Proline-Rich Specific Yeast Derivatives Enhance Grapevine (Vitis vinifera L.) Water Status and Enable Reduced Irrigation Volumes

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The article is interesting and presents relevant findings; however, it is based on only one growing season, and there are some issues regarding the naming and definition of treatments that are not entirely accurate. Although this may limit the robustness of the study, the results are novel, and the topic is of clear scientific interest. I think the study would be strengthened by including a correlation matrix to explore which physiological, physicochemical, or biochemical variables are most closely associated with proline content. I also recommend improving the methodology, particularly by providing a clearer explanation of the experimental design and treatment descriptions. The introduction should be refined, and a hypothesis explicitly stated based on the study objectives. Finally, the format of units and figures should be revised to comply with the journal’s author guidelines.

 

Abstract

The abstract is interesting but should better emphasize the study’s novelty and key findings. It currently reads as a general overview rather than a synthesis of results. I recommend highlighting how proline accumulation functions as an anti-stress mechanism in grapevines, including quantitative or physiological evidence supporting this role. The conclusion should stress the novel contribution and practical implications, such as using proline metabolism as a biomarker or management tool for stress resilience. A more result-focused and mechanistic tone would greatly enhance the impact and clarity of the abstract.

 

L10-11: I suggest improving this sentence for better understanding.

L13-16: I suggest explaining better or rewrite this phrase.

L18-30: I suggest improving the writing of these results avoiding the use of acronyms for better understanding.

 

Introduction

I suggest to deep introduction section in order to understand more in deep the role of proline biosynthesis in the enhance of abiotic stress in grapevines. In addition, I suggest reviewing the following papers for improve the section:

 

1)https://doi.org/10.1016/j.scienta.2008.07.031

2)https://onlinelibrary.wiley.com/doi/full/10.1111/ajgw.12523
3)https://www.sciencedirect.com/science/article/pii/S0304423822004940

L58-63: It would be good to add a hypothesis on this point.

 

Material and methods

I suggest to improve the treatments to an accurate terminology in defining water stress. The classification of treatments as “moderate” or “severe” water stress must be supported by quantitative physiological indicators, such as leaf or stem water potential, relative water content, or stomatal conductance. These parameters provide an objective measure of plant water status and ensure that stress levels are not arbitrarily assigned based solely on irrigation volume or visual symptoms. Without such measurements, it becomes difficult to compare results across studies or to interpret the physiological responses of grapevines with confidence. Therefore, redefining the treatments using measured data, or adopting neutral terminology (e.g., “low irrigation” or “deficit irrigation”), would strengthen the methodology of the study.

 

L67: The name of the specie must be written in italic.

L67-78: I suggest inserting a picture of the field trial and the experimental design for better understanding. I suggest also to add the number of replications by treatments in the trial and a better description of the experimental design.

L79-89: Why the treatments were called “moderate or severe water stress”? I suggest changing the names of treatments since there is no measurements of stem or leaf water potential to define the different levels of stress.

L79-89: Why you did not measure leaf or stem water potential? I think this parameter is of wide importance if you performed treatments with different amount of water applied to vines.

L90-94: This is not part of this section. I suggest to remove or replace by another sentence.

L99: The specie must be written in italic.

 

Results

The results section should be improved by including the full description of each acronym the first time it appears. In addition, some abbreviations are not clearly defined, such as CO₂ assimilation in certain parts of the text. It should also be specified whether the differences observed are statistically significant or not. Please revise and clarify these aspects for greater precision and readability. In addition, authors should uniform the units in whole document according to the journal guide of authors.

 

L222: Replace “A” by “A_N” or another term.

L283: I suggest replacing the term “proline abundance” by another. In this paragraph, the unit is showed in a different format than the previous paragraph, please, uniform it.

L314: Please, improve the format of tables in whole document according to the guide for authors.

 

Discussion

L330-333: This information does not contribute to the discussion and it should be deleted.

L335: Please, avoid the redundance in the term “consistent”.

L336-339: I think the most important is to mention if the treatments affected water potential.

L339: The decrease in water amount applied to vines could not be related to a severe water stress. Please, define it in the material and method section.

L330-352: This paragraph is a result description with scarcely discussion. I suggest to improve the deep of the discussion.

L354-370: How can the authors demonstrate that proline plays an active role in regulating the plant’s water status if this variable was not measured? Furthermore, since the applied product contains inactivated yeast cells, how can the authors distinguish between the specific effect of proline and the elicitor effect of the yeast components in this experiment? I recommend improving the discussion to address these points more clearly and to better substantiate the proposed physiological mechanisms.

L375-376: Please, describe the acronyms used in this phrase mainly, WUE and T.

L404-442: Please add more references according to the study of the effect of proline foliar applications on grape secondary metabolites. There are more studies published rather than the same research group.

 

Conclusion

I suggest to improve this section according to the results obtained and not the suggestion that can be obtained from them.

 

References

Please, the authors must check the references since there is some format mistakes.

