Genetic differentiation of neutral markers and quantitative traits in predominantly selfing metapopulations: confronting theory and experiments with Arabidopsis thaliana

The comparison of the genetic differentiation of quantitative traits (QST) and molecular markers (FST) can inform on the strength and spatial heterogeneity of selection in natural populations, provided that markers behave neutrally. However, selection may influence the behaviour of markers in selfing species with strong linkage disequilibria among loci, therefore invalidating this test of detection of selection. We address this issue by monitoring the genetic differentiation of five microsatellite loci (FST) and nine quantitative traits (QST) in experimental metapopulations of the predominantly selfing species Arabidopsis thaliana, that evolved during eight generations. Metapopulations differed with respect to population size and selection heterogeneity. In large populations, the genetic differentiation of neutral microsatellites was much larger under heterogeneous selection than under uniform selection. Using simulations, we show that this influence of selection heterogeneity on FST can be attributable to initial linkage disequilibria among loci, creating stronger genetic differentiation of QTL than expected under a simple additive model with no initial linkage. We found no significant differences between FST and QST regardless of selection heterogeneity, despite a demonstrated effect of selection on QST values. Additional data are required to validate the role of mating system and linkage disequilibria in the joint evolution of neutral and selected genetic differentiation, but our results suggest that FST/QST comparisons can be conservative tests to detect selection in selfing species.     

High quantitative and no molecular differentiation of a freshwater snail (Galba truncatula) between temporary and permanent water habitats

We investigate the variation in quantitative and molecular traits in the freshwater snail Galba truncatula, from permanent and temporary water habitats. Using a common garden experiment, we measured 20 quantitative traits and molecular variation using seven microsatellites in 17 populations belonging to these two habitats. We estimated trait means in each habitat. We also estimated the distributions of overall genetic quantitative variation (QST), and of molecular variation (FST), within and between habitats. Overall, we observed a lack of association between molecular and quantitative variance. Among habitats, we found QST>FST, an indication of selection for different optima. Individuals from temporary water habitat matured older, at a larger size and were less fecund than individuals from permanent water habitat. We discuss these findings in the light of several theories for life-history traits evolution.     

Genetic variability in natural populations of Arabidopsis thaliana in northern Europe

Ten populations of the model plant Arabidopsis thaliana were collected along a north-south gradient in Norway and screened for microsatellite polymorphisms in 25 loci and variability in quantitative traits. Overall, the average levels of genetic diversity were found to be relatively high in these populations, compared to previously published surveys of within population variability. Six of the populations were polymorphic at microsatellite loci, resulting in an overall proportion of polymorphic loci of 18%, and a relatively high gene diversity for a selfing species (HE = 0.06). Of the overall variability, 12% was found within populations. Two of six polymorphic populations contained heterozygous individuals. Both FST and phylogenetic analyses showed no correlation between geographical and genetic distances. Haplotypic diversity patterns suggested postglacial colonization of Scandinavia from a number of different sources. Heritable variation was observed for many of the studied quantitative traits, with all populations showing variability in at least some traits, even populations with no microsatellite variability. There was a positive association between variability in quantitative traits and microsatellites within populations. Several quantitative traits exhibited QST values significantly less than FST, suggesting that selection may be acting to retard differentiation for these traits.     

Evolutionary aspects of population structure for molecular and quantitative traits in the freshwater snail Radix balthica

Detecting the action of selection in natural populations can be achieved using the QST-FST comparison that relies on the estimation of FST with neutral markers, and QST using quantitative traits potentially under selection. QST higher than FST suggests the action of directional selection and thus potential local adaptation. In this article, we apply the QST-FST comparison to four populations of the hermaphroditic freshwater snail Radix balthica located in a floodplain habitat. In contrast to most studies published so far, we did not detect evidence of directional selection for local optima for any of the traits we measured: QST calculated using three different methods was never higher than FST. A strong inbreeding depression was also detected, indicating that outcrossing is probably predominant over selfing in the studied populations. Our results suggest that in this floodplain habitat, local adaptation of R. balthica populations may be hindered by genetic drift, and possibly altered by uneven gene flow linked to flood frequency.     

