Bot. Bull. Acad. Sin. (2003) 44: 329-336

Souto and Premoli — Divergence among A. aurea populations

Genetic divergence among natural populations of Alstroemeria aurea D. Don: A dominant clonal herb of the understory in subalpine Nothofagus forests

Cintia P. Souto* and A.C. Premoli

Laboratorio Ecotono, Centro Regional Universitario Bariloche, Universidad Nacional del Comahue, Quintral 1250, 8400 Bariloche, Argentina

(Received October 2, 2002; Accepted May 27, 2003)

Abstract. Alstroemeria aurea is a perennial herb with clonal rhizomatous growth, insect pollination, and ballistic seed dispersal that inhabits a range of different environments in the southern Andes. We evaluated the hypothesis that differential selective pressures act together with restricted among-population gene flow result in genetically divergent populations of A. aurea. Study sites were located within Nahuel Huapi National Park (41°8'S, 71°19'W) at two mountain ranges and two different elevations. These were Chall Huaco Valley, a pristine site, with populations at 1,250 and 1,100 m, respectively, and Cerro Otto, a disturbed site, with populations at 1,250 and 950 m, respectively. Seeds at each of the four populations were harvested from 20, 1 × 1 m plots. These seeds were counted, weighed, and germinated after a cold and humid stratification for 4 months. Within- and between-mountain ranges differences in seed traits were evaluated by ANOVA. We genetically characterized 30 plants of each population by allozyme electrophoresis and estimated levels of genetic variation and divergence. Seed traits showed different responses to elevation and site conditions. Total number of seeds was greater at low-elevation populations even though they had a higher number of undeveloped seeds. Reduced seed yield at high-elevation populations may result from a short growing season at higher altitudes. Additionally, seed weight, germination rates, and early vegetative spread were significantly greater at Otto, which may suggest a selective strategy to colonize disturbed sites under favorable physical conditions. These between-site differences were supported by allozyme data. High genetic divergence, and thus low gene flow, was estimated among Otto and Chall Huaco whereas within each mountain range among-population divergence depended upon site characteristics. Higher gene flow rates were found in the disturbed site Otto. Our results indicate that restricted pollen and seed dispersal, together with selective forces acting in different habitats, may produce genetic differentiation in populations of A. aurea.

Keywords: Alstroemeriaceae; Allozyme electrophoresis; Gene flow; Genetic structure; Germination; Patagonia; Seeds.


The ability of a plant species to occupy different environments could be the result of genetic adaptation, environmentally induced phenotypic plasticity, or a combination of these two processes (Jain, 1976). The relative importance of environment and genotype varies in the expression of a character. For example, differences in plant structure, leaf shape, and flowering time are often due to environmental factors while differences in pathogen resistance, flower architecture, and color are largely the result of genetic factors (Levin, 1984). Despite environmental influences, the genetic makeup of populations will directly affect their phenotypic characteristics such as the amount and kind of phenotypic variation, plant function including its interaction with the environment, and ecological tolerances of individuals together with their variation in time and space (Levin, 1984).

The amount and nature of genetic variation within and among populations is strongly affected by the mating system (Hamrick and Godt, 1990) and by the spatial relationship between plants and their parents (Hamrick and Nason, 1996). Populations where mating and seed dispersal occur over substantial spatial distances will have very different genetic properties as well as distinct ecological strategies from those in which these processes are restricted in space. Although random mating is a common assumption in many ecological and evolutionary models, it is unlikely to occur in plants, particularly when insect pollinators have restricted foraging behavior, such as bees tending to move from a plant to its near neighbors (Free, 1970; Levin and Kerster, 1974). In addition, for species with restricted seed dispersal, most progeny are established near their parents (Levin, 1984; Sobrevila, 1988; Redmond et al., 1989). Thus, the more restricted the dispersal of seeds and pollen, the stronger becomes the inverse relationship between distance and genetic kinship (Waser and Price, 1991; Souto et al., 2002). Spatial patterns of genetic variation for morphological, physiological, or phenological traits are the result not only of localized pollen and seed dispersal but also of different selection

*Corresponding author. Fax: +54-2944-422111; Phone: +54-2944-428505 Int. 508; E-mail:

Botanical Bulletin of Academia Sinica, Vol. 44, 2003

regimes under variable environments (Bazzaz, 1996). The objective of the present study was to analyze the degree of among-population genetic divergence in Alstroemeria aurea, a clonal species with restricted pollen movement and seed dispersal that inhabits different habitat types in northwestern Patagonia.

