Microbiological and ultrastructural evaluation of bacteriophage 191219 against planktonic, intracellular and biofilm infection with Staphylococcus aureus.

Infections of orthopaedic implants, such as fracture fixation devices and total-joint prostheses, are devastating complications. Staphylococcus aureus (S. aureus) is a predominant pathogen causing orthopaedic-implant biofilm infections that can also internalise and persist in osteoblasts, thus resisting antibiotic therapy. Bacteriophages are a promising alternative treatment approach. However, data on the activity of bacteriophages against S. aureus, especially during intracellular growth, and against in vivo biofilm formation on metals are scarce. Therefore, the present study evaluated the in vitro efficacy of S. aureus bacteriophage 191219, alone as well as in combination with gentamicin and rifampicin, to eradicate S. aureus strains in their planktonic stage, during biofilm formation and after internalisation into osteoblasts. Further, the invertebrate model organism Galleria mellonella was used to assess the activity of the bacteriophage against S. aureus biofilm on metal implants with and without antibiotics. Results demonstrated the in vitro efficacy of bacteriophage 191219 against planktonic S. aureus. The phage was also effective against in vitro S. aureus biofilm formation in a dose-dependent manner and against S. aureus internalised in an osteoblastic cell line. Transmission electron microscopy (TEM) analysis showed bacteriophages on S. aureus inside the osteoblasts, with the destruction of the intracellular bacteria and formation of new bacteriophages. For the Galleria mellonella infection model, single administration of phage 191219 failed to show an improvement in survival rate but appeared to show a not statistically significant enhanced effect with gentamicin or rifampicin. In summary, bacteriophages could be a potential adjuvant treatment strategy for patients with implant-associated biofilm infections.

Vertebral osteomyelitis is characterised by increased RANK/OPG and RANKL/OPG expression ratios in vertebral bodies and intervertebral discs.

Vertebral osteomyelitis (VO) is an infection of the spine mainly caused by bacterial pathogens. The pathogenesis leading to destruction of intervertebral discs (IVDs) and adjacent vertebral bodies (VBs) is poorly described. The present study aimed at investigating the connection between infection and bone/disc metabolism in VO patients. 14 patients with VO (infection group) and 14 patients with burst fractures of the spine (fracture group; control) were included prospectively. Tissue biopsies from affected IVDs and adjacent VBs were analysed by RT-qPCR for mRNA-expression levels of 18 target genes including chemokines, adipokines and genes involved in bone metabolism. Most importantly, the receptor activator of NF-κB/osteoprotegerin (RANK/OPG) expression ratio was drastically elevated in both VBs and IVDs of the infection group. In parallel, expression of genes of the prostaglandin-E2-dependent prostanoid system was induced. Such genes regulate tissue degradation processes via the triad OPG/RANK/RANKL as well as via the chemokines IL-8 and CCL-20, whose expression was also found to be increased upon infection. The gene expression of the adipokine leptin, which promotes inflammatory tissue degradation, was higher in IVD tissue of the infection group, whereas the transcription of omentin and resistin genes, whose functions are largely unknown in the context of infectious diseases, was lower in infected VBs. In summary, similar expression patterns of pro-inflammatory cytokines and pro-osteoclastogenic factors were identified in VBs and IVDs of patients suffering from VO. This suggests that common immuno-metabolic pathways are involved in the mechanisms leading to tissue degradation in VBs and IVDs during VO.

Extensor tendon ruptures in rheumatoid wrists.

