Arbuscular Mycorrhizal Fungi Inoculation Reduced the Growth of Pre-Rooted Olive Cuttings in a Greenhouse

Round 1

Reviewer 1 Report

Please see attachment.

Comments for author File: Comments.pdf

Author Response

The authors' responses were written in blue. Corrections in the manuscript, in red.

 

 

Line 19, 24, 50: Mycorrhized change to “Mycorrhizal”

The changes were made (lines 19, 24, 50)

 

Line 27: “that enter in their composition”-----unclear meaning

The sentence was rearranged (see line 27)

 

Introduction------Introduction should start generally and become more specific. It needs rearranging. Furthermore some background on olive would be good to include

We started introduction with general concepts of the mutualistic associations between plants and fungi. We provided data on the importance of mycorrhization to host plants. We stress the potential conflict between the benefits and the need of the plants to supply photosynthates to the fungi. We focused the last paragraph in olive, adding five more references and rearranging the sentences (lines 81-98).

 

Line 39: AM-----define: arbuscular mycorrhiza

Done (see line 39)

 

Line 45: best ----------most documented roles

Done (see line 45)

 

Line 47: in spite of most benefits to plants arise in P-limited soils --------unclear, rephrase

The sentence was rearranged (see line 47).

 

Line 55: Relevant change to “Of relevance”

Done (see line 55)

 

Line 56: have change to “provide”

Done (see line 56)

 

Line 119: is change to “was”

Done (see line 119)

 

Table 1 ------- n=? how many samples?which depth?use exch, for abbreviation

Please, see the new table 1.

 

Line 125: in change to “on”

Done (see line 158)

 

Line 159: kept free from weeds-------how?

By hand (see line 159)

 

Line 206: more change to “most”

That sentence was removed at the request of another reviewer

 

Table 2: leaf mass per area (LMA)----------- usually would express SLA, not LMA

Please see our argument. Leaf mass per area (LMA), the ratio of leaf dry mass to leaf area, is the inverse of specific leaf area (SLA). LMA presented in Table 2 and the respective units are correct​.

 

Table 5 and 6 ------------n = ?

n=6, please see the tables 5 and 6

 

Ref 11: Cooper, J.E., Scherer, H.W.---------- format problem

The format has been corrected (lines 481-482)

 

Final note

The authors also want to say that this manuscript was reviewed by four different reviewers who did a very valuable corrective work and from which resulted ~70 changes in the manuscript that were marked in red. Unfortunately, in these situations it is not always easy to answer exhaustively to the criticisms of each one of the reviewers. However, if any reviewer finds that the authors' work was unsatisfactory, we are available to quickly incorporate their further suggestions.

Reviewer 2 Report

 

Comments on manuscript soilsystems-1176253

 

The manuscript it is in this scope of the journal, regarding the effect of commercial mycorrhizal fungi on pre-rooted olive cuttings.

 

Tables 1 and 2 – Miss the titles of the second and fourth columns. Please correct in order to clarify what means the data of each one of this two columns.

The total amount of clay + silt + sand is high than 100%. Please correct.

 

Line 196 – Please add “… JMP14 software (city, country).”.

 

Figures 3 and 4 – The nutrient concentration is expressed on a dry matter basis? Please add to the to the YY axis (g kg-1 DM-1).

 

Author Response

The authors' responses were written in blue. Corrections in the manuscript, in red.

 

 

 

(x) I would not like to sign my review report
( ) I would like to sign my review report

English language and style

( ) Extensive editing of English language and style required
( ) Moderate English changes required
(x) English language and style are fine/minor spell check required
( ) I don't feel qualified to judge about the English language and style

 

	Yes	Can be improved	Must be improved	Not applicable
Does the introduction provide sufficient background and include all relevant references?	(x)	( )	( )	( )
Is the research design appropriate?	(x)	( )	( )	( )
Are the methods adequately described?	(x)	( )	( )	( )
Are the results clearly presented?	(x)	( )	( )	( )
Are the conclusions supported by the results?	(x)	( )	( )	( )

Comments and Suggestions for Authors

 

Comments on manuscript soilsystems-1176253

 

The manuscript it is in this scope of the journal, regarding the effect of commercial mycorrhizal fungi on pre-rooted olive cuttings.

