Co-immobilization of Ciprofloxacin and Chlorhexidine as a Broad-Spectrum Antimicrobial Dual-Drug Coating for Poly(vinyl chloride) (PVC)-Based Endotracheal Tubes.

The endotracheal tube (ETT) affords support for intubated patients, but the increasing incidence of ventilator-associated pneumonia (VAP) is jeopardizing its application. ETT surfaces promote (poly)microbial colonization and biofilm formation, with a heavy burden for VAP. Devising safe, broad-spectrum antimicrobial materials to tackle the ETT bioburden is needful. Herein, we immobilized ciprofloxacin (CIP) and/or chlorhexidine (CHX), through polydopamine (pDA)-based functionalization, onto poly(vinyl chloride) (PVC) surfaces. These surfaces were characterized regarding physicochemical properties and challenged with single and polymicrobial cultures of VAP-relevant bacteria (Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae, Staphylococcus aureus, Staphylococcus epidermidis) and fungi (Candida albicans). The coatings imparted PVC surfaces with a homogeneous morphology, varied wettability, and low roughness. The antimicrobial immobilization via pDA chemistry was still evidenced by infrared spectroscopy. Coated surfaces exhibited sustained CIP/CHX release, retaining prolonged (10 days) activity. CIP/CHX-coated surfaces evidencing no A549 lung cell toxicity displayed better antibiofilm outcomes than CIP or CHX coatings, preventing bacterial attachment by 4.1-7.2 Log10 CFU/mL and modestly distressingC. albicans. Their antibiofilm effectiveness was endured toward polymicrobial consortia, substantially inhibiting the adhesion of the bacterial populations (up to 8 Log10 CFU/mL) within the consortia in dual- and even inP. aeruginosa/S. aureus/C. albicans triple-species biofilms while affecting fungal adhesion by 2.7 Log10 CFU/mL (dual consortia) and 1 Log10 CFU/mL (triple consortia). The potential of the dual-drug coating strategy in preventing triple-species adhesion and impairing bacterial viability was still strengthened by live/dead microscopy. The pDA-assisted CIP/CHX co-immobilization holds a safe and robust broad-spectrum antimicrobial coating strategy for PVC-ETTs, with the promise laying in reducing VAP incidence.

Aptamer-based therapy for fighting biofilm-associated infections.

Biofilms are key players in the pathogenesis of most of chronic infections associated with host tissue or fluids and indwelling medical devices. These chronic infections are hard to be treated due to the increased biofilms tolerance towards antibiotics in comparison to planktonic (or free living) cells. Despite the advanced understanding of their formation and physiology, biofilms continue to be a challenge and there is no standardized therapeutic approach in clinical practice to eradicate them. Aptamers offer distinctive properties, including excellent affinity, selectivity, stability, making them valuable tools for therapeutic purposes. This review explores the flexibility and designability of aptamers as antibiofilm drugs but, importantly, as targeting tools for diverse drug and delivery systems. It highlights specific examples of application of aptamers in biofilms of diverse species according to different modes of action including inhibition of motility and adhesion, blocking of quorum sensing molecules, and dispersal of biofilm-cells to planktonic state. Moreover, it discusses the limitations and challenges that impaired an increased success of the use of aptamers on biofilm management, as well as the opportunities related to aptamers modifications that can significantly expand their applicability on the biofilm field.

Effect of ginger essential oil and 6-gingerol on a multispecies biofilm of Listeria monocytogenes, Salmonella Typhimurium, and Pseudomonas aeruginosa.

