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     

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 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     

Nitrate production in the rhizosphere of Plantago species

Summary The production of nitrate in an old established dune grassland soil and its uptake by plants was studied by comparing amounts of mineral nitrogen and numbers of nitrifying bacteria in the rhizosphere on the one hand, and on the other accumulated nitrate and levels of nitrate reductase (NaR) of individual plants of three Plantago species,i. e., P. major, P. lanceolata andP. coronopus. For these three Plantago species andP. media basal levels of NaR in the absence of nitrate were determined in plants grown in culture solutions. The basal NaR levels ofP. major andP. media (species occurring on nutrient-rich soils) were significantly higher than those ofP. lanceolata andP. coronopus (species found on nutrient-poor soils). NaR activity increased in the presence of nitrate and was suppressed by ammonium.From the numbers of nitrifying bacteria in the rhizosphere and NaR activity in the leaves it was concluded that nitrate was produced in the root environments of the three Plantago species and that the compound was taken up by the plants. NaR activities and numbers of nitrifying bacteria were higher for individuals ofP. major than for those ofP. lanceolata andP. coronopus. No correlation was found between the ammonium levels and the numbers of nitrifying bacteria in the soil, and no indications of inhibition of nitrifying bacteria in the rhizosphere were obtained. For individuals ofP. lanceolata a correlation was found between the numbers of nitrifying bacteria in the soil and NaR activity in the leaves. The results are discussed in relation to the ecological habitats of the three species.Grassland Species Research Group Publication No.38.     

Contrasting patterns of genetic diversity in two tropical pines: Pinus kesiya (Royle ex Gordon) and P. merkusii (Jungh et De Vriese)

We studied allozyme and chloroplast (cp) DNA variation in natural populations of Pinus kesiya and P. merkusii from Thailand and Vietnam. The results showed striking differences between the two species in the amount and distribution of allozyme variation. P. kesiya harboured considerable allozyme variation and showed weak interpopulational differentiation. In contrast, P. merkmii had very low intrapopulational variability but a high level of interpopulational differentiation. The average Nei's genetic distance separating the two species was exceptionally high (0.701) taking into account their close taxonomic placement in the same subsection Sylvestres. The constructed phylogenetic trees revealed very early divergence of P. kesiya and P. merkusii. The present analysis of cpDNA variation also confirmed the dissimilar character of these two species and was compatible with other evidence indicating the outstanding position of P. merkusii as compared to other Asian members of the subsection Sylvestres. Analysis of cpDNA variation in sympatric populations of P. kesiya and P. merkusii revealed that they are pure representatives of the species in question. This result indicates that despite an overlapping distribution P. kesiya and P. merkusii do not hybridise in nature. We suggest that the distinctive character of P. merkusii is a result of an early separation from other Eurasian pines. Despite spatial proximity, P. kesiya and P. merkusii are kept apart by strong reproductive barriers. The low genetic variability of P. merkusii may be explained by previous bottlenecks, reduced gene flow among populations, and an inbreeding due to small population size and asynchronous flowering.     

Ca2+ and Mg2+-Stimulated ATPases from Roots of Plantago lanceolata, Plantago media and Plantago coronopus: Response to Alterations of the Level of Mineral Nutrition and Ecological Significance

Growth of Plantago lanceolata L., P. media L. and P. coronopus L. was followed at two levels of mineral nutrition (low-salt and high-salt). In addition the response of transfer of plants from low-salt conditions to high-salt conditions and vice versa was studied. Growth of these Plantago species was not much affected by the nutritional level. P. coronopus showed the least dependence on the level of mineral nutrition. The Ca²⁺- and Mg²⁺-stimulated ATPase activity of microsomal preparations of the roots of Plantago was also studied. A pH optimum at pH 6.5 was observed in all species, together with a relatively high ATPase activation by Mg²⁺ in P. lanceolata, by Ca²⁺ in P. media, and by both ions in P. coronopus. The specific activity of the ATPases was highest in preparations from low-salt roots. The three species all occur in relatively nutrient-poor habitats, but they are at the same time particularly adapted to circumneutral soils (P. lanceolata), to Ca²⁺-rich soils (P. media) and to alternating levels of mineral nutrition (P. coronopus). The properties of the ATPases (K_m, V_max, protein content) and the growth are discussed in relation to these ecological properties of the species.     