BACKGROUND AND AIMS: Rheumatoid arthritis is a chronic inflammatory disease. The associated involvement of hands and tendons is over 90% and impairs overall function. In the course of the disease, the joints are often operated on. During this operation, ruptures of the extensor tendons are found by chance without the patients noticing them. The aim of this retrospective study is the prevalence of extensor tendon rupture. Which tendon is destroyed most frequently? How can the functional outcome be measured after reconstruction? MATERIALS AND METHODS: From 1572 operations on rheumatoid wrists, 61 extensor tendon ruptures were identified in 41 patients. The average time between the first rheumatic symptoms of the hand and surgery was 6.4 years. The average duration of RA was 7.8 years. 26 patients with 27 tendon reconstructions were included in the follow-up with an average postoperative duration of 4.6 years (3 to 14.2 years). RESULTS: Extensor tendons ruptures typically occurred at mechanically stressed sites. The most frequent rupture was found in the extensor pollicis longus tendon (21 tendons), followed by the small finger extensor tendon (14 tendons). A transfer was performed on 7 tendons. Fifty-five tendon lesions were sutured at other intact tendons. Free grafts were not used. The results in Clayton and QuickDASH scores were significantly different. Functional improvement was consistent with the results of tendon reconstructions in healthy control groups. CONCLUSION: In rheumatoid patients, a rupture of an extensor tendon must be expected at 4%. Patients tolerate and compensate this damage for a long time. The function of the hand including the tendon function is the most important factor in assessing the success of the operation. The subjective patient acceptance depends on the progress of the underlying disease, postoperative care (ergotherapy, physiotherapy, orthosis) and the patients' demands.

Successful Total Elbow Replacement after Septic Arthritis with Staphylococcus aureus- a Case Report and Review of the Literature.

Septic arthritis of the elbow seems to be a contraindication for total elbow arthroplasty (TEA). We here describe a 65-year-old male, American Society of Anesthesiologists (ASA) class 3 - patient, with a severely destructed right elbow due to septic arthritis with Staphylococcus aureus. His treatment consisted of multiple irrigation and debridement procedures including resection of the distal humerus, soft tissue coverage by local rotational flap and the use of a gentamicin-vancomycin loaded PMMA spacer, i.v. and oral antibiotics. After eradication of infection, a constraint cemented TEA could then successfully be performed eight months after the initial surgery and twenty-five weeks after the last debridement procedure. Twenty-one months after the TEA, the patient remained infect free and shows excellent functional results: Disabilities of the Arm, Shoulder and Hand (DASH) score: 38.3, Broberg and Morrey score: 91/100, Mayo elbow score: 95/100. To the best of our knowledge this is the first case in the literature that demonstrates TEA after septic elbow arthritis with Staphylococcus aureus. Although TEA is known as a typical surgical procedure with a low volume in numbers and higher complication rates, such as elevated infection rates compared to other types of arthroplasty, septic arthritis with Staphylococcus aureuscan successfully be performed after eradication of the infection and targeted antibiotic therapy. Key words:total elbow arthroplasty, total elbow replacement, septic arthritis, Staphylococcus aureus.

FOXO3-mediated chemo-protection in high-stage neuroblastoma depends on wild-type TP53 and SESN3.

Forkhead box O class transcription factors are homeostasis regulators that control cell death, longevity and therapy-resistance. In neuroblastoma (NB), nuclear FOXO3 correlates with stage M disease and poor prognosis. To analyze whether FOXO3 contributes to drug-resistance in this childhood cancer, we investigated how different high-stage-derived NB cells respond to the activation of an ectopic FOXO3 allele. We found endogenous FOXO3 mostly localized to the nucleus-upon activation of an ectopic, 4OHT-activated FOXO3(A3)ER fusion protein two of the cell lines underwent apoptosis, whereas in the others FOXO3-activation even increased survival during drug-treatment. In the latter cell type, FOXO3 did not induce the BH3-only protein BCL2L11/BIM due to impaired binding of FOXO3 to the BIM-promoter, but still activated other FOXO3 targets. It was shown before that FOXO3 and TP53 physically interact with each other at two different regions-the TP53-N-terminus binds to the FOXO3-DNA binding domain (DBD) and the FOXO3-C-terminus interacts with the TP53-DBD. Interestingly, cell lines that undergo FOXO3-induced cell death carry homozygous point mutations in the TP53-DBD near the structural hotspot-mutation-site R175H, which abrogated FOXO3-TP53 interaction. In contrast, in FOXO3-death-resistant cells no point mutations in the TP53-DBD were found-in these cells FOXO3-TP53 complexes are formed and FOXO3-binding to the BIM-promoter, but not the induction of the detoxifying protein SESN3, were prevented, which in turn increased chemo-protection in this type of high-stage-derived NB cells. Our combined data suggest that FOXO3 steps in as a death inducer in case of TP53-mutation, whereas functional TP53 alters FOXO3-target-promoter-recognition, which prevents death induction by FOXO3 and instead increases chemo-protection and survival of NB cells. This novel mechanism may explain the low incidence of TP53 mutation in high-stage NB at diagnosis and suggests FOXO3 as a therapeutic target for this childhood malignancy.

