Complete genome sequences of tolaasin-detoxifying Mycetocola species isolated from oyster mushroom
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Complete genome sequences of tolaasins-detoxifying Mycetocola spp., M. lacteus MAFF 211326, M. saprophilus MAFF 211324, and M. tolaasinivorans MAFF 211325, were determined. The length of circular chromosome of these strains was 3.47, 3.52, and 3.49 Mbp, respectively, and a 28,409-bp plasmid was found in M. lacteus MAFF 211326. Genomic comparisons of mushroom-associated, -pathogenic, and environmental strains of Mycetocola spp. showed that the genomic synteny of chromosomes was largely shared among Mycetocola spp. Furthermore, the previously estimated catalytic triad of tolaasin-degrading factor (Tdf) is conserved among the genomes of strains isolated from mushrooms and non-mushroom substrates. Lipopeptides, including tolaasins, are biologically active and involved in various biological interactions. The antagonistic traits of Mycetocola spp. might not be restricted to tolaasin detoxification; therefore, genome information of Mycetocola spp. will facilitate the mining and identification of enzymes that cleave cyclic lipopeptides acting in various biological interactions observed in agricultural and ecological contexts.
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Dear Dr Takashima, Thank you for your submission, following reviews i have selected major revisions. I'm pleased to say both reviewers where complementary of the manuscript however, some of the recommended changes may take some time to achieve. Please address reviewers comments. Best wishes, John.
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Comments to Author
Comments to authors Manuscript: "Complete genome sequences of tolaasin-detoxifying Mycetocola species isolated from oyster mushroom" In this manuscript, the authors present the complete genomes of three oyster mushroom-associated Mycetocola strains using ONT long-read sequencing. The key gene, tolaasin-degrading factor (Tdf), was characterized and compared across mushroom-associated, mushroom-pathogenic, and environmental strains. Overall, the concept of the manuscript is interesting, and the experiments are rigorously conducted; however, some aspects lack sufficient evidence or the use of reliable tools to support the conclusions. Given the availability of complete genomes in this study, the authors could also predict secondary metabolite-encoding genes that may be involved in detoxification …
Comments to Author
Comments to authors Manuscript: "Complete genome sequences of tolaasin-detoxifying Mycetocola species isolated from oyster mushroom" In this manuscript, the authors present the complete genomes of three oyster mushroom-associated Mycetocola strains using ONT long-read sequencing. The key gene, tolaasin-degrading factor (Tdf), was characterized and compared across mushroom-associated, mushroom-pathogenic, and environmental strains. Overall, the concept of the manuscript is interesting, and the experiments are rigorously conducted; however, some aspects lack sufficient evidence or the use of reliable tools to support the conclusions. Given the availability of complete genomes in this study, the authors could also predict secondary metabolite-encoding genes that may be involved in detoxification processes, which would either support or add value to the identification of putative Tdf genes. However, the writing remains unclear and requires better organization. The discussion is limited and lacks sufficient reasoning. Therefore, I suggest that the manuscript be majorly revised according to the comments and suggestions provided below. Materials and methods L89-90: How long was the sequencing run? (by default, 72 hours?) L89: Which PromethION platform? (P2, P2 solo or P24?) Please add more information. L90-97: Actually, a Q score ≥10 (indicating higher accuracy) is used by default for the SUP model (SUP v5.0.0), so it is unclear why the authors still used Q score 9 for this step. L102: Did the authors perform an error correction step after assembly with Flye? If not, the assembled genome may still contain a relatively high error rate, even though the latest SUP model (v5.0.0) was used for basecalling. L104-108: Which program did the authors use to predict the plasmid contigs from the assembled genome? I would suggest that the authors use MOB-suite or Plasmer, which are widely used in this field. Moreover, completeness and contamination should be mentioned in this section (DFAST?) L129-131: M&M should start with the objective of the experiment. Therefore, "Tolaasin-degrading factor (Tdf) was identified as a serine protease of the S9 prolyloligopeptidase family with a catalytic triad (Ser554, Asp641, and His680 in the positions of prolyl endopeptidase of Sus scrofa) [8]." Should be omitted. L131-133: "According to DFAST annotation of the MAFF strains and BLASTP searches during genome synteny analysis, putative Tdf proteins of the MAFF