The objective of this study was to evaluate the potential antimicrobial and antibiofilm effect of ginger essential oil (GEO) and 6-gingerol on a multispecies biofilm formed by Listeria monocytogenes, Salmonella Typhimurium, and Pseudomonas aeruginosa on a polypropylene surface. The minimum inhibitory concentration concentrations obtained for GEO were 100 and 50 mg/mL and for 6-gingerol 1.25 mg/mL. Sessile cell counts ranged within 5.35-7.35 log CFU/cm2 in the control biofilm, with the highest sessile growth at 72 h. GEO treatments acted on the total population regardless of concentration at 1 and 48 h. L. monocytogenes behaved similarly to the total population, showing GEO action at 1 h and keeping the same pattern at 48, 72, and 96 h. Better action on S. Typhimurium was obtained at times of 1, 72, and 96 h. P. aeruginosa showed logarithmic reduction only when treated with GEO 50 mg at 24 h. As for 6-gingerol, in general, there was no significant action (p > 0.05) on the evaluated sessile cells. GEO showed antimicrobial activity against L. monocytogenes, S. Typhimurium, and P. aeruginosa, acting as an inhibitor of biofilm formation. As for 6-gingerol, it was considered a possible antimicrobial agent but without efficacy during biofilm formation.

Evolving biofilm inhibition and eradication in clinical settings through plant-based antibiofilm agents.

BACKGROUND: After almost 100 years since evidence of biofilm mode of growth and decades of intensive investigation about their formation, regulatory pathways and mechanisms of antimicrobial tolerance, nowadays there are still no therapeutic solutions to eradicate bacterial biofilms and their biomedical related issues. PURPOSE: This review intends to provide a comprehensive summary of the recent and most relevant published studies on plant-based products, or their isolated compounds with antibiofilm activity mechanisms of action or identified molecular targets against bacterial biofilms. The objective is to offer a new perspective of most recent data for clinical researchers aiming to prevent or eliminate biofilm-associated infections caused by bacterial pathogens. METHODS: The search was performed considering original research articles published on PubMed, Web of Science and Scopus from 2015 to April 2023, using keywords such as "antibiofilm", "antivirulence", "phytochemicals" and "plant extracts". RESULTS: Over 180 articles were considered for this review with a focus on the priority human pathogens listed by World Health Organization, including Pseudomonas aeruginosa, Staphylococcus aureus, Klebsiella pneumoniae and Escherichia coli. Inhibition and detachment or dismantling of biofilms formed by these pathogens were found using plant-based extract/products or derivative compounds. Although combination of plant-based products and antibiotics were recorded and discussed, this topic is currently poorly explored and only for a reduced number of bacterial species. CONCLUSIONS: This review clearly demonstrates that plant-based products or derivative compounds may be a promising therapeutic strategy to eliminate bacterial biofilms and their associated infections. After thoroughly reviewing the vast amount of research carried out over years, it was concluded that plant-based products are mostly able to prevent biofilm formation through inhibition of quorum sensing signals, but also to disrupt mature biofilms developed by multidrug resistant bacteria targeting the biofilm extracellular polymeric substance. Flavonoids and phenolic compounds seemed the most effective against bacterial biofilms.

Antimicrobial materials for endotracheal tubes: A review on the last two decades of technological progress.

