Foliar and litter needle carbon and oxygen isotope compositions relate to tree growth of an exotic pine plantation under different residue management regimes in subtropical Australia

Background and aim

Significant differences in tree growth were observed in an exotic pine plantation under different harvest residue management regimes at ages 2–10 years. However, the variations in tree growth between residue management treatments could not be explained by soil and foliar nutrient analyses, except by potassium (K) concentration. Therefore, this study determined the carbon isotope composition (δ¹³C) and oxygen isotope composition (δ¹⁸O) of current and archived foliar samples from the exotic pine plantation to establish relationships with foliar K concentration and tree growth indices as a means to determine changes in stomatal conductance (g_s) and photosynthetic rate (A_max) or water use efficiency (WUE), and therefore understand the variations in tree growth across treatments.

Methods

The harvest residue treatments were: (1) residue removal, RR₀; (2) single level residue retention, RR₁; and (3) double level residue retention, RR₂. Foliar δ¹³C and δ¹⁸O were determined for samples at ages 2, 4, 6 and 10 years, and the atmospheric ¹³C discrimination (Δ¹³C), intercellular CO₂ concentration (C_i) and WUE were determined from the δ¹³C data. Litter needle δ¹³C and δ¹⁸O were also determined over 15 months between ages 9 and 10 years. These parameters or variables where correlated to each other as well as to the periodic mean annual increment of basal area (PAIB) and the periodic mean annual increment of tree diameter at breast height (PAID) across the treatments and over time. Foliar δ¹³C and δ¹⁸O were also related to published data of foliar K concentrations of the same trees.

Results

Significant variations of foliar δ¹³C, and therefore WUE and Δ¹³C, across treatments were only observed at ages 4 and 10 years old, and foliar δ¹⁸O at age 4 years old only. The results showed increasing foliar δ¹³C, δ¹⁸O and WUE, and decreasing Δ¹³C and Ci, from RR₀ to RR₂ treatments. However, while the WUE was positively related to the PAID and PAIB at age 4 years, it was negatively related to PAID and PAIB at age 10 years old. Litter needle δ¹³C, indicative of WUE, was also negatively related to the PAID at age 10 years old. . At age 4 years, foliar δ¹³C and δ¹⁸O were positively related with a steep slope of 7.70 ‰ across treatments, and that both isotopes were positively related to foliar K concentrations. Similarly, δ¹⁸O was negatively related to the Δ¹³C. No significant relationship can be determined between foliar δ¹³C, or Δ¹³C, and δ¹⁸O at age 10 years old. In addition, WUE was increasing (p?<?0.001) and Δ¹³C and C_i decreasing (p?<?0.001) with decreasing PAID over time.

Conclusions

The variations at age 4 years in foliar δ¹³C or Δ¹³C and δ¹⁸O and increasing WUE with increasing growth rate suggest growth induced water-stress with increasing residue-loading rate as a result of the nutritional effect of the harvest residues on tree growth. At age 10 years, the negative relationships between WUE and PAID indicate nutrient limitation has an over-riding effect on δ¹³C variations rather than g_s. This was due to the lack of a significant relationship between foliar Δ¹³C and δ¹⁸O at this age, as well as over time.     

Carbon isotope compositions (δ13C) of leaf,wood and holocellulose differ among genotypes of poplar and between previous land uses in a short‐rotation biomass plantation

The efficiency of water use to produce biomass is a key trait in designing sustainable bioenergy‐devoted systems. We characterized variations in the carbon isotope composition (δ¹³C) of leaves, current year wood and holocellulose (as proxies for water use efficiency, WUE) among six poplar genotypes in a short‐rotation plantation. Values of δ¹³C_wood and δ¹³C_{holocellulose} were tightly and positively correlated, but the offset varied significantly among genotypes (0.79–1.01‰). Leaf phenology was strongly correlated with δ¹³C, and genotypes with a longer growing season showed a higher WUE. In contrast, traits related to growth and carbon uptake were poorly linked to δ¹³C. Trees growing on former pasture with higher N‐availability displayed higher δ¹³C as compared with trees growing on former cropland. The positive relationships between δ¹³C_leaf and leaf N suggested that spatial variations in WUE over the plantation were mainly driven by an N‐related effect on photosynthetic capacities. The very coherent genotype ranking obtained with δ¹³C in the different tree compartments has some practical outreach. Because WUE remains largely uncoupled from growth in poplar plantations, there is potential to identify genotypes with satisfactory growth and higher WUE.     

