Quantitative size-resolved characterization of mRNA nanoparticles by in-line coupling of asymmetrical-flow field-flow fractionation with small angle X-ray scattering.

We present a generically applicable approach to determine an extensive set of size-dependent critical quality attributes inside nanoparticulate pharmaceutical products. By coupling asymmetrical-flow field-flow fractionation (AF4) measurements directly in-line with solution small angle X-ray scattering (SAXS), vital information such as (i) quantitative, absolute size distribution profiles, (ii) drug loading, (iii) size-dependent internal structures, and (iv) quantitative information on free drug is obtained. Here the validity of the method was demonstrated by characterizing complex mRNA-based lipid nanoparticle products. The approach is particularly applicable to particles in the size range of 100 nm and below, which is highly relevant for pharmaceutical products-both biologics and nanoparticles. The method can be applied as well in other fields, including structural biology and environmental sciences.

Eco-evolutionary dynamics modulate plant responses to global change depending on plant diversity and species identity.

Global change has dramatic impacts on grassland diversity. However, little is known about how fast species can adapt to diversity loss and how this affects their responses to global change. Here, we performed a common garden experiment testing whether plant responses to global change are influenced by their selection history and the conditioning history of soil at different plant diversity levels. Using seeds of four grass species and soil samples from a 14-year-old biodiversity experiment, we grew the offspring of the plants either in their own soil or in soil of a different community, and exposed them either to drought, increased nitrogen input, or a combination of both. Under nitrogen addition, offspring of plants selected at high diversity produced more biomass than those selected at low diversity, while drought neutralized differences in biomass production. Moreover, under the influence of global change drivers, soil history, and to a lesser extent plant history, had species-specific effects on trait expression. Our results show that plant diversity modulates plant-soil interactions and growth strategies of plants, which in turn affects plant eco-evolutionary pathways. How this change affects species' response to global change and whether this can cause a feedback loop should be investigated in more detail in future studies.

Over the last hundred years, human activities including burning of fossil fuels, clearing of forests, and fertilizer use have caused environmental changes that have resulted in many species of plants, animals and other forms of life becoming extinct. Loss of plant species can change the local environment by, for example, altering the availability of nutrients and local communities of microbes in the soil. This may, in turn, cause remaining plant species to develop differently: they may take up fewer resources or become more prone to pathogens, both of which may alter their physical appearance. However, little is known about whether this happens and, if so, how rapidly such changes occur. Since 2002, researchers in Germany have been running a long-term project known as the Jena Experiment to study how plants behave when they grow in communities with different numbers of other plant species. For the experiment, various species of grass and other plants commonly found in grasslands were grown together in different combinations. Some plots contained many species (referred to as "high diversity") and others contained only a few ("low diversity"). Here, Dietrich et al. collected seeds from four grasses grown for 12 years in Jena Experiment plots with two or six plant species. The seeds were then transferred to pots and grown in a greenhouse using soil either from the plot where the seeds originated or from another plot with a different diversity level. To simulate human-made changes in the environment, the team added nitrogen fertilizer or decreased how much they watered some of the plants. The greenhouse experiment showed that after receiving nitrogen fertilizer, the seeds from the high diversity Jena Experiment plots grew into larger plants than the seeds from the low diversity plots. But there was no difference in size when the plants were watered less. Moreover, both fertilizer and watering treatment had different effects on the plants' physical appearance (root and leaf architecture) depending on the soil in which they were growing in. The findings of Dietrich et al. suggest that plants may respond differently to changes in their environment based on their origins and the soil they are growing in. This study provides the first indication that species loss could accelerate a further loss of species due to changes in how the plants develop and the communities of organisms living in the soil.

3D Melanoma Cocultures as Improved Models for Nanoparticle-Mediated Delivery of RNA to Tumors.

Cancer therapy is an emergent application for mRNA therapeutics. While in tumor immunotherapy, mRNA encoding for tumor-associated antigens is delivered to antigen-presenting cells in spleen and lymph nodes, other therapeutic options benefit from immediate delivery of mRNA nanomedicines directly to the tumor. However, tumor targeting of mRNA therapeutics is still a challenge, since, in addition to delivery of the cargo to the tumor, specifics of the targeted cell type as well as its interplay with the tumor microenvironment are crucial for successful intervention. This study investigated lipoplex nanoparticle-mediated mRNA delivery to spheroid cell culture models of melanoma. Insights into cell-type specific targeting, non-cell-autonomous effects, and penetration capacity in tumor and stroma cells of the mRNA lipoplex nanoparticles were obtained. It was shown that both coculture of different cell types as well as three-dimensional cell growth characteristics can modulate distribution and transfection efficiency of mRNA lipoplex formulations. The results demonstrate that three-dimensional coculture spheroids can provide a valuable surplus of information in comparison to adherent cells. Thus, they may represent in vitro models with enhanced predictivity for the in vivo activity of cancer nanotherapeutics.

