Changes in structure of over‐ and midstory tree species in a Mediterranean‐type forest after an extreme drought‐associated heatwave

Worldwide, extreme climatic events such as drought and heatwaves are associated with forest mortality. However, the precise drivers of tree mortality at individual and stand levels vary considerably, with substantial gaps in knowledge across studies in biomes and continents. In 2010–2011, a drought‐associated heatwave occurred in south‐western Australia and drove sudden and rapid forest canopy collapse. Working in the Northern Jarrah (Eucalyptus marginata) Forest, we quantified the response of key overstory (E. marginata, Corymbia calophylla) and midstory (Banksia grandis, Allocasuarina fraseriana) tree species to the extreme climate event. Using transects spanning a gradient of drought impacts (minimal (50–100 m), transitional (100–150 m) and severe (30–60 m)), tree species mortality in relation to stand characteristics (stand basal area and stem density) and edaphic factors (soil depth) was determined. We show differential mortality between the two overstory species and the two midstory species corresponding to the drought‐associated heatwave. The dominant overstory species, E. marginata, had significantly higher mortality (~19%) than C. calophylla (~7%) in the severe zone. The midstory species, B. grandis, demonstrated substantially higher mortality (~59%) than A. fraseriana (~4%) in the transitional zone. Banksia grandis exhibited a substantial shift in structure in response to the drought‐associated heatwave in relation to tree size, basal area and soil depth. This study illustrates the role of climate extremes in driving ecosystem change and highlights the critical need to identify and quantify the resulting impact to help predict future forest die‐off events and to underpin forest management and conservation.     

Examining the response of needle carbohydrates from Siberian larch trees to climate using compound‐specific δ13C and concentration analyses

Little is known about the dynamics of concentrations and carbon isotope ratios of individual carbohydrates in leaves in response to climatic and physiological factors. Improved knowledge of the isotopic ratio in sugars will enhance our understanding of the tree ring isotope ratio and will help to decipher environmental conditions in retrospect more reliably. Carbohydrate samples from larch (Larix gmelinii) needles of two sites in the continuous permafrost zone of Siberia with differing growth conditions were analysed with the Compound‐Specific Isotope Analysis (CSIA). We compared concentrations and carbon isotope values (δ¹³C) of sucrose, fructose, glucose and pinitol combined with phenological data. The results for the variability of the needle carbohydrates show high dynamics with distinct seasonal characteristics between and within the studied years with a clear link to the climatic conditions, particularly vapour pressure deficit. Compound‐specific differences in δ¹³C values as a response to climate were detected. The δ¹³C of pinitol, which contributes up to 50% of total soluble carbohydrates, was almost invariant during the whole growing season. Our study provides the first in‐depth characterization of compound‐specific needle carbohydrate isotope variability, identifies involved mechanisms and shows the potential of such results for linking tree physiological responses to different climatic conditions.     

Seasonal dynamics of δ13C of C‐rich fractions from Picea abies (Norway spruce) and Fagus sylvatica (European beech) fine roots

The ^13/12C ratio in plant roots is likely dynamic depending on root function (storage versus uptake), but to date, little is known about the effect of season and root order (an indicator of root function) on the isotopic composition of C‐rich fractions in roots. To address this, we monitored the stable isotopic composition of one evergreen (Picea abies) and one deciduous (Fagus sylvatica), tree species' roots by measuring δ¹³C of bulk, respired and labile C, and starch from first/second and third/fourth order roots during spring and fall root production periods. In both species, root order differences in δ¹³C were observed in bulk organic matter, labile, and respired C fractions. Beech exhibited distinct seasonal trends in δ¹³C of respired C, while spruce did not. In fall, first/second order beech roots were significantly depleted in ¹³C, whereas spruce roots were enriched compared to higher order roots. Species variation in δ ¹³C of respired C may be partially explained by seasonal shifts from enriched to depleted C substrates in deciduous beech roots. Regardless of species identity, differences in stable C isotopic composition of at least two root order groupings (first/second, third/fourth) were apparent, and should hereafter be separated in belowground C‐supply‐chain inquiry.     

