Quantifying the contribution of soil organic matter turnover to forest soil respiration, using natural abundance δC

Quantifying the loss of soil carbon through respiration has proved difficult, due to the challenge of measuring the losses associated with the turnover of soil organic matter (SOM) as distinct from autotrophic components. In forest ecosystems the δ¹³C value of respiration from turnover of SOM (δ¹³CR_SOM) is typically 2-4‰ enriched compared with that from roots and associated microbes (δ¹³CR_ROOTS), with that from the litter (δ¹³CR_LITTER) lying between the two. We measured soil respiration at 50 locations in a forest soil and then used differences in isotopic signatures to quantify the proportion of soil respiration arising from the turnover of SOM (fR_SOM) at a subset of 30 locations, chosen randomly. The soil surface CO₂ efflux was collected using an open chamber system supplied with CO₂-free air and the δ¹³C signature (δ¹³CR_S) measured, giving a mean (±SD) value across the site of −26.1 ± 0.58‰. The values of δ¹³CR_ROOTS, δ¹³CR_LITTER and δ¹³CR_SOM were measured at each location by incubation of roots, litter and root-free soil and collection of the CO₂ for isotopic analysis. δ¹³CR_SOM became progressively depleted with length of incubation (1.5‰ after 8 h), so CO₂ was collected after 20 min. The mean value of δ¹³CR_LITTER was −27.2 ± 0.68 ‰, which was indistinguishable from δ¹³CR_ROOTS of −27.6 ± 0.51‰, while δ¹³CR_SOM was −25.1 ± 0.88‰. δ¹³CR_ROOTS and δ¹³CR_SOM measured at each location were used as the end points of a two component mixing model to calculate fR_SOM, giving a mean value for fR_SOM of 0.61 ± 0.28. It was not possible to estimate fR_SOM using the total C contents of the roots and soil which were significantly depleted in ¹³C in comparison to their respired CO₂. However, at seven locations the δ¹³CR_S was slightly enriched compared with δ¹³CR_SOM (mean 0.3‰), which was not considered significantly different so fR_SOM was constrained to 1.0. If these seven rings were excluded mean fR_SOM was 0.49 ± 0.20. We have shown the possibility of using natural abundance ¹³C discrimination to quantify fR_SOM in a forest soil with an input of carbon only from C₃ photosynthesis.     

Palatability trials on hardwood leaf litter grown under elevated CO2: a stable carbon isotope study

The palatability to isopods and microbes of a broad range of hardwood leaf litter, derived from three field CO₂-enrichment experiments in the USA, was investigated, using δ¹³C, to trace the C flow from litter to isopods and to CO₂ respired by microbial decomposition. Leaf litter grown under elevated CO₂ had δ¹³C values ranging from −39 to −45‰, which were significantly different from ambient litter δ¹³C values of around −30‰. Litter palatability to isopods of the Porcellio sp. was tested by incubating ambient- and elevated-CO₂ litter, and a mixture of the two, in the presence of isopods for 14 days, under environmentally controlled conditions; δ¹³C was measured on litter and isopods' body before and after incubation. In an additional experiment, litter was incubated in the absence of fauna for 30 days, and on five occasions the δ¹³C of the CO₂ respired from litter was measured. The ¹³C label was clearly carried from the litter source to the isopods' bodies, and their faeces. For microbial-respired CO₂, δ¹³C was significantly higher than that of the litter source, suggesting preferential degradation of substrates enriched in ¹³C as compared to those in the overall litter. With the exception of Quercus myrtifolia leaf litter, elevated CO₂ did not affect the palatability to isopods nor the microbial degradation of any of the litters, possibly as a result of unaltered litter N concentration. However, significant differences in litter palatability and decay rates were observed among the different species. With this study, the use of isotopically labelled litter material was confirmed as a key methodology that can significantly contribute to the advancement of the understanding of litter decomposition and of the quantification of C fluxes in the process.     

