Using a model-based framework for analysing genetic diversity during germination and heterotrophic growth of Medicago truncatula

Background and Aims

The framework provided by an emergence model was used: (1) for phenotyping germination and heterotrophic growth of Medicago truncatula in relation to two major environmental factors, temperature and water potential; and (2) to evaluate the extent of genetic differences in emergence-model parameters.

Methods

Eight cultivars and natural accessions of M. trunculata were studied. Germination was recorded from 5 to 30 °C and from 0 to −0·75 MPa, and seedling growth from 10 to 20 °C.

Key Results

Thermal time to reach 50 % germination was very short (15 °Cd) and almost stable between genotypes, while base temperature (2–3 °C) and base water potential for germination (−0·7 to −1·3 MPa) varied between genotypes. Only 35 °Cd after germination were required to reach 30 mm hypocotyl length with significant differences among genotypes. Base temperature for elongation varied from 5·5 to 7·5 °C. Low temperatures induced a general shortening of the seedling, with some genotypes more responsive than others. No relationship with initial seed mass or seed reserve distribution was observed, which might have explained differences between genotypes and the effects of low temperatures.

Conclusions

The study provides a set of reference values for M. trunculata users. The use of the ecophysiological model allows comparison of these values between such non-crop species and other crops. It has enabled phenotypic variability in response to environmental conditions related to the emergence process to be identified. The model will allow simulation of emergence differences between genotypes in a range of environments using these parameter values. Genomic tools available for the model species M. trunculata will make it possible to analyse the genetic and molecular determinants of these differences.Key words: Core collection, emergence, Medicago truncatula, modelling, seed, temperature, water potential     

The potential of plant viruses to promote genotypic diversity via genotype x environment interactions

Background and Aims

Genotype by environment (G × E) interactions are important for the long-term persistence of plant species in heterogeneous environments. It has often been suggested that disease is a key factor for the maintenance of genotypic diversity in plant populations. However, empirical evidence for this contention is scarce. Here virus infection is proposed as a possible candidate for maintaining genotypic diversity in their host plants.

Methods

The effects of White clover mosaic virus (WClMV) on the performance and development of different Trifolium repens genotypes were analysed and the G × E interactions were examined with respect to genotype-specific plant responses to WClMV infection. Thus, the environment is defined as the presence or absence of the virus.

Key Results

WClMV had a negative effect on plant performance as shown by a decrease in biomass and number of ramets. These effects of virus infection differ greatly among host genotypes, representing a strong G × E interaction. Moreover, the relative fitness and associated ranking of genotypes changed significantly between control and virus treatments. This shift in relative fitness among genotypes suggests the potential for WClMV to provoke differential selection on T. repens genotypes, which may lead to negative frequency-dependent selection in host populations.

Conclusions

The apparent G × E interaction and evident repercussions for relative fitness reported in this study stress the importance of viruses for ecological and evolutionary processes and suggest an important role for viruses in shaping population dynamics and micro-evolutionary processes.     

Biased morph ratios and skewed mating success contribute to loss of genetic diversity in the distylous Pulmonaria officinalis

Background and Aims

In heterostylous plant species, skewed morph ratios are not uncommon and may arise from a range of factors. Despite the recognized importance of skewed morph ratios on overall reproductive success within populations, little is known about the impact of skewed morph ratios on population genetic diversity and differentiation in heterostylous species. This study specifically aimed to clarify the effect of population size and morph bias on population genetic diversity and differentiation in the temperate forest herb Pulmonaria officinalis. This species is characterized by a distylous breeding system and shows morph-specific differences in reproductive success.

Methods

Genetic diversity was determined for 27 P. officinalis populations in northern Belgium by using eight recently developed microsatellite markers. Multiple regressions were used to assess the relationship between genetic diversity, morph bias and population size, and F_ST-values were calculated for short- and long-styled morphs separately to study genetic differentiation as a function of morph type.

