Critical analysis of methods to determine growth, control and analysis of biofilms for potential non-submerged antibiofilm surfaces and coatings.

The potential uses for antibiofilm surfaces reach across different sectors with significant resultant economic, societal and health impact. For those interested in using antibiofilm surfaces in the built environment, it is important that efficacy testing methods are relevant, reproducible and standardised where possible, to ensure data outputs are applicable to end-use, and comparable across the literature. Using pre-defined keywords, a review of literature reporting on antimicrobial surfaces (78 articles), within which a potential application was described as non-submerged/non-medical surface or coating with antibiofilm action, was undertaken. The most used methods utilized the growth of biofilm in submerged and static systems. Quantification varied (from most to least commonly used) across colony forming unit counts, non-microscopy fluorescence or spectroscopy, microscopy analysis, direct agar-contact, sequencing, and ELISA. Selection of growth media, microbial species, and incubation temperature also varied. In many cases, definitions of biofilm and attempts to quantify antibiofilm activity were absent or vague. Assessing a surface after biofilm recovery or assessing potential regrowth of a biofilm after initial analysis was almost entirely absent. It is clear the field would benefit from widely agreed and adopted approaches or guidance on how to select and incorporate end-use specific conditions, alongside minimum reporting guidelines may benefit the literature.

The denture microbiome in health and disease: an exploration of a unique community.

The United Nations suggests the global population of denture wearers (an artificial device that acts as a replacement for teeth) is likely to rise significantly by the year 2050. Dentures become colonized by microbial biofilms, the composition of which is influenced by complex factors such as patient's age and health, and the nature of the denture material. Since colonization (and subsequent biofilm formation) by some micro-organisms can significantly impact the health of the denture wearer, the study of denture microbiology has long been of interest to researchers. The specific local and systemic health risks of denture plaque are different from those of dental plaque, particularly with respect to the presence of the opportunist pathogen Candida albicans and various other nonoral opportunists. Here, we reflect on advancements in our understanding of the relationship between micro-organisms, dentures, and the host, and highlight how our growing knowledge of the microbiome, biofilms, and novel antimicrobial technologies may better inform diagnosis, treatment, and prevention of denture-associated infections, thereby enhancing the quality and longevity of denture wearers.

Spreading the message of antimicrobial resistance: a detailed account of a successful public engagement event.

The increase in Antimicrobial resistance (AMR) microorganisms has been exacerbated by exposure to antimicrobial drugs (e.g. antibiotics). A solution to AMR may require academic researchers to not only contribute to the drug discovery pipeline through laboratory research, but also to engage and inform non-specialist audiences using a variety of interventions in order to change behaviour towards our use of antibiotics. In this paper, the authors describe a hands-on public engagement event focusing on AMR. 'A Spoonful of Soil', was created by drawing on the past experiences of the delivering team (also described), with planning focusing on clear concise messages, selection of an appropriate audience and ensuring the event would be of significant interest to the audience. The event had a significant footfall of over 300 visitors. Key messages which aimed to raise awareness of AMR and educate visitors on the actions and behaviours that can help address the global issue of AMR were delivered by appropriate experts successfully, however success in reaching audience cannot be concluded from the feedback and evaluation gathered.

Fitting the message to the location: engaging adults with antimicrobial resistance in a World War 2 air raid shelter.

AIMS: There are many different initiatives, global and local, designed to raise awareness of antimicrobial resistance (AMR) and change audience behaviour. However, it is not possible to assess the impact of specific, small-scale events on national and international outcomes-although one might acknowledge some contribution to the individual and collective knowledge and experience-focused 'science capital' As with any research, in preparation for a public engagement event, it is important to identify aims, and appropriate methods whose results might help satisfy those aims. Therefore, the aim of this paper was to develop, deliver and evaluate an event designed to engage an adult audience with AMR. METHODS AND RESULTS: The venue was a World War 2 air raid shelter, enabling comparison of the pre- and postantibiotic eras via three different activity stations, focusing on nursing, the search for new antibiotics and investigations into novel antimicrobials. The use of observers released the presenters from evaluation duties, enabling them to focus on their specific activities. Qualitative measures of audience engagement were combined with quantitative data. CONCLUSIONS: The evaluation revealed that adult audiences can easily be absorbed into an activity-particularly if hands-on-after a brief introduction. SIGNIFICANCE AND IMPACT OF THE STUDY: This research demonstrates that hands-on practical engagement with AMR can enable high-level interaction and learning in an informal and enjoyable environment.

