Variation of Essential Oil Content and Composition,Phenolics, and Yield Related Traits Using Different Pollination Systems in Populations of Thymus Species

The production of self-pollinated plants could be important for improving medicinal plants secondary metabolites. In this study, 11 Thymus populations from eight species were evaluated to determine the effect of self and open pollination on agro-morphological characteristics, total phenolic content (TPC), essential oil (EO) content, and EO components. Inbreeding led to some positive effects of above mentioned traits in most of the studied populations. Total phenolic content ranged from 7.07 to 52.69 mg tannic acid equivalents (TAE) g⁻¹ dry weight (DW) in open pollinated derived populations, while it varied from 1.2 to 55.03 mg TAE g⁻¹ DW in self-pollinated ones. Under open and self-pollination condition, the highest EO content was obtained in T. trautvetteri (3.37 %) and T. pubescens (1.96 %), respectively. Gas chromatography-mass spectrometry (GC/MS) identified 42 compounds including thymol, carvacrol, linalool, p-cymene, γ-terpinene, terpinen-4-ol, and α-terpineol as the main compounds. In most cases, selfed plants compared to open pollinated ones, revealed higher thymol content. T. daenensis-1 showed a significant increase in thymol content (from 25.22 % to 74.3 %) due to self-pollination. Moreover, self-pollination led to emergence of some new compounds. Carvacrol methyl ether was the constituents of Thymus EO that are being reported in self-pollinated populations. Finally, inbreeding in Thymus might be suggested as a useful tool to increase genetic homogeneity for the selection of superior plants for improving secondary metabolite.     

Volatile Compounds in Thymus sect. Teucrioides (Lamiaceae): Intraspecific and Interspecific Diversity,Chemotaxonomic Significance and Exploitation Potential

Thymus sect. Teucrioides comprises three species, namely, T. hartvigii, T. leucospermus, and T. teucrioides, distributed in Greece and Albania. The volatile constituents of all species of the section were obtained by hydrodistillation and investigated by GC‐FID and GC/MS analyses. Twenty populations were sampled and a total of 103 compounds were identified, representing 98.0–99.9% of the oil compositions. The oils were mainly characterized by high contents of monoterpene hydrocarbons (42.7–92.4%), with the exception of three oils for which oxygenated monoterpenes were the dominating constituents, viz., that of T. hartvigii ssp. macrocalyx, with linalool as main compound (89.2±0.5%), and those of T. hartvigii ssp. hartvigii and of one population of T. teucrioides ssp. candilicus, containing thymol as major component (46.4±3.1 and 38.2±3.9%, resp.). The most common compound in the oils of the 20 populations of the section was p‐cymene. Considerable variation was detected within and among populations, and seven chemotypes were distinguished, i.e., p‐cymene, linalool, p‐cymene/thymol, p‐cymene/γ‐terpinene, p‐cymene/borneol, p‐cymene/γ‐terpinene/borneol, and p‐cymene/linalool chemotypes. Different chemotypes may exist in the same population. Multivariate statistical analyses enabled the segregation of the oils within Thymus sect. Teucrioides into two groups, one consisting of the three subspecies of T. teucrioides and the second comprising the species T. hartvigii and T. leucospermus. A linalool‐rich chemotype, unique within the section, distinguished the oil of T. hartvigii ssp. macrocalyx from all other oils. The high oil content of p‐cymene and the preference for serpentine substrates render T. teucrioides species promising for future exploitation.     

Genetic Diversity and Chemical Polymorphism of Some Thymus Species

To ascertain whether there are chemical and genetic relationships among some Thymus species and also to determine correlation between these two sets of data, the essential‐oil composition and genetic variability of six populations of Thymus including: T. daenensis ?elak. (two populations), T. fallax Fisch . & C.A.Mey ., T. fedtschenkoi Ronniger , T. migricus Klokov & Des .‐Shost ., and T. vulgaris L. were analyzed by GC and GC/MS, and also by randomly amplified polymorphic DNA (RAPD). Thus, 27 individuals were analyzed using 16 RAPD primers, which generated 264 polymorphic scorable bands and volatiles isolated by distillation extraction were subjected to GC and GC/MS analyses. The yields of oils ranged from 2.1 to 3.8% (v/w), and 34 components were identified, amounting to a total percentage of 97.8–99.9%. RAPD Markers allowed a perfect distinction between the different species based on their distinctive genetic background. However, they did not show identical clustering with the volatile‐oil profiles.     

