Xylaria furcata reconsidered and nine resembling species

Abstract

Background:
Xylaria collections from termite nests with dichotomously branched stromata have been identified as X. furcata. However, Léveillé's original material is no longer available, and the modern interpretation of X. furcata is based on a 1908 collection made by von Höhnel from termite nests at Buitenzorg Botanical Garden in Java. A packet of this von Höhnel material at FH was designated as the neotype by Rogers et al. in 2005.

Results:
We reexamined the neotype from FH and its duplicates from various herbaria and found that three different species were mixed in these specimens. Despite that all of them have dichotomously branched stromata and tiny ascospores, only one fits the 2005 neotypification of X. furcata, where exposed perithecial mounds on the stromatal surface were unambiguously indicated. This portion of material is redesignated as the neotype, while the other two species with immersed perithecia are described as new: X. hoehnelii and X. robustifurcata. The ITS sequence obtained from the neotype helped us designate a specimen with cultures obtained from it as the epitype. From specimens identifiable as X. furcata, we describe four new species: X. brevifurcata, X. furcatula, X. insignifurcata, and X. tenellifurcata. Additionally, we recognize X. furcata var. hirsuta at the species level as X. hirsuta and consider X. scoparia a distinct species rather than a synonym of X. furcata. Molecular phylogenetic analyses based on three protein-coding loci showed that X. furcata and resembling species were grouped into two clusters: the X. furcata cluster with half-exposed to fully exposed perithecial mounds and the X. hoehnelii cluster with largely immersed perithecial mounds.

Conclusion:
Ten species are recognized for X. furcata and resembling species, all of which could have been identified as X. furcata in the past. Its diversity has been overlooked primarily due to the small and similar stromata. Several additional species have been confirmed to be related to X. furcata by DNA sequences but are yet to be described due to the lack of mature stromata. While the species diversity of macrotermitine termites is equally high in Africa as in Asia, all of the species are primarily found in Asia, with X. hirsuta as the only exception. This suggests that there may be many more undiscovered species for this fungal group.

Full text

Juetal. Botanical Studies (2023) 64:21
https://doi.org/10.1186/s40529-023-00392-x
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Botanical Studies
Xylaria furcata reconsidered andnine
resembling species
Yu‑Ming Ju1* , Huei‑Mei Hsieh1 and Nuttika Suwannasai2
Abstract
Background Xylaria collections from termite nests with dichotomously branched stromata have been identified as X.
furcata. However, Léveillé’s original material is no longer available, and the modern interpretation of X. furcata is based
on a 1908 collection made by von Höhnel from termite nests at Buitenzorg Botanical Garden in Java. A packet of this
von Höhnel material at FH was designated as the neotype by Rogers et al. in 2005.
Results We reexamined the neotype from FH and its duplicates from various herbaria and found that three different
species were mixed in these specimens. Despite that all of them have dichotomously branched stromata and tiny
ascospores, only one fits the 2005 neotypification of X. furcata, where exposed perithecial mounds on the stromatal
surface were unambiguously indicated. This portion of material is redesignated as the neotype, while the other two
species with immersed perithecia are described as new: X. hoehnelii and X. robustifurcata. The ITS sequence obtained
from the neotype helped us designate a specimen with cultures obtained from it as the epitype. From specimens
identifiable as X. furcata, we describe four new species: X. brevifurcata, X. furcatula, X. insignifurcata, and X. tenellifurcata.
Additionally, we recognize X. furcata var. hirsuta at the species level as X. hirsuta and consider X. scoparia a distinct spe‑
cies rather than a synonym of X. furcata. Molecular phylogenetic analyses based on three protein‑coding loci showed
that X. furcata and resembling species were grouped into two clusters: the X. furcata cluster with half‑exposed to fully
exposed perithecial mounds and the X. hoehnelii cluster with largely immersed perithecial mounds.
Conclusion Ten species are recognized for X. furcata and resembling species, all of which could have been identi‑
fied as X. furcata in the past. Its diversity has been overlooked primarily due to the small and similar stromata. Several
additional species have been confirmed to be related to X. furcata by DNA sequences but are yet to be described
due to the lack of mature stromata. While the species diversity of macrotermitine termites is equally high in Africa
as in Asia, all of the species are primarily found in Asia, with X. hirsuta as the only exception. This suggests that there
may be many more undiscovered species for this fungal group.
Keywords Systematics, Termite nests, Xylariaceae
Background
Specimens of Xylaria Hill ex Schrank collected from
termite nests and featuring antler-like dichotomously
branched stromata have commonly been referred to as
X. furcata Fr. is name was proposed by Fries (1851) to
replace Sphaeria dichotoma Lév., which was published
by Léveillé (1845) on the basis of a collection made by
Pieter Willem Korthals from Java, Indonesia. Fries (1851)
replaced S. dichotoma with X. furcata because he had
recombined the epithet of Hypoxylon dichotomum Mont.
with Xylaria to form X. dichotoma (Mont.) Fr. and thus
*Correspondence:
Yu‑Ming Ju
yumingju@gate.sinica.edu.tw
1 Institute of Plant and Microbial Biology, Academia Sinica, Nankang,
Taipei 11529, Taiwan
2 Department of Microbiology, Faculty of Science, Srinakharinwirot
University, 114 Sukhumvit 23, Wathana, Bangkok 10110, Thailand

Page 2 of 27
Juetal. Botanical Studies (2023) 64:21
could not recombine S. dichotoma with Xylaria, which
would have created a later homonym. Léveillé (1845)
compared S. dichotoma with S. scopiformis Kunze, an
invalidly published name based on a Suriname collection
made by Christoph Weigelt in 1827, and stated that both
are sterile. e herbarium of Joseph-Henri Léveillé was
destroyed during the Franco-Prussian war in 1870–1871.
Searching for the Léveillé material of S. dichotoma at L
and PC or the Fries material of X. furcata at UPS was
unfortunately fruitless (Rogers etal. 2005).
e modern interpretation of X. furcata is rooted in
von Höhnel (1908) who believed that he had collected
X. furcata from termite nests at Buitenzorg Botani-
cal Garden of Java, currently Bogor Botanical Gardens,
regardless of the rather vague account on X. furcata [≡ S.
dichotoma] in Léveillé (1845), where the Korthals mate-
rial was recorded as being immature and associated with
trunk. von Höhnel (1908) sent portions of his ample X.
furcata collection made during 1907–1908 from Buiten-
zorg Botanical Garden to his contemporaries Heinrich
Rehm and Giacomo Bresadola and stated that his inter-
pretation of X. furcata was agreed by both of them. Rehm
kept a packet for his own herbarium and distributed the
rest of the von Höhnel collection as “1812. Xylaria (Xylo-
styla) furcata Fr.” in the exsiccati Ascomyceten that he
issued. A packet of this von Höhnel material at FH was
designated as the neotype by Rogers etal. (2005).
Petch (1906), who studied X. furcata in Sri Lanka,
believed that there was only one Xylaria species from
termite nests, X. nigripes (Klotsch) M. C. Cooke, and
thought that it could have many morphs, with stromata
branched or unbranched, robust or delicate, acute or
rounded at apices. However, Petch (1913) changed his
earlier view of one-species concept and accepted X. fur-
cata as a separate species from X. nigripes, apparently
influenced by von Höhnel (1908). Nonetheless, the two-
species concept of von Höhnel (1908) still underesti-
mated the diversity of Xylaria species from termite nests.
An example can be found in his collection of X. furcata
from Buitenzorg Botanical Garden, which contains stro-
mata quite variable in shape and provide a clue to the
mixed nature of his collection, as shown in Plates 3 and
4, in von Höhnel (1908). After studying the parts of the
von Höhnel collection of X. furcata stored in FH, K, and
S, we identified four different species: X. furcata, X. sco-
paria Pat., and two species described as new herein: X.
hoehnelii and X. robustifurcata.
While studying Xylariaceae in Congo, Dennis (1961)
stated that he followed Petch’s concept of X. furcata, but
the cited specimen Louis 14866 is unlike Petch’s material
from Sri Lanka in having hirsute stromata. Like Petch’s
material, von Höhnel’s X. furcata from Indonesia also
has glabrous stromata. erefore, Rogers et al. (2005)
described Louis 14866 as X. furcata var. hirsuta J. D. Rog-
ers & Y.-M. Ju.
We studied numerous specimens that were identified
or suspected as X. furcata and obtained cultures from
viable ones. After examining their teleomorphic and
anamorphic characteristics and analyzing the sequences
of three protein-coding loci and ITS, we concluded that
Xylaria species from termite nests with delicate forked
stromata consist of ten different species. Among these,
only X. furcata, its var. hirsuta, and X. scoparia haven
been described, while others are newly described herein.
e neotype designated for X. furcata in Rogers et al.
(2005) should be reconsidered because it was based on a
Höhnel packet in FH containing more than one species.
Methods
Fungal materials andobservations
Specimens labeled as X. furcata were loaned from vari-
ous herbaria. Culturable materials identifiable as X. fur-
cata were collected from nesting sites of Odontotermes
formosanus in Taiwan. Some of the stromata are fully
mature, bearing asci and ascospores, while others are
sterile or bear conidia only. Fresh stromatal tissue was
placed on SME medium (Kenerley and Rogers 1976)
to obtain cultures. If cultures could not be isolated in a
timely manner, freshly collected stromata were stored
in a moist condition in the fridge to prevent drying and
mold growth. Resulting colonies were transferred to
9-cm plastic Petri dishes containing 2% Difco oatmeal
agar (OMA) for culture descriptions and incubated at
20°C under 12h fluorescent light. Asci, ascospores, con-
idiogenous cells, and conidia were examined using differ-
ential interference contrast microscopy (DIC) and bright
field microscopy (BF). Material was mounted in water
and Melzer’s iodine reagent for examination by DIC and
BF. Cultures were deposited at BCRC (the Bioresource
Collection and Research Center, Hsin-chu, Taiwan). Only
a culture was deposited if cultures were obtained from
multiple stromata of a specimen. ose stromata with
cultures isolated or DNAs extracted were each labeled
with a number prefixed with “YMJ.”
Phylogenetic analyses
Sequences of the loci for β-tubulin (β-TUB) and α-actin
(α-ACT ) were obtained from X. furcata and resembling
species following Hsieh etal. (2005), while those of the
loci for the second largest subunit of RNA polymer-
ase II (RPB2) and nuc rDNA internal transcribed spac-
ers (ITS = ITS1-5.8S-ITS2) were obtained following
Hsieh etal. (2010) and Hsieh et al. (2009), respectively.
Obtained sequences from X. furcata and resembling spe-
cies were listed in Table 1 and deposited at GenBank.

