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: INTRASPECIFIC VARIATION IN THE COCHLEAE OF HARBOUR PORPOISES (PHOCOENA PHOCOENA) AND ITS IMPLICATIONS FOR COMPARATIVE STUDIES ACROSS ODONTOCETES
Review round: 2
Reviewer: 2

Output:
Basic reporting: Overall, I am very happy with the paper as it stands, and the authors made all of my suggested revisions. I don't really have much in the way of critical comments, other than some minor grammatical corrections, which I will list below.
Experimental design: again, all of my major criticisms were answered, and the inclusion of information on there samples as well as the criteria that was used to define different ecological classifications has greatly improved the manuscript.
Validity of the findings: The inclusion of the suggested information above does not change the overall findings, which remain strongly supported and which suggest that intraspecific variation in cochlea morphology is relatively minor compared to that between different species.
Additional comments: While the paper is well written, I did find a couple of minor typos and issues.

First, I am not sure what the difference is between a specimen from the UK versus the North Atlantic. Doesn't the UK border the North Atlantic? If the authors could clarify what this difference means it would greatly improve interpretation. I assume "North Atlantic" might just mean "we are not sure where it comes from other than general region", but again, that is an assumption on my part.

Under "Data Collection", "large availability of specimens" should be "large number of specimens"

The authors should overall double check formatting. While it might be a relic of the edited word document, I will note that between 159 and 160 there is a random change in line spacing, There is a change in font size at line 404, and it looks like some paragraphs may have extra spacing or not enough spacing between them.

Line 450: "It is also possible our data contain some cryptic species." This reads as a throwaway statement with no elaboration. While I would definitely buy this line of argument for say, members of Tursiops and Delphinus, I have never heard of any suggestions that North Atlantic populations of Phocoena phocoena may include cryptic species. There is some evidence that populations in different ocean basins, such as between Pacific and Atlantic, could be distinct, but these shouldn't influence your results. I would delete this sentence

Figures: I will note that the caption for the PC Plot for diving in the supplemental material still has diving depths randomly distributed, rather than going very deep, deep, etc (or vice versa). Make more sense to order them by depth, as the brain will probably initially want to read it that way.

These are all minor elements and should be quick to fix. I otherwise fully endorse this paper, and I look forward to reading the finished version


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

Input:
Title: ASSOCIATION OF WEANING PREPAREDNESS WITH EXTUBATION OUTCOME OF MECHANICALLY VENTILATED PATIENTS IN MEDICAL INTENSIVE CARE UNITS: A RETROSPECTIVE ANALYSIS
Review round: 1
Reviewer: 1

Output:
Basic reporting: For the most part, the article meets the standards.
The literature references use textbooks on some critical parts. I would recommend journal articles for these, even if only in addition, as textbooks are not peer reviewed and therefore not up-to-date per se.
Experimental design: The article falls within the aims and scope.
All technical and ethical standards were followed.
However, there is quite a difference in the patient characteristics in both groups that could lead to quite a big bias, and leading to wrong conclusions.
A more than tripling of the ventilation duration in the 'unsuccessful' group can lead to unacceptable bias. A better case-control or propensity-matching strategy should be (at least) considered.
Also the WEANS NOW criteria are not generally standardised. It would be better if the authors would elaborate on why the criteria were chosen 'as is', instead of just 'after a brainstorm session'. Especially as most of the criteria shown were not significantly attributing to the predictive value. Also, the verbal component of the GCS is a bit controversial for intubated patients.A bit more explanation on their standard would be advisable.
It is not stated whether or not more than one spontaneous breathing trials were (allowed to be) performed before actual weaning. It probably is, as the numbers in line 154 and 164 do not concur otherwise. This is an extra possible bit of bias.
Lastly, some more characteristics of the performance of the ICU would be appreciated. What is their current weaning failure? Then the conclusion would show whether or not the WEANSNOW could contribute to the performance.
Validity of the findings: See also 2.
All raw material is provided. The conclusions are quite a bit stronger than can be supported by the given material. The difference in patient characteristics is high between both groups. There are many reasons for higher failure to wean in longer ventiation duration, like ventilator associated pneumonia and muscle wasting to name only two. These can't be ignored by stating that the characteristics at the time of admission were comparable. Also the percentage of congestive heart failure was significantly higher in the unsuccessul group.
Moreover, even so the prediction cut off is 1, while the average of the successful group is about 0.54, and the unsuccessful is 1.18, a very small difference for a 8-point scale, and no patient is above 3 (and one of the 3 patients that scored three had a succesfull extubation).
Additional comments: I would think that with some elaboration on certain parts, it could be published. But the conclusions would probably quite be a bit less strong than it is now.


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

Input:
Title: ASSOCIATION OF WEANING PREPAREDNESS WITH EXTUBATION OUTCOME OF MECHANICALLY VENTILATED PATIENTS IN MEDICAL INTENSIVE CARE UNITS: A RETROSPECTIVE ANALYSIS
Review round: 1
Reviewer: 2

Output:
Basic reporting: 1.1.	Lines 48-49 reference an article over a decade old which describes different weaning processes. This seems like an arbitrary citation as this terminology is not commonly used internationally. Since differentiating between the three processes does not seem to be significant to the article, I suggest removing the sentence entirely.
1.2.	Several studies referenced are decades old and were not landmark studies in the field. It is important to build upon the current evidence base; updating references is highly recommended.
1.3.	Line 190: Respiratory therapists are mentioned here as the people who perform weaning assessments but it is unclear how that information is relevant. Is the WEANSNOW evaluation not performed by respiratory therapists?
1.3.1.	Are respiratory therapists commonly used in this geographical area? Having that bedside expertise (or lack of) may contribute to weaning readiness and success/failure. Clarification recommended.
1.4.	Table 3 and Table 4: RSBI > 105 is again referenced (see 3.1). RSBI should be less than 105 to align with the standard practice, unless the authors here are proposing something new.
Experimental design: 2.1.	It is unclear why participants were enrolled at a 2:1 ratio for success to failure. Was it only to get a larger sample size or some other reason?
Validity of the findings: 3.1.	Lines 169-172: An increased WEANSNOW score and RSBI > 105 should correlate with a decreased probability of extubation success. Is this a typo? RSBI < 105 is known to be the standard for assessing extubation readiness. How the article is currently written (suggesting RSBI > 105) would contradict a significant amount of published literature.
3.2.	Wakefulness seems to have substantial influence on patient performance. Should this be assessed independently or does research already exist? How valid is the WEANSNOW score if wakefulness is removed?
Additional comments: 4.1.	Lines 66-67 use the word evaluation twice in one sentence which may be confusing. Consider revision.
4.2.	Line 165: RSBI is listed as PSBI – needs typo correction
4.3.	Line 225: Use smaller internal diameter as it is more universal language.
4.4.	The focus on developing a weaning assessment bundle which accounts for airway condition, not just mechanics, is a worthy venture and is overdue.
4.5.	Several limitations are identified and thoroughly addressed.


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: ASSOCIATION OF WEANING PREPAREDNESS WITH EXTUBATION OUTCOME OF MECHANICALLY VENTILATED PATIENTS IN MEDICAL INTENSIVE CARE UNITS: A RETROSPECTIVE ANALYSIS
Review round: 2
Reviewer: 1

Output:
Basic reporting: Some typos. nothing major:
271-272: Typo, it should be “…WEANS NOW appear to be based…”.
295: Typo: experience should be experienced.

Perhaps some elucidation:
80: The Institutional Research Ethics Committee A of the National Taiwan University Hospital: is this correct? Is the Research Ethics Committee “A”? Or is it a typo? No need to change it per se if it is correct.

274-275: “…was also one of our original intentions…”. Intentions is probably not a correct word here, shouldn’t it be ‘reasons’ or ‘motivations’?
Experimental design: 171-173: what is the reason for mentioning the use of NIV? Is NIV to be viewed as an extubation-failure, or as a normal ‘step-down’ therapy or adjunct after MV? In my opinion it doesn’t really clarify anything, so I would just leave it out, and use a dichotomous extubation success or failure (unless WEANS NOW would predict usefulness of NIV after extubation, but I reckon this can’t be concluded from the current data). You talk about NIV as a step down in lines 283 and further, but the relation between the predictability of success of extubation using WEANS NOW with or without NIV is not researched here.

283-285: As stated before, the WEANS NOW wasn’t used with prediction of succes with or without NIV. I would rephrase, that it would be an interesting question whether or not patients with higher WEANS NOW scores can be succesfully extubated with NIV as a step down therapy.

287-288: I would elaborate on when a death or a transfer in a ventilator dependent state is viewed as still being a succesful extubation.You do this in lines 308-, but I think this should be done here, or it raises questions about almost 50% dying or needing ventilation after a successful extubation.
Validity of the findings: The conclusions reached are still a bit stronger than I would have reached myself. The use of WEANS NOW is not as strong a predictor as is suggested. The current conclusion is correct, but the discussion suggests something else:
263-264: “Otherwise, we strongly recommend…”

This seems like a bit of a strong recommendation, especially in the Discussion part of the article.

Also, I would address the differences in patient characteristics a little bit more in the Discussion. It is mentioned in the results, but it is not discussed as being a limitation (or not), or the possible effects on the interpretation.
Additional comments: The revisions so far have improved the article significantly. There is some difference in point of view with regards to the eventual conclusion. But with certain caveats it is defendable.
Most importantly I would like to see some discussion over the (possible extent or lack thereof of the) difference in patient characteristics.


