scholarly journals Role of CpxR in Biofilm Development: Expression of Key Fimbrial, O-Antigen and Virulence Operons of Salmonella Enteritidis

2019 ◽  
Vol 20 (20) ◽  
pp. 5146
Author(s):  
Deeksha Shetty ◽  
Juan Abrahante ◽  
Samuel Chekabab ◽  
Xuxiaochen Wu ◽  
Darren Korber ◽  
...  
Keyword(s):  
Sigma Factor ◽  
Salmonella Enteritidis ◽  
Constitutive Expression ◽  
Microtiter Plate ◽  
Two Component System ◽  
Swarming Motility ◽  
O Antigen ◽  
Wild Type Counterpart ◽  
Health Significance ◽  
Biofilm Development

Salmonella Enteritidis is a non-typhoidal serovar of great public health significance worldwide. The RpoE sigma factor and CpxRA two-component system are the major regulators of the extracytoplasmic stress response. In this study, we found that the CpxR has highly significant, but opposite effects on the auto-aggregation and swarming motility of S. Enteritidis. Auto-aggregation was negatively affected in the ∆cpxR mutant, whereas the same mutant significantly out-performed its wild-type counterpart with respect to swarming motility, indicating that the CpxR plays a role in biofilm-associated phenotypes. Indeed, biofilm-related assays showed that the CpxR is of critical importance in biofilm development under both static (microtiter plate) and dynamic (flow cell) media flow conditions. In contrast, the RpoE sigma factor showed no significant role in biofilm development under dynamic conditions. Transcriptomic analysis revealed that the cpxR mutation negatively affected the constitutive expression of the operons critical for biosynthesis of O-antigen and adherence, but positively affected the expression of virulence genes critical for Salmonella-mediated endocytosis. Conversely, CpxR induced the expression of curli csgAB and fimbrial stdAC operons only during biofilm development and flagellar motAB and fliL operons exclusively during the planktonic phase, indicating a responsive biofilm-associated loop of the CpxR regulator.

scholarly journals Rosmarinic Acid Interaction with Planktonic and Biofilm Staphylococcus aureus

2013 ◽  
Vol 8 (12) ◽  
pp. 1934578X1300801 ◽  
Author(s):  
Lívia Slobodníková ◽  
Silvia Fialová ◽  
Helena Hupková ◽  
Daniel Grančai
Keyword(s):  
Staphylococcus Aureus ◽  
Rosmarinic Acid ◽  
Antibacterial Activities ◽  
Microtiter Plate ◽  
Dependent Manner ◽  
Early Stages ◽  
Microtiter Plates ◽  
Sole Agent ◽  
The Subject ◽  
Biofilm Development

The subject of study was the evaluation of antibacterial activities of rosmarinic acid (RA) on clinical Staphylococcus aureus strains obtained from catheter-related infections. Minimal inhibitory (MIC) and minimal bactericidal concentrations (MBC) of RA were tested by broth microdilution assay. Biofilm-eradication activity was detected on 24-hour biofilm in microtiter plates using a regrowth technique; activity on biofilm formation was measured by a microtiter plate method after RA application to bacterial samples after 0, 1, 3 and 6 hours of biofilm development. RA had antimicrobial activity on all tested strains in concentrations from 625 to 1250 μg.mL−1 (MICs equal to MBCs). No biofilm-eradication activity on 24-hour biofilm was observed in the tested range of concentrations (from 156 to 5000 μg.mL−1). Subinhibitory RA concentrations suppressed the biofilm production, when applied at early stages of its development. Concentrations lower than subinhibitory stimulated the biofilm mass production in a concentration- and time-dependent manner. Considering our results, RA could be a candidate for a topical antimicrobial agent with killing activity on planktonic forms of bacteria and suppressing activity in the early stages of biofilm development, but probably not for the therapy of catheter-related infections as a sole agent.

scholarly journals Peptide Mix from Olivancillaria hiatula Interferes with Cell-to-Cell Communication in Pseudomonas aeruginosa

