Displays

Respiration is likely the most often measured process in soil ecology. It is used as a general measurement of soil activity, and physiologically related to microbial maintenance requirements, growth, and soil organic matter production via biochemical efficiency and CUE.

Genomic tools are increasingly used in soil ecology for measurement of community composition, and functional analysis of communities, and when combined with stable isotopes, can be used to infer activities, either of the whole community or of individual taxa. However, relating genomic or gene-expressed functions to whole ecosystem processes, such as respiration, remains a conceptual and practical problem.

We analyzed the biochemical processes related to respiration and determine how, during a short soil incubation experiment in the presence of glucose, these processes change. Furthermore, we will show how gene and transcript abundances of respiratory processes vary across more than 4000 soil and rhizosphere samples in forests and grasslands and other biomes.  

Results illustrate the treasure trove of biochemical information available to us in the form of metagenomes and metatranscriptomes.

How to cite: Dijkstra, P., Chuckran, P. F., Hungate, B. A., Schwartz, E., Glavina del Rio, T., and Eloe-Fadrosh, E.: What is respiration - response to glucose addition, presence of plant roots and differences across biomes, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-12072, https://doi.org/10.5194/egusphere-egu2020-12072, 2020.

The controlling factors and role of different soil microorganisms in rhizosphere priming effects (PE) are not yet well understood, especially the link between microbial growth and their carbon use efficiency (CUE) and PE remains poorly understood. We hypothesized that the positive PE (enhanced SOM decomposition) results from microbes using the additional C/energy from added labile substrates to decompose recalcitrant materials to release N (“microbial N mining hypothesis”). High CUE could lead to efficient growth of microbes and thus more decomposition of SOM and higher PE.

To test these hypothesis we assessed PE along a boreal forest gradient ranging from Estonia to Northern Finland, with soil C:N ratios and fungal to bacterial ratios increasing towards north. The soils received daily additions of ¹³C-labelled glucose during one week (dissolved in heavy water, 5 at% D₂O). Control soils received D₂O only. Respiration of glucose and respiration of SOM were distinguished by continuously measuring ¹³CO₂ using a Picarro analyzer. We also measured microbial incorporation (into PLFAs) of ¹³C and D to assess to which extent different microbial groups rely on labile C input (¹³C-labelled) and on SOM. We further used these results to calculate the CUE of glucose and of SOM decomposition.

Glucose additions induced PE (12-52% increase in SOM respiration) in all soils, but there was no linear relationship between PE and soil C:N ratio. Instead, cumulative PE (µg C g^-1 SOM) and the relative magnitude of the PE (%) were positively correlated with the average C:N imbalance experienced by the microbes (calculated as soil C:N ratio/microbial biomass C:N ratio). There was a positive relationship between the potential activity of total oxidative enzymes and the cumulative SOM respiration, but the same enzyme concentration resulted in higher SOM respiration in the glucose treatment. We suggest that glucose additions increased the activity of these enzymes rather than their concentrations.

Microbial incorporation of D and ¹³C into in PLFAs demonstrated that glucose additions stimulated both fungal and bacterial growth. Our results indicate that increased growth of fungi on the added ¹³C glucose was especially important for the PE, since the magnitude of PE was correlated with the ratio of fungal/bacterial growth on glucose and on SOM. High C:N ratio soils were fungal dominated, and there was a clear positive relationship between glucose CUE and fungal to bacterial ratio, indicating that fungal dominated communities had higher CUE. Bacteria were more affected by low N availability, since total bacteria growth and ¹³C uptake were lower in the high C:N ratio soils. When fungal growth was high relative to bacterial growth CUE was consistently higher, whether it was the total CUE, CUE of glucose or SOM respiration. Our results indicate that if fungal dominated communities can efficiently grow on the added glucose, they will have excess resources for decomposing N releasing recalcitrant substrates. This releases bioavailable C and N that can also increase bacterial respiration of SOM derived C.

How to cite: Karhu, K., Alaei, S., Li, J., and Bengtson, P.: Microbial carbon use efficiency and priming of soil organic matter mineralization by glucose additions in boreal forest soils with different C:N ratios, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-5775, https://doi.org/10.5194/egusphere-egu2020-5775, 2020.

Fertilization practices can influence the soil nutrients and fertility status, which subsequently induce changes in soil carbon (C):nitrogen (N) ratio and rebuilt C:N stoichiometric balances between microbial biomass and resources. In this study, we investigated how available resource C:N ratio can regulate the priming effect (PE) to maintain microbial C:N stoichiometric balance by adding ¹³C-labeled glucose to four long-term fertilized paddy soils [Control (no fertilization), NPK (fertilized with mineral NPK fertilizers) , NPKS (NPK combined with straw), NPKM (NPK combined with manure)]. Glucose addition significantly increased SOC mineralization and subsequently induced a positive priming effect at day 2 of incubation, whereas the PE became negative after 20 days. DOC contents were increased by more than 1000% with glucose addition at day 2, whereas they rapidly decreased to -10% to -50% compared with those in soils without glucose addition. With the changes in available and biomass C and N, the microbial C:N imbalance initially increased to 3.3–6.8, and then reduced to the level as that in the soils without glucose addition. At the end of incubation, the microbial C:N imbalance in the glucose-treated soils was ranked as Control < NPK < NPKM < NPKS. This suggested that, without organic fertilization, soils were highly susceptible to labile C and increased SOC mineralization, leading to C limitation. The PE was positively related to DOC and NH₄⁺ ratio, but negatively associated with microbial C:N imbalance, suggesting that the labile C supplied stimulated microbial stoichiometric decomposition of SOM. Glucose addition modified enzyme activities after 20 days, to allow the microorganisms to break up complex C compounds for C source. Our findings suggested that soil microorganisms could regulate extracellular hydrolytic enzyme production and their relative stoichiometric ratios to obtain necessary elements, thereby adjusting the microbial biomass C:N to the resource stoichiometry.

How to cite: Ge, T., Zhu, Z., and Wu, J.: Soil available resource C:N ratio regulates the priming effect by maintaining microbial C:N stoichiometric balance in long-term fertilized paddy soils, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-14449, https://doi.org/10.5194/egusphere-egu2020-14449, 2020.

Rhizosphere is the most biologically active region between the plant and the surrounding soil where plant release their fixed carbon into the soils. Depending on the availability and types of carbon compounds released from the plant, they can directly solubilize nutrient or indirectly influence nutrient cycling by promoting increased microbial activity in the rhizosphere. In this study we applied phosphate oxygen isotope ratios (d¹⁸O_P) to determine how root exudate influences temporal variation in microbial activities and P cycling in the rhizosphere. Rhizoboxes were filled with soils, watered to 75% water holding capacity and equilibrated for 10 days. After equilibration labeled phosphate isotopes synthesized using ¹⁸O labeled water was applied. Then a mixed exudate (i.e., glucose, alanine, and oxalate in the ratio of 1:1:1) was introduced into the soil for 4, 10, and 20 days via an artificial root. We used a sequential extraction technique (i.e., resin-Pi, NaHCO₃-P, NaOH-P, and HCl-P) to track the fate of applied P in bulk and rhizosphere soils. The root exudate effects on the rate of P cycling and microbial activity were investigated using phosphate oxygen isotope ratios in the resin-Pi pool. Microbial community structures was determined using phospholipid fatty acids (PLFA) profiles. After supplying root exudate for 4, 10, and 20 days, the results showed that bioavailable P (i.e., resin-Pi) concentration was always higher in the bulk soil compared to rhizosphere soil and originally bioavailable P transformed gradually into unavailable P (i.e., NaOH-P and HCl-P). After supplying exudate compound for 4 days, the applied PO₄ was mostly in the resin-Pi pool and its isotopic composition was heavier than the equilibrium isotopic composition suggesting that this Pi pool was not completely cycled by the microorganisms. As we continue supplying exudate compounds, the concentration of resin-Pi gradually decreased and as microbial activities increased, its isotopic composition got closer to the equilibrium isotopic composition. Further the microbial community structure in the rhizosphere soil after supply of root exudate were distinctly different then the bulk soil. Using phosphate oxygen isotopes this study shows the influence of root exudates on the rate of P cycling in rhizosphere soils.

