Prevalence of Pathogenic Microbes within the Endometrium in Normal Weight vs. Obese Women with Infertility

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The study conducted by King et al. aimed to determine the diferences in endometrial microbiome across body weight.

• Throughout the introduction, many more studies (references) are needed to support the information.
• Discuss chronic endometritis.
• The names of the microorganisms must be in italics.
• Introduce the clinical tests for microbiome detection used in the study. Include their limitations.
• Include the ethics committee approval number.
• Abbreviations must be defined the first time they are used.
• Analyze differences in terms of age among the different study groups.
• Figure 1 can be converted into percentages for clearer understanding.
• Redefine the groups based on BMI (not based on the uterine microenvironment).
• Given that there are different groups of pathology or etiology of infertility, the effect of BMI cannot be clearly observed.

Comments on the Quality of English Language

It is necessary to review the grammar and spelling.

Author Response

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections in track changes in the re-submitted files. 

Throughout the introduction, many more studies (references) are needed to support the information.

1. I changed the introduction to begin with a more general statement about obesity among women of reproductive age instead of directly addressing the prevalence of obesity in the United States because I wanted to provide a broader context before delving into specific details.
2. I added more information about the ongoing exploration of the relationship between obesity and reproductive function because I wanted to emphasize the dynamic nature of this research area.
3. I included a section highlighting recent studies on the influence of the endometrial microbiota on reproductive function because I wanted to draw attention to this important aspect that was not explicitly mentioned in the first version.
4. I listed specific pathogenic bacteria associated with impaired reproductive function to provide more clarity and detail on this aspect, which was not addressed in the first version.
5. I added information about potential mechanisms linking obesity to impaired reproductive function and its association with other reproductive pathologies to provide a more comprehensive overview of the topic.
6. I referenced existing studies exploring the relationship between obesity and the endometrial microbiome in post-menopausal women with endometrial cancer to support the discussion and provide context for the research gap addressed in the current study.
7. I explicitly stated the lack of publications focusing on the relationship between obesity and the endometrial microbiome in women of reproductive age without endometrial cancer to highlight the specific research gap that the current study aims to address.

• See lines 24-70

 

Discuss chronic endometritis.

• See lines 50-55
• Added section about chronic endometritis to tie together the introduction and the ALICE study specific results

 

The names of the microorganisms must be in italics.

• All microorganisms are in italics.
• Changed “lactobacillus” to “lactobacillus” throughout text

 

Introduce the clinical tests for microbiome detection used in the study. Include their limitations.

• See lines 83-100
• Limitations discussed in detail in “detailed EMMA/ALICE protocol” – line 96

 

Include the ethics committee approval number.

• It is included(IRB number) – line 82, 199

 

Abbreviations must be defined the first time they are used.

• in vitro fertilization now defined (IVF) – line 74
• all abbreviations are identified

 

Analyze differences in terms of age among the different study groups.

• See table 1
• age is first variable assessed and is not different among groups 

 

Figure 1 can be converted into percentages for clearer understanding.

• Changed figure 1 to highlight main point of article and used it to replace table 3
• Converted to percentages

 

Redefine the groups based on BMI (not based on the uterine microenvironment).

• Redefined and updated – see table 1

 

Given that there are different groups of pathology or etiology of infertility, the effect of BMI cannot be clearly observed.

• Added to discussion section
• see lines 182-184

 

 

 

 

 

 

 

Reviewer 2 Report

Comments and Suggestions for Authors

This article examines how BMI relates to the makeup of the endometrial microbiota in infertile women. Overall, the study makes sense. however, there are some shortages in the introduction and discussion sections. Therefore, my comments are as follows.

The title does not convey the aim of the study well.

The introduction appears to be too brief. Several articles have evaluated the endometrial microbiota composition and fertility outcome. I recommend incorporating some of these studies into the introductory section.

Please check the whole article for missing citations (e.g., lines (25-29))

In the Materials and Methods section, please outline your inclusion and exclusion criteria for subjects.

The sample size is small. Please note the limitations of this study.

The discussion section appears too brief. It would be valuable to compare and contrast your results with previous findings.

Author Response

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections in track changes in the re-submitted files. 

