Research Article |
Corresponding author: Saisamorn Lumyong ( scboi009@gmail.com ) Academic editor: Kentaro Hosaka
© 2019 Jaturong Kumla, Nakarin Suwannarach, Witchaphart Sungpalee, Kriangsak Sri-Ngernyuang, Saisamorn Lumyong.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Kumla J, Suwannarach N, Sungpalee W, Sri-Ngernyuang K, Lumyong S (2019) Clitopilus lampangensis (Agaricales, Entolomataceae), a new species from northern Thailand. MycoKeys 58: 69-82. https://doi.org/10.3897/mycokeys.58.36307
|
A new species of agaricomycetes, Clitopilus lampangensis, is described based on collections from northern Thailand. This species was distinguished from previously described Clitopilus species by its pale yellow to grayish yellow pileus with the presence of wider caulocystidia. Molecular phylogenetic analyses, based on the data of the internal transcribed spacers (ITS) and the large subunit (LSU) of the nuclear ribosomal DNA, and the second largest subunit of RNA polymerase II (rbp2) genes, also support the finding that C. lampangensis is distinct from other species within the genus Clitopilus. A full description, color photographs, illustrations and a phylogenetic tree showing the position of C. lampangensis are provided.
Agaricomycetes, gill mushroom, morphology, phylogeny, tropics
The genus Clitopilus was proposed by Kummer (1987) with C. prunulus (Scop.) P. Kummer as the type species. It belongs to the family Entolomataceae of the order Agaricales. This genus is saprotrophic and is widely distributed, especially in northern temperate areas (
Only six species, Clitopilus apalus (Berk. & Br.) Petch, C. crispus Pat. C. doimaesalongensis Jatuwong, Karun. & K.D. Hyde, C. chalybescens T.J. Baroni & Desjardin, C. peri (Berk. & Br.) Petch and C. prunulus, have been reported in Thailand (
Basidiocarps were collected in Mae Moh District, Lampang Province, northern Thailand in 2018. Basidiocarps were wrapped in aluminum foil and kept in plastic specimen boxes to be transported to the laboratory. Notes on the macromorphological features and photographs were obtained within 24 h of collection. The specimens were dried at 40–45 °C and deposited at the Herbarium of the Sustainable Development of Biological Resources Laboratory, Faculty of Science, Chiang Mai University (
Macromorphological data were recorded from fresh specimens. The recording of color names and codes followed
Genomic DNA of dry specimens (1–10 mg) was extracted using a Genomic DNA Extraction Mini-Kit (FAVORGEN, Taiwan). The ITS region of DNA was amplified by polymerase chain reactions (PCR) using ITS4 and ITS5 primers (
For phylogenetic analyses, the sequences from this study, previous studies and the GenBank database were used and provided in Table
Sequences used for phylogenetic analysis. The newly generated sequences are in bold.
Taxa | Voucher/strain | GenBank accession number | Refernces | ||
---|---|---|---|---|---|
ITS | LSU | rpb2 | |||
Clitopilus albidus | CAL 1320 | MF926596 | MF926595 | MF946579 |
|
CORT:26394WAT | – | KR869936 | KC816906 |
|
|
M536 | – | AF261287 | – |
|
|
Clitopilus austroprunulus | MEN2009062 | KC139085 | – | – |
|
MEN2009001 | KC139084 | – | – |
|
|
Clitopilus cf. argentinus | MTB480412 | – | – | KC816907 |
|
Clitopilus chalybescens | MFUCC130808 | KP938184 | – | – |
|
MFUCC130809 | KP938185 | – | – |
|
|
SDBR-CMUUP0039 | MK773645 | MK764940 | MK784129 | This study | |
Clitopilus chrischonensis | TOHG 1994 | HM623128 | HM623131 | – |
|
Clitopilus crispus | GDGM29931 | JQ281489 | – | – |
|
CORT:9982 | – | – | KC816910 |
|
|
