Regulation of extracellular protease activity in soil in response to different sources and concentrations of nitrogen and carbon

A large proportion of the nitrogen (N) in soil is in the form of proteinaceous material. Its breakdown requires the activity of extracellular proteases and other decomposing enzymes. The goal of our study was to better understand how carbon (C) and N availability affect soil protease activity. Several aerobic incubations were carried out with ammonium (NH₄⁺) and proteins as N sources and cellulose as the main C source. A strong increase in protease activity was observed when proteins were added, the increase depending on the amount of protein added and its solubility. Protease synthesis was clearly substrate induced, as NH₄⁺ had no effect. During this substrate induced phase, the addition of glucose but not NH₄⁺ resulted in protease repression, indicating that the level of protease synthesis was determined by the need for C rather than N. After 1 month of incubation, protease activity remained relatively constant over time and was closely related to microbial biomass N. Different concentrations of mineral N in soil solution had no direct effect on protease activity. However, during this stationary phase, protease activity could be repressed by glucose and NH₄⁺ in a treatment with low mineral N content while in treatments with a higher N availability no repression was observed. We hypothesize that the need for N determined protease activity in the treatment with limited N availability. The addition of NH₄⁺ allowed for reallocation of C and N away from protease synthesis, leading to the observed decrease in protease activity. The repression by glucose may be attributed to shifts in the pathway of microbial NH₄⁺ assimilation. The results emphasize the close links between the microbially mediated cycles of organic C and N.     

Seasonal protein dynamics in Alaskan arctic tundra soils

In the arctic tundra of Alaska, plant growth is limited by N supply, especially in tussock tundra. Because proteins are the predominant form of soil organic N, proteolysis is considered to be the rate-limiting step in both the release of amino acids and in N mineralization. To help understand the controls on N availability in tundra soils, and to determine whether proteolysis is controlled by enzyme activity or by substrate availability, we measured soil protein concentrations, and proteolysis rates with and without added protein, every 1-2 weeks through the summer of 2000 and twice in the summer of 2001. Protease activity with added protein (‘potential’) was higher than without added protein (‘actual’). However, differences between the two tended to be driven by relatively brief peaks in potential protease activity. In fact, actual and potential rates were usually similar, suggesting that much of the time proteolysis was not limited by substrate availability, but rather by enzyme activity. Our data suggest that protease activity was actually only substrate limited at the times when it was highest. Potential rates peaked at the same times that soluble proteins were also high. These increases in protease activity and soluble protein concentrations occurred when soil amino acid and NH₄⁺ concentrations were at their lowest, drawn down by the seasonal peaks in root growth. The fact that the peaks in protease activity coincided with the peak in soil amino acid and NH₄⁺ demand strongly suggests that proteolysis was stimulated by high soil amino acid demand, and resulted in increases in soluble protein concentrations caused by the solubilization of larger proteins.     

Microbial performance under increasing nitrogen availability in a Mediterranean forest soil

