Scale‐dependent thermal tolerance variation in Australian mountain grasshoppers

Physiological variation among and within species is thought to play a key role in determining distribution patterns across environmental gradients. We tested inter‐ and intraspecific variation in cold and heat tolerances for three grasshopper species (genus Kosciuscola) with overlapping elevation distributions, across their respective ranges in the Australian mountains. Of the three cold tolerance traits measured, the critical thermal minimum was the only trait to vary among species, with greater cold tolerance associated with a distribution extending to a higher elevation. Cold tolerance limits were regularly exceeded in exposed microhabitats, suggesting a role for cold adaptation in structuring species distribution patterns. In contrast to cold tolerance, heat tolerance variation was primarily partitioned within species. For two species, populations from treeless alpine habitat were more heat tolerant than their lower‐elevation counterparts, supporting recent models that suggest greater exposure to temperature extremes at higher elevations. These contrasting patterns of physiological variation among and within species emphasise the importance of considering variation within species when attempting to understand how species distributions are affected by thermal extremes.     

Intraspecific variation in thermal acclimation and tolerance between populations of the winter ant,Prenolepis imparis

Thermal phenotypic plasticity, otherwise known as acclimation, plays an essential role in how organisms respond to short‐term temperature changes. Plasticity buffers the impact of harmful temperature changes; therefore, understanding variation in plasticity in natural populations is crucial for understanding how species will respond to the changing climate. However, very few studies have examined patterns of phenotypic plasticity among populations, especially among ant populations. Considering that this intraspecies variation can provide insight into adaptive variation in populations, the goal of this study was to quantify the short‐term acclimation ability and thermal tolerance of several populations of the winter ant, Prenolepis imparis. We tested for correlations between thermal plasticity and thermal tolerance, elevation, and body size. We characterized the thermal environment both above and below ground for several populations distributed across different elevations within California, USA. In addition, we measured the short‐term acclimation ability and thermal tolerance of those populations. To measure thermal tolerance, we used chill‐coma recovery time (CCRT) and knockdown time as indicators of cold and heat tolerance, respectively. Short‐term phenotypic plasticity was assessed by calculating acclimation capacity using CCRT and knockdown time after exposure to both high and low temperatures. We found that several populations displayed different chill‐coma recovery times and a few displayed different heat knockdown times, and that the acclimation capacities of cold and heat tolerance differed among most populations. The high‐elevation populations displayed increased tolerance to the cold (faster CCRT) and greater plasticity. For high‐temperature tolerance, we found heat tolerance was not associated with altitude; instead, greater tolerance to the heat was correlated with increased plasticity at higher temperatures. These current findings provide insight into thermal adaptation and factors that contribute to phenotypic diversity by revealing physiological variance among populations.     

Acorn preference under field and laboratory conditions by two flightless Iberian dung beetle species (Thorectes baraudi and Jekelius nitidus): implications for recruitment and management of oak forests in central Spain

1. Recently, a mutualistic relationship has been described between some dung beetles (Thorectes lusitanicus and Mycotrupes lethroides) and oak species (Quercus suber, Q. canariensis, and Q. rubra), which could be crucial for ensuring seedling recruitment and sustaining the equilibrium of oak populations. For T. lusitanicus, a diet based on acorns during the reproductive period improved resistance to low‐temperature conditions and improved ovarian development. 2. In this paper, we conducted field and laboratory experiments to investigate the interaction between two potential acorn‐eating beetles, Thorectes baraudi and Jekelius nitidus, with Quercus suber. We determined the feeding preferences of both beetle species and estimated the rates of acorn manipulation by beetles according to habitat structure and several characteristics of the acorn, such as seed size and acorn infestation by weevils. 3. Results demonstrated the positive interaction between the dung beetle Thorectes baraudi and Quercus trees. Thorectes baraudi was clearly more attracted to volatiles of acorns than to dung. Jekelius nitidus, on the contrary, was either not or anecdotally attracted to acorns. On the contrary, in the case of Jekelius nitidus, the acorn attraction could be considered anecdotal or even accidental. Our field results demonstrated the acorn burying behaviour of T. baraudi in the oak forests of the Cabañeros National Park (Spain), suggesting a potential role of this beetle species as an active secondary acorn disperser. 4. This unexpected behaviour could be particularly important in Mediterranean oak forests and savannahs, where most Quercus species are strongly recruitment limited because of serious overgrazing problems.     

