Hirticlavula elegans, a new clavarioid fungus from Scandinavia

Abstract

Based on material from Denmark and Norway the new clavarioid genus Hirticlavula with one species, H. elegans, is described. It produces tiny, hyaline white basidiomata with upward pointing blunt ended hairs. Phylogenetic analysis of the LSU region of rDNA strongly supports the inclusion of this fungus in the Clavariaceae sensu stricto where it occupies a position sister to all members of Clavaria, Camarophyllopsis, and Hodophilus. It has been found from May to October fruiting on wet, rotten hardwood bark on the ground, and its lignicolous, saprophytic nutritional mode further distinguishes it from other members of the Clavariaceae.

Full text

Karstenia 54: 1–8, 2014
Hirticlavula elegans, a new clavarioid fungus from
Scandinavia
JENS H. PETERSEN, MARIE L. DAVEY and THOMAS LÆSSØE
PETERSEN, J.H., DAVEY, M.L. & LÆSSØE, T. 2014: Hirticlavula elegans, a new
clavarioid fungus from Scandinavia – Karstenia 54: 1–8. HELSINKI. ISSN 0453–3402.
Based on material from Denmark and Norway the new clavarioid genus Hirticlavula
with one species, H. elegans, is described. It produces tiny, hyaline white basidiomata
with upward pointing blunt ended hairs. Phylogenetic analysis of the LSU region of
rDNA strongly supports the inclusion of this fungus in the Clavariaceae sensu stricto
where it occupies a position sister to all members of Clavaria, Camarophyllopsis, and
Hodophilus. It has been found from May to October fruiting on wet, rotten hardwood
bark on the ground, and its lignicolous, saprophytic nutritional mode further distin-
guishes it from other members of the Clavariaceae.
Key words: Clavariaceae, taxonomy, phylogeny, Hirticlavula
Jens H. Petersen, MycoKey, Nøruplundvej 2, 8400 Ebeltoft, Denmark. Corresponding
author, e-mail & website: info@jenshpetersen.dk, www.mycokey.com.
Marie L. Davey, Department of Biosciences, University of Oslo, PO Box 1066
Blindern, 0316 Oslo, Norway; e-mail: m.l.davey@ibv.uio.no
Thomas Læssøe, Department of Biology/Natural History Museum of Denmark,
University of Copenhagen, Universitetsparken 15, 2100 København Ø; e-mail:
thomasl@bio.ku.dk
Introduction
One may think that the fungi of the well re-
searched Northern Europe can no longer offer
signifi cant surprises. But this is not so: my-
cologists and forayers continue to discover
remarkable new genera and species from this
area. Here, we describe a new clavarioid taxon
characterised by minute, somewhat typhuloid,
hairy fruit bodies on rotten bark. It is currently
known from several collections from Denmark
and Norway with the fi rst collection dating back
to 1995. It has been reported provisionally by
Jordal & Læssøe (2009), and has also featured
in various versions of MycoKey (Læssøe &
Petersen 2003). Initial attempts at sequenc-
ing material for phylogenetic reconstruction in
2005 were unsuccessful. Following successful
sequencing based on Norwegian material from
2011, a formal description can now be proposed.
Typhuloid fungi with or without sclerotia
have been monographed by Corner (1950, 1970)
and by Berthier (1976). In these works no fungi
with characters approaching those of the fun-
gus described below can be found. In addition,
the fungus was not covered in a comprehensive

