Translational Epigenetics
FraxPath
Innovative RNA interference (RNAi)-mediated control of Hymenoscyphus fraxineus, the pathogen causing ash dieback
It is being investigated whether RNA interference can be used as a non-chemical control method against Hymenoscyphus fraxineus. The aim is to stop an infection of H. fraxineus in infested ash seedlings.
First it is checked whether the essential enzymes of the RNAi mechanism are present in H. fraxineus. Next, molecular biological and subsequent bioinformatic methods will be used to describe the expression levels of genes essential for the fungus (transcriptome analysis) in order to identify suitable target genes. These genes are then to be switched off with the aid of RNAi, thus causing lethal damage to the fungus in the host.
Sequence-homologous double-stranded RNAs (dsRNA) are produced for the target genes and the efficacy of these dsRNAs as H. fraxineus growth inhibitors is tested with in vitro cultures (petri dish) of the fungus. In parallel, the ‘stem injection’ application method is being tested on ash seedlings. The dsRNA solution is injected into a hole drilled into the trunk. By detecting the dsRNA in apical leaves, the RNA transport via the pathways of the ash tree is verified.
This will be followed by efficacy studies of the most promising dsRNA candidates with infected ash seedlings. These studies are first carried out in the greenhouse and then under near-natural conditions. The dsRNA is introduced by stem application or spraying the leaves. In addition, various production methods for dsRNA will be tested and evaluated in terms of their efficiency, with the aim of possibly developing a new, optimised method.
Figure: Schematic representation of the application of double-stranded RNA. The dsRNA solution is introduced into the water channels (xylem) in a borehole. The molecules are then transported systemically in the plant tissue and thus also reach the fungi in the leaves. The uptake of dsRNA by the fungus leads to the degradation of vital genes. This could stop or minimise infection. Scheme: L. Hohenwarter; top photo: FVA; photo below: Robert Vidéki, Doronicum Kft, Bugwood.org (No. 5395833).