Developing technologies and resources to gain an accurate view of Canadian populations of Plasmodiophora brassicae
Term: 4 years
Status: Ongoing
Researcher(s): Fengqun Yu, AAFC
SaskOilseeds Investment: $183,330
Total Project Cost: $499,990
Funding Partners: ADF
Objective
Improve efficiency for producing near pure genotype isolates (NPGIs) to facilitate field surveys of pathogen race structures.
Collect clubroot gall samples in canola fields in Alberta, Saskatchewan and Manitoba annually.
Produce NPGIs from the galls collected in the canola fields in Western Canada.
Determine the frequency each Avirulence (Avr) gene and race structure of P. brassicae populations.
Perform studies on the pathogen’s genetic diversity and population structure through whole genome DNA sequencing.
Identify differential isolates that can accurately phenotype breeding lines carrying respective CR genes.
Project Description
Clubroot disease continues to spread on the Canadian prairies. Use of resistant cultivars combined with crop rotation is the only economical method to reduce production losses caused by the disease. A prerequisite of deployment of resistant cultivars for durable resistance is to determine if the corresponding Avr genes are present in the pathogen populations or not.
P. brassicae exists as mixtures of races in the soil, which can cause inaccuracy on resistance labeling and profiling race structure of the populations if the field collected clubroot galls are used as inoculum for experiments. Many attempts have been made to produce single spore derived isolates to obtain pure genotypes of P. brassicae. However, the practicability of the methods is poor and their infection rates are very low. To overcome the issues, we have developed a method to produce near-pure genotype isolates (NPGI) through isolation of resting spores from single root cells. Our data analyses show that almost no heterozygous alleles could be identified in the NPGIs compared to the high level of heterozygosity in the field-collected galls, indicating this method is reliable for producing NPGIs. We have achieved a success rate of almost 100% producing NPGIs with the commercial soilless mix Sunshine #3. However, Sunshine Mix #3 has been discontinued and substituted with other Sunshine Mixes. Unfortunately, the success rate was much lower (only 16% in average) with the substitute Mixes compared with Mix #3 (100%). Further studies are needed to find an alternative commercial soilless mix or develop a custom mix or a more efficient potting method to boost success so this method can be used for routine field monitoring aimed at determining changes in race structure of the pathogen populations, and other applications in science.
Several researchers classified P. brassicae into various pathotypes using differential reactions on Brassica cultivars such as Williams Differentials, European Clubroot Differential set and Canadian Clubroot Differential (CCD) set. Most of the differential cultivars used by the researchers are non-canola crops, belonging to vegetable or fodder Brassicas with little genetic information. In addition, the pathotypes defined by the differential sets do not reflect the variation of specific avirulence (Avr) genes in P. brassicae populations on canola, but rather reflect the biological outcome of the host/pathogen interaction with little or no information about the genetic factors conditioning the responses. Over the past ten years, with support from ADF, SaskCanola, WGRF and breeding companies, we have developed the first set of spring-type B. napus lines containing seven single clubroot resistance genes Rcr1, Rcr3, Rcr5, Rcr8, Rcr9, Rcr10 and Rcr11. Theoretically, this set of breeding lines has the ability to define 2 = 128 races.
This more informative system can show which resistance genes are currently effective against the pathogen, identify isolates which can discriminate among different resistance genes in registered cultivars, and provide the opportunity to deploy strategies for resistance that should be more durable for the future.
Plants and pathogens co-evolve through interactions with each other. This co-evolution is fueled by DNA variation underlying the recognition of pathogen proteins by the host and the defeat of host defenses by the pathogen. Adaptation is fueled by allelic diversity, sexual recombination, and, together with new mutations, leads to an on-going ebb-and-flow of Avr and R genes and their interactions. Genetic diversity in populations of both the host and pathogen represent a pool of possible variants to maintain adaptation via natural selection. Knowledge of the population genetic structure of the pathogen may offer insight into the best breeding strategy for durable resistance. However, genetic diversity of P. brassicae populations in western Canada has not been investigated so far.
Currently, canola breeding companies desperately need a set of pure genotype isolates that can consistently differentiate disease reactions on various clubroot resistance genes with reproducibility. With several hundreds of NPGIs produced from this project, we will be able to obtain a set of isolates that can accurate phenotype respective CR genes for their breeding programs.