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In this work, porous FeMn(-xAg) alloys are fabricated through powder metallurgy methods. The effects of porosity and Ag addition on the microstructure, biodegradability, magnetic and mechanical properties of the alloys are investigated. Studies on the cytocompatibility, inflammatory cytokine response and antibacterial effect are also conducted. The fabricated alloys exhibit a macro- and nanoporous structure, with uniformly distributed silver particles. The biodegradability tests reveal that the release of Mn to the Hank's solution is higher than that of Fe, without significant differences between the alloys. The degradation products consist mainly of Fe, Mn, O and compounds enriched in Ca, P and Cl. As-sintered alloys show a low saturation magnetization value (below 1 emu g−1), which does not increase significantly with immersion time. The results on biocompatibility indicate that all tested alloys are non-cytotoxic, but the addition of Ag might interfere with cell proliferation. However, the ions released by the FeMn(-xAg) alloys do not induce an inflammatory response in macrophages. The obtained results on microbiological interactions reveal that although no significant bactericidal effect is observed at 4 h between FeMn control and FeMn-5Ag, a significant reduction in the total biofilm biomass of both live and dead bacteria is observed after 24 h in Ag containing FeMn-5Ag surfaces.

IntroductionProsthetic joint infections (PJIs) are disastrous complications for patients and costly for healthcare organisations. They may promote bacterial resistance due to the extensive antibiotic use necessary in the PJI treatment. The PJI incidence is estimated to be 1%–3%, but the absolute numbers worldwide are high and increasing as large joint arthroplasties are performed by the millions each year. Current treatment algorithms, based on implant preserving surgery or full revision followed by a semitailored antibiotic regimen for no less than 2–3 months, lead to infection resolution in approximately 60% and 90%, respectively. Antibiotic choice is currently guided by minimum inhibitory concentrations (MICs) of free-living bacteria and not of bacteria in biofilm growth mode. Biofilm assays with relatively rapid output for the determination of minimum biofilm eradication concentrations (MBECs) have previously been developed but their clinical usefulness have not been established.Methods and analysisThis single-blinded, two-arm randomised study of hip or knee staphylococcal PJI will evaluate 6-week standard of care (MIC guided), or an alternative antibiotic regimen according to an MBEC-guided-based decision algorithm. Sixty-four patients with a first-time PJI treated according to the debridement, antibiotics, and implant retention principle will be enrolled at a single tertiary orthopaedic centre (Sahlgrenska University Hospital). Patients will receive 14 days of standard parenteral antibiotics before entering the comparative study arms. The primary outcome measurement is the proportion of changes in antimicrobial regimen from first-line treatment dependent on randomisation arm. Secondary endpoints are unresolved infection, how microbial properties including biofilm abilities and emerging antimicrobial resistance correlate to infection outcomes, patient reported outcomes and costs with a 12-month follow-up.Ethics and disseminationApproval is received from the Swedish Ethical Review Authority, no 2020-01471 and the Swedish Medical Products Agency, EudraCT, no 2020-003444-80.Trial registration numberClinicalTrials.gov ID:NCT04488458.

Staphylococcus aureus and Staphylococcus epidermidis are the bacteria that most frequently cause osteomyelitis. This study aimed to determine whether staphylococci isolated from osteomyelitis associated with septic loosening of orthopedic prostheses release extracellular vesicles (EVs) and, if so, to determine tentative immunomodulatory effects on the human monocytic cell line THP-1. EVs were isolated from bacterial cultures using filtration and ultracentrifugation and characterized by scanning electron microscopy, nanoparticle tracking analysis and Western Blot. The cytotoxic effect of EVs was analyzed by NucleoCounter and lactate dehydrogenase (LDH) analyses. Confocal laser scanning microscopy was employed to visualize the uptake of EVs by THP-1 cells. Activation of the transcription factor nuclear factor-κB (NF-κB) was determined in THP1-Blue™ NF-κB cells, and the gene expression and secretion of cytokines were determined by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. All investigated strains, irrespective of their biofilm formation ability, were able to secrete EVs in vitro. The S. aureus strains produced significantly more EVs than the S. epidermidis strains. Both S. aureus-derived EVs and S. epidermidis-derived EVs were internalized by THP-1 cells, upregulated Toll-like receptor 3 (TLR3) gene expression, activated NF-κB, and promoted the gene expression and secretion of interleukin (IL)-8, monocyte chemoattractant protein (MCP)-1, matrix metallopeptidase (MMP)-9 and IL-10. Whereas EVs from both staphylococcal species upregulated the proapoptotic DNA damage-inducible transcript 4 (DDIT4) gene and downregulated the antiapoptotic B-cell lymphoma 2 (Bcl-2) gene, cytolysis was preferentially induced in S. aureus EV-stimulated cells, possibly related to the expression of cytolytic proteins predominantly in S. aureus EVs. In conclusion, staphylococcal EVs possess potent cytolytic and immunomodulatory properties.

