Ammonia-oxidizing archaea and bacteria differentially contribute to ammonia oxidation in soil under precipitation gradients and land legacy.

Background: Global change has accelerated the nitrogen cycle. Soil nitrogen stock degradation by microbes leads to the release of various gases, including nitrous oxide (N2O), a potent greenhouse gas. Ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) participate in the soil nitrogen cycle, producing N2O. There are outstanding questions regarding the impact of environmental processes such as precipitation and land use legacy on AOA and AOB structurally, compositionally, and functionally. To answer these questions, we analyzed field soil cores and soil monoliths under varying precipitation profiles and land legacies. Results: We resolved 28 AOA and AOB metagenome assembled genomes (MAGs) and found that they were significantly higher in drier environments and differentially abundant in different land use legacies. We further dissected AOA and AOB functional potentials to understand their contribution to nitrogen transformation capabilities. We identified the involvement of stress response genes, differential metabolic functional potentials, and subtle population dynamics under different environmental parameters for AOA and AOB. We observed that AOA MAGs lacked a canonical membrane-bound electron transport chain and F-type ATPase but possessed A/A-type ATPase, while AOB MAGs had a complete complex III module and F-type ATPase, suggesting differential survival strategies of AOA and AOB. Conclusions: The outcomes from this study will enable us to comprehend how drought-like environments and land use legacies could impact AOA- and AOB-driven nitrogen transformations in soil.

Microbial Dispersal, Including Bison Dung Vectored Dispersal, Increases Soil Microbial Diversity in a Grassland Ecosystem.

Microbial communities display biogeographical patterns that are driven by local environmental conditions and dispersal limitation, but the relative importance of underlying dispersal mechanisms and their consequences on community structure are not well described. High dispersal rates can cause soil microbial communities to become more homogenous across space and therefore it is important to identify factors that promote dispersal. This study experimentally manipulated microbial dispersal within different land management treatments at a native tallgrass prairie site, by changing the relative openness of soil to dispersal and by simulating vector dispersal via bison dung addition. We deployed experimental soil bags with mesh open or closed to dispersal, and placed bison dung over a subset of these bags, to areas with three different land managements: active bison grazing and annual fire, annual fire but no bison grazing, and no bison grazing with infrequent fire. We expected microbial dispersal to be highest in grazed and burned environments, and that the addition of dung would consistently increase overall microbial richness and lead to homogenization of communities over time. Results show that dispersal rates, as the accumulation of taxa over the course of the 3-month experiment, increase taxonomic richness similarly in all land management treatments. Additionally, bison dung seems to be serving as a dispersal and homogenization vector, based on the consistently higher taxon richness and increased community similarity across contrasting grazing and fire treatments when dung is added. This finding also points to microbial dispersal as an important function that herbivores perform in grassland ecosystems, and in turn, as a function that was lost at a continental scale following bison extermination across the Great Plains of North America in the nineteenth century. This study is the first to detect that dispersal and vector dispersal by grazing mammals promote grassland soil microbial diversity and affect microbial community composition.

Experimental nitrogen and phosphorus enrichment stimulates multiple trophic levels of algal and detrital-based food webs: a global meta-analysis from streams and rivers.