Ventilator-associated pneumonia (VAP) is an unresolved problem in nosocomial settings, remaining consistently associated with a lack of treatment, high mortality, and prolonged hospital stay. The endotracheal tube (ETT) is the major culprit for VAP development owing to its early surface microbial colonization and biofilm formation by multiple pathogens, both critical events for VAP pathogenesis and relapses. To combat this matter, gradual research on antimicrobial ETT surface coating/modification approaches has been made. This review provides an overview of the relevance and implications of the ETT bioburden for VAP pathogenesis and how technological research on antimicrobial materials for ETTs has evolved. Firstly, certain main VAP attributes (definition/categorization; outcomes; economic impact) were outlined, highlighting the issues in defining/diagnosing VAP that often difficult VAP early- and late-onset differentiation, and that generate misinterpretations in VAP surveillance and discrepant outcomes. The central role of the ETT microbial colonization and subsequent biofilm formation as fundamental contributors to VAP pathogenesis was then underscored, in parallel with the uncovering of the polymicrobial ecosystem of VAP-related infections. Secondly, the latest technological developments (reported since 2002) on materials able to endow the ETT surface with active antimicrobial and/or passive antifouling properties were annotated, being further subject to critical scrutiny concerning their potentialities and/or constraints in reducing ETT bioburden and the risk of VAP while retaining/improving the safety of use. Taking those gaps/challenges into consideration, we discussed potential avenues that may assist upcoming advances in the field to tackle VAP rampant rates and improve patient care. STATEMENT OF SIGNIFICANCE: The use of the endotracheal tube (ETT) in patients requiring mechanical ventilation is associated with the development of ventilator-associated pneumonia (VAP). Its rapid surface colonization and biofilm formation are critical events for VAP pathogenesis and relapses. This review provides a comprehensive overview on the relevance/implications of the ETT biofilm in VAP, and on how research on antimicrobial ETT surface coating/modification technology has evolved over the last two decades. Despite significant technological advances, the limited number of gathered reports (46), highlights difficulty in overcoming certain hurdles associated with VAP (e.g., persistent colonization/biofilm formation; mechanical ventilation duration; hospital length of stay; VAP occurrence), which makes this an evolving, complex, and challenging matter. Challenges and opportunities in the field are discussed.

Colistin conditioning surfaces combined with antimicrobial treatment to prevent ventilator-associated infections.

Biofilm formation on endotracheal tubes (ETT) is an important factor in the development of ventilator-associated pneumonia (VAP). This work aimed to investigate the effectiveness of colistin (COL) against the early stages of biofilm formation by Pseudomonas aeruginosa. Two strategies were used: pre-conditioning the adhesion surfaces with COL before biofilm formation and growing biofilms in its presence. The combined effect of treating P. aeruginosa 24-hours old biofilms with Ciprofloxacin (CIP) or colistin (COL) on clean and COL-conditioned surfaces was also assessed. Random deposition of COL residues altered the physico-chemical properties of the adhesion surfaces and impaired biofilm formation. Moreover, as a consequence of the reduced amount of biofilms attached to COL conditioned surfaces, adhered cells became more exposed to the subsequent action of CIP or COL, suggesting a combined outcome of prophylactic and therapeutic COL-based strategies. Results highlighted the promising use of COL to prevent the establishment of biofilms on ETT.

Chemical Characterization of Sambucus nigra L. Flowers Aqueous Extract and Its Biological Implications.

The main goal of this study was to chemically characterize an aqueous S. nigra flower extract and validate it as a bioactive agent. The elderflower aqueous extraction was performed at different temperatures (50, 70 and 90 °C). The extract obtained at 90 °C exhibited the highest phenolic content and antiradical activity. Therefore, this extract was analyzed by GC-MS and HPLC-MS, which allowed the identification of 46 compounds, being quercetin and chlorogenic acid derivatives representative of 86% of the total of phenolic compounds identified in hydrophilic fraction of the aqueous extract. Naringenin (27.2%) was the major compound present in the lipophilic fraction. The antiproliferative effects of the S. nigra extract were evaluated using the colon cancer cell lines RKO, HCT-116, Caco-2 and the extract's antigenotoxic potential was evaluated by the Comet assay in RKO cells. The RKO cells were the most susceptible to S. nigra flower extract (IC50 = 1250 µg mL-1). Moreover, the extract showed antimicrobial activity against Gram-positive bacteria, particularly Staphylococcus aureus and S. epidermidis. These results show that S. nigra-based extracts can be an important dietary source of bioactive phenolic compounds that contribute to health-span improving life quality, demonstrating their potential as nutraceutical, functional foods and/or cosmetic components for therapeutic purposes.

Viable but non-cultivable state: a strategy for Staphylococcus aureus survivable in dual-species biofilms with Pseudomonas aeruginosa?

