Acta Botanica Brasilica, 2022, 36: e2021abb0382

doi: 10.1590/0102-33062021abb0382

Original article

Lepiota brunneogranulosa (Agaricaceae): a new species from Punjab,

Pakistan, based on integrated taxonomy
Muhammad Asif 1* , Aiman Izhar1 , Muhammad Haqnawaz1 , Abdul Rehman Niazi1 
and Abdul Nasir Khalid1 

Received: December 24, 2021

Accepted: August 29, 2022

ABSTRACT
.

During our surveys of mushrooms in 2019 and 2020, an interesting species of the genus Lepiota was collected from
three different areas of Punjab (Bahawalnagar, Sheikhupura, and Muzaffarghar), Pakistan. These areas vary in their
altitude and somewhat climatic conditions, nevertheless, morpho-anatomical comparisons and molecular analyses
proved that these collections represented the same taxon, described here as Lepiota brunneogranulosa. This new species
is characterized by a pileus with a yellowish-brown central disc, with brown colored granules on a very pale brown
background, dextrinoid, ellipsoid basidiospores, frequently bi-sporic basidia, and narrowly clavate cheilocystidia.
Because of the hymeniform pileus covering, and the placement in the phylogeny, based on Internal Transcribed
Spacer (nrITS) and Larger Subunit (LSU), it belongs to the Lepiota sect. Lilaceae.

Keywords: Bahawalnagar, Lepiotaceous fungi, litter-inhabiting, Phylogeny, taxonomy

Introduction species reported from Pakistan (Ahmad et al. 1997; Kirk et

al. 2008; Kumar & Manimohan 2009; Liang & Yang 2011;
Nawaz et al. 2013; Razaq et al. 2012; 2013; 2014; Qasim
Species of the genus Lepiota (Pers.) Gray (1821: 601) et al. 2015; Qasim et al. 2016; Bashir et al. 2020; Niazi et
are saprobes in nature (Singer 1986; Vellinga 2004) and al. 2021). The genus Lepiota is characterized by its white to
globally distributed from tropical to temperate areas pale, mostly dextrinoid basidiospores that have different
and are infrequent in deserts and arctic-alpine habitats shapes i.e. ellipsoid to spurred, fusiform or penguin-shaped,
(Vellinga 2004) with more than 500 species worldwide various pileus coverings, ranging from hymeniderm, cutis,
(Singer 1986; Kirk et al. 2008; Razaq et al. 2012; Nawaz et trichoderm, epithelium, a regular hymenophoral trama
al. 2013; Qasim et al. 2015; Sysouphanthong et. al 2016; and usually the presence of clamp connections (Candusso
Bashir et al. 2020; Niazi et al. 2021), including more than 30 & Lanzoni 1990; Vellinga 2003).

1 Fungal Biology and Systematics Research Laboratory, Institute of Botany, University of the Punjab, 54590, Lahore, Pakistan

*Corresponding author: [email protected]

 Muhammad Asif, Aiman Izhar, Muhammad Haqnawaz, Abdul Rehman Niazi and Abdul Nasir Khalid