CORT:10027 | – | – | KC816911 |
|
|
Clitopilus cystidiatus | 26 | – | GQ289147 | GQ289220 |
|
TOAV130 | HM623129 | HM623132 | – |
|
|
Clitopilus doimaesalongensis | MFUCC130806 | KP938183 | – | – |
|
Clitopilus fusiformis | SAAS1038 | KY385634 | – | KY385632 |
|
SAAS1892 | KU751777 | – | KY385633 |
|
|
Clitopilus giovanellae | SF14368 | EF413030 | EF413027 | – |
|
Clitopilus hobsonii | CBS 270.36 | FJ770395 | – | – |
|
CBS 445.86 | FJ770385 | – | – |
|
|
DLL9635 | – | – | KC816913 |
|
|
DLL9643 | – | – | KC816913 |
|
|
Clitopilus lampangensis | SDBR-CMUJK 0147 | MK764933 | MK764935 | MK784127 | This study |
SDBR-CMUNK 0047 | MK764934 | MK773856 | MK784128 | This study | |
Clitopilus kamaka | KA12-0364 | KR673433 | – | – |
|
Clitopilus orientalis | CAL 1616 | MG345134 | MG321558 | MG321559 |
|
Clitopilus passeckeriamus | CBS299.35 | MH855682 | MH867198 | – |
|
P78 | KY962494 | KY963078 | – | Unpublished | |
Clitopilus paxilloides | CORT:5809 | – | – | KC816919 |
|
Clitopilus peri | CORT:10033 | – | – | KC816920 |
|
CORT:10040 | – | – | KC816921 |
|
|
CORT:10041 | – | – | KC816922 |
|
|
Clitopilus pinsitus | CBS 623.70 | MH859879 | MH871665 | – |
|
Clitopilus prunulus | Champ-15 | KX449418 | – | – |
|
CBS 227.93 | FJ770408 | – | – |
|
|
Noordeloos 2003-09-14 | KR261096 | – | – | Unpublished | |
COPT:7003 | – | – | KC816925 |
|
|
TB9663 | – | – | GU384648 |
|
|
TB8229 | – | – | GU384650 |
|
|
COPT:REH8456 | – | – | KC816923 |
|
|
Clitopilus reticulosporus | DC-2010 | KC885966 | HM164414 | HM164416 |
|
Clitopilus scyphoides | CBS 127.47 | MH856181 | MH867707 | – |
|
CBS 400.79 | FJ770401 | – | – |
|
|
Clitopilus subscyphoides | CAL 1325 | MF927542 | MF946580 | MF946581 |
|
Clitopilus venososulcatus | CORT:8111 | – | – | KC816930 |
|
Rhodocybe griseoaurantia | CAL 1324 | KX083571 | KX83574 | KX083568 | Unpublished |
Rhodocybe pallidogrisea | CORT 013944 | NR154437 | – | KC816968 |
|
The topology of each single-gene of ITS and LSU, and the combined ITS and LSU phylograms were found to be similar. However, differences were observed in the topology of the rbp2 gene. Therefore, we present only the combined ITS and LSU gene phylogram (Fig.
Both the combined ITS and LSU, and the rbp2 phylograms indicated that the sequences were of a new species, C. lampangensis, that had formed a monophyletic clade with high BS (100 %) and PP (1.0) support (Figs
Phylogram derived from maximum likelihood analysis of the combined ITS and LSU region of nuclear rDNA of 34 sequences. Rhodocybe griseoaurantia and R. pallidogrisea were used as outgroup. The numbers above branches represent maximum likelihood bootstrap percentages (left) and Bayesian posterior probabilities (right). Only bootstrap values ≥ 50 % are shown, and the scale bar represents ten substitutions per nucleotide position. The fungal species obtained in this study are in bold.
Phylogram derived from maximum likelihood analysis of rpb2 gene of 27 sequences. Rhodocybe griseoaurantia and R. pallidogrisea were used as outgroup. The numbers above branches represent maximum likelihood bootstrap percentages (left) and Bayesian posterior probabilities (right). Only bootstrap values ≥ 50 % are shown, and the scale bar represents ten substitutions per nucleotide position. The fungal species obtained in this study are in bold.
Distinguished from other Clitopilus species by its pale yellow to grayish yellow pileus with the presence of caulocystidia, and from C. chalybescens by its wider caulocystidia, longer basidiospores, and lack of grayish blue color change on the pileus and stipe when bruised.