How to cite: Joshi, S. R. and McNear, D. H.: Root exudate induced microbial activities and phosphorus cycling in soil: An application of phosphate oxygen isotopes, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-12719, https://doi.org/10.5194/egusphere-egu2020-12719, 2020.

It has been assumed for a long time that stable soil organic carbon (SOC) results from selective preservation of plant residues. Yet, a new paradigm points to a more active role of microorganisms in building SOC storage. In this context, even labile C, such as sugars, may persist in soil for a long time due to their incorporation into microbial biomass and ultimately necromass. The latter is considered as a relatively stable pool. However, little is known about the cycling of labile C through the microbial biomass and the turnover time of its residues. Unraveling the mechanisms and regulating factors would be critical for understanding SOC stabilization in soil.

We assume that the fate of labile C is mainly driven by microbial nitrogen (N) demand and supply. Specifically, we hypothesize that (1) high N demand forces microbes to decompose N-rich substances (“microbial N mining”), such as amino sugars, leading to a rapid turnover of microbial necromass, and that (2) labile C is stabilized in microbial necromass when N demand is met.

To investigate these hypotheses, we set up a greenhouse pot experiment including four treatments: (1) bare soil, (2) bare soil+N, (3) tree, and (4) tree+N. The soil is a sandy and nutrient poor forest soil from southern Finland. Trees are 1 m high pines (Pinus Sylvestris), which are supposed to induce microbial N deficiency by exuding easily degradable C compounds and by competing with microbes for mineral N. In order to follow to fate of labile C, we added trace amounts of ¹³C labeled glucose to the soil (4 replicates per treatment). As a control to account for background variations in ¹³C, we added ¹²C glucose to another set of pots (4 replicates per treatment). Up to now, we sampled the soil 1 day, 3 days, 8 days, 1 month, 3 months, 6 months, 9 months, and 1 year after glucose addition. Measurements of the ¹³C recovery in soil, microbial biomass, water extractable C, PLFA, amino sugars, and DNA are in progress.

First results indicate that the largest loss of ¹³C tracer occurred in the unfertilized tree treatment, i.e., where N demand was high but N supply was low. Here, only 22% of the ¹³C glucose remained after 3 month, whereas 40% remained in the fertilized tree treatment. Only small proportions of the recovered ¹³C were present in the pool of water extractable C (<1%) and in living microbial biomass (8±3%, 3 days after glucose addition). As protection by clay minerals and aggregates is likely not a relevant process in this sandy soil, we suspect the remaining ¹³C to be stabilized in microbial residues, but depending on N demand. We assume that microbial necromass accounts for a considerable proportion to total SOC storage, especially under conditions of adequate nitrogen supply.

How to cite: Meyer, N., Sietiö, O.-M., Adamczyk, S., Biasi, C., Ambus, P., Mganga, K., Shrestha, R., Kalu, S., Martin, A., Glaser, B., and Karhu, K.: Stabilization of labile carbon in soil microbial biomass and necromass – A question of nitrogen deficiency?, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-7121, https://doi.org/10.5194/egusphere-egu2020-7121, 2020.

Temperate forests in Chile have experienced increasing temperatures and extreme climatic events, such as severe drought and short winters in unique Araucaria araucana forest in Nahuelbuta National Park. Therefore, it is relevant to understand the impact of drying and rewetting (D/R) or freezing and thawing (F/T) on SOM turnover in these ecosystems. Particularly important is the destabilization of soil organic matter (SOM) by microbial activity, which is highly heterogeneous and influenced by soil properties and water cycles. Drying and rewetting or F/T cycles accelerate particulate organic matter (POM) decomposition by aggregate disruption, thereby, decreasing carbon (C) availability for soil microorganism. We hypothesized that frequent D/R and F/T cycles release labile organic C locked away in the aggregates for microbial consumption. We assumed that a repeated number of D/R and F/T cycles enhance the preferential C utilization of fresh organic substrate. In the present study an incubation experiment was conducted for 27 days to assess the effect of F/T (-18 ºC to room temperature) and D/R (-500 kPa to 33 kPa, field capacity) cycles on labelled ¹⁴C glucose and ¹³C lignocellulose decomposition, soil aggregates size and POM fractions distributions. CO₂ efluxes and priming effect (PE), i.e. the turnover acceleration or retardation of native C mineralization, C use efficiency (CUE) and C allocation in soil aggregate classes as POM-light, POM-occluded and heavy fractions were also determined. Labelled glucose was mainly allocated in macro (> 250 mm) and microaggregates (< 250 mm) as part of the POM-light fraction. In contrast, labelled lignocellulose was allocated in microaggregate in the POM-occluded and heavy fraction. CUE was similar amongst all treatments. The PE was negative in soil with and without cycles and it was much more pronounced (-125 mg C kg^-1 soil) for F/T cycles than D/R (-50 mg C kg^-1 soil) at the end of incubation. The C:N ratio of soil following mining theory is further discussed. We conclude that D/R cycles clearly retarded the native C mineralization by preferential use of labelled ¹³C-lignocellulose, while F/T cycles led to preferential use of ¹⁴C-glucose.

How to cite: Nájera, F., Dippold, M., Boy, J., Seguel, O., Köster, M., Stock, S., Merino, C., Kuzyakov, Y., and Matus, F.: Soil carbon dynamic after freezing/thawing and drying/wetting in a temperate forest soil: Dual labeling of 13C and 14C, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-10528, https://doi.org/10.5194/egusphere-egu2020-10528, 2020.

Glacier and ice sheet beds represent important yet underresearched cryospheric ecosystems. Life in the subglacial environment is mostly dependent on organic matter (OM) overridden by ice during times of glacier advance, and the nature of subglacial OM is, therefore, likely to drive the ecosystem functionality. Here we describe the origin, degradation stage and environmental context of OM present underneath glaciers in the circum-Arctic, and their effects on the resident microbial communities.

In total, 19 glaciers from Alaska, Greenland, Iceland, Svalbard, and Norway were sampled for subglacial sediments. Biomarker analysis of the sediment samples was conducted using total solvent extraction, and copper (II) oxide (CuO) oxidation techniques yielding lipids and lignin-derived phenols. The extracts were analyzed by GC-MS to characterize the molecular-level composition of OM present. The biomarker data were then placed in the context of other environmental data, such as radiocarbon age, nutrient contents, and microbial community composition. The majority of OM in the samples was plant-derived, suggested by the dominance of long-chain n-alkanols over the microbial-specific short-chain n-alkanols. The composition of long-chain n-alkanes (≥C₂₀), used as biomarkers for vascular plant waxes, in the solvent extracts suggested grass sources for samples from most Greenland glaciers and conifer sources for some glaciers from Norway, Alaska, and Disko Island (Qeqertarsuaq) in West Greenland. The rest of the OM in the subglacial samples was identified to have more general tree sources. The carbon preference index (CPI) of long-chain n-alkanes suggested a high degradation stage in most samples and was correlated with the radiocarbon age of the sediments’ OC (r = -0.68). Sediments containing older and more degraded OM were found to host less diverse microbial communities compared to those of the younger sites.

In a rapidly warming climate, previously glacier-covered areas are being exposed as a consequence of glacier recession. This new land is standing at the onset of ecological succession and pedogenesis. Our results contribute to the understanding of the potential ecological function of subglacial OM as an important source of carbon and driver of microbial community development after deglaciation in the circum-Arctic region.

How to cite: Vinsova, P., Simpson, M. J., Yuan, T., Hajdas, I., Falteisek, L., Kohler, T. J., Yde, J. C., Zarsky, J. D., and Stibal, M.: The origin and fate of organic matter in circum-Arctic subglacial ecosystems, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-5530, https://doi.org/10.5194/egusphere-egu2020-5530, 2020.

