Insertional mutagenesis of Magnaporthe oryzae led to the identification of MCK1, a pathogenicity gene predicted to encode mitogen-activated protein kinase kinase kinase (MAPKKK) homologous to BCK1 in Saccharomyces cerevisiae. Targeted disruption of MCK1 resulted in the fungus undergoing autolysis and showing hypersensitivity to cell-wall-degrading enzyme. The mck1 produced significantly reduced numbers of conidia and developed appressoria in a slightly retarded manner compared with the wild type. Appressorium of the mck1 mutant was unable to penetrate into plant tissues, thereby rendering the mutant nonpathogenic. Cytorrhysis assay and monitoring of lipid mobilization suggested that the appressorial wall was altered, presumably affecting the level of turgor pressure within appressorium. Furthermore, the mck1 mutant failed to grow inside plant tissue. Complementation of the mutated gene restored its ability to cause disease symptoms, demonstrating that MCK1 is required for fungal pathogenicity. Taken together, our results suggest that MCK1 is an MAPKKK involved in maintaining cell wall integrity of M. oryzae, and that remodeling of the cell wall in response to host environments is essential for fungal pathogenesis.

Outbreaks of rice blast have been a threat to the global production of rice. Members of the Magnaporthe grisea species complex cause blast disease on a wide range of gramineous hosts, including cultivated rice and other grass species. Recently, based on phylogenetic analyses and mating tests, isolates from crabgrass were separated from the species complex and named M. grisea. Then other isolates from grasses including rice were named as M. oryzae. Here, we collected 103 isolates from 11 different species of grasses in Korea and analyzed their phylogenetic relationships and pathogenicity. Phylogenetic analyses of multilocus sequences and DNA fingerprinting revealed that the haplotypes of most isolates were associated with their hosts. However, six isolates had different haplotypes from the expectation, suggesting potential host shift in nature. Results of pathogenicity tests demonstrated that 42 isolates from crabgrass and 19 isolates from rice and other grasses showed cross-infectivity on rice and crabgrass, respectively. Interestingly, we also found that the isolates from rice had a distinct deletion in the calmodulin that can be used as a probe.

Two new pregnane glycosides, kidjoranine 3-O-β-D-glucopyranosyl-(1 → 4)-β-D-glucopyranosyl-(1 → 4)-α-L-cymaropyranosyl-(1 → 4)-β-D-cymaropyranosyl-(1→4)-α-L-diginopyranosyl-(1 → 4)-β-D-cymaropyranoside (5) and caudatin 3-O-β-D-glucopyranosyl-(1 → 4)-β-D-glucopyranosyl-(1 → 4)-α-L-cymaropyranosyl-(1 → 4)-β-D-cymaropyranosyl-(1 → 4)-α-L-diginopyranosyl-(1 → 4)-β-D-cymaropyranoside (6), were isolated from the roots of Cynanchum wilfordii along with four known compounds (1-4). The antifungal activities of the six compounds against barley powdery mildew caused by Blumeria graminis f. sp. hordei were compared to the antifungal activity of polyoxin B. The caudatin glycosides (1, 4, and 6) showed stronger antifungal activities than polyoxin B, whereas kidjoranine glycosides (2, 3, and 5) had weaker activities than polyoxin B. A wettable powder-type formulation (C. wilfordii-WP20) of the ethyl acetate extract from C. wilfordii roots prohibited the development of barley powdery mildew much more effectively than the commercial fungicide polyoxin B-WP10. In addition, C. wilfordii-WP20 effectively controlled strawberry powdery mildew caused by Sphaerotheca humuli under greenhouse conditions. Thus, the crude extract containing the pregnane glycosides can be used as a botanical fungicide for the environmentally benign control of powdery mildews.

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