Plastome sequencing of South American Podocarpus species reveals low rearrangement rates despite ancient gondwanan disjunctions.

BACKGROUND: Historical reconstructions within Podocarpaceae have provided valuable information to disentangle biogeographic scenarios that begun 65 Mya. However, early molecular phylogenies of Podocarpaceae failed to agree on the intergeneric relationships within the family. The aims of this study were to test whether plastome organization is stable within the genus Podocarpus, to estimate the selective regimes affecting plastome protein-coding genes, and to strengthen our understanding of the phylogenetic relationships and biogeographic history. METHODS AND RESULTS: We sequenced the plastomes of four South American species from Patagonia, southern Yungas, and Brazilian subtropical forests. We compared their plastomes to those published from Brazil, Africa, New Zealand, and Southeast Asia, along with representatives from other genera within Podocarpaceae as outgroups. The four newly sequenced plastomes ranged in size between 133,791 bp and 133,991 bp. Gene content and order among chloroplasts from South American, African and Asian Podocarpus were conserved and different from the plastome of P. totara, from New Zealand. Most genes showed substitution patterns consistent with a conservative selective regime. Phylogenies inferred from either complete sequences or protein coding regions were mostly congruent with previous studies, but showed earlier branching of P. salignus, P. totara and P. sellowii. CONCLUSIONS: Highly similar and conserved plastomes of African, South American and Asian species suggest that P. totara plastome should be revised and compared to other species from Oceanic distribution. Furthermore, given such structural conservation, we suggest plastome sequencing is not useful to test whether genomic order can be climatically or geologically structured.

A relic of the past: current genetic patterns of the palaeoendemic tree Nothofagus macrocarpa were shaped by climatic oscillations in central Chile.

BACKGROUND AND AIMS: The Mediterranean-type forest of central Chile is considered a 'biodiversity hotspot' and a relic of a wider ancient distribution produced by past climatic oscillations. Nothofagus macrocarpa, commonly known as 'roble de Santiago', is a threatened palaeoendemic of this forest, poorly represented in the protected area system. This tree has been repeatedly misidentified as the sister species N. obliqua, which has affected its recognition and protection. Only a few populations of N. macrocarpa remain within a matrix of intensive land use that has been affected by recent forest fires. We tested the hypothesis that current populations of N. macrocarpa are a relic state of a previously widespread range, with the aim of contributing to its identification, its biogeographical history and the design of conservation measures using genetic information. METHODS: We analysed remnant N. macrocarpa forests using nuclear (nDNA) and chloroplast DNA (cpDNA) sequences, conducted phylogenetic and phylogeographical analyses to reconstruct its biogeographical history, and assessed microsatellites [simple sequence repeats (SSRs)] to determine contemporary patters of diversity within and among all remnant populations. We also examined the degree of past, current and potential future isolation of N. macrocarpa populations using ecological niche models (ENMs). KEY RESULTS: The species N. macrocarpa was confirmed by nDNA sequences, as previously suggested by chromosomal analysis. Small isolated populations of N. macrocarpa exhibited moderate to high genetic diversity according to SSRs. cpDNA analysis revealed a marked past latitudinal geographical structure, whereas analysis of SSRs did not find such current structure. ENM analyses revealed local expansion-contraction of the N. macrocarpa range during warmer periods, particularly in the northern and central ranges where basal-most cpDNA haplotypes were detected, and recent expansion to the south of the distribution. CONCLUSIONS: Genetic patterns confirm that N. macrocarpa is a distinct species and suggest a marked latitudinal relic structure in at least two evolutionarily significant units, despite contemporary among-population gene flow. This information must be considered when choosing individuals (seeds and/or propagules) for restoration purposes, to avoid the admixture of divergent genetic stocks.

Refining predictions of population decline at species' rear edges.

