Applying insights from biofilm biology to drug development — can a new approach be developed?

Most of the research on bacterial pathogenesis has focused on acute infections, but much less is known about the pathogenesis of infections caused by bacteria that grow as aggregates in biofilms. These infections tend to be chronic as they resist innate and adaptive immune defence mechanisms as well as antibiotics, and the treatment of biofilm infections presents a considerable unmet clinical need. To date, there are no drugs that specifically target bacteria in biofilms; however, several approaches are in early-stage development. Here, we review current insights into biofilm physiology and pathology, and discuss how a deep insight into the physical and biological characteristics of biofilms can inform therapeutic strategies and molecular targets for the development of anti-biofilm drugs.

[1]  Thomas Bjarnsholt,et al.  Garlic blocks quorum sensing and promotes rapid clearing of pulmonary Pseudomonas aeruginosa infections. , 2005, Microbiology.

[2]  M. Hamilton,et al.  Measuring antimicrobial effects on biofilm bacteria: from laboratory to field. , 1999, Methods in enzymology.

[3]  S. Ankri,et al.  Antimicrobial properties of allicin from garlic. , 1999, Microbes and infection.

[4]  L. Rasmussen,et al.  Infection Risk with Nitrofurazone-Impregnated Urinary Catheters in Trauma Patients , 2007, Annals of Internal Medicine.

[5]  Thomas Bjarnsholt,et al.  Biofilms in chronic infections - a matter of opportunity - monospecies biofilms in multispecies infections. , 2010, FEMS immunology and medical microbiology.

[6]  Kathrine B. Christensen,et al.  Identity and effects of quorum-sensing inhibitors produced by Penicillium species. , 2005, Microbiology.

[7]  D. Pompliano,et al.  Drugs for bad bugs: confronting the challenges of antibacterial discovery , 2007, Nature Reviews Drug Discovery.

[8]  N. Bagge,et al.  Antibodies against β‐lactamase can improve ceftazidime treatment of lung infection with β‐lactam‐resistant Pseudomonas aeruginosa in a rat model of chronic lung infection , 2002, APMIS : acta pathologica, microbiologica, et immunologica Scandinavica.

[9]  Angela Lee,et al.  MexXY-OprM Efflux Pump Is Necessary for Adaptive Resistance of Pseudomonas aeruginosa to Aminoglycosides , 2003, Antimicrobial Agents and Chemotherapy.

[10]  J. Costerton,et al.  Scanning and transmission electron microscopy of in situ bacterial colonization of intravenous and intraarterial catheters , 1984, Journal of clinical microbiology.

[11]  B. Christensen,et al.  Molecular tools for study of biofilm physiology. , 1999, Methods in enzymology.

[12]  S. de Bentzmann,et al.  Hacking into bacterial biofilms: a new therapeutic challenge , 2011, Annals of intensive care.

[13]  D. Tifrea,et al.  A chemosensory system that regulates biofilm formation through modulation of cyclic diguanylate levels. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[14]  D. Hassett,et al.  Anaerobic metabolism and quorum sensing by Pseudomonas aeruginosa biofilms in chronically infected cystic fibrosis airways: rethinking antibiotic treatment strategies and drug targets. , 2002, Advanced drug delivery reviews.

[15]  J. Hahn,et al.  Tolerance of dormant and active cells in Pseudomonas aeruginosa PA01 biofilm to antimicrobial agents. , 2009, The Journal of antimicrobial chemotherapy.

[16]  Y. Hirakata,et al.  Azithromycin Exhibits Bactericidal Effects on Pseudomonas aeruginosa through Interaction with the Outer Membrane , 2005, Antimicrobial Agents and Chemotherapy.

[17]  C. Sternberg,et al.  An in vitro model of bacterial infections in wounds and other soft tissues , 2010, APMIS : acta pathologica, microbiologica, et immunologica Scandinavica.

[18]  C. Meisner,et al.  A double-blind randomized placebo-controlled phase III study of a Pseudomonas aeruginosa flagella vaccine in cystic fibrosis patients , 2007, Proceedings of the National Academy of Sciences.

[19]  N. Høiby,et al.  Pseudomonas aeruginosa biofilms in the respiratory tract of cystic fibrosis patients , 2009, Pediatric pulmonology.

[20]  P. Stewart,et al.  Gene expression and protein levels of the stationary phase sigma factor, RpoS, in continuously-fed Pseudomonas aeruginosa biofilms. , 2001, FEMS microbiology letters.

[21]  D. Hassett,et al.  Anaerobic killing of mucoid Pseudomonas aeruginosa by acidified nitrite derivatives under cystic fibrosis airway conditions. , 2006, The Journal of clinical investigation.

[22]  R. Darouiche,et al.  New strategies to prevent catheter-associated urinary tract infections , 2012, Nature Reviews Urology.

[23]  Thomas Bjarnsholt,et al.  Quorum Sensing Antagonism from Marine Organisms , 2007, Marine Biotechnology.

[24]  S. Lory,et al.  Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic pathogen , 2000, Nature.

[25]  A. Ojha,et al.  Targeting drug tolerance in mycobacteria: a perspective from mycobacterial biofilms , 2012, Expert review of anti-infective therapy.

[26]  H. Kristensen,et al.  Effects of Intratracheal Administration of Novispirin G10 on a Rat Model of Mucoid Pseudomonas aeruginosa Lung Infection , 2005, Antimicrobial Agents and Chemotherapy.

[27]  J. Costerton,et al.  Microbial Biofilms , 2011 .

[28]  J. Costerton,et al.  Establishment of aging biofilms: possible mechanism of bacterial resistance to antimicrobial therapy , 1992, Antimicrobial Agents and Chemotherapy.

[29]  C. van Delden,et al.  Swarming of Pseudomonas aeruginosa Is Dependent on Cell-to-Cell Signaling and Requires Flagella and Pili , 2000, Journal of bacteriology.

