Projected expansion of Trichodesmium's geographical distribution and increase in growth potential in response to climate change.
Estimates of marine N2 fixation range from 52 to 73 Tg N/year, of which we calculate up to 84% is from Trichodesmium based on previous measurements of nifH gene abundance and our new model of Trichodesmium growth. Here, we assess the likely effects of four major climate change-related abiotic factors on the spatiotemporal distribution and growth potential of Trichodesmium for the last glacial maximum (LGM), the present (2006-2015) and the end of this century (2100) by mapping our model of Trichodesmium growth onto inferred global surface ocean fields of pCO2 , temperature, light and Fe. We conclude that growth rate was severely limited by low pCO2 at the LGM, that current pCO2 levels do not significantly limit Trichodesmium growth and thus, the potential for enhanced growth from future increases in CO2 is small. We also found that the area of the ocean where sea surface temperatures (SST) are within Trichodesmium's thermal niche increased by 32% from the LGM to present, but further increases in SST due to continued global warming will reduce this area by 9%. However, the range reduction at the equator is likely to be offset by enhanced growth associated with expansion of regions with optimal or near optimal Fe and light availability. Between now and 2100, the ocean area of optimal SST and irradiance is projected to increase by 7%, and the ocean area of optimal SST, irradiance and iron is projected to increase by 173%. Given the major contribution of this keystone species to annual N2 fixation and thus pelagic ecology, biogeochemistry and CO2 sequestration, the projected increase in the geographical range for optimal growth could provide a negative feedback to increasing atmospheric CO2 concentrations.


Intraspecific evidence from guppies for correlated patterns of male and female genital trait diversification.
The role of sexual selection in fuelling genital evolution is becoming increasingly apparent from comparative studies revealing interspecific divergence in male genitalia and evolutionary associations between male and female genital traits. Despite this, we know little about intraspecific variance in male genital morphology, or how male and female reproductive traits covary among divergent populations. Here we address both topics using natural populations of the guppy, Poecilia reticulata, a livebearing fish that exhibits divergent patterns of male sexual behaviour among populations. Initially, we performed a series of mating trials on a single population to examine the relationship between the morphology of the male's copulatory organ (the gonopodium) and the success of forced matings. Using a combination of linear measurements and geometric morphometrics, we found that variation in the length and shape of the gonopodium predicted the success of forced matings in terms of the rate of genital contacts and insemination success, respectively. We then looked for geographical divergence in these traits, since the relative frequency of forced matings tends to be greater in high-predation populations. We found consistent patterns of variation in male genital size and shape in relation to the level of predation, and corresponding patterns of (co)variation in female genital morphology. Together, these data enable us to draw tentative conclusions about the underlying selective pressures causing correlated patterns of divergence in male and female genital traits, which point to a role for sexually antagonistic selection.


Extreme haplotype variation in the desiccation-tolerant clubmoss Selaginella lepidophylla.
Plant genome size varies by four orders of magnitude, and most of this variation stems from dynamic changes in repetitive DNA content. Here we report the small 109 Mb genome of Selaginella lepidophylla, a clubmoss with extreme desiccation tolerance. Single-molecule sequencing enables accurate haplotype assembly of a single heterozygous S. lepidophylla plant, revealing extensive structural variation. We observe numerous haplotype-specific deletions consisting of largely repetitive and heavily methylated sequences, with enrichment in young Gypsy LTR retrotransposons. Such elements are active but rapidly deleted, suggesting "bloat and purge" to maintain a small genome size. Unlike all other land plant lineages, Selaginella has no evidence of a whole-genome duplication event in its evolutionary history, but instead shows unique tandem gene duplication patterns reflecting adaptation to extreme drying. Gene expression changes during desiccation in S. lepidophylla mirror patterns observed across angiosperm resurrection plants.


Remote access and care: A comparison of Queensland women's maternity care experience according to area of residence.
This study fills a gap in the literature with a quantitative comparison of the maternity care experiences of women in different geographic locations in Queensland, Australia. Data from a large-scale survey were used to compare women's care experiences according to Australian Standard Geographical Classification (major city, inner regional, outer regional, remote and very remote). Compared to the other groups, women from remote or very remote areas were more likely to be younger, live in an area with poorer economic resources, identify as Aboriginal and/or Torres Strait Islander and give birth in a public facility. They were more likely to travel to another city, town or community for birth. In adjusted analyses women from remote areas were less likely to have interventions such as electronic fetal monitoring, but were more likely to give birth in an upright position and be able to move around during labour. Women from remote areas did not differ significantly from women from major cities in their satisfaction with interpersonal care. Antenatal and postpartum care was lacking for rural women. In adjusted analyses they were much less likely to have booked for maternity care by 18 weeks gestation, to be telephoned or visited by a care provider in the first 10 days after birth. Despite these differences, women from remote areas were more likely to be breastfeeding at 13 weeks and confident in caring for their baby at home. Findings support qualitative assertions that remote and rural women are disadvantaged in their access to antenatal and postnatal care by the need to travel for birth, however, other factors such as age were more likely to be significant barriers to high quality interpersonal care. Improvements to maternity services are needed in order to address inequalities in maternity care particularly in the postnatal period.


Corrosion of dental nickel-aluminum bronze with a minor gold content-mechanism and biological impact.
To study corrosion and to evaluate biological effects in vitro of corrosion products of a copper-aluminum-nickel alloy with 2% gold. The alloy NPGtrade mark+2 with the nominal composition Cu:77.3; Al:7.8; Ni:4.3; Fe:3.0; Zn:2.7; Au:2.0; and Mn:1.7 was characterized. Static immersion in acidic saline, pH 2.2-2.4, was used to determine release of metallic elements in a milieu simulating the condition of plaque build-up in interproximal areas of the tooth. Corrosion and surface reactions in saline and artificial saliva were studied by electrochemical techniques including registration of open-circuit-potentials, polarization curves and impedance spectra. Extracts were made in cell culture media and acidic saline and used for MTT test for cytotoxicity and HET-CAM method for irritation. The mean amount of elements released in the acidic saline were in microg cm(-2) : Cu:632; Al:210; Ni:144; Fe:122; Zn:48; Mn:52. No protective film was formed on the surface of the alloy, as extensive corrosion was observed in both saline and artificial saliva. The corrosion rate was higher in saline than in artificial saliva. Acidic extracts of the alloy diluted up to 64 times reduced cell viability with 80% or more. The extract induced coagulation of the blood vessels of the CAM and was rated as moderate irritant solution. The nickel-aluminum bronze showed high corrosion rate caused by an inability to create a protective surface layer. High levels of toxic elements were found after static immersion testing, and the corrosion products had a distinct adverse effect on the biological activity.


An Electrochemical DNA Biosensor Developed on a Nanocomposite Platform of Gold and Poly(propyleneimine) Dendrimer.
An electrochemical DNA nanobiosensor was prepared by immobilization of a 20mer thiolated probe DNA on electro-deposited generation 4 (G4) poly(propyleneimine) dendrimer (PPI) doped with gold nanoparticles (AuNP) as platform, on a glassy carbon electrode (GCE). Field emission scanning electron microscopy results confirmed the codeposition of PPI (which was linked to the carbon electrode surface by C-N covalent bonds) and AuNP ca 60 nm. Voltammetric interrogations showed that the platform (GCE/PPI-AuNP) was conducting and exhibited reversible electrochemistry (E°' = 235 mV) in pH 7.2 phosphate buffer saline solution (PBS) due to the PPI component. The redox chemistry of PPI was pH dependent and involves a two electron, one proton process, as interpreted from a 28 mV/pH value obtained from pH studies. The charge transfer resistance (Rct) from the electrochemical impedance spectroscopy (EIS) profiles of GCE/PPI-AuNP monitored with ferro/ferricyanide (Fe(CN)₆3-/4-) redox probe, decreased by 81% compared to bare GCE. The conductivity (in PBS) and reduced Rct (in Fe(CN)₆3-/4-) values confirmed PPI-AuNP as a suitable electron transfer mediator platform for voltammetric and impedimetric DNA biosensor. The DNA probe was effectively wired onto the GCE/PPI-AuNP via Au-S linkage and electrostatic interactions. The nanobiosensor responses to target DNA which gave a dynamic linear range of 0.01 - 5 nM in PBS was based on the changes in Rct values using Fe(CN)₆3-/4- redox probe.


Antisolvent crystallisation is a potential technique to prepare engineered lactose with promising aerosolisation properties: effect of saturation degree.
Engineered lactose particles were prepared by anti-solvent crystallisation technique using lactose solutions with different saturation degrees. In comparison to commercial lactose, engineered lactose particles exhibited less elongated and more irregular shape (large aggregates composed of smaller sub-units), rougher surface texture, higher specific surface area, and different anomer form. Engineered lactose powders demonstrated smaller bulk density, smaller tap density, and higher porosity than commercial lactose powder. Dry powder inhaler (DPI) formulations containing engineered lactose and salbutamol sulphate as a model drug demonstrated improved drug content homogeneity and higher amounts of drug delivered to lower airway regions. Higher fine particle fraction of drug was obtained in the case of lactose powders with higher porosity, higher specific surface area and higher fine particle content (<5 μm). The results indicated that the higher the saturation degree of lactose solution used during crystallisation the smaller the specific surface area, the higher the amorphous lactose content, and the higher the β-lactose content of engineered lactose particles. Also, lactose powders obtained from lactose solution with higher degree of saturation showed higher bulk and tap densities and smaller porosity. Engineered lactose powders crystallized from lower saturation degree (20% and 30% w/v) deposited higher amounts of drug on lower airway regions. In conclusion, this study demonstrated that it is possible to prepare engineered lactose particles with favourable properties (e.g. higher fine particle fraction and better drug content homogeneity) for DPI formulations by using lactose solutions with lower degree of saturation during crystallisation process.