Surface modifications for antimicrobial effects in the healthcare setting: a critical overview.

The spread of infections in healthcare environments is a persistent and growing problem in most countries, aggravated by the development of microbial resistance to antibiotics and disinfectants. In addition to indwelling medical devices (e.g. implants, catheters), such infections may also result from adhesion of microbes either to external solid-water interfaces such as shower caps, taps, drains, etc., or to external solid-gas interfaces such as door handles, clothes, curtains, computer keyboards, etc. The latter are the main focus of the present work, where an overview of antimicrobial coatings for such applications is presented. This review addresses well-established and novel methodologies, including chemical and physical functional modification of surfaces to reduce microbial contamination, as well as the potential risks associated with the implementation of such anticontamination measures. Different chemistry-based approaches are discussed, for instance anti-adhesive surfaces (e.g. superhydrophobic, zwitterions), contact-killing surfaces (e.g. polymer brushes, phages), and biocide-releasing surfaces (e.g. triggered release, quorum sensing-based systems). The review also assesses the impact of topographical modifications at distinct dimensions (micrometre and nanometre orders of magnitude) and the importance of applying safe-by-design criteria (e.g. toxicity, contribution for unwanted acquisition of antimicrobial resistance, long-term stability) when developing and implementing antimicrobial surfaces.

Rapid screening of the antimicrobial efficacy of Ag zeolites.

A semi-quantitative screening method was used to compare the killing efficacy of Ag zeolites against bacteria and yeast as a function of the zeolite type, crystal size and concentration. The method, which substantially reduced labor, consumables and waste and provided an excellent preliminary screen, was further validated by quantitative plate count experiments. Two pairs of zeolite X and zeolite beta with different sizes (ca. 200nm and 2µm for zeolite X and ca. 250 and 500nm for zeolite beta) were tested against Escherichia coli (E. coli) and Candida albicans (C. albicans) at concentrations in the range 0.05-0.5mgml-1. Reduction of the zeolite crystal size resulted in a decrease in the killing efficacy against both microorganisms. The semi-quantitative tests allowed convenient optimization of the zeolite concentrations to achieve targeted killing times. Zeolite beta samples showed higher activity compared to zeolite X despite their lower Ag content, which was attributed to the higher concentration of silver released from zeolite beta samples. Cytotoxicity measurements using peripheral blood mononuclear cells (PBMCs) indicated that Ag zeolite X was more toxic than Ag zeolite beta. However, the trends for the dependence of cytotoxicity on zeolite crystal size at different zeolite concentrations were different for the two zeolites and no general conclusions about zeolite cytotoxicity could be drawn from these experiments. This result indicates a complex relationship, requiring the necessity for individual cytotoxicity measurements for all antimicrobial applications based on the use of zeolites.

Effect of humidity and temperature on the survival of Listeria monocytogenes on surfaces.

Listeria monocytogenes is a pathogenic bacterium, with human disease and infection linked to dairy products, seafood, ready-to-eat meat and raw & undercooked meats. Stainless steel is the most common food preparation surface and therefore, it is important to understand how food storage conditions such as surface materials, temperature and relative humidity can affect survival of L. monocytogenes. In this study, survival of L. monocytogenes on stainless steel was investigated at three temperatures (4, 10 and 21°C), each approx. 11, 50 and 85% humidity. Results indicate that the lower the temperature, the more cells were recovered in all three humidity environments, while medium humidity enhances survival, irrespective of temperature. Lower humidity decreases recovery at all temperatures. These data support the guidance noted above that humidity control is important, and that lower humidity environments are less likely to support retention of viable L. monocytogenes on a stainless steel surface. SIGNIFICANCE AND IMPACT OF THE STUDY: Understanding survival of potential food-borne pathogens is essential for the safe production and preparation of food. While it has long been 'common knowledge' that relative humidity can affect the growth and survival of micro-organisms, this study systematically describes the survival of L. monocytogenes on stainless steel under varying humidity and temperatures for the first time. The outcomes from this paper will allow those involved with food manufacture and preparation to make informed judgement on environmental conditions relating to humidity control, which is lacking in the food standards guidelines.

Molybdenum doped titanium dioxide photocatalytic coatings for use as hygienic surfaces: the effect of soiling on antimicrobial activity.

