Peach Buds’ Microbiome Profiling Reveals Cultivar-Specific Signatures Associated with TCSB Susceptibility

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Article Peach buds’ microbiome profiling reveals cultivar-specific signatures associated with TCSB susceptibility by Antonella Cardacino, Taner Tastekin, Federico Brugneti, Marco Cirilli, Angelo Mazzaglia, Silvia Turco reveals the features of the peach tree microbiome depending on varietal characteristics.
The manuscript is formatted according to the rules and contains the necessary sections except for the conclusion section.

The presented data on diversity characterize the influence of the genotype on the composition of the microbiome and are quite convincing, the methods correspond to the questions posed.
However, it would be useful to improve the manuscript to characterize the differences in the surface structure, chemical composition of the cover or other parameters of the studied genotypes, for example, the presence of trichomes, surface features leading to such differences. It is optimal to provide photographs and parameters of the object under study. It is also unclear whether the location on the plant affects it and, if so, how. I recommend adding a section describing the plant material in detail, the characteristics of the sample produced with the help of statistics, and indicating the exact locations of the material collection. I recommend paying special attention to formulating the hypothesis and forming a conclusion with an assessment of the novelty of the results obtained and the prospects for using this study with the addition of the Conclusion section.

Author Response

Article Peach buds’ microbiome profiling reveals cultivar-specific signatures associated with TCSB susceptibility by Antonella Cardacino, Taner Tastekin, Federico Brugneti, Marco Cirilli, Angelo Mazzaglia, Silvia Turco reveals the features of the peach tree microbiome depending on varietal characteristics. The manuscript is formatted according to the rules and contains the necessary sections except for the conclusion section.

The presented data on diversity characterize the influence of the genotype on the composition of the microbiome and are quite convincing, the methods correspond to the questions posed.
However, it would be useful to improve the manuscript to characterize the differences in the surface structure, chemical composition of the cover or other parameters of the studied genotypes, for example, the presence of trichomes, surface features leading to such differences. It is optimal to provide photographs and parameters of the object under study. It is also unclear whether the location on the plant affects it and, if so, how. I recommend adding a section describing the plant material in detail, the characteristics of the sample produced with the help of statistics, and indicating the exact locations of the material collection. I recommend paying special attention to formulating the hypothesis and forming a conclusion with an assessment of the novelty of the results obtained and the prospects for using this study with the addition of the Conclusion section.

We thank the reviewer for their constructive suggestions, which have significantly improved our manuscript. In response to the recommendations, we have:

• Expanded the Materials and Methods section to provide a more detailed description of the plant material, including the exact collection locations and phenological stage.

• Added a paragraph in the Introduction discussing possible differences in surface features of the buds (e.g., trichomes, epidermal properties) that may influence microbial colonization, citing relevant literature. We note that direct morphological or chemical analyses were beyond the scope of this study.

• Clarified the sampling position on the plant to minimize variability; samples were consistently collected from one-year-old twigs from the median portion of the canopy.

• Added a dedicated Conclusion section summarizing the main findings, highlighting the novelty of our results, and discussing future perspectives for application in plant health and breeding.

We would also like to emphasize that detailed analyses of morphological structures, the chemical composition of the coverings, the presence of trichomes, and surface features that differentiate cultivars are outside the scope of this manuscript.

Reviewer 2 Report

Comments and Suggestions for Authors

Three peach tree varieties 'Catherina', 'Pavoro® Pav 1605'and  'Lami®.COM’ were selected    to analysis microbiome based on the differences in resistance to Twig Canker and Shoot Blight( TCSB). It analyzes the microbial diversity in peach buds through 16S analysis for bacteria and ITS analysis for fungi. The article is highly innovative and provides new insights into the research on varieties and resistance. 

1. The repeatability in the Pav 1605 of Figure 1 is poor，It is not as good as the repeatability of the fungal community. Why are there so few Sphingomonas in this  sample ? 

2. Is Alternaria in Lami®.COM endophytic or pathogenic?

Author Response

Three peach tree varieties 'Catherina', 'Pavoro® Pav 1605'and  'Lami®.COM’, were selected to analysis microbiome based on the differences in resistance to Twig Canker and Shoot Blight( TCSB). It analyzes the microbial diversity in peach buds through 16S analysis for bacteria and ITS analysis for fungi. The article is highly innovative and provides new insights into the research on varieties and resistance.

1. The repeatability in the Pav 1605 of Figure 1 is poor，It is not as good as the repeatability of the fungal community. Why are there so few Sphingomonasin this  sample ? 

We thank the reviewer for this insightful observation. We acknowledge that the bacterial community in ‘Pavoro® – Pav 1605’ exhibited higher variability across replicates compared to the fungal community. Such variability may reflect natural biological differences, as microbial communities in buds can be influenced by subtle environmental and physiological factors even under standardized sampling conditions. Alternatively, it could be influenced by technical factors such as uneven DNA pooling or sequencing variability. Regarding Sphingomonas, its consistently low abundance across replicates in ‘Pavoro® – Pav 1605’ suggests a genuine biological pattern rather than a technical artifact. The abundance of Sphingomonas is known to vary significantly among host genotypes and tissues, as reported in other fruit trees. We have now emphasized these points in the Discussion section.

