Bacterial succession and bio-induced carbonate precipitation following willow growth

Different species of willow (Salix sp.) were historically used to improve slope stability and as the first step in afforestation on sandy soils. Willows can be grown from cuttings that harbour microbial symbionts (bacteria, ectomycorrhizal, and arbuscular mycorrhizal fungi) and promote successful plant establishment.

In an indoor pot experiment, we studied successional changes in bacterial and fungal communities following planting of willow cuttings in clean sand. Physical, chemical and microbiological parameters of root-affected sand were monitored during the growth of willows. Samples were taken from different pots that were disassembled after 30, 60, 90, 150, and 180 days of growth. Sand aggregation around roots was observed in pots sampled at days 150 and 180. Sequencing of gene libraries and qPCR for bacterial 16S rRNA and fungal ITS2 marker genes were performed on DNA extracted from loose root-affected sand and sand aggregates.The concentrations of soil total and water-extractable organic carbon and several metals (Ca, Mg, Mn, Cr, Cu, Fe, and Zn) were measured by ICP-OES and ICP-MS.

Concentration of water-extractable organic carbon in root-affected sand increased twofold from day 30 to day 150. Bacterial communities exhibited a clear pattern of increasing gene abundance and alpha-diversity over time. Proteobacteria were dominant in all samples, while Bacteroidota, Planctomycetota, Chloroflexota abundances increased in samples of day 150. However, no clear trends could be observed for the taxonomic structure of fungal communities, as the distribution of fungal dominants in sand samples was more scattered.

Sand aggregates differed from loose sand samples in terms of Ca and Mg concentration (40-200-fold higher), as well as the number of fungal and bacterial marker genes. Bacterial communities in sand aggregates were dominated by plant-associated bacteria, such as Sphingobium sp., while Firmicutes were significantly reduced, compared to those in loose sand. Some endophytic and plant-pathogenic fungi were found in aggregates. SEM analysis showed that crystals were formed between aggregated sand grains. So presumably, calcium carbonate precipitated due to root respiration affecting local conditions, while bacterial biofilms on sand grains acted as crystallization points. Alternatively, oxalic acid could have been exuded by willows and either precipitated as calcium oxalate or converted by oxalotrophic bacteria. Bio-induced precipitation of carbonates around willow roots enhances slope stability and carbon sequestration.

Our findings show that the growth of willow cuttings may alter the properties of rooted sand not only through root reinforcement, but also through bio-induced sand aggregation that occurs after five months.

This work is part of the project Soil Is Alive (SoIA), which was granted by the Carlsberg Foundation as part of the Consolidator Excellence Grant, Semper Ardens: Accelerate.