WEBBIA
Journal of Plant
Taxonomy
and Geography
Firenze University Press
www.fupress.com/webbia
A taxonomic and phylogenetic study of some
Lecidella species from Pakistan
Citation: Rizwana Zulfiqar, Kamran
Habib, Memoona Khan, Abdul Nasir
Khalid (2020) A taxonomic and phylogenetic study of some Lecidella species
from Pakistan. Webbia. Journal of
Plant Taxonomy and Geography 75(2):
219-230. doi: 10.36253/jopt-9581
Received: August 8, 2020
Rizwana Zulfiqar*, Kamran Habib, Memoona Khan, Abdul Nasir
Khalid
Fungal Biology and Systematics Lab, Department of Botany, University of the Punjab,
Quaid-e-Azam Campus, Lahore, 54590, Pakistan
*Corresponding author. Email: rizwanamughal6@gmail.com
Accepted: September 9, 2020
Published: November 18, 2020
Copyright: © 2020 Rizwana Zulfiqar, Kamran Habib, Memoona Khan, Abdul
Nasir Khalid. This is an open access,
peer-reviewed article published by
Firenze University Press (http://www.
fupress.com/webbia) and distributed
under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and
reproduction in any medium, provided
the original author and source are
credited.
Data Availability Statement: All relevant data are within the paper and its
Supporting Information files.
Competing Interests: The Author(s)
declare(s) no conflict of interest.
Editor: Riccardo M. Baldini
Abstract. In this study, a number of species of the genus Lecidella were collected from
different areas of Pakistan and characterized using morpho-anatomical and molecular
techniques. The present work revealed that collected specimens belong to four species
of Lecidella. Among which L. tumidula is a new record for Pakistan while L. carpathica, L. patavina and L. stigmatea are being reported here from new localities, from Pakistan, representing their wider distribution. Complete morpho-anatomical descriptions,
ecology and distribution, along with ITS-based molecular analysis is provided.
Keywords: Khyber Pakhtunkhwa, Lecanoraceae, Lecidella, phylogeny, Pakistan.
INTRODUCTION
The genus Lecidella Korb. (Lecanoraceae), established by Korber in 1855,
is comprised of 80 species (Zhao et al. 2015). This genus of crustose lichens
is mainly characterized by black lecidiene apothecia with persistent proper
excipulum and Lecidella-type asci (Zhao et al. 2015). It is a cosmopolitan
genus, frequently found in temperate latitudes on wood, bark or rock (Kantvilas and Elix 2013). This group of lichenized ascomycetous fungi is usually
regarded as taxonomically difficult due to a high degree of variation in morphological characters. In such cases, molecular study has played a significant
role in systematics and species delimitation (Zhao et al. 2015).
Pakistan is located in western South Asia between 24–37o N latitude and
62–75o E longitudes. The country is well known for its geographical and climatic variations which is linked with rich biodiversity (IUCN 2006). The
lichen diversity in this region is probably very high but little known due to
the lack of surveys in many areas (Ahmad et al. 1997; Aptroot and Iqbal
2012). So far, 375 lichen species have been reported from Pakistan (Ahmad
1965; Aptroot and Iqbal 2012; Habib et al. 2017; Khan et al. 2018; Habib and
Khalid 2019). In the past, attempts were made to describe the lichen diversity
of the country using morpho-anatomical techniques (Ahmad 1965; Iqbal et
al. 1978; Aptroot and Iqbal 2012). Recently, molecular techniques have also
Webbia. Journal of Plant Taxonomy and Geography 75(2): 219-230, 2020
ISSN 0083-7792 (print) | ISSN 2169-4060 (online) | DOI: 10.36253/jopt-9581
220
Rizwana Zulfiqar, Kamran Habib, Memoona Khan, Abdul Nasir Khalid
been used to study lichen flora of Pakistan (Habib et al.
2017; Khan et al. 2018; Habib and Khalid 2019).