Material and Methods

The Species

Alstroemeria aurea, D. Don (Alstroemeriaceae) is a perennial herb that is geographically distributed in Argentina and Chile from 36° to 44° S latitude (Sanso, 1996). Outside its natural range, Alstroemeria aurea is used in breeding programs for cut flowers and pot plants. This species inhabits different plant communities ranging from woodlands, where mean annual precipitation of messic Nothofagus forests can exceed 4,000 mm, to the dry steppe with only c. 150 mm (Dimitri, 1972). It was suggested that the ability of A. aurea to occupy dissimilar habitats was environmentally induced, allowing different phenotypes to colonize areas ranging from pristine and successionally advanced to those altered by human and natural disturbances (Puntieri, 1991). This plasticity was related to its capacity to translocate available resources to different plant parts and, in particular, to its below ground storage organs, allowing the species to persist and establish itself as soon as environmental conditions are favorable (Puntieri, 1991).

In Northern Patagonia, Argentina, populations of A. aurea reach maximum size in high altitude forests of Nothofagus pumilio (southern beech), the dominant and, sometimes, nearly exclusive species of the understory. In these forests, Alstroemeria aurea is distributed between 750 and 1,250 m. Along this elevation gradient, different environmental variables—temperature, light intensity, wind exposure, and the amount and persistence of snow cover (Crawley, 1991)—are expected to affect the length of the growing season and, consequently, establishment and growth of A. aurea.

Alstroemeria aurea is a self-compatible herbaceous plant that reproduces vegetatively by clonal rhizomatous growth and sexually by seeds. Clone identification in the field is often difficult due to the continuous establishment of new individuals via seeds in patches already established (Puntieri, 1991). Using isozyme markers, it was estimated that only 17% of the ramets separated by 1 m belong to the same clone, suggesting a high degree of interdigitation among different genets (Souto et al., 2002). The main pollinators are bumblebees, such as the native Bombus dahlbomii (Apidae) and the exotic species B. ruderatus. Secondary in importance are the bee Apis melifera (Apidae) and a few other flies of the genus Tricophtalma sp. (Nemestrinidae) (Aizen, 1997). Average flight distances between consecutive flower visits of any of these pollinators do not exceed 1 m (Souto, 1999). Flowers that are successfully pollinated mature into capsules with explosive seed dehiscence (Aizen and Basilio, 1995).

In this study we tested the hypothesis that populations of A. aurea have a localized genetic structure and significant among-population genetic divergence. These would be the result of different evolutionary processes, such as restricted pollen movement, local seed dispersal, and the ability to propagate vegetatively, together with distinct selective regimes acting in different habitats. Seed characteristics were used as adaptive features to measure differential responses to elevation and site conditions. We used isozyme markers to estimate genetic divergence, useful as an indirect estimator of among-population gene flow.

Study Sites

Fieldwork was carried out at two mountain ranges within Nahuel Huapi National Park where A. aurea is the dominant species of the understory beneath pure N. pumilio forests. These are Chall Huaco Valley, a pristine site, and Cerro Otto, a disturbed site, located at 41°15'S / 71°18'W and 41°08'S / 71°21'W latitude/longitude, respectively. In each mountain range, two populations were chosen at different elevations: Chall Huaco high and low (CH and CL) located at 1,250 and 1,100 m, respectively, and Otto high and low (OH and OL) at 1,250 and 950 m, respectively.