Key Results

For all genetic measures used, morph bias was more important in explaining patterns of genetic diversity than population size, and in all cases patterns of population genetic diversity followed a quadratic function, which showed a symmetrical decrease in genetic diversity with increasing morph bias. However, probably due to the reproductive advantage of L-morphs relative to S-morphs, maximum genetic diversity was found in populations showing an excess of L-morphs (60·7 % L-morph). On the other hand, no significant difference in pairwise genetic distances between populations was observed between L- (0·107) and S-morphs (0·106).

Conclusions

Our results indicate that significant deviations from equal morph ratios not only affect plant reproductive success but also population genetic diversity of heterostylous plant species. Hence, when defining conservation measures for populations of heterostylous plant species, morph ratios should be considered as an important trait affecting their long-term population viability.     

High clonal diversity in threatened peripheral populations of the yellow bird's nest (Hypopitys monotropa; syn. Monotropa hypopitys)

Background and Aims

Peripheral populations of plant species are often characterized by low levels of genetic diversity as a result of genetic drift, restricted gene flow, inbreeding and asexual reproduction. These effects can be exacerbated where range-edge populations are fragmented. The main aim of the present study was to assess the levels of genetic diversity in remnant populations of Hypopitys monotropa (syn. Monotropa hypopitys; yellow bird''s nest) at the edge of the species'' European range in Northern Ireland, since these remnant populations are small and highly fragmented.

Methods

Every plant found through surveys of 21 extant populations was genotyped for eight microsatellite loci to estimate levels and patterns of genetic diversity and clonality.

Key Results

Levels of genetic diversity were relatively high in the populations studied, and the incidence of clonal reproduction was generally low, with a mean of only 14·45 % of clonal individuals. Clones were small and highly spatially structured. Levels of inbreeding, however, were high.

Conclusions

The observed low levels of clonality suggest that the majority of genets in the populations of H. monotropa studied are fertile and that reproduction is predominantly sexual. As the species is highly self-compatible, it is likely that the high levels of inbreeding observed in the populations in the present study are the result of self-pollination, particularly given the small numbers of individuals in most of the patches. Given this extent of inbreeding, further genetic monitoring would be advisable to ensure that genetic diversity is maintained.     

Centennial olive trees as a reservoir of genetic diversity

Background and Aims

Genetic characterization and phylogenetic analysis of the oldest trees could be a powerful tool both for germplasm collection and for understanding the earliest origins of clonally propagated fruit crops. The olive tree (Olea europaea L.) is a suitable model to study the origin of cultivars due to its long lifespan, resulting in the existence of both centennial and millennial trees across the Mediterranean Basin.

Methods

The genetic identity and diversity as well as the phylogenetic relationships among the oldest wild and cultivated olives of southern Spain were evaluated by analysing simple sequence repeat markers. Samples from both the canopy and the roots of each tree were analysed to distinguish which trees were self-rooted and which were grafted. The ancient olives were also put into chronological order to infer the antiquity of traditional olive cultivars.

Key Results

Only 9·6 % out of 104 a priori cultivated ancient genotypes matched current olive cultivars. The percentage of unidentified genotypes was higher among the oldest olives, which could be because they belong to ancient unknown cultivars or because of possible intra-cultivar variability. Comparing the observed patterns of genetic variation made it possible to distinguish which trees were grafted onto putative wild olives.

Conclusions

This study of ancient olives has been fruitful both for germplasm collection and for enlarging our knowledge about olive domestication. The findings suggest that grafting pre-existing wild olives with olive cultivars was linked to the beginnings of olive growing. Additionally, the low number of genotypes identified in current cultivars points out that the ancient olives from southern Spain constitute a priceless reservoir of genetic diversity.     

Genetic diversity for grain nutrients in wild emmer wheat: potential for wheat improvement

Background and Aims

Micronutrient malnutrition, particularly zinc and iron deficiency, afflicts over three billion people worldwide due to low dietary intake. In the current study, wild emmer wheat (Triticum turgidum ssp. dicoccoides), the progenitor of domesticated wheat, was tested for (1) genetic diversity in grain nutrient concentrations, (2) associations among grain nutrients and their relationships with plant productivity, and (3) the association of grain nutrients with the eco-geographical origin of wild emmer accessions.

