A Functional DNase I Coating to Prevent Adhesion of Bacteria and the Formation of Biofilm

Biofilms are detrimental in many industrial and biomedical applications and prevention of biofilm formation has been a prime challenge for decades. Biofilms consist of communities of adhering bacteria, supported and protected by extracellular-polymeric-substances (EPS), the so-called house of biofilm organisms. EPS consists of water, proteins, polysaccharides and extracellular-DNA (eDNA). eDNA, being the longest molecule in EPS, connects the different EPS components and therewith holds an adhering biofilm together. eDNA is associated with bacterial cell surfaces by specific and non-specific mechanisms, mediating binding of other biopolymers in EPS. eDNA therewith assists in facilitating adhesion, aggregation and maintenance of biofilm structure. Here, a new method is described to prevent biofilm formation on surfaces by applying a DNase I enzyme coating to polymethylmethacrylate, using dopamine as an intermediate. The intermediate coupling layer and final DNase I coating are characterized by water-contact-angle measurements and X-ray photoelectron-spectroscopy. The DNase I coating strongly reduces adhesion of Staphylococcus aureus (95%) and Pseudomonas aeruginosa (99%) and prevents biofilm formation up to 14 h, without affecting mammalian cell adhesion and proliferation. Also agarose-gel-electrophoresis indicates loss of enzyme activity between 8 and 24 h. This duration however, is similar to many local antibiotic-delivery devices, which makes it an ideal coating for biomaterial implants and devices, known to fail due to biofilm formation with disastrous consequences for patients and high costs to the healthcare system. With threatening increases in antibiotic resistance, the DNase I coating may provide a timely, potent new approach to biofilm prevention on biomaterial implants and devices.

[1]  L. Mashburn-Warren,et al.  Special delivery: vesicle trafficking in prokaryotes , 2006, Molecular microbiology.

[2]  Haeshin Lee,et al.  Mussel-Inspired Surface Chemistry for Multifunctional Coatings , 2007, Science.

[3]  W. Hinrichs,et al.  Formulation and process development of (recombinant human) deoxyribonuclease I as a powder for inhalation , 2009, Pharmaceutical development and technology.

[4]  J. M. Barrales-rienda,et al.  Validation and in vitro characterization of antibiotic-loaded bone cement release. , 2000, International journal of pharmaceutics.

[5]  Haeshin Lee,et al.  Facile Conjugation of Biomolecules onto Surfaces via Mussel Adhesive Protein Inspired Coatings , 2009, Advanced materials.

[6]  Annette Moter,et al.  Dental plaque biofilms: communities, conflict and control. , 2011, Periodontology 2000.

[7]  H. Busscher,et al.  DNA-mediated bacterial aggregation is dictated by acid–base interactions , 2011 .

[8]  J. Mattick,et al.  Extracellular DNA required for bacterial biofilm formation. , 2002, Science.

[9]  L. Cellini,et al.  Extracellular DNA in Helicobacter pylori biofilm: a backstairs rumour , 2011, Journal of applied microbiology.

[10]  S. Kjelleberg,et al.  A characterization of DNA release in Pseudomonas aeruginosa cultures and biofilms , 2006, Molecular microbiology.

[11]  R. Deora,et al.  Extracellular DNA Is Essential for Maintaining Bordetella Biofilm Integrity on Abiotic Surfaces and in the Upper Respiratory Tract of Mice , 2011, PloS one.

[12]  L. Hancock,et al.  Regulation of Autolysis-Dependent Extracellular DNA Release by Enterococcus faecalis Extracellular Proteases Influences Biofilm Development , 2008, Journal of bacteriology.

[13]  P. Holden,et al.  Extracellular DNA in Single- and Multiple-Species Unsaturated Biofilms , 2005, Applied and Environmental Microbiology.

[14]  H. Fuchs,et al.  Effect of aerosolized recombinant human DNase on exacerbations of respiratory symptoms and on pulmonary function in patients with cystic fibrosis. The Pulmozyme Study Group. , 1994, The New England journal of medicine.

