The Detection,Rate and Manifestation of Residual Sexuality in Apomictic Populations of Pilosella (Asteraceae,Lactuceae)

The effect of maternal, facultatively apomictic plants on population diversity was evaluated in seven hybridizing polyploid Pilosella populations, where apomictic (P. bauhini or P. aurantiaca) and sexual (P. officinarum) biotypes coexist. The ploidy level, reproductive system, morphology, clonal structure and chloroplast DNA haplotypes were used to characterize these plants and their hybrids. The reproductive origins of the progeny were assessed through either a flow cytometric seed screen and/or a comparison between the ploidy level of progeny embryos/seedlings and the maternal ploidy level. The cultivated progeny derived from residual sexuality in maternal apomicts were also identified based on their morphology and reproductive behaviour. The progeny different from their maternal parents (0.6?92.3 % of progeny embryos and 0?100 % of progeny seedlings) originated either sexually or via haploid parthenogenesis. Comparing the facultatively apomictic and sexual mothers, the progeny arrays generated in the field showed that apomictic mothers produce progeny that is more variable in ploidy level. This effect was demonstrated at both the embryonic and seedling stages of progeny development. Residual sexuality in apomicts was also effective in experimental crosses, generating progeny similar to spontaneous hybrids in the field. The 2n + n hybrids produced from an apomictic and a sexual parent displayed similar reproductive behaviour, producing polyhaploid, sexual and apomictic progeny in variable ratios. Repeated hybridizations between parental species and/or multi-step crosses can result in hybrid swarms rich in cytotypes and morphotypes. The variation recorded in these populations suggests prevailing introgressive hybridization towards the sexual species P. officinarum.     

Effect of pollen source and pollen ploidy on endosperm formation and seed set in pseudogamous apomictic Paspalum notatum

 It is generally accepted that most angiosperms require an accurate balance between maternal and paternal genome contribution for endosperm development. The endosperm balance number (EBN) hypothesis postulates that each species has an effective number which must be in a 2:1 maternal to paternal ratio for normal endosperm development and seed formation. The aim of this work was to investigate the effect of different sources and ploidy levels of pollen donors on endosperm formation and seed production of aposporous tetraploid (2n=4×=40) Paspalum notatum. Hand-emasculated spikelets of an apomictic 4× plant were dusted with pollen of 2×, 4×, 5×, 6× and 8× races of the same species; 3× and 4× races of a phylogenetically closely related species, P. cromyorrhizon; and 2× and 4× races of P. simplex, a species of a different subgenus. Experiments including self-pollination as well as emasculation without pollination were conducted for controls. Results indicated that apomictic 4×P. notatum is a pseudogamous species with effective fertilization of the two unreduced (2n) polar nuclei by a reduced (n) sperm. Endosperm development and seed production occurred independently of the species or the ploidy level of the pollen donor. However, seed germination rates were significantly lower than in the self-pollinated control when the pollen donor was 3×P. cromyorrhizon or 2× and 4×P. simplex. Aposporous embryo sacs in Paspalum contribute to endosperm formation with two unreduced (2n) polar nuclei, while the male contribution is the same as in sexual plants (n). Since sexual Paspalum plants fit the EBN hypothesis, the EBN insensitivity observed in apomictic plants might be a prerequisite for the spread of pseudogamous apomixis. The EBN insensitivity could have arisen as an imprinting consequence of a high maternal genome contribution. Received: 20 February 1998 / Revision accepted: 21 October 1998     

Reproductive differentiation into sexual and apomictic polyploid cytotypes in Potentilla puberula (Potentilleae, Rosaceae)

Background and Aims

Intraspecific reproductive differentiation into sexual and apomictic cytotypes of differing ploidy is a common phenomenon. However, mechanisms enabling the maintenance of both reproductive modes and integrity of cytotypes in sympatry are as yet poorly understood. This study examined the association of sexual and apomictic seed formation with ploidy as well as gene flow towards sexuals within populations of purely polyploid Potentilla puberula.