Methods

A total of 154 genotypes, including wild emmer accessions from across the Near Eastern Fertile Crescent and diverse wheat cultivars, were characterized in this 2-year field study for grain protein, micronutrient (zinc, iron, copper and manganese) and macronutrient (calcium, magnesium, potassium, phosphorus and sulphur) concentrations.

Key Results

Wide genetic diversity was found among the wild emmer accessions for all grain nutrients. The concentrations of grain zinc, iron and protein in wild accessions were about two-fold greater than in the domesticated genotypes. Concentrations of these compounds were positively correlated with one another, with no clear association with plant productivity, suggesting that all three nutrients can be improved concurrently with no yield penalty. A subset of 12 populations revealed significant genetic variation between and within populations for all minerals. Association between soil characteristics at the site of collection and grain nutrient concentrations showed negative associations between soil clay content and grain protein and between soil-extractable zinc and grain zinc, the latter suggesting that the greatest potential for grain nutrient minerals lies in populations from micronutrient-deficient soils.

Conclusions

Wild emmer wheat germplasm offers unique opportunities to exploit favourable alleles for grain nutrient properties that were excluded from the domesticated wheat gene pool.     

Tetraploidization events by chromosome doubling of nucellar cells are frequent in apomictic citrus and are dependent on genotype and environment

Background and Aims

Polyploidy is a major component of plant evolution. The citrus gene pool is essentially diploid but tetraploid plants are frequently encountered in seedlings of diploid apomictic genotypes. The main objectives of the present study were to establish the origin of these tetraploid plants and to ascertain the importance of genotypic and environmental factors on tetraploid formation.

Methods

Tetraploid seedlings from 30 diploid apomictic genotypes were selected by flow cytometry and genotyped with 24 single sequence repeat (SSR) markers to analyse their genetic origin. Embryo rescue was used to grow all embryos contained in polyembryonic seeds of ‘Tardivo di Ciaculli’ mandarin, followed by characterization of the plantlets obtained by flow cytometry and SSR markers to accurately establish the rate of tetraploidization events and their potential tissue location. Inter-annual variations in tetraploid seedling rates were analysed for seven genotypes. Variation in tetraploid plantlet rates was analysed between different seedlings of the same genotype (‘Carrizo’ citrange; Citrus sinensis × Poncirus trifoliata) from seeds collected in different tropical, subtropical and Mediterranean countries.

Key Results

Tetraploid plants were obtained for all the studied diploid genotypes, except for four mandarins. All tetraploid plants were identical to their diploid maternal line for SSR markers and were not cytochimeric. Significant genotypic and environmental effects were observed, as well as negative correlation between mean temperature during the flowering period and tetraploidy seedling rates. The higher frequencies (20 %) of tetraploids were observed for citranges cultivated in the Mediterranean area.

Conclusions

Tetraploidization by chromosome doubling of nucellar cells are frequent events in apomictic citrus, and are affected by both genotypic and environmental factors. Colder conditions in marginal climatic areas appear to favour the expression of tetraploidization. Tetraploid genotypes arising from chromosome doubling of apomictic citrus are extensively being used as parents in breeding programmes to develop seedless triploid cultivars and have potential direct use as new rootstocks.     

Evolution under domestication: ongoing artificial selection and divergence of wild and managed Stenocereus pruinosus (Cactaceae) populations in the Tehuac��n Valley, Mexico

Background and Aims

The Tehuacán Valley in Mexico is a principal area of plant domestication in Mesoamerica. There, artificial selection is currently practised on nearly 120 native plant species with coexisting wild, silvicultural and cultivated populations, providing an excellent setting for studying ongoing mechanisms of evolution under domestication. One of these species is the columnar cactus Stenocereus pruinosus, in which we studied how artificial selection is operating through traditional management and whether it has determined morphological and genetic divergence between wild and managed populations.

