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Phylogenetic relationships and morphological evolution in
Lentinus
,
Polyporellus
and
Neofavolus
, emphasizing southeastern Asian taxa
Jaya Seelan Sathiya Seelan
Biology Department, Clark University, 950 Main Street,
Worcester, Massachusetts 01610
Institute for Tropical Biology and Conservation (ITBC),
Universiti Malaysia Sabah, 88400 Kota Kinabalu,
Sabah, Malaysia
Alfredo Justo
Laszlo G. Nagy
Biology Department, Clark University, 950 Main Street,
Worcester, Massachusetts 01610
Edward A. Grand
Mahidol University International College (Science
Division), 999 Phuttamonthon, Sai 4, Salaya, Nakorn
Pathom 73170, Thailand
Scott A. Redhead
ECORC, Science & Technology Branch, Agriculture &
Agri-Food Canada, CEF, Neatby Building, Ottawa,
Ontario, K1A 0C6 Canada
David Hibbett
1
Biology Department, Clark University, 950 Main Street
Worcester, Massachusetts 01610
Abstract
:The genus
Lentinus
(Polyporaceae, Basi-
diomycota) is widely documented from tropical and
temperate forests and is taxonomically controversial.
Here we studied the relationships between
Lentinus
subg.
Lentinus
sensu Pegler (i.e. sections
Lentinus
,
Tigrini
,
Dicholamellatae
,
Rigidi
,
Lentodiellum
and
Pleuroti
and polypores that share similar morpholog-
ical characters). We generated sequences of internal
transcribed spacers (ITS) and partial 28S regions of
nuc rDNA and genes encoding the largest subunit of
RNA polymerase II (
RPB1
), focusing on
Lentinus
subg.
Lentinus
sensu Pegler and the
Neofavolus
group, combined these data with sequences from
GenBank (including
RPB2
gene sequences) and
performed phylogenetic analyses with maximum
likelihood and Bayesian methods. We also evaluated
the transition in hymenophore morphology between
Lentinus
,
Neofavolus
and related polypores with
ancestral state reconstruction. Single-gene phyloge-
nies and phylogenies combining ITS and 28S with
RPB1
and
RPB2
genes all support existence of
a
Lentinus
/
Polyporellus
clade and a separate
Neofavo-
lus
clade.
Polyporellus
(represented by
P. arcularius
,
P.
ciliatus
,
P. brumalis
) forms a clade with species
representing
Lentinus
subg.
Lentinus
sensu Pegler
(1983), excluding
L. suavissimus
.
Lentinus tigrinus
appears as the sister group of
Polyporellus
in the four-
gene phylogeny, but this placement was weakly
supported. All three multigene analyses and the
single-gene analysis using ITS strongly supported
Polyporus tricholoma
as the sister group of the
Lentinus/Polyporellus
clade; only the 28S rRNA
phylogeny failed to support this placement. Under
parsimony the ancestral hymenophoral configuration
for the
Lentinus/Polyporellus
clade is estimated to be
circular pores, with independent transitions to
angular pores and lamellae. The ancestral state for
the
Neofavolus
clade is estimated to be angular pores,
with a single transition to lamellae in
L. suavissimus
.
We propose that
Lentinus suavissimus
(section
Pleuroti
) should be reclassified as
Neofavolus suavissi-
mus
comb. nov.
Key words: Lentinus
sensu stricto,
Lentinus sua-
vissimus
, multigene phylogeny, PolyPEET, taxonomy
INTRODUCTION
Lentinus
Fr. is a widespread genus of wood-decaying
Agaricomycetes with tough basidiocarps, hyaline
spores and decurrent lamellae. Application of the
generic name
Lentinus
has been controversial (Pegler
1971, 1972, 1975, 1983a, b; Ku¨hner1980; Corner
1981; Pegler and Young 1983; Redhead and Ginns
1985; Singer 1986). A comprehensive world mono-
graph of
Lentinus
was published by Pegler (1983a),
but this concept of the genus is polyphyletic (Hibbett
and Vilgalys 1991, 1993; Hibbett et al. 1993a; Binder
et al. 2005; Binder et al. 2013).
Lentinus
sensu Pegler
(1983a) was subdivided into two subgenera,
Lentinus
subg.
Lentinus
and
Lentinus
subg.
Panus
,based
largely on anatomy of hyphal systems and hymeno-
phoral trama. Subgenus
Lentinus
included species
with skeleto-ligative hyphae with intercalary or termi-
nal branching, hyphal pegs (fascicles of sterile hyphae
protruding from the lamellae), hymenophoral trama
of descending, radiate or intermediate construction
and lacking metuloids and gloecystidia, whereas subg.
Panus
included species with skeletal hyphae (thick-
walled, typically unbranched), lacking hyphal pegs,
with metuloids and gloecystidia and hymenophoral
trama mostly of radiate construction. Subgenus
Lentinus
comprises six sections: sect.
Lentinus
sensu
Submitted 8 Apr 2014; accepted for publication 7 Jan 2015.
1
Corresponding author. E-mail: dhibbett@clarku.edu
Mycologia,
107(3), 2015, pp. 460–474. DOI: 10.3852/14-084
#2015 by The Mycological Society of America, Lawrence, KS 66044-8897
460
Pegler (eight species, including
L. crinitus
which was
accepted as lectotype by Pegler), sect.
Tigrini
Pegler
(six species, including
L. tigrinus
, which technically is
the correct lectotype of
Lentinus
[viz. Redhead and
Ginns 1985]), sect.
Dicholamellatae
Pegler (three
species), sect.
Rigidi
Pegler (five species), sect.
Lentodiellum
(Murr.) Pegler (four species) and sect.
Pleuroti
Sacc. (one species), and subgenus
Panus
includes nine sections: sect.
Pulverulenti
Fr. (three
species), sect.
Panus
(Fr.) Pegler (nine species), sect.
Cirrhosi
(two species), sect.
Velutini
Pegler (six
species), sect.
Gigantopanus
(Corner) Pegler (one
species), sect.
Squamosi
Fr. (six species), sect.
Tuberregium
(Singer) Pegler (four species), sect.
Prolifer
Pegler (four species) and sect.
Tenebrosi
Pegler (one species). The earliest acceptable lecto-
type by Clements and Shear (1931) for the generic
name
Lentinus
technically is
L. tigrinus
(viz. Redhead
and Ginns 1985), which would make
Lentinus
sect.
Tigrini
superfluous, and would require a new section-
al name for
Lentinus
sect.
Lentinus
sensu Pegler. For
the sake of convenience and consistency pending
a conservation, here we follow Pegler (1983a) and
accept as lectotype
L. crinitus
, which is in alignment
with most other classifications (see Redhead and
Ginns 1985).
