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Paula I. Watnick - One of the best experts on this subject based on the ideXlab platform.
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identification of novel stage specific genetic requirements through whole genome transcription profiling of vibrio cholerae Biofilm Development
Molecular Microbiology, 2005Co-Authors: Sudha Moorthy, Paula I. WatnickAbstract:Bacterial Biofilm formation has been described as a Developmental process. This process may be divided into three stages: the planktonic stage, the monolayer stage and the Biofilm stage. Bacteria in the planktonic stage are not attached to each other or to a surface; bacteria in the monolayer stage are attached to surfaces as single cells; and bacteria in the Biofilm stage are attached to surfaces as cellular aggregates. In a study limited to the Vibrio cholerae flaA, mshA and vps genes, we previously demonstrated that transcription in monolayer cells is distinct from that in Biofilm cells and that the genetic requirements of monolayer formation are distinct from those of Biofilm formation. In this work, we sought to identify additional stage-specific genetic requirements through microarray analysis of the V. cholerae transcriptome during Biofilm Development. These studies demonstrated unique patterns of transcription in the planktonic, monolayer and Biofilm stages of Biofilm Development. Based on our microarray results, we selected cheY-3 as well as two previously uncharacterized genes, bap1 and leuO, for targeted mutation. The ΔcheY-3 mutant displayed a defect in monolayer but not Biofilm formation, suggesting that chemotaxis plays a stage-specific role in formation of the V. cholerae monolayer. Mutants carrying deletions in bap1 and leuO formed monolayers that were indistinguishable from those formed by wild-type V. cholerae. In contrast, these mutants displayed greatly decreased Biofilm accumulation. Our microarray analyses document modulation of the transcriptome of V. cholerae as it progresses through the stages in Biofilm Development. These studies demonstrate that microarray analysis of the transcriptome of Biofilm Development may greatly accelerate the discovery of novel targets for stage-specific inhibition of Biofilm Development.
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Environmental determinants of Vibrio cholerae Biofilm Development.
Applied and environmental microbiology, 2003Co-Authors: Katharine Kierek, Paula I. WatnickAbstract:Vibrio cholerae is a versatile bacterium that flourishes in diverse environments, including the human intestine, rivers, lakes, estuaries, and the ocean. Surface attachment is believed to be essential for colonization of all of these natural environments. Previous studies have demonstrated that the vps genes, which encode proteins required for exopolysaccharide synthesis and transport, are required for V. cholerae Biofilm Development in Luria-Bertani broth. In this work, we showed that V. cholerae forms vps-dependent Biofilms and vps-independent Biofilms. The vps-dependent and -independent Biofilms differ in their environmental activators and in architecture. Our results suggest that environmental activators of vps-dependent Biofilm Development are present in freshwater, while environmental activators of vps-independent Biofilm Development are present in seawater. The distinct environmental requirements for the two modes of Biofilm Development suggest that vps-dependent Biofilm Development and vps-independent Biofilm Development may play distinct roles in the natural environment.
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Vibrio cholerae CytR is a repressor of Biofilm Development.
Molecular microbiology, 2002Co-Authors: Adam J. Haugo, Paula I. WatnickAbstract:Vibrio cholerae is both a human pathogen and a natural inhabitant of aquatic environments. In the aquatic environment, microorganisms are found attached to surfaces in structures known as Biofilms. We have identified a transcriptional repressor in V. cholerae that inhibits exopolysaccharide synthesis and Biofilm Development. Our studies show that this repressor is the V. cholerae homologue of Escherichia coli CytR, a protein that represses nucleoside uptake and catabolism when nucleosides are scarce. We propose that the role of CytR in V. cholerae Biofilm Development is to co-ordinate bacterial Biofilm accumulation with the presence of nucleosides. Thus, nucleosides may be a signal to planktonic cells to join the Biofilm.
Jorge A. Benitez - One of the best experts on this subject based on the ideXlab platform.
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role of methylthioadenosine s adenosylhomocysteine nucleosidase in vibrio cholerae cellular communication and Biofilm Development
Biochemical and Biophysical Research Communications, 2015Co-Authors: Anisia J. Silva, William B. Parker, Paula W. Allan, Julio C. Ayala, Jorge A. BenitezAbstract:In Vibrio cholerae, the genes required for Biofilm Development are repressed by quorum sensing at high cell density due to the accumulation in the medium of two signaling molecules, cholera autoinducer 1 (CAI-1) and autoinducer 2 (AI-2). A significant fraction of toxigenic V. cholerae isolates, however, exhibit dysfunctional quorum sensing pathways. It was reported that transition state analogs of the enzyme methylthioadenosine/S-adenosylhomocysteine nucleosidase (MtnN) required to make AI-2 inhibited Biofilm formation in the prototype quorum sensing-deficient strain N16961. This finding prompted us to examine the role of both autoinducers and MtnN in Biofilm Development and virulence gene expression in a quorum sensing-deficient genetic background. Here we show that deletion of mtnN encoding methylthioadenosine/S-adenosylhomocysteine nucleosidase, cqsA (CAI-1), and/or luxS (AI-2) do not prevent Biofilm Development. However, two independent mtnN mutants exhibited diminished growth rate and motility in swarm agar plates suggesting that, under certain conditions, MtnN could influence Biofilm formation indirectly. Nevertheless, MtnN is not required for the Development of a mature Biofilm.
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Role of methylthioadenosine/S-adenosylhomocysteine nucleosidase in Vibrio cholerae cellular communication and Biofilm Development.
Biochemical and biophysical research communications, 2015Co-Authors: Anisia J. Silva, William B. Parker, Paula W. Allan, Julio C. Ayala, Jorge A. BenitezAbstract:In Vibrio cholerae, the genes required for Biofilm Development are repressed by quorum sensing at high cell density due to the accumulation in the medium of two signaling molecules, cholera autoinducer 1 (CAI-1) and autoinducer 2 (AI-2). A significant fraction of toxigenic V. cholerae isolates, however, exhibit dysfunctional quorum sensing pathways. It was reported that transition state analogs of the enzyme methylthioadenosine/S-adenosylhomocysteine nucleosidase (MtnN) required to make AI-2 inhibited Biofilm formation in the prototype quorum sensing-deficient strain N16961. This finding prompted us to examine the role of both autoinducers and MtnN in Biofilm Development and virulence gene expression in a quorum sensing-deficient genetic background. Here we show that deletion of mtnN encoding methylthioadenosine/S-adenosylhomocysteine nucleosidase, cqsA (CAI-1), and/or luxS (AI-2) do not prevent Biofilm Development. However, two independent mtnN mutants exhibited diminished growth rate and motility in swarm agar plates suggesting that, under certain conditions, MtnN could influence Biofilm formation indirectly. Nevertheless, MtnN is not required for the Development of a mature Biofilm.
Staffan Kjelleberg - One of the best experts on this subject based on the ideXlab platform.
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Dynamic modelling of cell death during Biofilm Development
Journal of theoretical biology, 2011Co-Authors: Magnus Fagerlind, Jeremy S. Webb, Nicolas Barraud, Diane Mcdougald, Andreas Jansson, Patric Nilsson, Mikael Harlén, Staffan Kjelleberg, Scott A. RiceAbstract:Biofilms are currently recognised as the predominant bacterial life-style and it has been suggested that Biofilm Development is influenced by a number of different processes such as adhesion, detachment, mass transport, quorum sensing, cell death and active dispersal. One of the least understood processes and its effects on Biofilm Development is cell death. However, experimental studies suggest that bacterial death is an important process during Biofilm Development and many studies show a relationship between cell death and dispersal in microbial Biofilms. We present a model of the process of cell death during Biofilm Development, with a particular focus on the spatial localisation of cell death or cell damage. Three rules governing cell death or cell damage were evaluated which compared the effects of starvation, damage accumulation, and viability during Biofilm Development and were also used to design laboratory based experiments to test the model. Results from model simulations show that actively growing Biofilms develop steep nutrient gradients within the interior of the Biofilm that affect neighbouring microcolonies resulting in cell death and detachment. Two of the rules indicated that high substrate concentrations lead to accelerated cell death, in contrast to the third rule, based on the accumulation of damage, which predicted earlier cell death for Biofilms grown with low substrate concentrations. Comparison of the modelling results with experimental results suggests that cell death is favoured under low nutrient conditions and that the accumulation of damage may be the main cause of cell death during Biofilm Development.
