Jonathan Pratten

In vitro attachment of staphylococcus epidermidis to surgical sutures with and without Ag-containing bioactive glass coating

The ability of a silver-doped bioactive glass (AgBG) coating to prevent bacterial colonization on surgical sutures was investigated in vitro. Bioactive glass powders, in the form of 45S5 Bioglass® and AgBG, were used to coat Mersilk® sutures using an optimized ‘in house’ slurry-dipping process. In vitro experiments were carried out using Staphylococcus epidermidis under both batch and flow conditions. While the traditional batch culture testing was used to determine the number of viable cells adhered to the surface, the flow-cell was used to visualize attachment and detachment over time. Under batch conditions of up to 180 min, statistically significant differences were observed in the colony forming units (CFU) per suture for both the coated and uncoated Mersilk® sutures. The results showed that the AgBG coating had the greatest effect on limiting bacterial attachment (8 102 CFU) when compared to the 45S5 Bioglass® coating (3.2 103 CFU) and the uncoated Mersilk® (1.2 104 CFU). Also under flow conditions differences were seen between the coated and uncoated sutures. Therefore, this preliminary study has demonstrated the quantification and visualization of bacterial attachment onto sutures in order to compare the antibacterial properties of Ag-containing bioactive glass coatings. The bactericidal properties imparted by Ag-containing glass open new opportunities for use of the composite sutures in wound healing and body wall repair.

Cellulose Acetate Containing Toluidine Blue and Rose Bengal Is an Effective Antimicrobial Coating when Exposed to White Light

ABSTRACT Simple methods of reducing the microbial load on surfaces in hospitals are needed to reduce the risk of hospital-associated infections. Here we report on the ability of a cellulose acetate coating containing the photosensitizers toluidine blue and rose bengal to kill microbes (Staphylococcus aureus, Escherichia coli, Clostridium difficile, a bacteriophage, and Candida albicans) on its surface when illuminated with white light.

In vitro studies of the effect of antiseptic‐containing mouthwashes on the formation and viability of Streptococcus sanguis biofilms

The aims of this study were to evaluate the growth of Streptococcus sanguis on hydroxyapatite, bovine enamel and polytetrafluoroethylene substrata in a constant depth film fermentor, and to determine the effects of three antimicrobial‐containing mouthwashes on biofilm formation and bacterial viability on hydroxyapatite and enamel. There was little difference in the final cell density (5 × 104 cfu mm−2) of the Strep. sanguis biofilm on the three substrata. When hydroxyapatite‐grown biofilms were exposed to the mouthwashes for 1 min, the one containing triclosan (T) proved the most effective. The chlorhexidine‐containing mouthwash (CX) also achieved significant kills. The T‐containing mouthwash was the most effective at killing biofilms grown on enamel. Pre‐treatment of hydroxyapatite with CX, cetylpyridium chloride (CPC) or T for 1 min resulted in undetectable biofilm formation after 8 h. After 8 h of growth, only biofilms grown on enamel discs pre‐treated with CX showed a reduction in the number of viable organisms. In conclusion, the results of this study have shown that while growth of Strep. sanguis on hydroxyapatite and enamel were similar, the ability of antimicrobial agents to prevent the accumulation of viable bacteria depended on the nature of the substratum.

Effect of Silver Content on the Structure and Antibacterial Activity of Silver-Doped Phosphate-Based Glasses

ABSTRACT Staphylococcus aureus can cause a range of diseases, such as osteomyelitis, as well as colonize implanted medical devices. In most instances the organism forms biofilms that not only are resistant to the bodys defense mechanisms but also display decreased susceptibilities to antibiotics. In the present study, we have examined the effect of increasing silver contents in phosphate-based glasses to prevent the formation of S. aureus biofilms. Silver was found to be an effective bactericidal agent against S. aureus biofilms, and the rate of silver ion release (0.42 to 1.22 μg·mm−2·h−1) from phosphate-based glass was found to account for the variation in its bactericidal effect. Analysis of biofilms by confocal microscopy indicated that they consisted of an upper layer of viable bacteria together with a layer (∼20 μm) of nonviable cells on the glass surface. Our results showed that regardless of the silver contents in these glasses (10, 15, or 20 mol%) the silver exists in its +1 oxidation state, which is known to be a highly effective bactericidal agent compared to that of silver in other oxidation states (+2 or +3). Analysis of the glasses by 31P nuclear magnetic resonance imaging and high-energy X-ray diffraction showed that it is the structural rearrangement of the phosphate network that is responsible for the variation in silver ion release and the associated bactericidal effectiveness. Thus, an understanding of the glass structure is important in interpreting the in vitro data and also has important clinical implications for the potential use of the phosphate-based glasses in orthopedic applications to deliver silver ions to combat S. aureus biofilm infections.

