Review Reports

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The review addresses an important topic in plant pathology. Several aspects related to plant resistance to pathogens and their interactions are discussed. However, the manuscript requires significant improvement before publication. The authors should strengthen the introduction and provide clearer and more detailed descriptions of the pathogen domain examples. Some tables need to be restructured to better highlight key information. The discussion of certain strategies is limited and would benefit from further investigation and expansion. Additionally, the manuscript contains several typographical errors that must be corrected prior to publication.

 

Keywords: The two keywords (“LysM domain-containing proteins” and “plant pathogenic fungi”) should be different from those used in the title.

Lines 34–36: The authors should include the key review by Savary (2019), which addresses the different threats discussed in this report.

Line 36: The connection between the preceding and following ideas should be improved. The authors move directly to the main subject without providing a clear transition or outline that links the context of the review to its core topic.

For all scientific names used throughout the manuscript, it is recommended to include the authority names. The Catalogue of Life can be consulted to ensure the correct nomenclature.

Line 55: Are CEBiP and CERK1 genes? Please clarify this point and indicate the context in which they are discussed.

Line 66: Please use punctuation correctly and do not forget to include authority names for scientific species.

Line 83: The author needs to specify and clearly define HIGS.

Line 153: “oyher” changes other. Please revise.

Table 1: Please explain the meaning of “Sp” and “TM”.

Line 226: Replace “such as C. higginsianum ChElp1 and ChElp2” with “such as ChElp1 and ChElp2 in C. higginsianum”.

Lines 230–233: The authors could include additional experiments involving other pathogens using this novel strategy, which would strengthen the manuscript.

Author Response

Responses to Reviewer: 1

General comment:

The review addresses an important topic in plant pathology. Several aspects related to plant resistance to pathogens and their interactions are discussed. However, the manuscript requires significant improvement before publication. The authors should strengthen the introduction and provide clearer and more detailed descriptions of the pathogen domain examples. Some tables need to be restructured to better highlight key information. The discussion of certain strategies is limited and would benefit from further investigation and expansion. Additionally, the manuscript contains several typographical errors that must be corrected prior to publication.

Response: We thank the reviewer for the positive evaluation and for the insightful suggestions. We have thoroughly revised the manuscript according to the comments. The introduction has been strengthened by adding a recent global assessment of crop losses and by clarifying the distinction between our review and previous ones. The description of pathogen domain examples has been expanded, and Table 1 has been restructured with a clearer title and explanatory notes. The discussion on disease control strategies has been expanded with more examples and mechanistic details. All typographical errors have been corrected.

1. The two keywords ("LysM domain-containing proteins" and "plant pathogenic fungi") should be different from those used in the title.

Response: We agree with the reviewer. The keywords have been modified to avoid repetition with the title. The new keywords are: "LysM effectors; phytopathogenic fungi; virulence factor; chitin-triggered immunity; immune evasion; disease control".

2. Lines 34–36: The authors should include the key review by Savary (2019), which addresses the different threats discussed in this report.

Response: We thank the reviewer for this important reference. We have now included the Savary et al. (2019) paper in the Introduction (now in lines 41-43) and added it to the reference list as new reference. The sentence now reads: “A recent global assessment quantified the burden of pathogens and pests on major food crops, reporting yield losses ranging from 17.2% to 30.0% depending on the crop, with rice suffering the highest losses [3]. These figures underscore the critical need to understand the molecular mechanisms underlying fungal pathogenicity and plant immunity.”

3. Line 36: The connection between the preceding and following ideas should be improved. The authors move directly to the main subject without providing a clear transition or outline that links the context of the review to its core topic.

Response: We appreciate this observation. To improve the logical flow, we have added a paragraph at the end of the abstract (lines 28-28) and introduction (now in lines 94-110) that explicitly states the aims of this review, explains how it differs from previous reviews, and outlines the structure. This provides a clear transition from the general context to the specific focus on LysM proteins in plant pathogenic fungi.

4. For all scientific names used throughout the manuscript, it is recommended to include the authority names. The Catalogue of Life can be consulted to ensure the correct nomenclature.