Evaluation of a user guidance reminder to improve the quality of electronic prescription messages.

BACKGROUND: Prescribers' inappropriate use of the free-text Notes field in new electronic prescriptions can create confusion and workflow disruptions at receiving pharmacies that often necessitates contact with prescribers for clarification. The inclusion of inappropriate patient direction (Sig) information in the Notes field is particularly problematic. OBJECTIVE: We evaluated the effect of a targeted watermark, an embedded overlay, reminder statement in the Notes field of an EHR-based e-prescribing application on the incidence of inappropriate patient directions (Sig) in the Notes field. METHODS: E-prescriptions issued by the same exact cohort of 97 prescribers were collected over three time periods: baseline, three months after implementation of the reminder, and 15 months post implementation. Three certified and experienced pharmacy technicians independently reviewed all e-prescriptions for inappropriate Sig-related information in the Notes field. A physician reviewer served as the final adjudicator for e-prescriptions where the three reviewers could not reach a consensus. ANOVA and post hoc Tukey HSD tests were performed on group comparisons where statistical significance was evaluated at p<0.05. RESULTS: The incidence of inappropriate Sig-related information in the Notes field decreased from a baseline of 2.8% to 1.8% three months post-implementation and remained stable after 15 months. In addition, prescribers' use of the Notes decreased by 22% after 3 months and had stabilized at 18.7% below baseline after 15 months. CONCLUSION: Insertion of a targeted watermark reminder statement in the Notes field of an e-prescribing application significantly reduced the incidence of inappropriate Sig-related information in Notes and decreased prescribers' use of this field.

Lake Malawi cichlid evolution along a benthic/limnetic axis.

Divergence along a benthic to limnetic habitat axis is ubiquitous in aquatic systems. However, this type of habitat divergence has largely been examined in low diversity, high latitude lake systems. In this study, we examined the importance of benthic and limnetic divergence within the incredibly species-rich radiation of Lake Malawi cichlid fishes. Using novel phylogenetic reconstructions, we provided a series of hypotheses regarding the evolutionary relationships among 24 benthic and limnetic species that suggests divergence along this axis has occurred multiple times within Lake Malawi cichlids. Because pectoral fin morphology is often associated with divergence along this habitat axis in other fish groups, we investigated divergence in pectoral fin muscles in these benthic and limnetic cichlid species. We showed that the eight pectoral fin muscles and fin area generally tended to evolve in a tightly correlated manner in the Lake Malawi cichlids. Additionally, we found that larger pectoral fin muscles are strongly associated with the independent evolution of the benthic feeding habit across this group of fish. Evolutionary specialization along a benthic/limnetic axis has occurred multiple times within this tropical lake radiation and has produced repeated convergent matching between exploitation of water column habitats and locomotory morphology.

Design considerations to improve cognitive ergonomic issues of unmanned vehicle interfaces utilizing video game controllers.

Unmanned (UAVs, UCAVs, and UGVs) systems still have major human factors and ergonomic challenges related to the effective design of their control interface systems, crucial to their efficient operation, maintenance, and safety. Unmanned system interfaces with a human centered approach promote intuitive interfaces that are easier to learn, and reduce human errors and other cognitive ergonomic issues with interface design. Automation has shifted workload from physical to cognitive, thus control interfaces for unmanned systems need to reduce mental workload on the operators and facilitate the interaction between vehicle and operator. Two-handed video game controllers provide wide usability within the overall population, prior exposure for new operators, and a variety of interface complexity levels to match the complexity level of the task and reduce cognitive load. This paper categorizes and provides taxonomy for 121 haptic interfaces from the entertainment industry that can be utilized as control interfaces for unmanned systems. Five categories of controllers were based on the complexity of the buttons, control pads, joysticks, and switches on the controller. This allows the selection of the level of complexity needed for a specific task without creating an entirely new design or utilizing an overly complex design.

Determining skill transferability of action games as a method to reduce in-vehicle phone distractions.

