Growth kinetics, spatialization and quality of potato tubers monitored in situ by MRI - long-term effects of water stress.

Understanding the potato tuber development and effects of drought at key stages of sensitivity on yield is crucial, particularly when considering the increasing incidence of drought due to climate change. So far, few studies addressed the time course of tuber growth in soil, mainly due to difficulties in accessing underground plant organs in a non-destructive manner. This study aims to understand the tuber growth and quality and the complex long-term effects of realistic water stress on potato tuber yield. MRI was used to monitor the growth kinetics and spatialization of individual tubers in situ and the evolution of internal defects throughout the development period. The intermittent drought applied to plants reduced tuber yield by reducing tuber growth and increasing the number of aborted tubers. The reduction in the size of tubers depended on the vertical position of the tubers in the soil, indicating water exchanges between tubers and the mother plant during leaf dehydration events. The final size of tubers was linked with the growth rate at specific developmental periods. For plants experiencing stress, this corresponded to the days following rewatering, suggesting tuber growth plasticity. All internal defects occurred in large tubers and within a short time span immediately following a period of rapid growth of perimedullary tissues, probably due to high nutrient requirements. To conclude, the non-destructive 3D imaging by MRI allowed us to quantify and better understand the kinetics and spatialization of tuber growth and the appearance of internal defects under different soil water conditions.

Quantitative MRI imaging of parenchyma and venation networks in Brassica napus leaves: effects of development and dehydration.

BACKGROUND: Characterisation of the structure and water status of leaf tissues is essential to the understanding of leaf hydraulic functioning under optimal and stressed conditions. Magnetic Resonance Imaging is unique in its capacity to access this information in a spatially resolved, non-invasive and non-destructive way. The purpose of this study was to develop an original approach based on transverse relaxation mapping by Magnetic Resonance Imaging for the detection of changes in water status and distribution at cell and tissue levels in Brassica napus leaves during blade development and dehydration. RESULTS: By combining transverse relaxation maps with a classification scheme, we were able to distinguish specific zones of areoles and veins. The tissue heterogeneity observed in young leaves still occurred in mature and senescent leaves, but with different distributions of T2 values in accordance with the basipetal progression of leaf blade development, revealing changes in tissue structure. When subjected to severe water stress, all blade zones showed similar behaviours. CONCLUSION: This study demonstrates the great potential of Magnetic Resonance Imaging in assessing information on the structure and water status of leaves. The feasibility of in planta leaf measurements was demonstrated, opening up many opportunities for the investigation of leaf structure and hydraulic functioning during development and/or in response to abiotic stresses.

Quantitative magnetic resonance imaging of in vitro gastrointestinal digestion of a bread and cheese meal.

The monitoring of food degradation during gastrointestinal digestion is essential in understanding food structure impacts on the bioaccessibility and bioavailability of nutrients. Magnetic Resonance Imaging (MRI) has the unique ability to access information on changes in multi-scale structural features of foods in a spatially resolved and non-destructive way. Our objective was to exploit various opportunities offered by MRI for monitoring starch, lipid and protein hydrolysis, as well as food particle breakdown during the semi-dynamic in vitro gastrointestinal digestion of complex foods combined in a meal. The meal consisted of French bread, hard cheese and water (drink), with a realistic distribution of bolus particle sizes. The MRI approach was reinforced by parallel chemical analysis of all macronutrients in the supernatant. By combining different imaging protocols, quantitative MRI provided insights into a number of phenomena at the level of the cheese and bread particles and within the liquid phase that are hard to access through conventional approaches. MRI thus revealed the progressive ingress of fluids into the bread crust and the release of the gas trapped in the crumb, the erosion of cheese particles, the creaming of fat, the disappearance of small food particles and changes in liquid phase composition. Excellent agreement was obtained between the quantitative parameters extracted from the MRI images and the results of the chemical analysis, demonstrating the strong potential of MRI for the monitoring of in vitro gastrointestinal digestion. The present study proposes further improvements to fully exploit the capabilities of MRI and constitutes an important step towards the extension of quantitative MRI to in vivo studies.

