Identification of endangered or threatened Costa Rican tree species ...

Identification of endangered or threatened Costa Rican tree species by wood anatomy and fluorescence activity Róger Moya1*, Michael C. Wiemann2 & Carlos Olivares3 1.

Instituto Tecnológico de Costa Rica, Escuela de Ingeniería Forestal, Apartado 159-7050, Cartago, Costa Rica; [email protected] 2. Forest Products Laboratory, USDA Forest Service, One Gifford Pinchot Drive Madison, Wisconsin 53726-2398, USA; [email protected] 3. Instituto Tecnológico de Costa Rica, Escuela de Ingeniería Forestal, Apartado 159-7050, Cartago, Costa Rica; [email protected] * Corresponding author Received 10-I-2012.

Corrected 20-X-2012.

Accepted 12-XI-2012.

Abstract: Identificación de especies de árboles en peligro o amenazadas de Costa Rica basada en la anatomía de la madera y fluorescencia. A total of 45 native Costa Rican tree species are threatened or in danger of extinction, but the Convention on International Trade Endangered Species (CITES) includes only eight of these in its Appendices. However, the identification of other species based on their wood anatomy is limited. The present study objective was to describe and to compare wood anatomy and fluorescence activity in some endangered or threatened species of Costa Rica. A total of 45 (22 endangered and 23 threatened with extinction) wood samples of these species, from the xylaria of the Instituto Tecnológico de Costa Rica and the Forest Products Laboratory in Madison, Wisconsin, were examined. Surface fluorescence was positive in eight species, water extract fluorescence was positive in six species and ethanol extract fluorescence was positive in 24 species. Almost all species were diffuse porous except for occasional (Cedrela odorata, C. fissilis, Cordia gerascanthus) or regular (C. salvadorensis and C. tonduzii) semi-ring porosity. A dendritic vessel arrangement was found in Sideroxylon capari, and pores were solitary in Guaiacum sanctum and Vantanea barbourii. Vessel element length was shortest in Guaiacum sanctum and longest in Humiriastrum guianensis, Minquartia guianensis and Vantanea barbourii. Finally, anatomical information and fluorescence activity were utilized to construct an identification key of species, in which fluorescence is a feature used in identification. Rev. Biol. Trop. 61 (3): 1133-1156. Epub 2013 September 01. Key words: fluorescence, Costa Rican woods, tropical woods, wood identification key.

For its size (51 000km2) Costa Rica is one of the most biodiverse countries in the world. Of its 10 000 plant taxa, 1 300 are endemic and more than 25% of these are considered rare (Burger 1980). Nonetheless, intensive deforestation began at the start of the twentieth century, and reached levels in excess of 50 000ha per year in the 1970s (Alvarez 1986). Natural forest decreased to only 24.4% by 1987 (Sánchez et al. 2001). Tree timber species receive the highest degree of protection, and two actions are used to protect them: the promulgation of laws and

government decrees to stop the cutting of 18 commercial species (Costa Rica 1997) protected by international conventions such as CITES, and actions limiting harvesting in certain areas of the country (Costa Rica 1996a, 1996b). A number of important timber-producing species have been identified by some experts as threatened species due to their rarity (Jimenez 1999). Identification of these timber-producing species using floral and tree characteristics has been widely described (Jimenez 1999). In contrast, identification of these species based on their wood anatomy is limited to only a few

Rev. Biol. Trop. (Int. J. Trop. Biol. ISSN-0034-7744) Vol. 61 (3): 1133-1156, September 2013

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species: examples are the species included in CITES Appendices I and II. The CITES Identification Guide-Tropical Wood (CITES 2002) has a general identification key with illustrations of the transverse sections of each species. The book is intended for non-experts (Gasson et al. 2010) and its use is limited. More recently, the International Association of Wood Anatomists (IAWA) published several wood descriptions of tree timbers included in CITES (Gasson 2011, Gasson et al. 2011). Recently, wood anatomical identification has been accompanied by other analyses that together with special methods, can achieve precise identification that are useful in identification of endangered or threatened species. Some of these techniques are relatively easy to implement, for example, wood density, surface fluorescence, or fluorescence of water or ethanol extracts. (Miller & Wiemann 2006, Guzman et al. 2008, Wiemann & Ruffinatto 2012). Studies of anatomical features accompanied with complex statistical procedures, such as the use of multivariate analysis, helps to identify CITES species (Gasson et al. 2010, MacLanchlan & Gasson 2010). The objective of the present study was to describe and compare wood anatomy, fluorescence, and density of endangered or threatened species of Costa Rica, and to use these characteristics to build an identification key. The key will allow monitoring of the use and commercialization of these tree species (endangered or threatened) in Costa Rica, and perhaps will be also useful to other countries in the region. MATERIAL AND METHODS Wood samples and permanent slides: A total 45 timber species growing in Costa Rica were analyzed from March to December of 2010, 22 of which are considered as endangered species by the Costa Rican Government (Costa Rica 1996a and 1996b, Costa Rica 1997) (Table 1). Another 23 timber species studied are considered to be in threat of extinction in this country (Jimenez 1999) (Table 2). Wood samples of all the species were obtained 1134