2 KARSTENIA 54 (2014)
treatment of mainly Iberian cantharelloid and
clavariod fungi (Olariaga 2009) nor was any-
thing similar treated by Birkebak et al. (2013).
Material and methods
DNA sequencing and phylogenetic analysis
Three entire basidiomata were crushed in a single Eppen-
dorf tube using a Retsch ball mill (Retsch, Dusseldorf,
Germany) and their genomic DNA was extracted using a
modifi ed CTAB-based extraction protocol in accordance
to Murray & Thompson (1980) and Gardes & Bruns
(1993). The large subunit (LSU) region of ribosomal
DNA was amplifi ed with the forward and reverse primer
set LROR and LR7 (Vilgalys & Hester 1990) using illus-
tra PuReTaq Ready-To-Go PCR Beads (GE Healthcare,
Piscataway, NJ, USA) according to the manufacturer’s
instructions. Amplicons were sequenced with an ABI
3100 automated sequencer (Applied Biosystems Inc.,
Foster City, CA, USA) and the primers LROR, LR3R,
LR5 and LR7 (Vilgalys & Hester 1990). A data matrix
was assembled from LSU sequences of Hirticlavula el-
egans and members of the Clavariaceae sensu Dentinger
& McLaughlin (2006), as well as clavarioid fungi from
other groups (i.e. Mucronella, Macrotyphula, Typhula).
The matrix was aligned using MAFFT version 6.717 (Ka-
toh & Toh 2008), the resulting alignment was manually
verifi ed, and then Gblocks v0.91b (Castresana 2000) was
used to remove ambiguously aligned bases. The Bayesian
information criterion in jModelTest v0.1.1 (Guindon &
Gascuel 2003, Posada 2008) was used to determine the
best-fi t model of evolution, which was then implement-
ed in both maximum likelihood and Bayesian analyses.
GARLI v 1.0 (Zwickl 2006) was used to determine the
most likely tree and maximum likelihood bootstrap sup-
port for the dataset. Bayesian analyses were conducted us-
ing Mr.Bayes version 3.2 (Ronquist & Huelsenbeck 2003)
with two independent runs of four Markov Chain Monte
Carlo chains with 1.0 x 107 generations each, sampling
trees every 1000th generation. A fi nal standard deviation
of <0.01 for the split frequency was interpreted to refl ect
convergence. The fi rst 25% of sampled trees were dis-
carded as burn-in and posterior probabilities for each node
of the 50% majority rule consensus tree were recorded.
Morphology
The macroscopic images were taken with a Leica M 420
macroscope equipped with various analogue and digital
cameras. Pictures are stacked using Zerine Stacker and
post processed with Adobe Photoshop. Observations of
microscopic characters and micrographs were made with
Leica Orthoplan and Olympus BX50 microscopes. Mea-
surements were made on dead material in ca 5% ammo-
niacal congo red.
Results
The aligned matrix of LSU sequences included
94 taxa and 1288 characters, and the GTR+I+G
model was selected by jModelTest as the best-fi t
model of evolution for the data. Results of the
maximum likelihood bootstrap analyses and the
Bayesian inference are shown on the maximum
likelihood tree (-lnL -11336.51) (Fig. 1). Hirti-
clavula elegans is included within the well sup-
ported Clavariaceae sensu stricto (see Dentinger
& McLaughlin 2006). It is supported as sister to
a clade comprised of the genera Clavaria, Cama-
rophyllopsis, and Hodophilus (100% Bayesian
Posterior Probability, BPP and 78% Maximum
Likelihood Bootstrap Proportion, BP) that is
equivalent to Clade 4 as described in Birkebak et
al. (2013). Hyphodontiella is weakly supported
as sister to the Hirticlavula-Clavulariaceae clade
(59 BPP%, 53% BP). In all analyses, Hirticla-
vula is remote from all typhuloid taxa.
Hirticlavula J.H. Petersen & Læssøe, gen. nov.
MycoBank no.: MB 808059
Basidiomata club shaped, small, stipe hirsute,
without sclerotium. Hyphal system monomitic,
hyphae non-infl ated. Hairs unbranched, septate,
cyanophilous, up to 250 μm long, widening to-
wards the obtuse apex. Basidia holobasidia with-
out clamps. Spores hyaline, more or less cya-
nophilous, smooth.
Type: Hirticlavula elegans J.H. Petersen &
Læssøe
Hirticlavula elegans J.H. Petersen & Læssøe,
sp. nova – Figs. 2–5
MycoBank no.: MB 808060
Holotype: Norway. Vestfold. Sem, Gullk-
ronenes Naturreservat, Lille Gullkrone,
59.2854179 / 10.3826165, on Quercus bark on
the ground in mature oak woodland, 25.X.2013
Læssøe, JHP-13.364 (O, isotype in TUR).
PETERSEN ET AL. HIRTICLAVULA ELEGANS

KARSTENIA 54 (2014) 3
Fig 1. Maximum likelihood tree inferred from a dataset of large subunit rDNA sequences showing the placement of
Hirticlavula elegans among other clavarioid taxa. Support values are given above the branches as Bayesian posterior
probability/bootstrap proportion. Accession numbers for sequences retrieved from GenBank are given following the
sequence name.
PETERSEN ET AL. HIRTICLAVULA ELEGANS

