Induced Drought Stress Response of European Beech Seedlings Treated with Hydrogel and Ectomycorrhizal Inoculum
Round 1
Reviewer 1 Report
The manuscript contains interesting and important findings that hydrogel or the addition of mycorrhiza during planting does not much improve tree survival during drought. Even though the experiment is limited to a single soil type, the data provide a worthy contribution to our knowledge of how to increase the survival rates of European beech after planting.
The paper is rather well-written. Especially methodology is presented in detail. The experiment is well-designed, with over 700 trees being measured. Results are clearly shown, and the text describing graphs is more detailed than in most papers. The detailness also applies to the discussion section, which is quite lengthy and tedious: I would suggest shortening the sentences and the text.
I have only a few minor comments, highlighted in the attached PDF file. My main question would lead to the soil water content. It is hard to believe 40 % mortality in two of the treatments where the soil moisture never dropped below the wilting point. Is any information about leaf water potential available? Did the seedlings resprout after re-watering?
Comments for author File: 
Comments.pdf
Author Response
Cover letter
Manuscript ID: forests-2515923
Title: Induced Drought Stress Response of European Beech Seedlings Treated with Hydrogel and Ectomycorrhizal Inoculum
Response to the Reviewer's 1 review report:
Dear Reviewer,
Thank you very much for your comments, suggestions and in all appreciative report on the manuscript. We reply to your comments as follows:
Reviewer: The discussion section is quite lengthy and tedious: I would suggest shortening the sentences and the text.
Response: We have realized the Discussion section is extensive. Our ambition, encouraged by the Journal Instructions for Authors concerning discussion has been to discuss the study findings in the broadest context possible (explanations, implications, confrontations with previous results). There is difficult to avoid a long discussion not being tedious. We have aimed to make just minor revisions in the text of this section.
Reviewer: It is hard to believe 40 % mortality in two of the treatments where the soil moisture never dropped below the wilting point. Is any information about leaf water potential available? Did the seedlings resprout after re-watering?
Response: We are not able explain the seedling mortality you think is unexpectedly high. We can just uncertainly suppose that the long-term mild drought stress combined with specific substrate and seedling traits and environmental conditions could affect this result. Moreover, soil water potential (SWP) −1,500 kPa usually considered as corresponding to wilting point did not have to properly relate to wilting point in our experiment. As known from the literature, the value of SWP for wilting point varies with the type of plant, soil and climate, and ranges from -500 to -3,000 kPa. Beyond -300 kPa, the soil is too dry for the roots of most plants to extract water. On the other hand, the PWP was derived for agricultural plants, and according to Cocozza et al. (2016) young trees like European beech may deplete water resources near to the fine roots at lower soil water potentials than the PWP. However, Jamnická et al. (2013) found significant decline in physiological processes of beech seedlings as early as at SWP -900 kPa.
Leaf water potential was not measured. The dead seedlings resprouted after re-watering neither through experimental period in PW nor in subsequent spring.
Line 103: Is it 70% of relative extractable water? Or 70% of volume, weight?
The volumetric water content is the percentage of the volume of water from the volume of substrate (total pore space and volume of substrate solid phase). We have tried to explain water content related to the set up watering treatments in the subsection 2.4. More detailed information on relations among water retention, potential and volume content can be found in the studies cited in this subsection and another relevant literature, e.g., Michel J-C., Mires and Peat 2010, 6, 1–6.
Figure 1: Why is such high mortality in RW and PW? The soil dessicated only to wilting point in PW and was much higher in RW.
See the response to the related comment above please.
Figure 2: This figure repeats information from Figure 1. I suggest to delete it.
We have just wanted to emphasize survival and significance of differences among treatments at the end of the trial in the Figure 2. To show this in the Figure 1 has appeared rather complicated or even impossible. But we accept your suggestion to delete the figure.
Line 385: This last sentence belongs rather to the discussion.
The sentence was deleted.
Line 569: Also, FV/Fm is quite insensitive to mild drought stress. It changes only when the photosystem is severely damaged by drought.
Thank you for the comment. It confirms in other words written about this item in the discussion.
Line 591: Are there any results from fast kinetics? I do not remember them from the text. If not, this paragraph can be deleted.
Chl a fluorescence assessed in this study is known also as fast kinetics of chl a fluorescence (e.g., Coccoza et al., Front. Plant Sci. 2016, 7, 886; Jamnická et al., Cent.Eur.For.J. 2020, 66, 227-242). Inconsistent and thus confusing wording on this issue in the manuscript has been corrected. Since we would like shortly discuss effect of additives on chl fluorescence, we would rather the paragraph to be remained.
Reviewer 2 Report
This study examines the effect of different water treatments and applications (hydrogel, EMF) on the survival and growth of Fagus sylvatica L seedlings. Studies like this are extremely important to gain knowledge on the effects of droughts (especially in the context of climate change) and to seek adaptive measures. Overall, the manuscript is very well structured. I have some minor comment/suggestions given below:
- General recommendation: Add schematics, figures and tables which will help the reader to gain better understanding on the experimental design.
-Lines 99-114: A figure with the experimental design will be helpful for the reader.
-line 186: This is the “field capacity”. What do you mean by “poorly available water”?
