Generation of a ToxA knockout strain of the wheat tan spot pathogen Pyrenophora tritici-repentis

Abstract

The necrotrophic fungal pathogen Pyrenophora tritici-repentis causes tan spot, a major disease of wheat throughout the world. The proteinaceous effector ToxA is responsible for foliar necrosis on ToxA sensitive wheat genotypes. The single copy ToxA gene was deleted from a wild-type race 1 P. tritici-repentis isolate via homologous recombination of a knockout construct. Expression of the ToxA transcript was found to be absent in transformants (toxa), as was ToxA protein production in fungal culture filtrates. Plant bioassays were conducted to test transformant pathogenicity. The toxa strains were unable to induce necrosis on ToxA sensitivity wheat genotypes. To our knowledge, this is the first demonstration of a targeted gene knockout in P. tritici-repentis. The ability to undertake gene deletions will facilitate the characterisation of other pathogenicity effectors of this economically significant necrotroph.