choose to omit this information from the figure.
b)	I still do not understand why phylum relative abundance is different between weighted and unweighted Unifrac. Please explain this to me and the readers and double check your data. As no OTU table is provided I am unable to interrogate the data myself.

Figure 4:
The last sentence “The x-axis and the y-axis indicate groups and the indices of different diversity respectively. “ is not required.

Figure 5:
The red-green colour scheme is still not ideal.
Discussion:
The discussion now is more relevant to the results of the paper.

Conclusions
I am fine with the conclusion except from the following statement. “We infer that the same tendency of fecal microbiota might mainly result from genetic factors of leopard.” I do not think this can be inferred from the data as such. You may be able to speculate this but it is more likely the lack of significant differences is caused by the small sample size and too much variability within the groups.
Experimental design: The design is typical for studies of fecal samples from the wild. Limitations such as freshness and the limited number of samples are inherent and have been addressed.
Validity of the findings: The findings are observational. As I do not have access to the data I am unable to verify them.
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: CLINICOPATHOLOGICAL CHARACTERISTICS, TREATMENT AND SURVIVAL OF PULMONARY LARGE CELL NEUROENDOCRINE CARCINOMA: A SEER POPULATION-BASED STUDY
Review round: 1
Reviewer: 1

Output:
Basic reporting: 203: please fix the typo"respectable"
214: The authors keep referring that there is no standard treatment for pulmonary LCNEC. This is not true. There is no standard treatment for metastatic LCNEC and oncologists often use either NSCLC or SCLC chemotherapy regimens but early-stage pulmonary LCNEC is treated akin to NSCLC. Surgery and chemoradiation are standard treatments for Stage I-III pulmonary LCNEC.
229: Authors state that surgery and radiation are the favorable prognostic factor. This is expected since its the early stage disease that receive either surgery or chemoradiation and hence the prognosis is better. This needs to be mentioned or elaborated in the discussion. Surgery and radiation are in a way surrogates for early-stage disease in this analysis, but since authors did not analyze TNM staging this data is unavailable.
196: authors state that majority pulmonary LCNEC patients were either east coast or west/pacific coast residents and were Caucasians. This needs to be clarified since coastal areas are where the majority of the population lives. Was this data adjusted for population density? Similarly, the majority of the US population is Caucasian, can we get a comment on the incidence of LCNEC based on the race which is adjusted for population?
Experimental design: Experimental design is good.
Validity of the findings: Analysis is straightforward. Authors have used appropriate statistical tools.
Additional comments: The grammar and flow of article can, in general, be improved. Would recommend getting the manuscript proofread by a native english speaker.

Will compliment authors for taking up this project, review of pulmonary LCNEC epidemiological data is important in spreading awareness about this rare disease and to help foster therapeutic interventions. Will definitely advocate in favor of accepting this manuscript 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: CLINICOPATHOLOGICAL CHARACTERISTICS, TREATMENT AND SURVIVAL OF PULMONARY LARGE CELL NEUROENDOCRINE CARCINOMA: A SEER POPULATION-BASED STUDY
Review round: 1
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: In this manuscript, the Authors aim to investigate the clinical pathological characteristics, treatment and survival of pulmonary large cell neuroendocrine carcinoma. Using the SEER database data on 2556 patients were obtained. They found that LCNEC have a higher incidence in elderly people, males, late stage and peripheral localization. They also identified 9 variables that can be easily obtained in multivariate analyses.
1. The SEER is very poor database to use as the clinical detail in this administrative database is somewhat poor. Many factors are lacking which can only be obtained in institutional databases.
2. Just using “surgery” as the predictive variable does not well inform clinical practice. What type of surgery needs to be specified.
3. TNM staging is a very important prognostic factor for the prognosis of patients with LCNEC, however, the author did not take it into account.
4.The manuscript would likely benefit from copy edits since there are several misused and misspelled words. Attached Non-Native Speakers of English Editing Certificate should be attached.
5.The authors did not clarify how they dealt with missing data, particularly for the predictive variables.
6.Please report 3 and 5-year CSS.
7.Table 2: Please also report HRs for univariate analysis. How HRs were calculated? This was not reported in the manuscript.
8.Methods. There are 3 registries subgroups (SEER9, SEER13, SEER18). It is not clear to me the specific database used by Authors. ICD-O-3 codes used for SEER database query should be specified in a supplementary table, along with their explanation.


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: CLINICOPATHOLOGICAL CHARACTERISTICS, TREATMENT AND SURVIVAL OF PULMONARY LARGE CELL NEUROENDOCRINE CARCINOMA: A SEER POPULATION-BASED STUDY
Review round: 1
Reviewer: 3

Output:
Basic reporting: (1)	The total survival curve was not provided.
(2)	Table 2 is redundant since most information can be found in Figure 2.
(3)	The survival time in Figure 2 should better be with an interval of 12 months and patients at risk should be added.
(4)	There are some grammatical errors, such as in line 84, “This program contain” should be “contains”.
Experimental design: (1)	The authors investigated patients with LCNEC between 2000 and 2013 in SEER databases. However, SEER database recorded their data according to different modes as years went by. For example, the 6th and 7th TNM stages were provided in records after 2004 and 2010, respectively. The authors can restage the patients into the newest 8th TNM stage manually by carefully checking the records in recent years. They should better analyze the prognostic significance of T, N, M, and TNM stage instead of the old stage classification (local, regional, and distant).
(2)	The chemotherapy status, a quite important variable associated with prognosis, is also now provided in SEER database. The authors can visit the website (https://seer.cancer.gov/data/treatment.html), obtain the chemotherapy status and use it in the survival analyses.
(3)	Differentiation, marital status, and insurance were also frequently reported to be correlated with the prognosis, but not included in the analyses. The left or right lobes were not analyzed, either.
(4)	It is not appropriate that age only was divided into <60 and ≥60. Patients at the age of 80 couldn’t have a similar prognosis as those at the age of 60. The authors should better classify the age with an interval of 10 years.
(5)	The authors declared that 80.7% LNCEC patients were with peripheral localization, which is incorrect. The tumor was located in the upper, middle, or lower lobe did not mean it was peripheral. Peripheral lung cancer refers to lung cancer derived from respiratory bronchioles.
Validity of the findings: (1) The authors should better validate their results using their own records or data of other public databases.
Additional comments: In this manuscript, Dr. Gao et al. investigated the clinicopathological features, treatment, and survival of patients with lung large cell neuroendocrine carcinoma (LCNEC) using SEER data. They found that the survival of LCNEC was not well, and LCNEC was associated with elderly, male, peripheral localization, and so on. After univariate and multivariate analyses, the authors found that age, sex, stage, surgery, and radiation were independent prognostic factors of LCNEC.
There are several concerns as mentioned 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: CLINICOPATHOLOGICAL CHARACTERISTICS, TREATMENT AND SURVIVAL OF PULMONARY LARGE CELL NEUROENDOCRINE CARCINOMA: A SEER POPULATION-BASED STUDY
Review round: 2
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The revised manuscript is an improvement of the previous one. However, there are still some major concerns.
(1)	The TNM stages are very important prognostic variables, which is also mentioned by other reviewers. The authors can manually restage the stages using the variables CS tumor size,	CS extension, CS lymph nodes, CS mets at dx,	CS site-specific factor 1, and CS site-specific factor 2 recorded in SEER database. The details can be found on the website (https://seer.cancer.gov/archive/manuals/2016/appendixc.html).
(2)	The factors not significant in univariate analyses, such as race (p = 0.501), marital status (p = 0.215), laterality (p = 0.237), and chemotherapy (p = 0.796), should not be included in the multivariate analysis.


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: CLINICOPATHOLOGICAL CHARACTERISTICS, TREATMENT AND SURVIVAL OF PULMONARY LARGE CELL NEUROENDOCRINE CARCINOMA: A SEER POPULATION-BASED STUDY
Review round: 3
Reviewer: 1

Output:
Basic reporting: This is a revision; manuscript is well written. No issues
Experimental design: Methodology is appropriate.
Validity of the findings: Data is consistent with what should be expected.
Additional comments: Revisions are accurate and adequate. I have two minor suggestions.

1# I would recommend avoiding vague terms like "elderly" for e.g.  "Most of the patients were elderly people"

2# Authors report a low incidence 0.5% iof all pulmonary tumors as compared to single institutional series where in it is reported in about 2-3% of thoracic tumors. Authors suggests that "The main reason for this discrepancy is that previous research data were only based on post-operative data from a single center; however, our study is a multicenter study with a wide range of coverage, including both patients with surgery and without surgery. Therefore, the proportion of LCNEC in all lung cancer might be even lower than previously thought." This explanation might not be completely accurate. Pulmonary LCNEC diagnosis often needs a good quality biopsy or post op specimen. Inadequate FNAC or cytology can often miss the diagnosis. So I will recomend that authors should give both explanations and rather than stating that LCNEC incidence is low because SEER database analysis suggest so. They should also add that the incidence could be low in SEER due to underreporting/misdiagnosis in SEER.


