Dissection of Quantitative Blackleg Resistance Reveals Novel Variants of Resistance Gene Rlm9 in Elite Brassica napus

Blackleg is one of the major fungal diseases in oilseed rape/canola worldwide. Most commercial cultivars carry R gene-mediated qualitative resistances that confer a high level of race-specific protection against Leptosphaeria maculans, the causal fungus of blackleg disease. However, monogenic resistances of this kind can potentially be rapidly overcome by mutations in the pathogen’s avirulence genes. To counteract pathogen adaptation in this evolutionary arms race, there is a tremendous demand for quantitative background resistance to enhance durability and efficacy of blackleg resistance in oilseed rape. In this study, we characterized genomic regions contributing to quantitative L. maculans resistance by genome-wide association studies in a multiparental mapping population derived from six parental elite varieties exhibiting quantitative resistance, which were all crossed to one common susceptible parental elite variety. Resistance was screened using a fungal isolate with no corresponding avirulence (AvrLm) to major R genes present in the parents of the mapping population. Genome-wide association studies revealed eight significantly associated quantitative trait loci (QTL) on chromosomes A07 and A09, with small effects explaining 3–6% of the phenotypic variance. Unexpectedly, the qualitative blackleg resistance gene Rlm9 was found to be located within a resistance-associated haploblock on chromosome A07. Furthermore, long-range sequence data spanning this haploblock revealed high levels of single-nucleotide and structural variants within the Rlm9 coding sequence among the parents of the mapping population. The results suggest that novel variants of Rlm9 could play a previously unknown role in expression of quantitative disease resistance in oilseed rape.

Validating the Strategic Deployment of Blackleg Resistance Gene Groups in Commercial Canola Fields on the Canadian Prairies

Blackleg, caused by the fungal pathogen Leptosphaeria maculans, is a serious threat to canola (Brassica napus L.) production in western Canada. Crop scouting and extended crop rotation, along with the use of effective genetic resistance, have been key management practices available to mitigate the impact of the disease. In recent years, new pathogen races have reduced the effectiveness of some of the resistant cultivars deployed. Strategic deployment and rotation of major resistance (R) genes in cultivars have been used in France and Australia to help increase the longevity of blackleg resistance. Canada also introduced a grouping system in 2017 to identify blackleg R genes in canola cultivars. The main objective of this study was to examine and validate the concept of R gene deployment through monitoring the avirulence (Avr) profile of L. maculans population and disease levels in commercial canola fields within the Canadian prairies. Blackleg disease incidence and severity was collected from 146 cultivars from 53 sites across Manitoba, Saskatchewan, and Alberta in 2018 and 2019, and the results varied significantly between gene groups, which is likely influenced by the pathogen population. Isolates collected from spring and fall stubble residues were examined for the presence of Avr alleles AvrLm1, AvrLm2, AvrLm3, AvrLm4, AvrLm5, AvrLm6, AvrLm7, AvrLm9, AvrLm10, AvrLm11, AvrLepR1, AvrLepR2, AvrLep3, and AvrLmS using a set of differential host genotypes carrying known resistance genes or PCR-based markers. The Simpson’s evenness index was very low, due to two dominant L. maculans races (AvrLm2-4-5-6-7-10-11 and AvrLm2-5-6-7-10-11) representing 49% of the population, but diversity of the population was high from the 35 L. maculans races isolated in Manitoba. AvrLm6 and AvrLm11 were found in all 254 L. maculans isolates collected in Manitoba. Knowledge of the blackleg disease levels in relation to the R genes deployed, along with the L. maculans Avr profile, helps to measure the effectiveness of genetic resistance.

Status and advances in mining for blackleg (Leptosphaeria maculans) quantitative resistance (QR) in oilseed rape (Brassica napus)

Key message Quantitative resistance (QR) loci discovered through genetic and genomic analyses are abundant in the Brassica napus genome, providing an opportunity for their utilization in enhancing blackleg resistance. Abstract Quantitative resistance (QR) has long been utilized to manage blackleg in Brassica napus (canola, oilseed rape), even before major resistance genes (R-genes) were extensively explored in breeding programmes. In contrast to R-gene-mediated qualitative resistance, QR reduces blackleg symptoms rather than completely eliminating the disease. As a polygenic trait, QR is controlled by numerous genes with modest effects, which exerts less pressure on the pathogen to evolve; hence, its effectiveness is more durable compared to R-gene-mediated resistance. Furthermore, combining QR with major R-genes has been shown to enhance resistance against diseases in important crops, including oilseed rape. For these reasons, there has been a renewed interest among breeders in utilizing QR in crop improvement. However, the mechanisms governing QR are largely unknown, limiting its deployment. Advances in genomics are facilitating the dissection of the genetic and molecular underpinnings of QR, resulting in the discovery of several loci and genes that can be potentially deployed to enhance blackleg resistance. Here, we summarize the efforts undertaken to identify blackleg QR loci in oilseed rape using linkage and association analysis. We update the knowledge on the possible mechanisms governing QR and the advances in searching for the underlying genes. Lastly, we lay out strategies to accelerate the genetic improvement of blackleg QR in oilseed rape using improved phenotyping approaches and genomic prediction tools.

