The Wiggle Index: An Open Source Bioassay to Assess Sub-Lethal Insecticide Response in Drosophila melanogaster.
Toxicological assays measuring mortality are routinely used to describe insecticide response, but sub-lethal exposures to insecticides can select for resistance and yield additional biological information describing the ways in which an insecticide impacts the insect. Here we present the Wiggle Index (WI), a high-throughput method to quantify insecticide response by measuring the reduction in motility during sub-lethal exposures in larvae of the vinegar fly Drosophila melanogaster. A susceptible wild type strain was exposed to the insecticides chlorantraniliprole, imidacloprid, spinosad, and ivermectin. Each insecticide reduced larval motility, but response times and profiles differed among insecticides. Two sets of target site mutants previously identified in mortality studies on the basis of imidacloprid or spinosad resistance phenotypes were tested. In each case the resistant mutant responded significantly less than the control. The WI was also able to detect a spinosad response in the absence of the primary spinosad target site. This response was not detected in mortality assays suggesting that spinosad, like many other insecticides, may have secondary targets affecting behaviour. The ability of the WI to detect changes in insecticide metabolism was confirmed by overexpressing the imidacloprid metabolizing Cyp6g1 gene in digestive tissues or the central nervous system. The data presented here validate the WI as an inexpensive, generic, sub-lethal assay that can complement information gained from mortality assays, extending our understanding of the genetic basis of insecticide response in D. melanogaster.


An integrative approach for modeling and simulation of heterocyst pattern formation in cyanobacteria filaments.
Heterocyst differentiation in cyanobacteria filaments is one of the simplest examples of cellular differentiation and pattern formation in multicellular organisms. Despite of the many experimental studies addressing the evolution and sustainment of heterocyst patterns and the knowledge of the genetic circuit underlying the behavior of single cyanobacterium under nitrogen deprivation, there is still a theoretical gap connecting these two macroscopic and microscopic processes. As an attempt to shed light on this issue, here we explore heterocyst differentiation under the paradigm of systems biology. This framework allows us to formulate the essential dynamical ingredients of the genetic circuit of a single cyanobacterium into a set of differential equations describing the time evolution of the concentrations of the relevant molecular products. As a result, we are able to study the behavior of a single cyanobacterium under different external conditions, emulating nitrogen deprivation, and simulate the dynamics of cyanobacteria filaments by coupling their respective genetic circuits via molecular diffusion. These two ingredients allow us to understand the principles by which heterocyst patterns can be generated and sustained. In particular, our results point out that, by including both diffusion and noisy external conditions in the computational model, it is possible to reproduce the main features of the formation and sustainment of heterocyst patterns in cyanobacteria filaments as observed experimentally. Finally, we discuss the validity and possible improvements of the model.


A practical guide for the study of human and murine sebaceous glands in situ.
The skin of most mammals is characterised by the presence of sebaceous glands (SGs), whose predominant constituent cell population is sebocytes, that is, lipid-producing epithelial cells, which develop from the hair follicle. Besides holocrine sebum production (which contributes 90% of skin surface lipids), multiple additional SG functions have emerged. These range from antimicrobial peptide production and immunomodulation, via lipid and hormone synthesis/metabolism, to the provision of an epithelial progenitor cell reservoir. Therefore, in addition to its involvement in common skin diseases (e.g. acne vulgaris), the unfolding diversity of SG functions, both in skin health and disease, has raised interest in this integral component of the pilosebaceous unit. This practical guide provides an introduction to SG biology and to relevant SG histochemical and immunohistochemical techniques, with emphasis placed on in situ evaluation methods that can be easily employed. We propose a range of simple, established markers, which are particularly instructive when addressing specific SG research questions in the two most commonly investigated species in SG research, humans and mice. To facilitate the development of reproducible analysis techniques for the in situ evaluation of SGs, this methods review concludes by suggesting quantitative (immuno-)histomorphometric methods for standardised SG evaluation.


Influence of surface treatments and resin cement selection on bonding to zirconia.
This study aimed to evaluate the surface changes caused in zirconia by different surface treatments and the influence of the surface treatment and cement selection on bonding to zirconia under aging. Sintered zirconia specimens were divided into five groups (n = 31) based on the surface treatment, namely, control, air abrasion, silica coating, laser and air abrasion + laser. After surface treatment, surface roughness and microscope analyses were performed on one specimen of each group. Composite cylinders were then bonded to conditioned ceramics using RelyX U100 (RXU), Clearfil Esthetic Cement (CEC) and Panavia F (PF) (n = 10). After 24 h, the bonded specimens were subjected to thermal cycling (6,000 times), and then, a shear bond strength test was conducted. The roughness values were analysed using Kruskal-Wallis and Mann-Whitney U tests, and the bond strengths were analysed by two-way analysis of variance and Duncan's test. The relationship between the roughness and the bond strength was determined by Spearman's correlation analysis. Specimens subjected to surface treatments were rougher than the control specimen (p < 0.000). However, there were no significant differences between the air abrasion and air abrasion + laser groups and the silica coating and laser groups. Specimens treated with laser showed lower bond strengths irrespective of the resin cement used. CEC and/or PF showed higher bond strengths than RXU for each surface treatment group. No significant relationship was observed between the roughness and the bond strength. The results of this study showed that all the surface treatments, except for laser irradiation, were suitable for treating zirconia ceramics. Cement selection was found to be more important than surface treatment, and phosphate monomer-containing cements were suitable for cementing zirconia.


ERBB3 is a marker of a ganglioneuroblastoma/ganglioneuroma-like expression profile in neuroblastic tumours.
Neuroblastoma (NB) tumours are commonly divided into three cytogenetic subgroups. However, by unsupervised principal components analysis of gene expression profiles we recently identified four distinct subgroups, r1-r4. In the current study we characterized these different subgroups in more detail, with a specific focus on the fourth divergent tumour subgroup (r4). Expression microarray data from four international studies corresponding to 148 neuroblastic tumour cases were subject to division into four expression subgroups using a previously described 6-gene signature. Differentially expressed genes between groups were identified using Significance Analysis of Microarray (SAM). Next, gene expression network modelling was performed to map signalling pathways and cellular processes representing each subgroup. Findings were validated at the protein level by immunohistochemistry and immunoblot analyses. We identified several significantly up-regulated genes in the r4 subgroup of which the tyrosine kinase receptor ERBB3 was most prominent (fold change: 132-240). By gene set enrichment analysis (GSEA) the constructed gene network of ERBB3 (n = 38 network partners) was significantly enriched in the r4 subgroup in all four independent data sets. ERBB3 was also positively correlated to the ErbB family members EGFR and ERBB2 in all data sets, and a concurrent overexpression was seen in the r4 subgroup. Further studies of histopathology categories using a fifth data set of 110 neuroblastic tumours, showed a striking similarity between the expression profile of r4 to ganglioneuroblastoma (GNB) and ganglioneuroma (GN) tumours. In contrast, the NB histopathological subtype was dominated by mitotic regulating genes, characterizing unfavourable NB subgroups in particular. The high ErbB3 expression in GN tumour types was verified at the protein level, and showed mainly expression in the mature ganglion cells. Conclusively, this study demonstrates the importance of performing unsupervised clustering and subtype discovery of data sets prior to analyses to avoid a mixture of tumour subtypes, which may otherwise give distorted results and lead to incorrect conclusions. The current study identifies ERBB3 as a clear-cut marker of a GNB/GN-like expression profile, and we suggest a 7-gene expression signature (including ERBB3) as a complement to histopathology analysis of neuroblastic tumours. Further studies of ErbB3 and other ErbB family members and their role in neuroblastic differentiation and pathogenesis are warranted.


Modelling zoonotic diseases in humans: comparison of methods for hantavirus in Sweden.
Because their distribution usually depends on the presence of more than one species, modelling zoonotic diseases in humans differs from modelling individual species distribution even though the data are similar in nature. Three approaches can be used to model spatial distributions recorded by points: based on presence/absence, presence/available or presence data. Here, we compared one or two of several existing methods for each of these approaches. Human cases of hantavirus infection reported by place of infection between 1991 and 1998 in Sweden were used as a case study. Puumala virus (PUUV), the most common hantavirus in Europe, circulates among bank voles (Myodes glareolus). In northern Sweden, it causes nephropathia epidemica (NE) in humans, a mild form of hemorrhagic fever with renal syndrome.Logistic binomial regression and boosted regression trees were used to model presence and absence data. Presence and available sites (where the disease may occur) were modelled using cross-validated logistic regression. Finally, the ecological niche model MaxEnt, based on presence-only data, was used.In our study, logistic regression had the best predictive power, followed by boosted regression trees, MaxEnt and cross-validated logistic regression. It is also the most statistically reliable but requires absence data. The cross-validated method partly avoids the issue of absence data but requires fastidious calculations. MaxEnt accounts for non-linear responses but the estimators can be complex. The advantages and disadvantages of each method are reviewed.


Fast-food consumption, diet quality, and neighborhood exposure to fast food: the multi-ethnic study of atherosclerosis.
The authors examined associations among fast-food consumption, diet, and neighborhood fast-food exposure by using 2000-2002 Multi-Ethnic Study of Atherosclerosis data. US participants (n = 5,633; aged 45-84 years) reported usual fast-food consumption (never, <1 time/week, or > or =1 times/week) and consumption near home (yes/no). Healthy diet was defined as scoring in the top quintile of the Alternate Healthy Eating Index or bottom quintile of a Western-type dietary pattern. Neighborhood fast-food exposure was measured by densities of fast-food outlets, participant report, and informant report. Separate logistic regression models were used to examine associations of fast-food consumption and diet; fast-food exposure and consumption near home; and fast-food exposure and diet adjusted for site, age, sex, race/ethnicity, education, and income. Those never eating fast food had a 2-3-times higher odds of having a healthy diet versus those eating fast food > or =1 times/week, depending on the dietary measure. For every standard deviation increase in fast-food exposure, the odds of consuming fast food near home increased 11%-61% and the odds of a healthy diet decreased 3%-17%, depending on the model. Results show that fast-food consumption and neighborhood fast-food exposure are associated with poorer diet. Interventions that reduce exposure to fast food and/or promote individual behavior change may be helpful.


