A bioinspired approach for the modulation of electroosmotic flow and protein-surface interactions in capillary electrophoresis using silylated amino-amides blocks and covalent grafting.

We explore a bioinspired approach to design tailored functionalized capillary electrophoresis (CE) surfaces based on covalent grafting for biomolecules analysis. First, the approach aims to overcome well-known common obstacles in CE protein analysis affecting considerably the CE performance (asymmetry, resolution, and repeatability) such as the unspecific adsorption on fused silica surface and the lack of control of electroosmotic flow (EOF). Then, our approach, which relies on new amino-amide mimic hybrid precursors synthesized by silylation of amino-amides (Si-AA) derivatives with 3-isocyanatopropyltriethoxysilane, aims to recapitulate the diversity of protein-protein interactions (π-π stacking, ionic, Van der Waals…) found in physiological condition (bioinspired approach) to improve the performance of CE protein analysis (electrochromatography). As a proof of concept, these silylated Si-AA (tyrosinamide silylation, serinamide silylation, argininamide silylation, leucinamide silylation, and isoglutamine silylation acid) have been covalently grafted in physiological conditions in different amount on bare fused silica capillary giving rise to a biomimetic coating and allowing both the modulation of EOF and protein-surface interactions. The analytical performances of amino-amide functionalized capillaries were assessed using lysozyme, cytochrome C and ribonuclease A and compared to traditional capillary coatings poly(ethylene oxide), poly(diallyldimethylammonium chloride), and sodium poly(styrenesulfonate). EOF, protein adsorption rate, protein retention factor k, and selectivity were determined for each coating. All results obtained showed this approach allowed to modulate the EOF, reduce unspecific adsorption, and generate specific interactions with proteins by varying the nature and the amount of Si-AA in the functionalization mixture.

Shear Wave Elastography, a New Tool for Diaphragmatic Qualitative Assessment: A Translational Study.

Rationale: Prolonged mechanical ventilation is often associated with either a decrease (known atrophy) or an increase (supposed injury) in diaphragmatic thickness. Shear wave elastography is a noninvasive technique that measures shear modulus, a surrogate of tissue stiffness and mechanical properties. Objectives: To describe changes in shear modulus (SM) during the ICU stay and the relationship with alterations in muscle thickness. To perform a comprehensive ultrasound-based characterization of histological and force production changes occurring in the diaphragm. Methods: Translational study using critically ill patients and mechanically ventilated piglets. Serial ultrasound examination of the diaphragm collecting thickness and SM was performed in both patients and piglets. Transdiaphragmatic pressure and diaphragmatic biopsies were collected in piglets. Measurements and Main Results: We enrolled 102 patients, 88 of whom were invasively mechanically ventilated. At baseline, SM was 14.3 ± 4.3 kPa and diaphragm end-expiratory thickness was 2.0 ± 0.5 mm. Decrease or increase by more than 10% from baseline was reported in 86% of the patients for thickness and in 92% of the patients for SM. An increase in diaphragmatic thickness during the stay was associated with a decrease in SM (ß = -9.34 ± 4.41; P = 0.03) after multivariable analysis. In the piglet sample, a decrease in SM over 3 days of mechanical ventilation was associated with loss of force production, slow and fast fiber atrophy, and increased lipid droplets accumulation. Conclusions: Increases in diaphragm thickness during critical illness is associated with decreased tissue stiffness as demonstrated by shear wave ultrasound elastography, consistent with the development of muscle injury and weakness. Clinical trial registered with www.clinicaltrials.gov (NCT03550222).

Development of Amino Acids Functionalized SBA-15 for the Improvement of Protein Adsorption.

Ordered mesoporous materials and their modification with multiple functional groups are of wide scientific interest for many applications involving interaction with biological systems and biomolecules (e.g., catalysis, separation, sensor design, nano-science or drug delivery). In particular, the immobilization of enzymes onto solid supports is highly attractive for industry and synthetic chemistry, as it allows the development of stable and cheap biocatalysts. In this context, we developed novel silylated amino acid derivatives (Si-AA-NH2) that have been immobilized onto SBA-15 materials in biocompatible conditions avoiding the use of toxic catalyst, solvents or reagents. The resulting amino acid-functionalized materials (SBA-15@AA) were characterized by XRD, TGA, EA, Zeta potential, nitrogen sorption and FT-IR. Differences of the physical properties (e.g., charges) were observed while the structural ones remained unchanged. The adsorption of the enzyme lysozyme (Lyz) onto the resulting functionalized SBA-15@AA materials was evaluated at different pHs. The presence of different functional groups compared with bare SBA-15 showed better adsorption results, for example, 79.6 nmol of Lyz adsorbed per m2 of SBA-15@Tyr compared with the 44.9 nmol/m2 of the bare SBA-15.