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: CLINICOPATHOLOGICAL CHARACTERISTICS, TREATMENT AND SURVIVAL OF PULMONARY LARGE CELL NEUROENDOCRINE CARCINOMA: A SEER POPULATION-BASED STUDY
Review round: 3
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: it's now worth to be accepted 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: ASSOCIATED BACTERIA OF BOTRYOCOCCUS BRAUNII (CHLOROPHYTA)
Review round: 1
Reviewer: 1

Output:
Basic reporting: Excellent level and good clarity.
Suitable references.
Experimental design: Yes.  All good.
Validity of the findings: Conclusions are not all supported by the evidence and work.
Speculative comments not identified as such.
Additional comments: I have read with interest the manuscript by Gouveia et al., entitled “Associated Bacteria of Botryococcus braunii (Chlorophyta).
The paper describes a 16SrDNA barcode approach to describing the bacterial consortium that is present in different, long-term cultures of B. braunii.  Essentially, this study seems to follow on from previous work published by Sambles et al. (2017) in Microbiology Open (6; e482) on which Gouveia is an author but does not refer to this in any way.  This lack is surprising given that the earlier investigation demonstrated the adequacy of the 16S rDNA approach relative to whole-genome, shortgun sequencing, and resulted in an almost identical conclusion; that B. braunii appears closely associated with members of the rhizobiales that are possibly involved in nitrogen assimilation.
Another important observation in this paper is that there are more similarities in bacterial demographies within culture collections than between any B. braunii phylogeny – the exception being the aforementioned rhizobiales (lines 237-239).  This observation convinces me that the bacteria present is each culture are more probably due to culture artefacts than to any, normal microconsortium.  These artefacts are interesting, but well known in algal culture, and therefore the paper really does not present any new insight into algal biology on microbial associations.  Due to these artefacts, it is also impossible to assert, as the authors do, that “each strain has its own specific bacteria that are not shared between strains [of Botryococcus]” (lines 34-35); rather, it is clear that long-term, isolated culture leads to divergent representation in restricted populations that is dependent on the founder species – again a well-known tenet of ecology.
The paper, therefore, is underwhelming in its scope and conclusions, offering little new beyond the demographic description of bacteria present at a specific moment in long-term Botrycocccus cultures.  The science, however, is correct and well described, with no major flaws.  Equally, the manuscript is well-written and the figures are clear and representative of the data.
I cannot reject the paper on the basis of the experimentation or the methods, but I do not believe that it is, in of itself, worthy of publication, as the insights offered are much too slender and not generally applicable beyond the precise cultures used.


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: ASSOCIATED BACTERIA OF BOTRYOCOCCUS BRAUNII (CHLOROPHYTA)
Review round: 1
Reviewer: 2

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The manuscript is scientifically interesting and I recommend it for publication after minor revisions.
Specific comments
Line 62: “… (Jasti et al. 2005, Ramanan et al. 2015) (Eigemann et al. 2013, Hays et al. 2015)”. Is there any reason why the above citations are given in different parentheses?
line 78: “…B. braunii sp.” Delete the abbreviations ‘sp.’
line 204: “..Hydrogenophaga spp..”. Delete the second dot after spp.
line 274: “..were Hydrogenophaga spp.. Variovorax..” Replace the second dot with a comma.
Line 285: “..such as Chlorella spp. (Biondi et al. 2016).” Please correct, this citation refers to Tetraselmis not to Chlorella.
Lines 319-320. “ .. such as C. sorokiniana..” Please spell out the name of genus in full.

Lines 345-347.  The authors claim that “There is not a specific bacterial community
correlated to the different types of hydrocarbons produced by race A and B and mostly likely also not race L and S.” Why the authors refer to race S? This statement cannot be concluded with the analysis and results of the article.

Figure 3. I suggest to the authors to add the race subclassification letter symbols on plot (a) as they did in plot (b).

Table 1.  Please pay attention to: a) Citations not included in the reference list (e.g.  Nonomura 1988, Metzger et al 1985, Hilton et al 1989, Metzger and Casadevall 1987). b) Formatting of citations.


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: ASSOCIATED BACTERIA OF BOTRYOCOCCUS BRAUNII (CHLOROPHYTA)
Review round: 2
Reviewer: 1

Output:
Basic reporting: Clear and unambiguous
Experimental design: Methods described with sufficient detail.
Validity of the findings: Data is robust. Conclusions are well stated linked to original research question.
Additional comments: The revision on the paper and the responses to my comments are satisfactory.


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

Input:
Title: PIXEL: A CONTENT MANAGEMENT PLATFORM FOR QUANTITATIVE OMICS DATA
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript is overall well-written, figures are clear and easy to follow.  Although I like the general idea of organising secondary data more systematically, I was not fully convinced what Pixel’s USP was compared to popular and well-established tools like Galaxy which support many data types and offer data tagging.

1. Title is misleading. "Digital lab assistant" can mean anything. "Multi-omics": Pixel is made for a very specific type of omics data. There are many types of omics data that Pixel does not support, e.g. protein-protein interaction data. Similarly, the term data integration is misleading, as it usually implies the utility of ontologies, standard vocabularies, URI to integrate data - none of which are used in Pixel as far as I can tell. I suggest to use a different title, e.g.: “Pixel - a content management platform for quantitative omics data”.

2. Galaxy comparison. The latest version of Galaxy provides tags and group tags (https://docs.galaxyproject.org/en/release_18.09/releases/18.09_announce.html). Primary data can be tagged, and tags are propagated all the way to secondary data in Galaxy histories/ libraries. Galaxy has tools to filter, join and visualise tables. I like to know what the USP of Pixel is compared to Galaxy?
Experimental design: The software stack used to develop and deploy the software is cutting edge. The presented use case nicely illustrates the functionalities of Pixel. First, filters are used to identify two datasets tagged with the same species and “alkaline pH”. Second, genes/proteins are filtered if they contain “pathogenesis” in their description. It was not clear in Line 259-262 where the description came from.

My concerns are:
3. Reproducibility. Pixel clearly helps with data/content management, but I can’t figure out how it helps with reproducibility. As the authors clearly state, Pixel is not a tool for primary data analysis, it is for managing and visualising secondary data. When users upload data into Pixel, they can provide a description of the “Analysis” and “Experiment”. It is very optimistic to expect users would provide tool versions and parameters that created the secondary data. I would recommend to remove any references to reproducibility.

4. Metadata and Tags. The success of Pixel’s search depends on the richness and consistency of user-provided tags and metadata. Two users could choose different tags to describe the same dataset. How does Pixel cope with situations when different terms are used to describe the similar data, or with spelling mistakes?

5. Use case. It would be good if the author’s can illustrate why doing the same task in Galaxy would be more difficult: tag search, text search, join three tables and histogram visualisation.

6. Excel. Despite its universality it is notorious for identifier mangling (https://genomebiology.biomedcentral.com/articles/10.1186/s13059-016-1044-7). I would like to learn what safeguards there are against this.
Validity of the findings: Minor technical issues:
I got this following the README:
- ‘make bootsrap’ (first step):
- django.db.utils.ProgrammingError: relation "core_omicsarea" does not exist
	Went away after re-running the command
- Then I got: /bin/sh: yarn: command not found
- after installing 'yarn', it went fine

The step 'make dev', last message from 'make dev' is 'Creating pixel-dev_node_1 ... done'. At this point the server seems started, but the stdout isn't clear on this.
After this step, I can see a login page at http://127.0.0.1:8000, but I have no clue what to do from there.
There are requirements like Yarn, which the documentation should mention.
The documentation should say something about what to do once the application is deployed (is there a default user? Should I create it) and it should mention first steps I can do as user.
Moreover, regarding the step 'or run the Django development server and the css build watcher separately', it's not clear what to do as initial steps in either case (clone and cd).
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: PIXEL: A CONTENT MANAGEMENT PLATFORM FOR QUANTITATIVE OMICS DATA
Review round: 1
Reviewer: 2

Output:
Basic reporting: Denecker et al. describe the Pixel infrastructure, a digital lab assistant to manage, annotate and integrate datasets produced with a variety of omics technologies. In the first part of the manuscript, the authors describe the software in terms of underlying technology and design, and provide and overview of its usage. They then provide a short use case, leading to the identification of units potentially involved in the pathogenicity of Candida glabrata under alkaline stress using two omics datasets. The article is generally well written and conveys a good idea of the software's capabilities and the underlying technologies and development model.