In forest ecosystems, the external nitrogen (N) inputs mainly involve wet and dry depositions that potentially alter inorganic N availability in the soil and carbon (C) turnover. This study assesses the effect of a slow increase of inorganic N availability on microbial community activity and functionality in a Mediterranean forest soil. A four-month incubation experiment was performed with soil collected from the organic layer of a forest site and fertilized with a solution of ammonium nitrate. The fertilizer was supplied at an equivalent of 0, 10, 25, 50 and 75 kg N ha⁻¹ (0, 0.3, 0.7, 1.3 and 2 mg N g⁻¹ for control N0 and treatments N1, N2, N3 and N4, respectively). The incubation was carried out under optimal conditions, with the addition of the nutritive solution in small aliquots once a week to mimic the phenomenon of N deposition. In order to isolate the effect of N, the pH of the NH₄NO₃ solutions was adjusted to soil pH, and phosphorus was added in order to prevent any nutrient limitation effect. Inorganic N, C-mineralization, the activity of one oxidative enzyme (o-diphenol oxidase) and 8 hydrolitic enzymes (α-glucosidase, β-glucosidase, N-acetyl-β-d-glucosaminidase, cellulase, leucine amino-peptidase, acid phosphatase, butyric esterase and β-xylosidase) and the community level physiological profile (CLPP) were measured and analyzed during the whole incubation and at the end of the experiment as a proxy for microbial decomposition activity. In the first month, the highest N availability (N4) repressed the microbial respiration activity but stimulated microbial enzymatic activity, suggesting a change of C-pathways from spilling to enzymes and biomass investment. The treatments N1, N2 and N3 had no effect in the same period. Throughout the incubation, a general stress condition affected all the treated soils. As a consequence, treated soils exhibited higher respiration rates than the control. This was accompanied by a loss of functional diversity and an end-detected decline in biomass C. Although at the end of incubation most of the soil features showed a clear correlation with the inorganic N pool, the organic C content was strongly affected by different patterns of microbial activity during the experiment: the highest N treatment (N4) showed a lower C loss than the N3 treatment. Overall, the experiment showed how inorganic N availability can potentially alter the C cycle in a Mediterranean forest soil. The effect is non linear, depending on microbial community dynamics, on the community’s ability to adapt given the time scale of the process, and on N supply amount. Our study also revealed a common pattern in the short-term response to N addition in other, similar ecosystems with different climatic conditions.     

Response of enzyme activities to nitrogen addition in forest floors of different C-to-N ratios

The effect of different inputs of mineral N on several enzyme activities involved in the C and N cycles was investigated using Oa material of forest floors from four Norway spruce [ Picea abies (L.) Karst.] sites with different C-to-N ratios. The samples from each site were treated with five different concentrations of mineral N (as liquid NH ₄NO ₃). All samples were incubated aerobically for 15–20 weeks at 15°C and at field capacity. Respiration was measured weekly. At the end of the incubation period, four enzyme activities (endoglucanase, ß-glucosidase, polyphenol oxidase and ß-glucosaminidase) and microbial biomass were determined. Endoglucanase activity was increased and ß-glucosidase activity was decreased by N additions only in Oa material having a wide C-to-N ratio. In N-supplemented samples of low C-to-N ratio, increased polyphenol oxidase activities were often detected as a consequence of N addition. ß-Glucosaminidase activity responded positively to mineral N additions, particularly in Oa samples with low internal N concentration. The results of the present study indicate that the effects of N additions on enzymatic activities of organic matter in late stages of decomposition are related to the C-to-N ratio. Increasing inputs of mineral N to spruce ecosystems may especially affect C-hydrolyzing enzyme activities in soils with wide C-to-N ratio leading to an incomplete degradation of cellulose and thus reduced C availability to micro-organisms.     

Pore-scale view of microbial turnover: Combining 14C imaging, μCT and zymography after adding soluble carbon to soil pores of specific sizes

The location of microorganisms and substrates within soil pore networks plays a crucial role in organic carbon (C) processing, and its microbial utilization and turnover, and has direct consequences for C and nutrient cycling. An optimal approach to quantify responses to new C inputs from microorganisms residing in specific pores is the addition of new C to pores of target sizes in undisturbed soil cores. We used the matric potential approach to add ¹⁴C-labelled glucose to small (< 40 μm, root free) or large (60–180 μm, potentially inhabited by roots) pores of undisturbed soil cores. Localization of glucose-derived C via ¹⁴C imaging was related to pore size distributions and connectivity, characterized via X-ray computed microtomography (μCT), and to β-glucosidase activity, characterized via zymography. After 2-week incubations, 1.3 times more glucose was mineralized (¹⁴CO₂) when it was added to the large pores; however, more ¹⁴C remained in microbial biomass when glucose was added to the small pores. Consequently, although utilizing the same amounts of easily available C, the microorganisms localized in the large pores had faster turnover compared to microorganisms in small pores. Stronger associations between β-glucosidase activity and glucose-derived C were observed when glucose was added to the large pores. We conclude that (a) the matric potential approach allows placing, albeit not exactly, of soluble substrates into pores of target diameter range, and (b) microorganisms localized in large pores respond to new C inputs with faster turnover, greater growth and more intensive enzyme production compared to those inhabiting the small pores.     

Determining potential glutamine synthetase and glutamate dehydrogenase activity in soil

Ammonium (NH₄⁺), an important nitrogen (N) source for microorganisms, is assimilated via two major pathways. One route is catalyzed by glutamate dehydrogenase (GDH), while the other mechanism involves two enzymes, glutamine synthetase (GS) and glutamate synthase (GOGAT). The GS/GOGAT enzyme system requires more energy to operate, but has a much higher affinity for NH₄⁺ than GDH. We describe procedures to determine potential GS and GDH activity in soil samples. GS and GDH are intracellular enzymes. We used chloroform fumigation to make cell membranes permeable for substrates and products of the enzymes. Fumigation for 4 h increased GS activity almost ten-fold compared to the unfumigated control. Under optimized assay conditions, GS activity increased linearly for at least 80 min, indicating that the substrates were not limiting. In contrast to what was found for GS activity, direct addition of substrates to the soil to assay GDH activity did not result in a linear increase in GDH activity over time. A linear response for 3 h, however, resulted when the soil samples were first extracted with buffer solution and the reagents were added after centrifugation. The differences between the assays explain why fumigation for 3 d prior to the assay increased GDH activity by only 60%. In a microcosm study with glucose and NH₄⁺ addition, the activity of the two enzymes depended on the carbon (C) to N ratio of the amendment. With increasing C to N ratios from 5 to 120, GS activity doubled, while C to N ratios higher than 120 did not further increase GS activity. In contrast, GDH activity decreased by 13% with increasing C to N ratios from 5 to 200. The GDH to GS activity ratio in soil may therefore yield valuable information about the availability of N relative to C at a specific time.     

Formation and remobilisation of soil microbial residue. Effect of clay content and repeated additions of cellulose and sucrose

In two studies, we assessed the mass balance of added ¹⁴C-labelled sucrose and ¹⁵NH₄¹⁵NO₃ by measuring ¹⁴CO₂, ¹⁴C and ¹⁵N in soil microbial biomass (SMB) and ¹⁴C and ¹⁵N in soil solution. Specifically, we assessed the potential of recently added ¹⁴C to be re-mobilised by cryptic growth using subsequent additions of sucrose and cellulose and the effect of physical protection on the stabilisation of the labelled substrate. We used both a constructed soil with low soil organic matter content and varied the clay content as well as a natural soil. We observed a substantial initial as well as a later stage transfer of ¹⁴C into unidentifiable form, hypothesised to be microbial residues. When using a standard k _EC value of 0.45, only roughly 50% of the added labelled substrates were accountable and therefore we explored the full range of reported k _EC values to assess the mass balance. Subsequent application of unlabelled sucrose and cellulose did not substantially increase turnover ¹⁴C and ¹⁵N. Contrary to our expectation, there was no effect of clay content on the amount of unidentified ¹⁴C and ¹⁵N. The unidentified ¹⁴C and ¹⁵N is ascribed to formation of soil microbial residue. The low recovery of added isotope suggests that our mechanistic models are missing a large and important pool in order to realistically simulate organic matter turnover in soil.     

Soil moisture and plant residue addition interact in their effect on extracellular enzyme activity

Water availability strongly affects soil microbial activity and community composition. In a laboratory incubation we investigated the combined effect of soil moisture potential (−10 kPa, −135 kPa, and <−1500 kPa) and plant residue addition on soil enzyme activities (protease, β-glucosidase, β-glucosaminidase and exocellulase) and phospholipid fatty acid (PLFA) profiles. Soil respiration was positively correlated with soil moisture potential and significantly increased with the addition of residue. In the unamended soil, enzyme activities were little affected by soil moisture potential, nor did they change much over time. The addition of residue, however, significantly increased enzyme activity at each moisture level. Furthermore, all four enzyme activities were considerably higher in the amended dry soil than in amended samples with a higher moisture potential. In contrast, in the amended dry soil, respiration and microbial biomass were reduced compared to the amended samples with a higher moisture potential. The low microbial biomass in the amended dry soil was mainly due to a decrease in Gram-negative bacteria, while the fungal biomass reached similar levels at all water potentials. Therefore, shifts in microbial community composition alone cannot explain the increased enzyme activities in the dry soil. Other factors, such as increased fungal activity, stronger interactions between enzymes and soil particles due to thinner water films, may have contributed to the observed effects. Our results suggest that under dry conditions, potential enzyme activities may be decoupled from microbial biomass and respiration in the presence of substrates.     

Influence of organic by-products and nitrogen source on chemical and microbiological status of an agricultural soil

We assessed the influence of the addition of four municipal or agricultural by-products (cotton gin waste, ground newsprint, woodchips, or yard trimmings), combined with two sources of nitrogen (N), [ammonium nitrate (NH₄NO₃) or poultry litter] as carbon (C) sources on active bacterial, active fungal and total microbial biomass, cellulose decomposition, potential net mineralization of soil C and N and soil nutrient status in agricultural soils. Cotton gin waste as a C source promoted the highest potential net N mineralization and N turnover. Municipal or agricultural by-products as C sources had no affect on active bacterial, active fungal or total microbial biomass, C turnover, or the ratio of net C:N mineralized. Organic by-products and N additions to soil did not consistently affect C turnover rates, active bacterial, active fungal or total microbial biomass. After 3, 6 or 9 weeks of laboratory incubation, soil amended with organic by-products plus poultry litter resulted in higher cellulose degradation rates than soil amended with organic by-products plus NH₄NO₃. Cellulose degradation was highest when soil was amended with newsprint plus poultry litter. When soil was amended with organic by-products plus NH₄NO₃, cellulose degradation did not differ from soil amended with only poultry litter or unamended soil. Soil amended with organic by-products had higher concentrations of soil C than soil amended with only poultry litter or unamended soil. Soil amended with organic by-products plus N as poultry litter generally, but not always, had higher extractable P, K, Ca, and Mg concentrations than soil amended with poultry litter or unamende soil. Agricultural soil amended with organic by-products and N had higher extractable N, P, K, Ca and Mg than unamended soil. Since cotton gin waste plus poultry litter resulted in higher cellulose degradation and net N mineralization, its use may result in faster increase in soil nutrient status than the other organic by-products and N sources that were tested. Received: 15 May 1996     

Potential soil enzyme activities are decoupled from microbial activity in dry residue-amended soil

Extracellular enzymes play an important role in the microbial acquisition of carbon (C) and organically bound nutrients, such as nitrogen (N). The objective of the present study was to investigate the effect of different soil moisture contents on potential soil enzyme activities (β-glucosidase and protease), microbial biomass and activity. Soil incubations were carried out with gravimetric moisture contents (GMC) ranging from 0.8 (air-dry) to 30%. After 14 days, respiration, net N mineralization and potential enzyme activities were lowest at GMC below 10% in the unamended samples. In the residue-amended soil, however, respiration and net N mineralization were highest at GMC of 20% or more, while potential β-glucosidase and protease activity were highest at GMC of 10% or less. Increasing the moisture content of air-dry soil after 14 days of incubation resulted in significantly reduced β-glucosidase activity, but increased protease activity. With the exception of the high potential β-glucosidase activity in the residue-amended dry soil, enzyme activities were well correlated with microbial biomass and ergosterol, a biomarker for fungal biomass. Therefore, our results suggest that across the different GMC, protease activity was mainly dependent on the continuous production by microorganisms, while β-glucosidase accumulated in the dry soil due to an increased half-life, which was the result of interactions with soil colloids. Shifts in microbial community composition may also have contributed to the observed differences.     

Biochemical characteristics of solid fractions from animal slurry separation and their effects on C and N mineralisation in soil

Solid fractions from separated animal slurry can be used as organic fertilisers on agricultural land. Solid fractions contain variable amounts of inorganic and organic N, so it is important to synchronise their application in the field with crop demand to ensure N availability in the growing season. This study quantified C and N mineralisation for a wide range of solid fractions from slurry separation applied to soil and examined potential correlations between chemical and biochemical characteristics of solid fractions and their C and N turnover. The solid fractions were mixed with soil and incubated at 14°C for 120 days, during which CO₂ evolution and inorganic N content of the solid fractions were determined. A two-parameter exponential function fitted to the individual solid fraction C mineralisation patterns explained 98% of the data variation, while a three-parameter Monod-type equation fitted to the net N mineralisation patterns explained 89% of the variation. Between 5% and 45% of initially added C was mineralised within the incubation period, with the largest proportion tending to be mineralised from simple mechanically separated solid fractions (MEC). Nitrogen was initially immobilised by the majority of solid fractions. Solid fractions from decanter centrifuged, anaerobically digested slurry (DEC) and chemically pre-treated and separated slurry (KEM) began to re-mineralise N after 20?C40 days, whereas N was continuously immobilised from MEC solid fractions. The carbon mineralisation rate constant was correlated with the C content in the neutral detergent soluble (NDS), hemicellulose and cellulose fractions and the N content in NDS. Net nitrogen mineralisation was correlated with the C/Norg ratio of solid fractions, the N content of NDS and the C content of hemicellulose and cellulose.     

Logging residue harvest may decrease enzymatic activity of boreal forest soils

Nowadays conventional stem-only harvest where logging residues are left on the site is often displaced by whole-tree harvest, in which logging residues are harvested for use as bioenergy. Logging residues consist of tree branches and tops of stems with needles. The aim of this study was to evaluate the effect of logging residue harvest on soil enzyme activities involved in C, N and P cycling, namely β-glucosidase, β-glucosaminidase, protease and acid phosphatase in relation to other soil characteristics (i.e. soil respiration, net N mineralization, microbial biomass C and N). Soil samples were taken from the humus layer of five study sites, differing in fertility, dominating tree species and time elapsed after treatment. The study sites were Norway spruce (Picea abies, (L.) Karst) and Scots pine (Pinus sylvestris L.) stands in different parts of Finland. Four of the study sites were single-tree experiments, where thinning was performed 4–5 years before this study and 3–4 different doses of logging residues (from 0 up to 37.5 Mg ha⁻¹) were distributed on a circle around a single tree in 3 replicates. The last field experiment had been thinned twice, 23 and 13 years ago; the treatments in 3 replicates were whole-tree harvest and stem-only harvest. In the whole-tree harvest vs. stem-only harvest experiment, activities of β-glucosidase, β-glucosaminidase, acid phosphatase were similar in both treatments. In general, in the single-tree experiment with pine, enzymes raised the activity in response to increasing amount of logging residue. The pattern was less clear for the spruce single-tree experiment, but acid phosphatase and protease activities increased with the increase in amount of logging residue. In general, other soil characteristics were less affected than enzyme activities by logging residue removal; however, in some sites logging residues seemed to increase net C and N mineralization with increasing logging residue amount. Our results suggest that retaining logging residues on the site can increase soil enzyme activities and C and N mineralization.     

Long-term application of compost influences microbial biomass and enzyme activities in a tropical Aeric Endoaquept planted to rice under flooded condition

In a field study, long-term application of compost to a tropical Aeric Endoaquept under continuous rice growing in a rice-rice-fallow sequence resulted in the stimulation of microbial biomass and select soil enzyme activities. Mean seasonal soil microbial biomass-C (C_mic) increased by 42%, 39% and 89% in inorganic fertilizer, compost and compost+inorganic fertilizer treatments, respectively, over the unamended control. C_mic content was also influenced by the rice crop growth stage and was highest at maximum tillering stage irrespective of treatments and declined thereafter. Soil organic C (C_org) content showed highly significant positive correlation with dehydrogenase, urease, cellulase, β-glucosidase and fluorescein di-acetate (FDA) hydrolysis activity, and a positive but not significant correlation with invertase and amidase activity. C/N ratio which was lowest in unamended control plots showed a significant positive relationship with only the enzymes involved in C cycle. Stepwise regression analysis revealed that for prediction of both total organic C and total N, FDA hydrolysis activity contributed significantly for the variance and explained up to 85-96% variability. Results demonstrated that microbial biomass and soil enzyme activity is sensitive in discriminating between long-term organic residue amendment practices.     

Microbial respiration and growth during the decomposition of wheat straw

The effect of N on the disappearance of C from 1.5 g wheat (Triticum aestivum L. var. “Nugaines”) straw decomposing in sand and the response of the biomass to addition of N (adequate to bring the C:N ratio to 48:1) and C (200 mg of glucose-C) were determined. A concept was used that assumed the change in the microbial biomass was proportional to the change in acid hydrolyzable amino acid-N. Microbial respiration (CO₂ evolution and O₂ uptake) and growth were stimulated by the initial addition of N (which brought the original C:N ratio from 150:1 to 48:1), but the addition of the same amount of N to the system at 240 h that had received no N initially resulted in slight if any increase in respiration or microbial growth. The response of the microflora to the 200 mg of glucose-C additions after 240 h indicated the microbial populations were primarily limited by available C rather than available N after only 240 h incubation, even though about 95% of the original straw residue-C plus biomass C remained in the system. Respiratory quotients indicated a qualitative shift over time in the average oxidation state of the substrates being metabolized. It is postulated that the RQ shift resulted, at least in part, from death of the population rather than totally from the availability of the straw substrate.The initial addition of N resulted in 3.8 times the net amino acid production, but only 1.6 times the CO₂-C production over 240 h compared with the control without added N. These results suggest that N availability might result in a change in the growth yield of the microbial population.     

水分含量对秸秆还田土壤碳矿化和微生物特性的影响

An 80-d incubation experiment was conducted to investigate straw decomposition,the priming effect and microbial characteristics in a non-fertilized soil(soil 1) and a long-term organic manure-fertilized soil(soil 2) with and without13 C-labeled maize straw amendment under different moisture levels. The soil 2 showed a markedly higher priming effect,microbial biomass C(Cmic),and β-glucosidase activity,and more abundant populations of bacteria and fungi than the soil 1. Also,soil CO2 emission,Cmic,β-glucosidase activity,and bacterial and fungal population sizes were substantially enhanced by straw amendment. In the presence of straw,the amount of straw mineralization and assimilation by microbes in the soil at 55% of water holding capacity(WHC) were significantly higher by 31% and 17%,respectively,compared to those at 25% of WHC. In contrast,β-glucosidase activity and fungal population size were both enhanced as the moisture content decreased. Cmicdecreased as straw availability decreased,which was mainly attributed to the reduction of straw-derived Cmic. Amended soils,except the amended soil 2 at 25% of WHC,had a more abundant fungal population as straw availability decreased,indicating that fungal decomposability of added straw was independent of straw availability. Non-metric multidimensional scaling analysis based on fungal denatured gradient gel electrophoresis band patterns showed that shifts in the fungal community structure occurred as water and straw availability varied. The results indirectly suggest that soil fungi are able to adjust their degradation activity to water and straw availability by regulating their community structure.     

