Doctor of Philosophy / Department of Biology / John M. Blair / Juniperus virginiana, an important woody plant invader in the U.S. Central Plains, has increased considerably in density and cover in large areas previously dominated by tallgrass prairie. Change in the phenology and nitrogen use efficiency of the dominant plant communities as J. virginiana replaces native prairies may lead to increased plant productivity and biomass accumulation, but may also alter the microclimate and litter quality that affect soil microbial communities responsible for key soil processes. I have focused my investigations on changes in key soil processes that could lead to differences in soil N availability, as well as changes in ecosystem C and N pools and fluxes as J. virginiana expands into native grasslands. Juniperus virginiana forest soils exhibit greater cumulative annual net N mineralization (11.52 ± 0.38 µg N g¯1 soil y¯1) compared to prairie soils (7.90 ± 0.26 µg N g¯1 soil y¯1) (F = 60.67, P = 0.016), yet slightly reduced potential soil C flux. Examination of internal soil N cycling revealed that both J. virginiana and prairie soils minimize potential soil N losses, by rapid microbial immobilization of inorganic N, and constraining nitrification via substrate limitation or environmental constraints. Leaf-level photosynthetic nitrogen use efficiency (NUE) was over a magnitude higher in the dominant grass, Andropogon gerardii, but high annual ecosystem-level NUE and greater soil N availability may contribute to the higher productivity and rapid accrual of C in newly established J. virginiana forests. Increased plant productivity and elimination of fire in J. virginiana forests have allowed at least 80,000 kg ha-1 increase in ecosystem C storage in about half a century. Soil organic C, an important long-term sink, has also increased significantly in J. virginiana forests, with approximately 34% replacement of C4 grass-derived soil C with new C from trees in the A-horizon. The observed high productivity of J. virginiana and increased N availability necessary to support continued plant biomass accumulation are possible because of substantial (~ 44%) increase in ecosystem N in measured pools, which is a likely a result of reduced volatilization of N from biomass burning, possible increased exogenous N inputs, and/or N translocation from deeper soil horizons. Reduced fire return intervals in prairie provide an opportunity for J. virginiana to establish and facilitate N accrual, which may allow this species to accelerate is own establishment through creating conditions of increased N availability and efficient utilization of N.

Forested headwater stream ecosystems are important integrators of terrestrial and aquatic systems and their function depends greatly on water availability. In the southern Appalachians, models of future climate change predict alterations to the timing and intensity of storms such that most precipitation may be relegated to winter and spring. During the summer and fall, relatively less precipitation will translate to lower stream flows in systems that rarely experience such a lack of water. Given these predicted changes to the hydrologic cycle, I experimentally reduced flow to downstream sections of three streams at the Coweeta Hydrologic Laboratory in NC to assess changes to function in perennial ecosystems. The questions that I addressed included: 1) How is organic matter decomposition regulated by changes to the availability of water? and 2) How does the relationship between nutrient uptake and metabolism change under conditions of varying water availability? The availability of water (as discharge) was shown to be a major control of ecosystem function throughout these studies. Rates of leaf decomposition varied between red maple (Acer rubrum L.) and white oak (Quercus alba L.) with lower discharge in the early autumn regulating the breakdown trajectories of leaves through facilitation of colonization by microbes and macroinvertebrates. The return of water during the winter accelerated decomposition rates in the diverted sites such that mass of leaves remaining were similar to those in upstream sections. Colonization of decomposing organic matter by heterotrophic microbes (especially fungi) increased N immobilization leading to an increase in respiration per unit leaf standing stocks during the fall. Nitrification was detectable during summer low flows when leaf standing stocks were low. Changes in the timing and intensity of precipitation and thus discharge may in turn alter the temporal dynamics of ecosystem function. Leaves may remain in the stream unprocessed which will change the availability of food for macroinvertebrates, the production of which provides nutrition to higher trophic levels. Local-scale differences in organic matter processing and nutrient immobilization may translate to regional differences in food availability over both time and space. Hydrology not only acts as a local control of endogenous processes but acts also regionally through the transport of resources and nutrients to downstream reaches. / Ph. D.