Page 3 of 27
Juetal. Botanical Studies (2023) 64:21
Table 1 Species and isolates of the X. furcata and resembling species included in the phylogenetic analyses. Sequences in boldface
were generated in this study
Taxon Origin Collecting data GenBank Accession number
β-tubulin gene α-actin gene RPB2 gene ITS
X. brevifurcata Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ986 OQ818431 OQ818371 OQ851525 OQ845481
X. brevifurcata Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1381 from HOLOTYPE OQ818432 OQ818372 OQ851526 OQ845482
X. furcata Fr. Indonesia From NEOTYPE – – – OQ848122
X. furcata Fr. Indonesia Stromata emerging from fungus comb
(FH) – – – OQ848123
X. furcata Fr. Taiwan YMJ646, as X. sp. 4 in Hsieh et al. (2010) GQ502715 GQ853050 GQ853032 GU324757
X. furcata Fr. Taiwan YMJ956 OQ818449 OQ818389 OQ851543 OQ842973
X. furcata Fr. Taiwan YMJ978 OQ818450 OQ818390 OQ851544 OQ842974
X. furcata Fr. Taiwan YMJ979 OQ818451 OQ818391 OQ851545 OQ842975
X. furcata Fr. Taiwan YMJ990 OQ818452 OQ818392 OQ851546 OQ842976
X. furcata Fr. Taiwan YMJ993 OQ818453 OQ818393 OQ851547 OQ842977
X. furcata Fr. Taiwan YMJ994 OQ818454 OQ818394 OQ851548 OQ842978
X. furcata Fr. Taiwan YMJ1100 OQ818455 OQ818395 OQ851549 OQ842979
X. furcata Fr. Taiwan YMJ1101 OQ818456 OQ818396 OQ851550 OQ842980
X. furcata Fr. Taiwan YMJ1102 OQ818457 OQ818397 OQ851551 OQ842981
X. furcata Fr. Taiwan YMJ1120 OQ818458 OQ818398 OQ851552 OQ842982
X. furcata Fr. Taiwan YMJ1121 OQ818459 OQ818399 OQ851553 OQ842983
X. furcata Fr. Taiwan YMJ1122 OQ818460 OQ818400 OQ851554 OQ842984
X. furcata Fr. Taiwan YMJ1123 OQ818461 OQ818401 OQ851555 OQ842985
X. furcata Fr. Taiwan YMJ1125 OQ818462 OQ818402 OQ851556 OQ842986
X. furcata Fr. Taiwan YMJ1126 OQ818463 OQ818403 OQ851557 OQ842987
X. furcata Fr. Taiwan YMJ1127 OQ818464 OQ818404 OQ851558 OQ842988
X. furcata Fr. Taiwan YMJ1128 OQ818465 OQ818405 OQ851559 OQ842989
X. furcata Fr. Taiwan YMJ1129 OQ818466 OQ818406 OQ851560 OQ842990
X. furcata Fr. Taiwan YMJ1132 OQ818467 OQ818407 OQ851561 OQ842991
X. furcata Fr. Taiwan YMJ1133 OQ818468 OQ818408 OQ851562 OQ842992
X. furcata Fr. Taiwan YMJ1134 OQ818469 OQ818409 OQ851563 OQ842993
X. furcata Fr. Taiwan YMJ1135 OQ818470 OQ818410 OQ851564 OQ842994
X. furcata Fr. Taiwan YMJ1730 from EPITYPE OQ818471 OQ818411 OQ851565 OQ842995
X. furcata Fr. Taiwan YMJ1731 from EPITYPE OQ818472 OQ818412 OQ851566 OQ842996
X. furcata Fr. Taiwan YMJ1732 from EPITYPE OQ818473 OQ818413 OQ851567 OQ842997
X. furcata Fr. Taiwan YMJ1736 from EPITYPE OQ818474 OQ818414 OQ851568 OQ842998
X. furcata Fr. Taiwan YMJ1737 from EPITYPE OQ818475 OQ818415 OQ851569 OQ842999
X. furcata Fr. Taiwan YMJ1738 from EPITYPE OQ818476 OQ818416 OQ851570 OQ843000
X. furcatula Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1099 from HOLOTYPE OQ818433 OQ818373 OQ851527 OQ845483
X. furcatula Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1103 from HOLOTYPE OQ818434 OQ818374 OQ851528 OQ845484
X. furcatula Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1916 OQ818435 OQ818375 OQ851529 OQ845485
X. furcatula Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1917 OQ818436 OQ818376 OQ851530 OQ845486
X. furcatula Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1918 OQ818437 OQ818377 OQ851531 OQ845487
X. furcatula Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ2194 OQ818438 OQ818378 OQ851532 OQ845488
X. hoehnelii Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ642 from HOLOTYPE, as X. sp. 1
in Hsieh et al. (2010)GQ502719 GQ853054 GQ853036 GU324759
X. hoehnelii Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ643 from HOLOTYPE OQ818439 OQ818379 OQ851533 OQ845489
X. hoehnelii Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ644 from HOLOTYPE OQ818440 OQ818380 OQ851534 OQ845490
X. hoehnelii Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ645 from HOLOTYPE OQ818441 OQ818381 OQ851535 OQ845491
X. hoehnelii Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1151 OQ818442 OQ818382 OQ851536 OQ845492
X. hoehnelii Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1159 OQ818443 OQ818383 OQ851537 OQ845493
X. hoehnelii Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ2178 OQ818444 OQ818384 OQ851538 OQ845494

Page 4 of 27
Juetal. Botanical Studies (2023) 64:21
Other sequences included in the phylogenetic analyses
were listed in Additional file1: TableS1.
e ITS dataset (Additional file2) was formed using
ITS sequences from X. furcata and resembling species
as well as the neotype of X. furcata, with X. hypoxylon
(L.) Grev. as the outgroup. e concatenated dataset
RPB2-TUB-ACT (Additional file 3) was formed using
sequences of α-ACT , RPB2, and β-TUB from X. furcata
and resembling species (Table1) as well as various spe-
cies of Xylaria and closely related genera of Xylaria
(Additional file1: TableS1), with Biscogniauxia arima
F. San Martín, Y.-M. Ju & J. D. Rogers as the outgroup.
Both datasets were aligned using Clustal X 1.81 (omp-
son etal. 1997) with the “gap penalty” set to 10 and “gap
extension penalty” set to 0.2, and improved manually.
To calculate the similarities between pairs of ITS
sequences from X. furcata and resembling species, we
used DNADIST from the PHYLIP version 3.6 phyloge-
netic inference package (Felsenstein 2005).
Maximum-Likelihood (ML) and Bayesian Inference
(BI) analyses were performed on the aligned ITS data-
set (Additional file2) and the aligned RPB2-TUB-ACT
dataset (Additional file3). ML trees were generated using
RAxML analysis ver. 8.2.10 (Stamatakis 2014) with rapid
bootstrap support and 1000 replicates of bootstrap test to
assess the robustness of the tree topology. BI trees were
generated using MrBayes ver. 3.2.6 (Ronquist etal. 2012)
with a Markov Chain Monte Carlo (MCMC) algorithm.
Four MCMC chains (one cold and three heated) were run
for one million generations with the trees sampled every
100 generations to ensure convergence of the chains.
e first 25% trees were excluded as burn-in phases, and
posterior probability values were estimated with the 75%
remaining trees to assess the support for each clade in
the tree. Models of evolution for both ML and BI trees
were defined by MrModeltest 2.4 (Nylander 2004), and
consensus trees were viewed in FigTree ver. 1.4.4 (http://
tree. bio. ed. ac. uk/ softw are/ figtr ee/).
Results
Concept ofX. furcata reconrmed throughITS sequence
analysis oftheneotype andcomparison ofITS similarities
betweenspecies pairs
e ITS sequences from the neotype of X. furcata and
specimens identifiable as X. furcata (Table1) were ana-
lyzed using BI and ML phylogenetic methods. Both anal-
yses generated largely congruent trees, and only the ML
tree is shown (Fig.1). e ITS sequence from the neotype
Table 1 (continued)
Taxon Origin Collecting data GenBank Accession number
β-tubulin gene α-actin gene RPB2 gene ITS
X. hoehnelii Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ2179 OQ818445 OQ818385 OQ851539 OQ845495
X. insignifurcata Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ650 from HOLOTYPE, as X. sp. 5
in Hsieh et al. (2010)GQ502716 GQ853051 GQ853033 GU324758
X. insignifurcata Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ649 from HOLOTYPE OQ818446 OQ818386 OQ851540 OQ845496
X. insignifurcata Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1198 OQ818447 OQ818387 OQ851541 OQ845497
X. scoparia Pat. Taiwan YMJ958 OQ818477 OQ818417 OQ851571 OQ843001
X. scoparia Pat. Taiwan YMJ959 – – – OQ843002
X. scoparia Pat. Taiwan YMJ960 OQ818478 OQ818418 OQ851572 OQ843003
X. scoparia Pat. Taiwan YMJ982 OQ818479 OQ818419 OQ851573 OQ843004
X. scoparia Pat. Taiwan YMJ983 OQ818480 OQ818420 OQ851574 OQ843005
X. scoparia Pat. Taiwan YMJ989 OQ818481 OQ818421 OQ851575 OQ843006
X. scoparia Pat. Taiwan YMJ991 OQ818482 OQ818422 OQ851576 OQ843007
X. scoparia Pat. Taiwan YMJ992 OQ818483 OQ818423 OQ851577 OQ843008
X. scoparia Pat. Taiwan YMJ1158 OQ818484 OQ818424 OQ851578 OQ843009
X. scoparia Pat. Taiwan YMJ1423 OQ818485 OQ818425 OQ851579 OQ843010
X. scoparia Pat. Taiwan YMJ1424 OQ818486 OQ818426 OQ851580 OQ843011
X. scoparia Pat. Taiwan YMJ1427 OQ818487 OQ818427 OQ851581 OQ843012
X. scoparia Pat. Taiwan YMJ1428 OQ818488 OQ818428 OQ851582 OQ843013
X. scoparia Pat. Taiwan YMJ1435 from EPITYPE OQ818489 OQ818429 OQ851583 OQ843014
X. scoparia Pat. Taiwan YMJ2177 OQ818490 OQ818430 OQ851584 OQ843015
X. siamensis Wangsawat et al. Thailand SWUF17-20.2 from HOLOTYPE (Wang‑
sawat et al. 2021)OQ845437 OQ845429 OQ851589 MT622765
X. tenellifurcata Y.‑M. Ju & H.‑M. Hsieh Taiwan YMJ1070 from HOLOTYPE OQ818448 OQ818388 OQ851542 OQ845498