Methods

Semi-structured interviews were conducted with 83 households of three villages to investigate motives and mechanisms of artificial selection. Management effects were studied by comparing variation patterns of 14 morphological characters and population genetics (four microsatellite loci) of 264 plants from nine wild, silvicultural and cultivated populations.

Key Results

Variation in fruit characters was recognized by most people, and was the principal target of artificial selection directed to favour larger and sweeter fruits with thinner or thicker peel, fewer spines and pulp colours others than red. Artificial selection operates in agroforestry systems favouring abundance (through not felling plants and planting branches) of the preferred phenotypes, and acts more intensely in household gardens. Significant morphological divergence between wild and managed populations was observed in fruit characters and plant vigour. On average, genetic diversity in silvicultural populations (H_E = 0·743) was higher than in wild (H_E = 0·726) and cultivated (H_E = 0·700) populations. Most of the genetic variation (90·58 %) occurred within populations. High gene flow (Nm_FST > 2) was identified among almost all populations studied, but was slightly limited by mountains among wild populations, and by artificial selection among wild and managed populations.

Conclusions

Traditional management of S. pruinosus involves artificial selection, which, despite the high levels of gene flow, has promoted morphological divergence and moderate genetic structure between wild and managed populations, while conserving genetic diversity.     

Genetic evidence for predominantly hydrochoric gene flow in the invasive riparian plant Impatiens glandulifera (Himalayan balsam)

Background and Aims

Riparian systems are prone to invasion by alien plant species. The spread of invasive riparian plants may be facilitated by hydrochory, the transport of seeds by water, but while ecological studies have highlighted the possible role of upstream source populations in the establishment and persistence of stands of invasive riparian plant species, population genetic studies have as yet not fully addressed the potential role of hydrochoric dispersal in such systems.

Methods

A population genetics approach based on a replicated bifurcate sampling design is used to test hypotheses consistent with patterns of unidirectional, linear gene flow expected under hydrochoric dispersal of the invasive riparian plant Impatiens glandulifera in two contrasting river systems.

Key results

A significant increase in levels of genetic diversity downstream was observed, consistent with the accumulation of propagules from upstream source populations, and strong evidence was found for organization of this diversity between different tributaries, reflecting the dendritic organization of the river systems studied.

Conclusions

These findings indicate that hydrochory, rather than anthropogenic dispersal, is primarily responsible for the spread of I. glandulifera in these river systems, and this is relevant to potential approaches to the control of invasive riparian plant species.     

Investigating the effects of topography and clonality on genetic structuring within a large Norwegian population of Arabidopsis lyrata

Background and Aims

The gene flow through pollen or seeds governs the extent of spatial genetic structure in plant populations. Another factor that can contribute to this pattern is clonal growth. The perennial species Arabidopsis lyrata ssp. petraea (Brassicaceae) is a self-incompatible, clonal species found in disjunctive populations in central and northern Europe.

Methods

Fourteen microsatellite markers were employed to study the level of kinship and clonality in a high-altitude mountain valley at Spiterstulen, Norway. The population has a continuous distribution along the banks of the River Visa for about 1·5 km. A total of 17 (10 m × 10 m) squares were laid out in a north–south transect following the river on both sides.

Key Results

It is shown that clonal growth is far more common than previously shown in this species, although the overall size of the genets is small (mean diameter = 6·4 cm). Across the whole population there is no indication of isolation by distance, and spatial genetic structure is only visible on fine spatial scales. In addition, no effect of the river on the spatial distribution of genotypes was found.

Conclusions

Unexpectedly, the data show that populations of small perennials like A. lyrata can behave like panmictic units across relatively large areas at local sites, as opposed to earlier findings in central Europe.     

Polyembryony in non-apomictic citrus genotypes

Background and Aims

Adventitious embryony from nucellar cells is the mechanism leading to apomixis in Citrus sp. However, singular cases of polyembryony have been reported in non-apomictic genotypes as a consequence of 2x × 4x hybridizations and in vitro culture of isolated nucelli. The origin of the plants arising from the aforementioned processes remains unclear.