Temperature and photoperiod affect stress resistance traits in Drosophila melanogaster

The long‐term survival of species and populations depends on their ability to adjust phenotypic values to environmental conditions. In particular, the capability of dealing with environmental stress to buffer detrimental effects on fitness is considered to be of pivotal importance. Resistance traits are readily modulated by a wide range of environmental factors. In the present study, Drosophila melanogaster Meigen is used to investigate plastic responses to temperature and photoperiod in stress resistance traits. The results reveal that stress resistance traits (cold, heat, starvation and desiccation resistance) are affected by the factors temperature and sex predominantly. Cooler temperatures compared with warmer temperatures increase cold tolerance, desiccation and starvation resistance, whereas they reduce heat tolerance. Except for heat resistance, females are more stress‐resistant than males. Stress resistance traits are also affected by photoperiod. Shorter photoperiods decrease cold tolerance, whereas longer photoperiods enhance desiccation resistance. Overall, thermal effects are pervasive throughout all measured resistance traits, whereas photoperiodic effects are of limited importance in the directly developing (i.e. nondiapausing) flies used here, suggesting that pronounced photoperiodic effects on stress resistance traits may be largely limited to, and triggered by, diapause‐inducing effects.     

Surviving the heat: heterogeneity of response in Saccharomyces cerevisiae provides insight into thermal damage to the membrane

Environmental heat stress impacts on the physiology and viability of microbial cells with concomitant implications for microbial activity and diversity. Previously, it has been demonstrated that gradual heating of Saccharomyces cerevisiae induces a degree of thermal resistance, whereas a heat shock results in a high level of cell death. Here, we show that the impact of exogenous nutrients on acquisition of thermal resistance differs between strains. Using single‐cell methods, we demonstrate the extent of heterogeneity of the heat‐stress response within populations of yeast cells and the presence of subpopulations that are reversibly damaged by heat stress. Such cells represent potential for recovery of entire populations once stresses are removed. The results show that plasma membrane permeability and potential are key factors involved in cell survival, but thermal resistance is not related to homeoviscous adaptation of the plasma membrane. These results have implications for growth and regrowth of populations experiencing environmental heat stress and our understanding of impacts at the level of the single cell. Given the important role of microbes in biofuel production and bioremediation, a thorough understanding of the impact of stress responses of populations and individuals is highly desirable.     

Local coexistence and niche differences between the Lusitanian and Mediterranean pine voles (<Emphasis Type="Italic">Microtus lusitanicus</Emphasis> and <Emphasis Type="Italic">M. duodecimcostatus</Emphasis>)

In the present study, we analyzed the coexistence pattern of the Lusitanian pine vole (Microtus lusitanicus) and the Mediterranean pine vole (Microtus duodecimcostatus) in a potential area of sympatry in a Mediterranean landscape (Portugal). We also determined the relative contribution of local, landscape, and spatial factors explaining the differences in the distribution patterns of the two species in the region. Using a kriging interpolation method, we obtained a map of sympatric and allopatric areas of species occurrence. The estimated sympatry area corresponded to a northwest–southeast belt representing 11.3% of the study area. Habitat niche differences were assessed with binomial GLMs followed by a variance partitioning. At a local scale, higher altitude, higher cover of shrubs, lower clay content in the soil, and lower cover of tree canopy were the most important factors distinguishing M. lusitanicus presence sites from those with M. duodecimcostatus. At a larger scale, the presence of forest landscape units and the low abundance of “montado” units were the most influencing landscape factors in the identification of M. lusitanicus occurrence sites when compared to M. duodecimcostatus. Our results suggested that local coexistence of M. lusitanicus and M. duodecimcostatus in the field is a rare event. The differences in distribution patterns of the two pine vole species were mostly explained by fine-scale environmental factors and by shared spatial effects.     