In terms of spelling, I have two comments

- Line 79, possibly replace Importation with Import.
- Line 255, 'are search_ed_'
Experimental design: Here, I will comment on the software and its features:

- I struggled to install the software locally. Given the open development model used by the team, I will directly address this in a GitHub issue.

- As the authors acknowledge, installation of the software would require dedicated support, even if its use, later, is very intuitive. Given my own issues to install and test Pixel, and that potential users would want to try it out before installing it locally, it would be useful if a public instance could be used as test bench.

- I think further clarification or examples of what the Value and QS can be used for would be helpful. It becomes clear, later, in the example, that they can be used to store fold changes and p-values, but can any other data be used such as, for example classification or clustering results? Is there thus a limitation to 2 quantitative values per unit? How would one deal with a case where the experiment compared more than two groups and more fold changes and p-values are to be reported?

- Are the tags used to document Pixel sets free text, or is there some controlled vocabulary that will prevent different users to use different synonyms for the same term?

- On lines 188-190, I think there's a need for more information about 'user notebooks'. Are these dynamically executed scripts, or only provided as a documentation? Is this optional or mandatory?

- I would suggest to link to documentation in docs and docs-install in README.md file.

- Is it possible to export and update Pixel sets?

- On line 315, at the end of the discussion, the authors explain that a private web app contains 20000 pixels. Following the definition of a pixel being one line/entry in a pixel set, this seems very low.
Validity of the findings: Here, I will focus on the use case.

My main concern is that Pixel Set A zip file is missing and that the Pixel Set B isn't available at all. Hence, even if I had managed to install the software, I wouldn't be able to test it by reproducing the use case presented in the manuscript.

I also find the use case very simplistic, or lacking details, to make a convincing case. For example, the integration, based on common identifiers (unit names), which I assume, need to be mapped accordingly prior to importing the data into Pixel, is trivial. How would Pixel deal with metabolomics data, where liking between gene/protein names and metabolites isn't as direct? A more elaborated use case, that would illustrate a data analysis cycle as presented in figure 1C, would be useful.

In the use case, units matching the 'pathogenicity' term are searched for. How were these units annotated originally? This seems to be crucial to be able to perform a search and identify the units of interest. How could a user identify units matching a specific GO term, for example?

In the use case, the authors use p-values as an example. Are these not adjusted p-values, or shouldn't they be? This, in my opinion, also reduces the relevance of the use case.
Additional comments: To summarise, my main concerns are

- being able to test the software and either reproduce the use case, or at least use test data provided by the developers;
- the use case could be improved to make a stronger case in favour of Pixel.


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: DEVELOPMENT OF POLYMORPHIC EST-SSR MARKERS AND CHARACTERIZATION OF THE AUTOTETRAPLOID GENOME OF SAINFOIN (ONOBRYCHIS VICIIFOLIA)
Review round: 1
Reviewer: 1

Output:
Basic reporting: This article is mostly written in clear, unambiguous, professional English. Care should be given to "de novo" vs "denovo" used throughout the manuscript, referencing not consistent (et. al., 20xx vs et. al. 20xx), spacing between references (especially in Discussion) and other typing and spelling errors.

Literature are sited throughout. I would have preferred a bit more information on the uses of sainfoin, especially in animal husbandry and nutrition.

Figure 2 was not referenced in the text at all.

Reporting on the sequence, assembly and annotation of the transcriptome is sufficient and  well written. The validation of the wild sainfoin populations as well as the verification of sainfoin autoteraploidy is inconsistent between materials and methods, results and discussion and need more attention.

A better distinction between di/tri/tetra (etc)-nucleotides and the number of repeats should be made as it gets confusing what the authors mean.

The application of the EST-SSRs, once identified, is not clear. The authors only mention "...should benefit sainfoin improvement projects..." (lines 86-87) but more information on applications will give a clearer picture of the future application of the project.
The authors mention the objective is to "allow a better understanding of the diversity of sainfoin" (line 92). However, with 40 individuals from only 5 locations this could hardly be achieved.
Experimental design: The experiment was well designed. The aim (transcriptome sequencing) and two objectives (diversity and autotetraploidy) were clearly defined. However, more than 40 individuals should be tested before final conclusions on genomic diversities can be made. The authors can elaborate on why the use of sainfoin plants in agriculture is important and why this species was selected for sequencing, as well as the down-stream application of the identified EST-SSRs.

The transcriptome sequencing, assembly and annotation of EST-SSRs were well reported in materials and methods as well as results and discussion.

Sampling of wild populations, DNA extraction and amplification were well reported. The construction of T4 vectors were very poorly described (especially sequencing) and need more attention. Diversity analysis of wild populations were insufficient and inconsistent between materials and methods, results and discussion. For example, in materials and methods it is noted that population parameters were analysed using ATETRA 1.2, and in Discussion the authors mention that POPGENE were used to analysed some and other population parameters. No reference of which program was used is made in Results. No mention is made of which sequencing method is used (e.g. Sanger or NGS) of the T4 vectors and methods and results can be elaborated. The authors should pay attention to these sections and ensure correct reporting throughout.

The results from Figure 4 should be elaborated and better explained in materials and methods, results and discussion.
Validity of the findings: The transcriptome sequencing and EST-SSR identification data is well written, statistically sound and controlled.

The diversity analysis data in lines 225 - 236 and 237 - 249 should perhaps be given in table format as it is difficult to read.

EST-SSR identification was conclusive and compared well with results from similar projects.

Line 145-146 states the plants had different tannin contents and compositions. No data is given to state these claims.

The legends of Figures 2 and 3 are swapped and no mention of Figure 2 or an interpretation of the data is given. The authors should elaborate much more on these findings.

More detail on where EST-SSRs are located in the transcriptome (lines 235-236) can be given to show that included EST-SSRs cover the whole transcriptome.

The results of the autotetraploidy analysis is lacking and more detail should be given here.

The diversity analysis should be given more attention and it should be justified why only 40 individuals were used.

Conclusion is well stated.
Additional comments: Overall not a bad article but it needs more attention to the following:
- why sainfoin is important especially in agriculture and why this study was needed;
- diversity and autotetraploidy verification sections need more detail and attention to detail;
- whole article need to be checked for spelling errors, detail typing errors and inconsistent referencing;
- Figure 2 not addressed and legends incorrect;
- Figure 2 and 3 should be elaborated on;
-Re-write lines 80 - 81 as it is not clear;
- Line 52 - give SSR abbreviation, line 57 - write out EST;
- Lines 67 - 69: why is the current EST-SSR primers insufficient?


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: DEVELOPMENT OF POLYMORPHIC EST-SSR MARKERS AND CHARACTERIZATION OF THE AUTOTETRAPLOID GENOME OF SAINFOIN (ONOBRYCHIS VICIIFOLIA)
Review round: 1
Reviewer: 2

Output:
Basic reporting: Your title needs to be more clear and specific to match the content. In line 2, this is my suggestion: "...characterization of the autotetraploid genome of sainfoin..."
Experimental design: Lines 96 to 100. Were these tissues harvested from different saifoin plants or was it from the same plant? The reader will need to know this information.
If from different plants, how many were they and was it the same species/ clone? This will help the reader determine if the variation is a result of genotypic differences or not

When you were doing the cDNA library construction, how many illumina lanes were run? This information will give an idea of the sequence depth

Line 144. These wild saifoin plants; were they different species from O. viviifolia? Please write the species name. Are they also tetraploid, or this information is not known at this moment?

Line 157. How were the 200 primer pairs identified? Tell this to your readers please
Validity of the findings: When you were doing rigorous quality checks on the sequence reads, what did this rigor involve? What was the requirement?

In your results concluding paragraph (line 267, 268), you concluded that sainfoin is autotetraploid. However, you only used the number of PCR amplicons to come to this conclusion. I think you need to explain how much more powerful this method is compared to flow cytometry which is the norm when doing ploidy analysis?
Additional comments: I think you have done a good job and that this research will be very useful to the scientific community. My only striking observation that I noticed is that there are slight details which you have and need to include. Overall, your research adds valuable knowledge to the academic and scientific community.


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: DEVELOPMENT OF POLYMORPHIC EST-SSR MARKERS AND CHARACTERIZATION OF THE AUTOTETRAPLOID GENOME OF SAINFOIN (ONOBRYCHIS VICIIFOLIA)
Review round: 2
Reviewer: 1

Output:
Basic reporting: The authors addressed all my issues and I'm happy with the review.
Experimental design: The authors addressed all my issues and I'm happy with the review.
Validity of the findings: The authors addressed all my issues and I'm happy with the review.
Additional comments: I'm happy to accept this revised article.