‘lampangensis’, referring to Lampang Province, where the holotype was found.
THAILAND, Lampang Province, Mae Moh District, (18°24'21"N, 99°42'26"E, elevation 380 m), on ground in a tropical deciduous forest, May, 2018, J. Kumla & N. Suwannarach, SDBR-CMUJK 0147 and BBH 43590 (isotype).
MK764933 (ITS), MK764935 (LSU) and MK784127 (rbp2).
Basidiocarps small, clitocyboid. Pileus 35–50 mm diam., initially convex or somewhat plano-convex with or without a central depression, becoming deeply umbilicate with age; surface pale yellow (4A3) to greyish yellow (4B5), somewhat velutinous, finely pruinose all over; margin incurved to slightly inrolled, entire or slightly wavy. Lamellae subdecurrent to decurrent, white (1A1), crowded, up to 2.5 mm wide, with lamellulae of 1–3 lengths; edge entire or slightly wavy, concolorous with the sides. Stipe 20–25 × 5–8 mm, central, solid; surface white (1A1) to yellowish white (4A2), finely pruinose all over, densely so towards the apex; base with white cottony mycelium. Odor strong farinaceous. A pale pinkish spore print.
Basidiospores 7.0–9.0 × 3.0–5.0 μm, Q = 1.40–2.33, Q = 1.82 ± 0.27, ellipsoid in polar view, amygdaliform to limoniform in side view, with 6–8 prominent longitudinal ridges, colorless, thin-walled. Basidia 17.0–25.0 × 4.0–8.0 μm, clavate, colorless, thin-walled, 2- and 4-spored; sterigmata up to 4 μm long. Lamella-edge fertile. Pleurocystidia and cheilocystidia absent. Lamellar trama subregular; hyphae 2.5–4.0 μm wide, hyaline, thin-walled. Pileus trama compact, hyaline, cylindrical hyphae 5–10 μm wide. Pileipellis a cutis of loosely interwoven hyphae; 3–5 μm wide, hyaline, thin-walled, and terminal cells; subcylindric or narrowly clavate, 4–8 μm wide. Stipitipellis at stipe apex a layer of repent, hyaline, cylindrical hyphae 4–8 μm wide, thin-walled. Caulocystidia 25.5–42.5 × 8.0–15.0 μm, single or clustered, erect or repent, varying in shape from cylindrical to clavate, hyaline, slightly thick-walled. Clamp connections absent in all tissues.
Fruiting solitary or gregarious on soil in a tropical deciduous forest. Known only from northern Thailand
The present study has identified a new species of Clitopilus acquired from northern Thailand based on both morphological characteristics and phylogenetic analyses. Clitopilus lampangensis is characterized by its clitocyboid, pale yellow to grayish yellow basidiocarps, pinkish spore-print, ellipsoid basidiospores with longitudinal ridges and hyphae lacking clamp connections. Thus, these morphological characteristics support its placement into the genus Clitopilus (
Taxa | Origin | Pileus | Basidia | Caulocystidia | Basidiospores |
---|---|---|---|---|---|
C. lampangensis a | Thailand | 35–50 mm in diameter, pale yellow to greyish yellow | 17.0–25.0 × 4.0–8.0 μm, 2–4 streigmata | 25.5–42.5 × 8.0–15.0 μm | Ellipsoid, 7.0–9.0 × 3.0–5.0 μm, 6–8 longitudinal ridges |
C. chalybescens b, c | Thailand | 15–90 mm in diameter, white, yellowish white to greyish blue | 15.0–21.0 × 5.1–8.0 μm, 4 streigmata | 16.0–32.0 × 5.0–7.0 μm | Ellipsoid, 5.3–7.5 × 3.6–5.0 μm, 8–10 longitudinal ridges |
C. peri d,e | India, Sri Lanka, Thailand | 8–22 mm in diameter, white | 16.0–18.0 × 5.0–7.0 μm, 4 streigmata | Absent | Ellipsoid, 6.7–8.5 × 3.0–4.0 μm, 6–9 longitudinal ridges |
