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: EFFECT OF DIETARY CONCENTRATE TO FORAGE RATIO ON GROWTH PERFORMANCE, RUMEN FERMENTATION AND BACTERIAL DIVERSITY OF TIBETAN SHEEP UNDER BARN FEEDING ON THE QINGHAI-TIBETAN PLATEAU
Review round: 1
Reviewer: 3

Output:
Basic reporting: There were a lot of typos and wording mistakes along the whole manuscript found, for example:
line 158 ("Wiener" not "Weiner"), line 216, 370, Figure 3: NOT "Winner";
line 6: high-throughput [] was applied
line 19 Tibetan sheep
31 delete "a developed strategy" or rewrite it
34 certain instead of some ; rarely instead of seldom, use instead of with
35 this is not correct: concentrate feed would be even higher in certain nutrients as grassland, so rewrite the sentence in terms of: "could be disadvantageous in terms of rumen health, rumen acidosis etc."
35-37: if you DID already experiments, please give reference; if this is a grammar mistake, you need to write "in this study .. we performed.."
38 Start the sentence with "Rumen microbes [...]
39-40: reassess sentence: the microbes digest the plant materials and provide nutrient to the host, they probably perform their digestive metabolism without the host itself, the host only provides a adequate habitat for the microbes
41 not only CP, ADF and NDF: all nutrients that are ingested by the host! here you can be more general in writing protein, complex carbohydrates, starch, sugars, fat..
43 delete "crude", its just bacterial protein;
49 However, only few
76 change "non antibiotics" to "not under current antibiotic treatment"
79 are shown in Table 1
81 forage; from the same local herdsman
88 grind throug a 1mm sieve
92-93 electric pump, which was confirmed to show similar results for rumen microorganisms and fermentation parameters as samplig via a rumen cannula

line 158 indexes and species
line 160 weighted Unifrac
line 180 Body weight, grain ratio and ADG (--> Abrreviations need to spelled out at the beginning of the sentence
line 187 Rarefaction curves
throughout the manuscript: use for P either lowercase or uppercase letters P / p, don't switch
194 the highest relative abundance
213 [..] phyla. The relative [...]
217 -218 rarrange sentence: Both parameters were significantly higher in [..]
243 fermentation parameters pH, A:P and the CP of the diet
251 what do you mean by "positively jointly influenced" - rewrite this sentence
253 I would rather use "correlated" instead of "influenced" because we don't know for sure which parameter was the driving one.
259 treatment
266 and diets
283-285 Rearrange sentence: From a energy-utilization point of view, [...] means This can also explain that the high-concentrate diet
287 treatment (singular!)
296 In the current study
305 what do you mean by older bovines? old in terms of age, old in terms of DIM? please be more concise!
323 gourp (singular!)
330 according to your results, Prevotella_1 and CP were both increasing, so it ist not an "undulation", please rechekc and better use "increase"
332-336 Pseudobutyrivibrio genus / spp., Prevotellaceae! Correct mistaken genera names. Why do you cite Yange et al 2010 at this point in the sentence, they found Pseudobutyrivibrio in their study, but did not isolate and describe it; you should use citations to explain your findings. Rearrange this or delete the citation.
344 don't use "altered" in a manuscript, be concise with "increase" or "decrease"!
356-361 This abstract is very vague and general, please rewrite and be more concise in relation to your findings
365-366 [...] was significantly higher in groups HS3, HS4, and HS5 than in HS2 and HS2 (P<0.05).
375-376 Rearrange the sentence [Therefore, ...] to for example:  From our findings we can conclude that a better growth performance is not linked to a higher diversity of rumen microorganisms.
377-379: you cannot state that the microorganisms kept growing, you did not do any cultivation or counting; also correct for: when the C:F ratio ranged from 0:100 to 30:70, [..] With a C:F ratio of 45:55 and 60:40 the growth performance could be promoted.- suggesting that ...
lines 373, 388,395 - no figure and table references needed in the discussion section
387 there seems the beginning of the sentence missing: Therefore the inclusion of x% concentrate in the feed might be a good choice for [..], delete "than any of the other groups"
390 why do you only discuss cellulose digestive capacity? when the bacterial community is changing, all digestive processes are affected in the rumen.
390-391 delete first "growth performance", adapt sentence
395-398 rearrange sentence, as for example: Therefore, in our study the increasing abundance of B. and Pr. in the rumen of Tibetan sheep could possibly contribute to a higher nutrient availability for the host and consequently higher growht performance.
400 gain rate
401 you mention the cold season, please give more informations about the nutrient availablitiy during the warm season, would your findings also apply for the warm season?
403-404 that there is no better growth
407-408: Please name specific persons, institutes, teams and what they contributed!
236-237 [..] ruminal samples, being significantly lower in HS1 compared to the other HS groups.

A proper proof read by all authors is requested in order to correct for such kinds of writing mistakes! Moreover I suggest the proofreading of the manuscript by a native English speaker.

Please spell out all non-SI abbreviations at first usage in the main manuscript (line 5 HS, etc.); use the same nomeclature for the same meanings: as for "high concentrate" always use "high concentrate" etc.; this improves clarity about what you are talking about;

The introuction about the knowledge of rumen microbes and the influence of increasing amounts of concentrate feed is too general for this manuscript. It is well known that a change in diet changes the rumen microbes.  55-60: It is well known that NGS is used in studying rumen microbes, you don't need to state that. be more precice: as for example in line 44-46: do not write "useful products", which products? Propionate?. Focus on your special questions on Tibetan sheep. The gap of knowledge of rumen microbes in this sheep is only little supported by the background information given. The reviewer would like to see more informations about the specific region the sheep live in, their exact purpose, their normal growth performance, the health or management problems that occur with the grassland feeding, what you mean by scientific breeding (line 62), their special physiology, the cost effectiveness of a concentrate diet for the farmers to underline the need for reasearch in this field. Also differences of this sheep to other ruminants such as cows could underline the gap of knowledge. More references concerning studies with changes in diet in sheep are required (as for ex. Lettat et al, 2010 J. Anim. Sci.; Minuti et al 2014, Journal of Animal Science; M.M. Zein-Eldin et al, 2014, Biotechnology in Animal Husbandry; Sonia Andrés et al 2018,Small Ruminant Research, etc..)
The Tibetan sheep have been used in some studies about methane emmission, it would be worth looking in those studies for rumen fermentation parameters to discuss: Wang M et al., Front Microbiol 2016;7:850; Zhang et al. (2016) Curr. Biol. 26:1873–1879;

A clear Hypothesis is missing. Please state it!

Figures are tables: Altough the requirement for legends for figures and tables is optional in the Journal, at least the the feeding groups (HS) should shortly be described also in the figures so that they can be understand as standalone;
Table 4: The calculation of Spearman correlation needs also be mentioned in the statistical part in M&M of the manuscript. Rearragen the table, so that there are no blanc boxes;

Supplementary data: Table S2: change "nutrients" to "concentrate", Table S3: change "functional" to "selected" ; change "item" to "species",
Experimental design: There are many unclear analyses and statements along the manuscripts:

76-79 Did you perform a statistical power analysis to prove n=4 to be sufficient for this kind of research?

80 Table 1: give the exact ingredients of the concentrate feed; Please analyze the starch content of the fed diets. These parameters are important, since the most active nutrient according to concentrate feed is varying starch levels due to varying concentrate feedstuff. In M&M line 81 only "oat hay" is given as forage feed, in Table 1 also "Green hay" is stated. Please be clear along the manuscript what was exactly fed! Please define "green hay"
Also a separate analysis of the concentrate and the forage feed as supplemental material would improve the quality on a nutritional state of the manuscript, because oat hay also contains noteworth amounts of non-structural carbohydrates;

82 were the animals also housed separately? or only fed separately?
How much feed was offered? how was the feed mixed / processed? Who performed the feeding and the sampling?
74 and 84 which diet was fed during the 15 days adaptation? The same baselinediet for all groups, or the HS1-HS5 diets for each specific group?
90 for future studies I recommend to do weighting more often if you want to describe Average daily gain. because with n=4 you might have high variations, also state timepoint of weighting (before, after feeding?)
91 when exactly were the samples taken, before feeding, after feeding, which timepoint?
144-146 give more details about your quality filtering: which treshold did you use?
158 state all alpha diversity parameters you analysed. Also analyse for Goods_coverage, in order to prove sampling and sequence depth, which should be above 90% for each sample;
164 For stronger explanatory power of correlation I would recommend to do spearman correlation also for the microbiota and fermentation parameters instead of RDA plots.
165 why did you only perform analysis of the top 32 known genera?
167 State your spearman correlation analysis in the statistic section

