Publication Details

Category Text Publication
Reference Category Journals
DOI 10.1007/s11104-022-05573-4
Licence creative commons licence
Title (Primary) Development of micro-zymography: Visualization of enzymatic activity at the microscopic scale for aggregates collected from the rhizosphere
Author Ghaderi, N.; Schmidt, H.; Schlüter, S.; Banfield, C.C.; Blagodatskaya, E.
Source Titel Plant and Soil
Year 2022
Department BOOEK; BOSYS
Volume 478
Page From 253
Page To 271
Language englisch
Topic T5 Future Landscapes
Supplements https://static-content.springer.com/esm/art%3A10.1007%2Fs11104-022-05573-4/MediaObjects/11104_2022_5573_MOESM1_ESM.docx
Keywords Membrane-free micro-zymography; Exo-enzymes; Fixative
Abstract Aims

Visualization of enzymatic activity links microbial functioning to localization in heterogeneous soil habitats. To assess enzymatic reactions in soil thin layer at the microscopic level, we developed a micro-zymography approach and tested it by visualization of the potential activity of phosphomonoesterase for aggregates collected from the rhizosphere of Zea mays L.

Methods

We evaluated micro-zymography by applying fluorogenically-labeled substrate i) on individual soil aggregates freshly sampled from the rhizosphere, ii) on thin layers of aggregates (≈ 500 µm) saturated with substrate to assess the dynamics of phosphomonoesterase activity, and iii) on maize roots under laser scanning microscope upon the identified hotspots by membrane-based zymography.

Results

We found super transparent silicon as the most appropriate fixative to prevent sample drying. We optimized microscope settings to eliminate the soil auto-fluorescence. The fluorescent signal shifted from the free liquid phase towards the aggregate boundaries within 30 min after substrate addition and was finally detectable at the surface of a few aggregates. This was probably due to higher microbial abundance and enzymatic activity on the soil aggregates compared to the liquid phase. The enzymatic activity appeared patchy at the aggregate and root surfaces indicating heterogeneous distribution of hotspots.

Conclusions

The methodology including calibration, sample preparation, fixation, and monitoring was developed. The novel membrane-free micro-zymography approach is a promising tool to identify functional specificity and niche differentiation on roots and soil aggregates. This approach revealed unexplained complexity of competing processes (biochemical, hydrolytic, and physical) due to differently charged reaction products and enzyme-clay complexes.

Persistent UFZ Identifier https://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=26271
Ghaderi, N., Schmidt, H., Schlüter, S., Banfield, C.C., Blagodatskaya, E. (2022):
Development of micro-zymography: Visualization of enzymatic activity at the microscopic scale for aggregates collected from the rhizosphere
Plant Soil 478 , 253 - 271 10.1007/s11104-022-05573-4