Publications

The recently developed 3-source-partitioning approach: addition of 14C labeled organics to soil after C3–C4 vegetation changes, was used to distinguish C sources in three compartments, namely CO2, microbial biomass and dissolved organic C (DOC) during decomposition of labeled cellulose. Microbial community structure (based on PLFA composition) and functions (based on enzyme activities and on microbial growth parameters) revealed mechanisms and drivers of priming effects (PE) induced by cellulose addition. 14C-cellulose input caused negative PE within the first week and was accompanied by fast consumption of unlabelled DOC and its incorporation into microbial biomass. Microbial activation however, was not confirmed by substrate-induced respiration, nor by hydrolytic enzymes activity or by PLFA changes. A remarkable exception was a 2-fold increase in protozoan PLFA. Such an increase indicates that microorganisms feeding on cellulose and on DOC were quickly grazed by protozoans acting as a driver of microbial succession. This experimentally demonstrates the functioning of the microbial interactions: protozoan grazers provided for rapid recycling of nutrients and facilitated the succession of cellulose-degrading microorganisms during the second week of cellulose decomposition. An increase in the activity of cellulolytic enzymes caused short-term real PE accompanied by increase in abundance of slow-growing fungi and G(−) bacteria. Long-term real PE observed between 14 and 60 days after cellulose input was due to decomposition of SOM-originated hemicelluloses by fungi and G(+) bacteria. The CO2 released by primed soil organic matter (SOM) decomposition was originated mainly from C younger than 12 years (63%) and only 37% were older than 12 years despite the recent and old C contributed almost equally (51 and 49%, respectively) to SOM under Miscanthus giganteus. This indicates that the SOM pools are involved in PE according to their availability. Despite 71% of the applied cellulose-C was sequestered in the soil, the net soil C-gain amounted only for 28% of the applied cellulose-C after factoring in the C losses by the PE. Our study emphasizes the role of food webs in the PE dynamics: cellulose input served as a driver activating the food chain through the microbial loop.

ABSTRACT Soil viruses control the dynamics and metabolism of their hosts, strongly modifying carbon and nutrient cycling as well as soil biochemistry. Warming specifically affects viruses and their hosts, but the consequences of climate warming on the virus–host interactions, and for soil functions, remain unknown. We investigated the viral communities and the virus–host interactions under warming in situ based on a forest soil column translocation experiment. The abundance of the Petitvirales (DNA viruses) decreased by 25%, but that of the Durnavirales and Martellivirales (RNA viruses) strongly increased. The DNA viral lysogenic signals and RNA viral lytic proteins increased in soil, indicating the opposite lifestyles of DNA and RNA viruses. Correspondingly, the DNA abundance of viral hosts increased, whereas RNA viral hosts remained stable. The high DNA viruses/host ratios reflect very intensive interactions between the virus and host, leading to the drop in the host functions (such as carbon metabolism processes and nitrogen and phosphorus cycles) up to 43%. In contrast, the functions of the hosts for RNA viruses increased by up to 48%. The fundamental difference in behaviour of DNA and RNA viruses is that the former use mainly lysogenic, whereas the latter lytic, lifestyles and thus control the responses of host communities to warming. Conclusively, the opposite response of DNA and RNA viruses to warming in abundance, lifestyle, and interactions with hosts leads to divergent changes in nutrient fluxes in soil. These new perspectives on viral regulations of microbial communities and their function under soil warming reveal the undeniable role of viruses in microbial ecology.

