Review Reports

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Pesticides are a broad group that includes chemicals biologically active substances used in plant protection products. Distinctions include: zoocides to counter animal pests, herbicides to control weeds, bactericides to fight bacteria, and fungicides to eliminate microscopic fungi. With industrial, and also scientific and technological development, the use of pesticides in agriculture is increasing worldwide. Despite the effectiveness of these compounds in plant protection, there is still a lot of interest in the side effects and health consequences of pesticide use. The reviewed work fits into this direction. Main goal of reviewed manuscript is to provide an up-to-date, comprehensive review on the use, degradation, exposure and effects on gut health of these four EU approved pesticides, glyphosate, propamocarb, deltamethrin and tebuconazole.

This is a comprehensive review paper, including the authors' subjective view of the issues involved impacts on gut health primarily the gut microbiota, inflammation, metabolism, cancer and gut-brain axis selected pesticides such as glyphosate, propamocarb, deltamethrin and tebuconazole.

The paper is based on a carefully selected 154 papers from the last 10 years or so, although there are some cited papers from the early 2000s.
The reviewed paper is written in correct language and reads well. 
I have a few comments to the Authors:
1. why were these four pesticides chosen for discussion. We are in the year 2025, the time for three of them to be used passes this year, or next year. Considering the time of their perisistence in the environment, others could be indicated. Please explain in more detail the criteria for their selection.
2. line 178, Tert with a lowercase letter,
3. line 201, why is AMPA listed as the only degradation product besides the 4 basic pesticides discussed?
4. line 457, move the table heading lower, directly above the table,
5. line 1305, what does ...

 

 

Author Response

We sincerely appreciate the reviewers’ thoughtful questions and feedback, which have helped us improve the quality of our review.

1. why were these four pesticides chosen for discussion. We are in the year 2025, the time for three of them to be used passes this year, or next year. Considering the time of their perisistence in the environment, others could be indicated. Please explain in more detail the criteria for their selection.

The choice of these four specific pesticides was driven by several key considerations:

1. Relevance to Current Regulatory Timelines: While it is true that the EU approvals for glyphosate (2025), propamocarb (2025), deltamethrin (2025), and tebuconazole (2026) are nearing expiration, this makes them especially relevant for discussion. Regulatory authorities such as EFSA often reassess substances before renewing or banning the pesticide. By reviewing recent studies on how people are exposed to these pesticides and how they may affect gut health, our goal was to provide useful information that could help guide future regulatory decisions. Additionally, even if their approval expires, it is possible that agricultural workers may continue using existing stocks they have already purchased, extending potential environmental and health exposure beyond the official phase-out date.
2. Representation of Different Chemical Classes and Modes of Action: It is important to ensure a diverse representation of chemical classes with different biological targets. Glyphosate is an organophosphate herbicide, deltamethrin a pyrethroid insecticide, propamocarb a carbamate fungicide, and tebuconazole a triazole fungicide. This variety allows for a broader perspective on how different types of pesticides might differentially impact gut health.
3. Research Relevance and Knowledge Gaps: Some of these pesticides are also under investigation in our laboratory, making their selection particularly relevant to identify knowledge gaps in the literature. This helps us and potentially other researchers target areas that require further study.
4. Environmental and Health Persistence: All four pesticides have been detected in food or water samples in Europe as well as in human biomonitoring studies, emphasizing the real-world relevance of evaluating their potential effects on human health.

We agree that additional pesticides could have been included and encourage future reviews to build upon this work by incorporating other widely used or persistent pesticides. Our aim was to focus on a selected group of pesticides that represents different types and remains highly relevant and useful at this time.

2. line 178, Tert with a lowercase letter

Noted. 

3. line 201, why is AMPA listed as the only degradation product besides the 4 basic pesticides discussed?

We amended the sentence to: ‘Glyphosate, deltamethrin, propamocarb, tebuconazole and some of their degradation products have been detected in drinking water, fruits, vegetables, legumes, honey, cereals, infant formula, alcoholic beverages and animal products (Bellisai et al., 2023; Muñoz et al., 2023b; Nijssen et al., 2024; Nougadère et al., 2020).’

4. line 457, move the table heading lower, directly above the table

Due to the document's formatting and the table being embedded in a layout format, it is not possible to place the table title directly above the table.

5. The question is incomplete

 

 

Author Response File: Author Response.docx

Reviewer 2 Report

Comments and Suggestions for Authors

Pesticides and their metabolites can be found in soil, air, water, groundwater, and food products. The authors described the documented effects of compounds such as glyphosate, deltamethrin, propamocarb, and tebuconazole on human health, especially metabolism, gut microbiota, and the gut-brain axis.

The motivation for this review was based on the health risks associated with exposure to the pesticides mentioned above, which may contribute to a range of health problems, including inflammatory, metabolic, and neurodevelopmental disorders and carcinogenesis. The authors analyzed the diverse models (animal, human, epidemiological, and in vitro studies) and documented the effects of both short-term and long-term exposure at various doses, as reported over the last 10 years. I believe this work will be of interest to a broad range of readers due to the widespread use of pesticides in agriculture and their subsequent impact on human health, as well as the environment.