abstract: Nitrogen is an essential, often limiting, element for biological growth that can act as a pollutant if present in excess. Nitrogen is primarily transported by water from uplands to streams and eventually to recipient lakes, estuaries, and wetlands, but can be modulated by biological uptake and transformation along these flowpaths. As a result, nitrogen can accumulate in aquatic ecosystems if supply is high or if biological retention is low. Dryland and urban ecosystems offer interesting contrasts in water supply, which limits transport and biological activity in drylands, and nitrogen supply that increases with human activity. In my dissertation, I ask: What is the relative balance among nitrogen retention, removal, and transport processes in dryland watersheds, and what is the fate of exported nitrogen? My dissertation research demonstrates that water is a major control on where and when nitrogen is retained and removed versus exported to downstream ecosystems. I used a mass-balance model based on synoptic surveys to study seasonal and spatial patterns in nitrate loading to a dryland stream network. I found that irrigation diversions transport nitrate from agricultural areas to the stream network year-round, even during dry seasons, and are an important driver of nitrate loading. I further explored how seasonal precipitation influences flood nutrient export in an intermittent desert stream by coupling long-term data of flood-water chemistry with stream discharge and precipitation data. I found that higher precipitation prior to a flood fills water storage sites in the catchment, leading to larger floods. In addition, higher antecedent precipitation stimulates biological nitrogen retention in the uplands, leading to lower nitrogen concentration in floods. Finally, I evaluated the consequences of nitrogen export from watersheds on how urban wetlands attenuate nitrate through denitrification that permanently removes nitrogen, and dissimilatory nitrate reduction to ammonium (DNRA) that retains nitrogen in another biologically reactive form. I found that DNRA becomes proportionally more important with low nitrate concentration, thereby retaining nitrogen as ammonium. Collectively, my dissertation research addresses how dryland and urban ecosystems can be integrated into models of watershed nitrogen cycling. / Dissertation/Thesis / Doctoral Dissertation Biology 2019

Tropical coral reefs are highly productive ecosystems thriving in oligotrophic waters, a phenomenon facilitated by efficient but delicate biogeochemical cycling within reef communities. Global climate change and local stressors are driving phase shifts from coral- to non-calcifier-dominated states in reefs worldwide, substantially altering reef biogeochemical functioning. While major benthic players such as coral and macroalgae have been investigated in detail regarding carbon and nutrient dynamics, the less conspicuous “reef cryptobiome” (sensu Carvalho et al., 2019) – comprising most of reef diversity – has only recently gained attention. Autonomous Reef Monitoring Structures (ARMS) have recently been developed to sample coral reef cryptobenthic communities in a non-destructive and standardised way, allowing exploration of these often overlooked biota. Here, 16 ARMS were deployed for seven months in four distinct habitats dominated by different benthic players (i.e., four units per habitat) in a nearshore Red Sea coral reef to investigate the cryptobiome associated with proxies of varying benthic states. Two of these habitats were coral-dominated, and one each dominated by turf algae or coral rubble. To assess the biogeochemical fluxes of pioneering cryptobenthic communities, ARMS were incubated in situ prior to retrieval using customised chambers. Subsequently, 16S rRNA gene amplicon and shotgun metagenomic sequencing of the ARMS sessile (i.e., encrusting) fractions were performed to link observed fluxes with prokaryotic taxonomic and functional profiles, particularly regarding nitrogen cycling. The results show that the pioneering cryptobiome represents a significant source of inorganic nutrients and that its associated microbial communities facilitate the mineralisation and assimilation of organic matter and provide crucial genetic functional pathways for nitrogen cycling. Functional similarities among habitats suggested functional redundancy despite variation in bacterial community composition. Hence, the reef cryptobiome can be considered an important biogeochemical player in coral reefs, actively shaping the abiotic conditions within niches of the reef framework and driving the recruitment and persistence of crytobenthic and other reef organisms. As communities associated with the algae-dominated reef habitat were most distinct compositionally and biogeochemically, and as non-calcifiers are becoming more dominant in many reefs, this has implications for intensifying phase shifts in coral reefs worldwide. Future ARMS studies will also benefit from adjustment of sample processing and molecular protocols, resulting in higher sample throughput and lower costs in times of increased application of ARMS.