182. what is the difference between DMI and DMI consume?
188 please give the minimum sequence number you rarefied your rarefaction curves together with the %
192 41 phyla (and also 129 genera as in line 200) seems like a very high number, did you remove singletons from your sequence downstream analysis? Also give your phyla, genus and species talbes as supplementary material
195 change "less" to "moderate"
194 Please give information how you calculated realtive abundance: if the highest 10 phyla made 98% of the sequences, how can then for example Lentisphaere still have an abundance of 5.4%??
201: ist it really 0.3%? should that not be more higheer if they are the most important bacteria?
203 This demonstrates variabilit [..] -- belongs to discussin section
Table 3, Table 4, M&M section: In the material and methods section (line 163) the calculation of the feed efficiency (calculated by output/input) is claimed, however the results given in Table 3 as feed (conversion) efficiency (FCE) seem to be results of feed conversion ratio (FCR) calculation (calculated as output/input)!. Please clarify in the M&M section which of these two parameters was calculated and how, and correct your results data according to that. Also review the values used for calculation of the spearman correlation between FCE/ FCR(?) and the other parameters, since a decreasing FCR should result in a negative spearman correalation value when analysed with the increasing DMI, increasing BW, and ADG (and not positive, as given in table 4) and vice versa with Shannon, Observed species and Chao1!
214 do you have a certain reference for Prevotella_1 or was this just a blast result? Beucase in the discussion (329 and others) you discuss this Prevotella_1 very generously. I do not agree that there is so much knowledge about what this Blast result can degrade or not in the rumen.
232: state in which diets Prevotella_1 exactly increases!
232 and others: give the reason why you have chosen for example the top 12 genera? what were your criterions?
339 fibrolytic
Validity of the findings: 219: Be careful when interpreting your PcoA plot: the statement that all five groups were clearly distinguished is not supported by your plot: grouop HS1 and 2 cluster very closely together, so do HS4 and 5! and HS3 is ways more close to HS1 and 2 because PC2 represents only 7.14% of the variabilty. Reinterpret this section! also the numbers given in the text (line 222) do not fit to your figure!
316-318 Wang et al. 2016 did not describe any Acidobacteria, nor that they are inhibited in growth by proteins. Referring to general knowledge, dietary protein does not inhibit rumen bacteria.
366-368 You state that S.ruminatium was higher in the high-concentrate groups because they can withstand low ruminal pH and acid accumulation, but in your findings the pH and total VFA were not changing with concentrate feed. Please reconsider this discussion point!
393-395 You give the point that "although richness and diverstiy decrease" in your study, "Bacteroidetes increased" - this is not "although", it is well known that when diverstiy is decreasing, that certain bacterial groups can evolve, and so start to dominate the community, a decrease in diversity mostly goes hand in hand with a higher dominance index, and higher abundance of a certain group of bacteria.
270 Why is the protein level playing an important role in pH changes in the rumen? Starch would be the main driver for pH changes in the rumen. Please review this part and do proper literature research!
274-277 why is the rumen of Tibetan sheep special in terms of maintaining a stable pH? Please give better explanations for your findings! To my knowledge the general forestomach system is similar in all sheep.
287 "acute" is not appropiate for your research quetion: acute changes, including acute pH changes would clearly be seen in your samples. There is no acute change expected with a concentrate: forage ratio of maximum 60:40
290-292 a difference of 390 OTUs is not a noteworth difference between sequencing studies. remove or adapt this discussion point. also the reasons are very general. Do focus on the groups of bacteria found!
319 and following: Again: do not overinterpret protein related changes for prevotella. Better use correlation analysis for this data; prevotella can degrade everything than lignin and cellulose, they are not solely dependent on protein.
338- 343 the discussion about the ruminoccus species is not related to your research question. That R.albus and R.flavefaciens do compete in vitro is well known, but you need to discuss your findings in terms of an in vivo rumen study and to sheep production level
Figure 1: there seems to be a clustering of certain groups of sheep and the feeding groups, please state this in your results;
Additional comments: The authors investigated the effect of increasing replacement of the traditionally fed grass diet with concentrate based feed in Tibetan sheep. Analyses in respect to performance of the animals (DMI, BWG), rumen fermentation parameters (VFA, NH3-N) and rumen microbiome (16S rRNA gene sequencing) were performed. The trial setup is in the reviewers opinion sufficient to answer first questions of this research approach, analyses performed are well established methods in that field and the manuscript is well structured. However, there is a massive weakness in how the manuscript was written and results were interpreted. All the background information, description of the M&M and discussion are written too general, not focused on the research question, do have many unclear passages, writing typos and unclear, incoherent and weak descriptions of methods, results and discussion. I insist on revising the whole manuscript especially according to what is described above.


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

Input:
Title: EFFECT OF DIETARY CONCENTRATE TO FORAGE RATIO ON GROWTH PERFORMANCE, RUMEN FERMENTATION AND BACTERIAL DIVERSITY OF TIBETAN SHEEP UNDER BARN FEEDING ON THE QINGHAI-TIBETAN PLATEAU
Review round: 2
Reviewer: 1

Output:
Basic reporting: The revision is adequate. However, the English language still needs improvement. Numerous grammar errors existed in the revised manuscript.
Experimental design: No comment.
Validity of the findings: no comment.
Additional comments: Comments
The revision is acceptable.

The English language still need improvement. Below are some examples (though not the exhaustive list):
In Abstract:
1.	Line 30, “The results showed that increases dietary”, please consider to change “increases” to “increasing”
2.	Line 32, “The increases dietary concentrate feed level”, please consider to change “increases” to “elevated” (or something similar)
3.	Line 37, “In conclusion, increases dietary concentrate”, please consider to change “increases” to “increased”

In introduction:
1.	Line 60, “Seriously, irrational grazing during this period”. Please consider to remove “Seriously”
2.	Line 72, “can be affected by many factors such as diet”, please consider to revise the sentence into “can be affected by many factors, such as diet”

In Materials and Methods:
1.	Line 186, please use appropriate sign for “≧97%”.

In Results:
1.	Line 226, “The top ten phyla, which exhibited the highest relative abundances”, please consider to revised it to “The top ten phyla, which exhibited the highest relative abundances, were prevalent in all the samples, accounting for nearly 98% of all sequences.”
2.	Line 231-232, please rephrase this sentence, “10 genera with relative abundances that represented more than 1% of bacterial community were perceived”

In Discussion:
1.	Line 315, “This result might due to the”, please consider to change it to “This result might be due to the”
2.	Line 319, “NH3-N concentration, which consisted with former study”, please consider to change it to “NH3-N concentration, which is consistent with former study”


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

Input:
Title: THE ROLE OF PAST EXPERIENCE IN DEVELOPMENT OF FEEDING BEHAVIOR IN COMMON VAMPIRE BATS
Review round: 1
Reviewer: 1

Output:
Basic reporting: No comment
Experimental design: No comment
Validity of the findings: No comment
Additional comments: This manuscript is very clearly written. The methods used are adequate to the question being asked, and the results are very clear. The discussion is grounded in the results obtained, with little to no speculation. Thus, I really have no major comments. I have a couple of suggestions and comments on some very minor issues.

The first suggestion is to move the idea mentioned in lines 94-98 to the discussion. I think you have a very solid foundation for establishing predictions about learning in vampire bats without the ideas included here; in fact, these do not seem well supported, but I believe would be more valuable in your discussion.

My second minor suggestion is to remind the reader what each experiment is trying to accomplish in materials and methods. For example, either include some hints in the subtitle "Experiment x" that reminds us what you are trying to accomplish, or else start the first paragraph with something like "To asses if captive-born bats show a preference for warm blood, we presented an unfed...".

Third comment: line 257 should be mentioning table 2, not table 1.

In line 304 you say that porcupines and armadillos are exotic, which they are not. Do you mean here that they are rare, or perhaps not the vampire's regular prey?

In line 318, I would remove the word "null", and just leave it as "We had two unexpected results".

I have a minor confusion about your test subjects for experiment 2. You mention in the section "Animal subjects" (material and methods) that you only tested animals aged 6 to 16 months of age. However, it is clear that is experiment 2 you use some adult females (which you mention in the methods section for experiment 2, no doubt). So perhaps to avoid confusion, in the section animal subjects you can clarify this to avoid confusion later on?


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

Input:
Title: THE ROLE OF PAST EXPERIENCE IN DEVELOPMENT OF FEEDING BEHAVIOR IN COMMON VAMPIRE BATS
Review round: 1
Reviewer: 2

Output:
Basic reporting: The human evolution framing needs work. Please see my general comments.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: Review by Teague O'Mara

Quick summary:
The paper is nicely written and easy to read, the experiments seem to test what the authors set out to test, data and analysis are solid, but the framing of study is a big reach and its strategy should be rethought (although it is never really discussed later, for what it's worth). Even though the results are a bit underwhelming, it's great to see more developmental work on bats and I think this work adds excellent information regarding the peculiarities of vampire bats.

General Comments:
The way that the authors frame the study in the context of human learning over simplifies a large and complex literature on the interactions among life history, brain size, social complexity and the ratchet effects of cumulative culture, and food complexity. The study by Schuppli and colleagues is their citation for all of human life history evolution (and associated learning), but this only touches on many of these traits. Schuppli, Isler, & van Schaik target why humans have a late age of skill competency and tests this via several theoretical constructs, often referred to as 'needing to learn' or 'energetic constraints' related to brain sizes and ecological risks posed to juveniles. Schuppli really targets the length of the immature period / juvenility and subadulthood (measured by age at first reproduction, AFR, versus the age of adult skill acquisition, ASC). While most mammals have relatively early ASC relative to AFR, mammals with complex foraging niches have ASC more similar to AFR, and humans have ASC much later than AFR. Schuppli's reasoning was that the synergistic effects of complex social relationships, allomaternal care, post-weaning provisioning, and the cooperative pooling of food all come together to allow humans to reproduce much earlier than their foraging ecology should allow. van Schaik shows in other work that it was this social cooperation and complexity the allows for the expansion of large brains of humans, particularly the neocortex (and to some extent falls in line with Robert Barton's earlier proposals).

Vampires arguably have the most highly specialized feeding ecology of bats, and there has been an enormous amount of adaptation in their morphology, physiology, and neurology that has facilitated this. The main issue here is whether the feeding behavior of vampires is a complex skill that needs a lot of time to be learned, and is perhaps facilitated through adult food sharing (Schuppli's Fig 2C). Vampires do have many characters that make them similar to primates, but making the life history comparison directly between vampire bats and humans is a bit bold -- vampire bat specialization and life history is more similar to aye-ayes or some of the haplorhine primates than humans (or maybe just a capital breeder since blood is a relative poor resource).