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: INTERTHALAMIC ADHESION SIZE IN AGING DOGS WITH PRESUMPTIVE SPONTANEOUS BRAIN MICROHEMORRHAGES: A COMPARATIVE RETROSPECTIVE MRI STUDY OF DOGS WITH AND WITHOUT EVIDENCE OF CANINE COGNITIVE DYSFUNCTION
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript is well-written with clear professional English. The introduction and background sets the parallel between brain microhemorrhages in dogs and humans and states the potential association between brain microhemorrhages and dementia such as Alzheimer’s disease as well as pathologic mechanisms through cerebral amyloid angiopathy. While the association between microhemorrhages and disease states in humans in compelling and provides a rationale to undertake this study, the fact that the author do not have histopathology of the brain of the dogs included in this study is an important limitation which is acknowledged. Nevertheless, a lot of the introduction and discussion is based on the histopathological changes found in humans with microbleeds, cerebral amyloid angiopathy and/or Alzheimer disease which is not proven to happen in dogs with microbleeds based on this study as it lacks histopathology. I would recommend that the discussion is rephrased to discuss less the histopathological changes in humans and be more grounded into the results of this study.
Moreover, the authors emphasize that cerebral amyloid angiopathy may be a precursor stage to Alzheimer disease citing Ref 4. However, reference 4 is a metanalysis that showed no association between cerebral microbleeds and the risk of dementia. It should probably be emphasized more than the association between the two condition is controversial in humans.

The introduction also would benefit from being supported by more references. For example, the authors describe on line 69-73 that some geriatric dogs are presented with central vestibular dysfunction or recent onset of seizure activity, the vestibular disturbances being “much milder” and “tends to improve more rapidly” than dogs with the well described geriatric vestibular syndrome. However this is not supported by a reference and may only relate to the experience of the authors alone. Similarly, on lines 75-76, the authors claim that no metabolic cause has been identified in dogs with microbleeds although reference 17 showed an association between the presence of microbleeds and proteinuria while Lowrie et al., (vet radiol and ultrasound, 2012) observed that all dogs with putative microbleeds were hypertensive. Although this is not an identification of an absolute cause for the microbleeds these metabolic conditions should be discussed here.
Experimental design: The authors state the hypothesis at the end of the introduction. I would suggest that stating the hypothesis in the abstract would also help the reader to identify more clearly the goal of the study.
The hypothesis is: “that geriatric dogs with microhemorrhages without evidence of cognitive
dysfunction have a form of vascular -associated degenerative brain disease and that these dogs
have similar, abnormal, brain MRI features as dogs with cognitive dysfunction”. A second hypothesis is “More specifically, we hypothesized that interthalamic adhesion measurements in affected dogs would be smaller than that of aging geriatric control dogs (without evidence of encephalopathy or cognitive dysfunction).

	In order to answer the first hypothesis, meaning to prove that dogs with microhemorrhages have a form of vascular associated degenerative brain disease, this would definitely require histopathology. I would recommend that the hypothesis is changed and not include any histopathology related terms. Such as “vascular-associated degenerative”. Secondly, in order to compare the MRI features of between dogs with microhemorrhages and dogs without, it would require a control group which is strictly aged-matched. However, it is stated on line 135 that control dogs are significantly younger than dogs in the other two groups. On line 180 and 311 it is stated that the control are age-matched. This would need further clarification as if the control are age-matched they cannot be significantly younger at the same time. All the significant results found in this study could actually be the results of  affected dogs being older than control dogs as the interthalamic adhesion size as well as the presence of microbleeds have been associated with older age.

	Also the author took a novel approach to try and distinguish between dogs with and without CCD within all dogs that present with cerebral microbleeds, the control group is more arbitrary as it includes dogs with cerebral microbleeds as well but no evidence of encephalopathy. However the absence of clinical signs is solely based on the neurological examination which may not be able to detect changes in cognition in these dogs. It would be more objective to classify the dogs as having cerebral microbleeds or not and then within these dogs divide them into having such or such clinical signs. Especially if the dogs with cerebral microbleeds tend to experience quick clinical recovery of vestibular signs as stated by the author, is it possible that the control dogs with cerebral microbleeds have had some vestibular episodes that have now resolved and are just in a later phase when evaluated clinically?
For the inclusion criteria, some studies mention a size limit of  4-5.7mm of diameter to be classified as a microbleed. Has this been used in this study?
	Concerning the exclusion criteria, it is unclear in the methods if concurrent diseases that could cause vestibular signs or seizures have been ruled out in the affected dogs and if they have been ruled out, please explain using which tests.

The authors made some efforts to compare objectively their different groups. However some of the methods could be improved to make the study more reproducible and reduce bias:
On line 113, it is mentioned that two investigators have performed the measurement but it is unclear how this has been used in the study. Were the cases divided among the two? Has any inter-observer agreement been studied which would test the reproducibility of these results?
On line 120 it is mentioned that the number of microbleed was counted based on T2*GRE or SWI. However, there is a proven difference in sensibility between T2* GRE and SWI sequences may result in higher number of microhemorrhages in dogs evaluated by SWI (Guo et al., Clin Neuroradiol 2013). Can you comment on correlation between the two sequences? In order to make a rigorous investigation only one of those two sequences should be used in each dog.

Overall, the division in these 3 groups is not completely rigorous and based on subjective assessment (neurological examination which is insensitive to detect mild deficits in cognition or resolved clinical signs in diseases that improve spontaneously quickly). The way it is investigated without histopathology does not answer the primary hypothesis. A more rigorous analysis would probably involve looking at dogs with microbleeds only and compare dogs with and without CCD to investigate the relationship between these two conditions. Moreover, the control group being younger makes all of the results non-interpretable as they could be due to the age difference only.
Moreover, when looking at the raw data it seems that some dogs in the CCD group also presented for signs of central vestibular deficits and recent onset of seizures which makes the distinction between the three groups very unreliable and not clearly defined.
Validity of the findings: The lack of an appropriately age-matched control group is a major limitation preventing to reach a conclusion on the results observed.
On lines 161-164, the proportions of dogs with microhemorrhages in each group are presented which allows the construction of a contingency table for dogs with/without microhemorrhages and dogs with/without CCD which seems to show an association between presence of microhemorrages and presence of CCD. This finding would be worth mentioning and commenting.

In the Discussion section, the conclusions once again are hampered by the fact that the control group is younger and therefore all differences may be due to a difference in age between control and affected groups. It is also stated on lines 184-186 that: “dogs with microhemorrhage, with or without cognitive dysfunction, might share pathophysiologic mechanisms and might be manifestations along a spectrum of severity of a common disorder”. However, as this study lacks histopathology this conclusion cannot be reached based on the findings alone as it could be two separate diseases that happen concurrently in aged dogs.

The authors try to tackle the interesting question “what could be causing microhemorrages in dogs”. However this is not the stated purpose of this study, therefore the speculations provided in this section are not grounded in the results of this study and no results of this study suggest any potential cause. The reasoning used in the discussion, although interesting is based only on concurrent observation of the presence of hemorrhage on histopathology (and not MRI) and the presence of cerebral amyloid angiopathy in dogs which had no MRI evidence of microhemorrages similarly to this study which is therefore a weak link. In order to strengthen their argument, the authors draw an interesting parallel with the human conditions (line 206). It would be interesting to draw the parallel further by comparing which clinical data are different and which ones are similar between human and dogs with microbleeds.

The next question provided is “Could microhemorrhages in dogs be caused by cerebral amyloid angiopathy and could this, in turn, lead to cognitive dysfunction”. Similarly, with the lack of histopathology the answers to that question cannot be grounded in results from this study especially as no association between microhemorrhages and CCD is studied here (see contingency table above).
The discussion relies especially on histology features in humans which cannot be compared with the dogs of this study and this part of the discussion should therefore be removed.

One of the conclusion is that: “Our study suggests that the dogs with spontaneous brain microhemorrhages represent a distinct neurodegenerative brain disorder with some similarities to cognitive dysfunction, but other features more reminiscent of cerebral amyloid angiopathy.” This conclusion cannot be reached based on this study once again due to the lack of histopathology which would be necessary to state that this is a distinct neurodegenerative brain disorder with features of cerebral amyloid angiopathy. A disease may have many clinical signs and not all of them are always present in every case.
Similarly, the last sentence of the conclusion: “Our data support, but do not confirm the idea that spontaneous brain microhemorrhages in dogs might be a manifestation of amyloid angiopathy” cannot be supported by this study and should be removed in the absence of histopathology
Additional comments: Although this study describes a condition that has not been well described with only small case series reported, the construction of the three groups may artificially create the differences found as:
-	the control group is significantly younger and the diseases studied are strongly associated with older age
-	some dogs in the control group have microhemorrhages and could represent affected dogs that are not picked up on the neurological examination or affected dogs that have recovered clinically
-	some dogs in the CCD group present with similar clinical signs that the dogs with microhemorrhages and no CCD which blurs the distinction between these two groups.
-	Therefore, all the differences seen (or the absence of difference in between the two affected group) could be solely due to the group construction and may not have any biological relevance.

The lack of histopathology weakens this study as an important rationale used by the author is the link between microhemorrhage and cerebral amyloid angiopathy as is seen in humans.

On a side note, the consent form provided is not anonymous and contains private information from one of the participant.

In conclusion, this study does not provide a significant contribution to what is currently known about cerebral microhemorrhage as designed due to the important flaws in study design presented in this review. It may benefit from being reported as a case series of dogs with microhemorrhages. Some interesting information is present within this case series such as that the number of microhemorrhage (provided that it is counted only in T2*GRE or SWI for every dog and not one or the other) does not correlate with signs of CCD. Another interesting piece of information would be if location of the microbleed correlates with the presence of vestibular signs or recent onset of seizures as it is currently unknown if the microbleeds are actually the cause of these signs.


