Reduction of c-myc expression by an antisense approach under Cre/loxP switching induces apoptosis in human liver cancer cells.
c-Myc has been documented to be both a positive and a negative signal for the induction of apoptosis. It is well known that overexpression of the c-myc gene induces apoptosis of normal cells, but the result of a reduction in its expression is not fully understood. We examined whether a reduction in c-myc expression would induce apoptosis in human liver cancer cells. Specifically, antisense and sense oligodeoxynucleotides (oligos) against the human c-myc mRNA were synthesized, mixed with a liposome reagent at various ratios, and were applied to the liver cancer-derived cell lines, HCC-T, HepG2, and PLC/PRF/5. To exclude effects resulting from using oligos, plasmid vectors expressing the full-length c-myc cDNA in both sense and antisense orientations under the control of the Cre/loxP system were generated. Monoclonal cell lines including these plasmid vectors were produced and Cre was supplied by adenovirus infection. Apoptosis was determined morphologically and c-Myc and Bcl-2 expression was examined by Western blotting. The antisense myc significantly inhibited the proliferation of the cells within two days, while neither the liposome reagent alone nor sense myc did so. Most of the cells were rounded up by the antisense-treatment and nuclear fragmentation and DNA ladder formation were detected after two days in antisense c-myc-treated cells. Antisense c-myc largely reduced c-Myc and partially Bcl-2 expression; overexpression of Bcl-2 partially rescued from apoptosis in HCC-T and HepG2 cells. These results suggest that the massive reduction in c-myc mRNA induces apoptosis in liver cancer cell lines and consequent decrease in Bcl-2 enhances the cell death. c-Myc reduction under the Cre/loxP switching system may be a useful tool for the clarification of c-myc-related cellular mechanisms in differentiation and proliferation.


Sphaerostilbellins, New Antimicrobial Aminolipopeptide Peptaibiotics from Sphaerostilbella toxica.
Sphaerostilbella toxica is a mycoparasitic fungus that can be found parasitizing wood-decay basidiomycetes in the southern USA. Organic solvent extracts of fermented strains of S. toxica exhibited potent antimicrobial activity, including potent growth inhibition of human pathogenic yeasts Candida albicans and Cryptococcus neoformans, the respiratory pathogenic fungus Aspergillus fumigatus, and the Gram-positive bacterium Staphylococcus aureus. Bioassay-guided separations led to the purification and structure elucidation of new peptaibiotics designated as sphaerostilbellins A and B. Their structures were established mainly by analysis of NMR and HRMS data, verification of amino acid composition by Marfey's method, and by comparison with published data of known compounds. They incorporate intriguing structural features, including an N-terminal 2-methyl-3-oxo-tetradecanoyl (MOTDA) residue and a C-terminal putrescine residue. The minimal inhibitory concentrations for sphaerostilbellins A and B were measured as 2 μM each for C. neoformans, 1 μM each for A. fumigatus, and 4 and 2 μM, respectively, for C. albicans. Murine macrophage cells were unaffected at these concentrations.


Effects of red-cell storage duration on patients undergoing cardiac surgery.
Some observational studies have reported that transfusion of red-cell units that have been stored for more than 2 to 3 weeks is associated with serious, even fatal, adverse events. Patients undergoing cardiac surgery may be especially vulnerable to the adverse effects of transfusion. We conducted a randomized trial at multiple sites from 2010 to 2014. Participants 12 years of age or older who were undergoing complex cardiac surgery and were likely to undergo transfusion of red cells were randomly assigned to receive leukocyte-reduced red cells stored for 10 days or less (shorter-term storage group) or for 21 days or more (longer-term storage group) for all intraoperative and postoperative transfusions. The primary outcome was the change in Multiple Organ Dysfunction Score (MODS; range, 0 to 24, with higher scores indicating more severe organ dysfunction) from the preoperative score to the highest composite score through day 7 or the time of death or discharge. The median storage time of red-cell units provided to the 1098 participants who received red-cell transfusion was 7 days in the shorter-term storage group and 28 days in the longer-term storage group. The mean change in MODS was an increase of 8.5 and 8.7 points, respectively (95% confidence interval for the difference, -0.6 to 0.3; P=0.44). The 7-day mortality was 2.8% in the shorter-term storage group and 2.0% in the longer-term storage group (P=0.43); 28-day mortality was 4.4% and 5.3%, respectively (P=0.57). Adverse events did not differ significantly between groups except that hyperbilirubinemia was more common in the longer-term storage group. The duration of red-cell storage was not associated with significant differences in the change in MODS. We did not find that the transfusion of red cells stored for 10 days or less was superior to the transfusion of red cells stored for 21 days or more among patients 12 years of age or older who were undergoing complex cardiac surgery. (Funded by the National Heart, Lung, and Blood Institute; RECESS ClinicalTrials.gov number, NCT00991341.).


Optical mosaic gratings made by consecutive, phase-interlocked, holographic exposures using diffraction from latent fringes.
We propose and demonstrate a method for fabricating large optical mosaic gratings, i.e., gratings made by consecutive, phase-interlocked, holographic exposures on single substrates. It takes advantage of the latent fringes generated by the preceding exposure to align the substrate for the following exposure. Good angular alignment accuracy and period consistency between the fringes recorded by consecutive exposures can be achieved by using the large-area moiré pattern produced by the latent fringes and the alignment beams. Phase-interlocking accuracy of less than 4 degrees between consecutive exposures was experimentally achieved by using the heterodyne detection technique.


A survey of error-correction methods for next-generation sequencing.
Error Correction is important for most next-generation sequencing applications because highly accurate sequenced reads will likely lead to higher quality results. Many techniques for error correction of sequencing data from next-gen platforms have been developed in the recent years. However, compared with the fast development of sequencing technologies, there is a lack of standardized evaluation procedure for different error-correction methods, making it difficult to assess their relative merits and demerits. In this article, we provide a comprehensive review of many error-correction methods, and establish a common set of benchmark data and evaluation criteria to provide a comparative assessment. We present experimental results on quality, run-time, memory usage and scalability of several error-correction methods. Apart from providing explicit recommendations useful to practitioners, the review serves to identify the current state of the art and promising directions for future research. All error-correction programs used in this article are downloaded from hosting websites. The evaluation tool kit is publicly available at: http://aluru-sun.ece.iastate.edu/doku.php?id=ecr.


Site-specific occurrence of nonmelanoma skin cancers in patients with cutaneous melanoma.
In a registry-based case-control study, we compared the site-specific occurrence of nonmelanoma (keratinocytic) skin cancers among patients with cutaneous melanoma cases (cases, n = 3774) and solid tumours (controls, n = 349,923), respectively. Overall, patients with melanoma were almost five-fold more likely to develop keratinocytic cancers compared with solid tumour controls (adjusted OR 4.7, 95% CI 4.1-5.3), but the risks varied depending upon the site of melanoma. Whereas patients with melanoma of the head and neck had similarly increased risks of keratinocytic cancers across all body sites, patients with melanoma of the trunk were significantly more likely to develop keratinocyte cancer diagnosed on the trunk (adjusted OR 12.5, 95% CI 7.2-20.2) than on the head and neck (adjusted OR 3.0, 95% CI 2.2-4.3). Similar colocalisation of skin tumours was observed for patients with melanomas of the lower limb. These findings provide support for the hypothesis that skin cancers at different anatomical sites may arise through different causal pathways.


Effect of cotrimoxazole on mortality in HIV-infected adults on antiretroviral therapy: a systematic review and meta-analysis.
To determine whether cotrimoxazole reduces mortality in adults receiving antiretroviral therapy (ART) for human immunodeficiency virus (HIV) infection in low- and middle-income countries through a systematic review and meta-analysis. PubMed and Embase were searched for randomized controlled trials and prospective and retrospective cohort studies that compared mortality or morbidity in HIV-infected individuals aged ≥ 13 years on cotrimoxazole and ART and on ART alone. The Newcastle-Ottawa Quality Assessment Scale was used to assess selection, confounding and measurement bias. Publication bias was assessed using Egger's and Begg's tests. Sensitivity analysis was performed because the I-squared statistic indicated substantial heterogeneity in study results. A random-effects model was used for meta-analysis. Nine studies were included. Begg and Egger P-values for the seven that reported the effect of cotrimoxazole on mortality were 0.29 and 0.49, respectively, suggesting no publication bias. The I-squared statistic was 93.2%, indicating high heterogeneity in study results. The sensitivity analysis showed that neither the follow-up duration nor the percentage of individuals with World Health Organization stage 3 or 4 HIV disease at baseline explained the heterogeneity. The summary estimate of the effect of cotrimoxazole on the incidence rate of death was 0.42 (95% confidence interval: 0.29-0.61). Since most studies followed participants for less than 1 year, it was not possible to determine whether cotrimoxazole can be stopped safely after ART-induced immune reconstitution. Cotrimoxazole significantly increased survival in HIV-infected adults on ART. Further research is needed to determine the optimum duration of cotrimoxazole treatment in these patients.


Associative fear learning and perceptual discrimination: a perceptual pathway in the development of chronic pain.
Recent neuropsychological theories emphasize the influence of maladaptive learning and memory processes on pain perception. However, the precise relationship between these processes as well as the underlying mechanisms remain poorly understood; especially the role of perceptual discrimination and its modulation by associative fear learning has received little attention so far. Experimental work with exteroceptive stimuli consistently points to effects of fear learning on perceptual discrimination acuity. In addition, clinical observations have revealed that in individuals with chronic pain perceptual discrimination is impaired, and that tactile discrimination training reduces pain. Based on these findings, we present a theoretical model of which the central tenet is that associative fear learning contributes to the development of chronic pain through impaired interoceptive and proprioceptive discrimination acuity.


Statistical inference for stochastic simulation models--theory and application.
Statistical models are the traditional choice to test scientific theories when observations, processes or boundary conditions are subject to stochasticity. Many important systems in ecology and biology, however, are difficult to capture with statistical models. Stochastic simulation models offer an alternative, but they were hitherto associated with a major disadvantage: their likelihood functions can usually not be calculated explicitly, and thus it is difficult to couple them to well-established statistical theory such as maximum likelihood and Bayesian statistics. A number of new methods, among them Approximate Bayesian Computing and Pattern-Oriented Modelling, bypass this limitation. These methods share three main principles: aggregation of simulated and observed data via summary statistics, likelihood approximation based on the summary statistics, and efficient sampling. We discuss principles as well as advantages and caveats of these methods, and demonstrate their potential for integrating stochastic simulation models into a unified framework for statistical modelling.


