Print
Dryopteris wulingshanensis (Dryopteridaceae), a new species from Hunan, China
expand article infoJiang-Ping Shu§|, Zi-Yue Liu|, Zhi-Rong Gu#, Li-Jun Chen¤, Hong-Jin Wei«, Xi-Le Zhou», Yue-Hong Yan˄|, Rui-Jiang Wang|
‡ South China Botanical Garden, Chinese Academy of Sciences, Guangzhou, China
§ Shenzhen Key Laboratory for Orchid Conservation and Utilization, The National Orchid Conservation Centre of China, Shenzhen, China
| University of Chinese Academy of Sciences, Beijing, China
¶ Gannan Normal University, Ganzhou, China
# Badagongshan National Nature Reserve, Zhangjiajie, China
¤ Shenzhen Key Laboratory for Orchid Conservation and Utilization, the National Orchid Conservation Center of China, Shenzhen, China
« Shanghai Chenshan Botanical Garden, Shanghai, China
» Xiangxi Tujia and Miao Autonomous Prefecture Forest Resources Monitoring Center, Jishou, China
˄ Key Laboratory of National Forestry and Grassland Administration for Orchid Conservation and Utilization, Shenzhen, China
Open Access

Abstract

Dryopteris wulingshanensis, a new species growing on limestone in the Wulingshan Mountains, Hunan, China, is described and illustrated. This species is most similar to D. jishouensis and D. gymnophylla on general morphological traits, such as the form of scales, rhizome and sori, but differs by the number of vascular bundles at the base of the petiole, length to width ratio of lamina, stalk length of basal pinnae, division of the lamina, apex form of the pinnule and habitat. Moreover, molecular phylogenetic analysis using the chloroplast rbcL gene suggested that D. wulingshanensis, as the sister group of D. jishouensis, is a monophyletic clade. According to its restricted geographic range, small populations and few individuals, D. wulingshanensis should be considered endangered, according to the IUCN Red List criteria.

Keywords

New taxon, rbcL, subg. Dryopteris, phylogeny, fern

Introduction

Dryopteris Adans. (1763: 20, 551) is one of the largest fern genera with about 400 species, widely distributed all over the world (Wu et al. 2013). Based on molecular phylogenetic evidence, several genera are nested within Dryopteris, such as Acrophorus C. Presl, Acrorumohra (H. Itô) H. Itô, Diacalpe Blume, Dryopsis Holttum & P. J. Edwards, Nothoperanema (Tagawa) Ching and Peranema D. Don (Zhang and Zhang 2012; Zhang et al. 2012). Most species in Dryopteris share a short rhizome and catadromous arrangement of frond segments, compared to its sister genus, Arachniodes Blume, which has long-creeping rhizomes and anadromous laminae (Zhang et al. 2012; Wu et al. 2013). The species of this genus usually grow in forests, open vegetation and, occasionally, in the rocky area of temperate and tropical regions (Fraser-Jenkins 1986; Kramer et al. 1990; Wu et al. 2013). In China, the genus is widely distributed, especially in south-western regions, with about 167 species with 60 endemic species in four subgenera (D.subg. Pycnopteris, D.subg. Nothoperanema, D.subg. Dryopteris, and D.subg. Erythrovariae) (Wu et al. 2013).

During 2016–2021, we surveyed ferns in the Wulingshan Mountains, which occupy the border zone of four Provinces in China (Hubei, Chongqing, Guizhou and Hunan). This region, as one of the biodiversity hotspots, nurtures a large number of endemic plants and preserves many relict plants (Chen et al. 2004; Yan and Zhou 2021). When we arrived at the Pangu Peak of Dehang Scenic Area, Jishou City, Hunan, an epipetric species that grows in the limestone crevices caught our attention. It is most similar to Dryopteris gymnophylla (Baker) C. Chr. and Dryopteris jishouensis G.X. Chen & D.G. Zhang but differs by the length to width ratio of lamina, stalks of the basal pinnae, apex form of pinnules and habitat. Moreover, we found this unknown species was also distributed in Mt. Tianmenshan and Zhongli Grand Canyon of Zhangjiajie City, Hunan, China. In order to infer the phylogenetic position of this species, the chloroplast rbcL sequences of 32 individuals, representing 11 closely related species, were analyzed. Based on morphological and molecular phylogenetic evidence, we describe it as a new species, named Dryopteris wulingshanensis J.P. Shu, Y.H. Yan & R.J. Wang and illustrate it here.