Seasonal variations in leaf δ13C values: implications for different water-use strategies among species growing on continuous dolomite outcrops in subtropical China

Plant species growing in shallow-soil habitat are likely to experience water deficit especially in seasonally dry or arid regions. However, only scarce studies focused on their water-use strategies. The current study aimed to reveal water-use strategies of different species growing on continuous dolomite outcrops (a typical shallow-soil habitat) in subtropical China that relied on different water sources, and to investigate the differences between narrow endemic and widespread species, based on season variations in leaf δ¹³C values. Leaf samples of six plant species (Radermachera sinica, Sapium rotundifolium, Sterculia euosma, Schefflera octophylla, Alchornea trewioides, and Vitex negundo, in different life-forms and leaf phenologies) were collected for carbon isotope measurements in the wet and dry seasons, respectively. Contrary to the expectation, the evergreen big shrub species, S. octophylla, which always relied on deep water sources, exhibited the most positive δ¹³C values (high water-use efficiency, WUE), indicating more conservative water-use strategies. While the two deciduous small shrubs, A. trewioides and V. negundo, which always relied on shallow water sources, exhibited the most negative δ¹³C values (low WUE). This result was associated with their short life spans, indicating an opportunistic water-use strategy. Leaf δ¹³C values of almost all (except for S. octophylla) the selected species were significantly (P < 0.05) higher in the dry season than in the wet season. This indicated that it was a common strategy for species in rocky karst habitat to improve their WUE in dry season. Despite the similar water sources utilized by the selected three tree species, the widespread one (R. sinica) exhibited greater improvement in leaf δ¹³C values than the narrow endemic ones (S. rotundifolium and S. euosma). This suggested that the widespread tree species had more flexible water-use strategies. It was further speculated that broad spatial distribution of widespread species may contribute to their highly plastic responses to changes in environmental conditions rather than always maintaining high WUE.     

Species-specific differences in temporal and spatial variation in δ13C of plant carbon pools and dark-respired CO2 under changing environmental conditions

Stable carbon isotope signatures are often used as tracers for environmentally driven changes in photosynthetic δ¹³C discrimination. However, carbon isotope signatures downstream from carboxylation by Rubisco are altered within metabolic pathways, transport and respiratory processes, leading to differences in δ¹³C between carbon pools along the plant axis and in respired CO₂. Little is known about the within-plant variation in δ¹³C under different environmental conditions or between species. We analyzed spatial, diurnal, and environmental variations in δ¹³C of water soluble organic matter (δ¹³C_WSOM) of leaves, phloem and roots, as well as dark-respired δ¹³CO₂ (δ¹³C_res) in leaves and roots. We selected distinct light environments (forest understory and an open area), seasons (Mediterranean spring and summer drought) and three functionally distinct understory species (two native shrubs—Halimium halimifolium and Rosmarinus officinalis—and a woody invader—Acacia longifolia). Spatial patterns in δ¹³C_WSOM along the plant vertical axis and between respired δ¹³CO₂ and its putative substrate were clearly species specific and the most δ¹³C-enriched and depleted values were found in δ¹³C of leaf dark-respired CO₂ and phloem sugars, ~?15 and ~?33 ‰, respectively. Comparisons between study sites and seasons revealed that spatial and diurnal patterns were influenced by environmental conditions. Within a species, phloem δ¹³C_WSOM and δ¹³C_res varied by up to 4 ‰ between seasons and sites. Thus, careful characterization of the magnitude and environmental dependence of apparent post-carboxylation fractionation is needed when using δ¹³C signatures to trace changes in photosynthetic discrimination.     