Methods

The study is based on 22 populations representing various combinations of five polyploid cytotypes (tetraploid–octoploid) from East Tyrol, Austria. Embryo ploidy and the endosperm/embryo ploidy ratio obtained by a flow cytometric seed screen were used to infer reproductive modes of seed formation and to calculate the male and female genomic contributions to the embryo and endosperm. Self-incompatibility (SI) patterns were assessed and a new indirect approach was used to test for the occurrence of intercytotype matings based on the variation in the male genomic contribution to sexually derived embryos on the level of developed seed.

Key Results

Tetraploids formed seeds almost exclusively via sexual reproduction, whereas penta- to octoploids were preferentially apomictic. Non-random distribution of reproductive modes within maternal plants further revealed a tendency to separate the sexual from the apomictic mode among individuals. Self-incompatibility of sexuals indicated functionality of the gametophytic SI system despite tetraploidy of the nuclear genome. We found no indication for significant cross-fertilization of tetraploids by the high polyploids.

Conclusions

The study revealed a rare example of intraspecific differentiation into sexual and apomictic cytotypes at the polyploid level. The integrity of the sexual tetraploids was maintained due to reproductive isolation from the apomictic higher polyploids. Functionality of the gametophytic SI system suggested that the tetraploids are functional diploids.     

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     

Enriching Ploidy Level Diversity: the Role of Apomictic and Sexual Biotypes of Hieracium subgen. Pilosella (Asteraceae) that Coexist in Polyploid Populations

The capacity to generate variation in ploidy and reproductive mode was compared in facultatively apomictic versus sexual maternal plants that coexist in two model populations. The population structure was studied in polyploid hybrid swarms comprised of Hieracium pilosella (usually sexual, less commonly apomictic), H. bauhini (apomictic), and their hybrids (sexual, apomictic, or sterile). Relationships among established biotypes were proposed on the basis of their DNA ploidy level/chromosome number, reproductive mode and morphology. Isozyme phenotypes and chloroplast DNA haplotypes were assayed in the population that was richer in hybrids. The reproductive origin of seed progeny was identified in both sexual and apomictic mothers, using alternative methods: the karyological, morphological and reproductive characters of the cultivated progeny were compared with those of respective mothers, or flow cytometric seed screening was used. In both populations, the progeny of sexual mothers mainly retained a rather narrow range of ploidy level/chromosome number, while the progeny of facultatively apomictic mothers was more variable. The high-polyploid hybrids, which had arisen from the fertilization of unreduced egg cells of apomicts, mainly produced aberrant non-maternal progeny (either sexually and/or via haploid parthenogenesis). Apparently, such versatile reproduction resulted in genomic instability of the recently formed high-polyploid hybrids. While the progeny produced by both true apomictic and sexual mothers mostly maintained the maternal reproductive mode, the progeny of those ‘versatile’ mothers was mainly sexual. Herein, we argue that polyploid facultative apomicts can considerably increase population diversity.     

GENETICAL POPULATION STRUCTURE IN PLANTS: GENE FLOW BETWEEN DIPLOID SEXUAL AND TRIPLOID ASEXUAL DANDELIONS (TARAXACUM SECTION RUDERALIA)

Levels and distribution of genetic variation were studied in central and western European populations of Taraxacum section Ruderalia containing differing mixtures of sexual diploid and asexual triploid plants. All sexual populations were panmictic with their variation partitioned mainly among populations. Genotypic diversity in triploid samples was very high with few clones widespread and many clones restricted to one or a few populations. Extensive amounts of gene (pollen) flow between the diploid and triploid components of a population were inferred from the following data: (1) the two ploidy levels share all major allozyme polymorphisms; (2) the intrapopulational homogeneity in genic variation between diploids and triploids contrasts strongly with the geographic differentiation at each ploidy level separately; (3) population-unique alleles simultaneously occur at the two ploidy levels; (4) not only sexuals but also asexuals generally simulate Hardy-Weinberg expectations. Most likely, intrapopulational gene exchange occurs bidirectionally by mechanisms such as reductional pollen meiosis in apomictic plants, facultative apomixis, and formation of unreduced gametes in sexuals. Thus, diploid and triploid Taraxacum section Ruderalia are less genetically isolated than has previously been supposed and probably form a cohesive evolutionary unit with the level at which gene pools are shared differing by population.     

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.     

