Visualization of root extracellular traps in an ectomycorrhizal woody plant (Pinus densiflora) and their interactions with root-associated bacteria.

MAIN CONCLUSION: Extracellular traps in the primary root of Pinus densiflora contribute to root-associated bacterial colonization. Trapped rhizobacteria induce the production of reactive oxygen species in root-associated, cap-derived cells. Ectomycorrhizal (ECM) woody plants, such as members of Pinaceae and Fagaceae, can acquire resistance to biotic and abiotic stresses through the formation of mycorrhiza with ECM fungi. However, germinated tree seedlings do not have mycorrhizae and it takes several weeks for ectomycorrhizae to form on their root tips. Therefore, to confer protection during the early growth stage, bare primary roots require defense mechanisms other than mycorrhization. Here, we attempted to visualize root extracellular traps (RETs), an innate root defense mechanism, in the primary root of Pinus densiflora and investigate the interactions with root-associated bacteria isolated from ECM and fine non-mycorrhizal roots. Histological and histochemical imaging and colony-forming unit assays demonstrated that RETs in P. densiflora, mainly consisting of root-associated, cap-derived cells (AC-DCs) and large amounts of root mucilage, promote bacterial colonization in the rhizosphere, despite also having bactericidal activity via extracellular DNA. Four rhizobacterial strains retarded the mycelial growth of a pathogenic strain belonging to the Fusarium oxysporum species complex in dual culture assay. They also induced the production of reactive oxygen species (ROS) from host tree AC-DCs without being excluded from the rhizosphere of P. densiflora. Applying three Paraburkholderia strains, especially PM O-EM8 and PF T-NM22, showed significant differences in the ROS levels from the control group. These results reveal the indirect contributions of rhizobacteria to host root defense and suggest that root-associated bacteria could be a component of RETs as a first line of defense against root pathogens in the early growth stage of ECM woody plants.

Purification effects show seed and root mucilage's ability to respond to changing rhizosphere conditions.

Mucilage, a polysaccharide-containing hydrogel, is hypothesized to play a key role in the rhizosphere as a self-organized system because it may vary its supramolecular structure with changes in the surrounding solution. However, there is currently limited research on how these changes are reflected in the physical properties of real mucilage. This study examines the role of solutes in maize root, wheat root, chia seed, and flax seed mucilage in relation to their physical properties. Two purification methods, dialysis and ethanol precipitation, were applied to determine the purification yield, cation content, pH, electrical conductivity, surface tension, viscosity, transverse 1 H relaxation time, and contact angle after drying of mucilage before and after purification. The two seed mucilage types contain more polar polymers that are connected to larger assemblies via multivalent cation crosslinks, resulting in a denser network. This is reflected in higher viscosity and water retention ability compared to root mucilage. Seed mucilage also contains fewer surfactants, making them better wettable after drying compared to the two root mucilage types. The root mucilage types, on the other hand, contain smaller polymers or polymer assemblies and become less wettable after drying. However, wettability not only depends on the amount of surfactants but also on their mobility, as well as the strength and mesh size of the network structure. The changes in physical properties and cation composition observed after ethanol precipitation and dialysis suggest that the polymer network of seed mucilage is more stable and specialized in protecting the seeds from unfavorable environmental conditions. In contrast, root mucilage is characterized by fewer cationic interactions and its network relies more on hydrophobic interactions. This allows root mucilage to be more flexible in responding to changing environmental conditions, facilitating nutrient and water exchange between root surfaces and the rhizosphere soil.

Root hairs are the most important root trait for rhizosheath formation of barley (Hordeum vulgare), maize (Zea mays) and Lotus japonicus (Gifu).

