Dear Dr. Hirotsu,

Thank you very much for submitting your manuscript "Household transmission of SARS-CoV-2 R.1 lineage with spike E484K mutation in Japan" for consideration at PLOS Pathogens. As with all papers reviewed by the journal, your manuscript was reviewed by members of the editorial board and by several independent reviewers. The reviewers appreciated the attention to an important topic. Although this kind of observational study is not the traditional research type article that PLoS Pathog publishes, we are considering a short/discovery report type format for observations that are of particular significance and/or broad impact.  

We believe the authors' observation falls into that category. Based on the reviews, we are likely to accept this manuscript for publication, providing that you modify the manuscript according to the review recommendations.

Please address the comments of both reviewers constructively.  We concur that addition of a well-constructed  phylogenetic tree to show that the R1 lineage forms a monophyletic clade would be important (Reviewer #2 comments).  This editor also suggests that the authors discuss the potential significance of the W152L mutation. Note that W152C is a defining mutation in B.1.429, an emerging Variant of Interest, and is located in the NTD antigenic supersite i (PMID: 33761326, 33821281).  In addition, please ensure that the manuscript is carefully proof-read to correct for grammar, spelling and syntax. Some incomplete sentences (e.g line 152-154) detract from a major point that the authors might want to make. With regards to the Japanese sequence data in Fig. 2B, please update data to Mar 1 if available.   

Please prepare and submit your revised manuscript within 30 days. If you anticipate any delay, please let us know the expected resubmission date by replying to this email.

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[1] A letter containing a detailed list of your responses to all review comments, and a description of the changes you have made in the manuscript.

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Thank you again for your submission to our journal. We hope that our editorial process has been constructive so far, and we welcome your feedback at any time. Please don't hesitate to contact us if you have any questions or comments.

Sincerely,

Benhur Lee

Section Editor

PLOS Pathogens

Benhur Lee

Section Editor

PLOS Pathogens

Kasturi Haldar

Editor-in-Chief

PLOS Pathogens

​orcid.org/0000-0001-5065-158X

Michael Malim

Editor-in-Chief

PLOS Pathogens

orcid.org/0000-0002-7699-2064

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Reviewer Comments (if any, and for reference):

Reviewer's Responses to Questions

Part I - Summary

Please use this section to discuss strengths/weaknesses of study, novelty/significance, general execution and scholarship.

Reviewer #1: This is a straightforward case study describing household spread in Japan of the R.1 lineage of SARS-CoV-2. The study is generally well written. It is quite medically oriented and rather observational. There are no experiments.

Reviewer #2: This paper gives an initial description of a novel SARS-CoV-2 lineages, R.1, which appears to be mainly circulating in the USA and Japan, with a focus on the cases in Japan. R.1 carries the E484K mutation in spike, also shared by the 501Y.V2 and 501Y.V3 variants of concern, first identified in South Africa & Brazil, respectively, which is thought to be associated with possibly evading immune response, allowing reinfection and/or vaccine evasion.

This is a clear and well-written paper, presented in a level-headed manner. It is a descriptive work of a novel lineage, so there is no great detail about changes in viral behaviour or speculation of origins, but for a short initial description I do not think that is necessarily to be expected. The methods and results are detailed well and sufficient information is given to identify the lineage clearly in other sequences and to gain an understanding of the lineage and its spread so far.

I am not so familiar with the new format I believe this was submitted under therefore will leave to the editor to determine if this satisfied those requirements, but I believe this is a scientifically sound paper giving a good description of a novel SARS-CoV-2 lineage. I would recommend only 1 major and 1 minor changes before acceptance.

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Part II – Major Issues: Key Experiments Required for Acceptance

Please use this section to detail the key new experiments or modifications of existing experiments that should be absolutely required to validate study conclusions.

Generally, there should be no more than 3 such required experiments or major modifications for a "Major Revision" recommendation. If more than 3 experiments are necessary to validate the study conclusions, then you are encouraged to recommend "Reject".

Reviewer #1: Epidemiological event of R.1 lineage section should discuss sampling bias.

Reviewer #2: I believe this paper is a good description of the R.1 lineage, and useful to those who wish to identify it and understand its initial distribution and spread. However, I think this paper would benefit greatly from even a very basic phylogenetic tree of the lineage and the sequences it covers. I think this would allow even a quick look at the relationships between the global sequences, and to show that this is indeed one monophyletic clade.

