Soil evaporation and organic matter turnover in the Sub-Taiga and Forest-Steppe of southwest Siberia.

Southwest Siberia encompasses the forest-steppe and sub-taiga climatic zones and has historically been utilized for agriculture. Coinciding with predicted changes in climate for the region is the pressure of agricultural development; however, a characterization of the soil water and carbon dynamics is lacking. We assessed current soil water properties and soil organic carbon turnover in forests and grasslands for two sites that span the forest steppe and sub-taiga bioclimatic zones. Soil evaporation was 0.62 ± 0.17 mm d-1 (mean ± standard error) in grasslands and 0.45 ± 0.08 mm d-1 in the forests of the forest-steppe site. Evaporation at the sub-taiga site was 1.80 ± 1.70 mm d-1 in grasslands and 0.96 ± 0.05 mm d-1 in forest plots. Evaporation was significantly greater at the sub-taiga site than the forest-steppe site. The density of fine roots explained the soil water isotopic patterns between vegetation types and sites. We found soil organic matter turnover to be three times faster in the sub-taiga site than in the forest-steppe site. Our results show that while climate factors, in particular snow levels, between the two sites are drivers for water and carbon cycles, site level hydrology, soil characteristics, and vegetation directly interact to influence the water and carbon dynamics.

Genotypic variability enhances the reproducibility of an ecological study.

Many scientific disciplines are currently experiencing a 'reproducibility crisis' because numerous scientific findings cannot be repeated consistently. A novel but controversial hypothesis postulates that stringent levels of environmental and biotic standardization in experimental studies reduce reproducibility by amplifying the impacts of laboratory-specific environmental factors not accounted for in study designs. A corollary to this hypothesis is that a deliberate introduction of controlled systematic variability (CSV) in experimental designs may lead to increased reproducibility. To test this hypothesis, we had 14 European laboratories run a simple microcosm experiment using grass (Brachypodium distachyon L.) monocultures and grass and legume (Medicago truncatula Gaertn.) mixtures. Each laboratory introduced environmental and genotypic CSV within and among replicated microcosms established in either growth chambers (with stringent control of environmental conditions) or glasshouses (with more variable environmental conditions). The introduction of genotypic CSV led to 18% lower among-laboratory variability in growth chambers, indicating increased reproducibility, but had no significant effect in glasshouses where reproducibility was generally lower. Environmental CSV had little effect on reproducibility. Although there are multiple causes for the 'reproducibility crisis', deliberately including genetic variability may be a simple solution for increasing the reproducibility of ecological studies performed under stringently controlled environmental conditions.

On the contributions of photorespiration and compartmentation to the contrasting intramolecular 2H profiles of C3 and C4 plant sugars.

Compartmentation of C4 photosynthetic biochemistry into bundle sheath (BS) and mesophyll (M) cells, and photorespiration in C3 plants is predicted to have hydrogen isotopic consequences for metabolites at both molecular and site-specific levels. Molecular-level evidence was recently reported (Zhou et al., 2016), but evidence at the site-specific level is still lacking. We propose that such evidence exists in the contrasting 2H distribution profiles of glucose samples from naturally grown C3, C4 and CAM plants: photorespiration contributes to the relative 2H enrichment in H5 and relative 2H depletion in H1 & H6 (the average of the two pro-chiral Hs and in particular H6,pro-R) in C3 glucose, while 2H-enriched C3 mesophyll cellular (chloroplastic) water most likely contributes to the enrichment at H4; export of (transferable hydrogen atoms of) NADPH from C4 mesophyll cells to bundle sheath cells (via the malate shuttle) and incorporation of 2H-relatively unenriched BS cellular water contribute to the relative depletion of H4 & H5 respectively; shuttling of triose-phosphates (PGA: phosphoglycerate dand DHAP: dihydroacetone phosphate) between C4 bundle sheath and mesophyll cells contributes to the relative enrichment in H1 & H6 (in particular H6,pro-R) in C4 glucose.

Top canopy nitrogen allocation linked to increased grassland carbon uptake in stands of varying species richness.

