diversity

Article
Description of Lepiotaceous Fungal Species of the Genera
Chlorophyllum, Clarkeinda, Macrolepiota, Pseudolepiota, and
Xanthagaricus, from Laos and Thailand
Phongeun Sysouphanthong 1,2,3 , Naritsada Thongklang 1,2, *, Jian-Kui Liu 4 and Else C. Vellinga 5

1 Center of Excellence in Fungal Research, Mae Fah Luang University, Chiang Rai 57100, Thailand;
laofungi@[Link]
2 School of Science, Mae Fah Luang University, Chiang Rai 57100, Thailand
3 Ecology Division, Biotechnology and Ecology Institute, Ministry of Science and Technology,
Vientiane P.O. Box 2279, Laos
4 School of Life Science and Technology, Center for Informational Biology, University of Electronic Science and
Technology of China, Chengdu 611731, China; liujiankui@[Link]
5 UC Herbarium, UC Berkeley, Berkeley, CA 94720-2465, USA; ecvellinga@[Link]
* Correspondence: naritsada.t@[Link]; Tel.: +66-539-16996

Abstract: In our ongoing research on lepiotaceous taxa (Agaricaceae s.l.) in Laos and northern
Thailand, we focus here on Chlorophyllum, Clarkeinda, Macrolepiota, Pseudolepiota, and Xanthagaricus.
Collections were obtained from various habitats, including agricultural habitats, grasslands, and
rainforests. A total of 12 taxa were examined and investigated. Of these 12, two are new for science;





 viz. Xanthagaricus purpureosquamulosus with brownish-grey to violet-brown squamules on a pale-
violet to violet background; it shares the pileus color with X. caeruleus and X. ianthinus, but differs in
Citation: Sysouphanthong, P.;
other characters; and Macrolepiota excelsa, rather similar to M. procera but related to M. detersa. Two
Thongklang, N.; Liu, J.-K.; Vellinga,
E.C. Description of Lepiotaceous
species, Pseudolepiota zangmui and Xanthagaricus necopinatus are recorded for the first time in Thailand.
Fungal Species of the Genera Four species of Chlorophyllum and a total of four species of Macrolepiota were found, viz., C. demangei
Chlorophyllum, Clarkeinda, and C. hortense with white basidiospores, C. molybdites and C. globosum with green basidiospores, M.
Macrolepiota, Pseudolepiota, and detersa, M. dolichaula, the new M. excelsa, and M. velosa. Another rather common striking species is
Xanthagaricus, from Laos and Clarkeinda trachodes, with yellow-green basidiospores. Each species is described in detail, with color
Thailand. Diversity 2021, 13, 666. photographs and line drawings. Phylogenetic analyses based on internal transcribed spacer (nrITS)
[Link] region, the large subunit nuclear ribosomal (nrLSU) DNA and RNA polymerase II second largest
subunit (rpb2) genes provide evidence for the placement of the species covered.
Academic Editor: Michael Wink

Keywords: Agaricaceae; biodiversity; new taxa; new record; taxonomy; phylogeny; Southeast Asia
Received: 12 November 2021
Accepted: 7 December 2021
Published: 12 December 2021

1. Introduction
Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
As part of ongoing survey into the mycological diversity of Laos and northern Thai-
published maps and institutional affil- land [1], the focus of this article is on Macrolepiota Singer, and some genera in the ‘wider
iations. Agaricus clade’ (also considered to be Agaricaceae in the strict sense [2–4], in particular, the
genera Chlorophyllum Massee, Clarkeinda Kuntze, Pseudolepiota Z.W. Ge, and Xanthagaricus
(Heinem.) Little Flower, Hosag. and T.K. Abraham. Earlier, we described two new genera
in this clade, viz. Coniolepiota Vellinga and Eriocybe Vellinga [2] focused on the genus
Copyright: © 2021 by the authors.
Lepiota (Pers.) Gray [5–8] and documented the rarely recorded species Verrucospora vulgaris
Licensee MDPI, Basel, Switzerland.
Pegler [9].
This article is an open access article
Chlorophyllum Massee is widespread in tropical to temperate regions, often in anthro-
distributed under the terms and pogenic landscapes, with a total of 19 species [10–12]. The species are agaricoid or secotioid,
conditions of the Creative Commons the latter occurring in steppes, deserts, and other dry habitats. Until 2003 Chlorophyllum
Attribution (CC BY) license (https:// was restricted to the green-spored agaricoid species, whereas the white-spored agaricoid
[Link]/licenses/by/ taxa were included in Macrolepiota Singer, and the secotioid taxa were considered part of En-
4.0/). doptychum Czern. or Secotium Kunze. Molecular-phylogenetic research clearly showed that

Diversity 2021, 13, 666. [Link] [Link]

Diversity 2021, 13, 666 2 of 39

Chlorophyllum includes white-spored taxa with a smooth stipe and with or without a simple
germ pore, that Chlorophyllum and Macrolepiota s. str. are not closely related [2,13,14], and
that the secotioid taxa are relatively recent morphological adaptations (e.g., Vellinga et al.
2003 [12,13,15]). A proposal to conserve the name Chlorophyllum against Secotium was
published [16]. Only three Chlorophyllum species are listed in the checklist for Thailand [17].
Of these three, Chl. molybdites (G. Mey.) Massee seems to be common and widespread; Chl.
rhacodes (Vittad.) Vellinga was reported from the northeast, while Chl. hortense (Murrill)
Vellinga was only recorded from the southern part of Thailand [17–19]. However, these
reports are based on morphology alone, and in this group of similar-looking species, it is
better to have molecular and morphological evidence. The occurrence of Chl. rhacodes has
not yet been confirmed in Thailand, but in this article, we report Chl. globosum (Mossebo)
Vellinga with green basidiospores, and Chl. demangei (Pat.) Z.W. Ge and Zhu L. Yang with
white basidiospores as additions to the Thai mycoflora.
Clarkeinda trachodes (Berk.) Singer, originally described from Sri Lanka as Agaricus
trachodes Berk. [20], is commonly found in tropical regions of eastern and southern Asia;
there are reports from Bangladesh [21], India [22,23], Malaysia [24], Indonesia, and southern
China [25]. The species was also recorded from Thailand [2,17]. Here, we describe the Thai
collections in detail.
Macrolepiota Singer is also widespread, and members of this genus can be found
in various habitats, from grasslands to native forests. Till the early 2000s, the smooth-
stiped species, now accommodated in Chlorophyllum, were considered part of Macrolepiota.
However, Macrolepiota species differ from Chlorophyllum in the presence of a vestiture on the
stipe, resulting in a tiger pattern in the full-grown specimens, the trichodermal appearance
of the pileus covering, and the presence of a real germ pore in the spores [13,26,27]. The
two genera are not closely related, with Chlorophyllum in the Agaricus clade and Macrolepiota
close to Lepiota [13]. Similar to Chlorophyllum, Macrolepiota also harbors secotioid taxa [28].
Seven Macrolepiota taxa have been reported from Thailand, viz. M. africana (R. Heim)
Heinem, M. dolichaula (Berk. and Br.) Pegler and Rayner, M. excoriata (Schaeff.) Wasser, M.
gracilenta (Krombh.) Wasser, M. mastoidea (Fr.) Singer, M. procera var. procera and M. zeyheri
Heinem [17,18,29–31]. However, these records are based on morphology only; the presence
of some of these species in Thailand has to be confirmed by molecular means.
The monotypic genus Pseudolepiota Z.W. Ge was recently described from southern
China, based on Ps. zangmui Z.W. Ge [32]. This is one of the species superficially resembling
the European species Lepiota fuscovinacea F.H. Møller and J.E. Lange with a cutis-like pileus
covering and no clamp connections [2]. The species was found in two different locations in
northern Thailand, expanding the known distribution considerably.
Xanthagaricus (Heinem.) Little Flower, Hosag. and T.K. Abraham is thus far only
known from southern and eastern Asia and South Africa. It is a small genus characterized
by small basidiomata, a squamulose pileus, white to pale-yellow basidiospores, and the
absence of clamp connections. Heinemann and Little Flower [33] considered it a subgenus
of Hymenagaricus Heinem., but recent phylogenetic research showed that it did not form a
monophyletic group with Hymenagaricus species [34]. Two species were described from
Thailand, viz., X. thailandensis J. Kumla, N. Suwannarach and S. Lumyong [35] and X.
siamensis Yuan S. Liu and S. Lumyong [36], and here we describe a third species. We also
record X. necopinatus Iqbal Hosen, T.H. Li, and G.M. Gates, described from Bangladesh,
from Chiang Mai and Chiang Rai provinces in northern Thailand, expanding the known
distribution of that species.
In this study, we aim to give descriptions of new species and report new records and
extensive descriptions of some more common species for northern Thailand and Laos,
based on morphological studies and comparisons and molecular phylogenetic analyses.
Diversity 2021, 13, 666 3 of 39

2. Materials and Methods

2.1. Sample Collection and Morphological Study
The specimens were collected during the wet seasons of 2007–2019 in Laos and
Thailand. In the field, the forest type and habitat or substrate of mushrooms were recorded,
and the samples were photographed. Macro-morphology, examined on fresh samples,
included notes on shape, size, and color of the basidiomata, pileus, lamellae, stipe, annulus,
context, and spore print following the glossary of Vellinga and Noordeloos [26] and
Kornerup and Wanscher [37] for color annotations. Samples were dried on hot, dry air for
24 h and preserved in plastic bags with silica gel. Thai specimens were deposited in the
herbarium at Mae Fah Luang University (MFLU) with duplicates for the 2007 collections
made by Vellinga at UC; and Lao specimens were deposited in the National Herbarium of
Laos (HNL).
The micromorphology was illustrated from dried material: basidiospores, basidia, and
cheilocystidia from the lamellae and the structure of the pileus covering, using a drawing
tube attached to an Olympus CX-41 research compound microscope. The original color of
all characteristics was observed in water (H2 O) and 3–10% of potassium hydroxide (KOH),
and chemical reactions were observed using Melzer’s reagent, Cotton Blue, and Cresyl
Blue; line drawings were made of material mounted in Congo Red in ammonia. A total
of 25 basidiospores, 15 basidia, and cheilocystidia per collection were measured. Spores
were measured in side-view. The notation [50/2/2] indicates that the measurements were
made on 50 basidiospores in 2 samples in 2 collections; the size averages are given in the
description, while quotient (Q) of length and width average was calculated to indicate
the basidiospore shape. The technical terms used for the descriptions followed those of
Vellinga and Noordeloos [26].

2.2. Molecular Study

DNA was extracted from dried herbarium collections, according to the instructions of
the Biospin Fungus Genomic DNA Extraction Kit (Bioer Technology Co., Ltd., Hangzhou,
China). Primers ITS1 and ITS4 were used for the nrITS1, 5.8S, and nrITS2 regions; primers
LR0R and LR5 for large subunit region (LSU); primers fRPB2-6F and fRPB2-7R for poly-
merase II second largest subunit (rpb2) region; and PCR conditions followed common
protocols [38,39]. The PCR amplified products were cleaned and sequenced by Shanghai
Majorbio Bio-Pharm Technology Co., Ltd. (Shanghai, China). Sequences were edited, and
contigs were assembled using the SeqMan program (DNAStar, Madison, WI, USA), and all
new sequences were deposited in GenBank. The sequences were checked against existing
sequences at GenBank, and related sequences were obtained for each analysis.
Dataset 1, a combined dataset of nrITS-LSU genes (91 sequences), was composed
of 10 newly generated sequences (8 ITS, 2 LSU) and 81 sequences from GenBank (54 ITS,
27 LSU). Totally; 63 sequences in the Agaricus clade of the Agaricaceae were included,
representing Agaricus, Clarkeinda, Coniolepiota, Eriocybe, Heinemannomyces, Hymenagaricus,
Pseudolepiota, Micropsalliota, and Xanthagaricus; and Chlorophyllum rhacodes (Vittad.) Vel-
linga is set as outgroup (Table 1). Dataset 2, Chlorophyllum species, 125 sequences of nrITS
sequences, was composed of 10 new sequences and 115 sequences from GenBank repre-
senting all known species in the genus; Clarkeinda trachodes (Berk.) Singer and Agaricus
campestris L. are outgroups, and GenBank accession number and country of origin are
indicated after the species name in the tree (Figure 2). Dataset 3, a combined dataset of
nrITS-LSU-rpb2 genes (139 sequences) of Chlorophyllum species, was composed of 19 new
sequences (10 nrITS, 8 LSU, 1 rpb2) and 120 sequences from GenBank (48 nrITS, 40 LSU,
32 rpb2), with Clarkeinda trachodes and Agaricus campestris as outgroup (Table 2); all known
Chlorophyllum taxa are represented in this dataset as well. Dataset 4, a dataset of nrITS
gene regions (96 sequences) of Macrolepiota species, was composed of 17 new sequences
and 80 sequences from GenBank, with Leucoagaricus meleagris as the outgroup, and Gen-
Bank accession number and country of origin are indicated after the species name in the
tree (Figure 4).
Diversity 2021, 13, 666 4 of 39

Table 1. List of collections of taxa in the Agaricaceae and their GenBank accession numbers used in the molecular analyses
of dataset 1.

Country/Collection GenBAnk Accession Number

Taxon Voucher Number Number ITS LSU
Agaricus aff. campestris Murphy 6242 USA HM488744 -
Agaricus bisporatus Contu1 - AF432882 -
Agaricus bohusii LAPAG562 - KM657928 KR006613
Agaricus deserticola M. Smith USA HM488747 -
Agaricus diminutivus Vellinga 2360 USA AF482831 AF482877
Agaricus megacystidiatus MFLU 12-0137 Thailand NR_119953 -
Agaricus sp. NTS113 Thailand JF514531 -
Agaricus sp. C3182 Togo KJ540956 -
Agaricus sp. BAB5059 India KR155104 -
Chlorophyllum rhacodes Vellinga 2106 Netherlands AF482849 -
Clarkeinda trachodes Iqbal 806 Bangladesh - MG462712
Clarkeinda trachodes ecv3838 Thailand HM488750 HM488771
Clarkeinda trachodes ecv3550 Thailand HM488751 KY418837
Clarkeinda trachodes MFLUCC:100139 Sri Lanka KU845621 -
Clarkeinda trachodes MFLU 19-2351 Thailand MN099351 -
Coniolepiota spongodes ecv3898 Thailand HM488755 -
Coniolepiota spongodes HKAS 77574 Bangladesh KC625531 KC625530
Eriocybe chionea ecv3560 Thailand HM488752 HM488773
Heinemannomyces splendidissimus ecv3586 Thailand HM488760 HM488769
Heinemannomyces splendidissimus zrl3043 Thailand JF691559 -
Heinemannomyces splendidissimus GDGM 46633 China MF621038 MF621039
Hymenagaricus ardosiicolor LAPAF9 Togo JF727840 -
Hymenagaricus ardosiicolor isolateZ4 Tanzania KM360160 -
Hymenagaricus cf. kivuensis BR6089 Burundi KM982454 -
Hymenagaricus sp. CA833 Thailand JF727858 -
Hymenagaricus sp. CA801 Thailand JF727859
Hymenagaricus sp. zrl3103 Thailand KM982450 KM982452
Hymenagaricus sp. LD2012186 Thailand KM982451 KM982453
Micropsalliota arginophaea SFSU zrl 2089 China HM436616 HM436578
Micropsalliota pusillissima SFSU zrl 3047 China HM436645 HM436594
Pseudolepiota zangmui Z. W. Ge 3537 China KY768925 -
Pseudolepiota zangmui Z. W. Ge 2106 China KY768927 -
Pseudolepiota zangmui Z. W. Ge 2107 China KY768926 -
Pseudolepiota zangmui Z. W. Ge 2175 China KY768928 MG742049
Pseudolepiota zangmui MFLU 10-0515 Thailand KX904355 -
Pseudolepiota zangmui MFLU 10-0518 Thailand KX904356 -
Pseudolepiota zangmui MFLU 19-2355 Thailand MN099352 -
Xanthagaricus caeruleus GDGM 50651 China MF039088 MF039086
Xanthagaricus caeruleus GDGM 50794 China MF039089 MF039087
Xanthagaricus epipastus zrl 3045 Thailand HM436649 HM436609
Xanthagaricus flavosquamosus GDGM 50913 China MF351627 -
Xanthagaricus flavosquamosus GDGM 50924 China MF351628 -
Xanthagaricus flavosquamosus GDGM 50918 China MF351629 MF351631
Xanthagaricus ianthinus HMJAU45193 China MH166760 -
Xanthagaricus ianthinus HMJAU45192 China MH166761 -
Xanthagaricus ianthinus HMJAU45191 China MH166762 -
Xanthagaricus ianthinus HMJAU45194 China MH166764 -
Xanthagaricus necopinatus Iqbal821 Bangladesh MF351626 MF351630
Xanthagaricus necopinatus MFLU 19-2353 Thailand MN480544 -
Xanthagaricus necopinatus MFLU 19-2358 Thailand MN480545 -
Xanthagaricus pakistanicus LAH SH 207 Pakistan KY621555 KY621554
Xanthagaricus pakistanicus HUP SH 315 Pakistan KY621556 -
Xanthagaricus pakistanicus SWAT SH 389 Pakistan KY621557 -
Xanthagaricus purpureosquamulosus MFLU 19-2354 Thailand MN099353 MN097917
Xanthagaricus purpureosquamulosus MFLU 19-2356 Thailand MN099354 MN097918
Xanthagaricus siamensis MFLU 19-0575 Thailand MN176991 MN176981
Xanthagaricus siamensis MFLU 19-0574 Thailand MN176992 MN176982
Xanthagaricus siamensis MFLU 19-0576 Thailand MN176993 MN176983
Xanthagaricus sp. ecv3807 Thailand HM488761 HM488770
Xanthagaricus taiwanensis C.M. Chen 3636 Taiwan DQ006271 DQ006270
Xanthagaricus taiwanensis HKAS 42545 Taiwan DQ490633 -
Xanthagaricus thailandensis CMUJK010 Thailand MG256663 MG256665
Xanthagaricus thailandensis CMUJK0115 Thailand MG256664 MG256666
Note: New specimens and sequences are in bold.
Diversity 2021, 13, 666 5 of 39

Table 2. List of fungal taxa of second dataset of Chlorophyllum species and their GenBank accession numbers used in
molecular analysis of datasets 2.

