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: RANDOM FOREST AS A GENERIC FRAMEWORK FOR PREDICTIVE MODELING OF SPATIAL AND SPATIO-TEMPORAL VARIABLES
Review round: 2
Reviewer: 1

Output:
Basic reporting: I choose not to evaluate the 2nd version for reasons which I have communicated to the editor.
Experimental design: I choose not to evaluate the 2nd version for reasons which I have communicated to the editor.
Validity of the findings: I choose not to evaluate the 2nd version for reasons which I have communicated to the editor.
Additional comments: I choose not to evaluate the 2nd version for reasons which I have communicated to the editor. Having seen how the authors responded to my concerns about the original preprint, further comment would obviously be pointless and only lead to another argumentative response.


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: RANDOM FOREST AS A GENERIC FRAMEWORK FOR PREDICTIVE MODELING OF SPATIAL AND SPATIO-TEMPORAL VARIABLES
Review round: 2
Reviewer: 2

Output:
Basic reporting: Good
Experimental design: Good
Validity of the findings: Good
Additional comments: The authors have addressed most of the comments I raised in my initial review. A good reference that explains that KED conducts a local fitting of the regression model within each search window are the books by Hans Wackernagel (1995) and Pierre Goovaerts (1997).


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: DIVERSITY AND EVOLUTION OF THE ENDOSYMBIONTS OF BEMISIA TABACI IN CHINA
Review round: 1
Reviewer: 1

Output:
Basic reporting: The research is straight forward and not complicated. The manuscript is nicely written. How ever, the authors should pay more attention to some details.
For instance:
1 The genus and species names in literature references should be italic.
2 In figures 4, 5, 6, the names of symbionts should be explained well. The names of insect hosts should be italic. The authors should show full genus names of the insect host for the first time.
3 The new identified symbionts should have host information like "Bemisia tabaci China1 XXX".
4 The bold symbols had better have clearer notes on phylogenetic figures.
5 Line 69, Orientia is not the newly found S-symbiont in B. tabaci. It should be checked again.
Experimental design: I am curious why the authors only investigated 2 of the 7 secondary symbionts in B. tabaci. Do they have any special interest? They may be discussed in the revised manuscript.
Validity of the findings: 1 It's unprofessional to show "MEAM1/B" in the abstract. Full names should be shown for the first time shown in a paper. The biotype B or Q is not helpful here and can be deleted.
2 Figure 3, Bemisia samples should be noted so that they are different from T vaporariorum.
3 In the abstract, Line 27- 30, "phylogenetic analyses revealed that the P. aleyrodidarum and mtCO1 (mitochondrial cytochrome oxidase 1) phylograms were highly identical and corresponding with the five distinct cryptic species clades, indicating an ancient infection followed by strict vertical transmission and subsequent co-evolutionary diversification" . But,  this is not supported well by data shown in Figure 3 and Figure 4. The phylogeny of Portiera is not clustered well.
It should be better to build a figure that shows whether the two trees have evolutionary consistence.
Additional comments: The authors should discuss more on the findings. Although the text was corrected by native speakers,  the figures and details were obviously need improvement.


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: DIVERSITY AND EVOLUTION OF THE ENDOSYMBIONTS OF BEMISIA TABACI IN CHINA
Review round: 1
Reviewer: 2

Output:
Basic reporting: No comment
Experimental design: No comments
Validity of the findings: No comments
Additional comments: The research paper “Diversity and evolution of the endosymbionts of Bemisia tabaci in China” by authors Tang and Cia et al., describes the prevalence and diversity of one primary and two facultative symbionts in china. Authors samples 1501 B. tabaci individuals which belong to two invasive species (MEAM1 and MED) and three indigenous species (Asia II1, Asia II 3 and China 1). Though the authors collected good number of samples at 71 geographical locations across the China, my main concern of this study is why authors focused only on primary and two secondary endosymbionts (Arsenophonus and Cardinium), What about other secondary endosymbionts?. At present at least 40 morphologically indistinguishable species were reported from B. tabaci cryptic species (De Barro et al., 2011, Dinsdale et al., 2010 and Hu et al., 2017); authors should update their species information by citing above reference papers. As other studies have already shown endosymbiont diversity at an intra-species level, this study does not bring much information as other studies. Authors might have focused on all endosymbionts to study the strict vertical/horizontal transmission and co-evolution of endosymbionts in their samples collected. As a summary, no significant novel information are presented in this manuscript, except collecting and screening huge number of samples across the China. Also authors should have been performed phylogenetic analysis more than one more marker to represent evolutionary relationships of endosymbionts and B. tabaci. I would have made an interim decision for “major revision” if authors focused at least known five endosymbionts and more than one marker  to study the evolutionary relationships.


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: DIVERSITY AND EVOLUTION OF THE ENDOSYMBIONTS OF BEMISIA TABACI IN CHINA
Review round: 1
Reviewer: 3

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The manuscript by Tang and co-workers represents a systematic survey of three symbionts (Portiera, Arsenophonus and Cardinium) of Bemisia tabaci in China. The authors provide a valuable survey with wide coverage. The study is valuable for providing wide coverage and it is an effective supplement to the present situation of symbionts of B. tabaci in China.

Following, my comments on the manuscript:

In this study, the author rarely discussed other s-symbionts, such as Wolbachia and Hamiltonella, the later distributes widely in invasive species MED/Q and MEAM1/B.

As we known, some biotypes of whitefly have co-infection symbiont system. The dual endosymbionts showed important mutually dependent to provide a full complement of nutrients to their host whiteflies (ref.1), and effect on the fitness and biology of the Bemisia tabaci (ref.2). In the discussion, the relationship between the different symbionts might be discussed.
Ref.1 Rao Q, PA Rollat-Farnie, DT Zhu, et al. Genome reduction and metabolic complementation of the dual endosymbionts in the whitefly Bemisia tabaci，BMC Genomics，2015, 16:226.
Ref.2 Ghosh S, Bouvaine S, Richardson S C W, et al. Fitness costs associated with infections of secondary endosymbionts in the cassava whitefly species Bemisia tabaci. Journal of Pest Science, 2018, 91:17-28.

From figure 3 and figure 4, the relationship of clusters between P. aleyrodidarum and cryptic species is not obviously clear. The genetic variation of primary symbiont might not be an ideally reflect the genetic variation of this cryptic B. tabaci.

Line 155-157: “The results showed that 21 haplotypes were identified in whiteflies based on mtCO1 sequences, while 17, 7 and 8 symbiont haplotypes were defined based on analysis of P. aleyrodidarum, Arsenophonus and Cardinium sequences, respectively”. How to identify the haplotypes of whiteflies and symbiont haplotypes? What these claim are based on?

Line 170-172: “It is interesting that the phylogenetic tree of P. aleyrodidarum were highly similar to the mtCO1 tree and exhibited four groups corresponding to MEAM1/B+MED/Q, Asia II 1, Asia II 3 and China 1 clades (Fig. 4).” From figure 4, the sequences of P. aleyrodidarum of China1 are not clustered.

Line 222-224: “furthermore, the spread of these two invasive species in China has been frequently associated with founder effects that fix specific mtDNA variation(s) along with particular endosymbionts”, the sentence is not very clear.

Figure 2B and 2C: the total number of each cryptic B. tabaci on the bar graph is not necessary, because all samples are mentioned. The number of infection should more interest for audience.


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

Input:
Title: DIVERSITY AND EVOLUTION OF THE ENDOSYMBIONTS OF BEMISIA TABACI IN CHINA
Review round: 1
Reviewer: 4

Output:
Basic reporting: The manuscript by Tang et al. investigated the distribution frequency of the primary and two secondary endosymbionts of five species of Bemisia tabaci from China, and then outlined the co-evolution of the mutualists by phylogenetic analysis.
Experimental design: no comment
Validity of the findings: The data seem to provide some new insights into the endosymbiotic status and evolutionary relationships in whitefly.
Additional comments: 1.	In B. tabaci whitefly, seven secondary endosymbionts have been identified and many of them have abundant distribution, why the authors only concerned Arsenophonus and Cardinium rather than others in this study?

2.	Phylogenetic trees of Portiera and Cardinium were constructed with 16S rDNA sequences, while 23S rDNA was used for Arsenophonus. I wonder why different genes were chosen for phylogenetic analysis in different bacteria.

3.	Line 34-35: The vertical and horizontal transmission of symbionts was speculative and no direct evidences were provided in this study, please check your expression and make it unambiguous.

4.	Line 195-200: The interpretation of low frequency of symbionts was appeared to be not sound. Reproduction manipulation usually facilitates spread of symbionts in the hosts and sometimes may benefit their hosts, such as high proportion of female. Besides, the assumed negative effects of the symbionts can’t explain why those bacteria have high prevalence in native species but low frequency or absence in invasive species.

5.	Line 206-208: The diversity of symbionts has been well studied in B. tabaci, and the co-infection of the Cardnium and Arsenophonous has also been reported in previous studies (Bing et al., 2013; Chu et al., 2011; Parrella et al., 2014; Zchori-Fein et al., 2014).

6.	The text of scientific names of organisms should be italicized and the style of references should be consistent.

7.	In figure 1B, the native species can be labeled separately with different symbols.

8.	In table 2, the total number of  mtCOI haplotypes should be 21, please check and revise. How did authors calculate the percentage of each haplotype, by sequencing all samples or other methods?


