Online publication; available at: https://jats-truffles.org/truffology/

Truffology 5 (1): 23–32 (2022)

Original peer-reviewed article （原著論文 ； 査読有）Scleroderma capeverdeanum, a subhypogeous fungus new to Japan,
collected from the Eucalyptus plantation in Ibaraki Prefecture
茨城県のユーカリ植栽地において採集された日本新産の半地下生菌， Scleroderma capeverdeanum
Taiga Kasuya1*, Mihoko Uzawa2, Kentaro Hosaka3
糟谷 大河 1*, 鵜沢 美穂子 2, 保坂 健太郎 3
1

Department of Biology, Keio University, 4-1-1 Hiyoshi, Kohoku-ku, Yokohama-shi, Kanagawa 223-8521, Japan慶應義塾大学生物学教室， 〒 223-8521 神奈川県横浜市港北区日吉 4-1-1

2

Ibaraki Nature Museum, 700 Osaki, Bando-shi, Ibaraki 306-0622, Japan
ミュージアムパーク茨城県自然博物館， 〒 306-0622 茨城県坂東市大崎 700

3

Department of Botany, National Museum of Nature and Science, 4-1-1 Amakubo, Tsukuba-shi, Ibaraki 305-0005, Japan
国立科学博物館植物研究部，〒 305-0005 茨城県つくば市天久保 4-1-1* Corresponding author （主著者）

E-mail: tkasuya@keio.jp

Abstract
A subhypogeous sclerodermataceous fungus was collected in the Eucalyptus plantation of Ibaraki Prefecture,
central Japan. Based on morphological observations and phylogenetic analyses using nuclear ribosomal DNA
sequences, the present fungus was identified as Scleroderma capeverdeanum, belonging to Sclerodermataceae(Boletales). Scleroderma capeverdeanum is new to Japan and characterized by small-sized (4–10 mm in diam.),sessile basidiomata, yellowish to yellowish brown exoperidium with brownish, areolate to verrucose scales, andechinulate basidiospores. Japanese specimens of S. capeverdearnum are probably associated with introduced E.
globulus, and therefore they are most likely exotic species introduced from Australia.

要旨
茨城県のユーカリ植栽地において、 ニセショウロ型の半地下生菌の一種が採集された。 本菌について、 形態的特徴の観察および子実体より得られた核リボソーム DNA の塩基配列 を用いた系統解析を行った。その結果、本菌はイグチ目ニセショウロ科に属する Scleroderma capeverdeanum と同定された。 本種は日本新産であり、 子実体が小型 （直径 4–10 mm） で無柄である点、 外皮が黄色から黄褐色で、 網目状からいぼ状にひび割れ、 褐色を帯びた鱗片を有する点、 そして担子胞子が針状突起に覆われる点により特徴づけられる。 日本産標本は Eucalyptus globulus と共生し、 オーストラリアからの外来種であると推察される。 本菌の和名をユーカリキヒメニセショウロとする。
Article Info: Submitted: 14 December 2021

© The Japanese Association for Truffle Science (JATS), 2022

Accepted: 19 March 2022

Published: 31 March 2022

23

Kasuya et al.: Scleroderma capeverdeanum new to Japan

Truffology - Volume 5, Issue 1, 2022

Introduction

studies of macrofungal diversity at the Eucalyptus plantation in

The genus Scleroderma Pers. is currently classified in the order

Ishioka of Ibaraki Prefecture, central Honshu, Japan, three specimens

Boletales based on molecular phylogenetic analyses (Binder &of a subhypogeous sclerodermataceous fungus were collected. Based

Bresinsky, 2002; Binder & Hibbett, 2006; Watling, 2006). Previously,

on morphological observations and phylogenetic analyses usingca. 40 species of the genus were recognized from tropical, subtropical

nuclear ribosomal DNA sequences, the present fungus was identified

to temperate regions of the world (Baseia et al., 2016; Guzmán, 1970;

as Scleroderma capeverdeanum M.P. Martín, M. Dueñas & Telleria.