Voucher/Collection GenBank Accession Number

Taxon Country
Number ITS LSU rpb2
Agaricus campestris LAPAG370 China KM657927 KR006607 KT951556
Chlorophyllum africanum PREM 62140 South Africa MG741961 MG742041 MG742070
Chlorophyllum africanum PREM 62141 South Africa MG741963 MG742042 MG742071
Chlorophyllum agaricoides HKAS 101312 Russia MG742003 MG742020 MG742050
Chlorophyllum agaricoides HMAS 71678 China MG742004 MG742021 MG742051
Chlorophyllum arizonicum AH31724 Mexico KR233490 KR233499 -
Chlorophyllum arizonicum Trappe 11481 USA HQ020416 HQ020419 -
Chlorophyllum brunneum HKAS 101315 Italy MG742013 MG742022 MG742052
Chlorophyllum brunneum ecv2361 USA AY083206 AF482886 HM488804
Chlorophyllum demangei Z. W. Ge 3112 China MG741965 MG742027 MG742056
Chlorophyllum demangei Z. W. Ge 3574 China MG741964 MG742025 MG742055
Chlorophyllum demangei ecv3622 Thailand HM488765 - -
Chlorophyllum demangei MFLU 12-1769 Thailand KJ524556 MN097907 -
Chlorophyllum demangei MFLU 09-0005 Thailand KJ524555 MN097908 -
Chlorophyllum demangei ecv3557 Thailand MN582745 - -
Chlorophyllum globosum Z. W. Ge 2006-1 China MG741995 MG742023 -
Chlorophyllum globosum PREM 62147 South Africa MG742002 MG742024 MG742053
Chlorophyllum globosum MFLU 12-1815 Thailand KJ524553 MN097909 -
Chlorophyllum hortense HKAS 101317 China MG741967 MG742026 MG742054
Chlorophyllum hortense Z. W. Ge 3115 China MG741968 MG742028 MG742057
Chlorophyllum hortense HKAS 90470 China MG741971 MG742029 MG742058
Chlorophyllum hortense MFLU 12-1783 Thailand KJ524554 MN097910 -
Chlorophyllum hortense MFLU 19-2352 Thailand MN099355 MN097916 MN816433
Chlorophyllum hortense PC17 Philippines MN099356 MN097915 -
Chlorophyllum lusitanicum AH45540 Spain KR233482 KR233491 -
Chlorophyllum lusitanicum AH43927 Spain KR233483 KR233492 -
Chlorophyllum molybdites HKAS 45051 China MG741985 MG742030 MG742059
Chlorophyllum molybdites Z. W. Ge 3381 USA MG741993 MG742034 MG742063
Chlorophyllum molybdites Z. W. Ge 3146 China MG741987 MG742031 MG742060
Chlorophyllum molybdites HKAS 101322 Italy MG741988 MG742032 MG742061
Chlorophyllum molybdites Z. W. Ge 3377 USA MG741992 MG742033 MG742062
Chlorophyllum molybdites MFLU 12-1772 Thailand KJ524557 MN097911 -
Chlorophyllum molybdites MFLU 12-1765 Thailand KJ524559 MN097912 -
Chlorophyllum molybdites MFLU 12-1775 Thailand KJ524558 MN097913 -
Chlorophyllum molybdites S57 Sudan MK541941 - -
Chlorophyllum molybdites J. States AEF1097 USA AF482836 - -
Chlorophyllum molybdites DMSC09538 Thailand KP229775 - -
Chlorophyllum molybdites AM150 (CUH) India KM190077 - -
Chlorophyllum molybdites HKAS 101319 Thailand MG741994 - -
Chlorophyllum molybdites FS10 India MK855510 - -
Chlorophyllum neomastoideum HKAS 83208 China MG741976 MG742035 MG742064
Chlorophyllum nothorachodes H. Lepp 1142 Australia AF482855 - -
Chlorophyllum olivieri HKAS 31587 Germany MG742016 MG742036 MG742065
Chlorophyllum olivieri HKAS 53466 Germany MG742017 MG742037 MG742066
Chlorophyllum palaeotropicum PREM 62142 South Africa MG741978 MG742038 MG742067
Chlorophyllum palaeotropicum PREM 62145 South Africa MG741982 MG742039 MG742068
Chlorophyllum palaeotropicum HKAS 93747 Benin MG741983 MG742040 MG742069
Chlorophyllum palaeotropicum pc21 Philippines MN099357 MN097914 -
Chlorophyllum pseudoglobosum AM155 India KP642506 KR080484 -
Chlorophyllum rhacodes ecv2106 Netherlands AF482849 AY176345 -
Chlorophyllum rhacodes OKM19588 USA U85312 U85277 HM488803
Chlorophyllum sphaerosporum HMAS 66153 China MG742011 MG742043 MG742072
Chlorophyllum sphaerosporum HMAS 71683 China MG742012 MG742044 MG742073
Chlorophyllum subrhacodes Z. W. Ge 3411 USA MG741975 MG742045 MG742074
Chlorophyllum subrhacodes Z. W. Ge 3232 USA MG741973 MG742046 MG742075
Chlorophyllum subrhacodes Z. W. Ge 3385 USA MG741972 MG742048 MG742077
Chlorophyllum subrhacodes Z. W. Ge 3242 USA MG741974 MG742047 MG742076
Clarkeinda trachodes ecv3550 Thailand HM488751 KY418837 HM488802
Note: New specimens and sequences are in bold.

The datasets were aligned using MAFFT version 7.130-win32 ([Link]

[Link]/[Link]/MAFFT_7.130, accessed on 10 December 2021) [40,41]. The final align-
ments were submitted to the TreeBase website ([Link]
[Link], TreeBase No. 24630, accessed on 10 December 2021).
Diversity 2021, 13, 666 6 of 39

A Maximum likelihood (ML) analysis was performed in RAxML 7.2.6 [42] for the
individual dataset with GTRGAMMA + I as the model of evolution, and branch support
was estimated over 1000 bootstrap partitions (BP) with the rapid bootstrap option. For
Bayesian inferences (BI) analysis, the best substitution model of the individual dataset
was defined by using MrModelTest v.2.3 [43]. For dataset (1), the best-selected model
was GTR + I + G for both ITS and LSU. For dataset (2), the ITS dataset, the best-selected
model was HKY + I + G. For dataset (3), the best-selected model for ITS was HKY + I + G,
and GTR + I + G was the best for LSU and rpb2. For dataset (4), the best-selected model
was again GTR + I + G. A Bayesian inference (BI) analysis was performed with mrbayes
3.2.7a [44] at the CIPRES webserver (available at [Link] (accessed on
10 December 2021) with the suitable setting of 4 individual runs. The Bayesian infer-
ence analysis; each one beginning from random trees with 4 simultaneous independent
chains, each of 4 Markov chain Monte Carlo (MCMC) chains for 2 independent runs,
2,000,000 generations for datasets 1, 2, and 3, and 1,000,000 generations for dataset (4),
sampled every 1000 generations, the first 25% (0.25) of the sampled trees were discarded as
burn-in, and all sampled after the average standard deviation of split frequencies lowered
than 0.01 were used to reconstruct a 50% majority-rule consensus tree and to calculate the
Bayesian posterior probabilities (BPP) of the clades. The phylogenetic tree results of all
analyses were exported and edited in TreeView [Link]. [45]. The phylogenetic trees were
edited in the software of Adobe Illustrator CS3.

3. Results
3.1. Result of the Phylogenetic Analyses
Figure 1 shows the maximum likelihood phylogenetic tree of the extended Agaricus
clade based on a combined ITS and LSU data set. A total of 63 taxa were analyzed (Table 1).
The alignment dataset comprised 1560 characters (including the gaps). Seven different
major clades can be recognized in the phylogenetic tree (Figure 1). The Xanthagaricus clade
received high bootstrap support and was completely comprised of Xanthagaricus species;
the two Thai specimens of X. purpureosquamulosus had identical sequences and are basal to
a clade comprised of X. epipastus, X. necopinatus, and X. pakistanicus; the sequences of the
Thai specimens of Ps. Zangmui were identical with those from China. Clarkeinda also forms
a monophyletic clade, composed of a Thai specimen from this study, two Thai specimens
from Vellinga et al. [2], a specimen from Bangladesh Hosen et al. [34], and a specimen from
the type location in Sri Lanka [46].
Figure 2 indicates the maximum likelihood phylogenetic tree of Chlorophyllum species
based on nrITS sequences. Data from 126 collections were included, and the alignment
dataset comprised 809 characters (including the gaps). Six clades were distinguished, all
receiving high bootstrap support. Thai taxa clustered into two sections as in an analysis of
combined sequences data (Figure 3).
Figure 3 shows the maximum likelihood phylogenetic tree of Chlorophyllum species
based on multi-gene DNA data set of ITS, LSU, and rpb2 gene regions. The alignment
comprised 58 collections (Table 2), and this dataset comprised 2322 characters (including
the gaps). The six clades or sections in Chlorophyllum, (Chlorophyllum, Ellipsoidospororum,
Endoptychorum, Rhacodium, Parvispororum, and Sphaerospororum) received high bootstrap
support, and these results are consistent with those found by Ge et al. [10]. The Thai
specimens were distributed in two sections: C. demangei and C. hortense with white basid-
iospores without a germ pore in section Ellipsoidospororum, and the species with greenish
basidiospores and a germ pore, C. globosum and C. molybdites, in section Chlorophyllum.
Diversity 2021, 13, 666 7 of 39
Diversity 2021, 13, 666 8 of 43

Figure 1. Maximum
Figure 1. Maximum likelihood
likelihoodphylogenetic
phylogenetictree treeofof the
the Agaricus
Agaricus clade
clade in the
in the Agaricaceae
Agaricaceae basedbased on nrITS-LSU
on nrITS-LSU sequence
sequence data.
data. New sequences generated from Thailand are in bold. Bootstrap values (ML/B) are given above the branches. Boot-
New sequences generated from Thailand are in bold. Bootstrap values (ML/B) are given above the branches. Bootstrap
strap values for maximum likelihood equal to or greater than ≥60 and Bayesian posterior probabilities ≥ 0.95 are placed
values for maximum likelihood equal to or greater than ≥60 and Bayesian posterior probabilities ≥0.95 are placed above the
above the branches. Genus abbreviations are as follows: A = Agaricus, C = Chlorophyllum, Cl = Clarkeinda, Co = Coniolepiota,
branches.
E Genus
= Eriocybe, abbreviations are asHafollows:
H = Heinemannomyces, A = Agaricus,
= Hymenagaricus, Ps C = Chlorophyllum,
= Pseudolepiota, M =ClMicropsalliota,
= Clarkeinda, Co = Coniolepiota,
and E = Eriocybe,
X = Xanthagaricus. Chlo-
H = Heinemannomyces, Ha = Hymenagaricus,
rophyllum rachodes Vellinga is an outgroup. Ps = Pseudolepiota, M = Micropsalliota, and X = Xanthagaricus. Chlorophyllum
rachodes Vellinga is an outgroup.
Diversity 2021, 13, 666 8 of 39
Diversity 2021, 13, 666 9 of 43

[Link]
Figure Maximumlikelihood
likelihoodphylogenetic
phylogenetic tree
tree of of Chlorophyllum
Chlorophyllum based
based on on nrITS
nrITS sequences.
sequences. New New sequences
sequences generated
generated from
from Thailand are in bold. Bootstrap values for maximum likelihood equal to or greater than ≥ 60 and Bayesian posterior
Thailand are in bold. Bootstrap values for maximum likelihood equal to or greater than ≥60 and Bayesian posterior probabilities
probabilities ≥ 0.95 are placed above the branches. Genus abbreviations are as follows: A = Agaricus, C = Chlorophyllum, Cl
≥0.95 are placed above the branches. Genus abbreviations are as follows: A = Agaricus, C = Chlorophyllum, Cl = Clarkeinda, Thai
= Clarkeinda, Thai specimens from this study are in bold types. Clarkeinda trachodes (Berk.) Singer and Agaricus campestris
specimens from thisGenBank
L. are outgroup. study areaccession
in bold types.
numberClarkeinda trachodes
and country are(Berk.) Singer
indicated and
after Agaricus
species campestris L. are outgroup. GenBank
name.
accession number and country are indicated after species name.
Diversity 2021, 13, 666 9 of 39
Diversity 2021, 13, 666 10 of 43

Figure 3. Maximum Likelihood tree of Chlorophyllum based on analysis of nrITS-LSU-rpb2 sequence data. Bootstrap values
Figure 3. Maximum Likelihood tree of Chlorophyllum based on analysis of nrITS-LSU-rpb2 sequence data. Bootstrap values
(ML/B) are given above the branches. Bootstrap values for maximum likelihood equal to or greater than ≥60 and Bayesian
(ML/B) are given
posterior above the≥ branches.
probabilities Bootstrap
0.95 are placed values
above the for maximum
branches. likelihood
Abbreviations equal to
are as follows: [Link] greater than
= Agaricus, ≥60 and Bayesian
C = Chlorophyllum,
posterior
andprobabilities ≥Clarkeinda
Cl = Clarkeinda. 0.95 are placed above
trachodes the branches.
and Agaricus Abbreviations
campestris are as follows:
are used as outgroup. A. = Agaricus,
New sequences C = Chlorophyllum,
are in bold type.
and Cl = Clarkeinda. Clarkeinda trachodes and Agaricus campestris are used as outgroup. New sequences are in bold type.

Figure 4 shows the maximum likelihood phylogenetic tree of Macrolepiota species

based on nrITS sequences. A total of 97 sequences was analyzed; the aligned dataset
comprised 777 characters (including the gaps). Three major clades or sections (sect. Macrole-
piota, sect. Macrosporae, sect. Volvata) were recognized with high bootstrap support in the
phylogenetic tree.
Diversity 2021, 13, 666 10 of 39
Diversity 2021, 13, 666 11 of 43

[Link]
Figure Maximumparsimony
parsimonytree ofMacrolepiota
treeof Macrolepiota based
based on
on analysis of nrITS sequences.
sequences. Bootstrap
Bootstrapvalues
valuesfor
formaximum
maximum
likelihoodequal
likelihood equaltotoororgreater
greaterthan
than≥≥6060and
andBayesian
Bayesianposterior
posteriorprobabilities
probabilities≥≥0.95
0.95are
areplaced
placedabove
abovethe
thebranches.
[Link]-
Abbre-
viations
tions are
are as as follows:
follows: La. =La. = Leucoagaricus,
Leucoagaricus, L = Lepiota,
L = Lepiota, andand
M =MMacrolepiota.
= Macrolepiota.
All All taxon
taxon names
names in bold
in bold areare
newnew sequences
sequences in
in this
this study. Leucoagaricus meleagris (Gray) Singer and Lepiota cristata (Bolton) P. Kumm. are used as outgroup.
study. Leucoagaricus meleagris (Gray) Singer and Lepiota cristata (Bolton) P. Kumm. are used as outgroup. Country, type, and Country,
type, and GenBank accession number are indicated after each species name.
GenBank accession number are indicated after each species name.
 spherical with umbo, applanate to plano-concave, with straight to the deflexed margin;
surface smooth when young, brownish-grey (5D2), later breaking up into scales and leav-
ing light-brown to brown (5D4, 6D7–8) glabrous calotte at the center, with concolorous
with tufted patches around umbo and irregular patches scattered toward the margin, on
Diversity 2021, 13, 666 white to pale-yellow (5A3) background, turning orange-white (5A2) with time; margin 11 of 39
with strongly sulcate marginal zone, striate, up to 15 mm wide, with exceeding lamellae
when mature. Lamellae free, slightly or distinctly ventricose, up to 5 mm wide, white to
yellowish-white
3.2. Taxonomy (4A2), crowded, lamella-edge eroded. Stipe 10–90 × 6–7 mm, cylindrical,
or slightly wider at base; surface covered with white fibrils, turning orange-white (5A2)
3.2.1. Chlorophyllum Massee
when touched. Annulus superonate, attached at the upper side of the stipe, moveable
Chlorophyllum demangei (Pat.) Z.W. Ge & Zhu L. Yang, MycoKeys 33: 80 (2018)
when mature, with surface concolorous to patches on the pileus. The context in pileus
≡ Lepiota
white, in stipedemangei
white andPat., Bull. turning
hollow, trimest. orange-white
Soc. mycol. Fr. 23 (2):
(5A2) 78 (1907).
in both stipe and pileus con-
[Link]
TasteFungorum number:
peanut-like. IF 823863;
Smell mild. Facesoffungi
Spore number: FoF 07067; Figures 5 and 6.
print white.

