In Vitro Culture Strategies for the Conservation and Sustainable Use of Vanilla planifolia Andrews: A Comprehensive Review

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The purpose of this manuscript was to provide an overview of micropropagation of the vanilla orchid and options for explant types, culture media, hormones, and cultivation systems.  The introduction was particularly well written to provide historical and current context for vanilla cultivation.  It was very nice to see images of the plant itself, both in field cultivation and in the tissue culture container. 

 

I have just one larger suggestion for the manuscript.  It may be worth expanding a bit of context on micro-propagation for phytosanitary purposes.  Which are the main diseases/pathogens of concern for vanilla in this regard?  This approach may also be useful in phytosanitary distribution of plant materials to growers (either regionally or internationally).  Here, other crops could be discussed to illustrate the importance of this approach in general such as banana. 

 

A few minor changes would help polish the final manuscript.  Here are my suggestions.

 

1. A few areas of the text are missing citations, please add some to lines 66-70 (botanical description of vanilla), lines 75-80 (sites and climate of cultivation).
2. Please explain how cuttings are disinfected for planting and why (lines 115-116).
3. Is permission needed (or currently on hand) for Figure 3 panel D and Figure 4?
4. Please expand upon the source for artificial vanilla as this area would likely be of interest to readers.
5. Table 1 needs a few minor revisions to tidy up the formatting (may as well change them now before you get to the copy editor and proofs). If all concentrations are uM there is no need for that label in the cells themselves (the one example with a % could be labeled as it is currently shown).  One media type is listed as semisolido instead of semisolid, one is listed as medio, please make corrections as needed.  Citations 59 and 76 are in superscript.  Is fase (in the legend) meant to be phase? 
6. It may be more logical to list the Table 2 data by SIT type as the first column rather than by explant type. What does the dash (-) mean for the immersion frequency of the buds in the BIT system?  Citation 69 is in superscript here are well. 
7. Table 3 appears to be missing from the submitted version, please add it.
8. The section on mycorrhizal fungi is quite interesting. Please consider expanding this portion a bit to discuss if these endophytes survive the micropropagation process or if they need to be re-inoculated for optimal plant performance 

Author Response

Reviewer 1

The purpose of this manuscript was to provide an overview of micropropagation of the vanilla orchid and options for explant types, culture media, hormones, and cultivation systems.  The introduction was particularly well written to provide historical and current context for vanilla cultivation.  It was very nice to see images of the plant itself, both in field cultivation and in the tissue culture container. 

I have just one larger suggestion for the manuscript.  It may be worth expanding a bit of context on micro-propagation for phytosanitary purposes.  Which are the main diseases/pathogens of concern for vanilla in this regard?  This approach may also be useful in phytosanitary distribution of plant materials to growers (either regionally or internationally).  Here, other crops could be discussed to illustrate the importance of this approach in general such as banana. 

We are grateful for your insightful feedback. Accordingly, we have incorporated more detailed information concerning micropropagation and its role in phytosanitary management, particularly in lines 64–66 and 231–241, as well as in the concluding remarks.

A few minor changes would help polish the final manuscript.  Here are my suggestions.

1. A few areas of the text are missing citations, please add some to lines 66-70 (botanical description of vanilla), lines 75-80 (sites and climate of cultivation).

Thank you. The missing citations have been included in the revised manuscript.

 

2. Please explain how cuttings are disinfected for planting and why (lines 115-116).

Information regarding the disinfection of the cuttings has been included in lines 150–154.

 

3. Is permission needed (or currently on hand) for Figure 3 panel D and Figure 4?

The photographs are used with the authorization and courtesy of the respective authors, and the figure captions have been modified to reflect this.

 

4. Please expand upon the source for artificial vanilla as this area would likely be of interest to readers.

We appreciate the suggestion. We have added a brief mention of the market preference for natural vanilla, though we chose not to expand further to avoid discrediting synthetic vanilla production, which is outside the scope of this review.

 

5. Table 1 needs a few minor revisions to tidy up the formatting (may as well change them now before you get to the copy editor and proofs). If all concentrations are uM there is no need for that label in the cells themselves (the one example with a % could be labeled as it is currently shown).  One media type is listed as semisolido instead of semisolid, one is listed as medio, please make corrections as needed.  Citations 59 and 76 are in superscript.  Is fase (in the legend) meant to be phase? 

