Content uploaded by David Hawksworth
Author content
All content in this area was uploaded by David Hawksworth on Jul 17, 2021
Content may be subject to copyright.
J
SE Journal of Systematics
and Evolution doi: 10.1111/jse.12705
Research Article
Pleurocordyceps gen. nov. for a clade of fungi previously
included in Polycephalomyces based on molecular
phylogeny and morphology
Yong‐Hui Wang
1,2
, Sayaka Ban
3
, Wen‐Jing Wang
1
,YiLi
4
, Ke Wang
1
, Paul M. Kirk
1,5
, Kathryn E. Bushley
6
,
Cai‐Hong Dong
1
, David L. Hawksworth
7,8,9
, and Yi‐Jian Yao
1
*
1
State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China
2
University of Chinese Academy of Sciences, Beijing 100049, China
3
Medical Mycology Research Center, Chiba University, 1‐8‐1, Inohana, Chuo‐ku, Chiba 260‐8673, Japan
4
College of Food Science and Engineering, Yangzhou University, Yangzhou 225127, Jiangsu, China
5
Biodiversity Informatics & Spatial Analysis, Royal Botanic Gardens, Kew, Richmond, Surrey TW9 3AB, UK
6
Department of Plant and Microbial Biology, University of Minnesota, St. Paul, MN 55108, USA
7
Department of Life Sciences, The Natural History Museum, Cromwell Road, London SW7 5BD, UK
8
Comparative Plant and Fungal Biology, Royal Botanic Gardens, Kew, Surrey TW9 3DS, UK
9
Jilin Agricultural University, Changchun 130118, China
*Author for correspondence. E‐mail: yaoyj@im.ac.cn
Received 4 November 2019; Accepted 29 October 2020; Article first published online 9 November 2020
Abstract Since the discovery of the Pleurocordyceps/ “Polycephalomyces”clade unaffiliated with the clades of
Clavicipitaceae s. str., Ophiocordycipitaceae, and Cordycipitaceae of clavicipitaceous fungi, some taxa have been
published and more fungal material relevant to the group have become available for further study. Here, a
multigene phylogeny using nrSSU, nrLSU, tef1, rpb1, and rpb2 was constructed with some of the recently
discovered additional taxa using maximum likelihood and Bayesian analyses (BI) to test and refine the current
phylogenetic framework for Cordyceps s. lat. and other clavicipitaceous fungi. In addition to the well supported
major Pleurocordyceps/“Polycephalomyces”clade revealed previously, another clade with newly added taxa
referred to as “Polycephalomyces formosus‐like”from Japan was found to be sister to the Pleurocordyceps/
“Polycephalomyces”clade. Extensive investigation revealed that strains named “P. formosus‐like”grouped in this
new clade and do indeed represent the true P. formosus and that species previously included in the genus
Polycephalomyces required a new generic name. Based on the phylogenetic analyses and morphological
characteristics, including both sexual and asexual morphs when available, the new generic name Pleurocordyceps
is introduced and relevant new combinations are made. A newly designated lectotype and a supporting epitype
for P. formosus is selected and the circumscription of Polycephalomyces is discussed.
Key words: Clavicipitaceae, molecular systematics, new taxa, typification.
1 Introduction
When determining the systematic position of Paecilomyces
sinensis Q.T. Chen et al. (syn. Polycephalomyces sinensis (Q.T.
Chen et al.) W.J. Wang et al.), a new clade in the
clavicipitaceous fungi (including species referred to Cordy-
ceps s. lat.), the “Polycephalomyces”clade, was revealed
based on phylogenetic analyses of a multigene data set and
of ITS sequences, respectively. The clade comprised two
subgroups, I and II (Wang et al., 2012). In this paper, the
“Polycephalomyces”clade is now renamed the Pleuro-
cordyceps clade and temporally Pleurocordyceps/“Polycepha-
lomyces”clade. The existence of this new clade was
confirmed with new combinations of species being added
to the genus Polycephalomyces:P. kanzashianus (Kobayasi &
Shimizu) Kepler & Spatafora, P. nipponicus (Kobayasi) Kepler
& Spatafora and P. ramosopulvinatus (Kobayasi & Shimizu)
Kepler & Spatafora in subgroup I and P. cuboideus (Kobayasi
& Shimizu) Kepler & Spatafora, P. paracuboideus (S. Ban
et al.) Kepler & Spatafora, P. prolificus (Kobayasi) Kepler &
Spatafora, and P. ryogamiensis (Kobayasi & Shimizu) Kepler &
Spatafora in subgroup II (Kepler et al., 2013). Furthermore, a
new genus, Perennicordyceps, was proposed to accommo-
date those species in subgroup II (Matočec et al., 2014). Since
then, more new species have been discovered in subgroup I,
the genus Polycephalomyces:P. lianzhouensis W.M. Zhang &
L. Wang (Wang et al., 2014), P. yunnanensis Hong Yu bis et al.
(Wang et al., 2015a), P.agaricus Hong Yu bis & Y.B. Wang
(Wang et al., 2015b), P. onorei Kautman & Kautmanová
Month 2021
|
Volume 00
|
Issue 00
|
1–16 © 2020 Institute of Botany, Chinese Academy of Sciences
(Crous et al., 2017b), P. phaothaiensis Mongkols et al. (Crous
et al., 2017a), P. aurantiacus Y.P. Xiao et al., and P.
marginaliradians Y.P. Xiao et al. (Xiao et al., 2018). All these
species are listed in Table 1.
Although the two subgroups in the Pleurocordyceps/
“Polycephalomyces”clade have been separated into two
genera, Polycephalomyces and Perennicordyceps, and more
species have been described in the former, the clade remains
monophyletic. Some new species in Polycephalomyces were
described with both sexual and asexual morphs with the
support of molecular phylogeny, which provided strong links
between the two morphs (Wang et al., 2014; Crous
et al., 2017a; Xiao et al., 2018). The species in the two genera
are morphologically distinct. Those in Polycephalomyces are
characterized using a laterally expanded fertile part near to
the tip of stromata, and have immersed perithecia, bacilli-
form part‐ascospores on discharge and two types of conidia
in culture. In contrast, the four species in Perennicordyceps
have the fertile part expanded equally and surrounding the
upper portion of the stromata, superficial perithecia,
ascospores disarticulating into cuboid to narrowly prismatic
part‐spores, and a single type of conidia in culture.
However, the two types of conidia in culture found in the
asexual morph of all the species currently classified in
Polycephalomyces are not those described in the type species
of the genus, P. formosus, which produces only one type of
conidia on the capitellum of synnemata (Kobayasi, 1941). To
clarify this issue, we conducted an intensive literature search,
a morphological comparison of living strains and molecular
phylogenetic analyses. This established that the generic
name Polycephalomyces was not applicable to most of the
fungi placed under the name. The type species, the only
species at the time of publication, is morphologically distinct
from them and they group in different clusters in
phylogenetic analyses. This subgroup I of taxa in the
Pleurocordyceps/“Polycephalomyces”clade therefore re-
quired a new generic name. Here we introduce a new
generic name and provide full descriptions and a discussion
of the generic circumscription.
The type specimen of P. formosus was collected from Prov.
Musasi, Mt. Takao‐san, Japan (Kobayasi, 1941), but was
destroyed during World War II as mentioned by Seifert
(1985), who designated a neotype originally labeled
“Cordyceps falcata B & B, Hakagata, Feb. 1923, Herb. T. Petch
–Bequeathed 1949”, latter relabeled “Cordyceps falcata
Berk., Sri Lanka, Hakagata, Feb 1923, ex Herb. T. Petch, K(M)
187597”for the species. Later an nrLSU sequence from a
strain of P. formosus, ARS Collection of Entomopathogenic
Fungal (ARSEF) 1424, was obtained by Bischoffet al. (2003).
Since then, more gene fragments were sequenced from
ARSEF 1424 and widely used to represent Polycephalomyces
in phylogenetic analyses of the Cordyceps s. lat. group (e.g.,
Chaverri et al., 2005; Wang et al., 2012, 2014, 2015a, 2015b;
Kepler et al., 2013; Quandt et al., 2014; Sanjuan et al., 2015;
Ban, 2016; Crous et al., 2017a, 2017b; Xiao et al., 2018).
However, several strains representing Polycephalomyces
isolated from Japan were analyzed by Ban (2016) and two
species distinguished molecularly and morphologically were
revealed and named as P. formosus‐like and P. ramosus‐like.
Strikingly, strains of “P. formosus‐like”were found in a
distantly related clade from that of ARSEF 1424 in
phylogenetic analyses (Ban, 2016). It was therefore necessary
to clarify the true identity of the Pleurocordyceps/“Poly-
cephalomyces”clade. The nomenclature of Polycephalomyces
was investigated and strains of P. formosus‐like isolated from
Japan were studied to provide necessary information for the
generic concept of Polycephalomyces.
2 Material and Methods
2.1 Fungal materials
Living cultures of Polycephalomyces formosus‐like (NBRC 100686,
100687, 103843, 109993, 109994, and 109995) and P.ramosus‐
like (NBRC 101760, 109984, and 109985) strains reported in Ban
(2016) were obtained from the National Institute of Technology
and Evaluation Biological Resource Center (NBRC) in Japan. A
strain of P. fo rmosus (ARSEF 1424) was obtained from the USDA‐
ARS Collection of Entomopathogenic Fungal cultures (ARSEF) in
New York. All cultures were maintained at 4 °C on potato
dextrose agar (PDA) slants in the dark and incubated at
approximately 25 °C for 14–20 days for this study.
