Review Reports - The Main Compounds of Bio-Fumigant Plants and Their Role in Controlling the Root-Knot Nematode <i>Meloidogyne incognita</i> (Kofoid and White) Chitwood

Round 1

Reviewer 1 Report (New Reviewer)

Comments and Suggestions for Authors

It was with great interest that I agreed to review the article titled. However, after the very first sentences of the article, my enthusiasm gave way to irritation. The manuscript is written in a very poor style. The level of language makes it impossible to focus on the substantive content of the article. There are so many mistakes and poorly constructed sentences that the paper requires a huge amount of proofreading, practically writing from scratch. Some of the sentences look like they have been torn off in the middle, or joined together from two different sentences. The original topic and abstract in the system suggest the use of two plants as a source of potentially nematode-controlling materials. Only one plant already appears in the manuscript itself. The title of the article is very philosophical (almost like "to be or not to be") and in my opinion inappropriate for any scientific article. In my opinion, as presented, the manuscript is not suitable for publication in Agronomy.

Comments on the Quality of English Language

The manuscript is written in a very poor style. The level of language makes it impossible to focus on the substantive content of the article. There are so many mistakes and poorly constructed sentences that the paper requires a huge amount of proofreading, practically writing from scratch. Some of the sentences look like they have been torn off in the middle, or joined together from two different sentences.

Author Response

Answer: Thank you very much for your comments. Deficiencies related to the writing were removed based on the native English editing of the manuscript. The reviewed manuscript was resubmitted after major editing and revision based on the respected reviewers' comments. Please note that title and abstract of the previous version of the manuscript were incorrectly submitted, and all will be sent correctly by the second revision. Based on your comment, the title was improved as “The main compounds of bio-fumigant plants and their role in controlling root-knot nematode Meloidogyne incognita (Kofoid and White) Chitwood”.

Hope the major revision of the manuscript can removed your concerns.

Reviewer 2 Report (Previous Reviewer 4)

Comments and Suggestions for Authors

The manuscript has changed significantly. Ageratina adenophora (Syn. Eupatorium adenophorum) has been excluded as stated in the answer.But there are important aspects in which I do not agree and others that are not clear and some errors.

I do not agree with what is said in lines 17-21 of the abstract. In this work, the effect of the compounds methylene chloride, 2-decenal, linalool, and nonanal is studied. The composition and biofumigant effect of the leaves and stems-roots of tomato plants are also studied. But the effect of the compounds: α-pinene, d-limonene, butanol, cyclotetrasiloxane, alanine ethylamide, and hexanol, as stated in the abstract, is not particularly studied. These compounds are part of vegetables, but their particular effect is not studied. The correlations that may have been observed with Principal Component Analysis (PCA) do not necessarily imply causality. In observational data, correlation does not imply causation. It would be necessary to experiment with these substances. Therefore, it cannot be stated that α-pinene, d-limonene, had very significant nematicidal effects, or that butanol caused an increase in the population of the second-stage juveniles.

I also do not agree with part of what is indicated in the conclusions. In line 1555-1556 it is said that "The main compounds studied in this research showed that by having a special character such as nematicidal effect or attraction" or in line 1559 "we introduce linalool as a chemical compound with attraction properties". Or in the abstract "Linalool and nonanal have attraction properties". In this work, the attraction effect towards the crop has not been tested, only mortality has been tested. And what is observed is that an increase in the dose of linalool or nonanal represents a decrease in mortality, but not an attraction towards the crop. I consider that it can be expressed both in conclusions and in the abstract, since they are not derived from experimental measurements and observation

In materials and methods 2.5. Parameter measurement . The indicated mortality corrected formula is usually cited as the Schneider-Orelli formula (Schneider-Orelli O. Entomologisches Praktikum. 2nd ed. Sauerländer; Aarau, Switzerland: 1947. pp. 1–237) (as you can see at https://www.ehabsoft.com/ldpline/onlinecontrol.htm#SchneiderOrelli https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7411722/#B16-plants-09-00839.) However, it does not seem that in the ANOVAs (Tables 1 and 5) the mortalities corrected with the control were used, as indicated in 2.5. The degrees of degrees of freedom of the Concentration factor are 5 (so 6 dose levels are considered, therefore dose 0 has been included), but if mortality is corrected with the control (or dose 0) it should not be include this dose in the ANOVA. The mortality corrected with the control has not been used in the regression adjustments of figures 1 to 4 either, since at dose 0 there is mortality, but if it had been corrected with the control this would be 0.