BACKGROUND AND AIMS: Rhizosheaths are defined as the soil adhering to the root system after it is extracted from the ground. Root hairs and mucilage (root exudates) are key root traits involved in rhizosheath formation, but to better understand the mechanisms involved their relative contributions should be distinguished. METHODS: The ability of three species [barley (Hordeum vulgare), maize (Zea mays) and Lotus japonicus (Gifu)] to form a rhizosheath in a sandy loam soil was compared with that of their root-hairless mutants [bald root barley (brb), maize root hairless 3 (rth3) and root hairless 1 (Ljrhl1)]. Root hair traits (length and density) of wild-type (WT) barley and maize were compared along with exudate adhesiveness of both barley and maize genotypes. Furthermore, root hair traits and exudate adhesiveness from different root types (axile versus lateral) were compared within the cereal species. KEY RESULTS: Per unit root length, rhizosheath size diminished in the order of barley > L. japonicus > maize in WT plants. Root hairs significantly increased rhizosheath formation of all species (3.9-, 3.2- and 1.8-fold for barley, L. japonicus and maize, respectively) but there was no consistent genotypic effect on exudate adhesiveness in the cereals. While brb exudates were more and rth3 exudates were less adhesive than their respective WTs, maize rth3 bound more soil than barley brb. Although both maize genotypes produced significantly more adhesive exudate than the barley genotypes, root hair development of WT barley was more extensive than that of WT maize. Thus, the greater density of longer root hairs in WT barley bound more soil than WT maize. Root type did not seem to affect rhizosheath formation, unless these types differed in root length. CONCLUSIONS: When root hairs were present, greater root hair development better facilitated rhizosheath formation than root exudate adhesiveness. However, when root hairs were absent root exudate adhesiveness was a more dominant trait.

The wheat secreted root proteome: Implications for phosphorus mobilisation and biotic interactions.

Root secreted acid phosphatases and organic anions are widely perceived as major players of plant phosphorus (P) mobilisation from the rhizosphere under P limiting growth conditions. Previous research indicated that other mechanisms play a role, especially in species with fine roots, such as wheat. In this study we characterised the plant-derived extracellular proteome of wheat roots by profiling root tip mucilage, soluble root secreted and root tip proteomes. Extracellular acid phosphatases and enzymes of the central carbon metabolism were targeted using selected reaction monitoring. More than 140 proteins with extracellular localisation prediction were identified in mucilage. P starvation induced proteins predicted to be localised to the apoplast which are related to cell wall modification and defence in both, root tip and soluble root-secreted proteomes. Glycolytic enzymes were strongly increased in abundance by P limitation in root tips, as were PEPC and plastidial MDH. Soluble acid phosphatases were not identified in extracellular protein samples. Our results indicate that root tip mucilage contains proteins with the functional potential to actively shape their immediate environment by modification of plant structural components and biotic interactions. Wheat acid phosphatases appear to play a minor role in P mobilisation beyond the immediate root surface. SIGNIFICANCE: Phosphorus (P) is a plant growth limiting nutrient in many agricultural situations and the development of phosphorus efficient crops is of paramount importance for future agricultural management practices. As P is relatively immobile in soils, processes occurring at the root-soil interface, the rhizosphere, are suspected to play a key role in plant-induced P mobilisation. According to the current view, the secretion of extracellular acid phosphatases and organic anions enhances P mobilisation within several millimetres beyond the root surface, either directly or indirectly through the selection and appropriate soil microbes. However, the mechanisms of P mobilisation in species with fine roots, such as wheat, and the role of other secreted root proteins are poorly understood. Here, we carried out the profiling of wheat root tip mucilage, soluble root secreted and root tip proteomes. We analysed proteome changes in response to P starvation. We found that proteins with a predicted localisation to the apoplast made up a major proportion of stress-responsive proteins. Acid phosphatases were not identified within extracellular protein samples, which were enriched in proteins with predicted extracellular localisation. The absence of extracellular APases was further validated by multiple reaction monitoring. Our data indicates that wheat acid phosphatases play a minor role in P mobilisation beyond the immediate root surface and provides a resource for breeding strategies and further investigations of the functional roles of root tip-released proteins in the rhizosphere under P limitation.

Links between nanoscale and macroscale surface properties of natural root mucilage studied by atomic force microscopy and contact angle.