 

Tables 1 and 2 – Miss the titles of the second and fourth columns. Please correct in order to clarify what means the data of each one of this two columns.

 

We think the problem in only in table 1. We provided a set of changes in table 1 for a better clarification.

 

The total amount of clay + silt + sand is high than 100%. Please correct.

 

The reviewer is right. The sum of clay + silt + sand is equal to 100.1%. The problem was in the rounding. The situation has now been corrected. Please, see the new table 1.

 

Line 196 – Please add “… JMP14 software (city, country).”.

 

Missing data was added (lines 196-197)

 

 

Figures 3 and 4 – The nutrient concentration is expressed on a dry matter basis? Please add to the to the YY axis (g kg-1 DM-1).

 

DM was added. Please, check in the new figures.

 

 

Final note

The authors also want to say that this manuscript was reviewed by four different reviewers who did a very valuable corrective work and from which resulted ~70 changes in the manuscript that were marked in red. Unfortunately, in these situations it is not always easy to answer exhaustively to the criticisms of each one of the reviewers. However, if any reviewer finds that the authors' work was unsatisfactory, we are available to quickly incorporate their further suggestions.

 

Reviewer 3 Report

The authors focused on the effect of a commercial mycorrhizal fungi for growth of pre-rooted olive cuttings in a greenhouse. However, the statistical treatment should be reconsidered, and there is no evidence of AM fungi infection during pot cultivation experiments, which are discussed mainly in the manuscript. Major and minor comments are shown below.

#Major comments
3. Results: The authors compared the Dry matter yield among cultivars or soil treatments. In the soil treatments, the results contain dry matter yields from different cultivars. To discuss the AM effect by different soil treatments, authors should also show the statistical results from sole cultivar. At least, authors should show the results in a supplementary table. 

4. Discussion and 5. Conclusions: The authors mentioned several discussions about utilization of AM fungi and zeolite. The authors mentioned about the negative effect of AM fungi inoculation due to the competition for photosynthates between fungi and young plants during the formation of the arbuscular structures and hyphal network. However, this is overspeculation because the authors did not confirm the infection of AM fungi into three cultivars. Please reconsider the discussion and conclusions by considering the experimental results.

#Minor comments
L39: AM should be defined firstly.
L45: P should be defined as phosphorus (P).
Table 1: The table is not well described. For example, the Clay, Silt, and Sand show only P, K, and Ca, respectively. All soils should have P, K, and Ca information. And the pH, Organic C, and Extractable B are from mixed soil. Please reconsider the table format. And the annotation 1–7 should be described in materials and methods part.
Figures 1 and 2: Graph legends of Stems and Leaves are same. Please modify the graph legends. 
L236: Please delete the “Table 0.”.
L238–241: Please show the results in this description because the author mentioned that the trend of the results is similar to experiment 1. At least, authors can show the results as supplementary information.

Author Response

The authors' responses were written in blue. Corrections in the manuscript, in red.

 

 

 

 

(x) I would not like to sign my review report
( ) I would like to sign my review report

English language and style

( ) Extensive editing of English language and style required
( ) Moderate English changes required
( ) English language and style are fine/minor spell check required
(x) I don't feel qualified to judge about the English language and style

 

	Yes	Can be improved	Must be improved	Not applicable
Does the introduction provide sufficient background and include all relevant references?	( )	(x)	( )	( )
Is the research design appropriate?	( )	( )	(x)	( )
Are the methods adequately described?	( )	(x)	( )	( )
Are the results clearly presented?	( )	( )	(x)	( )
Are the conclusions supported by the results?	( )	( )	(x)	( )

Comments and Suggestions for Authors

The authors focused on the effect of a commercial mycorrhizal fungi for growth of pre-rooted olive cuttings in a greenhouse. However, the statistical treatment should be reconsidered, and there is no evidence of AM fungi infection during pot cultivation experiments, which are discussed mainly in the manuscript. Major and minor comments are shown below.