Soft synthetic microgels as mimics of mycoplasma.

Artificial model colloids are of special interest in the development of advanced sterile filters, as they are able to efficiently separate pleomorphic, highly deformable and infectious bacteria such as mycoplasma, which, until now, has been considered rather challenging and laborious. This study presents a full range of different soft to super soft synthetic polymeric microgels, including two types with similar hydrodynamic mean diameter, i.e., 180 nm, and zeta potential, i.e., -25 ± 10 mV, but different deformability, synthesized by inverse miniemulsion terpolymerization of acrylamide, sodium acrylate and N,N'-methylenebisacrylamide. These microgels were characterized by means of dynamic, electrophoretic and static light scattering techniques. In addition, the deformability of the colloids was investigated by filter cake compressibility studies during ultrafiltration in dead-end mode, analogously to a study of real mycoplasma, i.e., Acholeplasma laidlawii, to allow for a direct comparison. The results indicate that the variation of the synthesis parameters, i.e., crosslinker content, polymeric solid content and content of sodium acrylate, has a significant impact on the swelling behavior of the microgels in aqueous solution as well as on their deformability under filtration conditions. A higher density of chemical crosslinking points results in less swollen and more rigid microgels. Furthermore, these parameters determine electrokinetic properties of the more or less permeable colloids. Overall, it is shown that these soft synthetic microgels can be obtained with tailor-made properties, covering the size of smallest species of and otherwise similar to real mycoplasma. This is a relevant first step towards the future use of synthetic microgels as mimics for mycoplasma.

It is not just a 'trade-off': indications for sink- and source-limitation to vegetative and regenerative growth in an old-growth beech forest.

Controls on tree growth are key issues in plant physiology. The hypothesis of our study was that the interannual variability of wood and fruit production are primarily controlled directly by weather conditions (sink limitation), while carbon assimilation (source limitation) plays a secondary role. We analyzed the interannual variability of weather conditions, gross primary productivity (GPP) and net primary productivity (NPP) of wood and fruits of an old-growth, unmanaged Fagus sylvatica forest over 14 yr, including six mast years. In a multiple linear regression model, c. 71% of the annual variation in wood-NPP could be explained by mean air temperature in May, precipitation from April to May (positive influence) and fruit-NPP (negative influence). GPP of June to July solely explained c. 42% of the variation in wood-NPP. Fruit-NPP was positively related to summer precipitation 2 yr before (R2  = 0.85), and negatively to precipitation in May (R2  = 0.83) in the fruit years. GPP had no influence on fruit-NPP. Our results suggest a complex system of sink and source limitations to tree growth driven by weather conditions and going beyond a simple carbon-mediated 'trade-off' between regenerative and vegetative growth.

[Online survey for assessment of geriatric early rehabilitation complex treatment in geriatric trauma centers of the DGU by the medical services of the health funds]. / Onlinebefragung zur Prüfung der geriatrisch frührehabilitativen Komplexbehandlung durch den Medizinischen Dienst der Krankenversicherung in AltersTraumaZentren DGU®.

BACKGROUND: Orthogeriatric co-management of proximal femoral fractures has been proven to effectively reduce mortality rates. This involves extending resources in hospitals treating these patients as well as dealing with the possibility of prolonged periods of hospitalization. The increase in costs of orthogeriatric co-management are best illustrated by the implementation of geriatric early rehabilitation complex treatment. In view of the problems concerning billing this complex treatment, an online survey was carried among certified geriatric trauma centers of the German Trauma Society (DGU®). METHODS: Based on a trauma-geriatric consensus 20 questions were formulated by the Academy of Trauma Surgery (AUC) as an online questionnaire and sent to all 75 certified geriatric trauma centers. Apart from a description of the results, a subanalysis based on the figures presented by the case closing departments (geriatrics or trauma surgery) was included. The questions covered a 2-year period of experiences from 2016 to 2018. RESULTS: A total of 26 of the 75 certified geriatric trauma centers participated (35%). A continuous increase in cost analysis evaluations by the medical services of the health funds was observed. A rise from 38% in 2016 to 45% in 2018 was seen. An analogous rejection trend from 16% to 24% during this period was evident as well. Subanalysis revealed significantly higher cost evaluation by the medical services of the health funds and cost rejection rates if trauma departments were the case closing disciplines. CONCLUSION: The online survey revealed significantly higher assessment and rejection rates when compared to other hospital services. This could prove potentially detrimental to the future of orthogeriatric co-management.