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: INTERTHALAMIC ADHESION SIZE IN AGING DOGS WITH PRESUMPTIVE SPONTANEOUS BRAIN MICROHEMORRHAGES: A COMPARATIVE RETROSPECTIVE MRI STUDY OF DOGS WITH AND WITHOUT EVIDENCE OF CANINE COGNITIVE DYSFUNCTION
Review round: 1
Reviewer: 2

Output:
Basic reporting: The paper is well written. Investigation of the similarities between canine and human ageing processes are of great importance and interest.
Nevertheless, the scientific or clinical merit of the data as they are presented in this manuscript are not clear.
Microhemorrhages / microbleeds were previously described in the veterinary literature. Risk factors, possible causes, related pathological conditions and outcome were also discussed in a well controled study (ref 17) pointing out the fact that these microhemorrhages are increased in number with age.
The use of interthalamic adhesion size as a marker for brain atrophy is well established as w ell (ref 19 and others) and was also  demonstrated to be negatively correlated to age  and significantly reduced in dogs with cognitive dysfunction.
Experimental design: In this manuscript, the authors collected data from dogs diagnosed with microhemorrhages with and without clinical signs of cognitive dysfunction and compared them to control dogs. The inclusion criteria used to allocate each dog to its study group is not always clear.

      Moreover, comparing the three groups using same parameters that were used as inclusion criteria (eg -presence of microhemorrhages, brain atrophy) is not recommended.
The diagnosis of CCD is usually based on clinical signs and supported by typical MRI findings such as brain atrophy. I assume that the control group of dogs were dogs with normal cognitive function and normal MRI, therefore the idea of comparing the size of interthalamic adhesion or the presence or lack of microhemorrhages between these two group of dogs is not proper.

Material and methods:
1.	Inclusion criteria are not clear nor is the “search database from three institutions". – please clarify, how were dogs recruited to each of the study group. Please include the inclusion and exclusion criteria. (It seems that the inclusion / exclusion criteria of some of the groups  were then used to compare the three groups (eg. Intra-thalamic adhesion area and size and the presence of micro-bleeds)

2.	Line 95 -How were the cognitive dysfunction diagnosed?  please specify.

3.	Please provide reference of the definition of geriatric as dogs above 8 years?

4.	The differences between group 2 and 3 is not clear. Group 2 are dogs with microhemorrhages and no clinical signs. And group 3 should be no clinical abnormalities and no microhemorrhages on MRI? Please clarify were the MRI looked at before allocating dogs to the group? Was that a double blind study?

5.	Control: no age, weight- and breed-matched control were available.
6.	No consideration of breed and dog size is used for the statistical analysis although these were found to be significant risk factor in one of the references quoted (Kerwin SC et al 2017, ref 17)
Validity of the findings: 1.	The control group is not suitable if they are significantly younger than the affected dogs since we are looking at age related processes.

2.	Breed of dogs are given in each group but there were not matched even though small breed were found to have significantly higher chances to suffer microhemorrhages than larger breed of dogs.

3.    Figures 5, 6,7 & 8 present the same findings. Two figures should be enough. One that correlate height of interthalamic adhesion to its area and the other figure that uses either one of these parameters with or without normalisation (using the total brain volume) to correlate with number of microhemorrhages found.

4. Figure 9: Again, if control dogs were included based on normal MRI findings it is only expected that they will have significantly less microhemorrhages than both groups of affected dogs.
Additional comments: •	Not clear what the objective is. What is the importance of comparing these three groups of dogs?  They state that the objective is to compare these three group of dogs but the logic of doing this is not obvious and not provided.
•	The definition of geriatric dogs was previously discussed (Fortney WD. Implementing a successful senior/geriatrichealth care program for veterinarians, veterinary technicians, andoffice managers. Vet Clin North Am Small Anim Pract 2012; 42:823–834 ) were size of dog is taken under consideration when determining “geriatric” in dogs. Ageing dogs maybe a better term for 8 year and older rather than geriatric.


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: EVOLUTION OF SWITCHABLE APOSEMATISM: INSIGHTS FROM INDIVIDUAL-BASED SIMULATIONS
Review round: 1
Reviewer: 1

Output:
Basic reporting: see general comments
Experimental design: see general comments
Validity of the findings: see general comments
Additional comments: In this paper, Song et al. performed an exhaustive simulation analysis to dissect the conditions under which 'switchable aposematism' can be favoured. Switchable aposematism corresponds to an intriguing anti-predatory strategy where defended prey deter predators by suddenly switching warning signals that are normally hidden. While the benefit of switching conspicuous display is well understood in *undefended* prey (a sudden switch can surprise predators, allowing the prey to escape; a process called 'startling'), we lack a clear understanding of the underlying evolutionary forces favouring the evolution of such switching conspicuous display in *defended* prey. The evolution of aposematism and cryptism are rather complex by themselves, and switchable aposematism corresponds to a mixed anti-predatory strategy. Switchable aposematism can bring about multiple benefits that are not mutually exclusive ('startling', facilitation of aversive learning, reduction of the costs associated with signal display), and multiple costs (higher detectability, lower learning speed, intrinsic costs associated with switching). As such, I agree wholeheartedly with the authors that a theoretical investigation of the underlying evolutionary forces favouring 'switchable aposematism' is required.

In this study, the authors choose to constrain their sensitivity analysis to specific parameters linked to the costs associated with signaling or switching (the action of switching and the switching device alike), and highlight the conditions under which cryptism, constant aposematism or different kinds of switchable aposematism (pre- or post-attack switchable aposematism) can be favoured. I believe that the model is making important predictions. For instance, it shows that all sort of switchable aposematism cannot be favoured unless the costs associated with signaling or switching are explicitly considered. Intuitively, one might expect that accounting for one or the other cost is enough to explain the benefit of any kind of switchable aposematism. More importantly, they provide some theoretical evidence that the evolutionary transitions from cryptism to switchable aposematism or from constant aposematism to switchable aposematism are not equally likely, and depend on the intrinsic costs and on the nature of the predator community.

Overall, I think that the theoretical model is sound, and that the authors made a good job to tackle a rather complex question.

Nonetheless, I list below some important comments that the authors should tackle to make their case even stronger, and to make the paper easier to read.

1) I found the paper rather difficult to read, and I was a lot less enthusiastic at first read. The main issue is that the authors do not highlight their interesting results on switchable aposematism. They choose to emphasize the usefulness of their modeling software ('ApoSim'). I would strongly advice the authors to highlight their results instead. First, I am sure that theoretician and empiricists would be delighted to learn about the findings shown in this paper; this would make an overall more appealing paper. Second, I am afraid that very few researchers would use this simulation code because it is tackling a rather specific antipredatory strategy. If all theoretician would name their model and would ask other researchers to use it because it is "user friendly", I fear that it would not facilitate the reading of the theoretical literature. Third, all theoretician may not find such model with a graphic interface so 'user friendly'. Just to be clear, I am not stating that the model is useless. On the contrary, I hope that the authors of the paper (or other researchers) will use this same theoretical framework to tackle other questions related to switchable aposematism (e.g., deimatism and learning facilitation)! I think that the authors would emphasize the potential of the theoretical framework by being clear about the important and unexpected results they got.
To tackle this comment, the authors should consider to re-write almost all the manuscript to emphasize their results.

2) I appreciate that the simulation run time may have been limiting in such individual-based model. Yet, the authors may consider performing few additional analyses (as Supplementary Information) to test the robustness of the results to other parameters. In particular, I would be curious to see if the results hold for different values of intensity associated with signals 'L' and 'H'. Likewise, testing whether the results hold to variations (i.e., same figure as Figure 3, but with two different values around the default combination of parameter) of generation length, prey frequency, predator frequency... etc... would allow the reader to be more confident regarding the robustness of the main conclusions.

3) The authors should really think about removing the 3D representation. By distorting scales, 3D plots are very hard to read. In subfigures 3A, 3B and 3C, each subplot (for specific predator turnover values) could be represented as a separated 2D scatter plot.

4) Terminology "Cost"; "cost-reducing". In the introduction, the authors should be more specific and consistent concerning the terminology associated with costs. For instance, in line 122, I was very confused at first read because this is the first time the authors discuss the intrinsic costs associated with switchable aposematism, while the authors only discussed about 'cost-reduction' before. Given that these costs are the most important components of the model, the authors should be more explicit about all those costs and how this relates to the benefit/costs associated with switchable aposematism (for instance, signal cost brings about an advantage to switchable aposematism over constant aposematism but not over cryptism). I believe that introducing the concepts required to understand the results (instead emphasizing the novelty of the method), the introduction would be more easy to read, and less confusing on this aspect.

5) In the Material and Methods, the authors should state that all prey individuals are mimetic (i.e., there is one type of signal, with more or less intensity)

6) In the Material and Methods, I was confused about which condition each signal corresponds to (resting, approached, attacked). In particular, for the last 'prey react', it is not clear whether the prey re-adopt the resting signal. This should be clarified throughout the methods.

7) In the 'Overview of the model structure', it should be stated that the predator is tasting the prey upon attack ant that it may affect subsequent attacks.

