Catherine Santaella

Root Border-Like Cells of Arabidopsis. Microscopical Characterization and Role in the Interaction with Rhizobacteria

Plant roots of many species produce thousands of cells that are released daily into the rhizosphere. These cells are commonly termed border cells because of their major role in constituting a biotic boundary layer between the root surface and the soil. In this study, we investigated the occurrence and ultrastructure of such cells in Arabidopsis (Arabidopsis thaliana) using light and electron microscopy coupled to high-pressure freezing. The secretion of cell wall molecules including pectic polysaccharides and arabinogalactan-proteins (AGPs) was examined also using immunofluorescence microscopy and a set of anticarbohydrate antibodies. We show that root tips of Arabidopsis seedlings released cell layers in an organized pattern that differs from the rather randomly dispersed release observed in other plant species studied to date. Therefore, we termed such cells border-like cells (BLC). Electron microscopical results revealed that BLC are rich in mitochondria, Golgi stacks, and Golgi-derived vesicles, suggesting that these cells are actively engaged in secretion of materials to their cell walls. Immunocytochemical data demonstrated that pectins as well as AGPs are among secreted material as revealed by the high level of expression of AGP-epitopes. In particular, the JIM13-AGP epitope was found exclusively associated with BLC and peripheral cells in the root cap region. In addition, we investigated the function of BLC and root cap cell AGPs in the interaction with rhizobacteria using AGP-disrupting agents and a strain of Rhizobium sp. expressing a green fluorescent protein. Our findings demonstrate that alteration of AGPs significantly inhibits the attachment of the bacteria to the surface of BLC and root tip.

The exopolysaccharide of Rhizobium sp. YAS34 is not necessary for biofilm formation on Arabidopsis thaliana and Brassica napus roots but contributes to root colonization

Microbial exopolysaccharides (EPSs) play key roles in plant–microbe interactions, such as biofilm formation on plant roots and legume nodulation by rhizobia. Here, we focused on the function of an EPS produced by Rhizobium sp. YAS34 in the colonization and biofilm formation on non-legume plant roots (Arabidopsis thaliana and Brassica napus). Using random transposon mutagenesis, we isolated an EPS-deficient mutant of strain YAS34 impaired in a glycosyltransferase gene (gta). Wild type and mutant strains were tagged with a plasmid-born GFP and, for the first time, the EPS produced by the wild-type strain was seen in the rhizosphere using selective carbohydrate probing with a fluorescent lectin and confocal laser-scanning microscopy. We show for the fist time that Rhizobium forms biofilms on roots of non-legumes, independently of the EPS synthesis. When produced by strain YAS34 wild type, EPS is targeted at specific parts of the plant root system. Nutrient fluctuations, root exudates and bacterial growth phase can account for such a production pattern. The EPS synthesis in Rhizobium sp. YAS34 is not essential for biofilm formation on roots, but is critical to colonization of the basal part of the root system and increasing the stability of root-adhering soil. Thus, in Rhizobium sp. YAS34 and non-legume interactions, microbial EPS is implicated in root–soil interface, root colonization, but not in biofilm formation.

Rhizobium alamii sp. nov., an exopolysaccharide-producing species isolated from legume and non-legume rhizospheres.

A group of exopolysaccharide-producing bacteria was isolated from the root environment of Arabidopsis thaliana. The genetic diversity revealed by REP-PCR fingerprinting indicated that the isolates correspond to different strains. 16S rRNA gene sequence analysis showed that the isolates are closely related to the strains Rhizobium sp. YAS34 and USDA 1920, respectively isolated from sunflower roots and Medicago ruthenica nodules. These bacteria belong to the Rhizobium lineage of the Alphaproteobacteria, and the closest known species was Rhizobium sullae. DNA-DNA hybridization experiments and biochemical analysis demonstrated that the nine strains isolated from A. thaliana and Rhizobium strains YAS34 and USDA 1920 constitute a novel species within the genus Rhizobium, for which the name Rhizobium alamii sp. nov. is proposed. The type strain is GBV016(T) (=CFBP 7146(T) =LMG 24466(T)).

