Role of Respiratory Nitrate Reductase in Ability of Pseudomonas fluorescens YT101 To Colonize the Rhizosphere of Maize

Selection of the denitrifying community by plant roots (i.e., increase in the denitrifier/total heterotroph ratio in the rhizosphere) has been reported by several authors. However, very few studies to evaluate the role of the denitrifying function itself in the selection of microorganisms in the rhizosphere have been performed. In the present study, we compared the rhizosphere survival of the denitrifying Pseudomonas fluorescens YT101 strain with that of its isogenic mutant deficient in the ability to synthesize the respiratory nitrate reductase, coinoculated in nonplanted or planted soil. We demonstrated that under nonlimiting nitrate conditions, the denitrifying wild-type strain had an advantage in the ability to colonize the rhizosphere of maize. Investigations of the effect of the inoculum characteristics (density of the total inoculum and relative proportions of mutant and wild-type strains) on the outcome of the selection demonstrated that the selective effect of the plant was expressed only during the phase of bacterial multiplication and that the intensity of selection was dependent on the magnitude of this phase. Moreover, application of the de Wit replacement series technique to our results suggests that the advantage of the wild-type strain was maximal when the ratio between the two strains in the inoculum was close to 1:1. This work constitutes the first direct demonstration that the presence of a functional structural gene encoding the respiratory nitrate reductase confers higher rhizosphere competence to a microorganism.     

Preparation and characterization of F-protein vesicles isolated from Sendai virus by means of octyl glucoside

We have demonstrated that Triton X-100 is always present in F-protein vesicles at concentrations that can provoke cell lysis. In order to avoid any misinterpretation of the fusogenic capacity of this protein, we solubilized the Sendai virus using octyl glucoside, which can be totally removed from the F protein preparation in less than 16 h by dialysis in the presence of absorbent beads. F-glycoprotein preparations preserved their ability to lyse erythrocytes in the presence of lectins and to induce cell-vesicle fusion as demonstrated by ESR studies. These vesicles were characterized by electron microscopy and SDS-polyacrylamide gel electrophoresis. Lipid analysis of these preparations by thin-layer chromatography indicated that they had the same proportion of lipids as virus envelopes, with slight variations in the sphingomyelin content and the cholesterol/phospholipid molar ratio. F-protein vesicles of different sizes can be obtained by adding exogenous lipids before detergent removal. The hemolytic activity of the vesicles was retained over a large range of lipid concentrations. We conclude that F-protein vesicles prepared with octyl glucoside are convenient tools for studying the fusogenic mechanism of this protein and improving the fusion process between liposomes and cells.     

Abundance and activity of nitrate reducers in an arable soil are more affected by temporal variation and soil depth than by elevated atmospheric [CO2

Elevated atmospheric carbon dioxide concentrations ([CO(2) ]) might change the abundance and the function of soil microorganisms in the depth profile of agricultural soils by plant-mediated reactions. The seasonal pattern of abundance and activity of nitrate-reducing bacteria was studied in a Mini-FACE experiment planted with oilseed rape (Brassica napus). Three depths (0-10, 10-20 and 20-30 cm) were sampled. Analyses of the abundances of total (16S rRNA gene) and nitrate-reducing bacteria (narG, napA) revealed strong influences of sampling date and depth, but no [CO(2)] effects. Abundance and activity of nitrate reducers were higher in the top soil layer and decreased with depth but were not related to extractable amounts of nitrogen and carbon in soil. Dry periods reduced abundances of total and nitrate-reducing bacteria, whereas the potential activity of the nitrate reductase enzyme was not affected. Enzyme activity was only weakly correlated to the abundance of nitrate-reducing bacteria but was related to NH(4) (+) and NO(3) (-) concentrations. Our results suggest that in contrast to the observed pronounced seasonal changes, the elevation of atmospheric [CO(2) ] has only a marginal impact on nitrate reducers in the investigated arable ecosystem.     

