The prevention of infection: 12 modifiable risk factors.

AIMS: Prosthetic joint infection (PJI) remains a serious complication that is associated with high morbidity and costs. The aim of this study was to prepare a systematic review to examine patient-related and perioperative risk factors that can be modified in an attempt to reduce the rate of PJI. MATERIALS AND METHODS: A search of PubMed and MEDLINE was conducted for articles published between January 1990 and February 2018 with a combination of search terms to identify studies that dealt with modifiable risk factors for reducing the rate of PJI. An evidence-based review was performed on 12 specific risk factors: glycaemic control, obesity, malnutrition, smoking, vitamin D levels, preoperative Staphylococcus aureus screening, the management of anti-rheumatic medication, perioperative antibiotic prophylaxis, presurgical skin preparation, the operating room environment, irrigant options, and anticoagulation. RESULTS: Poor glycaemic control, obesity, malnutrition, and smoking are all associated with increased rates of PJI. Vitamin D replacement has been shown in preliminary animal studies to decrease rates of PJI. Preoperative Staphylococcus aureus screening and appropriate treatment results in decreased rates of PJI. Perioperative variables, such as timely and appropriate dosage of prophylactic antibiotics, skin preparation with chlorohexidine-based solution, and irrigation with dilute betadine at the conclusion of the operation, have all been associated with reduced rates of PJI. Similarly, aggressive anticoagulation and increased operating room traffic should be avoided to help minimize risk of PJI. CONCLUSION: PJI remains a serious complication of arthroplasty. Surgeons should be vigilant of the modifiable risk factors that can be addressed in an attempt to reduce the risk of PJI.

High-throughput screening of bacterial pathogens in clinical specimens using 16S rDNA qPCR and fragment analysis.

Molecular-based detection of bacterial pathogens directly from clinical specimens permits rapid initiation of effective antimicrobial treatment and adequate patient management. Broad-range polymerase chain reaction (PCR) amplification of the 16S rRNA gene (16S rDNA qPCR) is used in many diagnostic laboratories as a complement to cultural identification of bacterial pathogens. However, efforts for automation of 16S rDNA PCR workflows are needed in order to reduce turnaround times and to enhance reproducibility and standardization of the technique. In this retrospective method evaluation study, clinical specimens (N = 499) from patients with suspected bacterial infections were used to evaluate 2 diagnostic semiautomated workflows for rapid bacterial pathogen detection. The workflows included automated DNA extraction (QIASymphony), 16S rDNA qPCR, fragment or melting curve analysis, and amplicon sequencing. Our results support the use of the 16S rDNA qPCR and fragment analysis workflow as it enabled rapid and accurate identification of bacterial pathogens in clinical specimens.

What are the implications of withholding total joint arthroplasty in the morbidly obese? A prospective, observational study.

AIMS: The aim of this study was to observe the implications of withholding total joint arthroplasty (TJA) in morbidly obese patients. PATIENTS AND METHODS: A total of 289 morbidly obese patients with end-stage osteoarthritis were prospectively followed. There were 218 women and 71 men, with a mean age of 56.3 years (26.7 to 79.1). At initial visit, patients were given information about the risks of TJA in the morbidly obese and were given referral information to a bariatric clinic. Patients were contacted at six, 12, 18, and 24 months from initial visit. RESULTS: The median body mass index (BMI) at initial visit was 46.9 kg/m2 (interquartile range (IQR) 44.6 to 51.3). A total of 82 patients (28.4%) refused to follow-up or answer phone surveys, and 149 of the remaining 207 (72.0%) did not have surgery. Initial median BMI of those 149 was 47.5 kg/m2 (IQR 44.6 to 52.5) and at last follow-up was 46.7 kg/m2 (IQR 43.4 to 51.2). Only 67 patients (23.2%) went to the bariatric clinic, of whom 14 (20.9%) had bariatric surgery. A total of 58 patients (20.1%) underwent TJA. For those 58, BMI at initial visit was 45.3 kg/m2 (IQR 43.7 to 47.2), and at surgery was 42.3 kg/m2 (IQR 38.1 to 46.5). Only 23 patients (39.7%) of those who had TJA successfully achieved BMI < 40 kg/m2 at surgery. CONCLUSION: Restricting TJA for morbidly obese patients does not incentivize weight loss prior to arthroplasty. Only 20.1% of patients ultimately underwent TJA and the majority of those remained morbidly obese. Better resources and coordinated care are required to optimize patients prior to surgery. Cite this article: Bone Joint J 2019;101-B(7 Supple C):28-32.

Development and evaluation of a molecular assay for detection of nontuberculous mycobacteria by use of the cobas amplicor platform.

