MICROSTRUCTURED SURFACES FOR REDUCING BACTERIAL ADHESION

Background

Biofilms are structured communities of microorganisms encased in an extracellular polymeric matrix that typically are tenaciously adhered to the surface of biomaterials and host tissue. Bacterial biofilms are a significant issue in the development of materials that are exposed to aqueous and body fluids for prolonged periods for several different application areas: medical devices, filtration systems for food processing and other industrial applications, coatings for marine structures and other anti-fouling applications. Bacteria living in a biofilm are considerably more resistant to host defenses and antibiotic or antimicrobial treatments, when compared to "free" pathogens, and thereby increase the potential for infections during the use of in-dwelling and other tissue contacting devices.

Biofilms are believed to have a significant role in catheter associated urinary tract infections (CAUTI) and ventilator associated pneumonia (VAP). CAUTIs comprise the largest percentage of hospital acquired infections (HAIs) and are the second most common cause of nosocomial bloodstream infections. VAP has the highest morbidity of all HAIs, as roughly 15% of patients with VAP will die.

VAP may also be the most expensive HAI to treat ($20,000-$50,000 per episode), and has an incident rate between 25% and 40%.

By way of example, and without wishing to be bound by theory, biofilm formation on urinary catheter surfaces may proceed as follows: 1) The catheter surface is initially colonized by bacteria (some of them urease-producing bacteria) originally present on the periurethral skin and able to migrate into the bladder between the epithelial surface of the urethra and the catheter once the catheter is inserted. The adsorption of these cells to the catheter surface may be facilitated by the formation of an organic conditioning film made up largely of adsorbed proteins. 2) A bacterial biofilm community forms, encased primarily by a matrix of bacterial exopolysaccharide. The pioneer biofilm forming bacteria that initially cause urinary tract infections (UTIs) are typically S. epidermidis, E. coli or E. faecalis, with E. coli the overwhelming cause of CAUTI. At longer catheterization times, other species appear including P.

aeruginosa, P. mirabilis, and K. pneumoniae. These latter stage bacteria are more difficult to treat with antibiotics while the catheter is in place. 3) The presence of a growing biofilm, including bacterial species that are capable of producing urease, leads to an elevation of the urine's pH due to the action of urease on urea. 4) As the urine becomes alkaline, calcium phosphate and magnesium ammonium phosphate crystals precipitate and accumulate in the biofilm matrix growing on the catheter surface. 5) Continued crystal formation in the alkaline urine and continued growth of the biofilm lead to severe encrustation and eventually blockage of the device which necessitates re-catherization of the patient. Thus, preventing colonization and biofilm formation on the catheter could play a large role preventing CAUTIs.

Attempts have been made to provide surfaces that are inherently antimicrobial, either by composition or use of antimicrobial drug delivery systems. These surfaces can be insufficiently effective in reducing bio film formation for three important reasons: 1) when used as a delivery system, antimicrobial or active agents may be exhausted well before the end of the service lifetime of the medical article; 2) the surface antimicrobial properties are eventually impaired as dead cells, the high organic load in the urethra, and other adsorbed biomaterial mask the antimicrobial properties of that surface; and 3) antimicrobial agents in the catheter material or in an external coating fail to elute sufficiently.

Further attempts have been made to provide surfaces having an optimized engineered roughness index. Such attempts suggest that increased surface complexity is needed to sufficiently disrupt microorganism adhesion, indicating that simpler patterns are inadequate. While effective at reducing microbial adhesion in certain circumstances, these surfaces may be costly to reproduce on a sufficient scale. Effectively controlling and preventing biofilm formation long-term requires the creation of biomaterial surfaces that retain superior anti-adhesive properties throughout the useful life of the base medical article.

Summary

A medical article of the present disclosure includes a surface topography (i.e., the surface features of an object or region thereof) for resisting bioadhesion of microorganisms. The surface topography may be integral with or affixed to an exterior and/or interior surface of the article. The engineered surface has a topography comprising at least one pattern or arrangement defined by a plurality of unit cells. Each unit cell comprises at least one microstructure protruding from or projected into that surface. The unit cell typically includes at least one microstructure having a single size, shape and orientation. In certain embodiments, the unit cell is at least partially defined by a dimension at least approximating the pitch (i.e., distance between adjacent microstructures as measured centroid to centroid) between adjacent microstructures. In some embodiments of the present disclosure, the entire engineered surface is comprised of a single repeating unit cell geometry. In further embodiments, the plurality of unit cells are tiled and/or tessellated, in that the outer boundaries of a particular unit cell are directly adjacent the outer boundaries of any neighboring unit cell.

In certain embodiments, the microstructures defining the unit cell includes a base having a cross sectional dimension no less than 0.5 microns and no greater than 5 microns. To ensure adequate disruption of bacterial adhesion, the pitch between adjacent microstructures is at least the smallest dimension of the microstructure and may be no greater than 5 times said smallest dimension.

Surface topographies according to the invention resist bioadhesion as compared to a surface without such topography. Surface topographies according to the invention can be created by affixing a film or other substrate containing the plurality of microstructures to a target surface of the medical article or by microreplicating the structural features directly to the surface of the article. When microreplicated, the microstructures will be monolithically integrated with the underlying article. In other embodiments, the microstructures can be created by photolithography.

The engineered surfaces of the present invention can reduce the colonization of target microorganisms by at least 50% over 14 days compared to the colonization on a flat surface comprised of the same material, in the absence of antimicrobials and/or with any antimicrobial agents inactivated, according to the assay as set out in the Examples below. In preferred embodiments, the reduction in colonization can be at least 75%. In even more preferred embodiments, the reduction in colonization can be as high as 90%. In certain embodiments, the target organisms comprise Pseudomonas aeruginosa, Staphylococcus aureus; and methicillin-resistant Staphylococcus aureus.

Surprisingly, surfaces modified according to the present disclosure are equally or at least comparatively effective at resisting bioadhesion of target organisms in comparison to more complex topographies. The present inventors have found that both structure size and spacing impact the surface resistance to bioadhesion, particularly when both are commensurate with the size of the target microorganism. This discovery allows for the creation of repeating patterns of similarly sized microstructures in simplified arrays. These simplified arrays are expected to significantly improve the process efficiency by reducing shrinkage, providing more predictable and more uniform shrinkage, as well as providing more thermal uniformity in thermoplastic and thermoset molds. Thus, the preferred topographies of the present invention may be easily replicated with enhanced feature fidelity and manufactured on an industrial scale at appreciably reduced cost. The expected improvement in production cost and quality makes the engineered surfaces of the present invention particularly suited for a wide variety of potential applications.

In one aspect, the present disclosure provides a medical article including a surface for reducing bacterial adhesion. The article includes an engineered surface comprising a thermoplastic or thermoset material and a plurality of engineered microstructures on at least a portion of the surface, wherein each microstructure of the plurality of microstructures comprises a base having at least one microscale cross- sectional dimension. The aspect ratio of each microstructure is at least 0.5 and no greater than 10, each microstructure includes a height greater than 0.5 microns, the pitch between adjacent microstructures of the plurality of engineered microstructures is at least 1 time and no greater than 5 times than the smallest cross-sectional dimension, and no base comprises a cross-sectional dimension greater than 5 microns.

The arrangement of the microstructures on at least a portion of the engineered surface includes a plurality of unit cells, wherein each unit cell of the plurality of unit cells is at least partially defined by a dimension at least approximating the pitch and includes no more than one microstructure of the plurality of microstructures; each unit cell comprises a boundary and each unit cell boundary is directly adjacent the boundary of every neighboring unit cell; and the plurality of unit cells includes no more than three unique unit cell geometries.

In another aspect, the present disclosure provides a medical article including a surface for reducing bacterial adhesion; the article includes an engineered surface comprising a thermoplastic or thermoset material and a plurality of engineered microstructures on at least a portion of the surface, each microstructure of the plurality of microstructures including a base having at least one microscale cross- sectional dimension. . The aspect ratio of each microstructure is at least 0.5 and no greater than 10, each microstructure of the plurality of engineered microstructures includes a height greater than 0.5 microns, and no microstructure includes a base cross-sectional dimension greater than 5 microns. The arrangement of the microstructures on at least a portion of the engineered surface includes a plurality of unit cells including no more than three unique unit cell geometries. Each unit cell includes no more than one microstructure and a peripheral boundary defining the unit cell. Each unit cell boundary is directly adjacent the peripheral boundary of every neighboring unit cell (i.e., the unit cells are tiled).