C. prunulus f,g | Netherlands, Thailand, United State | 25–80 mm in diameter, white, yellowish white to grayish or yellow cream | 25.0–47.0 × 7.0–12.0 μm, 4 streigmata | Absent | Ellipsoid, 9.0–14.0 × 4.5–8.0 μm, 6–8 longitudinal ridges |
C. fasciculatus h | Netherlands, | 20–70 mm in diameter, pale brown | Sizes were not reported, 4 streigmata | Absent | Ellipsoid, 4.5–6.3 × 3.0–4.0 μm, 3–6 longitudinal ridges |
C. gallaecicus i | Spain | 80–90 mm in diameter, creamy, ochre to ochre-brown | 20.0–35.0 × 8.5–10.5 μm, 4 streigmata | Absent | Ellipsoid, 8.0–14.5 × 4.5–7.5 μm, 3–6 longitudinal ridges |
C. incrustatus j | Costa Rica, United State | 80–90 mm in diameter, grayish brown | 16.0–24.0 × 7.0–8.0 μm, 4 streigmata | Absent | Ellipsoid, 5.0–6.5 × 3.0–4.0 μm, 3–6 longitudinal ridges |
C. djellouliae k | France | 6–18 mm in diameter, light yellowish brown | 22.0–32.0 × 7.5–8.5 μm, 4 streigmata | Absent | Ellipsoid, 6.0–9.0 × 4.0–6.0 μm |
C. giovanellae l,m | Italy, Spain | 5–15 mm in diameter, grayish to light brown | 14.0–22.0 × 6.5–9.5 μm, 4 streigmata | Absent | Ellipsoid, 5.0–8.0 × 3.0–4.0 μm |
C. luteocinnamomeus n | Panama | 15–45 mm in diameter, ochre to light cinnamon-brown | 19.0–27.0 × 6.0–7.0 μm, 4 streigmata | Absent | Subglobose to ellipsoid, 4.5–6.0 × 3.5–5.0 μm |
C. catalonicus o | Panama | Up to 15mm in diameter, light yellowish brown | 32.0–40.0 × 6.4–8.0 μm, 4 streigmata | Absent | Ellipsoid, 5.3–7.5 × 3.7–4.5 μm |
The phylogenetic analyses of the combined ITS and LSU, and rpb2 sequences confirmed that C. lampangensis formed a monophyletic clade which clearly separated it from the other Clitopilus species. Clitopilus lampangensis forms a sister taxon to C. chalybescens and C. peri. Clitopilus peri differs from C. lampangensis by its smaller white basidiocarps (8–22 mm in diameter) and the absence of caulocystidia (
Therefore, a combination of the morphological characteristics and the molecular analyses strongly support recognition of a new fungus species. This discovery is considered important in terms of stimulating a deeper investigation of macrofungi in Thailand, and will help researchers to better understand the distribution and ecology of Clitopilus.
1 | Pileus white to chalk-white colors | 2 |
– | Pileus white or with other colors | 5 |
2 | Stipe ≥ 3 mm thick | 3 |
– | Stipe < 3 mm thick | C. peri |
3 | Basidia < 8 μm wide | 4 |
– | Basidia ≥ 8 μm wide, basidiospores 6.8–9.2 × 4.1–5.5 μm | C. doimaesalongensis |
4 | Basidia up to 25 μm, basidiospores 6–8.5 × 4.5–5.5 μm | C. apalus |
– | Basidia up to 30 μm, basidiospores 5.5–9 × 4–6 μm | C. cripus |
5 | Pileus white to pale grayish or yellowish cream colors | 6 |
– | Pileus pale yellow to greyish yellow colors, caulocystidia present, basidiospores 7.0–9.0 × 3.0–5.0 μm | C. lampangensis |
6 | Basidia ≥ 25 μm long, caulocystidia absent, basidiospores 8.0–12.0 × 4.0–6.5 μm | C. prunulus |
– | Basidia < 25 μm long, caulocystidia present, basidiospores 5.3–7.5 × 3.6–5.0 μm | C. chalybecens |
This work was supported by grants from Chiang Mai University and Center of Excellence on Biodiversity (BDC), Office of Higher Education Commission (BDC-PG3-161005), Thailand. We are grateful to staff of Mae Moh Forestry Industry Organization for their excellent field assistance, and Dr. Eric H.C. McKenzie for English proof reading.