Effects of nutrient enrichment on the decomposition of wood and associated microbial activity in streams

1. We determined the effects of nutrient enrichment on wood decomposition rates and microbial activity during a 3‐year study in two headwater streams at Coweeta Hydrologic Laboratory, NC, U.S.A. After a 1‐year pretreatment period, one of the streams was continuously enriched with inorganic nutrients (nitrogen and phosphorus) for 2 years while the other stream served as a reference. We determined the effects of enrichment on both wood veneers and sticks, which have similar carbon quality but differ in physical characteristics (e.g. surface area to volume ratios, presence of bark) that potentially affect microbial colonisation and activity. 2. Oak wood veneers (0.5 mm thick) were placed in streams monthly and allowed to decompose for approximately 90 days. Nutrient addition stimulated ash‐free dry mass loss and increased mean nitrogen content, fungal biomass and microbial respiration on veneers in the treatment stream compared with the reference. The magnitude of the response to enrichment was great, with mass loss 6.1 times, and per cent N, fungal biomass and microbial respiration approximately four times greater in the treatment versus reference stream. 3. Decomposition rate and nitrogen content of maple sticks (ca. 1–2 cm diameter) also increased; however, the effect was less pronounced than for veneers. Wood response overall was greater than that determined for leaves in a comparable study, supporting the hypothesis that response to enrichment may be greater for lower quality organic matter (high C : N) than for higher quality (low C : N) substrates. 4. Our results show that moderate nutrient enrichment can profoundly affect decomposition rate and microbial activity on wood in streams. Thus, the timing and availability of wood that provides retention, structure, attachment sites and food in stream ecosystems may be affected by nutrient concentrations raised by human activities.     

Nutrients and temperature additively increase stream microbial respiration

Rising temperatures and nutrient enrichment are co‐occurring global‐change drivers that stimulate microbial respiration of detrital carbon, but nutrient effects on the temperature dependence of respiration in aquatic ecosystems remain uncertain. We measured respiration rates associated with leaf litter, wood, and fine benthic organic matter (FBOM) across seasonal temperature gradients before (PRE) and after (ENR1, ENR2) experimental nutrient (nitrogen [N] and phosphorus [P]) additions to five forest streams. Nitrogen and phosphorus were added at different N:P ratios using increasing concentrations of N (~80–650 μg/L) and corresponding decreasing concentrations of P (~90–11 μg/L). We assessed the temperature dependence, and microbial (i.e., fungal) drivers of detrital mass‐specific respiration rates using the metabolic theory of ecology, before vs. after nutrient enrichment, and across N and P concentrations. Detrital mass‐specific respiration rates increased with temperature, exhibiting comparable activation energies (E, electronvolts [eV]) for all substrates (FBOM E = 0.43 [95% CI = 0.18–0.69] eV, leaf litter E = 0.30 [95% CI = 0.072–0.54] eV, wood E = 0.41 [95% CI = 0.18–0.64] eV) close to predicted MTE values. There was evidence that temperature‐driven increased respiration occurred via increased fungal biomass (wood) or increased fungal biomass‐specific respiration (leaf litter). Respiration rates increased under nutrient‐enriched conditions on leaves (1.32×) and wood (1.38×), but not FBOM. Respiration rates responded weakly to gradients in N or P concentrations, except for positive effects of P on wood respiration. The temperature dependence of respiration was comparable among years and across N or P concentration for all substrates. Responses of leaf litter and wood respiration to temperature and the combined effects of N and P were similar in magnitude. Our data suggest that the temperature dependence of stream microbial respiration is unchanged by nutrient enrichment, and that increased temperature and N + P availability have additive and comparable effects on microbial respiration rates.     

