The effects of nanopattern surface technology and targeted metabolic therapies on orthopaedic implant related infections

Bacterial biofilm infections cause significant morbidity in orthopaedic joint replacement. One of the most common bacteria in orthopaedic prosthetic infections is Staphylococcus aureus. Infection causes implant failure due to bacterial adherence and subsequent biofilm production. Nanotopography refers to the topography of a surface at the nanometre level and has major effects on cell behaviour. Studies suggest that surface nanotopography impacts the differential ability of staphylococci species to adhere, and may reduce orthopaedic implant infection rate. This research thesis focuses on bacterial adhesion on nanofabricated materials, and investigates the related metabolic changes and possible interventions. Staphylococcus aureus growth and quantification methods were optimised, with regard to growth media, incubation time and lysozyme incubation time. Both polystyrene and titanium (Ti) nanosurfaces were studied. Adhesion analysis was performed using fluorescence imaging, quantitative PCR, and bacterial percentage coverage. Metabolomic analysis was conducted by substitution with ‘heavy’ labelled glucose into growth medium, thus allowing for bacterial metabolomic analysis and identification of up-regulated, labelled metabolites and pathways. Bacterial growth was optimal using DMEM + supplement media, with adhesion occurring after 1hr bacterial incubation. Optimal lysozyme incubation for bacterial quantification using qPCR was 2hr. These parameters were used for all subsequent experimentation. Surface topography affects cell behaviour, bacterial adhesion and long term implant survival can be affected. This study found reduced bacterial adhesion on the SQ and HEX polystyrene patterns. While not found to be significant, this trend was supported by a lower average percentage bacterial coverage on both the SQ and HEX patterns (P=0.05 and P=0.01, respectively). It may be that the SQ and HEX nanopatterns are the optimal nanopit orientation required to prevent bacteria microcolony formation, keeping the bacteria in small, isolated clusters. In addition, this series of investigations showed an increase in bacterial concentrations on both the 2.5Hr and 3Hr treated Ti nanowire discs when compared to the polished Ti control disc, suggesting nanoroughness increases are associated with elevated bacterial adhesion. This theory was further supported by average percentage coverage, being significantly higher on the 2.5Hr and 3Hr treated discs. If, however, a disordered NC Ti nanopattern, hexagonal in nature, is used bacterial adhesion is significantly reduced when compared to a polished, control surface. The bacterial percentage coverage was also noted to be significantly lower on the NC surfaces, with over a 10-fold reduction when compared to the control surface. It is postulated that this reduction is through similar mechanisms to those described by Ivanova et al, and primarily related to altered surface interactions. Metabolomic analysis demonstrated increased intensity counts for key metabolites (pyruvate, aspartate, alanine and carbamoyl aspartate) involved in bacterial aggregation, proteoglycan and DNA synthesis. These pathways are also known to be important in bacterial biofilm production. Therapeutic targeting of these pathways was found to result in significantly reduced bacterial adhesion. This study shows that by altering nanotopography bacterial adhesion, and therefore, biofilm formation can be affected. Specific nanopatterned surfaces may reduce implant infection associated morbidity and mortality. The identification of metabolic pathways involved in adhesion allows for a targeted approach to biofilm eradication in S. aureus. This is of significant benefit to the patient, the surgeon and the NHS, and may well extend far beyond the realms of orthopaedics

Development and characterisation of a novel three-dimensional inter-kingdom wound biofilm model

Chronic diabetic foot ulcers are frequently colonised and infected by polymicrobial biofilms that ultimately prevent healing. This study aimed to create a novel in vitro inter-kingdom wound biofilm model on complex hydrogel-based cellulose substrata to test commonly used topical wound treatments. Inter-kingdom triadic biofilms composed of Candida albicans, Pseudomonas aeruginosa, and Staphylococcus aureus were shown to be quantitatively greater in this model compared to a simple substratum when assessed by conventional culture, metabolic dye and live dead qPCR. These biofilms were both structurally complex and compositionally dynamic in response to topical therapy, so when treated with either chlorhexidine or povidone iodine, principal component analysis revealed that the 3-D cellulose model was minimally impacted compared to the simple substratum model. This study highlights the importance of biofilm substratum and inclusion of relevant polymicrobial and inter-kingdom components, as these impact penetration and efficacy of topical antiseptics

Implications of antimicrobial combinations in complex wound biofilms containing fungi

Diabetic foot ulcer treatment currently focuses on targeting bacterial biofilms, while dismissing fungi. To investigate this we used an in vitro biofilm model containing bacteria and fungi, reflective of the wound environment, to test the impact of antimicrobials. Here we showed that while mono-treatment approaches influenced biofilm composition it had no discernible effect on overall quantity. Only by combining bacterial and fungal specific antibiotics were we able to decrease the biofilm bioburden, irrespective of composition