8) In the Material and Methods (including in the 'Overview'), the initial prey community should be described (I guess it is a mix of all strategies)

9) In the Material and Methods, I did not understand exactly when the instinctive repulsiveness is reset to 0. At each interaction frame? Note also that this terminology is confusing because it seems to refer to innate avoidance, which is not modeled (as said line 172)

10) I did not understand why a prey should do anything after the last attack within an interaction frame (step 3). Why switching signal (and paying a cost) if the predator is going to leave? I guess I missed something in term of process order.

11) It is very odd to refer that often to Figures in the Discussion. In particular, it makes the reading very unwieldy.

Other minor comments:

The authors should care about the use of "the". I found many sentences where it is misused (eg. lines 51, 52, 68, 69, 71, 75)
lines 55-56: This sentence is odd. How one can verify a theory? This sentence would be clearer if the authors were stating their results (cf my first point)
line 73: "one aspect" should be defined in this sentence
line 94: "startling" should be defined (as in lines 100-101)
line 96: "As introduced above": it has been done in the abstract. It may be worth to be more explicit
line 148: citations should be outside quotation marks
throughout the paper, it would help if the citations were ordered chronologically
Figure 3: the authors could consider adding a legend with the predatory strategies (even if it the same legend than in Figure 2)
line 228-230: this is very speculative. I would remove this sentence
Equation 1: What is the current survival chance initially? One?
Equation 9: Same for the initial current motivation? Zero?
line 325-329: This should be discussed further in the Discussion. It is an important assumption that leads to an overestimation of the parameter space with constant or switchable aposematism.
lines 426-427: This should be moved in the section 'Prey-pay-cost' below. Same lines 481-482
line 434: "after predator approach" is confusing because an attack has already occurred. "approach" should be clarified throughout the manuscript.
lines 580-584 and lines 585-588: these are repetitive paragraphs; this could be shortened


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: EVOLUTION OF SWITCHABLE APOSEMATISM: INSIGHTS FROM INDIVIDUAL-BASED SIMULATIONS
Review round: 1
Reviewer: 2

Output:
Basic reporting: Please see 'General comments for the author' below.
Experimental design: Please see 'General comments for the author' below.
Validity of the findings: Please see 'General comments for the author' below.
Additional comments: The authors develop a model called ApSim to study the different contexts in which the benefit of switchable aposematism can outweigh its cost and therefore become a winning evolutionary strategy. The model considers a large number of parameters that can constraint the output of prey-predator interaction, including predator turnover and learning speed, prey basal detectability and cost of switching on the anti-predator signal.

The modeling proposed by the authors is highly original, allows the full complexity of the interaction between several aspects of prey-predator interaction to be taken into account, and, importantly, does so by circumventing the hard question of measuring empirically the cost of all the behavioral traits involved in the interactions between prey and predator. The last point allows the authors to make several (qualitative) predictions regarding the situations in which switchable aposematism -and other alternative strategies- may be advantageous.

While I am totally convinced that the manuscript is worth publishing, for the reasons given above and below, I do think it could be greatly improved by being partly rewritten. Three types of readers may be interested by the study: 1. readers who will skim over the abstract, the introduction and jump to the conclusion to get the take-home message (in particular, something in the lines of “switchable aposematism may be advantageous when predator learning speed is high enough and the cost of switching low enough”); 2. readers who, in addition, will try to get an idea of the modeling used to show these theoretical results; 3. readers willing to, in addition, reproduce the results and explore related questions using ApSim. The authors give a very detailed description of the model and of the rationale for most of their choice in the modeling. However, the writing of the manuscript, as it is, would make the reading of categories 1 and 2 very hard, and the improvements I propose below would, I think, also benefit readers in the three categories, as well as boost citations.

The first improvement I would suggest is to start each of the very precise, long descriptions of the model setting (there are a lot given the complexity of the model) with one, or some, summary sentence(s) providing the meaning of what follows in plain, non-technical language. For doing so, a possible way is to first make the main claim of the paragraph and next back the claim up with detailed descriptions, rationale or references.

Studies based on computer simulation to understand anti-predator coloration are becoming more and more frequent. They are based on online experiments in which a great number of (human) observers play the role of predators and “forage” to determine the relative fitness of prey. This study is totally different. There is no “perceptual interaction” between prey and predators. Prey-predator interactions are determined computationally by the settings of the model. This should be spelled out very early in the text. I found it very hard to understand what the interaction between prey and predator was before the lines around line 270! I think, and that’s perhaps my main concern about the writing, that a clear, plain description of what ApoSim does in terms of prey and predator interaction should be provided very early on in the introduction. An ideal location would be on line 113, after the first sentence of the paragraph (“We developed ApoSim … this subject”). This added explanation would state that prey and predator interact according to a modeled predation that depends on several parameters etc., which is the core of what ApoSim does.

By reading the conclusion and the abstract only, one would get the feeling that the manuscript is solely about the presentation of a model, which is not true. The work realized offers much more: clear predictions about when switching aposematism should be expected according to the cost of different aspects of the problem. The main predictions of the modeling should spelled out in the abstract of the paper and in the conclusions to make sure readers in categories 1 and 2 (and perhaps 3) above will read the paper. It is clear that the modeling is purely theoretical in the sense that the values for the cost considered are abstract. However, the manuscript does a good job in explaining that these costs are very hard to measure empirically, so the theoretical conclusions of the modeling are a new contribution to the field and should be emphasized as much as the fact that a new model has been proposed, in both the abstract and the conclusion.

Other points:

-Lines 224-230: The short paragraph states first that coevolution is not considered, but goes on saying that the modeling can enhance our understanding of coevolution. I don’t understand how both statements can be true. Please clarify.

-Eq. 4, line 341: I find the negative interaction between signal intensity and basal detectability in the product “signal intensity * (1-basal detectability)” hard to understand. Is that arbitrary? I understand this formula guarantees that discovery chance will be between 0 and 1, but other formulae are possible such as, simply, “discovery chance = signal intensity * basal detectability”, and are, to my understanding, more intuitive. Could you clarify this point?

-Eq. 5, line 353: same as for Eq. 4. Why not, simply, “conditional stimulus = basal cognitive cue * signal intensity”? I don’t understand the negative interaction in the formula.

-A general remark saying that the model is fully qualitative is missing. Other choices for the parameters could lead to a totally different picture in Figures 2 and 3.

-Lines 531-533: “We asked … into the system”. This question is very important and really helps the reader to understand the study. Why not highlighting it earlier, in the introduction?

-Lines 616 and following lines of the Discussion: please explain what you mean by evolutionary transitions and how this idea relates to the modeling. It is not clear from the manuscript.

-Figure 3 really does a good job at summarizing the whole complexity of the study in a single visual representation! This figure should be displayed in the published manuscript on a full page. It would therefore be useful to show the color code and the short-three letters notation given in figure 2 in figure 3 as well.

-Line 731: “would be almost random” -> “wouldn’t favor any strategy”?

Minor point:

-Abstract, line 50: “it” is not precise enough and may be replaced by “the model” (“To illustrate how the model can be used”).
-Abstract, line 56: same remark for “its”.
-Line 130: “these mechanisms” instead of “those mechanisms”?
-Line 134: comma missing after Srygley 2004).
-Line 147: “In the literature the switchable aposematism…”
-Line 162: “cost-saving aspect of the switchable aposematism”.


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: EVOLUTION OF SWITCHABLE APOSEMATISM: INSIGHTS FROM INDIVIDUAL-BASED SIMULATIONS
Review round: 2
Reviewer: 1

Output:
Basic reporting: See General comments
Experimental design: See General comments
Validity of the findings: See General comments
Additional comments: The authors did a very good job in revising their manuscript that is now way easier to read.

The authors addressed my earlier concerns. I now only have few minor comments and editorial suggestions (mostly on the presentation of the Methods) that the authors may want to follow.

1) In the Introduction, I got a bit confused by what the authors refer to as 'signal penalty' (line 134). Intuitively I thought about costs associated with the signal even when it is hidden. I had to read until line 364 before understanding that the authors actually refer to signaling costs. I would suggest the authors (1) to be more explicit at line 134, and (2) to use the term 'signaling penalty' or 'signaling penalty' to avoid any confusion.

2) I find that the terminology about prey behavior is a bit confusing. In particular, the authors could use another expression to refer to the "post-attack behavior"; the sentence "the final attack envent occurred after the post-attack behavior" (line 247) sounds really weird. I understood that the "post-attack behavior" occurs after the initial attack, and before the final attack. Yet, I am afraid that other readers could be confused by this terminology. I would suggest either to change the terminology, or to draw a flowchart showing how the chronology of prey behavior and attack events.

3) In the Methods, I would suggest to explicitly state how the prey behavior is changing (e.g., from the resting state to the approached state). This would considerably help to understand the order of the processes in the main text

4) line 445-446 "it probabilistically decided to attack it or not", lines 453-454 "was the probability of deciding to attack the focal prey in the next time step", and line 459 "each predator conducted an action"; here the process order is not clear at all, and this is rather hard to understand when the attack take place. I guess that it does not really matter during the simulation; once you have update the motivation variable, you can update the survival chance variable at any time. Yet, it would be clearer for the reader to explicitly state that the predator attack (or the retreat) takes place at some point.
Note that the use of the past tense in the Methods does not ease the reading and the understanding of the process order because it make it hard to refer to past/future events. Personally, I would use the present tense but this is a style difference.