From Pakistan, five species of Lecidella have been
reported so far, viz; L. carpathica Korb., L. euphorea
(Florke) Hertel, L. pulveraceae (Florke) Th.Fr., L. patavina (A.Massal.) Knoph & Leuckert, and L. stigmatea
(Ach.) Hertel & Leuckert (Aptroot and Iqbal 2012). In
the present study, different species of Lecidella have
been collected from different areas of Pakistan. Use of
morpho-anatomical techniques along with phylogenetic
analysis led to identification of four different species. L.
tumidula (A.Massal.) Knoph & Leuckert has been collected and described for the first time from Pakistan
which made an addition to the lichen flora of this country. Now, the number of Lecidella species reported from
Pakistan has been raised from five to six.
MATERIALS AND METHODS
used to reassemble forward and reverse sequences (Hall
2005). Sequences of other Lecidella species based on initial BLAST searches and those used in a study on phylogeny of Lecidella by Zhao et al. (2015) were used in
phylogenetic analysis (Table 1). The multiple sequence
alignment was performed using MAFFT v7 with all
parameters set to default values (Katoh and Standley
2013). The ends of the alignment were trimmed to nearly
an equal number of sites for all sequences. All gaps were
treated as missing data. Maximum Likelihood analysis was performed with MEGA6 using a GTR model for
bootstrapping (Tamura et al. 2013). One thousand rapid
bootstrap replicates were run to infer the evolutionary
history of each species.
The length of the final aligned file was 541 nucleotides, of which 355 sites were conserved, 181 were variable,
156 were parsimony informative and 25 were singleton,
Rhizoplaca porterii and R. parilis (HM577376, HM577309)
were chosen as an outgroup (Zhao et al. 2015).
Morphological and chemical studies
Collections were made during a lichen survey of
Chikar, Muzaffarabad (Azad Jammu and Kashmir),
Parachinar, and Fairy Meadows (Gilgit Baltistan) in
2017 and 2018. Morphological characters were observed
under a stereomicroscope (Meiji Techno, EMZ-5TR,
Japan). Standard microscopy and spot tests (Hale 1979)
were used for identification. Measurements were made
from free hand section of apothecia mounted in water
on a glass slide. The sections were observed using a compound microscope (MX4300H, Meiji Techno Co., Ltd.,
Japan). Minimum twenty measurements in water were
made for each diagnostic feature.
DNA extraction and PCR amplification
We used thallus material along with apothecial
material to extract fungal DNA using a 2% CTAB protocol (Gardes and Bruns 1993). The primer pair ITS1F
(Grades and Bruns 1993) and ITS4 (White et al. 1990)
was used to amplify the internal transcribed spacer
(ITS) region under PCR conditions used by Khan et
al. (2018). PCR products were visualized in a 1 % agarose gel using ethidium bromide (Sambrook and Russell
2001). PCR products were sequenced from BGI, China.
Phylogenetic analysis
The ITS regions of all specimens were amplified
and sequenced. Bio-edit sequence alignment editor was
RESULTS
Lecidella tumidula (A.Massal.) Knoph & Leuckert,
Biblthca Lichenol. 68: 131. 1997. (Figure 1, A-E; Figure 5;
A-D).
Description
Thallus crustose, continuous, up to 3 cm in diameter, up to 0.6 mm thick, granulose to rimose. Colour:
light greyish green to light olive green, dull to weakly
glossy. Apothecia: rounded, semi-immersed, sessile, 0.5–
1 mm, frequently present, strongly constricted at base.
Margins: thin, continuous, concolorous to disc, distinct when young, indistinct when mature. Disc: black,
smooth, glossy, pruinose, flat to convex. Exciple: black,
40–50 µm thick. Epihymenium: blackish, 30–36 µm tall.
Hymenium: hyaline, 80–86 µm tall. Hypothecium: dark
reddish brown, 30–40 µm tall. Paraphyses: aseptate, hyaline, rarely anastomosing or branched, apex swollen, 2–4
µm wide. Asci: clavate, 8-spored, 58–78 µm × 10–14 µm.
Ascospores: simple, hyaline, narrowly ellipsoid to ovoid,
10–12 µm × 5–7 µm.