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Bacteriophage and Phenotypic Variation in Pseudomonas aeruginosa Biofilm Development
Journal of bacteriology, 2004Co-Authors: Jeremy S. Webb, Mathew Thye Ngak Lau, Staffan KjellebergAbstract:A current question in Biofilm research is whether Biofilm-specific genetic processes can lead to differentiation in physiology and function among Biofilm cells. In Pseudomonas aeruginosa, phenotypic variants which exhibit a small-colony phenotype on agar media and a markedly accelerated pattern of Biofilm Development compared to that of the parental strain are often isolated from Biofilms. We grew P. aeruginosa Biofilms in glass flow cell reactors and observed that the emergence of small-colony variants (SCVs) in the effluent runoff from the Biofilms correlated with the emergence of plaque-forming Pf1-like filamentous phage (designated Pf4) from the Biofilm. Because several recent studies have shown that bacteriophage genes are among the most highly upregulated groups of genes during Biofilm Development, we investigated whether Pf4 plays a role in SCV formation during P. aeruginosa Biofilm Development. We carried out immunoelectron microscopy using anti-Pf4 antibodies and observed that SCV cells, but not parental-type cells, exhibited high densities of Pf4 filaments on the cell surface and that these filaments were often tightly interwoven into complex latticeworks surrounding the cells. Moreover, infection of P. aeruginosa planktonic cultures with Pf4 caused the emergence of SCVs within the culture. These SCVs exhibited enhanced attachment, accelerated Biofilm Development, and large regions of dead and lysed cells inside microcolonies in a manner identical to that of SCVs obtained from Biofilms. We concluded that Pf4 can mediate phenotypic variation in P. aeruginosa Biofilms. We also performed partial sequencing and analysis of the Pf4 replicative form and identified a number of open reading frames not previously recognized in the genome of P. aeruginosa, including a putative postsegregational killing operon.
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Biofilm Development and cell death in the marine bacterium Pseudoalteromonas tunicata.
Applied and environmental microbiology, 2004Co-Authors: Anne Mai-prochnow, Jeremy S. Webb, Flavia F. Evans, Doralyn Dalisay-saludes, Sacha Stelzer, Suhelen Egan, Sally James, Staffan KjellebergAbstract:The newly described green-pigmented bacterium Pseudoalteromonas tunicata (D2) produces target-specific inhibitory compounds against bacteria, algae, fungi, and invertebrate larvae and is frequently found in association with living surfaces in the marine environment. As part of our studies on the ecology of P. tunicata and its interaction with marine surfaces, we examined the ability of P. tunicata to form Biofilms under continuous culture conditions within the laboratory. P. tunicata Biofilms exhibited a characteristic architecture consisting of differentiated microcolonies surrounded by water channels. Remarkably, we observed a repeatable pattern of cell death during Biofilm Development of P. tunicata, similar to that recently reported for Biofilms of Pseudomonas aeruginosa (J. S. Webb et al., J. Bacteriol. 185:4585-4595, 2003). Killing and lysis occurred inside microcolonies, apparently resulting in the formation of voids within these structures. A subpopulation of viable cells was always observed within the regions of killing in the Biofilm. Moreover, extensive killing in mature Biofilms appeared to result in detachment of the Biofilm from the substratum. A novel 190-kDa autotoxic protein produced by P. tunicata, designated AlpP, was found to be involved in this Biofilm killing and detachment. A ΔalpP mutant derivative of P. tunicata was generated, and this mutant did not show cell death during Biofilm Development. We propose that AlpP-mediated cell death plays an important role in the multicellular Biofilm Development of P. tunicata and subsequent dispersal of surviving cells within the marine environment.
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Quorum Sensing-Controlled Biofilm Development in Serratia liquefaciens MG1
Journal of bacteriology, 2004Co-Authors: Maurizio Labbate, Scott A. Rice, Shu Yeong Queck, Kai Shyang Koh, Michael Givskov, Staffan KjellebergAbstract:Serratia liquefaciens MG1 contains an N-acylhomoserine lactone-mediated quorum-sensing system that is known to regulate swarming motility colonization. In this study, we describe for S. liquefaciens MG1 the Development of a novel Biofilm consisting of cell aggregates and differentiated cell types, such as cell chains and long filamentous cells. Furthermore, quorum sensing is shown to be crucial for normal Biofilm Development and for elaborate differentiation. A mutant of S. liquefaciens MG1 that was incapable of synthesizing extracellular signal formed a thin and nonmature Biofilm lacking cell aggregates and differentiated cell chains. Signal-based complementation of this mutant resulted in a Biofilm with the wild-type architecture. Two quorum-sensing-regulated genes (bsmA and bsmB) involved in Biofilm Development were identified, and we propose that these genes are engaged in fine-tuning the formation of cell aggregates at a specific point in Biofilm Development.