Editorial suggestions

line 56: 'but' is misused given because the second part of the sentence does not conflict with the first part of the sentence
line 72: 'to accept'; should the authors use the word 'ingest' instead?
line 80: the authors can remove 'will'
line 100: To ease the reading, this sentence could be rephrased as "Recently, however, researchers experimentally explored the switchable visual displays of some chemically defended prey"
lines 113-114: The expression 'might also accelerate the predator nervous system's learning about the aposematic prey' sounds weird. This could be rephrased to ease the reading.
lines 112: I am not sure whether the authors start a new paragraph here. If not, this could be done to contrast costs associated with signaling and costs associated with switching.
line 162: I would suggest to add 'Notably'; i.e., "Notably, the interactions...."
line 170: 'the general cost of signaling'; do the authors refer to the cost of switchable signaling here, or to the cost of signaling in general?
lines 178-189: Here, the authors could cite the papers that suggested those existing hypotheses.
line 212 and elsewhere: I would avoid referring to 'silent' signals. This can be confusing; it sounds like the authors are describing sound signals.
lines 323-324: "the prey would assume the approached state instead"; This sounds weird. I guess that there is a typo in this sentence.
lines 422-423: "Please note that..."; I did not get the meaning of this sentence. Could the authors rephrase it?
line 475: The authors could write down explicitly the increment equation for *unconditional stimulus*. While reading, I used to look at the following equation to understand what is the use of variable in the subsequent time steps. I guess that writing explicitly the increment equations could really help other readers to understand the Methods.
line 742: To make their point about aposematism based on cues, the authors could refer to the recent opinion paper by de Solan & Aubier (2019 Front.Ecol.Evol. 7:283. doi:10.3389/fevo.2019.00283)
line 905: I would avoid referring to "intelligent predators", the notion of intelligence being rather difficult to define in animal behaviour.


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: EVOLUTION OF SWITCHABLE APOSEMATISM: INSIGHTS FROM INDIVIDUAL-BASED SIMULATIONS
Review round: 2
Reviewer: 2

Output:
Basic reporting: Please see "General comments for the author".
Experimental design: Please see "General comments for the author".
Validity of the findings: Please see "General comments for the author".
Additional comments: Dear Authors,

I am really pleased with the new version of the manuscript. The writing now reflects well the strength of the methods and the width of the results of the study. In particular, it is now clear that the paper provides not only a novel modeling software, but also, and arguably above all, insights on possible and testable routes for the evolution of switchable aposematism. The new abstract is clear and well written, and provides a clear description of the main question and the main results. It will make readers of the three categories of reader discussed in the first round of review willing to read the paper. In its new form, the introduction is also much easier to follow and well-structured. The methods have gained in both concision and clarity. Finally, the conclusion is very clear and provides a much broader message than the one in the former version of the manuscript, much closer to what the study deserves.

Overall, I think that the main problem of the paper in the first submitted version, namely that the writing didn’t reflect the wealth of theoretical results, which made the paper less appealing, has now been solved and well solved. I am also happy with the answers to my theoretical questions about the interaction between some of the variables (additivity, multiplicativity), and the way the corresponding information has been included in the manuscript. I have found a few instances in which the English language should be improved, in particular the use of “the”. I am not a native speaker, but I think it is worth looking at the points I have listed below to improve further the understandability of the manuscript. Taken together, I consider that the manuscript is sound, novel and interesting for different types of readers, and therefore worth publishing once some minor language corrections will have been carried out.

Best regards,
Olivier Penacchio-----

Minor comments on language:

-L. 49: The switchable aposematism has been relatively -> Switchable  aposematism.

-L. 51: Second, it could facilitate the aversive learning -> facilitate aversive learning.
Third, it could minimize the exposure or -> minimize exposure.

-L. 55: 88 128  -> no space, 88128.

-L. 63-64: explore the evolution of the switchable aposematism -> of switchable aposematism.

-In general: “the switchable aposematism”-> “switchable aposematism”.

-L. 83: We will call this form of aposematism as the “switchable aposematism.” -> remove “as the“.

-L. 84: “switchable aposematism”, without “the”.

-L. 84-85: the commas show come after the references in the whole sentence (e.g., with various adjectives such as “facultative” (Sivinski 1981; Grober 1988), “post-attack” (Umbers & Mappes 2015; Kang et al. 2016) “early-acting,” ),…”).

-L. 195: e. g.  -> no space.

-L. 268: hypotheses regarding evolution of the switchable aposematism…  -> hypotheses regarding the evolution of switchable aposematism…

-L. 270: formulated in -> remove “in”.

-L. 337: It should be noted that the signal intensity is a  -> remove “the”

-L. 464: The detailed simulation structure -> Detailed simulation structure

-L. 544: basal detectability and signal intensity has additive relationship, not multiplicative -> basal detectability and signal intensity have an additive, not a multiplicative, relationship

-L. 545-546: “The rationale behind this design is to prevent one side being zero from negating the other; each of them should be able to invoke predator discovery even if the other one is very small.”   Should be rephrased for clarity. Maybe something like: “The rationale for this design is that both basal detectability and signal intensity should be able to contribute to predator discovery even if the other variable is close to zero or zero.”

-L. 557: as the Eq. 4 -> “as in Eq. 4” is probably better.

-L. 701: Learning speed…:, without “the”

-L. 736-738: Consider inverting the order of the sentence for clarity.

-L. 839: May better: “Second, we assumed that increasing threat level can only be associated to increasing levels of signal intensity”

-L. 842: 88 128 -> no space.

-L. 857 and other instance below: e. g.  -> no space.

-L. 919: Maybe better: “The results of the model (Figure 3) help our understanding of possible evolutionary transitions…”

-L. 955: a higher chance.

-L. 1217: a more detailed discussion: the prey defense. -> no “the”. Same on line 1218.

-L. 1267-1268: may lead to disappearance of post-attack aposematism -> I think “may lead to the disappearance” works better.


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: VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES
Review round: 1
Reviewer: 1

Output:
Basic reporting: In my opinion, the research work can be accepted, but some issues need to be taken into account.
The work really, although interesting, does not contribute much. It is a further investigation on the use of portable meters to detect BOHB values and there are multiple publications on this aspect. Some of them comparing different meters, others comparing with more complex (and more expensive) laboratory techniques, others made in different animal species such as cattle, goats and sheep, and even comparing 2 and 3 portable devices (among them) existing in the market, to check its sensitivity and accuracy.

Really the only novelty is the method of induction of ketonemia, even causing severe ketonemias. In most of the publications, animals that present spontaneous disease are sampled, or induce ketonemia (or pregnancy toxemia) by combining food restriction, fasting and advanced gestation.
It is true that most publications have fewer samples, but in my opinion it is justified by the previous reason. And when cattle are used, the number of samples is usually even much wider. In my opinion, the sensitivity of the portable meter is indifferent that it is made for one species than for another.
It seems that there is interest in not mentioning the name of the meter tested (Optium Xceed®). I think it should be indicated at least in the Abstract, in the Discussion and in the Conclusions.
Experimental design: ..
Validity of the findings: ..
Additional comments: Specific comments

Abstract
Include the name of the hand meter tested (Optium Xceed®) in the Abstract.

Introduction
Βeta-hydroxybutyrate acid has not been correlated with the abbreviation BHB the first time it is used in the text (line 58). Yes it is done in the Abstrac, but I would recommend doing it in both places
Unify abbreviations and always use them. HM (as handheld meter) is used, but HH and HHM are also used in figure 1. SG (gold standard) and enzymatic colorimetic method (line 362, title Table 1, and within the same table 1). mM / L and mmol / L: unify
Remove capital letters that are not strictly necessary (Less, line 80; Handheld, line 362; Enzimatic line 362, etc)

Materials and Methods
How was the BHB cut point set? In addition to those indicated, I would recommend revising the reference Cal et al, 2015. In this publication, which is also taken as a reference for the diagnosis of subclinical pregnancy toxemia, the value of 2.26 mmol / l is set.
Clinical and Subclinical are continually referred to, and become synonymous with Severe and Moderate ketosis. It is an important mistake. The clinical and subclinical concept refers to the presence of visible or easily detectable clinical signs, not the more or less high BHB value. In Material and Methods there is no reference to this classification in clinical and subclinical.
Neither in Material and Methods is it indicated if other clinical signs are collected, in addition to the rectal temperature. Therefore it is difficult to classify the disease the clinical and subclinical.
Even on line 171 it is indicated: “… none presented any clinical sign of PT”. How can we talk about sheep with clinical signs in the rest of the manuscript, if it has been said that none presented clinical signs? Check.
In Material and Methods it is not indicated that ketoneuria is measured, but then it is indicated “… an intense ketonuria after 20 minutes” (line 172). Has ketoneuria really been measured? Check.
Line 145: “… containing sodium fluoride anticoagulant (Vacutainer®, ..)”. Sodium fluoride is not an anticoagulant, but in these vacuum tubes it is usually combined with potassium oxalate or disodium EDTA, which do act as anticoagulants. Sodium fluoride stabilizes glucose levels, preventing glycolysis.
Line 167: "... with GraphPad Prism software (GraphPad Inc., La Jola, CA, USA) ..." change to "... with GraphPad Prism software (GraphPad Inc., La Jolla, CA, USA)"
I would recommend making a specific section, or at least an individual paragraph, about the statistical study. How has specificity and sensitivity been assessed?

Figures and Tables
Review all titles in Tables and Figures. Take into account the Clinical and Subclinical indications, and possibly change for moderate and servere ketonemia, and Handheld meter (HM) and gold-standard enzymatic colorimetic test (GS).
I do not understand the titles in Table 3. Subclinical ketosis should be changed to BHB ≥ 0.8 mmol / l and BHB ≥ 1.6 mmol / l. Perhaps I would recommend removing the standard gold line and indicating it in the title of the figure.
Figure 3. Bellow is incorrect. Replace with below. (Lines 395 and 400)

Discussion
In general the discussion is correct, but I would recommend reviewing the references below and including them in the discussion.
Line 243 and following: The rectal temperature is increased, probably by a small quantity of pyrogen in the solution ... In my opinion the stress of manipulation, sampling and immobilization, may be sufficient to induce a slight elevation of the body temperature. And that would justify the slight hyperthermia more than the possible presence of pyrogens.