Seed Characteristics

At each population all mature fruits were harvested from each of 20 randomly selected plots (1 × 1 m). Fruits were stored under dark and dry conditions in labeled paper bags until their natural dehiscence. Seeds were then separated from their capsules and for each plot, we quantified the total number of seeds and the number of undeveloped seeds, those which were empty, poorly developed, and/or insect-attacked. Seeds were weighted to ± 0.1 mg in groups of 5 with 10 replicates per plot (N = 1000 seeds/population). To break dormancy, seeds were cold stratified at 4°C using a bed of humid cotton inside plastic bags following Premoli et al. (2000) for 4 months. They were germinated using soil collected from naturally occurring populations of A. aurea, assuring that no seeds of this species were present in the soil at the time of sowing. Germination capacity was evaluated at weekly intervals by quantifying the number of emerging shoots over a 150-day period. After 200 days from the initiation of the germination experiment, given that the number of sprouts exceeded the initial number of seeds, new emerging shoots were scored as the number of ramets, including both new emergence as well as possible vegetative spread at the seedling stage.

Seed and seedling characteristics of each plot (dependent variables) were analyzed in relation to elevation and mountain range (independent variables) by two-way ANOVA. Each of these independent variables had two levels: high and low elevation as well as disturbed and pristine, corresponding, respectively, to the Otto and Chall Huaco locations. Germination capacity and total number of ramets in different populations was analyzed by ANCOVA models using the number of pre-treated seeds as a covariate. For further comparisons of populations'

Souto and Premoli — Divergence among A. aurea populations

values, we statistically analyzed seed characteristics by the Tukey post-hoc test for multiple comparisons of means, adjusting probability levels by the Bonferroni test (Rice, 1989).

Isozyme Analysis

Genetic characteristics of different populations were analyzed by horizontal isozyme electrophoresis. Fresh leaf material (approximately 1 g) was harvested from 30 randomly selected seedlings grown from seeds collected at each of the four study populations. We extracted enzymes by grinding the leaf material with the buffer of Mitton et al. (1979). Homogenates were stored at -80°C until electrophoresis on 12% w/v starch gels. Seven enzymes coding for 14 putative genetic loci were resolved using two gel and buffer systems that were reliably scored in previous studies (Souto, 1999; Souto et al., 2002). These were: Isocitrate dehydrogenese (Idh), Malate dehydrogenase (Mdh-1, Mdh-2), Phosphoglucoisomerase (Pgi-1, Pgi-2), and 6-Phosphogluconate dehydrogenase (6Pgd) on the Morpholine-citrate system (MC) of Ranker et al. (1989) and Malic enzyme (Me-1, Me-2), Menadione reductase (Mnr), and Shikimate dehydrogenase (Skdh) on the Tris-histidine system (HC) by King and Dancik (1983). Electrophoresis was carried out at 4ºC with an ice bag on top of the gel until a bromophenol blue die migrated approximately 10 cm from the origin towards the anode. Staining schedules for particular enzymes followed standard procedures (Soltis et al., 1983). Alleles were sequentially numbered, with the lowest number assigned to the most anodal allozyme.

Isozyme data were used to calculate the following parameters of genetic variability at the population level: mean number of alleles per locus (A), the percentage of polymorphic loci (P) using the 95% criterion, and the observed (HO) and expected heterozygosity (HE). Isozyme differences among populations were analyzed by Nei's genetic identity (1978), which represents the proportion of genes that are alike between and within populations. In order to graphically portray genetic similarities among all populations, average linkage clustering using the unweighted pair-group method (UPGMA) was performed for all populations using modified Rogers distance coefficient (Wright, 1978).

Genetic diversity was analyzed following Nei (1973) by the total diversity (HT), which can be partitioned within (HS), and the proportion of the total genetic diversity found

among populations (GST). These parameters were estimated for polymorphic loci. The degree of among-population divergence was measured by Wright's FST, based on polymorphic loci using FSTAT v. 2.9.1. (Goudet, 2000). Means and confidence intervals (CI95%) for FST were computed by jackknifing and bootstrapping over polymorphic loci, respectively, following Weir and Cockerham (1984). FST was calculated among different mountain ranges and was compared with the value obtained for populations within each mountain range.


