Optimizing Microclonal Propagation of Red Currant Cultivars: The Role of Nutrient Media, Sterilizers, and LED Lighting in Plant Adaptation

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

This study is methodologically comprehensive, there are several areas where the clarity, contextualization, and practical relevance could be improved. The introduction effectively frames the significance of virus-free planting material in horticultural practices. However, it fails to highlight the novelty of the study sufficiently. The rationale for focusing on the selected red currant cultivars and the specific combinations of nutrient media and LED lighting is underdeveloped. Why were these cultivars chosen over others, and how do they represent broader trends or challenges in horticulture? Moreover, the manuscript does not adequately position its findings within the wider scope of current literature, particularly regarding the challenges of transitioning plants from in vitro to ex-vitro conditions.

Variations in light quality have been demonstrated to significantly influence plant morphology and stress responses, underscoring the role of precise spectral adjustments in enhancing overall plant performance (https://doi.org/10.3390/ijpb15030058). By linking these results with the existing literature, the manuscript reinforces the importance of optimizing light spectral combinations to advance controlled environment agriculture, in-vitro culture, and other propagation techniques.

The methodological section is detailed, clearly describing experimental conditions, including sterilization protocols and nutrient media formulations. However, the manuscript lacks justification for the choice of specific parameters, such as the timing of plant introduction into in vitro culture and the selected light spectra. For example, the exclusion of spring introduction as inefficient is noted, but the underlying physiological or environmental reasons are not fully elaborated. Additionally, while LED lighting is discussed extensively, the manuscript does not explore whether these specific spectral compositions are representative of commercially viable or scalable solutions for horticultural production. What do you think about it?

The results are rich with data, including survival rates, reproduction coefficients, and physiological parameters. However, the presentation is overly dense, requiring extensive cross-referencing between figures and tables to extract key findings. The study highlights the effects of nutrient media and LED lighting on various cultivars but stops short of connecting these observations to practical implications. For instance, while certain light spectra are shown to enhance pigment accumulation and plant vigor, the manuscript does not discuss the broader trade-offs involved in adopting these lighting conditions.

The discussion integrates the findings into the existing body of literature but remains largely descriptive. It emphasizes the benefits of specific nutrient media and LED lighting spectra but does not critically evaluate potential limitations, such as the restricted scope of cultivars or the lack of field validation. Additionally, the manuscript does not address how the findings could inform broader horticultural practices. What challenges might arise when implementing these protocols in commercial settings? Are there specific traits or genotypes in red currant cultivars that could be prioritized based on these findings?

The conclusion effectively summarizes the study's key findings but does not provide actionable recommendations for researchers or practitioners. While the study identifies effective nutrient media and LED spectra, it does not offer guidance on how these findings could be integrated into large-scale propagation systems. Furthermore, the proposed future research directions are too broad and lack specificity. What targeted studies or experiments could build upon these findings to enhance their applicability?

Clearly articulate the novelty of the study and the rationale behind the selection of cultivars, nutrient media, and LED lighting parameters. Expand the discussion to include practical implications and potential challenges for commercial adoption, such as cost-effectiveness and scalability. Address the limitations of the study more critically, including the restricted environmental conditions and the exclusive focus on a few cultivars. Consistency in uppercase and lowercase letters in the titles of references should be ensured. The style of the image looks rough, so I strongly recommend that you improve the quality. By addressing these areas, the manuscript could achieve greater clarity and practical relevance, making a more substantial contribution to the field of horticultural science.

Thank you.

Author Response

Dear Editor and Reviewers,

Thank you so much for the valuable comments provided to improve the manuscript.

We have thoroughly considered all the comments and are sincerely grateful.

Kindly see the below replies.

Thank you.

With respect,

 Dr. Olga Panfilova

Reviewer 1

1.This study is methodologically comprehensive, there are several areas where the clarity, contextualization, and practical relevance could be improved. The introduction effectively frames the significance of virus-free planting material in horticultural practices. However, it fails to highlight the novelty of the study sufficiently. The rationale for focusing on the selected red currant cultivars and the specific combinations of nutrient media and LED lighting is underdeveloped. Why were these cultivars chosen over others, and how do they represent broader trends or challenges in horticulture? Moreover, the manuscript does not adequately position its findings within the wider scope of current literature, particularly regarding the challenges of transitioning plants from in vitro to ex-vitro conditions.

