Potential Source of Resistance in Introgressed, Mutant and Synthetic Brassica juncea L. Lines against Diverse Isolates of White Rust Pathogen, Albugo candida
, Pooja Garg 1, Mahesh Rao 1, R. C. Bhattacharya 1, Jameel Akthar 3, Rashmi Yadav 3, Mamta Singh 3, Kartar Singh 3, P. Nallathambi 4, C. Uma Maheswari 4, P. D. Meena 5, Hari Singh Meena 5, P. K. Rai 5, Usha Pant 6, Mohd. Harun 7, Ravish Choudhary 8, Slavica Matic 9 and Ashish Kumar Gupta 1,*
Round 1
Reviewer 1 Report (New Reviewer)
Line 121: “13 prevalent isolates of white rust”. What is meant by prevalent? Are all 13 isolates race 7?
Line 171: Please add the manufacturer name of the Quick-DNA™ Fungal/Bacterial Miniprep Kit.
Line 182-183: Please add name of clean up kit ; add the name of the company that performed the sequencing.
Line 184: Please add blast website: http://blast.ncbi.nlm.nih.gov.
Line 194: Please further explain what gelatin capsules are.
Line 208: add “light” microscope and the microscope manufacturer's name.
Line 285: Most of the sequences deposited in GenBank of A. candida are between 750 and over 800 bp long. Why are your sequences only 542 bp long? Supplement the phylogenetic analysis with A. candida sequences downloaded from GenBank and add an outgroup.
Line 294: “Such variability might be the result of different selection pressure on pathogen due to availability of a host species for …”. It links up with what was said on line 121.
Author Response
Kindly find attached responses to the comments/suggestion
Author Response File: 
Author Response.pdf
Reviewer 2 Report (New Reviewer)
The authors screened the A. candida-resistant line from introgressed, mutant, and synthetic B. juncea lines using 13 field isolates of A. candida. This study showed that resistance varies by isolates and by the tissue to be inoculated. This study provides important information on resistance breeding for A. candida in B. juncea.
Several resistance genes of A. candida have been reported in B. juncea. Therefore, it is necessary to show information on the resistance genes in the materials used.
NJ tree of 13 isolates.
Presumably, the authors used a primer set that has been used in the previous study. If so, the previous study should be cited. Further, NJs should be made with known ITS sequences, especially ITS sequences of A. candida isolated from the genus Brassica and related species.
Author Response
Kindly find attached responses to the comments/suggestion
Author Response File: Author Response.pdf
Round 2
Reviewer 2 Report (New Reviewer)
I do not have further comments. It is ready for acceptance.
This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.
Round 1
Reviewer 1 Report
In this manuscript authors test different accessions of Brassica juncea for their resistance to Albugo candida. Accessions include resynthesized lines, introgression lines and mutated lines. They perform three different types of trials: at the cotyledon stage and true leaf stage in greenhouse under controlled conditions and under field conditions in different locations and years. They tested all the accessions against 13 isolates of the fungus, obtained from different locations.
The main strength of the paper is that resistance test have been carried out in different plant stages and different conditions. This makes that the results be quite robust. Authors have found several promising accessions with resistance at different stages and conditions.
However, two major concerns arise from the paper. Authors want to obtain lines of B. juncea with resistance to multiple isolates. My main concern here is that we do not have information about the isolates employed. Authors describe in the introduction that there are different races. Do they belong to different races?, or if they are from the same race, do they differ in their pathogenesis?. I think that this information is key in interpreting the results.
The other concern is that authors have made field evaluation in multiple locations and years. However, I do not know if the results they present in the tables are the average of the field trials and the years. They first present the data and later describe the ANOVA in the results, which for me it has no sense. They present an analysis of the GE interaction, however the interpretation of this data is poorly developed and is not even described as an objective of the paper. Do genotypes are stable across environments?, which accession is the most stable?.
Reviewer 2 Report
Dear Author
Please find my comments below
Page 3 line 120: from where the 13 prevalent isolates of white rust were obtained?
Page 3 line 123: B. juncea make it italic it’s a scientific name.
Page 3 line 131: The advanced introgressed (RLM 198) and it's mentioned in line 117 as 194? Please clarify.
Page 3 line 133: B. juncea make it italic it’s a scientific name.
Page 3 line 140: what is the total line for this population?
Page 3 line 141: B. juncea make it italic it’s a scientific name.
Page 3 line 147: what is the total line for this population?
Page 4 line 149: B. juncea make it italic it’s a scientific name.
Page 4 line 164: Are the preliminary screening studies for white rust isolate selected base one differential set? Are the isolate tested differential set?
Page 4 line 169: How the isolate has increased after a single pustule? Please describe more with details. How the isolation has been maintained during the increase?
Page 4 line 177: This is a confusing section. Please re write. “inoculum concentration to 2 ×104 zoosporangia mL−1before dilution with sterilized distilled water”
Page 4 line 178: Which one is the first step? Please make it clear for the reader. “The inoculum suspension was then kept at room temperature (20ËšC) for 15 min to release the zoospores after incubation at 13ËšC for 2 to 2.30 h.”
Page 5 line 184: Please put the measurement of the inoculum by using milliliter (ml). “micropipettes (15 ± l droplet size)”
Page 5 line 184: How many replications were used for this experiment?
Page 5 line 185: Where the plant was kept? Greenhouse or chamber? Please clarify. It's confusing for the reader.
“inoculated plants were kept at the ICAR-NIPB glass house facility in New Delhi 185 (Latitude: 28Ëš38′23” N, Longitude: 77Ëš09′27” E, Altitude: 228.61 m above) at 16/15 ± 1Ëš C 186 Day and night temperature, > 90% relative humidity, a 16-h photoperiod, and 8-h of dark- 187 ness. Furthermore, for successful infection and disease development, the entire chamber 188 was covered with a light-coloured tarpaulin sheet 24 h after inoculation.”
Page 5 line 191: Is this a range temp? Please confirm and clarify. “temperatures of 16-18 ËšC and 15ËšC,”
Page 5 line 192: Mention of the result (figure2) in material and method is not acceptable. Please move it to the result in the section.
Page 5 line 193: What disease scale was used for disease severity?
Page 5 line 194:What varieties were used for planting?
Page 5 line 203: This figure shows filed pictures, not a greenhouse as mentioned in line 192.
Page 5 line 217: Which isolate or race of white rust was used in the field?
Page 8 line 303: In table 2, what is the value of R equal to the disease index?
Page 11 line 323: The introgression lines are 194. Why are only 28 lines only showed in this table?
Page 13 line 349: B. juncea make it italic it’s a scientific name.
Page 20 line 507: Further molecular analysis is needed to confirm these sources of resistance are novel sources of resistance.
