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One of the most prominent changes in the rhizospheric soil structure is associated with the formation of a strongly bound soil layer in the surroundings of the root, which is named rhizosheath. In this study, we investigated how root hair elongation, a ubiquitous root morphological trait, affect the stability of rhizosheath aggregates. Using 13CO2 pulse labeling, we tracked the fate of root-derived 13C inputted into the rhizosheath of two Zea mays L. genotypes with contrasting root hair elongation: a mutant with root hair defective elongation (rth3) and a corresponding wild type (WT). In addition, we also investigated the differences between two 13CO2 labeling approaches (single vs. multiple pulse labeling) in the distribution of 13C in the rhizosheath aggregates. We were able to demonstrate that the rhizosheath aggregate stability and the resulting aggregate size distribution follows the same mechanisms irrespective of the root hair elongation. This result reinforces the assumption that other soil properties are more decisive for the soil structure formation in the rhizosheath in comparison to root hair elongation. The majority of recently deposited root-derived C (57%) was found in the macroaggregates. Increasing the number of pulses (multiple pulse labeling approach) resulted in a higher 13C enrichment of the rhizosheath aggregates fractions in comparison to the application of a single pulse. While both labeling approaches resulted in a similar distribution of 13C in the rhizosheath aggregates, the higher enrichment given by multiple pulse labeling allowed the separation of significant differences between the genotypes in plant C allocation in the rhizosheath.

Agroforestry systems are characterized by a high complexity between vegetation components and niche partitioning. In a crop-grass-tree agroforestry system, rape, willow, and grasses were in situ pulse labeled separately with 13CO2 for 6 h, and 13C was traced in shoots, roots, topsoil (0–15 cm) and subsoil (15–30 cm), microbial biomass carbon (C), and dissolved organic C, as well as respiration losses (CO2) up to 28 days after labeling to investigate the effects of vegetation components on C allocation belowground. 13C recovery in roots after 28 days was 7.0% of total assimilated C for grassland, which was 3.5- and 5.2-fold higher than that for rape and willow, respectively. The larger C allocation belowground in grassland was ascribed to its higher root/shoot ratio compared to willow and rape. Grassland facilitated higher accumulation of root-derived C in soil compared to rape (9.2% of recovered 13C) and compared to willow (1.6% of 13C). Willow retained more photosynthetic C aboveground and less was allocated to roots compared to rape. Although the C allocated to the top 15-cm soil was similar between willow and rape, willow facilitated C allocation in deeper soil compared to rape (0.6% vs. 0.2%). This could be explained by the lower microbial activity and subsequent weaker decomposition of rhizodeposits in 15–30-cm depth under willow. The net belowground C inputs in grassland, willow, and rape were 0.53, 0.06, and 0.10 g C m−2 month−1 of vegetation period, including rhizodeposition of 0.24, 0.05, and 0.04 g C m−2 month−1, respectively. Overall, integrating trees and grassland within cropland facilitates higher root-derived C input into soil, thus contributing to the soil C sequestration in agroforestry systems.

The spatial arrangement of the soil surrounding the root can improve plant resource acquisition under drought and is closely related to the fate of soil organic carbon (SOC). Thus, the formation of soil structure and the establishment of a stable rhizosheath can potentially improve plant drought resistance and contribute to maintained crop yields during drought events. Yet, soil structure formation is a complex process determined by the interaction between various functional plant and soil properties, such as the soil (micro)biome, root exudation, or root morphological characteristics. To date, it is not understood how water scarcity affects soil aggregation in the vicinity of roots, by which functional traits these drought effects can be modified, and how this feedbacks on the cycling of SOC. Thus, we investigated drought effects on rhizosheath properties and their link with functional plant traits. We conducted a greenhouse experiment with 38 maize varieties where half of the plants were grown under optimum moisture, while the second half of replicates were subjected to drought stress after an initial establishment phase. For each plant, the rhizosheath soil was sampled and its aggregate size distribution, carbon (C) and nitrogen (N) content, and the proportion of newly maize-derived C were analysed via natural abundance 13C. In addition, we recorded functional plant and rhizosphere traits, such as morphological and chemical root properties, microbial enzyme activities, and plant biomass.Drought-stressed plants formed lower amounts of rhizosheath, with a decreased physical aggregate stability and increased concentrations of SOC, N, and newly maize-derived C. Furthermore, under drought larger proportions of the elements were allocated into the microaggregate fractions. In particular, maize-derived C, along with N, accumulated under drought stress in the smaller aggregate size classes of the rhizosheath. Maize varieties forming larger amounts of roots under drought stress tended to maintain higher macroaggregate stability in the rhizosheath. In contrast, cultivars that invested little in root growth but promoted higher microbial enzyme activities in the rhizosheath and maintained root N contents under drought were associated with a strong accumulation of maize-C and N in the smaller aggregate size classes. Trait-based experimental approaches, such as the one presented here, are deepening our mechanistic understanding of drought effects in the crop rhizosheath and can thus help to guide future crop selection for improved drought resistance.

Intensified silage maize (Zea mays L.) cultivation has led to several environmental problems such as high greenhouse gas emissions and groundwater pollution, thus further amplifying climate change. Therefore, alternative crops with high carbon (C) sequestration capacities are crucial for sustainable bioenergy production, particularly those that can cope with extreme climate events, such as drought stress. Therefore, this study aimed to evaluate the potential of the perennial cup-plant (Silphium perfoliatum L.) as a promising alternative for silage maize to increase soil C input and reduce C output (as CO2) under optimum watering and drought conditions. For this purpose, a lysimeter experiment comparing these crops subjected to two watering regimes (well-watered and drought-stressed), was set up at the botanical garden of the University of Bayreuth, Germany. Soil respiration was correlated to soil moisture and temperature, biomass (aboveground and belowground), soil C and nitrogen (N), and dissolved organic C (DOC) and inorganic C in the leachates. Soil CO2 efflux was measured using a Licor-6400XT gas exchange system (LI-COR, Lincoln, NE, USA)) fitted with a soil chamber over three consecutive years. The final cumulative CO2 efflux was greater by 29% under drought-stressed cup-plant than in drought maize, which was explained by the greater belowground biomass production under cup-plant. Cup-plant increased root biomass by 143% (0.86±0.26 vs 2.08±0.33 kg m-2) compared to silage maize, thus, implying a higher contribution of root respiration to the total soil respiration under cup-plant. Despite the soil respiration and DOC leaching being higher for cup-plant than maize, cup-plant increased soil C (by 4%) and N content (by 14%), after only two years of cultivation. Even though root respiration and enhanced microbial activity result in higher soil respiration and C mineralization, the continuous supply of fresh C via the root litter and rhizodeposits of the perennial cup-plant suggests long-term effective C farming and demonstrates the potential of the cup-plant as an ecological alternative to silage maize.Keywords: Climate change; soil respiration; carbon sequestration; bioenergy crops; leachates

Modeling and quantification of carbon (C) allocation through the soil-plant-atmosphere continuum (SPAC) has received increasing attention in recent years. Although advanced imaging and numerical methods have boosted our knowledge, we still lack an experimental and mechanistic understanding of C flux and its partitioning across the SPAC. Here we combined a 13C pulse labeling technique with modeling of C transport across the SPAC to describe the flow of newly assimilated C from shoots to roots, to soils and then respired back to the atmosphere. To do so, different plants - maize (Zea mays L.), soybean (Glycine max (L.) merr. cv. sohae), and wheat (Triticum aestivum L.) - were exposed to a 13CO2 pulse for 2 hoursduring daytime. A CO2 Isotope Analyzer (CCIA-38d-EP, Los Gatos Research) was used to continuously monitor 13CO2 flux from the soils. Subsequently, after harvest the respective 13C contents of shoot and root biomass and of the soil was quantified.To model the C fluxes, we developed a simple multi-compartment domain, representing the SPAC. The SPAC was envisioned as four interconnected horizontal compartments namely atmosphere, shoots, roots, and soil compartment. The shoots and roots compartment were further simplified in three vertical compartments (phloem, storage, and structural pool). The C transport between these pools was represented by constant rates. These rates were inversely estimated by adjusting the model parameters to best reproduce the measured C flux and C contents in the various compartments. Our model described the allocation and transport of 13C within the shoots, roots, and soil well. The best-fitted coefficients of the model were reproducible among different replications of the same plant species. We also checked the sensitivity of our model to its parameters and observed a good sensitivity to most of the model parameters. In particular, our model was very sensitive to C loading and unloading in the phloem and also root exudation rates. The results of our study show that the combination of tracing 13C and modeling of 13C transport across SPAC is a promising tool to study C flux and its partitioning across SPAC.

Plants' roots promote changes in soil structure, forming a strongly-bound soil layer in the surroundings of the root, which is named as rhizosheath. Rhizosheath formation is attributed mainly to the root hairs' presence, that favors the enmeshment of the soil particles around the roots, and the release of mucilage and exudates, which acts as gluing agents of those soil particles. In the present work, we studied the rhizosheath aggregate formation of two Zea mays L. genotypes with contrasting root hair development: a mutant with root hair defective elongation (rth3) and a corresponding wild type (WT). We also tracked the fate of recently-deposited C in the rhizosheath aggregates using two 13CO2 pulse labeling approaches (single vs. multiple pulse labeling). The sampled rhizosheath aggregates were further separated using dry-sieving fractionation into three aggregate size classes: primary small particles and smaller microaggregates (250 µm). We observed that the aggregate size distribution followed the same pattern in both genotypes. This result reinforces the assumption that other soil properties are more important for rhizosheath aggregation than root hair elongation. We observed that the higher potion of the recently-deposited root-derived C (57%) was accumulated in the macroaggregates. Moreover, the multiple pulse labeling approach proportioned a higher 13C enrichment of the rhizosheath aggregates fractions than applying a single pulse. Despite both single and multiple labeling approaches have resulted in a similar distribution of 13C in the rhizosheath aggregates, multiple pulse labeling provided a higher enrichment in the rhizosheath aggregates, which allowed a better separation of significant differences between the genotypes.

Denitrification in agricultural crop production is one of the main sources of gaseous N2O and N2 losses to the environment. To successfully develop mitigation strategies, it is crucial to understand N2O production pathways, but also to quantify all other gaseous N losses, especially N2 emissions. Therefore, this project aimed (1) to identify the main drivers of denitrification during plant growth and in the post-harvest period, (2) to quantify denitrification derived N2O and N2 losses during the cropping season, and (3) to assess the interactions between plant litter quality, initial litter degradation, and formation of hotspots of N2O and N2 production. We conducted experiments on the laboratory, greenhouse, and field plot scale using the 15N gas flux method and HeO2 atmosphere to directly measure N2O and N2 losses and to determine the fraction of denitrification derived N losses. We worked with soils, crops, temperature and moisture conditions that are typical for our central German humid-temperate climate.Plant growth affected all controlling factors of denitrification, especially soil moisture, NO3− and Corg availability. Crop species differed in their growing patterns and N uptake throughout the growing season controlling both N and C availability in soil. Accordingly, N2O and N2 emission patterns differed between crop species. Overall, emissions were highest when plant N uptake was low, i.e., during early growth stages and ripening, and after harvest. On the field scale, soil moisture and temperature were major controls of N2O+N2 losses.In a climate chamber study under controlled temperature conditions, N2O and N2 fluxes mainly derived from denitrification of labeled 15NO3− in anoxic microsites, while nitrification simultaneously occurred in more oxic parts of the soil, potentially contributing to formation of unlabeled N2O. Increasing soil moisture with irrigation increased denitrification rates in anoxic hotspots, which corresponded with increasing N2O and especially N2 fluxes. At the same time, it restricted nitrification and thus decreased the share of nitrification-dependent processes contributing to N2O formation.Incorporation of plant litter increased CO2, N2O, and N2 losses irrespective of litter quality, soil moisture or soil type/SOM content. We found that under O2 limiting conditions (70 % WFPS), the fraction of easily degradable C controlled the magnitude of N2O and N2 losses after litter incorporation. Under moderate soil moisture (50-60 % WFPS), interactions between litter degradation and SOM turnover affected the time course and processes contributing to N2O formation. Overall, our high-resolution gas flux measurements showed that N2O+N2 emissions from harvest residues can contribute significantly to the total N loss.

