CO-OCCURRENCE OF NITROGEN FIXATION AND DENITRIFICATION ACROSS A STREAM NITROGEN GRADIENT IN A WESTERN WATERSHED

It is frequently assumed that N2 fixation and denitrification do not co-occur in streams because each process should be favored under different concentrations of reactive nitrogen. Yet, both N2 fixation and denitrification have been found to co-occur in marine and coastal ecosystems despite their differences in nitrogen requirements, and we cannot evaluate this assumption for streams because both processes are rarely quantified together. We asked if these processes could co-exist by measuring rates of N2 fixation using acetylene reduction, denitrification using acetylene block, and N2 flux using membrane inlet mass spectrometry on rocks and sediment in 8 southeastern Idaho streams encompassing a dissolved inorganic nitrogen (DIN) gradient of 6-615 µg/L. N2 flux rates on rocks had a mean of -12,000 ± 4,900 µg/m2/h and on sediment of -2,400 ± 12,000 µg/m2/h, which were significantly different. N2 fixation rates were not significantly different among rock and sediment substrate with means of 22.9 ± 54.4 and 2.2 ± 2.0 µg/m2/h, respectively. Unamended denitrification rates were significantly different among rock and sediment substrates with means of 3 ± 7 and 2248 ±1565 μg/m2/h, respectively. Amended denitrification rates were also significantly different among substrates with a mean of 352 ± 690 µg/m2/h on rocks and 18,100 ± 6287 µg/m2/h on sediment. DIN concentration was not a significant predictor of unamended denitrification rates, but was a significant predictor of N2 flux and N2fixation rates on rocks in 2016, and amended denitrification rates on sediments in 2015 and 2016, indicating that DIN concentration alone cannot predict occurrence of processes on all substrates at all times. Multiple linear regression models relating environmental variables to measured rates showed that carbon and phosphorus availability were important predictors of denitrification rates and phosphorus, carbon, and light availability were important predictors of N2 flux rates across all sites. No significant model was produced for N2 fixation rates. Environmental characteristics measured at the scale of entire stream-reaches may not be at a fine enough spatial scale to characterize and predict the co-occurrence of these processes within stream reaches. N2 flux is balanced by the rates of N2 fixation and denitrification, and in order to better understand the fluxes and cycling of N through stream ecosystems we need to examine the co-occurrence of these processes

SPATIAL HETEROGENEITY OF NITROGEN CYCLING WITHIN AND ACROSS FRESHWATER ECOSYSTEMS

Dinitrogen (N2) fixation and denitrification are two nitrogen (N) cycling processes that despite differences in environmental requirements and constraints, co-occur in aquatic ecosystems. The overall goal of this dissertation was to evaluate how spatial heterogeneity of environmental variables 1) drive hot spots of N2 fixation, denitrification rates and gene abundances in streams, 2) facilitate co-occurrence of these processes across wetland – stream – lake interfaces, and 3) affect differences in microbial community composition in streams across U.S. ecoregions. We found hot spots of both processes within 7 stream reaches in Michigan and Idaho, but rates of N2 fixation were not directly related to relative gene abundances of nifH, while denitrification rates were related to relative gene abundances of nirS. Spatial heterogeneity of organic matter and dissolved oxygen concentrations were important predictors of rates of both processes. In a survey across 5 wetland – stream – lake interfaces of Lakes Superior and Huron, we found that rates of N2 fixation and denitrification occurred across stream, wetland and shallow lake habitats and that phosphorus (P) availability was important for predicting rates of both processes, while N availability was an important predictor of denitrification and carbon (C) availability was important predictor of N2 fixation. Finally, in a survey of microbial assemblages from 30 streams across 13 U.S. ecoregions, we found that microbial community composition differed across ecoregions in alpha diversity and relative Class abundances, but little of this variation was explained by environmental variables. Together, these studies show that N2 fixation and denitrification co-occurred in stream and coastal ecosystems and across spatial scales from stream reaches to ecoregions. However, rates and microbial community composition are not explained fully xii by differences in environmental variables on the microhabitat, cross-habitat, or ecoregion scale. N alone was not always an important predictor of the processes despite N being thought of as the best indicator of these processes in the past. Overall, these studies highlight the need to include both N2 fixation and denitrification measurements in biogeochemical studies for a better understanding of the complexity of N cycling in aquatic ecosystems