Methods

The genetic structure (ploidy and allelic constitution with microsatellite markers) of plants obtained from polyembryonic seeds arising from 2x × 4x sexual hybridizations and those regenerated from nucellus culture in vitro was systematically analysed in different non-apomictic citrus genotypes. Histological studies were also conducted to try to identify the initiation process underlying polyembryony.

Key Results

All plants obtained from the same undeveloped seed in 2x × 4x hybridizations resulted from cleavage of the original zygotic embryo. Also, the plants obtained from in vitro nucellus culture were recovered by somatic embryogenesis from cells that shared the same genotype as the zygotic embryos of the same seed.

Conclusions

It appears that in non-apomictic citrus genotypes, proembryos or embryogenic cells are formed by cleavage of the zygotic embryos and that the development of these adventitious embryos, normally hampered, can take place in vivo or in vitro as a result of two different mechanisms that prevent the dominance of the initial zygotic embryo.     

ITIS,a bioinformatics tool for accurate identification of transposon insertion sites using next-generation sequencing data

Background

Transposable elements constitute an important part of the genome and are essential in adaptive mechanisms. Transposition events associated with phenotypic changes occur naturally or are induced in insertional mutant populations. Transposon mutagenesis results in multiple random insertions and recovery of most/all the insertions is critical for forward genetics study. Using genome next-generation sequencing data and appropriate bioinformatics tool, it is plausible to accurately identify transposon insertion sites, which could provide candidate causal mutations for desired phenotypes for further functional validation.

Results

We developed a novel bioinformatics tool, ITIS (Identification of Transposon Insertion Sites), for localizing transposon insertion sites within a genome. It takes next-generation genome re-sequencing data (NGS data), transposon sequence, and reference genome sequence as input, and generates a list of highly reliable candidate insertion sites as well as zygosity information of each insertion. Using a simulated dataset and a case study based on an insertional mutant line from Medicago truncatula, we showed that ITIS performed better in terms of sensitivity and specificity than other similar algorithms such as RelocaTE, RetroSeq, TEMP and TIF. With the case study data, we demonstrated the efficiency of ITIS by validating the presence and zygosity of predicted insertion sites of the Tnt1 transposon within a complex plant system, M. truncatula.

Conclusion

This study showed that ITIS is a robust and powerful tool for forward genetic studies in identifying transposable element insertions causing phenotypes. ITIS is suitable in various systems such as cell culture, bacteria, yeast, insect, mammal and plant.

Electronic supplementary material

The online version of this article (doi:10.1186/s12859-015-0507-2) contains supplementary material, which is available to authorized users.     

Somatic mutations contribute to genotypic diversity in sterile and fertile populations of the threatened shrub, Grevillea rhizomatosa (Proteaceae)

Background and Aims

Grevillea rhizomatosa is a spreading shrub which exhibits multiple breeding strategies within a narrow area in the fire-prone heathlands of eastern Australia. Reproductive strategies include self-compatibility, self-incompatibility and clonality (with and without sterility). The close proximity of contrasting breeding systems provides an opportunity to explore the evolution of sterility and to compare and contrast the origins of genotypic diversity (recombinant or somatic) against degrees of sexual expression.

Methods

ISSR markers for 120 band positions (putative loci) were used to compare genetic diversity among five populations at a macro-scale of 5 m between samples (n = 244 shrubs), and at a micro-scale of nearest neighbours for all plants in five 25-m² quadrats with contrasting fertilities (n = 162 shrubs). Nearest-neighbour sampling included several clusters of connected ramets. Matrix incompatibility (MIC) analyses were used to evaluate the relative contribution of recombination and somatic mutation to genotype diversity.

