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  • Bubai Bhakta1,
  • Shinzo Yamane2 and
  • Jatindra Nath Bhakta3,4
  • et al.

Reviewer 1: Anonymous Reviewer 2: Anonymous Reviewer 3: Alexander Smirnov

Round 1

Reviewer 1 Report

The manuscript “Microbial Control in Greenhouse by Spraying Slightly Acidic Electrolyzed Water” is within the scope of the journal. It presents the original research results. Conclusions result from the scope of the performed research, but in order for the manuscript to be published, it should be corrected taking into account the following comments

1.    ppm is a non-SI unit.

2.    Line 74 DW abbreviation not explained.

3.    Chapter 2.5. describe the growing conditions of plants and write what the measurement of plant growth consisted in. There is also no information about the number of repetitions.

4.    Chapter 2.8. should be moved to chapter 2.3.

5.    Chapter 2 is missing a section on statistical methods.

6.    The description under the title Figure 2 should be moved to chapter 2, while statistical calculations should be included in this figure.

7.    Why is Figure 4 shown after Figure 2, but Figure 3 is missing. Information contained in Figure 4 after the title of this Figure should be moved to Chapter 2. The same remark applies to the other Figures.

8.    In chapter 3.6. Figures 9A and 9B are described, and there is no such figure in the manuscript.

9.    In applications for the first time, write the full name of SAEW and put the abbreviation in parentheses. The note also applies to IPM

10. In the section Referrences, doi are missing after each reference. Also, too much literature is from five years ago.

11. Chapter 3. Results and discussion I propose to divide it into two separate chapters 3. Reuults, 4. Discussion

Author Response

Response to Reviewer 1

  1. ppm is a non-SI unit.

Response 1. We changed ppm to mg/L.

 

  1. Line 74 DW abbreviation not explained.

Response 2. We added the abbreviation as follows. distilled water (DW).

 

  1. Chapter 2.5. describe the growing conditions of plants and write what the measurement of plant growth consisted in. There is also no information about the number of repetitions.

Response 3. We modified the sentences as follows.

Cucumbers and eggplant were grown in the greenhouse. Tap water was sprinkled over the bedding once a day. Side shades were opened when the temperature was high. Six cucumbers and eggplant were selected for this experiment. The growth of four branches of cucumbers and two branches of eggplant from each plant was measured weekly for up to 70 days. The average of branch lengths was calculated.

 

  1. Chapter 2.8. should be moved to chapter 2.3.

Response 4. We moved chapter 2.8. to 2.3 and renumbered them.

 

  1. Chapter 2 is missing a section on statistical methods.

Response 5. We added chapter 2.9. for statical analysis as follows.

2.9. Statistical Analysis

All experiments were conducted with multiple replicates and data was analyzed using Student’s t-tests at the 1% level.

 

  1. The description under the title Figure 2 should be moved to chapter 2, while statistical calculations should be included in this figure.

Response 6. We moved some descriptions in the legend of Figure 2 to chapter 2.2 and rewrote the legend.

 

  1. Why is Figure 4 shown after Figure 2, but Figure 3 is missing. Information contained in Figure 4 after the title of this Figure should be moved to Chapter 2. The same remark applies to the other Figures.

Response 7. The submitted manuscript contained Figure 3. For some reason, Figure 3 was missing in the manuscript. We added Figure 3 to the text.

We moved some descriptions in Figure 4 to chapter 2.5. As the reviewer suggested, we moved detailed descriptions to the chapter on materials and methods.

 

  1. In chapter 3.6. Figures 9A and 9B are described, and there is no such figure in the manuscript.

Response 8. Sorry. We forgot to add Figure 9. We added Figure 9.

 

  1. In applications for the first time, write the full name of SAEW and put the abbreviation in parentheses. The note also applies to IPM

Response 9. We used the abbreviation when it appeared for the first time.

 

  1. In the section Referrences, doi are missing after each reference. Also, too much literature is from five years ago.

Response 10. We added doi for each reference and deleted some old references, if newer references have enough information.

 

  1. Chapter 3. Results and discussion I propose to divide it into two separate chapters 3. Reuults, 4. Discussion

Response 11. We divided into two chapters.

Author Response File: Author Response.docx

Reviewer 2 Report

Horticulturae-2115946 is very interesting and give the valuable information to the researchers and readers. The subject of the manuscript is consistent with the scope of the Journal. Thus, I suggested that the manuscript need to be major revised before it is accepted by this journal.