Distracted driving has been shown to be a safety issue in numerous studies. To combat this problem, in-vehicle technology, legislation, media interventions, and other methods have been proposed and attempted. However research indicates that the drivers themselves may circumvent, ignore, or not be able to react in time for these interventions to be effective. Therefore research into training programs for drivers may improve reaction time under distraction. Research indicates that action game players have faster reaction times and more attentional resources than non-players on paper-based tests. However, transferability to driving has not been studied yet. This paper outlines a study to determine if action game players perform better at a driving task based on frequency of game-play. Participants will be placed into two groups of play (high vs. low) and tested against two levels of distraction (none vs. phone conversation). It is expected that participants who play higher frequency of action games will perform better under distraction than lower frequency players. Driver performance, conversation recall, frequency and durations of eye fixations will be analyzed based on previous research which has validated those variables as a measure of distraction and higher workload.

Inappropriate intravascular device use: a prospective study.

Despite the clinical significance of complications due to intravascular catheters, the inappropriate use of intravascular catheters in hospitalised patients has not been adequately characterised. The objective of this prospective observational study was to develop definitions for appropriate intravascular device use, to estimate the frequency of inappropriate use of intravascular devices, and to examine risk factors and outcomes associated with inappropriate use in hospitalised patients. Among 436 patients admitted between October and December 2007, a total of 2909 hospitalisation days and use of 876 intravascular devices was observed. Of the 3806 total catheter-days recorded, 1179 (31%) were found to be inappropriate based on the study criteria. Logistic regression analysis indicated that age, total number of catheters used and total duration of catheterisation were risk factors for inappropriate device use (P<0.05). Inappropriate usage was strongly associated with increased intensive care unit admission (P<0.05) and length of hospital stay (4.9±4.3 days for appropriate vs 8.5±12.6 days for inappropriate; P<0.05). Use of central venous catheters was not a predictor for inappropriate device use. Inappropriate intravascular device use is a very common phenomenon in hospitalised patients and is strongly linked to adverse device-related outcomes. These results may be used to develop strategies to systematically reduce excessive intravascular device use which would be expected to reduce adverse events associated with morbidity, mortality, and excess healthcare costs.

The presence of icaADBC is detrimental to the colonization of human skin by Staphylococcus epidermidis.

Previous studies have demonstrated that Staphylococcus epidermidis isolates colonizing the skin of healthy humans do not typically encode icaADBC, the genes responsible for the production of polysaccharide intercellular adhesin or biofilms. It was therefore hypothesized that the presence of icaADBC was deleterious to the successful colonization of human skin by S. epidermidis. Using a human skin competition model, it was determined that the strong biofilm-producing S. epidermidis strain 1457 was outcompeted at 1, 3, and 10 days by an isogenic icaADBC mutant (1457 ica::dhfr), suggesting a fitness cost for carriage of icaADBC.

Staphylococcus epidermidis saeR is an effector of anaerobic growth and a mediator of acute inflammation.

The saeRS two-component regulatory system regulates transcription of multiple virulence factors in Staphylococcus aureus. In the present study, we demonstrated that the saePQRS region in Staphylococcus epidermidis is transcriptionally regulated in a temporal manner and is arranged in a manner similar to that previously described for S. aureus. Studies using a mouse foreign body infection model demonstrated that the virulence of strain 1457 and the virulence of a mutant, strain 1457 saeR, were statistically equivalent. However, histological analyses suggested that the polymorphonuclear neutrophil response at 2 days postinfection was significantly greater in 1457-infected mice than in 1457 saeR-infected mice, demonstrating that SaeR influences the early, acute phases of infection. Microarray analysis demonstrated that a saeR mutation affected the transcription of 65 genes (37 genes were upregulated and 28 genes were downregulated); in particular, 8 genes that facilitate growth under anaerobic conditions were downregulated in 1457 saeR. Analysis of growth under anaerobic conditions demonstrated that 1457 saeR had a decreased growth rate compared to 1457. Further metabolic experiments demonstrated that 1457 saeR had a reduced capacity to utilize nitrate as a terminal electron acceptor and exhibited increased production of lactic acid in comparison to 1457. These data suggest that in S. epidermidis SaeR functions to regulate the transition between aerobic growth and anaerobic growth. In addition, when grown anaerobically, 1457 saeR appeared to compensate for the redox imbalance created by the lack of electron transport-mediated oxidation of NADH to NAD+ by increasing lactate dehydrogenase activity and the subsequent oxidation of NADH.