3D finite element analysis of electrostatic deflection and shielding of commercial and FIB-modified cantilevers for electric and Kelvin force microscopy: II. Rectangular shaped cantilevers with asymmetric pyramidal tips.
This paper deals with an application of 3D finite element analysis to the electrostatic interaction between (i) a commercial rectangular shaped cantilever (with an integrated anisotropic pyramidal tip) and a conductive sample, when a voltage difference is applied between them, and (ii) a focused ion beam (FIB) modified cantilever in order to realize a probe with reduced parasitic electrostatic force. The 3D modelling of their electrostatic deflection was realized by using multiphysics finite element analysis software and applied to the real geometry of the cantilevers and probes as used in conventional electric and Kelvin force microscopy to evaluate the contribution of the various part of a cantilever to the total force, and derive practical criteria to optimize the probe performances. We report also on the simulation of electrostatic shielding of nanometric features, in order to quantitatively evaluate an alternative way of reducing the systematic error caused by the cantilever-to-sample capacitive coupling. Finally, a quantitative comparison between the performances of rectangular and triangular cantilevers (part I of this work) is reported.


Dental extraction for patients presenting at oral surgery student clinic.
The aim of this study was to examine the reasons for dental extraction and to determine the pattern of tooth loss in patients seeking care at the oral surgery teaching clinics in the Faculty of Dentistry of the University of Jordan, Amman, Jordan, over a 3-year period. Data pertaining to the dental extractions of 2435 patients were analyzed. The results showed that 63.8% of the teeth included in this study were extracted because of dental caries, 22.9% because of periodontal disease, and 11.0% for prosthetic reasons. Pericoronitis, orthodontic treatment, trauma, and eruption problems accounted for 2.4% of the reported extractions. The upper premolars were the teeth most commonly extracted, and the lower first and second molars were the teeth most commonly extracted because of dental caries. The logistic regression test revealed that extraction because of dental caries occurred mostly in the group aged 21 to 30 years (P < .001). Periodontal disease was not likely the cause of extraction in patients younger than 40 years. Mandibular incisors were the teeth least likely extracted because of dental caries (P < .001), but they were the teeth most commonly extracted because of periodontal disease (P < .001). Extraction for orthodontic reasons mostly involved the premolars (P < .05) and occurred in patients 20 years of age or younger (P < .001). Men were less likely to lose teeth because of caries and periodontal disease (P < .05 and P < .001, respectively) and were more likely to lose teeth for prosthetic reasons and trauma. The information gained from this study is useful to shift oral health planning toward emphasizing the importance of maintaining natural dentition and preventing dental disease.


Twelve tips for ethical approval for research in health professions education.
A growing number of faculty are engaging in research in health professions education. Suggestions continue to be made in the literature for a clear and less onerous pathway for the ethical review of this work. We aim to provide advice about the ethics application process for those conducting research in health professions education. We used critical reflection of our experiences as research ethics board (REB) members, applying for, reviewing and consulting about the ethics application process in both UK and Canadian health contexts in addition to evidence and advice that is available in the literature to inform the tips provided. Twelve tips are offered to help faculty understand and navigate through the ethics application process. Health professionals have an important role to play in advancing the field of health professions education, and despite issues identified with current review pathways, REB review is in place to ensure that this work is undertaken safely and ethically. We believe the tips offered in this article will help faculty identify, and devise plans to address, some ethical issues that are common in health professions education research.


Deficits in prospective memory following damage to the medial subdivision of the mediodorsal thalamic nucleus.
Identifying the neurocognitive mechanisms that lead individuals remembering to execute an intention at the right moment (prospective memory, PM) and how such mechanisms are influenced by the features of that intention is a fundamental theoretical challenge. In particular, the functional contribution of subcortical regions to PM is still unknown. This study was aimed at investigating the role of the medial subdivision of the mediodorsal thalamic nucleus (mMDT) in PM, with particular focus on the processes that are mediated by the projections from/to this structure. We analysed the performance of a patient (OG) with a right-sided lesion involving the mMDT in a series of PM tasks that varied for focality (i.e., overlapping of processes for the PM and ongoing tasks) and emotional valence of the stimuli, comparing the patient's performance with that of a control group. We found that the mMDT damage led to deficits in PM that were modulated by focality and emotional valence. OG indeed showed: a greater cost in the ongoing performance when a non-focal PM task was added; a slowing down in retrieving the intentions, in particular when these were associated with focal PM cues; an abnormal performance in the task with positive PM cues. Our findings provide evidence of a contribution of mMDT to PM and suggest a modulation of prefrontal-dependent strategic monitoring and a possible interaction with the limbic structures in the integration of emotion and PM processes. They also give support to the still controversial idea that connections with the perirhinal cortex mediate familiarity-based recognition.


Early oncological and functional outcomes following radical treatment of high-risk prostate cancer in men older than 70 years: A prospective longitudinal study.
Assess early oncological and functional outcomes following radical treatment of men with high-risk prostate cancer and aged more than 70 years. A total of 335 men with high-risk prostate cancer (prostate-specific antigen ≥20ng/ml or biopsy Gleason score 8 to 10 or≥cT2c) received radical treatment between 2007 and 2014. Men were identified from comprehensive clinical databases hosted at a tertiary cancer center in the UK. The data included basic demographics, and follow-up on functional and oncological outcomes using validated patient-reported outcome questionnaires. Univariate and multivariate analyses were used. In all, 117 patients received radical radiotherapy (RT) alone, 167 patients received neoadjuvant hormone therapy and RT, and 54 patients underwent radical prostatectomy with extended lymph node dissection. Mean age was 72.8, standard deviation (SD) = 2.1, mean follow-up of 40.9 months, SD = 25.5 months. Patients who underwent laparoscopic prostatectomy = 24 (44.4%) had positive surgical margins, and mean lymph nodes dissected were 18.7, SD = 6.7. Further, 5 men experienced postoperative complications in the form of pseudoaneurism of internal iliac branch, leg ischemia, high CO2 retention, and 2 men experienced sepsis. Incidence of biochemical recurrence was significantly lower at 16.7% in the surgery group, compared with RT 51.3% and RT and hormone therapy 30.5%, and Kaplan-Meier analysis P<0.001 over 3 years of follow-up Radical surgery with extended lymph nodes dissection appears to have good short-term oncological and functional outcomes compared with RT with or without hormones in high-risk men older than 70 years of age. Based on these findings, treatment decisions and surgical therapy should be considered on individual basis in older men with high-risk disease.


Probing amyloid protein aggregation with optical superresolution methods: from the test tube to models of disease.
The misfolding and self-assembly of intrinsically disordered proteins into insoluble amyloid structures are central to many neurodegenerative diseases such as Alzheimer's and Parkinson's diseases. Optical imaging of this self-assembly process in vitro and in cells is revolutionizing our understanding of the molecular mechanisms behind these devastating conditions. In contrast to conventional biophysical methods, optical imaging and, in particular, optical superresolution imaging, permits the dynamic investigation of the molecular self-assembly process in vitro and in cells, at molecular-level resolution. In this article, current state-of-the-art imaging methods are reviewed and discussed in the context of research into neurodegeneration.


Linking Gait Dynamics to Mechanical Cost of Legged Locomotion.
For millenia, legged locomotion has been of central importance to humans for hunting, agriculture, transportation, sport, and warfare. Today, the same principal considerations of locomotor performance and economy apply to legged systems designed to serve, assist, or be worn by humans in urban and natural environments. Energy comes at a premium not only for animals, wherein suitably fast and economical gaits are selected through organic evolution, but also for legged robots that must carry sufficient energy in their batteries. Although a robot's energy is spent at many levels, from control systems to actuators, we suggest that the mechanical cost of transport is an integral energy expenditure for any legged system-and measuring this cost permits the most direct comparison between gaits of legged animals and robots. Although legged robots have matched or even improved upon total cost of transport of animals, this is typically achieved by choosing extremely slow speeds or by using regenerative mechanisms. Legged robots have not yet reached the low mechanical cost of transport achieved at speeds used by bipedal and quadrupedal animals. Here we consider approaches used to analyze gaits and discuss a framework, termed mechanical cost analysis, that can be used to evaluate the economy of legged systems. This method uses a point mass perspective to evaluate the entire stride as well as to identify individual events that accrue mechanical cost. The analysis of gait began at the turn of the last century with spatiotemporal analysis facilitated by the advent of cine film. These advances gave rise to the "gait diagram," which plots duty factors and phase separations between footfalls. This approach was supplanted in the following decades by methods using force platforms to determine forces and motions of the center of mass (CoM)-and analytical models that characterize gait according to fluctuations in potential and kinetic energy. Mechanical cost analysis draws from these approaches and provides a unified framework that interprets the spatiotemporal sequencing of leg contacts within the context of CoM dynamics to determine mechanical cost in every instance of the stride. Diverse gaits can be evaluated and compared in biological and engineered systems using mechanical cost analysis.


Discovery of Time-Delayed Gene Regulatory Networks based on temporal gene expression profiling.
It is one of the ultimate goals for modern biological research to fully elucidate the intricate interplays and the regulations of the molecular determinants that propel and characterize the progression of versatile life phenomena, to name a few, cell cycling, developmental biology, aging, and the progressive and recurrent pathogenesis of complex diseases. The vast amount of large-scale and genome-wide time-resolved data is becoming increasing available, which provides the golden opportunity to unravel the challenging reverse-engineering problem of time-delayed gene regulatory networks. In particular, this methodological paper aims to reconstruct regulatory networks from temporal gene expression data by using delayed correlations between genes, i.e., pairwise overlaps of expression levels shifted in time relative each other. We have thus developed a novel model-free computational toolbox termed TdGRN (Time-delayed Gene Regulatory Network) to address the underlying regulations of genes that can span any unit(s) of time intervals. This bioinformatics toolbox has provided a unified approach to uncovering time trends of gene regulations through decision analysis of the newly designed time-delayed gene expression matrix. We have applied the proposed method to yeast cell cycling and human HeLa cell cycling and have discovered most of the underlying time-delayed regulations that are supported by multiple lines of experimental evidence and that are remarkably consistent with the current knowledge on phase characteristics for the cell cyclings. We established a usable and powerful model-free approach to dissecting high-order dynamic trends of gene-gene interactions. We have carefully validated the proposed algorithm by applying it to two publicly available cell cycling datasets. In addition to uncovering the time trends of gene regulations for cell cycling, this unified approach can also be used to study the complex gene regulations related to the development, aging and progressive pathogenesis of a complex disease where potential dependences between different experiment units might occurs.