[Link]
Figure Freshbasidiomata
basidiomataof of Chlorophyllum
Chlorophyllum demangei
demangei in situ.
in situ. (a,b)(a,b)
MFLU MFLU 09-0005.
09-0005. (c) MFLU
(c) MFLU 12-
12-1769.
1769.
Pileus 50–90 mm, subglobose when young, expanding to parabolic, convex-hemispherical
with umbo, applanate to plano-concave, with straight to the deflexed margin; surface
smooth when young, brownish-grey (5D2), later breaking up into scales and leaving light-
brown to brown (5D4, 6D7–8) glabrous calotte at the center, with concolorous with tufted
patches around umbo and irregular patches scattered toward the margin, on white to pale-
yellow (5A3) background, turning orange-white (5A2) with time; margin with strongly
sulcate marginal zone, striate, up to 15 mm wide, with exceeding lamellae when mature.
Lamellae free, slightly or distinctly ventricose, up to 5 mm wide, white to yellowish-white
(4A2), crowded, lamella-edge eroded. Stipe 10–90 × 6–7 mm, cylindrical, or slightly wider
at base; surface covered with white fibrils, turning orange-white (5A2) when touched.
Annulus superonate, attached at the upper side of the stipe, moveable when mature, with
surface concolorous to patches on the pileus. The context in pileus white, in stipe white
and hollow, turning orange-white (5A2) in both stipe and pileus context. Taste peanut-like.
Smell mild. Spore print white.
Basidiospores [100,4,4] 6.5–8.5 × 4.2–6.5 µm, avl × avw = 7.5 × 5.5 µm, Q = 1.3–1.67,
Qav = 1.4–1.55, in side-view ellipsoid to oblong-amygdaliform, in frontal view ellip-
soid, oblong, without germ pore, slightly thick-walled, hyaline, dextrinoid, congophilous,
cyanophilous, metachromatic in Cresyl Blue. Basidia 18–28 × 7.0–11.5 µm, clavate, hyaline
and thin-walled, 2-spored and 4-spored, occasionally 1-spored. Lamella edge sterile, with
abundant cheilocystidia. Cheilocystidia 24–55 × 6–9.5 µm, narrowly clavate, slightly nar-
rowly lageniform, cylindrical, colorless, and slightly thick-walled. Pleurocystidia absent.
Diversity 2021, 13, 666 12 of 39

Pileus covering a trichoderm made up of cylindrical, narrowly clavate elements with a long
stalk, 40–145 × 6–14 µm, colorless or with pale-brown parietal pigment, with the encrusted
Diversity 2021, 13, 666 wall in some elements and lower hyphae. Stipe covering a cutis made up of cylindrical
13 of 43
hyphae and elements, colorless, up to 12 µm wide. Clamp connections not found.

Figure [Link]
Figure Chlorophyllum demangei
demangei(ecv3622).
(ecv3622). (a)
(a)Basidiomata.
Basidiomata. (b)
(b) Basidiospores.
Basidiospores. (c)
(c) Basidia.
Basidia. (d)
(d) Cheilocystidia.
Cheilocystidia. (e)
(e) Pileus
Pileus
covering elements.
covering elements.

Basidiospores
Habitat [100,4,4] 6.5–8.5
and distribution: Solitary × 4.2–6.5 µm,groups,
or in large avl × avw = 7.5 × 5.5on
saprotrophic, µm, Q = 1.3–1.67,
decaying wood
Qavleaves
and = 1.4–1.55, in humus
on rich side-view ellipsoid
soil; to oblong-amygdaliform,
in deciduous rainforest, gardens, and in frontal view ellipsoid,
grasslands.
oblong, without
Material germ pore,
examined: slightlyChiang
Thailand, thick-walled, hyaline,Mae
Mai Province, dextrinoid, congophilous,
Taeng District, Pha Deng cy-
Village: 19◦ 07metachromatic
anophilous, 0 13.700 N; 98◦ 43in0 52.9 00 E, alt.,
Cresyl Blue.905Basidia 18–28
m, 6 July × 7.0–11.5
2007, µm, clavate, PNG05
P. Sysouphanthong, hyaline
and thin-walled,
(MFLU 2-spored
09-0005); ibidem, 12 and
July 4-spored, occasionally 1-spored.
2007, P. Sysouphanthong, PNG050 Lamella
(MFLUedge sterile,
09-0051); with
ibidem,
abundant cheilocystidia. Cheilocystidia 24–55 × 6–9.5 µm, narrowly
25 July 2007, E. C. Vellinga, ecv3557; ibidem, 15 July 2007, E. C. Vellinga, ecv3622; Chiangclavate, slightly nar-
rowly
Rai, lageniform,
Muang cylindrical,
District, Pong Phra colorless, and slightly
Bath Village, 21 Julythick-walled. Pleurocystidia absent.
2012, P. Sysouphanthong, 2012-6
Pileus covering
(MFLU 12-1769).a trichoderm made up of cylindrical, narrowly clavate elements with a
long Notes:
stalk, 40–145
the Thai× 6–14 µm, colorless
collections or with pale-brown
of Chlorophyllum demangei can parietal pigment,by
be recognized with the en-
brownish-
crusted
grey wall in some elements
to yellowish-brown and patches
irregular lower hyphae.
on white Stipe covering
pileus a cutis made
with sulcate up of
or striate cylin-
margin
in most
drical collections,
hyphae white and
and elements, free lamellae
colorless, up to 12with
µma wide.
whiteClamp
spore print, a simple
connections notannulus,
found.
oblong-ovoid
Habitat and to oblong amygdaliform,
distribution: Solitary orand hyaline
in large basidiospores
groups, saprotrophic, without a germ wood
on decaying pore,
clavate,
and leaves 4-spored
on richbasidia,
humus soil; narrowly clavate rainforest,
in deciduous to cylindrical cheilocystidia,
gardens, and [Link]
pileusMaterial
coveringexamined:
made up of cylindrical
Thailand, to narrowly
Chiang clavate elements,
Mai Province, Mae Taeng andDistrict,
absencePha of clamp
Deng
connections. It closely
Village: 19°07′13.7″ 98°43′52.9″C.E,hortense,
N;resembles alt., 905microscopically
m, 6 July 2007,because of the basidiospores
P. Sysouphanthong, PNG05
without
(MFLU a09-0005);
germ pore and the
ibidem, 12cylindrical
July 2007,cheilocystidia, but it differs
P. Sysouphanthong, in the(MFLU
PNG050 4-spored09-0051);
basidia.
Thus far, it seems to be much more restricted in its distribution than C.
ibidem, 25 July 2007, E. C. Vellinga, ecv3557; ibidem, 15 July 2007, E. C. Vellinga, ecv3622;hortense, as it is only
known from China, northern Thailand, and Vietnam [10,47]. This
Chiang Rai, Muang District, Pong Phra Bath Village, 21 July 2012, P. Sysouphanthong, is the first record of the
species from Thailand.
2012-6 (MFLU 12-1769).
Notes: the Thai collections of Chlorophyllum demangei can be recognized by brownish-
grey to yellowish-brown irregular patches on white pileus with sulcate or striate margin
in most collections, white and free lamellae with a white spore print, a simple annulus,
oblong-ovoid to oblong amygdaliform, and hyaline basidiospores without a germ pore,
clavate, 4-spored basidia, narrowly clavate to cylindrical cheilocystidia, trichodermal pi-
Diversity 2021, 13, 666 13 of 39

Chlorophyllum globosum (Mossebo) Vellinga, Mycotaxon 83: 416 (2002)

Diversity 2021, 13, 666 ≡ Macrolepiota globosa Mossebo, Mycotaxon 76: 268 (2000). 15 of 43
Index Fungorum number: IF 374394; Facesoffungi number: FoF 07069; Figures 7 and 8.

[Link]
Figure Freshbasidiomata
basidiomataof
ofChlorophyllum
Chlorophyllumglobosum
globosumininsitu.
situ.(a)
(a)MFLU
MFLU12-1815.
12-1815.(b,c)
(b,c)MFLU
MFLU192357.
192357.
(d) MFLU 10-0555.
(d) MFLU 10-0555.
Pileus 50–200 mm, subglobose to globose when young, expanding to parabolic to con-
vex, plano-concave when fully mature, with straight or slightly inflexed margin; glabrous
at the center, pale-orange to brownish-orange (5A2–3, 6C7–8), with concolorous patches
of irregular shapes around the center and scatted towards the margin and fragile when
mature, on white to yellowish-white (2A2, 3A2, 4A2) felted or fibrillose background and
turning pastel red to red (9A4–6) when touched; margin white, sulcate, but non-striate,
not fringed, exceeding lamellae when mature. Lamellae free, slightly remote from the
stipe, white to orange-white (5A1–2) when young, turning pastel red to red (9A4–6) when
touched, pastel green to grayish-green (29A4, 29B4) when fully mature, crowded, ven-
tricose, and narrowly to pileal margin, up to 20 mm wide, with the eroded edge. Stipe
85–210 × 12–31 mm, tapering to apex, with bulb-like, 30–34 mm wide base; white to
brownish-orange (6C3–6) background, sometimes with some white fibrils at apical zone,
turning pastel red to red (9A4–6) when touched, with white rhizomorphs connected to
the substrate. Annulus under-developed, cuff-like, moveable when mature, white and
fibrillose in the upper part, at underside with squamules similar to the patches on the
pileus. Context thick and white in pileus, white in the stipe, brownish-orange (6C3–6) at
apical zone, paler in the middle zone, and white toward the base, turning pastel red to red
(9A4–6) in both pileus and stipe context. Smell and taste not observed. Spore print dull
green to grayish-green (29D3–5, 29D5–6).
Diversity 2021, 13, 666 14 of 39
Diversity 2021, 13, 666 16 of

Figure 8. Chlorophyllum globosum (MFLU 12-1815). (a) Basidiomata. (b) Basidiospores. (c) Pileus covering elements.
Figure 8. Chlorophyllum globosum (MFLU 12-1815). (a) Basidiomata. (b) Basidiospores. (c) Pileus
covering elements.
Material examined: THAILAND, Chiang Mai Province, Mae Taeng District, Pon
duad Village:16°06′16.1″
Basidiospores [75,3,2] 9.0–12.0 × N; 99°43′07.9″
7.0–9.0 µm, avl E, ×alt.,
avw 780–805 m, 16 June
= 10.3–11.3 2010, µm,
× 7.4–8.3 P. Sysoupha
thong, P37 (MFLU 10-0555); Chiang Rai Province, Muang District, Ratjabhat Universi
Q = 1.3–1.57, avQ = 1.4, in side-view broadly ellipsoid to ellipsoid-amygdaliform, in frontal
Campus, 30 August 2012, P. Sysouphanthong, 2012-21 (MFLU 12-1815); Chiang Rai Pro
view broadly ellipsoid or amygdaliform, with truncate apex and germ pore, thick-walled,
ince, Muang District, Forest of Mae Fah Luang University Campus, 10 July 2019, P. Syso
hyaline to hyaline green, dextrinoid, congophilous, cyanophilous, metachromatic. Basidia
phanthong, 2019-24 (MFLU 19-2357). LAOS, Vientiane Capital, Xaythany, Houay Yan
22.0–34.0 × 9.0–14.0 µm, clavate, hyaline, thin-walled, 4-spored, often 2-spored, rarely
Preserve Forest, 16 June 2017, P. Sysouphanthong, PS2017–7 (HNL503445).
1-spored. Lamella edge sterile. Cheilocystidia 30.0–58.0 × 10.0–29.0 µm, essentially clavate,
Notes: Chlorophyllum globosum is recognized by medium to large-sized, white basi
occasionally with a slightly long stalk, rarely utriform and spheropedunculate, thin-walled
iomata, subglobose to globose pileus, which expands to convex when mature, pale-brow
and colorless. Pleurocystidia absent. Pileus covering a hymenoderm made up of layers of
to orange-brown patches on white pileus, a white stipe, a cuff-like annulus, and a redde
oblong to cylindrical, narrowly
ing reaction clavate
in most partselements, terminalwhen
of the basidioma elements 40.0–100
touched × 5.0–11.0 µm,
or damaged.
with rounded or attenuate apex, hyaline and thin-walled, sometimes
Chlorophyllum globosum was described from Cameroon as Macrolepiota with pale-yellowglobosa [4
intracellular pigment in upper elements. Clamp connections not found.
and the Thai material fits the description very well, except for the spore color, which w
Habitat and distribution:
originally given asgrowing
white. Thein small
species groups, saprotrophic,
was transferred on nutrient-rich
to Chlorophyllum by Vellinga [49
soil, in grassland in shade of Samanea saman
based on phylogenetic analyses of ITS and LSU. C. globosum isdeciduous
and on soil rich in humus in very close rain
to C. molybdit
forest during June to August inand
in morphology, thethey
wet are
season of taxa
sister northern
in theThailand and also
phylogenetic trees,known from
but they are definite
Vientiane Capital, [Link] [10,27].
different
Material examined: THAILAND,
The noticeable Chiangbetween
differences Mai Province, Mae Taengmolybdites
the Chlorophyllum District, Pongduad
and C. globosum sam
Village:16◦ 060 16.1 00 N; 99◦ 430 07.900 E, alt., 780–805 m, 16 June 2010, P. Sysouphanthong,
ples from Thailand and Laos are the whiter basidiomata, longer elements of the pile
P37 (MFLU 10-0555);
covering,Chiang Rai Province,
and longer Muangwith
cheilocystidia District, Ratjabhat
a distinct University
stalk in C. globosumCampus,
(Figures 7 and 8
30 August 2012, P. Sysouphanthong,
while cheilocystidia are2012-21 (MFLU
subglobose, 12-1815);
broadly Chiang
clavate Rai Province,
to clavate withoutMuang
a long stalk in
District, Forestmolybdites
of Mae Fah Luang11University
(Figures Campus,
and 12). However, 10 are
they Julyidentical
2019, [Link],
most other characteristics.
2019-24 (MFLUwe 19-2357).
had notLAOS,
obtainedVientiane
sequenceCapital, Xaythany,
data from Houay this
our collections, Yang Preserve
species Forest,
would most probab
16 June 2017, [Link] have been recognized,
Sysouphanthong, PS2017–7and(HNL503445).
the different morphology of the cheilocystidia tabled as
intraspecific globosum
Notes: Chlorophyllum [Link] recognized by medium to large-sized, white basid-
iomata, subglobose to globose pileus, which expands to convex when mature, pale-brown
to orange-brown patches on white pileus, a white stipe, a cuff-like annulus, and a reddening
reaction in most parts of the basidioma when touched or damaged.
Diversity 2021, 13, 666 15 of 39

Chlorophyllum globosum was described from Cameroon as Macrolepiota globosa [48],

and the Thai material fits the description very well, except for the spore color, which was
originally given as white. The species was transferred to Chlorophyllum by Vellinga [49],
based on phylogenetic analyses of ITS and LSU. C. globosum is very close to C. molybdites
in morphology, and they are sister taxa in the phylogenetic trees, but they are definitely
different species [10,27].
The noticeable differences between the Chlorophyllum molybdites and C. globosum
samples from Thailand and Laos are the whiter basidiomata, longer elements of the pileus
covering, and longer cheilocystidia with a distinct stalk in C. globosum (Figures 7 and 8),
while cheilocystidia are subglobose, broadly clavate to clavate without a long stalk in C.
molybdites (Figures 11 and 12). However, they are identical in most other characteristics. If
we had not obtained sequence data from our collections, this species would most probably
not have been recognized, and the different morphology of the cheilocystidia tabled as an
intraspecific variation.
Chlorophyllum globosum is now known from various countries in Africa (Benin, Cameroon,
and South Africa) and Asia (India, Laos, Thailand, and China). The occurrence in some
countries is based on ITS sequences in GenBank that are identical to those from Thailand
(Figure 2).

Chlorophyllum hortense (Murrill) Vellinga, Mycotaxon 83: 416 (2002)

≡ Lepiota hortensis Murrill, N. Amer. Fl. 10 (1): 59 (1914)
≡ Leucoagaricus hortensis (Murrill) Pegler, Kew Bull. add. Ser. 9: 414 (1983)
≡ Lepiota humei Murrill, Lloydia 6: 220 (1943)
= Chlorophyllum humei (Murrill) Vellinga, Mycotaxon 83: 416 (2002)
= Lepiota mammillata Murrill, Lloydia 6: 220 (1943)
= Chlorophyllum mammillatum (Murrill) Vellinga, Mycotaxon 83: 416 (2002)
= Lepiota subfulvidisca Murrill, Lloydia 6: 221 (1943)
= Chlorophyllum subfulvidiscum (Murrill) Vellinga, Mycotaxon 83: 416 (2002)
= Lepiota alborubescens Hongo, Mem. Fac. lib. Arts Educ. Shiga Univ., nat. Sci. 12: 40
= Macrolepiota alborubescens (Hongo) Hongo, Trans. Soc. mycol. Japan 27: 107 (1986)
= Chlorophyllum alborubescens (Hongo) Vellinga, Mycotaxon 83: 416 (2002)
= Leucoagaricus bisporus Heinem., Bull. Jard. bot. nat. Belg. 43: 8 (1973).
Index Fungorum number: IF 374396; Facesoffungi number: FoF 03444; Figures 9 and 10.
Pileus 72–98 mm, convex, expanding to umbonate with distinctly umbo, with straight
margin; surface covered with light-brown to yellow-brown (5D4–5) glabrous calotte at
center, with brownish-yellow (5C7–8) irregular patches or squamules toward the margin,
on orang white to light-orange (5A2–4) background; margin sulcate or slightly striate, white.
Lamellae free, narrowly fusiform, up to 7 mm wide, white, crowded, lamella-edge eroded.
Stipe 60–110 × 6–7 mm, cylindrical, slightly wider to base; surface smooth, white. Annulus
superonate, moveable, white, with brownish-yellow (5C7–8) on the upper part. Context
white in pileus, white in stipe and hollow. All parts of basidiomata turn orange-white (5A2)
when touched. Taste peanut-like. Smell mild. Spore print white.
Basidiospores [50,2,1] 6.0–8.5 × 5.3–6.5 µm, avl × avw = 7.5 × 5.8 µm, Q = 1.2–1.54,
Qav = 1.3, in side-view broadly ellipsoid to ellipsoid amygdaliform, in frontal view el-
lipsoid, oblong, without germ pore, hyaline, dextrinoid, congophilous, cyanophilous,
metachromatic. Basidia 25–30 × 8–11.5 µm, clavate, hyaline, 2-spored, occasionally 1-
spored. Lamella edge sterile, with abundant cheilocystidia. Cheilocystidia 35–50 × 7–9 µm,
narrowly clavate, cylindrical, sometimes with short apical excrescence or appendage, color-
less. Pleurocystidia absent. Pileus covering of scales a trichoderm made up of cylindrical,
narrowly clavate elements with long stalk, 30–155 × 6.3–16 µm, colorless or with pale-
brown parietal pigment, with an encrusted wall in some elements and lower hyphae. Stipe
covering a cutis made up of cylindrical hyphae and elements, colorless, 10 µm wide. Clamp
connections not observed.
 Diversity 2021, 13, 666 17 of 43
Diversity 2021, 13, 666 16 of 39