2019 ◽  
Vol 2019 ◽  
pp. 1-12
Author(s):  
Edward Ntim Gasu ◽  
Hubert Senanu Ahor ◽  
Lawrence Sheringham Borquaye
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Quorum Sensing ◽  
Biofilm Formation ◽  
Antimicrobial Agents ◽  
Cell Communication ◽  
Microtiter Plate ◽  
Antibiofilm Activity ◽  
Dependent Manner ◽  
Biofilm Inhibition ◽  
Swarming Motility

Bacteria in biofilms are encased in an extracellular polymeric matrix that limits exposure of microbial cells to lethal doses of antimicrobial agents, leading to resistance. In Pseudomonas aeruginosa, biofilm formation is regulated by cell-to-cell communication, called quorum sensing. Quorum sensing facilitates a variety of bacterial physiological functions such as swarming motility and protease, pyoverdine, and pyocyanin productions. Peptide mix from the marine mollusc, Olivancillaria hiatula, has been studied for its antibiofilm activity against Pseudomonas aeruginosa. Microscopy and microtiter plate-based assays were used to evaluate biofilm inhibitory activities. Effect of the peptide mix on quorum sensing-mediated processes was also evaluated. Peptide mix proved to be a good antibiofilm agent, requiring less than 39 μg/mL to inhibit 50% biofilm formation. Micrographs obtained confirmed biofilm inhibition at 1/2 MIC whereas 2.5 mg/mL was required to degrade preformed biofilm. There was a marked attenuation in quorum sensing-mediated phenotypes as well. At 1/2 MIC of peptide, the expression of pyocyanin, pyoverdine, and protease was inhibited by 60%, 72%, and 54%, respectively. Additionally, swarming motility was repressed by peptide in a dose-dependent manner. These results suggest that the peptide mix from Olivancillaria hiatula probably inhibits biofilm formation by interfering with cell-to-cell communication in Pseudomonas aeruginosa.

scholarly journals Homologous c-di-GMP-Binding Scr Transcription Factors Orchestrate Biofilm Development in Vibrio parahaemolyticus

2020 ◽  
Vol 202 (6) ◽  
Author(s):  
John H. Kimbrough ◽  
J. Thomas Cribbs ◽  
Linda L. McCarter
Keyword(s):  
Transcription Factors ◽  
Biofilm Formation ◽  
Vibrio Parahaemolyticus ◽  
Matrix Protein ◽  
Second Messenger ◽  
Minor Role ◽  
Binding Motif ◽  
Swarming Motility ◽  
Content Type ◽  
Biofilm Development

ABSTRACT The marine bacterium and human pathogen Vibrio parahaemolyticus rapidly colonizes surfaces by using swarming motility and forming robust biofilms. Entering one of the two colonization programs, swarming motility or sessility, involves differential regulation of many genes, resulting in a dramatic shift in physiology and behavior. V. parahaemolyticus has evolved complex regulation to control these two processes that have opposing outcomes. One mechanism relies on the balance of the second messenger c-di-GMP, where high c-di-GMP favors biofilm formation. V. parahaemolyticus possesses four homologous regulators, the Scr transcription factors, that belong in a Vibrio-specific family of W[F/L/M][T/S]R motif transcriptional regulators, some members of which have been demonstrated to bind c-di-GMP. In this work, we explore the role of these Scr regulators in biofilm development. We show that each protein binds c-di-GMP, that this binding requires a critical R in the binding motif, and that the biofilm-relevant activities of CpsQ, CpsS, and ScrO but not ScrP are dependent upon second messenger binding. ScrO and CpsQ are the primary drivers of biofilm formation, as biofilms are eliminated when both of these regulators are absent. ScrO is most important for capsule expression. CpsQ is most important for RTX-matrix protein expression, although it contributes to capsule expression when c-di-GMP levels are high. Both regulators contribute to O-antigen ligase expression. ScrP works oppositely in a minor role to repress the ligase gene. CpsS plays a regulatory checkpointing role by negatively modulating expression of these biofilm-pertinent genes under fluctuating c-di-GMP conditions. Our work further elucidates the multifactorial network that contributes to biofilm development in V. parahaemolyticus. IMPORTANCE Vibrio parahaemolyticus can inhabit open ocean, chitinous shells, and the human gut. Such varied habitats and the transitions between them require adaptable regulatory networks controlling energetically expensive behaviors, including swarming motility and biofilm formation, which are promoted by low and high concentrations of the signaling molecule c-di-GMP, respectively. Here, we describe four homologous c-di-GMP-binding Scr transcription factors in V. parahaemolyticus. Members of this family of regulators are present in many vibrios, yet their numbers and the natures of their activities differ across species. Our work highlights the distinctive roles that these transcription factors play in dynamically controlling biofilm formation and architecture in V. parahaemolyticus and serves as a powerful example of regulatory network evolution and diversification.

