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 ).

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.

Non-structural carbohydrates in woody plants compared among laboratories.

Non-structural carbohydrates (NSC) in plant tissue are frequently quantified to make inferences about plant responses to environmental conditions. Laboratories publishing estimates of NSC of woody plants use many different methods to evaluate NSC. We asked whether NSC estimates in the recent literature could be quantitatively compared among studies. We also asked whether any differences among laboratories were related to the extraction and quantification methods used to determine starch and sugar concentrations. These questions were addressed by sending sub-samples collected from five woody plant tissues, which varied in NSC content and chemical composition, to 29 laboratories. Each laboratory analyzed the samples with their laboratory-specific protocols, based on recent publications, to determine concentrations of soluble sugars, starch and their sum, total NSC. Laboratory estimates differed substantially for all samples. For example, estimates for Eucalyptus globulus leaves (EGL) varied from 23 to 116 (mean = 56) mg g(-1) for soluble sugars, 6-533 (mean = 94) mg g(-1) for starch and 53-649 (mean = 153) mg g(-1) for total NSC. Mixed model analysis of variance showed that much of the variability among laboratories was unrelated to the categories we used for extraction and quantification methods (method category R(2) = 0.05-0.12 for soluble sugars, 0.10-0.33 for starch and 0.01-0.09 for total NSC). For EGL, the difference between the highest and lowest least squares means for categories in the mixed model analysis was 33 mg g(-1) for total NSC, compared with the range of laboratory estimates of 596 mg g(-1). Laboratories were reasonably consistent in their ranks of estimates among tissues for starch (r = 0.41-0.91), but less so for total NSC (r = 0.45-0.84) and soluble sugars (r = 0.11-0.83). Our results show that NSC estimates for woody plant tissues cannot be compared among laboratories. The relative changes in NSC between treatments measured within a laboratory may be comparable within and between laboratories, especially for starch. To obtain comparable NSC estimates, we suggest that users can either adopt the reference method given in this publication, or report estimates for a portion of samples using the reference method, and report estimates for a standard reference material. Researchers interested in NSC estimates should work to identify and adopt standard methods.

Growth in two common gardens reveals species by environment interaction in carbon isotope discrimination of Eucalyptus.

One-year-old sun leaves of 60 species of Eucalyptus were collected in August 2005 at an arboretum in South Australia with a mean annual rainfall of 427 mm, and 14 of the same species were sampled at an arboretum in Western Australia with a mean annual rainfall of 216 mm. We determined the genetic and phenotypic variation in carbon isotope composition (delta13C), specific leaf area (SLA) and nitrogen content per unit area of the species at each site. There were very significant (P < 0.001) differences in delta13C among the species at both sites. The mean delta13C of the 60 species at the wetter site was -27.6 per thousand (from -25.8 per thousand in Eucalyptus youngiana to -29.9 per thousand in Eucalyptus salicola) and of the 14 species at the drier site was -25.3 per thousand (from -23.7 per thousand in Eucalyptus ravida to -27.3 per thousand in Eucalyptus ewartiana). Of the 14 species common to both sites, four species had similar values of delta13C at the two sites despite the differences in rainfall, whereas in others the values of delta13C were significantly (P < 0.001) lower (more negative) at the wet than at the dry site. The SLA and nitrogen content per unit leaf area also differed significantly among the species (P < 0.001), but there was not a common relationship between delta13C and SLA or between delta13C and nitrogen content at the two sites. The strong species by environment interaction resulted from some species demonstrating phenotypic plasticity for delta13C, while others were inherently stable across environments.

Annual rainfall does not directly determine the carbon isotope ratio of leaves of Eucalyptus species.

Leaf carbon isotope discrimination (delta13C) was widely considered to directly reflect the rainfall environment in which the leaf developed, but recent observations have queried this. The relationship between delta13C and rainfall was explored in Eucalyptus species growing along a rainfall gradient in Australia. The leaves of 43 species of Eucalyptus and the closely related Corymbia species produced in 2003 were sampled in September 2004 at 50 sites and grouped into 15 locations along a rainfall gradient in southwest Western Australia. At 24 sites, the same species and same trees were sampled as in a study in September 2003 when leaves produced in 2002 were sampled. The rainfall in 2004 was on average 190 mm (range 135-270 mm) higher at all locations than in 2003. In the leaves sampled in 2004, the mean carbon isotope discrimination (delta13C) across the 15 locations decreased 2.9 per thousand per 1000 mm of rainfall, the specific leaf area (SLA) increased by 2.9 m2 kg(-1) per 1000 mm of rainfall and the nitrogen (N) content decreased by 1.56 g m(-2) per 1000 mm of rainfall. In contrast, a comparison between the leaves produced in the drier 2002 year compared with the wetter 2003 year showed that there was a strong correlation (r2= 0.85) between the SLA values between years and a trend for higher values with increasing SLA, but the values of delta(13)C were on average only 0.38 per thousand lower (more negative) at all locations in the wetter year, equivalent to a decrease of 2.0 per thousand per 1000 mm of rainfall. The results suggest that while there may be constitutive differences in leaf morphology, SLA and N content per unit area, increasing rainfall or cloudiness associated with higher rainfall increases SLA and decreases N content per unit area. We conclude that rainfall does not directly influence delta13C, but induces leaf morphological and physiological changes that affect the resultant delta13C.

Laser ablation-combustion-GC-IRMS--a new method for online analysis of intra-annual variation of delta13C in tree rings.

We present a new, rapid method for high-resolution online determination of delta13C in tree rings, combining laser ablation (LA), combustion (C), gas chromatography (GC) and isotope ratio mass spectrometry (IRMS) (LA-C-GC-IRMS). Sample material was extracted every 6 min with a UV-laser from a tree core, leaving 40-microm-wide holes. Ablated wood dust was combusted to CO2 at 700 degrees C, separated from other gases on a GC column and injected into an isotope ratio mass spectrometer after removal of water vapor. The measurements were calibrated against an internal and an external standard. The tree core remained intact and could be used for subsequent dendrochronological and dendrochemical analyses. Cores from two Scots pine trees (Pinus sylvestris spp. sibirica Lebed.) from central Siberia were sampled. Inter- and intra-annual patterns of delta13C in whole-wood and lignin-extracted cores were indistinguishable apart from a constant offset, suggesting that lignin extraction is unnecessary for our method. Comparison with the conventional method (microtome slicing, elemental analysis and IRMS) indicated high accuracy of the LA-C-GC-IRMS measurements. Patterns of delta13C along three parallel ablation lines on the same core showed high congruence. A conservative estimate of the precision was +/- 0.24 per thousand. Isotopic patterns of the two Scots pine trees were broadly similar, indicating a signal related to the forest stand's climate history. The maximum variation in delta13C over 22 years was about 5 per thousand, ranging from -27 to -22.3 per thousand. The most obvious pattern was a sharp decline in delta13C during latewood formation and a rapid increase with spring early growth. We conclude that the LA-C-GC-IRMS method will be useful in elucidating short-term climate effects on the delta13C signal in tree rings.