Effects of increasing atmospheric CO2 on leaf water δ18O values are small and are attenuated in grasses and amplified in dicotyledonous herbs and legumes when transferred to cellulose δ18O values.

The oxygen isotope composition of cellulose (δ18O values) has been suggested to contain information on stomatal conductance (gs) responses to rising pCO2. The extent by which pCO2 affects leaf water and cellulose δ18O values (δ18OLW and δ18OC) and the isotope processes that determine pCO2 effects on δ18OLW and δ18OC are, however, unknown. We tested the effects of pCO2 on gs, δ18OLW and δ18OC in a glasshouse experiment, where six plant species were grown under pCO2 ranging from 200 to 500 ppm. Increasing pCO2 caused a decline in gs and an increase in δ18OLW, as expected. Importantly, the effects of pCO2 on gs and δ18OLW were small and pCO2 effects on δ18OLW were not directly transferred to δ18OC but were attenuated in grasses and amplified in dicotyledonous herbs and legumes. This is likely because of functional group-specific pCO2 effects on the model parameter pxpex. Our study highlights important uncertainties when using δ18OC as a proxy for gs. Specifically, pCO2-triggered gs effects on δ18OLW and δ18OC are possibly too small to be detected in natural settings and a pCO2 effect on pxpex may render the commonly assumed negative linkage between δ18OC and gs to be incorrect, potentially confounding δ18OC based gs reconstructions.

Water loss after stomatal closure: quantifying leaf minimum conductance and minimal water use in nine temperate European tree species during a severe drought.

Residual canopy transpiration (Emin_canop) is a key physiological trait that determines trees' survival time under drought after stomatal closure and after trees have limited access to soil water. Emin_canop mainly depends on leaf minimum conductance (gmin) and vapor pressure deficit. Here we determined the seasonal variation of gmin and how gmin is related to interspecies variation in leaf cuticular and stomatal traits for nine European tree species in a mature forest. In addition, we determined the species-specific temperature responses of gmin. With this newly obtained insight, we calculated Emin_canop for the nine species for one day at our research site during the 2022 central European hot drought. Our results show that at ambient temperatures gmin ranged from 0.8 to 4.8 mmol m-2 s-1 across the nine species and was stable in most species throughout the growing season. The interspecies variation of gmin was associated with leaf cuticular and stomatal traits. Additionally, gmin exhibited strong temperature responses and increased, depending on species, by a factor of two to four in the range of 25-50 °C. For the studied species at the site, during a single hot drought day, Emin_canop standardized by tree size (stem basal area) ranged from 2.0 to 36.7 L m-2, and non-standardized Emin_canop for adult trees ranged from 0.3 to 5.3 L. Emin_canop also exhibited species-specific rapid increases under hotter temperatures. Our results suggest that trees, depending on species, need reasonable amounts of water during a drought, even when stomates are fully closed. Species differences in gmin and ultimately Emin_canop can, together with other traits, affect the ability of a tree to keep its tissue hydrated during a drought and is likely to contribute to species-specific differences in drought vulnerability.

Uniform carbon reserve dynamics along the vertical light gradient in mature tree crowns.

Understanding the within-tree variability of non-structural carbohydrates (NSC) is crucial for interpreting point measurements and calculating whole-tree carbon balances. Yet, little is known about how the vertical light gradient within tree crowns influences branch NSC concentrations and dynamics. We measured NSC concentrations, irradiance and key leaf traits in uppermost, sun-exposed and lowest, shaded branches in the crowns of mature, temperate trees from nine species with high temporal resolution throughout one growing season. Measurements from two additional years allowed us to test the generality of our findings among climatically contrasting years. Despite the vertical light gradient, we found very similar seasonal NSC dynamics and concentrations between sun and shade branches in most species. This can at least partially be explained by acclimations in SLA and photosynthetic leaf traits compensating the different light availability between the top and bottom canopy. Only in the ring-porous species Quercus and Fraxinus, starch refilling after budbreak was slower in lower branches. End-of-season NSC concentrations were similar between canopy positions and among observation years. Only Fagus had 40 and 29% lower starch concentrations by the end of the extremely dry year 2020, relative to the other two years. We show that NSC measured anywhere in a tree crown is often representative of the whole crown. Overall, our results suggest that carbon reserve dynamics in trees are largely insensitive to both microclimatic gradients and inter-annual climatic variation, and only deviate under severe carbon deficits, as was presumably the case with Fagus in our study.