Page 5 of 27
Juetal. Botanical Studies (2023) 64:21
of X. furcata matched sequences from cultures obtained
from stromata of nine specimens, including YMJ646 of
specimen 95072001, YMJ990, 993 & 994 of specimen
97071501, YMJ978 of specimen 97072705, YMJ1101 of
specimen 98070101, YMJ1120, 1122 & 1123 of speci-
men 98072301, YMJ1125–1129 of specimen 98072401,
YMJ956 & 1135 of specimen 98073103, YMJ1132–1134
of specimen 98080301, and YMJ1730–1732 and 1736–
1738 of specimen 104072801. However, slight differences
(> 99.6% similarity) were observed with sequences from
three of the nine stromata, including YMJ979 of speci-
men 97072705, YMJ1100 & 1102 of specimen 98070101,
and YMJ1121 of specimen 98072301. Specimen
104072801 contains ample material and is designated as
the epitype of X. furcata herein. A packet attached on the
same herbarium sheet as the neotype from FH contain-
ing immature stromata emerging from a fungus comb
was also confirmed to be X. furcata.
e ITS similarities between X. furcata and resembling
species are shown in Table2. e species most similar
to X. furcata is X. brevifurcata with a similarity range of
96.5–97.0%, followed by X. tenellifurcata with a similar-
ity range of 94.1–94.3%, and X. scoparia with a similarity
range of 91.2–91.5%. e similarities between other pairs
of species are less than 93.80%, which is the similarity
shared between X. brevifurcata and X. tenellifurcata.
Fig. 1 Phylogenetic tree generated by ML analysis from the ITS dataset with the sequence from the neotype of X. furcata included. The
species newly described in the present study are in boldface. Numbers at internodes represent bootstrap values and are immediately followed
by the posterior probability values greater than 50% with BI analysis. Xylaria hypoxylon is the outgroup taxon
Table 2 Comparison of ITS similarities between species pairs of X. furcata and resembling species
Taxon X. brevifurcata
(%) X. furcata (%) X. furcatula
(%) X. hoehnelii
(%) X.
insignifurcata
(%)
X. scoparia
(%) X. siamensis
(%) X.
tenellifurcata
(%)
X. brevifurcata 100
X. furcata 96.5–97.0 99.6–100
X. furcatula 84.0–84.2 84.2–84.6 99.8–100
X. hoehnelii 83.7 83.1–83.5 89.3–89.7 99.8–100
X. insignifurcata 85.6 85.2–85.4 86.9–87.1 86.9–87.1 100
X. scoparia 91.4 91.2–91.5 83.2–83.4 83.2 84.5 99.8–100
X. siamensis 87.7 88.3–88.5 83.3–83.5 84.0–84.2 85.8 87.2 100
X. tenellifurcata 93.8 94.1–94.3 83.5–83.8 84.2–84.4 85.2 90.3 89.1 100

Page 6 of 27
Juetal. Botanical Studies (2023) 64:21
Molecular phylogenetic analyses based
ontheRPB2-TUB-ACT dataset
BI and ML analyses based on the RPB2-TUB-ACT
dataset were congruent and only the ML tree is pre-
sented (Fig.2). ese trees showed that X. furcata and
resembling species formed a clade with other Xylaria
species associated with termite nests and soil. is clade
corresponds with the TE clade in Hsieh etal. (2010) and
U’Ren et al. (2016) and labeled as such in Fig. 2. e
TE clade was divided into two major subclades: the X.
Fig. 2 Phylogenetic tree generated by ML analysis from the RPB2‑TUB‑ACT dataset. The subclade of the TE clade including X. furcata
and resembling species is in shade. The species newly described in the present study are in boldface. Numbers at internodes represent bootstrap
values and are immediately followed by the posterior probability values greater than 50% with BI analysis. Biscogniauxia arima is the outgroup taxon

Page 7 of 27
Juetal. Botanical Studies (2023) 64:21
furcata subclade, to which X. furcata and resembling
species belonged, and the X. nigripes subclade, to which
sclerotium-forming species (Ju et al. 2022) belonged.
Within the X. furcata subclade, X. furcata and resembling
species belonged to two clusters: the X. furcata cluster,
grouped with X. insolita Y.-M. Ju, H.-M. Hsieh & J.-C.
Chou and X. ochraceostroma Y.-M. Ju & H.-M. Hsieh in
one branch, and the X. hoehnelii cluster, grouped with
X. intraflava Y.-M. Ju & H.-M. Hsieh and subintraflava
Wangsawat, Y.-M. Ju, Phosri, Whalley & Suwannasai in
the other branch. e species of the X. furcata cluster,
including X. brevifurcata, X. furcata, X. insignifurcata,
Fig. 2 continued

Page 8 of 27
Juetal. Botanical Studies (2023) 64:21
X. tenellifurcata, X. scoparia, and X. siamensis, are char-
acterized by half-exposed to fully exposed perithecial
mounds on the stromatal surface, while those of the X.
hoehnelii cluster, including X. furcatula and X. hoehnelii,
are characterized by inconspicuous perithecial mounds.
Taxonomy
Description ofX. furcata
Xylaria furcata Fr., Nov. Act. Reg. Soc. Sci. Upsal., ser.
III, 1: 128. 1851; non Schwein. ex Berk. & M. A. Curtis.
Figs.3–4
≡ Sphaeria dichotoma Lév., Ann. Sci. Nat., Bot., sér. III,
3: 45. 1845; non Xylaria dichotoma (Mont.) Fr., 1851.
≡ Xylosphaera furcata (Fr.) Dennis, Bull. Jardin Botan.
État Brux. 31: 116. 1961.
≡ Podosordaria furcata (Fr.) P. M. D. Martin, J. S. Afri-
can Bot. 36: 131. 1970, nom. inval. ICN Art. 41.1 (Shenz-
hen Code); J. S. African Bot. 42: 79. 1976.
Typification INDONESIA. Java, Buitenzorg, on termite
nests, 1907–1908, von Höhnel, F. A4371 (neotype [redes-
ignated here, MycoBank Typification No. 10013825] of X.
furcata FH ex von Höhnel herb., isoneotypes S F57929 ex
Bresadola herb., FH ex von Höhnel herb. [in 3 packets],
K[M] ex von Höhnel herb.); Java, Buitenzorg, on termite
nests, 1908, von Höhnel, F., Rehm’s Ascomyceten 1812
(isoneotype of X. furcata S F57927 ex Rehm herb., K[M]).
TAIWAN. Tainan City, Hsin-shih District, Tan-ting,
on ground of mango orchard, 28 Jul 2015, Chou, K.-H.
104072801 (cultured from stromata YMJ1730, YMJ1731,
YMJ1732, YMJ1736, YMJ1737 & YMJ1738) (epitype [des-
ignated here, MycoBank Typification No. 10013826] of X.
furcata HAST 145909).
Stromata antler-like at fertile part, dichotomously
branched one to three times, with acuminate apices,
on a glabrous stipe, 2–3.5cm long above ground, 1.1–
1.7 cm long × 1–1.5 mm diam at fertile part; surface
dark brown, becoming black, with half-exposed to fully
exposed perithecial mounds but less so with close-set
perithecia, lacking an outer layer, underlain with a thin,
black layer ca. 10 µm thick; interior white, homogene-
ous, soft. Perithecia spherical, 150–250µm diam. Osti-
oles conical, 60–80µm high × 80–110µm broad at base.
Asci with eight ascospores arranged in uniseriate man-
ner, cylindrical, 55–75µm total length, the spore-bearing
part 25–40µm long × 3.5–4.5 µm broad, with an apical
ring staining blue in Melzer’s iodine reagent, inverted
hat-shaped, 1–1.5µm high × 1.5µm broad. Ascospores
light brown to brown, unicellular, short fusoid-inequi-
lateral, slightly laterally compressed, with narrowly
rounded ends, smooth, (3.5–)4–5.5(–6) × (2–)2.5–3 µm
(4.7 ± 0.6 × 2.6 ± 0.3 µm, N = 80), with a straight germ slit
spore-length or nearly so on the ventral side, lacking a
hyaline sheath; epispore smooth.
Cultures and anamorph: Colonies reaching the edge
of 9-cm Petri dish in 4 wk, whitish, mostly submerged,
azonate, with diffuse margins. Reverse uncolored.
Stromata arising copiously from the entire colonies,
antler-like, branched several times, up to 5cm long × 0.4–
0.6mm diam, white, immediately becoming black from
base upwards, overlain with smoke gray pustules on the
entire surface due to production of conidia. Anamo-
rph produced on the stromatal surface. Conidiophores
upright, mononematous; main axis unbranched, 200–
300 × 5–8µm, dichotomously branched two to five times
on top, smooth, hyaline. Conidiogenous cells 2–3 born
on each terminal short branch, initially ampulliform,
becoming cylindrical, closely geniculate at upper end
after producing multiple conidia in sympodial sequence,
8–12.5 × 3–4 µm, smooth, bearing terminal poroid
conidial secession scars. Conidia produced holoblasti-
cally, hyaline, smooth, subglobose to ellipsoid, (4.5–)
5–6.5(–7) × (3.5–)4–5(–5.5) µm (5.7 ± 0.6 × 4.4 ± 0.4 µm,
N = 40), with a flattened base indicating former point of
attachment to conidiogenous cell.
Additional specimens examined CHINA. Hainan,
Ting-an, on ground, Sep 1934, Deng, S. Q. 4219 (BPI
584600, BPI 584828, BPI 584607); Kiangsi, Tehsin, on
ground, 20 May 1935, Deng, S. Q. 9521 (BPI 584603),
mixed with X. scoparia; Kiangsi, Tehsin, on ground, 20
May 1935, Deng, S. Q. 9523 (BPI 584602, BPI 584829);
Kiangsi, Tehsin, on ground, 20 May 1935, Deng, S. Q.
9524 (BPI 584606); Nanking, Ling-ku-sze Woods, on
ground, 28 Jun 1936, Shen, H. N. 338 (BPI 584831); Nan-
king, Ling-ku-sze Woods, on ground, 13 Jun 1937, Teng,
S. C. 2743 (BPI 584601). TAIWAN. Tainan City, Hsin-
shih District, Tan-ting, on ground of bamboo plantation,
1 Jul 2009, Chou, K.-H. 98070101 (cultured from stro-
mata YMJ1100, YMJ1101 & YMJ1102) (HAST 145906);
Tainan City, Hsin-shih District, Tan-ting, on ground of
bamboo plantation, 24 Jul 2009, Chou, K.-H. 98072401
(cultured from stromata YMJ1125, YMJ1126, YMJ1127,
YMJ1128 & YMJ1129) (HAST 145907); Tainan City,
Hsin-shih District, Tan-ting, on ground of bamboo
plantation, 3 Aug 2009, Chou, K.-H. 98080301 (cultured
from stromata YMJ1132, YMJ1133 & YMJ1134) (HAST
145908); Tainan City, Hsin-shih District, Tan-ting, on
ground of mango orchard, 23 Jul 2009, Chou, K.-H.
98072301 (cultured from stromata YMJ1120, YMJ1121,
YMJ1122 & YMJ1123) (HAST 145910); Tainan City,
Shen-hua District, Liu-fen-liao, from fungus comb below
a Termitomyces mushroom, 31 Jul 2009, Chou, K.-H.
98073103 (cultured from stromata YMJ956 & YMJ1135)
(HAST 145947), culture only; Tainan City, Shen-hua