Dark, that is, nonphototrophic, microbial CO₂ fixation occurs in a large range of soils.
However, it is still not known whether dark microbial CO₂ fixation substantially contributes
to the C balance of soils and what factors control this process. Therefore,
the objective of this study was to quantitate dark microbial CO₂ fixation in temperate
forest soils, to determine the relationship between the soil CO₂ concentration and
dark microbial CO₂ fixation, and to estimate the relative contribution of different
microbial groups to dark CO₂ fixation. For this purpose, we conducted a ¹³C-CO₂ labeling
experiment. We found that the rates of dark microbial CO₂ fixation were positively
correlated with the CO₂ concentration in all soils. Dark microbial CO₂ fixation
amounted to up to 320 μg C kg⁻¹ soil day⁻¹ in the Ah horizon. The fixation rates were
2.8–8.9 times higher in the Ah horizon than in the Bw1 horizon. Although the rates of
dark microbial fixation were small compared to the respiration rate (1.2%–3.9% of the
respiration rate), our findings suggest that organic matter formed by microorganisms
from CO₂ contributes to the soil organic matter pool, especially given that microbial
detritus is more stable in soil than plant detritus. Phospholipid fatty acid analyses
indicated that CO₂ was mostly fixed by gram-positive bacteria, and not by fungi. In
conclusion, our study shows that the dark microbial CO₂ fixation rate in temperate
forest soils increases in periods of high CO₂ concentrations, that dark microbial CO₂
fixation is mostly accomplished by gram-positive bacteria, and that dark microbial
CO₂ fixation contributes to the formation of soil organic matter.

Reference

Spohn M, Müller K, Höschen C, Mueller CW, Marhan S. Dark microbial CO₂ fixation in temperate forest soils increases with CO₂ concentration.
Glob Change Biol. 2019;00:1–10. https ://doi.org/10.1111/gcb.14937

How to cite: Spohn, M., Müller, K., Höschen, C., Müller, C. W., and Marhan, S.: Dark microbial CO2 fixation in temperate forest soils increases with CO2 concentration, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-3537, https://doi.org/10.5194/egusphere-egu2020-3537, 2020.

Grazing is an important human activity affecting grassland ecosystems. Many studies have shown that grazing changed the carbon (C) cycle of grasslands, but it is still not clear how grazing will affect the recent photosynthetic C allocation in the temperate grasslands. To clarify this question, a situ field ¹³C labeling experiment was carried out in the temperate grasslands of Inner Mongolia, North China, in 2015. In this study. Grazing included 3 intensities of no grazing, medium grazing and heavy grazing. Eighty-one days after the labeling, the plants allocated more recent assimilated ¹³C (6.52% of recovered ¹³C) to shoots under medium grazing than that of no grazing (5.60%) and heavy grazing (5.40%). The most ¹³C was allocated to the belowground (roots, soil and soil respiration) under no grazing (40.68%). However, within the belowground pools, 1.36% and 17.33% of ¹³C were stored in roots and soil under medium grazing which was twice than that under no grazing and heavy grazing, which could be explained by intermediate disturbance hypothesis. ¹³C labeling experiment demonstrated medium grazing increased C assimilates by two processes:(Ⅰ) the highest total C input into plants and soil and (Ⅱ)the least C loss by soil microbial respiration (3.20%) than no grazing grassland (5.19%) and heavy grazing grassland (3.47%). The turnover rate of soil assimilates under the no grazing (0.25 ± 0.07 day^-1) was higher than that of grazing (medium grazing 0.059 ± 0.01 day^-1; heavy grazing 0.064 ± 0.02 day^-1). Overall, the no grazing isn’t the best for carbon accumulation and the medium grazing which promotes C input and C sequestration is the most suitable grazing intensity of temperate grassland in China.

How to cite: Zhao, Y. and Tian, Y.: Effect of grazing on photosynthetic carbon allocation in a temperate grassland, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-6814, https://doi.org/10.5194/egusphere-egu2020-6814, 2020.

Understanding the source partitioning of carbon dioxide (CO₂) and nitrous oxide (N₂O) fluxes from soil is integral for the characterization of total fluxes and the quantification of potential soil organic matter priming effects. Additionally, we utilized ¹⁵N-N₂O site preference data to analyze the process priming of microbial nitrification and denitrification on subsequent N₂O fluxes. A 32-day laboratory incubation was designed to examine the effects of artificial exudate, nitrogen fertilizer and their potential interactive effects on CO₂ and N₂O fluxes, soil organic matter source-priming and N₂O process-priming. Artificial root exudate (ARE) consisting of a mixture of 99 atom% ¹³C labelled compounds at three addition rates (0, 6.2, 12.5 mg C kg^-1 soil day^-1) was applied daily for 21 days to microcosms with or without urea fertilizer, a subset of which was labelled with 5 atom % ¹⁵N. Measurements of CO₂ and N₂O fluxes, isotopic composition and N₂O site preference were frequent throughout the duration of the experiment. Source partitioning of CO₂ fluxes showed that soil organic carbon (SOM-C) positive priming was significantly altered by additions of artificial exudate and urea (p < 0.001 and 0.001, respectively). When applied concurrently, urea addition had an antagonistic interactive effect on SOM-C sourced CO₂ fluxes (p < 0.001).  Source partitioning of N₂O flux data revealed that soil organic matter nitrogen (SOM-N) was positively primed for N₂O flux by the addition of urea fertilizer (p < 0.001), but positive SOM-N priming was reduced by an antagonistic interaction with artificial exudate application (p < 0.001). Further, examination of ¹⁵N-N₂O site preference found that the main processes by which N₂O is formed (nitrification and denitrification) were differentially process-primed by the addition or absence of ARE. Cumulative denitrification and nitrification contributions to total N₂O flux were both positively primed in the soils receiving both ARE and urea inputs relative to a control (50.0 ± 10.1 and 28.2± 8.0 μg N₂O-N kg^-1, respectively). In soils receiving only ARE application, denitrification-derived N₂O was negatively primed relative to a control and thus contributed less to overall N₂O flux (-9.5 ± 12.4 μg N₂O-N kg^-1) but nitrification-derived N₂O was positively primed (17.2 ± 9.0 μg N₂O-N kg^-1).

How to cite: Daly, E. and Hernandez Ramirez, G.: Source-process partitioning of soil N2O and CO2 production: nitrogen and simulated exudate additions, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-2221, https://doi.org/10.5194/egusphere-egu2020-2221, 2020.

In vegetated soils,plants naturally release root exudates, consisting of sugars, organic acids, and amino acids, into the soil increasing soil enzymatic activity.  Liberty State Park, located in Jersey City, New Jersey, is an industrial brownfield contaminated with heavy metals and organic pollutants.  Some sites have soils that function poorly, as indicated by low soil enzymatic activity, and do not support plant growth.  This study will determine whether different combinations of artificial root exudates increase soil enzymatic activity in these contaminated and low functioning soils. Different combinations of sugars, organic acids, and amino acids and will be added to barren, poorly functioning soil.  Three soil enzymatic activities will be examined at several time points over 120 days to assess the impacts of different combinations of root exudates on soil function. Further, soil microbial community composition will be determined to examine whether different artificial exudate solutions result in changes in soil microbial community.  Preliminary results suggest that the combination of sugars, organic acids, and amino acids greatly increased phosphatase, cellobiohydrolase, and L-leucine amino peptidase activity over time in poorly-functioning, barren soil from Liberty State Park. The other combinations (sugars and organic acids, sugars and amino acids, organic acids and amino acids) also increase the three enzyme activities more than the individual groups. Dormant microbes in barren soil can possibly be revived with the addition of artificial root exudates to mimic the presence of plants in revitalizing the microbial communities and improving soil function. 

How to cite: Hagmann, D., Krumins, J., Vaidya, B., and Goodey, N.: Activation of soil enzymes by addition of artificial root exudate combinations , EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-22169, https://doi.org/10.5194/egusphere-egu2020-22169, 2020.

Knowledge about the nexus between litter decomposition and soil organic matter formation is still scarce, likely because litter decomposition studies are often conducted in the absence of mineral soil. Even if mineral soil is considered, variations in soil texture, which should substantially influence decomposition and soil C sequestration via, e.g., different capacities to store C or host microbial communities, have been neglected. Here, we examined the effect of soil texture on litter decomposition and soil organic matter formation by incubating sand- and clay-rich soils. These soils, taken under C3 vegetation, were amended with C4 litter to trace the fate of organic matter newly entering the soil. While we found only small amounts of litter-derived carbon (C) in the mineral soils after our six-month experiment, the microbial activity and amount of remaining litter between the sand- and clay-rich soils substantially differed. A high microbial activity combined with higher amounts of litter-derived C and a higher remaining litter mass in the clay-rich soil indicate a more effective transformation of litter to soil organic matter as compared to the sand-rich soil. In the sand-rich soil, microbial activity was lower, less soil C was litter-derived, and the litter lost more of its mass. We explain the apparently contradictory results of higher microbial activity and concurrently higher C contents with a more effective microbial pathway of SOM formation in the clay-rich soil. Our results indicate that soil texture does not only play a role in the provision of reactive surfaces for the stabilization of C but will also affect the decomposition of litter via effects on microbial activity, ultimately determining if litter C is transferred to the soil or respired to the atmosphere.

