The quandary of sources and sinks of CO2 efflux in tree stems-new insights and future directions.

Stem respiration (RS) substantially contributes to the return of photo assimilated carbon to the atmosphere and, thus, to the tree and ecosystem carbon balance. Stem CO2 efflux (ECO2) is often used as a proxy for RS. However, this metric has often been challenged because of the uncertain origin of CO2 emitted from the stem due to post-respiratory processes. In this Insight, we (i) describe processes affecting the quantification of RS, (ii) review common methodological approaches to quantify and model RS and (iii) develop a research agenda to fill the most relevant knowledge gaps that we identified. Dissolution, transport and accumulation of respired CO2 away from its production site, reassimilation of respired CO2 via stem photosynthesis and the enzyme phosphoenolpyruvate carboxylase, axial CO2 diffusion in the gas phase, shifts in the respiratory substrate and non-respiratory oxygen (O2) consumption are the most relevant processes causing divergence between RS and measured stem gas exchange (ECO2 or O2 influx, IO2). Two common methodological approaches to estimate RS, namely the CO2 mass balance approach and the O2 consumption technique, circumvent some of these processes but have yielded inconsistent results regarding the fate of respired CO2. Stem respiration modelling has recently progressed at the organ and tree levels. However, its implementation in large-scale models, commonly operated from a source-driven perspective, is unlikely to reflect adequate mechanisms. Finally, we propose hypotheses and approaches to advance the knowledge of the stem carbon balance, the role of sap pH on RS, the reassimilation of respired CO2, RS upscaling procedures, large-scale RS modelling and shifts in respiratory metabolism during environmental stress.

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

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

Differences between tree stem CO2 efflux and O2 influx rates cannot be explained by internal CO2 transport or storage in large beech trees.

Tree stem respiration (RS ) is a substantial component of the forest carbon balance. The mass balance approach uses stem CO2 efflux and internal xylem fluxes to sum up RS , while the oxygen-based method assumes O2 influx as a proxy of RS . So far, both approaches have yielded inconsistent results regarding the fate of respired CO2 in tree stems, a major challenge for quantifying forest carbon dynamics. We collected a data set of CO2 efflux, O2 influx, xylem CO2 concentration, sap flow, sap pH, stem temperature, nonstructural carbohydrates concentration and potential phosphoenolpyruvate carboxylase (PEPC) capacity on mature beech trees to identify the sources of differences between approaches. The ratio of CO2 efflux to O2 influx was consistently below unity (0.7) along a 3-m vertical gradient, but internal fluxes did not bridge the gap between influx and efflux, nor did we find evidence for changes in respiratory substrate use. PEPC capacity was comparable with that previously reported in green current-year twigs. Although we could not reconcile differences between approaches, results shed light on the uncertain fate of CO2 respired by parenchyma cells across the sapwood. Unexpected high values of PEPC capacity highlight its potential relevance as a mechanism of local CO2 removal, which merits further research.

The size and the age of the metabolically active carbon in tree roots.

Little is known about the sources and age of C respired by tree roots. Previous research in stems identified two functional pools of non-structural carbohydrates (NSC): an "active" pool supplied directly from canopy photo-assimilates supporting metabolism and a "stored" pool used when fresh C supplies are limited. We compared the C isotope composition of water-soluble NSC and respired CO2 for aspen roots (Populus tremula hybrids) cut off from fresh C supply after stem-girdling or prolonged incubation of excised roots. We used bomb radiocarbon to estimate the time elapsed since C fixation for respired CO2 , water-soluble NSC and structural α-cellulose. While freshly excised roots (mostly <2.9 mm in diameter) respired CO2 fixed <1 year previously, the age increased to 1.6-2.9 year within a week after root excision. Freshly excised roots from trees girdled ~3 months ago had respiration rates and NSC stocks similar to un-girdled trees but respired older C (~1.2 year). We estimate that over 3 months NSC in girdled roots must be replaced 5-7 times by reserves remobilized from root-external sources. Using a mixing model and observed correlations between Δ14 C of water-soluble C and α-cellulose, we estimate ~30% of C is "active" (~5 mg C g-1 ).

Low-cost chamber design for simultaneous CO2 and O2 flux measurements between tree stems and the atmosphere.