"Compression-only" behavior: a second-order nonlinear response of ultrasound contrast agent microbubbles.
Oscillating phospholipid-coated ultrasound contrast agent microbubbles display a so-called "compression-only" behavior, where it is observed that the bubbles compress efficiently while their expansion is suppressed. Here, a theoretical understanding of the source of this nonlinear behavior is provided through a weakly nonlinear analysis of the shell buckling model proposed by Marmottant et al. [J. Acoust. Soc. Am. 118, 3499-3505 (2005)]. It is shown that the radial dynamics of the bubble can be considered as a superposition of a linear response at the fundamental driving frequency and a second-order nonlinear low-frequency response that describes the negative offset of the mean bubble radius. The analytical solution deduced from the weakly nonlinear analysis shows that the compression-only behavior results from a rapid change of the shell elasticity with bubble radius. In addition, the radial dynamics of single phospholipid-coated microbubbles was recorded as a function of both the amplitude and the frequency of the driving pressure pulse. The comparison between the experimental data and the theory shows that the magnitude of compression-only behavior is mainly determined by the initial phospholipids concentration on the bubble surface, which slightly varies from bubble to bubble.


Role of the snake venom toxin jararhagin in proinflammatory pathogenesis: in vitro and in vivo gene expression analysis of the effects of the toxin.
To assess the indirect effects of snake venom metalloproteinases (SVMP) on host tissue local necrosis, we investigated the effect of the SVMP jararhagin on the gene expression profiles of human fibroblasts in vitro and mouse tissue in vivo. Two functional classes of up-regulated proteins, cell death and inflammatory disease were identified as being significantly populated. The changes in gene expression observed by qRT-PCR on laser microdissected mouse muscle tissue treated with jararhagin were similar with significant up-regulation of proinflammatory transcripts such as IL-1 beta, IL-6, CXCL1, CXCL2, IL-8, and apoptosis, inflammation responsive transcripts such as TNF-alpha induced protein 6. Proteolytically inactive jararhagin had no effect on the gene expression profile of fibroblasts, indicating proteolysis as the primary mechanism affecting gene expression of cells and tissues resulting in a proinflammatory, pro-apoptotic host response which likely exacerbates the local necrosis frequently observed at the site of envenoming.


Effect of metformin exposure on growth and photosynthetic performance in the unicellular freshwater chlorophyte, Chlorella vulgaris.
Many pharmaceuticals have negative effects on biota when released into the environment. For example, recent work has shown that the commonly prescribed antidiabetic drug, metformin (N,N-dimethylbiguanide), has endocrine disrupting effects on fish. However, effects of metformin on aquatic primary producers are poorly known. We exposed cultured isolates of a freshwater chlorophyte, Chlorella vulgaris, to a range of metformin concentrations (0-767.9 mg L-1) to test the hypothesis that exposure negatively affects photosynthesis and growth. A cessation of growth, increase in non-photochemical quenching (NPQ, NPQmax), and reduced electron transport rate (ETR) were observed 24 h after exposure to a metformin concentration of 767.8 mg L-1 (4.6 mM). By 48 h, photosynthetic efficiency of photosystem II (Fv/Fm), α, the initial slope of the ETR-irradiance curve, and Ek (minimum irradiance required to saturate photosynthesis) were reduced. At a lower concentration (76.8 mg L-1), negative effects on photosynthesis (increase in NPQ, decrease in ETR) were delayed, occurring between 72 and 96 h. No negative effects on photosynthesis were observed at an exposure concentration of 1.5 mg L-1. It is likely that metformin impairs photosynthesis either through downstream effects from inhibition of complex I of the electron transport chain or via activation of the enzyme, SnRK1 (sucrose non-fermenting-related kinase 1), which acts as a cellular energy regulator in plants and algae and is an ortholog of the mammalian target of metformin, AMPK (5' adenosine monophosphate-activated protein kinase).


Raman-based distributed temperature sensor with 1 m spatial resolution over 26 km SMF using low-repetition-rate cyclic pulse coding.
We experimentally investigate the benefits of a new optical pulse coding technique for long-range, meter and submeter scale Raman-based distributed temperature sensing on standard single-mode optical fibers. The proposed scheme combines a low-repetition-rate quasi-periodic pulse coding technique with the use of standard high-power fiber lasers operating at 1550 nm, allowing for what we believe is the first long-range distributed temperature measurement over single-mode fibers (SMFs). We have achieved 1 m spatial resolution over 26 km of SMF, attaining 3°C temperature resolution within 30 s measurement time.


Incentives for preventing smoking in children and adolescents.
Adult smoking usually has its roots in adolescence. If individuals do not take up smoking during this period it is unlikely that they ever will. Further, once smoking becomes established, cessation is challenging; the probability of subsequently quitting is inversely proportional to the age of initiation. One novel approach to reducing the prevalence of youth smoking is the use of incentives. To determine whether incentives prevent children and adolescents from starting to smoke. We also attempted to assess the dose-response of incentives, the costs of incentive programmes, whether incentives are more or less effective in combination with other interventions to prevent smoking initiation and any unintended consequences arising from the use of incentives. We searched the Cochrane Tobacco Addiction Group Specialized Register, with additional searches of MEDLINE, EMBASE, CINAHL, CSA databases and PsycINFO for terms relating to incentives, in combination with terms for smoking and tobacco use, and children and adolescents. The most recent searches were in May 2012. We considered randomized controlled trials allocating children and adolescents (aged 5 to 18 years) as individuals, groups or communities to intervention or control conditions, where the intervention included an incentive aimed at preventing smoking uptake. We also considered controlled trials with baseline measures and post-intervention outcomes. Data were extracted by two authors and assessed independently. The primary outcome was the smoking status of children or adolescents at follow-up who reported no smoking at baseline. We required a minimum follow-up of six months from baseline and assessed each included study for risk of bias. We used the most rigorous definition of abstinence in each trial; we did not require biochemical validation of self-reported tobacco use for study inclusion. Where possible we combined eligible studies to calculate pooled estimates at the longest follow-up using the Mantel-Haenszel fixed-effect method, grouping studies by study design. We identified seven controlled studies that met our inclusion criteria, including participants with an age range of 11 to 14 years. Of the seven trials identified, only five had analysable data relevant for this review and contributed to the meta-analysis (6362 participants in total who were non-smokers at baseline; 3466 in intervention and 2896 in control). All bar one of the studies was a trial of the so-called Smokefree Class Competition (SFC), which has been widely implemented throughout Europe. In this competition, classes with youth generally between the ages of 11 to 14 years commit to being smoke free for a six month period. They report regularly on their smoking status; if 90% or more of the class is non-smoking at the end of the six months, the class goes into a competition to win prizes. The one study that was not a trial of the SFC was a controlled trial in which schools in two communities were assigned to the intervention, with schools in a third community acting as controls. Students in the intervention community with lower smoking rates at the end of the project (one school year) received rewards.Only one study of the SFC competition, a non-randomized controlled trial, reported a significant effect of the competition on the prevention of smoking at the longest follow-up. However, this study had a risk of multiple biases, and when we calculated the adjusted RR we no longer detected a statistically significant difference. The pooled RR for the more robust RCTs (3 studies, n = 3056 participants) suggests that, from the available data, there is no statistically significant effect of incentives to prevent smoking initiation among children and adolescents in the long term (RR 1.00, 95% CI 0.84 to 1.19). Pooled results from non-randomized trials also did not detect a significant effect, and we were unable to extract data on our outcome of interest for the one trial that did not study the SFC. There is little robust evidence to suggest that unintended consequences (such as youth making false claims about their smoking status and bullying of smoking students) are consistently associated with such interventions, although this has not been the focus of much research. There was insufficient information to assess the dose-response relationship or to report costs. To date, incentive programmes have not been shown to prevent smoking initiation among youth, although there are relatively few published studies and these are of variable quality. Trials included in this meta-analysis were all studies of the SFC competition, which distributed small to moderately sized prizes to whole classes, usually through a lottery system.Future studies might investigate the efficacy of incentives given to individual participants to prevent smoking uptake. Future research should consider the efficacy of incentives on smoking initiation, as well as progression of smoking, evaluate these in varying populations from different socioeconomic and ethnic backgrounds, and describe the intervention components in detail.


[Emission of aldehydes from light duty diesel vehicles].
To characterize acetaldehyde and formaldehyde emissions, which are harmful gases to human health and not yet regulated for diesel engines. Standardized tests were performed in four diesel light duty commercial vehicles, using a frame dynamometer and test procedure FTP-75. The pollutants were analyzed by high performance liquid chromatography. Results have shown acetaldehyde emission ranged from 5.9 to 45.4 mg/km, and formaldehyde emission from 16.5 to 115.2 mg/km. The average emission for aldehyde sum was 58.7 mg/km, ranging from 22.4 to 160.6 mg/km. The proportion between the two substances remained constant, close to 74% for formaldehydes and 26% for acetaldehydes. The emission of diesel vehicle aldehydes was significant when compared with actual spark ignition vehicle emissions, or with the foresee limit for Otto cycle vehicles in Brazilian legislation. Establishing emission limits for these substances also in diesel vehicles is imperative in the light of the vehicle fleet growth, toxicity of these compounds, and their contribution as precursors in ozone gas formation reactions in low troposphere.


IRE1α activation protects mice against acetaminophen-induced hepatotoxicity.
The mammalian stress sensor IRE1α plays a central role in the unfolded protein, or endoplasmic reticulum (ER), stress response by activating its downstream transcription factor XBP1 via an unconventional splicing mechanism. IRE1α can also induce the degradation of a subset of mRNAs in a process termed regulated IRE1-dependent decay (RIDD). Although diverse mRNA species can be degraded by IRE1α in vitro, the pathophysiological functions of RIDD are only beginning to be explored. Acetaminophen (APAP) overdose is the most frequent cause of acute liver failure in young adults in the United States and is primarily caused by CYP1A2-, CYP2E1-, and CYP3A4-driven conversion of APAP into hepatotoxic metabolites. We demonstrate here that genetic ablation of XBP1 results in constitutive IRE1α activation in the liver, leading to RIDD of Cyp1a2 and Cyp2e1 mRNAs, reduced JNK activation, and protection of mice from APAP-induced hepatotoxicity. A pharmacological ER stress inducer that activated IRE1α suppressed the expression of Cyp1a2 and Cyp2e1 in WT, but not IRE1α-deficient mouse liver, indicating the essential role of IRE1α in the down-regulation of these mRNAs upon ER stress. Our study reveals an unexpected function of RIDD in drug metabolism.


