Diversity and Function Potentials of Seed Endophytic Microbiota in a Chinese Medicinal Herb Panax notoginseng

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript titled 'Diversity and functional potentials of seed endophyticmicrobiota in a Chinese Medicinal herb Panax notoginseng' is fairly well written. This manuscript investigates the diversity and functional roles of seed endophytes in Panax notoginseng using culture dependent methods and amplicon sequencing. It shows that diverse seed endophytes, particularly antagonistic bacterial strains like Bacillus cereus and B. toyonensis can suppress root pathogens and promote seed germination and seedling growth, highlighting their potential for sustainable agriculture. 

May the authors consider the following comments to improve the manusxript;

Line 119 - may you include information on gel-running conditions including gel percentage used and the gel cutting process. Was it manual or automated?

Figure 1 - May the authors provide a brief description of the schematic. Although the information might be in the methods section,the figure legend must provide a clear description

Line 386 - remove the word 'In'

Line 448 - correct spelling to 'substantial'

Although the conclusion is concise and well written, it lacks recommendations for further research. Adding potential applications of the findings may also improve the manuscript. 

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript "Diversity and Function Potentials of Seed Endophytic Microbiota in a Chinese Medicinal Herb Panax notoginseng" written by Hong-Yan Hu et al. explores the diversity, community structure, and functional potential of seed endophytic microbiota in Panax notoginseng. Using both culture-dependent and amplicon sequencing approaches, the authors characterized microbial communities in seeds from different plants and evaluated the antagonistic activities of bacterial strains against Fusarium spp. Notably, Bacillus cereus and B. toyonensis showed biocontrol potential and enhanced plant growth, suggesting value for sustainable agriculture.

The manuscript is good and must be accepted to improve. However, some problems must be solved before.

Materials and Methods: some problems, as described bellow:

Surface Sterilization: Use of HgCl₂ is effective but toxic. No mention of residue removal validation (e.g., chemical neutralization).

No Control for Endophyte-Free Seeds: There’s no mention of a sterilization control to verify endogenous versus exogenous microbes.

Dual Culture Bioassays: Lacks clear justification for using only 11 strains from 256 isolates.

Statistical Power: No sample size calculation or statistical power assessment mentioned.

Use of Duncan’s Test: While valid, ANOVA followed by Tukey’s HSD may be more conservative and preferred in recent standards.

 

Results:

Over-reliance on relative abundance without absolute quantification. Maybe include qPCR validation or CFU counts for representative taxa.

Tables and figures are not always self-explanatory, then add explanatory notes or visual keys (e.g., define OTUs in figure legends).

Inhibition rates lack confidence intervals: Add error bars or statistical significance indicators in Table 5 and Figures.

The legends were too poor and did not fully explain the data and schemes.

 

Discussion:

Some data from results are restated without further interpretation.

Does not address ecological safety or potential risks of applying Bacillus spp.

Lacks consideration of limitations (e.g., localized sampling, only one growth environment).

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

Manuscript 'Diversity and Function Potentials of Seed Endophytic Microbiota in a Chinese Medicinal Herb Panax notoginseng' by Hu et al. is well written. I would like to comment on just a few points.

The introduction, although it covers a couple of issues that will be addressed, is very brief. Develop the topics so that the reader becomes more familiar with the subject matter.

Materials and Methods section is clear and sufficiently detailed so that other researchers can replicate the study. However, statistical indices should be described in detail.

Results: Graphs and tables are sufficient to highlight key results. Avoid repeating all the figures in the text.

Discussion: Begin by explaining what your conclusions mean in relation to your research questions or hypotheses.

Are there any limitations to the study?

Further, please provide suggestions on how to expand or improve this study.

Conclusions short and concise.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Reviewer 4 Report

Comments and Suggestions for Authors

The manuscript is descriptive, but offer the information of endophytic bacteria that could participate as bacterial growth promoting of the Chinese Medicinal Herb Panax notoginseng

1. include the main goal of the work
2. In general, in the figures, in the figure legend specify how many repetition were done (n=3, 4?)
3. the authors need to demonstrate that in vitro the interaction of B. cereus and  B. toyonensis vs. Fusarium oxysporum in confrontation assays, at least in agar plates, to demonstrate the inhibition of Fusarium by those bacterial strains
4. The authors need to demonstrate at least which is the factor that bacteria inhibits the fungal growth, by volatile or diffusible factor?
5. The conclusion needs to be more concise; it is too long.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

Dear authors, some suggestions and comments:

The manuscript was improved a little bit

The statistical test must be redone with SNK or Tukey, not Duncan.