Tree stem CO2 efflux is an important component of ecosystem carbon fluxes and has been the focus of many studies. While CO2 efflux can easily be measured, a growing number of studies have shown that it is not identical with actual in situ respiration. Complementing measurements of CO2 flux with simultaneous measurements of O2 flux provides an additional proxy for respiration, and the combination of both fluxes can potentially help getting closer to actual measures of respiratory fluxes. To date, however, the technical challenge to measure relatively small changes in O2 concentration against its high atmospheric background has prevented routine O2 measurements in field applications. Here, we present a new and low-cost field-tested device for autonomous real-time and quasi-continuous long-term measurements of stem respiration by combining CO2 (NDIR-based) and O2 (quenching-based) sensors in a tree stem chamber. Our device operates as a cyclic-closed system and measures changes in both CO2 and O2 concentration within the chamber over time. The device is battery powered with a >1-week power independence, and data acquisition is conveniently achieved by an internal logger. Results from both field and laboratory tests document that our sensors provide reproducible measurements of CO2 and O2 exchange fluxes under varying environmental conditions.

Starch and lipid storage strategies in tropical trees relate to growth and mortality.

Non-structural carbon (NSC) storage (i.e. starch, soluble sugras and lipids) in tree stems play important roles in metabolism and growth. Their spatial distribution in wood may explain species-specific differences in carbon storage dynamics, growth and survival. However, quantitative information on the spatial distribution of starch and lipids in wood is sparse due to methodological limitations. Here we assessed differences in wood NSC and lipid storage between tropical tree species with different growth and mortality rates and contrasting functional types. We measured starch and soluble sugars in wood cores up to 4 cm deep into the stem using standard chemical quantification methods and histological slices stained with Lugol's iodine. We also detected neutral lipids using histological slices stained with Oil-Red-O. The histological method allowed us to group individuals into two categories according to their starch storage strategy: fiber-storing trees and parenchyma-storing trees. The first group had a bigger starch pool, slower growth and lower mortality rates than the second group. Lipid storage was found in wood parenchyma in five species and was related to low mortality rates. The quantification of the spatial distribution of starch and lipids in wood improves our understanding of NSC dynamics in trees and reveals additional dimensions of tree growth and survival strategies.

Frost and drought: Effects of extreme weather events on stem carbon dynamics in a Mediterranean beech forest.

The effects of short-term extreme events on tree functioning and physiology are still rather elusive. European beech is one of the most sensitive species to late frost and water shortage. We investigated the intra-annual C dynamics in stems under such conditions. Wood formation and stem CO2 efflux were monitored in a Mediterranean beech forest for 3 years (2015-2017), including a late frost (2016) and a summer drought (2017). The late frost reduced radial growth and, consequently, the amount of carbon fixed in the stem biomass by 80%. Stem carbon dioxide efflux in 2016 was reduced by 25%, which can be attributed to the reduction of effluxes due to growth respiration. Counter to our expectations, we found no effects of the 2017 summer drought on radial growth and stem carbon efflux. The studied extreme weather events had various effects on tree growth. Even though late spring frost had a strong impact on beech radial growth in the current year, trees fully recovered in the following growing season, indicating high resilience of beech to this stressful event.

Probability distributions of nonstructural carbon ages and transit times provide insights into carbon allocation dynamics of mature trees.

●In trees, the use of nonstructural carbon (NSC) under limiting conditions impacts the age structure of the NSC pools. We compared model predictions of NSC ages and transit times for Pinus halepensis, Acer rubrum and Pinus taeda, to understand differences in carbon (C) storage dynamics in species with different leaf phenology and growth environments. ●We used two C allocation models from the literature to estimate the NSC age and transit time distributions, to simulate C limitation, and to evaluate the sensitivity of the mean ages to changes in allocation fluxes. ●Differences in allocation resulted in different NSC age and transit time distributions. The simulated starvation flattened the NSC age distribution and increased the mean NSC transit time, which can be used to estimate the age of the NSC available and the time it would take to exhaust the reserves. Mean NSC ages and transit times were sensitive to C fluxes in roots and allocation of C from wood storage. ●Our results demonstrate how trees with different storage traits are expected to react differently to starvation. They also provide a probabilistic explanation for the 'last-in, first-out' pattern of NSC mobilization from well-mixed C pools.

Winter's bite: beech trees survive complete defoliation due to spring late-frost damage by mobilizing old C reserves.

Late frost can destroy the photosynthetic apparatus of trees. We hypothesized that this can alter the normal cyclic dynamics of C-reserves in the wood. We measured soluble sugar concentrations and radiocarbon signatures (Δ14 C) of soluble nonstructural carbon (NSC) in woody tissues sampled from a Mediterranean beech forest that was completely defoliated by an exceptional late frost in 2016. We used the bomb radiocarbon approach to estimate the time elapsed since fixation of mobilized soluble sugars. During the leafless period after the frost event, soluble sugar concentrations declined sharply while Δ14 C of NSC increased. This can be explained by the lack of fresh assimilate supply and a mobilization of C from reserve pools. Soluble NSC became increasingly older during the leafless period, with a maximum average age of 5 yr from samples collected 27 d before canopy recovery. Following leaf re-growth, soluble sugar concentrations increased and Δ14 C of soluble NSC decreased, indicating the allocation of new assimilates to the stem soluble sugars pool. These data highlight that beech trees rapidly mobilize reserve C to survive strong source-sink imbalances, for example due to late frost, and show that NSC is a key trait for tree resilience under global change.

