Evolutionary dynamics of mixed-ploidy populations in an annual herb: dispersal, local persistence and recurrent origins of polyploids.

Background and Aims: Despite the recent wealth of studies targeted at contact zones of cytotypes in various species, some aspects of polyploid evolution are still poorly understood. This is especially the case for the frequency and success rate of spontaneous neopolyploidization or the temporal dynamics of ploidy coexistence, requiring massive ploidy screening and repeated observations, respectively. To fill this gap, an extensive study of spatio-temporal patterns of ploidy coexistence was initiated in the widespread annual weed Tripleurospermum inodorum (Asteraceae). Methods: DNA flow cytometry along with confirmatory chromosome counts was employed to assess ploidy levels of 11 018 adult individuals and 1263 ex situ germinated seedlings from 1209 Central European populations. The ploidy screening was conducted across three spatial scales and supplemented with observations of temporal development of 37 mixed-ploidy populations. Key Results: The contact zone between the diploid and tetraploid cytotypes has a diffuse, mosaic-like structure enabling common cytotype coexistence from the within-population to the landscape level. A marked difference in monoploid genome size between the two cytotypes enabled the easy distinction of neotetraploid mutants from long-established tetraploids. Neotetraploids were extremely rare (0·03 %) and occurred solitarily. Altogether five ploidy levels (2 x -6 x ) and several aneuploids were discovered; the diversity in nuclear DNA content was highest in early ontogenetic stages (seedlings) and among individuals from mixed-ploidy populations. In spite of profound temporal oscillations in cytotype frequencies in mixed-ploidy populations, both diploids and tetraploids usually persisted up to the last census. Conclusions: Diploids and tetraploids commonly coexist at all spatial scales and exhibit considerable temporal stability in local ploidy mixtures. Mixed-ploidy populations containing fertile triploid hybrids probaby act as effective generators of cytogenetic novelty and may facilitate inter-ploidy gene flow. Neopolyploid mutants were incapable of local establishment.

Small genomes and large seeds: chromosome numbers, genome size and seed mass in diploid Aesculus species (Sapindaceae).

Background and Aims: Aesculus L. (horse chestnut, buckeye) is a genus of 12-19 extant woody species native to the temperate Northern Hemisphere. This genus is known for unusually large seeds among angiosperms. While chromosome counts are available for many Aesculus species, only one has had its genome size measured. The aim of this study is to provide more genome size data and analyse the relationship between genome size and seed mass in this genus. Methods: Chromosome numbers in root tip cuttings were confirmed for four species and reported for the first time for three additional species. Flow cytometric measurements of 2C nuclear DNA values were conducted on eight species, and mean seed mass values were estimated for the same taxa. Key Results: The same chromosome number, 2 n = 40, was determined in all investigated taxa. Original measurements of 2C values for seven Aesculus species (eight taxa), added to just one reliable datum for A. hippocastanum , confirmed the notion that the genome size in this genus with relatively large seeds is surprisingly low, ranging from 0·955 pg 2C -1 in A. parviflora to 1·275 pg 2C -1 in A. glabra var. glabra. Conclusions: The chromosome number of 2 n = 40 seems to be conclusively the universal 2 n number for non-hybrid species in this genus. Aesculus genome sizes are relatively small, not only within its own family, Sapindaceae, but also within woody angiosperms. The genome sizes seem to be distinct and non-overlapping among the four major Aesculus clades. These results provide an extra support for the most recent reconstruction of Aesculus phylogeny. The correlation between the 2C values and seed masses in examined Aesculus species is slightly negative and not significant. However, when the four major clades are treated separately, there is consistent positive association between larger genome size and larger seed mass within individual lineages.

The intriguing complexity of parthenogenesis inheritance in Pilosella rubra (Asteraceae, Lactuceae).