Plant choice between arbuscular mycorrhizal fungal species results in increased plant P acquisition.

Arbuscular mycorrhizal fungi (AMF) are plant root symbionts that provide phosphorus (P) to plants in exchange for photosynthetically fixed carbon (C). Previous research has shown that plants-given a choice among AMF species-may preferentially allocate C to AMF species that provide more P. However, these investigations rested on a limited set of plant and AMF species, and it therefore remains unclear how general this phenomenon is. Here, we combined 4 plant and 6 AMF species in 24 distinct plant-AMF species compositions in split-root microcosms, manipulating the species identity of AMF in either side of the root system. Using 14C and 32P/33P radioisotope tracers, we tracked the transfer of C and P between plants and AMF, respectively. We found that when plants had a choice of AMF species, AMF species which transferred more P acquired more C. Evidence for preferential C allocation to more beneficial AMF species within individual plant roots was equivocal. However, AMF species which transferred more P to plants did so at lower C-to-P ratios, highlighting the importance both of absolute and relative costs of P acquisition from AMF. When plants had a choice of AMF species, their shoots contained a larger total amount of P at higher concentrations. Our results thus highlight the benefits of plant C choice among AMF for plant P acquisition.

Stem growth phenology, not canopy greening, constrains deciduous tree growth.

Canopy phenology is a widely used proxy for deciduous forest growth with various applications in terrestrial ecosystem modeling. Its use relies on common assumptions that canopy greening and stem growth are tightly coordinated processes, enabling predictions on the timing and the quantity of annual tree growth. Here, we present parallel observations of canopy and stem growth phenology and annual stem increment in around 90 deciduous forest trees with diffuse-porous (Fagus sylvatica, Acer pseudoplatanus, Carpinus betulus) or ring-porous (Quercus robur × petraea) wood anatomy. These data were collected in a mixed temperate forest at the Swiss-Canopy-Crane II site, in 4 years with strongly contrasting weather conditions. We found that stem growth resumption lagged several weeks behind spring canopy greening in diffuse-porous but not in ring-porous trees. Canopy greening and stem growth resumption showed no or only weak signs of temporal coordination across the observation years. Within the assessed species, the seasonal timing of stem growth varied strongly among individuals, as trees with high annual increments resumed growth earlier and also completed their main growth earlier. The length of main growth activity had no influence on annual increments. Our findings not only challenge tight temporal coordination of canopy and stem growth phenology but also demonstrate that longer main growth activity does not translate into higher annual increments. This may compromise approaches modeling tree growth and forest productivity with canopy phenology and growth length.

Carbon dynamics during long-term starving poplar trees-the importance of older carbohydrates and a shift to lipids during survival.

Carbon (C) assimilation can be severely impaired during periods of environmental stress like drought or defoliation, making trees heavily dependent on the use of C reserve pools for survival; yet, dynamics of reserve use during periods of reduced C supply are still poorly understood. We used stem girdling in mature poplar trees (Populus tremula L. hybrids), a lipid-storing species, to permanently interrupt phloem C transport and induced C shortage in the isolated stem section below the girdle and monitored metabolic activity during three campaigns in the growing seasons of 2018, 2019, and 2021. We measured respiratory fluxes (CO2 and O2), NSC concentration, the respiratory substrate (based on isotopic analysis and CO2/O2 ratio) and the age of the respiratory substrate (based on radiocarbon analysis). Our study shows that poplar trees can survive long periods of reduced C supply from the canopy by switching in metabolism from recent carbohydrates to older storage pools with a potential mixture of respiratory substrates, including lipids. This mechanism of stress resilience can explain why tree decline may take many years until death occurs.

Post-drought compensatory growth in perennial grasslands is determined by legacy effects of the soil and not by plants.

