Connecting structure and function from organisms to molecules in small-animal symbioses through chemo-histo-tomography

Our understanding of metabolic interactions between small symbi-otic animals and bacteria or parasitic eukaryotes that reside within their bodies is extremely limited. This gap in knowledge originates from a methodological challenge, namely to connect histologi -cal changes in host tissues induced by beneficial and parasitic (micro)organisms to the underlying metabolites. We addressed this challenge and developed chemo-histo-tomography (CHEMHIST), a culture-independent approach to connect anatomic structure and metabolic function in millimeter-sized symbiotic animals. CHEMHIST combines chemical imaging of metabolites based on mass spectrom-etry imaging (MSI) and microanatomy-based micro-computed X-ray tomography (micro-CT) on the same animal. Both high-resolution MSI and micro-CT allowed us to correlate the distribution of metab-olites to the same animal's three-dimensional (3D) histology down to submicrometer resolutions. Our protocol is compatible with tissue-specific DNA sequencing and fluorescence in situ hybridiza-tion for the taxonomic identification and localization of the associ-ated micro(organisms). Building CHEMHIST upon in situ imaging, we sampled an earthworm from its natural habitat and created an in-teractive 3D model of its physical and chemical interactions with bacteria and parasitic nematodes in its tissues. Combining MSI and micro-CT, we present a methodological groundwork for connecting metabolic and anatomic phenotypes of small symbiotic animals that often represent keystone species for ecosystem functioning

Plant Growth and Root Morphology Are Affected by Earthworm-Driven (Eisenia sp.) Changes in Soil Chemico-Physical Properties: a Mesocosm Experiment with Broccoli and Faba Bean

Earthworms are "ecosystem engineers" that improve soil water and nutrient content, soil macroporosity, and aeration, and provide suitable habitats for microbial populations. This study aimed at defining if the presence of epigeic earthworms (Eisenia sp.) affected the growth and development of two plant species (Brassica oleracea and Vicia faba) via the modifications of soil chemico-physical properties. A mesocosm experiment, in which plants were grown outdoors for 4 months with or without earthworms, was performed. The two plant species were selected based on their different habitus and root architecture and morphology. Soil macroporosity (M-soil) and water holding capacity (WHCsoil) were determined. Earthworm-driven bioturbation (B-soil) was measured by filling mesh bags with artificial soil. Earthworm abundance and biomass, together with plant morphometric parameters (root and leaf morphology by imaging and microscope techniques), were measured at the end of the trial. The presence of earthworms increased M-soil (on average +16%) and WHCsoil (on average +9%) and this was accompanied by a remarkable degree of B-soil. In most of the cases, earthworms enhanced plant growth in the two plant species studied, with a significant positive influence on the majority of the shoot and root traits. A significant increase of stomatal density (on average +24%) occurred in the leaves of both the plant species in the presence of earthworms. Our results confirmed the hypothesis that bioturbation by Eisenia sp. had a significant positive effect on plant growth, independently from the plant species cultivated, and that these growth-promoting effects were mediated by changes in soil chemico-physical parameters. By taking into account the essential role of earthworms in maintaining healthy soils and the vegetation they support, soils can become more resilient against environmental perturbations and climate change

A new emphasis on root traits for perennial grass and legume varieties with environmental and ecological benefits

Grasslands cover a significant proportion of the agricultural land within the UK and across the EU, providing a relatively cheap source of feed for ruminants and supporting the production of meat, wool and milk from grazing animals. Delivering efficient animal production from grassland systems has traditionally been the primary focus of grassland‐based research. But there is increasing recognition of the ecological and environmental benefits of these grassland systems and the importance of the interaction between their component plants and a host of other biological organisms in the soil and in adjoining habitats. Many of the ecological and environmental benefits provided by grasslands emanate from the interactions between the roots of plant species and the soil in which they grow. We review current knowledge on the role of grassland ecosystems in delivering ecological and environmental benefits. We will consider how improved grassland can deliver these benefits, and the potential opportunities for plant breeding to improve specific traits that will enhance these benefits whilst maintaining forage production for livestock consumption. Opportunities for exploiting new plant breeding approaches, including high throughput phenotyping, and for introducing traits from closely related species are discussed

Soil strength influences wheat root interactions with soil macropores

Deep rooting is critical for access to water and nutrients found in subsoil. However, damage to soil structure and the natural increase in soil strength with depth, often impedes root penetration. Evidence suggests that roots use macropores (soil cavities greater than 75μm) to bypass strong soil layers. If roots have to exploit structures, a key trait conferring deep rooting will be the ability to locate existing pore networks; a trait called trematotropism.In this study, artificial macropores were created in repacked soil columns at bulk densities of 1.6g cm‐3 and 1.2g cm‐3, representing compact and loose soil. Near isogenic lines of wheat, Rht‐B1a and Rht‐B1c, were planted and root‐macropore interactions were visualized and quantified using X‐ray Computed Tomography.In compact soil, 68.8% of root‐macropore interactions resulted in pore colonisation, compared to 12.5% in loose soil. Changes in root growth trajectory following pore interaction were also quantified, with 21.0% of roots changing direction (±3°) in loose soil compared to 76.0% in compact soil.These results indicate colonisation of macropores is an important strategy of wheat roots in compacted subsoil. Management practices to reduce subsoil compaction and encourage macropore formation could offer significant advantage in helping wheat roots penetrate deeper into subsoil

