Established vegetation can facilitate the ectomycorrhizal infection of seedlings, but it is not known whether this interaction is limited by the phylogenetic relatedness of trees and seedlings. We use a series of bioassay experiments to test whether soil modification by different ectomycorrhizal tree species causes different levels of seedling infection, whether the extent of seedling infection is a function of the relatedness of tree and seedling, and whether the effect of trees on seedlings is mediated by biotic or abiotic soil factors. We found that soils from under different tree species do vary in their mycorrhizal infectiveness. However, this variation is not related to the genetic relatedness of trees and seedlings but instead, appears to be an attribute of the overstory species, irrespective of seedling species, mediated through a suite of humus- and base-cation-related abiotic effects on soils. Modification of abiotic soil properties by overstory trees should be considered as an important factor in the effect of different overstory trees on the extent of seedling mycorrhizal infection.