Maybe the best way to present this would be instead of going directly to a simplified view of humans, instead present the suite of socioecological characters that make vampires unusual and show in various taxa how these characters generally accompany complex foraging skills and extended developmental periods needed to learn those skills. You could present humans as a special case where skill acquisition is even further extended beyond what is expected (= AFR), but this is only possible because of social features like resource pooling, allomaternal care, etc. Then why vampires are an interesting case because they show some, but not all of these characters and you want to test the role of preferences and learning in their feeding. Or something to that extent.

It would be interesting to then revisit some of these life history and behavioral traits in your discussion for why you didn't really find learning (particularly their ASC/AFR relationship -- seems a tidy heuristic at the moment). This would go beyond the current 'it's complicated' approach to learning in bats presented in the discussion/conclusions and suggesting the need for better experiments and larger sample sizes. My feeling is that since there is so much specialization in these bats that the motivation at the root of their feeding is instinctual, but that they need to practice to get really good at it. You have a tough mix of results. The young bats attempt to feed more on the chicken, but are pretty terrible at it -- which speaks to the importance of practice / learning. But there was no social facilitation observed in the data, just that some young bats try to feed on a prey item that only 2 of 3 wild bats fed on. It would be useful to know a couple of other developmental landmarks relative to their long lactation period e.g., when adult incisors are fully erupted & functional, and when young fledge, dispersal age, to understand the necessity for learning or how long there really is for these bats to practice before they are independent

Some specific comments:
As a matter of preference, double spacing a manuscript for review makes it easier to read. People who study this type of thing suggest that double spacing decreases read time and lateral masking, reduces the number of fixations, and results in more accurate return sweeps during reading. Especially for reading on a computer screen.

24. What is significant weight?

148-149. I'm not clear on the feeder setup. Were there 2 spouts per temp reservoir that were 5 cm apart or were the 2 feeders separated by 5 cm? Also, were these lixit type spouts, or did the blood pool out for them to feed from?

335-336. I disagree. I would not say that Trachops is highly adapted (at least not in the Desmodus specialization sense). It is very similar to the rest of the gleaners. It has _additional_ low-frequency hearing and many physiological and behavioral redundancies for when it captures toxic prey. It's an extremely flexible predator whose perception still plays by most cognitive rules. We still don't know what proportion of its natural diet really comprises frogs since they also are keen katydid hunters.

342-344. Disulfides are also likely essential for social learning via odor, at least in mice and rats (e.g., Munger et al 2010 DOI 10.1016/j.cub.2010.06.021), and it seems to also be the case for what we found using displaced cues in Uroderma bilobatum.


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

Input:
Title: THE ROLE OF PAST EXPERIENCE IN DEVELOPMENT OF FEEDING BEHAVIOR IN COMMON VAMPIRE BATS
Review round: 2
Reviewer: 1

Output:
Basic reporting: No comment
Experimental design: No comment
Validity of the findings: No comment
Additional comments: All my suggestions to the previous version of this manuscript have been addressed, and I have no further comments. In fact, the manuscript has improved significantly and the methods and results are much more straightforward. Great work!


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

Input:
Title: THE ROLE OF PAST EXPERIENCE IN DEVELOPMENT OF FEEDING BEHAVIOR IN COMMON VAMPIRE BATS
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 nicely addressed my concerns. Thank you for the thoughtful and clear responses to both reviewers.


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: INFLUENCE OF ACUTE COMBINED PHYSICAL AND COGNITIVE EXERCISE ON COGNITIVE FUNCTION: AN NIRS STUDY
Review round: 2
Reviewer: 1

Output:
Basic reporting: 1.	The English language of the article should be improved. Current English is hard to  understand and not clear. Also, some words (like influence and join) were not selected properly and it causes reading difficulties.
2.	Both introduction and discussion parts have some problems related with the references. Please cite the necessary articles for the fNIRS and NIRS systems (line 75-79) and check the bibliography for double cited references (Ludyga et al).
3.	There are many missing literatures in relation with the cognition and exercise such as Byun et al 2014, Lee et al 2016, Browne et al 2016, Netz et al 2016, and Peruyero 2017 (http://dx.doi.org/10.1016/j.neuroimage.2014.04.067, https://doi.org/10.1186/s13063-016-1650-4, http://dx.doi.org/10.1016/j.rppede.2016.01.005, 10.1016/j.bandc.2016.08.002, https://doi.org/10.3389/fpsyg.2017.00921). Please consider these articles when you are describing the gap in the literature.
4.	The authors should clearly describe the motivation behind the selection of older adults to the study and link that motivation with the hypothesis. In other words, why did the authors investigate the inhibition control with the older adults? Please justify your aim accordingly and re-write the introduction with that motivation.
Experimental design: 1.	The main research question is missing. Age of the participants should be noted.
2.	The authors should explain the cognitive task section better.  The readers want to know more detail without searching cited articles. A figure that describes the experimental procedure would be helpful. In current construction of the paper, it is hard to replicate the study.
3.	The authors stated that there are four conditions with randomized order in current experimental design. But, this in not clearly stated on the manuscript. From text, someone could easily understand that only three of them counterbalanced across the participants. Please explain it better.
4.	Please indicate the mean age on the text and describe how did you reach the participants.
5.	The authors stated that they used 20 channel fNIRS system but in that section only 14 of them were used. Please describe why you did not use all fNIRS channels? Also, the authors cited figure 1 in here, but figure 1 does not contain any fNIRS related elements. Please correct it and provide a figure, which shows the connected fNIRS probe in a participant or head model.
6.	Please describe the fNIRS analysis section better. For example, your sampling rate is 3.91 Hz and the duration of single stroop task is around 1 min. Please justify how did you get 20 values for each condition in your raw data? Did you average the oxyhemoglobin levels during single task?
Validity of the findings: 1.	In conclusion, the authors stated that “acute single exercise, single cognition and combined exercise enhanced the performance of the executive task”. How did the authors state this? It should be considered that there is slightly enhancement in the control condition as well. Please change that statement for the significant results or describe that better. Additionally, check the English on that section.
2.	The results section have some uncompleted sentences and problematic figures. The legends of figure 2 should be revised according to the text. Also, you provide figure 2a and 2b but there is only one legend for them, which is not descriptive. Please correct them. Also, the figure 2a contains same graph headings. Please correct them.
3.	The results are clear. However, the obtained results are not well discussed with the related literature in the discussion part. Especially, missing articles in parallel with current study is reducing the quality of this manuscript. Additionally, some of the literatures given in introduction is not discussed in the manuscript. Please re-write the discussion part accordingly.
4.	Another important point is the missing discussion of the current results in parallel with age. Please re-shape the discussion and include the age discussion to this section.
5.	Some of terms are really confusing in figures such as l-DLPFC, r-DLPFC. I suggest to use L-DLPFC and R-DLPFC instead of l-DLPFC, r-DLPFC.
6.	There are some missing statements such as pro-test. Please control the grammar and word mistakes.
Additional comments: 1. The aim of this study is to investigate the effects of different physical and cognitive load on inhibition control and cerebral oxygenation. This study is trying to establish the relationship between inhibition control and physical/cognitive loads. Additionally, PFC lateralization during the inhibition process is another topic in this manuscript. However, the manuscript has some serious problems in regard to figures and language errors (such as wrong words, incomplete sentences etc.). Also, some of the missing/not considered literatures are critically important for this study. The authors should re-write the introduction and discussion part accordingly.

2. In bibliography, the abbreviation of the journal in reference number 2 is correct. But, the PeerJ guidelines clearly indicated that the authors should write the full title of Journal. Please double check your bibliography according to that statement.


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: INFLUENCE OF ACUTE COMBINED PHYSICAL AND COGNITIVE EXERCISE ON COGNITIVE FUNCTION: AN NIRS STUDY
Review round: 2
Reviewer: 2

Output:
Basic reporting: The paper comes across as well considered and provides a nice overview of the relevant literature. There are sections where the sentence structure makes it hard to follow (especially in the discussion), which could benefit from English language editing. I have highlighted a few examples under general comments.

The use of abbreviations in the results section hampers readability; the C, CE, SC, SE conditions could easily be replaced by control condition, cognitive exercise, physical exercise, and cognitive+physical exercise.

The titles in Figure 2 seem incorrect (all plots are titled “L-VLPFC”, this needs to be corrected) and the bar subplots in Figures 2 and 3 do not seem to add anything to the more detailed subplots. I would suggest to remove these bar subplots, and add the symbols indicating significant differences to the remaining subplots.
Experimental design: Based on the introduction, the paper seems to provide novel insights. The research question is well defined and clear. The methodology seems rigorous, although I cannot comment on technical specifics. It is not entirely clear whether a baseline fNIRS measurement is repeated before every Stroop task condition; I can imagine that increased skin blood flow due to physical exercise could affect the fNIRS substantially. Please clarify if this was the case.

The paper does not mention ethical approval; a statement that ethical approval for the study was obtained from an appropriate committee would need to be added.
Validity of the findings: The study investigates oxygenated blood flow to the prefrontal cortex (using fNIRS) as a potential explanation for changes in cognitive function (mainly inhibitory function) after cognitive, physical or combined cognitive and physical exercise. A reason for recruiting (young) older people is not provided but could be added if relevant.
Additional comments: Major:
1.	It is unclear whether ethical approval was obtained for this research.
2.	It is unclear whether Oxy-hb is expressed as change compared to a baseline measurement right before the conditions of interest (to minimize increased blood flow artifacts). If not, this could drive the findings with respect to increased Oxy-hb after physical exercise.
3.	The English language should be improved to ensure that an international audience can clearly understand the text. Examples are: L78, L85-86, L145, L156, L254, L260, L261-267, L285 – the current phrasing makes comprehension difficult.