According to broad-scale application of biogeographical theory, widespread retractions of species' rear edges should be seen in response to ongoing climate change. This prediction rests on the assumption that rear edge populations are "marginal" since they occur at the limit of the species' ecological tolerance and are expected to decline in performance as climate warming pushes them to extirpation. However, conflicts between observations and predictions are increasingly accumulating and little progress has been made in explaining this disparity. We argue that a revision of the concept of marginality is necessary, together with explicit testing of population decline, which is increasingly possible as data availability improves. Such action should be based on taking the population perspective across a species' rear edge, encompassing the ecological, geographical and genetic dimensions of marginality. Refining our understanding of rear edge populations is essential to advance our ability to monitor, predict and plan for the impacts of environmental change on species range dynamics.

Niche squeeze induced by climate change of the cold-tolerant subtropical montane Podocarpus parlatorei.

Under changing climates, the persistence of montane subtropical taxa may be threatened as suitable habitats decrease with elevation. We developed future environmental niche models (ENNMs) for Podocarpus parlatorei, the only conifer from southern Yungas in South America, and projected it onto two greenhouse gas concentration scenarios based on 13 global climate models for the years 2050 and 2070. Modelling identified that P. parlatorei is sensitive and restricted to a relatively narrow range of both warm season temperature and precipitation. By the mid-late twenty-first century areas of high suitability for P. parlatorei will not migrate but overall suitability will become substantially reduced across its whole range and surrounding areas. Despite extensive areas in high mountain ranges where the species may encounter thermally optimal conditions to potentially allow upward local migration, these same areas will likely become strongly aridified under future conditions. On the other hand, in lowland locations where rainfall levels will not change substantially (e.g. northern range), excessive warming will likely generate abiotic and biotic restrictions (e.g. competition with lowland species) for this cold-tolerant species. Urgent measures should be developed for the local long-term preservation of the gene pool of the unique conifer that characterizes Yungas forests for reasons of biodiversity conservation and ecosystem services.

Climatic niche shift in the amphitropical disjunct grass Trichloris crinita.

Plant species disjunctions have attracted the interest of ecologists for decades. We investigated Trichloris crinita, a native C4 perennial grass with disjunct distribution between subtropical regions of North and South America, testing the hypothesis that the species has a similar realized climatic niche in both subcontinents. The climatic niche of T. crinita in North and South America was characterized and compared using presence records and five uncorrelated bioclimatic variables selected according to their ecological importance for the species. We used reciprocal modeling to make geographic projections of the realized niche within each subcontinent. Niche overlap between T. crinita distributions in North and South America was intermediate for the individual climatic variables and the multivariate space. In all cases the test of equivalence between climates inhabited by T. crinita indicated that the realized niche of the species differ significantly between subcontinents. Also, the similarity test showed that in the majority of cases the realized niche in both subcontinents was significantly different than that expected by chance. T. crinita occupied a greater diversity of environments in South than in North America, while in the latter its distribution was displaced to drier and warmer environments. The modeled geographic distribution using the actual occurrences of the species in North America did not accurately predict the distribution in South America, and vice versa. Together, these results led us to reject the hypothesis of similar niche of T. crinita in both subcontinents. This information may be useful to manage restoration efforts by presenting the suitable areas and climates for the species, and suggesting that translocation of individuals between subcontinents could only be recommended with caution because introduced genotypes can be potentially maladaptive, and could colonize sites actually not occupied by the species within each subcontinent.

Population Genetic Structure of the Giant Cactus Echinopsis terscheckii in Northwestern Argentina Is Shaped by Patterns of Vegetation Cover.