Grasslands recovering from drought have repeatedly been shown to outperform non-drought-stressed grasslands in biomass production. The mechanisms that lead to the unexpectedly high biomass production in grasslands recovering from drought are, however, not understood. To disentangle plant-intrinsic and plant-extrinsic (soil) drought legacy effects on grassland recovery from drought, we designed a factorial field experiment where Lolium perenne plants that were exposed to either a 2-month drought or to well-watered control conditions were transplanted into control and drought-stressed soil and rewetted thereafter. Drought and rewetting (DRW) resulted in negative drought legacy effects of formerly drought-stressed plants (DRWp ) compared with control plants (Ctrp ) when decoupled from soil-mediated DRW effects, with DRWp showing less aboveground productivity (-13%), restricted N nutrition, and higher δ13 C compared with Ctrp . However, plants grown on formerly drought-stressed soil (DRWs ) showed enhanced aboveground productivity (+82%), improved N nutrition, and higher δ13 C values relative to plants grown on control soil (Ctrs ), irrespective of the plants' pretreatment. Our study shows that the higher post-drought productivity of perennial grasslands recovering from drought relative to non-drought-stressed controls is induced by soil-mediated DRW legacy effects which improve plant N nutrition and photosynthetic capacity and that these effects countervail negative plant-intrinsic drought legacy effects.

Covariation between oxygen and hydrogen stable isotopes declines along the path from xylem water to wood cellulose across an aridity gradient.

Oxygen and hydrogen isotopes of cellulose in plant biology are commonly used to infer environmental conditions, often from time series measurements of tree rings. However, the covariation (or the lack thereof) between δ18 O and δ2 H in plant cellulose is still poorly understood. We compared plant water, and leaf and branch cellulose from dominant tree species across an aridity gradient in Northern Australia, to examine how δ18 O and δ2 H relate to each other and to mean annual precipitation (MAP). We identified a decline in covariation from xylem to leaf water, and onwards from leaf to branch wood cellulose. Covariation in leaf water isotopic enrichment (Δ) was partially preserved in leaf cellulose but not branch wood cellulose. Furthermore, whilst δ2 H was well-correlated between leaf and branch, there was an offset in δ18 O between organs that increased with decreasing MAP. Our findings strongly suggest that postphotosynthetic isotope exchange with water is more apparent for oxygen isotopes, whereas variable kinetic and nonequilibrium isotope effects add complexity to interpreting metabolic-induced δ2 H patterns. Varying oxygen isotope exchange in wood and leaf cellulose must be accounted for when δ18 O is used to reconstruct climatic scenarios. Conversely, comparing δ2 H and δ18 O patterns may reveal environmentally induced shifts in metabolism.

Daytime stomatal regulation in mature temperate trees prioritizes stem rehydration at night.

Trees remain sufficiently hydrated during drought by closing stomata and reducing canopy conductance (Gc ) in response to variations in atmospheric water demand and soil water availability. Thresholds that control the reduction of Gc are proposed to optimize hydraulic safety against carbon assimilation efficiency. However, the link between Gc and the ability of stem tissues to rehydrate at night remains unclear. We investigated whether species-specific Gc responses aim to prevent branch embolisms, or enable night-time stem rehydration, which is critical for turgor-dependent growth. For this, we used a unique combination of concurrent dendrometer, sap flow and leaf water potential measurements and collected branch-vulnerability curves of six common European tree species. Species-specific Gc reduction was weakly related to the water potentials at which 50% of branch xylem conductivity is lost (P50 ). Instead, we found a stronger relationship with stem rehydration. Species with a stronger Gc control were less effective at refilling stem-water storage as the soil dries, which appeared related to their xylem architecture. Our findings highlight the importance of stem rehydration for water-use regulation in mature trees, which likely relates to the maintenance of adequate stem turgor. We thus conclude that stem rehydration must complement the widely accepted safety-efficiency stomatal control paradigm.

Networking the forest infrastructure towards near real-time monitoring - A white paper.

