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Research Article
Aporosa tetragona Tagane & V. S. Dang (Phyllanthaceae), a new species from Mt. Hon Ba, Vietnam
expand article infoShuichiro Tagane, Van Song Dang§, Hironori Toyama, Akiyo Naiki|, Hidetoshi Nagamasu, Tetsukazu Yahara#, Hop Tran¤
‡ Kyushu University, Fukuoka, Japan
§ The VNM Herbarium, Institute of Tropical Biology, Ho Chi Minh, Vietnam
| Ryukyu University, Ishigaki, Japan
¶ The Kyoto Univesrity Museum, Kyoto, Japan
# Center for Asian Conservation Ecology, Kyushu University, Fukuoka, Japan
¤ University of Science Ho Chi Minh City, Ho Chi Minh, Vietnam
Open Access

Abstract

A new species, Aporosa tetragona Tagane & V. S. Dang, sp. nov., is described and illustrated from Mt. Hon Ba located in the Khanh Hoa Province, South Vietnam. This species is characterized by tetragonal pistillate flowers and fruits, which are clearly distinguishable from the other previously known species of the genus. The morphology and phylogeny based on rbcL and matK of this species indicated that the new species belongs to section Appendiculatae Pax & K. Hoffm.

Keywords

Aporosa, Hon Ba Nature Reserve, new species, Phyllanthaceae, Vietnam

Introduction

The genus Aporosa Blume (1825) (Phyllanthaceae) comprises 82 species of small to medium sized trees distributed in various environments in South and Southeast Asia. They can be found in both primary and secondary forests, from lowland rain to dry deciduous, and as well as montane forest (up to 2200 m altitude in New Guinea) (Gagnepain 1927, Hô 2003, Schot 2004, Schot and van Welzen 2005). The genus is characterized by a dioecy, indumentum of simple hairs, petioles pulvinate at both base and apex, two glands on adaxial base of lamina (occasional), often with scattered disk-like glands on lower surface of lamina, axillary inflorescences, absence of petals, styles and disks, tiny staminate flowers with a minute or absent pistillode, and dehiscent regmata with persistent stigmas, sometimes beaked and/or stiped (Pax and Hoffmann 1922, Schot 2004).

The recent revision (Schot 2004) classified the genus Aporosa into five sections based on morphological analyses: sect. Aporosa, sect. Appendiculatae Pax & K. Hoffm., sect. Benthamianae Schot, sect. Papuanae Schot and sect. Sundanenses Schot. In Vietnam, two sections including 11 species with two varieties of Aporosa are recorded (Gagnepain 1927, Hô 2003, Schot 2004). Nine species with two varieties are included in Aporosa sect. Appendiculatae: A. ficifolia Baill., A. macrophylla Müll. Arg., A. octandra var. octandra (Buch.-Ham. ex D. Don) A. R.Vickery (synonym: A. dioica Müll. Arg. and A. oblonga Müll. Arg.), A. octandra var. malesiana Schot (synonym, A. microcalyx (Hassk.) Hassk.), A. planchoniana Baill., A. serrata Gagnep., A. tetrapleura Hance, A. villosa Baill. (synonym: A. sphaerosperma Gagnep.), A. wallichii Hook. f. and A. yunnanensis (Pax & K. Hoffm.) F. P. Metcalf. Two species are included in Aporosa sect. Sundanenses: A. duthieana King ex Pax & K. Hoffm. and A. microstachya Müll. Arg.

During a botanical survey of Mt. Hon Ba in Khanh Hoa Province, South Vietnam in 2014, an undescribed species of Aporosa sect. Appendiculatae was found at the margin of a broad-leaved evergreen forest near a stream, at 200–400 m altitude. Here, we describe and illustrate this plant as a new species, Aporosa tetragona Tagane & V. S. Dang.

In addition to the morphological examination, DNA sequences and phylogenetic analysis are extremely helpful for delimiting species (Hebert and Gregory 2005, Dick and Webb 2012). Here, we sequenced two DNA barcode regions, the partial genes for the large subunit ribulose-1,5-bisphosphate carboxylase oxygenase (rbcL) and maturase K (matK) (CBOL Plant Working Group 2009) to compare with related taxa.

