Sustained release varnish containing chlorhexidine for prevention of Streptococcus mutans biofilm formation on voice prosthesis surface: an in vitro study.

OBJECTIVES: In this study, we aimed to develop a novel, sustained release varnish (SRV) for voice prostheses (VP) releasing chlorhexidine (CHX), for the prevention of biofilm formation caused by the common oral bacteria Streptococcus mutans on VP surfaces. METHODS: This study was performed in an in vitro model as a step towards future in vivo trials. VPs were coated with a SRV containing CHX (SRV-CHX) or SRV alone (placebo-SRV) that were daily exposed to S. mutans. The polymeric materials of SRV were composed of ethylcellulose and PEG-400. Biofilm formation was assessed by DNA quantification (qPCR), crystal violet staining, confocal laser scanning microscopy (CLSM), scanning electron microscopy (SEM), and kinetics experiments. RESULTS: The amount of DNA in the biofilms formed by S. mutans on VP surfaces coated once with SRV-CHX (1.024 ± 0.218 ng DNA/piece) was 58.5 ± 8.8% lower than that of placebo-SRV-coated VPs (2.465 ± 0.198 ng DNA/piece) after a 48-h exposure to S. mutans (p = 0.038). Reduced biofilm mass on SRV-CHX-coated VPs was visually confirmed by CLSM and SEM. CV staining of SRV-CHX single-coated VPs that have been exposed to S. mutans nine times showed a 98.1 ± 0.2% reduction in biofilm mass compared to placebo-SRV-coated VPs (p = 0.003). Kinetic experiments revealed that SRV-CHX triple-coated VPs could delay bacterial growth for 23 days. CONCLUSIONS: Coating VPs with SRV-CHX has an inhibitory effect on biofilm formation and prevents bacterial growth in their vicinities. This study is a proof-of-principle that paves the way for developing new clinical means for reducing both VPs' bacterial biofilm formation and device failure.

Tooth mousse containing casein phosphopeptide-amorphous calcium phosphate prevents biofilm formation of Streptococcus mutans.

BACKGROUND: Streptococcus mutans is a common cariogenic bacterium in the oral cavity involved in plaque formation. Casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) has been introduced into tooth mousse to encourage remineralization of dental enamel. The aim of this research was to study the effect of tooth mousse containing CPP-ACP (GC Tooth Mousse®) or CPP-ACP with 0.2% fluoride (CPP-ACPF; GC Tooth Mousse Plus®; GCP) on S. mutans planktonic growth and biofilm formation. METHODS: S. mutans was cultivated in the presence of different dilutions of the tooth mousse containing CPP-ACP or CPP-ACPF, and the planktonic growth was determined by ATP viability assay and counting colony-forming units (CFUs). The resulting biofilms were examined by crystal violet staining, MTT metabolic assay, confocal laser scanning microscopy (CLSM), and scanning electron microscope (SEM). RESULTS: The CPP-ACP tooth mousse (GC) at a dilution of 5-50 mg/ml (0.5-5%) did not inhibit planktonic growth, and even increased the ATP content and the number of viable bacteria after a 24 h incubation. The same was observed for the CPP-ACPF tooth mousse (GCP), except for the higher concentrations (25 and 50 mg/ml) that led to a drop in the bacterial count. Importantly, both compounds significantly decreased S. mutans biofilm formation at dilutions as low as 1.5-3 mg/ml. 12.5 mg/ml GC and 6.25 mg/ml GCP inhibited biofilm formation by 90% after 4 h. After 24 h, the MBIC90 was 6.25 mg/ml for both. CLSM images confirmed the strong inhibitory effect GC and GCP had on biofilm formation when using 5 mg/ml tooth mousse. SEM images of those bacteria that managed to form biofilm in the presence of 5 mg/ml tooth mousse, showed alterations in the bacterial morphology, where the streptococci appear 25-30% shorter on the average than the control bacteria. CONCLUSION: Our data show that the tooth mousse containing CPP-ACP reduces biofilm formation of the cariogenic bacterium S. mutans without killing the bacteria. The use of natural substances which inhibit biofilm development without killing the bacteria, has therapeutic benefits, especially in orthodontic pediatric patients.

Cannabigerol Prevents Quorum Sensing and Biofilm Formation of Vibrio harveyi.

Cannabigerol (CBG) is a non-psychoactive cannabinoid naturally present in trace amounts in the Cannabis plant. So far, CBG has been shown to exert diverse activities in eukaryotes. However, much less is known about its effects on prokaryotes. In this study, we investigated the potential role of CBG as an anti-biofilm and anti-quorum sensing agent against Vibrio harveyi. Quorum sensing (QS) is a cell-to-cell communication system among bacteria that involves small signaling molecules called autoinducers, enabling bacteria to sense the surrounding environment. The autoinducers cause alterations in gene expression and induce bioluminescence, pigment production, motility and biofilm formation. The effect of CBG was tested on V. harveyi grown under planktonic and biofilm conditions. CBG reduced the QS-regulated bioluminescence and biofilm formation of V. harveyi at concentrations not affecting the planktonic bacterial growth. CBG also reduced the motility of V. harveyi in a dose-dependent manner. We further observed that CBG increased LuxO expression and activity, with a concomitant 80% downregulation of the LuxR gene. Exogenous addition of autoinducers could not overcome the QS-inhibitory effect of CBG, suggesting that CBG interferes with the transmission of the autoinducer signals. In conclusion, our study shows that CBG is a potential anti-biofilm agent via inhibition of the QS cascade.