The influence of immune activation on thermal tolerance along a latitudinal cline

Global change is shifting both temperature patterns and the geographic distribution of pathogens, and infection has already been shown to substantially reduce host thermal performance, potentially placing populations at greater risk that previously thought. But what about individuals that are able to successfully clear an infection? Whilst the direct damage a pathogen causes will likely lead to reductions in host's thermal tolerance, the response to infection often shares many underlying pathways with the general stress response, potentially acting as a buffer against subsequent thermal stress. Here, by exposing Drosophila melanogaster to heat‐killed bacterial pathogens, we investigate how activation of a host's immune system can modify any response to both heat and cold temperature stress. In a single focal population, we find that immune activation can improve a host's knockdown times during heat shock, potentially offsetting some of the damage that would subsequently arise as an infection progresses. Conversely, immune activation had a detrimental effect on CT_max and did not influence lower thermal tolerance as measured by chill‐coma recovery time. However, we also find that the influence of immune activation on heat knockdown times is not generalizable across an entire cline of locally adapted populations. Instead, immune activation led to signals of local adaptation to temperature being lost, erasing the previous advantage that populations in warmer regions had when challenged with heat stress. Our results suggest that activation of the immune system may help buffer individuals against the detrimental impact of infection on thermal tolerance; however, any response will be population specific and potentially not easily predicted across larger geographic scales, and dependent on the form of thermal stress faced by a host.     

Keeping your options open: Maintenance of thermal plasticity during adaptation to a stable environment

Phenotypic plasticity may allow species to cope with environmental variation. The study of thermal plasticity and its evolution helps understanding how populations respond to variation in temperature. In the context of climate change, it is essential to realize the impact of historical differences in the ability of populations to exhibit a plastic response to thermal variation and how it evolves during colonization of new environments. We have analyzed the real‐time evolution of thermal reaction norms of adult and juvenile traits in Drosophila subobscura populations from three locations of Europe in the laboratory. These populations were kept at a constant temperature of 18ºC, and were periodically assayed at three experimental temperatures (13ºC, 18ºC, and 23ºC). We found initial differentiation between populations in thermal plasticity as well as evolutionary convergence in the shape of reaction norms for some adult traits, but not for any of the juvenile traits. Contrary to theoretical expectations, an overall better performance of high latitude populations across temperatures in early generations was observed. Our study shows that the evolution of thermal plasticity is trait specific, and that a new stable environment did not limit the ability of populations to cope with environmental challenges.     

Mechanisms of thermal adaptation and evolutionary potential of conspecific populations to changing environments

Heterogeneous and ever‐changing thermal environments drive the evolution of populations and species, especially when extreme conditions increase selection pressure for traits influencing fitness. However, projections of biological diversity under scenarios of climate change rarely consider evolutionary adaptive potential of natural species. In this study, we tested for mechanistic evidence of evolutionary thermal adaptation among ecologically divergent redband trout populations (Oncorhynchus mykiss gairdneri) in cardiorespiratory function, cellular response and genomic variation. In a common garden environment, fish from an extreme desert climate had significantly higher critical thermal maximum (p < .05) and broader optimum thermal window for aerobic scope (>3°C) than fish from cooler montane climate. In addition, the desert population had the highest maximum heart rate during warming (20% greater than montane populations), indicating improved capacity to deliver oxygen to internal tissues. In response to acute heat stress, distinct sets of cardiac genes were induced among ecotypes, which helps to explain the differences in cardiorespiratory function. Candidate genomic markers and genes underlying these physiological adaptations were also pinpointed, such as genes involved in stress response and metabolic activity (hsp40, ldh‐b and camkk2). These markers were developed into a multivariate model that not only accurately predicted critical thermal maxima, but also evolutionary limit of thermal adaptation in these specific redband trout populations relative to the expected limit for the species. This study demonstrates mechanisms and limitations of an aquatic species to evolve under changing environments that can be incorporated into advanced models to predict ecological consequences of climate change for natural organisms.     

Culturable aerobic and facultative bacteria from the gut of the polyphagic dung beetle Thorectes lusitanicus

Unlike other dung beetles, the Iberian geotrupid, Thorectes lusitanicus, exhibits polyphagous behavior; for example, it is able to eat acorns, fungi, fruits, and carrion in addition to the dung of different mammals. This adaptation to digest a wider diet has physiological and developmental advantages and requires key changes in the composition and diversity of the beetle's gut microbiota. In this study, we isolated aerobic, facultative anaerobic, and aerotolerant microbiota amenable to grow in culture from the gut contents of T. lusitanicus and resolved isolate identity to the species level by sequencing 16S rRNA gene fragments. Using BLAST similarity searches and maximum likelihood phylogenetic analyses, we were able to reveal that the analyzed fraction (culturable, aerobic, facultative anaerobic, and aerotolerant) of beetle gut microbiota is dominated by the phyla Proteobacteria, Firmicutes, and Actinobacteria. Among Proteobacteria, members of the order Enterobacteriales (Gammaproteobacteria) were the most abundant. The main functions associated with the bacteria found in the gut of T. lusitanicus would likely include nitrogen fixation, denitrification, detoxification, and diverse defensive roles against pathogens.     