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: DIVERSITY AND EVOLUTION OF THE ENDOSYMBIONTS OF BEMISIA TABACI IN CHINA
Review round: 2
Reviewer: 1

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


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

Input:
Title: DIVERSITY AND EVOLUTION OF THE ENDOSYMBIONTS OF BEMISIA TABACI IN CHINA
Review round: 2
Reviewer: 2

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


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

Input:
Title: CONTEXTUAL AND INDIVIDUAL DETERMINANTS OF ORAL HEALTH-RELATED QUALITY OF LIFE AMONG FIVE-YEAR-OLD CHILDREN: A MULTILEVEL ANALYSIS
Review round: 1
Reviewer: 1

Output:
Basic reporting: Except for minor points the english language used is clear and unambiguos.
the article  structure is fine. However,I have some suggestions for the improvement of the article:
Introduction:
1. The introduction is too long. An introduction should not exceed one page with 1.5 spacing.
Line 57: clinical conditions is very general,... do you mean oral diseases?
Line 59: The sentence about SOC is completely unclear, unspecific. Why and how is stress reflected in the oral health of an individual?
Line 61: which control of life events is related to oral health? to vague formulated.
Method
The part on Data collection should be sub-divided into individual determinants, contextual determinants and outcome.
Line 135: Why is the pilot study relevant for this research(-question)? Did you use for the calibration of researchers? No? why do you mention it, it does not add anything to this study...
Line 156: do you mean 'anytime'? or at least once? Unclear!
Line 204: code 1 is not used in epidemiological studies. This is a very common sentence... What did you do in your study?

Results:
The prevalences of oral health conditions (except for malocclusion) appear very high for 5-year old children. Table 1 could be presented stratified by private/public schools....This would already point towards the main result of the study and would make the data more transparant for the reader.

The ES for houshold income appears odd. How did you measure houshold income? Did you use it continously or categorical?You don't describe it at any point in your study, although you mention it as one of the most important determinants. You also describe it differently in model 1 and model 2 (monthly houshold income vs lower household income). Please stay consistent.

Tables:
The legends of the tables need to be more specific about which covariates are used in the model

I suppose you mean CI instead of IC?

Dont present intercept, this is not relevant information for the research question

Dont present Deviance in the table, this is not relevant information for the research question

What is the difference between model 1 and the RR given in table 2? Table 2 and 3 should be combined into one table

References

More diverse articles on the topic should be included, for example also from Europe. There are defenitely articles on the association of individual/environmental factors related to OHRQOL in children. Now only articles from Brazil are included... many based on the same data/from the same research groups....
Experimental design: Generally well done. However some suggestions/comments/questions:

Sampling: There are more private than public school registered with the ministry of education. Why in the end do you include more public than private schools in your study? Do you put your study at risk for selection bias in this way? Make this clear in table 1 and eventually discuss this in the limitations of your study.

Sample size: How did you derive at a 1.6 design effect? And what do you mean with a 50% prevalence of the impact on OHRQOL? Any impact? How many points under maximum? Unclear!

Statistical analysis: What about the correlations between individual and contextual factors? Have you test it? How did you deal with it?
have you tested for interaction between all covariates/ determinants and type of school? What were the results?
Validity of the findings: Generally good. However based on the comments above and some extra analysis they should (eventually) be adjusted. For now the validity of the findings are not yet sufficiently supported.
Additional comments: Nice and interesting study!
general comments:
1. In a sentence always first mention determinant and then outcome. In this way it becomes much easier to follow for the reader.
2. Why do you spent so much text on LOC and SOC? In the end it does not add anything to your study, as you even do not use it. You should only describe it in the Discussion.


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: CONTEXTUAL AND INDIVIDUAL DETERMINANTS OF ORAL HEALTH-RELATED QUALITY OF LIFE AMONG FIVE-YEAR-OLD CHILDREN: A MULTILEVEL ANALYSIS
Review round: 1
Reviewer: 2

Output:
Basic reporting: Introduction is well written. Literature is well referenced and relevant The subject studied is relevant and original.
Experimental design: Research question is well defined. Methods were described with sufficient detail e information to replicate.

Sample and study design

I did not understand the calculation of the minimum sample. The minimum
sample was determined to be 615 children. This value was increased to compensate for possible dropouts estimated at 20% resulting in a sample of 769 (it would not be 738?) five-year-old children. 769 children were the final sample and not minimum sample.

Clinical examination

I suggest clarify the criteria for the diagnosis of tooth wear.
Validity of the findings: Data is robust, statistically sound and controlled.

Results

Table 1:

Some variables don’t add 769 children. I suggest include the missing data in the footnote.

Discussion
I would like to know what the authors think about malocclusion is not associated with the dependent variable (OHRQoL based on the perceptions of children).
Additional comments: I accept the manuscript with minor changes.


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: CONTEXTUAL AND INDIVIDUAL DETERMINANTS OF ORAL HEALTH-RELATED QUALITY OF LIFE AMONG FIVE-YEAR-OLD CHILDREN: A MULTILEVEL ANALYSIS
Review round: 2
Reviewer: 1

Output:
Basic reporting: Introduction is well written. Literature is well referenced and relevant The subject studied is relevant and original.
Experimental design: Research question is well defined. Methods were described with sufficient detail e information to replicate.
I understand the calculation of the final sample now and the criteria for the diagnosis of tooth wear is clear too.
Validity of the findings: Data is robust, statistically sound and controlled.
A footnote has been added to Table 1 to clarify the missing answers.
The authors included some information explaining why malocclusion was not associated with the children’s perceptions regarding OHRQoL,
Additional comments: I accept the article without alteration


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

Input:
Title: EFFECTS OF MESOSCALE EDDIES ON BEHAVIOR OF AN OIL SPILL RESULTING FROM AN ACCIDENTAL DEEPWATER BLOWOUT IN THE BLACK SEA: AN ASSESSMENT OF THE ENVIRONMENTAL IMPACTS
Review round: 1
Reviewer: 1

Output:
Basic reporting: Basic reporting and general comments
The manuscript does not show clear, unambiguous, professional English language used throughout.
Intro & background show context. Literature is well referenced & relevant.
However, the manuscript is not well structured.
Some figures contain mistakes (e.g., Fig. 2, 7) and do not have high quality (e.g., Fig. 6, 9, 11). Figure captions are not always exhaustive (e.g., Fig. 6, 8, 9).
Some equations and model specifications are represented in a confusing way (e.g., Eq. 2. and boundary conditions in string 337).
Research question is well defined, relevant & meaningful. It is stated how the research fills an identified knowledge gap. But permanent pycnocline as an important Black Sea feature, that influences the vertical droplet transport, is not mentioned.
Rigorous investigation is performed to a high technical & ethical standard.
However, modeling method is not transparent, which makes it impossible to replicate.

Specific Comments
1.	String 111: Please explain the connection between cold methane seeps (yellow squares in Fig. 1) and possible deepwater oil blowouts.
2.	String 237: Which diameter droplets do raise at 0.07 ms-1?
3.	String 251: Please focus on the processes modeled by you not by Sokolofsky. Point out that near-field modeling the buoyant jet is beyond your model scope.
4.	String 275: Where is buoyancy flux?
5.	String 306: Please draw the profile/s of V3(x3) and K3(x3).
6.	Figure 2: Please correct: Bosphorus, Kizil Irmak. What are dotted arrows on the NW shelf denoted?
7.	Figure 7, 10: Please correct scale bar.
8.	Figure 9, 11: Please redraw in color.
9.	Some figures (e.g., Fig. 1-3, 8) have been already published e.g., in Korotenko (2016). Please insert the reference to the published graphics.
Experimental design: My opinion is expressed in the previous section.
Validity of the findings: My opinion is expressed in the previous section.
Additional comments: My opinion is expressed in the previous section.


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

Input:
Title: EFFECTS OF MESOSCALE EDDIES ON BEHAVIOR OF AN OIL SPILL RESULTING FROM AN ACCIDENTAL DEEPWATER BLOWOUT IN THE BLACK SEA: AN ASSESSMENT OF THE ENVIRONMENTAL IMPACTS
Review round: 1
Reviewer: 2

Output:
Basic reporting: No comment
Experimental design: No comment
Validity of the findings: No comment
Additional comments: 1. In the introduction the author should refer briefly to the “Oil Platform  Leaks”challenge of the EU EMODNET Black Sea Check point activities, which are of relevant to this ms.

2. In deep oil spill model section, provide the corresponding API number of the used light oil type for the simulations. This is important, especially with the  Fig. 11.

3. The sentence in lines 233-234 to be modified as: “It is apparent from Eq. 1, the more size of an oil droplet and greater density difference between oil and ambient water the more terminal velocity of the droplet”.

4. Provide an additional plot, similar to the Figure 11, indicating the % of the modeled fate parameters.

5. In line 321 correct the typo error : “released”

6. While in line 335 it is mentioned that the evaporation act within the first few days, in the Fig. 11 the plot concerning the evaporation shows that the evaporation is a process increased by time, more than few days. Explain or correct.

7. In lines 382-384, indicated the period/years of the used climatological T and S data for the hydrodynamical simulations.

8. In lines 470-474, the estimated time (3.7 hours) needed for  the first oil drop to reach the  sea surface from ~1000 meter seems to be large. I propose to look the recent paper by Lardner and Zodiatis 2017 mentioned below.

9. Studies regarding the oil plumes modeling and oil spill modeling in the Black Sea can be found in the following papers and I suggest to the author to have a look and include them at the references:

Lardner R. and G. Zodiatis (2017). Modelling oil plumes from subsurface spills, Marine Pollution Bulletin , http://dx.doi.org/10.1016/j.marpolbul.2017.07.018

Zodiatis G. and Lardner, R. and Solovyov, D. and Panayidou, X. and De Dominicis, M. (2012). Predictions for oil slicks detected from satellite images using MyOcean forecasting data. Ocean Science, Volume 8, Pages 1105-1115.