Chlorhexidine sustained-release varnishes for catheter coating - Dissolution kinetics and antibiofilm properties.

Catheter-associated urinary tract infections are difficult to eradicate or prevent, due to their biofilm-related nature. Chlorhexidine, a widely used antiseptic, was previously found to be effective against catheter-related biofilms. For the present study, we developed sustained-release chlorhexidine varnishes for catheter coating and evaluated their antibiofilm properties and chlorhexidine-dissolution kinetics under various conditions. The varnishes were based on ethylcellulose or ammonio methacrylate copolymer type A (Eudragit® RL). Chlorhexidine was released by diffusion from a heterogeneous matrix in the case of the ethylcellulose-based formulation, and from a homogeneous matrix in the case of Eudragit® RL. This dictated the release pattern of chlorhexidine under testing conditions: from film specimens, and from coated catheters in a static or flow-through system. Momentary saturation was observed with the flow-through system in Eudragit® RL-based coatings, an effect that might be present in vivo with other formulations as well. The coatings were retained on the catheters for at least 2weeks, and showed prolonged activity in a biological medium, including an antibiofilm effect against Pseudomonas aeruginosa. The current study demonstrates the potential of catheter coatings with sustained release of chlorhexidine in the prevention of catheter-associated urinary tract infections.

Pseudomonal Biofilm Topographic Distribution on Tympanostomy Tubes: An In Vitro Model.

In vitro growth of Pseudomonas aeruginosa biofilm on tympanostomy tubes showed that in Armstrong T-tubes, biofilm colonies were mainly located in the perpendicular junction between the body and the flanges, but in Paparella-type tube, they concentrated on the round rims. These "weakness" zones can be the future target areas for geometry changes, and can be specifically coated with antibiofilm materials.

Expression of tropoelastin in human periodontal ligament fibroblasts after simulation of orthodontic force.

The ability of elastic fibers to respond to mechanical stimuli suggests that they play a central role in physiological adaptation to external stimuli including application of orthodontic force. The purpose of this study was to examine the effect of external pressure simulating orthodontic force on tropoelastin gene expression in cultured human periodontal ligament fibroblasts (hPDLF). External pressure simulation was achieved by centrifugation for 10, 30, 60, 90 and 120 min of hPDLF in a horizontal microplate rotor. Semi-quantitative RT-PCR analysis of tropoelastin mRNA was performed and beta-actin was used as an internal invariant control. While centrifugal force on mRNA levels of beta-actin showed almost no change, the mRNA levels of tropoelastin increased significantly to a peak level of more than four-fold after 30 min. Thereafter, at 60 min, the mRNA levels remained at more than three-fold. After 90 min, mRNA levels decreased to control levels. The finding that no changes in mRNA levels of beta-actin occurred during the first 90 min of centrifugation validates its use as an invariant control gene in such an experimental model. This study demonstrated that tropoelastin is expressed in hPDLF and that the pressure caused significant time-dependent upregulation of the tropoelastin gene. The responsiveness of the tropoelastin gene to force shows its possible clinical importance in orthodontic tooth movement. Further studies, however, are essential in order to learn whether the high expression of the gene in vitro will also be followed by corresponding protein synthesis and deposition in vivo in the extracellular matrix (ECM) of the periodontal ligament (PDL).

Sustained release varnish containing chlorhexidine for prevention of biofilm formation on urinary catheter surface: in vitro study.

BACKGROUND AND PURPOSE: Biofilms on the surfaces of urinary catheters are among the pivotal factors for recurrent and persistent infections in urology. Many techniques have been investigated and applied for eradication of these biofilms--but with no full success. The aim of this study was to examine the effect of sustained release medicated varnish, releasing chlorhexidine, on the formation of biofilm on the urinary catheter surface in an in-vitro model. MATERIALS AND METHODS: A batch model was used to test the antibacterial/antibiofilm effect of the sustained release varnish: Catheter pieces coated with sustained release varnishes were placed in bacterial growth medium that was infected with Pseudomonas aeruginosa for 96 hours. Various concentrations of chlorhexidine impregnated in the varnish were tested. After the incubation period, the catheter pieces were assessed for biofilm formation by measuring the optical density, colony-forming units, and using confocal laser scanning microscopy, and electron scanning microscopy. RESULTS: Biofilm growth measurement (colony-forming units [CFU]) on the catheter surface coated with the various concentrations of chlorhexidine in sustained released varnish revealed a 94% reduction with 1% chlorhexidine (P<0.0001) and 43% reduction with 0.1% chlorhexidine (P=0.08) coated varnish in comparison with a positive control or the placebo varnish in preventing biofilm growth of P. aeruginosa. These biologic assays were confirmed using confocal and electron microscopy. CONCLUSIONS: Of the various tested concentrations of sustained release varnishes, the 1% chlorhexidine concentration has demonstrated the superior antibiofilm effect on urinary catheters with P. aeruginosa. Although similar varnishes are used in dentistry, it needs extended research in animals before applying this technology in human trials.