A Quantitative Index for Evaluating Maize Leaf Wilting and Its Sustainable Application in Drought Resistance Screening

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Article “Development of a High-Throughput Method for Assessing Leaf Wilting and Its Application in Maize Drought Resistance” has been planned and performed in nice way. Data generates is also fine with respect to the study finding requires.

 

But there are many mistakes in writing and some missing according to the journal.

 

My detailed comments are as follows.

 

1.     L48, “CO2” should be “CO2“

2.     Should check all the citation of references through the article. Like L69, Sirault et al. (2015)？？？  L73，Rascio et al. (2020)  L241, Sytar et al. (2018). L261,279…

3.     L94, 10-35 °C, there should not be space between the numbers and “°C”.

4.     All the references should check carefully according to the journal requirements.

5.     Research innovation points could be further clarified. The article abstract mentions the development of new methods, but can further emphasize their innovation and comparison with existing technologies.

6.     The discussion of the results can be in depth. The discussion section has already provided some analysis, but it can be further expanded.

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors This manuscript is a great start of a good study. The goal of the presented study is to develop a low cost low tech method to quantify wilting which indicates drought stress. This approach is straightforward, applicable and is interesting to a great audience. The study is well design and data collection was good for the most part. However, my major concern is the lack of statistical analyses in the study. The data are plot in graphs but were appropriately analyzed. Also, the Materials and Methods section of this manuscript is not clear, and in many cased conflicts with data presented in Results. More details and some other minor edits are listed below. Line 47-49: not sure if leaf wilting and curling lead to stomata closure.. Line 59: add citation. Line 94: 'precipitation of 50-70 mm', is this annual precipitation? Line 99-102: I suggest you specify which lines are drought sensitive and which are drought resistant. Line 109: The drought treatments 'normal', 'moderate' and 'severe' need to be more specific. Later in the paper these treatment was mentioned as 0%, 50% and 100% irrigation. This description doesn't match with Table 1. Legend of Table 1 needs more details. Line 112-120: This paragraph is very vague. Line 114 'for a total of 7-8 measurements' does it mean 7-8 time points through the entire growing period? But in Figure 5 there are 10 time points in 2020 and 9 time points in 2021. Line116 'A total of four measurements were taken' mean the four growing stages? This contradicts Line 114. Line 118 cab be changed to 'every 2 hours in 2020 and every 3 hours in 2021' for clarity. Line 119 'Agronomic traits' and 'conventional methods' need to be specified. This whole paragraph needs a lot more details and better clarity. Line 145: what is 'self-developed software'? Since this paper is about the method for monitoring wilting , I suggest you provide bit more details on this software. Line 152: I don't think this should be call 'wilting ratio', green area as mentioned previously is a better term. Line 165: period Line 166-169: need statistics. Figure 4: the mean Visual Green Area is the mean values of 100 inbred lines * 2 reps? So n= 200? All your tables and graphs need more details in the legends. I also suggest that you add temperature and solar radiation data to the graph to present the trend. You can also add a regression analysis or correlation to show that the leaf curling tracks air temperature and solar radiation. Line 192-193: 'there was a strong correlation between leaf wilt and the water sensitive period', I think if you state that there is a strong correlation you need a correlation test and p and r (greater than 0.6) values to support it. But I think here you can do an ANOVA to show Wr were different between different time points and test correlation between Wr and temperature (which you said are not correlated). Figure 5: The legend again needs more details, e.g. n = 200 (I think?), add description of red arrows point at flowering stage. Also I suggest you adding temperature to the data, as well as marking the four different stages to this graph. Line 207: the differences among varieties are small Figure 6: need statistics, since you mentioned that difference among varieties are small under 100% and 50% irrigation and the variation were greater under 0% irrigation, I suggest you use ANOVA to test the difference among lines. And did you not collect the yield data of there 10 lines? Figure 7: If you added trend lines here, you need to add p and r/r2 values to the graphs, and preferably add regression equations too, especially in Line 227 you mentioned 'there is a close relationship between wilting ratios, yield, and drought resistance...', you need to present those statistics. You also need to add A, B,C and D to label the sub-graphs. The y-axis titles of the left graphs need correction. You need to define drought resistance factor/coefficient, this is a new term that was not mentioned in previous text. And Figure 7 you used average wilting ratio, since you mentioned previously that flowering stage is the crucial for monitoring leaf wilting, so why not use Wr at flowering stage?     Discussion: I suggest that you add details on cost of your equipment especially this study focuses on developing a low cost low tech and high-throughput method. I like that in Line 134-136 you showed how efficient this system can be. So I recommend adding a bit more discussion on the low cost feature of your system. Comments on the Quality of English Language