Yapa, Poojitha, Zheng, Li, 1997. Simulation of oil spills from underwater accidents I:
model development. J. Hydraul. Res. 35, 673–687.

Zheng, Li, Yapa, Poojitha, 1998. Simulation of oil spills from underwater accidents II:  model verification. J. Hydraul. Res. 36, 117–134.


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

Input:
Title: EFFECTS OF MESOSCALE EDDIES ON BEHAVIOR OF AN OIL SPILL RESULTING FROM AN ACCIDENTAL DEEPWATER BLOWOUT IN THE BLACK SEA: AN ASSESSMENT OF THE ENVIRONMENTAL IMPACTS
Review round: 1
Reviewer: 3

Output:
Basic reporting: Language and Length of paper

There are some language problems. Already in the title:
Effects of mesoscale eddies on behavior and environmental impacts of oil spilled in result of an accidental deepwater blowout in the Black Sea
The phrase “In result” – should usually be “as a result”. E.g. [Title, plus Line 42, 65, 116, 141…]
I suggest: Effects of mesoscale eddies on THE behavior of oil spilled AS A RESULT of an accidental deepwater blowout in the Black Sea and its environmental impacts

[Lines 12-13]: employing numerical modeling is not exactly ‘easy’. I would suggest to cut that word, and rewrite the sentence.
Suggest: Employing numerical modeling as a prediction tool is one of the most efficient methods to understand oil spill behavior under various environmental forces.

[Lines 27-28] What does a 20-day scenario tell us? Explain that this is the time needed to see CNAEs evolve, and oil gets captured and transported by the AEs.
[Line 76]: “disastrous events, they seem to be inevitable due to a number of reasons”. …Such as? What could be a typical reason for a disastrous event in the Black Sea? This is not the same as e.g. the Gulf of Mexico, where many hurricanes hit.
[Line 79-82]: “The Black Sea is a water body that, owing to its semi-isolation from the open ocean, suffers from strong ecological disequlibria caused by pollution arising from many contaminants”,. Explain clearer what the disequlibria lie in? Rare flushing, strong stratification, …, thus contaminants may have a long-term effect…
[Line 83-88]: this should be rewritten, it does not flow well. Also, you mention that there are very many natural seeps in the BS, so obviously not only accidental oil spills posed a problem in the past.

[Lines 122-123]: repetition from the abstract.
[Lines 126-129]: repetition from the abstract

[Line 361]: high-resolution

I think that the scenarios chosen could be mentioned earlier, the introduction is too long and wordy. Tighten the introduction in order to get to the main interest of the paper. In general the paper is too long considering it’s in reality only describing a couple of scenarios
Experimental design: I cannot see that this paper lies within the scope of the PeerJ journal. Oil spill modeling is a physical/chemical discipline that involves numerical geophysical modeling of atmosphere, ocean, and waves, as well as a knowledge of oil chemistry and oil weathering processes.

The paper aims to address impacts on the marine environment, but only does so to the extent of evaluating whether certain regions of the coastlines of the Black Sea might be struck by oil in case of an accidental blowout under certain weather conditions. Even if this is an interesting subject matter, the study cannot really say anything about environmental impacts as long as it does not link to any scientific knowledge about the particularities of specific vulnerable ecosystems or fragile water systems around the Black Sea.

As such, I am not sure that this paper will find an interested audience within this particular journal. The author has previously published in scientific journals such as “Oceanology”, “Spill Science and Technology Bulletin”, “Terrestrial, Atmospheric and Oceanic Sciences”, “Journal of Marine Systems”, etc. Although Peer J aims to cover Environmental Sciences, the topics therein do not cover geophysics, but rather biology and ecology.

I would say that the methods have been sufficiently described to replicate, but are lacking in that other models and methods could have been used that would have provided more conclusive results. E.g. statistical runs (meaning many runs over several seasons) and more locations spread along interesting/potential drilling sites. This will hopefully be addressed in future studies.

Since the DieCast model does not include data assimilation, it can be very tricky to forecast the exact placement of eddies in time and space, and as such, the model is better used for process studies. For oil spill contingency and preparedness, it would be useful to have a more rigid assessment of the potential extent of shoreline contamination, combined with maps of ecologically vulnerable parts of the shoreline, as well as practical advice on where to place depots for oil spill cleanup efforts.

The intro and background show context, and the literature referenced is comprehensive, and shows that the author knows the context pretty well with regards to the geophysical modeling. However, the use of the oil spill modeling shows a simplistic approach and a basic knowledge of the field of oil weathering processes that have partly been taken from outdated literature (e.g. the oil is presented as a mixture of 8 hydrocarbon groups following Mackay & McAuliffe, 1988, where removal of oil through the evaporation process is linear over a time of 20 days (see figure 11), whereas in reality, a light, crude oil such as used in this study tends to evaporate away by 25% within the first few hours, 75% within the first few days. (E.g. D. Mackay, R.S. Matsugu, Evaporation rates of liquid hydrocarbon spills on land and water, Can. J. Chem.Eng. 51 (1973) 434-439.)
The mass balance of the oil therefore will necessarily be unrealistic after 20 days, and thus the assessment of the scale of contamination of marine environment, that was one of the aims of the study, will be wrong. Other important weathering processes, in particular emulsification, is not included in the model. As for the natural dispersion, he amount of dispersed oil seems very large. Is your droplet size distribution correct?

 [Lines 133-138]: You say that “in deep waters, the movement and fate of a multiphase plume is governed by the gas-oil separation process, rising velocity as well as background currents and stratification, while at the subsurface layer, the plume evolution and its fate experiences the influence of currents induced by local winds, Stokes drift and physicochemical processes which change the oil properties.”

I can’t see how the Stokes drift, or any wave parameters have been used at all in this study, although it is mentioned that waves are input to the to the Deepsea Oil Spill Model (lines 195-195).
I can’t see that you have discussed the effects of vertical mixing on the transportation of the oil, although this is very important for the overall transportation of oil as well as the oil mass budget.
Validity of the findings: The aims of the study are:
1)	To “elucidate the behavior of the subsea plume” – a simple plume model is set up, but the scope of the study does not add anything new to the general knowledge of deep sea plume modeling. (Multi phase aspects are lightly skipped, deep sea formation of gas hyrates is not mentioned as a problem in modeling)
2)	To assess scales of contamination of marine environment and coastlines in possible accidental blowouts – with two scenarios studied of a hypothetical deep sea oil well blowout placed at one location on the Shatsky Ridge, it is hard to say that any scale of contamination has been assessed. I’d rather say that a model has been set up that might work to do such an assessment in a much wider study.

I would not say that the investigation has been rigorous enough to address the research aims stated in the introduction, but would view it more as an initial effort giving some answers to how well the oil spill model performs.

The role of the CNAEs is illustrated well, in the way a “highly “oiled” eddy would extend and move near the coasts, depositing massive amounts of oil and polluting the Caucasian and Crimean beaches.
Why has only winter situation been studied in the 2 scenarios? [Lines 618-619]. For impact studies, it might be useful to only use climatological winds, and rather study different periods of the year with known differences in circulation patterns

Most of the figures are relevant and clear, but I find some figures missing for comparison between the climatological wind situation and the NCEP daily averaged wind situation. E.g. figure 9 showing successive vertical plots of the plume and surface spread of oil against the wind sticks for NCEP winds. Why not have the same plot for the climatological winds? What does that mean? Are the climatological winds not of high enough resolution in time?
Figure 11 is an important figure, showing the oil mass budget over the 20 day scenario. It would be easier to interpret this figure if it were in colour.
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: EFFECTS OF MESOSCALE EDDIES ON BEHAVIOR OF AN OIL SPILL RESULTING FROM AN ACCIDENTAL DEEPWATER BLOWOUT IN THE BLACK SEA: AN ASSESSMENT OF THE ENVIRONMENTAL IMPACTS
Review round: 2
Reviewer: 1

Output:
Basic reporting: I could not see any major revisions made by the author:
(1)	English language is really awful.
(2)	There are not any new findings in the manuscript. Horizontal transport of the oil droplets is well known. Any horizontal intrusions of droplets are not visible (Fig.6). Influence of the Black Sea permanent pycnocline is neglected because the rise velocities of droplets are very high, as the author confirms in his response below.

“5. String 306: Please draw the profile/s of V3(x3) and K3(x3).
A. Let I omit these profiles since for the used version of the model they have not much information but substantially boost the text.
Explanation: (1) Because the vertical velocity throughout the water column of is very small (~10-5 m/s) compared to the droplet terminal velocity, the former will have a little effect on the vertical displacements of oil droplets and everything would be determined by terminal velocity”

(3)	 V3(x3) and K3(x3) profiles, which are extremely uncertain in the Black Sea and are relevant to the vertical transport of small droplets, are not shown.
Unfortunately, my conclusion is to reject.
Experimental design: Everything I want to say is written in the Basic reporting Section
Validity of the findings: Everything I want to say is written in the Basic reporting Section
Additional comments: Everything I want to say is written in the Basic reporting Section


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

Input:
Title: EFFECTS OF MESOSCALE EDDIES ON BEHAVIOR OF AN OIL SPILL RESULTING FROM AN ACCIDENTAL DEEPWATER BLOWOUT IN THE BLACK SEA: AN ASSESSMENT OF THE ENVIRONMENTAL IMPACTS
Review round: 2
Reviewer: 2

Output:
Basic reporting: Figures have been improved, and the text is clearer. I assume that PeerJ has an editor that can help improve the English once the paper has been approved for publication.
Experimental design: No comment
Validity of the findings: No comment
Additional comments: I thank the author for his replies to my comments and questions. Several points have been improved and clarified in this latest version of the paper, and choices made have been justified.