Response: We thank the reviewer for this important suggestion. We have now added authority names for all fungal species at their first mention in the text (e.g., Magnaporthe oryzae B.C. Couch, Zymoseptoria tritici(Desm.) Quaedvl. & Crous, Cladosporium fulvum Cooke, Colletotrichum higginsianum Sacc., etc.). The Catalogue of Life was consulted to verify the correct nomenclature. These changes appear throughout the Introduction and subsequent sections.

5. Line 55: Are CEBiP and CERK1 genes? Please clarify this point and indicate the context in which they are discussed.

Response: We thank the reviewer for pointing out this ambiguity. In the revised manuscript (now in lines 57-62), we have clarified: “Chitin, a linear polymer of N-acetylglucosamine (GlcNAc) residues, is the major component of fungal cell walls and a typical PAMP that can be recognized by plant LysM domain containing PRRs, such as the rice chitin-elicitor binding protein (CEBiP) and chitin-elicitor receptor kinase-1 (OsCERK1). These two proteins, encoded by CEBiP and OsCERK1 genes, respectively, cooperatively regulate chitin-triggered immunity (CTI) in rice [9].”

6. Line 66: Please use punctuation correctly and do not forget to include authority names for scientific species.

Response: We have corrected punctuation and added authority names for the species mentioned in that line (now in lines 72). The sentence now reads: “Since the first fungal LysM domain-containing effector Ecp6 was identified from Cladosporium fulvum Cooke in 2008 [14]”

7. Line 83: The author needs to specify and clearly define HIGS.

Response: We have now defined HIGS (host-induced gene silencing) at its first appearance in the Introduction (now in lines 91). Additionally, in Section 4, we provide a more detailed explanation of the HIGS mechanism with a new figure (Figure 3A) and accompanying text (now in lines 268-278).

8. Line 153: "oyher" changes other. Please revise.

Response: We apologize for the typo. It has been corrected to “other” in the revised manuscript (now in line187, where the classification is described).

9. Table 1: Please explain the meaning of "Sp" and "TM".

Response: We have added a clear explanation in the table title and legend. The title now reads: “Table 1. LysM domain-containing proteins in four representative plant pathogenic fungi. The table lists gene identifiers, domain architectures (LysM copy number and additional domains), presence of signal peptide (SP) and transmembrane domain (TM), and relevant references for each protein. ”.

10. Line 226: Replace "such as C. higginsianum ChElp1 and ChElp2" with "such as ChElp1 and ChElp2 in C. higginsianum".

Response: We have made this change as suggested (now in line 293). The sentence reads: “...such as ChElp1 and ChElp2 in C. higginsianum [55].”

11. Lines 230–233: The authors could include additional experiments involving other pathogens using this novel strategy, which would strengthen the manuscript.

Response: We agree that including more examples of novel strategies applied to other pathogens strengthens the manuscript. In the revised Section 4, we have expanded the discussion to include additional studies, such as the use of RNA fungicides targeting Botrytis cinerea and the potential of LysM-fused chitinases (now in lines 261-308). We also discuss the need for further field studies, which is now included in the future perspectives.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

LysM domain-containing proteins are well-known to participate in plant-microbe interactions. This review summarize the advance of these proteins in their structures, mechanism of functions, and their potential usage in plant disease control. This is interesting to the readers. This review is generally well organized. However, some issues should be addressed before a recommendation of acceptance could be made.

Major:

1. Since LysM domain-containing proteins are famous in the field of plant pathology, I believe, reviews on these proteins have been made in literature. Have you cited them in your review? And how does your review differ from those previous reviews? 
2. What's the coverage of this review? all the plant fungal pathogens? what's the year spanning?
3. Judging from the text, only some species of fungi have been selected for investigating the LysM proteins. What's the criterion for the selection? In other words, why only select four fully sequenced species as shown in Table 1? Based on the crops? fungal taxonomy? This needs to be reconsidered. Even for wheat, Fusarium diseases such as Fusarium head blight are severe, not only the Septoria tritici blotch caused by Zymoseptoria tritici. More plant fungal pathogens are suggested to include for analysis.