Chloroplast DNA variation within and among five Plantago species

Chloroplast DNA variation was studied within and among five Plantago species. We determined polymorphism by surveying 86% of the chloroplast genome using 9 or 11 restriction enzymes for intra or interspecific variation, respectively. All five species were polymorphic for both site and length mutations. The outcrossing P. lanceolata has six chloroplast haplotypes of which some were found in the Netherlands as well as in the United States. The highly selfing P. major had five haplotypes and Dutch and United States populations were differentiated from each other in cpDNA. Plantago species are highly differentiated from each other. Of 252 restriction-sites, 52% were variable among species. Most of this variation was localized in part of the large single copy region. We derived a molecular phylogeny of the five species from restriction-site variability using PAUP 3.0. P. coronopus and P. maritima form a group as do P. major and P. media. P. lanceolata is more distantly related to the other four species. In a consensus tree both P. major haplotypes and both P. media haplotypes were connected to one node.     

Genetic structure of Polytrichum formosum in relation to the breeding system as revealed by microsatellites

Microsatellite variation was determined for three Danish and three Dutch populations of the haploid moss species Polytrichum formosum to gain insight into the relative importance of sexual vs. asexual reproduction for the amount and structure of genetic variation. In general, low levels of microsatellite variation were observed within this species. Even when estimated for polymorphic loci only, the levels of microsatellite variability (P=90.6, A=4.3 and H_S=0.468) within populations were on average lower than those reported for most other plant species. In contrast, genotypic diversity was high within each of the examined populations, indicating that sexual reproduction is a very important determinant of the genetic structure of P. formosum within populations. In agreement with previous findings for allozyme data, no significant genetic differentiation (F_ST=0.028, R_ST=0.015) was observed neither between populations nor between regions approximately 450 km apart (Denmark vs. the Netherlands). These low levels of population differentiation observed for both types of genetic markers are probably best explained by a high level of effective spore dispersal (gene flow) between populations. Therefore, also on a large geographical scale sexual reproduction is the most important determinant of the genetic structure of P. formosum, despite the high potential to reproduce clonally.     

Allozyme variation and population genetic structure of common wild rice Oryza rufipogon Griff. in China

In order to determine the genetic diversity and genetic structure of populations in common wild rice Oryza rufipogon, an endangered species, allozyme diversity was analyzed using 22 loci in 607 individuals of 21 natural populations from the Guangxi, Guangdong, Hainan, Yunnan, Hunan, Jiangxi and Fujian provinces in China. The populations studied showed a moderate allozyme variability (A=1.33, P=22.7%, Ho=0.033 and He=0.068), which was relatively high for the genus Oryza. The levels of genetic diversity for Guangxi and Guangdong were significantly higher than those for the other regions, and thus South China appeared to be the center of genetic diversity of O. rufipogon in China. A moderate genetic differentiation (F_ST=0.310, I=0.964) was found among the populations studied. Interestingly, the pattern of population differentiation does not correspond to geographic distance. An estimate of the outcrossing rate (t=0.324) suggests that the species has a typical mixed-mating system. The deficit of heterozygotes (F=0.511) indicates that some inbreeding may have taken place in outcrossing asexual populations because of intra-clone outcrossing events and ”isolation by distance” as a result of human disturbance. In order to predict the long-term genetic survival of fragmented populations, further studies on gene flow among the remaining populations and the genetic effects of fragmentation are proposed. Finally, some implications for the conservation of endangered species are suggested. Received: 22 June 1999 / Accepted: 20 December 1999     

THE INFLUENCE OF INTRASPECIFIC VARIATION IN DISPERSAL STRATEGIES ON THE GENETIC STRUCTURE OF PLANTHOPPER POPULATIONS

The hypothesis that levels of gene flow among populations are correlated with dispersal ability has typically been tested by comparing gene flow among species that differ in dispersal abilities, an approach that potentially confounds dispersal ability with other species-specific differences. In this study, we take advantage of geographic variation in the dispersal strategies of two wing-dimorphic planthopper species, Prokelisia marginata and P. dolus, to examine for the first time whether levels of gene flow among populations are correlated with intraspecific variation in dispersal ability. We found that in both of these coastal salt marsh–inhabiting species, population-genetic subdivision, as assessed using allozyme electrophoresis, parallels geographic variation in the proportion of flight-capable adults (macropters) in a population; in regions where levels of macroptery are high, population genetic subdivision is less than in regions where levels of macroptery are low. We found no evidence that geographic variation in dispersal capability influences the degree to which gene flow declines with distance in either species. Thus, both species provided evidence that intraspecific variation in dispersal strategies influences the genetic structure of populations, and that this effect is manifested in population-genetic structure at the scale of large, coastal regions, rather than in genetic isolation by distance within a region. This conclusion was supported by interspecific comparisons revealing that: (1) population-genetic structure (G_ST) of the two Prokelisia species correlated negatively with the mean proportion of flight-capable adults within a region; and (2) there was no evidence that the degree of isolation by distance increased with decreasing dispersal capability. Populations of the relatively sedentary P. dolus clustered by geographic region (using Nei's distances), but this was not the case for the more mobile P. marginata. Furthermore, gene flow among the two major regions we surveyed (Atlantic and Gulf Coasts) has been substantial in P. marginata, but relatively less in P. dolus. The results for P. marginata suggest that differences in the dispersal strategies of Atlantic and Gulf Coast populations occur despite extensive gene flow. We argue that gene flow is biased from Atlantic to Gulf Coast populations, indicating that selection favoring a reduction in flight capability must be intense along the Gulf. Together, the results of this study provide the first rigorous evidence of a negative relationship within a species between dispersal ability and the genetic structure of populations. Furthermore, regional variation in dispersal ability is apparently maintained by selective differences that outweigh high levels of gene flow among regions.     