Heat stress responses and population genetics of the kelp Laminaria digitata (Phaeophyceae) across latitudes reveal differentiation among North Atlantic populations

To understand the thermal plasticity of a coastal foundation species across its latitudinal distribution, we assess physiological responses to high temperature stress in the kelp Laminaria digitata in combination with population genetic characteristics and relate heat resilience to genetic features and phylogeography. We hypothesize that populations from Arctic and cold‐temperate locations are less heat resilient than populations from warm distributional edges. Using meristems of natural L. digitata populations from six locations ranging between Kongsfjorden, Spitsbergen (79°N), and Quiberon, France (47°N), we performed a common‐garden heat stress experiment applying 15°C to 23°C over eight days. We assessed growth, photosynthetic quantum yield, carbon and nitrogen storage, and xanthophyll pigment contents as response traits. Population connectivity and genetic diversity were analyzed with microsatellite markers. Results from the heat stress experiment suggest that the upper temperature limit of L. digitata is nearly identical across its distribution range, but subtle differences in growth and stress responses were revealed for three populations from the species’ ecological range margins. Two populations at the species’ warm distribution limit showed higher temperature tolerance compared to other populations in growth at 19°C and recovery from 21°C (Quiberon, France), and photosynthetic quantum yield and xanthophyll pigment responses at 23°C (Helgoland, Germany). In L. digitata from the northernmost population (Spitsbergen, Norway), quantum yield indicated the highest heat sensitivity. Microsatellite genotyping revealed all sampled populations to be genetically distinct, with a strong hierarchical structure between southern and northern clades. Genetic diversity was lowest in the isolated population of the North Sea island of Helgoland and highest in Roscoff in the English Channel. All together, these results support the hypothesis of moderate local differentiation across L. digitata's European distribution, whereas effects are likely too weak to ameliorate the species’ capacity to withstand ocean warming and marine heatwaves at the southern range edge.     

Thermal stress exposure of pupal oriental fruit fly has strong and trait-specific consequences in adult flies

Global climate change is projected to increase the incidence of heat waves, their magnitude and duration resulting in insects experiencing increasing environmental stress in both natural and managed ecosystems. While studies on insect thermal tolerance are rapidly increasing, variation across developmental or juvenile stress cross-stage effects within and across generations remain largely unexplored. Yet in holometabolous insects, heat stress at an early developmental stage may influence performance and survival during later stages. Here, we investigated the effects of pupal mild heat stress on the performance of laboratory-reared adult Bactrocera dorsalis (Hendel) (Diptera: Tephritidae) measured as longevity, critical thermal maximum (CT_max), critical thermal minima (CT_min), heat knockdown time (HKDT) and chill coma recovery time (CCRT). Pupal heat stress significantly influenced performance of B. dorsalis adults resulting in impaired longevity and heat tolerance (CT_max and HKDT) in both sexes with improved and compromised cold tolerance (CT_min and CCRT) in females and males, respectively. These findings highlight the role of juvenile stages in mediating stress responses at adult stages. For B. dorsalis, pupal heat stress largely compromised thermal tolerance implying that the species has limited potential to shift its geographic range in heat prone areas. Significant benefits in cold tolerance in females following heat stress may help in improving survival in the cold in the short-term despite restricted activity to the same traits in males. This study suggests that basal heat tolerance and not short-term compensatory thermal plasticity following heat stress may have aided the recent invasion of B. dorsalis in African landscapes.     

Weak functional response to agricultural landscape homogenisation among plants,butterflies and birds