The dried specimens constituting the neotype of P.
formosus,“Cordyceps falcata Berk., Sri Lanka, Hakagata,
Feb 1923, ex Herb. T. Petch, K(M) 187597”designated by
Seifert (1985) and the holotype of P. ramosus (Peck) Mains,
“Stilbum ramosum, holotype, leg. C. H. Peck, USA, N.Y.,
Cayunga County, Sterling (NYS).”were obtained on loan by
courtesy of the Fungarium of Royal Botanic Gardens, Kew (K
(M)) and the Herbarium of the New York State Museum
(NYS), respectively.
2.2 Culture and observation
Cultures were inoculated with a 1 cm diam. agar disk from a
colony of 7 cm diam. in a Petri dish and grown on PDA at
25 °C. Morphological examinations were performed every 2–5
days. For characterizing the strains, microscope slide cultures
were prepared using inoculating a small amount of mycelium
on a desired nutrient agar medium block of 1 cm
2
overlaid
using a coverslip of 2.2 cm as described in Wang et al. (2012).
Microscopic observations were carried out and photo-
graphed using a Zeiss Axioscope microscope equipped with
AxioCam MRc. Microscopic measurements were made using
AxioVision Rel. 4.6 software (Zeiss, Welwyn Garden City, UK).
Table 1 Species within the “Polycephalomyces”clade (Pleurocordyceps clade)
Subgroup I (Polycephalomyces) Subgroup II (Perennicordyceps)
“Polycephalomyces”clade
(Pleurocordyceps clade)
P. agaricus, P. aurantiacus, P. kanzashianu,
P. lianzhouensis, P. onorei, P. marginaliradians,
P. nipponicus, P. phaothaiensis, P. ramosopulvinatus,
P. sinensis, P. yunnanensis
P. cuboideus, P. paracuboideus,
P. prolificus, P. ryogamiensis
2 Wang et al.
J. Syst. Evol. 00 (0): 1–16, 2021 www.jse.ac.cn
2.3 DNA isolation, polymerase chain reaction (PCR)
amplification and sequencing
Fresh mycelia were harvested from strains grown on PDA at
25 °C for 14 days. Total genomic DNA was extracted using the
Fungi Genomic DNA Extraction Kit (Solarbio, Beijing, China),
and following the manufacturer's instructions. For dried
specimens, a method using Chelex‐100 (Bio‐Rad, Hercules,
CA, USA), which needs less material, was employed to
extract DNA, as described in Wang et al. (2012). Six nuclear
genes, including ITS, nrSSU, nrLSU, tef1, rpb1, and rpb2, were
amplified and sequenced. For the ITS region, primers ITS5
and ITS4 (White et al., 1990) were used and the PCR
amplification was conducted as described in Jiang & Yao
(2005). nrLSU and the nrSSU were amplified following the
PCR procedures of Sung et al. (2001) with primer pairs of
LR0R and LR5 (Vilgalys & Sun, 1994), and NS1 and NS4 (White
et al., 1990), respectively. tef1 was amplified with primers
983F and 2218R under the conditions of Rehner (2001).
Primers CRPB1A and RPB1Cr were employed for rpb1
amplification according to Castlebury et al. (2004). Amplifi-
cation of rpb2 with primers fRPB2‐5F and fRPB2‐7cR was
carried out using the protocol in Wang et al. (2012). PCR
amplification was performed on a thermocycler (ProFlex™
PCR System; Applied Biosystems, Foster City, CA, USA). The
25 μL PCR reactions contained 12.5 μL2×Taq MasterMix, 1 μL
each primer (10 μmol/L), 1 μL template DNA, and 9.5 μL
purified water.
DNA sequencing was carried out in both directions using
an automatic sequence analyzer from the Tsingke Biological
Technology (Beijing, China). The information and GenBank
accession numbers of taxa used in the ITS and multigene
phylogenetic analyses are provided in Tables S1 and S2,
respectively.
2.4 Phylogenetic analyses
In total, 163 sequences of nrSSU, nrLSU, tef1, rpb1, and rpb2
from species of Polycephalomyces and Perennicordyceps have
been submitted by various authors in recent years and were
retrieved from the GenBank database. Those sequences and
43 newly amplified sequences of nrSSU, nrLSU, tef1, rpb1, and
rpb2 from strains of P. formosus‐like and P. ramosus‐like
strains (see Table S2) were aligned to the multigene dataset
used by Sung et al. (2007b), Wang et al. (2012), Kepler et al.
(2013), and Quandt et al. (2014). Taxa from representative
families of Glomerellales, Glomerellaceae, and Plectosphaer-
ellaceae, were included as outgroups (see Table S2). In total,
61 ITS sequences, including 48 strains of Polycephalomyces
and Perennicordyceps and six of P. formosus‐like, were
assembled. Five strains of Ophiocordyceps were chosen as
outgroups (see Table S1).
Sequences of ITS and the multigene dataset including
nrSSU, nrLSU, tef1, rpb1,andrpb2 were aligned and manually
edited using BioEdit Version 7.0.9.0 software (Hall, 1999).
The matrixes and trees of ITS and the multigene were
deposited in TreeBASE (http://purl.org/phylo/treebase/
phylows/study/TB2:S26846). Maximum likelihood (ML) anal-
yses were performed with RAxML‐7.0.3‐WIN using a GTR‐
GAMMA model for evolution (Stamatakis, 2006). Nodal
support was assessed with non‐parametric bootstrapping
using 500 replicates. Bayesian analyses were run in the
MrBayes v3.1.2 program (Ronquist & Huelsenbeck, 2003) for
5×10
6
generations using two independent runs with four
chains until the average standard deviation of the split
frequencies between the simultaneous runs was below 0.01
and the log‐likelihood had reached stationary. A general
time reversible (GTR) model of DNA substitution with
gamma‐distributed rate variation across invariant sites as
determined by MrModeltest version 2.2 (Nylander, 2004)
was used for the multigene analyses, while the default F81
model was used for the ITS analyses. Trees were sampled
every 100 generations. The first 12 500 trees were discarded
from further analyses and the remaining ones used for
calculating posterior probabilities (PP) in the majority‐rule
consensustree.Treeswerefigured in FigTree v1.4.3
(Rambaut, 2014). Bootstrap proportions higher than 70%
of ML analyses (ML‐BP) and posterior probabilities of
Bayesian analyses (BI‐PP) greater than 95% are shown in
the phylogenetic trees.
3 Results
3.1 Morphological observations
Morphological observations were carried out on both
cultures and dried specimens.
3.1.1 Observation on living cultures
The morphological characteristics of the living cultures including
culture colony, hyphae, synnemata and conidial masses, and
microscopic characters of phialides and conidia for Polycepha-
lomyces formosus ARSEF 1424, P. ram osus‐like and P. formosus‐
like strains were observed and compared (Table 2). The
synnemata of P. for mosus ARSEF 1424 and P. ramosus ‐like
strains were almost simple and unbranched while those of P.
formosus‐like NBRC 109993 were often branched. In Petri dish
culture, two types of phialides and conidia were observed in P.
formosus ARSEF 1424 and P. ramosus ‐like NBRC 109985: α‐
phialides of the acremonium‐type producing subglobose to
ovoid α‐conidia at the terminal portion of the synnemata and
also at the edge of the colony, and β‐phialides of the hirsutella‐
type producing fusiform β‐conidia on the stipe of the
synnemata and also mycelium surface of the colony.
Conversely, α‐conidia were observed at the terminal portion
of synnemata from the P. formosus‐like strains but no β‐conidia
were found on the stipe of the synnemata from the Petri dish
cultures. However, in slide cultures, both P. formosus ARSEF
1424 and P. ramosus ‐like strains produced fusiform β‐conidia
ofteninchainsonphialides,whilethoseofP. form osus‐like
strains produced ovoid, oblong ellipsoidal to cylindric conidia,
often forming spore balls on the phialide.
Detailed descriptions of each strain are provided below.
Polycephalomyces formosus ARSEF 1424 Colonies growing
fairly well on PDA in Petri dish cultures, attaining 1.2 cm
diameter in 10 days at 25 °C, short floccose and white,
reverse reddish brown. Hyphae hyaline, branched,
smooth‐walled, 1.0–4.5 μm wide. Synnemata arising after
13 days on PDA; 1–15 mm long, clavate, pale yellow;
radiating with a ring‐like distribution (Fig. 1). Conidial
mass opaque, slimy, yellow‐orange, generated from the
apex of the synnema or on the surface of the colony
(Fig. 1). α‐Phialides verticillate and acropleurogenous on
conidiophores; cylindrical to subulate at the base,
3A new genus of Pleurocordyceps
J. Syst. Evol. 00 (0): 1–16, 2021www.jse.ac.cn
tapering gradually from the base to the apex; 6.5–71 µm
long, 0.8–1.5 µm wide at the base and 0.7–1.1 µm wide at
the apex. α‐Conidia subglobose to ovoid,
1.5–2.7 ×1.2–2.0 µm, formed in viscous pools located on
the agar and at the terminal portion of synnemata
forming conidial masses. β‐phialides acropleurogenous,
solitary on the hyphae, with swollen bases, tapering
abruptly from the base to the apex; 9.0–48 μmlong,
1.3–2.5 μmwideatthebaseand0.5–1.2 μmwideatthe
apex. β‐conidia fusiform, 3.5–4.5 (–6.0) ×1.2–2.5 (–3.5)
µm, produced by phialides on the stipe of the synnema
and mycelium surface of the colony, single or often in
chains on the phialides, sometimes aggregated to form
irregular spore balls (Fig. 1).