Bottlenecks, drift and differentiation: the fragmented population structureof the saltmarsh beetle Pogonus chalceus

We investigated the distribution of genetic variation within and between 10 fragmented populations of the saltmarsh beetle Pogonus chalceus in the region of Flanders (Belgium) representing all extant populations of the species in that region by using allozyme and microsatellite markers. Beetle population size and habitat area failed to explain a significant part of the genetic variability. Microsatellite allelic diversity was sensitive to population size differences but not to saltmarsh area estimates. Heterozygosities of both marker types and allozyme allelic diversity on the other hand showed no significant correlation to population size and saltmarsh area. There was also no correlation between geographical and genetic distances among populations. Evidence was found for past bottlenecks in some of the smallest populations. Maximum likelihood methods using the coalescent approach revealed that the proportion of common ancestors was also high in those small populations. 35% of our studied individuals, especially in the largest populations showed a relative wing size smaller than 70%. Moreover, only six out of the 10 studied populations showed a few individuals with functional flight musculature. In conclusion, the overall pattern of distribution of genetic variation and the low flight capacity did not support an equilibrium model of population structure in P. chalceus, but mainly suggested a lack of regional equilibrium with both drift and gene flow influences.     

Genetic differentiation and detection of cryptic species in the genus Lepismachilis (Insecta: Microcoryphia) from the Western Mediterranean region

Different species of the bristletail genus Lepismachilis were collected in 14 localities in Italy and Spain and an allozyme electrophoretic survey was carried out to estimate the degree of genetic variability and differentiation at intra- and interspecific levels. Four morphological species were initially identified (L osellai, L. y-signata, L. affinis, L. targionii), but the electrophoretic analysis demonstrated the presence of two additional species among the individuals of L. targionii (Lepismachilis spl and sp2). The validity of these species and their differentiation from L targionii were demonstrated by the fixation of alternative allelic patterns at several loci (7 in Lepismachilis spl and 8 in Lepismachilis sp2), coupled with fixed, previously undetected, morphological differences. In addition, Lepismachilis sp2 was sympatric with L. targionii in three collecting sites, where the fixation of alternative allelic patterns unequivocally demonstrated reproductive isolation. Genetic variability did not seem to be correlated with local ecological factors, and differences between species should rather be explained by different historical factors. Low levels of gene flow, estimated with two different indirect methods, were observed in L. targionii and L. y-signata, and were due to high levels of structuring among populations. Genetic differentiation among conspecific populations was not correlated to their geographical arrangement and the presence of loci fixed for different alleles among them suggested that stochastic factors (such as genetic drift) may have played a role in determining genetic differentiation of geographically isolated populations. Genetic divergence values indicated that the six species are well differentiated and allozyme profiles were diagnostic for all of them. On the other hand, allozyme data did not provide adequate information to resolve evolutionary relationships among the species, nor did they confirm the validity of the two subgenera (Lepismachilis and Berlesilis) in which the genus Lepismachilis is traditionally divided.     

Genetic Structure and Gene Flow in Elymus glaucus (blue wildrye): Implications for Native Grassland Restoration

