▪ Abstract The use of stable isotope techniques in plant ecological research has grown steadily during the past two decades. This trend will continue as investigators realize that stable isotopes can serve as valuable nonradioactive tracers and nondestructive integrators of how plants today and in the past have interacted with and responded to their abiotic and biotic environments. At the center of nearly all plant ecological research which has made use of stable isotope methods are the notions of interactions and the resources that mediate or influence them. Our review, therefore, highlights recent advances in plant ecology that have embraced these notions, particularly at different spatial and temporal scales. Specifically, we review how isotope measurements associated with the critical plant resources carbon, water, and nitrogen have helped deepen our understanding of plant-resource acquisition, plant interactions with other organisms, and the role of plants in ecosystem studies. Where possible we also...

Stable Isotopes in Plant Ecology

Equilibrium and kinetic isotope fractionations during incomplete reactions result in minute differences in the ratio between the two stable N isotopes, 15N and 14N, in various N pools. In ecosystems such variations (usually expressed in per mil [δ15N] deviations from the standard atmospheric N2) depend on isotopic signatures of inputs and outputs, the input–output balance, N transformations and their specific isotope effects, and compartmentation of N within the system. Products along a sequence of reactions, e.g. the N mineralization–N uptake pathway, should, if fractionation factors were equal for the different reactions, become progressively depleted. However, fractionation factors vary. For example, because nitrification discriminates against 15N in the substrate more than does N mineralization, NH4+ can become isotopically heavier than the organic N from which it is derived.Levels of isotopic enrichment depend dynamically on the stoichiometry of reactions, as well as on specific abiotic and biotic conditions. Thus, the δ15N of a specific N pool is not a constant, and δ15N of a N compound added to the system is not a conservative, unchanging tracer. This fact, together with analytical problems of measuring δ15N in small and dynamic pools of N in the soil–plant system, and the complexity of the N cycle itself (for instance the abundance of reversible reactions), limit the possibilities of making inferences based on observations of 15N abundance in one or a few pools of N in a system. Nevertheless, measurements of δ15N might offer the advantage of giving insights into the N cycle without disturbing the system by adding 15N tracer.Such attempts require, however, that the complex factors affecting δ15N in plants be taken into account, viz. (i) the source(s) of N (soil, precipitation, NOx, NH3, N2-fixation), (ii) the depth(s) in soil from which N is taken up, (iii) the form(s) of soil-N used (organic N, NH4+, NO3−), (iv) influences of mycorrhizal symbioses and fractionations during and after N uptake by plants, and (v) interactions between these factors and plant phenology. Because of this complexity, data on δ15N can only be used alone when certain requirements are met, e.g. when a clearly discrete N source in terms of amount and isotopic signature is studied. For example, it is recommended that N in non-N2-fixing species should differ more than 5‰ from N derived by N2-fixation, and that several non-N2-fixing references are used, when data on δ15N are used to estimate N2-fixation in poorly described ecosystems.As well as giving information on N source effects, δ15N can give insights into N cycle rates. For example, high levels of N deposition onto previously N-limited systems leads to increased nitrification, which produces 15N-enriched NH4+ and 15N-depleted NO3−. As many forest plants prefer NH4+ they become enriched in 15N in such circumstances. This change in plant δ15N will subsequently also occur in the soil surface horizon after litter-fall, and might be a useful indicator of N saturation, especially since there is usually an increase in δ15N with depth in soils of N-limited forests.Generally, interpretation of 15N measurements requires additional independent data and modelling, and benefits from a controlled experimental setting. Modelling will be greatly assisted by the development of methods to measure the δ15N of small dynamic pools of N in soils. Direct comparisons with parallel low tracer level 15N studies will be necessary to further develop the interpretation of variations in δ15N in soil–plant systems. Another promising approach is to study ratios of 15N[ratio ]14N together with other pairs of stable isotopes, e.g. 13C[ratio ]12C or 18O[ratio ]16O, in the same ion or molecules. This approach can help to tackle the challenge of distinguishing isotopic source effects from fractionations within the system studied.

Tansley Review No. 95 15N natural abundance in soil-plant systems

Boreal forest soils function as a terrestrial net sink in the global carbon cycle. The prevailing dogma has focused on aboveground plant litter as a principal source of soil organic matter. Using C-14 bomb-carbon modeling, we show that 50 to 70% of stored carbon in a chronosequence of boreal forested islands derives from roots and root-associated microorganisms. Fungal biomarkers indicate impaired degradation and preservation of fungal residues in late successional forests. Furthermore, 454 pyrosequencing of molecular barcodes, in conjunction with stable isotope analyses, highlights root-associated fungi as important regulators of ecosystem carbon dynamics. Our results suggest an alternative mechanism for the accumulation of organic matter in boreal forests during succession in the long-term absence of disturbance.

http://science.sciencemag.org/content/sci/339/6127/1615.full.pdf

Roots and associated fungi drive long-term carbon sequestration in boreal forest.

Our understanding of how saprotrophic and mycorrhizal fungi interact to re-circulate carbon and nutrients from plant litter and soil organic matter is limited by poor understanding of their spatiotemporal dynamics. In order to investigate how different functional groups of fungi contribute to carbon and nitrogen cycling at different stages of decomposition, we studied changes in fungal community composition along vertical profiles through a Pinus sylvestris forest soil. We combined molecular identification methods with 14C dating of the organic matter, analyses of carbon:nitrogen (C:N) ratios and 15N natural abundance measurements. Saprotrophic fungi were primarily confined to relatively recently (< 4 yr) shed litter components on the surface of the forest floor, where organic carbon was mineralized while nitrogen was retained. Mycorrhizal fungi dominated in the underlying, more decomposed litter and humus, where they apparently mobilized N and made it available to their host plants. Our observations show that the degrading and nutrient-mobilizing components of the fungal community are spatially separated. This has important implications for biogeochemical studies of boreal forest ecosystems.