A Cross-System Comparison of Bacterial and Fungal Biomass in Detritus Pools of Headwater Streams

The absolute amount of microbial biomass and relative contribution of fungi and bacteria are expected to vary among types of organic matter (OM) within a stream and will vary among streams because of differences in organic matter quality and quantity. Common types of benthic detritus [leaves, small wood, and fine benthic organic matter (FBOM)] were sampled in 9 small (1st-3rd order) streams selected to represent a range of important controlling factors such as surrounding vegetation, detritus standing stocks, and water chemistry. Direct counts of bacteria and measurements of ergosterol (a fungal sterol) were used to describe variation in bacterial and fungal biomass. There were significant differences in bacterial abundance among types of organic matter with higher densities per unit mass of organic matter on fine particles relative to either leaves or wood surfaces. In contrast, ergosterol concentrations were significantly greater on leaves and wood, confirming the predominance of fungal biomass in these larger size classes. In general, bacterial abundance per unit organic matter was less variable than fungal biomass, suggesting bacteria will be a more predictable component of stream microbial communities. For 7 of the 9 streams, the standing stock of fine benthic organic matter was large enough that habitat-weighted reach-scale bacterial biomass was equal to or greater than fungal biomass. The quantities of leaves and small wood varied among streams such that the relative contribution of reach-scale fungal biomass ranged from 10% to as much as 90% of microbial biomass. Ergosterol concentrations were positively associated with substrate C:N ratio while bacterial abundance was negatively correlated with C:N. Both these relationships are confounded by particle size, i.e., leaves and wood had higher C:N than fine benthic organic matter. There was a weak positive relationship between bacterial abundance and streamwater soluble reactive phosphorus concentration, but no apparent pattern between either bacteria or fungi and streamwater dissolved inorganic nitrogen. The variation in microbial biomass per unit organic matter and the relative abundance of different types of organic matter contributed equally to driving differences in total microbial biomass at the reach scale.     

Comparison of Fungal Activities on Wood and Leaf Litter in Unaltered and Nutrient-Enriched Headwater Streams

Fungi are the dominant organisms decomposing leaf litter in streams and mediating energy transfer to other trophic levels. However, less is known about their role in decomposing submerged wood. This study provides the first estimates of fungal production on wood and compares the importance of fungi in the decomposition of submerged wood versus that of leaves at the ecosystem scale. We determined fungal biomass (ergosterol) and activity associated with randomly collected small wood (<40 mm diameter) and leaves in two southern Appalachian streams (reference and nutrient enriched) over an annual cycle. Fungal production (from rates of radiolabeled acetate incorporation into ergosterol) and microbial respiration on wood (per gram of detrital C) were about an order of magnitude lower than those on leaves. Microbial activity (per gram of C) was significantly higher in the nutrient-enriched stream. Despite a standing crop of wood two to three times higher than that of leaves in both streams, fungal production on an areal basis was lower on wood than on leaves (4.3 and 15.8 g C m⁻² year⁻¹ in the reference stream; 5.5 and 33.1 g C m⁻² year⁻¹ in the enriched stream). However, since the annual input of wood was five times lower than that of leaves, the proportion of organic matter input directly assimilated by fungi was comparable for these substrates (15.4 [wood] and 11.3% [leaves] in the reference stream; 20.0 [wood] and 20.2% [leaves] in the enriched stream). Despite a significantly lower fungal activity on wood than on leaves (per gram of detrital C), fungi can be equally important in processing both leaves and wood in streams.     

Effects of agriculture on wood breakdown and microbial biofilm respiration in southern Appalachian streams

