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Biofilm formation by ΔEmbRS cells. Formation of bacterial veils around the toothpick scaffolds. Cells were inoculated in the liquid medium and grown photosynthetically in plates. The extracellular matrix polymerization and its anchorage to the toothpicks led to the formation of the polygonal networks. (See movie M2).
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Photosynthetic bacteria can switch from planktonic lifestyle to phototrophic biofilm in mats in response to environmental changes. The mechanisms of phototrophic biofilm formation are, however, not characterized. Herein, we report a two-component system EmbRS that controls the biofilm formation in a photosynthetic member of the Burkholderiales orde...
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... this end, DEmbRS cell aggregates were scattered within the medium in the plates containing toothpick scaffolds and allowed to grow by photosynthesis overnight. As shown in Figure 3, growth resulted in the formation of con- spicuous bacterial veils around the toothpick scaffolds. Cells and the matrix were associated with the toothpicks in large well-arranged polymeric structures. ...
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... prompted us to question about the rearrangement of the biofilm and to continuously follow its formation to capture the details of this rearrangement. As shown in movie M2 and in Figure S3, the cells/matrix grew until the matrix was associated with the plate borders; then rearrange- ment of the biofilm started when the matrix spontane- ously relaxed. The shrinking of the biofilm continued until the veil was in contact with the top of the tooth- picks, generating the organized polygonal network. ...
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... of acquisition of the same culture. Movie 2. Biofilm formation by the DEmbRS mutant. For- mation of the bacterial veil around the toothpick scaffold. Cells start growing in the whole plate; then the shrinking of the biofilm occurs and stops when the veil comes in contact with the top of the toothpicks, generating an organized polygonal network (Fig. 3). . Putative regulatory model of the biofilm for- mation and autoaggregation in Rubrivivax gelatinosus based on the genetic and molecular data in this study. EmbRS functions as a negative two-component system that control genes (cpsG, wcbM…) involved in the synthe- sis of exopolysaccharides (extracellular matrix). BmfSR is a second ...
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The dynamics of bacterial communities in a full-scale wastewater treatment plant (WWTP) were investigated using Illumina MiSeq sequencing and PICRUSt (phylogenetic investigation of communities by reconstruction of unobserved states) over a one-year period. Results showed that the temporal dynamics of structure, diversity and abundance of bacterial...
Citations
... Two extreme cases were sampled in our experiments of Hg(II) binding affinity of intact cells of Rvx. gelatinosus (Steunou et al. 2013). In the planktonic mode of living, the diffusion step is much faster than that through the membrane (k d >> k a ). ...
Mercury adsorption on the cell surface and intracellular uptake by bacteria represent the key first step in the production and accumulation of highly toxic mercury in living organisms. In this work, the biophysical characteristics of mercury bioaccumulation are studied in intact cells of photosynthetic bacteria by use of analytical (dithizone) assay and physiological photosynthetic markers (pigment content, fluorescence induction, and membrane potential) to determine the amount of mercury ions bound to the cell surface and taken up by the cell. It is shown that the Hg(II) uptake mechanism (1) has two kinetically distinguishable components, (2) includes co-opted influx through heavy metal transporters since the slow component is inhibited by Ca²⁺ channel blockers, (3) shows complex pH dependence demonstrating the competition of ligand binding of Hg(II) ions with H⁺ ions (low pH) and high tendency of complex formation of Hg(II) with hydroxyl ions (high pH), and (4) is not a passive but an energy-dependent process as evidenced by light activation and inhibition by protonophore. Photosynthetic bacteria can accumulate Hg(II) in amounts much (about 10⁵) greater than their own masses by well-defined strong and weak binding sites with equilibrium binding constants in the range of 1 (μM)⁻¹ and 1 (mM)⁻¹, respectively. The strong binding sites are attributed to sulfhydryl groups as the uptake is blocked by use of sulfhydryl modifying agents and their number is much (two orders of magnitude) smaller than the number of weak binding sites. Biofilms developed by some bacteria (e.g., Rvx. gelatinosus) increase the mercury binding capacity further by a factor of about five. Photosynthetic bacteria in the light act as a sponge of Hg(II) and can be potentially used for biomonitoring and bioremediation of mercury-contaminated aqueous cultures.
