The Prc and CtpA proteases modulate cell-surface signaling activity and virulence in Pseudomonas aeruginosa.

Cell-surface signaling (CSS) is a signal transfer system of Gram-negative bacteria that produces the activation of an extracytoplasmic function σ factor (σECF) in the cytosol in response to an extracellular signal. Activation requires the regulated and sequential proteolysis of the σECF-associated anti-σ factor, and the function of the Prc and RseP proteases. In this work, we have identified another protease that modulates CSS activity, namely the periplasmic carboxyl-terminal processing protease CtpA. CtpA functions upstream of Prc in the proteolytic cascade and seems to prevent the Prc-mediated proteolysis of the CSS anti-σ factor. Importantly, using zebrafish embryos and the A549 lung epithelial cell line as hosts, we show that mutants in the rseP and ctpA proteases of the human pathogen Pseudomonas aeruginosa are considerably attenuated in virulence while the prc mutation increases virulence likely by enhancing the production of membrane vesicles.

Clover Root Exudates Favor Novosphingobium sp. HR1a Establishment in the Rhizosphere and Promote Phenanthrene Rhizoremediation.

Rhizoremediation is based on the ability of microorganisms to metabolize nutrients from plant root exudates and, thereby, to cometabolize or even mineralize toxic environmental contaminants. Novosphingobium sp. HR1a is a bacterial strain able to degrade a wide variety of polycyclic aromatic hydrocarbons (PAHs). Here, we have demonstrated that the number of CFU in microcosms vegetated with clover was almost 2 orders of magnitude higher than that in nonvegetated microcosms or microcosms vegetated with rye-grass or grass. Strain HR1a was able to eliminate 92% of the phenanthrene in the microcosms with clover after 9 days. We have studied the molecular basis of the interaction between strain HR1a and clover by phenomic, metabolomic, and transcriptomic analyses. By measuring the relative concentrations of several metabolites exudated by clover both in the presence and in the absence of the bacteria, we identified some compounds that were probably consumed in the rhizosphere; the transcriptomic analyses confirmed the expression of genes involved in the catabolism of these compounds. By using a transcriptional fusion of the green fluorescent protein (GFP) to the promoter of the gene encoding the dioxygenase involved in the degradation of PAHs, we have demonstrated that this gene is induced at higher levels in clover microcosms than in nonvegetated microcosms. Therefore, the positive interaction between clover and Novosphingobium sp. HR1a during rhizoremediation is a result of the bacterial utilization of different carbon and nitrogen sources released during seedling development and the capacity of clover exudates to induce the PAH degradation pathway. IMPORTANCE The success of an eco-friendly and cost-effective strategy for soil decontamination is conditioned by the understanding of the ecology of plant-microorganism interactions. Although many studies have been published about the bacterial metabolic capacities in the rhizosphere and about rhizoremediation of contaminants, there are fewer studies dealing with the integration of bacterial metabolic capacities in the rhizosphere during PAH bioremediation, and some aspects still remain controversial. Some authors have postulated that the presence of easily metabolizable carbon sources in root exudates might repress the expression of genes required for contaminant degradation, while others found that specific rhizosphere compounds can induce such genes. Novosphingobium sp. HR1a, which is our model organism, has two characteristics desirable in bacteria for use in remediation: its ubiquity and the capacity to degrade a wide variety of contaminants. We have demonstrated that this bacterium consumes several rhizospheric compounds without repression of the genes required for the mineralization of PAHs. In fact, some compounds even induced their expression.

Degradation of phenanthrene by Novosphingobium sp. HS2a improved plant growth in PAHs-contaminated environments.

At the same time that the European Union (EU) policy recommend to direct efforts towards reductions of heavy metals, polycyclic aromatic hydrocarbons (PAHs) and mining residues, there is the need to increase the cultivable areas within Europe to cope with the increasing demands for food and energy crops. Bioremediation is a good technique for the restoration of contaminated soils; however, it has not been used extensively because of the variability of the outcome. This variability is frequently due to a bad establishment of foreign degrading populations in soil. We have demonstrated that Novosphingobium sp. HS2aR (i) is able to compete with other root colonizers and with indigenous bacteria, (ii) is able to establish in high numbers in the contaminated environments and (iii) is able to remove more than 90 % of the extractable phenanthrene in artificially contaminated soils. Furthermore, we have demonstrated that the capacity to remove phenanthrene is linked to the ability to promote plant growth in contaminated environments. The fact that the presence of Novosphingobium sp. HS2aR improves the growth of plants in contaminated soil suggests that it may be a useful strain for utilization in amelioration of soil quality while improving the growth of economically important energy crops, thus adding value to the bioremediation strategy.

The pGRT1 plasmid of Pseudomonas putida DOT-T1E encodes functions relevant for survival under harsh conditions in the environment.

Pseudomonas putida DOT-T1E has the capacity to grow in the presence of high concentrations of toluene. This ability is mainly conferred by an efflux pump encoded in a self-transmissible 133 kb plasmid named pGRT1. Sequence analysis of the pGRT1 plasmid revealed several key features. Most of the genes related to the plasmid maintenance functions show similarity with those encoded on pBVIE04 from Burkholderia vietnamensis G4, and knock-out mutants in several of these genes confirmed their roles. Two additional plasmid DNA fragments were incorporated into the plasmid backbone by recombination and/or transposition; in these DNA regions, apart from multiple recombinases and transposases, several stress-related and environmentally relevant functions are encoded. We report that plasmid pGRT1 not only confers the cells with tolerance to toluene but also resistance to ultraviolet light. We show here the implication of a new protein in solvent tolerance which controls the level of expression of the TtgGHI efflux pump, as well as the implication of a protein with homology to the universal stress protein in solvent tolerance and ultraviolet light resistance. Furthermore, this plasmid encodes functions that allow the cells to chemotactically respond to toluene and participate in iron scavenging.