1. Agriculture causes high sediment, nutrient and light input to streams, which may affect rates of ecosystem processes, such as organic matter decay. In the southern Appalachians, socioeconomic trends over the past 50 years have caused widespread abandonment of farmland with subsequent reforestation. Physical and chemical properties of streams in these reforested areas may be returning to pre‐agriculture levels thereby creating the potential for recovery of ecosystem processes. 2. We examined wood breakdown and microbial activity on wood substrata in streams with different historical and current agricultural activity in their catchments. We analysed historical (1950) and recent (1998) forested land cover from large areas of the southern Appalachians and categorized streams based on percent forested land cover in these two time periods. Categories included a gradient of current agriculture from forested to heavily agricultural and reforestation from agriculture due to land abandonment. We compared microbial respiration on wood veneer substrata and breakdown of wood veneers among these land‐use categories. We also compared temperature, sediment accumulation and nitrogen and phosphorus concentrations. 3. Streams with current agriculture had higher concentrations of dissolved inorganic nitrogen than forested streams. Despite reforestation from agriculture, nitrogen concentrations were also elevated in streams with agricultural histories relative to forested streams. Temperature was also higher in agricultural streams but appeared to recover from historical agriculture through reforestation and stream shading. 4. Wood breakdown rates ranged from 0.0015 to 0.0076 day^?1 and were similar to other studies using wood veneers to determine breakdown rate. Microbial respiration increased with incubation time in streams up to approximately 150 days, after which it remained constant. Neither wood breakdown nor microbial respiration was significantly different among land‐use categories, despite the observed physical and chemical differences in streams based on land‐use. Wood breakdown rates could be predicted by microbial respiration indicating microbial control of wood breakdown in these streams. Both breakdown and microbial respiration were negatively correlated with the amount of inorganic sediment accumulated on wood veneers. 5. Higher nutrients and temperature led us to expect faster breakdown and higher microbial respiration in agricultural streams, but sediment in these streams may be limiting microbial activity and breakdown of organic material resulting in little net effect of agriculture on wood breakdown. Wood may not be desirable as a tool for functional assessment of stream integrity due to its unpredictable response to agriculture.     

Microbial activity and soil respiration under nitrogen addition in Alaskan boreal forest

Climate warming could increase rates of soil organic matter turnover and nutrient mineralization, particularly in northern high‐latitude ecosystems. However, the effects of increasing nutrient availability on microbial processes in these ecosystems are poorly understood. To determine how soil microbes respond to nutrient enrichment, we measured microbial biomass, extracellular enzyme activities, soil respiration, and the community composition of active fungi in nitrogen (N) fertilized soils of a boreal forest in central Alaska. We predicted that N addition would suppress fungal activity relative to bacteria, but stimulate carbon (C)‐degrading enzyme activities and soil respiration. Instead, we found no evidence for a suppression of fungal activity, although fungal sporocarp production declined significantly, and the relative abundance of two fungal taxa changed dramatically with N fertilization. Microbial biomass as measured by chloroform fumigation did not respond to fertilization, nor did the ratio of fungi : bacteria as measured by quantitative polymerase chain reaction. However, microbial biomass C : N ratios narrowed significantly from 16.0 ± 1.4 to 5.2 ± 0.3 with fertilization. N fertilization significantly increased the activity of a cellulose‐degrading enzyme and suppressed the activities of protein‐ and chitin‐degrading enzymes but had no effect on soil respiration rates or ¹⁴C signatures. These results indicate that N fertilization alters microbial community composition and allocation to extracellular enzyme production without affecting soil respiration. Thus, our results do not provide evidence for strong microbial feedbacks to the boreal C cycle under climate warming or N addition. However, organic N cycling may decline due to a reduction in the activity of enzymes that target nitrogenous compounds.     

A closeup study of early beech litter decomposition: potential drivers and microbial interactions on a changing substrate

Aims

Litter decomposition and subsequent nutrient release play a major role in forest carbon and nutrient cycling. To elucidate how soluble or bulk nutrient ratios affect the decomposition process of beech (Fagus sylvatica L.) litter, we conducted a microcosm experiment over an 8 week period. Specifically, we investigated leaf-litter from four Austrian forested sites, which varied in elemental composition (C:N:P ratio). Our aim was to gain a mechanistic understanding of early decomposition processes and to determine microbial community changes.

Methods

We measured initial litter chemistry, microbial activity in terms of respiration (CO₂), litter mass loss, microbial biomass C and N (C_mic and N_mic), non purgeable organic carbon (NPOC), total dissolved nitrogen (TDN), NH₄ ⁺, NO₃ ^- and microbial community composition (phospholipid fatty acids – PLFAs).