Alternative generic classifications of species of
Lentinus
sensu Pegler (1983a) have been proposed
based on anatomy, mating systems (bipolar vs.
tetrapolar), decay types (white rot vs. brown rot),
nematode-trapping ability, hymenophore develop-
ment, post-meiotic nuclear behavior and molecular
phylogenies (Corner 1981; Redhead and Ginns 1985;
Singer 1986; Hibbett et al. 1993a, b; Hibbett and
Vilgalys 1993; Hibbett and Thorn 1994; Thorn et al.
2000; Hibbett and Donoghue 2011; Karunarathna
et al. 2011a; Binder et al. 2013). The "lentinoid fungi"
now are understood to be distributed across six
genera, including
Lentinus
and
Panus
(Polyporales),
Pleurotus
and
Lentinula
(Agaricales) and
Neolentinus
and
Heliocybe
(Gloeophyllales), as outlined by Red-
head and Ginns (1985) and refined by segregating
Panus
by Hibbett et al. (1993b). Excluding the five
other genera listed above, in the present study
"
Lentinus
" refers to
Lentinus
subg.
Lentinus
sensu
Pegler, which is roughly equivalent to
Lentinus
sensu
Corner (1981) and
Panus
sects.
Pleuroti
(Sacc.)
Singer and
Criniti
(Sacc.) Singer, both sensu Singer
(1986).
Lentinus
and certain polypores have a close re-
lationship based on their morphological character-
istics. Both genera contain dimitic and amphimitic
hyphal systems, cylindrical to subellipsoid and smooth
inamyloid basidiospores and hyphal pegs, which place
them in the Polyporaceae as traditionally defined
(Corner 1981, Pegler 1983a, Gilbertson and Ryvar-
den, 1987). The sub-poroid hymenophore of some
Lentinus
species also suggests a possible polyporoid
ancestry (Pegler 1983a). Developmental studies in
L.
tigrinus
have revealed the formation of both lamellae
and tangential ‘‘cross bridges’’, which might be
homologous to the tangential hymenophoral ele-
ments in polypores with angular pores such as
P.
arcularius
(Hibbett et al. 1993a).
Molecular studies have indicated that
Polyporus
is
polyphyletic and that species of
Lentinus
and
Polyporellus
(pileate-stipitate polypores, with angular
or circular pores, and relatively ephemeral fruiting
bodies) form a clade (Kru¨ ger 2002; Kru¨ger and
Gargas 2004; Grand 2004, 2011; Sotome et al. 2008;
Binder et al. 2013). The type species of
Polyporus
has
been interpreted variously as
P. brumalis
(Clements
and Shear 1931, Kru¨ger and Gargas 2004),
P.
squamosus
(Ryvarden 1978, Ryvarden and Gilbertson
1987) or
P. tuberaster
(Overholts 1953, Cunningham
1965, Singer 1986, Silveria and Wright 2005, Sotome
et al. 2008). Ryvarden (1991) preferred
P. tuberaster
as
lectotype of
Polyporus
while accepting
P. brumalis
as
the lectotype of
Polyporellus
. For the purpose of our
discussion we adopt
P. tuberaster
as type of
Polyporus
until this nomenclatural debate is settled and we
accept
P. brumalis
as type of
Polyporellus
.
Six prior phylogenetic studies using different genes in
Lentinus
,
Polyporellus
,
Neofavolus
(formerly known as
Polyporus
) and related polypores are summarized (TABLE
I). Five studies used sequences of the nuc rDNA internal
transcribed spacer regions 1 and 2 (ITS) and 28S
sequences for limited sections of
Lentinus
.Grandetal.
(2011) included representative members of five sections,
except section
Pleuroti
but analyzed only ITS sequences.
Meanwhile, Sotome et al. (2008) used 28S,
RPB2
and
ATP6
but sampled only two sections of
Lentinus
,along
with numerous polypores.
Species of
Lentinus
occur in boreal, temperate,
subtropical and tropical regions (Pegler 1983a, b;
Corner 1981). They play an important role in natural
ecosystems as wood decomposers and show potential
for seasonal food, medicine and alternative income
mainly in southeastern Asia and southern Africa
(Chin 1981, Watling 1993, Mossebo 2002, Bayramoglu
et al. 2006, Sysouphanthong et al. 2010, Njouonkou et
al. 2013a).
Lentinus
is widely documented within
southeastern Asia, including Indonesia, Thailand,
Laos, Peninsular Malaysia, Borneo, Philippines, China
and India (Manimohan and Leelavathy 1995, Huang
1998, Manimohan et al. 2004, Sumaiyah et al. 2007,
Nazura et al. 2010, Somchai 2010, Sudirman 2010,
Bolhassan et al. 2013). In Malaysia 20 species of
Lentinus
have been reported (Chipp 1921, Newsam et
al. 1967, Lim 1972, Corner 1981, Pegler 1983a,
SATHIYA SEELAN ET AL.: LENTINUS,POLYPORELLUS,NEOFAVOLUS 461
Oldridge et al. 1986, Lee et al. 1995, Salmiah and
Thillainathan 1998, Salmiah and Jones 2001, Noorli-
dah et al. 2005, Sumaiyah et al. 2007, Bolhassan et al.
2013). Seven new species of
Lentinus
have described
in the past 10 y (
L. parvus, L. bambusinus, L.
megacystidiatus, L. concentricus, L. roseus, L. alpacus,
L. cystidiatus
), all without molecular data. However
these taxa have greater resemblance to
Panus
based
on reported morphological characters (Arun Kumar
and Manimohan 2005, Manimohan et al. 2004,
Sumaiyah et al. 2007, Karunarathna et al. 2011b,
Senthilarasu et al. 2012, Drechsler-Santos 2012,
Njouonkou et al. 2013b) and new sequence data that
will be reported elsewhere (Seelan et al. unpubl).
This study aims to assess the limits of
Lentinus
and the
pattern of transitions between pores and gills. For this
purpose it is necessary to sample all sections of
Lentinus
sensu Pegler and a diverse assemblage of
polypores. By focusing on collections from Malaysia
this study also seeks to provide a framework for
taxonomic and phylogenetic studies of
Lentinus
from
southeastern Asia.
MATERIALS AND METHODS
Collections.—
They were made during the May-Jun 2009,
2010 rainy seasons and the Nov-Jan 2010, 2011 rainy seasons
from Sabah Park in northern Borneo, Malaysia. Fruiting
bodies of
Lentinus
were collected from highland and
lowland dipterocarp and heath, mainly in primary and
secondary forests around Kinabalu Park.