Biochemical and crystallographic investigations into isonitrile formation by a nonheme iron-dependent oxidase/decarboxylase.
The isonitrile moiety is found in marine sponges and some microbes, where it plays a role in processes such as virulence and metal acquisition. Until recently only one route was known for isonitrile biosynthesis, a condensation reaction that brings together a nitrogen atom of l-Trp/l-Tyr with a carbon atom from ribulose-5-phosphate. With the discovery of ScoE, a mononuclear Fe(II) α-ketoglutarate-dependent dioxygenase from Streptomyces coeruleorubidus, a second route was identified. ScoE forms isonitrile from a glycine adduct, with both the nitrogen and carbon atoms coming from the same glycyl moiety. This reaction is part of the nonribosomal biosynthetic pathway of isonitrile lipopeptides. Here, we present structural, biochemical, and computational investigations of the mechanism of isonitrile formation by ScoE, an unprecedented reaction in the mononuclear Fe(II) α-ketoglutarate-dependent dioxygenase superfamily. The stoichiometry of this enzymatic reaction is measured, and multiple high-resolution (1.45-1.96 Å resolution) crystal structures of Fe(II)-bound ScoE are presented, providing insight into the binding of substrate, (R)-3-((carboxylmethyl)amino)butanoic acid (CABA), cosubstrate α-ketoglutarate, and an Fe(IV)=O mimic oxovanadium. Comparison to a previously published crystal structure of ScoE suggests that ScoE has an "inducible" α-ketoglutarate binding site, in which two residues arginine-157 and histidine-299 move by approximately 10 Å from the surface of the protein into the active site to create a transient α-ketoglutarate binding pocket. Together, data from structural analyses, site-directed mutagenesis, and computation provide insight into the mode of α-ketoglutarate binding, the mechanism of isonitrile formation, and how the structure of ScoE has been adapted to perform this unusual chemical reaction.


The effect of exposure to biomass smoke on respiratory symptoms in adult rural and urban Nepalese populations.
Half of the world's population is exposed to household air pollution from biomass burning. This study aimed to assess the relationship between respiratory symptoms and biomass smoke exposure in rural and urban Nepal. A cross-sectional study of adults (16+ years) in a rural population (n = 846) exposed to biomass smoke and a non-exposed urban population (n = 802) in Nepal. A validated questionnaire was used along with measures of indoor air quality (PM2.5 and CO) and outdoor PM2.5. Both men and women exposed to biomass smoke reported more respiratory symptoms compared to those exposed to clean fuel. Women exposed to biomass were more likely to complain of ever wheeze (32.0 % vs. 23.5%; p = 0.004) and breathlessness (17.8% vs. 12.0%, p = 0.017) compared to males with tobacco smoking being a major risk factor. Chronic cough was similar in both the biomass and non-biomass smoke exposed groups whereas chronic phlegm was reported less frequently by participants exposed to biomass smoke. Higher PM2.5 levels (≥2 SDs of the 24-hour mean) were associated with breathlessness (OR = 2.10, 95% CI 1.47, 2.99) and wheeze (1.76, 1.37, 2.26). The study suggests that while those exposed to biomass smoke had higher prevalence of respiratory symptoms, urban dwellers (who were exposed to higher ambient air pollution) were more at risk of having productive cough.


Ca2+-mobility in the sarcoplasmic reticulum of ventricular myocytes is low.
The sarcoplasmic reticulum (SR) in ventricular myocytes contains releasable Ca(2+) for activating cellular contraction. Recent measurements of intra-SR (luminal) Ca(2+) suggest a high diffusive Ca(2+)-mobility constant (D(CaSR)). This could help spatially to unify SR Ca(2+)-content ([Ca(2+)](SRT)) and standardize Ca(2+)-release throughout the cell. But measurements of localized depletions of luminal Ca(2+) (Ca(2+)-blinks), associated with local Ca(2+)-release (Ca(2+)-sparks), suggest D(CaSR) may actually be low. Here we describe a novel method for measuring D(CaSR). Using a cytoplasmic Ca(2+)-fluorophore, we estimate regional [Ca(2+)](SRT) from localized, caffeine-induced SR Ca(2+)-release. Caffeine microperfusion of one end of a guinea pig or rat myocyte diffusively empties the whole SR at a rate indicating D(CaSR) is 8-9 microm(2)/s, up to tenfold lower than previous estimates. Ignoring background SR Ca(2+)-leakage in our measurement protocol produces an artifactually high D(CaSR) (>40 microm(2)/s), which may also explain the previous high values. Diffusion-reaction modeling suggests that a low D(CaSR) would be sufficient to support local SR Ca(2+)-signaling within sarcomeres during excitation-contraction coupling. Low D(CaSR) also implies that [Ca(2+)](SRT) may readily become spatially nonuniform, particularly under pathological conditions of spatially nonuniform Ca(2+)-release. Local control of luminal Ca(2+), imposed by low D(CaSR), may complement the well-established local control of SR Ca(2+)-release by Ca(2+)-channel/ryanodine receptor couplons.


Environmental filtering of eudicot lineages underlies phylogenetic clustering in tropical South American flooded forests.
The phylogenetic community assembly approach has been used to elucidate the role of ecological and historical processes in shaping tropical tree communities. Recent studies have shown that stressful environments, such as seasonally dry, white-sand and flooded forests tend to be phylogenetically clustered, arguing for niche conservatism as the main driver for this pattern. Very few studies have attempted to identify the lineages that contribute to such assembly patterns. We aimed to improve our understanding of the assembly of flooded forest tree communities in Northern South America by asking the following questions: are seasonally flooded forests phylogenetically clustered? If so, which angiosperm lineages are over-represented in seasonally flooded forests? To assess our hypotheses, we investigated seasonally flooded and terra firme forests from the Magdalena, Orinoco and Amazon Basins, in Colombia. Our results show that, regardless of the river basin in which they are located, seasonally flooded forests of Northern South America tend to be phylogenetically clustered, which means that the more abundant taxa in these forests are more closely related to each other than expected by chance. Based on our alpha and beta phylodiversity analyses we interpret that eudicots are more likely to adapt to extreme environments such as seasonally flooded forests, which indicates the importance of environmental filtering in the assembly of the Neotropical flora.


Pathways of infusate loss during convection-enhanced delivery into the putamen nucleus.
New strategies aiming to treat Parkinson's disease, such as delivery of trophic factors via protein infusion or gene transfer, depend upon localized intracerebral infusion, mainly into the putamen nucleus. Convection-enhanced delivery (CED) has been proposed as a method to improve intracerebral distribution of therapies. Yet analysis of controversial results during the clinical translation of these strategies suggests that intracerebral misdistribution of infusate may have affected the outcomes by limiting the amount of treatment into the target region. This study aimed to identify possible pathways of infusate loss and their relative impact in the success of targeted CED into the postcommissural ventral putamen nucleus. Thirteen adult macaque monkeys received intraputaminal CED infusions of 100 µl of 2.0 mM gadoteridol and bromophenol blue (0.16 mg/ml) solution at a rate of 1.0 µl/min under intraoperative magnetic resonance imaging (MRI) guidance. Quantitative maps of infusate concentration were computed at 10-min intervals throughout the procedure in a 3-Tesla MRI scanner. The fraction of tracer lost from the putamen as well as the path of loss were evaluated and quantified for each infusion. All injections (total 22) were successfully placed in the ventral postcommissural putamen nucleus. Four major paths of infusate loss from the putamen were observed: overflow across putamen boundaries, perivascular flow along large blood vessels, backflow along the inserted catheter and catheter tract leakage into the vacated catheter tract upon catheter removal. Overflow loss was observed within the first 30 µl of infusion in all cases. Measurable tracer loss following the path of an artery out of the putamen was observed in 15 cases, and in 8 of these cases, the loss was greater than 10% of infusate. Backflow that exited the putamen was observed in 4 cases and led to large loss of infusate (80% in 1 case) into the corona radiata. Loss into the vacated catheter tract amounted only to a few microliters. Our analysis demonstrates that after controlling for targeting, catheter type, infusion rate and infusate, the main issues during surgical planning are the identification of appropriate infusate volume that matches the target area, as well as mapping the regional vasculature as it may become a pathway for infusate loss. Most importantly, these results underscore the significance of presurgical planning for catheter placement and infusion, and the value of imaging guidance to ensure targeting accuracy.


An automated computerized method using Finapres for measuring cardiovascular reflexes.
1. The major drawback of the cardiovascular reflex tests used to study autonomic failure is the time involved in calculating the results. To overcome this disadvantage, we have developed an automated computerized program using a FINger Arterial PRESsure instrument for the measurement of beat-to-beat heart rate and blood pressure on a finger. 2. This program calculates heart rate variability during three standardized tests, forced breathing, standing up and the Valsalva manoeuvre, and records blood pressure values in response to standing up and sustained handgrip. The time taken to perform the test and to calculate the results is usually 25 min. 3. The reproducibility of the tests in 21 normal subjects was comparable with the reproducibility obtained with conventional test methods using an ECG and a sphygmomanometer. 4. In addition, we determined the age-dependent normal values of the seven test parameters in 124 subjects aged 20-90 years. 5. Using this program in 10 patients with longstanding (14-50 years) complicated diabetes, in each of them four or more abnormal test results were found.


Evaluation of a Triatoma infestans elimination program by the decrease of Trypanosoma cruzi infection frequency in children younger than 10 years, Chile, 1991-1998.
Chagas disease is widespread in Chile, distributed in rural and periurban areas in the 7 most northern regions of the country. The principal vector of Trypanosoma cruzi is Triatoma infestans. The interruption of the domestic cycle of transmission of T. cruzi has been attempted by health education, human housing improving, and elimination of the vector by means of systematic insecticide spraying of human dwellings. Spraying with insecticides has been supported by Chile's health authorities and has been carried out for the last 12 years. A total of 13,280 children (aged up to 10 years) were randomly selected from 47 counties in the area of Chile endemic for Chagas disease, and blood samples were collected to determine the levels of antibodies to T. cruzi by indirect hemagglutination and enzyme-linked immunosorbent assay tests. The results of the tests were analyzed to determine the changes that occurred after 12 years of insecticide spraying of dwellings to eliminate T. infestans infestation. A total of 142 (1.1%) samples of children showed antibodies to T. cruzi. This rate is significantly lower than the data generated in similar studies conducted in 1982-1985. The following reduction in prevalence rates were observed in each of the 7 endemic regions of the country: region I, 5.5-0.3%; region II, 6.6-0.3%; region III, 9.8-1.0%; region IV, 7.2-2.0%; region V, 5.2-1.9%, Metropolitan region, 1.4-0.6%; and region VI, 1.4-0.4%. Serovigilance of T. cruzi antibodies level represents a novel approach that may allow the evaluation of the impact of the vector elimination program. The results identify regions that need to strengthen the efforts to reduce the insect infestation of dwellings.


Association of Central Adiposity With Adverse Cardiac Mechanics: Findings From the Hypertension Genetic Epidemiology Network Study.
Central obesity, defined by increased waist circumference or waist:hip ratio (WHR), is associated with increased cardiovascular events, including heart failure. However, the pathophysiological link between central obesity and adverse cardiovascular outcomes remains poorly understood. We hypothesized that central obesity and larger WHR are independently associated with worse cardiac mechanics (reduced left ventricular strain and systolic [s'] and early diastolic [e'] tissue velocities). We performed speckle-tracking analysis of echocardiograms from participants in the Hypertension Genetic Epidemiology Network (HyperGEN) study, a population- and family-based epidemiological study (n=2181). Multiple indices of systolic and diastolic cardiac mechanics were measured. We evaluated the association between central obesity and cardiac mechanics using multivariable-adjusted linear mixed-effects models to account for relatedness among participants. The mean age of the cohort was 51±14 years, 58% were women, and 47% were black. Mean body mass index was 30.8±7.1 kg/m(2), waist circumference was 102±17 cm, WHR was 0.91±0.08, and 80% had central obesity based on waist circumference and WHR criteria. After adjusting for multiple potential confounders (including age, sex, race, physical activity, body mass index, heart rate, smoking status, systolic blood pressure, fasting glucose, total cholesterol, antihypertensive medication use, glomerular filtration rate, left ventricular mass index, wall motion abnormalities, and ejection fraction), central obesity and WHR remained associated with worse global longitudinal strain, early diastolic strain rate, s' velocity, and e' velocity (P<0.05 for all comparisons). There were no significant statistical interactions between WHR and obesity status. In this cross-sectional study of participants with multiple comorbidities, central obesity was found to be associated with adverse cardiac mechanics.


Phosphoinositides are essential coactivators for p21-activated kinase 1.
Phospholipid-enriched membranes such as the plasma membrane can serve as direct regulators of kinase signaling. Pak1 is involved in growth factor signaling at the plasma membrane, and its dysregulation is implicated in cancer. Pak1 adopts an autoinhibited conformation that is relieved upon binding to membrane-bound Rho GTPases Rac1 or Cdc42, but whether lipids also regulate Pak1 in vivo is unknown. We show here that phosphoinositides, particularly PIP(2), potentiate Rho-GTPase-mediated Pak1 activity. A positively charged region of Pak1 binds to phosphoinositide-containing membranes, and this interaction is essential for membrane recruitment and activation of Pak1 in response to extracellular signals. Our results highlight an active role for lipids as allosteric regulators of Pak1 and suggest that Pak1 is a "coincidence detector" whose activation depends on GTPases present in phosphoinositide-rich membranes. These findings expand the role of phosphoinositides in kinase signaling and suggest how altered phosphoinositide metabolism may upregulate Pak1 activity in cancer cells.


Menopausal status and distensibility of the common carotid artery.
Although several studies have shown that exogenous estrogens have beneficial effects on arterial characteristics, the effect of endogenous estrogen on the vascular system is still unknown. In this study, distensibility, an indicator of arterial elasticity, of the common carotid artery was compared in pre- and postmenopausal women. The study comprised 93 premenopausal and 93 postmenopausal women of similar age (range, 43 to 55 years). Women were selected from respondents to a mailed questionnaire about the menopause, which was sent to all women aged 40 to 60 years in the Dutch town of Zoetermeer (n=12 675). Postmenopausal women who were at least 3 years past natural menopause or whose menses had stopped naturally before age 48, were age-matched with premenopausal women with regular menses and without menopausal complaints. The selection aimed at maximizing the contrast in estrogen status between pre- and postmenopausal women of the same age. Distensibility of the carotid artery was measured noninvasively with B-mode ultrasound and a vessel wall movement detector system. Arterial distensibility is expressed as the change in arterial diameter (distension, DeltaD) with the cardiac cycle, adjusted for lumen diameter, pulse pressure, and mean arterial blood pressure. Compared with premenopausal women, postmenopausal women had significantly lower arterial distension (DeltaD 370.5 microm [SE 9.5] versus 397.3 microm [SE 9.6]). These results suggest that the distensibility of the common carotid artery is negatively affected by natural menopause in presumed healthy women.


Early stage kinetics of polyelectrolyte complex coacervation monitored through stopped-flow light scattering.
Polyelectrolyte complexes (PECs) between poly(acrylic acid) (PAA) and poly(diallyldimethylammonium chloride) (PDADMAC), a model system forming coacervate particles via electrostatic interaction at pH 10, were prepared by a stopped-flow (SF) fast mixing technique at different mixing charge ratios (z) and ionic strengths. Both PEC final morphologies prepared by either SF or manual one-shot mixing are similar at bench time. In situ light scattering combined with the SF technique pointed-out, however, the presence of three distinct early stage kinetic behaviors in the formation of PECs. The first stage observed at low z is ascribed to the relaxation/reorganization of soluble PECs. At higher z or in the presence of salt, a second stage is found corresponding to the aggregation and/or rearrangement of small soluble PECs into larger structures. Redistribution of excess charges among those PECs produces some neutral condensed coacervate droplets as well, coexisting with PECs in a wide range of mixing ratios. Finally, a last process featured with bell-shaped curves indicates the full coacervation that quickens while approaching charge neutrality and/or at higher ionic strength.


'Important… but of low status': male education leaders' views on gender in medicine.
The implementation of and communication about matters associated with gender in medical education have been predominantly perceived as women's issues. This study aimed to explore attitudes towards and experiences of gender-related issues among key male members of faculties of medicine. We conducted semi-structured interviews with 20 male education leaders from the six medical schools in Sweden. The interviews were analysed qualitatively using a modified grounded theory approach. The core category--'important… but of low status'--reflects ambivalent attitudes towards gender-related issues in medicine among male education leaders. All informants were able to articulate why gender matters. As doctors, they saw gender as a determinant of health and, as bystanders, they had witnessed inequalities and the wasting of women's competence. However, they had doubts about gender-related issues and found them to be overemphasised. Gender education was seen as a threat to medical school curricula as a consequence of the time and space it requires. Gender-related issues were considered to be unscientifically presented, to mostly concern women's issues and to tend to involve 'male bashing' (i.e. gender issues were often labelled as ideological and political). Interviewees asked for facts and knowledge, but questioned specific lessons and gender theory. Experiences of structural constraints, such as prejudice, hierarchies and homosociality, were presented, making gender education difficult and downgrading it. The results indicate that male faculty leaders embrace the importance of gender-related issues, but do not necessarily recognise or defend their impact on an area of significant knowledge and competence in medicine. To change this and to engage more men in gender education, faculty measures are needed to counteract prejudice and to upgrade the time allocation, merits and status of gender implementation work. Based on our findings, we present and discuss possible ways to interest more men and to improve gender education in medicine.


NTP Toxicology and Carcinogenesis Studies of Dichloromethane (Methylene Chloride) (CAS No. 75-09-2) in F344/N Rats and B6C3F1 Mice (Inhalation Studies).
Dichloromethane is widely used in industrial processes, food preparation, and agriculture. In industry, dichloromethane is used as a solvent in paint removers, degreasing agents, aerosol propellants, and triacetate solutions; as a blowing agent in flexible urethane foams; and as a process solvent in the manufacture of steroids, antibiotics, vitamins, and tablet coatings. The use of dichloromethane as an extraction solvent for spice oleoresins, hops, and caffeine from coffee has been approved by the U.S. Food and Drug Administration. Dichloromethane has been used as an inhalation anesthetic and as a fumigant for grain and strawberries. Toxicology and carcinogenesis studies of dichloromethane (99% pure) were conducted by inhalation exposure of groups of 50 male and 50 female F344/N rats and B6C3F1 mice 6 hours per day, 5 days per week, for 102 weeks. The exposure concentrations used (0, 1,000, 2,000, or 4,000 ppm for rats and 0, 2,000, or 4,000 ppm for mice) were selected on the basis of results from 13-week inhalation studies in which groups of 10 rats and 10 mice of each sex were exposed to dichloromethane at concentrations of 525-8,400 ppm 6 hours per day, 5 days per week. During the 2-year studies in rats, body weight gains for exposed males and females were comparable to those of the chamber controls. The survival of exposed male rats was comparable to that of the chamber controls; however, the survival of all groups of males at the termination of the study was low (control, 16/50; low dose, 16/50; mid dose, 17/50; high dose, 9/50). Most of the early deaths among male rats occurred during the final weeks of the study; the survival of male rats through week 86 of the study was 36/50, 39/50, 37/50, and 33/50. This decreased survival is believed to be related to the high incidence of leukemia (34/50; 26/50; 32/50; 35/50). Survival of female rats exposed at 4,000 ppm was reduced relative to that of the chamber controls (30/50; 22/50; 22/50; 15/50); leukemia occurred frequently in all female rat groups. Final mean body weights of high dose male mice and low and high dose female mice were 10%-17% lower than those of the chamber controls; these reductions occurred during the last 16 weeks of the study. The survival of dosed male mice and high dose female mice was reduced relative to that of the chamber controls (male: control, 39/50; low dose, 24/50; high dose, 11/50; female: 25/50; 25/50; 8/50). This reduced survival may have been due to the chemically induced development of liver and lung neoplasia in male and female mice. Increased incidences of benign mammary gland lesions (adenomas and fibroadenomas) occurred in male and female rats exposed to dichloromethane (male: 0/50; 0/50; 2/50; 5/50; female: 5/50; 11/50; 13/50; 23/50). The incidence of malignant mammary gland neoplasms was not increased in female rats (2/50; 2/50; 2/50; 0/50); none was observed in male rats. In addition, integumentary system tumors in the area of the mammary chain occurred with a positive trend in male rats (subcutaneous tissue fibroma or sarcoma: 1/50; 1/50; 2/50; 5/50); the combined incidence of all tumors in the mammary area in male rats was 1/50, 1/50, 4/50, and 9/50. Exposure to dichloromethane was associated with increased incidences of hepatic hemosiderosis, cytomegaly, cytoplasmic vacuolization, necrosis, granulomatous inflammation, and bile duct fibrosis in both male and female rats. There was a positive but marginal trend in the incidence of hepatocellular neoplastic nodules or hepatocellular carcinomas (combined) in female rats (2/50; 1/50; 4/50; 5/50). The incidence of squamous metaplasia of the nasal cavity was increased in female rats exposed at 4,000 ppm (1/50; 2/50; 3/50; 9/50) but not in males (4/50; 5/50; 3/50; 3/50). No nasal cavity tumors were observed in rats. The increased incidences of mononuclear cell leukemia in mid dose and high dose female rats (17/50; 17/50; 23/50; 23/50) were statistically significant by age-adjusted analyses. In male rats, mesotheliomas (arising primarily from the tunica vaginalis) occurred at increas) occurred at increased incidences (0/50; 2/50; 5/50; 4/50). Lung tumors occurred at increased incidences in male and female mice exposed to dichloromethane (alveolar/bronchiolar adenomas: male - 3/50; 19/50; 24/50; female - 2/50; 23/48; 28/48; alveolar/bronchiolar carcinomas: male - 2/50; 10/50; 28/50; female - 1/50; 13/48; 29/48). Cytologic degeneration of the liver was observed at increased incidences in high dose male and dosed female mice (male: 0/50; 0/49; 22/49; female: 0/50; 23/48; 21/48). Incidences of hepatocellular adenomas or hepatocellular carcinomas (combined) were increased in high dose male and dosed female mice (male: 22/50; 24/49; 33/49; female: 3/50; 16/48; 40/48). There were also dose-related increases in the numbers of mice bearing multiple lung or liver neoplasms. Dose-related increases were observed in the incidences of testicular atrophy in male mice and uterine and ovarian atrophy in female mice; these effects are considered to be secondary responses to neoplasia. An audit of the experimental data was conducted for the 2-year studies of dichloromethane. No data discrepancies were found that influenced the final interpretations. Under the conditions of these inhalation studies, there was some evidence of carcinogenicity of dichloromethane for male F344/N rats as shown by an increased incidence of benign neoplasms of the mammary gland. There was clear evidence of carcinogenicity of dichloromethane for female F344/N rats as shown by increased incidences of benign neoplasms of the mammary gland. There was clear evidence of carcinogenicity of dichloromethane for male and female B6C3F1 mice, as shown by increased incidences of alveolar/bronchiolar neoplasms and of hepatocellular neoplasms. Synonyms: DCM; methylene chloride