[15]  Y. Brun,et al.  A bacterial extracellular DNA inhibits settling of motile progeny cells within a biofilm , 2010, Molecular microbiology.

[16]  A. Kruse,et al.  Beta toxin catalyzes formation of nucleoprotein matrix in staphylococcal biofilms , 2010, Proceedings of the National Academy of Sciences.

[17]  E. Bakker,et al.  Pharmacology, clinical efficacy and safety of recombinant human DNase in cystic fibrosis , 2007, Expert review of respiratory medicine.

[18]  G. Tetz,et al.  Effect of DNase and Antibiotics on Biofilm Characteristics , 2008, Antimicrobial Agents and Chemotherapy.

[19]  Jeffrey B. Kaplan,et al.  Differential Roles of Poly-N-Acetylglucosamine Surface Polysaccharide and Extracellular DNA in Staphylococcus aureus and Staphylococcus epidermidis Biofilms , 2007, Applied and Environmental Microbiology.

[20]  S. Molin,et al.  A dual role of extracellular DNA during biofilm formation of Neisseria meningitidis , 2010, Molecular microbiology.

[21]  Joon-Seok Lee,et al.  Spatial control of cell adhesion and patterning through mussel-inspired surface modification by polydopamine. , 2010, Langmuir : the ACS journal of surfaces and colloids.

[22]  S. Tsuneda,et al.  Extracellular polymeric substances responsible for bacterial adhesion onto solid surface. , 2003, FEMS microbiology letters.

[23]  Haeshin Lee,et al.  General functionalization route for cell adhesion on non-wetting surfaces. , 2010, Biomaterials.

[24]  D. Allison,et al.  The Biofilm Matrix , 2003, Biofouling.

[25]  Ethan E. Mann,et al.  Modulation of eDNA Release and Degradation Affects Staphylococcus aureus Biofilm Maturation , 2009, PloS one.

[26]  R. Kolter,et al.  Biofilm formation as microbial development. , 2000, Annual review of microbiology.

[27]  Rosário Oliveira,et al.  Exopolymers in bacterial adhesion: interpretation in terms of DLVO and XDLVO theories , 1999 .

[28]  Zhixiong Xie,et al.  Role of DNA in Bacterial Aggregation , 2008, Current Microbiology.

[29]  Jie Li,et al.  Oxidant-induced dopamine polymerization for multifunctional coatings , 2010 .

[30]  Zhiqiang Qin,et al.  Role of autolysin-mediated DNA release in biofilm formation of Staphylococcus epidermidis. , 2007, Microbiology.

[31]  S. Molin,et al.  Role of Extracellular DNA during Biofilm Formation by Listeria monocytogenes , 2010, Applied and Environmental Microbiology.

[32]  Henk J. Busscher,et al.  Role of Extracellular DNA in Initial Bacterial Adhesion and Surface Aggregation , 2010, Applied and Environmental Microbiology.

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

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

[35]  S. Periasamy,et al.  How Staphylococcus aureus biofilms develop their characteristic structure , 2012, Proceedings of the National Academy of Sciences.

[36]  S. Goodman,et al.  Biofilms can be dispersed by focusing the immune system on a common family of bacterial nucleoid-associated proteins , 2011, Mucosal Immunology.

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

[38]  P. Marsh,et al.  A review of biofilms and their role in microbial contamination of dental unit water systems (DUWS) , 2004 .

[39]  R. Hancock A brief on bacterial biofilms , 2001, Nature Genetics.

[40]  T. Beveridge,et al.  Interactions of DNA with Biofilm-Derived Membrane Vesicles , 2009, Journal of bacteriology.

[41]  Norbert F Scherer,et al.  Single-molecule mechanics of mussel adhesion , 2006, Proceedings of the National Academy of Sciences.

[42]  J. Theron,et al.  DNA as an Adhesin: Bacillus cereus Requires Extracellular DNA To Form Biofilms , 2009, Applied and Environmental Microbiology.

[43]  T. Yasuda,et al.  Measurement of deoxyribonuclease I activity in human tissues and body fluids by a single radial enzyme-diffusion method. , 1993, Clinical chemistry.