Seed Characteristics

We found marked differences in seed characteristics among A. aurea natural populations studied. Within each mountain range, the total number of seeds produced per plot was significantly different for populations at different elevations. At both sites, high-elevation populations produced fewer seeds than low-elevation populations (Table 1). This pattern, consistent for Otto and Chall Huaco, resulted in non significant between-site differences (Table 2). Non significant effects, i.e. elevation, site or the interaction between them, were recorded for either the number of undeveloped seeds or seed weight. However, one-way ANOVA indicated that, within each mountain range, low-elevation populations had a greater number of undeveloped seeds, particularly at Otto (Table 1).

The Otto populations' seeds were significantly heavier than those at Chall Huaco (Table 1). In addition, significant site effects were recorded for the germination capacity measured after 150 d. as well as for the total growth measured as the number of ramets after 200 d. at each mountain range. Populations at Otto had significantly greater germination rates as well as a higher number of emerging seedlings and thus greater early vegetative spread than Chall Huaco populations (Tables 1 and 2). However, non significant differences with elevation were recorded for both variables within each mountain range (Table 1).

Genetic Analysis

Isozyme analysis showed that 5 out of a total of 14 resolved loci were polymorphic in at least one population. Overall, the analyzed populations were similar to each

Botanical Bulletin of Academia Sinica, Vol. 44, 2003

other in the levels of genetic variation they contained (Table 3). However, high-elevation populations showed a slightly greater mean number of alleles/locus (Otto population, OH) and higher polymorphism and heterozygosity (Chall Huaco population, CH). Genetic identities indicated that populations from the same location were more alike (Otto 0.999 and Chall Huaco 0.983) than were populations from different sites (average of all possible among-site comparisons 0.960; Table 4). This same pattern was obtained in the cluster analysis, where populations were grouped according to their location (Figure 1).

Elevated total genetic diversity was recorded in the studied populations, with HT = 0.310, most of which was partitioned within populations (HS=0.265 and GST=14.4 %). Divergence among mountain ranges yielded an FST value of 0.288 (95% confidence interval estimated by jacknifing 0.006-0.510). These values correspond to Nm = 1-FST/4 FST of about 0.6 migrants per generation, which means limited gene flow rates and thus elevated genetic divergence (Slatkin, 1985) among different mountain ranges. However,

Figure 1. Cluster analysis of the genetic relationships among populations of A. aurea using unweighted pair group method UPGMA based on modified Rogers distance coefficient (Wright, 1978). OH= Otto high; OL= Otto low; CH= Chall Huaco high; CL= Chall Huaco low populations.

greater gene flow rates were indirectly estimated by FST values within each mountain range. For Chall Huaco FST resulted 0.154 (CI95% = -0.012-0.223), corresponding to a gene flow rate of 1.4 migrants per generation, while FST of Otto populations was 0.011 (CI95% = -0.009-0.025), equivalent to 22.5 migrants per generation.


Isozyme and seed characteristics analyzed in this study indicate that among-population divergence exists in natural populations of A. aurea and occurs at two levels, site (i.e. mountain ranges) and elevation. Variations in seed weight, germination capacity, and early growth seem related to between-site differences. The studied sites represent different habitats for A. aurea. Otto can be characterized as a disturbed site, where individuals of A. aurea are weak, sparsely distributed, and share the understory with other species such as the bamboo Chusquea culeou. Populations at Chall Huaco, in contrast, can be described as successionally advanced where A. aurea grows vigorously in dense patches and is the dominant species of the understory. Under this scenario, greater seed weight and high germination capacity, as well as early growth may be beneficial in a competitive environment such as Otto, allowing A. aurea to rapidly occupy open habitats as soon as environmental conditions are favorable. Therefore, different selection regimes acting at each site may result in distinct establishment strategies. These differences appear to be genetic, a fact also suggested by

Souto and Premoli — Divergence among A. aurea populations

the isozyme analysis that indicated greater similarity between populations from a given site (Figure 1).