from the Forest Products Laboratory (FPL), Wisconsin-USA collection (MADw and SJRw) and the Instituto Tecnológico de Costa Rica (ITCR) collection (TECw). Their sample numbers are detailed in tables 1 and 2. In some cases only one sample and slide was available, so it was necessary to prepare additional permanent slides of these species. To do this, a block (1cm3) was obtained from a wood sample and was softened in hot water. Tangential, radial and transverse sections were cut (12-15μm thick). These sections were stained with safranin and dehydrated with a series of alcohol (5 minutes each in 50, 70 and 95%); finally, sections were rinsed and mounted on microscope slides. Furthermore, a small piece was cut from each wood block to prepare macerated wood using Franklin’s method (Ruzin 1999). Wood anatomical description: The IAWA list (IAWA 1989) was used as the basis for choosing identification characteristics, with some modifications to allow for increased accuracy and subsequent species level separation. The quantitative anatomical features that were measured were: length and diameter of the fibers, lumen diameter, cell wall thickness, vessel length, diameter and frequency of pores, solitary pore frequency, diameter of intervessel pits, and height and width of rays. Fiber dimensions and vessel lengths were measured on macerated wood. Permanent slides were used for measurement of the other anatomical characters. Qualitative anatomical features were also determined using the IAWA List as a guide (IAWA 1989). Additional information: Other important information about endangered or threatened species used was: endemic category if the species grows only in Costa Rica, wood density, traditional uses of species, and fluorescence. Wood density (weight/volume) was measured in the air-dry condition. The four traditional uses considered were light or heavy construction, flooring, furniture, and handicrafts. Heartwood fluorescence was observed directly the surface, in water extract, and in ethanol extract,



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Cordia gerascanthus L. Sclerolobium costaricense Zamora & Poveda Anthodiscus chocoensis Prance Caryodaphnopsis burgeri N. Zamora & Poveda Couratari scottmorii Prance Cedrela fissilis Vell. Cedrela salvadorensis Standl. Swietenia humilis Zucc. Swietenia macrophylla King Parkia pendula Benth. Dipteryx panamensis (Pittier) Record & Mell Hymenolobium mesoamericanum H.C. Lima Myroxylon balsamum (L.) Harms Paramachaerium gruberi Briz. Platymiscium curuense N. Zamora & Kiitgaard Platymiscium parviflorum Benth. Platymiscium pinnatum var. polystachyum (Jacq.) Dugand Platymiscium pinnatum (Jacq.) Dugand Podocarpus costaricensis de Laub. Podocarpus guatemalensis Standl. Podocarpus macrostachys Parl. Guaiacum sanctum L.

Species Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Gymnosperm Gymnosperm Gymnosperm Angiosperm

Wood species

Source: Law decree No. 23700-MINAE, No. 25167-MINAE and No. 25663-MINAE. Legend * Wood density was determined at 12% of moisture content. LC=Light construction, HC=Heavy construction, Fl=flooring, Fu=furniture and H=handicrafts.

Zygophyllaceae

Podocarpaceae

Mimosaceae Papilionaceae

Boraginaceae Caesalpiniaceae Caryocaraceae Lauraceae Lecythidaceae Meliaceae

Botanical Family No Yes Yes Yes Yes No No No No No No No No Yes Yes No Yes No Yes No No No

Endemic

Traditional used of wood Fu & LC HC & Fl HC HC HC Fu & H Fu & H Fu & H Fu & H LC HC & Fl LC HC & Fl HC Fu, Fl & H Fu, Fl & H Fu, Fl & H Fu, Fl & H Fu Fu Fu HC, Fl & H

TABLE 1 Endangered timber species in Costa Rica (decreed by government) Wood density* Wood samples Permanent (g/cm3) observed slides observed 0.72 6 6 0.74 1 1 0.88 1 1 0.78 1 1 0.79 2 1 0.54 7 7 0.56 2 2 0.75 6 6 0.60 7 12 0.59 7 5 0.85 1 3 0.70 1 1 0.91 7 7 0.82 2 2 0.78 1| 1 0.79 1 1 0.80 1 1 0.78 5 2 0.57 2 1 0.51 7 3 0.55 6 7 1.06 7 9

