Ectomycorrhizal community and extracellular enzyme activity following simulated atmospheric N deposition

Ectomycorrhizal (EM) fungi are abundant in temperate and boreal ecosystems and are understood to be an important means whereby plants can fulfill their nutrition requirements. The extent of the EM fungal involvement in accessing organic sources of N, however, remains unknown. Some EM fungi have been found to produce lignolytic and proteolytic enzymes which are necessary to depolymerize organic substrates, but this ability varies by species. Both EM fungal communities and the activities of lignolytic and proteolytic enzymes may be sensitive to changes in inorganic N availability such as through increased atmospheric deposition. Our objectives were to simulate an ecologically relevant increase in atmospheric N deposition in areas currently receiving very little exogenous N and examine changes in EM community composition, lignin degrading enzyme activity, and soil protein depolymerization. We found a distinct shift in the EM community composition following simulated atmospheric N deposition. Likewise, we found a significant decrease in the activity of lignin degrading enzymes, which could have important implications on ecosystem N and C cycling. Contrary to our hypotheses, proteolysis increased following N addition. The fact that lignolytic and proteolytic enzymes exhibit opposite responses is counterintuitive and suggests much is yet to be learned about how N addition affects global C storage by affecting the decomposition of organic matter. Our data suggest small increases in atmospheric N deposition could produce significant changes in communities of EM fungi and N and C cycles.     

Soil amino acid composition across a boreal forest successional sequence

Soil amino acids are important sources of organic nitrogen for plant nutrition, yet few studies have examined which amino acids are most prevalent in the soil. In this study, we examined the composition, concentration, and seasonal patterns of soil amino acids across a primary successional sequence encompassing a natural gradient of plant productivity and soil physicochemical characteristics. Soil was collected from five stages (willow, alder, balsam poplar, white spruce, and black spruce) of the floodplain successional sequence on the Tanana River in interior Alaska. Water-extractable amino acid composition and concentration were determined by HPLC. Irrespective of successional stage, the amino acid pool was dominated by glutamic acid, glutamine, aspartic acid, asparagine, alanine, and histidine. These six amino acids accounted for approximately 80% of the total amino acid pool. Amino acid concentrations were an order of magnitude higher in coniferous-dominated late successional stages than in early deciduous-dominated stages. The composition and concentration of amino acids were generally constant throughout the growing season. The similar amino acid composition across the successional sequence suggests that amino acids originate from a common source or through similar biochemical processes. These results demonstrate that amino acids are important components of the biogeochemical diversity of nitrogen forms in boreal forests.     

A liquid chromatographic/mass spectrometric method to evaluate <Superscript>13</Superscript>C and <Superscript>15</Superscript>N incorporation into soil amino acids

Purpose  

Amino acids are highly associated with biogeochemical cycling and represent an important potential source and sink of carbon (C) and nitrogen (N) in terrestrial ecosystems. Tracing the isotope dynamics of amino acids can improve the understanding of the origin and transformation of amino acids in soil matrix at process-levels; hence, the liquid chromatographic/mass spectrometric (LC/MS) method to evaluate ¹³C or ¹⁵N enrichment in amino acids is necessary to be established.     

Control of amino acid mineralization and microbial metabolism by temperature

Amino acids constitute a major reserve of soil organic-N and studies demonstrating direct uptake of amino acids by plants has indicated that understanding their bioavailability and fate in soil is important to understanding terrestrial N cycling. The aim of this study was to determine the effects of temperature and sorption on the mineralisation of three amino acids (glycine, lysine glutamate) in soil. Amino acid sorption followed the series lysine>glycine>glutamate, whereas mineralisation rate followed the series glutamate>glycine>lysine. These observations support the concept that sorption reduces the bioavailability of amino acids to the soil microbial population. Although the amino acids were used preferentially for making new biomass rather than respiration, differences were apparent between the individual amino acids with microbial assimilation efficiency (biomass production) following the series, lysine>glycine>glutamate. Our results suggest divergences in the uptake and metabolism of the individual amino acids with a rapid mineralisation of amino acids which readily enter general metabolic cycles (e.g. glutamate) compared to the amino acids which typically form the terminus of metabolic pathways (e.g. lysine). Temperature significantly affected the rate of amino acid mineralisation which increased up to 30°C (Q₁₀=2.0) followed by a decline as the temperature approached 40°C. Rapid mineralisation occurred even at very low temperatures (1°C). Amino acid mineralisation across three experimental soil treatments followed the trend acidified>control>eroded soil. In summary, the results indicate that mineralisation is highly amino acid species dependent, has a mesophilic optimum, is retarded by sorption and is most rapid in soils which are not degraded.     