Soil zymography is a new technique developed to visualize two‐dimensional distributions of enzyme activities. The method consists of incubating a membrane saturated with an enzyme‐specific fluorogenic substrate on a surface of the soil sample, followed by recording the membrane image generated by a fluorescent product (e.g. MUF: methylumbelliferone) in ultraviolet light. Despite its relative ease of use, performing zymography involves multiple user‐made decisions that might affect the accuracy of enzyme activity estimates. Therefore, unification of the zymography methodology is required for correct estimations and comparisons of various studies. We evaluated the following methodological aspects of the implementation of zymography: (a) camera settings and image processing, (b) effects of evaporation and (c) calibration procedures. Camera settings (shutter speeds or exposure time) affected the intensity of background fluorescence and signal‐to‐noise ratios (SNR). However, because their combined effects varied depending on MUF concentrations, light and camera setting need to be optimized for the expected range of MUF concentrations prior to zymography. Evaporation of MUF solution from the membrane had no effect on fluorescence. Relations between MUF concentration and intensity of fluorescence during calibrations demonstrated a saturated pattern and were strongly affected by image noise outside the optimal range (e.g. 8–14 μ m MUF pixel −1 ). We developed a new calibration approach that is based on a piecewise linear regression. The new approach accounted for specific ranges of MUF concentration and uses nonuniformly saturated membranes, reflecting the real distribution of enzyme activities in soil. The new calibration algorithm eliminated biases of the standard calibration and resulted in greater accuracy in predicting MUF concentrations. Highlights We developed a new approach to calibration for 2‐D soil zymography. The approach accounted for spatial nonuniformity of soil zymograms. Standard calibration resulted in systematic underestimation of enzyme activity. Soil zymography requires pixel‐based calibration with nonuniformly saturated membranes.

Nitrification acidifies soil, and the produced H+ are neutralized by inorganic carbon (C) in soil leading to irreversible CO2 emissions. CO2 released by nitrogen (N) fertilizer-induced acidification is partitioned between solid (CaCO3 re-precipitation), liquid (dissolved HCO3 − and CO3 2−) and gaseous (CO2) phases. Therefore, quantifying the effects of N fertilization on CO2 emissions from soil inorganic C is an enormous challenge. 14C-labeled CaCO3 was used as a model inorganic C to trace the released CO2 caused by acidification by five fertilizers: chicken manure, urea, KNO3, NH4NO3, and (NH4)2SO4 added at three N rates. Cropland soil was homogenously mixed with Ca14CO3 powder and fertilizers, and the emitted CO2 was trapped in NaOH solution to determine total CO2 and 14CO2 efflux originated from inorganic C. Fertilization, particularly ammonium-based fertilizers ((NH4)2SO4, NH4NO3), strongly decreased soil pH by 0.35 units over 40 days. All fertilizers except KNO3 increased total CO2 emissions by 21%–490% compared to the unfertilized control soil. The fertilization effects on cumulative 14CO2 emission induced by CaCO3 neutralization, corresponded to acidification and decreased in the order (NH4)2SO4 > NH4NO3 > urea > KNO3 > chicken manure. Ammonium-based fertilizers induced the strongest CO2 emissions originated from inorganic C, emitting 1.6–4.5 times more 14CO2 than non-fertilized soils during the first nine days. The total CO2 emissions from SIC were proportional to the fertilizer dose applied. Therefore, we conclude that both the choice of N fertilizers and their application rates need to be considered to control CO2 emissions originated from inorganic C. The soil inorganic C losses should be prevented not only because of their irreversible contribution to atmospheric CO2, but also to safeguard ecosystem services of CaCO3, such as organic matter preservation, soil structure stabilization, and C sequestration.

Silicon (Si) and arbuscular mycorrhizal fungi (AMF) increase plant resistance to various environmental stresses, including heavy metal (and metalloid) toxicity. Although Si and AMF each independently enhance plant tolerance, the nature of their interactions and their combined impacts on nutrient uptake, especially in the context of toxic elements such as arsenic (As), remains to be elucidated. This study investigated AMF-mediated regulation of plant nutrient uptake under As stress using rice, a model Si-accumulating plant. Experiments were conducted under As-free and As stress conditions, incorporating AMF inoculation and silicic acid application, with a focus on nutrient uptake and transporter expression. Without As, AMF inoculation increased shoot Si content by 44%, while invariance was common under As toxicity stress (10 μM of As(III)). Despite As presence, AMF increased Lsi1 expression with Si application, elevating As content in roots and shoots by 38% and 55%, respectively. Introduction of As stress amplified AMF role in phosphorus (P) uptake from 13% to 38%, correlating with up-regulated P transporter expression. Three-way ANOVA of interactions among As, Si, and AMF on P and As uptake by rice revealed that As amplified AMF potential to increase P uptake while weakening promotive effect on Si uptake. Silicon reduced As absorption, while AMF increased As uptake, but the elevated As were potentially retained within fungal hyphae, limiting transfer to rice plants. Overall, As toxicity stress had contrasting effects on P- and Si-promoting roles of AMF. These findings contribute to our understanding of plant-fungal interactions under heavy metal stress.