Caffeine Controls Glutamatergic Synaptic Transmission and Pyramidal Neuron Excitability in Human Neocortex.
Caffeine is the most widely used psychoactive drug, bolstering attention and normalizing mood and cognition, all functions involving cerebral cortical circuits. Whereas studies in rodents showed that caffeine acts through the antagonism of inhibitory A1 adenosine receptors (A1R), neither the role of A1R nor the impact of caffeine on human cortical neurons is known. We here provide the first characterization of the impact of realistic concentrations of caffeine experienced by moderate coffee drinkers (50 μM) on excitability of pyramidal neurons and excitatory synaptic transmission in the human temporal cortex. Moderate concentrations of caffeine disinhibited several of the inhibitory A1R-mediated effects of adenosine, similar to previous observations in the rodent brain. Thus, caffeine restored the adenosine-induced decrease of both intrinsic membrane excitability and excitatory synaptic transmission in the human pyramidal neurons through antagonism of post-synaptic A1R. Indeed, the A1R-mediated effects of endogenous adenosine were more efficient to inhibit synaptic transmission than neuronal excitability. This was associated with a distinct affinity of caffeine for synaptic versus extra-synaptic human cortical A1R, probably resulting from a different molecular organization of A1R in human cortical synapses. These findings constitute the first neurophysiological description of the impact of caffeine on pyramidal neuron excitability and excitatory synaptic transmission in the human temporal cortex, providing adequate ground for the effects of caffeine on cognition in humans.


Signal transducing molecules and glycosyl-phosphatidylinositol-linked proteins form a caveolin-rich insoluble complex in MDCK cells.
GPI-linked protein molecules become Triton-insoluble during polarized sorting to the apical cell surface of epithelial cells. These insoluble complexes, enriched in cholesterol, glycolipids, and GPI-linked proteins, have been isolated by flotation on sucrose density gradients and are thought to contain the putative GPI-sorting machinery. As the cellular origin and molecular protein components of this complex remain unknown, we have begun to characterize these low-density insoluble complexes isolated from MDCK cells. We find that these complexes, which represent 0.4-0.8% of the plasma membrane, ultrastructurally resemble caveolae and are over 150-fold enriched in a model GPI-anchored protein and caveolin, a caveolar marker protein. However, they exclude many other plasma membrane associated molecules and organelle-specific marker enzymes, suggesting that they represent microdomains of the plasma membrane. In addition to caveolin, these insoluble complexes contain a subset of hydrophobic plasma membrane proteins and cytoplasmically-oriented signaling molecules, including: (a) GTP-binding proteins--both small and heterotrimeric; (b) annex II--an apical calcium-regulated phospholipid binding protein with a demonstrated role in exocytic fusion events; (c) c-Yes--an apically localized member of the Src family of non-receptor type protein-tyrosine kinases; and (d) an unidentified serine-kinase activity. As we demonstrate that caveolin is both a transmembrane molecule and a major phospho-acceptor component of these complexes, we propose that caveolin could function as a transmembrane adaptor molecule that couples luminal GPI-linked proteins with cytoplasmically oriented signaling molecules during GPI-membrane trafficking or GPI-mediated signal transduction events. In addition, our results have implications for understanding v-Src transformation and the actions of cholera and pertussis toxins on hetero-trimeric G proteins.


Pulsed-field gel electrophoresis applied for comparing Listeria monocytogenes strains involved in outbreaks.
Recent food-borne outbreaks of human listeriosis as well as numerous sporadic cases have been mainly caused by Listeria monocytogenes serovar 4b strains. Thus, it was of interest to find out whether a certain clone or a certain few clones were responsible for these cases and especially for outbreaks. We used pulsed-field gel electrophoresis of large chromosomal DNA restriction fragments generated by ApaI, SmaI, or NotI to analyse 75 L. monocytogenes strains isolated during six major and eight smaller recent listeriosis outbreaks. These strains could be divided into 20 different genomic varieties. Thirteen of 14 strains isolated during major epidemics in Switzerland (1983-1987), the United States (California, 1985) and Denmark (1985-1987) demonstrated indistinguishable DNA restriction patterns. In contrast, strains responsible for the outbreaks in Canada (Nova Scotia, 1981), the United States (Massachusetts, 1983), France (Anjou, 1975-1976), New Zealand (1969), and Austria (1986) and some smaller outbreaks in France (1987, 1988, 1989) were each characterized by particular combinations of DNA restriction patterns. Seventy-seven percent of the tested strains could be classified into the previously described ApaI group A (Brosch et al. 1991), demonstrating a very close genomic relatedness. Because 49% of the epidemic strains selected for this study belonged to phagovar 2389/2425/3274/2671/47/108/340 or 2389/47/108/340, fifty-six additional strains of these phagovars, isolated from various origins, were also typed to determine whether differences in DNA restriction profiles between epidemic and randomly selected strains of the same phagovars could be pointed out. Variations in DNA patterns appeared more frequently within randomly selected strains than within epidemic strains.


Rearrangements and aberrant expression of the retinoic acid receptor alpha gene in acute promyelocytic leukemias.
Although acute promyelocytic leukemias (APLs) are consistently associated with a reciprocal chromosome 15;17 translocation, the gene(s) directly affected by the breakpoints have never been isolated. The chromosome 17 breakpoint maps to near the retinoic acid receptor alpha (RAR alpha) locus. Investigation of 20 APLs and a large series of other neoplastic patients and normal controls revealed RAR alpha gene rearrangements and aberrant transcripts only in the APL cases. These findings suggest that the RAR alpha gene is involved in the APL chromosome 17 breakpoint, is implicated in leukemogenesis, and could be used as a marker for identifying leukemic promyelocytes.


Structural and functional protection of photoreceptors from MNU-induced retinal degeneration by the X-linked inhibitor of apoptosis.
To evaluate the neuroprotective effects of adenoassociated virus delivery of XIAP in N-methyl-N-nitrosourea (MNU)-induced retinal degeneration in Sprague-Dawley rats. Sprague-Dawley rats were injected subretinally with recombinant adenoassociated virus (rAAV) encoding either XIAP or green fluorescent protein (GFP; injection control). Six weeks after injection, the animals received an intraperitoneal injection of MNU, a DNA methylating agent, at a dose of 60 mg/kg. Electroretinograms (ERGs) were recorded at 0, 24, 48 and 72 hours and 1 week after MNU. The rats were killed after the ERG was performed and were perfused with 4% paraformaldehyde. Eyes were then enucleated and embedded for cryosectioning. Eye sections were analyzed by TUNEL and histologic techniques. Real-time PCR and Western analysis were performed to confirm the overexpression of XIAP in injected eyes. Real-time PCR and Western analysis confirmed the overexpression of XIAP in virus-injected eyes in comparison to uninjected control eyes. At 24 hours after MNU injection, fewer cells had undergone apoptosis in the XIAP-treated eyes in comparison with GFP-injected or uninjected eyes. Hematoxylin and eosin staining revealed that the uninjected and GFP-injected photoreceptors were destroyed by 72 hours after injection of MNU, whereas the AAV-XIAP-injected eyes showed structural protection of the photoreceptors at all time points throughout the 1-week sampling period. ERGs showed functional protection up to 1 week after MNU injection in the AAV-XIAP-injected eye, whereas no response was observed in the control eye. The results suggest that XIAP is protective against this potent chemotoxic agent and holds promise as a therapeutic agent in gene therapy approaches to treating retinitis pigmentosa.


Radiological and chemical assessment of phosphate rocks in some countries.
In this study, the radiological, structural and chemical characterizations of Mardin-Mazidaği phosphate rock, which is an important phosphate fertilizer source in Turkey were investigated and compared to those of several different phosphate rocks of Tunisia, Egypt, Morocco, Algeria and Syria using gamma spectrometry, X-ray diffraction (XRD) and X-ray fluorescence (XRF) measurement techniques. Elemental analysis results of phosphate samples showed that they were mainly composed of CaO, P(2)O(5), SiO(2), Al(2)O(3), SO(3) and Fe(2)O(3). Elemental concentrations of U and Th were calculated using (226)Ra and (232)Th activity concentrations, respectively. As a result of XRD analysis, the main peaks of the samples were found to be Fluorapatite (Ca(5)(PO(4))(3)F). The radioactivity concentration levels for (226)Ra, (232)Th and (40)K in all phosphate samples ranged from 250 to 1029 Bq kg(-1) with a mean of 535 Bq kg(-1), from 5 to 50 Bq kg(-1) with a mean of 20 Bq kg(-1) and from 117 to 186 Bq kg(-1) with a mean of 148 Bq kg(-1), respectively. The computed values of annual effective doses ranged from 0.17 to 0.59 mSv, with a mean value of 0.33 mSv, which is lower than the recommended limit of 1 mSv y(-1) by the International Commission on Radiological Protection.


Quality of life in swallowing in healthy elderly.
To understand the self-perception of healthy elderly regarding possible swallowing difficulties. Participants were 104 seniors, 62 women and 42 men, ranging in age from 60 to 88 years (mean 70.6), who reported good general health, no history of degenerative diseases, neurological or any other diseases that could influence swallowing participated. All subjects underwent cognitive and emotional screening and presented satisfactory conditions for both. A questionnaire to gather general and health information, and the protocol SWAL-QOL were applied. The data were statistically analyzed. Most domains of the SWAL-QOL showed mean numbers close to the maximum score (100 points), indicating a positive perception of the elderly about quality of life related to swallowing. Individuals with dentures showed better rates in most domains when compared to non-users. Individuals who self-reported ill-fitting dentures had lower scores for the domains "burden", "eating duration" and "frequency of symptoms". Men and women differed only in the domains "sleep" and "fatigue", for which women had lower mean scores. There was no correlation between the SWAL-QOL scores obtained and the variables age and monthly income. Healthy elderly, in general, do not self-report significant changes in the quality of life related to swallowing. Moreover, there is no reduction of quality of life related to swallowing as age advances. Well-adjusted dentures minimize impairment related to swallowing.