Pileus covering and basidiospore shape are the key

characteristics that divide the genus Lepiota into several
Materials and methods
sections (Vellinga 2001a). Species within the genus Lepiota
show quite close affinities on morphological analysis and Locality description
have further been divided into several sections based on
molecular data. Lepiota species have been supported by The type specimen was collected from district
molecular studies indicating that the genus is polyphyletic, Bahawalnagar, Punjab, Pakistan during the rainy season
and thorough work focused on multigene phylogenetic of 2019. Geographically, District Bahawalnagar is situated
analyses has placed the members of the genus in the relevant in eastern Punjab (Coordinates: 30°33′02″ N, 73°23′26″ E,
sections. (Sysouphanthong et al. 2011). However, there are 163 m) on the bank of Satluj River. The temperature of the
still a lot of species that need to be identified and described. district varies from a minimum of 11 oC to a maximum of
Presently, there are six sections within the genus Lepiota 50 oC and the average annual precipitation is 119 mm
and each section has a unique structure of pileus covering (Ahmed et al. 2014a; 2014b). The second collection was
and basidiospore shape, viz., (1) species with fusiform- collected from district Sheikhupura, in central Punjab
amygdaliform spores and trichodermal pileus covering (Coordinates: 31°42’47” N, 73°58’ 41” E). The climatic
(sect. Lepiota), (2) species with subglobose to ellipsoid or conditions of the district remain moist sub-humid with
spurred spores and hymenidermal pileus covering (sect. annual rainfall from 250 to 500 mm which reaches a
Lilaceae M. Bon), (3) species with ovoid to ellipsoid spores maximum of 635 mm (Nawaz et al. 2017). Another collection
and cylindrical pileus covering elements without clamp- was made from District Muzaffargarh, in south-western
connections (sect. Fuscovinaceae (J. Lange) Kühner), (4) Punjab, (Coordinates: 30°4′10″ N, 71°11′39″ E, 65 m), which
species with ovoid to ellipsoid spores and trichodermal lies in a sub-humid region with 127-150 mm average annual
pileus covering (sect. Ovisporae (J. Lange) Kühner), (5) rainfall (Nickson et al. 2005; Akram et al. 2014; Zamir &
species with spurred spores and trichodermal or cutis pileus Kazmi 2014; Mahmood et al. 2019).
covering (sect. Stenosporae (J. Lange) Kühner) and (6) species
with ovoid to ellipsoid or cylindrical spores with a spurred
Morphological Protocols
base and epithelium pileus covering (elements agglutinated Basidiomata were collected, photographed, and the
in chains) (sect. Echinatae Fay.) (Candusso & Lanzoni 1990; main fleeting features and superficial characters along
Bon 1993; Vellinga 2001c). with habitat and geographical data were noted in the
Lepiota sect. Lilaceae is distinguished by a hymenidermal field. Basidiomata were described using the terminology
pileus covering which is made up of firmly packed clavate to of Vellinga (2001b). Munsell’s soil color chart (Munsell
narrowly clavate elements, and by subglobose to ellipsoid 1975) was used to describe the colors. The collected samples
or spurred basidiospores (Bon 1981; Vellinga 2001a). were brought to the laboratory and dried at 40-50 °C using a
Section Lilaceae is dissimilar from the other five sections fan heater. The dried specimens (Holotype) were deposited
of Lepiota based on morphological and molecular evidence in LAH Herbarium, Institute of Botany, University of the
(Vellinga 2001a; Vellinga 2003). However, some species, Punjab, Lahore, Pakistan, and holotype (=isotype) was
despite producing spurred spores like members of the sect. deposited in Islamabad Herbarium under herbarium voucher
Liliaceae, were placed in some other sections of the genus in no. ISL-F02.
having close phylogenetic relationships with those sections For anatomical analysis, dried tissues were rehydrated in
(Candusso & Lanzoni 1990; Bon 1981; Horak 1981). 5 % KOH, stained with 1 % Congo Red; Melzer’s reagent was
Morphological differences between Lepiota species are used for reactions of the spore walls. The microscopic features
often quite subtle (Vellinga & Huijser 1998). Fortunately, the such as shape and size of basidiospores, cheilocystidia,
separation of different species has become more precise and basidia, structure, and elements of pileus covering and stipe
exact based on nrITS phylogenetic analyses (Vellinga 2003). covering were observed using a light microscope (CXRII,
Lepiotaceous fungi are highly diverse group, especially Labomed Labo America Inc., Fremont, CA, USA) with an
in tropical and subtropical areas, and a huge amount of HDCE-X5 microscopic camera under 40X and oil immersion
work has been done on species from different sections 100X magnification. From each collection, a minimum of 25
of the genus, both based on morphology and phylogeny measurements were taken for each character. Abbreviation:
(Vizzini et al. 2014; Justo et al. 2015; Hosen et al. 2016; Dimension a–b × c–d = minimum and maximum values
Sysouphanthong et al. 2020). They are found in many of length × width, avw = average width, Lm × Wm = mean
localities of Punjab Pakistan. Here, a new species of the spore length × mean spore width, spore quotient Q =
genus Lepiota sect. Lilaceae, collected in three districts length/width, Qm = mean quotient of all basidiospores.
(Bahawalnagar, Sheikhupura, and Muzaffarghar) of Punjab, The notation ‘n/b/p’ indicates ‘n’ basidiospores measured
Pakistan, is presented with a description, illustrations, and from ‘b’ basidiomata from ‘p’ collections (Bas 1969; Yu et
phylogenetic analyses of its nrITS and LSU sequences. al. 2020).

