Polymorphic microsatellite loci in Plantago lanceolata

The genus Plantago is particularly interesting for evolutionary studies because of its wide range of mating systems. We have developed primers for five highly polymorphic microsatellite loci isolated from P. lanceolata. All five loci amplified and were polymorphic in the two populations examined, Lowsteads Beach in the United Kingdom and Duke in the United States. These new markers will allow a comparison of population structure between the outcrossing species P. lanceolata, and the highly selfing species P. major.     

Population and species variation of minisatellite DNA in Plantago

Three Plantago species were surveyed for within- and between-population variation at DNA sequences detected with the M13 minisatellite probe. The levels and patterns of variation detected by this probe correspond to those expected from the mating systems of the species. The highly-selfing species P. major has a relatively low variability of minisatellite sequences within populations and considerable differentiation between populations. The outcrossing species P. lanceolata exhibits higher minisatellite variability within populations and moderate differentiation between populations. P. coronopus, with a mixed mating system, has levels of variation intermediate between P. major and P. lanceolata. The levels of variation within and between populations corresponds, in general, to the levels of allozyme variation determined in an earlier study. Mating system and population structure appear to have a major influence on M13-detected fragment variation.     

Genetic variability in Plantago species in relation to their ecology

Summary Populations of the three Plantago species P. major, P. lanceolata and P. coronopus were scored for allozyme variability. They showed normal variability levels compared with other, similar plant species. Differentiation among populations appeared to be rather low in comparison with other species, probably due to a considerable amount of long distance seed transport. In order to be able to make an analysis of small-scale gene flow, all (sub)populations were critically checked for the existence of population structure equilibrium. The allozyme variation was tested for neutrality by testing homogeneity of F values among loci: between populations (Lewontin-Krakauer test) and within populations. No systematic deviations from the prediction of the neutral theory could be established. From the population structure analyses, little gene flow in the species P. major (with high selfing levels) and P. coronopus could be concluded, whereas P. lanceolata showed relatively high levels of gene flow. The degree of homozygosity in the latter species was too high to explain with the available data. In P. coronopus, on the other hand, an unusually high number of heterozygotes were observed.Grassland Species Research Group Publication No. 73     

Genetic analysis of morphological variability in three Plantago species with different mating systems

Summary Morphological variability in three Plantago species possessing different mating systems — P. Major, P. coronopus, and P. lanceolata — was analyzed. From each species several populations were analyzed. In comparing the species, it appeared that both the inbreeding P. major and the outbreeding P. lanceolata showed a high degree of morphological differentiation between populations. It was hypothesized that the absence of population differentiation for the species with a mixed mating system, P. coronopus, was caused by high adaptive plasticity. Plasticity is probably the predominant mechanism by which this species copes with different environments. There were considerable differences between populations within each species for their levels of variability and their heritabilities. It appeared that besides the influence of the mating system, selection may diminish morphological variability in the case of strong directional selection, especially in P. major and P. lanceolata.Grassland Species Research Group Publication No. 135     

Analysis of allozyme variability in three Plantago species and a comparison to morphological variability

Summary The level of electrophoretic variability in three Plantago species, P. major, P. coronopus, and P. lanceolata, was analyzed in relation to their breeding systems and compared with their morphological variability. From each species several populations were analyzed. The outcrossing P. lanceolata had the highest level of electrophoretic variability and the lowest population differentiation. The inbreeding P. major showed the opposite: a low level of electrophoretic variability and a high population differentiation. P. coronopus, with an intermediate breeding system, had an intermediate level of variability and differentiation. In comparing the species, it appeared that P. coronopus and P. major showed good concordance in the distribution of both kinds of variability, each having only a slightly higher morphological than electrophoretic differentiation between populations. P. lanceolata showed a higher morphological than electrophoretic differentiation between populations. A comparison of populations, within species, revealed good concordance of electrophoretic and morphological variability only within P. coronopus, while some populations of the other two species had relatively lower morphological variability compared with electrophoretic variability.Grassland Species Research Group Publication No. 182     