Guzmán et al., 2013; Nouhra et al., 2012; Ortiz-Rivero et al., 2021;

This species was originally described from Cape Verde, Macaronesia,

Zhang et al., 2013, 2020). In Japan, fifteen species of Scleroderma

off the coast of northwestern Africa (Crous et al., 2016) and it iswere hitherto recorded (Kasuya et al., 2002; Kasuya & Guzmán,

known only from the type locality. Our morphological observations

2007; Yoshimi, 2002). The genus forms epigeous, subhypogeus

and phylogenetic analyses of the Japanese specimens revealed thator hypogeous basidiomata (Beaton & Weste, 1982; Giachini et al.,it is the second distributional record of S. capeverdeanum in the

2000; Guzmán, 1970; Guzmán et al., 2013; Nouhra et al., 2012),

world. Moreover, our study demonstrates that S. capeverdeanum

and many species have been confirmed as ectomycorrhizal with ais presumably associated with Eucalyptus spp. In this paper,

wide range of host plants such as Pinaceae, Fagaceae, Salicaceae andmorphological, phylogenetic and ecological characteristics ofMyrtaceae (Mrak et al., 2017; Nouhra et al., 2012). Japanese species

Japanese specimens of S. capeverdeanum are discussed.

of Scleroderma were usually observed under betulaceous, fagaceousor pinaceous trees (Kasuya et al., 2002; Kasuya & Guzmán, 2007;
Yoshimi, 2002).

Materials and methods
Sample collecting and morphological observations

Some species of Eucalyptus (Myrtaceae) were introduced from

Fresh subhypogeous basidiomata were collected among plant

Australia to Japan early in the 20th century to meet the country’s

debris and rich soil in the exotic Eucalyptus plantation dominateddemand for pulpwood (Shibata & Yoshikawa, 1956). Recently, toby E. globulus Labill. at Otsuka, Ishioka, Ibaraki Prefecture (Fig. 1)

supply the demand for gardening, greening, and feeds of Koala which

during October to December, 2020. Specimens were photographed

are kept at zoos, several plantations of Eucalyptus spp. are maintained

and observed macroscopically. Fresh basidiomata of specimens

in Japan (Inuma et al., 2015; Komaki et al., 2000). Eucalyptus is anwere dried using a food dehydrator (Snackmaster Express FD-60;

ectomycorrhizal tree genus and several species of Scleroderma have

Nesco/American Harvest, Milwaukee, WI, USA) under 46 °C for

been known as its ectomycorrhizal symbionts (Bashir & Khalid,

46 hours. For light microscopy, hand-cut sections of both fresh and2015; Beaton & Weste, 1982; Giachini et al., 2000; Rose et al.,dried specimens were mounted in water, 3% KOH or 70% ethanol

1981). However, only two species, i.e., S. cepa Pers. and S. ﬂavidum

reagent. Dimensions of basidiospores were measured from water-

Ellis & Everh. have previously been recorded in exotic Eucalyptus

mounted sections. More than 50 randomly selected basidiospores

plantations in Japan (Yoshimi, 2002). During the course of our recent

were measured under a light microscope at 1000× magnification. All

Fig. 1. Habitat in the locality of Scleroderma capeverdeanum. A: Eucalyptus globulus trees. B: Numerous fallen leaves and old bark of E. globulus are densely
deposited on the ground.

24© The Japanese Association for Truffle Science (JATS), 2022

Truffology - Volume 5, Issue 1, 2022

Kasuya et al.: Scleroderma capeverdeanum new to Japanmeasurements were performed with Photoruler 1.1.3 (http://inocybe.

specimen were used for the phylogenetic analyses. Additionally,

info/_userdata/ruler/PhotoRuler.html). In addition, the surface features

42 ITS and 26 LSU sequences of sclerodermataceous fungi wereof basidiospores were observed by scanning electron microscopy

retrieved from the NCBI GenBank databases (https://www.ncbi.nlm.

(SEM). For SEM, a small portion from glebal tissue was put onto

nih.gov/) and included in the analyses (Table 1). DNA sequences weredouble-sided adhesive tape on a specimen holder and coated withinitially aligned using MUSCLE v.3.6 (Edgar, 2004a, b), followed byplatinum-palladium using a JFC-1600 Ion Sputter Coater (JEOL,

manual alignment in the data editor of BioEdit ver. 7.0.1 (Hall, 1999).