Thermal imaging and carbon isotope composition indicate variation amongst strawberry (Fragaria × ananassa) cultivars in stomatal conductance and water use efficiency

Cultivars with low stomatal conductance (g_s) may show high water use efficiency (WUE) under drought conditions, but under optimal conditions low g_s may result in low vigour. A combination of thermal imaging and carbon isotope composition (δ¹³C) analysis offers potential for screening simultaneously for both high g_s and high WUE. Ten cultivars of strawberry (Fragaria × ananassa Duch.) were grown in well watered or water limited conditions. Thermal images were taken of the plants, with various approaches to determine the optimal protocol for detecting variation in g_s, including use of reference leaves, grids to maintain leaves horizontal, and collection of meteorological data in synchrony with thermal images. δ¹³C of leaves, fruit, and crowns was determined. An index of g_s derived from the temperature of horizontal leaves and the temperature of wet and dry references showed significant differences between cultivars and between well watered and water limited plants, as did g_s estimated from leaf temperature, the temperature of a dry reference, and humidity. Thermal imaging indicated low g_s in ‘Elsanta’ and ‘Totem’ and relatively high g_s in well watered ‘Elvira’, ‘Florence’ and ‘Cambridge Favourite’. δ¹³C of all plant material was higher in water limited than well watered plants and showed significant differences between cultivars. In one experiment leaf δ¹³C indicated lowest WUE in ‘Elvira’ and highest WUE in ‘Totem’. δ¹³C was inversely correlated with an index of g_s derived from thermal imaging. Although the results indicate substantial variation in g_s and WUE between cultivars, generally all cultivars responded to water deficit by lowering g_s and hence increasing WUE.     

Does geographic origin dictate ecological strategies in Acacia senegal (L.) Willd.? Evidence from carbon and nitrogen stable isotopes

Background and aims

Acacia senegal, a leguminous dryland tree, is economically and ecologically important to sub-Saharan Africa. Water-use efficiency (WUE) and biological nitrogen fixation (BNF) are fundamental to plant productivity and survival. We quantify provenance differences in WUE, BNF, photosynthesis, biomass and gum arabic production from A. senegal assessing genetic improvement potential.

Methods

Using stable isotope ratios, we determined WUE (δ¹³C) and BNF (δ¹⁵N) from provenances of mature A. senegal in field-trials (Senegal), sampling leaves at the beginning (wet) and end (dry) of the rainy season. Seedling provenance trials (UK) determined photosynthesis, and biomass and δ¹³C in relation to water table. Environmental data were characterised for all provenances at their sites of origin.

Results

Provenances differed in both δ¹³C and δ¹⁵N. Gum yield declined with increasing WUE. Virtually no BNF was detected during the dry season and seedlings and mature trees may have different WUE strategies. Wind speed and soil characteristics at provenance origin were correlated with isotope composition and gum production.

Conclusion

Provenance differences suggest that selection for desirable traits, e.g., increased gum production, may be possible. As ecological strategies relate to native locality, the environmental conditions at plantation site and provenance origin are important in assessing selection criteria.     