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: ENERGY COST ASSOCIATED WITH MOVING PLATFORMS
Review round: 1
Reviewer: 1

Output:
Basic reporting: The introduction provides a good overview of the previous literature on platform motions and the impact on biomechanical, physiological, and psychological demands. Lines 67 - 76 provide a high-level descrpition of why motion-rich environments are important to examine. More detail should be provided early in the introduction as to which occupations are/would be greatly impacted. The information in these lines does not provide a clear picture to the reader which occupations the upcoming study would be most relevant to.

Line 93 - change increase to increased

Lines 103 - 105 - provide additional details as to the specific increases that were observed in oxygen uptake and what were the expected levels by the authors.

The authors provide a clear message as to the knowledge gap in this area with a clearly stated hypothesis.
Experimental design: Line 126 - should the university's entire name be listed, i.e Memorial University of Newfoundland?

Line 161 - change has to had

Line 166- add 'the' after task (throughout the task 'the' participant)

Line 166 - change 'the' to 'their'

Lines 172 - 174 - is it possible to include an image or images of the experimental setup revealing the various position the participants were placed in and the nature of the lifting tasks?

Line 183 - change increases to increase

Line 188 - suggest using significantly different throughout manuscript instead of statistically significantly difference

Line 187 - is the significant difference referring to the LM compared to the control condition and the HM compared to the control condition? Could the authors please explain why a students' t-tests were utilized instead of an ANOVA with 3 levels of conditions and 2 levels of direction?

Lines 203 – 211 – Please provide additional details as to how the VO2, VCO2, breathing frequency, and tidal volume data was analyzed. Data was collected using a breath-by-breath system, how was raw data handled, analyzed, and outliers removed?

Lines 245 – 248 – please reword sentence starting with “Significant differences in the main effect….”, confusing to follow what the significant differences are.
Validity of the findings: The authors provide a conclusion that seafarers may have significantly increased EC due to working in motion-rich environments. This conclusion is supported by the data presented on EC from each of the different conditions. However, the authors describe collecting data on VCO2, RER, breathing frequency, and tidal volume but do not represent the data in the manuscript. This information may provide additional insight into the energy requirements for the tasks during different moving environments. Please include this information or provide details as to why this data is not included.

Line 281 - provide, in parentheses, the specific tasks that were affected by the three different motion conditions.

Line 283 - end parenthesis missing after in-text citations.

Lines 335 - 356 - This paragraph is well-written but seems to be more appropriate earlier in the discussion section.

Lines 385 - 386 - please add the kcal/min for 40% and 33%, same as provided for 50%.

Line 392 - add commas to pulling pushing and carrying.
Additional comments: Thank you very much for the opportunity to review this scientific article. The authors provide a detailed description of the impacts of platform motion on the energy cost and heart rate of operators.

General Comments

Line 410 - change thanks to thank

Figure 2 - changes 10% increases to 10% increase

Figure 3 - Remove 'statistical', can state 'significant difference'; Panel B appears to be HR data and not EC data, as listed in the caption.


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: ENERGY COST ASSOCIATED WITH MOVING PLATFORMS
Review round: 1
Reviewer: 2

Output:
Basic reporting: No comment. Overall clear writing. Minor suggestions for improved clarity below.
Experimental design: No comment.
Validity of the findings: The study appears sound.
Additional comments: This is a straight-forward article on the energy costs of sitting, standing, and lifting when the support surface one is standing on moves. The experiments appear to be well-designed and the results are reported thoroughly. The claims seem to follow from evidence provided.

The article is well-written overall, but I'd suggest a proof-read for minor typos. I have a few suggestions or remarks.

I suggest that there is no need to use contractions such as EC, MIF, etc. The article is short enough and these words are important enough that these contractions reduce the readability of the article. Especially EC, which is so central to the article (energy costs).

Line 177. The English alphabet X need not be used for 'times'. Please re-typeset with an appropriate symbol or perhaps this can be taken care of by the journal.

I assume that the roll-pitch-heave convention is that standard in naval architecture. As the authors know, for Euler angles, the order of the rotation matters in deciding what the current orientation of the platform is.

It was unclear to me what the exact motions applied were. The authors say in Line 182: "Linear equations used to develop all motion profiles are detailed (Equations 1-5)." It is not stated where exactly Equations 1-5 come from and how exactly their amplitudes and frequencies are modified to obtain the low motion and high motion conditions. It may be better to add variables (representing amplitude and frequencies) in Equations 1-5 and then show (as in Table 1) what the values of these amplitudes and frequencies are under different conditions. Were all 5 degrees of freedome excited simultaneously?

Perhaps as an extra panel on figure 2, I suggest that providing example plots of the applied motion for the 5  degrees of freedom.

It may be useful to mention that the motions look complex enough (e.g., pseudo-random) that humans may not be able to entrain to it. This is useful because the following article considers models of humans walking on moving platforms and suggests that under certain conditions, they may actually experience reduction in energy costs (but resulting in increased costs under other conditions).
Joshi, V. and Srinivasan, M., 2015. Walking on a moving surface: energy-optimal walking motions on a shaky bridge and a shaking treadmill can reduce energy costs below normal. In Proc. R. Soc. A (Vol. 471, No. 2174, p. 20140662).
It may be worth mentioning that the current article operates far from such entrainment regimes, thereby generally resulting in increased energy cost. In other words, it is not always the case that there should always an increase in energy costs on a moving platform.

I found the presentation of the summary data in the supplementary excel file 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: ENERGY COST ASSOCIATED WITH MOVING PLATFORMS
Review round: 1
Reviewer: 3

Output:
Basic reporting: Clear and unambiguous, professional English used throughout.
Experimental design: Original primary research within Aims and Scope of the journal.
Validity of the findings: The results interpretation and its implications should be based on these effect sizes but not on p-values so that they provides practically meaningful conclusions.
Additional comments: Manuscript#: PeerJ-reviewing-23155-v0
Title: Energy cost associated with moving platforms

General Comments
This paper is an original work that evaluated the metabolic costs associated with maintaining postural stability in a motion-rich environment.  This study was designed with scientifically sound experimental methods and approaches.  However, the manuscript can be further improved and streamlined.  First, the introduction is not so convincing how increase EC is directly related to fatigue-related injuries and accidents.  Second, the results should provide effect sizes and its confidence intervals so that these differences are not only statistically significant, but also practically meaningful (some differences appears to be within a normal variation.  Specific comments are following:

Specific comments:
Comment #1: (Introduction) As the EC and fatigue are nonlinearly related, it is less convincing how the increased EC can be directly related to fatigue or increased risks of injuries and accidents, especially in the levels found in this study.

Comment #2:  (lines 163-164) Please justify how the weight and size were determined.

Comment #3: (lines 169) How many repetitions? a total of 60 lifting for 10 minute motion?

Comment #4: (lines 182-186) Please elaborate the justification for this choice.  Why low motion was only justed in frequency but not amplitude while high motion was done for both.

Comment #5: (lines 186-188)  Does it mean that it was practically significant? Please specify the meaningful effect sizes.

Comment #6: (line 200)  Please explain the reasons.  Assume it’s due to the actuator limitations?

Comment #7: (Results) Comparisons of the dependent variables (EC and HR) should include effect size and 95% CI of the effect size, not just F and p values.

Comment #7: (Conclusions) The results interpretation and conclusions should be based on these effect sizes but not on p-values.


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

Input:
Title: ENERGY COST ASSOCIATED WITH MOVING PLATFORMS
Review round: 2
Reviewer: 1

Output:
Basic reporting: The authors have satisfied all of the review comments and made the necessary changes.
Experimental design: The authors have satisfied all of the review comments and made the necessary changes.
Validity of the findings: The authors have satisfied all of the review comments and made the necessary changes.
Additional comments: Thank you to the authors for responding to each of the comments and making the necessary changes.


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: ENERGY COST ASSOCIATED WITH MOVING PLATFORMS
Review round: 2
Reviewer: 2

Output:
Basic reporting: No comment.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: The authors have reasonably addressed all my main remarks/suggestions. This is a clearly written paper.

For some clarification questions I asked, they have clarified in their review response instead of the main paper itself, but I will leave this up to the discretion/judgement of the authors.


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: COMPOSITIONAL AND PREDICTED FUNCTIONAL ANALYSIS OF THE GUT MICROBIOTA OF RADIX AURICULARIA (LINNAEUS) VIA HIGH-THROUGHPUT ILLUMINA SEQUENCING
Review round: 1
Reviewer: 1

Output:
Basic reporting: This manuscript examined the gut microbiome of Radix auricularia, in both juvenile and adult stages. I find the topic interesting, although the premise and importance of the study is not clearly described. The introduction should clearly state why is it important to study the microbiome of this snail, and why it is important to study various life stages. The main argument of most paragraphs was unclear. Paying attention to the flow, focus, and structure of each paragraph should improve the clarity of the writing. I recommend the authors focus upon a clear topic sentence for each paragraph, with only relevant supporting sentences in the remainder of the paragraph. For example, I am unclear as to how the four sentences in lines 85-91 are related to one another, and the main point the authors are trying to convey.

The language used in this manuscript is not quite clear or professional. For example, lines 58-59. The object that "this" refers to is unclear, and the sentence should be cited. Phrases such as "a few" (line 178), "fairly similar" (line 266), and "most other" (line 323) should be avoided.