Maladaptive Social Self-Beliefs in Alcohol-Dependence: A Specific Bias towards Excessive High Standards

Background

Emotional and interpersonal impairments associated with alcohol-dependence have been recently explored, but the distorted cognitive representations underlying these deficits remain poorly understood. The present study aims at exploring the presence of maladaptive social self-beliefs among alcohol-dependent individuals, as these biased self-beliefs have been recently shown to play a crucial role in the development and maintenance of other psychopathological states (social anxiety and depression).

Methodology/Principal findings

Twenty-five recently detoxified alcohol-dependent participants and 25 matched controls filled in self-report questionnaires evaluating maladaptive social self-beliefs, interpersonal problems and several comorbid states (anxiety, social anxiety, depression). As compared to controls, alcohol-dependent individuals showed higher scores than controls for the three subcategories of maladaptive social self-beliefs (high standards, conditional beliefs and unconditional beliefs). Our key finding was that when comorbidities were controlled for, alcohol-dependence was associated with a specific bias towards exaggerated high standards in social contexts. Moreover, these high standards beliefs were strongly correlated with interpersonal problems.

Conclusions/Significance

These results provide the first insights into the influence of cognitive biases on interpersonal problems in addictive states, and suggest that maladaptive self-beliefs could have a central influence on the development and maintenance of alcohol-dependence.     

Impact of Anodal and Cathodal Transcranial Direct Current Stimulation over the Left Dorsolateral Prefrontal Cortex during Attention Bias Modification: An Eye-Tracking Study

People with anxiety disorders show an attentional bias for threat (AB), and Attention Bias Modification (ABM) procedures have been found to reduce this bias. However, the underlying processes accounting for this effect remain poorly understood. One explanation suggests that ABM requires the modification of attention control, driven by the recruitment of the dorsolateral prefrontal cortex (DLPFC). In the present double-blind study, we examined whether modifying left DLPFC activation influences the effect of ABM on AB. We used transcranial direct current stimulation (tDCS) to directly modulate cortical excitability of the left DLPFC during an ABM procedure designed to reduce AB to threat. Anodal tDCS increases excitability, whereas cathodal tDCS decreases it. We randomly assigned highly trait-anxious individuals to one of three conditions: 1) ABM combined with cathodal tDCS, 2) ABM combined with anodal tDCS, or 3) ABM combined with sham tDCS. We assessed the effects of these manipulations on both reaction times and eye-movements on a task indexing AB. Results indicate that combining ABM and anodal tDCS over the left DLPFC reduces the total duration that participants’ gaze remains fixated on threat, as assessed using eye-tracking measurement. However, in contrast to previous studies, there were no changes in AB from baseline to post-training for participants that received ABM without tDCS. As the tendency to maintain attention to threat is known to play an important role in the maintenance of anxiety, the present findings suggest that anodal tDCS over the left DLPFC may be considered as a promising tool to reduce the maintenance of gaze to threat. Implications for future translational research combining ABM and tDCS are discussed.     

Response of total and nitrate-dissimilating bacteria to reduced N deposition in a spruce forest soil profile

A field-scale manipulation experiment conducted for 16 years in a Norway spruce forest at Solling, Central Germany, was used to follow the long-term response of total soil bacteria, nitrate reducers and denitrifiers under conditions of reduced N deposition. N was experimentally removed from throughfall by a roof construction ('clean rain plot'). We used substrate-induced respiration (SIR) to characterize the active fraction of soil microbial biomass and potential nitrate reduction to quantify the activity of nitrate reducers. The abundance of total bacteria, nitrate reducers and denitrifiers in different soil layers was analysed by quantitative PCR of 16S rRNA gene, nitrate reduction and denitrification genes. Reduced N deposition temporarily affected the active fraction of the total microbial community (SIR) as well as nitrate reductase activity. However, the size of the total, nitrate reducer and denitrifier communities did not respond to reduced N deposition. Soil depth and sampling date had a greater influence on the density and activity of soil microorganisms than reduced deposition. An increase in the nosZ /16S rRNA gene and nosZ/nirK ratios with soil depth suggests that the proportion of denitrifiers capable of reducing N₂O into N₂ is larger in the mineral soil layer than in the organic layer.     