Effects of amino-acid chemistry and soil properties on the behavior of free amino acids in acidic forest soils

Free amino acids (FAAs) in soil solution are increasingly recognized as a potentially important source of nitrogen (N) for plants, yet we are just beginning to understand the behavior of FAAs in soil. I investigated the effects of amino-acid chemistry and soil properties on mineralization, microbial assimilation and sorption of amino-acid N in soils from three ecosystems representing the two endpoints and mid point of a temperate forest fertility gradient ranging from low mineral N availability/high FAA oak forests to high mineral N availability/low FAA maple-basswood forests. Soils were amended with six ¹⁵N-labeled amino-acid substrates that ranged widely in chemical properties, including molecular weight, C:N ratio, average net charge, hydrophobicity, and polarity: Arginine (Arg), Glutamine (Gln), Glutamate (Glu), Serine (Ser), Glycine (Gly) and Leucine (Leu). Mineralization of amino-acid N accounted for 7-45% (18% avg.) of the added label and was most strongly affected by soil characteristics, with mineralization increasing with increasing soil fertility. Mineralization of amino-acid N was unrelated to amino-acid C:N ratio, rather, I observed greater N mineralization from polar FAAs compared to non-polar ones. Assimilation of amino-acid N into microbial biomass accounted for 6-48% (29% avg.) of the added label, and was poorly predicted by either intrinsic amino-acid properties or soil properties, but instead appeared to be explicable in terms of compound-specific demand by soil micoorganisms. Sorption of amino-acid N to soil solids accounted for 4-15% (7% avg.) of the added label and was largely controlled by charge characteristics of individual amino acids. The fact that both positively- and negatively-charged amino acids were more strongly sorbed than neutral ones suggests that cation and anion exchange sites are an important factor controlling sorption of FAAs in these acid forest soils. Together, the findings from this study suggest that there may be important differences in the behavior of free amino acids in sandy, acidic forest soils compared to generalizations drawn from finer-textured grassland soils, which, in turn, might affect the availability of some FAAs in soil solution.     

Amino Acids, Total Organic and Inorganic Nitrogen in Forest Floor Soil Solution at Low and High Nitrogen Input

It is now widely accepted that many plants and mycorrhizal fungi have the ability to take up organic nitrogen (N) in the form of amino acids, although the importance of this uptake in the field is less clear. In the laboratory it has been shown that uptake affinity and uptake kinetics of ammonium and some amino acids are comparable. The relative uptake of either N form from the soil solution would thus be related to the relative concentration in the soil solution accessed by roots. We sampled soil solution from the F- and H-layers under a Spruce stand in a fertilisation experiment in Flakaliden, northern Sweden. Tension lysimeters were installed in plots receiving irrigation (I) or irrigation plus liquid fertilisation (IL). The soil solution samples were analysed for ammonium, nitrate, free amino acids, hydrolysable amino acids, total organic N and total organic C. In I plots the concentrations of both ammonium and free amino acids were very low with no obvious dominance of either form. In IL plots inorganic N concentrations were higher and amino acid concentrations were lower compared to I plots, and thus the inorganic N dominated over amino acids. There was no difference in H-layer ammonium concentration between I and IL plots despite the high N addition rate on the soil surface during nights of sampling. The lower amino acid concentrations in IL plots might be an effect of a decreased proteolytic activity due to the documented shift in mycorrhizal fungi species composition at the site.     

Protein breakdown represents a major bottleneck in nitrogen cycling in grassland soils

Proteins represent the dominant input of organic N into most ecosystems and they also constitute the largest store of N in soil organic matter. The extracellular protease mediated breakdown of proteins to amino acids therefore represents a key step regulating N cycling in soil. In this study we investigated the influence of a range of environmental factors on the rate of protein mineralization in a grazed grassland and fallow agricultural soil. The protein turnover rates were directly compared to the rates of amino acid mineralization under the same conditions. Uniformly ¹⁴C-labelled soluble protein and amino acids were added to soil and the rate of ¹⁴CO₂ evolution determined over 30 d. Our results indicate that the primary phase of protein mineralization was approximately 20 ± 3 fold slower that the rate of amino acid mineralization. The addition of large amounts of inorganic NO₃⁻ and NH₄⁺ to the soil did not repress the rate of protein mineralization suggesting that available N does not directly affect protease activity in the short term. Whilst protein mineralization was strongly temperature sensitive, the presence of plants and the addition of humic and tannic acids had relatively little influence on the rate of soluble protein degradation in this fertile grassland soil. Our results suggests that the extracellular protease mediated cleavage of proteins to amino acids rather than breakdown of amino acids to NH₄⁺ represents the limiting step in soil N cycling.     