The figures are relevant, although I would consider placing Figure 1 in the supplemental material and only showing on taxonomic classification level in Figure 3.

Other specific comments:
Line 41,43,174- numbers at the beginning of sentences should be spelled out.
Line 42- species or OTUs?
Please fix types in line 52. Also, it is unclear what is meant by "Pulmonata Mollusca".
Line 84- I believe a snail species is supposed to be listed here?

Raw data is not shared.
Experimental design: As discussed above, the importance and purpose of the research questions are not well defined. With regard to sequencing and analysis, the experimental design is sufficient although replicate numbers are quite low (4 individuals per category). However, many more details are needed throughout the Materials and Methods section, particularly for the collection and assessment of the snails. For example, why were 40 individuals selected, but only 8 individuals sequenced? What methods were used to examine gonads? Where is this data? How do the authors know whether snails had aged over winter (line 107). What is meant by "gut" in line 114. Does this include the stomach? What PCR reagents were used, in what quantities, and what were the PCR conditions? What clustering methods were used? Were data rarefied? If not, please justify. Please also be sure the methods for all reported result are described. Not all diversity index calculations are described in the materials and methods. LEfSe analysis is also not described in the M&M. The abbreviations AS and JS were never defined in the text.

The description of the results is also a little ambiguous. For example, in line 208 I'm unclear what "composing 88.70% of all OTUs numbers" means. This applies to lines 205 and 214 as well.
Validity of the findings: I would recommend that the authors focus less upon the exact taxonomic composition and place more emphasis on the significance and ecological implications of the bacteria detected. For example, emphasis is placed on the presence cellulolytic species in Lines 290-293. The authors should discuss the ecological importance of cellulolytic species in the context of their results.

How is Radix auricularia different than those listed in lines 85-87,253-255? Why would their gut microbiota be different than the snails already studied? In lines 243-249, many assumptions are made that were not tested (such as similarity in diet composition). Please either prove these assumptions or remove them.
Additional comments: Assessment of functional genes was not actually carried out, rather predicted functional genes. This is an important distinction. Please change the title to read "… predicted functional analysis…" or something similar.

In line 132 QIIME is cited incorrectly.


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: COMPOSITIONAL AND PREDICTED FUNCTIONAL ANALYSIS OF THE GUT MICROBIOTA OF RADIX AURICULARIA (LINNAEUS) VIA HIGH-THROUGHPUT ILLUMINA SEQUENCING
Review round: 1
Reviewer: 2

Output:
Basic reporting: The authors describe bacterial communities from the gut of some juvenile and adult R. auricularia individuals. The authors used Illumina technology and 16S rRNA sequencing to determine the taxonomic profiles of these communities. I believe that the data are of interest as very little is known about snail microbial communities. Furthermore, a few other studies have shown that these animals are a promising source of new microbial enzymes with potential technological applications and of ecologic significance. Unfortunately, several language mistakes and other errors such as missing references were left in the text, many of which would have been easily avoided had the authors revised their work more carefully. Those that I could find are listed below:

Introduction:
Line 52: please change to: Radix auricularia (Linnaeus, 1758), a Pulmonata mollusk, is naturally…
Line 59: please change to: …have demonstrated that snail gut bacteria could assist…
Line 60: please change to: …vascular plants…
Line 67: please change to: …fix nitrogen…
Line 106: please change to: …adult snails were over 1 year old and juvenile snails were less than 1 year old.
Methods:
Line 120: Dennis et al (2013) is not on your references. Please correct this.
Line 123: Ma et al (2013) is not on your references list. Please correct this.
Line 128-129: please change to: spectrophotometry and paired-end sequences were generated on a Illumina Miseq platform (Illumina Corporation, San Diego, USA) with the 600-cycle MiSeq Reagent Kit v3 at the Shangai…
Line 132: The reference for Qiime that the authors provided (Lai et al. 2014) is not adequate. They should cite: QIIME allows analysis of high-throughput community sequencing data. J Gregory Caporaso, Justin Kuczynski, Jesse Stombaugh, Kyle Bittinger, Frederic D Bushman, Elizabeth K Costello, Noah Fierer, Antonio Gonzalez Pena, Julia K Goodrich, Jeffrey I Gordon, Gavin A Huttley, Scott T Kelley, Dan Knights, Jeremy E Koenig, Ruth E Ley, Catherine A Lozupone, Daniel McDonald, Brian D Muegge, Meg Pirrung, Jens Reeder, Joel R Sevinsky, Peter J Turnbaugh, William A Walters, Jeremy Widmann, Tanya Yatsunenko, Jesse Zaneveld and Rob Knight; Nature Methods, 2010; doi:10.1038/nmeth.f.303
Line 135: please change to: Using Usearch (version ….

Results:
Line 159: please change to: …between juvenile and adult populations.
Line 163 please change to: …are illustrated in Figure 2.
Line 184. This is an awkward sentence. I am not sure what it means. Please explain and rewrite it more clearly.
Line 197: please change to: …at both phylum and genus levels, there were no differences…
Line 219: please change to …KEGG pathways…
Line 225-226: Please change to: Figure S5 shows, according to KEGG mapping using the KO system, genetic pathways associated with xenobiotic biodegradation and metabolism.

Discussion:
Line 244: please remove the sentence: Furthermore…..similar. As this essentially says the same as the previous sentence.
Line 249: Please introduce a paragraph for: In this study.
Line 251: please remove the word taxa after phyla.
Line 256: Cardoso et al. 2012 did not study H. pomatia. They studied Achatina fulica.
Line 257: Please change to: Bacteroidetes
Line 282: Please change to: Pleurocapsa.
Line 286: Please change to: …lignocelluloses because of the presence of glycosyl hydrolases.
Line 287: Please change to: Many bacteria…
Line 294: Please change to: …most abundant OTU (OTU585) was affiliated with…
Line 309: Please change to: …account…
Line 312: Please change to: These bacteria….
Line 319: bacterial hosts is awkward.
Line 325: Please change to: …isolation of bacterial cellulases from snails….

Conclusion
Line 330: Please change to: …of individuals of the snail R. auricularia was described…
Line 332: Please change to: A high OTU and genus diversity were shown.
Line 333: Please change to: Growth and gonadal development may have influenced…

Suggestions to improve the Table and Figures:
Table and figures
Table 1- Please remove the text altogether and just leave the title of the table. You already mentioned in the manuscript text that there no statistically significant differences between the alpha diversity metrics and which text was used. I would also remove Sample/Estimators from the table as the headings are self-explanatory.
Figure 1- In the title please change to: …snail bacterial communities.
Figure 1- In the legend please change to: Four samples were collected from juvenile …
Figure 1- In the legend change to: sequencing of the V3-V4 region of the 16S rRNA gene.
Figure 2- In the legend change to: …taxa with abundance <0.01% are…
Figure 2- I suggest using only major taxa in the figure as there are too many taxonomic ids which makes the figure quite polluted. I would keep 1% for the phylum but increase to at least 5% for the family level and 10% for the genus level. You can give the taxonomy for every OTU and a full taxonomy table on supplementary files.
Figure 5- In the title please change to: PICRUSt predicts the functional composition of snail microbiomes.
Figure 5- In the legend please change to: represents instead of represent.

The references are sufficient but as outlined above some are missing in the reference list.
The authors supply a list of the representative sequences for some of the OTUs. However, they do not supply an accession number for their raw sequence data nor mention if they were submitted to a public database, This is not acceptable.
Experimental design: The work is within the aims and scope of PeerJ and the research questions are on the whole well defined, relevant and meaningful. As stated above there is still not enough data on snail gut microbiota and microbiomes even though these animals are a potential source of novel bacteria and enzymatic activities. Although the data are significant it would have been better if the authors had analyzed at least 10 animals from which group as groups with only 4 animals really make sound statistical analysis more difficult. But, as I said before, due to the scarcity of data on snail gut bacterial communities, especially when one considers the high biodiversity of this group, I still believe that data are useful as a preliminary report. I have some comments to improve the manuscript outlined below:
Methods:
Line 110-111: Please provide a drawing or photograph depicting the different parts of the snail gastrointestinal tract and indicate which parts were used for DNA extraction.
Line 126: The authors state that a library prep kit was used for making sequencing libraries but this strikes me as odd as amplicons under 500-600 bp are usually directly sequenced. The authors say that their amplicons had barcodes so I see no utility in using a library prep kit. The authors should clarify this.
Results
Line 186- What is this high abundance? More than what percentage?
Lines 191 -195- PcoA analysis- In contrast to what the authors state there are differences in community structure between juvenile and adult snail on both unweighted and weighted unifrac distances from what the graph shows. I suggest that a statistical analysis such as adonis, ANOSIM, BIO-ENV, Moran’s I, MRPP, PERMANOVA, PERMDISP, or db-RDA be used to on the distance matrices to confirm this observation.
Line 199-200: please indicate the abundance (%) of these genera in relation  to the total number of genera in juveniles and adults.
Line 216- PICRUSt does not estimate the number of sequences but estimates the number of proteins within pathways estimated from genomes from taxonomically related bacteria, so remove sequences. I suggest using genes instead of sequences. The same goes for line 220 where the authors say : sequences related to human…
Discussion
Line 244: please change to: …so diet was probably similar between sample groups.
As the authors did not show results for gut contents they cannot assume that the diet was highly similar. They can only suppose it was.
Validity of the findings: No comments as most of these points have been addressed in the previous sections.
Additional comments: All of the relevant comments are in sections 1 and 2.