Models predict that vertical gradients of foliar nitrogen (N) allocation, increasing from bottom to top of plant canopies, emerge as a plastic response to optimise N utilisation for carbon assimilation. While this mechanism has been well documented in monocultures, its relevance for mixed stands of varying species richness remains poorly understood. We used 21 naturally assembled grassland communities to analyse the gradients of N in the canopy using N allocation coefficients (K N ) estimated from the distribution of N per foliar surface area (KN-F) and ground surface area (KN-G). We tested whether: 1) increasing plant species richness leads to more pronounced N gradients as indicated by higher K N -values, 2) K N is a good predictor of instantaneous net ecosystem CO2 exchange and 3) functional diversity of leaf N concentration as estimated by Rao's Q quadratic diversity metric is a good proxy of K N . Our results show a negative (for KN-G) or no relationship (for KN-F) between species richness and canopy N distribution, but emphasize a link (positive relationship) between more foliar N per ground surface area in the upper layers of the canopy (i.e. under higher KN-G) and ecosystem CO2 uptake. Rao's Q was not a good proxy for either K N .

Variation in short-term and long-term responses of photosynthesis and isoprenoid-mediated photoprotection to soil water availability in four Douglas-fir provenances.

For long-lived forest tree species, the understanding of intraspecific variation among populations and their response to water availability can reveal their ability to cope with and adapt to climate change. Dissipation of excess excitation energy, mediated by photoprotective isoprenoids, is an important defense mechanism against drought and high light when photosynthesis is hampered. We used 50-year-old Douglas-fir trees of four provenances at two common garden experiments to characterize provenance-specific variation in photosynthesis and photoprotective mechanisms mediated by essential and non-essential isoprenoids in response to soil water availability and solar radiation. All provenances revealed uniform photoprotective responses to high solar radiation, including increased de-epoxidation of photoprotective xanthophyll cycle pigments and enhanced emission of volatile monoterpenes. In contrast, we observed differences between provenances in response to drought, where provenances sustaining higher CO2 assimilation rates also revealed increased water-use efficiency, carotenoid-chlorophyll ratios, pools of xanthophyll cycle pigments, ß-carotene and stored monoterpenes. Our results demonstrate that local adaptation to contrasting habitats affected chlorophyll-carotenoid ratios, pool sizes of photoprotective xanthophylls, ß-carotene, and stored volatile isoprenoids. We conclude that intraspecific variation in isoprenoid-mediated photoprotective mechanisms contributes to the adaptive potential of Douglas-fir provenances to climate change.

Land-use and hydroperiod affect kettle hole sediment carbon and nitrogen biogeochemistry.

Kettle holes are glaciofluvially created depressional wetlands that collect organic matter (OM) and nutrients from their surrounding catchment. Kettle holes mostly undergo pronounced wet-dry cycles. Fluctuations in water table, land-use, and management can affect sediment biogeochemical transformations and perhaps threaten the carbon stocks of these unique ecosystems. We investigated sediment and water of 51 kettle holes in NE Germany that differ in hydroperiod (i.e. the duration of the wet period of a kettle hole) and land-use. Our objectives were 1) to test if hydroperiod and land management were imprinted on the isotopic values (δ13C, δ15N) and C:N ratios of the sediment OM, and 2) to characterize water loss dynamics and kettle hole-groundwater connectivity by measuring the stable δ18O and δD isotope values of kettle hole water over several years. We found the uppermost sediment layer reflected recent OM inputs and short-term processes in the catchment, including land-use and management effects. Deeper sediments recorded the degree to which OM is processed within the kettle hole related to the hydroperiod. We see clear indications for the effects of wet-dry cycles for all kettle holes, which can lead to the encroachment of terrestrial plants. We found that the magnitude of evaporation depended on the year, season, and land-use type, that kettle holes are temporarily coupled to shallow ground water, and, as such, kettle holes are described best as partially-closed to open systems.

Circadian rhythms have significant effects on leaf-to-canopy scale gas exchange under field conditions.

BACKGROUND: Molecular clocks drive oscillations in leaf photosynthesis, stomatal conductance, and other cell and leaf-level processes over ~24 h under controlled laboratory conditions. The influence of such circadian regulation over whole-canopy fluxes remains uncertain; diurnal CO2 and H2O vapor flux dynamics in the field are currently interpreted as resulting almost exclusively from direct physiological responses to variations in light, temperature and other environmental factors. We tested whether circadian regulation would affect plant and canopy gas exchange at the Montpellier European Ecotron. Canopy and leaf-level fluxes were constantly monitored under field-like environmental conditions, and under constant environmental conditions (no variation in temperature, radiation, or other environmental cues). RESULTS: We show direct experimental evidence at canopy scales of the circadian regulation of daytime gas exchange: 20-79 % of the daily variation range in CO2 and H2O fluxes occurred under circadian entrainment in canopies of an annual herb (bean) and of a perennial shrub (cotton). We also observed that considering circadian regulation improved performance by 8-17 % in commonly used stomatal conductance models. CONCLUSIONS: Our results show that circadian controls affect diurnal CO2 and H2O flux patterns in entire canopies in field-like conditions, and its consideration significantly improves model performance. Circadian controls act as a 'memory' of the past conditions experienced by the plant, which synchronizes metabolism across entire plant canopies.

