Four mycelial strains of Entoloma clypeatum species complex form ectomycorrhiza-like roots with Pyrus betulifolia seedlings in vitro, and one develops fruiting bodies 2 months after inoculation

Abstract

Entoloma clypeatum species complex (ECSC) forms ectomycorrhiza-like roots (EMLR) with host plant species of Rosaceae or Ulmaceae. The EMLR colonized with ECSC are characterized by a thick fungal mantle, absence of a Hartig net structure, and collapse of the apical meristem caused by hyphal invasion. Some researchers have suggested parasitism of ECSC because of this unique mode of colonization; however, the nature of the interaction between ECSC and host plants has not been investigated in co-culture because of the difficulty of culturing this group of fungi. We established a procedure to synthesize EMLR of ECSC on pear seedlings using fungal cultures. Three conspecific strains of ECSC isolated from basidiospores and one strain isolated from EMLR were tested. Cultured mycelia were inoculated onto a modified Norkrans’ C (MNC) or Hyponex-yeast-glucose (HYG) medium slant on the bottom of a polycarbonate jar and covered with autoclaved andosol or a vermiculite/sphagnum moss mixture (VSM); an axenically cultivated Pyrus betulifolia seedling was then planted in the jar. Five months after inoculation, the formation of EMLR with Hartig net-like hyphae was confirmed in all of the experimental plots. However, the rate of root colonization was significantly higher in experimental plots using andosol than in those using VSM. The growth of pear seedlings was similar irrespective of the level of root colonization, suggesting commensalism rather than parasitism of ECSC. One experimental plot using strain A3, an MNC slant, and andosol as a substrate produced ECSC fruiting bodies with mature basidia and basidiospores. The results suggested that our procedure enables the synthesis of EMLR of ECSC and cultivation of their fruiting bodies.

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https://doi.org/10.1007/s00572-020-00994-4
ORIGINAL ARTICLE
Four mycelial strains ofEntoloma clypeatum species complex form
ectomycorrhiza‑like roots withPyrus betulifolia seedlings invitro,
andone develops fruiting bodies 2months afterinoculation
ManamiShishikura1· YoshihiroTakemura2· KozueSotome3· NitaroMaekawa3· AkiraNakagiri3· NaokiEndo3
Received: 23 March 2020 / Accepted: 12 October 2020
© Springer-Verlag GmbH Germany, part of Springer Nature 2020
Abstract
Entoloma clypeatum species complex (ECSC) forms ectomycorrhiza-like roots (EMLR) with host plant species of Rosaceae
or Ulmaceae. The EMLR colonized with ECSC are characterized by a thick fungal mantle, absence of a Hartig net structure,
and collapse of the apical meristem caused by hyphal invasion. Some researchers have suggested parasitism of ECSC because
of this unique mode of colonization; however, the nature of the interaction between ECSC and host plants has not been
investigated in co-culture because of the difficulty of culturing this group of fungi. We established a procedure to synthesize
EMLR of ECSC on pear seedlings using fungal cultures. Three conspecific strains of ECSC isolated from basidiospores
and one strain isolated from EMLR were tested. Cultured mycelia were inoculated onto a modified Norkrans’ C (MNC) or
Hyponex-yeast-glucose (HYG) medium slant on the bottom of a polycarbonate jar and covered with autoclaved andosol or a
vermiculite/sphagnum moss mixture (VSM); an axenically cultivated Pyrus betulifolia seedling was then planted in the jar.
Five months after inoculation, the formation of EMLR with Hartig net-like hyphae was confirmed in all of the experimental
plots. However, the rate of root colonization was significantly higher in experimental plots using andosol than in those using
VSM. The growth of pear seedlings was similar irrespective of the level of root colonization, suggesting commensalism
rather than parasitism of ECSC. One experimental plot using strain A3, an MNC slant, and andosol as a substrate produced
ECSC fruiting bodies with mature basidia and basidiospores. The results suggested that our procedure enables the synthesis
of EMLR of ECSC and cultivation of their fruiting bodies.
Keywords Cultivation· Edible mushroom· Hartig net-like hyphae· Japanese pear· Mycorrhizal synthesis· Parasitism or
commensalism
Introduction
Mycorrhizae are symbiotic organs comprising fungal myce-
lia and plant roots in which bidirectional nutrient transfer
occurs (Smith and Read 2008). Mycorrhizae are morpho-
logically and anatomically categorized into at least seven
types (Imhof 2009); the two major types are arbuscular
mycorrhiza (AM) and ectomycorrhiza (EM). The EM type
is characterized by a fungal sheath (or fungal mantle) and a
Hartig net with labyrinthine hyphae that penetrate the gaps
between cortical cells without directly penetrating those
cells (Smith and Read 2008). Many mycorrhizal fungi
belonging to Basidiomycota (e.g., Tricholoma matsutake
(S. Ito & S. Imai) Singer) form EM on the host plant (Smith
and Read 2008).
Entoloma clypeatum (L.) P. Kumm. and its related spe-
cies (E. clypeatum species complex; ECSC) are members
of the family Entolomataceae, order Agaricales, and phy-
lum Basidiomycota; these species are associated with eco-
nomically important rosaceous plant taxa, including Rosa,
Prunus, Pyrus, and Malus (Noordeloos 1981; Kobayashi
and Hatano 2001; Kobayashi etal. 2003). On such hosts,
ECSC form anatomically unique EM-like roots (EMLR)
* Naoki Endo
endo_nao@tottori-u.ac.jp; caesareae@yahoo.co.jp
1 Graduate School ofSustainable Science, Tottori University,
4-101 Koyama, Tottori680-8553, Japan
2 Faculty ofAgriculture, Tottori University, 4-101 Koyama,
Tottori680-8553, Japan
3 Fungus/Mushroom Resource andResearch Center,
Faculty ofAgriculture, Tottori University, 4-101 Koyama,
Tottori680-8553, Japan
Mycorrhiza (2021) 30:31–42
/Published online: 26 2020
October
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