Genetic characterization of apospory in tetraploid Paspalum notatum based on the identification of linked molecular markers

Tetraploid Paspalum notatum (bahiagrass) is a valuable forage grass with aposporous apomictic reproduction. In a previous study, we showed that apospory in bahiagrass is under the control of a single dominant gene with a distorted segregation ratio. The objective of this work was to identify molecular markers linked to apospory in tetraploid P. notatum and establish a preliminary syntenic relationship with the genomic region associated with apospory in P. simplex. A F₁ population of 290 individuals, segregating for apospory, was generated after crossing a completely sexual plant (Q4188) with a natural aposporous apomictic plant (Q4117). The whole progeny was classified as sexual or aposporous by embryo sacs analysis. A bulked segregant analysis was carried out to identify molecular markers co-segregating with apospory. Four hundred RAPD primers, 30 AFLP primers combinations and 85 RFLP clones were screened using DNA from both parental genotypes and aposporous and sexual bulks. Linkage analysis was performed with cytological and genetic information from the complete progeny. Cytoembryological analysis showed 219 sexual and 71 aposporous F₁ individuals. Seven different molecular markers (2 RAPD, 4 AFLP and 1 RFLP) were found to be completely linked to apospory. The RFLP probe C1069, mapping to the telomeric region of the long arm of rice chromosome 12, was one of the molecular markers completely linked to apospory in P. notatum. This marker had been previously associated with apospory in P. simplex. A preliminary map of the chromosome region carrying the apospory locus was constructed.     

Transference of natural diversity from the apomictic germplasm of Paspalum notatum to a sexual synthetic population

Genetic improvement in apomictic forage species has been restricted because of the absence of genetic variability in sexual germplasm with the same ploidy level. Following a new breeding scheme, a sexual synthetic tetraploid population (SSTP) of Paspalum notatum has been generated. The objectives of this work were: (a) to evaluate the genetic variability in SSTP by means of molecular markers, morphologic and agronomic traits, and seed fertility and quality traits and (b) to assess the transference of genetic variability from the apomictic germplasm to the sexual one. Molecular markers revealed a twofold higher level of variability in the SSTP in comparison with the sexual germplasm utilised for its generation, and similar levels with the apomictic ones; moreover, markers showed that most of the variability was inherited from the apomictic germplasm. Morphologic and agronomic traits and seed fertility and quality traits showed high levels of variation in the three groups of genotypes indicating that the new breeding scheme was effective in transferring variability from the apomictic germplasm to the SSTP. This new population will be useful in breeding of P. notatum, and the breeding scheme used for its generation may be used in other apomictic species.     

The chromosome segment related to apomixis in Paspalum simplex is homoeologous to the telomeric region of the long arm of rice chromosome 12

Apomixis is a form of asexual reproduction that in plants leads to the production of seed progeny that are exact copies of the mother individual. A mapping population generated by backcrossing a sexual with an apomictic genotype of Paspalum simplex, both at the tetraploid level, was used to find markers co-segregating with apomixis. Genetic analysis showed that apomixis is under the control of a single dominant allele assuming a random chromatid assortment. Five rice markers, mapped in the telomeric region of the long arm of rice chromosome 12, showed tight linkage with apomixis. Genetic and molecular data strongly indicate that the potentiality to express apomixis in P. simplex is given by a relatively large chromosome segment that is inherited as a single genetic unit.     

Molecular cytogenetics and DNA sequence analysis of an apomixis-linked BAC in Paspalum simplex reveal a non pericentromere location and partial microcolinearity with rice

Apomixis in plants is a form of clonal reproduction through seeds. A BAC clone linked to apomictic reproduction in Paspalum simplex was used to locate the apomixis locus on meiotic chromosome preparations. Fluorescent in situ hybridisation revealed the existence of a single locus embedded in a heterochromatin-poor region not adjacent to the centromere. We report here for the first time information regarding the sequencing of a large DNA clone from the apomixis locus. The presence of two genes whose rice homologs were mapped on the telomeric part of the long arm of rice chromosome 12 confirmed the strong synteny between the apomixis locus of P. simplex with the related area of the rice genome at the map level. Comparative analysis of this region with rice as representative of a sexual species revealed large-scale rearrangements due to transposable elements and small-scale rearrangements due to deletions and single point mutations. Both types of rearrangements induced the loss of coding capacity of large portions of the “apomictic” genes compared to their rice homologs. Our results are discussed in relation to the use of rice genome data for positional cloning of apomixis genes and to the possible role of rearranged supernumerary genes in the apomictic process of P. simplex. Ornella Calderini and Song B. Chang have contributed equally to this article     

