Isozymes,Speciation and Evolution in the Polyploid Complex Cochlearia L. (Brassicaceae)

Within the genus Cochlearia L. sect. Cochlearia three isozyme systems (aspartate aminotransferase, leucine aminopeptidase and phosphoglucomutase) have been analysed at the populational level for eleven species with different ploidy levels. Nine loci complexes were resolved with a total of 33 alleles from 89 populations. In general, polyploids displayed higher numbers of alleles per species and per population. A notable exception is diploid C. pyrenaica with as many alleles as some polyploids. Data support: (1) the origin of tetraploid C. officinalis from C. pyrenaica s.l., particularly from C. aestuaria; (2) the allopolyploid origin of octoploid C. anglica from C. officinalis; (3) the allopolyploid origin of hexaploid inland taxa C. polonica, C. tatrae and C. bavarica via hybridization between C. pyrenaica and C. officinalis; (4) a putative allopolyploid origin of hexaploid coastal species C. danica by hybridization between C. pyrenaica s.l. and C. officinalis accompanied by a shift in chromosome complement. A scenario summarizing our present understanding of the evolution within Cochlearia sect. Cochlearia is presented.     

Differences in the life histories of twoCochlearia species

Cochlearia bavarica Vogt andC. pyrenaica Dc. are closely related and highly endangered species in Bavaria (SE Germany).Cochlearia bavarica (2n=36) is a hybrid ofC. pyrenaica (2n=12) andC. officianalis (2n=24) stabilized by polyploidy.Cochlearia officinalis does not occur in Bavaria. The question being addressed is whether the present non-overlapping distribution of the two Bavarian species, both of which are restricted to calcareous springs with a continuous water supply or drainage ditches, is due to competitive exclusion or to subtle differences of ecological requirements. I did not find any evidence for the second case, i.e. niche differentiation, because both species occurred in all the habitat types that I could define by floristic and ecological criteria. The population structures of bothC. bavarica andC. pyrenaica vary in parallel with different habitat types. Both species have the greatest reproductive output (viable seeds) and the highest population densities in habitats with high light availability and/or low interspecific competition. Thus, I expected that one of the two species might be competitively superior over the other, but that it would fail to reach all its potential sites of occurrence. Cochlearia bavarica individuals (dry mass =5.75 g) grow significantly larger thanC. pyrenaica individuals (3.62 g;P<0.05) and produce more, larger and heavier seeds, possibly due to hybrid vigour. Flowering individuals ofC. bavarica also allocate more biomass to vegetative structures thanC. pyrenaica. The analysis of population dynamics reveals a higher life expectancy and reproductive output inC. bavarica. The calculated population growth rates although negative over the observation period in both species, suggest a significantly higher fitness ofC. bavarica (λ=0.66) compared withC. pyrenaica (λ=0.44). Thus, our data are in line with the competitive exclusion hypothesis andCochlearia bavarica appears as an example of a species with hybridogenous origin possessing higher fitness than its progenitor. However, considering the rarity of both species it seems at present unlikely that the former will displace the latter over a large distributional region.     

Large-scale geographic patterns of genetic variation in Melica nutans, a widespread Eurasian woodland grass

 The geographic distribution of allozyme variation within the Eurasian boreo-nemoreal woodland grass Melica nutans L. has been investigated together with a minor subset of other Melica species. Twenty alleles were found at nine polymorphic loci in M. nutans. Allelic richness was highest in areas central in the species' European distribution, i.e. in southern Fennoscandia. High population densities, reducing the effects of genetic drift, as well as accumulation of variation through long-distance gene-flow from different marginal populations, is proposed to explain high allelic richness in this area. Several alleles showed geographic patterns in distribution and frequency variation. However, these patterns were not congruent, e.g. some alleles appear to have migrated to northern Europe from the south-west whereas others may have spread from the east. Genetic distances between geographic regions, each consisting of 2–6 populations, were generally low between all Fennoscandian, Russian and Siberian regions, but much higher between western and continental European regions. On the population level, cluster analysis grouped populations from Siberia, Russia, coastal and lowland areas in Fennoscandia and British Cumbria into one subcluster whereas other subclusters contained mainly south-west European populations or populations from almost throughout the distribution range. A scenario with several independent glacial refugia in central Europe, south-western Siberia and possibly western Norway, and subsequent colonisation of Fennoscandia mainly from the east, but with some long-distance gene-flow from central Europe, is proposed. Received April 3, 2002; accepted September 17, 2002 Published online: December 11, 2002     