Karyotype analysis and DNA content in some species of Chrysolaena (Vernonieae,Asteraceae)

Cytogenetic characterization by karyotyping and determination of DNA content by flow cytometry of five species of Chrysolaena (Vernonieae, Asteraceae) was performed. This is the first study of nuclear DNA content realized in the genus. The 2C-values were compared with the ploidy level and the total karyotype length (TKL) of each species. Mitotic analysis revealed a base chromosome number x = 10 for all entities and different ploidy levels, from diploid (2n = 2x = 20) to octoploid (2n = 8x = 80). All species showed bimodal karyotypes composed of metacentric and submetacentric chromosomes. The average chromosome size (ML) varied from 1.86 μm to 2.70 μm, while the TKL ranged from 18.65 μm to 80.55 μm. The intrachromosomal asymmetry index (A₁) varied from 0.27 to 0.38, while the interchromosomal asymmetry index (A₂) ranged from 0.19 to 0.25. A new cytotype is reported for the first time for C. propinqua. Accessory chromosomes found in C. verbascifolia, C. cognata, C. flexuosa, and C. propinqua are also reported as new.     

Relationships among genetic distance,forage yield and heterozygosity in isogenic diploid and tetraploid alfalfa populations

Isogenic diploid and tetraploid alfalfa (Medicago sativa L.) was studied with molecular markers to help understand why diploid performance and breeding behavior does not always predict that of tetraploids. In a previous study of partially heterozygous alfalfa genotypes, we detected a low correlation between yields of isogenic diploid (2x) and tetraploid (4x) single-cross progenies, and genetic distances were more highly correlated with yields of tetraploids than diploids. These differences may be related to the level of RFLP heterozygosity expected among progenies derived from heterozygous parents at the two ploidy levels. The objectives of this study were to determine the relationships among genetic distance, forage yield and heterozygosity in isogenic 2 x and 4 x alfalfa populations. Four diploid genotypes were chromosome doubled to produce corresponding isogenic autotetraploids, and these genotypes were mated in 4 × 4 diallels to produce 6 single-cross families at each ploidy level for field evaluation. Allele compositions of parents were determined at 33 RFLP loci by monitoring segregation of homologous restriction fragments among individuals within progenies, and these were used to estimate RFLP heterozygosity levels for all single-cross progenies at both ploidy levels. RFLP heterozygosity rankings were identical between progenies of isogenic diploid and tetraploid parents; but significant associations (P < 0.05) between estimated heterozygosity levels and forage yield were detected only at the tetraploid level. Since tetraploid families were nearly 25% more heterozygous than the corresponding diploid families, inconsistencies in the association between molecular marker diversity and forage yields of isogenic 2 x and 4 x single crosses may be due to recessive alleles that are expressed in diploids but masked in tetraploids. The gene action involved in heterosis may be the same at both ploidy levels; however, tetraploids benefit from greater complementary gene interactions than are possible for equivalent diploids. Present address: AgResearch Grasslands, New Zealand Pastoral Agriculture Research Institute, Palmerston North, New Zealand     

Do polyploids require proportionally less rDNA loci than their corresponding diploids? Examples from Artemisia subgenera Absinthium and Artemisia (Asteraceae,Anthemideae)

