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The authors have addressed all comments either by altering the body of the manuscript or via the rubuttal
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The revised manuscript has already revised and the improvments are satisfied. My decision is the manuscript is suitable for publication.
Thank you very much for your submission, you have an interesting manuscript. However, as noted by 3 reviewers, the manuscript needs a lot of work. Please, address all comments point by point.
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Please find the full report below.
Please find the full report below.
Please find the full report below.
Jirayu Tanprasertsuk and colleagues investigated the effect of a mildly cooked diet for 4 weeks in healthy household dogs. I found the paper confusing and of limited relevance to current knowledge in the field. Dog owners’ perceptions of changes in health outcomes are ambiguous and almost impossible to interpret, as the authors noted it themselves. Moreover, the fact that some parameters were not available in all dogs (e.g. in Table 1) makes the results questionable. The paper is also often difficult to grasp, particularly the use of values from the PCo1 axis of Bray Curtis dissimilarity only, and the categorization of dogs based on the degree of PCo1 (or 4?, see line 775, label of Figure 2) axis values. I always encourage the use of novel alternatives to analyze the gut microbiota, but the authors give little information to back up their approach. Finally, there are sections in the manuscript that appear too careless (e.g. references, font types).
General comments
The authors use the word “species” a lot, and do not use (or even mention) OTU or amplicon sequence variants. Read this: https://pubmed.ncbi.nlm.nih.gov/12142474/ and consider rephrasing the entire manuscript.
The squares, for missing symbols, is very distracting. The authors should have reviewed the final PDF before submitting.
In the abstract generated by PeerJ, page 4 of PDF, is written the word “rho”, instead of the Greek letter. This is the only time they do so.
How can you discard the possibility, or the effect, of natural variation over time? See this paper in humans: https://doi.org/10.1186/gb-2011-12-5-r50.
The results need to be carefully re-organized, there are way to many headings (e.g. 3.4 Gut microbiome) and sub-headings (3.4.1 Diversity).
Title
In the title, “heterogeneity of responses” sounds ambiguous and confusing, please rephrase, focusing on a title that is both short and informative regarding the nature of this research work.
Abstract
Lines 56-57. This is confusing, is there any other way to write this? Please remember that the paper needs to be easy to understand even for people outside of the field of gut microbial ecology.
Line 59. Are these percentages before and after? If so, could you use, e.g., 61% vs 39%?
Line 62. What is the relevance of including these percentages here? Also, it is not clear for me whether these values come from day 28.
Line 74. This statement is too strong, even a strong correlation with dietary protein may be related to other parameters (e.g. liver metabolism etc) and not necessarily related to the possibility of personalized nutrition. This is concerning, because it goes along with one of the main purposes of nomnomnow.
Introduction
In the first paragraph of the introduction, where is reference 2?
Material and methods
Line 305. Could you please back up this approach?
Results
Line 316. What does “dog-weeks” mean exactly?
Lines 339-340. Could you add the numbers of dogs as well (e.g. 19 out of 31 dogs for the 61%?)? Also, and more importantly, these perceptions are ambiguous and almost impossible to interpret.
Line 350. This section is very confusing, the authors should make an effort to be clear and concise. From my point of view, the authors dug too deep.
Line 377. Why citing only these two studies? This can go into the discussion, with a more general overview of the many other studies that have studied the gut microbiota in dogs.
Line 380. Here it says abundances, and in 3.4.2 says relative abundances. I am not sure if the authors are aware of the issues related to the use of relative abundances: http://dx.doi.org/10.3402/mehd.v26.27663.
Line 391. This looks saturated, please be concise. Also, please note that the names are ultimately meaningless: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1857736/pdf/rstb20061914.pdf
Line 413. Do you really think is necessary to include all this information? Please back up your answer with appropriate references.
Line 422. I do not think this argument is strong enough to classify the subjects. Please provide useful references or arguments to back up this approach.
Line 457. Please see my comment above about heading and subheading. In this line, it looks strange that this subheading is not numbered. I would re-organize the entire manuscript, focusing on the most relevant results and observations only.
Line 512. All the subsections are confusing and out of place.
Line 524. This weakens the entire argument about the relationship with protein or any other nutrient in the diet.
Line 544. This is subjective and confusing, I would remove it.
Discussion
Eight and a half pages for the discussion is way too much. I admit that PeerJ do not have a page limit, but the authors should nonetheless be concise, clear, and explicit. I would re-organize the entire manuscript, and cut the discussion by half.
Line 568. In the title, you placed GM first. Please be consistent.