Lentinus sajor-caju
collections also were made on Gaya Island, which is about 6
km from the mainland of Kota Kinabalu, Sabah. Locality
information for the new specimens or cultures used for the
DNA extraction and sequencing in this study are presented
(TABLE II). All specimens of
Lentinus
were identified with
reference to (Pegler 1983a) and (Corner 1981).
Additional materials were collected in the Forest Research
Institute of Malaysia (FRIM) and Sungkai Wildlife Reserve
Forests (PERHILITAN) in peninsular Malaysia, mainly from
lowland dipterocarp forest. Specimens of
Lentinus suavissimus,
which is the only species in
Lentinus
sect.
Pleuroti
sensu Pegler
(1983a) and
Neofavolus sp.
(
Polyporus alveolaris
) were collected
on Chena Lakes Nature Trails, Alaska, and the Adirondack
Mountains, New York). A single specimen of
Lentinus crinitus
(AJ527) was collected at Virgin Islands National Park (St John,
US Virgin Islands). Specimens were dried and kept in
polyethylene bags with silica gel. One specimen of
Lentinus
squarrosulus
was obtained from the personal collection of
Professor Yu Cheng Dai (Academy of Sciences, China).
Specimens of
Polyporus arcularius
(DSH92-132),
Lentinus
crinitus
(DSH92N43C) and
Lentinus tigrinus
(DSH92D787)
were accessioned from the Clark University Herbarium.
Additional specimens obtained from the BIOTEC Bangkok
Herbarium (BBH), Thailand, and the Royal Botanical Gardens,
Kew (K), UK. Duplicate specimens of Malaysian collections
were deposited at the Sabah National Park Herbarium (SNP)
and at the Institute for Tropical Biology and Conservation,
Borneensis herbarium (BORH) at Universiti Malaysia Sabah.
Two cultures of
Polyporus ciliatus
were obtained from the
University of Tennessee Herbarium (TENN).
DNA extraction, PCR and sequencing.—
Cultures of
P.
ciliatus
were maintained 2–3 wk at 25–30 C on solid media
(MEA: 20 g malt extract, 0.5 g yeast extract, 20 g agar in1 L
water). When plates were covered with new mycelium, the
tissue was scraped with sterile scalpels and transferred to
a 1.5 mL microtube and ground with a sterile plastic pestle.
In the case of specimens, a small portion of the fruiting
TABLE I. Major molecular systematic studies for
Lentinus
,
Polyporellus
and related polypores showing the numbers of species
and individual (in parentheses) samples
Lentinus
subg.
Lentinus
/
section
a
Hibbett et al.
(1993)
Rolen, Tage
(2001)
Kru¨ ger et al.
(2008)
Sotome et al.
(2008)
Grand et al.
(2011)
Sotome et al.
(2013) This study
Lentinus
1(1) 3(60) 3(31) 3(8)
Tigrini
1(1) 1(1) 1(7) 1(1) 2(31) 1(1) 1(2)
Dicholamellatae
1(1) 1(4)
Rigidi
3(4) 2(3) 3(12)
Lentodiellum
2(4) 2(2) 2(2)
Pleuroti
1(10)
Related genera
Polyporellus
1(1) 6(56) 3(7) 4(12) 1(1) 4(9)
Neofavolus
1(1) 1(1) 3(15) 3(18)
Other Polyporales 6(6) 17(19) 27(64) 12(20) 40(50)
Gene (s) ITS, LSU ITS, LSU ITS, LSU LSU,
rpb2
,
ATP6
ITS ITS, LSU ITS, LSU,
rpb1
,
rpb2
a
Section
Lentinus
includes (
L. bertieri, L. crinitus
and
L. swartzii
); section
Tigrini
includes
L.tigrinus
and
L. glabratus
;
section
Rigidi
includes
L. polychrous, L. sajor-caju
and
L. squarrosulus
, section
Dicholamellatae
includes
L. badius
and section
Lentodiellum
includes
L. striatulus
and
L. scleropus. Polyporellus
group includes
Polyporus arcularius, P. brumalis, P. ciliatus
and
P. tricholoma
.
462 MYCOLOGIA
body was ground with liquid nitrogen. Cell lysis proceeded
for 1h at 65 C with the addition of 600 mL extraction buffer
(50 mM EDTA, 50 mM Tris-HCl, 3%SDS, pH 8). Cell
debris, polysaccharides and proteins were separated from
aqueous DNA portions through two purification steps with
equal volumes of phenol:chloroform (1 : 1) and chloro-
form:isoamylalcohol (24 : 1). Total DNA was precipitated
with the addition of 3 M sodium acetate (0.1 vol.) and
isopropanol (0.54 vol.) and incubated 30–60 min at 220 C.
The DNA pellets were washed in 1 mL 70%EtOH, dried at
65 C for 5–15 min and resuspended in 100 mL sterile H
2
O.
Dilutions of the original DNA extraction, usually 1 : 10–
1 : 500, were used in the polymerase chain reaction (PCR)
amplification. DNA was extracted from dried specimens
(approximately 0.2 g) with the EZNA Fungal DNA Kit
(Omega Bio-Tek, Norcross, Georgia) and following Hosaka
and Castellano (2008).
Three regions were sequenced, including those encoding
the internal transcribed spacers (ITS 5ITS1 +5.8S +ITS2) and
partial large subunit of 28S nuclear ribosomal RNA (28S) and
subunit 1 of RNA polymerase II (
RPB1
)(TABLE II). The ITS
(approx. 600–700 bp, including 157 bp 5.8S rRNA coding
TABLE II. Taxon sampling, geographic location, specimen-voucher information and GenBank accession numbers
GenBank accession Nos.