Page 9 of 27
Juetal. Botanical Studies (2023) 64:21
District, Liu-fen-liao, on ground of bamboo plantation,
27 Jul 2008, Chou, K.-H. 97072705 (cultured from stro-
mata YMJ978 & YMJ979) (HAST 145911), immature,
with anamorph only; Tainan City, Shen-hua District,
Niu-Chuang, on ground under Cordia dichotoma (Bor-
aginaceae), 15 Jul 2008, Chou, K.-H. 97071501 (cultured
from stromata YMJ990, YMJ993 & YMJ994) (HAST
145926), immature; Tainan City, Shen-hua District,
Fig. 3 Xylaria furcata (A–E from neotype, F–L from epitype). A, F Stromata. B, C, G, H. Stromatal surfaces. D, I Ascal apical rings. E, J Ascospores. K, L
Colony on 9‑cm Petri plate containing OA at 2 and 4 wk, respectively. Bars in F = 1 cm; A = 5 mm; B, C, G, H = 0.25 mm; D, E, I, J = 5 µm

Page 10 of 27
Juetal. Botanical Studies (2023) 64:21
Fig. 4 Xylaria furcata (from epitype). A Stromata produced in culture. B, C Stromatal surface bearing upright conidiophores. D–F Conidiophores.
G–I Conidiogenous cells. J Conidia. Bars in A = 1 mm; B, C = 100 μm; D = 20 μm; E, F = 10 μm; G–J = 5 µm

Page 11 of 27
Juetal. Botanical Studies (2023) 64:21
vicinity of Asian Vegetable Research and Development
Center, on termite fungus combs, 20 Jul 2006, Chou,
K.-H. 95072001 (cultured from stroma YMJ646) (HAST
145912), anamorph emerging from fungus combs, as X.
sp. 4 in Hsieh etal. (2010).
Notes Xylaria furcata is characterized by its delicate, gla-
brous stromata that branch dichotomously one to three
times. e stromata are topped with acuminate apices,
and roughened with partially or fully exposed perithe-
cial mounds on the fertile parts when fully mature. e
perithecial mounds are less evident where perithecia are
close-set. e mature stromatal surface is initially brown
but gradually blackens with time. is species is found
only in Asia thus far. Other species included in this study
can be easily misidentified as X. furcata or have been
confused with it. ese species can be separated from
X. furcata based on morphological features of the tele-
omorphs and anamorphs as outlined in the identification
key herein.
As mentioned above, the original material of X. fur-
cata seems to be missing. As a result, Rogers etal. (2005)
neotypified the name with a part of a von Höhnel collec-
tion made from Java, Indonesia. Five packets of the von
Höhnel collection can be located at FH and are attached
on the sheet 7472 in three rows, with one, two, and two
packets in the top, middle, and bottom rows, respectively.
e packet on the left in the middle row was designated
by Rogers etal. (2005) as the neotype for X. furcata but
contains three different Xylaria species. e material, on
which the above description of X. furcata is based, is now
redesignated as the neotype while the remaining material
belongs to two new species: X. hoehnelii and X. robusti-
furcata. e corresponding material in the packet in the
top row, right packet in the middle row, and right packet
in the bottom row are considered isoneotypes of X. fur-
cata while the remaining material belongs to X. hoehnelii
and/or X. robustifurcata. e left packet in the bottom
row contains long stromata emerging from an incubated
fungus comb shown in Plate IV of von Höhnel (1908),
with an ITS sequence slightly different from that of the
neotype (Fig.1). Other parts of the von Höhnel collection
were distributed as no. 1812 of Rehm’s exsiccati Ascomy-
ceten. Its duplicates at S and K contain both X. furcata
and X. hoehnelii, with the X. furcata stromata considered
isoneotypes while a duplicate at HBG contains X. hoeh-
nelii and X. robustifurcata but not X. furcata.
Dennis (1961) reported X. furcata from Congo. How-
ever, in a footnote on page 118, he mentioned that the
interpretation of X. furcata in Teng (1934) seemed dif-
ferent from his own. We have examined nine Chinese
specimens that Teng identified as X. furcata from BPI
and reconfirmed his identifications. e Congo specimen
that Dennis (1961) considered to be X. furcata is actually
X. hirsuta.
Xylaria cf. furcata reported from New Zealand by Rog-
ers and Samuels (1986) has ascospores measuring (7.0–)
7.5–8.5(–9.0) × 3.5–4.0(–4.5) μm, which are substantially
larger than those of X. furcata and resembling species. It
may represent an undescribed species of subg. Pseudox-
ylaria. However, it remains uncertain as to whether the
New Zealand collection is associated with soil or wood
buried in soil (Rogers and Samuels 1986).
Species resembling X. furcata
Xylaria brevifurcata Y.-M. Ju & H.-M. Hsieh, sp. nov.
Fig.5
MycoBank MB849229.
Typification. TAIWAN Hua-lien County, Shou-feng
Township, campus of National Dong Hwa University, on
ground, 26 Jun 2011, Chou, J.-C. 100062601 (cultured
from stroma YMJ1381) (holotype of X. brevifurcata
HAST 145904).
Etymology Referring to the short furcate stromata.
Stromata antler-like at fertile part, dichotomously
branched one to several times, with acuminate api-
ces, on a glabrous stipe, 0.9–1.5cm long above ground,
0.4–0.6cm long × 1.1–2.3 mm diam at fertile part; sur-
face brown to dark brown except for black perithecial
mounds, more light-colored at apices, with half-exposed
to fully exposed perithecial mounds, lacking an outer
layer, underlain with a thin, black layer ca. 10µm thick;
interior white, homogeneous, soft. Perithecia spherical,
150–250 µm diam. Ostioles conic-papillate, ca. 40 µm
high × ca. 50µm broad at base. Asci with eight ascospores
arranged in uniseriate manner, cylindrical, 55–80 µm
total length, the spore-bearing part 25–35µm long × 3.5–
4.5µm broad, with an apical ring staining light blue in
Melzer’s iodine reagent, inverted hat-shaped, 1.5–2µm
high × 1.5–2µm broad. Ascospores brown to dark brown,
unicellular, short fusoid-inequilateral, slightly later-
ally compressed, with narrowly rounded ends, some-
times slightly pinched on one end or both ends, smooth,
(4–)4.5–5(–5.5) × 2.5–3 µm (5.0 ± 0.2 × 2.6 ± 0.2 µm,
N = 40), with a straight germ slit spore-length or nearly
so on the ventral side, lacking a hyaline sheath; epispore
smooth.
Cultures and anamorph. Colonies reaching the edge
of 9-cm Petri dish in 4 wk, white, submerged, becom-
ing cottony at places, azonate, with diffuse margins.
Reverse uncolored. Stromata not produced or sporadi-
cally produced, cylindrical, tapering upwards, dichot-
omously branched one to several times, up to 3 cm
long × 0.4–0.9mm diam, white. Anamorph produced on

Page 12 of 27
Juetal. Botanical Studies (2023) 64:21
Fig. 5 Xylaria brevifurcata (from holotype). A Stromata. B Stromatal surface. C Ascal apical rings. D Ascospores. E, F Colony on 9‑cm Petri plate
containing OA at 2 and 4 wk, respectively. G, H Conidiophores. I Conidiogenous cells. J Conidia. Bars in A = 5 mm; B = 0.25 mm; G = 25 μm;
H = 10 μm; C, D, I, J = 5 µm

Page 13 of 27
Juetal. Botanical Studies (2023) 64:21
the colony surface. Conidiophores upright, mononema-
tous, 5–6 µm broad at base, dichotomously branched
several times from base, smooth, hyaline. Conidioge-
nous cells 2–3 born on each terminal short branch, ini-
tially ampulliform, becoming cylindrical, forming one
to several consecutive nodulose swellings at upper end
after producing multiple conidia in sympodial sequence,
9–15 × 3.5–4µm, smooth, bearing terminal poroid conid-
ial secession scars. Conidia produced holoblastically,
hyaline, smooth, highly variable in shape, subglobose,
obovoid, ellipsoid to oblong, equilateral or slightly to sig-
nificantly oblique, (4.5–)5.5–8.5(–11) × (3–)3.5–4.5(–5)
µm (7.0 ± 1.6 × 4.0 ± 0.3µm, N = 40), with a flattened base
indicating former point of attachment to conidiogenous
cell.
Additional specimen examined TAIWAN. Tainan
City, Hsin-shih District, Tan-ting, on ground of mango
orchard, 28 Jul 2008, Chou, K.-H. 97072805 (cultured
from stroma YMJ986) (HAST 145905), immature, with
anamorph and developing perithecia.
Notes Xylaria brevifurcata resembles a smaller version of
X. furcata, with stromata terminating into acuminate api-
ces and perithecial mounds that are half-exposed to fully
exposed. Stromata are infrequently produced in culture
but remain sterile. Conidia are produced over the entire
colony surface and vary greatly in shape.
Xylaria furcatula Y.-M. Ju & H.-M. Hsieh, sp. nov. Fig.6
MycoBank MB849230.
Typification TAIWAN. Tainan City, Hsin-shih District,
Tan-ting, on ground of bamboo plantation, 1 Jul 2009,
Chou, K.-H. 98070105 (cultured from stromata YMJ1099
& YMJ1103) (holotype of X. furcatula HAST 145914).
Etymology Referring to the short, furcate terminal
branchlets of stromata.
Stromata antler-like at fertile part, dichotomously
branched one to two times and further branched
at sterile apices, with delicate acuminate or forked
apices, on a glabrous stipe, 2.1–2.4 cm long above
ground, 1–1.2 cm long × 1.2–1.7 mm diam at fertile
part; surface tan-colored to brown except for sur-
rounding areas of ostioles, with inconspicuous peri-
thecial mounds, lacking an outer layer, underlain with
a thin layer concolorous with the surface, ca. 10µm
thick; interior white, homogeneous, soft. Perithe-
cia spherical, 150–200µm diam. Ostioles conical, ca.
50µm high × ca. 50µm broad at base. Asci with eight
ascospores arranged in uniseriate manner, cylindri-
cal, 70–90 µm total length, the spore-bearing part
35–45µm long × 3.5–4.5µm broad, with an apical ring
staining blue in Melzer’s iodine reagent, inverted hat-
shaped, 1.5–2 µm high × 1.5 µm broad. Ascospores
brown, unicellular, ellipsoid-inequilateral, with nar-
rowly rounded ends, smooth, 5.5–6(–6.5) × 3–3.5 µm
(5.8 ± 0.3 × 3.1 ± 0.3 µm, N = 40), with a straight germ
slit spore-length or nearly so on the ventral side, lack-
ing a hyaline sheath; epispore smooth.
Cultures and anamorph. Colonies reaching the edge
of 9-cm Petri dish in 4 wk, whitish, mostly submerged,
azonate, with diffuse margins. Reverse uncolored or pale
tan-colored. Stromata arising copiously from the entire
colonies, antler-like, branched several times, up to 5cm
long × 0.4–0.6mm diam, white, immediately becoming
black from base upwards, overlain with smoke gray pus-
tules on the entire surface due to production of conidia.
Anamorph produced on the stromatal surface. Conidi-
ophores upright, mononematous; main axis unbranched,
100–300 × 4.5–6µm, alternately branched several times
close to top, smooth, hyaline. Conidiogenous cells formed
singly or in whirls of 2–3 along each terminal branch, ini-
tially ampulliform, becoming cylindrical, slightly swollen
at upper end after producing multiple conidia in sympo-
dial sequence, 5.5–11 × 2.5–3.5µm, smooth, bearing ter-
minal poroid conidial secession scars. Conidia produced
holoblastically, hyaline, smooth, subglobose to obovoid,
3.5–5(–6) × 3–4µm (4.2 ± 0.6 × 3.5 ± 0.3µm, N = 40), with
a flattened base indicating former point of attachment to
conidiogenous cell.
Additional specimens examined TAIWAN. Tainan
City, Hsin-shih District, Tan-ting, on ground, 3 Jul 2017,
Chou, K.-H. 106070302 (cultured from stroma YMJ2194)
(HAST 145913); Tainan City, Shen-hua District, Niu-
Chuang, on ground under Cordia dichotoma (Boragi-
naceae), 16 Aug 2018, Chou, K.-H. 107081602 (cultured
from stromata YMJ1916, YMJ1917 & YMJ1918) (HAST
145915).
Notes Stromata of X. furcatula are characterized by hav-
ing short, dichotomous sterile branchlets on top, a tan to
brown surface, and inconspicuous perithecial mounds.
Xylaria hirsuta (J. D. Rogers & Y.-M. Ju) Y.-M. Ju &
H.-M. Hsieh, comb. nov. Fig.7A–D
MycoBank MB849231.
≡ Xylaria furcata Fr. var. hirsuta J. D. Rogers & Y.-M. Ju
in J. D. Rogers, Y.-M. Ju & Lehmann, Mycologia 97: 919.
2005.
Typification DEMOCRATIC REPUBLIC OF CONGO.
Yangambi, Isalowe, altitude 470 m, on soil, May 1939,