The physiological basis for genetic variation in water use efficiency and carbon isotope composition in Arabidopsis thaliana

Ecologists and physiologists have documented extensive variation in water use efficiency (WUE) in Arabidopsis thaliana, as well as association of WUE with climatic variation. Here, we demonstrate correlations of whole-plant transpiration efficiency and carbon isotope composition (δ¹³C) among life history classes of A. thaliana. We also use a whole-plant cuvette to examine patterns of co-variation in component traits of WUE and δ¹³C. We find that stomatal conductance (g _s) explains more variation in WUE than does A. Overall, there was a strong genetic correlation between A and g _s, consistent with selection acting on the ratio of these traits. At a more detailed level, genetic variation in A was due to underlying variation in both maximal rate of carboxylation (V _cmax) and maximum electron transport rate (Jmax). We also found strong effects of leaf anatomy, where lines with lower WUE had higher leaf water content (LWC) and specific leaf area (SLA), suggesting a role for mesophyll conductance (g _m) in variation of WUE. We hypothesize that this is due to an effect through g _m, and test this hypothesis using the abi4 mutant. We show that mutants of ABI4 have higher SLA, LWC, and g _m than wild-type, consistent with variation in leaf anatomy causing variation in g _m and δ¹³C. These functional data also add further support to the central, integrative role of ABI4 in simultaneously altering ABA sensitivity, sugar signaling, and CO₂ assimilation. Together our results highlight the need for a more holistic approach in functional studies, both for more accurate annotation of gene function and to understand co-limitations to plant growth and productivity.     

Responses of pigmentation and δ13C in Qilian juniper to cold and drought stresses under natural conditions in the Qilian Mountains,China

The seasonal changes of pigments and stable carbon isotope composition (δ¹³C values) were investigated in needles of Qilian juniper (Sabina przewalskii Kom.) from two typical sites, one drier and the other wetter, in the Qilian Mountains, China. The anthocyanins and rhodoxanthin content from both sites were much higher in winter than in summer. Plant needles contained more carotenoids and xanthophylls in winter at the wetter site, while no seasonal difference appeared at the drier site. However, lower chlorophyll content and higher proline and δ¹³C were observed at the drier site. Average tree height was shorter at the drier site trees than at the wetter site. Our results suggested that under natural conditions, pigmentation in S. przewalskii may serve to improve the energy balance of the photosynthetic apparatus under cold and drought stresses. Proline and δ¹³C could be used as drought indicators for S. przewalskii.     

Changes in leaf photosynthetic characteristics and water use efficiency along with tree height of 4 tree species

As main photosynthetic organs, leaves are very sensitive to exterior environments. Water deficiency obviously affects the biological and physiological characteristics of leaves. Xylem pathways increase when trees grow tall, which results in the increase in water gravity as well as pathway resistance. Accordingly, the physiological characteristics of leaves change along with tree height. In this research, the photosynthetic characteristics and carbon isotope ratio (δ¹³C) in the leaves of 4 tree species, Platanus hispanica, Robinia pseudoacacia, Fraxinus chinensis and Ginkgo biloba, were measured. The results showed that the leaf photosynthetic rate (Pn), transpiration rate (Tr), stomatal conductance (Cond) and internal CO₂ concentration (Ci) reduced along with tree height, while the leaf δ¹³C increased along with tree height. The One Way ANOVA and LSD tests showed that the leaf photosynthetic characteristics and δ¹³C varied significantly at different tree heights (P < 0.05). The decrease in leaf photosynthetic capability and the increase in δ¹³C along with tree height indicate that the leaves at the tree tops suffer from water stress. These results support the hydraulic limitation hypothesis.     

Correlating δ13C and δ18O in cellulose of trees

We measured the carbon and oxygen isotopic composition of stem cellulose of Pinus sylvestris, Picea abies, Fagus sylvatica and Fraxinus excelsior. Several sites along a transect of a small valley in Switzerland were selected which differ in soil moisture conditions. At every site, six trees per species were sampled, and a sample representing a mean value for the period from 1940 to 1990 was analysed. For all species, the mean site δ¹³C and δ¹⁸O of stem cellulose are related to the soil moisture availability, whereby higher isotope ratios are found at drier sites. This result is consistent with isotope fractionation models when assuming enhanced stomatal resistance (thus higher δ¹³C of incorporated carbon) and increased oxygen isotope enrichment in the leaf water (thus higher δ¹⁸O) at the dry sites. δ¹⁸ O-δ¹³C plots reveal a linear relationship between the carbon and oxygen isotopes in cellulose. To interpret this relationship we developed an equation which combines the above-mentioned fractionation models. An important new parameter is the degree to which the leaf water enrichment is reflected in the stem cellulose. In the combined model the slope of the δ¹⁸O-δ¹³C plot is related to the sensitivity of the p_i/p_a of a plant to changing relative humidity.     