Structural basis for selective vascular endothelial growth factor-A (VEGF-A) binding to neuropilin-1.
Neuropilin-1 (Nrp1) is an essential receptor for angiogenesis that binds to VEGF-A. Nrp1 binds directly to VEGF-A with high affinity, but the nature of their selective binding has remained unclear. Nrp1 was initially reported to bind to the exon 7-encoded region of VEGF-A and function as an isoform-specific receptor for VEGF-A(164/165). Recent data have implicated exon 8-encoded residues, which are found in all proangiogenic VEGF-A isoforms, in Nrp binding. We have determined the crystal structure of the exon 7/8-encoded VEGF-A heparin binding domain in complex with the Nrp1-b1 domain. This structure clearly demonstrates that residues from both exons 7 and 8 physically contribute to Nrp1 binding. Using an in vitro binding assay, we have determined the relative contributions of exon 7- and 8-encoded residues. We demonstrate that the exon 8-encoded C-terminal arginine is essential for the interaction of VEGF-A with Nrp1 and mediates high affinity Nrp binding. Exon 7-encoded electronegative residues make additional interactions with the L1 loop of Nrp1. Although otherwise conserved, the primary sequences of Nrp1 and Nrp2 differ significantly in this region. We further show that VEGF-A(164) binds 50-fold more strongly to Nrp1 than Nrp2. Direct repulsion between the electronegative exon 7-encoded residues of the heparin binding domain and the electronegative L1 loop found only in Nrp2 is found to significantly contribute to the observed selectivity. The results reveal the basis for the potent and selective binding of VEGF-A(164) to Nrp1.


RankExplorer: Visualization of Ranking Changes in Large Time Series Data.
For many applications involving time series data, people are often interested in the changes of item values over time as well as their ranking changes. For example, people search many words via search engines like Google and Bing every day. Analysts are interested in both the absolute searching number for each word as well as their relative rankings. Both sets of statistics may change over time. For very large time series data with thousands of items, how to visually present ranking changes is an interesting challenge. In this paper, we propose RankExplorer, a novel visualization method based on ThemeRiver to reveal the ranking changes. Our method consists of four major components: 1) a segmentation method which partitions a large set of time series curves into a manageable number of ranking categories; 2) an extended ThemeRiver view with embedded color bars and changing glyphs to show the evolution of aggregation values related to each ranking category over time as well as the content changes in each ranking category; 3) a trend curve to show the degree of ranking changes over time; 4) rich user interactions to support interactive exploration of ranking changes. We have applied our method to some real time series data and the case studies demonstrate that our method can reveal the underlying patterns related to ranking changes which might otherwise be obscured in traditional visualizations.


Differential interactions of cytochalasins with P-glycoprotein.
Cytochalasins are a family of structurally related natural product cytotoxins that selectively depolymerize microfilaments. In this study, the interaction between several cytochalasins and the drug transporter P-glycoprotein was investigated. Dihydrocytochalasin B and cytochalasin E consistently sensitized P-glycoprotein-overexpressing human breast carcinoma cells (MCF-7/ADR) to daunomycin, vinblastine, and actinomycin D without affecting the cytotoxicity of cisplatin. These compounds did not affect the sensitivities of the parental MCF-7 cells to anticancer drugs, indicating that their effects are due to P-glycoprotein inhibition. Effects of dihydrocytochalasin B and cytochalasin E were observed at concentrations as low as 2.5 and 5 microM, respectively. In contrast, cytochalasins A, B, C, D, H, and J did not sensitize MCF-7/ADR cells to any of the drugs. The accumulation of [3H]-vinblastine by MCF-7/ADR cells and by drug-resistant human ovarian carcinoma cells (SKVLB1) was increased to the greatest extent by verapamil, followed by dihydrocytochalasin B > cytochalasin E > cytochalasin B, whereas cytochalasins A, C, D, H, and J did not alter intracellular accumulation of the drug. Similarly to verapamil, dihydrocytochalasin B significantly stimulated the ATPase activity of P-glycoprotein, while other cytochalasins were ineffective. These results demonstrate that very closely related compounds can differentially interact with P-glycoprotein. For example, the only difference between cytochalasin B and dihydrocytochalasin B is the saturation of a carbon-carbon double bond in dihydrocytochalasin B. These structural differences may provide important insight into chemical determinants for drug interaction with P-glycoprotein.


Coronary Artery Calcification, Statin Use and Long-Term Risk of Atherosclerotic Cardiovascular Disease Events (from the Multi-Ethnic Study of Atherosclerosis).
The prognostic utility of coronary artery calcium (CAC) for individuals taking statins is unclear. We hypothesized that CAC remains associated with atherosclerotic cardiovascular disease (ASCVD) events in individuals using statins at baseline or among those started on statin at follow-up. The Multi-Ethnic Study of Atherosclerosis is a prospective cohort study of 6,814 participants who were enrolled between 2000 and 2002 and were free of clinical ASCVD at baseline. Four follow-up visits were conducted in 2002 to 2004, 2004 to 2006, 2005 to 2007, and 2010 to 2012. CAC was assessed at baseline and follow-up using either an electron-beam CT scanner or a multidetector CT system. Statin use at baseline and follow up was self-reported. Among 6,811 participants with complete information on statin use, mean age was 62 (SD = 10) years, 53% were women, 38% white, 12% Chinese-American, 28% African American, and 22% Hispanic. In multivariable analyses, CAC >0 was associated with a significantly higher risk of ASCVD events regardless of baseline or incident statin use. For example, hazard ratios (95% confidence interval) for the association between CAC >0 and ASCVD were 2.46 (1.41, 4.28) for baseline statin users, 2.08 (1.68, 2.57) for baseline-statin nonusers, and 2.21 (1.56, 3.15) for those started on a statin at follow-up. In conclusion, current statin use does not weaken the prognostic utility of CAC. CAC is associated with incident ASCVD regardless of baseline or incident statin use.


VoiceYourView: anytime, anyplace, anywhere user participation.
Citizens are increasingly called upon to comment on issues that directly concern them. However, such consultations may be tokenistic [1] as they occur with limited respondents, or may be at a time, or in a format which is inconvenient to the user. To encourage wider participation, the VoiceYourView project (vYv) has developed a system allowing people to make comments in a manner, time and place convenient to them. A real world trial of the prototype system was conducted at Coventry University campus to explore issues related to the system's usability and usage, as a means of enabling campus users to comment on their environment. Members of the university population were invited to comment on the university estate using one of five technologies (e-mail, online form, iPhone app, SMS message, or electronic kiosk). Although the immediate application area in this case was the design of public spaces, the approach can be transferred to other domains and thus provide a new way of gathering user information. Submitted comments were automatically analysed in terms of theme, sentiment, location and actionability and displayed online in a 2D visualisation. It is argued that that online data collection (crowd sourcing and skimming social networks) may provide a rich source of information for future ergonomists.


Maternal obesity and breastfeeding intention, initiation, intensity and duration: a systematic review.
This systematic review investigates the relationship between maternal obesity and breastfeeding intention, initiation, intensity, duration and milk supply. A comprehensive search was performed through three major databases, including Medline, Cochrane Library and Cumulative Index For Nursing and Allied Health Literature, and by screening reference lists of the relevant publications. Selection criteria were: report of original research, studies on low-risk obese mothers and the comparison with normal weight mothers which met at least two of the following primary outcomes: breastfeeding intention; initiation; intensity; duration and/or milk supply. Furthermore, the included reports had to contain a clear definition of pre-pregnant obesity, use compensation mechanisms for potential confounding factors, have a prospective cohort design and had to have been published between 1997 and 2011 and in English, French or Dutch. Effects of obesity on breastfeeding intention, initiation, intensity, duration and milk supply were analysed, tabulated and summarised in this review. Studies have found that obese women are less likely to intend to breastfeed and that maternal obesity seems to be associated with a decreased initiation of breastfeeding, a shortened duration of breastfeeding, a less adequate milk supply and delayed onset of lactogenesis II, compared with their normal weight counterparts. This systematic review indicates therefore that maternal obesity is an adverse determinant for breastfeeding success.


Ameliorative effects of Moringa oleifera leaf extract on levofloxacin-induced hepatic toxicity in rats.
Fluoroquinolones are some of the most common antibiotics used by clinicians all over the world. Levofloxacin, a fluoroquinolone, is used therapeutically in numerous countries; however, it can cause an increase in liver function tests and liver dysfunction. The current study was designed to determine the effect of Levofloxacin (40 mg/kg body weight (b.wt.) daily for 2 weeks) on rat liver function and oxidative stress markers as well as to evaluate the potential hepatoprotective effects of Moringa oleifera leaf extract as a known antioxidant herb. M. oleifera leaf extract was found to improve the hepatic dysfunction induced by Levofloxacin by recovering liver enzymatic activities (alanine aminotransferase [ALT], aspartate aminotransferase [AST] and gamma-glutamyl transferase [GGT]) to normal levels. The extract also reversed the antioxidant imbalance as measured by catalase and superoxide dismutase activities as well as by reduced glutathione and malondialdehyde levels. Moreover, M. oleifera leaf extract induced anti-inflammation by improving the production of interleukin (IL)-10. Additionally, its presence attenuated the downregulation of IL-1 induced by Levofloxacin alone from hepatic tissue. It can be concluded that M. oleifera extract can help reduce the side effects caused by Levofloxacin administration.


Extracellular vesicles of ETV2 transfected fibroblasts stimulate endothelial cells and improve neovascularization in a murine model of hindlimb ischemia.
Ischemia are common conditions related to lack of blood supply to tissues. Depending on the ischemic sites, ischemia can cause different diseases, such as hindlimb ischemia, heart infarction and stroke. This study aims to evaluate how extracellular vesicles (EVs) derived from ETV2 transfected fibroblasts affect endothelial cell proliferation and neovascularization in a murine model of hindlimb ischemia. Human fibroblasts were isolated and cultured under standard conditions and expanded to the 3th passage before use in experiments. Human fibroblasts were transduced with a viral vector containing the ETV2 gene. Transduced cells were selected by puromycin treatment. These cells were further cultured for collection of EVs, which were isolated from culture supernatant. Following co-culture with endothelial cells, EVs were evaluated for their effect on endothelial cell proliferation and were directly injected into ischemic tissues of a murine model of hindlimb ischemia. The results showed that EVs could induce endothelial cell proliferation in vitro and improved neovascularization in a murine model of hindlimb ischemia. Our results suggest that EVs derived from ETV2-transfected fibroblasts can be promising non-cellular products for the regeneration of blood vessels.