Author Response

Dear Editor and reviewers,

Thank you very much for the time and effort you dedicated to reviewing our manuscript. We have carefully considered all the comments provided, and in this resubmitted version, we did our best to address all your suggestions. We believe the manuscript has now been significantly improved. In the attached document, our replies are reported in blue, and all changes have been implemented in the revised manuscript using the MS Word Track Changes feature. Line references correspond to those in the marked version of the revised manuscript.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Based on which parameters you decided to set up the experiment this way. I mean the connection between ET and water stress. Also, since you've been finding levels of water stress why haven't you been finding stress enzymes like SOD,POX, CAT and APOX.

Author Response

Dear Editor and reviewers,

Thank you very much for the time and effort you dedicated to reviewing our manuscript. We have carefully considered all the comments provided, and in this resubmitted version, we did our best to address all your suggestions. We believe the manuscript has now been significantly improved. In the attached document, our replies are reported in blue, and all changes have been implemented in the revised manuscript using the MS Word Track Changes feature. Line references correspond to those in the marked version of the revised manuscript.

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

The investigation of proline-rich yeast derivatives (SYD) as a priming strategy to enhance grapevine water status and explore water-saving potential is innovative and holds practical significance. The experimental design is generally sound, incorporating a gradient from full irrigation to severe stress with appropriate treatment and control groups, enabling an effective evaluation of SYD under varying water regimes.

1. A notable limitation is the absence of a 'WS2-C' group (40% ET irrigation without SYD application). This omission makes it difficult to definitively attribute the effects observed under severe stress (WS2) solely to SYD, as they could be confounded by the plant's intrinsic stress response to extreme water deficit. While the comparison between WS2-T and WS1-C offers some insight, it is not a direct control.

The 'Materials and Methods' section should explicitly state the reason for omitting the WS2-C group (e.g., resource constraints) and acknowledge this as a study limitation.

The 'Discussion' must interpret the WS2-T results more cautiously. It should be clearly stated that, lacking direct control, the WS2-T performance likely reflects a combination of SYD's positive effects and the impact of severe stress itself. Interpretation should be carefully reasoned based on existing data (e.g., similar proline accumulation and H₂O₂ levels in WS2-T and WS1-T), avoiding direct attribution.

2. The results from Figure 4 (WS1-T exhibiting higher photosynthesis at higher stomatal conductance) provide strong evidence for a 'direct effect' of SYD. This point should be emphasized more prominently in the 'Discussion.' Consider a separate paragraph explaining how this demonstrates that SYD's benefits extend beyond merely improving water potential, potentially involving protection of the photosynthetic apparatus or enhanced biochemical efficiency.
3. Line 31, The applications of proline-rich SYD therefore improved the vine physiological and productive performance, effectively compensating for modulations in irrigation. The term 'modulations' is imprecise, as SYD compensates for the deficit caused by irrigation reduction, not the 'modulations' themselves. Suggested Revision: '... effectively mitigating the negative impacts of reduced irrigation.' or '... thereby compensating for the water deficit.'
4. Line 448, 'WS2-T vines, receiving half the water of WS1 vines, demonstrated identical physiology, yield, and fruit composition to WS1-C.' The word 'identical' is too absolute. Data in Table 1 show that WS2-T yield and berry weight were significantly lower than those of WS1-C, and physiological parameters (e.g., gₛ and A) also differed at some stages. Suggested Revision: '... WS2-T vines demonstrated broadly comparable physiology and fruit composition to WS1-C, despite a significant reduction in yield components.' This aligns better with the presented data.
5. Keywords:'Biosimulants' 应为 'Biostimulants'。
6. Line 216, 'WS1-T midday leaf Ψ was higher than WS1-C Ψ...' This phrasing is technically correct (less negative Ψ indicates better water status), but to avoid potential ambiguity, consistently using 'WS1-T exhibited a less negative midday leaf Ψ than WS1-C' is recommended.
7. Introduction: Consider briefly mentioning whether this proline-rich SYD is cost-effective, addressing its practical applicability for growers.
8. The fonts in the Figures are inconsistent. Please modify.
9. It is necessary to introduce how much ET is defined as deficit irrigation in the introduction or the material method section.

Author Response

Dear Editor and reviewers,

Thank you very much for the time and effort you dedicated to reviewing our manuscript. We have carefully considered all the comments provided, and in this resubmitted version, we did our best to address all your suggestions. We believe the manuscript has now been significantly improved. In the attached document, our replies are reported in blue, and all changes have been implemented in the revised manuscript using the MS Word Track Changes feature. Line references correspond to those in the marked version of the revised manuscript.

Author Response File: Author Response.pdf

Round 2

Reviewer 3 Report

Comments and Suggestions for Authors In the introduction part, the author describes the innovation of the research more clearly and makes a detailed supplement to the material method. The results are clearer, and the discussion has a certain depth, but the fonts in Fig. 6 are inconsistent and can be modified.    

Author Response

Thank you, Figure 6 is now uniform in fonts and style.