In another aspect, the present disclosure provides a medical article including a surface capable of reduced bacterial adhesion with an engineered surface including a thermoset material and a plurality of engineered microstructures on at least a portion of the engineered surface. Each microstructure of the plurality of microstructures includes a base having at least one cross sectional dimension, wherein the largest cross-sectional dimension of the base is at least 1 micron and no greater than 2 microns, wherein the aspect ratio of each microstructure is at least 0.5 and no greater than 10, and wherein the spacing between adjacent microstructures of the plurality of engineered microstructures is at least 1 time and no greater than 5 times than the smallest cross-sectional dimension. The arrangement of the microstructures on at least a portion of the engineered surface defines a periodic unit cell, each unit cell having the same or substantially same geometry and including no more than one microstructure.

In another aspect, the present disclosure provides a method of controlling microorganism adhesion to a medical article. The method includes providing a medical article having a surface including a polymeric material and a plurality of engineered microstructures on at least a portion of the surface, wherein each microstructure of the plurality of microstructures includes a base having at least one cross sectional microscale dimension, wherein the aspect ratio of each microstructure is at least 0.5 and no greater than 10, wherein the pitch between adjacent microstructures of the plurality of engineered microstructures is at least 1 time and no greater than 5 times the smallest cross-sectional dimension, wherein no base comprises a cross-sectional dimension greater than 5 microns, and wherein the arrangement of the microstructures on at least a portion of the surface includes a plurality of unit cells, wherein each unit cell of the plurality of unit is at least partially defined by a dimension at least approximating the pitch, and wherein each unit cell includes a boundary and each unit cell is directly adjacent the boundary of the nearest unit cell. The method further includes placing the surface proximate a tissue or fluid, wherein the colonization of a target microorganism on the portion of the surface including the plurality of engineered microstructures is reduced in comparison to a flat surface comprised of the same material.

In yet another aspect, the present disclosure provides a method of making a biofilm adhesion resistant medical article. The method includes providing a medical article having a surface comprising a polymeric material. The method further includes creating a plurality of engineered microstructures on at least a portion of the surface, wherein each microstructure of the plurality of microstructures includes a base having at least one cross sectional microscale dimension, wherein the aspect ratio of each microstructure is at least 0.5 and no greater than 10, wherein the pitch between adjacent microstructures of the plurality of engineered microstructures is at least 1 time and no greater than 5 times than the smallest cross-sectional dimension, wherein no base of any microstructure includes a cross-sectional dimension greater than 5 microns, and wherein the arrangement of the microstructures on at least a portion of the surface includes a plurality of unit cells, wherein each unit cell of the plurality of unit is at least partially defined by a dimension at least approximating the pitch, and wherein each unit cell comprises a boundary and each unit cell boundary is directly adjacent the boundary of the nearest unit cell.

As used herein "geometry" refers to the size and shape of a feature.

As used herein "base" of a structure is defined at the plane of the substrate surface from which the microstructure emerges. In "negative" microstructures which project into a substrate (e.g., holes) the base is defined at the plane of the substrate surface, i.e., at the entrance to the "hole".

As used herein, a "microstructure" is a structure or feature having a recognizable geometric shape defined by a volume that projects out the base plane of a surface or an indented volume which projects into the surface. Such structures typically include a base having cross sectional dimensions no less than 0.5 microns and no greater than 5 microns. In certain implementations, a microstructure can be a collection or amalgamation of filaments. In certain implementations, a microstructure can be porous.

As used herein, an "engineered microstructure" shall mean a microstructure deliberately formed into and integral with a surface. An engineered microstructure may be created, for example, by microreplicating a specific pattern unto a surface. An engineered microstructure is distinct from structures produced by random application of particles, by spraying, adhesive bonding, etc, to a surface.

As used herein, the term "microstructured surface" is generally used to refer to a surface that comprises microstructures or microstructured features.

The term "microreplicate" and derivatives thereof, is generally used to refer to the production of a microstructured surface through a process where the structured surface features retain an individual feature fidelity during and after manufacture.

As used herein, the term "pitch" identifies the distance between the centroids of adjacent microstructures. The pitch is measured from the centroid of a microstructure (i.e., the geometric center) to the centroid of an adjacent microstructure.

As used herein, the terms "height" , "base" and "top" are for illustrative purposes only, and do not necessarily define the orientation or the relationship between the surface and the microstructure. For example, the "height" of a microstructure projected into a surface can be considered the same as the depth of recess created, and the "top" the bottom said recess. Accordingly, the terms "height" and "depth", and "top" and "bottom" should be considered interchangeable.

The terms "comprises" and variations thereof do not have a limiting meaning where these terms appear in the description and claims.

The words "preferred" and "preferably" refer to embodiments of the invention that may afford certain benefits, under certain circumstances. However, other embodiments may also be preferred, under the same or other circumstances. Furthermore, the recitation of one or more preferred embodiments does not imply that other embodiments are not useful, and is not intended to exclude other embodiments from the scope of the invention.

As recited herein, all numbers should be considered modified by the term "about".

As used herein, "a," "an," "the," "at least one," and "one or more" are used interchangeably. Thus, for example, an article comprising an "engineered surface" can be interpreted to comprise one or more "engineered surfaces"

Also herein, the recitations of numerical ranges by endpoints include all numbers subsumed within that range (e.g., 1 to 5 includes 1, 1.5, 2, 2.75, 3, 3.80, 4, 5, etc.).

The above summary of the present invention is not intended to describe each disclosed embodiment or every implementation of the present invention. The description that follows more particularly exemplifies illustrative embodiments. In several places throughout the application, guidance is provided through lists of examples, which examples can be used in various combinations. In each instance, the recited list serves only as a representative group and should not be interpreted as an exclusive list.

Brief Description of the Drawings

The invention will be further described with reference to the drawings, wherein corresponding reference characters indicate corresponding parts throughout the several views, and wherein: Figs, la-c are cross-sectional views of an article including an engineered surface according to embodiments of the disclosure.

Fig. 2 illustrates a surface comprising a series of discrete recesses according to an embodiment of the disclosure.

Figs. 3a-c illustrate various shapes of microstructures according to the present description.

Figs. 4a-b illustrate perspective views of engineered surfaces according to embodiments of the disclosure. Figs. 5a-c illustrate a variety of microstructure configurations according to other embodiments of the disclosure.

Figs. 6a-d illustrate a variety of engineered surfaces according to certain embodiments of the disclosure. Figs. 7a-f illustrate configurations of repeating unit cells according to certain embodiments of the disclosure.

Figs. 8a-c illustrate configurations of repeating unit cells according to certain embodiments of the disclosure.

Fig. 9 illustrates an undulating substrate according to certain implementations of the disclosure. Detailed Description of Illustrative Embodiments

Medical articles of the present disclosure provide reduced colonization of target microorganisms, and thus biofilm formation, on surfaces of the article that include a plurality of engineered

microstructures. In certain embodiments, the engineered surface has at least one unit cell defined at least partially by the pitch between adjacent microstructures protruding from or projected into that surface. Each unit cell is defined in a single plane and includes at least one microstructure having a single geometry and orientation. The unit cells are arranged to be tiled or at least substantially tessellated, such that the boundary region of any one unit cell is directly adjacent to the boundary region of any neighboring unit cell (i.e., there is no deliberate space between the unit cells).

In some embodiments of the present disclosure, at least a portion of the engineered surface is comprised of a single repeating unit cell, and accordingly one microstructure geometry. In other embodiments, at least a portion of the surface comprises a plurality of unit cells that include a plurality of microstructure geometries.

