scholarly journals A Cardiolipin-Deficient Mutant of Rhodobacter sphaeroides Has an Altered Cell Shape and Is Impaired in Biofilm Formation

2015 ◽  
Vol 197 (21) ◽  
pp. 3446-3455 ◽  
Author(s):  
Ti-Yu Lin ◽  
Thiago M. A. Santos ◽  
Wayne S. Kontur ◽  
Timothy J. Donohue ◽  
Douglas B. Weibel
Keyword(s):  
Rhodobacter Sphaeroides ◽  
Biofilm Formation ◽  
Cell Shape ◽  
Bacterial Attachment ◽  
Growth Defect ◽  
Deficient Mutant ◽  
Membrane Composition ◽  
Anionic Phospholipid ◽  
Content Type ◽  
Altered Cell

ABSTRACTCell shape has been suggested to play an important role in the regulation of bacterial attachment to surfaces and the formation of communities associated with surfaces. We found that a cardiolipin synthase (Δcls) mutant of the rod-shaped bacteriumRhodobacter sphaeroides—in which synthesis of the anionic, highly curved phospholipid cardiolipin (CL) is reduced by 90%—produces ellipsoid-shaped cells that are impaired in biofilm formation. Reducing the concentration of CL did not cause significant defects inR. sphaeroidescell growth, swimming motility, lipopolysaccharide and exopolysaccharide production, surface adhesion protein expression, and membrane permeability. Complementation of the CL-deficient mutant by ectopically expressing CL synthase restored cells to their rod shape and increased biofilm formation. TreatingR. sphaeroidescells with a low concentration (10 μg/ml) of the small-molecule MreB inhibitorS-(3,4-dichlorobenzyl)isothiourea produced ellipsoid-shaped cells that had no obvious growth defect yet reducedR. sphaeroidesbiofilm formation. This study demonstrates that CL plays a role inR. sphaeroidescell shape determination, biofilm formation, and the ability of the bacterium to adapt to its environment.IMPORTANCEMembrane composition plays a fundamental role in the adaptation of many bacteria to environmental stress. In this study, we build a new connection between the anionic phospholipid cardiolipin (CL) and cellular adaptation inRhodobacter sphaeroides. We demonstrate that CL plays a role in the regulation ofR. sphaeroidesmorphology and is important for the ability of this bacterium to form biofilms. This study correlates CL concentration, cell shape, and biofilm formation and provides the first example of how membrane composition in bacteria alters cell morphology and influences adaptation. This study also provides insight into the potential of phospholipid biosynthesis as a target for new chemical strategies designed to alter or prevent biofilm formation.

scholarly journals Cardiolipin Alters Rhodobacter sphaeroides Cell Shape by Affecting Peptidoglycan Precursor Biosynthesis

mBio ◽  
2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Ti-Yu Lin ◽  
William S. Gross ◽  
George K. Auer ◽  
Douglas B. Weibel
Keyword(s):  
Cell Wall ◽  
Rhodobacter Sphaeroides ◽  
Cell Morphology ◽  
Cell Shape ◽  
Molecular Mechanisms ◽  
Wild Type ◽  
Anionic Phospholipid ◽  
Content Type ◽  
Lipid Ii ◽  
Topological Features

ABSTRACT Cardiolipin (CL) is an anionic phospholipid that plays an important role in regulating protein biochemistry in bacteria and mitochondria. Deleting the CL synthase gene (Δcls) in Rhodobacter sphaeroides depletes CL and decreases cell length by 20%. Using a chemical biology approach, we found that a CL deficiency does not impair the function of the cell wall elongasome in R. sphaeroides; instead, biosynthesis of the peptidoglycan (PG) precursor lipid II is decreased. Treating R. sphaeroides cells with fosfomycin and d-cycloserine inhibits lipid II biosynthesis and creates phenotypes in cell shape, PG composition, and spatial PG assembly that are strikingly similar to those seen with R. sphaeroides Δcls cells, suggesting that CL deficiency alters the elongation of R. sphaeroides cells by reducing lipid II biosynthesis. We found that MurG—a glycosyltransferase that performs the last step of lipid II biosynthesis—interacts with anionic phospholipids in native (i.e., R. sphaeroides) and artificial membranes. Lipid II production decreases 25% in R. sphaeroides Δcls cells compared to wild-type cells, and overexpression of MurG in R. sphaeroides Δcls cells restores their rod shape, indicating that CL deficiency decreases MurG activity and alters cell shape. The R. sphaeroides Δcls mutant is more sensitive than the wild-type strain to antibiotics targeting PG synthesis, including fosfomycin, d-cycloserine, S-(3,4-dichlorobenzyl)isothiourea (A22), mecillinam, and ampicillin, suggesting that CL biosynthesis may be a potential target for combination chemotherapies that block the bacterial cell wall. IMPORTANCE The phospholipid composition of the cell membrane influences the spatial and temporal biochemistry of cells. We studied molecular mechanisms connecting membrane composition to cell morphology in the model bacterium Rhodobacter sphaeroides. The peptidoglycan (PG) layer of the cell wall is a dominant component of cell mechanical properties; consequently, it has been an important antibiotic target. We found that the anionic phospholipid cardiolipin (CL) plays a role in determination of the shape of R. sphaeroides cells by affecting PG precursor biosynthesis. Removing CL in R. sphaeroides alters cell morphology and increases its sensitivity to antibiotics targeting proteins synthesizing PG. These studies provide a connection to spatial biochemical control in mitochondria, which contain an inner membrane with topological features in common with R. sphaeroides.