Page 14 of 27
Juetal. Botanical Studies (2023) 64:21
Fig. 6 Xylaria furcatula (from holotype). A Stromata. B Stromatal surface. C Ascal apical ring. D Ascospores. E, F Colony on 9‑cm Petri plate
containing OA at 2 and 4 wk, respectively. G Stromatal surface bearing upright conidiophores. H, I Conidiophores. J, K Conidiogenous cells. L
Conidia. Bars in A = 5 mm; B = 0.25 mm; G = 25 μm; H = 20 μm; I = 10 μm; C, D, J–L = 5 µm

Page 15 of 27
Juetal. Botanical Studies (2023) 64:21
Louis, F. 14866, as X. furcata Fr. by Dennis, R. W. G. (hol-
otype of X. furcata var. hirsuta K[M] 125997).
Stromata antler-like at fertile part, highly dichoto-
mously branched, with long acicular apices, on a tomen-
tose stipe, 5.2cm long above ground, 3cm long × 1.5mm
diam at fertile part; surface blackish brown, with fully
exposed perithecial mounds, overlain with a dark
brown tomentum, underlain with a thin, black layer ca.
10 µm thick; interior white, homogeneous, soft. Peri-
thecia spherical, 200–250µm diam. Ostioles conical, ca.
60µm high × ca. 100µm broad at base. Asci with eight
ascospores arranged in uniseriate manner, cylindrical,
45–65µm total length, the spore-bearing part 25–35µm
long × 3–4µm broad, with an apical ring staining light
Fig. 7 Xylaria hirsuta and X. robustifurcata. A–D X. hirsuta (from holotype). A Stroma. B Stromatal surface. C Ascal apical ring. D Ascospores. E–I X.
robustifurcata (from holotype except for I, which is from the holotype of X. nigripes var. trifida). E, I. Stromata. F Stromatal surface. G Ascal apical ring.
H Ascospores. Bars in A = 2.5 mm; B = 0.5 mm; E = 5 mm; F, I = 0.25 mm; C, D, G, H = 5 µm

Page 16 of 27
Juetal. Botanical Studies (2023) 64:21
blue in Melzer’s iodine reagent, inverted hat-shaped,
1–1.5 µm high × 1–1.5 µm broad. Ascospores light
brown to brown, unicellular, short fusoid, inequilateral
to slightly inequilateral, with narrowly rounded ends,
smooth, 4–5 × 2–2.5 µm (4.5 ± 0.3 × 2.3 ± 0.2 µm, N = 40),
with a straight germ slit spore-length or nearly so on the
ventral side, lacking a hyaline sheath; epispore smooth.
Cultures and anamorph. Unknown.
Notes Xylaria hirsuta is known only from its holotype,
which contains a stroma repeatedly dichotomously
branched up to seven times. It was collected from Afri-
can, featuring a hirsute stromatal surface roughened with
fully exposed perithecial mounds.
Xylaria hoehnelii Y.-M. Ju & H.-M. Hsieh, sp. nov.Fig.8
MycoBank MB849233.
Typification TAIWAN. Tainan City, Hsin-shih District,
Tan-ting, on ground of mango orchard, 20 May 2006, Ju,
Y.-M. & Hsieh, H.-M. 95052006 (cultured from stromata
YMJ642, YMJ643, YMJ644 & YMJ645) (holotype of X.
hoehnelii HAST 145916), as Xylaria sp. 1 in Hsieh etal.
(2010).
Etymology In honor of the Austrian mycologist Franz
Xaver Rudolf von Höhnel for his contribution to X. fur-
cata and similar fungi.
Stromata antler-like at fertile part, dichotomously
branched one to three times, with delicate acuminate
or forked apices, on a glabrous stipe, 2–3.5 cm long
above ground, 1.5–1.9 cm long × 1.5–2.1 mm diam at
fertile parts; surface tan-colored to brown except for
surrounding areas of ostioles, with inconspicuous peri-
thecial mounds, lacking an outer layer, underlain with a
thin layer concolorous with the surface, ca. 10µm thick;
interior white, homogeneous, soft. Perithecia spherical
to obovoid, 300–400µm diam × 400–500µm high. Osti-
oles conical, 60–100µm high × 60–80µm broad at base.
Asci with eight ascospores arranged in uniseriate man-
ner, cylindrical, 40–65µm total length, the spore-bearing
part 25–35µm long × 3.5–4.5 µm broad, with an apical
ring staining blue in Melzer’s iodine reagent, inverted
hat-shaped, 1.5 µm high × 1.5 µm broad. Ascospores
brown to dark brown, unicellular, ellipsoid-inequilateral,
with narrowly rounded ends, smooth, 4–5 × 2.5–3 µm
(4.5 ± 0.3 × 2.7 ± 0.2 µm, N = 40), with a straight germ slit
spore-length or nearly so on the ventral side, lacking a
hyaline sheath; epispore smooth.
Cultures and anamorph. Colonies reaching the edge of
9-cm Petri dish in 3 wk, white, submerged, immediately
becoming tufted over the entire surface, azonate, with
fimbriated margins. Reverse pale tan-colored. Stromata
arising copiously from the entire colonies, antler-like,
branched several times, up to 5 cm long × 0.4–0.9 mm
diam, white, immediately becoming black from base
upwards, overlain with smoke gray pustules on the
entire surface due to production of conidia. Anamo-
rph produced on the stromatal surface and hyphal
strands. Conidiophores upright, mononematous; main
axis unbranched, 120–200 × 4.5–6 µm, dichotomously
branched two to five times on top, smooth, hyaline.
Conidiogenous cells 2–3 born on each terminal short
branch, initially ampulliform, hyaline, becoming cylin-
drical and yellowish, slightly swollen at upper end after
producing multiple conidia in sympodial sequence,
5–7.5 × 2.5–3 µm, smooth, bearing terminal poroid
conidial secession scars. Conidia produced holoblasti-
cally, hyaline, smooth, subglobose to obovoid, (3–)3.5–
4(–4.5) × (2.5–)3–3.5µm (3.9 ± 0.3 × 3.3 ± 0.2µm, N = 40),
with a flattened base indicating former point of attach-
ment to conidiogenous cell.
Additional specimens examined INDONESIA. Java,
Buitenzorg, on termite nests, 1907–1908, von Höhnel, F.
A4371 (FH ex von Höhnel herb.); Java, Buitenzorg, Bot.
Garten, on termite nests, 1908, von Höhnel, F., Rehm’s
Ascomyceten 1812, as X. furcata (S F57927a, CWU [Myc]
AS716, BPI 584832, PC); Java, Mt. Salak, on ground?,
Zollinger, H., Planta Javanica 857a, as Sphaeria gracil-
lima (L 910.250–1438), immature. TAIWAN. Tainan
City, Hsin-shih District, Tan-ting, on ground of mango
orchard, 2 Oct 2009, Chou, K.-H. 98100204 (cultured
from stroma YMJ1159) (HAST 145948), immature;
Tainan City, Hsin-shih District, Tan-ting, on ground of
mango orchard, 2 Nov 2009, Chou, K.-H. 98111801 (cul-
tured from stroma YMJ1151) (HAST 145949); Tainan
City, Shen-hua District, Tung-shi, on ground, 25 Jun
2019, Chou, K.-H. 108062508 (cultured from stromata
YMJ2178 & YMJ2179) (HAST 145950).
Notes Among X. furcata and resembling species, X.
hoehnelii and X. furcatula are the only two with a tan to
brown stromatal surface and were shown to be closely
related in our phylogenetic analyses (Figs.1, 2). ey also
share similar colony features, conidial sizes, and Tricho-
derma-like conidiophore branching patterns. Xylaria
hoehnelii differs from X. furcatula by having larger peri-
thecia, 300–400 µm diam × 400–500µm high vs. 150–
200µm diam, and smaller ascospores, 4–5 × 2.5–3µm v s.
5.5–6(–6.5) × 3–3.5 µm. Numerous immature stromata
of X. hoehnelii are commonly found fruiting from fungus
combs excavated from the field in Taiwan. e holotype
of X. hoehnelii was referred to as Xylaria sp. 1 in Hsieh
etal. (2010).