#Major comments
3. Results: The authors compared the Dry matter yield among cultivars or soil treatments. In the soil treatments, the results contain dry matter yields from different cultivars. To discuss the AM effect by different soil treatments, authors should also show the statistical results from sole cultivar. At least, authors should show the results in a supplementary table. 

By our initial decision, we arranged the experiment 1 as a factorial design with two factors (cultivars and soil treatments) (two-way ANOVA), each factor with three levels (cultivars: Madural, Verdeal and Cobrançosa and soil treatments: Control, Zeolites and Mycorrhizal). By arranging the experiment in this way we can compare the effect of the cultivar, the effect of the soil treatments and the interaction of the two factors.

Experiments arranged as a factorial are probably the most widely used in agriculture. We thought that this was the best model for organizing this experiment.

We suspect that the presentation of results from sole cultivar is not the best solution. To present the results separated by cultivar, we should run 6 one-way ANOVAs after separating the factorial experiment into 6 experiments completely randomized. We will have the results of the three cultivars on each of the three growing mediums: control, zeolites and mycorrhizal. By other side, we also will have the results of the three growing mediums in each one of the three cultivars. We will need six different figures to present the results. We would have to exchange one factorial experiment for six completely randomized experiments. In our opinion, this would make it more difficult to interpret the results.

Furthermore, one of the four reviewers who reviewed this manuscript, made a data analysis proposal different from ours and different from the one that this reviewer is proposing. Faced with a somewhat confusing situation, the authors decided to maintain the data processing they had envisioned.

4. Discussion and 5. Conclusions: The authors mentioned several discussions about utilization of AM fungi and zeolite. The authors mentioned about the negative effect of AM fungi inoculation due to the competition for photosynthates between fungi and young plants during the formation of the arbuscular structures and hyphal network. However, this is overspeculation because the authors did not confirm the infection of AM fungi into three cultivars. Please reconsider the discussion and conclusions by considering the experimental results.

Yes, the reviewer is right. We did not use a methodology to confirm the infection of AM fungi. However, we present a set of analyzes of the performance of the plant (physiological, nutritional and dry matter yield) and of soil properties that gave consistency to the thesis that we defend. The loss of biomass of the three cultivars in the treatment with inoculation with mycorrhizal fungi was consistent, as well as in a second experiment. All data points to problems of competition for photosynthates, mainly because the greenhouse cultivation removes performance to the photosynthetic process and made the growth of the inoculated plants even more difficult. Similar situations were found in previous studies, but the authors were unable to find a good explanation for what happened. The entire text was revised and some modifications were introduced to reduce overspeculation (Please, see all the changes made in discussion section that were marked in red)

#Minor comments
L39: AM should be defined firstly.

The change was done, please see line 39

L45: P should be defined as phosphorus (P).

P was abbreviated in line 26.

Table 1: The table is not well described. For example, the Clay, Silt, and Sand show only P, K, and Ca, respectively. All soils should have P, K, and Ca information. And the pH, Organic C, and Extractable B are from mixed soil. Please reconsider the table format. And the annotation 1–7 should be described in materials and methods part.

Please, take a look to the new table 1. In the columns 1 and 3 we put the soil properties and in the columns 2 and 4 the corresponding values. We added “value” to the title of the columns 2 and 4 to reduce the risk of misinterpretation.

Annotation 1-7 was also described in materials and methods (lines 183-190). We put also the annotations here because for almost all journals this is mandatory because the tables have to be self-explanatory and should contain all information without having to read the text.

Figures 1 and 2: Graph legends of Stems and Leaves are same. Please modify the graph legends. 

We didn’t understand this issue, the legend is roots, stems and leaves (each one with a different pattern or color)! We don't know if there is a problem with the version the reviewer received.

L236: Please delete the “Table 0.”.

Yes, the error was removed (see line 234)

L238–241: Please show the results in this description because the author mentioned that the trend of the results is similar to experiment 1. At least, authors can show the results as supplementary information.

Following the suggestion of one of the other four reviewers of this manuscript, this paragraph has been completely eliminated.