Forest understory plant and soil microbial response to an experimentally induced drought and heat-pulse event: the importance of maintaining the continuum.

Drought duration and intensity are expected to increase with global climate change. How changes in water availability and temperature affect the combined plant-soil-microorganism response remains uncertain. We excavated soil monoliths from a beech (Fagus sylvatica L.) forest, thus keeping the understory plant-microbe communities intact, imposed an extreme climate event, consisting of drought and/or a single heat-pulse event, and followed microbial community dynamics over a time period of 28 days. During the treatment, we labeled the canopy with (13) CO2 with the goal of (i) determining the strength of plant-microbe carbon linkages under control, drought, heat and heat-drought treatments and (ii) characterizing microbial groups that are tightly linked to the plant-soil carbon continuum based on (13) C-labeled PLFAs. Additionally, we used 16S rRNA sequencing of bacteria from the Ah horizon to determine the short-term changes in the active microbial community. The treatments did not sever within-plant transport over the experiment, and carbon sinks belowground were still active. Based on the relative distribution of labeled carbon to roots and microbial PLFAs, we determined that soil microbes appear to have a stronger carbon sink strength during environmental stress. High-throughput sequencing of the 16S rRNA revealed multiple trajectories in microbial community shifts within the different treatments. Heat in combination with drought had a clear negative effect on microbial diversity and resulted in a distinct shift in the microbial community structure that also corresponded to the lowest level of label found in the PLFAs. Hence, the strongest changes in microbial abundances occurred in the heat-drought treatment where plants were most severely affected. Our study suggests that many of the shifts in the microbial communities that we might expect from extreme environmental stress will result from the plant-soil-microbial dynamics rather than from direct effects of drought and heat on soil microbes alone.

Water level changes affect carbon turnover and microbial community composition in lake sediments.

Due to climate change, many lakes in Europe will be subject to higher variability of hydrological characteristics in their littoral zones. These different hydrological regimes might affect the use of allochthonous and autochthonous carbon sources. We used sandy sediment microcosms to examine the effects of different hydrological regimes (wet, desiccating, and wet-desiccation cycles) on carbon turnover. (13)C-labelled particulate organic carbon was used to trace and estimate carbon uptake into bacterial biomass (via phospholipid fatty acids) and respiration. Microbial community changes were monitored by combining DNA- and RNA-based real-time PCR quantification and terminal restriction fragment length polymorphism (T-RFLP) analysis of 16S rRNA. The shifting hydrological regimes in the sediment primarily caused two linked microbial effects: changes in the use of available organic carbon and community composition changes. Drying sediments yielded the highest CO2 emission rates, whereas hydrological shifts increased the uptake of allochthonous organic carbon for respiration. T-RFLP patterns demonstrated that only the most extreme hydrological changes induced a significant shift in the active and total bacterial communities. As current scenarios of climate change predict an increase of drought events, frequent variations of the hydrological regimes of many lake littoral zones in central Europe are anticipated. Based on the results of our study, this phenomenon may increase the intensity and amplitude in rates of allochthonous organic carbon uptake and CO2 emissions.

Soil bacterial community structure responses to precipitation reduction and forest management in forest ecosystems across Germany.

Soil microbial communities play an important role in forest ecosystem functioning, but how climate change will affect the community composition and consequently bacterial functions is poorly understood. We assessed the effects of reduced precipitation with the aim of simulating realistic future drought conditions for one growing season on the bacterial community and its relation to soil properties and forest management. We manipulated precipitation in beech and conifer forest plots managed at different levels of intensity in three different regions across Germany. The precipitation reduction decreased soil water content across the growing season by between 2 to 8% depending on plot and region. T-RFLP analysis and pyrosequencing of the 16S rRNA gene were used to study the total soil bacterial community and its active members after six months of precipitation reduction. The effect of reduced precipitation on the total bacterial community structure was negligible while significant effects could be observed for the active bacteria. However, the effect was secondary to the stronger influence of specific soil characteristics across the three regions and management selection of overstorey tree species and their respective understorey vegetation. The impact of reduced precipitation differed between the studied plots; however, we could not determine the particular parameters being able to modify the response of the active bacterial community among plots. We conclude that the moderate drought induced by the precipitation manipulation treatment started to affect the active but not the total bacterial community, which points to an adequate resistance of the soil microbial system over one growing season.