1.     Manuscript needs through language editing.

2.     Line 11: ‘While’ is not black body.

3.     Line 20: ‘which is like killing two birds with one stone. ’should be deleted.

4.     Line 31: ‘organic acids’ should be listed in detail。

5.     Line 34-35: ‘However, these compounds are toxic to plants, animals, and humans [8,9].’ should be revised as ‘However, these compounds are potential toxic to plants, animals, and humans [8,9].’.

6.     Line 39: ‘considered an’ should be revised as ‘considered as an’.

7.     Line 46-47: These sentences should be revised as ‘SAEW has been paid more attention in agriculture, and shown to prevent and control bacterial infection [18-25].’.

8.     Line 74: ‘DW’ should be full name. Please check the full text for the first time abbreviations appear.

9.     Line 94-95: ‘minutes’ should be revised as ‘min’. Please check the full text

10.   Line 155: ‘E. coli’ should be italic. Please check the full text

11.   The logic and neat of introduction need to be further improved.

12.   Results and discussion must be written separately.

Author Response

Response to reviewer 2

 

  1. Manuscript needs through language editing.

Response 1. We did English editing with MDPI paid editing services.

 

  1. Line 11: ‘While’ is not black body.

Response 2. We modified it.

 

  1. Line 20: ‘which is like killing two birds with one stone. ’should be deleted.

Response 3. We deleted the sentence.

  1. Line 31: ‘organic acids’ should be listed in detail。

Response 4. We decided to remove the terms organic acids. The resulting sentence is as follows.

Sodium hypochlorite (NaOCl), hydrogen peroxide (H2O2), and chlorine dioxide (ClO2) are widely used as disinfectants

 

  1. Line 34-35: ‘However, these compounds are toxic to plants, animals, and humans [8,9].’ should be revised as ‘However, these compounds are potential toxic to plants, animals, and humans [8,9].’.

Response 5. We modified it as the reviewer suggested.

 

  1. Line 39: ‘considered an’ should be revised as ‘considered as an’.

Response 6. We modified it as the reviewer suggested.

 

  1. Line 46-47: These sentences should be revised as ‘SAEW has been paid more attention in agriculture, and shown to prevent and control bacterial infection [18-25].’.

Response 7. We modified it as the reviewer suggested.

 

  1. Line 74: ‘DW’ should be full name. Please check the full text for the first time abbreviations appear.

Response 8. We added the abbreviation distilled water.

 

  1. Line 94-95: ‘minutes’ should be revised as ‘min’. Please check the full text

Response 9. We modified it as the reviewer suggested.

 

  1. Line 155: ‘E. coli’ should be italic. Please check the full text

Response 10. We modified it in the figure legend.

 

  1. The logic and neat of introduction need to be further improved.

Response 11. We rewrote Introduction chapter.

 

  1. Results and discussion must be written separately.

Response 12. We separated into two chapters.

Author Response File: Author Response.docx

Reviewer 3 Report

The fight against microbes in the greenhouse is certainly an urgent task nowadays. In the manuscript, the authors used for this purpose a solution of slightly acidic electrolysis water. However, the article needs serious revision. The annotation should be expanded, the methods should be briefly described and you must indicate the research problem being solved. In the introduction, please clearly state the purpose and novelty of the study. Explain why solutions were chosen with a chlorine concentration of 5-25 ppm for in vitro experiments and 25-40 ppm for greenhouse. In the Materials and Methods section, describe how the results were statistically processed. Figure 3 is missing. Figure 4 does not show margins of error (error bars) for live cells. Explain why there are such large spreads of values for dead cells. Figure 9 is missing. There is no discussion chapter. The conclusion that SAEW successfully reduces airborne microorganisms is debatable, as Figure 4 shows that the statistically significant difference between treatment and control was not significant. In general, the manuscript looks raw and unfinished. In my opinion it should be strengthened by additional studies of the effect of slightly acidic electrolysis water solution of various concentrations on various microorganisms in greenhouse.

Author Response

Response to reviewer 3

 

  1. The annotation should be expanded, the methods should be briefly described and you must indicate the research problem being solved.

Response 1. Thank you for your suggestions.

We wrote the research problem in the text in lines 53-57 as follows.