Spot Tests
All negative (diploicin and lichexanthone reported
according to literature but not examined in our specimen).
Substrate and ecology
L. tumidula was found on bark of Quercus incana W.
Bartram in dry temperate forest, at an altitude of 1,705
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A taxonomic and phylogenetic study of some Lecidella species from Pakistan
Table 1. Specimens used in phylogenetic analysis of Lecidella species.
ITS Accession
NoKT453736
KT453737
HQ650596
KT453750
KT453746
KT453748
KT453747
AF517929
KT453743
KT453742
KX132994
KX132965
JN873903
MN387031
MN387029
HQ287871
JN873884
Specimen
Name
Lecidella
tumidula
Lecidella
tumidula
Lecidella
tumidula
Lecidella
elaeochromoid
Lecidella
elaeochromoid
Lecidella
effugiens
Lecidella
effugiens
Lecidella
meiococca
Lecidella
euphorea
Lecidella
euphorea
Lecidella
flavosorediata
Lecidella
flavosorediata
Lecidella
wulfenii
Lecidella
elaeochroma
Lecidella
elaeochroma
Lecidella
greenii
Lecidella
greenii
Country
China
China
USA
China
China
China
China
Sweden
China
China
Switzerland
Switzerland
Austria
Poland
Poland
Antarctica
Antarctica
Voucher NoZX XL0009
(Zhao et al. 2015)
ZX 20129166-2
(Zhao et al. 2015)
404720
(Schmull et al. 2011)
ZX 20141142
(Zhao et al. 2015)
ZX 20114966-2
(Zhao et al. 2015)
ZX 20141269-2
(Zhao et al. 2015)
ZX 20141148-2
(Zhao et al. 2015)
Ekman 3101 (BG)
(Ekman & Tonsberg, 2002)
ZX XL0387
(Zhao et al. 2015)
ZX 20140638
(Zhao et al. 2015)
980812
(Mark et al. 2016)
980812
(Mark et al. 2016)
Tuerk 39666
(Ruprecht et al. 2012)
272987
(Singh et al. 2019)
272987
(Singh et al. 2019)
Herbarium Tuerk 43015
(Lumbsch et al. 2011)
Tuerk 33612
(Ruprecht et al. 2012)
m, dominant vegetation including Pinus gerardiana Wall.
ex D. Don, Quercus ilex L., Juniperus macropoda Boiss.
and Picea smithiana Boiss, temperature ranges between
6–16 oC, average annual rainfall 300–500 mm.
Distribution
Temperate areas of Asia including China (Zhao et al.
2015), have also been reported from Europe and North
America (Nash et al. 2004). Here it is been reported for
the first time from Pakistan.
Material examined
PAKISTAN: Khyber Pakhtunkhwa Province: Kurram District, Parachinar: 33°90’ N, 70°08’ E; 1,705 m;
on bark of Quercus incana W. Bartram, 18 August 2018,
ITS Accession
NoMK970672
HQ605934
KT453767
MK620163
MK620136
KT453764
KT453765
KT453760
KT453763
KT695322
KT695353
Specimen
Name
Lecidella
greenii
Lecidella
patavina
Lecidella
patavina
Lecidella
stigmatea
Lecidella
stigmatea
Lecidella
stigmatea
Lecidella
stigmatea
Lecidella
stigmatea
Lecidella
stigmatea
Lecidella
carpathica
Lecidella
carpathica
Country
Antarctica
Turkey
China
Argentina
Argentina
China
China
China
China
Canada
Canada
JN873899
Lecidella siplei Antarctica
JN873896
Lecidella siplei Antarctica
JN873898
Lecidella siplei Antarctica
JN873897
Lecidella siplei Antarctica
HM577376
HM577309
Rhizoplaca
porterii
Rhizoplaca
parilis
USA
USA
Voucher NoT48787
(Wagner et al. 2019)
574971
(Basaran et al. 2014)
ZX 20140501-2
(Zhao et al. 2015)
UR00128
(Ruprecht et al. 2019)
UR00080
(Ruprecht et al. 2019)
ZX 20140086-2
(Zhao et al. 2015)
ZX 20140045-2
(Zhao et al. 2015)
ZX 20140519-2
(Zhao et al. 2015)
ZX 20140507-2
(Zhao et al. 2015)
BIOUG24047-H06
(Telfer et al. 2015)
BIOUG24047-E02
(Telfer et al. 2015)
Tuerk 35895
(Ruprecht et al. 2012)
Tuerk 32991
(Ruprecht et al. 2012)
Tuerk 33457
(Ruprecht et al. 2012)
Tuerk 33449
(Ruprecht et al. 2012)
55145 (BRY-C)
(Leavitt et al. 2011)
55078 (BRY-C)
(Leavitt et al. 2011)
A.N. Khalid & K. Habib; LAH36399; (GenBank accession no. MT139651).