SigmaB and SarA independently regulate polysaccharide intercellular adhesin production in Staphylococcus epidermidis.

The production of polysaccharide intercellular adhesin (PIA) is an essential process in foreign body infections mediated by Staphylococcus epidermidis. Transcriptional regulation of the icaADBC operon, the genes responsible for production of enzymes that synthesize PIA, is multi-factorial and involves at least SarA and sigmaB. Transcriptional and promoter fusion studies revealed that the decreased transcription of the icaADBC operon observed in a S. epidermidis 1457 sigB mutant is not mediated through a direct interaction of sigmaB-RNA polymerase at the icaADBC promoter region but instead through the upregulation of IcaR, a known repressor of icaADBC transcription. Transcriptional analysis of a 1457 sigB-icaR double mutant confirmed that the decreased icaADBC transcript in 1457 sigB is IcaR dependent. Furthermore, primer extension studies suggest that the icaR promoter appears to be sigmaA dependent, suggesting that sigmaB indirectly controls icaR transcription through an unknown pathway. In addition, it was confirmed that the loss of SarA results in the loss of icaADBC transcription and PIA production in S. epidermidis. It was further demonstrated, through the over-production of SarA in 1457 sigB, that the loss of sarP1 promoter activity in 1457 sigB has little or no effect on the loss of PIA production in this mutant. Finally, it was demonstrated that PIA production could be restored in both 1457 sigB and 1457 sarA by complementing these mutants with a full-length icaADBC operon controlled by a cadmium-inducible noncognate promoter. It is concluded that sigmaB and SarA operate independently of each other to regulate PIA production and biofilm development in S. epidermidis.

Adherence of Staphylococcus epidermidis to biomaterials is augmented by PIA.

Staphylococcus epidermidis is the most common cause of orthopaedic prosthetic device infections. Polysaccharide intercellular adhesin (PIA) is important in the pathogenesis of intravascular catheter-associated infection, and has an essential role in cellular aggregation and biofilm formation. However, the role of PIA in orthopaedic infections is less well understood. We used genetically defined strains of S. epidermidis in an in vitro adherence assay to assess the importance of PIA in the adherence to various orthopaedic biomaterials. On all biomaterials tested (zirconia, ultra-high molecular weight polyethylene, polymethylmethacrylate, cobalt chromium, titanium, stainless steel, and silastic), PIA-positive S. epidermidis 1457 exhibited greater levels of adherence thanS. epidermidis 1457 M10, an isogenic icaA Tn917 mutant. PIA appears to play a critical role in the adherence of S. epidermidis to orthopaedic biomaterials, and may serve as an important virulence determinant in orthopaedic prosthetic device infections.

Genetic and phenotypic analysis of biofilm phenotypic variation in multiple Staphylococcus epidermidis isolates.

Production of biofilm in Staphylococcus epidermidis is mediated through enzymes produced by the four-gene operon ica and is subject to phenotypic variation. The purpose of these experiments was to investigate the regulation of ica and icaR transcription in phenotypic variants produced by multiple unrelated isolates of S. epidermidis. Ten isolates were chosen for the study, four of which contained IS256. IS256 mediates a reversible inactivation of ica in approximately 30 % of phenotypic variants. All ten strains produced at least two types of phenotypic variant (intermediate and smooth) in which biofilm formation was significantly impaired. Reversion studies indicated that all phenotypic variants were stable after overnight growth, but began to revert to other phenotypic forms after 5 days of incubation at 37 degrees C. ica transcriptional analysis was performed on phenotypic variants from three IS256-negative isolates; 1457, SE5 and 14765. This analysis demonstrated that ica transcription was significantly reduced in the majority of phenotypic variants, although two variants from SE5 and 1457 produced wild-type quantities of ica transcript. Analysis of seven additional phenotypic variants from SE5 revealed that ica expression was only reduced in three. Expression of icaR transcript was unaffected in all smooth phenotypic variants. Mutations within ica were identified in two SE5 variants with wild-type levels of ica transcription. It is concluded that mutation and transcriptional regulation of ica are the primary mechanisms that govern phenotypic variation of biofilm formation within IS256-negative S. epidermidis.