Species
Specimen
voucher/cultures Location ITS nLSU
RPB1
Lentinus badius
JS0094 Crocker Range Park, Borneo KP283478 KP283512 KP325691
L. badius
JSKT5858 Sungkai, Perak, Peninsular Malaysia KP283479 KP283513 KP325690
L. badius
DED07668 Phuket, Thailand KP283480 KP283518 KP325692
L. badius
PU00436 Nakhon Ratchasima, Thailand KP283481
L. crinitus
DSH9243C Costa Rica KP283495 KP283523 KP325687
L. crinitus
AJ527 St John, US Virgin Islands KP283521 KP325688
L. polychrous
AH00024 Phang-Nga, Thailand KP283485
L. polychrous
JS00054 Malungong, Borneo KP283486
L. polychrous
KM141387 Thailand KP283487 KP283514
L. sajor-caju
FRI62056 FRIM, Peninsular Malaysia KP283492 KP283509 KP325677
L. sajor-caju
SNP24989 Gaya Island, Borneo KP283493 KP283510 KP325678
L. sajor-caju
JS0056 Kinabalu Park, Borneo KP283494 KP283511 KP325679
L. squarrosulus
CUI6513 Yunnan, China KP283482 KP283516 KP325680
L. squarrosulus
FRIM4180 Pahang, P. Malaysia KP283483 KP283517 KP325682
L. squarrosulus
BORHF0009 Sorinsim, Borneo KP283484 KP283515 KP325681
L. tigrinus
DSH92D787 North Carolina, USA KP283488 KP325689
L. suavissimus
ADD7 Adirondacks State Park, NY, USA KP283501 KP283527 KP325694
L. suavissimus
LE0791625 Germany KP283500
L. suavissimus
TENN19955 Great Smoky Mountain
National Park, TN, USA
KP283504
L. suavissimus
TENN11096 Great Smoky Mountain
National Park, TN, USA
KP283505 AY615970
L. suavissimus
TENN13225 Great Smoky Mountain
National Park, TN, USA
KP283503
L. suavissimus
TENN13316 Quebec, Canada KP283502
L. suavissimus
TENN11330 France KP283498 AY615969
L. suavissimus
TENN11129 France KP283499
L. suavissimus
LE127 Russia KP283497
L. suavissimus
DSH2011 (AL57) Fairbanks, AK, USA KP283496 KP283525 KP325693
Polyporus arcularius
DSH92-132 Taman Negara, Pahang, P. Malaysia KP283489 KP283522 KP325686
Polyporus brumalis
PB4 (EP4) Worcester, MA, USA KP283490 KP283519 KP325685
Polyporus brumalis
PB1 Newton hill, MA, USA KP283491 KP283520
Polyporus ciliatus
TFB10167 (SP3) Roskilde Amt, Denmark KP325684
Polyporus ciliatus
TFB7480 (SP28) Finland KP325683
Neofavolus
sp. MA672 Worcester, MA, USA KP283506 KP283524 KP325696
Neofavolus
sp. ADD5 Adirondacks State Park, NY, USA KP283508
Neofavolus
sp. SAV10 Savoy Mountain, Massachusetts, USA KP283507 KP283526 KP325695
Abbreviations: JS, author’s collection number; DED, PU, AH, Biotech Bangkok herbarium; SNP, Sabah National Park
herbarium; BORHF, BORNEENSIS herbaria collections; LE, Leiden Herbarium; SAV, MA, ADD, EP,PB, DSH, AJ, Clark
University Herbaria collections (different letters indicate different collectors or area collected); SP, Culture collection from
University of Tennessee Herbarium (TENN); CUI, Collection from Professor Yu Cheng Dai (Academy of Sciences, China);
FRI, Forest Research Institute of Malaysia (FRIM) Herbaria; K(M), Royal Botanical Garden Kew, London.
SATHIYA SEELAN ET AL.: LENTINUS,POLYPORELLUS,NEOFAVOLUS 463
region) was amplified with the primer pair ITS-1F/ITS4 (White
et al. 1990, Gardes and Bruns 1993) and the partial 28S region
(approx. 1300 bp) was amplified with the primer pair LR0R/
LR7 (Vilgalys and Hester 1990). For ribosomal DNA markers
this PCR protocol was used: (i) initial denaturation at 95 C for 2
min, (ii) denaturation at 94 C for 45 s, (iii) annealing at 50 C for
1 min 10 s, (iv) extension at 72 C for 2 min, (v) repeat for 34
cycles starting at step 2, (vi) leave at 72 C for 10 min (Binder et
al. 2010). Sequencing primers for ITS and 28S were the same
used for PCR and in the case of 28S with two additional internal
primers: LR3R and LR5 (Vilgalys and Hester 1990). A part of
the
RPB1
gene between conserved domains A and C of
RPB1
(approx. 1400 bp) was amplified with the primer pair RPB1-Af
and RPB1-Cr (Stiller and Hall 1997, Matheny et al. 2002). In
some cases the primer RPB1
-
2.2f (Binder et al 2010) was used as
an alternative to RPB1-Af, producing a slightly shorter product
(approx. 1000 bp). Additional sequencing primers were: RPB1-
2f, RPB1-2.1f, RPB1-2.2f and RPB1-2.1r (Frøslev et al. 2005).
For
RPB1
the following touchdown PCR protocol was
used: (i) initial denaturation at 94 C for 2 min, (ii)
denaturation at 94 C for 40 s, (iii) annealing at 60 C for 40 s
(minus 1 C per cycle), (iv) extension at 72 C for 2 min, (v)
repeat for nine cycles starting at step 2, (vi) denaturation at
94 C for 45 s, (vii) annealing at 53 C for 1 min 30 s, (viii)
extension at 72 C for 2 min, (ix) repeat for 36 cycles starting
at step 6, (x) leave at 72 C for 10 min. The amplification
products for all markers were sequenced with BigDye 3.1
terminator sequencing chemistry (Applied Biosystems,
Foster City, California) and run on an Applied Biosystems
3130 Genetic Analyzer at Clark University or analyzed by
Macrogen Inc. Rockville, Maryland.
Sequence alignment and phylogenetic analyses.—
In addition
to the sequences generated here, 72 were retrieved from
GenBank and come mainly from Sotome et al. (2013) and
Binder et al. (2013). Accession numbers of ITS, 28S,
RPB1
and
RPB2
sequences used in the analysis are provided (SUPPLE-
MENTARY TABLE I). Sequences were aligned with either
MUSCLE 3.8 (Edgar 2004), in the case of protein-coding
genes, or PRANK 130820 (Loytynoja and Goldmann 2008) for
the ITS sequences, becausee it has been shown to outperform
most other alignment algorithms for aligning ITS sequences
(Nagy et al. 2012). MUSCLE was launched with default
parameters, whereas in the case of PRANK we selected the +F
option. The alignments were manually corrected with Mac-
Clade 4.08 (Maddison and Maddison 2002; http://macclade.
org/). Overly variable intron regions from the
RPB1
and
RPB2
alignments were excluded. For the combined datasets each
marker was aligned separately and then concatenated in
MacClade. Four datasets were assembled for the phylogenetic
analyses (Figs. 1, 2; SUPPLEMENTARY FIGS.1,2),andsingle-gene
phylogenies provided (SUPPLEMENTARY FIGS.3–6).
Phylogenetically informative indels in the ITS alignment
were recoded as a matrix of binary characters and were
appended to the end of the concatenated matrix for the
Bayesian analyses. Indels were coded with the simple indel
coding algorithm (Simmons and Ochoterena 2000) using
the gapcode.by script (http://www.bioinformatics.org/
,rick/software.html).