Line 18 and others. It says "methylen chloride" it should say "methylene chloride".

Line 523. For which model has each software been used? What R packages and functions were the calculations performed with?

Line 527. The formula called lognormal is not written correctly. When substituting some of the values obtained in table 2 into the formula, for example Linalool 1 week (LNMRMax = 20.6, slope = 103.6, ECmax = 16.4), I cannot obtain a curve similar to the one obtained in graph 1.

Line 595 repeats "temperature × concentrations" interaction

Since the factors time, temperature and concentration and their double interactions are significant, I think that it would not be appropriate to make the adjustments indicated in Tables 2 and 3 and graphs 1 and 2 since the effect of the temperature and time factors respectively is confused. It would be more appropriate to make adjustments for each substance, temperature and time, as in figure 3, but also including linalool and nonanal.

Figure 3. In materials and methods it says that it is tested with concentrations of 400–25 μg ml-1os (line 497) . Why do concentrations of 80, 160, 240, 320 and 400 μg ml-1 appear in Figure 3?

Lines 892-896. All factors and interactions are significant according to Table 5. Review the wording of the paragraph.

Line 901 and 935. It says "Table 5" , it should say "Table 6"

Lines 936-938. This paragraph should come before the previous paragraph.

Lines 1107-1108. I don't understand what the percentages correspond to.

I think an in-depth review of the text is necessary.

Best regards

Author Response

The manuscript has changed significantly. Ageratina adenophora (Syn. Eupatorium adenophorum) has been excluded as stated in the answer. But there are important aspects in which I do not agree and others that are not clear and some errors.

Answer: Thank you very much for your valuable comments on the manuscript. We have considered your constructive comments to improve the quality of this manuscript.

Line 17-21: I do not agree with what is said in lines 17-21 of the abstract. In this work, the effect of the compounds methylene chloride, 2-decenal, linalool, and nonanal is studied. The composition and biofumigant effect of the leaves and stems-roots of tomato plants are also studied. But the effect of the compounds: α-pinene, d-limonene, butanol, cyclotetrasiloxane, alanine ethylamide, and hexanol, as stated in the abstract, is not particularly studied. These compounds are part of vegetables, but their particular effect is not studied.

Answer: The content of the abstract and keywords were changed based on your comment;

Meloidogyne spp. are important parasitic nematodes that attack various plant species in the world. The focus of the current research was on the study of the main compounds of bio-fumigant plants and the recognition of the characteristics of four compounds in nematode control, that were lin-alool, nonanal, methylene chloride, and 2-Decanal. In a laboratory study methylene chloride and2-Decenal caused decrease the population of the second-stage juveniles (J2s). In another part of study on the nematicidal effects of tomato Solanum lycopersicum L. on the J2s of M. incognita, it was found that leaves of tomato increased mortality percentage of J2s, however in root-stems treatment the concentration increase, decreased the mortality percentage in all times and tem-peratures. The maximum mortality response (MRmax) and half maximal effective concentration (EC50) were estimated about 100% and 4.0 µg/mg in temperature 35 °C and in week 8 in leaves treatment while corresponding values of root-stems treatment were 13. 5% and 3.0±1.7 µg/mg, respectively. In GC-MS analysis of both treatments dried root-stems and leaves of tomato main compounds, α-pinene, d-limonene, butanol, linalool, alanine ethylamide, cyclotetrasiloxane and hexanol, were identified. Results showed that tomato leaves can be used as bio-fumigants plant to control root-knot nematodes.