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Astronium graveolens Jacq. Tabebuia guayacan (Seem.) Hemsl. Copaifera aromatica Dwyer Copaifera camibar Poveda, N. Zamora & P.E. Sánchez Cynometra hemitomophylla (Donn. Sm.) Britton & Rose Mora oleifera (Triana) Ducke Peltogyne purpurea Pittier Prioria copaifera Griseb. Tachigali versicolor Standl. & L.O. Williams Caryocar costaricense Donn. Sm. Humiriastrum guianensis Cuatrec. Vantanea barbourii Standl. Oreomunnea pterocarpa Oerst. Couratari guianensis Aubl. Lecythis ampla Miers. Cedrela odorata L. Cedrela tonduzii C.DC. Minquartia guianensis Aubl. Dalbergia retusa Hemsl. Dussia macroprophyllata (Donn. Sm.) Harms Prumnopitys standleyi (Wild.) Ladd. Sideroxylon capari (A. DC.) Pittier Qualea paraensis Ducke

Species Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Angiosperm Gymnosperm Angiosperm Angiosperm

Wood species

Source: Jimenez 1999. Legend: * Wood density was determined at 12% of moisture content. LC=Light construction, HC=Heavy construction, Fl=flooring, Fu=furniture and H=handicrafts.

Podocarpaceae Sapotaceae Vochysiaceae

Olacaceae Papilionaceae

Meliaceae

Juglandaceae Lecythidaceae

Caryocaraceae Humiriaceae

Botanical Family Anacardiaceae Bignoniaceae Caesalpiniaceae No No No Yes No No No No No No No No Yes No No No No No No Yes Yes No No

Endemic

Traditional used of wood HC, Fl & H HC & Fl LC, Fl & H HC, Fl & Ha LC LC & Fl HC, Fl & H LC & Fl LC & Fl HC HC HC & Fl LC HC HC FU & H FU & H HC & Fl HC, Fl & H LC Fu HC & Fl LC & Fl

TABLE 2 Timber species considered to be in threat of extinction in Costa Rica Wood Density* (g/cm3) 0.95 1.01 0.68 0.78 0.78 0.75 0.98 0.48 0.64 0.72 0.66 0.91 0.53 0.69 0.78 0.44 0.44 0.89 1.01 0.59 0.63 0.79 0.37

Wood samples Permanent observed slides observed 8 9 7 5 3 2 1 1 2 2 1 1 7 3 6 9 1 2 6 2 2 1 6 1 3 8 5 1 5 1 7 9 9 2 1 6 7 6 1 3 3 1 1 1 2 8

as described in the IAWA List (IAWA 1989). Surface fluorescence was determined in a darkroom from freshly prepared (planed or scraped) transverse and/or longitudinal surfaces exposed to a low intensity, long wave ultraviolet light (around 365nm). The specimens were recorded as either fluorescente (noting color and intensity) or not fluorescente. Only specimens which exhibited a definite yellow, green, orange or blue fluorescence were recorded as fluorescente. For water and ethanol extracts, color and intensity of fluorescence were scored. Froth tests were conducted and were scored as positive (high intensity) if one minute after shaking vigorously, froth was present and covered the entire surface of the solution, negative if all froth had disappeared, and variable if froth was still present around the edge of the test tube but did not extend over the entire surface of the liquid column. RESULTS General aspects: 22% (ten species) were Papilionaceae, 18% (eight species) were Caesalpiniaceae, 13% (six species) were Meliaceae, 9% (four species) were Podocarpaceae, 7% (three species) were Lecythidaceae, 4% (two species) each were Caryocaraceae or Humiriaceae; other families represented the 27% (12 species) of endangered or threatened timber species endemic to Costa Rica. Most of these species are used in heavy or light construction or handicrafts; however, species in the Meliaceae are utilized for furniture manufacturing. High wood density was a characteristic of most of the species, and the main use of the lumber was construction (Table 1 and 2). Fluorescence test: Surface fluorescence was positive in 11 species. This represented 24% of the total number of species. Fluorescence in Caryocar costaricense, Copaifera aromatica, Astronium graveolens, Mora oleifera and Myroxylon balsamum was green, greenish, or weak green, whereas in Dussia macroprophyllata, Lecythis ampla, Hymenolobium mesoamericanum, Peltogyne purpurea