[30]  G. Jackson,et al.  Pathogenicity of staphylococci; a comparison of alpha-hemolysin production with the coagulase test and clinical observations of virulence. , 1955, The New England journal of medicine.

[31]  D. Maki,et al.  Are antimicrobial-impregnated catheters effective? When does repetition reach the point of exhaustion? , 2005, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[32]  Roberto Kolter,et al.  Initiation of biofilm formation in Pseudomonas fluorescens WCS365 proceeds via multiple, convergent signalling pathways: a genetic analysis , 1998, Molecular microbiology.

[33]  Federica Villa,et al.  Effects of Photoactivated Titanium Dioxide Nanopowders and Coating on Planktonic and Biofilm Growth of Pseudomonas aeruginosa , 2011, Photochemistry and Photobiology.

[34]  S. Molin,et al.  Biofilm formation by Pseudomonas aeruginosa wild type, flagella and type IV pili mutants , 2003, Molecular microbiology.

[35]  Peilin Wu,et al.  Preliminary Observations , 1830, The Medico-chirurgical review.

[36]  J. Carlet,et al.  Quorum sensing-dependent virulence during Pseudomonas aeruginosa colonisation and pneumonia in mechanically ventilated patients , 2010, Thorax.

[37]  J. Costerton,et al.  Bacterial biofilms: a common cause of persistent infections. , 1999, Science.

[38]  S. M. Kirov,et al.  Iron chelation directed against biofilms as an adjunct to conventional antibiotics. , 2009, American journal of physiology. Lung cellular and molecular physiology.

[39]  O. Ciofu,et al.  Development of antibiotic resistance in Pseudomonas aeruginosa during two decades of antipseudomonal treatment at the Danish CF Center , 1994, APMIS : acta pathologica, microbiologica, et immunologica Scandinavica.

[40]  A. Larsson,et al.  Good effect of IgY against Pseudomonas aeruginosa infections in cystic fibrosis patients , 2008, Pediatric pulmonology.

[41]  L. Eberl,et al.  Screening for Quorum-Sensing Inhibitors (QSI) by Use of a Novel Genetic System, the QSI Selector , 2005, Journal of bacteriology.

[42]  E. Greenberg,et al.  Influence of Quorum Sensing and Iron on Twitching Motility and Biofilm Formation in Pseudomonas aeruginosa , 2007, Journal of bacteriology.

[43]  S. Molin,et al.  In Situ Growth Rates and Biofilm Development of Pseudomonas aeruginosa Populations in Chronic Lung Infections , 2007, Journal of bacteriology.

[44]  J. Collins,et al.  Heterogeneous Bacterial Persisters and Engineering Approaches to Eliminate Them This Review Comes from a Themed Issue on Genomics Edited More than One Way to Make a Persister Persisters and Physiological Heterogeneity Engineering Treatments for Persisters , 2022 .

[45]  Forest Rohwer,et al.  Metagenomic Analysis of Respiratory Tract DNA Viral Communities in Cystic Fibrosis and Non-Cystic Fibrosis Individuals , 2009, PloS one.

[46]  A. Grant,et al.  Antimicrobial catheters for reduction of symptomatic urinary tract infection in adults requiring short-term catheterisation in hospital: a multicentre randomised controlled trial , 2012, The Lancet.

[47]  S. Kjelleberg,et al.  Eukaryotic interference with homoserine lactone-mediated prokaryotic signalling , 1996, Journal of bacteriology.

[48]  M. Enright,et al.  The role of regulated clinical trials in the development of bacteriophage therapeutics , 2012, Journal of molecular and genetic medicine : an international journal of biomedical research.

[49]  A. Birch‐Andersen,et al.  Morphology Of Pseudomonas aeruginosa phages from the sputum of cystic fibrosis patients and from the phage typing set , 1991, APMIS : acta pathologica, microbiologica, et immunologica Scandinavica.

[50]  Howard C. Berg,et al.  Genetic analysis , 1957, Nature Biotechnology.

[51]  Sanjay Saint,et al.  Guidelines for the prevention of intravascular catheter-related infections. , 2011, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[52]  H. Whitfield,et al.  Prophylactic efficacy of a new gentamicin‐releasing urethral catheter in short‐term catheterized rabbits , 2001, BJU International.

[53]  S. Molin,et al.  Phenotypes of Non-Attached Pseudomonas aeruginosa Aggregates Resemble Surface Attached Biofilm , 2011, PloS one.

[54]  J. Costerton,et al.  Bacterial biofilms in nature and disease. , 1987, Annual review of microbiology.

[55]  Hans-Curt Flemming,et al.  The EPS Matrix: The “House of Biofilm Cells” , 2007, Journal of bacteriology.

[56]  Efficacy of the lithotripsy in treating lower pole renal stones , 2013, Urolithiasis.

[57]  Martin Schuster,et al.  Pseudomonas aeruginosa Biofilms Exposed to Imipenem Exhibit Changes in Global Gene Expression and β-Lactamase and Alginate Production , 2004, Antimicrobial Agents and Chemotherapy.

[58]  H. Brüssow Pseudomonas Biofilms, Cystic Fibrosis, and Phage: a Silver Lining? , 2012, mBio.

[59]  P. Williams,et al.  The Pseudomonas aeruginosa sensor RetS switches type III and type VI secretion via c-di-GMP signalling. , 2011, Environmental microbiology.

[60]  P. Shaw,et al.  Role of hydrochloric acid in the treatment of central venous catheter infections in children with cancer , 2004, Cancer.

[61]  G. Pugliese,et al.  Biofilms: Survival Mechanisms of Clinically Relevant Microorganisms , 2002, Infection Control & Hospital Epidemiology.