How to cite: Angst, G., Pokorný, J., Meador, T., Hajek, T., Frouz, J., Prater, I., Mueller, C. W., van Buiten, G., and Angst, Š.: Soil texture mediated microbial activity affects the transfer of litter-derived carbon to soil organic matter, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-10898, https://doi.org/10.5194/egusphere-egu2020-10898, 2020.

Soil organic matter (SOM) originates predominantly from above and belowground OM inputs derived from plants. Although the formation of SOM is well studied, it still remains unclear how the biochemical composition of litter affects the formation of new SOM as well as the degradation of “native” SOM. In the present study we aimed to disentangle the effect of plant litter composition on C transference from different plant tissues into specific SOM fractions and to determine the magnitude of priming effect on native SOC caused by litter amendments. To this end, we individually incubated ¹³C enriched Eucalyptus spp. major litter types (bark, leaves, twigs and roots) in soil (0–20 cm) of a sandy-clay loam (Haplic Ferralsol - Brazil). Additionally, a soil sample without plant residue addition was incubated as a control. The samples were incubated at 80% of their water-holding capacity at 25 ºC for 200 days. Soil respiration was assessed along the incubation period through headspace gas sampling and ¹³C/¹²C–CO₂ analysis in a cavity ring-down spectrometer. After the incubation, soil subsamples were physically fractionated using a combined density-particle size separation method. The total C and the δ ¹³C of each soil organic matter fraction were measured and the litter-C contribution for each SOM fraction was assessed using a two-end member isotope mixing model. The molecular composition of the incubated plant material and SOM fractions were determined by solid-state ¹³C-CPMAS-NMR spectroscopy. Interestingly, we found no significant differences for total SOM contents among the different treatments. Conversely, incubation without litter amendment (control treatment) resulted in lower total SOM contents, indicating mineralization of “native” SOM along the incubation period. The partitioning of litter-derived C into SOM fractions indicated that leaves litter were preferentially transferred to mineral associated organic matter (MAOM), while roots contributed more to particulate organic matter (POM). Cumulative C-CO₂ evolution from the treatments over the incubation period increased in the following order: twigs > leaves > bark > roots > controls. Incubation with twigs, bark and roots significantly increased “native” SOM respiration, while the treatment with leaves addition did not differ from the control. When tracing the source of “native” SOM-derived CO₂, we observed a similar amount of C being respired from MAOM, regardless the treatment, while incubation with twigs, bark and roots resulted in higher respiration of “native” SOM-derived C from POM. Our data demonstrates that the biochemical composition of plant litter determines the fate of newly formed organic matter (MAOM or POM) and controls the degradation of “native” SOM. Therefore, plant residues enriched in more easily degradable compounds (leaves) are preferentially transferred to MAOM and causes less native SOC priming. On the other hand, plant residues enriched in structural compounds (twigs, bark and roots), are preferentially respired or allocated into the POM, also resulting in higher priming effect intensity.

How to cite: Januário Almeida, L. F., Colocho Hurtarte, L. C., Teixeira, P. P., Inagaki, T. M., de Souza, I. F., da Silva, I. R., and Mueller, C. W.: Biochemistry of plant litter types drives differentiation into particulate and mineral-associated soil organic matter and determines the magnitude of priming effect, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-11168, https://doi.org/10.5194/egusphere-egu2020-11168, 2020.

Approximately 50–70% of C stored in soil is derived from the roots or root-associated microorganisms, reflecting their importance for soil organic matter (SOM) formation. Microorganisms are the major driver for SOM decomposition and their activities are modified by the input of labile organics such as root exudates and low molecular weight organic substances derived from litter decomposition. Such short-term changes in the turnover of SOM caused by moderate organic additions into the soil were defined as priming effects (PE). Priming effects can influence global carbon (C) storage in soil and lead to climate feedbacks by accelerating the decomposition of organic matter (OM). In natural ecosystems, input of nitrate (NO₃⁻) and ammonium (NH₄⁺) into the soil can be derived from nitrogen (N) deposition and biological N fixation. Although availability N can alter the magnitude and direction of priming, it remains unclear whether additions of NO₃⁻ and NH₄⁺ have distinct effects on the decomposition of OM. Thus, the aims of this study were to investigate the responses of OM decomposition along a decay continuum (i.e. decreasing decomposition degree) to labile C and N inputs and determine the PE induced by the two N forms. Leaf litter, wood litter, organic soil horizon, and mineral soil, with a broad range of C:N ratios were collected along a decay continuum in a typical subtropical forest and incubated for 38 days with ¹³C labeled glucose and N (NO₃⁻) additions. Based on the very broad range of C:N ratios in OM in soil and inputs of labile C and N, we demonstrated the OM decomposition within a decay continuum as well as PE intensities and the thresholds for the switch of PE directions. In contrast to NH₄⁺ additions, NO₃⁻ generally accelerated the decomposition of all OM. Priming of plant litter was dependent on the C:N ratios of the labile inputs. However, leaf litter decomposition was more controlled by N addition than wood litter. Glucose addition greatly increased the priming of OM decomposition, demonstrating energy limitation for microorganisms. Distinct priming patterns were observed between NO₃⁻ and NH₄⁺ additions, both for the individual OM types and for all four types of OM. The PE induced by labile C and N inputs can increase or reduce C sequestration depending on C:N stoichiometric ratios of labile inputs. Net C losses caused by PE can be observed in organic soil and plant litter with low C and N additions, but all four OM substrates increased C sequestration under high C addition. Minor differences in priming along the continuum were observed where the OM C:N ratio was below 30 when NO₃⁻ was added and where the labile C:N ratio was less than 55 when NH₄⁺ was added. Thus, changes in the composition of deposited N (atmospheric deposition and fertilization) may induce distinct climate feedbacks. Under future climatic conditions by global warming and elevated CO₂, more labile C inputs via root exudates could accelerate litter decomposition. Effects of N however, depend on the N form: NH₄⁺ to NO₃⁻ due to the energy necessary for microorganisms for NO₃^- reduction.

How to cite: Liu, M., Qiao, N., Xu, X., Fang, H., Wang, H., and Kuzyakov, Y.: C:N stoichiometry of stable and labile organic compounds determine priming patterns, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-22306, https://doi.org/10.5194/egusphere-egu2020-22306, 2020.

Introduction & objectives: Over ten thousand years, soils have been formed through events of volcanic ash deposition in Hokkaido, Japan. The soil organic matter (SOM) in the past surface layer has been buried in the deeper soil. The buried humic horizons serve as a large carbon (C) reservoir. The SOM in the deeper soil horizons is preserved due to lower microbial activities and limited inputs of fresh organic matters. However, when the buried humic horizons are exposed to the surface by deep plowing and bottom plow tillage, decomposition of the exposed SOM may be accelerated through priming effects, due to the increased supply of low-molecular-weight (LMW) substances from fresh plant litter inputs. To test this, we examined glucose concentration dependency of priming effect and the change of SOC balance through priming effect using ¹³C tracer incubation.

Materials & methods: Soil samples were collected from the volcanic soil profiles in pasture site and adjacent forest sites in Hokkaido, Japan. The moist soils were sieved (< 4 mm) to eliminate plant debris and stones for the incubation study and the other analysis. A ¹³C-glucose solution (99 atom%; 0 – 3.9 mg glucose g^-1) was added to moist soil (equivalent to 10 g oven-dried weight) and incubated at 20ºC in the dark for 30 days. The head space gas sample was periodically taken into the vial, and ¹³CO₂ and ¹²CO₂ concentrations were determined by GC-MS. Priming effect (PE) was calculated by subtraction between the amounts of ¹²CO₂ with and without glucose. The head space gas in the bottle was flush out and replaced to CO₂-free-air every sampling time. We also measured soil microbial biomass C (MBC) by chloroform fumigation method, bacterial and fungal biomass by 16S and 18S rRNA genes targeted real-time PCR, SOC concentrations, inorganic N concentrations (ammonium and nitrate) and the other physicochemical properties of the soil profiles.