Polycephalomyces ramosus‐like Colonies on PDA in Petri
dish culture attaining a diameter of 1.0–1.6 cm in
10 days at 25 °C, cottony, white to orange‐yellow, and reverse
dry yellow. Hyphae hyaline, septate, branched, smooth‐
walled, 0.7–5.0 μm wide. Synnemata solitary, clavate,
unbranched 2–6 mm long. Conidial mass cream arising from
the apex of the synnemata or covering the surface of the
colony (Fig. 2). α‐Phialides verticillate and acropleurogenous,
in whorls of 1–3 on conidiophores, cylindrical to subulate at
the base, or occurring directly on the aerial hyphae;
7.0–59 μm long, 1.2–2.2 μm wide at the base and 0.5–1.1 μm
wide at the apex. α‐Conidia subglobose or ellipsoidal,
1.5–2.5 ×1.2–2.5 μm, produced on the edge of the colony. β‐
Phialides acropleurogenous, solitary, with swollen bases,
terminal phialides tapering abruptly, 8.7–41 μm long,
1.5–2.6 μm wide at the base and 0.7–1.1 μm wide at the
apex. β‐Conidia fusiform 3.5–6.5 ×1.2–3.0 μm, produced on
the mycelium surface of the colony, single or often in chains
on phialides (Fig. 2).
Polycephalomyces formosus‐like Colonies on PDA in Petri
dish cultures growing slowly, attaining 1.0 cm diameter in 10
days at 25 °C, dense, white, and reverse white or dull buff.
Hyphae hyaline, branched, smooth‐walled, 0.7–5.0 μmwide.
Synnemataemergingafter20days,formingtwotothree
branches, 0.5–30 mm long, arising as several radiating rings
on the colony (Fig. 3). Conidial masses pale yellow, arising
from the apex of synnemata or covering the surface of the
colony (Fig. 3). Phialides developing from the colony and
the terminal parts of synnemata; cylindrical to subulate at
the base; 5.5–75 μm long, tapering gradually from
0.9–1.8 μmatthebaseto0.8–1.5 μm at the apex. Conidia
of one type, one‐celled, smooth‐walled, ellipsoid to ovoid,
2.0–3.5 ×1.0–1.5 μm, arising in a conidial mass on the agar or
on the terminal portions of synnemata (Fig. 3). Conidia
absent from the stipes of synnemata. In slide culture,
phialides monothetic and solitary or acropleurogenous in
whorls of 1–4, narrowly lageniform or subulate, 7.0–51 μm
long, 1.0–2.5 μmwideatthebaseand0.5–1.0 μmwideat
the apex. Conidia obovoid to oblong ellipsoidal or
cylindrical, 2.0–6.0 ×1.2–1.8 μm; forming irregular spore
balls near the apex of phialides (Fig. 3).
3.1.2 Observations on the type material of Polycephalomyces
formosus and P. ramosus
Neotype of Polycephalomyces formosus The neotype
comprised two dried specimens, which were white and
broken into several fragments. Under the microscope, a
few conidia were observed, ellipsoidal, and measuring
1.5–3.5 ×1–1.5 μm (Fig. 4).
Holotype of Polycephalomyces ramosus The holotype on
loan from NYS consisted of a single microscopic slide,
mounted in an unknown medium and a tiny piece of
sample specified for DNA work. Examination of the slide
showed no discernible fungal structures under the micro-
scope (Fig. 5).
Table 2 Morphological comparison of species of Polycephalomyces formosus ARSEF 1424, P. ramosus‐like and P. formosus‐like
Species Colony Synnemata (mm) Phialides (µm) Conidia (µm)
P. formosus
ARSEF 1424
Colony short
floccose and
white, reverse
reddish brown.
Synnemata arising after
13 days, 1–15 long,
radiating with a ring‐
like distribution.
Conidial mass opaque,
slimy, yellow‐orange.
α‐Phialides 6.5–71 long,
0.8–1.4 wide at the base
and 0.7–1.1 wide at the
apex. β‐Phialides
9.0–47.9 long, 1.3–2.5
wide at the base and
0.5–1.2 wide at the apex.
α‐Conidia subglobose to
ovoid, 1.5–2.7 ×1.2–2.0.
β‐Conidia fusiform,
3.5–4.6 (–6.1)
×1.2–2.5 (–3.5).
P. ramosus‐like Colony cottony,
white to orange‐
yellow, reverse dry
yellow.
Synnemata solitary,
clavate, unbranched
2–6 long. Conidial mass
cream.
α‐Phialides 7.2–59 long,
1.2–2.2 wide at the base
and 0.5–1.1 wide at the
apex. β‐Phialides
8.7–40.8 long, 1.5–2.6
wide at the base and
0.7–1.1 wide at the apex.
α‐Conidia subglobose or
ellipsoidal,
1.5–2.5 ×1.2–2.2.
β‐Conidia fusiform,
3.5–6.2 ×1.2–3.0.
P. formosus‐like Colony dense, white,
and reverse white
or dull buff.
Synnemata emerging
after 20 days, 0.5–30
long, showing several
radiating ring‐like
distributions. Conidial
masses pale yellow.
Phialides 5.5–75 long,
tapering gradually from
0.9–1.8 at the base to
0.8–1.4 at the apex.
Conidia ellipsoidal, ovoid,
2.0–3.4 ×1.0–1.5.
4 Wang et al.
J. Syst. Evol. 00 (0): 1–16, 2021 www.jse.ac.cn
3.2 Phylogenetic analyses
Amplification and sequencing of dried specimen DNA
extraction, amplification and sequencing of the “neotype”of
P. formosu s and the holotype of P. ra mosus were performed with
a minute scraps of material. Amplification of the ITS sequence
failed for the neotype of P. formosus,butforP. r amosus resulted
in a sequence with a 97.60% similarity to accession KJ412030 in
GenBank named as Malassezia restricta strain ITC52‐1C.
Data sets The combined multigene data set with 187
taxa consisted of 4181 base pairs (nrSSU 998 bp, nrLSU
832 bp, tef1 898 bp, rpb1 559 bp, rpb2 894 bp) after the
exclusion of ambiguously aligned sites. In total,107 taxa
were complete for all the five gene sequences, 40 taxa were
complete for four genes, and 28 for three genes (Table S2).
The numbers of taxa for each gene were: nrSSU 159 taxa,
nrLSU 179, tef1 170, rpb1 158, and rpb2 135 (Table S2). ML
Fig. 1. “Polycephalomyces formosus”(ARSEF 1424). A, Obverse colony side on potato dextrose agar (PDA). B, Reverse colony
side on PDA. C, Synnemata growing on PDA medium. D,α‐Phialides from a conidial mass. E, Phialides and conidia. F,
β‐Phialides. G,α‐Conidia from conidia mass on synnemata. H,β‐Conidia. Scale bars: A, B =2 cm, C =500 μm, D, F–H=10 μm,
E=20 μm.
5A new genus of Pleurocordyceps
J. Syst. Evol. 00 (0): 1–16, 2021www.jse.ac.cn
analyses produced a tree with a log‐likelihood (–ln) of
89294.064046. Bayesian analyses revealed that the triple
repeated analyses converged on a stationary phase and the
50% majority‐rule consensus trees were topologically
identical. The tree topology produced in the ML and
Bayesian analyses was very similar and the Bayesian
consensus tree is provided in Fig. 6 with the BP values of
ML analyses and the PP values from one of the Bayesian
analyses cited as nodal support.
The ITS sequence data matrix included 520 nucleotide
characters after the exclusion of ambiguous sites
at both ends. ML analyses produced a tree with a log‐
likelihood (–ln) of 3352.132305. In the Bayesian analyses,
all three repeated analyses converged on a stationary
Fig. 2. Polycephalomyces ramosus‐like (NBRC 109985). A, Obverse colony side on potato dextrose agar (PDA). B, Reverse
colony side on PDA. C, Synnemata growing on PDA medium. D,α‐Phialides from a conidial mass. E, Phialides and conidia. F,
β‐Phialides. G,α‐Conidia from conidia mass on synnemata. H,β‐conidia. Scale bars: A, B =2 cm, C =1 mm, D, F–H=10 μm,
E=20 μm.
6 Wang et al.
J. Syst. Evol. 00 (0): 1–16, 2021 www.jse.ac.cn
phaseandthethree50%majority‐rule consensus trees
were topologically identical. There was no significant
difference in the tree topology produced using ML and
Bayesian analyses. The Bayesian consensus tree is
provided here with BP values of ML analyses and PP
values from one of the Bayesian analyses (Fig. 7).
Phylogeny Eight major clades representing currently
accepted families within the Hypocreales were recognized
from the multigene analyses. These clades include Bionec-
triaceae, Clavicipitaceae, Cordycipitaceae, Hypocreaceae, Nec-
triaceae, and Ophiocordycipitaceae as recognized by Sung
et al. (2007a, 2007b), the Pleurocordyceps/“Polycephalo-
myces”clade proposed by Wang et al. (2012), and the P.
formosus‐like clade revealed in our new study (Fig. 6). The
latter two clades could be viewed as possible family rank
clades. Six major clades, Bionectriaceae, Cordycipitaceae,
Hypocreaceae, Nectriaceae, Ophiocordycipitacea, and Pleuro-
cordyceps/“Polycephalomyces”, received support of 100% PP
Fig. 3. Polycephalomyces formosus‐like (CGMCC 5.2206 epitype). A, Obverse colony side on potato dextrose agar (PDA). B,
Reverse colony side on PDA. C, Synnemata growing on PDA medium. D,α‐Phialides from a conidial mass. E,F, Phialides and
conidia. G, Irregular conidia balls. H, Conidia from conidia mass on synnemata. Scale bars: A, B =2 cm, C =500 μm, D, G,
H=10 μm, E, F =20 μm.