Anthropogenic increases in nitrogen (N) and phosphorus (P) concentrations can strongly influence the structure and function of ecosystems. Even though lotic ecosystems receive cumulative inputs of nutrients applied to and deposited on land, no comprehensive assessment has quantified nutrient-enrichment effects within streams and rivers. We conducted a meta-analysis of published studies that experimentally increased concentrations of N and/or P in streams and rivers to examine how enrichment alters ecosystem structure (state: primary producer and consumer biomass and abundance) and function (rate: primary production, leaf breakdown rates, metabolism) at multiple trophic levels (primary producer, microbial heterotroph, primary and secondary consumers, and integrated ecosystem). Our synthesis included 184 studies, 885 experiments, and 3497 biotic responses to nutrient enrichment. We documented widespread increases in organismal biomass and abundance (mean response = +48%) and rates of ecosystem processes (+54%) to enrichment across multiple trophic levels, with no large differences in responses among trophic levels or between autotrophic or heterotrophic food-web pathways. Responses to nutrient enrichment varied with the nutrient added (N, P, or both) depending on rate versus state variable and experiment type, and were greater in flume and whole-stream experiments than in experiments using nutrient-diffusing substrata. Generally, nutrient-enrichment effects also increased with water temperature and light, and decreased under elevated ambient concentrations of inorganic N and/or P. Overall, increased concentrations of N and/or P altered multiple food-web pathways and trophic levels in lotic ecosystems. Our results indicate that preservation or restoration of biodiversity and ecosystem functions of streams and rivers requires management of nutrient inputs and consideration of multiple trophic pathways.

Historical Drought Affects Microbial Population Dynamics and Activity During Soil Drying and Re-Wet.

A history of drought exposure promoted by variable precipitation regimes can select for drought-tolerant soil microbial taxa, but the mechanisms of survival and death of microbial populations through the selective stresses of soil drying and re-wet are not well understood. We subjected soils collected from a 15-year field drought experiment ("Altered" precipitation history with extended dry periods, versus the "Ambient" field control) to a laboratory drying/re-wetting experiment, to learn whether selective population survival, death, or maintenance of protein synthesis potential and microbial respiration through variable soil water conditions was affected by field drought legacy. Microbial community composition, as measured by Illumina MiSeq sequencing of the 16S rRNA and 16S rRNA gene, shifted with laboratory drying/re-wet and field drought treatments. In Ambient soils, there was a higher proportion of reduced OTU abundance (indicative of mortality) during re-wet, whereas Altered soils had a greater proportion of stable OTU populations that did not change in abundance (indicative of survival) through drying/re-wet. Altered soils also had a lower proportion of rRNA:rRNA genes (lower protein synthesis potential) during dry-down, a greater weighted mean rRNA operon number (potential growth rate and r-selection) which was associated with higher abundance of Firmicutes (order Bacillales), and lower average microbial respiration rates. These data demonstrate that soils with a weaker historical drought legacy exhibit a higher prevalence of microbial water-stress mortality and differential survival and death at OTU levels following short-term dryingand re-wetting, concurrent with higher carbon loss potential. This work provides novel insight into the mechanisms and consequences of soil microbial changes resulting from extended drought conditions.

Microbial assemblages reflect environmental heterogeneity in alpine streams.

Alpine streams are dynamic habitats harboring substantial biodiversity across small spatial extents. The diversity of alpine stream biota is largely reflective of environmental heterogeneity stemming from varying hydrological sources. Globally, alpine stream diversity is under threat as meltwater sources recede and stream conditions become increasingly homogeneous. Much attention has been devoted to macroinvertebrate diversity in alpine headwaters, yet to fully understand the breadth of climate change threats, a more thorough accounting of microbial diversity is needed. We characterized microbial diversity (specifically Bacteria and Archaea) of 13 streams in two disjunct Rocky Mountain subranges through 16S rRNA gene sequencing. Our study encompassed the spectrum of alpine stream sources (glaciers, snowfields, subterranean ice, and groundwater) and three microhabitats (ice, biofilms, and streamwater). We observed no difference in regional (γ) diversity between subranges but substantial differences in diversity among (ß) stream types and microhabitats. Within-stream (α) diversity was highest in groundwater-fed springs, lowest in glacier-fed streams, and positively correlated with water temperature for both streamwater and biofilm assemblages. We identified an underappreciated alpine stream type-the icy seep-that are fed by subterranean ice, exhibit cold temperatures (summer mean <2°C), moderate bed stability, and relatively high conductivity. Icy seeps will likely be important for combatting biodiversity losses as they contain similar microbial assemblages to streams fed by surface ice yet may be buffered against climate change by insulating debris cover. Our results show that the patterns of microbial diversity support an ominous trend for alpine stream biodiversity; as meltwater sources decline, stream communities will become more diverse locally, but regional diversity will be lost. Icy seeps, however, represent a source of optimism for the future of biodiversity in these imperiled ecosystems.