Abstract

Fluorescent in situ hybridisation (FISH) of 5S and 18S-5.8S-26S ribosomal DNA was carried out in two species of the genus Artemisia, belonging to the subgenera Artemisia (A. medioxima) and Absinthium (A. lagocephala), each one showing both low and high ploidy levels (2x, 4x and 16x, and 2x and 6x, respectively). Both species have a base chromosome number of x = 9. Linkage of both rDNA genes has been observed confirming previous results. Diploid A. lagocephala (2n = 18) shows three rDNA loci, and the hexaploid six. Also in A. medioxima, the number of rDNA loci does not increase in the proportion given by the ploidy level, and a relative loss is found. In this species, the diploid population shows two rDNA loci, the tetraploid four, and the hexaidecaploid has around 20. The results evidence a relative loss of rDNA loci and heterochromatin, a phenomenon that is more pronounced at higher ploidy levels. Nevertheless, the DAPI banding pattern of A. lagocephala does not follow this trend, as it shows a spectacular increase of heterochromatic bands at the hexaploid level. These results are discussed in the light of possible chromosome restructuring and gene silencing mechanisms that take place during polyploidy, and more especially allopolyploid formation.     

Diploid endosperm formation in Tulipa spp. and identification of a 1:1 maternal-to-paternal genome ratio in endosperms of T. gesneriana L.

Most Liliaceae plants have the tetrasporic Fritillaria-type embryo sac and normally form diploid embryos and pentaploid endosperms derived from a 4:1 maternal-to-paternal genome ratio (4m:1p) after double fertilization. Here we characterize embryo sac and endosperm formation in Tulipa spp. of Liliaceae. Chromosome analysis using seeds derived from 2x × 2x crosses of Tulipa gesneriana (2n = 2x = 24) identified diploid chromosome number in the endosperm. Similarly, flow cytometric analysis confirmed diploid endosperm formation in T. gesneriana, T. fosteriana (2n = 2x = 24) and T. greigii (2n = 2x = 24). To further study the possible mechanism of diploid endosperm formation, we made interploidy crosses of triploid (2n = 3x = 36) × diploid in which aneuploid seeds with various chromosome numbers (2n = 25–36) were produced. Again, flow cytometric analysis confirmed the same ploidy level in both embryos and endosperms at all aneuploidy levels, suggesting that only a single haploid polar nucleus contributes to endosperm formation at fertilization. Histological observation further confirmed the physical separation of two polar nuclei by a large vacuole in the Fritillaria-type embryo sac of T. gesneriana that appeared to prevent the fusion of the two polar nuclei that originated at the micropylar and chalazal ends before fertilization. Taken together, these results indicate that diploid endosperms (1m:1p) are normally formed in Tulipa spp. by fusion of the micropylar polar nucleus (n) and a spermatid (n) but not by normal triple fusion. We also show that tulip endosperm partially overcomes the triploid block mechanism that occurs in interploidy crosses. Based on these observations, the possible role of triple nuclear fusion in double fertilization is discussed.     

Comparative analysis of essential oil compositions in seven populations of Bakhtiarian savory in natural and field conditions

Bakhtiarian savory (Satureja bachtiarica Bunge) is an endemic herb growing wild in Iran with interesting pharmacological and biological properties. In this research, the variability of essential oil content and phytochemical compositions among seven populations of this species was studied in both natural and field conditions. There was a wide interpopulation variability in phytochemical variation of studied populations under both natural and field conditions, indicating the existence of different chemotypes. Six chemical compositions showed high value in studied plants so that carvacrol and thymol were known as major compounds and confirmed by principal component analysis. Ilam population had the highest thymol in both field (77.10 %) and natural (42.32 %) conditions and was known as a thymol chemotype. Beside, Yazd and Shahrekord populations, respectively, showed the highest carvacrol percentages in both field (84.83 and 63.81 %) and natural (72.50 and 58.05 %) conditions and were known as carvacrol chemotypes. In other populations, carvacrol and thymol were the main components and showed partly equal percentage and these populations were known as carvacrol/thymol chemotypes. According to Pearson correlation analysis, carvacrol was negatively correlated with p-cymene, γ-terpinene and thymol (r = ?0.61, ?0.70 and ?0.98, respectively), indicating very high reverse correlation between carvacrol and thymol. In conclusion, the high value of the essential oil compositions in studied S. bachtiarica populations could provide useful information for conservation and selection of cross parents in breeding programs to develop improved cultivars.     