7A new genus of Pleurocordyceps
J. Syst. Evol. 00 (0): 1–16, 2021www.jse.ac.cn
in the Bayesian analyses, and the two clades of Clavicipita-
ceae and P. formosus‐like received 99% support. Most clades
had support of 100% BP in ML analyses except for that of
Clavicipitaceae (99%) and Nectriaceae (97%). Two species, P.
formosus‐like and Cordyceps pleuricapitata, formed a new
monophyletic clade with strong support in our Bayesian
analyses and ML (PP =99%, ML‐BP =100%). Among the eight
clades, only the group formed by Ophiocordycipitaceae, P.
formosus‐like, and Pleurocordyceps/“Polycephalomyces”
clades and its extended group forming the Clavicipitaceae
clade were supported by 100% PP in Bayesian analyses, but
<100% for the other groups. Both the Pleurocordyceps/
“Polycephalomyces”and P. formosus‐like clades were further
divided into two subclades; Pleurocordyceps/“Polycephalo-
myces”(subclade I in Wang et al., 2012) and Perennicordyceps
(subclade II in Wang et al., 2012) for the former, and
subclades P. formosus‐like and Cordyceps pleuricapitata for
the latter (Fig. 6). These subclades all received strong
support in the ML and Bayesian analyses.
In addition to the taxa in the Pleurocordyceps/“Polycephalo-
myces”cladeofthemultigenedataset,twotaxa,P. onorei and
P.sp.BRACR23906,wereincludedintheITSdataset.The
topology of the ITS phylogenetic tree is similar to that of the
Pleurocordyceps/“Polycephalomyces”subclade in the multigene
tree (Fig. 7). Most of the species were clearly separated in the
tree, but P. lianz houensis and P. ramos op ulvinatus were very
closely related. Based on both the ITS and multigene
phylogenetic analyses, these two species were considered as
distinct species. Therefore, 10 species can be recognized in the
Pleurocordyceps/“Polycephalomyces”subclade, that is, P. si -
nensis, P. lianzhouensis, P. ramosopulvinatus, P. yunnanensis, P.
aurantiacus, P. marginaliradians, P. phaothaiensis, P. nipponicus,
P. onorei,andP. agaricus.
3.3 Taxonomy
The new clade, the Polycephalomyces formosus‐like clade,
revealed in this study posed a taxonomic question on the
generic name to be used for the species included in
subclade I of the Pleurocordyceps/“Polycephalomyces”clade
(Fig. 7). Based on our morphological observations and
molecular analyses, the two clades, the P. fo rmosus‐like
clade and the Pleurocordyceps/“Polycephalomyces”clade,
represented two distinct groups of fungi both morpholog-
Fig. 4. The material proposed as a “neotype”of Polycephalomyces formosus by Seifert (1985). A,B, The loan of the neotype of
P. formosus.C,D, Conidia. Scale bars: A, B =2 cm, C, D =10 μm.
Fig. 5. Polycephalomyces ramosus (holotype).
8 Wang et al.
J. Syst. Evol. 00 (0): 1–16, 2021 www.jse.ac.cn
ically and molecularly. Separation in well resolved clades in
the phylogenetic analyses was supported by spore
production along the stipes of synnemata and the conidial
types, especially in culture. No conidia were formed on the
stipes of synnemata and only one conidial type was
produced on the capitate terminus of the synnemata in P.
formosus as reported by Kobayasi (1941) and confirmed by
Seifert (1985) and also in cultures of P. formos us‐like strains
Fig. 6. Phylogenetic tree from maximum likelihood (ML) and Bayesian analyses (BI) analyses of multigene (nrSSU, nrLSU, tef1,
rpb1 and rpb2) data set showing the relationships within Hypocreales. Bootstrap proportions (equal to or above 70%) of ML
analyses (ML‐BP) and posterior probabilities (PP) (equal to or above 95%) are shown above internodes before and after
backslash, respectively. The names of taxa (e.g., clades, families) are provided to the right of species names.
9A new genus of Pleurocordyceps
J. Syst. Evol. 00 (0): 1–16, 2021www.jse.ac.cn
as reported by Ban (2016) and confirmed in this study. The
production of a single conidial type in Petri dish cultures
reported by Ban (2016) for strains of P. formosus‐like was also
confirmed by us. In contrast, two types of conidia were found
in cultures in all the species previously assigned to
Polycephalomyces except for P. lianzhouensis, but the type of
conidia was not clearly described previously. There was only
one conidial type in a few species of Polycephalomyces found
in the wild, because of their structural features, for example,
P. aurantiacus (conidia oval to globose; Xiao et al., 2018) and P.
marginaliradians (conidia fusiform; Xiao et al., 2018), but two
types of conidia were later found to be produced in Petri dish
cultures (Xiao et al., 2018). Although a number of species have
been transferred to, and new species described in, Poly-
cephalomyces, it is apparently a misapplied generic name to
the group of fungi which produce two types of conidia in Petri
dish cultures. Therefore, we introduce a new generic name for
this group of fungi.
Pleurocordyceps Y.J. Yao, Y.H. Wang, S. Ban, W.J. Wang, Yi
Li, Ke Wang & P.M. Kirk, gen. nov.
Fungal Names FN570683
Diagnosis: Species in this new genus differ from
Perennicordyceps and Polycephalomyces formosus‐like fungi
(Polycephalomyces s. str.) in producing lateral fertile
pulvinate stromata close to the tip in the sexual morph
and two types of conidia in Petri dish culture in the asexual
morph.
Type:Paecilomyces sinensis Q.T. Chen et al. (syn. Pleuro-
cordyceps sinensis (Q.T. Chen et al.) Y.J. Yao et al. see below).
Etymology:Pleuro‐(Gk), lateral, in a sideways position;
referring to the fertile part with perithecia formed laterally
on stromata of the cordycepitoid sexual morph.
Hosts: Entomogenous and/or fungicolous.
Distribution: China, Ecuador, Japan, Thailand.
Description:Sexual morph:Stromata singular to numerous,
simple, fleshy, cylindrical, reddish brown, growing upwards
Fig. 7. Phylogeny of the Pleurocordyceps/“Polycephalomyces”and P. formosus‐like clades from maximum likelihood (ML) and
Bayesian analyses (BI) analyses using ITS sequences. Bootstrap proportions (equal to or above 70%) of ML analyses (ML‐BP)
and posterior probabilities (PP) (equal to or above 95%) are shown above internodes before and after backslash, respectively.
The names of taxa are provided to the right of species names.
10 Wang et al.
J. Syst. Evol. 00 (0): 1–16, 2021 www.jse.ac.cn
from the head or irregularly in various directions from the
whole body of the host; simple or branched. Stipes
cylindrical, tip pointed, perithecia forming laterally pad on
the stipe and often toward the apical parts of the stroma,
subterminal, sometimes appearing terminal when mature;
sterile tip curved or twisted, some missing when mature.
Perithecia pyriform and/or ovoid, with protruding ostioles,
immersed when young, emerged and separate when mature.
Asci cylindrical, wall thickened at the apex. Ascospores
hyaline, filiform, cylindrical, fragmenting to form small
truncate, bacilliform part‐spores.
Asexual morph:Colonies growing relatively well in culture.
Synnemata solitary to caespitose or crowded, unbranched or
rarely branched, arising from an insect corpse, other fungus
stroma, or in culture; stipes of the synnemata tomentose,
clavate or spatulate or/and cylindrical; a viscous spore mass
produced mostly on the terminal portions of the synnemata,
rarely in the median part of the synnemata and then
surrounding the stipe; some species produce unformed,
scab‐like aggregate conidial structure on the tawny stipe.
Conidiogenous cells phialides, of two types; α‐phialides
verticillate and acropleurogenous on conidiophores, cylin-
drical to subulate at the base or occurring directly on the
aerial hyphae; β‐phialides acropleurogenous and solitary on
the hyphae, narrow lageniform or subulate, tapering abruptly
from the base to the apex. Conidia one‐celled, hyaline and
smooth‐walled, of two types in culture; α‐conidia globose to
subglobose or ellipsoidal, in viscous pools located on the
colony in culture and at the terminal portion or occasionally
in the middle of the stipe of synnemata forming a conidial
mass; β‐conidia fusiform, produced along stipe of the
synnemata as well as on the mycelium surface of the colony,
single or often in chains on phialides.
Remarks:Polycephalomyces was first described as an
asexually typified genus (Kobayasi, 1941) and further species
with only asexual morphs were included in the genus, mostly
through combinations from other genera. The sexual morph
was not reported when the Pleurocordyceps/“Polycephalo-
myces”clade was revealed (Wang et al., 2012). Since then,
more species were included in Polycephalomyces and several
species with sexual morphs were combined with the genus
by Kepler et al. (2013). However, some of the species with
sexual morphs were latter removed from Polycephalomyces,
for example, the four species of Perennicordyceps (Matočec
et al., 2014). Subsequently, more species were newly
described in Polycephalomyces including some with sexual
morphs, that is P. lianzhouensis, P. onorei, P. phaothaiensis,
and P. marginaliradians (Wang et al., 2014; Crous
et al., 2017a, 2017b; Xiao et al., 2018). Currently, the two
genera within the Pleurocordyceps/“Polycephalomyces”clade,
representing subclades I and II, are Pleurocordyceps and
Perennicordyceps. These were well separated in the
phylogenetic analyses (Figs. 6 and 7) and also morpholog-
ically distinguished in both the sexual and asexual morphs,
for example, formation of fertile part and perithecia, shape
of part‐spores, and types of conidia as summarized above.
As the circumscription of the generic names has changed,
new combinations of species originally placed in Polycepha-
lomyces into Pleurocordyceps are made below, based on an
examination of type or other information in the protologue.