Host-specific competitiveness to form nodules in Rhizobium leguminosarum symbiovar viciae.

Fabeae legumes such as pea and faba bean form symbiotic nodules with a large diversity of soil Rhizobium leguminosarum symbiovar viciae (Rlv) bacteria. However, bacteria competitive to form root nodules (CFN) are generally not the most efficient to fix dinitrogen, resulting in a decrease in legume crop yields. Here, we investigate differential selection by host plants on the diversity of Rlv. A large collection of Rlv was collected by nodule trapping with pea and faba bean from soils at five European sites. Representative genomes were sequenced. In parallel, diversity and abundance of Rlv were estimated directly in these soils using metabarcoding. The CFN of isolates was measured with both legume hosts. Pea/faba bean CFN were associated to Rlv genomic regions. Variations of bacterial pea and/or faba bean CFN explained the differential abundance of Rlv genotypes in pea and faba bean nodules. No evidence was found for genetic association between CFN and variations in the core genome, but variations in specific regions of the nod locus, as well as in other plasmid loci, were associated with differences in CFN. These findings shed light on the genetic control of CFN in Rlv and emphasise the importance of host plants in controlling Rhizobium diversity.

Correction to: Real-time shear wave ultrasound elastography: a new tool for the evaluation of diaphragm and limb muscle stiffness in critically ill patients.

In the publication of this article [1], there was an error in the Family Name of one of the authors. This has now been updated in the original article.

Real-time shear wave ultrasound elastography: a new tool for the evaluation of diaphragm and limb muscle stiffness in critically ill patients.

BACKGROUND: Muscle weakness following critical illness is the consequence of loss of muscle mass and alteration of muscle quality. It is associated with long-term disability. Ultrasonography is a reliable tool to quantify muscle mass, but studies that evaluate muscle quality at the critically ill bedside are lacking. Shear wave ultrasound elastography (SWE) provides spatial representation of soft tissue stiffness and measures of muscle quality. The reliability and reproducibility of SWE in critically ill patients has never been evaluated. METHODS: Two operators tested in healthy controls and in critically ill patients the intra- and inter-operator reliability of the SWE using transversal and longitudinal views of the diaphragm and limb muscles. Reliability was calculated using the intra-class correlation coefficient and a bootstrap sampling method assessed their consistency. RESULTS: We collected 560 images. Longitudinal views of the diaphragm (ICC 0.83 [0.50-0.94]), the biceps brachii (ICC 0.88 [0.67-0.96]) and the rectus femoris (ICC 0.76 [0.34-0.91]) were the most reliable views in a training set of healthy controls. Intra-class correlation coefficient for inter-operator reproducibility and intra-operator reliability was above 0.9 for all muscles in a validation set of healthy controls. In critically ill patients, inter-operator reproducibility and intra-operator 1 and 2 reliability ICCs were respectively 0.92 [0.71-0.98], 0.93 [0.82-0.98] and 0.92 [0.81-0.98] for the diaphragm; 0.96 [0.86-0.99], 0.98 [0.94-0.99] and 0.99 [0.96-1] for the biceps brachii and 0.91 [0.51-0.98], 0.97 [0.93-0.99] and 0.99 [0.97-1] for the rectus femoris. The probability to reach intra-class correlation coefficient greater than 0.8 in a 10,000 bootstrap sampling for inter-operator reproducibility was respectively 81%, 84% and 78% for the diaphragm, the biceps brachii and the rectus femoris respectively. CONCLUSIONS: SWE is a reliable technique to evaluate limb muscles and the diaphragm in both healthy controls and in critically ill patients. TRIAL REGISTRATION: The study was registered (ClinicalTrial NCT03550222).

The genetics underlying natural variation of plant-plant interactions, a beloved but forgotten member of the family of biotic interactions.