Keywords: Solanum lycopersicum; Meloidogyne incognita; α-Pinene; D-limonene; Methylene chloride; Linalool; Nonanal; Butanol

Line 421 and Line 432: The correlations that may have been observed with Principal Component Analysis (PCA) do not necessarily imply causality. In observational data, correlation does not imply causation. It would be necessary to experiment with these substances. Therefore, it cannot be stated that α-pinene, d-limonene, had very significant nematicidal effects, or that butanol caused an increase in the population of the second-stage juveniles

Answer: Modified as these.

Line 426: “According to analysis (PCA), α-pinene, d-limonene and methylene chloride extracted from leaves of S. lycopersicum”

Line 437: “According to analysis (PCA), linalool and butanol extracted from root-stems of S. lycopersicum”

Answer: The content of the conclusions was re-written;

“Although RKN cause a large amount of damage and reduce the yield of tomato, interestingly, it was observed that the dried leaves of S. lycopersicum as a bio-fumigant have the potential to control J2s of M. incognita. Two main compounds studied in this research, methylene chloride and 2-Decenal, with their nematicidal effect character, are very suitable compounds for the production of non-chemical nematicides. Without any doubt, the use of non-chemical nematicides are eco-friendly products and are very suitable to be used in IPM program.”

Also, regarding this comment, part of the discussion was also changed;

“Line 453-458: In a recent study, the percentage mortality of M. incognita J2 was decreased in the root-stems treatment of S. lycopersicum. One of the main compounds identified in the root-stems of S. lycopersicum was linalool. The results obtained from of linalool research in previous studies indicated that linalool has attraction features. According to these findings, it can be assumed that the tomato root-stems due to having linalool has the potentiality to attract M. incognita J2”

And

“Line 477-479: Based on these trap plants studies, it can be assumed that the S. lycopersicum can be used as a trap plant in greenhouses. Of course, to prove this hypothesis, many laboratory and greenhouse studies are needed, which we hope be done in a very near future.”

In materials and methods 2.5. Parameter measurement. The indicated mortality corrected formula is usually cited as the Schneider-Orelli formula (Schneider-Orelli O. Entomologisches Praktikum. 2nd ed. Sauerländer; Aarau, Switzerland: 1947. pp. 1–237) (as you can see at https://www.ehabsoft.com/ldpline/onlinecontrol.htm#SchneiderOrelli https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7411722/#B16-plants-09-00839.). The degrees of degrees of freedom of the Concentration factor are 5 (so 6 dose levels are considered, therefore dose 0 has been included), but if mortality is corrected with the control (or dose 0) it should not be include this dose in the ANOVA. The mortality corrected with the control has not been used in the regression adjustments of figures 1 to 4 either, since at dose 0 there is mortality, but if it had been corrected with the control this would be 0.

Answer: Thank you so much for your best comment. The formula was changed. Accordingly (Schneider & Orelli 1947) it was re-written;

Mortality (%) = number of dead nematodes/total number of nematodes × 100

To quantify mortality changes using regression models, we required a control (zero) for each treatment. Consequently, deaths among the deceased were not compared to the control, and we needed concentrations ranging from 0 to 400 µg/ml in the laboratory section and from 0 to 100 in the second section to achieve a more accurate regression fit.

Line 18 and others: It says "methylen chloride" it should say "methylene chloride".

Answer: Corrected as "methylene chloride".

Line 523: For which model has each software been used? What R packages and functions were the calculations performed with?

Answer: We use of packages drc, tidyverse, ggplot2 and caret for fit regression and plots.

Line 527: The formula called lognormal is not written correctly. When substituting some of the values obtained in table 2 into the formula, for example Linalool 1 week (LNMRMax = 20.6, slope = 103.6, ECmax = 16.4), I cannot obtain a curve similar to the one obtained in graph 1.

Answer: It was selected from sigma plot to fit this model:

Line 595: repeats "temperature × concentrations" interaction

Answer: It was removed.

Line 249: Since the factors time, temperature and concentration and their double interactions are significant, I think that it would not be appropriate to make the adjustments indicated in Tables 2 and 3 and graphs 1 and 2 since the effect of the temperature and time factors respectively is confused. It would be more appropriate to make adjustment for each substance, temperature and time, as in figure 3, but also including linalool and nonanal.