Interest in using native grass species for restoration is increasing, yet little is known about the ecology and genetics of native grass populations or the spatial scales over which seed can be transferred and successfully grown. The purpose of this study was to investigate the genetic structure within and among populations of Elymus glaucus in order to make some preliminary recommendations for the transfer and use of this species in revegetation and restoration projects. Twenty populations from California, Oregon, and Washington were analyzed for allozyme genotype at 20 loci, and patterns of variation within and among populations were determined. Allozyme variation at the species level was high, with 80% of the loci polymorphic and an average expected heterozygosity (an index of genetic diversity) of 0.194. All but two of the populations showed some level of polymorphism. A high degree of population differentiation was found, with 54.9% of the variation at allozyme loci partitioned among populations (F_st= 0.549). A lesser degree of genetic differentiation among closely spaced subpopulations within one of the populations was also demonstrated (F_st= 0.124). Self-pollination and the patchy natural distribution of the species both likely contribute to the low level of gene flow (N_m= 0.205) that was estimated. Zones developed for the transfer of seed of commercial conifer species may be inappropriate for transfer of E. glaucus germplasm because conifer species are characterized by high levels of gene flow. Limited gene flow in E. glaucus can facilitate the divergence of populations over relatively small spatial scales. This genetic differentiation can be due to random genetic drift, localized selective pressures, or both. In order to minimize the chances of planting poorly adapted germplasm, seed of E. glaucus may need to be collected in close proximity to the proposed restoration site.     

Measures of gene flow in the Columbian ground squirrel

From analyses of published data and a review of the literature, I studied indirect and direct measures of gene flow among populations of Columbian ground squirrels, Spermophilus columbianus. New analyses were used to examine an allozyme data set (seven polymorphic loci) that had been collected by Zammuto and Millar (1985a) from six populations of ground squirrels that were spread over 183 km. G-tests indicated significant variation in allele frequencies among populations, but F-statistics revealed relatively little population differentiation (average F _ST=0.026). F _ST values were used to estimate rates of gene flow indirectly and indicated fairly high rates of gene flow (average N _e m=13.5). Recorded dispersal distances of individual ground squirrels were fairly short (most<4 km, maximum recorded distance was 8.5 km), and the minimum distance between populations used to create the allozyme data set was about 25 km. Thus, direct dispersal among the populations in the allozyme data set was highly unlikely. Small genetically effective populations may have experienced high rates of migration over short distances (about 43% of adults in local populations were immigrants), however, resulting in homogeneous allele frequencies over the geographic range. This explanation provides an alternative to invoking gene flow in the recent past to explain discrepancies between dispersal distances in the field and homogenization of allele frequencies over large ranges, Mammalian species that have virtually complete dispersal of subadult males from the natal area might be expected to exhibit relatively high rates of gene flow, regardless of actual dispersal distances. Genetically effective populations may be much smaller than more extensive ecological populations and experience higher rates of gene flow.     

Genetic structure of natural populations of Castanea sativa in Turkey: evidence of a hybrid zone

This study points out the evidence of a hybrid zone between two groups of genetically differentiated populations of chestnut (Castanea sativa Mill.) in Turkey. Genetic structure, gene flow and introgression levels, based on 16 allozyme loci, were investigated on 34 population samples spanning the entire C. sativa distribution area in this country. The occurrence of the hybrid zone, located in the Bithynian region, was inferred in a group of populations showing the following genetic characteristics: (i) enhanced genetic variability and intermediate allelic frequencies between those of the Western and Eastern groups of populations; (ii) sharp and concordant changes in allele frequencies; (iii) decreased gene flow with the Western and Eastern populations. Starting from the cline width estimated to be 324 km, strength of selection was evaluated from the gene flow distance, as indicated from the degree of genetic structuring outside the hybrid zone. Evolutionary processes shaping the observed genetic differentiation and introgression are discussed on the basis of palynological data, palaeoclimatic events and evidence of hybridization found in other plant and animal species in the same region.     

Gene flow measured by allozymic analysis in pesticide resistant Typhlodromus pyri occurring within and near apple orchards

Electrophoresis of allozymes was used to estimated gene flow among populations of Typhlodromus puri Scheuten from apple orchards and nearby blackberry plants from two valleys of western Oregon, USA. Four allozyme loci unaffected by pesticides were tested. Wright's coefficient of inbreeding, F _ST, for all populations was 0.115 and the proportion of populations that migrated per generation, Nm, was 2.08. These values were higher than expected for such locally collected mites. No allelic patterns could be discerned for populations among or within valleys: however, more variation was found for mites collected within than between valleys. From other studies of dispersal and pesticide resistance, we concluded that T. pyri had a low dispersal rate, but these data from allozymic analysis indicated there was moderate gene flow among populations. We concluded that the unique features of the population dynamics of T. pyri may account for the differences seen in estimating gene flow when using different types of population assessment (i.e., dispersal distances, resistance rates and allozyme frequency studies).     