Page 17 of 27
Juetal. Botanical Studies (2023) 64:21
Fig. 8 Xylaria hoehnelii (from holotype except for B, which is from von Höhnel A4371 [FH]). A, B Stromata. C Stromatal surface. D Ascal apical ring.
E Ascospores. F, G Colony on 9‑cm Petri plate containing OA at 2 and 4 wk, respectively. H Stromatal surface bearing upright conidiophores. I, J
Conidiophores. K Conidiogenous cell bearing a conidium. L Conidia. Bars in A, B = 1 cm; C, H = 1 mm; I = 25 μm; D, E, J, L = 5 µm; K = 2 μm

Page 18 of 27
Juetal. Botanical Studies (2023) 64:21
Xylaria insignifurcata Y.-M. Ju & H.-M. Hsieh, sp. nov.
Fig.9
MycoBank MB849234.
Typification TAIWAN. Tainan City, Hsin-shih District,
Tan-ting, on ground of bamboo plantation, 12 Jul 2006,
Chou, K.-H. 95071201 (cultured from stromata YMJ649
& YMJ650) (holotype of X. insignifurcata HAST 145918),
as Xylaria sp. 5 in Hsieh etal. (2010).
Etymology A remarkable furcate Xylaria species for
its anamorph superficially resembling an Aspergillus
species.
Stromata antler-like at fertile part, dichotomously
branched one to three times, with acuminate apices, on
a glabrous stipe, 5–9.6cm long above ground, 1–2.4cm
long × 3.6–4.8 mm diam at fertile part; surface dark
brown to blackish brown, with half-exposed perithecial
mounds, lacking an outer layer, underlain with a thin,
black layer ca. 10µm thick; interior white, homogeneous,
soft. Perithecia spherical, 300–400 µm diam. Ostioles
papillate, ca. 50µm high × ca. 120µm broad at base. Asci
with eight ascospores arranged in uniseriate manner,
cylindrical, 75–95µm total length, the spore-bearing part
30–50µm long × 3.5–4.5 µm broad, with an apical ring
staining light blue in Melzer’s iodine reagent, inverted
hat-shaped, 1.5µm high × 1–1.5µm broad. Ascospores
brown to dark brown, unicellular, ellipsoid-inequilat-
eral, with narrowly rounded ends, smooth, (4–)4.5–
5.5 × 2–2.5(–3) µm (5.0 ± 0.3 × 2.4 ± 0.2 µm, N = 40), with
a straight germ slit spore-length or nearly so on the ven-
tral side, lacking a hyaline sheath; epispore smooth.
Cultures and anamorph. Colonies reaching the edge of
9-cm Petri dish in 8 wk, white initially, becoming black-
ish immediately behind the growing margins, velvety,
appressed, zonate, with diffuse margins. Reverse tan-
colored. Stromata arising from concentric zones and
elongating much farther beyond the colonies, cylindrical,
tapering upwards, unbranched or branched once or twice
closer to the apex, up to 6cm long × 0.9–1.2 mm diam,
white, immediately becoming black from base upwards,
overlain with pale mouse gray pustules on the entire sur-
face due to production of conidia. Anamorph produced
on the stromatal surface. Conidiophores upright, monon-
ematous; main axis unbranched or branched once close
to top, 150–250 × 7–9µm, dichotomously branched three
to four times in short intervals on top, smooth, yellow-
ish to light brown. Conidiogenous cells 2–3 born on each
terminal short branch, initially ampulliform, becoming
cylindrical, unswollen at upper end after producing mul-
tiple conidia in sympodial sequence, 5–7.5 × 2.5–3 µm,
smooth, bearing terminal poroid conidial secession scars.
Conidia produced holoblastically, hyaline, smooth, sub-
globose to obovoid, (4.5–)5–6(–7) × (4–)4.5–5(–5.5) µm
(5.5 ± 0.6 × 4.7 ± 0.3 µm, N = 40), with a flattened base
indicating former point of attachment to conidiogenous
cell.
Additional specimens examined TAIWAN. Tainan
City, Shen-hua District, Liu-fen-liao, on ground of bam-
boo plantation, 3 Jul 2006, Chou, K.-H. 95070301 (HAST
145919), immature, with anamorph and developing peri-
thecia; Tainan City, Shen-hua District, Liu-fen-liao, on
ground of bamboo plantation, 28 Jun 2010, Chou, K.-H.
99062803 (cultured from stroma YMJ1198) (HAST
145920), immature, with anamorph only.
Notes Among X. furcata and resembling species, X. insig-
nifurcata stands out for its larger stromata and Aspergil-
lus-like conidiophores. It was referred to as Xylaria sp. 5
in Hsieh etal. (2010). e teleomorph of this species was
found in only one stroma, where the perithecia are sparse
and scattered.
Xylaria robustifurcata Y.-M. Ju & H.-M. Hsieh, sp. nov.
Fig. 7E–I
= Xylaria nigripes (Klotzsch) M. C. Cooke var. trifida
Pat., J. Bot. (Morot) 5: 317. 1891.
MycoBank MB849235.
Typification INDONESIA. Java, Buitenzorg, on termite
nests, 1907–1908, von Höhnel, F. A4371, as X. furcata Fr.
(holotype of X. robustifurcata FH ex von Höhnel herb.,
isotypes S F57928 ex Rehm herb., S F57929a ex Bresadola
herb.); Java, Buitenzorg, Bot. Garten, on termite nests,
1908, von Höhnel, F., Rehm’s Ascomyceten 1812, as X. fur-
cata (isotypes of X. robustifurcata HBG, PC).
Etymology Referring to the robust furcate stromata.
Stromata antler-like at fertile part, dichotomously
branched several times, with palmate apices, on a gla-
brous stipe, 3–4 cm long above ground, 1–2.4 cm
long × 2–2.6 mm diam at fertile part; surface grayish
brown, becoming blackish when outer layer worn off,
with inconspicuous to conspicuous perithecial mounds,
overlain with a grayish brown outer layer gradually
worn off during maturation, underlain with a thin, black
layer ca. 10µm thick; interior white, with a black core,
(See figure on next page.)
Fig. 9 Xylaria insignifurcata (from holotype). A Stroma. B, C Stromatal surfaces. D Ascal apical rings. E Ascospores. F, G Colony on 9‑cm Petri plate
containing OA at 2 and 4 wk, respectively. H Stromatal surface bearing upright conidiophores. I, J Conidiophores. K Conidia; the arrow pointing
towards a germinating conidium. Bars in A = 1 cm; B, C = 0.25 mm; H = 1 mm; I = 10 μm; D, E, J, K = 5 µm

Page 19 of 27
Juetal. Botanical Studies (2023) 64:21
Fig. 9 (See legend on previous page.)

Page 20 of 27
Juetal. Botanical Studies (2023) 64:21
coriaceous. Perithecia spherical, 300–400 µm diam.
Ostioles conical, 80–100µm high × 90–110µm broad at
base. Asci with eight ascospores arranged in uniseriate
manner, cylindrical, 60–85µm total length, the spore-
bearing part 30–40 µm long × 4–4.5µm broad, with an
apical ring staining light blue in Melzer’s iodine reagent,
inverted hat-shaped, 1µm high × 1µm broad. Ascospores
light brown to brown, unicellular, ellipsoid-inequilateral,
with narrowly to broadly rounded ends, smooth, (3.5–)
4–4.5 × 2–2.5µm (4.1 ± 0.2 × 2.3 ± 0.2 µm, N = 40), with a
straight germ slit spore-length or nearly so on the ventral
side, lacking a hyaline sheath; epispore smooth.
Cultures and anamorph. Unknown.
Additional specimen examined VIETNAM. Tonkin,
Ke’so’, vieilles souches, Bon, as X. escharoidea var. trifida
(holotype of X. nigripes var. trifida PC 0086053).
Notes Xylaria robustifurcata is characterized by its light
brown outer stromatal layer that persists into maturity
and short sterile stromatal apices. Unlike X. furcata, its
stromata are more robust, and its perithecia are largely
immersed, resulting in inconspicuous to conspicuous but
not exposed mounds on the stromatal surface. As already
mentioned in the notes for X. furcata, X. robustifurcata
is one of the three Xylaria species found in a von Höhnel
collection of X. furcata from Java. In addition to the five
packets stored in FH, the von Höhnel collection was also
distributed as Rehm’s exsiccata Ascomyceten 1812.
e holotype packet of X. nigripes var. trifida, filed as X.
escharoidea var. trifida in PC, contains a stroma, which
undoubtedly represents one of the two stromata depicted
in Patouillard (1891) and is thus considered the holo-
type. e stroma is overmature, having the outer layer no
longer detectable.
Xylaria scoparia Pat., J. Bot. (Morot) 5:318. 1891.Fig.10
Typification VIETNAM. Tonkin, on ground, Bon, H.
(lectotype [designated here, MycoBank Typification
No. 10013827] of X. scoparia FH ex Patouillard herb.);
Tonkin, Ha Noi, on ground, 1890, Bon, H. 4399 (syntype
of X. scoparia FH ex Patouillard herb., isosyntype PC),
immature; Tonkin, Ke’so’, on ground, 28 Jul 1890, Bon, H.
4470 (syntype of X. scoparia FH ex Patouillard herb., iso-
syntypes PC [in 2 packets]), immature. TAIWAN. Tainan
City, Shen-hua District, Liu-fen-liao, on ground of bam-
boo plantation, 3 Aug 2011, Chou, K.-H. 100080301 (cul-
tured from stroma YMJ1435) [epitype (designated here,
MycoBank Typification No. 10013828) of X. scoparia
HAST 145925].
Stromata antler-like at fertile part, dichotomously
branched one to, more frequently, many times, with long
acicular and curly apices, on a glabrous stipe, 2.4–4.2cm
long above ground, 1–1.5cm long × 0.7–3.3mm diam at
fertile part; surface brown to dark brown except for black
perithecial mounds, with half-exposed to fully exposed
perithecial mounds, lacking an outer layer, underlain
with a thin, black layer ca. 10 µm thick; interior white
or pale yellow, homogeneous, soft. Perithecia spheri-
cal, 150–200 µm diam. Ostioles depressed-conical, ca.
40µm high × ca. 80 µm broad at base. Asci with eight
ascospores arranged in uniseriate manner, cylindrical,
60–80µm total length, the spore-bearing part 30–40µm
long × 3.5–4.5µm broad, with an apical ring staining blue
in Melzer’s iodine reagent, inverted hat-shaped, 1.5µm
high × 1–1.5µm broad. Ascospores brown to dark brown,
unicellular, short fusoid-inequilateral, slightly later-
ally compressed, with narrowly rounded ends, smooth,
4.5–5 × 2.5–3µm (4.9 ± 0.3 × 2.6 ± 0.2 µm, N = 40), with a
straight germ slit spore-length or nearly so on the ventral
side, lacking a hyaline sheath; epispore smooth.
Cultures and anamorph. Colonies reaching the edge of
9-cm Petri dish in 8week, whitish, mostly submerged,
azonate, with diffuse margins. Reverse uncolored or
pale tan-colored. Stromata arising copiously from the
entire colonies, antler-like, branched several times, up to
5cm long × 0.7–1.3mm diam, white but frequently yel-
low on apices, immediately becoming black from base
upwards, overlain with pale mouse gray pustules on the
entire surface due to production of conidia. Anamo-
rph produced on the stromatal surface. Conidiophores
upright, mononematous; main axis unbranched, 150–
300 × 6–9µm, dichotomously branched two to five times
on top, smooth, hyaline. Conidiogenous cells 2–3 born
on each terminal short branch, initially ampulliform,
becoming cylindrical, closely geniculate at upper end
after producing multiple conidia in sympodial sequence,
8–18 × 3.5–4.5 µm, smooth, bearing terminal poroid
conidial secession scars. Conidia produced holoblasti-
cally, hyaline, smooth, subglobose to obovoid, (5–)6.5–
8.5(–10) × (4.5–)5–6(–6.5) µm (7.5 ± 1.1 × 5.3 ± 0.5 µm,
N = 40), with a flattened base indicating former point of
attachment to conidiogenous cell.
Additional specimens examined CHINA. Kiangsi, on
ground, 20 May 1935, Deng, S. Q. 4106, as X. furcata
(UPS). MALAYSIA. Malay Peninsula, Pulau Penang,
Fig. 10 Xylaria scoparia (from epitype except for B, which is from holotype). A, B Stroma. C, D Stromatal surfaces. E. Ascal apical ring. F Ascospores.
G Colony on 9‑cm Petri plate containing OA at 8 wk. H, I. Stromatal surface bearing upright conidiophores. J Conidiophore. K Conidiogenous cells
bearing conidia. L Conidia. Bars in A, B = 5 mm; C, D = 0.25 mm; H = 1 mm; I = 50 μm; J = 25 μm; E, F, K, L = 5 µm
(See figure on next page.)

