Niradha Withana Gamage

Canola cultivars carrying the blackleg resistance genes Rlm2 and Rlm7 were released in Western Canada in 2022. To determine the origin of virulence on these genes, we phenotyped and sequenced 21 Leptosphaeria maculans isolates collected before deployment (2010–2015) and seven isolates recovered from Rlm2 and Rlm7 plots. Three AvrLm2 variants differed only at residues 133 and 146: 13 of 14 isolates carrying the reference Gly133/Glu146 allele were avirulent on Rlm2, whereas 10 of 14 carrying His133/Gln146, Asn133/Gln146 or an allele inactivated by repeat-induced point mutation (RIP) were virulent or intermediate. Eight isolates carried RIP-inactivated AvrLm4-7 alleles sharing a premature stop at codon 35; seven were virulent or intermediate on Rlm7, and five were avirulent on Rlm3, consistent with masking of AvrLm3 by AvrLm4-7. Virulence alleles at both loci predated deployment: both post-pressure Rlm7 isolates carried an AvrLm4-7 allele identical to that of a 2015 isolate, and the His133/Gln146 allele of two post-pressure Rlm2 isolates occurred in four pre-pressure lineages. A core-genome phylogeny based on 39,012 SNPs placed the post-pressure isolates in at least five lineages of a single regional clade. Rlm2 and Rlm7 deployment is therefore selecting on standing variation that can be monitored before resistance genes are released.

Fusarium head blight is a limitation to grain production and can be caused by several different Fusarium species. We evaluated the ability of matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) to perform species identifications. The method generates a unique peptide mass fingerprint (PMF) for each sample that can be matched to a reference library. We first created a reference library of PMF profiles for Fusarium species from across Canada. Then, we tested the library to perform identifications using two validation panels. The first panel consisted of 820 fungal isolates from wheat (2021–2023 harvest years) and the second was 74 fungal isolates from oat and barley (2022 harvest year). The species identity of samples from the validation panels was confirmed with high-throughput quantitative PCR using species-specific DNA markers. The first validation panel was mostly F. graminearum and there was 95% overlap between the MALDI-TOF MS and DNA-based identifications. The second panel was mostly F. poae and the identifications from the two methods had 86% overlap. Our findings indicate that MALDI-TOF MS biotyping is sensitive enough to identify Fusarium strains to their species complexes and certain Fusarium strains to the species level.