Two phylogenetic analyses were performed in all the
datasets: (i) Maximum likelihood analyses (ML) were run in
RAxML 7.2.8 (http://phylobench.vitaleit.ch/raxmlebb/
index.php; Stamatakis et al. 2008) under the GTRGAMMA model
with 100 rapid bootstrap replicates. (ii) Bayesian analyses (BI)
were run with MrBayes 3.1.2 (Ronquist and Huelsenbeck
2003) at the Cipres Science Gateway (Miller et al 2010; http://
www.phylo.org/) for 10 000 000 generations, under a GTR
model, modeling rate heterogeneity by a discrete gamma
distribution. MrBayes was launched with two runs with four
chains each and trees were sampled every 100 generations.
For both ML and BI we partitioned the dataset into single
genes and estimated a partitioned model with unlinked model
parameters between the partitions. The recoded indels were
modeled with the likelihood model for binary characters
implemented in MrBayes. The burn-in was determined by
checking the convergence of log-likelihood values in Tracer
1.5 (Rambaut and Drummond 2007), and the first 30 000 trees
from each run were discarded. The remaining 140 000 trees
were used to compute a 50%majority rule consensus tree and
estimate posterior probabilities(PP) with the SumTrees script
of the Dendropy package (Sukumaran and Holder 2010).
Convergence of log likelihood scores (2Ln) was assessed with
TRACER 1.4 (Rambaut and Drummond 2007) and stationar-
ity was assumed when a stable equilibrium value was reached
(Ronquist and Huelsenbeck 2003). Individual nodes were
considered well supported when ML bootstrap values (BS)
were at least 70%and when PP values were at least 0.95.
Ancestral state reconstruction (ASR).—
To reconstruct the
evolution of hymenophoral transitions within the
Lentinus
and
Neofavolus
clades, we used parsimony and ML
optimization, implemented in MESQUITE 2.75 (Maddison
and Maddison 2011). Hymenophoral forms within
Lentinus
,
Neofavolus
and other remaining core polyporoid members
were assigned as discrete unordered character states:
Odontoid and tuberculate (outgroup) 50, circular pores
51, daedaleoid 52, angular pores 53, lamellate 54, sub-
poroid lamellae 55 and round to angular pores 56
(SUPPLEMENTARY TABLE II). Character coding 5 represents
the sub-poroid construction that arises from the base of the
stipe, which was considered as an important character to
differentiate
L. tigrinus
and
L. suavissimus
. Meanwhile,
coding 6 represents polymorphic characters within
Trametes
villosa
,
T. polyzona
,
Datronia scutellata
and
P. tricholoma
. All
character coding was produced based on Ryvarden (1991).
Ancestral states were estimated with 1000 rooted trees
drawn randomly from the post burn-in tree pool derived
from the MrBayes analysis of the four-gene 99 taxa dataset.
In MESQUITE, the option TRACE CHARACTER OVER TREES was
selected to reconstruct ancestral character states assuming
an Mk1 class model and unordered characters. Parsimony
reconstructions were optimized with the MOST PARSIMONI-
OUS RECONSTRUCTIONS (MPR) option.
RESULTS
Seventy new sequences were generated: 31 (ITS), 19
(28S) and 20 (
RPB1)
. Single-gene phylogenies from
ML analyses of the ITS, 28S,
RPB1
and
RPB2
datasets
464 MYCOLOGIA
indicated no strongly supported conflict, and are
provided (SUPPLEMENTARY FIGS. 3–6). Phylogenetic
relationships among members of the
Lentinus
and
Neofavolus
clades were estimated with three multi-
gene datasets, ITS+28S+
RPB1
+
RPB2
, ITS+28S+
RPB2
and ITS+28S+
RPB1
, and an ITS only dataset. ML and
BI trees generated from all analyses were largely
congruent; only the ML tree topologies are illustrated
here, and conclusions are based primarily on the
topology of the best tree from the ML analysis of the
combined ITS+28S+
RPB1
+
RPB2
dataset (FIG. 1). A
detailed analysis of relationships in the
Neofavolus
clade was produced using ITS alone (FIG.2). A
comparative overview of the different datasets used
for the phylogenetic analyses is provided (TABLE III),
and the alignments were deposited in TreeBASE
(http://purl.org/phylo/treebase/phylows/study/
TB2:S16854). Most available sequences from Gen-
Bank for
Lentinus
and
Neofavolus
were included
(SUPPLEMENTARY TABLE I), except an ITS sequence
deposited by Grand et al. (2011) GU207275 labeled as
Lentinus badius
, which was placed in the core
polyporoid clade but not in the
Lentinus
group in
preliminary analyses (not shown).
Single-gene phylogenies and phylogenies combin-
ing ITS and 28S with
RPB1
or
RPB2
genes all support
existence of a
Lentinus
/
Polyporellus
clade and a sepa-
rate
Neofavolus
clade (FIGS.1,2;SUPPLEMENTARY
FIGS. 1–6).
Polyporellus
(represented by
P. arcularius,
P. ciliatus
and
P. brumalis
) nested within a paraphy-
letic assemblage of species representing
Lentinus
subg.
Lentinus
sensu Pegler (1983) excluding
L.
suavissimus
.
Lentinus tigrinus
appears as the sister
group of
Polyporellus
in the four-gene phylogeny, but
this placement is weakly supported (FIG. 1). All three
multigene analyses and the single-gene analyses using
ITS and
RPB2
strongly support
Polyporus tricholoma
as
the sister group of
Lentinus
/
Polyporellus
; only 28S
fails to support this placement (
P. tricholoma
was not
sampled for
RPB1
;SUPPLEMENTARY FIG. 4).
Five sections of
Lentinus
sensu Pegler (1983) are
represented in the
Lentinus/Polyporellus
clade: sections
Rigidi
(
L. squarrosulus, L. polychrous, L. sajor-caju
),
Lentodiellum (L. scleropus, L. striatulus)
,
Lentinus (L.
crinitus, L. swartzii, L. bertieri), Dicholamellatae (L.
badius)
and
Tigrini (L. tigrinus)
.However
Lentinus
sect.
Pleuroti
, represented by
L. suavissimus (5
Neofavolus suavissimus)
, is strongly supported as
amemberofthe
Neofavolus
clade in all multigene
and single-gene analyses (except the single-gene
analysis of
RPB2
, which did not include
L. suavissi-
mus
)(FIGS.1, 2; SUPPLEMENTARY FIGS.1–6). All
datasets with appropriate sampling place
L. suavissi-
mus
as the sister group of
N. mikawai
(FIGS.1, 2;
SUPPLEMENTARY FIGS.1–6). Results from the
ITS+28S+
RPB1
+
RPB2
dataset are in general agree-
ment with analyses of Grand et al. (2004, 2011) and
Sotome et al. (2008).