In vitro induction and identification of tetraploid plants of <Emphasis Type="Italic">Paulownia tomentosa</Emphasis>

Polyploidization is a major trend in plant evolution that has many advantages over diploid. In particular, the enlargement and lower fertility of polyploids are very attractive traits in forest tree breeding programs. We report here a system for the in vitro induction and identification of tetraploid plants of Paulownia tomentosa induced by colchicine treatment. Embryonic calluses derived from placentas were transferred to liquid Murashige and Skoog (MS) medium containing different concentrations of colchicine (0.01, 0.05, or 0.1%) and incubated for 24, 48, or 72 h on an orbital shaker at 110 rpm. The best result in terms of the production of tetraploid plantlets was obtained in the 48 h + 0.05% colchicine treatment, with more than 100 tetraploid plantlets being produced. The ploidy level of plantlets was verified by chromosome counts, flow cytometry, and morphology. The chromosome number of tetraploids was 2n = 4x = 80 and that of diploid plantlets was 2n = 2x = 40. The relative fluorescence intensity of tetraploids was twofold higher than that of diploids. The tetraploid and diploid plantlets differed significantly in leaf shape, with those of the former being round and those of the latter pentagonal. The mean length of the stomata was longer in tetraploid plants than diploid plants, and stomatal frequency was reduced with the increased ploidy level. The tetraploids had large floral organs that were easily distinguishable from those of diploid plants.     

Essential Oil Composition of Five Thymus Species from Bulgaria

In this study, the essential oil composition of five Thymus species, belonging to Sect. Hyphodromi (A. Kerner) Halácsy – Thymus atticus Čelak., T. leucotrichus Halácsy, T. striatus Vahl, T. zygioides Griseb. and T. perinicus (Velen.) Jalas. was studied by GC/MS/FID. T. atticus, T. leucotrichus, and T. striatus were characterized by high amounts of sesquiterpenoids (57.7, 78.9 and 79.7 %, respectively) with β-caryophyllene and caryophyllene oxide as the main constituents. Aromatic compounds (61.2 %) were the most abundant group in T. zygioides essential oil, with thymol (51.2 %) as the principal component. The essential oil from the endemic species T. perinicus contained almost equal amounts of monoterpenoids (37.8 %) and aromatic compounds (36.0 %) with borneol (17.9 %) and thymol (20.9 %) as the major components. The obtained results revealed the existence of new chemotypes of T. atticus (caryophyllene oxide/β-caryophyllene), T. leucotrichus (β-caryophyllene/elemol/germacrene D) and T. striatus (β-caryophyllene/germacrene D/caryophyllene oxide). The essential oil content of endemic T. perinicus is reported for the first time. Principal component analysis (PCA) and cluster analysis (CA) were used to investigate the variations in the essential oils of different Thymus species from Sect. Hyphodromi (A. Kerner) Halácsy.     

青藏高原长花马先蒿的细胞地理学研究

对青藏高原长花马先蒿9个居群的核型及细胞地理学进行研究,所有居群的染色体数目均为2n=16,染色体基数为x=8。现有的细胞学资料表明:分布于云南中甸的居群可能为较原始的类群,而分布于西藏日土和青海门源的居群较为进化。长花马先蒿的9个居群均为二倍体,并未出现多倍化现象,可能是由于在末次冰期青藏高原存在广泛的避难所,二倍体得到很好的保存,冰期对它们的影响不是很大; 也可能是对环境的选择压力造成的。     

Karyotype analysis and new chromosome number reports in <Emphasis Type="Italic">Achillea</Emphasis> species

