Towards to physiological status of soil microorganisms determined by RNA:dsDNA ratio

Despite soil microorganisms spend most of their lifetime in a state of dormancy, they are quickly activated by substrate input and easily switch to growth. As both the dsDNA- and RNA- contents increase during microbial growth, the RNA:dsDNA ratio reflects a promising predictor, whether the response of a microbial community to environmental changes is due to an increase in population (by dsDNA) or due to an increase in activity (by RNA). This prediction of the RNA:dsDNA ratio can be accomplished by the comparison of microbial incubation approaches with and without addition of easily available substrates. We exhibited the RNA:dsDNA ratios for four contrasting soil types during substrate-induced growth. Overall, after glucose addition, a strong increase of dsDNA and RNA contents were determined in most of the soil types during 72 h of incubation. Furthermore, we identified distinct temporal soil-specific RNA:dsDNA patterns. The dsDNA- and RNA-contents yielded 26–174 and 0.3–30 µg g-1 soil, respectively. The soil texture was strongly associated with the reduction of RNA recovery, by means of an exponential decrease of RNA-content with increasing clay content. The lower RNA recovery in virgin and arable Chernozem (>30%) compared to soil types with lower clay contents (<17% for Retisol, Luvisol and Calcisol) suggests, that the undercount of RNA yields in clayey soils biased the RNA:dsDNA ratio, and subsequently the physiological state of the microbial community is not adequately represented in soils with clay contents exceeding 30%

Organic Nutrients Induced Coupled C- and P-Cycling Enzyme Activities During Microbial Growth in Forest Soils

Besides environmental and soil physical drivers, the functional properties of microbial populations, i. e., growth rate, enzyme production, and maintenance requirements are dependent on the microbes' environment. The soil nutrition status and the quantity and quality of the substrate input, both infer different growth strategies of microorganisms. It is uncertain, how enzyme systems respond during the different phases of microbial growth and retardation in soil. The objective of this study was to uncover the changes of microbial functioning and their related enzyme systems in nutrient-poor and nutrient-rich beech forest soil during the phases of microbial growth. We determined microbial growth via kinetic approach by substrate-induced respiratory response of microorganisms, enabling the estimation of total, and growing biomass of the microbial community. To induce microbial growth we used glucose, while yeast extract simulated additional input of nutrients and factors indicating microbial residues (i.e., necromass compounds). Microbial growth on glucose showed a 12–18 h delay in associated enzyme activity increase or the absence of distinct activity responses (Vmax). β-glucosidase and chitinase (NAG) demonstrated clear differences of Vmax in time and between P-rich and P-poor soils. However, during microbial growth on glucose + yeast extract, the exponential increase in enzymatic activity was clearly stimulated accompanied by a delay of 8–12 h, smoothing the differences in nutrient-acquisition dynamics between the two soils. Furthermore, cross-correlation of β-glucosidase and acid phosphatase between the two sites demonstrated harmonized time constraints, which reflected the establishment of comparable and balanced enzymatic systems within the decomposition network

Is soluble protein mineralisation and protease activity in soil regulated by supply or demand?

Protein represents a major input of organic matter to soil and is an important source of carbon (C) and nitrogen (N) for microorganisms. Therefore, determining which soil properties influence protein mineralisation in soil is key to understanding and modelling soil C and N cycling. However, the effect of different soil properties on protein mineralisation, and especially the interactions between soil properties, are poorly understood. We investigated how topsoil and subsoil properties affect protein mineralisation along a grassland altitudinal (catena) sequence that contained a gradient in soil type and primary productivity. We devised a schematic diagram to test the key edaphic factors that may influence protein mineralisation in soil (e.g. pH, microbial biomass, inorganic and organic N availability, enzyme activity and sorption). We then measured the mineralisation rate of 14C-labelled soluble plant-derived protein and amino acids in soil over a two-month period. Correlation analysis was used to determine the associations between rates of protein mineralisation and soil properties. Contrary to expectation, we found that protein mineralisation rate was nearly as fast as for amino acid turnover. We ascribe this rapid protein turnover to the low levels of protein used here, its soluble nature, a high degree of functional redundancy in the microbial community and microbial enzyme adaptation to their ecological niche. Unlike other key soil N processes (e.g. nitrification, denitrification), protease activity was not regulated by a small range of factors, but rather appeared to be affected by a wide range of interacting factors whose importance was dependent on altitude and soil depth [e.g. above-ground net primary productivity (NPP), soil pH, nitrate, cation exchange capacity (CEC), C:N ratio]. Based on our results, we hypothesise that differences in soil N cycling and the generation of ammonium are more related to the rate of protein supply rather than limitations in protease activity and protein turnover per se