Soil water repellency originating from organic coatings plays a crucial role for soil hydraulics and plant water uptake. Focussing on hydrophobicity in the rhizosphere induced by root-mucilage, this study aims to explore the link between macroscopic wettability and nano-microscopic surface properties. The existing knowledge of the nanostructures of organic soil compounds and its effect on wettability is limited by the lack of a method capable to assess the natural spatial heterogeneity of physical and chemical properties. In this contribution, this task is tackled by a geostatistical approach via variogram analysis of topography and adhesion force data acquired by atomic force microscopy and macroscopic sessile drop measurements on dried films of mucilage. The results are discussed following the wetting models given by Wenzel and Cassie-Baxter. Undiluted mucilage formed homogeneous films on the substrate with contact angles >90°. For diluted samples contact angles were smaller and incomplete mucilage surface coverage with hole-like structures frequently exhibited increased adhesion forces. Break-free distances of force curves indicated enhanced capillary forces due to adsorbed water films at atmospheric RH (35 ±â€¯2%) that promote wettability. Variogram analysis enabled a description of complex surface structures exceeding the capability of comparative visual inspection.

The epidermal surface of the maize root tip: I. Development in normal roots.

The surface of the meristematic epidermis of maize roots is tri-partite. A helicoidal primary wall follows the contours of the tops of the columnar epidermal cells and is continuous with their buttressed anticlinal walls. Two overlying layers form a smooth covering over the root which obscures the cell outlines. This compound surface is similar architecturally to outer epidermal surfaces of shoots. The two outer layers are distinct structurally and in their staining properties from the wall and are together here referred to as the pellicle. Both pellicle layers are fibrillar but not helicoidal. Their development begins in the boundary between the cap and the root proper and they reach maximum thickness over the meristematic region. The outer layer then disintegrates and is absent from the elongation zone. The inner layer thins irregularly as the columnar cells elongate to their final tabular form and usually persists only over the groove above anticlinal walls and at the base of root hairs. The cell wall thins to about half its maximum thickness during this elongation. Emerging root hairs broach the pellicle and the original primary wall. Remnants of both these layers form a short, tight collar at the base of each hair; this collar adheres to the primary wall of the hair which is continuous with a new, thin wall which is formed interior to the original outer wall of the parent cell. Failure to recognize the complex structure and transitory nature of the epidermal pellicle has led to confusion in the literature regarding the nature of root-surface and rhizosphere mucilaginous components and their origin. These interpretations are compared with those arising from this study.

Plant roots release phospholipid surfactants that modify the physical and chemical properties of soil.

• Plant root mucilages contain powerful surfactants that will alter the interaction of soil solids with water and ions, and the rates of microbial processes. • The lipid composition of maize, lupin and wheat root mucilages was analysed by thin layer chromatography and gas chromatography-mass spectrometry. A commercially available phosphatidylcholine (lecithin), chemically similar to the phospholipid surfactants identified in the mucilages, was then used to evaluate its effects on selected soil properties. • The lipids found in the mucilages were principally phosphatidylcholines, composed mainly of saturated fatty acids, in contrast to the lipids extracted from root tissues. In soil at low tension, lecithin reduced the water content at any particular tension by as much as 10 and 50% in soil and acid-washed sand, respectively. Lecithin decreased the amount of phosphate adsorption in soil and increased the phosphate concentration in solution by 10%. The surfactant also reduced net rates of ammonium consumption and nitrate production in soil. • These experiments provide the first evidence we are aware of that plant-released surfactants will significantly modify the biophysical environment of the rhizosphere.

Root mucilage enhances aluminum accumulation in Melastoma malabathricum, an aluminum accumulator.

Root mucilage is gelatinous polysaccharide-containing material exuded from the outer layers of the root cap. Although mucilage has been suggested to play several roles in plant growth, its role in mineral uptake has not been well understood. Melastoma malabathricum L. is an aluminum (Al) accumulator growing in tropical acid soils. This species accumulates more than 10 mg Al g(-1) DW in leaves and roots. Root mucilage is generally known to immobilize metal cations such as Al in the rhizosphere. However, we found that roots of M. malabathricum exuded large amounts of mucilage. Using the Zea mays L. mucilage as a control, we have recently shown that mucilage of M. malabathricum has unique physical and chemical characteristics, and facilitates Al uptake in this species. Since M. malabathricum cannot grow well in Al-deficient soil (nonacid soils), this species might have developed a mechanism for Al acquisition. We have also discussed the reason for this species' requirement of Al, a nonessential element.