Minor:
•	L71, “In recent studies assessed by …”: should this be “In recent studies that performed human brain imaging, a change in brain oxygenation it has been … intervention ...”.
•	L80: how does the higher activation related to function? Is more better or is there a saturation effect?
•	L82, “prefrontal”: should this be “prefrontal areas”?
•	L93, “demand of exercise”: “may be one of the potential …”
•	L95, “the question whether in combined exercise exceeds single exercise in acute exercise…”: please revise to clarify that this refers to a combination of (I assume) cognitive and physical exercise.
•	L98, “Dual-task …”: the link of this sentence to the rest of the section is unclear, consider revising.
•	L102, “According to neurovascular coupling …”: should this be “neurovascular coupling theory”? Plus: does the metabolic activity in cerebral blood flow increase or is the metabolic activity assessed as an increase in cerebral blood flow? Please revise this sentence for clarity.
•	L105, “the first aim of the study is the oxygenation …”: is it to “effect of exercise on the oxygenation …”?
•	L108, “while may be regulated”: please clarify what may be regulated.
•	L113, earlier research: please specify the “different benefits” of Tai Chi and brisk walking on cognition.
•	 L116: Please specify what Ludyga et al. found and detail its relevance to the current study.
•	L118, “…to address the gap which the effects …”: would this mean “to address the gap in knowledge on the effects …”?
•	L134: please briefly explain the PAR-Q here with appropriate referencing.
•	L137, “and cognitive function MMSE.”: “and cognitive function was assessed using the MMSE”.
•	L137, “… a reading control condition was complicated …”: “was completed”. Please add a rationale for this control condition.
•	L139: why was the control condition not randomized with the other 3 conditions? Could this have affected the results?
•	L143: Please clarify what VFT means.
•	L147: please add the rationale for allowing people to practice before the assessments. Please also clarify when the control condition was assessed.
•	L150: how was the fNIRS position standardized?
•	L157: which two heart rate values?
•	L161: please clarify the naming and executive tasks. Where these a simple color recognition task and the incongruent condition of the Stroop test?
•	L166: would this method of responding make the task even more difficult (since there is now a working memory component (recall of mapping) as well)? How would this affect the interpretation of the results? Would a difference score of “executive condition” minus “naming condition” provide a more pure measure of inhibition?
•	L167: were the cues always presented as the letters “XXX” displayed in different font colors? I am unfamiliar with this task, what was the rationale for not using the congruent condition of the Stroop test?
•	L169, “… and part of the stimuli is a rectangle appeared around the word, …”: I am not sure what this means, please clarify in the manuscript.
•	L187: please provide the appropriate references to the software including version numbers.
•	L190: would analysis of the error trials provide additional insight?
•	L197: please clarify that the 4 x 2 RM ANOVA was 4 (conditions) x 2 (assessments). Consider to give the “naming” and “executive” task a more intuitive name; I’d consider both to require executive functions (especially since the response needs to be provided by pressing a keyboard key).
•	L222-225: please clarify that the conditions were significantly higher “in oxy-hb levels”.
•	L222: Should the posthoc tests be corrected for multiple comparisons?
•	L226: “… there was a significant main effect …”.
•	L239: please discuss the specific findings here: which conditions had a higher oxy-hb level, and which conditions did not differ in oxy-hb.
•	L239, “… regardless of combined cognition”: “regardless of the addition of cognitive exercise”.
•	L242: Please discuss potential explanations of this finding.
•	L248 (and more often), “protest and posttest”: “pretest and posttest”.
•	L252: please specify what Ludya et al found and why it is relevant to this study.
•	L254: are these sensitivity results? Please clarify what was done, how and why.
•	L259: please provide a reference for this finding. I assume this might be Hsieh et al. 2016, which is mentioned in the next sentence, but this should be made clear in this sentence as well.
•	L274, “… reporting bilateral LPFC”: “LPFC activity”.
•	L284: Would this be “A potential explanation for this finding”? Plus: the reference style seems off here.
•	L285, “explicated”: “showed”?
•	L293, “that brings stimuli’s information”: “involved in processing/recognition of visual stimuli”?
•	L307, “only acute combined exercise”: both SE and CE show an effect, please revise.
4.	Discussion: The results indicate that both CE and SE improve performance on the cognitive tests and increase PFC oxygenation. However, the paper does not discuss whether the additional cognitive component in CE amplified the effects of acute physical exercise on cognitive performance while this would be of interest to readers. Consider to add discussion of CE vs SE.
•	Figure 1: What do the error bars depict, are these SD or SEM?
•	Figures 2 and 3: Please add a line depicting the average effect to the subplots.
•	Table: please correct the precision of BMI and VO2peak, 2 decimal digit seems sufficient. Should “MMSN” be “MMSE”? Please discuss the relevance of the VO2peak here and consider to add the heart rate measurements to this table.


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: INFLUENCE OF ACUTE COMBINED PHYSICAL AND COGNITIVE EXERCISE ON COGNITIVE FUNCTION: AN NIRS STUDY
Review round: 2
Reviewer: 3

Output:
Basic reporting: The standard of English was very poor, with grammatical errors, inappropriate use of words and many repetitions. In places it was difficult to understand what information was being conveyed.

The concept of 'inhibition control' which the study focuses on was not well defined.
A topic very relevant to the study that was not discussed adequately or referenced sufficiently is vascular regulation of the cortex and how exercise may change this long term.

'Acute' exercise needs to be defined.

Inadequate references are given. For example, in line 337, the MMSE was selected without appropriate referencing.
Experimental design: The research fits the Aims and Scope of the journal.

It is not clear how the fNIRS analysis was performed. In lines 191 - 192, it is stated that the change is calculated by 'subtracting the mean Oxy-Hb of the pre-intervention task period from the mean Oxy-Hb of the post-intervention period'. However in lines 182 - 184 it discusses changes in concentration compared to baseline.

How was the position of the optodes relative to the cortical region (i.e. DLPFC, VLPFC) assessed? Were MRIs taken or a 3D digitiser used?

Description of the naming and executive condition used in the Stroop test is needed.

How long after the acute exercise was the fNIRS measured?

Were any Bonferroni (or other) adjustments made for the multiple comparisons?
Validity of the findings: The standard deviations in figure 1B for the executive condition are very high. The significant main effect of time - p<0.001 is surprisingly. Were the data normally distributed and were there outliers which could account for the significant change reported?
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: INFLUENCE OF ACUTE COMBINED PHYSICAL AND COGNITIVE EXERCISE ON COGNITIVE FUNCTION: AN NIRS STUDY
Review round: 3
Reviewer: 1

Output:
Basic reporting: The improvement of the MS is very satisfactory now.
Experimental design: I still have some questions which were placed in general comments.
Validity of the findings: No comment
Additional comments: Thank you for your efforts to enhance the quality of the paper. Nearly, all of the suggestions from my side were completed accordingly. However, I have still further minor revisions for the paper.

Abstract
Line 58: The fNIRS findings show that acute single exercise influences oxygenation for executive tasks but not oxygenation for naming tasks. Should be replaced as “The fNIRS findings showed that acute single exercise influences oxygenation for executive tasks but not for naming tasks.

Line 59: Greater increment was observed in the post-exercise session of combined exercise during the modified Stroop. Replaced  with the “Greater improvement was observed in the post-exercise session of combined exercise during the modified Stroop.”

Introduction
Line 90: “improved” should be replaced with “improve”

Line 98: “Moreover, few studies studied the mode of acute exercise.” should be replaced with “Moreover, there are limited number of studies in which the effects of different types of exercise were investigated”

Methods
Line 177: VFT abbreviation is used first time in this section besides the abstracts. So, please add the “Verbal Fluency Task” before the VFT abbreviation.

Line 181-182: “The cognitive + physical exercise (CE + PE) paradigm combined walking at the same intensity as single exercise with cognitive + physical exercise.” This sentence is not clear. What do you mean with walking at the same intensity?

Line 209: Experimental procedure figure for whole paradigm would be still helpful. There is no explanation about the stages for combined exercise. When did you apply the cognitive tasks during the CE+PE condition? Please describe it better.

Line 229: The figure of the optodes were not good enough for the demonstration in regard to resolution. Please consider the literature you already referred in the manuscript. At least 5 or 6 of them represented the fNIRS optodes in a better way. You can easily take a photograph of a participant forehead and mark the ROI on that.

Line 233: “filtered” should be replaced with the “filter”

Line 243: Please correct the term as SPSS (v19.0).

Results
The resolutions of the figure 3 and 4 should be noticed and improved. Especially, figure 4 is hard to read due to this problem.
Line 249 and 261: Please revise the first sentence of the response time and accuracy related results section. For example: “When the response time was considered, in the naming task,…”
Line 257: “pro” will be “pre”

Discussion
Line 286: “modes of exercise” can be replaced with the “type of exercise”


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: INFLUENCE OF ACUTE COMBINED PHYSICAL AND COGNITIVE EXERCISE ON COGNITIVE FUNCTION: AN NIRS STUDY
Review round: 3
Reviewer: 2

Output:
Basic reporting: The English has been improved considerably. There are a few sentences that could be improved, and I hope that my co-reviewers point these out, but overall the paper is understandable to a general scientific audience.