Restricted seedling establishment is considered a conspicuous feature of clonal plant population dynamics (Eriksson, 1992). A literature review indicated that 60% of the 68 species for which demographic information was available do not have seed recruitment into established populations (Eriksson, 1989). In Trifolium repens module recruitment was shown to far exceed seedling establishment, but at least some new genets are added on a yearly basis (Barret and Silander, 1992). Seedling recruitment may increase particularly at disturbed sites (Cahn and Harper, 1976; Turkington et al., 1979; Ennos, 1981) although establishment rates are higher in undisturbed sites (Barret and Silander, 1992). This suggests that physical disturbances may open sites for seedling recruitment, but these sites are not necessarily favorable for establishment. More studies are needed in A. aurea to determine the adaptive features of long-term seedling persistence in different sites.

It has been suggested that seed traits are under maternal control, which in turn is affected by differential environmental conditions. This is because maternal plants growing under benign environments are expected to produce large seeds together with increased germination speed and seedling size. However, this may not be always the case, particularly when maternal ability is not correlated with seed characteristics (Roach and Wulff, 1987). Our results show that, especially at Otto, where such a positive correlation between maternal and seed size traits is not evident, maternal effects would be small. Therefore, seed traits in A. aurea may be more affected by the individual genotype and the environment in which it grows, as previously suggested for other species (Weiner et al., 1997).

In addition to between-site differences, within-site differences were also recorded among populations of A. aurea separated by short-distances, i.e. hundreds of meters at different elevations. High-elevation populations had smaller seed yields than low-elevation populations. Aizen (2001), for the same Chall Huaco populations of A. aurea studied here, found a decline of ~30-40% in seed output between early (e.g. low-elevation population) and late flowers (e.g. high-elevation population). He ascribes this pattern to increasing resource limitation or the short seed-filling period associated with late flowering plants (Schemske et al., 1978; Ågren and Willson, 1992; Bertin and Sholes, 1993; Kudo, 1997). Such seasonal decreases in seed set seem to be quite common among bee-pollinated spring and summer flowering plants (e.g. Schemske et al., 1978; Thomson and Barrett, 1981; Murcia, 1990; Ågren and Willson, 1992; Bertin and Sholes, 1993; Kudo, 1997).

Altitudinal gradients provide differential long-term selection regimes that may lead to changes in life history characteristics (e.g., Clausen et al., 1940; Neuffer and Hurka, 1986; Galen et al., 1991; Mayer and Poljakoff-Mayber, 1989). A few studies have documented intraspecific patterns of altitudinal variation in the southern Andes. In Nothofagus pumilio, the dominant overstory species with windborne

pollen and seeds studied at Chall Huaco, an altitudinal variation for several adaptive features including isozymes was reported (Premoli, 2003; in press). In Nothofagus antarctica, a species with marked variation in growth forms associated with different environments, reduced seed weight and decreased germination capacity was measured as elevation increased in northwestern Patagonia (Premoli, 1991). Moreover, isozyme evidence suggests that differences in the growth habit of N. antarctica may be genetically determined (Vidal Russell, 2000). Therefore, variable physical conditions probably exert differential selective pressures which, in combination with differences in flowering phenology, reproductively isolate nearby populations of different plant species along elevation gradients.

The heterogeneity recorded in seed traits throughout A. aurea populations at different elevations and mountain ranges was also reflected in their low levels of gene flow measured by FST. These values and the total genetic diversity were within the range of those obtained for other herbaceous species, particularly those with gravity dispersion of seeds, insect pollination, and sexual and asexual reproduction (Hamrick and Godt, 1990). However, most of the total genetic diversity of A. aurea was distributed within sites (75%), suggesting considerable gene exchange between populations within a given mountain range. In particular, Otto populations seem to be maintaining higher gene flow rates than Chall Huaco, as indirectly estimated by FST and Nm. This result is probably related to patch density and pollinator activities. In Chall Huaco, A. aurea occurs in large patches, resulting in pollinator foraging that could be restricted to dense flowering patches. In contrast, small and fragmented populations, such as those at Otto, consist of patches with low floral density, and thus flight distances tend to be longer because they do not retain pollinators to the degree larger populations do. Thus, pollinator behavior may result in elevated among-patch gene flow that, in combination with the significantly increased recruitment of seeds at Otto, would explain the greater gene flow measured here.