Minor:

1. Citations to each subfigure should be clearly indicated in the text. For instance, fig, 1A should be cited in the text, also that of fig. 1B, 1C, and 1D.
2. line 141, delete "…and below…", since no annotations are placed below the sequences.
3. lines 169-170, “…from 4 fully sequenced species…”So, which four?
4. line 207, “…(Table 1)”. Why place table 1 here? Obviously, table 1 has nothing to do with the functional mechanisms. 
5. lines 227-228, "...,the use of endolysins to hinder bacterial growth in transgenic plants, indicating it ..." what does that mean?
6. Many spelling errors, Such as line 212, should be “pathogenic”, but not “pathogenici”.

 

Author Response

Responses to Reviewer: 2

General comment: LysM domain-containing proteins are well-known to participate in plant-microbe interactions. This review summarizes the advance of these proteins in their structures, mechanism of functions, and their potential usage in plant disease control. This is interesting to the readers. This review is generally well organized. However, some issues should be addressed before a recommendation of acceptance could be made.

Response: We thank the reviewer for the positive assessment and for the constructive suggestions that have helped us improve the manuscript.

Major Comments 1: Since LysM domain-containing proteins are famous in the field of plant pathology, I believe, reviews on these proteins have been made in literature. Have you cited them in your review? And how does your review differ from those previous reviews?

Response: We thank the reviewer for this important question. We agree that several excellent reviews on LysM domain-containing proteins have been published over the past decade. In the revised manuscript, we have now explicitly cited these key reviews [11,29,30] and clearly articulated how our work differs from and advances beyond them.

Specifically, we have added the following paragraph in the Introduction (now in lines 94-110): "Over the past decade, the roles of LysM domain-containing proteins in microbial pathogenesis and symbiosis have been extensively reviewed [29]. These studies have established a broad framework for understanding LysM protein functions across kingdoms, particularly in the context of plant-microbe interactions [30]. However, existing reviews have primarily focused on evolutionary conservation or symbiotic signaling, with limited emphasis on the mechanistic diversity and virulence-specific roles of LysM effectors in plant pathogenic fungi. Recent advances in structural biology, comparative genomics, and effector biology have unveiled new layers of complexity, including intramolecular dimerization, host-specific recognition, and co-evolutionary dynamics between fungal effectors and plant receptors. Therefore, this review provides an updated synthesis focused exclusively on plant pathogenic fungi, systematically summarizing their structural characteristics, classification, functional mechanisms, representative species studies, application in disease control, and future perspectives, with an emphasis on the latest achievements from 2020-2025. This work aims to deepen the understanding of the molecular basis of fungal virulence and plant–fungal interactions, and to provide new ideas for the development of efficient and environmentally friendly fungal disease control strategies."

In this revision, we have (1) acknowledged and cited previous reviews, (2) clearly distinguished our review by its exclusive focus on plant pathogenic fungi and virulence-specific mechanisms, and (3) highlighted the inclusion of recent advances (2020-2025) that have not been systematically covered in earlier works. We believe this addresses the reviewer's concern and clarifies the unique contribution of our manuscript.Thank you again for this valuable suggestion, which has helped us strengthen the manuscript's framing and significance.

Major Comment 2: What's the coverage of this review? all the plant fungal pathogens? what's the year spanning?

Response: We thank the reviewer for this important question regarding the coverage of our review.This review focuses on plant pathogenic fungi but does not aim to cover all species exhaustively. Instead, we concentrate on representative model species from major ascomycete lineages (particularly Sordariomycetes and Dothideomycetes) for which LysM proteins have been functionally characterized, including Magnaporthe oryzae, Zymoseptoria tritici, Colletotrichum higginsianum, Verticillium dahliae, Cladosporium fulvum, and Botrytis cinerea. These species were selected because they represent distinct infection strategies (hemibiotrophic, necrotrophic, vascular) and possess the most comprehensive functional data on LysM effectors.

The literature coverage spans from foundational studies in the early 2000s to the most recent advances reported up to 2025, with emphasis on developments from 2020–2025 in structural biology, functional mechanisms, and disease control applications. Foundational studies are included to provide historical context.