Cytological analyses were performed in nineteen accessions belonging to seven Achillea species. The results indicate that two species, A. filipendulina and A. tenuifolia, are diploid (2n = 2x = 18) and three species, A. bieberstinii, A. pachycephala, and A. aucheri are tetraploid (2n = 4x = 36). For two latter species, the chromosome numbers are new. Two ploidy levels of 2n = 6x = 54 and 2n = 8x = 72 in A. millefolium and two ploidy levels of 2n = 2x = 18 and 2n = 4x = 36 in A. santolina species were found. Cluster analysis based on chromosomal characteristics and karyotype asymmetry indices clustered the studied genotypes into three major groups. The first group included the diploid species (A. filipendulina, A. santolina and A. tenufolia), while the second group comprised tetraploid species (A. santolina, A. aucheri and A. pachycephala). A. millefolium (hexaploid, octaploid) and A. bieberstinii (tetraploid) were classified in the third group. Total form percentage (TF%) in groups 1, 2 and 3 were 42.03, 42.15 and 41.08, respectively. Group 1 possessed the highest average of symmetry index (S% = 70.8). Stebbins classification method grouped all accessions in class A. Moreover, group 1 (diploid level species), had the most symmetric karyotype from point of view of centromere and chromosome length. Therefore, it can be concluded that the genotypes belonging to group 1 are the earliest evolutionary form.     

Cytogeography of Solidago gigantea (Asteraceae) and its invasive ploidy level

Aim Newly formed polyploids experience problems of establishment and spread similar to those faced by newly introduced alien species. To understand the significance of polyploidy in biological invasions, we mapped the distribution of ploidy levels in Solidago gigantea Aiton in its native range in North America, and in Europe and East Asia where it is invasive. Location North America, Europe and East Asia. Methods Flow cytometry was used to measure ploidy levels in a total of 834 plants from 149 locations. Together with data from contributors and a literature review, ploidy‐level data were assembled for 336 locations. Cytogeographical maps from North America and Europe were prepared, incorporating new and previously published ploidy‐level data. Results In the native range, diploid, tetraploid and hexaploid plants were found, and also one triploid and one pentaploid plant (2n = 3x and 2n = 5x, respectively, each being new reports for this species). There was a high degree of geographical separation among the ploidy levels, and populations with mixed ploidy were rare. However, four zones were identified where plants of different ploidy could come into contact. In Europe and East Asia, only tetraploid plants were found. Main conclusions The geographical pattern in North America suggests that the ploidy levels are ecologically differentiated, although further investigations are needed to identify the nature of these differences. Alien populations appear to be exclusively tetraploid, but it is not clear whether this is because tetraploids were selectively introduced or because diploids were unsuccessful. In any case, comparisons between native and introduced populations need to account for ploidy level.     

Geographical Distribution of Diploid and Tetraploid Cytotypes of Thymus sect. Mastichina (Lamiaceae) in the Iberian Peninsula, Genome Size and Evolutionary Implications

The occurrence of diploid and tetraploid cytotypes in Thymus section Mastichina (Lamiaceae) was investigated by chromosome counting and flow cytometric measurements of DNA content. To cover the entire native distribution of the section across the Iberian Peninsula, plant material was sampled from 29 populations, representing all three taxa (Thymus albicans, T. mastichina subsp. mastichina and T. mastichina subsp. donyanae). We provide first estimates of genome size for taxa of this section. Analyses revealed the existence of new cytotypes in T. mastichina subsp. mastichina (2n?=?2x?=?28, 30) and confirmed the existence of previous ones (2n?=?4x?=?56, 58, 60). We also confirmed the presence of exclusively diploid cytotypes (2n?=?2x?=?30) in the southwestern Iberian endemics T. albicans and T. mastichina subsp. donyanae. We conclude that the southwestern Iberian Peninsula acted as a Pleistocene glacial refugium that may have permitted speciation processes within this section. Several hypotheses concerning these processes and the origin and distribution of the studied taxa are discussed.     

Chromosome numbers and taxonomic notes for Mexican coreopsis,sections electra and pseudoagarista (compositae: heliantheae)

Notes on morphology and chromosome numbers are given for several species of MexicanCoreopsis, most of which were poorly known prior to recent collections.Coreopsis parvifolia (sectionElectra) is a large, shrubby, octoploid (2n=112) species apparently restricted to a small area in Puebla. It andC. cuneifolia (a diploid with 2n=28) appear to be closely related but differ by a number of morphological features in addition to ploidy level. A first chromosome report forC. pringlei (sectionPseudoagarista) shows this rare species to be diploid (2n=26), a fact which is in agreement with the base number of the section, i.e.,x=13. Additional collections of the very rareC. rudis andC. mcvaughii show them to be similar yet distinct species. Chromosome determinations forC. petrophila from Nayarit and Durango agree with previous counts for Jalisco populations of the species, i.e., 2n=26. Considerable morphological variation exists within this species but no subspecific entities are recognized.     

