Isolation, identification and characterization of Phytophthora katsurae, causing chestnut ink disease in Korea

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Since July 2005, survey of chestnut ink disease was carried out in chestnut stands located at southern parts of Korea. Dead chestnut trees showing inky ooze on necrotic trunks were found in two different locations. In order to isolate and identify the causal fungus, infected tissues and soil samples around dead or dying trees were collected and placed on Phytophthora-selective medium. Rhododendron and chestnut tree leaves were used as a bait to isolate the fungus from soil samples by attracting zoospores in soil suspensions. On V-8 culture medium, the isolates produced homothallic oogonia with protuberances (34.0-46.2×21.9-26.7㎛) abundantly, but did not produced sporangia. Mass production of sporangia was possible by immersing agar plugs with actively growing mycelium in the creek water at 18oC for 3 days. Sporangia were papillate, and ovoid to obpyriform (17.0-38.9×14.6-29.2㎛) in shape. Comparison of the ITS sequences revealed that the isolates had 100% identity to the P. katsurae isolates from Japan and New Zealand and 99.6% identity to other P. katsurae isolates. All of the examined isolates from Korea were completely identical to each other in ITS sequence. Numerous sporangia were formed in filtered as well as unfiltered creek water, but no sporangia formed in sterilized distilled water. Light induced sporangia formation, but has no influence on oospore formation. Amendments of β-sitosterol in culture media have no significant effect on mycelial growth but significantly stimulate oospore and sporangia formation.

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