All these have the unambiguous sexual morph and/or the
two types of conidia, especially in culture, and are supported
by molecular data; that is, species that display the consistent
characters of Pleurocordyceps, as circumscribed here. Note
that because of the lack of taxonomic evidence, some
species placed in Polycephalomyces remain as a residue
pending further investigation.
Pleurocordyceps sinensis (Q.T. Chen et al.) Y.J. Yao, Y.H.
Wang, S. Ban, W.J. Wang, Yi Li, Ke Wang & P.M. Kirk,
comb. nov.
Fungal Names FN570676
Basionym:Paecilomyces sinensis Q.T. Chen et al., Acta
Mycol. Sin. 3: 25 (1984).
Synonym:Polycephalomyces sinensis (Q.T. Chen et al.) W.J.
Wang et al., Syst. Biodiv. 10(2): 228 (2012).
Type: China, Sichuan Province, Kangding County, on larva
of Hepialus armocanus (Lepidoptera), 1980, HMAS 43720.
Pleurocordyceps sinensis grouped in a terminal clade with
other sequences, including Polycephalomyces formosus
ARSEF 1424, P. ramosus NBRC 109983, and P. tomentosus
BL 4 (Figs. 6 and 7).
The strain P. formosus ARSEF 1424 from Poland, reported
by Bischoffet al. (2003), has been widely used to represent
that species (e.g., Chaverri et al., 2005; Wang
et al., 2012, 2014, 2015a, 2015b; Kepler et al., 2013;
Quandt et al., 2014; Sanjuan et al., 2015; Ban, 2016; Crous
et al., 2017a, 2017b; Xiao et al., 2018). Morphological
observationonthisstrain,however, demonstrated that
two types of conidia (α‐and β‐conidia) were produced and
were unlike the true P. formosus,whichisdescribedas
producing “onetypeofconidiabyconidialmassatthe
apex of the synnemata”(Kobayasi, 1941). However, P.
formosus‐like strains, collected from Japan were much
closer to the type location, Prov. Musasi, Mt. Takao‐san,
than that of the neotype from Sri Lanka designated by
Seifert (1985), and displayed the same characters as
described by Kobayasi (1941) (Ban, 2016; this study).
Furthermore, strains ARSEF 1424 and P. fo rmosu s‐like,
clustered in distantly related clades (Ban, 2016; this study).
These are therefore different species and P. formo sus‐like
matches much more closely with the original concept of P.
formosus than ARSEF 1424, which we regard as incorrectly
identified.
Polycephalomyces ramosus (Peck) Mains (syn. Stilbum
ramosum Peck; Mains, 1948), was originally described as
“head subglobose whitish or pale yellow; stem thick, smooth,
branched, white above, pallid or brownish below, sometimes
creeping and sending up branches at intervals; spores minute,
oblong”(Peck, 1873) and “the conidia are hyaline, ellipsoidal to
obovoid, 2.2–3.3 ×1.1–1.5 μm and are covered by a mucus. They
are produced singly on subulate phialides which are 15–30 μm
long and up to 1.5 μmwideatthebase”based on the collection
by Mains (1948). Later, Seifert (1985) provided a description of
P.ramosus as “Phialides of two types, A‐phialides in capitulum,
B‐phialides on stipe, producing A‐and B‐conidia respectively.
A‐conidia obovoid to broadly obpyriform, 2–3×1–2μm.
B‐conidia fusiform, catenate, 3–4.5 ×1.5–2μm”with no
indication if it was based on the cited holotype material.
Polycephalomyces ramosus from Croatia was also reported with
two types of conidia (Matočec et al., 2014). Furthermore, two
strains named as P. ram osus, NBRC 100693 and NBRC 109983,
11A new genus of Pleurocordyceps
J. Syst. Evol. 00 (0): 1–16, 2021www.jse.ac.cn
with two types of conidia were cited by Ban (2016) and the
latter was accepted by Crous et al. (2017b) and Xiao et al.
(2018). Our microscopic examination of the slide from the
holotype of P. ra mo sus did not reveal any conidia. However, the
ITSsequenceobtainedinthisstudyfromtheholotypeofP.
ramosus was similar to that of KJ412030 in GenBank, close to a
sequence labeled as Malassezia restricta E. Guého et al. Due to
the lack of sufficient information on the application of the
name, the identity of P. ramosus requires further investigation.
Polycephalomyces tomentosus (Schrad.) Seifert (syn.
Stilbum tomentosum Schrad.) was excluded from Stilbum
by Sutton (1973) by recognizing a new genus, Blistum B.
Sutton. The first named strain of P. tomentosus in
phylogenetic analyses was obtained by Bischoffet al.
(2003) and considered to be Blistum, based on the 28S
rDNA phylogenetic analyses and its “myxomyceticolous
habit”(Bischoffet al., 2003; Wang et al., 2012). Later,
another strain of P. tomentosus, BL 4, was introduced in
phylogenetical analyses by Kepler et al. (2013) and since then
it has been accepted in Quandt et al. (2014), Wang et al.
(2015a), Crous et al. (2017a, 2017b), and Xiao et al. (2018).
However, there was not sufficient evidence to be clear on
the species identity of P. tomentosus BL 4 as the papers
mentioning it lacked morphological and ecological informa-
tion. As P. tomentosus was described as a parasite of slime
molds (Mycetozoa), we suspect it is probably not related to
the entomopathogenic fungi treated here.
Based on an analysis of all the names included in this
terminal clade, Polycephalomycesformosus,P.ramosus,P.
tomentosus and P. sinensis, the latter have been clearly
circumscribed both morphologically and molecularly by
Wang et al. (2012). Therefore, Polycephalomyces sinensis is
combined into Pleurocordyceps to typify this terminal
clade.
Pleurocordyceps agarica (Hong Yu bis & Y.B. Wang) Y.H.
Wang, S. Ban, W.J. Wang, Yi Li, Ke Wang, P.M. Kirk & Y.J. Yao,
comb. nov.
Fungal Names FN570677
Basionym:Polycephalomyces agaricus Hong Yu bis & Y.B.
Wang Mycol. Progr. 14 (No. 70): 4 (2015).
Type: China, Yunnan Province, Song ming County, Dashao
village, on the stroma of Ophiocordyceps sp. associated with
the melolonthid larva buried in the soil, 8 Aug. 2013, YHH
PA1305.
Pleurocordyceps aurantiaca (Y.P. Xiao et al.) Y.H. Wang, S.
Ban, W.J. Wang, Yi Li, Ke Wang, P.M. Kirk & Y.J. Yao,
comb. nov.
Fungal Names FN 570678
Basionym:Polycephalomyces aurantiacus Y.P. Xiao et al.,
Scientific Reports 8 (No. 18087): 2 (2018).
Type: Thailand, Prachuap Khiri Khan, on dead larvae
(Coleopteran), 29 Jul. 2015, MFLU 17‐1393.
Pleurocordyceps lianzhouensis (W.M. Zhang & L. Wang)
Y.H. Wang, S. Ban, W.J. Wang, Yi Li, Ke Wang, P.M. Kirk & Y.J.
Yao, comb. nov.
Fungal Names FN570679
Basionym:Polycephalomyces lianzhouensis W.M. Zhang & L.
Wang, Mycol. Progr. 13: 1093 (2014).
Type: China, Guangdong, Lianzhou, Dadongshan Nature
Reserve, on a Lepidoptera larva in fallen leaves, 18 Apr. 2002,
GDGM 20918.
The original description of Polycephalomyces lianz-
houensis followed the style of Kobayasi (1982) and Seifert
(1985) with photographs of synnemata growing on rice
medium, but the conidial types were unclear (Wang
et al., 2014), with only one type of conidia based on a
scanning electron micrograph. Nevertheless, based on
the sexual morph characters and our phylogenetic
analyses, this taxon belongs in the new genus.
Pleurocordyceps marginaliradians (Y.P. Xiao et al.) Y.H.
Wang, S. Ban, W.J. Wang, Yi Li, Ke Wang, P.M. Kirk & Y.J. Yao,
comb. nov.
Fungal Names FN570674
Basionym:Polycephalomyces marginaliradians Y.P. Xiao
et al., Scientific Reports 8 (No. 18087): 4 (2018).
Type: Thailand, Chiang Mai, The Mushroom Research
Center, on dead Cossidae larvae (Lepidoptera), 11 Jun. 2017,
MFLU 17‐1582.
Pleurocordyceps nipponica (Kobayasi) Y.H. Wang, S. Ban,
W.J. Wang, Yi Li, Ke Wang, P.M. Kirk & Y.J. Yao, comb. nov.
Fungal Names FN570680
Basionym:Cordyceps nipponica Kobayasi, Bull. Biogeogr.
Soc. Jap. 9: 151 (1939).
Synonym:Polycephalomyces nipponicus (Kobayasi) Kepler
& Spatafora, Fungal Biology 117: 618 (2013).
Type: Japan, Prov. Owari. Nisi‐kasngaigun. Sinkawa‐mati,
on Graptopsaltria nigrofuscata (Hemiptera), Jul. 1936, A.
Hayakawa, Type in Kobayasi Herb.
This species is combined into the new genus based on
characters of the sexual morph as illustrated in Kobayasi (1939)
and Shimizu (1994), displaying the typical fertile pads formed
laterally near to the top of the stromata. The two types of
conidia characteristically produced in the asexual morph have
recently been reported by Ban et al. (2020, unpublished data).
