Research Article |
Corresponding author: Yanfen Chang ( changyf2018@126.com ) Academic editor: Blanca León
© 2022 Yanfen Chang, Guocheng Zhang, Zhixin Wang, Limin Cao.
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:
Chang Y, Zhang G, Wang Z, Cao L (2022) Molecular and morphological evidence reveals a new fern species of Hymenasplenium (Aspleniaceae) from south and southwestern China. PhytoKeys 211: 93-106. https://doi.org/10.3897/phytokeys.211.90363
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Hymenasplenium obtusidentatum, a new fern species of the H. excisum subclade of Hymenasplenium (Aspleniaceae) from south and southwestern China was described. Molecular phylogenetic analyses and morphological observations of H. obtusidentatum and related species clearly indicated that this is a distinct taxonomic entity. Phylogenetically, H. obtusidentatum was confirmed to represent a diverging lineage in the H. excisum subclade of Hymenasplenium and was closely related to one lineage that includes accessions identified as H. obscurum, H. pseudobscurum and H. tholiformis. Morphologically, H. obtusidentatum can be distinguished by the combination of its lamina base truncate, stipe not shiny and with color of reddish brown to dark brown, and pinna marginal teeth that are not sharp, but blunt or rounded. A complete species description and comparison with related species in the H. excisum subclade were provided. The holotype of H. obtusidentatum was designated.
Hymenasplenium, H. excisum subclade, new taxon, species complex, taxonomy
Hymenasplenium Hayata (1927: 712) is one of the two genera in the nearly globally distributed and species-rich fern family Aspleniaceae (
The H. excisum subclade belongs to one of the six subclades in the Old World clade of Hymenasplenium (
As a continued effort to clarify the species diversity and taxonomy of Hymenasplenium, we conducted fieldworks in south and southwestern China. During these trips, we collected some specimens of a new taxon of H. excisum subclade that were obviously different from all other species of the subclade in both morphology and phylogeny analyses. To clarify its taxonomic status, we studied the morphological differences between this taxon and related species and investigated the distinction of this putative new species by analyzing chloroplast DNA sequences for members of the monophyletic paleotropical clade of Hymenasplenium (
The morphological characteristics of the new species were observed in the field. Herbarium specimens of Hymenasplenium at PE, KUN, HITBC and PYU were studied. Digital specimens of related species of the new taxon were examined from the online databases of CVH (https://www.cvh.ac.cn/) and JSTOR Global Plants (https://plants.jstor.org/).
Spores were obtained from newly collected specimens and examined with an Olympus BX-51 light microscope to examine aborted spores and determine the number of spores per sporangium. Mature sporangia from each specimen were removed and ruptured with a needle tip. The number of spores per sporangium was counted. The presence of 64 spores per sporangium was considered an indicator of the absence of apomictic reproduction (
To clarify the phylogenetic position of the new species, the taxa were sampled to include representatives of all the six subclades in the Old World clade of Hymenasplenium (
Genomic DNA was extracted from the silica gel-dried leaf material of each sampled individual using the modified CTAB method (
Sequences were edited using the Staden Package (
Like most species in Hymenasplenium, the new species has a long-creeping rhizome, once-pinnate laminae, asymmetrical pinnae, and elliptic to reniform spores. However, the new species can be distinguished from other species in the genus by the combined characteristics of truncate lamina base, reddish brown to dark brown stipe and rachis, stipe not shiny and with scale or subglabrous, and pinna marginal teeth not sharp, but blunt or rounded (Figs
Comparison of morphological characters to differentiate the new taxon (Hymenasplenium obtusidentatum) and the four described species in the H. excisum subclade.
Characters | H. obtusidentatum | H. excisum | H. obscurum | H. pseudobscurum | H. tholiformis |
---|---|---|---|---|---|
Size of lamina | 15–30 × 8–15 cm | 20–40 × 10–15 cm | 20–30 × 5–10 cm | 20–25 × 5–10 cm | 13–16 × 3–5 cm |
Lamina base | truncate | truncate and widest | truncate | truncate | truncate |
Rhizome size | 2–4 mm in diam. | 3–5 mm in diam. | 3–5 mm in diam. | 3–5 mm in diam. | 2 mm in diam. |
Stipe color | not shiny, reddish brown to dark brown | shiny, dark purple to black | not shiny, dull green to grayish green | not shiny, dull green to grayish green | shiny, black purple |
Pinna shape | trapeziform to falcate | trapeziform to falcate | trapeziform to falcate | trapeziform to falcate | trapeziform |
Size of middle pinnae | 3.5–8 × 1–1.8 cm | 5–10 × 1.3–2 cm | 3.5–7 × 0.8–1.3 cm | 2.5–4 × 0.8–1.8 cm | 2.5–8 × 0.6–1 cm |
Shape of pinna apex | acute or rarely obtuse | acute to obtuse | obtuse to subacute | obtuse to subacute | round |
Pinnae marginal teeth | blunt or rounded | sharp | sharp to blunt | sharp to blunt | sharp |
Number of basiscopic veins lacking | 4–6 | (3 or) 4–6 | 3–5 | 3–5 | 3–4 |
Sori position | inframedial to medial | medial | medial | medial to supramedial | medial |
Indusium | single | single | single | double | single |
The total length of the concatenated rbcL, rps4-trnS, and trnH-psbA alignment was 2527 bp. The alignment contained 138 variable characters, 98 of which were parsimony informative. The three phylogenetic analyses, MP, ML, and BI, of the combined chloroplast dataset recovered similar topologies (Fig.