Species inhabiting drylands commonly depend on the surrounding vegetation for recruitment under stress, while competition may affect populations in moister environments. Our objective was to analyze how different climates and vegetation affect the fine-scale spatial genetic structure (SGS) of the columnar cactus Echinopsis terscheckii. At 4 sites, we estimated vegetation cover by digitized patches and the normalized difference vegetation index (NDVI). We mapped 30 individuals per population and collected tissue for isozyme electrophoresis using 15 putative loci. Spatial autocorrelation between all possible genotype pairs and the number of genetically homogeneous groups and families were calculated for each population. Greater cover (66%) and average NDVI values were detected in the most humid habitat that consisted of fewer, larger, and more dispersed vegetation patches. All populations were genetically diverse and showed significant SGS. Positive correlations were found between the distance at which maximum autocorrelation and kinship values were reached and vegetation area and patch size. Also higher NDVI values were associated with lower number of patches. Populations exposed to higher precipitation and vegetation cover consisted of sparse individuals that clustered at larger distances whereas vegetation patches in arid climates produced groups of closely related genotypes at small distances. These results support the stress-gradient genetic hypothesis. Under water stress, facilitation promotes establishment underneath patchy vegetation resulting in fine-scale family structure. In moister xerophilous forests, competition for resources, that is, light, results in sparse individuals and thus coarse-scale neighborhoods. This information can guide conservation and/or restoration efforts, such as the spatial scale to be considered in germplasm collection.

Living on the edge: adaptive and plastic responses of the tree Nothofagus pumilio to a long-term transplant experiment predict rear-edge upward expansion.

Current climate change affects the competitive ability and reproductive success of many species, leading to local extinctions, adjustment to novel local conditions by phenotypic plasticity or rapid adaptation, or tracking their optima through range shifts. However, many species have limited ability to expand to suitable areas. Altitudinal gradients, with abrupt changes in abiotic conditions over short distances, represent "natural experiments" for the evaluation of ecological and evolutionary responses under scenarios of climate change. Nothofagus pumilio is the tree species which dominates as pure stands the montane forests of Patagonia. We evaluated the adaptive value of variation in quantitative traits of N. pumilio under contrasting conditions of the altitudinal gradient with a long-term reciprocal transplant experimental design. While high-elevation plants show little response in plant, leaf, and phenological traits to the experimental trials, low-elevation ones show greater plasticity in their responses to changing environments, particularly at high elevation. Our results suggest a relatively reduced potential for evolutionary adaptation of high-elevation genotypes, and a greater evolutionary potential of low-elevation ones. Under global warming scenarios of forest upslope migration, high-elevation variants may be outperformed by low-elevation ones during this process, leading to the local extinction and/or replacement of these genotypes. These results challenge previous models and predictions expected under global warming for altitudinal gradients, on which the leading edge is considered to be the upper treeline forests.

Identifying Genetic Hotspots by Mapping Molecular Diversity of Widespread Trees: When Commonness Matters.

Conservation planning requires setting priorities at the same spatial scale at which decision-making processes are undertaken considering all levels of biodiversity, but current methods for identifying biodiversity hotspots ignore its genetic component. We developed a fine-scale approach based on the definition of genetic hotspots, which have high genetic diversity and unique variants that represent their evolutionary potential and evolutionary novelties. Our hypothesis is that wide-ranging taxa with similar ecological tolerances, yet of phylogenetically independent lineages, have been and currently are shaped by ecological and evolutionary forces that result in geographically concordant genetic patterns. We mapped previously published genetic diversity and unique variants of biparentally inherited markers and chloroplast sequences for 9 species from 188 and 275 populations, respectively, of the 4 woody dominant families of the austral temperate forest, an area considered a biodiversity hotspot. Spatial distribution patterns of genetic polymorphisms differed among taxa according to their ecological tolerances. Eight genetic hotspots were detected and we recommend conservation actions for some in the southern Coastal Range in Chile. Existing spatially explicit genetic data from multiple populations and species can help to identify biodiversity hotspots and guide conservation actions to establish science-based protected areas that will preserve the evolutionary potential of key habitats and species.

How do cold-sensitive species endure ice ages? Phylogeographic and paleodistribution models of postglacial range expansion of the mesothermic drought-tolerant conifer Austrocedrus chilensis.