Characterization of Potato Tuber Tissues Using Spatialized MRI T2 Relaxometry.

Magnetic Resonance Imaging is a powerful non-destructive tool in the study of plant tissues. For potato tubers, it greatly assists the study of tissue defects and tissue evolution during storage. This paper describes the MRI analysis of potato tubers with internal defects in their flesh tissue at eight sampling dates from 14 to 33 weeks after harvest. Spatialized multi-exponential T2 relaxometry was used to generate bi-exponential T2 maps, coupled with a classification scheme to identify the different T2 homogeneous zones within the tubers. Six classes with statistically different relaxation parameters were identified at each sampling date, allowing the defects and the pith and cortex tissues to be detected. A further distinction could be made between three constitutive elements within the flesh, revealing the heterogeneity of this particular tissue. Relaxation parameters for each class and their evolution during storage were successfully analyzed. The work demonstrated the value of MRI for detailed non-invasive plant tissue characterization.

Characterization of the Water Shortage Effects on Potato Tuber Tissues during Growth Using MRI Relaxometry and Biochemical Parameters.

The potato is one of the most cultivated crops worldwide, providing an important source of food. The quality of potato tubers relates to their size and dry matter composition and to the absence of physiological defects. It depends on the spatial and temporal coordination of growth and metabolic processes in the major tuber tissues: the cortex, flesh and pith. In the present study, variations in the biochemical traits of each of these tissues were investigated during tuber growth under optimal and water-deficit conditions. MRI relaxometry was used as a non-invasive and quantitative method to access information on cellular water status. The presence of slight but significant variations in organic compound contents quantified in the cortex and flesh revealed a tissue-dependent metabolic pattern. The T2 and relative I0 of the bi-exponential relaxation signal allowed a distinction to be made between the pith and the cortex, whereas the flesh could be differentiated from these tissues only through its relative I0. T2 values did not vary significantly during tuber development, in accordance with the typical growth pattern of tubers, but were shown to be sensitive to water stress. The interpretation of the multi-exponential transverse relaxation times is discussed and could be further developed via microscopic analysis.

Multi-exponential MRI T2 maps: A tool to classify and characterize fruit tissues.

The estimation of multi-exponential relaxation time T2 and their associated amplitudes A0 at the voxel level has been made possible by recent developments in the field of image processing. These data are of great interest for the characterization of biological tissues, such as fruit tissues. However, they represent a high number of information, not easily interpretable. Moreover, the non-uniformity of the MRI images, which mainly directly impacts A0, could induce interpretation errors. In this paper, we propose a post-processing scheme that clusters similar voxels according to the multi-exponential relaxation parameters in order to reduce the complexity of the information while avoiding the problems associated with intensity non-uniformity. We also suggest a data representation suitable for the visualization of the multi-T2 distribution within each tissue. We illustrate this work with results for different fruits, demonstrating the great potential of multi-T2 information to shed new light on fruit characterization.

Quantitative MRI analysis of structural changes in tomato tissues resulting from dehydration.

A quantitative magnetic resonance imaging (MRI) analysis at 1.5T of the effects of different dehydration regimes on transverse relaxation parameters measured in tomato tissue is presented. Multi-exponential T2 maps have been estimated for the first time, providing access to spatialized microstructural information at voxel scale. The objective was to provide a better understanding of the changes in the multi-exponential transverse relaxation parameters induced by dehydration in tomato tissues and to unravel the effects of microstructure and composition on relaxation parameters. The results led to the hypothesis that the multi-exponential relaxation signal reflects cell compartmentation and tissue heterogeneity, even at the voxel scale. Multi-exponential relaxation times provided information about water loss from specific cell compartments and seem to indicate that the dehydration process mainly affects large cells. By contrast, total signal intensity showed no sensitivity to variations in water content in the range investigated in the present study (between 95% [fresh tissue] and 90% [after dehydration]). The variation in relaxation times resulting from water loss was due to both changes in solute concentration and compartment size. The comparative analysis of the two contrasted tissues in terms of microporosity demonstrated that magnetic susceptibility effects, caused by the presence of air in the placenta tissue, significantly impact the effective relaxation and might be the dominant effect in the variations observed in relaxation times in this tissue.