Forests account for nearly 90 % of the world's terrestrial biomass in the form of carbon and they support 80 % of the global biodiversity. To understand the underlying forest dynamics, we need a long-term but also relatively high-frequency, networked monitoring system, as traditionally used in meteorology or hydrology. While there are numerous existing forest monitoring sites, particularly in temperate regions, the resulting data streams are rarely connected and do not provide information promptly, which hampers real-time assessments of forest responses to extreme climate events. The technology to build a better global forest monitoring network now exists. This white paper addresses the key structural components needed to achieve a novel meta-network. We propose to complement - rather than replace or unify - the existing heterogeneous infrastructure with standardized, quality-assured linking methods and interacting data processing centers to create an integrated forest monitoring network. These automated (research topic-dependent) linking methods in atmosphere, biosphere, and pedosphere play a key role in scaling site-specific results and processing them in a timely manner. To ensure broad participation from existing monitoring sites and to establish new sites, these linking methods must be as informative, reliable, affordable, and maintainable as possible, and should be supplemented by near real-time remote sensing data. The proposed novel meta-network will enable the detection of emergent patterns that would not be visible from isolated analyses of individual sites. In addition, the near real-time availability of data will facilitate predictions of current forest conditions (nowcasts), which are urgently needed for research and decision making in the face of rapid climate change. We call for international and interdisciplinary efforts in this direction.

The metabolic sensitivity of hydrogen isotope fractionation differs between plant compounds.

Hydrogen stable isotope analyses (δ2H) of plant derived organic compounds are a useful tool for ecological, environmental, and palaeoclimatological research. However, during organic compound synthesis, variable biosynthetic 2H-fractionation has been suggested to occur as a result of changes in plant carbon fluxes. So far, inference has been based on examining the δ2H patterns of plant compounds along environmental gradients, among plant species, and between plant organs. In an alternative approach, we used four plant species with four different types of mutations that cause impaired starch synthesis to assess whether variability in carbon metabolism affects the biosynthetic 2H-fractionation during cellulose, phytol, and acetogenic lipid synthesis. We found that mutants with impaired starch synthesis always had higher cellulose and phytol δ2H values compared to the wild type. By contrast, 2H-fractionation during acetogenic lipid biosynthesis generally did not show strong metabolic sensitivity. We rationalise these differences by considering the biosynthetic pathway of each compound and the likely source of the variable isotope fractionation. In different organic compounds, the sensitivity of variable biosynthetic 2H-fractionation to changes in C-metabolism depends on incorporation of specific H atoms from precursor molecules. As such, we determined that the similar increase in cellulose and phytol δ2H values as an effect of impaired starch synthesis most likely originates in triose-phosphates.

Species variation in the hydrogen isotope composition of leaf cellulose is mostly driven by isotopic variation in leaf sucrose.

Experimental approaches to isolate drivers of variation in the carbon-bound hydrogen isotope composition (δ2 H) of plant cellulose are rare and current models are limited in their application. This is in part due to a lack in understanding of how 2 H-fractionations in carbohydrates differ between species. We analysed, for the first time, the δ2 H of leaf sucrose along with the δ2 H and δ18 O of leaf cellulose and leaf and xylem water across seven herbaceous species and a starchless mutant of tobacco. The δ2 H of sucrose explained 66% of the δ2 H variation in cellulose (R2 = 0.66), which was associated with species differences in the 2 H enrichment of sucrose above leaf water ( ε sucrose : -126% to -192‰) rather than by variation in leaf water δ2 H itself. ε sucrose was positively related to dark respiration (R2 = 0.27), and isotopic exchange of hydrogen in sugars was positively related to the turnover time of carbohydrates (R2 = 0.38), but only when ε sucrose was fixed to the literature accepted value of - 171 ‰. No relation was found between isotopic exchange of hydrogen and oxygen, suggesting large differences in the processes shaping post-photosynthetic fractionation between elements. Our results strongly advocate that for robust applications of the leaf cellulose hydrogen isotope model, parameterization utilizing δ2 H of sugars is needed.

Constraining parameter uncertainty for predicting oxygen and hydrogen isotope values in fruit.