Soil organic matter (SOM) and its fractions play key roles in optimizing crop yield and improving soil quality. However, how functional SOM fractions responded to long-term fertilization and their relative importance for C sequestration were less addressed. In this study, we determined the effects of long-term fertilization on six functional SOM fractions (unprotected, physically protected, physico-biochemically protected, physico-chemically protected, chemically protected, and biochemically protected) based on two long-term fertilization experiments carried out in South China. The unprotected coarse particulate organic matter (cPOM), the biochemically and chemically protected silt-sized fractions (NH-dSilt and H-dSilt) were the primary C storage fractions under long-term fertilization, accounting for 23.6–46.2%, 15.7–19.4%, and 14.4–17.4% of the total soil organic carbon (SOC) content in upland soil and 19.5–29.3%, 9.9–15.5%, and 14.2–17.2% of the total SOC content in paddy soil, respectively. Compared with the control treatment (CK) in upland soil, the application of manure combined with mineral NPK (NPKM) resulted in an increase in the SOC content in the cPOM, pure physically protected fraction (iPOM), the physico-chemically protected (H-μSilt), and the chemically protected (H-dSilt) fraction by 233%, 166%, 124%, and 58%, respectively. Besides, the SOC increase in upland soil expressed as SOC content per unit of total SOC for iPOM, H-μSilt, cPOM and H-dSilt were the highest and as large as 283%, 248%, 194%, and 105% respectively. In paddy soil, the highest increase per unit of total SOC was H-dSilt (190%), followed by H-dClay (156%) and H-μSilt (155%). These results suggested that the upland soil could stabilize more C through the pure physical, whereas the chemical protection mechanism played a more important role in paddy soil. Chemical protection mechanism within the microaggregates played important roles in sequestrating C in both upland and paddy soils. Overall, the different responses of functional SOM fractions to long-term fertilization indicate different mechanisms for SOM cycling in terms of C sequestration under upland and paddy systems.

Soil acidification is an ongoing problem in intensively cultivated croplands due to inefficient nitrogen (N) fertilization. We collected high-resolution data comprising 19,969 topsoil (0–20 cm) samples from the Land Use and Coverage Area frame Survey (LUCAS) of the European commission in 2009 to calculate the impact of N fertilization on buffering substances such as carbonates and base cations. We have only considered the impacts of mineral fertilizers from the total added N, and a N use efficiency of 60 %. Nitrogen fertilization adds annually 6.1 × 107 kmol H+ to European croplands, leading to annual loss of 6.1 × 109 kg CaCO3. Assuming similar acidification during the last 50 years, carbonates were completely removed from 3.4 × 106 ha of European croplands. In carbonate-free soils, annual loss of 2.1 × 107 kmol of basic cations will lead to strong acidification of at least 2.6 million ha of European croplands within the next 50 years. Inorganic carbon and basic cation losses at such rapid scale tremendously drop the nutrient status and production potential of croplands. Soil liming to ameliorate acidity increases pH only temporarily and with additional financial and environmental costs. Only the direct loss of soil carbonate stocks and compensation of carbonates-related CO2 correspond to about 1.5 % of the proposed budget of the European commission for 2023. Thus, controlling and decreasing soil acidification is crucial to avoid degradation of agricultural soils, which can be done by adopting best management practices and increasing nutrient use efficiency. Regular screening or monitoring of carbonate and base cations contents, especially for soils, where the carbonate stocks are at critical levels are urgently necessary.