Involvement of Nitrate Reductase and Pyoverdine in Competitiveness of Pseudomonas fluorescens Strain C7R12 in Soil

Involvement of nitrate reductase and pyoverdine in the competitiveness of the biocontrol strain Pseudomonas fluorescens C7R12 was determined, under gnotobiotic conditions, in two soil compartments (bulk and rhizosphere soil), with the soil being kept at two different values of matric potential (−1 and −10 kPa). Three mutants affected in the synthesis of either the nitrate reductase (Nar⁻), the pyoverdine (Pvd⁻), or both (Nar⁻ Pvd⁻) were used. The Nar⁻ and Nar⁻ Pvd⁻ mutants were obtained by site-directed mutagenesis of the wild-type strain and of the Pvd⁻ mutant, respectively. The selective advantage given by nitrate reductase and pyoverdine to the wild-type strain was assessed by measuring the dynamic of each mutant-to-total-inoculant (wild-type strain plus mutant) ratio. All three mutants showed a lower competitiveness than the wild-type strain, indicating that both nitrate reductase and pyoverdine are involved in the fitness of P. fluorescens C7R12. The double mutant presented the lowest competitiveness. Overall, the competitive advantages given to C7R12 by nitrate reductase and pyoverdine were similar. However, the selective advantage given by nitrate reductase was more strongly expressed under conditions of lower aeration (−1 kPa). In contrast, the selective advantage given by nitrate reductase and pyoverdine did not differ in bulk and rhizosphere soil, indicating that these bacterial traits are not specifically involved in the rhizosphere competence but rather in the saprophytic ability of C7R12 in soil environments.     

Effect of primary mild stresses on resilience and resistance of the nitrate reducer community to a subsequent severe stress

The factors regulating soil microbial stability (e.g. resistance and resilience) are poorly understood, even though microorganisms are essential for ecosystem functioning. In this study, we tested whether a functional microbial community subjected to different primary mild stresses was equally resistant or resilient to a subsequent severe stress. The nitrate reducers were selected as model community and analysed in terms of nitrate reduction rates and genetic structure by narG PCR-restriction fragment length polymorphism fingerprinting. Heat, copper and atrazine were used as primary stresses and mercury at a high concentration as a severe stress. None of the primary stresses had any significant impact on the nitrate reducer community. Although primary stress with heat, copper or atrazine had no effect on the resilience of the nitrate reducer activity to mercury stress, pre-exposure to copper, another heavy metal, resulted in increased resilience. In contrast, the resistance of both structure and activity of the nitrate reducer community to severe mercury stress was not affected by any of the primary stresses tested. Our experiment suggests that the hypothetical effect of an initial stress on the response of a microbial community to an additional stress is complex and may depend on the relatedness of the two consecutive stresses and the development of positive cotolerance.     

Role of respiratory nitrate reductase in ability of Pseudomonas fluorescens YT101 to colonize the rhizosphere of maize

Selection of the denitrifying community by plant roots (i.e., increase in the denitrifier/total heterotroph ratio in the rhizosphere) has been reported by several authors. However, very few studies to evaluate the role of the denitrifying function itself in the selection of microorganisms in the rhizosphere have been performed. In the present study, we compared the rhizosphere survival of the denitrifying Pseudomonas fluorescens YT101 strain with that of its isogenic mutant deficient in the ability to synthesize the respiratory nitrate reductase, coinoculated in nonplanted or planted soil. We demonstrated that under nonlimiting nitrate conditions, the denitrifying wild-type strain had an advantage in the ability to colonize the rhizosphere of maize. Investigations of the effect of the inoculum characteristics (density of the total inoculum and relative proportions of mutant and wild-type strains) on the outcome of the selection demonstrated that the selective effect of the plant was expressed only during the phase of bacterial multiplication and that the intensity of selection was dependent on the magnitude of this phase. Moreover, application of the de Wit replacement series technique to our results suggests that the advantage of the wild-type strain was maximal when the ratio between the two strains in the inoculum was close to 1:1. This work constitutes the first direct demonstration that the presence of a functional structural gene encoding the respiratory nitrate reductase confers higher rhizosphere competence to a microorganism.