We have developed and evaluated a semiautomated assay for detection of nontuberculous mycobacteria (NTM) from clinical samples based on the Cobas Amplicor Mycobacterium tuberculosis test (Roche Diagnostics, Switzerland). A capture probe, specific for mycobacteria at the genus level, was linked to magnetic beads and used for the detection of amplification products obtained by the Cobas Amplicor M. tuberculosis assay. We demonstrate that the analytical sensitivity of the genus assay is similar to that of Cobas Amplicor M. tuberculosis detection. Four hundred sixteen clinical specimens were evaluated for the presence of NTM DNA. Sensitivities for smear-positive and smear-negative specimens were found to be 100% and 47.9%, respectively. Specificity was 97.7%, the positive predictive value 84.6%, and the negative predictive value 93.1%. The genus assay is easy to perform, produces reliable results, and was found to be a valuable diagnostic tool for rapid diagnosis of infections with NTM. The genus assay has the potential to detect NTM not routinely recovered by culture and to discover new mycobacterial species.

Molecular detection of fungal pathogens in clinical specimens by 18S rDNA high-throughput screening in comparison to ITS PCR and culture.

The rising incidence of invasive fungal infections and the expanding spectrum of fungal pathogens makes early and accurate identification of the causative pathogen a daunting task. Diagnostics using molecular markers enable rapid identification of fungi, offer new insights into infectious disease dynamics, and open new possibilities for infectious disease control and prevention. We performed a retrospective study using clinical specimens (N = 233) from patients with suspected fungal infection previously subjected to culture and/or internal transcribed spacer (ITS) PCR. We used these specimens to evaluate a high-throughput screening method for fungal detection using automated DNA extraction (QIASymphony), fungal ribosomal small subunit (18S) rDNA RT-PCR and amplicon sequencing. Fungal sequences were compared with sequences from the curated, commercially available SmartGene IDNS database for pathogen identification. Concordance between 18S rDNA RT-PCR and culture results was 91%, and congruence between 18S rDNA RT-PCR and ITS PCR results was 94%. In addition, 18S rDNA RT-PCR and Sanger sequencing detected fungal pathogens in culture negative (N = 13) and ITS PCR negative specimens (N = 12) from patients with a clinically confirmed fungal infection. Our results support the use of the 18S rDNA RT-PCR diagnostic workflow for rapid and accurate identification of fungal pathogens in clinical specimens.

High affinity binding and direct antiproliferative effects of LHRH analogues in human ovarian cancer cell lines.

Recently, specific binding sites for luteinizing hormone releasing hormone (LHRH) and its analogues have been demonstrated in biopsy samples of human epithelial ovarian cancer. Their biological significance remained obscure. In this study we ascertained whether such LHRH-binding sites are also present in the human epithelial ovarian cancer cell lines EFO-21 and EFO-27 and if they could mediate antiproliferative effects of LHRH analogues. Using [125I, D-Trp6]LHRH, a high affinity/low capacity binding site was detected in both lines: EFO-21 (Kd1 = 1.5 x 10(-9) M; binding capacity (Bmax1) = 4.9 fmol/10(6) cells) and EFO-27 (Kd1 = 1.7 x 10(-9) M; Bmax1 = 3 fmol/10(6) cells). In addition, a second class of low affinity/high capacity binding sites (EFO-21: Kd2 = 7.5 x 10(-6) M; Bmax2 = 24 pmol/10(6) cells; EFO-27: Kd2 = 4.3 x 10(-6) M; Bmax2 = 14.5 pmol/10(6) cells) was demonstrated. Specific binding of [125I, D-Trp6]LHRH was displaced with nearly equal efficiency by unlabeled [D-Trp6]LHRH, the LHRH-antagonists SB-75 and Hoe-013, and by native LHRH but not by unrelated peptides such as oxytocin and somatostatin. In the presence of 10(-5) M agonist [D-Trp6]LHRH, the proliferation of both cell lines was significantly reduced to 77% of controls after 24 h and to approx. 60% after 6 days. Lower concentrations (10(-9) M) of the agonist, significantly decreased the proliferation to 87.5% for EFO-21 and 86% for EFO-27 after 6 days. These antiproliferative effects were enhanced by increasing doses of [D-Trp6]LHRH and were maximal at 10(-5) M (EFO-21: 65.5% of control, EFO-27: 68% of control). Similar dose-dependent antiproliferative effects were obtained in EFO-21 line with the LHRH-antagonists SB-75 and Hoe-013, while these analogues had no effects on the proliferation of EFO-27 cells. SB-75 partly antagonized the antiproliferative effect of [D-Trp6]LHRH in a dose dependent way in the EFO-27 line. These data suggest that LHRH analogues can directly inhibit the in vitro proliferation of human ovarian cancer cells. This effect might be mediated through the high affinity LHRH binding sites.

High-density miniaturized thermal shift assays as a general strategy for drug discovery.