Titanium dioxide (TiO2) surfaces doped with molybdenum (Mo) were investigated to determine if their photocatalytic ability could enhance process hygiene in the brewery industry. Doping TiO2 with Mo showed a 5-log reduction in bacterial counts within 4 to 24 h and a 1-log reduction in yeast numbers within 72 h. The presence of a dilute brewery soil on the surface did not interfere with antimicrobial activity. The TiO2-Mo surface was also active in the dark, showing a 5-log reduction in bacteria within 4 to 24 h and a 1-log reduction in yeast numbers within 72 h, suggesting it could have a novel dual function, being antimicrobial and photocatalytic. The study suggests the TiO2-Mo coating could act as a secondary barrier in helping prevent the build-up of microbial contamination on surfaces within the brewery industry, in particular in between cleaning/disinfection regimes during long production runs.

Silver confined within zeolite EMT nanoparticles: preparation and antibacterial properties.

The preparation of pure zeolite nanocrystals (EMT-type framework) and their silver ion-exchanged (Ag(+)-EMT) and reduced silver (Ag(0)-EMT) forms is reported. The template-free zeolite nanocrystals are stabilized in water suspensions and used directly for silver ion-exchange and subsequent chemical reduction under microwave irradiation. The high porosity, low Si/Al ratio, high concentration of sodium and ultrasmall crystal size of the EMT-type zeolite permitted the introduction of a high amount of silver using short ion-exchange times in the range of 2-6 h. The killing efficacy of pure EMT, Ag(+)-EMT and Ag(0)-EMT against Escherichia coli was studied semi-quantitatively. The antibacterial activity increased with increasing Ag content for both types of samples (Ag(+)-EMT and Ag(0)-EMT). The Ag(0)-EMT samples show slightly enhanced antimicrobial efficacy compared to that of Ag(+)-EMT, however, the differences are not substantial and the preparation of Ag nanoparticles is not viable considering the complexity of preparation steps.

The detection of food soils on stainless steel using energy dispersive X-ray and Fourier transform infrared spectroscopy.

Organic soiling is a major issue in the food processing industries, causing a range of biofouling and microbiological problems. Energy dispersive X-ray (EDX) and Fourier transform infra red spectroscopy (FT-IR) were used to quantify and determine the biochemical groups of food soils on stainless steel surfaces. EDX quantified organic material on surfaces where oily based residues predominated, but was limited in its usefulness since other food soils were difficult to detect. FT-IR provided spectral 'fingerprints' for each of the soils tested. Key soiling components were associated with specific peaks, viz. oils at 3025 cm(-1)-3011 cm(-1), proteins at 1698 cm(-1)-1636 cm(-1) and carbohydrates at 1658 cm(-1)-1596 cm(-1), 783 cm(-1)-742 cm(-1). High concentrations of some soils (10%) were needed for detection by both EDX and FT-IR. The two techniques may be of use for quantifying and identifying specific recalcitrant soils on surfaces to improve cleaning and hygiene regimes.

The influence of silver content on the tribological and antimicrobial properties of ZrN/Ag nanocomposite coatings.

ZrN/Ag nanocomposite coatings with varying silver contents were produced by co-deposition in a dual pulsed magnetron sputtering system. The coatings were characterised using scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD), scratch adhesion testing, thrust washer wear testing and nanoindentation. The hardness of the ZrN/Ag coatings and the friction coefficient running unlubricated against a steel counterface decreased with increasing silver content, whereas the coating-to-substrate adhesion increased for coatings with higher silver contents, compared to a 'pure' ZrN coating. The antimicrobial properties of the coatings were investigated using two well established microbiological assay techniques: zones of inhibition and a NBT (nitro-blue tetrazolium) redox dye. Zones of inhibition were used to determine the extent of silver ion release from the nanocomposite materials, and a NBT (nitro-blue tetrazolium) redox dye was used to determine the antimicrobial effectiveness of the surfaces following incubation. The microorganisms tested were Pseudomonas aeruginosa (P. aeruginosa), Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). Whilst no zones of inhibition were observed for S. aureus, on any of the surfaces, the diameter of the 'kill' zones generally increased with increasing silver content for the other microorganisms. For the NBT assays, after incubation, no P. aeruginosa colony forming units were observed on any surface and the number of viable cells of E. coli and S. aureus decreased with increasing silver content, compared to a 'pure' ZrN surface.

The retention of bacteria on hygienic surfaces presenting scratches of microbial dimensions.