Pseudomonas aeruginosa and Staphylococcus aureus are two of the most prevalent respiratory pathogens in cystic fibrosis patients. Both organisms often cause chronic and recalcitrant infections, in large part due to their ability to form biofilms, being these mixed-species infections correlated with poor clinical outcomes. In this study, the hypothesis that S. aureus adopts phenotypes allowing its coexistence with P. aeruginosa during biofilm growth was put forward. We noticed that S. aureus undergoes a viable but non-cultivable (VBNC) state in the dominated P. aeruginosa dual-species consortia, whatsoever the strains used to form the biofilms. Moreover, an increased expression of genes associated with S. aureus virulence was detected suggesting that the phenotypic switching to VBNC state might account for S. aureus pathogenicity and, in turn, influence the clinical outcome of the mixed-species infection. Thus, P. aeruginosa seems to induce both phenotypic and transcriptomic changes in S. aureus, helping its survival and coexistence in the dual-species biofilms. Overall, our findings illustrate how interspecies interactions can modulate bacterial virulence in vitro, contributing to a better understanding of the behaviour of P. aeruginosa-S. aureus dual-species biofilms.

Long-term coexistence of Pseudomonas aeruginosa and Staphylococcus aureus using an in vitro cystic fibrosis model.

Aim: To investigate the role of pre-established Staphylococcus aureus on Pseudomonas aeruginosa adaptation and antibiotic tolerance. Materials & methods: Bacteria were cultured mimicking the sequential pattern of lung colonization and exposure to ciprofloxacin. Results: In the absence of ciprofloxacin exposure, S. aureus and P. aeruginosa coexisted supported by the physicochemical characteristics of the artificial sputum medium. S. aureus had no role in P. aeruginosa tolerance against ciprofloxacin and did not select P. aeruginosa small-colony variants during antibiotic treatment. rhlR and psqE were downregulated after the contact with S. aureus indicating that P. aeruginosa attenuated its virulence potential. Conclusion:P. aeruginosa and S. aureus can cohabit in cystic fibrosis airway environment for long-term without significant impact on P. aeruginosa adaptation and antibiotic tolerance.

Tailoring the immobilization and release of chlorhexidine using dopamine chemistry to fight infections associated to orthopedic devices.

A crucial factor in the pathogenesis of orthopedics associated infections is that bacteria do not only colonize the implant surface but also the surrounding tissues. This study aimed to engineer an antimicrobial release coating for stainless steel (SS) surfaces, to impart them with the ability to prevent Staphylococci colonization. Chlorhexidine (CHX) was immobilized using two polydopamine (pDA)-based approaches: a one-pot synthesis, where CHX is dissolved together with dopamine before its polymerization; and a two-step methodology, comprising the deposition of a pDA layer to which CHX is immobilized. To modulate CHX release, an additional layer of pDA was also added for both strategies. Immobilization of CHX using a one-step approach yielded surfaces with a more homogenous coating and less roughness than the other strategies. The amount of released CHX was lower for the one-step approach, as opposed to the two-step approach yielding the higher release, which could be decreased by applying an outward layer of pDA. Both one and two-step approaches provided the surfaces with the ability to prevent bacterial colonization of the surface itself and kill most of bacteria in the bulk phase up to 10 days. This long-term antimicrobial performance alluded a stable and enduring immobilization of CHX. In terms of biocompatibility, the amount of CHX released from the one-step approach did not compromise the growth of mammalian cells, contrary to the two-step strategy. Additionally, the few bacteria that managed to adhere to surfaces modified with one-step approach did not show evidence of resistance towards CHX. Overall data underline that one-step immobilization of CHX holds great potential to be further applied in the fight against orthopedic devices associated infections.

Discerning the role of polymicrobial biofilms in the ascent, prevalence, and extent of heteroresistance in clinical practice.

