Genomics of Plasmodiophora brassicae

Plasmodiophora brassicae is a soil-borne pathogen that infects roots of plants in Brassicaceae and causes enlarged roots or clubs known as the clubroot disease. Details of its complex life cycle and particularly the molecular basis of its strategies to master defenses and alter metabolism of plant hosts are still largely unknown. The general aim of this thesis was to enhance the genomic knowledge on P. brassicae and its intimate relationship with a plant host. We developed a protocol for extraction of large amounts of high-quality DNA from pathogen resting spores that allowed us to apply long-read PacBio sequencing. De-novo assembly of the P. brassicae e3 strain generated a 25.2 Mb nuclear genome and a mitochondrial genome (114.6 kb). Twenty nuclear contigs were assembled of which 13 from telomere-to-telomere, thanks to the resolution of highly repetitive sequences. As much as 11.5% of the genome was assigned to repetitive sequences, a higher proportion than previously estimated. The most abundant were transposable elements (TEs) such as Copia and Gypsy and unclassified interspersed repeats. They were particularly clustered in telomeric, sub-telomeric and large regions with complex structural variation found on each contig. Among 9,231 predicted protein-coding genes in the nuclear genome, we identified 314 small, secreted proteins as P. brassicae effector candidates. They were distributed along all contigs. TEs and unclassified repeats were found within a 3 kb distance from more than a third of the predicted effectors. We further detected enrichment of the effector candidates with a motif rich in valine, leucine and alanine amino acids. Annotation of the circular mitochondrial genome revealed a compact and complex sequence organization with intron-rich genes, a new splicing mechanism and a previously not resolved 12 kb repetitive region. Our findings and the new genome sequences, currently representing the only P. brassicae long-read assembly, form a valuable resource for comparative and functional analyses