The Effects of High Temperature Stress and Its Mitigation Through the Application of Biostimulants in Controlled Environment Agriculture

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Dear editor

Thank you for the opportunity to review this manuscript. The role of biostimulants in alleviating abiotic stress in crops is a critical area of research. A great deal of previous work has focused on the application of biostimulants to address challenges like nitrogen deficiency and low temperatures. However, a systematic summary of how these compounds help plants respond to high-temperature stress has been lacking, which makes the topic of this paper particularly timely and significant. While the topic is excellent, I believe the manuscript requires further revision before it can be accepted for publication.

Title and introduction

My primary suggestion is to revise the title. There is considerable overlap between the terms 'Controlled Environment Agriculture' (CEA) and 'indoor farm'; in fact, many in the field would define indoor farming as a specific type of CEA (History of Controlled Environment Horticulture: Indoor Farming and Its Key Technologies (DOI: 10.21273/HORTSCI16159-21)). To address this, I recommend that the author begins the introduction by explicitly defining and distinguishing between these two concepts. If a clear distinction isn't central to the manuscript's thesis, it would be clearer and more accurate to simplify the title to include only 'CEA', which is generally understood to encompass all forms of facility-based agriculture.

Temperature Stress Effects

While the general effects of high temperature on plants are described, this information is quite fundamental. I would strongly recommend that the author narrows the focus to the specific impacts of high temperature on crop yield and quality in the context of CEA production systems. Such an analysis would be far more novel and would more effectively underscore the practical importance of applying biostimulants to address heat stress challenges unique to these advanced cultivation environments.

To achieve this, the author should detail the specific phenotypic, physiological, and biochemical changes that occur in crops under heat stress at different high-temperature ranges within CEA models. It would be particularly valuable to focus on representative CEA crops such as lettuce, tomato, cucumber, and pepper. Presenting these specific findings in a summary table would be an excellent way to organize this information for the reader.

Biostimulants

While the manuscript categorizes various biostimulants, it notably omits a discussion on humic substances (HS) regarding their role in mitigating high-temperature stress. Given that humic substances represent an important class of biostimulants with a significant body of research—especially concerning their application in CEA production systems—this is a considerable omission. I strongly recommend that the author adds a section to review the mechanisms by which HS help crops respond to heat stress, which would make this review more comprehensive.

Furthermore, there is a significant issue with the author's summary of the research on different biostimulants in Table 3. The information provided is incomplete, and the table's content requires optimization and adjustment.

First, the order of the biostimulant types in the table should be consistent with the order in which they are introduced in the main text. For example, the author first discusses the effects of seaweed extracts (SE) in the text, yet the table begins with results for amino acids.

Additionally, the application rate of biostimulants is critically important for crop growth; however, the author has omitted this data from the summary table. I believe the application dosages should be included.

Finally, for key data such as yield, the table should provide specific quantitative results, detailing exactly how much the yield was increased or decreased.

Conclusion

The "Conclusion" section currently reads more like a "Conclusion and Perspective". To enhance this summary, I strongly suggest that the author creates a schematic figure to illustrate the effects of biostimulants on horticultural crops under heat stress. This figure could visually summarize the impacts on different plant organs—such as the leaves, stems, roots, flowers, and fruits—which would help readers more intuitively understand how biostimulants assist crops in coping with high temperatures.

Regarding the future outlook, I agree with the author's general direction for future research, particularly the proposal to compare the potential of biostimulants in soil versus soilless systems; this is indeed an important research avenue. However, to better align this perspective with the manuscript's main theme, the author should provide a more in-depth discussion on future research directions specifically related to mitigating high-temperature stress. For example, what are the current deficiencies in the research on using biostimulants to help crops cope with high-temperature stress, and what are the main mechanisms that are still not clearly understood?

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript addresses a highly relevant and timely topic, given the increasing global adoption of Controlled Environment Agriculture (CEA) and the challenges posed by high-temperature stress. The focus on biostimulants as mitigation tools is well chosen and differentiates this work from broader reviews on CEA or biostimulants alone. The paper is generally well-structured, clearly written, and supported by extensive literature.

However, the manuscript suffers from repetition, lack of figures, and limited quantitative evidence to support claims. Additionally, many references are outdated (pre-2015), and recent omics-based and applied studies (2020–2025) are underrepresented. Tables are informative but could be restructured for better readability.