Increased anthropogenic deposition of nitrogen (N) and land-use changes associated with planted forests have important implications for sustainable forest management and associated water quality. The purpose of the research for this dissertation was to explore how N deposition will affect the long-term health, productivity, and carbon (C) and N sequestration of conifer and hardwood forest types by examining the mechanisms controlling N cycling and NO3-N production in two watersheds with contrasting vegetation at the Fernow Experimental Forest (FEF), West Virginia. I utilized watershed C and N budgets to account for differences in stream export of NO3-N from streams draining adjacent watersheds containing (i) planted Norway spruce (Picea abies) and (ii) native Appalachian hardwoods. I also investigated spatial and temporal patterns of dissolved C and N across both watersheds and identified key soil properties associated with NO3-N in soil solution and streamwater. In a third study, I performed a soil inoculation and incubation experiment, which utilized soil from both watersheds, mixed in ratios in order to create a gradient of soil chemical and biotic characteristics. Important differences in biogeochemical cycling of C and N were documented in the watersheds after nearly 40 years of influence by contrasting vegetation. Total C and N pools were 28% and 35% lower in the spruce watershed than the hardwood watershed, respectively. Results also identify vegetation-mediated differences in soil characteristics, with lower soil pH and base cations, and higher extractable aluminum and C:N ratios measured in the spruce soil as compared to the native hardwood soil. Establishment of a spruce monoculture at the FEF significantly altered N cycling, depleted N stores, increased soil acidity, and altered organic matter dynamics, thus leading to low net nitrification rates. Carbon and N properties and processes in the soil profile should be taken into consideration in forests managed for ecosystem services including C sequestration and improvement or maintenance of water quality through alleviation of N inputs into aquatic ecosystems. / Ph. D.

I investigated the drivers of nutrient cycling by heterotrophic microbes during leaf decomposition in streams. My research addressed two overarching questions: 1) how do exogenous and endogenous factors interact to drive microbial nitrogen (N) cycling during organic matter decomposition in stream ecosystems, and 2) what affect will the global increase in biologically active N have on these factors and resulting fluxes? I conducted studies in natural streams and laboratory mesocosms to address these questions and used general stoichiometric theory to conceptualize diverse microbial assemblages as a single functional unit within stream ecosystems. First, I described spatial and temporal patterns of N and phosphorus uptake and mineralization by leaf-associated microbial assemblages in five southern Appalachian streams which spanned a gradient of nitrate availability. I found wide variations in nutrient fluxes across time and space, perhaps due to macroinvertebrate-induced changes in microbial assemblage composition. Secondly, I explored the roles of endogenous and exogenous N in meeting microbial requirements. I isolated microbial biomass from leaves that had been labeled with N-15 and incubated in the same five Appalachian streams. The importance of exogenous N increased as decomposition progressed and was particularly important in streams with high N availability. Finally, I tested potential interactions between two exogenous drivers of microbial nutrient cycling: N availability and animal activity. I used mesocosms to test the effects of consumer nutrient recycling (CNR) and grazing by two shredders on microbial uptake under different N regimes. Animals only influenced microbial uptake under low N conditions. Shredder CNR generally stimulated uptake while grazing had a negative effect. My research provides a robust model describing N cycling by detritus-associated microbes over the course of decomposition. According to this model, microbes assimilate endogenous N during the initial stages of decomposition and immobilization of exogenous N becomes more important as decomposition progresses. The labeled substrate technique that I used to generate this model is an elegant way of testing the applicability of this model in other ecosystems. My results also suggest that anthropogenic activities that increase exogenous N availability have implications for N and C cycling in lotic systems. / Ph. D.

Continental margin sediments play an important role in marine biogeochemical cycles, partly due to high primary production rates and efficient export of organic matter to sediments in margin environments. This thesis presents studies of solute exchange in fine-grained sediments representative of slope and rise environments, and carbon and nitrogen cycling in sandy sediments dominant in continental shelves worldwide. Results of these studies advance understanding of the role of benthic processes on marine ecosystems. In fine-grained sediments, solute exchange by diffusion, biological mixing and bioirrigation can be quantified using in situ flux chambers with inert tracer additions. Mechanistic models of chamber tracer transport across the seabed indicate that in organic-rich sediments, bioirrigation and mixing dominate over a wide range of bottom water oxygen levels, reflecting the patchiness and versatility of benthic macrofaunal communities. Positive correlations between benthic oxygen and tracer fluxes appear site-specific. Reliable chamber volume estimates derived from mechanistic models reveal that empirical fits to chamber tracer datasets may overestimate chamber volume and benthic solute fluxes. The biogeochemistry of sandy, highly permeable sediments that dominate continental shelves is largely unknown because of the difficulty in sampling and studying them under natural conditions. Novel sediment reactors were developed and used to mimic in situ porewater advection and natural sedimentary conditions. Compositional changes of natural seawater, with and without the addition of