C signature of CO2 evolved from incubated maize residues and maize-derived sheep faeces

Analyses of the spatial and temporal variations in the natural abundance of ¹³C are frequently employed to study transformations of plant residues and soil organic matter turnover on sites where long continued vegetation with the C₃-type photosynthesis pathway has been replaced with a C₄-type vegetation (or vice versa). One controversial issue associated with such analyses is the significance of isotopic fractionation during the microbial turnovers of C in complex substrates. To evaluate this issue, C₃-soil and quartz sand were amended with maize residues and with faeces from sheep feed exclusively on maize silage. The samples were incubated at 15 °C for 117 days (maize residues) or 224 days (sheep faeces). CO₂ evolved during incubation was trapped in NaOH and analysed for C isotopic contents. At the end of incubation, 63 and 50% of the maize C was evolved as CO₂ in the soil and sand, respectively, while 32% of the faeces C incubated with soil and with sand was recovered as CO₂. Maize and faeces showed a similar decomposition pattern but maize decomposed twice as fast as faeces. The δ¹³C of faeces was 0.3‰ lower than that of the maize residue (δ¹³C −13.4‰), while the δ¹³C of the C₃-soil used for incubation was −31.6‰. The δ¹³C value of the CO₂ recovered from unamended C₃-soil was similar or slightly lower (up to −1.5‰) than that of the C₃-soil itself except for an initial flush of ¹³C enriched CO₂. The δ¹³C values of the CO₂ from sand-based incubations typically ranged −15‰ to −17‰, i.e. around −3‰ lower than the δ¹³C measured for maize and faeces. Our study clearly demonstrates that the decomposition of complex substrates is associated with isotopic fractionation, causing evolved CO₂ to be depleted in ¹³C relative to substrates. Consequently the microbial products retained in the soil must be enriched in ¹³C.     

Use of C abundance to study short-term pig slurry decomposition in the field

Applying pig slurry (PS) on agricultural soils is a common practice. However, its impact on soil organic C dynamics is not clear. This experiment investigated the use of natural ¹³C abundance to study the short-term C mineralization of anaerobically stored PS under field conditions. Measurements of δ¹³C-CO₂ were made on soil air samples obtained from a bare sandy loam during 22 d following incorporation of either PS alone, PS+barley straw, or barley straw alone; an unamended treatment was used as a control. Slurry C was enriched in ¹³C (−20.0‰) because of the high corn (Zea mays L.) content of the animal diet. This value contrasted with δ¹³C of −28.4‰ for the soil organic matter and of −29.0‰ for the barley straw. A peak of high δ¹³CO₂ values (average of −9.2‰) was observed on the day of PS application and was attributed to the dissociation of PS carbonates when mixed with the relatively acidic soil. After this initial burst, 36% of the evolved CO₂ originated from the decomposing PS. After 22 d of incubation, approx. 20% of the PS-C had been lost as CO₂. This short-term field study did not show any priming effect of PS on the mineralization of straw or native soil C. Due to its heterogeneity, the use of the isotopic composition of the evolved CO₂ for estimating PS decomposition requires precaution either through the use of a specific experimental design involving comparable C3 and C4 treatments, or calculations to account for the presence of ¹³C-enriched inorganic C in the PS.     

Translocation of surface litter carbon into soil by Collembola

Soil invertebrates are important in nutrient cycling in soils, but the degree to which mesofauna such as Collembola are responsible for the direct movement of carbon (C) from the litter layer into soil has not yet been ascertained. We used naturally occurring stable C isotopic differences between a C₄ soil and alder leaves (C₃) to examine the effect of the collembolan Folsomia candida on C translocation into soil in laboratory microcosms. Collembolan numbers greatly increased in the presence of alder, but despite large collembolan populations there were no changes in decomposition rate (measured as litter mass loss, cumulative respired CO₂ and alder C:N ratios). Small changes in the δ¹³C values of bulk soil organic matter were detected, but could not be assigned to collembolan activity. However, mean δ¹³C values of soil microbial phospholipid fatty acids (PLFAs) were significantly lower in the presence of alder and Collembola together, demonstrating that collembolan activities resulted in greater availability of litter-derived C to the soil microbial community. Additionally, the presence of Collembola resulted in the translocation of alder-derived compounds (chlorophyll and its breakdown product pheophytin) into soil, demonstrating that Collembola modify soil organic matter at the molecular level. These results are consistent with deposition of collembolan faeces in underlying soil and demonstrate that despite their small size, Collembola contribute directly to C transport in the litter-soil environment.     