 

1. The abstract is clear but overly descriptive. Background details on population growth could be shortened. The abstract is missing specific quantitative outcomes (e.g., “biostimulants increased tomato fruit set by 86% under 31°C”). Please add at least one or two quantitative results to highlight novelty.
2. The transition from energy costs in CEA to the introduction of biostimulants is abrupt. So, please provide smoother logical flow and include concrete energy cost data (e.g., percentage of operating costs in vertical farming) from recent studies (2020–2025). (State of art)

3.  For the section that related to temperature stress effects, this section lacks quantitative crop-specific thresholds. For instance, tomato, cucumber, and lettuce show different heat tolerances. In addition, for cellular responses, ROS and antioxidant mechanisms are explained twice; it needs condensation. A schematic figure of ROS generation, UPR, and antioxidant responses is essential.

4.  For hormonal and molecular responses, please add figures illustrating hormone interactions with HSPs/HSFs. Update with more recent omics studies.

5.  For the effect of the temperature on crops, the repetitive discussion of “food quality and quantity compromised.” Include quantitative nutritional data (e.g., % reduction in carotenoids, polyphenols under heat stress).

6. For the Biostimulants section, the seaweed extracts were well-detailed but lacked a critical comparison of different species and extraction methods. Also, for chitin and chitosan there is some redundancy; add emerging research on nano-chitosan formulations. For the protein hydrolysates part, there is a good cultivar-specific discussion, but it is fragmented. Needs stronger linkage to metabolomic studies (2020–2024). In the inorganic compounds there is a lack of clear differentiation between foliar and root application methods. Expand on this comparison.

7. Please consider inserting a new figure. The manuscript lacks visual aids. At minimum, include a schematic of ROS signaling and antioxidant responses, a figure linking hormone pathways with HSP/HSF regulation, and a conceptual model showing how biostimulants mitigate heat stress under CEA.

    

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Author Response

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Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors Congratulations on the comprehensive review of the topic. I would like to request the addition of a brief subchapter (3.7) on the positive biostimulant effects of magnetic water treatment and water treated with very low-frequency electromagnetic resonance fields. I believe it adds to your work and makes it more robust. Sincerely

Author Response

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Author Response File: Author Response.pdf

Reviewer 4 Report

Comments and Suggestions for Authors

1. Title: “…mitigation in through the application…”. I suggest dropping the word “in” to improve the readability of the title.
2. The title is a little redundant. Indoor farming is classified within the broad definition of CEA according to the University of California Davis definition (https://caes.ucdavis.edu/research/initiative/controlled-environment-agriculture). I suggest dropping “and Indoor Farming” from the title.
3. Line 16-17, Abstract section: The authors wrote “…by reducing reliance on cooling systems and allowing 16 temperatures to rise above optimum.”. While this is a valid viewpoint, indoor/vertical farms might have energy rebate or some other agreement with the local electric company (based on my personal experience) thus saving the company money on electric bill. I suggest framing this in a way that includes power outage due to external factors such as calamities like typhoons, hurricanes, tornadoes, etc. Another viewpoint is if the HVACD system broke down or need repairs and there are no redundant systems available to maintain the temperature in the plant cultivation area.
4. Line 67-68: minor grammatical error: “…variety of crops commonly grown __ different CEA systems…”
5. Line 108-109: “Heat stress decreases the number of pollen grains developed and released, reducing pollen viability and germination rates.” While the idea is true, how the information is not logically presented. This sentence reads like the decrease in pollen development and release is what is causing the reduction in pollen viability and germination. The authors should clarify that heat stress is causing the impairment or arrest of pollen development, and that heat stress is also what is causing the reduction of viability and germination rate of those pollen grains that were able to develop to maturity.
6. Line 115: “…due to changes in in energy resource…” To word “to” is missing here.
7. Section 2.1 should include the effect of heat waves in leafy greens growth in indoor farms since most indoor farms are growing leafy greens like lettuce and culinary herbs.
8. Line 193: “limiting resulting in reduced productivity”. Revise this sentence to improve readability.
9. Line 209: this sentence is clunky and needs revision.
10. Sections 2.2 and 2.3 are a nice addition to this review paper but it takes away from the focus of this manuscript on the role of biostimulants in mitigating heat stress in indoor production. The cellular response to heat stress and the hormone connection are extensively reported elsewhere so it doesn’t need to be included in here. Either significantly cut back these sections or remove them completely to focus this manuscript.

Comments on the Quality of English Language

There are some sentences that have missing prepositions so I urge the authors to go through the text again to ensure these are corrected.

Author Response

Please see the attachment

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The author has made extensive revisions to the manuscript, and all previously mentioned issues have been successfully addressed. However, there are still a few minor problems that need to be resolved before publication. For instance, some issues with verb tense consistency were noted, and several spelling errors were found.