Despite the importance of plant-plant interactions on crop yield and plant community dynamics, our understanding of the genetic and molecular bases underlying natural variation of plant-plant interactions is largely limited in comparison with other types of biotic interactions. By listing 63 quantitative trait loci (QTL) mapping and global gene expression studies based on plants directly challenged by other plants, we explored whether the genetic architecture and the function of the candidate genes underlying natural plant-plant interactions depend on the type of interactions between two plants (competition versus commensalism versus reciprocal helping versus asymmetry). The 16 transcriptomic studies are unevenly distributed between competitive interactions (n = 12) and asymmetric interactions (n = 4, all focusing on response to parasitic plants). By contrast, 17 and 30 QTL studies were identified for competitive interactions and asymmetric interactions (either weed suppressive ability or response to parasitic plants), respectively. Surprisingly, no studies have been carried out on the identification of genetic and molecular bases underlying natural variation in positive interactions. The candidate genes underlying natural plant-plant interactions can be classified into seven categories of plant function that have been identified in artificial environments simulating plant-plant interactions either frequently (photosynthesis, hormones), only recently (cell wall modification and degradation, defense pathways against pathogens) or rarely (ABC transporters, histone modification and meristem identity/life history traits). Finally, we introduce several avenues that need to be explored in the future to obtain a thorough understanding of the genetic and molecular bases underlying plant-plant interactions within the context of realistic community complexity.

Microfluidic Systems for Droplet Generation in Aqueous Continuous Phases: A Focus Review.

Microfluidics is one of the most fascinating fields that researchers have been trying to apply in a large number of scientific disciplines over the past two decades. Among them, the discipline of food and pharmaceutical formulation encountered several obstacles when combining microfluidics with aqueous media. Indeed, the physical properties of liquids at micrometric volumes being particular, the droplet generation within microfluidic devices is a big challenge to be met. This focus review is intended to be an initiation for those who would like to generate microdroplets in microfluidic systems involving aqueous continuous phases. It provides a state-of-the-art look at such systems while focusing on the microfluidic devices used, their applications to form a wide variety of emulsions and particles, and the key role held by the interface between the device channels and the emulsion. This review also leads to reflections on new materials that can be used in microfluidic systems with aqueous continuous phases.

Vegetable oil hybrid films cross-linked at the air-water interface: formation kinetics and physical characterization.

Vegetable oil based hybrid films were developed thanks to a novel solvent- and heating-free method at the air-water interface using silylated castor oil cross-linked via a sol-gel reaction. To understand the mechanism of the hybrid film formation, the reaction kinetics was studied in detail by using complementary techniques: rheology, thermogravimetric analysis, and infrared spectroscopy. The mechanical properties of the final films were investigated using nano-indentation, whereas their structure was studied using a combination of wide-angle X-ray scattering, electron diffraction, and atomic force microscopy. We found that solid and transparent films form in 24 hours and, by changing the silica precursor to castor oil ratio, their mechanical properties are tunable in the MPa-range by about a factor of twenty. In addition to that, a possible optimization of the cross-linking reaction with different catalysts was explored, and finally cytotoxicity tests were performed on fibroblasts proving the absence of film toxicity. The results of this work pave the way to a straightforward synthesis of castor-oil films with tunable mechanical properties: hybrid films cross-linked at the air-water interface combine an easy and cheap spreading protocol with the features of their thermal history optimized for possible future micro/nano drug loading, thus representing excellent candidates for the replacement of non-environmentally friendly petroleum-based materials.

Properties of dentin, enamel and their junction, studied with Brillouin scattering and compared to Raman microscopy.

OBJECTIVE: Dentin, enamel and the transition zone, called the dentin-enamel junction (DEJ), have an organization and properties that play a critical role in tooth resilience and in stopping the propagation of cracks. Understanding their chemical and micro-biomechanical properties is then of foremost importance. The aim of this study is to apply Brillouin microscopy on a complex biological structure, that is, the DEJ, and to compare these results with those obtained with Raman microscopy. DESIGN: Both techniques allow noncontact measurements at the microscopic scale. Brillouin microscopy is based on the interaction between acoustic phonons and laser photons and gives a relation between the frequency shift of the scattered light and the stiffness of the sample. Raman spectra contain peaks related to specific chemical bonds. RESULTS: Comparison of the Brillouin and Raman cartographies reveals correlations between mechanical and chemical properties. Indeed, the shapes of the phosphate content and stiffness curves are similar. The two spectroscopies give compatible values for the mean distance between two tubules, i.e., 4-6 µm. Moreover, for the first time, the daily cross striations of enamel could be studied, indicating a relationship between the variation in the phosphate concentration and the variation in the rigidity within the enamel prisms. CONCLUSIONS: We demonstrate here the possibility of using Brillouin scattering microscopy to both study complex biological materials such as the enamel-dentin junction and visualize secondary structures. Correlations between the chemical composition and mechanical properties could help in better understanding the tissue histology.

