Evaluation of Copper-Free Alternatives to Control Grey Mould in Organic Mediterranean Greenhouse Tomato Production

Round 1

Reviewer 1 Report

1.     The dosage in Table 1 is the dosage of the formulation or the active ingredient?

2.     Please add the type of the nozzle in the manuscript

3.     The units in Table 1 are more reasonably expressed as dilutions. And, the units in the text should be unified, for example, “g/hL” and “g/L”

4.     What is the purpose of analyze different pesticides in the selected formulations? Their composition is uncertain?

Author Response

Dear Reviewer,

We would like to thank your thorough work and constructive comments to improve this manuscript. All your comments and suggestions have been taken into account in the new version of the manuscript substantially improving its quality and clarity.

The revised manuscript contents the main changes. Changes have been highlighted by using the “track changes mode” in MS Word version before the edition of English (a certificate of English editing has been supplied with the new manuscript submission).

In this document, author responses are shown in red characters.

Furthermore, we would like to inform you that, at response to different requests of reviewer 2, 3 and 4 we have incorporated other changes. Please find in the new version of the manuscript our changes according with your comments.

Yours.

Response to Reviewer 1 Comments

Comments 1 to 4 have been responded in the new manuscript version as explained.

Additional response to Comment 4: Due to the high efficacy of some products in the in vitro tests, we used two multi-target analyses that are common in Almería to check the presence of pesticides in samples of fruit or plant tissues, searching for any pollution or inclusion of non-declared fungicides in the composition of the products.

Author Response File: Author Response.pdf

Reviewer 2 Report

Dear Authors

The subject matter discussed in the article is important and necessary because the European Union drastically limits the use of various types of chemical fungicides. At the same time, it recommends the use of natural preparations that are environmentally friendly, but their effectiveness in practice is very low.

In their work, the authors undertook to test the effectiveness of 12 biocidal compounds, already used on the market, in comparison to a copper-containing fungicide. Despite the interesting subject matter, the authors did not avoid some significant mistakes.

Details, for easier interpretation, are provided in the points:

- line 105-107 formulation based on basic substances / on plant extrakt.

Why was this division applied? After all, Equisetum arvense is also a plant, just like the aforementioned Mimosa. It should be remembered and it should be emphasized that its active substance / substances are responsible for the effectiveness of the biocides.

- The experiment described in point 2.1.3. is carried out incorrectly. The mycelium does not come into direct contact with the fungicyde. In addition, fungicides in the agar medium (PDA) can diffuse differently, which affects their bioavailability for the cell. Why was the sample in this experiment not performed without the use of a fungicide?

- line 134 Why were the plates incubated under UV light? After all, it is known that UV inhibits the growth of microorganisms, including mold. In addition, at this point of the methodology, not only a description should be prepared, but also a diagram/scheme- what samples were tested after how many days etc.

- line 209 Cinnamon 2 should be indicated because there were two samples called Cinnamon.

Summary:

 The article is more of an application and should be available as a valuable advice for farmers, but the scientific aspect here is small. It is a pity that the authors did not use, for example, some new active substance, e.g. a new extract, which has not been tested on tomato farms so far.

Author Response

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Reviewer 3 Report

Introduction section provides sufficient information regarding the importance and the justification of the study. The discussion is very well structured. After a careful reading, there are few observations that I had to do.

Lines 105-108 – What the diferences between basic substances and plant extracts? Equisetum arvense is also a plant.

Lines 162-163 – Do the authors have more detail of geographical coordinations.

Lines 201-203 – Did the climatic conditions the same/very similar internal and outside the greenhouse?

Line 289 – Which was the harvesting stage? There are six ripening stages of tomato. Did the authors use the Breaker Stage, turning, pink, red?

Line 299 – Did the authors used statistical analysis for in vitro tests?

Author Response

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Reviewer 4 Report

Please use a good scientific editor -  the reading of the work suffers from poor sentence construction, strange words, and repetition

Difficult paper to write with all the combinations. From the work not sure that the cinnamon treatment should rank that highly.  The PHC looks very promising though 

have tried to point out problems with sticky notes 

Comments for author File: Comments.pdf

Author Response

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Round 2

Reviewer 2 Report

The article has been substantially improved, but in my opinion there are still points with which I cannot agree.

1. I still stand by my position on the use of UV to incubate plates with molds Please check for example the abstract in the article Rotem J., H.J. Aust The Effect of Ultraviolet and Solar Radiation and Temperature on Survival of Fungal Propagules Journal of Phytopathology, 133(1):76-84 DOI:10.1111/j.1439-0434.1991.tb00139.x "Exposure to UV reduced survival much more than exposure in darkness."

2. The experiment described in point 2.1.3. is carried out incorrectly. The mycelium does not come into direct contact with the fungicide. In addition, fungicides in the agar medium (PDA) can diffuse differently, which affects their bioavailability for the cell. Why was the sample in this experiment not performed without the use of a fungicide?

The methodology used is common for In vitro mycelial growth inhibition tests (see Ebrahimi et al. (2017); Sernaite et al. (2020)). As described for these tests, the tested products were mixed homogeneously with the agar medium. Controls consisted of fungi transferred into non-amended Petri dishes containing only PDA. Controls were used to calculate the % Inhibition = ((D-d)/D x 100), where d is the diameter of the B. cinerea colony in the presence of the products tested, and D is the maximum Diameter of the B. cinerea colony of the controls, as appears in the text. Sernaite, L.; Rasiukevičiūtė, N.; Valiuškaitė, A. The extracts of cinnamon and cloves as potential biofungicides against strowbery gray mold. Plants 2020, 9, 613. https://doi.org/10.3390/plants9050613 Ebrahimi, L.; Jalali, H.; Etebarian, H.R.; Sahebani, N. Evaluation of antifungal activity of some plant essential oils against tomato gray mold disease. J Plant Pathol 2022, 104, 641–650. https://doi.org/10.1007/s42161-022-01029-x

When plant oils are used, their evaporation takes place and a micro-atmosphere is created on the Petri dishes. However, not all relationships work this way. Not all of them have the ability to diffuse in the medium. This is a common mistake made when checking the effectiveness of biocides. This method can be treated as only a screening method.

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Reviewer 4 Report

The paper although changed in places still has numerous examples of poor english expression which confound judgement of the science . Even more comments have been added. 

Many statements seem to conflict as the paper proceeds

What reads initially as a one type of greenhouse study  expands into discussion with other parameters  eg tomato shape etc      Such factors may be important but all of the changing parameters are not well introduced. 

Please please use professional editing.  

Comments for author File: Comments.pdf

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