scholarly journals Identification of three new GGDEF and EAL domain-containing proteins participating in the Scr surface colonization regulatory network in Vibrio parahaemolyticus

2020 ◽  
Author(s):  
John H. Kimbrough ◽  
Linda L. McCarter
Keyword(s):  
Regulatory Network ◽  
Vibrio Parahaemolyticus ◽  
Enteric Bacteria ◽  
Swarming Motility ◽  
Surface Colonization ◽  
Diguanylate Cyclases ◽  
Surface Sensing ◽  
Formation Programs ◽  
Catalytic Motif ◽  
Biofilm Development

Vibrio parahaemolyticus rapidly colonizes surfaces using swarming motility. Surface contact induces the surface sensing regulon including lateral flagellar genes, spurring dramatic shifts in physiology and behavior. The bacterium can also adopt a sessile, surface-associated lifestyle and form robust biofilms. These alternate colonization strategies are influenced reciprocally by the second messenger c-di-GMP. Although V. parahaemolyticus possesses 43 predicted proteins with the c-di-GMP-forming GGDEF domain, none have been previously been identified as contributors to surface colonization. We sought to explore this knowledge gap by using a suppressor transposon screen to restore swarming motility of a non-swarming, high c-di-GMP strain. Two diguanylate cyclases, ScrJ and ScrL, each containing tetratricopeptide repeat coupled GGDEF domains were demonstrated to contribute additively to swarming gene repression. Both proteins required an intact catalytic motif to regulate. Another suppressor mapped in lafV, the last gene in a lateral flagellar operon. Containing a degenerate phosphodiesterase (EAL) domain, LafV affected expression of multiple genes in the surface sensing regulon and required LafK, a primary swarming activator, to repress. Mutation of the signature EAL motif had little effect on LafV’s repressive activity, suggesting LafV belongs to the subclass of EAL-type proteins that are regulatory but not enzymatic. Consistent with these activities and their predicted effects on c-di-GMP, scrJ and scrL, but not lafV mutants affected transcription of the c-di-GMP-responsive, biofilm reporter cpsA::lacZ. Our results expand the knowledge of the V. parahaemolyticus GGDEF/EAL repertoire and their roles in this surface colonization regulatory network. Significance A key survival decision, in the environment or the host, is whether to emigrate or aggregate. In bacteria, c-di-GMP signaling almost universally influences solutions to this dilemma. In V. parahaemolyticus, c-di-GMP reciprocally regulates swarming and sticking (i.e., biofilm formation) programs of surface colonization. Key c-di-GMP degrading phosphodiesterases responsive to quorum and nutritional signals have been previously identified. c-di-GMP-binding transcription factors programming biofilm development have been studied. Here, we further develop the blueprint of the c-di-GMP network by identifying new participants involved in dictating the complex decision of whether to swarm or stay. These include diguanylate cyclases with tetratricopeptide domains and a degenerate EAL protein that serves, analogous to the negative flagellar regulator RflP/YdiV of enteric bacteria, to regulate swarming.

scholarly journals Evaluation of a Novel Enzyme-Linked Immunosorbent Assay for Detection of Antibodies against Salmonella, Employing a Stable Coating of Lipopolysaccharide-Derived Antigens Covalently Attached to Polystyrene Microwells

2000 ◽  
Vol 12 (2) ◽  
pp. 130-135 ◽  
Author(s):  
Camilla Wiuff ◽  
Eva Stenbæk Jauho ◽  
Henrik Stryhn ◽  
Lars Ole Andresen ◽  
Karina Thaulov ◽  
...  
Keyword(s):  
Microtiter Plate ◽  
Enzyme Linked Immunosorbent Assay ◽  
O Antigen ◽  
Salmonella Infections ◽  
Specificity And Sensitivity ◽  
Conventional Elisa ◽  
Polysaccharide Antigens ◽  
Serological Surveillance ◽  
Coated Surface ◽  
High Degree