-We add information and made changes in the text (Line 64-74, 77-92 red color)

2.Variations in light quality have been demonstrated to significantly influence plant morphology and stress responses, underscoring the role of precise spectral adjustments in enhancing overall plant performance (https://doi.org/10.3390/ijpb15030058). By linking these results with the existing literature, the manuscript reinforces the importance of optimizing light spectral combinations to advance controlled environment agriculture, in-vitro culture, and other propagation techniques.

- We add information and made changes in the text (Line 105-120, 124-126 red color)

3.The methodological section is detailed, clearly describing experimental conditions, including sterilization protocols and nutrient media formulations. However, the manuscript lacks justification for the choice of specific parameters, such as the timing of plant introduction into in vitro culture and the selected light spectra. For example, the exclusion of spring introduction as inefficient is noted, but the underlying physiological or environmental reasons are not fully elaborated

-We add information (Line 141-147 red color)

4.Additionally, while LED lighting is discussed extensively, the manuscript does not explore whether these specific spectral compositions are representative of commercially viable or scalable solutions for horticultural production. What do you think about it?

-We add this information to the Discussion (Line 473-488 red color)

5.The results are rich with data, including survival rates, reproduction coefficients, and physiological parameters. However, the presentation is overly dense, requiring extensive cross-referencing between figures and tables to extract key findings. The study highlights the effects of nutrient media and LED lighting on various cultivars but stops short of connecting these observations to practical implications. For instance, while certain light spectra are shown to enhance pigment accumulation and plant vigor, the manuscript does not discuss the broader trade-offs involved in adopting these lighting conditions

-We add information (Line 286-291, 295, 300-303, 344-347, 397-399 red color). In the Discussion we add information

6.The discussion integrates the findings into the existing body of literature but remains largely descriptive. It emphasizes the benefits of specific nutrient media and LED lighting spectra but does not critically evaluate potential limitations, such as the restricted scope of cultivars or the lack of field validation. Additionally, the manuscript does not address how the findings could inform broader horticultural practices. What challenges might arise when implementing these protocols in commercial settings?

-We add many information’s and made changes in the Discussion (Line 410-488, red, green and yellow colors)

7.Are there specific traits or genotypes in red currant cultivars that could be prioritized based on these findings?

- Our extensive research has identified variances in the micropropagation of Ribes spp. The primary distinctions observed in the propagation of redcurrants and blackcurrants pertain to the concentrations of BAP (6-benzylaminopurine). The optimal concentration for blackcurrants was identified as between 0.2 and 0.5 mg·L-1, while for red currants, effectiveness was noted at concentrations between 0.6 and 1.0 mg·L-1. These findings are consistent with those reported by Hautsalo, Juho, et al. (2017) (doi  10.17660/ActaHortic.2018.1224.28), Kukharchik, N.V. et al. 2013, 2016,  (Kukharchik N.V., Kolbanova E.V., Krasinskaya T.A., Malinovskaya A.M., Kastritskaya M.S., Tychinskaya L.Yu., Poleshko G.D. Mineral nutrition and morphogenesis of horticultural crops at in vitro cultivation. Fruit Growing. 2013;25(1):227-235.; Kukharchyk, N.; Kastritskaya, M.S.; Semenas, S.E.; Kolbanova, E.V.; Krasinskaya, T.A.; Volosevich, N.N.; Solovey, O.V.; Zmushko, A.A.; Bozhidai, T.N.; Rundia, A.P.; Malinovskaya, A.M. Reproduction of fruit and berry plants in vitro; Belarus science: Minsk, Belarus, 2016; рр. 72–81.).

There is no suitable general method for micropropagation available Ribes spp. for present day.