Molecular heterogeneity of Krabbe disease.
Krabbe disease (globoid cell leukodystrophy) is an autosomal recessive neurodegenerative disorder that affects both the central and peripheral nervous system due to an enzymatic defect of galactocerebrosidase (GALC). Following its cloning, many mutations in the galactocerebrosidase gene have been reported, but the correlation between phenotype and genotype was not clear in many cases. In this study we further investigated the molecular defects in another 10 patients (6 Japanese and 4 non-Japanese), using cultured skin fibroblasts, and found 10 mutations, of which 8 were novel, including a nonsense mutation (W647X) and 7 missense mutations (G43R, S52F, T262I, Y319C. W410G, R515H, T652R) in the coding region. Some phenotype-specific mutations were found but the other mutations were private. Mutations reported so far have been distributed over the whole GALC gene and it is difficult to speculate on functional domains of the GALC protein and phenotypically specific regions.


Costs of outpatient parenteral antimicrobial therapy (OPAT) administered by Hospital at Home units in Spain.
The aim of this study was to assess the direct healthcare costs of outpatient parenteral antimicrobial therapy (OPAT) administered by Hospital at Home (HaH) units in Spain. An observational, multicentre, economic evaluation of retrospective cohorts was conducted. Patients were treated at home by the HaH units of three Spanish hospitals between January 2012 and December 2013. From the cost accounting of HaH OPAT (staff, pharmacy, transportation, diagnostic tests and structural), the cost of each outpatient course was obtained following a top-down strategy based on the use of resources. Costs associated with inpatient stay, if any, were estimated based on length of stay and ICD-9-CM diagnosis. There were 1324 HaH episodes in 1190 patients (median age 70 years). The median (interquartile range) stay at home was 10 days (7-15 days). Of the OPAT episodes, 91.5% resulted in cure or improvement on completion of intravenous therapy. The mean total cost of each infectious episode was €6707 [95% confidence interval (CI) €6189-7406]. The mean cost per OPAT episode was €1356 (95% CI €1247-1560), mainly distributed between healthcare staff costs (46%) and pharmacy costs (39%). The mean cost of inpatient hospitalisation of an infectious episode was €4357 (95% CI €3947-4977). The cost per day of inpatient hospitalisation was €519, whilst the cost per day of OPAT was €98, meaning a saving of 81%. This study shows that OPAT administered by HaH units resulted in lower costs compared with inpatient care in Spain.


Achieving Accurate Sequence and Annotation Data for Caulobacter vibrioides CB13.
Annotated sequence data are instrumental in nearly all realms of biology. However, the advent of next-generation sequencing has rapidly facilitated an imbalance between accurate sequence data and accurate annotation data. To increase the annotation accuracy of the Caulobacter vibrioides CB13b1a (CB13) genome, we compared the PGAP and RAST annotations of the CB13 genome. A total of 64 unique genes were identified in the PGAP annotation that were either completely or partially absent in the RAST annotation, and a total of 16 genes were identified in the RAST annotation that were not included in the PGAP annotation. Moreover, PGAP identified 73 frameshifted genes and 22 genes with an internal stop. In contrast, RAST annotated the larger segment of these frameshifted genes without indicating a change in reading frame may have occurred. The RAST annotation did not include any genes with internal stop codons, since it chose start codons that were after the internal stop. To confirm the discrepancies between the two annotations and verify the accuracy of the CB13 genome sequence data, we re-sequenced and re-annotated the entire genome and obtained an identical sequence, except in a small number of homopolymer regions. A genome sequence comparison between the two versions allowed us to determine the correct number of bases in each homopolymer region, which eliminated frameshifts for 31 genes annotated as frameshifted genes and removed 24 pseudogenes from the PGAP annotation. Both annotation systems correctly identified genes that were missed by the other system. In addition, PGAP identified conserved gene fragments that represented the beginning of genes, but it employed no corrective method to adjust the reading frame of frameshifted genes or the start sites of genes harboring an internal stop codon. In doing so, the PGAP annotation identified a large number of pseudogenes, which may reflect evolutionary history but likely do not produce gene products. These results demonstrate that re-sequencing and annotation comparisons can be used to increase the accuracy of genomic data and the corresponding gene annotation.


Noelin1 Affects Lateral Mobility of Synaptic AMPA Receptors.
Lateral diffusion on the neuronal plasma membrane of the AMPA-type glutamate receptor (AMPAR) serves an important role in synaptic plasticity. We investigated the role of the secreted glycoprotein Noelin1 (Olfactomedin-1 or Pancortin) in AMPAR lateral mobility and its dependence on the extracellular matrix (ECM). We found that Noelin1 interacts with the AMPAR with high affinity, however, without affecting rise- and decay time and desensitization properties. Noelin1 co-localizes with synaptic and extra-synaptic AMPARs and is expressed at synapses in an activity-dependent manner. Single-particle tracking shows that Noelin1 reduces lateral mobility of both synaptic and extra-synaptic GluA1-containing receptors and affects short-term plasticity. While the ECM does not constrain the synaptic pool of AMPARs and acts only extrasynaptically, Noelin1 contributes to synaptic potentiation by limiting AMPAR mobility at synaptic sites. This is the first evidence for the role of a secreted AMPAR-interacting protein on mobility of GluA1-containing receptors and synaptic plasticity.


Antidepressants in the treatment of adult attention-deficit hyperactivity disorder: a systematic review.
Stimulant medications are the most effective drugs in the treatment of attention-deficit hyperactivity disorder (ADHD) in children and in adults. However, some patients do not respond to this treatment and other patients suffer from adverse effects. Very often there are also comorbid disorders that warrant treatment or there is somatic comorbidity that precludes the prescription of stimulants. As a result, alternative treatments for the treatment of ADHD have been explored, such as antidepressant agents. In this systematic review the evidence base for the effect of antidepressants for ADHD in adult patients is determined. Electronic and hand searches were conducted in order to identify clinical trials studying antidepressants for the treatment of ADHD in adult patients. The trials were screened for methodological characteristics and treatment-effect sizes. The odds ratio was calculated for randomized controlled trials with bupropion. A descriptive review of all the randomized controlled studies and an overview of the nonrandomized studies was developed. Only eight randomized controlled trials were retrieved with four different compounds. Five studies concerned bupropion and the meta-analysis indicates a beneficial effect for bupropion compared with placebo as measured with the Clinical Global Improvement Scale (odds ratio 2.42 [95% CI 1.09 to 5.36]). Several studies suffer from clinical and methodological shortcomings, such as exclusion of patients with comorbid disorders, short treatment duration, or a lack of information with respect to the randomization procedure. Although there is a need for alternative interventions for the treatment of ADHD, such as with antidepressant agents, the evidence base is not large. Only treatment with bupropion seems to have a medium-range effect size, but this is less than that of stimulant medications.


Biophysical properties of gap junctions between freshly dispersed pairs of mouse pancreatic beta cells.
Coupling between beta cells through gap junctions has been postulated as a principal mechanism of electrical synchronization of glucose-induced activity throughout the islet of Langerhans. We characterized junctional conductance between isolated pairs of mouse pancreatic beta cells by whole-cell recording with two independent patch-clamp circuits. Most pairs were coupled (67%, n = 155), although the mean junctional conductance (gj) (215 +/- 110 pS) was lower than reported in other tissues. Coupling could be recorded for long periods, up to 40 min. Voltage imposed across the junctional or nonjunctional membranes had no effect on gj. Up to several hours of treatment to increase intracellular cAMP levels did not affect gj. Electrically coupled pairs did not show transfer of the dye Lucifer yellow. Octanol (2 mM) reversibly decreased gj. Lower concentrations of octanol (0.5 mM) and heptanol (0.5 mM) than required to uncouple beta cells decreased voltage-dependent K+ and Ca2+ currents in nonjunctional membranes. Although gj recorded in these experiments would be expected to be provided by current flowing through only a few channels of the unitary conductance previously reported for other gap junctions, no unitary junctional currents were observed even during reversible suppression of gj by octanol. This result suggests either that the single channel conductance of gap junction channels between beta cells is smaller than in other tissues (less than 20 pS) or that the small mean conductance is due to transitions between open and closed states that are too rapid or too slow to be resolved.


Error-prone bypass of DNA lesions during lagging-strand replication is a common source of germline and cancer mutations.
Studies in experimental systems have identified a multitude of mutational mechanisms including DNA replication infidelity and DNA damage followed by inefficient repair or replicative bypass. However, the relative contributions of these mechanisms to human germline mutation remain unknown. Here, we show that error-prone damage bypass on the lagging strand plays a major role in human mutagenesis. Transcription-coupled DNA repair removes lesions on the transcribed strand; lesions on the non-transcribed strand are preferentially converted into mutations. In human polymorphism we detect a striking similarity between mutation types predominant on the non-transcribed strand and on the strand lagging during replication. Moreover, damage-induced mutations in cancers accumulate asymmetrically with respect to the direction of replication, suggesting that DNA lesions are resolved asymmetrically. We experimentally demonstrate that replication delay greatly attenuates the mutagenic effect of ultraviolet irradiation, confirming that replication converts DNA damage into mutations. We estimate that at least 10% of human mutations arise due to DNA damage.