From looking at this clade on the focal Nextstrain 484K build (https://nextstrain.org/groups/neherlab/ncov/S.E484?c=country&f_pango_lineage=R.1&label=mlabel:20B/C18877T), it does indeed seem to be, but sometimes in the paper it seems like perhaps this is presented as ambiguous (though this may not have been intentional, and I may be misinterpreting the writing - apologies if so). For example in lines 147-148, saying "...implying that [the lineage] may have emerged in several regions at approximately the same time."

Further a phylogeny would help to clarify the meaning of statement like in lines 168-169: "The circulating trend of R.1 lineage showed similar pattern (note: typo, should be 'patterns') in each country, implying that the ancestral strain acquired homegrown variants, and subsequently R.1 lineage are likely to emerge in several countries." Is there evidence of unique R.1 variants in different countries? (I actually think from the Nextstrain tree, there may be - but showing a tree would display this more clearly, if so.)

I think even a fairly basic phylogeny as an additional figure, with a small additional description of what can be seen, would add a lot of value to the paper in showing any clusters in any countries, the relationship between USA & Japanese sequences, and the genetic diversity of the cluster (including any sub-clusters within the lineage with S or other AA mutations).

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Part III – Minor Issues: Editorial and Data Presentation Modifications

Please use this section for editorial suggestions as well as relatively minor modifications of existing data that would enhance clarity.

Reviewer #1: There are a number of minor typos / grammar problems.

I also did not see a supplemental data table to conform with the GISAID terms of use.

Reviewer #2: I understand the world of SARS-CoV-2 research moves incredibly quickly, but I fear by the time this work is published it may seem quite outdated, given the data was collected through Feb 2021. As the work is descriptive I do think much of the value lies in the descriptions being as up-to-date as possible, so I would urge the authors to update the paper (counts, global coverage, etc) to cover data through as recent a period as is possible. This would also possibly shed some clarity onto statements like that of 153-154 ("it is possible that R.1 lineage is gradually increasing."

Minor typos/wording: Line 170 "acquired" should be "acquires". Line 174, should be "therefore, new emerging strain[s] [possibly] have been missed."

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Reviewer #1: No

Reviewer #2: No

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References:

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.

Dear Dr. Hirotsu,

We are pleased to inform you that your manuscript 'Detection of R.1 lineage severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with spike protein W152L/E484K/G769V mutations in Japan' has been provisionally accepted for publication in PLOS Pathogens.

Before your manuscript can be formally accepted you will need to complete some formatting changes, which you will receive in a follow up email. A member of our team will be in touch with a set of requests.

In addition, we would like to encourage the authors to deposit their whole genome sequences of R.1 lineage viruses into properly curated databases like GISAID so that your efforts can be properly acknowledged. We believe your findings should add to the usefulness of such databases.  

Please note that your manuscript will not be scheduled for publication until you have made the required changes, so a swift response is appreciated.

IMPORTANT: The editorial review process is now complete. PLOS will only permit corrections to spelling, formatting or significant scientific errors from this point onwards. Requests for major changes, or any which affect the scientific understanding of your work, will cause delays to the publication date of your manuscript.

Should you, your institution's press office or the journal office choose to press release your paper, you will automatically be opted out of early publication. We ask that you notify us now if you or your institution is planning to press release the article. All press must be co-ordinated with PLOS.

Thank you again for supporting Open Access publishing; we are looking forward to publishing your work in PLOS Pathogens.

Best regards,

Benhur Lee

Section Editor

PLOS Pathogens

Benhur Lee

Section Editor

PLOS Pathogens

Kasturi Haldar

Editor-in-Chief

PLOS Pathogens

​orcid.org/0000-0001-5065-158X

Michael Malim

Editor-in-Chief

PLOS Pathogens

orcid.org/0000-0002-7699-2064

***********************************************************

Reviewer Comments (if any, and for reference): The addition of the phylogenetic tree was edifying, in addition to the updated data showing the increasing spread of the R1 lineage in Japan. We encourage the authors to deposit  their full-length genomic sequence into properly curated databases like GISAID so that their efforts can be acknowledged.