8.The conclusion effectively summarizes the study's key findings but does not provide actionable recommendations for researchers or practitioners. While the study identifies effective nutrient media and LED spectra, it does not offer guidance on how these findings could be integrated into large-scale propagation systems. Furthermore, the proposed future research directions are too broad and lack specificity. What targeted studies or experiments could build upon these findings to enhance their applicability?

-We add many information’s and made changes in the Conclusions (Line 490-292, 498, 502, 503-510  red color).

9.Clearly articulate the novelty of the study and the rationale behind the selection of cultivars, nutrient media, and LED lighting parameters. Expand the discussion to include practical implications and potential challenges for commercial adoption, such as cost-effectiveness and scalability. Address the limitations of the study more critically, including the restricted environmental conditions and the exclusive focus on a few cultivars. Consistency in uppercase and lowercase letters in the titles of references should be ensured. The style of the image looks rough, so I strongly recommend that you improve the quality. By addressing these areas, the manuscript could achieve greater clarity and practical relevance, making a more substantial contribution to the field of horticultural science.

- We add many information’s and made changes in the manuscript (Line 63-72, 104-119, 410-488 red color). The manuscript's style has been reviewed, and necessary modifications have been applied.   The literature is designed in accordance with the recommendations of journal (pp.15-18)

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

This manuscript investigated the effects of reproduction duration, nutrient medium composition, sterilizing agents, and LED illumination on in vitro propagation and growth characteristics of four red currant cultivars. The results indicate that MS medium supplemented with 1 mg/L 6-benzylaminopurine demonstrated optimal efficacy, establishing a preliminary protocol for red currant micropropagation. This manuscript has a relatively comprehensive research content and a reasonable experimental plan. Here are some comments. The following revisions are recommended:

1. Figure 1 should be relocated to supplementary materials.

2. For consistency with Line 160 (Russia), add country designation (China) in Line 184.

3. Define a,b,c in Figure 2; verify superscripts and subscripts. The M HgCl₂ treatment group lacks a lowercase letter. Re-examine statistical significance in PL H₂O₂ treatment. Include standard deviations in the table.

4. Incorporate standard deviations in Table 4.

5. Upload Appendix A and B as supplementary files (Line 466).

6. Verify statistical significance tests in Tables 6 and 7.

7. Include explanatory notes for RWUV-A in Table 7.

8. Enhance Figures 2, 3, and 4 by removing gridlines and adding standard deviations to tabulated data.

9. It is recommended to use professional drawing software instead of Microsoft Excel for drawing the figure.

10. Strengthen the discussion section with more comprehensive analysis of experimental data.

11. Include additional photographic documentation of plantlets under various treatments.

Author Response

Dear Editor and Reviewers,

Thank you so much for the valuable comments provided to improve the manuscript.

We have thoroughly considered all the comments and are sincerely grateful.

Kindly see the below replies.

Thank you.

With respect,

 Dr. Olga Panfilova

Reviewer 2

1. Figure 1 should be relocated to supplementary materials.

We removed the figure 1 from the text and transferred it to supplementary materials. (Line 514-515 Yellow color)

2. For consistency with Line 160 (Russia), add country designation (China) in Line 184.

We add information (Line 203 Yellow color)

3. Define a,b,c in Figure 2; verify superscripts and subscripts. The M HgCl2 treatment group lacks a lowercase letter. Re-examine statistical significance in PL H2O2 treatment. Include standard deviations in the table.

We made changes in the Figure  (Line 264-271 Yellow color)

4. Incorporate standard deviations in Table 4.

We add information in Table 4 (Line 319-322 Yellow color)

5. Upload Appendix A and B as supplementary files (Line 466).

We made changes in the Text (Line 294, 339, 346 Yellow color)

6. Verify statistical significance tests in Tables 6 and 7.

We made changes in the Tables 6 and 7. (Line 350-352, 365-371 Yellow color)

7. Include explanatory notes for RWUV-A in Table 7.

We add information in Table 7. (Line 365-371 Yellow color)

8. Enhance Figures 2, 3, and 4 by removing gridlines and adding standard deviations to tabulated data.
9. It is recommended to use professional drawing software instead of Microsoft Excel for drawing the figure.