Results

At the beginning of the experiment microbial biomass increased and pools of inorganic nitrogen (N) decreased, followed by an increase in fungal PLFAs. Sites higher in NPOC:TDN (C:N of non purgeable organic C and total dissolved N), K and Mn showed higher respiration.

Conclusions

The C:N ratio of the dissolved pool, rather than the quantity of N, was the major driver of decomposition rates. We saw dynamic changes in the microbial community from the beginning through the termination of the experiment.     

Organic matter breakdown as a measure of stream health in New Zealand streams affected by acid mine drainage

Functional indicators of stream health have the potential to provide insights into stream condition that cannot be gained by traditional structural indices. We examined breakdown of leaves, wood, and cotton cloth strips at 18 sites along a gradient of effects of drainage from coal mines in New Zealand to determine the usefulness of these methods as functional indicators of stream health. The pH varied from 2.7 to neutral across the streams, and the more acidic streams typically had higher concentrations of aluminum, iron, zinc, and other metal ions. Precipitates of metal (mainly iron) hydroxides were present in most streams affected by mine drainage, especially in those with a pH of 4–5. Breakdown rates of all organic matter types were highest in several reference streams with neutral pH and lowest in sites with high rates of metal hydroxide deposition. Breakdown was relatively fast in the most acidic streams (pH < 3), in some cases as fast as at reference sites; these sites also had elevated nutrient concentrations. Shredding invertebrates were absent in litterbags from acidic streams and common at only 2 reference sites; their presence contributed to fast breakdown of leaves in the field and in lab microcosms. Microbial respiration was closely related to breakdown rates of leaves and wood; it was high at neutral and highly acidic streams, but lower at sites with pH 4–5, where metal hydroxides were precipitating onto solid surfaces. In these metal hydroxide-stressed streams, leaf and wood breakdown was slower, and associated biota, including microbes, were more affected than by water chemistry stressors (pH, dissolved metals) associated with mine drainage. Litter breakdown and microbial respiration provide insight into the functioning of streams, yielding different responses than traditional structural measures based on macroinvertebrates, which did not accurately distinguish impacts from acid mine drainage.     

Irrigation and enhanced soil carbon input effects on below-ground carbon cycling in semiarid temperate grasslands

Global climate change is generally expected to increase net primary production, resulting in increased soil carbon (C) inputs. To gain an understanding of how such increased soil C inputs would affect C cycling in the vast grasslands of northern China, we conducted a field experiment in which the responses of plant and microbial biomass and respiration were studied. Our experiment included the below-ground addition of particulate organic matter (POM) at rates equivalent to 0, 60, 120 and 240 g C m(-2), under either natural precipitation or under enhanced precipitation during the summer period (as predicted for that region in recent simulations using general circulation models). We observed that addition of POM had a large effect on soil microbial biomass and activity and that a major part of the added C was rapidly lost from the system. This suggests that microbial activity in the vast temperate grassland ecosystems of northern China is energy-limited. Moreover, POM addition (and the associated nutrient release) affected plant growth much more than the additional water input. Although we performed no direct fertilization experiments, the response of plant productivity to POM addition (and associated release of nutrients) leads us to believe that plant productivity in the semiarid grassland ecosystems of northern China is primarily limited by nutrients and not by water.     

Chemistry and microbial activity of forest and pasture riparian-zone soils along three Pacific Northwest streams

Throughout the United States, agricultural practices are responsible for large quantities of nutrients entering lakes and streams. Previous studies have shown that forested riparian areas can filter nutrients from surface runoff and groundwater that may potentially contaminate lakes and streams. This study examined seasonal differences in soil chemistry and soil microorganisms in paired mixed-forest riparian and pasture systems, the aim being to gain understanding of the sequestering of N and P. The forest soils retained higher levels of organic C and N, mineralizable N, extractable P, and fungal biomass, and had higher respiration rates than pasture soils. These findings suggest that forested riparian zones have a greater capacity than pasture soils to sequester C and retain nutrients. In past studies, fungal biomass has been shown to be less than bacterial biomass in grassland soils, but in this study, fungal biomass was greater than bacterial biomass throughout the year in both forest and pasture soils.     