2 Acta Botanica Brasilica, 2022, 36: e2021abb0382

 Lepiota brunneogranulosa (Agaricaceae): a new species from Punjab, Pakistan, based on integrated taxonomy

Molecular Protocols Phylogenetic analyses of LSU dataset

DNA extraction followed the 2 % CTAB protocol (Bruns The fragment size of the large subunit target region was
1995) using dried specimens. DNA was amplified using a 926 bp long. The ITS sequences of Lepiota brunneogranulosa
combination of primers (i.e. ITS1F/ITS4) for ITS and LR0R/ showed 97 % maximum similarity with Lepiota sp.
LR5 for 28S (Vilgalys & Hester 1990; Gardes & Bruns 1993; (OL652983), an unpublished species from Australia. In
White et al. 1990). PCR was done in 25 µl reaction volume this analysis, L. scaberula Vellinga (2001: 290) (MK278271)
following Gardes & Bruns (1993). The PCR products were reported from the USA is closely related to our new species
sequenced by a commercial laboratory and newly generated (Fig. 2).
sequences in this study were deposited in GenBank under
accession numbers nrITS OL331703 - OL331705 and LSU Taxonomy
OL331514 - OL331516.
Lepiota brunneogranulosa M. Asif, A. Izhar, Haqnawaz,
Phylogenetic analyses Niazi & Khalid sp. nov.
Figs. 3A-D, 4A-E
The three obtained sequences from both ITS and LSU
MycoBank No. MB841888
regions were prepared using BioEdit (Hall 1999). The final
Etymology: brunneogranulosa (Latin), referring to the
dataset of ITS sequences was retrieved from GenBank
brown-colored granules on the pileus surface.
based on maximum percentage identity from BLAST
Diagnosis: This species is characterized by brown granules
results and from phylogenetic studies on Lepiota (Hosen
on pileus, stipe turning yellowish on handling, narrowly
et al. 2016; Sysouphanthong et al. 2020) it consisted of
clavate cheilocystidia and hymeniderm pileus covering with
65 ingroup taxa and Macrolepiota dolichaula Pegler & R.W.
obovoid to utriform terminal cells.
Rayner (1969: 365) (JQ928939) as an outgroup. The
Type: Pakistan. Punjab: Haroonabaad, District
LSU final dataset retrieved from GenBank comprised 26 Bahawalnagar, (Coordinates: 30°33′02″ N, 73°23′26″ E;
sequences including Macrolepiota dolichaula (KY418836) 163 m above sea level (a.s.l.)), Scattered or in groups, on
as an outgroup taxon. We used the Basic Local Alignment nutrient-rich loamy soil, July 31, 2019, Muhammad Asif
Search Tool (BLAST) (Altschul et al. 1990) against BWN 23 (Holotype-LAH36803). GenBank: ITS = OL331703;
the GenBank database to find taxa with closely allied 28S = OL331514. (=Isotype ISL-F02).
sequences, which were then retrieved for addition in the Description: Pileus 1.7-2.6 cm, plano-convex at younger
phylogenetic analyses dataset. All the sequences were stage, later become plane at maturity, yellowish-brown
aligned using the Multiple Sequence Alignment search (10YR5/6) central disc with brown (7.5YR4/3) granules
tool (https://www.ebi.ac.uk/Tools/msa/muscle/) (Edgar on very pale brown (10YR8/4) surface, denser around the
2004) and then adjusted manually in BioEdit (Hall 1999). central disc and becoming sparse towards margins, showing
Final alignment was deposited in TreeBASE (http:// the pale brown surface; dull and dry; margin undulating
www.treebase.org/) under the accession numbers http:// and dentate. Lamellae free, sub-distant, broad, light
purl.org/phylo/treebase/phylows/study/TB2:S28978 brown (6YR7/1), wavy with eroded edges, a single tier of
and http://purl.org/phylo/treebase/phylows/study/ lamellulae, concolorous. Stipe 2.8-3.5 × 0.2-0.5 cm light
TB2:S28980. Maximum Likelihood (ML) analysis was brown (6YR7/1), becoming yellowish upon handling, central,
done using RAxML-HPC2 v. 8.2.12 (Stamatakis 2014) equal, with shiny surface, concolorous fibrils at the apex of
as applied on the CIPRES portal v 3.1 (Miller et al. 2010) stipe. Annulus sub-peronate, white (2.5YR5/12), single-
and with 1000 rapid bootstrap iterations for both genes. edged, upturned, attached at the upper part of the stipe,
disappearing at maturity. Volva absent. Smell acrid. Taste
Results mild
Basidiospores (100/4/4) (4.5-) 4.6-5.2 × 3.2-3.8 (-3.9)
µm, avl × avw = 4.9 × 3.5 µm, Q = 1.4 - 1.7, Qm = 1.5, thin-
Phylogenetic analyses of ITS dataset walled, pale yellow in KOH, slightly brownish in Melzer’s
reagent, but not so strong, ellipsoid, oblong in side view,
The fragment size of the target region was 660 bp rounded triangular in frontal view, non-guttulate, smooth
long. In BLAST results, the ITS sequences of Lepiota with a prominent apiculus. Basidia 18-22 × 6.8-9 µm, avl
brunneogranulosa showed 86 % maximum similarity with × avw = 20.19 × 7.69 µm, Q = 2.62, thick-walled, mostly
Lepiota sp. (AF391052) from the USA. In this analysis, L. clavate, rarely narrowly clavate, frequently bisporic, rarely
brunneogranulosa gets clustered within the L. sect. Lilaceae tri and tetra sporic, hyaline in KOH, non-guttulate. Lamella
close to L. lilacea (1892: 3) (GQ203822 & AY176379) edge sterile, cheilocystidia crowded. Cheilocystidia 17-23
reported from California, USA and L. bengalensis (2016: × 6-7.5 µm, avl × avw = 19.6 × 7.02 µm, Q = 2.8, thick-
188) (KU563149 & KU563148) reported from Bangladesh walled, narrowly clavate, hyaline in KOH, non-guttulate.
(Fig. 1). Pleurocystidia absent. Pileus covering hymeniderm,

