Rhizosheath formation occurs when soil particles, organic matter, and root exudates accumulate around plant roots, forming a protective sheath. This phenomenon plays a central role in plant-root interactions, nutrient uptake, and soil stabilization. In this study, the development of rhizosheath was significantly enhanced under wetter conditions as compared to DW cycles. In natural environment, several reasons can account for this: firstly, ample moisture promotes enhanced root growth, leading to more root surface area and increased release of exudates (Abdul-Jabbar et al., 1982; Mackay & Barber, 1985). These exudates act as a binding agent, causing soil particles to adhere to the roots and facilitating rhizosheath formation (Aslam et al., 2022). However, in this study, this reason does not apply, as we exclusively used model roots for our experiments.
The structure of the rhizosheath also creates an ecological niche with favorable microclimatic conditions, which in turn promote the growth and development of microbial populations, including nitrogen-fixing bacteria (Bergmann et al., 2009; Othman et al., 2003), as well as the production of related microbial gluing agents (Davinic et al., 2012; Six et al., 2004). Furthermore, excessive moisture reduces the cohesion of soil particles, making them more likely to detach and adhere to the roots (Bates & Lynch, 2001), thereby further enhancing rhizosheath formation. Additionally, constant moisture conditions may lead to an enhanced diffusion of mucilage into the surrounding soil, thus increasing the soil volume participating in mucilage-induced rhizosheath formation (Rahim et al., under review).
The constant moisture minimizes soil drying and subsequent cracking, ensuring the continuous accumulation of the rhizosheath around the roots. In contrast, alternate DW cycles of drying and wetting, characterized by intermittent moisture fluctuations, negatively affect rhizosheath formation because soil cracking and shrinkage occur during drying, disrupting the continuity of the rhizosheath, and resulting in reduced accumulation around the roots. Furthermore, the movement of soil particles during drying and subsequent rewetting events can wash away or displace the accumulated rhizosheath. Finally, mucilage might coagulate during the drying event, thus not diffusing into the surrounding soil and reducing the amount of mucilage-attached particles for rhizosheath formation (Rahim et al., under review). Noteworthy, similar effects can also be expected in natural soil environment: during drying cycles, when soil moisture decreases, roots may experience water stress and partial desiccation. This will additionally hinder root growth and reduce exudate release (Hinsinger et al., 2009), thus contributing to the reduced formation of rhizosheath. The fluctuating moisture conditions likely also impact soil microbial communities involved in rhizosheath formation. Yet, different processes interact, namely the microbial production of additional glues for rhizosheath formation, as well as the decomposition of mucilage and other gluing agents needed for aggregation and rhizosheath formation. The role of DW cycles is hard to predict in this context. It has been shown that DW cycles mighty restrict the availability of available C in the soil, thus limiting microbial decomposition processes (Fierer & Schimel, 2002; Mikha et al., 2005; Sommers et al., 1981); however, it is also well known that rewetting of dried soil usually leads to an increased CO2 flush (Birch effect), due to enhanced microbial activity (Birch, 1958; Franzluebbers et al., 1994; S. Zhang et al., 2020). Yet, we can decipher the underlying processes as we studied the rhizosheath development in soils in both sterilized and unsterilized soil. In line with Zhang et al., (2020), who reported higher rhizosheath formation and root biomass in rice plants in unsterilized soils, we also found that rhizosheath formation was enhanced when sterilization was lacking. Apparently, the production of microbial gluing agents outcompeted their potential role on the decomposition of mucilage. As similar effects were not recorded for the soils that underwent DW cycles, it seems at least reasonable to assume that microbial effects were less prominent in these samples than in those with constant moisture supply.