Global synthesis of the temperature sensitivity of leaf litter breakdown in streams and rivers.

Streams and rivers are important conduits of terrestrially derived carbon (C) to atmospheric and marine reservoirs. Leaf litter breakdown rates are expected to increase as water temperatures rise in response to climate change. The magnitude of increase in breakdown rates is uncertain, given differences in litter quality and microbial and detritivore community responses to temperature, factors that can influence the apparent temperature sensitivity of breakdown and the relative proportion of C lost to the atmosphere vs. stored or transported downstream. Here, we synthesized 1025 records of litter breakdown in streams and rivers to quantify its temperature sensitivity, as measured by the activation energy (Ea , in eV). Temperature sensitivity of litter breakdown varied among twelve plant genera for which Ea could be calculated. Higher values of Ea were correlated with lower-quality litter, but these correlations were influenced by a single, N-fixing genus (Alnus). Ea values converged when genera were classified into three breakdown rate categories, potentially due to continual water availability in streams and rivers modulating the influence of leaf chemistry on breakdown. Across all data representing 85 plant genera, the Ea was 0.34 ± 0.04 eV, or approximately half the value (0.65 eV) predicted by metabolic theory. Our results indicate that average breakdown rates may increase by 5-21% with a 1-4 °C rise in water temperature, rather than a 10-45% increase expected, according to metabolic theory. Differential warming of tropical and temperate biomes could result in a similar proportional increase in breakdown rates, despite variation in Ea values for these regions (0.75 ± 0.13 eV and 0.27 ± 0.05 eV, respectively). The relative proportions of gaseous C loss and organic matter transport downstream should not change with rising temperature given that Ea values for breakdown mediated by microbes alone and microbes plus detritivores were similar at the global scale.

Advancing the Food-Energy-Water Nexus: Closing Nutrient Loops in Arid River Corridors.

Closing nutrient loops in terrestrial and aquatic ecosystems is integral to achieve resource security in the food-energy-water (FEW) nexus. We performed multiyear (2005-2008), monthly sampling of instream dissolved inorganic nutrient concentrations (NH4-N, NO3-N, soluble reactive phosphorus-SRP) along a ∼ 300-km arid-land river (Rio Grande, NM) and generated nutrient budgets to investigate how the net source/sink behavior of wastewater and irrigated agriculture can be holistically managed to improve water quality and close nutrient loops. Treated wastewater on average contributed over 90% of the instream dissolved inorganic nutrients (101 kg/day NH4-N, 1097 kg/day NO3-N, 656 kg/day SRP). During growing seasons, the irrigation network downstream of wastewater outfalls retained on average 37% of NO3-N and 45% of SRP inputs, with maximum retention exceeding 60% and 80% of NO3-N and SRP inputs, respectively. Accurate quantification of NH4-N retention was hindered by low loading and high variability. Nutrient retention in the irrigation network and instream processes together limited downstream export during growing seasons, with total retention of 33-99% of NO3-N inputs and 45-99% of SRP inputs. From our synoptic analysis, we identify trade-offs associated with wastewater reuse for agriculture within the scope of the FEW nexus and propose strategies for closing nutrient loops in arid-land rivers.

Organic matter quantity and source affects microbial community structure and function following volcanic eruption on Kasatochi Island, Alaska.