[62]  P. Stewart,et al.  Arginine or Nitrate Enhances Antibiotic Susceptibility of Pseudomonas aeruginosa in Biofilms , 2006, Antimicrobial Agents and Chemotherapy.

[63]  E. Greenberg,et al.  Iron and Pseudomonas aeruginosa biofilm formation. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[64]  Søren Molin,et al.  Involvement of bacterial migration in the development of complex multicellular structures in Pseudomonas aeruginosa biofilms , 2003, Molecular microbiology.

[65]  J. Costerton,et al.  Antibiotic resistance of bacteria in biofilms , 2001, The Lancet.

[66]  P. Gilligan,et al.  Microbiology of airway disease in patients with cystic fibrosis , 1991, Clinical Microbiology Reviews.

[67]  T. Tolker-Nielsen,et al.  Proteins with GGDEF and EAL domains regulate Pseudomonas putida biofilm formation and dispersal. , 2006, FEMS microbiology letters.

[68]  A. Beauvais,et al.  Aspergillus fumigatus biofilms in the clinical setting. , 2011, Medical mycology.

[69]  K. Poole Bacterial stress responses as determinants of antimicrobial resistance. , 2012, The Journal of antimicrobial chemotherapy.

[70]  M. Givskov,et al.  The immune system vs. Pseudomonas aeruginosa biofilms. , 2010, FEMS immunology and medical microbiology.

[71]  L. Herzenberg,et al.  High-dose oral N-acetylcysteine, a glutathione prodrug, modulates inflammation in cystic fibrosis. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[72]  S. Sørensen,et al.  The interconnection between biofilm formation and horizontal gene transfer. , 2012, FEMS immunology and medical microbiology.

[73]  T. Coenye,et al.  Synergistic antibacterial efficacy of early combination treatment with tobramycin and quorum-sensing inhibitors against Pseudomonas aeruginosa in an intraperitoneal foreign-body infection mouse model , 2012 .

[74]  J. Martínez The role of natural environments in the evolution of resistance traits in pathogenic bacteria , 2009, Proceedings of the Royal Society B: Biological Sciences.

[75]  J. Faoagali,et al.  Ethanol lock therapy to treat tunnelled central venous catheter-associated blood stream infections: Results from a prospective trial , 2008, Scandinavian journal of infectious diseases.

[76]  K. M. Lee,et al.  Identification of genes controlled by quorum sensing in Pseudomonas aeruginosa. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[77]  S. Assinder,et al.  Mathematical model of β-lactam penetration into a biofilm of Pseudomonas aeruginosa while undergoing simultaneous inactivation by released β-lactamases , 1996 .

[78]  S. Kjelleberg,et al.  Cell Death in Pseudomonas aeruginosa Biofilm Development , 2003, Journal of bacteriology.

[79]  S. Lewenza,et al.  Extracellular DNA Chelates Cations and Induces Antibiotic Resistance in Pseudomonas aeruginosa Biofilms , 2008, PLoS pathogens.

[80]  S. Molin,et al.  The cep quorum-sensing system of Burkholderia cepacia H111 controls biofilm formation and swarming motility. , 2001, Microbiology.

[81]  K. Mathee,et al.  Panax ginseng has anti-infective activity against opportunistic pathogen Pseudomonas aeruginosa by inhibiting quorum sensing, a bacterial communication process critical for establishing infection. , 2010, Phytomedicine : international journal of phytotherapy and phytopharmacology.

[82]  P. Stewart,et al.  A genetic basis for Pseudomonas aeruginosa biofilm antibiotic resistance , 2003, Nature.

[83]  Hsin-Chieh Yeh,et al.  Effect of the 2011 vs 2003 duty hour regulation-compliant models on sleep duration, trainee education, and continuity of patient care among internal medicine house staff: a randomized trial. , 2013, JAMA internal medicine.

[84]  G. Weinstock,et al.  Effects of Enterococcus faecalis fsrGenes on Production of Gelatinase and a Serine Protease and Virulence , 2000, Infection and Immunity.

[85]  N. Raffaelli,et al.  Monitoring of diguanylate cyclase activity and of cyclic-di-GMP biosynthesis by whole-cell assays suitable for high-throughput screening of biofilm inhibitors , 2009, Applied Microbiology and Biotechnology.

[86]  Frank Cerny From laboratory to the “field” , 1996, Pediatric pulmonology.

[87]  Krasimir Vasilev,et al.  Antibacterial surfaces for biomedical devices , 2009, Expert review of medical devices.

[88]  D. Touw,et al.  Inhaled medication and inhalation devices for lung disease in patients with cystic fibrosis: A European consensus. , 2009, Journal of cystic fibrosis : official journal of the European Cystic Fibrosis Society.

[89]  P. Stewart New ways to stop biofilm infections , 2003, The Lancet.

[90]  P. Stewart,et al.  Theoretical aspects of antibiotic diffusion into microbial biofilms , 1996, Antimicrobial agents and chemotherapy.

[91]  J. Collins,et al.  A Common Mechanism of Cellular Death Induced by Bactericidal Antibiotics , 2007, Cell.

[92]  D. Davies,et al.  A Fatty Acid Messenger Is Responsible for Inducing Dispersion in Microbial Biofilms , 2008, Journal of bacteriology.

[93]  A. Weinberg,et al.  Antimicrobial activity of gentamicin palmitate against high concentrations of Staphylococcus aureus , 2011, Journal of materials science. Materials in medicine.

[94]  N. Schiller,et al.  Alginate lyase enhances antibiotic killing of mucoid Pseudomonas aeruginosa in biofilms , 2006, APMIS : acta pathologica, microbiologica, et immunologica Scandinavica.

[95]  S. Dye,et al.  An evolutionary perspective of the knee. , 1987, The Journal of bone and joint surgery. American volume.

[96]  Valeria Souza,et al.  Stress-Induced Mutagenesis in Bacteria , 2003, Science.

