Biocontrol of Biofilm Formation: Jamming of Sessile-Associated Rhizobial Communication by Rhodococcal Quorum-Quenching.

Biofilms are complex structures formed by a community of microbes adhering to a surface and/or to each other through the secretion of an adhesive and protective matrix. The establishment of these structures requires a coordination of action between microorganisms through powerful communication systems such as quorum-sensing. Therefore, auxiliary bacteria capable of interfering with these means of communication could be used to prevent biofilm formation and development. The phytopathogen Rhizobium rhizogenes, which causes hairy root disease and forms large biofilms in hydroponic crops, and the biocontrol agent Rhodococcus erythropolis R138 were used for this study. Changes in biofilm biovolume and structure, as well as interactions between rhizobia and rhodococci, were monitored by confocal laser scanning microscopy with appropriate fluorescent biosensors. We obtained direct visual evidence of an exchange of signals between rhizobia and the jamming of this communication by Rhodococcus within the biofilm. Signaling molecules were characterized as long chain (C14) N-acyl-homoserine lactones. The role of the Qsd quorum-quenching pathway in biofilm alteration was confirmed with an R. erythropolis mutant unable to produce the QsdA lactonase, and by expression of the qsdA gene in a heterologous host, Escherichia coli. Finally, Rhizobium biofilm formation was similarly inhibited by a purified extract of QsdA enzyme.

Selenium Nanocomposites in Natural Matrices as Potato Recovery Agent.

The paper presents a study of the effect of chemically synthesized selenium nanocomposites (Se NCs) in natural polymer matrices arabinogalactan (AG) and starch (ST) on the viability of the potato ring rot pathogen Clavibacter sepedonicus (Cms), potato plants in vitro, and the soil bacterium Rhodococcus erythropolis. It was found that the studied Se NCs have an antibacterial effect against the phytopathogenic Cms, reducing its growth rate and ability to form biofilms. It was revealed that Se NC based on AG (Se/AG NC) stimulated the growth and development of potato plants in vitro as well as their root formation. At the same time, Se did not accumulate in potato tissues after the treatment of plants with Se NCs. The safety of the Se NCs was also confirmed by the absence of a negative effect on the growth and biofilm formation of the soil bacterium R. erythropolis. The obtained results indicate that Se NCs are promising environmentally safe agents for the protection and recovery of cultivated plants from phytopathogenic bacteria.

Optimisation of biodegradation conditions for waste canola oil by cold-adapted Rhodococcus sp. AQ5-07 from Antarctica

BACKGROUND: The potential waste canola oil-degrading ability of the cold-adapted Antarctic bacterial strain Rhodococcus sp. AQ5-07 was evaluated. Globally, increasing waste from food industries generates serious anthropogenic environmental risks that can threaten terrestrial and aquatic organisms and communities. The removal of oils such as canola oil from the environment and wastewater using biological approaches is desirable as the thermal process of oil degradation is expensive and ineffective. RESULTS: Rhodococcus sp. AQ5-07 was found to have high canola oil-degrading ability. Physico-cultural conditions influencing its activity were studied using one-factor-at-a-time (OFAT) and statistical optimisation approaches. Considerable degradation (78.60%) of 3% oil was achieved by this bacterium when incubated with 1.0 g/L ammonium sulphate, 0.3 g/L yeast extract, pH 7.5 and 10% inoculum at 10°C over a 72-h incubation period. Optimisation of the medium conditions using response surface methodology (RSM) resulted in a 9.01% increase in oil degradation (87.61%) when supplemented with 3.5% canola oil, 1.05 g/L ammonium sulphate, 0.28g/L yeast extract, pH 7.5 and 10% inoculum at 12.5°C over the same incubation period. The bacterium was able to tolerate an oil concentration of up to 4.0%, after which decreased bacterial growth and oil degradation were observed. CONCLUSIONS: These features make this strain worthy of examination for practical bioremediation of lipid-rich contaminated sites. This is the first report of any waste catering oil degradation by bacteria originating from Antarctica.