Hard-to-heal wounds are typically infected with biofilm-producing microorganisms, such as Pseudomonas aeruginosa, which strongly contribute to delayed healing. Due to the global challenge of antimicrobial resistance, alternative treatment strategies are needed. Here, we investigated whether inhibition of quorum sensing (QS) by sodium salicylate in different P. aeruginosa strains (QS-competent, QS-mutant, and chronic wound strains) influences biofilm formation and tolerance to silver. Biofilm formation was evaluated in simulated serum-containing wound fluid in the presence or absence of sodium salicylate (NaSa). Biofilms were established using a 3D collagen-based biofilm model, collagen coated glass, and the Calgary biofilm device. Furthermore, the susceptibility of 48-h-old biofilms formed by laboratory and clinical strains in the presence or absence of NaSa towards silver was evaluated by assessing cell viability. Biofilms formed in the presence of NaSa were more susceptible to silver and contained reduced levels of virulence factors associated with biofilm development than those formed in the absence of NaSa. Biofilm aggregates formed by the wild-type but not the QS mutant strain, were smaller and less heterogenous in size when grown in cultures with NaSa compared to control. These data suggest that NaSa, via a reduction of cell aggregation in biofilms, allows the antiseptic to become more readily available to cells.

Antimicrobial peptides (AMPs) are seen as a promising replacement to conventional antibiotics for the prevention of skin wound infections. However, due to the short half-life of AMPs in biological environments, such as blood, their use in clinical applications has been limited. The covalent immobilization of AMPs onto suitable substrates is an effective solution to create contact-killing surfaces with increased long-term stability. In this work, an antimicrobial peptide, RRPRPRPRPWWWW-NH2 (RRP9W4N), was covalently attached to amphiphilic and ordered mesoporous Pluronic F127 hydrogels made of cross-linked lyotropic liquid crystals through 1-ethyl-3-(3-(dimethylamino)propyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) chemistry. The AMP-hydrogels showed high antibacterial activity against Staphylococcus epidermidis, Staphylococcus aureus, Pseudomonas aeruginosa, methicillin-resistant S. aureus (MRSA), and multidrug-resistant Escherichia coli for up to 24 h. Furthermore, the AMP-hydrogels did not present any toxicity to human fibroblasts. The AMPs retained their antimicrobial activity up to 48 h in human blood serum, which is a significant increase in stability compared to when used in dissolved state. A pilot in vivo rat model showed 10–100× less viable counts of S. aureus on AMP-hydrogels compared with control hydrogels during the first 3 days of infection. Studies performed on human whole blood showed that blood coagulated more readily in the presence of AMP-hydrogels as compared to hydrogels without AMPs, indicating potential hemostatic activity. Overall, the results suggest that the combination of amphiphilic hydrogels with covalently bonded AMPs has potential to be used as antibacterial wound dressing material to reduce infections and promote hemostatic activity as an alternative to antibiotics or other antimicrobial agents, whose use should be restricted.