In view of global climate change, it is important to understand the responses of tree species to climate changes in the past. Combinations of phylogeographic analysis of genetic evidence, coupled with species distribution models (SDMs), are improving our understanding on this subject. We combined SDMs and microsatellite data from populations of the entire range of Austrocedrus chilensis, a dominant mesotherm (cold-sensitive) conifer of dryland forests of the southern Andes, to test the hypothesis of long-distance postglacial migration from northern and warmer refugia at the Last Glacial Maximum (LGM). The SDM indicated suitable conditions for Austrocedrus in northern Chile (western) at the LGM and largely unsuitable conditions in Argentina (eastern). Population genetic diversity and effective population sizes within populations decreased southward along the Andes, consistent with the hypothesis of long-distance dispersal from a northern refugium. Results support the hypothesis of one (or a few) warmer (low latitude) refugia in Chile for Austrocedrus. On balance, the evidence suggests that in contrast to cold-tolerant tree taxa with the capacity to fast-track postglacial warming thanks to local refugia, cold-sensitive species might have undergone long-distance range expansion, lagging behind progressive climate change throughout the Holocene.

Fine-scale genetic structure of Nothofagus pumilio (lenga) at contrasting elevations of the altitudinal gradient.

Montane forests provide the natural framework to test for various ecological settings at distinct elevations as they may affect population demography, which in turn will affect the spatial genetic structure (SGS). We analyzed the fine-scale SGS of Nothofagus pumilio, which dominates mountain areas of Patagonia, in three pairs of sites at contrasting elevations (low- vs. high-elevation). Within a total area of 1 ha fresh leaf tissue from 90 individuals was collected at each of the six studied stands following a spatially explicit sampling design. Population genetic diversity parameters were analyzed for all sampled individuals using five polymorphic isozyme loci, and a subset of 50 individuals per stand were also screened for five microsatellite loci. The SGS was assessed on 50 individuals/stand, using the combined datasets of isozymes and microsatellites. Most low-elevation stands consisted of older individuals with complex age structures and genetically diverse plots. In contrast, high-elevation stands and one post-fire low-elevation population yielded even-aged structures with evidence of growth suppression, and were genetically homogeneous. All stands yielded significant SGS. Similarly to mature stands of the non-sprouter congener Nothofagus dombeyi, multi-age low-altitude N. pumilio yielded significant SGS weakened by competing species of the understory and the formation of seedling banks. Alike the sprouter Nothofagus antarctica, high-altitude stands produced significant SGS as a consequence of occasional seedling establishment reinforced by vegetative spread.

Shrinking forests under warming: evidence of Podocarpus parlatorei (pino del cerro) from the subtropical Andes.

Phylogeography in combination with ecological niche modeling (ENM) is a robust tool to analyze hypotheses on range shifts under changing climates particularly of taxa and areas with scant fossil records. We combined phylogeographic analysis and ENM techniques to study the effects of alternate cold and warm (i.e., glacial and interglacial) periods on the subtropical montane cold-tolerant conifer Podocarpus parlatorei from Yungas forests of the central Andes. Twenty-one populations, comprising 208 individuals, were analyzed by sequences of the trnL -trnF cpDNA region, and 78 sites were included in the ENM. Eight haplotypes were detected, most of which were widespread while 3 of them were exclusive of latitudinally marginal areas. Haplotype diversity was mostly even throughout the latitudinal range. Two distribution models based on 8 bioclimatic variables indicate a rather continuous distribution during cooling, while under warming remained within stable, yet increasingly fragmented, areas. Although no major range shifts are expected with warming, long-lasting persistence of cold-hardy taxa inhabiting subtropical mountains may include in situ and ex situ conservation actions particularly toward southern (colder) areas.

Phylogeographically concordant chloroplast DNA divergence in sympatric Nothofagus s.s. How deep can it be?

