Biotechnological Advances for Enhancing European Chestnut Resistance to Pests, Diseases, and Climate Change

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Comments for Authors:

This manuscript provides an extensive biotechnological advancements aimed at improving the resilience, productivity, and long-term sustainability of European chestnut. Given the significant ecological, economic, and cultural importance of this species coupled with the increasing threats posed by pests, emerging diseases, and climate change, the topic is highly relevant and of considerable scientific interest. The authors effectively highlight major biotechnological tools, including classical and assisted breeding, molecular markers, omics based analyses, micropropagation techniques, and recent progress in genetic modification approaches such as transgenic, cisgenic, and CRISPR/Cas9 technologies. While the manuscript is generally well organized and informative, several aspects would benefit from greater conceptual clarity, improved transitions between sections, and a more explicit articulation of current challenges and research gaps. Detailed comments and suggestions are provided below to help improve the overall quality and impact of the review.

Specific comments:    

1. The title is clear and relevant but somewhat long. Consider shortening or restructuring it for greater clarity (Biotechnological Advances for Enhancing European Chestnut Resistance to Pests, Diseases, and Climate Change) (Suggestion…..)
2. Add one sentence in the beginning or end clarifying the main aim of the review (In abstract section).
3. How does climate change specifically influence research priorities in chestnut biotechnology? Please add some detials.
4. Add a linking statement showing how classical breeding and advanced genomics interact (In abstract section).
5. Consider adding ‘genetic engineering’ or ‘breeding’ since these are major components (Keywords).
6. Consider condensing the historical and cultural background to maintain focus on the scientific context relevant to biotechnology and stress management.
7. Add a short bridging sentence that explains why discussing major diseases is essential for understanding the need for biotechnology.
8. You repeatedly mention poor drought adaptation of hybrids in southern areas; you could consolidate these points into a single explanatory paragraph to avoid redundancy.
9. Many examples come from Spain and Portugal; adding one or two more lines on advancements from Italy/France/US/Japan might give more international context.
10. Add brief concluding sentences at the end of each subsection to reinforce key takeaways.
11. Check numbering: section headers like “2.2.2.1.” should be consistent with journal formatting.
12. Consider adding subheadings or bullet points within long sections to improve readability.
13. Do you plan to add a future perspectives paragraph at the end of each major section?
14. Consider summarizing CRISPR/RNP results in a small comparative table (method, efficiency, challenges).
15. Add a brief statement linking cryopreservation with genetic editing workflows (e.g., long-term storage of edited lines).
16. Ensure consistent formatting of gene names (italic for genes, normal for proteins).
17. Many grammatical and language mistakes should be improved.
18. Please avoid symbols at the beginning of sentences.

Please arrange all the references according to the Journal format.

Author Response

PLEASE SEE FILE

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

This paper analyzes the main biotechnological strategies applied to the European chestnut (Castanea sativa) and presents future perspectives for the application of biotechnology in the recovery, improvement, and sustainability of chestnut in the face of current and future threats. Drawing on extensive data and literature, it holds significant academic value. However, the manuscript submitted appears to be a draft rather than the final version. Several issues require attention, and I look forward to reviewing the revised version from the authors.
1.The title specifies "European chestnut" (i.e., Castanea sativa), yet the main text extensively describes biotechnologies and pest/disease issues related to other chestnut species. Consider revising the title to ensure consistency.
2.The subsection heading "2.2.1.1. Genetic Basis of P. cinnamomi Resistance……" seems redundant. The content under Section 2.2.1 should be restructured with more refined subheadings for clarity.
3.Section 2.1 lists numerous technologies and procedures. As a review article, it should be more hierarchical, with effective connections between sections. Adding additional subheadings is recommended to enhance readability.
4.The authors focus primarily on descriptive summaries without emphasizing the logical relationships between topics. It is essential to synthesize existing findings, highlight controversies and research gaps, rather than merely listing technologies and results. Significant streamlining of the listed technologies and outcomes is advised.
5.The paper provides insufficient coverage of policies, promotion, and ethical considerations pertaining to transgenic plants.
6.The "Future Perspectives" section only mentions "integrating biotechnology with omics technologies." More specific, actionable research directions should be proposed.

Author Response

Please see file

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

The manuscript presents a comprehensive, timely, and well-structured review dedicated to the biotechnological approaches used for improving Castanea sativa in the context of major biotic (ink disease, blight, gall wasp) and abiotic stresses. The work is clearly written, rich in scientific detail, and supported by extensive literature. The manuscript is suitable, and its topic is highly relevant to forest biotechnology, conservation biology, and climate-resilient breeding.

However, while the review is scientifically strong, several improvements are necessary:

 

1/ The introduction should clearly state the added value of this review, especially since chestnut improvement has been reviewed recently.

 

2/ Please consider reducing historical details and focusing on problem statement and rationale.

 

3/ Large blocks of text, especially in Sections 2.2 and 2.2.2, contain excessive detail from primary research and could be more concise. A review should prioritize critical synthesis, knowledge gaps, and future challenges, not simply listing results.

 

4/ The manuscript does not discuss how the findings tie back to figures 1 and 2. Please add a discussion explaining how the “lab-to-field” pipeline (Figure 2) translates to practical breeding programs and which steps still lack technological optimization.

 

5/ The manuscript reviews each pathogen separately, but the introduction explicitly mentions combined stress under climate change. The review should include a dedicated subsection on combined P. cinnamomi × drought stress, heat waves, multi-stress phenotyping, and genomic selection models that incorporate both abiotic and biotic traits.

 

6/ In pest control, it would be appropriate to consider holistic strategies such as area-wide Integrated Pest Management (IPM) and other next-generation approaches: https://doi.org/10.1080/09670874.2024.2334230

 

7/ Please,include a forward-looking, visionary section to reinforce the review's value on regulatory challenges of CRISPR and transgenic chestnuts, socio-economic implications for European orchards, long-term eco-evolutionary risks, nursery adoption barriers, digital phenotyping and AI-guided selection techniques, and microbiome-based strategies (biocontrol consortia).

 

8/ A synthetic model illustrating C. sativa vs. C. crenata immune responses would be valuable.

 

9/ Please revise some typographical and grammatical issues, standardize units (°C, hours = h, etc.), and ensure all abbreviations are defined on first use.

 

10/ The reference list is generally extensive and well-selected; however, please ensure consistent formatting and adherence to the journal’s reference style throughout.

Author Response

Please see file.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

All the comments have been incorporated accordingly.

Author Response

Please see the attachment.

Reviewer 2 Report

Comments and Suggestions for Authors

This is an excellent review. The paper is rich in data and extensive in literature citations, boasting both academic value and practical guiding significance, with an overall outstanding performance. However, after the Introduction section in the main text, there is a lack of effective image illustrations. It is recommended to add figures in the section of "Molecular and Genomic Approaches" to demonstrate the Molecular Mechanisms of Castanea Defense.

Author Response

Please see the attachment

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

The authors have satisfactorily addressed the majority of the comments raised during the review process. The revisions have strengthened the manuscript, and the responses provided are appropriate and constructive. In its current form, the manuscript has improved substantially and could be considered for publication.

Comments on the Quality of English Language

improved

Author Response

This manuscript was edited for English language by the MDPI editing service.