Genetic variation in Scandinavian Anthericum liliago (Anthericaceae): allopolyploidy, hybridization and immigration history

 We investigated patterns of isozyme variation and the hierarchic structure of genetic diversity in 25 Scandinavian populations of the lilioid herb, Anthericum liliago. Isozyme data suggest that tetraploid A. liliago has an allopolyploid origin and that A. ramosum may be one of its diploid progenitors. Two populations contained known or suspected hybrids between A. liliago and A. ramosum. Isozymes show that one population from S Sweden contains both triploid (hybrid) and tetraploid individuals whereas a putatively hybrid Danish population represents diploid A. liliago. There is an overall northward and eastward decline in allelic richness in the tetraploid populations, with the highest numbers of alleles in Denmark and SW Sweden. This pattern is consistent with a progressive loss of allelic variation during the species' postglacial colonization of Scandinavia. The between-population component of genetic diversity is 4% (compared with 12% in diploid A. ramosum), the between-region diversity component is 7% and 89% of the total diversity is stored within populations. Received March 13, 2002; accepted September 24, 2002 Published online: December 11, 2002     

Diploid and autotetraploid sexuals and their relationships to apomicts in the Ranunculus cassubicus group: insights from DNA content and isozyme variation

 From the predominantly aposporous Ranunculus cassubicus group, a subgroup of the R. auricomus complex, two species with both diploid and tetraploid cytodemes (R. cassubicifolius W. Koch and R. carpaticola Soó) were known. Nine population samples of both species have been analyzed for variation of ploidy levels and isozymes. DNA image analysis showed that three populations of R. cassubicifolius from Bavaria and Salzburg are diploid, two from Lower Austria tetraploid. In R. carpaticola, two populations from Central Slovakia and one from Romania are diploid, one from northern Slovakia is hexaploid. The polyploid populations had somewhat smaller C-values than expected from diploids. Ploidy levels are consistent within populations, contradicting previous hypotheses that cycles of diploid-tetra-ploid-dihaploid apomictic individuals occur in natural populations of goldilocks. Isozyme-allozyme analysis of nine polymorphic loci showed that individual variation and genetic measures of di-ploid and tetraploid populations are equivalent to those of sexual taxa. Multiple allelism and unbalanced gene dosages in tetraploid R. cassubicifolius give evidence for autopolyploidy that is most probably of multiple origin. The observed excess of homozygotes and the high diversity between populations in the diploid populations of R. cassubicifolius are most probably due to geographical isolation of population groups, that might have happened during the Würm glaciation and might have promoted the separation of autotetraploids in the easternmost part of the distribution range. Genetic distance values analyzed by UPGMA of all sexual populations separated the two taxa and the cytodemes within R. cassubicifolius. Multidimensional scaling of individuals (including the apomicts) based on a presence/absence matrix of alleles confirmed this differentiation with an overlapping zone between R. cassubicifolius and R. carpaticola. The hexaploid R. carpaticola population showed reduced genotypic diversity, an increased number of heterozygotes and fixed heterozygosity as typical for apomictic mode of reproduction. This population shared alleles of both species without any specific ones, multidimensional scaling placed genotypes among R. cassubicifolius. Thus, the hexa-ploid apomicts might have originated from hybrids of R. cassubicifolius and R. carpaticola. In general, evolution of agamic lineages in the R. cassubicus complex might have been facilitated by autopoly-ploids, because they can provide a bridge between the reproductive systems that are otherwise isolated by ploidy levels, and also a starting point for spontaneous origin of apomixis. Received October 8, 2001; accepted May 10, 2002 Published online: November 7, 2002 Addresses of the authors: Elvia H?randl (E-mail: elvira.hoerandl@univie.ac.at), Department of Systematics and Evolution of Higher Plants; Johann Greilhuber (E-mail: Johann.Greilhuber@univie. ac.at), Department of Systematic Karyology and Embryology of Plants; both: Institute of Botany, University of Vienna, Rennweg 14, A-1030 Vienna, Austria.     