Controlled delivery of antimicrobial gallium ions from phosphate-based glasses

Gallium-doped phosphate-based glasses (PBGs) have been recently shown to have antibacterial activity. However, the delivery of gallium ions from these glasses can be improved by altering the calcium ion concentration to control the degradation rate of the glasses. In the present study, the effect of increasing calcium content in novel gallium (Ga2O3)-doped PBGs on the susceptibility of Pseudomonas aeruginosa is examined. The lack of new antibiotics in development makes gallium-doped PBG potentially a highly promising new therapeutic agent. The results show that an increase in calcium content (14, 15 and 16 mol.% CaO) cause a decrease in degradation rate (17.6, 13.5 and 7.3 microg mm(-2) h(-1)), gallium ion release and antimicrobial activity against planktonic P. aeruginosa. The most potent glass composition (containing 14 mol.% CaO) was then evaluated for its ability to prevent the growth of biofilms of P. aeruginosa. Gallium release was found to reduce biofilm growth of P. aeruginosa with a maximum effect (0.86 log(10) CFU reduction compared to Ga2O3-free glasses) after 48 h. Analysis of the biofilms by confocal microscopy confirmed the anti-biofilm effect of these glasses as it showed both viable and non-viable bacteria on the glass surface. Results of the solubility and ion release studies show that this glass system is suitable for controlled delivery of Ga3+. 71Ga NMR and Ga K-edge XANES measurements indicate that the gallium is octahedrally coordinated by oxygen atoms in all samples. The results presented here suggest that PBGs may be useful in controlled drug delivery applications, to deliver gallium ions in order to prevent infections due to P. aeruginosa biofilms.

Transfer of Tn916-Like Elements in Microcosm Dental Plaques

ABSTRACT Microcosm dental plaques were grown from an inoculum of human saliva in a constant-depth film fermentor. The inoculum contained four tetracycline-resistant streptococcal species, each of which contained a Tn916-like element. This element was shown to transfer to other streptococci both in filter-mating experiments and within the biofilms in the fermentor.

Staphylococcus aureus accessory regulators: expression within biofilms and effect on adhesion.

Many of the genes encoding the virulence factors for Staphylococcus aureus are controlled by the accessory gene regulator (agr) and staphylococcal accessory regulator (sar). This regulation may be affected by the environment in which the organisms are grown. In the majority of ecosystems, bacteria grow attached to surfaces and form biofilms. We used S. aureus strains containing mutations inactivating agr and sar to determine whether the presence of these genes influences the attachment of the bacterium to a surface. We also used strains harbouring reporter constructs of the agr and sar operons to determine their expression in biofilms. The attachment study results showed that the sarA mutant strain adhered better to glass than did the agrA mutant or the wild type. There was an increased adherence to fibronectin-coated glass for all three strains compared to glass. Thus, these adhesion studies demonstrate that agr and sar have pleiotrophic effects on the surface expression of molecules responsible for binding to different substrata. In the biofilms higher numbers of bacteria and the greatest expression were observed at the base, but there were no observable differences between the reporter constructs. Expression of the agr and sar reporter fusions was significantly higher in the deepest layers of the biofilms where the greatest numbers of bacteria were also observed, perhaps as one might expect for genes that are regulated in a cell density dependent fashion.

The effect of orthodontic bonding materials on dental plaque accumulation and composition in vitro

The aim of this study was to investigate the accumulation and composition of microcosm dental plaque on different orthodontic bonding materials using an in vitro model. Microcosm plaques were grown on discs of a range of bonding materials in a constant depth film fermentor. The biofilms were derived from human saliva and supplied with artificial saliva as a source of nutrients. The number of viable bacteria in the biofilms was determined and the streptococci present were identified to species level. The results showed that there was no significant difference in bacterial accumulation between different bonding materials, however, biofilms grown on materials which were fluoride releasing, did not contain Streptococcus mutans. This in vitro study has shown that the use of fluoride-releasing bonding materials may support the growth of supragingival plaque, which does not contain S. mutans.

An in vitro study of the effect of fluoridated milk on oral bacterial biofilms.

ABSTRACT Microcosmic dental plaques were grown in artificial saliva and supplemented with either milk or fluoridated milk. The presence of fluoride in the milk increased the pH of the biofilms and reduced the proportions of streptococci, demonstrating that in this model, fluoridation of milk produces biofilms with reduced cariogenic potential.

Association Between Interleukin-6 −174 Polymorphism and Aggregatibacter actinomycetemcomitans in Chronic Periodontitis

BACKGROUND We previously demonstrated an association between interleukin-6 (IL-6) polymorphisms and the detection of periodontopathogenic bacteria in aggressive and chronic periodontitis on a patient basis (pooled samples). The aim of this study is to comprehensively analyze the relative contribution of IL-6 genetic factors and local (tooth and site) factors on the detection of Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans) and Porphyromonas gingivalis in patients with chronic periodontitis. METHODS Forty patients diagnosed with severe chronic periodontitis had subgingival samples harvested from four sites (the deepest probing depths in each quadrant). All subjects had a blood sample taken to genotype the single nucleotide polymorphism at position -174 in the IL-6 gene promoter. Nested polymerase chain reaction analysis was performed on subgingival plaque samples for the detection of A. actinomycetemcomitans and P. gingivalis in each of the 160 sampled sites. The association between IL-6 -174 genotypes and bacterial detection was investigated with multilevel analysis accounting for the clustering of multiple sites analyzed within patients. RESULTS Respectively 60%, 62%, and 40% of subjects had A. actinomycetemcomitans, P. gingivalis, and both bacteria concomitantly detected in ≥1 site. The multilevel analysis confirmed that, among all site and subject factors, IL-6 -174 G homozygosity showed the strongest association with the presence of A. actinomycetemcomitans in all subjects and in the subgroup of whites only. No associations were detected for P. gingivalis. CONCLUSION This study provides further confirmatory evidence that the detection of A. actinomycetemcomitans is associated with IL-6 genetic factors in chronic periodontitis cases.