PONE-D-20-20588

Investigating hospital Mycobacterium chelonae infection using whole genome sequencing and hybrid assembly

PLOS ONE

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Reviewers' comments:

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Comments to the Author

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Reviewer #1: Partly

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

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5. Review Comments to the Author

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Reviewer #1: Good evening Authors,

Thank you for allowing me the chance to review this scientific research. I believe you have some very interesting data here. The use of the V. campbellii isolate as a model to determine the viability of calling identity on fully sequenced and assembled isolates is really cool. However, I do believe that the paper needs a bit more work as far as organization and clarity. Also, I think that some important points about your isolates and the comparison with the reference genomes are missing in the results as well as the discussion. A lot of the discussion comes off as a reiteration of the results. Thus, a re-tooling of the discussion to show the relevance of your findings in the current medical/microbial world or how your finding relate to other studies may be worthwhile. Please see the attached pdf with comments regarding the manuscript. I hope you find them helpful and use them as suggestions to increase the readers understanding of your novel findings.

Reviewer #2: The current study investigates the possible nosocomial transmission of Mycobacterium chelonae strains based on seven patient isolates collected during the period 2017 from the Hospital of the University of Pennsylvania (HUP). For the comparative genomic approach, they have used high-quality hybrid assemblies (Oxford Nanopore long reads and the Illumina as a high-quality short read). To rule out the possibility of sequencing errors Vibrio campbellii dataset has used as a positive control and sequenced 39 times. Single nucleotide variant (SNV) analysis and phylogenetic tree based on core genes (SNVs) reveal infections are possibly transmitted via environment source or other niche but not due to the outbreak at the hospital. Antibiotic resistance-related homolog search and their putative candidate genes list analysis provides more insights into the emerging nontuberculosis mycobacteria research. The manuscript is very clear, the experiments have been conducted diligently with the appropriate controls and overall supportive conclusions.

Minor

1) In Figure 1 a-b) it would be more useful to list all the species names or strain names.

2) In Figure 2) the resolution of the figure should be improved. As of now, it looks blurry image with the text font.

3) In Figure 3) Circosplot, the genome size of Myco1 has shown as 4958837. But for the same, in Table S1) it is 4968398 bp update the right genome size. Represent the outer ring with the genome reference scale (0.5, 1, 1.5,… 5 Mb) that would help readers to identify a unique region precise position.

4) Table S1) Write all the species names, strain names, source names, and accession number of newly sequenced genomes (Myco1-7). Also, Update with the genome coverage information for the Oxford Nanopore and Illumina datasets.

5) Ln 70, delete "that"

6) Update software version numbers for all the tools. For example Prokka ver---, SamTools ver---, CheckM ver---

7) Ln 152, rewrite as " 100% conserved, likely due to the draft genomes".

8) Ln 167, rewrite "45 to 64" instead "46 to 64"

9) Table 2) Footnote: Also write complete abbreviations for R, S and I

10) Ln 194, alignment to a reference genome (Myco1? it has not stated which was the reference was used to map SNVs as a reference).

11) Ln 210, we used all 43 genomes (as of now at NCBI,47 genomes available) possibly mention the latest sequence retrieved date. For these 43 genomes, is it PROKKA annotation or the NCBI annotation that has been used?

12) Ln 264, fewer core genes (3143 vs 3368). Not discussed how similar is this phylogeny compared to the hybrid assemblies phylogeny (if you have the data)? Fig 1a

13) Ln 270, "Providing strong evidence that they are different strains". possibly with the ANI values, this sentence makes strong evidence supporting different strains.

14) Ln 356, a suggestion to delete "Our investigation of the genetic basis of M. chelonae drug resistance was inconclusive". Line 378-379, the statement provides candidate genes.

15) It should be noteworthy, to discuss SNVs to their genomic positions such as is it belong to the coding region or intergenic sequence. Also, How many genes and which genes are affected, and their functional role.

16) SNVs per MB site was discussed in this work. It should be appropriate to compare how this SNVs per MB is in other mycobacteria eg. NTMs or pathogenic mycobacteria.

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Reviewer #1: No

Reviewer #2: No

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Attachments

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Submitted filename: PONE-D-20-20588_reviewer.pdf

Investigating hospital Mycobacterium chelonae infection using whole genome sequencing and hybrid assembly

PONE-D-20-20588R1

Dear Dr. Bushman,

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,

Herman Tse

Academic Editor

PLOS ONE

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