Aims Soil priming affects soil N transformation and plant N availability, but few studies have investigated these interactions to date. Methods To address this, we reviewed the literature for studies quantifying soil priming, soil N transformation and plant N uptake. Results Gross N mineralization was strongly controlled by soil priming in studies with plants, while abiotic factors had a minor influence on gross N mineralization. In contrast, soil priming was negatively related to gross N mineralization and had a low explanatory power in incubation studies where substrates are added as surrogates for root exudates. These results indicate that plants support increased N mineralization and that this is not adequately reflected in incubation studies. Additionally, we observed a positive relationship between soil priming and the % of Norg-derived N uptake as well as total N uptake, which demonstrates that priming enhances the availability of N that was previously organically bound and that at least part of the N mineralized during priming was available for plant uptake. Conclusion Our results show that the effect of roots and rhizodeposition leads to a number of processes supporting N mineralization and availability through priming that are not well reflected in incubation studies. To fully capture the interactions between plant roots and their associated microbiota, we recommend focusing research on systems with plants. Additionally, the strong correlation between C and N transformation should be considered in biogeochemical modelling.

Deadwood is a key component of forest ecosystems, but there is limited information on how it influences forest soils. Moreover, studies on the effect of thinning-derived deadwood logs on forest soil properties are lacking. This study aimed to investigate the impact of thinning-derived deadwood logs on the soil chemical and microbial properties of a managed spruce forest on a loamy sand Podzol in Bavaria, Germany, after about 15 years. Deadwood increased the soil organic carbon contents by 59% and 56% at 0–4 cm and 8–12 cm depths, respectively. Under deadwood, the soil dissolved organic carbon and carbon to nitrogen ratio increased by 66% and 15% at 0–4 cm depth and by 55% and 28% at 8–12 cm depth, respectively. Deadwood also induced 71% and 92% higher microbial biomass carbon, 106% and 125% higher microbial biomass nitrogen, and 136% and 44% higher β-glucosidase activity in the soil at 0–4 cm and 8–12 cm depths, respectively. Many of the measured variables significantly correlated with soil organic carbon suggesting that deadwood modified the soil biochemical processes by altering soil carbon storage. Our results indicate the potential of thinned spruce deadwood logs to sequester carbon and improve the fertility of Podzol soils. This could be associated with the slow decay rate of spruce deadwood logs and low biological activity of Podzols that promote the accumulation of soil carbon. We propose that leaving thinning-derived deadwood on the forest floor can support soil and forest sustainability as well as carbon sequestration.

Background and Aims Stomatal regulation allows plants to promptly respond to water stress. However, our understanding of the impact of above and belowground hydraulic traits on stomatal regulation remains incomplete. The objective of this study was to investigate how key plant hydraulic traits impact transpiration of maize during soil drying. We hypothesize that the stomatal response to soil drying is related to a loss in soil hydraulic conductivity at the root–soil interface, which in turn depends on plant hydraulic traits. Methods We investigate the response of 48 contrasting maize (Zea mays) genotypes to soil drying, utilizing a novel phenotyping facility. In this context, we measure the relationship between leaf water potential, soil water potential, soil water content and transpiration, as well as root, rhizosphere and aboveground plant traits. Key Results Genotypes differed in their responsiveness to soil drying. The critical soil water potential at which plants started decreasing transpiration was related to a combination of above and belowground traits: genotypes with a higher maximum transpiration and plant hydraulic conductance as well as a smaller root and rhizosphere system closed stomata at less negative soil water potentials. Conclusions Our results demonstrate the importance of belowground hydraulics for stomatal regulation and hence drought responsiveness during soil drying. Furthermore, this finding supports the hypothesis that stomata start to close when soil hydraulic conductivity drops at the root–soil interface.

Background and aims Plant growth affects soil moisture, mineral N and organic C availability in soil, all of which influence denitrification. With increasing plant growth, root exudation may stimulate denitrification, while N uptake restricts nitrate availability. Methods We conducted a double labeling pot experiment with either maize (Zea mays L.) or cup plant (Silphium perfoliatum L.) of the same age but differing in size of their shoot and root systems. The 15N gas flux method was applied to directly quantify N2O and N2 fluxes in situ. To link denitrification with available C in the rhizosphere, 13CO2 pulse labeling was used to trace C translocation from shoots to roots and its release by roots into the soil. Results Plant water and N uptake were the main factors controlling daily N2O + N2 fluxes, cumulative N emissions, and N2O production pathways. Accordingly, pool-derived N2O + N2 emissions were 30–40 times higher in the treatment with highest soil NO3− content and highest soil moisture. CO2 efflux from soil was positively correlated with root dry matter, but we could not detect any relationship between root-derived C and N2O + N2 emissions. Conclusions Root-derived C may stimulate denitrification under small plants, while N and water uptake become the controlling factors with increasing plant and root growth.

There is a trend of increasing woody biomass in tropical savannas. Here we ask what effect this increase may have on soil carbon pools and fluxes. Using a field experiment we determine the amount of soil carbon directly under grasses, a juvenile tree among grasses and a juvenile tree with no grasses. We also measure CO2 efflux at the soil surface and use gas wells to extract CO2 from several soil depths. Our results show that grasses contribute substantially more than trees to both soil carbon and soil respiration. Grasses also make a disproportionate contribution to the δ13C value of SOC in the tree only treatments. The surface gas efflux data show that soil respiration increased with an increase in volumetric soil moisture and temperature and plots with both grasses and trees had higher respiration rates than plots with trees only or with grasses only. The highest soil respiration is in the top 20 cm of the soil with grasses the primary contributors to both δ13CSOC and δ13CR. Any increase in woody biomass will result in a decline in SOM turnover and nitrogen mineralization rates resulting in higher SOC pools. The associated increases in SOC and above ground biomass will however be associated with negative economic and biodiversity impacts.

As the dynamics and magnitude of rhizodeposition vary considerably among cropping systems, we investigated effects of cropping system on the incorporation of root-derived carbon (C) into Collembola, a dominant taxon of soil microarthropods. In the field, we used 13CO2 to pulse label a crop monoculture (oilseed rape, Brassica napus L.), a mixed-grass community (dominated by Lolium perenne L. mixed with clover Trifolium repens L.), and a tree plantation (willow, Salix schwerinii E.L. Wolf and Salix viminalis L.). During 28 days, the incorporation of 13C was traced in nine species of Collembola including epedaphic (surface-dwelling), hemiedaphic (litter-dwelling), and euedaphic (soil-dwelling) functional groups. Incorporation of 13C into Collembola reached a plateau before day 3 after the labeling in grass and willow, but increased up to day 14 in rape. While euedaphic Collembola incorporated less root-derived C than epedaphic and hemiedaphic Collembola in rape and willow, the incorporation of 13C was similar among functional groups in grass. Differential incorporation of 13C in euedaphic species points to niche differentiation within the same functional group. Our findings highlight that cropping system not only affects the flux of root C into soil mesofauna, being slower in rape than in grass and willow, but also the utilization of root-derived resources by functional groups and species of Collembola. The results indicate that pronounced differences in belowground C inputs between cropping systems affect microbivores as basal species and thereby soil food webs and their functioning and services.

