Methods to isolate and maintain clubroot for improved resistance screening and labeling

Term: 3 years, beginning 2024
Status: Ongoing
Researcher(s): Stephen Strelkov, U of A
SaskCanola Investment: $56,925
Total Project Cost: $341,550
Funding Partners: WGRF, ACPC, MCGA

Objective

1. To develop best practices to maintain clubroot isolates in planta to avoid virulence shifts. 

2. To optimize microlaser technology as a fast and efficient way to obtain single-spore isolates for characterization and distribution. 

Project Description

Although planting resistant canola varieties is the primary approach for clubroot management, a growing number of clubroot pathotypes has emerged in recent years that can overcome host resistance, posing a significant challenge for growers. As such, it is critical to identify novel sources of resistance that are effective against these emerging strains of the pathogen. The identification of and breeding for resistance relies on testing host lines by inoculating them with the most prevalent and/or significant clubroot pathotypes on the Prairies. Unfortunately, several factors can hinder clubroot resistance screening. One major obstacle is the inability to grow the pathogen in vitro, which necessitates its maintenance and multiplication on plant hosts. These hosts can cause shifts in the virulence of the pathogen, so that breeders conducting resistance testing in the greenhouse may actually be using the “wrong” pathotypes. Further complicating matters is that the recovery of single-spores isolates of the clubroot pathogen is difficult and time-consuming, and thus “field isolates” often have to be used for screening purposes. These field isolates are usually heterogeneous and may consist of pathotype mixtures, producing inconsistent or fluctuating results. This study aims to improve the effectiveness and reliability of resistance screening and labelling. 

The proposed research stems from an identified need from industry and growers, and builds on the resistance labeling initiative currently underway. The Strelkov Lab at the University of Alberta is producing inoculum (via a limited service agreement with the Canola Council of Canada) of pathotypes 3A, 3D and 3H to share with industry for the first phase of resistance labeling. However, no single-spore isolates are available for pathotypes 3A and 3D, and hence field isolates are being used for this purpose; the scope of the service agreement does not cover costs for single-spore isolation or method development. In addition, as other important and/or emerging pathotypes are identified (such as pathotypes 8E and 9E) there will be a need for more efficient and high-throughput single-spore isolation protocols. Similarly, once inoculum is distributed to different companies or researchers, different approaches to maintain the isolates in planta may cause virulence shifts, so that over time different groups could no longer be working with the same material. This work will address both issues by enabling rapid isolation of many single-spores, while developing uniform and validated best practices for maintaining these isolates in different labs. 

Virulence shifts (or stability) can be monitored via RADseq genome sequencing of the pathogen to ensure pure pathotypes remain pure over time, and/or to further understand how virulence shifts are occurring in the lab. 

Our group is very well positioned to ensure that the solutions stemming from this work are adopted to the benefit of growers and the canola industry in general. Dr. Strelkov is a member of the Clubroot Steering Committee and contributed to development of the Clubroot Labeling Proposal. In addition, the Strelkov Lab is already producing the inoculum for the first phase of this initiative, so we will be able to share results from this work as they become available. The Lab already has Material Transfer Agreements (negotiated following approval from Alberta Agriculture & Irrigation) in place to share inoculum with numerous seed companies and is in the process of getting more agreements ready with others. As such, output from this research (e.g., single-spore isolates) will be shared immediately with stakeholders as it becomes available, expediting the benefits of the work. The information and protocols for improved maintenance of clubroot material in planta, as well as the detailed spore isolation protocols, will also be made available through the Clubroot Steering Committee, at grower and industry meetings, and via publication of reports and refereed papers.