The new Figure 1 is a great addition. The current figures 2 to 4 are of very low quality in the reviewer's version, please provide a high-resolution version with the revision.
Experimental design: I am still not convinced that the findings are not influenced by an artefact in fNIRS due to increased skin blood flow after physical exercise. It was unclear from the author's response if they considered or accounted for this. If they haven't, this should be discussed as a limitation to this study. If they have, this should be made more clear in the article.
Validity of the findings: No comments
Additional comments: No additional comments


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

Input:
Title: INFLUENCE OF ACUTE COMBINED PHYSICAL AND COGNITIVE EXERCISE ON COGNITIVE FUNCTION: AN NIRS STUDY
Review round: 3
Reviewer: 3

Output:
Basic reporting: Improved language structure.
More information on changes in cognitive function in ageing would be appropriate.
Figure 3 needs editing.
The aims need to be more clearly presented.
Experimental design: Pleas describe how the optodes were placed.
Validity of the findings: Limitations of fNIRS analysis should be added with discussion of artefacts related to talking and changes in haemodynamics on the different days.
Additional comments: General comments

The manuscript is much improved and many points clarified.
However, there are still grammatical errors eg. Lines 167-171.

All acronyms need to be defined eg VLPFC, DLPFC, LPFC

Methodology is not clearly described.  For example, how long after the exercise conditions was the modified Stroop condition applied?
The fNIRS was recorded at rest before the intervention and again for one minute after the intervention.  Was this during the modified Stroop test when the participant was talking, which would have introduced an artefact in the signal?

The naming condition needs to be better defined and more description added to Figure 1.

In response to the question regarding placement of electrodes the reply was ;’…the software integrates a 3D digitiser…’. Please add this to the text with reference.

Again, standard deviations are very high in Figure 3B and yet p < 0.001. Please provide a scatter plot of the individual data.

In figure 3 change the ‘C’ along the x-axis to ‘RC’. Change ‘SC’ to ‘CE’, ‘SE’ to ‘PE’ and ‘CE’ to ‘CE&PE’.

Table 3 – typos ‘Hight’, ‘MMSN’. Change 0.45 in M/F to number of male/female participants.


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: SPATIOTEMPORAL PATTERNS OF URBAN THERMAL ENVIRONMENT AND COMFORT ACROSS 180 CITIES IN SUMMER UNDER CHINA’S RAPID URBANIZATION
Review round: 1
Reviewer: 1

Output:
Basic reporting: Literature references, sufficient field background/context provided
Experimental design: Research question well defined, relevant & meaningful. It is stated how research fills an identified knowledge gap.
Validity of the findings: Conclusions are well stated, linked to original research question & limited to supporting results.
Additional comments: Major revision


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: SPATIOTEMPORAL PATTERNS OF URBAN THERMAL ENVIRONMENT AND COMFORT ACROSS 180 CITIES IN SUMMER UNDER CHINA’S RAPID URBANIZATION
Review round: 1
Reviewer: 2

Output:
Basic reporting: Please refer to the general comments.
Experimental design: Please refer to the general comments.
Validity of the findings: Please refer to the general comments.
Additional comments: The study has investigated thermal discomfort in 180 cities in China using a variety of datasets for 1990, 2005, and 2015. The topic is interesting and the findings are useful. However, the paper should be revised and the data/methods need to be clarified. There are many writing issues in the paper, and I tried to indicate some of them in the following, but please do a thorough writing check on the manuscript. Please find my comments as follows:

1. Line 64: “exhibit an decreasing” >> “exhibit a decreasing”
2. Line 65: “cities had positive” >> “cities that have positive”
3. Line 68: what does “GDP” stand for? Is it the gross domestic product? If so, the sentence does not read well; “the most important contribution to increasing PET is GDP”. GDP is a monetized measure and does not cause an increase in temperature.
4. Line 90: “become worsen” >> “become worse”
5. Line 112: “critically very necessary” >> “critically necessary”
6. Line 115: “have experienced” >> “has experienced”
7. Line 125: “were still” >> “are still”
8. Lines 126-127: the sentence does not read well. Please rephrase it.
9. Line 127: “an national” >> “a national”
10. Line 160: “was used” >> “were used”
11. Line 161: “Four variables” >> “the four variables”
12. Section 2.3, Calculation of urban thermal comfort: It is not clear how the authors have calculated the Physiological Equivalent Temperature (PET). They only mention that “PET was derived from the human energy balance” (Line 188). Is there an equation to calculate it? Or did the authors use a specific software package? Please explain how PET is derived in details.
13. Line 191: “monthy” >> “monthly”
14. Line 191: “The monthly PET index … was calculated” >> Does it mean that monthly data are used for calculating PET? Please explain the data in more details (temporal resolution that is used, period, variables). You may even consider summarizing the information in a small table.
15. Line 221: Fig.4 is introduced before Figures 2 and 3. The order of figures should be revised.
16. Fig.2 : It is better to revise “air humidity” to “relative humidity”. In addition, the legends do not read well. Please revise them and please include the legend for the green bar (2015) to all three plots.
17. Figures 5 and 6: In these two figures, what does “1990-2005” indicate? Does it show PET1995-PET2005? Based on the conclusions in the text, it seems that it should be 2005-1995, since the authors mention that “95% of cities had positive trend” (Line 241). Please revise the legend in both figures accordingly.
18. Discussion: It is important to discuss about the limitations of this study and their effect on the final conclusion. For instance, it has to be mentioned that the analyses are solely for three different years and the so-called “trends” do not necessarily indicate long-term changes, but they merely show year-to-year differences. Furthermore, it seemed that monthly data was used and then averaged over the season. Therefore, the study does not capture the severity of thermal discomfort neither its peaks, and it only provides a general condition for each city in each year.
Another important discussion to include in the paper is the expected impacts of climate change on the issue. I think the authors may find the following recently published study useful for this matter:
• Sun, Q., Miao, C., Hanel, M., Borthwick, A. G., Duan, Q., Ji, D., & Li, H. (2019). Global heat stress on health, wildfires, and agricultural crops under different levels of climate warming. Environment international, 128, 125-136.


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: SPATIOTEMPORAL PATTERNS OF URBAN THERMAL ENVIRONMENT AND COMFORT ACROSS 180 CITIES IN SUMMER UNDER CHINA’S RAPID URBANIZATION
Review round: 2
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The revisions are very good. I think the manuscript can be accepted 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: SPATIOTEMPORAL PATTERNS OF URBAN THERMAL ENVIRONMENT AND COMFORT ACROSS 180 CITIES IN SUMMER UNDER CHINA’S RAPID URBANIZATION
Review round: 2
Reviewer: 2

Output:
Basic reporting: Pass
Experimental design: Pass
Validity of the findings: Pass
Additional comments: Thank you for point by point response to all the comments. All my suggestions are addressed in the revised paper.


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: AGE-ASSOCIATED CHANGES OF CYTOCHROME P450 AND RELATED PHASE-2 GENE/PROTEINS IN LIVERS OF RATS
Review round: 1
Reviewer: 1

Output:
Basic reporting: No comments
Experimental design: No comments
Validity of the findings: No comments
Additional comments: A research manuscript by Shangfu Xu et al. entitled “Age-associated changes of cytochrome P450 and related phase-2 gene/proteins in livers of rats” submitted to Peer J for critical evaluation. The authors aim to increase our understanding concerning the dynamics of cytochrome P450 and other related genes/proteins in rat liver. The manuscript presents three patterns of gene expression in livers of male rat at fetus, neonates, weaning, puberty, adulthood and aging based on qPCR and western-blot analysis. Even though similar results have been reported in mice, this study focused on the effect of aging on cytochrome P450 in rat still looks relatively novel and might provide some useful information for other researchers to study age-related disorders. However, some concerns should be addressed before publication.
Major concern(s):
This study aims to elucidate the effect of aging on the dynamics of cytochrome P450 expression in rat liver. To eliminate gender-biased results and make the conclusions more convincing, female animals should also be included in this study.
Minor concerns:
1.	Line 104, in this part, the results from human mainly focused on neonates and young children, aging-induced changes of these enzymes in human should be added to make it more relevant to this study. Also, please check the mistake of the sentence in line 105.
2.	Line 60, please add reference(s) to this sentence.
3.	Did not find the supplemental Table 1 in supplemental materials, comments from DMD also missing in supplemental files.
4.	It’s a little difficult to follow the results by the figure legends, please show as A:…；B：…；C：…… to briefly introduce each panel.
5.	In figure legend of Fig 1, it seems CYP1A1 has been checked but the WB results are CYP1A2. Please confirm this.
6.	Why this study did not include the -2d, 1d and 7d samples for the western-blot?


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: AGE-ASSOCIATED CHANGES OF CYTOCHROME P450 AND RELATED PHASE-2 GENE/PROTEINS IN LIVERS OF RATS
Review round: 1
Reviewer: 2

Output:
Basic reporting: the ms needs to be rewritten since the present form is not acceptable.
Experimental design: This section should be rewritten and I have included my comments in this section
Validity of the findings: Needs more investigations
Additional comments: The study present by Shangfu et al., showed the changes in expression of different CYPs using western blotting and Real time PCR in livers of rats at different ages. The study is interesting and has a novelty that could be added to the field of drug metabolizing enzymes. However, I have major concerns regarding the following points:
1-	Introduction: The rationales of the study should be clear and texted in separate paragraph.
2-	Authors named CYP families in the introduction as 1, 2, 3, and 4, wheras in Materials and Methods, they named CYPs as CYP4A1; 7A1, 3A1; 2E1 and so on. Authors should clarify the families of CYP and their isozymes in the introduction.
3-	Different primers of   different CYP genes identified by Real time PCR should be included in Methods.
4-	The corresponding enzyme activity for each CYP isozyme should be included to give more evidence to the changes in its corresponding CYP isozyme. For example, dimethylnitrosamine N-demethylase is the corresponding enzyme of CYP2E1. Please see the following citations (PLoS One. 2016 Nov 1;11(11):e0165667; J Helminthol. 2002 Mar;76(1):71-8.; Curr Drug Metab. 2000 Sep;1(2):107-32. Review.)
5-	Authors mentioned in page 14 lined 186”The expression of CYP-1 family is shown in Figure 1” and should be corrected into CYP 1A2 since family 1 contains several of CYP isozymes.
6-	Authors mentioned in page 14 “Cyp2a1 increased dramatically after birth, reached 250-fold at 200 weanling (21 day), and peaked on 35 days of age (1100 fold). It was gradually declined afterwards.”. This is not true since no change in CYP 2a1 after day 28 until 800 d.
7-	Why authors did not study the gene and the protein expression for the same isozyme. They did RTPCR for 1A1 and Western blotting for another isozyme, Cyp2a1. It was very important to perform both expressions for the same CYP since both expressions could confirm each other.
8-	My comment No.7 is also the same for figures 2,3 and 4.
9-	In page 27, description of CYP 2 family is the same of figure 2 legend. Data of figure 2 should be described in details for each parameter.
10-	 The same in figure 3.
11-	Authors mentioned in Materials and methods that Band intensities were semi-quantified by densitometry using Quantity One® software (version 4.6.2, Bio Rad”. Where are the data of band densities? The band densities of each CYP should be presented as histograms in separate figures.
The manuscript in its present form needs extensive revision.