Transfer, Transformation, and Impacts of Ceria Nanomaterials in Aquatic Mesocosms Simulating a Pond Ecosystem

Mesocosms are an invaluable tool for addressing the complex issue of exposure during nanoecotoxicological testing. This experimental strategy was used to take into account parameters as the interactions between the NPs and naturally occurring (in)organic colloids (heteroaggregation), or the flux between compartments of the ecosystems (aqueous phase, sediments, biota) when assessing the impacts of CeO2 NPs in aquatic ecosystems. In this study, we determine the transfer, redox transformation, and impacts of 1 mg L(-1) of bare and citrate coated CeO2-NPs toward an ecologically relevant organism (snail, Planorbarius corneus) exposed 4 weeks in a complex experimental system mimicking a pond ecosystem. Over time, CeO2-NPs tend to homo- and heteroaggregate and to accumulate on the surficial sediment. The kinetic of settling down was coating-dependent and related to the coating degradation. After 4 weeks, Ce was observed in the digestive gland of benthic organisms and associated with 65-80% of Ce(IV) reduction into Ce(III) for both bare and coated CeO2 NPs. A transitory oxidative stress was observed for bare CeO2-NPs. Coated-NPs exposed snails did not undergo any lipid peroxidation nor change in the antioxidant contents, while Ce content and reduction in the digestive gland were identical to bare CeO2-NPs. We hypothesized that the presence of citrate coating enhanced the defense capacity of the cells toward the oxidative stress induced by the CeO2 core.

Fate of Ag-NPs in Sewage Sludge after Application on Agricultural Soils

The objective of this work was to investigate the fate of silver nanoparticles (Ag-NPs) in a sludge-amended soil cultivated with monocot (Wheat) and dicot (Rape) crop species. A pot experiment was performed with sludges produced in a pilot wastewater treatment plant containing realistic Ag concentrations (18 and 400 mg kg(-1), 14 mg kg(-1) for the control). Investigations focused on the highest dose treatment. X-ray absorption spectroscopy (XAS) showed that Ag2S was the main species in the sludge and amended soil before and after plant culture. The second most abundant species was an organic and/or amorphous Ag-S phase whose proportion slightly varied (from 24% to 36%) depending on the conditions. Micro and nano X-ray fluorescence (XRF) showed that Ag was preferentially associated with S-rich particles, including organic fragments, of the sludge and amended soils. Ag was distributed as heteroaggregates with soil components (size ranging from ≤0.5 to 1-3 μm) and as diffused zones likely corresponding to sorbed/complexed Ag species. Nano-XRF evidenced the presence of mixed metallic sulfides. Ag was weakly exchangeable and labile. However, micronutrient mobilization by plant roots and organic matter turnover may induce Ag species interconversion eventually leading to Ag release on longer time scales. Together, these data provide valuable information for risk assessment of sewage sludge application on agricultural soils.

Complete Genome Sequence of a Beneficial Plant Root-Associated Bacterium, Pseudomonas brassicacearum

To shed light on the genetic equipment of the beneficial plant-associated bacterium Pseudomonas brassicacearum, we sequenced the whole genome of the strain NFM421. Its genome consists of one chromosome equipped with a repertoire of factors beneficial for plant growth. In addition, a complete type III secretion system and two complete type VI secretion systems were identified. We report here the first genome sequence of this species.

Transformation of pristine and citrate-functionalized CeO2 nanoparticles in a laboratory-scale activated sludge reactor.

Engineered nanomaterials (ENMs) are used to enhance the properties of many manufactured products and technologies. Increased use of ENMs will inevitably lead to their release into the environment. An important route of exposure is through the waste stream, where ENMs will enter wastewater treatment plants (WWTPs), undergo transformations, and be discharged with treated effluent or biosolids. To better understand the fate of a common ENM in WWTPs, experiments with laboratory-scale activated sludge reactors and pristine and citrate-functionalized CeO2 nanoparticles (NPs) were conducted. Greater than 90% of the CeO2 introduced was observed to associate with biosolids. This association was accompanied by reduction of the Ce(IV) NPs to Ce(III). After 5 weeks in the reactor, 44 ± 4% reduction was observed for the pristine NPs and 31 ± 3% for the citrate-functionalized NPs, illustrating surface functionality dependence. Thermodynamic arguments suggest that the likely Ce(III) phase generated would be Ce2S3. This study indicates that the majority of CeO2 NPs (>90% by mass) entering WWTPs will be associated with the solid phase, and a significant portion will be present as Ce(III). At maximum, 10% of the CeO2 will remain in the effluent and be discharged as a Ce(IV) phase, governed by cerianite (CeO2).

