PONE-D-22-16144

The accuracy of PCR-based methods in predicting human intestinal nematode infections: a protocol for a systematic review and meta-analysis of the accuracy of available diagnostic tests

PLOS ONE

Dear Dr. Weerakoon,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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Please find the comments provided by the reviewers below this email.

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Please submit your revised manuscript by Sep 26 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

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We look forward to receiving your revised manuscript.

Kind regards,

Hesham

Hesham M. Al-Mekhlafi, PhD

Academic Editor

PLOS ONE

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2. Thank you for stating the following financial disclosure: 

 "This review is a part of an intestinal parasitoses community evaluation project funded by the National Research Council of Sri Lanka (NRC 20-118). However, this particular publication (review protocol) is not funded through this grant. The funder had and will not have a role in study design, data collection and analysis, decision to publish, or preparation of the manuscript."

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Does the manuscript provide a valid rationale for the proposed study, with clearly identified and justified research questions?

The research question outlined is expected to address a valid academic problem or topic and contribute to the base of knowledge in the field.

Reviewer #1: Partly

Reviewer #2: Yes

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2. Is the protocol technically sound and planned in a manner that will lead to a meaningful outcome and allow testing the stated hypotheses?

The manuscript should describe the methods in sufficient detail to prevent undisclosed flexibility in the experimental procedure or analysis pipeline, including sufficient outcome-neutral conditions (e.g. necessary controls, absence of floor or ceiling effects) to test the proposed hypotheses and a statistical power analysis where applicable. As there may be aspects of the methodology and analysis which can only be refined once the work is undertaken, authors should outline potential assumptions and explicitly describe what aspects of the proposed analyses, if any, are exploratory.

Reviewer #1: Partly

Reviewer #2: Yes

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3. Is the methodology feasible and described in sufficient detail to allow the work to be replicable?

Descriptions of methods and materials in the protocol should be reported in sufficient detail for another researcher to reproduce all experiments and analyses. The protocol should describe the appropriate controls, sample size calculations, and replication needed to ensure that the data are robust and reproducible.

Reviewer #1: No

Reviewer #2: Yes

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4. Have the authors described where all data underlying the findings will be made available when the study is complete?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception, at the time of publication. The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

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6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above and, if applicable, provide comments about issues authors must address before this protocol can be accepted for publication. You may also include additional comments for the author, including concerns about research or publication ethics.

You may also provide optional suggestions and comments to authors that they might find helpful in planning their study.

(Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: This manuscript describes a protocol to perform a systematic review, analysing the diagnostic accuracy of PCR-based methods for the detection of human intestinal nematodes. Taking into account the increasing use of nucleic acid amplification tests (NAAT’s) in the diagnosis of intestinal helminths, I think the authors have chosen a timely topic for their review. However, I see several flaws and inconsistencies in their approach.

First of all, what I understand from the methods section is that the Kato-Katz smear will be used as the reference test. Therefore, clarity should be given about the way the Kato-Katz has been performed in each study, i.e. how many smears have been examined, based on how many stool samples and were there appropriate quality control measures in place. Based on the described performance of the Kato-Katz smear examination the authors could rank each individual study included and give them a different weight in the overall analysis.

Furthermore, for Strongyloides and Enterobius, it is not clear to me how the authors want to compare the PCR outcome with Kato-Katz data. For these two species other microscopy procedures (Baermann, tape-test) are more appropriate.

In addition the authors should be more specific about the NAAT’s included. Even if limiting to a real-time PCR format, each test is different, be it either in design and target selection, as well as in procedures of sample preparation and DNA extraction, the output variable producing, and most importantly, the level of appropriate quality control measures implemented. How to interpret these differences between publications is not clarified in this protocol. The description of Line 187-188 I find therefore far from complete.

This lack of uniformity is not only a major struggle for the analysis of PCR performance, but also for serology. This brings me to another point. In the abstract I read: “We plan to analyse the efficacy of microscopic- and PCR-based approaches in detecting human intestinal nematode infections in different health care settings.” In the key-words I noticed “serology”, but nothing is mentioned in the abstract. Only from the search strategy (line 225) and description at line 181-186 I understand that several immune-diagnostic tests will be included as index test, including antibody tests and antigen tests. This sounds rather ambitious to me. The authors should be more clear what the relevance is of including immunodiagnostic test and how they want to summarise the different tests available. My advise is to leave the immunodiagnostic tests out of the analysis.

Also for the different microscopy-based procedures included, the descriptions are not very clear to me. In my opinion the protocol would improve if the focus would be on a comparison between real-time PCR as the index test and the most optimal microscopy procedure as a reference test. Concerning microscopy, this is Kato-Katz for most STH species and species-dedicated procedures for Strongyloides and Enterobius

Minor comments:

Abstract: the proposed reference test is not described (as it is in line 138-140).

Line 62: other Ancylostoma species should also be included .

Line 226: ELIZA should be ELISA.

Line 226: “immunofluorescent antibody” is not an appropriate search term.

Line 278: “The gold standard: As there is no universally accepted gold standard, we will use those in the published literature and the WHO recommended gold standard, which is Kato Katz.” I suggest to rephrase this as follows: “Reference test: As there is no universally accepted “gold” standard, we will use the Kato Katz as the reference test for the most common soil transmitted helminths, as this test is generally used in published literature and also recommended by the WHO as the most appropriate reference for these species.”

Reviewer #2: Title: The accuracy of PCR-based methods in predicting human intestinal nematode infections: a protocol for a systematic review and meta-analysis of the accuracy of available diagnostic tests.

Comments

This is a well thought out protocol, focusing on diagnostic techniques. It is well written with clear and concise methods which are adequate to realise the study goals. However, I wish to raise a few issues that the authors should consider before publication of the protocol.