Hard-to-heal wounds are typically infected with biofilm-producing microorganisms, such as Pseudomonas aeruginosa, which strongly contribute to delayed healing. Due to the global challenge of antimicrobial resistance, alternative treatment strategies are needed. Here, we investigated whether inhibition of quorum sensing (QS) by sodium salicylate in different P. aeruginosa strains (QS-competent, QS-mutant, and chronic wound strains) influences biofilm formation and tolerance to silver. Biofilm formation was evaluated in simulated serum-containing wound fluid in the presence or absence of sodium salicylate (NaSa). Biofilms were established using a 3D collagen-based biofilm model, collagen coated glass, and the Calgary biofilm device. Furthermore, the susceptibility of 48-h-old biofilms formed by laboratory and clinical strains in the presence or absence of NaSa towards silver was evaluated by assessing cell viability. Biofilms formed in the presence of NaSa were more susceptible to silver and contained reduced levels of virulence factors associated with biofilm development than those formed in the absence of NaSa. Biofilm aggregates formed by the wild-type but not the QS mutant strain, were smaller and less heterogenous in size when grown in cultures with NaSa compared to control. These data suggest that NaSa, via a reduction of cell aggregation in biofilms, allows the antiseptic to become more readily available to cells.

Introduction: The ability of bacteria to colonize implant surfaces and tissues as a biofilm plays a relevant role in medical-device-associated infections. Staphylococcus aureus strains can produce a biofilm matrix made of the poly-N-acetylglucosamine (PIA/PNAG) exopolysaccharide and/or proteins. PIA/PNAG is synthesised by enzymes encoded by the icaADBC operon whose expression is repressed by the transcriptional regulator IcaR, while the protein-dependent biofilm is commonly associated to fibronectin-biding proteins, FnBPA and FnBPB, encoded by fnbA and fnbB genes. The aim of this work was to identify common genetic features in the regulatory regions of biofilm-related genes among clinical S. aureus strains derived from periprosthetic joint infections (PJI). Material and Methods: Genomes of 45 S. aureus strains from PJI were sequenced. Firstly, the sequence comprising the entire icaADBC regulatory region (5’UTR of icaADBC and icaR, the icaR coding sequence and its 3’UTR region) and secondly, the sequence of the promoter region of fnbAB were compared to those of S. aureus MW2 strain. Regulatory regions containing distinctive features were identified, fused to a reporter gene and introduced in a reference strain to analyze differences in gene expression. Results: In the case of the icaADBC operon, single nucleotide polymorphisms (SNPs) in the icaADBC regulatory region allowed clustering of the strains in five groups from which a representative strain was chosen for further studies: S. aureus MIC 6924 (20% of isolates), MIC 6934 (13%), MIC 6936 (7%), MIC 6948 (2%) and MIC 7018 (4%). Of note, MICs 6948 and 7018 contained mutations in the icaR coding sequence. In this respect, a single nucleotide mutation in icaR (Val176Glu) caused a significant increase in icaADBC transcription and thus, in PIA/PNAG production and biofilm formation. In contrast, none of the rest of the SNPs found in the icaADBC regulatory region modified the transcription levels of the reporter gene. With respect to fnBPA and fnBPB genes and in agreement with previous studies, 100% of the strains contained the fnbA gene whereas only 69% contained the fnbB gene. The promoter region of fnbA was found to be highly conserved. SNPs in the promoter region of fnbB allowed clustering the strains in five groups. From these, the most frequently identified pattern was represented by S. aureus MIC 6948 (53%) and correlated with a lower level of reporter expression, whereas the group containing SNPs in the LexA binding sites was represented by MIC 7014 (4%) and correlated with higher expression levels. Conclusion: Our results suggest that S. aureus isolates from periprosthetic joint infections do not share specific features in cis regulatory regions of icaADBC and fnbB genes that may help to predict a higher expression levels of biofilm matrix compounds. Acknowledgement: This project has received funding from the European Union's H2020 research and innovation programme under Marie Sklodowska-Curie grant agreement No 801586.