We also found significant higher rhizosheath formation in unsterilized soil that had lower clay contents (22%). Rhizosheath formation constitutes a complex mechanism influenced by a multitude of biotic and abiotic factors. One pivotal factor contributing significantly to this process is soil texture. The SEM images revealed smaller aggregates, potentially enveloped by organic matter, in rhizosheath under wet conditions, particularly in unsterilized soil (22% clay content), characterized by a higher rhizosheath mass (see Fig. 1c). Conversely, in soils with a higher clay content (sterilized, 32%), we observe a diminished rhizosheath formation, corroborating our findings from SEM (see Fig. 1d). In the context of more clayey soils with reduced rhizosheath development, a single prominent aggregate predominates the image, underscoring the cohesive nature inherent to clayey soils and the substantial impact of abiotic factors on aggregate formation.
It is noteworthy that historical data have consistently indicated statistically significant instances of rhizosheath formation in sandy soils. Furthermore, in contrast to loamy and clayey soils, sandy soils, which have lower porosity and limited root-soil contact, typically exhibit the formation of larger rhizosheaths (Hallett et al., 2022). Similar results were reported by multiple studies where sandy soil depicted large and consolidated rhizosheaths (Bailey & Scholes, 1997; Leistner, 1967; Oppenheimer, 1960). These studies have indicated that roots grown in sandy soil tend to develop a greater number of epidermal hairs compared to roots of the same plant species grown in less sandy soil (Duell & Peacock, 1985). Here, however, only model roots were analyzed that did not develop additional root hairs. Nevertheless, our data confirm these findings that rhizosheath formation was facilitated at higher sand content. Two reasons might account for this: a) a better diffusion of mucilage into the surrounding soil when clay content was smaller, and b) less aggregation of the coarser surrounding soil facilitating attachment of particles to the (model) plant root.
To analyze the soil particle size distribution of the aggregate formed, we used laser diffraction. The median diameter of soil microaggregates was < 10 µm, in line with Krause et al. (2018). Yet, also some aggregates > 10 µm were found, possibly reflecting the presence of organic C in > 6.7 µm size fraction and inorganic size fractions like clay minerals and Fe/Al (oxyhydr)oxides in the form of silicate-oxide interaction (Churchman, 2018). The aggregates formed had higher stability when the soil contained more clay (Fig. 5, 1d). Clay particles have a high surface area and possess electrostatic charges that attract and bind soil particles together to aggregates, through cation bridging, and by interactions with organic gluing agents (Tisdall and Oades, 1982; Bronick and Lal, 2005; Totsche et al., 2018, Krause et al., 2019). Intriguingly, aggregate stability was also enhanced when the soils were sterilized (Fig. 4a). Less clayey soils typically have a coarser texture with larger sand and silt particles. These larger particles tend to form a variety of aggregates with more open structures (Tisdall & Oades, 1982). In contrast, clayey soils, with their fine-textured particles, are more prone to forming larger, compact aggregates due to their higher surface area and greater adhesion. This is further supported by the SEM images, where rhizosheath mass was higher in soils with lower clay content compared to soils with a higher clay content (see Fig. 1c and 1d). Indeed, the formation and stability of soil aggregates is influenced by an interplay between biotic factors (e.g., Extracellular Polymeric Substances - EPS and microorganisms) and abiotic factors (e.g., clay minerals, pedogenic oxides). As emphasized by Clarholm & Skyllberg (2013), the presence of EPS and microorganisms plays a pivotal role in the initial formation of aggregates owing to their adhesive properties. Nevertheless, when organic matter becomes depleted or in conditions of sterilization, aggregates may disintegrate due to the loss of the organic "glue" that binds them together (Siebers et al., 2023). This disintegration can lead to the emergence of smaller particles or aggregates that exhibit higher stability, primarily driven by the increased prevalence of abiotic components such as clay minerals and oxides. These abiotic factors, especially electrostatic interactions and cation-bridging, engender robust forces of attraction between soil particles, resulting in enhanced aggregate cohesion and overall stability (Brady et al., 2008; Rengasamy, 2006). The biotic and abiotic processes is crucial for maintaining soil structure and its capacity to provide vital ecosystem services. This phenomenon aligns with our findings that demonstrate a shift towards smaller particle sizes and concurrently increased stability under sterilization conditions (Fig. 5a).