Neither the genetic basis nor the inheritance of apomixis is fully understood in plants. The present study is focused on the inheritance of parthenogenesis, one of the basic elements of apomixis, in Pilosella (Asteraceae). A complex pattern of inheritance was recorded in the segregating F(1) progeny recovered from reciprocal crosses between the facultatively apomictic hexaploid P. rubra and the sexual tetraploid P. officinarum. Although both female and male reduced gametes of P. rubra transmitted parthenogenesis at the same rate in the reciprocal crosses, the resulting segregating F(1) progeny inherited parthenogenesis at different rates. The actual transmission rates of parthenogenesis were significantly correlated with the mode of origin of the respective F(1) progeny class. The inheritance of parthenogenesis was significantly reduced in F(1) n + n hybrid progeny from the cross where parthenogenesis was transmitted by female gametes. In F(1) n + 0 polyhaploid progeny from the same cross, however, the transmission rate of parthenogenesis was high; all fertile polyhaploids were parthenogenetic. It appeared that reduced female gametes transmitting parthenogenesis preferentially developed parthenogenetically and only rarely were fertilized in P. rubra. The fact that the determinant for parthenogenesis acts gametophytically in Pilosella and the precocious embryogenesis in parthenogenesis-transmitting megagametophytes was suggested as the most probable explanations for this observation. Furthermore, we observed the different expression of complete apomixis in the non-segregating F(1) 2n + n hybrids as compared to their apomictic maternal parent P. rubra. We suggest that this difference is a result of unspecified interactions between the parental genomes.

Expressivity of apomixis in 2n + n hybrids from an apomictic and a sexual parent: insights into variation detected in Pilosella (Asteraceae: Lactuceae).

Reproductive variation was studied in the tetraploid Pilosella aurantiaca, hexaploid P. rubra (both species with facultative autonomous apospory) and in their 2n + n hybrids, which were obtained by crossing with a sexual pollen parent (tetraploid P. officinarum). The different DNA content in P. aurantiaca and P. officinarum demonstrated the actual 2n + n origin, both spontaneous from the field and through experimental crosses, of their hexaploid hybrids. The octoploid 2n + n progeny were recovered from an experimental cross of P. rubra and P. officinarum. The reproductive pathways operating in two maternal facultatively apomictic species and in the hybrids were quantified using a flow cytometric analysis of seeds obtained from either open-pollinated or emasculated plants. Whereas both maternal species displayed a high penetrance of apomixis, the level of apomixis among the majority of 2n + n hybrids was much lower and variable. Some of the hexaploid hybrids had a reduced seed set. Compared to the respective maternal parents, the decrease in apomixis due to haploid parthenogenesis and/or n + n mating was evident in almost all unreduced hybrids, irrespective of their field/experimental origin and ploidy. Hence, the reproductive behaviour in the apomictic maternal parent was profoundly different from that of the 2n + n hybrids with a sexual parent in spite of the preservation of the complete maternal genome in the hybrids. The regulatory interactions in hybrid genomes, such as effects of modifiers, heterochrony, and epigenetic control, may be consistent with the different expressivity of apomixis observed under different genetic backgrounds.

Genome size variation and species relationships in Hieracium sub-genus Pilosella (Asteraceae) as inferred by flow cytometry.