Polysaccharides derived from Salmonella typhimurium lipopolysaccharide (LPS) representing the O-antigen factors 1, 4, 5, and 12 and the O-antigen factors 6 and 7 from Salmonella choleraesuis LPS were derivatized with the photoreactive compound anthraquinone and subsequently covalently coupled to microtiter polystyrene plates by ultraviolet irradiation. Both polysaccharide antigens could be coupled simultaneously to the same microtiter plate. The coated surface was used in indirect ELISA for the determination of serum antibodies from pigs infected with bacteria of the two Salmonella groups and from uninfected pigs. This ELISA proved itself by having a good long-term durability and a high degree of reproducibility, including low day-to-day variations and low interplate variations. Furthermore, the ELISA showed good specificity and sensitivity when data were compared with the optical density levels of a panel of pig sera as determined by a conventional ELISA on the basis of passive coating of the two Salmonella LPS antigens (the mix-ELISA). The covalent anthraquinone mix-ELISA shows promise as a stable and durable alternative to the existing conventional ELISA for serological surveillance of Salmonella infections in pigs.

scholarly journals Cryptococcus neoformans Biofilm Formation Depends on Surface Support and Carbon Source and Reduces Fungal Cell Susceptibility to Heat, Cold, and UV Light

2007 ◽  
Vol 73 (14) ◽  
pp. 4592-4601 ◽  
Author(s):  
Luis R. Martinez ◽  
Arturo Casadevall
Keyword(s):  
Cryptococcus Neoformans ◽  
Biofilm Formation ◽  
Capsular Polysaccharide ◽  
Uv Light ◽  
Surface Characteristics ◽  
Extracellular Polysaccharide ◽  
Microtiter Plate ◽  
Support Surface ◽  
Fungal Cell ◽  
Biofilm Development

ABSTRACT The fungus Cryptococcus neoformans possesses a polysaccharide capsule and can form biofilms on medical devices. We describe the characteristics of C. neoformans biofilm development using a microtiter plate model, microscopic examinations, and a colorimetric 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-2H-tetrazolium-hydroxide (XTT) reduction assay to observe the metabolic activity of cryptococci within a biofilm. A strong correlation between XTT and CFU assays was demonstrated. Chemical analysis of the exopolymeric material revealed sugar composition consisting predominantly of xylose, mannose, and glucose, indicating the presence of other polysaccharides in addition to glucurunoxylomannan. Biofilm formation was affected by surface support differences, conditioning films on the surface, characteristics of the medium, and properties of the microbial cell. A specific antibody to the capsular polysaccharide of this fungus was used to stain the extracellular polysaccharide matrix of the fungal biofilms using light and confocal microscopy. Additionally, the susceptibility of C. neoformans biofilms and planktonic cells to environmental stress was investigated using XTT reduction and CFU assays. Biofilms were less susceptible to heat, cold, and UV light exposition than their planktonic counterparts. Our findings demonstrate that fungal biofilm formation is dependent on support surface characteristics and that growth in the biofilm state makes fungal cells less susceptible to potential environmental stresses.

scholarly journals Quorum Sensing in Yersinia enterocolitica Controls Swimming and Swarming Motility

2006 ◽  
Vol 188 (4) ◽  
pp. 1451-1461 ◽  
Author(s):  
Steve Atkinson ◽  
Chien-Yi Chang ◽  
R. Elizabeth Sockett ◽  
Miguel Cámara ◽  
Paul Williams
Keyword(s):  
Mass Spectrometry ◽  
Yersinia Enterocolitica ◽  
Sigma Factor ◽  
Parent Strain ◽  
Homoserine Lactone ◽  
Swarming Motility ◽  
Acylhomoserine Lactone ◽  
Reverse Transcription Pcr ◽  
Factor Expression ◽  
Long Chain