Maximum likelihood phylogeny based on the concatenated plastid DNA sequence dataset. Maximum parsimony and Bayesian analyses recovered identical topologies with respect to the relationships among the main clades of the paleotropical Hymenasplenium. For each node, the following values are provided: maximum parsimony bootstrap (%) / maximum likelihood bootstrap (%) / and posterior confidence (p-value). Columns on the right refer to the subclades described in
The H. excisum subclade recovered in paleotropical Hymenasplenium included several well-supported lineages but represented only four described species (Fig.
In the present study, the Hymenasplenium excisum subclade was found to be polyphyletic (Fig.
Phylogenetically, the monophyly of H. obtusidentatum is strongly supported by the chloroplast sequences analyses (Fig.
Though H. obtusidentatum is distributed together with H. excisum and is morphologically similar to H. excisum, this new taxon was found to be closely related to H. obscurum and H. pseudobscurum in the chloroplast phylogenetic analyses. Different ploidy levels, including diploid, tetraploid, and even hexaploid, are found in H. excisum, H. obscurum and H. pseudobscurum. Besides, reticulations and allopolyploidizations were assumed to occur in Hymenasplenium (
China. Yunnan Province, Xishuangbanna, Menghai: Chang1258. 2019. (holotype, HITBC; isotype, KUN) (Figs
Plants 25–40 cm tall. Rhizomes long creeping, 2–4 mm in diameter, apex scaly; scales brown, lanceolate to triangular, entire. Fronds remote, up to 6 mm apart, grayish green when dry, herbaceous; stipe reddish brown to dark brown, 10–20 cm long, base ca. 1–1.5 mm in diam., subglabrous, base with sparse scales similar to those on rhizome; lamina one-pinnate, narrowly triangular to triangular, 15–30 × 8–15 cm, base truncate and reduced, apex acuminate to caudate; rachis reddish brown to dark brown, subglabrous; pinnae 15–21 pairs, trapeziform to falcate, basal pinnae nearly opposite, middle pinnae 3.5–8 × 1–1.8 cm, dimidiate, apex obtuse to acute, base asymmetrical, acroscopic side truncate and often almost parallel to rachis, basiscopic side of basal pinnae excavate, in middle pinnae narrowly cuneate and entire, acroscopic margin serrate, teeth blunt or rounded; pinnae spreading to ascending. Veins forking and terminating in marginal teeth, basiscopic side with 4–6 veins lacking. Sori inframedial to medial, linear, 3–5 mm, indusia persistent, pale brown, membranous, entire, opening toward the costa. Spores elliptic to reniform, perispore fimbriate-alate, 38–43 μm in diam.; 64 spores per sporangium.
Hymenasplenium obtusidentatum is similar to H. excisum. However, H. obtusidentatum has a thinner rhizome and stipe, not shiny and reddish brown to dark brown stipe and rachis, blunt or rounded pinna marginal teeth, and truncate lamina base.
Hymenasplenium obtusidentatum is currently known to coexist with H. excisum in south and southwestern China, and adjacent areas. It occurs in soil or on rocks near streams in half-shaded forests at alt. 1000–1500 m.
China. Yunnan Province, Menghai County, 2019; Chang1256, Chang1260; Chang1261, Chang1262; Chang1339 (HITBC); Pu’er City, Pu’er Sun River National Forest Park, 2018; Xu 324 (SYS). Hainan Province, Ledong County, 2019; Chang1284, Chang1285; Chang1286 (HITBC).
We are grateful to the editor and reviewers for their helpful comments. We thank several colleagues who assisted with the collections, including Changle Zhang and Huafeng Hong in China. This work was supported by the following projects: National Natural Science Foundation of China (31500171) and Science Foundation of Hengyang Normal University (2021HSKFJJ032).