Inter‐specific variability in organic nitrogen uptake of three temperate grassland species

We tested the inter‐specific variability in the ability of three dominant grasses of temperate grasslands to take up organic nitrogen (N) in the form of amino acids in soils of differing fertility. Amino acid uptake was determined by injecting dual labeled glycine‐2‐¹³C‐¹⁵N into the soil, and then measuring the enrichment of both ¹³C and ¹⁵N in plant tissue after 50 hours. We found enrichment of both ¹³C and ¹⁵N in root and shoot material of all species in both soils, providing first evidence for direct uptake of glycine. We show that there was considerable inter‐specific variability in amino acid uptake in the low fertility soil. Here, direct uptake of amino acid was greater in the grass Agrostis capillaris, which typically dominates low fertility grassland, than Lolium perenne, which inhabits more fertile sites. Direct uptake of amino acid for Holcus lanatus. was intermediate between the above two species. Unlike in the low fertility soil, there was no difference in uptake of either ¹³C or ¹⁵N by grasses in the high fertility soil, where uptake of mineral N is thought to be the major mechanism of N uptake of these grasses. Overall, our findings may contribute to our understanding of differences in competitive interactions between grasses in soils of different fertility status.     

Controls over mycorrhizal uptake of organic nitrogen

Mycorrhizal plants from a variety of ecosystems have the capacity to take up organic forms of nitrogen, yet the fraction of plant nitrogen demand met by organic N (ON) uptake remains unclear. ON uptake by mycorrhizal plants is a biochemical process that involves multiple steps, including breakdown and uptake of soil ON by mycorrhizal fungi, internal transformation of ON, and transfer of N to the host plant. We present hypothetical mechanisms controlling each of these steps and outline predictions for how these mechanisms structure patterns of ON uptake by mycorrhizal plants in ecosystems. Using a synthesis of published data, we found that uptake of amino acids by mycorrhizal fungi is related to the relative abundance, N content, and carbon structure of the amino acid. We hypothesize that the bond strength and structural diversity of soil ON controls the breakdown of polymeric ON by mycorrhizal fungi. In addition, the availability of carbon resources for the mycorrhizal fungus influences the capacity for mycorrhizal fungi to assimilate amino acids and produce extracellular enzymes that catalyze the breakdown of polymeric ON.     

Competition for amino acids between wheat roots and rhizosphere microorganisms and the role of amino acids in plant N acquisition

The direct uptake of organic nitrogen compounds from the soil solution by plant roots has been hypothesised to constitute a significant source of N to the plant particularly in N limiting ecosystems. The experiments undertaken here were designed to test whether wheat roots could out-compete the rhizosphere microflora for a pulse addition of organic N in the form of three contrasting amino acids, namely lysine, glycine and glutamate. Amino acids were added at a concentration reflecting reported soil solution concentrations (100 μM) and the uptake into either plant biomass or respiration or microbial biomass and respiration determined over a 24 h chase period. The results showed that the plant roots could only capture on average 6% of the added amino acid with the remainder captured by the microbial biomass. We therefore present direct in vivo evidence to support earlier work which has hypothesised that organic N may be of only limited consequence in high input agricultural systems. We suggest that this is a result of the higher concentrations of NO₃⁻ in agricultural soil solutions, the slow movement of amino acids in soil relative to NO₃⁻, the rapid turnover of amino acids by soil microorganisms, and the poor competitive ability of plant roots to capture amino acids from the soil solution.     

