Magnified view of an individual carpel of Uvaria valderramensis sp. nov. densely covered with trichomes. (10x) 

Context in source publication

Context 1

... sequences were then aligned with the acquired sequences from GenBank using the MacClade v.4.0 software. Aligned sequences were then subjected to parsimony analysis using the PAUP* v.4.0b (Swofford, 2000). Phylogenetic trees were obtained using the following settings: heuristic searches, MULTREES option on, tree-bisections- reconnection (TBR) branch swapping, using 5000 random addition sequences. The consistency and retention index (Farris, 1989) was calculated to measure homoplasy. Statistical support was derived through Bootstrap (BS) analysis using the following settings: 10,000 replicates, MULTREES option off, nearest neighbour interchange (NNI) branch swapping, and five random addition sequences. Clades having a BS value above 90% were considered to be strongly supported. Phytochemical Analysis . Crude extracts were obtained by grinding dried leaves using a Wiley mill. Powdered leaves were added in dichloromethane (DCM) and methanol (MeOH) (1:1) then drained after 3 d. Crude extracts were added with distilled water then petroleum ether (1:1). The water layer was extracted then added with DCM (1:1) further forming two layers. Water layer then was separated and extracted with butane. Layers with petroleum ether, DCM and butanol were developed into a 9:1 of ethanol-acetic acid and MeOH. Developed thin layer chromatography (TLC) plates were separately prepared for Vannilin reagent (Petroleum ether layer), Dragendorff’s reagent (Butanol layer), and potassium ferricyanide (DCM layer). The spotted TLC plates were allowed to stand with the solvent until the chromatograms of the extracts were obtained. Sequence variation of the two cpDNA markers . A total of 70 taxa was included for both trn L-F and mat K sequence analyses. Five new sequences from each gene markers are published here for the first time. The trn L-F region ranged from 560 base pair (bp) ( Goniothalamus elegans ) to 955 bp (Melodorum fruticosum ) of the unaligned data set. Uvaria grandiflora (VI014) had a total of 891 bp, while the unknown Uvaria (VI012a, VI012b, VI017) yielded an average of 901 bp. As for mat K, the unaligned sequence ranged from 771 bp ( U. angolensis ) to 842 bp (G. cheliensis ). All newly sequenced Uvaria species yielded a mean base content of 840 bp. The aligned data matrices for the trn L-F and mat K yielded a total length of 1094 and 849 bp, respectively. Out of the length of the trn L-F, 164 (14.99%) were parsimoniously informative characters, compared to mat K which had 182 (21.43%). Despite the length of the trn L-F against mat K, the latter had provided more informative sites. In the heuristic search of both gene markers, trn L-F resulted to a total of 3050 parsimonious trees, while mat K produced a total of 1831 equally parsimonious trees (Table 1). Consistency and retention indices and tree length of the trees provided with both trn L-F and mat K are presented in Table 1. Phylogenetic analysis . The combined data matrix had a total of 70 taxa included in the analysis. Out of the 1943 bp, 346 (17.81%) are parsimoniously informative characters. The heuristic search excluding the uninformative characters resulted to a total of 4881 equally parsimonious trees with a total length of 662 steps (Table 1). Consistency and retention indices were computed to be 0.678 and 0.859, respectively (Table 1.) In the separate (not shown here but available from the corresponding author) and combined tree analyses (Fig.1), Uvaria is resolved to be monophyletic with strong support (BS = 91%). The collected Uvaria spp. from Valderrama, Antique were shown in the combined tree (Fig. 1) to be nested in the Southeast (SE) Asian Uvaria group. Sampled Uvaria grandiflora formed a subgroup together with U. flava, U. grandiflora U. hirsuta and U. calamistrata with a high support (BS = 95%). The collected divergent yellow-flowered Uvaria formed a subgroup of its own with a strong support (BS = 99%) and it is not directly related to U. grandiflora (Fig. 1). The molecular data support the identity of yellow-flowered Uvaria as a true member of the genus but was not identical to U. grandiflora. Morphology of the divergent Uvaria. Congruent to the molecular analysis the distinctive Uvaria species collected from Valderrama revealed morphological features distinct to the genus. Among these characteristics sensu Ke β ler (1993) and Zhou et al. (2009) are the following: stellate hairs, valvate sepals, partially