Antimicrobial therapy is facing a worrisome and underappreciated challenge, the phenomenon of heteroresistance (HR). HR has been gradually documented in clinically relevant pathogens (e.g. Pseudomonas aeruginosa, Staphylococcus aureus, Burkholderia spp., Acinetobacter baumannii, Klebsiella pneumoniae, Candida spp.) towards several drugs and is believed to complicate the clinical picture of chronic infections. This type of infections are typically mediated by polymicrobial biofilms, wherein microorganisms inherently display a wide range of physiological states, distinct metabolic pathways, diverging refractory levels of stress responses, and a complex network of chemical signals exchange. This review aims to provide an overview on the relevance, prevalence, and implications of HR in clinical settings. Firstly, related terminologies (e.g. resistance, tolerance, persistence), sometimes misunderstood and overlapped, were clarified. Factors generating misleading HR definitions were also uncovered. Secondly, the recent HR incidences reported in clinically relevant pathogens towards different antimicrobials were annotated. The potential mechanisms underlying such occurrences were further elucidated. Finally, the link between HR and biofilms was discussed. The focus was to recognize the presence of heterogeneous levels of resistance within most biofilms, as well as the relevance of polymicrobial biofilms in chronic infectious diseases and their role in resistance spreading. These topics were subject of a critical appraisal, gaining insights into the ascending clinical implications of HR in antimicrobial resistance spreading, which could ultimately help designing effective therapeutic options.

Unraveling Pseudomonas aeruginosa and Candida albicans Communication in Coinfection Scenarios: Insights Through Network Analysis.

Modern medicine is currently facing huge setbacks concerning infection therapeutics as microorganisms are consistently knocking down every antimicrobial wall set before them. The situation becomes more worrying when taking into account that, in both environmental and disease scenarios, microorganisms present themselves as biofilm communities that are often polymicrobial. This comprises a competitive advantage, with interactions between different species altering host responses, antimicrobial effectiveness, microbial pathogenesis and virulence, usually augmenting the severity of the infection and contributing for the recalcitrance towards conventional therapy. Pseudomonas aeruginosa and Candida albicans are two opportunistic pathogens often co-isolated from infections, mainly from mucosal tissues like the lung. Despite the billions of years of co-existence, this pair of microorganisms is a great example on how little is known about cross-kingdom interactions, particularly within the context of coinfections. Given the described scenario, this study aimed to collect, curate, and analyze all published experimental information on the molecular basis of P. aeruginosa and C. albicans interactions in biofilms, in order to shed light into key mechanisms that may affect infection prognosis, increasing this area of knowledge. Publications were optimally retrieved from PubMed and Web of Science and classified as to their relevance. Data was then systematically and manually curated, analyzed, and further reconstructed as networks. A total of 641 interactions between the two pathogens were annotated, outputting knowledge on important molecular players affecting key virulence mechanisms, such as hyphal growth, and related genes and proteins, constituting potential therapeutic targets for infections related to these bacterial-fungal consortia. Contrasting interactions were also analyzed, and quorum-sensing inhibition approaches were highlighted. All annotated data was made publicly available at www.ceb.uminho.pt/ISCTD, a database already containing similar data for P. aeruginosa and Staphylococcus aureus communication. This will allow researchers to cut on time and effort when studying this particular subject, facilitating the understanding of the basis of the inter-species and inter-kingdom interactions and how it can be modulated to help design alternative and more effective tailored therapies. Finally, data deposition will serve as base for future dataset integration, whose analysis will hopefully give insights into communications in more complex and varied biofilm communities.

Fostering Innovation in the Treatment of Chronic Polymicrobial Cystic Fibrosis-Associated Infections Exploring Aspartic Acid and Succinic Acid as Ciprofloxacin Adjuvants.