Tokyo, Japan). Specimen was examined with a JSM-6480LV SEM

A total of 36 ITS and 89 LSU nucleotide positions were respectively(JEOL, Tokyo, Japan) operating at 5 kV. Three specimens examined

excluded from the analyses because of the presence of ambiguously

in this study were deposited at mycological herbaria of Ibaraki Naturealigned regions. Phylogenetic analyses were performed independently

Museum (INM) and National Museum of Nature and Science (TNS)

for ITS and LSU sequences under maximum parsimony (MP) andin Japan.

maximum likelihood (ML) criteria. MP analyses were conductedunder the equally weighted parsimony criterion using PAUP version

DNA preparation, PCR and sequencing

4.0b10 (Swofford, 2002). The analyses were performed using the

DNA extraction, PCR and DNA sequencing were carried out

heuristic search option with tree-bisection-reconnection (TBR)

according to the methods introduced by Kasuya et al. (2012). First,

and MULTREES option, and 1000 replicates of random addition

small fragments of glebal tissue from freshly collected samples

sequence were conducted. Support for the individual nodes waswere soaked in DMSO buffer (Seutin et al., 1991) with thetested by means of bootstrap analysis (Felsenstein, 1985) under the

addition of 100 mM Tris-HCl (pH 8.0) and 0.1 M sodium sulfite

equally weighted parsimony criterion. A bootstrap analysis was based(Na2SO3) at 4 °C, following the procedures of Hosaka (2009),

on 10,000 bootstrap replicates using the heuristic search option with

Hosaka & Castellano (2008), and Hosaka et al. (2010). DNA of

TBR and MULTREES options with ten random additional sequences.

the above specimens was extracted from the tissue fragments

ML analyses were performed using MEGA X (Kumar et al., 2018)

stored in DMSO buffer. DNA extractions used the modified

after testing the best models. According to the lowest BIC (Bayesian

CTAB extraction followed by glass milk purification methods as

Information Criterion) scores, Kimura 2-parameter (Kimura, 1980)

summarized by Hosaka (2009) and Hosaka & Castellano (2008).

with gamma distributed rate heterogenetic and a proportion of

DNA sequence data were obtained from the nuclear ribosomal

invariant sites (K2+G+I) and Tamura-Nei (Tamura & Nei, 1993) withITS region and a part of LSU. For amplifying the ITS region, thegamma distributed rate heterogenetic and a proportion of invariant

primer combination of ITS5 and ITS4 (White et al., 1990) was

sites (TN93+G+I) were chosen as the optimal substitution models

used. For amplifying the LSU, the combination of LR0R and LR5

for the analyses of the ITS and LSU datasets, respectively. Initial(Vilgalys & Hester, 1990) was used. PCR were carried out using

trees for the heuristic search were obtained automatically by applying

20 µl reaction volume, each containing 1 µl genomic DNA, 1 µl

Neighbor-Join and BioNJ algorithms to a matrix of pairwise distances

dNTP (4 mM), 1 µl each primer (8 µM), 0.5 units Taq polymerase

estimated using the Maximum Composite Likelihood (MCL)(Takara Bio Inc., Tokyo, Japan), 2 µl MgCl2 (25 mM), and 2 µl

approach, and then selecting the topology with superior log likelihood

bovine serum albumin (BSA). Cycling parameters for ITS region

value. For the ML analyses, clade robustness was assessed using

and LSU followed Kasuya et al. (2012). PCR products werea bootstrap analysis with 1000 replicates. Sequences of Pisolithus

electrophoresed in 1% agarose gels stained with ethidium bromide

arhizus (Scop.) Rauschert were selected for outgroups, which wereand visualized under UV light. When amplification bands werestrongly supported as the sister of the major clade containing theconfirmed, PCR products were then purified using the ExoSap-IT

genus Scleroderma (Baseia et al., 2016; Binder & Hibbett, 2006;(Millipore, Molsheim, France) and directly sequenced using the

Hosaka, 2009; Phosri et al., 2009).

Big Dye Terminator Cycle Sequencing Kit (Applied Biosystems,
Norwalk, CT, USA), following the manufacturer’s instructions.

Results

A total of two newly generated sequences from this study wereMorphological observations

deposited in GenBank (Table 1).

Japanese specimens were morphologically identical to S.
capeverdeanum in their sessile, small-sized basidiomata (Fig. 2A–

Phylogenetic analyses
Newly generated ITS and LSU sequences from the Japanese© The Japanese Association for Truffle Science (JATS), 2022B), yellowish to yellowish brown exoperidium covered by brownish,
areolate to verrucose scales (Fig. 2B–C), and globose, densely
25Truffology - Volume 5, Issue 1, 2022

