Transcriptional adaptation facilitates host specialization in a wheat pathogen

Host-pathogen co-evolutionary dynamics force microbial plant pathogens to constantly develop and adjust specific adaptations to thrive in their plant host, and therefore also act as strong drivers of divergence and speciation in pathogens. Factors that confer host specialization and determine host specificity are very diverse and range from molecular and morphological strategies to metabolic and reproductive adaptations. Identification of these key factors is a major goal in the study of pathogen evolution and may aid the development of sustainable crops and crop protection strategies. We here took a novel experimental approach and conducted comparative microscopy and transcriptome analyses of the closely related, recently diverged fungal pathogens Zymoseptoria tritici, Z. pseudotritici, and Z. ardabiliae that establish compatible and incompatible interactions with wheat. Although infections of the incompatible species induce plant defense response during invasion of stomatal openings, we found a highly conserved early-infection program among the three species. The transcriptional programs of the three pathogens are conserved to a large extent, as only 9.2% of the 8,885 orthologous genes are significantly differentially expressed during initial infection of wheat. The genes up-regulated in the compatible pathogen reflect adaptation to growth in wheat tissue e.g., by re-programming of fungal metabolism. In contrast, genes primarily involved in counteracting cell stress and damage are strongly induced in the incompatible species. Based on the species-specific gene expression profiles, we further identified nine candidate genes encoding putative effectors and host-specificity determinants in Z. tritici. These effectors are strongly induced in the compatible species and may interfere with host immune suppression. We also identify putative necrotrophic effectors which are induced at the onset of necrotrophic growth. Together, the results presented here indicate that host specialization has involved transcriptional adaptation of a relatively small number of genes. Our findings demonstrate the potential comparative analyses of compatible and incompatible infections present for identifying traits involved in pathogen evolution and host specialization. High-quality total RNA from wheat leaves inoculated with Zp13, and Za17 was isolated. Strand-specific RNA-seq libraries including polyA enrichment were prepared and sequenced using an Illumina HiSeq 2500 platform. We further used published in planta and in vitro RNA-seq data (Grandaubert et al., 2015; Haueisen et al., 2019b; Möller et al., 2019). Differential gene expression analyses for 8,885 orthologous Zymoseptoria spp. genes (Feurtey et al., 2020) were performed in R (R Core Team, 2017) using DESeq2 v1.28.1 (Love et al., 2014). Genes with Padj ≤ 0.01 and |log2 fold-change | ≥ 2 were considered significantly differentially expressed.