Lack of photosynthetic or stomatal regulation after 9 years of elevated [CO2] and 4 years of soil warming in two conifer species at the alpine treeline.

Alpine treelines are temperature-limited vegetation boundaries. Understanding the effects of elevated [CO2 ] and warming on CO2 and H2 O gas exchange may help predict responses of treelines to global change. We measured needle gas exchange of Larix decidua Mill. and Pinus mugo ssp. uncinata DC trees after 9 years of free air CO2 enrichment (575 µmol mol(-1) ) and 4 years of soil warming (+4 °C) and analysed δ(13) C and δ(18) O values of needles and tree rings. Tree needles under elevated [CO2 ] showed neither nitrogen limitation nor end-product inhibition, and no down-regulation of maximal photosynthetic rate (Amax ) was found. Both tree species showed increased net photosynthetic rates (An ) under elevated [CO2 ] (L. decidua: +39%; P. mugo: +35%). Stomatal conductance (gH2O ) was insensitive to changes in [CO2 ], thus transpiration rates remained unchanged and intrinsic water-use efficiency (iWUE) increased due to higher An . Soil warming affected neither An nor gH2O . Unresponsiveness of gH2O to [CO2 ] and warming was confirmed by δ(18) O needle and tree ring values. Consequently, under sufficient water supply, elevated [CO2 ] induced sustained enhancement in An and lead to increased C inputs into this ecosystem, while soil warming hardly affected gas exchange of L. decidua and P. mugo at the alpine treeline.

Soil warming alters microbial substrate use in alpine soils.

Will warming lead to an increased use of older soil organic carbon (SOC) by microbial communities, thereby inducing C losses from C-rich alpine soils? We studied soil microbial community composition, activity, and substrate use after 3 and 4 years of soil warming (+4 °C, 2007-2010) at the alpine treeline in Switzerland. The warming experiment was nested in a free air CO2 enrichment experiment using depleted (13)CO2 (δ(13)C = -30‰, 2001-2009). We traced this depleted (13)C label in phospholipid fatty acids (PLFA) of the organic layer (0-5 cm soil depth) and in C mineralized from root-free soils to distinguish substrate ages used by soil microorganisms: fixed before 2001 ('old'), from 2001 to 2009 ('new') or in 2010 ('recent'). Warming induced a sustained stimulation of soil respiration (+38%) without decline in mineralizable SOC. PLFA concentrations did not reveal changes in microbial community composition due to soil warming, but soil microbial metabolic activity was stimulated (+66%). Warming decreased the amount of new and recent C in the fungal biomarker 18:2ω6,9 and the amount of new C mineralized from root-free soils, implying a shift in microbial substrate use toward a greater use of old SOC. This shift in substrate use could indicate an imbalance between C inputs and outputs, which could eventually decrease SOC storage in this alpine ecosystem.

Tracing fresh assimilates through Larix decidua exposed to elevated CO2 and soil warming at the alpine treeline using compound-specific stable isotope analysis.

How will carbon source-sink relations of 35-yr-old larch trees (Larix decidua) at the alpine treeline respond to changes in atmospheric CO(2) and climate? We evaluated the effects of previously elevated CO(2) concentrations (9 yr, 580 ppm, ended the previous season) and ongoing soil warming (4 yr, + 4°C). Larch branches were pulse labeled (50 at% (13)CO(2)) in July 2010 to trace fresh assimilates through tissues (buds, needles, bark and wood) and non-structural carbon compounds (NCC; starch, lipids, individual sugars) using compound-specific isotope analysis. Nine years of elevated CO(2) did not lead to increased NCC concentrations, nor did soil warming increase NCC transfer velocities. By contrast, we found slower transfer velocities and higher NCC concentrations than reported in the literature for lowland larch. As a result of low dilution with older carbon, sucrose and glucose showed the highest maximum (13)C labels, whereas labels were lower for starch, lipids and pinitol. Label residence times in needles were shorter for sucrose and starch (c. 2 d) than for glucose (c. 6 d). Although our treatments showed no persistent effect on larch carbon relations, low temperature at high altitudes clearly induced a limitation of sink activities (growth, respiration, root exudation), expressed in slower carbon transfer and higher NCC concentrations.

An alpine treeline in a carbon dioxide-rich world: synthesis of a nine-year free-air carbon dioxide enrichment study.