Answer: In the case of chemical compounds, due to the fact that in the two compounds Methylene chloride and Decenal, the triple effects of time * temperature * concentration have become significant, it was possible to draw Figure 3, but in the case of Linalool and Nonanal, only the interaction of temperature * concentration and concentration *time became significant and it is possible to draw figures 1 and 2. In order to better compare all four compounds with each other, concentration*time and concentration*time models were drawn for all four compounds, but it is not possible to draw figures 3 for all compounds due to the significance of the three effects of time* temperature* concentration.

Line 305: Figure 3. In materials and methods it says that it is tested with concentrations of 400–25 μg ml-1os (line 497). Why do concentrations of 80, 160, 240, 320 and 400 μg ml-1 appear in Figure 3?

Answer: Modified as these

Lines 892-896: All factors and interactions are significant according to Table 5. Review the wording of the paragraph.

Answer: Modified as these.

Line 901 and 935: It says "Table 5", it should say "Table 6"

Answer: Modified as these.

Lines 936-938: This paragraph should come before the previous paragraph.

Answer: Modified as this.

Lines 1107-1108. I don't understand what the percentages correspond to.

Answer: It was modified.

I think an in-depth review of the text is necessary.

Answer: The text was thoroughly revised. Thank you again.

Reviewer 3 Report (New Reviewer)

Comments and Suggestions for Authors

The article titled ‘The Main Compounds of Bio-fumigant Plants and Their Important Role in Controlling or Not Controlling Root-knot Nematode Meloidogyne incognita (Kofoid and White) Chitwood’ extends an authors' previous work, presenting a comprehensive study on the nematicidal properties of various compounds extracted from biofumigant plants against the nematode Meloidogyne incognita. It builds upon a study published in this journal last year, employing a detailed methodology encompassing compound chemistry analysis and in vitro experimentation to assess biofumigants' effectiveness in managing nematodes in tomato plants. The study's implications are noteworthy for agricultural nematode control, particularly in tomato cultivation, suggesting more sustainable alternatives to traditional chemical treatments.

The manuscript offers a coherent and well-contextualized study, appropriately citing recent and relevant literature. The methodology is thorough, featuring a replicable experimental design and suitable statistical analyses, including ANOVA, nonlinear regression models, and Principal Component Analysis.

However, the manuscript requires attention to enhance its clarity and coherence. Specific areas needing improvement include:

Language and Grammar:

The manuscript contains grammatical errors and awkward sentence structures that hinder clarity. It would benefit from a thorough language review to correct these issues.

Consistency in Terminology:

I understand that the term 'methylen chloride' is a typographical error and should be replaced by 'methylene chloride'. There are also inconsistencies in chemical names (e.g., '2-decenal' vs. '2-Decenal' in L18 and L152, respectively) and usage of abbreviations. The term 'root-knot nematodes' and its abbreviation 'RKN' are used inconsistently. The first reference is in L43, but the abbreviation 'RKN' is noted for the first time in L84 without any association with the concept. Arbitrarily, root-knot nematodes are also mentioned without abbreviation in lines 71, 87, 460, 477, and 481. Similar issues are observed with second-stage juveniles (J2s), gas chromatography (GC), and Principal Component Analysis (PCA).

Formatting and Style:

Inconsistencies in formatting, such as different ways of referring to 'greenhouse soil', need to be standardized throughout the document. Also, attention should be paid to proper punctuation, especially in lists (e.g., '1, 2, 3, 4, and 8 weeks').

Content Balance and Tone:

The introduction and discussion sections are well-written, but certain colloquial expressions (e.g., 'plant nematologists have spent a lot of time studying...' in L51) should be replaced with more formal academic language. I also find the last two sentences of the introduction (L82 to L87) irrelevant.

Methodology and Results Sections:

These sections, in particular, suffer from long, convoluted sentences (e.g., between lines 261-266 or 278-283) and a higher frequency of typographical errors. A detailed review is necessary to ensure clarity and accuracy.

