Abstract
The obligate biotroph oomycete Plasmopara halstedii causes downy mildew on sunflower crop, Helianthus annuus. The breakdown of several Pl resistance genes used in sunflower hybrids over the last 25 years came along with the appearance of new Pl. halstedii isolates showing modified virulence profiles. In oomycetes, two classes of effector proteins, key players of pathogen virulence, are translocated into the host: RXLR and CRN effectors. We identified 54 putative CRN or RXLR effector genes from transcriptomic data and analyzed their genetic diversity in seven Pl. halstedii pathotypes representative of the species variability. Pl. halstedii effector genes were on average more polymorphic at both the nucleic and protein levels than random non-effector genes, suggesting a potential adaptive dynamics of pathogen virulence over the last 25 years. Twenty-two KASP (Competitive Allele Specific PCR) markers designed on polymorphic effector genes were genotyped on 35 isolates belonging to 14 Pl. halstedii pathotypes. Polymorphism analysis based on eight KASP markers aims at proposing a determination key suitable to classify the eight multi-isolate pathotypes into six groups. This is the first report of a molecular marker set able to discriminate Pl. halstedii pathotypes based on the polymorphism of pathogenicity effectors. Compared to phenotypic tests handling living spores used until now to discriminate Pl. halstedii pathotypes, this set of molecular markers constitutes a first step in faster pathotype diagnosis of Pl. halstedii isolates. Hence, emerging sunflower downy mildew isolates could be more rapidly characterized and thus, assessment of plant resistance breakdown under field conditions should be improved.
Highlights
The sunflower (Helianthus annuus) is the fourth most widely grown oil crop in the world after palm, soybean and rapeseed
We use Single Nucleotide Polymorphisms (SNPs) in effector genes to design 22 KASP (Competitive Allele Specific PCR) markers and genotype them in a set of 35 isolates belonging to 14 Pl. halstedii pathotypes
We identified 54 putative effector genes (Fig 1), hereafter named ‘effectors’, with 27 RXLR and 27 CRN listed on the Web page (S1 Table, Plasmopara halstedii effector polymorphism)
Summary
The sunflower (Helianthus annuus) is the fourth most widely grown oil crop in the world after palm, soybean and rapeseed. In contrast to other oil crops, sunflower is environmental friendly, since it grows under low inputs (water, soil fertilizers and fungicides). Downy mildew caused by the oomycete Plasmopara halstedii is one of the major diseases affecting sunflower crop production (for review [1]). Pl. halstedii has been described as a highly selfing species, and a high rate of homozygosity is observed in natural populations [1, 2]. First described in the United States [3], this disease was reported worldwide mostly in areas where sunflowers were largely cultivated. Pl. halstedii infection may severely impact sunflower seed yield through either damping off of seedlings and reduction of plant population size, or ongoing disease symptoms which in turn induce dwarfing and sterility of heads. Since 1992, Pl. halstedii has been submitted to quarantine regulation in the European Union (directive 92/103/CEE)
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