1. From the study objective, and the narrative in the whole protocol, it is clear that the goal of the systematic review is to evaluate different diagnostic techniques/ approaches that are available for evaluation of human intestinal nematodes. However, the title of the study is more biased to PCR-based approaches which are not the primary focus of the study. I will suggest that the title is reviewed to capture the whole study and not part of the study.

2. In the objective (Abstract section), the authors states that the study will specifically focus on microscopic and PCR based approaches, however in the Eligibility criteria (index tests Page 7 and 8), they propose to include serology based assays. This is a key area in diagnosis of human nematodes and needs to come out well in the objectives and also in the title.

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Reviewer #1: No

Reviewer #2: No

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Attachments

Attachment

Submitted filename: PLOS ONE. Reviewers comments.docx

PONE-D-22-16144R1The accuracy of nucleic acid amplification tests (NAATs) in detecting human intestinal nematode infections: a protocol for a systematic review and meta-analysisPLOS ONE

Dear Dr. Weerakoon,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

==============================Please find few minor comments by the reviewers at the end of this email. Moreover, please prepare the list of references according to the journal's instruction and style.==============================

Please submit your revised manuscript by Nov 26 2022 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

• A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Hesham

Hesham M. Al-Mekhlafi, PhD

Academic Editor

PLOS ONE

Journal Requirements:

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Does the manuscript provide a valid rationale for the proposed study, with clearly identified and justified research questions?

The research question outlined is expected to address a valid academic problem or topic and contribute to the base of knowledge in the field.

Reviewer #1: Yes

********** 

2. Is the protocol technically sound and planned in a manner that will lead to a meaningful outcome and allow testing the stated hypotheses?

The manuscript should describe the methods in sufficient detail to prevent undisclosed flexibility in the experimental procedure or analysis pipeline, including sufficient outcome-neutral conditions (e.g. necessary controls, absence of floor or ceiling effects) to test the proposed hypotheses and a statistical power analysis where applicable. As there may be aspects of the methodology and analysis which can only be refined once the work is undertaken, authors should outline potential assumptions and explicitly describe what aspects of the proposed analyses, if any, are exploratory.

Reviewer #1: Yes

********** 

3. Is the methodology feasible and described in sufficient detail to allow the work to be replicable?

Descriptions of methods and materials in the protocol should be reported in sufficient detail for another researcher to reproduce all experiments and analyses. The protocol should describe the appropriate controls, sample size calculations, and replication needed to ensure that the data are robust and reproducible.

Reviewer #1: Yes

********** 

4. Have the authors described where all data underlying the findings will be made available when the study is complete?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception, at the time of publication. The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

********** 

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

********** 

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above and, if applicable, provide comments about issues authors must address before this protocol can be accepted for publication. You may also include additional comments for the author, including concerns about research or publication ethics.

You may also provide optional suggestions and comments to authors that they might find helpful in planning their study.

(Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: I read the rebuttal and reviewed the changed made in the manuscript. In my opinion the authors responded well to all points mentioned by the reviewers. In particular I appreciate the more focused approach of the protocol.

Still, I have a number of technical comments. Most of these comments relate to the NAAT-procedures. The authors could be more precise in their wording and definitions at several points throughout the manuscript.

The line numbers mentioned refer to the manuscript including track changes.

Abstract (line 40): “… to more specialised high sequencing technologies such as the NAAT.” It is not clear to me what the authors try to say here. NAAT is a very broad definition of all tests related to nucleic acid amplification. Consequently “the” (?) NAAT is not an example of sequencing. More the other way around. Furthermore, the abbreviation NAAT is explained in the title. Still this should be repeated in the abstract (and in the introduction, see below).

Abstract (line 53): indeed, the details of the data bases are not so relevant for an abstract.

Objectives (line 151-152): I agree that the objectives improved by using NAATs and not “PCR-based methods”. But at the introduction section the terminology of NAATs have not been well introduced. In line 121-122 PCR is introduced as a method, but nothing is mentioned about other NAAT-methods, including isothermal amplification procedures, such as LAMP.

Inclusion criteria (line 187-188): LAMP is one example of several isothermal amplification procedures. It is not clear to me why only LAMP is mentioned an no other methods.

Search strategy (line 237-238): again it is not clear to me why only “loop mediated isothermal amplification” OR “LAMP” is included in the search terms for PubMed. I suggest to include the term “isothermal amplification” as well.

Data extraction (line 326-327): I appreciate the inclusion of a weight-factor, based on the quality of the used diagnostic procedures. But the description how this is done is completely lacking.

Data extraction (line 346-347): ”…the number of smears taken”. Please explain. I have never heard of “smears” in the context of a Baermann procedure. In addition ‘the standard technique” is not existing for the Baermann. Rather different approaches have been described.

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7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

**********

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

The accuracy of nucleic acid amplification tests (NAATs) in detecting human intestinal nematode infections: a protocol for a systematic review and meta-analysis

PONE-D-22-16144R2

Dear Dr. Weerakoon,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

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Kind regards,

Hesham

Hesham M. Al-Mekhlafi, PhD

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

This version has addressed all comments highlighted by the reviewers.

I would like to take this opportunity to express my deepest sympathy and condolences on the loss of Prof. Donald McManus of QIMR Berghofer Research Institute, Australia, to his family, friends, students, colleagues and the science community. Prof. Donald McManus was an esteemed parasitologist who dedicated his life to the prevention and elimination of neglected tropical diseases, and has made incredible contributions to the field of parasitology. Rest in paece!

Hesham Al-Mekhlafi

Reviewers' comments: