Ecoenzymatic Stoichiometry Research Articles

Intercropping with legumes alleviates soil N limitation but aggravates P limitation in a degraded agroecosystem as shown by ecoenzymatic stoichiometry

Agroforestry is a commonly applied restoration practice in nutrient-poor or degraded soils, and knowledge of soil nutrient limitations is critical for its successful implementation. Ecoenzymatic stoichiometric theory (EEST) models have been proposed as easy and fast methods to estimate soil nutrient limitations in terrestrial ecosystems; however, a comprehensive application of different models in agricultural studies is still lacking. To bridge these gaps, in this study, we analysed the nutrient limitation of three plantation types, Camellia oleifera monoculture (CK), C. oleifera–Arachis hypogaea (peanut) intercropping (CP), and C. oleifera–Senna tora intercropping (CS), within a fully controlled C. oleifera agroforestry system in nutrient-poor purple soils. We compared the estimates of the nutritional limitations of each plantation type using three different EEST models: vector, vector-threshold element ratio (V-T), and threshold. Estimates of nutrient limitation calculated using the threshold model recapitulated those calculated using the other models. Further, N and P limitations existed in all three plantation types, C limitation was not detected in any plantation type, and the CP plantation type alleviated N limitation and aggravated P limitation. These results were consistent with the actual nutrient characteristics of the purple soil, indicating that the microbial metabolic limitation profile estimated by the EEST model was reliable. The results of this study suggest that EEST models are valuable tools for estimating soil nutrient limitation in the context of agricultural ecosystem restoration, such as in agroforestry.

Exogenous nitrogen input skews estimates of microbial nitrogen use efficiency by ecoenzymatic stoichiometry

BackgroundEcoenzymatic stoichiometry models (EEST) are often used to evaluate microbial nutrient use efficiency, but the validity of these models under exogenous nitrogen (N) input has never been clarified. Here, we investigated the effects of long-term N addition (as urea) on microbial N use efficiency (NUE), compared EEST and 18O-labeling methods for determining NUE, and evaluated EEST’s theoretical assumption that the ratios of standard ecoenzymatic activities balance resource availability with microbial demand.ResultsWe found that NUE estimated by EEST ranged from 0.94 to 0.98. In contrast, estimates of NUE by the 18O-labeling method ranged from 0.07 to 0.30. The large differences in NUE values estimated by the two methods may be because the sum of β-N-acetylglucosaminidase and leucine aminopeptidase activities in the EEST model was not limited to microbial N acquisition under exogenous N inputs, resulting in an overestimation of microbial NUE by EEST. In addition, the acquisition of carbon by N-acquiring enzymes also likely interferes with the evaluation of NUE by EEST.ConclusionsOur results demonstrate that caution must be exercised when using EEST to evaluate NUE under exogenous N inputs that may skew standard enzyme assays.

Soil extracellular enzyme stoichiometry reveals the increased P limitation of microbial metabolism after the mixed cultivation of Korean pine and Manchurian walnut in Northeast China

Korean pine (Pinus koraiensis) and Manchurian walnut (Juglans mandshurica) are the main tree species used for plantation regeneration in Northeast China, and their mixed cultivation is typically used to address a decline in stand productivity in long-term monocultures. However, little is known about the changes in the C, N and P requirements of soil microbial metabolism between monocultures and mixed plantations. In this study, we examined the effect of plantation type on the soil microbial metabolic limitations by investigating the soil extracellular enzyme activities across three plantation types (P. koraiensis monocultures, J. mandshurica monocultures, P. koraiensis and J. mandshurica mixed plantations) of the same age (32 years old). We used ecoenzymatic stoichiometry methods to reveal the patterns of microbial metabolic C, N and P limitations along a soil depth gradient of 0–10, 10–20, and 20–40 cm by collecting a total of 81 composite soil samples including three plantation types. With the increased soil depth, the activities of C-, N- and P-acquiring enzymes in mixed plantations decreased from 93.60 nmol g−1 soil h−1 to 20.52 nmol g−1 soil h−1, 33.46 nmol g−1 soil h−1 to 22.09 nmol g−1 soil h−1, and 11.80 nmol g−1 soil h−1 to 7.30 nmol g−1 soil h−1, respectively. Enzymatic vector analysis (vector angle >45°) revealed P limitation rather than N limitation of microbial metabolism across all plantation types. Of the plantations, the mixed stands had the greatest vector length (1.15 ± 0.02) and vector angle (56.91 ± 1.47). The interactions of plantation type and soil depth had significant effects on microbial metabolic C and P requirements. Changes in soil available nutrient contents and stoichiometry ratios of C, N and P across different plantation types were the two most important soil variables affecting microbial metabolic C and P requirements.

Linking soil depth to aridity effects on soil microbial community composition, diversity and resource limitation

With ongoing climate change, aridity is increasing worldwide, affecting biodiversity and ecosystem function in drylands. However, how the depth-profile microbial community structure and metabolic limitations change along aridity gradients are still poorly explored. Here, 16S rRNA and ITS amplicon sequencing and ecoenzymatic stoichiometry analysis were used to investigate both bacterial and fungal diversities and resource limitations in 1 m depth profiles across a wide aridity gradient (0.51–0.78) in a semiarid region. Results showed a sharp decrease in microbial diversity with soil depth, accompanied by an increase in microbial phosphorus (P) vs. N (nitrogen) limitation and a decrease in microbial carbon (C) vs. nutrient limitation. Aridity led to a strong shift in microbial community composition, but aridity has a threshold effect on microbial resource limitation through impacts on soil pH and C/P or N/P. When the aridity threshold (1-precipitation/evapotranspiration) exceeds 0.65, relationship between aridity and microbial resource demand was decoupled; but at aridity threshold = 0.65, microbial relative C limitation and C-acquiring enzyme activity dropped. These results suggest that aridity might have a stronger influence on microbial community composition, than on diversity, shaped by inherent soil biotic factors (i.e., MBC:MBP or MBN:MBP). These findings suggest that soil microbial diversity or enzymatic stoichiometry may be not necessary to mirror changes in water availability in the drylands, while aridity would be well explained by microbial community composition.

Microbial carbon and nitrogen limitations with mulching of proso millet fields on the Loess Plateau: Evidence from soil ecoenzymatic stoichiometry

AbstractMulching measures can regulate soil properties; however, little is known about the effects metabolic limitations on farmland during the key growth stages of broomcorn millet in multiple regions of the Loess Plateau. We conducted field experiments to compare three techniques: flat planting with no mulching (TP), ridge–furrow mulching system (RF), and plastic film mulching (PFM). Soil extracellular enzymatic stoichiometry and physicochemical properties of three growth periods (jointing, flowering, and maturity) of proso millet (Panicum miliaceum L.) were measured to investigate microbial metabolic limitations and the relationship with soil moisture, temperature, and nutrients in the three regions of the Loess Plateau (Guyuan city, Huining County, and Yulin city). The results show that compared with TP, both PFM and RF techniques increased soil organic carbon (SOC), total nitrogen (TN), ammonium nitrogen (), and nitrate nitrogen () during the jointing period, but the levels decreased during the flowering period, and the activities of C‐, N‐, P‐acquiring enzymes were 29.02%, 33.68%, and 19.46% higher when using PFM, and 13.78%, 6.81%, and 6.52% higher when using RF. Meanwhile, RF treatment significantly increased the carbon metabolism limitation during the jointing, flowering, and maturity periods of proso millet in the three regions, and also improved the nitrogen metabolism limitation during the jointing and flowering periods of proso millet in the Huining and Yulin regions. Linear regression analysis showed that pH, SOC, and contents significantly affected carbon limitation, and nitrogen limitation was gradually alleviated with increases in SOC, TN, and contents in proso millet farmland soils. Partial least squares path modeling showed that soil moisture and nutrients differed significantly among the regions, and soil temperature positively regulated the soil nutrients. Mulching significantly improved the carbon limitation owing to increased soil temperature and moisture. These results provide important ideas for nutrient cycling and microbial metabolism of broomcorn millet farmland soil under mulching measures on the Loess Plateau.

Ecoenzymatic Stoichiometry in the Rhizosphere and Bulk Soil of a Larix principis-rupprechtii Plantation in North China

Soil extracellular enzymes play an important role in ecosystem energy conversion and material cycling. Ecoenzymatic stoichiometry can reflect the relationship between the soil’s microbial nutrient cycle and nutrient limitation. However, there have been few studies on the differences in ecoenzymatic stoichiometry and nutrient limitation between rhizosphere soil and bulk soil. This study examined soil nutrients and enzyme activities in rhizosphere soil and bulk soil in a Larix principis-rupprechtii plantation in north China. The results showed that the levels of soil organic carbon (C), total nitrogen (N), and available nutrients in the rhizosphere soil were significantly higher than those in the bulk soil, whereas the total potassium (TK) level was significantly lower. The soil C:N, C:P, and N:P ratios of the rhizosphere soil also exceeded those of the bulk soil. The acid phosphatase (ACP), urease (UE), and β-glucosidase (β-GC) activities in the rhizosphere soil exceeded those in the bulk soil, whereas the activities of N-acetyl-β-D-glucosidase (NAG), aminopeptidase (LAP), and nitrogenase (NA) were lower. The ratios of C, N, and P acquisition activities changed from 1:1.7:1 in the rhizosphere soil to 1:2:1 in the bulk soil. Redundancy analysis showed that the available K and soil water content in the rhizosphere soil were the most important soil factors affecting soil enzyme activities and ecoenzymatic stoichiometry; those in the bulk soil were soil N:P and soil water content. These results suggest that not all soil enzyme activities present rhizosphere effects and that bulk soil is more susceptible to N limitation in Larix principis-rupprechtii plantations. Plant roots play an important role in regulating soil nutrients and soil activities, and future studies should examine the underlying mechanisms in more detail.

Bacteria life-history strategies and the linkage of soil C-N-P stoichiometry to microbial resource limitation differed in karst and non-karst plantation forests in southwest China

Soil microbial resource-acquisition strategies play a crucial role in soil nutrient cycling and the accumulation of soil organic carbon (SOC) in vegetation restoration. Despite the growing interest in soil microbial resource limitation, the impact of lithology on microbial resource limitation and its relationship with soil carbon–nitrogen-phosphorus (C-N-P) stoichiometry is not well understood. Therefore, we investigated the soil C-N-P and ecoenzymatic stoichiometry, bacterial life-history strategies, and microbial resource limitation in two common plantation forests (Pinus yunnanensis Franch. (PY) and Eucalyptus maideni F. Muell. (EM)) in karst and non-karst areas in southwest China. These areas are characterized by soils derived from limestone and clastic rock, respectively. The results showed that (1) soil nutrients, SOC concentrations and ecoenzymatic activities were significantly higher in karst plantation forests compared to non-karst, except for dissolved inorganic phosphorus; (2) soil microorganisms in both lithology were largely co-limited by C and P in EM plantation while the PY plantation soil in organic horizon primarily limited by P, which might be due to a much higher ratio of soil C:P and N:P; (3) lithology affects the associations between soil C-N-P stoichiometry and microbial resource limitation; (4) redundancy analysis showed that the ratio of C:N acquiring enzyme was a substantially predictor for microbial resource limitation in both karst and non-karst soils; (5) karst soils had a higher proportion of species affiliated with oligotrophs bacteria. Overall, these findings improve our knowledge of microbial resource limitation over limestone and clastic rock and its relationship with soil C-N-P and ecoenzymatic stoichiometry, as well as the lithology effects on bacteria life-history strategies.

Ecoenzymatic Stoichiometry Reveals Microbial Carbon and Phosphorus Limitations under Elevated CO2, Warming and Drought at Different Winter Wheat Growth Stages

The use of microbial metabolic limitation techniques has the potential to provide insights into carbon and nutrient cycling in an ecosystem under the influence of climate change. This study aimed to determine the characteristics and potential mechanisms of microbial metabolic limitation at the different growth stages of winter wheat (Triticum aestivum L.) in response to elevated CO2 concentrations, warming and drought. Winter wheat plants were grown in artificial climate chambers, and a set of treatments were employed, including two levels of CO2 concentration (400 and 800 μmol·mol−1), a temperature regime (the current ambient temperature and a temperature increase of 4 °C) and water conditions (80% and 60% of the field water capacity). The results showed that the soil microbes were mainly limited by C and P. Microbial C limitation significantly decreased by 26.7% and 36.9% at the jointing stage and significantly increased by 47.6% and 42.6% at the grain filling stage in response to elevated CO2 and warming, respectively. The microbial P limitation significantly decreased by 10.9–13.0% under elevated CO2 at the anthesis and grain filling stages, while it was not affected by warming. Both microbial C and P limitations were unaffected by drought. The growth stage, soil dissolved organic carbon (DOC) and available phosphorus (AP) were the key factors affecting microbial C limitation, and microbial P limitation was mainly affected by the soil microbial biomass carbon (MBC), phosphorus (MBP) and microbial C:P ratio. Thus, the soil microbial C and P limitations differed with growth stages and were primarily indirectly affected by the available nutrients in the soil and the properties of the microbial biomass, respectively. These findings are important for understanding the mechanisms underlying microbe-mediated C and nutrient cycles. Overall, this study provides guidance for soil nutrient management in an agroecosystem experiencing climate change.

Ecoenzymatic stoichiometry reveals soil P limitation under biochar addition in a reclaimed mine area in Shanxi Province, China

Extracellular enzymatic activity plays an important role in carbon (C), nitrogen (N), and phosphorous (P) cycling, and consequently, serves as an indicator for soil function. However, microbial activities are inhibited in degraded lands due to nutrition limitations, which pose a challenge to ecological management. We investigated the effects of maize biochar addition (1, 3, and 5%) on soil and enzyme stoichiometries in a post‐mining area in Shanxi province. The results showed that soil organic C and total N increased significantly with increasing biochar addition, leading to significant increases in soil C:P (by 157.9%) and N:P (by 76.1%) under 5% of addition. Biochar addition stimulated enzyme activities significantly, and the maximum of enzyme activities occurred in treatment of 5% addition. Soil enzyme C:N increased significantly following biochar addition, while enzyme C:P and N:P decreased at 5% biochar addition. Vector analysis indicated that microbial activity tended to be limited by soil N before addition, whereas it was limited by P availability at high biochar addition. This was mainly because that biochar addition could promote sorption of P and lower P bioavailability in soil. Our research suggested that biochar application could substantially improve soil nutrition status and microbial activity in infertile land, however, it would likely result in imbalances between substrate stoichiometry and microbial demand. The result might provide a reference for the management of biochar application during soil restoration.

Soil extracellular enzymes characteristics and their controlling factors along the elevation gradient in Qinghai-Tibet Plateau, China

Soil extracellular enzyme activities (EEAs) and ecoenzymatic stoichiometry (EES) play an essential role in soil nutrient cycling and organic matter decomposition. Understanding EEAs and EES variation patterns and their influencing factors could offer direct information about the soil structure, function, and soil response to anthropogenic disturbances and climate change. This issue is noteworthy, especially in high-altitude areas where climate change is imminent and vegetation is diversified. This study measured different soil EEAs and EES characteristics and explored their key controlling factors along nine altitudes ranging from 2500 m to over 5200 m in the Qinghai-Tibet Plateau of western China. We also analyzed the effects of plant microhabitats on soil EEAs and EES. The results showed that most soil EEAs and EES had significant variability in spatial characteristics, and enzymatic activity increased with altitude. Compared to the soil nutrient distribution which also increased with altitude, this same change trend of soil EEAs and soil nutrients was inconsistent with the resource allocation theory. Microorganisms might mediate the effects of environmental factors on soil EEAs by altering the enzyme production efficiency. Specific soil EEAs (EEAs/g SOC), like soil enzyme carbon: phosphorus ratios (ECP), and nitrogen: phosphorus ratios (ENP), showed an opposing trend in variation, which decreased with increasing altitude. Plant microhabitats significantly promoted soil EEAs due to the accumulation of soil nutrients (carbon, nitrogen and phosphorus). Soil EEAs and EES's spatial variability was mainly determined by edaphic factors, accounting for >70.24 % and 55.67 % of latitudinal variations, respectively. Generally, carbon and nitrogen limitations were substantial in this area and gradually alleviated with increasing altitude. This study provided a data support for ecological protection of the Qinghai-Tibet Plateau based on the spatial variation of soil EEAs and nutrient limitation.

Imprint of tree species mycorrhizal association on microbial‐mediated enzyme activity and stoichiometry

Abstract Understanding the effects of tree species and their mycorrhizal association on soil processes is critical for predicting the ecosystem consequences of species shifts owing to global change and forest management decisions. While it is well established that forests dominated by different mycorrhizal types can vary in how they cycle carbon (C), nitrogen (N) and phosphorus (P), the degree to which these patterns are driven by microbial‐mediated enzyme activity (EA) and ecoenzymatic stoichiometry (ES) remains elusive. Here, we synthesized the effects of mycorrhizal association on seven soil enzymes involved in microbial C, N and P acquisition and ES using data from 56 peer‐reviewed papers. We found that relative to soil in ectomycorrhizal (EcM) trees, soil in arbuscular mycorrhizal (AM) trees exhibited greater activity of some C acquisition enzymes (e.g. beta‐glucosidase; BG) and higher ecoenzymatic ratios of BG/NAG (N‐acetyl‐glucosaminidase) and BG/AP (acid phosphatase). These results supported that AM trees had rapid C and nutrient turnover rates, inorganic nutrient economics and high soil microbial C limitation. We also found evidence for an organic nutrient economy and greater soil microbial demand for nutrients in EcM trees compared to AM trees. In addition, the effect of mycorrhizal association on the activity of certain soil enzymes and enzymatic stoichiometry (i.e. BG and BG/NAG ratio) appeared to be associated with the differences in soil pH, phylogenetic group (i.e. conifers and broadleaves) and leaf habit (i.e. evergreen and deciduous) between AM and EcM trees. The results from the global meta‐analysis suggested that soil EA and ES appear to play critical roles in shaping the differences in the nutrient economy between AM and EcM tree species, but leaf morphology and soil conditions should be considered in evaluations of soil processes in forests of different mycorrhizal associations. Given that most of the studies in the database were from the temperate and subtropical regions, further research in other biomes is needed to elucidate the underlying mechanisms driving the mycorrhizal effect at the global scale. Read the free Plain Language Summary for this article on the Journal blog.

Rural mixed pond water irrigation affected microbial community and eco-enzymatic stoichiometry of soil profiles

The application of rural mixed pond water (RMW) for irrigation has become a relatively normal agricultural practice in China. However, it is not clear whether soil microbial activity and soil stored nutrients are influenced by RMW. In this study, soil samples were collected from a depth of 0–60 cm for determining their basic physicochemical properties, microbial biomass and soil enzyme activities, moreover, the eco-enzymatic stoichiometry ratio was determined to evaluate microbial nutrient requirements. Our results showed that, with the increase in the soil depth, dissolved organic carbon, available phosphorus, microbial contents, and extracellular enzyme activities declined. We observed an increment in the microbial biomass nitrogen (MBN; 1.44 fold) and microbial biomass carbon (MBC; 1.23 fold) from 0 to 10 cm after RMW irrigation. The latter was more stable below 40 cm of the soil layer independent of irrigation water. The increase in carrier length and angular carrier indicated that the microorganisms were limited by carbon (C) and phosphorus (P). The RMW irrigation significantly decreased the β-1,4-Nacetylglucosaminidase (NAG), the leucine aminopeptidase (LAP), and the phosphatase (AP). In addition, RMW irrigation increased the α-diversity of surface soil microorganisms but suppressed them in deeper soil profiles. The RDA analysis suggested that random NAG of exoenzymes had the greatest relative contribution to microbial diversity, with bacteria and fungi accounting for 27.1 % and 25.1 %, respectively. This study showed that RMW irrigation led to changes the soil extracellular enzyme activities, resulting in alterations the enzyme stoichiometry ratios and microbial nutrient acquisition strategies.

Soil arthropods promote litter enzyme activity by regulating microbial carbon limitation and ecoenzymatic stoichiometry in a subalpine forest

Soil arthropods are crucial decomposers of litter at both global and local scales, yet their functional roles in mediating microbial activity during litter decomposition remain poorly understood. Here, we conducted a two-year field experiment using litterbags to assess the effects of soil arthropods on the extracellular enzyme activities (EEAs) in two litter substrates (Abies faxoniana and Betula albosinensis) in a subalpine forest. A biocide (naphthalene) was used to permit (nonnaphthalene) or exclude (naphthalene application) the presence of soil arthropods in litterbags during decomposition. Our results showed that biocide application was effective in reducing the abundance of soil arthropods in litterbags, with the density and species richness of soil arthropods decreasing by 64.18–75.45 % and 39.19–63.30 %, respectively. Litter with soil arthropods had a greater activity of C-degrading (β-glucosidase, cellobiohydrolase, polyphenol oxidase, peroxidase), N-degrading (N-acetyl-β-D-glucosaminidase, leucine arylamidase) and P-degrading (phosphatase) enzymes than litter from which soil arthropods were excluded. The contributions of soil arthropods to C-, N- and P-degrading EEAs in the fir litter were 38.09 %, 15.62 % and 61.69 %, and those for the birch litter were 27.97 %, 29.18 % and 30.40 %, respectively. Furthermore, the stoichiometric analyses of enzyme activity indicated that there was potential C and P colimitation in both the soil arthropod inclusion and exclusion litterbags, and the presence of soil arthropods decreased C limitation in the two litter species. Our structural equation models suggested that soil arthropods indirectly promoted C-, N- and P-degrading EEAs by regulating the litter C content and litter stoichiometry (e.g., N/P, LN/N and C/P) during litter decomposition. These results demonstrate that soil arthropods play an important functional role in modulating EEAs during litter decomposition.

Effects of global change on the ability of stream biofilm to dissipate the herbicide glyphosate

The herbicide glyphosate is contaminating a large number of freshwater ecosystems worldwide and its fate and effects remains uncertain in light of the effects of global change. The present study examines how variations in water temperature and light availability relative to global change affect the ability of stream biofilms to degrade the herbicide glyphosate. Biofilms were exposed in microcosms to two levels of water temperature simulating global warming (Ambient = 19–22 °C and Warm = 21–24 °C) and three levels of light representative of riparian habitat destruction due to land use change (Dark = 0, Intermediate = 600, High = 1200 μmol photons m−2 s−1). Biofilms were acclimated to six different experimental treatments, namely i) ambient temperature without light (AMB_D), ii) ambient temperature and intermediate light (AMB_IL), iii) ambient temperature and high light (AMB_HL), iv) warm temperature without light (WARM_D), v) warm temperature and intermediate light (WARM_IL) and vi) warm temperature and high light (WARM_HL). The ability of biofilms to degrade 50 μg L−1 of glyphosate was tested. Results showed that water temperature increase, but not light availability increase, significantly increased aminomethyl phosphonic acid (AMPA) production by biofilms. However, the combined increase of temperature and light generated the shortest time to dissipate half of the glyphosate supplied and/or half of the maximum AMPA produced (6.4 and 5.4 days, respectively) by biofilms. Despite light had a major effect in modulating biofilm structural and functional descriptors, the response of certain descriptors (i. e. chlorophyll-a concentration, bacterial density and diversity, nutrient content and PHO activity) to light availability increase depended on water temperature. Specifically, the biofilms in the WARM_HL treatment displayed the highest Glucosidase: Peptidase and Glucosidase: Phosphatase enzyme activity ratios and the lowest biomass C: N molar ratios compared to the other treatments. According to these results, warmer temperatures and high light availability could have been exacerbating the decomposition of organic C compounds in biofilms, including the use of glyphosate as a C source for microbial heterotrophs. This study shows that ecoenzymatic stoichiometry and xenobiotic biodegradation approaches can be combined to better understand the functioning of biofilms in pesticide-polluted streams.

Interpreting patterns of ecoenzymatic stoichiometry

The stoichiometry of extracellular enzyme activities has been both proposed and refuted to quantify resource use efficiency and resource limitations to soil microbial communities. The approach evaluates the acquisition of resources obtained through the enzyme-mediated catalysis of polymeric substrates. However, labile resources that do not require enzyme activity circumvent this catalytic pathway, can alter the balance of enzyme activities and skew their interpretation. More specifically, the microbial use of soluble resources can occur within minutes to hours of their addition whereas the use of polymeric resources depends on the production and turnover of extracellular enzymes (days to weeks). This temporal difference between potential, short-term microbial metabolic responses to soluble resources and longer-term community responses to enzyme-mediated acquisition of polymeric resources can produce seemingly conflicting interpretations of microbial resource limitations. However, stoichiometric tools provide insights to microbial resource use and limitations that differ in functional context from other measures of microbial community behavior.

Soil bacteria respond intensely to resource limitations regulated by edaphic properties during secondary succession on a semiarid abandoned farmland

Disproportionate inputs of carbon (C), nitrogen (N), and phosphorus (P) following plant succession typically induce microbial resource limitations. Such limitations are directly relevant to the fate of soil element cycles. However, it is not well understood how generalizable patterns of resource constraints act on soil microbial communities during the natural succession of abandoned farmlands, nor their driving forces. To this end, the potential activities of C-, N-, and P-acquiring enzymes, soil properties, plant characteristics, and soil microbial community composition and diversity were investigated along a 30-year successional chronosequence following agricultural abandonment on the Chinese Loess Plateau. Ecoenzymatic stoichiometry revealed a decrease in C limitation following succession, and soil microorganisms were primarily limited by N in the 10th year of succession but limited by P in the 20th and 30th years. Soil physicochemical properties cumulatively contributed 62.59% and 59.33% to the variations in microbial C and nutrient limitations, respectively. In particular, soil organic C content, C:N and C:P ratios, microbial biomass, and pH strongly affected microbial element limitations. Plant characteristics had minor effects on microbial resource constraints, accounting for 37.41% and 40.67% of the variance in microbial C and nutrient limitations, respectively. In comparison, plant diversity affected soil microbial element limitations more than plant family composition. Notably, the bacterial alpha and beta diversities, as well as the relative abundances of dominant phyla, such as Actinobacteria, Bacteroidetes, Proteobacteria, Acidobacteria, Gemmatimonadetes, and Planctomycetes, were significantly more associated with resource limitations than soil fungal community. Collectively, these findings suggest that the variable patterns of microbial element limitations were predominantly the result of changing soil properties across distinct successional stages, with soil bacteria responding more strongly than fungi to changes in microbial element limitations.

Features and driving factors of microbial metabolic limitation in mountain ecosystems in arid areas: A case study on the Helan Mountains, Northwest China

Insights into what limits the growth of soil microorganisms in mountain ecosystems increase our understanding of microbial functions and processes. Although the distribution pattern of soil microorganisms in mountain ecosystems has been widely studied, their role in biogeochemical cycles along elevation gradients of mountain ecosystems in arid regions is poorly understood. In this study we analyzed the soil physicochemical properties, soil microbial community structure, extracellular enzymatic activities, ecoenzymatic stoichiometry, microbial metabolism, and their relationships along the 1,300–2,500 m elevational gradient of the Helan Mountains, northwest China. The results showed that the total microbial biomass and its components did not significantly vary with elevation. The GP:GN (gram-positive: gram-negative bacteria) ratios at low elevations were higher than those at the mid and high elevations, indicating enrichment of oligotrophic bacteria at low elevations. The five extracellular enzymes significantly differed with elevation gradient, while the levels of carbon (C)- and nitrogen (N)- acquiring enzymes first increased and then decreased with increasing elevations. Ecoenzymatic stoichiometry indicated that significant limitation of microbial growth by carbon (C) and phosphorus (P) levels occurred at high and medium elevations. Soil physicochemical characteristics, microbial community composition, and ecoenzymatic activities accounted for 43.94 and 22.21% of the microbial C and P restriction, respectively. Our study suggests that mountain ecosystems with high organic C storage possess abundant microbial populations limited by relative C and P. The study also provides important insights linking microbial metabolisms to the environmental gradients in arid mountain ecosystems.

New insights into the patterns of ecoenzymatic stoichiometry in soil and sediment

Ecoenzymatic stoichiometry connects the elemental stoichiometry of microbial biomass and detrital organic matter to microbial nutrient assimilation and growth, and thus has been proposed and developed to estimate microbial nutrient limitations. However, the theories and methods developed so far have not yet clearly explored the critical thresholds of ecoenzymatic stoichiometry that identify whether nutrient limitation occurs or not. Here, we developed methods quantifying critical thresholds for microbial carbon (C), nitrogen (N) and phosphorus (P) limitations by centering on the potential responses of microbial resource use efficiency to nutrient limitations. We also provided new insight into the relationships of bivariate regressions between C-, N- and P-acquiring ecoenzymatic activities by reconsidering the intercept of regressions as an index to initial nutrient limitations, and consequently distinguished six initial limitation scenarios based on the range of values for these intercepts. According to the empirical values of the regression intercepts, the overall frequency of primary nutrient limitations in microbial communities across a global suite of soils and sediments is P limitation > N limitation > C limitation. The result implies that P and N availabilities rather than that of C could be the most important limitations on microbial production and metabolism in terrestrial soils and freshwater sediments. Nutrient limitations in heterotrophic microbial communities are fundamental characteristics of both soil and sediment systems, which regulate organic matter decomposition and element cycles. Our perspective provides a foundational framework and avenue for understanding and quantifying microbial nutrient limitations in studies of terrestrial C cycling and microbial ecology.

Soil aggregate development and associated microbial metabolic limitations alter grassland carbon storage following livestock removal

Natural restoration of degraded grasslands following livestock removal changed soil nutrient and carbon concentrations, soil aggregate distributions, and patterns of extracellular enzyme activities. Here, we used an ecoenzymatic stoichiometry model to quantify microbial resource limitations in semiarid grassland soil aggregates at 0, 11, 26, and 36 years after livestock removal and linked these limitations to microbial carbon use efficiency (CUE), which was estimated from stoichiometry theory (CUEST). Overall, livestock removal altered the size distribution of soil aggregates, increased microbial resources and stimulated extracellular enzyme activities. Long-term livestock removal increased the proportion of small macroaggregates (2–0.25 mm). Microbial carbon (C) and phosphorus (P) limitations in soil aggregates declined to minimum levels at 26 years following livestock removal and then increased after 36 years, with an opposite trend for CUEST. Additionally, the greatest resource limitations to microorganisms were found in smaller aggregates (<2 mm) under long-term livestock removal. Random forest and structural equation models revealed that soil abiotic factors, especially total nitrogen and pH, were key determinants of microbial resource limitation in soil aggregates. Moreover, microbial C and P limitations had significant direct effects on the CUEST. Thus, increasing microbial metabolic limitations after long-term livestock removal could reduce microbial C turnover, potentially reducing soil C sequestration. Overall, this study revealed that livestock removal altered soil aggregate development as well as the allocation of resources in aggregates and consequent microbial resource limitations, providing information that may be useful for developing grassland management strategies to enhance C sequestration.

Eco-enzymatic stoichiometry and microbial non-homeostatic regulation depend on relative resource availability during litter decomposition

The relationships between soil eco-enzymatic stoichiometry and substrate stoichiometry could be used to explain nutrient cycling processes regulated by microbial biomass stoichiometry. But what are the relationships between the stoichiometry of resources, microorganisms, and enzymes? We do not have a clear understanding yet, particularly in the different stages of litter decomposition. By analyzing the response of extracellular enzymatic stoichiometry to the imbalance between microbial and substrate stoichiometry, we explore the microbial-mediated litter transformation and microbial adaptation mechanisms in the transformation interface soil (TIS) layer. The results showed that soil microbial metabolism was limited by C and P during decomposition, peaking in the transition (middle) stage and then gradually decreasing. According to the judgment of microbial homeostasis (effect of substrate stoichiometry on microbial biomass stoichiometry), the microbial communities shift from homeostatic in the early stage to non-homeostatic in the middle stage and then return to homeostasis in the later stage. During the litter decomposition, the non-homeostatic period corresponded to the transition stage. That means severe microbial metabolic limitation is a potential determinant of microbial homeostasis. Soil microorganisms adapt and alleviate microbial metabolic limitations by adjusting extracellular enzyme activity and microbial non-homeostatic behavior when the available resources fail to satisfy microbial requirements.