A review of near patient testing in primary care.
The aim was to identify publications relating to near patient testing (NPT), the use of alternative delivery systems between laboratory and general practice, including electronic data interchange (EDI), and computerised diagnostic decision support (CDDS), in the primary care setting to answer the following questions. What is the availability of NPT for primary care? What evidence is available to support the clinical effectiveness of NPT? What evidence is available on the accuracy and reliability of NPT within primary care? What evidence is available on the cost-effectiveness of different NPTs? How may CDDS improve the effectiveness of NPT? What evidence is available that compares NPT and existing laboratory services? What evidence is available on the cost-effectiveness of EDI or alternative delivery systems? Eight databases were searched, and the bibliographies from relevant publications checked for completeness. Unpublished work and publications not included in the databases were obtained by personal contact with collaborators, and from a postal survey sent to heads of academic departments of general practice and clinical chemistry and to researchers active or interested in the field worldwide. Questionnaires were also sent to 150 commercial organisations. Publications that met agreed definitions and reported original data were included in the systematic review. Of the 1057 publications identified, 102 (92 related to NPT, eight to CDDS, and two to EDI) were passed to the reviewers for appraisal of validity. The limited amount of published research relating to any particular NPT prohibited meta-analysis. Scoring systems to assess the validity of evaluations were also difficult to apply. A wide variety of NPT systems have been developed. In general, the quality of the methods reported in the literature was poor. The issue of patient convenience and acceptability has not been adequately addressed. No evaluations of alternative delivery systems met the review criteria. No studies have evaluated the telephone or fax machine as a means of reporting results. For EDI, the majority of papers were descriptive. EDI and alternative delivery systems are not a replacement for NPT when the provision of an immediate result might have an impact on the quality of care. EDI may have clinical and cost advantages over traditional means of communication, but this has not been evaluated. The advisory role of the laboratory can be supported by CDDS. The use of CDDS and NPT has not, however, been fully evaluated. Few economic analyses have been conducted, and most were simple cost analyses. There are insufficient data for conclusions to be drawn on the cost-effectiveness of NPT in primary care. FURTHER SYSTEMATIC REVIEWS: Subject-specific systematic reviews are required that include laboratory and secondary care studies, and consider the potential for altering current management and patient acceptability. Priority topics include: biochemistry profiles on desktop analysers; cholesterol testing; urinalysis for the diagnosis of urinary tract infection; anticoagulation control; NPTs for the identification of acute infection. (ABSTRACT TRUNCATED)


A drone-based survey for large, basking freshwater turtle species.
Conservation concerns are increasing for numerous freshwater turtle species, including Pseudemys gorzugi, which has led to a call for more research. However, traditional sampling methodologies are often time consuming, labor intensive, and invasive, restricting the amount of data that can be collected. Biases of traditional sampling methods can further impair the quality of the data collected, and these shortfalls may discourage their use. The use of unmanned aerial vehicles (UAVs, drones) for conducting wildlife surveys has recently demonstrated the potential to bridge gaps in data collection by offering a less labor intensive, minimally invasive, and more efficient process. Photographs and video can be obtained by camera attachments during a drone flight and analyzed to determine population counts, abundance, and other types of data. In this study we developed a detailed protocol to survey for large, freshwater turtle species in an arid, riverine landscape. This protocol was implemented with a DJI Matrice 600 Pro drone and a SONY ILCE α6000 digital camera to determine P. gorzugi and sympatric turtle species occurrence across 42 sites in southwestern Texas, USA. The use of a large drone and high-resolution camera resulted in high identification percentages, demonstrating the potential of drones to survey for large, freshwater turtle species. Numerous advantages to drone-based surveys were identified as well as some challenges, which were addressed with additional refinement of the protocol. Our data highlight the utility of drones for conducting freshwater turtle surveys and provide a guideline to those considering implementing drone-mounted high-resolution cameras as a survey tool.


[Anisocoria: a suspicious sign of anisotropy and/or amblyopia].
A four-year retrospective study among children who attend a Primary Care Pediatric Service has been made, in search of a connection between the infant anisocoria and the anisometropia and/or amblyopia detected on the screening of visual acuity at the age of three-four years. Four out of five children with anisocoria (that is, the 80%) later suffered anisometropia and/or amblyopia, and, reciprocally, four out of eight children with anisometropia and/or amblyopia (50%) have previously presented anisocoria. The existence of a clear association is shown between both pathologies, and consequently, children with anisocoria (after putting aside other possible reasons, oculars or neurologicals) should be promptly examined on refraction and visual acuity in order to diagnose anisometropia and to prevent development of amblyopia.


Is guilt 'likely' or 'not certain'? Contrast with previous probabilities determines choice of verbal terms.
This research focuses on what determines speakers' choice of positive and negative probability phrases (e.g., "a chance" vs. "not certain") in a legal context. We argue that choice of phrase to describe an event's probability of occurrence can be determined by the contrast between its current p value and an earlier p value, and not by that current value alone. Three experiments were conducted describing scenarios where profilers communicated a suspect's probability of guilt to the police. In the first study, a probability estimate is revised upwards or downwards. In the second one, the probability estimate of a speaker is higher or lower than that given by a previous speaker. In both cases, participants expected upward trends to lead to positive phrases, whereas downward trends were associated with negative phrases. In a third study, participants had to select probability phrases to characterize two different suspects. No contrast effects were found. We conclude that verbal probability directionality has primarily an argumentative function, where positive phrases are selected when probabilities are contrasted with smaller p values, and negative when contrasted with higher p values.


Clinical evaluation of M43: a novel cancer-associated mucin epitope.
A monoclonal antibody to colon carcinoma mucin was found to react with a colon carcinoma-associated carbohydrate epitope. This antibody was used to develop a quantitative solid phase immunoassay, M43. We prospectively and retrospectively evaluated the assay in patients with and without gastrointestinal carcinoma and compared the sensitivity and specificity with that of carcinoembryonic antigen (CEA) and CA 19-9. One hundred ninety-two patients (181 with no evidence of malignancy) referred for upper or lower gastrointestinal endoscopy were prospectively studied. Sera from 172 patients with histologically confirmed gastrointestinal adenocarcinoma were retrospectively studied. Optimal discrimination cutoffs for M43 (5 units/ml), CEA (5 ng/ml), and CA 19-9 (30 units/ml) were determined by receiver operating characteristic curves analysis. M43 was positive in 112 of 151 patients with colorectal carcinoma (sensitivity 74%) and was negative in 167 of 181 patients without carcinoma (specificity 92%). Sensitivity and specificity were 77% and 93% for CEA and 60% and 83% for CA 19-9. Sixty-four % of 73 patients with colorectal carcinoma limited to the bowel wall had a positive M43 with a mean value of 178 units/ml. Eighty-one % of 27 patients with nonhepatic metastasis had a positive M43 with a mean value of 223 units/ml. Eighty-four % of 51 patients with hepatic metastasis had a positive M43 assay with a mean value of 2532 units/ml. Sensitivity in these three groups was 67%, 82%, and 82%, respectively, for CEA and 43%, 68%, and 79%, respectively, for CA 19-9. Of 38 carcinoma patients with a negative CEA, 45% had a positive M43. No correlation between the levels of M43 and CEA in patients with colorectal carcinoma was found. We conclude that M43 is positive in most patients with colorectal carcinoma, even in early stages. As a diagnostic test, its sensitivity and specificity are equivalent to those of CEA. However, the M43 assay is measuring a tumor antigen which is fundamentally different from CEA and which is present in a high percentage of CEA-negative patients.


First Report of the Root Knot Nematode, Meloidogyne inornata, on Common Bean in Paraná State, Brazil.
Common bean (Phaseolus vulgaris F.) is one of the most important crops in Paraná State, which is responsible for almost 10% of the Brazilian production (4). Root knot nematodes, Meloidogyne spp., are common parasites of this crop worldwide, but damage caused by Meloidogyne inornata has not been reported. During a survey of nematode species present on common bean fields in Paraná State, Brazil, galled root samples of cultivars Tuiuiú and Eldorado were submitted, in June 2012, in the Nematology Laboratory from IAPAR, collected in the municipalities of Araucária (25°35'34″S, 49°24'36″W) and Santana do Itararé (23°45'18″S, 49°37'44″W). Plants did not exhibit any above-ground symptoms. The specimens were identified through perineal patterns and esterase phenotypes of 20 adult females extracted from dissected roots (2,3). The population densities observed in the samples were 140 and 700 J2 and eggs per gram of roots, respectively, for both samples. Characteristics were consistent with those described for M. inornata. For example, perineal patterns of M. inornata showed a high dorsal arch, with smooth to wavy striae, similar to those of M. incognita; but no punctate markings between anus and tail terminus were observed. However, from the esterase electrophoresis we obtained the I3 (Rm = 0.83, 1.15, and 1.32) phenotype, typical of M. inornata, a species-specific phenotype used to differentiate this species from M. incognita (1). Moreover, the excretory pore of adult females was located 32.1 (± 5.4) μm from the anterior end, consistent with the M. inornata description (25 to 53 μm) (1). To the best of our knowledge, this is the first report of M. inornata parasitizing common bean roots. This finding has great importance for Brazilian agriculture, since this nematode may damage common bean plants and become an additional problem for this crop. Additional work is necessary in order to elucidate the losses caused by M. inornata on common bean. References: (1) R. M. D. G. Carneiro et al. Nematology 10:123, 2008. (2) P. R. Esbenshade and A. C. Triantaphyllou J. Nematol. 22:10, 1990. (3) K. M. Hartman and J. N. Sasser. Page 115 in: An Advanced Treatise on Meloidogyne, Volume II Methodology. K. R. Barker et al., eds. Raleigh: North Carolina State University Graphics, 1985. (4) MAPA. Feijão, Ministério da Agricultura, Brasil. Retrieved from http://www.agricultura.gov.br/vegetal/culturas/feijao September 05, 2012.


