Abstract
The Ascomycete fungus Phaeoacremonium minimum is one of the primary causal agents of Esca, a widespread and damaging grapevine trunk disease. Variation in virulence among Pm. minimum isolates has been reported, but the underlying genetic basis of the phenotypic variability remains unknown. The goal of this study was to characterize intraspecific genetic diversity and explore its potential impact on virulence functions associated with secondary metabolism, cellular transport, and cell wall decomposition. We generated a chromosome-scale genome assembly, using single molecule real-time sequencing, and resequenced the genomes and transcriptomes of multiple isolates to identify sequence and structural polymorphisms. Numerous insertion and deletion events were found for a total of about 1 Mbp in each isolate. Structural variation in this extremely gene dense genome frequently caused presence/absence polymorphisms of multiple adjacent genes, mostly belonging to biosynthetic clusters associated with secondary metabolism. Because of the observed intraspecific diversity in gene content due to structural variation we concluded that a transcriptome reference developed from a single isolate is insufficient to represent the virulence factor repertoire of the species. We therefore compiled a pan-transcriptome reference of Pm. minimum comprising a non-redundant set of 15,245 protein-coding sequences. Using naturally infected field samples expressing Esca symptoms, we demonstrated that mapping of meta-transcriptomics data on a multi-species reference that included the Pm. minimum pan-transcriptome allows the profiling of an expanded set of virulence factors, including variable genes associated with secondary metabolism and cellular transport.
Highlights
Grapevine trunk diseases (Esca, and Botryosphaeria, Eutypa, and Phomopsis-diebacks) are a significant threat to viticulture worldwide (Gramaje et al, 2018)
We identified a deletion in UCR-PA7, Pm448, and Pm449 that included five adjacent genes all belonging to BGC_22, which is potentially involved in polyketide synthesis (PKS) (Figure 3C)
We identified a total of 455 complete proteincoding sequences (CDS) encoding complete proteins that were not present in the Pm. minimum isolate 1119 (Pm1119) reference: 11 of these were shared by two isolates, whereas 195, 98, and 151 were found only in UCR-PA7, Pm1118, and Pm449, respectively (Supplementary Data S3 and Supplementary Data S1: Figure S9)
Summary
Grapevine trunk diseases (Esca, and Botryosphaeria-, Eutypa-, and Phomopsis-diebacks) are a significant threat to viticulture worldwide (Gramaje et al, 2018). Previous analyses of a draft genome assembly of Pm. minimum provided a glimpse of the large number and broad diversity of genes involved in secondary metabolism and cell wall degradation (Blanco-Ulate et al, 2013; Morales-Cruz et al, 2015). Gene families of these putative virulence factors have undergone distinctive patterns of expansion and contraction in Pm. minimum and another Esca pathogen Ph. chlamydospora, relative to the genomes of other trunk pathogens, which may explain the differences between Esca symptoms and those of the dieback-type trunk diseases (Morales-Cruz et al, 2015). We incorporated all core and variable transcripts into a pan-transcriptome, which provided a more comprehensive representation of the virulence repertoire of the species when used as reference for meta-transcriptomic analysis of naturally occurring Pm. minimum infections
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