Minor comments:

Ø L29: ‘root-knot nematodes’ instead of ‘root-knot nematode’

Ø L134: A comma is missing between ‘thermo-gravimetric’ and ‘first’

Ø L139: The plural form should not be used when using the term ‘J2’ as an adjective

Ø L156-166: Very confusing paragraph, perhaps because it is not yet the final edition?

Ø L310: It is advisable to avoid abbreviations in headings

Ø L346: ‘at all times and temperatures’ instead of ‘in all time and temperature’

Ø L366: ‘soil’ instead of ‘soli’

Ø L407: Remove the comma between ‘plants’ and ‘play’

Ø L410: ‘four’ instead of ‘4’

Ø L422: ‘α-pinene and d-limonene, extracted from leaves of S. lycopersicum, caused..’ instead of ‘α-pinene, d-limonene extracted from leaves of S. lycopersicum caused..’

Ø L423: Two verbs used in tandem: ‘reported’ and ‘introduced’. Choose one of them.

Ø L451: ‘and’ instead of ‘+’

Ø L457: A comma is missing between ’13.5%’ and ‘respectively’

Ø L481: The word 'nematodes' is redundant as it is included in the abbreviation ‘RKN’

Ø L485: ‘its capacity to decrease’ instead of ‘its decrease’

Ø L486: Unreadable line

In summary, while the study is promising and contributes valuable insights to the field, the manuscript requires substantial editing for language, consistency, and formatting. Attention to these details will significantly enhance the manuscript's readability and suitability for publication in Agriculture if considering the proposed suggestions.

Comments for author File: Comments.pdf

Comments on the Quality of English Language

Upon review, it is evident that the manuscript requires improving the quality of the English language and stylistic presentation. Here are some specific areas that need attention:

1. Grammar and Syntax: The text has grammatical errors and awkward sentence constructions. These issues distract from the content and can lead to misunderstandings of the scientific information presented.

2. Clarity and Conciseness: Several sentences are overly long and convoluted, making them challenging to follow. Shortening and simplifying these sentences can significantly improve the overall readability of your manuscript.

3. Formatting: The manuscript exhibits some inconsistencies in formatting, particularly in the punctuation in lists. Adhering to a standard format throughout will enhance the manuscript’s overall quality.

Improving the English language quality of your manuscript is crucial for its acceptance in an international journal, as it not only helps convey your research more effectively but also ensures that the focus remains on the scientific contributions of your work. I recommend engaging a professional scientific editing service or a native English-speaking colleague specializing in your field to thoroughly review and edit the manuscript. This step will significantly aid in clarifying your findings and arguments, enhancing the manuscript's chances for successful publication.

Author Response

However, the manuscript requires attention to enhance its clarity and coherence. Specific areas needing improvement include:

Answer: It is our pleasure that our findings are acceptable for you. We are very grateful for your constructive comments. All the comments have been considered, which was undoubtedly improved the quality of this manuscript.

Language and Grammar: The manuscript contains grammatical errors and awkward sentence structures that hinder clarity. It would benefit from a thorough language review to correct these issues.

Answer: The manuscript was thoroughly revised for English writing.

Consistency in Terminology:

Arbitrarily, root-knot nematodes are also mentioned without abbreviation in lines 71, 87, 460, 477, and 481. Similar issues are observed with second-stage juveniles (J2s), gas chromatography (GC), and Principal Component Analysis (PCA).

Answer: Modified as these.

Formatting and Style:

Answer: Modified as these

Content Balance and Tone:

Answer: Modified as these

Methodology and Results Sections:

Answer: Modified as this.

Minor comments:

Line 29: ‘root-knot nematodes’ instead of ‘root-knot nematode’

Answer: Modified as this.

Line 134: A comma is missing between ‘thermo-gravimetric’ and ‘first’

Answer: Modified as this.

Line 139: The plural form should not be used when using the term ‘J2’ as an adjective

Answer: Modified as this.

L156-166: Very confusing paragraph, perhaps because it is not yet the final edition?