Ploidy levels and reproductive behaviour in natural populations of five Paspalum species

Knowledge of variation in ploidy levels and reproductive behaviour in natural populations is essential in order to understand the functioning of agamic complexes. The aim of this study was to analyse the ploidy level and mode of reproduction in several wild Paspalum populations. A total of 19 populations representing five different species (P. alcalinum, P. denticulatum, P. lividum, P. nicorae, and P. rufum) were collected. Ploidy level was determined in 1,187 individuals by using flow cytometry. Among these individuals, 2x, 3x, 4x, 5x, 6x, and 7x chromosome constitutions were observed. Diploid sexual cytotypes of P. denticulatum were detected for the first time; this will allow the development of future breeding strategies for this particular species. Flow cytometry seed screen (FCSS) in bulked and single seeds revealed the reproductive diversity of these species, ranging from complete sexuality in diploids and varying levels of facultative apomixis in most tetraploids, to obligate apomixis in pentaploids and hexaploids. A fully sexual tetraploid plant was never detected. Nevertheless, most tetraploid genotypes produced both maternal (by apomixis) and non-maternal (by sexuality) progeny. This residual sexuality is very interesting from an evolutionary point of view, since it would allow the creation of new genotypic combinations in natural populations. In addition, the residual sexuality found in some apomictic tetraploid populations can be used as a source of variability for genetic improvement.     

Stomatal size,ploidy and polyembryony in Eriotheca Stellate Trichome Species Complex (Bombacoideae – Malvaceae) in the Cerrados of Brazil

• Geographic parthenogenesis, range expansion of apomictic plants after climate changes, has been described for Northern Hemisphere gametophytic apomicts. But similar trends have been observed for sporophytic apomicts of Cerrado, the savannas in Brazil. Eriotheca pubescens is a common Cerrado tree, an agamic complex of either hexaploid/polyembryonic apomicts or tetraploid/monoembryonic sexual individuals. Some populations have been described as a new species, Eriotheca estevesiae, all included in the Eriotheca Stellate Trichome Species Complex (ESTSC). Since breeding systems and ploidy are clearly associated with polyembryony and stomatal size, we used these ancillary features to map the reproductive and ploidy level traits of E. pubescens and E. estevesiae.
• Leaves and seeds were collected from individuals of 19 populations. Seeds were evaluated for the presence of polyembryony and leaves for stomatal measurements.
• Eight populations were monoembryonic while another eight were polyembryonic and for other three, the embryonic pattern was not readily verified. E. pubescens polyembryonic and hexaploid populations formed a homogeneous group, but monoembryonic plants were more variable. E. estevesiae populations were monoembryonic with smaller stomata. In contrast, some E. pubescens monoembryonic populations further south presented larger stomata. Despite these outliers, possibly mixed populations, stomatal size and embryonic pattern differed from northern to southern populations.
• Embryonic pattern and stomatal size indicated that northernmost populations of Eriotheca STSC (E. estevesiae) are diploid and sexual. Southernmost populations, mostly polyembryonic and with large stomata, are hexaploid and apomictic. This is in agreement with geographic parthenogenesis and range expansion of apomictic lineages to southern habitats available after the last glacial maximum.

     

Selection on apomictic lineages of Taraxacum at establishment in a mixed sexual–apomictic population

A species’ mode of reproduction, sexual or asexual, will affect its ecology and evolution. In many species, asexuality is related to polyploidy. In Taraxacum, apomicts are triploid, and sexuals are diploid. To disentangle the effects of ploidy level and reproductive mode on life‐history traits, we compared established apomictic Taraxacum genotypes with newly synthesized apomictic genotypes, obtained from diploid–triploid crosses. Diploid–triploid crossing is probably the way that most apomictic lineages originate. New genotypes had on average a much lower seed set than established genotypes. Established genotypes differed on average from new genotypes, in particular under shaded conditions: the established genotypes had longer leaves and flowered later. The differences between new and established triploids resembled the differences that have been found between sexual diploids and established apomictic triploids. We conclude that ploidy differences alone are not directly responsible for observed differences between sexual diploid and apomictic triploid dandelions.     