Lipid Composition of the Roots of Plantago Species: Response to Alteration of the Level of Mineral Nutrition and Ecological Significance

In a study of the adaptation of Plantago species to their specific environment the lipid composition of the roots of several species: Plantago major L. ssp. major, Plantago major L. ssp. pleiosperma Pilger, Plantago lanceolata L., Plantago media L., Plantago maritima L., Plantago coronopus L. was studied as well as the effect of the nutritional regime. Upon exposure to low-salt conditions Plantago major L. ssp. major L. and Plantago maritima L. maintained the level of free sterols in the roots, despite a depressed level of total sterols, and the root lipids were more saturated than under high-salt conditions. Both factors may reduce nutrient leakage from the roots to the low-salt condition. Upon exposure to low-salt conditions, all Plantago species showed a decreased level of galactolipid (exception: Plantago coronopus) and a decreased level of sitosterol (exception: Plantago maritima); the latter being compensated by an elevated level of cholesterol + tocopherol in Plantago major ssp. pleiosperma, Plantago lanceolata and Plantago media. Plantago coronopus was the only species which under low-salt conditions showed an increased level of free sterols, among which cholesterol was the most important; thus indicating a high degree of regulation of membrane permeability under alternating nutritional conditions. The level of sulfolipid was kept constant in all Plantago species, with the highest level observed in Plantago maritima. The role which various lipids may play in maintenance of membrane integrity under alternating nutritional conditions is discussed.     

The effect of nitrate concentration in a flowing solution system on growth and nitrate uptake of twoPlantago species

Summary Juvenile plants ofPlantago lanceolata andP. major ssp.major were grown in a flowing solution system at 7.5 mM or 9.5 M NO₃. The parameters investigated were: RGR, shoot weight percentage, leaf length, length of main root axis, shoot concentrations of major ions and organic N, and the specific uptake rate for NO₃. At 9.5 M NO₃ growth ofP. major was not hampered, whereas shoot growth and leaf length ofP. lanceolata were reduced. The NO₃ concentration ofP. lanceolata decreased more than that ofP. major. The different performances of the species at 9.5 M NO₃ were associated with different specific uptake rates. In both treatments the root system ofP. major was shorter than that ofP. lanceolata. P. lanceolata accumulated more NO₃ in the leaves. The performance of thePlantago species is discussed in relation to the availability of nutrients in their habitats.Grassland Species Research Group. Publication no. 37.     

Comparison of Plantago Species from Nutrient-rich and Nutrient-poor Conditions: Growth Response, ATPases and Lipids of the Roots, as Affected by the Level of Mineral Nutrition

Two groups of Plantago species, one of relatively nutrient-rich habitats (Plantago major L., P. major L. ssp. pleiosperma Pilger and P. martrima L.) and the other of relatively nutrient poor habitats (P. lanceolata L., P. media L. and P. coronopus L.), were compared with respect to the response of the plants to alterations of the level of mineral nutrition. The comparison deals with growth response and root cell membrane functions (Ca²⁺ and Mg²⁺ stimulated ATPases and lipids). As far as these parameters are concerned species from relatively nutrientpoor habitats are not much affected by bringing the plants to high-salt conditions, while the reverse was true for species from relatively nutrient-rich habitats when transferred to low salt medium.     

Host plant species affects the quality of the generalist Trichoplusia ni as a host for the polyembryonic parasitoid Copidosoma floridanum