Answer: The content of this paragraph was re-written;

“To obtain a stock solution of 1000 µg ml^-1, each of the chemical compounds was weighed by 0.01 g and dissolved in 10 ml of acetone. According to serial dilution 800–50 µg ml^-1 of stock solution was removed then it was used from 0.5 % Tween 80 as emulsifier to provide a volume of 10 ml. For the serial dilution, formula was used”

Line 310: It is advisable to avoid abbreviations in headings

Answer: Modified as this. Accordingly, it was re-written; In Vitro Mortality of second-stage juveniles

Line 346: ‘at all times and temperatures’ instead of ‘in all time and temperature’

Answer: Modified as this.

Line 366: ‘soil’ instead of ‘soli’

Answer: Corrected as soil.

Line 407: Remove the comma between ‘plants’ and ‘play’

Answer: Corrected as plants play.

Line 410: ‘four’ instead of ‘4’

Answer: Corrected as four

Line 422: ‘α-pinene and d-limonene, extracted from leaves of S. lycopersicum, caused..’ instead of ‘α-pinene, d-limonene extracted from leaves of S. lycopersicum caused..’

Answer: Corrected as α-pinene and d-limonene, extracted from leaves of S. lycopersicum, caused.

Line 423: Two verbs used in tandem: ‘reported’ and ‘introduced’. Choose one of them.

Answer: corrected as reported.

Line 451: ‘and’ instead of ‘+’

Answer: Corrected as and

Line 457: A comma is missing between ’13.5%’ and ‘respectively’

Answer: Corrected as ‘,”

Line 481: The word 'nematodes' is redundant as it is included in the abbreviation ‘RKN’

Answer: Modified as this.

Line 485: ‘its capacity to decrease’ instead of ‘its decrease’

Answer: Modified as this.

Line 486: Unreadable line

Answer: Modified as this. Thank you again.

Round 2

Reviewer 1 Report (New Reviewer)

Comments and Suggestions for Authors

No additional comments.

Comments on the Quality of English Language

No comments.

Author Response

Dear Editors,

We truly would like to thank you for your second review that helped us improve our manuscript. We have considered all comments and modified manuscript according to them. Please see the comments, our replies, and modifications as follows;

The modified parts were highlighted with two different colors: (Yellow) and (dark green) colors for the second and third reviewers, respectively.

Reviewer 2 Report (Previous Reviewer 4)

Comments and Suggestions for Authors

I have serious doubts about non-linear regression fits to the models. Either the indicated formulas are wrong, or the results in the tables are incorrectly expressed. I attach an Excel file where I put the formulas, with some of the parameters obtained by regression. The results I obtain do not coincide with those expressed in the graphs.

Also in Figure 3, in the previous version of the manuscript I showed my surprise that "The points of the observed values appeared at the concentrations of 80, 160, 240, 320 and 400 μg ml-1". Now in Figure 3, the concentrations are those indicated in materials and methods and the lines are said to be the values predicted with a Gompertz model. But they are not curves, they are simply straight lines that join the observed values.

I think that the non-linear regressions, the equations of lines 190-192, tables 2 to 4 and figures 1 to 3, and the changes that may arise from them, must be reviewed in depth.

Sincerely

Comments for author File: Comments.7z

Author Response

Also, in Figure 3, in the previous version of the manuscript I showed my surprise that "The points of the observed values appeared at the concentrations of 80, 160, 240, 320 and 400 μg ml-1". Now in Figure 3, the concentrations are those indicated in materials and methods and the lines are said to be the values predicted with a Gompertz model. But they are not curves, they are simply straight lines that join the observed values.

I think that the non-linear regressions, the equations of lines 190-192, tables 2 to 4 and figures 1 to 3, and the changes that may arise from them, must be reviewed in depth.

Sincerely

Answer: According to the opinions and comments of the respected reviewer related to the objections made to the non-linear regression models and the investigations, changes were made in the tables and figures characterized by yellow highlights.

The first change was made on in the Log-Normal model to Weibull. In this manuscript, we fitted several types of models and finally the best model was transferred to the manuscript. In the case of Linalool and Nonanal compounds, two Log-Normal and Weibull models were fitted. Unfortunately, in the material and methods and results, the Log-Normal formulas were mistakenly replaced by the Weibull model. According to the reviewer's comments and the excels he sent, a more detailed analysis was done on the initial analysis files and the problem was fixed. Material and methods were modified according to the Weibull model.