... Enfin, l'expression de BdcA augmente la motilité cellulaire et la production d'ADNe, alors qu'elle diminue l'agrégation cellulaire, la longueur des cellules et la production d'exopolysaccharides. La protéine BdcA contrôle ainsi la dispersion des biofilms d'E. coli en se liant au c-di-GMP et en diminuant la concentration intracellulaire du messager (Ma et al, 2011a). Bien que se liant au c-di-GMP, BdcA n'agit toutefois pas comme une PDE (Ma et al, 2011a) et bien que présentant une homologie avec les DGC de nombreuses bactéries, son activité synthétique n'est à ce jour pas encore démontrée (Steunou et al, 2013). BdcA a récemment été identifiée comme protéine appartenant à la superfamille des déshydrogénases/réductases NAD(P)(H) dépendantes (Lord et al, 2014). ...
Les biofilms sont des communautés de microorganismes adhérant à une surface et encastrées dans une substance polymérique produite par les cellules du système, dite matrice extracellulaire. Du fait de leur nature ubiquitaire, les biofilms colonisent de nombreux environnements et causent souvent de sérieux problèmes dans les secteurs de la santé et de l’industrie. La dispersion par ajout d’agent chimique est l’une des stratégies de lutte contre les biofilms. Un acide gras, l’acide cis-2-décénoique (CDA), semble être prometteur pour ce faire, grâce à l’étendue de son action dispersante sur les espèces et règnes du vivant. L’objectif de ce travail de thèse est d’investiguer les mécanismes de dispersion des biofilms de l’espèce bactérienne Escherichia coli (E. coli) par la molécule modèle CDA. Le CDA modifie-t-il les structures du biofilm ou induit-il une réponse génétique des bactéries lors de la dispersion ? Pour répondre à ces questions, la dispersion des biofilms d’E. coli a été étudiée in situ dans des chambres microfluidiques par microscopie confocale à balayage laser (CLSM). Des souches bactériennes spécifiques ont été construites par clonage de promoteurs d’intérêt en fusion transcriptionnelle avec un gène codant pour une protéine fluorescente verte. Les résultats confirment l’activité dispersante du CDA avec une réduction significative de la biomasse, de l’épaisseur moyenne et de l’aire de recouvrement par couche du biofilm. Un outil innovant d’analyse d’images CLSM a été développé en collaboration dans le but de déterminer les propriétés structurales du biofilm et l’intensité de fluorescence in situ du rapporteur étudié. Les résultats indiquent une augmentation de l’intensité moyenne de fluorescence des biofilms après dispersion avec le CDA, au niveau global en considérant tout le biofilm et au niveau local en considérant une segmentation du biofilm en microcolonies, ainsi qu’en profondeur. Ces résultats évoquent un changement d’expression génique des bactéries en présence de CDA. Par ailleurs, les résultats montrent que le CDA ne semble pas avoir d’effet en culture planctonique, ni sur la croissance bactérienne ni sur l’activité des promoteurs sélectionnés. Ceci suggère que les effets du CDA sont biofilm-dépendants.
Pollution by copper (Cu²⁺) extensively used as antimicrobial in agriculture and farming represents a threat to the environment and human health. Finding ways to make microorganisms sensitive to lower metal concentrations could help decreasing the use of Cu²⁺ in agriculture. In this respect, we showed that limiting iron (Fe) uptake makes bacteria much more susceptible to Cu²⁺ or Cd²⁺ poisoning. Using efflux mutants of the purple bacterium Rubrivivax gelatinosus, we showed that Cu⁺ and Cd²⁺ resistance relies on the expression of the Fur‐regulated FbpABC and Ftr iron transporters. To support this conclusion, inactivation of these Fe‐importers in the Cu⁺ or Cd²⁺‐ATPase efflux mutants gave rise to hypersensitivity towards these ions. Moreover, in metal overloaded cells the expression of FbpA, the periplasmic iron‐binding component of the ferric ion transport FbpABC system was induced, suggesting that cells perceived an ‘iron‐starvation’ situation and responded to it by inducing Fe‐importers. In this context, the Fe‐Sod activity increased in response to Fe homoeostasis dysregulation. Similar results were obtained for Vibrio cholerae and Escherichia coli, suggesting that perturbation of Fe‐homoeostasis by metal excess appeared as an adaptive response commonly used by a variety of bacteria. The presented data support a model in which metal excess induces Fe‐uptake to support [4Fe‐4S] synthesis and thereby induce ROS detoxification system.
