Multi-spectral imaging of rhizobox systems: New perspectives for the observation and discrimination of rhizosphere components

In this issue of Plant and Soil Nakaji et al. (Plant Soil, this volume, 2008) report a novel approach for automatically identifying roots and other rhizosphere components in rhizosphere images acquired using a multi-spectral (visible—VIS- and near-infrared—NIR-) imaging system. The images are acquired through a root-window observation device and the study highlights the perspectives offered by this imaging system. An outstanding outcome of this research is that the new approach can be applied to effectively separate soil litter from the purely mineral phase and distinguish root tissues that differ in physiological status, i.e. live (different age classes), senescent and dead. If achievable routinely, such a detailed classification of rhizosphere components could greatly improve our appraisal of root turnover and associated organic matter input to the soil, information of paramount importance for an improved understanding of many essential processes such as global geochemical cycles. Minirhizotrons (MR) systems have been increasingly used in global change studies because they are a convenient way to frequently and nondestructively quantify root length production and mortality (Norby and Jackson, New Phytol, 147:3–12, 2000; Hendrick and Pregitzer, Ecology, 73:1094–1104, 1992). However, the MR technique still has many limitations, including the lack of a standard, accurate and rapid procedure to extract and classify rhizosphere components from the MR images obtained. The recent work by Nakaji et al. (Plant Soil, this volume, 2008) provides convincing evidence that the inclusion of a VIS-NIR multi-spectral capability into conventional MR systems could substantially improve this method, and extend its adoption by the wider plant scientist community as a standard research tool.     

X-ray absorption and phase contrast imaging to study the interplay between plant roots and soil structure

Plant performance is, at least partly, linked to the location of roots with respect to soil structure features and the micro-environment surrounding roots. Measurements of root distributions from intact samples, using optical microscopy and field tracings have been partially successful but are imprecise and labour-intensive. Theoretically, X-ray computed micro-tomography represents an ideal solution for non-invasive imaging of plant roots and soil structure. However, before it becomes fast enough and affordable or easily accessible, there is still a need for a diagnostic tool to investigate root/soil interplay. Here, a method for detection of undisturbed plant roots and their immediate physical environment is presented. X-ray absorption and phase contrast imaging are combined to produce projection images of soil sections from which root distributions and soil structure can be analyzed. The clarity of roots on the X-ray film is sufficient to allow manual tracing on an acetate sheet fixed over the film. In its current version, the method suffers limitations mainly related to (i) the degree of subjectivity associated with manual tracing and (ii) the difficulty of separating live and dead roots. The method represents a simple and relatively inexpensive way to detect and quantify roots from intact samples and has scope for further improvements. In this paper, the main steps of the method, sampling, image acquisition and image processing are documented. The potential use of the method in an agronomic perspective is illustrated using surface and sub-surface soil samples from a controlled wheat trial. Quantitative characterization of root attributes, e.g. radius, length density, branching intensity and the complex interplay between roots and soil structure, is presented and discussed. This revised version was published online in June 2006 with corrections to the Cover Date.     

Estimating root elongation rates from morphological measurements of the root tip

To measure the elongation rate of individual roots in soil remains a challenge. A novel method for estimating elongation rates of excavated roots is presented. Morphological markers are identified along the tip of excavated roots, and their distance relative to the apex is measured. These markers correspond to developmental stages which follow known temporal patterns. Hence, their distance relative to the apex reflects root elongation during the period corresponding to their development. The method was tested on maize roots grown in a range of conditions and substrates. It was found that distances from markers to apices were proportional, with some variability, to elongation rates. Remarkably, the linear relationships between these distances were neither affected by substrate, nor by growing conditions. Using several markers allows covering time periods ranging from 0.3 day to 3 days as well as cross validation of estimates. Provided further testing, under a wider range of environmental conditions, is conducted, the concepts presented in this paper may serve to define a new measurement technique.     