The interaction between wheat roots and soil pores in structured field soil

Wheat (Triticum aestivum L.) root growth in the subsoil is usually constrained by soil strength, although roots can use macropores to elongate to deeper layers. The quantitative relationship between the elongation of wheat roots and the soil pore system, however, is still to be determined. We studied the depth distribution of roots of six wheat varieties and explored their relationship with soil macroporosity from samples with the field structure preserved. Undisturbed soil cores (to a depth of 100 cm) were collected from the field and then non-destructively imaged using X-ray computed tomography (at a spatial resolution of 90 µm) to quantify soil macropore structure and root number density (the number of roots cm–2 within a horizontal cross-section of a soil core). Soil macroporosity changed significantly with depth but not between the different wheat lines. There was no significant difference in root number density between wheat varieties. In the subsoil, wheat roots used macropores, especially biopores (i.e. former root or earthworm channels) to grow into deeper layers. Soil macroporosity explained 59% of the variance in root number density. Our data suggested that the development of the wheat root system in the field was more affected by the soil macropore system than by genotype. On this basis, management practices which enhance the porosity of the subsoil may therefore be an effective strategy to improve deep rooting of wheat

RootPainter: Deep Learning Segmentation of Biological Images with Corrective Annotation

We present RootPainter, a GUI-based software tool for the rapid training of deep neural networks for use in biological image analysis. RootPainter facilitates both fully-automatic and semiautomatic image segmentation. We investigate the effectiveness of RootPainter using three plant image datasets, evaluating its potential for root length extraction from chicory roots in soil, biopore counting and root nodule counting from scanned roots. We also use RootPainter to compare dense annotations to corrective ones which are added during the training based on the weaknesses of the current model

Measuring root system traits of wheat in 2D images to parameterize 3D root architecture models

Background and aimsThe main difficulty in the use of 3D root architecture models is correct parameterization. We evaluated distributions of the root traits inter-branch distance, branching angle and axial root trajectories from contrasting experimental systems to improve model parameterization.MethodsWe analyzed 2D root images of different wheat varieties (Triticum aestivum) from three different sources using automatic root tracking. Model input parameters and common parameter patterns were identified from extracted root system coordinates. Simulation studies were used to (1) link observed axial root trajectories with model input parameters (2) evaluate errors due to the 2D (versus 3D) nature of image sources and (3) investigate the effect of model parameter distributions on root foraging performance.ResultsDistributions of inter-branch distances were approximated with lognormal functions. Branching angles showed mean values <90°. Gravitropism and tortuosity parameters were quantified in relation to downwards reorientation and segment angles of root axes. Root system projection in 2D increased the variance of branching angles. Root foraging performance was very sensitive to parameter distribution and variance.Conclusions2D image analysis can systematically and efficiently analyze root system architectures and parameterize 3D root architecture models. Effects of root system projection (2D from 3D) and deflection (at rhizotron face) on size and distribution of particular parameters are potentially significant

From rhizosphere to detritusphere – Soil structure formation driven by plant roots and the interactions with soil biota

Roots and the associated soil directly affected by root activity, termed the rhizosphere, have both been extensively studied and recognized for their crucial role in soil functioning. The formation of the rhizosphere is primarily driven by the effect of roots on shaping the physical structure of the soil, which in turn has direct feedbacks on the interactions between physical, biological and chemical processes. As a result, the rhizosphere is a hot spot for microbial activity, cycling of nutrients and turnover of organic matter. Despite the pivotal role of soil structure in controlling rhizosphere processes, we still lack a quantitative description and understanding of the interrelationships of root-systems and soil in the creation and stabilization of soil structure. We provide a comprehensive review of current knowledge and novel insights into processes that drive the formation and stabilization of soil structure in the rhizosphere. These processes are regulated by multiple indirect and direct pathways, involving root growth, the production of rhizodeposits and root hairs, as well as the activity of soil microorganisms and fauna. Further, we highlight that rhizosphere processes may persist and evolve after root death to an extent currently largely unknown. Finally, we identify five pertinent challenges that should be addressed to fully apprehend rhizosphere processes and thus harness the potential resilience of plant-soil interactions. These challenges include refining structural assessment and sampling of rhizosheaths, examining the rhizosphere in-situ and bridging the gap between solid phase and pore scale research. In our view, overcoming these obstacles can be accomplished by combining the power of imaging and isotopic approaches, especially at the field scale, encompassing diverse soils and plant species. The ultimate objective of future research should be to upscale rhizosphere processes by conducting more field experiments in concert with modeling efforts, under the umbrella of collaborative interdisciplinary research