Assessing the bioactive profile of anti-fungal loaded calcium sulfate against fungal biofilms

Calcium sulfate (CS) has been used clinically as a bone or void filling biomaterial, and due to its resorptive properties have provided the prospect for its use as a release mechanism for local antibiotics to control biofilms. Here, we aimed to test CS beads loaded with three antifungal drugs against planktonic and sessile fungal species to assess whether these antifungal beads could be harnessed to provide consistent release of antifungals at biofilm inhibitive doses. A panel of different fungal species (n=15) were selected for planktonic broth microdilution testing with fluconazole (FLZ), amphotericin B (AMB) and caspofungin (CSP). After establishing planktonic inhibition, antifungal CS beads were introduced to fungal biofilms (n=5) to assess biofilm formation and cell viability through a combination of standard quantitative and qualitative biofilm assays. Inoculation of a hydrogel substrate, packed with antifungal CS beads, was also used to assess diffusion through a semi-dry material, to mimic active infection in-vivo. In general, antifungals released from CS loaded beads were all effective at inhibiting the pathogenic fungi over 7-days within standard MIC ranges for these fungi. We observed a significant reduction of pre-grown fungal biofilms across key fungal pathogens following treatment, with visually observable changes in cell morphology and biofilm coverage provided by scanning electron microscopy. Assessment of biofilm inhibition also revealed reductions in total and viable cells across all organisms tested. These data show that antifungal loaded CS beads produce a sustained antimicrobial effect, which inhibits and kills clinically relevant fungal species in-vitro as planktonic and biofilm cells

Géomorphologie des côtes rocheuses Arctiques

No abstract available

Estuarine and Coastal Geology and Geomorphology

[Extract] Coastal geodiversity, the geological and geomorphological processes and landforms of estuaries and coasts, is of vital importance as providing not only protection from marine processes such as storm waves but also the foundations for sustaining the integrity and biodiversity of ecosystems along our shores. Since the characteristics and functioning of coastal and estuarine ecosystems are addressed in subsequent volumes of the treatise, it is important now to contextualize coastal geodiversity as a key component in our understanding of how estuaries and coasts function, be it in relation to sandy beaches, tidal mud flats, or rocky coasts and in a range of latitudes from the tropical to polar shores. This volume, thus, sets out to capture some of the wide range of coastal geodiversity and set it within a framework that will be of relevance to the subsequent volumes of this treatise. In spite of this, we are aware that not all coastal contexts are represented here, and in future editions of the treatise we intend to take the opportunity to fill any gaps identified

Acute longitudinal ligament rupture following acute spinal trauma

The authors present a rare case of anterior longitudinal ligament (ALL) rupture in a 47- year-old gentleman following a bicycle accident. The ALL is a continuous band of a variable thickness that acts as a primary spinal stabiliser. Stress, strain or rupture of the ALL usually occurs as a result of hyperextension, with the primary perpetrator being whiplash injuries. Such injuries have been shown to result in cervical spine instability during extension, axial rotation, and lateral bending modes. Spine radiographs of such patients may be routinely assessed as normal, therefore this specific type of injury does not lend itself to identification by traditional imaging methods. This account demonstrates the importance of having a high index of suspicion of a ligamentous neck injury in the setting of normal plain radiographs but abnormal clinical examination

Géomorphologie des côtes rocheuses Arctiques

No abstract available

Knee Arthroscopy: The “Crevice Sign,” a New Pathognomonic Sign for Unstable Posterior Medial Meniscal Tear in Anterior Cruciate Ligament–Deficient Knees

There has been increased emphasis on medial meniscus repair in the anterior cruciate ligament–reconstructed knee, as this improves stability. We describe an arthroscopic sign of an unstable medial meniscal tear that is diagnostic. The “crevice sign” is a longitudinal fissure located on the distal medial femoral condyle. In the anterior cruciate ligament–deficient knee, there is increased strain on the medial meniscus. A posterior longitudinal medial meniscal tear can occur at the time of the index injury or with subsequent instability events. During this injury, the knee pivots and the anterior edge of the unstable medial meniscus digs into the articular cartilage of the medial femoral condyle, resulting in a longitudinal split of the distal femoral condyle articular cartilage. If this sign is observed during arthroscopy, it is recommended that surgeons thoroughly probe the medial meniscus to ensure no pathology is missed

Antifungal loaded calcium sulfate beads as a potential therapeutic in combating Candida auris

Candida auris provides a substantial global nosocomial threat clinically. With the recent emergence that the organism can readily colonize skin niches, it will likely continue to pose a risk in healthcare units, particularly to patients undergoing surgery. The purpose of this study was to investigate the efficacy of antifungal loaded calcium sulfate beads in combatting C. auris infection. We demonstrate that the CS-packed beads have the potential to interfere with planktonic and sessile C. auris