Cystic fibrosis (CF) disease provokes the accumulation of thick and viscous sputum in the lungs, favoring the development of chronic and polymicrobial infections. Pseudomonas aeruginosa is the main bacterium responsible for these chronic infections, and much of the difficulty involved in eradicating it is due to biofilm formation. However, this could be mitigated using adjuvant compounds that help or potentiate the antibiotic action. Therefore, the main goal of this study was to search for substances that function as adjuvants and also as biofilm-controlling compounds, preventing or dismantling P. aeruginosa biofilms formed in an in vitro CF airway environment. Dual combinations of compounds with subinhibitory (1 and 2 mg/L) and inhibitory concentrations (4 mg/L) of ciprofloxacin were tested to inhibit the bacterial growth and biofilm formation (prophylactic approach) and to eradicate 24-h-old P. aeruginosa populations, including planktonic cells and biofilms (treatment approach). Our results revealed that aspartic acid (Asp) and succinic acid (Suc) restored ciprofloxacin action against P. aeruginosa. Suc combined with 2 mg/L of ciprofloxacin (Suc-Cip) was able to eradicate bacteria, and Asp combined with 4 mg/L of ciprofloxacin (Asp-Cip) seemed to eradicate the whole 24-h-old populations, including planktonic cells and biofilms. Based on biomass depletion data, we noted that Asp induced cell death and Suc seemed somehow to block or reduce the expression of ciprofloxacin resistance. As far as we know, this kind of action had not been reported up till now. The presence of Staphylococcus aureus and Burkholderia cenocepacia did not affect the efficacy of the Asp-Cip and Suc-Cip therapies against P. aeruginosa and, also important, P. aeruginosa depletion from polymicrobial communities did not create a window of opportunity for these species to thrive. Rather the contrary, Asp and Suc also improved ciprofloxacin action against B. cenocepacia. Further studies on the cytotoxicity using lung epithelial cells indicated toxicity of Suc-Cip caused by the Suc. In conclusion, we provided evidences that Asp and Suc could be potential ciprofloxacin adjuvants to eradicate P. aeruginosa living within polymicrobial communities. Asp-Cip and Suc-Cip could be promising therapeutic options to cope with CF treatment failures.

Minimum information guideline for spectrophotometric and fluorometric methods to assess biofilm formation in microplates.

The lack of reproducibility of published studies is one of the major issues facing the scientific community, and the field of biofilm microbiology has been no exception. One effective strategy against this multifaceted problem is the use of minimum information guidelines. This strategy provides a guide for authors and reviewers on the necessary information that a manuscript should include for the experiments in a study to be clearly interpreted and independently reproduced. As a result of several discussions between international groups working in the area of biofilms, we present a guideline for the spectrophotometric and fluorometric assessment of biofilm formation in microplates. This guideline has been divided into 5 main sections, each presenting a comprehensive set of recommendations. The intention of the minimum information guideline is to improve the quality of scientific communication that will augment interlaboratory reproducibility in biofilm microplate assays.

Pseudomonas aeruginosa and Staphylococcus aureus communication in biofilm infections: insights through network and database construction.

The polymicrobial nature of most infections is often characterized by complex biofilm communities, where pathogen interactions promote infection progression and severity. Quorum-sensing, the major regulator of virulence and inter-species communication, is a promising target for new anti-infective strategies. This study aimed at collecting and analysing experimental information on the molecular basis of Pseudomonas aeruginosa and Staphylococcus aureus interactions in biofilms. Data were systematically annotated from relevant full-text papers optimally retrieved from PubMed, reconstructed as networks and integrated with specialized databases to identify promising antimicrobial targets. Network analysis revealed key entities regulating P. aeruginosa/S. aureus interactions, for instance the PqsABCDE/PqsR quorum-sensing system, which affects S. aureus growth and biofilm formation. By identifying the most reported P. aeruginosa virulence factors affecting S. aureus, for example, HQNO and siderophores, a list of experimentally validated agents affecting those factors, ranging from synthetic drugs to natural plant extracts, was constructed. The complex experimental data on P. aeruginosa/S. aureus interactions were for the first time systematically organized and made publically available in the new Inter-Species CrossTalk Database (www.ceb.uminho.pt/ISCTD).

RNA-based qPCR as a tool to quantify and to characterize dual-species biofilms.

