Biofilms in drug-coated medical devices: Mechanisms of antimicrobial resistance and perspectives on new approaches

Figures

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  Figure 1. Biofilm development in five stages. The development of bacterial biofilms is stereotyped by a 72+ hour’s developmental program. Stage 1 (0–2 h): surface binding due to van der Waals forces and hydrophobic interactions, which is reversible. Stage 1 (0–2 h): Surface binding is reversible, via van der Waals forces and hydrophobic interaction. Stage 2 (2–6 h): is irreversible, includes up-regulation of adhesins and fimbriae. Stage 3 (6–24 h): is the stage of the development of microcolonies, as bacteria divide actively and exopolysaccharides are produced in great amounts. Stage 4 (24–72 h): complete biofilm organism – tower and mushroom shape; water channels are formed to distribute nutrients. Stage 5 (>72 h): dispersal stage, produces motile and chemotactically active cells, stores metabolically dormant persister cells.

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  Figure 2. Quorum sensing mechanism. Bacterial communication through density-dependent signals leads to coordinated multicellular behavior in bacterial biofilms. Panel 1: Single bacteria generate small autoinducer molecules (N-3-oxododecanoyl-L-homoserine lactone when Las system is involved; N-butyryl-L-homoserine lactone when Rhl system is involved) which are small in size and diffusive. Autoinducers: Panel 2: There is an increase in accumulation of autoinducers with cell density in the biofilm microenvironment. If the autoinducers are allowed to reach a threshold concentration, then the cognate regulatory proteins (LasR, RhlR) will bind to them and will lead to a signal transduction cascade. Panel 4: Coordinated expression of biofilm-specific genes including virulence factors, EPS production, and, most importantly, genes encoding antimicrobial resistance determinants, which provide a direct molecular link between biofilm formation and the resistance phenotype.

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  Figure 3. Antimicrobial resistance through biofilm. Based on a 10–1,000-fold increase in resistance, this is achieved by a combination of six synergistic complementary mechanisms. Panel A: Extracellular polysaccharide matrix forms a physical barrier, which is only partially permeable to charge and hydrophobicity; negatively charged polymers bind cationic antibiotics (aminoglycosides, polymyxins) and inhibit penetration by about 1,000 times. Panel B: Oxygen concentration gradients developed in biofilm having aerobic outer and hypoxia-anaerobic inner layers. In anaerobic environments, microorganisms produce the alarmone (p)ppGpp, leading to the production of metabolically dormant persister cells that are resistant to antibiotics that require metabolism. Panel D: Biofilm-associated enzymes (β-lactamases, aminoglycoside-modifying enzymes) form degradation areas, destroying penetrating antibiotics. Biofilms upregulate expression of the efflux pumps (RND family, ABC transporters) which expel intracellular antibiotics. Resistance determinants are spread rapidly by horizontal gene transfer through the process of conjugation (Panel F) and by phage-mediated transduction (Panel G) (10–1,000-fold…

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  Figure 4. Left panels depict a traditional antibiotic-eluting device. Panel A: Surface adsorption approach with direct antibiotic coating with rapid elution (minocycline/rifampin systems); the mechanism generates high concentrations at the beginning but is rapidly depleted. Panel B: Surface erosion and bulk degradation of individual particles of polymer matrix (PLGA, PLA, PCL) that encapsulate antibiotics and exhibit sustained release over weeks. Panel C: Kinetics of release: burst release curve (surface adsorption) and sustained release profile (polymer matrix). The future approaches are shown in the right panels. Panel D: Polymers that are stimuli-responsive, releasing drugs in response to changes in pH (inflammatory microenvironment) or temperature. Panel E: Enzyme-immobilized surfaces and coatings that enable the enzyme (alginate lyase enzyme or proteases) to be bound to the surface to enable the degradation of the biofilm matrix and penetration of the antimicrobial. Panel F: Intelligent implants: Contain inbuilt biosensors, which can sense the quorum sensing molecules and only be able to release antimicrobials when the formation of biofilm begins.

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  Figure 5. New Advanced Biofilm Prevention technologies. Second-generation strategies combine a number of complementary mechanisms. Panel A: Bacteriophage-based therapy using depolymerases produced by the bacteriophage to break down the polysaccharides in the biofilm matrix, allowing antibiotics to penetrate mature biofilms; specificity of the bacteriophage prevents destruction of commensal microbiota. Panel B: Intelligent implants containing biofilm sensing, locally embedded biosensors which release increased amounts of antimicrobial agents on demand when transmitting the presence of active biofilm growth (bacterial quorum sensing molecules: N-acyl-homoserine lactones, autoinducer-2). The first panel has stimuli-responsive polymers which can provide temporal-spatial control: Panel C: pH-responsive release in the acid inflammatory microenvironment, temperature-responsive poly (N-isopropylacrylamide) triggers a phase transition in body temperature, light-responsive coatings provide external control. Panel D: Enzyme-immobilized surfaces with enzyme immobilized on surface which has alginate lyase from Pseudomonas and protease to matrix proteins which inhibited the formation of biofilms by…