Chlorophyllum globosum is now known from various countries in Africa (Benin, Cam-
eroon, and South
Habitat Africa) and Asia
and distribution: (India,
growing in Laos, Thailand,
large groups, inand China). The
grasslands, occurrence
saprotrophic onin
some countries
nutrient-rich [Link] based on ITS sequences in GenBank that are identical to those from
Thailand (Figure
Material 2).
examined: Thailand, Chiang Rai Province, Muang District, Mae Korn District,
22 July 2010, P. Sysouphanthong, BJP54 (MFLU 12-1783); Muang District, Pongphabath
Chlorophyllum
Village, hortense
1 July 2018, (Murrill) Vellinga,
P. Sysouphanthong, Mycotaxon
PS2018-31 (MFLU83: 416 (2002)
19-2352).
≡ Lepiota
Notes: Chlorophyllum hortense
hortensis Murrill, can [Link]
N. Amer. by light-brown to yellow-brown
10 (1): 59 (1914)
≡ Leucoagaricus
irregular patches on white to light-orange
hortensis pileus
(Murrill) Pegler, andBull.
Kew sulcate
[Link].
striate margin,
9: 414 (1983)white and
free lamellae
≡ Lepiotawith
humeia white spore
Murrill, print,6:
Lloydia an220
annulus,
(1943) oblong ovoid to oblong amygdaliform,
and hyaline basidiospores
= Chlorophyllum humeiwithout
(Murrill)aVellinga,
germ pore, 2-spored83:
Mycotaxon basidia, narrowly clavate to
416 (2002)
cylindrical cheilocystidia, trichodermal pileus
= Lepiota mammillata Murrill, Lloydia 6: 220 (1943)covering made up of cylindrical to narrowly
clavate= Chlorophyllum mammillatum (Murrill) Vellinga, Mycotaxon 83: 416 (2002) species
elements, and absence of clamp connections. It should be noted that most
of lepiotaceous fungi have 4-spored
= Lepiota subfulvidisca basidia;6:only
Murrill, Lloydia a handful of species in various genera
221 (1943)
are characterized by 2-spored
= Chlorophyllum basidia.
subfulvidiscum (Murrill) Vellinga, Mycotaxon 83: 416 (2002)
This species has been described
= Lepiota alborubescens Hongo, Mem. by several
Fac. [Link]
Arts Educ. [27,50], and because
Shiga Univ., nat. Sci. of
12:the
40
basidiospores without a germ pore, it was accommodated in Leucoagaricus.
= Macrolepiota alborubescens (Hongo) Hongo, Trans. Soc. mycol. Japan 27: 107 (1986) It is one of
the most widespread species in Chlorophyllum. A record from
= Chlorophyllum alborubescens (Hongo) Vellinga, Mycotaxon 83: 416 (2002)India [51] might refer to C.
demangei, because of the 4-spored basidia.
= Leucoagaricus bisporus Heinem., Bull. Jard. bot. nat. Belg. 43: 8 (1973).
The species is found throughout the tropics in diverse, mostly man-influenced habitats,
Index Fungorum number: IF 374396; Facesoffungi number: FoF 03444; Figures 9 and
on dung, compost, and soil [14,27].
10.

Figure 9. Fresh basidiomata of Chlorophyllum hortense. (a) MFLU 12-1783. (b,c) MFLU 19-2352.
Figure 9. Fresh basidiomata of Chlorophyllum hortense. (a) MFLU 12-1783. (b,c) MFLU 19-2352.
Diversity 2021, 13, 666 17 of 39
Diversity 2021, 13, 666 18 of 43

Figure 10. Microcharacters of Chlorophyllum hortense (MFLU 12-1783). (a) basidiospores. (b) basidia. (c) cheilocystidia. (d)
Figure 10. Microcharacters of Chlorophyllum hortense (MFLU 12-1783). (a) basidiospores. (b) basidia.
pileus covering.
(c) cheilocystidia. (d) pileus covering.
Pileus 72–98 mm, convex, expanding to umbonate with distinctly umbo, with straight
margin; surface
Chlorophyllum molybdites (G. covered with light-brown
Mey.) Massee, to yellow-brown
Kew Bull. (5D4–5) glabrous calotte at
1898: 136 (1898)
center, with brownish-yellow (5C7–8) irregular patches or squamules toward the margin,
≡ Agaricusonmolybdites
orang [Link]., Pr. Fl.(5A2–4)
light-orange essequ.: 300 (1818)
background; margin sulcate or slightly striate,
≡ Lepiota molybdites
white. Lamellae([Link],
Mey.) Sacc.,fusiform,
narrowly Syll. Fung.
up to 5: 30 wide,
7 mm (1887)white, crowded, lamella-edge
eroded. molybdites
= Mastocephalus Stipe 60–110 (G.
× 6–7Mey.)
mm, cylindrical,
Kuntze, Rev. slightly wider
Gen. [Link] 2:
base;
860surface
(1891)smooth, white.
Annulus superonate, moveable, white, with brownish-yellow
= Leucocoprinus molybdites (G. Mey.) Pat., Bull. Soc. mycol. Fr. 29: 215 (1913) (5C7–8) on the upper part.
Context white in pileus, white in stipe and hollow. All parts of basidiomata turn orange-
= Macrolepiota molybdites (G. Mey.) Moreno, Bañares and Heykoop, Mycotaxon 55: 467 (1995)
white (5A2) when touched. Taste peanut-like. Smell mild. Spore print white.
= Agaricus morganii Peck, [50,2,1]
Basidiospores Bot. Gaz. 4: ×137
6.0–8.5 (1879)
5.3–6.5 µm, avl × avw = 7.5 × 5.8 µm, Q = 1.2–1.54, Qav
= Lepiota morganii (Peck) Sacc.,
= 1.3, in side-view broadlySyll. Fung.
ellipsoid 5: 30 (1887)
to ellipsoid amygdaliform, in frontal view ellipsoid,
oblong, morganii
= Mastocephalus without germ pore,Kuntze,
(Peck) hyaline, dextrinoid,
Rev. [Link], cyanophilous, metachro-
Pl. 2: 860 (1891)
matic. Basidia 25–30 × 8–11.5 µm, clavate, hyaline, 2-spored, occasionally 1-spored. La-
= Chlorophyllum morganii (Peck) Massee, Kew Bull. 1898: 136 (1898)
mella edge sterile, with abundant cheilocystidia. Cheilocystidia 35–50 × 7–9 µm, narrowly
= Agaricusclavate,
glaziovii Berk., Vidensk.,
cylindrical, sometimes Meddel.
with short1879-1880: 32 (1880)
apical excrescence or appendage, colorless.
= Pholiota Pleurocystidia
glaziovii (Berk.) Sacc.,
absent. Syll.
Pileus Fung.
covering 5: 751a trichoderm
of scales (1887) made up of cylindrical, nar-
rowly clavate
= Lepiota ochrospora elements
Cooke andwith long stalk,
Massee, 30–155 × 21:
Grevillea 6.3–16
73µm, colorless or with pale-brown
(1893)
parietal esculentum
= Chlorophyllum pigment, withMassee,
an encrusted
Kew wall in some
Bull. elements
1898: and lower hyphae. Stipe cov-
136 (1898)
ering a cutis made up of cylindrical hyphae and elements, colorless, 10 µm wide. Clamp
= Lepiota esculenta (Massee) Sacc. and P. Syd., Syll. Fung. 16: 2 (1902)
connections not observed.
= Agaricus guadelupensis Pat., Bull. Soc. mycol. Fr. 15: 197 (1899)
= Annularia camporum Speg. in An. Mus. nac. Buenos Aires, Ser. II, 6: 117 (1899) [1898]
= Lepiota camporum (Speg.) Speg.,Bol. Acad. nac. Cienc. Cordoba 29: 114 (1926)
= Agaricus congolensis Beeli in Bull. Soc. roy. bot. Belg. 61: 92 (1928)
= Chlorophyllum molybdites var. congolense (Beeli) Heinem., Fl. Icon. Champ. Congo 16:
323 (1967).
Index Fungorum number: IF 604726; Facesoffungi number: FoF 03445; Figures 11 and 12.
Diversity 2021, 13, 666 18 of 39
Diversity 2021, 13, 666 20 of 43
Diversity 2021, 13, 666 20 of 43

Figure 11. Fresh basidiomata of Chlorophyllum molybdites

molybdites in situ.
situ. (a) MFLU
MFLU 12-1775. (b) MFLU 12-1819.
12-1819.
Figure 11. Fresh
Figure11. Freshbasidiomata ofChlorophyllum
basidiomataof Chlorophyllum molybditesinin situ.(a)
(a) MFLU12-1775.
12-1775.(b)
(b)MFLU
MFLU 12-1819.

Figure 12. Microcharacters of Chlorophyllum molybdites (MFLU 12-1783). (a) basidiospores. (b) cheil-
Figure 12.
12. Microcharacters of Chlorophyllum molybdites (MFLU 12-1783). (a) basidiospores. (b) cheil-
Figure
ocystidia. pileus [Link] Chlorophyllum molybdites (MFLU 12-1783). (a) basidiospores. (b)
(c)Microcharacters
ocystidia. (c) pileus covering.
cheilocystidia. (c) pileus covering.
Pileus 70–110 mm, globose to subglobose when young, expanding to convex, plano–
Pileus 70–110
Pileus 70–110 mm,
mm, globose
globose toto subglobose
subglobose when
when young,
young, expanding
expanding to to convex,
convex, plano–
plano–
convex, with low umbo, with straight margin, with glabrous calotte at center, light-brown
convex,
convex, with low umbo, with straight margin, with glabrous calotte at center, light-brown
to brownwith low umbo,
(7D5–6), with straightpatches
with concolorous margin,and
with glabrous scattered
squamules calotte at center,
towardlight-brown
margin, on
to brown
to brown (7D5–6),
(7D5–6), with
with concolorous
concolorous patches
patches and
and squamules
squamules scattered
scattered toward
towardmargin,
margin,onon
concentrically fibrillose background, orange-white (6A2) around umbo, and white around
concentricallyfibrillose
concentrically fibrillosebackground,
background,orange-white
orange-white (6A2)around
aroundumbo,
umbo,and andwhite
whitearound
around
marginal zone; margin fibrillose, fringed, exceeding(6A2)
lamellae when mature. Lamellae free,
marginal zone;
marginal zone; margin
margin fibrillose,
fibrillose, fringed,
fringed, exceeding
exceeding lamellae
lamellae when
when mature.
mature. Lamellae
Lamellae free,
free,
broad, crowded, white when young, becoming dull green (27D3) when mature, with an
broad, crowded,
broad, crowded, white
white when
when young,
young, becoming
becoming dull
dull green
green (27D3)
(27D3) when
when mature,
mature, with
with anan
eroded edge. Stipe 60–85 × 10–12 mm, widening downward base, 18–23 mm at the base,
eroded edge. Stipe 60–85 × 10–12 mm, widening downward base, 18–23 mm at the base,
Diversity 2021, 13, 666 19 of 39

eroded edge. Stipe 60–85 × 10–12 mm, widening downward base, 18–23 mm at the base,
white to yellowish-white (4A2), smooth, hollow. Annulus free, descending, with a cuff
around stipe, movable, white; underside concolorous with umbo. The context in pileus and
stipe white, turning brownish-orange (7C5). Smell carrot-like. Taste not observed. Spore
print grayish-green (27C3–4).
Basidiospores [100,5,5] 8.5–13 × 6–7.5 µm, avl × avw = 9.6 × 7.0 µm, Q = 1.3–1.5,
avQ = 1.4, ellipsoid-amygdaliform, with truncate apex and germ pore, hyaline or pale-
green, thick-walled, dextrinoid, congophilous, cyanophilous, and metachromatic in Cresyl
blue. Basidia 29–36 × 10–12 µm, 4-spored, some 1-spored or 2-spored, hyaline, slightly
thick-walled. Lamella edge sterile. Cheilocystidia 14–35 × 10–20 µm, clavate, broadly
clavate, hyaline, slightly thick-walled. Pileus covering a hymeniderm made up of tightly
packed clavate, narrowly clavate, oblong, cylindrical, elements; terminal elements 25–
62.5 × 5.0–12 µm, with hyaline to pale-brown parietal pigment. Stipe covering a cutis made
up of cylindrical to narrowly cylindrical elements, 4.5–7.5 µm wide, thin-walled, hyaline.
Clamp connections not found.
Habitat and distribution: growing in small groups to large groups or fairy rings
in grasslands and under shade trees along with grasslands, saprotrophic and terrestrial,
throughout northern Thailand.
Material examined: Thailand, Chiang Rai Province, Muang District, Mae Fah Luang
University Campus, 7 May 2009, P. Sysouphanthong and J-K. Liu, BJP0041; ibidem, 22 July
2012, P. Sysouphanthong, 2012-9 (MFLU 12-1772); ibidem, 05 August 2012, P. Sysouphan-
thong, 2012-12 (MFLU 12-1775); Chiang Rai Province, Muang District, Pongphrabath
Village, 17 July 2012, P. Sysouphanthong, 2012-2 (MFLU 12-1765); Pha Yao Province, Muang
District, Kuawn Pra Yuan Area, 12 June 2011, P. Callac and S.C. Karunarathna, 2011-11
(MFLU 12-1819).
Notes: Chlorophyllum molybdites are easy to recognize by the free and dull green
lamellae when fully mature, the grayish-green spore print, and the large basidiomata with
a light-pileus with brown squamules at the center. All parts of the basidioma turn orange
to red when damaged. It is often found in lawns and grasslands. The species might be
confused with Macrolepiota species when samples are collected in a young stage. It could
also be confused with Clarkeinda trachodes (Berk.) Sing, which has a yellow to olive-brown
spore print, a much flimsier annulus, and white velar remnants over the pileus scales. Cl.
trachodes grows in northern Thailand in the same area and habitat as C. molybdites, but
has not been found outside Asia yet. For differences with C. globosum, see notes under
that species.
Chlorophyllum molybdites is widespread, especially in subtropical and tropical re-
gions [10,14,27,49,52]. In northern-temperate regions in Europe, it was found only in-
doors [53], Watling [54]. It was first recorded in Thailand by Høiland and Schumacher
(1982), and since then, there have been several reports of the species throughout Thai-
land [17,18]. However, those studies were only based on their macro-morphological
characters. This is the first study on C. molybdites in Thailand, providing a full description
and illustrations. Læssøe et al. [55] recorded this species from Xiangkhouang Province of
northern Laos.
In the nrITS sequence analysis (Figure 2) and in the combined nrITS-rnLSU-rpb2
analysis (Figure 3), three sequences of Thai specimens from this study are clustered with
other specimens from other countries worldwide.
Chlorophyllum molybdites cause gastrointestinal distress, but the severity of the reactions
to this species differs among people and whether the mushrooms have been cooked or not.
The list of synonyms as given above is based on morphology; it is possible that some
of the names, such as Agaricus congolensis Beeli, in fact, refer to C. globosum, which is
morphologically very similar to C. molybdites [56,57].
 Diversity 2021, 13, 666 22 of 43
Diversity 2021, 13, 666 20 of 39
Diversity 2021, 13, 666 22 of 43

3.2.2. Clarkeinda Kuntze

3.2.2. Clarkeinda
3.2.2. Clarkeinda Kuntze
Kuntze
Clarkeinda trachodes (Berk.) Singer, Lilloa 22: 413 (1951) [1949]
Clarkeinda trachodes (Berk.) Singer, Lilloa 22: 413 (1951) [1949]
≡ Agaricus
Clarkeinda trachodes
trachodes (Berk.)Berk., London
Singer, Lilloa J.
22:Bot.
4136:(1951)
487 bis (1847)
[1949]
≡
≡ Agaricus trachodes
Chitoniella trachodes Berk., London
(Berk.) Petch, J. Bot.R.6:bot.
Ann. 487Gdns
bis (1847)
Peradeniya 4 (6): 396 (1909)
≡ Agaricus
≡ trachodes
Chitoniella Berk.,
trachodes London
(Berk.) J. Bot.
Petch, 6: 487
Ann. bis (1847)
R. bot. Gdns Peradeniya 4 (6): 396 (1909)
≡ Fungus trachodes (Berk.) Kuntze, Revis. gen. pl. (Leipzig)
≡ Chitoniella trachodes (Berk.) Petch, Ann. R. bot. Gdns Peradeniya 3 (3): 4480
(6):(1898).
396 (1909)
≡ Fungus
Index trachodes
Fungorum (Berk.) Kuntze, Revis. gen. pl. (Leipzig) 3 (3): 480 (1898). 13 and
≡ Fungus trachodesnumber: IF 294937;
(Berk.) Kuntze, Facesoffungi
Revis. number:
gen. pl. (Leipzig) FoF480
3 (3): 07066; Figures
(1898).
14. Index Fungorum number: IF 294937; Facesoffungi number: FoF 07066; Figures 13 and 14.
Index Fungorum number: IF 294937; Facesoffungi number: FoF 07066; Figures 13 and
14.

Figure13.
13. Fresh basidiomata
basidiomata of
of Clarkeinda
Clarkeinda trachodes
trachodesinin situ. (a,b) MFLU 19-2351.
Figure 13. Fresh
Figure Fresh basidiomata of Clarkeinda trachodes insitu.
situ. (a,b)
(a,b) MFLU
MFLU 19-2351.
19-2351.

Figure 14.
Figure
Figure 14. Microcharacters
Microcharactersof
Microcharacters Clarkeinda
ofofClarkeindatrachodes
Clarkeindatrachodes(MFLU
(MFLU
trachodes 192-351).
192-351).
(MFLU (a)(a)
192-351). basidiospores. (b)(b)
basidiospores. basidia.
(a) basidiospores. (c) (c)
basidia.
(b) basidia.
cheilocystidia. (d)
cheilocystidia. (d) pileus
pileus covering.
covering.
(c) cheilocystidia. (d) pileus covering.