Key Results

High levels of genotypic diversity were found in all populations regardless of fertilities (fertile populations, G/N ≥ 0·94; sterile populations, G/N ≥ 0·97) and most sterile populations had a unique genetic profile. Somatic mutations were detected along connected ramets in ten out of 42 ramet clusters. MIC analyses showed that somatic mutations have contributed to diversity in all populations and particularly so in sterile populations.

Conclusions

Somatic mutations contribute significantly to gene diversity in sterile populations of Grevillea rhizomatosa, the accumulation of which is the likely cause of male and female sterility. High levels of genetic diversity therefore may not always be synonymous with sexual fitness and genetic health. We hypothesize that frequent fires drive selection for clonal reproduction, at the cost of flowering such that sexual functions are not maintained through selection, and the build-up of somatic mutations in meristems results in high genotype diversity at the cost of pollen and ovule fertilities.     

Microsatellites reveal substantial among-population genetic differentiation and strong inbreeding in the relict fern Dryopteris aemula

Background and Aims

A previous study detected no allozyme diversity in Iberian populations of the buckler-fern Dryopteris aemula. The use of a more sensitive marker, such as microsatellites, was thus needed to reveal the genetic diversity, breeding system and spatial genetic structure of this species in natural populations.

Methods

Eight microsatellite loci for D. aemula were developed and their cross-amplification with other ferns was tested. Five polymorphic loci were used to characterize the amount and distribution of genetic diversity of D. aemula in three populations from the Iberian Peninsula and one population from the Azores.

Key Results

Most microsatellite markers developed were transferable to taxa close to D. aemula. Overall genetic variation was low (H_T = 0·447), but was higher in the Azorean population than in the Iberian populations of this species. Among-population genetic differentiation was high (F_ST = 0·520). All loci strongly departed from Hardy–Weinberg equilibrium. In the population where genetic structure was studied, no spatial autocorrelation was found in any distance class.

Conclusions

The higher genetic diversity observed in the Azorean population studied suggested a possible refugium in this region from which mainland Europe has been recolonized after the Pleistocene glaciations. High among-population genetic differentiation indicated restricted gene flow (i.e. lack of spore exchange) across the highly fragmented area occupied by D. aemula. The deviations from Hardy–Weinberg equilibrium reflected strong inbreeding in D. aemula, a trait rarely observed in homosporous ferns. The absence of spatial genetic structure indicated effective spore dispersal over short distances. Additionally, the cross-amplification of some D. aemula microsatellites makes them suitable for use in other Dryopteris taxa.     

The influence of cone age on the relative longevity of Banksia seeds

Background and Aims

Serotiny is common in the genus Banksia, so any seed collection is likely to be comprised of seeds that were produced in many different years. This study aimed to determine the impact of cone age and degree of serotiny on longevity in ex situ storage.

Methods

Cones of identifiable age classes were collected from three species of Banksia. Seeds were extracted from cones and the degree of serotiny calculated. An estimate of initial viability (K_i), the time for viability to fall by one probit (σ) and the relative longevity of seeds (p₅₀) for each species and cone age class was determined using a comparative longevity test (50 °C, 63 % relative humidity).

Key Results

The degree of serotiny ranged from moderate (7·9) for Banksia attenuata to strong (40·4) for B. hookeriana. Survival curves for all seed age classes within each species could be described by regressions with a common slope (1/σ), but with different values for K_i. The time taken for viability to fall by one probit (σ) could be described by a common value (29·1 d) for all three species.

Conclusions

Differences in seed longevity between cone age classes and species was related to variation in initial viability (K_i) rather than to differences in σ. While targeting the youngest mature seed cohort on a plant will maximize the viability of seeds collected, a wide range of age classes should be collected (but stored as separate cohorts if possible) for quality conservation/restoration seed collections where genetic diversity is important.     

Range expansion of a selfing polyploid plant despite widespread genetic uniformity

Background and Aims

Ongoing and previous range expansions have a strong influence on population genetic structure of plants. In turn, genetic variation in the new range may affect the population dynamics and the expansion process. The annual Ceratocapnos claviculata (Papaveraceae) has expanded its Atlantic European range in recent decades towards the north and east. Patterns of genetic diversity were investigated across the native range to assess current population structure and phylogeographical patterns. A test was then made as to whether genetic diversity is reduced in the neophytic range and an attempt was made to identify source regions of the expansion.