Cochlearia macrorrhiza (Brassicaceae): A bridging species between Cochlearia taxa from the Eastern Alps and the Carpathians?

Cochlearia macrorrhiza is one of the most highly endangered species in Central Europe and less than five individuals survived at its natural stand in a lowland area between the Eastern Alps and the Carpathians. Amplified fragment length polymorphisms (AFLPs) confirmed the status of C. macrorrhiza as a distinct taxon. Lowland C. macrorrhiza does not bridge the distribution of montainous and alpine Cochlearia species from the Eastern Alps and the Carpathians genetically, and C. macrorrhiza represents a separate lineage which evolved from diploid Cochlearia as C. excelsa in East Austrian high alpine regions did. Another species considered in this study, the Romanian C. borzaeana is more closely related to C. tatrae from the High Tatra mountains than to C. pyrenaica from Slovakia or Austria and the AFLP results suggest a single origin of alpine 2n=42 taxa. Genetic differentiation within and between populations is highly structured geographically, and the AFLP data favour a former widespread distribution of C. pyrenaica in mountainous regions and a parallel evolution of high alpine taxa in the Eastern Alps and the Carpathians, respectively.     

Reintroduction of the endangered and endemic plant species Cochlearia bavarica—Implications from conservation genetics

Population reintroduction is a common practice in conservation, but often fails, also due to the effects of inbreeding or outbreeding depression. Cochlearia bavarica is a strongly endangered plant species endemic to Bavaria in Germany, constantly declining since the late 1980s. Therefore, population reintroduction is intended. In this study, we analyzed genetic diversity within and genetic differentiation between all 32 remnant populations of the species in Swabia and Upper Bavaria using amplified fragment length polymorphisms. Our aim was to increase reintroduction success by providing data to avoid negative effects of inbreeding and outbreeding and to preserve the natural genetic pattern of the species. Genetic diversity within populations was low but similar to other rare and endemic species and varied strongly between populations but did not depend on population size. Our analysis revealed a strong geographic pattern of genetic variation. Genetic differentiation was strongest between Swabia and Upper Bavaria and at the population level, whereas differentiation between subpopulations was comparatively low. Isolation by distance and genetic differentiation was stronger among populations from Upper Bavaria than from Swabia. From the results of our study, we derived recommendations for a successful reintroduction of the species. We suggest using rather genetically variable than large populations as reintroduction sources. Moreover, the exchange of plant material between Swabia and Upper Bavaria should be completely avoided. Within these regions, plant material from genetically similar populations should preferably be used for reintroduction, whereas the exchange among subpopulations seems to be possible without a negative impact on genetic variation due to natural gene flow.     

Geographic variation and dispersal history in Fennoscandian populations of two forest herbs

 Carex digitata and Melica nutans are forest understorey herbs with wide European distributions and their northern range margins in Fennoscandia. The species have closely similar habitat requirements, occur in small populations in old forest stands on base-rich to neutral soils and have restricted dispersal abilities at the present day. This study investigates the structure of allozyme variation (12 and 8 loci, respectively) in material of both species (38 and 37 populations, respectively) from throughout southern Sweden and southern Finland. Both species show a relatively low overall genetic diversity (H_T excluding monomorphic loci=0.17 and 0.18, respectively). The hierarchic structuring of allelic diversity in the species is similar, with a relatively high between-population component of diversity (G_ST=0.36 and 0.37, respectively). Neither of the species shows a clear intraspecific pattern of geographic differentiation. The lack of large-scale patterns of geographic differentiation is not consistent with a simple scenario of discrete and independent waves of immigration into Fennoscandia. However, particularly in M. nutans, a group of populations from a lowland belt across southwestern Finland and southern central Sweden is somewhat differentiated from populations to the north and south. A number of rare alleles in both species are widely, but patchily distributed in low frequencies. Hybridization may account for the scattered occurence of some of the rare alleles in Carex digitata, but cannot explain the distribution of rare alleles in Melica nutans. Received July 23, 2001 Accepted December 6, 2001     