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 GENOMIC ANALYSIS OF THE IDD GENES IN FIVE ROSACEAE SPECIES AND EXPRESSION ANALYSIS IN CHINESE WHITE PEAR (PYRUS BRETSCHNEIDERI)
Review round: 1
Reviewer: 1

Output:
Basic reporting: Qverall, the results are informative. Because there are many grammatical errors in English writing, it is strongly recommended that manuscript needs to be polished by an English native speaker.

The logic was weaker in INTRODUCTION. One of the important content of this article is the identification of IDD genes from five Rosaceae species. However, only pear is introduced and no any introduction of other four Rosaceae species. You'd better briefly introduce these species and their genomes to give readers an idea of your research goals. Why did you do this? Functional comparison? Evolutionary comparison?
Experimental design: no comment
Validity of the findings: The functions of the IDD genes in pear are all speculated partially based on the expression levels. There is limited evidence showing that PbIDDs 3, 5 play a role in SCW formation and lignin biosynthesis in pear fruit and no evidence showing that PbIDDs 2, 8 are involved in the regulation of plant flowering induction in pear. Therefore, you should revise your conclusion.
Additional comments: The manuscript deals with the comparative genomic analysis of the IDD genes in five Rosaceae species and expression analysis in Chinese White Pear (Pyrus bretschneideri). In general, the results are informative. Although there are many grammatical errors in English writing, overall, the English of the manuscript is readable. However, it is strongly recommended that manuscript needs to be polished by an English native speaker. In addition, the functions of the IDD genes in pear are all speculated partially based on the expression levels. There is limited evidence showing that PbIDDs 3, 5 play a role in SCW formation and lignin biosynthesis in pear fruit and no evidence showing that PbIDDs 2, 8 are involved in the regulation of plant flowering induction in pear. Therefore, you should revise your conclusion.
There are other issues that need to be revised:
1)	The authors identified IDD genes from 5 Rosaceae species belonging to different genera. The author used both common names and scientific names. However, common names of species are not consistent with scientific names which create big confusion. A common name fruit trees may include different species, like pear (Pyrus communis, Pyrus pyrifolia…). Fragaria vesca refers to woodland strawberry, not strawberry which is assigned to Fragaria × ananassa. Prunus mume is not a plum, but scientific name of Japanese apricot. Although Prunus mume is sometimes called Chinese plum, which is not popular. The real Chinese plum is usually assigned to Prunus salicina. So, please confirm whether you identified IDD genes from plum or from Japanese apricot. Rubus occidentalis refers to common name black raspberry. Therefore, please revise the common names of fruit trees used in your study. In most cases, it is recommended that only using scientific names to replace the common names.
2)	Please use cultivar to replace variety. Variety is often used to refer to infraspecific taxon. While cultivar refers to the most basic classification category of cultivated plants.
3)	The logic was weaker in INTRODUCTION. One of the important content of this article is the identification of IDD genes from five Rosaceae species. However, only pear is introduced and no any introduction of other four Rosaceae species. You'd better briefly introduce these species and their genomes to give readers an idea of your research goals. Why did you do this? Functional comparison? Evolutionary comparison?
4)	–Line 68-69: you mentioned that ‘Dangshan Su’ had a high medicinal efficacy. What kinds of medicinal efficacy? Please cite references.
5)	–Line 69: please use pear flesh to replace pear pulp.
6)	–line 70: “they determine the quality of the pear fruit”? please specify the meaning of fruit quality. Stone cells may determine the fruit texture. However, fruit quality includes many aspects.
7)	–Line 89-90: please reword “Among the Rosaceae species, only apples have had their IDD genes identified genome-wide”.
8)	--Line 98, what does DAF mean? You should use the full name before the first abbreviation.
9)	--Line 104 -105: “The genome databases of the other four Rosaceae plants (Strawberry, plum, Raspberry and Cherry) were downloaded from GDR”. However, no genome sequence data of Prunus mume can be found in GDR website. You should check carefully.
10)	–Line 121: what is the meaning of “can from the study of Fan”?
11)	--Line 122:  Arabidopsis thaliana should be italic. In addition, all the gene name should be italic. Please check the text and revise.
12)	–Line 134: ” (http://mapinspect.software.informer.com) (Niu et al., 2016)” Please cite original reference to replace Niu et al., 2016.
13)	–Line 151: Suzhou County? Anhui Province?
14)	--Line 191: please reword the sentence: “2 pear genes, 2 strawberry, 2 plum, 2 raspberry, 1 cherry, and no apple”.
15)	In the section RESULTS, please avoid presenting any discussions and only describe your experimental data using past tense. Please remove sentence of line 183 to line 190, line 240 to line 242, line 245-246 line 259 to 2263, and so on.
16)	–Line 271：Chinese pear should be replaced by Chinese white pear.
17)	Please perform the significance analysis in Fig.7
18)	In the section DISCUSSION, when you cite your experiment results, you should indicate which figure or table you refer to.
19)	Please remove the repeated results in the discussion. For instance: Line 335-338.
20)	--Line 367:  ‘group4’ should be ‘group 4’. Please also check other parts of the text.
21)	In section CONCLUSION, please revise “we believe that PbIDD2, 8 are involved in the regulation of plant flowering induction in pear and that PbIDD3, 5 are involved in the regulation of SCW formation and lignin biosynthesis in pear fruit”. In this study you don’t have solid evidences to support these conclusions.


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 GENOMIC ANALYSIS OF THE IDD GENES IN FIVE ROSACEAE SPECIES AND EXPRESSION ANALYSIS IN CHINESE WHITE PEAR (PYRUS BRETSCHNEIDERI)
Review round: 1
Reviewer: 2

Output:
Basic reporting: As a Chinese writer, the Chinglish in this manuscript is obvious. The language statement is not refined. There are some repeated sentences can be found both in Result and Discussion. Most of these kinds of language problems I found have been labeled with wavy underline or linear underline in the PDF.
Experimental design: no comment
Validity of the findings: Conclusion are not well stated (See the review tips)
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 GENOMIC ANALYSIS OF THE IDD GENES IN FIVE ROSACEAE SPECIES AND EXPRESSION ANALYSIS IN CHINESE WHITE PEAR (PYRUS BRETSCHNEIDERI)
Review round: 2
Reviewer: 1

Output:
Basic reporting: Although both reviewers requested that the manuscript should be polished by an English native speaker or English-editing Service Company, there are still many many English grammar mistakes. I strongly suggest the manuscript should be polished before acceptance.
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The revised manuscript has been significantly improved and the authors have addressed my major concerns. However, there are still some problems to be revised.
1．	Although both reviewers requested that the manuscript should be polished by an English native speaker or English-editing Service Company, there are still many English grammar mistakes. I strongly suggest the manuscript should be polished before acceptance.
2．	Line 17: What is Chinese white Pyrus bretschneideri;
3．	Line 20: please use different organs to replace different tissues, because fruit, flower and bud you used in the experiment are organs, not tissues.
4．	Line 55: please revise: “the variety of pear cultivars grown is very wide”. I cannot understand the meaning of this sentence. The variety generally refers to Infrageneric taxon. You may mean that there are lots of different pear cultivars grown in China??
5．	Lines 55-56: There are several points that need to be clarified or modified. When you use the expression “’Dangshan’ pear (Pyrus bretschneideri)”, you can’t further use Pyrus bretschneideri cv. Dangshan Su. The term “White pear” is often used by Chinese researchers. However, in Asia, there are also other types of pear cultivars or cultivar groups native to different countries: Japanese pear, Chinese sand pear…. Thus you had better use Chinese white pear to replace white pear. Furthermore, if you have read Dr. Teng and his colleague’s papers, you should have known that Pyrus bretschneideri has been misused to refer to the cultivars of the Chinese white pear. They proposed the name for Chinese white pear: Pyrus pyrifolia White Pear Group to replace misused Pyrus bretschneideri. In this paper, you can still use Pyrus bretschneideri. But you are recommended to read papers to understand what is Chinese white pear, what is P. bretschneideri. In addition, what is the meaning of “a characteristic pear type”?
6．	Line 58: “The tone cells …. pear's unique taste”? Stone cells are related to flesh texture, not to taste. Please revise the sentence.
7．	Line 59: “However, there are few reports on fruit development;” I don’t think so. There are many papers dealing with fruit development in many fruit trees.
8．	Line 70: tissues or organs?
9．	Line 74: Sweet cherry (Prunus avium) is originated form Europe, Anatolia, Maghreb, and western Asia, not only from Europe.
10．	Lines 78-79:  the four species included in your paper belong to rose family, but they belong to different subfamily, which means that they don’t have a close evolutionary relationship.
11．	Line 367: what is ” the developmental law”?
12．	Lines 290-291: the authors attributed different number of IDD genes identified in Malus and Pyrus to origin of the IDD gene family. This may be true. But the authors should also consider the quality of genome sequence of P. bretschneideri used in this study.
13．	Acknowledgements: the authors expressed their gratitude to themselves? This is very funny.