 

Final note

The authors also want to say that this manuscript was reviewed by four different reviewers who did a very valuable corrective work and from which resulted ~70 changes in the manuscript that were marked in red. Unfortunately, in these situations it is not always easy to answer exhaustively to the criticisms of each one of the reviewers. However, if any reviewer finds that the authors' work was unsatisfactory, we are available to quickly incorporate their further suggestions.

Reviewer 4 Report

Comments to the Authors

 

GENERAL COMMENTS

In my opinion, the manuscript is very valuable. Describes the scale of mycorrhiza influence on the growth and development of olive seedlings. The introduction is easily written. It has well-chosen content. However, there was no information about the species of olive trees or a few information about olive trees that would emphasize their specificity. The "Materials and methods" chapter is, in my opinion, sometimes described too precisely or the description of the methods is a bit chaotic and therefore incomprehensible. Statistical data analysis is at a sufficient level, although it would be good if innovative graphic solutions appeared. The results are also quite chaotic. Well-described results should at least put a lot of emphasis or describe only the relevant results. The chapters "Discussion" and "Conclusions" are important and interesting elements of the publication.

 

SPECIFIC COMMENTS

The publication is valuable. Describes the scale of mycorrhiza influence on the growth and development of olive seedlings.

chapter "Introduction"

The introduction is easily written. It has well-chosen content. However, there was no information about the species of olive trees, or some information about olive trees, emphasizing their specificity.

Chapter “Materials and methods”

The methodology is described too precisely in my opinion. The exact dates for setting up the experiments are not that important. In turn, there are also elements of the method description written chaotically and some parts are incomprehensible. That is why I have a suggestion that may help in redrafting this chapter.

In subsection 2.1 the description of the first experiment is a bit complicated, and it is especially unclear what exactly three treatments were applied. I propose to edit this paragraph. Experiment 2 was described in much clearer language.

Subsection 2.2. Again, it should be indicated whether measurements were made in both experiments.

Subsection 2.3 I do not know what are the exact dates of the next stages of research? Is it not enough to say that the experiment ended during the period when seedlings usually reach farmers?

Subsection 2.4

A shortcoming in the manuscript is the lack of a graphical, slightly more innovative presentation of the data. I think that the PCA - Principal component analysis method would show the correlations selected from the results by the authors in a very interesting way. Is it possible to propose newer statistical solutions?

Chapter “Results”

The results are also quite chaotic. It is difficult to pinpoint the relevant results, as there is much information in each description of the determinations that the results were not relevant, especially regarding the differences between the species of olive seedlings. This gives you the feeling that the research design was not set up correctly and it lowers the value of the work. Well-described results should only describe the relevant results.

Chapter “Discussion”

The discussion chapter is the best written chapter of all chapters. At this stage of the publication, it is possible to understand what specific research results were obtained by the authors. Which confirms the belief that all the described data, irrelevant to the results chapter, make the results unclear. The discussion is built on 29 literature items, including only 11 which appeared after 2015.

Chapter “Conclusions”

They are written correctly in my opinion.

References

I also think that a scientific work should be based on a larger pool of the latest publications.

Author Response

The authors' responses were written in blue. Corrections in the manuscript, in red.

 

 

 

Open Review

(x) I would not like to sign my review report
( ) I would like to sign my review report

English language and style

( ) Extensive editing of English language and style required
( ) Moderate English changes required
( ) English language and style are fine/minor spell check required
(x) I don't feel qualified to judge about the English language and style

 

	Yes	Can be improved	Must be improved	Not applicable
Does the introduction provide sufficient background and include all relevant references?	( )	(x)	( )	( )
Is the research design appropriate?	(x)	( )	( )	( )
Are the methods adequately described?	( )	(x)	( )	( )
Are the results clearly presented?	( )	(x)	( )	( )
Are the conclusions supported by the results?	(x)	( )	( )	( )

Comments and Suggestions for Authors

Comments to the Authors

 

GENERAL COMMENTS

In my opinion, the manuscript is very valuable. Describes the scale of mycorrhiza influence on the growth and development of olive seedlings. The introduction is easily written. It has well-chosen content. However, there was no information about the species of olive trees or a few information about olive trees that would emphasize their specificity. The "Materials and methods" chapter is, in my opinion, sometimes described too precisely or the description of the methods is a bit chaotic and therefore incomprehensible. Statistical data analysis is at a sufficient level, although it would be good if innovative graphic solutions appeared. The results are also quite chaotic. Well-described results should at least put a lot of emphasis or describe only the relevant results. The chapters "Discussion" and "Conclusions" are important and interesting elements of the publication.