Pileus 50–220 mm, first hemispherical to campanulate, expanding to convex or um-

bonate, applanate to plano-concave, with straight or uplifted margin, with a large irregular
patch at the center covering up to 1/3 of pileus, smooth or glabrous, light-brown (6D5–8)
Diversity 2021, 13, 666 21 of 39

to brown (6E7–8), with yellowish-white or pale-yellow (4A3–4) to light-brown (6D5–8)

squamules around patch toward the margin, on yellowish-white or pale-yellow (4A3–4)
fibrillose background, with white velar remnants on top the central patch; margin split,
with concolorous squamules same as on rest of pileus. Lamellae L = around 180, l = 1–3,
7–12 mm wide, free and remote (2.5 mm) from the stipe, white when young, becoming dull
yellow (3B2) to greyish-yellow (4B2) and finally greyish-yellow (3D4) to olive (3C4), with a
concolorous or white eroded edge. Stipe 80–150 × 10–28 mm (apex) × 15–28 mm (base),
wider at bulbous up to 25 mm wide base, tapering to apex, with white to yellowish-white or
pale-yellow (4A3–4) surface, with concolorous squamules same as those on pileus, turning
reddish-brown (8E5–7) when touched, hollow. Annulus up to 6 cm wide, attached at the
upper part of the stipe, membranous, with eroded margin, fragile and ragged with age;
upper side smooth or fibrillose, white to pale-yellow (4A3–4); underside covered with
concolorous squamules as on pileus or stipe. Volva present, white. Context white and
up to 8 mm thick in pileus with turning orange or reddish-brown (8E5–7), white and
hollow in stipe with turning reddish-brown (8E5–7). Odor strong, a mixture of artificial
fruit flavor and rubber. Taste not tried. Spore print greyish-yellow (3D4) to olive (3C4) or
olivaceous yellow.
Basidiospores [25,1,1] 5.0–6.0 × 3.5–4.0 µm, avl × avw = 5.36 × 3.74 µm, Q = 1.32–1.57,
avQ = 1.43, ellipsoid ovoid in side-view, broadly to ellipsoid ovoid in frontal view, with trun-
cate apex and germ pore, hyaline, thick-walled. Basidia 19–28 × 5–8 µm, clavate, hyaline,
thin-walled, 4-spored. Pleurocystidia absent. Cheilocystidia abundant, 20–50 × 12–35 µm,
clavate to broadly clavate, often subglobose to sphaeropedunculate with a short or long
stalk, hyaline, slightly thick-walled. Pileus covering in squamules an epithelium composed
of 4–5 layers of clavate or short clavate elements, often oblong to fusiform, rarely subclavate
in the upper layer, 30–50 × 15–25 µm; subglobose, oblong to clavate in the lower layer,
18–45 × 15–25 µm; elements, with pale-brown to brown partial and intracellular pigment.
Stipe covering in squamules same as those on pileus, stipe surface not observed. Clamp
connections absent.
Habitat and distribution: growing solitary or in small groups, saprotrophic, on
nutrient-rich soil, in the deciduous rain forest of northern Thailand.
Material examined: Thailand, Chiang Mai Province, Mae Taeng District, Pha Deng
Village, alt. 900–950 m, 15 August 2011, P. Sysouphanthong, PS2011-12 (MFLU 19-2351);
ibidem, 22 August 2007, P. Sysouphanthong (MFLU 09-0059); ibidem, 25 June 2007, E.C.
Vellinga 3550 (UC); ibidem, 4 July 2008, E.C. Vellinga 3838 (MFLU 08-1227). Laos, Oudomxay
Province, Xay District, Houay Houm Village, 16 August 2014, PS2014-752 (HNL502423).
Notes: Clarkeinda trachodes is commonly found in tropical regions of eastern and
southern Asia. It has been recorded in Thailand before [2,17,55] and Laos [55]. This study
provides a full description, line drawing, and color photos taken in the field. A new nrITS
sequence data analysis showed that our new collection clustered together with specimens
collected from Thailand [2], Bangladesh [21], and the type location in Sri [46] (Figure 1).

3.2.3. Macrolepiota Singer

Macrolepiota detersa Z. W. Ge, Zhu. L. Yang and Vellinga, Fungal Diversity 45: 83 (2010)
Index Fungorum number: IF 518349; Facesoffungi number: FoF 09698; Figures 15 and 16.
Diversity 2021, 13, 666 22 of 39
Diversity 2021, 13, 666 24 of 43

Diversity 2021, 13, 666 25 of 43

Figure 15. Fresh basidiomata of Macrolepiota detersa in situ. (a,b) MFLU 12-1784. (c–e) MFLU 12-
Figure 15. Fresh basidiomata of Macrolepiota detersa in situ. (a,b) MFLU 12-1784. (c–e) MFLU 12-1772.
1772.

Figure 16. Microcharacters of Macrolepiota detersa (MFLU 192-351). (a) basidiospores. (b) cheilocystidia. (c) pileus cover-
ing.
Figure 16. Microcharacters of Macrolepiota detersa (MFLU 192-351). (a) basidiospores. (b) cheilocys-
tidia. (c) pileus covering.
Pileus 60–160 mm, when young globose to subglobose, expanding to convex, umbo-
nate, broadly campanulate, plane-umbonate, with straight or decurrent margin; when
young glabrous, brownish-orange to light-brown or dark red-brown (7C5–6, 8–9 F7–8),
surface-breaking around the center, with concolorous squamules or patches around gla-
brous calotte at the center toward the margin, crowded, with distantly fragile patches to-
ward margin), with concolorous brownish-orange (7C5–6) fibrillose squamules or
squamules between patches towards the margin, on white to yellowish-white (4A2) mat-
Diversity 2021, 13, 666 23 of 39

Pileus 60–160 mm, when young globose to subglobose, expanding to convex, umbon-
ate, broadly campanulate, plane-umbonate, with straight or decurrent margin; when young
glabrous, brownish-orange to light-brown or dark red-brown (7C5–6, 8–9 F7–8), surface-
breaking around the center, with concolorous squamules or patches around glabrous calotte
at the center toward the margin, crowded, with distantly fragile patches toward margin),
with concolorous brownish-orange (7C5–6) fibrillose squamules or squamules between
patches towards the margin, on white to yellowish-white (4A2) matted-fibrillose to the
felted background; margin splitted, covered with light-grey (6C3) to grey orange (6B3)
granular-floccose Universal Veil elements. Lamellae free, slightly remote from the stipe,
white, sometimes with reddish spots, ventricose to broad, 8–16 mm wide, close to crowded,
with the eroded edge. Stipe 80–400 × 12–18 mm, cylindrical with slight wider downward
base, with bulb at base, 20–30 mm; dull, dry, scurfy to squamulose overall, minute squa-
mules at apex zone to middle and squamules downward base, concolorous with those
squamules on pileus, on brown-grey (6C3) background. Annulus superior, membranous,
thin, descending, double, persistent but easily removable, pendulous, 30–40 mm long, with
concolorous squamules underside of internal cuff-liked. Context white and thick in pileus;
stipe white and hollow, fibrous, turning pinkish-grey when cut, not changing color in bulb.
Taste not observed. Smell strong, pleasant. Spore print white.
Basidiospores [50,2,2] 14–16.2 × 9.5–11 µm, avl × avw = 15.2 × 10.1 µm, Q = 1.45–1.62,
avQ = 1.52, ellipsoid amygdaliform, ellipsoid to oblong in frontal view, with apical germ
pore and hyaline cap, hyaline and thick-walled, dextrinoid, congophilous, cyanophilous,
metachromatic in Cresyl Blue. Basidia 38–41 × 12–14 µm, clavate, slightly thick-walled,
hyaline, 4-spored. Lamella edge sterile. Cheilocystidia 20–25 × 7–14 µm, short clavate to
clavate, irregular clavate, rarely utriform, sometime branched, hyaline, thin-walled. Pleuro-
cystidia absent. Pileus covering a hymenoderm made up of layers of oblong to cylindrical
elements, branched, terminal cylindrical elements cylindrical or fusiform, oblong, with
rounded or attenuate to apex, 10–80 × 7–16 µm, slightly thick-walled, with hyaline to
pale-brown parietal pigment. Stipe covering not observed. Clamp-connections not found.
Habitat and distribution: solitary, saprotrophic, and terrestrial, growing on the ground
beside a grassland or yard, and ground with decomposing deciduous leaves from the
mixed rain forest, dominant species being Castanopsis spp. in northern Thailand and in
northern Laos. This is proposed as a new record for tropical Southeast Asia.
Material examined: Thailand, Chiang Rai, Muang District, Pong Phra Bath Village,
8 June 2011, P. Sysouphanthong, 2011-10 (MFLU 12-1772); Chiang Rai, Mae Fah Luang
District, Mae Sa Long Nok Village, 9 July 2012, P. Sysouphanthong, 20112-19 (MFLU
12-1784). Laos, Oudomxay Province, Xay District, Houay Houm Village, 28 July 2014, P.
Sysouphanthong, PS2014-567 (HNL502238).
Notes: Macrolepiota detersa is recognized by medium to large and very tall basidiomata,
white to yellowish-white pileus covering with brownish-orange to light-brown central
calotte and patches or squamules, white and free lamellae, long stipe with a draping white
membranous annulus attached at the upper part, amygdaliform basidiospores with germ
pore and hyaline cap, clavate to narrowly clavate cheilocystidia, and the hymenidermal
pileus covering made up of oblong to cylindrical elements.
Macrolepiota detersa was originally described, from a temperate region in the Anhui
province of eastern China, as an edible species by Ge et al. [58]. It was already known from
Japan (as an undescribed species; Vellinga et al. [13] and was later reported from South
Korea [59]. Here we record its presence in northern Thailand and Laos [55].
The ITS sequence of the Thai collection is identical to materials from China, Japan,
and South Korea with 100% bootstrap support, and it is sister to M. excelsa (Figure 4).

Macrolepiota dolichaula (Berk. and Broome) Pegler and Rayner, Kew Bull. 23: 365 (1969)
≡ Agaricus dolichaulus Berk. and Broome, Trans. linn. Soc., Bot. 27: 150 (1870)
≡ Lepiota dolichaula (Berk. and Broome) Sacc., Syll. Fung. 5: 32 (1887)
Diversity 2021, 13, 666 24 of 39

≡ Leucocoprinus dolichaulus (Berk. and Broome) Pat., Bull. trimest. Soc. mycol. Fr. 29:
215 (1913)
= Agaricus beckleri Berk., J. linn. Soc., Bot. 13: 156 (1872)
= Lepiota beckleri (Berk.). Sacc., Syll. Fung. 5: 56 (1887)
= Agaricus stenophyllus Cooke and Massee, Grevillea 15: 98 (1887)
= Lepiota stenophylla (Cooke and Massee) Sacc., Syll. Fung. 9: 4 (1891)
= Leucocoprinus dolichaulus var. cryptocyclus Pat., Bull. trimest. Soc. mycol. Fr. 29: 215 (1913)
Diversity 2021, 13, 666 27 of 43
= Lepiota dolichaula var. cryptocycla (Pat.) Sacc. and Trotter, Syll. Fung. 23: 23 (1925.)
Index Fungorum number: IF 333540; Facesoffungi number: FoF 09699; Figures 17 and 18.

Diversity 2021, 13, 666 27 of 43

Figure 17. Fresh

FigureFigure
17. Fresh basidiomata
17. Fresh
basidiomata ofMacrolepiota
basidiomata of
of Macrolepiota
Macrolepiota dolichaula
dolichaula in
in situ. in
dolichaula situ. (a)
(a) situ.
MFLU (a) MFLU
12-1816.
MFLU 12-1816.
(b) MFLU (b)
12-1776.
12-1816. MFLU
(b) (c) 12-1776. (c)
HNL503204.
MFLU 12-1776. (c) HNL503204.
HNL503204.

Figure 18. Microcharacters of Macrolepiota dolichaula (MFLU 12-1816). (a) pileus covering. (b) basid-
Figure 18. Microcharacters of Macrolepiota dolichaula (MFLU 12-1816). (a) pileus covering. (b) basidiospores.
iospores. (c) cheilocystidia.
(c) cheilocystidia.
Pileus 75–175 mm, first subglobose to globose, soon expanding to campanulate, um-
bonate with high umbo, with inflexed margin, with brownish-orange to light-brown
(6C5–6, 6D6–7) glabrous surface and smooth when young, soon surface radially broken
Diversity 2021, 13, 666 25 of 39

Pileus 75–175 mm, first subglobose to globose, soon expanding to campanulate, um-
bonate with high umbo, with inflexed margin, with brownish-orange to light-brown (6C5–6,
6D6–7) glabrous surface and smooth when young, soon surface radially broken from mar-
gin to center, with light-brown (6C5–6, 6D6–7) radially crowded squamules around central
calotte toward margin; squamules fragile and distant or completely disappearing when ma-
ture, on white fibrillose background; marginal zone with white fibrils, sulcate when mature.
Lamellae free, white, crowded, ventricose, wider at pileus margin and narrow to pileus
center, 20–28 mm wide. Stipe 140–260 × 10–15 mm, cylindrical, white background, white
and smooth from apex to the middle, covered with orange-white to pale-orange (5A2–3)
minute squamules downward base, turning pale-red to pastel (7A3–5) when touched, with
crowded white rhizomorphs at the base. Annulus membranous, descendent, white on the
upper side, with concolorous squamules underside as those on pileus, moveable when
mature. Context, white and moderately thick in pileus, white and hollow in the stipe,
slowly turning pale-red to pastel (7A3–5) in both pileus and stipe. Taste and smell note
observed. Spore print whitish to white.
Basidiospores [100,4,4] 12.5–15 × 8.5–11.5 µm, avl × avw = 13.86 × 10.2 µm, Q = 1.31–1.5,
avQ = 1.37, ellipsoid-ovoid, ellipsoid, to ellipsoid-amygdaliform in the side view, ellipsoid
in frontal view, hyaline and thick-walled, with germ pore and hyalinous cap over germ pore,
dextrinoid, congophilous, cyanophilous, metachromatic. Basidia 30–45 × 10–15 µm, clavate, hya-
line, and thin-walled, 4-spored. Lamella edge sterile. Cheilocystidia 20–30 × 10–14 µm, clavate,
rarely spheropedunculate, slightly thick-walled, hyaline. Pleurocystidia absent. Pileus
covering a hymeniderm made up of oblong to cylindrical elements, terminal elements
oblong, cylindrical, short clavate, with rounded or attenuate apex, 15–55 × 6–9 µm, hyaline
to pale-brown parietal pigment. Stipe covering not observed. Clamp-connections are
present and abundant at the base of basidia and cheilocystidia.
Habitat and distribution: saprotrophic and terrestrial growing in grasslands, un-
der shade trees beside grassland, in pine forests, and on cow dung; solitary or with
many basidiomata spread over a wide area; widely distributed and found throughout
northern Thailand.
Material examined: Thailand, Chiang Rai, Muang District, Mae Fah Luang University
Campus: 20◦ 030 16.900 N; 99◦ 530 42.600 E, alt., 410 m, 7 August 2011, P. Sysouphanthong, 2011-
15 (MFLU 12-1816); Chiang Rai, Muang District, Pong Phra Bath Village, 15 July 2012, P.
Sysouphanthong, 2012-1 (MFLU 12-1764); ibidem, 20 August 2012, P. Sysouphanthong, 2012-
13 (MFLU 12-1776); ibidem, 27 August 2012, P. Sysouphanthong, 2012-15 (MFLU 12-1778);
Chiang Rai, Muang District, Mae Pu Ka Village, 14 June 2009, K. Anchalee, BJP020, culture
(MFLUCC 10-0366); ibidem, 9 July 2009, K. Anchalee, BJP0046 (MFLU 12-1782); Chiang Rai,
Mae Fah Luang District, Mae Salong Nok Village: 20◦ 090 121.700 N; 99◦ 390 26.600 E, alt., 985 m,
22 July 2012, P. Sysouphanthong, 2012-8 (MFLU 12-1771); Chiang Mai, Mae Taeng District,
Pha Deng Village: 19◦ 060 49.100 N; 98◦ 430 33.500 E, alt., 971 m, 16 June 2009, P. Sysouphan-
thong, BJP017, culture (MFLUCC 10-0363). Laos, Xiang Khouang Province, Pek District,
Mak Khay Village, 8 June 2016, P. Sysouphanthong, PS2016-81 (HNL503204); Champasak
Province, Pak Song District, 3 October 2015, P. Sysouphanthong, PS50 (HNL503277).
Notes: All specimens of M. dolichaula are characterized by large basidiomata, a pileus
with small, brownish-orange to light-brown, crowded squamules on a white fibrillose
background, a long stipe with pale-orange minute squamules, and membranous annulus
attached at the upper part of stipe, ellipsoid to ellipsoid-amydaliform spores with germ pore
and cap, clavate cheilocystidia, and oblong to cylindrical elements in the pileus covering.
This species is widespread all through tropical Asia and has been reported from Sri
Lanka [60–63], India [64], and Vietnam [65], both tropical and temperate China [58,66],
Laos [55,67], and is also known from Australia [27,68,69] (Figure 4).
Macrolepiota dolichaula is common in Thailand [17], and it was also recorded from Laos
by Læssøe et al. [55], however, these studies were based on macromorphology only. Thus,
we redescribe this species based on morphology and molecular evidence. In Figure 4, the
nrITS sequences from Thailand and Laos group with specimens from China and Australia
 Diversity 2021, 13, 666 29 of 43
Diversity 2021, 13, 666 26 of 39

Australia with high bootstrap support. Macrolepiota aberdarensis Mbaluto and Otieno, a
with
closehigh bootstrap
relative from support. Macrolepiota
Kenya, does aberdarensis
not differ much from Mbaluto and Otieno,
M. dolichaula a close relative
in morphology and
from
nrITSKenya, does analysis
sequences not differ[70].
much from M. adolichaula
However, multipleingenes
morphology
analysisand nrITS sequences
is required to verify
analysis [70]. However,
both species. a multiple
It is well possible thatgenes analysis
the species is required
recorded to verify
by Pegler andboth species.
Rayner It is
[71] from
East Africa is M. aberdarensis, and not M. dolichaula proper. One should also note thatM.
well possible that the species recorded by Pegler and Rayner [71] from East Africa is M.
aberdarensis, and not
abararensis was M. dolichaula
not validly proper.
published as aOne should also
herbarium note
for the M. abararensis
thatcollections
type was was not
not indi-
validly
cated. published as a herbarium for the type collections was not indicated.