An unbiased method to estimate individual specialisation from multi‐tissue isotopic data

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Changes in wood and foliar δ13C in sugar maple at Gatineau Park,Quebec, Canada

Leaf samples and tree rings formed between the mid‐1960s and mid‐1990s from sugar maple (Acer saccharum Marsh.) at Gatineau Park (45°30′ N, 75°54′ W), Quebec were analysed for δ¹³C. Leaf samples were collected at ground level (1–2 m above ground) at monthly intervals during the summer, whereas tree cores were extracted from the largest trees (d.b.h. > 30 cm) in the young deciduous forest in August 1998. Significant linear decreases in δ¹³C over time were found in foliage and tree rings, but the decrease in δ¹³C was significantly greater in foliage than in the wood. The apparent isotopic discrimination (Δ) of tree rings varied insignificantly around a mean of 18‰, whereas foliar Δ increased significantly from 19‰ in the 1960s to around 23‰ by the mid 1990s, likely as a result of an increasing canopy effect as the forest matured. Using models of carbon discrimination and Δ‐values of the tree rings, we calculate that the intrinsic water use efficiency of mature sugar maple has increased by approximately 4% over the study period.     

Auxin biosynthetic gene TAR2 is involved in low nitrogen‐mediated reprogramming of root architecture in Arabidopsis

In plants, the plasticity of root architecture in response to nitrogen availability largely determines nitrogen acquisition efficiency. One poorly understood root growth response to low nitrogen availability is an observed increase in the number and length of lateral roots (LRs). Here, we show that low nitrogen‐induced Arabidopsis LR growth depends on the function of the auxin biosynthesis gene TAR2 (tryptophan aminotransferase related 2). TAR2 was expressed in the pericycle and the vasculature of the mature root zone near the root tip, and was induced under low nitrogen conditions. In wild type plants, low nitrogen stimulated auxin accumulation in the non‐emerged LR primordia with more than three cell layers and LR emergence. Conversely, these low nitrogen‐mediated auxin accumulation and root growth responses were impaired in the tar2‐c null mutant. Overexpression of TAR2 increased LR numbers under both high and low nitrogen conditions. Our results suggested that TAR2 is required for reprogramming root architecture in response to low nitrogen conditions. This finding suggests a new strategy for improving nitrogen use efficiency through the engineering of TAR2 expression in roots.     

Dual‐tracer‐based isotope turnover rates in a highly invasive mysid Limnomysis benedeni from Lake Constance

Understanding the ecological patterns of invasive species and their habitats require an understanding of the species’ foraging ecology. Stable carbon (δ¹³C) and nitrogen (δ¹⁵N) isotope values provide useful information into the study of animal ecology and evolution, since the isotope ratios of consumers reflect consumer's dietary patterns. Nevertheless, the lack of species‐ and element‐specific laboratory‐derived turnover rates could limit their application. Using a laboratory‐based dual stable isotope tracer approach (Na¹⁵NO₃ and NaH¹³CO₃), we evaluated the δ¹⁵N and δ¹³C isotope turnover rates in full‐grown adult invasive Limnomysis benedeni from Lake Constance. We provide δ¹⁵N and δ¹³C turnover rates based on nonlinear least‐squares regression and posterior linear regression models. Model precisions and fit were evaluated using Akaike's information criterion. Within a couple of days, the δ¹⁵N and δ¹³C of mysids began to change. Nevertheless, after about 14 days, L. benedeni did not reach equilibrium with their new isotope values. Since the experiment was conducted on adult subjects, it is evident that turnover was mainly influenced by metabolism (in contrast to growth). Unlike traditional dietary shifts, our laboratory‐based dual stable isotope tracer approach does not shift the experimental organisms into a new diet and avoids dietary effects on isotope values. Results confirm the application of isotopic tracers to label mysid subpopulations and could be used to reflect assimilation and turnover from the labeled dietary sources. Field‐based stable isotope studies often use isotopic mixing models commonly assuming diet‐tissue steady state. Unfortunately, in cases where the isotopic composition of the animal is not in equilibrium with its diet, this can lead to highly misleading conclusions. Thus, our laboratory‐based isotopic incorporation rates assist interpretation of the isotopic values from the field and provide a foundation for future research into using isotopic tracers to investigate invasion ecology.     