Life history variation between species of the relictual genus Borderea (Dioscoreaceae): phylogeography, genetic diversity, and population genetic structure assessed by RAPD markers

The genus Borderea consists of two species, B. pyrenaica and B. chouardii, taxa which have been previously considered as conspecific due to their overall close morphology. These two sole species of the rare genus of Dioscoreaceae are endemic to the Pyrenees (Spain, France). This mountain range likely operated as a refugium for these plants during the last glaciations. B. chouardii is only known from a single population in the Spanish Prepyrenees and has been classified as at risk of extinction in the Red List of Endangered Species (IUCN); B. pyrenaica shows a narrow distribution range in the central Pyrenees and Prepyrenees. We analysed genetic variation, population structure and differentiation in these two taxa using RAPD markers. Our study was conducted on the same seven populations for which very low levels of genetic differentiation were detected previously through allozyme analysis. By contrast, high levels of genetic variability were detected through the RAPD hypervariable markers. Twelve RAPD primers produced 112 distinct bands in the 397 surveyed individuals, totalling 395 different RAPD phenotypes. Only four bands were monomorphic across all samples of Borderea, whereas 21 of the polymorphic bands were species‐specific (20 for B. chouardii, and one for B. pyrenaica). The largest genetic distances were those between the B. chouardii and the B. pyrenaica phenotypes. An analysis of molecular variance showed greater variance between groups (B. chouardii vs. B. pyrenaica, 76.08%) than within groups (3.60%). RAPD band specificity, phenotypic distances, and the partitioning of variance all support the taxonomic separation of the two species. Statistical evaluation of within‐ and among‐population RAPD genetic variability in B. pyrenaica showed that genetic variability was higher within populations (>80%) than among them. No clear pattern of RAPD differentiation could be observed among the six studied populations of this taxon though slight differences in genetic diversity could be observed in the more isolated Prepyrenean populations compared with the more widespread Pyrenean ones. These results suggest a recent postglacial origin of the present B. pyrenaica populations. © 2003 The Linnean Society of London, Biological Journal of the Linnean Society, 2003, 80 , 483–498.     

Evolutionary history of the polyploid complex of Cardamine amara (Brassicaceae): isozyme evidence

 In samples from 56 populations of Cardamine amara, representing four diploid subspecies (subspp. amara, opicii, balcanica, and pyrenaea) and two tetraploid subspecies (subspp. austriaca and olotensis) from different parts of the European distribution area, four enzyme systems with 23 alleles were studied. These data, together with previous morphological and karyological data, suggest that the distribution and variation pattern within the species was strongly influenced by the last glacial period and postglacial migrations. Cardamine amara subsp. pyrenaea is monomorphic for a unique allele, and subsp. balcanica has a unique allele, too which, however, is not fixed in all populations of the taxon. Both taxa seem to be relic ones, although otherwise subsp. balcanica in respect of allelic spectrum much resembles subsp. amara. The other two diploid subspecies, subsp. amara and subsp. opicii, are not characterised by presence of unique alleles but differ in allele frequencies. The two tetraploid subspecies have different evolutionary histories. C. amara subsp. austriaca seems to be an autopolyploid derivative of subsp. amara which colonised open space offered by retreating glaciers in the Eastern Alps. C. amara subsp. olotensis from the Iberian Peninsula represents most probably a polyploid of preglacial time. Received June 22, 2001; accepted May 17, 2002 Published online: September 13, 2002     

Autotetraploids of Vicia cracca show a higher allelic richness in natural populations and a higher seed set after artificial selfing than diploids

Background and Aims

Despite the great importance of autopolyploidy in the evolution of angiosperms, relatively little attention has been devoted to autopolyploids in natural polyploid systems. Several hypotheses have been proposed to explain why autopolyploids are so common and successful, for example increased genetic diversity and heterozygosity and the transition towards selfing. However, case studies on patterns of genetic diversity and on mating systems in autopolyploids are scarce. In this study allozymes were employed to investigate the origin, population genetic diversity and mating system in the contact zone between diploid and assumed autotetraploid cytotypes of Vicia cracca in Central Europe.

