PONE-D-24-10270Minding the margins: Evaluating the impact of COVID-19 among Latinx and Black communities with optimal qualitative serological assessment tools.PLOS ONE

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Reviewer #2: Yes

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Reviewer #2: Yes

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Reviewer #2: Yes

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Reviewer #2: Yes

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5. Review Comments to the Author

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Reviewer #1: Reviewer comments:

The authors have made a detailed qualitative serological assessment to investigate the impact of COVID-19 among the Latinx and Black communities. After going through the manuscript, I have found that the parameters for participant recruitment, sample collection, and sample processing had been performed meticulously (as the study involved culturally sensitive-marginalized communities) and were lucidly justified in the manuscript. The following of concerned protocols regarding the handling of human samples and due approval of the concerned authority/ethical committee is appreciated. The survey was performed to answer mainly three important investigations, i.e. (i) the impact of COVID-19 and vaccine uptake among marginalized communities, (ii) the utility of using saliva for serosurveys, (iii) a comparison of the utility of a bead-based multiplex assay vs. a point-of-care (POC) test for SARS-CoV-2 antibody measurements, and (iv) demonstration of the benefit of developing and using classification boundary methods for optimal interpretation of serological assays. The main research techniques used in the study were: Multiplex Luminex assay, POC test, across-plate normalization, and Qualitative assessment using statistical analysis.

The following few points need to be highlighted by the authors to aid the readers in perceiving the statements made thereof.

Methods (Qualitative serological assessment)

The authors have reflected on the use of alternative control samples from a Kenyan study for qualitative serological assessment which is elaborated in the supplementary information. However, I feel the need to cite the reference of that particular Kenyan study in the methods section itself for the ease of the readers.

Results

Demographics and Vaccine/Infection History: The authors have elaboratively discussed the population demographics and the history of COVID-19 vaccinations in the Latinx and Black communities. As the study is highly based on the self-declaration of health status by individuals of these communities with a very limited sample size, I feel the need to mention/compare the demographics with previous studies of COVID-19 on the Latinx and Black communities (as some more references are available).

Blood-based SARS-CoV-2 Antibodies test:

The POC test covered both SARS-CoV-2 N and S antigens, while the multiplex assay allowed measuring the presence of antibodies based on individual antigens and therefore distinguishing between vaccine and infection-induced antibodies. The POC test was found reliable for detecting the RBD/S antibodies measured by the multiplex assay. This section needs no further changes.

Discussion:

As mentioned by the authors, the antigen-specific outcomes between serum and saliva did not correlate, even though the antigen-specific IgG seroprevalences aligned. However, the authors have been able to ascertain the use of saliva as a less-invasive and accessible sample. Furthermore, this research highlights the need of hCoV cross-reactivity to be evaluated for reliable SARS-CoV-2 serosurvey results.

Final comment: This research investigation is of potential importance as it makes important remarks on the impact of the COVID-19 disease among marginalized ethnic races using serological assessment tools and also devices some qualitative assessment parameters that will serve as effective investigation methods for further large-scale population-based surveys. The authors were able to answer the questions addressed initially in the investigation. No significant grammatical/English corrections were noticed in the manuscript. The manuscript is suitable for publication with very minor changes as mentioned above.

Reviewer #2: The authors show the feasibility of using saliva and matching blood samples to determine SARS-CoV-2 antibody seroprevalence among a predominantly female and Hispanic population. They found good correlation between self-reported vaccination status (most were vaccinated against COVID) and a POC SARS-CoV-2 antibody device they employed on-site during the study visit and a multiplex assay that was used to test both blood and saliva for SARS-CoV-2-specific and endemic coronavirus IgG and IgA with the exception for salivary IgA.

My main comment is that the authors should address the discrepancy they see between prior self-reported COVID-19 infection and discrepant classification by blood and maybe also saliva test. ~40% of participants reported having had COVID-19 (test confirmed, n = 121 participants) but about 50 of them did not test positive for anti-N IgG. Similarly, ~60 out of ~160 participants who reported not having had COVID-19 do test positive for anti-N IgG. It is a bit odd to stratify Table 1 by this outcome and then not address it.

Another comment is that while normalization of saliva antibody signals with total Ig CONCENTRATION is often used, normalization with MFI values only works if both, the pathogen-specific signal and the total Ig MFI signal are within the linear range of the assay. The non-transformed (raw) data is not provided.