Synergistic effects of water temperature and dissolved nutrients on litter decomposition and associated fungi

In woodland streams, the decomposition of allochthonous organic matter constitutes a fundamental ecosystem process, where aquatic hyphomycetes play a pivotal role. It is therefore greatly affected by water temperature and nutrient concentrations. The individual effects of these factors on the decomposition of litter have been studied previously. However, in the climate warming scenario predicted for this century, water temperature and nutrient concentrations are expected to increase simultaneously, and their combined effects on litter decomposition and associated biological activity remains unevaluated. In this study, we addressed the individual and combined effects of water temperature (three levels) and nutrient concentrations (two levels) on the decomposition of alder leaves and associated aquatic hyphomycetes in microcosms. Decomposition rates across treatments varied between 0.0041 day^?1 at 5 °C and low nutrient level and 0.0100 day^?1 at 15 °C and high nutrient level. The stimulation of biological variables at high nutrients and temperatures indicates that nutrient enrichment of streams might have a higher stimulatory effect on fungal performance and decomposition rates under a warming scenario than at present. The stimulation of fungal biomass and sporulation with increasing temperature at both nutrient levels shows that increases in water temperature might enhance fungal growth and reproduction in both oligotrophic and eutrophic streams. The stimulation of fungal respiration and litter decomposition with increasing temperature at high nutrients indicates that stimulation of carbon mineralization will probably occur at eutrophied streams, while oligotrophic conditions seem to be ‘protected’ from warming. All biological variables were stimulated when both factors increased, as a result of synergistic interactions between factors. Increased water temperature and nutrient level also affected the structure of aquatic hyphomycete assemblages. It is plausible that if water quality of presently eutrophied streams is improved, the potential stimulatory effects of future increases in water temperature on aquatic biota and processes might be mitigated.     

Leaf litter decomposition and microbial activity in nutrient-enriched and unaltered reaches of a headwater stream

SUMMARY 1. Decomposition of red maple ( Acer rubrum ) and rhododendron ( Rhododendron maximum ) leaves and activity of associated microorganisms were compared in two reaches of a headwater stream in Coweeta Hydrologic Laboratory, NC, U.S.A. The downstream reach was enriched with ammonium, nitrate, and phosphate whereas the upstream reach was not altered.
2. Decomposition rate, microbial respiration, fungal and bacterial biomass, and the sporulation rate of aquatic hyphomycetes associated with decomposing leaf material were significantly higher for both leaf types in the nutrient-enriched reach. Species richness and community structure of aquatic hyphomycetes also exhibited considerable changes with an increase in the number of fungal codominants in the nutrient-enriched reach.
3. Fungal biomass was one to two orders of magnitude greater than bacterial biomass in both reaches. Changes in microbial respiration rate corresponded to those in fungal biomass and sporulation, suggesting a primary role of fungi in leaf decomposition.
4. Nutrient enrichment increased microbial activity, the proportion of leaf carbon channelled through the microbial compartment and the decomposition rate of leaf litter.     

Breakdown of tussock grass in streams along a gradient of agricultural development in New Zealand

Summary 1. We measured the breakdown rate of tussock grass in 12 New Zealand streams in catchments that provided a gradient of agricultural development. We also examined the microbial and invertebrate communities associated with decomposing tussock litter.
2. Pristine streams in the study had low concentrations of dissolved inorganic nitrogen (<10  μ g L⁻¹) and dissolved reactive phosphate (<3  μ g L⁻¹), whereas streams in the most developed catchments had high concentrations of nitrate (>2500  μ g L⁻¹) and phosphate (35  μ g L⁻¹), as well as greater amounts of suspended sediment and fine sediment covering the streambed.
3. Breakdown rate and microbial respiration were significantly related across the sites, and both were positively related to concentrations of nitrate and phosphate. Fungal biomass, measured as ergosterol, was positively related to microbial respiration and was also higher at sites with higher concentrations of nutrients. Total and shredding invertebrates were most abundant at the sites with high nutrient concentrations, but abundance of shredding invertebrates was not significantly related to breakdown rate. Amphipods were the most common shredding invertebrate at most sites, but probably did not contribute greatly to high rates of breakdown in streams in agricultural catchments.
4. With the exception of one site, nutrients from agricultural development appeared to have larger positive effects on litter breakdown than negative effects from sedimentation. Litter breakdown can serve as a functional measure of ecosystem health in streams, but caution should be exercised when a stress, such as land use, can have both positive (nutrients) and negative (sedimentation) effects.     

