The 863C>A and 1031T>C Single Nucleotide Polymorphisms (SNPs) in the Tumor Necrosis Factor Alpha (TNF-α) Promoter Gene May Not Be Putative Predictors of HBV Endemicity

Round 1

Reviewer 1 Report

Dear authors,

I extend my heartfelt gratitude to you for submitting your work to Livers (MDPI), and I am honored to have been entrusted with reviewing your manuscript. 

I assure you that I approached the evaluation of your manuscript with utmost impartiality, conducting a thorough and critical analysis. 

The manuscript exhibits exceptionally high-quality content. Nonetheless, it is important to address a few limitations beforehand.

Please, ensure that all the suggested modifications have been diligently incorporated. 

I kindly request that you consider having your manuscript reviewed by native English speakers, as there appear to be moderate English editing issues.

Best regards,

The Reviewer

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ABSTRACT

Comment 1: Omit the pre-print information from the abstract, refraining from indicating its prior publication as a pre-print, and remove the corresponding reference within the abstract. If you intend to include this information, please make sure to place it towards the conclusion of the introduction section.

Comment 2: Omit the web address from line 31. While this aspect couldn't be discussed in the abstract, it should be addressed in the methodology section.

INTRODUCTION

Comment 1: In the transition between the first and second paragraphs, kindly introduce a new passage illustrating instances where HBV infection dynamics were influenced by genetic polymorphisms within populations of different nations. Please incorporate concise examples, along with appropriate citations of the authors and their respective references. Cite the countries and the study types.

Comment 2: (Line 65) Reactive oxygen species (ROS) and free radicals should be rephrased. ROS cannot be indiscriminately equated with free radicals for all purposes. Please insert an "and" between ROS and free radicals.

MATERIALS AND METHODS

Comment 1: Please carefully observe the titles of the subsections and eliminate the commas after their phrases (as exemplified in subsection title 2.1).

Comment 2: The content spanning lines 124 to 136 may be more suitable for the Results section rather than the Methodology. This portion provides details about the patients screened and included in the study. Therefore, I recommend rephrasing these sentences and relocating them to the Results section.

Comment 3 (Section 2.2): Mention any statistical considerations that influenced the decision for why you chose to arrive at a sample size of 384.

Comment 4 (Section 2.3, Line 151): You haven't explained how the sample collection methods were carried out; I could not locate this information. Instead, you've referenced the source. Could you kindly elaborate on the sample collection methods in detail within this manuscript?

Comment 5 (Section 2.4, Lines 173-174): You've mentioned that your study utilized the 20,000 IU/mL cutoff, in line with the Ugandan Ministry of Health's criteria for treatment decisions. It would be valuable to briefly highlight the alignment of this threshold with those from prominent international organizations. Could you please provide citations from sources such as WHO, World Hepatitis Alliance, Hepatitis B Foundation, and Health-EU portal, among others? This will enhance the generalizability of your study for endemic areas.

Comment 6 (Section 2.9): In section 2.9, it's important to provide a more detailed description of the statistical analyses conducted. Ensure that you explicitly mention all the statistical tests employed and elaborate on how each of these tests contributes to substantiating your points. Including references supporting the appropriateness and adequacy of these chosen statistical tests would be beneficial.

RESULTS

In this section, incorporate relevant information (such as the names and relevance) about the statistical tests employed for analyzing each result. Integrating this information will elevate the overall quality of your presentation.

CONCLUSION

Kindly integrate a conclusive segment into this manuscript that skillfully summarizes the addressed issues, acknowledges relevant constraints within the scope of the investigation, and sheds light on future directions. Emphasize the most noteworthy findings and their implications.

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Thank you, esteemed authors!

The manuscript requires some moderate English editing. I suggest that the authors either have native speakers review the language used in their manuscript or consider utilizing a paid service such as MDPI or PaperTrue.

Author Response

The changes are highlighted in the manuscript and the step-by-step response to the reviewer's comment has been given at the end of the revised manuscript 

Author Response File: Author Response.docx

Reviewer 2 Report

The TNF-α has been implicated in the pathogenesis of inflammatory, malignancies and auto immune diseases. It is an example of the inflammatory cytokine and is important in the prognosis of the liver diseases following infection with hepatitis B virus. This paper presented the study guide future research in establishing a plausible explanation for the differences in endemicity of the virus in Uganda.  The results inform health care providers when designing interventions that can bridge the HBV endemicity gap in this country. This will provide a corner stone in the journey towards the elimination of the virus by 2030 as highlighted by the United Nations sustainable development goals. This is a very interesting, useful and up-to-date article.

The literature cited in this paper is sufficient in volume and content (58). All 5 figures are of sufficiently high quality. The 4 tables are well presented. I recommend that the journal editors accept the article with reg. no.2581898 in  this form.

I have no questions, remarks or recommendations for the authors.

Author Response

The reviewer endorsed the publication of the manuscript in submitted state. So the reviewer never gave any comments 

Reviewer 3 Report

In the current manuscript, the authors discuss whether or not the prediction of HBV endemicity could be made by 863C>A and 1031T>C single nucleotide polymorphisms in the TNF-α promoter or not in Negros' patients in Uganda. The manuscript is among a list of studies for the same research group. The study is good, and a few comments were addressed, including the following:

- No reference in the abstract; the authors may transform the last sentence in the abstract into the introduction section.

- In the introduction section:

- Line number 57 "It is an example of the inflammatory cytokine and is…" It is better to replace "It" with "TNF-α" to be clear to the reader.

- In line number 90 "TNF-α − 863 C/A SNPs" the full name must be written first, then the abbreviation is used.

- In the methods and materials section:

- In lines number 220-221 "incubated for 15minutes at 37oC and the at 80oC for a minute to terminate the reaction" Please revise.

- In Figure 3, "Lanes 1-4, 5, 12-14, and 18 are negative samples for the HBV pol gene." Please revise the lane numbers.

- In the results section:

- In line number 341, CHB is written without its full name.

- In the discussion section, the second paragraph must start with "in our study, or our data revealed, etc. " to clarify that it is your study.

- The authors claimed, "Second, the differences in the environment between our study and the published studies since the environment is a key driver of mutations and evolutionary changes." It is better to explain the differences in environment, such as whether the pollution in the study region is greater or lesser than in other study areas, the habits, the diet, etc….

- "Finally, in our systematic review on the prevalence and predictors of HBV in East Africa, our data synthesis has shown that the most important risk factor to infection has been body scarification (50); a vice that is common among the persons from the high endemic region compared to those fem the low endemic regions(51)" Please revise the risk factor to infection in the other study (reference 50) and the spelling.

- The written limitation of the study is critical in explaining the drawbacks of the data, but as the authors listed the levels of TNF-α is so important and the study results could be tentative for further studies.

- The references are recent and sufficient.

Author Response

The changes in the manuscript have been highlighted and a step-by-step response has been given at the end of the manuscript. 

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

The manuscript has been significantly improved based on my suggestions.

I recommend publication for the manuscript, which has been significantly improved based on my suggestions.