Fine-Scale Genetic Structure and Gene Flow Within Costa Rican Populations

Heredity (2003) 90, 268–275 & 2003 Nature Publishing Group All rights reserved 0018-067X/03 $25.00 www.nature.com/hdy Fine-scale genetic structure and gene flow within Costa Rican populations of mahogany (Swietenia macrophylla) AJ Lowe1, B Jourde1, P Breyne2,4, N Colpaert2,5, C Navarro3, J Wilson1 and S Cavers1 1Centre for Ecology and Hydrology-Edinburgh, Bush Estate, Penicuik, Midlothian EH26 0QB, Scotland, UK; 2Vlaams Instituut voor Biotechnologie (VIB), Department of Plant Genetics, University of Gent, K. L. Ledeganckstraat 35, B-9000 Gent, Belgium; 3Centro Agrońomico Tropical de Investigacioń y Ensenãnza, Cartago, Turrialba 7170, Costa Rica Fine-scale structure of genetic diversity and gene flow were flow, via either pollen or seed, is occurring between blocks of analysed in three Costa Rican populations of mahogany, mahogany within a continuous or disturbed forest landscape. Swietenia macrophylla. Population differentiation estimated Thus, once diversity is removed from a forest population of using AFLPs and SSRs was low (38.3 and 24%) and only mahogany, these data suggest that recovery would be slightly higher than previous estimates for Central American difficult via seed or pollen dispersal, and provides an populations based on RAPD variation (20%). Significant fine- explanation for mahogany’s apparent susceptibility to the scale spatial structure was found in all of the surveyed pressures of logging. Evidence is reviewed from other mahogany populations and is probably strongly influenced by studies of gene flow and seedling regeneration to discuss the limited seed dispersal range of the species. Furthermore, alternative extraction strategies that may maintain diversity a survey of progeny arrays from selected mother trees in two or allow recovery of genetic resources. of the plots indicated that most pollinations involved Heredity (2003) 90, 268–275. doi:10.1038/sj.hdy.6800247 proximate trees. These data indicate that very little gene Keywords: AFLP; genetic structure; gene flow; mahogany; microsatellites; Swietenia macrophylla Introduction the southern Amazon basin in Brazil (Pennington, 1981), and inhabits both wet and dry forests (rainfall typically To maintain natural populations as evolutionarily viable 1000–2000 mm) and a range of soils up to 1400 m altitude units able to adapt to changing conditions in the long (Whitmore, 1983). Although population densities are term, genetic variation must exist. For selectively typically less than one individual per hectare (Barros et al, exploited tropical tree species, genetic erosion can 1992), up to eight trees per hectare have also been become an important consideration and sustainable recorded. Mahogany has unisexual, staminate and production can be threatened if genetically determined pistillate flowers (Styles and Khosla, 1976), which are wood quality or pest resistance characters are under- insect-pollinated (bees, moths and thrips), and have a mined (Newton et al, 1996). A genetic resource manage- functional self-incompatibility system (Bawa et al, 1985; ment strategy for such species needs to be based on Mayhew and Newton, 1998), making it an obligate research data examining the extent of genetic differentia- outbreeder. Although little information is available on tion within and between populations, and on under- phenology, flowering appears to be seasonal and standing the processes maintaining this variation. simultaneous in most trees within a population. Leaf Mahogany, Swietenia macrophylla King (Meliaceae, fall tends to occur at the onset of the dry season, leaving Swietenioideae), is one of the most important timber the mature seed capsules on the tree. Seed production species in the world (Rodan et al, 1992), and over- from mahogany, in both natural and plantation trees, exploitation and high rates of deforestation have led to it fluctuates considerably from year to year and may reflect becoming the focus of increased conservation concern variation in flowering phenology, or failure of pollination (Newton et al, 1996). Mahogany matures to a large, or fertilisation. The seeds are winged and dispersed by deciduous tree able to reach heights of 40 m, a diameter wind, but are relatively heavy, and tend to fall close to of over 2 m and may live for several centuries. It has a the maternal tree (32–36 m median distance). However, a wide distribution, stretching from southern Mexico to distance of over 80 m was estimated for seed dispersal in a Bolivian population (Gullison et al, 1996). Correspondence: AJ Lowe, Centre for Ecology and Hydrology-Edinburgh, In Meso-America deforestation rates are particularly Bush Estate, Penicuik, Midlothian EH26 0QB, Scotland, UK. rapid, where approximately 4% of forest is removed E-mail: [email protected] annually (Myers, 1989), but a large proportion of those 4Current address: Institute for Forestry and Game Management (IBW), Gaverstraat 4, B-9500 Geraardsbergen, Belgium forests that once contained mahogany has already been 5Current address: Institute Plant Biotechnology for Developing Countries, felled. A recent study using random amplified poly- University of