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 GENOMIC ANALYSIS OF THE IDD GENES IN FIVE ROSACEAE SPECIES AND EXPRESSION ANALYSIS IN CHINESE WHITE PEAR (PYRUS BRETSCHNEIDERI)
Review round: 2
Reviewer: 2

Output:
Basic reporting: The English is largely improved and I have highlighted the words or sentence that are wrong or difficult to understand in the PDF version. The authors have done a lot of effort to improve the quality of the manuscript according to our suggestions, but minor revision is required.
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: ROBUST AND AUTOMATIC DEFINITION OF MICROBIOME STATES
Review round: 1
Reviewer: 1

Output:
Basic reporting: Generally, the syntax of this manuscript needs to be improved. It is often casual in tone and rife with colloquial word choice. Interrupters, parenthetical statements, and transition statements (eg. ‘such as’, ‘here’, ‘for example’) are used too liberally. There are grammatical errors. The entire paper needs a heavy edit for professional language. In the spirit of Strunk and White, needless words should be deleted.

The materials and methods section is well written and clear.

Specific comments:
Lines 28-29: cavity does not describe where microbiomes are, yet there is a parenthetical statement regarding distinct environments directly following. This second statement should be used to describe the microbiome more generally.

Lines 33-34 sentences starting “Albeit…” and “Some studies…” are fragments and should be combined into a complete thought. The following statements 37-46 should be revised to fit with the earlier changes suggested in 33-34.

Line 60 contains an example of a distracting and non-contributing parenthetical statement.

Lines 238-240 are not needed, as the idea contained within is implicit in the following text.

Lines 328-329 contain an introductory sentence too casual for a manuscript.

Lines 367-370 contain a run-on sentence which obfuscates your main point.

		Lines 390-392 contain further casual language.

Line 427, ‘an standard’ is a mistake and should be corrected.

The tables should both be reformatted and given proper legends. They are difficult to understand quickly and would benefit from better labels and graphic design. I personally do not like labelling the datasets by the author, but it would not prevent me from accepting the manuscript. However, ‘no.clusters’ and ‘no.taxa’ should be written out in English to prevent confusion. It is very frustrating that the headers contain back slashes and yet the cells contain forward slashes.

The figures should be labeled better – rather than describing where a panel is in the figure legend, please label it and use the legend to describe what the figure is, rather than where it is in space.

The conclusion section needs to be edited for brevity.
Experimental design: The logic of this study is very clear and I think that this protocol and algorithm fills an important knowledge gap / tool niche. As stated in the manuscript, enterotyping is often ad hoc and this may be a solution to the softer end of the science.

The methods section is very clear and well cited.
Validity of the findings: I am cautious to agree with the assertion that this algorithm ‘broadly applicable’ as stated in the abstract and implied in the body. While there are high value datasets included in the study, four sets is but a small fraction of the data extant. It would be helpful to further validate these findings in other diverse populations, other models or nonhuman systems before claiming broadness or generalizability.

I do, however, think that this algorithm will contribute to the conversation regarding microbial states, enterotypes, or gradients. Without standardization of workflow, it is certainly a challenge to assess change over time and differences between populations or localities.

The transparency of the description of the algorithm, specifically figure 1, should be commended. Its sometimes a challenge to reconstruct the logic of tool design and, here, the reader is shown the plan with great clarity.
Additional comments: The approach is interesting and needed to help resolve outstanding tension in the field as well articulated in the Patrick O’Toole’s 2012 comment (doi:10.1038/nrmicro2859) and the cited Costea//Bork perspective from 2017 (10.1038/s41564-017-0072-8). The point, however, is often muddied by poorly presented figures and tables and bad writing.

I would like to see this paper largely reconstructed for clarity and brevity in the case that the methods described within become part of the standards repertoire as you suggest in your discussion, lines 380-385. Reanalysis of Human Microbiome Project data would certainly help bolster the position that this is a generalizable tool. I recognize that it is no easy task to amass set after set of data and that this concern will be mitigated upon publication, adoption in the field, and iterative design. However, in this case more is better, or it should be deemed a proof of concept for a useful new standard.


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: ROBUST AND AUTOMATIC DEFINITION OF MICROBIOME STATES
Review round: 1
Reviewer: 2

Output:
Basic reporting: Honestly, this manuscript looks like an unfinished work in progress and needs to be re-thought from the basic message to reporting. For example, I could not find references to Table 2 or Figure 3 in the text.
Experimental design: In my opinion, the best bet to get this work published is to reinvent is as an application note rather than original research. There's probably most value in the algorithm as a collection of working standard R code than in any of the results reported.
Validity of the findings: Claim: Generic, domain-independent and automatic procedure to determine a reliable set of microbiome sub-states within dataset and with respect to the conditions of the study.

The authors try to make a point about the difference between states and sub-states, however, the exact difference between the two escapes this reviewer, especially, given the fact that the methodology to define both of these are very similar. Further, the microbiome field is converging on the community state type (CST); perhaps, the authors could use more standard terminology here.

How do the authors define reliability? Although the manuscript defines some measures of robustness, it is never made explicit what it means for CSTs to be reliable.

What does domain-independent mean?

The methods presented are all unsupervised, therefore, I do not see how these approaches define CSTs with respect to conditions of the study. Unsupervised approaches may result in CSTs that are coincidentally related to conditions of a study, but there is no explicit guarantee.

It is not clear why the authors are focusing on longitudinal datasets. The analyses do not in any way exploit the fact that the data are longitudinal and/or sampled multiple times from the same individuals.

Methodologically, no novel approaches are proposed. The manuscript proposes to exploit best practices in clustering and uses a few established techniques that have appeared elsewhere in microbiome literature. Therefore, it is not clear what audience will find this information useful. Nonetheless, there is nothing wrong with the approaches used by the authors.

I could not follow the definition of Jaccard similarity clustering assessment.

The most important unaddressed question by this manuscript and in the field is the evidence for clustering. The evaluation is performed on k>=2 number of clusters; it is never checked whether k=1 is the optimal clustering.

Cluster interpretation results could benefit from formal statistical testing of association between the clusters and the putative experimental variable/factor.

"The score is the frequency of correct classification pairs." => "The score is the frequency of matching classification pairs." ??
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: ROBUST AND AUTOMATIC DEFINITION OF MICROBIOME STATES
Review round: 1
Reviewer: 3

Output:
Basic reporting: The english grammar is lacking.

It is difficult to understand how to use these "cluster" results.  So what if they found two clusters -- what are they and how can we use this to analyze the results?

The description of the pipeline is vague.

In general, it is hard to understand the relevance of this paper because the authors do not explain how their results can be used.
Experimental design: Is agglom clustering the right approach for measuring the evolution of “substates” if the clusters are discrete in that the OTUs do not overlap from cluster to cluster? One would presume that a transition from substate 1 to substate 2 may involve shifts in the relative abundance of certain taxa that fall within a cluster and not a shift from OTUs specific to cluster x to OTUs specific to cluster y.

Just validating their results against the original papers is not enough.
 The authors need to show the advantages/disadvantages of their approach by generating simulated data and see if they can capture the predefined substates, and then they need to describe at what point their approach fails and to what parameters their approach is sensitive to.

They also claim they want to describe the dynamics of these clusters – i.e., how they behave over time. This was not shown.

In Section MATERIALS AND METHODS, authors mentioned they used SI to determine best number of clusters. But the authors didn’t discuss the reason why this SI works well for this task expect for barely mentioned that it was from a previous research. It would be beneficial if the authors could provide some insights about why SI is helpful to find good number of clusters in microbiome dataset.

In Section MATERIALS AND METHODS, authors mentioned the threshold for PS is 0.8 and the threshold of Jaccard is 0.75. But the authors didn’t discuss the selection of threshold in this section.

In Section RESULTS, by looking at Figure 1, it seems that Jaccard similarity criterion is redundant as it is always better than PS. Did the author also evaluated the efficacy of the metrics used in the pipeline to avoid unnecessary computation?
Validity of the findings: Figure 2 needs to be redone. It’s too difficult to distinguish shapes. Decide to plot ellipses or to use facets.