Understanding the spatial organization of genetic diversity within and among plant populations is of critical importance for the development of strategies designed to preserve genetic variation (Hamrick, 1983; Brown and Briggs, 1991; Hamrick et al., 1991). It has been shown that species with limited gene flow, i.e. with restricted seed and/or pollen movement, have considerably more among-population variation for total amount of genetic diversity (Schoen and Brown, 1991). Thus, conservation strategies for such species as A. aurea should be developed with the dispersal ability of the species in mind.

Acknowledgements. We thank M. Aizen and one anonymous reviewer for helpful comments on early versions of this manuscript and E. Raffaele who provided statistical assistance. This work was supported by Grant No. B-036 from Centro Regional Universitario Bariloche, Universidad Nacional del Comahue and by a postdoctoral fellowship to A.P. from the National Research Council (CONICET) of Argentina.

Botanical Bulletin of Academia Sinica, Vol. 44, 2003

Literature Cited

Ågren, J. and M.F. Willson. 1992. Determinants of seed production in Geranium maculatum. Oecologia 92: 177-182.

Aizen, M. 2001. Flower sex ratio, pollinator abundance, and the seasonal pollination dynamics of a protandrous plant. Ecology 82: 127-144.

Aizen, M. 1997. Influence of local floral density and sex ratio on pollen receipt and seed output: empirical and experimental results in dichogamous Alstroemeria aurea (Alstroemeriaceae). Oecologia 111: 404-412.

Aizen, M. and A. Basilio. 1995. Within and among flower sex-phase distribution in Alstroemeria aurea (Alstroemeriaceae). Can. J. Bot. 73: 1986-1994.

Barret, J.P. and J. A. Silander, Jr. 1992. Seedling recruitment limitation in white clover (Trifolium repens; Leguminosae). Am. J. Bot. 79: 643-649.

Bazzaz, F.A. (ed.). 1996. Plants in Changing Enviroments, Linking physiological, population, and community ecology. Cambridge University Press, Cambridge.

Bertin, R.I. and O.D.V. Sholes. 1993. Weather, pollination and the phenology of Geranium maculatum. Am. Mid. Nat. 129: 52-66.

Brown, A.H. and J.D. Briggs. 1991. Sampling strategies for genetic variation in ex situ collections of endangered plant species. In D.A. Falk and K.F. Holsinger (eds.), Genetics and conservation of rare plants. Oxford University Press, New York, pp. 99-119.

Cahn, M.G. and J.L. Harper. 1976. The biology of the leaf mark polymorphism in Trifolium repens L. J. Ecol. 71: 307-330.

Clausen, J., D.D. Keck, and W.M. Hiesey. 1940. Experimental studies on the nature of species. I. Effect of varied environments in western North American plants. Carnegie Institute of Washington Publication 520, Washington D.C.

Crawley, M.J. 1991. Life history and environment. In M.J. Crawley (ed.), Plant Ecology. Blackwell Scientific Publications, London, pp. 253-290.

Dimitri, M.J. 1972. La Región de los bosques Andino - Patagónicos. Sinopsis General. Colección Científica del INTA. Tomo X. Buenos Aires.

Ennos, R.A. 1981. Detection of selection in populations of white clover (Trifolium repens L.). Biol. J. Linn. Soc. 15: 75-82.

Eriksson, O. 1989. Seedling dynamics and life hystories in clonal plants. Oikos 55: 235-238.

Eriksson, O. 1992. Evolution of seed dispersal and recruitment in clonal plants. Oikos 63: 439-448.

Free, J. B. 1970. Insect pollination of crops. Academic Press, New York.

Galen, C., J.S. Shore, and H. Deyoe. 1991. Ecotypic divergence in Alpine Polemonium viscosum: genetic structure, quantitative variation, and local adaptation. Evolution 45: 1218-1228.