Materials and methods

DNA extraction and sequencing

A total of 32 samples, representing 11 species of the genus Dryopteris, were analyzed to infer the phylogenetic relationships amongst the unknown species and its closest relatives. Dryopteris aemula (Aiton) Kuntze was sampled as an outgroup based on the previous phylogenetic studies of the genus Dryopteris (Zhang and Zhang 2012; Zhang et al. 2012). The rbcL gene of 18 individuals were newly sequenced and submitted on the GenBank (Table 1), and the others were obtained from GenBank database. Total genomic DNA was extracted from silica gel-dried leaves by using a DNA secure Plant Kit (Tiangen Biotech, Beijing, China) according to the manufacturer’s protocols. The primers and amplification reaction of rbcL gene followed the protocols of Shu et al. (2017). Sequencing reactions were set up to obtain both the forward and reverse sequences, and then sequenced on an ABI 3730xl DNA Analyzer (Applied Biosystems, Foster City, California, USA).

Table 1.

Information of 18 samples newly sequenced in this study.

Taxon Voucher specimen Locality rbcL
Dryopteris jishouensis JSL3612 Guangxi, China MZ444597
JSL3610 Guangxi, China MZ444596
JSL3607 Guangxi, China MZ444595
ZXL6320 Hunan, China MZ444598
ZXL6317 Hunan, China MZ444593
YYH7842 Guizhou, China MZ444594
Dryopteris wulingshanensis ZXL6320-3 Hunan, China MZ444607
ZXL6320-1 Hunan, China MZ444606
JSL3935 Hunan, China MZ444605
JSL3926 Hunan, China MZ444604
Dryopteris gymnophylla ZXL6626 Jiangxi, China MZ444602
JSL4354 Zhejiang, China MZ444599
JSL3320 Anhui, China MZ444600
JSL3393 Anhui, China MZ444601
JSL3949 Anhui, China MZ444603
Dryopteris chinensis JSL3359 Anhui, China MZ444610
JSL3329 Anhui, China MZ444609
JSL2983 Anhui, China MZ444608

Molecular phylogenetic analysis

The consensus sequences were generated using SeqMan v7.1.0 (DNASTAR, USA) and then 32 sequences used for phylogenetic analysis were aligned with BioEdit v7.2.0 (Hall 1999). The Maximum Likelihood (ML) phylogenetic tree was constructed by IQ-TREE v2.1.3 (Minh et al. 2020), the best-fit model (K2P+I) was chosen according to Bayesian Information Criterion (BIC) with ModelFinder (Kalyaanamoorthy et al. 2017) and the branch support values of ultrafast bootstrap (UFBoot) approximation was performed with 1000 repetitions. Each bootstrap tree was optimized using a hill-climbing Nearest Neighbor Interchange (NNI) search, based directly on the corresponding bootstrap alignment to reduce the risk of overestimating branch supports with UFBoot (Hoang et al. 2017). The Bayesian Inference (BI) species tree was constructed by MrBayes v3.2.7 (Ronquist et al. 2012) with the GTR + I + G model. Four Markov Chain Monte Carlo (MCMC) chains were run simultaneously for two million generations, and sampled every 100 generations. The convergence was assessed with the average standard deviation of split frequencies lower than 0.01.

Results and discussion

A total of 32 samples were used for phylogenetic analysis, based on chloroplast rbcL gene and the length of sequence alignments was 1,204 bp after removing the missing or gap sites. The phylogenetic relationships amongst D. wulingshanensis and its relatives based on ML and BI algorithms, showed a tree topology to be nearly the same as previous studies (Zhang and Zhang 2012; Zhang et al. 2012). Our results indicated the new species D. wulingshanensis was a monophyletic clade (BS/PP = 100/-) and was sister to D. jishouensis (Fig. 1). Morphologically, the new species was similar to D. jishouensis and D. gymnophylla on the general morphological characteristics, but differs by the vascular bundles at the base of petiole, length/width ratio of the fronds, stalk length of basal pinnae, division of the lamina, pinnules and habitat (Fig. 2, Table 2).