Clustering samples from subject A and B together makes no sense for David data because we are not interested in how well we can separate two subjects. Instead, the goal of this paper is to identify sub-states that can capture transitions in longitudinal microbiome data. Clustering samples together will reduce each resolution because the intra class distance between samples from two subject A and B is too big.

Why only 2 clusters are found for David et al. (2014)?  There is a pre-state, a dysbiosis state, and a post-state (Figure 3F of the original paper).. so there should be at least 3, which shows that this paper doesn't produce proper results.  How about looking at Fig 4B in https://www.biorxiv.org/content/biorxiv/early/2017/08/15/176412.full.pdf which shows 5 clusters... with 3 of them being major, with corresponding taxonomic relative abundance and posterior probability of cluster over time?  This is a much more in-depth analysis than just determining the number of clusters.

The authors previously mentioned that their pipeline can provide new insight into the data structure. But from the result, readers couldn’t really find interesting and novel explanations respect to the clusters the pipelines found. Basically, the pipeline can recover metadata information by the clusters to some extent, but the resolution is relatively low because there is no further analysis followed up by digging into each cluster and try to interpret the intrinsic relationship between clusters (OTU profiles that are similar, for example).
Additional comments: Line 66: cite https://genomebiology.biomedcentral.com/articles/10.1186/gb-2014-15-7-r89 and https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5621509/


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: ROBUST AND AUTOMATIC DEFINITION OF MICROBIOME STATES
Review round: 2
Reviewer: 1

Output:
Basic reporting: Dear authors,

This is such a vast improvement on the previous manuscript. Thank you for your careful work and heavy edits - it really clarifies your contributions documented in this manuscript. There appears to be a very minor line splice over lines 481-484 where something may have been transposed from your clipboard. The sentence will make more sense once this is corrected.

Otherwise, all comments have been addressed or rewritten.
Experimental design: The design of the experiments is repetitive and clear and spans a broad swath of both microbiomes and the mathematical evaluation of their emergent clusters. I am satisfied that they indeed cluster microbiomes in a data driven fashion. I don't think you include a link to your github - obviously you are findable, but including a link to the code would be helpful / thorough.
Validity of the findings: This manuscript seems to have improved significantly, and for the better, with revision. Based on the information provided in the manuscript and the supplemental files I believe that their approach provides biology-agnostic, data driven clustering. Based on their github repository, I see the r scripts to be well documented and clearly useful to the microbiome computational biology community.
Additional comments: Thank you for taking all three reviewers comments so seriously. You took a good idea and made it into a clear paper. The only reason that I have not accepted this paper outright is because of the presumed typo above and my hope that the editor will suggest a place where it is appropriate to link your github or r scripts.  I wish you the best in all your future endeavors.

Sincerely,
Jonathan LoTempio Jr


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: ROBUST AND AUTOMATIC DEFINITION OF MICROBIOME STATES
Review round: 2
Reviewer: 2

Output:
Basic reporting: The manuscript was drastically changed and hard to compare with the previous version and almost warrants a resubmit.

"There are relatively few studies about temporal dynamics within any microbiome site" -- I feel like the authors are leaving out numerous studies.  Off the top of my head, I know this one:  https://www.nature.com/articles/ncomms5344

Their approach -- clustering and optimally determining the number of clusters based on different distance measures etc. -- does not seem very novel.  And they fail to really compare their methods and results to other papers' methods and results.  Like, we had you site Ananke (that and other methods exist)... how does your method compare, even if it does something different -- compare and contrast to other algorithms.
Experimental design: A state should be defined for the input data and desired output.  The following definition is confusing:  "In this manuscript, we define a microbiome ”state” as a collection of constraints satisfied by the58microbiota of some given samples, that are not satisfied by other samples, allowing them to be reliably59distinguished from one another."  -- So, is a state a group of samples or is it a group of OTUs?  This explanation does not help:  "Therefore, our approach to defining states, described in detail below, consists100of applying a clustering algorithm to the OTU data, taking a metagenomics beta diversity metric as the101distance measure between samples." ... sounds like states are clustering of samples.

The authors use k from 2 to 10 in their algorithm as the number of clusters. However, since you are working on longitude data, there is a chance that there are more than 10 changes in a long period of time. Therefore, it makes more sense to increase k until the clustering assessment scores stays the same or drops (thus you could have a way of smartly choosing k).

Your algorithm assumes that there are at least 2 clusters in the data, what if there is only 1 cluster?

For the David et al. data:  As the authors wrote in their responses when they apply their algorithm to subject A only, they could find 2 clusters: “for subject A we get 2 clusters, that is in agreement with Figure 3B and 3D of the original study, with one cluster for samples during travel (approx. 70-120 collection days) and other cluster for the remaining samples”. Whereas in the manuscript, the authors just show the results of clustering A and B together and in this setting, the authors can’t find the cluster that correspond to A’s travel period. I still suggest the authors show the results of their clustering experiment on subject A and B separately in the paper.

In line 358-370, the authors talk about the composition of the clusters they found. I suggest to provide a figure to illustrate the composition. For example, a phylum level distribution for each cluster. They can also integrate this in their algorithm to help users to explore the clusters produced.
Validity of the findings: I am glad the authors brought up their recently published paper. I think the algorithm presented in this paper is a parameter sweep tool to find the best number of cluster because no further analysis is followed by the algorithm hence this algorithm itself can’t really identify biological meaningful states. However, when this algorithm is followed by the other method proposed in the authors’ recently published paper, we can identify interesting clusters as well as understand their transitions. That makes the clusters valuable and interpretable.

The authors should have included this algorithm in the previous paper in the methods.  Without comparing this algorithm to other algorithms, this cannot be a standalone paper.
Additional comments: The writing is still poor, e.g. "'habits' such as lifestyle."


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 RELATIONSHIP BETWEEN SERUM URIC ACID WITHIN THE NORMAL RANGE AND Β-CELL FUNCTION IN CHINESE PATIENTS WITH TYPE 2 DIABETES: DIFFERENCES BY BODY MASS INDEX AND GENDER
Review round: 1
Reviewer: 1

Output:
Basic reporting: In this study the Authors aim to demonstrate the relationship between serum uric acid (SUA) within the normal range and the beta cell function (HOMA 2B) and insulin resistance indexes (HOMA IR) in type 2 diabetic Chinese population.
Experimental design: In my opinion, the Authors should also keep in mind that Asian population have different cut offs for BMI, HOMA-IR, HOMA 2B and other metabolic indexes. In this regard, it would be necessary to make a new additional table and to re-evaluate the association between SUA levels and cut off point of BMI recommended for the Asian population.
Validity of the findings: The results of this study evaluate for the first time the association between normal range of SUA with beta cell function and insulin resistance in diabetic Chinese population. It is interesting to note that normal SUA levels were associated with beta cell function in overweight/ obese people, while no association was seen in normal weight people. However, it would be attractive deepened even with further studies the molecular mechanisms underlying these observations of associations.
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 RELATIONSHIP BETWEEN SERUM URIC ACID WITHIN THE NORMAL RANGE AND Β-CELL FUNCTION IN CHINESE PATIENTS WITH TYPE 2 DIABETES: DIFFERENCES BY BODY MASS INDEX AND GENDER
Review round: 1
Reviewer: 2

Output:
Basic reporting: Not completely convincig results
Experimental design: Sample size adequate
Validity of the findings: I find a little bit contradictory that UA levels are associated with beta cells function and contextually do not mirror IR in T2DM patients.
The normal range of UA is different for gender.