Limited water supply is one of the largest impediments to food production worldwide in the light of climate change and increasing food demand. Stomatal regulation allows plants to promptly react to water stress and regulate water use. Although the coordination between stomatal closure and aboveground hydraulics has extensively been studied, our understanding of the impact of belowground hydraulics on stomatal regulation remains, as yet, incomplete. The overall objective of this study was to investigate the impact of belowground hydraulic conductivity as affected by differences in expressions of root and rhizosphere traits on the water use regulation of different maize genotypes.We have utilized a novel phenotyping facility to investigate the response of a selection of 48 maize (Zea mays L.) genotypes exhibiting different root and rhizosphere traits to soil drying. We measured the relation between leaf water potential, soil water potential, soil water content and transpiration rate, as well as root and rhizosphere traits (e.g. root length, rhizosheath mass) between genotypes. Our hypothesis is that stomatal response to soil drying is related to a loss in soil hydraulic conductivity and that key root and rhizosphere hydraulic traits affect such relation.We found that the genotypes differed in their responsiveness to drought and that such differences were related to belowground hydraulic traits. The critical soil water content at which plants started to decrease transpiration was related to a combination of plant- and rhizosphere traits (namely plant hydraulic conductance, maximum transpiration rate, root length and rhizosheath mass). Genotypes with a higher maximum transpiration rate and a higher plant hydraulic conductance and a smaller root system closed stomata in wetter soil conditions, meaning earlier in the drying process. This finding is explained by a soil-plant hydraulic model that assumes that stomata start to close when the soil hydraulic conductance of the soil-plant continuum starts to decline. Those findings stress the importance of belowground hydraulic properties on stomatal regulation and thereby drought responsiveness.

Soil C priming is a short-term change in the turnover of soil organic matter caused by the addition of easily available organic C to the soil. The increase in SOM decomposition during priming is likely to affect not only C- but also gross N mineralization from SOM because large amounts of soil N is stored in SOM that is decomposed during priming. In order to assess whether soil C priming results in an increase in gross and mineralization and finally in enhanced plant N availability and uptake, we searched the literature for studies relating soil C priming to soil N cycling. In order to assess the effect of soil priming on soil N cycling we included studies quantifying soil C priming (PE) and gross N mineralization (GNM) in plant systems and in incubation setups. Secondly, we searched for studies measuring GNM in dependence to addition of C to the system. The third data set comprised studies, quantifying PE and the % of soil N derived N uptake as well as total N uptake of plants. Finally we included studies that quantified soil priming and enzyme activities in the respective soil samples. In order to be able to compare PE to GNM, % of soil N derived N uptake and soil enzyme activities respectively, we calculated the excess of GNM, % of soil N derived N uptake and soil enzyme activities by subtracting the parameter values of the control from the treatment values. We found a significant positive relation between soil C priming and GNM for studies with plants (R2=0.21) indicating that soil priming caused by root exudation increased soil N mineralization. In agreement with this, activities of enzymes related to the N cycle were positively related to priming (p=0.09), though, due to the small number of studies, the enzyme results must be interpreted with caution. In contrast to plant studies, the relation between soil C priming and GNM was significantly negative for incubation studies (R2=0.06). These contrasting results for plant and incubation studies indicate that incubation studies might not adequately reflect processes occurring in the rhizosphere. It is possible that plants attract particularly N mineralizing microbes for example by exudation of signaling compounds, a process that would not be reflected in incubation studies. We also found a significant positive relation between soil C priming and the % of soil N derived N uptake by plants (R2=0.56) and total plant N uptake (R²=0.21) indicating that at least part of the N mineralized during priming was available to, and taken up by the plants. In conclusion, the results of our meta-analysis indicate that rhizosphere C priming positively feeds back to plant N nutrition by causing increased N mineralization in the rhizosphere that facilitates plant N uptake.

The occurrence of drought is likely to increase and intensify as a result of climate change, which poses a great challenge to agriculture. It is thus crucial to enhance agronomic resilience to secure food and feed production. Roots and root functioning as well as the interplay of roots with the surrounding soil, the rhizosphere, plays a key role in water acquisition of plants. Investigating rhizosphere traits is hence promising to shed light on future crops that better adapt to drought stress. A great strength of this study is the screening of various varieties which is facilitated by the high-throughput phenotyping method. It allows a wider coverage of traits and especially the genetic and phenetic diversities preserved in landraces.Maize (Zea mays L.), being one of the major cereal crops worldwide, was selected as the plant of study. A total of 38 varieties, which encompasses hybrid varieties, open pollinated varieties, and landraces, were screened in the “Moving Fields”, a greenhouse equipped with the high-throughput phenotyping facility in the Bavarian State Research Center for Agriculture. Maize plants were grown in mesocosms filled with loamy soil. Plants were exposed to two water treatments, well-watered and drought-stressed, during vegetative stem extension stage. Dynamic plant development was captured by continuous image acquisition. A visible light (RGB) camera was used to document the size and architecture of shoots and roots, while a chlorophyll fluorescence camera recorded the metabolic activity of shoots.Using shoot images, we compared variety-specific plant growth curves under well-watered and drought-stressed conditions to highlight the growth strategy of plants towards drought stress. The results reveal differences in growth inhibition during drought across varieties. In addition, differences in shoot and root dry weights are found between landraces and modern varieties. More analyses are in progress in search of rhizosphere traits and their influences on agronomic resilience.

Alternate wet-drying (AWD) and sulfate fertilization have been considered as effective management practices for lowering CH4 emissions from paddy soils. However, the effects of management practices on in situ belowground CH4 turnover (production and oxidation) are not yet fully understood. Here, soil CO2 and CH4 concentrations and their C isotope compositions were measured at three rice growing stages in straw-amended paddy soils with and without sulfate fertilization under continuously flooded conditions and two wet-dry-cycles. CH4 concentration reached 51.0 mg C L-1 at flowering stage under flooded conditions, while it decreased to 0.04 mg C L-1 under AWD. Relative enrichment of δ13C in CH4 and depletion of δ13C in CO2 under AWD indicated CH4 oxidation. Sulfate addition had no significant effect on CH4 concentration. The ample substrate supply might have prevented sulfate-reducing bacteria from out-competing methanogenic archaea and could therefore explain the absence of a fall in CH4 production. The δ13C-CO2 enrichment over time (7 ‰ and 5‰ with and without sulfate fertilizer, respectively) under flooded conditions likely indicates an increasing contribution of hydrogenotrophic methanogenesis to CH4 production with ongoing rice growth. Overall, the results showed that AWD could more efficiently reduce CH4 production than sulfate fertilization in rice-straw-amended paddy soils.

Various studies showed a decrease of drought stress specific parameters of plants after silicon (Si) fertilization. But all studies differed in soil Si concentration between the control and Si treatments. As amorphous silica (ASi) was recently found to cause a strong increase of water holding capacity and plant available water in soils, a combined effect of soil moisture and plant response due to Si addition was assumed. In this study, the influence of the soil Si content was excluded by using the same Si enriched soil for treatments of two rice lines, lsi1 mutant defective in Si uptake and its wild-type rice. Most plant parameters, such as nutrient contents, biomass, specific leaf area, specific root length, leaf water content and C allocation did not differ significantly between the genotypes neither under flooded conditions, nor under drought conditions. Only photosynthesis and stomatal conductance were slightly higher for the wild type in both drought and flooded treatments. Overall, our data showed that Si accumulation within the plant tissues has only a minor effect on plant performance under drought stress. Hence, existing studies should be reinterpreted in light of the fact that Si additions may increase soil water availability.

Microbial turnover processes are typically restricted by low substrate availability in the subsoil. We hypothesized that SOM decomposition increases with plant density and decreases with N fertilization: We expected a greater rate of C allocation to the rhizosphere in the topsoil than in the subsoil treatments. In order to simulate different degrees of rhizodeposition, wheat was planted in pots at four different densities. The plants were continuously labeled with 13C-depleted CO2. Soil CO2 efflux was partitioned for root- and SOM-derived CO2. Moreover, we determined the enzyme kinetics by measuring catalytic efficiency and enzyme stoichiometry in both topsoil and subsoil. Shoot biomass and the shoot to root ratio were significantly higher for plants grown in the topsoil compared with the subsoil, which demonstrated higher relative C allocation to root biomass in the subsoil treatment. Despite the similar size of the rhizosphere, root-derived CO2 was always higher in the topsoil compared with the subsoil treatment, indicating enhanced root exudation. Effect sizes of all enzyme activities showed stronger magnitudes for the subsoil treatments. This was in line with a two-times increase of the effect size of SOM decomposition in the subsoil relative to topsoil. Overall, the plants in the subsoil treatments allocated more C to root biomass, less C to shoot biomass, and substantially less C to root exudates. However, the effect sizes of both SOM decomposition and enzyme activities were higher in the subsoil than in the topsoil, reflecting a stronger sensitivity to C inputs.

Numerous 13CO2 labeling studies have traced the flow of carbon from fresh plant exudates into rhizosphere bacterial communities. However, the succession of the uptake of carbon leaving the roots by distinct rhizosphere microbiota has rarely been resolved between microbial kingdoms. This can provide valuable insights on the niche partitioning of primary rhizodeposit consumption, as well as on community interactions in plant-derived carbon flows in soil. Here, we have traced the flow of fresh plant assimilates to rhizosphere microbiota of maize (Zea mays L.) by rRNA-stable isotope probing (SIP). Carbon flows involving bacteria, unicellular fungi, as well as protists were observed over 5 and 8 days. Surprisingly, labeling of Paraglomerales and several bacteria including Opitutus, Mucliaginibacter and Massilia spp. was especially apparent in soil surrounding the strict rhizosphere after 5 d. This highlights the central role of arbuscular mycorrhizal fungi (AMF) as a shunt for fresh plant assimilates to soil microbes not directly influenced by root exudation. Distinct trophic webs involving different flagellates, amoeba and ciliates were also observed in rhizosphere and surrounding soil, while labeling of filamentous saprotrophic Ascomycota or Basidiomycota was not apparent. This challenges the proposed “sapro-rhizosphere” concept and demonstrates the utility of rRNA-SIP to disentangle inter-kingdom microbial relationships in the rhizosphere.

Plant roots interact with rhizosphere microorganisms to accelerate soil organic matter (SOM) mineralization for nutrient acquisition. Root-mediated changes in SOM mineralization largely depend on root-derived carbon (root-C) input and soil nutrient status. Hence, intraspecific competition over plant development and spatiotemporal variability in the root-C input and nutrients uptake may modify SOM mineralization. To investigate the effect of intraspecific competition on SOM mineralization at three growth stages (heading, flowering, and ripening), we grew maize (C4 plant) under three planting densities on a C3 soil and determined in situ soil C- and N-mineralization by 13C-natural abundance and 15N-pool dilution approaches. From heading to ripening, soil C- and N-mineralization rates exhibit similar unimodal trends and were tightly coupled. The C-to-N-mineralization ratio (0.6 to 2.6) increased with N availability, indicating that an increase in N-mineralization with N depletion was driven by microorganisms mining N-rich SOM. With the intraspecific competition, plants increased specific root lengths as an efficient strategy to compete for resources. Root morphologic traits rather than root biomass per se were positively related to C- and N-mineralization. Overall, plant phenology and intraspecific competition controlled the intensity and mechanisms of soil C- and N- mineralization by the adaptation of root traits and nutrient mining.

The main prerequisites for denitrification are availability of nitrate (NO3-) and easily decomposable organic substances, and oxygen deficiency. Growing plants modify all these parameters and may thus play an important role in regulating denitrification. Previous studies investigating plant root effects on denitrification have found contradictive results. Both increased and decreased denitrification in the presence of plants have been reported and were associated with higher Corg or lower NO3- availability, respectively. Accordingly, it is still unclear whether growing plants stimulate denitrification through root exudation or restrict it through NO3- uptake. Furthermore, reliable measurements of N2 fluxes and N2O/(N2O+N2) ratios in the presence of plants are scarce.Therefore, we conducted a double labeling pot experiment with either maize (Zea mays L.) or cup plant (Silphium perfoliatum L.) of the same age but differing in size of their shoot and root systems. The 15N gas flux method was applied to directly quantify N2O and N2 fluxes in situ. To link denitrification with available C in the rhizosphere, 13CO2 pulse labeling was used to trace C translocation from shoots to roots and its release by roots into the soil.Plant water uptake was a main factor controlling soil moisture and, thus, daily N2O+N2 fluxes, cumulative N emissions, and N2O production pathways. However, N fluxes remained on a low level when NO3- availability was low due to rapid plant N uptake. Only when both N and water uptake were low, high NO3- availability and high soil moisture led to strongly increased denitrification-derived N losses.Total CO2 efflux was positively correlated with root dry matter, but there was no indication of any relationship between recovered 13C from root exudation and cumulative N emissions. We anticipate that higher Corg availability in pots with large root systems did not lead to higher denitrification rates, as NO3- was limiting denitrification due to plant N uptake. Overall, we conclude that root-derived C stimulates denitrification only when soil NO3- is not limited and low O2 concentrations enable denitrification. Thus, root-derived C may stimulate denitrification under small plants, while N and water uptake become the controlling factors with increasing plant and root growth.

Growing plants affect soil moisture, mineral N and organic C (Corg) availability in soil and may thus play an important role in regulating denitrification. The availability of the main substrates for denitrification (Corg and NO3-) is controlled by root activity and higher denitrification activity in rhizosphere soils has been reported. We hypothesized that (I) plant N uptake governs NO3- availability for denitrification leading to increased N2O and N2 emissions, when plant N uptake is low due to smaller root system or root senescence. (II) Denitrification is stimulated by higher Corg availability from root exudation or decaying roots increasing total gaseous N emissions while decreasing their N2O/(N2O+N2) ratios.We tested these assumptions in a double labeling pot experiment with maize (Zea mays L.) grown under three N fertilization levels S / M / L (no / moderate / high N fertilization) and with cup plant (Silphium perfoliatum L., moderate N fertilization). After 6 weeks, all plants were labeled with 0.1 g N kg-1 (Ca(15NO3)2, 60 at%), and the 15N tracer method was applied to estimate plant N uptake, N2O and N2 emissions. To link denitrification with available C in the rhizosphere, 13CO2 pulse labeling (5 g Na213CO3, 99 at%) was used to trace C translocation from shoots to roots and its release by roots into the soil. CO2 evolving from soil was trapped in NaOH for δ13C analyses, and gas samples were taken for analysis of N2O and N2 from the headspace above the soil surface every 12 h.Although pots were irrigated, changing soil moisture through differences in plant water uptake was the main factor controlling daily N2O+N2 fluxes, cumulative N emissions, and N2O production pathways. In addition, total N2O+N2 emissions were negatively correlated with plant N uptake and positively with soil N concentrations. Recently assimilated C released by roots (13C) was positively correlated with root dry matter, but we could not detect any relationship with cumulative N emissions. We anticipate that higher Corg availability in pots with large root systems did not lead to higher denitrification rates as NO3- was limited due to plant uptake. In conclusion, plant growth controlled water and NO3- uptake and, subsequently, formation of anaerobic hotspots for denitrification.

The projected global warming risks due to high emissions of greenhouse gases, mainly from anthropogenic activities, increases the need for an agricultural practice with high carbon sink capacity and low water requirements without compromising on environment and productivity. On one hand, it’s well accepted that soil moisture directly affects microbial activity, whereas on the other hand, drought stress was recently postulated to increase root exudates, which in turn will accelerate soil organic matter mineralization “priming effects”. Thus, the objective of this study was to investigate the interplay between soil moisture (well-watered and drought stressed) and maize (Zea mays L.) root exudates on soil CO2 efflux. The experiment consisted of three treatments, which are well-watered, drought stressed maize plus a control (without plants) lysimeters (1 m3), Soil CO2 efflux, soil temperature and moisture content were measured weekly during the growing season (April to September) and monthly in the fallow period. Under well-watered conditions, the annual average of CO2 efflux was 0.12 g CO2-C m-2 hr-1, which was 24.5 and 20% significantly higher than under drought stressed and the control, respectively. Moreover, well-watered treatment had significantly greater primed carbon than drought stressed maize. Soil temperature in deeper soil layers (25, 50 and 75 cm) correlated positively (with the CO2 efflux, while soil moisture correlated negatively at the 5 cm and 25 cm. Overall, these results suggested that the root exudates decreased under drought conditions, which decreasing soil respiration. Drought tolerance varieties could be an option to decrease soil respiration and maintain productivity.

Data for stable C and N isotope natural abundances of arbuscular mycorrhizal (AM) fungi are currently sparse, as fungal material is difficult to access for analysis. So far, isotope analyses have been limited to lipid compounds associated with fungal membranes or storage structures (biomarkers), fungal spores and soil hyphae. However, it remains unclear whether any of these components are an ideal substitute for intraradical AM hyphae as the functional nutrient trading organ. Thus, we isolated intraradical hyphae of the AM fungus Rhizophagus irregularis from roots of the grass Festuca ovina and the legume Medicago sativa via an enzymatic and a mechanical approach. In addition, extraradical hyphae were isolated from a sand-soil mix associated with each plant. All three approaches revealed comparable isotope signatures of R. irregularis hyphae. The hyphae were 13C- and 15N-enriched relative to leaves and roots irrespective of the plant partner, while they were enriched only in 15N compared with soil. The 13C enrichment of AM hyphae implies a plant carbohydrate source, whereby the enrichment was likely reduced by an additional plant lipid source. The 15N enrichment indicates the potential of AM fungi to gain nitrogen from an organic source. Our isotope signatures of the investigated AM fungus support recent findings for mycoheterotrophic plants which are suggested to mirror the associated AM fungi isotope composition. Stable isotope natural abundances of intraradical AM hyphae as the functional trading organ for bi-directional carbon-for-mineral nutrient exchanges complement data on spores and membrane biomarkers.

While it has long been held that invasive plants alter ecosystem processes, the magnitude and direction of these effects have rarely been quantified in situ. We measured the effects of an invasive C4 grass (Microstegium vimineum) on soil organic matter (SOM) decomposition in a deciduous forest in south-central Indiana, USA. The unique 13C signature of the C4 grass relative to the C3 trees allowed us to partition soil CO2 fluxes and estimate M. vimineum effects on decomposition. The magnitude and direction of priming effects hinged on the soil characteristics, which related to the mycorrhizal association of dominant trees. In forest plots dominated by ectomycorrhizal trees, with low nitrogen availability and most SOM in particulate (i.e., unprotected) forms, M. vimineum increased SOM decomposition by 58%. In contrast, in plots dominated by arbuscular mycorrhizal trees, characterized by high nitrogen availability and most SOM in mineral-associated (i.e., protected) forms, M. vimineum decreased decomposition by 14%. Collectively, our results demonstrate that invasive species can play a large role in altering ecosystem processes and suggest that the magnitude and direction of such effects depend on the dominant trees and edaphic characteristics of the stand.