Kasuya et al.: Scleroderma capeverdeanum new to Japan

Table 1. Specimen identification, voucher specimen and/or isolate numbers, origin and GenBank accession numbers for ITS and LSU sequences used forthe present phylogenetic analyses
Species names
Pisolithus arhizus
P. arhizus
Scleroderma areolatum
S. areolatum
S. areolatum
S. aurantium
S. aurantium
S. bermudense
S. bermudense
S. bovista
S. bovista
S. bovista
S. bovista
S. bovista
S. bovista
S. bovista
S. bovista
S. bovista
S. bovista
S. capeverdeanum
S. capeverdeanum
S. cepa
S. cepa
S. citrinum
S. citrinum
S. columnare
S. columnare
S. dictyospora
S. dunensis
S. dunensis
S. dunensis
S. echinatum
S. laeve
S. laeve
S. laeve
S. laeve
S. laeve
S. meridionale
S. michiganense
S. michiganense
S. polyrhizum
S. polyrhizum
S. polyrhizum
S. radicans
S. radicans
S. septentrionale
S. septentrionale
S. sinnamariense
S. sinnamariense
S. sinnamariense
S. verrucosum
S. verrucosum
S. verrucosum
S. xanthochroum
S. yunnanense
S. yunnanense
S. yunnanense
Scleroderma sp.
Scleroderma sp.
Uncultured ectomycorrhizal fungus

GenBank accession no.
ITS
LSU
Italy
FR748132n/a
Wat.139161; 92fbisPISOLIn/an/a
AF336262
588; n/an/a; O3C_4
USA: New York
JX030282n/an/a; Scl_1-1-Qm1H
USA: New York
JX030288
n/a
AWW211; n/aUSAn/aEU718149
n/a; 8_5
China: Sichuan
HM237174 n/a
HB#E02; HBS2
Pakistan
KF802172n/a
BZ3961; n/aBelizen/a
DQ644137n/a; BA 06.04.22
Guadeloupe
FR682092
n/aMG061001_01; n/aGermany: Thuringia
JQ669943
n/a
n/a
Japan
AB099901
n/aNara_ScB84e3; n/aJapan: Shizuoka, Gotenba
AB211267
n/a
TNS-F-82197 (= Kasuya B251); n/a Japan: Yamanashi, Narusawa OL764485
OL752409
TNS-F-82195 (= Kasuya B236); n/a Japan: Nagano, Nagano
OL764484
OL752408
TNS-F-82200 (= Kasuya B804): n/a Japan: Toyama, Toyama
OL764487
OL752412
K80S09; n/a
New Zealand
GQ267487
n/aTrappe26575; n/aUSA
EU819517
EU718178n/a; O1Q_1
USA: New York
JX030277
n/a
W#1149; n/an/an/aAF336264
TNS-F-82205 (= Kasuya B4288); n/a Japan: Ibaraki, Ishioka
OL764491
OL752414
MA-Fungi 87406*; n/aCape Verde: Santiago Island
KU747111
KU747110
JMP0081; n/aUSA
EU819439n/a184; n/an/an/aAF336265n/a; ecmSC2
Czech
JX679368
n/a
AWW212; n/aUSAn/aEU718151
CUB Microbiology KHS3; n/a
Thailand: Chachoengsao
AB459512n/aMS43; n/an/an/aAF336273
MS55; n/an/an/aAF336267
UFRN-Fungus 2551; n/aBrazilKU747116n/a
UFRN-Fungus 2553; n/aBrazilKU747118n/a
UFRN-Fungus 2206; n/aBrazilKU747121n/aMS34; n/an/an/aAF336268
ZLR46; n/a
China
MW553325 MW553729
TNS-F-82199 (= Kasuya B803); n/a Japan: Ishikawa, Komatsun/aOL752411
MCA242; n/aUSA
EU718117n/a
OSC27936; n/aUSA
EU718120n/a27936; n/an/an/aDQ683003
AWW218; n/aUSA
EU718121
EU718152
E00278311; MICSCL2
USA:Wisconsin, Sheboygan
FM213347n/an/a; Scl_3-1-2L
USA: New York
JX030206n/an/a; 11_3
China: Sichuan
HM237173 n/an/a; Scl_1-1-4Nc
USA: North Carolina
JX030195n/a594; n/aUSA
n/a
DQ683000
PDD103563; n/a
New Zealandn/a
KP191692
PDD103558; n/a
New Zealandn/a
KP191693
J. Nitare (12.9.1986); SEPSCL2
Sweden: Norrbotten
FM213336
n/a
A.D. Parker (2.10.1997); SEPSCL3
USA: Wisconsin, Lone RockFM213338
n/a
SINSCL_8; n/a
Thailand: Pranomprai, Roi Et
FM213363
n/a
NAST-FB11; CMU53-210-2
Thailand
HQ687222n/aMS46; n/an/an/aAF336270
SV-5602; n/a
Burkina Faso
FJ840461
n/a
BCC-MPM2605; n/a
Spain: Catalonia
AJ629886n/a
5; n/an/an/aAF336271
AWW311; n/a
Malaysia
EU718126
EU718154
KUN-HKAS 79633A; Ji001A
China: Yunnan
JQ639040
n/a
KUN-HKAS 79633B; Ji001B
China: Yunnan
JQ639041
n/a
KUN-HKAS 79633C; Ji001C
China: Yunnan
JQ639042
n/a
TNS-F-82198 (= Kasuya B714); n/a Japan: Ishikawa, Komatsu
OL764486
OL752410
MEL2295738A; n/aAustralia
n/aEU718181n/a; L5001_Sclerod_Cam_Euc02
Cameroon
FR731681
n/aHerbarium voucher; isolate