We made changes in the Figures 2, 3, and 4 (Line 264-271, 387-394, 403-409 Yellow color)

10. Strengthen the discussion section with more comprehensive analysis of experimental data.

We made changes in the discussion section (Line 410-488 Yellow color)

11. Include additional photographic documentation of plantlets under various treatments.

We add photographic documentation in the manuscript (Line 514-522 Yellow color). I moved new photo graphics (Figure S2, Figure S3) to folder «Supplementary Materials».

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

Experience with many varieties and treatments. The values ​​of the results obtained should be provided in the abstract and in the description of the results. Explain why these varieties were studied. In the methodology, specify how many plants the measurements concerned, whether all of them, and how many repetitions of chlorophyll and carotenoid measurements there were. Why are the results of some varieties included in the tables and charts while others are omitted?  The description of the results needs to be improved. In the discussion, indicate which species the research of other authors concerned and what results were obtained.

Comments for author File: Comments.pdf

Author Response

Dear Editor and Reviewers,

Thank you so much for the valuable comments provided to improve the manuscript.

We have thoroughly considered all the comments and are sincerely grateful.

Kindly see the below replies.

Thank you.

With respect,

 Dr. Olga Panfilova

Reviewer 3

1.Experience with many varieties and treatments. The values ​​of the results obtained should be provided in the abstract and in the description of the results. Explain why these varieties were studied. 

-We add information (Line 64-73 red color)

2.In the methodology, specify how many plants the measurements concerned, whether all of them, and how many repetitions of chlorophyll and carotenoid measurements there were. Why are the results of some varieties included in the tables and charts while others are omitted? 

-We add information (Line 156-161, 211-215, 216, 228, 249, green color)

3.The description of the results needs to be improved.

-We add information and made changes in the text (Line 129-132, 157-162, 276-224, 296, 299-300, 305, 326, 338-339, 358, 360, 364, 373-375, 377 green color)

4.In the discussion, indicate which species the research of other authors concerned and what results were obtained

-We add information and made changes in the text (Line 411-420 green color, 420-443, 459-474 Yellow color, 445-458 red color)

-We add information in the Abstract (Line 1-25, 31-32 green color), and made changes in the Keywords

-We made changes in the References and in the text

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

Many figures lack the necessary clarity and precision to effectively convey the results. For instance, statistical annotations are either absent or insufficient in several figures, which undermines the reliability of the visualized data. The authors should consider including detailed statistical markers, such as "asterisks" or "p-values", to denote significance levels (especially Fig. 2 and 3). Additionally, the figures appear to be of a quality that is not up to standard for publication. A format that seems to combine figures and tables doesn't look good. For example, numerical data can be placed on top of each bar, and there is also a way to rotate the angle by 180 degrees. I strongly recommend the authors to improve the quality of the figures included in this manuscript.

The tables, while informative, often present an overload of data without adequate synthesis or focus on the most critical outcomes. Table 4 shows the results of the post-tests from "a" to "v". Many readers may find this very confusing. I think authors should seriously consider whether there is another way to express it. This makes it challenging for readers to extract meaningful conclusions. Streamlining the data presentation and emphasizing the most significant findings would enhance readability and impact.

The rationale for certain choices, such as specific wavelengths of LED light or the concentrations of growth regulators, remains underexplored. The authors must address why these parameters were chosen and how they align with broader trends or previous studies in plant micropropagation.

The results section is comprehensive but overly descriptive in several areas. The narrative often reiterates data from tables and figures without offering a critical analysis or broader interpretation. For instance, while the positive effects of RWUV-A lighting on physiological parameters are noted, the underlying mechanisms remain speculative. Furthermore, the discussion on the impact of nutrient media lacks depth, particularly regarding their scalability and cost-effectiveness in commercial applications.

The discussion section effectively integrates the study's findings with existing literature but remains largely descriptive. A more critical evaluation of the limitations, such as the exclusive focus on specific cultivars or the reliance on controlled conditions, would add depth to the manuscript. Additionally, the discussion could benefit from exploring the broader implications of the findings for commercial horticulture.

