Generating a rapid and low-cost diagnosis of fungi on wheat

Posted on 14.02.2022 | Last Modified 18.03.2024
Lead Researcher (PI): Sean Walkowiak
Institution: Canadian Grain Commission
Total WGRF Funding: $133,550
Co-Funders: Manitoba Crop Alliance, Saskatchewan Wheat Development Commission
Start Date: 2021
Project Length: 3 Years

Construct a 'reference panel' of Fusarium and rust fungi. Optimize sample preparation and generate MALDI-TOF profiles for the 'reference panel'. Detailed profiling of the 'reference panel' using genetics, genomics, and analytical chemistry.

Project Summary:

The project was successful at completing its objective in developing novel Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) methods for Fusarium and rust identification. The work was conducted by technical staff at the Canadian Grain Commission and a graduate student at the University of Manitoba, with contributions from five undergraduate trainees in term appointments (Appendix 1), and samples from Agriculture and Agri-Food Canada and The University of British Columbia. We constructed a ‘reference panel’ of Fusarium and rust isolates that are of known species, chemotypes, or races that have been provided by expert pathologists from across Canada – many of these strains are widely used in research in Canada or in disease nurseries used for varietal registration. We optimized methods for sample preparation and achieved reproducible MALDI-TOF MS fingerprints/profiles for the ‘reference panel’ and completed the construction of MALDI-TOF MS reference libraries. We collected samples for the ‘validation panel’, which were treated as ‘unknown’ samples and compared to the MALDI-TOF MS reference libraries for identification. The results support that the MALDI-TOF MS method and reference libraries are able to properly distinguish species of Fusarium and rust. To ensure the success of the project, we added to the Fusarium ‘validation panel’ an additional harvest year and an additional panel of recent isolates from oat and barley containing more diverse species, effectively tripling the originally proposed size of the ‘validation panel’. Due to the low cost of the MALDI-TOF MS method, this did not significantly impact our operational costs. For the Fusarium testing, true species identity was confirmed by DNA-based testing methods. The analysis of the Fusarium samples of the ‘validation panel’, which originated from the 2021-2023 harvests, also revealed interesting surveillance information on the trends of Fusarium species prevalence in response to the 2021 drought conditions, where there was increased Fusarium species diversity observed on harvested wheat in the drought year of 2021. The main conclusion of the study is that MALDI-TOF MS can identify Fusarium and rust species that are impacting Canadian cereals, and can be used to complement existing microbial identification tools such as morphology analyses and DNA-based testing.