[97]  S. Molin,et al.  Heterogeneity of Biofilms Formed by Nonmucoid Pseudomonas aeruginosa Isolates from Patients with Cystic Fibrosis , 2005, Journal of Clinical Microbiology.

[98]  Chengping Lu,et al.  Application of a Bacteriophage Lysin To Disrupt Biofilms Formed by the Animal Pathogen Streptococcus suis , 2011, Applied and Environmental Microbiology.

[99]  H. Kristensen,et al.  Plectasin Shows Intracellular Activity against Staphylococcus aureus in Human THP-1 Monocytes and in a Mouse Peritonitis Model , 2009, Antimicrobial Agents and Chemotherapy.

[100]  G. Volckaert,et al.  Quality-Controlled Small-Scale Production of a Well-Defined Bacteriophage Cocktail for Use in Human Clinical Trials , 2009, PloS one.

[101]  M. Jadoul,et al.  Preventing haemodialysis catheter-related bacteraemia with an antimicrobial lock solution: a meta-analysis of prospective randomized trials. , 2008, Nephrology, dialysis, transplantation : official publication of the European Dialysis and Transplant Association - European Renal Association.

[102]  Michael J. MacCoss,et al.  Aminoglycoside antibiotics induce bacterial biofilm formation , 2005, Nature.

[103]  Qun Ma,et al.  Engineering a novel c-di-GMP-binding protein for biofilm dispersal. , 2011, Environmental microbiology.

[104]  P Stoodley,et al.  Survival strategies of infectious biofilms. , 2005, Trends in microbiology.

[105]  I. Chopra,et al.  Increased mutability of Pseudomonas aeruginosa in biofilms. , 2008, The Journal of antimicrobial chemotherapy.

[106]  B. Iglewski,et al.  Azithromycin Retards Pseudomonas aeruginosa Biofilm Formation , 2004, Journal of Clinical Microbiology.

[107]  P. Williams,et al.  Controlling infection by tuning in and turning down the volume of bacterial small-talk. , 2002, The Lancet. Infectious diseases.

[108]  T. Tolker-Nielsen,et al.  The metabolically active subpopulation in Pseudomonas aeruginosa biofilms survives exposure to membrane-targeting antimicrobials via distinct molecular mechanisms. , 2012, FEMS immunology and medical microbiology.

[109]  Richard C Boucher,et al.  Effects of reduced mucus oxygen concentration in airway Pseudomonas infections of cystic fibrosis patients. , 2002, The Journal of clinical investigation.

[110]  N. Høiby,et al.  Avidity of anti-P aeruginosaantibodies during chronic infection in patients with cystic fibrosis , 1999, Thorax.

[111]  L. J. Douglas,et al.  Effects of Aspirin and Other Nonsteroidal Anti-Inflammatory Drugs on Biofilms and Planktonic Cells of Candida albicans , 2004, Antimicrobial Agents and Chemotherapy.

[112]  I. Chopra,et al.  Targeting bacterial membrane function: an underexploited mechanism for treating persistent infections , 2010, Nature Reviews Microbiology.

[113]  C. Desnues,et al.  Bacteriophages as vehicles of the resistome in cystic fibrosis. , 2011, The Journal of antimicrobial chemotherapy.

[114]  N. Høiby,et al.  Interactions between Polymorphonuclear Leukocytes and Pseudomonas aeruginosa Biofilms on Silicone Implants In Vivo , 2012, Infection and Immunity.

[115]  T. Mah,et al.  Involvement of a Novel Efflux System in Biofilm-Specific Resistance to Antibiotics , 2008, Journal of bacteriology.

[116]  D. Kadouri,et al.  Growing and Analyzing Static Biofilms , 2006, Current protocols in microbiology.

[117]  N. Chauhan,et al.  Sensitization of Candida albicans biofilms to various antifungal drugs by cyclosporine A , 2012, Annals of Clinical Microbiology and Antimicrobials.

[118]  Weiling Fu,et al.  Bacteriophage Cocktail for the Prevention of Biofilm Formation by Pseudomonas aeruginosa on Catheters in an In Vitro Model System , 2009, Antimicrobial Agents and Chemotherapy.

[119]  G. Pier,et al.  Inactivation of the rhlA gene in Pseudomonas aeruginosa prevents rhamnolipid production, disabling the protection against polymorphonuclear leukocytes , 2009, APMIS : acta pathologica, microbiologica, et immunologica Scandinavica.

[120]  Mirian Domenech,et al.  In Vitro Destruction of Streptococcus pneumoniae Biofilms with Bacterial and Phage Peptidoglycan Hydrolases , 2011, Antimicrobial Agents and Chemotherapy.

[121]  M. Shirtliff,et al.  Resolution of Staphylococcus aureus Biofilm Infection Using Vaccination and Antibiotic Treatment , 2011, Infection and Immunity.

[122]  G. Anderson,et al.  Targeting iron uptake to control Pseudomonas aeruginosa infections in cystic fibrosis , 2012, European Respiratory Journal.

[123]  M. Maciá,et al.  Dynamics of Mutator and Antibiotic-Resistant Populations in a Pharmacokinetic/Pharmacodynamic Model of Pseudomonas aeruginosa Biofilm Treatment , 2011, Antimicrobial Agents and Chemotherapy.

[124]  A. Filloux,et al.  Key two-component regulatory systems that control biofilm formation in Pseudomonas aeruginosa. , 2011, Environmental microbiology.

[125]  Leo Eberl,et al.  Inhibition of quorum sensing in Pseudomonas aeruginosa biofilm bacteria by a halogenated furanone compound. , 2002, Microbiology.

[126]  Kwang-tae Choi,et al.  Botanical characteristics, pharmacological effects and medicinal components of Korean Panax ginseng C A Meyer , 2008, Acta Pharmacologica Sinica.