2. Is Alternariain Lami®.COM endophytic or pathogenic?

We appreciate this important question. In our dataset, Alternaria was detected as part of the fungal community inhabiting dormant buds of ‘Lami®.COM’. The genus Alternaria includes both latent pathogens and endophytes, depending on the host and environmental conditions. Since our study used metabarcoding, we cannot infer the functional lifestyle of Alternaria in this specific context. However, given that the buds were asymptomatic at the time of collection, it is likely that Alternaria was present in an endophytic or latent form rather than actively causing disease. We have clarified this point in the Discussion.

Reviewer 3 Report

Comments and Suggestions for Authors

The article by Cardacino et al. is devoted to the topical issue of analyzing the natural microbiome of plants in order to identify the possible participation of indigenous microflora in plant resistance to phytopathogens. The work is quite compact, logical, and performed using adequately selected methods. Discussion of the results allows us to understand the authors' hypothesis about the relationship between the diversity of plant microflora and the resistance of peach plants to diseases.

 

The authors have done a lot of work to identify patterns in the formation of microbiota (bacteria and fungi) in peach buds from a variety of plant. The main goal was to identify the presence and even dominance of pathogenic microflora and to link this fact with the genetic characteristics of peach varieties.

 

This is a fairly popular and relevant scientific direction. Obviously, it is not without justification, since everyone knows that some varieties of cultivated plants are quite sensitive to phytopathogens. Is it original? Within the framework of the study, yes. Within the framework of the scientific direction, it has rather become routine. Does this topic fill a gap in this area of ​​research. Rather, this work should be the beginning of a cycle of further, more in-depth studies.

 

I like the rationale behind the main approach to the study and the number of different peach varieties selected, as well as the methods used for the analysis. In this case, since the search is for the root cause of the damage to peach fruits, the study of the buds is clearly justified. The authors are trying to link this with the genetic characteristics of the varieties.

 

In my opinion, this work has a weak point. Of course, cultivated plants have varietal variability in disease resistance. But we understand that with phytopathogenic diseases, it is necessary to take into account climatic and weather conditions, agricultural technologies used, etc. I would like to identify genetic patterns of disease sensitivity. How quickly can this problem be solved? An analysis of the scientific literature shows that it is unlikely to be anytime soon. But each study in this area allows us to move forward, albeit little by little. 

The main results of the study are clearly displayed in illustrations, which allows us to assess their reliability.

The references do not cause any comments.

 

Author Response

The article by Cardacino et al. is devoted to the topical issue of analyzing the natural microbiome of plants in order to identify the possible participation of indigenous microflora in plant resistance to phytopathogens. The work is quite compact, logical, and performed using adequately selected methods. Discussion of the results allows us to understand the authors' hypothesis about the relationship between the diversity of plant microflora and the resistance of peach plants to diseases.

 The authors have done a lot of work to identify patterns in the formation of microbiota (bacteria and fungi) in peach buds from a variety of plant. The main goal was to identify the presence and even dominance of pathogenic microflora and to link this fact with the genetic characteristics of peach varieties.

 This is a fairly popular and relevant scientific direction. Obviously, it is not without justification, since everyone knows that some varieties of cultivated plants are quite sensitive to phytopathogens. Is it original? Within the framework of the study, yes. Within the framework of the scientific direction, it has rather become routine. Does this topic fill a gap in this area of ​​research. Rather, this work should be the beginning of a cycle of further, more in-depth studies.

 I like the rationale behind the main approach to the study and the number of different peach varieties selected, as well as the methods used for the analysis. In this case, since the search is for the root cause of the damage to peach fruits, the study of the buds is clearly justified. The authors are trying to link this with the genetic characteristics of the varieties.

 In my opinion, this work has a weak point. Of course, cultivated plants have varietal variability in disease resistance. But we understand that with phytopathogenic diseases, it is necessary to take into account climatic and weather conditions, agricultural technologies used, etc. I would like to identify genetic patterns of disease sensitivity. How quickly can this problem be solved? An analysis of the scientific literature shows that it is unlikely to be anytime soon. But each study in this area allows us to move forward, albeit little by little. 

The main results of the study are clearly displayed in illustrations, which allows us to assess their reliability.

The references do not cause any comments.

 

We thank the reviewer for the thoughtful and constructive comment. We fully agree that the variability in disease susceptibility among peach cultivars is influenced not only by genotype, but also by environmental conditions and management practices. 

We acknowledge that the originality of the study lies in being the first characterization of bacterial and fungal communities in dormant buds of multiple peach cultivars in relation to Diaporthe amygdali. We have now clarified this in the Introduction and Conclusion sections, highlighting the study as a foundation for future in-depth research.

As you also mentioned, this work marks the beginning of a cycle aimed at highlighting potential genotype-associated patterns in the bud microbiome. In parallel (and in extreme confidence), we are conducting a broad susceptibility test of different cultivars to be coupled with more advanced techniques like transcriptomics and GWAS. 

We also appreciate the positive feedback regarding the clarity of the figures, the selection of cultivars, and the methodological approach.