Low field, time domain NMR in the agriculture and agrifood sectors: An overview of applications in plants, foods and biofuels.

In this contribution, a selective overview of low field, time-domain NMR (TD-NMR) applications in the agriculture and agrifood sectors is presented. The first applications of commercial TD-NMR instruments were in food and agriculture domains. Many of these earlier methods have now been recognized as standard methods by several international agencies. Since 2000, several new applications have been developed, using state of the art instruments, new pulse sequences and new signal processing methods. TD-NMR is expected, in the coming years, to become even more important in quality control of fresh food and agricultural products, as well as for a wide range of food-processed products. TD-NMR systems provide excellent means to collect data relevant for use in the agricultural environment and the bioenergy industry. Data and information collected by TD-NMR systems thus may support decision makers in business and public organizations.

Impact of chemical exchange on transverse relaxation at low and moderate magnetic field strengths for sugar solutions representative of fruit tissues analyzed by simulation and MRI experiments.

Proton exchange effects on transverse relaxation rate were studied at low and moderate magnetic fields. Analysis was conducted on low-concentrate simple sugar (fructose and glucose) solutions modeling the vacuolar liquid in fruits. Simulated data obtained from Carver and Richards equations were used to analyze the effects of temperature and pH on parameters involved in the chemical exchange mechanisms. The results have highlighted that measurement conditions and tissue characteristics (echo time, magnetic field strength, temperature, pH, etc.) significantly impact the transverse relaxation rate via chemical exchange, even for low and mid magnetic fields and the narrow echo time ranges of TD-NMR and MRI measurements. This explains some of the divergent relaxation data from plant tissue NMR measurements reported in the literature and emphasizes the importance of taking experimental conditions and tissue characteristics into account when interpreting results. It also clearly demonstrates that the appropriate choice of experimental conditions can emphasize the effects of sugar concentration or pH variations on transverse relaxation in vacuolar liquids, making it possible to monitor these parameters accurately via transverse relaxation measurements. The impact of concentration, solute type, pH and temperature on transverse relaxation of sugar solutions were demonstrated experimentally at 1.5 T with an MRI scanner. These data were interpreted using the Carver-Richards model which was effective in estimating parameters involved in chemical exchange mechanism in the imaging experiment.

Spatial and temporal evolution of quantitative magnetic resonance imaging parameters of peach and apple fruit - relationship with biophysical and metabolic traits.

Fruits are complex organs that are spatially regulated during development. Limited phenotyping capacity at cell and tissue levels is one of the main obstacles to our understanding of the coordinated regulation of the processes involved in fruit growth and quality. In this study, the spatial evolution of biophysical and metabolic traits of peach and apple fruit was investigated during fruit development. In parallel, the multi-exponential relaxation times and apparent microporosity were assessed by quantitative magnetic resonance imaging (MRI). The aim was to identify the possible relationships between MRI parameters and variations in the structure and composition of fruit tissues during development so that transverse relaxation could be proposed as a biomarker for the assessment of the structural and functional evolution of fruit tissues during growth. The study provides species-specific data on developmental and spatial variations in density, cell number and size distribution, insoluble and soluble compound accumulation and osmotic and water potential in the fruit mesocarp. Magnetic resonance imaging was able to capture tissue evolution and the development of pericarp heterogeneity by accessing information on cell expansion, water status and distribution at cell level, and microporosity. Changes in vacuole-related transverse relaxation rates were mostly explained by cell/vacuole size. The impact of cell solute composition, microporosity and membrane permeability on relaxation times is also discussed. The results demonstrate the usefulness of MRI as a tool to phenotype fruits and to access important physiological data during development, including information on spatial variability.

Multi-exponential Transverse Relaxation Times Estimation from Magnetic Resonance Images under Rician Noise and Spatial Regularization.