Introduction. An important factor for delayed healing of chronic wounds is the presence of bacteria. Quorum sensing (QS), a cell density-dependent signalling system, controls the production of many virulence factors and biofilm formation in Pseudomonas aeruginosa . Aim. Inhibition by sodium salicylate (NaSa) of QS-regulated virulence expression was evaluated in QS-characterized clinical wound isolates of P. aeruginosa, cultured in serum-containing medium. Methodology. Fourteen clinical P. aeruginosa strains from chronic wounds were evaluated for the production of QS signals and virulence factors. Inhibition of QS by NaSa in P. aeruginosa clinical strains, wild-type PAO1 and QS reporter strains was evaluated using in vitro assays for the production of biofilm, pyocyanin, siderophores, alkaline protease, elastase and stapholytic protease. Results. Six clinical strains secreted several QS-associated virulence factors and signal molecules and two were negative for all factors. Sub-inhibitory concentrations of NaSa downregulated the expression of the QS-related genes lasB, rhlA and pqsA and reduced the secretion of several virulence factors in PAO1 and clinical strains cultured in serum. Compared to serum-free media, the presence of serum increased the expression of QS genes and production of siderophores and pyocyanin but decreased biofilm formation. Conclusions. Pseudomonas aeruginosa from chronic wound infections showed different virulence properties. While very few strains showed no QS activity, approximately half were highly virulent and produced QS signals, suggesting that the targeting of QS is a viable and relevant strategy for infection control. NaSa showed activity as a QS-inhibitor by lowering the virulence phenotypes and QS signals at both transcriptional and extracellular levels.

Purpose In this prospective clinical pilot study, abutments with different topologies (machined versus polished) were compared with respect to the clinical outcome and the microbiological profile. Furthermore, three different sampling methods (retrieval of abutment, collection of peri-abutment exudate using paper-points, and a small peri-abutment soft-tissue biopsy) were evaluated for the identification and quantification of colonising bacteria. Methods Twelve patients, seven with machined abutment and five with polished abutment, were included in the analysis. Three different sampling procedures were employed for the identification and quantification of colonising bacteria from baseline up to 12 months, using quantitative culturing. Clinical outcome measures (Holgers score, hygiene, pain, numbness and implant stability) were investigated. Results The clinical parameters, and total viable bacteria per abutment or in tissue biopsies did not differ significantly between the polished and machined abutments. The total CFU/mm2 abutment and CFU/peri-abutment fluid space of anaerobes, aerobes and staphylococci were significantly higher for the polished abutment. Anaerobic bacteria were detected in the tissue biopsies before BAHS implantation. Anaerobes and Staphylococcus spp. were detected in all three compartments after BAHS installation. For most patients (10/12), the same staphylococcal species were found in at least two of the three compartments at the same time-point. The common skin coloniser Staphylococcus epidermidis was identified in all patients but one (11/12), whereas the pathogen Staphylococcus aureus was isolated in five of the patients. Several associations between clinical and microbiological parameters were found. Conclusions There was no difference in the clinical outcome with the use of polished versus machined abutment at 3 and 12 months after implantation. The present pilot trial largely confirmed a suitable study design, sampling and analytical methodology to determine the effects of modified BAHS abutment properties. Level of evidence 2. Controlled prospective comparative study. Electronic supplementary material The online version of this article (10.1007/s00405-018-4946-z) contains supplementary material, which is available to authorized users.

The combination of increased healthcare access, universal aging, and infallible therapy demands, synergistically drive the need for the development of biomaterial technologies that mitigate the challenge of biomaterial-associated infections (BAI). Staphylococcus epidermidis and Staphylococcus aureus account for the majority of BAI due to their ability to accumulate in adherent multilayered biofilm. This investigation details the development of gene expression assays to evaluate the genetic processes of attachment, accumulation, maturation, and dispersal phases of biofilms on biomaterials in vitro, while abiding by the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines. The biofilm formation of S. epidermidis on polyurethane (PU) central venous catheters and S. aureus on machined titanium (Ti) was examined in terms of gene expression at early and late time points. The results provided insight into how each stage of biofilm formation is orchestrated over time on these biomaterials in vitro. Furthermore, the results suggested that mechanical RNA extraction, organic solvents, elimination of genomic DNA, and preamplification are advisable strategies to implement for biofilm gene expression analysis. It is concluded that this method can be employed for the assessment of biofilm-biomaterial interactions at the molecular level. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3400-3412, 2017.

Analysis of the implant-tissue interface is important for an understanding of the cellular response to biomaterials with different surface characteristics. However, inaccessibility to the site has restricted the detailed evaluation of the tissue surrounding the implant. Laser microdissection enables the isolation of specific cells and tissues for subsequent DNA, RNA, or protein analysis. The present experimental study employed laser microdissection to analyze tissue-specific differences in gene expression in cells around infected or control titanium implants 72 h after subcutaneous implantation in a rat model. Three different tissue zones located 0-800 μm away from the implant-tissue interface were analyzed. Implant sites challenged with a dose of 106 CFU Staphylococcus epidermidis demonstrated higher gene expression of selected markers for inflammation (TNF-α, IL-6), cell recruitment (MCP-1, IL-8, IL-8 R), infection (TLR2), and tissue remodeling (MMP-9) compared with control implants. Furthermore, the gene expression analysis of the three extracted tissue zones revealed marked spatial differences, depending on the distance to the implant. Control implants continuously induced higher cell gene expression in the implant-tissue interface compared with cells 200-800 μm away from the implant, whereas the sites inoculated with S. epidermidis resulted in high gene expression further away from the implant as well. In conclusion, this study demonstrates that laser microdissection is an interesting tool, revealing both gene- and site-specific gene expression patterns in the implant-tissue interface. The technique provides an opportunity for detailed molecular dissection of the biological events related to the implant but occurring at different distances from the implant. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2210-2217, 2017.

Osseointegrated transfemoral amputation prostheses have proven successful as an alternative method to the conventional socket-type prostheses. The method improves prosthetic use and thus increases the demands imposed on the bone-implant system. The hypothesis of the present study was that the loads applied to the bone-anchored implant system of amputees would result in locations of high stress and strain transfer to the bone tissue and thus contribute to complications such as unfavourable bone remodeling and/or elevated inflammatory response and/or compromised sealing function at the tissue-abutment interface. In the study, site-specific loading measurements were made on amputees and used as input data in finite element analyses to predict the stress and strain distribution in the bone tissue. Furthermore, a tissue sample retrieved from a patient undergoing implant revision was characterized in order to evaluate the long-term tissue response around the abutment. Within the limit of the evaluated bone properties in the present experiments, it is concluded that the loads applied to the implant system may compromise the sealing function between the bone and the abutment, contributing to resorption of the bone in direct contact with the abutment at the most distal end. This was supported by observations in the retrieved clinical sample of bone resorption and the formation of a soft tissue lining along the abutment interface. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1113-1122, 2017.

Commercially pure titanium (cp-Ti) and titanium alloys (typically Ti6Al4V) display excellent corrosion resistance and biocompatibility. Although the chemical composition and topography are considered important, the mechanical properties of the material and the loading conditions in the host have, conventionally, influenced material selection for different clinical applications: predominantly Ti6Al4V in orthopaedics while cp-Ti in dentistry. This paper attempts to address three important questions: (i) To what extent do the surface properties differ when cp-Ti and Ti6Al4V materials are manufactured with the same processing technique?, (ii) Does bone tissue respond differently to the two materials, and (iii) Do bacteria responsible for causing biomaterial-associated infections respond differently to the two materials? It is concluded that: (i) Machined cp-Ti and Ti6Al4V exhibit similar surface morphology, topography, phase composition and chemistry, (ii) Under experimental conditions, cp-Ti and Ti6Al4V demonstrate similar osseointegration and biomechanical anchorage, and (iii) Experiments in vitro fail to disclose differences between cp-Ti and Ti6Al4V to harbour Staphylococcus epidermidis growth. No clinical comparative studies exist which could determine if long-term, clinical differences exist between the two types of bulk materials. It is debatable whether cp-Ti or Ti6Al4V exhibit superiority over the other, and further comparative studies, particularly in a clinical setting, are required.