Low levels of quantitative and molecular genetic differentiation among natural populations of Medicago ciliaris Kroch. (Fabaceae) of different Tunisian eco-geographical origin

In this paper, we analyze the genetic variability in four Tunisian natural populations of Medicago ciliaris using 19 quantitative traits and six polymorphic microsatellite loci. We investigated the amplification transferability of 30 microsatellites developed in the model legume M. truncatula to M. ciliaris. Results revealed that about 56.66% of analyzed markers are valuable genetic markers for M. ciliaris. The most genetic diversity at quantitative traits and microsatellite loci was found to occur within populations (>80%). Low differentiations among populations at quantitative traits Q _ST  = 0.146 and molecular markers F _ST  = 0.18 were found. The majority of measured traits exhibited no significant difference in the level of Q _ST and F _ST . Furthermore, significant correlations established between these traits and eco-geographical factors suggested that natural selection should be invoked to explain the level of phenotypic divergence among populations rather than drift. There was no significant correlation between population differentiation at quantitative traits and molecular markers. Significant spatial genetic structure consistent with models of isolation by distance was detected within all studied populations. The site-of-origin environmental factors explain about 9.07% of total phenotypic genetic variation among populations. The eco-geographical factors that influence more the variation of measured traits among populations are the soil texture and altitude. Nevertheless, there were no consistent pattern of associations between gene diversity (He) and environmental factors.     

Latitudinal divergence of common frog (Rana temporaria) life history traits by natural selection: evidence from a comparison of molecular and quantitative genetic data

The relative roles of natural selection and direct environmental induction, as well as of natural selection and genetic drift, in creating clinal latitudinal variation in quantitative traits have seldom been assessed in vertebrates. To address these issues, we compared molecular and quantitative genetic differentiation between six common frog (Rana temporaria) populations along an approximately 1600 km long latitudinal gradient across Scandinavia. The degree of population differentiation (QST approximately 0.81) in three heritable quantitative traits (age and size at metamorphosis, growth rate) exceeded that in eight (neutral) microsatellite loci (FST = 0.24). Isolation by distance was clear for both neutral markers and quantitative traits, but considerably stronger for one of the three quantitative traits than for neutral markers. QST estimates obtained using animals subjected to different rearing conditions (temperature and food treatments) revealed some environmental dependency in patterns of population divergence in quantitative traits, but in general, these effects were weak in comparison to overall patterns. Pairwise comparisons of FST and QST estimates across populations and treatments revealed that the degree of quantitative trait differentiation was not generally predictable from knowledge of that in molecular markers. In fact, both positive and negative correlations were observed depending on conditions where the quantitative genetic variability had been measured. All in all, the results suggest a very high degree of genetic subdivision both in neutral marker genes and genes coding quantitative traits across a relatively recently (< 9000 years) colonized environmental gradient. In particular, they give evidence for natural selection being the primary agent behind the observed latitudinal differentiation in quantitative traits.     

Morphological and microsatellite diversity associated with ecological factors in natural populations of Medicago laciniata Mill. (Fabaceae)

Genetic variability in 10 natural Tunisian populations of Medicago laciniata were analysed using 19 quantitative traits and 12 polymorphic microsatellite loci. A large degree of genetic variability within-populations and among-populations was detected for both quantitative characters and molecular markers. High genetic differentiation among populations for quantitative traits was seen, with Q _ST = 0.47, and F _ST = 0.47 for microsatellite markers. Several quantitative traits displayed no statistical difference in the levels of Q _ST and F _ST . Further, significant correlations between quantitative traits and eco-geographical factors suggest that divergence in the traits among populations may track environmental differences. There was no significant correlation between genetic variability at quantitative traits and microsatellite markers within populations. The site-of-origin of eco-geographical factors explain between 18.13% and 23.40% of genetic variance among populations at quantitative traits and microsatellite markers, respectively. The environmental factors that most influence variation in measured traits among populations are assimilated phosphorus (P₂0₅) and mean annual rainfall, followed by climate and soil texture, altitude and organic matter. Significant associations between eco-geographical factors and gene diversity, H _e , were established in five-microsatellite loci suggesting that these simple sequence repeats (SSRs) are not necessarily biologically neutral.     

