The genome assembly of the fungal pathogen Pyrenochaeta lycopersici from Single-Molecule Real-Time sequencing sheds new light on its biological complexity

Abstract

The first draft genome sequencing of the non-model fungal pathogen Pyrenochaeta lycopersici showed an expansion of gene families associated with heterokaryon incompatibility and lacking of mating-type genes, providing insights into the genetic basis of this “imperfect” fungus which lost the ability to produce the sexual stage. However, due to the Illumina short-read technology, the draft genome was too fragmented to allow a comprehensive characterization of the genome, especially of the repetitive sequence fraction. In this work, the sequencing of another P. lycopersici isolate using long-read Single Molecule Real-Time sequencing technology was performed with the aim of obtaining a gapless genome. Indeed, a gapless genome assembly of 62.7 Mb was obtained, with a fraction of repetitive sequences representing 30% of the total bases. The gene content of the two P. lycopersici isolates was very similar, and the large difference in genome size (about 8 Mb) might be attributable to the high fraction of repetitive sequences detected for the new sequenced isolate. The role of repetitive elements, including transposable elements, in modulating virulence effectors is well established in fungal plant pathogens. Moreover, transposable elements are of fundamental importance in creating and re-modelling genes, especially in imperfect fungi. Their abundance in P. lycopersici, together with the large expansion of heterokaryon incompatibility genes in both sequenced isolates, suggest the presence of possible mechanisms alternative to gene re-assorting mediated by sexual recombination. A quite large fraction (~9%) of repetitive elements in P. lycopersici, has no homology with known classes, strengthening this hypothesis. The availability of a gapless genome of P. lycopersici allowed the in-depth analysis of its genome content, by annotating functional genes and TEs. This goal will be an important resource for shedding light on the evolution of the reproductive and pathogenic behaviour of this soilborne pathogen and the onset of a possible speciation within this species.

Long-read only assembly of Drechmeria coniospora genomes reveals widespread chromosome plasticity and illustrates the limitations of current nanopore methods

“…On the basis of our results, a re-analysis of the completeness of these “nanopore-only” genomes is merited, to confirm that they are indeed of low quality. Similar concerns do not apply to fungal genomes assembled using only long reads generated with Pacific Biosciences technology [ 23 ] because these are not hindered by the intrinsic problem of homopolymer length errors that we found to be the most significant quality barrier when using ONT reads. On the basis of our detailed analysis and in line with the consensus regarding de novo assembly with ONT long reads (e.g., [ 24 ]), we polished our 3 assemblies with short reads.…”

Long-read only assembly of Drechmeria coniospora genomes reveals widespread chromosome plasticity and illustrates the limitations of current nanopore methods

“…This is problematic as low-quality genome sequences compromise confirm that they are indeed low quality. Similar concerns do not apply to fungal genomes assembled using only long reads generated with Pacific Bioscience technology[23] as these do not suffer from the intrinsic problem of homopolymer length errors that we found to be the most significant quality barrier when using ONT reads.On the basis of our detailed analysis and in line with the consensus regarding de novo assembly with ONT long reads (e.g [24]…”

Host-specific subtelomere: Genomic architecture of pathogen emergence in asexual filamentous fungi

“…Chromosome 12 was flanked on either side by nearly complementary sequences of 12,772 and 11,195 bp; an alignment of the sequences in opposite orientation contained two sections of similar sequences (at 98.14% identity) that were separated by a section of different sequences of 1,926 and 341 bp (at overall 83% identity over the three sections). This subtelomeric element is called a 11-kb element, based on the number of exact base matches (10,653) in the alignment. And chromosome 11 was similarly associated with 18-kb complementary sequences at 99.87% complementarity.…”

“…Heterokaryon incompatibility involves a protein partner with a HET domain as a trigger of programmed cell death (Paoletti and Clavé 2007). A large number of HET domain genes were found in the genomes of asexual filamentous fungal pathogens, for example, 231 and 324 in two isolates of Pyrenochaeta lycopersici (Dal Molin et al 2018). HET domain genes possess some characteristics of pathogenicity islands: they are highly variable between closely related isolates from different incompatibility groups, but they display trans-species polymorphism, where a HET domain gene from an isolate is more similar to one from an isolate of a different species than to ones from isolates of the same species (Muirhead et al 2002).…”