Emergence and spread of drug resistant influenza: A two-population game theoretical model.
The potential for emergence of antiviral drug resistance during influenza pandemics has raised great concern for public health. Widespread use of antiviral drugs is a significant factor in producing resistant strains. Recent studies show that some influenza viruses may gain antiviral drug resistance without a fitness penalty. This creates the possibility of strategic interaction between populations considering antiviral drug use strategies. To explain why, we develop and analyze a classical 2-player game theoretical model where each player chooses from a range of possible rates of antiviral drug use, and payoffs are derived as a function of final size of epidemic with the regular and mutant strain. Final sizes are derived from a stochastic compartmental epidemic model that captures transmission within each population and between populations, and the stochastic emergence of antiviral drug resistance. High treatment levels not only increase the spread of the resistant strain in the subject population but also affect the other population by increasing the density of the resistant strain infectious individuals due to travel between populations. We found two Nash equilibria where both populations treat at a high rate, or both treat at a low rate. Hence the game theoretical analysis predicts that populations will not choose different treatment strategies than other populations, under these assumptions. The populations may choose to cooperate by maintaining a low treatment rate that does not increase the incidence of mutant strain infections or cause case importations to the other population. Alternatively, if one population is treating at a high rate, this will generate a large number of mutant infections that spread to the other population, in turn incentivizing that population to also treat at a high rate. The prediction of two separate Nash equilibria is robust to the mutation rate and the effectiveness of the drug in preventing transmission, but it is sensitive to the volume of travel between the two populations. Model-based evaluations of antiviral influenza drug use during a pandemic usually consider populations in isolation from one another, but our results show that strategic interactions could strongly influence a population's choice of antiviral drug use policy. Furthermore, the high treatment rate Nash equilibrium has the potential to become socially suboptimal (i.e. non-Pareto optimal) under model assumptions that might apply under other conditions. Because of the need for players to coordinate their actions, we conclude that communication and coordination between jurisdictions during influenza pandemics is a priority, especially for influenza strains that do not evolve a fitness penalty under antiviral drug resistance.


The mutation spectrum in genomic late replication domains shapes mammalian GC content.
Genome sequence compositions and epigenetic organizations are correlated extensively across multiple length scales. Replication dynamics, in particular, is highly correlated with GC content. We combine genome-wide time of replication (ToR) data, topological domains maps and detailed functional epigenetic annotations to study the correlations between replication timing and GC content at multiple scales. We find that the decrease in genomic GC content at large scale late replicating regions can be explained by mutation bias favoring A/T nucleotide, without selection or biased gene conversion. Quantification of the free dNTP pool during the cell cycle is consistent with a mechanism involving replication-coupled mutation spectrum that favors AT nucleotides at late S-phase. We suggest that mammalian GC content composition is shaped by independent forces, globally modulating mutation bias and locally selecting on functional element. Deconvoluting these forces and analyzing them on their native scales is important for proper characterization of complex genomic correlations.


A phase I/II trial of pazopanib in combination with lapatinib in adult patients with relapsed malignant glioma.
Increased mitogenic signaling and angiogenesis, frequently facilitated by somatic activation of EGF receptor (EGFR; ErbB1) and/or loss of PTEN, and VEGF overexpression, respectively, drive malignant glioma growth. We hypothesized that patients with recurrent glioblastoma would exhibit differential antitumor benefit based on tumor PTEN/EGFRvIII status when treated with the antiangiogenic agent pazopanib and the ErbB inhibitor lapatinib. A phase II study evaluated the antitumor activity of pazopanib 400 mg/d plus lapatinib 1,000 mg/d in patients with grade 4 malignant glioma and known PTEN/EGFRvIII status not receiving enzyme-inducing anticonvulsants (EIAC). The phase II study used a two-stage Green-Dahlberg design for futility. An independent, parallel phase I component determined the maximum-tolerated regimen (MTR) of pazopanib and lapatinib in patients with grade 3/4 glioma receiving EIACs. The six-month progression-free survival (PFS) rates in phase II (n = 41) were 0% and 15% in the PTEN/EGFRvIII-positive and PTEN/EGFRvIII-negative cohorts, respectively, leading to early termination. Two patients (5%) had a partial response and 14 patients (34%) had stable disease lasting 8 or more weeks. In phase I (n = 34), the MTR was not reached. On the basis of pharmacokinetic and safety review, a regimen of pazopanib 600 mg plus lapatinib 1,000 mg, each twice daily, was considered safe. Concomitant EIACs reduced exposure to pazopanib and lapatinib. The antitumor activity of this combination at the phase II dose tested was limited. Pharmacokinetic data indicated that exposure to lapatinib was subtherapeutic in the phase II evaluation. Evaluation of intratumoral drug delivery and activity may be essential for hypothesis-testing trials with targeted agents in malignant glioma.


Ready for the frontline: is early thoracoscopic decortication the new standard of care for advanced pneumonia with empyema?
Video-assisted thoracoscopic decortication (VATD) has been established as an effective and potentially less morbid alternative to open thoracotomy for the management of empyema. However, the timing and role of VATD for advanced pneumonia with empyema is still controversial. In assessing surgical outcome, the authors reviewed their VATD experience in children with empyema or empyema with necrotizing pneumonia. The charts of 42 children who underwent VATD at our institution between July 2001 and July 2005 were retrospectively reviewed for surgical outcome. For purposes of analysis, patients were cohorted into four classes with increasing severity of pneumonia: 1 (-) intraoperative pleural fluid cultures, (-) necrotizing pneumonia, 18 (43%); 2 (+) pleural fluid cultures, (-) necrotizing pneumonia, 10 (24%); 3 (-) pleural fluid cultures, (+) necrotizing pneumonia, 6 (14%); 4 (+) pleural fluid cultures, (+) necrotizing pneumonia, 8 (19%). A P value of < 0.05 via Student's t test or Fischer's exact analysis was considered an indicator of significant difference in the comparison of group outcomes. VATD was successfully completed in all 42 patients with no mortality and without significant morbidity (82% had less than 20 cc blood loss). There was found to be no significant difference (p = NS) in time to surgical discharge (removal of chest tube) among all groups. Hospital length of stay postsurgery was found to be significantly increased between 1 and 4 (6 days vs 9 days; P = 0.038). 14/14 (100%) of children with necrotizing pneumonia were found to have evidence of lung parenchymal preservation with improved aeration on follow-up CT scan and/or chest x-rays. The authors conclude that early VATD in children with advanced pneumonia with empyema is indicated to avoid unnecessarily lengthy hospitalization and prolonged intravenous antibiotic therapy. Furthermore, early VATD can be safely performed in various stages of advanced pneumonia with empyema, promoting lung salvage, and accelerating clinical recovery.


A single amino acid substitution converts a carboxylesterase to an organophosphorus hydrolase and confers insecticide resistance on a blowfly.
Resistance to organophosphorus (OP) insecticides is associated with decreased carboxylesterase activity in several insect species. It has been proposed that the resistance may be the result of a mutation in a carboxylesterase that simultaneously reduces its carboxylesterase activity and confers an OP hydrolase activity (the "mutant ali-esterase hypothesis"). In the sheep blowfly, Lucilia cuprina, the association is due to a change in a specific esterase isozyme, E3, which, in resistant flies, has a null phenotype on gels stained using standard carboxylesterase substrates. Here we show that an OP-resistant allele of the gene that encodes E3 differs at five amino acid replacement sites from a previously described OP-susceptible allele. Knowledge of the structure of a related enzyme (acetylcholinesterase) suggests that one of these substitutions (Gly137 --> Asp) lies within the active site of the enzyme. The occurrence of this substitution is completely correlated with resistance across 15 isogenic strains. In vitro expression of two natural and two synthetic chimeric alleles shows that the Asp137 substitution alone is responsible for both the loss of E3's carboxylesterase activity and the acquisition of a novel OP hydrolase activity. Modeling of Asp137 in the homologous position in acetylcholinesterase suggests that Asp137 may act as a base to orientate a water molecule in the appropriate position for hydrolysis of the phosphorylated enzyme intermediate.