Carbon isotope ratios of photolithotrophs from allt meall nan damh,a burn at ardeonaig,Perthshire, and their ecophysiological significance

Summary

δ¹³C measurements were made of dissolved inorganic C, and of submerged benthic cyanobacteria, algae and bryophytes, from Allt Meall nan Damh, a burn at Ardeonaig, Perthshire. The δ¹³C of the CO₂, HCO3/- and CO3/2- components of the inorganic C were computed, and the Δ values of the organic C in the photolithotrophs were then calculated relative to dissolved CO₂. The decreasing order of A values in the Ardeonaig Burn is Lemanea and bryophytes ≥ green macroalgae and Audouinella > diatom mats, which is the same as in the Dighty Burn. However, the Δ values of Lemanea and the bryophytes, which depend on diffusive CO₂ entry, are lower at Ardeonaig than in the Dighty Burn, suggesting greater diffusive limitation to photosynthesis in the Ardeonaig Burn. It is not easy to relate this difference in Δ values in Lemanea to the higher C:N atomic ratio in the Ardeonaig Burn (21.2 ± 0.64) than in the Dighty Burn (9.5–11.0). The Δ values relative to HCO3/- for the HCO3/--using diatom mat in the Ardeonaig Burn is also lower than that in the Dighty Burn; this is consistent with a greater diffusion limitation of photosynthesis in the thicker mats in the Ardeonaig Burn. The δ¹³C of a Lyngbya mat overlying a Lemanea population stranded by low summer water levels indicates that some of the C fixed by the HCO3/--using Lyngbya comes from respiration of low-δ¹³C inorganic C by the Lemanea which is shaded by the Lyngbya. The δ¹³C values of Mesotaenium in its mucilage sheath on a thinly vegetated bank is suggestive of predominant use of the higher CO₂ concentrations with lower δ¹³C from groundwater rather than of atmospheric CO₂ yielding lower dissolved CO₂ concentrations with a higher δ¹³C value.     

Identifying drivers of leaf water and cellulose stable isotope enrichment in <Emphasis Type="Italic">Eucalyptus</Emphasis> in northern Australia

Several previous studies have investigated the use of the stable hydrogen and oxygen isotope compositions in plant materials as indicators of palaeoclimate. However, accurate interpretation relies on a detailed understanding of both physiological and environmental drivers of the variations in isotopic enrichments that occur in leaf water and associated organic compounds. To progress this aim we measured δ¹⁸O and δ²H values in eucalypt leaf and stem water and δ¹⁸O values in leaf cellulose, along with the isotopic compositions of water vapour, across a north-eastern Australian aridity gradient. Here we compare observed leaf water enrichment, along with previously published enrichment data from a similar north Australian transect, to Craig–Gordon-modelled predictions of leaf water isotopic enrichment. Our investigation of model parameters shows that observed ¹⁸O enrichment across the aridity gradients is dominated by the relationship between atmospheric and internal leaf water vapour pressure while ²H enrichment is driven mainly by variation in the water vapour—source water isotopic disequilibrium. During exceptionally dry and hot conditions (RH < 21%, T > 37 °C) we observed strong deviations from Craig–Gordon predicted isotope enrichments caused by partial stomatal closure. The atmospheric–leaf vapour pressure relationship is also a strong predictor of the observed leaf cellulose δ¹⁸O values across one aridity gradient. Our finding supports a wider applicability of leaf cellulose δ¹⁸O composition as a climate proxy for atmospheric humidity conditions during the leaf growing season than previously documented.     