In August 2008, Kasatochi volcano erupted and buried a small island in pyroclastic deposits and fine ash; since then, microbes, plants and birds have begun to re-colonize the initially sterile surface. Five years post-eruption, bacterial 16S rRNA gene and fungal internal transcribed spacer (ITS) copy numbers and extracellular enzyme activity (EEA) potentials were one to two orders of magnitude greater in pyroclastic materials with organic matter (OM) inputs relative to those without, despite minimal accumulation of OM (< 0.2%C). When normalized by OM levels, post-eruptive surfaces with OM inputs had the highest ß-glucosidase, phosphatase, NAGase and cellobiohydrolase activities, and had microbial population sizes approaching those in reference soils. In contrast, the strongest factor determining bacterial community composition was the dominance of plants versus birds as OM input vectors. Although soil pH ranged from 3.9 to 7.0, and %C ranged 100×, differentiation between plant- and bird-associated microbial communities suggested that cell dispersal or nutrient availability are more likely drivers of assembly than pH or OM content. This study exemplifies the complex relationship between microbial cell dispersal, soil geochemistry, and microbial structure and function; and illustrates the potential for soil microbiota to be resilient to disturbance.

Solute Concentrations Influence Microbial Methanogenesis in Coal-bearing Strata of the Cherokee Basin, USA.

Microorganisms have contributed significantly to subsurface energy resources by converting organic matter in hydrocarbon reservoirs into methane, the main component of natural gas. In this study, we consider environmental controls on microbial populations in coal-bearing strata of the Cherokee basin, an unconventional natural gas resource in southeast Kansas, USA. Pennsylvanian-age strata in the basin contain numerous thin (0.4-1.1 m) coalbeds with marginal thermal maturities (0.5-0.7% R o ) that are interbedded with shale and sandstone. We collected gas, water, and microbe samples from 16 commercial coalbed methane wells for geochemical and microbiological analysis. The water samples were Na-Cl type with total dissolved solids (TDS) content ranging from 34.9 to 91.3 g L(-1). Gas dryness values [C1/(C2 + C3)] averaged 2640 and carbon and hydrogen isotope ratios of methane differed from those of carbon dioxide and water, respectively, by an average of 65 and 183‰. These values are thought to be consistent with gas that formed primarily by hydrogenotrophic methanogenesis. Results from cultivation assays and taxonomic analysis of 16S rRNA genes agree with the geochemical results. Cultivable methanogens were present in every sample tested, methanogen sequences dominate the archaeal community in each sample (avg 91%), and few archaeal sequences (avg 4.2%) were classified within Methanosarcinales, an order of methanogens known to contain methylotrophic methanogens. Although hydrogenotrophs appear dominant, geochemical and microbial analyses both indicate that the proportion of methane generated by acetoclastic methanogens increases with the solute content of formation water, a trend that is contrary to existing conceptual models. Consistent with this trend, beta diversity analyses show that archaeal diversity significantly correlates with formation water solute content. In contrast, bacterial diversity more strongly correlates with location than solute content, possibly as a result of spatial variation in the thermal maturity of the coalbeds.

Stream microbial diversity in response to environmental changes: review and synthesis of existing research.

The importance of microbial activity to ecosystem function in aquatic ecosystems is well established, but microbial diversity has been less frequently addressed. This review and synthesis of 100s of published studies on stream microbial diversity shows that factors known to drive ecosystem processes, such as nutrient availability, hydrology, metal contamination, contrasting land-use and temperature, also cause heterogeneity in bacterial diversity. Temporal heterogeneity in stream bacterial diversity was frequently observed, reflecting the dynamic nature of both stream ecosystems and microbial community composition. However, within-stream spatial differences in stream bacterial diversity were more commonly observed, driven specifically by different organic matter (OM) compartments. Bacterial phyla showed similar patterns in relative abundance with regard to compartment type across different streams. For example, surface water contained the highest relative abundance of Actinobacteria, while epilithon contained the highest relative abundance of Cyanobacteria and Bacteroidetes. This suggests that contrasting physical and/or nutritional habitats characterized by different stream OM compartment types may select for certain bacterial lineages. When comparing the prevalence of physicochemical effects on stream bacterial diversity, effects of changing metal concentrations were most, while effects of differences in nutrient concentrations were least frequently observed. This may indicate that although changing nutrient concentrations do tend to affect microbial diversity, other environmental factors are more likely to alter stream microbial diversity and function. The common observation of connections between ecosystem process drivers and microbial diversity suggests that microbial taxonomic turnover could mediate ecosystem-scale responses to changing environmental conditions, including both microbial habitat distribution and physicochemical factors.