A new method to uncover signatures of divergent and stabilizing selection in quantitative traits

While it is well understood that the pace of evolution depends on the interplay between natural selection, random genetic drift, mutation, and gene flow, it is not always easy to disentangle the relative roles of these factors with data from natural populations. One popular approach to infer whether the observed degree of population differentiation has been influenced by local adaptation is the comparison of neutral marker gene differentiation (as reflected in FST) and quantitative trait divergence (as reflected in QST). However, this method may lead to compromised statistical power, because FST and QST are summary statistics which neglect information on specific pairs of populations, and because current multivariate tests of neutrality involve an averaging procedure over the traits. Further, most FST-QST comparisons actually replace QST by its expectation over the evolutionary process and are thus theoretically flawed. To overcome these caveats, we derived the statistical distribution of population means generated by random genetic drift and used the probability density of this distribution to test whether the observed pattern could be generated by drift alone. We show that our method can differentiate between genetic drift and selection as a cause of population differentiation even in cases with FST=QST and demonstrate with simulated data that it disentangles drift from selection more accurately than conventional FST-QST tests especially when data sets are small.     

Analysis of Genetic Variation in Natural Populations of Medicago truncatula of Southern Tunisian Ecological Areas, Using Morphological Traits and SSR Markers

We used 19 quantitative traits and 14 microsatellite markers (SSRs) to analyze the genetic variation in four natural populations of the model legume Medicago truncatula sampled in southern Tunisia. The greatest genetic variation of quantitative traits and molecular markers occurred within populations (>71%). In contrast to quantitative population differentiation (Q _ST ?=?0.09), a high level of molecular differentiation (F _ST ?=?0.23) was found among populations. The majority of quantitative traits exhibited Q _ST values significantly less than F _ST values, suggesting that selection may be acting to suppress differentiation for these traits. There was no significant correlation between genetic variation of quantitative traits and molecular markers within populations. On the other hand, significant correlations were found between measured quantitative characters and the site-of-origin environmental factors. The eco-geographical factors with the greatest influence on the variation of measured traits among populations were altitude, followed by soil texture, assimilated phosphorus (P₂O₅) and organic matter. Nevertheless, there were no consistent patterns of associations between gene diversity (He) and eco-geographical factors.     

Comparison of rhizobia that nodulate Medicago laciniata and Medicago truncatula present in a single Tunisian arid soil

The rhizobia present in a single arid region Tunisian soil that nodulate Medicago laciniata and Medicago truncatula were compared. All isolates, 40 from each host, were Sinorhizobium meliloti based on 16S rRNA polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) patterns and subsequent confirmation by sequence analysis of the 16S rRNA genes in four representatives from each host species. There was no apparent relationship between Medicago host species of isolation and the nodulating rhizobial genome as determined by repetitive extragenic palandromic PCR. The isolates of M. laciniata were distinguished from those of M. truncatula present in the same soil by variation in PCR-RFLP of nifDK, indicating that this dissimilarity is originally genetic and not geographic. While forming effective symbioses with their own respective isolates, both M. laciniata and M. truncatula formed ineffective true nodules, nodule-like structures, or no nodules at all in cross-inoculation tests, as confirmed by the histological observations.     

