Bram Neirinck

The Nonsteroidal Antiinflammatory Drug Diclofenac Potentiates the In Vivo Activity of Caspofungin Against Candida albicans Biofilms

In this study, we demonstrated that in vitro Candida albicans biofilms grown in the presence of diclofenac showed increased susceptibility to caspofungin. These findings were further confirmed using a catheter-associated biofilm model in rats. C. albicans-inoculated catheters retrieved from rats that were treated with both diclofenac and caspofungin contained significantly fewer biofilm cells and showed no visible biofilms inside the catheter lumens, as documented by scanning electron microscopy, as compared to catheters retrieved from rats receiving only caspofungin or diclofenac. This report indicates that diclofenac could be useful in combination therapy with caspofungin to treat C. albicans biofilm-associated infections.

A Current Opinion on Electrophoretic Deposition in Pulsed and Alternating Fields

Electrophoretic deposition (EPD) is a colloidal production process developed in the early 20th century. Industrial scale EPD for the production of electronic components and phosphorescent screens and in the form of cataphoretic painting has known some success. Despite its limited practical applications, the inherent versatility of EPD has never ceased to fuel research into this technique. One of the major drives of this research was to render the method more environmentally friendly by enabling deposition from aqueous suspensions. One particular route, suggested to circumvent the problems caused by the use of water in EPD, is the use of alternating or pulsed fields. Recently, the use of alternating fields in EPD has been investigated for the deposition of biological matter in the form of cells and molecules. With this new avenue of research opening up and coinciding with a rise in biotechnological processes, one can expect a renewed interest in traditional EPD and fundamental research on the use of pulsed and alternating fields in this technique. Hence, this review attempts to summarize a centurys worth of both fundamental and applied research for scientists venturing into the field of EPD.

In vivo Candida glabrata biofilm development on foreign bodies in a rat subcutaneous model

OBJECTIVES Biofilm studies have been mostly dedicated to the major human fungal pathogen Candida albicans, whereas much less is known about this virulence factor in Candida glabrata, certainly under in vivo conditions. This study provides a deeper understanding of the biofilm development of C. glabrata, its architecture and susceptibility profile to fluconazole and echinocandins. METHODS In vitro and in vivo C. glabrata biofilms were developed inside serum-coated triple-lumen catheters placed in 24-well polystyrene plates or implanted subcutaneously in the back of a rat, respectively. Scanning electron microscopy and confocal scanning laser microscopy were used to visualize the biofilm architecture. Quantitative real-time PCR was used to demonstrate the expression profile of EPA1, EPA3, EPA6 and AWP1-AWP7 during in vivo biofilm formation. RESULTS Mature biofilms were observed within the first 48 h and the amount of biofilm reached its maximum by 6 days. Architecturally, mature C. glabrata biofilms consisted of a thick network of yeast cells embedded in an extracellular matrix. Moreover, in vivo biofilms were susceptible to echinocandin drugs, whereas fluconazole remained ineffective. Gene expression profiling revealed that EPA3, EPA6, AWP2, AWP3 and AWP5 were up-regulated in in vivo biofilms compared with in vitro biofilms. CONCLUSIONS C. glabrata is a unique microorganism, which, despite the lack of transition to the hyphal form, formed thick biofilms inside foreign bodies in vivo. To our knowledge, this is the first study that has described in vivo C. glabrata biofilm development and its architectural changes in detail and provides an insight into the susceptibility profile, as well as the gene expression machinery, of biofilm-associated infections.

Novel anti-infective implant substrates: Controlled release of antibiofilm compounds from mesoporous silica-containing macroporous titanium

Bone implants with open porosity enable fast osseointegration, but also present an increased risk of biofilm-associated infections. We design a novel implant material consisting of a mesoporous SiO2 diffusion barrier (pore diameter: 6.4 nm) with controlled drug release functionality integrated in a macroporous Ti load-bearing structure (fully interconnected open porosity: 30%; pore window size: 0.5-2.0 μm). Using an in vitro tool consisting of Ti/SiO2 disks in an insert set-up, through which molecules can diffuse from feed side to release side, a continuous release without initial burst effect of the antibiofilm compound toremifene is sustained for at least 9 days, while release concentrations (up to 17 μM daily) increase with feed concentrations (up to 4mM). Toremifene diffusivity through the SiO2 phase into H2O is estimated around 10(-13)m(2)/s, suggesting configurational diffusion through mesopores. Candida albicans biofilm growth on the toremifene-release side is significantly inhibited, establishing a proof-of-concept for the drug delivery functionality of mesoporous SiO2 incorporated into a high-strength macroporous Ti carrier. Next-generation implants made of this composite material and equipped with an internal reservoir (feed side) can yield long-term controlled release of antibiofilm compounds, effectively treating infections on the implant surface (release side) over a prolonged time.