A putative transporter is essential for integrating nutrient and hormone signaling with lateral root growth and nodule development in Medicago truncatula.

Legume root architecture involves not only elaboration of the root system by the formation of lateral roots but also the formation of symbiotic root nodules in association with nitrogen-fixing soil rhizobia. The Medicago truncatula LATD/NIP gene plays an essential role in the development of both primary and lateral roots as well as nodule development. We have cloned the LATD/NIP gene and show that it encodes a member of the NRT1(PTR) transporter family. LATD/NIP is expressed throughout the plant. pLATD/NIP-GFP promoter-reporter fusions in transgenic roots establish the spatial expression of LATD/NIP in primary root, lateral root and nodule meristems and the surrounding cells. Expression of LATD/NIP is regulated by hormones, in particular by abscisic acid which has been previously shown to rescue the primary and lateral root meristem arrest of latd mutants. latd mutants respond normally to ammonium but have defects in responses of the root architecture to nitrate. Taken together, these results suggest that LATD/NIP may encode a nitrate transporter or transporter of another compound.

IPD3 controls the formation of nitrogen-fixing symbiosomes in pea and Medicago Spp.

A successful nitrogen-fixing symbiosis requires the accommodation of rhizobial bacteria as new organelle-like structures, called symbiosomes, inside the cells of their legume hosts. Two legume mutants that are most strongly impaired in their ability to form symbiosomes are sym1/TE7 in Medicago truncatula and sym33 in Pisum sativum. We have cloned both MtSYM1 and PsSYM33 and show that both encode the recently identified interacting protein of DMI3 (IPD3), an ortholog of Lotus japonicus (Lotus) CYCLOPS. IPD3 and CYCLOPS were shown to interact with DMI3/CCaMK, which encodes a calcium- and calmodulin-dependent kinase that is an essential component of the common symbiotic signaling pathway for both rhizobial and mycorrhizal symbioses. Our data reveal a novel, key role for IPD3 in symbiosome formation and development. We show that MtIPD3 participates in but is not essential for infection thread formation and that MtIPD3 also affects DMI3-induced spontaneous nodule formation upstream of cytokinin signaling. Further, MtIPD3 appears to be required for the expression of a nodule-specific remorin, which controls proper infection thread growth and is essential for symbiosome formation.

The Rise of the OM-LoC: Opto-Microfluidic Enabled Lab-on-Chip.

The integration of optical circuits with microfluidic lab-on-chip (LoC) devices has resulted in a new era of potential in terms of both sample manipulation and detection at the micro-scale. On-chip optical components increase both control and analytical capabilities while reducing reliance on expensive laboratory photonic equipment that has limited microfluidic development. Notably, in-situ LoC devices for bio-chemical applications such as diagnostics and environmental monitoring could provide great value as low-cost, portable and highly sensitive systems. Multiple challenges remain however due to the complexity involved with combining photonics with micro-fabricated systems. Here, we aim to highlight the progress that optical on-chip systems have made in recent years regarding the main LoC applications: (1) sample manipulation and (2) detection. At the same time, we aim to address the constraints that limit industrial scaling of this technology. Through evaluating various fabrication methods, material choices and novel approaches of optic and fluidic integration, we aim to illustrate how optic-enabled LoC approaches are providing new possibilities for both sample analysis and manipulation.

Cultivar Grain Yield in Durum Wheat-Grain Legume Intercrops Could Be Estimated From Sole Crop Yields and Interspecific Interaction Index.