Effects of spawning Pacific salmon on the isotopic composition of biota differ among southeast Alaska streams

1. Adult Pacific salmon (Oncorhynchus spp.) transport marine nutrients to fresh waters and disturb sediments during spawning. The relative importance of nutrient fertilisation and benthic disturbance by salmon spawners can be modulated by environmental conditions (e.g. biological, chemical and physical conditions in the catchment, including human land use). 2. To determine the importance of the environmental context in modifying the uptake and incorporation of salmon‐derived material into stream biota, we measured the nitrogen (δ¹⁵N) and carbon (δ¹³C) isotopic composition of benthic algae (i.e. epilithon) and juvenile coho salmon (Oncorhynchus kisutch) in seven streams across a timber‐harvest gradient (8–69% catchment area harvested), both before and during the salmon run. Conditional bootstrap modelling simulations were used to assess variability in the response of epilithon and juvenile coho salmon to spawning salmon. 3. In response to spawning salmon, epilithon exhibited enrichment in both δ¹⁵N (mean: 1.5‰) and δ¹³C (2.3‰). Juvenile coho were also enriched in both δ¹⁵N (0.7‰) and δ¹³C (1.4‰). Conditional bootstrap models indicate decreased variation in data as spatial replication increases, suggesting that the number of study sites can influence the results of Pacific salmon isotope studies. 4. Epilithon isotopic enrichment was predicted by environmental conditions, with δ¹⁵N enrichment predicted by stream temperature and timber harvest (R² = 0.87) and δ¹³C enrichment by discharge, sediment size, timber harvest and spawner density (R² = 0.96). Furthermore, we found evidence for a legacy effect of salmon spawners, with pre‐spawner δ¹⁵N and δ¹³C of both epilithon and juvenile coho predicted by salmon run size in the previous year. 5. Our results show that the degree of incorporation of salmon‐derived nitrogen and carbon differs among streams. Furthermore, the environmental context, including putative legacy effects of spawning salmon, can influence background isotopic concentrations and utilisation of salmon‐derived materials in southeast Alaska salmon streams. Future studies should consider the variation in isotopic composition of stream biota when deciding on the number of study sites and samples needed to generate meaningful results.     

Small‐mammal isotope ecology tracks climate and vegetation gradients across western North America

Stable carbon, nitrogen, hydrogen and oxygen isotopes have been used to infer aspects of species ecology and environment in both modern ecosystems and the fossil record. Compared to large mammals, stable isotopic studies of small‐mammal ecology are limited; however, high species and ecological diversity within small mammals presents several advantages for quantifying resource use and organism–environment interactions using stable isotopes over various spatial and temporal scales. We analyzed the isotopic composition of hair from two heteromyid rodent species, Dipodomys ordii and Perognathus parvus, from localities across western North America in order to characterize dietary variation in relation to vegetation and climatic gradients. Significant correlations between the carbon isotopic composition (δ¹³C) of these species and several climatic variables imply that seasonal temperature and precipitation control the composition and distribution of dietary resources (grass seeds). Our results also suggest a moisture influence on the nitrogen isotopic composition (δ¹⁵N) of heteromyid diets. Population‐ and species‐level variation in δ¹³C and δ¹⁵N values record fine‐scale habitat heterogeneity and significant differences in resource use between species. Using classification and regression‐tree techniques, we modeled the geographic variation in heteromyid δ¹³C_diet values based on 10 climatic variables and generated an isotope landscape model (‘isoscape’). The isoscape predictions for δ¹³C_diet differ from expectations based on observed C₄ distributions and instead indicate that D. ordii and P. parvus record seasonally abundant grass resources, with additional model deviations potentially attributed to geographic variation in dietary selection. The oxygen and hydrogen isotopic composition of D. ordii is enriched relative to local meteoric water and suggests that individuals rely on highly evaporated water sources, such as seed moisture. Based on the climatic influences on vegetation and diet documented in this study, the isotopic composition of small mammals has high potential for recording ecological responses to environmental changes over short and long time scales.     