4 KARSTENIA 54 (2014)
Basidiomata 800–1100 μm high, translucent
white, with a 600–800 μm long and 40–60
μm thick, hairy stem abruptly widening into a
230–260 x 90–200 μm large, solid, fertile head.
Dried material with a cream tinge and hardly
collapsed. Hyphal structure monomitic, without
clamps; hyphae in stem parallel, non-infl ated,
1–2 μm wide, more intertwined and gnarled at
insertion point. Hairs straight, pointing upwards
at an oblique angle, unbranched, slightly thick-
walled, remotely septate, 150–250 × 1.5–2.5
μm (base) and 3–4.5 μm thick at the rounded
± clavate apex; cells 10–30 μm long. Basidia
without clamps, cyanophilous, 14–22 × 3–5.5
μm, with four, curved and remarkably thin, 4–5
μm long sterigmata. Spores hyaline, cyanophi-
lous or with a few cyanophilous particles inside,
smooth, broadly ellipsoid, 4.5–6.5 × 3–4.2 μm,
Q= 1.2–1.5, mean Q= 1.36 (n = 15), apicu-
lus up to 0.8 μm long. Crystals not prominent.
The basidiomata do not emerge from a
sclerotium, but arise directly on bark or rare-
ly wood from hard wood trees, two collec-
tions on Quercus, another on Salix, one on
Corylus and one on undetermined hardwood.
Additional specimens studied: DENMARK.
Jutland. Skindbjerglund, south of Aalborg,
56.88889 / 9.923832, on bark on Quercus
trunk on the ground, 30.IX.1995 Vesterholt,
JHP-95.177 (C); Store Vildmose, Sandelsb-
jerg, 57.220723 / 9.759979, on rotten Salix bark
on the ground in moist Salix carr, 16.V.2005
Huhtinen JHP-05.067/JHP-05.068 (C, TUR).
NORWAY. Akershus. Frogn, Håøya, 59.104221
/ 10.212019, on rotten hard wood bark (Betula?),
2.IX.2011 Læssøe. Møre & Romsdal. Nes-
set, under Rangåfjellet, on decayed Corylus
wood, 62.461407 / 8.221528; MQ 59499 26079,
26.IX.2008 Læssøe & Nielsen TL-13513 (C).
Discussion
The distinguishing characters of this species are
its very small basidiomata and peculiar hairs. The
up to 1.1 mm high basidiomata are well outside
PETERSEN ET AL. HIRTICLAVULA ELEGANS
Fig. 2. Hirticlavula elegans (holotype). – Photo: J.H. Petersen.

KARSTENIA 54 (2014) 5PETERSEN ET AL. HIRTICLAVULA ELEGANS
Fig. 3. Basidiomata of Hirticlavula elegans. a = JHP-95.117, b = JHP-05.068, c-d = JHP-13.364 (holotype). – Photos:
J.H. Petersen
abc
d

6 KARSTENIA 54 (2014)PETERSEN ET AL. HIRTICLAVULA ELEGANS
abc
de
g
f
h
Fig. 4. Light microscope photographs of Hirticlavula elegans. a-c = JHP-95.117, d-h = JHP-13.364.
a = whole basidiome, b = head with basidia and spores, c = rounded apices of hairs, d = spiralling hyphae of stem base,
e = whole basidiome, f = hairs, g = hairs raising from stipe tissue, h = basidia and spores; a-g in cotton blue, h in water.
– Photos: J.H. Petersen & T. Læssøe.

KARSTENIA 54 (2014) 7PETERSEN ET AL. HIRTICLAVULA ELEGANS
Fig. 5. Hirticlavula elegans (JHP-95.117). a = whole ba-
sidiome, b = hairs, c = basidia, d = spores. – Drawings:
J.H. Petersen.
ab
c
d
the range of all other known members of the Cla-
variaceae sensu stricto but are found elsewhere
among more distantly related clavarioid fungi,
e.g. the genera Typhula, Pistillina and Pterula.
The hairs of Hirticlavula elegans originate from
the outer hyphae of the stipe and point upwards,
as if aiming to surround the fertile head. They are
not merely branched hyphae, as the sparsely sep-
tate, rather irregular hairs often seen on the stipes
of species of Typhula, Pistillina and Pterula, but
are highly specialised structures with numerous
septa and ending in a characteristic, rounded
apex. While the hairs of all other known cla-
varioid fungi narrow towards the apex, the hairs
of this species widen. Hirticlavula elegans is
strongly supported as sister to all other members
of Clavaria, Camarophyllopsis, and Hodophilus
in phylogenetic analyses of the LSU region, and
can be distinguished from these genera not only
based on its minute size and diagnostic hairs, but
also based on its ecology. While members of the
Clavariaceae sensu stricto can be classifi ed as bio-
trophic based on stable isotope analysis (Birke-
bak et al. 2013), H. elegans is lignicolous and
saprotrophic. Given that Hyphodontiella, anoth-
er wood-inhabiting genus within the clavarioid
clade, is identifi ed as sister to the Hirticlavula-
Clavariaceae clade, further multigene analysis of
the systematic positions within this group could
provide valuable insight into the evolution of nu-
tritional mode transitions in the clavarioid fungi.
It is rather surprising that Hirticlavula can
produce basidiomata from May through to
October. We do not know other tiny clavari-
oid fungi with such a long fruiting period. The
unique morphology, ecology, and phyloge-
netic position of this fungus all suggest that
it represents an undescribed genus closely
related to the Clavaria/Camarophyllopsis-
clade within the Clavariaceae sensu stricto.
Acknowledgements: Jan Vesterholt (†) and Seppo
Huhtinen are thanked for placing their material at our
disposal. Risken, Molde- og omegn Soppforening and
Wenche Johansen are thanked for inviting and hosting
TL at the Eikesdalen Foray. Ernest Emmett is thanked for
checking the English language.
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