It may be more logical to list the Table 2 data by SIT type as the first column rather than by explant type. What does the dash (-) mean for the immersion frequency of the buds in the BIT system?  Citation 69 is in superscript here are well. 

We are grateful for these helpful comments. Tables 1 and 2 have been updated to reflect the suggested changes.

 

6. Table 3 appears to be missing from the submitted version, please add it.

Thank you. Table 3 has been included in the revised manuscript.

 

7.  The section on mycorrhizal fungi is quite interesting. Please consider expanding this portion a bit to discuss if these endophytes survive the micropropagation process or if they need to be re-inoculated for optimal plant performance 

We have updated the section 'Vanilla planifolia and orchid mycorrhizal fungi', rewriting several paragraphs to include new information (Lines 517–562).

Reviewer 2 Report

Comments and Suggestions for Authors

This paper discusses various biotechnological approaches for the in vitro propagation of vanilla, including micropropagation, temporary immersion systems, light conditions, genetic stability, and the role of orchid mycorrhizal fungi. While the overall structure of the article is well-organized, several aspects could be refined to enhance its academic rigor and clarity. The following suggestions are provided for further improvement：

1. Abstract: Should be more concise and highlight the review's value. The current abstract is overly descriptive. It is recommended to explicitly state the academic contributions and practical implications of this review at its conclusion.
2. Introduction: The elaboration of the scientific problem and rationale for the review needs strengthening. It is suggested to clearly articulate the core objectives and structural organization of the review at the end of the introduction.
3. Paper structure: The structure of the paper needs to be clearly redefined and outlined. The current text only lists "1. Introduction," lacking a clear delineation of subsequent sections.
4. Colloquial expressions: Excessive use of colloquial language should be avoided. For example,"helps to reduce costs" should be revised to "contributes to cost reduction."
5. Standardize terminology and abbreviations: Consistent use of "temporary immersion system (TIS)" throughout the text is recommended, avoiding the mixed use of "TISs." "Plant growth regulators (PGRs)" should be abbreviated after its first full mention.
6. Figure and table captions: Captions should be self-explanatory. It is advised to provide more detailed descriptions for Figures 1, 2, etc., including sample sources, treatment conditions, and statistical information.
7. Optimize table design: For Table 1, the "Shoots per explant" column should adopt the "Mean ± SD" format, with the sample size indicated. Consider merging tables with similar content to enhance the readability of the paper.
8. Provide source information for figures and tables: All images must cite their source (e.g., "Adapted from…" or "Original data") in accordance with academic publishing standards.
9. Strengthen comparative analysis and identify research gaps: When discussing sections such as TIS and light quality regulation, contradictory findings across studies or existing knowledge gaps should be explicitly pointed out.
10. Emphasize applicability assessment over mere technical description: When comparing different propagation systems (e.g., semi-solid media vs. TIS), a systematic analysis of their advantages and disadvantages regarding cost, scalability, and genetic stability should be provided.
11. Deepen the mechanistic discussion on mycorrhizal Fungi: The current description focuses on phenomena. It is recommended to supplement how mycorrhizal fungi influence vanilla growth and stress resistance through signaling pathways or metabolic regulation.
12. Clarify technical limitations and challenges, proposing specific future directions: The conclusion or discussion section should address common limitations of current in vitro propagation techniques, such as somaclonal variation, high costs, and low transplantation survival rates, and propose specific future research directions accordingly.
13. Standardize units and numerical formats: Consistent use of "μM" or "μmol/L" throughout the text is necessary. 
14. Check reference format consistency: Ensure uniformity in journal title abbreviations, author name formats, and the accessibility of DOI links within the references.
15. Add summary paragraphs at appropriate junctures: It is suggested to include brief summaries at the end of each major section to help readers grasp the core message of that part.

Comments on the Quality of English Language

The language style in certain sections lacks formal academic rigor, with some expressions being overly colloquial. It is recommended that the text undergo native-level polishing to enhance its academic tone and fluency.

Author Response

Reviewer 2

We are grateful for the reviewer's time and for the valuable comments, suggestions, and corrections provided. Our point-by-point responses to each of them are presented below.