• Here, we performed phylogenetic analyses and estimated the divergence times on mostly sympatric populations of five species within subgenus Nothofagus. We aimed to investigate whether phylogenetic relationships by nuclear internal transcribed spacer (ITS) and phylogeographic patterns by chloroplast DNA (cpDNA) mirror an ancient evolutionary history that was not erased by glacial eras. Extant species are restricted to Patagonia and share a pollen type that was formerly widespread in all southern land masses. Weak reproductive barriers exist among them. • Fifteen cpDNA haplotypes resulted from the analysis of three noncoding regions on 330 individuals with a total alignment of 1794 bp. Nuclear ITS data consisted of 822 bp. We found a deep cpDNA divergence dated 32 Ma at mid-latitudes of Patagonia that predates the phylogenetic divergence of extant taxa. Other more recent breaks by cpDNA occurred towards the north. • Complex paleogeographic features explain the genetic discontinuities. Long-lasting paleobasins and marine ingressions have impeded transoceanic dispersal during range expansion towards lower latitudes under cooler trends since the Oligocene. • Cycles of hybridization-introgression among extant and extinct taxa have resulted in widespread chloroplast capture events. Our data suggest that Nothofagus biogeography will be resolved only if thorough phylogeographic analyses and molecular dating methods are applied using distinct genetic markers.

Oldies (but goldies!): extracting DNA from cryopreserved allozyme homogenates.

In this work, we describe a method that allows the extraction of plant genomic DNA from frozen enzyme homogenates. To test if reliable DNA was recovered from allozyme extracts, we compared these results with that obtained from fresh frozen and dried material. Sequencing and microsatellite fragments were analyzed for samples from 5 plant families in order to check for chloroplast and nuclear DNA quality. High-molecular weight DNA was recovered from frozen homogenates, and it was found that the quality was independent of storage time, which varied from 1 to 9 years. High-quality PCR products along with sequences and microsatellite fragments were obtained for nearly all analyzed samples. Frozen protein homogenates from previous population studies could thus be used for further phylogeographical and phylogenetic analyses using DNA molecular tools. This simple method provides an opportunity to take advantage of stored cryopreserved samples.

Evidence of chloroplast capture in South American Nothofagus (subgenus Nothofagus, Nothofagaceae).

Subgenus Nothofagus, although geographically restricted at present to temperate areas of South America, has captured much attention in discussions of plant biogeography due to its widespread distribution through Gondwanan continents during the Tertiary. However, phylogenetic relationships within the subgenus Nothofagus have not yet been resolved. We examined geographic patterns of intraspecific and interspecific genetic variation to detect whether incongruences in nuclear or plastid DNA phylogenies occur, in order to better understand the evolutionary history of the subgenus Nothofagus. We conducted spatially-explicit sampling at 10 distinct locations throughout the range of austral South American forests and sampled all present Nothofagus species. We used ITS and chloroplast DNA sequences to estimate phylogenetic relationships. A phylogeny constructed from nuclear genes resolved the subgenus Nothofagus as monophyletic. We found that N. antarctica was a sister to a clade of evergreen species (N. betuloides, N. dombeyi, and N. nitida), while N. pumilio likely diverged earlier. Nine cpDNA haplotypes were distinguished in the subgenus Nothofagus which were associated to geographic locations rather than to taxonomic relationships. This species-independent cpDNA phylogeographic structures within the subgenus Nothofagus may be related to repeated chloroplast capture events over geological time in Patagonia.

Out in the cold: genetic variation of Nothofagus pumilio (Nothofagaceae) provides evidence for latitudinally distinct evolutionary histories in austral South America.