Lecidella carpathica Korb., Parerga lichenol. (Breslau) 3:
212. 1861. (Figure 2, A-F; Figure 5; E-H).
Description
Thallus crustose, effuse, rimose areolate to subsquamulose, dull, up to 5 cm wide, up to 300 µm thick.
Areoles: scattered to crowded, smooth or rough, up to
0.5 mm in diameter, irregular in outline. Colour: pale
green to grey green. Apothecia: sessile, semi-immersed,
constricted at base, scattered to crowded, up to 1 mm
in diameter. Disc: black, plane, flat to moderately con-
222
Rizwana Zulfiqar, Kamran Habib, Memoona Khan, Abdul Nasir Khalid
Figure 1. Lecidella tumidula. (A & B) showing crustose thallus and abundant apothecia (C) pruinose apothecia; (D) Cross section of an
apothecium (H: Hymenium; E: Epihymenium); (E) showing ascus and paraphyses (A: Ascus; P: Paraphyses).
vex, pruinose. Margins: distinct, entire or flexuose, up to
0.9 mm wide. Exciple: bluish green to black, 20–30 µm
thick. Epihymenium: pale brown to dark brown, 15–20
µm tall. Hymenium: hyaline, 65–90 µm tall. Hypothecium: pale brown to brown, 30–45 µm tall. Paraphyses: aseptate, hyaline, rarely anastomosing or branched,
sometimes slightly swollen apically, up to 2 µm wide.
Asci: clavate, 45–60 x 13–15 µm, 8-spored. Ascospores:
simple, hyaline, ellipsoid to ovoid, 10–14 x 6–8 µm.
Spot tests
K+ yellow, C–, KC–, P–
Distribution
Widespread including Africa, Europe, Australia,
Macaronesia, New Zealand (Smith et al. 2009), UK,
USA, Ukraine (Oxner 1968; Kondratyuk et al. 1998;
2003), temperate parts of Asia (Nash et al. 2004), China
(Zhao et al. 2015), India (Singh & Sinha 2010) and Pakistan (Nasim et al. 2004).
Material examined
PAKISTAN: Azad Jammu & Kashmir: Chikar,
34° 9’ N, 73° 41’ E, 234 m; on rock; 18 August 2018, K.
Habib; LAH36400; (Genbank accession no. MT139649).
Substrate and ecology
L. carpathica was found on rock (saxicolous), in
moist temperate forest at an altitude of 234 m, temperature ranges between -2–37 oC, average annual rainfall
1,500–1,600 mm, with dominant tree species Pinus wallichiana A.B. Jacks., Picea smithiana Boiss., Abies pindrow Royle., Quercus incana W. Bartram and Q. dilatata
Lindl. ex Royle.
Lecidella patavina (A.Massal.) Knoph & Leuckert, in
Knoph, Bibliotheca Lichenol. 36: 116. 1990. (Figure 3,
A-G; Figure 6; A-D).
Description
Thallus indistinctly crustose, inconspicuous, intermingled with rock particles, up to 4 cm in diameter.
A taxonomic and phylogenetic study of some Lecidella species from Pakistan
223
Figure 2. Lecidella carpathica. (A & B) Crustose thallus showing abundant apothecia; (C) black, pruinose apothecia; (D) Cross section of an
apothecium (H: Hymenium; E: Epihymenium); (E) Ascus; (F) Paraphyses.