Diet of herbivorous insects can influence both the herbivores and their natural enemies. We examined the direct and indirect effects of diet on the interactions between the polyphagous herbivore Trichoplusia ni Hübner (Lepidoptera: Noctuidae) and its polyembryonic parasitoid Copidosoma floridanum Ashmead (Hymenoptera: Encyrtidae). To determine how host plant species and host plant iridoid glycoside content affect host caterpillars and their parasitoids, parasitized and unparasitized T. ni were given leaves of either Plantago lanceolata L., which contains the iridoid glycosides aucubin and catalpol, Plantago major L. (Plantaginaceae), which contains only aucubin, or Taraxacum officinale F.H. Wigg (Asteraceae), which contains neither. Survival of unparasitized T. ni was much lower when fed P. major compared with the other two host plants, whereas pupae were smallest when fed T. officinale and developed most slowly when fed P. lanceolata as larvae. Neither aucubin nor catalpol were detected in intact Plantago‐fed T. ni larvae or their hemolymph, and only trace amounts of aucubin were detected in frass, suggesting that these compounds are mostly metabolized in the midgut and are not encountered by the parasitoid. Copidosoma floridanum clutch size was almost doubled when reared from P. lanceolata‐fed T. ni compared with T. officinale‐fed larvae and tripled compared with P. major‐fed larvae, although the percent of parasitoids surviving to adulthood was uniformly high regardless of host diet. The observed variation in C. floridanum fitness among host diets is likely mediated by the effect of the diets on host quality, which in turn may be influenced more by other factors in the host plants than their iridoid glycoside profiles. Interactions between plant metabolites, generalist herbivores like T. ni, and their parasitoids may be predominantly indirect.     

Establishment of Plantago lanceolata L. and Plantago major L. among grass

Summary Establishment of Plantago lanceolata and P. major ssp major among grass was studied in a field experiment in which survival and selection on date of seedling emergence and plant size was investigated in relation to the vegetation structure. P. major — in contrast to P. lanceolata — was not able to establish itself in grass because of its lower competitive ability caused by later germination, smaller seedling size, and shorter leaves. In both species there was selection for early germination. For P. lanceolata a significant correlation was found between the strength of selection and the light climate, determined by the structure of the grass sward. Plants that germinated early were at an advantage because they were larger, especially the leaves, when compared with plants that germinated late. It seems likely that selection was mainly by competition for light. Contrary to expectation P. major-seedlings had a higher shade tolerance than those of P. lanceolata. The performance of both species is discussed in relation to their different life strategies.Grassland species research group publication no 142     

Variations in pollen proportions of Plantago lanceolata and P. major/media at a Neolithic lake dwelling,Lake Chalain,France

Detailed pollen analytical investigations at a Neolithic lake dwelling site on Lake Chalain, Jura, France, show very characteristic variation in the Plantago lanceolata and P. major/media pollen curves in the period during which settlements are recorded (3030–2630 cal. B.C.). At first, P. lanceolata is the more important taxon but P. major/media representation gradually rises, to become the more important taxon in the uppermost settlement phase. After considering the present day ecology and phytosociology of the Plantago species in question, i.e. P. lanceolata, P. major and P. media, and the available archaeozoological and archaeological information, it is suggested that the changes in the representation of these two pollen taxa are the result of a change in the farming economy, at ca. 2800 cal. B.C., which involved a substantial rise in the numbers of domesticated grazing animals and more intensive land use.     

Genetic systems of six species ofPlantago (Plantaginaceae)

Investigation of the genetic system of six species ofPlantago has revealed striking differences in their breeding and meiotic systems.Plantago patagonica is an inbreeder on account of preanthesis cleistogamy, whereasP. lanceolata is an obligate outbreeder, as it is self-incompatible.Plantago drummondii, P. lagopus, P. ovata, andP. major show mixed mating but in varying proportions. In terms of their energy budgets, outbreeding species invest more in floral advertisement and male function, while inbreeders invest more in female function. The contribution of the meiotic system to genetic variability, as revealed through recombination index, is more important in the inbreeding species.     