More general and universally applicable drug discovery assay technologies are needed in order to keep pace with the recent advances in combinatorial chemistry and genomics-based target generation. Ligand-induced conformational stabilization of proteins is a well-understood phenomenon in which substrates, inhibitors, cofactors, and even other proteins provide enhanced stability to proteins on binding. This phenomenon is based on the energetic coupling of the ligand-binding and protein-melting reactions. In an attempt to harness these biophysical properties for drug discovery, fully automated instrumentation was designed and implemented to perform miniaturized fluorescence-based thermal shift assays in a microplate format for the high throughput screening of compound libraries. Validation of this process and instrumentation was achieved by investigating ligand binding to more than 100 protein targets. The general applicability of the thermal shift screening strategy was found to be an important advantage because it circumvents the need to design and retool new assays with each new therapeutic target. Moreover, the miniaturized thermal shift assay methodology does not require any prior knowledge of a therapeutic target's function, making it ideally suited for the quantitative high throughput drug screening and evaluation of targets derived from genomics.

Gene Replacement in Mycobacterium tuberculosis and Mycobacterium bovis BCG Using rpsL as a Dominant Negative Selectable Marker.

Much progress has been made in mycobacterial research in general and in mycobacterial genetics in particular during the past 10 yr. The complete genome sequences of two isolates of Mycobacterium tuberculosis, the widely distributed laboratory strain M. tuberculosis H37Rv (1) and a clinical isolate CDC 1551 ( http://www.tigr.org ), have recently been determined, thus providing a complete blueprint of the genetic make-up of this pathogen.

A prospective study of modified Ottawa ankle rules in a military population. Interobserver agreement between physical therapists and orthopaedic surgeons.

To determine the necessity of ankle and foot radiographs, we used modified Ottawa Ankle Rules to evaluate all cadets seen with an acute ankle or midfoot injury at the United States Military Academy. This scoring system determines the need for radiographs. Each patient was independently examined and the decision rules were applied by a physical therapist and an orthopaedic surgeon. Ankle and foot radiographs were obtained for all subjects. Sensitivity, specificity, and the positive predictive value were calculated in 153 patients. There were six clinically significant ankle fractures and three midfoot fractures, for a total incidence of 5.8%. For physical therapists, the sensitivity was 100%, the specificity for ankle injuries was 40%, and the specificity for foot injuries was 79%. For orthopaedic surgeons, the sensitivity was also 100%, the specificity for ankle injuries was 46%, and the specificity for foot injuries was 79%. Interobserver agreement between the orthopaedic surgeons and physical therapists regarding the overall decision to obtain radiographs was high, with a kappa coefficient value of 0.82 for ankle injuries and 0.88 for foot injuries. There were no false-negative results. Use of the modified Ottawa Ankle Rules would have reduced the necessity for ankle and foot radiographs by 46% and 79%, respectively.

Current measurement in applied behavior analysis.

The analysis of behavior began with a form of data, rate of responding, which allowed for efficient study and for the description of the basic principles of behavior. Especially important were the facts that rate of responding was a direct reflection of fundamental properties of behavior, and that rate of responding was measured continuously within an experimental session. As behavior analysts moved from purely experimental to applied settings, discontinuous, time-based methods of measurement evolved, which neither directly reflect fundamental properties of behavior nor continuously record behavior within an experimental session. This paper offers a critical discussion of current measurement practices, and discusses factors possibly related to the use of discontinuous, time-based observing/recording procedures. A theoretical basis for observing/recording procedures is presented which emphasizes continuous measurement of response dimensions directly related to fundamental properties of behavior.

Analysis of Campylobacter jejuni isolates of various sources for loci associated with Guillain-Barré Syndrome.

Campylobacter jejuni is a major cause of the Guillain-Barré syndrome (GBS) and related diseases. These autoimmune diseases are caused by antibodies cross-reacting with the peripheral (GBS) and central neural tissue (Miller Fisher syndrome - MFS, Bicker-staff's brainstem encephalitis - BBE), leading to acute polyneuropathy. Recently, specific gene loci in C. jejuni have been distinguished which are associated with the onset of GBS, despite a molecular or phenotypic clustering. In this study, we used PCR to analyse C. jejuni isolates of different origin (i.e. bovine, poultry, human) for these genes. A total of 196 isolates were tested for cst-II and neuA. Of these, 101 isolates harboured the cst-II locus and 102 the neuA locus. Eighty-six isolates (44%) hold both genes. The frequency of cst-II in different sources of isolates of bovine, poultry and human isolates did not vary significantly (52, 50 and 52%, respectively). In contrast, the neuA locus was less often found in poultry isolates. Two human strains - from a family outbreak of campylobacteriosis (in 1989 in Austria) in which one person developed MFS - harboured both genes. Thus, although only one in more than 3000 patients with Campylobacter-associated enteritis develop GBS, about half of Campylobacter jejuni strains found in different environments are possibly able to cause GBS. These strains almost equally distributed in bovine, poultry and human isolates. Our results suggest that isolates associated with GBS are not selected by environmental or host-specific factors. Accordingly, this study indicates that host factors such as humoral and cellular immunity are possibly responsible for the development of these autoimmune diseases.