224
Rizwana Zulfiqar, Kamran Habib, Memoona Khan, Abdul Nasir Khalid
Figure 3. Lecidella patavina. (A & B) Crustose thallus showing abundant apothecia; (C) black, pruinose fascicle of apothecia; (D) Cross section of an apothecium (H: Hymenium; E: Epihymenium); (E) Ascus; (F) Ascospores; (G) Paraphyses.
Colour: light green to dark green. Apothecia: rounded
to irregular, 0.2–1.5 mm in diameter, sessile, constricted
at base, frequent, separate, rarely fascicle. Margins: thin,
continuous, concolorous to disc, distinct when young,
indistinct when mature. Disc: black, flat to strongly convex, smooth and slightly glossy, slightly pruinose. Exci-
A taxonomic and phylogenetic study of some Lecidella species from Pakistan
225
ple: bluish green, 40–50 µm thick. Epihymenium: dark
blue with blackish tint, 20–30 µm tall. Hymenium: hyaline to grey, 110–120 µm tall, inspersed. Hypothecium:
hyaline to brown, 90–110 µm tall. Paraphyses: hyaline,
aseptate, apically branched, not anastomosing, apex
slightly swollen, 1–2 µm wide. Asci: clavate, 8-spored,
30–67 µm × 12–20 µm. Ascospores: hyaline, simple, narrowly ellipsoid to ovoid, 9–13 µm × 5–6 µm.
78–88 µm thick, bluish green to brown. Epihymenium:
dark brown, 12–16 µm high. Hymenium: hyaline, 75–80
µm tall, not inspersed. Hypothecium: hyaline to brown,
65–75 µm tall. Paraphyses: aseptate, hyaline, rarely anastomosing or branched, slightly swollen apically, up to 1.2
µm wide. Asci: lecanoral-type, clavate, 8-spored, 40–57
x 14–18 µm; Ascospores: simple, hyaline, thick and
smooth wall, ellipsoid to ovoid, 9–14 x 5–8 µm.
Spot Test
Spot tests
K+ (Slight yellow), C–, KC–, P– (atranonin or lichexanthone might be predicted).
K+ yellow, C–, KC–, P– (atranonin, zeorin, lichexanthone or norlichexanthone might be predicted).
Substrate and ecology
Substrate and ecology
L. patavina was found on nutrient enriched siliceous
rocks near water falls, in moist temperate forest at an
altitude of 1,705 m, temperature ranges between -2–37
oC, average annual rainfall 1,500– 1,650 mm, with dominant tree species Cedrus deodara (Roxb. ex Lambert)
G.Don, Pinus wallichiana A.B. Jacks., Picea smithiana
Boiss., Abies pindrow Royle., Quercus incana W.Bartram,
Q. dilatata Lindl. ex Royle. and Q. semecarpifolia Sm.
L. stigmatea was found on bark of Pinus wallichiana
A.B. Jacks., in moist temperate forest, with dominant
tree species Cedrus deodara (Roxb. ex Lambert) G.Don,
Pinus wallichiana A.B. Jacks., Picea smithiana Boiss.,
Abies pindrow Royle., Quercus incana W. Bartram, Q.
dilatata Lindl. ex Royle. and Q. semecarpifolia Sm. at an
altitude of 3,300 m, temperature ranges between -2–37
oC, average annual rainfall 1650 mm.
The second collection of L. stigmatea was found on
a rock (saxicolous), close to water falls, in moist temperate forest at an altitude of 2,900 m, temperature ranges
between -2–37 oC, average annual rainfall 1,500– 1,650
mm.
Distribution
Mainly arctic-alpine, UK, N.W. Scotland (Smith et
al. 2009), Africa, Antarctica, Europe, USA, (Knoph and
Leuckert 2004; Knudsen and Kocourková 2012), Ukraine
(Vondrak et al. 2010), temperate parts of Asia (Nash et
al. 2004), China (Zhao et al. 2015) and Pakistan (Aptroot
and Iqbal 2012).