scholarly journals Rcs Phosphorelay Activation in Cardiolipin-DeficientEscherichia coliReduces Biofilm Formation

2019 ◽  
Vol 201 (9) ◽  
Author(s):  
Julia F. Nepper ◽  
Yin C. Lin ◽  
Douglas B. Weibel
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Lipid Membrane ◽  
Protein Translocation ◽  
Membrane Composition ◽  
Anionic Phospholipid ◽  
Content Type ◽  
Envelope Stress ◽  
Biofilm Development

ABSTRACTBiofilm formation is a complex process that requires a number of transcriptional, proteomic, and physiological changes to enable bacterial survival. The lipid membrane presents a barrier to communication between the machinery within bacteria and the physical and chemical features of their extracellular environment, and yet little is known about how the membrane influences biofilm development. We found that depleting the anionic phospholipid cardiolipin reduces biofilm formation inEscherichia colicells by as much as 50%. The absence of cardiolipin activates the regulation of colanic acid synthesis (Rcs) envelope stress response, which represses the production of flagella, disrupts initial biofilm attachment, and reduces biofilm growth. We demonstrate that a reduction in the concentration of cardiolipin impairs translocation of proteins across the inner membrane, which we hypothesize activates the Rcs pathway through the outer membrane lipoprotein RcsF. Our study demonstrates a molecular connection between the composition of membrane phospholipids and biofilm formation inE. coliand suggests that altering lipid biosynthesis may be a viable approach for altering biofilm formation and possibly other multicellular phenotypes related to bacterial adaptation and survival.IMPORTANCEThere is a growing interest in the role of lipid membrane composition in the physiology and adaptation of bacteria. We demonstrate that a reduction in the anionic phospholipid cardiolipin impairs biofilm formation inEscherichia colicells. Depleting cardiolipin reduced protein translocation across the inner membrane and activated the Rcs envelope stress response. Consequently, cardiolipin depletion produced cells lacking assembled flagella, which impacted their ability to attach to surfaces and seed the earliest stage in biofilm formation. This study provides empirical evidence for the role of anionic phospholipid homeostasis in protein translocation and its effect on biofilm development and highlights modulation of the membrane composition as a potential method of altering bacterial phenotypes related to adaptation and survival.

scholarly journals Acquisition and Role of Molybdate in Pseudomonas aeruginosa

2014 ◽  
Vol 80 (21) ◽  
pp. 6843-6852 ◽  
Author(s):  
Victoria G. Pederick ◽  
Bart A. Eijkelkamp ◽  
Miranda P. Ween ◽  
Stephanie L. Begg ◽  
James C. Paton ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Fatty Acid Composition ◽  
Nitrate Reduction ◽  
Biofilm Formation ◽  
Energy Production ◽  
Anaerobic Growth ◽  
Membrane Composition ◽  
Content Type ◽  
High Affinity

ABSTRACTIn microaerophilic or anaerobic environments,Pseudomonas aeruginosautilizes nitrate reduction for energy production, a process dependent on the availability of the oxyanionic form of molybdenum, molybdate (MoO42−). Here, we show that molybdate acquisition inP. aeruginosaoccurs via a high-affinity ATP-binding cassette permease (ModABC). ModA is a cluster D-III solute binding protein capable of interacting with molybdate or tungstate oxyanions. Deletion of themodAgene reduces cellular molybdate concentrations and results in inhibition of anaerobic growth and nitrate reduction. Further, we show that conditions that permit nitrate reduction also cause inhibition of biofilm formation and an alteration in fatty acid composition ofP. aeruginosa. Collectively, these data highlight the importance of molybdate for anaerobic growth ofP. aeruginosaand reveal novel consequences of nitrate reduction on biofilm formation and cell membrane composition.