Another important point about this model is that in the outputs of our model, only the parameters that are important for interpretation and also have a biological definition were used. Due to reviewer Excel model for the correctness, and to respect him, parameters such as c and MR_min, width of the origin of the model, not being used before interpretation, was added to the tables.

Regarding the second objection to Figure 3 and the mistakes in the numbers of concentrations,80, 160, 240, 320 and 400 in the initial version, the wrong numbers were corrected. Accurate and correct concentrations obtained from the original stock, which was in the range of 25 to 400, were used and the models were based on this in Figure 3. In re-examining this figure and the outputs of the model in the new version of Figure 3, the models are fitted again and the new outputs are included in Table 4.

The Excel file related to the regression models was completed and sent to the honorable reviewer, please find in the attachment. It should be mentioned that the regression models, in the manuscript, were performed by Sigmaplot software. Being thank full to the reviewer, all his opinions and comments were employed to correct the mistakes.

We would like to sincerely thank and appreciate the reviewer for his comments and feedback.

Reviewer 3 Report (New Reviewer)

Comments and Suggestions for Authors

Thank you for your thorough response to the review comments on your manuscript titled "The Main Compounds of Bio-fumigant Plants and Their Important Role in Controlling or Not Controlling Root-knot Nematode Meloidogyne incognita (Kofoid and White) Chitwood."

I am pleased you have corrected the grammatical errors, terminology inconsistencies, formatting issues, and other aspects highlighted in the review. Your effort in revising the language, ensuring consistency in terminology, and refining the content structure is commendable and has undoubtedly improved the overall quality of your manuscript.

However, I want to reemphasize the importance of further improving the English language quality of the text. While the revisions have addressed many initial concerns, enhancing the manuscript's readability and flow through additional language editing will be beneficial, being particularly important for ensuring that your valuable research is accessible and clearly understood by an international audience.

I am enclosing the manuscript with some suggestions and corrections, including the editing of some paragraphs, which will help the authors improve the quality of the text.

Comments for author File: Comments.pdf

Comments on the Quality of English Language

To this end, I recommend a final manuscript review by a professional, scientific editing service or a native English-speaking colleague with expertise in your field. This step will help to polish the language further and ensure that your study is presented in the best possible manner.

Author Response

Due to the lengthy reply, I have added it to the attachment. Thank you！

Author Response File: Author Response.pdf

This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

My specific comments and corrections can be found in the attached PDF. The text is generally understandable, but the manuscript would benefit from editing by a language expert or native speaker.

In summary, after major revisions, the manuscript can be considered for publication in Agriculture.

Comments for author File: Comments.pdf

Comments on the Quality of English Language

The manuscript would benefit from editing by a language expert or native speaker

Reviewer 2 Report

Comments and Suggestions for Authors

The subject is essential and worth publication. Yet, the authors should have examined their data and methodology more thoroughly. For example, nothing was reported about the greenhouse experiment in materials and methods that need to be rewritten in detail. Moreover, the writing style is awkward and more difficult to understand also, the grammar is abysmal in this manuscript. The authors present statistical analysis methods more than research results, which readers find difficult to understand. How to use natural sources and then use commercial compounds? Why weren't those compounds extracted, separated, and used? Also, the discussion is misleading and needs to be clearer. See also the comments inside the manuscript fileز

Note: in the article title: the Nematicidal not Nemticidal

Comments for author File: Comments.pdf

Comments on the Quality of English Language

I strongly suggest that the writer of this manuscript give it to the English editing office for assistance in grammar, style, and sentence structure.