Functional groups differ in trait means, but not in trait plasticity to species richness in local grassland communities.

Despite growing interest in incorporating intraspecific variation of functional traits in community-level studies, it remains unclear whether species classified into functional groups based on interspecific trait differences are similar regarding their variation in trait expression in response to varying plant diversity and composition in local communities. In a large biodiversity experiment (Jena Experiment) designed on a trait-based a priori definition of functional groups (grasses, legumes, small herbs, tall herbs), we studied means, extent of variation (coefficient of variation across communities) and plasticity to increased plant diversity (slopes over a logarithmic species richness ranging from 1, 2, 4, 8 and 16 to 60 species) for nine functional traits. Species means and extent of variation in traits related to nitrogen (N) acquisition and N use differed among functional groups and were more similar in phylogenetically closely related species than expected by chance. Species in the same functional group showed a weak phylogenetic signal and varied widely in means and extent of variation in traits related to shoot architecture and to a smaller extent in leaf traits related to carbon acquisition. This indicated that functional groups were less distinguishable in light than in nitrogen acquisition strategies. The direction and degree of trait plasticity to increasing species richness did not show a phylogenetic signal and were not different among functional groups, but varied largely among species within functional groups. Correlation structures in trait means, extent of trait variation and trait plasticity revealed functional tradeoffs in the acquisition of nitrogen and light across species. While correlations between trait means and extent of trait variation varied from trait to trait (positive, negative or unrelated), trait means and trait plasticity were mostly unrelated. Our results suggest that the concept of functional groups is viable, but context-specific trait measurements are required to improve our understanding about the functional significance of intraspecific trait variation and interspecific trait differences in local plant communities.

Widespread expression of perilipin 5 in normal human tissues and in diseases is restricted to distinct lipid droplet subpopulations.

Diseases associated with the accumulation of lipid droplets are increasing in western countries. Lipid droplet biogenesis, structure and degradation are regulated by proteins of the perilipin family. Perilipin 5 has been shown to regulate basal lipolysis in oxidative tissues. We examine perilipin 5 in normal human tissues and in diseases using protein biochemical and microscopic techniques. Perilipin 5 was constitutively located at small lipid droplets in skeletal myocytes, cardiomyocytes and brown adipocytes. In addition, perilipin 5 was detected in the epithelia of the gastrointestinal and urogenital tract, especially in hepatocytes, the mitochondria-rich parietal cells of the stomach, tubular kidney cells and ductal cells of the salivary gland and pancreas. Granular cytoplasmic expression, without a lipid droplet-bound localization was detected elsewhere. In cardiomyopathies, in skeletal muscle diseases and during hepatocyte steatogenesis, perilipin 5 was upregulated and localized to larger and more numerous lipid droplets. In steatotic human hepatocytes, perilipin 5 was moderately increased and colocalized with perilipins 1 and 2 but not with perilipin 3 at lipid droplets. In liver diseases implicated in alterations of mitochondria, such as mitochondriopathies, alcoholic liver disease, Wilson's disease and acute liver injury, perilipin 5 was frequently localized to small lipid droplets and less in the cytoplasm. In tumorigenesis, perilipin 5 was especially upregulated in lipo-, leio- and rhabdomyosarcoma and hepatocellular and renal cell carcinoma. In summary, our study provides evidence that perilipin 5 is not restricted to certain cell types but localizes to distinct lipid droplet subpopulations reflecting a possible function in oxidative energy supply in normal tissues and in diseases.

Enhanced stability of a chimeric hepatitis B core antigen virus-like-particle (HBcAg-VLP) by a C-terminal linker-hexahistidine-peptide.

