Metagenomics-Based Analysis of the Effect of Rice Straw Substitution for a Proportion of Whole-Plant Corn Silage on the Rumen Flora Structure and Carbohydrate-Active Enzymes (CAZymes)

Round 1

Reviewer 1 Report (Previous Reviewer 3)

My remarks were taken into consideration. OK to publish in the present form.

Author Response

Dear reviewer 1,

Thank you for reviewing our manuscript entitled " Metagenomics-based analysis of the effect of rice straw substi-tution for a proportion of whole-plant corn silage on the rumen flora structure and carbohydrate-active enzymes (CAZymes)" (fermentation-2688171) and expressing your approval of our revisions. We have learned a lot from your previous comments, which have given us a deeper understanding of the rigor and logic of scientific research. Thank you very much for your help in refining this article.

We made some minor changes and ensured that these modifications do not affect the content and structure of the paper. We have not listed these changes here but have highlighted them in red and purple in the revised paper

Please allow me to express my gratitude to you once again. I wish you good health and a happy life.

Kind regards,
Mr. Yubin

Author Response File: Author Response.pdf

Reviewer 2 Report (New Reviewer)

In this manuscript, the authors used metagenomics sequencing to analyze the rumen microbiota of cows fed a diet that replaced a portion of whole-plant corn silage with rice straw. Their study provides information for the potential use of rice straw to reduce costs. The experiments were designed and conducted systematically and convincingly. The explanations were clear, and the conclusions are well-supported by the data. A minor revision is sufficient prior to publication.

#Line 110: The method to measure the concentration of NH3-N should be briefly described, even though you have a citation.

#Line 112: Although the authors describe the parameters of GC-MS, they should mention how they identified and quantified the chemicals.

#Line 123: Providing a supplementary figure to show the degree of DNA degradation can make the data more convincing.

Author Response

Dear reviewer 2,

Thank you very much for your professional advice and your recognition of our article. These suggestions have contributed to enhancing the academic rigor of our paper. In accordance with your advice and requests, we have made revisions to the manuscript. We hope that our work meets your approval once again. The details of the changes are in the appendix for your perusal.

We sincerely appreciate your dedicated work and hope that the revisions will be recognized. Thank you again for your comments and suggestions.I wish you good health and all the best!

Kind regards,

Mr.Yubin

Author Response File: Author Response.pdf

This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.

Round 1

Reviewer 1 Report

v This study aimed to investigate the effects of replacing a portion of whole-plant corn silage with straw on rumen microbial community structure and carbohydrate-active enzyme activity in dairy cows. It is an interesting study and fits the scope of the journal. However, a major revision is required before the final judgment. Please address the appended comments.

v Line 27: Please increase keywords including different words not mentioned in the title of the manuscript.

v Line 30-40: Please mention these roles.

v Line 34: Several researchers found.......

v Line 36: Please mention the kind of these relationships.

v Line 41: Feed resources?

v Line 44-45: In which animal species?

v Line 51-60: This should be combined with the previous paragraph.

v Line 61-64: The rationale of the study is very poor. Please try to enhance it.

v Line 68-68: late lactation: please specify more.

v Line 68-69: Body weight, body condition score, and age (mean, SD) should be specified.

v Line 74-75: The authors should explain why they selected these time points for the study.

v Line 80-81: animal ethical approval number?

v Line 83: I recommend a schematic figure for the experimental procedures.

v Line 83-89: Please add an appropriate reference(s).

v Line 101-102: Please describe more.

v The methodology section needs moderate language editing.

v Also, please provide the appropriate references.

v Line 186: Statistical section should be provided as a single subsection.

v Line 197-208: The result section should have digital values for the obtained data. Please improve this section using the p-value.

v Line 211-213: The figure legend should be more descriptive.

v Line 308-311: should be at the end.

v Line 251-341: The discussion section needs rewriting. Please try to explain your findings simply. Avoid very long paragraphs.

Moderate editing of English language required

Author Response

Dear Editors and Reviewers: 

Thank you for your letter and for the reviewers’ comments concerning our manuscript entitled “Macrogenomic-based analysis of the effect of rice straw sub-stitution for partial proportion of whole maize silage on rumen flora structure and carbohydrate-active enzymes (CAZymes)” (ID: fermentation-2522411). Those comments are all valuable and very helpful for revising and improving our paper, as well as the important guiding significance to our researches. We have studied comments carefully and have made correction which we hope meet with approval. Revised portion are marked in red in the paper.Major corrections in the paper and responses to reviewers' comments can be found in the Appendix.

Author Response File: Author Response.pdf

Reviewer 2 Report

The study compared the rumen metagenome of two small groups (4/group) of lactating dairy cows under the feedings of corn silage (CS) or rice straw (RS) replacement. The manuscript only has only two pieces of metagenomic data, microbial taxa and the genes of carbohydrate-active enzymes, but lack other data, such as performance (milk, intake, etc) and metabolites (blood biochemicals, rumen fermentation products, etc), for examining the correlations and the significance of observed differences in rumen metagenome. This clear defect minimize the scientific value of this manuacript, making it just a thin report from a single experiment.

Besides the major concern above, there are some additional concerns on experimental design and writing.

---In the Introduction, the authors stated that "Low-quality feed has a high content of crude fiber and fewer available nutrients". I suppose rice straw is considered as a low-quality ingredient in this study. However, RS feed had lower percentages in both NDF and ADF than CS feed in (Figure S1. Note: it should be called as Table S1) . In addition, nutrient composition of two feeds were not provided.

---Macrogenomics is mentioned in the title and keywords, but is absent in the body text. This is a metagenomics study. Why use "macrogenomics".