Ensuring food security for a world population projected to reach over nine billion by 2050 while mitigating the environmental impacts and climate change represent the major agricultural challenges. Diversification of the cropping systems using notably cereal-legume mixtures is one key pathway for such agroecological intensification. Indeed, intercropping is recognised as a practice having the potential to increase and stabilise the yields in comparison with sole crops while limiting the use of inputs notably when species exploit resources in a complementary way. However, predicting intercropped species grain yield remains a challenge because the species respond to competition through complex genotype x cropping mode interactions. Here, we hypothesised that the grain yield achieved by a cultivar in low nitrogen input durum wheat-grain legume intercrops (ICs) could be estimated using a few simple variables. The present work is based on a 2-year field experiment carried out in southwestern France using two durum wheat (Triticum turgidum L.), four winter pea (Pisum sativum L.), and four winter faba bean (Vicia faba L.) genotypes with contrasting characteristics, notably in terms of height and precocity, to explore a wide range of durum wheat-grain legume phenotypes combinations to generate variability in terms of yield and species proportion. The major result is that the yield of durum wheat-grain legume IC component in low nitrogen input conditions could be correctly estimated from only three variables: (i) wheat cultivar full density sole crop (SC) yield, (ii) legume cultivar half density sole crop (SC½) yield, and (iii) an indicator of legume cultivar response to interspecific competition. The latter variable, the interspecific interaction index (IE), reveals cultivars' competitive abilities and tolerance to competition. However, to propose generic IC design and management procedures, further mechanistic understanding is required to better understand the links between tolerance to interspecific competition and cultivar phenotype characteristics. In particular, a special emphasis on the grain legume is needed as their response to interspecific competition appears less predictable than that of durum wheat. Cultivar choice is a key element to optimise the functional complementarity and subsequent IC advantages. This work proposes a simple tool to assist the design of specific breeding programs for cultivars ideotypes adapted to intercropping.

Adaptation of Medicago truncatula to nitrogen limitation is modulated via local and systemic nodule developmental responses.

Adaptation of Medicago truncatula to local nitrogen (N) limitation was investigated to provide new insights into local and systemic N signaling. The split-root technique allowed a characterization of the local and systemic responses of NO(3)(-) or N(2)-fed plants to localized N limitation. (15)N and (13)C labeling were used to monitor plant nutrition. Plants expressing pMtENOD11-GUS and the sunn-2 hypernodulating mutant were used to unravel mechanisms involved in these responses. Unlike NO(3)(-)-fed plants, N(2)-fixing plants lacked the ability to compensate rapidly for a localized N limitation by up-regulating the N(2)-fixation activity of roots supplied elsewhere with N. However they displayed a long-term response via a growth stimulation of pre-existing nodules, and the generation of new nodules, likely through a decreased abortion rate of early nodulation events. Both these responses involve systemic signaling. The latter response is abolished in the sunn mutant, but the mutation does not prevent the first response. Local but also systemic regulatory mechanisms related to plant N status regulate de novo nodule development in Mt, and SUNN is required for this systemic regulation. By contrast, the stimulation of nodule growth triggered by systemic N signaling does not involve SUNN, indicating SUNN-independent signaling.

Fractal Structure in Silica and Composites Aerogels.

Silica aerogels are known to be materials with exceptional characteristics, such as ultra-low density, high surface area, high porosity, high adsorption, and low-thermal conductivity. In addition, these unique properties are mainly related to their specific processing. Depending on the aerogel synthesis procedure, the aerogels texture can be tailored with meso and/or macroporosity. Fractal geometry has been observed and used to describe silica aerogels at nanoscales in certain conditions. In this review paper, we describe the fractal structure of silica aerogels that can develop depending on the synthesis conditions. X-ray and neutron scattering measurements allow to show that silica aerogels can exhibit a fractal structure over one or even more than two orders of magnitude in length. The fractal dimension does not depend directly on the material density but can vary with the synthesis conditions. It ranges typically between 1.6 and 2.4. The effect of the introduction of silica particles or of further thermal treatment or compression of the silica aerogels on their microstructure and their fractal characteristics is also resumed.

Developmental Modulation of Root Cell Wall Architecture Confers Resistance to an Oomycete Pathogen.