Soil- and enantiomer-specific metabolism of amino acids and their peptides by Antarctic soil microorganisms

Most nitrogen (N) enters many Arctic and Antarctic soil ecosystems as protein. Soils in these polar environments frequently contain large stocks of proteinaceous organic matter, which has decomposed slowly due to low temperatures. In addition to proteins, considerable quantities of d-amino acids and their peptides enter soil from bacteria and lengthy residence times can lead to racemisation of l-amino acids in stored proteins. It has been predicted that climate warming in polar environments will lead to increased rates of soil organic N turnover (i.e. amino acids and peptides of both enantiomers). However, our understanding of organic N breakdown in these soils is very limited. To address this, we tested the influence of chain length and enantiomeric composition on the rate of breakdown of amino acids and peptides in three contrasting tundra soils formed under the grass, moss or lichen-dominated primary producer communities of Signy Island in the South Orkney Islands. Both d- and l-enantiomers of the amino acid monomer were rapidly mineralized to CO₂ at rates in line with those found for l-amino acids in many other terrestrial ecosystems. In all three soils, l-peptides were decomposed faster than their amino acid monomer, suggesting a different route of microbial assimilation and catabolism. d-peptides followed the same mineralization pattern as l-peptides in the two contrasting soils under grass and lichens, but underwent relatively slow decomposition in the soil underneath moss, which was similar to the soil under the grass. We conclude that the decomposition of peptides of l-amino acids may be widely conserved amongst soil microorganisms, whereas the decomposition of peptides of d-amino acids may be altered by subtle differences between soils. We further conclude that intense competition exists in soil microbial communities for the capture of both peptides and amino acids produced from protein breakdown.     

Soil nitrogen pools and turnover in native woodland and managed pasture soils

Dissolved organic nitrogen (DON) is a significant nitrogen (N) pool in most soils and is considered to be important for N cycling. The present study focused on paired sites of native remnant woodland and managed pasture at three locations in south-eastern Australia. Improved understanding of N cycling is important for assessing the impact of agriculture on soil processes and can guide conservation and restoration soil management strategies to maintain remnant native woodland systems, which currently exist as small pockets of woodland within extensive managed pasture landscapes. Organic and inorganic N pools were quantified, as well as the rates of amino acid and peptide mineralisation in the paired native woodland and managed pasture systems. Soil DON dominated the soil N pool in both land uses, and the proportion of DON to other N pools was greatest at the most N-limited site (up to ∼70% of extractable N). In both land uses soil ammonium and free amino acid concentrations were similar (∼20% of extractable N), and soil nitrate formed the smallest N pool (<∼5% of extractable N). Mineralisation of ¹⁴C-labelled amino acid and peptide substrates was rapid (<3 h), and more amino acid was respired than peptide in both the native woodland and managed pasture soils. Soil C:N ratio was important in separating site and land use differences, and contrasting relationships between soil physico-chemical properties and organic N uptake rates were identified across sites and land uses.     

Dissolved organic carbon and nitrogen dynamics in temperate coniferous forest plantations

Dissolved organic matter (DOM) plays a central role in driving many chemical and biological processes in soil; however, our understanding of the fluxes and composition of the DOM pool still remains unclear. In this study we investigated the composition and dynamics of dissolved organic carbon (DOC) and nitrogen (DON) in five temperate coniferous forests. We subsequently related our findings to the inputs (litterfall, throughfall, atmospheric deposition) and outputs (leaching, respiration) of C and N from the forest and to plant available sources of N. With the exception of NO₃^?, most of the measured soil solution components (e.g. DOC, DON, NH₄⁺, free amino acids, total phenolics and proteins) progressively declined in concentration with soil depth, particularly in the organic horizons. This decline correlated well with total microbial activity within the soil profile. We calculated that the amount of C lost by soil respiration each day was equivalent to 70% of the DOC pool and 0.06% of the total soil C. The rapid rate of amino acid mineralization and the domination of the low molecular weight soluble N pool by inorganic N suggest that the microbial community is C‐ rather than N‐limited and that C‐limitation increases with soil depth. Further, our results suggest that the forest stands were not N‐limited and were probably more reliant on inorganic N as a primary N source rather than DON. In conclusion, our results show that the size of the DON and DOC pools are small relative to both the amount of C and N passing through the soil each year and the total C and N present in the soil. In addition, high rates of atmospheric N deposition in these forests may have removed competition for N resources between the plant and microbial communities.     

Analysis and behavior of soluble organic nitrogen in forest soils

Background, aim, and scope  

A large proportion of soil nitrogen (N; >80%) is present in organic form. Current research on plant N uptake in terrestrial ecosystems has focused mainly on inorganic N such as ammonium (NH₄ ⁺) and nitrate (NO₃ ⁻), while soluble organic N (SON) has received little attention. In recent years, the increasing evidence showing the direct uptake of various amino acids by plants and the predominance of the organic form in N loss by leaching in many forest ecosystems has drawn attention to critically re-examine the nature and the ecological role of soil SON in terrestrial N cycling. However, little is known about the sources and dynamics, chemical nature, and ecological functions of soil SON in forest ecosystems. This paper reviews recent advances in the areas of research on current techniques for characterizing soil SON and the size, nature, and dynamics of soil SON pools in forest ecosystems.     

