Ornithine decarboxylase knockout in Tapesia yallundae abolishes infection plaque formation in vitro but does not reduce virulence toward wheat

A knockout strain of Tapesia yallundae lacking the single ornithine decarbo xylase (ODC) allele has been created by targeted gene replacement. A centra l region of the ODC gene was isolated by polymerase chain reaction with deg enerate oligonucleotides and used to probe a lambda genomic library. The ge ne was sequenced, and the encoded ODC protein sequence was shown to be simi lar to those from other fungi. The functionality of the T. yallundae ODC wa s confirmed by complementation of an Aspergillus nidulans mutant (puA) stra in devoid of ODC activity, restoring growth in the absence of exogenous pol yamines. Transformation-mediated gene replacement was used to create strain s that were auxotrophic for putrescine and lack ODC coding sequences. ODC k nockout strains were unable to differentiate infection structures after in vitro induction and showed an abnormal hyphal branching phenotype. Pathogen icity studies on these mutants showed that, surprisingly, they are not redu ced in virulence com pared with nondisrupted transformants. This suggests t hat the strains carrying an ODC disruption can obtain sufficient polyamines from the host plant for normal growth and differentiation and, therefore, that fungal ODC may not be a suitable target for fungicides.