Rajendar Deora - One of the best experts on this subject based on the ideXlab platform.
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Transcriptome Profiling Reveals Stage-Specific Production and Requirement of Flagella during Biofilm Development in Bordetella bronchiseptica
PloS one, 2012Co-Authors: Tracy L. Nicholson, Matt S. Conover, Rajendar DeoraAbstract:We have used microarray analysis to study the transcriptome of the bacterial pathogen Bordetella bronchiseptica over the course of five time points representing distinct stages of Biofilm Development. The results suggest that B. bronchiseptica undergoes a coordinately regulated gene expression program similar to a bacterial Developmental process. Expression and subsequent production of the genes encoding flagella, a classical Bvg− phase phenotype, occurs and is under tight regulatory control during B. bronchiseptica Biofilm Development. Using mutational analysis, we demonstrate that flagella production at the appropriate stage of Biofilm Development, i.e. production early subsequently followed by repression, is required for robust Biofilm formation and maturation. We also demonstrate that flagella are necessary and enhance the initial cell-surface interactions, thereby providing mechanistic information on the initial stages of Biofilm Development for B. bronchiseptica. Biofilm formation by B. bronchiseptica involves the production of both Bvg-activated and Bvg-repressed factors followed by the repression of factors that inhibit formation of mature Biofilms.
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Role of a Putative Polysaccharide Locus in Bordetella Biofilm Development
Journal of bacteriology, 2006Co-Authors: Gina Parise, Meenu Mishra, Yoshikane Itoh, Tony Romeo, Rajendar DeoraAbstract:Bordetellae are gram-negative bacteria that colonize the respiratory tracts of animals and humans. We and others have recently shown that these bacteria are capable of living as sessile communities known as Biofilms on a number of abiotic surfaces. During the Biofilm mode of existence, bacteria produce one or more extracellular polymeric substances that function, in part, to hold the cells together and to a surface. There is little information on either the constituents of the Biofilm matrix or the genetic basis of Biofilm Development by Bordetella spp. By utilizing immunoblot assays and by enzymatic hydrolysis using dispersin B (DspB), a glycosyl hydrolase that specifically cleaves the polysaccharide poly-β-1,6-N-acetyl-d-glucosamine (poly-β-1,6-GlcNAc), we provide evidence for the production of poly-β-1,6-GlcNAc by various Bordetella species (Bordetella bronchiseptica, B. pertussis, and B. parapertussis) and its role in their Biofilm Development. We have investigated the role of a Bordetella locus, here designated bpsABCD, in Biofilm formation. The bps (Bordetella polysaccharide) locus is homologous to several bacterial loci that are required for the production of poly-β-1,6-GlcNAc and have been implicated in bacterial Biofilm formation. By utilizing multiple microscopic techniques to analyze Biofilm formation under both static and hydrodynamic conditions, we demonstrate that the bps locus, although not essential at the initial stages of Biofilm formation, contributes to the stability and the maintenance of the complex architecture of Bordetella Biofilms.
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The BvgAS Signal Transduction System Regulates Biofilm Development in Bordetella
Journal of bacteriology, 2005Co-Authors: Meenu Mishra, Gina Parise, Daniel J Wozniak, Kara D. Jackson, Rajendar DeoraAbstract:The majority of Bordetella sp. virulence determinants are regulated by the BvgAS signal transduction system. BvgAS mediates the control of multiple phenotypic phases and a spectrum of gene expression profiles specific to each phase in response to incremental changes in the concentrations of environmental signals. Studies highlighting the critical role of this signaling circuitry in the Bordetella infectious cycle have focused on planktonically growing bacterial cells. It is becoming increasingly clear that the major mode of bacterial existence in the environment and within the body is a surface-attached state known as a Biofilm. Biofilms are defined as consortia of sessile microorganisms that are embedded in a matrix. During routine growth of Bordetella under agitating conditions, we noticed the formation of a bacterial ring at the air-liquid interface of the culture tubes. We show here that this surface adherence property reflects the ability of these organisms to form Biofilms. Our data demonstrate that the BvgAS locus regulates Biofilm Development in Bordetella. The results reported in this study suggest that the Bvg-mediated control in Biofilm Development is exerted at later time points after the initial attachment of bacteria to the different surfaces. Additionally, we show that these Biofilms are highly tolerant of a number of antimicrobials, including the ones that are currently recommended for treatment of veterinary and human infections caused by Bordetella spp. Finally, we discuss the significance of the Biofilm lifestyle mode as a potential contributor to persistent infections.