Allocate carbon for a reason: priorities are reflected in the ¹³C/¹²C ratios of plant lipids synthesized via three independent biosynthetic pathways.

It has long been theorized that carbon allocation, in addition to the carbon source and to kinetic isotopic effects associated with a particular lipid biosynthetic pathway, plays an important role in shaping the carbon isotopic composition ((13)C/(12)C) of lipids (Park and Epstein, 1961). If the latter two factors are properly constrained, valuable information about carbon allocation during lipid biosynthesis can be obtained from carbon isotope measurements. Published work of Chikaraishi et al. (2004) showed that leaf lipids isotopic shifts from bulk leaf tissue Δδ(13)C(bk-lp) (defined as δ(13)C(bulkleaftissue)-δ(13)C(lipid)) are pathway dependent: the acetogenic (ACT) pathway synthesizing fatty lipids has the largest isotopic shift, the mevalonic acid (MVA) pathway synthesizing sterols the lowest and the phytol synthesizing 1-deoxy-D-xylulose 5-phosphate (DXP) pathway gives intermediate values. The differences in Δδ(13)C(bk-lp) between C3 and C4 plants Δδ(13)C(bk-lp,C4-C3) are also pathway-dependent: Δδ(13)C(ACT)(bk-lp,C4-C3) > Δδ(13)C(DXP(bk-lp,C4-C3) > Δδ(13)C(MVA)(bk-lp,C4-C3). These pathway-dependent differences have been interpreted as resulting from kinetic isotopic effect differences of key but unspecified biochemical reactions involved in lipids biosynthesis between C3 and C4 plants. After quantitatively considering isotopic shifts caused by (dark) respiration, export-of-carbon (to sink tissues) and photorespiration, we propose that the pathway-specific differences Δδ(13)C(bk-lp,C4-C3) can be successfully explained by C4-C3 carbon allocation (flux) differences with greatest flux into the ACT pathway and lowest into the MVA pathways (when flux is higher, isotopic shift relative to source is smaller). Highest carbon allocation to the ACT pathway appears to be tied to the most stringent role of water-loss-minimization by leaf waxes (composed mainly of fatty lipids) while the lowest carbon allocation to the MVA pathway can be largely explained by the fact that sterols act as regulatory hormones and membrane fluidity modulators in rather low concentrations.

Douglas-fir seedlings exhibit metabolic responses to increased temperature and atmospheric drought.

In the future, periods of strongly increased temperature in concert with drought (heat waves) will have potentially detrimental effects on trees and forests in Central Europe. Norway spruce might be at risk in the future climate of Central Europe. However, Douglas-fir is often discussed as an alternative for the drought and heat sensitive Norway spruce, because some provenances are considered to be well adapted to drier and warmer conditions. In this study, we identified the physiological and growth responses of seedlings from two different Douglas-fir provenances to increased temperature and atmospheric drought during a period of 92 days. We analysed (i) plant biomass, (ii) carbon stable isotope composition as an indicator for time integrated intrinsic water use efficiency, (iii) apparent respiratory carbon isotope fractionation as well as (iv) the profile of polar low molecular metabolites. Plant biomass was only slightly affected by increased temperatures and atmospheric drought but the more negative apparent respiratory fractionation indicated a temperature-dependent decrease in the commitment of substrate to the tricarboxylic acid cycle. The metabolite profile revealed that the simulated heat wave induced a switch in stress protecting compounds from proline to polyols. We conclude that metabolic acclimation successfully contributes to maintain functioning and physiological activity in seedlings of both Douglas-fir provenances under conditions that are expected during heat waves (i.e. elevated temperatures and atmospheric drought). Douglas-fir might be a potentially important tree species for forestry in Central Europe under changing climatic conditions.

Drought response of mesophyll conductance in forest understory species--impacts on water-use efficiency and interactions with leaf water movement.