AIMS: To produce surfaces of defined linear topographical features which reflect those found on worn and new stainless steel, to monitor the effect of feature dimensions on the retention of Listeria monocytogenes and Staphylococcus sciuri. METHODS AND RESULTS: Surfaces were fabricated with parallel linear features of 30 microns or of microbial dimensions (1.02 and 0.59 microm width) and used in microbial retention assays with Staph. sciuri and L. monocytogenes. Retained cells were distributed uniformly across the smooth 30 micron featured surfaces but were retained in high numbers on microtopographies at the 'peaks' between the wide grooves. On smaller features, retention was attributed to the maximum area of contact between cells and substratum being attained, with cocci being embedded in 1.02-microm-width grooves, and rods aligned along (and across) the densely packed parallel 0.59-microm grooves. CONCLUSIONS: The dimensions of surface features may enhance or impede cell retention. This phenomenon is also related to the size and shape of the microbial cell. SIGNIFICANCE AND IMPACT OF STUDY: Findings may help describe and evaluate properties of hygienic and easily cleanable surfaces.

Evaluation of in vivo denture plaque assessment methods.

BACKGROUND: Measurement and assessment of denture plaque can provide valuable information regarding an individual's oral health status and assessment of new treatments or products. Current methods tend to rely on subjective indices or image analysis derived planimetric (area measurement) assessment of stained plaque on dentures. Plaque indices are most commonly used to assess plaque coverage without image capture. This is not ideal because the methods are subjective, examiner bias may occur, there is no reproducibility between studies, the methods have lower accuracy and sensitivity than image analysis, and there is no record. To the authors' knowledge, no standardised published method of denture plaque assessment is currently employed for product development and testing. METHOD: In this study visual and planimetric plaque assessment methods were compared using reference dentures. In addition, an in vivo study compared these methods for evaluating denture cleanser efficacy. RESULTS AND CONCLUSIONS: The results show that blinded image scoring is more representative of the true plaque area coverage than 'live' denture scoring, detecting significant decreases in plaque coverage. Planimetric analysis provides a more sensitive and less subjective technique with greater differentiation between treatments. However, analysis is very time consuming. Thus, a number of recommendations are made regarding quantification of denture plaque for the assessment of cleanser products.

QLF is not readily suitable for in vivo denture plaque assessment.

OBJECTIVES: Current methods available for denture plaque assessment utilise visual and planimetric techniques. This paper evaluates the use of the Quantitative Light-induced Fluorescence system (QLF) in image capture of denture plaque and the suitability of these images for planimetric plaque measurement. It is proposed that fluorescence imaging could provide a valuable and sensitive standardising method for plaque assessment in clinical trials for denture cleansing products and denture hygiene. Indeed, the detection of red fluorescent plaque using the QLF system is indicative of black-pigmented obligate anaerobes and mature plaque. METHODS: The QLF system was evaluated in a clinical study for use in denture plaque assessment in comparison to white light based image capture. RESULTS: Despite appearing as a promising system for denture plaque quantification, this study revealed numerous problems associated with the QLF system including small focal depth, thus large numbers of images and processing time were required. In addition, differential fluorescence of acrylic made images unsuitable for plaque quantification. CONCLUSION: QLF is unsuitable for in vivo denture plaque assessment. However, the visualisation of red autofluorescence, indicating mature plaque, remains an important clinical use of QLF for denture hygiene assessment.

Review of mobile communication devices as potential reservoirs of nosocomial pathogens.

Innovation in mobile communication technology has provided novel approaches to the delivery of healthcare and improvements in the speed and quality of routine medical communication. Bacterial contamination of mobile communication devices (MCDs) could be an important issue affecting the implementation of effective infection control measures and might have an impact on efforts to reduce cross-contamination. This review examines recent studies reporting bacterial contamination of MCDs, most demonstrating that 9-25% of MCDs are contaminated with pathogenic bacteria. We examine previously investigated risk factors for MCD contamination in addition to work on surface decontamination of the device. Recommendations to reduce contamination risks include staff education, strict hand hygiene measures, guidelines on device cleaning and consideration of the restrictions regarding use of mobile phone technology in certain high risk areas, for example, operating theatres, intensive care units and burns units. Further work is required to evaluate the benefit of such interventions on MCD contamination and to determine whether a link exists between contamination and subsequent patient infection.

The use of physicochemical methods to detect organic food soils on stainless steel surfaces.

