Cross-kingdom small RNA (sRNA) silencing has recently emerged as a mechanism facilitating fungal colonisation and disease development in plants. Here we characterise RNAi pathways in Zymoseptoria tritici, a major fungal pathogen of wheat, and assess their contribution to pathogenesis. Computational analysis of fungal sRNA and wheat mRNA sequencing datasets was used to define the global sRNA populations in Z. tritici and predict putative mRNA targets in wheat. In total, 389 in planta-induced sRNA loci were identified in Z. tritici. sRNAs generated from some of these loci were predicted to target wheat mRNAs, including some that have previously been implicated in defence against pathogens. However, biochemical approaches were unable to successfully validate targeting of selected wheat mRNAs by fungal sRNAs. Z. tritici gene deletion strains deficient for key RNAi components were generated and virulence bioassays suggested that these are dispensable for full infection of wheat. Nonetheless, our results do point to the existence of non-canonical Dicer-independent pathway(s) for sRNA biogenesis in Z. tritici. dsRNA applied in vitro or generated from an RNA virus vector in planta was ineffective at triggering gene silencing or reducing growth of Z. tritici. We conclude that neither in vitro nor in planta RNAi approaches are likely to be useful for gene function analyses or as a viable control measure
for this pathogen