Factors Controlling Soil Microbial Biomass and Bacterial Diversity and Community Composition in a Cold Desert Ecosystem: Role of Geographic Scale.

Understanding controls over the distribution of soil bacteria is a fundamental step toward describing soil ecosystems, understanding their functional capabilities, and predicting their responses to environmental change. This study investigated the controls on the biomass, species richness, and community structure and composition of soil bacterial communities in the McMurdo Dry Valleys, Antarctica, at local and regional scales. The goals of the study were to describe the relationships between abiotic characteristics and soil bacteria in this unique, microbially dominated environment, and to test the scale dependence of these relationships in a low complexity ecosystem. Samples were collected from dry mineral soils associated with snow patches, which are a significant source of water in this desert environment, at six sites located in the major basins of the Taylor and Wright Valleys. Samples were analyzed for a suite of characteristics including soil moisture, pH, electrical conductivity, soil organic matter, major nutrients and ions, microbial biomass, 16 S rRNA gene richness, and bacterial community structure and composition. Snow patches created local biogeochemical gradients while inter-basin comparisons encompassed landscape scale gradients enabling comparisons of microbial controls at two distinct spatial scales. At the organic carbon rich, mesic, low elevation sites Acidobacteria and Actinobacteria were prevalent, while Firmicutes and Proteobacteria were dominant at the high elevation, low moisture and biomass sites. Microbial parameters were significantly related with soil water content and edaphic characteristics including soil pH, organic matter, and sulfate. However, the magnitude and even the direction of these relationships varied across basins and the application of mixed effects models revealed evidence of significant contextual effects at local and regional scales. The results highlight the importance of the geographic scale of sampling when determining the controls on soil microbial community characteristics.

Dynamics of ammonia-oxidizing archaea and bacteria populations and contributions to soil nitrification potentials.

It is well known that the ratio of ammonia-oxidizing archaea (AOA) and bacteria (AOB) ranges widely in soils, but no data exist on what might influence this ratio, its dynamism, or how changes in relative abundance influences the potential contributions of AOA and AOB to soil nitrification. By sampling intensively from cropped-to-fallowed and fallowed-to-cropped phases of a 2-year wheat/fallow cycle, and adjacent uncultivated long-term fallowed land over a 15-month period in 2010 and 2011, evidence was obtained for seasonal and cropping phase effects on the soil nitrification potential (NP), and on the relative contributions of AOA and AOB to the NP that recovers after acetylene inactivation in the presence and absence of bacterial protein synthesis inhibitors. AOB community composition changed significantly (P0.0001) in response to cropping phase, and there were both seasonal and cropping phase effects on the amoA gene copy numbers of AOA and AOB. Our study showed that the AOA:AOB shifts were generated by a combination of different phenomena: an increase in AOA amoA abundance in unfertilized treatments, compared with their AOA counterparts in the N-fertilized treatment; a larger population of AOB under the N-fertilized treatment compared with the AOB community under unfertilized treatments; and better overall persistence of AOA than AOB in the unfertilized treatments. These data illustrate the complexity of the factors that likely influence the relative contributions of AOA and AOB to nitrification under the various combinations of soil conditions and NH(4)(+)-availability that exist in the field.

Bacterial community structure along moisture gradients in the parafluvial sediments of two ephemeral desert streams.