BACKGROUND: Virus-like-particles (VLPs) are attractive nanoparticulate scaffolds for broad applications in material/biological sciences and medicine. Prior their functionalization, specific adaptations have to be carried out. These adjustments frequently lead to disordered particles, but the particle integrity is an essential factor for the VLP suitability. Therefore, major requirements for particle stabilization exist. The objective of this study was to evaluate novel stabilizing elements for functionalized chimeric hepatitis B virus core antigen virus-like particles (HBcAg-VLP), with beneficial characteristics for vaccine development, imaging or delivery. RESULTS: The effects of a carboxy-terminal polyhistidine-peptide and an intradimer disulfide-bridge on the stability of preclinically approved chimeric HBcAg-VLPs were assessed. We purified recombinant chimeric HBcAg-VLPs bearing different modified C-termini and compared their physical and chemical particle stability by quantitative protein-biochemical and biophysical techniques. We observed lower chemical resistance of T = 3- compared to T = 4-VLP (triangulation number) capsids and profound impairment of accessibility of hexahistidine-peptides in assembled VLPs. Histidines attached to the C-terminus were associated with superior mechanical and/or chemical particle stability depending on the number of histidine moieties. A molecular modeling approach based on cryo-electron microscopy and biolayer interferometry revealed the underlying structural mechanism for the strengthening of the integrity of VLPs. Interactions triggering capsid stabilization occur on a highly conserved residue on the basis of HBcAg-monomers as well as on hexahistidine-peptides of adjacent monomers. This new stabilization mechanism appears to mimic an evolutionary conserved stabilization concept for hepadnavirus core proteins. CONCLUSIONS: These findings establish the genetically simply transferable C-terminal polyhistidine-peptide as a general stabilizing element for chimeric HBcAg-VLPs to increase their suitability.

Origin context of trait data matters for predictions of community performance in a grassland biodiversity experiment.

Plant functional traits may explain the positive relationship between species richness and ecosystem functioning, but species-level trait variation in response to growth conditions is often ignored in trait-based predictions of community performance. In a large grassland biodiversity experiment (Jena Experiment), we measured traits on plants grown as solitary individuals, in monocultures or in mixtures. We calculated two measures of community-level trait composition, i.e., community-weighted mean traits (CWM) and trait diversity (Rao's quadratic entropy; FD) based on different contexts in which traits were measured (trait origins). CWM and FD values of the different measurement origins were then compared regarding their power to predict community biomass production and biodiversity effects quantified with the additive partitioning method. Irrespective of trait origin, models combining CWM and FD values as predictors best explained community biomass and biodiversity effects. CWM values based on monoculture, mixture-mean or community-specific trait data were similarly powerful predictors, but predictions became worse when trait values originated from solitary-grown individuals. FD values based on monoculture traits were the best predictors of community biomass and net biodiversity effects, while FD values based on community-specific traits were the best predictors for complementarity and selection effects. Traits chosen as best CWM predictors were not strongly affected by trait origin but traits chosen as best FD predictors varied strongly dependent on trait origin and altered the predictability of community performance. We conclude that by adjusting their functional traits to species richness and even specific community compositions, plants can change community-level trait compositions, thereby also changing community biomass production and biodiversity effects. Incorporation of these plastic trait adjustments of plants in trait-based ecology can improve its predictive power in explaining biodiversity-ecosystem functioning relationships.

Gastroresistant oral peptide for fluorescence imaging of colonic inflammation.

The use of molecular markers for inflammation in the gastrointestinal tract could empower optical imaging modalities for early diagnosis and eventually personalized timely treatments. A major hurdle to the widespread use of functional fluorescence imaging is the absence of suitable contrast agents, in particular to be administered via the oral route due to the usual proteolytic susceptibility of the biomarkers. By designing a retro-inverso peptide, starting from a previously described sequence specific for N-cadherin, we achieved resistance to gastrointestinal degradation and even slightly improved specificity towards the target, both in ex vivo and in vivo experimental colitis. Simulations at fundamental molecular level suggested that the introduced retro-inverso modifications did not affect the folding of the peptide, leaving its ability to interact with the binding pocket of the monomeric N-cadherin unaltered, even when fluorescently labeled. Possible further derivatization of this sequence could be envisaged to further extend the potential of the designed retro-inverso peptide as diagnostic or theranostic agent for the oral route.

Fabrication of nanoporous graphene/polymer composite membranes.

Graphene is currently investigated as a promising membrane material in which selective pores can be created depending on the requirements of the application. However, to handle large-area nanoporous graphene a stable support material is needed. Here, we report on composite membranes consisting of large-area single layer nanoporous graphene supported by a porous polymer. The fabrication is based on ion-track nanotechnology with swift heavy ions directly creating atomic pores in the graphene lattice and damaged tracks in the polymer support. Subsequent chemical etching converts the latent ion tracks in the supporting polymer foil, here polyethylene terephthalate (PET), into open microchannels while the perfectly aligned pores in the graphene top layer remain unaffected. To avoid unintentional damage creation and delamination of the graphene layer from the substrate, the graphene is encapsulated by a protecting poly(methyl methacrylate) (PMMA) layer. By this procedure a stable composite membrane is obtained consisting of nanoporous graphene (coverage close to 100%) suspended across selfaligned track-etched microchannels in a polymer support film. Our method presents a facile way to create high quality suspended graphene of tunable pore size supported on a flexible porous polymeric support, thus enabling the development of membranes for fast and selective ultrafiltration separation processes.