References

I would recommend reviewing the following references, and perhaps including them in the discussion of this manuscript

Cal-Pereyra L, Benech A, González-Montaña JR, Acosta-Dibarrat J, Da Silva S, Martín A. 2015. Changes in the metabolic profile of pregnant ewes to an acute feed restriction in late gestation. New Zealand Veterinary Journal, 63:3, 141-146;
Carrier J, Stewart S, Godden S, Fetrow J, Rapnicki P. Evaluation and use of three cowside tests for detection of subclinical ketosis in early postpartum cows. J DairySci 2004;87(11):3725-3735.
Firat A, Ozpinar A. 2002. Metabolic profile of pre-pregnancy, pregnancy and early lactation in multiple lambing Sakiz ewes 1. Changes in plasma glucose, 3-hydroxybutyrate and cortisol levels. Annals of Nutrition and Metabolism 46, 57–61.
Kupczynski R, Cupok A. Sensitivity and specificity of various tests determining beta-hydroxybutyrate acid in diagnosis of ketosis in cows. Electronic Journal of Polish Agricultural Universities Series Veterinary Medicine 2007;3(10):1-15.
Mair, B., Drillich, M., Klein-Jöbstl, D., Kanz, P., Borchardt, S., Meyer, L., ... & Iwersen, M. (2016). Glucose concentration in capillary blood of dairy cows obtained by a minimally invasive lancet technique and determined with three different hand-held devices. BMC veterinary research, 12(1), 34.
Nielen M, Aarts MG, Jonkers AG, Wensing T, Schukken YH. Evaluation of two cowside tests for the detection of subclinical ketosis in dairy cows. Canadian Vet J 1994;35(4):229.


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: VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES
Review round: 1
Reviewer: 2

Output:
Basic reporting: This research paper presents the test of a handheld “ewe-side” ketometer device. There are already published relevant data concerning the same measurements in ruminants, but this research offers an additional approach to a known subject. The innovative approach of the paper is that the ewes were administered beta-hydroxybutyrate acid (BHB) intravenously and the artificial hyperketonemia enables the researchers to test for high blood BHB values, not easily found in clinical practice and natural occurring cases. Study design is good and the idea of BHB infusion and measurements over time is interesting. Study was performed well, but the results of the study are not presented well, on the contrary, the results are not clear and reader cannot actually understand the paper. Significant corrections and changes are suggested in the presentation and the selection of data that are the results of the study. In the form that are presented now, results and data do not comply with the study design, which is promising and well performed, and cannot justify publication of this work.

English language is adequately used, although some mistakes could be corrected L54, L57, L86, L103, L127,…
Literature is adequate in selection, use and presentation and format.
Tables are not presetned well, are confusing and do not help the presentation of results.
Data selection and presetnation is not adequate.
Detailed comments:
Abstract: Wrong numbers of ewes and samples are written (comment below).
L81, though the most common is US dollar, it would be better to define it in the text as US dollar (USD)
L101, it is mentioned that 19 multiparous ewes, but in abstract (L27-28) it is claimed that 24 ewes were used which is not correct. In the same way the total samples were 13X19=247 (L152-153 and as seen in Table 3) and not 312 (13X24) as claimed in L29

L128. Physical evaluation could be described, as it is important either for the welfare of animals and for the interesting clinical point of artificial hyperketonemia induction. For example the ketonuria mentioned later on, how was measured? Were there changes in heart/respiratory rate? Preliminary nervous signs? Any clinical changes in a ewe with BHB >3 mM/L would be interesting to be mentioned. It is also useful to mention the climate condition, especially temperature during the sampling time.
L139-L143 This part does not offer anything significant, it can be omitted

L362 replace Table 1 with Table 2
The results as described in test and presented in Tables are not comprehensive and actually confusing.
L174-177 these results are not presented in table 1, difficult to understand where are obtained from. For example: The overall mean value of BHB concentration using the GS was 1.85±1.45 mM/L, (according to Table 1 is 1.90±1.56!!) while the HM presented a higher mean of 3.10 ± 2.90 mM/L (P <0.05) (according to table 1 is the opposite, HM is lower in mean 1.25 ± 0.89 mM/L!). The BHB concentration of the GS ranged from 0.10 to 5.65 mM/L (according to table 1 ranged from 0.10 to 7.10 mM/L!), while the results of the HM ranged from 0.10 to 10.90 mM/L. (according to table 1 ranged from 0.1050 to 3.9360 mM/L!)
In other words, according to Table 1, GS method obtains significant higher values, which is in contrast to the results presented in Table 2. Moreover, in this part of the text results of both tables (1,2) are presented, and in making confusion bigger table 2 is named table 1 in L 362

L182-184 and Table2
In Table 2, authors allocate the mean values obtained according to BHB range in 3 categories (0-79 mΜ/L)/ 0.8-1.6 mΜ/L and >1.6 mΜ/L and then present the mean values obtained from HM and GS measurements. The results present the big difference between the mean values obtained from the 2 different methods of BHB measurement. But, in case the difference of mean values is so wide e.g.1.82 vs 1.27 for the category (0.8-1.6 mΜ/L) is questionable how is possible to have the same number N=61 of values in both measurement methods. If Handmeter (HM) gives constantly higher results, than GS, it is curious to have the same number N=61 of measurements for both method in the same wide range (0.8-1.6 mΜ/L) category. It could be assumed that due to wider range of valued obtained from HM method, the numbers of results in each category would differ between the methods. Anyway, this Table (table 20 is more confusing than helping for the reader, only the overall –line results make sense.
It would be more informative to compare in the same sample the different values obtained from HM and GS. Also it would be more adequate to compare mean values of each sampling time T1-T13 and present the differences in values measured with HM and GS method. It would be also more interesting from clinical point of view, as to see how infusion and excretion of BHB in blood changes. Authors designed a trails with BHB IV infusion and repeated measurements on certain times frames and do not present these results! On the contrary they mix mean results of both methods (table 2) offering confusion to the reader.

Again, in table 3 the way columns and line are allocated is impossible for the reader to understand which are positive and negative by which method, so as to understand false and true positive ones.

L205-207: These results are not presented and are actually a basic finding according to the design of the study.
L208, L172: Ketonuria is mentioned here, but it was not mentioned in the material and methods. Urine BHB was measured to all animals in each sampling time? It could be added in the relevant section
L210-211: These results are not presented anywhere in the study
L233: sensibility?
L241-243 and L247-249. As mentioned above these results are not presented anywhere in the paper and are actually important and one of the innovation of this study, but authors simply omitted form results and add suddenly this information in the discussion part.

Paper is self-contained.
Experimental design: Experimental design is adequate for the well defiend reseasrch quastion. Presentation of the design in the material and methods section needs slight improovement, but in general are described well.
Statistical analysis is adequate and accurate.
Adequate and relevant to the aims of the study, methods followed, technical and ethical standards are fulfiiled.
Validity of the findings: The major problem of the submitted paper is the selection of data that authors presented and the way these data are presented.
Desing of the study enables repated measurement before during and after intravenous infusion of BHB in ewes. According to the study desing, research could harvest valuable measurements of variable blood BhB concentrations including high and very high values and their change in certain time frames. Unfortunately authors do not present these results, that could be the nevel thing of thiw research. On the contrary present insuffuciently a mixture of mean values, with no time reference in a very confusing and disrupted way.
This makes thiw work incomprehensive and actually hinters the soundness of the findings/
Under this point of view discussion looks poor and also the conclusions.
Additional comments: A nice idea and well designed study is actually wrong by the selected results The data selected for presetnation and the way that are presented, make it very diffcult to understand the findings of the study.
More details ans suggestion are writen in the basic reporting section of the review.


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: VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES
Review round: 1
Reviewer: 3