Microbial biomass and mineralization-immobilization of nitrogen in some agricultural soils

Summary The chloroform fumigation-incubation method (CFIM) was used to measure the microbial biomass of 17 agricultural soils from Punjab Pakistan which represented different agricultural soil series. The biomass C was used to calculate biomass N and the changes occurring in NH₄ ⁺-N and NO₃ ^–-N content of soils were studied during the turnover of microbial biomass or added C source. Mineral N released in fumigated-incubated soils and biomass N calculated from biomass C was correlated with some N availability indexes.The soils contained 427–1240 kg C as biomass which represented 1.2%–6.9% of the total organic C in the soils studied. Calculations based on biomass C showed that the soils contained 64–186 kg N ha^–1 as microbial biomass. Immobilization of NCO₃ ^–-N was observed in different soils during the turnover of microbial biomass and any net increase in mineral N content of fumigated incubated soils was attributed entirely to NH₄ ⁺-N.Biomass N calculated from biomass C showed non-significant correlation with different N availability indexes whereas mineral N accumulated in fumigated-incubated soils showed highly significant correlations with other indexes including N uptake by plants.     

Significance of organic nitrogen uptake from plant residues by soil microorganisms as affected by carbon and nitrogen availability

Soil microorganisms can use a wide range of nitrogen (N) compounds. When organic N sources are degraded, microorganisms can either take up simple organic molecules directly (direct route), or organic N may be mineralized first and taken up in the form of mineral N (mineralization-immobilization-turnover [MIT] route). To determine the importance of the direct route, a microcosm experiment was carried out. Two types of wheat residue were added to soil samples, including younger residue with a carbon (C) to N ratio of 12 and older residue with a C to N ratio of 29. Between days 1 and 4, the gross N mineralization rate reached 8.4 and 4.0 mg N kg⁻¹ dry soil day⁻¹ in the treatment with younger and older residue, respectively. During the same period, there was no difference in protease activity between the two residue amended treatments. The fact that protease activity was not related to gross N mineralization, even though the products of protease activity are the substrates for N mineralization, suggests that not all organic molecules released from residue or soil N passed through the soil mineral N pool. In fact, when leucine and glycine were added, only 10 and 53% of the amino acid-N, respectively, was mineralized. The fraction of N taken up via the direct route was estimated to be 55 and 62% for the young and older residue, respectively. After 28 days of incubation, the proportion of amino acid-N mineralized had increased especially in the soil amended with older residue, suggesting that the MIT route became increasingly important. This result is supported by an increase in the activities of enzymes responsible for the intracellular assimilation of ammonium (NH₄⁺). Our results suggest that in contrast to what is proposed by many models of soil N cycling, both the direct and MIT routes were operative, with the direct route being the preferred route of residue N uptake. The direct route became less important over time and was more important in soil amended with older residue, suggesting that the direct route is favored by lower mineral N availabilities. An important implication of these findings is that when the direct route is dominant, gross N mineralization underestimates the amount of N made available from the residue.     

Carbon mineralization is promoted by phosphorus and reduced by nitrogen addition in the organic horizon of northern hardwood forests

Limitations to the respiratory activity of heterotrophic soil microorganisms exert important controls of CO₂ efflux from soils. In the northeastern US, ecosystem nutrient status varies across the landscape and changes with forest succession following disturbance, likely impacting soil microbial processes regulating the transformation and emission of carbon (C). We tested whether nitrogen (N) or phosphorus (P) limit the mineralization of soil organic C (SOC) or that of added C sources in the Oe horizon of successional and mature northern hardwood forests in three locations in central New Hampshire, USA. Added N reduced mineralization of C from SOC and from added leaf litter and cellulose. Added P did not affect mineralization from SOC; however, it did enhance mineralization of litter- and cellulose- C in organic horizons from all forest locations. Added N increased microbial biomass N and K₂SO₄-extractable DON pools, but added P had no effect. Microbial biomass C increased with litter addition but did not respond to either nutrient. The direction of responses to added nutrients was consistent among sites and between forest ages. We conclude that in these organic horizons limitation by N promotes mineralization of C from SOC, whereas limitation by P constrains mineralization of C from new organic inputs. We also suggest that N suppresses respiration in these organic horizons either by relieving the N limitation of microbial biomass synthesis, or by slowing turnover of C through the microbial pool; concurrent measures of microbial growth and turnover are needed to resolve this question.     