Differential In Vitro and In Vivo Toxicities of Antimicrobial Peptide Prodrugs for Potential Use in Cystic Fibrosis.
There has been considerable interest in the use of antimicrobial peptides (AMPs) as antimicrobial agents for the treatment of many conditions, including cystic fibrosis (CF). The challenging conditions of the CF patient lung require robust AMPs that are active in an environment of high proteolytic activity but that also have low cytotoxicity and immunogenicity. Previously, we developed prodrugs of AMPs that limited the cytotoxic effects of AMP treatment by rendering the antimicrobial activity dependent on the host enzyme neutrophil elastase (NE). However, cytotoxicity remained an issue. Here, we describe the further optimization of the AMP prodrug (pro-AMP) model for CF to produce pro-WMR, a peptide with greatly reduced cytotoxicity (50% inhibitory concentration against CFBE41o- cells, >300 μM) compared to that of the previous group of pro-AMPs. The bactericidal activity of pro-WMR was increased in NE-rich bronchoalveolar lavage (BAL) fluid from CF patients (range, 8.4% ± 6.9% alone to 91.5% ± 5.8% with BAL fluid; P = 0.0004), an activity differential greater than that of previous pro-AMPs. In a murine model of lung delivery, the pro-AMP modification reduced host toxicity, with pro-WMR being less toxic than the active peptide. Previously, host toxicity issues have hampered the clinical application of AMPs. However, the development of application-specific AMPs with modifications that minimize toxicity similar to those described here can significantly advance their potential use in patients. The combination of this prodrug strategy with a highly active AMP has the potential to produce new therapeutics for the challenging conditions of the CF patient lung.


Late entry to antenatal care in New South Wales, Australia.
This study aimed to assess the prevalence of women who entered antenatal care (ANC) late and to identify factors related to the late entry to ANC in New South Wales (NSW) in 2004. The NSW Midwives Data Collection contained data of 85,034 women who gave birth in 2004. Data were downloaded using SAS and transferred to STATA 8.0. Entering ANC after 12 weeks of gestation was classified as late. The Andersen Health Seeking Behaviour Model was used for selection and analyses of related factors. Regression and hierarchical analyses were used to identify significant factors and their relative contributions to the variation of pregnancy duration at entry to ANC. 41% of women commenced ANC after 12 weeks of gestation. Inequality existed between groups of women with predisposing characteristics and enabling resources contributed more to the variation in pregnancy duration at entry to ANC than needs. The groups of women with highest risk were teenagers, migrants from developing countries, women living in Western Sydney, Aboriginal and Torres Strait Islanders, women with three or more previous pregnancies and heavy smokers. The high risk groups with largest number of women were migrants from developing countries and women living in Western Sydney. A large number of women in NSW entered ANC late in their pregnancies. Efforts to increase early entry to ANC should be targeted on identified high risk groups of women.


Involvement of Mitochondrial Dysfunction, Endoplasmic Reticulum Stress, and the PI3K/AKT/mTOR Pathway in Nobiletin-Induced Apoptosis of Human Bladder Cancer Cells.
Nobiletin (NOB) is a polymethoxylated flavonoid isolated from citrus fruit peel that has been shown to possess anti-tumor, antithrombotic, antifungal, anti-inflammatory and anti-atherosclerotic activities. The main purpose of this study was to explore the potential of using NOB to induce apoptosis in human bladder cancer cells and study the underlying mechanism. Using an MTT assay, agarose gel electrophoresis, a wound-healing assay, flow cytometry, and western blot analysis, this study investigated the signaling pathways involved in NOB-induced apoptosis in BFTC human bladder cancer cells. Our results showed that NOB at concentrations of 60, 80, and 100 μM inhibited cell growth by 42%, 62%, and 80%, respectively. Cells treated with 60 μM NOB demonstrated increased DNA fragmentation, and flow cytometry analysis confirmed that the treatment caused late apoptotic cell death. Western blot analysis showed that mitochondrial dysfunction occurred in NOB-treated BFTC cells, leading to cytochrome C release into cytosol, activation of pro-apoptotic proteins (caspase-3, caspase-9, Bad, and Bax), and inhibition of anti-apoptotic proteins (Mcl-1, Bcl-xl, and Bcl-2). NOB-induced apoptosis was also mediated by regulating endoplasmic reticulum stress via the PERK/elF2α/ATF4/CHOP pathway, and downregulating the PI3K/AKT/mTOR pathway. Our results suggested that the cytotoxic and apoptotic effects of NOB on bladder cancer cells are associated with endoplasmic reticulum stress and mitochondrial dysfunction.


Composite tissue allotransplantation and dysregulation in tissue repair and regeneration: a role for mesenchymal stem cells.
Vascularized composite tissue allotransplantation is a rapidly evolving area that has brought technological advances to the forefront of plastic surgery, hand surgery, and transplant biology. Composite tissue allografts (CTAs) may have profound functional, esthetic, and psychological benefits, but carry with them the risks of life-long immunosuppression and the inadequate abilities to monitor and prevent rejection. Allografts may suffer from additional insults further weakening their overall benefits. Changes in local blood flow, lack of fully restored neurologic function, infection, inflammation with subsequent dysregulated regenerative activity, and paucity of appropriate growth factors may all be involved in reducing the potential of CTAs and therefore serve as new therapeutic targets to improve outcomes. Strategies involving minimized immunosuppression and pro-regenerative therapy may provide a greater path to optimizing long-term CTA function. One such strategy may include mesenchymal stem cells (MSCs), which can provide unique anti-inflammatory and pro-regenerative effects. Insights gained from new studies with MSCs on composite allografts, advances in tissue regeneration reported in other MSC-based clinical studies, as well as consideration of newly described capacities of MSCs, may provide new regenerative based strategies for the care of CTAs.


Cardiometabolic Abnormalities Among Normal-Weight Persons From Five Racial/Ethnic Groups in the United States: A Cross-sectional Analysis of Two Cohort Studies.
The relationship between body weight and cardiometabolic disease may vary substantially by race/ethnicity. To determine the prevalence and correlates of the phenotype of metabolic abnormality but normal weight (MAN) for 5 racial/ethnic groups. Cross-sectional analysis. 2 community-based cohorts. 2622 white, 803 Chinese American, 1893 African American, and 1496 Hispanic persons from MESA (Multi-Ethnic Study of Atherosclerosis) and 803 South Asian participants in the MASALA (Mediators of Atherosclerosis in South Asians Living in America) study. Prevalence of 2 or more cardiometabolic abnormalities (high fasting glucose, low high-density lipoprotein cholesterol, and high triglyceride levels and hypertension) among normal-weight participants was estimated. Correlates of MAN were assessed by using log-binomial models. Among normal-weight participants (n = 846 whites, 323 Chinese Americans, 334 African Americans, 252 Hispanics, and 195 South Asians), the prevalence of MAN was 21.0% (95% CI, 18.4% to 23.9%) in whites, 32.2% (CI, 27.3% to 37.4%) in Chinese Americans, 31.1% (CI, 26.3% to 36.3%) in African Americans, 38.5% (CI, 32.6% to 44.6%) in Hispanics, and 43.6% (CI, 36.8% to 50.6%) in South Asians. Adjustment for demographic, behavioral, and ectopic body fat measures did not explain racial/ethnic differences. After adjustment for age, sex, and race/ethnicity-body mass index (BMI) interaction, for the equivalent MAN prevalence at a BMI of 25.0 kg/m2 in whites, the corresponding BMI values were 22.9 kg/m2 (CI, 19.5 to 26.3 kg/m2) in African Americans, 21.5 kg/m2 (CI, 18.5 to 24.5 kg/m2) in Hispanics, 20.9 kg/m2 (CI, 19.7 to 22.1 kg/m2) in Chinese Americans, and 19.6 kg/m2 (CI, 17.2 to 22.0 kg/m2) in South Asians. Cross-sectional study design and lack of harmonized dietary data between studies. Compared with whites, all racial/ethnic minority groups had a statistically significantly higher prevalence of MAN, which was not explained by demographic, behavioral, or ectopic fat measures. Using a BMI criterion for overweight to screen for cardiometabolic risk may result in a large proportion of racial/ethnic minority groups being overlooked. National Institutes of Health.


A side by side comparison of cytology and biomarkers for bladder cancer detection.
The identification of accurate bladder tumor markers/tests could improve diagnosis, recurrence monitoring and treatment in patients with bladder cancer. In this study we compared the efficacy of the hyaluronic acid (HA)-hyaluronidase (HAase), BTA-Stat (Bard/Bion Diagnostics, Redmond, Washington), Hemastix (Bayer Corp., Elkhart, Indiana) (hematuria detection) and UBC-Rapid (IDL Biotech, Borlänger, Sweden) tests, and cytology to detect bladder cancer. The HA-HAase test measures urinary HA and HAase levels, BTA-Stat detects complement factor-H and H related protein in urine, the Hemastix hemoglobin dipstick detects hematuria and UBC-Rapid detects cytokeratin 8 and 18 fragments in urine. A total of 138 urine specimens from 115 patients were collected at University Hospital Hamburg-Eppendorf, including 59 with active bladder cancer and 79 with a history of bladder cancer (73) or with benign genitourinary conditions (6). Specimens were assayed by the HA-HAase test, BTA-Stat, Hemastix (hemoglobin dipstick) and UBC-Rapid. Cystoscopy and histological findings were used to make the clinical diagnosis. Cytology results were available on 92 patients. In a side by side comparison the HA-HAase test, cytology, BTA-Stat, Hemastix and UBC-Rapid had 88.1%, 70.6%, 52.5%, 50.8% and 35.6% sensitivity, and 81%, 81%, 76.7%, 78.2% and 75% specificity, respectively. The accuracy, and negative and positive predictive values of the HA-HAase test were the highest (84.1%, 90.1% and 77.6%), followed by cytology (77.2%, 82.5% and 68.6%), Hemastix (66.4%, 67.8% and 63.8%), BTA-Stat (66.2%, 67.8% and 63.3%) and UBC-Rapid (57.8%, 60% and 52.5%), respectively. : The HA-HAase test is superior to cytology, BTA-Stat, Hemastix and UBC-Rapid for detecting bladder cancer recurrence. A side-by-side comparison of tumor markers should help identify a marker for monitoring bladder cancer recurrence.


