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Microorganisms interact with different soil components, such as varying substrates and soil minerals, to drive soil processes and functionality. They can be influenced by the environment, but they themselves can influence their environment by their activities, for example through the production of extracellular enzymes and extracellular polymeric substances (EPS). The formation and stability of aggregates as the backbone of the soil structure, for instance, are thought to be largely influenced by soil microorganisms, or vice versa. There remain, however, open questions as to whether and how microorganisms can influence soil aggregation. While microbes are influencing their environment their interaction with the soil minerals could also change their responses upon adsorption - affecting their influence on soil aggregation. Therefore, the overarching goal of this thesis was to investigate the effect of soil minerals, in particular clay content, on the composition and activity of soil microbial community, with a specific focus on enzyme activities and EPS. Finally, the microbial control of soil aggregation through the influence of substrate availability was explored. In total, two adsorption experiments and two incubation experiments were conducted using soils manipulated experimentally with increasing clay content. The sandy soil was amended with different amounts of soil minerals (i.e. montmorillonite) to achieve a gradient in clay content. For the first incubation experiment, organic substrates differing in decomposability (i.e., starch and cellulose) were added to the soil to stimulate microbial activities and incubated for 80 days. Soil samples from the first incubation experiment were analysed after 0, 3, 10, 20, 40 and 80 days for enzyme activities, microbial community composition, biomass C, EPS-protein and polysaccharide. Additionally, the geometric mean diameter and mean weight diameter of the soil aggregates were determined as measures of aggregate formation

Microorganisms interact with different soil components, such as varying substrates and soil minerals, to drive soil processes and functionality. They can be influenced by the environment, but they themselves can influence their environment by their activities, for example through the production of extracellular enzymes and extracellular polymeric substances (EPS). The formation and stability of aggregates as the backbone of the soil structure, for instance, are thought to be largely influenced by soil microorganisms, or vice versa. There remain, however, open questions as to whether and how microorganisms can influence soil aggregation. While microbes are influencing their environment their interaction with the soil minerals could also change their responses upon adsorption - affecting their influence on soil aggregation. Therefore, the overarching goal of this thesis was to investigate the effect of soil minerals, in particular clay content, on the composition and activity of soil microbial community, with a specific focus on enzyme activities and EPS. Finally, the microbial control of soil aggregation through the influence of substrate availability was explored. In total, two adsorption experiments and two incubation experiments were conducted using soils manipulated experimentally with increasing clay content. The sandy soil was amended with different amounts of soil minerals (i.e. montmorillonite) to achieve a gradient in clay content. For the first incubation experiment, organic substrates differing in decomposability (i.e., starch and cellulose) were added to the soil to stimulate microbial activities and incubated for 80 days. Soil samples from the first incubation experiment were analysed after 0, 3, 10, 20, 40 and 80 days for enzyme activities, microbial community composition, biomass C, EPS-protein and polysaccharide. Additionally, the geometric mean diameter and mean weight diameter of the soil aggregates were determined as measures of aggregate formation

Nitrification is the microbial conversion of reduced forms of nitrogen (N) to nitrate (NO3-), and in fertilized soils it can lead to substantial N losses via NO3- leaching or nitrous oxide (N2O) production. To limit such problems, synthetic nitrification inhibitors have been applied but their performance differs between soils. In recent years, there has been an increasing interest in the occurrence of biological nitrification inhibition (BNI), a natural phenomenon according to which certain plants can inhibit nitrification through the release of active compounds in root exudates. Here, we synthesize the current state of research but also unravel knowledge gaps in the field. The nitrification process is discussed considering recent discoveries in genomics, biochemistry and ecology of nitrifiers. Secondly, we focus on the 'where' and 'how' of BNI. The N transformations and their interconnections as they occur in, and are affected by, the rhizosphere, are also discussed. The NH4+ and NO3- retention pathways alternative to BNI are reviewed as well. We also provide hypotheses on how plant compounds with putative BNI ability can reach their targets inside the cell and inhibit ammonia oxidation. Finally, we discuss a set of techniques that can be successfully applied to solve unresearched questions in BNI studies.

Nitrification is the microbial conversion of reduced forms of nitrogen (N) to nitrate (NO3−), and in fertilized soils it can lead to substantial N losses via NO3− leaching or nitrous oxide (N2O) production. To limit such problems, synthetic nitrification inhibitors have been applied but their performance differs between soils. In recent years, there has been an increasing interest in the occurrence of biological nitrification inhibition (BNI), a natural phenomenon according to which certain plants can inhibit nitrification through the release of active compounds in root exudates. Here, we synthesize the current state of research but also unravel knowledge gaps in the field. The nitrification process is discussed considering recent discoveries in genomics, biochemistry and ecology of nitrifiers. Secondly, we focus on the ‘where’ and ‘how’ of BNI. The N transformations and their interconnections as they occur in, and are affected by, the rhizosphere, are also discussed. The NH4+ and NO3− retention pathways alternative to BNI are reviewed as well. We also provide hypotheses on how plant compounds with putative BNI ability can reach their targets inside the cell and inhibit ammonia oxidation. Finally, we discuss a set of techniques that can be successfully applied to solve unresearched questions in BNI studies.