14C-Free Carbon Is a Major Contributor to Cellular Biomass in Geochemically Distinct Groundwater of Shallow Sedimentary Bedrock Aquifers.

Despite the global significance of the subsurface biosphere, the degree to which it depends on surface organic carbon (OC) is still poorly understood. Here, we compare stable and radiogenic carbon isotope compositions of microbial phospholipid fatty acids (PLFAs) with those of in situ potential microbial C sources to assess the major C sources for subsurface microorganisms in biogeochemical distinct shallow aquifers (Critical Zone Exploratory, Thuringia Germany). Despite the presence of younger OC, the microbes assimilated 14C-free OC to varying degrees; ~31% in groundwater within the oxic zone, ~47% in an iron reduction zone, and ~70% in a sulfate reduction/anammox zone. The persistence of trace amounts of mature and partially biodegraded hydrocarbons suggested that autochthonous petroleum-derived hydrocarbons were a potential 14C-free C source for heterotrophs in the oxic zone. In this zone, Δ14C values of dissolved inorganic carbon (-366 ± 18‰) and 11MeC16:0 (-283 ± 32‰), an important component in autotrophic nitrite oxidizers, were similar enough to indicate that autotrophy is an important additional C fixation pathway. In anoxic zones, methane as an important C source was unlikely since the 13C-fractionations between the PLFAs and CH4 were inconsistent with kinetic isotope effects associated with methanotrophy. In the sulfate reduction/anammox zone, the strong 14C-depletion of 10MeC16:0 (-942 ± 22‰), a PLFA common in sulfate reducers, indicated that those bacteria were likely to play a critical part in 14C-free sedimentary OC cycling. Results indicated that the 14C-content of microbial biomass in shallow sedimentary aquifers results from complex interactions between abundance and bioavailability of naturally occurring OC, hydrogeology, and specific microbial metabolisms.

Living on borrowed time - Amazonian trees use decade-old storage carbon to survive for months after complete stem girdling.

Nonstructural carbon (NSC) reserves act as buffers to sustain tree activity during periods when carbon (C) assimilation does not meet C demand, but little is known about their age and accessibility; we designed a controlled girdling experiment in the Amazon to study tree survival on NSC reserves. We used bomb-radiocarbon (14 C) to monitor the time elapsed between C fixation and release ('age' of substrates). We simultaneously monitored how the mobilization of reserve C affected δ13 CO2 . Six ungirdled control trees relied almost exclusively on recent assimilates throughout the 17 months of measurement. The Δ14 C of CO2 emitted from the six girdled stems increased significantly over time after girdling, indicating substantial remobilization of storage NSC fixed up to 13-14 yr previously. This remobilization was not accompanied by a consistent change in observed δ13 CO2 . These trees have access to storage pools integrating C accumulated over more than a decade. Remobilization follows a very clear reverse chronological mobilization with younger reserve pools being mobilized first. The lack of a shift in the δ13 CO2 might indicate a constant contribution of starch hydrolysis to the soluble sugar pool even outside pronounced stress periods (regular mixing).

How fresh is maple syrup? Sugar maple trees mobilize carbon stored several years previously during early springtime sap-ascent.

While trees store substantial amounts of nonstructural carbon (NSC) for later use, storage regulation and mobilization of stored NSC in long-lived organisms like trees are still not well understood. At two different sites with sugar maple (Acer saccharum), we investigated ascending sap (sugar concentration, δ(13) C, Δ(14) C) as the mobilized component of stored stem NSC during early springtime. Using the bomb-spike radiocarbon approach we were able to estimate the average time elapsed since the mobilized carbon (C) was originally fixed from the atmosphere and to infer the turnover time of stem storage. Sites differed in concentration dynamics and overall δ(13) C, indicating different growing conditions. The absence of temporal trends for δ(13) C and Δ(14) C indicated sugar mobilization from a well-mixed pool with average Δ(14) C consistent with a mean turnover time (TT) of three to five years for this pool, with only minor differences between the sites. Sugar maple trees hence appear well buffered against single or even several years of negative plant C balance from environmental stress such as drought or repeated defoliation by insects. Manipulative investigations (e.g. starvation via girdling) combined with Δ(14) C measurements of this mobilized storage pool will provide further new insights into tree storage regulation and functioning.

Non-structural carbon dynamics and allocation relate to growth rate and leaf habit in California oaks.