Coping and defending: age differences in maturity of defence mechanisms and coping strategies.
Previous studies have examined either coping strategies or defence mechanisms; however, few have considered both. This research examined age differences in the type of defence mechanisms and coping strategies that people employ. In addition, gender differences, personality, and environmental variables were taken into account. The three age groups used in this study included: 17-23 year olds, 40-47 year olds, and 63-70 year olds. The youngest participants used significantly less mature defence mechanisms and significantly more immature defence mechanisms than the middle-aged and the oldest group. However, there was no significant difference in maturity of defence mechanisms between the middle-aged and the oldest group. In contrast, there were no age differences revealed for effectiveness of coping strategies people employ. One further interesting finding was that people with a higher purpose in life were more likely to use mature defence mechanisms. This research concludes that when developing theories on stress, psychologists might benefit from considering both defence mechanisms and coping strategies.


Melanoma adjuvant therapy.
The risk of recurrence and death after complete surgical resection of clinically detectable primary cutaneous melanoma ranges from low, intermediate to high risk depending on the stage of disease at diagnosis. This is defined by the depth, ulceration status and mitotic rate of the primary tumor, the presence of regional nodal disease or distant metastasis. For high-risk melanoma, adjuvant therapy aims at eradicating melanoma micrometastases in those patients that carry an unacceptable risk of mortality from melanoma recurrence. The ultimate goal of adjuvant therapy is to provide a potential cure before progression into advanced inoperable stages. After decades of randomized clinical trials, interferon-α alone has been shown to have a significant impact on relapse free survival and at high dosage on overall survival. These survival benefits were seen in comparison to observation (E1684 trial) and the ganglioside GMK vaccine (E1694 trial). Advances in our understanding of melanoma molecular biology and host immunology have given ways to novel targets and therapeutic agents that have demonstrated unprecedented results in the management of metastatic disease and currently are being tested in the adjuvant stetting where recent data support a significant clinical impact of adjuvant ipilimumab. Here, we review the standard of care melanoma adjuvant therapy along with the main completed, current and planned clinical trials.


Direct-access to sonographic diagnosis of deep vein thrombosis in general practice: a descriptive cohort study.
Suspicion of deep vein thrombosis (DVT) is common and requires urgent and efficient investigation due to hazardous prognosis. The traditional diagnostic pathway can be complex and time-consuming, and innovative solutions may provide easy access to diagnostics and better use of healthcare resources. We aimed to describe use, clinical outcomes and time used when providing general practitioners (GPs) with a direct-access pathway to hospital-based, single whole-leg compression ultrasound (CUS) for patients with suspected DVT. Furthermore we aimed to describe the resources used in the new direct-access pathway and compare it with the previous pathway. We conducted a 2-year descriptive cohort study (2016-2017) including 449 consecutively referred patients for diagnosis of DVT in a Danish regional hospital. The previous pathway included pre-test at the medical department, a proximal leg CUS if required based on the pre-test and a re-scan if the first CUS was negative. The new pathway included two strategies: 1) a 'yes-no strategy', where GPs referred patients directly to whole-leg CUS and if positive, treated at the medical department and if negative, discharged to the GP, 2) a 'follow-up strategy' where GPs could require that patients were seen at the medical department, irrespective of the CUS result. Data included extractions from the Radiology Information and Patient Administrative Systems, and mean salaries of healthcare professionals at Silkeborg Regional Hospital, Denmark. Descriptive statistics were used to describe prevalence, timelines and costs. GPs referred 318 (71%) patients through the yes-no strategy and 131 (29%) via the follow-up strategy with DVT diagnosed in 48 (15%) and 51 (39%) patients, respectively (p < 0.001). For the 263 patients completed after CUS in the yes-no strategy, median pathway time was 24 min (IQI: 16-36), and for those with DVT (including both strategies) 202 min (IQI: 158-273). Direct-access pathway costs were €49.7 less per patient than the previous pathway. Direct-access to CUS for suspected DVT was achievable, had short time intervals and required fewer resources. The difference in DVT prevalence indicates that GPs distinguish between patients with low and high risk of DVT.


In vitro osteoclast resorption of bone substitute biomaterials used for implant site augmentation: a pilot study.
This observational study examined the resorptive behavior of normal neonatal rabbit osteoclasts grown on slices of bovine cortical bone as compared to samples of commercially available bone substitute biomaterials. It also examined the surface characteristics of these materials. The 11 materials tested fell into 3 groups: (1) bone-derived, including freeze-dried human rib block, human demineralized freeze-dried bone, and deproteinated bovine bone; (2) synthetic hydroxyapatites (HA); and (3) synthetic non-HA, including coated methacrylates and coated silica glass. After 4 days in culture, 1 group of samples of each material underwent scanning electron microscopy (SEM) to evaluate resorptive pitting versus controls, while another group underwent tartrate-resistant acid phosphatase staining and light microscopy to examine osteoclast numbers and morphology. The 2 bovine-derived HA materials also underwent immunohistochemical staining and surface chemistry analysis. While most of these materials supported osteoclast attachment, some spreading, and survival in culture, only the bone-derived materials, with the exception of sintered deproteinated bovine bone, showed large scalloped-edged resorption pits with trails and exposed collagen when examined by SEM, although not to the same extent as unprocessed natural bone material. The HA materials and the sintered deproteinated bovine bone showed evidence of etching with smaller pits but no evidence of resorptive trail formation. The non-HA materials showed no evidence of pit formation or trails. Under immunohistochemical staining, Bio-Oss appeared to be positive for type I collagen after osteoclast activity on its surface, while Osteograf/N showed no positive staining. Surface chemistry analysis revealed nitrogen present in Bio-Oss specimens (0.17% to 0.47%), while there was no nitrogen detected in the Osteograf/N (0.00%); the percent nitrogen observed in normal bovine bone controls was 6.01% to 9.25%. The bone-derived materials supported osteoclast activity on the material surface in a way that facilitated formation of the more complex resorption pits in vitro. Assuming the rate of pit formation observed in vitro mimics that observed in vivo, the quantity and type of osteoclastic remodeling seen on non-bone-derived materials--and perhaps sintered bone-derived materials--would be extremely slow to negligible. Physiologic removal of non-bone-derived bone substitutes in vivo may occur by methods other than osteoclast resorption. Allogenous and xenogenous bone-derived materials that undergo delayed physiologic resorption may be more appropriately used with a staged surgical approach when used in sites intended to support osseointegrated dental implants. The combination of collagen staining and the presence of nitrogen suggest that there may be residual protein in Bio-Oss.


Exploring pre-school vision screening in primary care offices in Alabama.
To investigate practices, barriers, and facilitators of universal pre-school vision screening (PVS) at pediatric primary care offices. Focus group sessions (FGS) were moderated on-site at nine pediatric practices. A semi-structured topic guide was used to standardize and facilitate FGS. Discussions were audiotaped, and transcriptions were used to develop themes. All authors reviewed and agreed on the resultant themes. FGS included 13 physicians and 32 nurses/certified medical assistants (CMAs), of whom 82% personally conducted some facet of PVS. In all practices, nurses/CMAs tested visual acuity (most using a non-recommended test), and physicians completed vision screening with external observation, fix/follow, red reflex, and cover test. Facilitators included (1) accepting that PVS is a routine part of the well-child visit, and (2) using an electronic medical record with prompts to record acuity (eight of nine practices). Barriers were related to difficulty testing pre-schoolers, distractions in the office setting, time constraints, and limited reimbursement. Responsibility for PVS is shared by physicians and nurses/CMAs; thus, interventions to improve PVS should target both. Few practices are aware of new evidence-based PVS tests; thus, active translational efforts are needed to change current primary care practices.


DNA minor groove sensing and widening by the CCAAT-binding complex.
The CCAAT box is a frequent element of eukaryotic promoters, and its specific recognition by the conserved heterotrimeric CCAAT-binding complex (CBC) constitutes a key step in promoter organization and regulation of transcription. Here, we report the crystal structures of the CBC from Aspergillus nidulans in the absence and in complex with double-stranded DNA at 1.8 Å resolution. The histone-like subunits HapC and HapE induce nucleosome-like DNA bending by interacting with the sugar-phosphate backbone. Minor groove sensing and widening by subunit HapB tightly anchor the CBC to the CCAAT box, as shown by structural and biochemical data. Furthermore, crucial interactions of the DNA duplex with subunit HapB provide an explanation for the sequence specificity of the CBC. The herein-described mode of transcription factor binding answers the question of how histone proteins gained sequence specificity for the CCAAT box.


The actin cytoskeleton modulates the activation of iNKT cells by segregating CD1d nanoclusters on antigen-presenting cells.
Invariant natural killer T (iNKT) cells recognize endogenous and exogenous lipid antigens presented in the context of CD1d molecules. The ability of iNKT cells to recognize endogenous antigens represents a distinct immune recognition strategy, which underscores the constitutive memory phenotype of iNKT cells and their activation during inflammatory conditions. However, the mechanisms regulating such "tonic" activation of iNKT cells remain unclear. Here, we show that the spatiotemporal distribution of CD1d molecules on the surface of antigen-presenting cells (APCs) modulates activation of iNKT cells. By using superresolution microscopy, we show that CD1d molecules form nanoclusters at the cell surface of APCs, and their size and density are constrained by the actin cytoskeleton. Dual-color single-particle tracking revealed that diffusing CD1d nanoclusters are actively arrested by the actin cytoskeleton, preventing their further coalescence. Formation of larger nanoclusters occurs in the absence of interactions between CD1d cytosolic tail and the actin cytoskeleton and correlates with enhanced iNKT cell activation. Importantly and consistently with iNKT cell activation during inflammatory conditions, exposure of APCs to the Toll-like receptor 7/8 agonist R848 increases nanocluster density and iNKT cell activation. Overall, these results define a previously unidentified mechanism that modulates iNKT cell autoreactivity based on the tight control by the APC cytoskeleton of the sizes and densities of endogenous antigen-loaded CD1d nanoclusters.