A single-gene phylogeny of the
Neofavolus
group
was constructed with 33 ITS sequences, including 20
sequences from Sotome et al. (2013), focusing on the
position of
L. suavissimus
(5
N. suavissimus
, which
was not sampled by Sotome; FIG. 2). The ML analysis
provides strong bootstrap support for monophyly of
10 individuals of
L. suavissimus
(FIG. 2). However the
L. suavissimus
group is divided into two distinct
lineages with a strong geographic pattern; one
includes collections from Tennessee, New York and
Quebec, and the other includes collections from
France, Germany, Russia and Alaska (FIG.2). In addition
three unidentified collections of
Neofavolus
from
NewYorkandMassachusettswereplacedasapara-
phyletic assemblage, with a nested clade of Japanese
collections of
N. alveolaris
and
N. cremeoalbidus
from
TABLE III. Phylogenetic datasets used in this study
Dataset Ingroup sequences Outgroup
Parsimony-
informative
characters
(including gap)
Aligned
length (bp)
nLSU+ITS+
rpb1
+
rpb2
40
Lentinus
/
Polyporellus
taxa
and 59 other Polyporales
Dendocorticium sulphurellum
and
Lopharia cinerascens
(Polyporales)
3728 6402
nLSU+ITS+
rpb1
40
Lentinus
/
Polyporellus
taxa
and 58 other Polyporales
Dendocorticium sulphurellum
and
Lopharia cinerascens
(Polyporales)
2334 4553
nLSU+ITS+
rpb2
37
Lentinus
/
Polyporellus
taxa
and 58 other Polyporales
Dendocorticium sulphurellum
and
Lopharia cinerascens
(Polyporales)
2272 5025
ITS 10
L. suavissimus
taxa, 18
Neofavolus
and 5
Favolus
taxa
Favolus
brasilliensis
(Polyporales)
238 711
SATHIYA SEELAN ET AL.: LENTINUS,POLYPORELLUS,NEOFAVOLUS 465
FIG. 1. Phylogenetic relationships of members of the
Lentinus
and
Neofavolus
clades inferred from 28S, ITS,
RPB1
and
RPB2
sequences. Topology from ML analysis. Support values along branches are from ML bootstrap ($70) and BI analyses (PP $0.95)
respectively. Symbols on branches indicate transitions in hymenophoral form estimated with parsimony. Sections of
Lentinus
sensu
Pegler (1983) are indicated. Hymenophore character states occurring within
Lentinus
,
Polyporellus
and
Neofavolus
are illustrated.
466 MYCOLOGIA
FIG. 2. ML analysis of the ITS dataset of the
Neofavolus
clade. Support values along branches are from ML bootstrap ($70)
and BI analyses (PP $0.95), respectively.
SATHIYA SEELAN ET AL.: LENTINUS,POLYPORELLUS,NEOFAVOLUS 467
Sotome et al. (2013). The topology of the
Neofavolus
clade is consistent in the ITS and multigene analyses
(FIGS.1,2).
Ancestral-state reconstruction suggests a complex
pattern of transitions between round pores, angular
pores and lamellae. Under ML the pattern of
hymenophoral transitions is largely equivocal (results
not shown). Under parsimony the ancestral hymeno-
phoral configuration for the
Lentinus
/
Polyporellus
clade is estimated to be circular pores, with in-
dependent transitions to angular pores and lamellae
(FIG. 1). The ancestral state for the
Neofavolus
clade is
estimated to be angular pores, with a single transition
to lamellae in
L. suavissimus
(5
N. suavissimus
).
TAXONOMY
Neofavolus suavissimus
(Fr.) J. S. Seelan, Justo and
Hibbett, comb. nov.
Lentinus suavissimus
Fries., Synopsis Generis Lentinorum
13. 1836. Basionym.
;
Pocillaria suavissima
(Fr.) Kuntze,
Revisio generum
plantarum
2: 866 (1891)
;
Hemicybe suavissima
(Fr.) P. Karst.: 249. 1897.
;
Panus suavissimus
(Fr.) Singer, Lilloa 22: 274. 1951.
5
Lentinus anisatus
Henn., Verhandlungen des
Botanischen Vereins der Provinz Brandenburg
39: 95. 1898.
Notes.
Four individuals of
L. suavissimus
(L09791624, L09791625, TMI18871, DSH2011) con-
formed to Pegler’s (1983) description of the species.
However one collection of
L. suavissimus
differed
from the description in Pegler as follows: Fruiting
bodies of collection ADD7 were slightly larger than
those reported for
L. suavissimus
by Pegler (pileus 3–
8 cm diam, stipe 1–3 cm 35–8 mm in ADD7, vs.
pileus 0.5–5 cm diam, stipe 0.5–2 cm 34–6 mm). The
pileus of ADD7 was white when fresh, whereas Pegler
(1983) reported that pileus of
L. suavissimus
is yellow
to tawny ochraceous (FIG. 3A, B). The pale appear-
ance of ADD7 might be due to rain. In addition the
stipe base is red in ADD7, which has not been noted
in
L. suavissimus
(FIG.3A).
Lentinus suavissimus
is uncommon, but it has been
widely reported (sometimes under the synonyms
Poccularia haematopus
[Berk.] Kuntze,
Panellus hae-
matopus
[Berk.] Murr. and
Lentinus haematopus
Berk.) in North America, including Canada (Ontario,
Quebec, Saskatchewan) and USA (Maine, Michigan,
New Hampshire, New York, North Carolina, Tennes-
see, Vermont, Virginia) (Murrill 1915, Kauffman 1918,
Mains et al. 1939, Bigelow, 1959, Bigelow and Barr
1962, Miller and Manning 1976, Pomerleau 1980,
McNeil 2006), Europe, including Austria, Czech Re-
public, Denmark, Estonia, Finland, France, Germany,
Latvia, Norway, Poland and Sweden (Pila´t, 1946, Pegler
1983a, Knudsen et al. 2012), and Japan (Kobayashi
2007). The precise locality of LE127 is not known (but
might be Russia). Specimen DSH2011 from Alaska
represents a significant westward range extension for
the species in North America.
Specimens examined.
UNITED STATES. New York: Adir-
ondack mountains, 44u069450N73u559260W, 800 ft. On dead
wood, 15 Oct 2012, Jaya Seelan (ADD7); Alaska: Fairbanks,
on dead branch, 06 Jul 2011, David Hibbett, (DSH2011);
GERMANY: Bayern: Pensenberg. On dead branch, 24 Aug
1960, Donk, MA (L0791625).CZECH REPUBLIC: Bohe-
man, Sobeslav, South of Bohemia. On dead log, 06 Aug
1958, Kotlaba, F (L0791624). JAPAN. Mount Otyonosen,
Tottori. On fallen branch, 02.10.1994, Nagasawa, E
(TMI18871).
