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© 2016 Springer-Verlag Berlin Heidelberg We compared modifications of soil zymography, a new in situ technique to visualize enzyme activities, based on contact of fluorgenic substrate-saturated membranes with soil either through the gel layer (gel zymography) or without gel application (direct zymography). We coupled zymography with quantitative measurements of enzyme kinetics to characterize catalytic mechanisms of β-glucosidase activity at the plant-soil interface including root surface (rhizoplane), rhizosphere, and bulk soil. Direct zymography refined and focused image resolution. The area of hotspots (i.e., spots with most intensive enzyme activity) as well as color intensity ratios estimated using direct zymography exceeded by a factor of 2 the corresponding values obtained with gel zymography. As determined by direct zymography, the percentage of hotspots associated to root surfaces was 58–68 % of total hotspot area. Hotspot area comprised only 6.8 ± 0.1 % of the total area of an image and 9.0 ± 3 % of the root surface area. The intensity of β-glucosidase activity, however, was up to 20 times higher in the hotspots versus bulk soil. The contribution of rhizosphere to β-glucosidase activity of the whole image (77–82 %) was four times higher than the contribution of the root surface. Enzyme kinetic parameters indicated different enzyme systems in bulk and rhizosphere soil. Higher substrate affinity and catalytic efficiency in bulk than in rhizosphere soil suggested relative domination of microorganisms with more efficient enzyme systems in the former. Coupling direct zymography and kinetic assays enabled mapping the two-dimensional (2D) distribution of enzyme activity at the root-soil interface and estimating the catalytic properties of root-associated and soil-associated enzymes.

© 2016, Springer-Verlag Berlin Heidelberg.We compared modifications of soil zymography, a new in situ technique to visualize enzyme activities, based on contact of fluorgenic substrate-saturated membranes with soil either through the gel layer (gel zymography) or without gel application (direct zymography). We coupled zymography with quantitative measurements of enzyme kinetics to characterize catalytic mechanisms of β-glucosidase activity at the plant-soil interface including root surface (rhizoplane), rhizosphere, and bulk soil. Direct zymography refined and focused image resolution. The area of hotspots (i.e., spots with most intensive enzyme activity) as well as color intensity ratios estimated using direct zymography exceeded by a factor of 2 the corresponding values obtained with gel zymography. As determined by direct zymography, the percentage of hotspots associated to root surfaces was 58–68 % of total hotspot area. Hotspot area comprised only 6.8 ± 0.1 % of the total area of an image and 9.0 ± 3 % of the root surface area. The intensity of β-glucosidase activity, however, was up to 20 times higher in the hotspots versus bulk soil. The contribution of rhizosphere to β-glucosidase activity of the whole image (77–82 %) was four times higher than the contribution of the root surface. Enzyme kinetic parameters indicated different enzyme systems in bulk and rhizosphere soil. Higher substrate affinity and catalytic efficiency in bulk than in rhizosphere soil suggested relative domination of microorganisms with more efficient enzyme systems in the former. Coupling direct zymography and kinetic assays enabled mapping the two-dimensional (2D) distribution of enzyme activity at the root-soil interface and estimating the catalytic properties of root-associated and soil-associated enzymes.

© 2016 Springer-Verlag Berlin Heidelberg We compared modifications of soil zymography, a new in situ technique to visualize enzyme activities, based on contact of fluorgenic substrate-saturated membranes with soil either through the gel layer (gel zymography) or without gel application (direct zymography). We coupled zymography with quantitative measurements of enzyme kinetics to characterize catalytic mechanisms of β-glucosidase activity at the plant-soil interface including root surface (rhizoplane), rhizosphere, and bulk soil. Direct zymography refined and focused image resolution. The area of hotspots (i.e., spots with most intensive enzyme activity) as well as color intensity ratios estimated using direct zymography exceeded by a factor of 2 the corresponding values obtained with gel zymography. As determined by direct zymography, the percentage of hotspots associated to root surfaces was 58–68 % of total hotspot area. Hotspot area comprised only 6.8 ± 0.1 % of the total area of an image and 9.0 ± 3 % of the root surface area. The intensity of β-glucosidase activity, however, was up to 20 times higher in the hotspots versus bulk soil. The contribution of rhizosphere to β-glucosidase activity of the whole image (77–82 %) was four times higher than the contribution of the root surface. Enzyme kinetic parameters indicated different enzyme systems in bulk and rhizosphere soil. Higher substrate affinity and catalytic efficiency in bulk than in rhizosphere soil suggested relative domination of microorganisms with more efficient enzyme systems in the former. Coupling direct zymography and kinetic assays enabled mapping the two-dimensional (2D) distribution of enzyme activity at the root-soil interface and estimating the catalytic properties of root-associated and soil-associated enzymes.

© The Author(s) 2017. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. Background and Aims Although it is commonly accepted that root exudation enhances plant-microbial interactions in the rhizosphere, experimental data on the spatial distribution of exudates are scarce. Our hypothesis was that root hairs exude organic substances to enlarge the rhizosphere farther from the root surface. Methods Barley (Hordeum vulgare 'Pallas' - wild type) and its root-hairless mutant (brb) were grown in rhizoboxes and labelled with 14CO2. A filter paper was placed on the soil surface to capture, image and quantify root exudates. Key Results Plants with root hairs allocated more carbon (C) to roots (wild type: 13 %; brb: 8 % of assimilated 14C) and to rhizosheaths (wild type: 1.2 %; brb: 0.2 %), while hairless plants allocated more C to shoots (wild type: 65 %; brb: 75 %). Root hairs increased the radial rhizosphere extension three-fold, from 0.5 to 1.5 mm. Total exudation on filter paper was three times greater for wild type plants compared to the hairless mutant. Conclusion Root hairs increase exudation and spatial rhizosphere extension, which probably enhance rhizosphere interactions and nutrient cycling in larger soil volumes. Root hairs may therefore be beneficial to plants under nutrient-limiting conditions. The greater C allocation below ground in the presence of root hairs may additionally foster C sequestration.