Relaxation signal inside each voxel of magnetic resonance images (MRI) is commonly fitted by a multi-exponential decay curve. The estimation of a discrete multi-component relaxation model parameters from magnitude MRI data is a challenging nonlinear inverse problem since it should be conducted on the entire image voxels under non-Gaussian noise statistics. This paper proposes an efficient algorithm allowing the joint estimation of relaxation time values and their amplitudes using different criteria taking into account a Rician noise model, combined with a spatial regularization accounting for low spatial variability of relaxation time constants and amplitudes between neighboring voxels. The Rician noise hypothesis is accounted for either by an adapted nonlinear least squares algorithm applied to a corrected least squares criterion or by a majorization-minimization approach applied to the maximum likelihood criterion. In order to solve the resulting large-scale non-negativity constrained optimization problem with a reduced numerical complexity and computing time, an optimization algorithm based on a majorization approach ensuring separability of variables between voxels is proposed. The minimization is carried out iteratively using an adapted Levenberg-Marquardt algorithm that ensures convergence by imposing a sufficient decrease of the objective function and the non-negativity of the parameters. The importance of the regularization alongside the Rician noise incorporation is shown both visually and numerically on a simulated phantom and on magnitude MRI images acquired on fruit samples.

NMR study of fresh cut salads: Influence of temperature and storage time on leaf structure and water distribution in escarole.

Consumption of fresh-cut vegetables has rapidly increased over the past decades. Among salads, escarole is one of the most popular varieties. Specific packaging limits gas exchange and consequently water loss and bacterial respiration, increasing the shelf life of salads. Although the major cause of quality loss for minimally processed salads is the leaf textural changes, this aspect has rarely been investigated. Therefore, investigating structural changes of leaves during storage is important in order to understand and minimize quality loss of salads. In this study, we focused on the impact of storage duration and temperature on the escarole leaf structure. The complex leaf structure was investigated by relaxation NMR, via transverse relaxation times, which allows the specific description of vacuolar water compartment of the cell. The storage duration (maximum 12 days) and temperatures (4°C, 7°C, 10°C, and 12°C) have been chosen in order to represent the conditions registered in factory. The results showed that the temperature did not have significant impact on the salad structure during the first week. During the second week, changes in the water distribution and changes in the relaxation time T2 have been observed. The changes in transverse relaxation times associated with vacuolar water are related to lost of cell membrane and wall integrity. The NMR results confirmed the effect of storage temperature on the degradation process of the cell before visual detection of the salad leaf degradation. The present study confirmed the sensibility of NMR relaxometry for monitoring water changes in the leaf.

A mobile NMR lab for leaf phenotyping in the field.

BACKGROUND: Low field NMR has been used to investigate water status in various plant tissues. In plants grown in controlled conditions, the method was shown to be able to monitor leaf development as it could detect slight variations in senescence associated with structural modifications in leaf tissues. The aim of the present study was to demonstrate the potential of NMR to provide robust indicators of the leaf development stage in plants grown in the field, where leaves may develop less evenly due to environmental fluctuations. The study was largely motivated by the need to extend phenotyping investigations from laboratory experiments to plants in their natural environment. METHODS: The mobile NMR laboratory was developed, enabling characterization of oilseed rape leaves throughout the canopy without uprooting the plant. The measurements made on the leaves of plants grown and analyzed in the field were compared to the measurements on plants grown in controlled conditions and analyzed in the laboratory. RESULTS: The approach demonstrated the potential of the method to assess the physiological status of leaves of plants in their natural environment. Comparing changes in the patterns of NMR signal evolution in plants grown under well-controlled laboratory conditions and in plants grown in the field shows that NMR is an appropriate method to detect structural modifications in leaf tissues during senescence progress despite plant heterogeneity in natural conditions. Moreover, the specific effects of the environmental factors on the structural modifications were revealed. CONCLUSION: The present study is an important step toward the selection of genotypes with high tolerance to water or nitrogen depletion that will be enabled by further field applications of the method.

Nitrogen deficiency impacts on leaf cell and tissue structure with consequences for senescence associated processes in Brassica napus.