strains and the other three complete genomes used for genome synteny were found." Since this information appears to be part of the Results section, I would suggest revising the "Alignment of putative tolaasin-degrading factor (Tdf)" section accordingly to avoid redundancy. Results and discussion L14-145: Please consider rewriting this section to improve readability. Since the numerical data are already presented in Table 1, it may be unnecessary to restate them in the text in order to avoid redundancy. L155: In this section, I would recommend that the authors discuss the host- or environment-specific genomic signatures within Mycetocola strains in more detail. Upon examining Fig. 1, it appears that the strains cluster into approximately 15 subgroups based on ANIb relationships. Therefore, the authors should additionally describe this observation in the manuscript. L161-164: Please consider rewriting this section for improved clarity and readability. For example, our three assembled genomes showed 100% ANIb values when compared to previously deposited wild strains, clearly indicating that they belong to the same species. L165: What do the colours (red, blue, green, orange…) next to the dendrograms mean? Is there any significant in the group? L175-176: I do not fully understand what the authors intended to convey with the sentence, "Complete genome sequences of Mycetocola spp. have been determined for three strains: M. spongiae MSC19, Mycetocola sp. JXN-3, and M. zhujimingii CGMCC 1.16372 [11, 14, 15]." This paragraph should be rewritten for clarity. For example, the authors could state, "Among the 31 genome sequences available for the Mycetocola genus (based on NCBI database records), only three genomes — M. spongiae MSC19, Mycetocola sp. JXN-3, and M. zhujimingii CGMCC 1.16372 — have been fully completed [11, 14, 15]. Among these, a 32,244-bp plasmid (CP026950.1) has been identified only in M. zhujimingii CGMCC 1.16372. Therefore, ..." L179-180: How many genes are shared among the species? Please specify and discuss more about this point. L189-190: Change to "Mobilizable plasmids typically require at least an oriT region and a relaxase for transfer [26]. However, no oriT region was detected in the M. lacteus plasmid, suggesting that its potential for mobilization is uncertain." Minor points L85: day L88: (SQK-RBK114.24) L168: (15 Liu et al., 2020)? L229: or famide A L226: "Mycetocola spp." changes to "this bacteria" Figures Fig. 2 and Supplementary Fig. 1 appear to be identical; therefore, Supplementary Fig. 1 should be omitted from the manuscript. Please check the entire manuscript carefully to ensure consistency.
Please rate the manuscript for methodological rigour
Satisfactory
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Partially support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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Comments to Author
This is a well written a robust paper. The methodology is clear and sound. All raw data is accessible. There are however some minor grammatical errors and figure legends could be better clarified: Line 26: "tolaasins-detoxifying" should be corrected to "tolaasin-detoxifying" Line 27/28: Change "The length of circular chromosome" to "The length of the circular chromosomes" Line 55: "as a biological control agent" should be corrected to "as biological control agents" Line 85: "4 d" should be written as "4 days" Line 166: Consider removing "certainly" as it overstates certainty based only on ANI values Additionally clarification should be added to Figure 1. A scale bar may be useful to show genetic distance. Furthermore, the coloured bars at the tips of the dendrogram branches are not explained. …
Comments to Author
This is a well written a robust paper. The methodology is clear and sound. All raw data is accessible. There are however some minor grammatical errors and figure legends could be better clarified: Line 26: "tolaasins-detoxifying" should be corrected to "tolaasin-detoxifying" Line 27/28: Change "The length of circular chromosome" to "The length of the circular chromosomes" Line 55: "as a biological control agent" should be corrected to "as biological control agents" Line 85: "4 d" should be written as "4 days" Line 166: Consider removing "certainly" as it overstates certainty based only on ANI values Additionally clarification should be added to Figure 1. A scale bar may be useful to show genetic distance. Furthermore, the coloured bars at the tips of the dendrogram branches are not explained. What are they there for? Clarification could also be added to Figure 2. The "flipped" and "regular" legend is not clear. Clarify what is being flipped and what each classification means in biological or visual terms.
Please rate the manuscript for methodological rigour
Very good
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Strongly support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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