Acta Botanica Brasilica, 2022, 36: e2021abb0382 3

 Muhammad Asif, Aiman Izhar, Muhammad Haqnawaz, Abdul Rehman Niazi and Abdul Nasir Khalid

Figure 1. Molecular phylogenetic placement of Lepiota brunneogranulosa based on maximum likelihood (ML) method of ITS sequences.

4 Acta Botanica Brasilica, 2022, 36: e2021abb0382

 Lepiota brunneogranulosa (Agaricaceae): a new species from Punjab, Pakistan, based on integrated taxonomy

Figure 2. Molecular phylogenetic placement of Lepiota brunneogranulosa based on maximum likelihood (ML) method of LSU sequences.

thick-walled, hyaline in KOH, septate, irregularly arranged m a.s.l) scattered on loamy soil, September 20, 2019, Aiman
with obovoid, clavate to broadly clavate or utriform with Izhar SKP 1211 (LAH36974). Pakistan-Punjab: Easan Wala
median constriction terminal cells 25-47 × 17-27 µm, hyphae Forest, District Muzaffargarh, (Coordinates: 30°4′10″N,
8.7-6.3 µm, avl × avw = 19.3 × 11.5 µm. Stipitipellis a 71°11′39″E; 123 m a.s.l) August 29, 2020, Muhammad
cutis, hyphae 6.6 – 13.2 µm, avw = 8.4 µm, thick-walled, Haqnawaz EF 02, (LAH36954). GenBank: ITS = OL331705;
hyaline in KOH, septate, parallel arrangement, long and 28S = OL331515.
narrow. Caulocystidia absent. Clamp connections are

Discussion
present in all tissues.
Habitat and Ecology: Scattered or in groups, on nutrient-
rich loamy soil, under Eucalyptus camaldulensis Dehnh.,
Aug to Sep. Lepiota brunneogranulosa is characterized by pileus with
Geographical distribution range: Known only from Punjab, brown granules on a very pale brown background, presence of
Pakistan. annulus on stipe, becoming yellowish on handling, narrowly
Additional specimens examined: PAKISTAN. Punjab: clavate cheilocystidia, ellipsoid to broadly basidiospores,
Haroonabad, District Bahawalnagar, at 163 m a.s.l, September and obvoid, clavate to broadly clavate or utriform terminal
07, 2020, Muhammad Asif BWN 142 (LAH36804). GenBank: elements in pileus covering (Tab. 1).
ITS = OL331704; 28S = OL331516. Pakistan-Punjab: District Lepiota lilacea (GQ203822 & AY176379) and L.
Sheikhupura, (Coordinates: 31° 71′67″N, 73°98 50″E; 236 bengalensis (KU563149 & KU563148) are found to be