Materials and methods

Morphological observations

The new species was recognized by detailed comparisons with morphologically similar species through literature review, dry specimens from the herbaria ANDA, BK, BKF, BO, HN, KYO, SING, TNS, VNM, and digitized plant specimens available on the web (e.g. JSTOR Global Plants (https://plants.jstor.org/)).

DNA barcoding

Total DNA was extracted from silica-gel dried leaves collected in the field. DNA extraction was performed by a modified CTAB protocol (Doyle and Doyle 1987), as described in detail in Toyama et al. (2015). Amplification and sequencing of the two DNA barcodes regions, rbcL and matK, were performed according to published protocols (Kress et al. 2009, Dunning and Savolainen 2010).

Phylogenetic analysis

In total, 22 accessions representing 14 species of Aporosa were included in phylogenetic analyses using DNA barcoding regions of rbcL (362 bp) and matK (392 bp) (Table 1). In addition to the new species, Aporosa tetragona Tagane & V. S. Dang, four species, A. aurea Hook. f., A. microstachya (Tul.) Müll. Arg., A. penangensis (Ridl.) Airy Shaw and A. tetrapleura Hance, were newly sequenced in the present study. The remaining sequences were obtained from GenBank. Phyllanthus bokorensis Tagane was used as an outgroup. The sequence alignment was performed by ClustalW with default parameter implemented in MEGA v 6.06 (Tamura et al. 2013).

List of taxa used in this study with vouchers and GenBank accession number.

Section Species Vouchers GenBank accession no.
rbcL matK
Sect. Aporosa Aporosa frutescens Blume BT0095962054 KJ594599 KJ708827
Sect. Appendiculatae Aporosa aurea Hook. f. Tagane et al. T4249, FU LC089033 LC089037
Aporosa ficifolia Baill. KYUM:5 AB925289 AB924682
Aporosa octandra var. octandra (Buch.-Ham. ex D. Don)
A. R.Vickery
SCBG007-1 KP094163 KP093256
Aporosa octandra var. octandra (Buch.-Ham. ex D. Don)
A. R.Vickery
SCBG007-2 KP094164 KP093257
Aporosa planchoniana Baill. ex Müll. Arg. KYUM:315 AB925549 AB924927
Aporosa planchoniana Baill. ex Müll. Arg. KYUM:945 AB925759 AB925129
Aporosa planchoniana Baill. ex Müll. Arg. KYUM:29 AB925313 AB924702
Aporosa tetrapleura Hance Toyama et al. 1426, FU LC089030 LC089034
Aporosa tetragona Tagane & V. S. Dang Tagane et al. V1976, FU LC050338 LC050339
Aporosa villosa (Lindl.) Baill. KYUM:994 AB925783 AB925152
Aporosa villosa (Lindl.) Baill. KYUM:127 AB925406 AB924795
Aporosa yunnanensis (Pax & K.Hoffm.) F. P. Metcalf J578 KR528750 KR530383
Aporosa yunnanensis (Pax & K.Hoffm.) F. P. Metcalf BB0195 KR528747 KR530380
Aporosa yunnanensis (Pax & K.Hoffm.) F. P. Metcalf G202 KR528748 KR530381
Aporosa yunnanensis (Pax & K.Hoffm.) F. P. Metcalf BB0194 KR528746 KR530379
Sect. Benthamianae Aporosa benthamiana Hook. f. BT0070230656 KJ594594 KJ708826
Aporosa lunata (Miq.) Kurz BT0070234186 KJ594600 KJ708829
Sect. Sundanenses Aporosa microstachya (Tul.) Müll. Arg. BT0070234330 KJ594602 KJ708830
Aporosa microstachya (Tul.) Müll. Arg. Tagane et al. T4172, FU LC089032 LC089036
Aporosa penangensis (Ridl.) Airy Shaw Tagane et al. T4012, FU LC089031 LC089035
Sect. Papuanae Aporosa papuana Pax & K. Hoffm. Damas 004, KYO AB233915 AB233811
Outgroup Phyllanthus bokorensis Tagane Toyama et al. 1740, FU AB936022 AB936023

The Neighbor-joining methods (Saitou and Nei 1987) with Maximum Composite Likelihood distance matrix (Tamura et al. 2004) implemented in MEGA v 6.06 was used to construct the phylogenic trees. Confidence values for individual branches were determined by bootstrap analysis with 10,000 repeated samplings of the data.