The term "microorganism" is generally used to refer to any prokaryotic or eukaryotic microscopic organism, including without limitation, one or more of bacteria (e.g., motile or vegetative, Gram positive or Gram negative), bacterial spores or endospores, algae, fungi (e.g., yeast, filamentous fungi, fungal spores), mycoplasmas, and protozoa, as well as combinations thereof. In some cases, the microorganisms of particular interest are those that are pathogenic, and the term "pathogen" is used to refer to any pathogenic microorganism. Examples of pathogens can include, but are not limited to, both Gram positive and Gram negative bacteria, fungi, and viruses including members of the family

Enter obacteriaceae, or members of the family Micrococaceae, or the genera Staphylococcus spp., Streptococcus, spp., Pseudomonas spp., Enterococcus spp., Salmonella spp., Legionella spp., Shigella spp., Yersinia spp., Enterobacter spp., Escherichia spp., Bacillus spp., Listeria spp., Campylobacter spp., Acinetobacter spp., Vibrio spp., Clostridium spp., Klebsiella spp., Proteus spp. and Corynebacterium spp. Particular examples of pathogens can include, but are not limited to, Escherichia coli including enterohemorrhagic E. coli e.g., serotype 0157:H7, 0129:H1 1 ; Pseudomonas aeruginosa; Bacillus cereus; Bacillus anthracis; Salmonella enteritidis; Salmonella enterica serotype Typhimurium; Listeria monocytogenes; Clostridium botulinum; Clostridium perfringens; Staphylococcus aureus; methicillin- resistant Staphylococcus aureus; Campylobacter jejuni; Yersinia enterocolitica; Vibrio vulnificus;

Clostridium difficile; vancomycin-resistant Enterococcus; Klebsiella pnuemoniae; Proteus mirabilus and Enterobacter [Cronobacter] sakazakii.

A medical article 100 including at least one engineered surface 1 10 for contact with a tissue or fluid is depicted in Figure 1 a. In some embodiments, the engineered surface defines a portion of the exterior surface of medical article 100. In other embodiments, the engineered surface 1 10 defines a portion of the interior surface of a medical article 100. In yet other embodiments, the engineered surface may comprise at least a portion of both the interior and exterior surfaces of the medical article 100.

Suitable medical articles for use with the invention include, but are not limited to: nasal gastric tubes, wound contact layers, blood stream catheters, stents, pacemaker shells, heart valves, orthopedic implants such as hips, knees, shoulders, etc., periodontal implants, orthodontic brackets and other orthodontic appliances, dentures, dental crowns, contact lenses, intraocular lenses, soft tissue implants (breast implants, penile implants, facial and hand implants, etc.), surgical tools, sutures including degradable sutures, cochlear implants, tympanoplasty tubes, shunts including shunts for hydrocephalus, post surgical drain tubes and drain devices, urinary catheters, endotraecheal tubes, heart valves, wound dressings, other implantable devices, and other indwelling devices.

The medical article 100 includes a plurality of engineered microstructures 120 molded or integral with at least a portion of the engineered surface 1 10. In Figure la, the engineered microstructures 120 are depicted as post-like features projecting or protruding from the engineered surface 1 10. The engineered microstructure 120 can be substantially solid throughout (Figure la) or include pores 122 (Figure lc). In other implementations (Figure lb), the engineered microstructure 120 can be a collection of a plurality of fibers or filaments 121. In other embodiments as depicted in Figure 2, it may be preferred that the plurality engineered microstructures 220 are projected into the engineered surface 210 at height (i.e., depth) 214, creating a series of disconnected or discrete recesses. In such embodiments the projected or "negative" microstructures may improve the anti-adhesion capabilities of the engineered surfaces in comparison to a projected or "positive" equivalent microstructure. Without wishing to be bound by theory, the improved reduction in microorganism adhesion for negative microstructures could be due to a difference in how negative and positive structures are wetted by fluid (growth media) covering those surfaces. Whereas a positive structure is defined by an interconnected or continuous channel structure, the equivalent negative pattern is defined by a disconnected or discrete recess or channel structure (e.g., the pockets may form discontinuous channels). A somewhat high surface tension liquid contacting a positive structure may be able to fully wet that surface because the entrapped air may be displaced more readily due to the presence of an interconnected channel structure. The same liquid on a negative structure, with discrete micron sized recesses, may not be able to fully wet that surface because the surface tension over an opening of that size is difficult to break, and the air or other gasses entrapped in those recesses cannot be easily displaced. The net result is a lower fractional area available for bacterial contact and subsequent adhesion.

Generally, the microstructures may be composed of all or substantially all of the same material. More specifically, the microstructures may be made of a curable, thermoset material. In some embodiments, that material is a majority silicone polymer by weight. In at least some embodiments, the silicone polymer will be polydialkoxysiloxane such as poly(dimethylsiloxane) (PDMS), such that the microstructures are made of a material that is a majority PDMS by weight. More specifically, the microstructures may be all or substantially all PDMS. For example, the microstructures may each be over 95wt.% PDMS. In certain embodiments the PDMS is a cured thermoset composition formed by the hydrosilylation of silicone hydride (Si-H) functional PDMS with unsaturated functional PDMS such as vinyl functional PDMS. The Si-H and unsaturated groups may be terminal, pendant, or both. In other embodiments the PDMS can be moisture curable such as alkoxysilane terminated PDMS.

In some embodiments, other silicone polymers besides PDMS may be useful, for example, silicones in which some of the silicon atoms have other groups that may be aryl, for example phenyl, alkyl, for example ethyl, propyl, butyl or octyl, fluororalkyl, for example 3,3,3-trifluoropropyl, or arylalkyl, for example 2-phenylpropyl. The silicone polymers may also contain reactive groups, such as vinyl, silicon-hydride (Si-H), silanol (Si-OH), acrylate, methacrylate, epoxy, isocyanate, anhydride, mercapto and chloroalkyl. These silicones may be thermoplastic or they may be cured, for example, by condensation cure, addition cure of vinyl and Si-H groups, or by free-radical cure of pendant acrylate groups. They may also be cross-linked with the use of peroxides. Such curing may be accomplished with the addition of heat or actinic radiation. Other useful polymers may be thermoplastic or thermoset and include polyurethanes, polyolefins including metallocene polyolefins, polyesters such as elastomeric polyesters (e.g., Hytrel), biodegradable polyesters such as polylactic, polylactic/glycolic acids, copolymers of succinic acid and diols, and the like, fluoropolymers including fluoroelastomers, polyacrylates and polymethacrylates. Polyurethanes may be linear and thermoplastic or thermoset.

Polyurethanes may be formed from aromatic or aliphatic isocyanates combined with polyester or polyether polyols or a combination thereof. In another embodiment, polymers with a glass transition temperature of less than 25°C are useful. Particularly useful are polymers with a glass transition temperature of less than about 10°C. In at least some embodiments, the microstructures may be an elastomer. An elastomer may be understood as a polymer with the property of viscoelasticity (or elasticity) generally having suitably low Young's modulus and high yield strain compared with other materials. The term is often used interchangeably with the term rubber, although the latter is preferred when referring to cross-linked polymers. Polymers may also be filled with suitable organic or inorganic fillers and for certain applications the fillers are radioopaque. The polymers may contain other additives such as antimicrobial agents (including antiseptics and antibiotics), dyes, mold release agents, antioxidants, plasticizers, and the like. Suitable antimicrobials can be incorporated into or deposited onto the polymers. Suitable preferred antimicrobials include those described in US Publication Nos.

2005/0089539 and 2006/0051384 to Scholz et al. and US Publication Nos. 2006/0052452 and

2006/0051385 to Scholz. The microstructures of the present invention also may be coated with antimicrobial coatings such as those disclosed in International Application No. PCT/US201 1/37966 to Ali et al.

Referring again to Figure 1 , engineered surface 1 10 of the medical article 100 comprises a substrate. The substrate may planar, substantially planar, or included varying topography (e.g., undulations) as depicted in Figure 9. The substrate may be made from any number of suitable materials typically used to form medical articles. The substrate can be formed from a metal, alloy, polymer, biologic scaffolding, or a combination comprising at least one of the foregoing. The thickness of the substrate can vary depending on the use of the medical article. For example, in some embodiments, the substrate may be made from the same materials as microstructures 102, including those described above. In such embodiments, the substrate may be a material that is a majority silicone polymer by weight. In one exemplary embodiment, the substrate may be made of PDMS. In other exemplary embodiments, the substrate may be made of other commonly used substrates. Specifically, glass, ceramic, metal or polymeric substrates may be appropriate, as well as other suitable alternatives and combinations thereof such as ceramic coated polymers, ceramic coated metals, polymer coated metals, metal coated polymers and the like. The substrate can, in some implementations, include discrete pores and/or pores in communication.