Figure 1. 

The phylogenetic relationships amongst the new species Dryopteris wulingshanensis and its relatives. The topology was the Maximum Likelihood (ML) tree and bootstraps support values (BS) and posterior probability (PP) are showed on the branches.

Figure 2. 

Morphological comparison amongst Dryopteris wulingshanensis, D. jishouensis and D. gymnophylla A–C D. jishouensis (type locality, Hunan A plants and habitats B stalks of basal pinnae C scales at base of stipe) D–F D. wulingshanensis (Zhangjiajie, Hunan D plants and habitats E stalks of basal pinnae F scales at base of stipe) G–I D. gymnophylla (Zhejiang G plants and habitats H stalks of basal pinnae I scales at base of stipe).

Table 2.

The morphological comparison of Dryopteris wulingshanensis, D. jishouensis and D. gymnophylla.

Species characters D. jishouensis D. wulingshanensis D. gymnophylla
Vascular bundles at the base of petiole 2–3 About 5 7–8
Length/width of the fronds 1.7–2.0 times 1.3–1.6 times 1–1.2 times
Stalk of basal pinnae Usually shorter than 1.5 cm With a long stalk, usually up to 3cm or more With a long stalk, usually up to 3cm or more
Division of the lamina 3× pinnate 4× pinnate- pinnatipartite 3× pinnate-pinnatipartite
Pinnules Obtuse Acuminate Acuminate
Habitat Epipetric Epipetric Terrestrial

Taxonomic treatment

Dryopteris wulingshanensis J.P.Shu, Y.H.Yan & R.J.Wang, sp. nov.

Figs 3, 4 武陵山鳞毛蕨

Type

China. Hunan: Wulingshan Mountains Zhongli Grand Canyon, Sangzhi County, Zhangjiajie City. 29°39'10.08"N, 110°37'04.29"E, 900 m alt., 26 June 2021, Y.-H. Yan & Z.-R. Gu, YYH24468 (holotype, IBSC; isotypes, NOCC!, CSH!,PE!).

Diagnosis

The morphology of D. wulingshanensis was intermediated between D. jishouensis and D. gymnophylla, but more similar to the former. Dryopteris wulingshanensis and D. jishouensis both grow in alkaline soil, but the former’s fronds are ovate, length/width 1.3–1.6 times, tripinnate to quadripartite; the latter’s fronds are ovate-lanceolate to triangular-lanceolate, length/width 1.7–2.0 times or more, bipinnate to tripinnatisect. Dryopteris gymnophylla grows in acidic soil, the fronds are pentagonal, usually length/width 1–1.2 times, tripinnate to quadripinnnatisect (Fig. 2, Table 2).

Figure 3. 

Dryopteris wulingshanensis J.P. Shu, Y.H. Yan & R.J. Wang A habit B pinnule with sori C scale at base of stipe D sporangium E spore (drawn by Li-Jun Chen, based on the type material at IBSC).

Figure 4. 

The habitat and morphological characters of Dryopteris wulingshanensis J.P. Shu, Y.H. Yan & R.J. Wang A habitat B lamina C basal pinna D sori E rhizome F base of stipe G spore (Left distal pole, Right proximal pole) H sporangium I scale.