High Plasma Erythropoietin Predicts Incident Fractures in Elderly Men with Normal Renal Function: The MrOS Sweden Cohort.
Preclinical studies on the role of erythropoietin (EPO) in bone metabolism are contradictory. Regeneration models indicate an anabolic effect on bone healing, whereas models on physiologic bone remodeling indicate a catabolic effect on bone mass. No human studies on EPO and fracture risk are available. It is known that fibroblast growth factor 23 (FGF23) affects bone mineralization and that serum concentration of FGF23 is higher in men with decreased estimated glomerular filtration rate (eGFR). Recently, a direct association between EPO and FGF23 has been shown. We have explored the potential association between EPO and bone mineral density (BMD), fracture risk, and FGF23 in humans. Plasma levels of EPO were analyzed in 999 men (aged 69 to 81 years), participating in the Gothenburg part of the population-based Osteoporotic Fractures in Men (MrOS) study, MrOS Sweden. The mean ± SD EPO was 11.5 ± 9.0 IU/L. Results were stratified by eGFR 60 mL/min. For men with eGFR ≥60 mL/min (n = 728), EPO was associated with age (r = 0.13, p < 0.001), total hip BMD (r = 0.14, p < 0.001), intact (i)FGF23 (r = 0.11, p = 0.004), and osteocalcin (r = -0.09, p = 0.022). The association between total hip BMD and EPO was independent of age, body mass index (BMI), iFGF23, and hemoglobin (beta = 0.019, p < 0.001). During the 10-year follow-up, 164 men had an X-ray-verified fracture, including 117 major osteoporotic fractures (MOF), 39 hip fractures, and 64 vertebral fractures. High EPO was associated with higher risk for incident fractures (hazard ratio [HR] = 1.43 per tertile EPO, 95% confidence interval [CI] 1.35-1.63), MOF (HR = 1.40 per tertile EPO, 95% CI 1.08-1.82), and vertebral fractures (HR = 1.42 per tertile EPO, 95% CI 1.00-2.01) in a fully adjusted Cox regression model. In men with eGFR<60 mL/min, no association was found between EPO and BMD or fracture risk. We here demonstrate that high levels of EPO are associated with increased fracture risk and increased BMD in elderly men with normal renal function. © 2019 American Society for Bone and Mineral Research.


Development of an accelerated functional rehabilitation protocol following minimal invasive Achilles tendon repair.
Surgical repair after acute Achilles tendon rupture leads to lower re-rupture rates than non-surgical treatment. After open repair, early functional rehabilitation improves outcome, but there are risks of infection and poor wound healing. Minimal invasive surgery reduces these risks; however, there are concerns about its stability. Consequently, physicians may have reservations about adopting functional rehabilitation. There is still no consensus about the post-operative treatment after minimal invasive repair. The aim of this study was to define the most effective and safe post-operative rehabilitation protocol following minimal invasive repair. A systematic literature search in Embase, MEDLINE and Cochrane Library for prospective trials reporting on early functional rehabilitation after minimal invasive repair was performed. Seven studies were included. One randomized controlled trail, one prospective comparative and five prospective non-comparative studies were identified. Four studies performed full weight bearing, all demonstrating good functional results, an early return to work/sports and high satisfaction. One study allowed early mobilization leading to excellent subjective and objective results. The only randomized controlled trial performed the most accelerated protocol demonstrating a superior functional outcome and fewer complications after immediate full weight bearing combined with free ankle mobilization. The non-comparative study reported high satisfaction, good functional results and an early return to work/sports following combined treatment. Immediate weight bearing in a functional brace, together with early mobilization, is safe and has superior outcome following minimally invasive repair of Achilles tendon rupture. Our recommended treatment protocol provides quality assurance for the patient and reliability for the attending physician. II.


Avian SERPINB11 gene: a marker for ovarian endometrioid cancer in chickens.
As serine and cysteine proteinase inhibitors, serpins, such as SERPINB5, cause ovarian, colorectal and pancreatic adenocarcinomas. We identified SERPINB11 as a novel estrogen-induced gene in chickens during oviduct development. The chicken is a unique animal model for research on human ovarian cancer, because it spontaneously develops epithelial cell-derived ovarian cancer as in women. Therefore, this study investigated the expression pattern, CpG methylation status, and miRNA regulation of the SERPINB11 gene in normal and cancerous ovaries from chickens. Our results indicate that SERPINB11 is most abundant in the glandular epithelium of endometrioid adenocarcinoma of cancerous, but not normal, ovaries of hens. In addition, bisulfite sequencing revealed that about 30% of -110 CpG sites are methylated in ovarian cancer cells, whereas -110 CpG sites are demethylated in normal ovarian cells. Next, we determined whether miR-1582 influences SERPINB11 expression via its 3'UTR and found that it does not directly target the 3'UTR of SERPINB11 mRNA. Therefore, it is unlikely that post-transcriptional regulation influences SERPINB11 expression in the chicken ovary. On the other hand, in human ovarian cancer cells such as OVCAR-3, SKOV-3 and PA-1 cells, immunoreactive SERPINB11 protein was predominant in the cytoplasm and had a similar expression pattern to that in chicken ovarian cancer cells. Collectively, these results suggest that SERPINB11 is a biomarker for chicken ovarian endometrioid carcinoma that could be used for diagnosis and monitoring effects of therapies for the disease in women.


Different proctolin neurons elicit distinct motor patterns from a multifunctional neuronal network.
Distinct motor patterns are selected from a multifunctional neuronal network by activation of different modulatory projection neurons. Subsets of these projection neurons can contain the same neuromodulator(s), yet little is known about the relative influence of such neurons on network activity. We have addressed this issue in the stomatogastric nervous system of the crab Cancer borealis. Within this system, there is a neuronal network in the stomatogastric ganglion (STG) that produces many versions of the pyloric and gastric mill rhythms. These different rhythms result from activation of different projection neurons that innervate the STG from neighboring ganglia and modulate STG network activity. Three pairs of these projection neurons contain the neuropeptide proctolin. These include the previously identified modulatory proctolin neuron and modulatory commissural neuron 1 (MCN1) and the newly identified modulatory commissural neuron 7 (MCN7). We document here that each of these neurons contains a unique complement of cotransmitters and that each of these neurons elicits a distinct version of the pyloric motor pattern. Moreover, only one of them (MCN1) also elicits a gastric mill rhythm. The MCN7-elicited pyloric rhythm includes a pivotal switch by one STG network neuron from playing a minor to a major role in motor pattern generation. Therefore, modulatory neurons that share a peptide transmitter can elicit distinct motor patterns from a common target network.


Sealing ability of mineral trioxide aggregate and super-EBA when used as furcation repair materials: a longitudinal study.
Immediate sealing of furcation perforations enhances the repair process. The purpose of this study was to longitudinally compare the ability of mineral trioxide aggregate (MTA) and Super-EBA to seal furcation perforations. Fifty-one extracted human maxillary molars were decoronated 3 mm above the CEJ, and the roots were amputated 3 mm below the furcation. A #2 high-speed round bur was used to perforate the center of the furcations. The canals were obturated with gutta-percha, and the root ends were sealed with C&B Metabond. Three experimental groups of 15 teeth each were restored with MTA, Super-EBA, or a combination of MTA in the perforation and a Super-EBA dome on the pulpal floor. Six teeth served as controls. Each tooth was affixed to a fluid filtration device and subjected to a pressure of 20 cm H2O. The integrity of the perforation seal was evaluated initially at 30 min for the Super-EBA and the combination groups and at 4 h for the MTA group. Additional measurements were then made at 24 h, 1 week, and 1 month. The controls behaved as expected. A two-way ANOVA revealed a significant difference (p = 0.01) between materials. Tukey's test isolated the difference to Super EBA as producing a superior seal but only at 24 h. There was no significant effect with time (p = 0.57) or the interaction of the materials with time (p = 0.66). All materials sealed the perforations very well. The maximum leakage of all materials was <0.007 microL min(-1) cm H2O(-1).


Coordination of Orofacial Motor Actions into Exploratory Behavior by Rat.
The delineation of sensorimotor circuits that guide exploration begins with an understanding of the pattern of motor outputs [1]. These motor patterns provide a clue to the form of the underlying circuits [2-4] (but see [5]). We focus on the behaviors that rodents use to explore their peripersonal space through goal-directed positioning of their nose, head, and vibrissae. Rodents sniff in response to novel odors, reward expectation, and as part of social interactions [6-12]. Sniffing serves olfaction [13, 14], while whisking synchronized to sniffing serves vibrissa-based touch [6, 15, 16]. We quantify the ethology of exploratory nose and head movements in relation to breathing. We find that sniffing is accompanied by prominent lateral and vertical deflections of the nose, i.e., twitches, which are driven by activation of the deflector nasi muscles [17]. On the timescale of individual breaths, nose motion is rhythmic and has a maximum deflection following the onset of inspiration. On a longer timescale, excursions of the nose persist for several breaths and are accompanied by an asymmetry in vibrissa positioning toward the same side of the face. Such directed deflections can be triggered by a lateralized source of odor. Lastly, bobbing of the head as the animal cranes and explores is phase-locked to sniffing and to movement of the nose. These data, along with prior results on the resetting of the whisk cycle at the onset of inspiration [15, 16, 18], reveal that the onset of each breath initiates a "snapshot" of the orofacial sensory environment. VIDEO ABSTRACT.


Compensatory load redistribution of horses with induced weight-bearing forelimb lameness trotting on a treadmill.
The study was performed to obtain a detailed insight into the load and time shifting mechanisms of horses with unilateral weight-bearing forelimb lameness. Reversible lameness was induced in 11 clinically sound horses by applying a solar pressure model. Three degrees of lameness (subtle, mild and moderate) were induced and compared with sound control measurements. Vertical ground reaction force-time histories of all four limbs were recorded simultaneously on an instrumented treadmill. Four compensatory mechanisms could be identified that served to reduce structural stress, i.e. peak vertical force on the affected limb: (1) with increasing lameness, horses reduced the total vertical impulse per stride; (2) the diagonal impulse decreased selectively in the lame diagonal; (3) the impulse was shifted within the lame diagonal to the hindlimb and in the sound diagonal to the forelimb; (4) the rate of loading and the peak forces were reduced by prolonging the stance duration. Except in the diagonal hindlimb, where peak vertical forces increased slightly in the moderate lameness condition, no equivalent compensatory overload situation was observed in the other limbs. Specific force and time information of all four limbs allow the unequivocal identification of the affected limb.