Methods

Samples were taken from 55 populations in the native and 34 populations in the neophytic range (Sweden, north-east Germany). Using amplified fragment length polymorphism markers an analysis was made of genetic variation and population structure (Bayesian statistical modelling) and population differentiation was quantified. Pollen/ovule ratio was analysed as a proxy for the breeding system.

Key Results

Genetic diversity at population level was very low (mean H_e = 0·004) and two multilocus genotypes dominated large parts of the new range. Population differentiation was strong (F_ST = 0·812). These results and a low pollen/ovule ratio are consistent with an autogamous breeding system. Genetic variation decreased from the native to the neophytic range. Within the native range, H_e decreased towards the north-east, whereas population size increased. According to the Bayesian cluster analysis, the putative source regions of the neophytic range are situated in north-west Germany and adjacent regions.

Conclusions

Ceratocapnos claviculata shows a cline of genetic variation due to postglacial recolonization from putative Pleistocene refugia in south-west Europe. Nevertheless, the species has expanded successfully during the past 40 years to southern Sweden and north-east Germany where it occurs as an opportunistic neophyte. Recent expansion was mainly human-mediated by single long-distance diaspore transport and was facilitated by habitat modification.     

Dynamics of polyploid formation and establishment in the allotetraploid rock fern Asplenium majoricum

Background and Aims

Successful establishment of newly formed polyploid species depends on several interlinked genetic and ecological factors. These include genetic diversity within and among individuals, chromosome behaviour and fertility, novel phenotypes resulting from novel genomic make-up and expression, intercytotypic and interspecific competition, and adaptation to distinct habitats. The allotetraploid rock fern Asplenium majoricum is known from one small population in Valencia, Spain, and several larger populations on the Balearic island of Majorca. In Valencia, it occurs sympatrically with its diploid parents, A. fontanum subsp. fontanum and A. petrarchae subsp. bivalens, and their diploid hybrid A. × protomajoricum. This highly unusual situation allowed the study of polyploid genetic diversity and its relationship to the formation and establishment of nascent polyploid lineages.

Methods

Genetic variation for isozyme and chloroplast DNA markers was determined for A. majoricum and A. × protomajoricum sampled thoroughly from known sites in Majorca and Valencia. Results were compared with variation determined previously for the diploid parent taxa.

Key Results

A highly dynamic system with recurring diploid hybrid and allotetraploid formation was discovered. High diversity in the small Valencian A. majoricum population indicates multiple de novo origins from diverse parental genotypes, but most of these lineages become extinct without becoming established. The populations on Majorca most probably represent colonization(s) from Valencia rather than an in situ origin. Low genetic diversity suggests that this colonization may have occurred only once.

Conclusions

There is a striking contrast in success of establishment of the Majorcan and Valencian populations of A. majoricum. Chance founding of populations in a habitat where neither A. fontanum subsp. fontanum nor A. petrarchae subsp. bivalens occurs appears to have been a key factor enabling the establishment of A. majoricum on Majorca. Successful establishment of this polyploid is probably dependent on geographic isolation from diploid progenitor competition.     

Mixed mating in androdioecious Mercurialis annua inferred using progeny arrays and diploid-acting microsatellite loci in a hexaploid background

Background and Aims

The frequency at which males can be maintained with hermaphrodites in androdioecious populations is predicted to depend on the selfing rate, because self-fertilization by hermaphrodites reduces prospective siring opportunities for males. In particular, high selfing rates by hermaphrodites are expected to exclude males from a population. Here, the first estimates are provided of the mating system from two wild hexaploid populations of the androdioecious European wind-pollinated plant M. annua with contrasting male frequencies.