Material examined
PAKISTAN: Khyber Pakhtunkhwa Province: Kurram District, Parachinar: 33°90’ N, 70°08’ E; 1705 m;
on rock; 18 August 2018, A.N. Khalid & K. Habib;
LAH3640; (GenBank accession no. MT139652).
Lecidella stigmatea (Ach.) Hertel & Leuckert, Willdenowia 5: 375. 1969. (Figure 4, A-G; Figure 6; E-H).
Description
Thallus crustose, areolate or rimose-areolate to verruculose-areolate, 7–8 cm in diameter, up to 0.2 mm
thick, rarely thick up to 0.6 mm. Areoles: indistinct to
distinct, flat to slightly convex, angular to irregular in
outline, 0.2–0.6 mm in diameter. Surface: finely granulose, rough, dull. Colour: greyish to grey to brownish
grey. Apothecia: lecideine, black, sessile, 0.4–0.9 mm in
diameter. Disc: black, flat to somewhat convex, pruinose. Margins: distinct, thin, becoming excluded. Exciple:
Distribution
Probably cosmopolitan, including UK, USA, Antarctica, Australia (Smith et al. 2009), Ukraine (Oxner 1968,
Kondratyuk et al. 1998, 2003), temperate parts of Asia
(Nash et al. 2004), China (Zhao et al. 2015), India (Singh
& Sinha 2010) and Pakistan (Ahmad 1965).
Material examined
PAKISTAN: Gilgit Baltistan, Fairy Meadows: 35°23’
N, 74°35’ E; 3300 m; on bark of Pinus wallichiana A.B.
Jacks., 18 July 2017, A.N. Khalid & M. Khan; LAH36402;
(GenBank accession no. MT139653); Azad Jammu &
Kashmir: District Muzaffarbad, Peer Chanasi: 34°2’ N,
73°33’ E; 2900 m; on rock; 18 August 2018, K. Habib;
LAH36403; (Genbank accession no. MT139650).
Key to Genus Lecidella in Pakistan
1a. On Bark ................................................................. L. tumidula
1b. On rock, Cortex K-, C ............................................................ 2
1c. On rock, or on bark, Cortex K+, C ...................................... 3
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Rizwana Zulfiqar, Kamran Habib, Memoona Khan, Abdul Nasir Khalid
Figure 4. Lecidella stigmatea. (A & B) Crustose thallus showing abundant apothecia; (C) black, pruinose disc of apothecia; (D) Cross section of an apothecium (H: Hymenium; E: Epihymenium); (E) Ascus; (F) Ascospores; (G) Paraphyses.
1d. On rock, Cortex K+, C ........................................................... 4
2a. Thallus crustose, areolate, epihymenium 15–20 µm tall,
hypothecium brownish grey, on rock ...............L. carpathica
2b. Thallus crustose, granulose, epihymenium 30–36 µm tall,
hypothecium dark reddish brown, on bark ....... L. tumidula
3a. Thallus rimose-areolate, paraphyses rarely branched, exciple greenish black, hymenium not inspersed ... L. stigmatea
A taxonomic and phylogenetic study of some Lecidella species from Pakistan
227
Figure 5. (A-D): Line drawings of Lecidella tumidula A: Granulose
thallus; B: Ascus; C: Ascospores; D: Paraphyses; (E-H): Line drawings of Lecidella carpathica E: Crustose-areolate thallus; F: Ascus;
G: Paraphyses; H: Ascospores.
Figure 6. (A-D): Line drawings of Lecidella patavina A: crustose
thallus B: Ascus; C: Ascospores; D: Paraphyses; (E-H): Line drawings of Lecidella stigmatea E: Rimose-areolate thallus; F: Ascus; G:
Paraphyses; H: Ascospores.
3b. Thallus inconspicuous, paraphyses mostly branched, exciple bluish green, hymenium inspersed ................L. patavina
In the ITS-based phylogenetic analysis, the Pakistani
collection of L. tumidula (PR–112) clustered with specimens of the same taxon reported from China (Accession
no. KT453736, KT453737) with strong support (99%, Fig.