Therefore, I would recommend this work for publication after the minor revisions noted below are properly taken into consideration.

Where in the text is the reference to Fig. 1?

The Latin names of bacterial species should be written in italic, e.g., Page 5, line 166 B. amyloliquefaciens; Page 18, line 546 Roseburia; Page 20, line 647 Helicobacter pylori, etc.

Some abbreviation is missing, for example, CRC (colorectal cancer) Page 20,

 Page 5, line 178: What is the meaning of HWG?

Line 114: Please specify what kind of plant is more accurate in the phrase: This inhibits protein synthesis and secondary metabolites, causing the plant to die within 1 to 3 weeks. Herbicides are applied to weeds, not to all plants.

I cannot agree with the statement: Page 11, line 428: However, one study also reported a rise in Helicobacter levels. At the end are placed two references (Ku 428 et al., 2023; Liu et al., 2021).

Page 10, line 400: explain/add the active compound in Roundup- not all Readers are familiar with commercially available products containing pesticides.

Correct the references:

Page 19, line 599: The study by Zhang et al. (2019) on adult male zebrafish exposed to 100 and 1000 μg/L of 600 propamocarb for 7 days revealed significant disruptions in glucose and lipid metabolism. Should be [151]. Zhang et al. (2018) https://doi.org/10.1093/abbs/gmy153

Page 22, line 794: Which publication of Munoz should be cited? [96] or [97]?

Page 24, line 857: Where in the references part is the cited publication reported by Wu et al. (2023)?

Finally, to improve the manuscript, I recommend a minor revision to address some typographical and other errors present in the manuscript:

The applied units: ug should be corrected to μg; superscripts should be used for example 1x10-3 M: Page 22, lines 751, 755, 758;

Page 23/24: Correct the name of the author: Othmene et al. or Othmène et al.,

Page 5, line 160. correct the name of Propamacarb;

no brackets (line 227)

Please ensure this correction is applied throughout the manuscript.

Author Response

We sincerely appreciate the reviewer's thoughtful questions and feedback, which have helped us improve the quality of our review.

1. Where in the text is the reference to Fig. 1?

Line 334

2. The Latin names of bacterial species should be written in italic, e.g., Page 5, line 166 B. amyloliquefaciens; Page 18, line 546 Roseburia; Page 20, line 647 Helicobacter pylori, etc.

Noted and amended

3. Some abbreviation is missing, for example, CRC (colorectal cancer) Page 20

CRC is first mentioned in full on page 4 line 646

4. Page 5, line 178: What is the meaning of HWG?

HWG is a code assigned to tebuconazole-related metabolites to help track and identify its degradation products during environmental and toxicological studies. (https://sitem.herts.ac.uk/aeru/ppdb/en/Reports/610.htm) We removed it from the text to avoid confusion and kept the chemical name.

5. Line 114: Please specify what kind of plant is more accurate in the phrase: This inhibits protein synthesis and secondary metabolites, causing the plant to die within 1 to 3 weeks. Herbicides are applied to weeds, not to all plants.

We agree that herbicides are applied to unwanted plants (weeds), not to all plants. Our intention was to refer to the targeted plant species affected by the herbicide. We have revised the sentence for clarity as follows:

“This inhibits protein synthesis and the production of secondary metabolites, causing the targeted plant to die within 1 to 3 weeks.”

6. I cannot agree with the statement: Page 11, line 428: However, one study also reported a rise in Helicobacter levels. At the end are placed two references (Ku 428 et al., 2023; Liu et al., 2021).

This was taken from Ku et al., 2023, as also noted in Table 3. We removed the citation for Liu et al., 2021.

7. Page 10, line 400: explain/add the active compound in Roundup- not all Readers are familiar with commercially available products containing pesticides.

We added in parentheses that Roundup is “ a formulation of glyphosate consisting of the potassium salt of glyphosate together with surfactants”)

8. References:

Line 601 – Zhang et al., 2018

Lines 787 and 796 - Woźniak et al., 2021

Line 750 - Muñoz et al. (2023a)

Ref 135 amended

Line 858 – Ku et al., 2023

9. Typographical errors fixed

 

 

 

 

Author Response File: Author Response.docx

Reviewer 3 Report

Comments and Suggestions for Authors

• Consider deleting Table 1, as the information it contains is already adequately covered in the introduction.

• For clearer visualization, it is recommended to add a figure illustrating the degradation pathways of pesticides.

• Please also include a brief section discussing the toxicity of pesticide degradation products.

• Many sections of the review mainly summarize previously reported research (e.g., on inflammation, metabolic disorders, cancer, etc.). It is recommended to include statements highlighting the key findings and major insights from the literature.

• Additionally, elaborate on how the pathways affected by these pesticides contribute to inflammation, metabolic disorders, and cancer.

Author Response

We sincerely appreciate the reviewer’s thoughtful questions and feedback, which have helped us improve the quality of our review.