Reviewer 3 Report

Comments and Suggestions for Authors

see comments inside document

Comments for author File: Comments.pdf

Reviewer 4 Report

Comments and Suggestions for Authors

The topic is interesting, since it tries to provide knowledge for the control of Meloidogyne, one of the main phytopathological problems of many crops. However, the experimental design does not seem the most appropriate. I do not understand the selection of the chemical compounds tested, taking into account that previous research has been carried out (https://www.mdpi.com/2077-0472/13/6/1109) in which it is revealed that the J2 of M .incognita were more sensitive to the root-stem treatment than to the leaves and, however, instead of carrying out tests with the chemical compounds present in the root-stem, they tested with the compounds extracted from the leaves of E. adenophorum, which they are not present in the root-stem. In materials and methods important information is omitted. And the discussion is poor and barely reports previous results, and does not contribute new ideas additional to those of the previous work.

Aspects that need to be reviewed and clarified.

Eupatorium adenophorum is a synonym of Ageratina adenophora (Spreng.) R.M.King & H.Rob. as you can see in https://www.worldfloraonline.org/taxon/wfo-0000059952 and https://powo.science.kew.org/taxon/urn:lsid:ipni.org:names:6811-2 You must put the current name.

Line 59. It should be indicated that it belongs to the Asteraceae family.

Line 85-86. Check parenthese.

Line 98. It is not specified which parts of the tomato samples are analyzed, nor what extraction protocol is used.

Line 118. "250 m2 of soil", isn't it 250 g or 250 cm3?

Lines 139-140. It is not indicated whether the same is done with Eupatorium adenophorum.

Line 142. Are the tubes incubated open or closed? Are they incubated in light or in the dark?.

Lines 143-144. I don't understand what "The tubes were incubated at 25 and 35 °C in two cycles including 4 weeks" means. Does it mean that two cycles have been tested? What is a cycle? What is the relationship between cycle 1 and cycle 2? Or that all the samples have first been incubated at 25 and then at 35? It should be clarified.

Lines 152-160. The composition of the emulsion is very confusing. Why is the concentration of the four chemical compounds not expressed by weight of soil, in the same way that the plant material is expressed?.

Line 161. "... stored for 2 cycles including 4 weeks" Same comment as the one made for lines 143-144.

Lines 163-164. "The percent of M. incognita J2 juvenile mortality was calculated for each chemical using the equation according to Mao et al. 2014 [25] method" can be eliminated. It is explained in point 2.5.

Line 178. Abbreviate to M. incognita and in italics

Lines 175 - 193. Item 2.6 Data analysis.

It does not indicate how the anova of tables 1 and 3 is performed. Is mortality in % used as response variable or is some transformation performed? What software is used? It has been verified whether the assumptions of the anova of homoscedasticity and normality of the residuals are met?.

Nor are they explained with which software and methods or functions the nonlinear regression models are adjusted.

Line 180. Sigmoid is a generic type of curves. The equation shown corresponds to Gompertz curve.

Line 181. This equation corresponds to log-logistic curve. I recommend you consult Ritz (2010) (At https://doi.org/10.1002/etc.7 and at https://setac.onlinelibrary.wiley.com/doi/10.1002/etc.7) to know the formulas in detail of the dose–response models that are used in ecotoxicology.

Line 182. The Weibull equation is incorrect. See Ritz (2010). I believe that it would not be necessary to include the three aforementioned formulas, as long as the models used, the statistical packages, and the options used for their calculation are detailed.

Line 183-184 (and line 231) It says "EC50: half maximal effective concentration". It shoul say "EC50: Concentration producing a response half-way between the minimum and the maximum". It should not be confused with the dose that causes mortality in 50% of the individuals, since in some of the cases shown in the work 100% mortality is not reached, and this can be misleading.

Lines 203 and 270. It says: "Table X. In vitro analysis of variance....". It should say "Table X. Analysis of variance...."

Line 208, 214, 242, 275, 294. It says "comperze". It should say "Gompertz"

Tables 2 and 4. Figures 1 to 4. It should be indicated to which model the estimated parameters and the curves represented correspond.

Figures 1 to 4. They should be made larger, and with the same scale on the Y axis for visual comparison. Why don't the three replicates appear in each concentration?

Line 339. It should be indicated in the legend of table 5 that the results for E. adenophorum come from previous research [13].

Line 355. Where it says "lg" it should say "µg".

Lines 360-366. They don't contribute much to the discussion.

References. Check the scientific names of the species, there are some that are not written in italics.

Best regards