In vivo efficacy and pharmacokinetics of AC98-6446, a novel cyclic glycopeptide, in experimental infection models.
AC98-6446 is a novel semisynthetic derivative of a natural product related to the mannopeptimycins produced by Streptomyces hygroscopicus. Naturally occurring esterified mannopeptimycins exhibited excellent in vitro activity but only moderate in vivo efficacy against staphylococcal infection. The in vivo efficacy and pharmacokinetics of AC98-6446 were investigated in murine acute lethal, bacterial thigh and rat endocarditis infections. Pharmacokinetics were performed in mice, rats, monkeys, and dogs. Acute lethal infections were performed with several gram-positive isolates: Staphylococcus aureus (methicillin-susceptible and methicillin-resistant staphylococci), vancomycin-resistant Enterococcus faecalis, and penicillin-susceptible and -resistant Streptococcus pneumoniae. The 50% effective dose for all isolates tested ranged from 0.05 to 0.39 mg/kg of body weight after intravenous (i.v.) administration. Vancomycin was more than fivefold less efficacious against all of these same infections. Results of the thigh infection with S. aureus showed a static dose for AC98-6446 of 0.4 mg/kg by i.v. administration. Reduction of counts in the thigh of >2 log(10) CFU were achieved with doses of 1 mg/kg. i.v. administration of 3 mg/kg twice a day for 3 days resulted in a >3 log(10) reduction in bacterial counts of vancomycin-susceptible and -resistant E. faecalis in a rat endocarditis model. Pharmacokinetics of AC98-6446 showed an increase in exposure (area under the concentration-time curve) from mouse to dog species. The i.v. half-life (t(1/2)) increased threefold between rodents and the higher species dosed. Efficacy of AC98-6446 has been demonstrated in several models of infection with resistant gram-positive pathogens. This glycopeptide exhibited bactericidal activity in these models, resulting in efficacy at low doses with reduction in bacterial load.


Whole-genome sequence analysis shows that two endemic species of North American wolf are admixtures of the coyote and gray wolf.
Protection of populations comprising admixed genomes is a challenge under the Endangered Species Act (ESA), which is regarded as the most powerful species protection legislation ever passed in the United States but lacks specific provisions for hybrids. The eastern wolf is a newly recognized wolf-like species that is highly admixed and inhabits the Great Lakes and eastern United States, a region previously thought to be included in the geographic range of only the gray wolf. The U.S. Fish and Wildlife Service has argued that the presence of the eastern wolf, rather than the gray wolf, in this area is grounds for removing ESA protection (delisting) from the gray wolf across its geographic range. In contrast, the red wolf from the southeastern United States was one of the first species protected under the ESA and was protected despite admixture with coyotes. We use whole-genome sequence data to demonstrate a lack of unique ancestry in eastern and red wolves that would not be expected if they represented long divergent North American lineages. These results suggest that arguments for delisting the gray wolf are not valid. Our findings demonstrate how a strict designation of a species under the ESA that does not consider admixture can threaten the protection of endangered entities. We argue for a more balanced approach that focuses on the ecological context of admixture and allows for evolutionary processes to potentially restore historical patterns of genetic variation.


Comparison of the effect of Yoga, Zumba and Aerobics in controlling blood pressure in the Indian population.
This study aims at comparing the effect of Yoga, Zumba dance, and Aerobic exercises in controlling blood pressure among the Indian population without using hypertensive drugs. This study is designed to analyze the effect of Yoga, Zumba, and Aerobics in controlling blood pressure among the Indian population and to determine which of the three is better to treat hypertension without using hypertensive drugs. Hypertension is a disease which is nowadays most commonly seen among children and adolescents and has been found that regular exercise tends to reduce the levels of high blood pressure in a very effective way among which the effects of aerobics exercise on reducing high blood pressure in hypertensive patients had been more convincing that regular aerobics exercise reduces the blood pressure by > 1.5 mmHg.


Properties of virion transactivator proteins encoded by primate cytomegaloviruses.
Human cytomegalovirus (HCMV) is a betaherpesvirus that causes severe disease in situations where the immune system is immature or compromised. HCMV immediate early (IE) gene expression is stimulated by the virion phosphoprotein pp71, encoded by open reading frame (ORF) UL82, and this transactivation activity is important for the efficient initiation of viral replication. It is currently recognized that pp71 acts to overcome cellular intrinsic defences that otherwise block viral IE gene expression, and that interactions of pp71 with the cell proteins Daxx and ATRX are important for this function. A further property of pp71 is the ability to enable prolonged gene expression from quiescent herpes simplex virus type 1 (HSV-1) genomes. Non-human primate cytomegaloviruses encode homologs of pp71, but there is currently no published information that addresses their effects on gene expression and modes of action. The UL82 homolog encoded by simian cytomegalovirus (SCMV), strain Colburn, was identified and cloned. This ORF, named S82, was cloned into an HSV-1 vector, as were those from baboon, rhesus monkey and chimpanzee cytomegaloviruses. The use of an HSV-1 vector enabled expression of the UL82 homologs in a range of cell types, and permitted investigation of their abilities to direct prolonged gene expression from quiescent genomes. The results show that all UL82 homologs activate gene expression, and that neither host cell type nor promoter target sequence has major effects on these activities. Surprisingly, the UL82 proteins specified by non-human primate cytomegaloviruses, unlike pp71, did not direct long term expression from quiescent HSV-1 genomes. In addition, significant differences were observed in the intranuclear localization of the UL82 homologs, and in their effects on Daxx. Strikingly, S82 mediated the release of Daxx from nuclear domain 10 substructures much more rapidly than pp71 or the other proteins tested. All UL82 homologs stimulated the early release of ATRX from nuclear domain 10. All of the UL82 homolog proteins analysed activated gene expression, but surprising differences in other aspects of their properties were revealed. The results provide new information on early events in infection with cytomegaloviruses.


Determinants of fruit and vegetable consumption among children and adolescents: a review of the literature. Part I: Quantitative studies.
In order to more effectively promote fruit and vegetable intake among children and adolescents, insight into determinants of intake is necessary. We conducted a review of the literature for potential determinants of fruit and vegetable intake in children and adolescents. Papers were identified from Medline and PsycINFO by using all combinations of the search terms: "fruit(s) or vegetable(s)" and "children or adolescents". Quantitative research examining determinants of fruit and/or vegetable intake among children and adolescents aged 6-18 years were included. The selection and review process was conducted according to a four-step protocol resulting in information on country, population, design, methodology, theoretical basis, instrument used for measuring intake, statistical analysis, included independent variables, and effect sizes. Ninety-eight papers were included. A large number of potential determinants have been studied among children and adolescents. However, for many presumed determinants convincing evidence is lacking, mostly because of paucity of studies. The determinants best supported by evidence are: age, gender, socio-economic position, preferences, parental intake, and home availability/accessibility. Girls and younger children tend to have a higher or more frequent intake than boys and older children. Socio-economic position, preferences, parental intake, and home availability/accessibility are all consistently positively associated with intake. The determinants most consistently supported by evidence are gender, age, socio-economic position, preferences, parental intake and home availability/accessibility. There is a need for internationally comparative, longitudinal, theory-based and multi-level studies taking both personal and environmental factors into account. This paper is published as part of the special Pro Children series in the International Journal of Behavioral Nutrition and Physical Activity. Please see [http://www.ijbnpa.org/content/3/1/26] for the relevant editorial.


Inhibition of TGF-beta-induced collagen production in rabbit flexor tendons.
Postoperative adhesions frequently compromise the success of flexor tendon repair. Manipulation of growth factors responsible for scar formation may be a method of decreasing adhesion formation. Transforming growth factor beta (TGF-beta) is a key cytokine in the pathogenesis of tissue fibrosis. The purpose of this study was to examine the effectiveness of TGF-beta neutralizing antibody in blocking TGF-beta-induced collagen I production in rabbit flexor tendons in vitro. Sheath fibroblasts, epitenon tenocytes, and endotenon tenocytes were obtained from rabbit flexor tendons. Each cell culture was supplemented with 1 ng/mL of TGF-beta along with increasing doses of TGF-beta neutralizing antibody (0.1-2.0 microg/mL). Collagen I production was measured by enzyme-linked immunoabsorbent assay and TGF-beta bioactivity was measured by the luciferase assay. Results were compared with TGF-beta alone and unsupplemented controls. The addition of neutralizing antibody significantly reduced TGF-beta-induced collagen I production in a dose-dependent manner in all 3 cell cultures. TGF-beta bioactivity was also reduced by its neutralizing antibody. This study shows that TGF-beta inhibition through its neutralizing antibody was effective in cultured flexor tendon cells. The results encourage further experiments that use such agents to modulate flexor tendon wound healing in in vivo models in the hope of eventually blocking the effect of TGF-beta on flexor tendons clinically.


Genome sequencing supports a multi-vertex model for Brassiceae species.
The economically important Brassica genus is a good system for studying the evolution of polyploids. Brassica genomes have undergone whole genome triplication (WGT). Subgenome dominance phenomena such as biased gene fractionation and dominant gene expression were observed in tripled genomes of Brassica. The genome of radish (Raphanus sativus), another important crop of tribe Brassiceae, was derived from the same WGT event and shows similar subgenome dominance. These findings and molecular dating indicate that radish occupies a similar evolutionary origin as that of Brassica species. Here, we extended the Brassica "triangle of U" to a multi-vertex model. This model describes the relationships or the potential of using more Brassiceae mesohexaploids in the creation of new allotetraploid oil or vegetable crop species.


Impact of magnetic resonance imaging on preoperative planning for breast cancer surgery.
To review the impact of preoperative breast magnetic resonance imaging on the management of planned surgery, and the appropriateness of any resulting alterations. Retrospective review. A private hospital in Hong Kong. PATIENTS; For the 147 consecutive biopsy-proven breast cancer patients who underwent preoperative magnetic resonance imaging to determine tumour extent undergoing operation by a single surgeon between 1 January 2006 and 31 December 2009, the impact of magnetic resonance imaging findings was reviewed in terms of management alterations and their appropriateness. The most common indication for breast magnetic resonance imaging was the presence of multiple indeterminate shadows on ultrasound scans (53%), followed by ill-defined border of the main tumour on ultrasound scans (19%). In 66% (97 out of 147) of the patients, the extent of the operation was upgraded. Upgrading entailed: lumpectomy to wider lumpectomy (23 out of 97), lumpectomy to mastectomy (47 out of 97), lumpectomy to bilateral lumpectomy (15 out of 97), and other (12 out of 97). Mostly, these management changes were because magnetic resonance imaging showed more extensive disease (n=29), additional cancer foci (n=39), or contralateral disease (n=24). In five instances, upgrading was due to patient preference. In 34% (50 out of 147) of the patients, there was no change in the planned operation. Regarding 97 of the patients having altered management, in 12 the changes were considered inappropriately extensive (due to false-positive magnetic resonance imaging findings). In terms of magnetic resonance imaging detection of more extensive, multifocal, multicentric, or contralateral disease, the false-positive rate was 13% and false-negative rate 7%. Corresponding rates for sensitivity and specificity were 95% and 81%, using the final pathology as the gold standard. Preoperative magnetic resonance imaging had a clinically significant and mostly correct impact on management plans. Magnetic resonance imaging should be included as part of the preoperative investigation in patients planned for breast-conserving surgery, in whom there are doubts about the extent of the tumours based on conventional assessment.