“In this study, SAEW mist was sprayed in the greenhouse to control airborne microorganisms. In addition, this study combined integrated pest management (IPM) and humidity using SAEW spraying. We also investigated the effects of SAEW spraying on the environment in the greenhouse, such as plant growth and bacterial community structures in the soil and on plant leaf surfaces.”

  1. In the introduction, please clearly state the purpose and novelty of the study.

Response 2. Thank you for your comments.

As indicated in response 1, we indicated the purpose in the last part of the introduction.

 

  1. Explain why solutions were chosen with a chlorine concentration of 5-25 ppm for in vitro experiments and 25-40 ppm for greenhouse.

Response 3. In vitro experiments, we changed the chlorine concentration at a different level to know which concentration was effective in killing microorganisms. As a result, at least 25 ppm was proved to be necessary. We used a minimum 25 ppm chlorine concentration in the greenhouse. The maximum level of chlorine concentration, that the SAEW generating machine could be produced, was 40 ppm. So, we used 25-40 ppm for greenhouse spray.

 

  1. In the Materials and Methods section, describe how the results were statistically processed.

Response 4. We added one section, section 2.9. Statical analysis.

 

  1. Figure 3 is missing.

Response 55. The submitted manuscript contained Figure 3. However, it was missing for some reason. We added Figure 3.

 

  1. Figure 4 does not show margins of error (error bars) for live cells.

Response 6. Live cells were calculated based on the dead cell numbers. So, we could not show the standard errors for live cells.

 

  1. Explain why there are such large spreads of values for dead cells.

Response 7. We counted the total and dead cells on the membrane under the fluorescent microscope. Cells were not distributed evenly on the membrane. Cell numbers differed from microscope images to images, which made a large variation.

 

  1. Figure 9 is missing.

Response 9. Sorry for it. We forgot to add Figure 9 in the text. We added Figure 9.

 

  1. There is no discussion chapter.

Response 9. We made a discussion section.

 

  1. The conclusion that SAEW successfully reduces airborne microorganisms is debatable, as Figure 4 shows that the statistically significant difference between treatment and control was not significant.

Response 10. We believe that SAEW spraying increased dead cells dramatically. Different from in vitro experiments, it was impossible to kill microorganisms completely in the greenhouse. In comparison with tap water spraying, SAEW spraying significantly increased dead cells. All indicated that SAEW spraying is an effective method to reduce the microorganism in the greenhouse.

 

  1. In general, the manuscript looks raw and unfinished. In my opinion it should be strengthened by additional studies of the effect of slightly acidic electrolysis water solution of various concentrations on various microorganisms in the greenhouse.

Response 11. We used enough chlorine concentration calculated from in vitro experiments. As a control, tap water spraying was appropriate, so we do not think to lower the chlorine concentration in the greenhouse experiment.

We cannot control the species of microorganisms in the greenhouse. SAEW has no selectivity of microorganisms for its effect. We used a small container with a specific bacterium, such as E. coli, and sprayed SAEW. E. coli cells were reduced very much. This kind of experiment was not applicable in the actual greenhouse.

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

The current version of the manuscript is much better than the previous one, but the discussion section needs to be expanded. A discussion based on 3 literature items is not enough.

 

Author Response

Thank you for your review. 

We rewrote the discussion part with several references according to your comments.  Please see line 226-234, line 238-243 and line 250-252. The modified parts are shown in red.

Reviewer 2 Report

I have no comments, thanks!

Author Response

Thank you for your review. We are very much appreciated.

Reviewer 3 Report

The authors revised the article, but did not take into account all the comments.

The annotation should be expanded. Background: Place the question addressed in a broad context and highlight the purpose of the study.

Please check the text formatting for MDPI format.

Author Response

Thank you for your review. We modified the manuscript according to your comments. The modified parts are shown in red.

1. The annotation should be expanded.

Response 1. We are sorry, but we could not catch the point. If this means the abbreviations, we added several abbreviations in the text.

Luria-Bertani (LB, 10 g bacto peptone, 5g yeast extract, 5 g NaCl in 1 L)

phosphate-buffered saline (PBS)

2. Background: Place the question addressed in a broad context and highlight the purpose of the study.

Response 2. We rewrote the Introduction.

3. Please check the text formatting for MDPI format.

Response 3. We asked English editing supplied by MDPI and checked the MDPI format.

 

 

Round 3

Reviewer 1 Report

The manuscript has been revised as suggested by me. It is printable in its present form.