A Cross-Ecoregion Evaluation of Nitrogen Fixation and Denitrification in Streams and Rivers of the United States of America

It is typically assumed that dinitrogen (N2) fixation and denitrification are mutually exclusive processes in riverine ecosystems because N2 fixation is favored in high light, low nitrogen (N) environments but denitrification is favored under anoxic, high N conditions. Yet recent work in marine and lake ecosystems has demonstrated that N2 fixation can happen under high N conditions and in sediments, challenging this assumption. We conducted a cross-ecoregion study to test the hypothesis that N2 fixation and denitrification would co-occur in streams and rivers across a range of reactive N concentrations. Between 2017 and 2019, we sampled 30 streams in 13 ecoregions, using chambers to quantify N2 flux using membrane inlet mass spectrometry, N2 fixation using acetylene reduction, denitrification using acetylene block, and microbial diversity using 16S gene sequencing. 25 of the study streams were part of the National Ecological Observatory Network or the StreamPULSE network, which provided data on water temperature, light, nutrients, discharge and metabolism. We found that N2 fixation rates were detectable in half of the streams surveyed, and were most frequently detected on rock, wood, and/or macrophyte substrates. Denitrification potential was detected in all streams, with rates 1-2 orders of magnitude higher than N2 fixation rates and the highest rates measured in sediments. Substrate heterogeneity, and associated variation in environmental conditions, appeared to facilitate the coexistence of N2 fixation and denitrification in the study streams. Rates of denitrification were significantly positively related to streamwater nitrate concentrations (r2 = 0.35), but N2 fixation rates were not, despite the common simplifying assumption that denitrification dominates the N2 flux in streams under high N and N2 fixation only occurs under low N conditions. Additional analyses are exploring reach to watershed characteristics, and metabolic regimes as drivers of cross-ecoregion patterns in processes

Biogeochemistry of upland to wetland soils, sediments, and surface waters across Mid-Atlantic and Great Lakes coastal interfaces

Transferable and mechanistic understanding of cross-scale interactions is necessary to predict how coastal systems respond to global change. Cohesive datasets across geographically distributed sites can be used to examine how transferable a mechanistic understanding of coastal ecosystem control points is. To address the above research objectives, data were collected by the EXploration of Coastal Hydrobiogeochemistry Across a Network of Gradients and Experiments (EXCHANGE) Consortium – a regionally distributed network of researchers that collaborated on experimental design, methodology, collection, analysis, and publication. The EXCHANGE Consortium collected samples from 52 coastal terrestrial-aquatic interfaces (TAIs) during Fall of 2021. At each TAI, samples collected include soils from across a transverse elevation gradient (i.e., coastal upland forest, transitional forest, and wetland soils), surface waters, and nearshore sediments across research sites in the Great Lakes and Mid-Atlantic regions (Chesapeake and Delaware Bays) of the continental USA. The first campaign measures surface water quality parameters, bulk geochemical parameters on water, soil, and sediment samples, and physicochemical parameters of sediment and soil

Co-occurrence of in-stream nitrogen fixation and denitrification across a nitrogen gradient in a western U.S. watershed

It is frequently assumed that nitrogen (N2) fixation and denitrification do not co-occur in streams because each process should be favored under different concentrations of dissolved inorganic nitrogen (DIN), and therefore these processes are rarely quantified together. We asked if these processes could co-exist by conducting a spatial survey of N2 fixation using acetylene reduction and denitrification using acetylene block [with and without amendments of carbon (C) as glucose and nitrogen (N) as nitrate]. Rates were measured on rocks and sediment in 8 southeastern Idaho streams encompassing a DIN gradient of 26–615 µg L−1. Sampling at each site was repeated in summer 2015 and 2016. We found that both denitrification and N2fixation occurred across the gradient of DIN concentrations, with N2 fixation occurring primarily on rocks and denitrification occurring in sediment. N2 fixation rates on rocks significantly decreased 100× across the DIN gradient in 1 year of the study, and amended (with N and C) denitrification rates increased 10× across the DIN gradient in both years. Multiple linear regression and partial least squares models with environmental characteristics measured at the scale of entire stream reaches showed that C and phosphorus were positive predictors of amended and unamended denitrification rates, but no significant model could explain N2fixation rates across all streams and years. This, coupled with the observation that detectable rates of N2 fixation occurred primarily on rocks and denitrification occurred primarily on sediment, suggests that microhabitat scale factors may better predict the co-occurrence of these processes within stream reaches. Overlooking the potential co-occurrence of N2 fixation and denitrification in stream ecosystems will impede understanding by oversimplifying the contribution of each process to the N cycle