Risks of respiratory disease in the heavy clay industry.
Little information is available on the quantitative risks of respiratory disease from quartz in airborne dust in the heavy clay industry. Available evidence suggested that these risks might be low, possibly because of the presence in the dust of other minerals, such as illite and kaolinite, which may reduce the harmful effects of quartz. The aims of the present cross sectional study were to determine among workers in the industry (a) their current and cumulative exposures to respirable mixed dust and quartz; (b) the frequencies of chest radiographic abnormalities and respiratory symptoms; (c) the relations between cumulative exposure to respirable dust and quartz, and risks of radiographic abnormality and respiratory symptoms. Factories were chosen where the type of process had changed as little as possible during recent decades. 18 were selected in England and Scotland, ranging in size from 35 to 582 employees, representing all the main types of raw material, end product, kilns, and processes in the manufacture of bricks, pipes, and tiles but excluding refractory products. Weights of respirable dust and quartz in more than 1400 personal dust samples, and site histories, were used to derive occupational groups characterised by their levels of exposure to dust and quartz. Full size chest radiographs, respiratory symptoms, smoking, and occupational history questionnaires were administered to current workers at each factory. Exposure-response relations were examined for radiographic abnormalities (dust and quartz) and respiratory symptoms (dust only). Respirable dust and quartz concentrations ranged from means of 0.4 and 0.04 mg.m-3 for non-process workers to 10.0 and 0.62 mg.m-3 for kiln demolition workers respectively. Although 97% of all quartz concentrations were below the maximum exposure limit of 0.4 mg.m-3, 10% were greater than this among the groups of workers exposed to most dust. Cumulative exposure calculations for dust and quartz took account of changes of occupational group, factory, and kiln type at study and non-study sites. Because of the importance of changes of kiln type additional weighting factors were applied to concentrations of dust and quartz during previous employment at factories that used certain types of kiln. 85% (1934 employees) of the identified workforce attended the medical surveys. The frequency of small opacities in the chest radiograph, category > or = 1/0, was 1.4% (median reading) and seven of these 25 men had category > or = 2/1. Chronic bronchitis was reported by 14.2% of the workforce and breathlessness, when walking with someone of their own age, by 4.4%. Risks of having category > or = 0/1 small opacities differed by site and were also influenced by age, smoking, and lifetime cumulative exposure to respirable dust and quartz. Although exposures to dust and to quartz were highly correlated, the evidence suggested that radiological abnormality was associated with quartz rather than dust. A doubling of cumulative quartz exposure increased the risk of having category > or = 0/1 by a factor of 1.33. Both chronic bronchitis and breathlessness were significantly related to dust exposure. Although most quartz concentrations at the time of this study were currently below regulatory limits in the heavy clay industry, high exposures regularly occurred in specific processes and occasionally among most occupational groups. However, there are small risks of pneumoconiosis and respiratory symptoms in the industry, although frequency of pneumoconiosis is low in comparison to other quartz exposed workers.


Axonal loss in multiple sclerosis: a pathological survey of the corticospinal and sensory tracts.
Clinical, imaging, and pathological studies in multiple sclerosis have generally emphasized the relative preservation of axons in comparison with myelin. Recent evidence, however, demonstrates that axonal loss is also significant, affects long tracts such as the corticospinal and sensory tracts and relates closely to functional disability. Accordingly, the distribution and extent of this axonal loss is the focus of the current investigation. Post-mortem material of 55 multiple sclerosis patients and 32 matched controls was used to examine quantitatively the population of axons in the corticospinal tracts from the medulla to the lumbar spinal cord and the sensory tracts from the lumbar to the upper cervical spinal cord. Myelin- and axon-stained sections have been prepared to estimate the notional area and axon density, respectively of both tracts. Our results indicate that in the corticospinal tracts there is a significant reduction of the area and axon density at all levels investigated in multiple sclerosis cases when compared with controls. In contrast, the sensory tracts in multiple sclerosis cases showed a significant reduction in area and axon density only in the upper regions of the spinal cord. As has been found with MRI plaque load and T2 burden, correlations of axonal loss with duration of disease were not strong. Of the fibres lost in multiple sclerosis, we have found that small fibres (<3 microm2) seem to be particularly affected, with large fibres remaining relatively preserved in both the corticospinal and sensory tracts. In multiple sclerosis, axonal loss is widespread, and its extent is tract specific and size selective.


Colors of the daytime overcast sky.
Time-series measurements of daylight (skylight plus direct sunlight) spectra beneath overcast skies reveal an unexpectedly wide gamut of pastel colors. Analyses of these spectra indicate that at visible wavelengths, overcasts are far from spectrally neutral transmitters of the daylight incident on their tops. Colorimetric analyses show that overcasts make daylight bluer and that the amount of bluing increases with cloud optical depth. Simulations using the radiative-transfer model MODTRAN4 help explain the observed bluing: multiple scattering within optically thick clouds greatly enhances spectrally selective absorption by water droplets. However, other factors affecting overcast colors seen from below range from minimal (cloud-top heights) to moot (surface colors).


Alpha 1-adrenergic and muscarinic cholinergic stimulation of phosphoinositide hydrolysis in adult rat cardiomyocytes.
Cardiac alpha-adrenergic receptors mediate cellular responses to norepinephrine through an undefined series of molecular events. We examined the possibility that phosphoinositide hydrolysis was stimulated through alpha-adrenergic receptors in cardiomyocytes purified from adult rat ventricle. Phosphoinositide stores were labeled with [3H]inositol, and [3H]inositol phosphate formation was assessed after the addition of lithium chloride and norepinephrine. Norepinephrine increased the accumulation of [3H]inositol phosphate by approximately 5-fold, giving a maximal response at approximately 30 microM and a half-maximal response at approximately 1 microM. There was a significant increase in [3H]inositol phosphate formation in response to norepinephrine at 5 minutes, and the response was linear over 40 minutes. Norepinephrine-stimulated [3H]inositol phosphate formation was not blocked by propranolol (1 microM) or yohimbine (0.1 microM) but was completely antagonized by the alpha 1-selective antagonist prazosin (0.1 microM). Muscarinic cholinergic receptor activation by carbachol also stimulated [3H]inositol phosphate formation in rat ventricular myocytes. The maximal effect of carbachol (approximately 2-fold) was always less than that of norepinephrine. The combined effects of norepinephrine and carbachol were additive, suggesting that the two hormones do not share a common rate-limiting step. Removal of extracellular calcium and addition of ethylene glycol bis(beta-amino ether)-N,N'-tetraacetic acid, attenuated, but did not abolish, norepinephrine- or carbachol-stimulated [3H]inositol phosphate formation. Neither the calcium ionophore A23187 nor the calcium channel blockers verapamil and nifedipine had any effect on basal or hormone-stimulated [3H]inositol phosphate formation. We suggest that some of the physiological and metabolic effects of adrenergic and cholinergic stimulation on the rat myocardium are secondary to receptor-mediated hydrolysis of phosphoinositides.


A Probable Case of Incipient Speciation in Schizocosa Wolf Spiders Driven by Allochrony, Habitat Use, and Female Mate Choice.
There is growing evidence that speciation can occur between populations that are not geographically isolated. The emergence of assortative mating is believed to be critical to this process, but how assortative mating arises in diverging populations is poorly understood. The wolf spider genus Schizocosa has become a model system for studying mechanisms of assortative mating. We conducted a series of experiments to identify the factors that control mate pair formation in a Schizocosa population that includes both ornamented and nonornamented males. We show that the population also includes two previously unrecognized female phenotypes. One female phenotype mates mostly or exclusively with ornamented males, and the other mates mostly or exclusively with unornamented males. Assortative mating within these groups is maintained by differences in maturation time, microhabitat use, and female mate preference. We conclude that the population is not a single species, as previously believed, but rather an incipient species pair with multiple overlapping mechanisms of reproductive isolation. The identification of a new incipient species pair in the well-studied and rapidly speciating Schizocosa clade presents new opportunities for the study of speciation without geographic isolation.


shRNA-induced saturation of the microRNA pathway in the rat brain.
RNA interference (RNAi) is a powerful strategy for unraveling gene function and for drug target validation, but exogenous expression of short hairpin RNAs (shRNAs) has been associated with severe side effects. These may be caused by saturation of the microRNA pathway. This study shows degenerative changes in cell morphology and intrusion of blood vessels after transduction of the ventromedial hypothalamus (VMH) of rats with a shRNA expressing adeno-associated viral (AAV) vector. To investigate whether saturation of the microRNA pathway has a role in the observed side effects, expression of neuronal microRNA miR-124 was used as a marker. Neurons transduced with the AAV vector carrying the shRNA displayed a decrease in miR-124 expression. The decreased expression was unrelated to shRNA sequence or target and observed as early as 1 week after injection. In conclusion, this study shows that the tissue response after AAV-directed expression of a shRNA to the VMH is likely to be caused by shRNA-induced saturation of the microRNA pathway. We recommend controlling for miR-124 expression when using RNAi as a tool for studying (loss of) gene function in the brain as phenotypic effects caused by saturation of the RNAi pathway might mask true effects of specific downregulation of the shRNA target.


Is adolescent internet use a risk factor for the development of depression symptoms or vice-versa?
The extent to which digital media use by adolescents contributes to poor mental health, or vice-versa, remains unclear. The purpose of the present study is to clarify the strength and direction of associations between adolescent internet use and the development of depression symptoms using a longitudinal modeling approach. We also examine whether associations differ for boys and girls. Data are drawn from (N = 1547) participants followed for the Quebec longitudinal Study of Child Development (QLSCD 1998-2020). Youth self-reported internet use in terms of the average hours of use per week at the ages of 13, 15, and 17. Youth also self-reported depression symptoms at the same ages. After testing sex-invariance, random intercepts cross-lagged panel models stratified by sex, revealed that internet use by girls was associated with significant within-person (time-varying) change in depression symptoms. Girl's internet use at age 13 was associated with increased depression symptoms at age 15 (ß = 0.12) and internet use at age 15 increased depression at age 17 (ß = 0.10). For boys, internet use was not associated with significant time varying change in depression symptoms. The present findings support the hypothesis that internet use by adolescents can represent a significant risk factor for the development of depressive symptoms, particularly in girls.


Cognitive-behavior therapy resolves implicit fear associations in generalized anxiety disorder.
Cognitive schema theories postulate that anxiety disorders are associated with excessive fear associations in memory. For generalized anxiety disorder (GAD), it has been shown that patients not only exhibit negative implicit evaluations of clearly negative worry words (e.g., cancer), but also a generalization of this effect to neutral words (e.g., diagnosis). This study assessed the sensitivity of this bias, which has been interpreted as an indicator of a pathologically broadened fear structure, to cognitive-behavioral therapy (CBT). An Extrinsic Affective Simon task was used to measure implicit associations with idiosyncratic neutral and negative worry words in 23 GAD patients and 25 healthy controls (HC). Patients were tested before and after CBT, and half of them were additionally tested while waiting for treatment. Clinical symptoms were measured before and after treatment, and at 6-months follow-up. CBT normalized bias for neutral words, and the extent of bias reduction during treatment predicted the extent of additional symptom improvement during the 6 months following intervention. Furthermore, the amplitude of pre-treatment bias predicted the onset of CBT response, with lower bias predicting immediate symptom improvement at the end of treatment, and higher bias predicting delayed treatment effects during the 6 months follow-up. Biased implicit evaluation of neutral worry targets does not represent an enduring vulnerability factor for the development of GAD but is related to heightened levels of state worry. Furthermore, the normalization of this bias might be a crucial factor in the therapeutic action of CBT.