Results & discussion: Glucose addition induced the positive PEs in the buried humic soil samples of both sites, and the magnitudes of PEs (cumulative primed-CO₂ amounts) in the buried humic soil samples were 0.4 to 1.5 times as those in the surface soils. However, the negative PEs were detected in the forest surface soil, probably because of low soil pH and relatively high inorganic N concentration. The magnitudes of PEs were dependent on added glucose concentrations for all the soils, and the threshold between negative and positive PEs corresponded to 3.5 % of glucose-C relative to MBC in the forest surface soil. The positive correlation between evolution rates of primed-CO₂ significantly and bacterial or fungal biomass suggests both bacteria and fungi contributes to PE in the soils studied. Even if glucose addition induced PE, total SOC after incubation increased when glucose-C was added more than 0.5 mg C g^-1 in the all soils. This implies that the optimized fresh litter input can control priming effects and C sequestration in volcanic soils.

How to cite: Hayakawa, C., Kobayashi, T., Fujii, K., Inagaki, Y., and Senoo, K.: Glucose concentration controls priming effects and soil carbon storage under pasture and forest in volcanic ash soils of Hokkaido, Japan, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-11990, https://doi.org/10.5194/egusphere-egu2020-11990, 2020.

Soil organic matter (SOM) stabilization plays an important role in long-term storage of carbon (C). However, now many ecosystems are experiencing global climate change, which could change soil C balance through affecting the C input via plant community shifts, and C losses via SOM decomposition. In subarctic ecosystems, plant community composition and productivity are shifting because of climate change. This change of above-ground communities will affect rhizosphere input such as low molecular weight organic substances (LMWOS), which can affect microbial decomposer activities and subsequent contribution to SOM mineralization (priming effect). In the present study, we simulated climate change with N fertilization, to represent a warming enhanced nutrient cycling, and litter input, to simulate arctic greening, to evaluate the effect of a changing climate on subarctic ecosystems in Abisko, Sweden. The 6 sampled field treatments included three years of chronic N addition (5 g N m^-2 y^-1), three years of chronic litter addition (90 g m^-2 y^-1), three years of chronic N and litter additions, one year of high N addition (15 g N m^-2 y^-1), one year of high litter addition (270 g m^-2 y^-1) and a control treatment. All treatments were established in 1×1 m experimental squares and had 6 replicates. We resolved effects on plant community (NDVI), SOM mineralization, microbial composition, bacterial and fungal growth rates, and soil properties.

We found that N treatments changed plant community and stimulated productivity and that the associated increase in belowground LMWOS induced shifts in the soil microbial community. This coincided with a tendency for a shift towards bacterial dominated decomposition (low fungi/bacterial growth ratio) and a microbial community that had shifted from gram-positive bacteria to gram-negative bacteria; a shift often observed when comparing bulk with rhizosphere conditions. However, N treatments had no effect on SOC mineralization, but did increase soil gross N mineralization. This shift in the C/N of mineralisation might be because N treatments accelerated the growth of fast growing plant species with higher nutrient content, whose litter input provided microbes with fresh OM richer in N.

These responses in belowground community and processes driven by rhizosphere input prompted the next question: how did the simulated climate change affect the susceptibility of SOM to priming by LMWOS? To assess this question and explore the microbial mechanisms underpinning priming of SOM mineralization, we added a factorial set of additions including ¹³C-glucose with and without mineral N, and ¹³C-alanine semicontinously to simulate the effect of belowground LMWOS input on SOM mineralization and microbial activity, and investigate how the SOM priming was linked to the actively growing microorganisms. Therefore, we incubated these samples for 7 days, treated with ¹³C LMWOS, and measured SOC and SON mineralization to assess SOM priming, bacterial and fungal growth rates, microbial phospholipid fatty acids (PLFAs) and ¹³C-PLFA enrichment, as well as the microbial C use efficiencies to assess microbial responses to LMWOS additions.

How to cite: Na, M., Yuan, M., Hicks, L., and Rousk, J.: How does simulated climate change affect the susceptibility of SOM to priming by LMWOS in the Subarctic?, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-4817, https://doi.org/10.5194/egusphere-egu2020-4817, 2020.

Extreme precipitation events resulting from climate change have strong impact on structure and functions of grassland ecosystems. The extreme climate events may shift plant productivity and nutrient acquisition preferences by roots and microorganisms.We conducted an extreme precipitation simulation experiment and used in-situ ¹⁵N labeling of the three N forms to investigate N acquisition (N uptake rate, ¹⁵N recovery and preference for N form) by the dominant plant species Stipa grandis and soil microorganisms.Increased rain frequency raised the growth and N acquisition of S. grandis, while microbial N uptake remains unaffected. Microorganisms strongly outcompeted S. grandis for total ¹⁵N acquisition, however such superiority decreased in higher extreme precipitation frequency. Plant and microorganisms converged their N demands from distinct to similar preferences for N forms with high precipitation frequency. Such chemical niche partitioning by extreme precipitation effectively reduced root and microbial competition for each N form. Overall, important mechanistical insights into chemical niche differentiation by the effects of extreme climate events and their effects on structure, functions and plant-microbial interactions in temperate grasslands were explained.

How to cite: Tian, Y.: Extreme precipitation increases plant biomass through altering nitrogen acquisition by grasses and soil microorganisms, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-3243, https://doi.org/10.5194/egusphere-egu2020-3243, 2020.

Boreal forest productivity on permafrost is limited by availability of soil nitrogen (N) in the active layer. Low soil temperature and summer flooding limit microbial N mineralization on shallow permafrost table. Uptake of amino acids by plant root-mycorrhizal association is known to mitigate N limitation in boreal forest soils. However, amino acid hypothesis can not fully explain advantage of black spruce trees in drunken forests due to competition of amino acids between plants, bryophytes, and microbes. Based on the observation of urea accumulation in deeper soil, we test another hypothesis that black spruce trees take up intact urea in deeper soil. Mixture solutions (glutamic acid, urea, ammonium, nitrate), with only one N form labeled by ¹³C and/or ¹⁵N, was injected into the organic/mineral soil layers. We compared two black spruce forest sites with/without shallow permafrost table in northern Canada. We found that black spruce trees take up intact urea as well as amino acids in the shallow permafrost sites. Urea accumulation is explained by low microbial activities to mineralize ¹⁴C-labeled urea. The other plants or bryophyte compete with black spruce for amino acids, but not for urea. Since the other black spruce trees in the deeper soil sites rely on amino acids and inorganic N, urea uptake strategy is specific to black spruce trees on shallow permafrost table. The root expansion on hummocky microrelief provides opportunity for leaning trees to access urea. The uptake of intact urea could be one of strategy of black spruce trees to mitigate N limitation in permafrost-affected hummocky soils.

How to cite: Fujii, K., Matsuura, Y., Inagaki, Y., and Hayakawa, C.: Uptake of urea by “drunken” trees on permafrost , EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-6271, https://doi.org/10.5194/egusphere-egu2020-6271, 2020.

Keywords: Soil reaction, analytical pyrolysis, soil respiration, carbon stabilization

During the last decade, soil organic matter dynamics and its determining factors have received increased attention, mainly due to the evident implication of these parameters in climate change understanding, predictions and possible management. High-mountain soil could be considered as hotspot of climate change dynamic since its high carbon accumulation and low organic matter degradation rates could be seriously altered by slight changes in temperature and rainfall regimes associated to climate change effects. In the particular case of Sierra Nevada National Park, this threat could be even stronger due to its Southern character, although its elevated biodiversity could shed some light on how could we predict and manage climate change in the future.

In this study, a quantitative and qualitative organic matter characterization was performed and soil microbial activity measured to evaluate the implication of pH and vegetation in soil organic matter dynamics.