OnesequenceunderthenamePolycephalomyces
kanzashianus (GenBank Accession: AB027371, submitted
by Nikoh & Fukatsu, 2000) is included in the Pleuro-
cordyceps nipponica clade (Fig. 6). Based on the
characters of their sexual morph, they appear to be
different species (cf. Shimizu, 1994, figs. 35 and 37). The
sexual and asexual morphs of P. nippo ni cus (Shimizu,
1994;Ban,2016)areinaccordwiththecircumscriptionof
Pleurocordyceps and the new combination is therefore
made here. If Polycephalomyces kanzashinatus later
proves to be conspecificwithPleurocordyceps nipponica,
the new combination would stand because of the priority
of publication of the epithets.
Pleurocordyceps onorei (Kautman & Crous) Y.H. Wang, S.
Ban, W.J. Wang, Yi Li, Ke Wang, P.M. Kirk & Y.J. Yao,
comb. nov.
Fungal Names FN570675
Basionym:Polycephalomyces onorei Kautman & Crous,
Persoonia 38: 367 (2017).
Type: Ecuador, Cotopaxi Province, Union de Toachi village,
on caterpillar of Lepidoptera (cf. Arctinae), on stem of
Etlingera sp., 27 Mar. 2011, BRA CR23902.
12 Wang et al.
J. Syst. Evol. 00 (0): 1–16, 2021 www.jse.ac.cn
Pleurocordyceps phaothaiensis (Mongkols. et al.) Y.H.
Wang, S. Ban, W.J. Wang, Yi Li, Ke Wang, P.M. Kirk & Y.J.
Yao, comb. nov.
Fungal Names FN570684
Basionym:Polycephalomyces phaothaiensis Mongkols et al.,
Persoonia 39: 327 (2017).
Type: Thailand, Phitsanulok Province, Noen Maprang district,
on Coleoptera larva, buried in soil, 27 Jun. 2017, BBH42883.
Pleurocordyceps ramosopulvinata (Kobayasi & Shimizu)
Y.H. Wang, S. Ban, W.J. Wang, Yi Li, Ke Wang, P.M. Kirk & Y.J.
Yao, comb. nov.
Fungal Names FN570682
Basionym:Cordyceps ramosopulvinata Kobayasi & Shimizu,
Bull. natn. Sci. Mus., Tokyo, B 9(1): 2 (1983).
Synonym:Polycephalomyces ramosopulvinatus (Kobayasi &
Shimizu) Kepler & Spatafora, Fungal Biology 117: 618 (2013).
Type: Japan, Yamagata Pref. Isl. Tobishima, on nymph of
Cicadidae (Hemiptera), 19 Aug. 1980, TY 141.
The combination of the species in Polycephalomyces known to
have sexual morphs was mainly based on DNA sequence
analyses (Kepler et al., 2013). As in the case of P. nipponicus,this
requires further morphological support. Both these species are
accepted as members of Pleurocordyceps based on the
characteristics of the fertile pads formed near to the top of
the stromata, as illustrated by Kobayasi (1939) and Kobayasi &
Shimizu (1983) as well as by the DNA data. These characteristics
of the fertile structures are similar to that of these newly
described species with sexual morphs described here despite a
significant difference in the size of stromata and fertile pads.
Pleurocordyceps yunnanensis (Hong Yu bis et al.) Y.H.
Wang, S. Ban, W.J. Wang, Yi Li, Ke Wang, P.M. Kirk & Y.J. Yao,
comb. nov.
Fungal Names FN570681
Basionym:Polycephalomyces yunnanensis Hong Yu bis
et al., Phytotaxa 208: 39 (2014).
Type: China, Yunnan Province, Kunming, the Wild Duck
Lake Forest Park, on the stroma of Ophiocordyceps nutans on
the ground, 28 Jul. 2010, YHH PY1006.
In the phylogenetic analyses (Figs. 6 and 7), sequences
from this species clustered with those from strains named P.
ramosus‐like by Ban (2016). The name P. yunnanensis used for
this terminal clade is selected because the identity of P.
ramosus required clarification (this study). However, if the P.
ramosus‐like strains proved to be identical to the type of P.
ramosus, that name would have priority over P. yunnanensis.
Key to accepted species of the genus Pleurocordyceps
In order to facilitate species identification, a key to the
species recognized in Pleurocordyceps is provided here based
on morphological and ecological characters. Note that the
asexual morph is generally seen in culture.
1a. Ascomata present ……………………………………….. 2
1b. Ascomata absent ………………………………………… 7
2a. Growing on nymph of Cicadidae ……………………….. 3
2b. Growing on other hosts or substrates ………………… 4
3a. Perithecia partly immersed, 750–925 ×275–300 μm…
………………………………………… P. ramosopulvinata
3b. Perithecia immersed, 800–950 ×300–370 μm…………
………………………………………………… P. nipponica
4a. Perithecia (900–) 950–1067(–1100) ×(350–)352–429
(–450) µm; only known from Coleoptera larvae
.……………………………………………P. phaothaiensis
4b. Perithecia shorter than 950 μm; only known from
Lepidoptera larvae ….…………………………………....5
5a. Perithecia 355–473 ×158–197 μm; Asci 89–194 ×2.0–
4.0 μm……………………………………... P. lianzhouensi s
5b. Perithecia longer than 500 μm; Asci longer than
200 μm………………………………………………….…6
6a. Perithecia 676–803 ×246–328 μm; Asci 459–556 ×
3.0–4.5 μm; Ascospores easily breaking into cylindrical
part‐spores…………………….…….. P. marginaliradians
6b. Perithecia 854–950 ×330–395 μm; Asci shorter than
510 μm; Ascospore part‐spores bacilliform, sometimes
not divided……..……………………………….. P. onorei
7a. Synnemata mushroom‐shaped, 0.5–12 mm long……….
….………………………………………………… P. agarica
7b. Synnemata with other shapes ………………………….8
8a. Synnemata 50–60 mm long; α‐conidia ovoid……...….
…………………………………………..……… P. sinensis
8b. Synnemata shorter than 15 mm long; α‐conidia globose to
subglobose or ellipsoidal…… ….……………………….…9
9a. α‐Phialides 10.5–18.5 ×0.8–1.8 μm, β‐phialides narrowly
slender, 22–64 ×1–1.5 μm………..………… P. aurantiaca
9b. α‐Phialides 20–57 ×1.0–2.3 μm, β‐phialides narrowly
lageniform or subulate, 7.0–30 ×2.3–3.7 μm…………
…………………………………………….P. yunnanensis
3.4 Polycepholomyces
The genus Polycephalomyces was established with a single
species, P. formosu s, by Kobayasi (1941). The type
specimen of the species and genus, collected from Prov.
Musasi, Mt. Takao‐san, Japan, was destroyed and a
collection was designated as a “neotype”from a
collectiononinsectlarvafromSriLanka(Seifert,1985).
The “neotype”was originally labeled “Cordyceps falcata B
& B, Hakagata, Feb. 1923, HERB. T. Petch—Bequeathed
1949”, latterly relabeled “Cordyceps falcata Berk., Sri
Lanka,Hakagata,Feb1923,exHerb.T.Petch,K(M)
187597”and is now in poor condition, not suitable for
both morphological and molecular studies. In designating
a“neotype”, Seifert had overlooked that there was an
illustration that should have been selected as a lectotype
as that is “original material.”This “neoptypification”can
therefore be set aside and so an epitype, to support the
lectotype, is needed to stabilize the use of both the
generic and species names.
When selecting the epitype, living strains recently obtained
from Japan by Ban (2016) were considered. A collection from
Hara, Takatsuki‐shi, Osaka Pref., Japan, in 2005, near to the
type locality “Prov. Musasi, Mt. Takao‐san”(Kobayasi, 1941)
was chosen to support the lectotype of P. formosus as
discussed above. The epitype is derived from a living strain
named as P. formosus‐like by Ban (2016) and as P. formosus in
the Nite Biological Resource Center (NBRC 109993), where it
is specified “on Coleopteran larva from Hara, Takatsuki‐shi,
Osaka Pref., Japan, in 2005.”Details of the nomenclature and
typification are presented below.
Polycephalomyces Kobayasi, Sci. Rep. Tokyo Bunrika Daig.,
Sect. B 5: 245 (1941).
Type:Polycephalomyces formosus Kobayasi.
13A new genus of Pleurocordyceps
J. Syst. Evol. 00 (0): 1–16, 2021www.jse.ac.cn
Polycephalomyces formosus Kobayasi, Sci. Rep. Tokyo
Bunrika Daig., Sect. B 5: 245 (1941).
Holotype: Japan, Prov. Musasi, Mt. Takao‐san, on larvae of
Coleoptera, 15 Jul. 1936, Y. Kobayasi (herb. Kobayasi
destroyed).
Lectotype (Fungal Names FN570686): Figures A–Cof
Polycephalomyces formosus Y. Kobayasi in Kobayasi (1941,
Sci. Rep. Tokyo Bunrika Daig., Sect. B 5: 246) (reproduced
here as Fig. 8), lectotype designated here.
Epitype (Fungal Names FN570687):Asubculture,
derived from a living strain (NBRC 109993) originated
from Japan, Hara, Takatsuki‐shi, Osaka Pref., on coleop-
teran larva, 2005 (NBRC H‐12427), now preserved in the
China General Microbiological Culture Collection Center
(CGMCC) in a metabolically inactive state using lyophiliza-
tion and deep‐freezing in liquid nitrogen and accessed as
CGMCC 5.2206 is designated here as epitype to support
the lectotype (Fungal Names FN570686) designated
above. A dried culture (with 20 plates), prepared at the
same time was preserved in the Fungarium (HMAS),
Institute of Microbiology, Chinese Academy of Sciences
andaccessedasHMAS248264.
Description: Sexual morph: unknown. Asexual morph:A
description of the epitype, cultured on PDA at 25 °C for 10–20
days, is provided above in the morphological observations
under Polycephalomyces formosus‐like.
