Philippine Journal of Science 145 (3): 259-269, September 2016 ISSN 0031 - 7683 Date Received: ?? Feb 20?? Xerophytic Characteristics of Tectona philippinensis Benth. & Hook. f. Jonathan O. Hernandez1, Pastor L. Malabrigo Jr.1, Marilyn O. Quimado1*, Lerma SJ. Maldia1, and Edwino S. Fernando1 1Department of Forest Biological Sciences, College of Forestry and Natural Resources, University of the Philippines Los Baños, College, Laguna Tectona philippinensis Benth. & Hook.f. is one of only three species in the genus Tectona (Lamiaceae) restricted to the Asian tropics. It is endemic to Ilin Island and Batangas Province on Luzon Island, Philippines and is regarded as a critically endangered species. While role of xerophytic characteristics of plants are very important for their survival and growth under various environmental pressures, such characteristics in native tree species remain unclear. In this study, the anatomy of the species was analyzed to determine the xerophytic characteristics of T. philippinensis. Histological paraffin technique was used to examine the anatomical structures of leaf and young stem of the species. The anatomical structures of T. philippinensis have the characteristics typical of xerophytic plants. This includes the presence of four types of trichomes, extended and well-developed vascular system, and multiple layers of palisade and sclerenchyma cells. Extension of extended vascular bundles to both non-glandular hairs on the adaxial surface and glandular hairs on the abaxial surface of leaf is reported for the first time in this study. Therefore, anatomical structures of this species suggest its ability to survive under marginal conditions. However, studies on ecophysiology, pot experiments/field trials, phenology, and associated vegetation of the species are suggested to further understand its habitat preference and adaptation mechanisms. Key words: anatomy, arid or semi-arid, endemic, Lamiaceae, restoration, xerophytes. INTRODUCTION T. philippinensis is known only from Ilin Island and Batangas Province on Luzon Island, usually along dry The genus Tectona L.f. (Lamiaceae) includes only three hills and exposed limestone ridges along the coasts and species of trees restricted to the Asian tropics, viz., is also deciduous (Caringal et al. 2015). It is commonly Tectona grandis L.f. occurring in India, Laos, Mynamar, called Philippine teak, but is also known locally by the and Thailand; Tectona hamiltoniana Wall., endemic to vernacular names malabayabas and bunglas. The species Myanmar; and Tectona philippinensis Benth. & Hook.f., is regarded as critically endangered (Fernando et al. 2008, endemic to the Philippines. T. hamiltoniana occurs in the Madulid et al. 2008). The few remaining populations central dry zone of Myanmar (Kiyono et al. 2007; Aye et have been reported to be threatened by habitat destruction al. 2014), while T. grandis is known from a wider range through land conversion and development. Significant of climatic conditions, including dry areas, throughout its conservation efforts of the species include the Biodiversity natural range (Kaosa-ard 1981; Gyi & Tint 1998). Both Management Bureau (BMB) initiated project on ex-situ these species are known to be deciduous trees. conservation areas for the Philippine teak (PAWB-DENR *Corresponding author: [email protected] 259 Philippine Journal of Science Hernandez et al.: Morpho-anatomy of Tectona Vol. 145 No. 3, September 2016 philippinensis Leaf and Stem 1998) and non-government organizations and academe apical portion of an orthotropic branch. For leaf sample, initiated certain in-situ conservation strategies (Agoo & a small piece measuring 1mm2 was transversely cut in Oyong 2008). the median to include the midrib. For stem sample, on the other hand, approximately 1-2 mm long was also Many anatomical characteristics have been recognized transversely cut along the main axis of the stem using as protective mechanisms that allow the plants to survive a sharp Gillete razor blade. The illustrations of these against various levels of environmental pressure. For procedures are presented in Figure 1. example, the seven types of trichomes and their density through in vivo leaves of T. grandis were linked to extreme Histological paraffin technique was used (Johansen 1940) dependence of the species, especially those young ones, (Figure 2). Samples were fixed in 1:1 mixture of FAA-A for storing water during the developmental stage. In (12ml 37% Formaldehyde, 88ml 95% Ethanol) and vitro leaves, on the other hand, due to poor development FAA-B (10ml Glacial Acetic Acid, 88ml, and 90ml water) of epidermal structures (e.g. trichomes) were reported for three weeks. They were dehydrated following series of to have higher water loss than those of in