The figures are too small and the fonts are smaller. Please see the attached manuscript. Figure 6 should be drawn in color or with another type of marking. Grayscale confuses treatments.

I disagree with the authors when they mention that qPCR could interfere with the other results. The authors could have had two subsamples, applying qPCR to one and the validations presented in the manuscript to the other. I believe this explanation is very flawed, and I urge the authors to rethink this strategy in future manuscripts.

I suggest that authors make corrections in the next version using red fonts as this makes the review process easier and speeds up the analysis of the manuscript.

Comments for author File: Comments.pdf

Author Response

Dear editor and reviewers,

Thank you very much for giving us the opportunity to further improve our manuscript. Having carefully considered the comments raised, we have revised the manuscript as far as possible and attached a point-by-point response to the comments below.

Response to Reviewer 2’ Comments

Comments 1: Dear authors, some suggestions and comments:

The manuscript was improved a little bit

The statistical test must be redone with SNK or Tukey, not Duncan.

Response 1: The manuscript has been revised accordingly. Specifically, the statistical analysis has been re-performed using Tukey's HSD test instead of the Duncan test, as recommended. Thank you again for your valuable feedback.

Comments 2: The figures are too small and the fonts are smaller. Please see the attached manuscript. Figure 6 should be drawn in color or with another type of marking. Grayscale confuses treatments.

Response 2: We sincerely thank the reviewer for these helpful suggestions. We have carefully improved all figures in accordance with your comments. Specifically, the sizes and fonts of Figures 2, 3, 4, and A1 have been enlarged to enhance readability, and the legends in Figure 6 have been modified to use distinct colors and marker types, avoiding grayscale to ensure clear differentiation between treatments. Thank you again for your valuable suggestion.

Comments 3: I disagree with the authors when they mention that qPCR could interfere with the other results. The authors could have had two subsamples, applying qPCR to one and the validations presented in the manuscript to the other. I believe this explanation is very flawed, and I urge the authors to rethink this strategy in future manuscripts.

Response 3: We apologise for not understanding your suggestion in the previous response. Using a qPCR validation method on one set of samples would better illustrate the issue. Thank you very much for this valuable suggestion. We will try it out in our future research.

Comments 4: I suggest that authors make corrections in the next version using red fonts as this makes the review process easier and speeds up the analysis of the manuscript.

Response 4: We sincerely appreciate your suggestion. In the revised version, all changes have been highlighted in red font to facilitate your review and ensure a more efficient evaluation process. Thank you for this helpful recommendation.

Reviewer 3 Report

Comments and Suggestions for Authors

Avoid commenting on the results in the Results section and move them to the next section, Discussions. Otherwise, the authors provided responses to the reviewer's comments and the manuscript was improved.

Author Response

Dear editor and reviewers,

Thank you very much for giving us the opportunity to further improve our manuscript. Having carefully considered the comments raised, we have revised the manuscript as far as possible and attached a point-by-point response to the comments below.

Response to Reviewer 3’ Comments

Comments 1: Avoid commenting on the results in the Results section and move them to the next section, Discussions. Otherwise, the authors provided responses to the reviewer's comments and the manuscript was improved.

Response 1: We sincerely thank the reviewer for this constructive suggestion. In accordance with your comment, we have moved all interpretive and explanatory remarks regarding the results from the Results section to the Discussion section. All these changes have been carefully implemented and are now highlighted in red in the revised manuscript to facilitate your review. We believe this adjustment significantly improves the clarity and structural integrity of our manuscript. Thank you again for your valuable guidance.

Reviewer 4 Report

Comments and Suggestions for Authors

In this new version of the manuscript, the authors improve the original manuscript. For this reason, I consider that it is suitable for its publication in this journal.

Author Response

Dear editor and reviewers,

Thank you very much for giving us the opportunity to further improve our manuscript. Having carefully considered the comments raised, we have revised the manuscript as far as possible and attached a point-by-point response to the comments below.

Comments: In this new version of the manuscript, the authors improve the original manuscript. For this reason, I consider that it is suitable for its publication in this journal.

Response: Thank you very much!