1. Consider deleting Table 1, as the information it contains is already adequately covered in the introduction.

We have retained Table 1  because it provides a concise and clear overview of the chemical structure and chemical name of each pesticide. However, to avoid redundancy in the introduction, we have deleted lines 87–91

2. For clearer visualization, it is recommended to add a figure illustrating the degradation pathways of pesticides.

In this review, the section on transformation and degradation is intended primarily to provide background context for later sections, particularly in relation to biomonitoring and health effects studies where certain metabolites or degradation products are discussed. As this is not a central focus of the review, we believe that including a detailed degradation pathway figure may not add substantial value for the reader. 

3. Please also include a brief section discussing the toxicity of pesticide degradation products.

Where relevant studies were available; such as those on AMPA (e.g. lines 616, 710, 736 etc.), we have included this information in the text. However, as indicated in the “Transformation and degradation of pesticides” section, the parent compound generally represents the main residue of concern, and the majority of available studies focus on the toxicity of the parent pesticide. For this reason, the discussion in our review is centred primarily on the parent compound, while including degradation product toxicity where data permit. 

4. Many sections of the review mainly summarize previously reported research (e.g., on inflammation, metabolic disorders, cancer, etc.). It is recommended to include statements highlighting the key findings and major insights from the literature.

Section 5 presents recent documented effects of glyphosate, deltamethrin, propamocarb, and tebuconazole on gut microbiota, gut inflammation, metabolism, cancer, and the gut–brain axis. This section was presented with limited author interpretation to allow readers to assess the findings objectively and draw their own conclusions.  Our interpretation, along with key insights, limitations, and considerations for future research, is provided later in the review (Sections 6 and 7).

To directly address the reviewer’s suggestion, we have strengthened Section 7 by adding an integrated synthesis (lines 937–954) that highlights the key findings and major insights from the literature covered in Sections 5 and 6. This new paragraph draws connections across disease areas, and summarises proposed mechanisms of action. The added text reads:

“Overall, the evidence reviewed highlights several key trends. These pesticides can induce shifts in gut microbial composition, often reducing beneficial taxa and altering SCFA production, with effects varying by compound, dose, sex, and study model. Chronic exposure in animal and in vitro models can trigger gut and systemic inflammation, often through disruption of the intestinal barrier and upregulation of pro-inflammatory pathways, with potential to exacerbate pre-existing inflammatory conditions. Pesticide exposure can also disrupt lipid and glucose metabolism, alter bile acid homeostasis, and promote metabolic disorders such as obesity, type 2 diabetes, and atherosclerosis, frequently through gut microbiota–mediated mechanisms. Evidence further suggests that glyphosate, propamocarb, tebuconazole, and deltamethrin may contribute to cancer risk via gut dysbiosis, modulation of oncogenic signalling, chronic inflammation, metabolic disruption, oxidative stress, endocrine modulation, and epigenetic alterations, though the strength of evidence varies across compounds and exposure contexts. Additionally, pesticide-induced gut microbiota alterations may influence neurological health by disrupting neurotransmitter metabolism, increasing neuroinflammation, and impairing synaptic function, thereby contributing to behavioural changes and cognitive deficits; however, these effects appear to vary by pesticide type, exposure timing, and host susceptibility.”

5. Additionally, elaborate on how the pathways affected by these pesticides contribute to inflammation, metabolic disorders, and cancer.

The aim of this review is to examine how pesticide exposure can influence disease, with an emphasis on the biological pathways and markers through which these effects occur, rather than providing exhaustive mechanistic detail. The review already discusses relevant pathways and markers, including pro-inflammatory cytokine release, oxidative stress, oncogenic signalling, endocrine modulation, and epigenetic alterations, that are implicated in disease pathogenesis. However, we acknowledge that in these sections, the links between these pathways and specific disease outcomes could be made more clear. To address this, we have added clarifying statements in the relevant subsections of Section 5 to connect each pathway to its role in inflammation, metabolic dysregulation, or cancer progression. For example:

line 488 “Mechanistically, deltamethrin activated the p53 signalling pathway in colonic epithe-lial cells, driving cellular senescence. Senescent cells adopted a senescence-associated secretory phenotype (SASP), characterised by the persistent release of pro-inflammatory cytokines, chemokines, metalloproteinases, and reactive oxygen species, thereby sustaining and amplifying colonic inflammation (Ma et al., 2024).”

Line 593: Meng et al. (2022) found that exposure to tebuconazole in the mice caused significant changes in three metabolic pathways involved in glycolipid metabolism, amino acid metabolism and tricarboxylic acid (TCA) cycle indicating disruption of core energy and nutrient metabolic pathways.

Line 670: These cytokines contribute to the growth of neoplasia by activating signalling pathways such as JAK/STAT3 that promote tumour cell proliferation and survival (Ullman & Itzkowitz, 2011).

 

 

 

 

 

 

Author Response File: Author Response.docx

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

I have no additional comments or remarks

Reviewer 3 Report

Comments and Suggestions for Authors

The authors have appropriately revised the manuscript; therefore, I recommend it for acceptance in its present form.