A sustained CYCLINB1;1 and STM expression in the neoplastic tissues induced by Rhodococcus fascians on Arabidopsis underlies the persistence of the leafy gall structure.

RHODOCOCCUS FASCIANS: is a gram-positive phytopathogen that infects a wide range of plant species. The actinomycete induces the formation of neoplastic growths, termed leafy galls, that consist of a gall body covered by small shoots of which the outgrowth is arrested due to an extreme form of apical dominance. In our previous work, we demonstrated that in the developing gall, auxin drives the transdifferentiation of parenchyma cells into vascular elements. In this work, with the use of transgenic Arabidopsis thaliana plants carrying molecular reporters for cell division (pCYCB1;1:GUS) and meristematic activity (pSTM:GUS), we analyzed the fate of cells within the leafy gall. Our results indicate that the size of the gall body is determined by ongoing mitotic cell divisions as illustrated by strong CYCB1;1 expression combined with the de novo formation of new meristematic areas triggered by STM expression. The shoot meristems that develop in the peripheral parts of the gall are originating from high ectopic STM expression. Altogether the presented data provide further insight into the cellular events that accompany the development of leafy galls in response to R. fascians infection.

Improving soybean growth under arsenic stress by inoculation with native arsenic-resistant bacteria.

Certain metal (loid)-resistant bacteria that inhabit the rhizosphere have shown to improve plant growth and tolerance under toxic metal stress. In this study, we tested if six native, arsenic-resistant and plant growth promoting bacteria (PGPB) were able to enhance soybean (Glycine max L.) growth and modulate arsenic (As) uptake. As a previous work, we tested all single isolates and all possible binary combinations without arsenic stress to identify the combinations that would have the greatest plant growth promoting effect. In this study, a screening assay was performed with only five inoculation options selected after first stage (Pseudomonas sp. AW4, Pseudomonas sp. AW6, AW4+AW6, Rhodococcus sp. AW3+Pseudomonas sp. AW5 and Enterobacter sp. AW1+AW6). In both stages, inoculation was implemented by imbibition of soybean seeds with bacterial suspensions, and plant growth was carried out in pots using perlite as substrate in a chamber with controlled conditions. In the third stage, we performed similar assays, under As stress, using the three most promising inoculation options (AW4, AW6 and AW3+AW5). Treatments were performed by irrigation with 25 µM arsenite (As3+), 25 µM arsenate (As5+), 25 µM equimolar As3+/As5+ solution or water (control). Biometric and biochemical parameters indicated that inoculation with Pseudomonas sp. AW4 significantly promoted soybean growth under As3+/As5+ treatment and did not modified As accumulation pattern. Further field studies are needed to determine if some of these inoculation options are useful to improve in situ soybean growth under arsenic stress and could become a tool for the development of sustainable agriculture in As-impacted environments.

Effect of rhamnolipid on the physicochemical properties and interaction of bacteria and fungi.

Bacterial adhesion on surfaces is an essential initial step in promoting bacterial mobilization for soil bioremediation process. Modification of the cell surface is required to improve the adhesion of bacteria. The modification of physicochemical properties by rhamnolipid to Pseudomonas putida KT2442, Rhodococcus erythropolis 3586 and Aspergillus brasiliensis ATCC 16404 strains was analysed using contact angle measurements. The surface energy and total free energy of adhesion were calculated to predict the adhesion of both bacteria strains on the A. brasiliensis surface. The study of bacterial adhesion was carried out to evaluate experimental value with the theoretical results. Bacteria and fungi physicochemical properties were modified significantly when treated with rhamnolipid. The adhesion rate of P. putida improved by 16% with the addition of rhamnolipid (below 1 CMC), while the increase of rhamnolipid concentration beyond 1 CMC did not further enhance the bacterial adhesion. The addition of rhamnolipid did not affect the adhesion of R. erythropolis. A good relationship has been obtained in which water contact angle and surface energy of fungal surfaces are the major factors contributing to the bacterial adhesion. The adhesion is mainly driven by acid-base interaction. This finding provides insight to the role of physicochemical properties in controlling the bacterial adhesion on the fungal surface to enhance bacteria transport in soil bioremediation.