Kinetics of soil microbial uptake of free amino acids

 Amino acids and proteins typically form the biggest input of organic-N into most soils and provide a readily available source of C and N for soil microorganisms. Amino acids can also be taken up directly by plant roots, providing an alternative source of available soil N. However, the degree to which plants can compete against the soil microbial population for amino acids in soil solution remains poorly understood. The aim of this study was to measure the rate of microbial uptake of three contrastingly charged ¹⁴C-labelled amino acids (glutamate^1–, glycine⁰, lysine^0.9+) over a wide concentration range (0.1–5 mM) and in two contrastingly managed soils varying in their degree of erosion, organic-C content and microbial biomass. Amino acid uptake was concentration dependent and conformed to a single Michaelis-Menten equation. The mean maximum amino acid uptake rate (V _max) for the non-eroded (control) soil (high organic-C, high biomass) was 0.13±0.02 mmol kg^–1 h^–1, while half maximal uptake occurred at a concentration (K _m) of 2.63±0.07 mM. Typically, V _max was fourfold lower and K _m twofold lower in the eroded soil (low available organic-C, low biomass) compared to the non-eroded (control) soil. Amino acid substrate concentration had little effect on the proportion of amino acid utilized in catabolic versus anabolic metabolism and was similar for both. While the results obtained here represent the summation of kinetics for a mixed soil population, they indicate that amino acid uptake is saturated at concentrations within the millimolar range. Because the affinity constants also were similar to those described for plant roots, we hypothesized that competition for amino acids between plants and microbes will be strong in soil but highly dependent upon the spatial distribution of roots and microbes in soil. Received: 2 March 2000     

Interference by amino acids during the determination of N ammonium in soil

In the study of terrestrial N cycling, NH₄⁺ concentration and ¹⁵N enrichment are routinely determined by colorimetric continuous flow analysis and microdiffusion methods. Amino acids can interfere in these determinations; consequently the aim of the present study was to evaluate the significance of the interference. Glycine and glutamine are key amino acids in soil and were therefore used as ‘models’. Both glycine and glutamine interfered during continuous flow analysis, whereas interference during microdiffusion was of little importance. The effects of interference can be significant, e.g. estimates of gross mineralisation rate were reduced up to 33%, where we allowed for amino acid interference during determination of NH₄⁺ concentration. The potential influence of amino acid interference emphasises that development of continuous flow analysis to increase NH₄⁺ specificity is needed.     

Amino acids and amino sugars extracted by EUF from a sandy soil incubated with green manure,bacterial biomass or cellulose

The extraction of soils by the electro-ultrafiltration (EUF) method yields organic N which has been used as an index for mineralisable N in soils. This EUF extractable organic fraction contains a mixture of various N compounds not yet completely identified. It has been proposed that the amino N compounds are more indicative for the potentially mineralisable N in soils than the total organic N extracted (Mengel et al., 1999). An amendment of soils with easily mineralisable organic matter may, therefore, alter the amino N concentrations of the organic N extracted. Our determination of the amino N compounds aimed to prove this hypothesis. The principle of our experiment was to mix soil with green manure, bacterial biomass and cellulose, respectively, and to incubate the treated soil aerobically for 80 days at 20°C in the laboratory. Control treatments without organic amendment were also incubated. Soil samples were taken several times during the incubation period and analysed for the inorganic N (NO₃⁻-N and NH₄⁺-N) and for the EUF extractable organic N. Amino acids and amino sugars were determined in the hydrolysed EUF extracts. The concentrations of amino acids and amino sugars in the organic N extracted varied with time and differed between the treatments. Glutamic acid has been found to be the most relevant amino acid in the EUF extracts and was particularly indicative for the existence of mineralisable green manure in the soil. Glucosamine was the most relevant amino sugar in the EUF extracts and this amino sugar appears to be indicative for the easily mineralisable relics of microbial cells in the soil.     

Can ectomycorrhizal fungi circumvent the nitrogen mineralization for plant nutrition in temperate forest ecosystems?

