Characterization of the Mutants of Alternaria alternata Japanese Pear Pathotype Deficient in Melanin Production and Their Pathogenicity.

Abstract

Alternaria alternata (Fr.) Keissler is known to produce a dark gray pigment, melanin, from acetate via 1, 8-dihydroxynaphthalene (DHN). We isolated three phenotypic classes of color mutants from A. alternata Japanese pear pathotype by treatment of the spores with N-methyl-N'-nitro-N-nitrosoguanidine. These mutants, AKT88-1 (Alm-), AKT88-2 (Brm1-) and AKT88-3 (Brm2-), formed albino, light-brown and brown mycelia, respectively, on potato-dextrose agar plates, whereas the parent strain formed black mycelia. A mutant AKT88-2 accumulated scytalone in culture medium, a key intermediate of the direct pathway of DHN melanin, and mutant AKT88-3 accumulated 2-hydroxyjuglone (2-HJ), a characteristic shunt product from 1, 3, 8-trihydroxynaphthalene (1, 3, 8-THN). Mutant AKT88-1 did not accumulate scytalone, 2-HJ, or flaviolin, a shunt product from 1, 3, 6, 8-tetrahydroxynaphthalene (1, 3, 6, 8-THN). Mutant AKT88-1 was melanized when it was grown on medium supplemented with scytalone, but not AKT88-2 and AKT88-3. These results suggest that mutant AKT88-1 is deficient in the synthesis of 1, 3, 6, 8-THN from acetate, while mutants AKT88-2 and AKT88-3 are deficient in the steps from scytalone to 1, 3, 8-THN and from 1, 3, 8-THN to vermelone, respectively. All the melanin-deficient mutants and the parent strain developed approximately same numbers of lesions with similar size on susceptible pear leaves in laboratory tests. Tricyclazole, a known inhibitor of the biosynthesis of DHN melanin, inhibited pigmentation of cultured mycelia of the parent strain, but not the ability of the spores to penetrate artificial membranes or their ability to infect host leaves. These results indicate that the ability to produce melanin may not be relevant to the pathogenicity of A. alternata Japanese pear pathotype.