scholarly journals Chicken Juice Enhances Surface Attachment and Biofilm Formation of Campylobacter jejuni

2014 ◽  
Vol 80 (22) ◽  
pp. 7053-7060 ◽  
Author(s):  
Helen L. Brown ◽  
Mark Reuter ◽  
Louise J. Salt ◽  
Kathryn L. Cross ◽  
Roy P. Betts ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Food Chain ◽  
Biofilm Formation ◽  
Cell Attachment ◽  
Bacterial Attachment ◽  
Poultry Meat ◽  
Abiotic Surface ◽  
Surface Attachment ◽  
Aerobic Conditions ◽  
Content Type

ABSTRACTThe bacterial pathogenCampylobacter jejuniis primarily transmitted via the consumption of contaminated foodstuffs, especially poultry meat. In food processing environments,C. jejuniis required to survive a multitude of stresses and requires the use of specific survival mechanisms, such as biofilms. An initial step in biofilm formation is bacterial attachment to a surface. Here, we investigated the effects of a chicken meat exudate (chicken juice) onC. jejunisurface attachment and biofilm formation. Supplementation of brucella broth with ≥5% chicken juice resulted in increased biofilm formation on glass, polystyrene, and stainless steel surfaces with fourC. jejuniisolates and oneC. coliisolate in both microaerobic and aerobic conditions. When incubated with chicken juice,C. jejuniwas both able to grow and form biofilms in static cultures in aerobic conditions. Electron microscopy showed thatC. jejunicells were associated with chicken juice particulates attached to the abiotic surface rather than the surface itself. This suggests that chicken juice contributes toC. jejunibiofilm formation by covering and conditioning the abiotic surface and is a source of nutrients. Chicken juice was able to complement the reduction in biofilm formation of an aflagellated mutant ofC. jejuni, indicating that chicken juice may support food chain transmission of isolates with lowered motility. We provide here a useful model for studying the interaction ofC. jejunibiofilms in food chain-relevant conditions and also show a possible mechanism forC. jejunicell attachment and biofilm initiation on abiotic surfaces within the food chain.

scholarly journals Multiple Drug-Induced Stress Responses Inhibit Formation of Escherichia coli Biofilms

2020 ◽  
Vol 86 (21) ◽  
Author(s):  
Nataliya A. Teteneva ◽  
Sergey V. Mart’yanov ◽  
María Esteban-López ◽  
Jörg Kahnt ◽  
Timo Glatter ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Stress Responses ◽  
Biofilm Formation ◽  
Molecular Mechanisms ◽  
Drug Repurposing ◽  
Bacterial Attachment ◽  
Bacterial Biofilm ◽  
Multiple Drug ◽  
Content Type ◽  
Approved Drugs

ABSTRACT In most ecosystems, bacteria exist primarily as structured surface-associated biofilms that can be highly tolerant to antibiotics and thus represent an important health issue. Here, we explored drug repurposing as a strategy to identify new antibiofilm compounds, screening over 1,000 compounds from the Prestwick Chemical Library of approved drugs for specific activities that prevent biofilm formation by Escherichia coli. Most growth-inhibiting compounds, which include known antibacterial but also antiviral and other drugs, also reduced biofilm formation. However, we also identified several drugs that were biofilm inhibitory at doses where only a weak effect or no effect on planktonic growth could be observed. The activities of the most specific antibiofilm compounds were further characterized using gene expression analysis, proteomics, and microscopy. We observed that most of these drugs acted by repressing genes responsible for the production of curli, a major component of the E. coli biofilm matrix. This repression apparently occurred through the induction of several different stress responses, including DNA and cell wall damage, and homeostasis of divalent cations, demonstrating that biofilm formation can be inhibited through a variety of molecular mechanisms. One tested drug, tyloxapol, did not affect curli expression or cell growth but instead inhibited biofilm formation by suppressing bacterial attachment to the surface. IMPORTANCE The prevention of bacterial biofilm formation is one of the major current challenges in microbiology. Here, by systematically screening a large number of approved drugs for their ability to suppress biofilm formation by Escherichia coli, we identified a number of prospective antibiofilm compounds. We further demonstrated different mechanisms of action for individual compounds, from induction of replicative stress to disbalance of cation homeostasis to inhibition of bacterial attachment to the surface. Our work demonstrates the potential of drug repurposing for the prevention of bacterial biofilm formation and suggests that also for other bacteria, the activity spectrum of antibiofilm compounds is likely to be broad.

scholarly journals Disruption of L-Rhamnose Biosynthesis Results in Severe Growth Defects in Streptococcus mutans