Leaf and Stem Spot Caused by Ramularia sphaeroidea on Purple and Lana Woollypod Vetch (Vicia spp.) Cover Crops in California.
Vetches (Vicia spp.) are planted alone or in combination with other plants as cover crops in vegetable production areas of California. December 2001 through February 2003, purple (V. benghalensis) and lana woollypod (V. villosa subsp. varia) vetches in the Salinas Valley (Monterey County) developed a foliar disease. Symptoms were small (≤5 mm in diameter), circular to oblong, purple brown-to-red brown spots that were visible from the adaxial and abaxial leaf sides, and occurred lower in the plant canopy. White sporulation was visible in the spot centers. Stems were infected and had elongated, irregularly shaped, brown lesions that were <5 mm long and had white sporulation. When fungal masses or tissues from lesions were placed on acidified potato dextrose agar (LA-PDA), a fungus was consistently recovered. On LA-PDA, the isolates produced slow-growing (30 mm colony diameter in 45 days), irregularly raised, light pink and white colonies that produced dark exudates. The undersurfaces of cultures were gray black. The growth on lesions consisted of fascicles of conidiophores that were hyaline, smooth, flexuous, distinctly geniculate, and measured 20 to 120 × 2.5 to 6 µm. Conidia were hyaline, subglobose, smooth, aseptate, measured 9 to 15 × 8 to 13 µm, and formed singly. The internal transcribed spacer rDNA sequence of a representative strain (CBS 112891) was determined using standard protocols (GenBank Accession No. AY352584). A nucleotide BLAST search revealed a 94 to 97% similarity to other species of Ramularia (GenBank Accession Nos. AF222848, AF173310, AJ417496, AF362060, and AF297235). On the basis of these morphological and molecular data, the fungus was identified as Ramularia sphaeroidea Sacc. (= Ovularia sphaeroidea (Sacc.) Sacc.) (1). Pathogenicity of six isolates grown on LA-PDA was confirmed by spraying conidial suspensions (1.0 × 105 conidia per ml) onto direct-seeded, 8-week-old, potted purple and lana woollypod vetch (12 plants each). Plants were kept in a dew chamber for 48 h and maintained in a greenhouse (23 to 25°C). After 7 to 10 days, all plants developed the characteristic leaf spots and stem lesions, and R. sphaeroidea was reisolated from such symptoms. Plants treated with only water did not develop symptoms. However, because disease incidence on test plants was low, inoculum was also prepared in water amended with 1.0 ppm of Tween 20. Four pots each of purple and lana woollypod vetch were sprayed with amended or nonamended inocula, and plants were handled as described. After 10 days, plants inoculated with Tween 20 amended inoculum had significantly higher disease incidence and severity (purple = 83% of leaflets infected with a mean of 3.4 spots per leaflet; lana = 83% infected with a mean of 3.2 spots) than did plants inoculated with water-only conidial suspensions (purple = 27% and a mean of 0.4 spots; lana = 38% and a mean of 0.6 spots). Finally, two other vetches used in the Salinas Valley were inoculated with the two suspensions. After 2 weeks, common (V. sativa) and languedoc (V. sativa 'Languedoc') vetch showed no symptoms, and control plants of purple and lana vetch developed disease. All inoculation tests were repeated, and results were similar. To our knowledge, this is the first report of R. sphaeroidea as a pathogen of purple and lana woollypod vetches in California. Reference: (1) U. Braun. A Monograph of Cercosporella, Ramularia, and Allied Genera (Phytopathogenic Hyphomycetes) Vol. 2. IHW-Verlag, Eching, Germany, 1998.


Changes in blood pressure, vascular reactivity and inflammatory biomarkers following consumption of heated corn oil.
Consumption of corn oil for cooking purpose is gaining popularity. The present study examined the effect of heated corn oil on blood pressure and its possible mechanism in experimental rats. Thirty male Sprague-Dawley rats were randomly divided into 5 groups and were fed with the following diets, Group I was fed with basal diet only; whereas group II,III,IV and V were fed with basal diet fortified with 15% (w/w) either fresh, once-heated, five-times-heated or ten-times-heated corn oil, respectively for 16 weeks. Body weight, blood pressure were measured at baseline and weekly interval for 16 weeks. Inflammatory biomarkers which included soluble intracellular adhesion molecules (sICAM), soluble vascular adhesion molecules (sVCAM) and C reactive protein (CRP), were measured at baseline and the end of 16 weeks. The rats were sacrificed and thoracic aorta was taken for measurement of vascular reactivity. There was significant increase in the blood pressure in the groups fed with heated once, five-times (5HCO) and ten-times-heated corn oil (10-HCO) compared to the control. The increase in the blood pressure was associated with an increase in CRP, sICAM and sVCAM, reduction in vasodilatation response to acetylcholine and greater vasoconstriction response to phenylephrine. The results suggest that repeatedly heated corn oil causes elevation in blood pressure, vascular inflammation which impairs vascular reactivity thereby predisposing to hypertension. There is a need to educate people not to consume corn oil in a heated state.


A MALDI-Mass Spectrometry Imaging method applicable to different formalin-fixed paraffin-embedded human tissues.
Recent advancements in Matrix Assisted Laser Desorption/Ionisation (MALDI) Mass Spectrometry Imaging (MSI) technology have enabled the analysis of formalin-fixed paraffin-embedded (FFPE) tissue samples, unlocking a wealth of new proteomic information and facilitating the possibility of performing studies with higher statistical power as well as multi-centric collaborations within the field of proteomics research. However, current methods used to analyse these specimens are often time-consuming and they need to be modified when applied to human tissues of different origin. Here we present a reproducible and time-effective method that could address these aforementioned issues and widen the applicability of this technology to a number of challenging tissue types. Additionally, tissue molecular images show high spatial resolution and a strong correlation with the morphological features, enabling the identification of tissue morphology using statistically derived visualisation, without any prior knowledge.


Sirtuin Lipoamidase Activity Is Conserved in Bacteria as a Regulator of Metabolic Enzyme Complexes.
Lipoic acid is an essential metabolic cofactor added as a posttranslational modification on several multimeric enzyme complexes. These protein complexes, evolutionarily conserved from bacteria to humans, are core regulators of cellular metabolism. While the multistep enzymatic process of adding lipoyl modifications has been well characterized in Escherichia coli, the enzyme required for the removal of these lipoyl moieties (i.e., a lipoamidase or delipoylase) has not yet been identified. Here, we describe our discovery of sirtuins as lipoamidases in bacteria and establish their conserved substrates. Specifically, by using a series of knockout, overexpression, biochemical, in vitro, proteomic, and functional assays, we determined the substrates of sirtuin CobB in E. coli as components of the pyruvate dehydrogenase (PDH), α-ketoglutarate dehydrogenase (KDH), and glycine cleavage (GCV) complexes. In vitro assays provided direct evidence for this specific CobB activity and its NAD+ dependence, a signature of all sirtuins. By designing a targeted quantitative mass spectrometry method, we further measured sirtuin-dependent, site-specific lipoylation on these substrates. The biological significance of CobB-modulated lipoylation was next established by its inhibition of both PDH and KDH activities. By restricting the carbon sources available to E. coli, we demonstrated that CobB regulates PDH and KDH under several growth conditions. Additionally, we found that SrtN, the sirtuin homolog in Gram-positive Bacillus subtilis, can also act as a lipoamidase. By demonstrating the evolutionary conservation of lipoamidase activity across sirtuin homologs, along with the conservation of common substrates, this work emphasizes the significance of protein lipoylation in regulating central metabolic processes.IMPORTANCE Here, we demonstrate that sirtuin lipoamidase activity exists in both Gram-positive and Gram-negative bacteria and establishing its conservation from bacteria to humans. Specifically, we discovered that CobB and SrtN act as lipoamidases in E. coli and B. subtilis, respectively. Intriguingly, not only is this sirtuin enzymatic activity conserved, but also the lipoylated substrates and functions are conserved, as bacterial sirtuins negatively regulate the lipoylation levels and activities of PDH and KDH. Considering that PDH and KDH regulate two carbon entry points into the tricarboxylic acid cycle, our finding highlights lipoylation as a conserved molecular toggle that regulates central metabolic pathways. Indeed, our findings from tests in which we limited nutrient availability support this. Furthermore, this study illustrates how the integration of technologies from different disciplines provides avenues to uncover enzymatic activities at the core of cellular metabolism regulation.


Carrying asymmetric loads while walking on an uneven surface.
Individuals often carry asymmetric loads over challenging surfaces such as uneven or irregular terrain, which may require a higher demand for postural control than walking on an even surface. The purpose of this study was to assess postural stability in the medial-lateral (ML) direction while carrying unilateral versus bilateral loads when walking on even versus uneven surfaces. Nineteen healthy young adults walked on even and uneven surface treadmills under three load conditions: no load, 20% body weight (BW) bilateral load, and 20% BW unilateral load. A Pedar in-shoe pressure system (Novel, Munich, Germany) was used to evaluate center of pressure (COP)-based parameters. Carrying 20% BW bilateral or unilateral loads significantly increased double support ratio. In addition, carrying a 20% BW unilateral load significantly increased coefficient of variation (CV) of double support ratio, CV of ML COP excursion, and CV of ML COP velocity. Walking on an uneven surface significantly increased double support ratio, ML COP excursion, ML COP velocity, and CV of double support ratio. When carrying a 20% BW unilateral load, unloaded limb stance had significantly increased double support ratio and ML COP velocity, although it appears that the loaded limb may be used to make step-by-step adjustments as evidenced by the higher CV of ML COP velocity. Unilateral load carriage, walking on uneven surfaces, and unloaded leg stance are of particular concern when considering postural stability.