Understanding δ18O and δ2H values of agricultural products like fruit is of particular scientific interest in plant physiology, ecology, and forensic studies. Applications of mechanistic stable isotope models to predict δ18O and δ2H values of water and organic compounds in fruit, however, are hindered by a lack of empirical parameterizations and validations. We addressed this lack of data by experimentally evaluating model parameter values required to model δ18O and δ2H values of water and organic compounds in berries and leaves from strawberry and raspberry plants grown at different relative humidities. Our study revealed substantial differences between leaf and berry isotope values, consistent across the different relative humidity treatments. We demonstrated that existing isotope models can reproduce water and organic δ18O and δ2H values for leaves and berries. Yet, these simulations require organ-specific model parameterization to accurately predict δ18O and δ2H values of leaf and berry tissue and water pools. We quantified these organ-specific model parameters for both species and relative humidity conditions. Depending on the required model accuracy, species- and environment-specific model parameters may be justified. The parameter values determined in this study thus facilitate applications of stable isotope models where understanding δ18O and δ2H values of fruit is of scientific interest.

Explicitly accounting for needle sugar pool size crucial for predicting intra-seasonal dynamics of needle carbohydrates δ18 O and δ13 C.

We explore needle sugar isotopic compositions (δ18 O and δ13 C) in boreal Scots pine (Pinus sylvestris) over two growing seasons. A leaf-level dynamic model driven by environmental conditions and based on current understanding of isotope fractionation processes was built to predict δ18 O and δ13 C of two hierarchical needle carbohydrate pools, accounting for the needle sugar pool size and the presence of an invariant pinitol pool. Model results agreed well with observed needle water δ18 O, δ18 O and δ13 C of needle water-soluble carbohydrates (sugars + pinitol), and needle sugar δ13 C (R2 = 0.95, 0.84, 0.60, 0.73, respectively). Relative humidity (RH) and intercellular to ambient CO2 concentration ratio (Ci /Ca ) were the dominant drivers of δ18 O and δ13 C variability, respectively. However, the variability of needle sugar δ18 O and δ13 C was reduced on diel and intra-seasonal timescales, compared to predictions based on instantaneous RH and Ci /Ca , due to the large needle sugar pool, which caused the signal formation period to vary seasonally from 2 d to more than 5 d. Furthermore, accounting for a temperature-sensitive biochemical 18 O-fractionation factor and mesophyll resistance in 13 C-discrimination were critical. Interpreting leaf-level isotopic signals requires understanding on time integration caused by mixing in the needle sugar pool.

Do 2 H and 18 O in leaf water reflect environmental drivers differently?

We compiled hydrogen and oxygen stable isotope compositions (δ2 H and δ18 O) of leaf water from multiple biomes to examine variations with environmental drivers. Leaf water δ2 H was more closely correlated with δ2 H of xylem water or atmospheric vapour, whereas leaf water δ18 O was more closely correlated with air relative humidity. This resulted from the larger proportional range for δ2 H of meteoric waters relative to the extent of leaf water evaporative enrichment compared with δ18 O. We next expressed leaf water as isotopic enrichment above xylem water (Δ2 H and Δ18 O) to remove the impact of xylem water isotopic variation. For Δ2 H, leaf water still correlated with atmospheric vapour, whereas Δ18 O showed no such correlation. This was explained by covariance between air relative humidity and the Δ18 O of atmospheric vapour. This is consistent with a previously observed diurnal correlation between air relative humidity and the deuterium excess of atmospheric vapour across a range of ecosystems. We conclude that 2 H and 18 O in leaf water do indeed reflect the balance of environmental drivers differently; our results have implications for understanding isotopic effects associated with water cycling in terrestrial ecosystems and for inferring environmental change from isotopic biomarkers that act as proxies for leaf water.

Lack of hydraulic recovery as a cause of post-drought foliage reduction and canopy decline in European beech.