---A focus on carbohydrate-active enzymes was not well justified. Do CS and RS have great differences in carbohydrate composition, such as arabinoxylan and arabinan mentioned in the Abstract?

There are clear issues in grammar and writing style.

Author Response

Dear Editors and Reviewers: 

Thank you for your letter and for the reviewers’ comments concerning our manuscript entitled “Macrogenomic-based analysis of the effect of rice straw sub-stitution for partial proportion of whole maize silage on rumen flora structure and carbohydrate-active enzymes (CAZymes)” (ID: fermentation-2522411). Those comments are all valuable and very helpful for revising and improving our paper, as well as the important guiding significance to our researches. We have studied comments carefully and have made correction which we hope meet with approval. Revised portion are marked in red in the paper.Major corrections in the paper and responses to reviewers' comments can be found in the Appendix.

Author Response File: Author Response.pdf

Reviewer 3 Report

This paper is interesting because the valorization of raw materials rich in cellulose by the ruminants  is very important from an economic point of view.

The design is good and the methods used, especially as I can judge seem innovative to me.

I have only few comments :

lines 71-73 and figure S1 it would have been good to have a system a little better adjusted at least in CP and possibly NEl so that variations in the microbiota can only be justified by a change in the fibre source.

Line 78 S1 is not a figure but a table please correct

Line 262 "observed" please precise "in the RS group"

This paper is written in correct English

Lines 40-41 The term "low quality feed" does not seem appropriate to me, we should find another term, "low quality raw material", "low cost co-products", or something else.

Line 46 : idem as above

Author Response

Dear Editors and Reviewers: 

Thank you for your letter and for the reviewers’ comments concerning our manuscript entitled “Macrogenomic-based analysis of the effect of rice straw sub-stitution for partial proportion of whole maize silage on rumen flora structure and carbohydrate-active enzymes (CAZymes)” (ID: fermentation-2522411). Those comments are all valuable and very helpful for revising and improving our paper, as well as the important guiding significance to our researches. We have studied comments carefully and have made correction which we hope meet with approval. Revised portion are marked in red in the paper.Major corrections in the paper and responses to reviewers' comments can be found in the Appendix.

Author Response File: Author Response.pdf

Reviewer 4 Report

I have reviewed the manuscript, and here are the comments:-

English needs to improve; there are some sentences and parts of the manuscript that are difficult to understand

Many abbreviations in the manuscript need to be cleared, and add more results in the abstract.

Unclassified Bacteria Words: these should not write in italics; only scientific names need to be written in italics

Please arrange all keywords in alphabetical order.

The materials and methods are difficult to follow in many cases. Extensive revisions are required. The authors should read the manuscript carefully and provide more details in any analysis. In addition, provide the proper reference.

Provide the origin and model of all devices.

Section: 2.3. Macrogenomics sequencing: Needs critical revision; check and rewrite

To improve the quality of the paper, update the reference list by adding 2022 and 2023 references, especially in the discussion section.

Enhance the resolution of Figure 4

The references section needs critical revision. The majority of the journal names are NOT capitalized as below. References that contain many authors must be shortened according to the journal format; Please correct them.

In addition, many scientific names in the references must be written in italic format.

Please follow the authors' instructions in writing the reference in the list. For references about textbooks, please add the page numbers of the textbook. Also, please add the city of the publisher.

English needs to improve; there are some sentences and parts of the manuscript that are difficult to understand

Author Response

Dear Editors and Reviewers: 

Thank you for your letter and for the reviewers’ comments concerning our manuscript entitled “Macrogenomic-based analysis of the effect of rice straw sub-stitution for partial proportion of whole maize silage on rumen flora structure and carbohydrate-active enzymes (CAZymes)” (ID: fermentation-2522411). Those comments are all valuable and very helpful for revising and improving our paper, as well as the important guiding significance to our researches. We have studied comments carefully and have made correction which we hope meet with approval. Revised portion are marked in red in the paper.Major corrections in the paper and responses to reviewers' comments can be found in the Appendix.

Author Response File: Author Response.pdf

Reviewer 5 Report

ok.

Macrogenomic-based analysis of the effect of rice straw substi- 2 tution for partial proportion of whole maize silage on rumen 3 flora structure and carbohydrate-active enzymes (CAZymes)

Keywords: straw; Macrogenomic; CAZymes; rumen flora structure;

There are repeated words in the title and as keywords. Must appear in only one place

Author Response

Dear Editors and Reviewers: 

Thank you for your letter and for the reviewers’ comments concerning our manuscript entitled “Macrogenomic-based analysis of the effect of rice straw sub-stitution for partial proportion of whole maize silage on rumen flora structure and carbohydrate-active enzymes (CAZymes)” (ID: fermentation-2522411). Those comments are all valuable and very helpful for revising and improving our paper, as well as the important guiding significance to our researches. We have studied comments carefully and have made correction which we hope meet with approval. Revised portion are marked in red in the paper.Major corrections in the paper and responses to reviewers' comments can be found in the Appendix.

Author Response File: Author Response.pdf

Round 2

Reviewer 2 Report

The authors added the rumen metabolite data in the revision while other issues in my previous comments remain unaddressed (such as the lack of correlation analysis between microbes and other parameters). More importantly, the response on dietary composition further indicates the poor execution in this research project, and inadequate responses to other comments may also imply inadequate engagement of senior investigators in the effort.

Careful proofreading on grammar and typos is needed.

Reviewer 4 Report

The authors have carefully processed all comments. The quality of the manuscript has increased significantly. I have no further comments.

 Minor editing of the English language is required.