Differentiation of soil properties related to the spatial association of wheat roots and soil macropores

Under certain soil conditions, e.g. hardsetting clay B-horizons of South-Eastern Australia, wheat plants do not perform as well as would be expected given measurements of bulk soil attributes. In such soils, measurement indicates that a large proportion (80%) of roots are preferentially located in the soil within 1 mm of macropores. This paper addresses the question of whether there are biological and soil chemical effects concomitant with this observed spatial relationship. The properties of soil manually dissected from the 1–3 mm wide region surrounding macropores, the macropore sheath, were compared to those that are measured in a conventional manner on the bulk soil. Field specimens of two different soil materials were dissected to examine biological differentiation. To ascertain whether the macropore sheath soil differs from rhizosphere soil, wheat was grown in structured and repacked cores under laboratory conditions. The macropore sheath soil contained more microbial biomass per unit mass than both the bulk soil and the rhizosphere. The bacterial population in the macropore sheath was able to utilise a wider range of carbon substrates and to a greater extent than the bacterial population in the corresponding bulk soil. These differences between the macropore sheath and bulk soil were almost non-existent in the repacked cores. Evidence for larger numbers of propagules of the broad host range fungus Pythium in the macropore sheath soil were also obtained. This revised version was published online in June 2006 with corrections to the Cover Date.     

Efficient and sensitive detection of residues of nine coccidiostats in egg and muscle by liquid chromatography-electrospray tandem mass spectrometry

We present a method based on electrospray liquid chromatography tandem mass spectrometry (LC-MS/MS) for determining in muscle and eggs the following nine coccidiostats: halofuginone, diclazuril, dinitrocarbanilide (the main metabolite of nicarbazin), robenidine, monensin, lasalocid, narasin, salinomycin, and maduramicin. Dinitrocarbanilide-d8, nigericin, and diclazuril-bis were used as internal standards. The method uses extraction in acetonitrile followed by a clean-up on an SiOH solid-phase extraction column. High-performance liquid chromatography (HPLC) separation was performed on a Purospher C(18) column (125 mm x 3 mm i.d.) protected by a guard column, the mobile phase being a water-acetonitrile gradient (each gradient component containing 0.1% formic acid) at a flow rate of 1 ml min(-1). For unequivocal identification of each analyte, two ions were detected and chosen for multiple reaction monitoring (MRM). Validation was carried out on spiked muscle and egg samples. The method described meets all the criteria of Decision 2002/657/EC and is easy to use in routine analysis. Validation results are presented with the measured CCalpha and CCbeta values. This whole method allows extraction and analysis of up to 24 samples per day.     

Water velocity dependence of phosphate uptake on a coral-dominated fringing reef flat,La Réunion Island,Indian Ocean

Phosphate uptake (P-uptake) into coral reef communities has been hypothesized to be mass-transfer limited. One method of demonstrating mass-transfer limitation of P-uptake is to show dependence of P-uptake on water velocity. Water velocity across reef flats varies with tides and swell; thus, we measured P-uptake over the entire reef flat on eight different days, representing a range in water velocities. P-uptake was calculated from changes in P concentration of the water column. Changes in P concentration were measured by water sampling at six sites along a 300-m cross-reef transect while simultaneously measuring water velocity. To smooth the variability in phosphate concentrations, peristaltic pumps were used to get time-integrated water samples for 4–6 h at each site. Water velocities were measured in the middle of the transect using an acoustic Doppler current profiler and were averaged to match the time-integrated water sampling. Depth-averaged cross-reef water velocities were 0.031 ± 0.013 m s⁻¹ (mean ± SD), while the root-mean-square water velocities, accounting for oscillatory flow, averaged 3.3 times higher, 0.101 ± 0.021 m s⁻¹ (mean ± SD). Phosphate decreased along all transects. The first-order rate constant for P-uptake (S) was 8.5 ± 2.4 m d⁻¹ (mean ± SD) and increased linearly with root-mean-square water velocity. The Stanton number derived from oscillatory flow, the ratio of the first-order rate constant for P-uptake to the root-mean-square water velocity (S/U _rms), was (9.4 ± 1.2) × 10⁻⁴ (mean ± SD). P-uptake ranged from 0.2 to 1.1 mmol P m⁻² d⁻¹, demonstrating that P-uptake is variable on short time scales and is directly related to P concentration and water velocity.