Macrolepiota
Macrolepiotaexcelsa
excelsaVellinga,
Vellinga,Sysouph.,
Sysouph.,Thongkl.
[Link]
andK.D.
[Link]
Hydesp.
[Link].
nov.
Mycobank umbero:
Mycobank umbero: MB 838685; Facesoffungi number: FoF 09700; Figures19
MB 838685; Facesoffungi number: FoF 09700; Figures 19and
and20.
20.

Figure19.
Figure [Link]
Fresh basidiomata
basidiomata of Macrolepiota
of Macrolepiota excelsa
excelsa in [Link](a)
situ. (a) 09-0203.
MFLU MFLU 09-0203. (b) (c)
(b) ecv3553. ecv3553. (c)
ecv3572
ecv3572 (type). (d,e)
(type). (d,e) HNL501921.HNL501921.

Etymology: The Latin name ‘excelsa’ refers to ‘high, lofty, distinguished.’

Diagnosis: Macrolepiota excelsa is recognized by large brown to dark brown pileus, free
and white lamellae, brown to dark brown cylindrical stipe with the white membranous
annulus, broadly ellipsoid to ellipsoid basidiospore with a germ pore, utriform to narrowly
clavate cheilocystidia, a hymeniderm made up of clavate to cylindrical elements in pileus
covering.
Holotype: ecv3572 (UC).
Pileus 120–220 mm, when young globose to subglobose, paraboloid, expanding to
campanulate, convex, umbonate or not, sometimes slightly depressed at the center, when
young completely brown (6E5), later covering broken and with dark brown, irregular
calotte (6F4–8; Mu. 7.5 YR 5/4, 6/6) at the center, with concolorous big patches and
Diversity 2021, 13, 666 27 of 39

squamules around calotte toward the margin, patches fragile when mature; background
radially fibrillose and light to greyish-brown (6D3–5; 10 YR 7/4–7.5 YR 8/6–4), slightly
squarrose especially in the outer part, where the fibrils are more spread out and show
the white background in between; margin irregular and exceeding lamellae. Lamellae,
L = 80–130, l = 0 or 1, moderately crowded to crowded, free and remote (up to 4 mm) from
stipe, off-white, broad and ventricose, up to 12 mm wide, with white floccose (cystidiose)
edge. Stipe 130–350 × 7–10 mm at apex, 10–18 mm wide in the center, cylindrical or slightly
widening downward, 23–30 mm wide at base; base in some specimens bulbous; completely
covered in brown tomentose-velvety light-brown to brown, or reddish-brown (6D4–5,
6E5–6, 7.5 YR 7–6/6) covering, which breaks open into small very fine to fine horizontal
bands that can form zig-zag patterns, hollow with central white cottony strand, protruding
into pileus. Annulus a white descending or ascending cuff, and a flaring double part, white
to yellowish-white (4A2) membranous on the upper side with a fringed edge; underside
with squamules as on pileus; movable with age. Context thick and soft, white and dull in
Diversity 2021, 13, 666 30 of 43
pileus; in stipe buff and hollow. Smell fungoid. Taste not observed. Spore print white.

Figure 20. Microcharacters of Macrolepiota excelsa (ecv3572, Type). (a) basidiospores. (b) Cheilocys-
Figure 20. Microcharacters of Macrolepiota excelsa (ecv3572, Type). (a) basidiospores. (b) Cheilocys-
tidia. (c) pileus covering.
tidia. (c) pileus covering.
Etymology: The Latin name ‘excelsa’ refers to ‘high, lofty, distinguished.’
Diagnosis: Macrolepiota
Basidiospores excelsa11.8–16.5
[90,3,3] × 8.3–13.5
is recognized by large brown
µm, avl × avw
to dark brown 13.2–15.3 × 9.2–11.2 µm,
= pileus,
free and white lamellae, brown to dark brown cylindrical stipe with the white membra-
Q = 1.2–1.51, avQ = 1.3–1.45, in side-view broadly ellipsoid to ellipsoid, with germ pore
nous annulus, broadly ellipsoid to ellipsoid basidiospore with a germ pore, utriform to
covered by thecheilocystidia,
narrowly clavate hyaline cap, thick-walled,
a hymeniderm madehyaline to pale-yellow,
up of clavate dextrinoid, congophilous,
to cylindrical elements
in pileus covering. Basidia 30–40 × 11–19 µm, clavate, slightly thick-walled, 4-spored, rarely
metachromatic.
Holotype:
2-spored, ecv3572
with basal(UC).
clamp-connection. Lamella edge sterile. Cheilocystidia in chains of
Pileus 120–220 mm, when young globose to subglobose, paraboloid, expanding to
up to 5 elements; terminal elements 7–40 × 7–30 µm, variable in size and shape, usually
campanulate, convex, umbonate or not, sometimes slightly depressed at the center, when
subglobose,
young completely occasionally
brown (6E5),clavate, withbroken
later covering rarely cylindrical,
and hyaline.
with dark brown, Pleurocystidia
irregular ca- not found.
Pileus covering
lotte (6F4–8; at umbo
Mu. 7.5 YR 5/4,and 6/6)calotte
at the a hymenoderm
center, made up
with concolorous big of hemispherical
patches and to cylindrical
squamules around
elements, calotte toward
thick-walled, the margin,
hyaline patches fragile 20–92.5
to pale-brown, when mature; background
× 10–12.5 µm. Stipe covering a
radially fibrillose and light to greyish-brown (6D3–5; 10 YR 7/4–7.5 YR 8/6–4), slightly
hymeniderm made up of cylindrical elements and hyphae, 8.7–15 µm wide. Clamp-
squarrose especially in the outer part, where the fibrils are more spread out and show the
connections
white background at in
the base of
between; basidia
margin andand
irregular cheilocystidia.
exceeding lamellae. Lamellae, L = 80–
130, l =Habitat and distribution:
0 or 1, moderately crowded to growing solitary
crowded, free to small
and remote (up togroup, saprotrophic,
4 mm) from stipe, and terrestrial
off-white,
in broad forests
half-open and ventricose,
and on up the
to 12edge
mm wide, with white floccose
of grasslands; known (cystidiose) edge.
from Chiang Mai province of
Stipe 130–350 × 7–10 mm at apex, 10–18 mm wide in the center, cylindrical or slightly
northern Thailand and Oudomxay province of North Laos.
widening downward, 23–30 mm wide at base; base in some specimens bulbous; com-
pletely covered inexamined:
Material Thailand, Chiang
brown tomentose-velvety MaitoProvince,
light-brown Mae Taeng District, Pha Deng
brown, or reddish-brown
Village, 25 June
(6D4–5, 6E5–6, 7.5 YR2007,
7–6/6) E.C. Vellinga
covering, 3553 open
which breaks (UC); ibidem,
into 3 July
small very 2007,
fine to R. Walleyn (coll. E.C.
fine hor-
izontal bands that can form zig-zag patterns, hollow with central white cottony
Vellinga 3599) (UC); ibidem, 28 September 2008, P. Sysouphanthong (MFLU 09-0203); ibidem, strand,
protruding into pileus. Annulus a white descending or ascending cuff, and a flaring dou-
ble part, white to yellowish-white (4A2) membranous on the upper side with a fringed
edge; underside with squamules as on pileus; movable with age. Context thick and soft,
white and dull in pileus; in stipe buff and hollow. Smell fungoid. Taste not observed.
Spore print white.
Diversity 2021, 13, 666 28 of 39

20 June 2009, P. Sysouphanthong and J.K. Lui, BJP0023, culture (MFLUCC 10-0369); Chiang
Mai province, Mae Taeng District, Pongduad Village, 22 June 2009, P. Sysouphanthong
and J.K. Lui, BJP0029, culture (MFLUCC 10-0375). Laos, Oudomxay Privince, Xay District,
Houy Houm Village, 30 June 2014, P. Sysouphanthong, PS2014-260 (HNL501921); ibidem,
16 July 2014, P. Sysouphanthong, PS2014-437 (HNL502108); ibidem, 05 August 2014, P.
Sysouphanthong, PS2014-648 (HNL502319).
Notes: Macrolepiota excelsa is a new species found in northern Thailand and northern
Laos. Macroscopically this species resembles M. procera (Scop.: Fr.) Singer very much in
general appearance, but it differs in the brown, not grey-brown, colors on pileus and stipe.
The ITS sequence, however, does not place this species in the vicinity of M. procera, but
closely related to M. detersa, M. dolichaula, the Australian sequestrate species M. turbinata
T. Lebel, and M. aberdarensis Mbaluto and Otieno from Kenya (Figure 4). Macrolepiota
detersa differs in brownish-orange to light-brown squamules on pileus and yellowish-white
background, the very well-developed hanging annulus, and the branched cheilocystidia
(Figures 15 and 16); the holotype from China showed more clavate to broadly clavate to
pyriform and rarely subfusiform cheilocystidia [58]. The pileus of M. dolichaula is much
lighter colored with small brown squamules on a white background, and more clavate to
sphaeropedunculate cheilocystidia (Figures 19 and 20). Macrolepiota aberdarensis, a species
resembling M. dolichaula, is distinghuised from M. detersa by the lighter color of squamules
on pileus and white background and the cheilocystidia not arranged in chains [70]. Another
species in the same clade, Macrolepiota umbonata H. J. Cho, H. Lee, and Y.W. Lim, described
from South Korea differs from M. excelsa by the distinct umbo on the pileus, the white
background of the pileus surface, and superior and non-membranous annulus [59].
The stipes of M. excelsa have a central cord, similar to the stipes of Coprinus comatus
(O.F. Müll.: Fr.) Pers. and Montagnea arenaria (DC.) Zeller [72]. This is the first report of this
feature outside the Coprinus comatus clade.

Macrolepiota velosa Vellinga and Zhu L. Yang, Mycotaxon 85: 184 (2003)
Diversity 2021, 13, 666 32 of 43
Index Fungorum number: IF 373847; Facesoffungi number: FoF 09701; Figures 21 and 22.

Figure 21. Fresh basidiomata of Macrolepiota velosa in situ. (a,b) HNL502655. (c) MFLU 09-0052. (d)
Figure 21. Fresh basidiomata of Macrolepiota velosa in situ. (a,b) HNL502655. (c) MFLU 09-0052.
HNL502371. (e) MFLU 12-1818. (f) MFLU 09-0055.
(d) HNL502371. (e) MFLU 12-1818. (f) MFLU 09-0055.
Diversity 2021, 13, 666 29 of 39
Figure 21. Fresh basidiomata of Macrolepiota velosa in situ. (a,b) HNL502655. (c) MFLU 09-0052. (d)
HNL502371. (e) MFLU 12-1818. (f) MFLU 09-0055.

Figure 22. Microcharacters

Figure 22. Microcharactersof
ofMacrolepiota
Macrolepiotavelosa
velosa (MFLU
(MFLU 09-0052).
09-0052). (a) (a) basidiospores.
basidiospores. (b) cheilocys-
(b) cheilocystidia.
tidia. (c) pileuscovering.
(c) pileuscovering.

Pileus 100–140 mm, slightly globose when young, expanding to hemispherical, parabolic
to campanulate with wide umbo, straight margin, center with a round to star shape or
calotte, 10–15 mm diam., with uplifted margin, dark brown (8F4) to dark, with dark brown
(8F4) warts or squamules around center toward the margin, squamules long around the
marginal zone, with uplift tip, with orange write (5A2) radially fibrillose, background, with
white to orange (5A2) white remnant partial veil at the center and scattered on the surface,
in mature sample 4–6 grooves, with split margins, exceeding lamellae, fringe. Lamellae
free, white, remote from the stipe, close to crowded, ventricose, up to 13 mm wide with
white fine-floccose at lamellae surface and marginal edge turning brownish-orange at the
edge. Stipe 80–160 × 8–115 mm, long cylindrical, wider at the base, with 20 mm wide
bulb, white, smooth, yellowish-white (4A2) fibrillose at apex, with brown (7E4) fibrillose at
base zone, with white membranous volva at base. Annulus ascending, movable, with cuff,
white to cream at the upper side, grayish-orange (5B3) at underside, thick, with broken
brownish beige (6E3) margin. The context in pileus white, 4–6 mm wide, in stipe white.
Odor such as vitamin B. Taste mild, sweet.
Basidiospores [75,3,3] 9–11.8 × 6.5–7.5 µm, avl × avw + 9.5 × 7.0 µm, Q = 1.28–1.68,
avQ = 1.35, in the side view broadly to ellipsoid-amygdaliform, in frontal view ellip-
soid, with germ pore covered by hyaline cap, thick-walled, dextrinoid, congophilous,
cyanophilous, metachromatic in Cresyl Blue. Basidia 28–35 × 11–13 µm, 4-spored, clavate,
hyaline. Lamella edge sterile, with crowded cheilocystidia. Cheilocystidia 20–85 × 5.5–8 µm,
cylindrical, wide at apex, with granular content at apex; clamp-connections absent. Pleu-
rocystida absent. Pileus covering a trichoderm, in upper layer or squamules, made up of
clavate to long clavate terminal elements, 20.0–120 × 4–13 µm, with brown parietal and
intraocular pigment. Stipe covering a cutis composed of hyaline hyphae and cylindrical
elements up to 7 µm wide. Clamp-connections absent.
Diversity 2021, 13, 666 30 of 39

Habitat and distribution: solitary or in small groups, saprotrophic and terrestrial on

soil or ground with decomposing leaves mixed with humus; distributed in the rainforest in
the highlands in Chiang Mai and Chiang Rai Provinces.
Material examined: Thailand, Chiang Rai Province, Mueng District, Hua Doi Village,
15 August 2009, P. Sysouphanthong and Jainkui Lui, BJP0072, culture (MFLUCC 10-0417);
Chiang Rai, Mae Fah Luang District, Pa Kluay Village, 22, August 2011, P. Sysouphanthong,
2011-14 (MFLU 12-1781); Chiang Mai, Mae Taeng District, Pha Deng Village, 12 August
2007, P. Sysouphanthong, PNG055 (MFLU 09-0055); ibidem, 12 August 2012, P. Sysouphan-
thong, PNG052 (MFLU 09-0052), ibidem, 19 July 2007, P. Sysouphanthong, PNG015 (MFLU
09-0015). Laos, Oudomxay Province, Xay District, Houay Houm Village, 6 August 2014,
P. Sysouphanthong, PS2014-700 (HNL502371); Oudomxay Province, Xay District, Houay
Hoom Village, 6 September 2014, P. Sysouphanthong, PS2014-984 (HNL502655); Xekong
Province, Tha Taeng District, Dongmakjong Forest, 30 September 2015, P. Sysouphanthong,
PS1 (HNL503228).
Notes: Macrolepiota velosa differs from other Macrolepiota species as it is covered with
dark brown to dark squamules on pileus and has a white volva at the basal stipe bulb,
cylindrical to narrowly clavate cheilocystidia; the pileus covering is a hymeniderm made
up of long, narrowly clavate terminal elements, with pale-brown parietal and intracellular
pigment; clamp-connections were not found in the basidiomata. This is also the smallest of
the Thai Macrolepiota species.
Ge et al. [58] and Vellinga et al. [2] already mentioned the occurrence of M. velosa in
northern Thailand. [73] Vellinga and Yang (2003) discussed all volvate Macrolepiota species.
Diversity 2021, 13, 666 The nrITS sequences of the Thai species are grouped with specimens from China 34 of
and43
form a sister group with the volvate M. eucharis from Australia (Figure 4). Sysouphan-
thong et al. [67] recorded M. velosa in central Laos, and here we report the species also from
northern
3.2.4. and southern
Pseudolepiota parts
Z.W. Ge of Laos.

3.2.4. Pseudolepiota
Pseudolepiota zangmuiZ.W.
[Link]
Ge in Ge and Yang, Phytotaxa 312: 252 (2017).
Pseudolepiota zangmui Z.W. Ge in Ge and Yang, Phytotaxa 312: 252 (2017)
Index Fungorum number: IF 820482; Facesoffungi number: FoF 07068; Figures 23 and
24. Index Fungorum number: IF 820482; Facesoffungi number: FoF 07068; Figures 23 and 24.