Output:
Basic reporting: -In general, the syntax is not clear. It is difficult to follow the text. The structure of the sentences should be improved. For example, keep sentences shorter (about two lines).
-Please, check layout for submitting to this journal
-Lines 1 to 20: Please check the journal layout to identify affiliation, how to report an affiliation, and how to indicate and the corresponding authors and the information to be included. (See template of the journal: https://peerj.com/about/author-instructions/
-Line3 to 5: Use numbers instead of letters for the affiliation of the authors.
-Not sure that the Short title is needed for that journal (Line 21)
-Check layout required for the abstract: https://peerj.com/about/author-instructions/
Subheadings must be bold, followed by a period, and start a new paragraph e.g.
Background. The background section text goes here...
-Line 24: Please add information regarding the Background of the question, instead of starting with the aim of the paper.
-Line 27: Be consistent with the abbreviation selected to use. BHB or BHBA? Or define both if needed.
-Line 33-36: Sentence not clear
-Line 33: Please, report the p-value in the format required by the journal. If not clear, in the journal guidelines, please keep a space at both sides of the “<”.
-Line 37: What do you mean “in practical conditions”?
-Line 38-39: Start each Keyword with capital letters. Check the layout required by the journal.
-Line 41: This journal does not require “Implications”. Please check the sections required. However, it cannot be a unique sentence and it is too long.
-Line 51: Do not start a sentence with an abbreviation.
-Line 62-63: The reference from Benedet et al. 2019 is not correctly used because it is a review in cows, not in sheep or in ruminants in general. Please check that the references in the text are correctly used and placed.
-Line 78-80: Include a references for that sentence.
-Line 80: Do not use the capital letters in a sentence between brackets.
-Line 82-84: Check the structure of the sentence.
-Line 98: Make subsections to make it easier to follow. For example: animals, sampling collection, analysis, statistical analysis, etc.
-Line 101: Specify the country.
-Line 103: Revise the sentence.
-Line 159: Indentation of the paragraph.
-Line 167: Check how to report states and countries and keep it through all the test.
-Line 200: the term “somehow” is colloquial, be more specific.
-Line 172: If none of the animals presented any clinical sign of PT, the study does not respond correctly to the objective presented in Line 95.
-Some references are very old.
Experimental design: -Line 24: Abstract information does not match the material and methods. In the abstract, it is indicated 24 ewes, but in the material and methods, they are indicated 19. The number of samples in the abstract also is for 24 ewes, not for 19. Moreover, the Approved Ethical Committee is for 20 ewes. Also, the blood samples indicated in the abstract (312) are consistent with 24 ewes, not 19.
-Line 102: Indicate also the body condition score.
-Line 102: It was measured at the beginning of the adaptation period? Please, specify.
-Line 105: Provide chemical composition of the hay and the composition and chemical composition of the concentrate.
-Line 111: Check sentence. The use of “elsewhere” is not adequate for that sentence. Better refer directly to the reference.
-Line 111-112: Check sentence to make it easier to follow.
-Line 112: “Then, each ewe received”
-Line 112-115: Check punctuation to make it easier to follow.
-Line 111-119: Not clear the steps followed. Reconsider the order of the sentences.
-Line 123: Where does also in the left external jugular.
-Line 123-124: Not clear. Are those sampling times reported as minutes after the infusion?
-Line 126: What means SP? It also indicates the country.
-Line 150: Check sentence. The use of “elsewhere” is not adequate for that sentence. Better refer directly to the reference.
-Line 152-153: Number of samples and ewes do not match with the ones in the abstract.
-Line 172: Ketonuria refers to urine. Was urine check for that? How?
Validity of the findings: -Table 1: Explain all abbreviations
-Error in the number of the Tables. Please check it.
-The second “Table 2”, delete the comma after “et al.,”.
-Figure 1. Abbreviation of the y-axis is not defined in the caption of the figure. Also indicate the number of samples.
-Figure 4: x-axis indicates that is “sampling time”. Indicate when the infusion started and finished.
-Line 199-200: Is the temperature pre-infusion significantly different from the temperature infusion and post-infusion?
-Line 200 to 202: Sentence not clear.
-Line 201: Which times were classified at false-positive samples?
-Line 205-207: Sentence not clear.
-Line 208: How was ketonuria evaluated? It is not described in the material and methods section.
-A graph with the BHB concentration at each sampling point for both methods could be helpful to follow the paper.
-Line 210: Why? Which is the differences between the studies?
-Line 216: Any explication for that?
-Line 230: This is a result sentence not a discussion.
-Conclusions are poor.
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: VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES
Review round: 2
Reviewer: 1

Output:
Basic reporting: Peer J; 43743v1 VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES

The manuscript has improved substantially with the revision. Practically the authors have taken into account the indications of all the reviewers and made the appropriate modifications, and in my opinion it is ready to be published.
I would only recommend some small changes, which would improve the article, some referring to the figures and tables and other grammatical errors.
Experimental design: ..
Validity of the findings: ..
Additional comments: Line 28: "... Βeta-hydroxybutyrate acid (BHB), ..." replace with "... beta-hydroxybutyrate acid" or "... β-hydroxybutyrate acid (BHB) ...". Do not capitalize B. Use this same criterion on line 59 and whenever this word is used.
Line 175. Animals, italicize. Unify with other subtitles
Line 114: remove “Chemical composition of the diet”
The word “bellow” continues to appear in the title of Figure 3. It is an error that has not been corrected. Replace with below.
After the modifications made to the tables and figures, it is still difficult to understand them. Especially review table 2, try to make them easier to understand. I propose the following Table 2.
Classification in moderate (BHB: 0.8-1.6 mmol / L) and severe (BHB ≥ 1.6 mmol / L) ketosis. Take into account the entire manuscript. Especially check line 282 (should put BHB ≥ 1.6 mmol/L), Table 2, Figure 2, etc.


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: VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES
Review round: 2
Reviewer: 2

Output:
Basic reporting: This is the second revision of the manuscript.
Authors, paid attention and made special effort to answer all comments and questions and made significant corrections and changes in the manuscript. In the form it is now it is much more comprehensive and to the point. In general, the revised manuscript presents clearly the work and stresses the results and the novelty of the study. As it was stated and in the first revision, a direct comparison of BHB values measured by 2 methods and obtained each sampling time, would be of interest, since the induced hyperketonaemia is the novel thing of the study. Authors claimed this was published elsewhere, which also diminishes the value of the manuscript. I didn’t notice any changes in tables and figures in the revised submission, as they are exactly as submitted for first time. Given that it is still not possible to read the Table 2, which is positive or negative by which method. It is not clear in the way presented.
Experimental design: Experimental design is as at the first submission, but authors made all the recommended changed in the presentation of the design and the material and methods section, so as to be clear and comprehensive. Experimental design is simple, clear and adequate for the aims of the study, so are the laboratory techniques and the statistical methods performed.
Validity of the findings: Discussion is improved and remains relevant, supports the findings and enlightens the results of the study. Conclusions are adequate and literature is up to date and representative of the study.
Additional comments: The re-submitted revised version of the manuscript offers significant changes and improvements, following reviewer’s suggestions and comments. In general, manuscript is better, sufficient and adequate considering its aims, its methods and results. There are some minor comments, that would meke the manuscript better:
(Line numbers refer to tracked changes word archive)
L26-28. Language needs improvement as to be more comprehensive. A suggestion:
The end of pregnancy is the period with the highest risk of occurrences of ketosis and pregnancy toxaemia due to fat mobilization and increasing non-esterified fatty acids in the liver which are converted in ketone bodies

L125 “Chemical composition of the diet” This is not presented or discussed.

It is still not possible to read the Table 2, which is positive or negative by which method.It is not clear to the reader, the positive and negative results should be presented clearly for both methods used.

L232-234 It is noteworthy that the ketonemia induction model resulted in 61 points of data (24.7%) with BHB concentration above 3.0 mmol/L.
Difficult to understand what is noteworthy. According to raw data se provided In the raw there are 63 points –values higher than 3,0 mmol/lt, but almost all (52) of them  are referred to HHM. This could be stress also in discussion, as enlighten that HHM measure and give result in high BHB values, but there are usually false positive! according to GS method

L210 where instead of were.


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: VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES
Review round: 2
Reviewer: 3

Output:
Basic reporting: -	Some words seem to be in grey (for example L28 and Lines 30, or Line 60).
-	Check punctuation used (see Line 28).
-	Sentence too long (Line 26-28).
-	Line 28: “is” or “are”?
-	Line 30: indicate the brand.
-	Line 30: Instead of indicating here the brand of the handheld meter, I think it is better to indicate it on the Methods (Line 37).
-	Line 40: specify “respectively” also for 0.98 and 0.89.
-	Revise punctuation and sentences too longs. That makes difficult to follow the paper. For example, Line 39-42, or Line 55-56.
-	Check English grammar (Line 43: revise “at the field conditions”
-	Line 51: Rewrite as suggested “and, consequently, of the highest risk of occurrences…”
-	Line 53: What do you mean with “conditioned”? Sentence not clear.
-	Line 56: Not clear. The disease reduces the DM intake, or predisposes the disease because of the decrease of DM intake?
-	Line 61: To which ketone body is referring?
-	Line 71: Revise “Monitoring of BHB concentrations”
-	Line 78: “is most” or “is more”?
-	Line 78-79: “in blood”
-	Line 79: Revise the sentence. Suggestion: “it is not volatile and more abundant than the other two ketone bodies”
-	Line 81-83: Sentence not clear. Check the correct place to indicate the name of the handheld meter used, and to what is referring “this”.
-	Line 85: Confusing structure of the sentence. Revise “It has recently also been validated for use”
-	Check extra spaces between words, such the one in Line 85 (for use  in dogs”).
-	Line 92-95: Sentence difficult to follow.
-	Line 95-97: Sentence difficult to follow. Not clear “with lower blood concentrations of BHB”. Maybe it is easier to indicate the range where it has been validated.
-	Line 98: Aim not clear. “wide” or “wider”? What means “from predictive values to ewes with clinical signs of PT”?
Experimental design: -	Line 104: Although I imagine that the University of Sao Paulo is in Sao Paulo (Brasil), you think that you need to indicate the city (or state) and the country.
-	Line 108: Revise sentence “a 30 days adaptation period of 30 days”.
-	Line 111-Line 112: Sentence not clear.
-	Line 114: Not clear: What do you mean “to correct the diet”?
-	Line 113: I’m not familiar with this specific breed. Is the average BW reported in agreement with that breed? Or they presented under or over BW?
-	Line 115: Sentence not finished.
-	Line 115: The chemical composition of the coast-cross grass and the commercial concentrate is relevant for the readers. So they can determinate by themselves if the animals were correctly fed. I strongly recommend adding as much as you can regarding that point. At least, for the commercial concentrate, it should be easy to ask the company that produced it to provide that information. About the hay, I do not know if you produced it, or you buy it for the experiment, but usually, the chemical composition is determined to see if animals’ requirements are fulfilled.
-	Line 118-125: Did the animals had free access to mineral salt block and water?
-	Line 119-120: Check sentence order and the use of a comma.
-	Line 120: “&” or “and”?
-	Line 124: administered?
-	Line 128-129: Sentence not clear: “with a total of 120 minutes of infusion for all sheep”
-	Line 130-132: I think it is missing a comma “During and after the infusion, blood”. No need for the “:”.
-	Line 133: I think that “samplings” is a more adequate word than “collections”.
-	Line 134: “(T8 to T13)”.
-	Line 136: Why you indicate here the State, but you did not indicate it in Line 111?
-	Line 136: Sao Paulo is not written always in the same way.
-	Line 140: missing a stop before “All sheep”
-	Line 140: Fully recovered from what?
-	Line 142: “from …. to …”
-	Line 147: See comments for Line 136.
-	Line 145: Check rules around commas and time phrases and correct that trough all the manuscript.
-	Line 162-164: Do not use single-sentence paragraph.
-	Line 164: To which techniques are referring?
Validity of the findings: Results
-	In general, two decimals are enough, report the results between brackets in a way that they can be understood, use point for decimals. For example, lines 198 to 203.
-	Line 185-186: Sentence not clear: “non presented any”.
-	Line 188-189: Missing space between numbers and units.
-	Line 192: maybe better refer to methods instead of tests?
-	Line 194: usually 2 decimals are enough, and makes easy to follow.
-	Line 194: information between parenthesis difficult to follow, maybe rewrite/reorder.
-	Line 194: write the equation to make it easy to follow. For example, add the ×, put a comma to separate the equation from the information between brackets, etc.
-	Line 197: Do not start a sentence with an abbreviation.
-	Line 197: Check if Bias needs to go with capital letters.
-	Line 210: What means 61 points of data?
-	Line 212 and Line 215: higher instead of high.
-	Line 214: I think it is better “greater” than “further”.
-	Line 212-216: Still not clear enough.