Gent, K.L. Ledeganckstraat 35, B-9000 Gent, Belgium morphic DNA (RAPD) markers, found that 80% of the Mahogany gene diversity and dynamics Lowe et al 269 variation detected across Central American populations repeats, or SSRs, Goldstein and Schlo¨tterer, 1999), target of S. macrophylla was maintained within populations individual loci of short tandem DNA repeats. The (Gillies et al, 1999). This study also showed that a history codominant and multiallelic nature of SSRs make them of logging was correlated with a significant reduction in ideal for studying outcrossing rate and gene flow. population diversity. However, reasons for the popula- For the present study, three populations of S. macro- tion level decline in genetic diversity following logging phylla from northern Costa Rica were chosen, and were were not examined in detail. It is therefore a priority to among those previously analysed by Gillies et al (1999) examine the fine scale structure of genetic diversity and for RAPD variation. The aim of the investigation was to extent of gene flow within and between populations as a use AFLP analysis to examine the extent of fine-scale first step to understanding the potential impact of such structure within all populations. In addition, primer landscape changes. Without this knowledge, it will be pairs previously developed for microsatellite loci in impossible to implement sustainable utilisation pro- S. humilis, but known to amplify in S. macrophylla (White grammes for mahogany (Newton et al, 1999). and Powell, 1997), were used to assess outcrossing rate A range of powerful molecular genetic techniques, based and potential gene flow in progeny cohorts from two of on the polymerase chain reaction (PCR; Newton and the populations. A combination of AFLP and SSR data Graham, 1994), are available for studying biodiversity, will provide information on the population dynamics of genetic structure and mating systems. In particular, the S. macrophylla, and such investigations will contribute to amplified fragment length polymorphism (AFLP) techni- the development of an integrated strategy for the que (Vos et al, 1995) targets random loci from across the sustainable resource management of S. macrophylla, plant genome, which are in general dominantly expressed. which is an urgent priority. Owing to the large number of loci that can be screened using this technique, AFLPs are particularly applicable to Materials and methods studies of genomewide genetic diversity, structure and population differentiation. Another set of molecular All populations selected for this study are located in markers, microsatellites (also called simple sequence northern Costa Rica (Figure 1). Pocosol is separated from Figure 1 Map of Costa Rica showing location of populations. Inserts show detailed mapping positions of trees sampled from each population, open circles indicate those trees from which open pollinated progeny arrays were collected. Heredity Mahogany gene diversity and dynamics Lowe et al 270 the other two populations by an assemblage of moun- tion PCR was carried out using an EcoRI primer tains, the ‘Cordillera de Guanacaste’. Canõ Negro and (GACTGCGTACCAATTC) without additional nucleo- San Emilio are located in the wet region of Alajuela, east tides and an MseI primer (GATGAGTCCTGAGTAA) of the mountain range, while Pocosol is situated in the carrying two selective nucleotides (AT, AC, GC and CA). dry region of Guanacaste on the west coast. All Final selective PCR reactions used EcoRI primers with populations have suffered some form of disturbance in two nucleotides and MseI primers with four nucleotides their past. Canõ Negro has been intensively logged, and in the following combinations: Eco+GC/Mse+ATAG, most trees with superior form (ie tall, straight trunk) Eco+GT/Mse+ACCC, Eco+CG/Mse+GCGA, Eco+CG/ have been removed. The forest therefore only contains a Mse+ACCG, Eco+GC/ Mse+CACG. All oligos and low density of mahogany, but a few large trees remain in primers were obtained from Genset (Paris, France). The a disturbed forest landscape. San Emilio also has a EcoRI primers were end labelled with [g-33P] ATP and history of logging. A proportion of trees of superior form amplification products were visualised by polyacryla- have been removed, but the forest has been left to mide gel electrophoresis and exposure to Kodak X-ray regenerate and forms a continuous canopy containing film. clusters of regenerating mahogany. The Pocosol study PCR amplification reactions using SSR primers con- plot is located within a block of continuous forest with tained: 10 mM of each of dATP, dTTP, dCTP and dGTP; 1 intact canopy, but is partially surrounded by grazing unit of Taq polymerase (Promega); 1/10th volume 10 Â pasture. The Guanacasten name for the open woodland buffer (supplied with the enzyme); 200 pM microsatellite used for traditional grazing systems that characterise the primers (MAC49, 54 and 58 from White and Powell, Pocosol area are ‘sitios’, and it is likely that this area has 1997); 1–10 ng of DNA; and sterile, deionised water to a been exploited over a long period of time (J Cornelius, final volume of 25 ml.

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