[127]  Philip S. Stewart,et al.  Contributions of Antibiotic Penetration, Oxygen Limitation, and Low Metabolic Activity to Tolerance of Pseudomonas aeruginosa Biofilms to Ciprofloxacin and Tobramycin , 2003, Antimicrobial Agents and Chemotherapy.

[128]  Garth D Ehrlich,et al.  Physiology of Pseudomonas aeruginosa in biofilms as revealed by transcriptome analysis , 2010, BMC Microbiology.

[129]  N. Høiby,et al.  Pharmacokinetics/Pharmacodynamics of Colistin and Imipenem on Mucoid and Nonmucoid Pseudomonas aeruginosa Biofilms , 2011, Antimicrobial Agents and Chemotherapy.

[130]  R. Morton,et al.  Therapeutic ultrasound as treatment for chronic rhinosinusitis: preliminary observations. , 2010, The Journal of laryngology and otology.

[131]  P. Williams,et al.  Rapid necrotic killing of polymorphonuclear leukocytes is caused by quorum-sensing-controlled production of rhamnolipid by Pseudomonas aeruginosa. , 2007, Microbiology.

[132]  J. Vleugels,et al.  The nonsteroidal antiinflammatory drug diclofenac potentiates the in vivo activity of caspofungin against Candida albicans biofilms. , 2012, The Journal of infectious diseases.

[133]  K. Sauer,et al.  SagS Contributes to the Motile-Sessile Switch and Acts in Concert with BfiSR To Enable Pseudomonas aeruginosa Biofilm Formation , 2011, Journal of bacteriology.

[134]  D. Maki,et al.  Use of vancomycin-containing lock or flush solutions for prevention of bloodstream infection associated with central venous access devices: a meta-analysis of prospective, randomized trials. , 2006, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[135]  T. Bjarnsholt,et al.  A UVC Device for Intra‐luminal Disinfection of Catheters: In Vitro Tests on Soft Polymer Tubes Contaminated with Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli and Candida albicans , 2011, Photochemistry and photobiology.

[136]  D. McDougald,et al.  Pseudomonas aeruginosa PAO1 Preferentially Grows as Aggregates in Liquid Batch Cultures and Disperses upon Starvation , 2009, PloS one.

[137]  Blaise R. Boles,et al.  Endogenous oxidative stress produces diversity and adaptability in biofilm communities , 2008, Proceedings of the National Academy of Sciences.

[138]  Viktoria Hancock,et al.  Inactivation of Efflux Pumps Abolishes Bacterial Biofilm Formation , 2008, Applied and Environmental Microbiology.

[139]  R. Kobza,et al.  Bacterial Colonization and Infection of Electrophysiological Cardiac Devices Detected With Sonication and Swab Culture , 2010, Circulation.

[140]  L. Zeef,et al.  Characterization of Nutrient-Induced Dispersion in Pseudomonas aeruginosa PAO1 Biofilm , 2004, Journal of bacteriology.

[141]  P. Stewart,et al.  Role of Antibiotic Penetration Limitation in Klebsiella pneumoniae Biofilm Resistance to Ampicillin and Ciprofloxacin , 2000, Antimicrobial Agents and Chemotherapy.

[142]  Lindsey A. Lorenz,et al.  A method for growing a biofilm under low shear at the air–liquid interface using the drip flow biofilm reactor , 2009, Nature Protocols.

[143]  Anders Folkesson,et al.  Adaptation of Pseudomonas aeruginosa to the cystic fibrosis airway: an evolutionary perspective , 2012, Nature Reviews Microbiology.

[144]  N. Høiby,et al.  Occurrence of Hypermutable Pseudomonas aeruginosa in Cystic Fibrosis Patients Is Associated with the Oxidative Stress Caused by Chronic Lung Inflammation , 2005, Antimicrobial Agents and Chemotherapy.

[145]  C. von Buchwald,et al.  Decreased mucosal oxygen tension in the maxillary sinuses in patients with cystic fibrosis. , 2011, Journal of cystic fibrosis : official journal of the European Cystic Fibrosis Society.

[146]  F. Malouin,et al.  Iron transport-mediated drug delivery using mixed-ligand siderophore-beta-lactam conjugates. , 1996, Chemistry & biology.

[147]  M. Parsek,et al.  Bacterial biofilms: an emerging link to disease pathogenesis. , 2003, Annual review of microbiology.

[148]  T. Wilt,et al.  Systematic Review: Antimicrobial Urinary Catheters To Prevent Catheter-Associated Urinary Tract Infection in Hospitalized Patients , 2006, Annals of Internal Medicine.

[149]  B. Marquez Bacterial efflux systems and efflux pumps inhibitors. , 2005, Biochimie.

[150]  Philip S. Stewart,et al.  Stratified Growth in Pseudomonas aeruginosa Biofilms , 2004, Applied and Environmental Microbiology.

[151]  N. Høiby,et al.  PSEUDOMONAS AERUGINOSA INFECTION IN CYSTIC FIBROSIS , 2009 .

[152]  Thomas Bjarnsholt,et al.  Quorum sensing inhibitory drugs as next generation antimicrobials: Worth the effort? , 2008, Current infectious disease reports.

[153]  J. Costerton,et al.  Observations of fouling biofilm formation. , 1981, Canadian journal of microbiology.

[154]  I. Sutherland,et al.  Biofilm susceptibility to bacteriophage attack: the role of phage-borne polysaccharide depolymerase. , 1998, Microbiology.

[155]  A. Gristina,et al.  Biomaterial-centered infection: microbial adhesion versus tissue integration. , 1987, Science.

[156]  C. Potera Studying slime. , 1998, Environmental health perspectives.

[157]  J. Alder,et al.  Bactericidal Action of Daptomycin against Stationary-Phase and Nondividing Staphylococcus aureus Cells , 2007, Antimicrobial Agents and Chemotherapy.