The reference ranges for uric acid are as follows:
Men: 2.5-8 mg/dL
Women: 1.9–7.5 mg/d

Gender plays a key role also for other aspects as ... Cardiol. 2016 Feb;67(2):170-6.
Gender differences in the association between serum uric acid and prognosis in patients with acute  coronary syndrome.
Additional comments: I find a little bit contradictory that UA levels are associated with beta cells function and contextually do not mirror IR in T2DM patients.
Probably, this is due to the imprecision of surrogate markers used for the assessment.......my hypothesis!
As the authors know very well.....The normal range of UA is different for gender and thus there are renal factors involved.
Authors should give values also in mg/dL for the sake of completeness.
The reference ranges for uric acid are as follows:
Men: 2.5-8 mg/dL
Women: 1.9–7.5 mg/dL
Gender plays a key role also for other aspects as ... Cardiol. 2016 Feb;67(2):170-6.
Gender differences in the association between serum uric acid and prognosis in patients with acute  coronary syndrome.
The ACS is the main complication of T2DM patients,


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 RELATIONSHIP BETWEEN SERUM URIC ACID WITHIN THE NORMAL RANGE AND Β-CELL FUNCTION IN CHINESE PATIENTS WITH TYPE 2 DIABETES: DIFFERENCES BY BODY MASS INDEX AND GENDER
Review round: 1
Reviewer: 3

Output:
Basic reporting: Good
Experimental design: Good
Validity of the findings: Good
Additional comments: The current study suggested that: 1) SUA levels are significantly associated with HOMA2%B in T2DM patients with overweight/obesity group, but not in normal weight group; 2) other islet function indexes, such as FCPI, PPCPI, and ΔC-peptide, did correlate with SUA levels in T2DM patients with overweight/obesity group; 3) the absence of a relationship between SUA and HOMA2IR after adjustment for Cr, BMI, sex, HbA1c, and diabetic duration in T2DM patients with overweight/obesity.
          To define Metabolic Syndrome, the presence of at least three of the proposed criteria is required, and sometimes it is sufficient to have only one laboratory value, modified by diet or drugs, for the classification of Metabolic Syndrome. [1]. Recently, long term high fructose diet induced metabolic syndrome with increased blood pressure and proteinuria in rats [2]. Metabolic syndrome was associated with dual increase in renal glucose and uric acid transporters, including SGLT1, SGLT2, GLUT2, GLUT9 and UAT [2]. Dapagliflozin, a novel oral diabetic drug - Sodium-glucose cotransporter 2 inhibitor, could improve glycemic control and lower SUA levels in hospitalized patients with uncontrolled T2DM [3].
             Authors are kindly requested to emphasize the current concepts about these issues in the context of recent knowledge and the available literature. This articles should be quoted in the References list.
References
1.	What about non-alcoholic fatty liver disease as a new criterion to define metabolic syndrome? World J Gastroenterol. 2013 Jun 14; 19 (22): 3375-84. doi: 10.3748/wjg.v19.i22.3375.
2.	Alterations of Renal Epithelial Glucose and Uric Acid Transporters in Fructose Induced Metabolic Syndrome. Kidney Blood Press Res. 2018;43(6):1822-1831. doi: 10.1159/000495814.
3.	Effects of dapagliflozin on serum uric acid levels in hospitalized type 2 diabetic patients with inadequate glycemic control: a randomized controlled trial. Ther Clin Risk Manag. 2018 Dec 11;14:2407-2413. doi: 10.2147/TCRM.S186347.


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 RELATIONSHIP BETWEEN SERUM URIC ACID WITHIN THE NORMAL RANGE AND Β-CELL FUNCTION IN CHINESE PATIENTS WITH TYPE 2 DIABETES: DIFFERENCES BY BODY MASS INDEX AND GENDER
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: 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 RELATIONSHIP BETWEEN SERUM URIC ACID WITHIN THE NORMAL RANGE AND Β-CELL FUNCTION IN CHINESE PATIENTS WITH TYPE 2 DIABETES: DIFFERENCES BY BODY MASS INDEX AND GENDER
Review round: 2
Reviewer: 2

Output:
Basic reporting: Fine
Experimental design: Fine
Validity of the findings: Valid
Additional comments: Manuscript improved


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

Input:
Title: THE RELATIONSHIP BETWEEN SERUM URIC ACID WITHIN THE NORMAL RANGE AND Β-CELL FUNCTION IN CHINESE PATIENTS WITH TYPE 2 DIABETES: DIFFERENCES BY BODY MASS INDEX AND GENDER
Review round: 2
Reviewer: 3

Output:
Basic reporting: Good
Experimental design: Good
Validity of the findings: Good
Additional comments: Homeostatic model assessment for insulin - resistance (HOMA-IR), a relation between fasting glucose and insulin has been recognized as the most sensitive and specific method for measuring insulin resistance and risk for T2D, therefore Metabolic Syndrome. If it speaks of insulin - resistance the authors should specify that to define Metabolic Syndrome, the presence of at least three of the proposed criteria is required, and sometimes it is sufficient to have only one laboratory value, modified by diet or drugs, for the classification of Metabolic Syndrome [1].
Authors are kindly requested to emphasize the current concepts about these issues in the context of recent knowledge and the available literature. This article should be quoted in the References list.
Reference
1. What about non-alcoholic fatty liver disease as a new criterion to define metabolic syndrome? World J Gastroenterol. 2013 Jun 14; 19 (22): 3375-84. doi: 10.3748/wjg.v19.i22.3375.


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: LOANCORHYNCHUS CATRILLANCAI GEN. ET SP. NOV., A NEW SWORDFISH (XIPHIOIDEI, BLOCHIIDAE) FROM THE MIDDLE EOCENE OF CENTRAL CHILE
Review round: 1
Reviewer: 1

Output:
Basic reporting: The ms. is generally readable and readily understood, but there are some awkwardly worded passages and misspelled words reflecting the fact that English is a second language for the author. I recommend that the author have someone completely fluent in English take one more editorial pass through the ms.

For ease of use by readers, I recommend that the author spell out and explain abbreviations for institutional collections when they are first used in the text (e.g. ‘GS 13924’ on p. 7 of the ms. pdf) rather than just listing them in one place.

Family name should be BLOCHIIDAE, not BLOCHIDAE as it is spelled on ms. p. 14. Likewise XIPHIAS not XIPHIUS (p. 6).

The author should define and cite a source or sources to better explain the Weddellian Biogeographic Province.

I don’t understand what ‘strongly attached … because of taphonomic reasons’ means, in the description of the opercular series on p. 19.

The Taxonomic Placement section that is in the Discussion should be in the Systematic Paleontology section, it is much more description than discussion.

I found Fig. 1A a bit difficult to interpret – is the coastline of Chile indicated on this map on the left edge of the gray-shaded area? If so it would be helpful to just add ‘Pacific Ocean’ to the map to make that clear.

On Fig. 2, the abbreviation dr is not explained in the caption, and the abbreviation fr is not in bold as are the other abbreviations in the caption. Also, what is meant by ‘anterior bulk?’

On Fig. 4, I suggest using the wording ‘line drawing interpretation of element’ rather than the awkward phrase ‘scheme of the previous.’

On Fig. 7, it would be helpful to see the supporting characters (by number) mapped on, or at least an explanation of where in the paper that information is provided.
Experimental design: The scope of comparisons in the Diagnosis needs to be expanded – in other parts of the paper the author makes comparisons between the new taxon and several previously described billfish taxa including Aglyptorhynchus and Hemingwaya, but then only makes comparisons with Blochius, Xiphias and Xiphiorhynchus in the Diagnosis itself.

The author needs to include a clear statement to justify and support his placing the new taxon in the Family Blochiidea as he makes several comparisons between his new taxon and other genera that are assigned to a variety of billfish families. I found it difficult to follow his train of thought (e.g. see pp. 15 and 16, and then p. 22) as he goes from the individual features he describes on the new taxon, to the comparisons with other billfish taxa that each share a different subset of those features, to his ultimate taxonomic decision on where to assign his new taxon. The phylogenetic analysis does not help as the new taxon is part of an unresolved polytomy in the strict consensus cladogram.

The statement on p. 16 that the postorbital skull elements are well-preserved and undistorted is overstated and inconsistent with the actual details of the Description, which notes incomplete and somewhat crushed/distorted, and/or fragmentary, elements, and well as the fact that many skull elements are simply not represented at all.
Validity of the findings: I am very skeptical that a billfish estimated to be nearly three meters in length is exhibiting a LARVAL condition with respect to the presence of villiform teeth, as the author suggests on p.  22.

As noted in other sections of the review, I believe a clearer and more comprehensive case can be made for the taxonomic decisions arrived at, with respect to the phylogenetic conclusions, and a more comprehensive Diagnosis should be provided.
Additional comments: Overall this is a well-executed paper describing  a noteworthy specimen that should be published. The anatomical descriptions are generally solid and the figures are clear and at a publishable standard. I do believe that the paper needs one more editorial pass by someone completely fluent in English to edit out some awkward wording and misspellings, and as I noted I am still a bit confused on the rationale for the family-level assignment and how that assignment relates to the phylogenetic analysis that is presented, and I believe the Diagnosis should be strengthened by including all mentioned and relevant taxa.


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: LOANCORHYNCHUS CATRILLANCAI GEN. ET SP. NOV., A NEW SWORDFISH (XIPHIOIDEI, BLOCHIIDAE) FROM THE MIDDLE EOCENE OF CENTRAL CHILE
Review round: 1
Reviewer: 2

Output:
Basic reporting: This is an excellent article dealing with the description of a new genus and a new species of a Blochidae xiphioid. The article is clear, with interesting results, update references (but see below) and excellent figures.