Figure 23.
Figure 23. Fresh
Fresh basidiomata
basidiomata of
of Pseudolepiota
Pseudolepiota zangmui.
zangmui. (a)
(a) MFLU
MFLU 10-0515.
10-0515. (b)
(b) MFLU
MFLU 10-0518.
10-0518. (c,d) MFLU 192-355.
(c,d) MFLU 192-355.
Diversity 2021, 13, 666 31 of 39
Diversity 2021, 13, 666 35 of 43

Figure 24. Microcharacters of Pseudolepiota zangmui (MFLU 10-0515). (a) basidiospores. (b) basidia.
Figure 24. Microcharacters of Pseudolepiota zangmui (MFLU 10-0515). (a) basidiospores. (b) basidia.
(c) cheilocystidia. (d) pileus covering.
(c) cheilocystidia. (d) pileus covering.
Pileus 20–50 mm, at first hemispherical, campanulate, expanding to convex, umbo-
nate Pileus
or often20–50 mm, at with
plano-convex first hemispherical,
an umbo and inflexedcampanulate, expanding
margin; surface coveringto violet-
convex, umbonate
or often(10F5-8,
brown plano-convex withatan
11F-8), darker umbo
umbo and inflexed
or center, dark ruby margin;
to dark surface
(12F4–8),covering
smooth orviolet-brown
rough, later
(10F5-8, splitting
11F-8), off and
darker at becoming
umbo orirregularly
center, dark concolorous
ruby tosquamulose,
dark (12F4–8),with larger
smooth or rough,
scales around the umbo and with smaller squamules toward
later splitting off and becoming irregularly concolorous squamulose, with the margin, with dull redlarger scales
(9B3–4) fibrillose or fibrillose squamules at the marginal zone on a white to orange-white
around the umbo and with smaller squamules toward the margin, with dull red (9B3–4)
(6A2) background; squamules are fragile when mature; margin appendiculate, with white
fibrillose or fibrillose squamules at the marginal zone on a white to orange-white (6A2)
to dull red (9B3–4) fibrillose remnants. Lamellae free, at first white, turning yellowish-
background;
white when mature squamules are fragile (5B3–4)
and greyish-orange when mature;
when dried, margin appendiculate,
3.5–5 mm wide, crowded, with white to
dull
with red
4–5 (9B3–4)
lamellulae fibrillose
in betweenremnants. Lamellae
lamellae, slightly free,with
fragile at first
age, white,
with an turning yellowish-white
eroded edge.
when mature
Stipe 25–40 × 5–8and
mm,greyish-orange
equal, slightly wider(5B3–4)
at basewhen dried, 3.5–5
zone, completely mmby
covered wide,
whitecrowded,
to with
yellowish-white
4–5 lamellulae fibrils, sometimes
in between with white
lamellae, remnants
slightly at the
fragile with annular zone. an
age, with Annulus
eroded a edge. Stipe
white annular
25–40 × 5–8 zone.
mm,Context
equal,white in pileus,
slightly widerup toat4 base
mm wide,
zone, white in the stipe,covered
completely changing by white to
to reddish with age, hollow. Spore print whitish. Taste and smell not observed.
yellowish-white fibrils, sometimes with white remnants at the annular zone. Annulus a
Basidiospores [2,2,50] 4.8–5.2 × 3.5–4.5 µm, avl × avw = 5.0 × 4.0 µm, Q = 1.15–1.37,
white annular
Qav = 1.25, zone. Context
in side-view white intopileus,
broadly ellipsoid up to 4 mm
ellipsoid-ovoid, wide,view
in frontal white in the
broadly stipe, changing
ellip-
to reddish
soid withhyaline
to ellipsoid, age, hollow. Spore in
to pale-yellow print
waterwhitish.
and KOH, Taste andthick-walled
slightly smell not observed.
without
a germBasidiospores
pore, dextrinoid,[2,2,50] 4.8–5.2 ×
congophilous, 3.5–4.5 µm,not
cyanophilous, avl × avw = 5.0Basidia
metachromatic. × 4.015–18
µm, ×Q = 1.15–1.37,
Qav = 1.25, in side-view broadly ellipsoid to ellipsoid-ovoid, in frontal view broadly
ellipsoid to ellipsoid, hyaline to pale-yellow in water and KOH, slightly thick-walled
without a germ pore, dextrinoid, congophilous, cyanophilous, not metachromatic. Basidia
15–18 × 4.8–5.0 µm, clavate, 4-spored, some 2-spored. Lamella edge sterile. Cheilocystidia
15–24 × 6–10 µm, most clavate, some with a long stalk, rarely utriform and sphaeropedun-
culate, thin-walled, hyaline. Pleurocystidia absent. Pileus covering hymenoderm made
up of 3–4 layers of elements, oblong to cylindrical in the lower layer, mostly with clavate,
rarely subclavate, elements in the upper layer, 12–30 × 5.0–10 µm, thick-walled, smooth
Diversity 2021, 13, 666 32 of 39

and with hyaline to pale-yellow parietal pigments, with brown intracellular pigments and
encrusted walls in some lower elements. Clamp connections absent.
Habitat and distribution: in small to large groups, saprotrophic and terrestrial in
deciduous forests dominated by Lithocarpus spp. and Castanopsis spp. at 650–950 m alt.
Found in China and Thailand.
Material examined: THAILAND, Chiang Rai Province, Muang District, Forest of
Hua Doi Village, 23 September 2009, P. Sysouphanthong, PHO25 (MFLU 10-0515); ibidem,
25 September 2009, P. Sysouphanthong, PHO28 (MFLU 10-0518); Chiang Rai Province,
Muang District, Mae Fah Luang University Campus, 13 August 2019, P. Sysouphanthong,
PS2019-74 (MFLU 19-2360). Chiang Mai Province, Mae Taeng District, Pha Deng Village,
alt. 900–950 m, 7 August 2018, P. Sysouphanthong, PS2018-84 (MFLU 19-2355).
Notes: Pseudolepiota zangmui is characterized by squat basidiomata in which the pileus
diameter is equal or slightly longer than the stipe length; the pileus is covered in violet-
brown to dark ruby squamules with dull red fibrils between squamules, and it has a white
appendiculate margin; the basidiospores are pale-yellow, and oblong-ovoid in side-view,
the cheilocystidia are clavate, and the pileus covering is hymenidermal.
The type of Pseudolepiota zangmui was described from Xishuangbanna, Yunnan Province,
Southwest Yunnan, China [59]; it closely resembles Xanthagaricus species but differs in
the white to pale-cream spore print and the squat basidiomata. Based on phylogenetic
analyses of combined ITS and LSU data with a wide sampling of Xanthagaricus species, it
is closely related and basal to Xanthagaricus with good support (Figure 1). However, in a
phylogenetic analysis including protein-coding genes [32], Pseudolepiota is clearly separated
from Xanthagaricus.

3.2.5. Xanthagaricus (Heinem.) Little Flower, Hosag. and T.K. Abraham

Xanthagaricus necopinatus Iqbal Hosen, T.H. Li, and G.M. Gates in Hosen, Song, Gates,
Karunarathna and Li, MycoKeys 28: 9 (2017).
Index Fungorum number: IF 820482; Facesoffungi number: FoF 07068; Figures 25 and 26.
Pileus 10–20 mm, first paraboloid, expanding to convex, plano-convex, with or without
umbo, with straight or slightly incurved margin; when young covered with crowded
squamules, greyish-yellow to yellow (3B3–8), with brownish-yellow squamules (5C7–8)
at center; when mature with yellowish-white to yellow (3A2–6) squamules on yellowish-
white to pastel yellow (3A2–4) background; margin with concolorous squamules as on
pileus surface and with light-orange (6A4–5) appendiculate velar remnants. Lamellae,
l = 3 series, free, 2–4 mm wide, yellowish-white to orange-white (4A2, 5A2) when young,
becoming orange-grey to greyish-orange (5B2–3) when mature, broadly ventricose, with
white, eroded edge. Stipe 20–45 × 2–3 mm, cylindrical, white fibrillose at apex zone, with
yellowish-white to yellow (3A2–6) squamules below annulus to base, with orange-grey
to greyish-orange (5B2–3) background, turning greyish red (7B4–5). Annulus attached to
the apical zone of the stipe, membranous, greyish-orange (6B4–6), sometimes fragile with
age. Context white to orange-white (5A2) in pileus, up to 2 mm thick at the center; hollow
in stipe and concolorous with the surface. Odor and taste not observed. Spore pint not
observed.
Basidiospores [75,3,3] 4.0–5.2 × 2.5–3.5 µm, avl × avw = 4.71 × 2.90 µm, Q = 1.42–1.80,
avQ = 1.62, ellipsoid to oblong-ovoid in frontal view, ellipsoid to oblong in side-view, thick-
walled, smooth, hyaline to pale-brown, without a germ pore. Basidia 15–20 × 4.5–7 µm,
short clavate to clavate, thin-walled, hyaline, 4-spored. Pleurocystidia absent. Cheilocys-
tidia 17–25 × 5–10 µm, short clavate to clavate, often ellipsoid, with 1 or 2 septa at the base,
slightly thick-walled, hyaline. Pileus covering an irregular epithelium composed of oblong
to short clavate elements in the upper layer, 10–20 × 5–15 µm, with irregular globose to
subglobose elements in the lower layer, 5–15 µm wide, slightly thick-walled, rough-walled,
encrusted, with pale-brown to brown parietal and intracellular pigments, with hyaline
to brown rough-walled and encrusted, up to 8 µm wide hyphae at base of the epithelial
layers. Stipe covering an epithelium similar to that on pileus. Clamp connections absent.
Diversity 2021, 13, 666 33 of 39
Diversity 2021, 13, 666 37 of 43

Diversity 2021, 13, 666 37 of 43

Figure 25. Basidiomata of Xanthagaricus necopinatus in situ. (a) MFLU 19-2353. (b) MFLU 19-2359.
Figure 25. Basidiomata of Xanthagaricus necopinatus in situ. (a) MFLU 19-2353. (b) MFLU 19-2359.
(c) MFLU 19-2358.
(c) MFLU 19-2358. Figure 25. Basidiomata of Xanthagaricus necopinatus in situ. (a) MFLU 19-2353. (b) MFLU 19-2359.
(c) MFLU 19-2358.

Figure 26. Microcharacters of Xanthagaricus necopinatus (MFLU 19-2353). (a) basidiospores. (b) basidia. (c) cheilocystidia.
Figure
(d) 26. Microcharacters of Xanthagaricus necopinatus (MFLU 19-2353). (a) basidiospores.
pileus covering.
Figure 26. Microcharacters(b)
of Xanthagaricus necopinatus (MFLU
basidia. (c) cheilocystidia. 19-2353).
(d) pileus (a) basidiospores. (b) basidia. (c) cheilocystidia.
covering.
(d) pileus covering.
Habitat and distribution: growing in large groups, saprotrophic on humus-rich soil
with dead leaves and wood under trees or in grassland; commonly found in Chiang Mai
and Chiang Rai Provinces, northern Thailand.
Material examined: Thailand, Chiang Mai Province, Muang District, Chiangmai
University Campus, 28 July 2018, P. Sysouphanthong, PS2018-56 (MFLU 19-2353); Chiang
Rai Province, Muang District, Forest of Mae Fah Luang University Campus, 8 June 2019,
Diversity 2021, 13, 666 34 of 39

P. Sysouphanthong, PS2019-45 (MFL 19-2358); ibidem, 8 August 2019, P. Sysouphanthong,

PS2019-67 (MFL 19-2359).
Notes: Xanthagaricus necopinatus has the slender basidiomata characteristic for the
genus, and a pale-yellow pileus and stipe with squamules, velar remnants on the pileus
margin, hyaline to pale-brown basidiospores, clavate to ellipsoid cheilocystidia, and an
irregular epithelium on pileus and stipe; clamp-connections are absent. The basidiospores
are smooth.
Xanthagaricus necopinatus was originally described from Bangladesh [34], and this is
the first report of this species outside that country. Thai specimens completely resemble the
type specimens, but the stipe covering of type specimens is a cutis while the Thai specimens
have an epithelium on the stipe. However, the nrITS sequences of Thai specimens are
clustered with the type specimens with high BS (Figure 1).
Xanthagaricus necopinatus can be confused with X. flavosquamosus T.H. Li, Iqbal Hosen
and Z.P. Song, described from China, but differs from that species in the smoother pileus
and the smooth (vs. rough) basidiospores. The differences in the width and shape of the
cheilocystidia as mentioned by Hosen et al. [74], are not supported by the data from the
Thai collections.

Diversity 2021, 13, 666 Xanthagaricus purpureosquamulosus Sysouph., Thongkl. and K.D. Hyde sp. nov. 39 of 43
Mycobank umbero: MB 833740; Facesoffungi number: FoF 07071; Figures 27 and 28.

Figure27.
Figure 27. Fresh
Fresh basidiomata
basidiomata of
of Xanthagaricus
Xanthagaricus purpureosquamulosus.
purpureosquamulosus.(a–c)
(a–c)MFLU
MFLU19-2354
19-2354(holotype).
(holotype).
(d) MFLU 192356.
(d) MFLU 192356.
Diversity 2021, 13, 666 35 of 39
Figure 27. Fresh basidiomata of Xanthagaricus purpureosquamulosus. (a–c) MFLU 19-2354 (holotype).
(d) MFLU 192356.

Figure 28. Microcharacters of Xanthagaricus purpureosquamulosus (MFLU 19-2354, holotype). (a) basidiospores.
basidiospores. (b)
(b) basidia.
basidia.
(c) cheilocystidia. (d) pileus covering.
(c) cheilocystidia. (d) pileus covering.

Etymology: from purpureus (L) purple, and squamulosus (L) with small scales, as the
basidiomata are covered with purple squamules.
Diagnosis: Xanthagaricus purpureosquamulosus has slender basidiomata, characterized
by a pileus with brownish-grey to violet-brown squamules on a pale-violet to violet
background, brownish-grey to greyish-brown appendiculate velar remnants, white to
greyish lamellae, a stipe with greyish-yellow to light-brown squamules, and a small apical
annulus, ellipsoid-ovoid basidiospores, which are convex on adaxial side, with hyaline to a
pale-brown wall, short clavate to ellipsoid cheilocystidia, an irregular epithelium as pileus
and stipe covering, and no clamp connections.
Holotype: MFLU 19-2354.
Pileus 4.5–10 mm, first paraboloid or hemispherical, expanding to convex, plano-
convex, with or without low umbo, when young completely covered with crowded squa-
mules, brownish-grey (9D2–3) to violet-brown (10D4–8), soon breaking up into greyish-
brown (9E2, 9D3) to violet-brown (10E4) squamules, crowded around center or umbo and
with radially arranged squamules toward the margin, on pale-violet to violet (17A3–7)
background and paler when mature; margin incurved, with brownish-grey (9D2–3) to
greyish-brown (9E2, 9D3) appendiculate triangular velar remnants. Lamellae, l = 3–4 series,
free, 1.5–2 mm wide, white to yellowish-white (4A2), becoming yellowish-grey to greyish-
yellow (4B2–3) when mature, broadly ventricose, with white, smooth to slightly eroded
margin. Stipe 15–30 × 1–1.5 mm, slender and tapering downwards, covered with white
to yellowish-white (4A2) squamules at apex, bellow annulus with crowded squamules
toward the base, greyish-yellow, greyish-brown to light-brown (6D3–5), on yellowish-white
to pale-yellow (4A2–3) background, darker at base zone, with greyish-brown (7E3–4) squa-
mules. Annulus attached to apical zone of stipe, with velar remnants and concolorous to
those on pileus margin, sometimes fragile with age. Context white in pileus, up to 1 mm
thick at center; hollow in stipe and concolorous with surface. Odor and taste not observed.
Spore pint greyish-orange (5B3).
Basidiospores [50,2,2] 4.5–6 × 3.5–4 µm, avl × avw = 5.34 × 3.78 µm, Q = 1.3–1.5,
avQ = 1.4, ellipsoid-ovoid in frontal view, ellipsoid in side-view, thick-walled, smooth,
hyaline to pale-brown, without a germ pore. Basidia 10–12 × 5.5–7 µm, obovoid, short
clavate to ellipsoid, thin-walled, hyaline, 4-spored. Pleurocystidia absent. Cheilocystidia
10–16 × 5–8 µm, short clavate to clavate, ellipsoid, some branched, slightly thick-walled,
hyaline. Pileus covering an irregular epithelium composed of globose to subglobose (7.5–
Diversity 2021, 13, 666 36 of 39

25 µm wide) cells in the upper layer, broadly ellipsoid to oblong cells (10–62 × 5–15 µm) in
the lower layer, thin-walled, smooth, with pale-brown to brown parietal and intracellular
pigments, with pale-brown to hyaline, up to 7 µm wide hyphae at base of the epithelial
layers. Stipe covering an irregular epithelium same as on pileus. Clamp connections absent.
Habitat and distribution: growing in small to large groups, saprotrophic on humus-
rich soil with dead leaves and wood under trees of Samanea saman or beside grassland;
commonly found on Mae Fah Luang University Campus, Chiang Rai, Thailand.
Additional material examined: Thailand, Chiang Rai Province, Muang District, Mae
Fah Luang University Campus, 26 October 2018, P. Sysouphanthong, PS2018-217 (MFL19-
2356, paratype).
Notes: there are only two species of Xanthagaricus, which resemble X. purpureosquamu-
losus in pileus color. X. caeruleus Iqbal Hosen, T.H. Li and Z.P. Song, described from China
has pale-lilac to grayish-lilac or grayish-violet squamules on a pale-grayish-lilac to violet
white background, but it differs from X. purpureosquamulosus by having white lamellae,
which turn light-blue or pastel blue to grayish and finally ink-blue or blackish blue [74].
Based on the phylogenetic analysis of ITS and LSU sequence data, X. caeruleus is not closely
related to X. purpureosquamulosus (Figure 1). The second species with similar colors is X.
ianthinus Y. Li and F.J. Wang, also from China, which differs from X. purpureosquamulosus
by having bluish-violet to violet, more or less violet-brown squamules, white velar rem-
nants, and yellowish-white to light-pinkish-white (4A2) lamellae [75]. It is not close to
X. purpureosquamulosus either in the phylogenetic tree based on ITS and LSU sequences
(Figure 1).
Xanthagaricus thailandensis J. Kumla, N. Suwannarach, and S. Lumyong, recently
described from Thailand, differs in the larger pileus (30−45 mm), and the pale-orange to
grayish-orange, large squamules on the pileus [35].
The phylogenetic analysis of the ITS and LSU sequences (Figure 1) shows that X.
purpureosquamulosus is closely related to a group of species with yellow to olive-brown
squamules, viz. X. epipastus (Berk. and Broome) S. Hussain, known from Sri Lanka [33,76],
X. pakistanicus Hussain, Afshan and Ahmad from Pakistan [77], and X. necopinatus.