Biomonitoring of urban habitat quality by anatomical and chemical leaf characteristics

This study focused on the potential of specific leaf area, stomatal density and stomatal pore surface as easy-to-measure plant parameters in low cost biomonitoring of urban habitat quality with a high spatial resolution. The study area (81.5 km²) was the city of Gent, Belgium. In the study area 148 sampling locations were identified within 4 land use classes. Specific leaf area, stomatal density, stomatal pore surface, minimal stomatal resistance, chlorophyll a and b, C and N content, δ¹³C and δ¹⁵N in the leaf samples of a common herbaceous plant Taraxacum officinalis were measured. The stomatal pore surface and minimal stomatal resistance of T. officinalis varied significantly between land use classes. In the harbor and industry land use class and the urban land use class a 27% and 21% lower mean stomatal pore surface at the abaxial leaf surface, and a 29% and 27% lower mean stomatal pore surface at the adaxial leaf surface was observed compared to that in the pasture land use class. The minimal stomatal resistance at the abaxial leaf surface was significantly higher in the urban land use class and harbor and industry land use class by 28% and 29%, respectively compared to that in the pasture land use class. In addition, urbanized and industrial land use classes as the harbour and industry and the urban land use classes showed significantly lower δ¹³C values compared to pasture land use class. The specific leaf area, stomatal parameters and δ¹³C data were geostatistically analyzed to understand their spatial variation. The spatial distributions of stomatal pore surface and minimal stomatal resistance varied considerably across the study area, indicating a different habitat quality from the harbour area in the north, over the city centre in the middle and the industrial areas in the south, compared to off city areas. Spatial patterns of δ¹³C showed depleted δ¹³C levels in city areas indicating the diluted δ¹³C in the urban atmosphere by fuel combustion. We concluded that stomatal characteristics seem to be the most promising parameter for estimating urban habitat quality.     

Comparison of seasonal variations in water-use efficiency calculated from the carbon isotope composition of tree rings and flux data in a temperate forest

Tree-ring δ(13) C is often interpreted in terms of intrinsic water-use efficiency (WUE) using a carbon isotope discrimination model established at the leaf level. We examined whether intra-ring δ(13) C could be used to assess variations in intrinsic WUE (W(g), the ratio of carbon assimilation and stomatal conductance to water) and variations in ecosystem WUE (W(t) , the ratio of C assimilation and transpiration) at a seasonal scale. Intra-ring δ(13) C was measured in 30- to 60-μm-thick slices in eight oak trees (Quercus petraea). Canopy W(g) was simulated using a physiologically process-based model. High between-tree variability was observed in the seasonal variations of intra-ring δ(13) C. Six trees showed significant positive correlations between W(g) calculated from intra-ring δ(13) C and canopy W(g) averaged over several days during latewood formation. These results suggest that latewood is a seasonal recorder of W(g) trends, with a temporal lag corresponding to the mixing time of sugars in the phloem. These six trees also showed significant negative correlations between photosynthetic discrimination Δ calculated from intra-ring δ(13) C, and ecosystem W(t), during latewood formation. Despite the observed between-tree variability, these results indicate that intra-ring δ(13) C can be used to access seasonal variations in past W(t).     

Effect of climatic variability on δ13C and tree-ring growth in piñon pine (Pinus edulis)

Understanding the response of long-lived species to natural climatic variability at multiple scales is a prerequisite for forecasting ecosystem responses to global climate change. This study investigated the response of piñon pine (Pinus edulis) to natural climatic variability using information on physiology and growth as recorded in leaves and tree rings. δ¹³C of annual leaf cohorts (δ¹³C_leaf) and tree rings (δ¹³C_ring) were measured at an ecotonal/xeric site and a mid-range/mesic site. Ring width indices (RWI) were used to estimate annual growth of individual trees. Relationships between seasonal and annual climate parameters and δ¹³C and growth were investigated. δ¹³C–climate relationships were stronger for δ¹³C_leaf than for δ¹³C_ring especially at the xeric site. The mean monthly maximum summer temperatures over May through September (summer T _max) had the strongest influence on δ¹³C_leaf. There was a strong negative relationship between RWI with summer T _max and a strong positive relationship between RWI with October to October precipitation (water–year PPN) at both sites. This suggests that piñon pine populations could be vulnerable to decreased growth and, perhaps mortality, in response to warmer, drier conditions predicted by models of global climate change.     