European beech (Fagus sylvatica) was among the most affected tree species during the severe 2018 European drought. It not only suffered from instant physiological stress but also showed severe symptoms of defoliation and canopy decline in the following year. To explore the underlying mechanisms, we used the Swiss-Canopy-Crane II site and studied in branches of healthy and symptomatic trees the repair of hydraulic function and concentration of carbohydrates during the 2018 drought and in 2019. We found loss of hydraulic conductance in 2018, which did not recover in 2019 in trees that developed defoliation symptoms in the year after drought. Reduced branch foliation in symptomatic trees was associated with a gradual decline in wood starch concentration throughout summer 2019. Visualization of water transport in healthy and symptomatic branches in the year after the drought confirmed the close relationship between xylem functionality and supported branch leaf area. Our findings showed that embolized xylem does not regain function in the season following a drought and that sustained branch hydraulic dysfunction is counterbalanced by the reduction in supported leaf area. It suggests acclimation of leaf development after drought to mitigate disturbances in canopy hydraulic function.

Effects of phenotypic variability on the oxygen and hydrogen isotope compositions of grains in different winter wheat varieties.

Stable isotope analyses are the leading method for geographic origin determination, especially of plant-based agricultural products. Origin analysis is typically done by comparing a suspicious sample to reference materials with known geographic origin. Reference materials are usually collected at the species level, assuming different varieties of a species to have comparable isotope compositions within a given location. We evaluated whether different phenotypes that are expressed in different varieties of winter wheat (Triticum aestivum L.) influence the oxygen (δ18O) and hydrogen (δ2H) isotope composition of plant tissue water and organic compounds. We found that mean δ18O and δ2H values among winter wheat varieties did not differ significantly in leaf water, however, differed significantly in bulk dried grain tissue. The differences in bulk dried grain δ18O and δ2H values among varieties can be related to differences in phenotypic trait expression among varieties. Despite this substantial phenotypic variability, the overall variability of bulk dried grain δ18O and δ2H values among varieties was small (SD 0.54 ‰ for oxygen, 3.60 ‰ for hydrogen). We thus conclude that reference materials collected at the species level should be sufficient for geographic origin analysis of winter wheat and possibly other cereals using δ18O and δ2H values.

Number of growth days and not length of the growth period determines radial stem growth of temperate trees.

Radial stem growth dynamics at seasonal resolution are essential to understand how forests respond to climate change. We studied daily radial growth of 160 individuals of seven temperate tree species at 47 sites across Switzerland over 8 years. Growth of all species peaked in the early part of the growth season and commenced shortly before the summer solstice, but with species-specific seasonal patterns. Day length set a window of opportunity for radial growth. Within this window, the probability of daily growth was constrained particularly by air and soil moisture, resulting in intermittent growth to occur only on 29 to 77 days (30% to 80%) within the growth period. The number of days with growth largely determined annual growth, whereas the growth period length contributed less. We call for accounting for these non-linear intra-annual and species-specific growth dynamics in tree and forest models to reduce uncertainties in predictions under climate change.

Climate Change Modulates Multitrophic Interactions Between Maize, A Root Herbivore, and Its Enemies.

How climate change will modify belowground tritrophic interactions is poorly understood, despite their importance for agricultural productivity. Here, we manipulated the three major abiotic factors associated with climate change (atmospheric CO2, temperature, and soil moisture) and investigated their individual and joint effects on the interaction between maize, the banded cucumber beetle (Diabrotica balteata), and the entomopathogenic nematode (EPN) Heterorhabditis bacteriophora. Changes in individual abiotic parameters had a strong influence on plant biomass, leaf wilting, sugar concentrations, protein levels, and benzoxazinoid contents. Yet, when combined to simulate a predicted climate scenario (Representative Concentration Pathway 8.5, RCP 8.5), their effects mostly counter-balanced each other. Only the sharp negative impact of drought on leaf wilting was not fully compensated. In both current and predicted scenarios, root damage resulted in increased leaf wilting, reduced root biomass, and reconfigured the plant sugar metabolism. Single climatic variables modulated the herbivore performance and survival in an additive manner, although slight interactions were also observed. Increased temperature and CO2 levels both enhanced the performance of the insect, but elevated temperature also decreased its survival. Elevated temperatures and CO2 further directly impeded the EPN infectivity potential, while lower moisture levels improved it through plant- and/or herbivore-mediated changes. In the RCP 8.5 scenario, temperature and CO2 showed interactive effects on EPN infectivity, which was overall decreased by 40%. We conclude that root pest problems may worsen with climate change due to increased herbivore performance and reduced top-down control by biological control agents.