The sampling areas were selected according to vegetation and soil pH; with distinct soil pH (area A with pH<7 and area B with pH>7) and vegetation (high-mountain shrubs and pine reforested area). Soil samples were collected under the influence of several plant species representatives of each vegetation series. Six samples were finally obtained (five replicates each); three were collected in area A under Juniperus communis ssp. Nana (ENE), Genista versicolor (PIO) and Pinus sylvestris (PSI) and other three were collected in area B under Juniperus Sabina (SAB), Astragalus nevadensis (AST) and Pinus sylvestris (PCA).

Qualitative and quantitative analyses of soil organic matter were made to establish a possible relationship with microbial activity estimated by respiration rate (alkali trap) and fungi-to-bacteria ratio using a plate count method. Soil easily oxidizable organic carbon content was determined by the Walkley-Black method (SOC %) and organic matter amount was estimated by weight loss on ignition (LOI %). Analytical pyrolysis (Py-GC/MS) was used to analyse in detail the soil organic carbon composition.

Our results showed that the microbial and therefore the dynamics of organic matter is influenced by both, soil pH and soil of organic matter. So that the pH in acidic media prevail as a determining factor of microbial growth over soil organic matter composition conditioned by vegetation.

Acknowledgement: Ministerio de Ciencia Innovación y Universidades (MICIU) for INTERCARBON project (CGL2016-78937-R). N.T. Jiménez-Morillo and L. San Emeterio also thanks MICIU for funding FPI research grants (BES-2013-062573 and Ref. BES-2017-07968). Mrs Desiré Monis is acknowledged for technical assistance.

How to cite: González-Pérez, J. A., Bárcenas.Moreno, G., Jiménez-Morillo, N. T., Colchero-Asensio, M., San Emeterio, L. M., and de la Rosa, J. M.: Effect of Ph and vegetation cover in soil organic matter structure at a high-mountain ecosystem (Sierra Nevada National Park, Granada, Spain), EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-8246, https://doi.org/10.5194/egusphere-egu2020-8246, 2020.

Afforestation with pure and mixed-species is an important strategy to improve soil organic carbon (SOC) stocks and restore degraded lands. However, what remains unclear is the stability of SOC to microbial degradation after afforestation and the effect of tree species composition. Moreover, it is important to reveal how sensitive the SOC in afforestation lands is to environmental changes, such as warming. To study the combined effects of warming and the tree species composition on decomposition of SOC by microorganisms and enzyme activities, soils were collected from the monocultural and mixtures of Silver birch (Betula Pendula) and European beech (Fagus Silvatica) (BangorDiversity, UK, 12 years since afforestation) and were incubated for 169 days at 0, 10, 20, 30 °C at 60 % of WHC. The field experiment is arranged into a completely randomized design with n=4. The CO₂ efflux was measured constantly, whereas activities of β-glucosidase, chitinase and acid phosphatase, and content of microbial biomass C (MBC) were obtained at the end of the incubation. Results showed that soil cumulative CO₂ efflux increased by 34.7–107% with the temperature. Potential enzyme activities were dependent on tree species composition. Warming, but not tree species exhibited a significant impact on the temperature sensitivity (Q10) of soil cumulative CO₂ efflux and enzyme activities. The greatest temperature sensitivity (Q₁₀) of total CO₂ efflux was found at 10–20 °C and was 2.0–2.1, but that of enzyme activities were found as 0.9–1.1 at 0–10 °C. These results suggest that warming has an asynchronous effect on the SOC decomposition and enzyme activity, and enzymes cannot account for the temperature sensitivity of soil respiration. Thus, thermal adaptations of SOC mineralization is independent of the adaptation of the enzyme pool.

How to cite: Jiang, Z., Gunina, A., Merz, L., Yang, Y., Kuzyakov, Y., Jones, D., Smith, A. R., and Ludwig, B.: Loss of available soil organic carbon from afforestation plots: effect of tree species composition and warming, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-19705, https://doi.org/10.5194/egusphere-egu2020-19705, 2020.

Soil microbial necromass represents a significant proportion (>50%) of soil organic matter (SOM). Microbial necromass consists mainly of particulate organic residues from fragmented cells walls and other slow turnover cytoplasmic components of dead fungi and bacteria. Some of the key components of microbial cell walls, such as peptides and amino sugar polymers, can remain and accumulate in the soil over prolonged times. Amino sugars have been used as biomarkers to quantify the contribution of microbial necromass to stabilized SOM. The different amino sugars present in polymeric form in soils can be released by acid hydrolysis and allow the estimation of the contribution of both fungal and bacterial necromass to the SOM pool. Among the amino sugars, hexosamine isomers (glucosamine, galactosamine, mannosamine) and muramic acid (the ether of lactic acid and glucosamine) are the most abundant ones. Muramic acid is specific to bacterial peptidoglycan while glucosamine is an abundant cell wall component of both, fungal chitin and bacterial peptidoglycan.

There are several chromatographic methods to measure free and bound amino sugars and amino acids in soil extracts and soil hydrolysates, but none of them allow the combined determination of amino sugar biomarkers and amino acids simultaneously in a single assay for rapid analysis. This is important as a large fraction of soil necromass N (>50%) consists of non-amino sugar-N, such as proteins and nucleic acids. In this study we therefore adopt a method based on 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AccQ.Tag) derivatization of amino compounds and optimized chromatographic (reversed phase) separation to simultaneously measure amino sugars (isomers) and amino acids in soil extracts and soil hydrolysates using ultra-high-performance liquid chromatography coupled to fluorescence or UV detection.

The use of this method allows for fast, robust and highly sensitive quantification of amino acids and amino sugars in environmental samples at sub-micromolar levels. This approach will help to improve our understanding of soil microbial necromass dynamics and their inherent effect on soil C and N sequestration. The AccQ.Tag chemistry also allows compound detection by electrospray ionization (ESI)-mass spectrometry, enabling isotope (¹³C, ¹⁵N) tracing applications.

How to cite: Salas, E., König, A., Kaiser, C., and Wanek, W.: A new method to measure amino sugar isomers and amino acids in soil extracts and soil hydrolysates based on AccQ.Tag-chemistry and reversed phase ultra-high-performance liquid chromatography, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-8200, https://doi.org/10.5194/egusphere-egu2020-8200, 2020.

The activities of soil microorganisms drive soil carbon (C) and nutrient cycling and therefore play an important role in local and global terrestrial C dynamics and nutrient cycles. Unfortunately, soil microbial activities have been defined mostly by measurements of heterotrophic respiration, potential enzyme activities, or net N processes. However, soil microbial activities comprise more than just catabolic processes such as respiration and N mineralization. Recently anabolic processes (biosynthesis and growth) and the partitioning between anabolic and catabolic processes in soil microbial metabolism have gained more attention as they control microbial soil organic matter formation. Understanding the controls on these processes allows an improved understanding of the key roles that soil microbes play in organic matter decomposition (catabolic processes) and soil organic matter sequestration (anabolic processes leading to growth, biomass and necromass formation), and their potential feedback to global change.

Generally, there are two approaches to study the metabolism of soil microbial communities: First, position-specific isotope labeling is a tool that allows the tracing of ¹³C-atoms in organic molecules on their way through the network of metabolic pathways and second, metabolomics and fluxomics approaches can enable disentangling the highly complex metabolic networks of microbial communities, which however have rarely (metabolomics) or never (fluxomics) been applied to soils.

In this study we developed a targeted soil metabolomics approach coupled to ¹³C isotope tracing (fluxomics), in which we extract, purify and measure a preselected set of key metabolites. Our aim was to cover the wide spectrum of soil microbial metabolic pathways based on the analysis of biomarker metabolites being unique to specific metabolic pathways such as  glycolysis/gluconeogenesis (e.g. fructose 1,6-bisphosphate), the pentose phosphate pathway (ribose-5-phosphate), the citric acid cycle (α-ketoglutaric acid), purine and pyrimidine metabolism (UMP, AMP, allantoin), amino acid biosynthesis and degradation (10proteinogenic amino acids and their intermediates), the urea cycle (ornithine), amino sugar metabolism (N-Acetyl-D-Glucosamine and –muramic acid) and the shikimate pathway (shikimate). The minute concentrations of these primary metabolites are extracted from soils by 1 M KCl including 5 % chloroform, salts are removed by freeze-drying, methanol dissolution and cation-/anion-exchange chromatography and the metabolites and their isotopomers quantified by UPLC-Orbitrap mass spectrometry. To cover the wide range of metabolites, compound separations are performed by  hydrophilic interaction chromatography (HILIC) for metabolites such as amino acids, (poly-)amines, nucleosides and nucleobases and by Ion chromatography (IC), to separate charged molecules like amino sugars, sugar phosphates and organic acids.  Here we will show fluxomics results from a laboratory soil warming experiment where we added ¹³C-glucose to a temperate forest soil as a proof of concept.