Nothofagus pumilio is the dominant and almost ubiquitous tree species in mountainous environments of temperate South America. We used two types of molecular markers (cpDNA and isozymes) to evaluate the effects of the Paleogene paleogeography of Patagonia and more recent climatic oscillations of the Neogene on such cold-tolerant species' genetic makeup. Phylogeographic analysis on sequences of three cpDNA non-coding regions at 85 populations yielded two latitudinally disjunct monophyletic clades north and south of c. 42 degrees S containing 11 and three haplotypes, respectively. This indicates a long-lasting vicariant event due to the presence of an extended open paleobasin at mid latitudes of Patagonia. Also distribution patterns of cpDNA haplotypes suggest regional spread following stepping-stone models using pre-Cenozoic mountains as corridors. Comparable genetic diversity measured along 41 sampled populations using seven polymorphic isozyme loci provides evidence of local persistence and spread from multiple ice-free locations. In addition, significantly higher heterozygosity and allelic richness at high latitudes, i.e. in areas of larger glacial extent, suggest survival in large and isolated refugia. While, higher cpDNA diversity in lower latitudes reflects the complex orogeny that historically isolated northern populations, lower isozyme diversity and reduced F(ST) values provide evidence of local glacial survival in numerous small locales. Therefore, current genetic structure of N. pumilio is the result of regional processes which took place during the Tertiary that were enhanced by contemporary local effects of drift and isolation in response to Quaternary climatic cycles.

Genetics of sprouting: effects of long-term persistence in fire-prone ecosystems.

Fire functional traits (postfire resprouting and seeding) are considered to be adaptations for persisting in fire-prone environments. Although ecological and evolutionary consequences of sprouting have been extensively discussed, within-species genetic variability and structure are unknown. Here we report levels and distribution patterns of genetic polymorphisms in postfire stands of the predominant sprouter Nothofagus antarctica. Fresh foliage of 50 individuals was collected following a spatially explicit sampling design for isozyme analysis from two replicates of each of four habitat types inhabited by the species in northwestern Patagonia, Argentina: matorral, high elevation, forest, and temporally flooded basins. Average polymorphism per population ranged from 44% to 78% and mean gene diversity per site H(S )varied from 0.187 to 0.274. These results show that sprouter populations hold considerable genetic variation. Significant genetic structure over short distances (< 50 m) was found at all locations. Ancient fine-scale genetic structure is preserved by occasional seedling establishment that results in high co-ancestry coefficients. Sprouter populations growing in suboptimal habitats such as matorral, high elevation or basins consist of pairs of heterozygous genets that occur at larger spatial scales as a result of micro-environmental heterogeneity and/or local competition between near neighbour genotypes. In contrast, homozygous pairs of individuals for distinct isozyme loci occurred at larger spatial scales in forest stands. This indicates that biparental inbreeding due to local propagule establishment may take place to some extent in sprouters growing under favourable conditions. Our results show that sprouters follow a long-lasting genet persistence strategy which most probably is selected under unpredictable disturbance regimes, such as fire.

Genetic structure and early effects of inbreeding in fragmented temperate forests of a self-incompatible tree, Embothrium coccineum.

Deforestation of temperate forests has created landscapes of forest remnants in matrices of intense human use. We studied the genetic effects of fragmentation in southern Chile on Embothrium coccineum J.R. et G. Forster, an early colonizing, bird-pollinated tree. We tested the hypothesis that, because of its self-incompatibility and life-history strategy, E. coccineum is less strongly affected by fragmentation. We studied the effects of reduced population size and increased isolation on population genetic structure and early performance of progeny. Samples were collected from spatially isolated trees and six fragments of differing sizes (small, 1 ha; medium, 20 ha; large, >150 ha). Based on isozyme polymorphisms we estimated parameters of genetic diversity, divergence, and inbreeding for adults and greenhouse-grown progeny. We also measured germination, seedling growth, and outcrossing rates on progeny arrays. Genetic variation of adults did not correlate significantly with population size, as expected, given that fragmentation occurred relatively recently. Weak effects of fragmentation were measured on progeny. Only adults yielded significant inbreeding. Similar total genetic diversity was found in adults and progeny. Low but significant genetic differentiation existed among adult and progeny populations. Seedling growth correlated positively with the effective number of alleles, showing deleterious effects of inbreeding on progeny. Seeds from small fragments had the highest outcrossing rates and germination success, indicating that higher pollinator activity in such fragments reduced selfing, thereby buffering genetic erosion and maintaining adaptive variation. The effects of forest fragmentation were detectable in E. coccineum, but these effects will probably not be detrimental to the viability of remnant populations because small, fragmented populations demonstrated higher levels of gene flow and lower inbreeding than larger stands. Pioneer species that are insensitive to forest clearing may be crucial in recovery plans to facilitate the establishment of species intolerant to such disturbance.