Measures of functional diversity are expected to predict community responses to land use and environmental change because, in contrast to taxonomic diversity, it is based on species traits rather than their identity. Here, we investigated the impact of landscape homogenisation on plants, butterflies and birds in terms of the proportion of arable field cover in southern Finland at local (0.25 km²) and regional (> 10 000 km²) scales using four functional diversity indices: functional richness, functional evenness, functional divergence and functional dispersion. No uniform response in functional diversity across taxa or scales was found. However, in all cases where we found a relationship between increasing arable field cover and any index of functional diversity, this relationship was negative. Butterfly functional richness decreased with increasing arable field cover, as did butterfly and bird functional evenness. For butterfly functional evenness, this was only evident in the most homogeneous regions. Butterfly and bird functional dispersion decreased in homogeneous regions regardless of the proportion of arable field cover locally. No effect of landscape heterogeneity on plant functional diversity was found at any spatial scale, but plant species richness decreased locally with increasing arable field cover. Overall, species richness responded more consistently to landscape homogenisation than did the functional diversity indices, with both positive and negative effects across species groups. Functional diversity indices are in theory valuable instruments for assessing effects of land use scenarios on ecosystem functioning. However, the applicability of empirical data requires deeper understanding of which traits reliably capture species’ vulnerability to environmental factors and of the ecological interpretation of the functional diversity indices. Our study provides novel insights into how the functional diversity of communities changes in response to agriculturally derived landscape homogenisation; however, the low explanatory power of the functional diversity indices hampers the ability to reliably anticipate impacts on ecosystem functioning.     

Parallel trait adaptation across opposing thermal environments in experimental Drosophila melanogaster populations

Thermal stress is a pervasive selective agent in natural populations that impacts organismal growth, survival, and reproduction. Drosophila melanogaster exhibits a variety of putatively adaptive phenotypic responses to thermal stress in natural and experimental settings; however, accompanying assessments of fitness are typically lacking. Here, we quantify changes in fitness and known thermal tolerance traits in replicated experimental D. melanogaster populations following more than 40 generations of evolution to either cyclic cold or hot temperatures. By evaluating fitness for both evolved populations alongside a reconstituted starting population, we show that the evolved populations were the best adapted within their respective thermal environments. More strikingly, the evolved populations exhibited increased fitness in both environments and improved resistance to both acute heat and cold stress. This unexpected parallel response appeared to be an adaptation to the rapid temperature changes that drove the cycling thermal regimes, as parallel fitness changes were not observed when tested in a constant thermal environment. Our results add to a small, but growing group of studies that demonstrate the importance of fluctuating temperature changes for thermal adaptation and highlight the need for additional work in this area.     

Altitudinal variation for stress resistance traits and thermal adaptation in adult Drosophila buzzatii from the New World

Multiple stress resistance traits were investigated in the cactophilic fly Drosophila buzzatii. Adults from seven populations derived from North-Western Argentina were compared with respect to traits relevant for thermal stress resistance and for resistance to other forms of environmental stress. The populations were collected along an altitudinal gradient spanning more than 2000 m in height, showing large climatic differences. The results suggest that knock-down resistance to heat stress, desiccation resistance and Hsp70 expression at a relatively severe stressful temperature best reflect thermal adaptation in this species. Furthermore, cold resistance seemed to be of less importance than heat resistance, at least for the adult life stage, in these populations. Clinal variation in thermal resistance traits over short geographical distances suggests relatively strong adaptive differentiation of the populations. This study provides the first evidence for altitudinal differentiation in stress-related traits, and suggests that Hsp70 expression level can be related to altitudinal clines of heat-stress resistance.     

Differential effects of short term winter thermal stress on diapausing tiger swallowtail butterflies (Papilio spp.)

Abstract It is generally thought that insects inhabiting lower latitudes are more severely impacted by changes in their thermal environment than are high latitude species. This is attributed to the wider range of temperatures to which high‐latitude species are exposed. By contrast, low‐latitude species have typically evolved in more thermally stable environments with a narrower range of temperature variation. However, deviation from this pattern can occur and here we report that under variable winter conditions a higher latitude species may be more sensitive to thermal variation than its lower latitude sister species. Using split broods, we examined the survival and adult emergence success of diapausing pupae of Papilio canadensis and P. glaucus, as well as a unique, recombinant hybrid population (“late‐flight”) to short periods of mid‐winter cold and heat stress. Our results indicate that the higher latitude, univoltine populations (P. canadensis and late‐flights) exhibit lower pupal survival than the lower latitude, facultative diapauser (P. glaucus) for all mid‐winter thermal stress treatments, both high and low. Size differences alone do not appear to account for the observed differences in survival or metabolic costs in these three phenotypes, as late‐flight individuals are similar in size to P. glaucus. We attribute the observed differences in survival and weight loss to potential metabolic differences and variation in the intensity of diapause, in addition to divergent adaptation to winter precipitation levels (e.g. snow cover) and the influences this may have on microhabitat temperature moderation.     