in vivo leaves solutions of water, ethyl, and tertiary butyl alcohols from (Bandyopadhyay et al. 2004). Stephanou & Manetas 50% to 100% for four days. Gradual infiltration followed (1997) reported the features of leaves enable plants to using a 1:1 mixture of paraffin oil and tertiary butyl tolerate adverse conditions in the site such as drought, high alcohol for three days in the oven at 650C. Embedding the air temperature, UV-B radiation, among others. Plants that samples in the melted condition of paraffin wax followed. are well-adapted to such conditions commonly referred The samples were then mounted into 1.5cm x 1.5cm x 2cm to as xerophytes exhibit certain adaptive mechanisms to wooden blocks. Mounted samples were sectioned using a complete their life cycle in dry environments (Atia et al. rotary microtome (American Optical 820) at a thickness of 2014). They have special modifications such as leaves 10µm. Cross sections were mounted on microscope slides that are trichomous, with thick cuticle (Richardson & coated with Haupt’s solution, air-dried for three days, and Berlyn 2002), high palisade tissue/spongy tissue ratio, stained with 1% Safranin and were counter stained with and well developed water-storing and water-transporting 0. 5% Fast green. tissues to minimize the rate of transpiration. Many of the species in the family Lamiaceae have long been reported Microscopic examination and analysis to have xerophytic characteristics such as in the case of The typologies of anatomical structures were identified Salvia sclarea L. (Ozdemir & Senel 1999), Teucrium following the manual on anatomy of dicot plants. The montanum L. and Teucrium polium L. (Dinç et al. 2011). thicknesses of all visible dermal, ground, and vascular There is no report yet on xerophytic characteristics of tissues were measured. Characteristics of other structures T. philippinensis. This study analysed the anatomical such as stomata and trichomes were also examined. structures (leaf and young stem) of T. philippinensis to determine the species’ xerophytic characteristics. All the cross sections obtained were observed under a compound microscope (Euromex 0112987, manufacturer: BlueLine Holland) equipped with a camera which was connected to a desktop computer. The scale of all the MATERIALS AND METHODS measurements was calibrated at 40x magnifications. The mean thicknesses of the observed anatomical Place and duration of the study structures for both species were calculated using some The anatomical examination of leaf and stem was functions in MS Excel. Comparison of anatomical conducted at the Microtechnique Laboratory of the structures between T. philippinensis and C. ramiflora Department of Forest Biological Sciences (DFBS), was made. College of Forestry and Natural Resources (CFNR), University of the Philippines Los Baños (UPLB) from August to October 2015. RESULTS Preparation of specimens Three sample replicates for each leaf and stem of T. Stem philippinensis were collected from Lobo, Batangas, The stem of T. philippinensis is six-angled (Figure 3) and located at 400 masl. Samples of a non-xerophytic plant, its surface is occupied with glandular trichomes – capitate, Cynometra ramiflora L. were collected from Arbor peltate, and branched (Figure 5). The hypodermis is four Square, CFNR - UPLB. The leaf and/or stem sample for to six-layered of collenchyma cells. The rest of the cortex both species was obtained from c.a. 6-8 cm long from the is composed of 591.2µm thick, oval to round parenchyma 260 Philippine Journal of Science Hernandez et al.: Morpho-anatomy of Tectona Vol. 145 No. 3, September 2016 philippinensis Leaf and Stem Figure 1. Young leaf and stem of an orthotropic branch as the plant material of the study showing (a) length of sample used (b) part of leaf where the samples were obtained (c) size of cross section samples put inside the microcentrifuge tubes and (d) size of stem samples put inside the microcentrifuge tubes. Figure 2. Procedures of paraffin technique used in this study showing (a) fixation (b) dehydration (c) infiltration (d) embedding (e) microtoming (f) mounting on slide (g) staining (h) microscopic examination which were conducted at Microtechnique Laboratory of CFNR-UPLB. cells with intercellular spaces. The vascular tissue is of The pith enclosed by the vascular cylinder is built up collateral bundle type, measuring 1504.2µm thick, where of round and polygonal parenchymatous cells and 4-5 the xylem is of endarch configuration (Figure 3). Xylem clumps of compactly arranged thick walled sclerenchyma measures 383.0µm. Xylem fibres and xylem parenchyma cells (Figure 3). were also present. The phloem cells are small, polygonal, measuring 323.6µm in thickness. There are 2-3 layers The mean thickness of each of