Biocompatible metals for use as the substrate include stainless steel alloys such as type 316 L, chromium-cobalt-molybdenum alloys titanium alloys such as TiAl ₄ V, zirconium alloys, shape memory nickel-titanium alloys, super elastic nickel-titanium alloys, and combinations thereof.

The polymers used to form the substrate can be biodegradable, non-biodegradable, or combinations thereof. In addition, fiber- and/or particle-reinforced polymers can also be used. Non- limiting examples of suitable non-biodegradable polymers include polyisobutylene copolymers and styrene-isobutylene-styrene block copolymers, such as styrene-isobutylene-styrene tert-block copolymers (SIBS); polyvinylpyrrolidone including cross-linked polyvinylpyrrolidone; polyvinyl alcohols;

copolymers of vinyl monomers such as EVA; polyvinyl ethers; polyvinyl aromatics; polyethylene oxides; polyesters such as polyethylene terephthalate; polyamides; polyacrylamides; polyethers such as polyether sulfone; polyolefins such as polypropylene, polyethylene, highly crosslinked polyethylene, and high or ultra high molecular weight polyethylene; polyurethanes; polycarbonates; silicones; siloxane polymers; natural based polymers such as optionally modified polysaccharides and proteins including, but not limited to, cellulosic polymers and cellulose esters such as cellulose acetate; and combinations comprising at least one of the foregoing polymers. Combinations may include miscible and immiscible blends as well as laminates.

Non-limiting examples of suitable biodegradable, bioabsorbable, bioerodible polymers include polycarboxylic acid; polyanhydrides such as maleic anhydride polymers; polyorthoesters; poly-amino acids; polyethylene oxide; polyphosphazenes; polylactic acid, polyglycolic acid, and copolymers and mixtures thereof such as poly(L-lactic acid) (PLLA), poly(D,L,-lactide), poly(lactic acid-co-glycolic acid), and 50/50 weight ratio (D,L-lactide-co-glycolide); polydioxanone; polypropylene fumarate;

polydepsipeptides; polycaprolactone and co-polymers and mixtures thereof such as poly(D,L-lactide-co- caprolactone) and polycaprolactone co-blutylacrylate; polyhydroxybutyrate valerate and mixtures thereof; polycarbonates such as tyrosine-derived polycarbonates and arylates, polyiminocarbonates, and polydimethyltrimethylcarbonates; cyanoacrylate; calcium phosphates; polyglycosaminoglycans;

macromolecules such as polysaccharides (including hyaluronic acid, cellulose, and hydroxypropylmethyl cellulose; gelatin; starches; dextrans; and alginates and derivatives thereof, proteins and polypeptides; and mixtures and copolymers of any of the foregoing. The biodegradable polymer can also be a surface erodible polymer such as polyhydroxybutyrate and its copolymers, polycaprolactone, polyanhydrides (both crystalline and amorphous), and maleic anhydride.

Although Figures la-c illustrate post-shaped microstructures 120, a number of different microstructure shapes are contemplated. For example, as illustrated in Fig. 3a-d, Microstructures may be dome-shaped as illustrated in SEM image 3a, post-shaped, as shown on SEM image in Fig. 3b, a pattern that reportedly mimics shark skin, as shown in Fig. 3c. A wide variety of additional shapes are also contemplated.

Further examples of suitable microstructure shapes can include, but are not limited to, a variety of polyhedral shapes, parallelepipeds, prismatoids, prismoids, etc., and combinations thereof. For example, the microstructures can be polyhedral, conical, frusto-conical, pyramidal, frusto-pyramidal, spherical, partially spherical, hemispherical, ellipsoidal, dome-shaped, cylindrical, and combinations thereof.

Additional suitable microstructure shapes include irregular geometries that can be described by non-Euclidean mathematics. Non-Euclidean mathematics is generally used to describe those structures whose mass is directly proportional to a characteristic dimension of the spaced feature raised to a fractional power (e.g., fractional powers such as 1.34, 2.75, 3.53, or the like). Examples of geometries that can be described by non-Euclidean mathematics include fractals and other irregularly shaped

microstructures.

Generally, microstructures according to the present invention comprise a base 1 12 adjacent the engineered surface 1 10 and a top surface 108 separated from base 1 12 by a height 1 14. It should be appreciated that the terms "height" , "base" and "top" are for used illustrative purposes only, and do not necessarily define the relationship between the surface and the microstructure. For example, the "height" of a microstructure projected into a surface can be considered the same as the depth of recess created, and the "top surface" the bottom of said recess.

The base 1 12 of each engineered microstructure 120 may comprise a variety of cross-sectional shapes including, but not limited to, parallelograms, parallelograms with rounded corners, rectangles, squares, circles, half-circles, ellipses, half-ellipses, triangles, trapezoids, stars, other polygons (e.g., hexagons), etc., and combinations thereof.

Regardless of cross-sectional shape, each microstructure comprises a smallest cross-sectional dimension at the base 1 12. The smallest cross-sectional dimension of the base may be no greater than 20 microns, in some embodiments no greater than 10 microns, and in some embodiments no greater than 5 microns. The smallest cross-sectional dimension may be at least 0.1 microns, in some embodiments at least 0.5 microns, and in some embodiments at least 1 micron.

In certain preferred embodiments, no cross-sectional dimension at the base 1 12 exceeds 5 microns. Microstructures having cross-sectional dimensions no greater than 5 microns arranged according to the present disclosure are believed to substantially interfere with the settlement and adhesion of target bacteria most responsible for HAIs or other biofouling problems such as increased drag, reduced heat transfer, filtration fouling, etc. Generally, each micro-structure or the plurality of microstructures has a height 1 14 that is at least

0.5 microns. In some embodiments, each microstructure of the plurality of microstructures has a height of at least 1 micron, in other embodiments at least 1.5 microns, in other embodiments at least 2 microns, in other embodiments at least 3 microns and in other embodiments at least 5 microns.

In certain embodiments, the microstructure height is no greater than 100 microns, in some embodiments no greater than 50 microns, in some embodiments no greater than 30 microns, in some embodiments no greater than 20 microns, and in certain preferred embodiments no greater than 10 microns.

Microstructures including a height or smallest cross-sectional dimension less than 0.5 microns may not sufficiently reduce surface contact and subsequent adhesion of certain gram negative bacteria.

Whether protruding from or projecting into the engineered surface, each microstructure of the plurality of microstructures includes a particular aspect ratio. For microstructures comprising regular (e.g., Euclidean) and irregular (e.g., Non-Euclidean) cross-sectional shapes substantially throughout the height of the microstructure, the aspect ratio is defined herein as the ratio of the height to the smallest cross-sectional dimension (e.g., width, length, diameter) at the base. For irregularly shaped bases (bases which are not parallelograms or circles) the smallest cross-sectional dimension will be understood to be the diameter of a circle of equivalent area. Regardless of microstructure geometry, each microstructure of the plurality of microstructures includes an aspect ratio of at least 0.5, and in some embodiments at least

1. The aspect ratio of each microstructure may be no greater than 15, and in some embodiments no greater than 10.

In certain preferred embodiments, the plurality of engineered microstructures comprises an array of posts as depicted in Figure 1 and Figure 3b. The base 1 12 of the post may comprise a variety of cross- sectional shapes including, but not limited to, parallelograms, parallelograms with rounded corners, rectangles, squares, circles, half-circles, ellipses, half-ellipses, triangles, trapezoids, stars, other polygons (e.g., pentagons, hexagons, octagons), etc., and combinations thereof.

In certain embodiments, the cross-sectional area of the post does not substantially change in a vertical direction as the top surface 108 is approached. For example, if the base comprises a circular cross-section, the diameter will not substantially change between the top surface and the base. In other embodiments, the cross-sectional area may increase or decrease in a vertical direction relative to the engineered surface.

As depicted in Figure 5a, the top surface 520 of the post 500 is substantially planar and is substantially parallel to the engineered surface. It is further contemplated, however, that the top surface 520 include at least a portion that is concave (Fig. 5b) or convex (Fig. 5c), such that the post appears rounded or dimpled on the top surface. Though not depicted, the top surface 520 of the post 500 can include additional features having one of the myriad shapes discussed above.