/pdf/spatial-separation-of-litter-decomposition-and-mycorrhizal-57wp06kh9t.pdf

Spatial separation of litter decomposition and mycorrhizal nitrogen uptake in a boreal forest

https://www.cell.com/trends/plant-science/pdf/S1360-1385(01)01889-1.pdf

Physiological mechanisms influencing plant nitrogen isotope composition.

Preliminary attempts to make retrospective studies of N balances and water stress in forest fertilization experiments by analyzing changes in the abundances of 15N and 13C, respectively, are discussed. Most evidence is from the Swedish Forest Optimum Nutrition Experiments, which have been running for two decades. Annual additions of N have been given either alone or in combination with other elements, notably P and K, every third year. Processes leading to loss of N, e.g. volatilization of ammonia, nitrification followed by leaching or denitrification, and denitrification alone, discriminate against the heavy isotope 15N. A correlation was found between fractional losses of added N and the change in δ15N (‰) during 19 years in current needles in a Scots pine forest, irrespective of source of N. Isotope effects were larger on urea than on ammonium nitrate plots (2 as compared to 9 δ15N (‰)) because of ammonia volatilization and higher rates of nitrification. They developed gradually over time, which opens possibilities to analyse the development of N saturation. However, the analysis may be confounded by shifts in 15N abundance of fertilizer N. In another trial, N isotope effects could be seen in both plants and soils 10 years after the last fertilization; they were smaller in soils because of a large pretreatment memory effect, but we expect them to persist there for decades.

Measurements of abundances of 15N and 13C as tools in retrospective studies of N balances and water stress in forests: A discussion of preliminary results

Analysis of the activity of the substrate‐induced enzyme nitrate reductase in plants can be used to indicate the mode of nitrogen nutrition and the presence of nitrate in the soil. We made two surv...

Shoot nitrate reductase activities of field‐layer species in different forest types: I. Preliminary surveys in Northern Sweden

The reversibility of induced N saturation was investigated in a 46-yr-old pine (Pinus sylvestris L.) forest in northern Sweden. Ammonium nitrate has been applied annually since 1971 to plots at average dosages of 36 (N1), 72 (N2), and 108 (N3) kg N ha{sup {minus}1} yr{sup {minus}1}, with or without P and K addition (background N deposition is < 4 kg ha{sup {minus}1} yr{sup {minus}1}). In 1990, after two decades of treatment, the largest N application (N3) was suspended, while N1 and N2 still received ammonium nitrate applications. Seven years after the last application in N3, the N availability measured as N concentration in plants [pine roots and needles and in leaves of the grass Deschampsia flexuosa (L.) Trin] and activity of the enzyme nitrate reductase in leaves of D. flexuosa, and {sup 15}N uptake by excised pine roots, was at the same levels as in N1, although more than twice the amount of N has been applied in total to N3. The arginine concentrations in pine needles, concentrations of exchangeable mineral N in the organic layer, and the uppermost 20 cm of the mineral soil were at the same levels as in the control plots. Thus, an experimentally induced Nmore » excess was, according to these measurements, to a high degree reversed 7 yr after the last N application. However, the composition of the understory vegetation still differed markedly from the untreated control 8 yr after the last N3 application.« less

Responses of a Nitrogen‐Saturated Forest to a Sharp Decrease in Nitrogen Input

High δ15N of tree foliage in forests subject to high N supply has been attributed to 15N enrichment of plant available soil N pools after losses of N through processes involving N isotope fractionation (ammonia volatilization, nitrification followed by leaching and denitrification, and denitrification in itself). However, in a long-term experiment with high annual additions of NH4NO3, we found no change in the weighted average δ15N of the soil, but attributed the high δ15N of trees to loss of ectomycorrhizal fungi and their function in tree N uptake, which involves redistribution of N isotopes in the ecosystem (Hogberg et al. New Phytol 189:515–525, 2011), rather than a loss of isotopically light N. Here, we compare the effects of additions of urea and NH4NO3 on the δ15N of trees and the soil profile, because we have previously found higher δ15N in tree foliage in trees in the urea plots. Doing this, we found no differences between the NH4NO3 and urea treatments in the concentration of N in the foliage, or the amounts of N in the organic mor-layer of the soil. However, the foliage of trees receiving the highest N loads in the urea treatment were more enriched in 15N than the corresponding NH4NO3 plots, and, importantly, the weighted average δ15N of the soil showed that N losses had been associated with fractionation against 15N in the urea plots. Thus, our results in combination with those of Hogberg et al. (New Phytol 189:515–525, 2011) show that high δ15N of the vegetation after high N load may be caused by both an internal redistribution of the N isotopes (as a result of change of the function of ectomycorrhiza) and by losses of isotopically light N through processes fractionating against 15N (in case of urea ammonia volatilization, nitrification followed by leaching and denitrification).

/pdf/is-the-high-15n-natural-abundance-of-trees-in-n-loaded-2mudw1ils5.pdf

Is the high 15N natural abundance of trees in N-loaded forests caused by an internal ecosystem N isotope redistribution or a change in the ecosystem N isotope mass balance?

Christian Johannisson

Papers

Measurements of abundances of 15N and 13C as tools in retrospective studies of N balances and water stress in forests: A discussion of preliminary results

Shoot nitrate reductase activities of field‐layer species in different forest types: I. Preliminary surveys in Northern Sweden

Responses of a Nitrogen‐Saturated Forest to a Sharp Decrease in Nitrogen Input

Is the high 15N natural abundance of trees in N-loaded forests caused by an internal ecosystem N isotope redistribution or a change in the ecosystem N isotope mass balance?