Methods

Four enzyme systems resolved in six putative loci were investigated in ten diploid, ten tetraploid and five mixed-ploidy populations. Genetic diversity and heterozygosity, partitioning of genetic diversity among populations and cytotypes, spatial genetic structure and fixed heterozygosity were analysed. These studies were supplemented by a pollination experiment and meiotic chromosome observation.

Key Results and Conclusions

Weak evidence of fixed heterozygosity, a low proportion of unique alleles and genetic variation between cytotypes similar to the variation among populations within cytotypes supported the autopolyploid origin of tetraploids, although no multivalent formation was observed. Tetraploids possessed more alleles than diploids and showed higher observed zygotic heterozygosity than diploids, but the observed gametic heterozygosity was similar to the value observed in diploids and smaller than expected under panmixis. Values of the inbreeding coefficient and differentiation among populations (ρ_ST) suggested that the breeding system in both cytotypes of V. cracca is mixed mating with prevailing outcrossing. The reduction in seed production of tetraploids after selfing was less than that in diploids. An absence of correlation between genetic and geographic distances and high differentiation among neighbouring tetraploid populations supports the secondary contact hypothesis with tetraploids of several independent origins in Central Europe. Nevertheless, the possibility of a recent in situ origin of tetraploids through a triploid bridge in some regions is also discussed.     

Contrasting patterns of spatial genetic structure of diploid and triploid populations of the clonal aquatic species,<Emphasis Type="Italic">Butomus umbellatus (Butomaceae)</Emphasis>, in Central Europe

Genetic diversity in a sample of an aquatic plantButomus umbellatus from 37 localities in Czechia and Slovakia was studied by analyzing six polymorphic loci in three enzymatic systems (SKDH, PGD and AAT). Diversity among ramets was low in eight populations with relatively extensive sampling (only one population possessed more than one multilocus genotype), suggesting high clonality of reproduction in these populations. However, among-population diversity was high: G = 0.782 and 0.881 for the samples of diploid and triploid populations, respectively. Heterozygosity of individual plants averaged over variable loci was also high: H = 0.554 for diploids and 0.453 for triploids. Genetic differentiation among populations was additionally studied using cluster analysis. Several populations of diploids clustered separately from all other populations, whereas another group of diploid populations clustered with some triploid populations, indicating the possibility of relatively recent, probably multiple origin of these triploid populations from their diploid progenitors. Association between matrices of Nei’s genetic distances among populations from different localities and matrices of geographic distances among these localities revealed highly significant correlation for the sample of diploid populations (r = 0.60,P < 0.001) but no significant correlation for the sample of triploid populations (r = 0.02,P = 0.593). These results indicate a spatial structure of diploid populations in accordance with the isolation by distance model, and a random distribution of genotypes among triploid populations ofB. umbellatus.     

Allozyme variation in Carex sect. Digitatae – Evidence of introgression, genetic distinctiveness and evolution of taxa

 A subset of populations of the European taxa of Carex sect. Digitatae, including Carex digitata, C. pallens, C. ornithopoda, C. pediformis ssp. rhizodes and C. humilis have been investigated for allozyme variation. The (presumably) distantly related C. pendula was used as a reference taxon. Carex digitata, C. pallens and C. ornithopoda on the one hand, and C. humilis and C. pediformis on the other, shared the majority of the alleles. Cluster analyses based on genetic distances grouped populations of C. digitata and C. ornithopoda in a mixed subcluster whereas all populations of C. pallens formed a subcluster distinct from the digitata/ornithopoda cluster. This in spite of the fact that C. ornithopoda is morphologically clearly distinct from C. digitata whereas C. pallens is barely distinguishable from the latter. Carex pediformis and C. humilis appeared not more genetically similar to C. digitata than to the reference taxon. It is concluded that 1) C. digitata, C. pallens and C. ornithopoda are closely related and most probably forms a monophyletic group, 2) C. pallens is a genetically distinct species, 3) C. pediformis and C. humilis are not closely related to the rest of C. sect. Digitatae, 4) C. pediformis and C. humilis are relatively closely related, 5) introgression do occur between the investigated species but not to such an extent that it can explain major geographic patterns of variation in C. digitata. Received December 21, 2001; accepted November 14, 2002 Published online: March 24, 2003     