Where the microstructures 420 are prism-shaped as in Figure 4, the pyramids may have a peak angle Θ _Ρ (or angle between the two facets of a prism) of 90 degrees. As the prism-shaped microstructures 420 are depicted are isosceles triangles, the angle of intersection of the two facets with the plane of the film will then be an angle of 45 degrees. In other embodiments, the peak angle may be greater or less than 90 degrees. For example, the peak angle may be between 90 degrees and 100 degrees, or between 80 degrees and 90 degrees.

Turning to Figures 6a - 6d, a plurality of microstructures may be arranged on an engineered surface according to any number of patterns. The engineered surface may be planar (as depicted in Figures 6a-6d), substantially planar, or included varying topography (e.g., undulations) as depicted in Figure 9. More generally, microstructures may be created that vary in one, two or three dimensions. For example in Fig. 6a, the microstructures may be structures that identically run a length 680 of the film at the same height along the vertical direction 690 without any segmentation. However, across the width 670 of the portion of the engineered surface, or across a first dimension, the film is segmented into different discrete microstructures. In addition, as shown in Fig. 6b, the microstructures may vary in two directions. For example, the microstructures may be segmented as in Fig. 6a along the width 670 (i.e., the x-axis) of a portion of the engineered surface, but also be segmented along the length 680 (i.e., y-axis) of that portion of the engineered surface. In such a case, discrete posts are located along both the x and y axes. Here, however, the structures are all the same height in the vertical direction (or third dimension) 690. Further, as shown in Fig. 6c, the structures may be segmented along both the width and length of the film and may also vary in the height of the microstructures across the surface in the z-axis (or third dimension). Finally, the engineered surface may comprise a plurality of microstructure shapes, including combinations of regular and irregular microstructures in any of the patterns described above. In any of these scenarios the microstructures may be directly adjacent to one another or may be spaced apart by some portion of the engineered surface that is substantially flat.

The microstructures can be useful t in certain implementations to control friction during the intended use of a medical article including an engineered surface. Friction may depend on a variety of factors, including but not limited to orientation, aspect ratio, and material compliance/deformability of the microstructures. These factors may be considered relative to the engineered surface, the neighboring microstructures, or both. Structures such as those shown in Fig. 6a have different frictional properties in at least two major directions (670 and 680). For example, it may be beneficial to have the different frictional directions oriented along the length of a catheter (urinary or venous) or an endotracheal tube.

As illustrated by certain previous figures herein (e.g., Fig. 4), the microstructures may be directly adjacent to one another, such that the base of a microstructure is directly in contact with the base of an adjacent microstructure. However, it should be understood that the microstructures may be further spaced apart, such that the facets or perimeters of the microstructures are not in contact and are spaced apart by a segment of engineered surface that may, for example, be flat. The configuration of microstructures in any given region should be chosen, however, so that the average pitch (i.e., the centroid to centroid distance between adjacent microstructures) is at least as large as the smallest dimension of the smallest microstructure and no greater than 5 times the smallest dimension of the microstructure. Surfaces having pitches outside this range may result in topographies that reduce bioadhesion in certain areas, but at least fail to impede or potentially promote bioadhesion in other areas.

In certain preferred embodiments, the microstructures are segmented in two dimensions, preferably the along the x-axis and the y-axis (i.e., the height of each microstructure along the z-axis is substantially the same). The average pitch may be the same in both dimensions. In other embodiments, the pitch along the x-axis is less than the pitch along the y-axis, and vice versa.

The arrangement of microstructures on the engineered surface may comprise a particular density of microstructures per square centimeter. In some implementations, the engineered surface comprises at least 1,000,000 microstructures per square centimeter, in some embodiments, at least 4,000,000, and in yet other embodiments at least 25,000,000 microstructures per square centimeter. The engineered surface may comprise no greater than 400,000,000-microstructures per square centimeter, in some embodiments no greater than 250,000,000, in some embodiments no greater than 100,000,000 , in some embodiments no greater than 75,00,000, and in other embodiments no greater than 50,000,000 microstructures/cm ² . Without wishing to be bound by theory, surfaces with microstructure densities less than 1,000,000 and greater than 400,000,000may not sufficiently disrupt biofilm formation in certain implementations, as such surfaces may provide greater area for attachment and are believed to act essentially as a flat surface.

As can be appreciated by reference to Figures 7a-f, the arrangement of microstructures 710 on at least a portion of the engineered surface 700 comprises a plurality of unit cells740. Each unit cell 740 of the plurality of unit cells 730 exists in a single plane (i.e., two dimensions), is at least partially defined by a dimension at least approximating the pitch, and contains one microstructure. In certain embodiments, the unit cell is at least partially defined by a dimension equal to the pitch. As depicted in Fig. 7a, each unit cell 740 is entirely defined by the pitch between adjacent microstructures. Each unit cell includes a boundary 750 defining the perimeter of the unit cell. Each boundary 750 is directly adjacent the boundary of a neighboring unit cell, so that the plurality of unit cells resemble, e.g., a grid or tessellation.

In other embodiments, as depicted in Fig. 7f, unit cells can include certain dimensions that are greater than the pitch. As can be appreciated, the length 770 of unit cell 748 is greater than the pitch 720.

In certain embodiments, a single unit cell geometry is repeated over at least a portion of the engineered surface 700. Preferably, each unit cell includes no more than one microstructure and all microstructures of the plurality of microstructures 740 share the substantially same or same geometry and/or orientation. In certain advantageous embodiments, such as the one depicted in Fig. 7a, at least a portion of the engineered surface comprises posts having the same geometry and orientation.

In another embodiment depicted in Fig. 7b, at least a portion of the engineered surface 700 comprises a plurality of microstructure geometries. For example, a unit cell including a pyramidal projections is directly adjacent a unit cell including a post. Each unit cell is still defined at least partially by a dimension at least approximating, preferably equal to, the pitch and includes a single microstructure. A variety of shapes may be used to define the unit cell around a single microstructure. Suitable unit cells may comprise rectangles, circles, half-circles, ellipses, half-ellipses, triangles, trapezoids, and other polygons (e.g., pentagons, hexagons, octagons), etc., and combinations thereof. Regardless of regular unit cell shape, the boundaries of any unit cell are directly adjacent the boundary of a neighboring unit cell and the unit cell is at least partially defined by a dimension equal to the pitch, such that the arrangement resembles a tessellation.

The arrangement of unit cells on any portion (or entire portion) of the engineered surface can resemble a structured or unstructured grid array. Exemplary structured arrays or tessellations include a Cartesian grid array as depicted in Figs. 7a-b and a regular grid array as depicted in Fig. 7c. In other embodiments, the arrangement of unit cells resembles a tessellation as depicted in Fig. 7d, with adjacent unit cells offset or alternating in one dimension. An exemplary unstructured array featuring three unique unit cell geometries (743, 745, and 747) is depicted in Figure 7e.

In certain embodiments, at least a portion of the engineered surface includes no more than three unique unit cell geometries. In other embodiments, at least a portion of the engineered surface includes no more than two unique unit cell geometries. In certain preferred embodiments, at least a portion of the engineered surface includes only one unit cell geometry repeated over said surface portion. As described above, the unit cell in such embodiments can contain microstructures having substantially similar geometry or microstructures having a different geometry. Engineered surfaces having more than three distinct unit cell geometries, as shown in Figure 7f as unit cells 742, 744, 746, and 748, render replication of the microstructures substantially more difficult due to e.g., shrinkage of material and mold replication difficulties.

In yet other embodiments, at least portions of the engineered surface may be defined by irregular unit cells containing one or more microstructure geometries and/or orientations. The unit cell in such embodiments is not necessarily at least partially defined by the pitch (but can be in certain embodiments). For example, the irregular unit cell may include the smallest dimension that is shared by one or more adjacent microstructures or by two adjacent unit cells. As another example as depicted in Figure 7e, the irregular unit cells can be drawn around the base shape of the microstructure and still resemble a tessellation. The microstructures of such irregular unit cells can have variety of geometries and can exist in one, two or three dimensions or any dimensions therebetween. The microstructures in such unit cells can have similar geometries with different dimensions, similar geometries with similar dimensions, or can have different geometries with different dimensions.