Regulation of stomatal (gs ) and mesophyll conductance (gm ) is an efficient means for optimizing the relationship between water loss and carbon uptake in plants. We assessed water-use efficiency (WUE)-based drought adaptation strategies with respect to mesophyll conductance of different functional plant groups of the forest understory. Moreover we aimed at assessing the mechanisms of and interactions between water and CO2 conductance in the mesophyll. The facts that an increase in WUE was observed only in the two species that increased gm in response to moderate drought, and that over all five species examined, changes in mesophyll conductance were significantly correlated with the drought-induced change in WUE, proves the importance of gm in optimizing resource use under water restriction. There was no clear correlation of mesophyll CO2 conductance and the tortuosity of water movement in the leaf across the five species in the control and drought treatments. This points either to different main pathways for CO2 and water in the mesophyll either to different regulation of a common pathway.

The oxygen isotope enrichment of leaf-exported assimilates--does it always reflect lamina leaf water enrichment?

The oxygen stable isotope composition of plant organic matter (OM) (particularly of wood and cellulose in the tree ring archive) is valuable in studies of plant-climate interaction, but there is a lack of information on the transfer of the isotope signal from the leaf to heterotrophic tissues. We studied the oxygen isotopic composition and its enrichment above source water of leaf water over diel courses in five tree species covering a broad range of life forms. We tracked the transfer of the isotopic signal to leaf water-soluble OM and further to phloem-transported OM. Observed leaf water evaporative enrichment was consistent with values predicted from mechanistic models taking into account nonsteady-state conditions. While leaf water-soluble OM showed the expected (18)O enrichment in all species, phloem sugars were less enriched than expected from leaf water enrichment in Scots pine (Pinus sylvestris), European larch (Larix decidua) and Alpine ash (Eucalyptus delegatensis). Oxygen atom exchange with nonenriched water during phloem loading and transport, as well as a significant contribution of assimilates from bark photosynthesis, can explain these phloem (18)O enrichment patterns. Our results indicate species-specific uncoupling between the leaf water and the OM oxygen isotope signal, which is important for the interpretation of tree ring data.

The long way down--are carbon and oxygen isotope signals in the tree ring uncoupled from canopy physiological processes?

The carbon (δ(13)C) and oxygen (δ(18)O) stable isotope composition is widely used to obtain information on the linkages between environmental drivers and tree physiology over various time scales. The tree-ring archive can especially be exploited to reconstruct inter- and intra-annual variation of both climate and physiology. There is, however, a lack of information on the processes potentially affecting δ(13)C and δ(18)O on their way from assimilation in the leaf to the tree ring. As a consequence, the aim of this study was to trace the isotope signals in European beech (Fagus sylvatica L.) from leaf water (δ(18)O) and leaf assimilates (δ(13)C and δ(18)O) to tree-ring wood via phloem-transported compounds over a whole growing season. Phloem and leaf samples for δ(13)C and δ(18)O analyses as well as soil water, xylem water, leaf water and atmospheric water vapour samples for δ(18)O analysis were taken approximately every 2 weeks during the growing season of 2007. The δ(13)C and δ(18)O samples from the tree rings were dated intra-annually by monitoring the tree growth with dendrometers. δ(18)O in the phloem organic matter and tree-ring whole wood was not positively related to leaf water evaporative enrichment and δ(18)O of canopy organic matter pools. This finding implies a partial uncoupling of the tree-ring oxygen isotopic signal from canopy physiology. At the same time, internal carbon storage and remobilization physiology most likely prevented δ(13)C in tree-ring whole wood from being closely related to intra-annual variation in environmental drivers. Taking into account the post-photosynthetic isotope fractionation processes resulting in alterations of δ(13)C and δ(18)O not only in the tree ring but also in phloem carbohydrates, as well as the intra-annual timing of changes in the tree internal physiology, might help to better understand the meaning of the tree-ring isotope signal not only intra- but also inter-annually.

Soil moisture effects on the carbon isotope composition of soil respiration.