Ecology: entomogenous, parasitic on a Coleoptera larvae.
Distribution: Japan.
4 Discussion
All the eight major clades included in this multigene analysis
received very strong support. Further grouping of these
clades mostly received less than 80% BP support, except the
extended group including Ophiocordycipitaceae, Polycepha-
lomyces formosus‐like, Pleurocordyceps/“Polycephalomyces”
and Clavicipitaceae clades received support of 96% BP and
also 100% PP (Fig. 6). This showed that members of these
clades were more closely related than the other clades,
even some of the former Cordyceps s. lat. group, that is, the
Cordycipitaceae clade. However, the relationships among
the Clavicipitaceae, Ophiocordycipitaceae, Pleurocordyceps/
“Polycephalomyces”and P. fo rmosus‐like clades were not
resolved in ML analysis. Each of them was more or less
equally independent from each other. The possible family
clade of Pleurocordyceps/“Polycephalomyces”proposed by
Wang et al. (2012) is confirmed and a further possible family
clade of P. formo sus‐like is suggested in this study
(Figs. 6 and 7).
The phylogenetic analyses revealed more molecular
variations within the P. formosus‐like clade, which were
divided into two subclades (Figs. 6 and 7). This study
included only one taxon in each of the subclades, P. formosus
and Cordyceps pleuricapitata. With more taxon sampling with
the P. formosus‐like clade, it is likely to differentiate those
two subclades as separate genus or even higher taxonomy
rank. The similar scenario has resulted in the Pleurocordyceps/
“Polycephalomyces”clade. The two subclades were now
recognized as two genera, Pleurocordyceps and Perennicor-
dyceps, with strong support by morphology in both the
sexual and asexual morphs and phylogeny in the ML and
Bayesian analyses.
Polycephalomyces was based on P. form os us collected
from Japan (see above), with only one type of conidia
described as ellipsoidal to ovoid, 2.5–3.2 ×1–1.2 μm,
produced on the head of synnemata (Kobayasi, 1941). A
further description of the species was provided by Seifert
(1985, see above), gave the conidia as obovoid to
ellipsoidal, 1.5–3×1–1.5 μm based on a collection from
Sri Lanka. Later, another strain identified as P. fo rmosus,
ARSEF 1424, was obtained from Poland by Bischoffet al.
(2003) and has widely been used to represent Polycepha-
lomyces in phylogenetic analyses (e.g., Chaverri et al., 2005;
Fig. 8. Polycephalomyces formosus (Kobayasi, 1941: Pl. s.n. and A–C, lectotype)
14 Wang et al.
J. Syst. Evol. 00 (0): 1–16, 2021 www.jse.ac.cn
Wang et al., 2012; Kepler et al., 2013; Quandt et al., 2014;
Wangetal.,2014,2015a,2015b;Sanjuanetal.,2015;
Ban, 2016; Crous et al., 2017a, 2017b; Xiao et al., 2018).
While the strain ARSEF 1424 produced both αand βtypes
of conidia (Bischoffet al., 2003), it differed in the type of
conidia (αconidia) as described by Kobayasi (1941) and
Seifert (1985) and therefore was a misapplication of the
name. A number of new species were subsequently
described with two types of conidia in Polycephalomyces
(Wang et al., 2012, 2015a, 2015b; Crous et al., 2017a; Xiao
et al., 2018). However, species with one, two and three
types of conidia were included Polycephalomyces by Seifert
(1985); P.formosus,P.cylindrosporusSamson & H.C. Evans
(one type of conidia); P.ramosus (two types of conidia);
and P. tomentosu s (three types of conidia). He could not
have appreciated the generic significance of conidal types
at that time in the absence of DNA data. Recently,
P. formosu s‐like strains with one type of conidia, from
Japan have been recovered and demonstrate that they are
phylogenetically distinguished from those species having
two types of conidia (Ban, 2016, fig. 3). Ban (2016) provided
convincing evidence of the importance of conidial types in
phylogenetic analyses and morphological taxonomy (fig. 2
and table 2 in Ban, 2016). The P. formosus‐like strains
displayed typical characters including only one type of
conidia (αconidia) produced in conidial mass at the apex of
synnema (Ban, 2016; this study) of P. for mosus, which
typified the genus Polycephalomyces (Kobayasi, 1941). The
lectotype and supporting epitype designated in this study
for P. formosus and further for Polycephalomyces have one
type of conidia.
There are currently 23 names published under Poly-
cephalomyces (see Index Fungorum, accessed on 14
October 2020). Among them, 10 have been combined
with the new genus, Pleurocordyceps,asshownabove;four
were previously combined in Perennicordyceps.Polycepha-
lomyces orbicularis, was a taxonomic synonym of Stilbella
byssiseda (Seifert, 1985; also see Species Fungorum:
Stilbella byssiseda;Hypocreomycetidae)whileP. f ormosus
remains as the true Polycephalomyces in our study. There
are six names which need further taxonomic studies. Most
of them were described as having one type of conidia,
except P. kanzashianus which was described from only the
sexual morph (Shimizu, 1994). As shown above, sequences
named as both P. kanzashianus and P. nip po ni ca were
clustered in the same terminal species clade sharing a high
similarity, but their sexual morphs are distinguished
significantly in morphology.
Among five species with one conidial type in the
protologue, two were later described as having two or three
conidial types; P. ramosus (two; Seifert, 1985; Bischoffet al.,
2003; Matočec et al., 2014; and Ban, 2016 for both
P. ramosus and P. ramosus‐like) and P. tomentosus (three;
Seifert, 1985; and one; Sutton, 1973; Bischoffet al., 2003;
Ban, 2016). Although the sequences have been used for
phylogenetic analyses, the identity of both P. ramosus and
P. tomentosus require further investigation. The remaining
three, P. cylindrosporus, P. ditmarii Van Vooren & Audibert and
P. paludosus Mains, might possibly share the same mor-
phology and molecular features, but they were not adequately
studied and there is no DNA sequence available in GenBank.
Acknowledgements
This work is supported by the Ministry of Science and
Technology (2018YFD0400201, 2012FY111600), the Chinese
Academy of Sciences (XDA19050201) and the Ministry of
Ecology and Environment of China (2019HJ2096001006).
References
Ban S. 2016. Re‐classification of family Ophiocordycipitaceae, the
entomopathogenic fungi. Microbial Resources and Systematics 32:
105–113. (in Japanese)
BischoffJ, Sullivan R, Hywel‐Jones N, Struwe L, White J. 2003.
Resurrection of Blistum tomentosum and its exclusion from
Polycephalomyces (hyphomycetes, Deuteromycota) based on 28S
rDNA sequence data. Mycotaxon 86: 433–444.
Castlebury LA, Rossman AY, Sung G‐H, Hyten AS, Spatafora JW.
2004. Multigene phylogeny reveals new lineage for Stachybotrys
chartarum, the indoor air fungus. Mycological Research 108:
864–872.
Chaverri P, BischoffJF, Evans HC, Hodge KT. 2005. Regiocrella, a new
entomopathogenic genus with a pycnidial anamorph and its
phylogenetic placement in the Clavicipitaceae. Mycologia 97:
1225–1237.
Crous PW, Wingfield MJ, Burgess TI, Carnegie AJ, Hardy G, Smith D,
Summerell BA, Cano Lira JF, Guarro J, Houbraken J, Lombard L,
Martin MP, Sandoval‐Denis M, Alexandrova AV, Barnes CW,
Baseia IG, Bezerra JDP, Guarnaccia V, May TW, Hernandez‐
Restrepo M, Stchigel AM, Miller AN, Ordonez ME, Abreu VP,
Accioly T, Agnello C, Agustin Colman A, Albuquerque CC, Alfredo
DS, Alvarado P, Araujo‐Magalhaes GR, Arauzo S, Atkinson T,
Barili A, Barreto RW, Bezerra JL, Cabral TS, Camello Rodriguez F,
Cruz R, Daniels PP, Da Silva BDB, De Almeida DAC, De Carvalho
Junior AA, Decock CA, Delgat L, Denman S, Dimitrov RA,
Edwards J, Fedosova AG, Ferreira RJ, Firmino AL, Flores JA,
Garcia D, Gene J, Giraldo A, Gois JS, Gomes AAM, Goncalves CM,
Gouliamova DE, Groenewald M, Gueorguiev BV, Guevara‐Suarez
M, Gusmao LFP, Hosaka K, Hubka V, Huhndorf SM, Jadan M,
Jurjevic Z, Kraak B, Kucera V, Kumar TKA, Kusan I, Lacerda SR,
Lamlertthon S, Lisboa WS, Loizides M, Luangsa‐Ard JJ, Lyskova
P, Mac Cormack WP, Macedo DM, Machado AR, Malysheva EF,
Marinho P, Matocec N, Meijer M, Mesic A, Mongkolsamrit S,
Moreira KA, Morozova OV, Nair KU, Nakamura N, Noisripoom W,
Olariaga I, Oliveira RJV, Paiva LM, Pawar P, Pereira OL, Peterson
SW, Prieto M, Rodriguez‐Andrade E, Rojo De Blas C, Roy M,
Santos ES, Sharma R, Silva GA, Souza‐Motta CM, Takeuchi‐
Kaneko Y, Tanaka C, Thakur A, Smith MT, Tkalcec Z, Valenzuela‐
Lopez N, Van Der Kleij P, Verbeken A, Viana MG, Wang XW,
Groenewald JZ. 2017a. Fungal Planet description sheets: 625–715.
Persoonia 39: 270–467.