As depicted in Figures 8a and 8b, when a plurality of regular or irregular unit cells are arranged as a tessellation according to the present disclosure, multiple unit cells 840 can be grouped into larger, mother cells 850. The mother cells 850 are also arranged in a tessellation such that the boundaries of any mother cell 850 are directly adjacent the boundary of a neighboring mother cell. In contrast, and as depicted in 8c, certain arrangements of microstructures including four (4) or more distinct unit cell geometries include mother cells 850 that cannot be tessellated. In a different aspect, the present disclosure relates to a method of producing anti-adhesion surfaces. In such embodiments, the engineered surface 1 10 can be formed by a variety of methods, including a variety of microreplication methods, including, but not limited to, casting, coating, and/or compressing techniques. For example, microstructuring of the engineered surface 1 10 can be achieved by at least one of (1) casting a molten thermoplastic using a tool having a microstructured pattern, (2) coating of a fluid onto a tool having a microstructured pattern, solidifying the fluid, and removing the resulting film, (3) passing a thermoplastic film through a nip roll to compress against a tool having a

microstructured pattern (i.e., embossing), and/or (4) contacting a solution or dispersion of a polymer in a volatile solvent to a tool having a microstructured pattern and removing the solvent, e.g., by evaporation. The tool can be formed using any of a number of techniques known to those skilled in the art, selected depending in part upon the tool material and features of the desired topography. Illustrative techniques include etching (e.g., chemical etching, mechanical etching, or other ablative means such as laser ablation or reactive ion etching, etc., and combinations thereof), photolithography, stereolithography,

micromachining, knurling (e.g., cutting knurling or acid enhanced knurling), scoring, cutting, etc., or combinations thereof.

Alternative methods of forming the engineered surface 1 10 include thermoplastic extrusion, curable fluid coating methods, and embossing thermoplastic layers, which can also be cured. Additional information regarding the substrate material and various processes for forming the engineered surface 1 10 can be found, for example, in Halverson et al., PCT Publication No. WO 2007/070310 and US

Publication No. US 2007/0134784; Hanschen et al., US Publication No. US 2003/0235677; Graham et al., PCT Publication No. WO2004/000569; Ylitalo et al., US Patent No. 6,386,699; Johnston et al., US Publication No. US 2002/0128578 and US Patent Nos. US 6,420,622, US 6,867,342, US 7,223,364 and Scholz et al., US Patent No. 7,309,519 .

With microreplication, the engineered surface 110 can be mass produced without substantial variation from product-to-product and without using relatively complicated processing techniques. In some embodiments, microreplication can produce an engineered surface that retains an individual feature fidelity during and after manufacture, from product-to-product, that varies by no more than about 50 microns. In some embodiments, the engineered surface 1 10 retains an individual feature fidelity during and after manufacture, from product-to-product, which varies by no more than 5 microns, in some embodiments no more than 0.5 microns, and in preferred embodiments no more than 0.2 microns. In some embodiments, the engineered surface 1 10 comprises a topography that has an individual feature fidelity that is maintained with a resolution of between about 5 microns and 0.05 microns, and in some embodiments, between about 2.5 microns and 1 micron.

Alternatively, the engineered surface including a plurality of microstructures may be provided as a film and affixed to the substrate. In such embodiments, the microstructures may be made of the same or different material as the substrate. Fixation may be provided using mechanical coupling, an adhesive, a thermal process such as heat welding, ultrasonic welding, RF welding and the like, or a combination thereof.

As a final optional step, surface energy modifying coating may be applied to the microstructures. For example, a low surface energy coating may be desired. A low surface energy coating may generally be understood as a coating that, on a flat surface, has a water contact angle of greater than 1 10 degrees.

Such a coating may not be necessary to achieve highly hydrophobic performance. Exemplary low surface energy coating materials that may be used may include materials such as hexafluoropropylene oxide (HFPO), or organosilanes such as, alkylsilane, alkoxysilane, acrylsilanes, polyhedral oligomeric silsequioxane (POSS) and fluorine-containing organosilanes, just to name a few. A number of other suitable low surface energy coatings may also be used to further enhance the hydrophobicity of the film. Examples of particular coatings known in the art may be found, e.g., in US Publication No.

2008/0090010, and commonly owned publication, US Publication No. 2007/0298216. Where a coating is applied to the microstructures, it may be applied by any appropriate coating method, such as sputtering, vapor deposition, spin coating, dip coating, roll-to-roll coating, or any other number of suitable methods.

It also is possible and often preferable in order to maintain the fidelity of the microstructures to put a surface energy modifying compound in the composition used to form the microstructures. In this manner, the surface energy modifying compound is at least partially deposited on the surface of the microstructure thereby modifying the surface energy. In certain instances it may be necessary to add one or more bloom additive compounds to enhance the mobility of the surface energy modifying compound in order to get it to the surface in sufficient amount. For example, the surface energy modifying compound may have greater solubility in the bloom additive than in the base microstructure composition. In other cases, the bloom additive may retard or prevent crystallization of the base composition. Suitable bloom additives may be found, for example, in International Publication No. WO2009/152345 to Scholz et al. and US Patent No. 7,879,746 to Klun et al.

Advantages of this invention are further illustrated by the following examples, but the particular materials and amounts thereof recited in these examples, as well as other conditions and details, should not be construed to unduly limit this invention. Unless otherwise indicated, all parts and percentages are by weight.

EXAMPLES

Creating Microstructured Surfaces

To fabricate the master for a microstructure, a photoresist (PR) pattern was fabricated on a silicon wafer by optical lithography. A 1500 nm thick layer of silicon dioxide was coated onto the PR microstructures by plasma-enhanced chemical vapor deposition (PECVD), using a Model PlasmaLab System 100, available from Oxford Instruments, Yatton, UK, using the parameters listed in TABLE 1. A fast deposition rate was used in order to create surface roughness. TABLE 1. Conditions used for de ositin SiQ ₂ la er

standard cubic centimeter

The master, as prepared above, was next adhered to a stainless steel disk using double-stick tape. It was then made conductive by plating a thin layer of silver. This was followed by electroforming a nickel layer from a sulfamate nickel bath at a temperature of 54.4°C (130° F) and using a current density of 18 ampere per square foot (ASF). The thickness of the resulting nickel deposit was about 51 μηι (0.02 inch). The fidelity of the microstructure pattern was maintained through these metal plating steps. After electroforming was completed, the nickel deposit was separated from the original silicon wafer master, and used as a mold to prepare PDMS replicates.

The Ni mold was then treated with a release agent consisting of 0.1 wt % HFPO

(hexafluoropropyleneoxide) phosphonic acid delivered from a 49: 1 HFE7100:IPA solution. The HFE7100 was supplied by 3M Company of St. Paul, MN as 3M™ Novec™ 7100 Engineered Fluid, methoxy-nonafluorobutane (C4F90CH3),. The procedure for treating the mold was as follows:

1) The mold was cleaned in a plasma cleaner for 5 minutes.

2) The mold was dip coated in the 0.1 wt % HFPO release agent solvent solution.

3) The coated mold was heated in an oven for 1 hour at 120°C 4) After cooling, the mold was rinsed for 1 minute in fresh 0.1 wt % HFPO release agent solvent solution.

After this treatment, the mold was ready for use in the replication process. To prepare a polydimethyl siloxane (PDMS) replica of the microstructure, Sylgard 184 PDMS (available as SYLGARD 184 Silicone Elastomer Kit, available from Dow Corning, Midland, MI) and its curing agent were first thoroughly mixed in a 10: 1 weight ratio. Air bubbles trapped in the mixture were removed by degassing for 30 min at low vacuum. The degassed mixture was then poured onto the Ni microstructure (mold) master, further degassed for another 30 min, and then cured on a hot plate at 80°C for 1 hour. After curing, the PDMS replica was peeled off the Ni mold. High-quality microstructure replicas of the mold were produced. Figures 3a-3c and Figure 4 show the shapes and patterns of prepared microstructure surfaces. A flat "control" surface of the same material was also prepared. Microstructured Surfaces

All of the microstructured surfaces used in the examples consisted of SYLGARD 184

polydimethylsiloxane (PDMS) and were created according to the methods describe above unless otherwise noted. The following patterns tested comprised a unit cell with a single microstructure geometry and orientation.