Acarbose reduces the risk for myocardial infarction in type 2 diabetic patients: meta-analysis of seven long-term studies.

AIMS: To assess if treatment with the alpha-glucosidase inhibitor acarbose can reduce cardiovascular events in type 2 diabetic patients. METHODS AND RESULTS: This meta-analysis included seven randomized, double-blind, placebo-controlled acarbose studies with a minimum treatment duration of 52 weeks. Type 2 diabetic patients valid for safety were randomized to either acarbose (n=1248) or placebo (n=932). The primary outcome measure was the time to develop a cardiovascular event. Primary analysis was conducted using Cox regression analysis. The effect of acarbose on metabolic parameters was also investigated. Acarbose therapy showed favourable trends towards risk reduction for all selected cardiovascular event categories. The treatment significantly reduced the risk for "myocardial infarction" (hazards ratio=0.36 [95% Cl 0.16-0.80], P=0.0120) and "any cardiovascular event" (0.65 [95% Cl 0.48-0.88], P=0.0061). Glycaemic control, triglyceride levels, body weight and systolic blood pressure also improved significantly during acarbose treatment. CONCLUSION: Intervention with acarbose can prevent myocardial infarction and cardiovascular disease in type 2 diabetic patients while most of them are already on intensive concomitant cardiovascular medication.

Comparative molecular analysis of community- or hospital-acquired methicillin-resistant Staphylococcus aureus.

Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) is a growing public health concern that has been associated with pediatric fatalities. It is hypothesized that the evolution of CA-MRSA is a recent event due to the acquisition of mec DNA by previously methicillin-susceptible strains that circulated in the community. This study investigated the genetic relatedness between CA-MRSA, hospital-associated MRSA (HA-MRSA), and nonmenstrual toxic shock syndrome (nmTSS) isolates. Thirty-one of 32 CA-MRSA isolates were highly related as determined by pulsed-field gel electrophoresis and spa typing yet were distinguishable from 32 HA-MRSA strains. The 31 related CA-MRSA isolates produced either staphylococcal enterotoxin B (n = 5) or C (n = 26), and none made TSS toxin 1. All CA-MRSA isolates tested contained a type IV staphylococcal cassette chromosome mec (SCCmec) element. In comparison, none of the HA-MRSA isolates (n = 32) expressed the three superantigens. Antibiotic susceptibility patterns were different between the CA-MRSA and HA-MRSA isolates; CA-MRSA was typically resistant only to beta-lactam antibiotics. Six of twenty-one nmTSS isolates were indistinguishable or highly related to the CA-MRSA isolates. MnCop, an nmTSS isolate obtained in Alabama in 1986, was highly related to the CA-MRSA isolates except that it did not contain an SCCmec element. These data suggest that CA-MRSA strains may represent a new acquisition of SCCmec DNA in a previously susceptible genetic background that was capable of causing nmTSS. CA-MRSA poses a serious health risk not only because it is resistant to the antibiotics of choice for community-acquired staphylococcal infections but also because of its ability to cause nmTSS via superantigen production.

Health-related quality of life and healthcare resource utilization by COPD patients: a comparison of three instruments.

The aim of this study was to compare the SF-36 and two disease-specific instruments--St. George's Respiratory Questionnaire (SGRQ) and Chronic Respiratory Disease Questionnaire (CRDQ) among chronic obstructive pulmonary disease (COPD) patients using healthcare utilization data. The three instruments were administered twice by telephone interview to 40 COPD patients separated by a 3-month interval. Healthcare utilization data were obtained for the 3 months immediately prior to each administration. Reliabilities for sub-scales on the three instruments ranged from 0.70 to 0.90 for the SF-36, from 0.78 to 0.85 for the SGRQ, and from 0.64 to 0.91 for the CRDQ. In general, the study sample had poor physical health, and somewhat better mental health. None of the demographic variables were significantly associated with either health status or utilization. Among the three instruments, the SF-36 explained the most variance in healthcare utilization. The aggregate scores such as the summary scores of the SF-36 and the total score of the SGRQ did not perform as well as the individual sub-scales of the respective instruments in explaining the variation in utilization. The conclusion that the SF-36 provides the best association to healthcare utilization data from among the three instruments studied should be interpreted with caution given the small sample size and relatively brief duration of the study.