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: COMPOSITIONAL AND PREDICTED FUNCTIONAL ANALYSIS OF THE GUT MICROBIOTA OF RADIX AURICULARIA (LINNAEUS) VIA HIGH-THROUGHPUT ILLUMINA SEQUENCING
Review round: 1
Reviewer: 3

Output:
Basic reporting: The overall structure of the manuscript is sound, and the figures and data reporting is acceptable. The authors should consider the following:

1. The internal structure of the manuscript makes it extremely difficult to follow. Each method, software program, experimental step, etc. needs to be clearly stated; the "how" and "why" are often confounded or absent. The authors must make their narrative more accessible to the reader by following a consistent mention of the methods both in terms of the order of presentation and in completeness. As a reviewer I have enough experience with what the authors are doing to mentally fill in the gaps as I read, but readers of the manuscript should not be expected to do so. A specific example: the authors used STAMP (the figures are easily recognizable) but there is no mention of it.

2. Many method and literature sources are explicitly or implicitly referenced throughout the document are not referenced and/or listed in the Literature Cited. The literature that is cited is not in alphabetical order.

3. The authors should have any future re-submissions checked by a native English speaker to improve basic grammar and punctuation.

4. I would recommend that the authors other relevant and/or recent published snail gut papers, including Joynson et al. (2017), Aronson et al. (2016), Van Horn et al. (2012) to name a few. These either represent the most recent publications or similar analyses from other freshwater snails.

5. Figure 2 needs to be reworked for easier interpretation, as does Figure 4.
Experimental design: The general experimental design is acceptable and consistent with other published works on snail gut microbe studies. My concerns with the experimental design are:

1. The authors crushed the snails to remove the tissue for analysis. This poses a few issues. First, Radix are large enough that when anesthetized they can be removed from the shell intact and the needed parts dissected. Second, although the shells may have been cleaned, snail shells harbor bacterial biofilms on the surfacce and in the fluid trapped in and around the body inside. By crushing the shell, the authors risked contaminating the gut flora with non-gut flora.

2. The sample size the authors used is very small, which is evident in their rarefaction curves.

3. In reporting the microbial results, the authors make no mention of separating the identified bacteria by possible origin. For example, cyanobacteria are most likely dietary and not enteric. The same holds for functional results; infectious and human diseases are identifiable KEGG pathways but are they biologically relevant to the analysis?

4. The 'standard' LEfSe cutoff for reporting is +/- 2 on the log LDA score. The authors used 3.5. Why?
Validity of the findings: I find the findings to be acceptable, the data properly generated and analyzed, and the conclusions to be supported by the results.
Additional comments: This is a sound study in theory, but isn't anywhere near ready for publication as it stands currently. The version sent in for review reads and looks like a very early draft of a manuscript that needs a lot of revision and improvement. I think there is value in the project's conception and execution, but the text belies this fact. I think with major revisions and extensive editing, the authors can have a quality publication on their hands, but not at this point.


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: COMPOSITIONAL AND PREDICTED FUNCTIONAL ANALYSIS OF THE GUT MICROBIOTA OF RADIX AURICULARIA (LINNAEUS) VIA HIGH-THROUGHPUT ILLUMINA SEQUENCING
Review round: 2
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: I am satisfied with the modifications undertaken by the authors and I think the paper is now suitable 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: COMPOSITIONAL AND PREDICTED FUNCTIONAL ANALYSIS OF THE GUT MICROBIOTA OF RADIX AURICULARIA (LINNAEUS) VIA HIGH-THROUGHPUT ILLUMINA SEQUENCING
Review round: 2
Reviewer: 2

Output:
Basic reporting: This study seeks to evaluate the microbiome composition and diversity of juvenile and adult Radix auricularia snails and to predict the effect that the gut bacteria may have on host functions. The study is very interesting, is generally well done, and helps fill an important gap in our knowledge. For being the second most specious phylum on earth, mollusks receive very little attention, and their microbiomes are relatively uncharacterized. This study provides important new data and insights in this regard.

There are still quite a few English language problems throughout the manuscript and I think that the manuscript would benefit from another round of editing from a native/proficient English speaker. There are also some instances where I feel that additional citations are necessary. The figures in general look great, there are a few edits that need to be made to some legends (detailed below), and the font in Figure 5 does not match the rest of the figures.
Experimental design: This article is in line with the aims and scope of PeerJ and I feel that this paper meaningfully addresses their questions. I think that this paper addresses an important knowledge gap and authors could provide a greater emphasis on this aspect of their work. In general, I feel that the methods used were rigorous and that results were evaluated accurately.

I only have a couple issues with experimental design. First, the authors need to provide further explanation as to why they chose to only use 4 samples from each developmental stage. Second, the unweighted and weighted analysis give different results. However, this discrepancy is never addressed or discussed in the text, and it needs to be. Otherwise, the methods are explained very well and in a way that would be conducive to replication of the study.
Validity of the findings: The data presented appear to be robust. After the authors address the issue of the weighted vs. unweighted analysis, I think that the results are statistically sound and interpreted in a reasonable manner. The authors generated novel data and interesting new conclusions that increase what we know about snail microbiomes, which thus far have been largely unstudied.
Additional comments: 1. Throughout the manuscript the authors use a lot of passive voice. I recommend revising a bit to include more active voice. This will increase readability. e.g. L167-169 could say: “We used principal coordinates analysis (PcoA) to calculate beta diversity, and subsequently used ANOSIM to confirm findings from the distance matrices.”

2. Make sure to use consistent fonts in all figures. Some fonts use serif fonts, others use sans serif fonts. Being consistent in figure formatting with increase aesthetic appeal of the paper.

3. The weighted analysis did not show this same pattern as the unweighted analysis, that analysis found no difference between JS and AS, suggesting no difference in microbiome composition based on developmental stage. Please acknowledge and explain this discrepancy

L21: type in title: should say “microbiota”

L46: the term “data information” is awkward, and not specific enough. Be more specific: e.g. “A large number of species could not be successfully identified, which could indicate the detection of novel ribotypes or result from insufficient availability of snail microbiome data.”

L48-50: this is an awkward sentence and I’m not totally sure what the authors are trying to say. predicted functions of what? What “genes” are the authors referring to if they only did16S sequencing of the microbiome, or are they referring to the snail?

L49: And do you mean “enriched” instead of “rich”?

L53: should say: “searching for xenobiotic…” please revise

L57: change “Pulmonata mollusk” to “pulmonate snail”

L62: harmful to

L66: The capacity

L66-69: the grammar of this sentence it awkward (particularly the term “aids their life”), please revise

L78: “previous”, not “precious”

L84: change “are even stronger impacted by” to “are more strongly influenced by”

L86-L89: or that reproductive mode influences microbiome composition, which way this relationship goes is still unclear

L98: “freshwater”

L102: add a sentence here briefly summarizing what you found and why it’s important.

L111: For grouping based on developmental stage? please specify

L112-115: Is there literature to cite to support these groupings?

L116-117: Please justify/explain why only 4 snails were used for each developmental stage. Of the 40 snails collected, did only 4 meet the criteria for each of the AS/JS groupings?

L121: change to “removed to expose”

L142: “products” not “productions”, and change “through a” to “using a”

L153: change “merged to a” to “merged into a”

L161: how was the single rarefaction chosen? At random?, change represent to generate

L162: change “sequences in all” to “sequence to be used in all”

L 185: The two instances of +/- have different formatting. Is one in bold?

L186: What is the standard deviation for sequence length?

L186-187: awkward sentence, change to something like: “the 214,584 rarefied sequences were clustered into 1,196 OTUs…”

L202-203: include the numbers here

L205-209: these sentences could be more concise, combined into a single sentence, and added to the previous paragraph

L210-211: change “at least one of samples” to “at least one of the samples”

L235: this sentence is unnecessarily complicated, change to something like: “Venn analyses found that 1060 OTUs (88.7% of the 1,196 OTUS identified) were shared between JS and AS.

L244: do the authors mean the predicted genomic functions of the bacteria in the snails’ guts/the snail functional pathways that the bacteria are predicted to influence? This sentence/figure 5 need clarification

L250: should say “enriched” not “rich”

L255: add the word “pathways” after metabolism

L255-257: this sentence could be part of the previous paragraph

L259: change “may be” to “maybe”

L265: do you mean present in high abundance?

L267: change would to could

L277: get rid of “In this study”

L278: “next generation” not “new generation”

L279: add the word “bacterial” before “community”

L281: add the word “community” before “structures”

L282: change to “indicating that the developmental stage may have an effect…”

L282-284: however, your weighted analysis did not show this same pattern, that analysis found no difference, suggesting no difference in microbiome composition based on developmental stage. Please acknowledge and explain this discrepancy

L285-288: this is an awkward sentence, please revise

L289: change different to differences and before to previously

L295: change bacteria to bacterial

L298: bacteria are NOT closely related to snails, I do not think this is what the authors mean to say here. The sentence needs to be revised and clarified

L307-310: please provide citations for this information

L236: change “not only” to “may not only”

L338: change indicated to implicated

L340-341: this sentence is poorly written, please revise to something like: “The predicted functions of infectious and human diseases that were identified among the KEGG pathways could potentially be associated with the genus Mycoplasma.”

L341-343: this could be moved to the section about Cyanobacteria. As they are both potentially dietary, it makes sense to discuss them together.

L347: change “several” to “other”

L348-350: Again, the weight analysis gave a different result. This discrepancy needs to be acknowledged and discussed.

L354-L356: please provide a citation for this sentence.