Close-up on a bacterial informational war in the geocaulosphere.

The geocaulosphere is home to microbes that establish communication between themselves and others that disrupt them. These cell-to-cell communication systems are based on the synthesis and perception of signaling molecules, of which the best known belong to the N-acyl-homoserine lactone (AHL) family. Among indigenous bacteria, certain Gram-positive actinobacteria can sense AHLs produced by soft-rot Gram-negative phytopathogens and can degrade the quorum-sensing AHL signals to impair the expression of virulence factors. We mimicked this interaction by introducing dual-color reporter strains suitable for monitoring both the location of the cells and their quorum-sensing and -quenching activities, in potato tubers. The exchange of AHL signals within the pathogen's cell quorum was clearly detected by the presence of bright green fluorescence instead of blue in a portion of Pectobacterium-tagged cells. This phenomenon in Rhodococcus cells was accompanied by a change from red fluorescence to orange, showing that the disappearance of signaling molecules is due to rhodococcal AHL degradation rather than the inhibition of AHL production. Rhodococci are victorious in this fight for the control of AHL-based communication, as their jamming activity is powerful enough to prevent the onset of disease symptoms.

Influence of pressure and dispersant on oil biodegradation by a newly isolated Rhodococcus strain from deep-sea sediments of the gulf of Mexico.

A new Rhodococcus strain, capable of degrading crude oil, was isolated from the Gulf of Mexico deep-sea sediment and was investigated for its biodegradation characteristics under atmospheric as well as under deep-sea pressure (1500 m = 15 MPa). Additionally, the effect of dispersant (Corexit EC9500A) addition was studied. Rhodococcus sp. PC20 was shown to degrade 60.5 ±â€¯10.7% of the saturated and aromatic fraction of crude oil at atmospheric pressure and 74.2 ±â€¯9.1% at deep-sea level pressure within 96 h. Degradation rates, especially for monoaromatic hydrocarbons, were significantly higher at elevated pressure compared to atmospheric pressure. This study found a growth inhibiting effect at a dispersant to oil ratio of 1:100 and higher. This effect of the dispersant was enhanced when elevated pressure was applied.

Phenotypic Adaptations Help Rhodococcus erythropolis Cells during the Degradation of Paraffin Wax.

During crude oil extraction, the reduction in temperature and pressure results in the precipitation of paraffin wax that contains 20-40 carbon chain hydrocarbons. The paraffin wax may accumulate inside production tubes, pipelines, and processing facilities, and also in tankers during petroleum transportation. There are few bacterial strains that are able to degrade solid substrates. In the present study, the biodegradation of paraffin is evaluated using Rhodococcus erythropolis cells. This bacterium is able to grow using paraffin wax from an oil refinery plant as the sole carbon source. The cells grow as a thick biofilm over the solid substrate, make scale-like structures that increase the area of the initially smooth surface of paraffin, produce biosurfactants, and become more negatively charged than ethanol- or glucose-grown cells. When paraffin wax is supplied as microparticles, to increase the cell-substrate contact area and to simulate paraffin precipitation, the cells also adjust the composition of the fatty acids of the phospholipids of the cellular membrane to decrease its fluidity and paraffin biodegradation increases considerably. The study suggests that the phenotypic adaptation of R. erythropolis cells may be used to degrade paraffin wax under real conditions.

Biocontrol of Soft Rot: Confocal Microscopy Highlights Virulent Pectobacterial Communication and Its Jamming by Rhodococcal Quorum-Quenching.

Confocal laser-scanning microscopy was chosen to observe the colonization and damage caused by the soft rot Pectobacterium atrosepticum and the protection mediated by the biocontrol agent Rhodococcus erythropolis. We developed dual-color reporter strains suited for monitoring quorum-sensing and quorum-quenching activities leading to maceration or biocontrol, respectively. A constitutively expressed cyan or red fluorescent protein served as a cell tag for plant colonization, while an inducible expression reporter system based on the green fluorescent protein gene enabled the simultaneous recording of signaling molecule production, detection, or degradation. The dual-colored pathogen and biocontrol strains were used to coinoculate potato tubers. At cellular quorum, images revealed a strong pectobacterial quorum-sensing activity, especially at the plant cell walls, as well as a concomitant rhodococcal quorum-quenching response, at both the single-cell and microcolony levels. The generated biosensors appear to be promising and complementary tools useful for molecular and cellular studies of bacterial communication and interference.

Biodegradation of high-molecular weight PAHs by Rhodococcus wratislaviensis strain 9: Overexpression of amidohydrolase induced by pyrene and BaP.

A Gram-positive bacterium, Rhodococcus wratislaviensis strain 9, completely degraded 280 µM of phenanthrene, 40% of 50 µM pyrene or 28% of 40 µM benzo[a]pyrene (BaP), each supplemented in M9 medium, within 7 days. PCR screening with gene-specific primers indicated that the strain 9 harbors genes which code for 2,3-dihydroxybiphenyl 1,2-dioxygenase (bphC), 4-nitrophenol 2-monooxygenase component B (npcB) as well as oxygenase component (nphA1), 4-hydroxybenzoate 3-monooxygenase (phbH), extradiol dioxygenase (edo), and naphthalene dioxygenase (ndo), all of which are largely implicated in biodegradation of several aromatic hydrocarbons. An orthogonal design experiment revealed that BaP biodegradation was greatly enhanced by surfactants such as Tween 80, Triton X-100 and linoleic acid, suggesting that bioavailability is the major limiting factor in bacterial metabolism of BaP. Both pyrene and BaP induced the overexpression of amidohydrolase, a metallo-dependent hydrolase, possibly involved in their biodegradation by strain 9. The up-regulation of amidohydrolase gene induced by BaP, in particular, was also confirmed by semi-quantitative RT-PCR. Catechol 2,3-dioxygenase and the large subunit of ndo, but not amidohydrolase, accumulated when the strain 9 was grown on phenanthrene. To our knowledge, this is the first report on overexpression of amidohydrolase and its possible implication in bacterial degradation of high-molecular weight PAHs.

Characterization of Rhodococcus sp. A5wh isolated from a high altitude Andean lake to unravel the survival strategy under lithium stress / Caracterización de las estrategias seguidas por Rhodococcus sp. A5wh proveniente de laguna de altura en su adaptación a la presencia de litio

Lithium (Li) is widely distributed in nature and has several industrial applications. The largest reserves of Li (over 85%) are in the so-called "triangle of lithium" that includes the Salar de Atacama in Chile, Salar de Uyuni in Bolivia and Salar del Hombre Muerto in Argentina. Recently, the use of microorganisms in metal recovery such as copper has increased; however, there is little information about the recovery of lithium. The strain Rhodococcus sp. A5wh used in this work was previously isolated from Laguna Azul. The assays revealed that this strain was able to accumulate Li (39.52% of Li/g microbial cells in 180 min) and that it was able to grow in its presence up to 1 M. In order to understand the mechanisms implicated in Li tolerance, a proteomic approach was conducted. Comparative proteomic analyses of strain A5wh exposed and unexposed to Li reveal that 17 spots were differentially expressed. The identification of proteins was performed by MALDI-TOF/MS, and the obtained results showed that proteins involved in stress response, transcription, translations, and metabolism were expressed under Li stress. This knowledge constitutes the first proteomic approach to elucidate the strategy followed by Rhodococcus to adapt to Li.