Neighbor species differentially alter resistance phenotypes in Plantago

In this study, we investigated how neighbors (i.e., competitors) altered resistance phenotypes, namely plant size and levels of secondary compounds (iridoid glycosides), of individual plants and specifically tested whether neighbor identity mattered. We conducted a greenhouse experiment with Plantago lanceolata and Plantago major (Plantaginaceae) in which each species served as focal plants as well as neighbors in a factorial design. In addition, we harvested plants six and nine weeks after transplantation to test whether effects changed as plants grew. In both species, competition reduced plant size, and this effect increased over time. Plantago lanceolata neighbors suppressed growth of both focal plant species more than P. major neighbors. Effects of competition on levels of secondary compounds were more complex. Concentrations of iridoid glycosides were increased by competition in both species at harvest one. By the second harvest, an effect of competition on iridoid glycosides was found only in P. major. Neighbor identity influenced levels of iridoid glycosides in P. lanceolata at harvest one; concentrations were higher in plants grown with P. lanceolata neighbors than in plants grown with P. major neighbors. We also tested whether there was a trade-off between growth (biomass) and defense (levels of iridoid glycosides). Biomass and iridoid glycoside content were significantly correlated only in plants grown with competition and harvested at nine weeks, and this relationship was positive in both species, indicating that there was no trade-off between growth and defense. This study suggests that neighbor identity could play an important role in interspecific interactions, including the interactions of plants with other trophic levels.     

High thermal acclimation potential of both photosynthesis and respiration in two lowland Plantago species in contrast to an alpine congeneric

Thermal acclimation of photosynthesis and respiration can enable plants to maintain near constant rates of net CO₂ exchange, despite experiencing sustained changes in daily average temperature. In this study, we investigated whether the degree of acclimation of photosynthesis and respiration of mature leaves differs among three congeneric Plantago species from contrasting habitats [two fast‐growing lowland species (Plantago major and P. lanceolata), and one slow‐growing alpine species (P. euryphylla)]. In addition to investigating some mechanisms underpinning variability in photosynthetic acclimation, we also determined whether leaf respiration in the light acclimates to the same extent as leaf respiration in darkness, and whether acclimation reestablishes the balance between leaf respiration and photosynthesis. Three growth temperatures were provided: constant 13, 20, or 27°C. Measurements were made at five temperatures (6–34°C). Little acclimation of photosynthesis and leaf respiration to growth temperature was exhibited by P. euryphylla. Moreover, leaf masses per area (LMA) were similar in 13°C‐grown and 20°C‐grown plants of the alpine species. In contrast, growth at 13°C increased LMA in the two lowland species; this was associated with increased photosynthetic capacity and rates of leaf respiration (both in darkness and in the light). Alleviation of triose phosphate limitation and increased capacity of electron transport capacity relative to carboxylation were also observed. Such changes demonstrate that the lowland species cold‐acclimated. Light reduced the short‐term temperature dependence (i.e. Q₁₀) of leaf respiration in all three species, irrespective of growth temperature. Collectively, our results highlight the tight coupling that exists between thermal acclimation of photosynthetic and leaf respiratory metabolism (both in darkness and in the light) in Plantago. If widespread among contrasting species, such coupling may enable modellers to assume levels of acclimation in one parameter (e.g. leaf respiration) where details are only known for the other (e.g. photosynthesis).     

Ammonium and nitrate nutrition inPlantago lanceolata L. andPlantago major L. ssp.major

With the aims (1) to test whether the different natural occurrence of twoPlantago species in grasslands is explained by a different preference of the species for nitrate or ammonium; (2) to test whether the different occurrence is explained by differences in the flexibility of the species towards changes in the nitrogen form; (3) to find suitable parameters as a tool to study ammonium and nitrate utilization of these species at the natural sites in grasslands, plants ofPlantago lanceolata andP. major ssp.major were grown with an abundant supply of nitrate, ammonium or nitrate+ammonium as the nitrogen source (0.5 mM). The combination of ammonium and nitrate gave a slightly higher final plant weight than nitrate or ammonium alone. Ammonium lowered the shoot to root ratio inP. major. Uptake of nitrate per g root was faster than that of ammonium, but from the mixed source ammonium and nitrate were taken up at the same rate. In vivo nitrate reductase activity (NRA) was present in both shoot and roots of plants receiving nitrate. When ammonium was applied in addition to nitrate, NRA of the shoot was not affected, but in the root the activity decreased. Thus, a larger proportion of total NRA was present in the shoot than with nitrate alone. In vitro glutamate dehydrogenase activity (GDHA) was enhanced by ammonium, both in the shoot and in the roots.In vitro glutamine synthetase activity (GSA) was highest in roots of plants receiving ammonium. Both GDHA and GSA were higher inP. lanceolata than inP. major. The concentration of ammonium in the roots increased with ammonium, but it did not accumulate in the shoot. The concentration of amino acids in the roots was also enhanced by ammonium. Protein concentration was not affected by the form of nitrogen. Nitrate accumulated in both the shoot and the roots of nitrate grown plants. When nitrate in the solution was replaced by ammonium, the nitrate concentration in the roots decreased rapidly. It also decreased in the shoot, but slowly. It is concluded that the nitrogen metabolism of the twoPlantago species shows a similar response to a change in the form of the nitrogen source, and that differences in natural occurrence of these species are not related to a differential adaptation of nitrogen metabolism towards the nitrogen form. Suitable parameters for establishing the nitrogen source in the field are thein vivo NRA, nitrate concentrations in tissues and xylem exudate, and the fraction of total reduced nitrogen in the roots that is in the soluble form, and to some extent thein vitro GDHA and GSA of the roots. Grassland Species Research Group. Publ. no 118.     