Discussion
-	Line 219-212: Sentence not clear.
-	Line 212: Not clear. “blood concentration was greater than 1.0 mmol/L BHB”?
-	Line 225-227: Better to indicate “(61 out of XX; 24.7%)”. Also for the referenced papers in Line 226 and 227.
-	Line 231: Revise the sentence.
-	Line 234: Remember to use the past tense. (Check all the manuscript for that).
-	Line 234-238: Sentence too long
-	Line 245: that this was in cattle was already indicated at the beginning of the sentence.
-	Line 245-247: Check sentence. Place the comma after BHB.
-	Line 247-249: Sentence difficult to follow. Maybe you need a comma in “, and the sensibility”, or you should try to present the same structure in both parts of the sentence “, and high sensibility”.
-	Line 250-252: Sentence not clear. To what is referring “the and perfect agreement”? seems something is missing.
-	Line 250-253: Avoid creating a single-sentence paragraph.
-	Line 261: Missing comma before “the greater”?
-	The term “somehow” feels not accurate. I recommend to find another, or at least, do not use it so close in the text. For example, it is in Line 256, and then in Line 262.
-	Check the use of the verb “evoke”.
-	Line 272-277: Rewrite conclusions. Sentence too long (Line 272-276), and not clear what means the last sentence.
Additional comments: Although the authors have improved the manuscript, there are still some issues they need to improve. I strongly suggest revising carefully the English (in particular the grammar and the way to structure sentences). Moreover, American and British English are mixed in the document. The idea of the material and methods is that other researchers would be able to replicate the study’s conditions, because of that it is important to provide information regarding the composition and chemical analysis of the feed administered.


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: VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES
Review round: 3
Reviewer: 1

Output:
Basic reporting: In my opinion the authors made a special effort to respond to all comments, made the corrections and changes suggested by the reviewers in the manuscript.
I wish to congratulate the authors for the work done, for the innovative experimental design and for the results obtained that are reflected in the current manuscript.
Therefore the manuscript meets conditions to be published.
Experimental design: None
Validity of the findings: none
Additional comments: In my opinion the authors made a special effort to respond to all comments, made the corrections and changes suggested by the reviewers in the manuscript.
I wish to congratulate the authors for the work done, for the innovative experimental design and for the results obtained that are reflected in the current manuscript.
Therefore the manuscript meets conditions to be published.


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: VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES
Review round: 3
Reviewer: 2

Output:
Basic reporting: The document has improved. However, some issues are still not clear or need to be revised.
-Check American/British spelling also in tables and figures.
-Reduce decimals in Tables and Figures, and report in the same way the formula (equation) in Tables and in the text.
-Check Tables and Figures for the comma and spot to report decimals.
-Figure 1: Indicate in the title what is the blue line, and the red one. Add in the figure the equation and the r.
-Figure 4: Indicate in the figure when the infusion started and when it finished. Could you report the ES instead of SD, so it is more clear when point differs significantly, or, if you keep using SD indicate which point significantly with the baseline.
-When reporting XX ± XX it is not clear if you are reporting ES or SD. Please make it clear. Moreover, if a P-value is associated with that information it is better to indicate the ES and not the SD.
-Line 39: Here you are reporting a range, not only the threshold. I suggest “BHB concentration range”.
-Line 41: Start a new sentence for severe ketosis.
-Line 83: Correct capital letter used for “It”
-Line 91: sentence not clear, something is lacking.
-Line 110: to what is “(2.7%)” referring?
-Line 117: Could the editor check if this is correct for that journal. Even if using software for adding the references, the journal guidelines (or recently published papers) have to been checked to see how they report citations within the text. Usually “&” is only used inside a parenthesis.
-Line 118: Check the need of adding a comma after “Before the BHB infusion”
-Line 149: there is an extra stop at the end of the sentence
-Line 161-163: A unique sentence cannot be a paragraph. That was not corrected from the previous submission.
-Line 164: Still not clear to which techniques are referring. I suggest stating their name again.
-Line 170: This does not agree with the following sentences. I suggest to delete it.
-Line 192: use the symbol ×, not the x.
-Line 194: showed instead of show.
-Line 197: Not clear “for normal and moderate”.
-Line 227: use “It was” or “they were”.
-Line 232: Use “There are no previous studies”
-Line 242: “this could increase”
-Line 244-245: Difficult to follow.
-Line 246-250: A unique sentence cannot be a paragraph. That was not corrected from the previous submission.
Experimental design: N/A
Validity of the findings: N/A
Additional comments: N/A


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

Input:
Title: VALIDATION OF A HANDHELD Β-HYDROXYBUTYRATE ACID METER TO IDENTIFY HYPERKETONAEMIA IN EWES
Review round: 4
Reviewer: 1

Output:
Basic reporting: Some (very) small comments regarding Table 1:
-Table 1 is indicated in the M&M but it was not at the end of the manuscript with the other Tables.
-Table 1 needs to indicate the units of the parameters (%) and is they are calculated on a DM basis.
-I think that the paper is using British spelling, but in Table 1, “fiber” is written with the American Spelling. In British is “fibre”.

No further comments to add.
Experimental design: No comment
Validity of the findings: No comment
Additional comments: No comment


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

Input:
Title: EFFECTS OF PRESERVATION DURATION AT 4 °C ON THE QUALITY OF RNA IN RABBIT BLOOD SPECIMENS
Review round: 1
Reviewer: 1

Output:
Basic reporting: The manuscript entitled “Effects of preservation duration at 4 ℃ on the quality of RNA in rabbit blood specimens “ deals with the quality of RNA from whole blood and leukocytes isolated from rabbit blood in regard to prolonged storage. The obtained results suggest that the purity of whole blood RNA and leukocyte RNA does not indicate significant change after rabbit blood is stored at 4 ℃ for different preservation durations (from 1 hour to 7 days). The integrity of leukocyte RNA indicates the same result as above, but the integrity of whole blood RNA is significantly decreased after rabbit blood is stored at 4 ℃ for over 3 days. As for the expression of SMAD7, MKI67, FOS, TGFβ1 and HIF1α of whole blood RNA and leukocyte RNA it remained stable, but PCNA expression of whole blood RNA or leukocyte RNA is significantly decreased after rabbit blood is stored at 4 ℃ for over 24 hours or 7 days. Authors concluded that high-quality RNA is obtained from the fresher blood specimens, and if blood specimens are stored for over 3 days at 4 ℃, the quality of leukocyte RNA is more stable and of better quality than that of whole blood RNA.
This is a relevant methodological paper dealing with the storage conditions which present relevant and much needed data for this particular field.
Experimental design: Methods section is very well described with sufficient data for the study to reproducible. Since the study involves animal it seems that all the ethical standards were fully obeyed.
Please be consistent in the writing of either "hours" or "h" throughout the manuscript.
Validity of the findings: Data are very comprehensive and are graphically presents and easy to read.
Letters and numbers on the x and y axes are a bit small and difficult to read.
Additional comments: This is a very interesting methodological paper dealing with the impact of storage conditions of the quality of RNA.
This could be very interesting data for the ones working in the field of gene expression.
The first paragraph of the Discussion section is a bit of a repetition of what was already stated and could be shortened a bit.
If the Authors noted that that the mRNA of PCNA may be influenced by the preservation duration of blood specimens at 4 ℃ which obviously needs more research and they emphasize that it is something that should be done in the future, is it possible to do this additional experiments to resolve this issues already in this study and present this data in the submitted publication. It will certainly be an added value for the current manuscript.
Please use capital letter at the beginning of the first sentence of the Conclusion section.