Revista de Biología Tropical ISSN: 0034-7744 [email protected] Universidad de Costa Rica Costa Rica de Viana, Marta L.; Giamminola, Eugenia; Russo, Roberta; Ciaccio, Mirella Morphology and genetics of Anadenanthera colubrina var. cebil (Fabaceae) tree from Salta (Northwestern Argentina) Revista de Biología Tropical, vol. 62, núm. 2, junio-, 2014, pp. 757-767 Universidad de Costa Rica San Pedro de Montes de Oca, Costa Rica Disponible en: http://www.redalyc.org/articulo.oa?id=44931383029 Cómo citar el artículo Número completo Sistema de Información Científica Más información del artículo Red de Revistas Científicas de América Latina, el Caribe, España y Portugal Página de la revista en redalyc.org Proyecto académico sin fines de lucro, desarrollado bajo la iniciativa de acceso abierto Morphology and genetics of Anadenanthera colubrina var. cebil (Fabaceae) tree from Salta (Northwestern Argentina) Marta L. de Viana1*, Eugenia Giamminola1, Roberta Russo2 & Mirella Ciaccio2 1. Banco de Germoplasma de Especies Nativas, Instituto de Ecología y Ambiente Humano. Universidad Nacional de Salta. Avda. Bolivia 5150, 4400, Salta, Argentina; [email protected], [email protected] 2. Istituto di Biomedicina e Immunologia Molecolare “A. Monroy”. Consiglio Nazionale delle Ricerche (CNR), Via Ugo La Malfa 153, 90146 Palermo, Italy; [email protected], [email protected] * Correspondence Received 13-VI-2013. Corrected 20-X-2013. Accepted 22-XI-2013. Abstract: Anadenanthera colubrina var. cebil is an important tree species for its cultural, economic, and medicinal uses in South America. In order to characterize A. colubrina populations, we collected fruits from four different sites (San Bernardo, El Cebilar, Metán and El Gallinato) within the species distribution area in Salta Province, Northwestern Argentina. For this, a total of 75 fruits and seeds per site were collected and described using morphological (fruits size and weight; seed weight and number per fruit) and genetic descriptors (ribo- somic DNA extraction and PCR; nucleotide alignment and phylogenetic analysis) with standard protocols. Our results showed that the San Bernardo population had the heaviest fruits and seeds (7.89±0.2g and 0.19±0.002, respectively), and the Cebilar population the lightest (6.25±0.18g and 0.15±0.002g, respectively). Fruits and seeds from Metán and El Gallinato showed similar and intermediate values. The proportion viable (39 to 55%) and aborted (43 to 57%) seeds was different, while the proportion of predated (1.7 to 4.2%) seeds was similar among populations. The genetic analysis showed variability of ITS sequences within the especies, and also when compared with the same Brazilian species. Both, morphologic and genetic descriptors showed a high level of similarity between San Bernardo and Metán, and between El Cebilar and El Gallinato populations. Further stud- ies are needed to assess levels of phenotypic and genetic variability within and between populations of different plant species, since this information is crucial for biodiversity and germplasm long-term conservation. Rev. Biol. Trop. 62 (2): 757-767. Epub 2014 June 01. Key words: biodiversity, germplasm bank, molecular markers, nucleotidic sequence, plant conservation. Anadenanthera colubrina var. cebil (Vell.) in diameter. The wood contains tannins, is Brenan belongs to the Fabaceae/Leguminosae hard and resistant to termites and is used in family (Lewis, Schrire, MacKinder, & Lock, construction and furniture, as poles and for 2005), Mimosoideae subfamily. This tree spe- firewood. In popular medicine, it is used to cies has economic, medicinal, and cultural treat respiratory problems and inflammations. applications in South America. It is widely It is considered a sacred tree by local cultures. distributed in Brazil, Paraguay, Bolivia and Its seeds have been used for over 3 000 years Argentina, where it is found in eleven prov- by shamans in rituals and popular medicine as inces in the Northeast, Northwest and center, they contain alkaloids derived from dimethyl in mountain and transition forests and in Chaco tryptamine (Carod-Artal & Vásquez-Cabrera, Serrano (Siri Von Reis, 1964). It measures 2007; Demaio, Karlin, & Medina, 2002; Jus- up to 35m in height and can reach 60-80cm tiniano & Fredericksen, 1998; Martinez & Rev. Biol. Trop. (Int. J. Trop. Biol. ISSN-0034-7744) Vol. 62 (2): 757-767, June 2014 757 Andrade, 2006; Monteiro et al., 2006). This Argentina (Barrandeguy, Prinz, García, & Fin- species is categorized as “least concern” with keldey, 2012). low risk of extinction (IUCN, 2012). Different molecular markers have been The characterization of germplasm collec- used to study plant genetic diversity, such as tions allows the assessment of the variability ISSR (Inter