Good

Author Response

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Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

The manuscript presents an optical camera-based method for quantitatively assessing the wilting status of maize in field conditions. Its basic technique principle is not really novel. Meanwhile, there are notable shortcomings in the study, primarily relating to quality control measures and standardization protocols.

Lack of quality control: The absence of comparison between the "leaf wilting ratio" calculated through the presented method and ground truth data obtained from alternative methodologies or reference approaches is a significant flaw. Comparative analyses are crucial for validating the accuracy and reliability of the proposed method.

Environmental variation standardization: Figure 3 highlights notable lighting variations, including harsh shadows and glare during noon hours, which obscure details within the maize canopy. These variations can significantly impact the accuracy of wilting status assessment.

Phenology monitoring and standardization: The manuscript lacks clarity regarding how the phenology of different maize cultivars is monitored and standardized during wilting ratio evaluation. Standardized protocols for phenological monitoring are essential for ensuring uniformity and comparability across 100 maize cultivars.

Author Response

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Author Response File: Author Response.pdf

Reviewer 4 Report

Comments and Suggestions for Authors

Introduction

The introduction lacked a paragraph dedicated to corn - the crop chosen to carry out the experiment. Why corn? Corn is a plant from the C4 group, highly efficient in using light. Corn is one of the most planted crops in the world. Many studies and development of new varieties, techniques, and by-products involve corn cultivation. Widely used in human and animal nutrition...

MM

Line 96 - what is the soil classification? Reference. What is the sand, silt and clay content?

Line 102 - provide more irrigation data. What type of drip hose is used? 16mm? What is the spacing between emitters? What is the flow rate of each emitter? Self-compensating dripper?

Line 115 - what irrigation management method was used? Was it based on reference evapotranspiration? Which equation was used for the calculations? Penman-Monteith-FAO???

I missed more details about water management. The plants were stressed. After taking photographs, was irrigation done to hydrate the plants? Or were you letting the stress go to the point of permanent withering? There is a lack of information regarding the conduct of the study.

Results

Line 215 - curve adjustment has been made. However, the value of the adjustment coefficient (R2) was not presented. What is the validation of the adjustments? Was it significant? I ask this because apparently, the values are very dispersed.

References

 

Of 28 references, only 7 are recent, less than 5 years old. There are many recent works in this area in the literature. Authors need to use more recent references from 2020, 2021, 2022, 2023, and 2024.

Comments for author File: Comments.pdf

Comments on the Quality of English Language

ok

Author Response

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Author Response File: Author Response.pdf

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

You added some good statistics that strongly supported the argument in this manuscript. Some more minor comments are listed below: 

Line118: I meant to add the drought resistance of the ten varieties 006, 019, 115 011, 038, 078, 081, 095, PH4CV, PH6WC, and Zheng 58 since Fig. 8 suggested an interactive effect between drought treatment and variety, not the 100 inbred lines..... Also legend needs to explain Dc and Lv. 

Line 156-169: Still need more details. What are' effective plants'? Survived plants? 

How was the moisture content measured? 

I recommend changing 'A: grain dry weight' to 'A: weight of air-dried' grains to avoid confusion. 

What is the 14%? Harvest index? Plz specify. 

Figure 5: it looks very nice! And it clearly showed that temperature negatively correlated with visible green area. I think it would be even better if you could test correlation between temperature and visible green area. 

Line 279-282: I think it would look better if you can overlay the daily temperature in Fig. 6 the show the trends in wilting and temperature did not closely follow each other. 

Figure 6: add description of red arrows pointing at flowering stage in the graph legend

Figure 7: It is probably self-evident but still need description of which bars/lines correspond to which y axis.

Line 288-302: Can remove the asterisks in Table 4 since p values are listed. 

Figure 9: The y axis title of the left graphs still need correct: Grain weight per (WHAT) in high drought/g. And p values? 