Acta Botanica Brasilica, 2022, 36: e2021abb0382 5

 Muhammad Asif, Aiman Izhar, Muhammad Haqnawaz, Abdul Rehman Niazi and Abdul Nasir Khalid

close relatives of our newly described taxon in ITS based Lepiota bengalensis reported from Bangladesh differs
phylogenetic analysis (Fig. 1). But both species can be from L. brunneogranulosa in having pastel red to brownish
differentiated from our new species morpho-anatomically. red, conico-convex to hemispherical pileus with the dark
Lepiota lilacea is widely reported around temperate areas. ochraceous or reddish-brown central disc, and brownish red
It was originally described and widely reported in Europe to reddish-orange appressed or radially arranged squamules
and is also present in North America. It differs from L. on the pileus and no color change on touching (Hosen et
brunneogranulosa macroscopically in having plano-convex to al. 2016). Clamp connections are only present in pileus
applanate pileus with dark violet central disc that has radially covering (Tab. 1) (Hosen et al. 2016).
arranged scales, white lamellae with free margins, and In LSU based phylogenetic analysis (Fig. 2), Lepiota
microscopically in having ellipsoid to oblong basidiospores scaberula, a species originally reported from the California,
with 5.2 × 3.1 µm average size (Tab. 1) (Vellinga 2001ab; USA, is the closest relative to L. brunneogranulosa; it differs
Vellinga et al. 2011). from latter by its scurfy pileus surface with fibrillose v-shaped

Figure 3. Basidiomata of Lepiota brunneogranulosa (LAH36803, Holotype) Photos by Muhammad Asif & Aiman Izhar.

6 Acta Botanica Brasilica, 2022, 36: e2021abb0382

 Lepiota brunneogranulosa (Agaricaceae): a new species from Punjab, Pakistan, based on integrated taxonomy

Figure 4. Microscopic characters of Lepiota brunneogranulosa (LAH36803, Holotype) A. Basidia, B. Basidiospores,

C. Cheilocystidia, D. Pileus covering, E. Stipitipellis, Bars: A-E = 10 µm. Drawings by Aiman Izhar.

Acta Botanica Brasilica, 2022, 36: e2021abb0382 7

 Muhammad Asif, Aiman Izhar, Muhammad Haqnawaz, Abdul Rehman Niazi and Abdul Nasir Khalid

Table 1. Comparison of the diagnostic characteristics of L. brunneogranulosa with closely related species (Vellinga 2001b; 2001d;
Hosen et al. 2016; Orton 1960).
Characters L. brunneogranulosa L. lilaceae L. bengalensis L. scaberula L. ochraceofulva
Pastel red to brownish
Paler around centre
red, campanulate
and whistish
Violet, plano-convex to to conicoconvex, Orange brown to
Pileus Plane, very pale brown at margins,
applanate, expanding to leather brown
hemispherical to
hemispherical to
plano-convex
applanate with age

Dark ochraceous or Cream to yellow-

Central disc Yellowish-brown Dark violet Reddish brown
reddish brown brownish

Brownish red to
Violet, radially arranged reddish orange, Irregularly fibrillose Pileus broken
Squamules Brown granules
appressed scales appressed or radially v-shaped squamules into squamules
arranged squamules

Distant to sub-
crowded, pale yellow Moderately crowded
Sub-distant, light
Lamellae Crowded, white to yellowish white to moderately Crowded
brown
or light ochraceous- distant
cream

Ellipsoid to broadly
Ellipsoid to oblong, Ellipsoid to cylindrical, Ellipsoid to oblong, Broadly ellipsoid,
Basidiospores ellipsoid or oblong,
av = 5.2 × 3.1 µm av = 5.6 × 3 µm av = 5.7 × 3.6 µm av = 5.5 × 3.7 µm
av = 4.9 × 3.5 µm,