Variation for neutral markers is correlated with variation for quantitative traits in the plant pathogenic fungus Mycosphaerella graminicola

We compared genetic variation and population differentiation at RFLP marker loci with seven quantitative characters including fungicide resistance, temperature sensitivity, pycnidial size, pycnidial density, colony size, percentage of leaves covered by pycnidia (PLACP) and percentage of leaves covered by lesions (PLACL) in Mycosphaerella graminicola populations sampled from four regions. Wide variation in population differentiation was found across the quantitative traits assayed. Fungicide resistance, temperature sensitivity, and PLACP displayed a significantly higher Q(ST) than G(ST), consistent with selection for local adaptation, while pycnidial size, pycnidial density and colony size displayed a lower or significantly lower Q(ST) than G(ST), consistent with constraining selection. There was not a statistical difference between Q(ST) and G(ST) in PLACL. We also found a positive and significant correlation between genetic variation in molecular marker loci and quantitative traits at the multitrait scale, suggesting that estimates of overall genetic variation for quantitative traits in M. graminicola could be derived from analysis of the molecular genetic markers.     

Neutral additive genetic variance in a metapopulation

For neutral, additive quantitative characters, the amount of additive genetic variance within and among populations is predictable from Wright's FST, the effective population size and the mutational variance. The structure of quantitative genetic variance in a subdivided metapopulation can be predicted from results from coalescent theory, thereby allowing single-locus results to predict quantitative genetic processes. The expected total amount of additive genetic variance in a metapopulation of diploid individual is given by 2Ne sigma m2 (1 + FST), where FST is Wright's among-population fixation index, Ne is the eigenvalue effective size of the metapopulation, and sigma m2 is the mutational variance. The expected additive genetic variance within populations is given by 2Ne sigma e2(1-FST), and the variance among demes is given by 4FSTNe sigma m2. These results are general with respect to the types of population structure involved. Furthermore, the dimensionless measure of the quantitative genetic variance among populations, QST, is shown to be generally equal to FST for the neutral additive model. Thus, for all population structures, a value of QST greater than FST for neutral loci is evidence for spatially divergent evolution by natural selection.     

Inferring local adaptation from QST-FST comparisons: neutral genetic and quantitative trait variation in European populations of great snipe

We applied a phenotypic QST (PST) vs. FST approach to study spatial variation in selection among great snipe (Gallinago media) populations in two regions of northern Europe. Morphological divergence between regions was high despite low differentiation in selectively neutral genetic markers, whereas populations within regions showed very little neutral divergence and trait differentiation. QST > FST was robust against altering assumptions about the additive genetic proportions of variance components. The homogenizing effect of gene flow (or a short time available for neutral divergence) has apparently been effectively counterbalanced by differential natural selection, although one trait showed some evidence of being under uniform stabilizing selection. Neutral markers can hence be misleading for identifying evolutionary significant units, and adopting the PST-FST approach might therefore be valuable when common garden experiments is not an option. We discuss the statistical difficulties of documenting uniform selection as opposed to divergent selection, and the need for estimating measurement error. Instead of only comparing overall QST and FST values, we advocate the use of partial matrix permutation tests to analyse pairwise QST differences among populations, while statistically controlling for neutral differentiation.     

Cross-species amplification of 92 microsatellites of Medicago truncatula

Medicago species are important genetic sources for forage crops and nitrogen sources for various ecosystems. The ongoing genome sequencing of the model legume, Medicago truncatula, provides a wealth of genetic markers potentially useful for characterizing the population genetic structure and evolutionary history, and the potential of the wild Medicago species. Here we tested the PCR amplification of 92 microsatellites developed from M. truncatula in six other Medicago species, and found that the cross-species transferability, ranging from 53.26% to 61.96%, is comparable with those reported in other angiosperm genera. This article thus reports a number of microsatellites that are potentially useful for large-scale ecological and evolutionary genetic studies of wild Medicago species.     