Recent Findings Unravel Genes and Genetic Factors Underlying Leptosphaeria maculans Resistance in Brassica napus and Its Relatives

Among the Brassica oilseeds, canola (Brassica napus) is the most economically significant globally. However, its production can be limited by blackleg disease, caused by the fungal pathogen Lepstosphaeria maculans. The deployment of resistance genes has been implemented as one of the key strategies to manage the disease. Genetic resistance against blackleg comes in two forms: qualitative resistance, controlled by a single, major resistance gene (R gene), and quantitative resistance (QR), controlled by numerous, small effect loci. R-gene-mediated blackleg resistance has been extensively studied, wherein several genomic regions harbouring R genes against L. maculans have been identified and three of these genes were cloned. These studies advance our understanding of the mechanism of R gene and pathogen avirulence (Avr) gene interaction. Notably, these studies revealed a more complex interaction than originally thought. Advances in genomics help unravel these complexities, providing insights into the genes and genetic factors towards improving blackleg resistance. Here, we aim to discuss the existing R-gene-mediated resistance, make a summary of candidate R genes against the disease, and emphasise the role of players involved in the pathogenicity and resistance. The comprehensive result will allow breeders to improve resistance to L. maculans, thereby increasing yield.

In-silico identification and differential expression of putative disease resistance-related genes within the collinear region of Brassica napus blackleg resistance locus LepR2’ in Brassica oleracea

Blackleg disease, caused by Leptosphaeria maculans, greatly affects the production of cabbage (Brassica oleracea). However, definitive R-gene(s) are yet to be identified in this crop. In contrast, a number of R-loci have been identified in A- or B-genome crops. Identification of few resistant cabbage genotypes indicates the presence of R-genes in this C-genome crop. High ancestral synteny between Brassica genomes suggests that the collinear regions of known A- or B-genome R-loci may also contain functional R-genes in the C-genome. Strong resistance was observed in the cotyledons of cabbage inbred line SCNU-98 against two L. maculans isolates, 03–02 s and 00–100 s. We investigated the collinear region of the Brassica napus blackleg resistance locus LepR2’ in B. oleracea since both isolates of L. maculans contain corresponding avirulence genes. The locus was collinear to a 5.8 Mbp genomic segment of B. oleracea chromosome C09 containing 13 genes that have putative disease resistance-related domains. High expression of genes Bo9g117290 and Bo9g111510 against isolate 00–100 s, and high expression of genes Bo9g126150 and Bo9g111490 against both isolates in the resistant-line SCNU-98 indicate their putative roles in blackleg resistance, which remained to be functionally verified. This work enhances our understanding of R-gene-mediated resistance to blackleg in cabbage.

Assessing progress in breeding to improve grain yield, quality and blackleg (Leptosphaeria maculans) resistance in selected Australian canola cultivars (1978–2012)

Canola breeding in Australia began in the early 1970s with the first cultivars being released in the late 1970s. Thirty-four non-herbicide-tolerant canola cultivars, released in Australia between 1978 and 2012, were evaluated for improvements in yield, quality, blackleg resistance and adaptation to Australian environments. The cultivars were sown at three sites in 2008 and one site in 2014. In addition, blackleg susceptibility was assessed in two independent blackleg experiments in 2008. Yield improvement averaged 21.8 kg ha–1 year–1 (1.25% year–1) but ranged from 8 to 39.1 kg ha–1 year–1 at the lowest to the highest yielding sites, respectively. Although the yield gain shown by our study was for conventional canola only, the different herbicide-tolerant types are derived by incorporating the herbicide tolerance genes into Australian germplasm and so the rate of genetic gain would be expected to be similar for all herbicide tolerance types. Oil and protein concentrations have increased by 0.09% year–1 and 0.05% year–1, respectively, whereas glucosinolate concentration was reduced to between 7 and 16 μmoles per gram of meal by the mid-1990s. Cultivars released before 2002 all had low to moderate resistance to the blackleg isolates present in the fields during the experimental period but more recent releases had improved survival under heavy blackleg pressure due to the incorporation of additional or different resistance genes. The data suggests that at least 25% of the yield improvement achieved by the breeding programs over 30 years was associated with improved blackleg resistance and the remainder with gains in other aspects of potential grain yield. The private breeding companies in Australia will need to continue to produce cultivars with high yield potential and deploy blackleg resistance genes wisely in order to maintain the yield improvements required to remain competitive in global markets.