Results and discussion

The new species belongs to the section Appendiculatae as the leaf lamina has basal adaxial glands (Fig. 2C), disc-like glands unevenly scattered within the arches of the marginal veins throughout the abaxial surface of the lamina (Fig. 2B), papillate stigmas (Fig. 2D), and pubescent septae and column in the ovary (Fig. 2E) (Schot 2004), but is distinguished from previously known species by its tetragonal ovary of the pistillate flower and the fruit.

Figure 1.

Aporosa tetragona sp. nov. A Leafy branch B Fruits and portion of abaxial surface of young leave C Apical bud and pulvinate petiole at both base and apex D Pistillate inflorescence E Fruits F, G Transverse section of fruits H Seeds taken from fruits. Materials: Toyama et al. V1976.

Figure 2.

Aporosa tetragona sp. nov. A Fruiting branch B Schematic of the placement of the disc-like glands on the lower side of the leaf C Apex of branch D Pistillate inflorescence E Longitudinal section of fruits. Materials: AC, E from Toyama et al. V1976 (KYO), D from Toyama et al. V829 (FU).

The Neighbor-joining tree based on rbcL and matK supports the separation of each morphologically defined section and the monophyly of sect. Appendiculatae, sect. Benthamianae and sect. Sundanenses with 98 %, 76 % and 85 % bootstrap probability, respectively (Fig. 3). The new species was placed in sect. Appendiculatae and clearly separated from other species of this section with a sister relationship to the clade including A. ficifolia, A. odctandra var. octandra, A. planchoniana and A. villosa.

Figure 3.

NJ tree of Aporosa species based on rbcL and matK. Branches are labeled with bootstrap support (% of 10,000 replicates).

Thus, the species is morphologically distinct from known taxa and the phylogeny supports the separation from related species. Here, we define the new species Aporosa tetragona Tagane & V. S. Dang.

Taxonomy

Aporosa tetragona Tagane & V. S. Dang, sp. nov.

Figs 1, 2

Diagnosis

Aporosa tetragona is distinct from all other Aporosa species by having a tetragonal ovary and fruit. The leaves are similar to Aporosa acuminata Thwaites, but differing in not only fruit shape but also wider leaves (vs. 2–4 cm wide), larger pistillate flowers (vs. 2–3.5 mm long), and glabrous fruits (vs. sparsely puberulous).

Type

VIETNAM. Khanh Hoa Province, Mt. Hon Ba, edge of evergreen forest near stream, 12°06'30.60"N, 108°59'15.70"E, alt. 393 m, 22 November 2014, with female fl. and fr., Toyama H., Tagane S., Dang. V. S., Nagamasu H., Naiki A., Tran H., Yang C. J. V1976 (holotype KYO!, isotypes BKF!, FU!, K, NTU!, P, VNM!, the herbarium of Hon Ba Nature Reserve!).