Microorganisms inhabiting stream sediments mediate biogeochemical processes of importance to both aquatic and terrestrial ecosystems. In deserts, the lateral margins of ephemeral stream channels (parafluvial sediments) are dried and rewetted, creating periodically wet conditions that typically enhance microbial activity. However, the influence of water content on microbial community composition and diversity in desert stream sediments is unclear. We sampled stream margins along gradients of wet to dry sediments, measuring geochemistry and bacterial 16S rRNA gene composition, at streams in both a cold (McMurdo Dry Valleys, Antarctica) and hot (Chihuahuan Desert, New Mexico, USA) desert. Across the gradients, sediment water content spanned a wide range (1.6-37.9% w/w), and conductivity was highly variable (12.3-1,380 µS cm(-2)). Bacterial diversity (at 97% sequence similarity) was high and variable, but did not differ significantly between the hot and cold desert and was not correlated with sediment water content. Instead, conductivity was most strongly related to diversity. Water content was strongly related to bacterial 16S rRNA gene community composition, though samples were distributed in wet and dry clusters rather than as assemblages shifting along a gradient. Phylogenetic analyses showed that many taxa from wet sediments at the hot and cold desert site were related to, respectively, halotolerant Gammaproteobacteria, and one family within the Sphingobacteriales (Bacteroidetes), while dry sediments at both sites contained a high proportion of taxa related to the Acidobacteria. These results suggest that bacterial diversity and composition in desert stream sediments is more strongly affected by hydrology and conductivity than temperature.

Bacterial and archaeal amoA gene distribution covaries with soil nitrification properties across a range of land uses.

Ammonia-oxidizing bacteria and ammonia-oxidizing archaea are commonly found together in soils, yet the factors influencing their relative distribution and activity remain unclear. We examined archaeal and bacterial amoA gene distribution, and used a novel bioassay to assess archaeal and bacterial contributions to nitrification potentials in soils spanning a range of land uses (forest, pasture, cultivated and long-term fallowed cropland) along a 10 km transect. The assay, which quantifies the extent to which acetylene-inactivated soil nitrification potential recovers (RNP) in the presence of bacterial protein synthesis inhibitors, indicated a significant archaeal contribution to the nitrification potentials of the pasture and long-term fallowed soils. Archaeal amoA gene abundance did not vary significantly among the soils, but bacterial amoA gene abundance did, resulting in archaeal : bacterial amoA abundance ratios ranging from 1.1 ± 0.8 in cultivated soils to 396 ± 176 in pasture soils. Both archaeal and bacterial amoA gene compositions were heterogeneous across the landscape, but differed in their patterns of variability. Archaeal amoA gene distributions were distinct among each of the three main land-use types: forest, pasture and cropland soils. In contrast, bacterial amoA gene composition was distinct in forest and in cultivated cropland, while pasture and long-term fallowed cropland soils were similar. In both pasture and long-term fallowed cropland soils, one phylotype of Nitrosospira cluster 3a was highly abundant. This distinct bacterial amoA gene fingerprint correlated with significant contributions of archaea to RNP of both soils, despite differences in archaeal amoA gene composition between the pasture and fallowed soils. This observation suggests that the factors driving the development of ammonia-oxidizing bacteria community composition might influence the extent of archaeal contribution to soil nitrification.

Evidence for different contributions of archaea and bacteria to the ammonia-oxidizing potential of diverse Oregon soils.

A method was developed to determine the contributions of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) to the nitrification potentials (NPs) of soils taken from forest, pasture, cropped, and fallowed (19 years) lands. Soil slurries were exposed to acetylene to irreversibly inactivate ammonia monooxygenase, and upon the removal of acetylene, the recovery of nitrification potential (RNP) was monitored in the presence and absence of bacterial or eukaryotic protein synthesis inhibitors. For unknown reasons, and despite measureable NPs, RNP did not occur consistently in forest soil samples; however, pasture, cropped, and fallowed soil RNPs commenced after lags that ranged from 12 to 30 h after acetylene removal. Cropped soil RNP was completely prevented by the bacterial protein synthesis inhibitor kanamycin (800 µg/ml), whereas a combination of kanamycin plus gentamicin (800 µg/ml each) only partially prevented the RNP (60%) of fallowed soils. Pasture soil RNP was completely insensitive to either kanamycin, gentamicin, or a combination of the two. Unlike cropped soil, pasture and fallowed soil RNPs occurred at both 30°C and 40°C and without supplemental NH(4)(+) (≤ 10 µM NH(4)(+) in solution), and pasture soil RNP demonstrated ∼ 50% insensitivity to 100 µM allyl thiourea (ATU). In addition, fallowed and pasture soil RNPs were insensitive to the fungal inhibitors nystatin and azoxystrobin. This combination of properties suggests that neither fungi nor AOB contributed to pasture soil RNP and that AOA were responsible for the RNP of the pasture soils. Both AOA and AOB may contribute to RNP in fallowed soil, while RNP in cropped soils was dominated by AOB.