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: AGE-ASSOCIATED CHANGES OF CYTOCHROME P450 AND RELATED PHASE-2 GENE/PROTEINS IN LIVERS OF RATS
Review round: 1
Reviewer: 3

Output:
Basic reporting: In my opinon, this manuscript had been written in English clearly, and the structure of this manuscript is good for publishing in PeerJ.
Experimental design: in the experiment design, the major problem is that the enzyme activities of these CYPs in rats should been detected by using their selective probes. Only detecting  proteins levels by WB or PCR is not enough to support the results.
Validity of the findings: The data is statistically sound, and the conclusions was appropriately stated, but experiemntal data did not support the conlusion of authors in the present form. The real activities of CYP isoforms should be measured in the revision manuscript to prove the conlusion.
Additional comments: In my opinon, this manuscript had been written in English clearly, and the structure of this manuscript is good for publishing in PeerJ.
But in the experiment design, the major problem is that the enzyme activities of these CYPs in rats should been detected by using their selective probes. Only detecting  proteins levels by WB or PCR is not enough to support the results.


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: AGE-ASSOCIATED CHANGES OF CYTOCHROME P450 AND RELATED PHASE-2 GENE/PROTEINS IN LIVERS OF RATS
Review round: 2
Reviewer: 1

Output:
Basic reporting: no comment
Experimental design: no comment
Validity of the findings: no comment
Additional comments: The authors have addressed most of the concerns, I think this revised manuscript is suitable to be published in Peer J.


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: HISTOLOGICAL EVALUATION OF FIVE SUTURE MATERIALS IN THE TELSON LIGAMENT OF THE AMERICAN HORSESHOE CRAB (LIMULUS POLYPHEMUS)
Review round: 1
Reviewer: 1

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


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: HISTOLOGICAL EVALUATION OF FIVE SUTURE MATERIALS IN THE TELSON LIGAMENT OF THE AMERICAN HORSESHOE CRAB (LIMULUS POLYPHEMUS)
Review round: 1
Reviewer: 2

Output:
Basic reporting: No comment.
Experimental design: No comment.
Validity of the findings: No comment.
Additional comments: The authors have incorporated all suggested revisions along with including additional necessary clarifications in the resubmitted manuscript. Excellent work.

There are a few minor revisions needed:
Line 112 – Change Fiberglas to fiberglass.

Line 156 – ‘Biopsies were performed using metzenbaum scissors to take a 3-5 mm snip of tissue by one of two…’ Recommend reword biopsy description… Consider instead…’Excisional biopsies were performed using metzenbaum scissors, a 3-5 mm diameter tissue sample was obtained by one of two surgeons (AEK or AH) for the control and experimental groups..’

Line 168 – Change semi colon to comma after ‘et al.’

Line 179-181 - Prior to systemic clove oil injection, heart rate was measured at 8 bpm and 30 bpm in the two subjects of the experimental group where heart rate was monitored. Consider change sentence to ‘Prior to systemic clove oil injection, heart rate was measured in two of the experimental group subjects. Heart rates were 8 bpm and 30 bpm.’

Line 187-188 - In three subjects, where it took up to 15 minutes for cessation
Of movements, up to 4 mL of clove oil was required. Recommend reword sentence to ‘Three subjects had prolonged limb/gill movements and required higher doses of intravascular clove oil, up to 4 ml were administered (or list a range used ‘3-4 ml’)... or something similar.

Line 390 – Insert space after ‘hemolymph,’

Figures 4 and 5 caption abbreviations not evident in the histopath photomicrograph images.


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: TAXONOMIC REVISION OF BLACK SALAMANDERS OF THE ANEIDES FLAVIPUNCTATUS COMPLEX (CAUDATA: PLETHODONTIDAE)
Review round: 1
Reviewer: 1

Output:
Basic reporting: This is an excellent paper overall and should be published once concerns about the multivariate statistical analysis have been addressed (see Experimental design, below). The results clarify the taxonomy of the Aneides flavipunctatus complex to better reflect species-level diversity and evolutionary history. The sampling is impressive and undoubtedly represents thousands of hours of fieldwork and labwork over the years. The manuscript is clearly written, although I urge the authors to consider using the “active voice” in order to improve the strength and impact of the writing. It may not be the authors’ style, but in my opinion would greatly improve the manuscript. I’ve made minor comments and corrections throughout the pdf, which I’ve included. The figures are nice and photos are awesome, but there is a mistake in the legend in Figure 5.
Experimental design: The research question is well-defined and this work clearly fills a knowledge gap. In terms of analysis, the phylogenetic analyses were performed well and support the authors’ conclusions. With respect to the multivariate analysis of morphological data, I have some concerns. I don’t think it makes sense to perform a PCA here. PCA is used as an exploratory method to recover patterns in the data. So, for example, if you didn’t have any pre-defined groups (i.e., you didn’t know the species boundaries from the genetics), you could perform a PCA on the entire complex to see what groupings you found. In this case, however, you already have your 4 well-defined groups based on genetics (species) that you wish to test for differences among. The PCA is unnecessary.

With respect to the discriminant analysis, the canonical portion of the discriminant analysis is appropriate as you’ve performed it. However, the classification function is not (at least as described). The classification function should be validated with a completely separate set of samples, for example, specimens that were not used in creating the canonical function. The “% classified correctly” will always be high (deceptively so) when the prediction is made on the same sample that was used to develop the function. So, I would either take out the classification portion of the discriminant analysis, or perform it correctly by validating it using a resampling procedure (e.g., jackknifing). Alternatively, with a large enough sample, you can use split-sample validation to randomly divide the total sample into two groups, one for analysis, one for validation. I’m not sure your sample size is large enough to do this, however.
Validity of the findings: The findings are valid, though there is one speculative statement (lines 477-478): "...likely insufficient to ever merge these species." I'd take that last bit out. Yes, the genetic estimates suggest a narrow contact zone and low migration, but that's a bit of a leap in my opinion.
Additional comments: This is a great paper, very well done.


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

Input:
Title: TAXONOMIC REVISION OF BLACK SALAMANDERS OF THE ANEIDES FLAVIPUNCTATUS COMPLEX (CAUDATA: PLETHODONTIDAE)
Review round: 1
Reviewer: 2

Output:
Basic reporting: No comment
Experimental design: See my comments and suggestions listed in the General Comments for the Author section.
Validity of the findings: Strengths of the Manuscript:
Taxonomy is often overlooked or relegated to journals specializing in taxonomic descriptions. This paper represents an ideal blend of historical and recent genetic studies all of which point to a conclusion of multiple species of these salamanders. These three new species represent a significant increase to a morphologically and ecologically specialized genus of salamanders with relatively few members making the description important for conservation purposes and to provide a framework for studying diversification. The authors were able to weave in the discussion of species relationships and other important considerations within the species descriptions themselves making the descriptions a key part of understanding the evolution of these taxa.

The historical facts discussed are an important part of both the species descriptions and key to understanding the species names and previous ideas regarding their status. I have included some suggestions regarding some of the historical details below.

The writing is excellent and clear with the exceptions noted in comments for the author.
Additional comments: Review of Taxonomic revision of black salamanders of the 1 Aneides flavipunctatus complex (Caudata:Plethodontidae) (#36410) by Sean B. Reilly and David B. Wake:

Points by importance, then in order:
1) For the morphological analyses, I assume that all of the specimens listed (the paratypes and holotypes) constitute the points in Figure 4 and the numbers in Table 2. If so, this should be made more explicit both in the methods and materials and in the results. In addition, being able to identify the individuals present in the contact zone in Figure 2 would help to either show whether individuals close the contact zone are distinct in morphometric space, or not. I suggest circling or numbering the individuals in Figure 4 that are from populations listed in Figure 2. Perhaps a subscript number with the population numbers from figure 2.
For table 2, it would be useful to know if the misidentified specimens were in the contact zone, or not.
In addition, I would like to see the distinction (or lack thereof) morphometrically (specifically referring to Figure 4 and Table 2) in the contact zone discussed somewhere in the manuscript. If animals close to the contact zone are more distinct ten elsewhere, this raises the possibility of reproductive character displacement and/or reinforcement.
2) Figure 5
Aneides flavipunctatus is mislabeled as Aneides niger (repeated twice on the figure)
3) Figure 3 - The red coloration of the new samples is confusing as they appear to indicate Aneides klamathensis sequences because that species is indicated with red - I suggest a symbol like an asterisk to indicate new samples
The figure legend should state clearly that there is no mitochondrial introgression between geographic localities.
Within the box, “Populations 7-12” in the upper box and 1-6 in the lower box would help to clarify the distinct nature of the populations.