Diagnosis of Pneumocystis pneumonia: evaluation of four serologic biomarkers.
The diagnosis of Pneumocystis pneumonia (PCP) relies on microscopic visualization of Pneumocystis jirovecii organisms or DNA detection in pulmonary specimens. This study aimed to assess the usefulness of (1-3)-β-d-glucan (BG), Krebs von den Lungen-6 antigen (KL-6), lactate dehydrogenase (LDH) and S-adenosyl methionine (SAM) as serologic biomarkers in the diagnosis of PCP. Serum levels of BG, KL-6, LDH and SAM were investigated in 145 Portuguese patients, 50 patients from the Netherlands, 25 Spanish patients and 40 Portuguese blood donors. Data on clinical presentation, chest imaging and gasometry tests were available. PCP cases were confirmed by microscopy and PCR techniques. A cost-effectiveness analysis was performed. BG was found to be the most reliable serologic biomarker for PCP diagnosis, followed by KL-6, LDH and SAM. The BG/KL-6 combination test was the most accurate serologic approach for PCP diagnosis, with 94.3% sensitivity and 89.6% specificity. Although less sensitive/specific than the reference standard classic methods based on bronchoalveolar lavage followed by microscopic or molecular detection of P. jirovecii organisms, the BG/KL-6 test may provide a less onerous procedure for PCP diagnosis, as it uses a minimally invasive and inexpensive specimen (blood), which may be also a major benefit for the patient's care. The BG/KL-6 combination test should be interpreted within the clinical context, and it may be used as a preliminary screening test in patients with primary suspicion of PCP, or as an alternative diagnostic procedure in patients with respiratory failure or in children, avoiding the associated risk of complications by the use of bronchoscopy.


Effect of temperature on pH and electrolyte concentration in air-breathing ectotherms.
The aim of this study was to determine the effects of temperature upon pH, protein charge and acid-base-relevant ion exchange in air-breathing ectotherms. Plasma and skeletal muscles in cane toads (Bufo marinus) and bullfrogs (Rana catesbeiana) were examined at 30, 20 and 10 degrees C. In addition, skeletal muscle ion concentrations were examined in black racer snakes (Coluber constrictor) at 30 and 10 degrees C. Cooling the amphibians produced a reduction in most of the plasma ion concentrations (Na(+), K(+), Ca(2+), Cl(-), SO(4)(2)(-)) and in protein concentration because of increased hydration. Between 30 and 10 degrees C, total plasma osmolality fell by 14 % in the toads and by 5 % in the frogs. Plasma protein charge, calculated using the principle of electroneutrality, was unaffected by temperature, except possibly for the toads at 10 degrees C. The in vivo skeletal muscle capdelta pHi/ capdelta T ratio, where pHi is intracellular pH and T is temperature, between 30 and 20 degrees C averaged -0.014 degrees C(-)(1) in the toads and -0.019 degrees C(-)(1) in the frogs. Between 20 and 10 degrees C, there was no change in pHi in the toads and a -0.005 degrees C(-)(1) change in the frogs. The in vitro skeletal muscle capdelta pHi/ capdelta T averaged -0.011 degrees C(-)(1) in both toads and frogs. In all three species, skeletal muscle inulin space declined with cooling. Intracellular ion concentrations were calculated by subtracting extracellular fluid ion concentrations from whole-muscle ion concentrations. In general, temperature had a large effect upon intracellular ion concentrations (Na(+), K(+), Cl(-)) and intracellular CO(2) levels. The relevance of the changes in intracellular ion concentration to skeletal muscle acid-base status and protein charge and the possible mechanisms producing the adjustments in intracellular ion concentration are discussed. It is concluded that ion-exchange mechanisms make an important contribution to adjusting pH with changes in temperature.


Toxoplasmosis.
Toxoplasma gondii, an Apicomplexan, is a pathogic protozoan that can infect the central nervous system. Infection during pregnancy can result in a congenial infection with severe neurological sequelae. In immunocompromised individuals reactivation of latent neurological foci can result in encephalitis. Immunocompetent individuals infected with T. gondii are typically asymptomatic and maintain this infection for life. However, recent studies suggest that these asymptomatic infections may have effects on behavior and other physiological processes. Toxoplasma gondii infects approximately one-third of the world population, making it one of the most successful parasitic organisms. Cats and other felidae serve as the definite host producing oocysts, an environmentally resistant life cycle stage found in cat feces, which can transmit the infection when ingested orally. A wide variety of warm-blooded animals, including humans, can serve as the intermediate host in which tissue cysts (containing bradyzoites) develop. Transmission also occurs due to ingestion of the tissue cysts. There are three predominant clonal lineages, termed Types I, II and III, and an association with higher pathogenicity with the Type I strains in humans has emerged. This chapter presents a review of the biology of this infection including the life cycle, transmission, epidemiology, parasite strains, and the host immune response. The major clinical outcomes of congenital infection, chorioretinitis and encephalitis, and the possible association of infection of toxoplasmosis with neuropsychiatric disorders such as schizophrenia, are reviewed.


The community impact of the 2009 influenza pandemic in the WHO European region: a comparison with historical seasonal data from 28 countries.
The world has recently experienced the first influenza pandemic of the 21st century that lasted 14 months from June 2009 to August 2010. This study aimed to compare the timing, geographic spread and community impact during the winter wave of influenza pandemic A (H1N1) 2009 to historical influenza seasons in countries of the WHO European region. We assessed the timing of pandemic by comparing the median peak of influenza activity in countries of the region during the last seven influenza seasons. The peaks of influenza activity were selected by two independent researchers using predefined rules. The geographic spread was assessed by correlating the peak week of influenza activity in included countries against the longitude and latitude of the central point in each country. To assess the community impact of pandemic influenza, we constructed linear regression models to compare the total and age-specific influenza-like-illness (ILI) or acute respiratory infection (ARI) rates reported by the countries in the pandemic season to those observed in the previous six influenza seasons. We found that the influenza activity reached its peak during the pandemic, on average, 10.5 weeks (95% CI 6.4-14.2) earlier than during the previous 6 seasons in the Region, and there was a west to east spread of pandemic A(H1N1) influenza virus in the western part of the Region. A regression analysis showed that the total ILI or ARI rates were not higher than historical rates in 19 of the 28 countries. However, in countries with age-specific data, there were significantly higher consultation rates in the 0-4 and/or 5-14 age groups in 11 of the 20 countries. Using routine influenza surveillance data, we found that pandemic influenza had several differential features compared to historical seasons in the region. It arrived earlier, caused significantly higher number of outpatient consultations in children in most countries and followed west to east spread that was previously observed during some influenza seasons with dominant A (H3N2) ifluenza viruses. The results of this study help to understand the epidemiology of 2009 influenza pandemic and can be used for pandemic preparedness planning.


Annoyance caused by sounds of wheeled and tracked vehicles.
In a laboratory experiment, the relationship between the annoyance caused by sounds of tracked and wheeled vehicles was investigated. Sounds of various vehicles, ranging from a main battle tank (MBT) to a passenger car, were recorded at several distances. Subjects were presented with these sounds and were asked to judge the annoyance if they were exposed to them at home on a regular basis. Overall, sounds emanated from military tracked vehicles were judged to be less annoying than civil passenger cars, provided that the indoor A-weighted sound-exposure levels (ASELs) were the same. It should be noted that receiver distance and operating conditions were not specifically matched for military and civil vehicles but were chosen to represent typical ranges of values found in practice. For steady-speed conditions, the sounds were judged to be equally annoying if the (indoor) ASELs of the tracked armored personnel carrier (APC) and the MBT exceeded that of a passenger car by 5 and 10 dB, respectively. In specific driving conditions such as alternate accelerations and decelerations, however, these differences were smaller. For ASELs below 55 dB, passenger cars were found to be more annoying than heavier wheeled vehicles, such as a bus or a truck with a trailer. The data indicate that the difference between the high-frequency part and the low-frequency part of the spectrum might play a role in the annoyance. With respect to noise-zoning procedures it is highly interesting that the outdoor ASEL could predict the annoyance, as rated indoors, better than could any indoor noise measure. The 5- and 10-dB bonuses to the indoor level of the APC and the MBT, respectively, were reduced to 1 and 3 dB if the dose was measured outdoors.


Relating neural dynamics to neural coding.
We demonstrate that two key theoretical objects used widely in computational neuroscience, the phase-resetting curve (PRC) from dynamics and the spike triggered average (STA) from statistical analysis, are closely related when neurons fire in a nearly regular manner and the stimulus is sufficiently small. We prove that the STA due to injected noisy current is proportional to the derivative of the PRC. We compare these analytic results with numerical calculations for the Hodgkin-Huxley neuron and we apply the method to neurons in the olfactory bulb of mice. This observation allows us to relate the stimulus-response properties of a neuron to its dynamics, bridging the gap between dynamical and information theoretic approaches to understanding brain computations and facilitating the interpretation of changes in channels and other cellular properties as influencing the representation of stimuli.


Divergent evolution in reverse transcriptase (RT) of HIV-1 group O and M lineages: impact on structure, fitness, and sensitivity to nonnucleoside RT inhibitors.
Natural evolution in primate lentiviral reverse transcriptase (RT) appears to have been constrained by the necessity to maintain function within an asymmetric protein composed of two identical primary amino acid sequences (66 kDa), of which one is cleaved (51 kDa). In this study, a detailed phylogenetic analysis now segregates groups O and M into clusters based on a cysteine or tyrosine residue located at position 181 of RT and linked to other signature residues. Divergent evolution of two group O (C181 or Y181) and the main (Y181 only) HIV-1 lineages did not appreciably impact RT activity or function. Group O RT structural models, based on group M subtype B RT crystal structures, revealed that most evolutionarily linked amino acids appear on a surface-exposed region of one subunit while in a noncatalytic RT pocket of the other subunit. This pocket binds nonnucleoside RT inhibitors (NNRTI); therefore, NNRTI sensitivity was used to probe enzyme differences in these group O and M lineages. In contrast to observations showing acquired drug resistance associated with fitness loss, the C181Y mutation in the C181 group O lineage resulted in a loss of intrinsic NNRTI resistance and was accompanied by fitness loss. Other mutations linked to the NNRTI-resistant C181 lineage also resulted in altered NNRTI sensitivity and a net fitness cost. Based on RT asymmetry and conservation of the intricate reverse transcription process, millions of years of divergent primate lentivirus evolution may be constrained to discrete mutations that appear primarily in the nonfunctional, solvent-accessible NNRTI binding pocket.