Origin*Holotype of Scleroderma capeverdeanum.
"n/a" means information not available. Sequences newly generated in the present study were shown in bold.

References
Phosri et al. (2012)
Binder & Bresinsky (2002)
n/a
n/a
Wilson et al. (2011)
n/aBashir & Khalid (2015)
Louzan et al. (2007)
Sene et al. (2015)
Gube & Dörfelt (2012)
Kanchanaprayudh et al. (2003)
Nara (2006)
Present study
Present study
Present study
Walbert et al. (2010)
Wilson et al. (2011)
n/aBinder & Bresinsky (2002)
Present study
Crous et al. (2016)
Hosaka (2009)
Binder & Bresinsky (2002)
n/a
Wilson et al. (2011)
Disyatat et al. (2016)
Binder & Bresinsky (2002)
Binder & Bresinsky (2002)
Crous et al. (2016)
Crous et al. (2016)
Crous et al. (2016)
Binder & Bresinsky (2002)
Huang et al. (2021)
Present study
Wilson et al. (2011)
Wilson et al. (2011)
Louzan et al. (2007)
Wilson et al. (2011)
Phosri et al. (2009)
n/a
n/a
n/a
Louzan et al. (2007)
n/a
n/a
Phosri et al. (2009)
Phosri et al. (2009)
Phosri et al. (2009)
Siri-in et al. (2014)
Binder & Bresinsky (2002)
Sanon et al. (2009)
Phosri et al. (2007)
Binder & Bresinsky (2002)
Wilson et al. (2011)
Zhang et al. (2013)
Zhang et al. (2013)
Zhang et al. (2013)
Present study
Wilson et al. (2011)
Tedersoo et al. (2011)

echinulate basidiospores (Fig. 2D–E). However, subhypogeous

It had an aligned length of 865 characters including gaps, of which

habit of Japanese specimens is different from the type specimen of

36 characters were constant, 509 variable and phylogeneticallyS. capeverdeanum, which was described as epigeous (Crous et al.,uninformative, and 320 phylogenetically informative. The MP2016). A detailed description and illustrations of the salient features

analysis of the ITS dataset yielded 10,000 most parsimonious

of Japanese specimens are given below.

trees, of which 149 trees were found in the first step of the heuristic
search. Consistency index (CI), retention index (RI) and rescaled

Phylogenetic analyses
The ITS dataset was consisted by 42 ingroup and 1 outgroup taxa.
26

consistency index (RC) of the most parsimonious trees are 0.4487,
0.7205, 0.3233, respectively. The highest log likelihood of the© The Japanese Association for Truffle Science (JATS), 2022

Truffology - Volume 5, Issue 1, 2022

Kasuya et al.: Scleroderma capeverdeanum new to Japan

Fig. 2. Morphological features of Scleroderma capeverdeanum. A: Basidiomata in the natural habitat (INM-2-217835). B: Surface of mature basidioma (INM2-217835). C: Vertical sections of mature basidiomata showing exoperidium and gleba (TNS-F-82205). D: Basidiospores (TNS-F-82205). E: Scanning electron
microscopy image of basidiospores (TNS-F-82205). Bars: A–B = 1 cm; C = 5 mm; D = 10 µm; E = 5 µm.