Soil biochemical and microbial indices in wet tropical forests: Effects of deforestation and cultivation

Little information is available about the long‐term effects of deforestation and cultivation on biochemical and microbial properties in wet tropical forest soils. In this study, we evaluated the general and specific biochemical properties of soils under evergreen, semi‐evergreen, and moist deciduous forests and adjacent plantations of coconut, arecanut, and rubber, established by clear felling portions of these forests. We also examined the effects of change in land use on microbial indices and their interrelationships in soils. Significant differences between the sites occurred for the biochemical properties reflecting soil microbial activity. Microbial biomass C, biomass N, soil respiration, N mineralization capacity, ergosterol, levels of adenylates (ATP, AMP, ADP), and activities of dehydrogenase and catalase were, in general, significantly higher under the forests than under the plantations. Likewise, the activities of various hydrolytic enzymes such as acid phosphomonoesterase, phosphodiesterase, casein‐protease, BAA‐protease, β‐glucosidase, CM‐cellulase, invertase, urease, and arylsulfatase were significantly higher in the forest soils which suggested that deforestation and cultivation markedly reduced microbial activity, enzyme synthesis and accumulation due to decreased C turnover and nutrient availability. While the ratios of microbial biomass C : N and microbial biomass C : organic C did not vary significantly between the sites, the ratios of ergosterol : biomass C and ATP : biomass C, qCO2 and AEC (Adenylate Energy Charge) levels were significantly higher in the forest sites indicating high energy requirements of soil microbes at these sites.     

Responses of soil microbial communities and functions associated with organic carbon mineralization to nitrogen addition in a Tibetan grassland

Alpine grasslands with a high soil organic carbon(SOC)storage on the Tibetan Plateau are experiencing rapid climate warming and anthropogenic nitrogen(N)deposition;this is expected to substantially increase the soil N availability,which may impact carbon(C)cycling.However,little is known regarding how N enrichment influences soil microbial communities and functions relative to C cycling in this region.We conducted a 4-year field experiment on an alpine grassland to evaluate the effects of four different rates of N addition(0,25,50,and 100 kg N ha^-1 year^-1)on the abundance and community structure(phospholipid fatty acids,PLFAs)of microbes,enzyme activities,and community level physiological profiles(CLPP)in soil.We found that N addition increased the microbial biomass C(MBC)and N(MBN),along with an increased abundance of bacterial PLFAs,especially Gram-negative bacterial PLFAs,with a decreasing ratio of Gram-positive to Gram-negative bacteria.The N addition also stimulated the growth of fungi,especially arbuscular mycorrhizal fungi,reducing the ratio of fungi to bacteria.Microbial functional diversity and activity of enzymes involved in C cycling(β-1,4-glucosidase and phenol oxidase)and N cycling(β-1,4-N-acetyl-glucosaminidase and leucine aminopeptidase)increased after N addition,resulting in a loss of SOC.A meta-analysis showed that the soil C/N ratio was a key factor in the response of oxidase activity to N amendment,suggesting that the responses of soil microbial functions,which are linked to C turnover relative to N input,primarily depended upon the soil C/N ratio.Overall,our findings highlight that N addition has a positive influence on microbial communities and their associated functions,which may reduce soil C storage in alpine grasslands under global change scenarios.