A new species of Eustigma (Hamamelidaceae) from Hon Ba Nature Reserve, Vietnam

Abstract A new species of Hamamelidaceae, Eustigma honbaense H.Toyama, Tagane & V.S.Dang, sp. nov., is described from Hon Ba Nature Reserve, Vietnam. This species is similar to Eustigma oblongifolium Gardner & Champ., but differs from it in having entire leaves, longer infructescences, capsules with a longer apical part and seeds with a larger hilum. A description, preliminary conservation assessment, illustration and photographs of the new species are provided, as well as an updated key to the genus Eustigma.


Introduction
Eustigma Gardner & Champ. (Gardner 1849) is a small genus of the Hamamelidaceae, distinguished from other genera by small auriculate petals and the greatly enlarged stigma (Harms 1930;Zhang et al. 2003). Currently, three species are known in the genus: E. balansae Oliv., E. lenticellatum C.Y.Wu and E. oblongifolium Gardner & Champ., distributed in mainland China, Laos, Taiwan and Vietnam, among which E. balansae is the only species recorded in Vietnam (Tardieu 1965;Hsieh 1993;Hô 2003;Zhang et al. 2003;Newman et al. 2007).
From 2013 to 2014, as part of a collaborative programme to document the biodiversity and ecology of Southeast Asia, Kyushu University (FU) together with Institute of Tropical Biology, Vietnam (VNM) carried out botanical fi eld surveys in Hon Ba Nature Reserve, in South Vietnam (Figure 1) and found a species of Eustigma that was distinct from any of the three known species. Here, we describe this plant as a new species, Eustigma honbaense H.Toyama, Tagane & V.S.Dang, and place the species within the wider generic context by providing an updated identifi cation key to all known species of Eustigma. Our conclusion is based on observations of specimens in the herbaria, BKF, E, HN, K, KAG, KYO, L, P, TI, and VNM, and specimen images on the website of JSTOR Global Plants (https://plants.jstor.org/). We also provide DNA sequences of 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); established protocols were used to determine the sequences of these regions (Kress et al. 2009;Dunning and Savolainen 2010 Diagnosis. Th is species is similar to Eustigma oblongifolium Gardner & Champ., but distinguished from that species by having entire elliptic to oblong leaves (vs. partly dentate oblong-lanceolate leaves), infructescences 5-10.3 cm long (vs. 3-5 cm long), apical part (above tepal scar) of capsules 6.5-9 mm long (vs. 3-5 mm long) and seed hilum 4 mm long and 3 mm wide for bigger side (vs. 2-3 mm long and 1-1.5 mm wide).
Other Distribution and habit. Th is species is only known from Hon Ba Nature Reserve of southern Vietnam. We found only three individuals in the evergreen forest along a stream at ca. 400 m alt. Th e fl ora of this area is reported in Choudhary et al. (2012), Schuiteman et al. (2013) and Tagane et al. (2015a, b).
Phenology. Flowering season is unknown. Immature fruits and capsules were observed in July and November.
Etymology. Eustigma honbaense is named after the type locality, Hon Ba Nature Reserve.
Preliminary conservation status. Eustigma honbaense was collected from a single locality in the Hon Ba Nature Reserve, where only three individuals (one reproductive tree and two young trees) were found in evergreen forest along a streamside. Th e forest around the habitat was frequently logged and disturbed. Th erefore, this species is assessed as Critically Endangered (CR) using the criterion D of the Red List Categories (IUCN 2012), although more individuals could be discovered by more thorough surveys.
Note. Eustigma honbaense has terminal naked buds and axillary scaled buds that are in contrast with the description of Eustigma having naked-buds as given in the Flora of China (Zhang et al. 2003). Th e naked bud is covered either with immature leaves that develop to foliage leaves or with their stipules, while the scaled bud is covered with cataphylls or stipules that are highly modifi ed to protect the shoot tip (Nitta and Ohsawa 1998). Th e terminal buds of E. honbaense are classifi ed as naked buds because they are incompletely covered by 2 stipules (Fig. 4A, B, E & F), while the axillary buds are classifi ed as scaled buds because they are covered by 2 cataphylls that are subsequently shed or remained at the tip of previous shoot without further growth (Fig. 4C, D, E & F). Th e terminal bud of Fig. 4B shows alternate leaf arrangement from the axillary bud, but same direction is also observed.
Th e description of bracts and bracteoles is insuffi cient because most of the bracts and bracteoles had been shed. Eustigma oblongifolium and E. lenticellatum have 3-bracteate fl owers (Gardner 1849;Hsieh 1993;Wu 1977), while E. balansae has 2-bracteate fl owers (Oliver 1891;Tardieu 1965). Th is could be a key trait to distinguish between species. Further collections of E. honbaense are needed.
Th e matK sequences between E. honbaense and E. oblongifolium (AF013043) diff er in 6 bases of the 781 total. Th e rbcL and matK sequences between E. honbaense and E. balansae (rbcL: HQ415214, matK: HQ415379) diff er in 1 base of the 517 total and 5 bases of the 761 total, respectively.  An updated key to the species of Eustigma