resulting ML tree of the ITS dataset is -2446.64. A discrete Gamma

informative. The MP analysis of the LSU dataset yielded 10,000

distribution was used to model evolutionary rate differences among

most parsimonious trees, of which 36 trees were found in the firstsites [5 categories (+G, parameter = 0.5939)]. The rate variation

step of the heuristic search. CI, RI and RC of the most parsimonious

model allowed for some sites to be evolutionarily invariable [(+I),

trees are 0.3631, 0.5807, 0.2108, respectively. The highest log

41.58% sites]. The MP and ML analyses resulted in trees that werelikelihood of the resulting ML tree of the LSU dataset is -2656.84.

identical in topology. Hence, only the MP tree of the ITS dataset

A discrete Gamma distribution was used to model evolutionary rateis shown in Fig. 3. By MP and ML, ITS sequences generated fromdifferences among sites [5 categories (+G, parameter = 0.4036)].

Japanese and Cape Verdean samples of S. capeverdeanum were

The rate variation model allowed for some sites to be evolutionarily

placed within a strongly supported clade (MP BS = 100%, ML BS

invariable [(+I), 63.46% sites]. The MP and ML analyses resulted= 99%; Fig. 3) and were distinct from those of the other members ofin trees that were identical in topology. Hence, only the MP tree

Scleroderma.

of the LSU dataset is shown in Fig. 4. By MP and ML, LSU

The LSU dataset was consisted by 26 ingroup and 1 outgroupsequences generated from Japanese and Cape Verdean samples of S.

taxa. It had an aligned length of 1006 characters including

capeverdeanum were placed within a strongly supported clade (MPgaps, of which 89 characters were constant, 836 variable and

BS = 96%, ML BS = 100%; Fig. 4) and were distinct from those ofphylogenetically uninformative, and 81 phylogeneticallythe other members of Scleroderma.

© The Japanese Association for Truffle Science (JATS), 2022

27

Kasuya et al.: Scleroderma capeverdeanum new to Japan

Truffology - Volume 5, Issue 1, 2022

Fig. 3. Phylogenetic tree generated from maximum parsimony (MP) analysis based on the nuclear ribosomal ITS region of
selected sclerodermataceous species. Bootstrap support values (BS) of MP and maximum likelihood (ML) greater than 50% areshown for each node (MP/ML), and BS less than 50% are indicated by an asterisk (*).

Fig. 4. Phylogenetic tree generated from maximum parsimony (MP) analysis based on the nuclear ribosomal LSU of selectedsclerodermataceous species. Bootstrap support values (BS) of MP and maximum likelihood (ML) greater than 50% are shown foreach node (MP/ML), and BS less than 50% are indicated by an asterisk (*).

28

© The Japanese Association for Truffle Science (JATS), 2022

Truffology - Volume 5, Issue 1, 2022

Kasuya et al.: Scleroderma capeverdeanum new to Japan

Taxonomyof S. capeverdeanum were observed among these litter and rich

Scleroderma capeverdeanum M.P. Martín, M. Dueñas & Telleria,soil, as subhypogeous nature. Although the detailed environmental

in Crous et al., Persoonia 36: 413 (2016)

information on the type locality of S. capeverdeanum was not shown
Fig. 2.

by Crous et al. (2016), subhypogeous or epigeous nature of its

Description: Basidiomata (Fig. 2A–B) subhypogeous, globose,basidiomata presumably depend on their habitat. The present species

depressed globose to subglobose, small-sized, 4–10 mm broad,is morphologically similar to S. ﬂavidum in its subhypogeous, sessile

sessile, arising from whitish mycelial strands attached to plant debris

basidiomata with yellowish surface of peridium and echinulate

and soil. Peridium ca. 0.5–1 mm thick, two-layered. Exoperidium

basidiospores (Yoshimi, 2002). However, S. flavidum is clearly(Fig. 2B–C) very thin, yellow to yellowish brown, covered by brown

distinguishable from S. capeverdeanum by its larger basidiomata

to dark brown, areolate to verrucose scales. Endoperidium papery,(3–5 cm in diam.; Yoshimi, 2002) and larger basidiospores (12–16

firm but brittle when dry, whitish. Apical part of peridium irregularlyµm broad including surface ornamentation; Coccia et al., 1990).

dehiscent and opening lacerate pore when mature. Gleba (Fig. 2C)

Another species having yellowish basidiomata, S. sinnamariense

compact when young, becoming powdery when mature, grayish.