The active force-length relationship is invisible during extensive eccentric contractions in skinned skeletal muscle fibres.

In contrast to experimentally observed progressive forces in eccentric contractions, cross-bridge and sliding-filament theories of muscle contraction predict that varying myofilament overlap will lead to increases and decreases in active force during eccentric contractions. Non-cross-bridge contributions potentially explain the progressive total forces. However, it is not clear whether underlying abrupt changes in the slope of the nonlinear force-length relationship are visible in long isokinetic stretches, and in which proportion cross-bridges and non-cross-bridges contribute to muscle force. Here, we show that maximally activated single skinned rat muscle fibres behave (almost across the entire working range) like linear springs. The force slope is about three times the maximum isometric force per optimal length. Cross-bridge and non-cross-bridge contributions to the muscle force were investigated using an actomyosin inhibitor. The experiments revealed a nonlinear progressive contribution of non-cross-bridge forces and suggest a nonlinear cross-bridge contribution similar to the active force-length relationship (though with increased optimal length and maximum isometric force). The linear muscle behaviour might significantly reduce the control effort. Moreover, the observed slight increase in slope with initial length is in accordance with current models attributing the non-cross-bridge force to titin.

Development of a Novel Immobilization Method by Using Microgels to Keep Enzyme in Hydrated Microenvironment in Porous Hydrophobic Membranes.

Using colloidal polyacrylamide (PAAm) microgels as carriers, a novel strategy for covalent immobilization of enzymes maintained in hydrated microenvironment on/in a macroporous surface-functionalized hydrophobic polyvinylidene fluoride (PVDF) membrane is developed. The PAAm microgels are synthesized by inverse miniemulsion polymerization, and first the parameters are investigated which are suited to obtain particles in the desired size range, 100-200 nm, with narrow size distribution. Amino functions are then imparted to the microgels applying the Hofmann reaction. The modification is confirmed by Fourier-transform infrared spectroscopy analysis, ninhydrin test, and elemental analysis. In addition, functionalized microgels are characterized by dynamic light scattering. The amino-functionalized PAAm microgels are then immobilized on pre-modified PVDF membrane having aldehyde functionalities on the surface. Afterward, unreacted aldehyde groups still present on the membrane where quenched by ethanolamine and the enzyme lipase from Candida rugosa (LCR) is subsequently immobilized on the microgels loaded PVDF membrane via glutaraldehyde cross-linking, exploiting the free amino groups on immobilized microgels. Catalytic efficiency of LCR immobilized by this strategy is evaluated using para-nitrophenyl palmitate as substrate and compared with LCR directly immobilized on PVDF membrane without microgels. Results show that LCR immobilized by means of microgels exhibits better performance with a 2.3-fold higher specific biocatalytic activity.

Resource Availability Alters Biodiversity Effects in Experimental Grass-Forb Mixtures.

Numerous experiments, mostly performed in particular environments, have shown positive diversity-productivity relationships. Although the complementary use of resources is discussed as an important mechanism explaining diversity effects, less is known about how resource availability controls the strength of diversity effects and how this response depends on the functional composition of plant communities. We studied aboveground biomass production in experimental monocultures, two- and four-species mixtures assembled from two independent pools of four perennial grassland species, each representing two functional groups (grasses, forbs) and two growth statures (small, tall), and exposed to different combinations of light and nutrient availability. On average, shade led to a decrease in aboveground biomass production of 24% while fertilization increased biomass production by 36%. Mixtures were on average more productive than expected from their monocultures (relative yield total, RYT>1) and showed positive net diversity effects (NE: +34% biomass increase; mixture minus mean monoculture biomass). Both trait-independent complementarity effects (TICE: +21%) and dominance effects (DE: +12%) positively contributed to net diversity effects, while trait-dependent complementarity effects were minor (TDCE: +1%). Shading did not alter diversity effects and overyielding. Fertilization decreased RYT and the proportion of biomass gain through TICE and TDCE, while DE increased. Diversity effects did not increase with species richness and were independent of functional group or growth stature composition. Trait-based analyses showed that the dominance of species with root and leaf traits related to resource conservation increased TICE. Traits indicating the tolerance of shade showed positive relationships with TDCE. Large DE were associated with the dominance of species with tall growth and low diversity in leaf nitrogen concentrations. Our field experiment shows that positive diversity effects are possible in grass-forb mixtures irrespective of differences in light availability, but that the chance for the complementary use of resources increases when nutrients are not available at excess.