Mineral absorption in the black rhinoceros (Diceros bicornis) as compared with the domestic horse.
To test whether mineral recommendations for horses are likely to guarantee adequate mineral provision for black rhinoceroses (Diceros bicornis), we investigated the apparent absorption (aA) of macro- and microminerals in eight black rhinoceroses from three zoological institutions in a total of 32 feeding trials with total faecal collection, with additional data from three unpublished studies (18 feeding trials). Feeds and faeces were analysed for Ca, P, Mg, Na, K, Fe, Mn, Cu, Zn and Co. The resulting aA coefficients, and the linear relationships of apparently absorbable dietary mineral content to total dietary mineral content [per 100 g dry matter (DM)], were compared with data for domestic horses. Rhinoceroses had significantly higher aA coefficients for Ca and Mg (because of a higher calculated 'true' absorption), and lower ones for Na and K (because of calculated higher endogenous faecal losses). High absorption efficiency for divalent cations is hypothesized to be an adaptation to a natural diet of particularly high Ca:P ratio (approximately 14:1); an effective removal of Ca from the ingesta guarantees sufficient P availability at the fermentation site in the hindgut. Higher faecal losses of Na and K are hypothesized to be linked to a higher faecal bulk per DM intake in black rhinoceroses as compared with horses because of a generally lower digestive efficiency. There were no relevant differences in the absorption patterns of microminerals. In particular, there were no discernable differences in Fe absorption within the rhinoceroses for diets with and without tannin supplementation. Several of the zoo diets assessed in this study were deficient in Cu, Mn or Zn, and most contained excessive levels of Fe when compared with horse requirements. The findings of this study indicate that differences in mineral absorption between occur even between species of similar digestive anatomy; that in particular, Ca absorption might vary between hindgut fermenters with Ca:P ratio in their natural diet; that Na might be a particularly limiting factor in the ecology of free-ranging rhinoceroses; that moderate doses of tannins do not seem to markedly influence mineral absorption; and that diets for captive animals should contain adequate, but not excessive mineral levels.


Thermoelectric current in topological insulator nanowires with impurities.
In this paper we consider charge current generated by maintaining a temperature difference over a nanowire at zero voltage bias. For topological insulator nanowires in a perpendicular magnetic field the current can change sign as the temperature of one end is increased. Here we study how this thermoelectric current sign reversal depends on the magnetic field and how impurities affect the size of the thermoelectric current. We consider both scalar and magnetic impurities and show that their influence on the current are quite similar, although the magnetic impurities seem to be more effective in reducing the effect. For moderate impurity concentration the sign reversal persists.


Impact of the ASA Physical Status Score on Adjuvant Chemotherapy Eligibility and Survival of Upper Tract Urothelial Carcinoma Patients: a Multicenter Study.
The aim of the present multi-institutional study was to assess the influence of the American Society of Anesthesiologists Physical Status (ASA-PS) classification on adjuvant chemotherapy eligibility and survival in a multi-institutional cohort of patients treated with radical nephroureterectomy (RNU) for upper tract urothelial carcinoma (UTUC). We retrospectively reviewed data from 416 patients who underwent RNU for UTUC at four Korean institutions between 2001 and 2013. The ASA-PS classification was obtained from the anesthesia chart. Locally advanced UTUC was defined as ≥ pT3 and/or pN1 disease. The influence of ASA-PS score on survival was evaluated by Kaplan-Meier analyses and a multivariate Cox regression model. Patients with a higher ASA-PS class were less likely to be eligible for adjuvant chemotherapy in locally advanced UTUC (P = 0.016). Kaplan-Meier estimates showed that the high-risk ASA-PS group has a poorer overallsurvival (OS) and cancer-specific survival (CSS) compared to low risk ASA-PS groups in both the total and locally advanced UTUC cohorts. Based on multivariate Cox regression analysis, the high-risk ASA-PS category was an independent predictor for overall mortality (OM) (hazard ratio [HR], 1.919; 95% confidence interval [CI], 1.017-3.619; P = 0.044) and cancer-specific mortality (CSM) (HR, 2.120; 95% CI, 1.023-4.394; P = 0.043). In conclusion, high-risk ASA-PS score was independently associated with a lower survival rate in patients with UTUC after RNU. However, the influence of ASA-PS classification on survival was limited to locally advanced UTUC. The lower eligibility of patients in the high-risk ASA category for adjuvant chemotherapy may contribute to the lower survival rate in this group.


Genome of Wild Mandarin and Domestication History of Mandarin.
Mandarin (Citrus reticulata) is one of the most important citrus crops worldwide. Its domestication is believed to have occurred in South China, which has been one of the centers of mandarin cultivation for four millennia. We collected natural wild populations of mandarin around the Nanling region and cultivated landraces in the vicinity. We found that the citric acid level was dramatically reduced in cultivated mandarins. To understand genetic basis of mandarin domestication, we de novo assembled a draft genome of wild mandarin and analyzed a set of 104 citrus genomes. We found that the Mangshan mandarin is a primitive type and that two independent domestication events have occurred, resulting in two groups of cultivated mandarins (MD1 and MD2) in the North and South Nanling Mountains, respectively. Two bottlenecks and two expansions of effective population size were identified for the MD1 group of cultivated mandarins. However, in the MD2 group there was a long and continuous decrease in the population size. MD1 and MD2 mandarins showed different patterns of interspecific introgression from cultivated pummelo species. We identified a region of high divergence in an aconitate hydratase (ACO) gene involved in the regulation of citrate content, which was possibly under selection during the domestication of mandarin. This study provides concrete genetic evidence for the geographical origin of extant wild mandarin populations and sheds light on the domestication and evolutionary history of mandarin.


A changing time: the International Society for Cellular Therapy embraces its industry members.
The last decade has seen a dramatic rise in the development of new cellular therapeutics in a wide range of indications. There have been acceptable safety profiles reported in early studies using blood-derived and adherent stem cell products, but also an inconsistent efficacy record. Further expansion has been hindered in part by a lack of capital (both private and public) and delayed entry into the cell therapy space by large healthcare and pharmaceutical companies, those members of the industry most reliably able to initiate and maintain advanced-phase clinical trials. With recognition that the International Society for Cellular Therapy (ISCT) is uniquely positioned to serve the global translational regenerative medicine research community as a network hub for scientific standards and policy, the ISCT commissioned the establishment of an Industry Task Force (ITF) to address current and future roles for industry. The objectives of the ITF were to gather information and prioritize efforts for a new Commercialization Committee (CC) and to construct innovative platforms that would foster constructive and synergistic collaborations between industry and ISCT. Recommendations and conclusions of the ITF included that the new CC: (1) foster new relationships with therapeutic and stem cell societies, (2) foster educational workshops and forums to cross-educate and standardize practices, (3) create industry subcommittees to address priority initiatives, with clear benchmarks and global implementation, and (4) establish a framework for a greater industry community within ISCT, opening doors for industry to share the new vision for commercialization of cell therapy, emphasizing the regenerative medicine space.


Operative outcomes beyond 30-day mortality: colorectal cancer surgery in oldest old.
Resections for elderly colorectal cancer (CRC) are forecasted to grow, particularly in those beyond the age limit of screening (>80 years). However, literature on operative outcomes after CRC procedures in the oldest old is focused primarily on operative mortality. We hypothesize that older age will additionally impact operative morbidity after CRC resections in a multihospital, risk-adjusted database. We identified 19,375 patients >40 years who underwent CRC procedures in the 2005 to 2008 American College of Surgeons National Surgical Quality Improvement Program (ACS NSQIP) database. Pre-, intra-, and postoperative factors were compared by age groups. Multivariable techniques were used to assess the effects of older age on operative outcome measures, adjusting for covariates. Over 20% of our cohort was older than 80 years. Of those, 17% developed major complications and 29% experienced prolonged length of stay (LOS). Older patients also experienced higher rates of 30-day operative mortality (>80 years vs. 45-55 years; 6% vs. <1%), major complications (>80 years vs. 45-55 years; 21% vs. 14%), and prolonged LOS after open (>80 years vs. 45-55 years; 37% vs. 24%) and laparoscopic procedures (>80 years vs. 45-55 years; 40.5% vs. 18%). These unadjusted comparisons persisted in multivariable analyses demonstrating that older age independently predicted worse operative outcomes after CRC procedures. The effects of older age extend to other important outcome measures after CRC procedures beyond operative mortality. As one of the largest multihospital studies, our study identified increased morbidity in the oldest old, a growing population. Our results should stimulate review of current policy and resource allocation.


Comparison of the Framingham Risk Score, SCORE and WHO/ISH cardiovascular risk prediction models in an Asian population.
Cardiovascular risk-prediction models are used in clinical practice to identify and treat high-risk populations, and to communicate risk effectively. We assessed the validity and utility of four cardiovascular risk-prediction models in an Asian population of a middle-income country. Data from a national population-based survey of 14,863 participants aged 40 to 65 years, with a follow-up duration of 73,277 person-years was used. The Framingham Risk Score (FRS), SCORE (Systematic COronary Risk Evaluation)-high and -low cardiovascular-risk regions and the World Health Organization/International Society of Hypertension (WHO/ISH) models were assessed. The outcome of interest was 5-year cardiovascular mortality. Discrimination was assessed for all models and calibration for the SCORE models. Cardiovascular risk factors were highly prevalent; smoking 20%, obesity 32%, hypertension 55%, diabetes mellitus 18% and hypercholesterolemia 34%. The FRS and SCORE models showed good agreement in risk stratification. The FRS, SCORE-high and -low models showed good discrimination for cardiovascular mortality, areas under the ROC curve (AUC) were 0.768, 0.774 and 0.775 respectively. The WHO/ISH model showed poor discrimination, AUC=0.613. Calibration of the SCORE-high model was graphically and statistically acceptable for men (χ(2) goodness-of-fit, p=0.097). The SCORE-low model was statistically acceptable for men (χ(2) goodness-of-fit, p=0.067). Both SCORE-models underestimated risk in women (p<0.001). The FRS and SCORE-high models, but not the WHO/ISH model can be used to identify high cardiovascular risk in the Malaysian population. The SCORE-high model predicts risk accurately in men but underestimated it in women.