Improvement of nutrient use efficiency is a major goal for several crop plants, especially Brassica napus. Indeed, the low nitrogen use efficiency (NUE) in this crop results in negative economic and ecological consequences. The low NUE of oilseed rape is mainly due to low remobilization of nitrogen from vegetative parts to growing organs. Remobilization of leaf nitrogen takes place during senescence, a process known to strongly modify cell and tissue structure. This study focused on the impact of moderate N depletion, expected to induce 30 % reduction of seed yield, on these structural modifications. Two genotypes (Aviso and Express) were studied, with different tolerance of nitrogen depletion, evaluated through seed yield and dry mass production. Structural modifications of leaf cells and tissues were investigated through NMR relaxometry and light microscopy. Lower tolerance of N depletion was associated with higher impact on senescence associated structural modification pattern. The link between leaf structure modifications and nutrient remobilization is discussed. It is proposed that leaf structure monitoring during senescence through NMR device could be developed to select genotypes with high NUE.

MRI investigation of subcellular water compartmentalization and gas distribution in apples.

Water status and distribution at subcellular level in whole apple fruit were evaluated by Magnetic Resonance Imaging (MRI) measurement of the multi-exponential transverse (T2) relaxation of water protons. Apparent microporosity, also estimated by MRI, provided mapping of gas distribution in fruit tissues. Measuring for the first time the multi-exponential relaxation of water and apparent tissue microporosity in whole fruit and combining these with histological measurements provided a more reliable interpretation of the origins of variations in the transverse relaxation time (T2) and better characterization of the fruit tissue. Measurements were performed on 54 fruits from 3 different cultivars. Fruits of different sizes were selected for each cultivar to provide tissues with cells of different dimensions. Macrovision measurements were carried out on parenchymal tissue from all fruits to investigate the impact of cell size on T2 value. The results showed that the MRI transverse relaxation signal is well fitted by a tri-exponential decay curve that reflects cell compartmentalization. Variations in cell size partially explained the different T2 observed. This study highlighted the heterogeneity of apple tissues in terms of relaxation parameters, apparent microporosity and cell morphology and in relation to specific variations between fruit of different cultivars.

Assessment of nutrient remobilization through structural changes of palisade and spongy parenchyma in oilseed rape leaves during senescence.

MAIN CONCLUSION: Differential palisade and spongy parenchyma structural changes in oilseed rape leaf were demonstrated. These dismantling processes were linked to early senescence events and associated to remobilization processes. During leaf senescence, an ordered cell dismantling process allows efficient nutrient remobilization. However, in Brassica napus plants, an important amount of nitrogen (N) in fallen leaves is associated with low N remobilization efficiency (NRE). The leaf is a complex organ mainly constituted of palisade and spongy parenchyma characterized by different structures and functions concerning water relations and carbon fixation. The aim of the present study was to demonstrate a specific structural evolution of these parenchyma throughout natural senescence in B. napus, probably linked to differential nutrient remobilization processes. The study was performed on 340 leaves from 32 plants during an 8-week development period under controlled growing conditions. Water distribution and status at the cellular level were investigated by low-field proton nuclear magnetic resonance (NMR), while light and electron microscopy were used to observe cell and plast structure. Physiological parameters were determined on all leaves studied and used as indicators of leaf development and remobilization progress. The results revealed a process of hydration and cell enlargement of leaf tissues associated with senescence. Wide variations were observed in the palisade parenchyma while spongy cells changed only very slightly. The major new functional information revealed was the link between the early senescence events and specific tissue dismantling processes.

Structural changes in senescing oilseed rape leaves at tissue and subcellular levels monitored by nuclear magnetic resonance relaxometry through water status.