Discussion: In the last version I recommended adding adding a bit more discussion on the "LOW COST" feature of your system. Then you "add some discussion on the innovation and practicability of this method" and the title also changed. I guess the aim of this study moved away from being low cost to being innovative? 

Author Response

Thank you for your decision and constructive comments on my manuscript. We have carefully considered the suggestion of Reviewer and make some changes. We have tried our best to improve and made some changes in the manuscript.

Comments 1: [Line118: I meant to add the drought resistance of the ten varieties 006, 019, 115 011, 038, 078, 081, 095, PH4CV, PH6WC, and Zheng 58 since Fig. 8 suggested an interactive effect between drought treatment and variety, not the 100 inbred lines..... Also legend needs to explain Dc and Lv. ]

Response 1: Thank you for pointing this out. we have revised it in the manuscript. The change can be found – L118.

Comments 2: [Line 156-169: Still need more details. What are' effective plants'? Survived plants? 

How was the moisture content measured? 

I recommend changing 'A: grain dry weight' to 'A: weight of air-dried' grains to avoid confusion. 

What is the 14%? Harvest index? Plz specify. ]

Response 2: Thank you for pointing this out.  the numbers of effective plants are the number of survived plants，we have revised it in the manuscript. The change can be found – L147-157.

Comments 3: [Figure 5: it looks very nice! And it clearly showed that temperature negatively correlated with visible green area. I think it would be even better if you could test correlation between temperature and visible green area.  ]

Response 3: Thank you for pointing this out.  we have revised it in the manuscript. The change can be found – Figure 5.

Comments 4: [Line 279-282: I think it would look better if you can overlay the daily temperature in Fig. 6 the show the trends in wilting and temperature did not closely follow each other. 

Figure 6: add description of red arrows pointing at flowering stage in the graph legend ]

Response 4: Thank you for pointing this out.  we have revised it in the manuscript. The change can be found – Figure 6.

Comments 5: [Figure 7: It is probably self-evident but still need description of which bars/lines correspond to which y axis.]

Response 5: Thank you for pointing this out.  we have revised it in the manuscript. The change can be found – L266-268.

Comments 6: [Line 288-302: Can remove the asterisks in Table 4 since p values are listed. ]

Response 6: Thank you for pointing this out.  we have revised it in the manuscript. The change can be found – Table 4.

Comments 7: [Figure 9: The y axis title of the left graphs still need correct: Grain weight per (WHAT) in high drought/g. And p values? ]

Response 7: Thank you for pointing this out.  we have revised it in the manuscript. The change can be found – Figure 9.

Comments 8: [Discussion: In the last version I recommended adding adding a bit more discussion on the "LOW COST" feature of your system. Then you "add some discussion on the innovation and practicability of this method" and the title also changed. I guess the aim of this study moved away from being low cost to being innovative?  ]

Response 8: Thank you for pointing this out.  This study has done a lot of work on the development of low-cost devices, and has also made some progress, which is conducive to high throughput data determination. In this study, the wilting degree was proposed on the quantitative index of leaf wilting, and the correlation analysis of drought-resistance yield was carried out, which has certain innovation..

 

 

 

Reviewer 3 Report

Comments and Suggestions for Authors

The clarity and credibility of the paper have been significantly enhanced by the addition of supplementary data and supporting evidence provided by the authors.

Author Response

Thank you for your decision and constructive comments on my manuscript.

Reviewer 4 Report

Comments and Suggestions for Authors

A paragraph dedicated to corn was inserted in the introduction – the crop chosen to carry out the experiment.

What about soil classification? Reference. There was no response from the classification system used.

More irrigation data was provided. And the irrigation management method used.

Validation of adjustments was presented

The authors have inserted more recent references.

I give merit to the new work. There was a lot of modification and addition of valuable information.

Comments on the Quality of English Language

ok

Author Response

Thank you for your decision and constructive comments on my manuscript. We have carefully considered the suggestion of Reviewer and make some changes. We have tried our best to improve and made some changes in the manuscript.

Comments 1: [What about soil classification? Reference. There was no response from the classification system used.]

 Response 1: Thank you for pointing this out. we have revised it in the manuscript. The change can be found-L111-112.