TABLE 2. Exam le Microstructure Patterns

Preparation of S. aureus bacterial suspension

S. aureus bacteria were obtained from The American Type Culture Collection (Rockville, MD), under the trade designation ATCC 25923. The bacteria were grown overnight (17-22 hours at 37°C) in broth cultures prepared by inoculating 12 milliliters of prepared, sterile Tryptic Soy Broth (Hardy Diagnostics, Santa Maria, CA) with the bacteria.

Preparation of P. aeruginosa bacterial suspension

P. aeruginosa bacteria were obtained from The American Type Culture Collection (Rockville, MD), under the trade designation ATCC 15144. The bacteria were grown overnight (17-22 hours at 37°C) in broth cultures prepared by inoculating 12 milliliters of prepared, sterile Tryptic Soy Broth (Hardy Diagnostics, Santa Maria, CA) with the bacteria.

Static Biofilm Assay

The static biofilm assay was conducted as follows: 1. To begin the assay (Day 0), the culture tube containing the bacterial suspension as prepared above (S. aureus or P. aeruginosa) was mixed well by vortexing for 30-60 seconds to mix and suspend the bacteria. An amount of 10 mL of the incubated bacterial suspension was then added to 90 mL of sterile Tryptic Soy Broth and vortexed. The bacterial concentration of this sample was approximately lxl 0 ⁷ CFUs/mL.

2. To precisely quantify the bacterial concentration of the stock sample prepared in step 1. above, the sample was enumerated by performing serial ten-fold dilutions of the stock solution and plating on blood agar plates (Hardy Diagnostics, Santa Maria, CA) lOOuL of the -4, -5, and -6 dilutions. The plates are incubated overnight at 37°C, and the number of colony formed units were counted the next day (Day 1) to determine the exact concentration in CFUs/mL of the stock sample prepared in step 1. The colonies are visible can be manually counted.

3. Using a 1.27cm (1/2-inch) diameter punch, sample coupons of the surfaces to be tested are punched out. The samples are rinsed in isopropyl alcohol and allowed to dry in a biological hood.

4. The samples are then placed into a polycarbonate 24-well titer and identified by marking the lid of the titer plate. Each microstructure type was included in at least duplicates, or triplicates depending on the availability of the sample.

5. On "Day 0", 2mL of the bacterial suspension prepared in step 1 was added to each well of the titer plate. The plate was then incubated at 37°C overnight.

6. Every day, the titer plate(s) were removed from the incubator and placed on a rocker shaker for 1 minute. In a biological hood, the growth media was removed from each well.

7. For one set of samples (herein referred to as post-rinsed samples), 2mL of fresh TSB growth media were added to each well using a repeat pipette.

8. For a duplicate set of samples (herein referred to as "rinsed" samples), 2mL of sterile water were added to each well. A pipette was then used to rinse the samples by withdrawing and expelling the rinse water 5-6 times for each sample, and finally removing the rinse water. This wash procedure was repeated two more times for each sample, using fresh sterile water, resulting in a total of 3 washes per sample. Finally, 2mL of fresh TSB growth media was added to each well using a repeat pipette.

9. The titer plate(s) where then placed on the rocker shaker for 1 minute, and returned to the incubator overnight.

10. On days 7 and 14, samples were removed for analysis. For all samples removed ("rinsed" and "post-rinsed"), a total of 3 washes using 2 mL of sterile water were performed according to the procedure described in step 8, above.

1 1. The removed and washed samples were placed into a new 24-well titer plate and allowed to dry.

12. Once dry, the samples were stained according to the Gram-Staining Protocol, below. Gram- Staining Protocol

Samples from the Static Biofilm Assay described above were stained in preparation for analysis by microscopy using the following protocol:

Fixing Samples:

1. The sample were completely air-dried prior to fixing.

2. A large beaker was filled with methanol (Alfa Aesar, Stock #19393) and using a pair of tweezers, each sample was submerged completely for 1 minute and 10 seconds.

3. The sample was removed from methanol and allowed to air dry on a clean surface. Gram-Staining S. aureus Samples

1. A 30 mL beaker was filled with approximately 8mL of Gram crystal violet stain (Becton Dickson Co.).

2. A second beaker was filled with approximately 8mL of stabilized Gram iodine (Becton Dickson Co.

Franklin Lakes, NJ).

3. A third beaker was filled with approximately 8mL of Gram decolorizer (Becton Dickson Co.).

4. Using a pair of tweezers, the sample was first submerged completely into the Gram crystal violet stain for exactly 1 minute. After 1 minute, the sample was rinsed thoroughly under a slow and gentle running stream of deionized water.

5. The sample was then submerged completely in the stabilized Gram iodine for exactly 1 minute, followed by a rinse in deionized water as described above.

6. The sample was finally submerged completely in the Gram decolorizer for exactly 10 seconds, followed by a final rinse in deionized water.

7. Excess water was removed by blotting and the sample was allowed to air-dry on a clean surface. Gram-Staining P. aeruginosa Samples

1. A 30mL beaker was filled with approximately 8mL of Gram safrin stain (Becton Dickson Co.).

2. Using a pair of tweezers, the sample was first submerged completely into the Gram safrin stain for exactly 2 minutes. After 2 minutes, the sample was rinsed thoroughly under a slow and gentle running stream of deionized water.

3. Excess water was removed, and the sample was allowed to air-dry on a clean surface.

Analysis of Sample Surfaces for Biofilm Formation

Determination of the surface area covered by a given organism for the samples resulting from the Static

Biofilm Assay(above) was conducted by analyzing at least five micrographs for each sample. The micrographs were obtained using a Leica microscope (Model DM4000B, available from Leica

Microsystems Inc., Bannockburn, IL) outfitted with a CCD camera. The micrographs used for analysis were taken at a magnification of 40x. At this magnification (40x), the field of view for the microscope setup was 345 μηι x 290 μηι. The fractional area of the surface covered by bacteria was determined using SigmaScan software (available from Systat Software Inc., San Jose, CA). This image analysis process involved taking a given micrograph, converting the image to a grayscale version, deriving an intensity histogram of the image, and setting an intensity threshold on the histogram to isolate the stained bacterial cells from the rest of the image. The software was then capable of automatically calculating the total pixel area of the image that comprises the stained cells. That area was then divided by the pixel size of the entire image to arrive at the fractional area covered by the bacteria. For each sample, a total of at least 5 fields (micrographs) were analyzed in this manner. The std. dev. comes from the analysis of the 5 micrographs times the number of replicates. The data from each field was used to calculate an average value of the fractional area covered by the bacteria as well as the 1σ standard deviation (error).

TABLE 3. % Surface Coverage by P. aeruginosa

TABLE 4. % Surface Covera e b S. aureus

TABLE 5. Rinsin Effect on % Surface Covera e b P. aeru inosa 14 da s

TABLE 6. Rinsin Effect on % Surface Covera e b S. aureus - 14 da s

Embodiments

1. A medical article including a surface for reducing bacterial adhesion, the article comprising: an engineered surface comprising a thermoplastic or thermoset material; a plurality of engineered microstructures on at least a portion of the surface, wherein each microstructure of the plurality of microstructures comprises a base having at least one microscale cross-sectional dimension, wherein the aspect ratio of each microstructure is at least 0.5 and no greater than 10, wherein each microstructure of the plurality of engineered microstructures comprises a height greater than 0.5 microns, wherein the pitch between adjacent microstructures of the plurality of engineered microstructures is at least 1 time and no greater than 5 times than the smallest cross-sectional dimension, and wherein no base comprises a cross- sectional dimension greater than 5 microns, and wherein the arrangement of the microstructures on at least a portion of the engineered surface includes a plurality of unit cells, wherein each unit cell of the plurality of unit cells is at least partially defined by a dimension at least approximating the pitch and includes no more than one microstructure, wherein the plurality of unit cells are tiled, and wherein the plurality of unit cells includes no more than three unique unit cell geometries.

2. The medical article of embodiment 1, wherein the colonization of a target organism over 14 days as tested via Assay 1 on the portion of the engineered surface comprising the plurality of engineered microstructures is at least 50% less than the colonization on a flat surface comprised of the same biocompatible material. 3. The medical article of embodiment 1, wherein the engineered surface comprises at least

4,000,000 and no greater than 50,000,000 microstructures per square centimeter of surface.