or fully imbricate and subequal whorls of leathery petals. Furthermore, the presence of solitary, leaf-opposed inflorescences (Fig. 2a), numerous stamens and pistils (Fig. 2b), and fused sepals (Fig. 2c) are features that are shared by other Philippine Uvaria species. Among the Philippine species, the divergent Uvaria highly resembles U. panayensis on the basis of leaf shape, texture and venation pattern, and fruits (Fig. 3). However, the divergent Uvaria species is different by having ovate to obtuse sepals (Fig. 2c) and densely pubescent carpels (Fig. 4) in contrast to the reniform sepals and glabrous carpels of U. panayensis . With these distinctions, we propose a new name for the divergent Uvaria species, Uvaria valderramensis Cabuang, Exconde & Alejandro. Uvaria valderramensis Cabuang, Exconde & Alejandro sp. nov. (Fig. 5) Type: Philippines, Visayas, Province of Antique, Municipality of Valderrama, Barangay Bugnay (11 o 00’05.60”N; 122 o 07’45.58”E) 14 May, 2011, Cabuang P.G., Exconde B., Lemana B.O.C . VI011 (holotype, USTH!; isotypes, PNH! USTH!) Shrubs, ca. 3 to 7 m tall, branches cylindrical, greenish- brown with lenticels; young branches green. Leaves glabrous on the upper surface while sparsely pubescent along the midrib on the lower surface; subcoriaceous; blades ovate, 15-31 cm long, 6-13 cm wide; margin entire, apex acute, obtuse, or mostly acuminate; base slightly cordate; leaf venation brochidodromous with 15-18 (-21) secondary veins; petioles 6-11 mm. Inflorescence leaf-opposed, axillary, solitary or simple corymbs with 2 to 4 flowers per node. Flowers pedicellate, 1.0-3.5 cm long, 2.0-4.5 cm diameter. Sepals ovate to obtuse, united at the base. Outer petals ovate, 3 to 4 per flower, yellow, 1-2 cm long, 1.0-1.8 cm wide; inner petals 3-5 per flower somewhat smaller than the former. Stamens and carpels numerous, forming a round, globose structure. Anthers brown, basifixed, flattened, oblong to somewhat discoid in shape, and leathery. Carpel arranged in aggregates. Pseudostyle apical in position, cylindrical. Stigma dark brown, flattened, oblong, leathery, covered usually with nectar in their receptive pistillate phase. Fruit monocarps ca. 10-20+, torulose, with slight constrictions in between seeds, each segment obovoid in shape, yellow- green with a smooth, fleshy pericarp. Fruits 16-23 mm in length, 8.0-8.65 mm in diameter with 3-4 seeds. Seeds globose to obovoid, flattened, 6.0-6.5 mm in diameter, exotesta reddish brown. Notes. Uvaria valderramensis is closely allied to U. panayensis based on similarities in the leaf blade shape and corolla lobe shapes. The species is however different from U. panayensis by having a large ovate to obovate leaves, distinctly yellow corolla lobes, and densely pubescent carpels. Phenology . Flowering May to June and fruiting from July to November Distribution and Habitat . Endemic to the Municipality of Valderrama, Antique, Panay Island, Visayas, Philippines. In riverine forests, shaded areas, hanging on trees, in clay soils, 20-30 m away from Bugnay Elementary School. Local name. Usog (Kinaray-a) Etymology. The specific epithet is derived from the type locality. Additional specimens examined. Visayas: Province of Antique, Municipality of Valderrama, 14 May, 2011. Cabuang P.G., Exconde B., Lemana B.O.C. VI012, VI017. (syntypes, PNH!; USTH! <2 sheets>) Phytochemical analysis of U. valderramensis . The phytochemical assay on the leaf extracts of U. valderramensis showed the presence of phenols, alkaloids, flavonoids, steroids, tannins and terpenes. (Table 2) Several of these compounds found in the leaf extract had already been proven to yield various medicinal and biological properties. Triterpenes for instance were found to possess anticancer properties (Liby et al., 2007). Phenols and flavonoids can confer anti-oxidant properties on plants, while the latter also provide a strong potential anti-microbial agent (Aminimoghadamfarouj et al., 2011). Flavonoids have been proven to demonstrate anti-inflammatory properties (Okwu & Iroabuchi, 2008). The present molecular and morphological study confirmed the placement of both Uvaria grandiflora and the unknown Uvaria collected in Valderrama, Antique to be members of the genus Uvaria. Maximum parsimony analyses and the morphological features of the unknown Uvaria species corroborated the distinction of this species from the rest of the SE Asian species. With this, we proposed a new species, Uvaria valderramensis Cabuang, Exconde & Alejandro sp. nov. This new species of Uvaria has yielded several phytochemical ...