How to cite: Böckle, T., Hu, Y., Schnecker, J., and Wanek, W.: A novel fluxomics approach to decipher the flux partitioning between anabolic and catabolic processes in soil microbial communities, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-9150, https://doi.org/10.5194/egusphere-egu2020-9150, 2020.

Phospholipid fatty acids (PLFA) are widely used as biomarkers for soil microbial biomass. In more recent years, neutral lipid fatty acids (NLFA) have additionally been used as storage biomarkers. Both lipid classes are usually separated via silica solid phase extraction (SPE) after extraction with a mixture of chloroform, methanol and citric acid buffer. However, in recent years several studies reported incomplete or inconsistent separation of lipid classes, depending on minor differences in the polarity of the eluents used during the SPE. Moreover, while PLFA profiles have been tested on microbial pure cultures, the taxonomic specificity of NLFA is only assumed to equal that of PLFA.

Complementary to fatty acid based biomarkers, many studies quantify ergosterol as a reliable indicator for fungal biomass because the fungal-specific PLFA 18:1ω9 and 18:2ω6,9 also occur in plants, which compromises their use for detecting fungal biomass in plant tissue (for example mycorrhizal fungi in plant roots). Measuring ergosterol requires an additional extraction method, but existing protocols include silylation for further gas chromatography analysis and are thus not compatible with determining ¹³C by IRMS.

Here, we aimed to quantify the recovery of polar and non-polar lipid classes as well as ergosterol following lipid extraction and silica SPE fractionation. We used pure standards of representative phospholipids, glycolipids and neutral lipids with unique fatty acid chain lengths for unambiguous identification of the lipid class after SPE. Lipid fractionation was tested on a 96-well SPE plate with different eluents. Subsequently, we applied the modified method to characterize lipid fractions in microbial pure cultures from bacteria (Proteobacteria, Firmicutes, Actinobacteria), and saprotrophic and ectomycorrhizal fungi (Ascomycota, Basidiomycota).

Separation of lipid classes was achieved by successively eluting NLFA and sterols with a mixture of chloroform and ethanol (v:v = 98:2), glycolipid fatty acids (GLFA) with acetone, and PLFA with a mixture of methanol, chloroform and water (v:v:v = 5:5:1). GLFA were partially recovered in the NLFA or PLFA fraction depending on the nature of the lipid, which should be considered when interpreting PLFA data. Ergosterol recovery was unaffected by subsequent mild alkaline methanolysis of the NLFA fraction in which it was collected, allowing further analysis of both lipid classes in the same mixture. The gas-chromatographic method may be extended to elute both NLFA and (non-silylated) sterols in one run, assuming that the concentration of ergosterol in soil samples is high enough. Therefore, the method can be optimized by using an internal standard added to the NLFA fraction and simultaneously quantify ergosterol. Finally, we show how different lipid classes and attached fatty acid chains distribute in pure cultures of soil micro-organisms.

How to cite: Gorka, S., Canarini, A., Imai, B., Teischinger, G., Darcy, S., and Kaiser, C.: A unified protocol for the high-throughput measurement of PLFA, NLFA, GLFA and sterols from soil, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-17791, https://doi.org/10.5194/egusphere-egu2020-17791, 2020.

Soil lipids encompass substances of mainly plant or microbial origin that are insoluble in water and soluble in organic solvents such as ether, hexane, benzene, chloroform or dichloromethane. This soil organic fraction is of great interest because it encompasses biomarkers associated to soil microbial communities, i.e. Gram positive/negative bacteria, mycorrhizae, actinomycetes, etc. and because of its transitory nature that provides insights into soil organic matter (SOM) dynamics and soil carbon turnover. Compound-specific isotope analysis (CSIA) have been used in biomarker studies to investigate the assimilation of carbon from external inputs into SOM. This study determined the distribution and d13C composition of fatty acids as dominant part of the soil lipid fraction to assess turnover times in agricultural practice.

Soil samples were taken from three depth intervals (0-5, 5-20, 20-40 cm) from a Mediterranean agricultural soil at “La Hampa” experimental station used for a crop rotation experiment with wheat (C3 plant) and maize (C4 plant). Using the C4 biosynthetic pathway, maize discriminates less strongly against 13C, i.e. d13C values of fatty acids originating from maize are less negative than those of fatty acids from wheat.

Soil lipids were extracted using a DCM:MeOH (3:1) solvent mixture. Fatty acids were transmethylated with MeOH:acetyl chloride (30:1) to form fatty acid methyl esters (FAMEs) while the hydroxy groups of hydroxy acids, alcohols, sterols and other compounds were silylated using BSTFA prior to analysis by gas-chromatography combustion chamber isotope ratio mass spectrometry (GC-C-IRMS) for carbon isotope ratios. Compounds were identified through their mass spectra by gas-chromatography mass spectrometry (GC-MS) and quantified by gas chromatography with flame ionization detection (GC-FID).

Only two maize harvests after wheat cultivation, a significant 13C enrichment of up to 2 ‰ was found in the saturated C20, C22 and C23 FAMEs and the mono-unsaturated C22 FAME and of up to 5 ‰ in the leaf wax-derived C29 and C31 n-alkanes relative to the control treatments without maize input. No significant differences, however, were found for alcohols and hydroxy acids. These differences may respond to the high specificity of the long-chain n-alkanes from plant origin, whereas the other compounds FAMEs, and mainly alcohols and hydroxyl acids are less specific plant markers and may have a diverse origin.

No significant differences in the isotopic composition were observed at different depths within treatments apart from a slight d13C enrichment of 1.5 ‰ in the upper soil layer (0-5 cm) in the maize plots relative to the deeper layers. It is worth noticing that SOM content remained very low (< 1.3%) over the entire duration of the experiment, with no significant differences despite the high amount of C4 biomass presumably added to the soil during the two growth periods. Together with the d13C enrichment observed in the maize plots, this points to high mineralization rates in these soils and implies both a rapid turnover of plant debris into the SOM.

Acknowledgement: Ministerio de Ciencia Innovación y Universidades (MICIU) for INTERCARBON project (CGL2016-78937-R). L. San Emeterio also thanks MICIU for funding FPI research grants (BES-2017-07968). Mrs Desiré Monis is acknowledged for technical assistance.

How to cite: M. San-Emeterio, L., Bull, I. D., Holtvoeth, J., and González-Pérez, J. A.: Compound-specific isotopic analysis of fatty acids in three soil profiles to estimate organic matter turnover in agricultural soils., EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-18526, https://doi.org/10.5194/egusphere-egu2020-18526, 2020.

Dehesa are woodlands typical of southern Mediterranean climate species modified by human to seasonal wood-pastures adapted to the unpredictability of the Mediterranean climate. Changes in climatic and environmental conditions can affect both, plant biomass chemical and isotope composition that will eventually be reflected in soil organic matter (SOM). Nowadays, many ecological studies use bulk isotope values, which represent a weighted mean average of the different necromass compounds. An isotopic characterization of individual compounds is desirable to differentiate the isotopic composition of the main plant components. Soil organic matter is composed mainly of high MW biopolymers i.e. polysaccharides, polypeptides, polypeptides, polyesters, etc. not amenable to most chromatographic techniques without the use of intense extraction and sample preparation steps.

Here, an analytical pyrolysis technique combining Py-GC with a continuous flow isotope ratio mass spectrometer (IRMS) (Py-CSIA) is described and validated for the direct study of compound specific isotope composition in soil samples.

The consistency of the Py-CSIA was tested using a standard n-alkanes mixture (dissolved C16 to C30 series with increasing concentrations along three pentads, Indiana Univ. SIL mix. Type B). The values obtained fitted to a straight line (R² > 0.999). No induced thermal cracking nor deviations from the acclaimed isotope composition (fractionation) was observed up to high pyrolysis temperature (< 500 °C).