Goudet, J. 2000. FSTAT. A program to estimate and test gene diversities and fixation indices. Release 2.9.1. Dorigny, Switzerland, Université de Lausanne. Available from

Hamrick, J.L. 1983. The distribution of genetic variation within and among natural plant populations. In C.M. Schoenwald-Cox et al. (eds.), Genetics and Conservation. Benjaminn Cummings: Menlo Park, NJ, pp. 335-348.

Hamrick, J.L. and M.J. Godt. 1990. Allozyme diversity in plant species. In A.D.H. Brown, M. T. Clegg, A.L. Kahler, and B.S. Weir (eds.), Plant population genetics, breeding, and genetic resources. Sinauer: Sunderland, MA, pp. 43-63.

Hamrick, J.L. and J.D. Nason. 1996. Consequences of dispersal in plants. In O.E. Rhodes, R.K. Chesser, and M.H. Smith (eds.), Population dynamics in ecological space and time. University of Chicago Press: Chicago, IL, pp. 203-236.

Hamrick, J.L., M.J. Godt, D.A. Murawski, and M.D. Loveless. 1991. Correlations between species traits and allozyme diversity: Implications for conservation biology. In D.A. Falk and K.E. Holsinger (eds.), Genetic and conservation of rare plants. Oxford University Press, New York, pp. 75-86.

Jain, S.K. 1976. The evolution of inbreeding in plants. Annu. Rev. Ecol. Syst. 7: 469-495.

King, J.N. and B.P. Dancik. 1983. Inheritance and linkage of isozymes in white spruce (Picea glauca). Can. J. Genet. Cytol. 25: 430-436.

Kudo, G. 1997. Sex expression and fruit set of an andromoecious herb, Peucedanum multivittatum (Umbelliferae) along a snowmelt gradient. Op. Bot. 132: 121-128.

Levin, D.A. 1984. Inbreeding Depression and Proximity-Dependent Crossing success in Phlox drummondii. Evolution 38: 116-127.

Levin, D.A. and H.W. Kerster. 1974. Gene flow in seed plants. Evol. Biol. 30: 139-220.

Mayer, A.M. and A. Poljakoff-Mayber. 1989. The Germination of Seeds. Pergamon Press, New York.

Mitton, J.B., Y. B Linhart, K.B. Sturgeon, and J.L. Hamrick. 1979. Allozyme polymorphisms detected in mature needle tissue of ponderosa pine. J. Hered. 70: 86-89.

Murcia, C. 1990. Effect of floral morphology and temperature on pollen receipt and removal in Ipomoea trichocarpa. Ecology 71: 1098-1109.

Nei, M. 1973. Analysis of gene diversity in subdivided populations. Proc. Nat. Acad. Sci. USA.70: 3321-3323.

Nei, M. 1978. F-statistics and analysis of gene diversity in subdivided populations. Ann. Hum. Genet. 41: 225-233.

Neuffer, B. and H. Hurka. 1986. Variation of development time until flowering in natural populations of Capsella bursa-pastoris (Cruciferae). Plant Syst. Evol. 152: 277-296.

Premoli, A.C. 1991. Morfología y capacidad germinativa en poblaciones de Nothofagus antactica (Forster) Oerst. Del noroeste andino patagónico. Bosque 12: 53-59.

Premoli, A.C., T. Kitzberger , M. Caldiz, C. Souto, S. Arana, F. García, and C. Palópoli. 2000. Cultivemos nuestras plantas nativas: Técnicas de Germinación. Patagonia Silvestre 7: 3-7.

Premoli, A.C. 2003. Nothofagus caducifolios de hoja pequeña y de distribución preferentemente meridional o de altura: Nothofagus pumilio. In: Variación en especies nativas de los bosques templados chileno-argentinos. In C. Donoso, A.C. Premoli, L. Gallo, and R. Ipinza (eds.), Santiago, Chile: Editorial Universitaria, in press.

Premoli, A.C. 2003. Isozyme polymorphisms provide evidence of clinal variation with elevation in Nothofagus pumilio. J. Hered. 94: 218-226.