Fine‐scale variation in natural nitrogen isotope ratios of ants

Stable isotopes provide a powerful means of elucidating the trophic ecology of organisms. Analyses of variation in the ratio of nitrogen isotopes (δ¹⁵N) can provide insights into the trophic position of species with broad diets and the ability to occupy multiple positions in food webs, such as ants. The most powerful studies compare subjects across various spatial scales, but to do so, local variation in δ¹⁵N baselines must be taken into account. To date, a wide variety of baseline calibration methods have been employed, leading some authors to suggest that a standard approach is needed, and that the reality of environmental variation necessitates that this should be at fine scales. In this study, we examine the fine‐scale variation in δ¹⁵N value of colonies of the ant Formica kozlovi Dlussky (Hymenoptera: Formicidae: Formicini) along a sloped transect in Mongolia, and compare these with values for associated soils in an effort to shed further light on this issue. We find variation in ant δ¹⁵N to the order of one trophic level (ca. 3‰), over a distance of only 1 km. Ant δ¹⁵N was highly correlated with soil δ¹⁵N, and variation in mineral soil δ¹⁵N explained ca. 81% of the variation in ant δ¹⁵N. This study underlines the importance of local‐scale baseline corrections for isotopic studies, particularly in environments where baseline variation might be expected. It further suggests that δ¹⁵N of mineral soils may provide a suitable baseline for ecological studies of terrestrial arthropods.     

Changes in isotopic signatures of soil carbon and CO2 respiration immediately and one year after Miscanthus removal

The removal of perennial bioenergy crops, such as Miscanthus, has rarely been studied although it is an important form of land use change. Miscanthus is a C4 plant, and the carbon (C) it deposits during its growth has a different isotopic signature (¹²/¹³C) compared to a C3 plant. Identifying the proportion of C stored and released to the atmosphere is important information for ecosystem models and life cycle analyses. During a removal experiment in June 2011 of a 20‐year old Miscanthus field (Grignon, France), vegetation was removed mechanically and chemically. Two replicate plots were converted into a rotation of annual crops, two plots had Miscanthus removed with no soil disturbance, followed by bare soil (set‐aside), one control plot was left with continued Miscanthus cultivation, and an adjacent field was used as annual arable crops control. There was a significant difference in the isotopic composition of the total soil C under Miscanthus compared with adjacent annual arable crops in all three measured soil layers (0–5, 5–10 and 10–20 cm). Before Miscanthus removal, total C in the soil under Miscanthus ranged from 4.9% in the top layer to 3.9% in the lower layers with δ¹³C values of ?16.3 to ?17.8 while soil C under the adjacent arable crop was significantly lower and ranged from 1.6 to 2% with δ¹³C values of ?23.2. This did not change much in 2012, suggesting the accumulation of soil C under Miscanthus persists for at least the first year. In contrast, the isotopic signals of soil respiration 1 year after Miscanthus removal from recultivated and set‐aside plots were similar to that of the annual arable control, while just after removal the signals were similar to that of the Miscanthus control. This suggests a rapid change in the form of soil C pools that are respired.     

From pattern to process: linking intrinsic water‐use efficiency to drought‐induced forest decline

The rise in atmospheric CO₂ concentrations (Ca) has been related to tree growth enhancement and increasing intrinsic water‐use efficiency (iWUE). However, the extent that rising Ca has led to increased long‐term iWUE and whether climate could explain deviations from expected Ca‐induced growth enhancement are still poorly understood. The aim of this research was to use Ca and local climatic variability to explain changes during the 20th century in growth and tree ring and needle δ¹³C in declining and nondeclining Abies alba stands from the Spanish Pyrenees, near the southern distribution limit of this species. The temporal trends of iWUE were calculated under three theoretical scenarios for the regulation of plant‐gas exchange at increasing Ca. We tested different linear mixed‐effects models by multimodel selection criteria to predict basal area increment (BAI), a proxy of tree radial growth, using these scenarios and local temperature together with precipitation data as predictors. The theoretical scenario assuming the strongest response to Ca explained 66–81% of the iWUE variance and 28–56% of the observed BAI variance, whereas local climatic variables together explained less than 11–21% of the BAI variance. Our results are consistent with a drought‐induced limitation of the tree growth response to rising CO₂ and a decreasing rate of iWUE improvement from the 1980s onward in declining A. alba stands subjected to lower water availability.     