This paper discusses various biotechnological approaches for the in vitro propagation of vanilla, including micropropagation, temporary immersion systems, light conditions, genetic stability, and the role of orchid mycorrhizal fungi. While the overall structure of the article is well-organized, several aspects could be refined to enhance its academic rigor and clarity. The following suggestions are provided for further improvement:

1. Abstract: Should be more concise and highlight the review's value. The current abstract is overly descriptive. It is recommended to explicitly state the academic contributions and practical implications of this review at its conclusion. Response: We appreciate your insightful comments. The abstract has been rewritten to be more concise and to better highlight the key contributions of this review.

2. Introduction: The elaboration of the scientific problem and rationale for the review needs strengthening. It is suggested to clearly articulate the core objectives and structural organization of the review at the end of the introduction. Response: Thank you once more. Following the suggestions of the reviewers, the introduction has been updated to better reflect the article’s objective and structure.

3. Paper structure: The structure of the paper needs to be clearly redefined and outlined. The current text only lists "1. Introduction," lacking a clear delineation of subsequent sections. Response: We have restructured the article and numbered the sections for better organization.

4. Colloquial expressions: Excessive use of colloquial language should be avoided. For example, "helps to reduce costs" should be revised to "contributes to cost reduction." Response: The manuscript was reviewed to remove informal language and improve the overall writing style.

5. Standardize terminology and abbreviations: Consistent use of "temporary immersion system (TIS)" throughout the text is recommended, avoiding the mixed use of "TISs." "Plant growth regulators (PGRs)" should be abbreviated after its first full mention. Response: The pertinent changes have been made throughout the manuscript.

6. Figure and table captions: Captions should be self-explanatory. It is advised to provide more detailed descriptions for Figures 1, 2, etc., including sample sources, treatment conditions, and statistical information. Response: Figure captions and table headings have been revised to provide the reader with more detailed information.

7. Optimize table design: For Table 1, the "Shoots per explant" column should adopt the "Mean ± SD" format, with the sample size indicated. Consider merging tables with similar content to enhance the readability of the paper. Response: Tables 1 and 2 have been revised in accordance with the comments from all reviewers.

8. Provide source information for figures and tables: All images must cite their source (e.g., "Adapted from…" or "Original data") in accordance with academic publishing standards. Response: Each figure caption now specifies when photographs were provided courtesy of their respective authors. In other cases, we have identified the co-author responsible for designing the image and providing the photographs.

9. Strengthen comparative analysis and identify research gaps: When discussing sections such as TIS and light quality regulation, contradictory findings across studies or existing knowledge gaps should be explicitly pointed out. Response: The content of section '2. Stage II: Propagule multiplication' was expanded to include further relevant details that better reflect the aim of the review article.

10. Emphasize applicability assessment over mere technical description: When comparing different propagation systems (e.g., semi-solid media vs. TIS), a systematic analysis of their advantages and disadvantages regarding cost, scalability, and genetic stability should be provided. Response: We have refined the corresponding sections to ensure the information is clearer and more prominent, while avoiding any significant increase in text length.

11. Deepen the mechanistic discussion on mycorrhizal Fungi: The current description focuses on phenomena. It is recommended to supplement how mycorrhizal fungi influence vanilla growth and stress resistance through signaling pathways or metabolic regulation. Response: Thank you for your suggestion. We have updated Section 4 with pertinent information; however, we chose not to go into exhaustive detail, as we believe this topic merits a separate, more comprehensive review.

12. Clarify technical limitations and challenges, proposing specific future directions: The conclusion or discussion section should address common limitations of current in vitro propagation techniques, such as somaclonal variation, high costs, and low transplantation survival rates, and propose specific future research directions accordingly.Response: Revisions were made throughout the manuscript, and Section 5, 'Conclusions and Perspectives,' has been rewritten to incorporate more pertinent and relevant information.

13. Standardize units and numerical formats: Consistent use of "μM" or "μmol/L" throughout the text is necessary.Response: Thank you. The pertinent changes have been made.

14. Check reference format consistency: Ensure uniformity in journal title abbreviations, author name formats, and the accessibility of DOI links within the references. Response: Thank you. The pertinent changes have been made.