Knowledge remains limited on how the structure of microbial community in paddy soils changes in relation to different types of fertilizers with same amount of nutrients. Thus, here, soil samples were collected at 0–10, 10–20, 20–30, and 30–40 cm depths from a paddy field subjected to four long-term fertilization treatments (no fertilization, mineral fertilization, mineral fertilization combined with rice straw, and chicken manure) and analyzed for microbial biomass and community composition. In unfertilized soils, microbial biomass decreased from 0 to 40 cm (with actinomycetes

Most subsoil carbon (C) turnover occurs in biopore hotspots such as root channels and earthworm burrows. Biopores allocate large C amounts into the subsoil, where a vast capacity for long-term C sequestration is predicted. We hypothesise that organic matter (OM) cycling in biopores depends on their origin. Earthworm and root biopores were induced under field conditions and were sampled from the subsoil (45–75 and 75–105 cm) after two years of biopore formation. The effects of biopore formation on OM decomposition were studied by biomarkers: neutral sugars, cutin and suberin-derived lipids, lignin-derived phenols and free lipids. The degradation stage of OM was biopore type-specific but was only governed by the soil depth in root biopores. Degradation of OM increased from earthworm biopores to root biopores and bulk soil. Hemicelluloses (GM/AX ratio) were more strongly degraded than lignin side-chains (relative change from initial values). Two years of microbial processing during biopore formation increased the GM/AX ratio in earthworm biopores from 0.65 to 1.05 and in root biopores from 0.15 to 1.35 (both relative to source biomasses). Root biopores and bulk soil had the highest GM/AX ratios (1.2–1.3), hinting to rapid processing of plant residues and accumulation of microbial residues. The regular, frequent OM inputs by earthworms stimulated microbial growth and processing of mostly bioavailable OM and, thus, relatively enriched more persistent OM (e.g. lignin). Syringyl subunits of lignin underwent low (ratio changed from 0.35 to 0.55 relative to initial input) and vanillyl subunits underwent almost no processing in earthworm biopores indicating the preferential microbial utilisation of the easily available compounds frequently replenished by earthworm activity. After two years of decomposition of the root detritus, mainly structural plant material was enriched in root biopores. Short periods (6 months) of earthworm activity effectively recharged the highly processed OM in root biopores with fresh OM. In total, deep-rooting catch crops and short-term earthworm activities promote C accumulation in the subsoil followed by biopore-specific microbial processing predominantly governed by the C input frequency. As root biopores are up to 40 times more common than earthworm biopores, they dominate the OM input into subsoils. Such C inputs create several years lasting hotspots for preferential root growth and nutrient mobilisation in the subsoil. We conclude that root- and earthworm-derived biopores are vertical pathways for plant C from the soil surface into the subsoil and for intensive processing of litter C and sequestration of microbial necromass.

This study assessed the carbon (C) budget and the C stocks in major compartments of the soil food web (bacteria, fungi, protists, nematodes, meso- and macrofauna) in an arable field with/without litter addition. The C stocks in the food web were more than three times higher in topsoil (0–10 cm) compared to subsoil (>40 cm). Microorganisms contained over 95% of food web C, with similar contributions of bacteria and fungi in topsoil. Litter addition did not alter C pools of soil biota after one growing season, except for the increase of fungi and fungal feeding nematodes in the topsoil. However, the C budget for functional groups changed with depth, particularly in the microfauna. This suggests food web resilience to litter amendment in terms of C pool sizes after one growing season. In contrast, the distinct depth dependent pattern indicates specific metacommunities, likely shaped by dominant abiotic and biotic habitat properties.

Hydrological cycle is expected to become the primary cause of ecosystem's degradation in near future under changing climate. Rain manipulation experiments under field conditions provide accurate picture on the responses of biotic processes to changed water availability for plants. A field experiment, mimicking expected changes in rain patterns, was established in a Mediterranean shrub community at Porto Conte, Italy, in 2001. In November 2011 Cistus monspeliensis, one of the dominating shrub species in the Mediterranean basin, was 13C labelled on plots subjected to extended rain shortage period and on control non manipulated plots. Carbon (C) allocation was traced by 13C dynamics in shoots, shoot-respired CO2, roots, microbial biomass, K2SO4-extractable C and CO2 respired from soil. Most of the recovered 13C (60%) was respired by shoots within 2 weeks in control plots. In rain shortage treatment, 13C remained incorporated in aboveground plant parts. Residence time of 13C in leaves was longer under the rain shortage because less 13C was lost by shoot respiration and because 13C was re-allocated to leaves from woody tissues. The belowground C sink was weak (3–4% of recovered 13C) and independent on rain manipulation. Extended rain shortage promoted C exudation into rhizosphere soil in expense of roots. Together with lowered photosynthesis, this “save” economy of new C metabolites reduces the growing season under rain shortage resulting in decrease of shrub cover and C losses from the system on the long-term.

Kobresia grasslands on the Tibetan Plateau comprise the world’s largest pastoral alpine ecosystem. Overgrazing-driven degradation strongly proceeded on this vulnerable grassland, but the mechanisms behind are still unclear. Plants must balance the costs of releasing C to soil against the benefits of accelerated microbial nutrient mineralization, which increases their availability for root uptake. To achieve the effect of grazing on this C-N exchange mechanism, a 15NH4+ field labeling experiment was implemented at grazed and ungrazed sites, with additional treatments of clipping and shading to reduce belowground C input by manipulating photosynthesis. Grazing reduced gross N mineralization rates by 18.7%, similar to shading and clipping. This indicates that shoot removal by grazing decreased belowground C input, thereby suppressing microbial N mining and overall soil N availability. Nevertheless, NH4+ uptake rate by plants at the grazed site was 1.4 times higher than at the ungrazed site, because plants increased N acquisition to meet the high N demands of shoot regrowth (compensatory growth: grazed > ungrazed). To enable efficient N uptake and regrowth, Kobresia plants have developed specific traits (i.e., efficient above-belowground interactions). These traits reflect important mechanisms of resilience and ecosystem stability under long-term moderate grazing in an N-limited environment. However, excessive (over)grazing might imbalance such C-N exchange and amplify plant N limitation, hampering productivity and pasture recovery over the long term. In this context, a reduction in grazing pressure provides a sustainable way to maintain soil fertility, C sequestration, efficient nutrient recycling, and overall ecosystem stability.