Sexual tetraploid and apomictic pentaploid populations ofHieracium pilosella (Compositae)

Five species are recognized inHieracium subgen.Pilosella sect.Pilosellina Fries. Four are diploid (2x, 2n = 18), one (H. pilosella L.) is highly variable morphologically and cytologically (from 2x to 10x), in its mode of reproduction (self-incompatibility, agamospermy, amphimixis, apo-amphimixis) and in its hybridization pattern. A part of this huge agamic complex was analysed by comparing sexual 4x and apomictic 5x plants (crossing and germination experiments, measurements of vegetative reproduction by stolons etc.). In the experimental garden apomictic 5x produced more stolons than the sexual 4x plants and the total length of the stolons per rosette was greater. However, in nature, the competitive potential of the sexual plants seems to be higher, presumably as a result of the higher mortality of ramets in 5x. Sexual 4x plants often grow in dense and grazed grass vegetation, whereas 5x apomicts often occur in dunes with patchy vegetation. Apomicts produce more capitula per rosette, and sexual rosettes form only about 60% of the number of viable achenes as compared to apomictic ones. Therefore, apomicts appear to be characterized by a greater colonizing ability than sexual plants. Apomictic plants produce equal numbers of viable achenes under conditions of both open pollination and isolation. Sexual plants do not form any viable achenes after isolation and produce a somewhat lower percentage of achenes after open pollination than do apomictics. 5xreproduce exclusively apomictically. Apo-amphimixis was never observed in pentaploids and only very rarely in tetraploids. Addition hybrids are very rare. The cross sexual 4x × apomictic 5x failed in 70% of the attempts, but the recombination of genomes carrying genes for apomixis is possible and results in apomictic 4x and sexual 5x, both with a reduced number of viable achenes. In nature sexual and apomictic plants may occur in close proximity. In such cases the germination rate of the achenes of 4x and 5x is lower; this may indicate that apomictic plants fertilize sexual plants in nature (unidirectional gene-flow). 5x plants form euploid gametes carrying two or three genomes. The results of the crossing experiments can be explained in terms ofNogler's theory of monogenic inheritance of apospory.Variation and evolution inHieracium subg.Pilosella sect.Pilosellina I.     

When sexual meets apomict: genome size,ploidy level and reproductive mode variation of Sorbus aria s.l. and S. austriaca (Rosaceae) in Bosnia and Herzegovina

Background and Aims Allopolyploidy and intraspecific heteroploid crosses are associated, in certain groups, with changes in the mating system. The genus Sorbus represents an appropriate model to study the relationships between ploidy and reproductive mode variations. Diploid S. aria and tetraploid apomictic S. austriaca were screened for ploidy and mating system variations within pure and sympatric populations in order to gain insights into their putative causalities.Methods Flow cytometry was used to assess genome size and ploidy level among 380 S. aria s.l. and S. austriaca individuals from Bosnia and Herzegovina, with 303 single-seed flow cytometric seed screenings being performed to identify their mating system. Pollen viability and seed set were also determined.Key Results Flow cytometry confirmed the presence of di-, tri- and tetraploid cytotype mixtures in mixed-ploidy populations of S. aria and S. austriaca. No ploidy variation was detected in single-species populations. Diploid S. aria mother plants always produced sexually originated seeds, whereas tetraploid S. austriaca as well as triploid S. aria were obligate apomicts. Tetraploid S. aria preserved sexuality in a low portion of plants. A tendency towards a balanced 2m : 1p parental genome contribution to the endosperm was shared by diploids and tetraploids, regardless of their sexual or asexual origin. In contrast, most triploids apparently tolerated endosperm imbalance.Conclusions Coexistence of apomictic tetraploids and sexual diploids drives the production of novel polyploid cytotypes with predominantly apomictic reproductive modes. The data suggest that processes governing cytotype diversity and mating system variation in Sorbus from Bosnia and Herzegovina are probably parallel to those in other diversity hotspots of this genus. The results represent a solid contribution to knowledge of the reproduction of Sorbus and will inform future investigations of the molecular and genetic mechanisms involved in triggering and regulating cytotype diversity and alteration of reproductive modes.     