Reviewer 3 Report

Comments and Suggestions for Authors

Comments for Authors:

The manuscript entitled “Biotechnological Approaches for the in vitro Propagation of Vanilla planifolia Andrews: A Review” addresses an important topic of both scientific and commercial relevance. The authors compile recent literature on in vitro propagation techniques, culture systems, genetic stability, acclimatization, and mycorrhizal associations in V. planifolia. The review is generally well organized and provides useful information for researchers and practitioners working on vanilla biotechnology. However, improvements are required in language quality, consistency, and clarity of presentation. Several grammatical errors, formatting issues, and unclear sentences, particularly in the abstract, should be corrected to enhance readability and professional standards.

Specific comments:    

1. Line 15# Replace with “has significant commercial importance worldwide” for academic tone.
2. Line 18# revision: “the high labor costs associated with fruit harvesting”.
3. Can the authors revise the abstract to improve its clarity, grammar, and overall readability?
4. The Introduction provides a thorough background on the origin, biology, cultivation systems, pollination, curing process, and economic importance of Vanilla planifolia. However, the section is overly descriptive for a review focused on biotechnological and in vitro propagation approaches. The Introduction would benefit from improved conciseness, better focus, and a clearer transition toward the core topic of in vitro culture. Additionally, several grammatical, formatting, and stylistic issues should be addressed to enhance readability.
5. Line 186-194# Suggestion: Cite the correct spelling of Gottlieb Haberlandt (Line 189).
6. Standardise units (μm, °C, %, mm).
7. Avoid excessive semicolons; use commas or full stops where appropriate.
8. Can the transition between conventional cultivation challenges and the need for in vitro culture be made clearer and more concise?
9. Minor language, formatting, and clarity issues are also present and should be addressed.
10. Avoid long sentences exceeding 3–4 clauses.
11. Can the authors further condense the micropropagation section to emphasise key biotechnological advances and reduce repetition?
12. Many grammatical and language mistakes should be improved.
13. Please avoid symbols at the beginning of sentences.
14. Please arrange all the references according to the Journal format.

Author Response

Reviewer 3

We are grateful for the reviewer's time and for the valuable comments, suggestions, and corrections provided. Our point-by-point responses to each of them are presented below.

The manuscript entitled “Biotechnological Approaches for the in vitro Propagation of Vanilla planifolia Andrews: A Review” addresses an important topic of both scientific and commercial relevance. The authors compile recent literature on in vitro propagation techniques, culture systems, genetic stability, acclimatization, and mycorrhizal associations in V. planifolia. The review is generally well organized and provides useful information for researchers and practitioners working on vanilla biotechnology. However, improvements are required in language quality, consistency, and clarity of presentation. Several grammatical errors, formatting issues, and unclear sentences, particularly in the abstract, should be corrected to enhance readability and professional standards.

We appreciate your kind and insightful comments. Several sections of the manuscript, including the abstract, introduction, and conclusions and perspectives, have been revised and updated accordingly.

Specific comments:    

1. Line 15# Replace with “has significant commercial importance worldwide” for academic tone.

Response: Thank you. The pertinent changes have been made.

 

2. Line 18# revision: “the high labor costs associated with fruit harvesting”.

Response: Thank you. The pertinent changes have been made.

 

3. Can the authors revise the abstract to improve its clarity, grammar, and overall readability?

Response: Thank you. We have revised the abstract to incorporate the feedback and suggestions from all the reviewers.

4. The Introduction provides a thorough background on the origin, biology, cultivation systems, pollination, curing process, and economic importance of Vanilla planifolia. However, the section is overly descriptive for a review focused on biotechnological and in vitro propagation approaches. The Introduction would benefit from improved conciseness, better focus, and a clearer transition toward the core topic of in vitro culture. Additionally, several grammatical, formatting, and stylistic issues should be addressed to enhance readability.

Response: Thank you. The introduction has been rewritten to address the comments and suggestions from all reviewers.

 

5. Line 186-194# Suggestion: Cite the correct spelling of Gottlieb Haberlandt (Line 189).

Response: Corrected

6. Standardise units (μm, °C, %, mm).

Response: Corrected

7. Avoid excessive semicolons; use commas or full stops where appropriate.

Response: Corrected

8. Can the transition between conventional cultivation challenges and the need for in vitro culture be made clearer and more concise?