Description

Small tree, 3 m tall. Twigs glabrous, young branchlets green in vivo, dull yellowish green to pale yellow in sicco, old branchlets light grayish brown. Stipules caducous, not seen. Leaves: petiole 0.8–1.7 cm long, sunken above, rounded below, pulvini distinct, glabrous; blade ovate to elliptic, (6.8–)9–16.5 × 3.9–7.0 cm, length/width ratio 2.0–2.9, chartaceous to subcoriaceus, completely glabrous, dull yellowish green to dull pale yellow above and beneath in sicco, base cuneate to rounded, or shallowly subcordate, basal glands present, margin entire, foliar glands abaxially scattered mostly within the arches of the marginal veins, apex acuminate, acumen up to 2.3 mm long; midrib prominent on both surfaces, or rarely sunken only on the upper surface, secondary veins 10–14 pairs, raised on the lower surface, tertiary veins reticulate, visible on both surfaces of young leaves in sicco, inconspicuous on lower surface of old leaves. Staminate inflorescences not seen. Pistillate inflorescences in axils of leaves near the top of branchlets, solitary, flowers up to 7, rachis 2–5 mm long, densely pubescent; bracts broadly triangular, ca. 1 × 1.1 mm, margin ciliate, very sparsely pubescent outside, glabrous inside. Pistillate flowers (6–)8–10 mm long, (1.8–)2.5–3 mm in diam., sessile, yellowish in vivo, reddish brown in sicco; sepals 4, triangular, 0.8–1.1 × 1.1 mm, glabrous to very sparsely hairy outside, glabrous inside except near base, margin ciliate; ovary obclavate, 5–9 mm long, tetragonal, 2-locular, glabrous outside; ovules 2 per locule; stigmas slightly raised, elongated, ascending from the top of the ovary, stigma bilobed, lobes ca. 0.6–1 mm long, each stigma lobe apically deeply bifid, papillate and hairless above, smooth and very sparsely hairy beneath, style remnant present. Fruits tetragonal ellipsoid with sharp ridges, 21–25 × 7–9 mm, stiped, beaked, fleshy, reddish in vivo, pinkish orange to reddish brown in sicco, glabrous; septae and column pubescent with hairs of 0.4–0.6 mm long. Seeds 2 or 3, ellipsoid, flattened, ca. 9.0 × 5.0 × 3.5–4 mm, covered by fleshy, yellow aril in vivo, yellowish brown in sicco.

Other specimen examined

Vietnam. Khanh Hoa Province, Mt. Hon Ba, in evergreen forest near river, 12°06'33.41"N, 108°59'24.89"E, alt. 367 m, 19 Feb. 2014, with female fl., Toyama et al. V829 (FU!, VNM!, the herbarium of Hon Ba Nature Reserve!).

Phenology

Flowering specimens were collected in July and November; fruiting in November.

Distribution and habitat

This species is currently known only from Hon Ba Nature Reserve, Khanh Hoa Province, South Vietnam. The small populations were found at the edge of humid broad-leaved evergreen forest close to a stream, altitude 200–400 m.

Etymology

The specific epithet tetragona reflects the quadrangular shape of the ovaries in the pistillate flowers and fruits.

GenBank accession No

Toyama et al. V1976: LC050338 (rbcL), LC050339 (matK).

Conservation status

The species is known only from the type locality in Mt. Hon Ba at 200–400 m altitude. It is suggested that Aporosa tetragona should be placed under the IUCN category ‘Critically Endangered’ (CR) (IUCN 2012) because of its limited distribution with an area of occupancy estimated to be less than 10 km2 (criterion B2 a) and a small number of individuals estimated to be less than 250. Recent botanical inventories carried out in this narrow area along stream discovered several new species, including Dillenia tetrapetala Joongku Lee, T. B. Tran & R. K. Choudhary (Choudhary et al. 2012), Goniothalamus flagellistylus Tagane & V. S. Dang (Tagane et al. 2015) and Vanilla atropogon Schuit., Aver. & Rybková (Schuiteman et al. 2012), all of which are rare and endemic to Mt. Hon Ba. Therefore further collection efforts around this area are necessary to accurately understand the flora there and to update the conservation status of the species.

Acknowledgements

Authors cordially thank the staff of the Hon Ba Nature Reserve for permitting and supporting our field surveys in the protected area. We are grateful to Keiko Mase for her help with DNA barcoding, and to the curators and staff of the following herbaria: ANDA, BK, BKF, BO, HN, KYO, SING, TNS, VNM for allowing us to examine their collections. The present study was supported by the Environment Research and Technology Development Fund (S9) of the Ministry of the Environment, Japan and was partially supported by JSPS KAKENHI Grant Number 15H02640.