Microbial community structure mediates response of soil C decomposition to litter addition and warming

Microbial activity has been highlighted as one of the main unknowns controlling the fate and turnover of soil organic matter (SOM) in response to climate change. How microbial community structure and function may (or may not) interact with increasing temperature to impact the fate and turnover of SOM, in particular when combined with changes in litter chemistry, is not well understood. The primary aim of this study was to determine if litter chemistry impacted the decomposition of soil and litter-derived carbon (C), and its interaction with temperature, and whether this response was controlled by microbial community structure and function. Fresh or pre-incubated eucalyptus leaf litter (¹³C enriched) was added to a woodland soil and incubated at 12, 22, or 32 °C. We tracked the movement of litter and soil-derived C into CO₂, water-extractable organic carbon (WEOC), and microbial phospholipids (PLFA). The litter additions produced significant changes in every parameter measured, while temperature, interacting with litter chemistry, predominately affected soil C respiration (priming and temperature sensitivity), microbial community structure, and the metabolic quotient (a proxy for microbial carbon use efficiency [CUE]). The direction of priming varied with the litter additions (negative with fresh litter, positive with pre-incubated litter) and was related to differences in the composition of microbial communities degrading soil-C, particularly gram-positive and gram-negative bacteria, resulting from litter addition. Soil-C decomposition in both litter treatments was more temperature sensitive (higher Q₁₀) than in the soil-only control, and soil-C priming became increasingly positive with temperature. However, microbes utilizing soil-C in the litter treatments had higher CUE, suggesting the longer-term stability of soil-C may be increased at higher temperature with litter addition. Our results show that in the same soil, the growth of distinct microbial communities can alter the turnover and fate of SOM and, in the context of global change, its response to temperature.     

Effect of Collembola on mineralization of litter and soil organic matter

Although soil Collembola are known to contribute to soil carbon (C) cycling, their contribution to the mineralization of C sources that differ in bioavailability, such as soil organic C (SOC) and leaf litter, is unknown. Stable C isotopes are often used to quantify the effects of both soil C and litter C on C mineralization. Here, ¹³C-labeled litter was used to investigate the effects of Collembola (Folsomia candida) on the mineralization of both SOC and litter C in laboratory microcosms. The three microcosm treatments were soil alone (S); soil treated with δ¹³C-labeled litter (SL); and soil treated with δ¹³C-labeled litter and Collembola (SLC). The presence of Collembola did not significantly affect soil microbial biomass or litter mass loss and only had a small effect on CO₂ release during the first week of the experiment, when most of the CO₂ was derived from litter rather than from SOC. Later, during the experiment (days 21 and 63), when litter-derived labile C had been depleted and when numbers of Collembola had greatly increased, Collembola substantially increased the emission of SOC-derived CO₂. These results suggest that the effect of Collembola on soil organic C mineralization is negatively related to C availability.     

Spatial heterogeneity in the relocation of added C within the structure of an upland grassland soil

A pulse of ¹³CO₂ was added to the above ground vegetation in an upland grassland to determine the effects of faunal diversity on the flux of carbon to the surface horizons of the soil. Faunal diversity was manipulated by liming and biocide treatments for three years prior to the pulse addition. The relocation of ¹³C within roots and rhizosphere soil was determined by analysis of samples of bulk soil and of specific features identified on soil thin sections on four dates after the addition of the ¹³CO₂ pulse. Analysis of bulk soils showed only a small enrichment in ¹³C and no significant effects of the treatments. Analysis by isotope ratio mass spectrometry of the products of in situ laser combustion of root material and aggregates formed from faunal excrement showed that the distribution of the newly photosynthesised ¹³C is very localised, with large spatial variability in soil and root δ¹³C at scales of less than 1 mm. δ¹³C values ranged from the natural abundance level of around −28‰ to −4.9‰ in roots and to −8.4‰ in aggregates. The small pulse and large spatial variability masked any effects of the liming and biocide treatments in these soils. However, the variability in the relocation of newly photosynthesised carbon may help to explain the large spatial variability found in bacterial numbers at the sub-mm scale within soils and emphasises the importance of the accessibility of substrates to decomposers in undisturbed structured soils.     

C fractionation at the root-microorganisms-soil interface: A review and outlook for partitioning studies