     

Simultaneous X-ray imaging of plant root growth and water uptake in thin-slab systems

Direct and simultaneous observation of root growth and plant water uptake is difficult because soils are opaque. X-ray imaging techniques such as projection radiography or Computer Tomography (CT) offer a partial alternative to such limitations. Nevertheless, there is a trade-off between resolution, large field-of-view and 3-dimensionality: With the current state of the technology, it is possible to have any two. In this study, we used X-ray transmission through thin-slab systems to monitor transient saturation fields that develop around roots as plants grow. Although restricted to 2-dimensions, this approach offers a large field-of-view together with high spatial and dynamic resolutions. To illustrate the potential of this technology, we grew peas in 1 cm thick containers filled with soil and imaged them at regular intervals. The dynamics of both the root growth and the water content field that developed around the roots could be conveniently monitored. Compared to other techniques such as X-ray CT, our system is relatively inexpensive and easy to implement. It can potentially be applied to study many agronomic problems, such as issues related to the impact of soil constraints (physical, chemical or biological) on root development.     

Transfer of rare earth elements (REE) from natural soil to plant systems: implications for the environmental availability of anthropogenic REE

Background and aims

Rare Earth Elements (REE) are widely used to trace natural geochemical processes. They are also increasingly used by man (electronics industry, medicine, agriculture) and therefore considered as emerging pollutants. The present study documents REE mobility in non-polluted natural soil-plant systems in order to characterize their environmental availability for future anthropogenic pollution.

Methods

The study is based on a field approach in non-polluted natural sites with contrasting geological environments (limestone, granite, and carbonatite) and highly variable REE contents.

Results

REE concentrations in soils do not directly reflect bedrock concentrations, but depend largely on pedogenetic processes and on the mineralogy of bedrock and soil. The soils of all sites are with respect to bedrock enriched in heavy REE. The REE uptake by plants is not primarily controlled by the plant itself, but depends on the concentration and the speciation in the soil and the adsorbed soil water pool.

Conclusions

REE uptake by plant roots are linked with those of Fe. Roots absorb preferentially the light REE. Before translocation, REE are retained by the Casparian strip leading to much lower concentrations in the aerial parts. The transport of the REE within the xylem is associated with the general nutrient flux.     

Influence of xylem development on axial hydraulic conductance within Prunus root systems

The effect of secondary growth on the distribution of the axial hydraulic conductance within the Prunus root system was investigated. Secondary growth resulted in a large increase in both the number (from about 10 to several thousand) and diameter of xylem vessels (from a few micrometres to nearly 150 µm). For fine roots (<3 mm), an increase in root diameter was correlated with a slight increase in the number of xylem vessels and a large increase in their diameter. Conversely, for woody roots, an increase in root diameter was associated with a dramatic increase in the number of xylem vessels, but little or no change in vessel diameter. The theoretical axial conductivity (Kh, m⁴.s^-1.MPa^-1) of root segments was calculated with the Poiseuille-Hagen equation from measurements of vessel diameter. Kh measured using the tension-induced technique varies over several orders of magnitude (7.4᎒^-11 to 5.7᎒^-7 m⁴.s^-1.MPa^-1) and shows large discrepancies with theoretical calculated Kh. We concluded that root diameter is a pertinent and useful parameter to predict the axial conductance of a given root, provided the root type is known. Indeed, the relationship between measured Kh and root diameter varies according to the root type (fine or woody), due to differences in the xylem produced by secondary growth. Finally, we show how the combination of branching pattern and axial conductance may limit water flow through root systems. For Prunus, the main roots do not appear to limit water transfer; the axial conductance of the main axes is at least 10% higher than the sum of the axial conductance of the branches.