Relationships between segregated afferents and postsynaptic neurones in the optic tectum of three-eyed frogs.
In 3-eyed frogs, afferents from 2 eyes converge on an optic tectum that normally receives input from only 1 eye. This produces an interdigitating series of stripes, resembling the ocular dominance columns in cats and monkeys. The consequences of this induced striping on the behavior of tectal dendrites was investigated in an in vitro preparation of the tectum. Stripes were labeled by anterograde transport of a fluorescent dye (rhodamine) and postsynaptic tectal cells labeled by intracellular injections of Lucifer yellow. The same types of cells were present in both normal and striped tecta, but dendritic arbors were altered in 2 ways. In normal tecta, dendrites were most frequently biased in a rostral direction. In striped tecta, dendrites were more frequently unbiased: fewer arbors had a strong rostral bias. The second effect of stripes was on the behaviors of individual dendrites of certain cell types. Some cells, primarily those with small, highly branched arbors, had dendrites that abruptly terminated at the borders between stripes. Other cells, with larger arbors, maintained "clumps" of dendrites in both eye's stripes. While these cells had portions of their dendritic arbor in more than one stripe, each individual dendrite was restricted to a single stripe. However, the processes of many cells, especially those with extensive, medial-laterally oriented dendrites, did not respect stripe boundaries in any obvious fashion. At the border between 2 stripes, there is an abrupt discontinuity in the patterns of activity in afferent axons. The dendritic alterations seen in striped tecta suggest that correlated activity can, in some cells, modulate the spatial arrangement of dendrites, such that an individual dendrite preferentially arborizes within such areas, but not between them. These cells as a whole can accommodate uncorrelated inputs, if these are segregated onto separate dendrites. This implies that local interactions between presynaptic terminals and postsynaptic dendrites, rather than action potentials in the postsynaptic cells, may furnish important signals for the modulation of dendritic arbor shape.


Safe administration of the measles vaccine to children allergic to eggs.
The safety of administering the combined measles-mumps-rubella (MMR) vaccine to patients who are allergic to eggs has been debated for decades because of concern about potential anaphylaxis, since the live attenuated virus used in the vaccine is grown in cultured chick-embryo fibroblasts. We recruited 54 children (mean age, 18.5 months) who had not previously been vaccinated and were allergic to eggs. The children's histories of allergy were confirmed with skin tests and double-blind, placebo-controlled food-challenge tests; some children also underwent skin testing with the MMR vaccine. We then routinely administered the vaccine to the children in one subcutaneous (0.5-ml) dose. All 54 children had positive results on skin testing with egg. Allergy to eggs was confirmed in 26 of the children by convincing histories of anaphylaxis after the ingestion of eggs, in 22 children by food-challenge tests, and in 6 patients by convincing histories of recent allergic reactions occurring after the ingestion of eggs. Of the 17 children who underwent skin testing with the MMR vaccine, 3 had positive results. All 54 children received the MMR vaccine as a single subcutaneous injection; none had an immediate or delayed adverse reaction. The MMR vaccine can be safely administered in a single dose to children with allergy to eggs, even those with severe hypersensitivity.


Genetic counseling following direct-to consumer genetic testing: Consumer perspectives.
As the use and scope of direct-to-consumer genetic testing (DTC GT), also becoming known as consumer-driven genetic testing, increases, consumers may seek genetic counseling to understand their results and determine healthcare implications. In this study, we interviewed individuals who sought genetic counseling after receiving DTC GT results to explore their motivations, expectations, and experiences. Participants were recruited from the Impact of Personal Genomics (PGen) Study, a longitudinal cohort study of DTC GT customers. We interviewed 15 participants (9 females, mean age = 38 years) by telephone and analyzed the double-coded transcripts using qualitative methods. Motivations for genetic counseling included family and personal health histories, concern and confusion about results, and information-seeking; of note, one-third of our interview participants had Ehlers-Danlos syndrome Type III (hypermobility type). Expectations of genetic counseling sessions were high. Participants generally saw DTC GT results as valid and potentially impactful for their healthcare, wanted more thorough explanations in "layman's terms," a pooling of their results with their family and personal health history and a "game plan." Several participants had already accessed online resources, including resources typically used by genetics clinicians. Our results point to several elements of a successful DTC GT genetic counseling session: 1) effective contracting when starting the clinic visit, especially determining motivations for genetic counseling, results that are concerning/confusing and resources already accessed; 2) ascertainment and management of expectations and clearly communicating if and why all results may not be reviewed; 3) explaining how DTC GT differs from clinical genetic testing and why additional testing may not be indicated and 4) listening to (not dismissing) patient concerns about their results. For those patients who seek genetic counseling about DTC GT results, the findings from our study can help inform case preparation and provision of genetic counseling.


Accurate genome-wide predictions of spatio-temporal gene expression during embryonic development.
Comprehensive information on the timing and location of gene expression is fundamental to our understanding of embryonic development and tissue formation. While high-throughput in situ hybridization projects provide invaluable information about developmental gene expression patterns for model organisms like Drosophila, the output of these experiments is primarily qualitative, and a high proportion of protein coding genes and most non-coding genes lack any annotation. Accurate data-centric predictions of spatio-temporal gene expression will therefore complement current in situ hybridization efforts. Here, we applied a machine learning approach by training models on all public gene expression and chromatin data, even from whole-organism experiments, to provide genome-wide, quantitative spatio-temporal predictions for all genes. We developed structured in silico nano-dissection, a computational approach that predicts gene expression in >200 tissue-developmental stages. The algorithm integrates expression signals from a compendium of 6,378 genome-wide expression and chromatin profiling experiments in a cell lineage-aware fashion. We systematically evaluated our performance via cross-validation and experimentally confirmed 22 new predictions for four different embryonic tissues. The model also predicts complex, multi-tissue expression and developmental regulation with high accuracy. We further show the potential of applying these genome-wide predictions to extract tissue specificity signals from non-tissue-dissected experiments, and to prioritize tissues and stages for disease modeling. This resource, together with the exploratory tools are freely available at our webserver http://find.princeton.edu, which provides a valuable tool for a range of applications, from predicting spatio-temporal expression patterns to recognizing tissue signatures from differential gene expression profiles.


Microbial L-phenylalanine ammonia-lyase. Purification, subunit structure and kinetic properties of the enzyme from Rhizoctonia solani.
1. Phenylalanine ammonia-lyase (EC 126.96.36.199) was purified to homogeneity from the acetone-dried powders of the mycelial felts of the plant pathogenic fungus Rhizoctonia solani. 2. A useful modification in protamine sulphate treatment to get substantial purification of the enzyme in a single-step is described. 3. The purified enzyme shows bisubstrate activity towards L-phenylalanine and L-tyrosine. 4. It is sensitive to carbonyl reagents and the inhibition is not reversed by gel filtration. 5. The molecular weight of the enzyme as determined by Sephadex G-200 chromatography and sucrose-density-gradient centrifugation is around 330000. 6. The enzyme is made up of two pairs of unidentical subunits, with a molecular weight of 70000 (alpha) and 90000 (beta) respectively. 7. Studies on initial velocity versus substrate concentration have shown significant deviations from Michaelis-Menten kinetics. 8. The double-reciprocal plots are biphasic (concave downwards) and Hofstee plots show a curvilinear pattern. 9. The apparent Km value increases from 0.18 mM to as high as 5.0 mM with the increase in the concentration of the substrate and during this process the Vmax, increases by 2-2.5-fold. 10. The value of Hill coefficient is 0.5. 11. Steady-state rates of phenylalanine ammonia-lyase reaction in the presence of inhibitors like D-phenylalanine, cinnamic, p-coumaric, caffeic, dihydrocaffeic and phenylpyruvic acid have shown that only one molecule of each type of inhibitor binds to a molecule of the enzyme. These observations suggest the involvement of negative homotropic interactions in phenylalanine ammonia-lyase. 12. The enzyme could not be desensitized by treatment with HgCl2, p-chloromercuribenzoic acid or by repeated freezing and thawing.


Folic acid therapy and cardiovascular disease in ESRD or advanced chronic kidney disease: a meta-analysis.
The efficacy of folic acid therapy to lower homocysteine (Hcy) levels in an effort to reduce cardiovascular disease (CVD) risk in patients with ESRD or advanced chronic kidney disease (ACKD; creatinine clearance, <30 ml/min) remains inconclusive. We conducted a meta-analysis of relevant randomized trials to further examine this issue. This meta-analysis included 3886 patients with ESRD/ACKD from seven qualified randomized trials using folic acid therapy and with CVD reported as one of the end points. When pooling the seven trials, folic acid therapy reduced the risk of CVD by 15% (RR, 0.85; 95% CI, 0.76 to 0.96; P = 0.009). A greater beneficial effect was observed among those trials with a treatment duration >24 months (RR, 0.84; 95% CI, 0.72 to 0.98; P = 0.02), a decrease in Hcy level >20% (RR, 0.83; 95% CI, 0.73 to 0.95; P = 0.007), and no or partial folic acid fortification (RR, 0.80; 95% CI, 0.65 to 0.99; P = 0.04). The beneficial effect also was seen when Hcy levels decreased >20%, even in the presence of folic acid fortification (RR, 0.85; 95% CI, 0.73 to 0.99; P = 0.04). In the corresponding comparison groups, the estimated RRs were attenuated and insignificant. Folic acid therapy can reduce CVD risk in patients with ESRD/ACKD by 15%. A greater beneficial effect was observed among those trials with no or partial folic acid fortification or a decrease in Hcy level >20% regardless of folic acid fortification.


Interaction of meso-tetrakis (4-N-methylpyridyl) porphyrin in its free base and as a Zn(II) derivative with large unilamellar phospholipid vesicles.
Our aim was to investigate the interaction of the cationic meso-tetrakis (4-N-methylpyridyl) porphyrin, a photosensitizer used for photodynamic therapy, in its free base form (TMPyP) and complexed with Zn(II) (ZnTMPyP), with large unilamellar vesicles (LUVs), as a model for the gram-negative bacterial cell wall. Mixtures of the zwitterionic 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and anionic 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (POPG) phospholipids, at different molar percentages, were used as LUVs. A significant increase of porphyrin affinity at higher POPG molar concentrations was observed from the binding constant values, K b, estimated by optical absorption and steady-state fluorescence. Besides, as demonstrated by time-resolved fluorescence, this affinity increase is also followed by a higher fraction of vesicle-bound porphyrin in the LUVs. Moreover, based on the K b values, we have observed a higher affinity of the ZnTMPyP to the POPG containing LUVs as compared to the TMPyP. Steady-state fluorescence quenching and zeta potential studies revealed that both porphyrins are possibly located at the LUVs Stern layer region. Therefore, the electrostatic attraction between the positively charged porphyrin peripheral groups and the negatively charged outer surface of the LUVs plays an important role in porphyrin association and localization. Our results have improved the understanding of the successful application of cationic porphyrins on the photo-inactivation of gram-negative bacteria. Since a higher accumulation of the ZnTMPyP in the bacterial cell wall would be expected, this porphyrin could be a more efficient therapeutic drug for this treatment.