Voucher specimens and GenBank accession numbers for the DNA sequences used in this study. Information is presented in the following order: species, voucher, locality, GenBank numbers for rbcL , rps4 & rps4-trnS , trnH - psbA . “*” represents the newly published sequences in this study; “–” represents no data. Herbaria acronyms follow Index Herbariorum (
Hymenasplenium adiantifrons (Hayata) Viane & S. Y. Dong; Knapp 3904 (P); Taiwan, China; –, MH065323, –. H. apogamum (N. Murak. & Hatan.) N. Murak. & Hatan.; Chang1040 (HITBC); xishuangbanna, Yunnan, China; MH884808, MH884830, MH884838. H. apogamum; ChenCC1106 (HITBC); Taiwan, China; MH884813, MH884832, MH884839. H. cardiophyllum (Hance) Nakaike; Sysu 2; Cult. in Sun-yat Sen University; MH065387, MH065306, –. H. cheilosorum (Kunze ex Mettenius) Tagawa; Xu 102 (SYS); Hainan, China; MH065385, MH065346, –. H. cheilosorum; Xu 299 (SYS); Guangxi, China; MH065402, MH065350, –. H. cheilosorum; Xu 322 (SYS); Yunnan, China; MH065405, MH065352, –. H. cheilosorum; Xu GX022 (SYS); Guangxi, China; MH065381, MH065343, –. H. cheilosorum; Zhang et al. 8629 (CDBI, MO, PHH); Lam Dong, Vietnam; MH065415, MH065357, –. H. chingii K. W. Xu, Li Bing Zhang & W. B. Liao; Zhang et al. 9455 (CDBI); Guizhou, China; MH065430, MH065333, –. H. excisum (C. Presl) S. Lindsay; Brownsey & Perrie FIJI 190 (WELT); Fiji; KP774884, KP851882, –. H. excisum; Chang1045 (HITBC); Yunnan, China; OP375802*, OP375808*, –. H. excisum; Chang992 (HITBC); Yunnan, China; OP375801*, OP375807*, –. H. excisum; DJ0305; Kaaruu Creek, Queensland, Australia; KP774930, –, KP851883. H. excisum; Xu 274 (SYS); Guangdong, China; MH065389, –, –. H. excisum; KH17; Indonesia; GU586828, –, –. H. excisum; Ranker 1786 (COLO); Hawaii; AY549728, –, –. H. excisum; TNS:764169; Japan; AB574888, –, –. H. excisum; Xu 100 (SYS); Hainan, China; MH065383, MH065344, –. H. excisum; Xu 110 (SYS); Hainan, China; MH065384, MH065345, –. H. excisum; Xu 198 (SYS); Hainan, China; MH065394, MH065341, –. H. excisum; Xu 196 (SYS); Hainan, China; MH065390, –, –. H. excisum; Xu 197 (SYS); Hainan, China; MH065391, –, –. H. excisum; Zhang et al. 6714 (CDBI, MO, VNMN); Bac Kan, Vietnam; MH065436, MH065375, –. H. excisum; Zhang et al. 8039 (CDBI, MO, VNMN); Quang Nam, Vietnam; MH065419, MH065361, –. H. filipes (Copeland) Sugimoto, Murakami 596902 (TI); Kagoshima, Japan; U30605, –, –. H. filipes; Murakami 92-J012; Kagoshima, Japan; AB016176, –, –. H. filipes; Schuettpelz_1084 (HITBC); Taiwan, China; MW194197, –, –. H. furfuraceum (Ching) Viane & S. Y. Dong; Chang1072 (HITBC); Yunnan, China; MW194198, MW194221, MW194182. H. furfuraceum; Xu 304 (SYS); Yunnan, China; MH065406, MH065353, –. H. hastifolium Ke Wang Xu, Li Bing Zhang &W. B. Liao; Xu 282-1 (SYS); Guangxi, China; MH065398, MH065313, –. H. hondoense (N. Murak. & Hatan.) Nakaike; Chang1062 (HITBC); zhaotong,Yunnan, China; MH884814, MH884833, MH884840. H. laetum (Sw.) L. Regalado & Prada; Jones 1084 (TUR); KJ628715, –, –. H. laetum; NM N293; Ecuador; AB014707, –, –. H. laterepens N. Murak. & X. Cheng ex Y.Fen Chang & K. Hori; Chang1039 (HITBC); xishuangbanna, Yunnan, China; MH884807, MH884829; MH884837. H. latidens (Ching) Viane & S. Y. Dong; Chang1029 (HITBC); Yunnan, China; MW194204, MW194219, MW194180. H. latidens (Ching) Viane & S. Y. Dong; Xu 309 (SYS); Yunnan, China; MH065407, MH065318, –. H. murakami-hatanakae Nakaike; ChenCC1089 (HITBC); Taiwan, China; MH884823, –, –. H. neocaledonicum Li Bing Zhang & K. W. Xu; PerrieNC177 (WELT); New Caledonia; KP774896, KP851878, –. H. ngheanense Li Bing Zhang, K. W. Xu & N. T. Lu; Zhang et al. 6532 (CDBI, MO, VNMN); Phu Tho, Vietnam; MH065426, MH065331, –. H. obliquissimum (Hayata) Sugimoto; Iwatsuki et al. 94-V302; Hoang Lien Son, Vietnam; AB016187, –, –. H. obliquissimum; Murakami & J. Yokoyarna 94-T628; Chiang Mai, Thailand; AB016178, –, –. H. obscurum (Blume) Tagawa; Fraser-Jenkins30715; Bhutan; MH884826, MH884834, –. H. obscurum; Zhang et al. 7715 (CDBI, MO, VNMN); Thanh Hoa, Vietnam; MH065411, MH065354, –. H. obscurum; Wu et al. WS2176 (MO); Xiangkhoang, Laos; ON85986, ON859875, –. H. obscurum; Wu et al. WS2552 (MO); Louangphrabang, Laos; ON859870, ON859876, –. H. obtusidentatum Y.Fen Chang & G.Cheng Zhang; Chang1256 (HITBC); Yunnan, China; OP375803*, OP375809*, OP375813*. H. obtusidentatum; Chang1258 (HITBC); Yunnan, China; OP375804*, OP375810*, OP375814*. H. obtusidentatum; Chang1285 (HITBC); Hainan, China; OP375805*, OP375811*, OP375815*. H. obtusidentatum; Chang1339 (HITBC); Yunnan, China; OP375806*, OP375812*, OP375816*. H. perriei Li Bing Zhang & K. W. Xu; Brownsey & Perrie FIJI 13 (WELT); Fiji; KP774885, KP851880, –. H. phamhoanghoi Li Bing Zhang, K. W. Xu & T. T. Luong; Zhang et al. 8819 (CDBI, MO, PHH); Khanh Hoa, Vietnam; MH065432, MH065334, –. H. pseudobscurum Viane; Chang1010 (HITBC); Yunnan, China; MH884827, MH884835, MH884841. H. pseudobscurum; Chang1047 (HITBC); Yunnan, China; MH884828, MH884836, MH884842. H. pseudobscurum; Xu 004 (SYS); Hong Kong, China; MH065380, MH065342, –. H. queenslandicum Li Bing Zhang & K. W. Xu; Ohlsen 252 (MELU); Queensland, Australia; KP774849, KP851879, –. H. riparium (Liebm.) L. Regalado & Prada; NM and M. Grayum 281; Virgen del Socorro, Costa Rica; AB014708, –, –. H. sinense K. W. Xu, Li Bing Zhang & W. B. Liao; Xu 134 (SYS); Jiangxi, China; MH065388, MH065348, –. H. sinense; Xu PB001 (SYS); Yunnan, China; MH065386, MH065347, –. H. sp8; Xu 324 (SYS); Yunnan, China; MH065404, MH065351, –. H. subnormale (Copel.) Nakaike; Schuettpelz 874; Malay Peninsula; MH884824, –, –. H. tholiformis Liang Zhang, K. W. Xu & W. B. Ju; Zhang Liang 4781 (KUN); Tibet, China; ON859868, –, ON859874. H. triquetrum (N.Murak. & R.C.Moran) L.Regalado & Prada; L.Sylvestre 2208 (RB); Brazil; KT329398, −, –. H. unilaterale (Lam.) Hayata; Hemp A. 18 (BM); Kenya; AF240652, –, –. H. unilaterale; Hennequin S. R144 (BM, REU); Reunion; KF992497, –, –. H. unilaterale; RV8344; Reunion; GU929873, –, –. H. unilaterale; SH408; Mauritius; MH884825, –, –. H. vanuatuense Li Bing Zhang & K. W. Xu; Ohlsen 392 (MELU); Tanna, Vanuatu; KP774898, KP85188, –. H. wangpeishanii Li Bing Zhang & K. W. Xu; Zhang et al. EM03 (CDBI); Sichuan, China; MH065410, MH065333, –. H. wildii (F. M. Bailey) D. Ohlsen; DJO246 (MELU); Queensland, Australia; KP774927, KP851877, –. H. wuliangshanense (Ching) Viane & S. Y. Dong; Chang1102 (HITBC); Yunnan, China; MW194208, MW194224, MW194184. H. wuliangshanense; Chang1104 (HITBC); Yunnan, China; MW194209, MW194225, MW194185.