Regeneration mode affects spatial genetic structure of Nothofagus dombeyi forests.

Disturbance may generate population bottlenecks by reducing population size and the number of founders establishing a new colony. We tested the hypothesis that the scale of disturbance affects the levels of genetic diversity and the spatial distribution of genotypes in naturally regenerating stands of Nothofagus dombeyi, an evergreen angiosperm tree, in northwestern Patagonia. At similar spatial scales, we predicted that old-growth stands characterized by fine-scale gap phase dynamics would be genetically diverse due to restricted gene flow among temporal and spatially isolated gaps. In contrast, young massively regenerated postfire cohorts resulting from coarse-scale disturbances would be genetically more homogeneous. At each of three paired old-growth and postfire stands a minimum of 50 trees were mapped and sampled within 1 ha. Fresh tissue was collected for isozyme analysis from a total of 361 trees along with tree cores and diameters. Tree age distributions reflected the dominant modes of regeneration. Six out of nine analysed loci were polymorphic. Mean genetic diversity parameters were greater but not significant in mature stands. Fixation indices suggested significant heterozygous deficit at two-thirds of possible tests indicating a Wahlund effect due to local recruitment of related seeds. F(ST) indicated moderate between-stand divergence. Mature stands concentrated half of positively like joins and yielded significant (P < 0.05) autocorrelation coefficients at small distance classes (< 20 m). Fine-scale patch dynamics within mature stands favours the maintenance of fine-scale genetic structure as a result of shade intolerance and local seed dispersal. Conversely, postfire stands suffer the effects of genetic drift given that a few reproductive trees produce a somewhat impoverished and genetically uniform progeny. Bottleneck effects will depend upon the density of remnant trees which could also be a function of the severity of fire.

Effects of crossing distance and genetic relatedness on pollen performance in Alstroemeria aurea (Alstroemeriaceae).

Prezygotic barriers may represent effective mechanisms to avoid the deleterious effects of inbreeding. This study reports the existence of distance-dependent prezygotic barriers in self-compatible Alstroemeria aurea, a clonal herb native to temperate forests of the southern Andes. We analyzed pollen germination and tube growth as indicators of donor-recipient affinity using crossing distances of 1, 10, and 100 m. We used allozyme electrophoresis to determine the actual genetic relatedness between donor and recipient ramets. Pollen germination was not affected by distance between mates, but the number of pollen tubes reaching the base of the style increased strongly with distance between donor and recipient. This pattern was related to an increase in genetic dissimilarity with distance between mates. In contrast, pollen tube-style interactions did not change with distance when we restricted analysis to individuals at different distances that appeared to be genetically identical. This test implied genetic dissimilarity as the critical factor affecting pollen performance. We propose that the existence of prezygotic barriers might contribute to the high degree of genetic mixing exhibited by some clonal species.

GENETIC VARIATION IN ALETES ACAULIS AND ITS RELATIVE, THE NARROW ENDEMIC A. HUMILIS (APIACEAE).

We compared the organization of genetic variability in Aletes humilis, a species restricted to seven populations in north-central Colorado, with A. acaulis, a closely related widespread species. Genetic variability was scored at 11 electrophoretically detectable protein loci. Nine of these loci were polymorphic (i.e., P > 0.95) in both taxa, and were used for interpopulation and interspecific comparisons. Levels of genetic variability and patterns of organization of this variability are very comparable in both species. In contrast to many other narrow endemics, A. humilis is not genetically depauperate when compared to its presumed progenitor. Both biochemical and morphological evidence suggest that A. humilis is derived from A. acaulis; the comparable levels of variability in the two species suggest that A. humilis was derived from a source containing a substantial portion of the original A. acaulis genome, because there is no evidence of a genetic bottleneck or appreciable loss of allelic variability.