Affordable and robust phenotyping framework to analyse root system architecture of soil‐grown plants

The phenotypic analysis of root system growth is important to inform efforts to enhance plant resource acquisition from soils; however, root phenotyping remains challenging because of the opacity of soil, requiring systems that facilitate root system visibility and image acquisition. Previously reported systems require costly or bespoke materials not available in most countries, where breeders need tools to select varieties best adapted to local soils and field conditions. Here, we report an affordable soil‐based growth (rhizobox) and imaging system to phenotype root development in glasshouses or shelters. All components of the system are made from locally available commodity components, facilitating the adoption of this affordable technology in low‐income countries. The rhizobox is large enough (approximately 6000 cm² of visible soil) to avoid restricting vertical root system growth for most if not all of the life cycle, yet light enough (approximately 21 kg when filled with soil) for routine handling. Support structures and an imaging station, with five cameras covering the whole soil surface, complement the rhizoboxes. Images are acquired via the Phenotiki sensor interface, collected, stitched and analysed. Root system architecture (RSA) parameters are quantified without intervention. The RSAs of a dicot species (Cicer arietinum, chickpea) and a monocot species (Hordeum vulgare, barley), exhibiting contrasting root systems, were analysed. Insights into root system dynamics during vegetative and reproductive stages of the chickpea life cycle were obtained. This affordable system is relevant for efforts in Ethiopia and other low‐ and middle‐income countries to enhance crop yields and climate resilience sustainably.     

Exploring cycad foliage as an archive of the isotopic composition of atmospheric nitrogen

Molecular nitrogen (N₂) constitutes the majority of Earth's modern atmosphere, contributing ~0.79 bar of partial pressure (pN₂). However, fluctuations in pN₂ may have occurred on 10⁷–10⁹ year timescales in Earth's past, perhaps altering the isotopic composition of atmospheric nitrogen. Here, we explore an archive that may record the isotopic composition of atmospheric N₂ in deep time: the foliage of cycads. Cycads are ancient gymnosperms that host symbiotic N₂‐fixing cyanobacteria in modified root structures known as coralloid roots. All extant species of cycads are known to host symbionts, suggesting that this N₂‐fixing capacity is perhaps ancestral, reaching back to the early history of cycads in the late Paleozoic. Therefore, if the process of microbial N₂ fixation records the δ¹⁵N value of atmospheric N₂ in cycad foliage, the fossil record of cycads may provide an archive of atmospheric δ¹⁵N values. To explore this potential proxy, we conducted a survey of wild cycads growing in a range of modern environments to determine whether cycad foliage reliably records the isotopic composition of atmospheric N₂. We find that neither biological nor environmental factors significantly influence the δ¹⁵N values of cycad foliage, suggesting that they provide a reasonably robust record of the δ¹⁵N of atmospheric N₂. Application of this proxy to the record of carbonaceous cycad fossils may not only help to constrain changes in atmospheric nitrogen isotope ratios since the late Paleozoic, but also could shed light on the antiquity of the N₂‐fixing symbiosis between cycads and cyanobacteria.     

Evidence for seasonal variation in δ13C and δ18O profiles of Baculites and implications for growth rate

The life histories of ammonites and the life strategies they employed are difficult to assess without robust modern analogues but placing constraints on ammonite growth rates provides a fundamental first step to understanding this abundant, but poorly understood, fossil group. Here we interpret periodic variations in carbon and oxygen stable isotope profiles from Campanian and Maastrichtian ammonites (Baculites) as seasonally driven and use these records to determine their rate of shell precipitation. Several of these samples are housed in museums and were originally prepared using sealants for display and preservation but testing of these sealants indicated no alteration of the isotopic values of treated carbonate. Diagenetic alteration, as determined by shell microstructure, affected the preservation of isotopic signals, resulting in the loss of seasonal variation in less well‐preserved specimens, and the δ¹³C signal is more robust than δ¹⁸O. The periodicity of isotopic profiles from Baculites shells presented here suggest that these organisms grew at rapid rates (c. 340 mm per year), which may imply an r‐type life strategy in which the animals reach maturity quickly, spawn large quantities of progeny, and die at a young age. Because of the potential mobility of Baculites, reconstructing palaeoenvironmental conditions from these isotopic records is challenging and should be conducted cautiously. Unfortunately, well‐preserved Baculites shells much longer than 350 mm are rarely recovered, which complicates the statistical treatment of potential periodicity in isotopic profiles.     