Differentiating Thymus vulgaris chemotypes with ISSR molecular markers

Common thyme (Thymus vulgaris L.) is a Mediterranean evergreen shrub of the Lamiaceae that shows chemical polymorphism in its natural populations. In Mediterranean habitats six basic chemotypes have been described according to the main component in its essential oil: thymol, carvacrol, linalool, geraniol, α-terpineol, thuyanol-4, of which the thymol chemotype is widely used in cultivation and as a spice. In this study ten selected clones were analysed in terms of chemical composition of the volatile oil and genetic fingerprint. The study question was whether individual chemotypes can be distinguished by molecular markers. Chemical composition was analysed by GC-MS, and for assessing the genetic polymorphism ISSR marker system was chosen as it is reliable and easy to use even in case of less studied species. The studied T. vulgaris clones represented five different chemotypes and could be distinguished using 12 ISSR primers. Clones belonging to the thymol chemotype showed the most distinct separation. The study shows the potential of using molecular markers in breeding and selection of T. vulgaris, being able to differentiate different chemotypes.     

Pollen and stomata morphometrics and polyploidy in Eriotheca (Malvaceae‐Bombacoideae)

Approximately 70% of the angiosperm species are polyploid, an important phenomenon in the evolution of those plants. But ploidy estimates have often been hindered because of the small size and large number of chromosomes in many tropical groups. Since polyploidy affects cell size, morphometric analyses of pollen grains and stomata have been used to infer ploidy level. Polyploidy is present in many species of the Cerrado, the Neotropical savanna region in Central Brazil, and has been linked to apomixis in some taxa. Eriotheca gracilipes and Eriotheca pubescens are common tree species in this region, and present cytotypes that form reproductive mosaics. Hexaploid individuals (2n = 6x = 276) are polyembryonic and apomictic, while tetraploid and diploid individuals (2n = 2x = 92, 2n = 4x = 184) are sexual and monoembryonic. We tested whether morphometric analysis can be used to estimate ploidy levels in E. gracilipes and E. pubescens individuals. Pollen material from diploid and hexaploid individuals of E. gracilipes, and tetraploid and hexaploid individuals of E. pubescens, were fixed in 50% FAA, and expanded leaves were dried in silica gel. Pollen grains and stomata of at least five individuals from each population were measured. The results demonstrate that all measures were significantly different among cytotypes. Individuals with higher levels of ploidy (hexaploid) all presented measurements that were higher than those with lower levels (diploid and tetraploid). There was no overlap between ploidy levels in each species at 95% confidence interval. Thus, the size of the pollen grains and stomata are effective parameters for analysis of ploidy levels in E. gracilipes and E. pubescens.     

Pollinator-mediated assortative mating in mixed ploidy populations of Chamerion angustifolium (Onagraceae)

Establishment of polyploid individuals within diploid populations is theoretically unlikely unless polyploids are reproductively isolated, pre-zygotically, through assortative pollination. Here, we quantify the contribution of pollinator diversity and foraging behaviour to assortative pollen deposition in three mixed-ploidy populations of Chamerion angustifolium (Onagraceae). Diploids and tetraploids were not differentiated with respect to composition of insect visitors. However, foraging patterns of the three most common insect visitors (all bees) reinforced assortative pollination. Bees visited tetraploids disproportionately often and exhibited higher constancy on tetraploids in all three populations. In total, 73% of all bee flights were between flowers of the same ploidy (2x–2x, 4x–4x); 58% of all flights to diploids and 83% to tetraploids originated from diploid and tetraploid plants, respectively. Patterns of pollen deposition on stigmas mirrored pollinator foraging behaviour; 73% of all pollen on stigmas (70 and 75% of pollen on diploid and tetraploid stigmas, respectively) came from within-ploidy pollinations. These results indicate that pollinators contribute to high rates of pre-zygotic reproductive isolation. If patterns of fertilization track pollen deposition, pollinator–plant interactions may help explain the persistence and spread of tetraploids in mixed-ploidy populations.     