Effects of Nutrient Enrichment on Decomposition and Fungal Colonization of Sweet Chestnut Leaves in an Iberian Stream (Central Portugal)

This study assessed the effect of nutrient enrichment on rates of decomposition, ergosterol concentrations (as a measure of fungal biomass), and rates of fungal sporulation of sweet chestnut (Castanea sativa Miller) leaves in a 3rd order stream (Central Portugal), with medium to high background values of nutrients. Coarse and fine mesh leaf bags were attached to nutrient diffusing substrata containing NaNO₃, KH₂PO₄, both nutrients, or no additions. Leaf breakdown rates were similar in the four treatments and in the two mesh sizes (k=−0.0155 to −0.0219 day⁻¹). Phosphorus content of P or N + P enriched leaves was higher than in the other treatments after 28 days, but there were no differences in N concentrations. Ergosterol concentrations associated with decomposing leaves were similar among treatments. The peak sporulation rates of aquatic hyphomycetes were stimulated by the addition of N + P and N but not by P alone. Results from the experiment provide evidence that leaf breakdown in the study stream, as a model for streams with naturally medium to high level of nutrients, was not nutrient-limited, and that fungal reproductive activity was limited by dissolved N but not by dissolved P in stream water.     

Tadpoles enhance microbial activity and leaf decomposition in a neotropical headwater stream

1. Relationships between biodiversity and ecosystem function are of increasing interest, particularly in freshwater ecosystems where species losses are occurring at unprecedented rates. Amphibian declines have been associated with a loss of ecosystem function in neotropical streams, but little is known of the potential roles of stream‐dwelling tadpoles in leaf decomposition. Leaf litter is an important energy source to streams, and the breakdown of this material to fine particulate organic matter (FPOM) is a key ecosystem function. 2. We used mesocosms in a natural stream setting to quantify the effects of grazing tadpoles, shredding macroinvertebrates and a combination of the two on leaf decomposition and associated microbial activity. We measured respiration rates of decomposing leaves, particulate organic matter (POM) and leaf biofilm biomass and C : N : P ratios, and leaf area loss in 4 treatments: Control, tadpole only (TP), tadpole and shredding macroinvertebrates (TP + INV) and shredding macroinvertebrates only (INV). We hypothesised that tadpoles would enhance leaf decomposition by changing nutrient availability and stimulating microbial activity. 3. Respiration rates ranged from 3.1 to 6.0 mg O₂ dry mass^?1 h^?1 and were significantly higher in the TP and TP + INV treatments than in the control. The TP + INV treatment had significantly higher POM in chambers than the control and INV treatments. The TP treatment had significantly lower leaf biofilm biomass than the control and INV treatments. 4. Tadpoles influenced the elemental balance of C and N in POM and leaf biofilm. In contrast to our prediction, molar C : N ratios were higher in the TP + INV treatment than in the control. Mean molar N : P ratios in POM were higher in the TP + INV treatment than in any other treatment. Leaf biofilm followed a similar pattern, but both TP and TP + INV had significantly higher N : P ratios than the control and INV treatments. Leaf area loss was greatest when tadpoles and invertebrates were together (TP + INV = 0.6% leaf area loss per mg organism) than separate (TP = 0.1%, INV = 3%), indicating facilitation. 5. Tadpoles indirectly affected leaf decomposition by influencing microbial communities and macroinvertebrate feeding. As such, ongoing amphibian declines may adversely affect a critical ecosystem function in freshwater habitats.     