Age‐related variation in the trophic characteristics of a marsupial carnivore,the Tasmanian devil Sarcophilus harrisii

Age‐related changes in diet have implications for competitive interactions and for predator–prey dynamics, affecting individuals and groups at different life stages. To quantify patterns of variation and ontogenetic change in the diets of Tasmanian devils Sarcophilus harrisii, a threatened marsupial carnivore, we analyzed variation in the stable isotope composition of whisker tissue samples taken from 91 individual devils from Wilmot, Tasmania from December 2014 to February 2017. Both δ¹³C and δ¹⁵N decreased with increasing age in weaned Tasmanian devils, indicating that as they age devils rely less on small mammals and birds, and more on large herbivores. Devils <12 months old had broader group isotopic niches, as estimated by Bayesian standard ellipses (SEA_B mode = 1.042) than devils from 12 to 23 months old (mode = 0.541) and devils ≥24 months old (mode = 0.532). Devils <24 months old had broader individual isotopic niches (SEA_B mode range 0.492–1.083) than devils ≥24 months old (mode range 0.092–0.240). A decrease in δ¹⁵N from the older whisker sections to the more recently grown sections in devils <24 months old likely reflects the period of weaning in this species, as this pattern was not observed in devils ≥24 months old. Our data reveal changes in the isotopic composition of devil whiskers with increasing age, accompanied by a reduction in isotopic variation both among population age classes and within individuals, reflecting the effect of weaning in early life, and a likely shift from an initially diverse diet of small mammals, birds, and invertebrates towards increasing consumption of larger herbivores in adulthood.     

Carbon sequestration in soil in a semi‐natural Miscanthus sinensis grassland and Cryptomeria japonica forest plantation in Aso,Kumamoto, Japan

Although Miscanthus sinensis grasslands (Misc‐GL) and Cryptomeria japonica forest plantations (Cryp‐FP) are proposed bioenergy feedstock systems, their relative capacity to sequester C may be an important factor in determining their potential for sustainable bioenergy production. Therefore, our objective was to quantify changes in soil C sequestration 47 years after a Misc‐GL was converted to a Cryp‐FP. The study was conducted on adjacent Misc‐GL and Cryp‐FP located on Mt. Aso, Kumamoto, Japan. After Cryp‐FP establishment, only the Misc‐GL continued to be managed by annual burning every March. Mass C and N, δ¹³C, and δ¹⁵N at 0–30 cm depth were measured in 5 cm increments. Carbon and N concentrations, C:N ratio, δ¹³C, and δ¹⁵N were measured in litter and/or ash, and rhizomes or roots. Although C input in Misc‐GL by M. sinensis was approximately 36% of that in Cryp‐FP by C. japonica, mean annual soil C sequestration in Misc‐GL (503 kg C ha^?1 yr^?1) was higher than that in Cryp‐FP (284 kg C ha^?1 yr^?1). This was likely the result of larger C input from aboveground litter to soil, C‐quality (C:N ratio and lignin concentration in aboveground litter) and possibly more recalcitrant C (charcoal) inputs by annual burning. The difference in soil δ¹⁵N between sites indicated that organic C with N had greater cycling between heterotrophic microbes and soil and produces more recalcitrant humus in Misc‐GL than in Cryp‐FP. Our data indicate that in terms of soil C sequestration, maintenance of Misc‐GL may be more advantageous than conversion to Cryp‐FP in Aso, Japan.     