Genetic diversity and morphological differentiation in Liatris helleri (Asteraceae), a threatened plant species

Liatris helleri (Asteraceae) is an insect-pollinated herbaceous perennial endemic to several high-elevation sites in the Blue Ridge Mountains of North Carolina. Allozymes were used to describe the genetic diversity and population structure in nine populations of this rare, federally listed species. Differences in leaf morphology were also examined for greenhouse-grown plants representing several populations. The proportion of the total genetic diversity found among populations, as indicated by the allozyme data, was 16%. Higher levels of population differentiation were found for differences in leaf shape; population of origin accounted for 37% of the variation in maximum leaf width, while families within populations accounted for 7%. In contrast to many endemic species, L. helleri maintains fairly high levels of genetic diversity. For the species, the percent polymorphic loci was 87.5, the average number of alleles at variable loci was 3.00 and the gene diversity was 0.276. Mean population values were percent polymorphic loci =58.4, mean number of alleles per polymorphic locus =2.59 and gene diversity =0.219. The estimated gene flow was low (Nm=0.85–1.32) and a relatively high correlation (r=0.55; p<0.005) was found between linear geographic and genetic distance. This suggests that the populations are partially isolated by distance, despite the limited range (<60 km) of the species. We recommend that population distinetiveness be maintained in restoration efforts.     

Low population genetic differentiation in the Orchidaceae: implications for the diversification of the family

A leading hypothesis for the immense diversity of the Orchidaceae is that skewed mating success and small, disjunct populations lead to strong genetic drift and switches between adaptive peaks. This mechanism is only possible under conditions of low gene flow that lead to high genetic differentiation among populations. We tested whether orchids typically exhibit high levels of population genetic differentiation by conducting a meta‐analysis to compare mean levels of population genetic differentiation (F_ST) between orchids and other diverse families and between rare and common orchids. Compared with other families, the Orchidaceae is typically characterized by relatively low genetic differentiation among populations (mean F_ST = 0.146) at allozyme loci. Rare terrestrial orchids showed higher population genetic differentiation than common orchids, although this value was still lower than the mean for most plant families. All lines of evidence suggest that orchids are typically characterized by low levels of population genetic differentiation, even in species with naturally disjunct populations. As such, we found no strong evidence that genetic drift in isolated populations has played a major role in the diversification of the Orchidaceae. Further research into the diversification of the family needs to unravel the relative roles of biotic and environmental selective pressures in the speciation of orchids.     

Population genetic structure in Myrtus communis L. in a chronically fragmented landscape in the Mediterranean: can gene flow counteract habitat perturbation?

Ancient managed landscapes provide ideal opportunities to assess the consequences of habitat fragmentation on the patterns of genetic diversity and gene flow in long-lived plant species. Using amplified fragment length polymorphism (AFLP) and allozyme markers, we quantified seed-mediated gene flow and population genetic diversity and structure in 14 populations of Myrtus communis (myrtle), a common endozoochorous shrub species of forest patches in lowland agricultural Mediterranean areas. Overall, allozyme diversity for myrtle was low (P₉₅   =   25%; A   =   1.411; H_e = 0.085) compared to other known populations, and a significant portion of populations (57%) had lower levels of allelic diversity and/or heterozygosity than expected at random, as shown by simulated resampling of the whole diversity of the landscape. We found significant correlations between allozyme variability and population size and patch isolation, but no significant inbreeding in any population. Genetic differentiation among populations for both allozyme and AFLP markers was significant (Φ_ST = 0.144 and Φ_ST = 0.142, respectively) but an isolation-by-distance pattern was not detected. Assignment tests on AFLP data indicated a high immigration rate in the populations ( ca. 20–22%), likely through effective seed dispersal across the landscape by birds and mammals. Our results suggest that genetic isolation is not the automatic outcome of habitat destruction since substantial levels of seed-mediated gene flow are currently detectable. However, even moderate rates of gene flow seem insufficient in this long-lived species to counteract the genetic erosion and differentiation imposed by chronic habitat destruction.     

Genetic analysis of ecological relevant morphological variability in Plantago lanceolata L.

Summary Morphological variability was analysed in an F₂-generation derived from crosses between two ecotypes of Plantago lanceolata L. Six allozyme loci, localised in five linkage groups, were used as markers. For two marker loci, Got-2 and Gpi-1, segregations did not fit monogenic ratios. In the linkage groups to which these two loci belonged, male sterility genes appeared to be present. In these crosses, male sterility (type 3, as described by Van Damme 1983) may be determined by two recessive loci located in the linkage groups of Got-2 and of Gpi-1. Many correlations of morphological and life history characters with allozyme markers were observed. The quantitative trait loci did not appear to be concentrated in major gene complexes. Often many loci were involved, sometimes with effects opposite to those expected from the population values. Main effects of the linkage groups appeared to be more important than interaction effects in determining variability. It also appeared that there is a positive correlation between the number of heterozygous allozyme loci and generative growth.Grassland Species Research Group Publication No. 115