Nutrient Uptake and Mineralization during Leaf Decay in Streams – a Model Simulation

We developed a stoichiometrically explicit computer model to examine how heterotrophic uptake of nutrients and microbial mineralization occurring during the decay of leaves in streams may be important in modifying nutrient concentrations. The simulations showed that microbial uptake can substantially decrease stream nutrient concentrations during the initial phases of decomposition, while mineralization may produce increases in concentrations during later stages of decomposition. The simulations also showed that initial nutrient content of the leaves can affect the stream nutrient concentration dynamics and determine whether nitrogen or phosphorus is the limiting nutrient. Finally, the simulations suggest a net retention (uptake > mineralization) of nutrients in headwater streams, which is balanced by export of particulate organic nutrients to downstream reaches. Published studies support the conclusion that uptake can substantially change stream nutrient concentrations. On the other hand, there is little published evidence that mineralization also affects nutrient concentrations. Also, there is little information on direct microbial utilization of nutrients contained in the decaying leaves themselves. Our results suggest several directions for research that will improve our understanding of the complex relationship between leaf decay and nutrient dynamics in streams. (© 2009 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Nitrogen processing by grazers in a headwater stream: riparian connections

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Microbial Decomposer Communities Are Mainly Structured by Trophic Status in Circumneutral and Alkaline Streams

In streams, the release of nitrogen and phosphorus is reported to affect microbial communities and the ecological processes they govern. Moreover, the type of inorganic nitrogen (NO₃, NO₂, or NH₄) may differently impact microbial communities. We aimed to identify the environmental factors that structure aquatic microbial communities and drive leaf litter decomposition along a gradient of eutrophication. We selected five circumneutral (Portuguese) and five alkaline (French) streams differing in nutrient concentrations to monitor mass loss of alder leaves, bacterial and fungal diversity by PCR-denaturing gradient gel electrophoresis, fungal biomass and reproduction, and bacterial biomass during 11 weeks of leaf immersion. The concentrations of inorganic nutrients in the stream water ranged from 5 to 300 μg liter⁻¹ soluble reactive phosphorus, 0.30 to 5.50 mg liter⁻¹ NO₃-N, 2 to 103 μg liter⁻¹ NO₂-N, and <4 to 7,100 μg liter⁻¹ NH₄-N. Species richness was maximum in moderately anthropized (eutrophic) streams but decreased in the most anthropized (hypertrophic) streams. Different species assemblages were found in subsets of streams with different trophic statuses. In both geographic areas, the limiting nutrient, either nitrate or phosphate, stimulated the microbial activity in streams of intermediate trophic status. In the hypertrophic streams, fungal biomass and reproduction were significantly lower, and bacterial biomass dramatically decreased at the site with the highest ammonium concentration. The limiting nutrients that defined the trophic status were the main factor structuring fungal and bacterial communities, whatever the geographic area. A very high ammonium concentration in stream water most probably has negative impacts on microbial decomposer communities.There is evidence that increases in nitrate and phosphate concentrations stimulate microbial respiration and fungal and bacterial activity (biomass buildup, sporulation, and/or productivity) on plant litter, leading to faster leaf decomposition in freshwaters (16, 17, 26, 34). However, fungal demands of nitrate and phosphate are reported to be fulfilled at relatively low levels (1, 12), and further increases in these nutrients in the stream water do not necessarily result in enhanced fungal activity. Besides, the form in which inorganic nutrients are present in streams, their biological availability, and even their toxicity have different ecological consequences. In densely anthropized hypertrophic streams, high levels of nitrate and phosphate were associated with decreased fungal biomass and leaf breakdown, most probably because of the high concentrations of ammonium and ammonia (2). On the other hand, the positive effects of nutrients on biomass and productivity of leaf-associated fungi can be offset by other factors, such as low oxygen concentration and sedimentation, leading to retarded decomposition (26, 33, 34).Changes in inorganic nutrient concentrations in the stream water were reported to alter the structure of fungal communities on plant litter (16, 36). Nutrient additions to moderate levels increased the diversity of fungal communities in circumneutral soft-water Appalachian mountain streams (18) but not in a Mediterranean alkaline stream (1). Moreover, fungal diversity was lower in circumneutral eutrophic streams than in reference streams (10, 35). Fungal diversity has been assessed mostly through the morphological analysis of produced conidia, not taking into account nonsporulating fungi. This raises the question of whether the differential impacts of eutrophication on fungal diversity could be due partly to difficulties in measuring actual diversity. Besides, the study of bacterial diversity on decomposing leaves has been strongly restricted to a few cultivable bacteria (<1%). Molecular typing, such as denaturing gradient gel electrophoresis (DGGE) of a specific rRNA gene region, has proved useful for assessing diversity in both leaf-associated fungi and bacteria (7, 8, 9, 11, 30).We aimed to identify the environmental factors that drive the ecological processes in freshwaters impacted by eutrophication through examination of leaf litter decomposition and associated microbial communities along a gradient of nutrient enrichment. Specifically, we addressed the following two questions: (i) which are the environmental factors that mainly structure the fungal and bacterial communities and (ii) what are the relationships between concentrations of inorganic nutrients in the stream water, leaf litter decomposition, and the activity of associated microbes? We selected 10 stream sites spanning wide concentration ranges of dissolved inorganic nitrogen (NO₃-N, NO₂-N, NH₄-N, and NH₃-N) and soluble reactive phosphorus (SRP), including 5 in northwestern Portugal with circumneutral pH and 5 in southwestern France with an alkaline pH. With these two groups of stream sites, we assessed the structure of and diversity in both sporulating and nonsporulating fungal communities, using asexual spore morphology and DGGE fingerprints of the ITS2 region, and in bacterial communities, using DGGE fingerprints of the 16S rRNA gene region. Additionally, we examined leaf mass loss and microbial activity on decomposing leaves by determining bacterial and fungal biomass and fungal reproduction.     