Carbon isotope discrimination and stomatal responses of mature Pinus sylvestris L. trees exposed in situ for three years to elevated CO2 and temperature

The Climate Change Experiment (CLIMEX) is a unique large scale facility in which an entire undisturbed catchment of boreal vegetation has been exposed to elevated CO₂ (560 ppm) and temperature (+3°C summer, +5°C winter) for the past three years with all the soil-plant-atmosphere linkages intact. Here, carbon isotope composition and stomatal density have been analysed from sequential year classes of needles of mature Scots pine trees (Pinus sylvestris L.) to investigate the response of time-integrated water-use efficiency (UWE) and stomatal density to CO₂ enrichment and climate change. Carbon isotope discrimination decreased and WUE increased in cohorts of needles developing under increased CO₂ and temperature, compared to needles on the same trees developing in pretreatment years. Mid-season instantaneous gas exchange, measured on the same trees for the past four years, indicated that these responses resulted from higher needle photosynthetic rates and reduced stomatal conductance. Needles of P. sylvestris developing under increased CO₂ and temperature had consistently lower stomatal densities than their ambient grown counterparts on the same trees. The stomatal density of P. sylvestris needles was inversely correlated with δ¹³C-derived WUE, implying some effect of this morphological response on leaf gas exchange. Future atmospheric CO₂ and temperature increases are therefore likely to improve the water economy of P. sylvestris, at least at the scale of individual needles, by affecting stomatal density and gas exchange processes.     

Comparison of water-use efficiency and internal leaf carbon dioxide concentration in juvenile leaves and phyllodes of Acacia koa (leguminosae) from Hawaii,estimated by two methods

To gain further insight into comparative ecophysiology of different leaf types, water-use efficiency (WUE) and internal leaf carbon dioxide concentration (C_i) were estimated in the field for juvenile leaves and phyllodes of Acacia koa by carbon dioxide and water vapor exchange using a closed system infrared gas analyzer and humidity sensor, and by δ¹³C measurements. Both methods indicate that phyllodes possessed higher WUE and lower C_i than juvenile leaves. However, C_i predicted by δ¹³C for juvenile leaves and phyllodes was lower than the average gas exchange estimated values of C_i and closer to minimal gas exchange estimated values of C_i. It is suggested that δ¹³C may be influenced more during times of maximal assimilation and leaf expansion than during maintenance.     

Stable isotopic (δ13C and δ18O) signatures of biogenic calcretes marking discontinuity surfaces: a case study from Upper Cretaceous carbonates of central Dalmatia and eastern Istria,Croatia