Trees contain non-structural carbon (NSC), but it is unclear for how long these reserves are stored and to what degree they are used to support plant activity. We used radiocarbon ((14)C) to show that the carbon (C) in stemwood NSC can achieve ages of several decades in California oaks. We separated NSC into two fractions: soluble (∼50% sugars) and insoluble (mostly starch) NSC. Soluble NSC contained more C than insoluble NSC, but we found no consistent trend in the amount of either pool with depth in the stem. There was no systematic difference in C age between the two fractions, although ages increased with stem depth. The C in both NSC fractions was consistently younger than the structural C from which they were extracted. Together, these results indicate considerable inward mixing of NSC within the stem and rapid exchange between soluble and insoluble pools, compared with the timescale of inward mixing. We observed similar patterns in sympatric evergreen and deciduous oaks and the largest differences among tree stems with different growth rates. The (14)C signature of carbon dioxide (CO2) emitted from tree stems was higher than expected from very recent photoassimilates, indicating that the mean age of C in respiration substrates included a contribution from C fixed years previously. A simple model that tracks NSC produced each year, followed by loss (through conversion to CO2) in subsequent years, matches our observations of inward mixing of NSC in the stem and higher (14)C signature of stem CO2 efflux. Together, these data support the idea of continuous accumulation of NSC in stemwood and that 'vigor' (growth rate) and leaf habit (deciduous vs evergreen) control NSC pool size and allocation.

Carbon dioxide emitted from live stems of tropical trees is several years old.

Storage carbon (C) pools are often assumed to contribute to respiration and growth when assimilation is insufficient to meet the current C demand. However, little is known of the age of stored C and the degree to which it supports respiration in general. We used bomb radiocarbon ((14)C) measurements to determine the mean age of carbon in CO2 emitted from and within stems of three tropical tree species in Peru. Carbon pools fixed >1 year previously contributed to stem CO2 efflux in all trees investigated, in both dry and wet seasons. The average age, i.e., the time elapsed since original fixation of CO2 from the atmosphere by the plant to its loss from the stem, ranged from 0 to 6 years. The average age of CO2 sampled 5-cm deep within the stems ranged from 2 to 6 years for two of the three species, while CO2 in the stem of the third tree species was fixed from 14 to >20 years previously. Given the consistency of (14)C values observed for individuals within each species, it is unlikely that decomposition is the source of the older CO2. Our results are in accordance with other studies that have demonstrated the contribution of storage reserves to the construction of stem wood and root respiration in temperate and boreal forests. We postulate the high (14)C values observed in stem CO2 efflux and stem-internal CO2 result from respiration of storage C pools within the tree. The observed age differences between emitted and stem-internal CO2 indicate an age gradient for sources of CO2 within the tree: CO2 produced in the outer region of the stem is younger, originating from more recent assimilates, whereas the CO2 found deeper within the stem is older, fueled by several-year-old C pools. The CO2 emitted at the stem-atmosphere interface represents a mixture of young and old CO2. These observations were independent of season, even during a time of severe regional drought. Therefore, we postulate that the use of storage C for respiration occurs on a regular basis challenging the assumption that storage pools serve as substrates for respiration only during times of limited assimilation.

Manipulative lowering of the water table during summer does not affect CO2 emissions and uptake in a fen in Germany.

We simulated the effect of prolonged dry summer periods by lowering the water table on three manipulation plots (D(1-3)) in a minerotrophic fen in southeastern Germany in three years (2006-2008). The water table at this site was lowered by drainage and by excluding precipitation; three nonmanipulated control plots (C(1-3)) served as a reference. We found no significant differences in soil respiration (R(Soil)), gross primary production (GPP), or aboveground respiration (R(AG)) between the C(1-3) and D(1-3) plots in any of the measurement years. The water table on the control plots was naturally low, with a median water table (2006-2008) of 8 cm below the surface, and even lower during summer when respiratory activity was highest, with median values (C(1-3)) between 11 and 19 cm below the surface. If it is assumed that oxygen availability in the uppermost 10 cm was not limited by the location of the water table, manipulative lowering of the water table most likely increased oxygen availability only in deeper peat layers where we expect R(Soil) to be limited by poor substrate quality rather than anoxia. This could explain the lack of a manipulation effect. In a second approach, we estimated the influence of the water table on R(Soil) irrespective of treatment. The results showed a significant correlation between R(Soil) and water table, but with R(Soil) decreasing at lower water tables rather than increasing. We thus conclude that decomposition in the litter layer is not limited by waterlogging in summer, and deeper peat layers bear no significant decomposition potential due to poor substrate quality. Consequently, we do not expect enhanced C losses from this site due to increasing frequency of dry summers. Assimilation and respiration of aboveground vegetation were not affected by water table fluctuations between 10 and >60 cm depth, indicating the lack of stress resulting from either anoxia (high water table) or drought (low water table).