The effect of spatial heterogeneity on nitrate reduction in soil systems

Nitrogen is not only an abundant element on earth, making up roughly 80%of the earth's atmosphere, it is also essential for life, and a functional nitrogen cycle is of great importance to human activities and our ecosystems. The nitrogen cycle ultimately returns reactive nitrogen, which was chemically or biochemically fixed from inert nitrogen, back into the atmosphere as inert nitrogen. Over the last century, the excess of anthropogenically fixed nitrogen has put increasing pressures on the nitrogen cycle. Nitrate is a central molecule in the nitrogen cycle. Its concentration is, on the one hand governed by formation by oxidation of ammonia-N, and on the other hand by removal a removal by two dissimilatory nitrate reduction processes:denitrification, in which nitrate is converted to the gaseous compounds dinitrogen and nitrous oxide, and dissimilatory nitrate reduction to ammonium, DNRA. While both processes bring about the reduction of nitrate, their impact on ecosystems is radically different – especially in soil environments. Nitrate itself is poorly retained in soils, and its conversion to gaseous dinitrogen and nitrous oxide through denitrification only serves to further the loss of reactive nitrogen from the system. On top of that nitrous oxide is an important air pollutant and greenhouse gas, with a global warming potential per unit mass 300 times higher than carbon dioxide. DNRA, on the other hand, converts nitrate to ammonium, which is more easily retained in soils than nitrate, and can be assimilated into organic matter, effectively bypassing both denitrification and dinitrogen fixation and conserving nitrogen in the ecosystem. It is well established that soil is an extremely heterogeneous environment, not merely on a macroscopic level, but also on a microscopic level. Spatial heterogeneity and diffusive limitations result in the formation of specialized niches. It is becoming increasingly clear that these factors are of great importance for biogeochemical processes such as the carbon cycle. Studying the heterogeneity of soil and its impact on ecological processes is not merely a fascinating scientific activity, it may very well be central to gaining insights to influence fundamental soil processes such as nitrogen metabolism, promising advancement of agricultural and pollution prevention and remediation techniques.A number of conceptual and quantitative frameworks have been developed to assess the impact of mass transfer kinetics on biotransformation rates in various environments. One such approach uses the dimensionless parameter Da3: a Damköhler number which for a given system quantifies the relative impact of diffusive limitation on biotransformation. During this PhD project, we specifically examined the incidence of denitrification and DNRA in soil systems and studied the impact of diffusive limitation on their relative occurrence. An array of column-based soil microcosms was set up to look at the incidence and magnitude of DNRA vs. denitrification in Fagus sylvatica forest soil litter, and to investigate the effect of electron donor abundance and bacterial inoculum size on nitrate reduction. Increasing the electron donor abundance increased both DNRA and denitrification, allowing both processes to coexist in the system. At reduced biokinetic limitations, obtained by increasing the initial inoculum size, nitrate reduction was barely affected, but DNRA increased substantially by 71%. Additionally, nitrite-, ammonium-, and nitrous oxide were sequentially produced during nitrate reduction: an initial burst of nitrite production led to DNRA, and for the microcosms which became mass transfer limited also to nitrous oxide production.To allow application of the Damköhler number framework, a well-controlled experimental system was required where interfacial areas of diffusion are well defined. Hence, a protocol for encasing soil material in alginate aggregates with well-defined size and geometry was developed. The degree of diffusive limitation was then imposed using soil-alginate aggregates with different defined Da3 values. These were applied in an array of column microcosms to investigate the effect of diffusive limitation on nitrate reduction processes. Going from a high to a low degree of diffusive limitation shifted the system from denitrification, with significant release of nitrous oxide, to DNRA. Carefully imposed degrees of diffusive limitations are powerful tools to studying environmental processes. These results clearly reveal heterogeneity in the form of diffusive limitation can impact nitrate reduction processes. Our results also indicate that a simple management scenario that would allow retention of reactive nitrogen in soil (favouring DNRA over denitrification) would involve adequate soil mixing after addition of excess of electron donor substrate. In addition to a contribution to the primary literature, an exhaustive review was conducted on tools applicable to collecting geochemical data at the microscale in soil and sediment. The review examined their ability to provide spatially resolved data with microscale resolution, focusing on their performance characteristics, the degree of physical disruption they inflict on the system being studied, the potential for repeated measurements and the accessibility of the tools. In addition to providing an overview of existing tools, the review revealed that many parts of the microscale toolkit have become increasingly accessible and affordable. However work remains to be done to facilitate simultaneous measurements of multiple analytes, and to expand the array of potential analytes for the various techniques. In addition, during this PhD study, a nitrate sensitive planar optode was developed along with a tool for producing large, smooth planar optode sensor foil sheets. This is the first planar optode of its kind, and it exhibits a linear response to nitrate from 1 to 50 mM at pH 8.0, a fast response time of < 10sand good lifetime, allowing for fast two dimensional measurements of nitrate distributions over long periods of time. This new sensor technology allows dynamic two dimensional measurements of nitrate at microscale spatial resolution. Unfortunately, time did not permit to use the optode after its development.