7) and formed a sister group relationship with L. tumidula
collected from USA (Accession no. HQ650596) that was
wrongly reported in GenBank (Schmull et al. 2011), but
after re-examination it was L. tumidula (Zhao et al. 2015).
Morphological comparison also confirms its identity as L. tumidula (Nash et al. 2004) except the presence
of a pruinose disc in the specimen collected from Pakistan. There is only one nucleotide difference between the
Pakistani collection and the Chinese L. tumidula specimen. It is also the second report of this taxon from Asia
after China (Zhao et al. 2015).
4a. Thallus crustose, granulose to rimose-areolate, hymenium
55–100 µm tall ....................................................... L. euphorea
4b. Thallus sorediate, farinose, hymenium 60–70 µm tall, .........
........................................................................ L. pulveraceae
Discussion
The present study of the genus Lecidella from northern Pakistan revealed one new record for the lichen flora
of Pakistan while other Lecidella species have been collected from new localities thus indicating their wide distributional range in Pakistan.
228
Rizwana Zulfiqar, Kamran Habib, Memoona Khan, Abdul Nasir Khalid
Figure 7. Phylogenetic analysis of species of Lecidella, comprised of 39 sequences. This tree has been inferred using maximum likelihood method. The bootstrap values based on 1000 replicates are shown below the branches. Sequences generated from local collection are
marked with .
In the phylogenetic analysis, the ITS sequence
of Lecidella carpathica (CHK-04) clustered with ITS
sequences of L. carpathica collected from Canada
(KT695322, KT695353). Morphological comparison also
confirms its identity as L. carpathica (Nash et al. 2004).
There is only one nucleotide difference between Pakistani and Canadian L. carpathica (KT695322, KT695353).
Previously, it was reported from dry temperate forest of
Kalam (Swat), Pakistan at an elavation of about 2,001 m.
a.s.l. The new collection is from moist temperate forest of
AJK state, found at an elevation of 2,900 m. a.s.l.
The ITS sequence of Lecidella patavina (PR-11) clustered with a Chinese collection of L. patavina (Accession
no. KT453767) in the phylogenetic analysis and made a
sister branch with L. stigmatea (Ach.) Hertel & Leuckert. Morphologically, both Lecidella species are similar
in having the same chemistry, a crustose epilithic thallus
with a black disc of apothecia (Basaran et al. 2014) but
differ due to the presence of the conspicuous and thicker
thallus in the latter.
There are also two nucleotide difference between
Pakistani and Chinese collection of L. patavina
(KT453767). From Pakistan, it was previously reported
but locality was not recorded. Here it is described from
high elevation in Parachinar i.e. 1,705 m. a.s.l., where it
is widespread in warm and temperate climates.
A taxonomic and phylogenetic study of some Lecidella species from Pakistan
The ITS sequence of Lecidella stigmatea (MKF–7
and PC–34), clustered with L. stigmatea reported from
China (KT453763, KT453764, KT453765). The L. stigmatea (MK620163, MK620136) in our tree formed separate subclade. Results from phylogenetic analysis of L.
stigmatea requires revision between specimens identified as L. stigmatea from Argentina, are distantly related to samples of our clade. Morphologically our specimen is similar to the Sonoran L. stigmatea, in having
rimose-areolate thallus and lecideine apothecia (Nash
et al. 2004). From Pakistan, this taxon has previously been reported from dry temperate forest of Kalam
(Swat), Pakistan, at an elavation of about 2,001 m. a.s.l.
but recent collections are from moist temperate forests
of Azad Jammu & Kashmir and Gilgit Baltistan, at high
elevations 2,900 m and 3,300 m. a.s.l. respectively.
ACKNOWLEDGEMENTS
We are sincerely thankful to Prof. Dr. Sci. Sergey
Yakovych Kondratyuk, Kholodny Institute of Botany,
National Academy of Sciences of Ukraine, for presubmission review of the manuscript and Dr. Francis
Q. Brearley, School of Science and the Environment,
Manchester Metropolitan University, for linguistic revision of the manuscript.
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