Lines 63-64 This sentence should be re-worded to make it clearer that Storer considered Aneides iecanus synonymous with Aneides flavipuctatus and therefore considered all previous references to Plethodon iecanus, Aneides iecanus and Autodax iecanus to all be considered Aneides flavipunctatus.

From Storer 1925:
Plethodon flavipunctatus Strauch (1870, pp. 71-72). Original description, type from New Albion [probably the coastal portion of Sonoma County], California.
Plethodon flavipunctatus, Boulenger (1882b, pp. 55-56).
Plethodon iecanus Cope (1883, pp. 24-25). Type locality, Baird, Shasta
County, California.
Aneides iecanus, Cope (1886, p. 526). Generic allocation.
Plethodon iecanus, Townsend (1887, pp. 240-241).
Plethodon flavipunctatus, Cope (1889, p. 145).
Autodax iecanus, Cope (1889, pp. 187-189, text fig. 46). General account. Autodax iecanus, Van Denburgh (1895b, pp. 776-778). Range; breeding
habits; eggs.
Autodax iecanus, Ritter and Miller (1899, p. 696).
Plethodon flavipunctatus, Yan Denburgh (1916, p. 221).
Aneides iecanus, Grinnell and Camp (1917, pp. 135-136, fig. 2). Range. Aneides iecanus, Stejneger and Barbour (1917, p. 21; 1923, p. 18). Range.

Cope had moved Plethodon iecanus to Aneides iecanus in 1886 (see above) - perhaps this should be added to the history of the taxonomic changes.

Line 441 -"manuscript name" does that mean a new name in the Lowe manuscript?

Lines 604-605 Did Cope move iecanus to Aneides in 1886 on page 576? see above comment in Storer 1925:
From Cope 1886:
Notes. — I add here that the Plethodon iecanus Cope proves to be a well- marked species of Anaides. The species was described from a young one.

Cope 1886. Synonymic list of the North American species of Bufo and Rana, with
descriptions of some new species of Batrachia, from specimens in
the National Museum. Proc. Am. Philos. Soc., 23, 514-526.

Line 653-654:
Ieka was recognized as the word for Mt. Shasta in a few other sources:

A History of Northern California; A Memorial and Biographical History
Chicago, IL: Lewis Publishing Co., 1891.
Page 241: "...the tribe of lndians that inhabit Scott and Shasta valleys and the mountains to tbe north. The true name of their tribe they have forgotten or will not tell, having been called Shasta for half a century; but the name of their beautiful patron mountain still remains to us, Ieka, the white."

Also, the town of Yreka was reported to have been originally Ieka --
Snopes:
https://www.snopes.com/fact-check/yreka-bakery/
An 1876 article from the Yreka Journal quoted on Snopes.com:
"It was intended the county seat should bear the name of Ieka, the Indian name of Mount Shasta, but by mistake the name of Wyreka was substituted and the error continued, with the exception of dropping the letter W, thought to be superfluous."

Figure 10
The spots on this specimen as well as living specimens I can recall look blueish - is this incorrect? Is it worth mentioning that the small spotting looks blue from a distance, but is white when examined close up?


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: FLEAS OF BLACK RATS (RATTUS RATTUS) AS RESERVOIR HOST OF BARTONELLA SPP. IN CHILE
Review round: 1
Reviewer: 1

Output:
Basic reporting: This is a descriptive study where authors detected DNA of Bartonella spp. in extracts of fleas collected on Rattus rattus. The structure and logical flow is there, however authors must pay more attention to the correct usage of the bacteria they are talking about (I assume authors talk about for example Bartonella koehlerae and Bartonella clarridgeiae). There are typos in many of the scientific names, which does not shed a good light on the rigor of the study. Overall there is evidence of understanding of the context of the flea/Bartonella knowledge. There are minor syntax and English problems. Sequence data are shared but I am puzzled by the length of DNA submitted. GenBank (BN: not GeneBank) accepts only sequences >200nt, but the suppl. sequences claimed to be submitted to GenBank are <200nt.
Experimental design: The sampling is commendably performed. I am not sure how authors assured that they trapped only Rattus rattus, based on my experience some native rodents will always be captured as well.When collecting fleas how did you assure that you captured all fleas on the rats?

Authors designed new sets of primers for both genes of Bartonella, why did they not use already established sets of primers? Anyhow, it is not clear how long are the fragments amplified esp if the comparison is based on <200bp. It would be desirable that longer fragments are produced.

How did authors prevent cross-contamination? Did you run blanks while isolating DNA?

Was sequencing done bidirectionally. It is highly desirable that it is done so.
It would be good that the alignment is included as suppl material.
Validity of the findings: The sequence length as outlined above needs to be clarified. If the generated sequences are >200nt it should be fine, however if not I would strongly suggest that authors consider amplifying more DNA. Consider protocols outlines in this: https://doi.org/10.1017/S0031182017001263

I found the discussion rather long for the amount of new data. Authors could be bit more succinct. Authors should refrain from repeating the results in discussion.
Additional comments: The references are in rather disorganized state, authors needs to pay more attention to the journal style and be consistent.


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: FLEAS OF BLACK RATS (RATTUS RATTUS) AS RESERVOIR HOST OF BARTONELLA SPP. IN CHILE
Review round: 1
Reviewer: 2

Output:
Basic reporting: •The English language should be improved to ensure that a general audience can clearly understand your text. Major issues include run-on sentences and double-negatives (e.g., lines 53-59, lines 282-285). Minor issues include grammar, pluralization, spacing, and spelling errors throughout the manuscript (e.g., lines 45, 300, Tables 2-4).

•In the introduction (lines 53-53), there are more recent review articles that list Bartonella species detected in vectors for various mammalian hosts (e.g., Tsai et al. 2011., Cheslock & Embers. 2019.). The authors should cite the most recent work in the field. Also, Breitschwerdt & Kordick (2000) is not listed in alphabetical order in the references.

•The authors should consider reducing the amount of text in the discussion. Although I appreciate that they are providing information for each Bartonella species/strain, it is quite laborious to read through and takes away from the overall findings of the manuscript.
Experimental design: •There’s a disconnect between the introduction and the sample localities associated with hydrographic zones and seasonality. The introduction states the aim of the study is to investigate the “prevalence of Bartonella spp. in Rattus rattus fleas in areas with different human population density throughout Chile,” but makes no mention as to why the authors are also investigating different hydrographic zones and seasons. Please explain why these parameters were also examined.

•Please provide more details about the institutional animal care and use regulations (e.g., animal use was conducted in accordance with the Animal Welfare Act, protocol approval number, etc.).

•How were the rodents identified? Please reference taxonomic keys if used.

•Were the fleas collected immediately in the field or later in the lab? If later, were the rodents placed in separate bags or were they all placed together? My concern is that the prevalence and intensity data would be skewed if animals were placed together prior to ectoparasite removal. Please clarify.
Validity of the findings: •Some of the text in the results section does not correspond to the data presented in the tables. For example:
-Lines 188-189: A total of 179 fleas is listed in Table 2 when analyzed by locality (as opposed to the "177" mentioned in text). This number will change the data given in lines 188-189, 226-227.
-Lines 196-198: Plocopsylla wolffsohni is listed under the village locality too, not just in wild areas, according to Table 2.

•Fifteen species of fleas seems like a lot for R. rattus. Similar studies report one to five species of fleas (Loftis et al. 2006, Marie et al. 2006, Li et al. 2007, Reeves et al. 2007, Tsai et al. 2010). The authors do not address this finding in the manuscript. Also, are any of these species new host records? If yes, then there need to be voucher specimens.

•Please provide the confidence intervals for prevalence, intensity, and abundance values to indicate the accuracy of the estimation.

•In the abstract, the authors conclude that “villages in the arid zone correspond to areas with higher infection risk;” however, there are no statistical tests to support this claim in the results section (only prevalence of fleas was significantly higher in the villages, no stats presented in text on prevalence of Bartonella). Additionally, it states in the discussion that “no pattern is observed” between the prevalence of Bartonella in the different hydrographic zones.

•Again, the discussion states “in our study, a significantly higher prevalence of Bartonella DNA in the arid zone than in the other zone,” but there are no stats to back up this claim.
Additional comments: •The references need to be updated to reflect the articles cited in the text. For example, Zouari et al. (2017) is cited, but not listed in the reference section.

•Line 78: The common name for Pulex irritans is human flea, not dog flea.

•Line 82: Define “occupational risk”.


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: FLEAS OF BLACK RATS (RATTUS RATTUS) AS RESERVOIR HOST OF BARTONELLA SPP. IN CHILE
Review round: 2
Reviewer: 1

Output:
Basic reporting: ok
Experimental design: ok
Validity of the findings: ok
Additional comments: The manuscript was in general improved but remains rather long with rather hard to read and understand the details of the results.