Biogenic calcretes associated with a regional Cretaceous to Paleogene subaerial unconformity and an intraformational composite (polygenic) surface in Upper Cretaceous intra-platform peritidal successions in central Dalmatia and eastern Istria, Croatia (Adriatic-Dinaridic Carbonate Platform), were analyzed for their δ¹³C and δ¹⁸O signatures in order to provide insight into the conditions of subaerial exposure and calcrete development. The distinctly negative δ¹³C signatures of biogenic calcretes marking the regional subaerial unconformity differ considerably from the δ¹³C values of the host marine limestones. This indicates carbon isotope exchange of primary marine CaCO₃ with CO₂ released by root and rhizomicrobial respiration and subsequent precipitation of pedogenic calcrete. The range of δ¹³C (from ?13.1 to ?8.2 ‰ Vienna PeeDee Belemnite standard, VPDB) and δ¹⁸O (from ?10.1 to ?6.1 ‰ VPDB) values of calcretes are similar to those reported from calcretes elsewhere, and the δ¹³C values of biogenic calcretes with typical Microcodium aggregates (?13.1 to ?12.3 ‰ VPDB) at the ?ibenik locality are very close to, or at the lower limit of, values for soil carbonates formed in isotopic equilibrium with soil CO₂. These values are expected for authigenic pedogenic carbonates formed under the influence of C₃ plant communities, without influence from heavier carbon from pre-existing carbonate and lack of input of atmospheric CO₂. Such low δ¹³C values support the interpretation of Microcodium aggregates as being precipitated under a direct biological control within the soil, although the relationship between formation mechanisms and stable isotope signatures of Microcodium needs further investigation. The δ¹³C values (?4.4 to ?3.6 ‰ VPDB) of rhizogenic calcretes formed inside firmground Thalassinoides burrows of the composite surface at the ?ibenik locality are more negative than the δ¹³C values of the host marine limestones, which confirms that the composite surface went through a phase of meteoric pedo(dia)genesis. However, the overall δ¹³C values of calcretes are less negative than expected, which might reflect contamination from associated primary marine carbonate. This study represents the first detailed stable isotope investigation of calcretes from carbonate successions of the External Dinarides, and the results may be applied to discontinuities present in other shallow-water carbonate rock successions.     

Relative humidity- and ABA-induced variation in carbon and oxygen isotope ratios of cotton leaves

Cotton (Gossypium hirsutum L. cv. CS50) plants were grown at two levels of relative humidity (RH) and sprayed daily with abscisic acid (ABA) at four concentrations. Plants grown at lower humidity had higher transpiration rates, lower leaf temperatures and lower stomatal conductance. Plant biomass was also reduced at low humidity. Within each humidity environment, increasing ABA concentration generally reduced stomatal conductance, evaporation rates, superficial leaf density and plant biomass, and increased leaf temperature and specific leaf area. As expected, decreased stomatal conductance resulted in decreased carbon isotope discrimination in leaf material ( Δ ¹³C_l). Plants grown at low humidity were more enriched in ¹⁸O than those grown at high RH, as theory predicts. Within each humidity environment, increasing ABA concentration increased oxygen isotope enrichment of leaf cellulose ( Δ ¹⁸O_c) and whole‐leaf tissue ( Δ ¹⁸O_l). Values of Δ ¹³C_l and Δ ¹⁸O_l predicted by theoretical models were close to those observed, accounting for 79% of the measured variation in Δ ¹³C_l and 95% of the measured variation in Δ ¹⁸O_l. Supporting theory, Δ ¹³C_l and Δ ¹⁸O_l in whole‐leaf tissue were negatively related.     

Stable nitrogen and carbon isotope ratios in wild native honeybees: the influence of land use and climate

The eastern hive bee Apis cerana is a major honeybee species in Asia providing numerous ecosystem services. Understanding how much the honeybees depend on natural and human-influenced plants and landscapes in different climates is important could contribute to evaluate how wild honeybees use food resources and to measure the ecosystem services. We investigated the effects of land use and climate changes on stable nitrogen and carbon isotope ratios in wild populations of A. cerana. In populations from 139 individual sites throughout Japan, we measured nitrogen (δ¹⁵N) and carbon (δ¹³C) stable isotope ratios and analyzed the effects of land use and climate. Our results showed that forested areas and annual precipitation had significant effects on δ¹⁵N, and that paddy fields and urban areas had significant effects on δ¹³C. These results suggest that A. cerana sensibly uses available food resources in the various environments and that stable nitrogen and carbon isotope ratios clearly reflect the effects of land use and climate changes on the populations of A. cerana. Thus, stable nitrogen and carbon isotope ratios in A. cerana, which widely distributes in Asia, can be used as indicators of the environments, such as land use and climate, of an area within its foraging range.