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Relatively few ectomycorrhizal fungal species are known to form sclerotia. Usually, sclerotia are initiated at the extraradical mycelium. In this study, we present anatomical and ultrastructural evidence for the formation of sclerotia directly in the hyphal mantle of the mycorrhizal morphotype Pinirhiza sclerotia. A dark-pigmented fungal strain was isolated from Pinirhiza sclerotia and identified by molecular tools as Acephala macrosclerotiorum sp. nov., a close relative of Phialocephala fortinii s.l. As dark septate fungi are known to be mostly endophytic, resyntheses with Pinus sylvestris and A. macrosclerotiorum as well as Populus tremula x Populus tremuloides and A. macrosclerotiorum or P. fortinii s.l. were performed under axenic conditions. No mycorrhizas were found when hybrid aspen was inoculated with A. macrosclerotiorum or P. fortinii. However, A. macrosclerotiorum formed true ectomycorrhizas in vitro with P. sylvestris. Anatomical and ultrastructural features of this ectomycorrhiza are presented. The natural and synthesized ectomycorrhizal morphotypes were identical and characterized by a thin hyphal mantle that bore sclerotia in a later ontogenetic stage. The Hartig net was well-developed and grew up to the endodermis. To our knowledge, this is the first evidence at the anatomical and ultrastructural level that a close relative of P. fortinii s.l. forms true ectomycorrhizas with a coniferous host.