Biomaterials are frequently used in various clinical applications, improving function, mobility, and the overall quality of life for patients. The clinical performance of bone-anchored implants is typically assessed by X-ray-based analytical technologies and various methods to evaluate the stability of the implant–bone unit. For eventual clinical translation of preclinical knowledge of tissue response to various biomaterials, it is essential to evaluate implants retrieved with associated biological tissues from humans. Information pertaining to causes of failure mechanisms and factors contributing to successful tissue integration can thus be obtained. This chapter highlights the process of implant retrieval analysis, with a focus on (1) reasons that necessitate implant retrieval, (2) recommended sample handling and preparation procedures, and (3) state-of-the-art ex vivo assessment. Furthermore, typical analyses of three different clinical applications are exemplified where microstructural and ultrastructural analysis and microbiological evaluation are performed on the same bone–implant specimen. While valuable knowledge could be gained, often only a limited amount of retrieved material is available. Therefore a carefully planned sample preparation and analytical approach is needed.

Infection constitutes a major risk for implant failure, but the reasons why biomaterial sites are more vulnerable than normal tissue are not fully elucidated. In this study, a soft tissue infection model was developed, allowing the analysis of cellular and molecular responses in each of the sub-compartments of the implant-tissue interface (on the implant surface, in the surrounding exudate and in the tissue). Smooth and nanostructured titanium disks with or without noble metal chemistry (silver, gold, palladium), and sham sites, were inoculated with Staphylococcus epidermidis and analysed with respect to number of viable bacteria, number, viability and gene expression of host cells, and using different morphological techniques after 4 h, 24 h and 72 h. Non-infected rats were controls. Results showed a transient inflammatory response at control sites, whereas bacterial administration resulted in higher recruitment of inflammatory cells (mainly polymorphonuclear), higher, continuous cell death and higher gene expression of tumour necrosis factor-alpha, interleukin-6, interleukin-8, Toll-like receptor 2 and elastase. At all time points, S. epidermidis was predominantly located in the interface zone, extra- and intracellularly, and lower levels were detected on the implants compared with surrounding exudate. This model allows detailed analysis of early events in inflammation and infection associated to biomaterials in vivo leading to insights into host defence mechanisms in biomaterial-associated infections.

Aim This study evaluates biofilm formation and barrier function against Streptococcus oralis of nonresorbable polytetrafluoroethylene (PTFE) guided bone regeneration membranes having expanded (e-PTFE) and dense (d-PTFE) microstructure. Materials and methods Three e-PTFE membranes of varying openness, one d-PTFE membrane, and commercially pure titanium discs were evaluated. All e-PTFE membranes consisted of PTFE nodes interconnected by fibrils. The d-PTFE membrane was fibril-free, with large evenly spaced indentations. The surfaces were challenged with S. oralis and incubated statically for 2-48h. Bacterial colonization, viability, and penetration were evaluated. Results S. oralis numbers increased over time on all surfaces, as observed using scanning electron microscopy, while cell viability decreased, as measured by colony forming unit (CFU) counting. At 24h and 48h, biofilms on d-PTFE were more mature and thicker (tower formations) than on e-PTFE, where fewer layers of cells were distributed mainly horizontally. Biofilms accumulated preferentially within d-PTFE membrane indentations. At 48h, greater biofilm biomass and number of viable S. oralis were found on d-PTFE compared to e-PTFE membranes. All membranes were impermeable to S. oralis cells. Conclusions All PTFE membranes were effective barriers against bacterial passage in vitro. However, d-PTFE favored S. oralis biofilm formation.

The combination of increased healthcare access, universal aging, and infallible therapy demands, synergistically drive the need for the development of biomaterial technologies that mitigate the challenge of biomaterial-associated infections (BAI). Staphylococcus epidermidis and Staphylococcus aureus account for the majority of BAI due to their ability to accumulate in adherent multilayered biofilm. This investigation details the development of gene expression assays to evaluate the genetic processes of attachment, accumulation, maturation, and dispersal phases of biofilms on biomaterials in vitro, while abiding by the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines. The biofilm formation of S. epidermidis on polyurethane (PU) central venous catheters and S. aureus on machined titanium (Ti) was examined in terms of gene expression at early and late time points. The results provided insight into how each stage of biofilm formation is orchestrated over time on these biomaterials in vitro. Furthermore, the results suggested that mechanical RNA extraction, organic solvents, elimination of genomic DNA, and preamplification are advisable strategies to implement for biofilm gene expression analysis. It is concluded that this method can be employed for the assessment of biofilm-biomaterial interactions at the molecular level. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3400-3412, 2017.