 

This article examines how BMI relates to the makeup of the endometrial microbiota in infertile women. Overall, the study makes sense. however, there are some shortages in the introduction and discussion sections. Therefore, my comments are as follows.

The title does not convey the aim of the study well.

• Changed the title
• Prevalence of Pathogenic Microbes within the Endometrium in Normal Weight vs. Obese Women

The introduction appears to be too brief. Several articles have evaluated the endometrial microbiota composition and fertility outcome. I recommend incorporating some of these studies into the introductory section.

- 14, 15, 16, 18, 19 (lines 42-49) are the references I added due to this recommendation. Please see the comments about the other changes I made below 

1. I changed the introduction to begin with a more general statement about obesity among women of reproductive age instead of directly addressing the prevalence of obesity in the United States because I wanted to provide a broader context before delving into specific details.
2. I added more information about the ongoing exploration of the relationship between obesity and reproductive function because I wanted to emphasize the dynamic nature of this research area.
3. I included a section highlighting recent studies on the influence of the endometrial microbiota on reproductive function because I wanted to draw attention to this important aspect that was not explicitly mentioned in the first version.
4. I listed specific pathogenic bacteria associated with impaired reproductive function to provide more clarity and detail on this aspect, which was not addressed in the first version.
5. I added information about potential mechanisms linking obesity to impaired reproductive function and its association with other reproductive pathologies to provide a more comprehensive overview of the topic.
6. I referenced existing studies exploring the relationship between obesity and the endometrial microbiome in post-menopausal women with endometrial cancer to support the discussion and provide context for the research gap addressed in the current study.
7. I explicitly stated the lack of publications focusing on the relationship between obesity and the endometrial microbiome in women of reproductive age without endometrial cancer to highlight the specific research gap that the current study aims to address.

• See lines 24-70

In the Materials and Methods section, please outline your inclusion and exclusion criteria for subjects.

• See lines 73-80
• Outlined inclusion and exclusion more specifically

The sample size is small. Please note the limitations of this study.

• See Lines 179-186
• added comment about the sample size

The discussion section appears too brief. It would be valuable to compare and contrast your results with previous findings.

• I re-worked the entire discussion section, please see lines 149-195

Reviewer 3 Report

Comments and Suggestions for Authors

Dear author. Thank you for a valuable analysis on microbiota in different BMI women. I find several issues that need to be addressed prior to publishing.

1. How were samples obtained? You say frozen-thawed embryo transfers, RPL and failed IUI cycles. This is quite heterogeneous group. I would guess that samples were probably collected as a part of another study. Additionally, it should be stated how they were collected? Biopsy? What kind of biopsy, what equipment was used and what was the procedure? Contamination is a serious issue in microbiota analysis and the method of sample collection is extremely important. Since contamination usually occurs with bacteria from cervix or vagina, several studies collect additional samples from those locations to exclude contamination. 

2. If the study was retrospective, samples should have been collected before as a part of something else.

3. The observed group were patients with fertility issues. This is not a normal or lets say control group. Such group of patients is expected to have altered microbiota which was not taken into consideration. This is serious bias that has to be addressed.

Author Response

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections in track changes in the re-submitted files. 

 

How were samples obtained? You say frozen-thawed embryo transfers, RPL and failed IUI cycles. This is quite heterogeneous group. I would guess that samples were probably collected as a part of another study. Additionally, it should be stated how they were collected? Biopsy? What kind of biopsy, what equipment was used and what was the procedure? Contamination is a serious issue in microbiota analysis and the method of sample collection is extremely important. Since contamination usually occurs with bacteria from cervix or vagina, several studies collect additional samples from those locations to exclude contamination. 

• Samples were obtained through endometrial pipelle with transabdominal ultrasound guidance
• We do not have cervical or vaginal samples to exclude contamination but pipelle did not touch vagina
• Cervical contamination is a limitation of this study
• See lines 83-89
• See lines 184-186

If the study was retrospective, samples should have been collected before as a part of something else.

• clarified in lines 80-82

The observed group were patients with fertility issues. This is not a normal or lets say control group. Such group of patients is expected to have altered microbiota which was not taken into consideration. This is serious bias that has to be addressed.

• addressed in the limitations section + agree with this point
• see lines 182-184