The importance of root hairs and root radius for exudation and nutrient acquisition by plants is known mainly from nutrient solution studies. The in situ effects of root hairs and root radius on the spatial distribution of enzyme activity in the rhizosphere of various plants are unknown. Four plants with contrasting root morphology (maize, wheat, lentil and lupine) were chosen to test the effects of root hairs and root radius on the spatial distribution of β-glucosidase, cellobiohydrolase, leucine aminopeptidase and acid phosphatase. We combined zymography with enzyme kinetics to evaluate the effects of root hairs on the rhizosphere extent and on substrate turnover. The extent of enzyme activity in the rhizosphere of four plants ranged from 0.55 to 2 mm. The extent of β-glucosidase was 1.5 times broader (1.2 mm versus 0.8 mm) and the substrate turnover was 2-fold faster around wheat root regions with hairs than hairless locations. The rhizosphere extent relative to root radius and the enzyme activity per root surface area were plant and enzyme specific: the rhizosphere extent was 1.5–2 times broader and the enzyme activity was 2–8-fold higher in wheat (with thin roots and long root hairs) compared to maize, lentil and lupine. The rhizosphere extent of acid phosphatase (1.1–2.0 mm) was 1.5–2-fold broader than that of other enzymes (0.5–1.0 mm). For the first time, we showed that the rhizosphere extent relative to root radius was 20–100% broader and enzyme activity per surface area was 4–7-fold higher around thin roots (wheat) than around thick roots (maize). Moreover, the rhizosphere extent relative to root radius was 10–30% broader and enzyme activity per root area was 2–7 times higher around roots with long and dense hairs (lupine) than around roots with short and sparse hairs (lentil). We conclude that root hairs and root radius shape the rhizosphere: root hairs contributed mainly to the rhizosphere extent, while root radius more strongly affected the enzyme activity per root surface area.

Background and Aims Although it is commonly accepted that root exudation enhances plant-microbial interactions in the rhizosphere, experimental data on the spatial distribution of exudates are scarce. Our hypothesis was that root hairs exude organic substances to enlarge the rhizosphere farther from the root surface. Methods Barley (Hordeum vulgare 'Pallas' - wild type) and its root-hairless mutant (brb) were grown in rhizoboxes and labelled with 14CO2. A filter paper was placed on the soil surface to capture, image and quantify root exudates. Key Results Plants with root hairs allocated more carbon (C) to roots (wild type: 13 %; brb: 8 % of assimilated 14C) and to rhizosheaths (wild type: 1.2 %; brb: 0.2 %), while hairless plants allocated more C to shoots (wild type: 65 %; brb: 75 %). Root hairs increased the radial rhizosphere extension three-fold, from 0.5 to 1.5 mm. Total exudation on filter paper was three times greater for wild type plants compared to the hairless mutant. Conclusion Root hairs increase exudation and spatial rhizosphere extension, which probably enhance rhizosphere interactions and nutrient cycling in larger soil volumes. Root hairs may therefore be beneficial to plants under nutrient-limiting conditions. The greater C allocation below ground in the presence of root hairs may additionally foster C sequestration.

In arable fields, biopores are primarily formed by taproots, but may also be bored by earthworms. Irrespective of the pore origin, repeated use by anecic earthworms yields a wall coating that is rich in carbon, nutrients and microorganisms. However, this effect is halted by routine tillage, and it remains unclear how quickly earthworms are able to alter biopore properties in subsoil. We conducted an earthworm incubation field experiment in arable soil to test the capacity of Lumbricus terrestri s to i. increase total nutrient contents including plant available P, ii. alter the microbial community and iii. increase enzyme activities in biopore walls over one vegetation period. Firstly, biopores that contained chicory roots were identified on a plot scale (4.2 × 1.5 m). After two years under fallow, roots were decomposed. We then inserted individual earthworms at 45 cm depth into a subset of these pores, afterwards refilling with topsoil. After six months, earthworms were removed and soil was opened at 45–75 cm and 75–105 cm soil depth layers. The inner pore wall (1 mm) of individual root biopores (‘RBP’) or root biopores modified by earthworms (‘EBP’) as well as the bulk soil were sampled in 6 depth intervals of 10 cm each and analyzed for total C, N, S content, plant available P, microbial biomass, phospholipid fatty acids (PLFA) and enzyme activity. Biochemical properties of bulk soil, RBP and EBP clearly differed after one vegetation period as indicated by principal component analysis. PLFA markers of fungi and protozoa were detected only in biopores. Compared with the bulk soil, total C, N, S were enriched in RBP by a factor of 2.0–3.1, plant available P by a factor of 8–10, and microbial biomass by a factor of 12–36. In EBP, all of these parameters were as in RBP or elevated even further (C, N, S: factor 1.0–1.4, plant available P: factor 1.3–1.5, microbial biomass: factor 1.5–2.0, PLFA markers of fungi: factor 2.6–4.4, PLFA markers of protozoa: factor 9.2–14.2). PLFA markers indicative of the ratio of Gram-positive to Gram-negative bacteria (G+: G−) were 5–10 fold lower in RBP than in bulk soil, the microbial metabolic quotient ( q CO 2 ) was 0.4–0.6 times as high. In EBP, these parameters were further reduced (ratio G+: G−: factor 0.7, q CO 2 : factor 0.7–0.8). RBP were particularly characterized by high contents of 10-methyl branched fatty acid indicators of actinobacteria. Activities of enzymes involved in the C-cycle (xylanase, cellobiohydrolase, s-glucosidase) and N-cycle (chitinase, chitotriosidase, leucine aminopeptidase) were also elevated in RBP as compared to the bulk soil (factor 1.1–3.6) and further increased in EBP (factor 1.2–3.7). All these effects were more pronounced in the 45–75 cm soil layer. We conclude that, in only six months, L. terrestris in arable fields modified ordinarily nutrient-rich biopores into ‘super-hotspots’ of microbial biomass, enzyme activity and nutrient availabilities. Hence, even short-term promotion of earthworm populations by agricultural management practices can increase microbial biomass and enzyme activity in biopores and its coupling to nutrient mobilization in the subsoil.