Cylindrical and
Narrowly cylindrical, Fasciculate, clavate to
Cheilocystidia Narrowly clavate slightly widened at Clavate
narrowly utriform narrowly clavate
apex

Hymeniderm,
Hymeniform,
irregularly arranged Hymeniderm made up of Cylindrical to
composed of clavate
Pileus covering with obvoid, clavate clavate to broadly clavate slightly inflated Hymeniform
to broadly clavate or
to broadly clavate or cells hyphae
narrowly clavate cells
utriform terminal cells

Gregarious in small
Scattered or in groups, Solitary, caespitose Gregarious,
groups, on irrigated soil, On loamy soil,
on nutrient-rich loamy or gregarious in small saprotrophic on
Habitat on rich nitrophilous in gardens and
soil under Eucalyptus groups on debris of soil in Sequoia
shrubs in woods under parks
camaldulensis Albizia and Eucalyptus sempervirens forests
Sequoia sempervirens

squammules, moderately crowded and pinkish lamellae, Acknowledgments

whitish-wooly stipe and non-dextrinoid basidiospores
(Tab. 1) (Vellinga 2001d).
We are sincerely thankful to Dr. Else C. Vellinga (University
Lepiota ochraceofulva, which is originally reported from
of California, Berkeley, USA) for her critical review and
Europe, differs from L. brunneogranulosa by its crowded
valuable comments and suggestions on an earlier version
lamellae, long and broad stipe 52-70 × 6-9 mm with an of this manuscript which helped us to improve the article.
almost clavate bulbous base, tinged ochraceous-cream color We thank all the anonymous reviewers for their corrections
at stipe apex, and slightly wider basidiospores 5.5-7 × 3.5- and suggestions to improve this paper.
4.5 µm (Tab. 1) (Orton 1960).
Previously, 32 species of Lepiota have been reported from
Pakistan (Ahmad et al. 1997; Razaq et al. 2012; Nawaz et al. References
2013; Qasim et al. 2015; 2016; Bashir et al. 2020; Niazi et al.
Ahmad S, Iqbal S, Khalid AN. 1997. Fungi of Pakistan. Lahore, Sultan
2021; Haqnawaz et al. 2022). Our current study, based on
Ahmad Mycological Society of Pakistan.
morphological and molecular data of the species collected Ahmed N, Mahmood A, Tahir SS, et al. 2014a. Ethno-medicinal knowledge
from Punjab, Pakistan, indicated that this is a genuine and relative importance of indigenous medicinal plants of Cholistan
member of the genus Lepiota and also a new taxon to science. desert Punjab Province. Pakistan Journal of Ethno-pharmacology
155: 1263-1275.
So, future mycological expeditions are appreciated to explore Ahmed N, Mahmood A, Tahir SS, et al. 2014b. Relative importance of
more hidden species of the genus, their variabilities, and indigenous medicinal plants from Layyah district, Punjab Province.
ecology from Pakistan. Pakistan Journal of Ethno-pharmacology 155: 509-523.

8 Acta Botanica Brasilica, 2022, 36: e2021abb0382

 Lepiota brunneogranulosa (Agaricaceae): a new species from Punjab, Pakistan, based on integrated taxonomy