Food processing surfaces fouled with organic material pose problems ranging from aesthetic appearance, equipment malfunction and product contamination. Despite the importance of organic soiling for subsequent product quality, little is known about the interaction between surfaces and organic soil components. A range of complex and defined food soils was applied to 304 stainless steel (SS) surfaces to determine the effect of type and concentration of soil on surface physicochemical parameters, viz surface hydrophobicity (DeltaG(iwi)), surface free energy (gamma(s)), Lifshitz van der Waals (gamma_LW(s)), Lewis acid base (gamma_AB(s)), electron acceptor (gamma_+(s) ) and electron donor (gamma_-(s) ) measurements. When compared to the control surface, changes in gamma_AB(s), gamma_+(s) and gamma_-(s) were indicative of surface soiling. However, soil composition and surface coverage were heterogeneous, resulting in complex data being generated from which trends could not be discerned. These results demonstrate that the retention of food soil produces changes in the physicochemical parameters of the surface that could be used to indicate the hygienic status of a surface.

Combating implant infections. Remarks by a women's team.

Research on implant infections requires cooperative efforts and integration between basic and clinical expertises. An international group of women scientists is acting together in this field. The main research topics of the participants of this group are described. Formation of bacterial biofilms, antibiotic resistance and production of virulence factors like adhesins and toxins are investigated. New biomaterials, coatings and drugs designed to inhibit microbial adhesion are evaluated, and infection-resistant biomaterials are under study, such as a novel heparinizable polycarbonate-urethane (Bionate) or incorporation of diamino-diamide-diol (PIME) to reduce bacterial attachment. The correlation between biofilm production and the accessory-gene-regulator (agr) is investigated in Staphylococcus aureus. The ability to form biofilm has also been shown to be one of the important virulence factors of Enterococcus faecalis, favouring colonization of inert and biological surfaces. The study of quorum sensing has led to the discovery of a quorum sensing inhibitor termed RIP that suppresses staphylococcal biofilm and infections. The immune response and the local defence mechanisms of the host against implant-associated infections, activation and infiltration of immunocompetent cells into the sites of infection have been studied in patients with implant-associated osteomyelitis. Production of monoclonal antibodies (mAbs) as possible vaccines against the staphylococcal collagen-binding MSCRAMMs is in progress.

Development of an in vitro denture plaque biofilm to model denture malodour.

This study aimed to develop an in vitro denture plaque biofilm to model denture malodour. No previous studies have attempted to characterize the malodour associated with dentures and the effect of Candida spp. (main aetiological agent of denture-related stomatitis) on malodour. Pooled denture plaque microcosms and 'model' denture plaque biofilms (pooled saliva supplemented with additional microbial species) with and without addition of candida were grown aerobically at 37 °C for up to 13 days in a constant depth film fermenter (CDFF) on denture acrylic discs. Sample discs were removed, rinsed in sterile water and placed in phosphate buffered saline (PBS). The discs were vortex mixed to remove the biofilms, diluted in PBS and plated in duplicate onto general and selective media. The composition and stability of the biofilms over time were assessed. CDFF-grown microcosms and 'model' denture plaque biofilms were relatively stable in composition, with streptococci remaining the dominant microbial group. Model denture plaque biofilms were comparable in composition to denture plaque microcosms. This model system has the potential for evaluation of agents that might affect these parameters such as denture cleansers and other oral hygiene treatments.

Potential use of copper as a hygienic surface; problems associated with cumulative soiling and cleaning.

It has been suggested that antibacterial copper could be used in place of stainless steel to help reduce the occurrence of hospital-acquired infections. The antibacterial activity of copper has been clearly demonstrated when using cell suspensions held in prolonged contact with copper or copper alloys. The aim of this study was to evaluate the antimicrobial properties of copper in comparison with stainless steel in a generally dry environment. Three stainless steels of varying surface finish and polished copper were soiled with Staphylococcus aureus suspended in a protein-based organic soil (bovine serum album), dried rapidly, and then incubated for 24 h. Surfaces were then wiped clean using a standardised wiping procedure with two cleaning agents recommended by UK National Health Service guidelines. This soiling/cleaning procedure was carried out daily over five days. After each cleaning cycle the amount of residual soil and live cells was assessed using direct epifluorescence microscopy. All materials were easily cleaned after the first soiling episode but a build-up of cells and soil was observed on the copper surfaces after several cleaning/wiping cycles. Stainless steel remained highly cleanable. Accumulation of material on copper is presumably due to the high reactivity of copper, resulting in surface conditioning. This phenomenon will affect subsequent cleaning, aesthetic properties and possibly antibacterial performance. It is important to select the appropriate cleaning/disinfecting protocols for selected surfaces.