Some minor comments:

Line 235 “[…] indicating past infection rather than vaccination and mirrored self- reported exposures.”

The percentage of self-reported infection may be similar to the percentage of anti-N positive blood samples but Table 1 appears to suggest that only ~50% of participants who reported a prior SARS-CoV-2 positive test also tested anti-N positive. Conversely, ~ one third of those who did not report a prior SARS-CoV-2 positive test had a anti-N positive result. The authors may want to acknowledge this here or omit the “mirroring” statement if addressed in the discussion.

Line 242 As for SARS-CoV-2 variants, the delta variant had the most abundant antibodies among our study population, see Fig 1.

Higher MFI does not necessarily mean most abundant antibody. MFI signals are influenced by a lot of factors including quality of the antigen, orientation of immunogenic (antibody-binding sites) regions of the antigen on the bead, antigen density on the bead, orientation of the bound antibody on the bead, etc. The study took place during the Omicron wave and, as the authors mentioned, most participants were vaccinated (i.e., “primed” with the Wuhan strain). Consider acknowledging that MFI signal strength alone does not mean that most participants were infected with Delta or revise sentence accordingly.

Line 263 “[…] indicating that the POC test was reliably detecting the RBD/S antibodies measured by the multiplex assay.”

Are the authors trying to say that the POC assay does only classify blood samples with high/higher anti-S MFI as positive? Please clarify. Also, a brief description about what is meant with 2/3 antigen positivity (S/N; RBD/N, etc.) would be helpful. I.e., positive for IgG to all antigens or at least one of them, etc.? It might also be helpful to indicate the multiplex assay outcome (for one of the algorithms?) in Figure 2, e.g., coloring the dots according to the multiplex assay result or by using different (larger) symbols according to result and/or adding a threshold for S MFI / RBD MFI / N MFI.

Line 266 “negative for the POC test across all antigen combinations”

Does the POC test detect antibodies against multiple antigens?

Line 300 While anti-SARS-CoV-2 IgA has been shown to resolve faster than IgG

Maybe “wane” or “decline” would be the better word?

Line 350 “the first COVID-19 case in MA was confirmed on Feb 1st 2022”

Should this be 2020?

Line 363 groups as a risk factor, although and comorbidities did not correlate with lack of COVID-9 symptom resolution,

Grammar? Delete “and” I assume.

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Reviewer #2: No

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PONE-D-24-10270R1Minding the margins: Evaluating the impact of COVID-19 among Latinx and Black communities with optimal qualitative serological assessment tools.PLOS ONE

Dear Dr. Binder,

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 address the following minor issues: Re-number supplemental tables and figures. In the current version, Table S5 is the first supplemental table referenced in the Supplemental methods while Figure S11 is the first supplemental figure referenced in the text on page 12.

Please check the names of companies and products. It should be Sino Biological rather than Sino Biology in Table S5.  

Please make sure to use consistent and logical terminology and definitions throughout the text. For example, please correct statements in Lines 98-101 and in 2^nd paragraph in the Supplemental Methods. The POC test detects IgG and IgM antibodies to S and N antigens (anti-SARS-CoV-2 antibodies), rather than anti-immunoglobulin antibodies in blood samples. Also, this test does not detect S and N antigens in blood.

Lines 358 – 361. It appears that previously infected (“exposed”) individuals include those who tested seropositive to the N antigen as well as those seronegative individuals who had been diagnosed with SARS-COV-2 infection (Table 1).  

Lines 395 and 450. No data to compare sensitivity values of Luminex and POC tests is provided in the paper. Please provide such data or remove statements about Luminex assay being more sensitive.

Supplemental Information, Multiplex Assay – Saliva: Please confirm that undiluted saliva samples were tested for total IgG and IgA. The concentration of total Ig in saliva is so high that it would be reasonable to use highly diluted saliva in this analysis.

Please submit your revised manuscript by Aug 18 2024 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,

Andrey I Egorov

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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Minding the margins: Evaluating the impact of COVID-19 among Latinx and Black communities with optimal qualitative serological assessment tools.

PONE-D-24-10270R2

Dear Dr. Binder,

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Andrey I Egorov

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