ABSTRACT The Yersinia enterocolitica LuxI homologue YenI directs the synthesis of N-3-(oxohexanoyl)homoserine lactone (3-oxo-C6-HSL) and N-hexanoylhomoserine lactone (C6-HSL). In a Y. enterocolitica yenI mutant, swimming motility is temporally delayed while swarming motility is abolished. Since both swimming and swarming are flagellum dependent, we purified the flagellin protein from the parent and yenI mutant. Electrophoresis revealed that in contrast to the parent strain, the yenI mutant grown for 17 h at 26°C lacked the 45-kDa flagellin protein FleB. Reverse transcription-PCR indicated that while mutation of yenI had no effect on yenR, flhDC (the motility master regulator) or fliA (the flagellar sigma factor) expression, fleB (the flagellin structural gene) was down-regulated. Since 3-oxo-C6-HSL and C6-HSL did not restore swimming or swarming in the yenI mutant, we reexamined the N-acylhomoserine lactone (AHL) profile of Y. enterocolitica. Using AHL biosensors and mass spectrometry, we identified three additional AHLs synthesized via YenI: N-(3-oxodecanoyl)homoserine lactone, N-(3-oxododecanoyl)homoserine lactone (3-oxo-C12-HSL), and N-(3-oxotetradecanoyl)homoserine lactone. However, none of the long-chain AHLs either alone or in combination with the short-chain AHLs restored swarming or swimming in the yenI mutant. By investigating the transport of radiolabeled 3-oxo-C12-HSL and by introducing an AHL biosensor into the yenI mutant we demonstrate that the inability of exogenous AHLs to restore motility to the yenI mutant is not related to a lack of AHL uptake. However, both AHL synthesis and motility were restored by complementation of the yenI mutant with a plasmid-borne copy of yenI.

scholarly journals Two GacA-Dependent Small RNAs Modulate the Quorum-Sensing Response in Pseudomonas aeruginosa

2006 ◽  
Vol 188 (16) ◽  
pp. 6026-6033 ◽  
Author(s):  
Elisabeth Kay ◽  
Bérénice Humair ◽  
Valérie Dénervaud ◽  
Kathrin Riedel ◽  
Stéphanie Spahr ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Quorum Sensing ◽  
Small Rnas ◽  
Homoserine Lactone ◽  
Two Component System ◽  
Bond Forming ◽  
Small Regulatory Rnas ◽  
Regulatory Rnas ◽  
Chitin Binding Protein ◽  
Biofilm Development

ABSTRACT In Pseudomonas aeruginosa, the GacS/GacA two-component system positively controls the quorum-sensing machinery and the expression of extracellular products via two small regulatory RNAs, RsmY and RsmZ. An rsmY rsmZ double mutant and a gacA mutant were similarly impaired in the synthesis of the quorum-sensing signal N-butanoyl-homoserine lactone, the disulfide bond-forming enzyme DsbA, and the exoproducts hydrogen cyanide, pyocyanin, elastase, chitinase (ChiC), and chitin-binding protein (CbpD). Both mutants showed increased swarming ability, azurin release, and early biofilm development.

scholarly journals Oxygen-Mediated Regulation of Biofilm Development Is Controlled by the Alternative Sigma Factor σB in Staphylococcus epidermidis

2008 ◽  
Vol 75 (1) ◽  
pp. 261-264 ◽  
Author(s):  
John J. Cotter ◽  
James P. O'Gara ◽  
Dietrich Mack ◽  
Eoin Casey
Keyword(s):  
Staphylococcus Epidermidis ◽  
Sigma Factor ◽  
Rotating Disk ◽  
Negative Regulator ◽  
Alternative Sigma Factor ◽  
Polysaccharide Intercellular Adhesin ◽  
Anaerobic Conditions ◽  
Rotating Disk Reactor ◽  
Biofilm Development

ABSTRACT Using a modified rotating-disk reactor to sparge oxygen to Staphylococcus epidermidis cultures, we found that oxygen negatively regulates biofilm development by influencing the activity of σB. Under anaerobic conditions, increased σB activity activates icaADBC, which encodes enzymes responsible for polysaccharide intercellular adhesin synthesis, by repressing transcription of the negative regulator icaR.

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