Puntieri, J.G. 1991. Vegetation response on a forest slope cleared for a skin-run with special reference to the herb Alstroemeria

Souto and Premoli — Divergence among A. aurea populations

Souto, C.P. 1999. Barreras precigóticas y distancias de cruzamiento en el éxito reproductivo de Alstroemeria aurea: Una hipótesis genético - ecológica. Licenciatura Thesis, Universidad Nacional del Comahue. Bariloche, Argentina.

Souto, C.P., M. Aizen, and A. Premoli. 2002. Effects of crossing distance and genetic relatedness on pollen performance in Alstroemeria aurea (Alstroemeriaceae). Am. J. Bot. 89: 427-432.

Thomson, J.D. and S.C. H. Barrett. 1981. Temporal variation in Aralia hispida Vent. (Araliaceae). Evolution 35: 1094-1107.

Turkington, R., M.A. Cahn, A. Vardy, and J.L. Harper. 1979. The growth distribution and neighbor relationships of Trifolium repens in a permanent pasture. III. The establishment and growth of Trifolium repens in natural and perturbed sites. J. Ecol. 67: 231-243.

Vidal Russell, R. 2000. Evidencias de resistencia en Nothofagus al Misodendrum: patrones de infección y consecuencias sobre la estructura genética de la planta parásita. Licenciatura Thesis, Universidad Nacional del Comahue, Bariloche, Argentina.

Waser, N. and M. Price. 1991. Outcrossing distance effects in Delphinium nelsonii: pollen loads, pollen tubes and seed set. Ecology 72: 171-179.

Weiner, J., S. Martinez, H. Müller-Schärer, P. Stoll, and B. Schmid. 1997. How important are environmental maternal effects in plants? A study with Centaurea maculosa. J. Ecol. 85: 133-142.

Weir, B.S. and C.C. Cockerham. 1984. Estimating F-statistics for the analysis of population structure. Evolution 38: 1358-1370.

Wright, S. 1978. Evolution and the Genetics of Populations. Vol. 4. Variability Within and Among Natural Populations. University of Chicago Press, Chicago, IL, USA.

aurea Graham (Alstroemeriaceae), Argentina. Biol. Conserv. 56: 207-221.

Ranker, T.A., C.H. Haufler, P.S. Soltis, and D.E. Soltis. 1989. Genetic evidence for allopolyploidy in the neotropical fern Hemionitis pinnitifida (Adiantaceae) and the reconstruction of an ancestral genome. Sys. Bot. 14: 439-447.

Redmond, A., J. Robbins, and L. Travis. 1989. The effects of pollination distance on seed production in three populations of Amianthium muscaetoxicum (Liliaceae). Oecologia 9: 260-264.

Rice, W.R. 1989. Analyzing tables of statistical tests. Evolution 43: 223-225.

Roach, D.A. and R. Wulff. 1987. Maternal effects in plants. Ann. Rev. Ecol. Syst. 18: 209-235.

Sanso, A.M. 1996. El género Alstroemeria (Alstroemeriaceae) en Argentina. Darwiniana 34: 349-382.

Schemske, D.W., M.F. Willson, M.N. Melampy, L.J. Miller, L. Verner, K.M. Schemske, and L.B. Best. 1978. Flowering ecology of some spring woodland herbs. Ecology 59: 361-366.

Schoen, D.J. and A.H.D. Brown. 1991. Intraspecific variation in population gene diversity and effective population size correlates with the mating system in plants. Proc. Nat. Acad. Sci. USA 88: 4494-4497.

Slatkin, M. 1985. Gene flow in natural populations. Ann. Rev. Ecol. Syst. 16: 393-430.

Sobrevila, C. 1988. Effects of distance between pollen donor and pollen recipient on fitness components in Espeletia schultzii. Amer. J. Bot. 75: 701-724.

Soltis, D.E., C.H. Haufler, D.C. Darrow, and G.J. Gastony. 1983. Starch gel electrophoresis of ferns: A compilation of grinding buffers, and staining schedules. Amer. Fern. J. 73: 9-27.

Botanical Bulletin of Academia Sinica, Vol. 44, 2003