Mont. is also distinguished from S. capeverdeanum by its substipitate

Basidiospores (Fig. 2D–E) globose, 8.5–11 µm in diam. including

basidiomata and subreticulate basidiospores (Guzmán, 1970; Guzmán

surface ornamentation, dark brown in 3% KOH, dark reddish brown in& Ovrebo, 2000). Although basidiospores of S. cepa are echinulate

70% ethanol reagent; surface densely echinulate composed of hyaline

and it forms sometimes yellowish basidiomata, it is distinct from S.

spines up to 1.5 µm long. Under SEM, spines of basidiospore surface

capeverdeanum because it has larger basidiomata (up to 4 cm broad;irregularly covered with scaly to tabular materials (Fig. 2E). Basidia

Guzmán, 1970), and thicker peridium (up to 3 mm; Coccia et al.,not observed. Rudimentary hyphae interwoven, septate, thin-walled,1990). Brownish scales on peridium appear in mature basidiomata

4–8 µm broad, hyaline in 3% KOH and 70% ethanol reagent; surfaceof S. capeverdeanum. Although S. verrucosum (Bull.) Pers. also has

sparsely covered with amorphous, hyaline gelatinous materials.

similar verrucose scales on the peridium (Sims et al., 1995), it has

Habitat: Subhypogeous, usually solitary or small groups among plantlarger basidiomata (up to 4 cm broad; Coccia et al., 1990; Guzmán,debris and rich soil under Eucalyptus globulus trees (Fig. 1). Fruiting1970) and larger basidiospores (11–15 µm in diam. including surface

in the Japanese localities occurs in autumn to winter (October toornamentation; Yoshimi, 2002) than S. capeverdeanum.

December).

Results of our phylogenetic analyses (Figs. 3, 4) show theSpecimens examined: JAPAN, Ibaraki Prefecture, Ishioka,

monophyly of the present species, as indicated by Crous et al. (2016).

Otsuka (alt. approx. 140 m asl.), October 12, 2020, coll. T.

Crous et al. (2016) reported that the holotype of S. capeverdeanum

Kasuya, INM-2-217835; same place, October 12, 2020, coll. T.

was collected under Furcraea foetida (L.) Haw. (Asparagaceae)

Kasuya, TNS-F-82205, GenBank accession no.: OL764491 (ITS),

and Lantana camara L. (Verbenaceae), and no relationships with

OL752414 (LSU); same place, December 21, 2020, coll. T. Kasuya,

Eucalyptus were shown in their original description. However,INM-2-217836.

the ITS sequence of S. capeverdeanum from the root sample of E.

Known distribution: Cape Verde (Crous et al., 2016) and Japan (newgrandis W. Hill in Cameroon (GenBank FR731681; Tedersoo etrecord).

al., 2011) was identical with that of fruit body data (Fig. 3). In CapeJapanese name: Yûkari-ki-hime-nise-shoro (“yûkari” = Eucalyptus;

Verde, there are exotic Eucalyptus plantations in wide range of islands“ki-hime” = yellowish, small-sized; “nise-shoro” = the Japanese namein addition to introduced F. foetida and L. camara communities

of Scleroderma).

(Frahm et al., 1996). While it is unclear whether the mycorrhizal hostof the present species is restricted to Eucalyptus, S. capeverdeanum

Discussion

presumably prefers the habitat dominated by eucalypt trees. Their

As outlined above, studied Japanese specimens were morphologicallypotential association with F. foetida and L. camara, however, warrants

almost identical to the original description of S. capeverdeanum (Crous

further studies.

et al., 2016). However, basidiomata of Japanese materials were

Based on the present ITS phylogeny (Fig. 3), S. capeverdeanum

subhypogeous while the type specimen of the present species wasclusters with two sequences of root sample collected from E.

described as epigeous (Crous et al., 2016). Numerous fallen leaves

grandis in China (GenBank HM237173 and HM237174), as S.

and old bark of E. globulus are densely deposited on the groundpolyrhizum (J.F. Gmel.) Pers. and S. aurantium (L.) Pers., and one

in the locality of Japanese specimens (Fig. 1B) and basidiomata

sequence from Pakistan as S. aurantium (GenBank KF802172;© The Japanese Association for Truffle Science (JATS), 2022

29

Truffology - Volume 5, Issue 1, 2022

Kasuya et al.: Scleroderma capeverdeanum new to Japan
Bashir & Khalid, 2015), as a sister group. Moreover, phylogenetic
tree of the LSU dataset (Fig. 4) shows that three sequences fromgrowing in different areas of the Punjab Province, Pakistan. Australian
Journal of Botany 62: 689–697.