Analysis of the genetic diversity and relationships among and within species of Hippophae (Elaeagnaceae) based on RAPD markers

RAPD markers were used to assess the genetic diversity and inter- and intra-specific relationships of the genus Hippophae L. and to study the correlation between genetic distances and geographic distances among populations of H. rhamnoides ssp. sinensis. The results analyzed by the percentage of polymorphic loci and Shannon information index indicated that a high level of genetic diversity existed both among and within species of the genus Hippophae. In the UPGMA dendrogram, the species or subspecies were clustered into two main groups but not strictly grouped according to sect. Hippophae and sect. Gyantsensis Lian. The multiple regression analysis and Mantel test both indicated a significant correlation between genetic distance and altitude distance among populations of H. rhamnoides ssp. sinensis, and the cluster analysis suggested that the genetic variation among populations of H. rhamnoides ssp. sinensis was linked to their monophyletic origin. Moreover, some degree of genetic differentiation was found among samples collected at different times.     

Morphological differentiation within the Ranunculus cassubicus group compared to variation of isozymes, ploidy levels, and reproductive systems: implications for taxonomy

 In the partly apomictic Ranunculus cassubicus group, a subgroup of the R. auricomus complex, two species were studied by morphometric analyses: R. cassubicifolius W. Koch (with three diploid and two autotetraploid sexual populations), and R. carpaticola Soó (with three diploid sexual populations and a hexaploid apomictic one). Multidimensional scaling analyses (MDS) of individuals, boxplots and cluster analyses of populations revealed a differentiation of R. cassubicifolius and R. carpaticola, whereby in MDS the hexaploid apomictic individuals are partly intermediate between R. cassubicifolius and R. carpaticola. The cytodemes of R. cassubicifolius showed no morphological and only a weak genetic differentiation. A comparison of morphology, isozymes, reproductive system and ploidy levels showed only partly congruence of data sets in respect of grouping populations, thus illustrating the problem to find criteria for a taxonomic concept. A treatment of the apomictic population as a separate group is indicated by all data sets, afterwards R. cassubicifolius and diploid R. carpaticola represent two other well-defined groups. Canonical variate analysis including all characters confirmed the three suggested groups as significantly different and showed that a total of 89.3% of individuals are correctly classified; number of teeth of stem leaf segments and number of petals are the most discriminating characters. Herbarium studies confirm the morphological differentiation yielded from population samples. The three population groups are even better separated in a canonical variate analysis of isozyme data (presence/absence of 25 alleles) of the same material, here 92.6% of individuals are correctly classified. Morphology and isozyme data suggest that the hexaploid apomict originated from hybrids of R. cassubicifolius and diploid R. carpaticola and must be excluded from the sexual taxa; the final classification and naming of the apomicts must be left for further studies on a larger material. The sexual taxa should be classified as separate species. Herbarium studies indicate that R. carpaticola s.str. is widespread over the Carpathians and might include other populations hitherto ascribed to other microspecies as well. Received November 20, 2001; accepted May 10, 2002 Published online: September 13, 2002     

Length polymorphism and allele structure of trinucleotide microsatellites in natural accessions of Arabidopsis thaliana

 The objective of this work was to assess the degree of trinucleotide microsatellite length polymorphism in the selfing species Arabidopsis thaliana. PCR amplifications of 12 microsatellite loci among 49 natural populations revealed between one to eight length variants (alleles) for each locus. The average number of alleles per locus was four and the average genetic diversity index was 0.43. Divergence between length variants was investigated at the nucleotide level. Several observations emerge from the sequence data: (1) for most loci, length polymorphism results only from variations in the number of trinucleotide repeats; (2) for a few others, some variability was noted in the flanking sequences; (3) for compound and interrupted loci containing two arrays of trinucleotide repeats, length variations preferentially affect the longest one. Five of the Arabidopsis thaliana accessions were clearly composed of two sublines. In 2 other accessions, some heterozygous individual plants, probably resulting from recent outcrosses, were found. A phylogenetic tree constructed on the basis of trinucleotide microsatellite allelic diversity shows that genetic relationships among the accessions are not correlated with their geographic origin. Received: 4 November 1997 / Accepted: 3 March 1998     

RAPD variation within and among natural populations of outcrossing willow-leaved foxglove (Digitalis obscura L.)