The choice of material for implanted prostheses is of great importance concerning bacterial colonization and biofilm formation. Consequently, methods to investigate bacterial behavior are needed in order to develop new infection resistant surfaces. In this study, different methodological setups were used to evaluate the antimicrobial effect of photocatalytic titanium oxide and silver surfaces. Biofilm formation and eradication under static and dynamic culture conditions were studied with the use of the following analytical techniques: viable colony-forming unit (CFU) counting, imprinting, fluorescence, and bioluminescence. The present study demonstrates that different methods are needed in order to evaluate the prophylactic and treatment effects on planktonic and biofilm bacteria and to assess the antimicrobial effect of different surface treatments/coatings. Choosing the right antibacterial testing model for the specific application is also of great importance. Both in situ approaches and indirect methods provide valuable complementary information. © 2014 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2014.

Staphylococci and enterococci account for most deep infections associated with bone-anchored percutaneous implants for amputation treatment. Implant-associated infections are difficult to treat; therefore, it is important to investigate if these infections have a biofilm origin and to determine the biofilm antimicrobial susceptibility to improve treatment strategies. The aims were: (i) to test a novel combination of the Calgary biofilm device and a custom-made susceptibility MIC plate (Sensititre

Osseointegrated transfemoral amputation prostheses have proven successful as an alternative method to the conventional socket-type prostheses. The method improves prosthetic use and thus increases the demands imposed on the bone-implant system. The hypothesis of the present study was that the loads applied to the bone-anchored implant system of amputees would result in locations of high stress and strain transfer to the bone tissue and thus contribute to complications such as unfavourable bone remodeling and/or elevated inflammatory response and/or compromised sealing function at the tissue-abutment interface. In the study, site-specific loading measurements were made on amputees and used as input data in finite element analyses to predict the stress and strain distribution in the bone tissue. Furthermore, a tissue sample retrieved from a patient undergoing implant revision was characterized in order to evaluate the long-term tissue response around the abutment. Within the limit of the evaluated bone properties in the present experiments, it is concluded that the loads applied to the implant system may compromise the sealing function between the bone and the abutment, contributing to resorption of the bone in direct contact with the abutment at the most distal end. This was supported by observations in the retrieved clinical sample of bone resorption and the formation of a soft tissue lining along the abutment interface. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1113-1122, 2017.

Summary: Autologous bone or inert alloplastic materials used in cranial reconstructions are techniques that are associated with resorption, infection, and implant exposure. As an alternative, a calcium phosphate–based implant was developed and previously shown to potentially stimulate bone growth. We here uncover evidence of induced bone formation in 2 patients. Histological examination 9 months postoperatively showed multinuclear cells in the central defect zone and bone ingrowth in the bone-implant border zone. An increased expression of bone-associated markers was detected. The other patient was investigated 50 months postoperatively. Histological examination revealed ceramic materials covered by vascularized compact bone. The bone regenerative effect induced by the implant may potentially improve long-term clinical outcome compared with conventional techniques, which needs to be verified in a clinical study.

The choice of material for implanted prostheses is of great importance concerning bacterial colonization and biofilm formation. Consequently, methods to investigate bacterial behavior are needed in order to develop new infection resistant surfaces. In this study, different methodological setups were used to evaluate the antimicrobial effect of photocatalytic titanium oxide and silver surfaces. Biofilm formation and eradication under static and dynamic culture conditions were studied with the use of the following analytical techniques: viable colony-forming unit (CFU) counting, imprinting, fluorescence, and bioluminescence. The present study demonstrates that different methods are needed in order to evaluate the prophylactic and treatment effects on planktonic and biofilm bacteria and to assess the antimicrobial effect of different surface treatments/coatings. Choosing the right antibacterial testing model for the specific application is also of great importance. Both in situ approaches and indirect methods provide valuable complementary information.