Phylogeography of Eriotheca species complex: insights into the origin and range expansion of apomictic and polyploid trees in Neotropical Savannas

• Polyploidy and whole genome duplication are major evolutionary drivers in plants. Climate variations during the Pleistocene have influenced distribution and range expansion worldwide. Similar trends have been reported for Cerrado plants, but no attempt has been made to link phylogeography with ploidy and breeding changes. Thus, we aimed to (i) assess ploidy and genome size of Eriotheca estevesiae Carv.-Sobr., and compare it with E. pubescens (Mart.) Schott & Endl. (Both included into the Eriotheca Stellate Trichome Species Complex – ESTSC). (ii) Subsequently, we investigated their phylogeography to see whether genetic structure and range expansion trends were similar to those previously described for the Cerrado biome. Finally (iii), we discuss whether ESTSC phylogeographic patterns could be associated with geographic parthenogenesis processes.
• Common cytogenetic techniques and flow cytometry were used to confirm chromosome number and genome size of E. estevesiae. We used three cpDNA regions to analyse 14 ESTSC Cerrado populations, for which we also obtained ploidy level and breeding information. We investigated haplotype diversity, population structure and tested neutrality, aiming to reconstruct phylogeographic scenarios.
• We found three ploidy levels and eight cpDNA haplotypes in ESTSC, one shared by most populations. Haplotype and ploidy distribution corroborated that E. pubescens, the widely distributed polyploid and apomictic species, may have originated from northern diploid and probably sexual E. estevesiae.
• Matrilinear cpDNA links support the idea that apomixis and polyploidy in ESTSC may have allowed range expansion during the Pleistocene, in a process analogous to the geographic parthenogenesis described elsewhere.

     

A molecular map of the apomixis-control locus in <Emphasis Type="Italic">Paspalum procurrens</Emphasis> and its comparative analysis with other species of <Emphasis Type="Italic">Paspalum</Emphasis>

Since apomixis was first mapped in Paspalum, the absence of recombination that characterizes the related locus appeared to be the most difficult bottleneck to overcome for the dissection of the genetic determinants that control this trait. An approach to break the block of recombination was developed in this genus through an among-species comparative mapping strategy. A new apomictic species, P. procurrens (Q4094) was crossed with a sexual plant of P. simplex and their progeny was classified for reproductive mode with the aid of morphological, embryological and genetic analyses. On this progeny, a set of heterologous rice RFLP markers strictly co-segregating in coupling phase with apomixis was identified. These markers were all located on the telomeric region of the long arm of the chromosome 12 of rice. In spite of the lack of recombination exhibited by the apomixis-linked markers in P. procurrens, a comparative mapping analysis among P. simplex, P. malacophyllum, P. notatum and P. procurrens, allowed us to identify a small group of markers co-segregating with apomixis in all these species. These markers bracketed a chromosome region that likely contains all the genetic determinants of apomictic reproduction in Paspalum. The implications of this new inter-specific approach for overcoming the block of recombination to isolate the genetic determinants of apomixis and gain a better comprehension of genome structure of apomictic chromosome region are discussed.     

Cytogeography of Pilosella officinarum (Compositae): altitudinal and longitudinal differences in ploidy level distribution in the Czech Republic and Slovakia and the general pattern in Europe

