Isolation and characterisation of Gaeumannomyces graminis var. graminis melanin mutants

Gaeumannomyces graminis var. graminis produces 1,8-dihydroxynaphthalene (DHN) melanin in its hyphal and hyphopodial cell walls. We isolated G. graminis mutants that were affected in their melanin biosynthesis. One was unable to synthesize DHN-melanin and, because it accumulated 2-hydroxyjuglone, a DHN melanin pathway shunt product, it is most likely to be defective in the reductase that catalyzes the conversion of 1,3,8-trihydroxynaphthalene to vermelone, the penultimate reaction in DHN synthesis. Genetic crosses with our wild-type strain indicated that this trihydroxynaphthalene reductase deficiency was the result of a single mutation. Another mutant constitutively synthesized DHN melanin and genetic crosses with our wild-type strain suggested that this heavily melanized mutant had a single mutation responsible for its phenotype. This mutant produced more melanin than the wild-type strain as measured by Azure A binding to melanin. The wild type and constitutively melanized mutant hyphae were more hydrophobic and more resistant to lyric enzymes, benomyl, restrictive temperature, and uv light than the non-melanized mutant, which also autolysed more readily. The non-melanized mutant was not more sensitive to heavy metal than the melanized strains. In addition, the non-melanized mutant was unaltered in pathogenicity to rice, whereas the constitutively melanized mutant was less pathogenic. The constitutively melanized mutant produced less extracellular lytic enzymes than the wild-type and the non-melanized mutant, which may explain its reduced virulence.