Line 577. I am not sure whether this is true, particularly the point on “taxonomic resolution and accuracy”. Besides the reference provided above, please note that even identical marker genes can be found in bacteria with highly divergent ecophysiologies (DOI: 10.1128/AEM.70.8.4831–4839.2004) and this reference is better for taxonomic resolution: https://doi.org/10.1038/s41587-020-0501-8 (please us this).
Line 589. Cite those other studies.
Line 681. Do not use flora.
Line 682. It is surprising that this wonderful reference is only used here, and in that context.
References
Be consistent, e.g. all first letters capitalized or not, same font type (ref. 62).
Figures
There is way too much emphasis on HR, MR, and LR, e.g. in figure 2C. I personally would not place too much emphasis on such ambiguous categorization.
Tables
The information presented in tables 3, 5, 6 and 8 is excessive. It is strange that the font of the title of Tables 3, 4, 5, 6 and 8 is different. Even more strange, the mixing of font types in the title of Table 3.
This paper studies the impact (compared to a baseline kibble/extruded diet) of feeding a mildly cooked diet on the microbiome, and owner-assessed health outcomes in the dog.
The paper is well written and considerable thought appears to have gone into the statistical analysis and interpretation of the results.
The authors have used the available literature widely.
In order to support the hypothesis, some background/introduction on the effects of mildly cooked/fresh diets on health aspects, even if anecdotally, could be included (I note it is discussed in the literature).
The experiment utilises dogs in home, and therefore relies on owner assessment and reporting. The diets used are commercial diets, and as such there is not treatment per say, instead a baseline sample (taken from dogs consuming a variety of commercial extruded/kibbled diets). This lends the study to make generalisations about what was observed; a fact acknowledged by the authors in L747-754.
Methodology: I would like to see data on the cooking process/nutrient composition of the MC diets provided as main Tables rather than supplemental data. This is because the diet is the main driver of the changes in the microbiome. Sequencing/analytical methodology is described well and would be repeatable for any study.
Regardless the authors have considered their data set in novel ways (ie the identification of low and high responders) which adds to our knowledge base on the impacts of diet on the microbiome of the dog.
Overall, the findings reported link back to the hypothesis/study undertaken.
Major comment:
Regarding the term 'mildly cooked' - this needs to be defined (especially in the materials and methods). Similarly, when referencing the literature that use the term 'mildly cooked' the authors should stipulate what this was defined as if the temperatures etc differ from the current study. My preference would be to remove the statement on L121 "also commonly known as fresh diets" as the words cooked and fresh contradict each other. This may seem pedantic, however one issue arising from assessing the impacts of diet on the microbiome is that we as a research community use a variety of terms which, when reading articles in depth, do not reflect the treatments investigated.
Questions arising were:
The age of the dogs is large (1-14 years); given age affects the microbiome, and 'ageing' is affected by body size (as a rough guide) were any considerations taken to assess either breed or 'dog size' in terms of the results?
The MC diets had a range of primary proteins used; studies in the dog/cat and rat have shown that protein source can affect the microbiome; was this taken into consideration?
Minor comments:
The term 'transition' is used (eg L568-569); this (to me at least) implies that the microbiome was assessed at multiple time points AND/OR the microbiome is not considered to be stable. Indeed, your study of 28 days would we would expect to see a stable population.
In your discussion on L625-640: I would expect most high-quality kibbled diets to include many of these ingredients.
I understand the process around looking at the baseline/kibble protein content of the diet on the results is and agree with it in principle. However given 'on pack'/reported protein levels can vary up to 10% from actual it would be good to see a statement along those lines.
In general, I look forward to seeing this work published, and seeing more research into the concept of 'high' and 'low responders' in the future.
The authors aimed to determine the effect of variety of differences between diets on dog microbiome. The study also observed a shift in the diversity and abundance of GM upon diet change adn also the fecal outputamong the enrolled dogs. The resuts indicates a a significant shift in the GM profile of 73 healthy dogs. The magnitude of shift was associated with baseline GM and dietary protein, and 74 may provide a basis of personalized nutrition in companion dogs. These results support future 75 investigation of the impact of a MC diet on health maintenance given its increasing popularity. These findings maybe useful in the understanding for canine foods.
The experimental designs were clear and the statistical analysis was suitable. However, the sample size of the dogs needd to be calculated, in orer to obtain the significant sample sizze for analysis.
The diversity of the gut microbe will be a good indications for the health status. However, the quantitative analysis of the gut microbe maybe needed to be determined because these microbes are associated with the efficacy of microbe to utilize and synthesize the essential elements for dog.
The study sounds clear and useful in term of canine gut microbiome. However, the quantitative analysis of the gut microbe maybe needed to be determined because these microbes are associated with the efficacy of microbe to utilize and synthesize the essential elements for dog.
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