Response: We have modified the manuscript to improve clarity on this and other points.

9. Minor language, formatting, and clarity issues are also present and should be addressed.

Response: Revised and corrected accordingly.

10. Avoid long sentences exceeding 3–4 clauses.

Response: Revised and corrected accordingly.

11. Can the authors further condense the micropropagation section to emphasise key biotechnological advances and reduce repetition?

Response: Thank you for your comment. We have modified the introduction to clarify the article's structure and highlight micropropagation as its core theme. Additionally, all points raised by the four reviewers have been addressed.

12. Many grammatical and language mistakes should be improved.

Response: The text has been revised and corrected.

13. Please avoid symbols at the beginning of sentences.

Response: Corrected

14. Please arrange all the references according to the Journal format.

Response: The references has been revised and corrected.

Reviewer 4 Report

Comments and Suggestions for Authors

A critical evaluation of the limitations of current protocols could enrich the article, which primarily consists of a compilation of studies.
It is recommended that the Conclusions and Perspectives section be strengthened with concrete proposals and reflections on scalability and technology transfer.

Review and standardize all references according to the journal's style. Ensure that each entry includes the year and DOI (when available), and that there are no duplicates in the numbering.

 

Comments on the Quality of English Language

Although the English is generally clear, there are minor errors in writing and punctuation (e.g., spaces before commas, inconsistent use of "and" vs. "&"). Professional proofreading of scientific English is recommended before publication.

 

Author Response

Reviewer 4

We sincerely thank the reviewer for their time and for the constructive feedback provided. Detailed, point-by-point responses to each of the comments are presented below.

A critical evaluation of the limitations of current protocols could enrich the article, which primarily consists of a compilation of studies.
It is recommended that the Conclusions and Perspectives section be strengthened with concrete proposals and reflections on scalability and technology transfer.

Response: Revisions were made throughout the manuscript, and Section 5, 'Conclusions and Perspectives,' has been rewritten to incorporate more pertinent and relevant information.

Review and standardize all references according to the journal's style. Ensure that each entry includes the year and DOI (when available), and that there are no duplicates in the numbering.

Response: The references has been revised and corrected.

Although the English is generally clear, there are minor errors in writing and punctuation (e.g., spaces before commas, inconsistent use of "and" vs. "&"). Professional proofreading of scientific English is recommended before publication.

Response: We appreciate your observations, all of which have been addressed. The introduction, several sections of the text, and the conclusions and perspectives have been modified and strengthened. The manuscript has undergone a thorough English language revision to ensure consistency in punctuation and style.

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

The purpose of this review is to provide an overview In Vitro Culture Strategies for the Conservation and Sustainable Use of Vanilla planifolia Andrews. However, there are too many descriptions of the basic characteristics of Vanilla planifolia, and it is recommended to appropriately reduce this part of the content.

Comments on the Quality of English Language

The language style in certain sections lacks formal academic rigor, with some expressions being overly colloquial. It is recommended that the text undergo native-level polishing to enhance its academic tone and fluency.

Author Response

We would like to thank the reviewers for their thoughtful comments and for the time invested in reviewing the revised version of our manuscript. We have addressed each of the observations to further improve the quality and academic rigor of the article.

1. The purpose of this review is to provide an overview In Vitro Culture Strategies for the Conservation and Sustainable Use of Vanilla planifolia Andrews. However, there are too many descriptions of the basic characteristics of Vanilla planifolia, and it is recommended to appropriately reduce this part of the content.

Answer: Certain sentences and paragraphs have been condensed to preserve clarity, whilst emphasizing the importance of establishing the background of the current vanilla situation, its conservation status, and, importantly, laying the foundation for understanding the necessity of exploring alternatives such as in vitro cultivation for mass propagation, conservation, and sustainable utilization.

Similarly, this section of the article was expanded based on the observations and recommendations from the four reviewers during the initial review process.  

 

2. The language style in certain sections lacks formal academic rigor, with some expressions being overly colloquial. It is recommended that the text undergo native-level polishing to enhance its academic tone and fluency.

 

Answer: To ensure that the article meets the journal's formal academic rigor and style, we rely on MDPI's expert editors, who provide professional language editing through their service.