Australia (GenBank EU718181; Wilson et al., 2011) as Scleroderma

Beaton G., Weste G. (1982) A new species of Scleroderma from Victoria,

sp., and New Zealand (GenBank KP191692 and KP191693) as

Australia. Transactions of the British Mycological Society 79: 41–43.

S. radicans Lloyd, are sister to S. capeverdeanum. Australian

Binder M., Bresinsky A. (2002) Derivation of a polymorphic lineage of

specimen housed at MEL (2295738A) is the voucher of EU718181,
and it was collected on soil (grass nearby) under a rosaceous plant,
Prunus cerasifera Ehrh. var. atropurpurea H. Jaeger. Althoughrelationships between the above Australian specimen and any
myrtaceous plants is not shown, KP191692 and KP191693 wereobtained from two specimens (103558 and 103563 housed at PDD,
respectively) identified as S. radicans, and both were collected

Gasteromycetes from boletoid ancestors. Mycologia 94: 85–98.
Binder M., Hibbett D.S. (2006) Molecular systematics and biologicaldiversification of Boletales. Mycologia 98: 971–981.
Coccia M., Migliozzi V., Lavorato C. (1990) Studio sul genere
Scleroderma Persoon. Bollettino dell’Associazione Micologia ed
Ecologica Romana 20–21: 3–59.
Crous P.W., Wingfield M.J., Richardson D.M., Le Roux J.J., Strasberg
D., Edwards J., Roets F., Hubka V., Taylor P.W.J., Heykoop M.,

from under Leptospermum (Myrtaceae) trees. These facts suggest

Martín M.P., Moreno G., Sutton D.A., Wiederhold N.P., Barnes

that the S. capeverdeanum and its phylogenetically related taxa

C.W., Carlavilla J.R., Gené J., Giraldo A., Guarnaccia V., Guarro

presumably have some type of preference to myrtaceous plants forJ., Hernández-Restrepo M., Kolařík M., Manjón J.L., Pascoe I.G.,

their habitat. Although further studies on the ecological nature of S.

Popov E.S., Sandoval-Denis M., Woudenberg J.H.C., Acharya K.,

capeverdeanum and its relatives are needed to clarify the relationship

Alexandrova A.V., Alvarado P., Barbosa R.N., Baseia I.G., Blanchette

between them and their associated plants, we treat S. capeverdeanum

R.A., Boekhout T., Burgess T.I., Cano-Lira J.F., Čmoková A., Dimitrov

as an introduced fungal species to Japan from outside because
any native species of Eucalyptus, Leptospermum, Furcraea andLantana are not known in the country. While it is unknown whether
the origin of S. capeverdeanum is Cape Verde or not, this species
was probably introduced to Japan from Australia with E. globulus.
Further investigations of Scleroderma in worldwide eucalypt

R.A., Dyakov M.Y., Dueñas M., Dutta A.K., Esteve-Raventós F.,
Fedosova A.G., Fournier J., Gamboa P., Gouliamova D.E., Grebenc
T., Groenewald M., Hanse B., Hardy G.E.St.J., Held B.W., Jurjević
Ž., Kaewgrajang T., Latha K.P.D., Lombard L., Luangsa-Ard J.J.,
Lysková P., Mallátová N., Manimohan P., Miller A.N., Mirabolfathy M.,
Morozova O.V., Obodai M., Oliveira N.T., Ordóñez M.E., Otto E.C.,
Paloi S., Peterson S.W., Phosri C., Roux J., Salazar W.A., Sánchez

forests will demonstrate the origin and geographic distribution of S.

A., Sarria G.A., Shin H.-D., Silva B.D.B., Silva G.A., Smith M.Th.,

capeverdeanum.

Souza-Motta C.M., Stchigel A.M., Stoilova-Disheva M.M., Sulzbacher
M.A., Telleria M.T., Toapanta C., Traba J.M., Valenzuela-Lopez N.,

Acknowledgements
We are very much obliged to Ms. Kyung-Ok Nam and Ms. Megumi
Otsuka of the National Museum of Nature and Science for help withassisting molecular experiments. We also thank Mr. Minoru Nakajima,
Mr. Wajiro Yoshitake and Mr. Yoshinori Inaba for facilitating thefieldwork. We are grateful to Dr. Tamao Hatta of Chiba Institute of
Science for his skillful technical assistance and for helpful suggestions
to SEM observation. This work was supported by research grants
from the committee of the introduced species management of Ibaraki
Prefectural Government and the comprehensive surveys of plants,
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