Natural variations of the ¹³C/¹²C ratio have been frequently used over the last three decades to trace C sources and fluxes between plants, microorganisms, and soil. Many of these studies have used the natural-¹³C-labelling approach, i.e. natural δ¹³C variation after C₃-C₄ vegetation changes. In this review, we focus on ¹³C fractionation in main processes at the interface between roots, microorganisms, and soil: root respiration, microbial respiration, formation of dissolved organic carbon, as well as microbial uptake and utilization of soil organic matter (SOM). Based on literature data and our own studies, we estimated that, on average, the roots of C₃ and C₄ plants are ¹³C enriched compared to shoots by +1.2 ± 0.6‰ and +0.3 ± 0.4‰, respectively. The CO₂ released by root respiration was ¹³C depleted by about −2.1 ± 2.2‰ for C₃ plants and −1.3 ± 2.4‰ for C₄ plants compared to root tissue. However, only a very few studies investigated ¹³C fractionation by root respiration. This urgently calls for further research. In soils developed under C₃ vegetation, the microbial biomass was ¹³C enriched by +1.2 ± 2.6‰ and microbial CO₂ was also ¹³C enriched by +0.7 ± 2.8‰ compared to SOM. This discrimination pattern suggests preferential utilization of ¹³C-enriched substances by microorganisms, but a respiration of lighter compounds from this fraction. The δ¹³C signature of the microbial pool is composed of metabolically active and dormant microorganisms; the respired CO₂, however, derives mainly from active organisms. This discrepancy and the preferential substrate utilization explain the δ¹³C differences between microorganisms and CO₂ by an ‘apparent’ ¹³C discrimination. Preferential consumption of easily decomposable substrates and less negative δ¹³C values were common for substances with low C/N ratios. Preferential substrate utilization was more important for C₃ soils because, in C₄ soils, microbial respiration strictly followed kinetics, i.e. microorganisms incorporated heavier C (? = +1.1‰) and respired lighter C (? = −1.1‰) than SOM. Temperature and precipitation had no significant effect on the ¹³C fractionation in these processes in C₃ soils. Increasing temperature and decreasing precipitation led, however, to increasing δ¹³C of soil C pools.Based on these ¹³C fractionations we developed a number of consequences for C partitioning studies using ¹³C natural abundance. In the framework of standard isotope mixing models, we calculated CO₂ partitioning using the natural-¹³C-labelling approach at a vegetation change from C₃ to C₄ plants assuming a root-derived fraction between 0% and 100% to total soil CO₂. Disregarding any ¹³C fractionation processes, the calculated results deviated by up to 10% from the assumed fractions. Accounting for ¹³C fractionation in the standard deviations of the C₄ source and the mixing pool did not improve the exactness of the partitioning results; rather, it doubled the standard errors of the CO₂ pools. Including ¹³C fractionations directly into the mass balance equations reproduced the assumed CO₂ partitioning exactly. At the end, we therefore give recommendations on how to consider ¹³C fractionations in research on carbon flows between plants, microorganisms, and soil.     

Effluxed CO2-C from sterilized and unsterilized treatments of a calcareous soil

Soil inorganic carbon (C) represents a substantial C pool in arid ecosystems, yet little data exist on the contribution of this pool to ecosystem C fluxes. A closed jar incubation study was carried out to test the hypothesis that CO₂-¹³C production and response to sterilization would differ in a calcareous (Mojave Desert) soil and a non-calcareous (Oklahoma Prairie) soil due to contributions of carbonate-derived CO₂. In addition to non-sterilized controls, soils were subjected to sterilization treatments (unbuffered HgCl₂ addition for Oklahoma soil and unbuffered HgCl₂ addition, buffered HgCl₂ addition, and autoclaving for Mojave Desert soil) to decrease biotic respiration and more readily measure abiotic CO₂ flux. Temperature and moisture treatments were also included with sterilization treatments in a factorial design.The rate of CO₂ production in both soils was significantly decreased (36-87%) by sterilization, but sterilization treatments differed in effectiveness. Sterilization had no significant effect on effluxed CO₂-¹³C values in the non-calcareous Oklahoma Prairie soil and autoclaved Mojave Desert soil as compared to their respective non-sterilized controls. However, sterilization significantly altered CO₂-¹³C values in Mojave Desert soil HgCl₂ sterilization treatments (both buffered and non-buffered). Plots of 1/CO₂ versus CO₂-δ¹³C (similar to Keeling plots) indicated that the source CO₂-δ¹³C value of the Oklahoma Prairie soil treatments was similar to the δ¹³C value of soil organic matter [(SOM); −17.76‰ VPDB] whereas the source for the (acidic) unbuffered-HgCl₂ sterilized Mojave Desert soil was similar to the δ¹³C value of carbonates (−0.93‰ VPDB). The source CO₂-δ¹³C value of non-sterilized and autoclaved (−18.4‰ VPDB) Mojave Desert soil treatments was intermediate between SOM (−21.43‰ VPDB) and carbonates and indicates up to 13% of total C efflux may be from abiotic sources in calcareous soils.