Heterogeneity in habitat and nutrient availability facilitate the co-occurrence of N2 fixation and denitrification across wetland–stream–lake ecotones of Lakes Superior and Huron

Great Lakes coastlines are mosaics of wetland, stream, and lake habitats, characterized by a high degree of spatial heterogeneity that may facilitate the co-occurrence of seemingly incompatible biogeochemical processes due to variation in environmental factors that favor each process. We measured nutrient limitation and rates of N2 fixation and denitrification along transects in 5 wetland–stream–lake ecotones with different nutrient loading in Lakes Superior and Huron. We hypothesized that rates of both processes would be related to nutrient limitation status, habitat type, and environmental characteristics including temperature, nutrient concentrations, and organic matter quality. We found that median denitrification rates (914 μg N m−2 h−1) were 166 × higher than N2 fixation rates (5.5 μg N m−2 h−1), but the processes co-occurred in 48% of 83 points measured across all 5 transects and habitat types. N2 fixation occurred on sediment and macrophyte substrate, while denitrification occurred mostly in sediment. Nutrient-diffusing substrate experiments indicated that biofilm chlorophyll-a was limited by N and/or P at 55% and biofilm AFDM was limited at 26% of sample points. N2 fixation and denitrification rates did not differ significantly with differing nutrient limitation. Predictive models for N2 fixation and denitrification rates both included variables related to the composition of dissolved organic matter, while the model for N2 fixation also included P concentrations. These results demonstrate the potential for heterogeneity in habitat characteristics, nutrient availability, and organic matter composition to lead to biogeochemical complexity at the local scale, despite overall N removal at broader scales

Co-occurrence of in-stream nitrogen fixation and denitrification across a nitrogen gradient in a western U.S. watershed, 2015 - 2016

This data was collected as part of the NSF CAREER Grant DEB 14-51919 to Amy Marcarelli to evaluate if nitrogen fixation and denitrification co-occur in streams across a gradient of nitrate concentrations. This data was collected during summer 2015 and 2016 in the Portneuf River watershed near Pocatello, ID. In summer 2015, rates of nitrogen fixation were only measured on rock substrate and denitrification only on sediment substrate. In summer 2016, both process rates were measured on both rock and sediment substrate to more accurately capture variation in rates

Do nitrogen fixation and denitrification co-occur across a gradient of stream nitrogen concentrations in a western watershed?

It is frequently assumed that N2 fixation and denitrification do not co-occur in streams because they are favored under different concentrations of reactive nitrogen, yet we cannot evaluate this assumption because both processes are rarely quantified in the same stream. We measured rates of each process, as well as N2 flux, in 7 southeastern Idaho streams encompassing a DIN gradient from 6 to 582 µg/L. N2 fixation rates measured using acetylene reduction were 0-198 µg/m2/hr while denitrification rates measured using acetylene block were 0.59-5.66 µg/m2/hr. The highest N2 fixation rates occurred when DIN \u3c 13 µg/L, but low rates were observed with DIN \u3e 200 µg/L. N2 fixation rates were not significantly related to P concentrations or N:P, but N:P was \u3c 20 in all study streams. Denitrification was linearly related to DIN, but the trend appeared asymptotic. These results suggest stream nutrient concentrations alone cannot explain the distribution of N2 fixation and denitrification. We are exploring the role of small-scale variation in environmental conditions and microbial assemblages for facilitating the co-occurrence of these processes