[158]  L. Eberl,et al.  Transcriptome analysis of Pseudomonas aeruginosa biofilm development: anaerobic respiration and iron limitation , 2005 .

[159]  H. Budzikiewicz Siderophore-antibiotic conjugates used as trojan horses against Pseudomonas aeruginosa. , 2001, Current topics in medicinal chemistry.

[160]  K. Krogfelt,et al.  Why chronic wounds will not heal: a novel hypothesis , 2008, Wound repair and regeneration : official publication of the Wound Healing Society [and] the European Tissue Repair Society.

[161]  A. Gristina Biomaterial-centered infection: microbial adhesion versus tissue integration. 1987. , 2004, Clinical orthopaedics and related research.

[162]  M. Hentzer,et al.  Azithromycin Blocks Quorum Sensing and Alginate Polymer Formation and Increases the Sensitivity to Serum and Stationary-Growth-Phase Killing of Pseudomonas aeruginosa and Attenuates Chronic P. aeruginosa Lung Infection in Cftr−/− Mice , 2007, Antimicrobial Agents and Chemotherapy.

[163]  N. Høiby,et al.  Effect of aerosolized rhDNase (Pulmozyme®) on pulmonary colonization in patients with cystic fibrosis , 2006, Acta paediatrica.

[164]  O. Ciofu Pseudomonas aeruginosa chromosomal beta-lactamase in patients with cystic fibrosis and chronic lung infection. Mechanism of antibiotic resistance and target of the humoral immune response. , 2003, APMIS. Supplementum.

[165]  S. Molin,et al.  Colistin-tobramycin combinations are superior to monotherapy concerning the killing of biofilm Pseudomonas aeruginosa. , 2010, The Journal of infectious diseases.

[166]  K. Marchal,et al.  The small regulatory RNA molecule MicA is involved in Salmonella enterica serovar Typhimurium biofilm formation , 2010, BMC Microbiology.

[167]  F. D. De Rosa,et al.  Early experience with high-dosage daptomycin for prosthetic infections. , 2009, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[168]  P. Stewart,et al.  Comparison of recalcitrance to ciprofloxacin and levofloxacin exhibited by Pseudomonas aeruginosa bofilms displaying rapid-transport characteristics , 1997, Antimicrobial agents and chemotherapy.

[169]  T. B. Rasmussen,et al.  Quorum-sensing inhibitors as anti-pathogenic drugs. , 2006, International journal of medical microbiology : IJMM.

[170]  R. Darouiche,et al.  Treatment of infections associated with surgical implants. , 2004, The New England journal of medicine.

[171]  N. Høiby,et al.  Antibodies against Pseudomonas aeruginosa chromosomal β-lactamase in patients with cystic fibrosis are markers of the development of resistance of P. aeruginosa to β-lactams , 1995 .

[172]  Chih-kuan Tung,et al.  Acceleration of Emergence of Bacterial Antibiotic Resistance in Connected Microenvironments , 2011, Science.

[173]  K. Lewis,et al.  Persister cells. , 2010, Annual review of microbiology.

[174]  S. Molin,et al.  Synergistic Activities of an Efflux Pump Inhibitor and Iron Chelators against Pseudomonas aeruginosa Growth and Biofilm Formation , 2010, Antimicrobial Agents and Chemotherapy.

[175]  Garth D. Ehrlich,et al.  Oxygen Limitation Contributes to Antibiotic Tolerance of Pseudomonas aeruginosa in Biofilms , 2004, Antimicrobial Agents and Chemotherapy.

[176]  S. Kjelleberg,et al.  Attenuation of Pseudomonas aeruginosa virulence by quorum sensing inhibitors , 2003, The EMBO journal.

[177]  M. Cuéllar-Cruz,et al.  Candida species: new insights into biofilm formation. , 2012, Future microbiology.

[178]  R. Darouiche,et al.  Antibacterial activity of antibiotic coated silicone grafts. , 1998, The Journal of urology.

[179]  L. Eberl,et al.  Ajoene, a Sulfur-Rich Molecule from Garlic, Inhibits Genes Controlled by Quorum Sensing , 2012, Antimicrobial Agents and Chemotherapy.

[180]  A. Grossman,et al.  Biochemical and genetic characterization of a competence pheromone from B. subtilis , 1994, Cell.

[181]  M. Parsek,et al.  Pseudomonas aeruginosa recognizes and responds aggressively to the presence of polymorphonuclear leukocytes. , 2009, Microbiology.

[182]  Leo Eberl,et al.  Surface Motility of Serratia liquefaciens MG1 , 1999, Journal of bacteriology.

[183]  Paul Stoodley,et al.  Bacterial biofilms: from the Natural environment to infectious diseases , 2004, Nature Reviews Microbiology.

[184]  J W Costerton,et al.  How bacteria stick. , 1978, Scientific American.

[185]  M. Hentzer,et al.  Dynamics and Spatial Distribution of β-Lactamase Expression in Pseudomonas aeruginosa Biofilms , 2004, Antimicrobial Agents and Chemotherapy.

[186]  H. Ceri,et al.  The Calgary Biofilm Device: New Technology for Rapid Determination of Antibiotic Susceptibilities of Bacterial Biofilms , 1999, Journal of Clinical Microbiology.

[187]  A. Chakraborty Electron Microscopy Study , 2014 .

[188]  T. Tolker-Nielsen,et al.  Insight into the microbial multicellular lifestyle via flow‐cell technology and confocal microscopy , 2009, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[189]  Bonnie L. Bassler,et al.  Quorum-sensing regulators control virulence gene expression in Vibrio cholerae , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[190]  Frederick M. Ausubel,et al.  Pseudomonas biofilm formation and antibiotic resistance are linked to phenotypic variation , 2002, Nature.