In response to environmental changes, the photosynthetic bacterium Rubrivivax gelatinosus (Rvx.) can switch from a planktonic lifestyle to a phototrophic biofilm. Like in critical phenomena, the colonization and sedimentation of the cells is abrupt and hard to predict causally, and the underlying biophysics of the mechanisms involved is not known. Herein, we report basic experimental observations and quantitative explanations as keys to understanding microbial turnover of aggregates. (1) The moment of sedimentation can be controlled by the height of the tube of cultivation, by the concentrations of externally added Ficoll (a highly branched polymer) and/or of internally produced polysaccharides (constituents of the biofilm). (2) The observed translational diffusion coefficient of the planktonic bacteria is the sum of diffusion coefficients coming from random Brownian and twitching movements of the bacteria and amounts to 14 (μm)²/s. (3) This value drops hyperbolically with the association number of the cell aggregates and with the concentration of the exopolysaccharides in the biofilm. In the experiments described herein, their effects could be separated. (4) The critical conditions of colonization and sinking of the cells will be achieved if the height of the tube meets the scale height that is proportional to the ratio of the diffusion coefficient and the net mass of the bacterium. The decisive role of the web-like structure of a biofilm, the organization of bacteria from loose cooperativity to solid aggregation, and the possible importance of similar controls in other phototrophic microorganisms are discussed.
Thauera aminoaromatica
MZ1T, a floc-forming bacterium isolated from an industrial activated sludge wastewater treatment plant, overproduces exopolysaccharide (EPS) leading to viscous bulking. This phenomenon results in poor sludge settling and dewatering during the clarification process. To identify genes responsible for bacterial flocculation, a whole genome phenotypic sequencing technique was applied. Genomic DNA of MZ1T flocculation-deficient mutants were subjected to massively parallel sequencing. The resultant high-quality reads were assembled and compared to the reference genome of the wild type. We identified nine nonsynonymous mutations and one nonsense mutation putatively involved in EPS biosynthesis. Complementation of the nonsense mutation located in an EPS deacetylase gene restored the flocculating phenotype. The FTIR spectra of EPS isolated from the wild-type showed reduced C=O peak of the
N
-acetyl group at 1665 cm
-1
as compared to the spectra of MZ1T floc-deficient mutant EPS, suggesting that the WT EPS was partially deacetylated. Gene expression analysis also demonstrated the putative deacetylase gene transcript increased before flocculation occurred. These data suggest that targeting deacetylation processes via direct chemical modification of EPS or enzyme inhibition may prove useful in combating viscous bulking in this and related bacteria.
In this study, potential application of phototaxis to promote the formation of anoxygenic photosynthetic bacteria (APB) biofilm and the hydrogen production was evaluated in batch and continuous experiments. The batch experiments showed that the biofilm biomass on the illuminated sidewall reached 0.86 mg cm2, which was 26-fold higher than that of the biofilm on the unlighted sidewall. The analyses of living cell fluorescence labeling and intracellular c-d-GMP suggested that the sidewall illumination aggregated APB cells to form flocculations on illuminated sidewall by upregulating intracellular c-di-GMP levels, but not on illuminated liquid surface. These findings suggested that phototaxis and surface sensing modulated the biofilm formation of APB. Further, a continuous photorotating biological contactor reactor with mirror coating was developed to improve APB biofilm formation. The APB biomass and hydrogen production were significantly higher than those in the control without mirror coating, indicating the feasibility of applying reflected light-induced biofilm formation of APB for the improvement of hydrogen production.
Literature published during the year 2013 regarding the use of biofilms and bioreactors to treat water and wastewater is revie\ ed herein. The review is organized into the following sections: biofilm fornl. ationand factors Ulat impact biofilm fom13tion; xtracellular polymeric substance and its e rtraction from biofihns; biofilm consortia and quorum sensing; biofilm imaging teclmiques; biofilm carriers and biofilm reactors.
Background
Most photobioreactors used for wastewater treatment with anoxygenic photosynthetic bacteria (APB) are based on suspended cultures because APB have poor biofilm-forming ability. In this study, a photo-rotating biological contactor (PRBC) was applied for the culture of APB biofilm using azo wastewater, with a particular emphasis on the formation of APB biofilm and its relationship to other microorganisms and exopolysaccharides (EPS). ResultsPRBC was successfully used in the cultivation of APB biofilm and in the removal of chemical oxygen demand and color. In-situ analysis showed that APB biofilm formation can be divided into two distinct stages: development and differentiation. In the development stage, nonphotosynthetic filamentous microorganisms initially proliferated on a disc surface and promoted the adherence of APB. In the differentiation stage, filamentous microorganisms and their EPS matrices caged APB in the inner layer and stabilized the APB biofilm. During biofilm stratification, the biofilm biomass and APB number reached 24.2 ± 0.63 g L−1 and 4.8 ± 0.6 × 108 CFU (g dry biofilm)−1, respectively. The dominant APB in the mature biofilm were identified as Rhodopseudomonas, Rhodomicrobium, and Chlorobium. ConclusionAPB with poor biofilm-forming ability could form a steady biofilm in PRBC with the aid of nonphotosynthetic microorganisms.