Furthermore, a key request from the last review was to replace all instances of "indoor farm" with "CEA". The author has not yet made this change in the concluding section of the paper. We ask the author to please update this as requested.

Author Response

Thank you for your feedback and your patience whilst we implemented the changes suggested. We have worked through the document and amended grammar to bring more consistency and clarity to the work, specifically focusing on the tenses as suggested.

The author has made extensive revisions to the manuscript, and all previously mentioned issues have been successfully addressed. However, there are still a few minor problems that need to be resolved before publication. For instance, some issues with verb tense consistency were noted, and several spelling errors were found.

Thank you for highlighting this, we have made further amendments to the manuscript to address the verb tense consistency and spelling errors.

Furthermore, a key request from the last review was to replace all instances of "indoor farm" with "CEA". The author has not yet made this change in the concluding section of the paper. We ask the author to please update this as requested.

Thank you for highlighting this again, we have now removed the term “indoor farm” from the manuscript completely.

Reviewer 2 Report

Comments and Suggestions for Authors

Dear authors, Please do the correction and highlighted 

  Comment 4: For the effect of the temperature on crops, the repetitive discussion of “food quality and quantity compromised.” Include quantitative nutritional data (e.g., % reduction in carotenoids, polyphenols under heat stress).

Comment 5: For the Biostimulants section, the seaweed extracts were well-detailed but lacked a critical comparison of different species and extraction methods. Also, for chitin and chitosan there is some redundancy; add emerging research on nano-chitosan formulations. For the protein hydrolysates part, there is a good cultivar-specific discussion, but it is fragmented. Needs stronger linkage to metabolomic studies (2020–2024). In the inorganic compounds there is a lack of clear differentiation between foliar and root application methods. Expand on this comparison.

Comment 6: Please consider inserting a new figure. The manuscript lacks visual aids. At minimum, include a schematic of ROS signaling and antioxidant responses, a figure linking hormone pathways with HSP/HSF regulation, and a conceptual model showing how biostimulants mitigate heat stress under CEA.

Author Response

Thank you for your feedback and your patience whilst we implemented the changes suggested.

For the effect of the temperature on crops, the repetitive discussion of “food quality and quantity compromised.” Include quantitative nutritional data (e.g., % reduction in carotenoids, polyphenols under heat stress).

Thank you for this feedback, we have now included some quantitative nutritional data as requested in section 2.4.

For the Biostimulants section, the seaweed extracts were well-detailed but lacked a critical comparison of different species and extraction methods.

We appreciate this input, and as such have included some discussion on the different species and extraction methods (see lines 707-711 and 721-727) into the SE section.

For the protein hydrolysates part, there is a good cultivar-specific discussion, but it is fragmented. Needs stronger linkage to metabolomic studies (2020–2024).

In the protein hydrolysate section, we have attempted to remove fragmentation further by restructuring the section. We have also discussed metabolomic studies in lines 937-939.

Also, for chitin and chitosan there is some redundancy; add emerging research on nano chitosan formulations.

We have addressed nano-chitosan formulations in lines 835 - 839.

In the inorganic compounds there is a lack of clear differentiation between foliar and root application methods. Expand on this comparison.

We have included a small section discussing the impacts of Silicon applications on soil in addition to the information in table 2, which can be found in lines 1086-1089. However, we have kept this brief as other sections do not largely have discussions regarding the differentiation between foliar and root applications to maintain consistency between the different sections.

Please consider inserting a new figure. The manuscript lacks visual aids. At minimum, include a schematic of ROS signaling and antioxidant responses, a figure linking hormone pathways with HSP/HSF regulation, and a conceptual model showing how biostimulants mitigate heat stress under CEA.

We appreciate this feedback and have created a new figure (figure 1) which can be found at line 305/306 which we hope will be satisfactory. It covers the cellular responses of heat stress and how these link with the core regulators of the molecular stress response pathways. Hormone responses are summarised in table 2, which summarises the key effects and mechanisms of hormone responses, with references to comprehensive reviews which summarise these pathways in lines 455-471. Figure 2 includes a conceptual model for how biostimulants mitigate heat stress, found in section 4.

We thank both the editorial team and the reviewers again for their time and patience with the team. We hope that this revisions will enhance the contribution of this paper to the literature available on both biostimulants and Controlled Environment Agriculture, and we await to hear the response regarding its publication in this special issue.

Yours sincerely,

Anna Gardiner-Piggott

On behalf of the authors