 Random amplified polymorphic DNA (RAPD) markers were used to assess levels and patterns of genetic diversity in Digitalis obscura L. (Scrophulariaceae), an outcrossing cardenolide-producing medicinal plant species. A total of 50 plants from six natural populations on the Iberian Peninsula were analysed by six arbitrarily chosen decamer primers resulting in 96 highly reproducible RAPD bands. To avoid bias in parameter estimation, analyses of population genetic structure were restricted to bands (35 of 96) whose observed frequencies were less than 1–3/n in each population. The analysis of molecular variance (AMOVA) with distances among individuals corrected for the dominant nature of RAPDs (genotypic analysis) showed that most of the variation (84.8%) occurred among individuals within populations, which is expected for an outcrossing organism. Of the remaining variance, 9.7% was attributed to differences between regions, and 5.5% for differences among populations within regions. Estimates of the Wright, Weir and Cockerham and Lynch and Milligan F_ST from null-allele frequencies corroborated AMOVA partitioning and provided significant evidence for population differentiation in D. obscura. A non-parametric test for the homogeneity of molecular variance (HOMOVA) also showed significant differences in the amount of genetic variability present in the six populations. UPGMA cluster analyses, based on Apostol genetic distance, revealed grouping of some geographically proximate populations. Nevertheless, a Mantel test did not give a significant correlation between geographic and genetic distances. This is the first report of the partitioning of genetic variability within and between populations of D. obscura and provides important baseline data for optimising sampling strategies and for conserving the genetic resources of this medicinal species. Received: 7 September 1998 / Accepted: 28 November 1998     

Microsatellite polymorphism in natural populations of wild emmer wheat,Triticum dicoccoides,in Israel

Diversity in 20 microsatellite loci of wild emmer wheat, Triticum dicoccoides, was examined in 15 populations (135 genotypes) representing a wide range of ecological conditions of soil, temperature, and water availability, in Israel and Turkey. An extensive amount of diversity at microsatellite loci was observed despite the predominantly selfing nature of this plant species. The 20 Gatersleben wheat microsatellites (GWM), representing 13 chromosomes of genomes A and B of wheat, revealed a total of 364 alleles, with an average of 18 alleles per GWM marker (range: 5–26). The proportion of polymorphic loci per population averaged 0.90 (range: 0.45– 1.00); genic diversity, He, averaged 0.50 (range 0.094– 0.736); and Shannon’s information index averaged 0.84 (range 0.166–1.307). The coefficients of genetic distance between populations were high and averaged D=1.862 (range 0.876–3.320), an indication of sharp genetic divergence over short distances. Interpopulation genetic distances showed no association with geographic distance between the population sites of origin, which ruled out a simple isolation by distance model. Genetic dissimilarity values between genotypes were used to produce a dendrogram of the relationships among wild wheat populations by the unweighted pair-group method with arithmetic averages (UPGMA). The results showed that all the wild emmer wheat populations could be distinguished. Microsatellite analysis was found to be highly effective in distinguishing genotypes of T. dicoccoides, originating from diverse ecogeographical sites in Israel and Turkey, with 88% of the 135 genotypes correctly classified into sites of origin by discriminant analysis. Our present microsatellite results are non-random and in agreement with the previously obtained allozyme and RAPD patterns, although the genetic-diversity values obtained with microsatellites are much higher. Significant correlates of microsatellite markers with various climatic and soil factors suggest that, as in allozymes and RAPDs, natural selection causes adaptive microsatellite ecogeographical differentiation, not only in coding, but most importantly in non-coding genomic regions. Hence, the concept of ”junk DNA” needs to be replaced by at least partly regulatory DNA. The obtained results suggest that microsatellite markers are useful for the estimation of genetic diversity in natural populations of T. dicoccoides and for the tagging of agronomically important traits derived from wild emmer wheat. Received: 27 February 2001 / Accepted: 22 March 2001     