Crous PW, Wingfield MJ, Burgess TI, Hardy G, Barber PA, Alvarado P,
Barnes CW, Buchanan PK, Heykoop M, Moreno G, Thangavel R,
Van Der Spuy S, Barili A, Barrett S, Cacciola SO, Cano‐Lira JF,
Crane C, Decock C, Gibertoni TB, Guarro J, Guevara‐Suarez M,
Hubka V, Kolarik M, Lira CRS, Ordonez ME, Padamsee M,
Ryvarden L, Soares AM, Stchigel AM, Sutton DA, Vizzini A, Weir
BS, Acharya K, Aloi F, Baseia IG, Blanchette RA, Bordallo JJ,
Bratek Z, Butler T, Cano‐Canals J, Carlavilla JR, Chander J,
Cheewangkoon R, Cruz R, Da Silva M, Dutta AK, Ercole E, Escobio
V, Esteve‐Raventos F, Flores JA, Gene J, Gois JS, Haines L, Held
BW, Jung MH, Hosaka K, Jung T, Jurjevic Z, Kautman V,
Kautmanova I, Kiyashko AA, Kozanek M, Kubatova A, Lafourcade
M, La Spada F, Latha KPD, Madrid H, Malysheva EF, Manimohan
P, Manjon JL, Martin MP, Mata M, Merenyi Z, Morte A, Nagy I,
15A new genus of Pleurocordyceps
J. Syst. Evol. 00 (0): 1–16, 2021www.jse.ac.cn
Normand AC, Paloi S, Pattison N, Pawlowska J, Pereira OL,
Petterson ME, Picillo B, Raj KNA, Roberts A, Rodriguez A,
Rodriguez‐Campo FJ, Romanski M, Ruszkiewicz‐Michalska M,
Scanu B, Schena L, Semelbauer M, Sharma R, Shouche YS, Silva
V, Staniaszek‐Kik M, Stielow JB, Tapia C, Taylor PWJ, Toome‐
Heller M, Vabeikhokhei JMC, Van Diepeningen AD, Van Hoa N,
Van Tri M, Wiederhold NP, Wrzosek M, Zothanzama J,
Groenewald JZ. 2017b. Fungal Planet description sheets:
558–624. Persoonia 38: 240–384.
Hall T. 1999. BioEdit: A user‐friendly biological sequence alignment
editor and analysis program for Windows 95/98/NT. Nucleic Acids
Symposium Series 41: 95–98.
Jiang Y, Yao Y‐J. 2005. ITS sequence analysis and ascomatal
development of Pseudogymnoascus roseus.Mycotaxon 94: 55–73.
Kepler R, Ban S, Nakagiri A, BischoffJ, Hywel‐Jones N, Alisha Owensby
C, Spatafora J. 2013. The phylogenetic placement of hypocrealean
insect pathogens in the genus Polycephalomyces:Anapplicationof
One Fungus One Name. Fungal Biology 117: 611–622.
Kobayasi Y. 1939. On the genus Cordyceps and its allies on cicadae from
Japan. Bulletin of the Biogeographical Society of Japan 9: 145–176.
Kobayasi Y. 1941. The genus Cordyceps and its allies. Science Reports
of the Tokyo Bunrika Daigaku, Section B No. 84 5: 53–260.
Kobayasi Y. 1982. Keys to the taxa of the genera Cordyceps and
Torrubiella.Transactions of the Mycological Society of Japan 23:
329–364.
Kobayasi Y, Shimizu D. 1983. Cordyceps species from Japan 6. Bulletin
of the National Science Museum, Series B 9: 1–21.
Mains EB. 1948. Entomogenous fungi. Mycologia 40: 402–416.
Matočec N, Kušan I, Ozimec R. 2014. The genus Polycephalomyces
(Hypocreales) in the frame of monitoring Veternica cave
(Croatia) with a new segregate genus Perennicordyceps.
Ascomycete.org 6: 125–133.
Nylander JAA. 2004. MrModeltest 2.2. Program distributed by the
author [online]. Evolutionary Biology Centre, Uppsala University,
Uppsala. Available from https://github.com/nylander/MrModeltest2
[accessed 14 October 2020].
Peck CH. 1873. Descriptions of new species of fungi. Bulletin of the
Buffalo Society of Natural Sciences 1: 41–72.
Quandt CA, Kepler RM, Gams W, Araujo JP, Ban S, Evans HC, Hughes
D, Humber R, Hywel Jones N, Li Z, Luangsa‐Ard JJ, Rehner SA,
Sanjuan T, Sato H, Shrestha B, Sung GH, Yao YJ, Zare R,
Spatafora JW. 2014. Phylogenetic‐based nomenclatural pro-
posals for Ophiocordycipitaceae (Hypocreales) with new combi-
nations in Tolypocladium.IMA Fungus 5: 121–34.
Rambaut A 2014. FigTree v1.4.2 molecular evolution, phylogenetics
and epidemiology [online]. Available from http://tree.bio.ed.ac.
uk/software/figtree/ [accessed 14 October 2020].
Rehner SA 2001. Primers for Elongation Factor 1‐a (EF1‐a) [online].
Available from https://www.docin.com/p-1613748809.html [ac-
cessed 14 October 2020].
Ronquist F, Huelsenbeck JP. 2003. MrBayes 3: Bayesian phylogenetic
inference under mixed models. Bioinformatics 19: 1572–1574.
Sanjuan TI, Franco‐Molano AE, Kepler RM, Spatafora JW, Tabima J,
Vasco‐Palacios AM, Restrepo S. 2015. Five new species of
entomopathogenic fungi from the Amazon and evolution of
neotropical Ophiocordyceps.Fungal Biology 119: 901–916.
Seifert KA. 1985. A monograph of Stilbella and some allied
hyphomycetes. Studies in Mycology 27: 1–235.
Shimizu D. 1994. Color iconography of vegetable wasps and plant
Worms. Tokyo: Seibundo Shinkosha. (in Japanese)
Stamatakis A. 2006. RAxML‐VI‐HPC: Maximum likelihood‐based
phylogenetic analyses with thousands of taxa and mixed models.
Bioinformatics 22: 2688–2690.
Sung G‐H, Hywel‐Jones NL, Sung J‐M, Luangsa‐Ard JJ, Shrestha B,
Spatafora JW. 2007a. Phylogenetic classification of Cordyceps
and the clavicipitaceous fungi. Studies in Mycology 57: 5–59.
Sung G‐H, Spatafora J, Zare R, Hodge KT, Gams W. 2001. A revision of
Verticillium sect. Prostrata. II. Phylogenetic analyses of SSU and
LSU nuclear rDNA sequences from anamorphs and teleomorphs
of the Clavicipitaceae.Nova Hedwiga 72: 311–328.
Sung GH, Sung JM, Hywel‐Jones NL, Spatafora JW. 2007b. A multi‐gene
phylogeny of Clavicipitaceae (Ascomycota, Fungi): Identification of
localized incongruence using a combinational bootstrap approach.
Molecular Phylogenetics and Evolution 44: 1204–1223.
Sutton BC. 1973. Hyphomycetes from Manitoba and Saskatchewan,
Canada. Mycological Papers 132: 16–20.
Vilgalys R, Sun BL. 1994. Ancient and recent patterns of geographic
speciation in the oyster mushroom Pleurotus revealed by
phylogenetic analysis of ribosomal DNA sequences. Proceedings
of the National Academy of Sciences of the United States of
America 91: 4599–4603.
Wang L, Li H‐H, Chen Y‐Q, Zhang W‐M, Qu L. 2014. Polycephalomyces
lianzhouensis sp. nov., a new species, co‐occurs with Ophiocor-
dyceps crinalis.Mycological Progress 13: 1089–1096.
Wang W‐J, Wang X‐L, Li Y, Xiao S‐R, Kepler RM, Yao Y‐J. 2012.
Molecular and morphological studies of Paecilomyces sinensis
reveal a new clade in clavicipitaceous fungi and its new
systematic position. Systematics and Biodiversity 10: 221–232.
Wang Y‐B, Yu H, Dai Y‐D, Chen Z‐H, Zeng W‐B, Yuan F, Liang Z‐Q.
2015a. Polycephalomyces yunnanensis (Hypocreales), a new
species of Polycephalomyces parasitizing Ophiocordyceps nutans
and stink bugs (hemipteran adults). Phytotaxa 208: 34.
Wang Y‐B, Yu H, Dai Y‐D, Wu C‐K, Zeng W‐B, Yuan F, Liang Z‐Q. 2015b.
Polycephalomyces agaricus, a new hyperparasite of Ophiocordy-
ceps sp. infecting melolonthid larvae in southwestern China.
Mycological Progress 14: 70.
White TJ, Bruns T, Lee S, Taylor J. 1990. Amplification and direct
sequencing of fungal ribosomal RNA genes for phylogenetics. In:
Innis MA, Gelfand DH, Sninsky JJ, White TJ eds. PCR protocols: A
guide to methods and applications. San Diego: Academic Press.
315–322.
Xiao YP, Wen TC, Hongsanan S, Jeewon R, Luangsa‐Ard JJ, Brooks S,
Wanasinghe DN, Long FY, Hyde KD. 2018. Multigene phyloge-
netics of Polycephalomyces (Ophiocordycipitaceae, Hypocreales),
with two new species from Thailand. Scientific Reports 8: 18087.
Supplementary Material
The following supplementary material is available online for
this article at http://onlinelibrary.wiley.com/doi/10.1111/jse.
12705/suppinfo:
Table S1. Taxa used in ITS phylogenetic analyses.
Table S2. Taxa used in 5‐gene phylogenetic analyses.
16 Wang et al.
J. Syst. Evol. 00 (0): 1–16, 2021 www.jse.ac.cn