BACKGROUND AND AIMS: Hieracium sub-genus Pilosella (hawkweeds) is a taxonomically complicated group of vascular plants, the structure of which is substantially influenced by frequent interspecific hybridization and polyploidization. Two kinds of species, 'basic' and 'intermediate' (i.e. hybridogenous), are usually recognized. In this study, genome size variation was investigated in a representative set of Central European hawkweeds in order to assess the value of such a data set for species delineation and inference of evolutionary relationships. METHODS: Holoploid and monoploid genome sizes (C- and Cx-values) were determined using propidium iodide flow cytometry for 376 homogeneously cultivated individuals of Hieracium sub-genus Pilosella, including 24 species (271 individuals), five recent natural hybrids (seven individuals) and experimental F(1) hybrids from four parental combinations (98 individuals). Chromosome counts were available for more than half of the plant accessions. Base composition (proportion of AT/GC bases) was cytometrically estimated in 73 individuals. KEY RESULTS: Seven different ploidy levels (2x-8x) were detected, with intraspecific ploidy polymorphism (up to four different cytotypes) occurring in 11 wild species. Mean 2C-values varied approx. 4.3-fold from 3.53 pg in diploid H. hoppeanum to 15.30 pg in octoploid H. brachiatum. 1Cx-values ranged from 1.72 pg in H. pilosella to 2.16 pg in H. echioides (1.26-fold). The DNA content of (high) polyploids was usually proportional to the DNA values of their diploid/low polyploid counterparts, indicating lack of processes altering genome size (i.e. genome down-sizing). Most species showed constant nuclear DNA amounts, exceptions being three hybridogenous taxa, in which introgressive hybridization was suggested as a presumable trigger for genome size variation. Monoploid genome sizes of hybridogenous species were always between the corresponding values of their putative parents. In addition, there was a good congruency between actual DNA estimates and theoretical values inferred from putative parental combinations and between DNA values of experimental F(1) hybrids and corresponding established hybridogenous taxa. CONCLUSIONS: Significant differences in genome size between hawkweed species from hybridogenous lineages involving the small-genome H. pilosella document the usefulness of nuclear DNA content as a supportive marker for reliable delineation of several of the most problematic taxa in Hieracium sub-genus Pilosella (including classification of borderline morphotypes). In addition, genome size data were shown to have a good predictive value for inferring evolutionary relationships and genome constitution (i.e. putative parental combinations) in hybridogenous species.

Reproduction mode in the allopolyploid facultatively apomictic hawkweed Hieracium rubrum (Asteraceae, H. subgen. Pilosella).

The versatility of the breeding system in the hybridogenous hexaploid, Hieracium rubrum, was demonstrated in emasculation and crossing experiments. The flow-cytometric ploidy analysis of 1095 seedlings amongst its' progeny enabled the determination and quantification of the reproductive pathway (apospory, haploid parthenogenesis, cross-fertilization of both reduced and unreduced female gametes) responsible for each progeny class. The progeny profiles were stable between two successive years. The percentage of hybrids arisen from crosses with tetraploid sexual H. pilosella was 7.9%. The rate of trihaploids generated by H. rubrum when crossed to H. pilosella was 3.95%, similar to that of pentaploid (n + n) and octoploid (2n + n) hybrids (3.95% and 3.39%). Unreduced pollen from H. pilosella contributed to hybridization much less frequently (0.56% of all progeny), than unreduced eggs of H. rubrum. The increased frequency of trihaploids formed by crossed compared to emasculated plants suggests an interaction between the presence of pollen and the autonomous development of meiotic embryo sacs. Although the environmental conditions (garden compared to unheated glasshouse) influenced the progeny following the emasculation, no corresponding response to environment was detected amongst the autonomously derived progeny following pollination with H. pilosella. However, there was an influence of pollen parent on progeny which varied under the different environmental treatments. The importance of residual sexuality in reproduction of apomicts is evident. The non-maternal progeny was generated with highly variable frequency especially in the glasshouse, with some capitula reaching up to 50%. Having an autonomous endosperm development, Hieracium subgen. Pilosella is almost unique among other aposporous genera, in which such variable reproduction mode is connected with pseudogamy.

Variation in DNA-ploidy Levels of Reynoutria Taxa in the Czech Republic.

The genus Reynoutria is represented by four taxa in the Czech Republic: Reynoutria japonica var. japonica, R. japonica var. compacta, R. sachalinensis and R. xbohemica. By using flow cytometry, cytological variability within the genus is described based on 257 Reynoutria samples. The varieties of R. japonica are cytologically uniform, var. japonica is exclusively octoploid (2n = 8x = 88) and var. compacta occurs only as a tetraploid (2n = 4x = 44), but R. sachalinensis and R. xbohemica exhibit some variation in chromosome numbers. Reynoutria sachalinensis is predominantly tetraploid (2n = 4 x = 44), but also occurs occasionally as hexaploid and octoploid cytotypes. The most common ploidy level in R. xbohemica is hexaploid (2n = 6x = 66), but tetraploid and octoploid clones were also found. The four taxa occurring in the Czech Republic are described briefly and the possible origins of the cytotypes discussed.