 

SPECIFIC COMMENTS

The publication is valuable. Describes the scale of mycorrhiza influence on the growth and development of olive seedlings.

chapter "Introduction"

The introduction is easily written. It has well-chosen content. However, there was no information about the species of olive trees, or some information about olive trees, emphasizing their specificity.

We added five new references on olive tree and texts were rearranged (see all the red, but in particular the last paragraph of the introduction section)

 

Chapter “Materials and methods”

The methodology is described too precisely in my opinion. The exact dates for setting up the experiments are not that important. In turn, there are also elements of the method description written chaotically and some parts are incomprehensible. That is why I have a suggestion that may help in redrafting this chapter.

In subsection 2.1 the description of the first experiment is a bit complicated, and it is especially unclear what exactly three treatments were applied. I propose to edit this paragraph. Experiment 2 was described in much clearer language.

We removed several sentences and rephrased part of the texts in particular were we put a red mark (lines 103, 109, 119, 125, 158)

Subsection 2.2. Again, it should be indicated whether measurements were made in both experiments.

We rephrased the text (see lines 164-165)

 

Subsection 2.3 I do not know what are the exact dates of the next stages of research? Is it not enough to say that the experiment ended during the period when seedlings usually reach farmers?

We tried to clarify the sentence (see line 173)

 

Subsection 2.4

A shortcoming in the manuscript is the lack of a graphical, slightly more innovative presentation of the data. I think that the PCA - Principal component analysis method would show the correlations selected from the results by the authors in a very interesting way. Is it possible to propose newer statistical solutions?

We have two reviewers proposing different approaches for the analysis of results. Of course, we agreed that PCA could be included in the data processing. However, we follow a classic line of data treatment with analysis of variance and presentation of data in figures and tables. Authors are in a difficult situation when two reviewers refer to the same topic but propose things that are valid but do not go in the same direction. Thus, the authors decided to maintain the data analysis that was devised by them and that shows the results in a satisfactory way.

Chapter “Results”

The results are also quite chaotic. It is difficult to pinpoint the relevant results, as there is much information in each description of the determinations that the results were not relevant, especially regarding the differences between the species of olive seedlings. This gives you the feeling that the research design was not set up correctly and it lowers the value of the work. Well-described results should only describe the relevant results.

We removed several part of the texts (lines 205, 207, 225, 241, 246, 276) and an entire paragraph (line 236)

Chapter “Discussion”

The discussion chapter is the best written chapter of all chapters. At this stage of the publication, it is possible to understand what specific research results were obtained by the authors. Which confirms the belief that all the described data, irrelevant to the results chapter, make the results unclear. The discussion is built on 29 literature items, including only 11 which appeared after 2015.

We updated the reference list with more six references and several changes were made on the texts (see lines 323-325, 330, 338-342, 363, 420-422)

 

Chapter “Conclusions”

They are written correctly in my opinion.

 

References

I also think that a scientific work should be based on a larger pool of the latest publications.

Several new references were added (lines 492-494, 503-505, 507-509, 530-532, 539-541, 554-555) and texts rearranged accordingly (see the new introduction and discussion sections)

 

Final note

The authors also want to say that this manuscript was reviewed by four different reviewers who did a very valuable corrective work and from which resulted ~70 changes in the manuscript that were marked in red. Unfortunately, in these situations it is not always easy to answer exhaustively to the criticisms of each one of the reviewers. However, if any reviewer finds that the authors' work was unsatisfactory, we are available to quickly incorporate their further suggestions.

Round 2

Reviewer 3 Report

I understand the difficulty to be consistent modification from 4 reviewers’ comments. The modified manuscript is suitable for this journal. And I agree to authors' modification.