[191]  S. Molin,et al.  Synthetic furanones inhibit quorum-sensing and enhance bacterial clearance in Pseudomonas aeruginosa lung infection in mice. , 2004, The Journal of antimicrobial chemotherapy.

[192]  Blaise R. Boles,et al.  agr-Mediated Dispersal of Staphylococcus aureus Biofilms , 2008, PLoS pathogens.

[193]  Bonnie L Bassler,et al.  Chemical communication among bacteria , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[194]  L. Fernández,et al.  Adaptive and Mutational Resistance: Role of Porins and Efflux Pumps in Drug Resistance , 2013, Clinical Microbiology Reviews.

[195]  T. Tolker-Nielsen,et al.  The implication of Pseudomonas aeruginosa biofilms in infections. , 2011, Inflammation & allergy drug targets.

[196]  Stefan Wuertz,et al.  High Rates of Conjugation in Bacterial Biofilms as Determined by Quantitative In Situ Analysis , 1999, Applied and Environmental Microbiology.

[197]  S. Molin,et al.  Mucoid Pseudomonas aeruginosa isolates maintain the biofilm formation capacity and the gene expression profiles during the chronic lung infection of CF patients , 2011, APMIS : acta pathologica, microbiologica, et immunologica Scandinavica.

[198]  R. Samudrala,et al.  Pseudomonas aeruginosa uses a cyclic-di-GMP-regulated adhesin to reinforce the biofilm extracellular matrix , 2010, Molecular microbiology.

[199]  D. Haas,et al.  The ArgR Regulatory Protein, a Helper to the Anaerobic Regulator ANR during Transcriptional Activation of thearcD Promoter in Pseudomonas aeruginosa , 1999, Journal of bacteriology.

[200]  M. Leunig,et al.  Improved Diagnosis of Periprosthetic Joint Infection by Multiplex PCR of Sonication Fluid from Removed Implants , 2010, Journal of Clinical Microbiology.

[201]  A. Filloux,et al.  Two-component regulatory systems in Pseudomonas aeruginosa: an intricate network mediating fimbrial and efflux pump gene expression , 2011, Molecular microbiology.

[202]  N. Høiby,et al.  Genetic adaptation of Pseudomonas aeruginosa during chronic lung infection of patients with cystic fibrosis: strong and weak mutators with heterogeneous genetic backgrounds emerge in mucA and/or lasR mutants. , 2010, Microbiology.

[203]  A. Dominguez,et al.  The role of diclofenac sodium in the dimorphic transition in Candida albicans. , 2010, Microbial pathogenesis.

[204]  A. Gristina Microbial Adhesion versus Tissue Integration , 2004 .

[205]  J. Oliver,et al.  Recent findings on the viable but nonculturable state in pathogenic bacteria. , 2010, FEMS microbiology reviews.

[206]  L. Touqui,et al.  Pulmonary Bacteriophage Therapy on Pseudomonas aeruginosa Cystic Fibrosis Strains: First Steps Towards Treatment and Prevention , 2011, PloS one.

[207]  M. Hentzer,et al.  Pharmacological inhibition of quorum sensing for the treatment of chronic bacterial infections. , 2003, The Journal of clinical investigation.

[208]  Christophe Pichon,et al.  Small RNA gene identification and mRNA target predictions in bacteria , 2008, Bioinform..

[209]  H. Buchholz,et al.  [Experimental and clinical studies on the release of gentamicin from bone cement]. , 1972, Der Chirurg; Zeitschrift fur alle Gebiete der operativen Medizen.

[210]  M. Givskov,et al.  In vitro screens for quorum sensing inhibitors and in vivo confirmation of their effect , 2010, Nature Protocols.

[211]  Thomas Bjarnsholt,et al.  Towards diagnostic guidelines for biofilm-associated infections. , 2012, FEMS immunology and medical microbiology.

[212]  D. Hassett,et al.  Involvement of Nitric Oxide in Biofilm Dispersal of Pseudomonas aeruginosa , 2006, Journal of bacteriology.

[213]  T. Tolker-Nielsen,et al.  Quorum Sensing and Virulence of Pseudomonas aeruginosa during Lung Infection of Cystic Fibrosis Patients , 2010, PloS one.

[214]  C. Hill,et al.  Bacteriophages ϕMR299-2 and ϕNH-4 Can Eliminate Pseudomonas aeruginosa in the Murine Lung and on Cystic Fibrosis Lung Airway Cells , 2012, mBio.

[215]  N. Høiby Pseudomonas aeruginosa infection in cystic fibrosis. Diagnostic and prognostic significance of pseudomonas aeruginosa precipitins determined by means of crossed immunoelectrophoresis. A survey. , 1977, Acta pathologica et microbiologica Scandinavica. Supplement.

[216]  L. Jacobsen,et al.  Pseudomonas aeruginosa infection in cystic fibrosis. Diagnostic and prognostic significance of Pseudomonas aeruginosa precipitins determined by means of crossed immunoelectrophoresis. , 1977, Scandinavian journal of respiratory diseases.

[217]  A. Oliver,et al.  High β-Lactamase Levels Change the Pharmacodynamics of β-Lactam Antibiotics in Pseudomonas aeruginosa Biofilms , 2012, Antimicrobial Agents and Chemotherapy.

[218]  N. Høiby,et al.  Biological Trojan Horse: Antigen 43 Provides Specific Bacterial Uptake and Survival in Human Neutrophils , 2006, Infection and Immunity.

[219]  H. Schweizer,et al.  Molecular Basis of Azithromycin-Resistant Pseudomonas aeruginosa Biofilms , 2005, Antimicrobial Agents and Chemotherapy.

[220]  M. Rohde,et al.  Pseudomonas aeruginosa cupA‐encoded fimbriae expression is regulated by a GGDEF and EAL domain‐dependent modulation of the intracellular level of cyclic diguanylate , 2007 .