Variation in the carbon and oxygen isotope composition of plant biomass and its relationship to water‐use efficiency at the leaf‐ and ecosystem‐scales in a northern Great Plains grassland

Measurements of the carbon (δ¹³C_m) and oxygen (δ¹⁸O_m) isotope composition of C₃ plant tissue provide important insights into controls on water‐use efficiency. We investigated the causes of seasonal and inter‐annual variability in water‐use efficiency in a grassland near Lethbridge, Canada using stable isotope (leaf‐scale) and eddy covariance measurements (ecosystem‐scale). The positive relationship between δ¹³C_m and δ¹⁸O_m values for samples collected during 1998–2001 indicated that variation in stomatal conductance and water stress‐induced changes in the degree of stomatal limitation of net photosynthesis were the major controls on variation in δ¹³C_m and biomass production during this time. By comparison, the lack of a significant relationship between δ¹³C_m and δ¹⁸O_m values during 2002, 2003 and 2006 demonstrated that water stress was not a significant limitation on photosynthesis and biomass production in these years. Water‐use efficiency was higher in 2000 than 1999, consistent with expectations because of greater stomatal limitation of photosynthesis and lower leaf c_i/ca during the drier conditions of 2000. Calculated values of leaf‐scale water‐use efficiency were 2–3 times higher than ecosystem‐scale water‐use efficiency, a difference that was likely due to carbon lost in root respiration and water lost during soil evaporation that was not accounted for by the stable isotope measurements.     

Extraction of water from Eucalyptus trees for analysis of deuterium and oxygen-18: laboratory and field techniques

This study investigated a methodology for sampling and extracting water from Eucalyptus spp. for analysis of stable isotopes of water. Firstly, water of known isotopic composition was extracted from stems of seedlings of Eucalyptus spp. by azeotropic distillation using three solvents (hexane, toluene and kerosene) in laboratory and glasshouse experiments. Kerosene and toluene gave the most accurate extractions, with a small negative bias (～-2‰ for δ²H and ～-0.4‰ for δ¹⁸O) due to incomplete extraction. This bias was not evident in further experiments, using kerosene, where water was extracted from stems and roots that had been pre-treated in a manner that more closely reflected field conditions. Secondly, variation in δ²H in trees was investigated in the field to determine an accurate and convenient place for sampling. There was no significant difference in δ²H between water extracted from trunk sapwood and whole twigs, even twigs close to leaves, in samples taken from four Eucalyptus spp. at two field sites. The results mean that the most convenient part of the tree can be sampled. Also, there was little difference in δ²H between sapwood and heart-wood samples, although small increases (～ 2‰) in δ²H were observed in bark of trunks and twigs. Tree roots (15–25 mm diameter) had similar δ²H values to the surrounding soil in many cases, especially surface (<0.2 m depth) roots, rather than having δ²H values similar to twigs of the trees. These roots were absorbing water from surrounding soil, but not contributing significant quantities of water to the tree canopy. Thus, the δ²H of root water is not a good indicator of the isotopic composition of water in the upper parts of Eucalyptus trees.     

Nitrogen and carbon isotope variability in the green‐algal lichen Xanthoria parietina and their implications on mycobiont–photobiont interactions

Stable isotope patterns in lichens are known to vary largely, but effects of substrate on carbon and nitrogen stable isotope signatures of lichens were previously not investigated systematically. N and C contents and stable isotope (δ¹⁵N, δ¹³C) patterns have been measured in 92 lichen specimens of Xanthoria parietina from southern Bavaria growing on different substrates (bark and stone). Photobiont and mycobiont were isolated from selected populations and isotopically analyzed. Molecular investigations of the internal transcribed spacer of the nuclear ribosomal DNA (ITS nrDNA) region have been conducted on a subset of the specimens of X. parietina. Phylogenetic analysis showed no correlation between the symbionts X. parietina and Trebouxia decolorans and the substrate, isotope composition, or geographic origin. Instead specimens grown on organic substrate significantly differ in isotope values from those on minerogenic substrate. This study documents that the lichens growing on bark use additional or different N sources than the lichens growing on stone. δ¹⁵N variation of X. parietina apparently is controlled predominantly by the mass fraction of the mycobiont and its nitrogen isotope composition. In contrast with mycobionts, photobionts of X. parietina are much more ¹⁵N‐depleted and show less isotopic variability than mycobionts, probably indicating a mycobiont‐independent nitrogen acquisition by uptake of atmospheric ammonia.