4. The medical article of embodiment 1, wherein the plurality of unit cells comprises one type of unit cell geometry that repeats regularly over the engineered surface.

5. The medical article of any of the preceding embodiments, wherein the microstructure is selected from the group consisting of a post, a pyramid, a rib, a diamond, a dome, and combinations thereof.

6. The medical article of any of preceding embodiments, wherein each microstructure comprises the same thermoplastic or thermoset material as the engineered surface.

7. The medical article of embodiment 1 or 6, wherein each microstructure of the plurality of microstructures comprises an elastomeric microstructure.

8. The medical article of embodiments 1-7, wherein the plurality of microstructures protrude from the engineered surface.

9. The medical article of embodiments 1-7, wherein the plurality of microstructures are projected into the engineered surface.

10. The medical article of embodiment 9, wherein the plurality of microstructures comprise a plurality of discontinuous recesses.

1 1. The medical article of embodiments 1-10, wherein the plurality of unit cells includes no more than two unit cells having unique geometries.

12. The medical article of embodiment 1-1 1, wherein the geometry of any unit cell of the plurality of unit cells is substantially identical to at least all neighboring unit cells.

13. The medical article of embodiments 1-12, wherein the cross section comprises a Euclidean geometrical shape.

14. The medical article of embodiments 1- 13, wherein the colonization of S. aureus and P.

aeruginosa on the portion of the engineered surface comprising the plurality of engineered

microstructures is significantly reduced according to the Static Biofilm Assay compared to the colonization on a flat surface comprised of the same biocompatible material.

15. The medical article of embodiment 1-14, wherein the engineered surface is on at least a portion of a urinary catheter.

16. The medical article of embodiment 15, wherein the engineered surfaces is on at least a portion of the exterior of a urinary catheter.

17. The medical article of embodiment 15 wherein the engineered surfaces is on at least a portion of the interior of a urinary catheter.

18. The medical article of embodiment 15 wherein the engineered surfaces is on at least a portion of both the interior and the exterior of a urinary catheter

19. The medical article of embodiments 1-14, wherein the medical article is a wound dressing, wound absorbent, or wound contact layer. 20. The medical article of embodiments 1-19, wherein each microstructure has a base and the largest cross-sectional dimension of the base is at least 1 micron and no greater than 2 microns.

21. The medical article of embodiments 1 -20, wherein at least a portion of the engineered surface comprises an antimicrobial in or on a microstructure.

22. A method of controlling microorganism adhesion to a medical article, the method comprising: providing a medical article having a surface comprising a polymeric material and a plurality of engineered microstructures on at least a portion of the surface, wherein each microstructure of the plurality of microstructures comprises a base having at least one cross sectional microscale dimension, wherein the aspect ratio of each microstructure is at least 0.5 and no greater than 10, wherein the pitch between adjacent microstructures of the plurality of engineered microstructures is at least 1 time and no greater than 5 times than the smallest cross-sectional dimension, wherein no base comprises a cross- sectional dimension greater than 5 microns, and wherein the arrangement of the microstructures on at least a portion of the surface includes a plurality of unit cells, wherein each unit cell of the plurality of unit cells is at least partially defined by a dimension at least approximating the pitch, and wherein each unit cell comprises a boundary and each unit cell is directly adjacent the boundary of the nearest unit cell and wherein the plurality of unit cells includes no more than three unique unit cell geometries; and

placing the surface proximate a tissue or fluid, wherein the colonization of a target

microorganism on the portion of the surface comprising the plurality of engineered microstructures is reduced in comparison to a flat surface comprised of the same material.

23. The method of embodiment 22, wherein reducing the colonization of a target microorganism comprises reducing the colonization of S. aureus and P. aeruginosa by at least 50% over 14 days compared to the colonization on a flat surface comprised of the same biocompatible polymeric material according to the Static Biofilm Assay.

24. The method of embodiments 22-23, wherein each unit cell includes no more than one microstructure.

25. The method of embodiments 22-24, wherein the plurality of engineered microstructures are secured to the plurality of engineered microstructures to the engineered surface.

26. The method of embodiments 22-25, wherein the plurality of engineered microstructures are replicated directly onto the engineered surface.

27. The method of embodiments 22-26, wherein the microstructure is selected from the group consisting of a post, a pyramid, a rib, a diamond, a dome and combinations thereof.

28. The method of embodiments 22-27, wherein the microstructures comprise a plurality of protrusions from the surface.

29. The method of embodiments 22-27, wherein the microstructures comprise a plurality of discrete recesses in the surface.

30. The method of embodiment 29, wherein the plurality of discrete recesses comprises a plurality of discontinuous channels. 31. The method of embodiment 28, wherein creating the plurality of protrusions are created by microreplication.

32. The method of embodiment 22-31, wherein the engineered surface comprises a first polymeric material and wherein the plurality of engineered microstructures comprise a second polymeric material, and wherein the first and second material are the same.

33. The method any of the preceding embodiments, wherein the plurality of microstructures comprises a plurality of elastomeric microstructures.

34. The method of any of the preceding embodiments, wherein the medical article is an implantable or indwelling device.

35. The method of any of the preceding embodiments, wherein the medical article is a wound dressing, wound contact layer, or wound absorbent.

36. A medical article including a surface for reducing bacterial adhesion, the article comprising: an engineered surface comprising a thermoplastic or thermoset material;

a plurality of engineered microstructures on at least a portion of the surface, wherein each microstructure of the plurality of microstructures comprises a base having at least one microscale cross- sectional dimension, wherein the engineered surface comprises at least 4,000,000 and no greater than 50,000,000 microstructures per square centimeter of surface,

wherein the aspect ratio of each microstructure is at least 0.5 and no greater than 10,

wherein each microstructure of the plurality of engineered microstructures comprises a height greater than 0.5 microns, wherein no microstructure comprises a base cross-sectional dimension greater than 5 microns,

wherein the arrangement of the microstructures on at least a portion of the engineered surface includes a plurality of unit cells, each unit cell comprising a single microstructure and a peripheral boundary, wherein the unit cells are arranged in a tessellation, and wherein the plurality of unit cells includes no more than three unique unit cell geometries.

37. A method of making a biofilm adhesion resistant medical article, the method comprising:

providing a medical article having a surface comprising a polymeric material; and

creating a plurality of engineered microstructures on at least a portion of the surface, a plurality of engineered microstructures on at least a portion of the surface, wherein each microstructure of the plurality of microstructures comprises a base having at least one cross sectional microscale dimension, wherein the aspect ratio of each microstructure is at least 0.5 and no greater than 10, wherein the pitch between adjacent microstructures of the plurality of engineered microstructures is at least 1 time and no greater than 5 times than the smallest cross-sectional dimension, wherein no base of any microstructure comprises a cross-sectional dimension greater than 5 microns, and wherein the arrangement of the microstructures on at least a portion of the surface includes a plurality of unit cells, wherein each unit cell of the plurality of unit is at least partially defined by a dimension at least approximating the pitch, and wherein the plurality of unit cells includes no more than three unique unit cell geometries, and wherein the plurality of unit cells are tiled.

38. A medical article including a surface having reduced bacterial adhesion, the article comprising: a engineered surface comprising a thermoplastic or thermoset material;

a plurality of engineered microstructures on at least a portion of the engineered surface, wherein each microstructure of the plurality of microstructures comprises a base having at least one cross sectional dimension, wherein the largest cross-sectional dimension of the base is at least 1 micron and no greater than 2 microns,

wherein the aspect ratio of each microstructure is at least 0.5 and no greater than 10,

wherein the spacing between adjacent microstructures of the plurality of engineered

microstructures is at least 1 time and no greater than 5 times than the smallest cross-sectional dimension, and

wherein the arrangement of the microstructures on at least a portion of the engineered surface includes a plurality of tiled unit cells, each unit cell having the same or substantially same geometry and including no more than one microstructure.

The complete disclosures of the patents, patent documents, and publications cited herein are incorporated by reference in their entirety as if each were individually incorporated. Various

modifications and alterations to this invention will become apparent to those skilled in the art without departing from the scope and spirit of this invention. It should be understood that this invention is not intended to be unduly limited by the illustrative embodiments and examples set forth herein and that such examples and embodiments are presented by way of example only with the scope of the invention intended to be limited only by the claims set forth herein as follows.