DISCUSSION
Relationships between the agaricoid genus
Lentinus
and certain polypores have long been suspected
based on anatomical features (Corner 1981, Pegler
1983a) and phylogenetic analyses with scattered
sampling of both genes and species (Hibbett et al.
1993, Tage Roland 2001, Kru¨ger et al. 2008, Sotome
et al. 2008, Grand et al. 2011, Sotome et al. 2013).
The present study includes the most comprehensive
phylogenetic analysis of
Lentinus
so far, with a focus
on southeastern Asian taxa. Most species of
Lentinus
subg.
Lentinus
sensu Pegler (1983a) form a mono-
phyletic group along with the pileate-stipitate
Poly-
porellus
(FIG. 1), but
Lentinus suavissimus
is not in
this group; the new combination
Neofavolus suavissi-
mus
is proposed. Relationships among the species of
Polyporellus
,which form a strongly supported clade,
and
Lentinus
sects.
Lentinus, Rigidi, Lentodiellum,
Dicholamellatae
and
Tigrini
sensu Pegler are not well
resolved (FIG. 1). The taxonomic disposition of the
Lentinus
/
Polyporellus
clade will await improved phy-
logenetic resolution, perhaps from genomic analyses
(complete genomes are available for
L. tigrinus
and
P. arcularius
). If the topology of the four-gene tree is
upheld, one option would be to combine
Polyporellus
into
Lentinus
. In the meantime we discuss
Lentinus
and
Polyporellus
as separate genera.
There have been parallel transformations between
angular pores and sub-poroid lamellae in the
Lentinus
/
Polyporellus
clade and the
Neofavolus
clade
(FIG.1).In
Neofavolus
the topology suggests one most
parsimonious reconstruction, implying derivation of
sub-poroid lamellae from angular pores (FIG.1).The
hymenophore of
N. suavissimus
is sub-poroid only at
the apex of the stipe, whereas in
L. tigrinus
the
hymenophore is sub-poroid across the entire width of
the pileus (FIG. 1; Hibbett et al. 1993a). These
structural differences reflect the convergent origins
of sub-poroid lamellae from poroid ancestors in
468 MYCOLOGIA
Neofavolus
and the
Lentinus
/
Polyporellus
clade
(Pegler 1983a, Hibbett et al. 1993a).
The precise pattern of transformations in hymeno-
phore configurations in the
Lentinus
/
Polyporellus
clade is not well resolved, in part due to uncertainty
about the branching order at the base of the clade.
Polyporus tricholoma
, which is reported as having
circular or angular pores, is strongly supported as
FIG.3.
Neofavolus suavissimus (5L. suavissimus)
.A,B.
Neofavolus suavissimus
from Adirondack Park, New York (ADD7,
Photo by Jaya Seelan). C, D. Sub-poroid hymenophore in young basidiocarp (Photo by Jiri Lastuvka, Bohemia). E, F.
Lentinus
badius
from Borneo (JS0094). G, H.
Lentinus badius
from Peninsular Malaysia (JSKT5858). I.
Lentinus sajor-caju
from mainland
Borneo (SNP24989). J.
Lentinus sajor-caju
from Gaya Island (JS0056). K.
Lentinus polychrous
from Borneo (JS0054). L.
Lentinus
squarrosulus
(BORHF0009).
SATHIYA SEELAN ET AL.: LENTINUS,POLYPORELLUS,NEOFAVOLUS 469
the sister group of the
Lentinus
/
Polyporellus
clade.
The most parsimonious reconstruction of character
states suggested that the plesiomorphic condition
for the
Lentinus/Polyporellus
clade is to have circular
pores (FIG.1,SUPPLEMENTARY FIG.7).Onsometree
topologies, a single transition from circular to
angular pores is reconstructed in the lineage leading
to
P. arcularius
and
P. brumalis
,butinothertrees
these two species do not form a monophyletic group
(FIG.1, SUPPLEMENTARY FIG. 7). In all trees the
species of
Lentinus
other than
L. tigrinus
form
a clade (albeit weakly supported), which implies
a single transition to wholly lamellate hymeno-
phores. The sub-poroid hymenophore of
Lentinus
tigrinus
is morphologically intermediate between
lamellae and angular pores, but the precursor to
the wholly lamellate condition in the
Lentinus/
Polyporellus
clade is uncertain.
Relationships of Lentinus sect. Dicholamellatae.—
Pegler (1983a) placed three species in section
Dicholamellatae: L. araucariae
,
L. brunneofloccosus
and
L. badius.
Manimohan et al. (2004) and
Njouonkou et al. (2013) described
L. cystidiatus
and
L. dicholamellatus
respectively as members of this
section. However morphological features reported for
L. cystidiatus
(cheilocystidia and absence of hyphal
pegs) suggest that it is probably a
Panus
and ITS
sequence data from a specimen identified as
L.
dicholamellatus
(TENN060790) suggest that it is in
fact
L. sajor-caju
, which will be discussed in a forth-
coming study focused on the
L. sajor-caju
complex
(Seelan et al. unpubl).
Morphological characters distinguishing sect.
Di-
cholamellatae
include a verrucose-squamose pileal
surface, wide-angled skeleto-ligative hyphae and
non-inflated generative hyphae, radiate hymeno-
phoral trama, abundant hyphal pegs, often dichoto-
mously furcate hymenophore, without true lamellu-
lae, ellipsoid to cylindrical spores and large basidia
(17–20 33.5–4.5 mm). Species of section
Dicholamel-
latae
exhibit metavelangiocarpic development, with
a universal veil eventually reduced to verrucose
squamules, which differs from the gymnocarpic
development represented in other
Lentinus
species
like
L. squarrosulus
,
L. sajor-caju
,
L. tigrinus
,
L.
striatulus
,
L. scleropus
,
L. bertieri
,
L. swartzii
and
L.
polychrous
.
Lentinus araucariae
is similar to
L. badius
,
except that it has subdistant and furcating lamellae.
Section
Dicholamellatae
is represented here by four
isolates that form a strongly supported group (FIG.1).
Isolate JS0094 from Borneo is macromorphologically
similar to
L. araucariae
as described in Pegler (1983a)
(FIG. 3E, F). The pilea surface was thin and with
subdistant, dichotomizing lamellae compared to
other
L. badius
collections. However microscopic
characters of this isolate conform to the description
of
L. badius
and were indistinguishable from those
of the isolate JSKT5858 from peninsular Malaysia
(FIG. 3G, H). All four specimens of section
Dichola-
mellatae
had abundant hyphal pegs. Pegler (1983a)
reported that
L. badius
contains more abundant
hyphal pegs than
L. araucariae
.