The burden of bacterial antimicrobial resistance in the WHO European region in 2019: a cross-country systematic analysis.
Antimicrobial resistance (AMR) represents one of the most crucial threats to public health and modern health care. Previous studies have identified challenges with estimating the magnitude of the problem and its downstream effect on human health and mortality. To our knowledge, this study presents the most comprehensive set of regional and country-level estimates of AMR burden in the WHO European region to date. We estimated deaths and disability-adjusted life-years attributable to and associated with AMR for 23 bacterial pathogens and 88 pathogen-drug combinations for the WHO European region and its countries in 2019. Our methodological approach consisted of five broad components: the number of deaths in which infection had a role, the proportion of infectious deaths attributable to a given infectious syndrome, the proportion of infectious syndrome deaths attributable to a given pathogen, the percentage of a given pathogen resistant to an antimicrobial drug of interest, and the excess risk of mortality (or duration of an infection) associated with this resistance. These components were then used to estimate the disease burden by using two counterfactual scenarios: deaths attributable to AMR (considering an alternative scenario where infections with resistant pathogens are replaced with susceptible ones) and deaths associated with AMR (considering an alternative scenario where drug-resistant infections would not occur at all). Data were solicited from a wide array of international stakeholders; these included research hospitals, surveillance networks, and infection databases maintained by private laboratories and medical technology companies. We generated 95% uncertainty intervals (UIs) for final estimates as the 25th and 975th ordered values across 1000 posterior draws, and models were cross-validated for out-of-sample predictive validity. We estimated 541 000 deaths (95% UI 370 000-763 000) associated with bacterial AMR and 133 000 deaths (90 100-188 000) attributable to bacterial AMR in the whole WHO European region in 2019. The largest fatal burden of AMR in the region came from bloodstream infections, with 195 000 deaths (104 000-333 000) associated with resistance, followed by intra-abdominal infections (127 000 deaths [81 900-185 000]) and respiratory infections (120 000 deaths [94 500-154 000]). Seven leading pathogens were responsible for about 457 000 deaths associated with resistance in 53 countries of this region; these pathogens were, in descending order of mortality, Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa, Enterococcus faecium, Streptococcus pneumoniae, and Acinetobacter baumannii. Methicillin-resistant S aureus was shown to be the leading pathogen-drug combination in 27 countries for deaths attributable to AMR, while aminopenicillin-resistant E coli predominated in 47 countries for deaths associated with AMR. The high levels of resistance for several important bacterial pathogens and pathogen-drug combinations, together with the high mortality rates associated with these pathogens, show that AMR is a serious threat to public health in the WHO European region. Our regional and cross-country analyses open the door for strategies that can be tailored to leading pathogen-drug combinations and the available resources in a specific location. These results underscore that the most effective way to tackle AMR in this region will require targeted efforts and investments in conjunction with continuous outcome-based research endeavours. Bill & Melinda Gates Foundation, Wellcome Trust, and Department of Health and Social Care using UK aid funding managed by the Fleming Fund.


What Are the Factors Identifying Caregivers Who Need Help in Managing Medications for Palliative Care Patients at Home? A Population Survey.
Background: For most people, the last 12 months of life are spent living in the community, with the support of family and friends for a number of caregiving functions. Previous research has found that managing medicines is challenging for caregivers. Currently there is little information describing which caregivers may struggle with tasks associated with managing a loved one's medicines. Aim: The aim of this study was to identify factors that flag caregivers who are likely to experience problems when managing someone else's medications. Setting/Participants: The annual South Australian Health Omnibus Survey provides a face-to-face, cross-sectional, whole-of-population view of health care. Structured interviews, including questions covering palliative care and end-of-life care, were conducted with 14,625 residents in their own homes. Results: Of the 1068 respondents who had provided care for someone who died of a terminal illness in the last five years, 7.4% identified that additional support with medicine management would have been beneficial. In addition, three factors were predictive of the need for additional support in managing medicines: aged <65 years; lower household income; and living in a metropolitan region. Conclusion: The findings of this study provide insights to inform the development of palliative care service models to support informal caregivers in the management of medications for people with a life-limiting illness.


Xpert® MTB/RIF for national tuberculosis programmes in low-income countries: when, where and how?
Xpert ® MTB/RIF offers new and important possibilities for the diagnosis of sputum smear-negative tuberculosis (TB) and/or rifampicin (RMP) resistance, and many are encouraging rapid and widespread implementation. This simple test can be implemented almost everywhere, and it provides results within a few hours. In low-income countries (LICs), however, its cost, environmental limitations (stable and regular electricity, adequate room temperature) and difficulties involved in supply and maintenance are major obstacles. While it may be suitable for major reference hospitals, operational research is needed to evaluate the test and its additional yield above high-quality smear microscopy and clinical algorithms before its use at the peripheral level. In the meantime, direct microscopy should remain the initial diagnostic test for TB suspects. In most LICs, the prevalence of RMP resistance among new TB patients is very low; an Xpert MTB/RIF result indicating RMP resistance will thus always need confirmation by another test. In a population at high risk of RMP resistance (> 15%), however, the positive predictive value for RMP resistance by Xpert MTB/RIF is high, and identification of RMP resistance is an excellent proxy for multidrug-resistant TB (MDR-TB). The assay should be widely used for this purpose if, and only if, excellent MDR-TB management is available, both for ethical reasons and to reduce the risk of extensively drug-resistant TB.


Effects of physical activity on life expectancy with cardiovascular disease.
Physical inactivity is a modifiable risk factor for cardiovascular disease. However, little is known about the effects of physical activity on life expectancy with and without cardiovascular disease. Our objective was to calculate the consequences of different physical activity levels after age 50 years on total life expectancy and life expectancy with and without cardiovascular disease. We constructed multistate life tables using data from the Framingham Heart Study to calculate the effects of 3 levels of physical activity (low, moderate, and high) among populations older than 50 years. For the life table calculations, we used hazard ratios for 3 transitions (healthy to death, healthy to disease, and disease to death) by levels of physical activity and adjusted for age, sex, smoking, any comorbidity (cancer, left ventricular hypertrophy, arthritis, diabetes, ankle edema, or pulmonary disease), and examination at start of follow-up period. Moderate and high physical activity levels led to 1.3 and 3.7 years more in total life expectancy and 1.1 and 3.2 more years lived without cardiovascular disease, respectively, for men aged 50 years or older compared with those who maintained a low physical activity level. For women the differences were 1.5 and 3.5 years in total life expectancy and 1.3 and 3.3 more years lived free of cardiovascular disease, respectively. Avoiding a sedentary lifestyle during adulthood not only prevents cardiovascular disease independently of other risk factors but also substantially expands the total life expectancy and the cardiovascular disease-free life expectancy for men and women. This effect is already seen at moderate levels of physical activity, and the gains in cardiovascular disease-free life expectancy are twice as large at higher activity levels.


The impact of parents' medication beliefs on asthma management.
Previous studies suggest a relationship between parental beliefs about asthma medications and medication adherence. It is not clear how parents' positive and negative feelings about medications interact to influence medication adherence. The objectives of this study were to describe parents' perceived need for and concerns about their child's asthma medications and to assess the weighted impact of these positive and negative beliefs on parent-reported adherence. We conducted a cross-sectional survey of parents of children with asthma in southeast Michigan; response rate was 71%. Children with reported use of a preventive asthma medication were included (n = 622). We used a validated Beliefs About Medications Questionnaire (2 subscales: necessity and concern) to assess parents' positive and negative attitudes about their child's medications. To measure how parents weigh these beliefs, we also calculated a necessity-concern differential score (difference between necessity and concern subscales). We used a 4-item parent-report scale to measure medication adherence. The majority of children were nonminority. Overall, 72% of parents felt that their child's asthma medications were necessary, and 30% had strong concerns about the medications. For 77% of parents, necessity scores were higher than concern scores, and for 17%, concern exceeded necessity. Nonminority parents were more likely to have necessity scores exceed concern scores compared with minority parents (79% vs 68%). Mean adherence scores increased as the necessity-concern differential increased. In a multivariate mixed-model regression, a greater necessity-concern differential score and being nonminority predicted better adherence. These findings confirm a relationship between medication beliefs and adherence among parents of children with asthma. A better understanding of parents' medication beliefs and their impact on adherence may help clinicians counsel effectively to promote adherence.


Bursaries, writing grants and fellowships: a strategy to develop research capacity in primary health care.
General practitioners and other primary health care professionals are often the first point of contact for patients requiring health care. Identifying, understanding and linking current evidence to best practice can be challenging and requires at least a basic understanding of research principles and methodologies. However, not all primary health care professionals are trained in research or have research experience. With the aim of enhancing research skills and developing a research culture in primary health care, University Departments of General Practice and Rural Health have been supported since 2000 by the Australian Government funded 'Primary Health Care Research Evaluation and Development (PHCRED) Strategy'. A small grant funding scheme to support primary health care practitioners was implemented through the PHCRED program at Flinders University in South Australia between 2002 and 2005. The scheme incorporated academic mentors and three types of funding support: bursaries, writing grants and research fellowships. This article describes outcomes of the funding scheme and contributes to the debate surrounding the effectiveness of funding schemes as a means of building research capacity. Funding recipients who had completed their research were invited to participate in a semi-structured 40-minute telephone interview. Feedback was sought on acquisition of research skills, publication outcomes, development of research capacity, confidence and interest in research, and perception of research. Data were also collected on demographics, research topics, and time needed to complete planned activities. The funding scheme supported 24 bursaries, 11 writing grants, and three research fellows. Nearly half (47%) of all grant recipients were allied health professionals, followed by general practitioners (21%). The majority (70%) were novice and early career researchers. Eighty-nine percent of the grant recipients were interviewed. Capacity, confidence, and level of research skills in ten core areas were generally considered to have improved as a result of the award. More than half (53%) had presented their research and 32% had published or submitted an article in a peer-reviewed journal. A small grant and mentoring scheme through a University Department can effectively enhance research skills, confidence, output, and interest in research of primary health care practitioners.


Complete Mapping of Mutations to the SARS-CoV-2 Spike Receptor-Binding Domain that Escape Antibody Recognition.
Antibodies targeting the SARS-CoV-2 spike receptor-binding domain (RBD) are being developed as therapeutics and are a major contributor to neutralizing antibody responses elicited by infection. Here, we describe a deep mutational scanning method to map how all amino-acid mutations in the RBD affect antibody binding and apply this method to 10 human monoclonal antibodies. The escape mutations cluster on several surfaces of the RBD that broadly correspond to structurally defined antibody epitopes. However, even antibodies targeting the same surface often have distinct escape mutations. The complete escape maps predict which mutations are selected during viral growth in the presence of single antibodies. They further enable the design of escape-resistant antibody cocktails-including cocktails of antibodies that compete for binding to the same RBD surface but have different escape mutations. Therefore, complete escape-mutation maps enable rational design of antibody therapeutics and assessment of the antigenic consequences of viral evolution.