Nitrification is the microbial conversion of reduced forms of nitrogen (N) to nitrate (NO3−), and in fertilized soils it can lead to substantial N losses via NO3− leaching or nitrous oxide (N2O) production. To limit such problems, synthetic nitrification inhibitors have been applied but their performance differs between soils. In recent years, there has been an increasing interest in the occurrence of biological nitrification inhibition (BNI), a natural phenomenon according to which certain plants can inhibit nitrification through the release of active compounds in root exudates. Here, we synthesize the current state of research but also unravel knowledge gaps in the field. The nitrification process is discussed considering recent discoveries in genomics, biochemistry and ecology of nitrifiers. Secondly, we focus on the ‘where’ and ‘how’ of BNI. The N transformations and their interconnections as they occur in, and are affected by, the rhizosphere, are also discussed. The NH4+ and NO3− retention pathways alternative to BNI are reviewed as well. We also provide hypotheses on how plant compounds with putative BNI ability can reach their targets inside the cell and inhibit ammonia oxidation. Finally, we discuss a set of techniques that can be successfully applied to solve unresearched questions in BNI studies.

Stabilization of C in soil is important for minimizing greenhouse gas emission into the atmosphere and for improving the soil fertility. The physical, chemical, and biological properties of soil greatly influence the C protection capacity. Both the physical and chemical properties of soils are directly or indirectly governed by clay minerals, which are the most reactive soil particles. Apart from the amount of clays, clay types are also very important in protecting soil C. Clay minerals provide both permanent and variable surface charges and different specific surface areas that are crucial to determine the C protection capacity of soils. They form organo–mineral complexes, promote aggregate establishment, and protect soil organic matter against microbial decomposition. This chapter aims to provide an overview of the role of various clay minerals in protecting organic C in soils and highlights the mechanisms of C sequestration by clay minerals.

Stabilization of C in soil is important for minimizing greenhouse gas emission into the atmosphere and for improving the soil fertility. The physical, chemical, and biological properties of soil greatly influence the C protection capacity. Both the physical and chemical properties of soils are directly or indirectly governed by clay minerals, which are the most reactive soil particles. Apart from the amount of clays, clay types are also very important in protecting soil C. Clay minerals provide both permanent and variable surface charges and different specific surface areas that are crucial to determine the C protection capacity of soils. They form organo–mineral complexes, promote aggregate establishment, and protect soil organic matter against microbial decomposition. This chapter aims to provide an overview of the role of various clay minerals in protecting organic C in soils and highlights the mechanisms of C sequestration by clay minerals.

Direct extraction and characterization of microbial community DNA through PCR amplicon surveys and metagenomics has revolutionized the study of environmental microbiology and microbial ecology. In particular, metagenomic analysis of nucleic acids provides direct access to the genomes of the “uncultivated majority.” Accelerated by advances in sequencing technology, microbiologists have discovered more novel phyla, classes, genera, and genes from microorganisms in the first decade and a half of the twenty-first century than since these “many very little living animalcules” were first discovered by van Leeuwenhoek.

This study was carried out to understand the interplay of plant Nitrogen Utilizing Efficiency (NUE) with protease activity and microbial proteolytic community composition in the rhizosphere and bulk soils. Protease activity, diversity and abundance of protease genes (using DGGE and qPCR respectively of two key bacterial protease encoding genes: alkaline metallo-peptidase (apr) and neutral-metallopeptidases (npr)) were monitored in both rhizosphere and bulk soils from two maize in-bred lines L05 and T250 with higher and lower NUE respectively, using a rhizobox approach. Illumina sequencing was employed to assess the diversity of proteolytic communities encoding for the above-mentioned protease genes. Our results show higher enzyme activity, higher abundance and diversity of proteolytic genes in L05 maize rhizosphere, with higher NUE than in T250 maize rhizosphere.