[221]  N. Høiby,et al.  Early aggressive eradication therapy for intermittent Pseudomonas aeruginosa airway colonization in cystic fibrosis patients: 15 years experience. , 2008, Journal of cystic fibrosis : official journal of the European Cystic Fibrosis Society.

[222]  Liang Yang,et al.  Effects of ginseng on Pseudomonas aeruginosa motility and biofilm formation. , 2011, FEMS immunology and medical microbiology.

[223]  L. J. Douglas,et al.  Prostaglandin production during growth of Candida albicans biofilms. , 2005, Journal of medical microbiology.

[224]  G. O’Toole,et al.  Tobramycin and FDA-approved iron chelators eliminate Pseudomonas aeruginosa biofilms on cystic fibrosis cells. , 2009, American journal of respiratory cell and molecular biology.

[225]  S. Molin,et al.  Involvement of N‐acyl‐l‐homoserine lactone autoinducers in controlling the multicellular behaviour of Serratia liquefaciens , 1996, Molecular microbiology.

[226]  Abdelwahab Omri,et al.  Importance of DNase and alginate lyase for enhancing free and liposome encapsulated aminoglycoside activity against Pseudomonas aeruginosa. , 2009, The Journal of antimicrobial chemotherapy.

[227]  T. Tolker-Nielsen,et al.  Detection of Bacteria by Fluorescence in Situ Hybridization in Culture‐Negative Soft Tissue Filler Lesions , 2009, Dermatologic surgery : official publication for American Society for Dermatologic Surgery [et al.].

[228]  E. Greenberg,et al.  Chelator-Induced Dispersal and Killing of Pseudomonas aeruginosa Cells in a Biofilm , 2006, Applied and Environmental Microbiology.

[229]  T. Bauer,et al.  Brief Ultrasonication Improves Detection of Biofilm-formative Bacteria Around a Metal Implant , 2006, Clinical orthopaedics and related research.

[230]  J. Aronson,et al.  What is a clinical trial? , 2004, British journal of clinical pharmacology.

[231]  T. Tolker-Nielsen,et al.  Tolerance to the antimicrobial peptide colistin in Pseudomonas aeruginosa biofilms is linked to metabolically active cells, and depends on the pmr and mexAB‐oprM genes , 2008, Molecular microbiology.

[232]  T. Tolker-Nielsen,et al.  Effects of Antibiotics on Quorum Sensing in Pseudomonas aeruginosa , 2008, Antimicrobial Agents and Chemotherapy.

[233]  A. Marchese,et al.  Activity of daptomycin on biofilms produced on a plastic support by Staphylococcus spp. , 2008, International journal of antimicrobial agents.

[234]  A. Baccarelli,et al.  Internally coated endotracheal tubes with silver sulfadiazine in polyurethane to prevent bacterial colonization: a clinical trial , 2008, Intensive Care Medicine.

[235]  L. Sampath,et al.  Evaluation of the Antimicrobial Efficacy of Urinary Catheters Impregnated With Antiseptics in an In Vitro Urinary Tract Model , 2003, Infection Control & Hospital Epidemiology.

[236]  D. Warren,et al.  Clinical practice guidelines for the diagnosis and management of intravascular catheter-related infection: 2009 Update by the Infectious Diseases Society of America. , 2009, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[237]  T. Tolker-Nielsen,et al.  Interference of Pseudomonas aeruginosa signalling and biofilm formation for infection control , 2010, Expert Reviews in Molecular Medicine.

[238]  L. Saiman The use of macrolide antibiotics in patients with cystic fibrosis , 2004, Current opinion in pulmonary medicine.

[239]  T. Tolker-Nielsen,et al.  Distribution, Organization, and Ecology of Bacteria in Chronic Wounds , 2008, Journal of Clinical Microbiology.

[240]  Alan D. Lopez,et al.  The Global Burden of Disease Study , 2003 .

[241]  E. Greenberg,et al.  The potential of desferrioxamine-gallium as an anti-Pseudomonas therapeutic agent , 2008, Proceedings of the National Academy of Sciences.

[242]  Pradeep K. Singh,et al.  The transition metal gallium disrupts Pseudomonas aeruginosa iron metabolism and has antimicrobial and antibiofilm activity. , 2007, The Journal of clinical investigation.

[243]  S. Molin,et al.  Pseudomonas aeruginosa tolerance to tobramycin, hydrogen peroxide and polymorphonuclear leukocytes is quorum-sensing dependent. , 2005, Microbiology.

[244]  E. Änggård,et al.  A controlled clinical trial of a therapeutic bacteriophage preparation in chronic otitis due to antibiotic‐resistant Pseudomonas aeruginosa; a preliminary report of efficacy , 2009, Clinical otolaryngology : official journal of ENT-UK ; official journal of Netherlands Society for Oto-Rhino-Laryngology & Cervico-Facial Surgery.

[245]  Alan D. Lopez,et al.  Alternative projections of mortality and disability by cause 1990–2020: Global Burden of Disease Study , 1997, The Lancet.

[246]  E. Witsø,et al.  Sonication is superior to scraping for retrieval of bacteria in biofilm on titanium and steel surfaces in vitro , 2009, Acta orthopaedica.

[247]  M. Elkins,et al.  Antibiotic Susceptibilities of Pseudomonas aeruginosa Isolates Derived from Patients with Cystic Fibrosis under Aerobic, Anaerobic, and Biofilm Conditions , 2005, Journal of Clinical Microbiology.

[248]  M. Sugai,et al.  Effect of the Growth Rate of Pseudomonas aeruginosa Biofilms on the Susceptibility to Antimicrobial Agents: β-Lactams and Fluoroquinolones , 1999, Chemotherapy.

[249]  S. Molin,et al.  Mutations in 23S rRNA Confer Resistance against Azithromycin in Pseudomonas aeruginosa , 2012, Antimicrobial Agents and Chemotherapy.