El litio (Li) es un elemento químico con múltiples aplicaciones industriales. Es considerado uno de los minerales más ampliamente distribuidos en la naturaleza. Sus mayores reservas (más del 85%) se encuentran en el llamado «triángulo de litio¼: salar de Atacama, en Chile; salar de Uyuni, en Bolivia, y salar del Hombre Muerto, en Argentina. En los últimos años, el empleo de microorganismos en la recuperación de metales se ha visto incrementado; sin embargo, hay muy poca información sobre la recuperación de Li por esta vía. En este estudio se trabajó con Rhodococcus sp. A5wh, cepa aislada de Laguna Azul. Los ensayos revelaron que este microorganismo fue capaz de acumular Li (39,52% de Li/g de biomasa en 180 min) y de crecer en presencia de este metaloide hasta una concentración de 1 M. Para comprender los mecanismos implicados en la tolerancia al Li, se llevó a cabo el análisis proteómico comparativo de esta cepa expuesta o no expuesta al Li. Los resultados revelaron 17 spots expresados en forma diferencial. La identificación de las proteínas se realizó porMALDI-TOF/MS. Este estudio constituye el primer enfoque proteómico para dilucidar la estrategia seguida por Rhodococcus en su adaptación al estrés.

Current taxonomy of Rhodococcus species and their role in infections.

Rhodococcus is a genus of obligate aerobic, Gram-positive, partially acid-fast, catalase-positive, non-motile, and none-endospore bacteria. The genus Rhodococcus was first introduced by Zopf. This bacterium can be isolated from various sources of the environment and can grow well in non-selective medium. A large number of phenotypic characterizations are used to compare different species of the genus Rhodococcus, and these tests are not suitable for accurate identification at the genus and species level. Among nucleic acid-based methods, the most powerful target gene for revealing reliable phylogenetic relationships is 16S ribosomal RNA gene (16S rRNA gene) sequence analysis, but this gene is unable to differentiation some of Rhodococcus species. To date, whole genome sequencing analysis has solved taxonomic complexities in this genus. Rhodococcus equi is the major cause of foal pneumonia, and its implication in human health is related to cases in immunocompromised patients. Macrolide family together with rifampicin is one of the most effective antibiotic agents for treatment rhodococcal infections.

Biodegradation pathway of di-(2-ethylhexyl) phthalate by a novel Rhodococcus pyridinivorans XB and its bioaugmentation for remediation of DEHP contaminated soil.

A novel bacterial strain designated as Rhodococcus pyridinivorans XB, capable of utilizing various endocrine disruptor phthalates or phthalic acid (PA) as sole source of carbon and energy, was isolated from activated sludge. Under the optimal culture conditions (pH 7.08, 30.4 °C, inoculum size (OD600 nm) of 0.6) obtained by response surface methodology, di-(2-ethylhexyl) phthalate (DEHP, 200 mg/L) could be degraded by strain XB with a removal rate of 98% within 48 h. Under the observation of an atomic force microscope, it was confirmed that DEHP did not inhibit the growth of strain XB which might produce some extracellular polymeric substances as a response to DEHP stress, resulting in rapid degradation of DEHP. At initial concentrations of 50-800 mg/L DEHP, its degradation curves were well fitted with the first-order kinetic model, and the half-life of DEHP degradation varied from 5.44 to 23.5 h. The degradation intermediates of DEHP were identified by both GC-MS and high performance liquid chromatography-time of flight-mass spectrometry (HPLC-TOF-MS). Significant up-regulation was observed for the relative expression levels of genes (i.e., phthalate hydrolase, PA 3,4-dioxygenase, protocatechuate 3,4-α and 3,4-ß dioxygenase) involved in DEHP degradation determined by real-time quantitative PCR (RT-qPCR). A DEHP biodegradation pathway by strain XB was proposed based on the identified intermediates and the degrading genes. Bioaugmentation of DEHP-contaminated soils with strain XB could efficiently promote DEHP removal, offering great potential in bioremediation of DEHP-contaminated environment.

Biodegradation of gaseous emissions of 2-chlorotoluene by strains of Rhodococcus sp. in polyurethane foam packed biotrickling filters.