2019 ◽  
Vol 202 (6) ◽  
Author(s):  
Andrew P. Bischer ◽  
Christopher J. Kovacs ◽  
Roberta C. Faustoferri ◽  
Robert G. Quivey
Keyword(s):  
Cell Wall ◽  
Dental Caries ◽  
Oral Cavity ◽  
Streptococcus Mutans ◽  
Biofilm Formation ◽  
Etiologic Agent ◽  
Growth Defect ◽  
Cell Wall Polysaccharide ◽  
Content Type ◽  
Severe Growth

ABSTRACT The rhamnose-glucose cell wall polysaccharide (RGP) of Streptococcus mutans plays a significant role in cell division, virulence, and stress protection. Prior studies examined function of the RGP using strains carrying deletions in the machinery involved in RGP assembly. In this study, we explored loss of the substrate for RGP, l-rhamnose, via deletion of rmlD (encoding the protein responsible for the terminal step in l-rhamnose biosynthesis). We demonstrate that loss of rhamnose biosynthesis causes a phenotype similar to strains with disrupted RGP assembly (ΔrgpG and ΔrgpF strains). Deletion of rmlD not only caused a severe growth defect under nonstress growth conditions but also elevated susceptibility of the strain to acid and oxidative stress, common conditions found in the oral cavity. A genetic complement of the ΔrmlD strain completely restored wild-type levels of growth, whereas addition of exogenous rhamnose did not. The loss of rhamnose production also significantly disrupted biofilm formation, an important aspect of S. mutans growth in the oral cavity. Further, we demonstrate that loss of either rmlD or rgpG results in ablation of rhamnose content in the S. mutans cell wall. Taken together, these results highlight the importance of rhamnose production in both the fitness and the ability of S. mutans to overcome environmental stresses. IMPORTANCE Streptococcus mutans is a pathogenic bacterium that is the primary etiologic agent of dental caries, a disease that affects billions yearly. Rhamnose biosynthesis is conserved not only in streptococcal species but in other Gram-positive, as well as Gram-negative, organisms. This study highlights the importance of rhamnose biosynthesis in RGP production for protection of the organism against acid and oxidative stresses, the two major stressors that the organism encounters in the oral cavity. Loss of RGP also severely impacts biofilm formation, the first step in the onset of dental caries. The high conservation of the rhamnose synthesis enzymes, as well as their importance in S. mutans and other organisms, makes them favorable antibiotic targets for the treatment of disease.

scholarly journals The Transcriptional Regulator BpsR Controls the Growth ofBordetella bronchisepticaby Repressing Genes Involved in Nicotinic Acid Degradation

2018 ◽  
Vol 200 (12) ◽  
pp. e00712-17 ◽  
Author(s):  
Manita Guragain ◽  
Jamie Jennings-Gee ◽  
Natalia Cattelan ◽  
Mary Finger ◽  
Matt S. Conover ◽  
...  
Keyword(s):  
Nicotinic Acid ◽  
Biofilm Formation ◽  
Transcriptional Control ◽  
Bordetella Bronchiseptica ◽  
Negative Regulator ◽  
Growth Defect ◽  
Content Type ◽  
Polysaccharide Production ◽  
Expression Of Genes

ABSTRACTMany of the pathogenic species of the genusBordetellahave an absolute requirement for nicotinic acid (NA) for laboratory growth. These Gram-negative bacteria also harbor a gene cluster homologous to theniccluster ofPseudomonas putidawhich is involved in the aerobic degradation of NA and its transcriptional control. We report here that BpsR, a negative regulator of biofilm formation and Bps polysaccharide production, controls the growth ofBordetella bronchisepticaby repressing the expression ofnicgenes. The severe growth defect of the ΔbpsRstrain in Stainer-Scholte medium was restored by supplementation with NA, which also functioned as an inducer ofnicgenes at low micromolar concentrations that are usually present in animals and humans. Purified BpsR protein bound to thenicpromoter region, and its DNA binding activity was inhibited by 6-hydroxynicotinic acid (6-HNA), the first metabolite of the NA degradative pathway. Reporter assays with the isogenic mutant derivative of the wild-type (WT) strain harboring deletion innicA, which encodes a putative nicotinic acid hydroxylase responsible for conversion of NA to 6-HNA, showed that 6-HNA is the actual inducer of thenicgenes in the bacterial cell. Gene expression profiling further showed that BpsR dually activated and repressed the expression of genes associated with pathogenesis, transcriptional regulation, metabolism, and other cellular processes. We discuss the implications of these findings with respect to the selection of pyridines such as NA and quinolinic acid for optimum bacterial growth depending on the ecological niche.IMPORTANCEBpsR, the previously described regulator of biofilm formation and Bps polysaccharide production, controlsBordetella bronchisepticagrowth by regulating the expression of genes involved in the degradation of nicotinic acid (NA). 6-Hydroxynicotinic acid (6-HNA), the first metabolite of the NA degradation pathway prevented BpsR from binding to DNA and was the actualin vivoinducer. We hypothesize that BpsR enablesBordetellabacteria to efficiently and selectively utilize NA for their survival depending on the environment in which they reside. The results reported herein lay the foundation for future investigations of how BpsR and the alteration of its activity by NA orchestrate the control ofBordetellagrowth, metabolism, biofilm formation, and pathogenesis.