P2CS: updates of the prokaryotic two-component systems database

The P2CS database (http://www.p2cs.org/) is a comprehensive resource for the analysis of Prokaryotic Two-Component Systems (TCSs). TCSs are comprised of a receptor histidine kinase (HK) and a partner response regulator (RR) and control important prokaryotic behaviors. The latest incarnation of P2CS includes 164 651 TCS proteins, from 2758 sequenced prokaryotic genomes. Several important new features have been added to P2CS since it was last described. Users can search P2CS via BLAST, adding hits to their cart, and homologous proteins can be aligned using MUSCLE and viewed using Jalview within P2CS. P2CS also provides phylogenetic trees based on the conserved signaling domains of the RRs and HKs from entire genomes. HK and RR trees are annotated with gene organization and domain architecture, providing insights into the evolutionary origin of the contemporary gene set. The majority of TCSs are encoded by adjacent HK and RR genes, however, ‘orphan’ unpaired TCS genes are also abundant and identifying their partner proteins is challenging. P2CS now provides paired HK and RR trees with proteins from the same genetic locus indicated. This allows the appraisal of evolutionary relationships across entire TCSs and in some cases the identification of candidate partners for orphan TCS proteins.

Deciphering the Responses of Root Border-Like Cells of Arabidopsis and Flax to Pathogen-Derived Elicitors

Root border-like cells of flax and Arabidopsis activate innate immunity responses to elicitors involving both callose deposition and cell wall extensin reorganization. Plant pathogens including fungi and bacteria cause many of the most serious crop diseases. The plant innate immune response is triggered upon recognition of microbe-associated molecular patterns (MAMPs) such as flagellin22 and peptidoglycan. To date, very little is known of MAMP-mediated responses in roots. Root border cells are cells that originate from root caps and are released individually into the rhizosphere. Root tips of Arabidopsis (Arabidopsis thaliana) and flax (Linum usitatissimum) release cells known as “border-like cells.” Whereas root border cells of pea (Pisum sativum) are clearly involved in defense against fungal pathogens, the function of border-like cells remains to be established. In this study, we have investigated the responses of root border-like cells of Arabidopsis and flax to flagellin22 and peptidoglycan. We found that both MAMPs triggered a rapid oxidative burst in root border-like cells of both species. The production of reactive oxygen species was accompanied by modifications in the cell wall distribution of extensin epitopes. Extensins are hydroxyproline-rich glycoproteins that can be cross linked by hydrogen peroxide to enhance the mechanical strength of the cell wall. In addition, both MAMPs also caused deposition of callose, a well-known marker of MAMP-elicited defense. Furthermore, flagellin22 induced the overexpression of genes involved in the plant immune response in root border-like cells of Arabidopsis. Our findings demonstrate that root border-like cells of flax and Arabidopsis are able to perceive an elicitation and activate defense responses. We also show that cell wall extensin is involved in the innate immunity response of root border-like cells.

Aged TiO2-Based Nanocomposite Used in Sunscreens Produces Singlet Oxygen under Long-Wave UV and Sensitizes Escherichia coli to Cadmium

TiO2-based nanocomposite (NC) are widely used as invisible UV protectant in cosmetics. These nanomaterials (NMs) end in the environment as altered materials. We have investigated the properties of T-Lite SF, a TiO2-NC used as sunscreen, after weathering in water and under light. We have examined the formation of ROS and their consequences on cell physiology of Escherichia coli. Our results show that aged-T-Lite SF produced singlet oxygen under low intensity long wave UV and formed hydroxyl radicals at high intensity. Despite the production of these ROS, T-Lite SF had neither effect on the viability of E. coli nor on mutant impaired in oxidative stress, did not induce mutagenesis and did not impair the integrity of membrane lipids, thus seemed safe to bacteria. However, when pre-exposed to T-Lite SF under low intensity UV, cells turned out to be more sensitive to cadmium, a priority pollutant widely disseminated in soil and surface waters. This effect was not a Trojan horse: sensitization of cells was dependent on the formation of singlet oxygen. These results provide a basis for caution, especially on NMs that have no straight environmental toxicity. It is crucial to anticipate indirect and combined effects of environmental pollutants and NMs.