Composite dehesa (Pozoblanco , Córdoba, Spain) surface soil samples were taken under evergreen oak canopy . A detailed SOM study was performed using conventional analytical pyrolysis (Py-GC/MS) and δ¹³C for specific compounds released after pyrolysis was done using Py-CSIA.

Well-resolved chromatograms were obtained by Py-GC/MS and a total of 40 pyrolysis compounds were detected that represented the chemical variability of soil organic matter and consisted mainly of polysaccharide, lignin-derived compounds (G- and S- units), fatty acids and n-alkanes. When coupling Py with GC-C-IRMS, many c peaks were well resolved and with a sufficient chromatographic separation to give accurate δ¹³C readings. Nonetheless, there were compounds with high δ¹³C standard deviations considered not sufficiently resolved for a reliable estimation of their isotope composition due to coelution and were discarded.

The δ¹³C for specific biomass compounds released by pyrolysis of soil was in line with the expected values for C3 plants i.e. Quercus spp. Polysaccharide derived products (furans, cyclopentanones), showed slightly enriched δ¹³C values (-26.0 ± 0.47 ‰) in accordance with their naturally ¹³C enriched composition. Although no statistical differences were found, lignin-derived units showed slightly depleted δ¹³C ( -27.4 ± 0.78 ‰). Accordingly, depleted δ¹³C values for lipids (-35.1 ± 2.41 ‰) and alkanes (-35.5 ± 2.20 ‰) were found, the latter with lighter isotope composition with increasing the hydrocarbon length.

Here we show the possibility of using this particular analytical pyrolysis technique (Py-CSIA) for the direct measurement of δ¹³C in relevant specific soil organic matter components including those from polysaccharides (cellulose/hemicellulose), lignin, lipid/waxes and also peptide/protein-derived compounds.

Acknowledgement: Ministerio de Ciencia Innovación y Universidades (MICIU) for INTERCARBON project (CGL2016-78937-R) DECAFUN (CGL2015-70123-R). L. San Emeterio also thanks MICIU for funding FPI research grants (BES-2017-07968). Mrs Desiré Monis & Mr Eduardo Gutiérrez González are acknowledged for technical assistance.

How to cite: González-Pérez, J. A., San Emeterio, L. M., González-Vila, F. J., Domínguez-Núñez, M. T., and de la Rosa, J. M.: Direct soil organic matter compound specific δ13C analysis using pyrolysis (Py-CSIA): identification of biomarkers in a dehesa from Southern Spain, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-19596, https://doi.org/10.5194/egusphere-egu2020-19596, 2020.

It is well known, that phospholipid fatty acids (PLFAs) are very dynamic, and reflect the living microbial community. The vast majority of previous studies limited its turnover determination to the lipid moiety (“the tail”) of the phospholipids. Thus, it remains unclear how dynamic the head groups of phospholipids are, and whether environmental conditions, i.e. amount of available carbon (C) have an effect on the dynamics of parts of the phospholipid molecule. To answer these questions, the double-labeling ¹⁴C/³³P-was used in the present experiment. 

The soil was collected from a 45-75 cm depth at the Klein-Altendorf experimental research station Bonn, Germany. The site is an agricultural field for more than 100 years. Formation of PLFA was traced for the two conditions: C limited (2.5 mg glucose-C kg^-1 soil added, 1% from microbial biomass C) and C rich (250 glucose-C kg^-1 soil added, 100% of MBC). For both conditions, ¹⁴C labeled glucose and ³³P-K₂HPO₄ (12.5 mg P kg^-1 soil) were added with 1 mL of water and supplemented with (NH₄)₂SO₄ (25 mg N kg^-1 soil). These ratios of C/P and C/N were chosen relative to a 100% glucose-C application to reach a ratio of C:P=20:1 and C:N=10:1. The soil was incubated for 10 d, and destructive samplings were performed after 5 h, 19 h, 1, 3, 5, 7 and 10 days, and each time four replicates were harvested. Soils were extracted for PLFAs in 2 steps: first PLFAs were obtained following the standard procedure, but both phospholipid tails and head groups were collected for further ¹⁴C and ³³P counting. The second step included separate extraction and compound-specific PLFA analysis by GC-MS to reveal changes in the community composition induced by C, N, P addition that might explain de-novo formation of phospholipids.

The peak of ³³P incorporation into headgroups under high glucose addition was after 19 h, and accounted 0.3% from the applied tracer, whereas it was up to 1% after low glucose addition and peaked in the middle of incubation time. Incorporation of ¹⁴C into the head groups and tails after high glucose addition showed an identical temporal dynamic and was 1.5 times higher in heads than in tails. Both, ³³P and ¹⁴C incorporation into head and tail had a temporal minimum at day 3 and increased afterwards suggesting two different underlying processes: direct incorporation versus C recycling. After low glucose addition, ¹⁴C incorporation was maximum on day 5 but was 3 times lower compared to growth conditions. This shows that even under limited C supply microorganisms construct new phospholipids from available glucose. Irrespective of the C supply, the ratio of head to tail incorporation relative to the ratio of head-to-tail C atoms demonstrates a significantly higher turnover of headgroup C than lipid C, suggesting recycling as an important process to cover microbial lipid demand. Thus, for the first time the different dynamics of phospholipid heads and tails was found and suggest recycling as an important process for growth and maintenance lipid formation.

How to cite: Zhao, Z., Gunina, A., and Dippold, M.: Does C accessibility have an effect on the formation of microbial cell membranes?, EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-19738, https://doi.org/10.5194/egusphere-egu2020-19738, 2020.

The ability of an agricultural soil to function and sustainably provide an increasing food supply for a rapidly increasing global population has become of vital worldwide importance. Traditionally, soil health has been determined on a physico-chemical basis with biological characteristics often being ignored. Although several biological methods have been proposed, to date, none of these methods adequately indicate soil health. One method proposed to correct these circumstances is profiling or fingerprinting the volatile organic compounds (VOCs) from soil. VOCs in soils originate from a large variety of biological sources; microbial, fungal, animal- and plant-derived. These volatilomes are vital to plant/fungi-microbe and animal/human-microbe interactions and therefore offer a potential reactive, functional diagnostic tool to determine soil health by investigating the intra and interspecies interactions.

The standard methodology for VOC profiling has been solid phase microextraction (SPME). This automated VOC extraction method allows the monitoring of the community structure, physiological state, and activity of any microbial community in a soil without the need of manual extraction or cultivation procedures. Other common techniques that could be used to monitor the VOC fingerprints from soils include high capacity sorptive extraction (HCSE) or thermal desorption using sorbent-packed tubes for passive, in-situ sampling of soil gas.

Combining each of these techniques with an innovative cryogen-free focussing and pre-concentration trap has two main advantages:

1. All extraction techniques can run on a single platform without the need to change the hardware.
2. Single (SPME-trap) and multiple extractions (SPME-trap with enrichment) can be carried out automatically on a single sample to increase the analytical sensitivity, thus achieving a comprehensive VOC profile.

In this microcosm study, soils were treated in three different ways and their VOC profiles investigated. A ‘good’ soil comprised of brown earth and compost, a ‘medium’ soil of unaltered brown earth and a ‘bad’ soil of brown earth held under eutrophic anaerobic conditions. 2 g of each soil was analysed with SPME-trap, SPME-trap with enrichment, HCSE and sorbent tubes. Both a targeted (phenol, p-cresol, isophorone, indole and trans-β-ionone) and untargeted approach indicates that there are significant differences between the different soil types. By increasing the sensitivity of the untargeted approach with SPME-trap enrichment, this study was able to extend the number of VOCs identified, allowing a much more comprehensive VOC profile and possibility to determine the actual functions of specific VOC produced by the soil microbial community.

How to cite: Brown, R., Mayser, J. P., Widdowson, C., Chadwick, D., and Jones, D.: VOC analysis in soils – Extending SPME to SPME-trap and SPME-trap with enrichment., EGU General Assembly 2020, Online, 4–8 May 2020, EGU2020-22358, https://doi.org/10.5194/egusphere-egu2020-22358, 2020.