About 60,000-70,000 tons of 2-chlorotoluene, which shows high toxicity in aquatic ecosystems, are produced worldwide and used in a tremendous field of applications. However, clear proofs of biodegradation were only presented for Comamonas testosteroni KT5 and Rhodococcus sp. OCT10. Hence, this study aims on the isolation of additional strains and their characterization in pilot-scale biotrickling filters. Three strains named OCT2, OCT9, and OCT14 of the genus Rhodococcus were isolated, able to mineralize gaseous 2-chlorotoluene like the previously isolated strain Rhodococcus sp. OCT10. The performance levels of these strains were tested in four biotrickling filters each containing 18.8 L of polyurethane foam package, showing elimination capacities of carbon (C) of 30.9 (OCT2), 30.1 (OCT9), 32.2 (OCT10), and 3.9 g C·m-3·h-1 (OCT14) at an average crude gas level of 397.6 mg C·m-3 and an empty bed residence time (EBRT) of 22.6 s. Since OCT10 showed the highest performance levels, this strain was characterized in a second biotrickling filter configuration at long-term conditions of 985 days, varying crude gas levels, EBRT and nutrient supply. Chloride balancing showed a recovery of 94.4% of 2-chlorotoluene eliminated out of the gas phase, pointing out mineralization of 2-chlorotoluene. German emission limit values were met at crude gas levels up to 750 mg C·m-3 at EBRTs of 120 s or higher. The maximum elimination capacity was 51.2 g C·m-3·h-1 at a specific freight of 51.9 g C·m-3·h-1 and an EBRT of 254 s. Performance levels were strongly boosted by addition of ammonia as nutrient and stabilized at efficiency levels higher than 90% at a feed rate of 4 g ammonium sulfate per week and 100 L of package volume. Repetitive monitoring of the established 2-chlorotoluene degrading community by BOX-PCR fingerprinting revealed a high long-term stability of OCT10, underlining its suitability in this kind of application.

Hydroxysteroid dehydrogenase family proteins on lipid droplets through bacteria, C. elegans, and mammals.

Lipid droplets (LDs) are the main fat storing sites in almost all species from bacteria to humans. The perilipin family has been found as LD proteins in mammals, Drosophila, and a couple of slime molds, but no bacterial LD proteins containing sequence conservation were identified. In this study, we reported that the hydroxysteroid dehydrogenase (HSD) family was found on LDs across all organisms by LD proteomic analysis. Imaging experiments confirmed LD targeting of three representative HSD proteins including ro01416 in RHA1, DHS-3 in C. elegans, and 17ß-HSD11 in human cells. In C. elegans, 17ß-HSD11 family proteins (DHS-3, DHS-4 and DHS-19) were localized on LDs in distinct tissues. In intestinal cells of C. elegans, DHS-3 targeted to cytoplasmic LDs, while DHS-9 labeled nuclear LDs. Furthermore, the N-terminal hydrophobic domains of 17ß-HSD11 family were necessary for their targeting to LDs. Last, 17ß-HSD11 family proteins induced LD aggregation, and deletion of DHS-3 in C. elegans caused lipid decrease. Independent of their presumptive catalytic sites, 17ß-HSD11 family proteins regulated LD dynamics and lipid metabolism through affecting the LD-associated ATGL, which was conserved between C. elegans and humans. Together, these findings for HSDs provide a new insight not only into the mechanistic studies of the dynamics and functions of LDs in multiple organisms, but also into understanding the evolutionary history of the organelle.

Properties of Probiotics Kocuria SM1 and Rhodococcus SM2 Isolated from Fish Guts.