Laboratory maintenance does not alter ecological and physiological patterns among species: a Drosophila case study

Large comparative studies in animal ecology, physiology and evolution often use animals reared in the laboratory for many generations; however, the relevance of these studies hinges on the assumption that laboratory populations are still representative for their wild living conspecifics. In this study, we investigate whether laboratory‐maintained and freshly collected animal populations are fundamentally different and whether data from laboratory‐maintained animals are valid to use in large comparative investigations of ecological and physiological patterns. Here, we obtained nine species of Drosophila with paired populations of laboratory‐maintained and freshly collected flies. These species, representing a range of ecotypes, were assayed for four stress‐tolerance, two body‐size traits and six life‐history traits. For all of these traits, we observed small differences in species‐specific comparisons between field and laboratory populations; however, these differences were unsystematic and laboratory maintenance did not eclipse fundamental species characteristics. To investigate whether laboratory maintenance influence the general patterns in comparative studies, we correlated stress tolerance and life‐history traits with environmental traits for the laboratory‐maintained and freshly collected populations. Based on this analysis, we found that the comparative physiological and ecological trait correlations are similar irrespective of provenience. This finding is important for comparative biology in general because it validates comparative meta‐analyses based on laboratory‐maintained populations.     

Low genetic diversity contrasts with high phenotypic variability in heptaploid Spartina densiflora populations invading the Pacific coast of North America

Species can respond to environmental pressures through genetic and epigenetic changes and through phenotypic plasticity, but few studies have evaluated the relationships between genetic differentiation and phenotypic plasticity of plant species along changing environmental conditions throughout wide latitudinal ranges. We studied inter‐ and intrapopulation genetic diversity (using simple sequence repeats and chloroplast DNA sequencing) and inter‐ and intrapopulation phenotypic variability of 33 plant traits (using field and common‐garden measurements) for five populations of the invasive cordgrass Spartina densiflora Brongn. along the Pacific coast of North America from San Francisco Bay to Vancouver Island. Studied populations showed very low genetic diversity, high levels of phenotypic variability when growing in contrasted environments and high intrapopulation phenotypic variability for many plant traits. This intrapopulation phenotypic variability was especially high, irrespective of environmental conditions, for those traits showing also high phenotypic plasticity. Within‐population variation represented 84% of the total genetic variation coinciding with certain individual plants keeping consistent responses for three plant traits (chlorophyll b and carotenoid contents, and dead shoot biomass) in the field and in common‐garden conditions. These populations have most likely undergone genetic bottleneck since their introduction from South America; multiple introductions are unknown but possible as the population from Vancouver Island was the most recent and one of the most genetically diverse. S. densiflora appears as a species that would not be very affected itself by climate change and sea‐level rise as it can disperse, establish, and acclimate to contrasted environments along wide latitudinal ranges.     

Diversity and Frequency of Clonal Traits Along Natural and Land-Use Gradients in Grasslands of the Swiss Alps

The frequency of clonal plants in different vegetation types is known to be influenced by environmental and land-use factors. However, the underlying behavior of individual clonal traits or clonal trait diversity has received little attention. Here, we assess for species- and trait-diverse grasslands of the Swiss Alps the relative importance of temperature, soil moisture, land use and species richness on the diversity and frequency of individual compared with all clonal traits. We further analyzed how cover-weighted data alters the relationships found with commonly used presence-absence data. We combined species compositional, land-use and environmental data from 236 28-m² grassland plots with clonal trait information for 527 species following the Clonal Growth Organ (CGO) classification. Test results are based on linear models, ANOVAs and ANCOVAs. The grassland sites were 84% dominated by clonal species. Drought-prone grasslands harbored the least clonal species. No increase in clonality was detected with decreasing temperature (= altitude). Mown or pastured grasslands had more clonal species than fallows. Certain sets of traits were correlated. Rhizomatous species especially reacted strongly to climatic and land-use gradients and had highest frequencies in cold, moist and disturbed sites. Clonal diversity was strongly dependent on species richness. Cover-weighted and presence-absence based estimates were largely similar. Overall, our data outlined that common clonal traits react differently to natural and land-use gradients as well as differently to the sum of clonal traits. Also, soil moisture was more decisive than temperature (= altitude) for the presence of clonal species. Lastly, the strong correlation between species-richness and clonal trait diversity needs to be accounted for when interpreting the functional role of clonal traits.