Alexander R. Horswill - One of the best experts on this subject based on the ideXlab platform.
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Impact of Environmental Cues on Staphylococcal Quorum Sensing and Biofilm Development
The Journal of biological chemistry, 2016Co-Authors: Jeffrey S. Kavanaugh, Alexander R. HorswillAbstract:Staphylococci are commensal bacteria that colonize the epithelial surfaces of humans and many other mammals. These bacteria can also attach to implanted medical devices and develop surface-associated Biofilm communities that resist clearance by host defenses and available chemotherapies. These communities are often associated with persistent staphylococcal infections that place a tremendous burden on the healthcare system. Understanding the regulatory program that controls staphylococcal Biofilm Development, as well as the environmental conditions that modulate this program, has been a focal point of research in recent years. A central regulator controlling Biofilm Development is a peptide quorum-sensing system, also called the accessory gene regulator or agr system. In the opportunistic pathogen Staphylococcus aureus, the agr system controls production of exo-toxins and exo-enzymes essential for causing infections, and simultaneously, it modulates the ability of this pathogen to attach to surfaces and develop a Biofilm, or to disperse from the Biofilm state. In this review, we explore advances on the interconnections between the agr quorum-sensing system and Biofilm mechanisms, and topics covered include recent findings on how different environmental conditions influence quorum sensing, the impact on Biofilm Development, and ongoing questions and challenges in the field. As our understanding of the quorum sensing and Biofilm interconnection advances, there are growing opportunities to take advantage of this knowledge and develop therapeutic approaches to control staphylococcal infections.
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Temporal and Stochastic Control of Staphylococcus aureus Biofilm Development
mBio, 2014Co-Authors: Derek E. Moormeier, Jeffrey L. Bose, Alexander R. Horswill, Kenneth W. BaylesAbstract:Biofilm communities contain distinct microniches that result in metabolic heterogeneity and variability in gene ex- pression. Previously, these niches were visualized within Staphylococcus aureus Biofilms by observing differential expression of thecidandlrgoperons during tower formation. In the present study, we examined early Biofilm Development and identified two new stages (designated "multiplication" and "exodus") that were associated with changes in matrix composition and a distinct reorganization of the cells as the Biofilm matured. The initial attachment and multiplication stages were shown to be protease sensitive but independent of most cell surface-associated proteins. Interestingly, after6hofgrowth, an exodus of the Biofilm population that followed the transition of the Biofilm to DNase I sensitivity was demonstrated. Furthermore, disruption of the gene encoding staphylococcal nuclease (nuc) abrogated this exodus event, causing hyperproliferation of the Biofilm and disrupt- ing normal tower Development. Immediately prior to the exodus event, S. aureus cells carrying a nuc::gfp promoter fusion dem- onstrated Sae-dependent expression but only in an apparently random subpopulation of cells. In contrast to the existing model for tower Development in S. aureus, the results of this study suggest the presence of a Sae-controlled nuclease-mediated exodus of Biofilm cells that is required for the Development of tower structures. Furthermore, these studies indicate that the differential expression ofnucduring Biofilm Development is subject to stochastic regulatory mechanisms that are independent of the forma- tion of metabolic microniches. IMPORTANCE In this study, we provide a novel view of four early stages of Biofilm formation by the human pathogen Staphylococ- cus aureus. We identified an initial nucleoprotein matrix during Biofilm Development that is DNase I insensitive until a critical point when a nuclease-mediated exodus of the population is induced prior to tower formation. Unlike the previously described dispersal of cells that occurs after tower Development, we found that the mechanism controlling this exodus event is dependent on the Sae regulatory system and independent of Agr. In addition, we revealed that the gene encoding the secreted staphylococcal nuclease was expressed in only a subpopulation of cells, consistent with a model in which Biofilms exhibit multicellular charac- teristics, including the presence of specialized cells and a division of labor that imparts functional consequences to the remain- der of the population.