and Tachigali versicolor it was yellow, yellowish or weak yellow (Table 3). Water extract fluoresced in only nine species (Table 3). The fluorescence was yellow in Copaifera species, H. mesoamericanum and P. purpurea and green in Cordia gerascanthus, A. graveolens, M. balsamum and Platymiscium species (Table 3). C. gerascanthus and Platymiscium species did not show surface fluorescence, but they did show water extract fluorescence (Table 3). Ethanol extract fluorescence was found in 31 of the species. The fluorescence was greenish blue or yellow, weak or light green, purple, yellowish or bluish (Table 3). Several species (Cedrela odorata, Cedrela fissilis, Cedrela tonduzii, Couratari guianensis, Couratari scottmorii, Dalbergia retusa, Dipteryx panamensis, Guaiacum sanctum, Oreomunnea pterocarpa, Parkia pendula, Prioria copaifera, Qualea paraensis, Swietenia humilis, Swietenia macrophylla, Tabebuia guayacan and Vantanea barborii) had positive ethanol extract fluorescence, but negative surface or water extract fluorescence (Table 3). General features of endangered or threatened species: Porosity: Diffuse porosity was common in endangered or threatened species, being found in 39 species. Three species were both diffuse and semi-ring porous (C. odorata, C. fissilis and C. gerascanthus) and two species were semi-ring porous (Cedrela salvadorensis and C. tonduzii). Sideroxylon capari was unique with a radial or diagonal pore pattern (Fig. 1a). Almost all of the species had solitary pores and pore multiples. However, the percentage of solitary pores was higher than 85% in G. sanctum and V. barbourii, so these species can be classified as pores exclusively solitary. Higher frequency of multiple pores (>15 pores/mm2) was measured in three species: C. guianensis, P. purpurea and T. guayacan. Pores frequency was the highest in Guaiacum sanctum, and M. balsamo. Lower pore frequencies (1 440 380 306 134 194 323 67 360 400 480 >1 330 170 535 190 224 >1 202

RW 1-3, U (>) 1-3, 4-10 1-3 3-10 1-3 1-3 1-3, 4-10 1-3 4-10 1-3 4-10 1-3, 4-10 1-3 1-3 1-3 1-3 1-3 1 1-3, 4-10 1-3, U (>) 1-3 1-3, U (>) 1-3 1-3 1-3 1-3 1-3 4-10 1

Ray parenchyma RF RT 6.1 Ht 3.2 Ht 7.8 Ht 2.0 Ht 3.5 Ho-Ht 3.6 Ht 4.8 Ht 3.7 Ht 6.7 Ho-Ht 6.4 Ho-Ht 6.8 Ht 6.2 Ho-Ht 6.8 Ho-Ht 14.9 Ht 16.6 Ho-Ht 6.6 Ho 6.6 Ho-Ht 14.4 Ho 5.9 Ht 19.7 Ht 2.8 Ho-Ht 7.0 Ht 10.0 Ho 11.6 Ht 9.5 Ht 12.4 Ho 5.8 Ho 5.7 Ho 10.4 Ho

TABLE 4 Fiber or tracheid and ray parenchyma characteristics of species that are endangered or considered to be in threat of extinction in Costa Rica

RS + + ++ + + + ++

R (C) S(+) U+ P(-) U+P(-) U+ P(-) U+ P(-) U+ P(-) U+ P(-) U+ P(+) S(+) S(+) S(+) U+ P(-) U+ P(-) -


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FL 1.2 1.0 1.1 2.4 3.2 2.9 1.0 2.5 1.1 1.1 0.8 1.0 0.8 0.9 0.9 1.6

LD 21.5 18.4 19.9 31.5 35.4 26.5 21.8 31.4 18.3 14.6 17.0 18.8 21.0 6.2 17.7 33.1

WCT 3.6 4.4 4.9 6.3 3.1 4.6 2.9 5.7 4.4 2.7 8.9 2.9 3.6 3.8 3.1 6.8

Fibers or tracheids FC SF + -+ GTF SBD SBD SBD T T T BD T SBD SBD SBD SBD SBD SBD SBD BD

PFRT +

FS + -+ -+ + + + -+ -+ + -

RH 170 160 160 71 55 111 550 90 260, >1 250 224 330 147 190 227 650

RW 1 1 1 1 1 1 1-3 1 1-3 1 1-3 1-3, 4-10 1-3, 4-10 1-3 1-3 1-3

Ray parenchyma RF RT 8.9 Ho 6.0 Ho 9.6 Ho 4.2 Ho 5.6 Ho 7.7 Ho 5.9 Ht 4.7 Ho 4.1 Ho-Ht 11.0 Ho 12.1 Ht 7.0 Ht 8.1 Ht 9.5 Ho 8.2 Ho 11.1 Ht

