Studies on the Identification of Resistance to Fusarium oxysporum (Schlecht.) in Different Genetic Backgrounds of Asparagus officinalis (L.) and Its Defense Responses
Round 1
Reviewer 1 Report
The article is devoted to an important problem - the search for genotypes resistant to Fusarium in Asparagus. Since Asparagus officinalis cultivars are susceptible to Fusarium, it is a challenge to find donors of resistance. Therefore, the task of the study is very relevant. My biggest question to this work is what determines the choice of particular genotypes and why they differ in the first and second parts of the work.
The part with microsatellite analysis is not clear at all. Why the set of species differs from the first part of the work. Why are these markers chosen? What is known about the genus system? What would the tree look like for basic ITS and chloroplast markers?. Why was this approach not used?
All these should be corrected
Author Response
Thank you for taking the time to read and comment on the paper. Below are the explanations and changes in the order of the comments.
Comments and Suggestions for Authors
The article is devoted to an important problem - the search for genotypes resistant to Fusarium in Asparagus. Since Asparagus officinalis cultivars are susceptible to Fusarium, it is a challenge to find donors of resistance. Therefore, the task of the study is very relevant. My biggest question to this work is what determines the choice of particular genotypes and why they differ in the first and second parts of the work.
In order to study as wide a range of cultivars as possible, we wanted to study older cultivars such as RvB, but also modern cultivars such as Ramada. For the wild species, we mainly chose available genotypes, i.e. those from which we could get enough seeds and which germinated in sufficient numbers. Because of these problems, it is sometimes very difficult to obtain enough seeds for multiple tests. Therefore, by comparing the complete and partial screening, we wanted to show that one screening already has some significance. Although it is always better to do as many replicates as possible.
The part with microsatellite analysis is not clear at all. Why the set of species differs from the first part of the work. Why are these markers chosen? What is known about the genus system? What would the tree look like for basic ITS and chloroplast markers?. Why was this approach not used?
We have added this part mainly to show in advance the possibilities of later crossings. We do not intend to use it to determine possible Fusarium resistance based on a phylogenetic tree. It is likely that the cultivars cannot be sufficiently differentiated using ITS markers. This is due to their genetic similarity based on breeding history. Chloroplast markers occur mainly on the maternal alleles. Although these two markers are widely used, we felt that multiallelic SSR markers were the most appropriate in our case.
Please see the attachment.
Author Response File: 
Author Response.docx
Reviewer 2 Report
Lines 9-11 are not required in the abstract. the focus should be given to results and a few lines as a conclusive remark
Line 25 use standard units of area
Line 41 correct the sentence
Line 64 provide the source of the collection of these isolates
The cultivars used in the study are too less and authors should make a good pool of cultivars and extend the study.
Authors should also perform the NBT staining
Discussion should be improved
a retest of materials is always suggested in control conditions to rely on results.
Author Response
Thank you for taking the time to read and comment on the paper. Below are the explanations and changes in the order of the comments.
Comments and Suggestions for Authors:
Lines 9-11 are not required in the abstract. the focus should be given to results and a few lines as a conclusive remark
Improved: Abstract: Due to the restricted genetic diversity among current asparagus cultivars, wild relatives are of particular interest as a source of resistance. In this study, seventeen cultivars …
Line 25 use standard units of area
In the original source (FOA) the cultivated area is given in hectares and to avoid confusion we would like to stay with the unit.
Line 41 correct the sentence
Improved: There are some studies on the use of fungicides and biological control agents to control Fusarium crown and root rot, but the results are mostly inconclusive or contradictory, especially in field trials.
Line 64 provide the source of the collection of these isolates
Improved: The original material was isolated in 2011 from asparagus shoots in experimental fields at the JKI in Quedlinburg (Germany).
The cultivars used in the study are too less and authors should make a good pool of cultivars and extend the study.
Other studies and our own findings have shown that resistance is mainly found in wild species (Lewis and Shoemaker 1964, Kathe et al. 2019, Sonoda et al. 2001). For wild species, we selected mainly available genotypes, i.e., those from which we could obtain sufficient seed and which germinated in sufficient numbers. Because of these problems, it is sometimes very difficult to obtain enough seeds for multiple testing. It´s known that asparagus varieties have a narrow genetic base (Knaflewski 1996, Mercati et al. 2015), but we did not want to exclude them from the outset. Therefore, we included older varieties such as RvB, but also modern varieties such as Ramada in the set. However, a larger number of varieties did not seem reasonable to us for the reasons mentioned above.
Authors should also perform the NBT staining
In this study, we aimed only to compare the resistance mechanism of wild species A. densiflorus and A. aethiopicus using the staining method after determining the possible source of resistance. For this purpose, we used a well-established method (Serfling et al. 2016). In other studies, comparison of DAB staining with NBT staining has not revealed significant differences, even though NBT staining localizes ROS more precisely (Javvaji et al. 2020). NBT staining will definitely be included for a more detailed study of the resistance mechanisms of these two wildtype species.
Discussion should be improved
Improved
a retest of materials is always suggested in control conditions to rely on results.
Please see the attachment.
Author Response File: Author Response.docx
Round 2
Reviewer 1 Report
The authors took into account the main comments. The text has become clearer. The article can be accepted
Reviewer 2 Report
The authors have revised the manuscript as per suggestions.