Nitrogen use efficiency is relatively low in oilseed rape (Brassica napus) due to weak nitrogen remobilization during leaf senescence. Monitoring the kinetics of water distribution associated with the reorganization of cell structures, therefore, would be valuable to improve the characterization of nutrient recycling in leaf tissues and the associated senescence processes. In this study, nuclear magnetic resonance (NMR) relaxometry was used to describe water distribution and status at the cellular level in different leaf ranks of well-watered plants. It was shown to be able to detect slight variations in the evolution of senescence. The NMR results were linked to physiological characterization of the leaves and to light and electron micrographs. A relationship between cell hydration and leaf senescence was revealed and associated with changes in the NMR signal. The relative intensities and the transverse relaxation times of the NMR signal components associated with vacuole water were positively correlated with senescence, describing water uptake and vacuole and cell enlargement. Moreover, the relative intensity of the NMR signal that we assigned to the chloroplast water decreased during the senescence process, in agreement with the decrease in relative chloroplast volume estimated from micrographs. The results are discussed on the basis of water flux occurring at the cellular level during senescence. One of the main applications of this study would be for plant phenotyping, especially for plants under environmental stress such as nitrogen starvation.

MSE-MRI sequence optimisation for measurement of bi- and tri-exponential T2 relaxation in a phantom and fruit.

The transverse relaxation signal from vegetal cells can be described by multi-exponential behaviour, reflecting different water compartments. This multi-exponential relaxation is rarely measured by conventional MRI imaging protocols; mono-exponential relaxation times are measured instead, thus limiting information about of the microstructure and water status in vegetal cells. In this study, an optimised multiple spin echo (MSE) MRI sequence was evaluated for assessment of multi-exponential transverse relaxation in fruit tissues. The sequence was designed for the acquisition of a maximum of 512 echoes. Non-selective refocusing RF pulses were used in combination with balanced crusher gradients for elimination of spurious echoes. The study was performed on a bi-compartmental phantom with known T2 values and on apple and tomato fruit. T2 decays measured in the phantom and fruit were analysed using bi- and tri-exponential fits, respectively. The MRI results were compared with low field non-spatially resolved NMR measurements performed on the same samples. The results demonstrated that the MSE-MRI sequence can be used for up to tri-exponential T2 quantification allowing for estimation of relaxation times from a few tens of milliseconds to over a second. The effects of the crusher moment and the TE value on T2 measurements were studied both on the bi-compartmental phantom and on the fruit tissues. It was demonstrated that the sequence should be optimised with regard to the characteristics of the tissue to be examined by considering the effects of water molecular diffusion in the presence of both imaging gradients and gradients produced by susceptibility inhomogeneities.

Temperature mapping in bread dough using SE and GE two-point MRI methods: experimental and theoretical estimation of uncertainty.

Two-dimensional (2D)-SE, 2D-GE and tri-dimensional (3D)-GE two-point T(1)-weighted MRI methods were evaluated in this study in order to maximize the accuracy of temperature mapping of bread dough during thermal processing. Uncertainties were propagated throughout each protocol of measurement, and comparisons demonstrated that all the methods with comparable acquisition times minimized the temperature uncertainty to similar extent. The experimental uncertainties obtained with low-field MRI were also compared to the theoretical estimations. Some discrepancies were reported between experimental and theoretical values of uncertainties of temperature; however, experimental and theoretical trends with varying parameters agreed to a large extent for both SE and GE methods. The 2D-SE method was chosen for further applications on prefermented dough because of its lower sensitivity to susceptibility differences in porous media. It was applied for temperature mapping in prefermented dough during chilling prior to freezing and compared locally to optical fiber measurements.

Quantification of microporosity in fruit by MRI at various magnetic fields: comparison with X-ray microtomography.

Microstructure determines the mechanical and transport properties of fruit tissues. One important characteristic of the microstructure is the relative volume fraction of gas-filled intercellular spaces, i.e., the tissue microporosity. Quantification of this microporosity is fundamental for investigating the relationship between gas transfer and various disorders in fruit. We present a new method for quantifying the apparent microporosity using magnetic resonance imaging (MRI). The method is based on the differences in magnetic susceptibility between gas-filled intercellular spaces and their environment inside fruit tissues. It was tested at two different magnetic fields (1.5 and 0.2 T) on apple and tomato fruit. The method was validated by comparing the MRI results with estimation of local tissue porosity using X-ray microtomography experiments. MRI was shown to be effective in determining the distribution of apparent microporosity in fruit.