Prediction of nighttime driving visibility from laboratory data.
The application of laboratory threshold visibility data to the subject of driving visibility with heat absorbing glass has been reviewed in an attempt to resolve excessive differences between calculated predictions and road test observations. New calculations are described that yield predicted losses of visibility distance due to the use of heat absorbing glass rather than regular glass in automobile windshields. The predicted losses agree satisfactorily with the observed losses for road tests, which average proximately 3%. The new calculations have made use of a revised visual exposure interval of ? sec corresponding with five visual fixational pauses per second and a new simulation model that assumes that the target-to-background contrast increases with reduced headlamp-to-target distance.


Research resource: Gene profiling of G protein-coupled receptors in the arcuate nucleus of the female.
The hypothalamic arcuate nucleus controls many critical homeostatic functions including energy homeostasis, reproduction, and motivated behavior. Although G protein-coupled receptors (GPCRs) are involved in the regulation of these functions, relatively few of the GPCRs have been identified specifically within the arcuate nucleus. Here, using TaqMan low-density arrays we quantified the mRNA expression of nonolfactory GPCRs in mouse arcuate nucleus. An unprecedented number of GPCRs (total of 292) were found to be expressed, of which 183 were known and 109 were orphan GPCRs. The known GPCR genes expressed were classified into several functional clusters including hormone/neurotransmitter, growth factor, angiogenesis and vasoactivity, inflammation and immune system, and lipid messenger receptors. The plethora of orphan genes expressed in the arcuate nucleus were classified into 5 structure-related classes including class A (rhodopsin-like), class B (adhesion), class C (other GPCRs), nonsignaling 7-transmembrane chemokine-binding proteins, and other 7-transmembrane proteins. Therefore, for the first time, we provide a quantitative estimate of the numerous GPCRs expressed in the hypothalamic arcuate nucleus. Finally, as proof of principle, we documented the expression and function of one of these receptor genes, the glucagon-like peptide 1 receptor (Glp1r), which was highly expressed in the arcuate nucleus. Single-cell RT-PCR revealed that Glp1r mRNA was localized in proopiomelanocortin neurons, and using whole-cell recording we found that the glucagon-like peptide 1-selective agonist exendin-4 robustly excited proopiomelanocortin neurons. Thus, the quantitative GPCR data emphasize the complexity of the hypothalamic arcuate nucleus and furthermore provide a valuable resource for future neuroendocrine/endocrine-related experiments.


Fundus autofluorescence in age-related macular disease imaged with a laser scanning ophthalmoscope.
To image and quantify the spatial distribution of fundus autofluorescence in normal subjects, to determine its age dependence, and to document the deviation from normal in patients with age-related macular disease. Using a confocal laser scanning ophthalmoscope (cLSO), the intensity and spatial distribution of fundus autofluorescence was studied in 33 normal subjects, 97 eyes with drusen only, and 111 eyes with visual loss caused by age-related macular disease. Fundus autofluorescence intensity in normal subjects was highest at the posterior pole and dipped at the fovea. Autofluorescence increased with age at the posterior pole. Fundus in eyes with age-related maculopathy showed localized high autofluorescence that did not correspond with drusen. Linear pigmentation at the level of the retinal pigment epithelium (RPE), whether detached or flat, fluoresced brightly, whereas plaques of melanin did not. Areas of low and high levels of autofluorescence were seen in lesions containing choroidal new vessels. In areas of geographic atrophy, autofluorescence was low. The spatial distribution of background fundus autofluorescence and the correlation of autofluorescence with age in normal subjects imply that autofluorescence is derived from lipofuscin at the level of the RPE. Focal accumulation of autofluorescent material occurs at the level of the RPE in patients with drusen, but the drusen do not show marked increases in autofluorescence. It is likely that melanolipofuscin accounts for the high levels of autofluorescence, corresponding to linear pigmentation at the level of the RPE. Low-intensity autofluorescence occurs in the presence of retinal photoreceptor loss, and variable levels over disciform lesions probably relate to variations in metabolic activity of the RPE.


Room temperature hydrophosphination using a simple iron salen pre-catalyst.
Phosphines are fundamentally important to the fine chemicals, pharmaceutical and agrochemical industries. Reported is the first example of alkene hydrophosphination using a designed iron pre-catalyst which yields the anti-Markovnikov products in high yield at room temperature. The phosphine products are excellent pro-ligands for Fe-catalyzed Negishi cross-coupling.


Bacillus rigiliprofundi sp. nov., an endospore-forming, Mn-oxidizing, moderately halophilic bacterium isolated from deep subseafloor basaltic crust.
A facultatively anaerobic bacterium, designated strain 1MBB1T, was isolated from basaltic breccia collected from 341 m below the seafloor by seafloor drilling of Rigil Guyot during Integrated Ocean Drilling Program Expedition 330. The cells were straight rods, 0.5 μm wide and 1-3 μm long, that occurred singly and in chains. Strain 1MBB1T stained Gram-positive. Catalase and oxidase were produced. The isolate grew optimally at 30 °C and pH 7.5, and could grow with up to 12 % (w/v) NaCl. The DNA G+C content was 40.5 mol%. The major cellular fatty acids were C16:1ω11c (26.5 %), anteiso-C15:0 (19.5 %), C16:0 (18.7 %) and iso-C15:0 (10.4 %), and the cell-wall diamino acid was meso-diaminopimelic acid. Endospores of strain 1MBB1T oxidized Mn(II) to Mn(IV), and siderophore production by vegetative cells was positive. Phylogenetic analysis of the 16S rRNA gene indicated that strain 1MBB1T was a member of the family Bacillaceae, with Bacillus foraminis CV53T and Bacillus novalis LMG 21837T being the closest phylogenetic neighbours (96.5 and 96.2 % similarity, respectively). This is the first novel species described from deep subseafloor basaltic crust. On the basis of our polyphasic analysis, we conclude that strain 1MBB1T represents a novel species of the genus Bacillus, for which we propose the name Bacillus rigiliprofundi sp. nov. The type strain is 1MBB1T ( = NCMA B78T = LMG 28275T).


Chronic Meningitis Investigated via Metagenomic Next-Generation Sequencing.
Identifying infectious causes of subacute or chronic meningitis can be challenging. Enhanced, unbiased diagnostic approaches are needed. To present a case series of patients with diagnostically challenging subacute or chronic meningitis using metagenomic next-generation sequencing (mNGS) of cerebrospinal fluid (CSF) supported by a statistical framework generated from mNGS of control samples from the environment and from patients who were noninfectious. In this case series, mNGS data obtained from the CSF of 94 patients with noninfectious neuroinflammatory disorders and from 24 water and reagent control samples were used to develop and implement a weighted scoring metric based on z scores at the species and genus levels for both nucleotide and protein alignments to prioritize and rank the mNGS results. Total RNA was extracted for mNGS from the CSF of 7 participants with subacute or chronic meningitis who were recruited between September 2013 and March 2017 as part of a multicenter study of mNGS pathogen discovery among patients with suspected neuroinflammatory conditions. The neurologic infections identified by mNGS in these 7 participants represented a diverse array of pathogens. The patients were referred from the University of California, San Francisco Medical Center (n = 2), Zuckerberg San Francisco General Hospital and Trauma Center (n = 2), Cleveland Clinic (n = 1), University of Washington (n = 1), and Kaiser Permanente (n = 1). A weighted z score was used to filter out environmental contaminants and facilitate efficient data triage and analysis. Pathogens identified by mNGS and the ability of a statistical model to prioritize, rank, and simplify mNGS results. The 7 participants ranged in age from 10 to 55 years, and 3 (43%) were female. A parasitic worm (Taenia solium, in 2 participants), a virus (HIV-1), and 4 fungi (Cryptococcus neoformans, Aspergillus oryzae, Histoplasma capsulatum, and Candida dubliniensis) were identified among the 7 participants by using mNGS. Evaluating mNGS data with a weighted z score-based scoring algorithm reduced the reported microbial taxa by a mean of 87% (range, 41%-99%) when taxa with a combined score of 0 or less were removed, effectively separating bona fide pathogen sequences from spurious environmental sequences so that, in each case, the causative pathogen was found within the top 2 scoring microbes identified using the algorithm. Diverse microbial pathogens were identified by mNGS in the CSF of patients with diagnostically challenging subacute or chronic meningitis, including a case of subarachnoid neurocysticercosis that defied diagnosis for 1 year, the first reported case of CNS vasculitis caused by Aspergillus oryzae, and the fourth reported case of C dubliniensis meningitis. Prioritizing metagenomic data with a scoring algorithm greatly clarified data interpretation and highlighted the problem of attributing biological significance to organisms present in control samples used for metagenomic sequencing studies.


Ocular injury reduction from ocular protection use in current combat operations.
The mechanisms of modern warfare unfortunately have lead to many ocular injuries. This study attempts to quantify the effect of military combat eye protection on the incidence of ocular injury among US service members engaged in current combat operations. We retrospectively gathered data from the Joint Theater Trauma Registry from patients entering level III hospital facilities in Operations Iraqi and Enduring Freedom from March 2003 to September 2006. The presence of ocular injury was the primary outcome measure; whereas severity of injury and the effect of eye protection education on compliance with wearing this protection were secondary outcome measurements. Ocular protection status at the time of injury was documented in 3,276 casualties during the collection period. Of these, 451 of 2,671 (17%) casualties who reported wearing ocular protection sustained an ocular injury and 155 of 605 (26%) casualties who reported not wearing eye protection suffered an eye injury (p < 0.01). After an intense ocular protection education program, there was a 16% increase in compliance of eye protection use in combat. Finally, when the severity of eye injuries are compared using an abbreviated injury scale, casualties with eye protection had decreased injury severity in addition to incidence (p < 005). This study suggests that the military combat eye protection used by military personnel during current combat operations has resulted in significantly fewer and less severe ocular injuries. Further, results from this study may also suggest that Department of Defense educational programs may have been successful in increasing eye protection compliance.