RS + + + +++ -

R (C) S(+)v U+ P(-) U+ P(-) U+ (-) S(+) -

Fiber: FL=Fiber or tracheids length (mm), LD=lumen or tracheids diameter (µm), WCT=wall cell thick (µm), FC=Fibers crystals, SF=Septate fibers, GTF=ground tissue fiber, SBD=simple to minutely bordered pits, BD=Fibers with distinctly bordered pit, T=tracheids, PFRT=Fiber pits common in both radial and tangential, FS=Fibers stored. Ray parenchyma: RH=ray height (µm), RW=ray width cell (µm), U(>)=mainly uniseriate, RF=ray frequency (ray/mm), RT=ray type, Ht=heterogeneous, Ho=homogenous, RS=ray stored, R(C)=crystals presence in ray, U=crystals in upright cells, P=procumbent cells, S=Silica bodies presence, “-“ anatomical feature absent, “+”anatomical feature present.

Platymiscium parviflorum Platymiscium pinnatum P. pinnatum var. polystachyum Podocarpus costaricensis Podocarpus guatemalensis Podocarpus macrostachys Prioria copaifera Prumnopitys standleyi Qualea paraensis Sclerolobium costaricense Sideroxylon capari Swietenia humilis Swietenia macrophylla Tabebuia guayacan Tachigali versicolor Vantanea barbourii

Spcies

TABLE 4 (Continued) Fiber or tracheid and ray parenchyma characteristics of species that are endangered or considered to be in threat of extinction in Costa Rica

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Anthodiscus chocoensis Astronium graveolens Caryocar costaricense Caryodaphnopsis burgeri Cedrela odorata Cedrela fissilis Cedrela salvadorensis Cedrela tonduzii Copaifera aromatica Copaifera camibar Cordia gerascanthus Couratari guianensis Couratari scott-morii Cynometra hemitomophylla Dalbergia retusa Dipteryx panamensis Dussia macroprophyllata Guaiacum sanctum H. mesoamericanum Humiriastrum guianensis Lecythis ampla. Minquartia guianensis Mora oleifera Myroxylon balsamum Oreomunnea pterocarpa Paramachaerium gruberi Parkia pendula Peltogyne purpurea Platymiscium curuense

AP + + + + + + + + + + + + + -

Type Diff Diff-Agg Diff Diff Diff Diff Diff Diff Diff-Agg Diff-Agg Diff Diff-Agg Diff -

Axial parenchyma Parenchyma arrangement Banded Parenchyma Vc Sca & Vc Mar Sca Sca Nar Sca-Vc Mar Sca-Vc-Loz Mar Vc-Con Mar Sca & Vc Mar Vc Mar Vc, Loz-alif & Con Wid Sca, Vc & Con Mar Wid-Nar-Ret-Scal Nar-Ret-Scal Nar-Ret-Mar Sca & Vc Nar-Mar Loz-alif, win-alif & Uni Mar Con Wid Sca Nar Loz-alif, Con, Vc Wid Vc, Loz-alif, Win-alif & Uni Wid-Ret-Mar Sca Vc, Loz-alif, win-alif & Con Mar Sca, Vc, Con & Uni Nar-Ret-Mar Sca, Loz-alif & wing-alif Nar Loz-alif & Con Loz-alif & Con & Uni Mar Loz-alif & Con Nar Stored + ves + ves + wae + wae + ves + wae +- wae + wae + ves -

Cell >8 3-8 3-8 3-8 3-8 3-8 3-8 3-8 3-8 3-8 2 & 3-4 3-8 3-8 5-8 2 & 3-4 2 & 3-4 3-8 & >8 2 2 & 3-4 3-8 & >8 3-8 5-8 3-5 2, 3-4 5-8 2 3-5 3-4 2

AP (C) + (+) + (+) + (-+) + (-+) + (-+) + (-+) + (+) + (+) + (+) + (+), S(+) + (+) + (+) + (+) + (+) + (+) + (+) + (+) + (+) + (+) + (+) + (+) + (+) + (-+) + (-+) + (+) + (+)

Radial canals

Radial canals

Others anatomical features

Crystals in enlarged cells

Vasicentric tracheids

Traumatic canals

Axial & ray canals & sheath cells Axial & ray canals & sheath cells

TABLE 5 Fiber, ray parenchyma and axial parenchyma characteristics of endangered and threatened species in Costa Rica


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Type Diff Sca Diff Sca Diff Diff-Agg Diff Diff Diff

Axial parenchyma Parenchyma arrangement Banded Parenchyma Stored Cell Loz-alif & Con Nar + ves 2 Loz-alif & Con Nar + ves 2 Loz-alif & Con Nar + ves 2