Akram Z, Hussain S, Mansoor M, et al. 2014. Soil fertility and salinity Orton PD. 1960. New check list of British Agarics and Boleti: part III.
status of Muzaffargarh District, Punjab Pakistan. Universal Journal Notes on genera and species in the list. Transactions of the British
of Agriculture Research 2: 242-249. Mycological Society 43: 159-384.
Altschul SF, Gish W, Miller W, et al. 1990. Basic local alignment search Pegler DN, Rayner RW. 1969. A contribution to the agaric fora of Kenya.
tool. Journal of Molecular Biology 215: 403-410. Kew Bulletin 23: 347-412
Bas C. 1969. Morphology and subdivision of Amanita and a monograph Qasim T, Khalid AN, Vellinga EC, et al. 2015. Lepiota albogranulosa sp.
of its section Lepidella. Persoonia 5: 96-97. nov. (Agaricales, Agaricaceae) from Lahore, Pakistan. Mycological
Bashir H, Usman M, Khalid AN. 2020. Lepiota cholistanensis a new species of Progress 14: 24.
Lepiota (Agaricaceae: Basidiomycota) from Cholistan desert, Pakistan. Qasim T, Khalid AN, Vellinga EC. 2016. A new species of Lepiota, Lepiota
Phytotaxa 455: 267-276. lahorensis, from Lahore, Pakistan. Turkish Journal of Botany 40: 419-426.
Bon M. 1981. Clé monographique des “Lépiotes” d’Europe. Documents Razaq A, Khalid AN, Ilyas S. 2013. Molecular Identification of Lepiota
Mycologiques 11: 1-77. acutesquamosa and L. cristata (Basidiomycota, Agaricales) based on
Bon M. 1993. Flore mycologique d’Europe 3. Les Lépiotes. Documents ITS-rDNA barcoding from Himalayan Moist Temperate Forests of
Mycologiques Mémoire hors série 3: 1-153. Pakistan. International Journal of Agriculture & Biology 15: 313-318.
Bruns TD. 1995. Thoughts on the processes that maintain local species Razaq A, Khalid AN, Vellinga EC. 2012. Lepiota himalayensis (Basidiomycota,
diversity of ectomycorrhizal fungi. Plant and Soil 170: 63-73. Agaricales), a new species from Pakistan. Mycotaxon 121: 319-325.
Candusso M, Lanzoni G. 1990. Fungi Europaei 4: Lepiota s.l. Alassio, Razaq A, Vellinga EC, Ilyas S, et al. 2014. Lepiota brunneoincarnata and L.
Giovanna Biella. subincarnata: distribution and phylogeny. Mycotaxon 126: 133-141.
Edgar RC. 2004. MUSCLE: multiple sequence alignment with high accuracy Singer R. 1986. The Agaricales in modern taxonomy. 4th edn. Koenigstein,
and high throughput. Nucleic Acids Research 32: 1792-1797. Koeltz Scientific Books.
Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for Stamatakis A. 2014. RAxML version 8: a tool for phylogenetic analysis and
Basidiomycetes-Application to the identification of mycorrhizae and post–analysis of large phylogenies. Bioinformatics 30: 1312-1313.
rusts. Molecular Ecology 2: 113118. Sysouphanthong P, Guo JY, Hyde K, et al. 2016. Lepiota thailandica
Gray SF. 1821. A Natural Arrangement of British Plants. Vol. II According to (Agaricaceae) a new species from Thailand. Phytotaxa 245: 262-270.
Their Relations to Each Other as Pointed Out by Jussieu, De Candolle, Sysouphanthong P, Hyde KD, Chukeatirote E, et al. 2011. A review of genus
Brown, et al.. Baldwin,Cradock, and Joy. pp. 601. Lepiota and its distribution in Asia. Current Research in Environmental
Hall TA. 1999. BioEdit: a user-friendly biological sequence alignment & Applied Mycology 1: 161-176.
editor and analysis program for Windows 95/98/NT/7. Nucleic Acids Sysouphanthong P, Thongklang N, Karunarathna SC, et al. 2020. Lepiota
Symposium Series 41: 95-98. condylospora, a new species with nodulose spores in section Lilaceae
Haqnawaz M, Niazi AR, Usman M, et al. 2022. Lepiota sindhudeltana sp. from northern Thailand. Phytotaxa 455: 61-69.
nov. (Agaricaceae; Basidiomycota) from Punjab, Pakistan. Phytotaxa Vellinga EC, Huijser HA. 1998. Studies in Lepiota I. Species with a
550: 253-262. hymeniform pileus covering. Belgian Journal of Botany 131: 191-210.