L'augmentation de la dégradation des sols et ses conséquences sur les services écosystémiques nécessite le développement de stratégies de restauration de ces sols. La constitution de Technosols, résultant de l’assemblage de sols pollués et de déchet industriels, est une approche innovatrice pouvant à la fois permettre de restaurer les sols et de recycler des sous-produits industriels. Des études récentes ont mis en évidence que les Technosols pouvaient assurer des services écosystémiques tels que la production primaire. Toutefois, notre connaissances des autres services écosystémiques rendus par les Technosols tels que les cycles biogéochimiques est limitée. En raison de la contribution significative des communautés microbiennes aux cycles biogéochimiques dans les sols, l’objectif de ce travail de thèse était l’effet du type de Technosol sur les communautés microbiennes et plus particulièrement les communautés fonctionnelles impliquées dans le cycle de l’azote. Dans ce contexte, (i) la densité et la diversité de la communauté bactérienne totale, (ii) la densité de la communauté crenarchéenne ainsi que (iii) la densité et les activités des communautés nitrifiante et dénitrifiante ont été étudiées dans deux types de Technosols différents. Les résultats obtenus montrent que la diversité et la composition de la communauté bactérienne des deux Technosols n'étaient pas significativement différentes entre elles, et similaires à celles de ‘sols naturels’, les Proteobacteria étant le phylum dominant (50-80%). La densité de la communauté bactérienne oxydant l’ammonium était non seulement plus importante que celle des crenarcheae qui oxyde l’ammonium, mais est également corrélée avec l'activité potentielle de nitrification suggérant ainsi que les bactéries sont responsables de l’oxydation de l’ammonium dans les Technosols. La densité des dénitrifiants et de la communauté oxydant l’ammonium sont du même ordre de grandeur que celles observées dans les sols agricoles. L’analyse de la distribution de l'activité et de la densité des communautés nitrifiante et dénitrifiante dans les différents horizons des Technosols montre un effet négatif de la profondeur, cet effet étant plus marqué que l’effet du type de Technosol étudié. Les propriétés physico-chimiques des Technosols et la densité de la communauté bactérienne oxydant l’ammonium sont corrélés à l'activité de nitrification alors que l'activité de dénitrification était contrôlée principalement par les propriétés physico-chimiques des Technosols, et dans une moindre mesure par la densité de la communauté dénitrifiant possédant le gène nirS. L'estimation de la stabilité fonctionnelle du processus de dénitrification vis-à-vis de périodes stress hydrique et thermique a montré que les Technosols présentaient une plus haute résistance et une meilleure résilience que des sols remédiés par traitement thermique uniquement. Ce travail souligne le potentiel des Technosols à assurer les services écosystémiques tels que le cycle de l’azote, ainsi que leur forte capacité à résister et à se remettre de stress environnementaux. Tout ceci semble donc indiquer que la construction de Technosols est une technologie prometteuse qui pourrait permettre la restauration de friches industrielles et le recyclage des déchets industriels / Increasing soil degradation and its consequences on overall ecosystem services urge for restoration strategies. Construction of Technosols through assemblage of treated soil and industrial wastes is an innovative technology for the restoration of polluted land and re-use of industrial by-products. Recent studies have evidenced that Technosols could support ecosystemic services such as primary production but the knowledge about other soil functions, such as biogeochemical cycling, is limited. Due to the significant contribution of microbial communities to soil functioning, this PhD work was carried out to study the effect of the type of Technosol on microbial communities with a focus on functional guilds involved in N cycling. For this purpose, the abundance and diversity of the total bacterial community and the abundance of crenarchaeal community together with the abundance and activities of the nitrifying and denitrifying communities were investigated in two types of Technosols. Results demonstrated that diversity and composition of the bacterial community were similar to ‘natural soils’ and were not significantly different between the two Technosols with Proteobacteria being the dominant phylum (50-80%). The bacterial ammonia oxidizers were greater in number than crenarchaeal ammonia oxidizers but also correlated to the potential nitrification activity suggesting that bacteria are the dominant ammonia oxidizers in Technosols. The abundance of both the ammonia oxidizers and the denitrifiers were in the same range than that observed in other soil systems. Analyses of the vertical distribution of the activity and abundance of N-cycling communities in the Technosols showed a significant depth-effect, which was more important than the Technosol type-effect. Technosols physicochemical properties and the abundance of the bacterial ammonia oxidizers were the main drivers of the nitrification activity whereas the denitrification activity was controlled mainly by the Technosols physicochemical properties and, to a minor extent, by the abundances of the nirS denitrifiers. The estimation of the functional stability of the denitrification process against the heat-drought stresses revealed that Technosol exhibited the high resistance and resilience in comparison to the thermally treated soil. This work highlighted the potential of constructed Technosols to ensure the N cycling ecosystem services, along with a high capacity to resist and recover from environmental stresses, suggesting that construction of Technosols is a promising technology and a solution for the restoration of industrial wastelands and waste recycling

