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Developing canola germplasms with multiple resistance mechanisms for durable clubroot control

Posted on 06.02.2017 | Last Modified 07.05.2019
Lead Researcher (PI): Peng, Gary
Institution: Agriculture & Agri-Food Canada
Total WGRF Funding: $169,143
Co-Funders: Saskatchewan Canola Development Commission
Start Date: 2015
Project Length: 3 Years
Objectives:

To identify club-root resistance (CR) genes effective against newly identified clubroot pathotypes. To characterize and differentiate resistance mechanisms for selected CR lines/genes and to incorporate effective and versatile CR genes into canola breeding lines.

Project Summary:

Cultivar resistance is considered the most effective and practical approach for clubroot management. However, current resistant canola cultivars, available in Canada since 2009, were based on a single clubroot resistance (CR) gene. In 2013, some fields in Alberta with a resistant canola cultivar showed substantial levels of clubroot damage, with further testing showing that the pathogen strains from fields were virulent to each of the clubroot resistant cultivars in the marketplace, including a new pathotype 5x. In 2014, researchers initiated a four-year project to assess if any of the previously identified CR genes would show efficacy against pathotype 5x of P. brassica, to explore advanced technologies for better understanding CR mechanisms and to develop more diverse CR germplasm for sustainable clubroot resistance. As a result of the project, researchers have successfully developed a range of new and improved canola germplasms carrying single/multi CR genes against the new strain of P. brassicae pathotypes 5x.

Cultivar resistance is considered the most effective and practical approach for clubroot management, with resistant canola cultivars available in Canada since 2009. However, all of these cultivars are based on a single clubroot resistance (CR) gene, which can be eroded when the pathogen changes in virulence. In 2013, some fields in Alberta with a resistant canola cultivar showed substantial levels of clubroot damage. Further testing showed that the pathogen strains from these fields were virulent to each of the clubroot resistant cultivars in the marketplace, indicating a potential breakdown of clubroot resistance by new pathogen strains referred tentatively as pathotype 5x.

Researchers at AAFC had previously screened a large number brassica accessions for resistance against the pathotype 3 of (3H in the new system based on Canadian Clubroot Differentials) Plasmodiophora brassicae, the most common strain on the prairies, and identified more than 20 resistant candidates. Some of them were even resistant to all pathotypes (2, 3, 5, 6, 8) found in Canada previously. However, there seemed to be further variations among different populations of the new pathotype 5x that showed different virulence toward resistant canola cultivars. It was not clear if any of the CR genes identified would be effective against all pathotype 5x populations.

In 2014, researchers initiated a new four-year project, with the goal to assess if any of the previously identified CR genes would show efficacy against pathotype 5x of P. brassica. Researchers also wanted to explore molecular and biochemical tools that can be used for studying CR mechanisms, and to develop canola germplasm carrying more diverse CR genes for sustainable clubroot resistance.

The initial assessment of CR candidates against pathotype 5x of P. brassicae began with screening of 24 resistant candidates (to pathotype 3) against mixed and single populations of pathotype 5x from Alberta fields where resistant cultivars failed in 2013. Several candidates were resistant to mixed 5x populations, and the CR genes also showed resistance to all other Pb pathotypes found earlier in Canada (pathotype 2, 3, 5, 6, 8). The work at University of Alberta showed that pathotype 5x is a mixed group of races not separable with the current host differentials.

Researchers then explored advanced technologies to better understand the resistance mechanisms associated with specific CR genes, including transcriptome (RNA-seq), proteome, metabolome (performed at the McGill University), fluorescent microscopy and synchrotron-based chemometrics. These technologies were used to identify metabolic or signaling pathways relating to clubroot resistance, potentially differentiating the modes of action among different CR genes and guiding CR-gene deployment judiciously to enhance the resistance durability. The results also provided the first report on proteomics on clubroot resistance, which provides insights into the resistance conferred by the CR gene Rcr1. The results were published in Frontiers in Plant Science (Song et al. 2016). The final project step was the development of germplasms carrying different CR genes.

Overall, researchers successfully developed a range of new and improved canola germplasms carrying single/multi CR genes against the new strain of P. brassicae pathotypes 5x. Several stable populations of clubroot resistant B. napus germplasm carrying a single CR gene, including the CR genes Rcr1, Rcr2, Rcr3 and Rcr4, have been transferred to seven breeding companies for the production of new resistant canola hybrids. The project has also generated new materials, especially two resynthesized species and two B. napus breeding lines carrying double CR genes, for further studies on resistance mechanisms and durability in new projects.

Resistance to clubroot caused by pathotype 5x with newly developed canola germplasm

 

B. napus germplasm carrying double CR genes (A3, A8) has been produced through the collaboration with Crop Production Services. Two of the double CR-gene lines consistently showed a moderate level of resistance to the most virulent pathotype 5x populations. Their efficacy against other pathotype variants and potential for durable disease resistance is being assessed. Researchers also developed a stable elite line of B. carinata carrying the CR gene Rcr6 that can be used for commercial variety development. Genetic markers have been validated for marker-assisted selection. Elite AAFC B. napus lines (black- and yellow-seeded) carrying the CR gene Rcr1or Rcr3 have been produced and field testing has been carried out with the black-seeded lines. These CR genes can be pyramided in commercial breeding lines for canola hybrids with resistance against a broader range of pathotypes.

Scientific publications.
Lahlali1 R, Song T, Chu M, Yu F, Kumar S, Karunakaran C, Peng G. 2017. Evaluating changes in cell-wall components associated with clubroot resistance using fourier transform infrared spectroscopy and RT-PCR. Int. J Mol. Sci. 18(10), 2058.
Song T, Chu M, Lahlali R, Yu F, Peng G. 2016a. Shotgun label-free proteomic analysis of clubroot (Plasmodiophora brassicae) resistance conferred by the gene Rcr1 in Brassica rapa. Frontiers in Plant Science 7: 1013.
Song T, Chu M, Lahlali R, Karunakaran C, Yu F, Peng G. 2016b. Using “omics” approaches to decipher resistance mechanisms of Rcr1. In: Proc. Brassica 2016.
Oct. 3 – 6.
Yu F, Zhang XG, Huang Z, Chun MG, Song T, Chang A, Deora A, Chen Q, Zhang Y, McGregor L, Falk KC, Gossen BD, McDonald MR, Jing MY, Peng G. 2016. Identification of SNP markers associated with clubroot resistance gene Rcr1 through bulked segregant RNA sequencing. PLoS One 11(4): e0153218.
Yu F, Zhang XG, Peng G, Falk KC, Strelkov SE, Gossen BD. 2017. Genotyping-by-sequencing reveals three QTL for clubroot resistance to six pathotypes of Plasmodiophora brassicae in Brassica rapa. Scientific Reports 7: 4516.
Zhen Huang, Gary Peng, Xunjia Liu, Abhinandan Deora, Kevin Falk, Bruce D. Gossen, Mary Ruth McDonald, Fengqun Yu. 2017. Fine mapping of a clubroot resistance gene in Chinese cabbage using SNP markers identified from bulked segregant RNA sequencing. Frontiers in Plant Sci 8: 1448.