Horak E. 1981. On Australasian species of Lepiota S.F. Gray (Agaricales) Vellinga EC, Sysouphanthong P, Hyde KD. 2011. The family Agaricaceae:
with spurred spores. Sydowia 33: 111-144. phylogenies and two new white-spored genera. Mycologia, 103: 494-509.
Hosen MI, Li TH, Ge ZW, et al. 2016. Lepiota bengalensis, a new species Vellinga EC. 2001a. Lepiota. In: Noordeloos ME, Kuyper TW, Vellinga
with first generic report for Bangladesh. Sydowia 68: 187-192. EC (eds.).Flora Agaricina Neerlandica 5. Swets & Zeitlinger Lisse.
Justo A, Angelini C, Bizzi A. 2015. Two new species and a new record of pp. 109-151.
Lepiota (Basidiomycota, Agaricales) from the Dominican Republic. Vellinga EC. 2001b. Studies in Lepiota IV. Lepiota cristata and L. castaneidisca.
Mycological Progress 14: 56. Mycotaxon 80: 297-306.
Kirk PM, Cannon PF, Minter DW, et al. 2008. Ainsworth and Bisby’s Vellinga EC. 2001c. Agaricaceae. In: Noordeloos ME, Kuyper TW, Vellinga EC
Dictionary of the fungi, Wallingford, International Mycological (eds.). Flora Agaricina Neerlandica 5. Rotterdam, Balkema Publishers,
Institute. pp. 220.
Kumar TA, Manimohan P. 2009. The genus Lepiota (Agaricales, Vellinga EC. 2001d. Studies in Lepiota III. Some species from California,
Basidiomycota) in Kerala State, India. Mycotaxon 107: 105-138. USA. Mycotaxon 80: 285-295.
Liang JF, Yang ZL. 2011. A new species of Lepiota (Agaricaceae) from south Vellinga EC. 2003. Phylogeny of Lepiota (Agaricaceae) evidence from nrITS
western China. Mycotaxon 117: 359-363. and nrLSU sequences. Mycological Progress 2: 305-322.
Mahmood S, Rahman AU, Sajjad A. 2019. Assessment of 2010 flood Vellinga EC. 2004. Ecology and distribution of lepiotaceous fungi
disaster causes and damages in district Muzaffargarh, Central Indus (Agaricaceae) - a review. Nova Hedwigia 78: 273-299.
Basin, Pakistan. Environmental Earth Sciences 78: 1-11. Vilgalys R, Hester M. 1990. Rapid genetic identification and mapping of
Miller MA, Pfeiffer W, Schwartz T. 2010. Creating the CIPRES Science enzymatically amplified ribosomal DNA from several Cryptococcus
Gateway for Inference of Large Phylogenetic Trees. Gateway Computing species. Journal of Bacteriology 172: 4239-4246.
Environments Workshop: 1 – 8. Vizzini A, Liang JF, Jancovicová S. 2014. Lepiota coloratipes, a new species
Munsell A. 1975. Munsell soil color charts. Macbeth Division of Kollmorgen for Lepiota rufipesss. auct. europ. non ss. orig. Mycological Progress
Instruments Corporation, New York. 13: 171-179.
Nawaz M, Wahla AJ, Kashif MS, et al. 2017. Effects of exogenous nitrogen White TJ, Bruns T, Lee S, et al. 1990. Amplification and direct sequencing
levels on the yield of rice grain in Sheikhupura, Pakistan. Pakistan of fungal ribosomal RNA genes for phylogenetics. In: Innis MA,
Journal of Agricultural Research 30: 85-92. Gelfand DH, Sninsky JJ, White TJ (eds.) PCR protocols: a guide to
Nawaz R, Khalid AN, Hanif M, et al. 2013. Lepiota vellingana sp. nov. methods and applications. New York, Academic Press. pp. 315-322.
(Basidiomycota, Agaricales) a new species from Lahore, Pakistan. Yu WJ, Chang C, Qin LW, Zeng NK, Wang SX, Fan YG. 2020. Pseudosperma
Mycological progress 12: 727-732. citrinostipes (Inocybaceae), a new species associated with Keteleeria
Niazi AR, Asif M, Izhar A, et al. 2021. A new species of Lepiota (Agaricaceae) from southwestern China. Phytotaxa 450: 8-16.
from Punjab, Pakistan. Phytotaxa 511: 163-174. Zamir UB, Kazmi JH. 2014. Automated method for delineating watershed,
Nickson RT, McArthur JM, Shrestha B, et al. 2005. Arsenic and other drainage pattern and calculation of flow accumulation in Punjab
drinking water quality issues, Muzaffargarh District, Pakistan. Applied province using digital elevation model. Pakistan Journal of Scientific
geochemistry 20: 55-68. & Industrial Research Series A: Physical Sciences 58: 90-98.

Acta Botanica Brasilica, 2022, 36: e2021abb0382 9