scholarly journals Self-Regulation and Interplay of Rsm Family Proteins Modulate the Lifestyle of Pseudomonas putida

2016 ◽  
Vol 82 (18) ◽  
pp. 5673-5686 ◽  
Author(s):  
Óscar Huertas-Rosales ◽  
María Isabel Ramos-González ◽  
Manuel Espinosa-Urgel
Keyword(s):  
Pseudomonas Putida ◽  
Biofilm Formation ◽  
Expression Patterns ◽  
Self Regulation ◽  
Bacterial Attachment ◽  
Triple Mutant ◽  
Content Type ◽  
Global Regulators ◽  
Glass Surfaces ◽  
Plastic Surfaces

ABSTRACTIn the plant-beneficial bacteriumPseudomonas putidaKT2440, three genes have been identified that encode posttranscriptional regulators of the CsrA/RsmA family. Their regulatory roles in the motile and sessile lifestyles ofP. putidahave been investigated by generating single-, double-, and triple-null mutants and by overexpressing each protein (RsmA, RsmE, and RsmI) in different genetic backgrounds. Thersmtriple mutant shows reduced swimming and swarming motilities and increased biofilm formation, whereas overexpression of RsmE or RsmI results in reduced bacterial attachment. However, biofilms formed on glass surfaces by the triple mutant are more labile than those of the wild-type strain and are easily detached from the surface, a phenomenon that is not observed on plastic surfaces. Analysis of the expression of adhesins and exopolysaccharides in the different genetic backgrounds suggests that the biofilm phenotypes are due to alterations in the composition of the extracellular matrix and in the timing of synthesis of its elements. We have also studied the expression patterns of Rsm proteins and obtained data that indicate the existence of autoregulation mechanisms.IMPORTANCEProteins of the CsrA/RsmA family function as global regulators in different bacteria. More than one of these proteins is present in certain species. In this study, all of the RsmA homologs inP. putidaare characterized and globally taken into account to investigate their roles in controlling bacterial lifestyles and the regulatory interactions among them. The results offer new perspectives on how biofilm formation is modulated in this environmentally relevant bacterium.

scholarly journals Biofilm Formation by Psychrobacter arcticus and the Role of a Large Adhesin in Attachment to Surfaces

2013 ◽  
Vol 79 (13) ◽  
pp. 3967-3973 ◽  
Author(s):  
Shannon M. Hinsa-Leasure ◽  
Cassandra Koid ◽  
James M. Tiedje ◽  
Janna N. Schultzhaus
Keyword(s):  
Biofilm Formation ◽  
Time Course ◽  
Microtiter Plate ◽  
Bacterial Attachment ◽  
Wild Type ◽  
Microtiter Plate Assay ◽  
Content Type ◽  
Reading Frame ◽  
Transposon Mutants

ABSTRACTPsychrobacter arcticusstrain 273-4, an isolate from a Siberian permafrost core, is capable of forming biofilms when grown in minimal medium under laboratory conditions. Biofilms form at 4 to 22°C when acetate is supplied as the lone carbon source and with 1 to 7% sea salt.P. arcticusis also capable of colonizing quartz sand. Transposon mutagenesis identified a gene important for biofilm formation byP. arcticus. Four transposon mutants were mapped to a 20.1-kbp gene, which is predicted to encode a protein of 6,715 amino acids (Psyc_1601). We refer to this open reading frame ascat1, for cold attachment gene 1. Thecat1mutants are unable to form biofilms at levels equivalent to that of the wild type, and there is no impact on the planktonic growth characteristics of the strains, indicating a specific role in biofilm formation. Through time course studies of the static microtiter plate assay, we determined thatcat1mutants are unable to form biofilms equivalent to that of the wild type under all conditions tested. In flow cell experiments,cat1mutants initially are unable to attach to the surface. Over time, however, they form microcolonies, an architecture very different from that produced by wild-type biofilms. Our results demonstrate that Cat1 is involved in the initial stages of bacterial attachment to surfaces.

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