New chromosome counts and evidence of polyploidy in Haageocereus and related genera in tribe Trichocereeae and other tribes of Cactaceae

Chromosome numbers for a total of 54 individuals representing 13 genera and 40 species of Cactaceae, mostly in tribe Trichocereeae, are reported. Five additional taxa examined belong to subfamily Opuntioideae and other tribes of Cactoideae (Browningieae, Pachycereeae, Notocacteae, and Cereeae). Among Trichocereeae, counts for 35 taxa in eight genera are reported, with half of these (17 species) for the genus Haageocereus. These are the first chromosome numbers reported for 36 of the 40 taxa examined, as well as the first counts for the genus Haageocereus. Both diploid and polyploid counts were obtained. Twenty nine species were diploid with 2n=2x=22. Polyploid counts were obtained from the genera Espostoa, Cleistocactus, Haageocereus, and Weberbauerocereus; we detected one triploid (2n=3x=33), nine tetraploids (2n=4x=44), one hexaploid (2n=6x=66), and three octoploids (2n=8x=88). In two cases, different counts were recorded for different individuals of the same species (Espostoa lanata, with 2n=22, 44, and 66; and Weberbauerocereus rauhii, with 2n=44 and 88). These are the first reported polyploid counts for Haageocereus, Cleistocactus, and Espostoa. Our counts support the hypothesis that polyploidy and hybridization have played prominent roles in the evolution of Haageocereus, Weberbauerocereus, and other Trichocereeae.     

Towards resolving the Knautia arvensis agg. (Dipsacaceae) puzzle: primary and secondary contact zones and ploidy segregation at landscape and microgeographic scales

Background and Aims

Detailed knowledge of variations in ploidy levels and their geographic distributions is one of the key tasks faced in polyploid research in natural systems. Flow cytometry has greatly facilitated the field of cytogeography by allowing characterization of ploidy levels at both the regional and population scale, and at multiple stages of the life cycle. In the present study, flow cytometry was employed to investigate the patterns and dynamics of ploidy variation in the taxonomically challenging complex Knautia arvensis (Dipsacaceae) and some of its allies (K. dipsacifolia, K. slovaca) in Central Europe.

Methods

DNA ploidy levels were estimated by DAPI flow cytometry in 5205 adult plants, 228 seedlings and 400 seeds collected from 292 Knautia populations in seven European countries. The flow cytometric data were supplemented with conventional chromosome counts. A subset of 79 accessions was subjected to estimation of the absolute genome size using propidium iodide flow cytometry.

Key Results and Conclusions

Five different ploidy levels (from 2x to 6x) were found, with triploids of K. arvensis being recorded for the first time. The species also exhibited variation in the monoploid genome size, corresponding to the types of habitats occupied (grassland diploid populations had larger genome sizes than relict and subalpine diploid populations). Disregarding relict populations, the distribution of 2x and 4x cytotypes was largely parapatric, with a diffuse secondary contact zone running along the north-west margin of the Pannonian basin. Spatial segregation of the cytotypes was also observed on regional and microgeographic scales. The newly detected sympatric growth of diploids and tetraploids in isolated relict habitats most likely represents the primary zone of cytotype contact. Ploidy level was found to be a major determinant of the strength of inter-cytotype reproductive barriers. While mixed 2x + 4x populations virtually lacked the intermediate ploidy level at any ontogenetic stage, pentaploid hybrids were common in 4x +6x populations, despite the cytotypes representing different taxonomic entities.Key words: Contact zone, cytogeography, flow cytometry, genome size, hybridization, Knautia arvensis, ploidy mixture, polyploidy, relict, reproductive isolation, serpentine