The isolates of Borneo and peninsular Malaysia
form a sister clade to the two isolates from Thailand.
Isolate PU00436 was identified as
L. araucariae
in the
herbarium collection, but it matches Pegler’s de-
scription of
L. badius
. We examined the holotype of
L. araucariae
(PC, New Caledonia); none of the
collections from this study match the holotype. Thus
at present we find evidence of at least two species that
conform morphologically to Pegler’s concept of
L.
badius,
one in Malaysia and another in Thailand.
Pegler placed into synonymy with
L. badius
the
names
Agaricus verrucarius
Berk. (West Bengal,
Darjeeling),
Lentinus inquinans
Berk. (Nepal),
Lenti-
nus brevipes
Cooke (Malay Peninsula, Perak),
Lentinus
fuscus
Lloyd (Singapore), and
Lentinus inverseconicus
(Vietnam), and he placed into synonymy with
L. araucariae
, the name
Panus verruciceps
Hongo
(from Papua New Guinea). Pegler reported that the
type locality of
L. badius
(synonym
Panus badius
)is
in the Philippines. Meanwhile the type locality for
L. araucariae
is recorded from New Caledonia and this
species is mainly restricted to Australasia and Sabah,
Malaysia. Resolving the species of section
Dicholamel-
latae
will require additional sampling from across
the geographic range of
L. badius
and
L. araucariae
,
including as many type specimens as possible.
Relationships of Lentinus sect. Rigidi.—
Section Ri-
gidi comprises
L. sajor-caju
,
L. squarrosulus
and
L.
polychrous
according to Pegler (1983a). Pegler
(1983a) placed
L. sajor-caju
as the type species for
section
Rigidi
. Section
Rigidi
is represented by four
isolates of each species.
Lentinus sajor-caju
is the only species reported in
the section that has an annulus and gills with
abundant hyphal pegs. Meanwhile
L. squarrosulus
,
which has a slender stipe, and
L. polychrous
, which has
a short, thick stipe, lack an annulus (Corner 1981,
Pegler 1983a).
Grand (2011) and Sotome et al. (2008) said that
this section was poorly resolved due to lack of
sampling. Our study suggests that the section
Rigidi
is monophyletic. The combined four-gene phylogeny
moderately supported the monophyly of
L. sajor-caju
,
L. squarrosulus
and
L. polychrous
as in Pegler’s
classification.
Lentinus sajor-caju
forms a clade that
470 MYCOLOGIA
is sister to a clade containing
L. squarrosulus
and
L.
polychrous
(FIG. 1).
Lentinus sajor-caju
revealed wide variation in
morphological features in collections from different
areas (Pegler 1983a). According to Pegler (1983a), 26
synonyms of
L. sajor-caju
have been reported in prior
classifications. Two isolates (SNP24989, JS0056) from
Borneo form a clade that is sister to a clade contain-
ing two isolates (FRI62056, TENN59793) from pen-
insular Malaysia and Thailand respectively. The
Bornean isolates, from the mainland (JS0056) and
Gaya Island (SNP24989), are morphologically differ-
ent based on the pileus shape. The pileus of isolate
JS0056 is larger (7–9 cm) diam compared to isolate
SNP24989, which is 3–5 cm diam. Both isolates had
abundant hyphal pegs and an annulus. Isolate
SNP24989 had a lobed margin (FIG. 3I), which is
not present in isolate JS0056 (FIG. 3J). It is not clear
whether these morphological variations correspond
to species limits.
Lentinus polychrous
resembles
L. badius
based on
their pilea surface (squamules and warts present as in
L. badius
) and forked hymenophore. However
Lentinus polychrous
has dimitic to trimitic hyphal
construction, rarely forking lamellae and a reduced
number of hyphal pegs, which is different from
L.
badius
, which has dimitic hyphal construction, strong-
ly forking lamellae and abundant hyphal pegs, and
explains why this species was not placed by Pegler
(1983a) in section
Dicholamellatae
.
Lentinus poly-
chrous
often has distinctive coloration (ochraceous
to brown) on the pileus surface with scattered
squamules as described in Pegler (1983a), which we
observed in some of the herbarium specimens or
fresh materials from Indonesia (AR618), Thailand
and Malaysia.
L. polychrous
has a pileus surface that resembles
chamois leather in both color and texture, which
contrasts with dark brown lamellae with a reddish or
purplish tint, according to Pegler (1983a). This
morphological character was observed in isolate
AH00024 from Thailand. Meanwhile isolate
KM141387 from Thailand did not have the reddish
lamellae but it had a greenish to dark brown pileus
and rusty brown lamellae. Isolate JS0054 from Borneo
had light to cream brown pileus with scattered
squamules as in
L. badius
(FIG. 3K).
Lentinus tigrinus
is reported frequently in tropical
regions especially in southeastern Asia (Sumaiyah et al.
2007, Dulay et al. 2012, Bolhassan et al. 2013). Pegler
(1983a) reported that
L. tigrinus
has essentially a north
temperate distribution. He also added that this species
often is confused with
L. squarrosulus
, which is mainly
found in paleotropical and Australasian regions. The
main cause of confusion between
L. tigrinus
(section
Tigrini
)and
L. squarrosulus
(section
Rigidi
)istheir
similar scabrous pilea surfaces (FIG.3L). Grand etal.
(2011) generated an ITS phylogeny and performed
mating studies of section
Lentinus
emphasizing
L.
tigrinus
, confirming that it has only a Eurasian and
north temperate distribution.
ACKNOWLEDGMENTS
The authors thank the Ministry of Higher Education of
Malaysia (MOHE) and Universiti Malaysia Sabah (UMS) for
providing KPT scholarship to JSS. Special thanks to Sabah
Parks (SP) and Sabah Biodiversity Council (SaBC) for
providing the permits for the collections in Kinabalu Park
area throughout 2010–2013. The authors also thank Dr Lee
Su See (Forest Research Institute of Malaysia), Atika
Retinowati (Herbarium Bogoriense, Indonesia), Professor
Yu Cheng Dai (Chinese Academy of Sciences, China) and
Professor Ronald H. Petersen and Professor Karen Hughes
(University of Tennessee, Knoxville) for providing loans of
specimens, Dr Maklarin Lakim (Sabah Parks), Dr Abdul
Hamid (ITBC, UMS) and Remi Ripin (SP) for arranging
the logistics during the field trip in Borneo. Many thanks to
Yabainus Juhailin, Siti Karim, Joumin Rangkasan and
Rollinus Paulous who assisted in field collections. Financial
support from NSF through the PolyPEET grant
(DEB0933081 to DSH) is gratefully acknowledged.
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