BACKGROUND AND AIMS: Pilosella officinarum (syn. Hieracium pilosella) is a highly structured species with respect to the ploidy level, with obvious cytogeographic trends. Previous non-collated data indicated a possible differentiation in the frequency of particular ploidy levels in the Czech Republic and Slovakia. Therefore, detailed sampling and ploidy level analyses were assessed to reveal a boundary of common occurrence of tetraploids on one hand and higher ploids on the other. For a better understanding of cytogeographic differentiation of P. officinarum in central Europe, a search was made for a general cytogeographic pattern in Europe based on published data. METHODS: DNA-ploidy level and/or chromosome number were identified for 1059 plants using flow cytometry and/or chromosome counting on root meristem preparations. Samples were collected from 336 localities in the Czech Republic, Slovakia and north-eastern Hungary. In addition, ploidy levels were determined for plants from 18 localities in Bulgaria, Georgia, Ireland, Italy, Romania and Ukraine. KEY RESULTS: Four ploidy levels were found in the studied area with a contrasting pattern of distribution. The most widespread cytotype in the western part of the Czech Republic is tetraploid (4x) reproducing sexually, while the apomictic pentaploids and mostly apomictic hexaploids (5x and 6x, respectively) clearly prevail in Slovakia and the eastern part of the Czech Republic. The boundary between common occurrence of tetraploids and higher ploids is very obvious and represents the geomorphologic boundary between the Bohemian Massif and the Western Carpathians with the adjacent part of Pannonia. Mixed populations consisting of two different ploidy levels were recorded in nearly 11% of localities. A statistically significant difference in a vertical distribution of penta- and hexaploids was observed in the Western Carpathians and the adjacent Pannonian Plain. Hexaploid populations tend to occur at lower elevations (usually below 500 m), while the pentaploid level is more or less evenly distributed up to 1000 m a.s.l. For the first time the heptaploid level (7x) was found on one site in Slovakia. In Europe, the sexual tetraploid level has clearly a sub-Atlantic character of distribution. The plants of higher ploidy level (penta- and hexa-) with mostly apomictic reproduction prevail in the northern part of Scandinavia and the British Isles, the Alps and the Western Carpathians with the adjacent part of Pannonia. A detailed overview of published data shows that extremely rare records on existence of diploid populations in the south-west Alps are with high probability erroneous and most probably refer to the closely related diploid species P. peleteriana. CONCLUSIONS: The recent distribution of P. officinarum in Europe is complex and probably reflects the climatic changes during the Pleistocene and consequent postglacial migrations. Probably both penta- and hexaploids arose independently in central Europe (Alps and Carpathian Mountains) and in northern Europe (Scandinavia, Great Britain, Ireland), where the apomictic plants colonized deglaciated areas. We suggest that P. officinarum is in fact an amphidiploid species with a basic tetraploid level, which probably originated from hybridizations of diploid taxa from the section Pilosellina.     

Inheritance of parental genomes in progenies of Poa pratensis L. from sexual and apomictic genotypes as assessed by RAPD markers and flow cytometry

 Moving gene(s) responsible for the apomictic trait into crop plants that naturally reproduce through a sexual process would open up new areas in plant breeding and agricultural systems. Kentucky bluegrass (Poa pratensis L.) is one of the most important forage and turf grasses in temperate climates. It reproduces through facultative aposporous parthenogenesis, but the reproductive behaviour ranges naturally from nearly obligate apomixis to complete sexuality. In addition to apomictic reproduction, sexual hybridization may take place. Selfing may also occur, and occasionally reduced egg cells may develop through parthenogenesis generating (poly)haploids. The inheritance of parental genomes was assessed in Kentucky bluegrass progenies by employing RAPD markers in combination with flow cytometry (FCM). Nine progenies from different crosses carried out between completely sexual and highly apomictic genotypes were evaluated in order to probe the reproductive behaviour of the mother plants and to distinguish the different classes of aberrant plants. Not only were maternals and balanced B_II hybrids recorded, but so were (poly)triploid B_III hybrids, selfs, and (poly)haploids. The application of these techniques demonstrated that FCM analysis accurately distinguishes the n, 2n, and 3n ploidy levels of progenies, and that RAPD markers unequivocally recognize progenies of apomictic and hybrid origin. The occurrence of aneusomaty was documented in one of the selected sexual genotypes, whose crossed progeny plants manifested two distinct classes of ploidy. The nomenclature B_I was adopted to refer to hybrids with a hypodiploid nuclear condition. On the whole, the FCM analysis confirmed most of the RAPD data. The combined evaluation of DNA markers and DNA contents proved to be an efficient screening tool for scoring maternal plants, assessing the genetic origin of aberrant plants, and quantifying the inheritance of parental genomes in Kentucky bluegrass. Hybrid populations from sexual×apomictic matings that segregate for the mode of reproduction represent a valuable basis for attempting to identify molecular markers linked to the apomixis gene(s). Received: 11 November 1996/Accepted: 22 November 1996