We have clarified this scope in the revised Introduction (now in lines 94-110). Due to the vast diversity of phytopathogenic fungi, a comprehensive analysis of all species is not feasible; instead, our focused approach enables meaningful comparative analysis across distinct infection strategies.

Major Comment 3: Judging from the text, only some species of fungi have been selected for investigating the LysM proteins. What's the criterion for the selection? In other words, why only select four fully sequenced species as shown in Table 1? Based on the crops? fungal taxonomy? This needs to be reconsidered. Even for wheat, Fusarium diseases such as Fusarium head blight are severe, not only the Septoria tritici blotch caused by Zymoseptoria tritici. More plant fungal pathogens are suggested to include for analysis.

Response: We thank the reviewer for this valuable suggestion. In the revised manuscript, we have clarified the criteria for species selection in Table 1, including economic importance, phylogenetic diversity, genome availability, and prior functional evidence for LysM effectors. To address the concern about limited representation, we expanded Table 1 from 4 to 10 species, adding Fusarium verticillioides, Cladosporium fulvum, Colletotrichum gloeosporioides, C. lindemuthianum, C. fructicola, and Rhizoctonia solani. In the main text, we have updated the classification and functional mechanisms sections to incorporate discussions on those newly added species for which sufficient functional data exist (e.g., C. fulvum Ecp6, C. gloeosporioides Cg2LysM, R. solani RsLysM). For species with limited mechanistic studies (e.g., C. lindemuthianum CIH1, C. fructicola CfLysM2, F. verticillioides FvLcp1), we include them in Table 1 to provide a comprehensive inventory and direct readers to relevant references, while keeping the text focused on well-characterized examples. This approach balances completeness with clarity, and we believe it significantly enhances the comparative value of the review.

Minor Comment 1: Citations to each subfigure should be clearly indicated in the text. For instance, fig. 1A should be cited in the text, also that of fig. 1B, 1C, and 1D.

Response: We have carefully reviewed the text and added explicit citations to each subfigure.

Minor Comment 2: line 141, delete "...and below...", since no annotations are placed below the sequences.

Response: We have removed “and below” from the sentence. The revised sentence reads: “Structural annotations are indicated above the sequences for MoCVNH3, Ecp6 and Mg1LysM, respectively.”

Minor Comment 3: lines 169-170, "...from 4 fully sequenced species..."So, which four?

Response: We have explicitly named the four species in the figure legend of Figure 2B (now in lines 217-218): “...from 4 fully sequenced plant pathogenic fungal species: Magnaporthe oryzae 7015, Zymoseptoria tritici (strain ST99CH_3D7), Colletotrichum higginsianum (strain IMI 349063), and Verticillium dahliae (strain VdLs.17).”

Minor Comment 4: line 207, "...(Table 1)". Why place table 1 here? Obviously, table 1 has nothing to do with the functional mechanisms.

Response: We agree that the original placement was suboptimal. In the revised manuscript, we have moved the citation of Table 1 to the end of the classification section (line 256), where it now serves to illustrate the domain architectures of the different types of LysM proteins. The text now reads: “Representative LysM proteins from four major plant pathogenic fungi are listed, illustrating the domain architectures corresponding to each type (Tab.1).” This provides a logical connection between the classification and the table.

Minor Comment 5: lines 227-228, "...the use of endolysins to hinder bacterial growth in transgenic plants, indicating it..." what does that mean?

Response: We have rewritten this sentence for clarity (lines 293-297): “ The application of LysM domains for improving plant resistance also includes the use of endolysins to inhibit bacterial growth in transgenic plants, suggesting that chimeric chitinases fused to LysM domains could potentially be exploited to hinder fungal growth in a diverse range of crop plants [60].” We hope this is now clear.

Minor Comment 6: Many spelling errors, Such as line 212, should be "pathogenic", but not "pathogenici".

Response: We apologize for these spelling errors. Now we have corrected this and other spelling errors throughout the manuscript.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

Thank you for the corrections made in the document and for the improvements.