Nitrogen (N) limits plant growth in many forest ecosystems. The largest N pool in the plant-soil system is typically organic, contained primarily within the living plants and in the humus and litter layers of the soil. Understanding the pathways by which plants obtain N is a priority for clarifying N cycling processes in forest ecosystems. In this review, the interactions between saprotrophic microorganisms and ectomycorrhizal fungi in N nutrition with a focus on the ability of ectomycorrhizal fungi to circumvent N mineralization for the nutrition of plants in forest ecosystems will be discussed. Traditionally, it is believed that in order for plants to fulfill their N requirements, they primarily utilize ammonium (NH₄⁺) and nitrate (NO₃⁻). In temperate forest ecosystems, many woody plants form ectomycorrhizas which significantly improves phosphorus (P) and N acquisition by plants. Under laboratory conditions, ectomycorrhizal fungi have also been proven to be able to obtain N from organic sources such as protein. It was thus proposed that ectomycorrhizal fungi potentially circumvent the standard N cycle involving N mineralization by saprotrophic microorganisms. However, in many forest ecosystems the majority of the proteins in the forest floor form complexes with polyphenols. Direct access of N by ectomycorrhizal fungi from a polyphenol-protein complex may be limited. Ectomycorrhizal fungi may depend on saprotrophic microorganisms to liberate organic N sources from polyphenol complexes. Thus, interactions between saprotrophic microorganisms and ectomycorrhizal fungi are likely to be essential in the cycling of N within temperate forest ecosystems.     

Dynamics of soil carbon to nitrogen ratio changes under long‐term fertilizer addition in wheat‐corn double cropping systems of China

Soil carbon to nitrogen (C:N) ratio is one of the important properties of terrestrial ecosystems. Here, we report a study of soil C:N ratio dynamics in wheat‐corn double cropping systems based on four long‐term experimental sites in China: three in the temperate zone and one in the sub‐tropical zone. We evaluate effects of long‐term fertilizer input on soil organic carbon (SOC) and total nitrogen (TN) by comparing three treatments: no added fertilizer (the control), added nitrogen‐phosphorus‐potassium chemical fertilizers (NPK), and chemical fertilizers combined with manure (NPKM). Our study shows that SOC and TN had different responses to the treatments. There was an increasing trend in SOC, even without fertilizer. However, applying inorganic fertilizers only (NPK) did not maintain TN contents at some sites. The NPKM treatment resulted in a large increase in both SOC (35–147%) and TN (33 to 10%) contents, relative to the initial values. The soil C:N ratio showed a significant increase over time at the sub‐tropical site but little change at the three temperate sites. Our analysis showed similar C:N ratios (37–38) in gross input of organic materials under the NPK treatments. However, the estimated C:N ratio during decomposition was much smaller at the sub‐tropical site (23.7) than at the three temperate sites (44.0–48.2) under the NPK treatments, which may explain the increased soil C:N ratio at the sub‐tropical site. Thus, we conclude that variations in soil C:N ratio are not caused by organic matter inputs but by decomposition in the wheat‐corn double cropping systems.     

Nitrogen deposition and dissolved organic carbon production in northern temperate forests

Deposition of anthropogenic nitrogen (N) alters the decomposition of organic matter in forest ecosystems by changing the expression of key microbial enzymes. We investigated the effects of experimental N deposition on dissolved organic matter (DOM) in soils of three forest ecosystems representative of the upper Great Lakes region: the sugar maple/basswood (SMBW), sugar maple/red oak (SMRO) and white oak/black oak (WOBO) ecosystems. Mineral soil samples were collected on five dates from ambient and N-amended plots (80 kg N ha⁻¹ yr⁻¹) in three replicate stands of each forest type. DOM was extracted (2:1, water:soil) from each soil sample and analyzed for dissolved organic carbon (DOC). DOC concentration was significantly greater in the N-amended soils (on average: 24% higher for SMBW, 9% for SMRO, and 40% for BOWO). In June and October 2002, bioassays were performed to assess N treatment effects on the composition of DOM and its interacting bacterial community. Within each site, DOM extracts from the ambient and N-amended plots were reciprocally inoculated with bacteria from each plot. After a 48 h incubation at 20 °C, community activity in each microcosm was profiled by measuring 10 extracellular enzyme activities (EEA). MANOVA showed that ecosystem type, sampling date, DOM source (ambient or N-amended plot) and inoculum source (ambient or N-amended plot) all had significant effects on bioassay EEA. Post hoc tests (Tukey's HSD) found significant reductions in oxidative enzyme activity as a result of the N treatment. In general, the bioassay results corroborated a previous report describing losses in soil oxidative enzyme activity in response to N saturation. However, it is not clear whether increased DOC concentration is the direct result of reduced oxidative activity.