We evaluated the impacts of elevated CO2 in a treeline ecosystem in the Swiss Alps in a 9-year free-air CO2 enrichment (FACE) study. We present new data and synthesize plant and soil results from the entire experimental period. Light-saturated photosynthesis (A max) of ca. 35-year-old Larix decidua and Pinus uncinata was stimulated by elevated CO2 throughout the experiment. Slight down-regulation of photosynthesis in Pinus was consistent with starch accumulation in needle tissue. Above-ground growth responses differed between tree species, with a 33 % mean annual stimulation in Larix but no response in Pinus. Species-specific CO2 responses also occurred for abundant dwarf shrub species in the understorey, where Vaccinium myrtillus showed a sustained shoot growth enhancement (+11 %) that was not apparent for Vaccinium gaultherioides or Empetrum hermaphroditum. Below ground, CO2 enrichment did not stimulate fine root or mycorrhizal mycelium growth, but increased CO2 effluxes from the soil (+24 %) indicated that enhanced C assimilation was partially offset by greater respiratory losses. The dissolved organic C (DOC) concentration in soil solutions was consistently higher under elevated CO2 (+14 %), suggesting accelerated soil organic matter turnover. CO2 enrichment hardly affected the C-N balance in plants and soil, with unaltered soil total or mineral N concentrations and little impact on plant leaf N concentration or the stable N isotope ratio. Sustained differences in plant species growth responses suggest future shifts in species composition with atmospheric change. Consistently increased C fixation, soil respiration and DOC production over 9 years of CO2 enrichment provide clear evidence for accelerated C cycling with no apparent consequences on the N cycle in this treeline ecosystem.

Evaluation of a liquid chromatography method for compound-specific δ13C analysis of plant carbohydrates in alkaline media.

RATIONALE: Isotope analysis of carbohydrates is important for improved understanding of plant carbon metabolism and plant physiological response to the environment. High-performance liquid chromatography/isotope ratio mass spectrometry (HPLC/IRMS) for direct compound-specific δ(13)C measurements of soluble carbohydrates has recently been developed, but the still challenging sample preparation and the fact that no single method is capable of separating all compounds of interest hinder its wide-spread application. Here we tested in detail a chromatography method in alkaline media. METHODS: We examined the most suitable chromatographic conditions for HPLC/IRMS analysis of carbohydrates in aqueous conifer needle extracts using a CarboPac PA20 anion-exchange column with NaOH eluent, paying specific attention to compound yields, carbon isotope fractionation processes and the reproducibility of the method. Furthermore, we adapted and calibrated sample preparation methods for HPLC/IRMS analysis. OnGuard II cartridges were used for sample purification. RESULTS: Good peak separation and highly linear and reproducible concentration and δ(13)C measurements were obtained. The alkaline eluent was observed to induce isomerization of hexoses, detected as reduced yields and (13)C fractionation of the affected compounds. A reproducible pre-purification method providing ~100% yield for the carbohydrate compounds of interest was calibrated. CONCLUSIONS: The good level of peak separation obtained in this study is reflected in the good precision and linearity of concentration and δ(13)C results. The data provided crucial information on the behaviour of sugars in LC analysis with alkaline media. The observations highlight the importance for the application of compound-matched standard solution for the detection and correction of instrumental biases in concentration and δ(13)C analysis performed under identical chromatographic conditions. The calibrated pre-purification method is well suited for studies with complex matrices that disable the use of a spiked internal standard for the detection of procedural losses.

The influence of increasing temperature and CO2 concentration on recent growth of old-growth larch: contrasting responses at leaf and stem processes derived from tree-ring width and stable isotopes.

Time series of tree-ring growth show significant increases since the early 1970s at the alpine tree line, with simultaneously increasing temperatures and atmospheric CO2 concentration. For a comprehensive understanding of this growth change, the physiological response patterns at both the leaf and stem level need to be separately analyzed and identified, and can be retrieved from tree-ring growth and isotope (δ13C, δ18O) series. In this study, we assessed the relative contribution of environmental factors to interannual tree-ring variability by multivariate linear mixed-effects models and the dual isotope approach on a dataset of tree-ring records of ~400-year-old larch (Larix decidua Mill.) from a non-water-limited high-elevation site in the Swiss Alps. The models suggest that summer temperatures and the recent lack of larch budmoth outbreaks were most important for explaining growth variations and trends, while a significant direct effect of the continuously increasing CO2 concentration could not be confirmed. In contrast, δ13C and δ18O, which are strongly influenced by fractionation changes in the leaf, clearly reflected the impact of air humidity (precipitation and vapor pressure deficit) and CO2 concentration: the increase in (δ13C-derived) intrinsic water-use efficiency over the second half of the 20th century suggests an increase in carbon assimilation as a result of enhanced CO2 concentration. The tree-ring δ18O largely reflected recent precipitation as source water, thus indicating a low variability in stomatal conductance, which was confirmed by the dual isotope approach. These leaf-level effects were not reflected in stem growth as they may have been masked by the temperature-caused growth limitation controlling the allocation of increased amounts of photosynthates into wood growth. Our approach demonstrates that the identification of different roles of environmental factors on leaf and stem processes helps to improve the assessment of site-specific changes of carbon fluxes and growth performance under future environmental conditions.