However, I would like to suggest that:
1- References should be checked;
2 - After the description of the new taxon in the "Systematic Palaeontology" (line 218), the specimen ceased to be treated by the number SGO.PV.6634 and started being treated by the generic or (better) the specific name Loancorhynchus catrillancai.
Experimental design: No comments.
Validity of the findings: Nothing to add.
Additional comments: I would suggest to the author to check the references because I did not find some in the text (cf.  Cappetta 1987, Chirino-Galvez 1993, Fierstine 2oo7, Fierstine and Starnes 2005, Kummel 1964, Muizon and DeVries, 1985....) or others in the references (Momsch and Fierstine, 2002).

Also, that after the first description of the new taxon, the specimen ceased to be treated by the number SGO.PV.6634 and started being treated by the generic or (better) the specific name Loancorhynchus catrillancai.

There are still some minor typos throughout the text.

Finally, I would like to congratulate the author for 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: LOANCORHYNCHUS CATRILLANCAI GEN. ET SP. NOV., A NEW SWORDFISH (XIPHIOIDEI, BLOCHIIDAE) FROM THE MIDDLE EOCENE OF CENTRAL CHILE
Review round: 1
Reviewer: 3

Output:
Basic reporting: The article meets the editorial standards in my opinion. It is properly organized in standard format for a paleontological descriptive work, including the systematic portion. I agree that it qualifies as a new genus and species; the description meets prevailing standards. The bibliography is generally sufficient, but requires some relatively minor revisions, as noted in my attached word file. That is, in fact, where most revisions are needed. I have also noted other typographical and style comments, notably for standard English expressions. If the editors wish me to do so, I will be pleased to review the article again following revision.

	REVIEW OF: LOANCORHYNCHUS CATRILLANCAI MANUSCRIPT
TYPOGRAPHICAL AND STYLISTIC NOTES:
Line 18: these being (conventional English sequence)
Line 22: Omit the word “studied”
Line 23: such a unique (conventional expression)
Line 24: Omit the word “studied”
Line 28: Omit the word “studied”
Line 71: crop out (conventional expression)
Line 79: Is Sernageomin, 2003 in the bibliography?
Line 114: Spell out generic name.
Line 114: phragmocone (usual English spelling)
Line 133: collection (not recollection)
Line 159: spelling of Schultz?
Line 169: What is the date of the Nakamura reference 1985? 1995?
Line 173: Should this be Fierstine and Monsch?
Line 181: spelling of Schultz?
Line 263: Abbreviation should be “approx.”
Line 265: based on the (conventional expression)
Line 267: Should this be Fierstine and Monsch?
Line 274: Prefer “nutrient” to “nutritious”.
Line 302: there (spelling)
Line 310: “the”, rather than “their”
Line 355: Is Fierstine, 1972 in the bibliography? (also Lines 347, 348, 355.)
Line 378: Should this be Fierstine and Monsch? (also Lines 382 and 394.)
Line 394: Check Nakamura publication date (as for Line 169).
Line 401: Omit word “shows”.
Line 419: Was Paleogene intended (instead of Paleocene)
Line 421: Should this be Gottfried et al.?
Line 425: Special thanks are due to H. Fierstine (conventional expression)
Line 429: for valuable literature (conventional expression)
Line 449: was this cited?
Line 458: Was this cited?
Line 459: Kent, Ohio, U.S.A.
Line 458: Was this cited?
Line 473: Capitalize Scombroidei, Middle, Eocene, Monte, Miscellanea
Line 476: Was this cited?
Line 506: Was this cited?
Line 519: This is not entered in alphabetical order.
Line 533: Check date of reference.
Line 541: Was this cited?
Line 550: Was this cited?Line 560: Was this cited?
Line 563: Enter entire reference.
Line 577: Was this cited?
Line 599: Should this be Fierstine and Monsch?
Figure 2 Caption. Should this be Fierstine and Monsch?
Experimental design: As noted previously, the organization of the paper is standard for descriptive and systematic works. The description is very thorough.
Validity of the findings: I agree with the conclusions, that is, that it is a new genus and the corollary conclusions are valid.
Additional comments: I have no further 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: CAPACITY FOR HEAT ABSORPTION BY THE WINGS OF THE BUTTERFLY TIRUMALA LIMNIACE (CRAMER)
Review round: 1
Reviewer: 1

Output:
Basic reporting: The article is written in clear professional English and is clearly understandable throughout. The background of the study is properly introduced and the literature cited fits the broader field of the study. A formal analysis of butterfly thermoregulation is currently lacking in literature, as well as a quantification of the contribution of dark wing areas to the thermoregulatory process. Hence, the article clearly contributes to fill a gap in literature. However, in its current state the article has some remarkable flaws in the way results are presented making it unsuitable for publication.
Experimental design: Methods are described in sufficient detail. However, in some cases it is not completely clear whether the butterflies used were alive or dead. Hence, more details should be provided regarding the condition of the butterflies tested.
Validity of the findings: The structure of the article mostly conforms to the standards of scientific publications, but there are few remarkable exceptions: the second half of the results section, which is also the one including the focal findings of this paper, is built upon data inferred from figures rather than based on a statistical analysis formally quantifying the effects asserted. Based on the raw data provided, the authors have the data for a thorough statistical analysis, but this was not performed. For example, the fact that the so called heat storage areas heat more than other wing areas should be statistically tested, and figure 11 should only serve as an example. For all the data referring to figures 12 to 14, the authors should perform a time-series analysis, or at least compare the slopes of the different curves during the linear phase, and compare the final temperatures after they plateau. A statistical analysis formally supporting the findings is essential for publication.
Another weakness of the paper is the lack of references to the ecological relevance of the measurements assayed. First of all, it is not clear (at least to me) why this butterfly is a good model organism to study heat absorption. In addition, the authors test light intensities ranging from 600 to 60 000 lx, even though all butterflies die at intensities higher than 40 000 lx. This result should be extensively discussed in the light of the natural conditions the butterfly faces in the wild. Does this mean that butterflies do not bask during the warmest times of the day because of lethal light intensity? Or could it be that the halogen lamps impact the butterflies more heavily than natural conditions? I encourage the authors to discuss these points as critically relevant to the focus of the paper.
Conclusions are well stated and linked to the research question, but they are flawed to the extent that they are partially based on results not supported by an adequate statistical analysis. But this issue will disappear once the authors provide adequate statistical tests for their findings.
Finally, there was little or no attempt to link the findings to the natural basking and thermoregulatory behaviors of the butterfly (e.g. are the optimal wing opening angles found here observed more commonly by naturally basking butterflies?).
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: CAPACITY FOR HEAT ABSORPTION BY THE WINGS OF THE BUTTERFLY TIRUMALA LIMNIACE (CRAMER)
Review round: 1
Reviewer: 2

Output:
Basic reporting: Expression could be revised for minor grammar points. The results are horrendously complex and laboriously presented - some sections are almost impenetrable for the repetition of raw statistical tests.
Experimental design: See general comments.
Validity of the findings: See general comments.
Additional comments: I was interested in this manuscript in terms of how it seeks to define exactly how heat exchange occurs in butterflies. The question is of general curiosity for workers in Lepidoptera. The focus is heavily mechanistic; therefore the title may better reflect this. Overall there are useful insights here, but the presentation is horrendously complex and laborious. The results section “Effects of light intensity on the heat absorption capacity” is simply unreadable. Very few readers could be expected to extract the principal findings. I strongly suggest that this material be better incorporated in Figures or Tables. The authors have obviously invested an enormous amount of work in this study, which is highly commendable. The presentation however may not generate the highest or broadest impact because it is highly specific to the taxon, but it is worthy of publication in my view.

Minor:
Line 34 (and throughout): It is not just flight that body temperature determines in these insects – indeed it is their greater metabolic rate, and hence all metabolic processes (in females this may determine the rate of ovarian development, for example).

Line 46: the extensive literature on “reflectance basking” by Kingsolver and colleagues is mentioned later, which is great, but it could perhaps be also stated here.

“Light intensity” should be simply referred to as such, and not “The light intensity”

Line 271: The term “double-factorial analysis of variance” is unclear if not unconventional. The statistics should be phrased in terms of general linear modelling, which more broadly encompasses what used to be referred to as one-way & factorial ANOVA, etc.

The study relies on light intensity in lux as the basic unit of illumination. Lux is a measure of intensity that is “calibrated” for human perception. It would be unequivocal to use this under various sunlight conditions, but it may deviate from heat gain under artificial lighting. The authors should recognize this caveat in their argument.