Whole-stream nitrate addition affects litter decomposition and associated fungi but not invertebrates

We assessed the effect of whole-stream nitrate enrichment on decomposition of three substrates differing in nutrient quality (alder and oak leaves and balsa veneers) and associated fungi and invertebrates. During the 3-month nitrate enrichment of a headwater stream in central Portugal, litter was incubated in the reference site (mean NO₃-N 82 μg l⁻¹) and four enriched sites along the nitrate gradient (214–983 μg NO₃-N l⁻¹). A similar decomposition experiment was also carried out in the same sites at ambient nutrient conditions the following year (33–104 μg NO₃-N l⁻¹). Decomposition rates and sporulation of aquatic hyphomycetes associated with litter were determined in both experiments, whereas N and P content of litter, associated fungal biomass and invertebrates were followed only during the nitrate addition experiment. Nitrate enrichment stimulated decomposition of oak leaves and balsa veneers, fungal biomass accrual on alder leaves and balsa veneers and sporulation of aquatic hyphomycetes on all substrates. Nitrate concentration in stream water showed a strong asymptotic relationship (Michaelis–Menten-type saturation model) with temperature-adjusted decomposition rates and percentage initial litter mass converted into aquatic hyphomycete conidia for all substrates. Fungal communities did not differ significantly among sites but some species showed substrate preferences. Nevertheless, certain species were sensitive to nitrogen concentration in water by increasing or decreasing their sporulation rate accordingly. N and P content of litter and abundances or richness of litter-associated invertebrates were not affected by nitrate addition. It appears that microbial nitrogen demands can be met at relatively low levels of dissolved nitrate, suggesting that even minor increases in nitrogen in streams due to, e.g., anthropogenic eutrophication may lead to significant shifts in microbial dynamics and ecosystem functioning. Electronic Supplementary Material Supplementary material is available to authorised users in the online version of this article at .