The carbon isotopic composition (delta(13)C) of recently assimilated plant carbon is known to depend on water-stress, caused either by low soil moisture or by low atmospheric humidity. Air humidity has also been shown to correlate with the delta(13)C of soil respiration, which suggests indirectly that recently fixed photosynthates comprise a substantial component of substrates consumed by soil respiration. However, there are other reasons why the delta(13)CO(2) of soil efflux may change with moisture conditions, which have not received as much attention. Using a combination of greenhouse experiments and modeling, we examined whether moisture can cause changes in fractionation associated with (1) non-steady-state soil CO(2) transport, and (2) heterotrophic soil-respired delta(13)CO(2). In a first experiment, we examined the effects of soil moisture on total respired delta(13)CO(2) by growing Douglas fir seedlings under high and low soil moisture conditions. The measured delta(13)C of soil respiration was 4.7 per thousand more enriched in the low-moisture treatment; however, subsequent investigation with an isotopologue-based gas diffusion model suggested that this result was probably influenced by gas transport effects. A second experiment examined the heterotrophic component of soil respiration by incubating plant-free soils, and showed no change in microbial-respired delta(13)CO(2) across a large moisture range. Our results do not rule out the potential influence of recent photosynthates on soil-respired delta(13)CO(2), but they indicate that the expected impacts of photosynthetic discrimination may be similar in direction and magnitude to those from gas transport-related fractionation. Gas transport-related fractionation may operate as an alternative or an additional factor to photosynthetic discrimination to explain moisture-related variation in soil-respired delta(13)CO(2).

Bias and uncertainty of delta13CO 2 isotopic mixing models.

Patterns in the isotopic signal (stable C isotope composition; delta(13)C) of respiration (delta(13)C(R)) have led to important gains in understanding the C metabolism of many systems. Contained within delta(13)C(R) is a record of the C source mineralized, the metabolic pathway of C and the environmental conditions during which respiration occurred. Because gas samples used for analysis of delta(13)C(R) contain a mixture of CO(2) from respiration and from the atmosphere, two-component mixing models are used to identify delta(13)C(R). Measurement of ecosystem delta(13)C(R), using canopy airspace gas samples, was one of the first applications of mixing models in ecosystem ecology, and thus recommendations and guidelines are based primarily on findings from these studies. However, as mixing models are applied to other experimental conditions these approaches may not be appropriate. For example, the range in [CO(2)] obtained in gas samples from canopy air is generally less than 100 micromol mol(-1), whereas in studies of respiration from soil, foliage or tree stems, the range can span as much as 10,000 micromol mol(-1) and greater. Does this larger range in [CO(2)] influence the precision and accuracy of delta(13)C(R) estimates derived from mixing models? Does the outcome from using different regression approaches and mixing models vary depending on the range of [CO(2)]? Our research addressed these questions using a simulation approach. We found that it is important to distinguish between large (>1,000 micromol mol(-1)) and small (<100 micromol mol(-1)) ranges of CO(2) when applying a mixing model (Keeling plot or Miller-Tans) and regression approach (ordinary least squares or geometric mean regression) combination to isotopic data. The combination of geometric mean regression and the Miller-Tans mixing model provided the most accurate and precise estimate of delta(13)C(R) when the range of CO(2) is >or=1,000 micromol mol(-1).

A laboratory comparison of two methods used to estimate the isotopic composition of soil delta13CO2 efflux at steady state.

The stable isotopic composition of soil (13)CO(2) flux is important for monitoring soil biological and physical processes. While several methods exist to measure the isotopic composition of soil flux, we do not know how effective each method is at achieving this goal. To provide clear evidence of the accuracy of current measurement techniques we created a column filled with quartz sand through which a gas of known isotopic composition (-34.2 per thousand) and concentration (3,000 ppm) diffused for 7 h. We used a static chamber at equilibrium and a soil probe technique to test whether they could identify the isotopic signature of the known gas source. The static chamber is designed to identify the source gas isotopic composition when in equilibrium with the soil gas, and the soil probe method relies on a mixing model of samples withdrawn from three gas wells at different depths to identify the gas source. We sampled from ports installed along the side of the sand column to describe the isotopic and concentration gradient as well as to serve as a control for the soil probe. The soil probe produced similar isotopic and concentration values as the control ports, as well as Keeling intercepts. The static chamber at equilibrium did not identify the source gas but, when applied in a two end-member mixing model, did produce a similar Keeling intercept produced from the control ports. Neither of the methods was able to identify the source gas via the Keeling plot method probably because CO(2) profiles did not reach isotopic steady state. Our results showed that the static chamber at equilibrium should be used only with a Keeling plot approach and that the soil probe is able to provide estimates of uncertainty for the isotopic composition of soil gas as well as information pertinent to the soil profile.