L357: change expose to understand

L359: Do you mean TAXA, rather than GENES? All of the results are based on the sequence of a single gene (16S). This is unclear and greatly needs to be addressed.

L363-372: Again, I think that you mean the bacterial taxa are associated with these functions, or that there is some evidence that the bacteria affect host genes involved in these processes. Please clarify what you mean by this relationship…because you only sequenced a single gene from a wide diversity of taxa.

L380: high number of OTUs

Figure 1: should go in the supplement

Figure 3: change “were showed”, to “are shown”

Figure 4 legend, change get rid of “taxon” in “bacterial taxon phyla” and “bacterial taxon core genera” and change “bacterial taxon and snail population” to “bacterial taxa and snail population”

Figure 5: change to a sans serif font so that it matches the other figures


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: COMPOSITIONAL AND PREDICTED FUNCTIONAL ANALYSIS OF THE GUT MICROBIOTA OF RADIX AURICULARIA (LINNAEUS) VIA HIGH-THROUGHPUT ILLUMINA SEQUENCING
Review round: 3
Reviewer: 1

Output:
Basic reporting: This new version of the manuscript is much improved. There are still many English language/grammatical errors detail below. Most importantly, the authors still need to address one of the previous reviewer comments regarding genes and their functional analysis. The authors only sequenced 16S and used taxonomic assignments to predict the functional effects of the bacteria themselves and the KEGG pathways that the bacteria are expected to influence. However, in the results section these data are still described as genes that are enriched in the bacteria, which is not what was done. The authors did a great job of addressing the weighted vs. unweighted analysis differences. After the one major issue mentioned above and the minor grammatical errors are addressed I believe that this manuscript will be ready for publication.
Experimental design: None
Validity of the findings: None
Additional comments: L34: change “The gut microbiota is involved” to “The gut microbiota are involved”
L78: change “and previous study showed” to “and previous studies have shown”
L85: change “Gut microbiota is” to “Gut microbiota are,” remove “the” before reproductive, and change “process” to “processes”
L101: change “The results of study here may fill” to “The results of this study fill”
L102: change “of technological” to “for technological”
L113: change “preliminary” to “preliminarily”
L115: change “preliminary” to “preliminarily”
L116: change to “juvenile snails”
L117: change to “adult snails”
L119: change to “Snails were”
L123: change to  “washed with sterile water”
L110-L128: In this section the authors need to say how many snails they ended up with after sampling and classifying (4 from each group).
LL160-162: This sentence does not belong in this section; it should be in the “DNA extraction and PCR amplification section.”
L171: change “co-ordinate” to “coordinate”
L173: change “To inspect the statistically significant differences of taxonomic groups between JS and AS” to “To identify statistically significant taxonomic groups that differ between JS and AS”
L217: change “identified” to “identifiable”
L230: change “level” to “levels”
237: specify whether “these OTUs” refers to the 469 core OTUs, or to the 70 JS and 65 AS unique OTUs.
L243: change “pathway” to “pathways”
L248: change “enrich” to “enriched”
L242-264: As noted in the previous round of reviews, “genes” is not the appropriate term here, and is actually misleading. The authors only sequenced 16S from the gut bacteria. The authors should be referring to predicted functions of the bacteria themselves and the PATHWAYS that they might influence, not specific genes. This needs to be changed throughout this entire section, as it is not an accurate representation of what was actually done in this study. The title of this section (16S rRNA gene functional prediction) also needs to be changed, as it is also not an accurate portrayal of what was actually done here. A more accurate section heading would be something like “Functional predictions of bacterial communities.”
L280-281: “partly due…are present”,  the grammar in this part of the sentence is awkward. Change to “this is at least in part due to abundance information, which can obscure significant patterns of variation in the taxa that are present.”
L285: change to “At the phylum level”
L286: change “ as the dominant”
L299: get rid of phrase “bacteria related to”
L325: change to “of the cellulolytic”
L340: change “was affiliated” to “were affiliated”
Fig. 4: change “PICRUSt predict the” to “PICRUSt predictions of the.” In the legend change “represent” to “represents” in all occurrences.


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

Input:
Title: A SIMPLE METHOD FOR DATA PARTITIONING BASED ON RELATIVE EVOLUTIONARY RATES
Review round: 1
Reviewer: 1

Output:
Basic reporting: see below
Experimental design: see below
Validity of the findings: see below
Additional comments: Rota and colleagues present a generally well written manuscript that formalizes, tests, and makes publicly available a novel method that they have used in some recently published papers.  I think the manuscript merits publication in PeerJ, but only after the authors have rigorously addressed my six sets of comments below.

-- Mark P. Simmons, 16 November 2017

Major comments:

I’m glad to see that the authors appropriately cite John Gatesy’s and my 2016 paper criticizing TIGER (and OV).  But so far as I can tell they have not addressed the implications of our study on their favored method of using TIGER to partition characters by rate.  Even when all characters are sampled, isn’t the approach still biased in favoring assignment of informative characters with highly asymmetrical partitions to the slowest rates?  The authors can test for this using Farris’ (1989) amount of possible synapomorphy.  Also, do the trees generated using their partition-delimitation method have more cherries than the trees using the other four partitioning methods they applied?  See Simmons and Gatesy (2016) for additional details.  I think the authors need to address this bias before recommending their methodology for general use.

“However, the k-means algorithm has been found to place all invariable characters into one partition (Baca et al., 2017), which leads to biased likelihood values."  The point is to partition characters by rate, and the simplest explanation for invariable characters is that they have not evolved at all, and hence should be assigned to the lowest rate partition.  State explicitly why you believe this approach is problematic.  The authors cite biased likelihood values, but why does this approach lead to biased likelihood values and why does the approach fail based on theory rather than in an operational context?

The main results of the paper are presented in Table 1.  First, why wasn't TIGER, without the authors' method tested?  Second, given that these models invoke different numbers of partitions and hence parameters, how did they control for this effect on their Y-axis?  So far as I can tell there is no control and so it's perfectly reasonable to expect more partitions to lead to greater likelihoods.

Minor comments:

“We do not carry out phylogenetic analyses and compare
resulting topologies because it has been previously established that partitioning does affect
topology, branch support, and branch lengths (see Kainer & Lanfear, 2015 and references
therein), and since true phylogenies in all of these cases are unknown, we can only select the best
partitioning strategy using statistical model evaluation metrics, such as e.g. BIC."  Break that up into two separate sentences.

Where is the description of how tree searches were performed?  I don't see this information in the methods.

“BIC was chosen as a statistical model evaluation metric because it has been shown to perform well in model selection for phylogenetic analysis (Abdo et al., 2005)."  The same can certainly be said for the AIC and the LRT.  State explicitly why you chose to use the BIC rather than these alternatives.


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

Input:
Title: A SIMPLE METHOD FOR DATA PARTITIONING BASED ON RELATIVE EVOLUTIONARY RATES
Review round: 1
Reviewer: 2

Output:
Basic reporting: Two minor comments:

1. The figures need axis labels at all axes.

2. l. 104-106 "One of these properties is that with standard DNA sequence data of protein-coding genes, one to two thirds of the data consist of invariable characters." This seems to depend on the specific data set analyzed. Maybe write "typically consist of".
Experimental design: One comment:

An improved BIC score does not in any way guarantee that the inference result is closer to the truth. This is particularly the case when the model is misspecified, which is essentially always the case when working with real (as opposed to simulated) data.

I think that, at a minimum, the authors should also perform a test on simulated data and show how their method affects actual quantities of interest, such as the distance between the inferred and the true tree.
Validity of the findings: Two comment:

1. As mentioned under "Experimental design", improvement in BIC does not guarantee that the inferred result is actually closer to the truth. This needs to be discussed, and the method needs to be tested with some simulations where the ground truth is known.

2. I would like to see the authors think more carefully about why different sites evolve at different rates. It's not clear to me that all sites that evolve at the same rate can be thrown together into one group. For example, second codon positions may evolve rapidly if the corresponding amino acid lies on the surface of the folded protein [1], and third codon positions may evolve slowly due to various causes of codon bias [2]. Is there any good reason to believe that a fast-evolving second position and a slow-evolving third position should be grouped together?

[1] J. Echave, S. J. Spielman, C. O. Wilke (2016). Causes of evolutionary rate variation among protein sites. Nature Rev. Genet. 17:109–121

[2] J. B. Plotkin, G. Kudla (2011). Synonymous but not the same: the causes and consequences of codon bias. Nature Rev. Genet. 12:32–42
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: A SIMPLE METHOD FOR DATA PARTITIONING BASED ON RELATIVE EVOLUTIONARY RATES
Review round: 1
Reviewer: 3

Output:
Basic reporting: Fine
Experimental design: Needs simulations and code tests, see below.
Validity of the findings: Depends on simulations and comparisons with related methods (free-rates models). See below.
Additional comments: This paper presents an approach to automatically partition datasets based on TIGER rates.

I am intrigued by the approach, but I have a couple of concerns which mean that I remain to be convinced that the method is useful.

First, I think it’s vital that we see some simulations here. I say this because we have tried a range of very similar approaches to this, and in every case we have found insurmountable problems with the approach on suitably simulated data. The authors correctly point out that our previous attempt at a very similar approach (the k-means method) had some unforeseen properties. In developing that method we tried it on a range of real and simulated datasets, and never found any odd behaviour. As the authors (and we) have noted, this was premature: our method turns out to do some odd things.