Thank you.

Author Response

Dear Reviewer,

Thanks for your valuable comments, we went through them and we tried to improve the quality of our manuscript.

Many figures lack the necessary clarity and precision to effectively convey the results. For instance, statistical annotations are either absent or insufficient in several figures, which undermines the reliability of the visualized data. The authors should consider including detailed statistical markers, such as "asterisks" or "p-values", to denote significance levels (especially Fig. 2 and 3). Additionally, the figures appear to be of a quality that is not up to standard for publication. A format that seems to combine figures and tables doesn't look good. For example, numerical data can be placed on top of each bar, and there is also a way to rotate the angle by 180 degrees. I strongly recommend the authors to improve the quality of the figures included in this manuscript.

- We improved the quality of the figures 3, 4 and add information (Line 280-284, 400-404, 417-422 blue color).

The tables, while informative, often present an overload of data without adequate synthesis or focus on the most critical outcomes. Table 4 shows the results of the post-tests from "a" to "v". Many readers may find this very confusing. I think authors should seriously consider whether there is another way to express it. This makes it challenging for readers to extract meaningful conclusions. Streamlining the data presentation and emphasizing the most significant findings would enhance readability and impact.

- We changed the table to Figure 2 and added the information (Line 325-333 blue color).

The rationale for certain choices, such as specific wavelengths of LED light or the concentrations of growth regulators, remains underexplored. The authors must address why these parameters were chosen and how they align with broader trends or previous studies in plant micropropagation.

-We added the information in the text (Line 96-99, 118-121 blue color) and added new references №35, 36

-The results section is comprehensive but overly descriptive in several areas. The narrative often reiterates data from tables and figures without offering a critical analysis or broader interpretation. For instance, while the positive effects of RWUV-A lighting on physiological parameters are noted, the underlying mechanisms remain speculative. Furthermore, the discussion on the impact of nutrient media lacks depth, particularly regarding their scalability and cost-effectiveness in commercial applications.

-We added the information in the text (Line 442-443, 451-455, 470-472 blue color) and added new references № 58, 60, 61, 71, 72. Moreover, the results section was corrected as recommended by other reviewers and the changes were approved.

- The discussion section effectively integrates the study's findings with existing literature but remains largely descriptive. A more critical evaluation of the limitations, such as the exclusive focus on specific cultivars or the reliance on controlled conditions, would add depth to the manuscript. Additionally, the discussion could benefit from exploring the broader implications of the findings for commercial horticulture.

-In the discussion, we focused a lot on our results and compared them with the results of other scientists. We also added new information ( Line 442-443, 451-455, 470-472, 502-504 blue color).

In addition, the discussion section was modified in accordance with recommendations from other reviewers and the changes were approved.

With respect,

Dr. Olga Panfilova

Reviewer 2 Report

Comments and Suggestions for Authors

The author has responded to all comments and made revisions to the manuscript.

Author Response

Dear Reviewer,

Thank you so much for reviewing my paper. 

With respect,

Dr. Olga Panfilova

Reviewer 3 Report

Comments and Suggestions for Authors

The abstract should include the most important results obtained on the influence of the studied factors on the propagation of selected red currant varieties. Minor errors were noticed in the publication, e.g. .3 should be 3.3, in the summary instead of redcurrant it should be red currant and similar errors. The authors took into account the remaining comments of the reviewer.

Author Response

Dear Reviewer,

Many thanks for your positive and constructive comments. We have edited the manuscript as follows and trust it improved considering focus and clarity.

The abstract should include the most important results obtained on the influence of the studied factors on the propagation of selected red currant varieties. Minor errors were noticed in the publication, e.g. .3 should be 3.3, in the summary instead of redcurrant it should be red currant and similar errors. The authors took into account the remaining comments of the reviewer.

-We add information in the Abstract (Line 20-25, 36-38 green color) and made changes in the text (Line 16, 70, 147, 226, 303, 317, 350, 354, 409, 412, 511, 518, 530, 532 537 Green color)

With respect,

Dr. Olga Panfilova