-Lines 191 – 193: Can you elaborate on the method used to estimate the wilting point?
-Section 2.5.3: Please provide some references for the methods described.
- Section 2.5.4: Please elaborate further on the method used to measure chlorophyll a
-Figure 1: What are the units of X-axis?
Author Response
Cover letter
Manuscript ID: forests-2515923
Title: Induced Drought Stress Response of European Beech Seedlings Treated with Hydrogel and Ectomycorrhizal Inoculum
Response to the Reviewer's 2 review report:
Dear Reviewer,
Thank you very much for your comments, suggestions and in all appreciative report on the manuscript. We reply to your comments as follows:
- General recommendation: Add schematics, figures and tables which will help the reader to gain better understanding on the experimental design. Lines 99-114: A figure with the experimental design will be helpful for the reader.
A schematic experimental design has been added to the manuscript.
- Line 186: This is the “field capacity”. What do you mean by “poorly available water”?
To our knowledge, volumetric water content (VWC) and field capacity (FC) are at once related but different soil water categories. VWC is parameter quantifying soil water content; it is the ratio/percentage of the volume of water to/from the volume of soil (total pore space and soil solid phase). FC, saturation, and wilting point are soil water thresholds (status), specific values of water content indicating water availability for plants. We derived water contents representing watering treatments from the fully saturated substrate (all pores filled with water). VWC at the saturation point is equivalent to the total pore space. Unlike VWC, FC is the upper limit (content) of the available water at which water in larger pores have been drained away by gravity after soil had been saturated. We have done an attempt to rephrase and improve clarity of the sentence on VWC.
Based on the relevant literature, our meaning of poorly available water is amount of available water in growth medium corresponding to soil water potential range between −10 and −1,500 kPa. Water content within this range of water potential does not have to be sufficient for supplying water demands of plants and water deficit can hamper physiological processes and overall development of plants.
- Lines 191 – 193: Can you elaborate on the method used to estimate the wilting point?
We apologize, but we are not sure what you exactly mean by this comment. We used conventional thresholds of water potentials to set the watering treatments including wilting point, except of no watering. A water potential value adequate to wilting point of our seedlings in the particular growth substrate was not searched and estimated. We believe that procedures concerning setting and maintenance of the watering treatments were described concisely and clearly. We really have no idea how we could develop the method used to estimate the wilting point.
- Section 2.5.3: Please provide some references for the methods described.
The standardized routine well-known methods have been stated in the manuscript and the analyses performed in the accredited Central Forestry Laboratory of National Forest Centre according to procedures involved in the relevant standards and manuals. These methods were already introduced and used in several previous studies whereby no referenced to any references. Accordingly, we have thought that our presentation of the methods coincides with usual approach to this methodology, provides sufficient information on the foliar analyses carried out and the citing of the above mentioned sources would be useless and even inappropriate.
- Section 2.5.4: Please elaborate further on the method used to measure chlorophyll a
Minor corrections have been made in the manuscript.
- Figure 1: What are the units of X-axis?
The unit of axis X is Date. The axis depiction has been changed and its comprehensibility (we hope) improved.
Reviewer 3 Report
The manuscript showed the results from Drought Stress Response of European Beech Seedlings Treated with Hydrogel and Ectomycorrhizal Inoculum, it is interesting in for grow of this species. However, it is too simple for physiological and molecular responses at different treats.
1. The different watering regimes are too simple, because only FW, RW, PW and NW, often 5-7 treatments will be designed.
2. Molecular analysis in the abstract, but I did find the detailed contents in the text.
Author Response
Cover letter
Manuscript ID: forests-2515923
Title: Induced Drought Stress Response of European Beech Seedlings Treated with Hydrogel and Ectomycorrhizal Inoculum
Response to the Reviewer's 3 review report:
Dear Reviewer,
Thank you very much for your comments, suggestions and in all appreciative report on the manuscript. We reply to your comments as follows:
Although you kindly recommended any sections can (but not must) be improved, by reason you did not identify particular issues to be improved, and these recommendations contradict to those of at least one of the other two reviewers, we have afforded to let these sections in general unchanged. Please see revised manuscript to become familiar with the revisions made.
- The different watering regimes are too simple, because only FW, RW, PW and NW, often 5-7 treatments will be designed.
Of course, we agree more treatments – better design and more outcomes. But we also have assumed that 4 watering treatments might be an acceptable treatment range. Four or less treatments were designed in studies with topic similar to that solved in our manuscript, e.g., Orikiriza et al., J. Environ. Protect. 2013, 4, 713–721; Jamnicka et al., Plant Soil Environ. 2013, 59, 446–541; Coccoza et al., Front. Plant Sci. 2016, 7, 886; Robakowski et al., Forests 2020, 11, 1011;
- Molecular analysis in the abstract, but I did find the detailed contents in the text.
We do not understand comprehensively what you mean by this comment, but you did not find a description of the methods used for molecular analysis as we have decided that in this study is enough to reference the molecular methods to the literature. However, we add a basic information on the methods to the revised manuscript (the subsection 2.5.2.).
Round 2
Reviewer 3 Report
The authors have addressed my comments, it could be accepted for publish.
