which could be identified, formed sclerotia in culture. Thus, the sclerotia examined
in this study might have been formed either by C. geophilum or by other fungal taxa.
Keywords Sclerotia · Cenococcum geophilum · T-RFLP · Clone library analysis ·
DSE fungi
2.1 Introduction
Fungal species can respond to environmental stress such as desiccation by forming
resting structures composed of compact masses of hardened mycelia known as
sclerotia. One of the most common soil fungal species that forms sclerotia is
Cenococcum geophilum, which is distributed worldwide (Jany et al. 2002; Dickie
2007) and has more than 200 species of host plants (LoBuglio et al. 1996). The
sclerotia of C. geophilum are abundant and have been examined in many parts of the
world. The biomass of C. geophilum sclerotia was estimated to be 440 kg ha
À1 in
old-growth Norway spruce forests in southern Sweden (Dahlberg et al. 1997) and
2785 kg ha
À1 in a second-growth Douglas fir stand in the Oregon Coast Range
(Fogel and Hunt 1979). The distribution of C. geophilum sclerotia has also been
examined in forest soils of the Harz Mountains in Germany (Watanabe et al. 2004;
Sakagami 2009a). The sclerotia of C. geophilum are distributed in Andosols in
central Japan (Watanabe et al. 2002) and are also abundant in Pinus thunbergii
forests in coastal area of Japan (Matsuda et al. 2009). These studies tentatively
identified sclerotia as the resting bodies of C. geophilum according to a description
of morphological characteristics provided by Trappe (1969) and Massicotte et al.
(1992) without further molecular identification; however, a broad range of fungal
species have the ability to form sclerotia (Smith et al. 2015). Identification of
sclerotia can be difficult mainly because they grow slowly in culture (Chen et al.
2007) and are occasionally difficult to isolate in axenic culture (Obase et al. 2014).
Furthermore, it is difficult to extract DNA from sclerotial cell because they contain a
non-hydrolyzable residue consisting of a highly resistant melanin-like pigment,
which plays an important role in the resistance of sclerotia to chemical and biological
degradation (Chen et al. 2007). On the other hand, sclerotia accumulate relatively
high concentrations of carbohydrates, fats, and proteins during their growth, which
may be a good source of nutrition for the development of other microorganisms
associated with sclerotia (Willetts 1971). These sclerotia-associated microorganisms
provide valuable information for studies on microbial diversity in the rhizosphere,
biocontrol for plant pathogens (Zachow et al. 2011), and functional heterogeneity in
fungal adaptations to drought (Homma 1937). We aimed to identify the fungi that
formed the sclerotia collected from Japanese forests, if possible, and to identify the
members of the sclerotia-associated fungal community.
18
K. Narisawa et al.
in this study might have been formed either by C. geophilum or by other fungal taxa.
Keywords Sclerotia · Cenococcum geophilum · T-RFLP · Clone library analysis ·
DSE fungi
2.1 Introduction
Fungal species can respond to environmental stress such as desiccation by forming
resting structures composed of compact masses of hardened mycelia known as
sclerotia. One of the most common soil fungal species that forms sclerotia is
Cenococcum geophilum, which is distributed worldwide (Jany et al. 2002; Dickie
2007) and has more than 200 species of host plants (LoBuglio et al. 1996). The
sclerotia of C. geophilum are abundant and have been examined in many parts of the
world. The biomass of C. geophilum sclerotia was estimated to be 440 kg ha
À1 in
old-growth Norway spruce forests in southern Sweden (Dahlberg et al. 1997) and
2785 kg ha
À1 in a second-growth Douglas fir stand in the Oregon Coast Range
(Fogel and Hunt 1979). The distribution of C. geophilum sclerotia has also been
examined in forest soils of the Harz Mountains in Germany (Watanabe et al. 2004;
Sakagami 2009a). The sclerotia of C. geophilum are distributed in Andosols in
central Japan (Watanabe et al. 2002) and are also abundant in Pinus thunbergii
forests in coastal area of Japan (Matsuda et al. 2009). These studies tentatively
identified sclerotia as the resting bodies of C. geophilum according to a description
of morphological characteristics provided by Trappe (1969) and Massicotte et al.
(1992) without further molecular identification; however, a broad range of fungal
species have the ability to form sclerotia (Smith et al. 2015). Identification of
sclerotia can be difficult mainly because they grow slowly in culture (Chen et al.
2007) and are occasionally difficult to isolate in axenic culture (Obase et al. 2014).
Furthermore, it is difficult to extract DNA from sclerotial cell because they contain a
non-hydrolyzable residue consisting of a highly resistant melanin-like pigment,
which plays an important role in the resistance of sclerotia to chemical and biological
degradation (Chen et al. 2007). On the other hand, sclerotia accumulate relatively
high concentrations of carbohydrates, fats, and proteins during their growth, which
may be a good source of nutrition for the development of other microorganisms
associated with sclerotia (Willetts 1971). These sclerotia-associated microorganisms
provide valuable information for studies on microbial diversity in the rhizosphere,
biocontrol for plant pathogens (Zachow et al. 2011), and functional heterogeneity in
fungal adaptations to drought (Homma 1937). We aimed to identify the fungi that
formed the sclerotia collected from Japanese forests, if possible, and to identify the
members of the sclerotia-associated fungal community.
18
K. Narisawa et al.