Nanometer scale surface features on implants and prostheses can potentially be used to enhance osseointegration and may also add further functionalities, such as infection resistance, to the implant. In this study, a nanostructured noble metal coating consisting of palladium, gold and silver, never previously used in bone applications, was applied to machined titanium screws to evaluate osseointegration after 6 and 12weeks in rabbit tibiae and femurs. Infection resistance was confirmed by in vitro adhesion test. A qualitatively and quantitatively similar in vivo bone response was observed for the coated and uncoated control screws, using histology, histomorphometry and electron microscopy. The bone-implant interface analysis revealed an extensive bone formation and direct bone-implant contact. These results demonstrate that the nanostructured noble metal coating with antimicrobial properties promotes osseointegration and may therefore be used to add extra implant functionality in the form of increased resistance to infection without the use of antibiotics. From the Clinical Editor The authors of this paper demonstrate that nanostructured noble metal coating of implants and prostheses used in orthopedic procedures promotes osseointegration and may be used to add extra implant functionality in the form of increased resistance to infection without the use of antibiotics.

The sul2 gene encodes sulphonamide resistance (SulR) and is commonly found in Escherichia coli from different hosts. We typed E. coli isolates by multilocus sequence typing (MLST) and compared the results to sequence variation of sul2, in order to investigate the relation to host origin of pathogenic and commensal E. coli strains and to investigate whether transfer of sul2 into different genomic lineages has happened multiple times. Sixty-eight E. coli isolated in Denmark and Norway from different hosts and years were MLST typed and sul2 PCR products were sequenced and compared. PFGE was performed in a subset of isolates. All isolates were divided into 45 different sequence types (STs), with clonal complexes CC10, CC23, CC168, CC350 and CC69 being the most frequent. The sul2 gene from the majority of E. coli strains had only two point mutations, at positions 159 and 197, leading to a synonymous and a non-synonymous change, respectively. Five strains had extra single mutations. All poultry, poultry meat, and Danish human blood isolates had the same sul2 ST and some of these strains clustered under the same MLST STs, indicating that they shared habitats. Most PFGE profiles clustered according to source, but some included different sources. SulR E. coli from different animals, food, human faeces and infections did not cluster according to their origin, suggesting that these habitats share E. coli and sul2 gene types. However, while pig isolates on one occasion clustered with urinary tract infection isolates, poultry isolates seemed more related to isolates from bloodstream infections in humans. Presence of mainly two types of the sul2 gene in both human and animal isolates, irrespective of date and geography, and the presence of both types in the same clonal lineages, suggest horizontal transfer of sul2.

Objectives: The aim of this study was to investigate whether the sulphonamide resistance gene sul2 could be transferred between Escherichia coli in the human gut. Methods: Nine volunteers ingested a 10 9 cfu suspension of sulphonamide-susceptible, rifampicinresistant E. coli recipients of human origin. Three hours later, they ingested a 10 7 cfu suspension of a sulphonamide-resistant (MIC >1024 mg/L) E. coli donor of pig origin. Stool samples were collected 24 h prior to ingestion, daily for 7 days and at days 14 and 35. Samples were plated on selective plates and monitored for the acquisition of sulphonamide-resistance by the recipient from the indigenous or administrated donor E. coli. Possible transconjugants were typed by PFGE and tested for the presence of plasmids containing the sul2 gene, which was also sequenced. Results: Concentrations of the human and animal E. coli reached a maximum of 7.5 3 10 6 cfu/g faeces and colonized for more than 7 days, and 2 3 10 8 cfu/g for more than 14 days, respectively. On day 2, a transconjugant was detected in one volunteer. This volunteer was colonized with sulphonamideresistant E. coli at day 0. The transconjugant was sul2-positive, had an MIC >1024 mg/L for sulfamethoxazole and the same PFGE profile as the recipient. The resident E. coli transferred a plasmid (>63 kb), containing the sul2 gene, to the recipient. The sul2 sequence of the transconjugant was identical to that of the volunteer’s own E. coli from day 0, but differed from the animal strain. Co-transfer of ampicillin resistance was also demonstrated. Conclusions: Transfer of sul2 was observed between E. coli bacteria in the human intestine. The transconjugant’s sul2 gene came from the volunteer’s own flora. The origin of the E. coli donor is unknown.