Low levels of intraspecific genetic variation at a rapidly evolving chloroplast dna locus in North American duckweeds (Lemnaceae)

Although most previous studies on chloroplast (cp) DNA variation in plants have concentrated on systematics and evolution above the species level, intraspecific variation in cpDNA is common and has provided useful insights into population-level evolutionary processes. Polymerase chain reaction methods were used to examine restriction site and sequence variation in the chloroplast rpLI6 gene within and among populations of duckweed species (Spirodela and Lemna) from the southern and eastern United States. To our knowledge, the rpL16 region has not previously been used to investigate cpDNA variation in nature. While considerable restriction site and sequence variation were detected among species, no variation was found within populations of either of the two species (S. punctata and L. minor) selected for sequence analysis, and S. punctata showed no interpopulational variation. Two cpDNA haplotypes were identified in L. minor, with one haplotype restricted to three sites in Louisiana and the other found in all other populations sampled. This paucity of variation cannot be readily explained as the result of a low mutation rate. In general, group II introns appear to be subject to very little functional constraint, and extensive sequence differences have been found between species in the chloroplast rpL16 intron in particular. However, factors such as historical range expansions and contractions, founding effects, fluctuations in local population size, and natural selection may play a role in reducing cpDNA sequence variability in these species.     

Genetic variation and biogeography of the disjunct Vitis subg. Vitis (Vitaceae)

Aim Vitis subg. Vitis provides an example of a plant disjunction occurring in the Northern Hemisphere. It shows broad morphological variation but is assumed to be a species complex with limited genetic differentiation. Based on a comprehensive sampling of taxa and polymorphism in both chloroplast and nuclear DNA, we assessed genetic variation within this subgenus. Our aims were to clarify the relationships among species and to examine their historical biogeography. Location Asia, Europe, North America. Methods We analysed a total of 30 species and putative hybrids from subgenus Vitis and examined the infra‐specific variation in some species. Polymorphism in chloroplast DNA was assessed in trnL and trnH–psbA–trnK sequences (c. 2170 bp) and in 15 microsatellite loci. We also obtained nuclear data for size variation at 24 microsatellite loci. Phylogenetic inference was performed with Bayesian analyses. A maximum parsimony network was constructed to depict the evolutionary relationships among haplotypes, and microsatellite data were also subjected to hierarchical clustering analysis using the Ward distance. In addition, we assessed size homoplasy by sequencing both chloroplast and nuclear microsatellite loci. Results Chloroplast polymorphisms resolved subgenus Vitis as a monophyletic group with limited genetic variation. The ancestral haplotypes were found in Eurasia. American taxa all harboured derived haplotypes. Most of them formed a monophyletic group that did not include Vitis californica. The four main haplotypes in Vitis vinifera corresponded to two different origins. Nuclear microsatellites indicated that genetic variation was especially large in North America. Asian species exhibited a lower level of nuclear divergence and the European V. vinifera corresponded to a differentiated nuclear lineage. Main conclusions We obtained some evidence that subgenus Vitis has an Asian origin and then dispersed to Europe and North America. Geographic separation was followed by diversification, presumably during the Pleistocene, resulting in phylogeographic patterns similar to other biota. In contrast to chloroplast DNA, nuclear DNA shows a larger than expected genetic variation. Our molecular data also highlight the need to re‐examine certain aspects of the current subgeneric classification.     