Author Contributions: Conceptualization, P.S., E.C.V., N.T. and J.-K.L.; methodology, E.C.V., P.S.,
J.-K.L. and N.T.; formal analysis, J.-K.L., E.C.V., P.S. and N.T.; data curation, E.C.V., P.S., N.T.
and J.-K.L.; supervision, N.T. and E.C.V.; writing—original draft, P.S., E.C.V., N.T. and J.-K.L.;
writing—review and editing, E.C.V., P.S., N.T. and J.-K.L. All authors have read and agreed to the
published version of the manuscript.
Funding: We were supported by Thailand research fund grants “Study of saprobic Agaricales in
Thailand to find new industrial mushroom products” (Grant No. DBG6180015) and Thailand Science
Research and Innovation (TSRI) grant, Macrofungi diversity research from the Lancang-Mekong
Watershed and surrounding areas (Grant No. DBG6280009). The fieldwork of ECV in 2007 was
conducted under an MTA agreement with Dr. Kasem Soytong of KMITL. Financial support of ECV
and PS was from NSF grant DEB 0618293.
Institutional Review Board Statement: Not applicable.
Informed Consent Statement: Not applicable.
Data Availability Statement: Data can be found within the manuscript.
Acknowledgments: Kevin D. Hyde is thanked for his advising and providing a laboratory for
fieldwork at the Mushroom Research Centre in Chiang Mai, Thailand. Mae Fah Luang University
is thanked for providing laboratory facilities for microscopical study and herbarium. Biotech-
nology and Ecology Institute, Ministry of Science and Technology of Laos is also thanked for
depositing herbarium.
Conflicts of Interest: The authors declare no conflict of interest.
Diversity 2021, 13, 666 37 of 39

References
1. Hyde, K.D.; Norphanphoun, C.; Chen, J.; Dissanayake, A.J.; Doilom, M.; Hongsanan, S.; Jayawardena, R.S.; Jeewon, R.; Perera,
R.H.; Thongbai, B.; et al. Thailand’s amazing diversity—Up to 96% of fungi in northern Thailand are novel. Fungal Divers. 2018,
93, 215–239. [CrossRef]
2. Vellinga, E.C.; Sysouphanthong, P.; Hyde, K.D. The family Agaricaceae: Phylogenies and two new white-spored genera. Mycologia
2011, 103, 494–509. [CrossRef] [PubMed]
3. He, M.Q.; Zhao, R.L.; Hyde, K.D.; Begerow, D.; Kemler, M.; Yurkov, A.; McKenzie, E.H.C.; Raspé, O.; Kakishima, M.; Sánchez-
Ramírez, S.; et al. Notes, outline and divergence times of Basidiomycota. Fungal Divers. 2019, 99, 105–367. [CrossRef]
4. Kalichman, J.; Kirk, P.M.; Matheny, P.B. A compendium of generic names of agarics and Agaricales. Taxon 2020, 69, 425–447.
[CrossRef]
5. Sysouphanthong, P.; Hyde, K.D.; Chukeatirote, E.; Bahkali, A.H.; Vellinga, E.C. Lepiota (Agaricales) in northern Thailand-1 Lepiota
section Stenosporae. Mycotaxon 2011, 117, 53–85. [CrossRef]
6. Sysouphanthong, P.; Hyde, K.D.; Chukeatirote, E.; Bahkali, A.H.; Vellinga, E.C. Lepiota (Agaricales) in northern Thailand–2 Lepiota
section Lepiota. Cryptogam. Mycol. 2012, 33, 25–42. [CrossRef]
7. Sysouphanthong, P.; Hyde, K.D.; Vellinga, E.C.; Chukeatirote, E. Diversity of Lepiota (Agaricales) in northern Thailand. Mycol. Int.
J. Fungal. Biol. 2013, 4, 22–28.
8. Sysouphanthong, P.; Guo, J.; Hyde, K.D.; Xu, J.; Mortimer, P.E. Lepiota thailandica (Agaricaceae), a new species from Thailand.
Phytotaxa 2016, 245, 262–270. [CrossRef]
9. Sysouphanthong, P.; Hyde, K.D.; Chukeatirote, E.; Bahkali, A.H.; Vellinga, E.C. Verrucospora vulgaris (Agaricaceae, Agaricales), a
rare tropical species and a new record for Thailand. Chiang Mai J. Sci. 2013, 40, 289–293.
10. Ge, Z.W.; Jacobs, A.; Vellinga, E.C.; Sysouphanthong, P.; van der Walt, R.; Lavorato, C.; An, Y.F.; Yang, Z.L. A multi-gene phylogeny
of Chlorophyllum (Agaricaceae, Basidiomycota): New species, new combination and infrageneric classification. MycoKeys 2018, 32,
65–90. [CrossRef] [PubMed]
11. Dutta, A.K.; Bera, S.; Paloi, S.; Rakshit, S.; Tarafder, E.; Sherpa, A.R.; Acharya, K. Lepiotaceous fungi of West Bengal, India: The
genus Chlorophyllum. Phytotaxa 2020, 451, 113–131. [CrossRef]
12. Loizides, M.; Alvarado, P.; Dimou, D.; Zervakis, G.I.; Thines, M.; Telle, S.; Konstantinou, G.; Gube, M. Multiple evolutionary
origins of sequestrate species in the genus Chlorophyllum. Mycologia 2020, 112, 400–422. [CrossRef] [PubMed]
13. Vellinga, E.C.; de Kok, R.P.J.; Bruns, T.D. Phylogeny and taxonomy of Macrolepiota (Agaricaceae). Mycologia 2003, 95, 442–456.
[CrossRef] [PubMed]
14. Vellinga, E.C. Genera in the family Agaricaceae—Evidence from nrITS and nrLSU sequences. Mycol. Res. 2004, 108, 354–377.
[CrossRef] [PubMed]
15. Gube, M. Ontogeny and Phylogeny of Gasteroid Members of Agaricaeae (Basidiomycetes). Ph.D. Thesis, Friedrich-Schiller-
Universität, Jena, Germany, 2009. Available online: [Link] (accessed on
10 December 2021).
16. Gube, M.; Vellinga, E.C.; Alvarado, P.; Polemis, E.; Dimou, D.M.; Zervakis, G.I.; Loizides, M. (2749) Proposal to conserve
Chlorophyllum against the additional name Secotium (Agaricaceae). Taxon 2020, 69, 819–820. [CrossRef]
17. Chandrasrikul, A.; Suwanarit, P.; Sangwanit, U.; Lumyong, S.; Payapanon, A.; Sanoamuang, N.; Pukahuta, C.; Petcharat, V.;
Sardsud, U.; Duengkae, K.; et al. Checklist of Mushrooms (Basidiomycetes) in Thailand; Office of Natural Resources and Environmental
Policy and Planning: Bangkok, Thailand, 2011; 448p.
18. Soytong, K. Mushrooms and Macrofungi in Thailand; Siritham Offset Publishers Ltd.: Ubonratchathani, Thailand, 1994; 151p. (In Thai)
19. Høiland, K.; Schumacher, T. Agarics, clavarioid and some heterobasidiomycetous fungi from Northern Thailand. Nord. J. Bot.
1982, 2, 265–271. [CrossRef]
20. Berkeley, M.J. Decades of fungi. Decade XV-XIX. Ceylon fungi. Lond. J. Bot. 1847, 6, 479–514.
21. Hosen, M.I.; Ge, Z.W. Clarkeinda trachodes (Agaricales, Basidiomycetes), first record from Bangladesh. Mycotaxo 2011, 118, 331–336.
[CrossRef]
22. Leelavathy, K.K.; Zachariah, S.; Sankaran, K.V. Clarkeinda trachodes—An agaric new to India. Mycologia 1981, 73, 204–207.
23. Verma, R.K.; Tiwari, C.; Parihar, J.; Kumar, S. Diversity of macro-fungi in central India-II: Clarkeinda trachodes. Van Sangyan. 2016,
3, 17–20.
24. Pegler, D.N. Agaric Flora of Sri Lanka. Kew Bull. Addit. Ser. 1986, 12, 1–519.
25. Yang, Z.L. Clarkeinda trachodes an agaric new to China. Acta Bot. Yunnanica 1991, 13, 279–282.
26. Vellinga, E.C.; Noordeloos, M.E. Glossary. In Flora Agaricina Neerlandica 5; Noordeloos, M.E., Kuyper, T.W., Vellinga, E.C., Eds.;
A.A. Balkema Publishers: Tokyo, Japan, 2001; pp. 6–11.
27. Vellinga, E.C. Chlorophyllum and Macrolepiota (Agaricaceae) in Australia. Aust. Syst. Bot. 2003, 16, 361–370. [CrossRef]
28. Lebel, T.; Syme, A. Sequestrate species of Agaricus and Macrolepiota from Australia: New species and combinations and their
position in a calibrated phylogeny. Mycologia 2012, 104, 496–520. [CrossRef] [PubMed]
29. Junopat, M.; Junopat, W.; Jongjitvimol, T.; Chiengkul, A.; Sanpote, P. Survey of macrofungi in Doi Vieng La Wildlife reserve, Mae
Hong Son Province. Naresuan Univ. Sci. J. 2006, 2, 175–181.
30. Klinhom, U. Checklist of edible mushrooms in Northeast Thailand of survey from 2002–2007 (I). Inf. Mushroom Cultiv. 2008, 13,
8–14. (In Thai)
Diversity 2021, 13, 666 38 of 39

31. Sanoamuang, N. Wild Mushrooms of Thailand: Biodiversity and Utilization; Khon Kaen University: Universal Graphic & Trading
Ltd.: Bangkok, Thailand, 2010; 424p. (In Thai)
32. Ge, Z.W.; Yang, Z.L. Pseudolepiota zangmui gen. et sp. nov. (Agaricaceae, Basidiomycota), a new white-spored mushroom from
China. Phytotaxa 2017, 312, 247–255. [CrossRef]
33. Heinemann, P.; Little Flower, S.R. Hymenagaricus (Agaricaceae) de Kerala (Inde) et de Sri Lanka. Bull. Du Jard. Bot. Natl. De Belg.
1984, 54, 151–182. [CrossRef]
34. Hosen, M.I.; Song, Z.P.; Gates, G.; Karunarathna, S.C.; Chowdhury, M.S.M.; Li, T.H. Two new species of Xanthagaricus and some
notes on Heinemannomyces from Asia. MycoKeys 2017, 28, 1–18. [CrossRef]
35. Kumla, J.; Suwannarach, N.; Sringernyuang, K.; Lumyong, S. Xanthagaricus thailandensis sp. nov. (Agaricales, Basidiomycota),
from northern Thailand. Phytotaxa 2018, 348, 109–117. [CrossRef]
36. Liu, S.Y.; Liu, J.K.; Syouphanthong, P.; Hyde, K.D.; Lumyong, S. Xanthagaricus siamensis sp. nov. (Agaricaceae), a new species
with dull green lamellae from northern Thailand. Phytotaxa 2020, 437, 14–22. [CrossRef]
37. Kornerup, A.; Wanscher, J.H. Methuen Handbook of Colour, 3rd ed.; Eyre Methuen: London, UK, 1978; 243p.
38. Gardes, M.; Bruns, T.D. ITS primers with enhanced specificity for basidiomycetes—Application to the identification of mycorrhizae
and rusts. Mol. Ecol. 1993, 2, 113–118. [CrossRef] [PubMed]
39. White, T.J.; Bruns, T.; Lee, S.; Taylor, J.W. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics.
In PCR Protocols: A Guide to Methods and Applications; Innis, M.A., Gelfand, D.H., Sninsky, J.J., White, T.J., Eds.; Academic Press:
San Diego, CA, USA, 1990; pp. 315–322. [CrossRef]
40. Katoh, K.; Misawa, K.; Kuma, K.; Miyata, T. MAFFT: A novel method for rapid multiple sequence alignment based on fast Fourier
transform. Nucleic Acids Res. 2002, 30, 3059–3066. [CrossRef] [PubMed]
41. Katoh, K.; Toh, H. Recent developments in the MAFFT multiple sequence alignment program. Brief. Bioinform. 2008, 9, 286–298.
[CrossRef] [PubMed]
42. Stamatakis, A.; Hoover, P.; Rougemont, J. A rapid bootstrap algorithm for the RAxML webservers. Syst. Biol. 2008, 75, 758–771.
[CrossRef]
43. Nylander, J.A.A. Modeltest v2. Program Distributed by the Author; Evolutionary Biology Centre, Uppsala University: Uppsala,
Sweden, 2004.
44. Ronquist, F.; Huelsenbeck, J.P. MrBayes 3: Bayesian phylogenetic inference under mixed models. Bioinformatics 2003, 19,
1572–1574. [CrossRef] [PubMed]
45. Page, R.D. TreeView: An application to display phylogenetic trees on personal computers. Comput. Appl. Biosci. 1996, 12, 357–358.
46. Li, G.J.; Hyde, K.D.; Zhao, R.L.; Hongsanan, S.; Abdel-Aziz, F.A.; Abdel-Wahab, M.A.; Alvarado, P.; Alves Silva, G.; Ammirati,
J.F.; Ariyawansa, H.A.; et al. Fungal diversity notes 253–366: Taxonomic and phylogenetic contributions to fungal taxa. Fungal
Divers. 2016, 78, 1–237. [CrossRef]
47. Patouillard, N. Champignons nouveaux du Tonkin. Bull. Trimest. De La Soc. Mycol. De Fr. 1907, 23, 69–79.
48. Mossebo, D.C.; Akoa, A.; Eteme, R.A. Macrolepiota globosa et Nothopanus nsimalenensis, deux nouvelles espèces fongiques du
Cameroun. Mycotaxon 2000, 76, 267–278.
49. Vellinga, E.C. New combinations in Chlorophyllum. Mycotaxon 2002, 83, 415–417.
50. Akers, B.P.; Sundberg, W.J. Leucoagaricus hortensis: Some synonyms from Florida and taxonomic observations. Mycotaxon 1997, 62,
401–419.
51. Kumari, B.; Atri, N.S.; Kaur, M. Chlorolepiota indica sp. nov. A new species from India. Mycoscience 2013, 54, 257–259. [CrossRef]
52. Reid, D.A.; Eicker, A. A comprehensive account of Chlorophyllum molybdites. Bot. Bull. Acad. Sin. 1991, 32, 317–333.
53. Vellinga, E.C. Een tropisch paddestoelenparadijs. Coolia 1990, 33, 78–79.
54. Watling, R. A striking addition to the British mycoflora. Mycologist 1991, 5, 23. [CrossRef]
55. Læssøe, T.; Pedersen, O.S.; Sysouphanthong, P. An Introduction to the Edible, Poisonous and Medicinal Fungi of Northern Laos;
Privately Published: Bangkok, Thailand, 2019; 192p.
56. Bijeesh, C.; Vrinda, K.B.; Pradeep, C.K. Mushroom poisoning by Chlorophyllum molybdites in Kerala. J. Mycopathol. Res. 2017,
54, 477–483.
57. Wang, N.; Zhao, Z.; Gao, J.; Tian, E.; Yu, W.; Li, H.; Zhang, J.; Xie, R.; Zhao, X.; Chen, A. Rapid and Visual Identification of
Chlorophyllum molybdites With Loop-Mediated Isothermal Amplification Method. Front. Microbiol. 2021, 12, 638315. [CrossRef]
58. Ge, W.; Yang, Z.L.; Vellinga, E.C. The genus Macrolepiota (Agaricaceae, Basidiomycota) in China. Fungal Divers. 2010, 45, 81–98.
[CrossRef]
59. Cho, H.J.; Lee, H.; Park, M.S.; Kim, C.; Wisitrassameewong, K.; Lupala, A.; Park, K.H.; Kim, M.J.; Fong, J.J.; Lim, Y.W. Macrolepiota
in Korea: New records and a new species. Mycobiology 2019, 47, 310–368. [CrossRef]
60. Pegler, D.N. A revision of the genus Lepiota from Ceylon. Kew Bull. 1972, 27, 155–202. [CrossRef]
61. Pegler, D.N. A preliminary Agaric Flora of East Africa; Kew Bull. Additional Series 6: 1–615; Her Majesty’s Stationery Office:
London, UK, 1977.
62. Petch, T. Revision of Ceylon Fungi. Annals royal botanic gardens. Peradeniya 1907, 4, 21–68.
63. Petch, T.; Bisby, G.R. The fungi of Ceylon. Perad. Manua 1950, 6, 1–111.
64. Natarajan, K.; Manjula, P. South Indian Agaricales. Indian J. Bot. 1981, 4, 50–59.
65. Yang, Z.L. Type studies on Agarics described by N. Patouillard (and his co-authors) from Vietnam. Mycotaxon 2000, 75, 431–476.
Diversity 2021, 13, 666 39 of 39

66. Chiu, W.F. The Amanitaceae of Yunnan. Sci. Rep. Natl. Tsing Hua Univ. Ser. B Biol. Psychol. Sci. 1948, 3, 165–178.
67. Sysouphanthong, P.; Bouamanivong, S.; Salichan, T. Some Mushrooms in Houayyang Preserves; Biotechnology and Ecology Institute:
Vientiane, Laos, 2018; 87p. (In Lao)
68. Aberdeen, J.E.C. Notes on Lepiota in the Kew Herbarium. Kew Bull. 1962, 16, 129–137. [CrossRef]
69. Grgurinovic, C.A. Larger Fungi of South Australia; Botanic Gardens of Adelaide; State Herbarium; Flora of South Australia
Handbooks Committee: Adelaide, Australia, 1997.
70. Mbaluto, C.; Runo, S.; Wanyoike, W.; Onyango, C.; Kimani, W.; Jagger, H.; Otieno, D.O. Macrolepiota aberdarense, a new edible
mushroom from Kenya. Curr. Res. Environ. Appl. Mycol. J. Fungal. Biol. 2018, 8, 247–253. [CrossRef]
71. Pegler, D.N.; Rayner, R.W. A contribution to the Agaric flora of Kenya. Kew Bull. 1969, 23, 347–412. [CrossRef]
72. Redhead, S.A. Bully for Coprinus—A story of manure, minutiae, and molecules. McIlvainea 2001, 14, 5–14. [CrossRef]
73. Vellinga, E.C.; Yang, Z.L. Volvolepiota and Macrolepiota—Macrolepiota velosa, a new species from China. Mycotaxon 2003, 85,
183–186.
74. Hosen, M.I.; Song, Z.; Gates, G.; Li, T.H. Xanthagaricus caeruleus, a new species with ink-blue lamellae from southeast China.
Mycoscience 2017, 59, 190–194. [CrossRef]
75. Wang, F.; Qi, L.L.; Zhou, X.; Li, Y. A new species and a new record of Xanthagaricus (Agaricaceae, Agaricales) from China.
Phytotaxa 2018, 371, 241–250. [CrossRef]
76. Berkeley, M.J.; Broome, C. The fungi of Ceylon (Hymenomycetes, from Agaricus to Cantharellus). Bot. J. Linn. Soc. 1871, 11,
494–567. [CrossRef]
77. Hussain, S.; Afshan, N.S.; Ahmad, H.; Sher, H.; Khalid, A.N. Xanthagaricus pakistanicus sp. nov. (Agaricaceae), the first report of
the genus from Pakistan. Turk. J. Bot. 2018, 42, 123–133. [CrossRef]