<p>Thermokarst lakes formed form permafrost thawing under the global warming are an important source of greenhouse gases (GHG). However, the driving mechanisms and temperature sensitivity (Q<sub>10</sub>) of GHG emissions from the sediments of thermokarst lakes require deeper understanding. From existing studies of organic matter (OM) turnover and thermodynamic theory, it is known that more refractory OM has a higher temperature sensitivity of decomposition. To test the relevance of such effects in thermokarst lakes, sediments of two differently sized lakes (small = young, DOC rich; large = mature, DOC poor) from Western Siberia were anoxically incubated under three temperatures (4, 10, 16°C) for 49 days. We hypothesized that the Q<sub>10</sub> of CO<sub>2</sub>, CH<sub>4</sub> and N<sub>2</sub>O production increases with lake size as OM becomes increasingly refractory. Rates of CO<sub>2</sub> production increased exponentially with temperature in sediments from lakes of both sizes, whereas the highest rates were observed for sediments of the small lake (4.2-9.7 μg C g<sup>-1</sup> day<sup>-1</sup>), as expected for the more labile OM. However, the Q<sub>10</sub> of CO<sub>2</sub> production (1.8-2.2) was unexpectedly similar between two lakes. The small lake sediment emitted 2-3 orders of magnitude larger amount of CH<sub>4</sub> (20-583 ng C g<sup>-1</sup> day<sup>-1</sup>) as compared with large lake. The Q<sub>10</sub> values and activation energy (E<sub>a</sub>) of CH<sub>4</sub> production in small lake sediment significantly decreased from 4-10°C (Q<sub>10</sub> = 6.7; E<sub>a</sub> = 124 kJ mol<sup>-1</sup>) to 10-16°C (Q<sub>10</sub> = 3.1; E<sub>a</sub> = 76 kJ mol<sup>-1</sup>). This suggests that methanogenesis is a strongly temperature-dependent process that is more sensitive in the low-temperature range. However, Q<sub>10</sub> of CH<sub>4</sub> production in the large lake did not reveal a sensitivity to temperature probably due to too low CH<sub>4</sub> concentrations. In contrast to low CH<sub>4</sub> production, the N<sub>2</sub>O emission rates were dramatically high (0.1-1.3 μg N g<sup>-1</sup> day<sup>-1</sup>) in the sediment of the large lake. Interestingly, there was no N<sub>2</sub>O detected in the small lake sediment. Presumably, intensive denitrification in the large lake sediment outcompeted methanogenesis for substrate and energy, or enhanced CH<sub>4</sub> oxidation occurred with NO<sub>3</sub><sup>-</sup> as the electron acceptor. In summary, the temperature sensitivity of GHG production in thermokarst lake sediments depended more on gas species than on lake size. Nevertheless, the size of thermokarst lakes can serve as an indicator of biogeochemical processes in the sediments, as the small lakes are hotspots of CH<sub>4</sub> and the large lakes are hotspots of N<sub>2</sub>O production.</p>

Direct plant uptake of organic nitrogen (N) is often studied using the dual-labeling approach (15N + 13C or 15N + 14C). However, the method might be hampered by uptake of labeled inorganic carbon (C) produced by mineralization of labeled organic compounds. Here we report the results from a triple labeling experiment (15N + 13C + 14C) investigating whether root uptake of labeled inorganic C can bias the results obtained in studies of organic N uptake using dual-labeled amino acids (15N, 13C). In a rhizosphere tube experiment we investigated 13C and 14C uptake by maize either supplied with labeled glycine or CO 3 2 − , but found no differences in uptake rates between these C-sources. The uptake of inorganic C to the shoot tissue was higher for maize grown in full light compared to shading, which indicates a passive uptake of inorganic C with water. We conclude that uptake of inorganic C produced by mineralization of amino acids can significantly bias the interpretations of organic N uptake studies using dual-labeling.

Global warming is accelerating glacial retreat and leaving open areas for vegetation succession on young developing soils. Soil microbial communities interact with plants affecting vegetation succession, but the specific microbial groups controlling these interactions are unclear. We tested whether plant–soil–microbial interactions explain plant primary succession in the Gongga Mountain glacial retreat chronosequence. The direction and intensity of plant–soil–microbial interactions were quantified by comparing the biomass of one early-, two mid- and two late-succession plant species under sterilized vs. live, and inter- vs. intra-specific competition. The performance of most plant species was negatively affected by soil biota from early habitats (5–10 yr), but positively by soil biota from mid- (30–40) and late-succession (80–100) habitats. Two species of Salicaceae from middle habitats, which are strong competitors, developed well on the soils of all successional stages and limited the establishment of later serial plant species. The strongest microbial drivers of plant–microbial interactions changed from i) saprophytic fungal specialists during the early stage, to ii) generalists bacteria and arbuscular mycorrhizal fungi in the middle stage, and finally to iii) ectomycorrhizal fungal specialists in the late stage. Microbial turnover intensified plant–soil–microbial interactions and accelerated primary succession in the young soils of the glacial retreat area.