Genetic Markers and Quantitative Genetic Variation in Medicago Truncatula (Leguminosae): A Comparative Analysis of Population Structure

Two populations of the selfing annual Medicago truncatula Gaertn. (Leguminoseae), each subdivided into three subpopulations, were studied for both metric traits (quantitative characters) and genetic markers (random amplified polymorphic DNA and one morphological, single-locus marker). Hierarchical analyses of variance components show that (1) populations are more differentiated for quantitative characters than for marker loci, (2) the contribution of both within and among subpopulations components of variance to overall genetic variance of these characters is reduced as compared to markers, and (3) at the population level, within population structure is slightly but not significantly larger for markers than for quantitative traits. Under the hypothesis that most markers are neutral, such comparisons may be used to make hypotheses about the strength and heterogeneity of natural selection in the face of genetic drift and gene flow. We thus suggest that in these populations, quantitative characters are under strong divergent selection among populations, and that gene flow is restricted among populations and subpopulations.     

PATTERNS OF GENETIC ARCHITECTURE FOR LIFE-HISTORY TRAITS AND MOLECULAR MARKERS IN A SUBDIVIDED SPECIES

Abstract Understanding the utility and limitations of molecular markers for predicting the evolutionary potential of natural populations is important for both evolutionary and conservation genetics. To address this issue, the distribution of genetic variation for quantitative traits and molecular markers is estimated within and among 14 permanent lake populations of Daphnia pulicaria representing two regional groups from Oregon. Estimates of population subdivision for molecular and quantitative traits are concordant, with Q _ST generally exceeding G _ST. There is no evidence that microsatellites loci are less informative about subdivision for quantitative traits than are allozyme loci. Character-specific comparison of Q _ST and G _ST support divergent selection pressures among populations for the majority of life-history traits in both coast and mountain regions. The level of within-population variation for molecular markers is uninformative as to the genetic variation maintained for quantitative traits. In D. pulicaria , regional differences in the frequency of sex may contribute to variation in the maintenance of expressed within-population quantitative-genetic variation without substantially impacting diversity at the genic level. These data are compared to an identical dataset for 17 populations of the temporary-pond species, D. pulex .     

Does natural selection promote population divergence? A comparative analysis of population structure using amplified fragment length polymorphism markers and quantitative traits

Divergent natural selection is considered an important force in plant evolution leading to phenotypic differentiation between populations exploiting different environments. Extending an earlier greenhouse study of population differentiation in the selfing annual plant Senecio vulgaris, we estimated the degree of population divergence in several quantitative traits related to growth and life history and compared these estimates with those based on presumably neutral molecular markers (amplified fragment length polymorphisms; AFLPs). This approach allowed us to disentangle the effects of divergent selection from that of other evolutionary forces (e.g. genetic drift). Five populations were examined from each of two habitat types (ruderal and agricultural habitats). We found a high proportion of total genetic variance to be among populations, both for AFLP markers (phiST = 0.49) and for quantitative traits (range of QST: 0.26-0.77). There was a strong correlation between molecular and quantitative genetic differentiation between pairs of populations (Mantel's r = 0.59). However, estimates of population differentiation in several quantitative traits exceeded the neutral expectation (estimated from AFLP data), suggesting that divergent selection contributed to phenotypic differentiation, especially between populations from ruderal and agricultural habitats. Estimates of within-population variation in AFLP markers and quantitative genetic were poorly correlated, indicating that molecular marker data may be of limited value to predict the evolutionary potential of populations of S. vulgaris.     

Contrasting patterns of divergence in quantitative traits and neutral DNA markers: analysis of clinal variation