Description

Rhizome short-creeping, apex scaly; scales dark brown, lanceolate, margin entire or 1–2 dentate at base. Frond approximate, (40–)65–70 cm; stipe (23–)31–36 cm, medial diameter 1.5–2 mm, basal scales similar to rhizome scales, antrorse, glabrous, stramineous to brown-stramineous, ventrally grooved; lamina ovate, (19–)32–36 × (12–)19–28 cm, about 1.3–1.6 times as long as wide, base round or cordate, apex acuminate, tripinnate to quadripartite (premature lamina only bipinnate to tripinnatisect); pinnae 6–8 pairs, oblique, distant, falcate, stipitate, basal pair longest and largest, opposite to nearly opposite, significantly falcate, stipe (1.2–)3–4.5(–7) cm, deltate-lanceolate, (8–)12–23 × (3.6–)7–11 cm, apex long-acuminate, base broad-cuneate to round, asymmetric, basiscopic pinnules longer than acroscopic pinnules, suprabasal pinnae with pinnules similar; pinnules 7–8 pairs, discrete, oblique to explanate, falcate, base broadly cuneate, pedicellate, basiscopic ones largest, (3–)5–10 × (1.5–)2.2–4 cm, stipe (1.5–)4–9 mm, trigonal oblong, apex obtuse, ultimate pinnules (lobe) 7–10 pairs, oblique to explanate, trigonal oblong, basiscopic ones longer than acroscopic ones, exstipitate, round-obtuse, base broad-cuneate, connate, pinnatifid to partite, (9–)14–20 (4)8–10 mm; lobes 2–7 pairs, 1.5–2 mm wide, oblong, entire, round, others ultimate pinnules lobate to pinnatifid or crenate, base broad-cuneate to decurrent; other pinnae decrescent, opposite to alternate, trigonal lanceolate to oblong lanceolate; rachis and costa glabrous, stramineous, adaxially sulcate, lamina papyraceous, glabrous; veins pinnate, single or dichotomous; sori round, on apices of veinlets, nearly incision; indusia orbicular-reniform, margin premorse, brown, persistent.

Additional specimens examined (paratypes)

China. Hunan: Pangu Peak, Dehang Scenic Area, Jishou City, Xiangxi Tujia and Miao Autonomous Prefecture. 28°20'50.046"N, 109°34'34.522"E, 914 m alt., 26 Jun 2016, X.-L. Zhou et al. ZXL6320-1, ZXL6320-3 (CSH!); Tianmenshan Scenic Area, Zhangjiajie City. 5 April 2016, H.-J. Wei et al. JSL3926, JSL3935 (CSH!).

Distribution and habitat

Dryopteris wulingshanensis is endemic to the Wulingshan mountains in Jishou and Zhangjiajie Cities, Hunan, China. It is epipetric in limestone crevices at an elevation of 700–1000 m in evergreen and deciduous broad-leaved mixed forest. The associated ferns include: Cheilanthes patula Baker, Cyrtomium fortunei J. Sm., Cyrtomium nephrolepioides (Christ) Copel., Lemmaphyllum drymoglossoides (Baker) Ching, Polystichum tsus-simense (Hook.) J. Sm., Pronephrium penangianum (Hook.) Holttum, Pyrrosia petiolosa (Christ) Ching and Woodwardia unigemmata (Makino) Nakai and associated seed plants include: Celtis sinensis Pers., Choerospondias axillaris (Roxb.) B.L. Burtt & A.W. Hill, Davidia involucrata Baill., Ficus sarmentosavar. henryi (King ex Oliv.) Corner, Hydrangea strigosa Rehder, Loropetalum chinense (R. Br.) Oliv., Mallotus philippinensis (Lam.) Muell. Arg., Miscanthus sinensis Andersson, Platycarya strobilacea Siebold & Zucc., Rhus chinensis Mill. and Viburnum cylindricum Buch.–Ham. ex D. Don.

Etymology

The specific epithet “wulingshanensis” is derived from the name of type locality Wulingshan Mountains, where the new species is found.

IUCN Conservation Assessment

EN(B1ab(iii)). Dryopteris wulingshanensis is only known from three locations of Wulingshan Mountains in Jishou and Zhangjiajie Cities, Hunan, China. Based on its restricted geographic range, small populations and few individuals, Dryopteris wulingshanensis should be considered endangered under the IUCN Red List criteria (IUCN 2019).

Acknowledgements

This research was funded by the Strategic Priority Research Program of the Chinese Academy of Sciences (XDA19050404). We are very grateful to the subject editor Blanca León and anonymous reviewers for their helpful comments and modification assistance. We thank Prof. Gong-Xi Chen from Jishou University and Mr. Yang Xiang from Badagongshan National Nature Reserve, Hunan Province, China, for assisting in the field surveys.