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: BIOREMEDIATION OF ENGINE-OIL CONTAMINATED SOIL USING LOCAL RESIDUAL ORGANIC MATTER
Review round: 1
Reviewer: 1

Output:
Basic reporting: In this work, authors have carried out a study on bioremediation of engine-oil contaminated soil using local residual organic matter. The study design considered four treatments notably; ramial chipped wood (RCW), horse manure (MANR), brewers ‘spent grain (BSG), and inorganic fertilizer (FERT). Authors sufficiently used many analytical parameters to support the study. This manuscript would be recommended for publication, however, I would recommend some minor corrections prior to acceptance.
Comments
Abstract
(1) L21 – 22, authors should consider using either “framework of circular economy” or “circular economy’s framework”

Introduction
(2) In L57 – 58, some of the citations used by authors are way too old and not recent enough to warrant a novel study.
(3) Authors are invited to consider the following recommendations wherein strength of locally generated fertilizing materials has been recently demonstrated.
Onwosi et al., 2018
Onwosi, C.O., Nwankwegu, A.S., Enebechi, C.K., Odimba, J.N., Nwuche, C.O., Igbokwe, V.C., 2018.	Bioremediation of Soil Contaminated with Diesel Using Inorganic Nitrogen Sources: Incorporating nth-	Order Algorithm in the Evaluation of Process Kinetics. Soil and sediment contamination. 1 – 9.	https://doi.org/10.1080/15320383.2018.1423023.
Onwosi et al., 2017
Onwosi, C.O., Odibo, F.J.C., Enebechi, C.K., Nwankwegu, A.S., Ikele, A.I., Okeh, O.C. 2017. Bioremediation of 	diesel-contaminated soil by composting with locally generated bulking agents. Soil and Sediment 	Contamination. 26(4), 438–456.

Nwankwegu et al., 2016a

Nwankwegu, A.S., Onwosi, C.O., Orji, M.U., Anaukwu, C.G., Okafor, U.C., Azi, F., Martins, P.E., 2016a 	Reclamation of DPK hydrocarbon polluted agricultural soil using a selected bulking agent. J Environ Manag.  	172, 136 – 142.
(4) L62. Circular economy’s framework. Revise.
(5) L82 – 84 essentially needs citation (s). The following or another would be appropriate;
Nwankwegu et al., 2016b
Nwankwegu, A.S., Orji, M.U., Onwosi, C.O., 2016b. Studies on organic and in-organic biostimulants in 	bioremediation of diesel-contaminated arable soil. Chemosphere.162, 148 – 156.

Nwankwegu et al., 2017

Nwankwegu, A.S., Onwosi, C.O., Azi, F., Azumini, P., Anaukwu, C.G., 2017.  Use of rice husk as bulking agent in 	bioremediation of automobile gas oil impinged agricultural soil. Soil Sediment Contam. 26, 96 – 114.
(6) Consider removing Hattab et al., 2015 in L108 its appearance in L107 is already an ideal. This should be effected where applicable in the entire manuscript.
(7) In L116, authors should replace the verb “has” with have.
(8) The citation “Abioye (2011) as it appeared in L119 should be revised.
(9) L127 – 128, what are those legal regulations? A mention of two or more would sufficiently increase the strength of assertion.
(10) Authors considered more than one treatments in the study so the word “between” as used in L138 should be replaced with “among”
Experimental design: Materials and methods
(11) In L156 according to authors ‘explanation, are they part of the RFM not locally sourced?
(12) In L236, it is not clear if the authors stated how the protocol is done rather than what was done in the present study. Saying “extraction is done” is like stating a general procedure instead of what was done.
(13) L249 – 250, it is noteworthy for attention why the authors chose the use of old citations, whereas more recent studies which discussed the similar approach abound. For example;
Nwankwegu and Onwosi, 2017
Nwankwegu, A.S., Onwosi, C.O., 2017. Bioremediation of gasoline contaminated agricultural soil by 	bioaugmentation. Environ. Technol. Innovat. 7, 1 – 11.
(14) In the whole methodology, it is clear that authors did not monitor the pH fluctuations in the different amendments. Studies have shown that no bioremediation approach is 100% efficient because of different metabolite that maybe produced as bioremediation event lasts. It is also demonstrated that pH in different microcosms under this condition is usually the function of dominant metabolite arising from the amendments. It is actually surprising that the pH of the inorganic fertilizer amended options did not significantly decrease relative to the organic material amended systems but remained stable throughout the incubation. Technically, one may wish to know why? For examples; Orji et al., 2012 (Malaysian Journal of Microbiology), Nwnakwegu et al., 2018 (Environmental Technology), Nwankwegu et al., 2016 (Chemosphere), etc. have attributed substantial contribution to pH dynamics. The pH is therefore as important as temperature and other physicochemical parameters. This could be the first report of pH stability under different treatments.
(16) What do the authors mean by the acronym PCN in L299? Is it similar to PHC as previously used in the previous sections? If yes then it is crucial to maintain consistency in the acronyms if no then initial definition would be more compelling.
(17) In L300, authors may not necessary introduce citations in the result section since it is separated from the discussion and not results and discussion. If this is a statement that needs to be backed by citations then I suggest you lift them to the discussion section.
(18) The word “hovering” in L319 needs to be revised. Consider “was within the commercial & Industrial sites’ regulatory guideline (3500 mg kg-1) instead of hovering. The personification is way too animate.
(19) In L328, replace “between” with “among”
(20) In L338, initial definition of AWCD is important prior to subsequent acronym. AWCD could mean Average Well Color Development.
(21) It is further more surprising that considering what happened from L338 – L351, pH stability was reported. I would therefore challenge the authors to show previous reports of pH stability under similar scenarios. If not I kindly request authors to revise and include pH dynamics as it certainly would affect microbial amensalism and bioremediation efficiency.
Discussion
(22) The study did not consider metabolomics evaluation of PHC degradation hence the statement in L356 –L357 lacks authority
(23) Consider revising this word “hovering” in L415
(24) L422 –L427, what kind of citation is this e.g. Agamuthu et al. (2010)et al.
423 (2010)?
(25) L432 – L433, statement not clear. Revise
(26) Consider changing least amount of time to shortest time in L438.
(27) What do the authors actually mean by the use of the acronym PCH in L441 without initial definition?
(28) L450 – L451, what about cost?
Conclusion
(29) L465, consider changing the infinitive” to source” to “to sourcing”
Validity of the findings: I strongly advocate that the study demonstrated significant validity of finding while I want the authors to further address the effect of pH on PHC removal efficiency
Additional comments: Generally, the whole work especially the conclusion showed great strength but I sincerely recommend more recent citations. This would do pretty much work on the overall strength and novelty.


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: BIOREMEDIATION OF ENGINE-OIL CONTAMINATED SOIL USING LOCAL RESIDUAL ORGANIC MATTER
Review round: 1
Reviewer: 2

Output:
Basic reporting: Line 55-57, should these two sentences being switched? "Several studies demonstrate the potential of RFM as bioremediation agents..However, the MELCC (2014) reports that only 8% of salvaged RFM were used on degraded sites and usage on PHC-contaminated soils was not mentioned. ". So it helps emphasize the importance of this work. Also, authors should add more details of how those studies demonstrate the potential of RFM as bioremediation agents by not actually using RFM on PHC-contaminated soils?

Line 67, what about US? and Canada? I think the whole world is working on circular economy now.

Line 78, typo, "to be covered..."

Line 82, the first sentence can be more specific. appropriate nutrient input in terms of amount and time, right?

Line 86, what is "this parameter"? total N, or different forms of N (inorganic, organic)?

Line 138, "among treatments".
Experimental design: Line 143, "and operated by.." no need using "but"

Line 146, why mentioning "Fe and P" but not other elements (e.g. N, P)? I don't expect to see those because the context of "importance of Fe and P" was not introduced in the Introduction.

Line 177, omit "in"

Line 170-187, if authors could create and add a diagram showing the "flow" of this method, it would help improve readers to understand those steps.

Line 193-194, change to "Samples were taken from 30 cm and 60 cm depths in the first hole, and from 60 cm and 90 cm depths in the second hole"

Line 208-245, this section is a little long, probably have several paragraphs instead of just one. Also, group sentences about ammonia, nitrate and nitrite ions together because they are all about nitrogen.

Line 266, "among treatments".

I appreciate those clear sentences with enough details in the statistical section from the authors!
Validity of the findings: Line 300, I did not expect to see references in the Result section, if there is, it is more like a discussion point.

Line 301-302, "The horse manure (MANR) contained the most potassium, and ramial chipped wood (RCW) contained the least nutrients overall" this sentence is inappropriate, authors comparing "MANR's potassium" vs. "RCW's all nutrients"?

Line 302, "available nitrogen" is nitrogen in forms that ready to be uptake by plants. I assume NH3 is not one of them?

Line 305, how did authors define the nitrogen loss is "low"? within 10%? or this is comparing to the loss of nitrogen for BSG-F?

Line 342, "some differences" this wording is not professional, edit it

Line 360, could be 0 instead of zero

Line 361, omit "seemed to", if that's their results not from their discussion (assumption)

Line 366, it might be interesting to develop a parameter "temperature/moisture" to see the combine effects of temperature and moisture to the treatment since authors have examined their impacts individually.

Line 407, the choice of degrading which PHC molecules by bacteria might depend on the total energy source they need. I guess the assumption of "larger PHC molecules degraded at slower rates" is assuming the energy source (or C:N balance?) is not enough?

Line 418, what kind of "optimize the treatments"? adjust the temperature or moisture? or changing to a more efficient treatment?

Line 470-481, this paragraph of discussing cost and accessibility of some treatments is very valuable, but might be better to move up to section of line 445? In the Conclusion section, the statement should be more broad saying "Choice of the treatment should be considering multiple factors treatment effects, cost, time, accessibility by users, sustainable supplying, etc,  to fall into the concept of circular economy"?
Additional comments: This paper examined the bioremediation of engine-oil contaminated soils using residual fertilizing matter additions, and compared three types of matters (ramial chipped wood, horse manure and brewer spent grain) with a baseline fertilizer treatment. It measured multiple variables including VOCs, O2, CO2, PHC fractions, Microbial community level physiological profiling, etc. It reported that addition of horse manure had the best performance, but other types of residual fertilizing matters might also have the potential (e.g. smaller proportion of the dense and nutritious BSG with ramial chipped wood). I found this paper is very valuable to the society.