Enzyme properties down the soil profile - A matter of substrate quality in rhizosphere and detritusphere

© 2016 Elsevier LtdThe decomposition of soil organic matter depends strongly on its availability to microorganisms and their enzymes. The rhizosphere and detritusphere are microbial hot spots due to additional substrate input, leading to high abundance, specific species diversity and functional diversity of microbial communities. However, rhizosphere and detritusphere differ in substrate quality, localization, and duration of input. We hypothesized that the contrasting substrate availability between rhizosphere and detritusphere affects the activity of microorganisms and associated enzymes. Organic carbon (C) from the rhizosphere and detritusphere decreases with soil depth and, consequently, microbial hot spots become rarer and competition for C and nutrients increases. In deeper soil (>40 cm depth) the amount and quality of substrates is expected to decrease and, therefore, the effect of contrasting substrate input to disappear. Plant N uptake is expected to reduce N availability in the rhizosphere of maize compared to the detritusphere and bare fallow. These hypotheses were tested in a factorial field experiment with 1) maize-planted, 2) maize litter-amended, and 3) bare sites. Enzyme kinetic parameters (Vmax, Km, Ka), extractable organic C and microbial biomass C were compared in soil affected by rhizosphere and detritusphere throughout the profile to 70 cm depth, to assess microbial C and nutrient limitations. A decrease in enzyme activity with depth due to resource scarcity and lower substrate quality appeared in planted and litter-amended soil. N limitation in planted soil increased the activity and substrate affinity of proteolytic enzymes to provide for microbial N demand through SOM decomposition. This was in line with lower Vmax ratios (Vmax for C-cycling enzymes divided by Vmax for N-cycling enzymes) in planted relative to litter-amended topsoil. The catalytic efficiency of enzymes decreased 2- to 20-fold from top- (40 cm), irrespective of the substrate input. Substrate quality in the rhizosphere and detritusphere affected enzyme activities only in the topsoil, whereas a sharp decline of C input with depth led to similar activities in the subsoil. Most of the enzyme indexes reflected shifts in allocation of C and nutrients in the rhizosphere and detritusphere. The presented results underline the role of microorganisms as critical links in the C and nutrient transfers in the rhizosphere and detritusphere

Enzyme properties down the soil profile - A matter of substrate quality in rhizosphere and detritusphere

© 2016 Elsevier LtdThe decomposition of soil organic matter depends strongly on its availability to microorganisms and their enzymes. The rhizosphere and detritusphere are microbial hot spots due to additional substrate input, leading to high abundance, specific species diversity and functional diversity of microbial communities. However, rhizosphere and detritusphere differ in substrate quality, localization, and duration of input. We hypothesized that the contrasting substrate availability between rhizosphere and detritusphere affects the activity of microorganisms and associated enzymes. Organic carbon (C) from the rhizosphere and detritusphere decreases with soil depth and, consequently, microbial hot spots become rarer and competition for C and nutrients increases. In deeper soil (>40 cm depth) the amount and quality of substrates is expected to decrease and, therefore, the effect of contrasting substrate input to disappear. Plant N uptake is expected to reduce N availability in the rhizosphere of maize compared to the detritusphere and bare fallow. These hypotheses were tested in a factorial field experiment with 1) maize-planted, 2) maize litter-amended, and 3) bare sites. Enzyme kinetic parameters (Vmax, Km, Ka), extractable organic C and microbial biomass C were compared in soil affected by rhizosphere and detritusphere throughout the profile to 70 cm depth, to assess microbial C and nutrient limitations. A decrease in enzyme activity with depth due to resource scarcity and lower substrate quality appeared in planted and litter-amended soil. N limitation in planted soil increased the activity and substrate affinity of proteolytic enzymes to provide for microbial N demand through SOM decomposition. This was in line with lower Vmax ratios (Vmax for C-cycling enzymes divided by Vmax for N-cycling enzymes) in planted relative to litter-amended topsoil. The catalytic efficiency of enzymes decreased 2- to 20-fold from top- (40 cm), irrespective of the substrate input. Substrate quality in the rhizosphere and detritusphere affected enzyme activities only in the topsoil, whereas a sharp decline of C input with depth led to similar activities in the subsoil. Most of the enzyme indexes reflected shifts in allocation of C and nutrients in the rhizosphere and detritusphere. The presented results underline the role of microorganisms as critical links in the C and nutrient transfers in the rhizosphere and detritusphere