Chloroplast evidence for geographic stasis of the Australian bird‐dispersed shrub Tasmannia lanceolata (Winteraceae)

Few chloroplast‐based genetic studies have been undertaken for plants of mesic temperate forests in the southern hemisphere and fossil‐based models have provided evidence of vegetation history only at the broadest scales in this region. This study investigates the chloroplast DNA phylogeography of Tasmannia lanceolata (Winteraceae), a fleshy‐fruited, bird‐dispersed shrub that is widespread in the mountains of southeastern Australia and Tasmania. Thirty haplotypes were identified after sequencing 3206 bp of chloroplast DNA in each of 244 individuals collected across the species’ range. These haplotypes showed unexpectedly strong phylogeographic structuring, including a phylogeographic break within a continuous part of the species’ range, with the distribution of four major clades mostly not overlapping, and geographic structuring of haplotypes within these clades. This strong geographic patterning of chloroplast DNA provided evidence for the survival of T. lanceolata in multiple putative wet forest refugia as well as evidence for additional wet forest species refugia in southeastern Australia. In western Tasmania lower haplotype diversity below the LGM tree line compared to above the LGM tree line suggests that glacial refugia at high altitudes may have been important for T. lanceolata. The level of geographic structuring in T. lanceolata is similar to gravity dispersed southern hemisphere plants such as Nothofagus and Eucalyptus. Behavioural traits of the birds transporting seed may have had a strong bearing on the limited transport of T. lanceolata seed, although factors limiting establishment, possibly including selection, may also have been important.     

Changes in ion composition and hexitol content of differentPlantago species under the influence of salt stress

Summary The effect of salinity on the ionic balance and the hexitol content of the halophyteP. maritima L. and the nonhalophytesP. major L. ssp.major, P. lanceolata L., andP. media L. was studied in a culture experiment. In response to salt application the nonhalophilous species increased their internal electrolyte content to a considerably greater extent at high transpiration thanP. maritima. Excessive uptake of Cl and SO₄ following salt-treatment was in most cases compensated by a decline in the total organic anion content in all species under investigation. Na was strongly accumulated in the shoots ofP. maritima when subjected to salt-stress, while the nonhalophytic species tended to exclude this ion from leaf tissue enhancing Mg-uptake for charge balance. Acyclic polyhydric alcohols (sorbitol and mannitol), the dominant soluble carbohydrates in allPlantago species studied, increased, with one exception, in all plants under saline conditions. The results indicate marked physiological differences between the halophyteP. maritima and the nonhalophytic members of the genusPlantago in their reaction to salinity being perhaps partially responsible for differences in the degree of salt tolerance.     

Patterns of chloroplast diversity among western European Dactylorhiza species (Orchidaceae)

In Europe, the genus Dactylorhiza comprises a bewildering variety of forms that are difficult to sort into discrete species. Most Dactylorhiza species are diploid or tetraploid and contrasting hypotheses have been proposed to explain the complex variation within this group. Using PCR-RFLP analysis in eight putative species, we could identify four highly differentiated chloroplast DNA lineages. The first lineage (clade A) included the unique haplotype found in D. sambucina. Clade B grouped four haplotypes belonging mostly to D. incarnata. Clades C and D included 27 haplotypes detected in diploid D. fuchsii and in all tetraploid species investigated. Eighty percent of the chloroplast variation were consistent with currently accepted species boundaries. The imperfect agreement between chloroplast variation and species boundaries may be ascribed to incomplete lineage sorting and/or reticulation. Our cpDNA results provide strong evidence that the allotetrapolyploids have been formed through asymmetric hybridization with a member of the D. fuchsii / maculata group as the maternal parent.