Stoichiometry of soil enzyme activity at global scale.

Extracellular enzymes are the proximate agents of organic matter decomposition and measures of these activities can be used as indicators of microbial nutrient demand. We conducted a global-scale meta-analysis of the seven-most widely measured soil enzyme activities, using data from 40 ecosystems. The activities of beta-1,4-glucosidase, cellobiohydrolase, beta-1,4-N-acetylglucosaminidase and phosphatase g(-1) soil increased with organic matter concentration; leucine aminopeptidase, phenol oxidase and peroxidase activities showed no relationship. All activities were significantly related to soil pH. Specific activities, i.e. activity g(-1) soil organic matter, also varied in relation to soil pH for all enzymes. Relationships with mean annual temperature (MAT) and precipitation (MAP) were generally weak. For hydrolases, ratios of specific C, N and P acquisition activities converged on 1 : 1 : 1 but across ecosystems, the ratio of C : P acquisition was inversely related to MAP and MAT while the ratio of C : N acquisition increased with MAP. Oxidative activities were more variable than hydrolytic activities and increased with soil pH. Our analyses indicate that the enzymatic potential for hydrolyzing the labile components of soil organic matter is tied to substrate availability, soil pH and the stoichiometry of microbial nutrient demand. The enzymatic potential for oxidizing the recalcitrant fractions of soil organic material, which is a proximate control on soil organic matter accumulation, is most strongly related to soil pH. These trends provide insight into the biogeochemical processes that create global patterns in ecological stoichiometry and organic matter storage.

Microbial responses to nitrogen addition in three contrasting grassland ecosystems.

The effects of global N enrichment on soil processes in grassland ecosystems have received relatively little study. We assessed microbial community response to experimental increases in N availability by measuring extracellular enzyme activity (EEA) in soils from three grasslands with contrasting edaphic and climatic characteristics: a semiarid grassland at the Sevilleta National Wildlife Refuge, New Mexico, USA (SEV), and mesic grasslands at Konza Prairie, Kansas, USA (KNZ) and Ukulinga Research Farm, KwaZulu-Natal, South Africa (SAF). We hypothesized that, with N enrichment, soil microbial communities would increase C and P acquisition activity, decrease N acquisition activity, and reduce oxidative enzyme production (leading to recalcitrant soil organic matter [SOM] accumulation), and that the magnitude of response would decrease with soil age (due to higher stabilization of enzyme pools and P limitation of response). Cellulolytic activities followed the pattern predicted, increasing 35-52% in the youngest soil (SEV), 10-14% in the intermediate soil (KNZ) and remaining constant in the oldest soil (SAF). The magnitude of phosphatase response did not vary among sites. N acquisition activity response was driven by the enzyme closest to its pH optimum in each soil: i.e., leucine aminopeptidase in alkaline soil, beta-N-acetylglucosaminidase in acidic soil. Oxidative enzyme activity varied widely across ecosystems, but did not decrease with N amendment at any site. Likewise, SOM and %C pools did not respond to N enrichment. Between-site variation in both soil properties and EEA exceeded any treatment response, and a large portion of EEA variability (leucine aminopeptidase and oxidative enzymes), 68% as shown by principal components analysis, was strongly related to soil pH (r = 0.91, P < 0.001). In these grassland ecosystems, soil microbial responses appear constrained by a molecular-scale (pH) edaphic factor, making potential breakdown rates of SOM resistant to N enrichment.