Page 21 of 27
Juetal. Botanical Studies (2023) 64:21
Fig. 10 (See legend on previous page.)

Page 22 of 27
Juetal. Botanical Studies (2023) 64:21
Waterfall Gardens, on termite nests, 23 Jan 1920, Noor,
M. 15366, as X. furcata (BPI 584609), immature; Malay
Peninsula, Pulau Penang, Waterfall Gardens, on termite
nests, 23 Jan 1920, Noor, M. 5625, as X. furcata (BPI
714719 ex Lloyd herb. 10425, BPI 584610 ex Reinking
herb.), immature. PHILIPPINES. Leyte, Palo, on ground,
Jan 1906, Elmer, A. D. E. 7233 (BPI 584827 ex Bresadola
herb.), immature. SINGAPORE. on ground, 27 Dec
1919, Chipp, T. F., as X. furcata (BPI 713933 ex Lloyd
herb. 11865), immature. SRI LANKA. on ground, Petch,
T. 21, as X. furcata (BPI 713932 ex Lloyd herb. 11858).
TAIWAN. Hua-lien County, Shou-feng Township, cam-
pus of National Dong Hwa University, on ground, 20 Jul
2011, Chou, J.-C. 100072002 (cultured from stromata
YMJ1427, YMJ1428 & YMJ1429) (HAST 145921); Hua-
lien County, Shou-feng Township, campus of National
Dong Hwa University, on ground, 3 Sep 2011, Chou,
J.-C. 100092103 (cultured from stromata YMJ1423 &
YMJ1424) (HAST 145922), immature; Tainan City, Hsin-
shih District, Tan-ting, on ground of mango orchard, 18
Jul 2016, Chou, K.-H. 105071801 (cultured from stroma
YMJ2177) (HAST 145951); Tainan City, Hsin-shih Dis-
trict, Tan-ting, on ground of mango orchard, 28 Jul 2008,
Chou, K.-H. 97072802 (cultured from stromata YMJ982
& YMJ983) (HAST 145923), immature, with anamorph
and developing perithecia; Tainan City, Hsin-shih Dis-
trict, Tan-ting, on ground of mango orchard, 2 Oct 2009,
Chou, K.-H. 98100203 (cultured from stroma YMJ1158)
(HAST 145924); Tainan City, Shen-hua District, Niu-
Chuang, on ground under Cordia dichotoma (Bor-
aginaceae), 26 Jul 2008, Chou, K.-H. 97072601 (HAST
145927); Tainan City, Shen-hua District, Niu-Chuang,
on ground under Cordia dichotoma (Boraginaceae), 15
Jul 2008, Chou, K.-H. 97071504 (cultured from stromata
YMJ958, YMJ959, YMJ960, YMJ989, YMJ991& YMJ992)
(HAST 145952), immature.
Notes Xylaria scoparia was considered a synonym of X.
furcata by Rogers etal. (2005). However, our molecular
phylogenetic analyses (Figs. 1, 2) confirmed that both
species are distinct. Compared to X. furcata, X. scoparia
has stromata that are frequently repeatedly branched
many times, long acicular and curly stromatal apices,
and darker ascospores, thus resembling X. tenellifurcata,
which differs mainly by having smaller conidia and lack-
ing the yellow apices frequently found on immature stro-
mata produced in cultures of X. scoparia (Fig.10H).
Xylaria siamensis Wangsawat, Y.-M. Ju, Phosri, Whalley
& Suwannasai, Biology (Basel) 10: 575: 21. 2021.
For descriptions of the teleomorph, cultures, and
anamorph, see Wangsawat et al. (2021) where illus-
trations of stromata, ascus, ascospores, colony,
conidiophores, and conidia are also provided. Xylaria
siamensis is characterized by the following features: stro-
mata antler-like at fertile part, dichotomously branched
one to many times, with long acicular and curly apices, on
a glabrous stipe, 2–6.7cm long above ground, 0.3–1cm
long × 0.2–1.5mm diam at fertile part; stromatal surface
dark brick to black, with conspicuous to half-exposed
perithecial mounds, lacking an outer layer, with a white,
soft interior; perithecia spherical, 200–400 µm broad,
with a coarsely conic-papillate ostiole ≤ 100µm broad at
base; ascospores brown to dark brown, ellipsoid-inequi-
lateral, with narrowly rounded ends, smooth, 5–6 × 2.5–
3.5 µm, with a straight germ slit spore-length on the
ventral side.
Notes Xylaria siamensis is currently known only from
ailand. It resembles X. scoparia and X. tenellifurcata
in having stromata with exposed perithecial mounds on
the surface and long acicular and curly apices. However,
it differs from the latter two species mainly by its coarsely
conic-papillate ostioles and slightly longer ascospores.
Wangsawat etal. (2021) obtained cultures, where stro-
mata and anamorph were not produced. e anamorph
found on the surface of young stromata from nature is
much like that of X. furcata, except for having smaller
conidia 4–4.5(–5) × 3–4 µm vs. (4.5–)5–6.5(–7) × (3.5–)
4–5(–5.5) µm in X. furcata.
Xylaria tenellifurcata Y.-M. Ju & H.-M. Hsieh, sp. nov.
Fig.11
MycoBank MB849236.
Typification TAIWAN. Tainan City, Hsin-shih District,
Tan-ting, on ground of bamboo plantation, 20 May 2009,
Chou, K.-H. 98052001 (cultured from stroma YMJ1070)
(holotype of X. tenellifurcata HAST 145928).
Etymology Referring to the delicate furcate stromata.
Stromata antler-like at fertile part, dichotomously
branched one to many times, with long acicular and
curly apices, on a glabrous stipe, 1.9–3 cm long above
ground, 0.6–1cm long × 1.2–1.6mm diam at fertile part;
surface brown to dark brown except for black perithe-
cial mounds, with 2/3-exposed to fully exposed peri-
thecial mounds, lacking an outer layer, underlain with
an extremely thin, black layer less than 10 µm thick;
interior white, homogeneous, soft. Perithecia spheri-
cal, 200–250 µm diam. Ostioles depressed-conical, ca.
40µm high × ca. 80 µm broad at base. Asci with eight
ascospores arranged in uniseriate manner, cylindrical,
55–70µm total length, the spore-bearing part 35–40µm

Page 23 of 27
Juetal. Botanical Studies (2023) 64:21
Fig. 11 Xylaria tenellifurcata (from holotype). A Stromata. B, C Stromatal surfaces. D Ascal apical rings. E, F Ascospores; ascospores showing
a straight to slightly sigmoid germ slit in (F). G Colony on 9‑cm Petri plate containing OA at 4 wk. H Stromata produced in culture. I, J. Stromatal
surface bearing upright conidiophores. K, L Conidiophore. M, N Conidiogenous cells bearing conidia. O Conidia. Bars in A = 5 mm; B, C = 0.25 mm;
H = 1 mm; I = 0.5 mm; J = 50 μm; K = 25 μm; L = 10 μm; D, E, M–O = 5 µm; F = 2.5 μm

Page 24 of 27
Juetal. Botanical Studies (2023) 64:21
long × 3.5–4.5µm broad, with an apical ring staining blue
in Melzer’s iodine reagent, inverted hat-shaped, 1.5µm
high × 1–1.5µm broad. Ascospores brown to dark brown,
unicellular, short fusoid-inequilateral, slightly later-
ally compressed, with narrowly rounded ends, smooth,
4.5–5(–5.5) × 2.5–3 µm (5.0 ± 0.2 × 2.7 ± 0.2 µm, N = 40),
with a straight to slightly sigmoid germ slit spore-length
or nearly so on the ventral side, lacking a hyaline sheath;
epispore smooth.
Cultures and anamorph. Colonies reaching the edge
of 9-cm Petri dish in 6 wk, whitish, mostly submerged,
azonate, with diffuse margins. Reverse pale tan-colored.
Stromata arising copiously from the entire colonies, ant-
ler-like, branched several times, up to 5cm long × 0.4–
0.6mm diam, white, immediately becoming black from
base upwards, overlain with pale mouse gray pustules
on the entire surface due to production of conidia.
Anamorph produced on the stromatal surface. Conidi-
ophores upright, mononematous; main axis unbranched,
100–200 × 6–7µm, dichotomously branched two to five
times on top, smooth, hyaline. Conidiogenous cells 2–3
born on each terminal short branch, initially ampulli-
form, becoming cylindrical, forming one to several con-
secutive nodulose swellings at upper end after producing
multiple conidia in sympodial sequence, 7.5–13 × 3.5–
4.5µm, smooth, bearing terminal poroid conidial seces-
sion scars. Conidia produced holoblastically, hyaline,
smooth, subglobose to obovoid, 4.5–5.5 × 3.5–4.5(–5)
µm (5.0 ± 0.3 × 4.0 ± 0.3µm, N = 40), with a flattened base
indicating former point of attachment to conidiogenous
cell.
Notes Xylaria tenellifurcata is similar to a smaller ver-
sion of X. scoparia. Both species have long acicular and
curly stromatal apices but can be distinguished primar-
ily by the smaller conidia in X. tenellifurcata and by the
yellow apices frequently found on immature stromata
produced in cultures of X. scoparia (Fig.10H). Xylaria
siamensis also has long acicular and curly stromatal api-
ces but differs in having coarsely conic-papillate ostioles
and slightly longer ascospores. Multi-locus sequence data
confirms these three species are distinct.
Identication key toX. furcata andresembling species
1. Stromata black or tan-colored at center, overlain with
a grayish brown outer layer, becoming dull black-
ish when outer layer worn off; ascospores ellipsoid-
inequilateral, (3.5–)4–4.5 × 2–2.5 µm…X. robustifur-
cata
1ʹ. Stromata with a homogenous interior, lacking an
outer layer…2
2. Perithecia immersed, with inconspicuous perithecial
mounds…3
2ʹ. Perithecial mounds conspicuous, half-exposed to
fully exposed…4
3. Stromata 1.2–1.7 mm diam at fertile part; ascospores
5.5–6(–6.5) × 3–3.5 µm…X. furcatula
3ʹ. Stromata 1.5–2.1 mm diam at fertile parts;
ascospores 4–5 × 2.5–3 µm…X. hoehnelii
4. Stromatal surface hairy on fertile parts; ascospores
4–5 × 2–2.5 µm…X. hirsuta
4ʹ. Stromatal surface glabrous on fertile parts…5
5. Stromata 5–9.6 cm long above ground, 3.6–4.8 mm
diam at fertile part; ascospores ellipsoid-inequilateral
(4–)4.5–5.5 × 2–2.5(–3) µm; conidiophores dichoto-
mously branched 3–4 times in short intervals on
top…X. insignifurcata
5ʹ. Stromata mostly less than 5 cm long above ground,
less than 3.5 mm diam; conidiophores not branched
in short intervals on top…6
6. Stromata terminating into acuminate apices…7
6ʹ. Stromata terminating into long acicular and curly
apices…8
7. Stromata 2–3.5 cm long above ground; ascospores
(3.5–)4–5.5(–6) × (2–)2.5–3 µm; conidia subglo-
bose to ellipsoid, (4.5–)5–6.5(–7) × (3.5–)4–5(–5.5)
µm…X. furcata