What the authors couldn’t have known is that in light of the problems identified with our method, we tried a range of approaches that are very similar to that presented here. These included (but weren’t limited to) lumping together invariant sites and slow sites; removing invariant sites from the analysis and using a bias correction when inferring trees; and lumping every invariant site with the physically closest variable site. In all cases, the methods seemed to perform just fine on most datasets. But in all cases we found that they had inexplicable issues in some corner cases. Because we were unable to resolve (or properly understand) these issues, we have yet to publish these results.

The relevance of this for the current ms is simple: we need to see a good range of simulations in which the authors simulate appropriate datasets (more on that below), and test whether using their partitioning method one can recover the correct tree and branch lengths. Given the history of these methods, it’s absolutely crucial that this is done before the method is published and promoted. To be clear – I absolutely hope that this method works great. It is certainly a little different from what we proposed in print, or have tried since. But to be frank, I suspect that it will suffer the same limitations. And if it does, I don’t think it should be published or promoted for use unless those limitations can be clearly understood, and crucially predicted accurately for empirical datasets. It’s the latter limitation which led us to remove k-means from PartitionFinder. All of our evidence suggested that the odd behaviour was very very rare. But unless we can predict which empirical datasets will suffer from the odd behaviour, we cannot (and the authors should not, if simulations show their method to also be inadequate) suggest that this is a useful method.

The simplest simulation conditions for which I have been able to show that the k-means algorithm (and our various modifications of it, which are similar to the work presented here) suffered from odd innacuracies are as follows:

1.	Simulate a phylogram with rate variation, with 4-20 taxa
2.	Simulate a 4kb alignment under a JC+G model  on that phylogram
3.	Reconstruct the tree (using e.g. IQtree) with the true model (i.e. JC+G)
4.	Perform the TIGER partitioning described here
5.	Reconstruct the tree under the TIGER partitioning scheme

If one repeats this ~10 times for each set of simulation conditions (e.g. varying the number of taxa, the total substitution rate on the tree, and the alpha parameter of G), one has a baseline set of simulations where rate variation varies along (because of variation in G). Of course, one will also need to vary the parameter for rate splitting presented in the current method.

Reconstructing the tree with the true model tells one how accurate one can hope to be (on average across the 10 simulations). And one can then ask whether the TIGER rate partitioning is as accurate as analysing the tree with the true model, or less accurate. Of course, we would expect it to be a little less accurate. But what we observed with the k-means approach (especially on datasets with very few taxa) was that the reconstructed k-means trees often completely collapsed the internal branch.

I have already written all of the code for these simulations (except the TIGER parts, which would be simple to add) and would be happy to share this with the authors if that’s of use. I really, genuinely, hope that their method overcomes the limitation of the method we proposed. But in the circumstances I think it’s very important to check with some rigorous simulations. Crucially (and where we failed in our ms) these simulations have to include tests of the reconstruction of the tree itself.

Of note is that these simulations are extremely fast to do, even without access to HPC resources it would be possible to do 10s of thousands of simulations.

A few other comments:

Line 84: I don’t know of any evidence that the likelihood values are biased. The likelihood values make perfect sense – an invariant site can be perfectly explained with a zero-length tree, so gets a likelihood of 1. I think the issue comes in model-misspecification on the remaining subsets (those without invariant sites included). This is likely to be an issue for the current method too, because it removes invariant sites from all of the faster-evolving subsets. All the more reason for a rigorous set of simulations.

Lines 96-99. For all the reasons above, I think it is critically important to compare phylogenies. Both of the simulated data (see above) and the empirical datasets.

Lines 108-109: invariant partitions do contain information: on the distribution of rates across sites and on the base composition. These inform the model, which informs the tree.

Line 109: What is the evidence that it’s advisable to include some slowly-evolving sites in a partition of invariant sites?

Line 143-145: This sentence is unclear. In addition, no justification is provided for removing sites. Also, my (perhaps incorrect) understanding was the TIGER did not cope with missing data. Is this wrong? Can the authors explain how a TIGER rate is calculated when data is missing here.

General point: I think one should really compare this approach to the newly-implemented free-rate models in IQtree. These are mixture models which nominally do exactly what the authors are proposing, but without the headache of running TIGER (which is exceptionally slow for larger datasets) and the author’s script. This could easily be incorporated in the simulation pipeline, and also in the analysis of empirical datasets, simply by adding an analysis in which one ran IQtree with automated model selection. This is important, because it may be that the authors method outperforms (or is outperformed by) these methods. The advantage of the free-rate models is that they have a rigorous statistical underpinning, whereas the current method (and my own work on k-means) is much more of a hack to a solution. I have no problem with the hacked version, but the target audience for this method (empirical phylo folks) should be given a straightforward comparison of methods.

The Python script seems to work fine (I tried it). However, I have a few concerns given that the paper is proposing that this is a script that others should use. First, the script should ideally be placed in a repository (e.g. GitHub) where people can find it. Second, and more importantly, we need to know how the script was tested, and we need to see the tests. There should be a series of unit tests, and some end-to-end tests, which demonstrate that the script works as intended.

Yours,

Rob Lanfear


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

Input:
Title: A SIMPLE METHOD FOR DATA PARTITIONING BASED ON RELATIVE EVOLUTIONARY RATES
Review round: 2
Reviewer: 1

Output:
Basic reporting: N/A
Experimental design: N/A
Validity of the findings: I don't feel the authors have adequately addressed my comments. I had two comments under this topic, one of which was "I would like to see the authors think more carefully about why different sites evolve at different rates" (reiterated by the handling editor). The authors essentially dismissed this point and did not make any relevant changes that I could see. I think this is insufficient.

I understand that this is a complex topic and there's only so much one can do in a paper, but at the same time I think it is critical to not pretend that actual genetic sequences in actual organisms are just random strings generated from some uniform stochastic process (see also: http://journals.plos.org/ploscompbiol/article?id=10.1371/journal.pcbi.1002572). There is real biophysics underlying the evolutionary divergence of genetic sequences, and at a minimum the authors need to think under what conditions their work may or may not violate known facts about biophysical constraints on molecular evolution. Pretending these issues can safely be ignored is insufficient.

What I would like to see is about a paragraph of a careful discussion of these issues.
Additional comments: N/A


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

Input:
Title: ANATOMICAL NOTES AND DISCUSSION OF THE FIRST DESCRIBED AETOSAUR STAGONOLEPIS ROBERTSONI (ARCHOSAURIA: SUCHIA) FROM THE UPPER TRIASSIC OF EUROPE, AND THE USE OF PLESIOMORPHIES IN AETOSAUR BIOCHRONOLOGY
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript by Parker & Martz is a solid description and discussion of some previously overlooked anatomical features of Stagonolepis, a classic aetosaur taxon from the Late Triassic of Scotland, with a discussion of the taxonomy and phylogenetic position of this taxon. There are no substantial problems with this work - it is generally well written and adequately referenced and appropriately organised. I have a number of minor comments on typographical errors and grammar that are marked directly on the attached PDF.
Experimental design: This paper contains original research, and there are no problems with research design.
Validity of the findings: Results and conclusions are well supported by the presented data.
Additional comments: I don't have any substantial edits to suggest. I have made a number of minor comments on typographical errors and grammar on the attached PDF. There are also some minor comments on terminology and the correct names and abbreviations of repositories for the described fossil specimens.


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: ANATOMICAL NOTES AND DISCUSSION OF THE FIRST DESCRIBED AETOSAUR STAGONOLEPIS ROBERTSONI (ARCHOSAURIA: SUCHIA) FROM THE UPPER TRIASSIC OF EUROPE, AND THE USE OF PLESIOMORPHIES IN AETOSAUR BIOCHRONOLOGY
Review round: 1
Reviewer: 2

Output:
Basic reporting: The paper provides an update on the earlier classic monograph by Walker (1961), and confirms that that older work was largely accurate. It might then seem unnecessary to publish a further paper when there is no new material. However, since 1961, many new aetosaurian taxa have been found and, in particular, these have been subject to several cladistics analyses – the current authors have been world leaders in this endeavour, and so their comments on the classic material are enlightening. The restudy has allowed the authors to confirm the correct coding of phylogenetically informative characters in Stagonolepis robertsoni, and provide a diagnosis. They further resolve issues including mis-assignment of some American material to Stagonolepis, a mis-assignment that had been used as a biostratigraphic indicator, but which now fails. Detailed comparisons are given between S. robertsoni and other aetosaurs, most of which were not known in 1961.

In particular, the osteoderms are described more fully, in terms not considered important in 1961, but now critical in determining the phylogenetic position of the taxon.
Experimental design: Anatomical description, identification of phylogenetically informative characters, and phylogenetic analysis all properly carried out.
Validity of the findings: Findings are all well justified.
Additional comments: 37	the despite = despite
77	is the = is to
81	instead = but instead
90	are in = is in
106	NHMUK R4787; Sandstone = NHMUK R4787, sandstone
122–128 insert taxonomic commas, after author names
308	36mm = 36 mm
407	S. robertsoni = S. robertsoni
421	uppermost which = uppermost of which
435	Stagonolepididae as = Stagonolepididae, as
485	apomorphy based = apomorphy-based
513	drop shaped = drop-shaped
533	status = status,
587	poisons = poisons
587	grés = grès
587	Dévonian = Dévonien
587	ed de = et de [if you don’t know French, it’s best simply to copy and paste from a reputable source: https://scholar.google.co.uk/scholar?hl=en&as_sdt=0%2C5&q=Monographie+des+poisons+fossiles+du+vieux+grés+rouge+ou+Système+Dévonian+%28Old+Red+Sandstone%29+des+Isles+Britanniques+ed+de+Russie.&btnG=.]
753	Triassic Reptiles from the Elgin Area = Triassic reptiles from the Elgin area