A expansão agrícola tem provocado modificações expressivas na dinâmica de nitrogênio (N) em sistemas tropicais. No Brasil, a expansão dos cultivos de soja é uma realidade e, portanto, investigações a cerca dos processos que controlam o ciclo de nitrogênio nestes sistemas são fundamentais. A fixação biológica de nitrogênio (FBN) por leguminosas pode promover aportes significativos de N nos sistemas agrícolas em solos tropicais, no entanto, o destino destes aportes e o balanço entre entradas e saídas de N não é completamente entendido. Este trabalho teve como objetivo investigar comparativamente a dinâmica de nitrogênio em cultivos de soja e floresta no estado de Mato Grosso, sudeste da Amazônia. Foram determinados o \'delta\'15N e %N do solo e da vegetação, estoques de N e C, N-NO3-, N-NH4+, bem como outras propriedades químicas e físicas do solo em áreas de floresta e em áreas submetidas a cultivos de soja ao longo de uma cronosequência (1, 2, 5 e 6 anos de cultivo). Foram realizadas estimativas de FBN (Fixação Biológica de Nitrogênio) em cultivos de soja utilizando a abundância natural de 15N sob condições de campo. A conversão das áreas de cultivo partiu de pastagem, sendo esta área utilizada como referência inicial quanto aos estoques de N e ao \'delta\'15N do solo. Foi observado um aumento significativo nos estoques de nitrogênio do solo (0 a 10cm) ao longo dos anos de cultivo de soja, estes no entanto, foram menores que os estoques encontrados na floresta. Os estoques de N no solo (0-10cm) variaram de 1230 kg N ha-1 na pastagem a 1370 kg N ha-1 nos cultivos mais antigos. O acúmulo anual de N pela soja foi de 158,6 kg N ha-1 nos cultivos mais antigos, do qual 79% foi derivado da FBN. Não foram encontradas diferenças significativas nas taxas de mineralização e nitrificação líquida entre as áreas, no entanto, altos valores de N-NO3- foram encontrados nas camadas mais profundas de solo em cultivos de soja. Apesar de não serem observadas diferenças significativas no \'delta\'15N do solo entre os cultivos, estes, no entanto, apresentaram valores de \'delta\'15N intermediários entre a pastagem e a floresta. Os resultados indicaram um padrão de acúmulo de nitrogênio ao longo da cronosequência de cultivos de soja, indicando um possível retorno gradual dos estoques de N e do sinal isotópico do solo que ocorriam na floresta antes da conversão para pastagem e cultivo de soja, este retorno, no entanto, não parece acontecer a médio-prazo / Agricultural expansion has greatly changed the nitrogen (N) dynamics in tropical systems. The expanding soybean frontier in Brazil is a reality, and investigations of the processes driving N dynamics in these systems are needed to minimize environmental impacts and to promote the sustainability of agricultural systems. Biological nitrogen fixation (BNF) by legumes can provide significant N inputs to crop systems on highly weathered tropical soils, although the fate of these inputs in the environment and the balance between inputs and outputs of N in these systems is poorly understood. This work investigated N dynamics in a chronosequence of soybean fields (1, 2, 5 and 6 years of cultivation) and mature forest in the Brazilian state of Mato Grosso, which is at the southern limit of the Amazon forest. We measured soil N and C stocks, N-NO3-, N-NH4+ concentration and soil \'delta\'15N as well as biological nitrogen fixation (BNF) inputs by soybean, which were assessed using the 15N natural abundance technique under field conditions. Additional measurements of physical and chemical properties of soils were also provided. The land-use-conversion started from pasture, so this site was used as an initial reference for soil N stocks and soil \'delta\'15N. Mature forest stands on the ranch were also used as an additional reference. Soil N stocks (top 10cm) ranged from 1230 kg N ha-1 in the pasture to 1370 kg N ha-1 in the oldest soybean fields. The trend of increasing N stocks with field age was statistically significant. The annual N accumulation by soybean plant biomass was 158,6 kg N ha-1 in the oldest soy fields, of which 79% was estimated to be derived by BNF, based on the natural abundance technique. There was no statistically significant trend in the mineralization and nitrification rates among the areas. However, extracts of soil profiles showed significant increases in deep soil nitrate concentrations in soybean soils compared to forest soils. There was no statistically significant trend in the soil \'delta\'15N within the chronosequence of soybean fields, although the values were intermediate between the soil \'delta\'15N values found in the pasture and the forest. These results showed a pattern of nitrogen accumulation in the soil along the chronosequence of soybean fields, indicating a possible gradual return to soil N stocks and isotopic signatures occurring in the forest soil before the conversion to pasture and soybean, although this may not happen in the near future

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