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    In three related experiments, root systems of 2-year-old Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) seedlings were dip-inoculated with a viscous blend of Leptographium wageneri var. pseudotsugae Harrington and Cobb spores and hyphal fragments and planted in a sterile potting medium. Infection frequency and points of entry were evaluated for dormant seedlings and seedlings that had been active for 4 and 8 weeks. All putative black stain infections and other areas of sapwood discoloration were free-hand sectioned and examined with a bright-field microscope and phase-contrast optics. This technique was shown to be 100% reliable in a prior experiment with 223 inoculated Douglas-fir seedlings that paired microscope examinations with pathogen isolation. In this study, all lesions were sectioned and examined at 250–1000 diameters magnification for the presence of L. wageneri var. pseudotsugae hyphae and characteristic pathological anatomy. Complete root system dissections revealed that L. wageneri var. pseudotsugae infected roots through wounds and natural openings where a direct path to the xylem was exposed and never penetrated living cortical or cambial tissues to infect its host. Among the dormant inoculated seedlings, 63% of infections occurred through wounds associated with nursery handling. Wound infection frequency decreased to zero in seedlings inoculated 8 weeks after coming out of dormancy. Seedlings inoculated 4 and 8 weeks after coming out of dormancy were most frequently infected through openings occurring at sites of new lateral root initiation. Infection of dead fine root stubs suggested that during periods of increased fine root mortality, these sites may be important for the new infection of healthy trees and egress from already diseased trees.

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    Hessburg, Paul F.; Hansen, Everett M. 2000. Infection of Douglas-fir by Leptographium wageneri. Canadian Journal of Botany. 78: 1254-1261

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