Clinal variation in quantitative traits is often attributed to the effects of spatially varying selection. However, identical patterns can be produced by the interplay between purely stochastic processes (i.e. drift in combination with spatially restricted gene flow). One means of distinguishing between adaptive and nonadaptive causes of geographical variation is to compare relative levels of between-population divergence in quantitative traits and neutral DNA markers. Such comparisons can be used to test whether levels of trait divergence attributable to additive genetic effects (as measured by QST) exceed null expectations based on the level of divergence at neutral marker loci (as measured by FST). The purpose of this study was to use an approach based on 'QST vs. FST' contrasts to test for evidence of diversifying selection on body size of an Indian fruit bat, Cynopterus sphinx (Chiroptera: Pteropodidae). Specifically, relative levels of between-population divergence in body size and microsatellite DNA markers were compared to assess whether the observed pattern of clinal size variation could be explained by a neutral model of isolation by distance. QST for body size was calculated using unbiased estimators of within- and between-population variance of principal component scores. The association between body size variation and geographical/environmental distance was tested using pairwise and partial matrix correspondence tests (MCTs). Independent variables (representing causal hypotheses) were constructed as between-locality distance matrices. The effects of neutral genetic divergence were assessed by including a matrix of pairwise FST as an independent variable. Partial MCTs revealed highly significant associations between phenotypic divergence (QST) and both geographical and environmental distance, even when the effects of neutral genetic divergence (FST) were partialled out. Results of the tests confirmed that migration-drift equilibrium is not a sufficient explanation for the latitudinal pattern of clinal size variation in C. sphinx. The geographical patterning of pairwise QST is most likely attributable to spatially varying selection and/or the direct influence of latitudinally ordered environmental effects.     

Local adaptation in brown trout early life-history traits: implications for climate change adaptability

Knowledge of local adaptation and adaptive potential of natural populations is becoming increasingly relevant due to anthropogenic changes in the environment, such as climate change. The concern is that populations will be negatively affected by increasing temperatures without the capacity to adapt. Temperature-related adaptability in traits related to phenology and early life history are expected to be particularly important in salmonid fishes. We focused on the latter and investigated whether four populations of brown trout (Salmo trutta) are locally adapted in early life-history traits. These populations spawn in rivers that experience different temperature conditions during the time of incubation of eggs and embryos. They were reared in a common-garden experiment at three different temperatures. Quantitative genetic differentiation (QST) exceeded neutral molecular differentiation (FST) for two traits, indicating local adaptation. A temperature effect was observed for three traits. However, this effect varied among populations due to locally adapted reaction norms, corresponding to the temperature regimes experienced by the populations in their native environments. Additive genetic variance and heritable variation in phenotypic plasticity suggest that although increasing temperatures are likely to affect some populations negatively, they may have the potential to adapt to changing temperature regimes.     

DOES SELECTION ON FLORAL ODOR PROMOTE DIFFERENTIATION AMONG POPULATIONS AND SPECIES OF THE SEXUALLY DECEPTIVE ORCHID GENUS OPHRYS?

Sexually deceptive orchids from the genus Ophrys attract their pollinators primarily through the chemical mimicry of female hymenopteran sex pheromones, thereby deceiving males into attempted matings with the orchid labellum. Floral odor traits are crucial for the reproductive success of these pollinator-limited orchids, as well as for maintaining reproductive isolation through the attraction of specific pollinators. We tested for the signature of pollinator-mediated selection on floral odor by comparing intra and interspecific differentiation in odor compounds with that found at microsatellite markers among natural populations. Three regions from southern Italy were sampled. We found strong floral odor differentiation among allopatric populations within species, among allopatric species and among sympatric species. Population differences in odor were also reflected in significant variation in the attractivity of floral extracts to the pollinator, Colletes cunicularius. Odor compounds that are electrophysiologically active in C. cunicularius males, especially alkenes, were more strongly differentiated among conspecific populations than nonactive compounds in the floral odor. In marked contrast to these odor patterns, there was limited population or species level differentiation in microsatellites (FST range 0.005 to 0.127, mean FST 0.075). We propose that the strong odor differentiation and lack of genetic differentiation among sympatric taxa indicates selection imposed by the distinct odor preferences of different pollinating species. Within species, low FST values are suggestive of large effective population sizes and indicate that divergent selection rather than genetic drift accounts for the strong population differentiation in odor. The higher differentiation in active versus non-active odor compounds suggests that divergent selection among orchid populations may be driven by local pollinator preferences for those particular compounds critical for pollinator attraction.