CoCr F75 scaffolds produced by additive manufacturing: Influence of chemical etching on powder removal and mechanical performance

Additive manufacturing techniques such as Selective Laser Melting (SLM) allow carefully controlled production of complex porous structures such as scaffolds. These advanced structures can offer many interesting advantages over conventionally produced products in terms of biological response and patient specific design. The surface finish of AM parts is often poor because of the layer wise nature of the process and adhering particles. Loosening of these particles after implantation should be avoided, as this could put the patients health at risk. In this study the use of hydrochloric acid and hydrogen peroxide mixtures for surface treatment of cobalt-chromium F75 scaffolds produced by SLM is investigated. A 27% HCl and 8% H2O2 etchant proved effective in removing adhering particles while retaining the quasi-static and fatigue performance of the scaffolds.

Influence of short chain organic acids and bases on the wetting properties and surface energy of submicrometer ceramic powders.

The effect of short chained organic acids and bases on the surface energy and wetting properties of submicrometer alumina powder was assessed. The surface chemistry of treated powders was determined by means of Diffuse Reflectance Infrared Fourier Transform spectroscopy and compared to untreated powder. The wetting of powders was measured using a modified Washburn method, based on the use of precompacted powder samples. The geometric factor needed to calculate the contact angle was derived from measurements of the porous properties of the powder compacts. Contact angle measurements with several probe liquids before and after modification allowed a theoretical estimation of the surface energy based on the surface tension component theory. Trends in the surface energy components were linked to observations in infrared spectra. The results showed that the hydrophobic character of the precompacted powder depends on both the chain length and polar group of the modifying agent.

Alternating Current Electrophoretic Deposition for the Immobilization of Antimicrobial Agents on Titanium Implant Surfaces

One prominent cause of implant failure is infection; therefore, research is focusing on developing surface coatings that render the surface resistant to colonization by micro-organisms. Permanently attached coatings of antimicrobial molecules are of particular interest because of the reduced cytoxicity and lower risk of developing resistance compared to controlled release coatings. In this study, we focus on the chemical grafting of bioactive molecules on titanium. To concentrate the molecules at the metallic implant surface, we propose electrophoretic deposition (EPD) applying alternating current (AC) signals with an asymmetrical wave shape. We show that for the model molecule bovine serum albumin (BSA), as well as for the clinically relevant antifungal lipopeptide caspofungin (CASP), the deposition yield is drastically improved by superimposing a DC offset in the direction of the high-amplitude peak of the AC signal. Additionally, in order to produce immobilized CASP coatings, this experimental AC/DC-EPD method is combined with an established surface activation protocol. Principle component analysis (PCA) of time-of-flight secondary ion mass spectrometry (ToF-SIMS) data confirm the immobilization of CASP with higher yield as compared to a diffusion-controlled process, and higher purity than the clinical CASP starting suspensions. Scratch testing data indicate good coating adhesion. Importantly, the coatings remain active against the fungal pathogen C. albicans as shown by in vitro biofilm experiments. In summary, this paper delivers a proof-of-concept for the application of AC-EPD as a fast grafting tool for antimicrobial molecules without compromising their activities.

Throwing Power during Electrophoretic Deposition

The deposit can induce an extra potential drop near the electrode, depending on the suspension composition. This can result in a levelling off of the deposition rate in a constant-voltage deposition process. The magnitude of the extra voltage drop determines the uniformity of the deposit as function of the uniformity of the electric field present at the deposition electrode. It was experimentally proven that a uniform Al2O3 coating thickness was obtained in a non-homogeneous electrical field in ethanol with addition of HNO3, while the coating thickness varied uniformly with the E-field strength for a MEK with n-butylamine based suspension. The uniformity of the coating deposited from these suspensions was related to the measured potential drop over the deposit during electrophoretic deposition.

Alternating Current Electrophoretic Deposition of Bovine Serum Albumin onto Magnesium

Magnesium and magnesium alloys are gaining considerable attention for use in biomedical applications due to their capability to completely resorb in the human body without noticeable side effects. For structural biomedical applications however, the resorption rate is too large. In order to decrease this rate researchers are investigating magnesium alloys with an increased corrosion resistance and/or biodegradable coatings, such as dense protein layers, which retard the resorption.In this work, we demonstrate the electrophoretic deposition of Bovine Serum Albumin (BSA) directly onto pure magnesium substrates using unbalanced alternating fields (AC-EPD). The effect of the obtained coatings on the corrosion behavior of the substrates was evaluated by potentiodynamic polarization. The results show that an albumin layer deposited by AC-EPD from a 50/50 ethanol/H2O medium significantly reduces the corrosion rate.

Aqueous Electrophoretic Deposition at High Electric Fields

From an environmental, safety and economic perspective water should be the solvent of choice for electrophoretic deposition under industrial circumstances. However, because of the electrolytic decomposition of water under the influence of direct current, the majority of EPD is carried out in non-aqueous solvents. In this work, experiments prove that deposits can be obtained from aqueous alumina suspensions while avoiding electrolysis of the medium by using unbalanced alternating current fields [1]. In addition it is shown that the formed deposits have a green density which is intrinsically higher than those formed by traditional DC EPD from ethanol based suspensions. A theoretical basis for both electrophoretic deposition by means of unbalanced alternating fields and the higher density of deposits formed by application of such fields is provided.