Foodborne gastroenteritis outbreak in an Austrian healthcare facility caused by asymptomatic, norovirus-excreting kitchen staff.

An outbreak of norovirus GGII.4 2006b affected an Austrian 600-bed healthcare facility from 15 to 27 March 2009. A total of 204 patients, residents and staff fitted the outbreak case definition; 17 (8.3%) were laboratory-confirmed. Foodborne origin was suspected in the 114 patient and resident cases with onset 15-18 March. A case-cohort study was performed to test the hypothesis that consumption of dishes offered on 14, 15 and 16 March (risk days) was associated with increased risk of infection. Data on food exposure of 62% (317/510) of the patient and resident cohort were available for a simultaneous retrospective cohort study. The case-cohort analysis revealed that consumption of sliced cold sausage offered on 15 March [odds ratio (OR): 3.98; 95% confidence interval (CI): 1.18-14.1], a meat dish with salad (adjusted OR: 2.2; 95% CI: 1.19-4.08) and a rolled spinach pancake (adjusted OR: 2.17; 95% CI: 1.27-3.71) on 16 March were independent risk factors. It is likely that one of the five asymptomatic excretors among the kitchen staff on duty on the risk days was the source of food contamination. The case-cohort study design was found to be a valid alternative to the retrospective cohort study design for the investigation of a suspected foodborne outbreak in a large cohort.

Rapidly destructive Staphylococcus epidermidis endocarditis.

A 29-year-old man with rapidly destructive Staphylococcus epidermidis endocarditis after mitral valve reconstruction is presented. Resistance to rifampin and teicoplanin occurred during antibiotic treatment resulting in clinical failure and valve destruction. Subsequently, the patient was successfully treated, by combining valve replacement with antibiotic therapy including quinupristin/dalfopristin, levofloxacin, and vancomycin. In conclusion, S. epidermidis can cause rapid valve destruction with large vegetations, and combination of surgery and antibiotic therapy may be necessary.

[A critical consideration of the combined radiochemotherapy of head and neck tumors]. / Kritische Betrachtung der kombinierten Radiochemotherapie von Kopf-Hals-Tumoren.

PURPOSE: Advanced tumors of the head and neck are often treated with combined radiochemotherapy. The chemotherapeutic agents used should be active in this tumor type and its adverse effects should not overlap with those of the radiation treatment. METHODS: Published studies were reviewed and discussed on the basis of these principles. RESULTS: Initially, single agents such as 5-fluorouracil (5-FU) and methotrexate were used in combination with radiotherapy. These combinations caused an improved local tumor control but also an increased mucosal reaction. For mitomycin C it has been shown in a randomized trial that local tumor control was improved without concomitant increased normal tissue toxicity. Also cisplatin and carboplatin were studied in combination with radiotherapy. Unfortunately, there are no results of randomized studies available but these agents do not seem to increase mucosal toxicity. The standard chemotherapy of squamous cell carcinomas of the head and neck is cisplatin and 5-FU. Many studies have been conducted with this chemotherapy in combination with radiotherapy. To this day it has not been shown that the results of an effective radiation treatment or an effective chemotherapy can be improved by these experiments. The explanation for that is that either the chemotherapy or the radiotherapy cannot be given at full dose because the regimen would become too toxic. CONCLUSION: 5-FU containing polychemotherapy regimens should not be combined with radiation any more because it is known that 5-FU increases the mucosal reaction. Agents that could be studied in the future either alone or in combination with cisplatin or carboplatin are etoposide and taxol.

High-level expression of sperm whale myoglobin in Escherichia coli.

Sperm whale myoglobin was expressed in Escherichia coli from a totally synthetic gene inserted in the expression vector pUC19. The gene was constructed as 23 overlapping oligonucleotides encoding both strands of the DNA. Gene synthesis provides several advantages over traditional eukaryotic gene-cloning techniques, allowing the incorporation of an efficient ribosome binding site, appropriate initiation and termination sequences, restriction enzyme sites for convenient subcloning and future mutagenesis, and frequently used codons for highly expressed E. coli genes. The sperm whale myoglobin expressed from the synthetic gene constituted approximately 10% of the total soluble protein as holo-protein, indicating that iron-protoporphyrin IX biosynthesis and prosthetic-group incorporation are not limiting in the high-level expression of this heme protein in E. coli. We credit the use of frequently used E. coli codons for the observed high-level expression. The sperm whale myoglobin produced is stable, easily purified to homogeneity, and indistinguishable from commercially available sperm whale myoglobin by optical and magnetic spectroscopic methods.