While considerable research has focused on studying individual-species, we now face the challenge of determining how interspecies interactions alter bacterial behaviours and pathogenesis. Pseudomonas aeruginosa and Staphylococcus aureus are often found to co-infect cystic-fibrosis patients. Curiously, their interaction is reported as competitive under laboratory conditions. Selecting appropriate methodologies is therefore critical to analyse multi-species communities. Herein, we demonstrated the major biases associated with qPCR quantification of bacterial populations and optimized a RNA-based qPCR able not only to quantify but also to characterize microbial interactions within dual-species biofilms composed by P. aeruginosa and S. aureus, as assessed by gene expression quantification. qPCR quantification was compared with flow-cytometry and culture-based quantification. Discrepancies between culture independent and culture dependent methods could be the result of the presence of viable but not-cultivable bacteria within the biofilm. Fluorescence microscopy confirmed this. A higher sensitivity to detect viable cells further highlights the potentialities of qPCR approach to quantify biofilm communities. By using bacterial RNA and an exogenous mRNA control, it was also possible to characterize bacterial transcriptomic profile, being this a major advantage of this method.

Antimicrobial resistance three ways: healthcare crisis, major concepts and the relevance of biofilms.

Worldwide, infections are resuming their role as highly effective killing diseases, as current treatments are failing to respond to the growing problem of antimicrobial resistance (AMR). The social and economic burden of AMR seems ever rising, with health- and research-related organizations rushing to collaborate on a worldwide scale to find effective solutions. Resistant bacteria are spreading even in first-world nations, being found not only in healthcare-related settings, but also in food and in the environment. In this minireview, the impact of AMR in healthcare systems and the major bacteria behind it are highlighted. Ecological aspects of AMR evolution and the complexity of its molecular mechanisms are explained. Major concepts, such as intrinsic, acquired and adaptive resistance, as well as tolerance and heteroresistance, are also clarified. More importantly, the problematic of biofilms and their role in AMR, namely their main resistance and tolerance mechanisms, are elucidated. Finally, some of the most promising anti-biofilm strategies being investigated are reviewed. Much is still to be done regarding the study of AMR and the discovery of new anti-biofilm strategies. Gladly, considerable research on this topic is generated every day and increasingly concerted actions are being engaged globally to try and tackle this problem.

Design of an Antifungal Surface Embedding Liposomal Amphotericin B Through a Mussel Adhesive-Inspired Coating Strategy.

Microbial colonization of urinary catheters remains a serious problem for medicine as it often leads to biofilm formation and infection. Among the approaches reported to deal with this problem, surfaces functionalization to render them with antimicrobial characteristics, comprises the most promising one. Most of these strategies, however, are designed to target bacterial biofilms, while fungal biofilms are much less taken into account. In real-life settings, fungi will be inevitably found in consortium with bacteria, especially in the field of biomaterials. The development of antifungal coating strategies to be combined with antibacterial approaches will be pivotal for the fight of biomaterial-associated infections. The main goal of the present study was, therefore, to engineer an effective strategy for the immobilization of liposomal amphotericin B (LAmB) on polydimethylsiloxane (PDMS) surfaces to prevent Candida albicans colonization. Immobilization was performed using a two-step mussel-inspired coating strategy, in which PDMS is first immersed in dopamine solution. Its polymerization results in the deposition of a thin adherent film, called polydopamine (pDA), which allowed the incorporation of LAmB, afterwards. Different concentrations of LAmB were screened in order to obtain a contact-killing surface with no release of LAmB. Surface characterization confirmed the polymerization of dopamine and further functionalization with LAmB yielded surfaces with less roughness and more hydrophilic features. The proposed coating strategy rendered the surfaces of PDMS with the ability to prevent the attachment of C. albicans and kill the adherent cells, without toxicity toward mammalian cells. Overall results showed that LAmB immobilization on a surface retained its antifungal activity and reduced toxicity, holding therefore a great potential to be applied for the design of urinary catheters. Since the sessile communities commonly found associated to these devices exhibit a polymicrobial nature, the next challenge will be to co-immobilize LAmB with antibacterial agents to prevent the establishment of catheter-associated urinary tract infections (CAUTI).