This study characterized probiotics Kocuria SM1 and Rhodococcus SM2, which were recovered from the intestinal microbiota of rainbow trout (Oncorhynchus mykiss, Walbaum). The cultures were Gram-positive, non-motile, catalase-positive and oxidase-negative cocci or rods. Cell multiplication of SM1 and SM2 was observed at 4-37 °C (45 °C for SM1), in 0-20% (w/v) NaCl and at pH 2-11. The viability was not affected when exposed to pepsin at pH 2.0 and 3.0, and pancreatin at pH 8.0. Neither isolates were chrome azurol S-positive for siderophore production. Of the 19 common enzymes analysed using the API-ZYM system, only 8 were evident in the culture of SM1 compared to 11 enzymes for SM2. The secondary metabolites of both probiotics were inhibitory to Acinetobacter baumannii, Vibrio anguillarum and V. ordalii; SM2 inhibited Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Staphylococcus aureus. SM2 was resistant to penicillin and sulphatriad, out of six antimicrobial agents; SM1 was resistant to sulphatriad. These results suggest that Kocuria SM1 and Rhodococcus SM2 are able to grow over a wide range of temperature, salinity and pH, including in conditions that mimic the gastrointestinal environment of fish and produce extracellular enzymes that may have a role in the host digestive processes. Importantly, Rhodococcus SM2 displays a high degree of bacteriocinogenic potential against multi-drug-resistant human pathogens that have never been documented among the gut microbiota of fish.

Characterization of Rhodococcus sp. A5wh isolated from a high altitude Andean lake to unravel the survival strategy under lithium stress.

Lithium (Li) is widely distributed in nature and has several industrial applications. The largest reserves of Li (over 85%) are in the so-called "triangle of lithium" that includes the Salar de Atacama in Chile, Salar de Uyuni in Bolivia and Salar del Hombre Muerto in Argentina. Recently, the use of microorganisms in metal recovery such as copper has increased; however, there is little information about the recovery of lithium. The strain Rhodococcus sp. A5wh used in this work was previously isolated from Laguna Azul. The assays revealed that this strain was able to accumulate Li (39.52% of Li/g microbial cells in 180min) and that it was able to grow in its presence up to 1M. In order to understand the mechanisms implicated in Li tolerance, a proteomic approach was conducted. Comparative proteomic analyses of strain A5wh exposed and unexposed to Li reveal that 17 spots were differentially expressed. The identification of proteins was performed by MALDI-TOF/MS, and the obtained results showed that proteins involved in stress response, transcription, translations, and metabolism were expressed under Li stress. This knowledge constitutes the first proteomic approach to elucidate the strategy followed by Rhodococcus to adapt to Li.

Neovascularization during leafy gall formation on Arabidopsis thaliana upon Rhodococcus fascians infection.

MAIN CONCLUSION: Extensive de novo vascularization of leafy galls emerging upon Rhodococcus fascians infection is achieved by fascicular/interfascicular cambium activity and transdifferentiation of parenchyma cells correlated with increased auxin signaling. A leafy gall consisting of fully developed yet growth-inhibited shoots, induced by the actinomycete Rhodococcus fascians, differs in structure compared to the callus-like galls induced by other bacteria. To get insight into the vascular development accompanying the emergence of the leafy gall, the anatomy of infected axillary regions of the inflorescence stem of wild-type Arabidopsis thaliana accession Col-0 plants and the auxin response in pDR5:GUS-tagged plants were followed in time. Based on our observations, three phases can be discerned during vascularization of the symptomatic tissue. First, existing fascicular cambium becomes activated and interfascicular cambium is formed giving rise to secondary vascular elements in a basipetal direction below the infection site in the main stem and in an acropetal direction in the entire side branch. Then, parenchyma cells in the region between both stems transdifferentiate acropetally towards the surface of the developing symptomatic tissue leading to the formation of xylem and vascularize the hyperplasia as they expand. Finally, parenchyma cells in the developing gall also transdifferentiate to vascular elements without any specific direction resulting in excessive vasculature disorderly distributed in the leafy gall. Prior to any apparent anatomical changes, a strong auxin response is mounted, implying that auxin is the signal that controls the vascular differentiation induced by the infection. To conclude, we propose the "sidetracking gall hypothesis" as we discuss the mechanisms driving the formation of superfluous vasculature of the emerging leafy gall.