References

  • Chen GX, Liu SB, Ao CQ, Liao WB (2004) On endemic genera to China of spermatophytic flora from Mt. Wulingshan region. Xibei Zhiwu Xuebao 24(5): 865–871.
  • Fraser-Jenkins CR (1986) A classification of the genus Dryopteris (Pteridophyta: Dryopteridaceae). Bulletin of the British Museum (Natural History). Botany 14: 183–218.
  • Hall TA (1999) BioEdit: A user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series 41: 95–98.
  • Hoang DT, Chernomor O, von Haeseler A, Minh BQ, Vinh LS (2017) UFBoot2: Improving the Ultrafast Bootstrap Approximation. Molecular Biology and Evolution 35(2): 518–522. https://doi.org/10.1093/molbev/msx281
  • IUCN (2019) guidelines for using the IUCN red List Categories and Criteria. Version 14. Prepared by the Standards and Petitions Subcommittee. Available from: http://www.iucnredlist.org/documents/ RedListGuidelines.pdf [accessed: 6 January 2020]
  • Kalyaanamoorthy S, Minh BQ, Wong TKF, von Haeseler A, Jermiin LS (2017) ModelFinder: Fast model selection for accurate phylogenetic estimates. Nature Methods 14(6): 587–589. https://doi.org/10.1038/nmeth.4285
  • Kramer KU, Holttum RE, Moran RC, Smith AR (1990) Dryopteridaceae. In: Kubitzki K (Ed.) the families and genera of vascular plants, vol. 1. Pteridophytes and gymnosperms. Springer-Verlag, Berlin, 101–144.
  • Minh BQ, Schmidt HA, Chernomor O, Schrempf D, Woodhams MD, von Haeseler A, Lanfear R (2020) IQ-TREE 2: New Models and Efficient Methods for Phylogenetic Inference in the Genomic Era. Molecular Biology and Evolution 37(5): 1530–1534. https://doi.org/10.1093/molbev/msaa015
  • Ronquist F, Teslenko M, van der Mark P, Ayres DL, Darling A, Höhna S, Larget B, Liu L, Suchard MA, Huelsenbeck JP (2012) MrBayes 3.2: Efficient Bayesian phylogenetic inference and model choice across a large model space. Systematic Biology 61(3): 539–542. https://doi.org/10.1093/sysbio/sys029
  • Shu JP, Shang H, Jin D, Wei HJ, Zhou XL, Liu HM, Gu YF, Wang Y, Wang FG, Shen H, Zhang R, Adjie B, Yan YH (2017) Re-establishment of species from synonymies based on DNA barcoding and phylogenetic analysis using Diplopterygium simulans (Gleicheniaceae) as an example. PLoS One 12(3): e0164604. https://doi.org/10.1371/journal.pone.0164604
  • Wu SG, Xiang JY, Lu SG, Wang FG, Xing FW, Dong SY, He H, Zhang LB, David SB, Maarten JMC (2013) Dryopteris. In: Wu ZY, Raven PH, Hong DY (Eds) Flora of China vols. 2–3. Science Press, Beijing, 571–628.
  • Yan YH, Zhou XL (2021) The pteridophytes in Mt. Wulingshan region, China. China Forestry Publishing House, Beijing, 320 pp.
  • Zhang LB, Zhang L (2012) The inclusion of Acrophorus, Diacalpe, Nothoperanema, and Peranema in Dryopteris: The molecular phylogeny, systematics, and nomenclature of Dryopterissubg. Nothoperanema (Dryopteridaceae). Taxon 61(6): 1199–1216. https://doi.org/10.1002/tax.616003
  • Zhang LB, Zhang L, Dong SY, Sessa EB, Gao XF, Ebihara A (2012) Molecular circumscription and major evolutionary lineages of the fern genus Dryopteris (Dryopteridaceae). BMC Evolutionary Biology 12(1): 180. https://doi.org/10.1186/1471-2148-12-180