Simple Sequence Repeat) (Lom- of the preserved material, although this infor- bardo, Schicchi, Marino, & Palla, 2012). Other mation is poorly documented in seed banks. molecular markers of great importance are the When the characterizations are morphological, Intergenic Transcripted Spacer (ITS) regions descriptors that exhibit high heritability are of ribosomal rRNA (rRNA). ITS refers to a used due to their stability in different environ- piece of non-functional RNA situated between ments (de Viana, Morandini, Giamminola, & rRNAs. This polycistronic rRNA transcript Diaz, 2011; Souza & Sorrells, 1991). contains the 5’ external transcribed sequence Studies carried out on wild plant species (ETS), 18S rRNA, ITS1, 5.8S rRNA, ITS2, have revealed that the distribution of morpho- 28S rRNA and finally the 3’ ETS. Genes logical and chemical characteristics present encoding rRNA occur in tandem repeats that patterns can be related to geographic regions are in thousands of copies long, each separated by non-transcribed DNA regions termed inter- (Nevo, Beiles, & Krugman, 1988; Nevo, Noy- genic spacers (IGS) or non-transcribed spacers Meir, Beiles, Krugman, & Agami, 1991). These (NTS) (Rogers & Bendich, 1987). They have variations may reflect the phenotypic plasticity an important biological meaning in rRNA pro- of the individuals, genetic adaptations of the cessing and in RNAs during ribosome matura- populations to different environments or both tion (Hausner & Wang, 2005). In genomes, the (Bradshaw, 1984; Matthies; Schmid, & Smid- ITS regions vary greatly in size and sequence Hempel, 1995). For instance, Wulff (1986) (Korabecna, 2007). reported that the degree of differentiation in Sequence comparison of the ITS region populations of the genus Plantago was great- is widely used in taxonomy and molecular er for morphological than for isoenzymatic phylogeny. This can be explained by the rela- markers, due to its great phenotypic plastic- tively low evolutionary pressure acting on such ity in response to environmental differences. non-functional sequences. For example, ITS Makkar & Becker (1998) found that plants in has been proven especially useful for eluci- semiarid environments contain more phenolic dating relationships among congeneric spe- compounds than in humid African environ- cies and closely related genera in Asteraceae ments. Morphological differences were also and Cyperaceae (Baldwin, 1992; Shekhovtsov, reported in relation to genetic and environmen- Shekhovtsova, & Peltec, 2012). tal characteristics, such as soil moisture, nutri- The aim of this work was to study the mor- ent levels, precipitation, radiation, temperature phological descriptors and the genetic diversity and elevation (Baker, 1972; Kuiper, 1985; of A. colubrina var. cebil, collected from differ- Bond, Hoing, & Maze, 1999; Bu et al., 2007) ent locations over its distribution area in Salta and Monteiro et al (2006) reported a positive Province (North Argentina) and conserved relationship between A. colubrina bark tannin in the Germplasm Bank of Native Species content and environmental humidity. (BGEN-INEAH) of the National University of Morphological characteristics are related Salta by analyzing, for the first time, the ITS of to genetic traits, but this information is poorly rDNA region. documented. The genetic diversity of the Bra- zilian A. colubrina was studied using micro- MATERIALS AND METHODS satellite markers, and polymorphisms were found in the studied populations (Feres et al., Collection of fruits: Initially four popu- 2012), as well as in A. colubrina var. cebil in lations of A. colubrina var. cebil (Ac) were 758 Rev. Biol. Trop. (Int. J. Trop. Biol. ISSN-0034-7744) Vol. 62 (2): 757-767, June 2014 selected from different sites over the distribu- thickness) and weighed. The weight distribu- tion area of the species in Salta province: El tions of fruits and seeds were analyzed with Gallinato (G), San Bernardo (B), El Cebilar (C) the Shapiro - Wilks normality test as well as and Metán (M) (Fig. 1), all with different eleva- skewness and kurtosis tests. The morphological tion and rainfall (Table 1). From each popula- descriptors were compared between popula- tion, we collected mature fruits from 10 trees tions with ANOVA and analysis of conglomer- (separated by at least 30m from each other). ates (Euclidean distance) using InfoStat (2009). The fruits were processed in the INEAH labo- ratory, separating two groups: one for long term Isolation and sequencing of ITS-rDNA conservation in the seed bank, and the other for regions: Genomic DNA was extracted from morphological and genetic characterization. the seeds of one tree from each population following the protocol of Doyle & Doyle Morphological approach of fruits and (1990). PCR