Methods

Four diploid microsatellite loci were used to genotype 19–20 progeny arrays from two populations of M. annua, one with males and one without. Mating-system parameters were estimated using the program MLTR.

Key Results

Both populations had similar, intermediate outcrossing rates (t_m = 0·64 and 0·52 for the population with and without males, respectively). The population without males showed a lower level of correlated paternity and biparental inbreeding and higher allelic richness and gene diversity than the population with males.

Conclusions

The results demonstrate the utility of new diploid microsatellite loci for mating system analysis in a hexaploid plant. It would appear that androdioecious M. annua has a mixed-mating system in the wild, an uncommon finding for wind-pollinated species. This study sets a foundation for future research to assess the relative importance of the sexual system, plant-density variation and stochastic processes for the regulation of male frequencies in M. annua over space and time.     

Interspecific gene flow in a multispecies oak hybrid zone in the Sierra Tarahumara of Mexico

Background and Aims

Interspecific gene flow can occur in many combinations among species within the genus Quercus, but simultaneous hybridization among more than two species has been rarely analysed. The present study addresses the genetic structure and morphological variation in a triple hybrid zone formed by Q. hypoleucoides, Q. scytophylla and Q. sideroxyla in north-western Mexico.

Methods

A total of 247 trees from ten reference and 13 presumed intermediate populations were characterized using leaf shape variation and geometric morphometrics, and seven nuclear microsatellites as genetic markers. Discriminant function analysis was performed for leaf shape variation, and estimates of genetic diversity and structure, and individual Bayesian genetic assignments were obtained.

Key Results

Reference populations formed three completely distinct groups according to discriminant function analysis based on the morphological data, and showed low, but significant, genetic differentiation. Populations from the zone of contact contained individuals morphologically intermediate between pairs of species in different combinations, or even among the three species. The Bayesian admixture analysis found that three main genetic clusters best fitted the data, with good correspondence of reference populations of each species to one of the genetic clusters, but various degrees of admixture evidenced in populations from the contact area.

Conclusions

The three oak species have formed a complex hybrid zone that is geographically structured as a mosaic, and comprising a wide range of genotypes, including hybrids between different species pairs, backcrosses and probable triple hybrids.     

Diverse spore rains and limited local exchange shape fern genetic diversity in a recently created habitat colonized by long-distance dispersal

Background and Aims

Populations established by long-distance colonization are expected to show low levels of genetic variation per population, but strong genetic differentiation among populations. Whether isolated populations indeed show this genetic signature of isolation depends on the amount and diversity of diaspores arriving by long-distance dispersal, and time since colonization. For ferns, however, reliable estimates of long-distance dispersal rates remain largely unknown, and previous studies on fern population genetics often sampled older or non-isolated populations. Young populations in recent, disjunct habitats form a useful study system to improve our understanding of the genetic impact of long-distance dispersal.

Methods

Microsatellite markers were used to analyse the amount and distribution of genetic diversity in young populations of four widespread calcicole ferns (Asplenium scolopendrium, diploid; Asplenium trichomanes subsp. quadrivalens, tetraploid; Polystichum setiferum, diploid; and Polystichum aculeatum, tetraploid), which are rare in The Netherlands but established multiple populations in a forest (the Kuinderbos) on recently reclaimed Dutch polder land following long-distance dispersal. Reference samples from populations throughout Europe were used to assess how much of the existing variation was already present in the Kuinderbos.

Key Results

A large part of the Dutch and European genetic diversity in all four species was already found in the Kuinderbos. This diversity was strongly partitioned among populations. Most populations showed low genetic variation and high inbreeding coefficients, and were assigned to single, unique gene pools in cluster analyses. Evidence for interpopulational gene flow was low, except for the most abundant species.

Conclusions

The results show that all four species, diploids as well as polyploids, were capable of frequent long-distance colonization via single-spore establishment. This indicates that even isolated habitats receive dense and diverse spore rains, including genotypes capable of self-fertilization. Limited gene flow may conserve the genetic signature of multiple long-distance colonization events for several decades.