References

  • Blume C (1825) Bijdragen tot de Flora van Nederlandsch Indië : 514. Lands drukkerij, Batavia.
  • CBOL Plant Working Group (2009) A DNA barcode for land plants. Proceedings of the National Academy of Sciences of the United States of America 106: 12794–12797. doi: 10.1073/pnas.0905845106
  • Choudhary RK, Bach TT, Nong LV, Hai DV, Quang BH, Lee YM, Park SH, Lee C, Lee J (2012) Dillenia tetrapetala (Dilleniaceae), a new species from Hon Ba Nature Reserve, Vietnam. Annales Botanici Fennici 49: 369–376. doi: 10.5735/085.049.0608
  • Dick CW, Webb CO (2012) Plant DNA barcodes, taxonomic management, and species discovery in tropical forests. In: Kress WJ, Erickson DL (Eds) DNA Barcodes. Springer, New York, NY. Methods and Protocols 858: 379–393. doi: 10.1007/978-1-61779-591-6_18
  • Doyle JJ, Doyle JL (1987) A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochemical Bulletin 19: 11–15.
  • Dunning LT, Savolainen V (2010) Broad-scale amplification of matK for DNA barcoding plants, a technical note. Botanical Journal of the Linnean Society 164: 1–9. doi: 10.1111/j.1095-8339.2010.01071.x
  • Gagnepain F (1927) Aporosa. In: Lecomte MH, Gagnepain F (Eds) Flore générale de l’Indo-Chine. Masson, Paris, 5: 552–563.
  • Hebert PDN, Gregory TR (2005) The promise of DNA barcoding for taxonomy. Systematic Biology 54: 852–859. doi: 10.1080/10635150500354886
  • Hô PH (2003) Cay Co Viet Nam: An Illustrated Flora of Vietnam Vol. 2. Montreal. Published by the author, 951 pp.
  • IUCN (2012) IUCN red list categories and criteria, version 3.1. IUCN Species Survival Commission, Gland/Cambridge, 1–32.
  • Kress WJ, Erickson DL, Jones FA, Swenson NG, Perez R, Sanjur O, Bermingham E (2009) Plant DNA barcodes and a community phylogeny of a tropical forest dynamics plot in Panama. Proceedings of the National Academy of Sciences of the United States of America 106(44): 18621–18626. doi: 10.1073/pnas.0909820106
  • Pax F, Hoffmann K (1922) EuphorbiaceaePhyllanthoideaePhyllantheae. In: Engler A (Ed.) Das Pflanzenreich 4, 147, 15: 80–105. Leipzig.
  • Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution 4: 406–425.
  • Schot AM (2004) Systematics of Aporosa (Euphorbiaceae). Blumea Suppl. 17: 1–377. Nationaal Herbarium Nederland.
  • Schot AM, van Welzen PC (2005) Aporosa. In: Chayamarit K, van Welzen PC (Eds) Euphorbiaceae (Genera A-F). Flora of Thailand 8(1): 1–303.
  • Schuiteman A, Averyanov L, Rybková R (2013) Vanilla atropogon, a new species from Vietnam. Orchideen Journal 1: 10–16.
  • Tagane S, Dang VS, Yahara T, Toyama H, Tran H (2015) Goniothalamus flagellistylus Tagane & Dang VS (Annonaceae), a new species from Mt. Hon Ba, Vietnam. PhytoKeys 50: 1–8. doi: 10.3897/phytokeys.50.4427
  • Tamura K, Nei M, Kumar S (2004) Prospects for inferring very large phylogenies by using the neighbor-joining method. Proceedings of the National Academy of Sciences of the United States of America 101: 11030–11035. doi: 10.1073/pnas.0404206101
  • Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: Molecular Evolutionary Genetics Analysis Version 6.0. Molecular Biology and Evolution 30: 2725–2729. doi: 10.1093/molbev/mst197
  • Toyama H, Kajisa T, Tagane S, Mase K, Chhang P, Samreth V, Ma V, Sokh H, Ichihasi R, Onoda Y, Mizoue N, Yahara T (2015) Effects of logging and recruitment on community phylogenetic structure in 32 permanent forest plots of Kampong Thom, Cambodia. Philosophical Transactions of the Royal Society B: Biological Sciences 370(1662): 20140008. doi: 10.1098/rstb.2014.0008