Rhizospheric microbial composition plays an important role in plant growth and nutrient metabolism. Under the effect of root exudates, soil in rhizosphere is different from bulk soil in its microbial composition and functions from bulk soil. Effect of root exudates on microbial communities had been investigated by various researchers, still leaving a gap in study of effect on functions related to soil. One such important soil function is proteolysis. Proteases and peptidases are important enzymes that bring about N mineralization and in turn N uptake to facilitate plant growth and development. Understanding of the relationship between soil microbial communities expressing protease activity and nitrogen in the rhizosphere is still a challenge, due to the difficulties of sampling the rhizosphere microenvironment. This study investigated the interplay of plant nitrogen utilizing efficiency with N mineralization brought about by proteolysis with special focus on bacterial extracellular soil proteases. We studied changes in the biochemical activity and microbial community structure in the rhizosphere of the inbred maize (Zea mais L.) lines Lo5 and T250 characterized by high and low Nitrogen utilizing effeciencies (NUE) using rhizobox experiments. Plants were regularly regularly monitored for the inorganic N (NH4+-N and NO3--N) concentration in the rhizosphere by ion selective electrodes (ISE). At the stage of N depletion soil was sampled from rhizosphere and bulk compartments. Cellular biomass was estimated as a measure of ATP and protease activity was assayed as a measure on caseinate hydrolysis. Soil DNA was extracted and was used for molecular studies. Two bacterial genes coding for alkaline proteases (apr) and neutral protease (npr) were selected for studies. Bacterial gene abundance was measured using qPCR. To study diversity of these genes, amplicons were sequenced by Illumina high-throughput technology. Plant L05 with higher NUE, had shown a significant difference

Soil as a biological system is characterized by: i) the presence of a remarkable diversity since thousands of bacterial genomes can be present in one gram of soil. In addition microbial biomass is huge; ii) only a minor proportion of the available space is occupied by microorganisms in soil (microbiological space); iii) soil colloids can adsorb important biological molecules such as proteins and nucleic acids. Nucleic acids can adsorbed and retain their biological activity; iv). soil components show enzyme-like activities. Unfortunately there is no methods to distinguish enzyme from enzyme-like reactions but these methods are needed to quantify both contributions; v) virus are more abundant than in other systems such as aquatic ones. A book “Omics in Soil Science” (Nannipieri et al 2014) has been recently published; it presents the state-of-the-art of omics in soil science, a field that is advancing rapidly on many fronts. The various omics (mainly metagenomics, metatranscriptomics, proteomics and proteogenomics) approaches hold much promise but also await further refinement before they are ready for widespread adaptation. One way to judge their readiness is to compare them to methods that have become standards for soil microbiology research. Methods become standards because they provide useful information quickly and inexpensively. There is no question that omics can provide useful information, some of which cannot be obtained with traditional techniques, and integration of omics methods may provide insights into ecosystem functioning. In particular, the potential for omics to provide comprehensive coverage of genes and genes products make them well-suited for the study of general soil microbiological phenomena, such as decomposition, response to water stress

Soil as a biological system is characterized by: i) the presence of a remarkable diversity since thousands of bacterial genomes can be present in one gram of soil. In addition microbial biomass is huge; ii) only a minor proportion of the available space is occupied by microorganisms in soil (microbiological space); iii) soil colloids can adsorb important biological molecules such as proteins and nucleic acids. Nucleic acids can adsorbed and retain their biological activity; iv). soil components show enzyme-like activities. Unfortunately there is no methods to distinguish enzyme from enzyme-like reactions but these methods are needed to quantify both contributions; v) virus are more abundant than in other systems such as aquatic ones. A book “Omics in Soil Science” (Nannipieri et al 2014) has been recently published; it presents the state-of-the-art of omics in soil science, a field that is advancing rapidly on many fronts. The various omics (mainly metagenomics, metatranscriptomics, proteomics and proteogenomics) approaches hold much promise but also await further refinement before they are ready for widespread adaptation. One way to judge their readiness is to compare them to methods that have become standards for soil microbiology research. Methods become standards because they provide useful information quickly and inexpensively. There is no question that omics can provide useful information, some of which cannot be obtained with traditional techniques, and integration of omics methods may provide insights into ecosystem functioning. In particular, the potential for omics to provide comprehensive coverage of genes and genes products make them well-suited for the study of general soil microbiological phenomena, such as decomposition, response to water stress

Both classical and molecular approaches in soil enzymology are discussed. The detection of genes encoding enzymes combined with measurements of the relative soil enzyme activity has given insights on the origin of the measured enzymes. It is also discussed the main findings and future research about assays, kinetics, use of enzyme activities as indicators of soil functions, and location, state and role of extracellular stabilized enzymes in soil. Most of these aspects have been debated in the extensive bibliography, which is nowadays sometime ignored, probably because published in books, which, particularly those published before 1990s, are not accessible by the electronic searches. It is still problematic to use enzyme activities as indicators of soil functions because: i) present enzyme assays determine potential and not real enzyme activities; ii) the meaning of measured enzyme activities is not known; iii) the biochemistry of the process is often neglected when the target enzyme activity is assumed be an indicator of the nutrient dynamics in soil; iv) spatio-temporal variations are not always considered in situ experiments; v) many direct and indirect effects make difficult the interpretation of the response of the enzyme activity to perturbations, changes in the soil management, changes in the plant cover of soil, etc. However, combining measurements of enzyme activity in soil with expression (transcriptomics and proteomics) of genes encoding the relative enzymes may contribute to understanding the mode and timing of microbial communities responses to the substrate availability and the persistence and the stabilization of enzymes in the soil, and detecting the loci of enzyme production.