Root-o-Mat: A novel tool for 2D image processing of root-soil interactions and its application in soil zymography

We developed a software tool enabling user-friendly and standardized pre- and post-processing of images of rooted soil by combining image processing techniques such as image registration, calibration, and segmentation in a graphical user interface. The added benefits of this image processing approach include an improved workflow in soil zymography. For evaluation, we conducted a rhizobox experiment with maize and determined the activity of leucine-aminopeptidase before and after glucose addition based on soil zymography. The temporal and spatial distribution of enzyme activity at the root-soil interface can be visualized by Root-o-Mat which offers 1) standardized image pre-processing, 2) calibration, 3) identification of hotspots of various intensity thresholds, 4) spatial analysis for selected roots, 5) inter-active illustration of enzyme activity profile lines, 6) image viewer, and 7) detection of temporal changes of enzyme activity. Registering images of the same rhizobox taken in successive periods allows further temporal and spatial analysis. We conclude that Root-o-Mat simplifies and firmly anchors image processing and image analyses in soil zymography. The new software can be downloaded for free (www.root-o-mat.de). © 2021 Elsevier Lt

Root-o-Mat: A novel tool for 2D image processing of root-soil interactions and its application in soil zymography

We developed a software tool enabling user-friendly and standardized pre- and post-processing of images of rooted soil by combining image processing techniques such as image registration, calibration, and segmentation in a graphical user interface. The added benefits of this image processing approach include an improved workflow in soil zymography. For evaluation, we conducted a rhizobox experiment with maize and determined the activity of leucine-aminopeptidase before and after glucose addition based on soil zymography. The temporal and spatial distribution of enzyme activity at the root-soil interface can be visualized by Root-o-Mat which offers 1) standardized image pre-processing, 2) calibration, 3) identification of hotspots of various intensity thresholds, 4) spatial analysis for selected roots, 5) inter-active illustration of enzyme activity profile lines, 6) image viewer, and 7) detection of temporal changes of enzyme activity. Registering images of the same rhizobox taken in successive periods allows further temporal and spatial analysis. We conclude that Root-o-Mat simplifies and firmly anchors image processing and image analyses in soil zymography. The new software can be downloaded for free (www.root-o-mat.de)

Organic Nutrients Induced Coupled C- and P-Cycling Enzyme Activities During Microbial Growth in Forest Soils

Besides environmental and soil physical drivers, the functional properties of microbial populations, i. e., growth rate, enzyme production, and maintenance requirements are dependent on the microbes' environment. The soil nutrition status and the quantity and quality of the substrate input, both infer different growth strategies of microorganisms. It is uncertain, how enzyme systems respond during the different phases of microbial growth and retardation in soil. The objective of this study was to uncover the changes of microbial functioning and their related enzyme systems in nutrient-poor and nutrient-rich beech forest soil during the phases of microbial growth. We determined microbial growth via kinetic approach by substrate-induced respiratory response of microorganisms, enabling the estimation of total, and growing biomass of the microbial community. To induce microbial growth we used glucose, while yeast extract simulated additional input of nutrients and factors indicating microbial residues (i.e., necromass compounds). Microbial growth on glucose showed a 12-18 h delay in associated enzyme activity increase or the absence of distinct activity responses (V-max). beta-glucosidase and chitinase (NAG) demonstrated clear differences of V(max)in time and between P-rich and P-poor soils. However, during microbial growth on glucose + yeast extract, the exponential increase in enzymatic activity was clearly stimulated accompanied by a delay of 8-12 h, smoothing the differences in nutrient-acquisition dynamics between the two soils. Furthermore, cross-correlation of beta-glucosidase and acid phosphatase between the two sites demonstrated harmonized time constraints, which reflected the establishment of comparable and balanced enzymatic systems within the decomposition network. The network accelerated nutrient acquisition to maintain microbial growth, irrespective of the contrasting soil properties and initial nutrient stocks, indicating similar tradeoffs of C- and P- cycling enzymes in both soils. This reflects comparable temporal dynamics of activities in nutrient-poor and nutrient-rich soil when the glucose + yeast extract was added. During lag phase and phase of exponential microbial growth, substrate turnover time of all enzymes was shortened in nutrient-poor forest soil exclusively, reflecting that the quality of the added substrate strongly matters during all stages of microbial growth in soil