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Juetal. Botanical Studies (2023) 64:21
7ʹ. Stromata 0.9–1.5 cm long above ground; ascospores
(4–)4.5–5(–5.5) × 2.5–3 µm; conidia highly variable
in shape, subglobose, obovoid, ellipsoid to oblong,
equilateral or slightly to significantly oblique, (4.5–)
5.5–8.5(–11) × (3–)3.5–4.5(–5) µm…X. brevifurcata
8. Ostioles coarsely conic-papillate; ascospores 5–6 ×
2.5–3.5 μm; conidia 4–4.6(–5) × 3–4 μm…X. siamen-
sis
8ʹ. Ostioles depressed-conical; ascospores slightly
shorter…9
9. Ascospores 4.5–5 × 2.5–3 µm; stromata produced
in culture frequently with yellow apices; conidia
(5–)6.5–8.5(–10) × (4.5–)5–6(–6.5) µm…X. scoparia
9ʹ. Ascospores 4.5–5(–5.5) × 2.5–3 µm; stromata pro-
duced in culture with white apices; conidia 4.5–5.5 ×
3.5–4.5(–5) µm…X. tenellifurcata
Discussion
Species diversity of Xylaria subg. Pseudoxylaria had
not been much investigated until the study of Rogers
et al. (2005), where 11 taxa were recognized. e spe-
cies number has since been greatly enriched, with 24
more taxa added to the subgenus (Chou etal. 2017; Hsieh
etal. 2020, 2022; Ju and Hsieh 2007; Ju etal. 2011, 2022;
Kim etal. 2016; Wangsawat etal. 2021). Xylaria collec-
tions from termite nests with delicate, dichotomously
branched stromata are commonly identified as X. fur-
cata, but our study showed that multiple species can be
delimited by morphological features of teleomorphs and
anamorphs as well as sequences of multiple DNA loci.
Species delimitation amongX. furcata andresembling
species
Xylaria furcata and resembling species are all antler-
like and share similar sizes of asci, ascal apical rings, and
ascospores. Xylaria robustifurcata can be readily sepa-
rated from the other species by having a dark stromatal
core and an outer splitting layer overlying the stromatal
surface. Its perithecia are entirely immersed within stro-
matal tissue, a feature also found in X. furcatula and X.
hoehnelii. Other species, including X. furcata, X. brevifur-
cata, X. hirsuta, X. insignifurcata, X. scoparia, X. siamen-
sis, and X. tenellifurcata, normally have conspicuously
exposed perithecial mounds and were shown more
closely related among themselves (Figs.1, 2). All of the
species are glabrous on the stromatal surface and have
geographic distributions in Asia except for X. hirsuta,
which has a hirsute stromatal surface and is known only
from Africa. e average size of ascospores among the
species ranges from 4.1 to 5.8 µm in length and 2.3 to
3.1µm in width, with little variation between species. On
the other hand, conidial sizes vary greatly, with averages
ranging from 3.9 to 7.5µm in length and 3.3 to 5.3µm
in width. is makes conidial sizes useful for distinguish-
ing between species that are difficult to separate by their
teleomorphs, such as X. furcata and X. brevifurcata or X.
scoparia and X. tenellifurcata.
Anamorph ofX. furcata
Petch (1913) was probably the first to attempt to link an
anamorphic Xylaria to X. furcata, describing Botrytis-
like conidiophores formed on the stromata. ese conidi-
ophores are highly branched and terminate into a lobed
head on each branch. A whorl of flask-shaped conid-
iogenous cells is born on these heads. e conidia were
reported as being born in catenulate chains on the flask-
like conidiogenous cells. is is exceptional for a Xylaria
anamorph, where conidia are normally produced holo-
blastically in sympodial sequence. It should be noted that
Petch (1906) may have already described and illustrated
the Botrytis-like conidiophores as Piptocephalis-like from
stromata induced from fungus combs kept in moisture
chambers but did not specifically link it to X. furcata.
Dixon (1965) observed an anamorphic Xylaria species
from fungus combs of an African termite and considered
it the anamorph of X. furcata. is fungus produces two
conidia in acropetal chain on each conidiogenous cell
and released conidia ballistically. It was subsequently
named as Padixonia bispora Subram. by Subramanian
(1972). Dixon (1985) later considered the fungus not the
anamorph of X. furcata but a new Xylaria species.
ese earlier attempts to connect the teleomorph and
anamorph of X. furcata were equivocal and inconsist-
ent likely because their studied materials were not actu-
ally X. furcata. ose observations of Petch (1906, 1913)
were based on Xylaria stromata emerging from fungus
combs, where a variety of other fungi coexist with the
Xylaria stromata sympatrically and some of them pro-
duce conidia in catenulate chains (personal observation
of Y-MJ).
e anamorphs of the species being studied have
poroid conidial secession scars on top of swollen con-
idiogenous cells after producing multiple conidia. is
differs from anamorphs of most Xylaria species, which
produce denticulate conidial secession scars that are not
grouped at the top of conidiogenous cells. Anamorphs of

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Juetal. Botanical Studies (2023) 64:21
species with a teleomorph similar to that of X. furcata,
including X. brevifurcata, X. scoparia, X. siamensis, and
X. tenellifurcata, can be confused with the anamorph of
X. furcata. Despite having distinct anamorphic features
from each other and from X. furcata, these species can
still be mistaken for X. furcata especially when similar
stromata from more than one Xylaria species appear
together on a fungus comb.
Diculties encountered duringour study ofX. furcata
andresembling species
During our study of X. furcata and resembling species,
several difficulties were encountered, including: (i) a
large number of the collected stromata being immature,
(ii) stromata being mostly small and delicate, easily over-
looked in the field, (iii) poor ascospore germination, (iv)
difficulty in managing fragile stromatal tissue in small
quantity during isolation, (v) small stromata drying
quickly within hours and losing viability not long after,
and (vi) presence of multiple similar species commonly
found in a field collection. Obtaining pure cultures from
multiple stromata in a collection is crucial, particularly
when the presence of similar species is suspected. All
of the cultures that we studied were initiated from fresh
stromatal tissue before drying. Our numerous attempts
to germinate ascospores on agar media were unsuccess-
ful, as previously reported by Ju and Hsieh (2007) for
other Xylaria species associated with termite nests in
Taiwan. It is possible that the ascospore germination is
triggered or significantly promoted by passage through
termite guts or landing on termite fungus gardens.
Conclusion
Macrotermitine termites are found in Africa and Asia,
with a greater generic diversity in Africa. Ten species
were revealed among the specimens identifiable as X.
furcata, with nine known from Asia and only X. hirsuta
from Africa. e diversity of X. furcata and resembling
species was previously overlooked due to the small
size and similarity of their stromata, resulting in X.
furcata being the only recognized species. Additional
undescribed species related to X. furcata have been
confirmed through sequencing collections containing
immature stromata (unpublished data of H-MH), indi-
cating that the diversity is not limited to the ten spe-
cies included in the present study. In fact, these ten
species likely represent only a small fraction of the true
diversity, with most species yet to be described. In Tai-
wan, Odontotermes formosanus is the only macroter-
mitine termite species; however, X. furcata and seven
resembling species are associated with its nests. Mac-
rotermitine termites have a known species diversity
of 332, with 165 each in Africa and Asia and two from
Madagascar (Kambhampati and Eggleton 2000). With
the known diversity of macrotermitine termites far
greater than that of X. furcata and resembling species,
it is foreseeable that many more species of this fungal
group are yet to be collected and described.
Supplementary Information
The online version contains supplementary material available at https:// doi.
org/ 10. 1186/ s40529‑ 023‑ 00392‑x.
Additional le1: TableS1. List of isolates and taxa other than X. furcata
and resembling species included in the phylogenetic analyses.
Additional le2: Aligned ITS dataset.
Additional le3: Aligned concatenated dataset of α‑ACT, RPB2, and
β‑TUB (RPB2‑TUB‑ACT dataset).
Acknowledgements
We dedicate this study to the memory of the late Prof. Jack D. Rogers of
Washington State University, Pullman. We honor his contributions to the
family Xylariaceae and other ascomycetes, as well as his specific interests in
the genus Xylaria. Prof. Rogers had a particular fondness for Xylaria species
associated with termite nests and initiated an effort to sort out this fungal
group in 2004 in collaboration with Y‑MJ. During his visit to Taiwan in 2008
with Mrs. Belle Rogers, he specifically requested to visit termite nesting areas
to see the Xylaria species found in these habitats. We would like to express our
gratitude to K.‑H. Chou for her invaluable assistance in collecting specimens.
We also thank the curators of BPI, FH, HBG, NY for locating and loaning speci‑
mens. Special thanks go to Prof. Don Pfister of FH for his kindness in allowing
us to extract DNA from the neotype of X. furcata. Finally, we are indebted to
Mei‑Jane Fang and Chun‑Ru Lin for their technical assistances in obtaining
DNA sequences.
Author contributions
Y‑MJ recorded morphological traits, obtained cultures, wrote the manuscript,
and prepared figure plates; H‑MH obtained DNA sequences and conducted
molecular phylogenetic analyses; and NS provided cultures and DNA
sequences of Thai species. Y‑MJ served as the project leader. All authors read
and approved the final manuscript.
Funding
This work was supported by Grant MOST 111–2311‑B‑001–038‑MY2 from
National Science and Technology Council of Taiwan to Y‑MJ.
Availability of data and materials
Specimens have been deposited at the HAST herbarium, and cultures are
available at BCRC. DNA sequences have been deposited at GenBank. Newly
described species and new type designations were registered at MycoBank.
Declarations
Ethics approval and consent to participate
Not applicable.
Consent for publication
Not applicable.
Competing interests
The authors declare that they have no competing interests.
Received: 11 May 2023 Accepted: 29 June 2023

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Juetal. Botanical Studies (2023) 64:21
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