Biodegradation in a Partially Saturated Sand Matrix: Compounding Effects of Water Content, Bacterial Spatial Distribution, and Motility

Bacterial pesticide degraders are generally heterogeneously distributed in soils, leaving soil volumes devoid of degradation potential. This is expected to have an impact on degradation rates because the degradation of pollutant molecules in such zones will be contingent either on degraders colonizing these zones or on pollutant mass transfer to neighboring zones containing degraders. In a model system, we quantified the role exerted by water on mineralization rate in the context of a heterogeneously distributed degradation potential. Alginate beads colonized by Pseudomonas putida KT2440 were inserted at prescribed locations in sand microcosms so that the initial spatial distribution of the mineralization potential was controlled. The mineralization rate was strongly affected by the matric potential (decreasing rate with decreasing matric potential) and by the initial distribution of the degraders (more aggregated distributions being associated with lower rates). The mineralization was diffusion-limited, as confirmed with a mathematical model. In wet conditions, extensive cell dispersal was observed for the flagellated wild type and, albeit to a lesser extent, for a nonflagellated mutant, partially relieving the diffusion limitation. Dry conditions, however, sustained low mineralization rates through the combined effects of low pollutant diffusivity and limited degrader dispersal

Outer membrane Modifications of <i>Pseudomonas fluorescens</i> MF37 in Response to Hyperosmolarity

The effect of hyperosmotic condition on the outer membrane protein (Omp) composition of Pseudomonas fluorescens was investigated by proteomic analyses. The abundances of 12 proteins, including porins, lipoproteins, and the flagella subunit FliC, were modified. This was at least partly explained by altered gene expression, as shown by mRNA level study. In agreement with Omp changes, hyperosmotic condition resulted in vesicle formation and modifications of mobility and antibiotic susceptibility

Functional classes of SigX-regulated genes identified by expression profiling on DNA array.

<p>All 307 genes that had a significant difference in expression between wildtype and mutant strains (Fold change ≥2,<i>p</i>-value ≤0.05 as determined by Empirical Bayes) were included and classified according to their function. Functional classes were determined using the <i>Pseudomonas</i> Genome Project website (<a href="http://www.pseudomonas.com" target="_blank">www.pseudomonas.com</a>; Winsor et al., 2011), among which these framed in grey were discussed.</p

Altered levels of (A) exotoxinA, (B) pyocyanin and (C) siderophores in the culture supernatants of the <i>sigX</i> mutant.

<p>H103 (black), PAOSX (white) and PAOSX+ (grey) culture supernatants were obtained from overnight cultures in (A) LB, (B) King A or (C) King B media. The relative amounts of exotoxin A, pyocyanin and the total siderophores were assayed at least three times independently for each strain and means and standard deviations are presented. For the Chrome Azurol S (CAS) assay, the haloes around the wells in the CAS plate show siderophore production in sample supernatant. Statistics were done by pairwise strain comparisons (<i>t</i> test). *<i>p</i>-value<0.05, **<i>p</i>-value<0.01, ***<i>p</i>-value<0.001, **** <i>p</i>-value<0.0001, NS no significant difference.</p

Involvement of SigX in (A) twitching and (B) swarming motilities, attachment to (C) glass slides and to (D) Caco2/TC7 cells, and (E) biofilm formation.

<p>Twitching motility was assayed on solidified M9G medium containing 1% agar. Swarming motility was assayed on M9G containing 1% casamino acids as nitrogen source and solidified with 0.5 % agar. For adherence onto glass slides, mid-log phase cultures of GFP expressing bacteria were diluted in 0.9% NaCl to an OD₅₈₀ of 0.6 and allowed to adhere for 2h. Attached cells were observed using a confocal laser scanning microscope and a binding index was calculated (value on each slide). Binding of bacteria onto Caco2/TC7 cells: each bar represents the mean number of adherent bacteria per cell (±SD) calculated by direct microscopic counting of 100 cells and expressed as a percentage compared to the binding of the wildtype H103 strain. For biofilm assay, bacteria were allowed to form a pellicle for 24h at 37°C. Biofilms were quantified by measuring the absorbance at 595 nm after crystal violet (CV) staining. Relative biofilm formation was determined by comparison to the wildtype strain (±SD). Each experiment was performed at least three times. Statistics were done by pairwise strain comparisons (<i>t</i> test). *<i>p</i>-value<0.05, **<i>p</i>-value<0.01, ***<i>p</i>-value<0.001.</p

The absence of SigX modulated <i>P</i>. <i>aeruginosa</i> virulence in the <i>C</i>. <i>elegans</i> model.

<p>Kaplan-Meier survival plots of <i>C</i>. <i>elegans</i> nematodes fed with the wild type strain <i>P</i>. <i>aeruginosa</i> H103 (n = 210), the <i>sigX</i> mutant PAOSX (n = 257), and the SigX-complemented mutant strain PAOSX+ (n = 201). Each value reported for the assay is the mean of measurements of eight samples from three independent experiments. Pairwise strain comparisons (log rank test) were as follows: H103 <i>versus</i> PAOSX, <i>p</i>-value< 0.0001; PAOSX <i>versus</i> PAOSX+, <i>p</i>-value< 0.0001; H103 <i>versus</i> PAOSX+, <i>p</i>-value <0.001. Four independent experiments were performed. </p