A Mutagenomic Dissection of Virulence in the Fungal Wheat Pathogen Zymoseptoria tritici

Zymoseptoria tritici is the causal agent of Septoria tritici leaf blotch (STB), an economically significant disease that can reduce wheat (Triticum aestivum) yields. Due to this fungus' in vitro growth capabilities, functional genomics studies can isolate morphogenic switching genes that play a crucial role in fungal virulence. Depending on in vitro nutrient availability and temperature, Z. tritici can grow and switch between a “yeast-like” budding form or a hyphal form. As no specialised infection structures have been identified in Z. tritici infection, the genes involved in morphogenic switching and successful hyphal extension are essential for virulence, enabling leaf penetration through stomatal openings or wounds. “Mutagenomics” is a combination approach of mutagenesis, phenotypic screening, and whole-genome re-sequencing. Advancements in genomic sequencing technologies have eased the process of identifying candidate virulence and morphogenic switching associated genes in Z. tritici.
The mutagenomic study described herein identified strains impaired in full virulence through in planta and in vitro screening of two independent mutant libraries, one generated by restriction enzyme-mediated integration and the other by Agrobacterium-mediated T-DNA mutagenesis. From the nineteen re-sequenced mutant strains, a list of nearly 100 genes affected either by plasmid/T-DNA integration or untagged variation have been identified and prioritised using a combination of approaches and datasets. Functional complementation assays on three candidate genes resulted in restored hyphal growth and restoration of full virulence. These genes include one involved in the purine biosynthesis pathway, a mitogen-activated protein kinase kinase kinase (MAPKKK) which functions upstream of HOG1 MAPK, and a predicted cell cycle control protein involved in pre-mRNA splicing. The data presented support the use of mutagenomics approaches to identify new genes implicated in fungal plant pathogen virulence and identify new Achilles heels in the infection biology of Z. tritici.