The Pakistan National Emergency Department Surveillance Study (Pak-NEDS): Introducing a pilot surveillance.
Evidence-based decision making is essential for appropriate prioritization and service provision by healthcare systems. Despite higher demands, data needs for this practice are not met in many cases in low- and middle-income countries because of underdeveloped sources, among other reasons. Emergency departments (EDs) provide an important channel for such information because of their strategic position within healthcare systems. This paper describes the design and pilot test of a national ED based surveillance system suitable for the Pakistani context. The Pakistan National Emergency Department Surveillance Study (Pak-NEDS) was pilot tested in the emergency departments of seven major tertiary healthcare centres across the country. The Aga Khan University, Karachi, served as the coordinating centre. Key stakeholders and experts from all study institutes were involved in outlining data needs, development of the study questionnaire, and identification of appropriate surveillance mechanisms such as methods for data collection, monitoring, and quality assurance procedures. The surveillance system was operational between November 2010 and March 2011. Active surveillance was done 24 hours a day by data collectors hired and trained specifically for the study. All patients presenting to the study EDs were eligible participants. Over 270,000 cases were registered in the surveillance system over a period of four months. Coverage levels in the final month ranged from 91-100% and were highest in centres with the least volume of patients. Overall the coverage for the four months was 79% and crude operational costs were less than $0.20 per patient. Pak-NEDS is the first multi-centre ED based surveillance system successfully piloted in a sample of major EDs having some of the highest patient volumes in Pakistan. Despite the challenges identified, our pilot shows that the system is flexible and scalable, and could potentially be adapted for many other low- and middle-income settings.


Community based actions save Yellow-spotted river turtle (Podocnemis unifilis) eggs and hatchlings flooded by rapid river level rises.
The conservation and recovery of increasingly threatened tropical freshwater turtle populations depends on effective management plans and actions. Here we show that community-based actions saved Yellow-spotted river turtle (Podocnemis unifilis) eggs submerged by unseasonal flooding and ensured the release of hatchlings. We recovered 926 eggs and 65 premature hatchlings from 74 submerged nests at 16 flooded nesting areas along 75 km of waterways. The rescued eggs were transferred to a rearing center and incubated. Hatchlings emerged from eggs that had remained underwater for up to two days. Hatchlings were maintained in 250-500 L nursery tanks until yolk sac scars had closed. Healthy hatchlings were then immediately released around the original nesting areas. We released 599 hatchlings (60.4%) from 991 submerged eggs and hatchlings. Egg survival (61.7% (571/926)) was substantially less than hatchling survival (94.2% (599/636)) but within the expected range of values reported for this species. These findings suggest that Yellow-spotted river turtle eggs and embryos are resistant to short-term submersion, which could help explain the widespread distribution of this species across highly seasonal Amazonian rivers. Management plans should take the possible survival of submerged eggs into consideration as part of species conservation and recovery actions.


In vivo attenuation of simian immunodeficiency virus by disruption of a tyrosine-dependent sorting signal in the envelope glycoprotein cytoplasmic tail.
Attenuated simian immunodeficiency viruses (SIVs) have been described that produce low levels of plasma virion RNA and exhibit a reduced capacity to cause disease. These viruses are particularly useful in identifying viral determinants of pathogenesis. In the present study, we show that mutation of a highly conserved tyrosine (Tyr)-containing motif (Yxxphi) in the envelope glycoprotein (Env) cytoplasmic tail (amino acids YRPV at positions 721 to 724) can profoundly reduce the in vivo pathogenicity of SIVmac239. This domain constitutes both a potent endocytosis signal that reduces Env expression on infected cells and a sorting signal that directs Env expression to the basolateral surface of polarized cells. Rhesus macaques were inoculated with SIVmac239 control or SIVmac239 containing either a Tyr-721-to-Ile mutation (SIVmac239Y/I) or a deletion of Tyr-721 and the preceding glycine (DeltaGY). To assess the in vivo replication competence, all viruses contained a stop codon in nef that has been shown to revert during in vivo but not in vitro replication. All three control animals developed high viral loads and disease. One of two animals that received SIVmac239Y/I and two of three animals that received SIVmac239DeltaGY remained healthy for up to 140 weeks with low to undetectable plasma viral RNA levels and normal CD4(+) T-cell percentages. These animals exhibited ongoing viral replication as determined by detection of viral sequences and culturing of mutant viruses from peripheral blood mononuclear cells and persistent anti-SIV antibody titers. In one animal that received SIVmac239Y/I, the Ile reverted to a Tyr and was associated with a high plasma RNA level and disease, while one animal that received SIVmac239DeltaGY also developed a high viral load that was associated with novel and possibly compensatory mutations in the TM cytoplasmic domain. In all control and experimental animals, the nef stop codon reverted to an open reading frame within the first 2 months of inoculation, indicating that the mutant viruses had replicated well enough to repair this mutation. These findings indicate that the Yxxphi signal plays an important role in SIV pathogenesis. Moreover, because mutations in this motif may attenuate SIV through mechanisms that are distinct from those caused by mutations in nef, this Tyr-based sorting signal represents a novel target for future models of SIV and human immunodeficiency virus attenuation that could be useful in new vaccine strategies.


Characterization of echinocandin-resistant mutants of Candida albicans: genetic, biochemical, and virulence studies.
The pneumocandins are potent antifungal agents of the echinocandin class which are under development for use as broad-spectrum antimycotic therapy. One important consideration for any new therapeutic class for treating serious fungal infections is the potential for drug resistance development. In this study we have isolated and characterized four independent spontaneous Candida albicans mutants resistant to the potent semisynthetic pneumocandin L-733,560. These mutants have many of the properties of FKS1/ETG1 echinocandin-resistant mutants of Saccharomyces cerevisiae, including (i) cross-resistance to other 1,3-beta-D-glucan synthase inhibitors, such as papulacandin and echinocandins, but no change in sensitivity to other antifungal agents; (ii) in vitro glucan synthase activity that is more resistant to pneumocandins than the wild-type parent enzyme; and (iii) semidominant drug resistance in spheroplast fusion strains. The mutants were compared with C. albicans echinocandin-resistant mutants isolated by mutagenesis by L. Beckford and D. Kerridge (mutant M-2) (abstr. PS3.11, in Proceedings of the XI Congress of the International Society for Human and Animal Mycology, Montreal, Canada, 1992) and by A. Cassone, R. E. Mason, and D. Kerridge (mutant CA-2) (Sabouraudia 19:97-110, 1981). All of the strains had resistant enzyme activity in vitro. M-2 grew poorly and had low levels of enzyme activity. In contrast, CA-2 and the spontaneous mutants grew as well as the parents and had normal levels of glucan synthase activity. These results suggest that these resistant mutants may have alterations in glucan synthase. CA-2 was unable to form germ tubes, an ability retained by the spontaneous mutants. The virulence of the spontaneous mutants was unimpaired in a mouse model of disseminated candidiasis, while M-2 and CA-2 were 2 orders of magnitude less virulent than their parent strains. Significantly, mice challenged with the spontaneous mutant CAI4R1 responded therapeutically to lower levels of L-733,560 than would he predicted by the increase in in vitro susceptibility.


Predictive factors of health-related quality of life in patients with polycystic ovary syndrome: a structural equation modeling approach.
To determine association between health-related quality of life (HRQOL) and psychosexual variables in patients with polycystic ovary syndrome (PCOS). Cross-sectional study. Two private gynecology clinics. A sample of 300 women with PCOS were entered into the study and were subdivided into three groups: hyperandrogenism (HA) and PCO morphology; menstrual dysfunction and PCO morphology; menstrual dysfunction and HA and/or PCO morphology. Then each patient completed the following questionnaires: the Hospital Anxiety and Depression Scale, the Body Image Concern Inventory, the Rosenberg Self-Esteem Scale, the Short-Form Health Survey, and the Female Sexual Function Index. None. Both direct and indirect relationships among clinical symptoms, psychologic status, self-esteem, body image, and sexual function as independent predictors of HRQOL were examined with the use of structural equation modeling analysis. The clinical variables and psychologic distress had the strongest indirect relationships with HRQOL in the HA phenotype. The highest effect of PCOS symptoms on HRQOL impairment in patients with menstrual irregularities along with HA and patients with menstrual irregularities along with polycystic ovaries was exerted by clinical variables, poor perception of self-worth, negative body image, and sexual dysfunction. In patients with various phenotypes of PCOS, the effects of mediating variables on HRQOL are different. In patients with PCOS, the findings suggest that mediating factors, especially psychologic distress, self-esteem, body image, and sexual function, play an important role and should be taken into consideration and adequately treated if present.


[Analysis of proteomic spectra in serum from patients with laryngeal carcinoma by SELDI-TOF-MS technology].
To screen the tumor markers of laryngeal carcinoma and to investigate their expression in preoperative and postoperative serum. The distinct protein in serum was detected in 32 cases of laryngeal carcinoma and 38 healthy people by IMAC-Cu proteinchip array and surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS). The distinct proteins in serum were detected in 32 cases of laryngeal carcinoma preoperation and within 10 days 15 cases of laryngeal carcinoma postoperation with the same methods. The discriminatory profiling between preoperative and postoperative patients was analyzed by Biomarker Wizard software and Biomarker Pattern software. The results showed that fifteen differentially expressed proteins in serum were screened by analysis of proteomic spectra of preoperative patients and normal subjects. Seventeen kinds of protein differentially expressed in serum were screened by analysis of proteomic spectra of preoperative and postoperative patients. Six kinds of protein (2958.52, 3796.89, 5148.86, 6115.57, 4052.18, and 7770.76) were obtained for making up patterns that was able to class the preoperative-team and postoperative-team. Corresponding correct ratio were 84.38% (27/32) and 73.33% (11/15). The preliminary results suggest that classification system will provide a highly accurate and innovative approach for the early diagnosis of laryngeal carcinoma and judgement of prognosis. SELDI-TOF-MS technology is a useful tool for a high throughput screening of large-sized serum samples to discover potential biomarkers for laryngeal carcinoma.