Phenotypic plasticity to light and nutrient availability alters functional trait ranking across eight perennial grassland species.

Functional traits are often used as species-specific mean trait values in comparative plant ecology or trait-based predictions of ecosystem processes, assuming that interspecific differences are greater than intraspecific trait variation and that trait-based ranking of species is consistent across environments. Although this assumption is increasingly challenged, there is a lack of knowledge regarding to what degree the extent of intraspecific trait variation in response to varying environmental conditions depends on the considered traits and the characteristics of the studied species to evaluate the consequences for trait-based species ranking. We studied functional traits of eight perennial grassland species classified into different functional groups (forbs vs. grasses) and varying in their inherent growth stature (tall vs. small) in a common garden experiment with different environments crossing three levels of nutrient availability and three levels of light availability over 4 months of treatment applications. Grasses and forbs differed in almost all above- and belowground traits, while trait differences related to growth stature were generally small. The traits showing the strongest responses to resource availability were similarly for grasses and forbs those associated with allocation and resource uptake. The strength of trait variation in response to varying resource availability differed among functional groups (grasses > forbs) and species of varying growth stature (small-statured > tall-statured species) in many aboveground traits, but only to a lower extent in belowground traits. These differential responses altered trait-based species ranking in many aboveground traits, such as specific leaf area, tissue nitrogen and carbon concentrations and above-belowground allocation (leaf area ratio and root : shoot ratio) at varying resource supply, while trait-based species ranking was more consistent in belowground traits. Our study shows that species grouping according to functional traits is valid, but trait-based species ranking depends on environmental conditions, thus limiting the applicability of species-specific mean trait values in ecological studies.

Contrasting effects of intraspecific trait variation on trait-based niches and performance of legumes in plant mixtures.

Niche differentiation, assumed to be a key mechanism of species coexistence, requires that species differ in their functional traits. So far it remains unclear to which extent trait plasticity leads to niche shifts of species at higher plant diversity, thereby increasing or decreasing niche overlap between species. To analyse this question it is convenient to measure niches indirectly via the variation in resource-uptake traits rather than directly via the resources used. We provisionally call these indirectly measured niches trait-based niches. We studied shoot- and leaf-morphological characteristics in seven legume species in monoculture and multi-species mixture in experimental grassland. Legume species varied in the extent of trait variation in response to plant diversity. Trait plasticity led to significant shifts in species niches in multiple dimensions. Single-species niches in several traits associated with height growth and filling of canopy space were expanded, while other niche dimensions were compressed or did not change with plant diversity. Niche separation among legumes decreased in dimensions related to height growth and space filling, but increased in dimensions related to leaf size and morphology. The total extent of occupied niche space was larger in mixture than in the combined monocultures for dimensions related to leaf morphology and smaller for dimensions related to whole-plant architecture. Taller growth, greater space filling and greater plasticity in shoot height were positively, while larger values and greater plasticity in specific leaf area were negatively related with increased performance of species in mixture. Our study shows that trait variation in response to plant diversity shifts species niches along trait axes. Plastically increased niche differentiation is restricted to niche dimensions that are apparently not related to size-dependent differences between species, but functional equivalence (convergence in height growth) rather than complementarity (divergence in traits associated with light acquisition) explains increased performance of legumes in mixture.

Correction of Discretization Errors Simulated at Supply Wells.

Many hydrogeology problems require predictions of hydraulic heads in a supply well. In most cases, the regional hydraulic response to groundwater withdrawal is best approximated using a numerical model; however, simulated hydraulic heads at supply wells are subject to errors associated with model discretization and well loss. An approach for correcting the simulated head at a pumping node is described here. The approach corrects for errors associated with model discretization and can incorporate the user's knowledge of well loss. The approach is model independent, can be applied to finite difference or finite element models, and allows the numerical model to remain somewhat coarsely discretized and therefore numerically efficient. Because the correction is implemented external to the numerical model, one important benefit of this approach is that a response matrix, reduced model approach can be supported even when nonlinear well loss is considered.