Understanding ploidy complex and geographic origin of the Buchloe dactyloides genome using cytoplasmic and nuclear marker systems

Characterizing and inferring the buffalograss [Buchloe dactyloides (Nutt.) Engelm.] genome organization and its relationship to geographic distribution are among the purposes of the buffalograss breeding and genetics program. This buffalograss study was initiated to: (1) better understand the buffalograss ploidy complex using various marker systems representing nuclear and organelle genomes; (2) determine whether the geographic distribution was related to nuclear and organelle genome variation; and (3) compare the genetic structure of accessions with different ploidy levels. The 20 buffalograss genotypes (15 individuals from each genotype) that were studied included diploid, tetraploid, pentaploid, and hexaploid using nuclear (intersimple sequence repeat (ISSRs), simple sequence repeat (SSRs), sequence related amplified polymorphism (SRAPs), and random amplified polymorphic DNA (RAPDs)) and cytoplasmic markers (mtDNA and cpDNA). There was a significant correlation between the ploidy levels and number of alleles detected using nuclear DNA (ISSR, SSR, and SRAP, r=0.39, 0.39, and 0.41, P<0.05, respectively), but no significant correlation was detected when mitochondrial (r=0.17, P<0.05) and chloroplast (r=0.11, P<0.05) DNA data sets were used. The geographic distribution of buffalograss was not correlated with nuclear and organelle genome variation for the genotypes studied. Among the total populations sampled, regression analysis indicated that geographic distance could not explain genetic differences between accessions. However, genetic distances of those populations from the southern portion of buffalograss adaptation were significantly correlated with geographic distance (r= 0.48, P<0.05). This result supports the hypothesis that genetic relationship among buffalograss populations cannot be estimated based only on geographic proximity.     

Comparative analysis of genetic diversity in the mangrove species Avicennia marina (Forsk.) Vierh. (Avicenniaceae) detected by AFLPs and SSRs

Avicennia marina is an important mangrove species with a wide geographical and climatic distribution which suggests that large amounts of genetic diversity are available for conservation and breeding programs. In this study we compare the informativeness of AFLPs and SSRs for assessing genetic diversity within and among individuals, populations and subspecies of A. marina in Australia. Our comparison utilized three SSR loci and three AFLP primer sets that were known to be polymorphic, and could be run in a single analysis on a capillary electrophoresis system, using different- colored fluorescent dyes. A total of 120 individuals representing six populations and three subspecies were sampled. At the locus level, SSRs were considerably more variable than AFLPs, with a total of 52 alleles and an average heterozygosity of 0.78. Average heterozygosity for AFLPs was 0.193, but all of the 918 bands scored were polymorphic. Thus, AFLPs were considerably more efficient at revealing polymorphic loci than SSRs despite lower average heterozygosities. SSRs detected more genetic differentiation between populations (19 vs 9%) and subspecies (35 vs 11%) than AFLPs. Principal co-ordinate analysis revealed congruent patterns of genetic relationships at the individual, population and subspecific levels for both data sets. Mantel testing confirmed congruence between AFLP and SSR genetic distances among, but not within, population comparisons, indicating that the markers were segregating independently but that evolutionary groups (populations and subspecies) were similar. Three genetic criteria of importance for defining priorities for ex situ collections or in situ conservation programs (number of alleles, number of locally common alleles and number of private alleles) were correlated between the AFLP and SSR data sets. The congruence between AFLP and SSR data sets suggest that either method, or a combination, is applicable to expanded genetic studies of mangroves. The codominant nature of SSRs makes them ideal for further population-based investigations, such as mating-system analyses, for which the dominant AFLP markers are less well suited. AFLPs may be particularly useful for monitoring propagation programs and identifying duplicates within collections, since a single PCR assay can reveal many loci at once. Received: 3 October 2000 / Accepted: 19 February 2001