The cell wall is the primary interface between plant cells and their immediate environment and must balance multiple functionalities, including the regulation of growth, the entry of beneficial microbes, and protection against pathogens. Here, we demonstrate how API, a SCAR2 protein component of the SCAR/WAVE complex, controls the root cell wall architecture important for pathogenic oomycete and symbiotic bacterial interactions in legumes. A mutation in API results in root resistance to the pathogen Phytophthora palmivora and colonization defects by symbiotic rhizobia. Although api mutant plants do not exhibit significant overall growth and development defects, their root cells display delayed actin and endomembrane trafficking dynamics and selectively secrete less of the cell wall polysaccharide xyloglucan. Changes associated with a loss of API establish a cell wall architecture with altered biochemical properties that hinder P. palmivora infection progress. Thus, developmental stage-dependent modifications of the cell wall, driven by SCAR/WAVE, are important in balancing cell wall developmental functions and microbial invasion.

Evidence for the involvement in nodulation of the two small putative regulatory peptide-encoding genes MtRALFL1 and MtDVL1.

Nod factors are key bacterial signaling molecules regulating the symbiotic interaction between bacteria known as rhizobia and leguminous plants. Studying plant host genes whose expression is affected by Nod factors has given insights into early symbiotic signaling and development. Here, we used a double supernodulating mutant line that shows increased sensitivity to Nod factors to study the Nod factor-regulated transcriptome. Using microarrays containing more than 16,000 70-mer oligonucleotide probes, we identified 643 Nod-factor-regulated genes, including 225 new Nod-factor-upregulated genes encoding many potential regulators. Among the genes found to be Nod factor upregulated, we identified and characterized MtRALFL1 and MtDVL1, which code for two small putative peptide regulators of 135 and 53 amino acids, respectively. Expression analysis confirmed that these genes are upregulated during initial phases of nodulation. Overexpression of MtRALFL1 and MtDVL1 in Medicago truncatula roots resulted in a marked reduction in the number of nodules formed and in a strong increase in the number of aborted infection threads. In addition, abnormal nodule development was observed when MtRALFL1 was overexpressed. This work provides evidence for the involvement of new putative small-peptide regulators during nodulation.

api, A novel Medicago truncatula symbiotic mutant impaired in nodule primordium invasion.

Genetic approaches have proved to be extremely useful in dissecting the complex nitrogen-fixing Rhizobium-legume endosymbiotic association. Here we describe a novel Medicago truncatula mutant called api, whose primary phenotype is the blockage of rhizobial infection just prior to nodule primordium invasion, leading to the formation of large infection pockets within the cortex of noninvaded root outgrowths. The mutant api originally was identified as a double symbiotic mutant associated with a new allele (nip-3) of the NIP/LATD gene, following the screening of an ethylmethane sulphonate-mutagenized population. Detailed characterization of the segregating single api mutant showed that rhizobial infection is also defective at the earlier stage of infection thread (IT) initiation in root hairs, as well as later during IT growth in the small percentage of nodules which overcome the primordium invasion block. Neither modulating ethylene biosynthesis (with L-alpha-(2-aminoethoxyvinylglycine or 1-aminocyclopropane-1-carboxylic acid) nor reducing ethylene sensitivity in a skl genetic background alters the basic api phenotype, suggesting that API function is not closely linked to ethylene metabolism or signaling. Genetic mapping places the API gene on the upper arm of the M. truncatula linkage group 4, and epistasis analyses show that API functions downstream of BIT1/ERN1 and LIN and upstream of NIP/LATD and the DNF genes.

Purification of Nongreen Plastids (Proplastids and Amyloplasts) from Angiosperms, and Isolation of Their Envelope Membranes.

Plastids, a wide family of plant specific organelles, exist in all plant cells in a number of different forms with different functions essential for plant life. Among them, chloroplasts are by far the more extensively studied owing to their central role in photosynthesis. However, other plastid family members, often referred to as nongreen plastids, play also major roles in the physiology of higher plants and could be better suited for studies of specific metabolic processes in heterotrophic plant cells. Unfortunately, serious technical problems are frequently encountered with separating intact, active nongreen plastids from contaminating membranes and mitochondria. Here, we provide detailed protocols suitable for the large scale preparation of intact and highly pure proplastids from cauliflower buds, as well as amyloplasts from sycamore cultured cells, and for the subsequent separation of their surrounding envelope membranes from the stroma and other plastid fractions. Both methods proved to be highly reliable and have been instrumental for in-depth investigations on biochemistry and physiology of nongreen plastids.